Luc Lambotte

Saccharomyces boulardii upgrades cellular adaptation after proximal enterectomy in rats.

BACKGROUND Saccharomyces boulardii is a non-pathogenic yeast which exerts trophic effects on human and rat small intestinal mucosa. AIMS To examine the effects of S boulardii on ileal adaptation after proximal enterectomy in rats. METHODS Wistar rats, aged eight weeks, underwent 60% proximal resection or transection and received by orogastric intubation either 1 mg/g body wt per day lyophilised S boulardii or the vehicle for seven days. The effects on ileal mucosal adaptation were assessed eight days after surgery. RESULTS Compared with transection, resection resulted in mucosal hyperplasia with significant decreases in the specific and total activities of sucrase, lactase, and maltase. Treatment of resected animals with S boulardii had no effect on mucosal hyperplasia but did upgrade disaccharidase activities to the levels of the transected group. Enzyme stimulation by S boulardii was associated with significant increases in diamine oxidase activity and mucosal polyamine concentrations. Likewise, sodium dependentd-glucose uptake by brush border membrane vesicles, measured as a function of time and glucose concentration in the incubation medium, was significantly (p<0.05) increased by 81% and three times respectively in the resected group treated withS boulardii. In agreement with this, expression of the sodium/glucose cotransporter-1 in brush border membranes of resected rats treated with S boulardii was enhanced twofold compared with resected controls. CONCLUSION Oral administration of S boulardii soon after proximal enterectomy improves functional adaptation of the remnant ileum.

Steatosis is not sufficient to cause an impaired regenerative response after partial hepatectomy in rats.

BACKGROUND/AIMS Fatty liver is known to be associated with increased mortality and morbidity after liver resection. The ability of fatty liver to regenerate after two-thirds partial hepatectomy was studied in three different models of steatosis in rats: obese Zucker rats, orotic acid-fed Wistar rats and Wistar rats fed a methionine-low, choline-deficient diet. METHODS Liver regeneration was assessed 24 h after partial hepatectomy by bromodeoxyuridine incorporation (immunohistochemistry), proliferating cell nuclear antigen, cyclin E and cyclin-dependent kinase 2 protein expression (Western blot analysis) and cyclin-dependent kinase 2 activity (kinase assays using histone H1 as a substrate). RESULTS No significant difference of proliferative response was found between orotic acid or methionine-low, choline-deficient diet-fed and control Wistar rats 24 h after partial hepatectomy. In contrast, hepatocyte proliferation in obese Zucker rats after partial hepatectomy was significantly reduced when compared with their lean controls. CONCLUSIONS Steatosis per se does not impair liver regeneration. The reduced liver regeneration observed in obese Zucker rats may not be due to fatty infiltration itself but to other factors such as leptin receptor dysfunction.

In vivo IgM depletion by anti-mu monoclonal antibody therapy. The role of IgM in hyperacute vascular rejection of discordant xenografts.

Xenoreactive natural antibodies (XNA) and complement activation are thought to be the 2 main factors responsible for the hyperacute vascular rejection (HVR) of discordant xenografts. The aim of this work was to study the role of IgM XNA in the HVR of guinea pig to rat cardiac xenografts, a discordant model. Adult LOU/C rats were depleted of circulating IgM and therefore of IgM XNA using an anti-mu mAb (mouse anti-rat IgM mAb 7 [MARM-7]). Rats were injected with 10 mg and 5 mg of MARM-7 at days -3 and -1, respectively, and guinea pig cardiac xenografts were performed on day 0. Control animals were injected on the same days with 10 mg and 5 mg of anti-alpha mAb (MARA-1) or equivalent volumes of PBS. Xenografts were performed on day 0. Guinea pig cardiac xenograft survival time was significantly prolonged in IgM-depleted animals (62 min, P < 0.01) compared with controls using PBS (18 min) or MARA-1 mAb (12 min). This prolongation was not due to a decrease in the complement activity in IgM-depleted rats, since no significant variation of the C1q, C4, C3, and C5 complement hemolytic activity was observed between control and treated animals before HVR. Prolongation of the xenograft survival time in the MARM-7-treated group was correlated with an undetectable serum level of IgM and IgM XNA and a lack of IgM XNA deposits on the rejected xenograft vascular endothelium. Contrarily, both IgM-depleted and control animals showed C3 deposits on the rejected xenograft vascular endothelium and myocardium, as well as diffuse deposits of IgG2a XNA. Although HVR was not abrogated by the depletion of IgM XNA, our data indicate that IgM is implicated in the HVR and that the anti-mu approach is a potential therapeutic treatment for discordant xenografts. Finally, we suggest that other factors such as IgM-independent activation of complement might be one of the mechanisms responsible for the pathogenesis of HVR in the guinea pig to rat xenograft model.

Role of Metabolic Overload in the Initiation of DNA Synthesis following Partial Hepatectomy in the Rat

Changes in hepatic ATP energy charge (EC = ATP +0.5 ADP/ATP + ADP AMP) as an expression of metabolic overload and oxidative phosphorylation were studied conjointly with DNA synthesis after partial hepatectomy (PH) in the male rat. ATP and EC showed a significant decrease while mitochondrial phosphorylative activity was enhanced within 24 h. Confronted with the pattern of DNA synthesis, the above changes were clearly separated in time from the actual process of DNA synthesis. Fasting delayed the recovery of EC as well as the peak value in the rate of thymidine incorporation. Extended glucose infusion prevented the drop of ATP during the entire period of treatment and considerably reduced fat infiltration and glycogen breakdown. In these glucose-infused rats, unchanged blood sugar was associated with tendency for plasma insulin to rise and suppression of the usual posthepatectomy hyperglucagonemia. With these metabolic and hormonal changes, an important delay in the onset and modification of the whole pattern of DNA synthesis were observed. The latter process began consistently only after a late fall of ATP which followed the cessation of glucose infusion. It is suggested that changes in energy metabolism, taken as an expression of hepatocyte metabolic overload following PH, account for the early events involved in the initiation of DNA synthesis, and probably regulate hepatocyte response to systemic hepatotrophic factors.

Arterialization of the portal vein in conjunction with a therapeutic portacaval shunt. Hemodynamic investigations and results in 75 patients.

Seventy-five cirrhotic patients were submitted to peroperative hemodynamic investigations including flow and pressure studies. Sixty-two patients with a hepatopedal portal flow underwent a therapeutic end-to-side portacaval shunt (PC) in conjunction with arterialization of the portal vein and 13 with a stagnant flow a PC shunt alone. Thirty-five patients were operated on in emergency and 40 electively. In 61 patients portal flow was correlated with maximum perfusion pressure (r = 0.66), and in 33 patients with the reduction of corrected sinusoidal pressure induced by the occlusion of the portal vein (r = 0.72). Operative mortality, which was 3.5% for 57 class A and B patients and 55.5% for 18 class C patients, differed significantly (p < 0.05) in emergency between arterialized (14.8%) and nonarterialized patients (62.5%). At the time this study was ended on July 15, 1981, the follow-up was over two years for all the patients. The five-year actuarial survival rate of the arterialized patients was 48% for the whole group and 56% for class A and B patients; the overall incidence of chronic encephalopathy was 20%. It is concluded that arterialization is a safe surgical procedure that could be beneficial in respect with operative mortality in emergency, late survival, and toierance to portacaval shunt. However, a prospective randomized study such as the one undertaken in December 1979 is the only method to prove clearly that arterialization is really able to minimize the risk of encephalopathy and to prolong the long-term survival after portacaval shunt.

Measurement of the vasoconstrictive substances endothelin, angiotensin II, and thromboxane B2 in cold storage solution can reveal previous renal ischemic insults.

Abstract In a rat model, the left kid ney was subjected to 60 min of normothermic ischemia followed by 15 min of reperfusion, whereas the right kidney, serving as a paired control, was not rendered ischemic. Both kidneys were then perfused in situ with either Euro‐Collins (EC) solution (n=12) or University of Wisconsin (UW) solution (n=6) for 10 min. Each kidney was then harvested and stored at 4°C in its respective solution. After 24 and 48 h of cold storage, the following vasoactive substances were measured in the preservation media: endothelin (ET), angiotensin II (A‐II), thromboxane (B2) (TxB2), and prostaglan‐din I2 (PGI2). After 24 h in EC solution, left kidneys uniformly produced significantly higher concentrations of each vasoactive substance than right kidneys: ET 1.64 ± 0.3 pg/ml vs 0.82 ± 0.1 pg/ml (P 0.009); A‐II 20.8 ± 6.2 pg/ml vs 7.75 + 2.3 pg/ml (P 0.007); TxB2, 100.8 ± 17.7 pg/ml vs 40.1 f 11.7 pg/ml (P 0.04); PGI2, 638.3 ± 41.1 pg/ml vs 318.3 ± 36.4 pg/ml (P 0.001), respectively. At 48 h, a similar pattern of results was obtained as the kidney continued to produce TxB2 and prostacyclins during the 24–48 h period. In the UW solution, basal levels of ET and A‐II were lower than those in EC solution, but similarly increased after initial ischemia. At 24 h, the concentrations produced by the left and right kidneys were as follows: ET 0.66 ± 0.1 pg/ml vs 0.48 ± 0.1 pg/ml (P 0.14); A‐II 10.36 f 3.7 pg/ml vs 2.14 ± 0.7 pg/ml (P 0.006); TxB2 178 ± 53 pg/ml vs 52 ± 23.1 pg/ml (P 0.001); and PGI2 448.3 ± 49 pg/ml vs 323 ± 44.3 pg/ml (P 0.01), respectively. After 48 h, the range of concentrations of each substance was similar to that obtained after 24 h. In further studies, the concentrations of ET and A‐II were measured in solution previously used to preserve human kidneys (n=7). The mean concentration of ET and A‐II in these samples was 3.82 ± 1.14 pg/ml and 21.3 ± 9.2 pg/ml, respectively, whereas in control media both substances were below the limits of detection. These results demonstrate that vaso‐constrictive substances can be measured in the preservation media after a kidney has been stored cold and that higher concentrations are found when the organ has been subjected to prior normothermic ischemia. The measurement of these vasoactive substances before transplantation may reveal that the kidney has been subjected to previous ischemic events. Moreover, these vasoactive substances could be involved in the early recovery of renal function after kidney transplantation.

Early hepatocyte, endothelial, and bile duct cell injury after pediatric liver transplantation from cadaveric or living-related donors.

BACKGROUND When compared with cadaveric grafts (Cad), the potential advantages of pediatric orthotopic liver transplantation (OLT) from living-related (LR) donors may include better graft quality, shorter ischemic time, appropriate preparation of the recipient, and better immunologic compatibility. METHODS The aim of this study was to analyze early hepatocyte, endothelial, and bile duct cell injury following pediatric OLT using LR (n=15) or uncomplicated Cad reduced-size (n=10) grafts. Median (range) total ischemic times were 190 min (105-261) versus 760 min (418-948) in LR and Cad groups, respectively (P<0.001). RESULTS The post-OLT cytolytic profile, assessed daily during the first 7 days using both plasma glutamate-pyruvate transaminase and serum alpha-glutathione S-transferase, showed significantly higher levels of both parameters for the 10 uncomplicated Cad cases when compared with the 15 LR grafts (P<0.001). The evaluation of hepatic endothelial cell function during the first week after OLT, using serum hyaluronic acid levels, suggested lower endothelial injury in the LR grafts, when compared with the Cad grafts (P=0.059). Bile duct cell injury, as assessed using plasma gamma-glutamyl transferase levels, was similar in both groups, with a progressive increase at the end of the first week after OLT, which was correlated with a similar incidence of early acute rejection in both groups (80% in the LR group vs. 62% in the Cad group, NS). CONCLUSION (1) The hepatocellular and endothelial cell damage was reduced after OLT with LR grafts, which may be related to shorter ischemic time when compared with Cad grafts; (2) the putative immunologic advantage for LR grafts was not confirmed in terms of incidence of acute rejection.

The unreliability of the lidocaine/monoethylglycinexylidide test for assessment of liver donors.

The serum monoethylglycinexylidide (MEGX) level 15 min (t15) after i.v. administration of lidocaine (1 mg/kg) in liver donors was retrospectively correlated with graft outcome and early hepatic function. Among the 35 orthotopic liver transplants studied, 4 recipients had to be retransplanted within 10 days post-OLT because of early graft nonfunction or dysfunction, and 3 recipients died, with a median (range) donor MEGX t15 (ng/ml) of 100 (86–119) and 169 (146–182), respectively. The remaining 28 OLT patients living with functioning grafts had a donor MEGX of 87 (18–245). No significant correlations could be found between donor MEGX t15 and recipient mean and peak glutamic-oxaloacetic and pyruvic transaminases, total serum bilirubin, or mean and minimum prothrombin time values studied from day 1 to day 5 post-OLT. Moreover, categorization of donors using the MEGX t15 cut-off point of 80 ng/ml could not predict liver graft quality, as previously suggested. In summary, MEGX t15 in liver donors correlated neither with graft outcome nor with early functional parameters. Accordingly, the MEGX test should not be used as an isolated discriminatory evaluation for organ utilization.

Noninvasive measurement of liver regeneration with positron emission tomography and (2-11C)thymidine

The feasibility of liver regeneration determination with [2-11C]thymidine and positron emission tomography was investigated in partially hepatectomized rats. Serial tomographic scans were performed over a 120-minute period after injection of [2-11C]thymidine together with tritium-labeled thymidine. Within 10 minutes after injection, positron emission tomography scans showed a twofold higher hepatic uptake in regenerating than in nonregenerating livers. Time-activity curves over the liver area indicated that the maximal uptake was followed by a faster decrease of 11C radioactivity in controls than in regenerating animals, so that total 11C activity remaining in the liver at 120 minutes accounted for 68% of maximum in regenerating and only 38% in controls. Tissue distribution studies performed at 120 minutes showed that total 11C radioactivity, expressed in percent injected dose per gram, was six times higher in regenerating livers than in controls (0.62% +/- 0.07% in regenerating livers and 0.10% +/- 0.03% in nonregenerating livers; P less than 0.001) and correlated with 3H radioactivity measured in the nuclear fraction (r = 0.92; P less than 0.001). When the hepatic uptake was expressed in percent of dose per organ, the difference between both groups increased (2.31% +/- 0.23% in regenerating livers and 0.29% +/- 0.02% in nonregenerating livers; P less than 0.001) because of higher weight of regenerating livers than of nonregenerating livers (3.83 +/- 0.11 g in regenerating livers and 2.96 +/- 0.16 g in nonregenerating livers; P less than 0.001). In other organs examined, no difference in 11C radioactivity was found between the two groups of rats. These results indicated the potential usefulness of [2-11C]thymidine and positron emission tomography for noninvasive measurement of liver regeneration.

Prevention of acute cyclosporine nephrotoxicity by atrial natriuretic factor after ischemia in the rat.

The present experimental study investigates whether the atrial natriuretic factor (ANF) is able to prevent the nephrotoxic effects of cyclosporine infused after 30 min of warm renal ischemia in the rat. At 2 hr after the end of ischemia, the glomerular filtration rate was improved by an ANF infusion: 390 +/- 19 microliters/min/100 g versus 298.3 +/- 31 microliters/min/100 g in ANF and saline-infused rats, respectively (P less than 0.05). Intravenous CsA infusion at a dose of 2.5 mg/kg/day produced a more pronounced fall in GFR when compared with the control: 205.4 +/- 19.7 microliters/min/100 g versus 298.3 +/- 31 microliters/min/100 g in CsA and saline, respectively (P less than 0.05). In contrast, a synthetic rat atriopeptin III (0.5 microgram/kg/min) infusion after ischemia given together with CsA prevented its deleterious effects upon GFR: 316 +/- 22 microliters/min/100 g versus 205.4 +/- 19 microliters/min/100 g in ANF/CsA versus CsA alone (P less than 0.001). Moreover, the natriuretic ANF effects remained unaffected by high plasma CsA peak levels: indeed, other parameters of renal function--urinary flow, urinary sodium concentration and excretion rates, and urinary sodium reabsorption and fractional excretion rates, were significantly increased in ANF alone or CsA/ANF groups. These preliminary results suggest that ANF may be useful in renal transplantation or in the management of patients given large doses of CsA (liver or heart transplant) since, despite nephrotoxic CsA levels (greater than 1500 ng/ml), ANF provides an improved GFR and tubular function after ischemia.