Dorothy M. Painter

Early up-regulation of macrophages and myofibroblasts : A new marker for development of chronic renal allograft rejection

BACKGROUND Increased numbers of macrophages and myofibroblasts are observed to occur in chronic renal allograft rejection (CR). The aim of this study was to examine the expression of cellular markers for the macrophage and myofibroblast in early renal transplant biopsy specimens and correlate these findings with allograft outcome. METHODS The first postengraftment biopsy specimens from 53 patients who underwent renal transplantation between January 1993 and December 1995 were studied using immunohistochemistry with antibodies to alpha-smooth muscle actin, which identifies myofibroblasts and CD68, a marker for monocytes and macrophages. Patients were followed until December 1998 (mean follow-up 4.7+/-1.2 years). RESULTS Nine patients had progressed to CR by the time of the study, whereas 44 patients continued to have stable renal function. A marked increase in both macrophages (P=0.02) and myofibroblasts (P=0.04) was noted in the first biopsy specimen obtained after engraftment in the patients who developed CR compared with those with stable allograft function. There was a positive correlation between alpha-smooth muscle actin and collagen expression (P=0.0001). CONCLUSION Significant increases in macrophages and myofibroblasts occur in the first renal biopsy specimen in those patients who later develop CR.

Evidence that portal tract microvascular destruction precedes bile duct loss in human liver allograft rejection.

In liver allograft rejection, interlobular bile ducts are thought to be the main target of rejection. In contrast, in other organ allografts the capillary bed of the graft appears to be the primary target. To determine whether portal tract microvasculature is destroyed in liver allografts during rejection, we have identified portal tract microvasculature in 11 normal livers and 38 liver allograft biopsy specimens using monoclonal antibodies to capillary endothelium in immunohistochemical staining. El.5, CD31 and EL-4 antibodies identified portal microvascular endothelium in normal liver that had the morphology of capillaries. In allograft biopsies the number of microvascular structures per portal tract was reduced markedly in acute cellular rejection to 1.1 ±0.6 (n=25) and to 0.65±0.9 (n=15) in chronic ductopenic rejection compared with nonrejecting allografts (2.8± 0.6)(n=4) or normal liver (3.8±0.7)(n=11). To determine whether loss of microvascular structures preceded bile duct destruction in rejection, sections were double-stained to identify both microvasculature and bile ducts. The number of microvascular structures per bile duct was significantly lower in acute cellular rejection (0.5± 0.4)(n=18) or chronic rejection (0.3±0.4)(n=8) compared with normal liver (2.3±0.6)(n=7) (P<0.0001), demonstrating that components of the portal vasculature are destroyed prior to bile ducts. There was a correlation between the severity of rejection and the loss of microvascular structures per bile duct (P<0.001). In conclusion, in common with other allografted organs, the microvasculature of liver allografts appears to be an early target of rejection.

P53, deleted in colorectal cancer gene, and thymidylate synthase as predictors of histopathologic response and survival in low, locally advanced rectal cancer treated with preoperative adjuvant therapy

AbstractPURPOSE: Adjuvant therapy, either preoperatively or postoperatively, and modifications of surgery have been used to try to improve outcome of surgery for rectal cancer in regard to both local recurrence and survival. Assessment of prognosis in patients after resection is currently primarily based on clinicopathologic factors. These predict the subsequent behavior of the tumor only imperfectly. The aim of this study was to evaluate three potential molecular genetic markers of prognosis (p53, deleted in colorectal cancer gene, and thymidylate synthase) in Dukes Stage B and C low rectal tumors treated with adjuvant therapy and to determine whether they correlate with survival, local recurrence, or the pathologic response to adjuvant therapy (assessed by extent of tumor regression and tumor down-staging). METHODS: Sixty locally advanced low rectal tumors resected after preoperative chemoradiotherapy or radiotherapy alone were studied by immunohistochemical staining for p53, deleted in colorectal cancer gene, and thymidylate synthase. In addition, p53 gene mutations were sought by polymerase chain reaction–single-strand conformation polymorphism analysis. These results were correlated with survival, local recurrence, and pathologic response to adjuvant therapy. RESULTS: Lack of thymidylate synthase staining by immunohistochemistry was associated with tumor down-staging after preoperative chemoradiotherapy but not after radiotherapy or for these two combined groups. There was no correlation between p53, deleted in colorectal cancer gene, or thymidylate synthase immunohistochemical staining or between p53 polymerase chain reaction–single-strand conformation polymorphism and local recurrence or survival in locally advanced low rectal cancers treated with preoperative adjuvant therapies. CONCLUSION: Prediction of prognosis in patients with locally advanced low rectal cancers treated with preoperative adjuvant therapies continues to be problematic. Thymidylate synthase immunohistochemistry appears to be the most promising factor of those assessed in predicting tumor down-staging after preoperative chemoradiotherapy for locally advanced low rectal cancers.

Vascular endothelial growth factor expression in human chronic renal allograft rejection.

BACKGROUND Chronic renal allograft rejection is characterized by interstitial fibrosis and vasculopathy. Vascular endothelial growth factor (VEGF) is an endothelial mitogen with increased expression in inflammation and vasculopathy. METHODS Renal tissue from 17 patients with chronic rejection was examined for VEGF protein and the presence of CD 68-positive macrophages, and compared to biopsies from patients with temporary allograft dysfunction, acute rejection, and native kidneys with thin membrane disease. RESULTS In the chronic rejection group, there was markedly increased expression of VEGF protein in the interstitium (P<0.0001). In serial sections, VEGF colocalized with the expression of CD 68-positive macrophages. Significantly more macrophages were in the tubulointerstitium in tissue with chronic rejection than in those with temporary allograft dysfunction (P<0.005). Additionally, VEGF protein expression in the glomeruli and the vascular compartment of patients with chronic rejection was increased. CONCLUSION The up-regulation of VEGF in chronic renal allograft rejection may be important in inflammation and development of fibrosis.

A Large, Single Center Investigation Of The Immunogenetic Factors Affecting Liver Transplantation

BACKGROUND Reports on the relevance of immunogenetic factors in liver transplantation are often conflicting or inconclusive. We have, therefore, investigated a range of factors that may underlie liver graft survival. METHODS The influences of HLA, flow cytometric, and enhanced cytotoxic crossmatching and immunoglobulin (Ig)A levels on graft survival, and acute and chronic rejection were investigated for a single center involving 446 patients over 13 years. RESULTS The effect of HLA mismatching on graft survival was significant (P<10(-2)) and was reversed in recipients with autoimmune diseases (P<0.5x10(-2)), whereas the effect of HLA mismatches on the level of acute rejection was detrimental in all recipients. There was a significant effect of a positive cytotoxic crossmatch on 3-month (P<10(-5)) and 1-year (P<10(-4)) graft survival, and an additional effect of the flow cytometric crossmatch was seen for chronic rejection (P<10(-2)) and acute rejection (P<10(-2)). Recipients with HLA-A1,B8,DRB1*0301 had higher levels of acute rejection (P<0.5x10(-2)), and recipients who received an ABO compatible-nonidentical transplant have a significantly higher risk (P<10(-2)) of developing chronic rejection. Finally, the beneficial effect of high serum IgA and, specifically, IgA anti Fab, seen in renal transplants was not evident in liver transplants, and in fact the opposite may be true, at least for acute rejection (P<0.5x10(-2)). CONCLUSIONS By separating the recipients with autoimmune disease from other patients and by including acute and chronic rejection as outcome parameters, we have used the power of a large single-centre study to delineate the significance of some of the important immunogenetic factors involved in liver transplantation.

Post-transplant quasispecies pattern remains stable over time in patients with recurrent cholestatic hepatitis due to hepatitis C virus

BACKGROUND/AIMS Several studies have shown that cholestatic recurrent hepatitis is associated with very high HCV RNA loads in liver transplant recipients. The aim of this study was to investigate whether a correlation exists between cholestatic hepatitis post-transplant and the population of viral quasispecies. METHODS One hundred and nine serial sera samples were tested from 15 recurrent HCV patients. Four of these patients showed severe cholestatic recurrent hepatitis, 11 patients demonstrated non-cholestatic recurrent hepatitis post-transplant. Quasispecies were detected by RT-PCR amplification of the HVR1 followed by single-stranded conformational polymorphism analysis. RESULTS Forty-one samples from four cholestatic patients were tested. All four patients showed very stable quasispecies patterns post-transplant. One cholestatic patient also showed a stable quasispecies band pattern following retransplantation, again associated with severe cholestatic hepatitis. Sixty-eight samples were tested from the 11 non-cholestatic patients. In contrast, these patients showed significantly more quasispecies bands than the cholestatic patients. The noncholestatic patients also displayed fluctuating band patterns post-transplant. Serial samples were tested after retransplantation in one non-cholestatic patient, with a fluctuating pattern again seen. There was a negative correlation between the HCV RNA load in serum and the number of quasispecies bands. CONCLUSIONS Stable hepatitis C viral quasispecies associated with persistently high viral load in post-transplant cholestatic hepatitis suggest that viral escape from immune pressures may play a role in the pathogenesis of this condition.

Analysis of initial poor graft function after orthotopic liver transplantation: experience of an australian single liver transplantation center

IN ORTHOTOPIC LIVER transplantation (OLTx), initial poor graft function (IPGF) may be related to the quality of the donor organ, long cold and warm ischemic times, the medical status of the recipient, the surgical technique used, reduced size liver grafts. With IPGF, subsequent patient morbidity and mortality is increased. Primary graft nonfunction (PNF) is the most serious end result of initial poor allograft function and may occur in 1.4% to 8.5% of cases after OLTx and may require urgent re-transplantation to avoid patient mortality. Despite developments in preservation solutions, operative procedures, and new immunosuppressive agents, IPGF continues to be a major complication immediately after OLTx. The definition and criteria used to diagnose IPGF following OLTx is inconsistent in the literature. Evaluation of early graft dysfunction determined by a high level of transaminases [aspartate aminotransferase (AST) or alanine aminotransferase (ALT)] and coagulation activity are often used as appropriate criteria within 7 days after OLTx.) The present study was undertaken as a retrospective analysis of donor and recipient demographics and intraand postoperative biochemical and pathological features to determine the factors leading to IPGF at a single transplant center in Sydney, Australia.

Posttransplant Administration of Donor Leukocytes Induces Long-Term Acceptance of Kidney or Liver Transplants by an Activation-Associated Immune Mechanism

Donor leukocytes play a dual role in rejection and acceptance of transplanted organs. They provide the major stimulus for rejection, and their removal from the transplanted organ prolongs its survival. Paradoxically, administration of donor leukocytes also prolongs allograft survival provided that they are administered 1 wk or more before transplantation. Here we show that administration of donor leukocytes immediately after transplantation induced long-term acceptance of completely MHC-mismatched rat kidney or liver transplants. The majority of long-term recipients of kidney transplants were tolerant of donor-strain skin grafts. Acceptance was associated with early activation of recipient T cells in the spleen, demonstrated by a rapid increase in IL-2 and IFN-γ at that site followed by an early diffuse infiltrate of activated T cells and apoptosis within the tolerant grafts. In contrast, IL-2 and IFN-γ mRNA were not increased in the spleens of rejecting animals, and the diffuse infiltrate of activated T cells appeared later but resulted in rapid graft destruction. These results define a mechanism of allograft acceptance induced by donor leukocytes that is associated with activation-induced cell death of recipient T cells. They demonstrate for the first time that posttransplant administration of donor leukocytes leads to organ allograft tolerance across a complete MHC class I plus class II barrier, a finding with direct clinical application.

Placental endothelial nitric oxide synthase localization and expression in normal human pregnancy and pre-eclampsia

1. The aim of the present study was to investigate whether pre‐eclampisa, a state of placental hypoxia, is associated with placental abnormalities in the amount, distribution and expression of enothelial nitric oxide synthase (eNOS).

Histological tumour response to pre-operative combined modality therapy in locally advanced rectal cancer

Pre‐operative combined modality therapy (CMT) is used in locally advanced rectal cancer. Its use affects the clinicopathological staging based on the resected specimen. Assessment of the tumour response in the resected specimen may provide prognostic information. This study was undertaken to determine the histological response to pre‐operative chemoradiation and to assess the interobserver reliability of a newly developed tumour response grading system for rectal cancer.