A potential role for beta- and gamma-crystallins in the vascular remodeling of the eye.

We demonstrate that expression of beta- and gamma-crystallins is associated with intraocular vessels during normal vascular development of the eye and also in the Nuc1 rat, a mutant in which the hyaloid vascular system fails to regress normally. Real-Time RT PCR, Western blot and metabolic labeling studies indicate an increased expression of beta- and gamma-crystallins in Nuc1 retina. The increased expression of crystallins was localized to the astrocytes surrounding the intraocular vessels. A similar pattern of crystallin expression was also observed in the retinal vessels during normal development. Cultured human astrocytes exposed to 3-nitropropionic acid, an established model of neuronal hypoxia, increased VEGF expression, as expected, but also increased expression of crystallins. Our data suggest that crystallins may function together with VEGF during vascular remodeling. Interestingly, in human PFV (persistent fetal vasculature) disease, where the hyaloid vasculature abnormally persists after birth, we show that astrocytes express both VEGF and crystallins.     

Clade analysis and surface antigen polymorphism of hepatitis B virus American genotypes

Eight genotypes (A-H) of hepatitis B virus (HBV) have been described, HBV genotypes F and H being autochthonous to America. HBV genotype F has been classified in four clusters. The objective of this study was to gain insight into the molecular epidemiology of HBV American genotypes, as well as to analyze the genotype-related polymorphism in some functional domains of the surface proteins. The sequences of the S region of 106 isolates genotype F and H were analyzed, out of which 47 isolates genotype F circulated in different Venezuelan populations. Most of the Venezuelan isolates genotype F were grouped in cluster III (n = 39) and 7 in cluster II. One isolate obtained from a blood donor could not be classified in any clade and harbored amino acid substitutions characteristic of a vaccine escape mutant (G145R) and a stop codon in the surface antigen. Amino acid analysis of the PreS and S gene products showed unique genetic characteristics in genotype F and H sequences in some important domains involved in the early steps of infection. Out of 30 available sequences, two complete genome sequences of HBV genotype F from Venezuela were obtained. Phylogenetic analysis of these complete genomes confirmed the presence of four clusters inside genotype F, differing in more than 4% nucleotide divergence. Our extended analysis showed that genotype F clades Ia, III, and IV exhibit a restricted geographic distribution (Central America, the North and the South of South America, respectively) while clades Ib and II are found in all the Americas except in the Northern South America and North America respectively.     

Molecular and serological evaluation of surface antigen negative hepatitis B virus infection in blood donors from Venezuela

Surface antigen negative hepatitis B virus (HBV) infection was evaluated in Venezuela, by molecular characterization of blood samples positive for antibodies to core antigen (anti-HBc) and negative for surface antigen (HBsAg) in blood donors (residual infections). HBV DNA was found in 11/258 samples (4.3%), and was significantly associated with high levels of anti-HBc antibodies (>25 UI/ml, P < 0.05), while no correlation was found between the presence of HBV DNA and the levels of anti-HBs. Synonymous and non-synonymous mutations were found in the HBV surface region (but not vaccine escape mutants) and in the precore/core region (precore mutants in 2/7 samples and 33-45 bp deletions near the N-terminal core region in 4/19 samples). While HBV genotype F prevails among HBsAg positive samples from blood donors in Venezuela, residual infection isolates were mainly genotypes A and D. Phylogenetic analysis of viral surface and core region revealed discrepancies in genotype designation in 6/9 samples, suggesting the presence of mixed infection or recombination. In conclusion, HBV residual infection in Venezuela does not seem to be frequently observed in HBV genotype F. This type of infection is frequently associated with variants exhibiting mutations in the surface gene that might be affecting the correct recognition by commercial tests, with precore mutants and with core internal deletions. These variants do not seem to cause severe liver disease, and on the contrary, were found circulating at low viremia.     

Psychosocial predictors of long-term adjustment to lower-limb amputation and phantom limb pain

PURPOSE: To evaluate the utility of a biopsychosocial model to predict long-term adjustment to lower-limb amputation and phantom limb pain (PLP). METHOD: One month after lower-limb amputation, 70 participants completed measures of PLP intensity, cognitions (catastrophizing, perceived control over pain), coping (pain-contingent rest), social environment (social support, solicitous responding), and functioning (pain interference, depressive symptoms). The measures of functioning were administered again at 1- and 2-years post-amputation. Multiple regression analyses were used to examine the ability of the psychosocial variables at 1-month post-amputation to predict changes in the functioning measures over time. RESULTS: The psychosocial variables at 1-month post-amputation, controlling for initial PLP intensity, accounted for 21% of the variance in change in depressive symptoms at 1-year (p < 0.05), and 27% and 22% (p's < 0.01 and 0.05, respectively) of the variance in change in pain interference and depressive symptoms, respectively, at 2-years post-amputation. Catastrophizing and social support were associated with decreases (improvement) in both criterion measures, while solicitous responding was associated with increases (worsening) in both measures. DISCUSSION: The findings support a biopsychosocial model of long-term adjustment to amputation and PLP. In addition, results suggest that some psychosocial variables are more important than others for predicting adjustment, providing important implications for early interventions after amputation.     

Welcome back survey: exploring concerns impacting HIV care engagement and retention

The current study describes the development of a short pre-clinic survey that helped multidiscipline providers to elicit patient perspective on barriers to HIV primary care. The survey was piloted with 318 patients returning to care after being lost to care for at least 12 months. Reasons for breaks in care were dependent on age, gender, and race. Concerns about confidentiality in care were more commonly reported by African-American, Latino, and younger patients, while concerns relating to acceptance of diagnosis and side effects were greater for women, African-American, and Latino participants. Further, Intimate Partner Violence (IPV) and transportation were greater concerns for women and younger patients in the sample.     

In vivo analysis of intestinal permeability following hemorrhagic shock

AIM: To determine the time course of intestinal permeability changes to proteolytically-derived bowel peptides in experimental hemorrhagic shock. METHODS: We injected fluorescently-conjugated casein protein into the small bowel of anesthetized Wistar rats prior to induction of experimental hemorrhagic shock. These molecules, which fluoresce when proteolytically cleaved, were used as markers for the ability of proteolytically cleaved intestinal products to access the central circulation. Blood was serially sampled to quantify the relative change in concentration of proteolytically-cleaved particles in the systemic circulation. To provide spatial resolution of their location, particles in the mesenteric microvasculature were imaged using in vivo intravital fluorescent microscopy. The experiments were then repeated using an alternate measurement technique, fluorescein isothiocyanate(FITC)-labeled dextrans 20, to semi-quantitatively verify the ability of bowel-derived low-molecular weight molecules( 20 k D) to access the central circulation.RESULTS: Results demonstrate a significant increase in systemic permeability to gut-derived peptides within 20 min after induction of hemorrhage(1.11 ± 0.19 vs 0.86 ± 0.07, P 0.05) compared to control animals. Reperfusion resulted in a second, sustained increase in systemic permeability to gut-derived peptides in hemorrhaged animals compared to controls(1.2 ± 0.18 vs 0.97 ± 0.1, P 0.05). Intravital microscopy of the mesentery also showed marked accumulation of fluorescent particles in the microcirculation of hemorrhaged animals compared to controls. These results were replicated using FITC dextrans 20 [10.85 ± 6.52 vs 3.38 ± 1.11 fluorescent intensity units(× 105, P 0.05, hemorrhagic shock vs controls)], confirming that small bowel ischemia in response to experimental hemorrhagic shock results in marked and early increases in gut membrane permeability. CONCLUSION: Increased small bowel permeability in hemorrhagic shock may allow for systemic absorption of otherwise retained proteolytically-generated peptides, with consequent hemodynamic instability and remote organ failure.     

Plasma homocysteine concentration and its relationship with the development of preeclampsia. Effect of prenatal administration of folic acid

The increase of plasmatic homocysteine (Hc) in pregnant women, who later develop preeclampsia/eclampsia, the cause of this increment and its pathogenic role in toxemia of pregnancy, are still controversial. The objectives of the present research were to determine the plasmatic He concentrations during the first and second trimesters of pregnancy and the effect of folic acid administration on these values, and in the prevention of preeclampsia. Ninety six pregnant women of low economic background were studied on the first prenatal consultation: 27 women in the first trimester of pregnancy and 59 in the second. After 8 hours of fasting, venous blood was extracted and each patient was provided with 1 mg folic acid tablets and instructed to ingest one tablet daily, and to come back to the laboratory after three months. Plasma homocysteine and serum folic acid were determined for each patient before and after the folic acid treatment, by using the IMX system (Abbott Lab) and radioimmunoassay, respectively. Basal homocysteine concentrations were 4.0 +/- 2.1 micromol/L and 4.8 +/- 2.1 micromol/L in the first and second trimesters respectively, with no significant modifications after three months of folic acid. Although the degree of desertion from the study was high, it was possible to determine the evolution of 65 pregnancies. Ten of them developed preeclampsia (15.4%). No significant differences were found in Hc concentrations, or the frequency of hyperhomocysteinemia in the different stages of pregnancy, between women with normal gestation and those who developed preeclampsia. The small sample size of these groups, preclude any valid conclusion, however the results do not suggest that Hc concentration or folic acid administration influence the development of toxemia of pregnancy.     

cGMP/PKG pathway mediates myocardial postconditioning protection in rat hearts by delaying normalization of intracellular acidosis during reperfusion

Ischemic postconditioning has been demonstrated to limit infarct size in patients, but its molecular mechanisms remain incompletely understood. Low intracellular pH (pHi) inhibits mitochondrial permeability transition, calpain activation and hypercontracture. Recently, delayed normalization of pHi during reperfusion has been shown to play an important role in postconditioning protection, but its relation with intracellular protective signaling cascades is unknown. The present study investigates the relation between the rate of pHi normalization and the cGMP/PKG pathway in postconditioned myocardium. In isolated Sprague-Dawley rat hearts submitted to transient ischemia both, postconditioning and acidic reperfusion protocols resulted in a similar delay in pHi recovery measured by ³¹P-NMR spectroscopy (3.6 ± 0.2 min and 3.5 ± 0.2 min respectively vs. 1.4 ± 0.2 min in control group, P < 0.01) and caused equivalent cardioprotection (48% and 41% of infarct reduction respectively, P < 0.01), but only postconditioning increased myocardial cGMP levels (P = 0.02) and activated PKG. Blockade of cGMP/PKG pathway by the addition of the guanylyl cyclase inhibitor ODQ or the PKG inhibitor KT5823 during reperfusion accelerated pHi recovery and abolished cardioprotection in postconditioned hearts, but had no effect in hearts subjected to acidic reperfusion suggesting that PKG signaling was upstream of delayed pHi normalization in postconditioned hearts. In isolated cardiomyocytes the cGMP analog 8-pCPT-cGMP delayed Na⁺/H⁺-exchange mediated pHi normalization after acidification induced by a NH₄Cl pulse. These results demonstrate that the cGMP/PKG pathway contributes to postconditioning protection at least in part by delaying normalization of pHi during reperfusion, probably via PKG-dependent inhibition of Na⁺/H⁺-exchanger.