Livebirth and utero-placental insufficiency in Papio hamadryas baboons with uterus angiosome perfused by bilateral utero-ovarian microsurgical anastomoses alone.

STUDY QUESTION: Can the baboon uterus support a gestation to livebirth with an angiosome using microsurgically anastomosed utero-ovarian vessels and lacking uterine arteries and veins? SUMMARY ANSWER: Our angiosome model allows healthy livebirth albeit with risk of fetal growth restriction and stillbirth. WHAT IS KNOWN ALREADY: Uterine transplant can provide livebirth in humans, but requires a living donor to undergo a prolonged laparotomy for hysterectomy. In an attempt to avoid the time-consuming dissection of the uterine vein, our group has previously shown maintenance of baboon uterine menstrual function after ligation of the uterine vein and after ligation of both the uterine artery and uterine vein. STUDY DESIGN, SIZE, DURATION: In a 19-month timespan, three baboons underwent laparotomy to surgically alter uterine perfusion, and pregnancy outcomes were monitored after spontaneous mating in a breeding colony. PARTICIPANTS/MATERIALS, SETTING, METHODS: Three nulligravid female Papio hamadryas baboons in a breeding colony underwent laparotomy to ligate uterine arteries and veins along with colpotomy and cervico-vaginal anastomosis. During the same surgery, the utero-ovarian arteries and veins were microsurgically transected and re-anastomosed to themselves. Intraoperative organ perfusion was confirmed with laser angiography. After a recovery period, monitoring of menstrual cycling via menstrual blood flow and sex-skin cycling occurred, as well as uterine viability via sonography and cervical biopsy. Each baboon was released to the breeding colony for spontaneous mating and pregnancies dated by menstrual calendar and compared with early ultrasound. Delivery outcomes were monitored in each including neonate weight and placental pathology. In the event of a stillbirth, the animal was returned to the breeding colony for repeat mating attempts. After achieving a livebirth, the maternal baboon was removed from the study. MAIN RESULTS AND THE ROLE OF CHANCE: Each baboon in the trial underwent successful surgery with all uteri demonstrating viability and return of menstrual function within 10 weeks of surgery. Pregnancies occurred within two menstrual cycles in breeding colony. Baboons one and two initially had vaginal breech stillbirths, both with appearance of placental insufficiency, and one with fetal growth restriction. Baboon three underwent scheduled cesarean delivery resulting in a normally grown livebirth. Baboon one had a subsequent pregnancy resulting in a livebirth via cesarean delivery. LIMITATIONS, REASONS FOR CAUTION: Stillbirth in two of four gestations, and fetal growth restriction in one of four, are the largest concerns in our perfusion model. It remains uncertain whether the stillbirths resulted from placental insufficiency, or birth trauma from breech deliveries. WIDER IMPLICATIONS OF THE FINDINGS: The success of two livebirths warrants further attempts at improving consistency of our proposed uterine angiosome. This may allow living uterine donors to undergo less-invasive and shorter donor hysterectomy procedures. STUDY FUNDING/COMPETING INTEREST(S): The study had no external sponsors, and was supported by the Cleveland Clinic Foundation. Some equipment was loaned without cost to the research team including a laser angiography system courtesy of Novadaq Technologies, Inc. (Missaugua, ON, Canada) and a surgical microscope courtesy of DB Surgical (Coral Springs, FL, USA). B.B., K.A., M.S., K.R., M.M., P.F.E., A.T. and T.F. have no conflicts of interest. M.L.S. and S.Z. report activity as consultants for Medtronic-Covidien, and S.Z. also is a consultant to Applied Medical.

Liver, pancreas and kidney transplantation for the treatment of Wolcott-Rallison syndrome.

We present the case of a child who underwent a combined liver, pancreas and double kidney transplant following complications of Wolcott-Rallison syndrome (WRS) a rare genetic disorder that causes infantile insulin-dependent diabetes mellitus (IDDM) and often death in childhood from fulminant liver and concomitant kidney failure. WRS is characterized clinically through infantile IDDM, propensity for liver failure following viral infections, bone dysplasia and growth failure and developmental delay. Fewer than 60 cases with WRS are reported in the literature, mostly from consanguineous parents. Future episodes of liver failure, the main contributor to the increased mortality in WRS, may be prevented through timely liver transplantation. To the best of our knowledge, transplantation has not been utilized to manage complications of WRS prior to this report.

Eculizumab Salvage Therapy for Antibody-Mediated Rejection in a Desensitization-Resistant Intestinal Re-Transplant Patient.

The presence of elevated calculated panel reactive antibody (cPRA) and anti-HLA donor specific antibodies (DSA) are high risk factors for acute antibody-mediated rejection (AAMR) in intestinal transplantation that may lead to graft loss. Eculizumab has been used for the treatment of AAMR in kidney transplantation of sensitized patients that do not respond to other treatment. Here, we report a case where eculizumab was used to treat AAMR in a desensitization-resistant intestinal re-transplant patient. A male patient lost his intestinal graft to AAMR 8.14 years after his primary transplant. He received a second intestinal graft that had to be explanted a month later due to refractory AAMR. The patient remained highly sensitized despite multiple treatments. He received a multivisceral graft and presented with severe AAMR on day 3 posttransplantation. The AAMR was successfully treated with eculizumab. The patient presently maintains an elevated cPRA level above 90% but his DSAs have decreased from 18 000 MFI (mean fluorescent intensity) to below the positive cut-off value of 3000 MFI and remains rejection free with a 2-year follow-up since his multivisceral transplant. Eculizumab offers an alternative to treat AAMR in intestinal transplantation in desensitization-resistant patients.

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.

Multivisceral ex vivo surgery for tumors involving celiac and superior mesenteric arteries.

Abdominal tumors involving both roots of the celiac and superior mesenteric artery are deemed unresectable by conventional surgical methods. We performed three cases of multivisceral ex vivo surgery involving temporary removal of the entire abdominal viscera followed by vascular reconstruction, ex vivo tumor resection and autotransplantation of excised organs. We achieved a complete tumor resection with negative margins in all cases. All patients have survived with no tumor recurrence to date at 17-, 27- and 38-month follow-up. Postoperative complications included diarrhea, sphincter of Oddi dysfunction and arterial stenosis; all responded to directed treatments. Multivisceral ex vivo surgery applying techniques of deceased donor multivisceral transplantation is feasible in achieving local control of otherwise unresectable abdominal tumors. This surgery is best suitable for locally invasive tumors unresectable because of location and vascular involvement.

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.

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.

Predisposing factors of diminished survival in simultaneous liver/kidney transplantation.

Since the adoption of the Model for End-Stage Liver Disease, simultaneous liver/kidney transplants (SLKT) have substantially increased. Recently, unfavorable outcomes have been reported yet contributing factors remain unclear. We retrospectively reviewed 74 consecutive adult SLKT performed at our center from 2000 to 2010 and compared with kidney transplant alone (KTA, N = 544). In SLKT, patient and death-censored kidney graft survival rates were 64 ± 6% and 81 ± 5% at 5 years, respectively (median follow-up, 47 months). Multivariable analyses revealed three independent risk factors affecting patient survival: hepatitis C virus positive (HCV+, hazard ratio [HR] 2.9, 95% confidence interval [CI] 1.1-7.9), panel reactive antibody (PRA) > 20% (HR 2.8, 95% CI 1.1-7.2) and female donor gender (HR 2.9, 95% CI 1.1-7.9). For death-censored kidney graft survival, delayed graft function was the strongest negative predictor (HR 8.3, 95% CI 2.5-27.9), followed by HCV+ and PRA > 20%. The adjusted risk of death-censored kidney graft loss in HCV+ SLKT patients was 5.8 (95% CI 1.6-21.6) compared with HCV+ KTA (p = 0.008). Recurrent HCV within 1 year after SLKT correlated with early kidney graft failure (p = 0.004). Careful donor/recipient selection and innovative approaches for HCV+ SLKT patients are critical to further improve long-term outcomes.

Uterine transplantation research: laboratory protocols for clinical application.

The aim of this review is to summarize the state-of the-art methods that are used in clinical organ transplantation today, as well as the major findings of recent experimental uterus transplantation (UTx) research regarding organ donation/retrieval, ischemic preservation, surgical techniques for anastomosis, immunosuppression and pregnancy. Absolute uterine factor infertility lacks treatment despite the major developments in infertility treatment and assisted reproduction. Concerning uterine factor infertile patients, genetic motherhood is only possible through gestational surrogacy. The latter can pose medical, ethical and legal concerns such as lack of control of life habits during surrogate pregnancy, economic motives for women to become surrogate mothers, medical/psychological pregnancy-related risks of the surrogate mother and uncertainties regarding the mother definition. Thus, surrogacy is non-approved in large parts of the world. Recent advances in the field of solid organ transplantation and experimental UTx provide a favourable and safe background in a scenario in which a human clinical UTx trial can take place. Protocols based on animal research over the last decade are described with a view to providing a scientifically guided approach to human UTx as an experimental procedure in the future.

Chronic kidney disease after orthotopic liver transplantation in recipients receiving tacrolimus.

INTRODUCTION: Chronic kidney disease (CKD) is common in liver transplant recipients receiving calcineurin inhibitors. METHOD AND POPULATION: The goals of this case-control study were to identify risk factors associated with CKD and its effect on mortality in 294 liver transplant recipients receiving calcineurin inhibition with tacrolimus. RESULTS: Hepatitis C virus (HCV) was the most common indication (42%) for transplantation. CKD 4 and 5 (estimated glomerular filtration rate (eGFR) of <=29 ml/min/1.73 m2) developed in 10.8% of recipients during a mean follow-up of 52 months. The incidence density of CKD was 2.56 per 100 patient-years. End-stage renal disease developed in 2.7%. By univariate analysis, CKD patients were older (mean±sd, 57±10 vs. 51±11, p<0.05) with hypertension (56 vs. 32%, p<0.05), had lower preoperative hematocrit (31±6 vs. 34±5, p<0.05), alanine aminotransferase (median (95% confidence limit) 46 (34­80) vs. 68 (56­77), <0.05) and eGFR (56±28 vs. 91±35 ml/ min/1.73 m2, p<0.05), had higher preoperative prothrombin time (16.1 (14.6­17.2) vs. 14.8 (14.5­15.1) seconds, p<0.05), and required more perioperative renal replacement therapy (RRT) (41% vs. 6.5%, p<0.05) compared to controls. Perioperative need for RRT (hazard ratio (95% CI) 2.72 (1.05­7.03)) and lower preoperative eGFR: 60­89 (4.08 (1.23­13.5)), 30­59 (4.26 (1.18­15.36)), and<=29 (5.91 ((1.28­27.19)) vs. eGFR>=90 ml/min/1.73 m2 were independently associated with development of CKD adjusting for important covariates. The development of CKD (2.36 (1.22­4.59)) was independently associated with late mortality with an attributable risk of 12.8%. CONCLUSION: Data demonstrate that CKD is an important clinical event associated with increased risk for death after primary liver transplantation.

Hyperinfection strongyloidiasis in a liver transplant recipient treated with parenteral ivermectin.

Severe strongyloidiasis, including hyperinfection and dissemination, is a recognized complication of solid organ transplantation. However, the development of strongyloidiasis in a liver transplant recipient has not been previously described. We present a case of severe strongyloidiasis occurring in a patient 4 months after liver transplantation and 1 month after receiving treatment for acute rejection. We assess the management challenges in this patient who remained symptomatic despite oral treatment with ivermectin and albendazole and eventual successful treatment with parenteral ivermectin. We review the published experience with alternative methods of ivermectin administration. We also investigate the possible source of infection, as the patient was not from an endemic area.

Mycobacterium abscessus infection in solid organ transplant recipients: report of three cases and review of the literature.

Mycobacterium abscessus is an ubiquitous organism found in the environment. This rapidly growing mycobacterium infrequently causes disease in humans; however, in immunocompromised hosts, disease can range from localized cutaneous lesions to disseminated infection. The organism is resistant to most antimycobacterial drugs and therapy can be limited by drug interactions. The exact incidence of M. abscessus infection among solid organ transplant (SOT) recipients is unknown; data are only available from previously reported cases in the literature. We describe 3 cases of M. abscessus infection in SOT recipients diagnosed within a 5-month period. One of the cases followed multi-visceral transplantation, the first such case to be reported in the literature. An epidemiological investigation did not reveal significant commonalities among the cases, and pulsed-field gel electrophoresis of genomic DNA of the case isolates confirmed their non-identity. All cases improved with antibiotic therapy, most notably with the new glycylcycline, tigecycline, along with surgical intervention in 2 of the cases. In addition, we review features and characteristics of M. abscessus infections in recipients of SOT reported in the literature from 1992 to 2008 and summarize some selected therapeutic concerns and issues related to treatment.

Surgical considerations in liver transplantation: small for size syndrome.

The chronic shortage of cadaveric grafts for patients on the liver transplant list has resulted in wide implementation of living donor liver transplant (LDLT) and split cadaveric liver transplantation (SLT). Small for size syndrome (SFSS) is a significant complication that can occur during LDLT or SLT. It is generally defined as the presence of prolonged cholestasis, coagulopathy and ascites within the first week from transplant. Multiple factors contribute to the pathogenesis of SFSS, such as overall graft size, portal hyperperfusion, impaired venous outflow, as well as donor and recipient factors. Strategies utilized to minimize or resolve SFSS include the use of right lobe grafts, modulation of portal flow by splenic artery ligation, splenectomy or porto-systemic bypass, and optimization of venous outflow. Additional surgical techniques to avoid SFSS include the use of auxiliary orthotopic liver grafts and dual liver graft transplantation. Careful consideration of risk to the LDLT donor has to be taken whenever right lobe graft is utilized, especially if the middle hepatic vein (MVH) is going to be included in the graft.

In situ quantitative immunoprofiling of regulatory T cells using laser scanning cytometry.

Laser scanning cytometry (iCys; CompuCyte, Cambridge, Mass) has recently been developed to use fluorescence-based quantitative measurements on tissue sections or other cellular preparations at a single-cell level. The purpose of this study was to develop objective, quantitative immunoprofiling of regulatory T cells (T regs) on formalin-fixed/paraffin-embedded (FFPE) biopsy samples from transplanted allografts using iCys. We sought to evaluate the usefulness of iCys to analyzes the population of CD4 (+) Foxp3 (+) T regs among CD4 (+) T-cell and the entire T-cell (the total of CD4 [+] and CD8 [+] populations in human intestinal allograft biopsy samples. Primary antibodies (Foxp3 and CD4) which had been labeled using Alexa Fluoro 488 (Foxp3 Alexa488) and 647 (CD4 Alexa647) with polymer horseradish peroxidase and catalyzed signal amplification were incubated on 1 section. On the other section, CD8 and CD4 were labeled using Alexa488 and Alexa647 using the same protocol. Data acquisition was performed using iCys. The signal intensities of Alexa488 and Alexa647 were sufficient to analyze by iCys. Distribution of the integrals of Alexa488 and Alexa647 to visualize each cell population enabled calculation of the population of T reg among CD4 (+) T cells, CD4 (+) T cells among total T cells, and T reg among entire T cells. iCys and signal amplified immunofluorescent staining allowed objective quantitative immunoprofiling of in situ T reg populations, with precise quantitative analysis at a single-cell level on FFPE sections. This objective method may be applied on biopsy samples from various transplanted organs.

Abdominal wall transplantation: surgical and immunologic aspects.

Abdominal wall transplantation is a type of composite tissue allograft that can be utilized to reconstitute the abdominal domain of patients undergoing intestinal transplantation. We have presented herein combined experience and long-term follow-up results of a series of abdominal wall transplants performed at 2 institutions. A total of 15 abdominal wall transplants from cadaveric donors were performed in 14 patients at the end of intestinal transplant surgery or, in 2 cases, a few days after the primary intestinal transplant. The vascular supply was through the inferior epigastric vessels, from the iliac vessels in 12 cases and via a microsurgical technique in 3 cases. Immunosuppression consisted of induction with alemtuzumab and maintenance treatment with tacrolimus monotherapy. Two grafts lost to vascular thrombosis were removed. Five patients are still alive, although all deaths were unrelated to the abdominal wall transplant. There were 3 episodes of abdominal wall graft rejection, treated with steroids; the abdominal wall graft and the intestinal grafts experienced rejection independent from each other. In summary, abdominal wall transplantation is a feasible technique for recipients of intestinal or multivisceral transplants, when the closure of the abdominal cavity by primary intention is technically impossible.

Partial bladder transplantation with en bloc kidney transplant--the first case report of a 'bladder patch technique' in a human.

Transplantation of the urinary bladder has not been reported in humans. We transplanted a portion of the donor bladder with an en bloc kidney graft in a 12-month-old girl. The child had a congenital hypoplastic single kidney with an ectopic ureteral opening into the vagina. Her native bladder was extremely small. Bilateral kidneys were transplanted en bloc with their ureters connected to a patch of the donor bladder, which encompassed the bilateral ureterovesical junctions (UVJs) (bladder patch technique). Approximately one-third of the donor bladder wall was used. The bladder patch reperfused well via blood supply from the ureters. Posttransplant cystoscopy with retrograde cystogram revealed a viable transplanted bladder with normal emptying of transplanted ureters. No reflux across the donor UVJs was seen in a voiding cystourethrogram. The child is doing well with normal renal function at 18-month follow-up.

Hepatic gas gangrene following orthotopic liver transplantation: three cases treated with re-transplantation and a review of the literature.

Gas gangrene is a rare and devastating infectious process that can occur after liver transplantation, most often following hepatic artery thrombosis. We here report 3 cases of gas gangrene following orthotopic liver transplantation. Blood cultures were positive for Clostridium clostridiiforme in one case. In 2 other cases liver tissue from explanted specimens was positive for Enterobacter cloacae. Ultrasound demonstrated hepatic artery thrombosis and computed tomography imaging revealed diffuse liver necrosis with gas formation in each case. All 3 patients were successfully treated with a combination of antibiotics and emergent re-transplantation. We review previously published cases of gas gangrene after liver transplant and emphasize the importance of hepatic artery thrombosis in the development of this syndrome as well as the frequent involvement of non-clostridial organisms. Early diagnosis and aggressive combined medical and surgical treatment including re-transplantation are essential for successful treatment of these rare and catastrophic infections.

Effect of pretransplant hepatitis C virus RNA status on posttransplant outcome.

Undetectable hepatitis C virus (HCV) RNA [RNA(-)] before liver transplantation (OLT) has been shown to decrease the rates of disease recurrence. We sought to determine whether RNA(-) subjects differ in post-OLT recurrence (virological/VR, histological/HR), graft failure (GF), or patient survival from RNA(+) patients using a retrospective review. From 1995 to 2004, a total of 49 patients were RNA(-) at OLT as a result of interferon-based therapy: 22 SVR and 27 with end-of-treatment response (ETR) transplanted when RNA(-) within 6 months of ET. Forty-eight RNA(+) patients were analyzed as controls. Virological recurrence (VR) was seen in 55% of RNA(-) subjects with no difference in HR between RNA(-) vs (+) groups, namely 36.7% versus 56.3% (P = .068), respectively. The RNA(+) subjects showed a lower time to HR (5.6 vs 11 months; P = .027). The SVR subjects displayed lower VR (36.4%) and histological recurrence (HR) (13.6%) compared to ETR (VR 70.4%, P = .023; HR 55.6%, P = .003) or RNA(+) (HR 56%, P = .0008). The SVR subjects, who were identified with a sensitive assay (SVR(S), lower limit <600 IU/mL) showed no VR, HR, or GF. The 1- and 5-year survivals were 87.8%/75.6% and 89.6%/77.8% for RNA(-) and (+) groups, respectively (P = .77). In conclusion, RNA(-)-transplanted patients displayed lower VR and longer time to HR. The SVR patients showed lower VR and HR compared to ETR and RNA(+) patients.

The effects of chimeric cells following donor bone marrow infusions as detected by PCR-flow assays in kidney transplant recipients.

40 recipients of first cadaver kidney transplants were given perioperative donor vertebral bone marrow infusions (DBMC), compared with 100 controls who did not receive donor bone marrow. The immunosuppressive regimen included OKT3, Tacrolimus, and steroid maintenance therapy, and, in some patients, newly introduced mycophenolate mofetil. This report describes the 24-mo actuarial follow-up and several immunological monitoring studies including sequential measurements of donor bone marrow lineage subset chimerism by the recently reported PCR-flow assay. This is a sensitive in situ PCR detection system for donor versus recipient histocompatibility genes as well as cell surface CD epitope markers using flow cytometry. The results indicate (a) the stabilization of the donor CD3+ and CD34+ cells in recipient peripheral blood at levels below 1% between 6 mo and 1 yr postoperatively, with a 10-fold higher level of donor cell chimerism of these lineages in recipient iliac crest marrow; (b) significantly lower levels of chimerism in peripheral blood up to 6 mo postoperatively in patients who had early acute (reversible) rejection episodes compared with those who did not; (c) a higher degree of chimerism seen in patients who were class II MHC HLA DR identical with their donors; (d) the identification of a high proportion of the donor bone marrow derived CD3 dimly staining subset of T cells (to which regulatory functions have been ascribed) in recipient peripheral blood and especially in recipient bone marrow; and (e) an unexpectedly increased susceptibility to clinically significant infections (primarily viral), and even death in the DBMC-infused group, compared with controls, but no graft losses because of rejection in the DBMC-infused group. Mixed lymphocyte culture assays showed a trend toward a greater number of nonspecifically low reactors in the DBMC group, as well as a greater number of nonspecifically high reactors in the controls (P = 0.058). The autologous mixed lymphocyte reaction also indicated a trend towards nonspecific immune activation in the DBMC group. Finally, anti-cytomegaloviral IgG antibody reactivity was significantly inhibited in the DBMC group 4-6 mo postoperatively (P = < 0.05). In the controls, there were no donor cell lineages detected by PCR-flow in the peripheral blood. These rather unexpected findings, indicating a more depressed cellular and humoral immune capacity in the DBMC cadaver kidney transplant recipients in this relatively early follow-up period, are discussed relevant to chimerism, MHC restriction, and suppressor activity brought about by specialized DBMC subsets, which still need to be defined.