Role of Intrinsic (Graft) Versus Extrinsic (Host) Factors in the Growth of Transplanted Organs Following Allogeneic and Xenogeneic Transplantation.

In our studies of life-supporting α-1,3-galactocyltransferase knockout (GalT-KO) pig-to-baboon kidneys, we found that some recipients developed increased serum creatinine with growth of the grafts, without histological or immunological evidence of rejection. We hypothesized that the rapid growth of orthotopic pig grafts in smaller baboon recipients may have led to deterioration of organ function. To test this hypothesis for both kidneys and lungs, we assessed whether the growth of outbred (Yorkshire) organ transplants in miniature swine was regulated by intrinsic (graft) or extrinsic (host environment) factors. Yorkshire kidneys exhibited persistent growth in miniature swine, reaching 3.7 times their initial volume over 3 mo versus 1.2 times for miniature swine kidneys over the same time period. Similar rapid early growth of lung allografts was observed and, in this case, led to organ dysfunction. For xenograft kidneys, a review of our results suggests that there is a threshold for kidney graft volume of 25 cm3 /kg of recipient body weight at which cortical ischemia is induced in transplanted GalT-KO kidneys in baboons. These results suggest that intrinsic factors are responsible, at least in part, for growth of donor organs and that this property should be taken into consideration for growth-curve-mismatched transplants, especially for life-supporting organs transplanted into a limited recipient space.

Identification of cancer stem cells derived from a canine lung adenocarcinoma cell line.

Accumulating evidence supporting the cancer stem cell (CSC) hypothesis is based on the finding that tumors contain a small population of self-renewing cells that generate differentiated progeny and thereby contribute to tumor heterogeneity. CSCs are reported to exist in several human cancers, yet only a few reports demonstrate the existence of CSCs in primary lung cancer in dogs. In this study, the authors established a cancer cell line derived from a canine primary lung adenocarcinoma and identified a side population (SP) of cells that displayed drug-resistant features. To confirm the characteristics of these SP cells, the authors investigated the tumorigenicity of the cells in vivo by using a nude mouse xenograft model. Only 100 SP cells were able to give rise to new tumors, giving a 10-fold enrichment over the main population (MP) of cells, suggesting that these cells have the cancer-initiating ability of CSCs. Further studies characterizing CSCs in canine lung adenocarcinoma might contribute to the elucidation of the mechanisms of tumorigenesis and to the establishment of novel therapeutic strategies.

The indirect alloresponse impairs the induction but not maintenance of tolerance to MHC class I-disparate allografts.

We studied the effects of indirect allorecognition on the induction and maintenance phases of tolerance in miniature swine cotransplanted with heart and kidney allografts. MHC class I-mismatched heart and kidney grafts were cotransplanted in recipients receiving CyA for 12 days. Recipients were unimmunized or immunized with a set of donor-derived or control third-party MHC class I peptides either 21 days prior to transplantation or over 100 days after transplantation. T-cell proliferation, delayed type hypersensitivity reaction (DTH) and antibody production were assessed. All animals injected with donor MHC class I peptides developed potent indirect alloresponses specific to the immunizing peptides. While untreated recipients developed stable tolerance, all animals preimmunized with donor allopeptides rejected kidney-heart transplants acutely. In contrast, when peptide immunization was delayed until over 100 days after kidney-heart transplantation, no effects were observed on graft function or in vitro measures of alloimmunity. Donor peptide immunization prevented tolerance when administered to recipients pre transplantation but did not abrogate tolerance when administered to long-term survivors post transplantation. This suggests that the presence of T cells activated via indirect allorecognition represent a barrier to the induction but not the maintenance of tolerance.

Thymectomy does not abrogate long-term acceptance of MHC class I-disparate lung allografts in miniature Swine.

UNLABELLED: We have previously reported that tolerance to class I disparate lung allografts in miniature swine could be induced using an intensive 12-day course of tacrolimus and that pretransplant sensitization with immunogenic MHC class I allopeptides failed to block the induction of tolerance. We also have previously reported the importance of the presence of the thymus in the induction of tolerance to isolated heart, kidney, and combined heart-kidney transplants. In this study, we examined the impact of thymectomy on tolerance induction in lung transplantation. METHODS: Orthotopic left lung transplantation was performed using MHC class I-disparate donors. The recipients received a 12-day course of high-dose tacrolimus (n = 6). Total thymectomies were performed in three of the swine 21 days prior to transplantation. Lung grafts were monitored by chest radiography and serial open lung biopsy. RESULTS: All euthymic recipients maintained their grafts for over 1 year. None of the thymectomized recipients has experienced graft loss in the 6 to 10 months following transplantation. Although isolated lesions of obliterative bronchiolitis were occasionally seen in one thymectomized animal on biopsy, donor-specific unresponsiveness has been observed on assays of cell-mediated lymphocytotoxicity in all recipients. Moreover, co-culture assays have shown that recipient lymphocytes can strongly inhibit the normally robust response of naïve recipient-matched lymphocytes to donor antigen. This inhibition was not seen when using stimulators primed with third-party antigens against appropriate targets. CONCLUSIONS: These data suggest that thymus-independent peripheral regulatory mechanisms may be sufficient to induce and maintain long-term acceptance of the lung allografts.

The role of indirect recognition of MHC class I and II allopeptides in a fully mismatched miniature swine model of lung transplantation.

UNLABELLED: Considerable evidence suggests that indirect recognition of MHC allopeptides plays an important role in solid-organ rejection. Here, we examine whether immunization with class I or class II allopeptides accelerates rejection in a fully MHC-mismatched lung transplant model in miniature swine. METHODS: Recipients were immunized with either donor-derived class I or class II peptides. Sensitization to the peptides was confirmed by DTH testing and in vitro proliferation assays. Nonimmunized control (n = 6), class I peptide-immunized (n = 3), and class II peptide-immunized (n = 3) swine were transplanted with fully mismatched lungs using only a 12-day course of tacrolimus. RESULTS: One control animal rejected its graft on postoperative day 103, while the others maintained their grafts for over 1 year. In the class I peptide-immunized group, two recipients rejected their grafts (days 14 and 52). The third animal has not rejected the graft (day 120, experiment is ongoing). In contrast, in the class II-peptide immunized group, only one animal rejected its graft on day 52, while the others maintained their grafts over 1 year. Both anti-donor IgM and IgG antibodies were detectable in all acute rejectors, although no alloantibody was detectable in long-term acceptors. Regardless of the fate of the graft, all animals have maintained their proliferative responses to the peptides. However, only acceptors maintained donor-specific hyporesponsiveness in cell-mediated lymphocytotoxity and mixed lymphocyte reaction assays. CONCLUSIONS: Pretransplant sensitization of lung allograft recipients to donor allopeptides accelerates graft rejection. This appears particularly true for class I-derived allopeptides, suggesting that class II molecules may be less antigenic when presented indirectly.

An MHC class II disparity raises the threshold for tolerance induction in pulmonary allografts in miniature swine.

OBJECTIVES: The mechanisms and treatment of chronic rejection in pulmonary allotransplantation remain elusive. We have induced robust tolerance to class I-disparate lung allografts in miniature swine using an intensive 12-day course of tacrolimus. Here, we tested whether a tolerant state can be induced in swine receiving fully mismatched lung allografts. METHODS: Orthotopic left lung allografts were performed using MHC class I-disparate (group 1: n = 3) or fully disparate (group 2: n = 6) donors. The recipients received a 12-day postoperative course of tacrolimus (continuous intravenous infusion; target level = 35-50 ng/mL) as their only immunosuppression. RESULTS: All swine in group 1 maintained their grafts long term without developing any lesions of chronic rejection (>497, >432, >451 days). These recipients exhibited donor-specific hyporesponsiveness in cell-mediated lymphocytotoxity (CML) and mixed lymphocyte reaction (MLR) assays. In group 2, five of the six recipients maintained their grafts long term (sacrificed on postoperative days 515, 389, 429, 481, and 438 with viable grafts). Isolated lesions of obliterative bronchiolitis were occasionally seen on biopsy, and donor-specific hyporesponsiveness on assays was consistently observed. The remaining recipient rejected its graft on day 103 with histologic findings of obliterative bronchiolitis. CONCLUSIONS: We report long-term graft acceptance without chronic immunosuppression in five of six recipients across a full MHC disparity, albeit with some evidence of obliterative bronchiolitis. These data suggest that the class II disparity inherent in a fully mismatched transplant increases the requirement for tolerance induction.

Radioresistance of cancer stem-like cell derived from canine tumours.

Cancer stem-like cells (CSCs)/cancer-initiating cells (CICs) are a small subpopulation of cancer cells that are responsible for the initiation, recurrence and metastasis of cancer. We previously demonstrated that, using the Hoechst 33342 dye-based side population technique, CSCs/CICs in canine lung adenocarcinoma cell line exist. In this study, as CSCs/CICs are known to form spheres in anchorage-independent environment in vitro, we evaluated the stemness of spheroid cells derived from canine lung adenocarcinoma and osteosarcoma cells by expression of stemness markers, and investigated radioresistance. Spheroid cells showed greater expression of stemness markers Oct-4 and CD133 gene than those of adherent-cultured cells. In nude mouse xenograft models, spheroid cells showed higher tumourigenic ability than adherent-cultured cells. In addition, spheroid cells showed significantly resistant against radioactivity as compared with adherent-cultured cells. These results suggest that spheroid cells could possess stemness and provide a CSCs/CICs research tool to investigate CSCs/CICs of canine tumour cells.

Development of the Intestinal Transplantation Model With Major Histocompatibility Complex Inbred CLAWN Miniature Swine.

BACKGROUND: Clinical intestinal transplantation (Int-Tx) is associated with some problems such as rejection, infection, graft-versus-host disease, and ischemia-reperfusion injury (IRI). To determine mechanisms of rejection as well as to develop treatment strategies for Int-Tx, this study was designed to establish both heterotopic and orthotropic Int-Tx models using major histocompatibility antigen complex (MHC) inbred CLAWN miniature swine. MATERIALS AND METHODS: Eleven CLAWN miniature swine received MHC matched but minor antigen mismatched allogenic intestinal grafts. Four animals received intestinal grafts heterotopically and kept host intestine intact. The remaining 7 animals received intestinal grafts orthotopically and resected host small intestine. Continuous infusion of tacrolimus was given from day 0 for 12 days. RESULTS: Heterotopically transplanted small intestine were well perfused after revascularization; however, grafts easily underwent ischemic changes during or soon after abdomen closure due to oppression of the grafts in the limited abdominal space. In contrast, all of 7 orthotopically transplanted intestinal grafts in which recipients' small intestine was removed from the jejunum to the ileum had no signs of severe ischemia associated with compartment syndrome. Elevation of the serum concentration of inflammatory cytokines and the progression of lethal acidosis seen in recipients of heterotipic transplantation were markedly less in the case of orthotopic transplantation. Two recipients survived more than 30 days, and 1 long-term survivor showed no evidence of rejection at day 90 despite the fact that tacrolimus was stopped at day 12. CONCLUSIONS: In this study, we demonstrated the establishment of a clinically relevant orthotopic Int-Tx model with long survival in MHC inbred CLAWN miniature swine. We believe that this unique MHC inbred swine Int-Tx model is useful for developing treatment strategies for clinical Int-Tx.

Treatment with beta-SQAG9 prevents rat hepatic ischemia-reperfusion injury.

BACKGROUND: Ischemia-reperfusion (I/R) injury occurs in various situations, including transplantation, trauma, and shock. We previously reported that the synthetic beta-SQDG (18:0), which was derived from sulfoquinovosyl diacylglycerol of the sea urchin, possessed immunosuppressive effects, such as inhibition of T-cell responses in human allogenic human mixed lymphocyte reactions (MLR) and skin allograft survival in rats. beta-SQAG9 was synthesized from beta-SQDG (18:0) to improve structural stability in aqueous solution with the same biological activities to bind to CD62L (L-selectin) and CD62P (P-selectin) in vitro. We hypothesized that beta-SQAG9 might attenuate leukocyte rolling on the endothelium and neutrophil infiltration in which L-selectin and P-selectin are key molecules. We investigated the protective effect of beta-SQAG9 against hepatic I/R injury. METHODS: Male Lewis rats were divided into 6 groups: sham, control, and treatment. Rats in the control, and the treatment groups were subjected to hepatic ischemia for 30 minutes. They were injected with PBS or beta-SQAG9 at doses of 5, 10, 25, and 50 mg/kg into the penile vein immediately before reperfusion. To assess the damage to the hepatic parenchyma, aspartate aminotransferase (AST), alanine aminotransferase (ALT), and lactate dehydrogenase (LDH) were measured and histological evaluation was performed at 6 hours after reperfusion. RESULTS: In the group treated with beta-SQAG9 at a dose of 10 mg/kg, AST, ALT, and LDH were significantly reduced, and the amount of neutrophil infiltration also was significantly reduced. CONCLUSIONS: Our data suggest that SQAG-9 (10 mg/kg) reduces the warm hepatic I/R injury.

Mechanism of the immunosuppressive effect in vivo of novel immunosuppressive drug beta-SQAG9, which inhibits the response of the CD62L+ T-cell subset.

INTRODUCTION: We synthesized sulfo-glycolipid, beta-SQAG9 (designate square beta-SQAG9 liposome, because it efficiently forms a liposome structure) that possessed immunosuppressive effects such as inhibition of T-cell responses in human allogeneic MLR and skin allograft survival in rats, and bound to CD62L (L-selectin) in vitro. In this study, we further investigated the immunosuppressive mechanism in vivo by beta-SQAG9 liposome in a skin-allografted rat model. METHODS: ACI rats (RT1(a)) were grafted skin of LEW rats (RT1(1)) treated with PBS or beta-SQAG9 liposome IV once a day for 7 days. Subsequently, we investigated the population of T cells and CD62L(+) T-cell subset in the spleen, axillary lymph nodes (ALNs), and peripheral blood of skin-allografted rats by two-color flow cytometry. RESULTS: Five of 11 (45.5%) rats that were treated with 50 mg/kg beta-SQAG9 liposome showed graft survival and another showed moderate rejection in graft. The CD62L(+) T-cell subset population in ALNs of beta-SQAG9 liposome-treated rats decreased in a dose-dependent manner. No significant difference in the T-cell population was observed between the beta-SQAG9 and control groups. These data suggest that beta-SQAG9 could bind to the CD62L(+) T-cell subset in vivo as well as in vitro and affect T-cell migration, which might lead to T-cell tolerance in vivo.

Mechanisms of photoreceptor cell death in cancer-associated retinopathy.

PURPOSE: In a previous study, both recoverin and heat shock cognate protein (hsc) 70 were recognized as autoantigens by sera from patients with cancer-associated retinopathy (CAR), and retinal dysfunction similar to CAR was inducible by intravitreous injection of anti-recoverin and anti-hsc 70 antibodies to Lewis rat. The purpose of the present study was to elucidate the effects of these antibodies on retinal photoreceptor cell functions, the contribution of caspase during the photoreceptor degeneration, and the roles of aberrant expression of recoverin in tumor cells. METHODS: As photoreceptor functions, rhodopsin phosphorylation using freshly prepared rod outer segments (ROS) and electroretinogram (ERG) were studied. Expression of recoverin in several kinds of tumors was examined by reverse transcription-polymerase chain reaction and Western blot analysis. The effects of recoverin on calcium-dependent protein phosphorylation were studied using the A549 lung adenocarcinoma cell line, which does not express recoverin. RESULTS: Rhodopsin phosphorylation in bovine ROS was significantly promoted by the addition of anti-recoverin antibody. Similar effects on rhodopsin phosphorylation and ERG impairment were observed in rat eyes treated with anti-recoverin antibody. Co-injection of caspase inhibitors with anti-recoverin antibody inhibited ERG impairment and significantly suppressed the antibody-induced enhancement of rhodopsin phosphorylation. Aberrant expression of recoverin was found in 15 of 30 tumor tissues from patients with cancer without CAR. Profiles of calcium-dependent protein phosphorylation of cell lysate from A549 cells were modulated by the presence of purified recoverin. CONCLUSIONS: These observations suggest that anti-recoverin antibody is incorporated into rod photoreceptor cells and modulates rhodopsin phosphorylation, which in turn produces activation of caspase-dependent apoptotic pathways. Regarding antibody generation in CAR, a high incidence of aberrant expression of recoverin in cancer tissues is important, as suggested previously.

Studies on inhibitors of mammalian DNA polymerase alpha and beta: sulfolipids from a pteridophyte, Athyrium niponicum.

Three sulfolipid compounds, 1, 2, and 3, have been isolated from a higher plant, a pteridophyte, Athyrium niponicum, as potent inhibitors of the activities of calf DNA polymerase alpha and rat DNA polymerase beta. The inhibition by the sulfolipids was concentration dependent, and almost complete inhibition of DNA polymerase alpha and DNA polymerase beta was achieved at 6 and 8 microg/mL, respectively. The compounds did not influence the activities of calf thymus terminal deoxynucleotidyl transferase, prokaryotic DNA polymerases such as the Klenow fragment of DNA polymerase I, T4 DNA polymerase and Taq polymerase, the DNA metabolic enzyme DNase I, and even a DNA polymerase from a higher plant, cauliflower. Similarly, the compounds did not inhibit the activity of the human immunodeficiency virus type 1 reverse transcriptase. The kinetic studies of the compounds showed that DNA polymerase alpha was inhibited non-competitively with respect to the DNA template and substrate, whereas DNA polymerase beta was inhibited competitively with both the DNA template and substrate. The binding to DNA polymerase beta could be stopped with non-ionic detergent, but the binding to DNA polymerase alpha could not.

Role of the C-terminal region of beta-amylase from barley.

To investigate the role of the C-terminal region of barley beta-amylase, plasmid delta 54 was constructed with an expression vector (pBETA92) of barley beta-amylase by site-directed mutagenesis. Escherichia coli JM109 harboring plasmid delta 54 was expected to express delta 54 beta-amylase in which 54 amino acid residues were deleted from the C-terminus. The enzyme production started in the logarithmic phase, increased linearly, and reached a maximum after 12 h. delta 54 beta-amylase gave a single activity band on isoelectric focusing (pI 6.85). delta 54 beta-amylase was purified from the cells by consecutive alpha-cyclodextrin/Sepharose 6B column chromatography. A comparison of the properties of the mutant enzyme with those of the original recombinant beta-amylase [Biosci. Biotech. Biochem. (1994) 58, 1080-1086] revealed two major differences. First, the original recombinant beta-amylase showed heterogeneity on isoelectric focusing, but delta 54 beta-amylase gave a single main band of protein (pI 6.85). Therefore, the isoelectrophoretic heterogeneity of the original recombinant beta-amylase was apparently due to its C-terminal region. Secondly, delta 54 beta-amylase lacked thermostability. Therefore, it was concluded that the C-terminal region was significantly involved in the thermostability of beta-amylase.

PCR cloning and sequencing of the beta-amylase cDNA from barley.

Polymerase chain reaction (PCR) amplification of mRNA from developing barley (cultivar Haruna two-rows) endosperm was used to clone and sequence full-length cDNA encoding beta-amylase. The beta-amylase cDNA was 1,775 bp in length. The beta-amylase was deduced to be composed of 535 amino acid residues and its molecular weight was calculated to be 59,610. Kreis et al. reported that the beta-amylase cDNA from barley (cultivar Hiproly) was 1,754 bp in length and coded for a polypeptide of 535 amino acids [Eur. J. Biochem. (1987) 169, 517-525]. A comparison of the beta-amylase sequences from Haruna two-rows and Hiproly barleys revealed nine differences in the nucleotide sequence which resulted in three changes in the amino acid residues and 21 additional nucleotides at its 3'-end in the cultivar Haruna two-rows. The three changes were as follows: Ala-233, Ser-347, Met-527 (Haruna two-rows) and Val-233, Met-347, Ile-527 (Hiproly). Lundgard and Svensson pointed out that 23 amino acid residues of the peptide fragment derived from the COOH-terminal region of barley (cultivar Gula) beta-amylase were in agreement with the deduced amino acid sequence reported by Kreis et al., with the exception of a single position (Met-527 compared to Ile) [Carlsberg Res. Commun. (1986) 51, 487-491]. Our findings described above showed Met-527 is reasonable. In the cases of beta-amylases from soybean and sweet potato, the positions that corresponded to those at 233 and 347 in the amino acid sequence of beta-amylase from barley were Ala and Ser, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Construction of a plasmid used for the expression of a sevenfold-mutant barley beta-amylase with increased thermostability in Escherichia coli and properties of the sevenfold-mutant beta-amylase.

To increase the thermostability of beta-amylase, seven kinds of single-mutant plasmids were constructed with an expression vector of barley beta-amylase by mutagenesis. The remaining activity versus temperature curves were used to determine the temperatures (T50) at which 50% of the initial activity was lost during a 30-min heating period. These mutations increased the T50 values by amounts ranging from 0.8 to 3.2 degrees C. To express the sevenfold-mutant beta-amylase in Escherichia coli, plasmid pB927 was constructed. E. coli harboring plasmid pB927 produced sevenfold-mutant beta- amylase. The T50 value of purified sevenfold-mutant beta-amylase (69.0 degrees C) was higher than that of not only the original recombinant beta-amylase (57.4 degrees C) by 11.6 degrees C but also soybean beta-amylase (63.2 degrees C) by 5.8 degrees C. The intragenic amino acid replacements were found to have simple additive effects on the thermostability of beta-amylase. The sevenfold-mutant beta-amylase was found to be stable at pHs up to 12.5, while the original recombinant beta-amylase was unstable at pHs above 9.5. The data obtained from kinetics studies suggested that the sevenfold-mutant beta-amylase acquired enhanced thermostability, but its function as a beta-amylase remained unchanged.

Human CD8 and CD4 T cell epitopes of epithelial cancer antigens.

Recent human tumor immunology research has identified several genes coding immunogenic peptides recognized by CD8 cytotoxic T lymphocytes (CTLs) in melanoma tumors. Very recently, CD4 T cell antigenic epitopes were also determined in certain melanoma tumors. The use of these peptides in conjunction with human immunotherapy could prove to be of great benefit. However, such peptides in clinically common tumors of epithelial cell origin, such as of the stomach, colon, lung, etc., have not yet been determined extensively. We describe for the first time an HLA-A31 (A*31012)-restricted natural antigenic peptide recognized by the CD8 CTL TcHST-2 of gastric signet ring cell carcinoma cell line HST-2. We also identified the HLA-DRB1*08032-restricted peptide recognized by the CD4 T cell line TcOSC-20 of squamous cell carcinoma OSC-20 derived from the oral cavity. The antigenic peptide of HST-2, designated F4.2, is composed of 10 amino acid residues with two anchor motif residues necessary for binding to HLA-A31 molecules. The synthetic F4.2 peptide enhanced the reactivity of TcHST-2 against HST-2 cells. Furthermore, introduction of an expression minigene coding F4.2 peptide to HLA-A31(+) cells conferred cytotoxic susceptibility to TcHST-2 on the cells. Some stomach cancer lines into which the HLA-A31 gene had been introduced, such as MKN28-A31-2, were lysed by TcHST-2, suggesting the presence of F4.2 peptide in at least some HLA-A31(+) stomach cancers. Furthermore, F4.2 peptide induced an F4.2 peptide-specific CTL response in at least 30-40% of HLA-A31(+) peripheral blood lymphocytes from gastric cancer patients, suggesting that F4.2 peptide could be used as a cancer vaccine for gastric tumors. The natural antigenic peptide of OSC-20 was also determined using acid extraction and biochemical separation and by mass spectrometry. Consequently, OSC-20 peptide was designated as the 6-1-5 peptide, an HLA-DRB1*08032-restricted 16-mer peptide with two possible anchor motifs. It has an amino acid sequence identical to that of human alpha-enolase, suggesting that it was derived from the processed parental alpha-enolase protein. We are presently attempting to determine the genes that code tumor rejection antigens recognized by HLA-A24- and A26-restricted T cells, including those of pulmonary and pancreatic carcinomas. The search for these antigenic peptides may lead to the identification of immunogenic peptide antigens that would be suitable for clinical use in commonly occurring epithelial cancers.

Anomalous junction of the pancreaticobiliary duct accompanied by gallbladder cancer and obstructive jaundice in a patient with high serum and bile cytokine levels.

We report a case of anomalous junction of the pancreaticobiliary duct (AJPBD) associated with gallbladder cancer and obstructive jaundice in a patient with high serum and bile cytokine levels. The patient was a 63-year-old woman who complained of right hypochondralgia. Ultrasound, computed tomography, percutaneous transhepatic cholangiography, and endoscopic retrograde cholangio-pancreatatography revealed dilation of the bile ducts, an elevated lesion of the gallbladder, and AJPBD. She underwent percutaneous transhepatic cholangio-drainage (PTCD) for obstructive jaundice. However, the total bilirubin concentration remained high 7 days after PTCD. Her serum interleukin 6 level was 57,359 pg/ml before PTCD, and gradually decreased to 10 pg/ml after PTCD. Bile interleukin 6 level was 10 pg/ml before PTCD, 8997 pg/ ml 3 h after PTCD and gradually decreased there after. Serum and bile levels of tumor necrosis factor alpha and hepatocyte growth factor were high before and after PTCD. The patient underwent an extended cholecystectomy and resection of the extrahepatic bile duct. The resected specimen showed two elevated lesions of the gallbladder which, microscopically, revealed moderately differentiated tubular adenocarcinoma. These findings suggest that pre-existing inconspicuous inflammation of the biliary tract due to reflux of pancreatic juice is involved in elevation of serum and bile cytokines, and that cytokines may participate in gallbladder carcinogenesis associated with AJPBD.

Gallbladder cancer associated with cholesterosis.

We report herein two cases of carcinoma in situ of the gallbladder associated with cholesterosis. The patient in case 1 was an 81-year-old man who underwent a cholecystectomy for cholelithiasis. The resected specimens revealed gallbladder cancer in the fundus which was diagnosed histologically as mucinous carcinoma. Other findings included 13-mm, 12-mm, and 5-mm polypoid lesions in the neck of the gallbladder which macroscopically appeared to be cholesterol polyps, but histologically demonstrated carcinoma in situ with cholesterosis. The patient in case 2 was a 76-year-old man in whom ultrasonography revealed a highly echogenic, elevated lesion in the gallbladder. Cholecystectomy was performed, and a 33 x 28-mm papillary, elevated lesion with cholesterosis was resected from the neck of the gallbladder. Histologically, this was demonstrated to be papillary adenocarcinoma in situ with cholesterosis surrounded by glandular dysplasia. The distribution of the carcinomas and cholesterosis in both of these patients suggests that the adenoma or carcinoma of the gallbladder had occurred first. Then, the tumor epithelium absorbed cholesterol from the bile, and foamy cells were produced. Thus, when treating cholesterol polyps, it should be remembered that it is often difficult to distinguish between cholesterol polyp and gallbladder cancer with cholesterosis.

Synthetic sulfonolipids deduced from sulfonoquinovosyl diacylglycerols of sea urchin reduces hepatic ischemia-reperfusion injury in rats.

BACKGROUND: In hepatic surgery and liver transplantation, ischemia-reperfusion (I/R) is an unavoidable process, and protection against hepatic I/R injury is a major unresolved problem. In this study, we investigated whether 3-O-(6-deoxy-6-sulfono-beta-D-glucopyranosyl)-1,2-di-O-acylglycerol bound to saturated C18 fatty acids (beta-SQAG9), which was derived from sea urchin intestines, could reduce this injury. This agent was recently reported to have immunosuppressive effects in allogeneic rat skin grafts. MATERIALS & METHODS: Male Lewis rats were divided into two experimental groups. Group 1 rats were injected with SQAG9 (50 mg/kg) into the penile vein 15 minutes before the induction of ischemia and into the portal vein just reperfusion. The same amounts of normal saline were injected into rats in the control group (group 2). Each experimental groups included six rats. Seventy percent hepatic ischemia (20 minutes) was induced by occluding the blood vessels and bile duct with a vascular clamp. For examination of hepatic function, serum levels of aspartate aminotransferase, (AST) alanine transaminase (ALT), and lactic dehydrogenase (LDH) were measured. In addition, histological examination was also assessed. RESULTS: Three hours after reperfusion, the mean plasma concentration of AST, ALT, LDH in group 1 was suppressed compared with group 2. Six hours after reperfusion, the hepatic damage in group 1 was mild in comparison with that in group 2. CONCLUSIONS: Our data demonstrated that SQAG-9 reduced the warm hepatic I/R injury.

Serum and bile interleukin 6 after percutaneous transhepatic cholangio-drainage.

BACKGROUND/AIMS: This study was designed to assess the correlation of interleukin 6 (IL-6) with jaundice reduction by percutaneous transhepatic cholangio-drainage (PTCD) for obstructive jaundice. METHODOLOGY: We measured serum and bile IL-6 concentrations by enzyme linked immuno-sorbent assay before and after PTCD in 26 patients with obstructive jaundice. The patients were divided into two groups according to their bilirubin decrease rate. RESULTS: Serum IL-6 levels measured before and after PTCD in the slowly decreasing bilirubin group ("poor" group) were significantly higher than those of the rapidly decreasing bilirubin group ("good" group). Measures of bile IL-6 in the "poor" group remained high from 3 to 14 days after PTCD in comparison with the "good" group. CONCLUSIONS: These results suggest that circulating IL-6, as well as IL-6 in the bile, may be involved in the pathogenesis of obstructive jaundice, and play an important role in preventing the resolution of obstructive jaundice after PTCD.