N Ohkohchi

Surgical limitation for biliary atresia: indication for liver transplantation

Of 245 patients undergoing corrective operations for biliary atresia, jaundice was cleared in 113. In January 1988, 84 of them were living and free of jaundice and the other 19 were alive with jaundice. A vast majority of long-term survivors showed normal growth and development, and were leading normal lives for their respective ages. Portal hypertension, a common late complication, improved spontaneously or after sclerotherapy in jaundice-free patients. Therefore, liver transplantation is not recommended for jaundice-free patients even with esophageal varices. Patients with persistent severe jaundice (serum bilirubin over 10 mg/dL) and those with moderate jaundice (serum bilirubin 5 to 10 mg/dL) and severe esophageal varices require liver transplantation. Patients with moderate jaundice having no or slight varices should be carefully followed. When varices become worse or serum bilirubin rises, liver transplantation is indicated. Patients with mild jaundice (serum bilirubin lower than 5 mg/dL) have a possibility of improvement in their condition before the age of 15 years, and are not recommended for liver transplantation. The high value of the lowest postoperative bilirubin level suggests the necessity of liver transplantation in early childhood. Liver transplantation as the primary treatment for biliary atresia may be indicated only for patients over 120 days of age with an enlarged and hard liver.

Tumor necrosis factor-induced, superoxide-mediated neutrophil accumulation in cold ischemic/reperfused rat liver.

The mechanisms of hepatic ischemia/reperfusion injury are complicated and multifactorial. This study was designed to examine superoxide generation and neutrophil accumulation in cold ischemic‐reperfused rat livers after elimination of Kupffer cells and to determine the role of superoxide/tumor necrosis factor (TNF) interactions. Rat Kupffer cells were eliminated by liposome‐encapsulated dichloromethylene diphosphonate injected intravenously. Livers from control and treated rats were isolated and preserved in University of Wisconsin solution (4°C) for 0, 12, and 24 hours and then perfused for 60 minutes with oxygenated Krebs‐Henseleit bicarbonate buffer (37°C) by adding neutrophils into the perfusate. Superoxide generation was measured by using real‐time chemiluminescence (CL) during perfusion, and neutrophil accumulation was assessed by measuring myeloperoxidase activity in the liver tissue. In the control livers, CL intensity markedly increased on reoxygenation, and after neutrophil infusion it increased again with a lag period of 10 minutes. Total CL intensity and myeloperoxidase activity increased with the duration of cold preservation. TNF release into the effluent perfusate was detectable only after 24 hours of preservation, and lactate dehydrogenase release was high. Elimination of Kupffer cells attenuated CL intensity and TNF and lactate dehydrogenase release and resulted in reduced myeloperoxidase activity. Electron microscopy revealed amelioration of hepatocyte swelling and endothelial cell disruption when Kupffer cells were eliminated. After 24 hours of preservation, superoxide generation was inhibited in the control livers by anti‐TNF antiserum, whereas TNF release was not inhibited by superoxide dismutase. These results suggest that TNF induces superoxide generation by Kupffer cells, which mediates neutrophil accumulation and causes cellular injury in the initial phase of reperfusion.

New Technique for Gene Transfection Using Laser Irradiation

Background We have developed a gene transfection system using laser beams. The principle of this procedure is that a small hole is made in a cell membrane by pulse laser irradiation, and a gene contained in a medium is transferred into the cytoplasm through the hole. This hole disappears immediately with the application of laser irradiation of the appropriate power. Methods A pulse-wave Nd:YAG laser with a wavelength of 355 nm was used to make a hole in a cell membrane. To trap a cell, a continuous-wave Nd:YAG laser with a wavelength of 1015 nm was used. Plasmids that encode the enhanced green fluorescent protein (EGFP) gene were contained in a medium and transferred to HuH-7 and NIH/3T3 cells with pulse laser irradiation. We evaluated transfection efficiency on the basis of the number of cells that expressed EGFP. Stimulatory protein 2 cells in suspension were fixed using a trapping laser and the neomycin-resistance gene was transfected by pulse laser irradiation. We examined cell proliferation in the selection medium. Results Cells that expressed EGFP were recognized in the group that was irradiated by pulse laser. No cells expressed EGFP without irradiation. Transfection efficiency was ≈10% at a plasmid concentration of 10.0 μg/mL. At concentrations greater than 20 μg/mL, the transfection rate reached a plateau. We also successfully transfected neomycin-resistance genes to cells floating in suspension after fixation that was achieved with trapping laser irradiation. Conclusions This method enables us to transfect targeted cells, ie, cells in suspension as well as attached cells, with a simple technique that does not involve harmful vectors. The present method is very useful for gene transfection in cellular biotechnology.

Liver transplantation for severe hypoxemia caused by patent ductus venosus

The authors describe the case of a 9-year-old girl who underwent liver transplantation because she suffered from severe hypoxemia caused by patent ductus venosus (PDV). Generally, severe hypoxemia (PaO2 < 50 mm Hg in room air or < 300 mm Hg in pure oxygen) is not an indication for liver transplantation because the hypoxemia may not be improved, and may lead to a fatal outcome. PDV, which is associated with mild liver dysfunction, is not an indication for liver transplantation by itself. But in our patient, most of the mesenteric venous flow directly entered the systemic circulation through the PDV just like the portosystemic shunt, and this caused the pulmonary arteriovenous shunt and hypoxemia. Thus, the authors operated on the patient in an attempt to restore her pulmonary function. Nitric oxide (10 to 20 ppm) was added to the inhaled gas to dilate the functional pulmonary capillaries and to deliver sufficient oxygen after the transplantation. Although the patient suffered various complications after the operation, the final results were excellent.

Mechanism of D-xylose transport in human small intestine

Summary: The transport mechanism of D-xylose across the mucosal border of small intestine was investigated in humans both in vitro and in vivo. A comparative study was also made between infants and adults. We investigated the Na+ dependence of D-xylose influx from the mucosal solution into the epithelial cells, the phlorizin sensitivity of the influx, transport kinetics and electro-genie property of the D-xylose transport. The results of kinetic study indicated that the transport of D-xylose was completely diffusional. The influx of D-xylose across the mucosal border of human small intestine was Na+ independent and phlorizin insensitive. The electrical property of D-xylose transport could not be observed. There was no difference in the characteristics of D-xylose transport between small intestines of infants or adults. These results in the present study suggest that D-xylose does not share a common transport mechanism of D-glucose, and xylose tolerance test does not give any information about the absorptive capacity of D-glucose.

Disorder of bile acid metabolism in children with short bowel syndrome

The profile of fecal bile acids was examined in 13 children with short bowel syndrome; 7 of the 13 did not have diarrhea and the other 6 had intractable diarrhea. In children without diarrhea, no severe fat malabsorption was recognized, and the content of total bile acids in the feces was within the normal range or slightly higher. The ratio of primary to total bile acids showed various patterns. In children with intractable diarrhea, in contrast, fat malabsorption was observed and the fecal content of total bile acids in these patients was more than ten times higher than that of the control group, primary bile acids accounting for more than 95% of the total bile acids and taurine- or glycine-conjugated bile acids for 10%. In the children with intractable diarrhea, the values for thed-xylose absorption test were lower than the normal range. These results suggested that, in children with short bowel syndrome with diarrhea, the loss of bile acids was strongly associated with a decrease in the actual absorptive surface area of the residual small intestine, and the growth of the normal bacterial flora was disturbed in the residual intestine. Some children with or without dirrhea also had hyper bile acidemia. Ursodeoxycholic acid was not effective for the treatment of hyper bile acidemia or fat malabsorption.

Long-term follow-up study of patients with cholangitis after successful Kasai operation in biliary atresia: selection of recipients for liver transplantation.

Since the introduction of cyclosporine A, liver transplantation has become accepted as the therapy for end-stage liver disease. However, there are no definite criteria for liver replacement in biliary atresia. We investigated (a) the survival rate after hepatic portoenterostomy (n = 131), (b) liver function tests in fatal cases after an initially successful hepatic portoenterostomy (n = 9), and (c) liver function tests in the patients with episodes of cholangitis after a successful surgical treatment (n = 8). Patients with persisting jaundice after the surgery cannot be expected to survive long, and therefore they definitely should undergo liver transplantation. When total bilirubin concentration was above 10 mg/dl in patients with cholangitis after a successful operation, conservative therapy had almost no effect. Therefore, patients with total bilirubin levels above 10 mg/dl should be considered for liver transplantation. Of the liver function tests, only total bilirubin was reliable as a marker for hepatic failure in the end stage of biliary atresia. Prolongation of thrombo test and episodes of gastrointestinal bleeding also were used in selection of patients for liver replacement.

Reoperation in patients with biliary atresia.

Twenty-seven reoperations were done on 23 patients among 100 infants with biliary atresia who have been treated at Tohoku University Hospital between 1971 and 1981. Nineteen patients had a single reoperation and 4 patients had 2 reoperations. We present the results and the role of reoperation in biliary atresia patients in our institution. Excellent bile drainage after reoperation was obtained in 13 of 15 patients with good bile flow after the initial operation. On the contrary, good bile drainage was not obtained by reoperation in 8 of 12 cases without active bile flow after the initial operation. Cessation of bile flow after successful initial operation is an absolute indication for reoperation. Aggressive reoperations under proper indications improve the surgical results in biliary atresia patients.

Changes of serum cytokines associated with hepatic regeneration after living-related liver transplantation

IN CLINICAL orthotopic liver transplantation, the graft frequently deteriorates due to various mechanisms, including preservation injury, warm ischemia during implantation, and reperfusion injury. After hepatocyte damage, some cytokines stimulate hepatocytes and play key roles in the regulation of hepatic regeneration. In livingrelated liver transplantation (LRLT), we often must use partial livers, which are smaller than the standard liver volume of recipients. After transplantation, the size of the partial liver graft increases rapidly in a short period of time. During graft enlargement, it is believed that changes of serum cytokines reflect the status of the regeneration process of the liver graft. In this study, we measured serum cytokines that play a the role in the regulation of liver regeneration, (ie, hepatocyte growth factor [HGF], interleukin-6 [IL-6], and transforming growth factor-b1 [TGFb1]. The purpose of this study was to investigate the relationship between changes of serum cytokine levels and growth of the graft, as well as recovery from graft damage.

Mechanism of primary graft non-function in a rat model for fatty liver transplantation

Abstract We established a fatty liver model in rat suitable for the model of human liver with steatosis by cholesterol enriched chow, and investigated the mechanism of primary graft non‐function in fatty liver transplantation (LTx) using this model. Grafts with steatosis caused primary graft dysfunction after LTx following even short cold preservation; however, no significant difference was recognized in mitochondrial function of the graft during preservation. Morphological findings were not different at 1 h after reperfusion between non‐steatotic and steatotic livers. Focal necrosis of hepatocytes was seen and the sinusoidal endothelial cells were injured 24 h after reperfusion. In addition, the fluidity of the plasma membrane decreased in fatty liver. Our results indicate that deterioration of sinusoidal endothelial cells after reperfusion causes graft dysfunction in LTx of steatotic liver.