There is evidence that a humoral factor or factors in rats with one-kidney, one-clip (1K1C) hypertension increase growth of cultured vascular smooth muscle cells. Such humoral trophic factors may contribute to the abnormal growth of arterial muscle in hypertension. To further study the longitudinal expression of this trophic factor or factors, we prepared rats with 1K1C hypertension of different durations. To determine if the factor or factors are also expressed in other forms of experimental hypertension, we additionally prepared rats with two-kidney, one-clip (2K1C) hypertension and paired two-kidney (2K) normotensive controls; we also studied Spontaneously Hypertensive Rats (SHR) plus appropriate controls. In the presence of growth stimulated by background levels (1%) of fetal calf serum, 20% platelet-poor, plasma-derived serum (PDS) from 1K1C rats 8-14 days (n = 10) and 28 days (n = 12) after clipping increased [3H]-thymidine incorporation of growth-arrested cultured rat aortic smooth muscle cells more than the paired 1K PDS, by up to +67% and +40%, respectively (P < 0.01). However, with PDS from 1K1C rats 4 days (n = 11) and 38 days (n = 6) after clipping there was no evidence for a differential effect (P > 0.5 and P > 0.1, respectively). PDS from seven 2K1C rats (at 9 days) also increased [3H]-thymidine incorporation of the assay cells more than PDS from the paired 2K rats, by up to +19% (P < 0.05). However, there was no evidence that PDS from SHR differentially increased cellular thymidine incorporation. Thus, evidence from this study suggests that the humoral factor or factors trophic for vascular smooth muscle are expressed in both low- and high-renin forms of experimental renovascular hypertension, but not in the very early or in the late complicated stages of the hypertension, or in genetic hypertension in rats.
Macrophages in the liver are well known for their functional heterogeneity. However, subpopulations of the hepatic macrophages are not well defined.Two subsets of hepatic macrophages isolated from rats via FACS with immunolabeling of ED2 (anti-CD163) antibody were studied for phenotypic and functional characteristics.A subset showed an ED2(high) and autofluorescence(high) (ED2(high)/AF(high)) phenotype, exhibiting characteristics consistent with the description of the Kupffer cells (KC). A second subset, displaying an ED2(dim)/AF(dim) phenotype, was smaller in size, monocyte-like and weak in phagocytosis. Transmission electron microscopy demonstrated that both subsets are phagocytes. Quantitative RT-PCR revealed that in addition to expression of macrophage-related surface markers such as CD14, ED1 (CD68), fucose receptor, and CD163, the ED2(dim)/ AF(dim) cells expressed mRNA encoding for myeloid lineage differentiation markers ERMP12 (PECAM) and ERMP20 (Ly-6C). These two subsets exhibited differential in gene expression of selected cytokines, extracellular matrix proteinases, and Toll-like receptor in normal livers, as well as significantly upregulated expression in cholestatic livers induced by bile duct ligation.The data suggest that the ED2(high)/AF(high) population of the liver cells represent the conventional Kupffer cells. The ED2(dim)/AF(dim) cells, however, are small hepatic resident macrophages characteristically different from the conventional Kupffer cells.
Background. Natural antibodies that react with galactose-α(1,3)galactose [galα(1,3)gal] carbohydrate epitopes exist in humans and Old World primates because of the inactivation of the α1,3-galactosyltransferase (α1,3GT) gene in these species and the subsequent production of antibodies to environmental microbes that express the galα(1,3)gal antigen. The Gal knockout (Gal o/o) mouse, produced by homologous disruption of the α1,3GT gene, spontaneously makes anti-galα(1,3)gal antibodies and can be used to study the genetic control of humoral immune responses to this carbohydrate epitope. Methods. Six hybridomas that produce monoclonal antibodies (mAbs) to galα(1,3)gal were generated in Gal o/o mice. The mAbs were tested to characterize the binding activity with flow cytometry using pig aortic endothelial cells and ELISA with galα(1,3)gal carbohydrates. The VH and VK genes of these hybridomas were cloned, sequenced, and analyzed. Results. The mAbs showed distinct patterns of antibody binding to galα(1,3)gal antigens. The VH genes that encode the mAb binding activity were restricted to a small number of genes expressed in their germline configuration. Four of six clones used closely related progeny of the same VH germline gene (VH441). Comparison of the mouse gene VH441 to the human gene IGHV3-11, a gene that encodes antibody activity to galα(1,3)gal in humans, demonstrates that these two genes share a nonrandom distribution of amino acids used at canonical binding sites within the variable regions (complimentary determining regions 1 and 2) of their immunoglobulin VH genes. Conclusions. These results demonstrate the similarity of the Gal o/o mice and humans in their immune response to galα(1,3)gal epitopes. Gal o/o mouse can serve as a useful model for examining the genetic control of antibody/antigen interactions associated with the humoral response to pig xenografts in humans.
