[Development of an Analytical Method for Simultaneous Determinationof Quinolones and Tetracyclines in Livestock and Fishery Products].

In this study, we developed an analytical method for simultaneous determination of 14 quinolones and 4 tetracyclines in livestock and fishery products using LC-MS/MS. The analytes were extracted from food samples with citrate buffer (containing EDTA)-methanol-acetonitrile (3 : 1 : 1, v/v/v) in the presence of n-hexane, and the extract was purified with an Oasis PRiME HLB cartridge column. It was suggested that this analytical method can also extract analytes from solid samples containing fat by using n-hexane. In addition, using methanol-acetonitrile (3 : 7, v/v) containing 0.1 vol% formic acid as an eluent from the cartridge column, the purification effect could be improved, while minimizing the impairment of the recovery rate. As a result of the validation using six types of food samples, trueness (accuracy) was 70.6%-113.8%, the RSD of repeatability was 9.0% or less, and the RSD of within-laboratory reproducibility was 15.5% or less. Using this approach, the standard values mentioned in the Japanese guideline were successfully met.

Complex Modulating Effects of Dietary Calcium Intake on Obese Mice.

BACKGROUND/AIM: Οverweight and obesity are risk factors for chronic diseases. Dietary calcium has been reported to exert anti-obesity effects. However, the complex modulating effects of calcium intake on obese mice have not been clarified. MATERIALS AND METHODS: The effects of calcium intake on body weight/visceral fat mass were examined in the obese mouse model, KK-Ay Results: Body weight gain decreased in mice fed a diet containing 0.4 to 3.2% calcium at the age of 11 and 13 weeks, but not at 12 weeks after normalization for food intake. Calcium intake also decreased serum insulin levels and increased the amount of feces excreted. Fecal deoxycholate levels were lower in the high-calcium group than in the normal diet control group. Furthermore, the ratio of the deoxycholate-producing microbiome in feces decreased. CONCLUSION: Dietary calcium has anti-obesity effects in obese KK-Ay mice. Inhibition of insulin production and an increased amount of feces excreted with calcium intake may affect body weight.

12α-Hydroxylated bile acid induces hepatic steatosis with dysbiosis in rats.

There is an increasing need to explore the mechanism of the progression of non-alcoholic fatty liver disease. Steroid metabolism is closely linked to hepatic steatosis and steroids are excreted as bile acids (BAs). Here, we demonstrated that feeding WKAH/HkmSlc inbred rats a diet supplemented with cholic acid (CA) at 0.5 g/kg for 13 weeks induced simple steatosis without obesity. Liver triglyceride and cholesterol levels were increased accompanied by mild elevation of aminotransferase activities. There were no signs of inflammation, insulin resistance, oxidative stress, or fibrosis. CA supplementation increased levels of CA and taurocholic acid (TCA) in enterohepatic circulation and deoxycholic acid (DCA) levels in cecum with an increased ratio of 12α-hydroxylated BAs to non-12α-hydroxylated BAs. Analyses of hepatic gene expression revealed no apparent feedback control of BA and cholesterol biosynthesis. CA feeding induced dysbiosis in cecal microbiota with enrichment of DCA producers, which underlines the increased cecal DCA levels. The mechanism of steatosis was increased expression of Srebp1 (positive regulator of liver lipogenesis) through activation of the liver X receptor by increased oxysterols in the CA-fed rats, especially 4ß-hydroxycholesterol (4ßOH) formed by upregulated expression of hepatic Cyp3a2, responsible for 4ßOH formation. Multiple regression analyses identified portal TCA and cecal DCA as positive predictors for liver 4ßOH levels. The possible mechanisms linking these predictors and upregulated expression of Cyp3a2 are discussed. Overall, our observations highlight the role of 12α-hydroxylated BAs in triggering liver lipogenesis and allow us to explore the mechanisms of hepatic steatosis onset, focusing on cholesterol and BA metabolism.

Correlation between 12α-hydroxylated bile acids and insulin secretion during glucose tolerance tests in rats fed a high-fat and high-sucrose diet.

BACKGROUND: Previously, we found a significant relationship in a rat study between energy intake and bile acid (BA) metabolism especially 12α-hydroxylated (12αOH) BAs. The present study was designed to reveal relationships among BA metabolism, glucose tolerance, and cecal organic acids in rats fed a high-fat and high-sucrose diet (HFS) by using multivariate and multiple regression analyses in two types of glucose tolerance tests (GTTs). METHODS: Male WKAH/HkmSlc rats were fed with a control or a HFS for 13 weeks. Oral glucose tolerance test (OGTT) and intraperitoneal glucose tolerance test (IPGTT) were performed at week 9 and 11, respectively. BAs were analyzed by using ultra high-performance liquid chromatography-mass spectrometry. Organic acid concentrations in cecal contents were analyzed by using ultra high-performance liquid chromatography with post-column pH buffered electric conductivity method. RESULTS: A positive correlation of aortic 12αOH BA concentration was observed with energy intake and visceral adipose tissue weight. We found that an increase of 12αOH BAs in enterohepatic circulation, intestinal contents and feces in the HFS-fed rats compared to those in control rats regardless of no significant increase of total BA concentration in the feces in the test period. Fecal 12αOH BA concentration was positively correlated with maximal insulin level in OGTT and area under curve of insulin in IPGTT. There was a positive correlation between aortic 12αOH BAs concentration and changes in plasma glucose level in both OGTT and IPGTT. In contrast, a decrease in the concentration of organic acids was observed in the cecal contents of the HFS-fed rats. Multiple linear regression analysis in the IPGTT revealed that the concentrations of aortic 12αOH BA and cecal acetic acid were the predictors of insulin secretion. Moreover, there was a positive correlation between concentration of portal 12αOH BAs and change in insulin concentration of peripheral blood in the IPGTT. CONCLUSION: The distribution analysis of BA compositions accompanied by GTTs revealed a close relationship between 12αOH BA metabolism and insulin secretion in GTTs in rats.

Preparation and functional analysis of gossypols having two carbohydrate appendages with enaminooxy linkages.

We developed new gossypol (Gos)-based glycoconjugates through dehydration condensation of native Gos and chemically modified glycosides having aminooxy groups. The resultant glycoconjugates (glycoGos) were resistant to hydrolysis, although they were light-sensitive and slowly decomposed even under indoor lighting. The glycoGos also exhibited improved water solubility compared with native Gos, but their saturated concentrations in water were still low (6.4-17 µM), due to their hydrophobic naphthyl rings. We also carried out WST-8 assays to assess the anticancer activity of the glycoGos on DLD-1 and HepG2 cells and found that the glycoGos having ß-lactosides and having ß-galactosides (specific ligands for asialoglycoprotein receptors) showed enhanced anticancer activity on HepG2 cells.

Isolation of six novel 7-oxo- or urso-type secondary bile acid-producing bacteria from rat cecal contents.

Understanding the dynamics of secondary bile acid (SBA) formation in the gut by SBA-producing bacteria is important for host health, as SBAs have been shown to affect host pathophysiology and gut microbiota composition. However, our knowledge of SBA producers is limited in light of the diversity of gut microbes. Here, we isolated six novel SBA-producing bacteria from rat cecal contents, all of which were members of known species of gut microbes. Anaerostipes caccae D10, Bacteroides nordii C5, Clostridioides difficile D7, and Clostridium cadaveris G11 were capable of oxidizing cholic acid and chenodeoxycholic acid into 7-oxo-derivatives with varying yields. B. nordii C5 and its type strain JCM 12987T had the highest molar yield, ∼90%. Clostridium disporicum F4 and Clostridium subterminale C4 epimerized cholic acid into ursocholic acid with yields of ∼85%; the corresponding type strains lacked epimerization activity. Furthermore, although not novel as an SBA producer, Clostridium scindens G10 that produced deoxycholic acid from cholic acid was isolated for the first time from rodents. These findings will contribute to elucidation of SBA formation in the gut.

Activity of ERK regulates mucin 3 expression and is involved in undifferentiated Caco-2 cell death induced by 3-oxo-C12-homoserine lactone.

The signal molecule, 3-oxo-C12-homoserine lactone (3-oxo-C12-HSL), is similar to a mammalian hormone in bacteria. Although most studies have examined the effects of high 3-oxo-C12-HSL concentrations (>200 µM) on mammalian cellular functions because ~600 µM 3-oxo-C12-HSL can be secreted in biofilms of Pseudomonas aeruginosa grown in vitro, we previously showed that a low 3-oxo-C12-HSL concentration (30 µM) induces the apoptosis of undifferentiated Caco-2 cells through suppressing Akt activity. Here, we found that a low concentration of 3-oxo-C12-HSL-activated ERK1/2 in undifferentiated Caco-2 cells. Incubating cells with the ERK pathway inhibitor U0126 for 30 min alleviated the mucin 3 (MUC3) expression suppressed by 3-oxo-C12-HSL, and the upregulation of MUC3 expression induced by a 48-h incubation with U0126-reduced cell death. Thus, altered MUC3 expression caused by long-term attenuated ERK1/2 activity might correlate with the death of undifferentiated Caco-2 cells induced by 3-oxo-C12-HSL.

Diet supplementation with cholic acid promotes intestinal epithelial proliferation in rats exposed to γ-radiation.

Consumption of a high-fat diet increases some secondary bile acids (BAs) such as deoxycholic acid (DCA) in feces. DCA is derived from cholic acid (CA), a primary BA. We evaluated intestinal epithelial proliferation and BA metabolism in response to oral administration of cholic acid (CA) in rats to determine the influence of a CA diet on the responses of gut epithelia to γ-rays. WKAH/HkmSlc rats were divided into two dietary groups: control diet or CA-supplemented (2g/kg diet) diet. Some of the rats from each group were irradiated with γ-rays, and epithelial cell proliferation in the colon was analyzed histochemically. Unirradiated CA-fed rats had high levels of DCA and CA in the sera, as well as the presence of taurocholic acid in their feces. Significant increases were observed in both epithelial proliferation and the number of epithelial cells in the colon of the CA-fed rats, and this effect was observed at 8 weeks after γ-ray exposure. Furthermore, extracts from both cecal contents and sera of the unirradiated CA-fed rats promoted proliferation of IEC-6 cells. These results indicate that BAs in enterohepatic circulation promote proliferation and survival of the intestinal epithelium after receiving DNA damage.

Inhibition of nestin suppresses stem cell phenotype of glioblastomas through the alteration of post-translational modification of heat shock protein HSPA8/HSC71.

Nestin, a class VI intermediate filament, was first described as a neuronal stem/progenitor cell marker. We previously reported that knockdown of nestin expression in human glioblastoma cells suppresses cell proliferation, migration, and invasion. In the present study, we examined the effect of nestin on stemness, and identified molecules involved in modulating nestin function in glioblastoma cells. Nestin expression was shown to be higher in high-grade gliomas than in low-grade gliomas. Furthermore, compared with control cells, nestin short hairpin RNA (shRNA)-transfected glioblastoma cells exhibited reduced sphere formation, decreased expression of NANOG, N-cadherin, CD133, and Oct-4, and decreased tumor size in vivo. To examine the proteins regulated by nestin in glioblastomas, we carried out two-dimensional electrophoresis using nestin shRNA-transfected glioblastoma cells. As a result, nestin shRNA-transfected glioblastoma cells exhibited a decrease in the level of phosphorylation of heat shock cognate 71 kDa protein (HSC71; gene HSPA8). From immunoprecipitation experiments, we demonstrated the direct binding of nestin, HSC71, and cyclin D1 in vitro. Overexpression of nestin in glioblastoma cells increased cell growth, sphere formation, and cell invasion. Transfection with HSC71 siRNA restored nestin expression and cell behavior; therefore, HSC71 knockdown will interfere with enhanced tumorigenic properties of glioblastoma cells that ectopically overexpress nestin. We have demonstrated that HSC71 and nestin regulate each other's expression levels or patterns, and that cyclin D1 is located downstream of nestin and HSC71. In conclusion, nestin regulates stemness, cell growth, and invasion in glioblastoma cells through the alteration of HSC71. Inhibition of nestin and HSC71 may thus be a useful molecular target in the treatment of glioblastomas.

Long-term oral feeding of lutein-fortified milk increases voluntary running distance in rats.

To evaluate the effects of lutein-fortified milk administration on running exercise, a voluntary wheel-running model was performed in rats. Four-week-old F344 rats were administered test milk (10 mL/kg) daily following a 4-h fasting period, and their running distances were measured each day for a 9-week period. Total weekly running distance significantly increased from the sixth week until the end of the test period in lutein-supplemented rats (lutein-fortified milk administered) compared with control rats (vehicle administered). This increase was not apparent in rats administered lutein alone. In the lutein-fortified-milk exercise group compared with the sedentary control group, carnitine palitroyltransferase 1 (CPT-1), total AMP-activated protein kinase (tAMPK), and phosphorylated AMP-activated protein kinase (pAMPK) contents were significantly increased in the gastrocnemius muscle, with a concomitant decrease in triglyceride and total cholesterol levels in the blood and liver. Furthermore, the lutein level in blood of lutein-administered rats significantly decreased with exercise. These results suggest that lutein-fortified milk may enhance the effect of exercise by effective utilization of lipids when combined with voluntary running.

Deoxycholic acid is involved in the proliferation and migration of vascular smooth muscle cells.

Obesity is increasingly becoming associated with increased risk of atherosclerosis. Serum levels of the bile acid deoxycholic acid (DCA) are elevated in mice with obesity induced by a high-fat (HF) diet. Therefore, we investigated the influence of DCA on the functions of vascular smooth muscle cells (VSMCs) because the initiation and progression of atherosclerosis are associated with VSMC proliferation and migration. DCA induced c-jun N-terminal kinase (JNK) activation whereas a JNK inhibitor prevented DCA-induced VSMC proliferation and migration. Based on these findings, we examined whether DCA promotes the expression of platelet-derived growth factor ß-receptor (PDGFRß) that has a c-Jun binding site in its promoter region. The mRNA and protein expression levels of PDGFRß were upregulated in VSMCs after a 24- and 48-h incubation with DCA, respectively. The effects of PDGF such as proliferation and migration of VSMCs were promoted after a 48-h incubation with DCA despite the absence of DCA during PDGF stimulation. These findings suggest that elevated serum concentrations of DCA are involved in the pathogenesis of atherosclerosis in HF-induced obesity.

Contribution of dipeptidyl peptidase IV to the severity of dextran sulfate sodium-induced colitis in the early phase.

Dipeptidyl peptidase IV (DPPIV) degrades some peptide hormones and cytokines, resulting in homeostatic modulation. However, the role of DPPIV in inflammatory bowel diseases remains controversial. To determine the role of DPPIV in colitis, we used F344/DuCrlCrlj (F344/Du) rats as a DPPIV-deficient model. The serum DPPIV activity was much lower in the F344/Du rats than in F344/Jcl rats which were used as a DPPIV-positive model. Interestingly, the disease activity index (DAI) was different in the early phase of 2% dextran sulfate sodium (DSS)-induced colitis, as judged by the mucosal myeloperoxidase activity, colonic weight, and cecal fermentation. Similarly, retarded DAI was apparent in the DPPIV-deficient rats with 1% DSS-induced colitis. These findings suggest that a low level of DPPIV activity contributed to maintaining intestinal homeostasis by suppressing the cleavage of cytokines and growth hormones in DSS-induced colitis, especially in the early phase of colitis and with moderate inflammation.

Nestin: a novel angiogenesis marker and possible target for tumor angiogenesis.

Abnormal vasculature, termed tumor vessels, is a hallmark of solid tumors. The degree of angiogenesis is associated with tumor aggressiveness and clinical outcome. Therefore, exact quantification of tumor vessels is useful to evaluate prognosis. Furthermore, selective detection of newly formed tumor vessels within cancer tissues using specific markers raises the possibility of molecular targeted therapy via the inhibition of tumor angiogenesis. Nestin, an intermediate filament protein, is reportedly expressed in repair processes, various neoplasms, and proliferating vascular endothelial cells. Nestin expression is detected in endothelial cells of embryonic capillaries, capillaries of the corpus luteum, which replenishes itself by angiogenesis, and proliferating endothelial progenitor cells, but not in mature endothelial cells. Therefore, expression of nestin is relatively limited to proliferating vascular endothelial cells and endothelial progenitor cells. Nestin expression is also reported in blood vessels within glioblastoma, prostate cancer, colorectal cancer, and pancreatic cancer, and its expression is more specific for newly formed blood vessels than other endothelial cell markers. Nestin-positive blood vessels form smaller vessels with high proliferation activity in tumors. Knockdown of nestin in vascular endothelial cells suppresses endothelial cell growth and tumor formation ability of pancreatic cancers in vivo. Using nestin to more accurately evaluate microvessel density in cancer specimens may be a novel prognostic indicator. Furthermore, nestin-targeted therapy may suppress tumor proliferation via inhibition of angiogenesis in numerous malignancies, including pancreatic cancer. In this review article, we focus on nestin as a novel angiogenesis marker and possible therapeutic target via inhibition of tumor angiogenesis.

Nestin regulates epithelial-mesenchymal transition marker expression in pancreatic ductal adenocarcinoma cell lines.

Nestin, a class VI intermediate filament, is a neuronal stem/progenitor cell marker that is also expressed by various types of cancer, including pancreatic ductal adenocarcinoma (PDAC). We previously detected nestin expression in approximately 30% of PDAC cases, and found that nestin promotes the migration, invasion and metastasis of cells. Findings of recent studies have shown that epithelial mesenchymal transition (EMT) is important in the invasion and migration of cancer. In the present study, we investigated whether an altered nestin expression affected the expression levels of EMT markers in PDAC cells. Two human PDAC cell lines, PK-45H and KLM-1, in which nestin was suppressed and overexpressed, respectively, were used. The expression levels of the EMT-related molecules E-cadherin, Snail, Slug and Twist were analyzed using quantitative RT-PCR. Results showed that E-cadherin expression was decreased in nestin-overexpressed KLM-1 cells, and increased in nestin-suppressed PK-45H cells. Snail gene expression in the PDAC cells was altered concomitantly with the changes in nestin expression, while the Slug gene expression was significantly decreased in nestin-overexpressed KLM-1 cells. The Twist gene expression was below the detection limit in the two PDAC cell lines. The present findings indicated that nestin may be involved in the control of cancer behaviors in PDAC via the modulation of EMT-related molecules.

Expression of cancer stem cell markers in pancreatic intraepithelial neoplasias and pancreatic ductal adenocarcinomas.

Cancer stem cells (CSCs) play pivotal roles in cancer growth, invasion, metastasis and recurrence. Several proteins have been reported as CSC markers for pancreatic ductal adenocarcinoma (PDAC). In the present study, we examined the correlation between pancreatic intraepithelial neoplasias (PanINs) and CSC markers including CD24, CD44, CD133, CXCR4, ESA and nestin using immunohistochemical analysis. Furthermore, we examined the roles and clinical significance of these CSC markers in PDAC. CD24-, CD44-, CXCR4-, ESA- and nestin-positive cells were detected in the following tissues, listed in order of increasing percentage: normal ducts < low-grade PanINs < high-grade PanINs < PDACs. CD133 did not increase according to the malignancy grade. In PDAC, cells positive for each of the following CSC markers were detected, listed according to increasing percentage: nestin < CD133 < CD44 < CD24 < CXCR4 < ESA. CXCR4 and ESA expression correlated with well-differentiated PDAC. Venous invasion was positively associated with CD133 and inversely associated with ESA. CSC marker expression levels detected in PDAC cell lines using flow cytometry showed lowest expression of CD133 and highest of CD44, differing from the results obtained using immunohistochemistry. In two PDAC subtypes, adenosquamous carcinoma and anaplastic carcinoma, ESA was expressed more abundantly in adenocarcinoma components, whereas CD44 and nestin showed high expression in anaplastic components. Together, these results suggest that most CSC markers correlate with pancreatic carcinogenesis through the PanIN-to-PDAC sequence. Each CSC marker was related in a different manner with proliferation, differentiation, invasiveness or tissue type of PDAC.

Is bile acid a determinant of the gut microbiota on a high-fat diet?

Recently, we discovered that bile acid, a main component of bile, is a host factor that regulates the composition of the cecal microbiota in rats. Because bile secretion increases on a high-fat diet and bile acids generally have strong antimicrobial activity, we speculated that bile acids would be a determinant of the gut microbiota in response to a high-fat diet. The observed changes in the rat cecal microbiota triggered by cholic acid (the most abundant bile acid in human biliary bile) administration resemble those found in animals fed high-fat diets. Here, we discuss the rationale for this hypothesis by evaluating reported diet-induced gut microbiota alterations based on the postulate that bile acids worked as an underlying determinant. The identification of host factors determining the gut microbiota greatly contributes to understanding the causal relationships between changes in the gut microbiota and disease development, which remain to be elucidated.

Fibroblast growth factor receptor 2 IIIc as a therapeutic target for colorectal cancer cells.

A high percentage of colorectal carcinomas overexpress a lot of growth factors and their receptors, including fibroblast growth factor (FGF) and FGF receptor (FGFR). We previously reported that FGFR2 overexpression was associated with distant metastasis and that FGFR2 inhibition suppressed cell growth, migration, and invasion. The FGFR2 splicing isoform FGFR2IIIb is associated with well-differentiated histologic type, tumor angiogenesis, and adhesion to extracellular matrices. Another isoform, FGFR2IIIc, correlates with the aggressiveness of various types of cancer. In the present study, we examined the expression and roles of FGFR2IIIc in colorectal carcinoma to determine the effectiveness of FGFR2IIIc-targeting therapy. In normal colorectal tissues, FGFR2IIIc expression was weakly detected in superficial colorectal epithelial cells and was not detected in proliferative zone cells. FGFR2IIIc-positive cells were detected by immunohistochemistry in the following lesions, listed in the order of increasing percentage: hyperplastic polyps < low-grade adenomas < high-grade adenomas < carcinomas. FGFR2IIIc immunoreactivity was expressed in 27% of colorectal carcinoma cases, and this expression correlated with distant metastasis and poor prognosis. FGFR2IIIc-transfected colorectal carcinoma cells showed increased cell growth, soft agar colony formation, migration, and invasion, as well as decreased adhesion to extracellular matrices. Furthermore, FGFR2IIIc-transfected colorectal carcinoma cells formed larger tumors in subcutaneous tissues and the cecum of nude mice. Fully human anti-FGFR2IIIc monoclonal antibody inhibited the growth and migration of colorectal carcinoma cells through alterations in cell migration, cell death, and development-related genes. In conclusion, FGFR2IIIc plays an important role in colorectal carcinogenesis and tumor progression. Monoclonal antibody against FGFR2IIIc has promising potential in colorectal carcinoma therapy.

Glycochenodeoxycholic acid promotes proliferation of intestinal epithelia via reduction of cyclic AMP and increase in H2AX phosphorylation after exposure to γ-rays.

Bile acids (BAs) are considered to be promotive factors in colorectal carcinogenesis. We investigated whether BAs in the cellular environment influence proliferation of intestinal epithelial cell lines. Some BAs induced proliferation in several epithelial cell lines. In the proliferation assay, significant increases in IEC-6 cell proliferation were observed in response to glycodeoxycholic acid or glycochenodeoxycholic acid (GCDCA). Among the glycine-conjugated derivatives of BAs, especially GCDCA reduced cAMP production in IEC-6 cells. Pertussis toxin completely inhibited the GCDCA-induced increase in IEC-6 proliferation, suggesting GCDCA-induced proliferation required Gαi activation and cAMP reduction. Treatment with 3-isobutyl-1-methylxanthine, a phosphodiesterase inhibitor, also suppressed GCDCA-induced IEC-6 proliferation. We confirmed an increase in MEK1/2 phosphorylation in GCDCA-treated IEC-6 cells, and inhibition of MEK1/2 by U0126 clearly suppressed GCDCA-induced IEC-6 cell proliferation. A significant increase was observed in the phosphorylation of histone H2AX in GCDCA-treated IEC-6 cells after exposure to γ-rays. Cell cycle analysis revealed that GCDCA increased the proportion of cells in S phase only after γ-ray exposure. These results indicate that glycine-conjugated BAs in the cellular environment are potent inducers of cell proliferation accompanied by genomic instability in intestinal epithelia.

Clinicopathological features of 30 autopsy cases of pancreatic carcinoma.

The annual incidence of pancreatic carcinoma has been increasing worldwide, and the overall 5-year survival rate has remained at approximately 5%. We re-evaluated 30 autopsy cases histologically diagnosed as pancreatic carcinoma from 1994 through 2010 at Nippon Medical School Hospital. The mean patient age was 69.5 years, with no significant differences between male and female patients. The location of the primary tumor was most often the head of the pancreas (46.7%), followed by the body (36.7%) and tail (16.7%). All patients had advanced-stage pancreatic carcinoma at diagnosis, which limited the therapeutic options. Surgical resection, radiation, and surgical resection with chemotherapy were each performed for a single patient, and chemotherapy was performed for 5 patients. The other patients received only symptomatic therapy. The mean survival time from the first medical examination to death was short (5.5 months; range, 1-40 months). The cases were classified into 28 ductal adenocarcinomas, 1 acinar cell carcinoma, and 1 intraductal papillary mucinous neoplasm (IPMN) with an associated invasive carcinoma. Death in most cases was directly related to the pancreatic carcinoma, including cachexia, carcinomatous peritonitis and pleuritis, hepatic failure and ileus due to metastasis, and malignancy-related disorders, such as coagulation disorders and immunodeficiency. The most frequent site of metastasis was the lymph nodes, followed by the liver, peritoneum, spleen, lung and/or pleura, small intestine, adrenal gland, kidney, omentum, diaphragm, and bone. We classified the autopsy cases as showing distant metastasis or local infiltration. All cases with local infiltration were located in the pancreatic head, but no difference was seen in other clinicopathological features between cases with local infiltration and cases with distant metastasis. Thus, the autopsies revealed an extremely poor prognosis for pancreatic carcinoma due to the tumor itself and malignancy-related disorders. The progression pattern (i.e., local infiltration or distant metastasis) may correlate with the location of the primary tumor.

Bile acid is a host factor that regulates the composition of the cecal microbiota in rats.

BACKGROUND & AIMS: Alterations in the gastrointestinal microbiota have been associated with metabolic diseases. However, little is known about host factors that induce changes in gastrointestinal bacterial populations. We investigated the role of bile acids in this process because of their strong antimicrobial activities, specifically the effects of cholic acid administration on the composition of the gut microbiota in a rat model. METHODS: Rats were fed diets supplemented with different concentrations of cholic acid for 10 days. We used 16S ribosomal RNA gene clone library sequencing and fluorescence in situ hybridization to characterize the composition of the cecal microbiota of the different diet groups. Bile acids in feces, organic acids in cecal contents, and some blood parameters were also analyzed. RESULTS: Administration of cholic acid induced phylum-level alterations in the composition of the gut microbiota; Firmicutes predominated at the expense of Bacteroidetes. Cholic acid feeding simplified the composition of the microbiota, with outgrowth of several bacteria in the classes Clostridia and Erysipelotrichi. Externally administered cholic acid was efficiently transformed into deoxycholic acid by a bacterial 7α-dehydroxylation reaction. Serum levels of adiponectin decreased significantly in rats given the cholic acid diet. CONCLUSIONS: Cholic acid regulates the composition of gut microbiota in rats, inducing similar changes to those induced by high-fat diets. These findings improve our understanding of the relationship between metabolic diseases and the composition of the gastrointestinal microbiota.