Impact of hypothermic preservation on tissue yield and viability in pig pancreata.

INTRODUCTION: Chronic shortage of quality human cadaveric pancreata limits islet transplantation. Porcine islet xenotransplantation is being explored to increase the donor pool. For clinical-ready islets, centralized animal husbandry, Current Good Manufacturing Practice-regulated processing facilities, and organ transportation support are required. Amount of cold ischemia time (CIT) before isolation significantly affects transplantation. The goal of this study was to determine the maximum safe CIT of whole pancreata before islet isolation. MATERIALS AND METHODS: Pancreata were rapidly removed from Yorkshire pigs (age, 14-22 days) and stored in modified University of Wisconsin solution or in EuroCollins solution at 4(°)C. Pancreata were processed with <1 hour CIT (control) or stored for 4 or 12 hours before isolation. Islet yield and percent purity and viability were determined after 7 days of in vitro tissue culture and maturation. Samples from nonprocessed pancreata were collected and snap-frozen in liquid nitrogen at 0, 3, 6, 9, 12, 15, and 24 hours of preservation, then analyzed for adenosine diphosphate/adenosine triphosphate ratio as a measure of tissue energetics. RESULTS: Up to 12 hours in cold storage had no significant impact on overall islet yield after 7 days of in vitro culture compared with controls; islet yield at the end of the maturation process was 28,700 ± 500 islet equivalents per pancreas (mean ± SEM control yield, 30,300 ± 900 islet equivalents per pancreas); islet purity was 75 ± 5% compared with 74 ± 5% in controls. Islet viability was significantly reduced at 12 hours compared with controls (80 ± 6% vs 96 ± 5%; P < .05). The tissue adenosine diphosphate/adenosine triphosphate ratio was maintained within the first 6 hours (1.6 ± 0.1 to 1.8 ± 0.2; P = NS) but was markedly increased during the 24-hour study (3.3 ± 0.1 at 24 hours), indicating a progressive loss of adenosine triphosphate tissue stores. CONCLUSIONS: Young pig pancreata can be hypothermically stored for up to 12 hours without affecting islet yield and purity; however, islet viability is reduced. These data highlight the need for uniform shipping parameters to standardize islet quality, ideally with CIT <6 hours.

The impact of meeting donor management goals on the development of delayed graft function in kidney transplant recipients.

Many organ procurement organizations (OPOs) utilize preset critical care endpoints as donor management goals (DMGs) in order to standardize care and improve outcomes. The objective of this study was to determine the impact of meeting DMGs on delayed graft function (DGF) in renal transplant recipients. All eight OPOs of the United Network for Organ Sharing Region 5 prospectively implemented nine DMGs in every donor after neurologic determination of death (DNDD). "DMGs met" was defined a priori as achieving any seven of the nine DMGs and this was recorded at the time of consent for donation to reflect donor hospital ICU management, 12-18 h later, and prior to organ recovery. Multivariable analyses were performed to identify independent predictors of DGF (dialysis in the first week after transplantation) with a p<0.05. A total of 722 transplanted kidneys from 492 DNDDs were included. A total of 28% developed DGF. DMGs were met at consent in 14%, 12-18 h in 32% and prior to recovery in 38%. DGF was less common when DMGs were met at consent (17% vs. 30%, p=0.007). Independent predictors of DGF were age, Cr and cold ischemia time, while meeting DMGs at consent was significantly protective. The management of potential organ donors prior to consent affects outcomes and should remain a priority in the intensive care unit.

Induction therapy by anti-thymocyte globulin (rabbit) versus basiliximab in deceased donor renal transplants and the effect on delayed graft function and outcomes.

The use of induction therapy significantly reduces the incidence of acute rejection (AR) episodes posttransplantation and prevents delayed graft function (DGF). In our program, all adult deceased donor kidney transplant (DDKT) recipients receive immunosuppression induction therapy with either basiliximab (anti-CD25 Ab) or rabbit anti-thymocyte globulin (RATG). Our protocol is risk adjusted such that patients who are at a higher risk for DGF or AR received RATG and all other patients receive anti-CD25 Ab. We hypothesized that treating our higher-risk patients with RATG induction at the time of transplantation would lead to a lower rate of DGF and better outcomes. From August 1, 2005 through August 31, 2010, 116 consecutive adult patients received a DDKT in a single academic transplantation center. All DDKT patients received induction with RATG or anti-CD25 Ab. The induction decision was made prior to transplantation based on donor and recipient risk factors for AR and DGF. Transplants that were deemed at higher risk for DGF or AR based on donor factors or recipient factors received RATG. Medical records and patient databases were reviewed retrospectively. The use of RATG in higher-risk recipients for DGF and AR did not significantly reduce the DGF rate. At 6 months the function of the allograft function measured as creatinine clearance or serum creatinine was lower in the RATG group than the patients who received anti-CD25 Ab induction. The choice of induction therapy did not improve outcomes in high-risk patients in this short-term study.

Academic careers and lifestyle characteristics of 171 transplant surgeons in the ASTS.

This manuscript reports the demographics, education and training, professional activities and lifestyle characteristics of 171 members of the American Society of Transplant Surgeons (ASTS). ASTS members were sent a comprehensive survey by electronic mail. There were 171 respondents who were 49 ± 8 years of age and predominantly Caucasian males. Female transplant surgeons comprised 10% of respondents. ASTS respondents underwent 15.6 ± 1.0 years of education and training (including college, medical school, residency and transplantation fellowship) and had practiced for 14.7 ± 9.2 years. Clinical practice included kidney, pancreas and liver organ transplantation, living donor surgery, organ procurement, vascular access procedures and general surgery. Transplant surgeons also devote a significant amount of time to nonsurgical patient care, research, education and administration. Transplant surgeons, both male and female, reported working approximately 70 h/week and a median of 195 operative cases per year. The anticipated retirement age for men was 64.6 ± 8.6 and for women was 62.2 ± 4.2 years. This is the largest study to date assessing professional and lifestyle characteristics of abdominal transplant surgeons.

FTY720 and everolimus in de novo renal transplant patients at risk for delayed graft function: results of an exploratory one-yr multicenter study.

This exploratory, multicenter, open-label study evaluated the efficacy and safety of FTY720, as a part of an immunosuppressive regimen, in combination with everolimus and steroids in de novo renal transplant recipients at increased risk of delayed graft function (DGF). Patients received FTY720 (5 mg) and everolimus (4 mg) 2-12 h pre-transplantation, followed by 2.5 mg/d FTY720 and concentration-controlled everolimus (4-8 ng/mL) post-transplant for 12 months. Induction therapy was prohibited. After enrollment of 56 of the planned 200 patients between 2000 and 2002, the recruitment was terminated. The primary endpoint, rate of graft loss, or death at three months was 15.4% and the biopsy-confirmed acute rejection was 42.3%. Death or graft loss at 12 months in the DGF and non-DGF arms was 36.0% and 25.9%, respectively. The mean estimated creatinine clearance at three months was 63 and 55 mL/min in the non-DGF and DGF groups, respectively, while at 12 months it was 56 mL/min in both the groups. Although there was no comparator arm, the results from this exploratory study (compared with data from other phases II and III trials) indicated no apparent benefits of FTY720-based regimens for prevention of acute rejection and preservation of renal function in renal transplant recipients at high risk of DGF.

Simultaneous heart and kidney transplantation in patients with end-stage heart and renal failure.

Combined simultaneous organ transplantation has become more common as selection criteria for transplantation have broadened. Broadening selection criteria is secondary to improved immunosuppression and surgical techniques. The kidney is the most common extrathoracic organ to be simultaneously transplanted with the heart. A series of 13 patients suffering from both end-stage heart and renal failure underwent 14 simultaneous heart and kidney transplantations at Temple University Hospital between 1990 and 1999. This is the largest series reported from a single center. Three patients died during the initial hospitalization for an in-hospital mortality of 21%. Of 10 patients who left the hospital, 1-year survival was 100% and 2-year survival 75%. One patient required retransplant for rejection within the first year. Overall mortality at 1 and 2 years was 25 and 41%, respectively. Four out of nine (44%) patients greater than 5 years post-transplant were alive. Of the 10 patients who left the hospital, 66% were alive at 5 years. One patient succumbed to primary nonfunction of the cardiac allograft, while the four other deaths were secondary to bacterial or fungal sepsis. The patient's racial backgrounds were equally divided between African-American and white. These results are similar to those reported in a United Network of Organ Sharing Database (UNOS) registry analysis of 84 simultaneous heart and kidney transplants that found 1- and 2-year survival to be 76 and 67%, respectively. Simultaneous heart and kidney transplantation continues to be a viable option for patients suffering from failure of these two organ systems, although the results do not match those of heart transplant alone.

Structures of mammalian cytosolic quinone reductases.

The metabolism of quinone compounds presents one source of oxidative stress in mammals, as many pathways proceed by mechanisms that generate reactive oxygen species as by-products. One defense against quinone toxicity is the enzyme NAD(P)H:quinone oxidoreductase type 1 (QR1), which metabolizes quinones by a two-electron reduction mechanism, thus averting production of radicals. QR1 is expressed in the cytoplasm of many tissues, and is highly inducible. A closely related homologue, quinone reductase type 2 (QR2), has been identified in several mammalian species. QR2 is also capable of reducing quinones to hydroquinones, but unlike QR1, cannot use NAD(P)H. X-ray crystallographic studies of QR1 and QR2 illustrate that despite their different biochemical properties, these enzymes have very similar three-dimensional structures. In particular, conserved features of the active sites point to the close relationship between these two enzymes.

Simultaneous pancreas-kidney (SPK) and pancreas living-donor kidney (SPLK) transplantation at the University of Maryland.

The evolution of enteric and portal venous drainage, better immunosuppression, and better patient care has elevated pancreas transplantation with dramatically improved results. At our center, long-term graft survival and rejection has significantly improved with portal venous drainage, which has become our gold standard. This improvement is exemplified by the excellent one-year patient and graft survival rates for SPLK transplants. SPLK has proven to be an ideal approach in uremic Type 1 diabetic patients with living donors and should become the procedure of choice for that population. Moreover, the improved monitoring of rejection has allowed a similar success of pancreas transplantation alone in non-uremic patients with brittle diabetes. The treatment of diabetes mellitus has room for great improvement, however, and there is no question that islet transplantation, xenotransplantation, and the pursuit of immunologic tolerance will play an extremely important role in that endeavor.

Crystal structure of human quinone reductase type 2, a metalloflavoprotein.

In mammals, two separate but homologous cytosolic quinone reductases have been identified: NAD(P)H:quinone oxidoreductase type 1 (QR1) (EC 1.6.99.2) and quinone reductase type 2 (QR2). Although QR1 and QR2 are nearly 50% identical in protein sequence, they display markedly different catalytic properties and substrate specificities. We report here two crystal structures of QR2: in its native form and bound to menadione (vitamin K(3)), a physiological substrate. Phases were obtained by molecular replacement, using our previously determined rat QR1 structure as the search model. QR2 shares the overall fold of the major catalytic domain of QR1, but lacks the smaller C-terminal domain. The FAD binding sites of QR1 and QR2 are very similar, but their hydride donor binding sites are considerably different. Unexpectedly, we found that QR2 contains a specific metal binding site, which is not present in QR1. Two histidine nitrogens, one cysteine thiol, and a main chain carbonyl group are involved in metal coordination. The metal binding site is solvent-accessible, and is separated from the FAD cofactor by a distance of about 13 A.

General management of gastrointestinal fistulas. Recognition, stabilization, and correction of fluid and electrolyte imbalances.

Gastrointestinal fistulas are unfortunate complications of a number of disease states, such as inflammatory bowel disease and tumors, or may result from complications of surgical intervention. Fistulas may be associated with significant morbidity and mortality, much of which is a result of fluid losses and electrolyte imbalances. Thus, attention to these issues is a critical component of the management of patients with gastrointestinal fistulas. The management of gastrointestinal fistulas is divided into three phases: diagnosis/recognition, stabilization/investigation, and treatment. The major goal of the stabilization phase is the correction of fluid losses and electrolyte abnormalities. This phase must be carried out expeditiously to reduce the associated complications. Knowledge of the electrolyte content of various secretions of the gastrointestinal tract is essential to guide this phase of management. Early control of infectious foci, with drainage of abscesses if present, is of great importance. Esophageal fistulas most commonly result from instrumentation of the esophagus and are diagnosed by radiographic imaging studies. Nonoperative therapy is an option in select patients, but aggressive surgical intervention is often required. Dehydration is often associated with these injuries and must be corrected. Gastric and duodenal fistulas are most commonly iatrogenic and may be associated with significant fluid losses. Careful measurement of the fistula effluent is important. Nutritional support is begun following correction of fluid and electrolyte abnormalities. Pancreatic fistulas are often high volume fistulas and are associated with significant skin breakdown if they are cutaneous. The use of a somatostatin analogue may decrease the volume of the fistula to allow healing. Small intestinal fistulas often result from postoperative complications and require careful attention to electrolyte abnormalities. Spontaneous closure often obviates surgical intervention. Colonic fistulas are less often associated with complications than are other fistulas of the gastrointestinal tract. The stabilization phase in the management of patients with gastrointestinal fistulas is a critical time during which careful attention to fluid and electrolyte losses can result in reduced morbidity and mortality from these difficult management problems.

Hyperthermia increases intercellular adhesion molecule-1 expression and lymphocyte adhesion to endothelial cells.

BACKGROUND: We observed that the synergistic combination of immunotherapy and whole-body hyperthermia is active against large well-vascularized tumors but not microscopic tumors, and we therefore hypothesized that hyperthermia may act on lymphocyte-endothelial cell interactions. We undertook these studies to evaluate the effect of hyperthermia on lymphocyte-endothelial cell adhesion. METHODS: Cultured human umbilical vein endothelial cells (HUVEC) and normal peripheral blood lymphocytes were used. HUVEC were cultured to confluence. Treatment groups included control, hyperthermia alone (41 degrees C for 2 hours), interferon-gamma (IFN-gamma) alone, or hyperthermia + interferon-gamma. 51Cr-labeled peripheral blood lymphocytes were allowed to adhere to treated HUVEC, and nonadhering cells were washed away. Adherent cells were lysed and counted in a gamma-counter, calculating an adhesion index compared to controls. The experiment was then conducted with the addition of anti-intercellular adhesion molecule (ICAM) antibody. Cell surface ICAM expression was determined with double monoclonal antibody staining and fluorescence-activated cell sorter analysis, and soluble ICAM secretion was determined with enzyme-linked immunosorbent assay in each group. RESULTS: In a representative experiment, interferon-gamma increased adhesion by a factor of 1.81 (p < 0.05) compared with control and hyperthermia by 1.38 (p < 0.05) and combined treatment by a factor of 2.43 (p < 0.05). Anti-ICAM antibody abrogated the increased adhesion caused by hyperthermia but did not abrogate the effect of interferon-gamma. Although only 26% of control cells expressed ICAM-1 on the cell surface, interferon-gamma increased expression to 53% (p < 0.05), hyperthermia increased expression to 38% (p < 0.05), and combined treatment increased expression to 61% (p < 0.05). Soluble ICAM-1 was not increased 12 hours after treatment, but by 24 hours significant (p < 0.05) differences (control 0.262 ng/ml, IFN alone 1.50, hyperthermia alone 1.57, and combined 2.71) were noted. CONCLUSIONS: These results suggest that hyperthermia has a significant effect on lymphocyte adhesion to endothelial cells, at least in part mediated by ICAM-1. Cell surface ICAM-1 is increased at 12 hours, and soluble ICAM-1 is increased at 24 hours. These data suggest that hyperthermia may function by increasing lymphocyte adhesion, providing another locus of action to improve clinical results with immunotherapy.

Anthelmintic resistance in Australian sheep nematode populations.

The resistance status of gastro-intestinal nematodes to anthelmintics was evaluated on 881 sheep farms throughout Australia during 1991-92. Resistance was shown to be widespread. Overall, 85% of farms had sheep infected with nematodes resistant to benzimidazole, 65% to levamisole and 34% to combination (benzimidazole+levamisole) products. Resistance to ivermectin was not detected. On only 9% of farms did all anthelmintic groups reduce egg counts by greater than or equal to 95%. The culture of faeces from untreated sheep showed Telodorsagia circumcincta, Trichostrongylus spp, Chabertia ovina and Haemonchus contortus to be the principal species. The nature and prevalence of resistance was not significantly correlated with stocking rate. However, resistance to combination products was almost twice as prevalent on farms in areas with an average annual rainfall of greater than 500 mm.

Thallium scintigraphy in the evaluation of mass abnormalities of the breast.

Palpable mass abnormalities of the breast are often difficult to evaluate mammographically, especially in patients with fibrocystic change and dense breasts. The current study evaluates 201TI scintigraphy as a potential test in detecting malignancy and in differentiating malignant from benign masses. Eighty-one female patients underwent thallium scintigraphy of the breast because of palpable breast masses. An additional 30 females with no palpable breast abnormalities were also studied using 201TI. Of 44 patients with palpable breast carcinomas, 42 carcinomas (96%) were detected using 201TI scintigraphy. Three of three patients had other primary breast malignancies that were also detected. In contrast, 19 patients with palpable breast abnormalities shown on biopsy to be benign fibrocystic disease processes were not detectable on thallium studies. Of two patients with fat necrosis, none were detectable. Three of 13 patients had adenomas of the breast (23%) that were detected. The three detectable adenomas were all highly cellular. The smallest detectable carcinoma was an adenocarcinoma measuring 1.3 x 1.1 x 0.9 cm. Thallium-201 scintigraphy of palpable breast lesions is an effective test for evaluation of palpable masses. Sensitivity for detection of malignant masses greater than 1.5 cm is high. Highly cellular adenomas, however, may demonstrate significant 201TI uptake. Benign fibrocystic disease is not detectable with thallium scintigraphy. Thallium scintigraphy of breast lesions is an effective means of differentiating benign from malignant lesions.

Periampullary duodenal diverticulum in a patient undergoing laparoscopic cholecystectomy.

Duodenal diverticulae are acquired lesions usually found near the papilla of Vater and associated with significant symptoms. We present a case of a patient with obstructive jaundice who had a diverticulum and cholelithiasis. The management of this scenario is explained in the era of laparoscopic cholecystectomy.

Primary endodermal sinus (yolk sac) tumor of the falciform ligament.

Extragonadal yolk sac tumors (YSTs) are uncommon and YSTs of the liver are exceedingly rare, with only three reported cases in the literature. A case is described of primary YST of the falciform ligament extending into the left lobe of the liver in a 14-month-old boy. This is the first reported case of primary YST arising within the falciform ligament. The patient underwent an exploratory laparotomy after presenting with hemoperitoneum. An extremely friable and necrotic tumor was found extending from the falciform ligament into the liver. The tumor was debulked and the patient received 5 months of chemotherapy employing a modified Einhorn regimen. After a partial response to chemotherapy the patient had a second-look laparotomy, at which time a left hepatic lobectomy and en bloc resection of the falciform ligament was performed in order to remove residual tumor. At the present time the patient has no signs of metastases and is alive and well 2 years after his presentation.

Long-term follow-up of brachial artery ligation in children.

Ligation of the brachial artery in the antecubital fossa in children can be performed without limb loss, but the effect on subsequent limb growth is unknown. From 1969 to 1974, brachial artery ligation for insertion of a Scribner shunt was performed in 27 patients with a mean age of 8.4 years (range, 3 to 15 years). We examined 11 patients, all with functional renal transplants, a mean of 15.8 years (range, 13 to 18 years) after ligation. Nine patients had unilateral ligation and two had bilateral ligation. The patients were examined for arm length and circumference, resting blood pressure at the wrist, neurologic function of the arm, and exercise tolerance. A significant difference in resting arterial pressures in the ligated extremity was uniformly noted (mean systolic pressure 106 mmHg versus 123 mmHg in 7 patients, p less than 0.01). Although no patient specifically complained of problems with the ligated side, six of nine patients with unilateral ligation experienced arm discomfort on stress testing. There was no significant difference in limb circumference or length between the ligated and unligated extremity. Ligation of the brachial artery in growing children with renal disease caused diminished resting pressure and mildly decreased exercise tolerance but did not lead to limb loss or subsequent growth abnormalities.