Prolongation of survival of primary renal allografts by feeding of donor spleen cells.

We have found that feeding Brown Norway (BN) rat spleen cells to Lewis rats prior to transplanting BN kidneys prolongs allograft survival (mean: 8.8 days in unfed rats, 21 days in the BN cell-fed rats; longest survival: 11 days without allo-feeding vs. 37 days with feeding). We have also found that feeding BN cells both before and after transplantation further extends survival (mean: 38 days; longest survival: 105 days). We also examined the cells infiltrating the grafts during the early stages of the allograft response (day 5). Using flow cytometry, we found a significant decrease in the number of leukocytes infiltrating the transplanted kidneys of fed animals. This decrease was mainly due to a drop in the number of infiltrating T cells. We also found that cytokine mRNA production by the graft-infiltrating lymphocytes, assessed by reverse transcription polymerase chain reaction, showed a significant increase in interleukin-4 and transforming-growth factor-beta mRNA in the graft-infiltrating lymphocytes of fed animals compared with the controls.

Class II MHC expression on rat intestinal intraepithelial lymphocytes.

The expression of class II major histocompatibility complex (MHC) on rat intestinal intraepithelial lymphocytes (IELs) and mesenteric lymph node cells (MLNCs) were examined by immunofluorescence and flow cytometry. As expected, MLNCs contain eight small populations of CD4+ class II MHC+ T-cells in addition to classical antigen presenting cells. In contrast, rat IELs include a significant population of class II MHC+ T-cells, predominantly in the CD8+ CD4-alpha beta TCR+ subset. IEL samples with a relatively high percentage of class II MHC+ cells also include some CD4+ class II MHC+ cells; IEL samples with a low percentage of class II MHC+ cells also include some CD4- CD8- class II MHC+ cells. The role of lymphocyte subpopulations in the intestinal epithelium may need to be revisited in consideration of these findings.

Pattern of proinflammatory cytokine mRNA expression during Trichinella spiralis infection of the rat.

Trichinella spiralis occupies an intramulticellular niche in the small intestinal epithelium, and thus we examined the intestine and gut-associated tissues for proinflammatory cytokines during the infection. We document the patterns of interleukin-1 (IL-1), IL-6, gamma interferon, and tumor necrosis factor alpha mRNA expression in the duodenum, jejunum, Peyer's patches, mesenteric lymph node, spleen, and liver in T. spiralis-infected rats. By reverse transcription-PCR detection of mRNAs, IL-1beta was found increased in the jejunum but only on day 2. The jejunal IL-1beta increase was attributed to the epithelium by isolating epithelial cells and then depleting them of intraepithelial lymphocytes prior to analysis. The only cytokine for which mRNA was substantially increased in tissues later in infection was tumor necrosis factor alpha in the spleen and, to a lesser extent, in the mesenteric lymph node. In fact mRNA levels for some cytokines declined below uninfected levels in some organs during the infection. IL-1 may be important in the initiation of the intestinal inflammatory response to this infection.

Isolation and characterization of highly purified rat intestinal intraepithelial lymphocytes.

The study of intestinal intraepithelial lymphocytes (IEL) has been hindered by the difficulty of isolating a population of lymphocytes which is free of epithelial cell or lamina propria cell contaminants and representative of the in vivo population of IEL in both phenotype and function. We describe an improved technique for the extraction and purification of IEL from the proximal small intestine of the rat. This technique rapidly and reproducibly isolates 5-10 x 10(6) IEL/rat with 90-95% purity and viability without the use of enzymes which affect lymphocyte function. The resulting cell population, which is 75% alpha beta T cell receptor (TCR)+, 70% CD8+, and 33% CD4+ T cells, and only 5% B cells and 2% macrophages, is of suitable purity to allow for flow cytometric analysis of the entire population of cells without requiring gating on lymphocytes. IEL are comprised of a unique T cell repertoire in that 27% of cells co-express the CD4 and CD8 molecules, but only 11% of CD4+ cells co-express CD45RC. All CD4+ cells express the alpha beta TCR, but 9% of IEL are CD8+ CD4- alpha beta TCR-. The adhesion molecules alpha 4 integrin and L-selectin are expressed on 57% and less than 1% of IEL, respectively. The isolated IEL population contains mRNA for IL-1 alpha, IL-1 beta, IL-1R, IL-1RA, IL-2, IL-6R, IFN-gamma, TGF-alpha, TGF-beta 1, and TNF-alpha. Mesenteric lymph node cells (MLNC) were examined in parallel. This technique allows for the isolation of rat IEL appropriate for phenotypic analysis by flow cytometry and for cytokine analysis by reverse transcription/polymerase chain reaction.