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Document Management for Small Business
By Pam Baker | Posted December 13, 2013
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If you're confused over what document management really means, don't worry, you're not alone. It's a very broad category that refers to converting paper files to digital form, which makes all files searchable by type and keywords. It also lets you sync files when you make changes to a document and makes it easy for authorized employees to access documents at any time and on any device.
A document management system (DMS) can refer to any technologies needed to do any or all of these parts. Therefore, you must be careful in comparing and selecting products because the "document management solution" label doesn't tell you everything you need to know.
How to Prepare for Document Management
The first step in document management is to decide which documents you need to manage. And, no, that doesn't mean you should start by scanning all that paper bulging from your file cabinets or stacked to the ceiling of your storage container. Instead, proceed in manageable and less overwhelming chunks.
"Managing pools of documents that are categorized by the functional area of the business is a great use," says Joe Bachana, president of NYC-based DPCI, a content technologies solutions provider. "Also managing the version control of certain 'living' documents is a terrific use. Scanning records is helpful but sometimes onerous for administrative staff, so have a good business reason for doing so."
In other words, pick a category of documents that is most critical to your company, and start digitizing those. If they're already in digital form, look for ways to make them more manageable and useful. Also, remember that not all of your documents are on paper. Email is another source of documents, and so are files that you store in the cloud-based services such as Dropbox, Salesforce, or Office 365.
Once you know which documents you want to manage—and where all the documents reside—choosing a document management solution to handle it all becomes infinitely easier.
What to Look For In a Document Management System
Jeffrey Segarra, senior director of imaging product management at Nuance Communications, offered this checklist to help determine what you need from a document management system. Consider these 10 questions before you buy:
Do you need mobile or cloud access to these documents?
Consider your level of hardware/software expertise—do you have in-house capability or do you need a trusted IT vendor to handle it all?
Do you want to spend money upfront for a total solution or would you prefer to pay over time?
Do you need desktop tools such as scanning, PDF editing or publishing (PDF/A)?
Do you have security protocols that you must follow—HIPAA, for example?
Is full text search important or is simple folder or file browsing acceptable?
Do you need to connect to a database, customer relations management (CRM), or other sources for data look-up?
Do you need a completely internal system for employees only or do customers and/or suppliers need to participate too?
Is collaboration (real time document sharing, collective editing, change tracking and management) an important component to the system?
Do you require industry-specific capabilities—manufacturing, legal, healthcare, or insurance, for examples? Many specialty DMS system target specific industries.
"As you answer your DMS strategy questions, categorize your requirements into 'must have,' 'nice to have' and 'unnecessary,'" said Segarra. "Assess features that fit into the 'nice to have' category to determine whether the offering can demonstrate clear time savings and/or productivity gains to the business. That will move those items into the 'must have' category."
But there are a few more considerations worthy of adding to your checklist.
"Several other features are also crucial when choosing a cloud document management solution," says Aaron Weiss, director of Marketing for the HP LaserJet and Enterprise Solutions Business. "Solutions should offer encrypted data transfer, encrypted data at rest, and password protection. These features will help SMBs keep documents stored securely."
Weiss went on to say list more features that small business may want to consider, especially if they're looking to cloud-based systems. "Some cloud document management solutions offer optical character recognition optical character recognition (OCR) technology, document indexing, search engine capabilities and full audit trail, which allows employees and SMB owners to see who's accessed and modified a document from creation to deletion."


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Oda Krog (11. jun 1860 – 19. oktobar 1935) bila je norveška slikarka, kao i žena svog učitelja i kolege, Kristijana Kroga.

Biografija

Bila je druga ćerka javnog tužioca Kristijana Lasona i Aleksandre fon Munte af Morgenstierne. Njena baka po majci je bila ruska princeza. Odrasla je u liberalno-konzervativnoj porodici, zajedno sa osam sestara i dva brata. Njen brat Per Lason je postao poznat kompozitor, a njena sestra Karolina Boken Lason pevačica i spisateljica.

Godine 1881. se udala za biznismena Jorgen Engelharta (1852-1921) sa kojim je imala dvoje dece. Rastali su se 1883, a razveli 1888. Godine 1885. je postala učenica Erik Verenskiolda i Kristijana Kroga, za koga će se i udati u oktobru 1888. Dobili su ćerku Nanu i sina Pera, koji je takođe će bio poznat slikar.

Pariz
Porodica je živela u Pariz od 1901-1909. godine. Na gravuri "Kafeinteriør" (1893) od Edvard Munk, Oda je okružena boemima i ljudima bliskim njima: Munk, Kristijan Krog, Jape Nilsen, Hans Jaeger, Gunar Heiberg i Jorgen Engelhart. Veruje se da je Oda bila u vezi sa svima njima osim Munka. U svojoj knjizi "Syk Kjærlihet" ("Obolela ljubav", 1893), Hans Jæger opisuje ljubavni trougao gde je bio jako zaljubljen u ženu koja je trebalo da se uda slikara. Smatra se da je to bila Oda u leto i jesen 1888. godine.

Berlin
Devedesetih godina prošlog veka se seli u Berlin. Tokom svađe sa Augustom Strindbergom, koji je bio rezigniran kara je Oda naštelovala njegovu gitaru pred gostima u na zabavi u Strindbergovom omiljenom baru "Zum Schwarzen Ferkel", branio je dramski pisac Gunar Heiberg. August Strindberg je bio poznat po svojoj lošoj naravi loše i nekoliko sukoba sa drugima, bez nekog posebnog razloga.

Ponovo Pariz
Oda Krog i Heiberg su se ubrzo zavoleli da bi 1897. godine ona uzela svog sina Pera i preselila se u Pariz sa Heibergom. Njen muž je postao instruktor umetnosti na Akademiji Kolarosi u Parizu a Oda je ubrzo zatim dobila atelje u Monparnasu. Za kratko vreme se združila sa vodećim umetnicima u gradu, uključujući i Henri Matis. Svoje radove je predstavila 1903. godine na Pariskom salonu, a godinu dana kasnije imala svoju prvu izložbu u salonu d'Automne, gde je bila aktivna do 1909. godine. Za to vreme je bila u vezi sa pesnikom i likovnim kritičarom Jape Nilsenom. Oda se rastala od njega i vratila svom mužu. Malo kasnije su se vratili u Oslo.

Dela
Sa malo formalnog obrazovanja u umetnosti, ona je ubrzo pokupila znanje iz umetničkog okruženja kome je pripadala. Njen debi je bio 1886. godine sa Ved Kristianiafjordenom (Japansk lykt), koji se nalazi u Nacionalnoj galeriji Norveške. Njeni rani radovi predstavljaju primer novog romantizma u slikarstvu, dok njeni kasniji radovi ostavljaju snažniji utisak. Oda Krog je poznata po svojim pejzažima, među njima Ved Kristianiafjord i Ved Engen. Ona je takođe radila portrete, među kojima su: Asta Hansten, Ivar Arosenius, Gunar Heiberg, Johan Dibvad i Kristijan Krog.

Oda je takođe bila centralna figura u anti-kulturnom pokretu Boemi Kristijanije tokom 1880ih i 1890ih. («Kristiania-bohemen») in the 1880s and 1890s. Njen javni imidž kao "boemske princeze" je donekle zasenio nju kao kompetentnu slikarku.

Uprkos želji da postane spisateljica, vrlo malo toga je objavila tokom svog života.

Njen život je opisan u romanu Ketil Bjornstad "Oda!" (1983). Sahranjena je na nacionalnom groblju u Oslu.

Референце

Dodatna literatura

 Anne Wickstrøm. «Oda Krohg» In Norsk biografisk leksikon, vol. 5 (2002)

 Anne Wickstrøm. «Oda Krohg» In Norsk kunstnerleksikon, vol. 2, 1983
 Per Krohg. «Oda Krohg : Tale ved Chr. Krohgs fest 18. nov. 1952». In Kunst og kultur, 1953

Spoljašnje veze
  krus.com; Kunstnere i Akershus About her connection to Akershus
  Sørlandets kunstmuseum About the painting Aftenposten (1887)
  o-vaering.no Gallery

Рођени 1860.
Умрли 1935.
Норвешки сликари

redpajama_set_name: RedPajamaWikipedia


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Улица Циолковского () — улица в Новозаводском районе города Чернигова, исторически сложившаяся местность (район) Шерстянка. Пролегает от улиц Попова и Текстильщиков до перекрёстка улицы Ивана Мазепы и Михайло-Коцюбинского шоссе.

Примыкают переулки Дмитрия Самоквасова, Рабочая, Леонида Пашина, Рассветная, Славянская, Славутичская, Новый переулок (село Павловка).

История
Козлянская улица — в честь прежнего названия Михайло-Коцюбинского Козёл — проложена в 1930-е годы. После сооружения железной дороги до села Козёл, дорогу к Козлу проложили в другом месте, а старая дорога, которая уже именовалась Старокозлянская начала постепенно застраиваться.

В 1960 году улица получила современное название — в честь русского и советского философа, изобретателя, основоположника теоретического космонавтики Константина Эдуардовича Циолковского.

Застройка
Улица пролегает в юго-западном направлении, меняя углы направления между более западным или более южным. Улица имеет по два ряда движения в обе стороны.

Начало улицы (до примыкания улицы Леонида Пашина) занято территорией промышленных (завод стройматериалов № 2), коммунальных (ПМК - передвижная механизированная колона, БРУ - строительные организации), складских (базы снабжения), специальных предприятий, кроме парной стороны от начала до примыкания улицы Дмитрия Самоквасова — многоэтажной жилой застройкой (5-9-этажные дома). После примыкания улицы Леонида Пашина парная сторона занята территорией промышленных (асфальтобетонный завод), коммунальных (АТП - автотранспортные), складских предприятий, непарная — не застроена. Западнее данного участка расположен район с индивидуальной застройкой Астра и село Павловка, конец улицы служит административной границей Черниговского горсовета с Черниговским районом.

Северо-западный угол улиц Циолковского и Леонида Пашина занимает урочище и городской лесопарк Подусовка — охранная зона памятника истории «Мемориальный комплекс в честь мирных жителей, расстрелянных фашистами: братские могилы 15000 мирных жителей, расстрелянных в 1941-1943 годах и памятник жертвам фашизма», открытый в 1975 году.

Учреждения:
 дом № 6 — детсад № 2
 дом № 8 — комплексная детско-юношеская школа № 2
 дом № 9 — «Черниговглавснаб»
 дом № 11 — завод стройматериалов № 2
 дом № 30 — хлебозавод (Рига-хлеб)
 дом № 41 — асфальтобетонный завод

Примечания

Литература
 Чернігівщина: Енциклопедичний довідник, К.: УРЕ і м. М. П. Бажана, 1990. — с. 848 Ціолковського ВУЛИЦЯ
 Сапон В. . Вулиці старого Чернігова: Історико-краєзн. етюди. — Чернігів: РВК «Десн. правда», 2007. — 128 с.

Улицы Чернигова
Улицы, названные в честь Константина Циолковского


"A small town believing residents and businesses can help one another make life better together"
Long Lake is near and dear to me. I've been a member of the Long Lake Chamber Of Commerce for the past 6 years, serving as its President for 2 of those years. Our family business, Albers Jewelry, has made its home in Long Lake for 28 years. The Mayor, a long time friend and past baseball coach (Sorry if this dates you Tim!), is all about serving his citizens and supporting area businesses. Mayor Tim Hultmann was the catalyst for courting the organizers of the World Wake Surfing Championships. 2013 marked the 2nd annual Wake Surfing World Qualifying Event held at Nelson Lakeside Park, on Long Lake. We certainly look forward to ensuring a long-standing relationship with the organizers of this awesome young sport that has gone "viral"!
Long Lake residents are served by the Orono School District. I am a proud alumni of Orono schools. Superintendent, Dr. Karen Orcutt and her team of dedicated educators are committed to providing a welcoming and challenging teaching environment. Recently, the Orono school district was one of three in the country to be named one of the National Schools of Character. It's the first public school district in Minnesota to get the award. The athletic program is top notch and consistently competes at a very high level.
Long Lake and its surrounding communities offer boutique shopping, diverse dining opportunities and plenty of parks, trails, and lakes. Mark your calendar so you and your family can stop by our 4th annual SummerFest event June 2014. Its a blast. Welcome home!
What do you like most about living in Long Lake, MN?


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Will the general session be recorded?
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Can I buy a ticket to the general session?
Is there extra conference swag to send to attendees if they ask for it, such as backpacks, lanyards, and so on?
Why am I receiving mailings from exhibitors, and how can I stop this?
Seating will be on a first come, first served basis. We will not be able to allow attendees to hold seats for other colleagues who haven't arrived yet.


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Home Bible Prophecy Christian Persecution State of Georgia demands pastor turn over sermons
State of Georgia demands pastor turn over sermons
A lay minister who is suing the Georgia Department of Public Health for religious discrimination has been ordered by the state's attorney general to relinquish his sermons to the government, according to federal court documents.
"Please produce a copy of your sermon notes and/or transcripts," Attorney General Samuel Olens wrote to attorneys representing Dr. Eric Walsh.
Dr. Walsh said he will not comply with the request.
"No government has the right to require a pastor to turn over his sermons," Walsh said. "I cannot and will not give up my sermons unless I am forced to do so."
Walsh, a Seventh-day Adventist lay minister had been hired in May 2014 by as a District Health Director with the Georgia Department of Public Health. A week later, a government official asked him to submit copies of his sermons for review. He complied and two days later he was fired.
His attorneys said the government was curious about sermons Dr. Walsh delivered on health, marriage, sexuality, world religions, science and creationism. He also preached on what the Bible says regarding homosexuality.
He has since filed a federal lawsuit charging state officials with engaging in religious discrimination.
"He was fired for something he said in a sermon," attorney Jeremy Dys told me. "If the government is allowed to fire someone over what he said in his sermons, they can come after any of us for our beliefs on anything."
Dr. Walsh has assembled a powerhouse legal team comprised of Parks, Chesin & Walbert along with First Liberty Institute, one of the nation's most prominent religious liberty law firms.
"It's an incredible intrusion on the sanctity of the pulpit," Dys said. "This is probably the most invasive reach into the pulpit by the state that I've ever seen."
Neither the attorney general nor his communication director responded to telephone calls or email correspondence.
Dys said the government's request came as part of the process to build their case against his client. The request, he said, carries the same force of law as a subpoena.
"Pastors and rabbis across the country – especially in the state of Georgia should be frightened that a state would demand all the sermon notes and transcripts of a pastor," he said. "This is unprecedented."
It may be unprecedented, but is it illegal?
"I think it is wrong for a state to demand that a pastor turn over his sermons at any time," Dys said. "It was wrong for the state to ask Dr Walsh for his sermons as part of the job application and hiring process. It was wrong for them to turn around and fire him when they listened to those sermons. And it is wrong now for them to demand that he turn over all the sermons and sermon notes he ever produced going back to when he was 18 years old."
Family Research Council president Tony Perkins condemned the state's demand calling it "an alarming display of government intrusion into the sanctity of the church, pastor's study and pulpit."
"This is something that I would have expected to see in a communist country, not America," Perkins told me. "Government scrutiny of speech in the pulpit is unconstitutional, and unconscionable."
He called on Gov. Nathan Deal, a Republican, to "correct this egregious overreach of the state into church affairs."
Dr. Walsh also gained the support of Emir Caner, president of Truett-McConnell University, a Christian school based in Cleveland, Georgia.
"A new era has come to our shores, a time where government finds it acceptable to suppress the freedom of religion even to the extent of requesting a minister's sermons," Caner said. "As an ordained minister, I know that this is not merely an assault on the messenger, but on the very message of our Sacred Scriptures."
Caner correctly noted that the government seems to exclusively target one faith – biblical Christianity.
"What the government fails to recognize is that ministers of the Gospel are not hirelings of the state, but ambassadors for our Lord," he said.
Dr. Walsh was catapulted into the national spotlight when he was invited to deliver the 2014 commencement address at Pasadena City College in California.
Walsh, who was then the director of the city's public health department, came under fierce scrutiny from LGBT activists and students after someone found online copies of his sermons. He backed out of the speaking engagement – but critics persisted and he was forced to resign his post – after reaching a financial settlement with the city.
Shortly afterwards, he applied for the position in Georgia.
"The state of Georgia fired him for something he said in a sermon," Dys told me. "It's almost like the state ransacking the pastor's study trying to find evidence in a desperate attempt to justify their illegal behavior."
Dys said the attorney general could ask a judge to enforce their request. If Dr. Walsh still refused, he could face contempt of court.
"No one in America should want a pastor being held in contempt for refusing to give up one of his sermons to the state of Georgia," he said.
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Howell Solar Power
Howell solar power is a solar energy company located in Monmouth County in Howell, NJ, USA. Our company provides various kinds of solar energy solutions required to power your home or business. We are the one-stop for solar panels and all other kinds of inexpensive solar facilities.
1001 U.S. 9, Howell Township, NJ 07731, USA
Privacy Policy & Warranty: Howell Solar Power respects and is committed to protecting your privacy. We do not share any customer's personal information.
Copyright © 2023 Howell Solar Power


Copyright: © Komatsu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.
Bisphosphonates (BPs) are analogues of pyrophosphate that are known to prevent bone resorption by inhibiting osteoclast activity. Nitrogen-containing BPs, such as zoledronic acid (ZA), are widely used in the treatment of osteoporosis and bone metastasis. However, despite having benefits, ZA has been reported to induce BP-related osteonecrosis of the jaw (BRONJ) in cancer patients. The molecular pathological mechanisms responsible for the development of BRONJ, including necrotic bone exposure after tooth extraction, remain to be elucidated. In this study, we examined the effects of ZA on the transforming growth factor-β (TGF‑β)-induced myofibroblast (MF) differentiation of human gingival fibroblasts (hGFs) and the migratory activity of hGFs, which are important for wound closure by fibrous tissue formation. The ZA maximum concentration in serum (Cmax) was found to be approximately 1.47 µM, which clinically, is found after the intravenous administration of 4 mg ZA, and ZA at this dose is considered appropriate for the treatment of cancer bone metastasis or bone diseases, such as Erdheim-Chester disease. At Cmax, ZA significantly suppressed i) the TGF‑β-induced promotion of cell viability, ii) the TGF‑β-induced expression of MF markers such as α-smooth muscle actin (α-SMA) and type I collagen, iii) the TGF‑β-induced migratory activity of hGFs and iv) the expression level of TGF‑β type I receptor on the surfaces of hGFs, as well as the TGF‑β-induced phosphorylation of Smad2/3. Thus, ZA suppresses TGF‑β-induced fibrous tissue formation by hGFs, possibly through the inhibition of Smad‑dependent signal transduction. Our findings partly elucidate the molecular mechanisms underlying BRONJ and may prove to be beneficial to the identification of drug targets for the treatment of this symptom at the molecular level.
Bisphosphonates (BPs) are analogues of pyrophosphate that are known to prevent bone resorption by inhibiting osteoclast activity (reviewed in ref. 1). BPs are used for the treatment of various bone diseases, including osteoporosis and bone metastasis in cancer with or without hypercalcemia (2). Nitrogen-containing BPs, such as zoledronic acid (ZA), are used in the treatment of osteoporosis and bone metastasis (reviewed in ref. 3), as well as in the treatment of other bone diseases, such as Erdheim-Chester disease (4). In addition, they are more potent inhibitors of bone resorption than non-nitrogen-containing BPs (reviewed in ref. 5). However, despite their several benefits, ZA induces BP-related osteonecrosis of the jaw (BRONJ) in cancer patients (6,7). BRONJ is defined as necrotic bone exposure in the oral cavity continuing for >8 weeks in BP-treated patients who have not had received head and neck radiation therapy (8). The molecular mechanisms responsible for the development of BRONJ remain to be elucidated; however, some risk factors, including periodontitis with bacterial plaque in the oral cavity have been associated with this symptom (reviewed in ref. 7). Previous studies have suggested that infectious agents, including Actinomyces, play important roles in the etiology and progression of BRONJ (9–11). Interestingly, Kobayashi et al reported that ZA promoted the adherence of Streptococcus mutans to hydroxyapatite and the proliferation of oral bacteria obtained from healthy individuals, suggesting that ZA increases bacterial infection (12). Wound closure by oral epithelial cells (ECs) and gingival fibroblasts (GFs) is important, not only for successful wound healing, but also for the protection of the socket from oral bacterial infection following tooth extraction. Thus, it is possible that an oral bacterial infection may induce BRONJ in cooperation with other risk factors, such as diabetes mellitus with steroid intake (13) or with microvascular disease (14), smoking, prosthetic trauma, and implant treatment (15). In addition, the ability of GFs to migrate and synthesize type I collagen is essential for the formation of rigid gingival connective tissue to cover the intraoral bone exposure, which protects the alveolar bone of the maxilla and the jaw from infection with oral bacteria. However, the mechanisms through which ZA affects the ability of human GFs (hGFs) to migrate and synthesize the connective tissue at the molecular level remain to be elucidated.
When tissue injury occurs, the blood coagulation cascade is initiated, resulting in the formation of fibrous connective tissue, which functions as a scaffold for the influx of inflammatory cells. Platelets also aggregate at the site of injury, and become activated by binding to the negatively charged extravascular fibrous tissue, and release various growth factors, including transforming growth factor-β (TGF-β) (reviewed in ref. 16). TGF-β1 is known to be expressed in serveral types of cells, including hGFs, and is involved in the proliferation and differentiation of these cells (17,18). Thus, the functions of TGF-β seem to be autocrine or paracrine as regards the regulation of hGFs during oral inflammation and the wound healing processes at the site of injury. The TGF-β superfamily consists of two families: the TGF-β/activin/Nodal family and the bone morphogenetic protein (BMP)/growth and differentiation factor (GDF)/Mullerian inhibiting substance (MIS) family (reviewed in ref. 19).
The TGF-β superfamily ligands initiate a cascade of signaling events by binding to their respective type I and type II receptors in the extracellular space. Following this, two type I and two type II receptors form a tetrameric complex. In this ligand-bound complex of type I and type II receptors, the type II receptor kinase activates the type I receptor kinase. The type I receptor induces intracellular signal transduction by phosphorylating the receptor-regulated Smads (R-Smads) (reviewed in refs. 20–23). Smads are central signal transducers of TGF-β superfamily and are composed of 3 groups. The first group comprises the R-Smads; Smad1, Smad5 and Smad8 are primarily activated by the BMP-specific type I receptors, whereas Smad2 and Smad3 are activated by TGF-β-specific type I receptors. The second group contains the common mediator Smad (Co-Smad; e.g., Smad4). The third group comprises the inhibitory Smads (I-Smads; e.g., Smad6 and Smad7). Activated R-Smads form complexes with the Co-Smad, which enter the nucleus, and, together with other cooperative proteins, positively or negatively control the transcription of specific target genes. I-Smads suppress the activation of R-Smads by competing with R-Smads for type I receptor interaction and by recruiting specific ubiquitin ligases, resulting in their proteasomal degradation (reviewed in ref. 24).
TGF-β has the ability to induce the differentiation of various types of cells into myofibroblasts (MFs), which typically exhibit the formation of F-actin stress fibers (reviewed in ref. 25). We have previously demonstrated that TGF-β induces the expression of the MF markers, α-smooth muscle actin (α-SMA) and type I collagen, in fibroblastic cells derived from periodontal ligament (26). Sobral et al reported that TGF-β induced the differentiation of hGFs into MFs in a Smad-dependent manner (27). Thus, TGF-β is now known to induce MF differentiation from fibroblasts, which preferentially form a fibrous tissue. TGF-β is also known to induce migratory activity in various types of cells, including fibroblastic cells in a Smad-dependent manner (28–30). In addition, Bakin et al reported that TGF-β regulates the migratory activity of ECs in a p38 mitogen-activated protein kinase (MAPK)-dependent manner (31), suggesting that TGF-β induces cell migratory activity through MAPKs as opposed to Smads. Moreover, TGF-β plays important roles in physiological wound closure by promoting cell migration and type I collagen synthesis. On the other hand, the abnormal and persistent appearance of MFs causes scar formation or fibrosis (32,33). The abnormal potentiation of TGF-β signaling in MFs possibly causes fibrogenic diseases. Therefore, MFs represent key players in the physiological reconstruction of connective tissue following injury and in generating the pathological tissue deformations that characterize fibrosis (34).
In this study, we investigated the mechanisms through which ZA, at its maximum concentration in serum (Cmax), which clinically, is usually found after the intravenous administration of 4 mg ZA, which is the appropriate amount for the usual treatment of bone metastasis (35,36) or bone diseases such as Erdheim-Chester disease (4), affects TGF-β-induced intracellular signal transduction and MF differentiation of hGFs, which are important processes for the progression of fibrogenesis during inflammation.
Recombinant TGF-β was purchased from Peprotech, Inc. (Rocky Hill, NJ, USA). The TGF-β type I receptor inhibitor, SB-431542, which preferentially suppresses the activation of the intracellular signal transduction of Smad2 by this receptor (37,38), and sometimes broadly suppresses the TGF-β1-induced activation of p38 MAPK, and extracellular signal-regulated kinase (ERK) as opposed to Smad2 (30) was purchased from Calbiochem (La Jolla, CA, USA). Zometa® obtained from Novartis Pharmaceuticals (Tokyo, Japan) was used as ZA in all our experiments. The Cmax of ZA in an adult human body is approximately 1.47 µM within 15 min after its intravenous administration (4 mg/5 ml) according to the user instructions provided with Zometa®. This is the appropriate amount of ZA for the treatment of cancer bone metastasis (32,33), and is referred in the internal document ZOMU00007 belonging to Novartis Pharmaceuticals.
hGFs were isolated and cultured as described in our previous study (39). Briefly, the cells were isolated from the fresh gingival tissue biopsy samples of 3 volunteers, and were maintained in a Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal bovine serum (FBS) and penicillin-streptomycin (both from Invitrogen, Gaithersburg, MD, USA). Informed consent was obtained from all the volunteers prior to obtaining the samples, and the Ethics Committee of Iwate Medical University approved the research protocol (approval no. 1126). Subsequently, one hGF culture retaining a high proliferative potential among the 3 cultures was used. NIH3T3 mouse embryonic fibroblasts (used as a standard control for the fibroblasts), obtained from RIKEN Cell Bank (Tsukuba, Japan), were cultured with DMEM supplemented with 10% FBS and penicillin-streptomycin (both from Invitrogen).
The status of cell viability was evaluated using an alamarBlue assay (AbD Serotec, Oxon, UK) according to the manufacturer's instructions. This assay reagent includes an indicator that fluoresces and undergoes colorimetric changes when reduced by mitochondrial respiration, which is proportional to the number of living cells. For the viability assay, the cells were seeded in 96-well plates at a density of 2.48×103 cells/well and cultured for 48 h in medium containing 10% FBS, with or without ZA at the indicated concentrations (0.0147-147 µM). Some of the cells were subsequently treated with TGF-β1 (1–5 ng/ml) for 24 h following treatment with ZA. The medium was replaced with DMEM containing 10% alamarBlue solution to evaluate the viability of the cells, and the cells were cultured for an additional 4.5 h. The absorbance in each well was measured using an ELISA plate reader (Tosoh Corp., Tokyo, Japan). The data were presented as values of Abs570 − Abs600. Each experiment was repeated 3 times, with 5-wells dedicated for each time point.
Total RNA from the hGFs and NIH3T3 cells was isolated using ISOGEN II reagent (Nippon Gene, Toyama, Japan) according to the manufacturer's instructions. First-strand cDNA was synthesized from total RNA using the PrimeScript RT reagent kit (Takara Bio, Shiga, Japan). PCR was subsequently performed on a Thermal Cycler Dice Real-Time system using SYBR Premix Ex Taq II (both from Takara Bio) with specific oligonucleotide primers (human α-SMA forward, 5′-ATACAACATGGCATCATCACCAA-3′ and reverse, 5′-GGGCAACACGAAGCTCATTGTA-3′; mouse α-SMA forward, 5′-CAGATGTGGATACAGCAAACAGGA-3′ and reverse, 5′-GACTTAGAAGCATTTGCGGTGGA-3′; human TGF-β type I receptor forward, 5′-GCTGCTCCTCCTCGTGCT-3′ and reverse, 5′-TTGTCTTTTGTACAGAGGTGGC-3′; human TGF-β type II receptor forward, 5′-CTGCACATCGTCCTGTGG-3′ and reverse, 5′-GGAAACTTGACTGCACCGTT-3′; and human glyceraldehyde 3-phosphate dehydrogenase (GAPDH) forward, 5′-GCACCGTCAAGGCTGAGAAC-3′ and reverse, 5′-ATGGTGGTGAAGACCCCACT-3′; and mouse GAPDH forward, 5′-TGTGTCCGTCGTGGATCTG-3′ and reverse, 5′-TTGCTGTTGAAGTCGCAGGAG-3′). The mRNA expression levels of α-SMA, TGF-β type I receptor and TGF-β type II receptor were normalized to those of GAPDH, and the relative expression levels were presented as the fold increase or decrease relative to the control.
The cells were lysed in RIPA buffer [50 mM Tris-HCl (pH 7.2), 150 mM NaCl, 1% NP-40, 0.5% sodium deoxycholate and 0.1% SDS] or lysis buffer [20 mM HEPES (pH 7.5), 150 mM NaCl, 1 mM EDTA and 1% Triton® X-100] containing protease and phosphatase inhibitor cocktails (both from Sigma, St. Louis, MO, USA). The protein content of the samples was measured using BCA reagent (Pierce, Rockford, IL, USA). Samples containing equal amounts of protein were separated on 10% SDS-polyacrylamide gels and transferred onto a polyvinylidenedifluoride membrane (Millipore Corp., Bedford, MA, USA). After being blocked with 1% BSA or 1% skim milk in T-TBS (50 mM Tris-HCl, pH 7.2, 150 mM NaCl and 0.05% Tween-20), the membrane was incubated with primary antibodies including anti-α-SMA rabbit polyclonal antibody (1:1,000; ab5694; Abcam, Cambridge, UK), anti-Smad2/3 purified mouse monoclonal antibody (1:1,000; 610842; BD Transduction Laboratories™, Franklin Lakes, NJ, USA), anti-phospho-Smad2/3 (#8828), anti-p38 MAPK (#9212), anti-phospho-p38 MAPK (#9211), anti-c-Jun N-terminal kinase (JNK; #9251) and anti-phospho-JNK polyclonal antibodies (#9252) (1:1,000; all from Cell Signaling Technology, Inc., Beverly, MA, USA) and anti-α-tubulin mouse monoclonal antibody (1:25,000; #3873, Cell Signaling Technology, Inc.) as a loading control for normalization. The proteins of interest were then detected using appropriate horseradish peroxidase-conjugated secondary antibodies (Cell Signaling Technology, Inc.) and an Amersham ECL™ Prime Western Blotting Detection reagent (GE Healthcare Bio-Sciences, Pittsburgh, PA, USA).
For immunofluorescence analysis of the cultured cells, the hGFs were subcultured on non-coated cover glass slips (Matsunami Glass Ind., Ltd. Kishiwada, Japan) at a density of 2.5×104 cells/glass slip (BD Biosciences, Franklin Lakes, NJ, USA) and maintained in DMEM supplemented with 10% FBS, with or without ZA (1.47 µM) for 48 h. The culture medium was replaced with DMEM without FBS, supplemented with or without TGF-β1 (5 ng/ml). The cells were maintained in this medium for 4 or 5 days, fixed in 4% paraformaldehyde (Nacalai Tesque, Inc., Kyoto, Japan) for 15 min and permeabilized with Triton X-100 (Sigma). Following background reduction with normal goat serum, the cells were labeled with anti-α-SMA rabbit polyclonal antibody (1:100; ab5694; Abcam), or anti-collagen type I rabbit polyclonal antibody (1:100; 600-401-103-0.1; Rockland, Inc., Rockland, ME, USA) at room temperature for 1 h. After being washed with phosphate-buffered saline (PBS) to remove the excess primary antibody, the cells were incubated with Alexa Fluor® 488-conjugated goat anti-rabbit IgG (1:1,000; A-11034; Molecular Probes, Leiden, The Netherlands) and DAPI (1:1,000; KPL, Gaithersburg, MD, USA) for 30 min at room temperature. After being washed with PBS to remove the excess secondary antibody, the fluorescent signal was detected using an Olympus IX70 fluorescence microscope with the LCPIanFI 20 objective lens (Olympus Co., Tokyo, Japan).
Cell migration assays were performed using Transwell® membrane cell culture inserts (8 µm pore size; Corning Inc., Corning, NY, USA) according to the manufacturer's instructions. The hGFs were cultured with or without ZA (1.47 µM) in DMEM supplemented with 10% FBS for 48 h at a cell density of 6.0×105 cells in 10 cm culture dish. The cells were then placed at a density of 1.0×105 cells in the cell culture inserts in the each well of a 24-well cell culture plate in DMEM with 0.1% BSA. The cells were allowed to migrate through the porous membrane that bordered the upper cell culture insert and the lower well of 24-well culture plate which contained DMEM in the presence or absence of TGF-β (5 ng/ml) for 6 h at 37°C. TGF-β was added to the culture medium in the lower well of 24-well culture plate. In some cases, SB-431542 was added to both the upper and lower culture media. The cells on the upper side of the membrane were wiped, and the membrane was fixed in 4% paraformaldehyde in PBS. After being washed with PBS, the cells that had migrated onto the underside of the membrane were labeled with DAPI (1:1,000; KPL) and counted. The values were shown as average of those from 3 wells. The evaluation of the migratory activity of the NIH3T3 cells was performed as described above without pre-treatment of the cells with ZA.
The hGFs were cultured with or without ZA (1.47 µM) in DMEM supplemented with 10% FBS for 48 h at a density of 6.0×105 cells in a 10-cm culture dish. The cells (1.0×105 cells) were suspended in PBS containing 0.5% FBS and 2 mM EDTA and incubated with anti-TGF-β receptor type I (1:100; ab30103) or type II (1:50; ab78419) (both from Abcam) primary antibodies for 1 h at 4°C. For the negative control experiments, the cells were incubated with the same protein amount of normal control IgG (sc-2028; Santa Cruz Biotechnology, Inc., Santa Cruz, CA, USA) as each specific antibody. The cells were then incubated with PE-conjugated secondary antibodies (#732988 for ab31013; #732970 for ab78419) for 30 min in the dark. Acquisition was performed using an EPICS XL EXPO 32 ADC system (Beckman Coulter, Fullerton, CA, USA).
The data are presented as the mean ± SD (n=3 or 5 experiments). The data were statistically analyzed using the Student's t-test, and P<0.01 (indicated by an asterisk) was considered significant. The results shown in all experiments are representative of at least 3 separate experiments.
Treatment with ZA for 48 h in medium containing 10% FBS at the concentrations of 0.0147 µM (1/100 of Cmax), 0.147 µM (1/10 of Cmax) and 1.47 µM (Cmax) did not affect the viability of the hGFs (Fig. 1A) and the NIH3T3 cells which were used as a standard control for the fibroblasts (Fig. 1B). However, at 10-100-fold higher concentrations (14.7–147 µM) than the Cmax, ZA significantly suppressed the viability of the hGFs (35–58% suppression of the control) and that of the NIH3T3 cells (29–80% suppression of the control) in a dose-dependent manner. Thus, ZA (Cmax) does not affect the viability of hGFs and normal standard fibroblasts at 48 h following administration.
Zoledronic acid (ZA) suppresses the transforming growth factor-β1 (TGF-β1)-induced increase in the viability of human gingival fibroblasts (hGFs) at the maximum concentration in serum (Cmax). (A) hGFs and (B) NHI3T3 cells were seeded in 96-well plates at a density of 2.48×103 cells/well and cultured for 48 h in medium containing 10% fetal bovine serum (FBS) with or without ZA at the indicated concentrations (0.0147–147 µM). The medium was replaced with DMEM containing 10% alamarBlue solution, and the cells were cultured for an additional 4.5 h. To evaluate the viability of the cells, the absorbance (Abs570−Abs600) in each well was measured using a plate reader. (C) Firstly, hGFs were seeded in 96-well plates at a density of 2.48×103 cells/well and cultured for 48 h in medium containing 10% FBS with or without ZA (Cmax) (lower, and upper graphs, respectively). The cells were subsequently stimulated with TGF-β1 (1–5 ng/ml) for 24 h. The viability of the hGFs was then measured as described above. The data are presented as the means ± SD (n=5). *P<0.01 was considered significant.
We examined whether ZA (Cmax) affects the viability of hGFs stimulated with TGF-β1 (1–5 ng/ml). As shown in Fig. 1C (upper graph), TGF-β1 (1–5 ng/ml) increased the viability of the hGFs (23–47% promotion of the control). However, ZA (Cmax) suppressed the TGFβ1-induced increase in the viability of the hGFs (Fig. 1C, lower graph; compare same TGF-β1 concentrations between graphs).
TGF-β1 (5 ng/ml) significantly upregulated the α-SMA mRNA expression level in the hGFs (Fig. 2A, bars 1 and 2 from left). ZA (Cmax) completely suppressed the TGF-β1-induced increase in the mRNA expression of α-SMA in the hGFs (Fig. 2A, bars 2 and 4 from left). In addition, western blot analysis revealed that ZA (Cmax) clearly suppressed the TGF-β1-induced upregulation of α-SMA expression at the protein level in the hGFs (Fig. 2B). Immunofluorescence staining also revealed that ZA (Cmax) clearly suppressed the TGF-β1-induced upregulation of α-SMA expression (Fig. 2C, left panels) and type I collagen expression (Fig. 2C, right panels) at the protein level in the hGFs. We confirmed that the TGF-β1-induced increase in the mRNA expression of α-SMA in the hGFs was significantly inhibited by SB-431542, an inhibitor of TGF-β type I receptor (Fig. 2D), indicating that TGF-β1 specifically induced the MF differentiation of hGFs through ligand-receptor interaction. Thus, these results indicate that ZA (Cmax) suppresses the TGF-β-induced MF differentiation of hGFs.
Zoledronic acid (ZA) at the maximum concentration in serum (Cmax) inhibits the transforming growth factor-β (TGF-β)-induced myofibroblast (MF) differentiation of human gingival fibroblasts (hGFs). (A) hGFs were cultured for 48 h in growth medium supplemented with 10% fetal bovine serum (FBS) in the presence or absence of ZA (1.47 µM or Cmax value of an adult human; ZA administered intravenously at 4 mg/5 ml). The culture medium was replaced with DMEM without FBS supplemented with or without TGF-β1 (5 ng/ml) and maintained for 24 h. The relative expression level of MF marker α-smooth muscle actin (α-SMA) was analyzed by RT-qPCR. Data represent the means ± SD (n=3). *P<0.01 was considered significant. (B) The status of MF differentiation of hGFs at the protein level was evaluated by western blot analysis with anti-α-SMA antibody. The cells were cultured with or without ZA (Cmax) as described in (A), and were subsequently cultured in DMEM without FBS supplemented with or without TGF-β1 (5 ng/ml) for 4 days [α-SMA, upper panel; α-tubulin (internal control), lower panel]. (C) The cells were first cultured with or without ZA (Cmax) as described in (A), cultured in DMEM without FBS supplemented with or without TGF-β1 (5 ng/ml) for 4 (α-SMA; left panels) or 5 (type I collagen; right panels) days. The cells were then immunostained with anti-α-SMA (left panels) (green) and anti-type I collagen (right panels) (green) antibodies. Nuclei were fluorescently counterstained with DAPI (blue). Scale bar, 50 µm. (D) hGFs were cultured for 48 h in growth medium supplemented with 10% FBS, and the medium was replaced with DMEM without FBS supplemented with or without TGF-β1 (5 ng/ml). The cells were maintained for 24 h. Some of the cells were treated with the TGF-β receptor inhibitor, SB-431542 (10 µM), which was added to the culture medium 30 min prior to stimulation with TGF-β1. The relative mRNA expression level of MF marker α-SMA was analyzed by RT-qPCR. Data represent the means ± SD (n=3). *P<0.01 was considered significant.
As shown in Fig. 3A and C, the migration of the hGFs through the porous membrane that bordered the upper and lower chambers was significantly enhanced by stimulation with TGF-β1 (5 ng/ml). However, ZA (Cmax) completely suppressed the TGF-β1-induced migratory activity (Fig. 3A and B). In addition, we confirmed that the TGF-β-induced migratory activity was markedly inhibited by SB-431542 (Fig. 3C and D), indicating that TGF-β1 specifically induced the migration of hGFs through ligand-receptor interaction. These results indicate that ZA (Cmax) inhibits the TGF-β-induced migratory activity of hGFs.
Zoledronic acid (ZA) at the maximum concentration in serum (Cmax) inhibits the transforming growth factor-β (TGF-β)-induced migratory activity of human gingival fibroblasts (hGFs). Cell migration assays were performed using Transwell® membrane cell culture inserts as described in the Materials and methods. (A and B) hGFs were cultured with or without ZA (1.47 µM) for 48 h. The cells were placed in the cell culture inserts in the each well of 24-well cell culture plate. The cells were then allowed to migrate through the porous membrane that bordered the upper cell culture insert and the lower well of 24-well culture plate in the presence or absence of TGF-β1 (5 ng/ml). (C and D) hGFs were placed in the cell culture inserts as described above. The cells were allowed to migrate through the membrane in the presence or absence of TGF-β1 (5 ng/ml). In some cases, SB-431542 was added to both the upper and lower culture media. The cells that had migrated onto the underside of the membrane were labeled with DAPI and visualized using a fluorescent microscope (A and C) and counted (B and D). Data represent the means ± SD (n=3). *P<0.01 was considered significant.
In order to gain insight into the molecular mechanisms underlying the suppressive effects of ZA (Cmax) on TGF-β-induced fibrogenesis by hGFs, we investigated whether ZA affects the expression of TGF-β type I and II receptors on the surface of hGFs. In flow cytometric analysis, hGFs had two peaks in the histogram of TGF-β type I receptor-positive cells, that is, 2 groups of hGFs weakly or strongly expressed TGF-β type I receptors, respectively (Fig. 4A, upper left panel). Intriguingly, ZA (Cmax) lowered the ratio of the number of cells strongly expressing TGF-β type I receptors on their surface against that of total hGFs by 19.6 to 6.4% (Fig. 4A, upper panels). By contrast, hGFs had only one peak in the histogram of TGF-β type II receptor-positive cells (Fig. 4A, lower left panel). ZA (Cmax) did not change the position of the peak in the histogram of TGF-β type II receptor-positive cells (Fig. 4A, lower panels). Moreover, RT-qPCR analysis revealed that ZA (Cmax) did not decrease the total amount of both TGF-β receptor type (types I and II) expression at the mRNA level (Fig. 4B, upper and lower graphs, respectively), suggesting that ZA (Cmax) suppressed the expression of TGF-β type I receptor on the surface of hGFs. In addition, ZA (Cmax) suppressed the TGF-β1 (5 ng/ml)-induced phosphorylation of Smad2/3 (Fig. 4C). We confirmed these results by reproducing them in independent experiments.
Zoledronic acid (ZA) at the maximum concentration in serum (Cmax) suppresses the expression of transforming growth factor-β (TGF-β) type I receptor on the surface of human gingival fibroblasts (hGFs) and the phosphorylation of Smad2/3 in hGFs. (A) hGFs were cultured with or without ZA (Cmax) for 48 h as described in Fig. 2 at a density of 6.0×105 cells in a 10-cm culture dish. The expression levels of TGF-β type I (upper panels), and II (lower panels)-receptors on the surface of hGFs were evaluated by flow cytometry using anti-TGF-β type I receptor (red) and anti-TGF-β type II receptor (red) antibodies or the same amount of normal control IgG (blue). (B) hGFs were cultured for 48 h in growth medium supplemented with 10% fetal bovine serum (FBS) in the presence or absence of ZA (Cmax). The relative mRNA expression levels of TGF-β type I receptor (upper graph) and TGF-β type II receptor (lower graph) were analyzed by RT-qPCR. Data represent the means ± SD (n=3). *P<0.01 was considered significant. (C) hGFs were cultured with or without ZA (Cmax) as described in (A). The cells were subsequently cultured with or without TGF-β1 (5 ng/ml) for 1 h. The phosphorylation of Smad2/3 was evaluated by western blot analyses with anti-Smad2/3 and anti-phospho-Smad2/3 antibodies.
Wang et al reported that TGF-β1 increased the viability of human dermal fibroblasts (40). We found that ZA (Cmax) reduced the TGF-β1-induced increase in the viability of hGFs (Fig. 1C). These results suggest that ZA (Cmax) possibly suppresses the TGF-β1-induced fibrogenesis in oral gingival tissue. Lu et al reported that TGF-β1 promoted the viability of pulmonary artery endothelial cells through Smad2-mediated signal transduction (41). We found that ZA (Cmax) suppressed the TGF-β1 (5 ng/ml)-induced phosphorylation of Smad2/3 (Fig. 4C), implicating that ZA (Cmax) may possibly suppress the TGF-β1-induced promotion of the viability of hGFs through suppression of Smad2/3 activities.
Pan et al or Koch et al reported that 5 or 50 µM ZA induced the osteoblastic differentiation of osteoblast precursor cells (42,43). In addition, Chen et al reported that 10 µM ZA induced the dendritic cell differentiation of monocytes (44). However, the mechanisms through which ZA (Cmax) affected the differentiation ability of cells derived from the oral cavity remained to be elucidated. In this study, we demonstrated that 1.47 µM ZA (Cmax) significantly suppressed the TGF-β1-induced MF differentiation of hGFs (Fig. 2A, B, and C). These results strongly suggest that ZA (Cmax) inhibits wound healing in the oral cavity by the attenuation of type I collagen synthesis in MFs differentiated from hGFs following TGF-β1 stimulation. In addition, it is generally known that MFs play an important role in generating a contractile force, facilitating wound closure (reviewed in ref. 16), suggesting that ZA (Cmax) also interferes with this process through the suppression of the TGF-β1-induced MF differentiation of hGFs. Of note, we found that ZA (Cmax) clearly suppressed the TGF-β1-induced phosphorylation of Smad2/3 in hGFs (Fig. 4C). Smad2/3 are known to be key signaling molecules inducing the MF differentiation of fibroblasts (27). These results suggest that ZA (Cmax) suppressed TGF-β1-induced MF differentiation of hGFs through inhibition of Smad2/3 phosphorylation by TGF-β1.
Pabst et al reported that ZA at 50 µM, a much higher concentration than Cmax, significantly decreased the migratory activity of human oral keratinocytes at 72 h following treatment with ZA (45). However, it remained to be clarified whether ZA (Cmax) affected the migratory activity of various cells derived from the oral cavity, including hGFs. On the other hand, TGF-β is known to induce migratory activity in various types of cells, including fibroblastic cells, in a Smad-dependent manner (28–30). In this study, we demonstrated that ZA (Cmax) significantly suppressed the TGF-β1-induced migratory activity of hGFs (Fig. 3A and B). Wound healing or mucous re-epithelialization is a complex process involving the proliferation and migration of mesenchymal cells, such as fibroblasts and MFs, which construct granulation tissue for wound closure and then induce mucous re-epithelialization on it (reviewed in ref. 46). As described above, ZA (Cmax) suppressed TGF-β1-induced phosphorylation of Smad2/3 (Fig. 4C). These results suggest that ZA (Cmax) suppresses wound closure with granulation tissue in the oral cavity by the inhibition of the TGF-β1-induced migration of mesenchymal cells, such as hGFs by suppressing Smad2/3 phosphorylation by TGF-β1. On the other hand, Ozdamar et al (47) reported that the phosphorylation of polarity protein Par6 is required for TGF-β1-dependent epithelial-to-mesenchymal transition (EMT) in mammary gland ECs and controls the interaction of Par6 with the E3 ubiquitin ligase Smurf1. Smurf1, in turn, targets the guanosine triphosphatase RhoA for degradation, thereby leading to a loss of tight junctions (47). In addition, RhoA is generally known to be a regulator of cell migratory activity in various types of cells by promoting stress fiber assembly and the resultant cell adhesion (48). Intriguingly, serine-to-alanine point mutation [Par6(S345A)] of the TGF-β1-induced phosphorylation site in Par6 suppressed the TGF-β1-induced EMT in a Smad-independent manner. It remains to be clarified whether ZA (Cmax) suppresses the TGF-β1-induced migratory activity of hGFs through affecting Par6/Smurf1/RohA signal transduction pathway in a Smad-independent manner.
Saito et al demonstrated the negative effects of ZA on the re-epithelialization of oral mucosa in a three-dimensional in vitro oral mucosa wound healing model (49). They demonstrated histologically that ZA downregulated the expression of TGF-β type I and II receptors and Smad3 phosphorylation in oral keratinocytes. However, they used ZA at 10 µM, which was a much higher concentration than Cmax, for evaluating the effects on TGF-β1-induced signal transduction in human oral keratinocytes. Moreover, they did not elucidate how TGF-β-induced intracellular signals affect the function of human oral keratinocytes at the cellular and molecular levels. In this study, we demonstrated that ZA (Cmax) lowered the ratio of the number of cells strongly expressing TGF-β type I receptors on their surface against that of total hGFs, but did not affect the expression status of TGF-β type II receptors on hGFs (Fig. 4A). This finding suggests that ZA (Cmax) preferentially suppresses the expression of TGF-β type I receptor over that of TGF-β type II receptor on the surfaces of hGFs. By contrast, as described above, a high concentration of ZA (10 µM) seemed to downregulate the expression of both TGF-β receptor types (types I and II). In addition, we found that ZA (Cmax) suppressed the TGF-β1-induced phosphorylation of Smad2/3 in hGFs (Fig. 4C), implicating that the suppression of the TGF-β1-induced phosphorylation of Smad2/3 in hGFs by ZA (Cmax) may be caused by the inhibition of TGF-β type I receptor expression on the surfaces of hGFs. However, it remains to be clarified whether ZA (Cmax) directly suppresses the TGF-β1-induced phosphorylation of Smad2/3, or indirectly suppresses that through the inhibition of the TGF-β type I receptor expression on the surface of hGFs. On the other hand, Okamoto et al demonstrated that ZA (30-50 µM) induced apoptosis and S-phase arrest in mesothelioma by inhibiting the functions of Rab family proteins (50). The Rab family is generally known to control the endosomal trafficking of membrane molecules, such as growth factor receptors (reviewed in ref. 51). It is also generally known that Rab5, activated by a guanine nucleotide exchange factor (GEF), directs TGF-β-activated receptors into the endocytic pathway that promotes TGF-β-induced Smad2/3-dependent signals (52). In addition, Kardassis et al suggested that TGF-β receptors may be recycled on the cell membrane in a Rab-11-dependent manner following clathrin-dependent internalization of TGF-β receptors, which may positively affect the activities of Smad2/3-dependent TGF-β-induced intracellular signals mediated by these receptors (53). Our laboratory is currently investigating whether ZA (Cmax) suppresses the function of Rab family members that may upregulate the activity of TGF-β1-induced Smad2/3-dependent intracellular signals in hGFs at the receptor level, thereby resulting in downregulating TGF-β1-induced fibrogenic activity of the cells.
As described above, it has been reported that the TGF-β-induced activation of p38 MAPK positively regulates migratory activity (31). In fact, TGF-β1 upregulated the phosphorylation of p38 MAPK in hGFs; however, ZA (Cmax) did not suppress the TGF-β1 (5 ng/ml)-induced phosphorylation of p38 MAPK in hGFs (data not shown), suggesting that p38 MAPK activity is not related to the suppressive effects of ZA (Cmax) on TGF-β1-induced fibrogenic activity of hGFs. It is plausible that the suppression of type I receptor expression on the surface of hGFs by ZA (Cmax), as shown in Fig. 4A, may not be sufficient to suppress the TGF-β1-induced p38 MAPK phosphorylation. On the other hand, the activation of MAPKs such as JNK and p38 MAPK, but not ERK, is necessary for the progression of the hypoxia-induced MF differentiation of fibroblasts (54). Although, in western blot analysis, the phosphorylation of JNK was not detectable in hGFs before/after TGF-β1 (5 ng/ml) stimulation (data not shown), it remains to be clarified whether JNK affects the TGF-β1-induced fibrogenic and migratory activities of hGFs. In addition, the mechanisms through which p38 MAPK affects the TGF-β1-induced fibrogenic and migratory activities of hGFs also remains to be clarified.
In conclusion, in the present study, it was suggested that ZA (Cmax) attenuates TGF-β1-induced wound closure by inhibiting the formation of granulation tissue by hGFs stimulated with TGF-β1 that was derived from inflammatory tissue, possibly through the suppression of Smad2/3 signaling. Our findings partly clarify the molecular mechanisms underlying BRONJ and would benefit research into drug targets at the molecular level for the treatment of this symptom.
This study was supported in part by Grants-in-Aid for Scientific Research (KAKENHI) (grant nos. 24791981 awarded to M.I., 25463053 awarded to N.C., 26462823 awarded to S.K., 22592076 awarded to M.K., 26293426 awarded to T.S., 24593002 awarded to Y.S., and 26670852 awarded to A.I.) from the Ministry of Education, Culture, Sports, Science and Technology of Japan; Grant-in-Aid from the Dental Society of Iwate Medical University; and Grant-in-Aid for Strategic Medical Science Research Centre from the Ministry of Education, Culture, Sports, Science, and Technology of Japan, 2010–2014.
Komatsu, Y., Ibi, M., Chosa, N., Kyakumoto, S., Kamo, M., Shibata, T., Sugiyama, Y., Ishisaki, A."Zoledronic acid suppresses transforming growth factor-β-induced fibrogenesis by human gingival fibroblasts". International Journal of Molecular Medicine 38.1 (2016): 139-147.


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Q: node async function - get the result to the outside function I have the following async function and want to have the value data.
I tried with Promises and Async/Awaits. I always miss something. Would be happy for some help. Thanks!
Here the async function:
ad.getData("ID1", function (err, res) {
  let data = 11111;
  if ((res!= null) && (res.val != null)){
    data = res.val;
  } else {
    // store data
  }
});

and I tried:
async function getData(id){
  return await ad.getData(id, function (err, res) {
       let data = 11111;
       if ((res!= null) && (res.val != null)){
         data = res.val;
       } else {
         // store data
       }
         return data;
        })
}

let data = await getData("Id1");

console.log(data);

A: This is an asynchronous operation, and the code is written in callback style.
ad.getData("ID1", function (err, res) { // the function is a callback
  let data = 11111;
  if ((res!= null) && (res.val != null)){
    data = res.val;
  } else {
    // store data
  }
});

In order to get the result outside the function, or in general, write asynchronous code that look like a synchronous one, we can wrap callback style code in a Promise, like this :
function getDataAsync() {

  return new Promise ((resolve, reject) => {

  // callback style code is wrapped in the Promise
  ad.getData("ID1", function (err, res) {
    let data = 11111;
    if ((res!= null) && (res.val != null)){
      data = res.val;
    } else {
      // store data
    }

    resolve(data); // it means, when the Promise is resolved, it will give you the data
});

And you can get the output of getDataAsync like this :
let data = await getDataAsync("Id1");
// we use "await" keyword to wait for the Promise is resolve, data is the variable in the resolve
console.log(data);

You should use Promise instead of callback, and async/await will help you write Promise code easier.
Please take a deep look at https://scotch.io/courses/10-need-to-know-javascript-concepts/callbacks-promises-and-async. When you understand the concept, it's literally easy.

redpajama_set_name: RedPajamaStackExchange


Much of the documentation concerning RAF aircraft losses during WW2 no longer exists. When researching the details surrounding a specific event, it is largely a matter of chance as to whether any records survive - and down to the expertise of researchers to locate those surviving records!
resulted in the discovery of the AM1180 crash records. A copy of the aircraft AM78 maintenenance log was also uncovered.
The papers relating to the 'Court of Inquiry' that was convened 2 days after the crash were not discovered, and for some time the 756(c) initial report seemed to have been lost as well. In the end, a copy of that latter item was discovered in Canadian records (Sgt. McGregor was a Canadian), so we were grateful to obtain a copy of that, from the Canadian archives.
In fact, it appears that much of the detail from the missing documents has actually been entered, in summary form, onto the AM1180 form.
As a result, when added to the local press records from the time, together with a significant number of witness statements, and the additional records from RAF Hurn, we have been able to piece together a reasonably detailed idea, of the sequence of events that led up to the tragedy.
Left click to see the images in greater detail - remember to use your back button to get back to this page - or right click to open the image in a new tab).
A.M. Form 78 is the record card of each individual aircraft. This form shows the brief history of JP137.
It shows that Halifax Mk II serial no. JP137 was manufactured by the London Aircraft Production Company (part of London Transport!) as part of a batch of 250 aircraft, and that it had Rolls Royce Merlin 22 engines fitted.
It was delivered to the RAF from the manufacturers Leavesdon site in late November 1943. It then passed through various Maintenance Units ('MU') to be modified and 'kitted out' with equipment specific to it's forthcoming role, which was believed to be carrying out Special Operations Executive (SOE) 'drops' over Yugoslavia.
At the end of February onto No.1 Overseas Aircraft Preparation Unit (OAPU) at RAF Kemble, near Cirencester, and then to 301 Ferry Training Unit at RAF Lyneham in early March.
Finally, to No.3 Overseas Aircraft Dispatch Unit (OADU) at RAF Hurn on the 16th of March, 1944.
The next entry 'Dest. MAAF' shows that JP137 was intended to join the Mediterranean Allied Air Force, in North Africa.
Sadly, as we now know, it never arrived. The entry 'Cat E (burnt)' is the official way of describing a completely burnt out wreck. The final entry. 'SOC' (Struck Off Charge) describes the final removal from operational records.
The A.M. Form 1180 is the document that served as a common 'collection point' for other records and documents, that would be generated as a result of an RAF aircrash.
It is often the only record that survives, for many incidents from WW2.
We were fortunate ehough to obtain a copy of the AM1180 for JP137, and both sides of the form are shown here.
(Left click to see the images in greater detail - remember to use your back button to get back to this page - or right click to open the image in a new tab).
It also includes details of the incident location, and the name of the pilot.
Dotted around the edge of the document are a collection of boxes, some of which have crosses, ticks, and numbers referring back to other documents.
It is rather poignant to notice that it also includes the number of hours the pilot had flown - both in total, and for the aircraft type.
As you can read, Sgt. Evans only had 10 hours 'night solo' on a Halifax.
As we have discovered, the kind of problem he most likely encountered immediately after take off, would have been impossible to correct, even for an experienced test pilot.
So a 20 year old, with only a few hours, stood no chance, even though we believe that he was regarded as an exceptional pilot.
The reverse of the AM1180 gives witness detail of the crash, together with a brief description of the results of the Court of Inquiry held 2 days later.
This was of course quite impossible to verify - there were no survivors!
This is damning evidence. It is now known that the Halifax MKII aircraft had a serious tail design fault, which made the aircraft virtually impossible to fly under certain conditions.
The suggestion of a possible rudder failure, in such a short précis, indicates that this was a well known problem.
Indeed, significant amounts of data, secret at that time, had already been collected on the problem.
As a result, the subsequent MKIII Halifax aircraft was fitted with a seriously modified tail rudder!
Notice that the lowest box 'Blame and discip action' is empty. As it should be.
There were no survivors, and any undefendable suggestions of pilot error would have been totally inappropriate.
This fact became an important part of the project, as you will read in other notes.
The report form 765(c) shown below is not commonly located in these cases. It was a preliminary report on flying accidents not attributable to enemy action, and was usually prepared almost immediately after an incident.
As such, it often contained information that needed to be corrected later on, once further investigations - including a Court of Inquiry where appropriate - had been carried out.
The possibiblity of the 765(C) form being inaccurate is one of the main reasons given by the RAF for their general reluctance to release the form publicy.
Other organisations take a different attitude on these matters, and as one of the crew of JP137, Sgt. McGregor, was a Canadian, it was from the Canadian Authorities that our copy of the 765(c) form was acquired.
In fact, there is little extra information to be obtained from the form, in this case, but as the RAF suggested, it does contain some errors!
The first page suggests the incident took place at 'Kemborne Rd', for example.
One fact that was gratifying to read was the crew list. Although much careful research had been carried out to ensure we actually had the right names, this was the first official form we had seen, from the period, that listed all 7 names in one place.
Page 2 also states that there was a fire 'on the ground'. It makes no mention of a fire in the air, which seems most likely to have occured, based on witness reports.
Page 3 includes the interesting comment 'one wing was seen to drop'. This was of course, a significant comment, bearing in mind the subsequent events.
It is interesing to note that at this early stage (this report is dated 21.3.44 - the same day as the crash) it is reported that the aircraft 'crashed onto some trees'.
Not, as we now know, entirely accurate!
Why, you may well ask, did the RAF decide to prepare a report so early?
The answer, I suspect, was just to get the 'paper trail' started correctly.
There were all sorts of implications - for pensions, for civilian compensation claims, for probably a dozen other reasons - that required the documentation to 'be correct' (as far as possible) even at an early stage.


People die everyday. They die from car accidents and homicides and war and diseases. But one of these deaths can be prevented. One of these deaths doesn't have to happen. Heart disease is the leading cause of death in America, according to Medical News Today. At the top of the death list, heart disease may seem formidable, but it doesn't have to be, things can be done to educate people, to research, to save lives with better technology. This disease may seem foreign to the Leonardtown High School community, to but it doesn't have to be. Raiders, you can get involved with your high school Help A Heart Club.
Help A Heart aims to raise money for the American Heart Association, encourage heart health, and help "hearts" in the community through charity work. Within the club, students work together to make a difference well beyond their school community primarily through fundraisers and service events within the community.
The American Heart Association is the recipient of Help A Heart donations. The association aims "to improve people's health and reduce the occurrence of cardiovascular diseases and strokes." The American Heart Association is the nation's oldest and largest voluntary organization dedicated to fighting heart disease. Comprised of 22.5 million volunteers and supporters, the American Heart Association is able to fund innovative research and promote health education thanks to the donations of millions. Research and education conducted by the American Heart Association has saved lives after diagnosis and prevented the onset of heart disease.
The club hopes that events and fundraisers will not only allow the American Heart Association to advance medical treatment and education of potential patients, but will draw attention to the severity of the disease and the importance of heart health for all high schoolers.
Help A Heart is a rather new club being offered at Leonardtown High School. The club was started in the fall of 2015 by Amrik and Ramnik Gill, two members of the graduating class of 2016. Though the club found some success in donations with Popsicle and hot chocolate sales, it was never really able to diversify and expand its service and fundraisers. Karson Harris, current vice president and Leonardtown senior wants "to help the club get off its feet and do real things that help the American Heart Association." Members and leaders within the club have begun event planning and correspondence with the help of their sponsor, Julie Walsh, to achieve the goal of increasing service from last year.
Don't let all of the goals of Help A Heart deter you raiders, the club needs you to help them make a difference. Help A Heart is still in need of more members and is offering leadership positions, so make sure to join before it's too late. The club meets every other Friday in Mrs. Walsh's room, E11. If you have any further questions feel free to contact Sami Schwalbe or Karson Harris, the club's leaders.


INITION worked with the fuel marketing team at BP to produce their first virtual reality experience using the Samsung Gear VR. The experience allowed BP's stakeholders to take a journey following the oil from exploration through extraction to the end client.
The experience takes the viewer across vast areas of land and deep under the sea, narrating the key benefits of BP's processes in order to maximise efficiency in terms of current processes of oil extraction and also as they evolve in the future.
BP aimed to explain complex scientific and technical details in a visually attractive manner. Going beyond statistics and data, the virtual reality experience shows viewers what they would not usually see: the incredible work happening in remote locations over land and sea. By bringing the story of the journey to life, BP is able to connect the audience with the brand and resonate the company's message.
This experience was shown by BP in many different countries across the globe.
"For a while now we have tried to improve our physical 'reality' showcases/exhibitions by making them more immersive. VR provided the ideal way to deliver an immersive experience that is in a highly transportable and re-useable form.
The best thing about the virtual reality film is that almost 100% of our audience spend the whole time in the experience. This ensures that all of our message is communicated; a rare thing nowadays in today's busy world of advertising and communication.
The reception from the experience was fantastic. "Wow that was amazing" is by far and away the first phrase uttered, which says it all really.
using virtual reality ensures that all of our message is communicated; a rare thing nowadays in today's busy world of advertising and communication.


Revenge Is Teetering on Insanity. These 5 Things Need to Go. Now.
Lindsey U warned me that Revenge was soap-opera-level nuts last night, and still I was surprised by the sudden rabbit hole of craziness I found myself falling down.
I mean, Revenge is usually the good kind of ooh-crazy. But last night's ep was a tumble into, like, Cheri Oteri drugstore-lady crazy.
Everyone having twelve identities. Just kidding, the white-haired man's good! Just kidding, he's bad. Just kidding, deep cover! Just kidding, rogue! I mean... anyone can get away with anything if they have this many personas.
Ashley and Daniel. Ugh. Love you guys in real life, can't see your relationship end fast enough onscreen. I know where all this is headed, but their eventual betrayal-tinged split would have a lot more oomph if they'd ever had an ounce of chemistry in the first place.
This wedding thing. Is anyone else wanting this incredibly odd Conrad-Victoria wedding storyline over before it starts? Like any sane person, I want to look at the cast of Revenge in Tuscany, but I feel like we could have found an easier way to get there.
Remote-control revenge. Yes, we've discussed the ludicrous abilities of whale cams and such before. But it never bothered me until now: all of a sudden it feels like Emily is always behind the scenes, doing everything via clicker--iPads, ear buds, secret cameras. What, did she just get a new La-Z-Boy or something? I'd like to see a Revolution-Revenge crossover, in which the Hamptons loses power and Emily's forced to go back to full-time, face-to-face conniving.
SAD JACK. Guys. I cannot take anymore. Every time we go down to the docks it's nothing but moping and debts, debts and moping. Sad Jack's permanent sad Jack face reminds me of the dogs on that awful Sarah McLachlan ASPCA commercial. Enough is enough.
Are you maxing out on these Revenge habits, too? Or am I crazy and you think it's getting better and better?


text:
Thomas Grubb (4 August 1800 – 16 September 1878) was an Irish optician and founder of the Grubb Telescope Company.

He was born near Portlaw, County Waterford, Ireland, the son of William Grubb Junior, a prosperous Quaker farmer and his second wife, Eleanor Fayle.

Thomas started out in 1830 in Dublin as a metal billiard-table manufacturer.  He diversified into making telescopes and erected a public observatory near his factory at 1 Upper Charlemont Street, Portobello, Dublin.  As makers of some of the largest and best-known telescopes of the Victorian era, the company was at the forefront of optical and mechanical engineering. His innovations for large telescopes included clock-driven polar mounts, whiffletree mirror mounting cells and Cassegrain reflector optics. Later, the manufacturing firm changed its name to Grubb-Parsons in 1925.

Grubb helped build the famous telescope for William Parsons, 3rd Earl of Rosse, at Parsonstown (now known as Birr), County Offaly, Ireland.  One of his earliest instruments - the telescope for Markree Observatory in County Sligo in the West of Ireland, supplied in 1834 - was, until 1839, the largest refracting telescope in the world. It was used to sketch Halley's comet in 1835 and to view the solar eclipse of 15 May 1836.

Later he built telescopes for observatories worldwide, including Aldershot Observatory, Melbourne, Vienna, Madrid and Mecca and others.

Thomas died in 1878 in Monkstown, County Dublin, Ireland. He is buried at Mount Jerome Cemetery, Dublin, Ireland. He had married Sarah Palmer. Their youngest son was Sir Howard Grubb, who took over the optical business. Thomas Grubb's cousin, John Grubb Richardson (1813 - 1891) was a major Irish industrialist who founded the model village of Bessbrook.

References

External links

Grubb Telescope Company
List of Grubb telescopes

1800 births
1878 deaths
Irish inventors
People from County Waterford
Lens designers
Telescope manufacturers
Irish astronomers
Burials at Mount Jerome Cemetery and Crematorium
Fellows of the Royal Society


Law firm Gordon Dadds Group has struck a £34m deal to acquire rival practice Ince & Co International which will create the largest listed law firm in the UK by revenue.
The newly merged practice, which will trade as Ince Gordon Dadds and be one of the largest 40 UK law firms, will have combined revenues of over £110m with 100 partners and offices in nine countries.
The London-based group will be led by Gordon Dadds' CEO, Adrian Biles who said Ince is a "highly successful and well-respected business with an iconic brand".
"The merger will build upon the complementary strengths of the two firms in terms of industry expertise and range of services. Our management model will also allow Ince's partners and fee earners to focus even more on providing market leading legal advice to a stellar client base," he said.
Peter Rogan, chairman of the board of Ince, said the merger would enable the firm to extend its offering to client offering and invest further in its people.
Ince is a network of affiliated international commercial law firms, serving its global client base from offices across Asia, Europe and the Middle East. It focuses on four core sectors - transport, trade, energy & infrastructure and insurance.
The price paid will equate to a percentage of the turnover generated by the equity partners of Ince & Co International over the three years from completion.


text:
Welcome to The Venice West! Our venue is the culmination of a few longtime local friends who love live music and grew frustrated with having very few places on the west side that could deliver on a consistent basis. After many late nights and missed shows, we decided to stop complaining and do something about it. Five years (and a pandemic) later — after a long and twisty road — the project finally came to fruition and TVW was born.
"Venice West" was coined to describe the lesser known "Beat" scene in Venice and surrounding areas during the late 1950s and early '60s. This era helped define Venice as a global brand known for its cultural diversity, eccentricity, and expression through music and the arts, where places like "The Gas House" and "Venice West Cafe" created a haven for poets, musicians, and eccentric locals to share their words, sounds, and creativity.
There are a number of moments in time that contribute to Venice's storied history, but the "Beat Generation" certainly played a big part. We chose our name as a nod to this defining time and place in our town's unique past and to help keep some of the qualities that Venice is celebrated for alive and flourishing for years to come.
The Venice West is about community. Our goal is simple: create a welcome place for music fans and artists alike to enjoy consistently great food, music, and good times. We are humble in our approach and appreciative of your feedback so please don't be shy and let us know what you think. Thank you and we hope you enjoy the show!
-Rob, Allen, and The Venice West Team
Hand-Cut Garlic Parmesan Fries
@TheVeniceWest
1717 Lincoln Blvd. Venice, CA 90291