Monique T. Barakat

Learning from Jekyll to control Hyde: Hedgehog signaling in development and cancer

The Hedgehog (Hh) cascade controls cell proliferation, differentiation and patterning of tissues during embryogenesis but is largely suppressed in the adult. The Hh pathway can become reactivated in cancer. Here, we assimilate data from recent studies to understand how and when the Hh pathway is turned on to aid the neoplastic process. Hh signaling is now known to have a role in established tumors, enabling categorization of tumors based on the role Hh signaling plays in their growth. This categorization has relevance for prognosis and targeted therapeutics. In the first category, abnormal Hh signaling initiates the tumor. In the second category, Hh signaling helps maintain the tumor. In the third category, Hh signaling is implicated but its role is not yet defined.

Insulin-like Growth Factor 2 Is Required for Progression to Advanced Medulloblastoma in patched1 Heterozygous Mice

Medulloblastoma (MB) can arise in the cerebellum due to genetic activation of the Sonic Hedgehog (Shh) signaling pathway. During normal cerebellum development, Shh spurs the proliferation of granule neuron precursors (GNP), the precursor cells of MB. Mutations in the Shh receptor gene patched1 (ptc1+/-) lead to increased MB incidence in humans and mice. MB tumorigenesis in mice heterozygous for ptc1+/- shows distinct steps of progression. Most ptc1+/- mice form clusters of preneoplastic cells on the surface of the mature cerebellum that actively transcribe Shh target genes. In approximately 15% of mice, these preneoplastic cells will become fast-growing, lethal tumors. It was previously shown that the loss of function of insulin-like growth factor 2 (igf2) suppresses MB formation in ptc1+/- mice. We found that igf2 is not expressed in preneoplastic lesions but is induced as these lesions progress to more advanced MB tumors. Igf2 is not required for formation of preneoplastic lesions but is necessary for progression to advanced tumors. Exogenous Igf2 protein promoted proliferation of MB precursor cells (GNP) and a MB cell line, PZp53(MED). Blocking igf2 signaling inhibited growth of PZp53(MED) cells, implicating igf2 as a potential clinical target.

Phenotypic Differences in Reentrainment Behavior and Sensitivity to Nighttime Light Pulses in Siberian Hamsters

Spontaneous reentrainment to phase shifts of the photocycle is a fundamental property of all circadian systems. Siberian hamsters are, however, unique in this regard because most fail to reentrain when the LD cycle (16-h light/day) is phase delayed by 5 h. In the present study, the authors compared reentrainment responses in hamsters from 2 colonies. One colony descended from animals trapped in the wild more than 30 years ago (designated “nonentrainers”), and the other colony was outbred as recently as 13 years ago (designated “entrainers”). As reported previously, only 10% of hamsters from the nonentrainer colony reentrained to a 5-h phase delay of the LD cycle. By contrast, 75% of animals from the entrainer colony reentrained to the phase shift. Another goal of this study was to test the hypothesis that failure to reentrain was a consequence of light exposure during the middle of the night on the day of the 5-h phase delay. This hypothesis was tested by exposing animals to 2 h of light during the early, middle, or late part of the night and then subjecting them on the next day to a 3-h phase delay of the photocycle, which is a phase shift to which all hamsters normally reentrain. All animals from both colonies reentrained when light pulses occurred early in the night, but more animals from the entrainer colony, compared to the nonentrainer colony, reentrained when the light pulse occurred in the middle or late part of the night. The phenotypic variation in reentrainment responses is similar to the variation in photoperiodic responsiveness previously reported for these 2 colonies. Phenotypic variation in both traits is due to underlying differences in circadian organization and suggests a common genetic basis for reentrainment responses and photoperiodic responsiveness.

Kif3a is necessary for initiation and maintenance of medulloblastoma

Medulloblastoma (MB) cells arise from granule neuron precursors (GNPs) that have lost growth control. During normal development, GNPs divide in response to Sonic hedgehog (SHH), a ligand that binds to the patched (PTCH) receptor on GNPs. If one copy of the Ptch gene is lost, as in human Gorlins syndrome and in Ptch(+/-) mice, MBs may form. Proper transduction of the SHH signal critically depends on primary cilia. Loss of primary cilia results in improper signal reception and failure to properly activate SHH target genes. KIF3a, part of a kinesin motor, is required for formation of primary cilia. Here, we use tamoxifen-induced ablation of Kif3a in GNPs of postnatal Ptch(+/-) mouse cerebella to show that KIF3a is necessary for MB formation. To investigate the importance of primary cilia in established tumors, we deleted Kif3a from cultured cells and from tumor cell grafts. The loss of Kif3a from established tumors led to their growth arrest and regression. MBs behave as if they are addicted to the presence of primary cilia. These results underscore the potential utility of agents that disrupt cilia for the treatment of Hh pathway-related MBs.

Tail wags dog: primary cilia and tumorigenesis.

Aberrant activation of the Hedgehog (Hh) signaling pathway contributes to many forms of cancer. Primary cilia are Hh signal transduction centers. Two papers in a recent issue of Nature Medicine (Han et al., 2009; Wong et al., 2009) show that mutating cilia can increase or reduce the rates of tumorigenesis depending on how the Hh pathway is disrupted.

Light Pulses Do Not Induce C-Fos or Per1 in the SCN of Hamsters That Fail to Reentrain to the Photocycle

Circadian activity rhythms of most Siberian hamsters (Phodopus sungorus sungorus) fail to reentrain to a 5-h phase shift of the light-dark (LD) cycle. Instead, their rhythms free-run at periods close to 25 h despite the continued presence of the LD cycle. This lack of behavioral reentrainment necessarily means that molecular oscillators in the master circadian pacemaker, the SCN, were unable to reentrain as well. The authors tested the hypothesis that a phase shift of the LD cycle rendered the SCN incapable of responding to photic input. Animals were exposed to a 5-h phase delay of the photocycle, and activity rhythms were monitored until a lack of reentrainment was confirmed. Hamsters were then housed in constant darkness for 24 h and administered a 30-min light pulse 2 circadian hours after activity onset. Brains were then removed, and tissue sections containing the SCN were processed for in situ hybridization. Sections were probed with Siberian hamster c-fosand per1mRNA probes because light rapidly induces these 2 genes in the SCN during subjective night but not at other circadian phases. Light pulses induced robust expression of both genes in all animals that reentrained to theLDcycle, but no expression was observed in any animal that failed to reentrain. None of the animals exhibited an intermediate response. This finding is the first report of acute shift in a photocycle eliminating photosensitivity in the SCN and suggests that a specific pattern of light exposure may desensitize the SCN to subsequent photic input.

Affordable Care Act and healthcare delivery: A comparison of California and Florida hospitals and emergency departments

Importance The Affordable Care Act (ACA) has expanded access to health insurance for millions of Americans, but the impact of Medicaid expansion on healthcare delivery and utilization remains uncertain. Objective To determine the early impact of the Medicaid expansion component of ACA on hospital and ED utilization in California, a state that implemented the Medicaid expansion component of ACA and Florida, a state that did not. Design Analyze all ED encounters and hospitalizations in California and Florida from 2009 to 2014 and evaluate trends by payer and diagnostic category. Data were collected from State Inpatient Databases, State Emergency Department Databases and the California Office of Statewide Health Planning and Development. Setting Hospital and ED encounters. Participants Population-based study of California and Florida state residents. Exposure Implementation of Medicaid expansion component of ACA in California in 2014. Main outcomes or measures Changes in ED visits and hospitalizations by payer, percentage of patients hospitalized after an ED encounter, top diagnostic categories for ED and hospital encounters. Results In California, Medicaid ED visits increased 33% after Medicaid expansion implementation and self-pay visits decreased by 25% compared with a 5.7% increase in the rate of Medicaid patient ED visits and a 5.1% decrease in rate of self-pay patient visits in Florida. In addition, California experienced a 15.4% increase in Medicaid inpatient stays and a 25% decrease in self pay stays. Trends in the percentage of patients admitted to the hospital from the ED were notable; a 5.4% decrease in hospital admissions originating from the ED in California, and a 2.1% decrease in Florida from 2013 to 2014. Conclusions and relevance We observed a significant shift in payer for ED visits and hospitalizations after Medicaid expansion in California without a significant change in top diagnoses or overall rate of these ED visits and hospitalizations. There appears to be a shift in reimbursement burden from patients and hospitals to the government without a dramatic shift in patterns of ED or hospital utilization.

Effects of a Brief Educational Program on Optimization of Fluoroscopy to Minimize Radiation Exposure During Endoscopic Retrograde Cholangiopancreatography

Background & Aims Fluoroscopy during endoscopic retrograde cholangiopancreatography (ERCP) is increasingly performed by therapeutic endoscopists, many of whom have not received formal training in modulating fluoroscopy use to minimize radiation exposure. Exposure to ionizing radiation has significant health consequences for patients and endoscopists. We aimed to evaluate whether a 20‐minute educational intervention for endoscopists would improve use of fluoroscopy and decrease ERCP‐associated exposure to radiation for patients. Methods We collected data from 583 ERCPs, performed in California from June 2010 through November 2012; 331 were performed at baseline and 252 following endoscopist education. The educational intervention comprised a 20‐minute video explaining best practices for fluoroscopy, coupled with implementation of a formal fluoroscopy time‐out protocol before the ERCP was performed. Our primary outcome was the effect of the educational intervention on direct and surrogate markers of patient radiation exposure associated with ERCPs performed by high‐volume endoscopists (HVEs) (200 or more ERCPs/year) vs low‐volume endoscopists (LVEs) (fewer than 200 ERCPs/year). Results At baseline, total radiation dose and dose area product were significantly higher for LVEs, but there was no significant difference between HVEs and LVEs following education. Education was associated with significant reductions in median fluoroscopy time (48% reduction for HVEs vs 30% reduction for LVEs), total radiation dose (28% reduction for HVEs vs 52% for LVEs) and dose area product (35% reduction for HVEs vs 48% reduction for LVEs). All endoscopists significantly increased their use of low magnification and collimation following education. Conclusions A 20‐minute educational program with emphasis on ideal use of modifiable fluoroscopy machine settings results in an immediate and significant reduction in ERCP‐associated patient radiation exposure for low‐volume and high‐volume endoscopists. Training programs should consider radiation education for advanced endoscopy fellows.

A case of Meningococcal Pyomyositis in an Otherwise Healthy Adult

The clinical spectrum of Neisseria meningitidis can range from nasopharyngeal colonization to life-threatening invasive diseases such as meningitis. However, its etiologic role in invasive pyomyositis (PM) has never been reported before in the English language. In this study, we report the first case of PM in the English language and the second case in the literature caused by N meningitidis.

Plastic Surgery: Cholangioscopic Intra-stent Balloon Retrieval of a Proximally Migrated Biliary Stent

A 41-year-old man with a benign bile duct stricture was initially evaluated at our institution for endoscopic retrograde cholangiopancreatography (ERCP) with biliary stent exchange following plastic stent placement at an outside facility 2 months previously. His past medical history was notable for a motor vehicle accident resulting in abdominal crush injury and portal vein thrombosis. He then developed portal hypertension complicated by bleeding esophageal and gastric varices managed by splenectomy, with the creation of a mesocaval shunt. Due to subsequent stenosis of the mesocaval shunt, he underwent portal vein thrombectomy and remained on warfarin therapy. He additionally developed portal biliopathy with a distal common bile duct stricture managed by stent placement. Vital signs at the time of his procedure were notable for a heart rate of 82 beats/min, temperature of 37.2 C, blood pressure of 101/72 mm of Hg, and respiratory rate of 11/min. Physical examination revealed healed incisions consistent with his surgical history and mild right upper quadrant abdominal tenderness. His laboratory studies, including liver function tests, were unremarkable. At ERCP, the patient’s ampulla was located in a poorly accessible portion in the distal second part of the duodenum. There was no evidence of a biliary stent protruding into the ampulla. Fluoroscopy located a stent in the right upper quadrant consistent with proximal migration of the stent into the bile duct (Fig. 1). Several standard techniques typically utilized for the retrieval of proximally migrated stents were sequentially employed preceded by extending the existing sphincterotomy. Multiple attempts to grasp the stent with rat-tooth forceps (miniand full-sized) under fluoroscopic guidance were unsuccessful. Efforts at capture with a polypectomy snare, a basket, and a stone extraction balloon also failed. Attempts to cannulate and retrieve the stent using Soehendra stent extractors were similarly unsuccessful. Due to the length of the procedure, a decision was made to terminate with a plan to attempt cholangioscopy-assisted stent retrieval at a follow-up ERCP. An 8.5-Fr plastic stent was then placed alongside the proximally migrated stent. At the time of repeat ERCP for stent exchange 11 weeks later, the most recently placed stent was removed and the proximally migrated stent was still evident on fluoroscopy. A Spyglass cholangioscope (Boston Scientific, Marlborough, MA, USA) was advanced into the bile duct; the migrated stent’s distal end was embedded in the lateral wall of the distal bile duct with the proximal end impacted at the bile duct bifurcation, complicating attempts to directly cannulate the stent. The distal flange of the stent was grasped with a Spybite forceps, but due to the low closure force inherent with mini-forceps, the device slid off with each attempt to dislodge the embedded stent (Fig. 2a). A 0.03500 guidewire was then advanced through the cholangioscope into the stent’s flange side hole and through the stent under visual and fluoroscopic guidance. A 4 mm 9 2 cm biliary stone extraction balloon was advanced into the stent through the side flange, inflated, and then used to push and dislodge the embedded distal tip of the stent. Once the stent was floating freely in the bile duct, the cholangioscope was used to direct the 0.03500 guidewire into the stent through its distal tip (Fig. 2). The & Subhas Banerjee sbanerje@stanford.edu