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Inhibition of tumor necrosis factor-α does not prevent cardiac allograft arteriosclerosis in the rat
Inhibition of Tumor Necrosis Factor-␣ Does Not Prevent Cardiac Allograft Arteriosclerosis in the Rat V.P. Pulkkinen, R.K. Sihvola, P.K. Koskinen, and K.B. Lemstro¨m
T
UMOR NECROSIS FACTOR-␣ (TNF-␣), produced by activated macrophages, elicits a wide range of proinflammatory activities on target cells.1 It upregulates PDGF-R␣ on vascular smooth muscle cells (SMC) inducing the major pathway leading to cardiac allograft arteriosclerosis, (ie, chronic rejection).2 Therefore, we hypothesized that targeting to a single cytokine TNF-␣ may regulate arteriosclerotic lesion formation. MATERIALS AND METHODS Inbred DA and WF rat strains were used for heterotopic cardiac transplantations, as described previously.3 In an acute rejection model, the recipients were left nonimmunosuppressed and the grafts were removed 5 days after transplantation. In a chronic rejection model, allografts were given soluble TNF receptor p80/ IgG1 Fc fusion protein (TNF-RII:Fc) or human IgG (huIgG) at a dose of 2 mg/kg per day intravenously every other day on ten occasions. Subcutaneous CsA at 2 mg/kg per day was administered for 2 weeks and 1 mg/kg per day thereafter as background immunosuppression. The grafts were removed 60 days after transplantation and analyzed for histology. TNF-␣, TNF-RI, and TNFRII mRNA and protein levels were measured with RT-PCR and immunohistochemistry, respectively.
upregulated significantly (P ⬍ .05). Strong TNF-␣ and TNF-RII immunoreactivity was localized to the medial SMC of arteries in allografts with acute rejection and arteriosclerotic lesions, suggesting that TNF-RII regulates SMC activation in arteriosclerostic lesion formation. Inhibition of the TNF-␣–mediated pathway by TNF-RII:Fc did not affect graft survival or reduce the incidence or intensity of arteriosclerotic changes (87 ⫾ 3% and 1.4 ⫾ 0.2, respectively) compared with human IgG-treated allografts (94 ⫾ 2% and 1.8 ⫾ 0.2, respectively) demonstrating that expression of TNF-␣ is not rate-limiting for SMC proliferation. Treatment with TNF-RII:Fc significantly increased intragraft IL-1 mRNA expression, suggesting that the action of TNF-␣ may be replaced by synergically acting cytokine IL-1. REFERENCES 1. Ferrari R: Pharmacol Res 40:97, 1999 2. Sihvola RK, Koskinen PK, Mylla¨rniemi M, et al: Circulation 99:2295, 1999 3. Koskinen PK, Lemstro ¨m KB, Ha¨yry PJ: Am J Pathol 146:972, 1995
RESULTS AND DISCUSSION
In acutely rejecting rat cardiac allografts, an increase in intragraft TNF-␣ and TNF-RII mRNA expression, but not TNF-RI mRNA expression, was recorded. In long-term– surviving cardiac allografts with severe arteriosclerotic changes, mRNA expression of TNF-␣ and TNF-RII was
© 2001 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
From the Cardiopulmonary Research Group, Transplantation Laboratory, University of Helsinki, and Helsinki University Central Hospital, Helsinki, Finland. Address reprint requests to Dr K.B. Lemstro¨m, Transplantation Laboratory, University of Helsinki, PO Box 21 (Haartmaninkatu 3), University of Helsinki, FIN-00014 Helsinki, Finland.
0041-1345/01/$–see front matter PII S0041-1345(00)02041-8 347
Transplantation Proceedings, 33, 347 (2001)
T
UMOR NECROSIS FACTOR-␣ (TNF-␣), produced by activated macrophages, elicits a wide range of proinflammatory activities on target cells.1 It upregulates PDGF-R␣ on vascular smooth muscle cells (SMC) inducing the major pathway leading to cardiac allograft arteriosclerosis, (ie, chronic rejection).2 Therefore, we hypothesized that targeting to a single cytokine TNF-␣ may regulate arteriosclerotic lesion formation. MATERIALS AND METHODS Inbred DA and WF rat strains were used for heterotopic cardiac transplantations, as described previously.3 In an acute rejection model, the recipients were left nonimmunosuppressed and the grafts were removed 5 days after transplantation. In a chronic rejection model, allografts were given soluble TNF receptor p80/ IgG1 Fc fusion protein (TNF-RII:Fc) or human IgG (huIgG) at a dose of 2 mg/kg per day intravenously every other day on ten occasions. Subcutaneous CsA at 2 mg/kg per day was administered for 2 weeks and 1 mg/kg per day thereafter as background immunosuppression. The grafts were removed 60 days after transplantation and analyzed for histology. TNF-␣, TNF-RI, and TNFRII mRNA and protein levels were measured with RT-PCR and immunohistochemistry, respectively.
upregulated significantly (P ⬍ .05). Strong TNF-␣ and TNF-RII immunoreactivity was localized to the medial SMC of arteries in allografts with acute rejection and arteriosclerotic lesions, suggesting that TNF-RII regulates SMC activation in arteriosclerostic lesion formation. Inhibition of the TNF-␣–mediated pathway by TNF-RII:Fc did not affect graft survival or reduce the incidence or intensity of arteriosclerotic changes (87 ⫾ 3% and 1.4 ⫾ 0.2, respectively) compared with human IgG-treated allografts (94 ⫾ 2% and 1.8 ⫾ 0.2, respectively) demonstrating that expression of TNF-␣ is not rate-limiting for SMC proliferation. Treatment with TNF-RII:Fc significantly increased intragraft IL-1 mRNA expression, suggesting that the action of TNF-␣ may be replaced by synergically acting cytokine IL-1. REFERENCES 1. Ferrari R: Pharmacol Res 40:97, 1999 2. Sihvola RK, Koskinen PK, Mylla¨rniemi M, et al: Circulation 99:2295, 1999 3. Koskinen PK, Lemstro ¨m KB, Ha¨yry PJ: Am J Pathol 146:972, 1995
RESULTS AND DISCUSSION
In acutely rejecting rat cardiac allografts, an increase in intragraft TNF-␣ and TNF-RII mRNA expression, but not TNF-RI mRNA expression, was recorded. In long-term– surviving cardiac allografts with severe arteriosclerotic changes, mRNA expression of TNF-␣ and TNF-RII was
© 2001 by Elsevier Science Inc. 655 Avenue of the Americas, New York, NY 10010
From the Cardiopulmonary Research Group, Transplantation Laboratory, University of Helsinki, and Helsinki University Central Hospital, Helsinki, Finland. Address reprint requests to Dr K.B. Lemstro¨m, Transplantation Laboratory, University of Helsinki, PO Box 21 (Haartmaninkatu 3), University of Helsinki, FIN-00014 Helsinki, Finland.
0041-1345/01/$–see front matter PII S0041-1345(00)02041-8 347
Transplantation Proceedings, 33, 347 (2001)