Ileal mucosal bile acid absorption is increased in Cftr knockout mice.

BACKGROUND: Excessive loss of bile acids in stool has been reported in patients with cystic fibrosis. Some data suggest that a defect in mucosal bile acid transport may be the mechanism of bile acid malabsorption in these individuals. However, the molecular basis of this defect is unknown. This study examines the expression of the ileal bile acid transporter protein (IBAT) and rates of diffusional (sodium independent) and active (sodium dependent) uptake of the radiolabeled bile acid taurocholate in mice with targeted disruption of the cftr gene. METHODS: Wild-type, heterozygous cftr (+/-) and homozygous cftr (-/-) mice were studied. Five one-cm segments of terminal ileum were excised, everted and mounted onto thin stainless steel rods and incubated in buffer containing tracer 3H-taurocholate. Simultaneously, adjacent segments of terminal ileum were taken and processed for immunohistochemistry and Western blots using an antibody against the IBAT protein. RESULTS: In all ileal segments, taurocholate uptake rates were fourfold higher in cftr (-/-) and two-fold higher in cftr (+/-) mice compared to wild-type mice. Passive uptake was not significantly higher in cftr (-/-) mice than in controls. IBAT protein was comparably increased. Immuno-staining revealed that the greatest increases occurred in the crypts of cftr (-/-) animals. CONCLUSIONS: In the ileum, IBAT protein densities and taurocholate uptake rates are elevated in cftr (-/-) mice > cftr (+/-) > wild-type mice. These findings indicate that bile acid malabsorption in cystic fibrosis is not caused by a decrease in IBAT activity at the brush border. Alternative mechanisms are proposed, such as impaired bile acid uptake caused by the thick mucus barrier in the distal small bowel, coupled with a direct negative regulatory role for cftr in IBAT function.

Pitfalls in preparation of (3)H-unconjugated bilirubin by biosynthetic labeling from precursor (3)H-5-aminolevulinic acid in the dog.

We report problems encountered during preparation of tritium-labeled unconjugated bilirubin ((3)H-UCB) from precursor (3)H-5-aminolevulinic acid ((3)H-ALA) in 2 dogs with external biliary drainage installed into the animals under general anesthesia. Under prolonged sedation, 12.9 or 14.0 mCi of (3)H-ALA was administered intravenously in two divided doses, and bile was collected for 9 hours. In one animal, taurocholate (TC) infusion was needed to maintain bile flow. (3)H-UCB was isolated from the bile and recrystallized with the improved method of Webster et al (Webster CC, Tiribelli C, Ostrow JD. J Lab Clin Med 2001;137:370-3). Based on radioactivity and pigment content, hourly bile collections were pooled to optimize specific activities. Surprisingly, in the first dog, only 2.9% of injected radioactivity was recovered in bile and only 14.1% in urine, and the specific activities of the crystalline (3)H-UCB from the two pools were only 39.5 and 30.0 x 10(3) dpm/microg. High-performance liquid chromatography analysis revealed that only 4% of ALA degraded during 5 minutes in injection solution at pH 6.8. The low incorporation of (3)H-ALA and low specific activity of (3)H-UCB was apparently caused mainly by prior degradation and exchange of labile tritium of the (3)H-ALA and probably by enhanced endogenous ALA synthesis caused by the anesthetic/sedative agents. Revised procedures in the second dog improved the incorporation of (3)H-ALA to 11.9% excreted in bile and the specific activity of the crystalline (3)H-UCB to 122.0 and 50.8 x 10(3) dpm/microg, while urinary excretion of tritium increased to 28.5%. These experiences emphasize possible pitfalls in preparing (3)H-UCB by biosynthetic labeling from (3)H-ALA administered to dogs.

Systemic effects of acute terminal ileitis on uninflamed gut aggravate bile acid malabsorption.

BACKGROUND: Some patients with terminal ileitis suffer from significant bile acid malabsorption even if the inflammation is locally limited. We hypothesized that inflammation in the terminal ileum may lead to changes in mucosal absorption in more proximal intestinal segments and aggravate bile acid malabsorption. METHODS: Five hamsters underwent laparotomy and localized instillation of 2,4,6-trinitrobenzenesulfonic acid (TNBS) in 10% ethanol into the last 4 cm of ileum to create terminal ileitis. A control group (n = 5) underwent instillation of saline. Animals were sacrificed after 24 h. Active and passive transport of radiolabeled bile acids was measured in the proximal and terminal ileum and glucose absorption in the jejunum using an everted sleeve technique. Myeloperoxidase (MPO) activity and histomorphology were examined by standard methods. RESULTS: In animals with ileitis, active bile acid uptake decreased by 84% in the terminal ileum (t test, P <0.001) and by 58% in the proximal ileum (P < 0.05) compared with saline-treated controls. Jejunal glucose absorption decreased by 59% (P < 0.01). Passive bile acid and glucose absorption rates were not significantly changed in any segments of treated animals versus controls. Histological examination of the treated group revealed signs of acute terminal ileitis without changes in the proximal ileum and jejunum. All control tissues were uninflamed. MPO activity was 13-fold increased in the inflamed ileal samples compared with controls (P <0.001). No significant changes were seen in the proximal ileum and jejunum. There was no evidence of reflux of TNBS into proximal ileum. Nominal mucosal surface area values showed no significant changes between groups. Pretreatment of an additional group of hamsters (n = 5) with acetylsalicylic acid before TNBS instillation ameliorated the inflammatory response in the terminal ileum and largely abrogated the negative effects on ileal bile acid absorption. CONCLUSION: These data suggest that limited acute ileitis impairs active bile acid uptake in the terminal ileum. It also diminishes active bile acid and glucose absorption in more proximal segments of the small intestine, likely by a systemic effect. This systemic effect may aggravate bile acid malabsorption in patients with limited ileitis.

A simple method for measuring of intestinal solute transport in mucosal biopsy specimens.

Accurate in vitro measurements of intestinal mucosal solute uptake in humans are often difficult because only small amounts of tissue material are available. We describe a miniaturized everted sleeve method of measuring intestinal solute uptake in endoscopy biopsy samples that combines simplicity, good tissue viability and reproducibility. Biopsies were mounted on a dressmaker needle head stationed immediately over a stirring bar rotating at 1200 rpm. This approach was used to measure taurocholate uptake in sheep and human endoscopy biopsies. Comparison was made to conventional standardized everted sleeve preparations. Na+-dependent uptake rates correlated well among individual sheep (R2 = 0.88, P < 0.05). There was excellent correlation between conventional and biopsy preparations in humans (R2 = 0.98; P < 0.05). The biopsy method overestimated diffusional uptake rates in sheep and humans by two to three fold when compared to conventional everted sleeve preparations. We conclude that this method is valuable to measure Na+-dependent solute uptake rates in biopsy samples from human intestine.

Percutaneous cholecystostomy for acute cholecystitis in veteran patients.

BACKGROUND: Surgical cholecystostomy has been shown to carry a significantly higher mortality rate at Veterans Administration (VA) hospitals than at non-federal hospitals in the past. METHODS: A retrospective outcomes study was undertaken at a large VA medical center with a policy favoring radiologic over surgical cholecystostomy over the past 9 years. Records of 24 consecutive patients with acute cholecystitis were reviewed to evaluate the effectiveness of the procedure. RESULTS: Cholecystostomy was performed radiologically in 22 patients and surgically in 2 patients. Most (78%) of patients improved within 48 hours. The periprocedural mortality was 25%. The majority of these patients died from unrelated illnesses. Four patients developed complications, none of which required operative intervention. CONCLUSIONS: Comorbidities are the most important mortality factor for cholecystostomies in VA patients. Radiologic tube placement is effective and uncomplicated in most cases.

Atypical gastrointestinal symptoms are not associated with gallstones in patients with spinal cord injury.

OBJECTIVE: To determine if nonspecific gastrointestinal (GI) symptoms justify cholecystectomy in patients with spinal cord injury (SCI). DESIGN: The frequency of GI symptoms was determined in a sample of patients with SCI in whom the presence or absence of gallstones had been previously determined by screening ultrasonography or a known history of cholecystectomy. The prevalence of various symptoms in patients with and without gallstones was compared. SETTING: The Spinal Cord Injury Unit of the Veterans Affairs Puget Sound Health Care System, which provides rehabilitation and longitudinal primary care for SCI veterans. PATIENTS: Two hundred ninety-four patients who had undergone either right upper quadrant ultrasonography or cholecystectomy in the past, and who completed a questionnaire concerning GI symptoms. MAIN OUTCOME MEASURE: Bivariate logistic regression was used to calculate odds ratios (ORs) to determine the strength of associations between the presence of each symptom and the presence of gallstones. RESULTS: Pain in the right upper quadrant or epigastrium that occurred after meals or at night was significantly associated with gallstones (OR: 3.5; 95% confidence interval [CI] 1.02-11.73). Abdominal pain in other locations and nonspecific symptoms such as bloating and nausea, were not predictive of the presence of gallstones. CONCLUSIONS: Nonspecific symptoms in patients with SCI are not associated with gallstones and do not justify cholecystectomy in patients with otherwise asymptomatic gallstones.

Distribution of bile acid absorption and bile acid transporter gene message in the hamster ileum.

The apical, Na-dependent, ileal bile acid transporter (IBAT) is critical for the reabsorption of bile acids in the ileum. Bile acid transport capacities as well as the distribution of bile acid transporter messenger ribonucleic acid (mRNA) and transporter protein were studied along the axis of the ileum. Na-dependent and Na-independent taurocholate uptake was measured in the hamster ileum using an everted-sleeve technique. The distribution of IBAT mRNA and protein were mapped by in-situ hybridization, immunohistochemistry, and Western blotting. Na-dependent and Na-independent bile acid uptake rates were highest 1-4 cm before the ileocecal valve (maxima 780 and 120 pmol/mm2 per min, respectively) and decreased proximally and distally. Na-independent absorption was increased in the last 6 cm of the ileum. IBAT mRNA and protein expression were linked closely to the distribution of uptake capacity. IBAT mRNA was more abundant near the crypt-villus junction whereas the protein was expressed evenly along the villus axis. We conclude that Na-dependent and Na-independent bile acid absorption capacities both have distinct distribution curves in the hamster ileum. All ileocytes on villi in the high-uptake area of the ileum express IBAT mRNA and protein.

[Surgery education in the USA]. / Chirurgische Ausbildung in den USA.

Surgical education in the United States historically was influenced by the German educational system. Currently residents spend five years to become general surgeons. Education focuses on the teaching of surgical basic sciences and practical instruction in the operating room and at the bedside. Students interested in General Surgery are selected largely based on their performance in medical school. Admission into first-rate programs is highly competitive. In many university-based programs residents are expected to spend an additional two years in research as part of their training. While on their clinical rotations residents usually spend 80 to 100 hours per week in the hospital. Resident salaries are moderate.

The presentation of gallstones and results of biliary surgery in a spinal cord injured population.

BACKGROUND: Since spinal cord injured patients lack visceral sensation, their clinical manifestations of gallstones could be relatively occult. A higher proportion of these individuals may present with advanced biliary disease compared with the general population. Prophylactic cholecystectomy for asymptomatic stones may therefore be justified. METHODS: All spinal cord injured patients seen at the Seattle Veterans Hospital over a 5-year period were retrospectively surveyed to define a set of patients who had undergone a cholecystectomy. The operative indications and results were compared with those from a series of cholecystectomies in neurologically intact patients. RESULTS: The presentation of biliary disease in spinal cord injured patients was not more advanced than that of neurologically intact patients. Patients with high cord injuries presented in a similar fashion to those with low injuries. CONCLUSIONS: Since most spinal cord injured patients with biliary disease present with typical findings, prophylactic removal of gallstones in these patients is not warranted.

The prevalence and natural history of gallstones in spinal cord injured patients.

BACKGROUND: Individuals with a spinal cord injury are at increased risk for the development of gallstones. Because these patients cannot reliably manifest classic symptoms of biliary colic, they may be more likely to present with advanced biliary complications than patients with intact abdominal innervation. The natural history of gallstones in spinal cord injured patients has not been described. STUDY DESIGN: All spinal cord injured patients seen at the Seattle Veterans Affairs Medical Center from January 1, 1993, to December 31, 1997 were included in the study. For each patient, the presence or absence of gallstones had been determined previously through screening abdominal ultrasonographic evaluations. Pertinent demographic information was obtained from medical records and patient interviews. Patients with gallstones were followed until death, cholecystectomy, or the conclusion of the study, and the annual incidence of biliary complications and patients requiring a cholecystectomy were determined. The prevalence of gallstones was established by studying the subset of patients seen at the Seattle Spinal Cord Injury Unit from January 1, 1995 to December 31, 1997. RESULTS: Among the spinal cord injured patients, 31% either had gallstones or had undergone a cholecystectomy at some point after their injury. Increasing age, female gender, and greater severity of injury were risk factors for the formation of gallstones. Over the first 5 years after the diagnosis of gallstones, the annual incidence of cholecystectomy or biliary complications was 6.3% and 2.2%, respectively. CONCLUSIONS: Spinal cord injured patients are at increased risk for the development of gallstones. Patients with gallstones are at an increased risk for the development of biliary complications compared with neurologically intact patients, but the magnitude of this risk does not warrant prophylactic cholecystectomy.

[Telesurgery--experiences from the USA]. / Telechirurgie--Erfahrungen aus den USA.

Recent advances in telecommunications formed the basis for studies on the use of video and high resolution television for surgical teleconsulting. Experience from hospitals of the American Veterans Administration suggests the technology to be a valuable asset. Telesurgical techniques improve health care in remote areas, save costs by reducing the need for transfers, and ease the professional isolation of health care personnel. Clear recommendations for both primary care and referral centers about how to conduct telesurgical consulting are given in the text.

Colonic perforations after renal transplantation.

BACKGROUND: Perforation of the colon is a rare but serious complication in renal transplantation; allograft recipients constitute a patient population uniquely at risk with end-stage renal failure, maintenance therapy with dialysis before transplantation, and then chronic immunosuppression thereafter. STUDY DESIGN: In 1,401 consecutive transplants performed between 1951 and 1995 at the Brigham and Women's Hospital, 30 recipients (2.1 percent) experienced 34 episodes of colonic perforations, 13 of which (38 percent) were fatal. The medical records and clinic charts of each person were analyzed for variables between those who survived and those who died of the colon perforation. RESULTS: Significant differences in patient demography and clinical and laboratory findings including age, mean corticosteroid dose, and nutritional status were noted between the two groups. Early diagnosis and intervention improved the prognosis; 22 percent of those operated on within 24 hours died; 47 percent died after delayed intervention. Excision of the lesion with end-to-end anastomosis was effective in most right colon perforations, and removal of the lesion and formation of an end-colostomy and Hartmann pouch was preferred in rectosigmoid perforations. The incidence and outcome of posttransplant colonic perforations were associated with the intensity of immunosuppression, with 28 percent of perforations occurring within the first month after engraftment and 47 percent within the first 3 months; the mean daily dose of corticosteroids was three times higher in those who died compared with those who survived. The levels of serum albumin were significantly lower among nonsurvivors. CONCLUSIONS: Colon perforation is a catastrophic event in immunosuppressed renal transplant recipients. Prompt diagnosis and treatment are critical; because of the masking effects of corticosteroids on symptoms and signs, a high index of suspicion and urgent investigation are indicated. Prompt surgical intervention and attention to the nutritional status are essential for survival.

The neuronal and epithelial human high affinity glutamate transporter. Insights into structure and mechanism of transport.

High affinity transport of glutamate across plasma membranes of brain neurons and epithelial is mediated by a Na(+)- and K(+)-coupled electrogenic transporter. Here we report the primary structure and functional characterization of the human high affinity glutamate transporter (HEAAC1). A unique characteristic of HEAAC1-mediated transport is that the affinity for glutamate and the maximal transport rate are strongly dependent on membrane potential. Our data provide new insights into individual steps of high affinity glutamate transport and show that the transport mechanism is distinct from that of the gamma-aminobutyric acid transporter GAT-1 and the Na+/glucose transporter SGLT1. Under voltage clamp condition, HEAAC1 mediated large substrate-evoked inward currents (up to 1 microA). The substrate specificity, stereospecificity, the Km value (30 +/- 3 microM at -60 mV) of the L-glutamate-evoked current, and Northern analysis all agree with previously reported characteristics of high affinity glutamate transport in brain. In contrast to SGLT1 and GAT-1, voltage jump studies of HEAAC1 yielded only minor relaxation currents. Classic inhibitors of brain glutamate uptake such as DL-threo-beta-hydroxyaspartate, L-trans-pyrrolidine 2,4,-dicarboxylic acid (PDC), and dihydrokainate were found to be either transport substrates or to have no significant effect on glutamate transport. We also found that the maximal transport rate for PDC was markedly reduced compared to that for L-glutamate. We propose that PDC most likely reduces the turnover rate of the transporter. A search of the sequence data bases revealed weak homology of HEAAC1 to the H(+)-coupled vesicular monoamine transporter, suggesting an evolutionary link between plasma membrane and vesicular transporters.

Cloning and characterization of the vasopressin-regulated urea transporter.

Urea is the principal end product of nitrogen metabolism in mammals. Movement of urea across cell membranes was originally thought to occur by lipid-phase permeation, but recent studies have revealed the existence of specialized transporters with a low affinity for urea (Km > 200 mM)2. Here we report the isolation of a complementary DNA from rabbit renal medulla that encodes a 397-amino-acid membrane glycoprotein, UT2, with the functional characteristics of the vasopressin-sensitive urea transporter previously described in in vitro-perfused inner medullary collecting ducts. UT2 is not homologous to any known protein and displays a unique pattern of hydrophobicity. Because of the central role of this transporter in fluid balance and nitrogen metabolism, the study of this protein will provide important insights into the urinary concentrating mechanism and nitrogen balance.

Functional changes associated with short and long lateral ileal reservoirs in a canine model.

Colectomy and endorectal ileal pull-through with ileal reservoir (PTR) have become a standard operative procedure for severe ulcerative colitis and familial polyposis coli. Satisfactory results have been reported with a number of different reservoir designs; however, the optimal reservoir size and configuration remain controversial. Two groups of five dogs each, undergoing colectomy and PTR with construction of either 5-cm-long or 18-cm-long lateral ileal reservoirs, were studied to compare defecatory patterns, intestinal transit, and reservoir emptying. Three months postoperatively, dogs with long ileal reservoirs demonstrated a higher stool frequency and made more unsuccessful attempts at defecation. Evacuation of viscous polyethylene glycol solution from the long reservoirs was prolonged during the first hour compared with short reservoirs. Two of the five long reservoirs showed mucosal ulcerations on postmortem examination, whereas none were present in any of the short pouches. Increases in stool water content and subsequent reduction of urine volumes as well as a prolongation in oroanal transit times occurred to a similar degree in both groups. It is concluded that short lateral isoperistaltic reservoirs empty more effectively than do long reservoirs.

Expression of mRNA (D2) encoding a protein involved in amino acid transport in S3 proximal tubule.

A rat kidney- and intestine-specific cDNA (D2) that induces high-affinity, Na(+)-independent uptake of cystine and dibasic and neutral amino acids into cRNA-injected Xenopus oocytes was recently isolated by expression cloning in our laboratory (R. G. Wells and M. A. Hediger. Proc. Natl. Acad. Sci. USA 89: 5596-5600, 1992). At present it is not known whether the D2-encoded protein functions as a transporter or as a transporter activator. To gain more insight into the role of D2 in renal amino acid transport, we studied the site of its expression in the kidney. This was determined by Northern blot analysis and by using a combination of in situ hybridization and immunocytochemistry with antibodies that recognize specific proximal tubule segments. D2 antisense RNA hybridized to the same tubular segments that were strongly positive for anti-ecto-adenosinetriphosphatase but negative for carbonic anhydrase type IV and the facilitated glucose transporter GLUT2. We conclude that D2 mRNA is strongly expressed in the rat kidney proximal tubule S3 segment, although there is weak hybridization to the S1 and S2 segments. The signal is absent in all other parts of the kidney. The S3 specific expression of D2 mRNA coincides with the site of high-affinity transport of cystine and other amino acids, consistent with the proposed involvement of D2 in these processes.

Alterations in gastrointestinal motility during postoperative acute corticosteroid withdrawal.

Patients receiving exogenous corticosteroids may develop iatrogenic adrenal insufficiency, with resultant nausea, emesis, and abdominal distension if perioperative "stress steroid" dosages are inadequate. To investigate these gastrointestinal disturbances, motility measurements were obtained using perfused catheters in 10 dogs before steroid treatment (control), during administration of high-dose corticosteroids, and daily during 5 days of abrupt withdrawal. Withdrawal was characterized by a significant disruption in normal antral, duodenal, and jejunal motility with a prolongation of the migrating motor complex (MMC) and phases I and II, but not III (I = quiescence, II = irregular activity, and III = regular activity). Retrograde giant contractions (RGCs), giant migrating contractions (GMCs), and/or "intestinal fibrillation" were also observed during the first two withdrawal days. Adrenal weights and morphology did not change. We conclude: (1) high-dose corticosteroids can induce profound adrenal suppression in dogs without morphologic changes within 3 weeks; (2) high-dose steroid administration enhances gastrointestinal motility; and (3) acute withdrawal causes disappearance or shortening of MMC and the development of RGCs and GMCs with associated emesis.