INDUCTION OF TNFα AND TNFβ GENE EXPRESSION IN RAT CARDIAC TRANSPLANTS DURING ALLOGRAFT REJECTION

Pizarro, Theresa T.; Malinowska, Krystyna; Kovacs, Elizabeth J.; Clancy, John; Robinson, John A.; Piccinini, L. A.

doi:10.1097/00007890-199308000-00029

Cited by 63 publications

(28 citation statements)

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“…Accurate sample concentrations of TNF␣ were determined by comparing their respective absorbances with those obtained for the standards plotted on a standard curve. This immunoreactive method has been shown to be accurate for measuring TNF␣ in rats (19).…”

Section: Measurementsmentioning

confidence: 95%

Acute bacterial pneumonia in rats increases alveolar epithelial fluid clearance by a tumor necrosis factor-alpha-dependent mechanism.

Rezaiguia¹,

Garat²,

Delclaux³

et al. 1997

J. Clin. Invest.

160

118

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To study the rate and regulation of alveolar fluid clearance in acute pneumonia, we created a model of Pseudomonas aeruginosa pneumonia in rats. To measure alveolar liquid and protein clearance, we instilled into the airspaces a 5% bovine albumin solution with 1.5 Ci of 125 I-human albumin, 24 h after intratracheal instillation of bacteria. The concentration of unlabeled and labeled protein in the distal airspaces over 1 h was used as an index of net alveolar fluid clearance. Since there was histologic evidence of alveolar epithelial injury, several methods were used to measure alveolar fluid clearance, including the use of experiments in rats with blood flow and the use of experiments in rats without blood flow, so that movement across the epithelial barrier would be minimized in the latter group. The results with each method were identical. We found that P. aeruginosa pneumonia increased alveolar liquid clearance over 1 h by 48% in studies with blood flow, and by 43% in rats without blood flow, compared with respective controls ( P Ͻ 0.05). In both studies, this increase was inhibited with amiloride. However, propranolol had no inhibitory effect, thus ruling out a catecholamine-dependent mechanism to explain the increase in alveolar fluid clearance. An antitumor necrosis factor-␣ neutralizing antibody, instilled into the lung 5 min before bacteria, prevented the increase in alveolar liquid clearance in rats with pneumonia ( P Ͻ 0.05). Also, TNF ␣ (5 g) instilled in normal rats increased alveolar liquid clearance by 43% over 1 h compared with control rats ( P Ͻ 0.05). In normal rats instilled with TNF ␣ , propranolol had no inhibitory effect. In conclusion, gram-negative pneumonia markedly upregulates net alveolar epithelial fluid clearance, in part by a TNF ␣ -dependent mechanism. This finding provides a novel mechanism for the upregulation of alveolar epithelial sodium and fluid transport from the distal airspaces of the lung. ( J. Clin. Invest. 1997. 99:325-335.)

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Section: Measurementsmentioning

confidence: 95%

Acute bacterial pneumonia in rats increases alveolar epithelial fluid clearance by a tumor necrosis factor-alpha-dependent mechanism.

Rezaiguia¹,

Garat²,

Delclaux³

et al. 1997

J. Clin. Invest.

160

118

View full text Add to dashboard Cite

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“…Total RNA was extracted and prepared for Northern analysis as described earlier, using random primed cDNA probes for TNFa and GAPDH. Cytosolic TNFa protein was prepared using a modification of the method of Pizarro et al (31 ). Briefly, the cell pellet and supernatant were each subjected to two rapid freeze-thaw cycles, by rapidly immersing the cells in liquid nitrogen and warming the cells in a 37°C water bath.…”

Section: Cellular Source For Myocardial Tnfa Production In Vitromentioning

confidence: 99%

“…The cells were then centrifuged at 1,500 g for 10 min and the cell pellets frozen immediately in liquid nitrogen. The cytosolic portions of the diluent-and endotoxin-stimulated cells were prepared exactly as described earlier (31). The amount of TNFa per myocyte culture was normalized by the total amount of cytosolic protein per culture; protein determinations were performed using a commercially available assay (BCA; Pierce), using BSA as a standard.…”

Section: Cellular Source For Myocardial Tnfa Production In Vitromentioning

confidence: 99%

Tumor necrosis factor-alpha gene and protein expression in adult feline myocardium after endotoxin administration.

Kapadia¹,

Lee²,

et al. 1995

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TNFa mRNA and protein biosynthesis were examined in the adult feline heart after stimulation with endotoxin. When freshly isolated hearts were stimulated with endotoxin in vitro, de novo TNFa mRNA expression occurred within 30 min, and TNFa protein production was detected within 60-75 min; however, TNFa mRNA and protein production were not detected in diluent-treated hearts. Immunohistochemical studies localized TNFa to endothelial cells, smooth muscle cells, and cardiac myocytes in the endotoxin-treated hearts, whereas TNFa immunostaining was absent in the diluent-treated hearts. To determine whether the cardiac myocyte was a source for TNFa production, two studies were performed. First, in situ hybridization studies, using highly specific biotinylated probes, demonstrated TNFa mRNA in cardiac myocytes from endotoxin-stimulated hearts; in contrast, TNFa mRNA was not expressed in myocytes from diluent-treated hearts. Second, TNFa protein production was observed when cultured cardiac myocytes were stimulated with endotoxin, whereas TNFa protein production was not detected in the diluent-treated cells. The functional significance of the intramyocardial production of TNFa was determined by examining cell motion in isolated cardiac myocytes treated with superfusates from endotoxinand diluent-stimulated hearts. These studies showed that cell motion was depressed in myocytes treated with superfusates from the endotoxin-treated hearts, but was normal with the superfusates from the diluent-treated hearts; moreover, the negative inotropic effects of the superfusates from the endotoxin-treated hearts could be abrogated completely by pretreatment with an anti-TNFa antibody. Finally, endotoxin stimulation was also shown to result in the intramyocardial production of TNFa mRNA and protein in vivo.Thus, this study shows for the first time that the adult mammalian myocardium synthesizes biologically active TNFa. (J. Clin. Invest. 1995. 96:1042-1052

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“…Increased levels of TNF-␣ and LT␣ have been detected in rejecting allografts in both clinical studies and animal models (3)(4)(5)(6)(7)(8)(9), and inhibition of these cytokines has been associated with prolonged allograft survival (10 -12). However, the mechanism(s) by which TNF-␣ and LT␣ mediate allograft rejection remains incompletely understood.…”

mentioning

confidence: 99%

Prolonged Allograft Survival in TNF Receptor 1-Deficient Recipients Is Due to Immunoregulatory Effects, Not to Inhibition of Direct Antigraft Cytotoxicity

McKee

DeFina

et al. 2002

The Journal of Immunology

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TNF-α and lymphotoxin (LT)α have been shown to be important mediators of allograft rejection. TNF-R1 is the principal receptor for both molecules. Mice with targeted genetic deletions of TNF-R1 demonstrate normal development of T and B lymphocytes but exhibit functional defects in immune responses. However, the role of TNF-R1-mediated signaling in solid organ transplant rejection has not been defined. To investigate this question, we performed vascularized heterotopic allogeneic cardiac transplants in TNF-R1-deficient (TNF-R1−/−) and wild-type mice. Because all allografts in our protocol expressed TNF-R1, direct antigraft effects of TNF-α and LTα were not prevented. However, immunoregulatory effects on recipient inflammatory cells by TNF-R1 engagement was eliminated in TNF-R1−/− recipients. In our study, cardiac allograft survival was significantly prolonged in TNF-R1−/− recipients. Despite this prolonged allograft survival, we detected increased levels of CD8 T cell markers in allografts from TNF-R1−/− recipients, suggesting that effector functions, but not T cell recruitment, were blocked. We also demonstrated the inhibition of multiple chemokines and cytokines in allografts from TNF-R1−/− recipients including RANTES, IFN-inducible protein-10, lymphotactin, and IL-1R antagonist, as well as altered levels of chemokine receptors. We correlated gene expression with the physiologic process of allograft rejection using self-organizing maps and identified distinct patterns of gene expression in allografts from TNF-R1−/− recipients. These findings indicate that in our experimental system TNF-α and LTα exert profound immunoregulatory effects through TNF-R1.

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INDUCTION OF TNFα AND TNFβ GENE EXPRESSION IN RAT CARDIAC TRANSPLANTS DURING ALLOGRAFT REJECTION

Cited by 63 publications

References 0 publications

Acute bacterial pneumonia in rats increases alveolar epithelial fluid clearance by a tumor necrosis factor-alpha-dependent mechanism.

Acute bacterial pneumonia in rats increases alveolar epithelial fluid clearance by a tumor necrosis factor-alpha-dependent mechanism.

Tumor necrosis factor-alpha gene and protein expression in adult feline myocardium after endotoxin administration.

Prolonged Allograft Survival in TNF Receptor 1-Deficient Recipients Is Due to Immunoregulatory Effects, Not to Inhibition of Direct Antigraft Cytotoxicity

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