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Home Local Utah News Multiple agencies, searchers rescue young woman lost in Panguitch Lake area of...
Multiple agencies, searchers rescue young woman lost in Panguitch Lake area of Garfield County
Multiple agencies, hunters and family members worked together in the search and rescue of an 18-year-old woman lost in the Panguitch Lake area of Garfield County on Saturday, Oct. 3, 2020. Photo: Garfield County Sheriff's Office/Facebook
GARFIELD COUNTY, Utah, Oct. 3, 2020 (Gephardt Daily) — The Garfield County Sheriff's Office Search and Rescue team, along with several other agencies and searchers, responded to the Panguitch Lake area Saturday on a report of a missing 18-year-old woman.
A GCSO post on Facebook says the young woman, who has autism, was located at about 7 p.m.
"She was cold, scared and had a hurt ankle but other than that she was rescued and brought back to her family," the post says.
The post credits the many people who came to help, along with the Department of Public Safety Helicopter and crew:
The many people involved in the search were amazing and were never going to give up. Thank you for what you did.
I need to give a huge "Thank You" to the crew of the helicopter. They located the lost girl and hovered over her until searchers could get to her.
The department of Public Safety helicopter and crew have saved 148 people this year. Now they can claim 1 more person saved. They are invaluable in Searches.
Those who showed up to search were Garfield County Sheriff and Deputies, Iron County Deputies, Kane County Deputies, Department of Natural Resources, US Forest Service, elk hunters, campers and family members, K9's and of course the reliable and dedicated Garfield County Search and Rescue.
My apologies if I missed anyone involved.
To view a video of the helicopter and command post, click here.
The Department of Public Safety helicopter crew located a young woman in the Panguitch Lake area of Garfield County and hovered over her until rescuers on the ground could bring her to safety Saturday, Oct. 3, 2020. Photo: Garfield County Sheriff's Office/Facebook
Department of Natural Resources
Garfield County Sheriff's Office
Iron County
Panguitch Lake
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Child abuse includes any type of cruelty or violence against a child. For example, it may include neglect, physical abuse, sexual abuse, or mental abuse. Sometimes prosecutors also charge assault and battery in connection with these cases. There are numerous people who are mandated to report suspected child abuse, including teachers and doctors. However, their suspicions are not always correct. It is important to fight child abuse or aggravated child abuse charges, since they can have significant consequences if you are convicted. If you are accused of child abuse, you should contact Bradenton child abuse lawyer Will Hanlon at Hanlon Law. He takes pride in fighting for the rights of defendants.
You willfully or knowingly abused a child and in doing so caused permanent disfigurement, permanent disability, or great bodily harm to a child.
"Maliciously" under the statute means intentional, wrongful conduct that does not have a legal justification or excuse. A prosecutor can establish that your actions were malicious by showing circumstances from which somebody could conclude that a reasonable parent would not have committed the damaging acts toward his or her child for any legitimate reason, and the primary goal of the acts was to cause the child unjustifiable injury or pain. This is a challenging standard to meet, and a child abuse attorney can help Bradenton residents vigorously fight this type of charge.
There are several offenses that fall under the category of child abuse: child abuse, aggravated child abuse, and neglect. Aggravated child abuse is a first-degree felony. A conviction for aggravated child abuse is punishable by up to 30 years in prison, 30 years of probation, and a $10,000 fine. However, aggravated child abuse can be charged against anyone whose conduct meets the elements of the charge. Being a parent is not a prerequisite for being charged with aggravated child abuse. Sometimes a prosecutor may decide to charge felony battery rather than aggravated child abuse; felony battery is a slightly less serious charge.
Neglect of a child that causes great bodily harm, permanent disfigurement, or permanent disability is a second-degree felony. Neglect refers to a caregiver not providing a minor with the services, supervision, or care needed to maintain the minor's mental and physical health. For example, not taking your child to the doctor or providing him with insufficient food and supervision would be neglect. Neglect can also include a caregiver not making reasonable efforts to protect a child from abuse, exploitation, or neglect by somebody else. Second-degree felonies can be punished by a maximum of 15 years in prison, 15 years on probation, and a $10,000 fine, so retaining a Bradenton child abuse attorney is critical in attempting to avoid these harsh penalties. Neglect is only charged against people with a caregiver relationship to a child.
Knowing or willful abuse that does not cause permanent disfigurement, permanent disability, or great bodily harm is a third-degree felony. Willful or culpable negligence resulting in neglect of a child without causing permanent disfigurement, permanent disability, or great bodily harm can also be punished as a third-degree felony.
While children should not face abuse, there are many situations that are borderline, or in which a vindictive parent or other mandated reporter makes a report for inappropriate reasons. However, while child abuse is a very serious charge, it can sometimes be less serious than other potential charges that a prosecutor could bring. Will Hanlon can work to find an appropriate defense strategy for your case. It is important to retain an attorney as soon as you realize that you are being investigated. Often, the early involvement of an experienced criminal defense attorney can make a huge difference to the outcome of a case. Will Hanlon has aggressively and knowledgeably represented the accused in cases involving child abuse, domestic violence, and similarly fraught situations since 1994. Call Hanlon Law at 941.462.1789 or contact us through our online form.


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crosby composites home
Capabilities & Expertise
Crosby Composites is Britain's leading carbon fibre manufacturer. Established in 1982, we have almost 40 years' experience of innovation in this field.
Paul Field, Managing Director
Paul joined the team in 2009, bringing with him over 13 years of leadership experience in composites manufacturer and motorsports. He has been MD since 2014 and has led the growth of Crosby Composites into the automotive market.
Richard Drury, Senior Project & Design Manager
Richard started at Crosby Composites in 2007 through an industrial placement while completing his MEng in Motorsport Engineering at Brunel University. Starting as a Project Manager, he now leads a team of Project, Design and Technical staff. Richard works closely with Paul supporting Crosby Composites' customers with their design, technical, and planning needs.
Karen Mitchell, HR Director
Karen has been working with Crosby Composites since 2012 in a consulting role and became the HR Director in 2019. Karen brings with her over 20 years of HR experience mainly within the manufacturing arena with a proven ability to develop and implement successful HR strategies that support both the company and the employees.
At the heart of everything we do
Honesty and dependability in our everyday business
It's up to me and it's up to us, we all have to take responsibility for our actions
Always striving to improve in all we do
Never accepting the minimum standard
Driving safety standards
Looking out for ourselves and each other
We are committed to safeguarding the environment through adherence to ISO9001 and IATF16949 standards and ensuring we meet our environmental responsibilities. We comply with rigorous environmental standards, practising effective housekeeping, having accessible cleaning equipment and securely containing materials and recyclables.
Supporting all our customers in achieving industry requirements for sustainability, through providing custom-designed solutions, is important to us.
We recognise the necessity of safeguarding the health and safety of our own employees whilst at work. We provide a safe and comfortable working environment for employees, visitors and the public. Our policy is to manage our activities to avoid causing any unnecessary or unacceptable risks to health and safety. Our policies and associated management systems are reviewed regularly and when there are changes in the business, legislation or industry standards.
Heath & Safety
We endeavour to set the highest practical Health and Safety standards. Our Health and Safety committee meets regularly to discuss Health and Safety topics and concerns, make recommendations and implement and monitor any improvements or initiatives. Health and Safety issues and key measurements are also routinely reviewed and discussed at management meetings.
Ethical Trading and Anti-Slavery
We are committed to Ethical Trading and Anti-Slavery throughout our company and within our supply chain. We believe strongly in ethical principles and good stewardship and are committed to following a set of ethical trading criteria covering non-discriminatory practices, fair wages and working conditions. We adhere to the Modern Slavery Act of 2015 and a programme of action to ensure we do not knowingly support or trade with any business involved in slavery or human trafficking.
Gained IATF 16949 accreditation
Crosby Composites starts production at its 2nd site in Wellingborough
Paul Crosby becomes a Senior Advisor; Paul Field becomes MD
First large batch size order delivered
First automotive visual carbon component produced
Gained ISO 9001 accreditation
Crosby Composites doubles its factory space in Brackley
Start in-house tool design department
Start in-house machine shop
Business is renamed to Crosby Composites and moves to Brackley
First F1 customer and move into carbon fibre
Paul Crosby starts Crosby Composites, then named Crosby GRP, in Silverstone
© 2021 Crosby Composites

redpajama_set_name: RedPajamaCommonCrawl


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Abs and flow cytometry
T cell preparation
CFSE labeling
T cell activation
IL-2 ELISA
Cell transfection and mutagenesis
Characterization of BTLA-specific mAb
Differential effects of BTLA-specific mAb on CD4+ and CD8+ T cell proliferation
PK18-mediated inhibition of T cell activation does not involve regulatory T cells or induction of apoptosis
IL-2 production, but not CD69 induction, is suppressed by BTLA engagement
CD28 costimulation partially overcomes inhibition of T cell activation and proliferation by BTLA engagement
BTLA signal can inhibit proliferation of preactivated T cells
Research Article| November 15 2005
Functional Analysis of B and T Lymphocyte Attenuator Engagement on CD4+ and CD8+ T Cells1
Carsten Krieg;
Carsten Krieg
Department of Immunology, The Scripps Research Institute, La Jolla, CA 92037
Peggy Han;
Peggy Han
Roslynn Stone;
Roslynn Stone
Olivia D. Goularte;
Olivia D. Goularte
Jonathan Kaye
Address correspondence and reprint requests to Dr. Jonathan Kaye, Department of Immunology IMM-8, The Scripps Research Institute, 10550 North Torrey Pines Road, La Jolla, CA 92037. E-mail address: jkaye@scripps.edu
Published: November 15 2005
Copyright © 2005 by The American Association of Immunologists
J Immunol (2005) 175 (10): 6420–6427.
https://doi.org/10.4049/jimmunol.175.10.6420
Carsten Krieg, Peggy Han, Roslynn Stone, Olivia D. Goularte, Jonathan Kaye; Functional Analysis of B and T Lymphocyte Attenuator Engagement on CD4+ and CD8+ T Cells1. J Immunol 15 November 2005; 175 (10): 6420–6427. https://doi.org/10.4049/jimmunol.175.10.6420
T cell activation can be profoundly altered by coinhibitory and costimulatory molecules. B and T lymphocyte attenuator (BTLA) is a recently identified inhibitory Ig superfamily cell surface protein found on lymphocytes and APC. In this study we analyze the effects of an agonistic anti-BTLA mAb, PK18, on TCR-mediated T cell activation. Unlike many other allele-specific anti-BTLA mAb we have generated, PK18 inhibits anti-CD3-mediated CD4+ T cell proliferation. This inhibition is not dependent on regulatory T cells, nor does the Ab induce apoptosis. Inhibition of T cell proliferation correlates with a profound reduction in IL-2 secretion, although this is not the sole cause of the block of cell proliferation. In contrast, PK18 has no effect on induction of the early activation marker CD69. PK18 also significantly inhibits, but does not ablate, IL-2 secretion in the presence of costimulation as well as reduces T cell proliferation under limiting conditions of activation in the presence of costimulation. Similarly, PK18 inhibits Ag-specific T cell responses in culture. Interestingly, PK18 is capable of delivering an inhibitory signal as late as 16 h after the initiation of T cell activation. CD8+ T cells are significantly less sensitive to the inhibitory effects of PK18. Overall, BTLA adds to the growing list of cell surface proteins that are potential targets to down-modulate T cell function.
T cell specificity results from TCR-mediated recognition of specific Ag in conjunction with MHC on APCs (1). However, the functional outcome of this initial TCR engagement is modulated by secondary signals, which can have costimulatory or coinhibitory functions. The group of coinhibitory molecules includes CTLA-4 and programmed cell death-1 interacting with CD80 and CD86 or PD-L1 and PD-L2, respectively (reviewed in Ref.2).
We recently cloned a gene from differentiating thymocytes that encodes a member of the Ig superfamily of cell surface proteins (3). This protein, independently isolated from Th1 cells, has been named B and T lymphocyte attenuator (BTLA)3 (4). BTLA is most highly expressed by B cells, followed by T cells and APC (3), although its expression on macrophages has recently been reported to differ between mouse strains (5). The cytoplasmic tail of BTLA contains ITIM motifs, suggesting an inhibitory function for the protein. Indeed, lymphocytes from BTLA-deficient mice were found to be hyper-responsive to anti-CD3-mediated proliferation (3, 4). In contrast to CTLA-4 and programmed cell death-1, which are induced during T cell activation, BTLA is constitutively expressed on naive T cells and is further up-regulated upon T cell activation (3). Although BTLA was thought originally to be an additional member of the B7 family-recognizing T cell surface proteins, it has recently been demonstrated that herpes virus entry mediator (HVEM), a known ligand for the TNF family member LIGHT (derived from homologous to lymphotoxin, inducible expression), is the ligand for BTLA (6, 7). Like BTLA, HVEM is expressed by T and B cells although the hierarchy of expression on naive cells is reversed from that of BTLA (i.e., T>B) (8). A soluble form of HVEM has been shown to inhibit anti-CD3-induced proliferation of CD4+ T cells, and this effect could be reversed by anti-BTLA Ab (7). In addition, T cells from HVEM-deficient mice are hyperproliferative in response to Con A (9). Together, these data are consistent with a coinhibitory signal mediated by a BTLA-HVEM interaction.
We previously generated a panel of BTLA-specific mAb that preferentially react with the BTLA.2 allelic form of the protein (3). In this study we further characterize the functional properties of one of these Abs, PK18. This Ab has agonistic properties that allow analysis of the effects of BTLA activation and proliferation of naive CD4+ and CD8+ T cells.
C57BL/6J (B6), BALB.K, BTLA-deficient (BTLA−/−), and AND TCR-transgenic (Tg) mice on an H-2b background (10) were used in these studies. All animals were bred at The Scripps Research Institute and maintained under specific pathogen-free conditions. Experiments were conducted in accordance with National Institutes of Health Guidelines for the Care and Use of Animals and with an approved animal protocol from The Scripps Research Institute animal care and use committee.
BTLA-specific rat mAb PK3 (IgMκ), PK18 (IgG1κ), and mouse mAb PJ196 (IgG1κ) were used in these studies. The production of mAb was described previously (3). The following additional Abs were used (all from eBioscience, unless stated otherwise): FITC- or APC-conjugated anti-CD8 (clone 53-6.7), PE-conjugated anti-CD3 (clone 145-2C11), PE- or allophycocyanin-conjugated anti-CD4 (clone RM4-5), biotinylated anti-CD69 (clone 1.2F3), and allophycocyanin-conjugated anti-CD25 (clone PC61.5). Secondary staining reagents included PE-conjugated anti-rat IgM (clone G5-238; BD Pharmingen) and PE-labeled streptavidin (Biomeda). Cell surface staining was performed as described previously (3). In some instances cells were preincubated with anti-CD16/CD32 to block FcRs. For the study of T cell apoptosis, the annexin V-FITC apoptosis detection kit was used according to manufacturer's instructions (BD Pharmingen). Briefly, T cells were washed with ice-cold PBS (Invitrogen Life Technologies), resuspended in annexin V-binding buffer (10 mM HEPES/NaOH (pH 7.4), 140 mM NaCl, and 2.5 mM CaCl2), stained with annexin V-FITC and -propidium iodide, and assayed within 1 h by flow cytometry. As a positive control, T cell apoptosis was induced by dexamethasone (100 nM; Sigma-Aldrich). At least 20,000 viable cells were live-gated on a FACSCalibur or digital LSRII using CellQuest software (BD Biosciences) and were analyzed using FlowJo software (TriStar).
All culture experiments were performed in Click's medium (Irvine) containing 10% heat-inactivated FCS (Invitrogen Life Technologies), penicillin/streptomycin (Invitrogen Life Technologies), 200 mM glutamine, and 50 μM 2-ME (Sigma-Aldrich). In some cultures anti-CD28 (clone 37.51; eBioscience) was used at a concentration of 10 μg/ml, or recombinant murine IL-2 (eBioscience) was added at various concentrations. Cells were cultured under standard conditions at 37°C in 5% CO2. T cells from spleens or lymph nodes were enriched by negative selection using anti-B220 (clone RA3-6B2; eBioscience), anti-Ab class II MHC (clone Y3P) Abs, and anti-rat IgG magnetic beads (Qiagen). For Ag-specific proliferation assays, AND TCR-Tg mice were used as an enriched source of naive CD4+ T cells. For CD4+ T cell enrichments anti-CD8 (clone 53-6.7; eBioscience), Ab was added to the mixture. The purity of the resultant cell populations was >95%. In some experiments, naive CD4+CD25− T cells were purified by sorting on a FACSDiva or FACSAria (BD Biosciences) to a purity of >99%.
Total T cells were washed with PBS, resuspended at a density of 1 × 107 cells/ml in PBS containing 2 μM CFSE (Molecular Probes), and incubated for 5 min at room temperature. Labeling was terminated by addition of FCS to a final concentration of 10%, and cells were washed once in culture medium. The frequency of CD4+ or CD8+ T cell precursors that were activated to divide was calculated as the number of CFSE+ cells in each division/2n, where n is the number of divisions, divided by the input number of cells.
A total of 1 × 106 T cells/ml were cultured in 100 μl or 1 ml in 96- or 24-well, flat-bottom plates, respectively, coated with 1:1 ratios of anti-CD3 (clone 145-2C11; eBioscience) in combination with rat IgG (Jackson ImmunoResearch Laboratories), PK18, or PJ196 at the indicated total Ig concentration. In some experiments, T cells were stimulated with a combination of 50 ng/ml PMA and 500 ng/ml ionomycin (Sigma-Aldrich). For analysis of Ag-specific T cell proliferation, CD4+ T cells were cultured at 5 × 105 cells/ml with 2 × 106 cells/ml irradiated (3300 rad) BALB.K splenocytes in the presence of 100 nM pigeon cytochrome c peptide. For proliferation studies, T cells were cultured in 96-well plates for a total of 72 h and pulsed with 1 μCi/well [3H]thymidine (PerkinElmer) for the last 16 h. Cells were harvested onto glass-fiber filters (Brandel), and [3H]thymidine incorporation was determined. To analyze T cell activation markers, cultures were harvested at various times and were analyzed by flow cytometry. In other experiments, CFSE-labeled total T cells were stimulated for 72 h and then analyzed.
Culture supernatants were assayed for mouse IL-2 using a standard sandwich ELISA (eBioscience). Maxisorb plates (Nunc) were coated with anti-IL-2 mAb (clone JES6-1A12), and detection was performed using the corresponding biotinylated mAb. The plates were developed using streptavidin-HRP and tetramethxylbenzidine as substrate. Plates were read on a plate reader at 450 nm, and data were analyzed using Softmax software (reader and software from Molecular Devices). Samples were performed in duplicate. The assay was standardized with recombinant murine IL-2, and detection ranged from 1 to 1000 pg/ml.
Jurkat T cells were transfected with a construct encoding murine BTLA-yellow fluorescent protein (YFP) fusion protein (base vector pEYFP-N1; BD Clontech) by electroporation and were stained and analyzed by flow cytometry on day 2. PK18 does not stain untransfected Jurkat T cells. Site-specific mutations were made using the GeneTailor Site-Directed Mutagenesis System (Invitrogen Life Technologies) according to the manufacturer's instructions. All mutant constructs were sequence verified (TSRI Center for Nucleic Acids Research).
We previously generated BTLA-specific mAb by immunizing rats or mice with a recombinant soluble form of the protein (3). All Abs generated reacted preferentially with BTLA expressed on cells derived from B6, but not BALB/c, mice, the results of allelic variation in the BTLA external domain (3). We also demonstrated that coimmobilized rat anti-BTLA mAb PK18 is a potent inhibitor of anti-CD3 Ab-induced T cell proliferation (3) (Fig. 1,A). Soluble anti-BTLA Ab PK18 also significantly inhibited proliferation of CD4+ T cells at limiting concentrations of plate bound anti-CD3 Ab (Fig. 1,B). As a control, PK18 had no effect on the proliferation of CD4+ T cells derived from BTLA-deficient mice (Fig. 1,B). The PK18-mediated inhibition of T cell proliferation is not due to altered kinetics of the response (Fig. 1,C). Seven additional mAb (one rat- and six mouse-derived) had no effect on (or, in some experiments, enhanced) T cell proliferation (for example, Ab PJ196 in Fig. 1 A). The inability of other anti-BTLA Abs to inhibit T cell proliferation is not due to poor binding. In fact, PJ196 stains cells brightly even at extremely low concentrations of Ab, whereas PK18 is, somewhat surprisingly, a relatively poor staining reagent (data not shown). This suggested that PK18 and other Abs, such as PJ196, might differ in the respective epitopes on BTLA recognized.
Characterization of anti-BTLA mAb. A, T cells were cultured in plates coated with equal concentrations of anti-CD3 and rat IgG (▪), anti-CD3 and PK18 (□), or anti-CD3 and PJ196 (▴). Concentrations refer to total IgG. Data are expressed as the mean [3H]thymidine incorporation of triplicate cultures (±SD) minus the cpm of T cells cultured alone. B, CD4+ T cells isolated from BTLA−/− mice (top graph) or B6 mice (bottom graph) were stimulated with the indicated concentrations of plate-bound anti-CD3 in the presence of soluble rat IgG (▪) or soluble PK18 (•) at a concentration of 60 μg/ml. Proliferation was determined by [3H]thymidine incorporation, as described in A. C, T cells were stimulated with anti-CD3 and rat IgG (▪) or anti-CD3 and PK18 (□). [3H]Thymidine incorporation was determined at the indicated times as described in A. D, Jurkat T cells were transfected with constructs encoding a fusion of the wt or mutant BTLA.2 extracellular domain and YFP. Cells were stained with PK3, PK18, or PJ196 mAb as indicated and were analyzed by flow cytometry. The histograms shown are gated on a narrow range of YFP expression. Isotype control staining is shown as thin lines. Results are representative of three experiments.
We took advantage of the ability of PK18 and the other mAb to distinguish BTLA.2 (B6-derived) and BTLA.1 (BALB/c-derived) to map residues in the external domain of BTLA that are important for binding. BTLA.2 and BTLA.1 differ in 11 aa in the external domain of the cell surface protein (positions 41, 45, 47, 52, 55, 63, 74, 85, 91, 102, and 143, with the start methionine designated position 1). Murphy et al. (5) reported that residue 74 was an asparagine in both B6- and BALB/c-derived BTLA, whereas other strains had a histidine at this position. Our BALB/c-derived clone also had a histidine at this position, but whether this reflects differences in substrains is not known. A construct encoding a fusion of BTLA.2 and YFP was subjected to site-directed mutagenesis to allow the expression of mutants of BTLA.2-YFP that had individual residues changed to those in BTLA.1 (with the exception of codons encoding residues 45 and 47, which were mutagenized together). Individual BTLA-YFP constructs were transfected into Jurkat cells, the cells from each transfection were aliquoted for staining with various BTLA mAb, and the cells were subjected to two-color FACS analysis.
The cell surface expression of BTLA-YFP was quite variable between individual transfected cells despite similar expression of YFP (data not shown). Whether this reflects tight regulation of cell surface transport of the BTLA protein or a property of this particular fusion protein is not known. Nevertheless, cells were gated for a narrow range of expression of YFP, and then anti-BTLA mAb staining was assessed. All 10 mutant proteins were expressed on the cell surface (i.e., at least one anti-BTLA Ab could bind at near wild-type (wt) levels), but only three had significantly reduced PK18 staining compared with cells expressing wt BTLA-YFP (Fig. 1,D and data not shown). Mutation of residue 41 from BTLA.2 to BTLA.1 (P41E) completely abolished binding of PK18, whereas C85W and T45N/T47K mutations had lesser effects on Ab binding. Binding of PJ196 to mutant 45/47 was also reduced, as was binding to mutant 41, although to a somewhat lesser extent. A more modest reduction in binding of PJ196 to the 85 mutant was also observed. Staining of Ab PK3 to all three mutants is also shown for comparison, with little reduction in staining to mutants 41 and 45/47, and modest reduction in staining to mutant 85 (Fig. 1 D). Together, the data indicate that PK18 and PJ196 have overlapping, but distinct, epitopes that probably account for their distinct biological activities.
To better understand the functional consequences of BTLA engagement on CD4+ and CD8+ T cells, we took advantage of the agonistic ability of the PK18 mAb. T cells were labeled with CFSE and stimulated for 3 days with anti-CD3 in combination with PK18 or control rat IgG. The CFSE profile of total T cells revealed an almost complete block of T cell proliferation, consistent with results obtained by thymidine uptake (Fig. 2,A). However, when CD4+ and CD8+ T cells were individually analyzed in these assays, there was a near complete block of CD4+ T cell proliferation, whereas a significant proportion of CD8+ T cells had divided (Fig. 2,A). From these data we also calculated the precursor frequency of cells that were activated to divide in these cultures (Fig. 2 A). Only 2.2% of CD4+ precursor T cells underwent proliferation when activated by anti-CD3 in the presence of PK18, compared with 33.3% in control cultures. Moreover, the few CD4+ cells that did proliferate underwent a single division after anti-CD3 and PK18 stimulation, compared with up to three divisions in control cultures. In contrast, 57.9 and 28.5% of CD8+ precursors proliferated in the presence of anti-CD3 and rat IgG or PK18, respectively, and some cells underwent three or four divisions even in the presence of PK18.
Differential effects of BTLA engagement on CD4+ and CD8+ T cells. A, CFSE-labeled T cells were cultured in the presence of anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (▪). Plate-bound Abs were used at 20 μg/ml total Ig. On day 3 of culture, cells were stained for CD4 or CD8 and were assayed by flow cytometry. The distributions of precursor cell divisions in anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (▪) were also calculated. One representative experiment of four is displayed. B, T cells were cultured in medium (▦) or were stimulated for various times, as shown, with anti-CD3 (bold lines) or anti-CD3 and anti-CD28 (dotted lines). Thin lines and vertical bars indicate isotype controls. T cells were stained for CD4, CD8, and BTLA, the latter using the PK3 mAb, and were analyzed by flow cytometry. One representative experiment of three is shown.
Both CD4+ and CD8+ T cells express BTLA, although CD4+ T cells have modestly higher cell surface expression (3). In addition, TCR-mediated activation of T cells up-regulates BTLA (3). Thus, the more profound inhibition of proliferation of CD4+ compared with CD8+ T cells could result from differences in expression. To address this, we stained T cells activated by anti-CD3 in the presence or the absence of anti-CD28-mediated costimulation for expression of BTLA (Fig. 2 B). Up-regulation of BTLA was detected as early as day 1 after activation of CD4+ T cells and reached maximal levels by day 3. Costimulation had little effect early, but overall produced an increase in the number of responding cells. A similar pattern was observed for CD8+ T cells, although up-regulation lagged behind that in CD4+ T cells, and the level of expression was ∼2-fold lower. Thus, differential effects of PK18 on the proliferation of CD4+ or CD8+ T cells could, at least in part, result from differential amounts of cell surface protein and differing kinetics of up-regulation of BTLA surface expression. However, the fact that anti-BTLA can inhibit cytokine production before BTLA up-regulation (see below) suggests that this may not be the only factor involved in the differential effects of BTLA engagement on CD4+ and CD8+ T cells.
It was possible that the failure of CD4+ T cells to proliferate in the presence of PK18 was due to the induction of cell death. However, no significant loss of cell number was observed in PK18-treated cultures (data not shown). In addition, no increase in apoptotic cells as assessed by propidium iodide/annexin V staining was observed after activation of CD4+ T cells for 6 h with anti-CD3 in the presence of PK18 compared with that in cells activated in control cultures (Fig. 3 A). Analysis 16 h after stimulation yielded similar results (data not shown).
PK18-mediated inhibition does not involve induction of apoptosis or activation of Treg. A, T cells were activated in the presence or the absence of PK18 as described in Fig. 1F3. Apoptosis was measured after 6 h by propidium iodide/annexin V staining. As controls, apoptosis was also analyzed in the presence of dexamethasone or PMA and ionomycin. Bars represent the mean and SD of three independent experiments. B, Sorted CD4+CD25− T cells were activated by anti-CD3 and rat IgG (▪) or anti-CD3 and PK18 (□), and resulting proliferation was assessed as described in Fig. 1. Results are representative of three experiments.
We have previously reported that BTLA is expressed by CD4+CD25+ T cells, a subset that includes regulatory cells (Tregs) (reviewed in Ref.11). Thus, we also considered the possibility that PK18 was activating Tregs, which then acted to suppress the proliferation of CD4+ T cells. To test this, we purified CD4+CD25− T cells and tested the effect of BTLA engagement on anti-CD3-mediated proliferation as before. As observed for unseparated CD4+ T cells, proliferation of CD4+CD25− T cells was inhibited by PK18 (Fig. 3 B). Thus, the ability of anti-BTLA to inhibit CD4+ T cell proliferation is a direct effect on the CD4+ T cell itself.
Induction of proliferation is a relatively late event after T cell activation. To determine whether PK18 blocked all signals mediated by TCR engagement, we asked whether PK18 prevented the induction of other T cell activation markers. Interestingly, the induction of CD69 expression on both CD4+ and CD8+ T cells was not inhibited by PK18 (Fig. 4). In contrast, BTLA engagement led to inhibition of subsequent IL-2R (CD25) expression on both CD4+ and CD8+ T cells (Fig. 4). However, the decrease in CD25 expression was more pronounced on CD4+ T cells, consistent with the greater sensitivity of this population to PK18-mediated inhibition of proliferation. The increased frequency of CD25− T cells, particularly pronounced among CD8+ T cells, was due to reduced proliferation in the presence of PK18 and, thus, increased frequency of unactivated cells in these cultures. Because IL-2 is necessary for T cell proliferation in vitro as well as for maintenance of IL-2R expression (12), these results suggested that the primary defect in anti-BTLA engagement could be cytokine production. Indeed, anti-CD3- and PK18-stimulated T cells showed a 80-fold reduction in IL-2 secretion compared with anti-CD3- and rat IgG-stimulated T cells (Fig. 5 A).
PK18 inhibits the expression of CD25, but not CD69. T cells were stimulated, as described in Fig. 2, with anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (bold lines). Shown is expression of CD69 or CD25 after 16 or 48 h in culture, respectively, on gated populations of CD4+ or CD8+ T cells. Vertical bars indicate isotype controls. Experiments were repeated three times.
BTLA engagement inhibits IL-2 secretion. A, Supernatants were collected 16 h after stimulation by anti-CD3 in the presence of rat IgG or PK18 (as described in Fig. 2) and were analyzed for IL-2 secretion. B, T cells were stimulated in the presence or the absence of PK18, as described in A, with the addition in some cultures of the indicated concentration of recombinant murine IL-2. Proliferation was assessed as described in Fig. 1. C, T cells were stimulated with anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (bold lines) with the addition of recombinant murine IL-2 (800 pg/ml, high IL-2; 80 pg/ml, low IL-2). On day 2 of culture, cells were harvested, and gated populations of CD4+ and CD8+ T cells were analyzed for the expression of CD25 by flow cytometry. One representative experiment of three is shown.
To determine whether the decrease in IL-2 production was the sole cause of the inhibition of T cell proliferation in these cultures, we performed add-back experiments with exogenous IL-2 (Fig. 5,B). The addition of 80 pg/ml recombinant murine IL-2, comparable to that produced by anti-CD3-activated T cells themselves in these cultures (Fig. 5,A), was not sufficient to rescue the proliferation of T cells in the presence of PK18 (Fig. 5,B), nor did this concentration of IL-2 restore CD25 to wt levels (Fig. 5,C). Thus, the failure to produce IL-2 is not the sole cause of T cell inhibition mediated by PK18 under these conditions. Nevertheless, high concentrations (800 pg/ml) of IL-2 are sufficient to restore nearly 70% of the T cell proliferative response achieved in control cultures (Fig. 5,B). Despite this, high concentrations of IL-2 were not sufficient to return the expression of CD25 to wt levels on CD4+ or CD8+ T cells activated in the presence of PK18, although, again, the effects of PK18 were more pronounced on the CD4+ T cell population (Fig. 5 C).
CD28 costimulation is a strong enhancer of IL-2 production in T cells (13) and thus might be expected to overcome PK18 inhibition. At low concentrations of anti-CD3, PK18 was still inhibitory, even in the presence of CD28 (Fig. 6,A). However, at higher concentrations of anti-CD3, addition of CD28 costimulation restored T cell proliferation to near normal levels even in the presence of PK18 (Fig. 6,A). Similar results were found when tracking individual divisions of CFSE-labeled CD4+ and CD8+ T cells stimulated with saturating amounts of anti-CD3 (Fig. 6,B). Under these conditions, CD28 costimulation was also able to restore CD25 expression on CD4+ and CD8+ T cells activated in the presence of PK18 (Fig. 6,C). As expected, the addition of anti-CD28 increased IL-2 production in these cultures ∼50-fold (Figs. 5,A and 6,D). Thus, the ability of anti-CD28 to overcome PK18 inhibition mirrors that observed after adding high concentrations of exogenous IL-2 (Fig. 5,C). However, even in the presence of costimulation and a high concentration of anti-CD3, the presence of PK18 caused a nearly 13-fold reduction in IL-2 secretion (Fig. 6 D). Thus, CD28-mediated costimulation does not ablate the inhibitory signal initiated by BTLA engagement.
Effects of costimulation on BTLA engagement. A, T cells were activated by anti-CD3 and rat IgG (▴) or anti-CD3 and PK18 (▵) in the presence of anti-CD28, and proliferation was analyzed as described in Fig. 1. B, CFSE-labeled T cells were stimulated as described in Fig. 2 with anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (▪) in the presence of anti-CD28. On day 3, cells were stained for CD4 or CD8 and were analyzed by flow cytometry. C, T cells were cultured with anti-CD3 and rat IgG (▦) or anti-CD3 and PK18 (bold lines) in the presence of anti-CD28. On day 2 of culture, cells were harvested, and gated populations of CD4+ and CD8+ T cells were analyzed for the expression of CD25 by flow cytometry. D, IL-2 secretion was assayed in the supernatants of overnight cultures activated as described in B. E, Proliferation of AND TCR-Tg CD4+ T cells cultured in the presence of BALB.K APCs and specific peptide Ag with the indicated amount of soluble PK18 (□) or control rat IgG (▪) was measured by [3H]thymidine uptake as described in Fig. 1 A. Each graph represents one representative experiment of three.
Given that soluble PK18 could inhibit the proliferation of CD4+ T cells at limiting concentrations of plate-bound anti-CD3 in the presence of costimulation, we also tested the effect of PK18 on Ag-stimulated CD4+ T cells. AND TCR-Tg CD4+ T cells were stimulated in the presence of irradiated BALB.K splenocytes as a source of APCs. We have previously shown that BTLA is expressed on T cells and APC, and that PK18 binds preferentially to B6-derived splenocytes, but not BALB.K splenocytes (3). Thus, in these experiments PK18 would target the T cells and not the APC. Addition of PK18 to these cultures significantly inhibited T cell proliferation in response to peptide Ag (Fig. 6 E).
Our results demonstrated that BTLA engagement at the time of initial TCR activation was a potent inhibitor of aspects of T cell activation. However, the fact that CD69 induction was not affected by PK18 raised the possibility that the inhibitory effect of BTLA engagement might be a relatively late event. Thus, we isolated naive CD4+ T cells from B6 mice, stimulated them overnight with plate-bound anti-CD3, and then transferred the cells to fresh plates coated with additional anti-CD3 in combination with PK18 or rat IgG. Surprisingly, PK18 was able to inhibit CD4+ T cell proliferation even after an initial overnight activation with anti-CD3, similar to the inhibition observed for cells exposed to PK18 for the entire culture period (Fig. 7).
Proliferation in preactivated T cells can be blocked by a secondary BTLA signal. CD4+ T cells isolated from B6 mice were stimulated overnight with a titration of anti-CD3 and transferred to a new plate coated with the indicated concentrations of anti-CD3 and rat IgG (•) or anti-CD3 and PK18 (○). As controls, untransferred CD4+ T cells cultured in the presence of anti-CD3 and rat IgG (▪) or anti-CD3 and PK18 (□) are shown. Data are displayed as the mean [3H]thymidine incorporation of triplicate cultures (±SD) minus the cpm of T cells cultured alone. Two independent experiments gave similar results.
We have generated a panel of mAb specific for BTLA, a recently discovered member of the Ig superfamily of cell surface proteins (4). Only one of these Abs, PK18, was found to act as an agonist for this coinhibitory molecule, thereby inhibiting facets of T cell activation.
We were able to take advantage of the fact that PK18 preferentially binds the allele expressed in B6 mice, but not that in BALB mice (3), to perform initial epitope-mapping studies. Binding of PK18 to BTLA.2, but not BTLA.1, is critically dependent on residue 41, because the P41E substitution completely abolished Ab binding. Other Abs, for example, PK3, bound to the position 41 mutant at wt levels, indicating that the inability of PK18 to bind the mutant was not caused by a lack of surface expression. In addition, the binding of PK3 to mutant 41 indicates that other epitopes on the protein were not grossly distorted. The replacement of the proline at this position with a glutamic acid is likely to alter the local structure of the protein, but might also prevent Ab binding due to the addition of the negative charge.
Of the nine additional BTLA.2 mutants (encompassing 10 aa changes), only two, the substitution at position 85 and the double substitution at positions 45 and 47, had any effect on binding of PK18. Another allele-specific mAb, PJ196, which lacks inhibitory activity, was also affected to varying degrees by substitutions at these same positions. Substitution at 85 had a small effect on binding of PJ196, whereas mutants 41 and 45/47 had significantly greater effects on binding. Overall, however, the rank order of binding of PK18 and PJ196 to these mutants differed (for PK18 mutants, 41<45/47 and 85; for PJ196, 45/47<41≪85), consistent with overlapping, but distinct, epitopes recognized by the two Abs. Given that PJ196 is a better binder to cells than PK18 (data not shown), it seems likely that the dramatic difference in biological activity between these two mAb is due to the relatively subtle differences in the epitopes bound.
These data should be helpful in producing and screening for additional Abs that have agonist function. In an Ig fold, residues 41, 45, and 47 would be predicted to be on a solvent-exposed face of the protein (data not shown). Whether this region of BTLA is also involved in binding to HVEM, and whether there could be allelic differences in that binding, remain to be determined. Interestingly, two BTLA.2-specific Abs generated by Murphy et al. (5) appear to recognize epitopes distinct from that of PK18 or PJ196, although substitution at position 85 (our numbering) does eliminate binding of one of their Abs.
When assessed by uptake of thymidine, PK18 was found to be a potent inhibitor of T cell proliferation. However, single-cell analysis of cell division by CFSE staining revealed that CD4+ T cells were more susceptible to the inhibitory effects of PK18 than were CD8+ T cells. Similarly, PK18 also inhibited CD25 up-regulation on both CD4+ and CD8+ T cells, but the inhibition was again more pronounced on CD4+ T cells. Resting CD4+ T cells express modestly higher levels of BTLA than do CD8+ T cells (3) (Fig. 2 B). Moreover, as shown in this study, BTLA is up-regulated with faster kinetics on activated CD4+ compared with CD8+ T cells. Thus, we cannot rule out that differences in expression may be one factor in the increased susceptibility of CD4+ T cells to PK18-mediated inhibition. Alternatively, CD8+ T cells may be intrinsically less sensitive to the inhibitory effects of PK18, especially during their initial phase of proliferation, which may be IL-2 independent (14). We have seen similar results using a soluble form of HVEM (data not shown); thus, it is unlikely that these results are peculiar to the PK18 mAb.
Soluble PK18 also inhibited the proliferation of CD4+ T cells at limiting concentrations of plate-bound anti-CD3 (Fig. 1 B). This was somewhat surprising, because it was previously reported that BTLA phosphorylation, and thus presumably signaling, requires co-cross-linking with the TCR (4). In these latter experiments, T cell hybridomas overexpressing a Myc-tagged version of BTLA lacking the external domain were treated with anti-Myc and anti-CD3 Abs and cross-linked with a cross-reactive secondary Ab. Whether the lack of the BTLA external domain or the use of hybridoma cells affected the requirement for co-cross-linking in this system is not known. A soluble recombinant form of the BTLA receptor HVEM, however, was also reported to inhibit T cell proliferation induced by limiting concentrations of plate-bound anti-CD3 Ab, similar to the activity of PK18 (15, 16). Thus, PK18 appears to be a good mimic of the BTLA ligand.
We have found that BTLA engagement by PK18 is a potent inhibitor of IL-2 secretion. However, the addition of exogenous IL-2 at a concentration that is normally produced in these cultures was not able to rescue proliferation or CD25 expression on T cells activated in the presence of PK18. Thus, BTLA engagement has additional IL-2-independent inhibitory effects on T cell activation. In contrast to the effects on cytokine secretion, PK18 had no effect on the induction of CD69, indicating that BTLA engagement does not prevent initial activation of the T cell, but modifies the consequence of such activation. That BTLA might function as a response modifier is also supported by kinetic studies of susceptibility to a PK18-mediated inhibitory signal (see below). Although IL-2 is a potent mediator of initial T cell proliferation in vitro, IL-2 may play a role in vivo in subsequent phases of T cell clonal expansion and as a development of effector function in addition to its effects on Tregs (reviewed in Ref.17).
CD28-mediated costimulation lowers the threshold of T cell activation (18), potently enhances IL-2 production (13, 19) and IL-2R expression (20), and promotes progression through the cell cycle (21). At limiting concentrations of anti-CD3, PK18 was inhibitory even in the presence of anti-CD28. CD28 costimulation was able to counteract the inhibitory effects of PK18 on cell division in the presence of high concentrations of anti-CD3. Even under these conditions, however, IL-2 production was significantly inhibited by PK18. These findings are consistent with the ability of APC that coexpress HVEM and CD80 to inhibit the proliferation of TCR-Tg CD4+ T cells in response to low, but not high, concentrations of antigenic peptide (6). Similarly, high concentrations of PK18 can inhibit Ag-specific T cell responses (Fig. 6 D). This inhibitory effect was due to binding of PK18 to the T cell, because we used APC that would not be targeted by PK18. Thus, BTLA engagement under the limiting conditions of costimulation or TCR activation that may occur in vivo may deliver an inhibitory signal. In the presence of anti-CD3, -CD28, and -PK18, the remaining IL-2 produced was probably sufficient to induce T cell proliferation to near control levels, because similar concentrations of exogenously added IL-2 rescued the proliferation of T cells activated in the presence of PK18. It is also possible that the ability of CD28 costimulation to regulate progression through the cell cycle in an IL-2-independent manner (22) plays a role in counteracting the inhibitory effects of BTLA engagement on cell division.
BTLA up-regulation occurs over several days after T cell activation. CD28-mediated costimulation increased the frequency of cells that up-regulated BTLA, but did not alter these kinetics. Because the production of cytokine is a relatively early event, the ability of PK18 to modify this response suggests that BTLA engagement on naive T cells before up-regulation of the cell surface protein can still deliver an inhibitory signal. Nevertheless, up-regulation of BTLA in vivo may be an important factor in making T cells more susceptible to BTLA-mediated inhibition. Interestingly, the BTLA ligand HVEM, also expressed by naive T cells, is down-regulated with similar kinetics as those of BTLA up-regulation (9) (data not shown). This might suggest that a BTLA-HVEM interaction is unlikely to autonomously regulate T cell function.
Because PK18 failed to block CD69 up-regulation, we asked whether BTLA engagement on preactivated CD4+ T cells would still be inhibitory. Surprisingly, cell proliferation was inhibited when CD4+ T cells were exposed to PK18 as late as 16 h after initial culture with anti-CD3. Intravital imaging has recently shown that during the first hours of initial CD4+ T cell activation, serial short-lived interactions with APC bearing cognate peptide are sufficient to up-regulate CD69 (23). Subsequent phases associated with more stable T cell-APC contacts appear to be required for induction of cytokine and subsequent cell division. The fact that inhibitory signals mediated by BTLA can be temporally separated from initial TCR activation raises the possibility that BTLA engagement during this secondary phase of T cell-APC interaction could modify the outcome of the T cell response.
We thank Parinaz Aliahmad for critically reading this manuscript, and Alan Saluk, Cheryl Kim, and Kurt Van Gust for excellent technical assistance with cell sorting.
The authors have no financial conflict of interest.
This work was supported by National Institutes of Health Grant AI31231. This is Manuscript 17365-IMM from The Scripps Research Institute.
Abbreviations used in this paper: BTLA, B and T lymphocyte attenuator; HVEM, herpes virus entry mediator; LIGHT, derived from homologous to lymphotoxin, inducible expression; Tg, transgenic; Treg, regulatory T cell; YFP, yellow fluorescent protein; wt, wild type.
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In Pursuit of the Moon
by Bill Townsend (Author)
Nonfiction Science and technics Technology & engineering Mechanics Aeronautics & astronautics
In 2007, NASA released a request for proposals for the development of the avionics unit?the brains?of a new launch vehicle intended to return humans to the moon for the first time in forty years, before proceeding on to Mars.
This is the story of Ball Aerospace?s pursuit of this opportunity, while competing against four other major aerospace company teams, with almost $1 billion of new work at stake.
After forty-plus years with NASA, the author, having joined Ball Aerospace in Boulder, Colorado, in 2004, took thirty-five top-notch Birkenstock-wearing engineers to Huntsville, Alabama, to establish a new Ball Aerospace office and lead the pursuit of the ARES I Instrument Unit Avionics contract. Being less than a tenth the size of its competitors, Ball Aerospace was a certifiable underdog.
Once there, the Ball Aerospace team worked toward a common goal: pursuing and winning the contract by seeking to overcome the entrenched aerospace contractors who had dominated NASA?s human space flight program for decades.
Explore the inner workings of a real-life industry pursuit and see how a major government agency, NASA, behaves and conducts itself with its contractor base?sometimes in ways you would not expect.
iUniverse


A Lockport man has been charged with attempted murder after police say a woman was shot in the upper body.
Early Saturday morning at shortly after 2 a.m., police said officers were patrolling in the area of Andrews Street and Aberdeen Avenue when they came across a truck with a woman inside who had been shot in the upper body. Police said additional units were arriving on the scene when a man approached officers, threw something between two parked vehicles and then put his hands in the air. Officers took the man into custody and police said a gun was found near the vehicles.
The woman was taken to hospital in critical condition and was later upgraded to stable.
Upon further investigation, police said it is believed the man, who is the owner of the truck, got in an argument with the woman and an acquaintance immediately before the shooting took place.
Police say they have identified the acquaintance and have not identified the person as they are not a suspect in the incident.
Forty-year-old Donald Michael Kargus of Lockport faces a number of charges including attempted murder, discharging a restricted or prohibited firearm with intent, possession of a firearm knowing the serial number has been tampered with, and two counts of pointing a firearm.


We are celebrating an essential moment of this International Meeting of Prayer for Peace organized by the Community of Sant'Egidio. This meeting takes place year after year to keep the spirit of Assisi alive, after John Paul II started it with the representatives of the major world religions.
We gathered in the name of our Lord to pray, to faithfully and insistently ask Him to grant the gift of authentic peace to our world. We are all well aware that by world we mean the whole humanity, all men and women who received from our Lord Jesus the duty to be peacemakers. In this regard, we ask Him to fill our hearts with the gift of His peace. This is the only way for us to become people who sow peace built on love and justice in this world, while it stands unmoved by the horror of concentration camps, like Dachau, still dumbfounded while it commemorates the tenth anniversary of the attack on the Twin Towers, and suffering the severe consequences of the economic crisis.
This passage from the Letter to the Romans reminds us of peace, and offers us the most solid fundament to build peace everywhere in every time. With this letter, the Apostle mentions several problems that were troubling the Christian Community. Some of those problems were related to the daily life of its members. The community of Rome gathered together Christians of very different origins: it is left unsaid that the "strong" were gentiles and the "weak" the Judeo Christians. We can assume the problems were related to Judaic prescriptions, considered by the "weak" as compelling for all Christians, while seen as outstripped in Christ by the "strong".
The passage we listened to today has a rich moral content, evangelically moral, grounded in the Good News preached by Jesus. St Paul's exhortations remind us of the core message of the Sermon on the Mount: one should not return evil for evil, nor seek for his own justice, one should love his enemies and do them good, not be overcome by evil, but overcome evil with good. St Paul develops his thought and expects this behavior from us all, who are united to Him through Baptism, in His death and resurrection. Surrounded by Christ, we must imitate Him, living in the world as He did, always doing everyone good.
The four chapters of exhortations in the Letter to the Romans insist several times on the foundation of true peace, which is the new commandment of fraternal love the Lord has given to us. The Apostle connected this love to the prescriptions of morality. Love summarizes the various essential facets of moral rules in one higher attitude: whoever loves, starting with his inner joyful freedom, is able to give everything to the others, much more than what the prescription itself required. Those who are faithful in Christ, whether they are of gentile or Judaic origin, must be united in love and mutual support. These are the roots of the foundation of peace within our communities and societies.
By touching upon the deepest concerns and desires of the human being, the evangelical message shines today with a new clarity when it proclaims blessed the peacemakers, "for they shall be called sons of God" (Mt 5,9). This beatitude drives us to pursue the duty the Lord gave us all, members of different religions and peoples from around the world, the duty we are fulfilling in these days altogether. We are happy to work for peace, and thus shall be called sons of God.
Peace on earth, which stems from love for friends and enemies, is the image and consequence of Christ's peace, which comes from God our Father. His own incarnate Son, the Prince of Peace, reconciled humankind with God through the cross, and restoring unity to all in one body, He cancelled hatred in His flesh, and having risen with His Resurrection, He effused the Spirit of Love into the hearts of men and women.
Dear brothers and sisters gathered in the name of God, He is here among us; we ask him faithfully for the gift of peace He obtained through His death and resurrection. Jesus told us: "Ask and it will be given". Let us ask, and follow the exhortation of the Apostle, for peace is a gift and a reward. We must beg and work for it, offer it to our brothers and sisters and receive it back from them, as we are going to do in a symbolic gesture here, all of us believers in Christ, and on the square with all our other brothers and sisters.
It will be an opportunity to bear witness once again to how religions further peace around the world, like an expression of God's love for all humankind.


While photos capture a moment in time, they don't give the subjects an opportunity to speak and express their feelings. They also don't allow for captioning by those taking in the picture. What if you could make your pictures speak? What if you could find out what others are thinking about your wedding?
When ordering custom photo framing, individuals choose the right matting or border to surround their pictures. While there are lots of prints, patters, and textures to choose from, the matting also offers a great space for the pictures to have a voice. Here are three ideas for taking photo matting and turning it into something special.
Ask any married couple and you will find that a guest book is often packed away after the event, never to be seen again. Instead of relying on a book that isn't going to be opened, consider using photo matting as a way for wedding guests to sign in.
At the time of the signing, a bride and groom can place an engagement picture nearby. Every person in attendance can leave a note of well wishes or a simple signature, depending on the size of the matting. Once the wedding pictures are complete, the engagement photo can be replaced by one of the bride and groom on their special day.
Brides and grooms often choose to show their appreciation to parents or other loved ones before, during, or after the wedding. An engagement picture will work well if the framed picture is going to be presented to parents before or during the wedding. If the gift will be given after the wedding, the couple can arrange to have a wedding photo added.
Before turning the photo in for custom picture framing, the couple can write out a note that expresses their gratitude to the important people in their lives. For the recipients, the framed picture becomes more than just a gift. It becomes a treasured keepsake they will have for years to come.
Bridesmaids and groomsmen are usually presented with a gift to show the bride and groom's appreciation for their participation. While there are lots of traditional ideas for wedding party gifts, it is possible to create something personalized for each individual.
Choose a photo that best represents the person's relationship to the bride or the groom. There are lots of different photo ideas to choose from. Find a photo of the bride and her maid of honor during their school years. Track down a photo of the groom and his best man on vacation or out on the town.
In addition to presenting the wedding party with a memory, the bride and groom have the opportunity to express their appreciation by writing on the photo matting. It doesn't matter whether the message is simply, "thank you," or entire paragraphs are written around the matting. The personalization alone makes this a special gift.
Weddings offer a great opportunity for individuals to share an important moment with their closest friends and family. Take advantage of the experience and use it to communication something special with the help of places like Hoosier Highlander. Transform photo matting into a makeshift guest book or use it as a way to communicate to parents or members of the wedding party. This is your unique chance to make your photos speak.


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What the critics are saying about the 'Downton Abbey' movie: 'Ridiculous but fun'
Yahoo Movies UK 10 September 2019
Downton Abbey (Credit: Focus Features)
The Downton Abbey movie premiered in London last night, and now critics have had their say on how they feel to be back in the embrace of the Crawley family.
In short, it appears that while it might not sweep the boards during the forthcoming awards season, it's still a treat for fans.
It finds the denizens of Downton thrown into panic following news of a visit from King George V and Queen Mary.
Read more: All the gossip from our Downton Abbey set visit
The Guardian's Peter Bradshaw rather sums it up in a three-star notice, describing it as feeling like 'an intensely glucose Christmas special'.
"There are some films that you really have to see on the big screen," he adds. "Not this one, though. To get the full, authentic experience, you'll need to see it on the small screen, on 27 December, with quart of eggnog inside you and enough Quality Street to trigger a diabetic coma. It is at all times ridiculous – but, I have to admit, quite enjoyable."
(Credit: Focus Features)
Writes Caryn James for the BBC: "The film is delightful fun, even though the plot is obvious almost to the point of stupidity, and there are few surprises for the well-known characters, some of whom are handed unnecessary subplots to give everyone some screen time."
Adds Brian Viner in the Daily Mail: "There's absolutely no point seeing it if you don't already know who all these people are. If you do, how can you possibly resist?"
Empire calls it 'gentle, unchallenging drama for people who already know they like it, this is a nostalgic and rosy depiction of an England that was, surely, never so innocent'.
Thus far, it's draped in an 85 percent 'fresh' rating on Rotten Tomatoes, despite the vast majority of the reviews being a mediocre three stars.
But not everyone is bathing in the glow of nostalgia for the ITV series, which we all thought had wrapped up its affairs in 2015.
The Daily Telegraph questions 'does this really deserve to be in cinemas?'
"Ridiculously, it comes billed on buses as 'the cinematic event of the year' – true enough if it's the only film a person might opt to see," it adds.
The New York Post offers 'two thumbs Downton', branding it 'a lifeless blob of brass fixtures and aerial shots'.
Read more: Downton Abbey trailer drops and there's a Royal arrival
Variety is similarly unimpressed, adding: "It's reminiscent... of the 2008 film Sex and the City, which was strangely, lumpily paced and told a basically unnecessary story, but which was still true enough to its characters that it was embraced by fans.
"This journey to the past may end up feeling ultimately less nostalgic than backward-looking."
Whether the movie will draw fans of the series to the cinemas in any significant numbers remains to be seen, but it lands nationwide on September 13.
#maggie-smith
#hugh-bonneville
#downton-abbey
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text:
Sambizanga é um dos distritos que constituem a área urbana da cidade de Luanda, na província do mesmo nome, em Angola.

Sambizanga tem 14,5 km² e cerca de 244 mil habitantes. Limita-se a oeste com o Oceano Atlântico, a norte com o município do Cacuaco, a leste o município do Cazenga e a sul com os distritos da Ingombota e Rangel.

É subdividido nos bairros de Malanga, Sambizanga e Bairro Operário.

Ver também
 Comunas de Angola

Municípios extintos de Angola
Distritos do município de Luanda

redpajama_set_name: RedPajamaWikipedia


text:
Job Directory Microsoft Senior Program Manager
Senior Program Manager Microsoft
Microsoft is a technology company that develops and supports software, services, and devices.
Companies like Microsoft
Artificial Intelligence is a disruptive technology in the digital transformation of business. In Office, we are infusing AI and machine learning across the entire product suite to help our customers both at home and in the enterprise meet their goals more quickly and effectively. And all our applications and services will increase in their ability to anticipate user intention, automate basic actions, making Office more personalized than it is today. All these experiences become possible when we take a more platform-centric approach to Intelligence.
Over the last two years, Office has been powering scenarios through machine learning for a few years across areas like Search, Entity extraction, Designer, Assistive recommendations, and NL understanding to name a few. The scale and diversity of these scenarios requires large volumes of labeled training data (i.e., for supervised learning) to build AI models. Office data probably represents the world's largest productivity dataset annotated with usage signals from hundreds of millions of users. Imagine taking an arbitrary dataset and training an ML model by harnessing the power of distributed compute in the cloud. Imagine taking hundreds of millions of App usage streams from telemetry and training a DNN model using GPUs in Azure. And imagine the power of a end user-facing scenario that deploys these models seamlessly in the cloud or in the client to power recommendations, collect user labels and kickstart the flywheel of feedback and refinement! And to top it all, AI recommendations are personalized and made contextually relevant to each user. If this excites you, come chat with us!
The Office Language and Intelligence team is bringing the best technological investments from across Microsoft's Cloud AI Platform team, and make it relevant to every Office developer and machine learning scientist enabling them to light-up cutting edge AI in the product. We are looking for strong technical and highly driven program managers who can work and thrive in ambiguity, and hit the ground running to put together these systems of scale. Machine learning knowledge and experience is a huge plus. We work with several teams across the company such as Cloud + AI Platform, Substrate, Windows, and MSR to deliver best in class AI infrastructure for the scale of Office. In this role you will be having a big impact quickly and will be able to have your work cut across a variety of scenarios powering Office client and web apps. You will be part of a rare team that combines cutting-edge industry advancements with engineering excellence in an effort to enable Office shine in the next frontier of AI.
* Demonstrated customer focused innovation
* Strong problem solving skills and cross-group collaboration skills
* Minimum 5 years of Program or Product Management experience
* Strong communication skills and interpersonal awareness to work collaboratively with a diverse set of people across groups and organizational levels
* BA/BS degree in Engineering, Computer Science, or related discipline
Microsoft is an equal opportunity employer. All qualified applicants will receive consideration for employment without regard to age, ancestry, color, family or medical care leave, gender identity or expression, genetic information, marital status, medical condition, national origin, physical or mental disability, political affiliation, protected veteran status, race, religion, sex (including pregnancy), sexual orientation, or any other characteristic protected by applicable laws, regulations and ordinances. We also consider qualified applicants regardless of criminal histories, consistent with legal requirements. If you need assistance and/or a reasonable accommodation due to a disability during the application or the recruiting process, please send a request via the Accommodation request form.
Benefits/perks listed below may vary depending on the nature of your employment with Microsoft and the country where you work.
* Drive the charter of the team to ensure we land a holistic story in Office for the AI platform
* Understand research and industry trends and the AI landscape
* Advocate for Office developers and machine learning scientists to partners across the company
* Set and own the goals; define metrics and key performance indicators of success
* Driving execution, dependency management, and progress reporting
* Rapidly prototype and innovate on our existing tooling
* Build relationships with teams across the Cloud AI Platform and AI&R
More Jobs At Microsoft
Sr. Capability Delivery Manager
Principal Software Engineer, Backend
Microsoft Corporation One Microsoft Way


text:
The eldritch horror of capitalism
Recently I've been listening to Old Gods of Appalachia, a Lovecraft-inspired horror podcast set in my mountains. It's been a lot of fun to listen to, not least because this story is situated within the deep history of Appalachia — of labor unions, witching, and yes, the occasional spot of racism. It doesn't flinch from Appalachia's darkness, but it doesn't make sport of it either the way some big titles set here do. The people in this story are people, neither good nor bad (or sometimes a bit of both). But ultimately, the mines, the company, and to a lesser extent the church, become the embodiment of something too dark for our world.
This theme within the podcast got me to thinking of two other titles I've read in the past few years. The first is the well-known author N.K. Jemisin's The City We Became, a novel set in New York City that posits homogenizing capitalist entities such as Starbucks as arms of R'hley. Jemisin's work conflates whiteness with an eldritch horror, or at least a susceptibility to its influence, which is notable since the destruction of the town of Barlo, KY, in Old Gods is in part triggered by burying several Black scabs in an unmarked grave as burnt offerings to eldritch forces. But there is also a subtext here about money and power, as one of the key plot points in The City We Became involves the cooptation of a nonprofit art gallery via generous donations to the board. The forces of capitalism are definitely at work in this tale, and Jemisin sees them as uncaring, homogenizing, and fundamentally destructive. While the economic systems have certainly mutated, the way they are used to oppress is frighteningly familiar.
The other title that I found myself reflecting on while listening is not explicitly Lovecraftian, but shares some similarities so I thought it worth mentioning. Hunger Makes the Wolf is by Alex Wells, and tells a Firefly-esque story about a company planet. That company planet reminds me deeply of the company towns my ancestors lived in (my parents still live in the company shotgun house erected for my great-grandparents when they came to work the local mine, though we've added onto it a good deal). This work imagines a future where the forces of capitalism have once more overrun the ability for working people to survive, much less self-determine, aided by the isolation inherent in intergalactic travel. And of course those capitalist company interests have weaponized (or been weaponized by) something not quite of this dimension.
There's probably a reason these titles have been so enjoyable for me. Each is looking critically at different aspects of our global economy, and each is set in different time periods. But it seems clear that our past, our present, and our future are all defined by the concentration of power in the hands of the few, the pushing down of people based off their income and the color of their skin, and what solidarity we can provide for our neighbors in the face of such forces.
capitalism, fantasy, horror, hunger makes the wolf, labor unions, old gods of appalachia, science fiction, the city born great, the city we became
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New Members' Ministry — Christ Church Inc.
The vision of this ministry is to teach new members the basic foundations of Christianity, aide new members in building their relationship in Jesus Christ, and in building their relationship with Christ Church.
This ministry works by partnering each new member of Christ Church with a member of the New Member's Ministry for prayer and guidance. This partnership usually lasts a minimum of 3 months.
New members will also learn scriptures by receiving a different memory verse for each of the above lessons and create a plan to read the four gospels over a course of four months.
The New Members ministry is a go-to source for new members that may have questions about the church, or need items like directions to frequently visited churches. This ministry can provide information on recommended books, or reading materials and CDs to help with study and development of their relationship with Christ.


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In Nimble AMS, each product is set up with a default price and additional pricing details can be configured through price classes and special prices.
When you create a special price, you select one or more price classes to receive the price you set. A specific product is also tied to the special price, allowing you to set a price class to different products.
A special price for registration products allows for early, regular, and late pricing. The event determines date for each cut off by setting an Early Registration Cut Off Date and a Regular Registration Cut Off Date.
In Nimble AMS, you use products to represent the goods or services your association offers to your constituents. Your products are set up using product types which define what attributes the products have. For example, an event session product has a start and end date, but a merchandise product does not. Your products are purchased by your constituent using orders.
In Nimble AMS, constituents may receive a reduced price based on their involvement with your association or other attributes. A price class is a group of your constituents who should receive the same price. For example, members may pay a lower price for merchandise than non-members.
You can create a product with special prices, so that you can assign different prices by price class. When a constituent purchases a product, the constituent's price class is used to determine what price is charged.
Your constituents may be part of a price class in which they receive special pricing automatically. You can setup special pricing for different products in Staff View.
When a constituent logs into Community Hub, they will see either the retail price or special price of a product if they meet the requirements of the price class.
All products are set up with a single default price, or the List Price. When an order is processed in Staff View, the order processor displays special pricing based on the account's price class. If an account does not qualify for a special price, the list price is displayed.
Staff have the ability to set or change a special price in Staff View. When a special price is tied to a price class, the price will be driven from this. If an account does not meet the requirements of any price class, they will receive a default price of the product instead of the special price.
Staff may also change the price class on an order in the order process. In doing so, different special prices will display.
Both Community Hub guests and non members can only view retail prices for merchandise products, and must be logged in prior to adding a merchandise product to their cart. Logged in member users can view the retail and member price for merchandise products.


Congrats on finishing your undergrad. Getting a university degree isn't easy and is probably harder now than before with the pressures of part-time work, tenuous housing and future massive HELP debts.
Being smart – you're likely aware of the option to study for a PhD. Maybe your honours supervisor has approached you about doing one, or you've chatted with tutors, demonstrators and academics in your department. If you graduate with first class honours, many universities around Australia will guarantee you a scholarship. A PhD is worth considering – pushing the boundaries of human knowledge is fun. You'd develop serious skills in problem solving and you'd get to call yourself 'Dr'.
But! you should go forward with eyes open and with full awareness of options ahead.
I'm writing this letter to you as a young(ish) academic beginning to supervise PhD students for the first time. It's ace – PhD students for academics are like free worker bees; it's always in our interest to have more. Academics want to dish out scholarships like banks do credit cards. But I'm not sure that potential PhD students in Australia are fully informed of the prospects ahead of them.
This letter aims to address this imbalance.
In 1948 there were no PhDs awarded in Australia. In 1949 there were three. Now there are 7000 each year. And that's only Australia! The USA is graduating 49,000 per year, China more than 50,000, and India aims for 20,000 by 2020.
A PhD degree isn't as exceptional a qualification it once was, and I choose the word 'exceptional' carefully. I grew up reading the childhood classic "Little House on the Prairie" by Laura Ingalls Wilder. In it, Laura grows up to complete Year 7 and become a school teacher herself, something celebrated by the Ingalls family – teaching was a guarantee of secure, meaningful, well paid employment. That was in 1800: today around 7% of the world has an undergraduate university degree! Which is great! But it means that higher levels of education aren't an exception (which is a fantastic thing for the world).
A consequence is that university qualifications are now often a pre-requisite for jobs across many sectors. Perhaps one day post-graduate qualifications will be a pre-requisite for a job across many sectors.
A PhD (usually) takes between 3-4 years, often during some of the most energetic years of life and often on a lowish wage. There will always be a pressure for higher qualification, but at some point the marketplace will start selecting for 4 years of working experience instead of a highly-specialised higher degree by research (if it isn't already doing so).
To an honours student considering a PhD, one potential career path is academia. It's worth having a look at the history of academic employment in Australia.
Undergrad student numbers have doubled since 1989, PhD graduations are growing exponentially (quadrupled since 1989), but the number of full-time academic staff employed in Australia has increased by only 20%. A study of higher education outcomes in the UK found that 0.45% (that is, less than one in 200) of PhD graduates become a full-time professor in a university.
If you're doing a PhD for the money then that doesn't always add up either. The USA Bureau of Labor Statistics found those with a PhD earn on average slightly less than those with a Professional Degree. It's still on average more than a Bachelor, Mastes, or no degree, but the cashola isn't massive. And if you follow the academic route, you're likelygoing to be on short term contracts until you're in your mid-30s (if you're lucky).
There are other avenues of employment with a PhD – there are great initiatives like Teach for Australia, and rewarding opportunities in government and communications and consulting and industry, and of course plenty of opportunities in other countries.
You've got to follow your passions. There's no joy in life being a difference-engine living according to probabilities. And Fortune really does favour the brave. If research is what makes you tick, then there's no other option but to follow it. A PhD can be fun and exciting and interesting and fulfilling. But it's worth knowing the stats and it's worth knowing the options.
Authors note: The impetus for this letter was me commencing supervision for the first time of a new PhD student. Whilst the research we do during their PhD is obviously going to win us the Nobel Prize, I'm not sure how fully aware they are of the options available to the average PhD student upon graduating. And I reckon they should be.
I wanted to write it now while I'm still in academia though not yet tenured. This letter is of course influenced by my own age/stage, but not entirely… I feel that this advice doesn't make it out there from academia. We're either busy trying to recruit students, or writing grants, or perhaps feel like we'd be betraying the academic cause to highlight post-PhD issues. There's been fruitful discussion in the past about 'Whether Australia is producing too many PhDs', and 'What PhD graduates should expect', and the 'The uncertainty of a career in research', but I don't know if anyone is out there telling honours students about it. This letter is aimed at them.
Barro and Lee, "A new data set of educational attainment in the world, 1950-2010", National Bureau of Economic Research, Working Paper Series, Cambridge MA, 2010.
Triple bottom line academia: communicating in the face of research cuts.
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Next thing he knows was "Eh reach already ah?"
And so we started taking picture and Calvin got harassed by the aunties selling postcards. He bought some pretty cheap postcards from the lady.
We drove down a little bit where all the tourist was and took a few more shots before we left. It was cold there but everybody keep bugging you to buy something didn't make it pleasant at all. I love our pictures tho, its damn nice!
Next we go drink Coffee that came out of a musangs poop!


Jamyang Khyentsé Wangpo, to you I pray!
These verses of advice for renunicant meditators (spong ba bsam gtan pa) cover the entire Buddhist path, from instructions on following a spiritual friend through to the most advanced generation and perfection phase practices.
Alak Zenkar summarizes the remarkable life and liberation of Jamyang Khyentse Wangpo (1820–1892), showing how he mastered the so-called 'eight great chariots of the practice lineage' (sgrub brgyud shing rta chen po brgyad) and received the seven special transmissions or 'descents' (bka' babs bdun). Zenkar Rinpoche also briefly summarizes Khyentse Wangpo's collected writings, which are divided into nine main categories.
A brief text summarizing the five great logical arguments of the Madhyamaka, or Middle Way: 1) the investigation of the cause: the Diamond Splinters; 2) the investigation of the result: refuting existent or non-existent effects; 3) the investigation of both: refuting the four permutations of arising; 4) the investigation of essential identity: 'neither one nor many'; and 5) the logical argument of Great Interdependence.
A clear and concise commentary on the words of the most famous of praises to Mañjuśrī, Glorious Wisdom's Excellent Qualities (dPal ye shes yon tan bzang po), attributed to the Indian master Ācārya Vajrāyudha.
A simple guide to the practice known as Narak Kong Shak, or The Confession and Fulfilment that Empties the Hells from their Very Depths.
Jamyang Khyentse Wangpo recounts the origin of the teachings of Siṃhamukhā and how they have been subsequently passed down to him. He closely follows the story associated with the lineage of Bari Lotsawa (ba ri lugs).
Supplementary verses to be added to the prayer to the lineage of Parting from the Four Attachments composed by Ngorchen Kunga Zangpo (1382–1456).
A non-sectarian (ris med) prayer to 25 of the most important figures in Tibetan Buddhism, five for each of the five major traditions: Nyingma, Sakya, Kagyü, Kadam and Geluk.
A brief prayer to the lineage of Zabtik Drolchok, the practice of Green Tārā which was revealed as a mind terma by Chokgyur Dechen Lingpa.
Jamyang Khyentse composed this prayer for perfecting the Seven Points of Mind Training (blo sbyong don bdun ma) when he was in the presence of the famous Atiśa statue at the Tārā Temple in Nyethang (snye thang). The section headings were added by Dilgo Khyentse Rinpoche.
This experiential song (nyams dbyangs) based on 'Parting from the Four Attachments' (zhen pa bzhi bral) was composed in the cave of Rangjung Dorje, where Mañjuśrī is said to have delivered the original teaching to the Sakya patriarch Sachen Kunga Nyingpo (1092–1158).
This guide to the stages of visualization for the Longchen Nyingtik preliminary practices (sngon 'gro) is, as Khyentse Wangpo himself puts it, "brief, clear and essential." Some of its instructions differ slightly from those given by Patrul Rinpoche, so that it represents a distinct commentarial tradition.
This is a version of A Profound Concentration of Nectar, into which the root text of the Longchen Nyingtik preliminaries has been inserted. The text also includes several prayers that were not included in Jikmé Trinlé Özer's original version.
Sometimes referred to as the "short Longchen Nyingtik Ngöndro", this popular preliminary (sngon 'gro) liturgy by Jamyang Khyentse is remarkably concise.
This four-line prayer is sometimes used for the accumulation of maṇḍala offerings in the Longchen Nyingtik tradition. Khenpo Ngawang Palzang, for example, recommends accumulating this prayer seventy thousand times (following thirty thousand recitations of the three-kāya maṇḍala from the Longchen Nyingtik).
Jamyang Khyentse wrote this twelve-line prayer to Patrul Rinpoche during the festival of Chökhor Düchen in 1860. The text identifies Patrul as an emanation of Śāntideva and the early Dzogchen adept Aro Yeshe Jungne, and praises his qualities of renunciation, bodhicitta and wisdom.
Khyentse Wangpo composed this five-verse supplication prayer to Thangtong Gyalpo at the request of the Eighth Sangye Nyentrul Rinpoche by combining and supplementing previous prayers.
A simple four-line prayer to Buddha Śākyamuni and a retinue of the sixteen elders.
This popular liturgy for saving the lives of animals includes practices of taking refuge and generating bodhicitta, as well as the recitation of mantras and dhārāṇīs, visualization, and prayers of auspiciousness, dedication and aspiration.
For this 'means of recitation' (bklag thabs), which provides additional prayers and practices to be said before and after the root text of Chanting the Names of Mañjuśrī (Mañjuśrī-nāma-saṃgīti; 'jam dpal mtshan brjod), Khyentse Wangpo relied upon and adapted the writings of the great Sakya patriarchs Jetsün Drakpa Gyaltsen and Sakya Paṇḍita.
This prayer beautifully summarizes the Padma Kathang (The Chronicles of Padma), one of the most famous and influential of Guru Padmasambhava's many biographies. It reveals how Guru Rinpoche manifests in an infinite variety of forms in order to protect and spread the Buddhadharma.
This brief prayer to Guru Padmākara, which Khyentse Wangpo says came to him spontaneously, includes requests to dispel all obstacles and fulfil all wishes.
This poetic prayer relates the Lotus Light (padma 'od) pure realm of Guru Padmasambhava to the four visions of Dzogchen practice and contains the aspiration that we may all be reborn there, to advance through the four stages of a vidyādhara and swiftly reach the level of the Lake-born Guru himself.
These verses to be be recited before and after prayers to Guru Padmasambhava, especially the famous Prayer in Seven Chapters (le'u bdun ma), include the practices of taking refuge and arousing bodhicitta, a seven-branch offering, the generation and dissolution of a visualisation, and the dedication of merit.
This is a complete set of practices, including taking refuge, generating bodhicitta, visualization, invocation, seven-branch offering, heartfelt prayer—addressed to Guru Padmasambhava, his various manifestations and twenty-five disciples—mantra recitation, and dissolution.
Written in 1850, this is a prayer to the guru, who is understood to be inseparable from Guru Padmasambhava and the true nature of mind, and an aspiration to be reborn in the Lotus Light pure realm, if not already liberated during this life or when clear light dawns at the moment of death.
A prayer to the Precious Guru of Oḍḍiyāna as the embodiment of all sources of refuge (skyabs gnas kun 'dus) in order to avert all forms of obstacle and hindrance, on the outer, inner and innermost (or 'secret') levels.
The longer of two ritual texts by Jamyang Khyentse Wangpo featuring the sixteen arhats, both of which appear in recent editions of the extensive Nyingma Kama (bka' ma shin tu rgyas pa).
The shorter of two ritual texts by Jamyang Khyentse Wangpo featuring the sixteen arhats included in the extensive Nyingma Kama and elaborating on the famous text attributed to Śākyaśrībhadra.
This short Tārā feast-offering was composed for practitioners who wish to offer a simple gaṇacakra feast within a Tārā sādhana, such as the Zabtik Drolchok.
Jamyang Khyentse Wangpo composed this short aspiration prayer to be recited during the gaṇacakra. The prayer invokes the goal of the gaṇacakra, a six-fold satisfaction (tshim pa drug) of those assembled, i.e., the deities, teacher and vajra-brothers and sisters. Khyentse Wangpo dedicates one verse to each of these six satisfactions and concludes the prayer with an additional seventh verse of dedication.