Rho Inhibitor VX-210 in Acute Traumatic Subaxial Cervical Spinal Cord Injury: Design of the SPinal Cord Injury Rho INhibition InvestiGation (SPRING) Clinical Trial.

Traumatic spinal cord injury (SCI) is associated with a lifetime of disability stemming from loss of motor, sensory, and autonomic functions; these losses, along with increased comorbid sequelae, negatively impact health outcomes and quality of life. Early decompression surgery post-SCI can enhance patient outcomes, but does not directly facilitate neural repair and regeneration. Currently, there are no U.S. Food and Drug Administration-approved pharmacological therapies to augment motor function and functional recovery in individuals with traumatic SCI. After an SCI, the enzyme, Rho, is activated by growth-inhibitory factors and regulates events that culminate in collapse of the neuronal growth cone, failure of axonal regeneration, and, ultimately, failure of motor and functional recovery. Inhibition of Rho activation is a potential treatment for injuries such as traumatic SCI. VX-210, an investigational agent, inhibits Rho. When administered extradurally after decompression (corpectomy or laminectomy) and stabilization surgery in a phase 1/2a study, VX-210 was well tolerated. Here, we describe the design of the SPRING trial, a multicenter, phase 2b/3, randomized, double-blind, placebo-controlled clinical trial to evaluate the efficacy and safety of VX-210 (NCT02669849). A subset of patients with acute traumatic cervical SCI is currently being enrolled in the United States and Canada. Medical, neurological, and functional changes are evaluated at 6 weeks and at 3, 6, and 12 months after VX-210 administration. Efficacy will be assessed by the primary outcome measure, change in upper extremity motor score at 6 months post-treatment, and by secondary outcomes that include question-based and task-based evaluations of functional recovery.

Get Real: Evaluation of a Community-Level HIV Prevention Intervention for Young MSM Who Engage in Episodic Substance Use.

Young men who have sex with men (YMSM) have high rates of recreational drug use and binge drinking that are related to increases in unprotected sex and HIV risk. We describe the development of a community-level intervention focused on MSM ages 15 to 29 who identify as Black or White and who reported episodic use of alcohol and/or drugs. Intervention content included culturally-relevant role model stories and peer outreach. Outcome measures, including number of partners and a risk score indicating risk for HIV from protected or unprotected insertive or receptive anal sex, were examined over 36 months in a sample of YMSM in Philadelphia, the intervention site, and Baltimore, the comparison site. Decreases in risk scores over time were significantly larger in the intervention sample than in the comparison sample. Effects were greater for YMSM who engaged in binge drinking and/or marijuana use, compared to those who used other drugs.

Two pathways regulate cortical granule translocation to prevent polyspermy in mouse oocytes.

An egg must be fertilized by a single sperm only. To prevent polyspermy, the zona pellucida, a structure that surrounds mammalian eggs, becomes impermeable upon fertilization, preventing the entry of further sperm. The structural changes in the zona upon fertilization are driven by the exocytosis of cortical granules. These translocate from the oocyte's centre to the plasma membrane during meiosis. However, very little is known about the mechanism of cortical granule translocation. Here we investigate cortical granule transport and dynamics in live mammalian oocytes by using Rab27a as a marker. We show that two separate mechanisms drive their transport: myosin Va-dependent movement along actin filaments, and an unexpected vesicle hitchhiking mechanism by which cortical granules bind to Rab11a vesicles powered by myosin Vb. Inhibiting cortical granule translocation severely impaired the block to sperm entry, suggesting that translocation defects could contribute to miscarriages that are caused by polyspermy.

Rho kinase as a target for cerebral vascular disorders.

The development of novel pharmaceutical treatments for disorders of the cerebral vasculature is a serious unmet medical need. These vascular disorders are typified by a disruption in the delicate Rho signaling equilibrium within the blood vessel wall. In particular, Rho kinase overactivation in the smooth muscle and endothelial layers of the vessel wall results in cytoskeletal modifications that lead to reduced vascular integrity and abnormal vascular growth. Rho kinase is thus a promising target for the treatment of cerebral vascular disorders. Indeed, preclinical studies indicate that Rho kinase inhibition may reduce the formation/growth/rupture of both intracranial aneurysms and cerebral cavernous malformations.

Cervical spinal cord injury: tailoring clinical trial endpoints to reflect meaningful functional improvements.

Cervical spinal cord injury (SCI) results in partial to full paralysis of the upper and lower extremities. Traditional primary endpoints for acute SCI clinical trials are too broad to assess functional recovery in cervical subjects, raising the possibility of false positive outcomes in trials for cervical SCI. Endpoints focused on the recovery of hand and arm control (e.g., upper extremity motor score, motor level change) show the most potential for use as primary outcomes in upcoming trials of cervical SCI. As the field moves forward, the most reliable way to ensure meaningful clinical testing in cervical subjects may be the development of a composite primary endpoint that measures both neurological recovery and functional improvement.

Hypermasculinity and Sexual Risk Among Black and White Men Who Have Sex With Men and Women.

Men who have sex with men and women (MSMW), particularly Black MSMW, are at high risk for HIV. However, few studies have focused on factors that influence Black MSMW's specific HIV risk behaviors, and there are no evidence-based interventions specifically targeting this population. Some studies have suggested that masculine ideals are associated with high-risk sex practices and partners. Norms around masculinity in the social environments in which MSMW live may prohibit nonheterosexual identities and behaviors, may lead to internalized homophobia, and may promote high-risk strategies to seek male partners. Using data collected from 180 Black and 101 White MSMW recruited for a study to develop strategies for recruiting MSMW for research and services and to inform the content of HIV prevention messages, we examined the association between hypermasculinity ideals and sexual behaviors that may contribute to increased HIV risk among Black MSMW and a comparison group of White MSMW. Comparing Black and White MSMW, we explored how this association may differ by race. Multivariate models, controlling for sociodemographic and other covariates, indicate that hypermasculine ideals are associated with increased numbers of male and female partners among Black MSMW and an increased number of female partners among White MSMW. Hypermasculinity is important to address in programs that aim to reduce HIV risk among Black MSMW.

Are managed care organizations in the United States impeding the delivery of primary care by nurse practitioners? A 2012 update on managed care organization credentialing and reimbursement practices.

In 2014, the Affordable Care Act will create an estimated 16 million newly insured people. Coupled with an estimated shortage of over 60,000 primary care physicians, the country's public health care system will be at a challenging crossroads, as there will be more patients waiting to see fewer doctors. Nurse practitioners (NPs) can help to ease this crisis. NPs are health care professionals with the capability to provide important and critical access to primary care, particularly for vulnerable populations. However, despite convincing data about the quality of care provided by NPs, many managed care organizations (MCOs) across the country do not credential NPs as primary care providers, limiting the ability of NPs to be reimbursed by private insurers. To assess current credentialing practices of health plans across the United States, a brief telephone survey was administered to 258 of the largest health maintenance organizations (HMOs) in the United States, operated by 98 different MCOs. Results indicated that 74% of these HMOs currently credential NPs as primary care providers. Although this represents progress over prior assessments, findings suggest that just over one fourth of major HMOs still do not recognize NPs as primary care providers. Given the documented shortage of primary care physicians in low-income communities in the United States, these credentialing policies continue to diminish the ability of NPs to deliver primary care to vulnerable populations. Furthermore, these policies could negatively impact access to care for thousands of newly insured Americans who will be seeking a primary care provider in 2014.

Functional roles for myosin 1c in cellular signaling pathways.

Cellular signaling pathways underlie the transfer of information throughout the cell and to adjoining cells and so govern most critical cellular functions. Increasing evidence points to the molecular motor myosin 1c as a prominent player in many signaling cascades, from the integrin-dependent signaling involved in cell migration to the signaling events underlying insulin resistance. Myosin 1c functions on these pathways both via an important role in regulating lipid raft recycling and also via direct involvement in signaling cascades. This review provides an overview of the functional involvement of myosin 1c in cellular signaling and discusses the possible potential for myosin 1c as a target for drug-based treatments for human diseases.

Small-molecule inhibitors of myosin proteins.

Advances in screening and computational methods have enhanced recent efforts to discover/design small-molecule protein inhibitors. One attractive target for inhibition is the myosin family of motor proteins. Myosins function in a wide variety of cellular processes, from intracellular trafficking to cell motility, and are implicated in several human diseases (e.g., cancer, hypertrophic cardiomyopathy, deafness and many neurological disorders). Potent and selective myosin inhibitors are, therefore, not only a tool for understanding myosin function, but are also a resource for developing treatments for diseases involving myosin dysfunction or overactivity. This review will provide a brief overview of the characteristics and scientific/therapeutic applications of the presently identified small-molecule myosin inhibitors before discussing the future of myosin inhibitor and activator design.

Dynamic exchange of myosin VI on endocytic structures.

The actin-based molecular motor myosin VI functions in the endocytic uptake pathway, both during the early stages of clathrin-mediated uptake and in later transport to/from early endosomes. This study uses fluorescence recovery after photobleaching (FRAP) to examine the turnover rate of myosin VI during endocytosis. The results demonstrate that myosin VI turns over dynamically on endocytic structures with a characteristic half-life common to both the large insert isoform of myosin VI on clathrin-coated structures and the no-insert isoform on early endosomes. This half-life is shared by the myosin VI-binding partner Dab2 and is identical for full-length myosin VI and the cargo-binding tail region. The 4-fold slower half-life of an artificially dimerized construct of myosin VI on clathrin-coated structures suggests that wild type myosin VI does not function as a stable dimer, but either as a monomer or in a monomer/dimer equilibrium. Taken together, these FRAP results offer insight into both the basic turnover dynamics and the monomer/dimer nature of myosin VI.

Kinetic properties and small-molecule inhibition of human myosin-6.

Myosin-6 is an actin-based motor protein that moves its cargo towards the minus-end of actin filaments. Mutations in the gene encoding the myosin-6 heavy chain and changes in the cellular abundance of the protein have been linked to hypertrophic cardiomyopathy, neurodegenerative diseases, and cancer. Here, we present a detailed kinetic characterization of the human myosin-6 motor domain, describe the effect of 2,4,6-triiodophenol on the interaction of myosin-6 with F-actin and nucleotides, and show how addition of the drug reduces the number of myosin-6-dependent vesicle fusion events at the plasma membrane during constitutive secretion.

Myosin motor proteins are involved in the final stages of the secretory pathways.

In eukaryotes, the final steps in both the regulated and constitutive secretory pathways can be divided into four distinct stages: (i) the 'approach' of secretory vesicles/granules to the PM (plasma membrane), (ii) the 'docking' of these vesicles/granules at the membrane itself, (iii) the 'priming' of the secretory vesicles/granules for the fusion process, and, finally, (iv) the 'fusion' of vesicular/granular membranes with the PM to permit content release from the cell. Recent work indicates that non-muscle myosin II and the unconventional myosin motor proteins in classes 1c/1e, Va and VI are specifically involved in these final stages of secretion. In the present review, we examine the roles of these myosins in these stages of the secretory pathway and the implications of their roles for an enhanced understanding of secretion in general.

Myosin VI and its binding partner optineurin are involved in secretory vesicle fusion at the plasma membrane.

During constitutive secretion, proteins synthesized at the endoplasmic reticulum (ER) are transported to the Golgi complex for processing and then to the plasma membrane for incorporation or extracellular release. This study uses a unique live-cell constitutive secretion assay to establish roles for the molecular motor myosin VI and its binding partner optineurin in discrete stages of secretion. Small interfering RNA-based knockdown of myosin VI causes an ER-to-Golgi transport delay, suggesting an unexpected function for myosin VI in the early secretory pathway. Depletion of myosin VI or optineurin does not affect the number of vesicles leaving the trans-Golgi network (TGN), indicating that these proteins do not function in TGN vesicle formation. However, myosin VI and optineurin colocalize with secretory vesicles at the plasma membrane. Furthermore, live-cell total internal reflection fluorescence microscopy demonstrates that myosin VI or optineurin depletion reduces the total number of vesicle fusion events at the plasma membrane and increases both the proportion of incomplete fusion events and the number of docked vesicles in this region. These results suggest a novel role for myosin VI and optineurin in regulation of fusion pores formed between secretory vesicles and the plasma membrane during the final stages of secretion.

A targeted siRNA screen to identify SNAREs required for constitutive secretion in mammalian cells.

The role of SNAREs in mammalian constitutive secretion remains poorly defined. To address this, we have developed a novel flow cytometry-based assay for measuring constitutive secretion and have performed a targeted SNARE and Sec1/Munc18 (SM) protein-specific siRNA screen (38 SNAREs, 4 SNARE-like proteins and 7 SM proteins). We have identified the endoplasmic reticulum (ER)/Golgi SNAREs syntaxin 5, syntaxin 17, syntaxin 18, GS27, SLT1, Sec20, Sec22b, Ykt6 and the SM protein Sly1, along with the post-Golgi SNAREs SNAP-29 and syntaxin 19, as being required for constitutive secretion. Depletion of SNAP-29 or syntaxin 19 causes a decrease in the number of fusion events at the cell surface and in SNAP-29-depleted cells causes an increase in the number of docked vesicles at the plasma membrane as determined by total internal reflection fluorescence (TIRF) microscopy. Analysis of syntaxin 19-interacting partners by mass spectrometry indicates that syntaxin 19 can form SNARE complexes with SNAP-23, SNAP-25, SNAP-29, VAMP3 and VAMP8, supporting its role in Golgi to plasma membrane transport or fusion. Surprisingly, we have failed to detect any requirement for a post-Golgi-specific R-SNARE in this process.

Magic Johnson doesn't worry about how to pay for medicine: experiences of black men who have sex with men living with HIV.

Despite high and rapidly growing incidence of HIV, little is known about the everyday lived experiences of HIV-positive black men who have sex with men. Lack of empirical knowledge about members of this group is especially problematic as HIV-positive individuals continue to live in a world of hope, fear, waiting and wondering, which can heavily influence their everyday lives. In this exploratory study, we examine the everyday lives of HIV-positive black gay, bisexual and other men who have sex with men, particularly how being a racial minority may influence the ways that they manage living with the illness. Our goal was to provide a forum from which black men could share their personal experiences regarding the various aspects of living with HIV. In doing so, we identified five themes that may be unique to black men or experienced differently by black men due, in the USA, to their racial minority status.

Black men who have sex with men and the association of down-low identity with HIV risk behavior.

Black men "on the down low" have been considered prime agents of HIV transmission in the Black community despite little empirical evidence. We assessed the relationship between down-low identification and sexual risk outcomes among 1151 Black MSM. Down-low Identification was not associated with unprotected anal or vaginal sex with male or female partners. Future HIV prevention programs and research should target sexual risk behaviors of Black men, irrespective of identity, and not focus on the "down low."

Understanding differences in HIV sexual transmission among Latino and black men who have sex with men: The Brothers y Hermanos Study.

HIV sexual transmission risk behaviors were examined among 1,065 Latino and 1,140 black men who have sex with men (MSM). Participants completed a computer-administered questionnaire and were tested for HIV infection. Of men who reported that their last HIV test was negative or that they had never been tested or did not get the result of their last test, 17% of black and 5% of Latino MSM tested HIV-positive in our study. In both ethnic groups, the three-month prevalence of unprotected anal intercourse (UAI) with HIV-negative or unknown serostatus partners was twice as high among men unaware of their HIV infection than men who knew they were HIV seropositive at the time of enrollment. UAI exclusively with HIV-positive partners was more prevalent among HIV-positive/aware than HIV-positive/unaware men. The findings advance understanding of the high incidence of HIV infection among black MSM in the U.S.

Sexual risk behaviors of HIV-positive, HIV-negative, and serostatus-unknown Black men who have sex with men and women.

Black men who have sex with men and women (MSMW) are at high risk for HIV infection and transmission. This study compared the sexual risk behaviors of Black MSMW who self-reported being HIV-positive with those who reported being HIV-negative and those who did not know their HIV status. Respondent-driven sampling (RDS) was used to recruit 1,154 Black MSM in Philadelphia and New York who completed an audio computer-assisted self-interview (ACASI). Of these men, 212 had engaged in anal sex with male partners and vaginal or anal sex with female partners in the past 3 months. A quarter (23.6%; n = 50) of MSMW self-reported testing positive for HIV at their last test, 59.4% (n = 126) reported testing negative for HIV at their last test, and 17.0% (n = 36) reported never having an HIV test. Multivariate logistic regression analysis revealed that HIV-positive MSMW were much less likely than HIV-negative men and never-tested men to have engaged in unprotected intercourse with main male and main female partners perceived to be HIV-negative or of unknown serostatus. However, HIV-positive men were equally as likely as HIV-negative men to have unprotected intercourse with non-main male and non-main female partners perceived as HIV-negative or of unknown serostatus. Our findings indicate that some HIV-positive MSMW engage in unprotected sex that places female and male partners at risk for HIV infection. However, MSMW who have never taken an HIV test, or who have not been recently tested, may be a greater source of HIV transmission to their female and male partners.

Pericentric chromatin is organized into an intramolecular loop in mitosis.

BACKGROUND: Cohesin proteins link sister chromatids and provide the basis for tension between bioriented sister chomatids in mitosis. Cohesin is concentrated at the centromere region of the chromosome despite the fact that sister centromeres can be separated by 800 nm in vivo. The function of cohesin at sites of separated DNA is unknown. RESULTS: We provide evidence that the kinetochore promotes the organization of pericentric chromatin into a cruciform in mitosis such that centromere-flanking DNA adopts an intramolecular loop, whereas sister-chromatid arms are paired intermolecularly. Visualization of cohesin subunits by fluorescence microscopy revealed a cylindrical structure that encircles the central spindle and spans the distance between sister kinetochores. Kinetochore assembly at the apex of the loop initiates intrastrand loop formation that extends approximately 25 kb (12.5 kb on either side of the centromere). Two centromere loops (one from each sister chromatid) are stretched between the ends of sister-kinetochore microtubules along the spindle axis. At the base of the loop there is a transition to intermolecular sister-chromatid pairing. CONCLUSIONS: The C loop conformation reveals the structural basis for sister-kinetochore clustering in budding yeast and for kinetochore biorientation and thus resolves the paradox of maximal interstrand separation in regions of highest cohesin concentration.

Decisional balance, perceived risk and HIV testing practices.

Improving our understanding of how individuals decide to take an HIV test is essential for designing effective programs to increase testing. This paper assesses the relationship of decisional balance and perceived risk to HIV testing history in a cross-sectional community sample of 1523 HIV-negative men and women at risk due to drug use or sexual behavior. We developed scales to measure perceived advantages (pros) and perceived disadvantages (cons) of taking an HIV test and assessed their content using factor analysis. Perceived risk was highly related to the pros and cons scales. Multivariate analyses revealed that the pros scale had positive associations with having ever tested and the number of tests taken, while the cons scale had negative associations with these testing measures. Perceived risk was not related to testing practices. These results suggest that interventions to increase HIV testing need to address anticipated positive and negative outcomes of getting tested.