Longitudinal studies of a B cell-derived signature of tolerance in renal transplant recipients.

Biomarkers of transplant tolerance would enhance the safety and feasibility of clinical tolerance trials and potentially facilitate management of patients receiving immunosuppression. To this end, we examined blood from spontaneously tolerant renal transplant recipients and patients enrolled in two interventional tolerance trials using flow cytometry and gene expression profiling. Using a previously reported tolerant cohort as well as newly identified tolerant patients, we confirmed our previous finding that tolerance was associated with increased expression of B cell-associated genes relative to immunosuppressed patients. This was not accounted for merely by an increase in total B cell numbers, but was associated with the increased frequencies of transitional and naïve B cells. Moreover, serial measurements of gene expression demonstrated that this pattern persisted over several years, although patients receiving immunosuppression also displayed an increase in the two most dominant tolerance-related B cell genes, IGKV1D-13 and IGLL-1, over time. Importantly, patients rendered tolerant via induction of transient mixed chimerism, and those weaned to minimal immunosuppression, showed similar increases in IGKV1D-13 as did spontaneously tolerant individuals. Collectively, these findings support the notion that alterations in B cells may be a common theme for tolerant kidney transplant recipients, and that it is a useful monitoring tool in prospective trials.

Immunosuppression with belatacept-based, corticosteroid-avoiding regimens in de novo kidney transplant recipients.

Current immunosuppressive regimens in renal transplantation typically include calcineurin inhibitors (CNIs) and corticosteroids, both of which have toxicities that can impair recipient and allograft health. This 1-year, randomized, controlled, open-label, exploratory study assessed two belatacept-based regimens compared to a tacrolimus (TAC)-based, steroid-avoiding regimen. Recipients of living and deceased donor renal allografts were randomized 1:1:1 to receive belatacept-mycophenolate mofetil (MMF), belatacept-sirolimus (SRL), or TAC-MMF. All patients received induction with 4 doses of Thymoglobulin (6 mg/kg maximum) and an associated short course of corticosteroids. Eighty-nine patients were randomized and transplanted. Acute rejection occurred in 4, 1 and 1 patient in the belatacept-MMF, belatacept-SRL and TAC-MMF groups, respectively, by Month 6; most acute rejection occurred in the first 3 months. More than two-thirds of patients in the belatacept groups remained on CNI- and steroid-free regimens at 12 months and the calculated glomerular filtration rate was 8-10 mL/min higher with either belatacept regimen than with TAC-MMF. Overall safety was comparable between groups. In conclusion, primary immunosuppression with belatacept may enable the simultaneous avoidance of both CNIs and corticosteroids in recipients of living and deceased standard criteria donor kidneys, with acceptable rates of acute rejection and improved renal function relative to a TAC-based regimen.

Isolation and partial characterization of the pancreatic secretory trypsin inhibitor in the rat.

Isolation in a 55% yield of the low molecular weight pancreatic secretory trypsin inhibitor was achieved by gel filtration of an acid extract of whole inactive rat pancreas juice on Sephadex G-50 at pH 2.5 followed by desalting and ion-exchange chromatography on SP Sephadex C-50 at pH 4.5. Two distinct chromatographic fractions were obtained, labeled fraction 1 and 2. Fractions 1 and 2 showed three, respectively two, distinct closely migrating cationic bands on gel electrophoresis in barbital buffer, pH 8.6. Each fraction demonstrated one band on polyacrylamide disc electrophoresis at pH 4.6. The inhibitor is homogenous on gel filtration and on the basis of its stoichiometry with active site titrated rat anionic trypsin. Its molecular weight is approx. 6024. The amino acid composition is included. Rat pancreatic secretory trypsin inhibitor is trypsin-specific and interacts on a 1:1 molar basis with rat trypsin. It is a good inhibitor of bovine trypsin but a poor inhibitor of human cationic trypsin and its binding to trypsin is reversible by acidification. Like other inhibitors of this sort, it is present in about 0.1-0.2% of the total protein content of the juice, and normally exists in its free form. A simple procedure for the production of antiserum to the inhibitor which is a poor antigen is also described.

Small bowel allograft rejection detected by serum intestinal fatty acid-binding protein is reversible.

We hypothesized that, following experimental small bowel transplantation, immunosuppressive therapy initiated on the day of the initial rise in serum intestinal fatty acid-binding protein (I-FABP) would result in graft salvage. In previously published work, we showed that I-FABP was not detectable in the serum of isografted Lewis rats, but could be measured in the peripheral circulation during small bowel allograft rejection. A clinically useful method to monitor trans- planted allografts for rejection should detect the problem early in its evolution so that treatment to reverse the process would salvage a functional organ. Lewis rats served as recipients of LBNF1 out-of-continuity small bowel allografts and were studied in two groups: group I (control) received no immunosuppression and group II received cyclosporine (CsA, 15 mg/kg/d, p.o.) when I-FABP rose to > or = 80 ng/ml. Serum I-FABP was measured daily until the time of sacrifice. Full-thickness graft biopsies were obtained on postoperative days 3 (baseline), 6 or 7 (elevated I-FABP), 10, and 14 (sacrifice). Following transplantation baseline serum I-FABP (day 2 or 3) averaged < or = 10.0 ng/ml. I-FABP remained at baseline through day 5 (range 0-50 ng/ml) in all animals and then rose abruptly on either day 6 or 7 (range 86-150 ng/ml; P < 0.001 vs. baseline). Histology on day 6 or 7 revealed a mild-to-moderate cellular rejection. Cyclosporine therapy reversed the rejection reaction and restored the bowel to normal histology. Serum I-FABP returned to baseline. In untreated animals, serum I-FABP remained elevated for several days and then returned to baseline levels coincident with fulminant rejection and mucosal sloughing. I-FABP was released into the peripheral circulation early in the evolution of acute rejection in this model of small bowel transplantation. Immunosuppressive therapy initiated when elevated levels of I-FABP were detected in the serum resulted in graft salvage. Cyclosporine immunotherapy consistently reversed rejection in this model. This article represents the first report of salvage of small bowel allografts when immunosuppressive therapy was instituted prospectively on the basis of a serum marker. Immunoreactive I-FABP appears to hold significant potential as a biochemical screening tool for acute rejection occurring In small bowell allografts.

A three-year experience with serum anodal trypsinogen as a biochemical marker for rejection in pancreatic allografts. False positives, tissue biopsy, comparison with other markers, and diagnostic strategies.

Serum values of immunoreactive anodal trypsinogen (sAT) have been claimed to correlate well with rejection occurring in pancreatic allografts. We have studied the behavior of sAT in serial serum samples obtained from 39 type I diabetics undergoing whole-organ pancreas transplantation during the past 3 years. Patients had either received a pancreatic allograft simultaneously with a transplanted kidney (SPK, n = 33) or after a previous kidney transplant (pancreas after kidney [PAK] n = 6). The behavior of sAT was studied in relation to the clinical diagnosis of rejection. Graft amylase output for all 39 patients and serum creatinine for the 33 SPK recipients were also studied. Tissue biopsies were obtained from 11 patients with elevated sAT values and a presumptive diagnosis of rejection. Nine of these patients had SPK grafts and simultaneously elevated creatinine values. Tissue was obtained from the simultaneously transplanted kidney; all specimens revealed rejection. Two of the 11 patients had PAK allografts. Biopsies performed on the graft duodenum were consistent with acute rejection. Three additional patients with unchanged sAT values had biopsies for other reasons; these biopsies failed to demonstrate signs of acute rejection. Thus graft biopsy correlated exactly with sAT behavior in every case in which rejection was suspected. Five patients had elevations of sAT not associated with rejection: one resulted from direct trauma, two had outlet obstruction, and two had clinical diagnoses of graft pancreatitis. The sAT was more sensitive and specific than GAO and as sensitive as creatinine for SPK recipients. These studies confirm that sAT is a reliable, graft-specific biochemical marker for the early diagnosis of pancreatic rejection. The use of sAT should allow for the proper timing of graft biopsies and the judicious use of immunosuppressive agents, which will result in increased allograft survival for PAK and pancreas-alone allografts.

Asialoglycoprotein/asialoglycoprotein receptor (AGP-AGPr) interaction is an important mechanism for the uptake of FK506 by hepatocytes.

Hepatic tissue concentrations of FK506 have been correlated with early acute rejection following liver transplantation. Asialoglycoproteins (AGP) reputedly bind FK506 in blood. AGP are removed from the circulation by the liver via the AGP receptor (AGPr), which resides on hepatocytes. This study was undertaken to determine if the AGP-AGPr mechanism enhances the delivery of FK506 to hepatocytes. Human orosomucoid (OM) was used as a representative AGP. asialoOM (aOM) was prepared by desialation of OM. Fresh rat hepatocytes were isolated by collagenase digestion. Tritium labeled FK506 (FK) was used to identify and quantitate FK506. Quantitation of FK in serum and culture media was by direct counting. FK in animal tissues used a method developed in our laboratory for the purpose. AGPr on resting hepatocytes was demonstrated by flow cytometry using FITC-orosomucoid and FITC-BSA controls. AGPr were enhanced by 2 g glucose/L. Two serum FK-binding fractions, 44 kD and 15 kD, were identified by gel filtration. Exogenous OM avidly bound FK and displaced FK activity from the 15 kD fraction. Serum (1%) and the 44 kD fraction enhanced the uptake of FK by hepatocytes, while serum depleted of OM-aOM by affinity chromatography was only 72.5% as effective as control serum; aOM enhanced the uptake of FK by hepatocytes to a degree similar to that of control serum but OM did not significantly affect the uptake of FK. Cold FK506 blocked the uptake and was dose dependent; cold CsA had no effect. Affinity extraction of OM from serum to which FK had previously been added removed 28.4% of FK activity. Following i.v. infusion, the kidney had the highest and liver the lowest tissue concentration of FK at 1 hr and 3 hr. In contrast, after oral administration the liver had the highest concentrations of the other tissues tested. The AGP-AGPr mechanism plays a significant role in the delivery of FK506 to hepatocytes and is likely responsible for the differences in bioavailability observed after oral and i.v. administration. Factors governing the AGP-AGPr mechanism are germane to understanding both the efficacy and toxicity of FK506 and the development optimal therapeutic strategies.

Serum immunoreactive anodal trypsinogen and urinary amylase as biochemical markers for rejection of clinical whole-organ pancreas allografts having exocrine drainage into the urinary bladder.

Rejection of pancreas allografts is best measured today by co-monitoring the creatinine of a simultaneously transplanted kidney allograft from the same donor. This methodology discourages pancreas transplantation for patients who have previously received a kidney allograft and preuremic patients. Thus, an early, graft-specific marker of rejection is desirable. In this study we compared 2 putative biochemical markers for rejection of pancreas allografts, serum immunoreactive anodal trypsinogen and urinary amylase, with serum creatinine in 15 simultaneously transplanted type I diabetics. Serial values during hospitalizations were determined. Follow-up ranged from 18 to 134 postoperative days. Rejection was diagnosed clinically and considered real if the patient received a course of anti-rejection medication. Ten of these 15 patients experienced a total of 21 rejection episodes. For all episodes of rejection, serum trypsinogen rose from a baseline of 398.1 +/- 25 ng/ml to 1686.2 +/- 317.9 ng/ml (P less than 0.001) on the day of rejection. Urinary amylase fell from 88,310 +/- 7877 U/24 hr to 37,508 +/- 7142 U/24 hr (P less than 0.001). For 10 patients in whom rejection was diagnosed on the initial hospitalization so that serial prediagnosis sera and urines were available, anodal trypsinogen rose from a baseline of 756 +/- 263 ng/ml to 1936 +/- 582 ng/ml (P less than 0.001). Urinary amylase values for these same 10 patients did not change significantly (baseline = 55,788 +/- 18,404 U/24 hr, rejection = 47,133 +/- 14,737 U/24 hr, (P = 0.7). We conclude that serum anodal trypsinogen behaves as a graft-specific biochemical marker for rejection of vascularized pancreas allografts.

The immunochemical distribution of cyclophilin in normal mammalian tissues.

Cyclophilin, a 17 Kd proline cis-trans isomerase, high-affinity (Kd 10(-8) M) target for the immunosuppressive drug cyclosporine has ubiquitous phylogenic distribution, but its tissue localization in mammals has not been detailed. To explore a potential relationship between the multiple systemic effects of CsA and the cellular and tissue distribution of CYP, thirty-three different normal porcine tissues were examined using an immunohistochemical technique. Tissue was obtained from farmbred pigs, immediately fixed in buffered formalin, and prepared as embedded 5-mu sections. Immune-specific staining was accomplished using an ABC immunoperoxidase method and an affinity-purified, monospecific, rabbit anti-CYP IgG. Cut sections served as their own blanks and controls, and all tissues were stained in batch to minimize the effects of variation in technique. Consistent with earlier reports, CYP was present in all tissues studied, however, there was remarkable heterogeneity in CYP distribution. Renal parenchymal cells, cardiac and striated muscle, pulmonary and skin demonstrated cytoplasmic immunospecific CYP--however, the cellular localization varied. Cytoplasmic staining of endothelial, neural, and glandular elements was consistently observed. Contrasting with previous reports, CYP localized to the nucleus as well as the cytoplasm of some lymphoid cells, hepatocytes, and cells of the large intestine. Generally, greater CYP-specific staining was noted in organs amenable to CsA immunosuppression (heart, liver, kidney), compared with organs deemed more immunologically vulnerable when allografted under CsA (pancreas, lung, small bowel). Similarly, CYP-immunospecific staining was abundant in tissues susceptible to CsA toxicities (neural tissue, smooth muscle, kidney, liver). This detailed immunohistological examination affords a correlation between CYP content and sensitivity to CsA. It also raises some new questions about tissues with little extractable CYP but significant histological staining.

The effect of cyclosporine administration on the cellular distribution and content of cyclophilin.

Cyclophilin (CYP), an intracellular protein sharing amino acid sequence identity with the enzyme peptidyl-prolyl cis-trans isomerase has become the leading candidate for the receptor responsible for cyclosporine biological effects. Avid binding of CYP to cyclosporine and immunosuppressive cyclosporine metabolites has been demonstrated, while nonimmunosuppressive cyclosporine metabolites have tended not to bind to cyclophilin. A previous immunohistochemical analysis documented that CYP localized principally to the cytoplasmic cellular compartment, but nuclear staining was observed among some cells. This study was undertaken to more precisely define the ultrastructural distribution of CYP, and to determine whether CYP cellular content was affected by CsA therapy. Untreated Wistar rats or those receiving 7 days of CsA (15 mg/kg/day, i.p.) were anesthetized, perfusion-fixed in situ, and sacrificed. Analyses of lymph node, spleen, thymus, kidney, liver, heart, brain, and ileum used an affinity purified, rabbit anticyclophilin IgG. Transmission electron microscopy was performed after staining with anti-CYP using a horseradish peroxidase/biotin/avidin technique. Quantitative immunofluorescence was measured by confocal microscopy using anti-CYP, with a biotin/avidin/phycoerythrin technique. Cyclophilin localized to the cytoplasmic compartment--however, association with mitochondria endoplasmic reticulum, Golgi, and with the nuclear membrane among lymphocytes, as well as cells from kidney, liver and ileum--was documented. Cyclophilin was not identified within the nucleus proper. Tissues obtained from animals receiving CsA exhibited a generalized increase in CYP content compared with tissues from untreated controls, suggesting the possibility that CsA may exert a regulatory influence upon CYP gene activation. Collectively, the data were consistent with the hypothesis that CYP exerts a central role in cellular metabolism, and that CsA-mediated biologic effects result from the CsA/CYP interaction.

Successfully treated invasive pulmonary aspergillosis associated with smoking marijuana in a renal transplant recipient.

Invasive pulmonary aspergillosis (IPA) is often a lethal entity in transplant recipients (up to 90%). We report the successful treatment of a case of IPA in a renal transplant recipient whose only risk for exposure was habitual marijuana smoking. Although marijuana smoking has been linked to the development of IPA in patients immunosuppressed for a variety of reasons, this case is the first report involving a solid organ transplant recipient. The patient's clinical course and treatment are described and the literature is reviewed with respect to environmental and patient risk factors. In this case, IPA was associated with the patient's heavy usage of marijuana during the immediate posttransplant period. Treatment was successful and included the experimental amphotericin product amphotericin B colloidal dispersion. Contemporaneous exposure to a large amount of inocula of Aspergillus within 30 days of receiving high doses of steroids appeared to be the most important factor that predisposed this patient to IPA. Transplant recipients should be specifically proscribed from marijuana use during periods of high steroid administration.

Treatment alternatives in renal failure and renal transplantation patients with nonobstructive colonic dilatation.

The incidence of nonobstructive colonic dilatation (NCD) is unknown, but the attendant mortality associated with perforation is nearly 50%. Patients with chronic renal failure and transplant recipients may manifest many of the conditions that have been implicated in the development of NCD. Mechanical obstruction and ischemic bowel disease must be eliminated as causes for colon dilatation. Over a four-year period eight patients (mean age 50 years) were treated for presumed NCD. Six patients with a mean cecal diameter of 12.8 cm were treated initially with colonoscopy. Five patients (83%) had successful endoscopic decompression; of the three remaining patients, one underwent urgent ileocolectomy for cecal ischemia after unsuccessful endoscopic decompression, a second (cecal diameter 13 cm) had a tube cecostomy performed as an initial procedure, and the third (cecal diameter 9 cm) was managed successfully with enemas and nasogastric suction. Two deaths occurred in the series (25%), but both were unrelated to colon distension. No complications of colonoscopy were observed. The sequelae of massive NCD (cecal ischemia, perforation, and protracted sepsis) are poorly tolerated in the immunocompromised patient. Conservative management may be employed in patients with a cecal diameter of 9 cm, but urgent diagnostic and therapeutic colonoscopy is recommended for patients with a cecal diameter of 12 cm or greater. Operative tube cecostomy may be necessary if colonoscopic decompression is unsuccessful or cannot be performed.

Association of inclusion of the donor spleen in pancreaticoduodenal transplantation with rejection.

Clinical pancreas transplantation (txp) using a pancreaticoduodenal allograft (pda) and the donor spleen has been applied as treatment for insulin-dependent diabetes mellitus. Inclusion of the spleen in the pda is purported to protect against thrombosis and may be of possible immunological benefit. In pancreatectomized pigs, we compared the results of pda txp with (n = 8) and without (n = 10) the donor spleen. Animals were maintained on cyclosporine (6 mg/kg/day i.v.) and prednisolone (1.5 mg/kg/day i.v. tapered over 7 days to a maintenance dose of 0.5 mg/kg/day i.v.). Pancreatic exocrine enzyme replacement was administered daily. In 10 recipients of the technically successful pda without the spleen, rejection (blood glucose greater than 150 mg/dl) was not seen in the 28 days after txp. Mean daily blood glucose was 95 +/- 8 mg/dl during this period. In contrast, 5 of 8 recipients of the technically successful pda with the donor spleen rejected their grafts an average of 13 days (range 7-16) after txp (P less than 0.01). Mean daily blood glucose was 88 +/- 21 mg/dl prior to the declaration of rejection and increased to 275 +/- 59 mg/dl after rejection (P less than 0.0001). Rejection was histologically confirmed in all cases. None of the recipients of the pda with the donor spleen developed signs of graft-versus-host disease. Glucose tolerance testing carried out 28 days after transplantation in normoglycemic pigs from both experimental groups demonstrated no difference between the groups. In the porcine pda model used in this study, the inclusion of the spleen in the pda was associated with rejection of the transplant.

Hepatic retransplantation in New England--a regional experience and survival model.

Hepatic retransplantation (reTx) offers the only alternative to death for patients who have failed primary hepatic transplantation (PTx). Assuming a finite number of donor organs, reTx also denies the chance of survival for some patients awaiting PTx. The impact of reTx on overall survival (i.e., the survival of all candidates for transplantation) must therefore be clarified. Between 1983 and 1991, 651 patients from the New England Organ Bank underwent liver transplantation, and 73 reTx were performed in 71 patients (11% reTx rate). The 1-year actuarial survival for reTx (48%) was significantly less than for PTx (70%, P < 0.05). This survival varied, dependent on the interval of time following PTx in which the reTx was performed (0-3 days, 57% survival; 4-30 days, 24%; 30-365 days, 54%; and > 365 days, 83%). Patients on the regional waiting list had an 18% mortality rate while awaiting transplantation. These results were incorporated into a mathematical model describing survival as a function of reTx rate, assuming a limited supply of donor livers. ReTx improves the 1-year survival rate for patients undergoing PTx but decreases overall survival (survival of all candidates) for liver transplantation. In the current era of persistently insufficient donor numbers, strategies based on minimizing the use of reTx, especially in the case of patients in whom chances of success are minimal, will result in the best overall rate of patient survival.

Lack of utility of intestinal fatty acid binding protein levels in predicting intestinal allograft rejection.

INTRODUCTION: The enterocyte-specific protein, intestinal fatty acid binding protein (I-FABP), is detectable in serum only after intestinal injury. Previous studies in animals suggest that I-FABP might be a useful marker of intestinal allograft rejection. MATERIALS AND METHODS: I-FABP was repetitively measured in nine intestinal transplant recipients and correlated with findings of surveillance endoscopy. RESULTS: Average interval between I-FABP determination and biopsy was 3.4 days (SD=4.2 days). Average number of rejection episodes per patient totalled 1.6+/-1.2. General linear modeling demonstrated no tendency for increases in serum FABP to precede histologic graft rejection (P=0.263). Restriction of the analysis to I-FABP determinations 1 day before or on the day of biopsy failed to affect these results. Minor increases in I-FABP were often associated with histologically normal grafts, whereas rejection often occurred when I-FABP was not detectable. DISCUSSION: Serum I-FABP levels do not predict clinical intestinal allograft rejection.

Biochemical detection of small intestinal allograft rejection by elevated circulating levels of serum intestinal fatty acid binding protein.

BACKGROUND: Intestinal fatty acid binding protein (I-FABP) was investigated as a serum marker for acute intestinal allograft rejection. Its behavior was compared with that of another putative marker of intestinal damage, hexosaminidase. METHODS: Transplants were performed in three groups of rats: group 1, Lewis to Lewis; group 2, ACI to Lewis, no immunosuppression; and group 3, ACI to Lewis with cyclosporine given on posttransplant days 0 through 5. Daily serum I-FABP and hexosaminidase levels were quantitated and serial graft biopsy specimens were obtained. RESULTS: Serum I-FABP levels fell to 20 ng/ml or less in all animals between posttransplant days 3 and 4. In group 1, I-FABP levels remained at baseline throughout the experiment. In group 2, I-FABP levels rose dramatically on either day 6 or 7 and declined to baseline within 4 days of the peak. On the day that I-FABP levels increased, findings of biopsy specimens were consistent with early rejection. In group 3 the rise in serum I-FABP levels was delayed 2 to 10 days. Hexosaminidase did not correlate with rejection. CONCLUSIONS: Serum I-FABP content correlated with early histologic manifestations of rejection. Hexosaminidase was insensitive as a marker in this model. I-FABP, which has a human analog, has potential as a biochemical marker for early intestinal allograft rejection.

Intestinal fatty acid binding protein in serum and urine reflects early ischemic injury to the small bowel.

BACKGROUND: Intestinal fatty acid binding protein (I-FABP) is a 15 kd protein that constitutes 2% to 3% of enterocyte protein. Normally I-FABP is undetectable in serum. The purpose of this study was to determine whether I-FABP could be detected in peripheral serum and/or urine early in the evolution of intestinal ischemic injury. METHODS: I-FABP and two putative biochemical markers of mesenteric ischemia, hexosaminidase and lactate dehydrogenase, were quantitated in the serum of rats subjected to mesenteric ischemia produced by: (1) 0.5 hours, 1 hour, or 3 hours of superior mesenteric artery (SMA) occlusion followed by reperfusion; (2) 1 hour of mesenteric occlusion to a 10 cm segment of jejunum followed by reperfusion; and (3) arterial ligation to a 10 cm segment of jejunum without reperfusion. I-FABP was also quantitated in the urine and intestinal mucosa of these animals. RESULTS: The baseline serum I-FABP level was < or = 4.0 ng/ml in all animals. In control animals, I-FABP remained unchanged throughout the experiment. In the ischemia/reperfusion groups, I-FABP immediately appeared in the serum on reperfusion. With segmental arterial ligation, I-FABP was detected in the serum within 15 minutes. Urinary content of I-FABP rose 60 minutes after its initial appearance in the serum, and its elimination paralleled serum I-FABP levels. Serum hexosaminidase and lactate dehydrogenase levels only rose after 3 hours of SMA occlusion with reperfusion. One hour of SMA occlusion and reperfusion resulted in only mild to moderate mucosal injury, whereas 3 hours of SMA ischemia with reperfusion produced areas of transmural necrosis. CONCLUSIONS: I-FABP is released into the peripheral circulation after reversible intestinal ischemic injury and has potential as a biochemical marker to facilitate the early detection of mesenteric ischemia.

Human intestinal fatty acid binding protein: report of an assay with studies in normal volunteers and intestinal ischemia.

BACKGROUND: Human intestinal fatty acid binding protein (hIFABP) is a cytoplasmic protein of mature small intestinal epithelium. Work with the rat demonstrated that serum levels of IFABP correlated with early phases of intestinal mucosal injury. The aim of this study was to develop an assay for hIFABP and assess its usefulness as a marker for intestinal mucosal injury in human beings. METHODS: Recombinant hIFABP (r-hIFABP) was used to produce rabbit anti-hIFABP. Specificity and avidity of binding were tested with immunoprecipitation and Scatchard analysis. r-hIFABP was labeled with 125I, and a competitive assay was developed. Urine and serum from normal volunteers and from patients with necrotizing enterocolitis (NEC), acute thromboembolic related intestinal ischemia, and systemic inflammatory response syndrome were tested for hIFABP. RESULTS: Molecular weight was 10(-12) kd, limit of detection was 1.87 ng/ml, and no cross-reactivity occurred when tested against rat IFABP or human heart FABP. Mean levels of hIFABP (ng/ml) were controls (serum less than 1.87, urine less than 1.87), NEC (serum 14.7 ng/ml), intestinal ischemia (serum 50 ng/ml, urine 52.3 ng/ml), systemic inflammatory response syndrome (serum 5.3 ng/ml, urine 13.2 ng/ml). CONCLUSIONS: This assay is quantitative for hIFABP in serum and urine. Results from both normal persons and those with various causes of intestinal ischemia parallel our previous findings in the rat. Preliminary findings suggest that hIFABP may serve as a diagnostic marker for early intestinal mucosal compromise and, in addition, that it should prove useful as a tool in developing rationale therapeutic regimens to treat these complex clinical problems.

Immunoreactive anionic and cationic trypsins in serum after experimental porcine pancreatic transplantation.

Immunoreactive anionic and cationic trypsins (irAT and irCT) were measured in blood samples taken daily from 20 pigs during 60 days after whole organ pancreatic transplantation. Immunosuppression was discontinued on the twenty-eighth day after transplantation. IrAT concentrations showed three distinct peaks, the first immediately after operation, the second around the seventh day after operation, and the third irAT peak was seen 2 to 9 days after discontinuation of immunosuppression therapy. IrCT levels paralleled the irAT levels only in the first peak immediately after transplantation. After this there was a dissociation between the concentrations of the two immunoreactive trypsins, with low irCT levels and high irAT levels. Increased irAT levels heralded rejection of the pancreatic allograft by 4 to 30 days (median 20 days) in 13 of the 16 rejecting animals (80%). In addition, low irCT levels preceded hyperglycemia in 13 of the 16 rejecting pigs by 2 to 47 days (median 21 days). The first postoperative peak of immunoreactive trypsins is thought to be related to the operative and storage trauma, whereas the second and third peaks are thought to reflect a rejection process. The results suggest that immunoreactive trypsins can be used as markers for pancreatic allograft rejection.

Hepatic artery aneurysm associated with the mucocutaneous lymph node syndrome.

A case of a 4-year old child with a hepatic artery aneurysm after the mucocutaneous lymph node syndrome is reported. The child had obstructive jaundice and preoperative evaluation did not lead to the correct diagnosis. The aneurysm was resected and the postoperative course was unremarkable. The literature on this entity is reviewed.

Detrimental effects of four hours of cold storage on porcine pancreaticoduodenal transplantation.

Transplantation of the pancreaticoduodenal allograft (PDA) has recently been advocated as a technique that is superior to the use of the segmental allograft. However, the effect of simple cold storage preservation on the PDA has not been studied. We investigated the effect of 24 and 4 hours of cold storage in Eurocollins solution on porcine PDA function after transplantation in pancreatectomized pigs. A regimen of cyclosporine and prednisone was used, which prevented rejection for at least 28 days after transplantation. Cold storage preservation for 24 hours uniformly resulted in PDA failure. Compared with recipients of immediately transplanted PDA, recipients of PDA cold stored for 4 hours had marked plasma hyperamylasemia (10,000 U/L versus 1932 U/L), relative glucose intolerance (K value -2.15 versus -2.66), hypoinsulinemia (peak immunoreactive insulin 11.0 microU/ml versus 34.7 microU/ml), and an abnormal pattern of insulin secretion as demonstrated with intravenous glucose tolerance testing. There was also a higher incidence of technical complications in the group transplanted with cold-stored PDAs. Our results suggest that there is a detrimental effect on porcine PDA function after only 4 hours of cold storage in Eurocollins solution.