Peripheral blood CD4+CD8+ lymphocytes in cynomolgus monkeys are of resting memory T lineage

In this study, we analyzed peripheral blood CD4+CD8+ double-positive (DP) lymphocytes in adult cynomolgus monkeys (Macaca fascicularis). Forty of 55 monkeys had > 5% of the peripheral blood DP subpopulation (9.3 +/- 5.9%; mean +/- SD) in peripheral blood lymphocytes (PBL) in contrast to a low percentage of peripheral blood DP cells in humans and mice. In a cross-sectional study, the peripheral blood DP cells were found to increase in proportion with age. To clarify whether peripheral blood DP lymphocytes were immature precursors released from thymus without prior differentiation, the expressions of CD8 chains and CD1b on peripheral blood DP lymphocytes were compared with those on thymocytes. The peripheral blood DP lymphocytes were CD8 alpha + beta- and CD1b-, while thymic DP lymphocytes were CD8 alpha + beta + and CD1b +, suggesting that the peripheral blood DP cells are extrathymic T lymphocytes. Furthermore, the peripheral blood DP lymphocytes exhibited a resting memory T cell phenotype with CD2hiCD3+CD28-CD29hiCD49dhiCD69- CD80lo. Taken together, adult cynomolgus monkeys possess a unique peripheral blood DP T cell subpopulation which expresses a resting memory T cell phenotype. In addition, similar phenotypic properties of DP lymphocytes were distributed in the spleen and lymph nodes, although the proportion was less in the spleen and much less in lymph nodes than in PBL.      

Monoclonal and xenoantibodies specific for HLA-DR inhibit primary responses to HLA-D but fail to inhibit secondary proliferative (PLT) responses to allogeneic cells

Monoclonal antibodies (DA-2 and CA-2) and xenoantisera (rabbit anti-human, p23,30) specific for HLA-DR framework determinants were added to primary and secondary mixed lymphocyte cultures. Although such antisera were shown to inhibit primary MLC, primed lymphocytes were much less sensitive to the blocking effects of these antibodies. In the studies shown here, the concentration of antibody required to inhibit primary MLC reactions was 0.1-1.5 micrograms/ml).     

The dissociation of interleukin-2 production and antigen-specific helper activity by clonal analysis.

Influenza virus immune human T-lymphocyte clones maintained in continuous culture in TCGF were analysed for helper activity and interleukin-2 (IL-2) production. The clones that functioned as helper cells in the production of specific antibody failed to release detectable amounts of IL-2. Conversely, the T cells that produced IL-2 were unable to provide either specific or non-specific helper function. These findings indicated the IL-2 is not an essential component for helper activity. However, phenotypic analysis revealed that both the functional subsets of T-cell clones expressed the helper phenotype in that they were T4+, T3+ and T11+. Nevertheless analysis with other antibodies revealed differences in that the IL-2 releasing clone showed greater staining with the anti-T-cell subset antibodies 9.3 and Leu 8, confirming that there is phenotype as well as functional heterogeneity within the helper inducer T-cell population.     

Isolation of a temperature-sensitive dengue-2 virus under conditions suitable for vaccine development.

Dengue virus, type 2, in viremic human sera and after passage in cell cultures produces mixtures of small and large plaques when assayed in LLC-MK2 cells. Clones of dengue virus type 2 obtained by plaque selection in primary green monkey kidney cell cultures were tested for temperature sensitivity in vitro and for virulence by intracerebral inoculation of suckling mice. Sublines of a small-plaque clone were found to have lower nonpermissive temperatures than the parent virus by both plaque formation and release of infectious virus into the culture media. Small-plaque sublines were significantly less virulent in suckling mice than was the parent virus. Sublines from a large-plaque clone were not temperature sensitive and closely resembled parent virus mixed-plaque morphology. When small-plaque sublines were serially passaged using undiluted inocula, reversion occurred as evidenced by the appearance of large plaques and return of mouse virulence. Small-plaque virus could be maintained through several serial passages without reversion by using low-input inocula. Desirable passage history as well as temperature-sensitive and attentuation characteristics of the S-1 small-plaque subline make it appear suitable as a vaccine candidate virus.     

Temperature-sensitive events during the replication of the attenuated S-1 clone of dengue type 2 virus.

Temperature-sensitive events occurring during the replication of the attenuated S-1 clone of dengue type 2 virus were examined. The S-1 clone was more thermolabile than the parent virus at the nonpermissive temperature of 38.5 degrees C. Adsorption experiments in fetal rhesus monkey lung cells revealed an inefficient adsorption of S-1 at 38.5 degrees C compared with the parent virus, suggesting an alteration in a thermolabile virion protein important in adsorption. The production of S-1 viral RNA and antigen occurred at the nonpermissive temperature, which indicated that early events in the replication cycle of S-1 were not affected. Release of infectious virus at 38.5 degrees C was not impaired; however, lower amounts of infectious virus in infected cells at the nonpermissive temperature indicated that maturation of the S-1 clone was suppressed.     

Identification of antigenic escape variants in an immunodominant epitope of hepatitis C virus.

Numerous investigators have postulated that one mechanism by which hepatitis C virus (HCV) may evade the immune system is through the formation of escape mutants. This hypothesis is based largely on the observed mutability of the viral genome resulting in evolution of diverse quasispecies over the course of infection. That such diversification is a product of viral RNA polymerase infidelity, immune-driven selection or a combination of the two processes has not been addressed. We have examined sequence variability in a specific segment of HCV RNA encoding a known immunodominant region of the viral helicase, amino acids 358-375 of the non-structural 3 protein. Using sequence-specific oligonucleotide probe hybridization and automated DNA sequencing, we report a high frequency of mutations, essentially all of which result in amino acid replacements. To assess the biological impact of such mutations, corresponding chemically synthesized peptides were compared to wild-type peptide in T cell proliferation assays. We observed that a sizeable fraction of such peptides stimulated attenuated or negligible levels of proliferation by peripheral T cells from a chronically infected patient. This observation is consistent with expectations for immune-mediated selection of escape variants at the epitope level. We postulate that such a mechanism may be important in the immunopathogenesis of HCV infections.     

Incidence of diabetic retinopathy in a Hong Kong Chinese population

Background: The aim was to examine the progression and regression of diabetic retinopathy within a four‐year period in a Chinese population with type 2 diabetes mellitus in a community optometry clinic in Hong Kong. Methods: During the period May 2005 to November 2009, 5,160 patients with type 2 diabetes mellitus who had attended at least two diabetic retinopathy screening sessions at a community optometry clinic were included as subjects in this study. All had retinal photographs taken of both eyes, which were of sufficiently good quality for grading. For the purpose of this study, diabetic retinopathy grading was based on the results of the worst eye. The main outcomes were the within four‐year incidence of diabetic retinopathy and the incidence of progression and regression of diabetic retinopathy. Results: Of the 5,160 subjects in this study, 3,647 had no diabetic retinopathy, while 1,513 had diabetic retinopathy at the baseline visit. Of those 3,647 subjects with no diabetic retinopathy, the within four‐year cumulative incidence of any diabetic retinopathy, mild or moderate non‐proliferative diabetic retinopathy and sight‐threatening diabetic retinopathy was 15.16 per cent, 14.45 per cent, 0.69 per cent and 0.03 per cent, respectively. Of those 1,513 subjects with diabetic retinopathy at baseline, the within four‐year progression incidence of diabetic retinopathy was 6.61 per cent and the regression incidence of diabetic retinopathy was 45.54 per cent. Conclusion: The high regression incidence of diabetic retinopathy suggests that it might not be necessary for all patients with diabetes to be screened annually. Other methods to determine the screening frequency for an individual patient should be explored.