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What States Have Motorcycle Helmet Laws
Ramirez had seen the four-door sedan that summer morning as it swerved into oncoming traffic and struck the motorcycle.
Today, only three states have no laws relating to helmet use. Iowa, Illinois and New Hampshire do not require motorcyclists to wear helmets. Another 28 states have partial helmet laws, typically requiring riders under a certain age to wear a helmet. Georgia is one of 19 states that makes it clear riders should always wear a helmet.
Motorcycle Gas Tank Repair Near Me Costs of a mechanic or purchasing a new motorcycle fuel tank can run to hundreds of dollars. You can repair them yourself, however, and this article will show you how. Step 1 – Drain the Fuel Tank. In order to repair a leaky motorcycle fuel tank it first has to be drained. Make sure you
Nineteen states currently have a universal helmet law. This means that all motorcycle drivers on the road must wear a helmet regardless of their age. Arguments Against Motorcycle Helmet Laws in Illinois. The states that do not currently have motorcycle helmet laws include Illinois, Iowa, and New Hampshire.
States have been gradually repealing or weakening mandatory helmet laws for nearly two decades. In 1967, to increase motorcycle helmet use, the federal government required that states enact helmet.
Armed with lemons, roughly 500 motorcyclists rode to the Pennsylvania Capitol on Monday, where they rallied in the Capitol Rotunda to advocate for legislation that would protect motorcycle. we got.
Jun 1, 2012. In several states, the issue of legislating motorcycle helmet use has become controversial, with. Illinois currently does not have a helmet law.
Missouri law says anyone riding a motorcycle on Missouri roads has to wear a helmet. More than 15 other states have similar, universal helmet laws. In Kansas adults have the choice whether to wear a.
organized in groups, successfully lobbied to have these state laws repealed.3 Currently only. twenty states have universal motorcycle helmet laws.20.
Despite these facts, many states have repealed motorcycle helmet legislation. But what is the real dispute? Those who oppose helmet laws will undoubtedly refer to an individual's right to choose.
Jul 18, 2014. Iowa is off to its deadliest start to the year for motorcyclists since 2012, In all, 19 states have universal helmet laws. and 28 have age-based.
Wearing a bicycle helmet is always a good idea no matter what the law has to say, but bicycle helmet laws might also have an impact on an injury claim after a bicycle-car accident. No state currently requires helmets for adult bicyclists, but just under half of U.S. states require the use of helmets by riders under a certain age.
Mar 29, 2017. CHARLESTON, W.Va — The state Senate Wednesday defeated a bill that would have eliminated the state's motorcycle helmet law. SB 549.
During the pilot period, only riders under the age of 21 would have to wear wear one. A motorcyclist wears a patch on his riding vest declaring his opinion of Washington state's current mandatory.
U.S. and Canadian Motorcycle Helmet Use Laws. In some states, there are specific requirements which can change the application of the law. In Texas, for instance, riders 20 years-old and younger must wear a helmet, but those older must also wear one if they have not completed a motorcycle riding safety course or do not have health insurance.
"We have got to get over being a nanny state where we dictate every aspect of everybody's lives." Burlison and the bill's supporters contended that repealing the helmet law could also be good for the.
For more than 30 years, Umbenhauer has shook hands, attended fundraisers and talked to everyone who will listen about motorcycles — first as a state. the law change in 2003 to make helmets optional.
What US states do not have a motorcycle helmet law? Click on the link to your right for the answer. According to the American Motorcycle Association, a majority of states in the United States.
Jun 6, 2016. Live free and die: Michigan's motorcycle helmet law four years later. who meet certain requirements have had the freedom to ride without a helmet. According to Michigan State Police data, 138 persons were killed in 2015.
Understanding motorcycle helmet laws in Georgia On behalf of Cromartie Law posted in Motorcycle Accidents on Wednesday, February 15, 2017. In Georgia, the motorcycle helmet law has not changed since about 1969, but that is not true in all states.
Only 19 states and the District of Columbia require all motorcyclists. TELL US: Illinois currently doesn't have any motorcycle helmet laws on the books. Should that change, or should cyclists be.
Myth—Age-specific motorcycle helmet laws are effective Fact—Statistics tell us that the helmet use rate in states with age-specific helmet laws is usually the same as having no law at all. Currently 23 states have a law requiring helmet use for a specific portion.
A New York man died Sunday while participating in a ride with 550 other motorcyclists to protest the state's mandatory helmet law. Police said Philip A. Contos, 55, hit his brakes and his motorcycle.
Twenty-eight states have motorcycle helmet laws covering only some riders, usually for people younger than 18. Illinois, Iowa and New Hampshire don't require helmets for anyone riding motorcycles.
Aug 10, 2011 · Motorcycle helmet laws save lives: Death rates from head injuries, such as traumatic brain injury, are twice as high among motorcyclists in states without all-rider helmet laws. And motorcycle helmets are 37 percent effective in preventing motorcyclist deaths and 67 percent effective in preventing brain injuries.
Other states require head protection only for minors or. even when individual choices have collective costs. Motorcycle helmet laws are an unwarranted exception to our normal, sound approach, which.
In some countries, most notably the USA and India, there is some opposition to compulsory helmet use (see Helmet Law Defense League); not all USA states have a compulsory helmet law. Snell-and-DOT decal on an HJC motorcycle helmet in the U.S.
While the rider is expected to make a full recovery, police reported Tuesday, others have not been so lucky. It's been a particularly deadly summer for motorcycle riders. Safety Act required states.
And in every community in the United States. that motorcycle riders would have the common sense to wear helmets. Of course, we would also like to believe that all drivers would wear seat belts and.
Motorcycle operators who have had an operator's license for less than one year. FACT: In states without a universal helmet law, 65 percent of motorcyclists.
Jul 8, 2015. As you might have expected, the history of the United States' universal helmet laws, which is a more appetizing way of referring to mandatory.
May 15, 2019 · State Helmet Laws Based on Age and Additional Requirements. Twenty-eight states have age-based helmet laws, and 17 and younger is the most common age range for mandatory helmet use. Some states further refine their motorcycle helmet laws with safety training and/or medical insurance requirements.
Motorcyclists in our country usually take road safety laws for. by the Helmet Manufacturers Association to meet the demand of all motorcycle users in the country. India Union Budget 2019: What does.
Apr 24, 2017. Boone said he often plans trips to states that have looser helmet laws, and he thinks the bill would lead to increased tourism from bikers.
Michigan Motorcycle Accident Lawyers Create Helmet Law Graphic. The Helmet Laws infographic also displays which states have the highest fines and.
Today, only 19 states and the District of Columbia require all motorcycle operators to wear a helmet, the NCSL blog says. Other jurisdictions require only some, such as younger riders, to wear helmets. Three states – Illinois, Iowa and New Hampshire – have no helmet requirement at all, the NCSL says.
"Other states have shown reduction in car-motorcycle accidents that. According to the National Conference of State Legislatures, when Florida repealed its motorcycle helmet law for adults in 2000,
Dec 22, 2015. However, Ohio's motorcycle helmet laws require that all riders under the age of. Some states have different rules for low-power cycles (bikes,
all low-power cycles with an engine displacement greater than 50cc or can attain speeds greater than 30 mph are covered by the motorcycle helmet law: New York: all riders: all low-power cycles designed to travel at speeds of 20 mph or greater are covered by the motorcycle helmet law: North Carolina: all riders: yes: North Dakota: 17 and younger Footnote 8: yes: Ohio
The same weekend in New Hampshire, seven members of a motorcycle club – the Marine. New Hampshire doesn't have a helmet.
Jul 29, 2017. Motorcycle advocates have been effective in making this an issue. And while helmet laws were widespread in the 1970s, today just 19 states.
Jul 10, 2018 · Illinois, Iowa and New Hampshire are the only few without a helmet law set in place. Missouri helmet law. Motorcycle helmet laws vary state to state. There are currently three ways to identify states that do or do not participate, which include: Universal law: A law that requires all motorcyclists wear a helmet. Partial law: A law that requires only young riders (usually 17 and younger) and some.
Fact—The highest courts in more than 25 states have held motorcycle helmet laws to be constitutional. The Massachusetts motorcycle helmet law was affirmed by the U.S. Supreme Court. The Massachusetts motorcycle helmet law was affirmed by the U.S. Supreme Court.
Jan 11, 2018. Effort to repeal Nebraska's motorcycle helmet law fails in Legislature. "I'm sad for the motorcycle riders across the state," Lowe said. with a helmet, they ride with more safety than when they do not have that helmet in place.
Only a minority of states require helmet use by law. A mere nineteen-out-of-fifty mandate helmets for all riders, with two, Illinois and Iowa, holding tight to their free-choice helmet option. However, the remaining twenty-nine states have designed their motorcycle helmet laws, so be sure to look up the specific details that apply to where you.
per 10000 registered motorcycles and states without helmet laws experienced a median death rate of. medical literature have uniformly recommended helmet.
Hog Bikers Jan 26, 2019 · HOG: Harley Owners Group; Hack: A sidecar. Also a "side hack." Hamsters: A high-profile group of custom motorcycle enthusiasts, easily recognized at events by their distinctive uniform: A yellow t-shirt with the Hamsters' logo. Membership includes: Arlen Ness, Dave Perowitz, and Donnie Smith. BILLERICA — Engines roaring, members of a local motorcycle
May 18, 2017. Describing the Massachusetts helmet law as a "nanny state" approach, Gleason said motorcyclists have a voice at the State House and he.
Currently, 19 states and the District of Columbia have laws requiring all motorcyclists to wear a helmet which is known as universal helmet laws. Twenty-eight other states require only some motorcyclists to wear helmets. Three states, Illinois, Iowa and New Hampshire do not currently require motorcycle helmets while riding a motorcycle.
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Apr 11, 2018 · Helmet Laws in the United States. In three states namely New Hampshire, Illinois and Iowa, no motorcycle riders helmet law is in use. The Laws that require some bicycle riders to wear a helmet in twenty one states and the District of Columbia In twenty-nine states, no law was imposed for bicycle riders to wear helmet.
Currently, only 19 states and the District of Columbia have universal helmet laws; 28 states have laws requiring some bikers to use helmets; and three states have no helmet laws. Currently, Kansas and Missouri have different helmet laws, but that may change.
In 2012, the Michigan legislature repealed the state's motorcycle helmet law, a law that had been on the books for 35 years. In its place, the new law allowed riders 21 years old and older who have.
Nov 6, 2017. In 1975, nearly every state required all motorcyclists to wear helmets because of a federal law that tied highway construction funding to helmet.
The rider must have completed a motorcycle safety course. why don't they just mandate helmets for all riders like several other states already do? The law would be cut and dry, and police would.
states had universal helmet use laws in effect by the end of 1967 and 14 more states. Motorcycle helmet use laws have been one of the most contentious.
Finally, helmet law proponents argue that aside from the United States, most developed countries have mandatory motorcycle helmet laws in place. This supports the argument that if most other countries deem it necessary to have helmet laws to protect their citizens, the United States should follow suit.
Mar 31, 2014. More states are being pressured to repeal their universal motorcycle helmet laws. But when those laws are changed, the number of fatalities.
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arguing that "incredible strides" have been made to improve motorcycle safety and reduce deaths and injuries in Connecticut since the state's helmet law was repealed in 1976. And he said he has worked.
Abstract. The authors examine state motorcycle helmet laws in the United States, demonstrate their effectiveness, and tell how opponents have challenged their.


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Canine Albumin Polymorphisms and Their Impact on Drug Plasma Protein Binding
Ana P. Costa, Michael H. Court, Neal S. Burke, Zhaohui Zhu, Katrina L. Mealey and Nicolas F. Villarino
Drug Metabolism and Disposition October 2019, 47 (10) 1024-1031; DOI: https://doi.org/10.1124/dmd.119.087304
Ana P. Costa
Program in Individualized Medicine, Department of Veterinary Clinical Sciences, Washington State University College of Veterinary Medicine, Pullman, Washington
ORCID record for Ana P. Costa
Michael H. Court
Neal S. Burke
Zhaohui Zhu
Katrina L. Mealey
ORCID record for Katrina L. Mealey
Nicolas F. Villarino
PDF + SI
Drug binding to plasma proteins is routinely determined during drug development. Albumin polymorphisms c.1075G>T (p.Ala359Ser) and c.1422A>T (p.Glu474Asp) were previously shown to alter plasma protein binding of a drug candidate (D01-4582, 4-[1-[3-chloro-4-[N′-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidine-2-yl]methoxybenzoic acid) in a colony of Beagles. Our study investigated the hypothesis that drug-protein binding in plasma from dogs with the albumin H1 (reference) allele would be greater than in plasma from dogs with the albumin H2 allele (c.1075G>T and c.1422A>T) (n = 6 per group). The plasma protein binding extent of four drugs (D01-4582, celecoxib, mycophenolic acid, and meloxicam) was evaluated using ultracentrifugation or equilibrium dialysis. Free and total drug concentrations were analyzed by liquid chromatography–mass spectrometry. The albumin gene coding region was sequenced in 100 dogs to detect novel gene variants, and H1/H2 allele frequency was determined in a large and varied population (n = 1446 from 61 breeds and mixed-breed dogs). For meloxicam, H1 allele plasma had statistically significant higher free drug fractions (P = 0.041) than H2 allele plasma. No significant difference was identified for plasma protein binding of D01-4582, celecoxib, or mycophenolic acid. c.1075G>T and c.1422A>T were the most common single nucleotide polymorphisms in canine albumin, present concurrently in most study dogs and occasionally identified independently. Our findings suggest a potential influence of c.1075G>T and c.1422A>T on plasma protein binding. This influence should be confirmed in vivo and for additional drugs. Based on our results, albumin genotyping should be considered for canine research subjects to improve interpretation of pharmacokinetic data generated during the drug development process for humans and dogs.
For some drugs, the risk of adverse effects (toxicity or lack of efficacy) depends on the concentration of free drug achieved in plasma. In some cases, free drug concentrations and the extent of drug binding to plasma proteins can be affected by differences in the albumin genotype. Canine albumin polymorphisms c.1075G>T (p.Ala359Ser; rs852211303) and c.1422A>T (p.Glu474Asp; rs851238996) have been identified as a cause of pharmacokinetic variability of a drug candidate (D01-4582, 4-[1-[3-chloro-4-[N′-(2-methylphenyl) ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidine-2-yl]methoxybenzoic acid) in Beagle dogs (Ito et al., 2009). [To note, the description of protein sequence variants used by the authors is based on recommendations by the Human Genome Variation Society, with the position of the amino acid substituted calculated based on the precursor peptide (proalbumin) and not the mature peptide as in the article by Ito et al. (2009).] Specifically, dogs homozygous for both single nucleotide polymorphisms (SNPs) had an up to 6-fold greater unbound drug fraction compared with wild-type dogs, resulting in significantly greater drug clearance. Although the report by Ito et al. (2009) identified an association between canine albumin genotypes and high interindividual variability in drug disposition, the study was limited to only one experimental drug in a colony of research Beagles. The impact of canine albumin polymorphisms on plasma protein binding of other drugs and in a more heterogeneous dog population (outside of research colonies) remains to be investigated.
The primary objective of our study was to investigate the hypothesis that plasma samples collected from dogs with the albumin H1 allele (reference) have a higher extent of drug-protein binding than that of plasma collected from dogs with the albumin H2 allele (c.1075G>T and c.1422A>T). To test our hypothesis, we compared the extent of plasma protein binding of four highly albumin-bound drugs (D01-4582, meloxicam, celecoxib, and mycophenolic acid) in plasma obtained from dogs with H1 and H2 alleles. Secondary objectives of our study were to screen for other nonsynonymous variants in the coding region of the canine albumin gene that could influence protein tertiary structure and to describe c.1075G>T and c.1422A>T genotype frequencies among a large number of different dog breeds.
Reagents and Chemicals
Mycophenolic acid, meloxicam, and celecoxib, as well as their deuterated forms (mycophenolic acid-d3, meloxicam-d3, and celecoxib-d4), were purchased from Toronto Research Chemicals (Toronto, ON, Canada). D01-4582 was synthesized and generously provided by the Daiichi Sankyo Medicinal Chemistry Research Laboratory (Tokyo, Japan). Chemical structures of each analyte and its deuterated form are included in Fig. 1. High-performance liquid chromatography (HPLC)–grade methanol and acetonitrile (ACN) were from Fisher Scientific (Pittsburgh, PA). Milli‐Q water of at least 18 MΩ resistance was used to prepare the mobile phase. Potassium phosphate monobasic (99%) was from Fisher Chemical (Fair Lawn, NJ). Sodium chloride (99%) and potassium chloride (99%) were from Fisher Bioreagents (Fair Lawn, NJ). Sodium phosphate dibasic and formic acid (88%) were from J.T. Baker (Phillipsburg, PA).
Chemical structure for D01-4582, meloxicam, celecoxib, and mycophenolic acid.
Canine Plasma Samples
Blood samples were collected using procedures approved by the Washington State University (WSU) Institutional Animal Care and Use Committee (ASAF numbers 04662-004 and 04797-001). Owners provided written consent for their dogs' participation in the study. Pooled canine plasma collected into sodium citrate tubes from clinically healthy dogs (n = 6) was used for initial analytical assay validation (methods described below). The sample size requirement to evaluate the effect of albumin genotype (H1/H1 vs. H2/H2) on drug binding in plasma was determined based on mean ± S.D. results from a previous report (Ito et al., 2009). This analysis indicated that a minimum of five dogs per group was needed to achieve an α value of 0.05 with a power value of 0.80 for a 6-fold difference in mean free drug fraction. Therefore, we proposed to study six dogs per genotype group.
Privately owned dogs were recruited from the WSU student, faculty, and employee population and genotyped for the c.1075G>T and c.1422A>T variants (method described below). After initial screening, six dogs with the H1/H1 genotype (c.1075G and c.1422A) and six dogs with the H2/H2 genotype (c.1075T and c.1422T) were selected for our study. Breeds included in the H1/H1 genotype group were as follows: Labrador Retriever (n = 3), Afghan Hound (n = 1), English Pointer (n = 1), and mixed breed (n = 1). Breeds included in the H2/H2 genotype group were as follows: Golden Retriever (n = 2), Borzoi (n = 1), Pitbull (n = 1), German Shorthaired Pointer (n = 1), and mixed breed (n = 1). All study dogs (five castrated males and seven spayed females) were judged to be clinically healthy based on history and physical examination, with no medications received for at least 14 days before blood collection. Owners were asked to fast the dogs on the morning of blood sample collection. Blood samples were collected by venipuncture into tubes with sodium citrate as an anticoagulant. Plasma was obtained by centrifugation (1800g for 8 minutes) of blood samples, transferred to cryotubes, and stored at −80°C until analysis.
Determination of the Extent of Plasma Protein Binding of D01-4582, Celecoxib, Mycophenolic Acid, and Meloxicam
D01-4582 was included in our study because of its reported binding differences associated with canine albumin genotype. The remaining compounds were selected based on their reported high plasma protein binding (>95%), site-specific binding to human albumin (Er et al., 2013), and previous clinical use in dogs (Table 1). The method (ultracentrifugation or equilibrium dialysis) used to determine the extent of each drug plasma binding was primarily selected based on methods previously reported for that drug Jolliet et al., 1997; Lapicque et al., 2000; Ito et al., 2009; Cooper et al., 2014; Morassi et al., 2018).
Relevance of the drugs selected for our protein binding experiments in dogs
To control the potential effect of pH differences on drug-protein binding, individual plasma samples were adjusted with 1 N HCl to achieve a mean pH of 7.34 ± 0.04 (Mettler Toledo, Columbus, OH). The volume of acid used in these adjustments was <0.5% of the total mixture. The plasma albumin concentration for each genotyped plasma sample was determined using a standard bromocresol green method, and absorbance was monitored at 630 nm (SEKURE chemistry albumin assay; Sekisui Diagnostics, Lexington, MA).
Ultracentrifugation (for D01-4582).
Initially, 10 µl D01-4582 standard solution (2500 μg/ml) was added to 490 µl plasma for a final concentration of 50 ppm (92.6 µM). Samples were then incubated for 30 minutes at 37°C and 210 µl of each plasma sample was centrifuged in a 0.5-ml thick-wall polycarbonate tube on a fixed-angle rotor Thermo Sorvall MX-150 Micro-Ultracentrifuge (Sorvall Instruments, Newton, CT) at 200,000g for 16 hours at 4°C. The ultracentrifugation resulted in visually distinct layers: superficial (lipid), middle (clear), and deep (protein pellet). To minimize disruption of the lipid superficial layer, the supernatant sample was collected from the middle layer immediately after centrifugation, as recommended (Dr. Takashi Ito, personal communication). In addition, the rotor was kept in ice during collection to maintain sample temperature. A 50-μl aliquot of the supernatant was collected into a 0.6-ml Eppendorf tube and prepared for HPLC–tandem mass spectrometry (MS/MS) analysis, as described below. The unbound fraction of D01-4582 was estimated by dividing the unbound concentration by the nominal incubation concentration. Plasma from a nongenotyped dog was used to confirm acceptable inter- and intrarun precision [coefficient of variation (CV) < 15%]. The experiments for each dog were conducted in duplicate and repeated on 3 separate days.
High-Throughput Equilibrium Dialysis (for Celecoxib, Mycophenolic Acid, and Meloxicam).
A high-throughput equilibrium dialysis device (96-well, 75-μl half-cell capacity) and cellulose membranes (12–14 kDa molecular mass cutoff) (HT-Dialysis, Gales Ferry, CT) were used to determine the extent of protein binding for celecoxib, mycophenolic acid, and meloxicam. The dialysis membranes and device were conditioned and assembled as recommended by the manufacturer. Plasma samples were spiked with celecoxib (4 μM), mycophenolic acid (12.5 μM), and meloxicam (1.6 μM).
Compound spiked plasma samples were transferred to the plasma (donor) chambers of the dialysis device (75 μl per half-well), and an equal volume of phosphate buffer (15 mM, pH 7.3 ± 0.1) was placed in the buffer (receiver) chambers. The dialysis device was sealed using the kit adhesive (HT-Dialysis), and dialysis was conducted in an orbital shaker (100 rpm) maintained at 37°C. Complete equilibrium was achieved in the HT-Dialysis apparatus within 3.5 hours (data not shown) and samples were collected at the end of 4 hours. Samples were then processed for quantification by HPLC-MS/MS analysis as described below.
The extent of plasma protein binding of each analyte in all of the plasma samples was estimated using the following mathematical expressions:
where Cb and Cp represent the measured concentration of the compound from the buffer and plasma chambers of the dialysis device, respectively.
Quality controls (QCs) using mycophenolic acid (12.5 μM) (previously validated binding evaluation by equilibrium dialysis in canine plasma) were included in each row of the dialysis apparatus to monitor dialysis membrane integrity and assay performance. Experiments for each dog and compound were conducted in triplicate and repeated on 3 separate days.
Determination of Drug Concentrations
Chromatographic Conditions.
The concentration of each analyte was determined by liquid chromatography with mass spectrometry detection using an HPLC apparatus (Agilent 1100; Agilent Technologies, Santa Clara, CA) connected to a triple quadrupole mass spectrometry detector (AB Sciex API4000; Applied Biosystems/Life Technologies, Framingham, MA) operated in positive ion mode. The mobile phase consisted of 95% (v/v) water [containing 0.1% (v/v) formic acid] and 5% (v/v) ACN at 1 ml per minute through a Zorbax Eclipse XDB-C18 column (2.1 mm × 50 mm, 5 μm; Phenomenex, Torrance, CA). Mass transitions monitored included m/z 540.3→402.2 (D01-4582), m/z 382.2→362 (celecoxib), m/z 386.2→366 (celecoxib-d4), m/z 321.2→207.1 (mycophenolic acid), m/z 324.2→210.1 (mycophenolic acid-d3), m/z 352.1→115.1 (meloxicam), and m/z 355.1→115.1 (meloxicam-d3). Retention times for D01-4582, celecoxib, mycophenolic acid, and meloxicam were 2.20, 2.41, 2.11, and 2.14 minutes, respectively.
Calibration Standards.
Calibration standards and QC samples (n = 3 for each concentration) were prepared in canine plasma before protein precipitation (for celecoxib, mycophenolic acid, and meloxicam) or in the supernatant (D01-4582). The concentration ranges used for calibration curves and QCs for each analyte are listed in Table 2.
Concentrations used to validate the methods in the quantification of D01-4582, meloxicam, mycophenolic acid, and celecoxib in canine plasma
Sample Preparation for HPLC-MS/MS Analysis
For D01-4582 samples, methanol (50 µl) containing the internal standard (meloxicam-d3) was added to the supernatant sample (50 µl). The mixture was vortexed and centrifuged at 10,000g for 15 minutes at 4°C, with the supernatant (20 µl) introduced into the HPLC-MS/MS.
For the celecoxib, mycophenolic acid, and meloxicam samples, each 50-μl aliquot of plasma (calibration standard or QC samples) was transferred into a 2-ml Eppendorf tube with 50 μl phosphate buffer, 1000 μl precipitating solution [ACN for celecoxib and meloxicam; methanol/ACN 1:1 (v/v) for mycophenolic acid], and the respective internal standard (4 µl for celecoxib-d4, 3 µl for mycophenolic acid-d3, and 5 µl for meloxicam-d3). For dialysis samples, the matrix was equilibrated before protein precipitation by adding 50 μl plasma or phosphate buffer to the corresponding sample (e.g., plasma to dialysis buffer samples, and buffer to dialysis plasma samples) before protein precipitation and the addition of internal standard. Each extract was then vortexed for 30 seconds and centrifuged at 17,000g for 10 minutes. A 1050-μl aliquot of supernatant was evaporated to dryness using a Speed-Vac concentrator (Savant Instruments Inc., Farmingdale, NY). The residue was resuspended in 100 μl 95% (v/v) water [containing 0.1% (v/v) formic acid] and 5% (v/v) ACN, mixed, and centrifuged at 17,000g for 10 minutes for further sample cleanup. The second supernatant was injected into the chromatographic system.
Methods used for determination of D01-4582, celecoxib, mycophenolic acid, and meloxicam were validated according to the Guidelines for Bioanalytical Method Validation published by the U.S. Food and Drug Administration in May 2001. Quantification of all analytes was performed using internal standard calibration. All QC samples and calibration standards were prepared in triplicate in canine plasma (for celecoxib, mycophenolic acid, and meloxicam) or in the supernatant (for D01-4582). The lower limit of quantitation, the limit of detection, and the concentration ranges used for calibration curves and QCs for each analyte are listed in Table 2. Calibration curves were obtained with weighting of 1/x.
For all four analytes evaluated, the procedure produced a linear curve (r2 > 0.998) over the concentration ranges evaluated with a high degree of repeatability. Both intra- and interday precision CVs were <15%, with accuracy between −11% and 14%. All analytes were evaluated with run times of <5 minutes. All analytes were stable under the experimental conditions used in this study, with reproducible results (93%–100% of the nominal concentration) after 24 hours at room temperature. Experimentally determined concentration values in plasma samples were considered acceptable with precision and accuracy higher than 85%.
Canine Albumin Gene Sequencing
The canine albumin cDNA sequence has been reported for 26 Beagle dogs, identifying two linked SNPs at an allele frequency of 40% (Ito et al., 2009). Since that initial report included only one dog breed, we sequenced all 14 exons of the canine albumin gene using genomic DNA from 100 dogs consisting of 37 breeds. DNA samples were obtained from the WSU College of Veterinary Medicine DNA Bank. By sequencing a wider variety of breeds, we intended to identify previously unreported polymorphisms in the canine albumin gene.
Primers were designed to amplify all 14 canine albumin exons from genomic DNA. National Center for Biotechnology Information (http://www.ncbi.nlm.nih.gov) genomic DNA reference sequence NW_003726083 was used to design oligonucleotide primers and develop sequencing strategies for this study. The mRNA reference sequence (AB090854) was used for comparison. Primers were designed to anneal in the flanking intron DNA sequences to amplify each exon using the PrimerQuest online design tool (http://www.idtdna.com/Primerquest). Standard polymerase chain reaction (PCR) amplifications were carried out using 20 ng genomic DNA.
PCR products were visualized on a 1.2% agarose gel before the products were treated with ExoSAP-IT reagent (Affymetrix, Cleveland, OH) according to the manufacturer's directions. Treated PCR products were Sanger dye terminator sequenced with the same primers used for amplification using Big Dye 3.1 sequencing mix as directed (Applied Biosystems/Life Technologies, Grand Island, NY). Sequence data were analyzed using Sequencher 5.2 software (Gene Codes Corp., Ann Arbor, MI). Nucleotide sequences were evaluated for differences from the canine reference genome (CanFam3.1).
Canine Albumin c.1075G>T and c.1422A>T Genotyping
Canine albumin genotypes at two previously reported polymorphic sites (c.1075G>T and c.1422A>T) were determined using genomic DNA obtained from the WSU College of Veterinary Medicine DNA Bank. DNA samples were randomly selected to include 1446 dogs representing 61 different breeds (minimum of 10 dogs per breed) and 136 mixed-breed dogs. The breed of dog was based on owner designation at the time of admission to the WSU Veterinary Teaching Hospital. Genotyping was conducted using TaqMan allele discrimination assays (Applied Biosystems/Thermo Fisher Scientific, Foster City, CA) according to the manufacturer's directions. The primers and probes for c.1075G>T were 5′-CAA GTA AAA CTA TTT CAT TTT CAT CTG AAT CAG GT-3′ (forward primer), 5′-TGG CGA GTC TCA AAA GCA ATG A-3′ (reverse primer), 5′-TTG TAT GAA TAC GCA AGA AG-3′ (G-allele probe labeled with Vic), and 5′-TTT TGT ATG AAT ACT CAA GAA G-3′ (T-allele probe labeled with Fam). The primers and probes for c.1422A>T were 5′-GGC ACC AAA TGT TGT AAG AAA CCT-3′ (forward primer), 5′-GTT AGC TTT GTC CAT TTC TAA AGG CAA A-3′ (reverse primer), 5′-TGT GCT GAA GAC TTT-3′ (A-allele probe labeled with Vic), and 5′-CTG TGC TGA TGA CTT T-3′ (T-allele probe labeled with Fam). Genotype results for the dogs included in the study were confirmed with two separate runs.
All experimental measurements regarding the extent of plasma protein binding are reported as means ± S.D. (n = 12) unless otherwise noted. In vitro experiments were performed with two (D01-4582) or three (celecoxib, mycophenolic acid, and meloxicam) replicates per sample, randomly distributed in each run, on 3 separate days. Descriptive and comparative analysis was performed using Microsoft Office Professional Plus Excel 2013 (Microsoft Corporation, Redmond, WA) and SigmaPlot 12 software (Systat, San Jose, CA) to analyze and report study results. ChemDraw 18.1 software (Cambridge, MA) was used to construct Fig. 1. GraphPad Prism 3.03 software (La Jolla, CA) was used to construct Fig. 3. Normality of the unbound concentrations for each analyte was assessed with the Shapiro–Wilk test. Statistical power analysis was performed for sample size estimation, based on data from Ito et al. (2009), comparing dogs with and dogs without c.1075G>T and c.1422A>T variants. Median unbound fractions for all drugs were compared using the Mann–Whitney U test to determine differences in the extent of plasma protein binding between dogs with haplotypes H1 and H2. Statistical differences between the mean serum albumin concentration of each group were evaluated with the unpaired t test. For all comparisons, statistical significance was set at ≤0.05.
Canine Albumin Exon Sequencing
All 14 exons in the canine albumin gene were sequenced using DNA samples from 100 dogs representing 37 different breeds, and the results were compared with the reference sequence (NW_003726083). Six exons (2, 6, 7, 10, 12, and 14) showed no variation from the reference sequence, whereas eight exons (1, 3, 4, 5, 8, 9, 11, and 13) contained SNPs. Of the SNPs identified, 13 were synonymous (data not shown) and 3 were nonsynonymous (Table 3). Among the three nonsynonymous SNPs, c.268C>T was relatively uncommon. On the other hand, c.1075G>T and c.1422A>T were present in 90 of 100 dogs sequenced. Because of the low frequency of the uncommon nonsynonymous SNPs, they were not investigated further.
Canine albumin gene nonsynonymous SNPs identified by sequencing all exons in 100 dogs representing 37 different breeds
Effect of H1 and H2 Allelic Variation on Drug Binding to Plasma
Drug plasma protein binding was evaluated in plasma samples collected from dogs homozygous for the H1 allele (n = 6) and the H2 allele (n = 6). Plasma albumin concentrations in all dogs tested (n = 12) were considered within normal limits (range, 2.5–3.3 mg/dl), with no statistically significant difference identified between the H1 and H2 allele groups (P = 0.41). The extent of drug binding (expressed as the mean ± S.D. percent bound) for all 12 dogs was 99.93 ± 0.04 for D01-4582, 98.22 ± 0.3 for celecoxib, 95.36 ± 1.1 for mycophenolic acid, and 94.90 ± 1.72 for meloxicam.
Considering the high extent of protein binding of the compounds analyzed as well as the pharmacological importance of unbound (free) drug, free drug fractions for each genotype are reported (Fig. 2). Unbound fraction values calculated for each plasma-drug combination had a CV < 20% (data not shown). Median unbound drug fractions with D01-4582 and celecoxib were considerably smaller (<2%) than those determined for mycophenolic acid and meloxicam. Free drug fractions in plasma from dogs with the albumin H1 allele ranged from 0.02% to 0.13% (mean 0.06% ± 0.04%) for D01-4582, 1.72% to 2.43% (1.86% ± 0.34%) for celecoxib, 4.03% to 7.80% (6.10% ± 1.29%) for meloxicam, and 3.92% to 6.51% (5.17% ± 0.83%) for mycophenolic acid. Plasma from dogs with albumin H2 allele had free drug fractions between 0.04% and 0.18% (0.09% ± 0.05%) for D01-4582, 1.41% and 2.00% (1.69% ± 0.19%) for celecoxib, 2.38% and 5.93% (4.10% ± 1.55%) for meloxicam, and 2.75% and 5.35% (4.10% ± 1.11%) for mycophenolic acid. The difference in free drug fraction between albumin genotypes reached statistical significance for meloxicam only (P = 0.0411).
Scatter dot plots of unbound fractions of D01-4582 (A), celecoxib (B), meloxicam (C), and mycophenolic acid (D), in relation to the albumin haplotype (H1 and H2) for each dog evaluated (n = 6 per group; average of three replicates from three separate runs). Lines represent the median unbound fraction for each compound. The difference between the median unbound fraction for both haplotypes was statistically significant for meloxicam (P = 0.041) but not the other compounds analyzed (Mann–Whitney U test, significance set at *P < 0.05). ALB, albumin.
Canine Albumin c.1075G>T and c.1422A>T Haplotypes and Breed Distribution
c.1075G>T and c.1422A>T genotypes were determined in 1260 dogs from 61 different breeds and 186 mixed-breed dogs. Haplotypes that were resolved by inference are shown in Table 4 and haplotype frequencies for each breed are given in Supplemental Table 1. As shown in Fig. 3, the frequency distribution of albumin haplotypes varied greatly across dog breeds. H1 (reference) and H2 (variant for both SNPs) were the predominant haplotypes in all dog breeds examined. Haplotype H1 was most frequently detected in Cocker Spaniels (95%), Miniature Dachshunds (89.5%), and Italian Greyhounds (75.6%), whereas haplotype H2 was most common in Great Danes (100%), Basenjis (98%), Rottweilers (97.5%), Golden Retrievers (97.1%), Boxers (95%), and Yorkshire Terriers (95%). Median and interquartile range (IQR) frequencies for haplotypes H1 and H2 across all breeds were 39% (25%–50%) and 60% (50%–71%), respectively. By comparison, H1 and H2 haplotype frequencies in mixed-breed dogs were 37% and 63%, respectively. Haplotype H3 (c.1075G>T) was found in nine breeds, including Cirneco dell'Etna, Pomeranian, Portuguese Podengo, Weimaraner, Pembroke Welsh Corgi, Australian Shepherd, Rhodesian Ridgeback, Peruvian Inca Orchid, and Afghan Hound, with a median breed frequency of 5.9% (IQR, 2.8%–14%). Haplotype H4 (c.1422A>T) was found in 11 breeds, including Afghan Hound, Border Collie, Siberian Husky, Borzoi, Pharaoh Hound, Greyhound, Longhaired Whippet, Whippet, Italian Greyhound, Silken Windhound, and Scottish Deerhound, with a median breed frequency of 1.8% (IQR, 1.1%–4.0%). Afghan Hound was the only breed with all four haplotypes detected. Haplotypes H3 and H4 were not found in any of the 186 mixed-breed dogs tested.
Canine albumin c.1075G>T and c.1422A>T haplotypes
Albumin H1, H2, H3, and H4 allele frequencies determined for 61 dog breeds and mixed-breed dogs. The number of dogs sampled per breed is given in parentheses.
In our study, canine albumin genotype affected plasma protein binding of meloxicam, with no significant difference detected between dogs with mean free fractions of the albumin H1 allele (reference) and the H2 allele (c.1075G>T and c.1422A>T) for the other three drugs assessed (D01-4582, celecoxib, and mycophenolic acid). Drugs selected for our study had high preferential binding to albumin sites I or II and a narrow therapeutic index (Er et al., 2013). Meloxicam and mycophenolic acid were used in our experiments due to their preferential binding in humans to albumin site I and site II, respectively (Nowak and Shaw, 1995; Er et al., 2013). In addition, celecoxib and D01-4582 were used because of their reported pharmacokinetic variability in dogs that could be associated with differences in protein binding (Paulson et al., 1999a; Ito et al., 2009).
Our results were consistent with the mean extent of plasma protein binding previously reported for dogs (99.9% for D01-4582, 97% for meloxicam, 98.5% for celecoxib, and 95%–97% for mycophenolic acid) (Busch et al., 1998; Paulson et al., 1999b; Ito et al., 2009; Morassi et al., 2018). For D01-4582, our findings differ from a previous report (Ito et al., 2009) in which free drug fractions were greater in plasma from Beagles with the albumin H2 allele than in plasma from Beagles with the H1 allele. Free fractions of D01-4582 in Beagles were reported to be 0.029% ± 0.017% (CV = 59%, n = 35), with dogs with the albumin H1 allele having statistically significant lower values (0.015% ± 0.003%) compared with dogs with the albumin H2 allele (0.059% ± 0.016%) (Ito et al., 2009). In our study population, D01-4582 free fractions were somewhat higher (mean 0.07% ± 0.04%), with no statistically significant difference between dogs with the albumin H1 allele and those with the albumin H2 allele. There are some possible explanations for the contrasting results between the two studies. First, there are differences in the canine populations assessed. Whereas a single research Beagle colony was evaluated in the previously published study, our experiments included plasma from multiple dog breeds. Lower free fractions of D01-4582 may be common in Beagles; however, the inclusion of different breeds, representing greater genetic heterogeneity, likely increased the group biologic variability and resulted in higher free drug fractions. Evaluating plasma protein binding in dogs from multiple breeds likely provides a more comprehensive and representative assessment of the overall canine population; however, the higher variability in free drug fractions may have contributed to the lack of statistical differences between groups. A second explanation is associated with the potential variability of lipid fraction in the plasma from our study population. D01-4582 is a highly lipophilic drug with plasma protein binding >99%, which makes it likely to bind to lipoproteins in the plasma. Ultracentrifugation leads to the formation of a superficial chylomicron layer that can make the collection of supernatant difficult. In plasma samples, the presence and size of this chylomicron layer can lead to overestimation of free drug fraction. Even though plasma samples were collected while fasting, they were obtained from dogs from a variety of breeds, food regimen and body condition scores that likely contributed to a lipid profile more diverse than the one obtained from the research Beagle colony used by Ito et al. (2009). Breed, as well as diet and body condition score, has been identified as a source of difference in plasma lipids in dogs and could have contributed to the higher biologic variability in plasma lipid content in our samples compared with colony research animals (Downs et al., 1993; Jeusette et al., 2005). For celecoxib, our findings suggest that albumin genotype is unlikely to have caused the pharmacokinetic variability reported by Paulson et al. (1999a).
Multiple factors other than genotype can affect binding to albumin in plasma and may have contributed to the differences noted between dogs. Endogenous compounds in plasma (e.g., globulins, cells, or metabolites) can influence free drug fractions by competitively displacing drugs from albumin binding sites and/or inducing conformational changes in the protein (Otagiri, 2005). For example, bilirubin and free fatty acids can competitively bind to human albumin and increase free drug fractions of the antibiotic cefazolin in vitro (Decroix et al., 1988). Although the content of other plasma components in study samples was not controlled, we estimate their impact to be small for celecoxib, meloxicam, and celecoxib considering their high binding preference for albumin at the drug concentrations used in our experiments (below the reported Cmax in dogs). Lower albumin concentrations can also increase drug free fractions, in particular for drugs highly bound to albumin (Ikenoue et al., 2000). Plasma albumin concentrations in our study were considered normal and unlikely to affect our results, with no statistical difference in albumin concentrations between allele groups. Albumin can also undergo post-translational modifications (e.g., glycation, cysteinylation, and carbamoylation) that can cause structural changes to the molecule and affect albumin's ability to interact with some drugs (Lee and Wu, 2015). These were not assessed in our study population. Sex-related differences in our plasma protein binding experiments were not statistically significant (P > 0.05).
Albumin genotype seemed to impact the plasma protein binding of meloxicam in our ex vivo study, as dogs with the albumin H1 allele had a significantly higher free drug fraction than dogs with the albumin H2 allele (P = 0.041). This contrasts the initial hypothesis that the albumin H1 allele would be associated with lower free drug fractions (and therefore higher binding). Unlike the other three drugs tested in our study, meloxicam is the only one that binds to albumin site I in humans (Er et al., 2013). Although the preferential binding site for these drugs has not been extensively evaluated in canine albumin, it is possible that the role of albumin genotype in plasma protein binding is drug dependent. While the amino acid changes associated with the albumin H2 allele (c.1075G>T and c.1422A>T) may decrease binding of drugs commonly bound to site II, they may have the opposite effect on drugs bound to a different region of the albumin molecule in dogs. The difference in the mean free drug fraction of meloxicam between albumin genotypes noted ex vivo may be relevant and should be confirmed in vivo. Although changes in free drug fraction can affect total drug concentrations in vivo, the impact on free drug concentrations and pharmacologic effect is usually minor.
Another new finding of our study is that c.1075G>T and c.1422A>T polymorphisms are independent, occurring separately in a small percentage of dogs. Although median allele frequencies in Beagles in our study population were similar to those in a previous report (Ito et al., 2009), they varied considerably between dog breeds. This difference in allele frequencies may be particularly important for veterinary drug development programs since single-breed studies may introduce bias in preclinical results.
Interestingly, the presence of more than one form of canine albumin was first reported in 1985, with two albumin phenotypes ("slow" and "fast") identified based on their electrophoretic mobility in a large population of dogs (Christensen et al., 1985). The H2 allele breed frequencies from our study had a strong correlation (Rs = 0.86; P < 0.0001) with the corresponding frequencies for the slow form reported by Christensen et al. (1985) (Fig. 4). This indirectly suggests that the slow migration pattern of the S variant could result from the two predicted amino acid changes associated with the presence of the H2 allele. Unfortunately, we were unable to reproduce the starch gel electrophoresis technique used in the previous report to determine the electrophoretic mobility (phenotype) of the dogs used in our study.
Comparison of H2 haplotype frequencies determined in this study with frequencies of the albumin electrophoretic mobility slow (S) form allele reported previously by Christensen et al. (1985) for the same dog breeds. Also shown is the Spearman correlation coefficient and associated P value (significance set at P < 0.05).
The findings of our study should be interpreted considering its limitations. Comparing binding properties between genetic variants was previously done using a small number of marker ligands for different binding regions of human serum albumin (Kragh-Hansen et al., 1990; Vestberg et al., 1992). Although binding properties of human albumin have been extensively investigated, similar information regarding canine albumin is scarce. Canine albumin is expected to follow the same tertiary molecular structure as human albumin; however, it may differ in the number and specificity of binding sites (Kosa et al., 1997). Therefore, it is possible that the drugs we intended to interrogate sites I and II may actually bind to different sites on canine albumin. Our findings cannot predict the effect of c.1075G>T and c.1422A>T polymorphisms on specific binding sites and/or on plasma protein binding of other compounds. For that reason, additional studies evaluating the impact of albumin genotype on the extent of plasma protein binding of a large variety of drugs are recommended. Although the difference in plasma protein binding of meloxicam seen in our ex vivo study suggests a significant impact of albumin genotype on free drug fraction, this should be confirmed with in vivo studies. The lack of significant differences in free drug fractions of D01-4582, celecoxib, and mycophenolic acid between albumin genotypes in our study is likely unrelated to the number of dogs evaluated in each group (n = 6), since the previous report (Ito et al., 2009) included only five dogs with the H2 allele that showed statistically higher free drug fractions of D01-4582 compared with dogs with the H1 allele.
In conclusion, this study provides novel information on the frequency and pharmacokinetic impact of the most common nonsynonymous canine albumin polymorphisms (c.1075G>T and c.1422A>T) across multiple breeds. In our study, dogs with the albumin H1 allele had significantly higher free drug fractions (and consequently lower plasma protein binding) of meloxicam than dogs with the albumin H2 allele. Albumin alleles H1 and H2 seem to be the most frequent across breeds and c.1075G>T and c.1422A>T are not completely linked, a finding that has not been previously reported. Our findings suggest that albumin genotype may influence free drug fractions at physiologically relevant concentrations of select drugs in dogs. Because changes in plasma protein binding commonly do not affect free drug concentrations, the significance of the influence of albumin genotype in vivo will depend on the drug studied and its intrinsic clearance. However, albumin genotyping may improve the interpretation of pharmacokinetic data generated in canine research populations during the developmental process of human and veterinary drugs.
We thank Dr. Jane Wardrop and Dr. Leigh Knodler (WSU) for providing the canine plasma used for method development and access to the ultracentrifuge, respectively. We thank Dr. Ito (Daiichi Sankyo Co. Ltd.) for providing a sample of D01-4582 for our study, as well as Dr. Sol Rivera and Darren Schnider for technical assistance.
Authorship Contributions
Participated in research design: Costa, Court, Mealey, Villarino.
Conducted experiments: Costa, Burke, Zhu.
Contributed new reagents or analytic tools: Court, Mealey, Villarino.
Performed data analysis: Costa, Court, Mealey, Villarino.
Wrote or contributed to the writing of the manuscript: Costa, Court, Mealey, Villarino.
Received March 26, 2019.
Accepted June 7, 2019.
This work was supported by Wisdom Health, Mars Petcare, and the Washington State University Program in Individualized Medicine [(to A.P.C.)]. M.H.C. was supported by the National Institutes of Health National Institute of General Medical Sciences [Grant R01-GM-102130] and the William R. Jones Endowment to Washington State University College of Veterinary Medicine.
https://doi.org/10.1124/dmd.119.087304.
This article has supplemental material available at dmd.aspetjournals.org.
coefficient of variation
D01-4582
4-[1-[3-chloro-4-[N′-(2-methylphenyl) ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidine-2-yl]methoxybenzoic acid
high-performance liquid chromatography
tandem mass spectrometry
polymerase chain reaction
Copyright © 2019 by The Author(s)
This is an open access article distributed under the CC BY-NC Attribution 4.0 International license.
Busch U,
Schmid J,
Heinzel G,
Schmaus H,
Baierl J,
Huber C, and
Roth W
(1998) Pharmacokinetics of meloxicam in animals and the relevance to humans. Drug Metab Dispos 26:576–584.
Christensen K,
Arnbjerg J, and
Andresen E
(1985) Polymorphism of serum albumin in dog breeds and its relation to weight and leg length. Hereditas 102:219–223.
Cooper DL,
Wood RC 3rd.,
Wyatt JE, and
Harirforoosh S
(2014) Pharmacokinetic interactions between rebamipide and selected nonsteroidal anti-inflammatory drugs in rats. Eur J Pharm Sci 53:28–34.
Decroix MO,
Zini R,
Chaumeil JC, and
Tillement JP
(1988) Cefazolin serum protein binding and its inhibition by bilirubin, fatty acids and other drugs. Biochem Pharmacol 37:2807–2814.
Downs LG,
Bolton CH,
Crispin SM, and
Wills JM
(1993) Plasma lipoprotein lipids in five different breeds of dogs. Res Vet Sci 54:63–67.
Er JC,
Vendrell M,
Tang MK,
Zhai D, and
Chang YT
(2013) Fluorescent dye cocktail for multiplex drug-site mapping on human serum albumin. ACS Comb Sci 15:452–457.
Ikenoue N,
Saitsu Y,
Shimoda M, and
Kokue E
(2000) Disease-induced alterations in plasma drug-binding proteins and their influence on drug binding percentages in dogs. Vet Q 22:43–49.
Ito T,
Takahashi M,
Sudo K, and
Sugiyama Y
(2009) Interindividual pharmacokinetics variability of the alpha(4)beta(1) integrin antagonist, 4-[1-[3-chloro-4-[N′-(2-methylphenyl)ureido]phenylacetyl]-(4S)-fluoro-(2S)-pyrrolidine-2-yl]methoxybenzoic acid (D01-4582), in beagles is associated with albumin genetic polymorphisms. J Pharm Sci 98:1545–1555.
Jeusette IC,
Lhoest ET,
Istasse LP, and
Diez MO
(2005) Influence of obesity on plasma lipid and lipoprotein concentrations in dogs. Am J Vet Res 66:81–86.
Jolliet P,
Simon N,
Brée F,
Urien S,
Pagliara A,
Carrupt PA,
Testa B, and
(1997) Blood-to-brain transfer of various oxicams: effects of plasma binding on their brain delivery. Pharm Res 14:650–656.
Kosa T,
Maruyama T, and
Otagiri M
(1997) Species differences of serum albumins: I. Drug binding sites. Pharm Res 14:1607–1612.
Kragh-Hansen U,
Brennan SO,
Galliano M, and
Sugita O
(1990) Binding of warfarin, salicylate, and diazepam to genetic variants of human serum albumin with known mutations. Mol Pharmacol 37:238–242.
Lapicque F,
Vergne P,
Jouzeau JY,
Loeuille D,
Gillet P,
Vignon E,
Thomas P,
Velicitat P,
Türck D,
Guillaume C, et al.
(2000) Articular diffusion of meloxicam after a single oral dose: relationship to cyclo-oxygenase inhibition in synovial cells. Clin Pharmacokinet 39:369–382.
Lee P and
(2015) Review: modifications of human serum albumin and their binding effect. Curr Pharm Des 21:1862–1865.
OpenUrlCrossRef
Morassi A,
Rivera-Vélez SM,
Slovak JE,
Court MH, and
Villarino NF
(2018) Ex vivo binding of the immunosuppressant mycophenolic acid to dog and cat plasma proteins and the effect of co-incubated dexamethasone and prednisolone. J Vet Pharmacol Ther 41:513–521.
Nowak I and
Shaw LM
(1995) Mycophenolic acid binding to human serum albumin: characterization and relation to pharmacodynamics. Clin Chem 41:1011–1017.
(2005) A molecular functional study on the interactions of drugs with plasma proteins. Drug Metab Pharmacokinet 20:309–323.
Paulson SK,
Engel L,
Reitz B,
Bolten S,
Burton EG,
Maziasz TJ,
Yan B, and
Schoenhard GL
(1999a) Evidence for polymorphism in the canine metabolism of the cyclooxygenase 2 inhibitor, celecoxib. Drug Metab Dispos 27:1133–1142.
Kaprak TA,
Gresk CJ,
Fast DM,
Baratta MT,
Breau AP, and
Karim A
(1999b) Plasma protein binding of celecoxib in mice, rat, rabbit, dog and human. Biopharm Drug Dispos 20:293–299.
Vestberg K,
Galliano M,
Minchiotti L, and
Kragh-Hansen U
(1992) High-affinity binding of warfarin, salicylate and diazepam to natural mutants of human serum albumin modified in the C-terminal end. Biochem Pharmacol 44:1515–1521.
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Canine Albumin Polymorphisms and Plasma Drug Binding
Drug Metabolism and Disposition October 1, 2019, 47 (10) 1024-1031; DOI: https://doi.org/10.1124/dmd.119.087304

redpajama_set_name: RedPajamaCommonCrawl


The mental health epidemic that has exploded within the corporate behemoth is the direct result of the brutal exploitation of its workforce.
The censorship measures being deployed ineffectually against Tommy Robinson are being prepared for use against the left.
Джерри Уайт, 15 марта 2019 г.
On March 14, 1994, the labor and finance ministers of the Group of Seven leading industrialized countries met in Detroit, Michigan on the false promise of "creating jobs" amid the worst economic downturn and highest unemployment rates worldwide since the Great Depression.


Mixed Beans by Fran W.
The Imagination Tree: Grow a Bean Plant on Cotton Balls – Instructions and illustrations.
Oklahoma 4H: A Bean is a Seed – Life cycle of a plant – classroom activities.
Kean University: Bean Plant Activities – Site offers various activities.
Buzzle: Life Cycle of a Bean Plant – Site offers diagrams of various stages of a beans life cycle and offers explanations for each one.
Wikipedia: Bean – Site offers general information about beans, including cultivation and history.
Fran's World of Discovery: Three Sisters: Beans – Bean soup recipe and bean activities offered on this site.
Cook's Thesaurus: Dry Beans – This is a excellent resource for anyone interested in learning about various types of beans.
Enchanted Learning: Bean Diagram – Printable and ready to label. Word bank and answers available.
Kiddie Gardeners: Make a Bean Tee-pee – Instructions for making a bean tee-pee in your backyard.
Teaching Ideas: Bean Activities – Site offers bean activities for active young people.
Artists Helping Children: Seed and Bean Projects – Bean box, noise makers, crafts and other projects.
Bright Hub Education: Preschool Science Fun with a Lima Bean: Grow a Plant!
Wellness Proposals: Children's Learning Activities Using Dry Beans – Activities for very young children.


text:
China's Tianjin city to extend paternity leave
Authorities in the city have launched a revision on the local regulations of population and family planning.
By: Claire Lee | September 27, 2021
Topics: China | Compensation and Benefits | Employee Experience | Health and Wellness | News
Tianjin city in China is looking to extend paternity leave to encourage childbirth in response to China's new three-child policy.
Authorities in the Chinese city have launched a revision on the local regulations of population and family planning and aims to extend paternity leave for men. Currently, the city only awards seven days of paternity leave for fathers, writes the Global Times.
Paternity leave in China is stipulated by provincial family planning regulations, is not available in every region and differs from province to province.
Provinces like Henan, Gansu and Yunnan have the longest paternity leaves of 30 days, while Tianjin and Shandong Province have the shortest, with only seven days, according to Workers' Daily.
READ: 9.38 million jobs created in China this year till August
In May, China announced that it would allow couples to have up to three children. The policy was formally passed into law in August. The government is also set to roll out more support measures to ease the burden on families on childbearing, childcare, and education.
Indonesia plans wage subsidies for workers
The Ministry of Manpower is preparing the policy for distribution of the BSU wage subsidy programme for workers in 2021.
Wage growth in Australia unlikely despite labour shortage
The Reserve Bank of Australia noted that in sectors where wage growth was observed, it was largely due to earlier implemented wage cuts that were reversed.


Ignacio Ruvalcaba current General Manager over all youth development at professional Liga MX team Club Tijauana Xoloitzcuintles de Caliente, made his first trip to Utah to launch the newly organized official affialte Club Xoloitzcuintles Academy Utah(Xolos Utah).
The visit included attending many important events including press conferences, interviews with news outlets, official visits with state leaders as well as formal and informal meetings with club parents, players and staff.
"Professor" (title used in Mexico that shows respect for someone of great experience) Ignacio Ruvalcaba arrived Monday morning July 31 and was taken to Provo Utah to begin the 1st of 3 days of meetins with Xolos Utah. Over the next 3 days he interacted with the leadership of Club Xoloitzcuintles Utah including President Jon Baker, Sports Director, Salvador Perez, Academy Director Arturo Ochoa. He also met with board members and club administrators Renelle Smith and Veronica Cortez as well as individual team administrators.
The goal of the visit was to train Club Xoloitzcuintles Utah staff and coaches in the Club Tijuana Xoloitzcuintles methodology and as such the following took place.
General meeting with club club board, club administration and team administrators.
Given the feedback the club received Professor Ignacio Ruvalcaba left very impressed with the organization and is excited about the future of Club Xoloitzcuintles Academy Utah.
It was a great pleasure for Xoloitzcuintles Utah to host Professor Ignacio Ruvalcaba and we are left with a greater commitment to maximize the potential and talent that exists in the club, be it as players or as better people. With a strong group effort we will reach the goals we have set and will work to be the most successful club in Utah.


text:
import sys, re

import runtime, parsetree

def generate(inputfilename, outputfilename='', dump=0, **flags):
    """Generate a grammar, given an input filename (X.g)
    and an output filename (defaulting to X.py)."""

    if not outputfilename:
        if inputfilename.endswith('.g'):
            outputfilename = inputfilename[:-2] + '.py'
        else:
            raise Exception('Must specify output filename if input filename is not *.g')

    DIVIDER = '\n%%\n' # This pattern separates the pre/post parsers
    preparser, postparser = None, None # Code before and after the parser desc

    # Read the entire file
    s = open(inputfilename,'r').read()

    # See if there's a separation between the pre-parser and parser
    f = s.find(DIVIDER)
    if f >= 0: preparser, s = s[:f]+'\n\n', s[f+len(DIVIDER):]

    # See if there's a separation between the parser and post-parser
    f = s.find(DIVIDER)
    if f >= 0: s, postparser = s[:f], '\n\n'+s[f+len(DIVIDER):]

    # Create the parser and scanner and parse the text
    scanner = grammar.ParserDescriptionScanner(s, filename=inputfilename)
    if preparser: scanner.del_line += preparser.count('\n')

    parser = grammar.ParserDescription(scanner)
    t = runtime.wrap_error_reporter(parser, 'Parser')
    if t is None: return 1 # Failure
    if preparser is not None: t.preparser = preparser
    if postparser is not None: t.postparser = postparser

    # Check the options
    for f in t.options.keys():
        for opt,_,_ in yapps_options:
            if f == opt: break
        else:
            print >>sys.stderr, 'Warning: unrecognized option', f
    # Add command line options to the set
    for f in flags.keys(): t.options[f] = flags[f]

    # Generate the output
    if dump:
        t.dump_information()
    else:
        t.output = open(outputfilename, 'w')
        t.generate_output()
    return 0

if __name__ == '__main__':
    import doctest
    doctest.testmod(sys.modules['__main__'])
    doctest.testmod(parsetree)

    # Someday I will use optparse, but Python 2.3 is too new at the moment.
    yapps_options = [
        ('context-insensitive-scanner',
         'context-insensitive-scanner',
         'Scan all tokens (see docs)'),
        ]

    import getopt
    optlist, args = getopt.getopt(sys.argv[1:], 'f:', ['help', 'dump', 'use-devel-grammar'])
    if not args or len(args) > 2:
        print >>sys.stderr, 'Usage:'
        print >>sys.stderr, '  python', sys.argv[0], '[flags] input.g [output.py]'
        print >>sys.stderr, 'Flags:'
        print >>sys.stderr, ('  --dump' + ' '*40)[:35] + 'Dump out grammar information'
        print >>sys.stderr, ('  --use-devel-grammar' + ' '*40)[:35] + 'Use the devel grammar parser from yapps_grammar.py instead of the stable grammar from grammar.py'
        for flag, _, doc in yapps_options:
            print >>sys.stderr, ('  -f' + flag + ' '*40)[:35] + doc
    else:
        # Read in the options and create a list of flags
        flags = {}
        use_devel_grammar = 0
        for opt in optlist:
            for flag, name, _ in yapps_options:
                if opt == ('-f', flag):
                    flags[name] = 1
                    break
            else:
                if opt == ('--dump', ''):
                    flags['dump'] = 1
                elif opt == ('--use-devel-grammar', ''):
                    use_devel_grammar = 1
                else:
                    print >>sys.stderr, 'Warning: unrecognized option', opt[0], opt[1]

        if use_devel_grammar:
            import yapps_grammar as grammar
        else:
            import grammar

        sys.exit(generate(*tuple(args), **flags))

redpajama_set_name: RedPajamaGithub


to be staged at Ouseday raft race and regatta on Sunday July 1.
"shout out" at the start of the regatta at Malling Recreation Ground in Lewes.
the service's Saturday Music School.
and equipment for young bands and other musicians.
Ouseday will also be supporting A Band of Brothers, which mentors young men in crisis.
managed by Trevor Cutler, station officer for Newhaven Marine Coastguard.
On board Wil Wilson's fishing boat are: BHASVIC pupil Heather Owolabi, aged 16, flute; Gina Owolabi, 13, violin, Ringmer Academy; James West, 13, flute and recorder, Priory School; Julian Falkowski, seven, and Zofia Falkowski, 11, from Iford and Kingston; James Wilson, ten, violin, Harbour Primary; Brian Wilson, five, violin, Harbour; Thomas Fowler, 13, drums, violin and keyboards, Seahaven Academy; Louis Fisher, 16, saxophone, BHASVIC; Tom Van Howe, 16, sax, Priory; Maya Biddulph, 17, violin, Varndean; Miranda Raffo, 16, sax, Priory; Georgia Bickley, seven, violin, South Malling; Olive Reeves, eight, cello, South Malling; Jack Humberstone, 13, trombone,Willingdon Community College. Esme Parsons, six, violin.
Same children, also Esme Parsons, six, violin, LOGS, and teachers Jane Humberstone , cornet, and Malcolm Warnes, trumpet.


#customSBS Can you guess what ski this is destined for? @sbs_trailers @robertowenmarine #pwc trailer! Remember if you want a custom trailer for any PWC we can help you plan your custom SBS trailer! We have a full colour swatch card here and examples on site.
Just Landed! 2017 Seadoo Trixx with only 26hrs use £6995 fully serviced and 3 months warranty.


text:
Arcade Hall of…
Arcade Hall of Fame: Burger Time
Developed and published by Data East in 1982, Burger Time is an arcade game unlike any other you've probably played.
Combining elements of a platforming game with a puzzle game, Burger Time puts you in the chef's hat of Peter Pepper who is tasked with putting together mattress sized burgers while avoiding enemies with iconically cute names such as Mr. Hot Dog, Mr. Pickle, and Mr. Egg.
There are number of ways to avoid enemies as well as get rid of them if you need to do that. You can crush them beneath ingredients or send them falling to their demise. The possibilities are pretty endless. There is a method to the madness however because Burger Time is all about points – and lots of them.
You get more points for doing certain actions over others and this causes the player to modify their play style to accommodate this. For example, it may be easier for you to crush an enemy but you will get more points for "dropping" it so you will modify your play style to allow for this.
Peter Pepper can temporarily stun enemies using a shake of pepper but, aside from the above mentioned ways of dispatching with foes, is otherwise quite vulnerable. The game runs on a loop of six different stage variants that change as Peter Pepper completes each, cycling through the loop continuously with increasing difficulty as the player progresses. Like other platforming games, Peter Pepper's playtime is based upon a lives system which can be gained and lost in the course of the game.
Coming out in the same year as Zaxxon and Donkey Kong Jr., Burger Time had some tough competition when it hit the arcade scene. The arcades, largely driven by series that created fads that spawned endless variants on some theme, were not the easiest place for a new and novel concept like Burger Time to succeed. After all, this is the year that Super Pac Man, the third title in the series, was released and if you're thinking it was radically different from other Pac Man games then you'd be mistaken.
Variety was not the spice of life in the risky arcade business. Tried-and-true models were what many companies preferred, but Data East's gambit proved a success and Burger Time was not only a commercial success during its time in the arcades but also the recipient of great critical acclaim as well, garnering enough attention to make it a hot property for the home console market.
Today Burger Time exists as a kind of snapshot in time of what the video game industry was in the arcades. You see in Data East's design language for the game the trend towards more heavily graphical sprites and away from the more minimalistic approaches of the past. One could easily be forgiven for mistaking this game for a Famicom game though that system still had one year until it hit the streets but the spirit is there.
A common occurrence today, Burger Time's merger of two disparate genres in such an elegant way stands the test of time and sets the game apart as a true classic of the arcades.
Relive the iconic Burger Time arcade game with the Burger Time Mini Arcade Machine!
Headliners Retro Life
Guides Headliners


Rename the file "Full Name Course Code Repair Portrait Photo.jpg".
Using Affinity Photo or GIMP, edit out all the imperfections in the photo.
Hand in to the relevant handin folder.
Note: be sure to leave a copy of the original image as the background layer.
Note: for students who have done this assignment previously, use this sailor photo (credit: http://goo.gl/sgbhVV) instead (note: this image is only available to students on the class mail list).
Want some extra challenges? See this website.


text:
Corrie Chen, filmmaker: some advice for film and television graduates at the University of Melbourne
Filmmaker Corrie Chen. By Tammy Law. Image supplied.
Award-winning filmmaker and television director Corrie Chen graduated from the Victorian College of the Arts with a Bachelor Film and Television (Hons) in 2008, and a Master in Film and Television (Narrative) in 2011. Taiwanese by birth, her work spans comedy, drama and documentary, and often explores issues of identity and belonging. The following is an edited extract of her speech in December 2019 to graduating Film and Television students from the University of Melbourne.
By Corrie Chen
The last time I was at the Victorian College of the Arts (VCA) it was a very different place. The industry was a different place. There were fewer students and courses. We were sending out festival screeners on physical DVDs, in the post. TV was NOT cool; it was a pretty undesirable career. And I was one of maybe two people of colour across all the entire Film and Television cohort.
Like I said, it was a different time, and obviously, with the awareness for inclusion these days, we now understand that is not good enough – but you know, I will say it was excellent preparation for the Australian film and TV industry.
I come from a long line of Asian stereotypes. My parents are restaurant owners, my granddad was a rice farmer. I am an immigrant and I didn't speak English until I was 11. Filmmaking was actually my dad's idea, much to his dismay later on in life. I desperately wanted to be an astronaut and I was obsessed with space.
I was this weird kid, with a thick accent and braces, alone in my room with a telescope. So – the first time I coerced some white Australians to come over for my birthday sleepover ('cause I bragged about my parent's Laser disk player) my dad got super excited at the prospect of me finally making friends and gave me his old video camera – a JVC hi-8. And that party changed my life – out of boredom, pre-internet, in the late 90s. We were high on sugar, and had binged on a bunch of 90s horror flicks before we decided to make the opening sequence of Scream, shot for shot, all done with in-camera editing. It was the most genius and shit thing you'll ever see in your life. That was my first movie.
We all have one of these stories. An origin story, when you first caught the bug. I was intoxicated, probably by the power of it. The power of controlling my own narrative, of who gets to be on screen, and whose story gets told. But it would take another ten years before I got to the VCA. I spent all of high school doubting this thing I loved, and I was so embarrassed to even say I wanted to be a director.
Of course my parents tried to talk me out of it; of course they hated this dream. But the biggest resistance actually came from within. How dare I have this ambition that seemed so bold and so entitled? How dare I want something that is so unlike what an Asian woman was expected to do? My world indoctrinated me with this culture of permission, or being trained to ask for what I want instead of just taking it.
I applied to VCA because I was lucky enough to have a part-time tutor – a white Australian man – in my undergraduate media degree who saw something in one of my shorts, and he said, "Hey, I reckon you should give VCA a go." That was the permission I needed.
My short film that I applied with was creatively and technically inept, and I knew it. But it was raw and angry. And it was an opportunity. I somehow fumbled my way through an interview and got into the VCA.
Champions come in many forms, and opportunities present themselves in many shapes, and that is something I'm still reminded of today.
At the VCA, both my short films were about the immigrant experience – because that was what I knew, what I lived. I never thought about diversity (that stupid word) as a storytelling tool. Inclusive storytelling wasn't a buzzword to me, it was my lived reality. I definitely didn't sit down and go, "Well, time to grapple with some diverse themes". This movement did not exist nine years ago.
The VCA storytelling mantra, other than "show don't tell", is "write what you know". I was simply desperate to see myself on screen, and adamant that no kid should ever experience what I did – spending the entirety of my teenage years wishing I'd wake up white. And the VCA helped me realise that being different, doesn't make my voice any less worthy. It was the first step in defining what my cinematic voice is, and that is a journey that still goes on today.
History will tell you that my VCA shorts did not set the world on fire. I didn't win a single award at the VCA, not for screenwriting or directing, or even best student. It did not launch me out of film school the way it did some of my peers – the way that I thought was my ONLY path after film school. In fact, the morning after my final VCA opening night, I sat up in bed, and I wept non-stop for 20 minutes. It was like someone died (well yes, my dreams).
I don't say this out of bitterness, even though I do love revenge success. I say this because I've learnt that nothing will ever be handed to you, and that everyone's paths will be different. I have had to create the path I wish I had to follow, but I had to work hard to figure out what it is I want.
It took six years from when I left the VCA to getting my first job hired as a TV director to do three episodes of a matchbox show – and it was a show I loved and believed in.
How lucky am I! I thought. Then I remember, no, it was six years of really shit work – of notetaking, assisting, of applying for any piece of funding I was eligible to apply for, of attachments, of being ignored, of networking, of lots and lots of spec scripts that never went anywhere, of self doubt, poverty, more self doubt, lost friendships and relationships, of bringing shame to my parents cause I was working in the arts, and so on.
You will want to give up. THAT VOICE will tell you that you can't. "Nobody has ever done it like that. Nobody who looks like you has ever done it that way."
But finding out what you want, and what your path looks like – that is part of the work. There is a path for you, even if it's not immediately obvious. Even if it's not in what you trained in.
Start by looking around you. Find your tribe, your trusted collaborators, and hang on to them for dear life. There will be someone in this room I guarantee you'll be working with on your first feature, on your first show. Community is hard to build, but it's really the rocket fuel that propels this industry.
That's one of the most important things I learnt – that this is a relationship business, and you will only get traction if people want to work with you again. AKA, don't be a dick.
I was so privileged to have had champions along the way – from the teachers at the VCA, to funding bodies, to my agent, to producers – people who reminded me to keep persevering, that hope is not hopeless. People have to want to support you, to understand what you want, because EVERYTHING is made through collaboration. It can't be a "me against them" attitude. Now I'm a little bit older and wiser I've realised it's far easier to destroy than it is to build. It has taken the generosity of others to help me break through the door, get a seat at the table, and finally start building my own table.
I'm so excited and lucky to be a part of a time where anything feels possible. The industry definition of a director has traditionally not been someone who looks like me. The industry that I love is not one that statistically includes me. But a revolution is on and we're all a part of the new guard.
It's always been the responsibility of the minority to navigate the majority's world. We need to take the time to look at what it's like to be a minority, to decolonise the structure of story as we know it. The power of television and film is that we can write stories, which in turn, define us as people and a culture.
Human beings have always understood complex issues through the lens of the personal narrative – it's how we find the deep internal truth, and that is our responsibility as filmmakers.
The personal REALLY is political, especially in these times of urgent identity politics. If you are a woman, if you are of colour, if you are queer, if you are all of the above, if you're none of the above – if you are willing to tell your story and interrogate your position in it, and be vulnerable? That will end up being a revolutionary act.
Find out more about VCA Film & Television Graduate Screenings 2019 – showing in Melbourne until 15 December.
Visit Corrie Chen's website.
Find out more about Film & Television at the University of Melbourne.
Read VCA Digital Archive: multiculturalism in Australia.
Meet the Animation alumna shortlisted for the BAFTA Student Film Awards, Anastasia Dyakova
VCA alumni on the big screen at Palm Springs International ShortFest


Prior to Saturday's lunchtime Sky Bet League One clash against Barnsley, the Blades will be holding its annual 'Remembrance Scroll' as we pay our respects to Unitedites who have passed away during the course of the 2015/16 campaign.
Two seasons ago the club made a commitment for this recognition, to be known as 'Fan's Remembrance Time' to be a feature of our penultimate home fixture every season. This season, again has been a particularly tough one, as week by week we have announced the passing of so many loved ones.
The tribute, lead by Gary Sinclair, will be delivered at approximately 12.15pm, 15 minutes prior to the 12.30pm kick-off with the names of Blades supporters who the Club are aware have passed, being displayed on Bramall Lane's big screen.
Supporters are advised to arrive in good time to offer their applause and pay their respects to fellow fans or loved ones.


Neuronal structures at subcellular resolution in the cortex.
Lesions caused by traumatic brain damage, stroke and functional decline due to aging processes can disrupt the complex cellular network that constitutes the central nervous system, and lead to chronic pathologies, such as dementia, epilepsy and deleterious metabolic perturbations. "How exactly this happens is completely unknown," says Dr. Ali Ertürk, who heads a research group at the Institute for Stroke and Dementia Research at the LMU Medical Center. Ertürk and his team previously developed and have now refined a novel imaging technique that allows them to visualize and monitor these structural alterations in neuronal networks. The new findings appear in the journal Nature Methods.
Because essentially all cell types – including immune cells and tumor cells – can be specifically labeled with the aid of appropriate fluorescent markers or antibodies, the new method can be employed in a broad range of biomedical settings. "Since it allows individual cells to be localized, the method could be used to detect and characterize metastatic tumors at an earlier stage than is now feasible, or to monitor how stem cells behave in the body following a bone-marrow transplant," says Ertürk. Furthermore, the images obtained can be archived in a database and made available to other researchers, which should help reduce unnecessary duplication of studies. Ertürk and his colleagues are already planning to assemble such an online archive.


Daily Mom Military's mission is to provide military spouses and family's with the best resources, information, and most useful organizations that can help make their military life journey a little bit easier. Our partners are committed to this movement as well as committed to providing the best and most reliable resources for military families around the world. To our partners, we thank you for your support in our mission.


text:
var MongoClient = require('mongodb').MongoClient;
var DB_CONN_STR = 'mongodb://localhost:27017/csxiaoyao';

// 插入数据
// 插入两条数据
var insertData = function(db, callback) {
	//连接到表 site
	var collection = db.collection('site');
	//插入数据
	var data = [{"name":"sunshine","url":"www.csxiaoyao.com"},{"name":"csxiaoyao","url":"blog.csxiaoyao.com"}];
	collection.insert(data, function(err, result) {
		if(err){
			console.log('Error:'+ err);
			return;
		}
		callback(result);
	});
}

// 查询数据
// 检索 name 为 "csxiaoyao" 的数据
var selectData = function(db, callback) {
	//连接到表
	var collection = db.collection('site');
	//查询数据
	var whereStr = {"name":'csxiaoyao'};
	collection.find(whereStr).toArray(function(err, result) {
		if(err){
			console.log('Error:'+ err);
			return;
		}
		callback(result);
	});
}

// 更新数据
// 将 name 为 "csxiaoyao" 的 url 改为 "https://www.csxiaoyao.com"
var updateData = function(db, callback) {
	//连接到表
	var collection = db.collection('site');
	//更新数据
	var whereStr = {"name":'csxiaoyao'};
	var updateStr = {$set: { "url" : "https://www.csxiaoyao.com" }};
	collection.update(whereStr, updateStr, function(err, result) {
		if(err){
			console.log('Error:'+ err);
			return;
		}
		callback(result);
	});
}

// 删除数据
// 将 name 为 "csxiaoyao" 的数据删除
var MongoClient = require('mongodb').MongoClient;
var DB_CONN_STR = 'mongodb://localhost:27017/csxiaoyao';
var delData = function(db, callback) {
	//连接到表
	var collection = db.collection('site');
	//删除数据
	var whereStr = {"name":'csxiaoyao'};
	collection.remove(whereStr, function(err, result) {
		if(err){
			console.log('Error:'+ err);
			return;
		}
		callback(result);
	});
}

MongoClient.connect(DB_CONN_STR, function(err, db) {
	console.log("连接成功");
	// // 插入数据
	insertData(db, function(result) {
		console.log(result);
		db.close();
	});
	// 查询数据
	selectData(db, function(result) {
		console.log(result);
		db.close();
	});
	// // 更新数据
	updateData(db, function(result) {
		console.log(result);
		db.close();
	});
	// 删除数据
	delData(db, function(result) {
		console.log(result);
		db.close();
	});
});


text:
San Angelo Now
5 Unavoidable Car Maintenance Costs and How to Manage Them
A Quick Guide For Installing and Replacing Car Speakers
A Review Of Some Of The Most Popular Car Tires Available
Acquiring Mobility Ramps for Cars
13 favs
Air Deflectors - More Than Meets the Eye
The addition of a particular car part to our vehicle is usually designed to improve its performance, but much as we might try to deny it, appearances matter, and we also want our car to look good.
Auto Brakes Service - How To Avoid Getting Ripped Off
Having your car worked on is like going to the doctor. You never know how much it's going to cost, how bad it is and you are not sure if you can completely trust the guy. Finding a mechanic you can trust is like finding a doctor you can trust. You usually wind up going through several before you find the right one.
Auto Repairs To Prepare For Northern Winters
In the north, the winters months are not only hard on the body, but hard on the automobile as well. When the winds start howling and the snow starts falling, cars tend to work harder to warm up and travel to your normal destinations. It is important to keep your auto repair up to date, especially when being stranded could mean the difference between being safe and not being safe.
Better Gas Mileage - It's Easier Than You Think
Car whether it is there to merely serve your transportation needs or whether it is there to fulfill your desire of zooming down the streets in style, all cars have one part that determines its performance. Yes, we all know that is the engine. Although engines differ from car to car based on the make and detailed specification, but it is the "heart" of the car.
Buying New Shocks And Struts - What You Should Know
The first cars that were invented basically had tires that were essentially oversized rubber bands stretched over wooden frames. They didn't even think much about the suspension system either. Today however, suspension technology has improved to the point where riding in your car is just as comfortable as sitting at home on your living room couch and the car handles like it's running on rails.
Car Accident: What to Do After
No one really likes to think about having an accident with their vehicle, but if you drive, you're at risk for ending up in a crash, no matter how minor. Luckily, the majority of car accidents are fairly simple and end up only causing minimal damage to the vehicles, but it's a good idea to know exactly what you should do in the case of a car accident.1. Check if anyone is injured.
Car Advertising: A Low-Cost, Easy Solution For Small Businesses
For the small business owner, advertising can require a large portion of the budget, as well as being quite time consuming. Advertising on your car can create a much easier, lower cost solution to the problems involved with traditional print advertising. One of the first places small businesses go to advertise is the local yellow pages.
Car Seats For Dogs Buying Guide
Many dogs love to ride along in the car with their owners who also enjoy taking their beloved pet for a ride, however, as you may already know if you have ever taken your dog for a ride that many dogs tend to become excited at times while riding in the car and when unrestrained can cause distractions by wandering around the car and interfering with a drivers vision, steering, gear shift, and the p
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text:
Q: Simple JDialog causing Stack Overflow I'm writing a visual application in Netbeans, and came across the need for a dialog box. I decided to copy sections of code from the AboutBox that Netbeans provides automatically, only to discover that both of these are causing errors to be thrown!
I cannot seem to hunt down the source of the errors, although both are caused when the dialogs are closed.
For simplicity, since I believe that both bugs are related, here is my Dialog. I've omitted the initComponents() and main (which is never called) methods for brevity, but the defaultCloseOperation is DISPOSE_ON_CLOSE.
When running I get a stack overflow, which I'll post below.
public class AddRecipe extends javax.swing.JDialog {

/** Creates new form AddRecipe */
public AddRecipe(java.awt.Frame parent, boolean modal) {
    super(parent, modal);
    initComponents();
}

AddRecipe(java.awt.Frame parent) {
    super(parent);
    initComponents();

}

private void cancelButtonMouseClicked(java.awt.event.MouseEvent evt) {
    dispose();
}

// Variables declaration - do not modify
private javax.swing.JLabel Serves;
private javax.swing.JButton acceptButton;
private javax.swing.JButton cancelButton;
private javax.swing.JLabel jLabel1;
private javax.swing.JLabel jLabel2;
private javax.swing.JList jList1;
private javax.swing.JScrollPane jScrollPane1;
private javax.swing.JScrollPane jScrollPane2;
private javax.swing.JTextArea jTextArea1;
private javax.swing.JTextField nameTextField;
private javax.swing.JTextField servesTextField;
private javax.swing.JTextField timeTextField;
// End of variables declaration

}

Stack trace:
Exception in thread "AWT-EventQueue-0" java.lang.StackOverflowError
    at sun.reflect.GeneratedMethodAccessor1.invoke(Unknown Source)
    at sun.reflect.DelegatingMethodAccessorImpl.invoke(DelegatingMethodAccessorImpl.java:43)
    at java.lang.reflect.Method.invoke(Method.java:616)
    at sun.reflect.misc.MethodUtil.invoke(MethodUtil.java:262)
    at java.beans.Statement.invokeInternal(Statement.java:257)
    at java.beans.Statement.access$000(Statement.java:56)
    at java.beans.Statement$2.run(Statement.java:158)
    at java.security.AccessController.doPrivileged(Native Method)
    at java.beans.Statement.invoke(Statement.java:154)
    at java.beans.Expression.getValue(Expression.java:115)
    at java.beans.Encoder.getValue(Encoder.java:105)
    at java.beans.Encoder.get(Encoder.java:225)
    at java.beans.PersistenceDelegate.writeObject(PersistenceDelegate.java:110)
    at java.beans.Encoder.writeObject(Encoder.java:74)
    at java.beans.XMLEncoder.writeObject(XMLEncoder.java:274)
    at java.beans.Encoder.writeExpression(Encoder.java:304)
    at java.beans.XMLEncoder.writeExpression(XMLEncoder.java:389)
    at java.beans.DefaultPersistenceDelegate.doProperty(DefaultPersistenceDelegate.java:229)
    at java.beans.DefaultPersistenceDelegate.initBean(DefaultPersistenceDelegate.java:264)
    at java.beans.DefaultPersistenceDelegate.initialize(DefaultPersistenceDelegate.java:408)
    at java.beans.PersistenceDelegate.writeObject(PersistenceDelegate.java:116)
    at java.beans.Encoder.writeObject(Encoder.java:74)
    at java.beans.XMLEncoder.writeObject(XMLEncoder.java:274)
    at java.beans.Encoder.writeExpression(Encoder.java:304)
    at java.beans.XMLEncoder.writeExpression(XMLEncoder.java:389)
    at java.beans.DefaultPersistenceDelegate.doProperty(DefaultPersistenceDelegate.java:229)
    at java.beans.DefaultPersistenceDelegate.initBean(DefaultPersistenceDelegate.java:264)
    at java.beans.DefaultPersistenceDelegate.initialize(DefaultPersistenceDelegate.java:408)

(Stack trace repeats a few hundred more times).
And for completeness, here is the function that calls the Dialog. Again, this code is modeled off of the default Netbeans behavior, (even though the AboutBox Netbeans provided causes a java.lang.reflect.InvocationTargetException).
private void newRecipeItemActionPerformed(java.awt.event.ActionEvent evt) {
    if (addRecipe == null) {
        JFrame mainFrame = RecipesApp.getApplication().getMainFrame();
        addRecipe = new AddRecipe(mainFrame);
        addRecipe.setLocationRelativeTo(mainFrame);
    }
    RecipesApp.getApplication().show(addRecipe);
}

A: As Ashton K mentions in the comments, this problem can be resolved by moving to the Oracle version of the JDK.  So this problem is an issue with running at least his specific version of OpenJDK.

redpajama_set_name: RedPajamaStackExchange


My friend Jake has been traveling in South America for some time now. Shortly after his trip to Shanghai, China, he came back in time for our wedding (with a day or two to spare), only to them depart a week later for Bolivia.
Jake's been keeping a travel blog over at Travelpod, including both text and photos. Because his Internet access is somewhat limited at times, there are long stretches between posts. But I have to tell you – he's a great writer, and really brings his travels to life.
In February of 2005 I left Bolivia after spending three months volunteering at Inti Wara Yassi, a refuge for injured and orphaned wildlife. During the bus ride out of the jungle I silently vowed that one day I would return to this spectacular country and give more of myself to its people and animals. That day has finally come. Join me and my dog Tus as we head back to Bolivia, this time to La Senda Verde in Coroico, where I will once again spend my days amongst monkeys and wild cats, as well as local school children whom I hope to work with.
Image of an Aymara Noble, September 24th, 2009.
Not only is he descriptive in adding in details to his entries, he's also pretty open and honest at each step of the way. I think he's got a strong voice to his entries, that goes beyond merely documenting this neat site or that interesting spot.
Last time I visited Coroico it was after mountain biking down "The World´s Most Dangerous Road." More people die on other thoroughfares, but per capita this route straight down the Andes is the most fatal on the planet. Coroico is at five thousand feet and the drive is only three hours, so you can imagine how quickly it drops off. They have built a new road since I was last here, which is free from cliffs and is actually paved, so I was happy to learn that I would not be scared out of my wits within my first hour of entering Bolivia. But after the minibus left, the driver informed us that we were taking the old road because it is twenty minutes faster. Wonderful. Let´s legitimately risk our lives to save twenty minutes. The cheapness of life continually surprises me.
I was seated on the left hand side of the bus, which means I was on the cliff side. Many times I looked out and I couldn't see the road, just a several thousand foot drop straight down. Bus and truck bodies litter the ground below, and there are literally hundreds of crosses and plaques commemorating the fallen. The road is simply carved out of the mountain and snakes its way down through streams which only help to erode the mud under our tires. Frankly, it is terrifying.
Tus, hanging out by a waterfall, November 7th, 2009.
If you like any of his writing, also check out the travel blog he made for his Shanghai trip. He's also taken lots of photos, from both journeys.