Studies using a viral challenge and CD8 T cell depletions on the roles of cellular and humoral immunity in the control of an SHIV-89.6P challenge in DNA/MVA-vaccinated macaques

Here, we study immune responses in four DNA/MVA-vaccinated macaques following an SHIV-89.6P challenge and a subsequent CD8 cell depletion. Both post-challenge and post-depletion peaks of viremia contracted with the expansion, or re-emergence, of CD8 T cells. Post-depletion, CD8 cells expanded in the presence of higher levels of neutralizing Ab and CD4 help than post-challenge and had superior maturational characteristics as measured by expression of the anti-apoptotic protein Bcl-2, the IL-7 receptor CD127 and co-production of IFN-gamma and IL-2. Pre-challenge and pre-depletion titers of neutralizing Ab correlated inversely with peaks of viremia and directly with peaks for anti-viral CD4 cells. Thus, our results reveal CD8 cells playing a central role, and neutralizing Ab, a supporting role in SHIV-89.6P control. They also suggest a dynamic relationship between neutralizing Ab, antigen load and anti-viral CD4 cells in the maturation of high-quality anti-viral CD8 T cells.     

Retrospective-prospective study of safety and efficacy of sofosbuvir-based direct-acting antivirals in HIV/HCV-coinfected participants with decompensated liver disease pre– or post–liver transplant

Direct-acting antiviral (DAA) therapy has transformed the management of human immunodeficiency virus (HIV) and hepatitis C (HCV) coinfected patients with advanced liver disease. STOP-Coinfection was a multicenter prospective and retrospective, open-label study using sofosbuvir-based DAA therapy to treat HIV/HCV-coinfected participants pre– or post–liver transplant (LT). Sixty-eight participants with end-stage liver disease (Child-Turcotte-Pugh score ≥7 and Model for End-Stage Liver Disease score 6–29) were enrolled, 26 had hepatocellular carcinoma. Forty-two participants were treated pre–LT and 26 post–LT. All participants completed therapy without need for dose reduction or transfusion; eight required two or more courses of therapy. Ninety-three percent achieved a sustained virologic response and DAA therapy was well tolerated. Despite HCV cure, 12 end-stage liver disease participants required subsequent LT, 7 for decompensated liver disease. Thirteen participants died, 10 with decompensated liver disease pre–LT and three post–LT. Overall, transplant free survival was 42.8% at 4 years and post–LT survival was 87.9% at 5 years. We conclude that sofosbuvir-based DAA therapy is safe and highly effective in HCV-HIV patients with decompensated liver disease and post–LT, with post–LT survival rates comparable to other indications. This removes one of the last barriers to liver transplantation in this challenging cohort of recipients.     

Angiogenesis is an early event in the development of chemically induced skin tumors

In this study we have analyzed the vascular response induced in the two- stage carcinogenesis model in SENCAR mice. The role of angiogenesis has not been explored in this model, which is the paradigm of multistage carcinogenesis and a model for neoplastic lesions derived from exophytic premalignant lesions (e.g. colon carcinoma, bladder papilloma). We investigated if angiogenesis is involved in the formation of papillomas and in the progression from papilloma to carcinoma. To this end we analyzed the vasculature of normal and hyperplastic skin, focal epidermal hyperplasias that are precursors of papillomas, papillomas at different stages and squamous cell carcinomas. We also analyzed the vascularization of papillomas induced in two strains of mice that differ in their susceptibility to malignant progression. We show here that angiogenesis is turned on in the earliest stages of papilloma formation. In late stages, regardless of state of progression, the predominant response is an increase in the size of blood vessels. Thus, in the SENCAR mouse model, representative of exophytic tumors, the angiogenesis switch is a very early event, probably mechanistically related to the development of the primarily exophytic lesions. Therefore, the density of blood vessels cannot be used as a predictor of malignant progression in this model.      

Diffuse bipolar cells provide input to OFF parasol ganglion cells in the macaque retina

Parasol retinal ganglion cells are more sensitive to luminance contrast and respond more transiently at all levels of adaptation than midget ganglion cells. This may be due, in part, to differences between bipolar cells that provide their input, and the goal of these experiments was to study these differences. Midget bipolar cells are known to be presynaptic to midget ganglion cells. To identify the bipolar cells presynaptic to parasol cells, these ganglion cells were intracellularly injected with Neurobiotin, cone bipolar cells were immunolabeled, and the double-labeled material was analyzed. In the electron microscope, we found that DB3 diffuse bipolar cells labeled by using antiserum to calbindin D-28k were presynaptic to OFF parasol cells. In the confocal microscope, DB3 bipolars costratified with OFF parasol cell dendrites and made significantly more appositions with them than expected due to chance. Flat midget bipolar cells were labeled with antiserum to recoverin. Although they made a few appositions with parasol cells, the number was no greater than would be expected when two sets of processes have overlapping distributions in the inner plexiform layer. DB2 diffuse bipolar cells were labeled with antibodies to excitatory amino acid transporter 2, and they also made appositions with OFF parasol cells. These results suggest that DB2 bipolar cells are also presynaptic to OFF parasol ganglion cells, but midget bipolar cells are not. We estimate that midperipheral OFF parasol cells receive approximately 500 synapses from 50 DB3 bipolar cells that, in turn, receive input from 250 cones.     

Plackett–Burman randomization method for Bacterial Ghosts preparation form E. coli JM109

Plackett–Burman randomization method is a conventional tool for variables randomization aiming at optimization. Bacterial Ghosts (BGs) preparation has been recently established using methods other than the E lysis gene. The protocol has been based mainly on using critical concentrations from chemical compounds able to convert viable cells to BGs. The Minimum Inhibition Concentration (MIC) and the Minimum Growth Concentration (MGC) were the main guide for the BGs preparation. In this study, Escherichia coli JM109 DEC has been used to produce the BGs following the original protocol. The study contained a detail protocol for BGs preparation that could be used as a guide.     

Evaluation of alternative statistical models for crossover studies to demonstrate caffeine adjuvancy in the treatment of tension headache

This paper discusses alternative statistical models for the analysis of six crossover studies to determine whether better relief of tension headache occurs from treatment with an analgesic plus caffeine (C) than with the analgesic alone (A) or with placebo (P). Each patient in these crossover studies randomly received a pair of distinct medications in such a way as to treat the first two of four headaches with the initial medication in the pair and to treat the third and fourth headaches with the last medication in the pair. In order to have greater power for the C versus A comparison, three times as many patients were randomly assigned to the A:C and C:A sequence groups as to the A:P, C:P, P:A, and P:C sequence groups.

An issue of statistical interest for these crossover studies is the extent to which the possibility of unequal carryover effects of the three medications influences the roles of alternative models for data analysis and the interpretation of results. When carryover effects for all three medications are equal, univariate analysis of variance for the difference scores between the average response for the first two headaches and the average response for the third and fourth headaches for each patient provides nearly the same power for pairwise treatment comparisons as more comprehensive multivariate methods for all four headaches. However, for comparisons concerning carryover effects and for treatment comparisons with adjustment for carryover effects, multivariate methods encompassing all four headaches jointly can provide greater power than univariate analysis for difference scores, particularly when there is low intraclass correlation for responses within the same patient. Another noteworthy role for multivariate methods in situations with potentially unequal carryover effects is their capacity to clarify whether multiple types of carryover effects occur across the second, third, and fourth headaches in the respective sequence groups.

Multivariate models with alternative specifications of carryover effects are fit to the data from the six crossover studies to compare C, A, and P by weighted least squares. The role of potential variation among centers is addressed in these analyses by the use of stratified proportional means over centers, means of center means, and means ignoring centers. The primary focus of attention in the respective analyses is the evaluation of treatment comparisons with and without adjustment for potential differences among carryover effects of the treatments. Comparisons among carryover effects are assessed as well, but they mainly serve a background purpose since the principal issue is the extent to which findings for treatment comparisons are similar across alternative ways of accounting for potential carryover effects.

For all models, the average predicted response across all headaches treated with C was significantly better than that for A or P. For models that adjusted treatment effects for carryover effects in a statistically efficient way, the adjusted direct treatment effect of C was significantly better than that of A or P. Thus, the superiority of C over A found robust support from models both with and without adjustment for potential differences among carryover effects of the treatments.