Safety and efficacy analysis of liposomal insulin-like growth factor-1 in a fluid gel formulation for hair-loss treatment in a hamster model.

Insulin-like growth factor (IGF)-1 has shown some interesting results in studies examining its use as a hair-loss treatment. IGF-1 works by regulating cellular proliferation and migration during the development of hair follicles. Hepatotoxicity and myelotoxicity were evaluated in hamsters (Mesocricetus auratus) after topical application of the liquid gel vehicle (placebo), 1% IGF-1 or 3% IGF-1. No significant difference in the levels of aspartate aminotransferase or alanine aminotransferase was found between the control and treated groups. ELISA did not shown any increase in the plasma level of IGF-1. A haematopoietic niche was found, but it was not associated with myelotoxicity. Efficacy was determined by dermatoscopy analysis of hair density and microscopy analysis of hair diameter, with hair found to be thicker and with more rapid growth in the 3% group than in either the 1% group or the control group. These results strongly suggest that liposomal IGF-1 in a liquid gel formulation is a safe and efficient treatment for hair loss.

Cytotoxic T cell-resistant variants are selected in a virus-induced demyelinating disease.

C57BI/6 mice infected with mouse hepatitis virus, strain JHM (MHV-JHM) develop a chronic demyelinating encephalomyelitis. Infectious virus can be isolated only from symptomatic mice. In C57BI/6 mice, two CD8+ T cell epitopes within the MHV-JHM surface glycoprotein were previously identified. Here, we show that mutations in the RNA encoding the immunodominant of the epitopes are present in nearly all virus samples isolated from these mice. Mutations are not present in sequences flanking this epitope or in other CD8+ or CD4+ T cell epitopes. Furthermore, analysis of five peptides corresponding to variant epitopes in direct ex vivo cytotoxicity assays showed that each mutation caused a loss of epitope recognition. These results suggest that escape from CD8+ T cell recognition is necessary for enhanced virus replication and development of clinical disease in these MHV-JHM-infected mice.

Differential antigen recognition by T cells from the spleen and central nervous system of coronavirus-infected mice.

CD8+ cytotoxic T lymphocytes (CTLs) isolated from the central nervous system (CNS) of C57BI/6 mice acutely infected with mouse hepatitis virus, strain JHM (MHV-JHM), and analyzed in a direct ex vivo cytotoxicity assay recognize two epitopes (H-2Db- and H-2Kb-restricted encompassing amino acids 510-518 and 598-605, respectively) within the surface (S) glycoprotein. In contrast, CD8+ T cells isolated from the spleens of mice inoculated intraperitoneally with MHV-JHM and restimulated in vitro only respond to the H-2Db-restricted epitope. In this report, the preferential recognition of the H-2Db-restricted epitope is confirmed using splenocytes stimulated in vitro with either MHV-JHM-infected MC57 cells or with a cell line expressing the S protein and analyzed in secondary CTL assays. To determine whether these results represent a difference in epitope recognition between the spleen and CNS, secondary CTL assays were performed using spleen cells coated with peptides encompassing the CTL epitopes as stimulators. Under these conditions, both epitopes sensitized cells for lysis by spleen-derived CTLs, suggesting that both epitopes were recognized by splenic CD8+ T cells after infection in vivo. Furthermore, limiting dilution analysis indicated that the precursor frequency of splenic CD8+ T cells specific for both the H-2Kb- and H-2Db-restricted epitopes were not significantly different. Thus, the results suggest that in vitro stimulation of splenocytes specific for the H-2Kb-restricted epitope is inefficient after endogenous processing but that this inefficiency can be corrected if peptide is provided exogenously at sufficiently high concentrations. As a consequence, the results also show that cells responsive to both of the previously identified CNS-derived CD8+ T cell epitopes are present in the infected spleen at nearly the same frequency.

CD8+ T-cell epitopes within the surface glycoprotein of a neurotropic coronavirus and correlation with pathogenicity.

CD8+ T cells with cytotoxic activity against the surface glycoprotein (S) of mouse hepatitis virus, strain JHM, have been identified in the central nervous system (CNS) of both acutely and chronically infected C57BL/6 mice. In this report, two specific epitopes recognized by these CNS-derived cells were identified, using a panel of peptides chosen because they conformed to the allele-specific binding motif for MHC class I H-2Kb and H-2Db. The active peptides encompassed residues 510 to 518 (CSLWNGPHL, H-2Db) and 598 to 605 (RCQIFANI, H-2Kb). Both epitopes are located within the region of the S protein previously shown to be prone to deletion after passage in animals. These deleted strains are generally less neurovirulent than the wild-type virus but still are able to cause demyelination. Since C57BL/6 mice become persistently infected more commonly than many other strains of mice, these data are consistent with a role for CD8+ T-cell escape mutants in the pathogenesis of the demyelinating disease. This is the first report of CD8+ T-cell epitope localization within the S protein, the protein most strongly implicated thus far in pathogenesis in the host.

Activation of astrocytes in the spinal cord of mice chronically infected with a neurotropic coronavirus.

Mice infected with the neurotropic JHM strain of mouse hepatitis virus (MHV-JHM) develop a demyelinating encephalomyelitis several weeks after infection. Astrogliosis and infiltration of inflammatory cells are prominent findings in the brains and spinal cords of infected mice. In this report, astrocytes in infected spinal cords were analyzed for expression of three pleiotropic cytokines, TNF-alpha, IL-1 beta, and IL-6; Type 2 nitric oxide synthase (iNOS); and MHC class I and II antigen. The data show that all three cytokines and iNOS are expressed by astrocytes in chronically infected spinal cords. These activated astrocytes are localized to areas of virus infection and demyelination, although most of the astrocytes expressing these proteins are not MHV-infected. MHC class I and II antigen can be detected in these spinal cords as well, but not in cells with the typical morphology of astrocytes. TNF-alpha, IL-6, and iNOS are also evident in the brains of mice with MHV-induced acute encephalitis, but in marked contrast to the results obtained with the chronically infected mice, most of the cells expressing these cytokines or iNOS had the morphology of macrophages or other mononuclear cells and very few appeared to be astrocytes. Additionally, astrocytes and, most likely, oligodendrocytes are infected in the spinal cords of mice with chronic demyelination. These results are consistent with a role for both viral infection of glial cells and high localized levels of proinflammatory cytokines and nitric oxide in the demyelinating process in mice infected with MHV-JHM. They also show that analogously to the human demyelinating disease, multiple sclerosis, astrocytes are a major cellular source for these cytokines in mice with chronic, but not acute disease.

Coronavirus-induced demyelination occurs in the presence of virus-specific cytotoxic T cells.

C57BI/6, but not BALB/c, mice infected with mouse hepatitis virus strain JHM (MHV-JHM) develop a late onset, symptomatic demyelinating encephalomyelitis. In this report, we characterized anti-viral cytotoxic T cells in the central nervous system and spleen during the acute and chronic stages of the MHV infection. The data show that C57BI/6 mice display a cytotoxic T cell (CTL) response to the surface (S) glycoprotein and this response can be demonstrated in lymphocytes isolated from the brains and spinal cords of mice both acutely and persistently infected with MHV-JHM. Thus, the anti-S CTL activity present in the central nervous system of chronically infected animals is not sufficient to prevent the demyelinating process. BALB/c mice have been shown previously to mount a CTL response against the nucleocapsid (N) protein (Stohlman et al., 1992). Since C57BI/6 mice do not mount a response to the N protein, the role of the N-specific response in preventing the late onset disease was assessed using B10.A(18R) mice, recombinant in the H-2 locus. These mice contain the d alleles of the D and L loci and exhibit a CTL response against the N protein. However, unlike the BALB/c mice, these animals develop the late onset symptomatic disease. These results suggest that the N-specific response is partially protective against the development of the demyelinating disease, but that additional factors are also likely to be involved.

Estudo comparativo duplo-cego, randomizado, cruzado, controlado com placebo, da eficácia de nifedipina retard em pacientes refratários a terapêutica basal anti-hipertensiva / Comparative double-blind, randomized crossover study controlled with placebo, of the efficacy of nifedipine retard in patients refractory to basic antihypertensive therapy

Foi efetuado um estudo cruzado comparativo duplo-cego, randomizado, controlado com placebo, para avaliar a eficacia da nifedipina retard quando adicionada a terapeutica basal anti-hipertensiva que nao tenha conseguido baixar os niveis da pressao sanguinea diastolica ate valores normais.Trinta e seis pacientes de ambulatorio com hipertensao leve e moderada receberam nifedipina ou placebo durante 4 semanas apos um periodo de "washout" de 4 semanas. Este procedimento foi seguido por uma semana de placebo e 4 semanas de nifedipina ou placebo. Os pacientes foram avaliados semanalmente. Nifedipina retard foi administrada na dose de 20 mg, 2 vezes ao dia. O placebo foi administrado em regime identico. Em ambos os grupos, a nifedipina provocou uma reducao estatisticamente significativa (p < 0.01) nas pressoes sistolica e diastolica. Nao foram registradas alteracoes significantes na frequencia cardiaca com nifedipina nem alteracoes significantes da pressao sanguinea ou frequencia cardiaca quando se associou placebo a terapeutica basal. A tolerancia foi boa, porem houve casos de discreta cefaleia,edema maleolar e rubor, em alguns pacientes