Fred Sanfilippo

The effect of soluble complement receptor type 1 on hyperacute rejection of porcine xenografts

The use of xenografts (Xgs) from distantly related species to relieve the increasing shortage of organs for clinical transplantation is prevented by the occurrence of hyperacute rejection (HAR). This process, in which C activation plays a central role, cannot be inhibited with currently available immunosuppressants. In two clinically relevant xenotransplantation models, this study evaluated the effect of C inhibition using recombinant soluble complement receptor type 1 (sCR1) on HAR. In an ex vivo model in which porcine cardiac Xgs were perfused with human blood, cardiac function ceased within 34 min when the perfusate blood was untreated (n = 3). When the perfusate blood was treated with sCR1 (300 micrograms/ml), cardiac Xg function was maintained for up to 4 hr (n = 3). Immunohistologic examination of these Xgs demonstrated deposition of C3b/iC3b and C3d in Xgs perfused with untreated human blood but only C3d deposition in those Xgs perfused with sCR1-treated human blood. These findings are consistent with the cofactor activity of sCR1 for factor I-mediated degradation of deposited C3b/iC3b to C3d. Treatment with sCR1 also prevented the histopathologic changes of HAR observed when untreated blood was used as the perfusate. In an in vivo pig-to-primate heterotopic cardiac xenotransplantation model, in which porcine Xgs transplanted into untreated cynomolgus monkey recipients underwent HAR in 1 hr or less (n = 3), a single intravenous bolus of sCR1 (15 mg/kg) administered to the recipient immediately before Xg reperfusion markedly inhibited total and alternative pathway serum C activity and prolonged Xg survival to between 48 and 90 hr (n = 5). These studies confirm the important role of C activation in HAR of porcine cardiac Xgs by primates and indicate that sCR1 may be a useful agent for xenotransplantation.

Clinical and cost impact of second-opinion pathology : Review of prostate biopsies prior to radical prostatectomy

Despite numerous studies evaluating second-opinion surgical programs, we are unaware of work evaluating the cost effectiveness of a second opinion for pathology prior to surgery. One of six pathologists reviewed the pathology of the outside needle biopsies of 535 consecutive men referred to Johns Hopkins Hospital for radical prostatectomy over a 12-month period (from October 1993 until October 1994) before the men underwent surgery. Of the 535 needle biopsies initially diagnosed on the outside as adenocarcinoma of the prostate, seven (1.3%) were reclassified as benign upon pathology review at Johns Hopkins Hospital. The most common lesion misinterpreted as adenocarcinoma was adenosis or less pronounced examples of adenosis consisting of foci of crowded glands (five cases). Foci of atrophy in the remaining two cases were misdiagnosed as adenocarcinoma of the prostate. Upon subsequent clinical work up, six of seven men were considered not to have adenocarcinoma, and their surgery was cancelled. The cost for reviewing all 535 preoperative needle biopsies was

The detrimental effects of delayed graft function in cadaver donor renal transplantation.

44,883, which included the cost of immunohistochemical studies for high-molecular-weight cytokeratin and repeat biopsies and ultrasounds in men whose diagnoses were reversed. The total cost of the radical prostatectomies had the six men undergone surgery was estimated at

Complement deposition in early cardiac transplant biopsies is associated with ischemic injury and subsequent rejection episodes.

85,686, including hospitalization, anesthesia, radical prostatectomy pathology, and surgery. This cost savings did not include other costs resulting from lost wages, morbidity, or potential litigation. Second-opinion pathological evaluation of prostate biopsy before radical prostatectomy is cost effective and has a major impact on clinical treatment for a subset of patients.

The Expression of HLA Antigens by Cells in the Human Cornea

Data collected prospectively on over 3800 cadaveric renal transplants performed between June 1977 and July 1982 by the 41 member institutions of the South-Eastern Organ Procurement Foundation were analyzed to determine the influence of delayed graft function (DGF) on patient and graft outcome. Approximately 35% of first graft recipients and 47% of regrafted patients were found to have DGF, as determined by the necessity for dialysis at one week posttransplant. First-graft recipients with DGF tended to include more black recipients, patients with higher peak levels of panel reactive antibody (PRA), less use of antilymphocyte serum (ALS) posttransplant, slightly longer organ preservation times and the more frequent use of organs preserved by ice alone. Multivariate (Cox) regression analysis considering DGF simultaneously with ten other potentially confounding variables showed a highly significant association between DGF and overall graft loss from all causes (P<10−5), irreversible graft rejection (P<0.001) as well as patient death (P=0.012). The differences in graft survival between first graft recipients with DGF (n=961) versus those without DGF (n=1769) at one and four years posttransplant were 46%±2 vs. 60%±1 and 28%±3 vs. 40%±2, respectively. The detrimental effect of DGF was highly significant irrespective of the source of donor organs or the type of preservation used. For first transplant recipients who recovered good graft function by one month following DGF (n=564), there was a significant decrease in eventual graft survival, as compared with patients who had graft function at one month but no prior history of DGF (n=1407; P=0.008). However, patients with history of DGF who had good graft function at six months (n=361) showed no significant difference in longer-term graft survival when compared with similar patients with good graft function at six months but no history of DGF (n=912). Interestingly, first transplant recipients with DGF were found to have significantly better graft survival if they had received bilateral native nephrectomy at least one month prior to transplantation. These results indicate that delayed graft function following cadaver donor renal transplantation provides a significant risk for eventual graft and patient survival that is principally manifested during the first six months posttransplant. In addition, patients who recover graft function following DGF appear to also remain at higher risk for early graft loss, while pretransplant bilateral native nephrectomy may afford some protection against the detrimental effects of DGF.

Passive transfer of alloantibodies restores acute cardiac rejection in IgKO MICE

BACKGROUND Prolonged warm or cold ischemia is associated with poor survival of cardiac transplants, and ischemic changes in early posttransplantation endomyocardial biopsies correlate with the later development of chronic rejection. In animal models, tissue ischemia has been shown to activate complement. METHODS To determine whether ischemic changes in endomyocardial biopsies were associated with complement deposition, biopsies obtained 1-3 weeks after transplantation from 33 patients were evaluated immunohistologically for C4d and C3d deposition as well as for IgM, IgG, and IgA. The histological changes associated with ischemic injury were scored independently, using previously reported criteria without knowledge of the immunohistochemical results. RESULTS Diffuse capillary and pericapillary deposition of C4d or C3d were detected in endomyocardial biopsies of 14 of the 33 patients. The majority of biopsies (79%) with C4d or C3d deposits had histological evidence of ischemic injury, including eight of the nine biopsies containing both C4d and C3d deposition. In contrast, only 8 of 18 (45%) of the biopsies without C4d or C3d deposition had ischemic injury. Only trace amounts of IgM and no IgG or IgA were demonstrable in the biopsies. Only 2 of the 14 biopsies with C4d or C3d deposition had evidence of moderate acute rejection, whereas 5 of the 18 biopsies without C4d or C3d deposition had moderate acute rejection. However, C4d and C3d deposition did correlate with repeated acute rejection episodes on subsequent biopsies. CONCLUSIONS Thus, ischemic changes are associated with the activation of complement. Complement activation may in turn promote tissue injury and provide a potential target for future treatment.

Fate Of Four Cadaveric Donor Renal Allografts With Mesangial Iga Deposits

Although antigens of the human major histocompatibility complex, HLA, appear to be involved in the rejection of corneal allografts, the expression of these antigens by cells of the human cornea remains controversial. Using sensitive immunoperoxidase techniques, we readily demonstrated class I HLA antigens on corneal epithelial, stromal, and endothelial cells, regardless of donor age. Moreover, the expression of class I antigens by epithelium increased markedly from the central to the peripheral cornea. Finally, class II HLA antigens were found on cells scattered throughout both central and peripheral epithelium as well as the stroma. A cornea obtained from a donor of known HLA type expressed all expected and no unexpected polymorphic HLA determinants in the cornea, in a pattern similar to that seen for monomorphic HLA determinants.

Comparative effects of pregnancy, transfusion, and prior graft rejection on sensitization and renal transplant results.

Background. Alloantibody is an intrinsic component of the immune response to organ transplants. Although alloantibodies have been correlated with decreased graft survival, the mechanisms of alloantibody-mediated injury remain largely undefined in vivo. In the present study, we have established a model of alloantibody-mediated graft injury using B10.A (H-2a) hearts transplanted to wild type (WT) or immunoglobulin knock out (IgKO) C57BL-Igh-6 (H-2b) mice. Methods. Alloantibodies were measured in the circulation and graft by flow cytometry and in immunofluorescence staining, respectively. Intragraft cytokine mRNA expression was evaluated using a competitive template reverse transcriptase polymerase chain reaction (RT-PCR) technique. P-selectin and von Willebrand factor expression were localized by immunoperoxidase staining. The capacity of alloantibodies to restore acute cardiac allograft rejection was tested by passive transfer of monoclonal antibodies (mAbs) against donor major histocompatibility complex (MHC) class I antigens to IgKO recipients. Results. B10.A cardiac allografts are rejected acutely by WT C57BL/6 recipients, but over 50% of the cardiac allografts survived more than 50 days after transplantation in IgKO mice. Competitive template RT-PCR on the cardiac transplants demonstrated similar levels of IL-1-&agr;, IL-12 (p40), TNF-&agr;, IL-2, IFN-&ggr;, IL-4, and IL-10 mRNA in WT and IgKO recipients 8–10 days after transplantation, indicating that macrophage- and T-cell-dependent immune responses were intact in IgKO recipients. The rejection of B10.A hearts in WT recipients was characterized by interstitial and perivascular cellular infiltration; IgG, IgM, and complement (C3) deposition; vascular cell injury and intravascular platelet aggregation; and release of von Willebrand factor and P-selectin. In IgKO recipients the lower degree of vascular injury in the absence of alloantibody responses was reflected by the lack of release of von Willebrand factor and P-selectin, which remained confined to cytoplasmic storage granules of endothelial cells and platelets. Acute rejection of cardiac allografts was restored to IgKO recipients by passive transfer of proinflammatory IgG2b mAbs against donor MHC; recipients injected with isotype-matched control mAbs did not reject. In contrast, passive transfer of IgG1 mAbs against donor MHC failed to restore acute rejection of cardiac allografts to IgKO recipients. Passive transfer of IgG2b, but not IgG1 mAbs was associated with endothelial cell activation and platelet aggregation together with the release of preformed von Willebrand factor and P-selectin from storage granules. Conclusions. Acute rejection of cardiac allografts can be reconstituted in IgKO recipients by passive transfer of IgG2b, but not IgG1 antibody. This model allows the mechanism of alloantibody-mediate graft injury to be dissected in vivo.

Effect of continuous complement inhibition using soluble complement receptor type 1 on survival of pig-to-primate cardiac xenografts

In the course of routine pretransplant cadaveric donor kidney biopsy examination, specimens from two donors were found to exhibit intense mesangial localization of IgA by immunofluorescence, with the presence of large immune complex-type deposits in these areas confirmed ultrastructurally. Both kidneys from each donor were transplanted with the ultimate result that three of the four kidneys underwent early irreversible rejection and were removed within 3 months, while the fourth kidney has maintained normal function for a period of 8 months. Morphological and immunofluorescent evaluation of the rejected kidneys at the time of nephrectomy showed minimal residual IgA mesangial deposits, but all had changes indicative of severe acute allograft rejection. These findings suggest that glomerular lesions involving mesangial IgA deposition can resolve fairly quickly following transplantation, but that the risk of irreversible acute rejection might be greater in the recipients of such kidneys.

The contribution of terminal complement components to acute and hyperacute allograft rejection in the rat.

An analysis of all 2879 cadaveric donor transplants performed during the Southeastern Organ Procurement Foundation (SEOPF) Prospective Study from June 1977 to October 1981 was performed to determine the relative effects of previous pregnancy, pretransplant transfusion, and previous grafting on patient sensitization and renal transplant results. Previous transplantation was found to have the greatest quantitative effect on sensitization as measured by percentage of reactive antibody (PRA). Pregnancy had an intermediate effect while transfusion resulted in a very low, but statistically highly significant, increase in PRA levels. The sensitizing effect of transfusion was greatest in previous transplant recipients and minimal in parous females. Actuarial graft survival in males stratified for transfusion, transplant number, and PRA level showed significant enhancement in first graft recipients associated with transfusion regardless of PRA level. However, the benefit of transfusion was lower in regrafted recipients with PRA of ≤60% and not present at all in those with PRA of >60%. Prior graft loss did not significantly affect graft survival in unsensitized (PRA = 0) patients, but was associated with decreased graft survival in sensitized patients. Primary graft survival in females stratified by PRA level, transfusion, and prior pregnancy revealed a mildly detrimental effect of pregnancy in untransfused females, and no beneficial effect of pretransplant transfusion in nonpregnant females. However, prior pregnancy appeared to be beneficial in transfused females and transfusion was beneficial in previously pregnant females. Untransfused females without prior pregnancy had significantly better graft survival than untransfused males, irrespective of PRA level. The effect of sensitization on graft survival stratified for transfusion, prior pregnancy, and previous transplantation was of significant detriment only in transfused, regrafted males and transfused, previously pregnant females. This study, while limited only to patients who received transplants, indicates that transfusion, prior pregnancy, and previous transplantation should be considered as associated variables when their effects on sensitization and graft outcome are analyzed.