Akin Tekin

Heterotopic Uterus Transplantation in a Swine Model

Background. The aim of our study was to examine the feasibility of allogeneic uterine transplantation in a large animal model. Methods. We performed heterotopic uterine transplants in genetically defined mini-pigs. Immunosuppression was tacrolimus administered intravenously for the first 12 days posttransplantation followed by oral cyclosporine maintenance immunosuppression. The graft was transplanted heterotopically in the lower abdominal cavity of the recipient. The vaginal vault was exteriorized as a stoma in the lower right abdominal wall. The uterine grafts were followed with endoscopies and biopsies. Results. Ten transplants were performed. Follow-up was until July 2008. At the end of the follow-up period, 5 animals were alive and healthy, 0.5 to 12 months posttransplantation. There were 5 deaths due to pneumonia (n=1), intussusception of the graft (n=1), cardiorespiratory arrest during anesthesia (n=1), and complications of the stoma (n=2). Acute rejections of the graft presented during the 2nd and 3rd month posttransplantation were treated successfully with increase of the maintenance immunosuppression and steroids. Other complications included prolapse and infections of the graft stoma. Pathological changes seen in the endometrial biopsies included acute rejection and acute endometritis. Conclusion. These findings demonstrate that successful uterus transplantation in a large animal model (miniature swine) is feasible using this heterotopic model, and it can be useful for the study of these transplants.

When and why portal vein thrombosis matters in liver transplantation: A critical audit of 174 cases

Objective:To identify complications associated with different techniques utilized to treat portal vein thrombosis (PVT) during primary liver transplantation and their impact on survival. Background:PVT remains an intricate problem in liver transplantation, and the long-term outcomes of patients with PVT who undergo transplantation are not well defined. Methods:We performed a retrospective cohort analysis of all consecutive adult patients who underwent primary isolated liver transplantation from 1998 to 2009 (median follow-up period, 89 months). The outcomes of patients with PVT were compared with those without PVT. Results:Among 1379 recipients, 174 (12.6%) had PVT at the time of transplantation [83 (48%) complete and 91 (52%) partial]. Among PVT patients with reestablished physiological portal inflow (PVT: physiological group; n = 149), 123 underwent thrombectomies, 16 received interpositional vein grafts, and 10 received mesoportal jump grafts. In 25 patients, physiological portomesenteric venous circulation was not reconstituted (PVT: nonphysiological group; 18 underwent cavoportal hemitranspositions, 6 renoportal anastomoses, and 1 arterialization). The PVT: nonphysiological group suffered a significantly increased incidence of rethrombosis of the portomesenteric veins and gastrointestinal bleeding, with a marginal 10-year overall survival rate of 42% (no PVT, 61%; P = 0.002 and PVT: physiological, 55%; P = 0.043). The PVT: physiological and no PVT groups exhibited comparable survival rates (P = 0.13). No significant differences in survival were observed between complete and partial PVT as long as physiological portal flow was reestablished. Conclusions:The subset of PVT patients requiring nonphysiological portal vein reconstruction was associated with higher complication rates and suffered diminished long-term prognoses. For the most severe PVT cases, a comprehensive approach is critical to further improve outcomes.

Association Between Donor-Specific Antibodies and Acute Rejection and Resolution in Small Bowel and Multivisceral Transplantation

Background. Donor-specific antibodies (DSA) are associated with acute kidney graft rejection, but their role in small bowel/multivisceral allograft remains unclear. We carried out a prospective study to understand the impact of DSA in the setting of intestinal allograft rejection. Methods. Thirteen patients (15 grafts) were serially evaluated for DSA levels pre- and posttransplant. DSA was determined by Luminex and the results were interpreted as fluorescence intensity (FI), with FI more than 3000 considered positive. Results. The clinical rejection episodes in allografts were significantly associated with the presence of DSA (P=0.041).We obtained 291 biopsy samples from graft ileum and date-matched DSA assay reports. Sixty-three (21.65%) of the biopsies showed acute rejection. The appearance of DSA were preformed (n=5, anti-human leukocyte antigen class II=3, anti-class I and II=2), de novo (n=4, 15.25±4.72 days after transplantation, anti-class II=1, and anti-class I and II=3) and never (n=6). Among the 63 biopsies, 30(47.6%) had significant correlations with positive DSA (kappa=0.30, P<0.001) and manifested severe rejection grade (P=0.009). Conclusions. In this cohort of small bowel/multivisceral transplantation patients, there was a high incidence of DSA. The presence of DSA should alert the clinical team of a higher risk of rejection, and reduction of the FI is clinically associated with resolution. Serial endoscopy guided biopsies combined with simultaneous DSA measurement in postintestinal transplantation follow-up is an effective means of screening for cellular and humoral-based forms of acute rejection.

Preclinical report on allogeneic uterus transplantation in non-human primates

STUDY QUESTION Is it possible to perform allogeneic uterus transplantation (UTx) with a donation from a live donor in a non-human primate species and what immunosuppression is needed to prevent rejection? SUMMARY ANSWER Allogeneic UTx in the baboon is a donor- and recipient-safe surgical procedure; immunosuppression with induction therapy and a triple protocol should be used. WHAT IS KNOWN ALREADY UTx may become a treatment for absolute uterine factor infertility. Autologous UTx models have been developed in non-human primates with reports on long-term survival of the uterine grafts. STUDY DESIGN, SIZEAND DURATION: This experimental study included 18 female baboons as uterus donors and 18 female baboons as uterus recipients. The follow-up time was 5-8 weeks. PARTICIPANTS/MATERIALS, SETTING AND METHODS Uterus retrieval was performed with extended hysterectomy including bilateral uterine and internal iliac arteries and ovarian veins. After UTx, with vascular anastomoses unilateral to the internal iliac artery and the external iliac vein, the uterus recipients received one of the following: no immunosuppression (n = 4); monotherapy (oral slow release tacrolimus) (n = 4) or induction therapy (antithymocyte globulin) followed by triple therapy (tacrolimus, mycophenolate, corticosteroids; n = 10). Surgical parameters, survival, immunosuppression and rejection patterns were evaluated. MAIN RESULTS AND THE ROLE OF CHANCE The durations of uterus retrieval and recipient surgery were around 3 and 3.5 h, respectively. The total ischemic time was around 3 h. All the recipients and the donors survived the surgery. All the recipients presented rejection to some extent within the first weeks following UTx. In one recipient, the uterus was of normal appearance at the end of the study period. In spite of occasional high (>60 ng/ml) blood levels of tacrolimus, there was no evidence of nephrotoxicity. LIMITATIONS AND REASONS FOR CAUTION This initial non-human primate allogeneic UTx study indicates that further research is needed to optimize immunosuppression protocols in order to avoid uterine rejection. WIDER IMPLICATIONS OF THE FINDINGS The findings suggest that allogeneic UTx in primate species is feasible but continued work on this issue is needed. STUDY FUNDING/COMPETING INTEREST(S) The study was funded by the Swedish Research Council, ALF University of Gothenburg, Hjalmar Svensson Foundation and by Jane and Dan Olsson Research Foundation. The authors do not have any competing interest.

MicroRNA Signature of Intestinal Acute Cellular Rejection in Formalin-Fixed Paraffin-Embedded Mucosal Biopsies

Despite continuous improvement of immunosuppression, small bowel transplantation (SBT) is plagued by a high incidence of acute cellular rejection (ACR) that is frequently intractable. Therefore, there is a need to uncover novel insights that will lead to strategies to achieve better control of ACR. We hypothesized that particular miRNAs provide critical regulation of the intragraft immune response. The aim of our study was to identify miRNAs involved in intestinal ACR. We examined 26 small intestinal mucosal biopsies (AR/NR group; 15/11) obtained from recipients after SBT or multivisceral transplantation. We investigated the expression of 384 mature human miRNAs and 280 mRNAs associated with immune, inflammation and apoptosis processes. We identified differentially expressed 28 miRNAs and 58 mRNAs that characterized intestinal ACR. We found a strong positive correlation between the intragraft expression levels of three miRNAs (miR‐142‐3p, miR‐886‐3p and miR‐132) and 17 mRNAs including CTLA4 and GZMB. We visualized these miRNAs within cells expressing CD3 and CD14 proteins in explanted intestinal allografts with severe ACR. Our data suggested that miRNAs have a critical role in the activation of infiltrating cells during intestinal ACR. These differences in miRNA expression patterns can be used to identify novel biomarkers and therapeutic targets for immunosuppressive agents.

Liver transplantation using elderly donors: a risk factor analysis

Kim DY, Moon J, Island ER, Tekin A, Ganz S, Levi D, Selvaggi G, Nishida S, Tzakis AG. Liver transplantation using elderly donors: a risk factor analysis. 
Clin Transplant 2011: 25: 270–276.

Long-term follow-up of 23 operational tolerant liver transplant recipients.

Introduction. This is a follow-up of a withdrawal study that we previously performed on 104 liver transplant patients in which immunosuppression was gradually withdrawn over a period of 3 years. Eighty-one patients were not able to be withdrawn (rejectors), and 23 patients were successfully weaned off immunosuppression (tolerants). Methods. In this study, we present their follow-up after the end of the withdrawal study: we compared the results of the tolerant patients (n=23) with those of the rejectors (n=81). Follow-up was until February 2010. Results. Operational tolerant patients were off immunosuppression for an average of 7.27±0.28 years. Patient survival in the tolerant and the rejector groups was 63.66% and 74.25%, respectively (P=not significant). A patient in the rejector group received two retransplants for chronic rejection. In the rejector group, 19 patients presented 26 rejection episodes: clinically suspected (n=19) and biopsy-proven mild (n=4), moderate (n=2), and severe (n=1) rejection episodes. A tolerant patient had a moderate rejection episode of 5.3 years after immunosuppression withdrawal. In the rejector group, five patients received a kidney transplant and four more are on dialysis versus a tolerant patient on dialysis. Freedom from rejection in the tolerant and rejector groups was 95% and 73%, respectively (P<0.05), and freedom from renal replacement treatment was 83.33% vs. 44.58%, respectively (P=not significant). Conclusions. Long-term outcomes of operationally tolerant liver transplant patients are at least as good as those of control patients. Operational tolerance is not a permanent state, and continuous vigilance is required to detect rejection episodes.

Liver transplantation for hepatocellular carcinoma in the model for end-stage liver disease era.

BACKGROUND Since March 2002, the United Network for Organ Sharing liver allocation policy has given extra priority to patients with hepatocellular carcinoma (HCC) who meet specific medical criteria. This study reviews our experience with liver transplantation for HCC under this system. STUDY DESIGN Between March 2002 and April 2009, 244 patients with HCC underwent primary liver or liver-kidney transplantation under the current allocation system at the University of Miami. Outcomes including HCC recurrence-free survival (RFS) and patient survival (PS) were assessed retrospectively. Clinical variables that predicted outcomes were analyzed. RESULTS The median time from listing to transplantation was 48 days. The median follow-up was 27.4 months, with an observed recurrence rate of 10.7%. The RFS rates at 1, 3, and 5 years after transplantation were 96.0%, 89.0%, and 83.6%, respectively. The PS rates at 1, 3, and 5 years after transplantation were 86.3%, 71.5%, and 61.7%, respectively. Among patients diagnosed with T2 HCC, a trend toward improved RFS was observed for those who received preoperative ablative therapy; PS was similar (p > 0.05). Outcomes (RFS and PS) for patients with T3 HCC were similar to those in patients with T2 HCC (p > 0.05). Patients with an alpha-fetoprotein >100 ng/mL had an RFS that was inferior to that in patients with an alpha-fetoprotein < or =100 ng/mL (p < 0.0001). CONCLUSIONS Under the current allocation system, transplantation for HCC results in excellent RFS; PS depends on factors other than HCC; the value of preoperative ablative therapy for patients with T2 HCC is uncertain; the current criteria could be expanded to include selected patients with T3 HCC; and an elevated AFP level is associated with an increased risk of HCC recurrence after transplantation.

Clinical-scale isolation of interleukin-2-stimulated liver natural killer cells for treatment of liver transplantation with hepatocellular carcinoma.

Tumor recurrence is the main limitation of liver transplantation (LT) in patients with hepatocellular carcinoma (HCC) and can be promoted by immunosuppressants. However, there is no prevention or treatment for HCC recurrence after LT. Here we describe a clinical-scale method for an adoptive immunotherapy approach that uses natural killer (NK) cells derived from deceased donor liver graft perfusate to prevent tumor recurrence after LT. Liver mononuclear cells (LMNCs) that were extracted from deceased donor liver graft perfusate contained a high percentage of NK cells (45.0 ± 4.0%) compared with peripheral blood mononuclear cells (PBMCs) (21.8 ± 5.2%) from the same donor. The CD69 activation marker and the natural cytotoxicity receptors, NKp44 and NKp46, were expressed at high levels in freshly isolated liver NK cells. Furthermore, interleukin-2 (IL-2)-stimulated NK cells showed greater upregulation of activation markers and the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), which is critical for NK cell-mediated antitumor cell death and increased production of interferon. Moreover, IL-2 stimulation induced LMNCs to exhibit a strong cytotoxicity against NK-susceptible K562 target cells compared with PBMCs (p < 0.01). Finally, we also showed that the final product contained a very low T-cell contamination (0.02 ± 106 cells/kg−1), which reduces the risk of graft-versus-host disease (GVHD). Collectively, our results suggest that the adoptive transfer of IL-2-stimulated NK cells from deceased donor liver graft perfusate could be a promising treatment for LT patients with HCC.

Citrulline level is a potent indicator of acute rejection in the long term following pediatric intestinal/multivisceral transplantation.

Citrulline has been advocated as a marker for acute cellular rejection (ACR) in intestinal transplantation; however, its significance as a forewarning in the long‐term follow‐up remains unknown. This study aimed to investigate the association between citrulline levels and the grading of ACR to establish a cutoff point that accurately predicts ACR beyond 3 months posttransplant in the pediatric patient population. During a 16‐year period (1995–2011), a total of 13 499 citrulline samples were prospectively collected from 111 consecutive pediatric intestinal/multivisceral transplant recipients: 2155 were obtained concurrently with intestinal biopsies. There were 185 ACR episodes observed among 74/111 (67%) patients (median follow‐up: 4.4 years). Citrulline levels were inversely proportional to the severity of ACR. Negative predictive values for any type of ACR (cutoff, 20 μmol/L) and moderate/severe ACR (cutoff, 10 μmol/L) were 95% and 99%, respectively. When patients were divided according to graft size, diagnostic accuracy using the same cutoff was identical. Similarly, subgroup analysis by the timing of citrulline measurement prior to biopsy varying from 1 to 7 days demonstrated comparable results. Citrulline is a potent indicator as a danger signal for ACR, being an exclusionary, noninvasive biomarker with excellent negative predictive values in the long term after pediatric intestinal/multivisceral transplant.