Effects of MDR1/P-glycoprotein expression on prognosis in advanced colorectal cancer after surgery.

Resistance to chemotherapeutic agents is a major problem for successful cancer treatment. P-glycoprotein (Pgp), a product of the multidrug resistance (MDR)1 gene expressed in cancer cells, is one of the mechanism of MDR. However, there are few reports regarding the effects of Pgp on prognosis of colorectal cancer (CRC) after surgery. We examined a total of 80 patients (45 males and 35 females with an average age of 69 years) whose CRCs were classified into stage 2-4 and completely resected surgically in our institute between January 1990 and September 1999. To evaluate Pgp expression in CRC, immunohistochemical stain was performed with a monoclonal antibody. Relationships between Pgp expression and clinicopathological variables which may have affected prognosis were evaluated. Survival curves were calculated using the Kaplan-Meier method, and differences were evaluated with the log-rank test. The Cox's proportional hazards model was used in the univariate and multivariate survival analysis. Pgp expression showed a significant correlation with histological differentiation (p=0.023). However, no correlation was observed with gender, tumor location, lymph node metastasis, lymphatic invasion, venous invasion, and cancer stages. Survival rates after surgery tended (p=0.093) to be higher in Pgp (+) than Pgp (-) patients. Pgp was not a significant prognostic factor by univariate analysis and multivariate analysis adjusted for other clinicopathologic variables. Survival rates after surgery tended to be higher in Pgp (+) than Pgp (-) patients and Pgp expression was correlated with histological differentiation of CRC. Thus, a relative resistance of CRC to conventional chemotherapy may be partly caused by Pgp expressed in well or moderately differentiated CRC. However, Pgp expression was not a significant independent prognostic factor in advanced CRC after surgery.

Postsurgical sequential methotrexate/5-FU and leucovorin on outpatient basis for advanced colorectal carcinoma.

BACKGROUND/AIMS: The present study compared the effects of sequential methotrexate and 5-fluorouracil followed by leucovorin rescue (MFL), as an adjuvant chemotherapy versus a combination of UFT and mitomycin C (MMC), on patient survival and recurrence after surgery for colorectal carcinoma. METHODOLOGY: Between January 1990 and December 1997, a total of 55 patients with advanced colorectal cancer were treated postsurgically by adjuvant chemotherapy using MFL or UFT-MMC. Surgical treatment was performed according to standardized procedures for radical resection of colorectal cancer. The patients were divided into 2 groups after surgery. The MFL regimen consisted of MTX (100 mg/m2) and 5-FU (600 mg/m2) at hour 24, followed by leucovorin rescue. The UFT-MMC regimen consisted of MMC (12 mg/m2) intraoperatively and MMC (6 mg/m2) every other week after surgery for 2 months, and oral UFT (375 mg/m2/day), a combination of tegafur and uracil in a molar ratio of 1:4, was continued for 3 years or longer depending on the patients tolerance. RESULTS: The overall survival rates after surgery were significantly (P < 0.05) higher in the MFL than the UFT-MMC group. Recurrence rates were significantly lower in the MFL than the UFT-MMC group, especially for liver recurrence. Disease-free survival was significantly (P < 0.05) higher in the MFL than the UFT-MMC group. CONCLUSIONS: These results demonstrated the superiority of MFL therapy for improving postsurgical survival in patients with advanced colorectal cancer, in particular those patients with a high risk of recurrence following potential curative resection.

Sigmoid colon diverticulosis adherent to mesh plug migration after open inguinal hernia repair.

PURPOSE: Tension-free hernia repair with polypropylene mesh plug and patch is currently the most popular technique for open inguinal hernioplasty. It is well tolerated by most patients with few complications. Despite these excellent results, late-onset complications may occur. METHODS: An 83-year-old man reported 2 weeks of bloody stool. His medical history was significant for a left open inguinal herniorrhaphy with the mesh plug and patch technique. Barium enema revealed a stenotic segment in the sigmoid colon and multiple diverticulosis. Because a malignant lesion could not be ruled out, the patient underwent an operation. Laparotomy revealed an inflamed sigmoid colon with diverticulosis adherent to a hard tumor, which was mesh plug used for the previous open inguinal hernia repair. After mobilization of the adhesion between the mesh plug tumor and the sigmoid colon, sigmoidectomy was performed. The patient's postoperative course was uneventful. CONCLUSIONS: We reported a case of sigmoid colon diverticulosis adherent to mesh plug migration after open inguinal hernia repair. The potential risk of plug migration should be well understood by the surgeon. To avoid this risk completely, several methods have been proposed such as suturing the plug and patch together, or using an all-in-one design such as the Prolene Hernia System (Johnson and Johnson Co., Tokyo, Japan).

Hepatic undifferentiated (embryonal) sarcoma in an adult: a case report and review of the literature.

Undifferentiated (embryonal) sarcoma of the liver (USL) is a rare malignant tumour with a poor prognosis. The absence of specific symptoms, the rapid tumour growth, the normality of the common tumour markers, and the consequential delay in the diagnosis often result in significant enlargement of the tumour. To our knowledge, there have been only 42 reported cases of USL in adults worldwide during the 40 years since the clinicopathological entity of USL was defined. We report here a 27-year-old male with USL who has been treated successfully with surgical resection and adjuvant chemotherapy using ifosfamide, adriamycin and cisplatin. Although the prognosis of the disease remains generally poor, long-term survival has been achieved recently in patients with a combination of surgery and multi-agent chemotherapy.

Postsurgical sequential methotrexate, fluorouracil, and leucovorin for stages 3 and 4 gastric carcinoma: A preliminary study.

BACKGROUND AND OBJECTIVES: The present study compared the effects of sequential methotrexate and fluorouracil followed by leucovorin rescue (MFL), as an adjuvant chemotherapy vs. UFT (a combination of uracil and tegafur), on patient survival and recurrence following surgery for advanced gastric carcinoma. METHODS: Between July 1990 and June 1998, a total of 54 patients with advanced gastric cancer were treated postoperatively by adjuvant chemotherapy using MFL or UFT. Surgical treatment was performed according to standardized procedures for radical resection of gastric cancer. The patients were stratified into two groups following surgery. The MFL regimen consisted of methotrexate (100 mg/m2) and 5-fluorouracil (600 mg/m2) at hour 3, followed by leucovorin rescue. The oral UFT (375 mg/m2/day), a combination of tegafur and uracil in a molar ratio of 1:4, was continued for 3 years or longer depending on the patients tolerance. RESULTS: In stage 3 gastric cancer, the overall survival rates following surgery was significantly (p < 0.05) higher in the MFL than the UFT group. Difference in disease free survival was not statistically significant between the groups. Recurrence rates showed a trend (p = 0.08) to decrease in the MFL than the UFT group. In stage 4 gastric cancer, no significant difference was obtained in the overall survival rates between the groups. CONCLUSIONS: The present results suggested the superiority of MFL treatment for improving postoperative survival in patients with advanced gastric cancer, in particular for those patients with a high risk of recurrence following potential curative resection. In patients with stage 4 gastric cancer, however, MFL treatment showed similar effects as UFT on the postsurgical survival of the patients.

Density of Helicobacter pylori infection evaluated semiquantitatively in gastric cancer.

Helicobacter pylori infection may play a role in the development of gastric cancer; however, a quantitative evaluation of the density of H. pylori infection has not been reported previously in relation to the histologic stage and type of cancer. This study was designed to compare the density of H. pylori infection to the histologic stage and type of gastric cancer. Between March 1996 and March 1998, surgical resection of primary lesion was performed in 50 patients with gastric cancer (39 men and 11 women with a mean age of 67 years) at our institution. Using immunohistochemical stains, the density of H. pylori infection was evaluated semiquantitatively at cancer site as well as noncancerous mucosa adjacent to cancer. This density was compared with the histologic stage and the type of gastric cancer. The severity of the mucosal atrophy was evaluated using the updated Sydney System. The prevalences and density of H. pylori infection decreased in proportion to advances in the cancer stage and the mucosal atrophy. In early cancer of the intestinal- and diffuse-type, the prevalence of H. pylori in adjacent sites was almost 90% and was significantly higher (p < 0.01) than that seen in the advanced cancer lesions. In the intestinal-type early cancer, the prevalence and density of infection was higher (p < 0.05) in the adjacent mucosa than in the cancer site, whereas in the diffuse-type early cancer, H. pylori was found in all cases at the cancer site and the adjacent site. In advanced cancer, the prevalence of H. pylori was about 40% in the adjacent site and about 10% in the cancer site in both histologic types. These figures were significantly lower (p < 0.01) than in the early cancers. The prevalence and density of infection did not differ in the intestinal- and diffuse-type gastric cancers, but did decrease with more advanced cancer stages. The changes in local environment of the advanced cancer may not be conducive to the survival of H. pylori. Thus, the prevalence of H. pylori may be affected by the histologic stage rather than the histologic type of gastric cancer, and the organism may play a similar role, but through different pathways, in the pathogenesis of both types of cancer.

Modified rapid urease test for Helicobacter pylori detection in relation to an immunohistochemical stain.

BACKGROUND: The rapid urease test and touch cytology have been used for the rapid detection of Helicobacter pylori infection. Recently, a modified rapid urease (MRU) test, which provides results in 20 min has been available on a commercial basis. To date, few reports have evaluated the accuracy of this test. This study evaluated the sensitivity, specificity, and accuracy of the MRU test and touch cytology to detect H. pylori in relation to the density of H. pylori infection determined semi-quantitatively by using immunohistochemical stains. METHODS: Biopsy specimens obtained from a total of 60 patients who underwent endoscopy for evaluation of gastroduodenal diseases were studied by using the MRU test, Giemsa stain for touch smear tissue and histological methods. An immunohistochemical stain was used as a standard, and the density of H. pylori infection was graded according to the number of individual bacteria seen as follows: grade 0 = 0; grade 1+ = 1-9; grade 2+ = 10-29; grade 3+ = 30-99; grade 4+ > or = 100. The severity of gastritis was evaluated histologically in each specimen and compared with the density of H. pylori infection. RESULTS: The MRU test had an overall sensitivity of 73%, specificity of 100% and accuracy of 85%. The Giemsa stain had a sensitivity of 91%, specificity of 100% and accuracy of 95%.The sensitivities of the MRU test and Giemsa stain decreased in mild H. pylori infection. In the MRU test, the sensitivity was 47% when the density of H. pylori infection was 1+, while 80-100% sensitivities were obtained when the densities of infection were > or = 2+. With the Giemsa stain, the sensitivity was 80% when the density was 1+, while the sensitivity increased to 100% when the densities were > or = 2+. The severity of gastritis determined by the Rauws scores showed a positive correlation with the density of H. pylori infection as evaluated by immunohistochemical staining. CONCLUSIONS: The MRU test had high sensitivity and specificity for moderate to severe H. pylori infection, but it may result in false-negative results in tests for mild infection. As the MRU test has the advantages of shorter incubation times and low cost, a combination of the MRU test and the Giemsa stain for touch cytology may be the most time- and cost-efficient tests in a clinical setting for the diagnosis of H. pylori infection.

Jejunal pouch to avoid stricture after esophagojejunostomy with circular stapler.

BACKGROUND: Anastomotic stricture is one of the most common problems in esophagojejunostomy using the end-to-end anastomosing (EEA) instrument (Auto Suture Co, Norwalk, CT) after total gastrectomy. To alleviate the stricture, several methods, such as incision to the scar, balloon dilatation, and steroid injection are available. To avoid stricture, the jejunal pouch may allow use of a larger EEA than Roux-en-Y (ReY) reconstruction does. STUDY DESIGN: A total of 45 patients underwent curative total gastrectomy and esophagojejunostomy with jejunal pouch construction (27 patients) or ReY (18 patients), using the EEA. The effects of jejunal pouch construction with a large EEA on avoidance of stricture and benefit to nutritional status were investigated by comparing it with the ReY in terms of postoperative morbidity, postprandial symptoms, and nutritional parameters (serum protein, serum albumin, body weight). RESULTS: EEA28 or larger could be used in 25 patients in the pouch group and 8 patients in the ReY group (p < 0.05). Stricture developed in one patient in the pouch group and in four patients in the ReY group (p < 0.05). Postprandial symptoms were experienced less frequently (p < 0.05) in the pouch group than in the ReY group. When stricture and symptoms were analyzed according to the size of EEA, they occurred more frequently (p < 0.05) in the patients with EEA25 than those with EEA28 or EEA31. No significant differences were evident in nutritional parameters. CONCLUSIONS: The choice of jejunal pouch technique allowed the use of a larger EEA than that of ReY reconstruction, resulting in avoidance of anastomotic stricture and postprandial symptoms, though little benefit in nutritional status was evident to the patients after total gastrectomy.

A case of colon cancer recurrence with P-glycoprotein treated by methotrexate, fluorouracil, and leucovorin.

A 68 year-old female underwent right hemicolectomy for an advanced cecum cancer and had been well without any evidence of recurrence for a year after surgery. Despite post-operative treatment with oral Tegafur (400 mg/m2/day), CEA level increased gradually beginning 15 months after surgery. Sequential chemotherapy with methotrexate (MTX) and 5-Fluorouracil (5-FU), followed by leucovorin rescue (MFL) was started on an outpatient basis, and has been continued every 4 weeks since then. It consisted of MTX (100 mg/m2) and 5-FU (600 mg/m2) started 24 hours after MTX, followed by oral leucovorin (15 mg/body) started 30 hours after MTX 6 times at intervals of 6 hours. CEA level declined initially, but increased slowly for 3 years on MFL, although no evidence of recurrence was detected by imaging studies with computed tomography, ultrasound, and scintigram. Four years after surgery, a tumor recurrence developed in the abdominal wall. The patient underwent resection of the tumor, resulting in a decline of the CEA level. She has been alive and well for 5 years on MFL after the primary surgery. Both the original tumor and recurrent tumor showed immunoreactivity for P-glycoprotein. The present case demonstrates the feasibility of using MFL on an outpatient basis, and its potential to suppress the colon cancer growth with P-glycoprotein expression.

Density of Helicobacter pylori infection in patients with peptic ulcer perforation.

BACKGROUND: A lack of change in prevalence of severe ulcer complications requiring emergency operation has been reported, despite the common use of histamine-2 (H2)-receptor antagonists and proton pump inhibitors. This may be attributable to use of ulcerogenic drugs or Helicobacter pylori (HP) infection, or both. In this study, HP infection was evaluated semiquantitatively in patients with peptic ulcer who required surgery, and the severity of histologic change was investigated. METHODS: We reviewed a total of 113 consecutive patients (98 men and 15 women) operated on for perforation, hemorrhage, or stenosis of gastroduodenal ulcer between January 1986 and December 1995. Detection of HP was carried out by immunohistochemical staining. We graded the density of HP infection according to the number of individual HP bacteria counted in a highly magnified visual field (x 1,000 of light microscopy). The grade of HP infection was defined as follows: (0) = 0; (1+) = 1-9; (2+) = 10-29; (3+) = 30-99; (4+) > or = 100. The severity of gastritis was evaluated by histologic examination using the criteria of Rauws. RESULTS: Although the number of operations for gastroduodenal ulcer declined significantly, the rate of emergency operation for gastroduodenal ulcer increased from 60% to 90%, with the result that the frequency of operations for perforation or bleeding remained virtually constant and that for stenosis significantly decreased. HP infection was more prevalent in perforated ulcer (92%) than hemorrhagic ulcer (55%) or stenotic ulcer (45%). The grades of HP infection were 3.0 +/- 0.14 (mean +/- SEM) in perforated ulcer, 2.3 +/- 0.34 in hemorrhagic ulcer, and 2.5 +/- 0.22 in stenotic ulcer. Perforated ulcer was associated with significantly more severe HP infection and gastritis changes than hemorrhagic ulcer or stenotic ulcer. CONCLUSIONS: This study indicates that patients with perforated ulcer were infected with HP more severely than those with hemorrhagic ulcer or stenotic ulcer at the time of surgery. A close relationship was observed between the perforated ulcer and the density of HP infection determined semiquantitatively using immunohistochemical stain.

Analysis of factors affecting the graft oxygenation state in living related liver transplantation.

BACKGROUND/AIMS: In liver transplantation, graft dysoxia after reperfusion may lead to graft failure. The aim of this study is to investigate the relationship between the factors, which were supposed to affect the oxygen supply to the graft, and the oxygenation state of the graft in order to determine which factor is important to prevent the graft from dysoxia. MATERIALS AND METHODS: The relationship between oxygen supply and oxygenation state of the graft was investigated in 56 successful cases of living related liver transplantation. Factors affecting the oxygen supply to the graft were considered as follows; portal venous flow (PVF), mean velocity of the hepatic artery (HA-Vm), hemoglobin concentration in the peripheral blood (Hb), size of the graft liver relative to the recipient body weight (G/R ratio), partial oxygen pressure in the arterial blood (PaO2), and rate-pressure product (BP*PR). Oxygenation state of the graft was estimated by oxygen saturation of hemoglobin in the graft tissue (graft SO2) as measured by tissue near infrared spectroscopy. RESULTS: 1) Graft SO2 was rather independent of PVF and HA-Vm probably due to compensatory interrelation between the portal venous flow and hepatic arterial flow. 2) Significant correlation between G/R ratio and graft SO2 was observed after portal reflow (p < 0.01), but the correlation diminished after hepatic arterial reflow. Positive correlation between G/R ratio and AKBR after portal reflow suggested that the graft with large G/R ratio is likely to suffer dysoxia early after reperfusion. 3) Graft SO2 was positively correlated with Hb (p < 0.05), while there was no significant correlation between graft SO2 and PaO2 or BP*PR. CONCLUSION: This study clarified the contribution of the factors which were supposed to affect the oxygen supply to the graft and the oxygenation state of the graft, and which factor is important to prevent the graft from dysoxia.

Anatomical and physiological problems of Segment IV: liver transplants using left lobes from living related donors.

We reviewed the problem of Segment IV in using left lobes from living related donors, in 18 left-lobe transplants performed on pediatric patients who ranged in age from 6.0 to 17.3 yr, and in body weight from 19.8 to 58.0 kg. The separate monitoring of oxygen saturation of hemoglobin in the liver sinusoid of segments, using a spectrophotometric technique, demonstrated a delay in re-oxygenation of Segment IV after the portal reflow, and revealed its return to comparable oxygenation with Segments II and III by the re-arterialization. Hemoglobin content, which was determined by the same technique, occasionally increased in Segment IV during the operation, implying sluggish microcirculation caused by inadequate hepatic venous drainage. These characteristic profiles on tissue oxygenation and hemodynamics in Segment IV should be considered when using left lobes as living related liver grafts.

Graft size-matching in living related partial liver transplantation in relation to tissue oxygenation and metabolic capacity.

The influence of graft size-matching on tissue oxygenation and metabolic capability was studied in living related partial liver transplantations for 47 pediatric patients. Their age ranged from 4 months to 17 years 3 months, their body weight from 4.0 to 58.0 kg, graft weight from 191 to 440 g, and graft weight/recipient body weight ratio from 0.61% to 6.0%. Tissue oxygenation and its heterogeneity were investigated by measuring oxygen saturation of hemoglobin in the liver sinusoid (SO2), coefficient of variation of SO2, and arterial ketone body ratio. The metabolic capacity of the graft was investigated by measuring bilirubin clearance, recovery of cholesterol esterification, and ketone body production. In infants with a relatively large liver graft, both intra- and extracellular oxygenation remained low soon after reperfusion but recovered to the control value by the end of the operation. In adolescent recipients of a relatively small graft, by contrast, synthetic and detoxification capacities were relatively deficient; however, these improved with time. These results indicate that sufficient tissue oxygenation and liver regeneration are essential for successful liver transplantation with relatively large and small grafts, respectively.

Quantitative analysis of redox gradient within the rat liver acini by fluorescence images: effects of glucagon perfusion.

The redox gradient along the sinusoid in the rat liver was studied using a redox scanner, a device based on tissue fluorescence scanning spectroscopy measuring the fluorescence signals of oxidized flavoprotein (FP) and reduced pyridine nucleotide (PN). The FP/(FP+PN) ratio reflects the mitochondrial redox state in the liver tissue. The distribution of mitochondrial redox state on the scanned area is expressed as two-dimensional gray-scale images with a 20 micron resolution. Using this instrument, we have scanned a 2.5 x 2.5 mm area of the frozen rat liver sample to investigate the redox gradient within acini and the effects of glucagon on the changes in the redox distribution. The redox images obtained in the perfused livers showed mosaic patterns implicating a regular heterogeneity of redox state in an acinus. The analysis of gradient curve, furthermore, clarified that the redox level in an acinus decreased sigmoidally from the periportal to the pericentral region. Glucagon, which has been reported to reduce the intracellular redox state, decreased the redox potential in whole acini, especially, in the periportal region, when compared with the perfusion without glucagon. These results strongly indicate an intraacinus heterogeneity of glucagon function, with glucagon selectively operating in the upstream of the sinusoid.

Primary and reversible injury of H(+)-ATPase in warm ischemia and reperfusion of rat liver in relation to intramitochondrial adenine nucleotide.

The injury and recovery processes of complex reactions of liver mitochondrial ATP synthesis during warm ischemia and after reflow were studied separately in terms of the changes in oxidation (electron transfer system) and phosphorylation (H(+)-ATPase). Oxidative activity decreased significantly from the control value of 40 +/- 0.9 (mean +/- SEM, n = 5) to 31.5 +/- 1.13 (nanoatoms oxygen consumed/min/mg protein) after 40 min of warm ischemia, while phosphorylative activity decreased significantly from the control value of 1.06 +/- 0.12 to 0.42 +/- 0.03 (mumole ATP hydrolyzed/min/mg protein) after 20 min of warm ischemia. During 120 min of reflow after 20 min of warm ischemia, the decreased phosphorylation activity recovered to 0.52 +/- 0.01 concomitant with a recovery of intramitochondrial total adenine nucleotide and an increase in the ATP/ADP ratio, while oxidative activity decreased further to 23.9 +/- 0.81. These results indicate that H(+)-ATPase is more vulnerable to warm ischemia than the electron transfer system, but that it is restored concomitant with the recovery of intramitochondrial adenine nucleotide content.

Evaluation of water molecules in the cold-preserved rat liver by proton magnetic resonance imaging.

Hypothermically preserved rat livers were studied with proton magnetic resonance imaging (1H-MRI) under proton density-, spin-lattice relaxation time-, spin-spin relaxation time- and diffusion-weighted (P-W, T1-W, T2-W and D-W) conditions. Relative signal intensities (RSI) of the liver to distilled water in terms of P-W, T1-W, T2-W and D-W increased time-dependently during 12 h hypothermic (4 degrees C) preservation with saline, while these parameters did not increase during preservation with University of Wisconsin (UW) solution. One-hour Wiggers' hypotensive treatment before the harvesting increased the RSIs of P-W, T2-W and D-W, and the subsequent 12-hour preservation with UW solution did not improve the increased RSIs. These results suggest that 1H-MRI has potential application in evaluating the biophysical changes of water molecules in the liver graft, which were measured by placing the harvested liver in a plastic bag under a magnetic field at a low temperature.

Intraoperative measurement of the graft oxygenation state in living related liver transplantation by near infrared spectroscopy.

Graft oxygenation plays an important role in successful liver transplantation. Intraoperative changes in the oxygenation state of the liver graft were measured by near infrared spectroscopy in 28 cases of living related liver transplantation. Oxygen saturation of hemoglobin in the liver (hepatic SO2) changed from 81.2% +/- 1.5% (mean +/- SEM) before donation (in the donor) to 49.7% +/- 4.2% after portal reflow, to 58.4% +/- 5.0% after arterial reflow, and then to 71.4% +/- 3.9% before closure. Mean hepatic SO2 was positively correlated with portal flow rate as measured by duplex Doppler sonography. Cases with low portal flow rate showed a high coefficient of variation (SD/mean) of hepatic SO2, indicating heterogeneous tissue oxygenation. Though graft size was expected to affect the graft oxygenation state, hepatic SO2 was fairly independent of the graft-to-recipient weight ratio. In two cases with markedly low hepatic SO2, postoperative graft dysfunction occurred. This study suggest that the method of near infrared spectroscopy is reliable and useful for assessing the graft oxygenation state in liver transplantation.

Cold preservation-induced changes in oxygen radical generation between parenchymal and nonparenchymal cells in rat liver.

Many of the reports implicating the contribution of oxygen radicals to preservation-reperfusion injury have been based largely on indirect experiments demonstrating the effects or the consumption of various antioxidants. Investigations based on the direct measurement of the amounts of oxygen radicals that are actually formed during reoxygenation after preservation have not given satisfactory results. In this study, we attempted direct measurement of H2O2 from hepatocellular mitochondria and superoxide (O2-) from Kupffer cells, using the HRP method and cytochrome c perfusion method, respectively, for quantitative comparison of the cold preservation-induced changes in radical generation activity between these sources. H2O2 generation in mitochondria isolated after 24 h cold preservation decreased to 8% of non-preserved liver, but in the mitochondria isolated from the livers that were reperfused for 30 min after 24 h preservation H2O2 generation recovered to 60%. The respiratory control ratio also decreased significantly after 24 h preservation, and similarly recovered after an additional 30 min reperfusion. By contrast, O2- from Kupffer cells increased in time-dependent fashion until 12 h preservation and decreased after 24 h preservation. Although 12 h preservation did not cause an increase in LDH release, the lipid peroxide in the perfusate significantly increased after 12 h preservation, which indicated the occurrence of lipid peroxidation in the sinusoidal area. These results suggested that mitochondrial H2O2 was dependent upon the activity of respiratory function and so did not cause hepatocellular injury and that O2- from Kupffer cells contributed to oxidative injury to the sinusoidal lining cells. Our data support reports demonstrating the vulnerability of nonparenchymal cells.