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Contributing Voices
Medical Directors Explain Hows and Whys of Burnout
These high-achievers wrestle with competing interests as they operate both in the medical and business worlds
Frank Diamond
After 13 years in neonatal intensive care, Marjorie Schulman, MD, became a clinician executive, working as a medical director. She left Aetna at the end of 2011. She loved it there, calling it an ethical company that has done more than any other in the industry to allow work-life balance. Still, there was pressure, and balance in the lives of medical directors can be difficult to come by.
On her first day at Aetna, one of Schulman's new physician colleagues pulled her aside.
"'Your specialty is neonatal intensive care, right?'
'Yes,' I answered.
'Well, you are going to see a lot of hysteria in this industry. Everything is urgent, urgent, urgent when money is on the line. But remember, nothing in business is ever like medicine, especially the kind of medicine you practiced.'"
Hidden problem
Medical director burnout often goes unseen and undetected. Most professionals, not just medical directors, don't want to admit to burnout; it took a while to get anyone to speak to us about the problem. The attitude was "Medical director burnout? Never heard of it! Especially at my company!" Even an interview with a physican who works for one of the large consulting companies, someone you'd assume would be forthcoming, went nowhere.
A spokeswoman for America's Health Insurance Plans advised us that we should instead be writing about how excited medical directors are to go to work every day to face the challenges of the health reform era, about how much they love their jobs. While we have no doubt that most do, to suggest that medical director burnout doesn't exist pulls the reach of credulity out of its socket.
"The problem is quite real — not just for medical directors, but for any physician in a management role," says Brent James, MD, the chief quality officer and executive director of the Institute for Health Care Delivery Research at Intermountain Healthcare. "We tend to be really compulsive and over commit. The problem afflicts all ranks of management."
Despite appreciating her career, Schulman says that she hasn't met many medical directors who don't burn out at some point. Still, if she had it to do over again, she'd still become a medical director.
"My hope when I entered the health insurance field was to influence population health at the place where money drives clinical behavior," says Schulman. "I have been lucky to see that up close."
Now's the time to define terms. Managed care medical directors provide clinical leadership and expertise related to health insurance programs including — but not limited to — quality improvement, utilization management, network design, policy development, accreditation, credentialing, care delivery, and working in a team/collaborative relationship with other organization leaders, says Jaan Sidorov, MD, a former medical director at Geisinger Health Plan and a member of Managed Care's editorial board. Medical directors typically report to the chief medical officer.
Burnout is not a recognized psychiatric disorder, so there is no diagnosis, says Philip R. Muskin, MD, professor of clinical psychiatry at Columbia University and a distinguished life fellow of the American Psychiatric Association.
Sad every day
"Someone with burnout might be sad, or might be sad enough to qualify for a diagnosis of depression that has a main symptom of a pervasively sad mood for most of the day, every day, or significant loss of interest in usually enjoyable activities, plus other symptoms," says Muskin. "The person might be anxious and might be anxious enough for a diagnosis of generalized anxiety disorder that has a main symptom of worry. I would not want anyone to think he or she has a mental disorder if experiencing burnout."
Also, while someone might call himself a "burnout specialist," there is really "no such specialty," says Muskin.
Robert Forster, MD, is a health care consultant who has held various positions in his career, including chief medical officer (CMO) at BlueCross BlueShield of Florida for seven years. The medical director's role is far too restricted and inflexible, he says. "They are really not treated as a professional, if you will. You might as well put information in a computer and not have any judgment."
"Nearly all medical directors want to be on a professional advancement track, but that is not always possible," says Abbie Leibowitz, MD, of Health Advocate.
Abbie Leibowitz, MD, agrees. He is a former chief medical officer at Aetna and now is executive vice president and chief medical officer of a consumer advocacy company called Health Advocate. "Many medical director roles are pretty repetitive and don't require a lot of creativity or encourage much strategic thinking," says Leibowitz. "This is especially true of the utilization review jobs — a common entry point in many plans. Nearly all medical directors want to be on a professional advancement track, but that is not always available."
Likes going to work
One medical director, who asked not to be identified, recently changed jobs, moving from a regional to a national plan. An 11-year veteran, she now finds herself in an unusual state: She likes going to work. Not that it was torture before. "In the very beginning it's interesting because it's a learning curve, but after a while it just ends up becoming tedious," she says.
Some plans want medical directors to be involved in many extracurricular activities, such as meeting with specialty groups, serving on multiple internal committees, visiting practitioners and hospitals, "all the while being responsible for time sensitive turnaround cases without coverage from your peer physicians, which creates a huge amount of stress and job dissatisfaction.
"At the end of the day we're still responsible for the basic widgets. That is more highly valued. And if you do external activities, that is an addition, not a substitution. So there's less and less incentive to want to do outside activities."
Would more money help? "It's not really the compensation; it's more that for the most part, your basic activities take eight hours, so I would go to an external meeting, get back at 3 p.m., and find that I was going to be working until 8 p.m. to finish my core job."
Forster points out that medical directors' performances are often judged by nonclinical people. "Ultimately, as they move up the ladder, there's a non-clinician above them who has a difficult time grasping the subtleties."
Forster is a mentor to medical directors, and most of what he teaches has to do with explaining medical information in terms that nonclinician executives can understand. "You don't necessarily talk about the quality benefits that you can sustain by doing something in this method but about what it can bring in terms of dollars, in terms of saving the plan money, or allowing the plan to reduce premium and therefore become more competitive. Sometimes you have to talk their language."
Familiar symptoms
Thomas J. DeLong, PhD, points out that medical directors are high performers and that they burn out the way high performers in most professions burn out. They are addicted to achievement and sometimes become self-absorbed. "They don't understand how their behavior affects others," says DeLong, the Philip J. Stomberg Professor of Management Practice at Harvard Business School. "I believe, based on the interviews that I've done, that it is an addiction — as much of an addiction as being on prescription drugs or on heroin. It's that hard to break."
Physicians who become businesspeople might feel divided. The positive medical part is "being with the patient and forming an intimate relationship and seeing the patient get better," says DeLong. "But once you are a medical director, it becomes more about policy and procedures and numbers. That is degenerative. It doesn't refill the vessel by giving a feeling of growth and renewal and some sense of peace."
In the world of practicing physicians, corporate medical directors are viewed as the enemy, not as peers, which can be very depressing, says Schulman. "It is still the rule that practicing physicians carry stereotyped views of medical directors as paper pushers, care deniers, and little more than claims processors. Every medical director can cite dozens of incidents when network doctors or their former colleagues in practice accuse them of having gone over to the dark side."
Arlen Collins, MD, president and chief science officer at Remedia, a company that addresses problems of overuse and misuse of medications, has worked as a medical director at US HealthCare and also — as a chief medical officer — turned around several ailing health plans. He cites competing interests. "I was on a call the other day with some folks at a major national health plan and they invoked the lawyers at least six times as an impediment when discussing what everyone on the call agreed would be a real positive move for the plan and its members. Increasingly there are regulatory compliance issues that impede progress that drive many medical directors nuts."
He never experienced burnout, he says; his US Healthcare experience was constantly exciting and with the turnarounds, he was too busy trying to fix companies. "These plans were in such extremis that no one bothered me and I was able to do what needed to be done."
Marla Tobin, MD, a recently retired senior medical director at Aetna, thinks that burnout is rampant in the medical director community, especially among boomers.
Marla Tobin, MD, a recently retired senior medical director at Aetna, says that "physician burnout in the medical community is rampant — especially in the boomers." (See "Burnout Rooted in the System.")
The medical director community sees burnout, too, "and it often involves two main issues — task work and travel. Especially with the utilization review work and completing tasks in the queue, often physicians get into a production mode and have little personal contact or job satisfaction, as their feedback loop is often negative." In addition, she says, the peer-to-peer interaction involved in denials can get nasty.
Leibowitz contends that peer-to-peer review, where denials for coverage are examined and sometimes challenged, runs much more smoothly if the treating doctor or a specialty expert discusses the case with the medical director. "At Health Advocate we arrange these frequently to help our members. It is not common that the discussion between a treating physician and the plan's medical director gets confrontational, but as I am sure one can appreciate, that can happen."
Though the focus in this article is on medical director burnout, medical directors need to be able to spot it in their subordinates as well. Then it's time to have a talk, says DeLong.
"One thing I do is to say to a subordinate [showing signs of burnout], 'Number one, what's the desired outcome? Number two, how will you know when you've achieved success? Number three, if you were to throw a celebration in five years, what would you celebrate? How many people are you helping along the way other than just yourself?'"
Can be managed
DeLong says that there is hope, though it might be hard-earned. "I have seen 35-year-olds, 45-year-olds, and 55-year-olds make dramatic changes in managing this need for achievement and this overdrive. But you must manage it every day. This is not a problem that you fix and then do not think about."
More to burn out over
Medical directors once again find themselves in the position of gatekeeper, as techniques used and nearly abandoned in the 1990s in the managed care backlash make a comeback. Abbie Leibowitz, MD, says they include limited networks, step therapy, and prior authorization.
"That is an added burden and source of stress for medical directors," says Leibowitz, a former chief medical officer at Aetna and currently executive vice president and chief medical officer at a consumer advocacy company called Health Advocate. "They are on the front lines of setting these [systems] in place and then managing through the confrontations and issues that inevitably result."
Increase in claims
The American Medical Association's 2012 National Insurer Report Card shows a 23 percent increase in the number claims that require prior authorization — something that we featured in last month's Compensation Monitor department (http://www.nxtbook.com/nxtbooks/medimedia/managedcare_201208/index.php?startid=14).
A study by Athenahealth, a vendor that sells EHR and care coordination technology and services, shows that while the number of prior-authorization requests has gone up in recent years, the percentage of denials has gone down.
More haggling, more stress
Source: Athenahealth
How to spot it
Philip R. Muskin, MD, defines burnout as a special type of job stress. "It might be described as a state of physical, emotional, or mental exhaustion combined with doubts about the person's competence and/or the value of the person's work," says Muskin, professor of clinical psychiatry at Columbia University and a distinguished life fellow of the American Psychiatric Association.
Signs include feeling cynical or critical while at work, feeling that you have to drag yourself to work, becoming irritable or impatient with others, and becoming disillusioned with the job.
The main symptoms of burnout, says Marjorie Schulman, MD, a former medical director, are negativity and cynicism, often directed at the bureaucracy of one's own company. "In the case of medical directors who perform medical management, it is also easy to become very jaded and angry at the network physicians who are supposed to be one's peers. This is often because physicians can be extraordinarily rude and dismissive to medical directors when they are contacted."
Intense depression could be a sign that burnout either caused, or is caused by, the psychiatric disorder, says Philip R. Muskin, MD.
"Some people may notice a drop in their productivity or experience little satisfaction from realistic achievements," says Muskin. "Changes in sleep or appetite or new and/or unexplained physical symptoms such as headaches, backaches, or general aches and pains might occur. When the person notices he or she is using alcohol, food, prescription drugs, or illicit drugs to feel better or to create a sensation of not feeling at all, that is a serious warning sign that something is wrong and that intervention is necessary."
A classic sign of burnout is overreaction — to almost everything. "When something happens, you take it very personally," says Thomas J. DeLong, PhD, the Philip J. Stomberg professor of management practice at Harvard Business School. "There's a spillover effect in your private life where you find yourself being more curt and more judgmental, and you listen less. You find fault in others faster. You can't self-monitor."
Successful people are usually smart and welcome an overloaded agenda, says DeLong. "You define yourself on what you've accomplished, not on what kind of person you are. You define yourself based on how many things you cross off your to-do list."
Certain professions are predisposed to have such people in them, says DeLong; surgeons, professional athletes, and investment bankers for instance. The person knows something isn't right.
Less fun
"The fun is less," says Muskin. "Don't sweat the small stuff? Suddenly the small stuff looms much larger. There's a change in your behavior — inability to control your irritation with somebody. If you never had road rage, now you have road rage. You notice changes in yourself but not changes for the better. You notice that the creative stuff that got you where you are isn't functioning the way it should be."
What to do about it
Richard Rosen, MD, believes in the restorative power of sabbaticals. Rosen is retired and does some consulting work for the National Committee for Quality Assurance. In 1975, he began working for Rhode Island Group Health, which in the mid-1980s was bought by Harvard Pilgrim Health Care and became Harvard Pilgrim Health Care of New England.
Change of pace
"We created a sabbatical system at Harvard Pilgrim Health Care of New England," says Rosen. "Several physicians took advantage of the opportunity to have a change of pace and to take advantage of educational and travel opportunities."
Harvard Pilgrim Health Care of New England closed its doors in 1999, but the Harvard Vanguard Medical Group that contracts with today's Harvard Pilgrim Health Care still offers sabbaticals for physicians, says Rosen. "I know that many large groups have sabbatical plans, especially those that are part of academic university hospital medical centers. This allows for academic expansion, research, and development of publications. For many of us, it helps to prevent burnout."
Rosen recalls a trip to Russia with a group from Jewish Hospital of Louisville to be part of a medical exchange team for the Good Will Games in St. Petersburg. "We spent 10 days between Moscow and St. Petersburg, touring a bit and meeting with medical staffs at various facilities," says Rosen.
Energized
The next ten days "my wife and I went to Scandinavia, where I had a day tour of the Karolinska Institute, and we traveled all over Finland and Lapland, Sweden, and Norway in a bit of adventure and vacation. When we returned home, I spent the next three weeks doing intensive medical quality reviews for NCQA."
Thomas J. DeLong, PhD, the Philip J. Stomberg professor of management practice at Harvard Business School, says the change of pace need not be as extreme as a sabbatical. "Here's what helps: Stop and reflect. Make sacred time — an hour or two hours or three hours during the week where there are no PDAs, no telephones. Where you are having conversations with the most meaningful people in your life.
"That is not happening for most people, but if they were to do that, it would make a huge difference.
"Make sacred time — an hour or two hours or three hours during the week where there are no PDAs, no telephones. Where you are having conversations with the most meaningful people in your life."
"Take 10 minutes every day and write down five things that you are grateful for. It slows you down so that you can reflect. That is the first stage in individual change."
DeLong describes this as stopping the hamster wheel. "The trap is that one of the ways that high-achieving personalities try to solve the problem is to get busier and busier. That is a way to distract. It sets them up for some real problems later."
Philip R. Muskin, MD, agrees that taking time for yourself is crucial. "No lunch at the desk," says Muskin, professor of clinical psychiatry at Columbia University and a distinguished life fellow of the American Psychiatric Association. "Take a walk, go to the gym, listen to music. Learn a deep breathing or relaxation exercise to do during the day a few times."
Burnout rooted in the system
Burnout comes with the territory not only for medical directors, but for physicians as well, according to a study that made headlines last month. Published in the Archives of Internal Medicine, "Burnout and Satisfaction With Work-Life Balance Among U.S. Physicians Relative to the General U.S. Population," states that nearly half of doctors suffer from burnout. Burnout is not a recognized psychological disorder but, as the study's main author Tait D. Shanafelt, MD, points out, its ICD-10 code describes it as "Problems related to life-management difficulty." (It's an experience not a disorder.) Burnout among nearly 7,000 doctors is measured using the Maslach Burnout Inventory, a 22-item questionnaire. Authors describe it as "a syndrome characterized by a loss of enthusiasm for work (emotional exhaustion), feelings of cynicism (depersonalization), and a low sense of personal accomplishment."
Burnout rates
The study looks at various specialties, finding substantial differences in burnout rates. "Emergency medicine, general internal medicine, neurology, and family medicine had the highest rates of burnout, whereas pathology, dermatology, general pediatrics, and preventive medicine (including occupational health and environmental medicine) had the lowest rates."
One of the concluding points: "The fact that almost 1 in 2 U.S. physicians has symptoms of burnout implies that the origins of this problem are rooted in the environment and care delivery system rather in the personal characteristics of a few susceptible individuals."
That's something many medical directors can probably agree with.
Burnout by specialty
Source: "Burnout and Satisfaction With Work-Life Balance Among U.S. Physicians Relative to the General U.S. Population," Archives of Internal Medicine, Aug. 20, 2012
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Albinykus  — род тероподовых динозавров, принадлежащих к группе альваресзаврид и живших во время позднего мела (около 85—70 млн лет назад, сантон и кампан) на территории нынешней пустыни Гоби в Монголии. Окаменелости теропода были найдены в формации Javkhlant Formation в аймаке Дорноговь. Впервые описан группой палеонтологов: Sterling J. Nesbitt, Julia A. Clarke, Alan H. Turner и Mark A. Norell в 2011 году. Представлен одним видом — Albinykus baatar.

По кладистическому анализу Albinykus был помещён в группу альваресзаврид как сестринский таксон к другому виду: шувуйя из группы Parvicursorinae. Albinykus является одним из самых маленьких известных альваресзавридов. Этот динозавр настолько мал, что указывает на тенденцию уменьшения размеров тела в этой группе. Такая тенденция является единственным известным среди теропод и группы Paraves.

Примечания

Динозавры мелового периода
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Doom, Marvel, marvel comic book reviews, Marvel Comics
Doctor Doom #6 Review
Writers: Christopher Cantwell
Artist: Salvador Larroca
Colors: GURU eFX
Lettering: VC's Cory Petit
Cover: Salvador Larroca & GURU eFX
Release Date: March 4th, 2020
Reviewed by: Steve Baum
Doctor Doom and Kang the Conqueror have been twisted together throughout this series, and no one, including them, seems to know why. Not quite adversaries (for the moment) but not trusting each other either, all we can do is wait to see how this chronological chess match plays out. Doom gets his hands on some pretty serious weaponry at the conclusion of this issue. But that's just so he can use it for good… right?
"DYLAN!!! You sooonnnn ova BITCH!" (in Arnold Schwarzenegger accent) If that wasn't your first thought looking at this cover, then I pity your poor, poor childhood. Locked in biceps flashing action just like Arnold and Weathers (it's Predator if anyone still isn't following), Doom and Kang are poised for a face-off. But they've been getting along so well!
Poor Tom, the guy just wants to share a train car, and for his troubles, he gets launched through the roof 200 feet into the air. Just when you're starting to think Doom is turning a corner and acting like a nice guy… RIP Tom. As our duo continue their trek, some poor redneck pair makes the mistake of "joking" with them. Doom incinerates them in green cosmic energy. There's the Doom we know and love! Does it feel like nothing substantial has happened yet? Ok, as long as it's not just me. There is some comedy inherent in watching Doom and Kang 'Planes-Trains-and-Automobiles' it across Texas, and we get a rare moment of levity on the boat immediately followed by the two assuring each will destroy the other. Insult of the day goes to Kang for "toadfish romantic with a corkboard spine"; I might steal that one. After finally getting some action in the form of a brief and feeble ambush attempt, we're introduced to a fun new weapon that saps willpower and energy. Orb (the giant eyeball) is left questioning his life choices and what he's doing there.
Not a bad weapon for sure. But we need something better! Cue the grown-up Ash Ketchum dressed as a 'Weekend at Bernie's' extra delivering what even Galactus fears. That's right; the Ultimate Nullifier just showed up. Kang predictably turns on Doom, seeking the Nullifier for his own. Doom puts him down prison-shank style and makes a call to his bestie, Reed Richards. He has an idea for how to stop the black hole, but Blue Marvel isn't going to like it.
The art offers no complaints. The colors range from bright, harsh deserts to dark, moonlit canyon rivers. Doom and Kang are drawn to the character and are imposing at a distance and up close. If I had to nitpick, Doom's face just didn't seem quite right.
There didn't really seem to be much to this issue. Even though it elicited a few chuckles on my end, the banter between Doom and Kang and the vagrants they come across (and subsequently dispose of) are cookie-cutter. The last few pages seem to be all that matter and have any real ramifications. It's nice to see the Blue Marvel get some attention, he's a wildly powerful and drastically underutilized character in the Marvel universe. In the flow of this series, it's not a bad issue, just filler. In fairness, they can't all be 10s. Overall, I've enjoyed this series and will continue to follow it, but this issue just didn't seem to accomplish anything save for the last few pages. That's not to say that every panel has to be filled with world-threatening calamity and non-stop action, but there are more exciting or interesting ways to accomplish what this issue accomplished.
dispatchdcu March 6, 2020 Comic Book Reviews, comic books, comic reviews, comics, doctor doom, Doctor Doom #6, Doctor Doom #6 Review, Kang, Marvel Comic Reviews, marvel comics reviews, marvel reviews, review, Reviews
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The True Frat And Sorority Experience
Spoiler Alert: Not Like Those College Movies
By: William Morton
(Note: Article is written in May 2017)
The prestige of being in a fraternity or sorority is security that is near perfection. Sporting those Greek letters on clothing is the equivalent of feeling like royalty. Though the terms "fraternity" and "sorority" differ by gender, they both provide extracurricular education.
At the College of Staten Island, the frat and sorority representation glows with the confidence of students who value their education, and love interactions between their organization and the community. The fact that certain politicians and entrepreneurs are alumni of Greek organizations is no coincidence. To name some of the Greek organizations at the College of Staten Island, there is the nationally known brotherhood of Tau Kappa Epsilon and the longstanding sisterhood sororities of Delta Delta Omega and Chi Iota Omega. They boost up the vibe of the school.
Television programs like Blue Mountain State and a plethora of movies on Netflix make fraternities and sororities seem like they are irresponsible party enthusiasts that don't have respect for their peers. To the Contrary, Greek organizations, especially the ones at the College of Staten Island, balance researching in the library with volunteering at daycare centers.
Of course there are parties, but they are for charities such as St. Jude Children's Research Hospital and the protection of female rights; the motivating factor in the latter for a fraternity like Tau Kappa Epsilon being that they treasure acts that display gentlemanly characteristics.
Emmanuel Gbogboade, one of the most decorated members of the Upsilon-Lambda chapter of Tau Kappa Epsilon located at the College of Staten Island, confirmed the scholastic approach of Greek organizations by saying:
"The stereotype of frat and sororities is that Greek organizations are lazy and binge on alcohol, when in actuality they arrive to fields and buildings early to host philanthropy events. The experience of being in a fraternity and sorority is something often searched for but not often experienced, and it's the only opportunity in college that makes you feel connected to something."
Ask someone in Delta Delta Omega about someone in Tau Kappa Epsilon, and they are probably going to talk about how they found love at first sight or how they know they can rely on each other for comfort through humor and laughter. Sororities such as Chi Iota Omega and Delta Delta Omega have inspired Greek organizations around school to step up their efforts in enhancing and beautifying the college experience.
Though the prestige and pride of Greek organizations makes it seem as if they socialize at fancy mansions and experience things in luxury, the fact is they remain humble by fraternity brothers singing karaoke in the car and sorority sisters posting makeup tutorials on YouTube and Facebook. Though the administration at the College of Staten Island is hesitant to recognize the contributions of fraternities and sororities, there are scholastic forums such as the school radio station WSIA 88.9 FM and school publications such as The Banner in which both alumni and active members have found their niche. There are terminologies utilized by Greek organizations and stories that have connectivity to those who understand it.
It's like those postcards sent from someone on vacation that say "Wish You Were Here!" The love expressed in Greek organizations can be seen by others and can make miracles happen.
Tagged as: Frats, Greek, Greek Life, Lifestyles, Sororities
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Researching Cannabis for Therapeutic Purposes: The Potential Promise of Cannabidiol (CBD)
Three weeks ago I had the opportunity to testify before the Senate on cannabidiol, or CBD -- one of the main active ingredients in the cannabis plant, and a compound around which there is a growing amount of interest from the public as well as the scientific and medical communities. The intent of the hearing was to explore ways we can reduce obstacles to research on CBD, given that all chemicals in the marijuana plant are currently in the most restricted category of the Drug Enforcement Administration's schedule of controlled substances -- schedule I. This category is reserved for substances that have a high potential for abuse and no scientifically supported medical use. Medical research can and is being done with schedule I substances; however, there are strict regulations and administrative hurdles associated with this status.
Image Courtesy of NIDA
The chemical in cannabis that causes the high (and many of its other effects) is delta-9 tetrahydrocannabinol, or THC. But there are over 100 other cannabinoid chemicals in the plant; CBD is one of those. Different cannabinoids can have very different biological effects; CBD, for example, does not make people high and is not intoxicating (please see Bergamaschi et al. 2011). And, there is reason to believe it may have a range of uses in medicine, including in the treatment of seizures and other neurological disorders. In the impassioned, often emotional debates these days over medical marijuana, CBD is often cited as one of the main reasons restrictions on marijuana should be loosened.
A basic understanding of the way cannabinoids affect the body and the brain is helpful to making sense of some of the confusing and contradictory claims made by partisans on either side of those debates. Most cannabinoids (including THC) interact with specific targets on cells in the body, the CB1 and CB2 receptors. CB1 receptors are found mainly in the brain and are important for learning, coordination, sleep, pain, brain development, and other functions; CB2 receptors are found mostly in the immune system. The fact that chemicals in marijuana can speak the body's chemical language (and cause both beneficial and harmful effects) is not surprising and does not make marijuana special. Many plants, including the opium poppy, tobacco, and coca are similar in that regard. Extracting and amplifying the medicinal benefits of such plants and minimizing their potential harms can lead the way to effective medications, but are also a major scientific challenge.
In marijuana's case, future medicinal uses will most likely lie in drugs based on cannabinoid chemicals or extracts with defined concentrations that can be reliably produced. There are very few precedents for a whole plant being approved as a drug (only two "botanical drugs" are currently approved in the U.S.). This is because creating a standardized plant medicine poses major difficulties -- especially in a case like marijuana where there are countless, widely varying strains and large numbers of chemical components about which little is yet known. Marijuana has over 500 chemicals in total, including the 100 or so cannabinoids, so we will still be learning about this plant for years to come. However, we are quickly learning a lot about THC and CBD, and because of its unique properties, CBD, especially, may be poised to make great inroads into our pharmacopoeia.
CBD has very little effect on CB1 and CB2 receptors (please see Pertwee et al. 2008). This is probably why it does not make people high and is not mind-altering; in fact, it may even blunt some of THC's psychotropic effects. Most marijuana grown for recreational use is very low in CBD content (and high in THC). CBD's actions in the body are not well understood, but they appear to involve several signaling systems besides the CB receptors, including a serotonin receptor.
Although CBD first rose to public prominence two years ago as a result of a CNN piece on medical marijuana, CBD has been considered as a potential therapeutic agent since the 1970s, when its anti-seizure properties were first examined in animal studies. A few very small randomized clinical trials were also conducted in adults with epilepsy, some showing positive results. But what has generated the renewed interest and excitement are the anecdotal reports that some children with otherwise untreatable severe epilepsies respond well to CBD extracts and oils (please see Porter & Jacobson 2013; Hussain et al. 2015). A followup to the CNN piece depicted the struggles of parents of children with a seizure condition called Dravet syndrome, who moved their families to Colorado in order to get CBD-rich marijuana legally. Several drug companies have recently developed CBD-based medications, and one of these, called Epidiolex, is now being tested in clinical trials. Epidiolex was developed by GW Pharmaceuticals and has been given to more than 400 children under the FDA's expanded access ("compassionate use") program.
While the existing data show promise, it is still too soon to tell whether and for whom CBD will be effective (please see Welty et al. 2014). Like most medical treatments, it doesn't seem to work for everyone. Much more research needs to be done, but it should be done quickly. Parents can't wait -- their children are growing; their children's brains are actively developing; and the repeated seizures may be damaging this growth. Parents currently must navigate states' medical marijuana laws or defy the law to obtain CBD extracts -- which, in the current unregulated market, may be of questionable quality and content.
Besides the trials of Epidiolex, there are already many studies of CBD for other disorders being conducted, including at NIH. Studies related to its possible use in the treatment of substance use disorders are being funded by NIDA, while other NIH Institutes are funding work on the potential of CBD and other cannabinoids in treating neurological and psychiatric disorders, disorders of the immune system and metabolism, and cancer.
In short, CBD appears to be a safe drug with no addictive effects, and the preliminary data suggest that it may have therapeutic value for a number of medical conditions. Addressing barriers that slow clinical research with CBD would accelerate progress. NIDA will do what we can to address such barriers and expedite the study of this potentially valuable compound, as well as other components of the marijuana plant.
For the latest updates, follow the National Institute on Drug Abuse on Twitter @NIDAnews.
http://www.huffingtonpost.com/nora-volkow/cannabidiol_b_7834066.html
If you want the right to grow your own cannabis and to be able to obtain cannabis legally here in Australia then please SHARE. #BringBackCannabisMedicine
Drug driving charge thrown out in landmark case
Firefighters stumble across a huge cannabis crop as they fight horror bushfires in New South Wales
Cannabis compound could be weapon in fight against superbugs
Canberra's cannabis laws do not address supply problem, meaning buying the drug will remain illegal
Thais to have access to permanent official cannabis clinics
US: Illinois recreational cannabis sales bring in $3.2 million on day one
US: Illinois governor clears more than 11,000 cannabis convictions
NZ: Cannabis spray Sativex brings mobility back for Taupo toddler with tumour Elyse Johnson
'They're in desperation': Canberra women breaking the law to treat endometriosis pain with cannabis
Italy court rules home-growing cannabis is legal in landmark decision


text:
module Rack
  class GithubWebhooks
    VERSION = '0.5.0'
  end
end

redpajama_set_name: RedPajamaGithub


Newman Insurance offers commercial & personal services in FL. As an independent insurance agency, we have been representing dozens of insurance companies. Officers, Crime, Bonds, Automobile, Homeowners, Life, Health and Long Term Care. We're conveniently located in Hollywood, Florida. Current Weather.
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Assais-les-Jumeaux ist eine aus mehreren Weilern (hameaux) bestehende westfranzösische Gemeinde mit  Einwohnern (Stand ) im Département Deux-Sèvres in der Region Nouvelle-Aquitaine. Sie gehört zum Arrondissement Parthenay und zum Kanton Le Val de Thouet.

Lage
Assais-les-Jumeaux liegt etwa 33 Kilometer ostsüdöstlich von Bressuire und etwa 21 Kilometer nordöstlich von Parthenay. Umgeben wird Assais-les-Jumeaux von den Nachbargemeinden Airvault im Nordwesten und Norden, Moncontour im Nordosten, La Grimaudière im Nordosten und Osten, Craon im Osten, Thénezay im Südosten und Süden, Pressigny im Süden, Le Chillou im Süden und Südwesten sowie Saint-Loup-Lamairé im Westen.

Geschichte
1973 wurden die bis dahin eigenständigen Gemeinden Assais und Les Jumeaux zusammengelegt.

Bevölkerungsentwicklung

Sehenswürdigkeiten
 Tumulus de la Motte de Puytaillé, unklare Datierung, seit 1970 Monument historique
 Kirche Saint-Martin in Assais, Monument historique seit 1929
 Kirche Saint-Martin in Les Jumeaux aus dem 19. Jreahrhundert

Weblinks

Ort in Nouvelle-Aquitaine


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Kumu Kahua's Ua Pau: It is Finished, Over, Destroyed
By Noe Tanigawa • Aug 16, 2019
(l) Playwright Alani Apio with Uncle John Kaulupali and Lucky during one of Hui Aloha's cleanups of Kaka'ako Park. The cleanups happen every Saturday, 10am, all are welcome to participate.
Credit Noe Tanigawa / Hawai'i Public Radio
A new play opening in Honolulu links history, family, culture, and modern life in Hawai'i. It's the final episode in a trilogy that started 25 years ago, and has made some waves along the way. The author agreed to an interview in Kakaako park, well aware that many of the issues dealt with in the play come to a head in the park daily.
Maile Kapua'ala plays Stevie, the returning college student, in Alani Apio's new play, 'Ua Pau: It is Finished, Over, Destroyed.
Credit kumu kahua / Hawai'i Public Radio
Interviews about the Kamau Trilogy
Playwright Alani Apio discusses his work, friends, and sources of strength.
Ua Pau premieres at Kumu Kahua Theatre August 22nd, and runs through September 22nd, 2019.
The first part of Alani Apio's Kamau trilogy, debuted in 1994. It is the story of how Alika chooses to make a living as a tour guide after his family's fishing grounds are bought by a hotel. In the second play, Alika's cousin, Michael, returns to the family after a Malcolm X-style education on Hawaiian culture in prison.
I went to Kaka'ako park yesterday to talk with Apio, because he knows the park well.
Charles Kupahu Timtim and Annie Lokomaika'i Lipscomb
Credit kumu kahua
"I went homeless for as long as I could," Apio says, "To see what it was like, and it was the most terrifying, humiliating, degrading, humbling, and enlightening experience that I've ever had."
Apio says he learned what the human spirit is capable of from his friends on the street.
"It's life unobscured. There's no veil. There's no courteous curtain. It's just in-your-face life, and that helps to be honest in my own writing, and I always strive to do that."
"My students were blown away by the characters," says Gail Harada, who holds the literature seat on the Mayor's Culture and Arts Commission. Harada taught Kamau, the first play, for over 10 years in her literature class at KCC.
"They felt they could relate, they recognized those characters. It was just a well written play."
Harada says metaphor, history, pidgin and Hawaiian language are artfully entwined; there's a lot for students to unpack and chew on.
"It also brings in all the tensions caused by the struggles to survive economically, to preserve one's culture. I think it's important for people to see that kind of struggle."
Speaking of struggle, Harry Wong is directing part three, Ua Pau: It is Finished, Over, Destroyed. He directed the other two plays, Kamau and Kamau 'Ae, as well. Wong says the plays somehow get written in the theatre process.
"Because Alani, until he and I and the actors get together, he doesn't go to what he knows to be true, because he thinks it's going to be too much."
Wong points to the fact that the printed version of the first play, Kamau, ends differently from the staged version, which came later.
"It doesn't end the way the production did because we went into rehearsal with Alika not going back to work. Remember, Alika has to make the choice between supporting his family by fighting for their land, or going back to work to be able to provide for his family."
Wong continues, "In the original ending, he didn't go back to work to provide for them until we were getting ready to open. And I said, Alani come, on, tell me what you think he really does." Alani said, "Although I would like him to stand up for his land, I think he goes back to work." We said, Okay, then let's do that. And he wrote it and it was heartbreaking."
Wong says that through the acting, each character's truth becomes unavoidable. In this part three, as in a mythic hero's journey, the millennial daughter/niece, Stevie, returns from college and must find a way forward.
"Through what happens to Stevie in this last play you see that at last, the spirit is free to create again, to start anew, to no longer carry the burden, to no longer rage."
Harada says situations in the plays are not black and white, no easy answers are presented, but in the plays, family and culture remain a source of strength.
"The play (Kamau) starts with voices, and the first time you see Alika he's giving his canned speech about 'Iolani Palace. At the end of the play, he gives the same speech, and I ask students, Do you think he gives the speech the same way at the beginning and at the end? Opinions are divided, but one thing some students come to is, he might give the speech in the same way, but we don't see it the same way. And the teacher in me gets all happy."
All three parts of the Kamau trilogy are expected to be staged as part of the Pacific Islands Festival of the Arts, summer 2020.
kumu kahua
alani apio
local play
Eating Around: Foundations of Hawai'i's Cuisine
By Noe Tanigawa • Jul 2, 2019
Noe Tanigawa
eady availability of local produce is one lasting contribution consumers enjoy, thanks to recent combined efforts of chefs creating cachet and demand, farmers being encouraged to grow and expand offerings, and the counties and other entities that have supported farmers' markets.Credit Noe TanigawaEdit | Remove
Food is a shared national obsession these days, and the way we eat now has its roots in a movement that began in California in the 1970's. Increased awareness of fresh and local became the basis for Hawai'i Regional Cuisine in 1991, and it got us excited about Hawai'i's rich food culture. HPR's Noe Tanigawa looks at where we are, thirty years later.
Tattoo Today: A Piece of Your Skin for a Part of My Soul
By Noe Tanigawa • Aug 2, 2019
Jay Freestyle
This weekend, organizers expect about ten thousand people will attend the 8th annual Pacific Ink and Art Expo in Honolulu. More than 450 tattoo artists will be plying their trade. Why do some of these artists have more than three million followers on social media?
'Ohana Hale Marketplace: Where? Oh Yeah!!
If you're looking for a different place to shop and eat in Honolulu, 'Ohana Hale Marketplace might interest you. The Marketplace is basically street stalls in air conditioning, and more than fifty of the stalls are food vendors, inside the former Sports Authority on Ward Avenue. New shuttles from Waikīkī should improve traffic for the hopeful entrepreneurs.


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HALFTIME: Evansville 37, Indiana State 22
ST. LOUIS -- D.J.
HALFTIME: Evansville 37, Indiana State 22 ST. LOUIS -- D.J. Check out this story on courierpress.com: https://www.courierpress.com/story/sports/blogs/ue/2016/03/05/halftime-evansville-37-indiana-state-22/87423882/
Evansville Courier & Press Published 6:18 p.m. CT March 5, 2016
ST. LOUIS -- D.J. Balentine scored 10 points to lead a balanced effort as the University of Evansville took a 37-22 halftime lead over Indiana State on Saturday in a Missouri Valley Conference tournament semifinal at Scottrade Center.
Every Aces starter scored in the first 4:30 of the game and UE buried 5 of 10 3-point attempts in the half. Evansville shot 13 of 28 (46.4 percent) from the field, compared to Indiana State's 7 of 25.
Evansville took a 5-0 lead and never trailed. Devonte Brown had 10 points for ISU but no other Sycamore scored more than three.
There were no scoring runs larger than 5-0 in the half for UE, but the Aces steadily built the lead. Jaylon Brown had seven points, three assists and two steals with no turnovers, and Blake Simmons added eight points with a pair of 3-pointers.
Read or Share this story: https://www.courierpress.com/story/sports/blogs/ue/2016/03/05/halftime-evansville-37-indiana-state-22/87423882/


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Notch Signaling Induces Cell Cycle Arrest in Small Cell Lung Cancer Cells
Virote Sriuranpong, Michael W. Borges, Rajani K. Ravi, Dagmar R. Arnold, Barry D. Nelkin, Stephen B. Baylin and Douglas W. Ball
Virote Sriuranpong
Michael W. Borges
Rajani K. Ravi
Dagmar R. Arnold
Barry D. Nelkin
Stephen B. Baylin
Douglas W. Ball
DOI: Published April 2001
Among the various forms of human lung cancer, small cell lung cancer (SCLC) exhibits a characteristic neuroendocrine (NE) phenotype. Neural and NE differentiation in SCLC depend, in part, on the action of the basic-helix-loop-helix (bHLH) transcription factor human achaete-scute homologue-1 (hASH1). In nervous system development, the Notch signaling pathway is a critical negative regulator of bHLH factors, including hASH1, controlling cell fate commitment and differentiation. To characterize Notch pathway function in SCLC, we explored the consequences of constitutively active Notch signaling in cultured SCLC cells. Recombinant adenoviruses were used to overexpress active forms of Notch1, Notch2, or the Notch effector protein human hairy enhancer of split-1 (HES1) in DMS53 and NCI-H209 SCLC cells. Notch proteins, but not HES1 or control adenoviruses, caused a profound growth arrest, associated with a G1 cell cycle block. We found up-regulation of p21waf1/cip1 and p27kip1 in concert with the cell cycle changes. Active Notch proteins also led to dramatic reduction in hASH1 expression, as well as marked activation of phosphorylated extracellular signal-regulated kinase (ERK)1 and ERK2, findings that have been shown to be associated with cell cycle arrest in SCLC cells. These data suggest that the previously described function of Notch proteins as proto-oncogenes is highly context-dependent. Notch activation, in the setting of a highly proliferative hASH1-dependent NE neoplasm, can be associated with growth arrest and apparent reduction in neoplastic potential.
The four mammalian Notch genes encode 300 kDa single pass transmembrane receptors, which play important roles in a diverse group of developing tissues (1) . Binding of one of the Notch ligands, which include Delta1, Jagged1, and Jagged2, leads to a complex cleavage and activation of Notch proteins (1 , 2) . The released and activated COOH-terminal fragment of Notch translocates to the nucleus, in which it interacts with the transcription factor CBF1 (RBPjκ) to transactivate target genes including HES1 (2) . HES 3 family members and other targets of Notch signaling serve to down-regulate and counterbalance the effects of myogenic and neurogenic bHLH factors including MyoD and the achaete-scute complex. The net result of Notch signaling frequently is to preserve a small population of uncommitted multipotential cells within a differentiating tissue (3 , 4) .
Truncated, constitutively active forms of Notch have been shown to have oncogenic activity in acute lymphoblastic T-cell leukemia (5) , mouse mammary tumors (6) , and transformed kidney epithelial cells (7) . The in vitro transforming activity of Notch in rat kidney cells requires the cooperation of E1A oncogene and nuclear localization. Notch signaling may have opposite actions on cell growth, depending on poorly understood differences in cellular context. For example, in hematopoietic cell maturation, ligand activation of Notch1 inhibits the capability of myeloid precursor cells to differentiate in response to granulocyte colony-stimulating factor (8, 9, 10) . In B-cell development, chicken Notch1 has been shown to induce growth arrest and apoptosis (11) , whereas active Notch1 induces ectopic T-cell proliferation in the bone marrow (12) .
In the developing lung, Notch 1 and HES1 are strongly expressed in non-NE airway epithelial cells, whereas MASH1 is restricted to clustered PNECs (13 , 14) . In HES1 transgenic knockout mice, PNECs are markedly increased, with a parallel up-regulation of MASH1 (14) . Conversely, MASH1 transgenic knockout mice fail to develop PNECs (13 , 14) . Consistent with a well-established role in inhibiting commitment and differentiation in neuronal precursors, Notch signaling appears to play a critical role in restricting NE cell development within the airway epithelium.
Previous studies have revealed a tight concordance between the NE phenotype in lung cancer and hASH1. Disrupting hASH1 function by antisense treatment in SCLC cells results in attenuation of NE markers (13) . In a transgenic model simulating NE lung carcinogenesis, hASH1, in conjunction with SV40 large T antigen, caused airway epithelial cell proliferation, and the appearance of aggressive NE tumors when targeted to normal pulmonary epithelium (15) . We have previously shown that hASH1 may be transcriptionally down-regulated by HES1 in SCLC (16) . However, little is known about Notch action in lung cancer. It is conceivable that NE lung cancers have dysregulation of the normal mechanisms that normally extinguish hASH1 expression in fully differentiated neural and NE cells. Notch influences on SCLC proliferation are of particular interest, because there is evidence that some NE features may contribute to SCLC growth. In the present study, we explored whether activation of Notch signaling could disrupt hASH1 expression and interfere with growth of cultured SCLC. Overexpression of activated forms of Notch1 and Notch2 in SCLC cells caused profound growth suppression stemming from a G1 cell cycle arrest. The CDKIs p21waf1/cip1 and p27kip1 were up-regulated accompanying the arrest. Parallel to the growth arrest phenotype, activation of Notch signaling in the SCLC cells led to complete repression of hASH1 and induction of pMAPK. Interestingly, although HES1 was induced by Notch proteins, overexpression of HES1 alone could only partially replicate the Notch phenotype, implying an important role for HES1-independent actions of Notch in this model system.
DMS53 cells were grown in Waymouth's medium (Life Technologies, Rockville, MD) supplemented with 16% FBS, 100 units of penicillin and 100 μg of streptomycin per ml. NCI-H209 cells were grown in RPMI 1640 (Life Technologies) with 10% FBS. Low passage 293 cells were cultured in MEM with 10% FBS. 911 cells were maintained in DMEM (Life Technologies) with 10% FBS. NCI-H209/ΔRaf-1:ER cells were maintained and induced with 1 μm β-estradiol as previously described (17) .
Recombinant Adenovirus Generation and Infection Procedure.
Production of high titer recombinant adenovirus using the AdEasy system has been described previously (18) . Briefly, human intracellular Notch1 and Notch2, and HES1 were generated by PCR and subcloned into a pAdTrackCMV shuttle plasmid, which allows bicistronic expression of GFP and the inserted gene under cytomegalovirus promoters. The intracellular Notch1 fragment contains amino acids 1759–2556 and was amplified from plasmid TAN-1/pCDNA3; Notch2 contains amino acids 1700–2471 and was amplified from plasmid cytohN2pcAMP/pcDNAI/AMP (both gifts of S. Artavanis-Tsakonas, Massachusetts General Hospital Cancer Center, Charlestown, MA). The HES1 fragment was amplified from human lung cancer cDNA. Then the shuttle plasmids with inserted genes were cotransformed with pAdEasy1 into BJ5183 bacterial competent cells to generate the recombinant adenoviruses. High titer viral stocks were first transfected and then amplified in 911 cells. The control virus expresses the Escherichia coli β-galactosidase gene (AdβGal). Each viral stock was titered by plaque assay in low-passage 293 cells. A series of preliminary infections were performed in each cell line to determine the optimal dose of viruses, allowing at least 70% GFP reactive cells with acceptable cytotoxicity determined by growth rate compared with the mock-infected cells. The final doses were 2.5–5 plaque-forming units/cell for DMS53 or 10 plaque-forming units/cell in NCI-H209. In each experiment, the level of GFP expression at 48 h postinfection was assessed to confirm the efficiency of infection.
DNA Sequencing.
Vector constructs were verified by two strands sequencing at the Johns Hopkins Core Sequencing facility. HES1 genomic sequencing was performed with PCR amplification of exons 1–5, containing the entire coding sequence (19) .
Growth Assays.
The colorimetric assay using MTT (Sigma, St. Louis, MO) was performed as described previously (20 , 21) with some modification. Cells were passaged in phenol red-free RPMI 1640 with 10% FBS and antibiotics, then seeded into 24-well plates at 20,000 and 40,000 cells per well for DMS53 and NCI-H209 cells, respectively. For DMS53 cells, 250 μl of fresh medium containing 10% of MTT 5 mg/ml stock was replaced to each well at the day of harvesting. Plates were incubated at 37°C for 3 h; then 750 μl of DMSO (Sigma) was added to each well and shaken at room temperature for 10 min to dissolve intracellular MTT formazan crystals, followed by absorbance measurement at 540 nm. For NCI-H209 cells, the MTT stock was added at 10% of culture volume for each well, and plates were incubated at 37°C for 4 h. After brief centrifugation, all but ∼100 μl of medium was removed, 900 μl of DMSO was added per well, and absorbance was measured at 540 nm. Experiments were done in quadruplicate and were repeated at least twice. Statistical analysis was performed by GraphPad Prism software version 3.02 (GraphPad Software Inc., San Diego, CA) with two-way ANOVA.
Cell Cycle Analysis.
Cells were stained by propidium iodide according to Vindelov's method (22) . Briefly, DMS53 or NCI-H209 cells were harvest at 3 or 4 days after infection. Nuclei were prepared, stained with propidium iodide, and subjected to flow cytometric analysis with an EPICS 752 flow cytometer (Coulter Electronics, Hialeah, FL). The cell cycle parameters from 10,000 gated nuclei were determined with multicycle software (Phoenix Flow System, San Diego, CA). Experiments were done at least twice.
Immunoblotting.
Infected cells were harvested at the time indicated in each experiment. Cells were lysed in 1× SDS sample buffer (62.5 mm Tris, 2% SDS, 10% and glycerol) with aprotinin, leupeptin, pepstatin, and 4-(2-aminoethyl) benzenesulfonyl fluoride (AEBSF). Cell suspensions were briefly sonicated and protein concentration for each cell lysate was determined by DC Protein Assay (Bio-Rad, Hercules, CA) according to the manufacturer's instruction. Fifty to 100 μg of total protein from whole cell lysate were loaded to each lane during gel electrophoresis. Equivalent loading and transfer were verified by filter staining with Fast Green (Fisher Scientific, Pittsburgh, PA). Western blot analysis was performed by using 0.1 m Tris (pH 7.5), 0.9% NaCl, and 0.05% Tween 20 (TBST) with 5% nonfat dry milk as blocking and antibody dilution buffer. Working concentrations of antisera were as follows: Notch1, 1:1000 (Santa Cruz Biotechnology, Santa Cruz, CA); Notch2, 1:50 (anti-bhN6, gift from S. Artavanis-Tsakonas, Massachusetts General Hospital Cancer Center, Charlestown, MA); HES1, 1:10000 (gift from T. Sudo, Toray Industries, Inc., Kanagawa, Japan); MASH1, 1:1000 (BD PharMingen, San Diego, CA); p27, 1:500 (Santa Cruz Biotechnology); p53, 1:1000 (Labvision, Fremont, CA); p16, 1:1000 (Labvision); p21, 1:1000 (Labvision); pMAPK, 1:1000 (New England Biolabs, Beverly, MA); PARP, 1:5000 (BD PharMingen); and G3PDH, 1:5000 (Trevigen, Gaithersburg, MD). A Supersignal West Pico (Pierce, Rockford, IL) chemiluminescence kit was used for all antibodies with the exception that the Supersignal West Femto kit (Pierce, Rockford, IL) was used for antisera to MASH1, p16, and p21.
Activated Notch, but not HES1, Caused Morphological Changes and Growth Inhibition in SCLC Cells.
We selected two SCLC cell lines, NCI-H209 and DMS53, for this study. By immunoblotting, both NCI-H209 and DMS53 expressed endogenous Notch1 and Notch2 proteins at trace or undetectable levels (see control AdβGal lanes in Fig. 1A ⇓ ). We previously attempted to create stable cell lines expressing constitutively active Notch1 and Notch2, but noted a strong selection bias against clones expressing the Notch constructs (data not shown). Therefore, we elected to express Notch1 and Notch2 using recombinant adenoviruses. We were able to achieve at least 70% of cells expressing GFP at 48 h postinfection in both cell lines with minimal evidence of cytotoxicity. On adenovirus infection, DMS53 cells expressed GFP as early as 6 h and NCI-H209 cells as early as 24 h (Fig. 1C) ⇓ postinfection. Expression of the predicted forms of Notch1 and Notch2 was very robust in both cell lines by immunoblotting (Fig. 1A) ⇓ .
Overexpression of Notch adenoviruses caused HES1 induction and morphological changes in SCLC cells. In A, Notch1 and Notch2 expression were undetectable in control DMS53 and NCI-H209 cells by immunoblotting with antiNotch1 and Notch2. Adenovirus overexpression of Notch1 or Notch2 resulted in a strong expression of the expected Mr 116,000 activated form. Activated Notch showed multiple stable faster migrating bands similar to those described previously (7) . In B, overexpression of AdNotch1 and AdNotch2 in NCI-H209 resulted in induction of a Mr 31,000 HES1 band (∗, Lanes 3–4), which is otherwise undetectable (Lanes 1–2). Comparable levels of HES1 were observed when the protein was overexpressed with a HES1 adenovirus (Lane 5). MW, Mr in the thousands. C, left panels, phase-contrast morphological appearance; right panels, GFP expression in the same field. Notch1 overexpression in DMS53 caused rounding cells at 48 h, compared with the control virus. In NCI-H209, 96 h postinfection, Notch1 caused the appearance of flattened adherent clusters, compared with the floating spheroids in control virus.
As anticipated, adenoviral Notch1 and Notch2 induced HES1 protein expression (Fig. 1B) ⇓ . The entire coding sequence of the endogenous HES1 gene appeared to be intact in both DMS53 and NCI-H209 cells (data not shown). In NCI-H209, the level of HES1 induced by adenoviral HES1 was comparable with the level of endogenous HES1 induced by AdNotch1 or AdNotch2 (Fig. 1B ⇓ , compare Lanes 3–6). NCI-H209 cells normally grow in suspension as tight floating cell clusters. The AdNotch1 caused the cell clusters to adhere to the bottom of the plastic culture vessels. The peripheral cells of the adhering clusters had a tendency to spread out from the clusters with elongated cytoplasmic processes (Fig. 1C) ⇓ . The magnitude of changes was maximal beginning at 4 days after infection, when 70–80% of spheroids adhered to the plastic. The AdNotch2 infection also caused similar effects but to a lesser extent than the AdNotch1, with only 10–20% of cells showing the adherent morphology. DMS53 cells, which normally grow as a tightly adherent monolayer, exhibited rounding and detachment from the plastic starting from 24 h postinfection by AdNotch1 and AdNotch2 (Fig. 1C) ⇓ . None of these morphological changes were observed with the control virus or with AdHES1.
The growth rates of DMS53 cells infected by AdNotch1 or AdNotch2 markedly decreased compared with the rates from the control virus or the AdHES1-infected cells (Fig. 2A) ⇓ . In NCI-H209, AdNotch1 completely arrested cell proliferation (Fig. 2B) ⇓ . The effect of AdNotch2 on the growth of NCI-H209 was minimal, paralleling the weaker morphological effect of Notch2 in this cell line. In both cell lines, the effect of HES1 was indistinguishable from that of cells expressing the β-Gal control virus. To screen for a nonspecific viral cytotoxic effect, we compared the growth rate of mock-infected and control virus (AdβGal)-infected cells. We observed a small but statistically significant decrease of growth rate attributable to virus alone (Fig. 2, A and B) ⇓ .
Growth rate of Notch- or HES1-infected SCLC cells by MTT assays. In DMS53 cells, both AdNotch1 and AdNotch2 produced a significant decrease compared with the control virus (AdβGal) (P < 0.0001 for both). The growth of AdHES1 cells was similar to AdβGal (P = 0.3599). In NCI-H209 cells, AdNotch1 caused profound growth suppression, AdNotch2-infected cells had a slightly decreased growth rate that was significantly different from the control virus (P < 0.0001 and 0.0004, respectively), which had a growth rate similar to that of AdHES1 cells (P = 0.8023; experiment performed in quadruplicate, mean ± SD). PI, postinfection.
Activated Notch-induced G1 Cell Cycle Arrest Associated with p21waf1/cip1 and p27kip1 Induction.
To further investigate the growth inhibitory effect of active Notch in SCLC, we performed cell cycle analysis by propidium iodide staining and flow cytometry. These studies showed a typical G1 arrest pattern in both cell lines by Notch1, and in DMS53 cells but not in NCI-H209 cells by Notch2 (Fig. 3 ⇓ ; Table 1 ⇓ ). Although Notch1 exerted a stronger growth arrest effect than Notch2 in DMS53 cells by MTT, we were unable to detect a significant difference in their potency to induce the G1 arrest at 72 h after infection (compare Table 1 ⇓ and Fig. 3 ⇓ with Fig. 2 ⇓ results). In contrast to the reported induction of apoptosis by Notch1 in B cells (11) , we could not observe a significant sub-G0-G1 peak in our study, which suggested that there was not a significant fraction of apoptotic nuclei at that time point. Moreover, bis-benzimide trihydrochloride (hoechst 33342; Sigma) nuclear staining and immunoblotting to detect a PARP cleavage product did not reveal an obvious increase in apoptosis (data not shown).
Effect of the active Notch1 or HES1 on cell cycle distribution. DMS53 cells were harvested 3 days postinfection for cell cycle analysis using propidium iodide staining. X axis, DNA content; Y axis, the number of nuclei. Compared with the control virus, Notch1 caused G1 cell cycle arrest, whereas HES1 had no significant effect.
Summary of the cell cycle distribution by flow cytometry analysis
To further characterize the G1 arrest, we examined the level of expression of several known G1 cell cycle regulatory factors. In DMS53 cells, which have intact Rb protein (23) , both active Notch1 and Notch2 caused specific induction of p21waf1/cip1 as early as 24 h postinfection and correlating with the G1 arrest (Fig. 4A ⇓ , p21, Lanes 5 and 6). The levels of p27kip1, readily detectable in these cells, did not increase with Notch proteins. Activated Notch did not induce p16 or p53, nor did it down-regulate cyclinD1 or cyclinE (Fig. 4A ⇓ , p53, p27, p16; data not shown for cyclinD1 and cyclinE immunoblotting). In NCI-H209 cells, which have an inactive mutant Rb gene (24) , AdNotch1 caused a significant increase of both p21waf1/cip1 and p27kip1 corresponding to the G1 arrest, whereas the control viral infection did not cause induction of the CDKIs (Fig. 4B ⇓ , compare Lanes 6–9 versus 2–5). Significantly, adenoviral HES1 was unable to induce growth arrest, cell cycle changes, or CDKI expression in either cell line.
Expression of G1 cell cycle regulatory proteins. In A, DMS53 cells were harvested at 24 and 48 h postinfection. Both Notch1 and Notch2 caused significant up-regulation of p21waf1/cip1 (Lanes 5–6; data not shown for Notch2). The level of p53 was not significantly altered. In B, in NCI-H209, progressive increase of p21waf1/cip1 and p27kip1 by activated Notch1 was observed during a 4-day period postinfection (Lanes 6–9). G3PDH was used as an internal control for loading.
Repression of hASH1 Expression by Active Notch.
In a recent study, we showed that transgenic overexpression of hASH1 promotes airway epithelial cell hyperplasia, and cooperates with SV40 T antigen in promoting lung tumorigenesis (15) . In contrast, transgenic knockout of this factor produces a hypoproliferative phenotype in brain, olfactory epithelium, and the adrenal medulla, as well as an absence of lung NE cells (13) . We investigated whether the Notch-induced growth arrest correlated with any reduction in hASH1 expression. Overexpression of active Notch proteins caused a prompt and complete loss of hASH1 protein in both NCI-H209 and DMS53 cells (Figs. 5A ⇓ , Lanes 3–6, and 5B ⇓ , Lanes 6–9). In contrast, overexpression of HES1 caused a more modest and slower paced decrease in hASH1 protein abundance (Fig. 5A ⇓ , Lanes 7–9). These data correlate well with the slow decline observed in hASH1 mRNA over 96 h when we used an inducible rat HES1 vector in DMS53 cells (16) . The magnitude of reduction of hASH1 expression corresponded well with the extent of the morphological changes and the growth arrest in SCLC cells infected by the Notch adenoviruses.
Endogenous hASH1 expression was decreased by Notch and HES1. In A, in DMS53 cells, Notch1 and Notch2 completely reduced expression of the Mr 31,000–33,000 hASH1 species by 24 h postinfection (Lanes 2–6). With HES1 overexpression, the hASH1 level decreased slowly (Lanes 7–9). In B, in NCI-H209 cells, Notch1 down-regulated hASH1 expression in a comparable fashion to DMS53 cells (compare Lanes 6–9 to Lanes 1–5).
Activation of MAPK in SCLC by Notch Signaling.
In previous studies in SCLC cell lines, the activation of the raf/MEK/MAPK pathway resulted in a loss of anchorage-independent growth and cell cycle arrest, mediated primarily in NCI-H209 cells by p27kip1 (17) and by p16 in DMS53 cells (25) . Because we observed similar changes in the phenotype of Notch-infected cells, we then investigated the possibility that the raf/MEK/pMAPK pathway could be activated by Notch signaling. Similar to the effects of Notch, activation of raf resulted in down-regulation of hASH1 in NCI-H209 cells (Fig. 6A) ⇓ . Strikingly, Notch overexpression in both of the SCLC cell lines led to an increase in pMAPK, corresponding to the growth arrest phenotype (Fig. 6B ⇓ , Lanes 4–5, 11–13). Thus, in DMS53 cells, Notch1 markedly increased phosphorylated ERK1 and ERK2, whereas Notch2 had a weaker but still significant effect. In NCI-H209 cells, only Notch1 could induce MAPK activation. Treatment with the dual MEK inhibitor, U0126, resulted in toxicity in both DMS53 and NCI-H209 cells. Therefore, we were unable to definitively determine whether MAPK phosphorylation is necessary for the Notch-induced cell cycle block in these cells.
Raf activation in SCLC caused reduction of hASH1, and activated Notch caused induction of pMAPK in SCLC cells. In A, in NCI-H209/Δraf: ER cells, on activation of raf by β-estradiol, hASH1 abundance significantly decreased after 24 h. In B, immunoblotting using phosphospecific antibody to MAPK showed strong induction of phosphoERK1 (p44) and phosphoERK2 (p42) by Notch adenoviruses in both DMS53 and NCI-H209 (Lanes 4–5, 11–13). DMS53 cells were harvested at 72 h after infection. The basal levels of pMAPK in the SCLC cells are negligible and nondetectable by the antibody (Lanes 2–3, 7–10). U1752, a non-SCLC that has high basal levels of MAPK phosphorylation because of overexpression of epidermal growth factor receptor was used as a positive control (Lane 1).
In the present study, we showed novel effects of an active form of Notch in SCLC cells. Activation of Notch signaling suppressed growth and arrested the SCLC cells in the G1 phase. In a parallel fashion, Notch specifically induced p21waf1/cip1 and p27kip1. Overexpression of Notch caused abrupt and complete down-regulation of hASH1 in conjunction with induction of ras signaling pathway activity. Although HES1, a key downstream effector of Notch, could also decrease hASH1 expression, it was unable to suppress growth of SCLC, affect CDKIs, or induce MAPK activation. Overall, these data suggest that in the context of a proliferating NE tumor using bHLH factors such as hASH1, the net effect of high-level Notch signaling can be antiproliferative in a process mediated largely via HES1-independent mechanisms.
In a limited number of tumor types, including human T-cell leukemia and mouse mammary tumors, Notch signaling is oncogenic rather than antiproliferative (5 , 6 , 26) . However, a recent study showed opposite functions of Notch depending on the cell context. In the development of lymphoid cells, Notch promotes T-cell proliferation but induces apoptosis and growth arrest of B-cell precursors (11) . The cellular mechanisms that mediate growth arrest by Notch remain unclear. It is unlikely that an intact Rb pathway is necessary for Notch-induced arrest because this effect occurred in NCI-H209 cells, which have an inactive mutant Rb gene. The inability of HES1 to induce growth arrest indicates the contribution of HES1-independent mechanism(s). We observed consistently distinct responses to Notch1 versus Notch2 in the SCLC cells. With careful titering and expression studies, we believe that the differences stem from an intrinsically different response to the two forms of Notch rather than a bias in protein dosage. Interestingly, Notch2 was also less potent than Notch1 in inducing transformation of E1A-immortalized rat kidney cells (7) .
Activation of the ras/raf/MAPK pathway is frequently associated with transformation of primary cells and tumor progression of many established cancer cells (27 , 28) . However, an increasing body of evidence supports the idea that constitutive activation of this pathway may lead to growth arrest and terminal differentiation in some cellular contexts. This phenomenon has been observed in SCLC (17 , 25) and medullary thyroid cancer cells (29) , as well as in PC12 rat pheochromocytoma cells, (30) for example. In certain developmental contexts, there is evidence of an antagonistic interaction between Notch and ras signaling in a cell-autonomous manner (31, 32, 33) . However, a recent study in mammary tumors that were derived from transgenic activated Notch4 mice showed a possible cooperation between ras signaling and Notch-mediated tumorigenesis (34) . Although our findings indicate a positive interaction between the two signaling pathways, the cell cycle arrest induced by Notch may not be solely dependent on the MAPK activation. Under our experimental conditions, we have been unable to demonstrate reversal of this growth arrest phenotype by MEK inhibitors. Interestingly, the induction of pMAPK coincided well with the hASH1 reduction and cell cycle arrest by Notch. This association may represent a possible cooperative interaction between the Notch and ras signaling pathways in controlling growth and NE differentiation in SCLC cells. It will be of great interest to elucidate the exact mechanisms of the cross-talk between the two signaling pathways, which may potentially be applicable to other cell contexts.
Our data showed a correlation between the cell cycle arrest by Notch signaling and the induction of p21waf1/cip1 in two different SCLC lines with either wild-type or mutated Rb. Both of these cell lines have an intact p53. The potential interaction between Notch signaling and p21waf1/cip1 is fascinating in light of developmental parallels. Notch signaling is a hallmark of quiescent, multipotent cells in hematopoietic (35 , 36) and central nervous system development (37 , 38) , for example. In the maturation process of these tissues, activation of Notch signaling blocks the capability to respond to differentiating stimuli (39, 40, 41) . A recent study showed that p21waf1/cip1 plays a crucial role in maintaining the self-renewing properties of hematopoietic stem cells (42) . It will be interesting to determine whether Notch functions in maintaining multipotent progenitor cells through p21waf1/cip1.
In summary, Notch signaling in cultured SCLC cells results in a profound G1 cell cycle arrest associated with p21waf/cip1 induction, repression of hASH1, and induction of the downstream ras signaling pathway. Significantly, the current study suggests a direct link between Notch and the raf/MEK/MAPK pathway, although it is presently unclear at what level the MAPK pathway is being activated and what effectors downstream of Notch are implicated. In future studies, it will be important to characterize the signaling pathway responsible for Notch-dependent growth arrest in SCLC cells, understand the effect of Notch signaling in other NE neoplasms, and study the effects of Notch signaling using Notch ligands.
We thank Christopher Strock for critically reviewing the manuscript, Eric Nakakura, and D. Neil Watkins for helpful discussion, Tong Chuan He (Dr. Bert Vogelstein's laboratory, The Johns Hopkins University, Baltimore, MD) for the gift of the Spyros adenoviral vector and cell lines, Spyros Artavanis-Tsakonas (Massachusetts General Hospital Cancer Center, Harvard Medical School, Charlestown, MA) for helpful discussion and the gift of Notch vectors and Notch2 antisera.
↵1 Supported by the Royal Thai Government Scholarship, Chulalongkorn University, Bangkok, Thailand (to V. S.), and NIH-National Cancer Institute Grants RO1CA70244 (to D. W. B.) and RO1CA47480 (to B. D. N.), and a training grant in Endocrinology and Metabolism T32 DK07751 (to D. R. A.).
↵2 To whom requests for reprints should be addressed, at Johns Hopkins Oncology Center, 1650 Orleans Street, Room 553, Baltimore, MD 21231.
↵3 The abbreviations used are: HES, hairy enhancer of split; bHLH, basic helix-loop-helix; MASH1, mammalian achaete-scute homologue-1; NE, neuroendocrine; PNEC, pulmonary NE cell; hASH1, human achaete-scute homologue-1; SCLC, small cell lung cancer; CDKI, cyclin-dependent-kinase inhibitor; ERK, extracellular signal-regulated kinase; MEK, MAPK erk kinase; pMAPK, phosphorylated mitogen-activated-protein kinase; FBS, fetal bovine serum; GFP, green fluorescent protein; MTT, 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide; PARP, poly(ADP-ribose) polymerase; G3PDH, glyceraldehyde-3 phosphate dehydrogenase; Rb, retinoblastoma.
Received September 28, 2000.
Artavanis-Tsakonas S., Rand M. D., Lake R. J. Notch signaling: cell fate control and signal integration in development. Science (Washington DC), 284: 770-776, 1999.
Weinmaster G. The ins and outs of notch signaling. Mol. Cell. Neurosci., 9: 91-102, 1997.
Shawber C., Nofziger D., Hsieh J. J., Lindsell C., Bogler O., Hayward D., Weinmaster G. Notch signaling inhibits muscle cell differentiation through a CBF1-independent pathway. Development (Camb.), 122: 3765-3773, 1996.
Lewis J. Notch signalling and the control of cell fate choices in vertebrates. Semin. Cell Dev. Biol., 9: 583-589, 1998.
Ellisen L. W., Bird J., West D. C., Soreng A. L., Reynolds T. C., Smith S. D., Sklar J. TAN-1, the human homolog of the Drosophila notch gene, is broken by chromosomal translocations in T lymphoblastic neoplasms. Cell, 66: 649-661, 1991.
Uyttendaele H., Marazzi G., Wu G., Yan Q., Sassoon D., Kitajewski J. Notch4/int-3, a mammary proto-oncogene, is an endothelial cell-specific mammalian Notch gene. Development (Camb.), 122: 2251-2259, 1996.
Capobianco A. J., Zagouras P., Blaumueller C. M., Artavanis-Tsakonas S., Bishop J. M. Neoplastic transformation by truncated alleles of human NOTCH1/TAN1 and NOTCH2. Mol. Cell. Biol., 17: 6265-6273, 1997.
Milner L. A., Bigas A., Kopan R., Brashem-Stein C., Bernstein I. D., Martin D. I. Inhibition of granulocytic differentiation by mNotch1. Proc. Natl. Acad. Sci. USA, 93: 13014-9, 1996.
Bigas A., Martin D. I., Milner L. A. Notch1 and Notch2 inhibit myeloid differentiation in response to different cytokines. Mol. Cell. Biol., 18: 2324-2333, 1998.
Li L., Milner L. A., Deng Y., Iwata M., Banta A., Graf L., Marcovina S., Friedman C., Trask B. J., Hood L., Torok-Storb B. The human homolog of rat Jagged1 expressed by marrow stroma inhibits differentiation of 32D cells through interaction with Notch1. Immunity, 8: 43-55, 1998.
Morimura T., Goitsuka R., Zhang Y., Saito I., Reth M., Kitamura D. Cell-cycle arrest and apoptosis induced by Notch1 in B cells. J. Biol. Chem., 275: 36523-31, 2000.
Pui J. C., Allman D., Xu L., DeRocco S., Karnell F. G., Bakkour S., Lee J. Y., Kadesch T., Hardy R. R., Aster J. C., Pear W. S. Notch1 expression in early lymphopoiesis influences B versus T lineage determination. Immunity, 11: 299-308, 1999.
Borges M., Linnoila R. I., van de Velde H. J., Chen H., Nelkin B. D., Mabry M., Baylin S. B., Ball D. W. An achaete-scute homologue essential for neuroendocrine differentiation in the lung. Nature (Lond.), 386: 852-855, 1997.
Ito T., Udaka N., Yazawa T., Okudela K., Hayashi H., Sudo T., Guillemot F., Kageyama R., Kitamura H. Basic helix-loop-helix transcription factors regulate the neuroendocrine differentiation of fetal mouse pulmonary epithelium. Development (Camb.), 127: 3913-3921, 2000.
Linnoila R. I., Zhao B., DeMayo J. L., Nelkin B. D., Baylin S. B., DeMayo F. J., Ball D. W. Constitutive achaete-scute homologue-1 promotes airway dysplasia and lung neuroendocrine tumors in transgenic mice. Cancer Res., 60: 4005-4009, 2000.
Chen H., Thiagalingam A., Chopra H., Borges M. W., Feder J. N., Nelkin B. D., Baylin S. B., Ball D. W. Conservation of the Drosophila lateral inhibition pathway in human lung cancer: a hairy-related protein (HES-1) directly represses achaete-scute homolog-1 expression. Proc. Natl. Acad. Sci. USA, 94: 5355-5360, 1997.
Ravi R. K., Weber E., McMahon M., Williams J. R., Baylin S., Mal A., Harter M. L., Dillehay L. E., Claudio P. P., Giordano A., Nelkin B. D., Mabry M. Activated Raf-1 causes growth arrest in human small-cell lung cancer cells. J. Clin. Investig., 101: 153-159, 1998.
He T. C., Zhou S., da Costa L. T., Yu J., Kinzler K. W., Vogelstein B. A simplified system for generating recombinant adenoviruses. Proc. Natl. Acad. Sci. USA, 95: 2509-2514, 1998.
Feder J. N., Li L., Jan L. Y., Jan Y. N. Genomic cloning and chromosomal localization of HRY, the human homolog to the Drosophila segmentation gene, hairy. Genomics., 20: 56-61, 1994.
Carmichael J., DeGraff W. G., Gazdar A. F., Minna J. D., Mitchell J. B. Evaluation of a tetrazolium-based semiautomated colorimetric assay: assessment of chemosensitivity testing. Cancer Res., 47: 936-942, 1987.
Scudiero D. A., Shoemaker R. H., Paull K. D., Monks A., Tierney S., Nofziger T. H., Currens M. J., Seniff D., Boyd M. R. Evaluation of a soluble tetrazolium/formazan assay for cell growth and drug sensitivity in culture using human and other tumor cell lines. Cancer Res., 48: 4827-4833, 1988.
Vindelov L. L., Christensen I. J., Nissen N. I. A detergent-trypsin method for the preparation of nuclei for flow cytometric DNA analysis. Cytometry, 3: 323-327, 1983.
Helin K., Holm K., Niebuhr A., Eiberg H., Tommerup N., Hougaard S., Poulsen H. S., Spang-Thomsen M., N
rgaard P. Loss of the retinolastoma protein-related p130 protein in small cell lung carcinoma. Proc. Natl. Acad. Sci. USA, 94: 6933-6938, 1997.
Phelps R. M., Johnson B. E., Ihde D. C., Gazdar A. F., Carbone D. P., McClintock P. R., Linnoila R. I., Matthews M. J., Bunn P. A., Jr., Carney D., Minna J. D., Mulshine J. L. NCI-Navy Medical Oncology Branch cell line data base. J. Cell. Biochem., 24 (Suppl.): 32-91, 1996.
Ravi R. K., Thiagalingam A., Weber E., McMahon M., Nelkin B. D., Mabry M. Raf-1 causes growth suppression and alteration of neuroendocrine markers in DMS53 human small-cell lung cancer cells. Am. J. Respir. Cell Mol. Biol., 20: 543-549, 1999.
Pear W. S., Aster J. C., Scott M. L., Hasserjian R. P., Soffer B., Sklar J., Baltimore D. Exclusive development of T cell neoplasms in mice transplanted with bone marrow expressing activated Notch alleles. J. Exp. Med., 183: 2283-2291, 1996.
Marshall M. Interactions between Ras and Raf: key regulatory proteins in cellular transformation. Mol. Reprod. Dev., 42: 493-499, 1995.
Mitsudomi T., Viallet J., Mulshine J. L., Linnoila R. I., Minna J. D., Gazdar A. F. Mutations of ras genes distinguish a subset of non-small-cell lung cancer cell lines from small-cell lung cancer cell lines. Oncogene, 6: 1353-1362, 1991.
Chen H., Carson-Walter E. B., Baylin S. B., Nelkin B. D., Ball D. W. Differentiation of medullary thyroid cancer by C-Raf-1 silences expression of the neural transcription factor human achaete-scute homolog-1. Surgery, 120: 168-173, 1996.
Wood K. W., Qi H., D'Arcangelo G., Armstrong R. C., Roberts T. M., Halegoua S. The cytoplasmic raf oncogene induces a neuronal phenotype in PC12 cells: a potential role for cellular raf kinases in neuronal growth factor signal transduction. Proc. Natl. Acad. Sci. USA, 90: 5016-5020, 1993.
Miller D. T., Cagan R. L. Local induction of patterning and programmed cell death in the developing Drosophila retina. Development (Camb.), 125: 2327-2335, 1998.
Ordentlich P., Lin A., Shen C. P., Blaumueller C., Matsuno K., Artavanis-Tsakonas S., Kadesch T. Notch inhibition of E47 supports the existence of a novel signaling pathway. Mol. Cell. Biol., 18: 2230-2239, 1998.
Ikeya T., Hayashi S. Interplay of Notch and FGF signaling restricts cell fate and MAPK activation in the Drosophila trachea. Development (Camb.), 126: 4455-4463, 1999.
Fitzgerald K., Harrington A., Leder P. Ras pathway signals are required for notch-mediated oncogenesis. Oncogene., 19: 4191-4198, 2000.
Milner L. A., Kopan R., Martin D. I., Bernstein I. D. A human homologue of the Drosophila developmental gene, Notch, is expressed in CD34+ hematopoietic precursors. Blood, 83: 2057-2062, 1994.
Varnum-Finney B., Purton L. E., Yu M., Brashem-Stein C., Flowers D., Staats S., Moore K. A., Le Roux I., Mann R., Gray G., Artavanis-Tsakonas S., Bernstein I. D. The Notch ligand, Jagged-1, influences the development of primitive hematopoietic precursor cells. Blood, 91: 4084-4091, 1998.
Zhong W., Jiang M. M., Weinmaster G., Jan L. Y., Jan Y. N. Differential expression of mammalian Numb, Numblike and Notch1 suggests distinct roles during mouse cortical neurogenesis. Development (Camb.), 124: 1887-1897, 1997.
Henrique D., Hirsinger E., Adam J., Le Roux I., Pourquie O., Ish-Horowicz D., Lewis J. Maintenance of neuroepithelial progenitor cells by Δ-Notch signalling in the embryonic chick retina. Curr. Biol., 7: 661-670, 1997.
Walker L., Lynch M., Silverman S., Fraser J., Boulter J., Weinmaster G., Gasson J. C. The Notch/Jagged pathway inhibits proliferation of human hematopoietic progenitors in vitro. Stem Cells, 17: 162-171, 1999.
Carlesso N., Aster J. C., Sklar J., Scadden D. T. Notch1-induced delay of human hematopoietic progenitor cell differentiation is associated with altered cell cycle kinetics. Blood, 93: 838-848, 1999.
Culi J., Modolell J. Proneural gene self-stimulation in neural precursors: an essential mechanism for sense organ development that is regulated by Notch signaling. Genes Dev., 12: 2036-2047, 1998.
Cheng T., Rodrigues N., Shen H., Yang Y., Dombkowski D., Sykes M., Scadden D. T. Hematopoietic stem cell quiescence maintained by p21cip1/waf1. Science (Washington DC), 287: 1804-1808, 2000.
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Motionless in White to Dominate at HOB!
October is gearing up to be quite a badass month for shows! On October 9th; Motionless in White will be bringing their "The Trick 'R' Treat" tour to the House of Blues, Las Vegas. On this bill we will see performances by fellow metalcore contemporaries, We Came as Romans; genre riding deathcore/metalcore act, After the Burial; and the dynamic duo that is Twiztid.
While We Came as Romans haven't been back to Vegas since last year; I was fortunate enough to catch both After the Burial and Twiztid earlier this year on separate bills. They do not disappoint so make sure you pile in early for those two!
Past references
I mention this bill quite a bit… I've covered literally the entire line up since the 2009 ish show that Bleeding Through played at The Farm over on the west side of the city. Motionless in White played on this bill; and I believe it was the last time I caught an entire set of theirs. I did catch the very end of their set at last year's Van's Warped Tour downtown; but they were pretty hard to see from all the way in the back. Definitely one of those bands that exploded in popularity this last decade!
How did they blow up in popularity, anyway? My guess is that while the band started off as a traditional metalcore band, their drift towards a more industrial metal sound was more beneficial to them and their audience as a whole. If we go back and look at that golden era of the 2000's for metalcore; I can see why the need to go a different direction would be necessary. With a set few heavyweights dominating in that time period, I can imagine it'd be easy to get lost in the mix without shifting sound and demographics a bit.
I can only assume that this tour is in support of their most recent album release titled, "Disguise". The album was released in early June of this year. Upon listening to some of the tracks of this release, I can subtly hear the elements of metalcore that I've always been drawn to. And though I couldn't tell you a damn thing about that one show I saw them play a decade ago; from peer fan reviews and the abundant fan base they've acquired; I can't imagine being disappointed by these guys in the slightest. If anything, I wish I had kept up with them more through the years to full on experience their growth.
One for the books!
This show is going to be one hell of a night for the books! All four of these bands are highly successful in their niches! They have all graced the Warped Tour stages through the years; along with an uncountable number of big name stages around the world. Who are you most excited for on this bill? How do you feel about the diversity of this line up? We can't wait! See you in the pit, Vegas!
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Courtney Ware
After the Burial, House of Blues Las Vegas, Motionless in White, Twiztid, We Came As Romans


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Bad Santa? Navy officer accused of Christmas party misconduct
By Mackenzie Wolf
The Pentagon with the Washington Monument and National Mall in the background. (Staff Sgt. Perry Aston/U.S. Air Force)
A Navy commander who was accused of sexual misconduct at a Pentagon Christmas party last year stayed on for months as the spokesman for the Navy's top admiral before being reassigned recently, USA Today reported on Thursday.
Cmdr. Chris Servello, 41, was dressed as Santa Claus at an alcohol-fueled party in December inside the Navy's public affairs headquarters at the Pentagon. Servello was accused of sexual misconduct while at the party, allegedly slapping a woman on the buttocks and making unwanted sexual advances, according to documents obtained by USA Today.
At the time, Servello was serving as the spokesman for Adm. John Richardson, the chief of naval operations. Servello kept his influential job for months despite written warnings about his behavior.
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Navy Capt. Peter Hudson completed the initial inquiry into the incident and recommended that Servello be immediately reassigned to "a billet with no supervisory responsibilities and with dedicated oversight."
"I have cooperated fully with all investigations conducted by the Navy," Servello told USA Today in a statement. "Ultimately, the allegations were unproven and charges were not filed. I should have never put myself into a situation where my judgment or my military bearing could be called into question. I deeply regret this and have been held to account for my decision making. Indeed, the overall effect of the administrative counseling makes it highly unlikely that my career will recover."
"The most important thing I can tell you is that the allegations were unproven and charges were never filed," Servello told Navy Times via email Thursday.
"I don't think this story accurately captures the events of that afternoon, nor does it fully convey the findings of the final investigation," he said.
A Naval Criminal Investigative Service investigation was completed in the spring.
"I followed the prosecutor's recommendations to the letter, imposing several adverse measures including dismissing Cdr. Servello from the staff and reassigning him to a non-supervisory position," Richardson said in a statement. "I was advised by legal counsel throughout, and due process was strictly followed to safeguard both the dignity and safety of the complainants and the rights of Cdr. Servello."
About Mackenzie Wolf
Mackenzie Wolf is an editorial intern for Military Times.
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