Rachel M. McKenna

Anti-HLA antibodies after solid organ transplantation.

We have cited more than 23 studies showing that de novo development of anti-HLA antibodies is associated with increased acute and chronic rejection and decreased graft survival in kidney, heart, lung, liver, and corneal transplants. Antibodies to both HLA class I and class II antigens seem to be detrimental. Antibodies of the IgG isotype and possibly the IgM isotype were clinically relevant. Most studies showed that donor-specific antibodies were associated with rejection and graft loss. Therefore, HLA antibodies provide a clinical readout for patient alloreactivity that may have the ability to distinguish graft dysfunction due to immunologic and nonimmunologic causes. Antibody may act as a critical trigger for rejection of allografts and may serve as an early indicator of a slowly smoldering chronic rejection that is not manifested at a given time by biochemical measures such as serum creatinine levels. The effectiveness of various drugs on chronic rejection should be evaluable by their effects on HLA antibody production. We predict that recently developed ELISA and flow cytometry techniques using purified HLA antigen will increase the clinical relevance of posttransplantation HLA antibody monitoring by (1) allowing the detection of low levels of donor antibody; (2) easily distinguishing the isotype and target (HLA class I or class II) of the antibodies; and (3) correlating the antibody with specific graft pathology.

Neointimal and Tubulointerstitial Infiltration by Recipient Mesenchymal Cells in Chronic Renal-Allograft Rejection

BACKGROUND Tissue remodeling depends on mesenchymal cells (fibroblasts and myofibroblasts) and is a prominent feature of chronic renal-transplant rejection. It is not known whether the mesenchymal cells that participate in remodeling originate locally or from circulating precursor cells. METHODS We obtained biopsy specimens of renal allografts from six male recipients of an allograft from a female donor, four female recipients of an allograft from a male donor, two male recipients of an allograft from a male donor, and two female recipients of an allograft from a female donor. All the allografts were undergoing chronic rejection. All but two specimens were obtained within six months after transplantation. We used immunohistochemical methods to identify mesenchymal cells with smooth-muscle alpha-actin and in situ hybridization to identify mesenchymal cells with Y-chromosome DNA. RESULTS No Y-chromosome bodies were identified in the case of the two renal-allograft specimens in which both the donor and the recipient were female. In the case of the two renal-allograft specimens in which both the donor and the recipient were male, approximately 40 percent of mesenchymal cells contained a Y-chromosome body. In the case of the six specimens in which the donor was female and the recipient was male, a mean (+/-SD) of 34+/-16 percent of mesenchymal cells in the neointima, 38+/-12 percent of such cells in the adventitia, and 30+/-7 percent of such cells in the interstitium contained the Y-chromosomal marker, indicating that they originated from the recipient rather than the donor. In the case of the four renal-allograft specimens in which the donor was male and the recipient was female, the respective values were 24+/-15 percent, 33+/-9 percent, and 23+/-8 percent, indicating a persistent population of donor mesenchymal cells. CONCLUSIONS The presence of mesenchymal cells of host origin in the vascular and interstitial compartments of renal allografts undergoing chronic rejection provides evidence that a circulating mesenchymal precursor cell has the potential to migrate to areas of inflammation.

Computerized Image Analysis of Sirius Red–Stained Renal Allograft Biopsies as a Surrogate Marker to Predict Long-Term Allograft Function

Chronic allograft nephropathy (CAN) is a major problem in posttransplant management. The lack of a reliable and early surrogate marker of CAN has hampered patient care and research. In this study, the Cortical Fractional Interstitial Fibrosis Volume (V(IntFib)), quantitated with computerized image analysis of Sirius Red-stained protocol biopsies, was examined as a potential surrogate for time to graft failure (TTGF) in 68 renal allograft recipients. At 6 mo posttransplant, V(IntFib) was highly correlated with TTGF (r = 0.64, P < 0.001). Both the Banff Chronic Sum and the Acute Sum Scores were also correlated with TTGF, but less strongly (r = 0.28, P < 0.02; r = 0.35, P < 0.003, respectively). As V(IntFib) was not correlated with the Banff Chronic Score, a multivariate model was created that incorporated V(IntFib) and both Acute and Chronic Banff pathology. This model was highly correlated with TTGF (r = 0.7, P < 0.0001). These findings suggest that V(IntFib) determined by computerized image analysis of Sirius Red-stained protocol biopsies at 6 mo posttransplant, with or without incorporation of Banff acute and chronic scoring, may provide an early surrogate for time to graft failure in renal allograft recipients.

Quantitation of allograft fibrosis and chronic allograft nephropathy

Abstract: Despite improvements in the prevention and treatment of acute renal allograft rejection, the long‐term survival of renal transplants has not increased. Immunologic and non‐immunologic factors contribute to the gradual deterioration of graft function and to the histologic lesion characterized by vascular and interstitial fibrosis (‘chronic rejection’). Quantitation of this process has been attempted using various invasive and non‐invasive methods. These methods, performed at different times post‐transplant, are reviewed in this article. In particular, pathology scoring systems and the potential of using computerized image analysis of biopsy material are discussed.

Peritubular capillary basement membrane reduplication in allografts and native kidney disease: A clinicopathologic study of 278 consecutive renal specimens

Background. An association has been found between transplantglomerulopathy (TG) and reduplication of peritubular capillary basementmembranes (PTCR). Although such an association is of practical and theoreticalimportance, only one prospective study has tried to confirmit. Methods. We examined 278 consecutive renal specimens (from 135transplants and 143 native kidneys) for ultrastructural evidence of PTCR. Inaddition to renal allografts with TG, we also examined grafts with acuterejection, recurrent glomerulonephritis, chronic allograft nephropathy andstable grafts (“protocol biopsies”). Native kidney specimensincluded a wide range of glomerulopathies as well as cases of thromboticmicroangiopathy, malignant hypertension, acute interstitial nephritis, andacute tubularnecrosis. Results. We found PTCR in 14 of 15 cases of TG, in 7 transplantbiopsy specimens without TG, and in 13 of 143 native kidney biopsy specimens.These 13 included cases of malignant hypertension, thrombotic microangiopathy,lupus nephritis, Henoch-Schonlein nephritis, crescentic glomerulonephritis,and cocaine-related acute renal failure. Mild PTCR in allografts without TGdid not predict renal failure or significant proteinuria after follow-upperiods of between 3 months and 1year. Conclusions. We conclude that in transplants, there is a strongassociation between well-developed PTCR and TG, while the significance of mildPTCR and its predictive value in the absence of TG is unclear. PTCR alsooccurs in certain native kidney diseases, though the association is not asstrong as that for TG. We suggest that repeated endothelial injury, includingimmunologic injury, may be the cause of this lesion both in allografts andnativekidneys.

PROTOCOL BIOPSIES IN RENAL TRANSPLANTATION : RESEARCH TOOL OR CLINICALLY USEFUL ?

Early protocol biopsies of stable, well functioning renal allografts reveal a high prevalence of clinically unsuspected acute and chronic pathology. It is becoming increasingly apparent that these histopathological findings are both pathogenic and predictive of long-term allograft outcome. Therefore, protocol biopsies may be required for optimal post-transplant surveillance until non-invasive methods to detect allograft inflammation are developed.

Production of Tumor Necrosis Factor Alpha and Hemodialysis

The production of TNF alpha by peripheral blood mononuclear cells (PBMC) was determined in 18 hemodialysis (HD) patients. Blood was taken from each patient before and after an HD treatment. Both pre- and post-HD PBMC produced significantly more TNF alpha than controls (TNF alpha units/ml; mean +/- SEM; controls 3.1 +/- 0.7; pre-HD 9.7 +/- 3.9; post-HD 19.8 +/- 7.7, p < 0.05). In addition, post-HD PBMC produced significantly more TNF alpha than pre-HD PBMC suggesting that the HD procedure itself may activate cytokine production. This was true when PBMC were cultured in serum free medium as well as on culture with non-HD sera (human AB) and autologous sera. A positive correlation was also found between the production of TNF alpha and age in HD patients (r = 0.58; p < 0.01). Finally, normal PBMC cultured in post-HD sera produced significantly less TNF alpha than when cultured in the same sera pre-HD (p < 0.02). These findings suggest that PBMC of HD patients are chronically stimulated to produce TNF alpha which may contribute to some of the short-term and long-term complications of HD.

Interleukin 2, interferon, and lymphotoxin in renal transplant recipients

The immunocuppressive action of cyclosporine in transplantation (Tx) is thought to be due to its potent inhibition of lymphokine production by T cell. Several studies have shown a decrease in interleukin 2 (IL–2) adn interferon-Gamma (IFN-G) production of renal Tx recipients on CsA treatment and have suggested that increased in lymphokine production can be correlatede with rejection epidosodes.In this study we measured IL-2, IFN-G, and lymphotoxin (LT) production by mitogen-stmulated peripheral blood lymphocytes in eight renal Tx recipients before and at various times after Tx. IL-2 production was significantly (P<0.05) decresed by one week post-Tx compared with pre-Tx and normal levels. IFN-G production was significantly (P<0.05) decreased by one week post-Tx, after which time it returned to normal. LT production was not decreased post-Tx compared with pre-Tx or normal levels. Lymphokine production was measured every 48–72 hr in the first month post-Tx, when we failed to detect any correlation between incresed in productinin any of the three lymphokines and rejection episodes. A further group of patients were studied in whom the production of all three lymphokines was measured at the time of diagnosis of rejection adn after treatment for rejection. In only 3/5 patients did IL-2 production decrease with a return to stable graft functio, while IFN-G production did not alter in these patients. Interestingly LT production increased significantly (P<0.05) after treatment. We conclude from these studies that the usefulness of lymphokine determinations for the diagnosis of allograft rejection remians unproved.

Low frequency of infiltrating cells intensely expressing T cell cytokine mRNA in human renal allograft rejection.

Immunosuppressive drugs used in clinical transplantation block cytokine mRNA transcription in vitro, but clinical rejection episodes are common. An understanding of what cytokine message is transcribed would be helpful in determining what contributes to the success of immunosuppression and provide directions for further research aimed at targeting specific cytokines. Previous studies have examined cytokine mRNA in rejecting solid organ biopsies by the reverse transcriptase polymerase chain reaction (RT-PCR) with variable results. We used nonradioactive in situ hybridization with cytokine-specific riboprobes to determine the frequency of cells expressing cytokine mRNA in the allograft infiltrate. Kidney biopsies were obtained from patients receiving protocol biopsies and with clinical evidence of rejection. Fourteen biopsies with a pathologic diagnosis of rejection were studied. Eight showed no cytokine staining, 2 expressed IL-2, and 3 expressed IL-4 and IFN-gamma. The positive cells were present at a low frequency (mean 2, range 1-5 per 10 high-power fields). The proportion of kidney biopsies expressing detectable message for interleukin-2 (IL-2), interleukin-4 (IL-4), and interferon-gamma (IFN-gamma) by in situ hybridization were similar to those reported using RT-PCR. The novel finding is that these cytokines are expressed in a few strongly positive cells in the allograft infiltrate. The vast majority of infiltrating cells are negative. This suggests that either the biopsies were performed when cytokine message was not expressed at a high level or that in human allograft recipients the sustained expression of the cytokines IL-2, IL-4, and IFN-gamma may not be necessary for graft rejection.

Increased Production of Tumor Necrosis Factor Activity by Hemodialysis but Not Peritoneal Dialysis Patients

The production of tumor necrosis factor alpha (TNF alpha) activity by peripheral blood mononuclear cells (PBMC) was determined in uremic patients on chronic hemodialysis (HD; n = 27), continuous ambulatory peritoneal dialysis (CAPD; n = 19), and in patients with chronic renal failure who were not yet on dialysis (CRF-ND; n = 18). In the HD group blood was taken immediately prior to and immediately following an HD session utilizing a cellulose acetate dialyzer. Post-HD PBMC spontaneously (i.e. in serum free media) produced significantly more TNF alpha activity than the PBMC of all other patient groups as well as those of the normal controls (n = 41) (p < 0.003). Post-HD PBMC produced significantly more TNF alpha activity than pre-HD PBMC both spontaneously and in the presence of nonuremic sera (p < 0.003). PBMC prior to HD also produced significantly more. TNF alpha activity than CAPD PBMC and normal PBMC in the presence of autologous heat inactivated sera (p < 0.03). Under some culture conditions (i.e. in the presence of nonuremic sera) normal PBMC produced significantly (p < 0.003) more TNF alpha activity than CAPD PBMC. Finally, a positive correlation was found between PBMC TNF alpha activity and age for HD patients (r = 0.7, p < 0.004) but not for CAPD or CRF-ND patients. These findings suggest that PBMC of HD but not CAPD or CRF-ND patients are chronically stimulated to produce TNF alpha activity.(ABSTRACT TRUNCATED AT 250 WORDS)