Comparison of granulocyte-macrophage colony stimulating factor and interleukin-1 production from human peripheral blood mononuclear cells as measured by specific radioimmunoassays.

Attention has focused on cytokine networks in which gene and protein expression of some cytokines is under the influence of other cytokines. In the present studies, we addressed the relationship between the synthesis of granulocyte-macrophage colony stimulating factor (GM-CSF) and interleukin-1 (IL-1) in human peripheral blood mononuclear cells (PBMC) stimulated with mitogens. Since bioassays for cytokines are sensitive to more than one of these factors, it was necessary to measure the amounts of IL-1 and GM-CSF independent of bioassays. A specific and sensitive (40 pg/ml) radioimmunoassay (RIA) was developed for human GM-CSF. The sensitivity of the RIA was greater when lysine residues were iodinated with Bolton-Hunter reagent than tyrosine residues using chloramine T. After stimulating PBMC with concanavalin A (Con A), the biological activity of GM-CSF was determined in bone marrow cultures and compared to immunoreactive GM-CSF; GM-CSF levels detected by bioassays and RIAs were highly correlated in two separate sets of experiments (r2 = 0.95 and 0.43). Incubation with Con A for 48 h induced more GM-CSF than stimulation by phytohemagglutinin (PHA) despite the fact that PHA stimulates large amounts of IL-1 alpha; indomethacin had no effect on Con A stimulated synthesis of GM-CSF or IL-1 alpha. In two separate studies, PBMC from 14 donors and a second group of 12 donors were incubated with Con A for 48 h and the total amount of immunoreactive IL-1 alpha and GM-CSF was determined in the same cell cultures. There was no correlation of the amount of either cytokines in these cultures.(ABSTRACT TRUNCATED AT 250 WORDS)

Magnetic resonance imaging in ischemic injury after heart transplantation in rats.

Magnetic resonance imaging with and without gadolinium (Gd)-DTPA has been shown to enable detection of coronary occlusive ischemic injury and heart transplant rejection. This study was performed to examine findings on magnetic resonance images associated with ischemic injury after heart transplantation in rats. Magnetic resonance imaging was performed immediately before death in 22 rats, between 1 and 90 days after isogeneic (Lewis grafts, Lewis host; or Fischer graft, Fischer host) heterotopic heart transplantation. Ischemic injury, characterized histologically by cellular infiltration or myocyte necrosis, correlated inversely with graft duration. It was graded as moderate to severe in 5 of 5 rats killed at 1 to 2 days, and in 0 of 9 animals killed at greater than or equal to 30 days. T2-weighted myocardial signal intensity (TR = 2.3 seconds; TE = 90 milliseconds) correlated inversely with graft duration and was significantly greater in grafts with moderate or severe histologic abnormalities than in grafts with absent or minimal changes. GD-DTPA-induced myocardial enhancement was judged on T1-weighted images (TR = 0.5 seconds, TE = 25 milliseconds). Areas of intense enhancement were present in all seven grafts with severe histologic abnormalities, but in only 3 of 15 grafts with absent to moderate histologic abnormalities. In conclusion, after heart transplantation in rats, ischemic injury causes increased T2-weighted signal intensity and Gd-DTPA-induced T1-weighted signal enhancement--findings similar to those described in transient coronary occlusive ischemia and in graft rejection. Abnormalities seen on magnetic resonance images during the first few posttransplant weeks may represent ischemic injury rather than rejection.

Total lymphoid irradiation and discordant cardiac xenografts.

Total lymphoid irradiation can prolong concordant cardiac xenografts. The effects of total lymphoid irradiation in a discordant xenograft model (guinea pig to rat) were studied with and without adjuvant pharmacologic immunosuppression. Inbred Lewis rats were randomly allocated to one of four groups. Group 1 (n = 6) served as a control group and rats received no immunosuppression. Group 2 (n = 5) received triple-drug therapy that consisted of intraperitoneal azathioprine (2 mg/kg), cyclosporine (20 mg/kg), and methylprednisolone (1 mg/kg) for 1 week before transplantation. Group 3 animals (n = 5) received 15 Gy of total lymphoid irradiation in 12 divided doses over a 3-week period. Group 4 (n = 6) received both triple-drug therapy and total lymphoid irradiation as described for groups 2 and 3. Complement-dependent cytotoxicity assay was performed to determine if a correlation between complement-dependent cytotoxicity and rejection-free interval existed. Rejection was defined as cessation of graft pulsation and was confirmed by histologic test results. Only groups 1 and 2 showed a difference in survival (group 1, 6.9 +/- 1.0 minutes; group 2, 14.2 +/- 2.7 minutes, p = 0.02). Although total lymphoid irradiation did decrease complement-dependent cytotoxicity, linear regression revealed no correlation between complement-dependent cytotoxicity and graft survival (coefficient of correlation, 0.30). Unlike concordant cardiac xenografts, total lymphoid irradiation with or without triple-drug therapy does not prolong graft survival.

Magnetic resonance imaging with gadolinium-DTPA for detecting cardiac transplant rejection in rats.

To date, no noninvasive tool has gained widespread acceptance as an adequate substitute for endomyocardial biopsy for the diagnosis and grading of cardiac transplant rejection. We examined the potential role of magnetic resonance imaging with gadolinium (Gd)-diethylenetriamine penta-acetic acid (DTPA) image enhancement for the diagnosis of cardiac graft rejection. We studied 15 rats with heterotopic cardiac transplants, nine of which received no immunosuppression, and six of which received cyclosporine, azathioprine, and methylprednisolone. The animals underwent magnetic resonance imaging, which was immediately followed by sacrifice (2-12 days after transplant). Myocardial image enhancement was assessed on T1-weighted images performed before and after administration of Gd-DTPA, 0.5 mmol/kg. Histological specimens were graded I, II, or III to indicate increasing severity of rejection. In the absence of rejection, Gd-DTPA induced mild homogeneous myocardial enhancement. Ten of 11 cases with Grade II or III rejection manifested one or more areas of intense myocardial enhancement. The extent and distribution of intense myocardial enhancement corresponded to the severity and distribution of histological rejection. Quantitative myocardial enhancement, expressed as the ratio of maximal signal intensity after Gd-DTPA to signal intensity before Gd-DTPA administration, separated Grade I animals (1.61 +/- 0.27; mean +/- SD) from Grades II (2.89 +/- 0.58) and III (3.10 +/- 0.77; p less than 0.01) animals. In conclusion, cardiac transplant rejection is characterized by intense T1-weighted image enhancement after administration of Gd-DTPA. Magnetic resonance imaging with Gd-DTPA thus has potential application in the clinical diagnosis of cardiac transplant rejection.