CD40 regulates the processing of NF-kappaB2 p100 to p52.

The nf-kb2 gene encodes the cytoplasmic NF-kappaB inhibitory protein p100 from which the active p52 NF-kappaB subunit is derived by proteasome-mediated proteolysis. Ligands which stimulate p100 processing to p52 have not been defined. Here, ligation of CD40 on transfected 293 cells is shown to trigger p52 production by stimulating p100 ubiquitylation and subsequent proteasome-mediated proteolysis. CD40-mediated p52 accumulation is dependent on de novo protein synthesis and triggers p52 translocation into the nucleus to generate active NF-kappaB dimers. Endogenous CD40 ligation on primary murine splenic B cells also stimulates p100 processing, which results in the delayed nuclear translocation of p52-RelB dimers. In both 293 cells and primary splenic B cells, the ability of CD40 to trigger p100 processing requires functional NF-kappaB-inducing kinase (NIK). In contrast, NIK activity is not required for CD40 to stimulate the degradation of IkappaBalpha in either cell type. The regulation of p100 processing by CD40 is likely to be important for the transcriptional regulation of CD40 target genes in adaptive immune responses.

Laparoscopic-assisted renal biopsy: an alternative to open approach.

An open renal biopsy frequently is performed when the conventional percutaneous approach is contraindicated. Although many of the surgical procedures, previously performed by a traditional open technique, are now successfully accomplished by laparoscopic technique, there remains a paucity of reports on laparoscopic kidney biopsy. We describe the use of laparoscopic technique in performing native kidney biopsy in three patients and review the potential safety and accuracy of this approach. To date, laparoscopic-assisted renal biopsy has been reported in six patients. The average length of stay in our small series was 1.3 days, and there were no major or catastrophic complications. Adequate numbers of glomeruli for confirmation of microscopic diagnosis were obtained in all cases. The review of literature, in addition to our encouraging preliminary results of a small group of patients, shows a possible role of laparoscopic-assisted renal biopsy in place of an open renal biopsy.

New strategies using 'low-dose' mycophenolate mofetil to reduce acute rejection in patients following kidney transplantation.

CONTEXT: Tacrolimus, microemulsion cyclosporine (Neoral), and mycophenolate mofetil (MMF) at 2 and 3 grams daily have demonstrated superior immunosuppressive properties in several recent clinical trials involving solid-organ transplants. An effective immunosuppression may be maintained with lower doses of MMF administered with either tacrolimus or microemulsion cyclosporine. OBJECTIVE: To compare tacrolimus plus "low-dose" MMF-based immunosuppressive regimen (TMBIR) with Neoral plus "low-dose" MMF-based immunosuppressive regimens (NMBIR) among kidney transplant recipients. DESIGN: Prospective, randomized study. PATIENTS: 53 consecutive adult recipients of kidney transplant. Both groups (TMBIR and NMBIR) were equally matched on demographic characteristics. INTERVENTIONS: Participants were randomized to receive orally either tacrolimus (0.08 mg/kg twice daily) (n = 27) or Neoral (4 mg/kg twice daily) (n = 26). Both regimens were started before surgery and continued when allograft demonstrated no postoperative acute tubular necrosis. Both groups received similar "low-dose" MMF (500 mg twice daily) and prednisone (2 mg/kg/day to taper off after 1 year). Switch from tacrolimus to Neoral or vice versa was allowed after refractory rejection or serious adverse events. MAIN OUTCOME MEASURE: Acute rejection and patient and graft survival 1 year following kidney transplant. RESULTS: One-year patient survival rates were 88.9% for the TMBIR group and 100% for the NMBIR group; 1-year graft survival rates were 88.9% for the TMBIR group and 96.1% for the NMBIR group. No significant differences were found in the incidence of biopsy-confirmed acute rejection (14.8% TMBIR vs 23% NMBIR). Steroid-resistant rejections requiring cytolytic antibody therapy were higher in the NMBIR group (50% vs 25%). Three patients crossed over from NMBIR to TMBIR for refractory rejections and 1 patient crossed over from TMBIR to NMBIR for new onset seizure. Three episodes of cytomegalovirus infection were observed in the TMBIR group. Other adverse events were similar in both groups. CONCLUSIONS: Both tacrolimus and microemulsion cyclosporine combined with "low-dose" MMF and corticosteroids provide effective immunosuppression and have similar adverse events in kidney transplant recipients.

MAPkinase: a second site of G-protein regulation of B-cell activation via the antigen receptors.

Ligation of the antigen receptors on B cells transduces transmembrane signals leading to the induction of DNA synthesis. We now show that a pertussis toxin-sensitive heterotrimeric G-protein(s) of the Gi class plays a key role in the regulation of surface immunoglobulin (sIg)-mediated DNA synthesis in B cells. This site of G-protein regulation is distinct from that we have previously reported to govern the coupling of the antigen receptors on B cells to the phospholipase C-mediated hydrolysis of phosphatidylinositol-4,5-bisphosphate. We have, moreover, identified a candidate target for this new G-protein regulation by showing that mitogen-activating protein kinase (MAPkinase) activity, which plays a key role in the transduction of sIg-mediated proliferative signals in B cells, is abrogated by pre-exposure to pertussis toxin that covalently modifies and inactivates heterotrimeric G-proteins of the Gi class. Furthermore, our data suggest that this pertussis toxin-sensitive G-protein couples the antigen receptors to MAPkinase activation, at least in part, by regulating sIg-coupling to Lyn, Syk and perhaps Blk and Fyn activity, results consistent with studies in other systems which show that classical G-protein-coupled receptors recruit such protein tyrosine kinases to tranduce MAPkinase activation. Interestingly, however, this G-protein plays no apparent role in the control of up-regulation of major histocompatibility complex class II expression on B cells, suggesting that such G-protein-regulated-tyrosine kinase and MAPkinase activation is not required for the induction of this biological response following antigen receptor ligation.

Peloisis hepatis due to Bartonella henselae in transplantation: a hemato-hepato-renal syndrome.

BACKGROUND: Bacillary peliosis hepatis is an uncommon but well recognized disease due to disseminated Bartonella infections occurring predominantly in immunocompromised individuals infected with human immunodeficiency virus, type 1. A similar condition in the absence of Bartonella infection when described in organ transplant patients was felt to be secondary to azathioprine and/or cyclosporine. METHODS: Herein, we report the first case of bacillary peliosis hepatis due to systemic Bartonella henselae infection in a patient after kidney transplant. The patient presented with severe anemia, persistent thrombocytopenia, and hepato-renal syndrome. DNA-based polymerase chain reactions (PCR), which allowed direct detection of both B henselae and quintana DNA in patient's peripheral blood and liver tissue, were used. Indirect immunofluorescence assay for Bartonella serology was performed on peripheral blood. RESULTS: Histopathology of the liver biopsy demonstrated peliosis hepatis. Indirect immunofluorescence assay for Bartonella serology was positive, and B henselae DNA was identified by PCR in the peripheral blood and liver tissue. Treatment with a 3-month course of oral erythromycin resulted in an excellent clinical response. CONCLUSIONS: The present case suggests that although various anti-rejection therapies and opportunistic infections are associated with hepatic and renal dysfunction along with bone marrow suppression, the diagnostic evaluation in this situation should include liver biopsy and a careful search for evidence of systemic Bartonella infection, e.g., exposure to cats, Bartonella serology, and Bartonella DNA by PCR. A reduction in immunosuppression and prolonged therapy with antibiotics such as erythromycin will often result in early recovery.

Efficacy of oral ganciclovir in prevention of cytomegalovirus infection in post-kidney transplant patients.

Unlike parenteral gancicovir, the efficacy of oral ganciclovir in the prevention and treatment of cytomegalovirus (CMV) infection in kidney transplantation has not been well documented. This study prospectively evaluated the episodes of CMV infection within the first nine months after transplantation in renal transplant recipients treated prophylactically with oral ganciclovir (750 mg twice a day) over a period of 3 months (oral ganciclovir, N = 22), compared with patients randomly assigned as controls (controls, N = 22) who did not receive any anti-viral prophylaxis. Diagnosis of CMV infection at presentation included serological determination of CMV-specific immunoglobulin M antibodies, CMV immunofluorescence assay [standard culture tube and shell vial] (blood) and virus isolation (urine and tissue). CMV infection occurred in one patient (5%) in the oral ganciclovir group and 6 patients (27%) in the control group (p < 0.05). The episodes of biopsy proven allograft rejections were 5% (1/21) and 18% (4/22) in the oral ganciclovir and control groups, respectively. Except for one, none of these patients developed CMV infection either before or after rejection(s). Controlling for the reason (induction or treatment of rejection) for using cytolytic antibody therapies, we found that prophylactic oral ganciclovir was protective against CMV infection (adjusted risk reduction 0.83; 95% confidence interval, 0.33-0.98; p < 0.05). Neither, the CMV status of donors and recipients nor the treatment for acute rejection had any significant impact on the occurrence of CMV infections. Our results show that oral ganciclovir is an effective and well tolerated therapy in the prevention of CMV infection in renal transplant patients.

Pure red cell aplasia due to parvovirus B19 infection in solid organ transplantation.

Human parvovirus B19 disease is an infrequent but recognized rare cause of anemia in immunocompromised patients. A few cases of parvovirus B19 infections have been reported in transplant recipients, of those only four patients underwent renal transplantation. The primary immunosuppressive therapy in these patients included prednisone with either cyclosporine or tacrolimus. In one patient the disease was self-limiting, while in three others the hematocrit improved following 10-15 d of treatment with commercial intravenous immunoglobulin (IVIG). Herein, we report the fifth case of pure red cell aplasia due to parvovirus B19 infection in a renal transplant recipient who responded to a 5-d course of IVIG. To our knowledge, this is the first case of parvovirus B19 infection in a patient with solid-organ transplantation whose immunosuppressive regimen included both mycophenolate mofetil and tacrolimus and in whom an excellent clinical response was achieved with a short course of IVIG infusion.

Blood transfusions in liver recipients: a conundrum or a clear benefit in the cyclosporine/tacrolimus era?

Blood transfusions are common in patients with end-stage liver disease (ESLD), and their effects on sensitization, rejection, and liver graft survival are not well known. These effects were examined in 121 recipients of primary liver grafts, surviving > or = 30 days. Ninety-six (79%) patients received transfusions before transplantation. Transfusion recipients had significantly fewer severe or recurrent rejection episodes (18%), compared with patients who did not receive transfusions (42%, P=0.006), if the first transfusion was > or = 90 days before the transplant. Patients with alcoholic ESLD (n=49) had significantly fewer severe rejection episodes when compared with the nonalcoholic (n=72) patients (12% vs. 35%, P=0.004). The transfusion benefit was, however, more apparent and significant in the nonalcoholic (26% vs. 56% in nontransfused, P=0.02) than among the alcoholic recipients (6% vs. 25%, P=0.1). This finding is, most likely, due to a combination of a higher rate of severe rejection and the statistical power of the larger number of recipients in the nonalcoholic group. This finding is further corroborated by a multivariate analysis in which blood transfusions retained their benefit (P<0.05) independent of recipient's age and diagnosis. Graft and patient survival were not significantly different in the transfused versus nontransfused groups. Transfusion recipients had a higher panel antibody (11.4+/-23.4 vs. 2.7+/-8.1, P<0.02) but no increased risk of a positive crossmatch. In liver recipients, blood transfusions diminish the risk of rejection independent of recipient's age and the cause of ESLD.

Successful reversal of acute vascular rejection in a renal allograft with combined mycophenolate mofetil and tacrolimus as primary immunotherapy.

Despite the use of newer immunosuppressive drugs, early allograft rejection is still a major cause of graft loss and diminished graft survival. The pathological picture of acute interstitial rejection usually responds to high dose steroids, whereas the treatment of acute vascular (AVR) rejection characterized by an intense vasculitis and/or endotheliolitis is more formidable. The use of newer monoclonal antibodies specific for T lymphocytes and their subsets has only been occasionally successful in reversing AVR. Here we report a case of successful reversal of acute vascular renal allograft rejection with mycophenolate mofetil (MMF) and tacrolimus as primary therapy.

Induction of NF-AT in normal B lymphocytes by anti-immunoglobulin or CD40 ligand in conjunction with IL-4.

We show here that ligation of surface immunoglobulin or CD40 receptors in conjunction with interleukin-4 induces the nuclear factor of activated T cells (NF-AT) in normal murine B cells, which is inhibited by cyclosporin (CsA). Lipopolysaccharide, which activates B cells by a Ca(2+)-independent, CsA-resistant pathway, does not induce NF-AT. The NF-AT complex in T cells and B cells appears to be identical, comprising both Fos and Jun proteins and the 120 kDa cytosolic component of NF-AT (NF-ATp). Our transfection experiments using a trimerized NF-AT site linked to the minimal IL-2 promoter driving luciferase activity demonstrate that NF-AT is functional in A20 B-lymphoma cells. These results therefore suggest that the induction of NF-AT forms part of the B cell response to both cross-linking antigens and T cell-generated signals.

Plastic-immobilized anti-mu or anti-delta antibodies induce apoptosis in mature murine B lymphocytes.

It is widely accepted that extensive cross-linking of surface immunoglobulin (sIg) receptors on mature B cells promotes their activation and progression through the cell cycle. A commonly employed method to maximize receptor cross-linking via anti-receptor antibodies is to immobilize them on tissue culture plastic. We show here that immobilizing monoclonal anti-mu or anti-delta antibodies, which are mitogenic in solution, on plastic abrogates their capacity to induce DNA synthesis in mature murine B cells, even in the presence of interleukin-4 (IL-4). The cells do become abortively activated, as evidenced by up-regulation of major histocompatibility complex class II antigen levels, but subsequently virtually all of them die, manifesting DNA fragmentation characteristic of apoptosis. The induction of apoptosis is abrogated by the inclusion of either IL-4 or anti-CD40 antibodies in the cultures, with the two stimuli acting in concert. We believe that the system represents a polyclonal model of clonal deletion tolerance in mature B cells, such as may be induced under physiological conditions by antigens with repeating epitopes.

Liver transplant in complete situs inversus: a case report.

BACKGROUND: Several technical solutions have been proposed for patients with situs inversus (SI) needing liver transplantation. This report describes the hepatic replacement in a patient with complete SI with dextrocardia. In the only other reported patient with complete SI the donor right lobe was placed over the vertebral column and the stomach to align the donor cava with that of the recipient. METHODS: A 45-year-old woman with complete SI, suffering from Laënnec's cirrhosis with frequent upper gastrointestinal hemorrhages, underwent transplantation in June 1991. The recipient weighed 48.2 kilograms and was 155 centimeters tall. The donor weighed 77.3 kilograms and was 188 centimeters tall. The weight of the native liver was 1934 grams, and the donor liver weighed 1595 grams. RESULTS: At hepatectomy of the native liver an intact vena cava was left behind. Donor liver was rotated 90 degrees to the left, making the donor left lobe point into the left iliac fossa and the donor right lobe fall into the recipient hepatic fossa. Donor infrahepatic vena cava was sewn end-to-side to the recipient vena cava. Suprahepatic vena cava was oversewn. Donor and recipient hila were well aligned, allowing a standard arterial reconstruction and a choledocholedochostomy. Patient's recovery was uneventful with no problems during the following 6 months. CONCLUSIONS: The proposed technique for a patient with complete SI and dextrocardia offers several advantages: no need to downsize the donor in comparison with the recipient; no need for cutdown of the liver; no risk of kinking of the venous outflow; hepatic hila are aligned; and it allows for a standard arterial and biliary reconstruction. We recommend this technique as a procedure of choice for patients with SI and an intact vena cava.

The impact of operative bleeding on outcome in transplantation of the liver.

Excessive operative blood transfusion has been correlated with an increased rate of infectious complications and lower survival rate after transplantation of the liver. Two hundred and five consecutive transplants of the liver, performed between January 1988 and December 1989, were studied retrospectively to determine preoperative risk factors associated with an increased operative blood loss and to evaluate the impact of operative transfusion on the outcome of transplantation. Preoperative clinical and laboratory parameters in patients who required 10 units or more of banked erythrocytes were compared with those in patients who received less than ten units of erythrocytes. In evaluating the outcome, the two groups were compared for infection, rejection and graft and patient survival rates. The median operative blood loss for 205 patients was 5 units of banked erythrocytes (range of zero to 52, mean of 6.9 units). Only 41 patients (20 percent) required 10 units or more of erythrocytes. The significant factors on univariate analysis that were associated with an increased operative blood loss were hospitalized patients (United Network for Organ Sharing Status > or = 3), fulminant hepatic failure, previous portosystemic shunt and complete ABO mismatch. Patients who required more blood had higher incidence of coagulation abnormalities, renal dysfunction and high bilirubin levels. A stepwise logistic regression analysis model using all these parameters identified an elevated serum creatinine, decreased platelets and a prolonged partial thromboplastin time as being the strongest risk factors. Using these variables, operative bleeding of more than 10 units could be predicted accurately only 60 percent of the time (sensitivity 60.0 percent with a specificity of 69.1 percent). Septic episodes occurred more frequently in patients with an excessive operative blood loss (p < 0.05), and these patients also tended to have a higher rate of severe cytomegalovirus infections and a lower incidence of acute rejection. Patients who required more blood also had significantly prolonged stays in the intensive care units postoperatively (18.3 versus 6.3 days, p < 0.002) and lower graft and patient survival rates (p < 0.001 and p < 0.05, respectively). We conclude that intraoperative bleeding has remained a significant problem affecting the immediate outcome after transplantation of the liver. Preoperative parameters cannot predict operative bleeding accurately and the mainstay to prevent bleeding is a meticulous surgical technique during the hepatectomy and correction of coagulation abnormalities throughout the procedure.

OKT3 rescue for steroid-resistant rejection in adult liver transplantation.

The results of OKT3 use for steroid-resistant rejection rescue in adult liver transplantation were analyzed retrospectively from a single transplant center. Comparison was made with concurrent patients who had no rejection (NR) or steroid-responsive rejections (SR). The records of 290 patients who underwent 323 liver transplants from April 1985 to December 1989 were examined. The first technically successful grafts were used for this analysis (265 grafts). Follow-up was a minimum of 1 year, or until death or loss of graft. All patients received triple-drug induction immunosuppression (CsA, Aza, steroids). Initial rejection was treated with 1 g methylprednisolone bolus i.v., followed by a 5-day taper of steroids from 200 mg to 20 mg. No rejection occurred in 108 (40.8%) and SR in 86 (32.4%), and OKT3 was given for persistent rejection in 71 (26.8%). The age, sex distribution, mean follow-up, and preoperative status were similar in all three groups. The preoperative diagnoses were similar, except for fulminant liver failure, in which 19 of 20 patients experienced rejection (P < 0.0001). The median hospitalization stay was 37 days for OKT3, 27 days for SR, and 21 days for NR (P < 0.0001). The median ICU stay was similar in the three groups (OKT3, 4; SR, 4; NR, 3). Infections in the first 6 weeks, and in the period of 6 weeks to 1 year posttransplant, were of similar frequency for all three groups. By the Kaplan-Meier estimation, the graft and patient actuarial survival rates were comparable. At 1 year, the graft survival rate was 79.6% for NR, 79.8% for SR, and 67.6% for OKT3. The 1-year patient survival rate was 85.2% for NR, 83.7% for SR, and 84.5% for OKT3. Following treatment by OKT3, rejection was permanently reversed in 42 patients. A temporary response occurred in 12 patients, 16 patients failed to respond to OKT3, 2 patients died during therapy, and 6 of the nonresponders died within 12 months. Additional OKT3 treatment was attempted in 6 patients for persistent rejection within a 1-month interval from the previous OKT3 course. Of these 6, 4 developed lymphoproliferative disorder, and only 1 survived in response to drastic reduction of immunosuppression. In conclusion, OKT3 was effective as rescue therapy for adult liver transplant steroid-resistant rejection. Because of the associated morbidity and expense, OKT3 should be used in a selective fashion. Failure to respond to OKT3 is a serious complication, and should not be managed by prolonged or repeated courses, but rather by alternative means.