Identification and HLA-Tetramer-Validation of Human CD4(+) and CD8(+) T Cell Responses against HCMV Proteins IE1 and IE2

Human cytomegalovirus (HCMV) is an important human pathogen. It is a leading cause of congenital infection and a leading infectious threat to recipients of solid organ transplants as well as of allogeneic hematopoietic cell transplants. Moreover, it has recently been suggested that HCMV may promote tumor development. Both CD4+ and CD8+ T cell responses are important for long-term control of the virus, and adoptive transfer of HCMV-specific T cells has led to protection from reactivation and HCMV disease. Identification of HCMV-specific T cell epitopes has primarily focused on CD8+ T cell responses against the pp65 phosphoprotein. In this study, we have focused on CD4+ and CD8+ T cell responses against the immediate early 1 and 2 proteins (IE1 and IE2). Using overlapping peptides spanning the entire IE1 and IE2 sequences, peripheral blood mononuclear cells from 16 healthy, HLA-typed, donors were screened by ex vivo IFN-γ ELISpot and in vitro intracellular cytokine secretion assays. The specificities of CD4+ and CD8+ T cell responses were identified and validated by HLA class II and I tetramers, respectively. Eighty-one CD4+ and 44 CD8+ T cell responses were identified representing at least seven different CD4 epitopes and 14 CD8 epitopes restricted by seven and 11 different HLA class II and I molecules, respectively, in total covering 91 and 98% of the Caucasian population, respectively. Presented in the context of several different HLA class II molecules, two epitope areas in IE1 and IE2 were recognized in about half of the analyzed donors. These data may be used to design a versatile anti-HCMV vaccine and/or immunotherapy strategy

The association between renal function and structural parameters: a pig study

<p>Abstract</p> <p>Background</p> <p>The objective was to investigate the association between renal structural parameters and renal function. The structural parameters were renal cortical volume, total renal volume, number of glomeruli, and total glomerular volume, and renal function was expressed by the single kidney GFR (skGFR). Investigations were performed using both healthy and chronically diseased kidneys. We investigated which of the structural parameters showed the best correlation to renal function and evaluated the possibility of predicting the renal function from structural parameters.</p> <p>Methods</p> <p>Twenty-four pigs, twelve with healthy kidneys and twelve with diseased kidneys, underwent skGFR measurements. Nephrectomies were performed and structural parameters were estimated using stereological procedures. The correlation between the structural parameters and skGFR was analysed by Pearson's correlation test. The prediction of skGFR from structural parameters was analysed by a linear regression test.</p> <p>Results</p> <p>In general, we demonstrated a good correlation between structural parameters and skGFR. When all kidneys were evaluated together Pearson's correlation coefficient between skGFR and any stereological parameter was above 0.60 and highly significant (p < 0.001), and with r-values ranging from 0.62 regarding number of glomeruli, to 0.78 regarding cortical volume. The best correlation was found between cortical volume and skGFR. Prediction of single kidney GFR from any structural parameter showed to be quite imprecise.</p> <p>Conclusion</p> <p>The observed correlations between structural parameters and renal function suggest that these parameters may potentially be useful as surrogate markers of the renal function. At present, however, precise prediction of renal function based on a single structural parameter seems hard to obtain.</p

The Acute Effect of Insulin on Capillary Endothelial Cells

A quantitative morphologic study of the capillary endothelium of striated muscle was performed in diabetic rats one hour after injection of insulin. Diabetic and nondiabetic rats injected with saline constituted the reference groups. In a randomized sample of capillary profiles, morphometric data on micropinocytotic vesicles (MPV) and on various capillary characteristics were collected. The numeric density of MPV was higher in diabetic rats injected with insulin than in diabetic rats injected with saline (44.2 ± 8.5 versus 34.0 ± 4.5 MPV/μ2 endothelial cytoplasm (mean ± S.D.), 2p = 0.045), thereby approaching the control value of 48.0 ± 7.3, 2p = 0.51. The insulin-injected animals, furthermore, differed markedly from the diabetic reference group as regards the ratio of free-to-attached vesicles, since they showed a large increase in the numeric density of free MPV (1.40 ± 0.18 versus 1.00 ± 0.23/μ circumference, 2p = 0.015, control value 1.59 ± 0.20) and a small decrease in the density of attached MPV (0.19 ± 0.03 versus 0.23 ± 0.02/μ circumference, 2p = 0.022; controls: 0.15 ± 0.04). Both the height of the endothelial cytoplasm and the size of the MPV were similar in all groups. These findings may be the morphologic counterpart of the clinically demonstrated decrease in plasma volume and in the intravascular pool of albumin in diabetic patients after insulin injection. The present results demonstrate that insulin has an acute effect on the endothelial cells, including a change in the kinetics of the vesicular transport pathway</jats:p

Irreversibility of Glomerular Basement Membrane Accumulation Despite Reversibility of Renal Hypertrophy with Islet Transplantation in Early Experimental Diabetes

A quantitative morphologic study of the glomeruli in rats after 4 wk of streptozotocin-induced diabetes showed a number of glomerular changes, as previously described. Of particular interest was the increase in the total amount of glomerular basement membrane material [from 0.94 ± 0.13 (SD) mm3 to 1.26 ± 0.14 mm3 per kidney]. This parameter did not change after 4 wk of normoglycemia following islet cell transplantation (1.19 ± 0.17 mm3), nor was the total glomerular volume normalized. The contralateral kidney was weighed and used for estimating the total amounts of protein, RNA, and DNA. Four weeks of diabetes expectedly resulted in a 50% increase in kidney weight, and islet cell transplantation diminished this to 15% in excess of normal. The average cell size (protein/DNA ratio) paralleled the kidney size after diabetes and following transplantation. The average amount of RNA per cell (RNA/DNA) increased significantly after induction of diabetes and was totally normalized after transplantation. Kidney protein concentration (mg protein/mg kidney) remained constant throughout the experiment. Considering that a few weeks of diabetes provokes a large increase in basement membrane material, it is especially noteworthy that 1 mo of normoglycemia is quite insufficient to reverse the accumulation.</jats:p