Extracellular acidosis enhances Zika virus infection both in human cells and ex-vivo tissue cultures from female reproductive tract.

Zika virus (ZIKV) is a flavivirus transmitted by mosquitoes of the genus Aedes, but unlike other flaviviruses, ZIKV can be sexually transmitted by vaginal intercourse. The healthy vaginal pH ranges from 4.0 to 6.0, reaching values of 6.0-7.0 after semen deposition. Here, we report that low extracellular pH values (range 6.2-6.6) dramatically increase ZIKV infection on cell lines of different origin including some derived from the female genital tract and monocyte-derived macrophages. Furthermore, low pH significantly increased ZIKV infection of human ectocervix and endocervix cultured ex-vivo. Enhancement of infection by low pH was also observed using different ZIKV strains and distinct methods to evaluate viral infection, i.e. plaque assays, RT-PCR, flow cytometry, and fluorescence microscopy. Analysis of the mechanisms involved revealed that the enhancement of ZIKV infection induced by low pH was associated with increased binding of the viral particles to the heparan sulphate expressed on the target cell surface. Acidosis represents a critical but generally overlooked feature of the female genital tract, with major implications for sexual transmission diseases. Our results suggest that low vaginal pH might promote male-to-female transmission of ZIKV infection.

Impact of colonoscopic screening in male and female Lynch syndrome carriers with an MSH2 mutation.

The lifetime risk of developing colorectal cancer (CRC) in Lynch syndrome (LS) carriers is very high. To determine the impact of colonoscopic screening in 54 male and 98 female MSH2 mutation carriers, outcomes were compared with 94 males and 76 females who were not screened. CRC incidence and survival in the screened group were compared to that expected, derived from the non-screened group. To correct for survivor bias, controls were matched for age at entry into screening and also for gender. In males, median age to CRC was 58 years, whereas expected was 47 years (p = 0.000), and median survival was 66 years vs 62 years (p = 0.034). In screened females, median age to CRC was 79 years compared to 57 years in the non-screened group (p = 0.000), and median survival was 80 years compared with expected of 63 years (p = 0.001). Twenty percent of males and 7% of females developed an interval CRC within 2 years of previous colonoscopy. Although colonoscopic screening was associated with decreased CRC risk and better survival, CRCs continued to occur. CRC development may be further reduced by decreasing the screening interval to 1 year and improving quality of colonoscopy.

HIV-1 sexual transmission: early events of HIV-1 infection of human cervico-vaginal tissue in an optimized ex vivo model.

Infection and dissemination of human immunodeficiency virus (HIV)-1 through the female body after vaginal intercourse depends on the activation/differentiation status of mucosal CD4 T cells. In this study, we investigated this status and the susceptibility to HIV-1 infection of human cervico-vaginal tissue ex vivo. We found that virtually all T cells are of the effector memory phenotype with broad CC chemokine receptor 5 (CCR5) expression. As it does in vivo, human cervico-vaginal tissue ex vivo preferentially supports the productive infection of R5 HIV-1 rather than that of X4 HIV-1 in spite of the broad expression of CXC chemokine receptor 4 (CXCR4). X4 HIV-1 replicated only in the few tissues that were enriched in CD27(+)CD28(+) effector memory CD4 T cells. Productive infection of R5 HIV-1 occurred preferentially in activated CD38(+)CD4 T cells and was followed by a similar activation of HIV-1-uninfected (bystander) CD4 T cells that may amplify viral infection. These results provide new insights into the dependence of HIV-1 infection and dissemination on the activation/differentiation of cervico-vaginal lymphocytes.

CNS stem and progenitor cell differentiation into functional neuronal circuits in three-dimensional collagen gels.

The mammalian central nervous system (CNS) has little capacity for self-repair after injury, and neurons are not capable of proliferating. Therefore, neural tissue engineering that combines neural stem and progenitor cells and biologically derived polymer scaffolds may revolutionize the medical approach to the treatment of damaged CNS tissues. Neural stem and progenitor cells isolated from embryonic rat cortical or subcortical neuroepithelium were dispersed within type I collagen, and the cell-collagen constructs were cultured in serum-free medium containing basic fibroblast growth factor. The collagen-entrapped stem and progenitors actively expanded and efficiently generated neurons, which developed neuronal polarity, neurotransmitters, ion channels/receptors, and excitability. Ca2+ imaging showed that differentiation from BrdU+/TuJ1- to BrdU-/TuJ1+ cells was accompanied by a shift in expression of functional receptors for neurotransmitters from cholinergic and purinergic to predominantly GABAergic and glutamatergic. Spontaneous postsynaptic currents were recorded by patch-clamping from precursor cell-derived neurons and these currents were partially blocked by 10-microM bicuculline, and completely blocked by additional 10 microM of the kainate receptor antagonist CNQX, indicating an appearance of both GABAergic and glutamatergic synaptic activities. Staining with endocytotic marker FM1-43 demonstrated active synaptic vesicle recycling occurring among collagen-entrapped neurons. These results show that neural stem and progenitor cells cultured in 3D collagen gels recapitulate CNS stem cell development; this is the first demonstration of CNS stem and progenitor cell-derived functional synapse and neuronal network formation in a 3D matrix. The proliferative capacity and neuronal differentiating potential of neural progenitors in 3D collagen gels suggest their potential use in attempts to promote neuronal regeneration in vivo.

Dissolved elemental mercury investigations in Long Island Sound using on-line Au amalgamation-flow injection analysis.

A novel semiautomatic dissolved elemental mercury analyzer (DEMA) was developed for investigating dissolved elemental Hg (DEM) in natural waters. This on-line setup couples the main analytical steps from sample introduction, gas-liquid separation, and Au amalgamation/separation to final detection/data acquisition using flow injection techniques. This approach provides ease of operation and high analytical performance and is suitable for shipboard use. The analyzer can be fully automated and also be modified to examine other Hg species (e.g., reactive and total Hg and monomethyl-Hg). Here, we present the results of laboratory performance tests and make a comparison with a traditional manual method. DEM measured by both manual and the DEMA show good agreement. Representative field DEM data from spring and summer 1999 in Long Island Sound, U.S.A. (LIS) are presented. Spatial and temporal DEM variations were evident. Rapid and accurate determinations of DEM are necessary to observe its distribution dynamics, evaluate emissions, and assess its role in the aquatic biogeochemical cycling of Hg.

Suppression of CCR5- but not CXCR4-tropic HIV-1 in lymphoid tissue by human herpesvirus 6.

HIV-1 infects target cells via a receptor complex formed by CD4 and a chemokine receptor, primarily CCR5 or CXCR4 (ref. 1). Commonly, HIV-1 transmission is mediated by CCR5-tropic variants, also designated slow/low, non-syncytia-inducer or macrophage-tropic, which dominate the early stages of HIV-1 infection and frequently persist during the entire course of the disease. In contrast, HIV-1 variants that use CXCR4 are typically detected at the later stages, and are associated with a rapid decline in CD4+ T cells and progression to AIDS (refs. 2,7-11). Disease progression is also associated with the emergence of concurrent infections that may affect the course of HIV disease by unknown mechanisms. A lymphotropic agent frequently reactivated in HIV-infected patients is human herpesvirus 6 (HHV-6), which has been proposed as a cofactor in AIDS progression. Here we show that in human lymphoid tissue ex vivo, HHV-6 affects HIV-1 infection in a coreceptor-dependent manner, suppressing CCR5-tropic but not CXCR4-tropic HIV-1 replication, as shown with both uncloned viral isolates and isogenic molecular chimeras. Furthermore, we demonstrate that HHV-6 increases the production of the CCR5 ligand RANTES ('regulated upon activation, normal T-cell expressed and secreted'), the most potent HIV-inhibitory CC chemokine, and that exogenous RANTES mimics the effects of HHV-6 on HIV-1, providing a mechanism for the selective blockade of CCR5-tropic HIV-1. Our data suggest that HHV-6 may profoundly influence the course of HIV-1 infection.

Bayesian restoration of ion channel records using hidden Markov models.

Hidden Markov models have been used to restore recorded signals of single ion channels buried in background noise. Parameter estimation and signal restoration are usually carried out through likelihood maximization by using variants of the Baum-Welch forward-backward procedures. This paper presents an alternative approach for dealing with this inferential task. The inferences are made by using a combination of the framework provided by Bayesian statistics and numerical methods based on Markov chain Monte Carlo stochastic simulation. The reliability of this approach is tested by using synthetic signals of known characteristics. The expectations of the model parameters estimated here are close to those calculated using the Baum-Welch algorithm, but the present methods also yield estimates of their errors. Comparisons of the results of the Bayesian Markov Chain Monte Carlo approach with those obtained by filtering and thresholding demonstrate clearly the superiority of the new methods.

Formation of artifact methylmercury during extraction from a sediment reference material.

We examined monomethylmercury (MMHg) in sediment reference material IAEA-405 and sediments from Long Island Sound. MMHg was extracted by both aqueous distillation and leaching with dilute solutions of nitric acid. MMHg was formed from both ambient and added inorganic mercury in extracts of IAEA-405 by either technique. Artifact MMHg in IAEA-405 was related linearly with ambient reactive mercury (HgR) in leachates having > 1.2 M acid, but little or no artifact was measured in less acidic extracts. Addition of potassium chloride enhanced extraction of HgR from LAEA-405 but had no effect on artifact MMHg for each leachate molarity tested. Mercury methylation occurred in solution and was a function of both the availability of HgR and a methylation potential, being limited by HgR in less acidic solutions (0.5-1.2 M) and by the methylation potential in more acidic ones (> 1.2 M). Formation of artifact MMHg in Long Island Sound sediments was inconclusive, but additions of inorganic mercury demonstrated that a potential exists. A potential for abiotic mercury methylation seems always present in sediment, and the availability of HgR appears to control MMHg production under environmental conditions. Abiotic methylation of mercury may occur in environs where the reactivity of inorganic mercury is enhanced, such as river-seawater mixing zones.

Explaining the variety of human sexuality.

In this paper, the author formulates a theory to explain why human sexual orientation seems to run amok. The 'psychic instrument', as he terms it, is the baby's dreaming mind which interprets or misinterprets input from its sociocultural sexual environment. The baby, already born an omnisexual being, then develops a fantasy life with socially sanctioned or unsanctioned fetishes which are likely to be expressed when certain triggering situations arise.

Legal and ethical considerations in the treatment of psychosis.

A variety of legal and ethical issues surround any decision about the treatment of patients with psychosis. These issues have come to the forefront with the introduction of the atypical antipsychotic agents. The law defines the minimum expected level of conduct for a health care professional, and where the law ends, ethics begin. Adverse drug reactions are a leading cause of death in the United States, and medication error is a common reason for liability claims against health care professionals. Patients alleging negligence must prove that the health care professional owed a duty to the patient, that the duty was breached, that the patient was injured, and that the breach of duty was the legal cause of the injury. Professional ethics are governed by various models for ethical decision making. The principles model, which can be readily applied to the patient with mental illness, is based on the ethical principles of beneficence, nonmaleficence, autonomy, utility, and justice.

CD4(+) T-lymphocyte depletion in human lymphoid tissue ex vivo is not induced by noninfectious human immunodeficiency virus type 1 virions.

We tested infectious human immunodeficiency virus type 1 (HIV-1), noninfectious but conformationally authentic inactivated whole HIV-1 virions, and purified gp120 for the ability to induce depletion of CD4(+) T cells in human lymphoid tissues ex vivo. Infectious CXCR4-tropic HIV-1, but not matched inactivated virions or gp120, mediated CD4(+) T-cell depletion, consistent with mechanisms requiring productive infection.

The biogeochemistry of an ombrotrophic bog: evaluation of use as an archive of atmosphere mercury deposition.

The utility of ombrotrophic bogs as archives of atmospheric mercury deposition was assessed with an investigation in Arlberg Bog, Minnesota, U.S.A. Since the use of ombrotrophic bogs as archives depends on the immobility of deposited trace metals, we examined the postdepositional transport processes revealed by the solid-phase distributions of mercury and ancillary metals (Fe, Al, Mn, and Pb) in this bog. We modeled metal speciation in bog porewaters as a function of pe in order to understand metal behavior in ombrotrophic peat. Specifically, we considered the effect of water movement and resultant shifts in redox potential gradients on metal retention. Our results indicate that Hg and Pb are immobile in ombrotrophic peat, so their distribution can be used to determine temporal changes in deposition. To substantiate the deposition estimates determined in this study, we emphasized the importance of confirming the validity of the dating scheme, assessing the degree of horizontal homogeneity in the accumulation record, and providing evidence for retention of Hg based on geochemical modeling. As recorded in Arlberg Bog, historic atmospheric Hg deposition increased gradually after the mid-1800s, peaked between 1950 and 1960, and may have declined thereafter. Preindustrial deposition was about 4 micrograms/m2 year and recent deposition about 19 micrograms/m2 year. The results of this study indicate that deposition at Arlberg Bog has been influenced by a regional and/or local-scale source.

Evidence for the HIV-1 phenotype switch as a causal factor in acquired immunodeficiency.

Both cellular and humoral immunodeficiency develop in vivo after prolonged infection with HIV-1, but the mechanisms are unclear. Initial infection with HIV-1 is transmitted by macrophage (M)-tropic/non-syncytia-inducing (NSI) viruses, which hyperactivate the immune system, and, in one view, cause immunodeficiency by "exhaustion" of lymphoid tissue. An alternative hypothesis is that immunodeficiency is caused by the replacement of M-tropic viruses by T cell (T)-tropic/syncytia-inducing (SI) viruses, which are known to be highly cytopathic in vitro and emerge late in infected individuals around the time of transition to AIDS (refs. 1, 7-9). To test these two possibilities, we have developed an ex vivo model of humoral immunity to recall antigens using human lymphoid tissue. This tissue supports productive infection with both M- and T-tropic HIV-1 isolates when cultured ex vivo. We found that specific immune responses were enhanced by productive infection of the tissue with M-tropic/NSI HIV-1 isolates, but were blocked by T-tropic/SI HIV-1 isolates. The mechanism involves specific irreversible effect on B-cell activity. Our results support the hypothesis that the phenotype switch to T-tropic viruses is a key determinant of acquired humoral immunodeficiency in patients infected with HIV.

Lymphocyte trafficking and HIV infection of human lymphoid tissue in a rotating wall vessel bioreactor.

The pathogenesis of HIV infection involves a complex interplay between both the infected and noninfected cells of human lymphoid tissue, the release of free viral particles, the de novo infection of cells, and the recirculatory trafficking of peripheral blood lymphocytes. To develop an in vitro model for studying these various aspects of HIV pathogenesis we have utilized blocks of surgically excised human tonsils and a rotating wall vessel (RWV) cell culture system. Here we show that (1) fragments of the surgically excised human lymphoid tissue remain viable and retain their gross cytoarchitecture for at least 3 weeks when cultured in the RWV system; (2) such lymphoid tissue gradually shows a loss of both T and B cells to the surrounding growth medium; however, this cellular migration is reversible as demonstrated by repopulation of the tissue by labeled cells from the growth medium; (3) this cellular migration may be partially or completely inhibited by embedding the blocks of lymphoid tissue in either a collagen or agarose gel matrix; these embedded tissue blocks retain most of the basic elements of a normal lymphoid cytoarchitecture; and (4) both embedded and nonembedded RWV-cultured blocks of human lymphoid tissue are capable of productive infection by HIV-1 of at least three various strains of different tropism and phenotype, as shown by an increase in both p24 antigen levels and free virus in the culture medium, and by the demonstration of HIV-1 RNA-positive cells inside the tissue identified by in situ hybridization. It is therefore reasonable to suggest that gel-embedded and nonembedded blocks of human lymphoid tissue, cocultured with a suspension of tonsillar lymphocytes in an RWV culture system, constitute a useful model for simulating normal lymphocyte recirculatory traffic and provide a new tool for testing the various aspects of HIV pathogenesis.

Observations on sleeping position and essential hypertension.

A hypertensive black male, at risk for episodic attacks of pseudo-malignant hypertension and self-induced atrial fibrillation, seeks to discover possible clues to the pathogeneses of these strange disorders through self-study and concludes they might be associated with impaired oxygen intake, secondary to sleeping position in bed.

Induction of carcinoembryonic antigen expression in a three-dimensional culture system.

MIP-101 is a poorly differentiated human colon carcinoma cell line established from ascites that produces minimal amounts of carcinoembryonic antigen (CEA), a 180 kDa glycoprotein tumor marker, and nonspecific cross-reacting antigen (NCA), a related protein that has 50 and 90 kDa isoforms, in monolayer culture. However, MIP-101 produces CEA when implanted into the peritoneum of nude mice but not when implanted into subcutaneous tissue. We tested whether three-dimensional (3D) growth was a sufficient stimulus to produce CEA and NCA 50/90 in MIP-101 cells, because cells grow in 3D in vivo rather than in two-dimensions (2D) as occurs in monolayer cultures. To do this, MIP-101 cells were cultured on microcarrier beads in 3D cultures, either in static cultures as nonadherent aggregates or under dynamic conditions in a NASA-designed low shear stress bioreactor. MIP-101 cells proliferated well under all three conditions and increased CEA and NCA production three- to four-fold when grown in 3D cultures compared to MIP-101 cells growing logarithmically in monolayers. These results suggest that 3D growth in vitro simulates tumor function in vivo and that 3D growth by itself may enhance production of molecules that are associated with the metastatic process.

Competency, board certification, credentialing, and specialization: who benefits?

Pharmacists are concerned with the rapid changes in the healthcare system and what the requirements will be for a pharmacist in the near future. The emergence of board certification, credentialing, and other certification programs for pharmacists are causing significant concern among pharmacists. Pharmacists must assess certification programs and decide on the value of certification to their careers and to the patients they serve. Employers of pharmacists and those paying for healthcare and pharmacy services must also evaluate the value of pharmacists certification. Perhaps the most direct and significant benefit of pharmacist certification lies in the ability of the pharmacist to provide better and more comprehensive care to patients or selected groups of patients (eg, diabetic patients). Better and more comprehensive care provided by a pharmacist benefits the patient, other healthcare professionals, the healthcare system generally, and payers of healthcare and pharmacy services. Demonstrated competence of the pharmacist to provide pharmaceutical care is essential to achieving this benefit. Board certification of pharmacists in current board-recognized specialty areas of nutrition support pharmacy, pharmacotherapy, psychiatric pharmacy, nuclear pharmacy, and oncology pharmacy totaled 2075 board certified pharmacists in the United States as of January 1997.

Historical vignette: cerebral cortical stimulation and surgery for epilepsy.

In 1909, in an isolated community hospital, on the northern tip of the Province of Newfoundland and Labrador, Canada, Dr. John Mason Little, Jr. performed electrical stimulation of the cerebral cortex, prior to cortical excision, as treatment of recurrent cerebral seizures in three patients. Extracts from Dr. Little's written records of the clinical features, the neurosurgical procedures and cerebral cortical stimulation are summarised. A brief review of the contemporaneous history of neurosurgical procedures for epilepsy provides a prospective of Dr. Little's remarkable surgical virtuosity.

[MPR14]-rADM(14-50), a novel analog of adrenomedullin, possesses potent vasodilator activity in the hindlimb vascular bed of the cat.

Responses to [Mpr14]-ADM(14-50), a novel analog of adrenomedullin, were investigated in the hindlimb vascular bed of the cat under conditions of controlled blood flow. Intraarterial injections of [Mpr14]-rADM(14-50) in doses of 0.003-1 nmol caused dose-related decreases in hindlimb perfusion pressure. In terms of relative vasodilator activity, [Mpr14]-rADM(14-50) was more potent than human synthetic adrenomedullin (hADM) in doses of 0.003-0.1 nmol. The recovery half-times (T 1/2) for the vasodilator response to [Mpr14]-rADM(14-50) were significantly greater than the recovery half-times for hADM in all doses studied. Decreases in hindlimb perfusion pressure in response to [Mpr14]-rADM(14-50) were not altered by the calcitonin gene-related peptide receptor antagonist rCGRP(8-37) at the same time vasodilator responses to calcitonin gene-related peptide were significantly reduced. The present data demonstrate that [Mpr14]-(14-50) has potent and long-lasting vasodilator activity when compared to hADM, and that vasodilator responses to [Mpr14]-rADM(14-50) are not dependent on the activation of CGRP receptors in the hindlimb vascular bed of the cat.