During cardiac rejection we studied the kinetics of IL-2 and IL-4 mRNA and subsequent protein production by
in vivo primed graft-infiltrating lymphocytes (GIL), using semiquantitative RT-PCR and ...ELISA. Following
in vitro stimulation with either donor or third-party antigens, mRNA expression of IL-2 and IL-4 were already detectable 1–2 h after stimulation, while their protein production could be measured from 4 h onwards at least until 48 h. At both the mRNA and protein level, we measured a donor-specific signal for IL-2 and for IL-4 production (
p = 0.02), while the relative donor-specific IL-2 mRNA level was significantly higher than the relative IL-4 mRNA level (
p = 0.002). These observations suggest that after
in vitro challenge with donor antigens, GIL obtained from rejecting cardiac allografts predominantly produce IL-2 mRNA and protein.
Sixteen monoclonal antibodies (Mcabs) were prepared against infectious bronchitis virus strain M41, all of them reacting with the peplomer protein. One of them, Mcab 13, was able to neutralize the ...virus and to inhibit hemagglutination. Competition binding assays allowed the definition of five epitopes, designated as A, B, C, D, and E, of which epitopes A and B are overlapping. Furthermore, the binding of Mcab 13 (epitope E) could be enhanced by the addition of Mcabs from group B, C, and D. A dot immunoblot assay was used to analyze the effect of denaturation on antibody recognition of the epitopes. Only the binding of Mcab 13 was affected, indicating that the epitope involved in neutralization and hemagglutination is conformation dependent. The epitopes A to D were highly conserved among IBV strains, while epitope E was specific for strains M41 and D3896. In this last strain, however, this epitope was not involved in neutralization.
This study was to determine whether the growth factors platelet-derived growth factor-alpha (PDGF-α) and transforming growth factor-betal (TGF-β1) contribute to the development of graft vascular ...disease (GVD) after clinical heart transplantation. We analysed intragraft PDGF-α and TGF-β1 messenger RNA (mRNA) expression levels by competitive template reverse transcriptase polymerase chain reaction (RT-PCR). Endomyocardial biopsies (EMB) were obtained at 1 and 9 months post-transplant from cardiac allograft recipients with (
n = 11) and without (
n = 11) angiographic evidence of GVD at 1 year. In 1-month EMB, comparable TGF-β1 mRNA levels were found in patients with and without GVD at 1 year (
p = 0.84, Mann-Whitney U-test). In contrast, in 9-month EMB during the development of GVD, intragraft mRNA levels of both PDGF-α (
p = 0.08) and TGF-β1 (
p = 0.03) were higher in patients with GVD after the first year compared to patients without GVD. These results suggest that intragraft PDGF-α and TGF-β1 play a role in the pathogenesis of accelerated GVD after clinical heart transplantation.
: Because production of immune regulatory proteins may play a role in early graft dysfunction after heart transplantation, we analyzed whether intragraft cytokine messenger RNA (mRNA) expression ...levels are associated with diastolic left ventricular function in cardiac allografts. We intensively monitored 16 cardiac allograft recipients during the first 3 months after transplantation. The mRNA expression levels of tumor necrosis factor (TNF‐α), monocyte chemoattractant protein‐1 (MCP‐1), transforming growth factor (TGF‐β), platelet derived growth factor (PDGF‐A), and basic fibroblast growth factor (bFGF) were measured in endomyocardial biopsies (n = 123) by quantitative RT‐PCR. To determine diastolic allograft function, concurrent M‐mode and two‐dimensional Doppler echocardiograms were analyzed for the following parameters: left ventricular total wall thickness, maximal early and atrial mitral flow velocity, deceleration time of maximal early mitral flow velocity, and isovolumetric relaxation period. During the first 3 months post‐transplant an overall decrease in mRNA expression levels of almost all measured cytokines was observed, which paralleled an improvement in diastolic left ventricular wall thickness and function. However, no straightforward relationship could be found between a specific cytokine mRNA expression pattern and the studied echocardiographic parameters. Our data suggest that the improvement in diastolic left ventricular function is associated with a general reduction of inflammation within the allograft, rather than related to a specific cytokine expression pattern.
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