Cytomegalovirus infection induces anti-endothelial cell antibodies in cardiac and renal allograft recipients.

Cytomegalovirus (CMV) infection represents a significant morbidity factor for transplant recipients. CMV infection has an association with the development of allograft rejection (AR) through graft endothelial cell (EC) damage, but the mechanisms are not yet clear. There are few reports addressing the role of humoral immunity in vascular EC injury mediated by CMV infection whereas many reports are available regarding the mechanism(s) of CMV-associated allograft EC injury mediated by cellular immunity. Here we examine the incidence of CMV infection in 40 cardiac and 25 renal allograft recipients using polymerase chain reaction (PCR) techniques. We also monitored sera for the development of anti-EC antibodies (AECA) using an ELISA with human umbilical vein ECs as targets, and IL-2 levels using an ELISA. AECA levels (immunoglobulin-G and immunoglobulin-M) were significantly elevated in allograft recipients who demonstrated CMV-PCR positivity when compared with the CMV-PCR negative group (IgG: 23.1 +/- 16.4 vs 4.7 +/- 4.5, p < 0.0001; IgM: 47.0 +/- 53.6 vs 7.0 +/- 11.2, p < 0.0001). Serum AECA (IgG and IgM) levels increased one to four weeks after CMV DNA was detected and elevated AECA levels persisted for at least one to two months, and sometimes for several months. Elevated AECA levels correlated well with serum IL-2 levels. These results suggest that CMV infection is associated with an increased humoral immune response to EC antigens, which may be a risk factor for vascular rejection, chronic rejection and decreased allograft survival.

Correlation of cytomegalovirus DNA levels with response to antiviral therapy in cardiac and renal allograft recipients.

BACKGROUND: Cytomegalovirus (CMV) infection represents a significant morbidity factor for transplant recipients. A rapid, sensitive, specific, and reliable test is desirable for early detection of CMV infection and for monitoring the efficacy of antiviral therapy. METHODS: We examined the incidence of CMV infection in 95 cardiac and 25 renal allograft recipients followed for up to 3 years using qualitative and quantitative polymerase chain reaction (PCR) techniques. Results were subsequently correlated with clinical findings. Of the 236 samples analyzed by the CMV PCR, 84 and 20 were also analyzed by blood buffy coat culture and anti-CMV antibody IgM assays, respectively. RESULTS: The sensitivity of the CMV PCR was found to be superior to that of the other assays, although the specificity of the blood buffy coat culture is as good as that of the CMV PCR, which is higher than that of the anti-CMV antibody IgM assay. CMV infection was detected by the CMV PCR in 17 of 95 cardiac and 9 of 25 renal transplant recipients. Clinical symptoms were observed when > or =500 copies of CMV DNA/1 microg of total DNA were detected by a quantitative CMV PCR assay using an external control CMV plasmid; however, some patients had symptoms when 50-100 copies were present. The levels of CMV DNA detected varied (50-1000 copies) in patients who developed asymptomatic CMV infection. The CMV DNA levels decreased to 50-100 copies 1-2 weeks after antiviral therapy was initiated and correlated well with disappearance of clinical symptoms. CMV DNA levels decreased to < or =5 copies at 4-7 weeks after treatment. This contrasts with patients who were unresponsive to anti-CMV therapy, in whom high levels of CMV DNA (> or =500 copies) persisted for at least 5 weeks and significant levels of CMV DNA (50-100 copies) were detected for several months afterward, despite multiple courses of anti-CMV therapy. Clinical symptoms also did not disappear during this period of observation. CONCLUSIONS: (1) The CMV PCR represents a rapid, sensitive, specific, reliable test for detection of CMV infection, especially for detection of virus replication in an incipient phase. (2) The quantitative CMV PCR is useful for monitoring the efficacy of antiviral therapy to distinguish patients who respond to therapy from those who do not. (3) CMV DNA levels > or =500 copies/1 microg of total DNA analyzed by the quantitative CMV PCR can be used to differentiate CMV infection from other infections and rejection.

Modulation of immunoglobulin production and cytokine mRNA expression in peripheral blood mononuclear cells by intravenous immunoglobulin.

Intravenous immunoglobulin (IVIG) has the potential to regulate Ig production, but the mechanism(s) responsible for this effect is unknown. In experiments reported here, we examined the ability of IVIG to regulate Ig production in human peripheral blood mononuclear cells (PBMCs) stimulated with pokeweed mitogen (PWM). IVIG (2-10 mg/ml) showed a potent (80-85%) inhibition of PWM-stimulated IgG, IgM, and IgA production. To determine more precisely how IVIG mediated the inhibition of Ig production, we studied Ig promoting cytokine gene expression after PWM stimulation with or without IVIG (2 and 10 mg/ml) using dot-blot techniques. RNA was isolated from PBMCs at predetermined time points and probed with cDNAs specific for human cytokines (IL-1 beta, IL-2, IL-2R, IL-4, IL-5, IL-6, gamma-IFN, and TNF-alpha). IL-6 mRNA accumulation was maximal at 4.5 hr post-PWM stimulation and was inhibited 64-75% when IVIG (10 mg/ml) was present. gamma-IFN mRNA levels peaked at 72 hr poststimulation and were also 68-75% inhibited by IVIG. IL-2 mRNA levels peaked at 4.5 hr and were 23-46% inhibited by IVIG. The inhibitory effect of IVIG on production of these cytokines (IL-6 and gamma-IFN) was also observed at the protein level in sonicated PBMCs after incubation with PWM and IVIG. The mRNA levels for other cytokines were not or only minimally inhibited by IVIG. Addition of IL-6, gamma-IFN, or IL-2 partially restored Ig production in IVIG-treated PWM-stimulated cultures, suggesting that inhibition of other cytokines or another mechanism(s) independent of cytokine inhibition might also be involved, although inhibition of IL-6, gamma-IFN, and IL-2 may be one of the critical factors in the suppression of Ig production by IVIG.