Indeed, in 267

Indeed, in 267 HDAC inhibitor treatment-naïve Asian patients with CHB under entecavir treatment, steatosis has recently been reported to represent an

independent predictor of viral response, which, if confirmed by independent studies, would advise for a specific antiviral strategy in CHB patients with steatosis.[53] Despite the limitations related to the cross-sectional design and the limited number of subjects considered with coexistent genetic and acquired risk factors for steatosis, strenghts of our study consist in the possibility to analyze one of the largest series of well-characterized biopsied CHB patients of Western countries with systematic assessment of liver steatosis and fibrosis as well as to evaluate, for the first time, the effect of the I148M PNPLA3 polymorphism on steatosis in CHB. In conclusion, the PNPLA3 I148M polymorphism is an independent predictor of steatosis and, especially, of severe steatosis in patients with CHB. The study also suggests that steatosis is highly

prevalent in Italian CHB patients with indications for liver biopsy and is related to genetic and metabolic, but not to viral, factors. “
“Obesity is associated with chronic inflammation and contributes to the development of insulin resistance and nonalcoholic fatty liver disease. The suppressor of cytokine signaling-3 (SOCS3) protein is increased in inflammation and is thought to contribute to the pathogenesis of insulin resistance by inhibiting insulin and leptin signaling. Therefore, we studied the metabolic effects of liver-specific SOCS3 deletion in vivo. We fed wild-type (WT) and liver-specific SOCS3 knockout (SOCS3 LKO) Trametinib order mice either a control diet or a high-fat diet (HFD) for 6 weeks and examined their metabolic phenotype.

We isolated hepatocytes from WT and SOCS3 LKO mice and examined the effects of tumor necrosis factor α and insulin on Akt phosphorylation and fatty acid metabolism and lipogenic gene expression. Branched chain aminotransferase Hepatocytes from control-fed SOCS3 LKO mice were protected from developing tumor necrosis factor α–induced insulin resistance but also had increased lipogenesis and expression of sterol response element–binding protein-1c target genes. Lean SOCS3 LKO mice fed a control diet had enhanced hepatic insulin sensitivity; however, when fed an HFD, SOCS3 LKO mice had increased liver fat, inflammation, and whole-body insulin resistance. SOCS3 LKO mice fed an HFD also had elevated hypothalamic SOCS3 and fatty acid synthase expression and developed greater obesity due to increased food intake and reduced energy expenditure. Conclusion: Deletion of SOCS3 in the liver increases liver insulin sensitivity in mice fed a control diet but paradoxically promotes lipogenesis, leading to the development of nonalcoholic fatty liver disease, inflammation, and obesity. (HEPATOLOGY 2010.) Obesity is associated with type 2 diabetes and the metabolic syndrome and is a major cause of morbidity and mortality.


had a larger effect than injury severity on WMS-II


had a larger effect than injury severity on WMS-III scores (average Cohen’s d=−1.27). Clinical implications of these findings are discussed. “
“The specificity of the Word Memory Test (WMT) effort indices was examined in 48 individuals with minimal to mild head injury (MHI) in the acute stages post-injury. None of the individuals was involved in litigation or disability claims. At the established cut-offs, the WMT had an unacceptable false-positive rate (18%). T test analysis was also carried out for WMT passers and failures on a battery of neuropsychometric measures and across a range of demographic variables. The WMT was performed at a significantly lower level on the Wechsler Memory Scale – III word list sub-tests and verbal fluency tests (p < .05). This suggests that WMT failure may be indicative of a specific deficit in verbal processing in the acute phase of MHI. "
“In this paper, the effectiveness of interventions for SB203580 in vivo executive disorders

was reviewed. The objective was to evaluate the internal and external validity of intervention studies. A total of 46 papers, describing 54 studies, conducted in the last two decades meeting several preset inclusion criteria, was included in this review. The studies were categorized into three treatment approaches in order to enhance comparability. The overall results show that many interventions yield positive outcomes and seem to be effective in reducing executive problems in brain-injured subjects. However, several studies have only an explorative intent and are based on less sophisticated experimental designs. The verification of their results is generally this website more tenuous. The internal validity, or the set-up of experimental conditions Succinyl-CoA necessary to draw valid conclusions about treatment effectiveness, including the choice of well-matched control groups, or the randomization of patients over treatment and control conditions, is not always strong. The same conclusion can be drawn for the external validity of a number of the intervention studies; often evidence of generalization to real-life situations, long-term follow-up, and

transfer to non-trained situations, were (partially) lacking in the studies under review. The authors are aware that the design of proper randomized controlled trials for the investigation of the treatment effectiveness of executive disorders is cumbersome and time consuming. Nonetheless, the provisional results of several well-designed studies described in this review make the effort worthwhile. “
“Data for copying and delayed recall (after a 15-min delay) of the Modified Taylor Complex Figure (MTCF), an alternative form of the Rey-Osterrieth Complex Figure (ROCF), were collected from 290 healthy participants. Normative data are provided. Age and education were significantly correlated with MTCF scores and must be corrected for to interpret results accurately.

Then proliferation of LoVo cells was determined by methyl thiazol

Then proliferation of LoVo cells was determined by methyl thiazol tetrazolium (MTT) assay, and observed the changes of cell morphology by inverted microscope. The activities

of motility and invasion of LoVo cells were assessed by transwell chamber inwasion assay in vitro. Flow cytometry was used for cell cycle analysis. Reverse transcription-polymerase chain reaction (RT-PCR) was used to semi-quantify the RIP1 mRNA level. Results: The abilities of proliferation were inhibited (0.560 ± 0.023 vs 0.930 ± 0.034, P < 0.05), and typical apoptosis cellular morphology of LoVo cells was observed under the inverted microscope, but the nonspecific RIP1 siRNA and the blank control group had no effect on LoVo learn more cells. The motility and inwation of LoVo cells were inhibited significantly (21 ± 2.731 vs 43 ± 2.064), and the colorectal cancer cells penetrated polycarbonate membrane notable reduction. The nonspecific siRNA and the blank control group had no effect

on LoVo cells. The number of cells in G0∼G1 phrase increased in RIP1 siRNA group 58.28% selleck products in comparison with negative control 48.88% and blank control groups 43.82%. The nonspecific RIP1 siRNA and the blank control group had no effect on LoVo cells. In the RIP1 siRNA group, the RIP1 mRNA level was down-regulaed remarkably (P < 0.05). Conclusion: After the transfection of the RIP1-targeted siRNA into LoVo cells, the expression of RIP1 gene has been knockoutted effectively. Silencing RIP1 could regulae the malignant biological behavrors of colon cancer cell line LoVo effectively. The abilities NADPH-cytochrome-c2 reductase of proliferation and the motility and inwation of LoVo cells

were inhibited. The results shown that RIP1 gene played the important role of the proliferation and apoptosis regulation in colorectal carcinoma cells. Key Word(s): 1. RNA interference; 2. RIP1 gene; 3. Colorectal carcinoma; Presenting Author: YU-RONG WENG Additional Authors: YA-NAN YU, LIN-LIN REN, YUN CUI, YOU-YONG LV, WEIBIAO CAO, JIE HONG, JING-YUAN FANG Corresponding Author: JIE HONG, JING-YUAN FANG Affiliations: Renji Hospital, Shanghai Jiao-Tong University School of Medicine; Peking University; The Warren Alpert Medical School of Brown University & Rhode Island Hospital Objective: C9orf140 is a newly identified and characterized gene which is associated with cell proliferation and tumorigenicity. Expression of C9orf140 is upregulated in human gastric cancer and colorectal cancer (CRC); however, little is known about its role in CRC invasion. Methods: We have investigated the clinical significance, biological effects and mechanisms of C9orf140 signaling. Results: Our finding showed that knockdown of C9orf140 significantly reduced cell invasion in vitro and dramatically increased overall survival and decreased lung metastasis in vivo.

Acknowledgement: The first author would like to thank Prof Eui-H

Acknowledgement: The first author would like to thank Prof. Eui-Hong (Sam) Han and Prof. Peng Zhang for providing the ROCK and HD implementations, respectively. SEPAHAN was financially supported by Vice Chancellery for Research and Technology, Isfahan University of Medical Sciences (IUMS). We wish to thank all staff

of SEPAHAN project. Key Word(s): 1. FGID; 2. SEPAHAN project; 3. Clustering; 4. data mining; Presenting Author: PEYMAN ADIBI Additional Authors: MARJAN MANSOURIAN, HAMID REZA MARATEB, HAMED DAGHAGHZADEH, AMMAR HASSANZADEH KESHTELI Corresponding Author: HAMID REZA MARATEB Affiliations: Isfahan University of Medical Sciences; University of Isfahan;, Isfahan University of Medical Sciences; University of Alberta Objective: Functional bowel disorders (FBDs) are functional gastrointestinal disorders (FGIDs) with symptoms Ivacaftor clinical trial related to the middle or lower gastrointestinal CX-4945 tract. One of which is the IBS, in which discomfort is associated with defecation or a change in bowel habit, and with features of disordered defecation. According to the Rome III survey, a symptom-based classification is necessary for clinical diagnosis of IBS. However, in SEPAHAN project, a large-sample Iranian cross-sectional study, we

studied the feasibility of identifying the subtypes of IBS as groups (clusters) identified based on an unsupervised classification. Methods: Four-item rating scales of 37 click here selected head-questions were converted to interval data. Then a density-based clustering method was used to generate groups of people having similar symptoms. The representative of each group (cluster) was used for further clinical validation

and interpretation. Results: Three of the detected clusters (C15, C18, C23), could be classified as IBS-U, IBS-D ad IBS-C. In all of these clusters, people often had pain relieve after defecation, pain changes bowel habit, bloating and abdominal pain. However, hard and loose stools were frequent in clusters no. 18 (C18) and C23 respectively. None of these symptoms were frequent in C15, at all. Also, none of the clusters could be classified as IBS-M. People in these three clusters were also complaining of belching, fullness and dyspepsia and other frequent symptoms. Conclusion: Having used unsupervised classification, it is possible to study the groups of similar subjects e. g. subtypes of IBS. The clustering might end up with identifying new sub-groups of FGIDs. Acknowledgement: SEPAHAN was financially supported by Vice Chancellery for Research and Technology, Isfahan University of Medical Sciences (IUMS). Key Word(s): 1. IBS; 2. FGID; 3. clustering; 4.

In contrast, where increasing group size has little effect on the

In contrast, where increasing group size has little effect on the intensity of breeding competition between group members, females may form large groups whose size is ultimately limited by the effects of competition for resources on fecundity and survival (Prins, 1996; Moss & Lee, 2011). Differences find more in female group size resulting from variation in female competition affect the potential for polygyny, which in turn influences the degree of reproductive skew among males, the intensity of mating competition and the strength of sexual selection for traits that increase competitive success in

males such as body size and weapon development (Clutton-Brock, Harvey & Rudder, 1977; Clutton-Brock & Albon, 1989;

Lindenfors, Gittleman & Jones, 2007; Clutton-Brock, 2009b). An additional consequence of contrasts in female group size is that it influences the frequency of competitive interactions between males and affects the tenure and longevity of resident males (Clutton-Brock & Isvaran, 2007) with important consequences for average relatedness between group members and the genetic JNK inhibitor structure of populations (Clutton-Brock, 2009b). The intensity of female competition for breeding opportunities also affects the degree of reproductive skew among females. The medroxyprogesterone highest levels of reproductive skew in female mammals are found in singular cooperative breeders where dominant females suppress the fertility of subordinate females (Clutton-Brock et al., 2006; Clutton-Brock, 2009b,c). In these species, females can produce large litters at frequent intervals because their young are protected and fed by other group members, and variance

in breeding success is often larger in females than in males (Hauber & Lacey, 2005; Clutton-Brock et al., 2006). For example, in wild meerkats, the majority of females fail to breed while successful breeders can rear more than 80 offspring (Clutton-Brock, 2009b). Reproductive success in both sexes is closely related to whether or not individuals acquire breeding roles and their length of tenure in breeding groups; and as tenure is shorter in males than in females, standardized variance in lifetime breeding success is higher in females than males (Clutton-Brock et al., 2006). Reproductive skew can also be high in plural breeders where the rank of females affects their breeding success and the survival of their offspring, like spotted hyenas (Holekamp et al., 1996) and savannah baboons (Silk, 2009; Pusey, 2012), but it is unlikely to approach levels observed in singular cooperative breeders.

522 4(p=0 443),Alb:3 473 48g/dl(p=0 898), Prealb:17 817 4 mg/dl(p

522.4(p=0.443),Alb:3.473.48g/dl(p=0.898), Prealb:17.817.4 mg/dl(p=0.822).T. chol: 168168 mg/dl(p=0.973)In case (2). Before and 8 weeks after start taking orally of Pancrelipase; BMI: (n=14):19.919.6(p=0.420), Selleckchem GSK-3 inhibitor Alb:3.363.36g/dl(p=0.948), PreAlb(n=14):20.514.4 mg/dl(p=0.286), T. chol(n=17):163148 mg/dl (p=0.099). In case (3).Before and 8weeks after start chemotherapy; BMI:(n=77):21.120.7(p=0.0025), Alb:(n=79)3.683.52g/dl(p=0.0006), At case (1) and (3), BMI(after 8weeks)/BMI(when start) ratio is 1.00:0.98(p=0.176), alb(after 8 weeks)/alb(when start)ratio is 1.01:0.96(p=0.072). Conclusion: There are possibilitys that nutrition conditions become improvement when start

chemotherapy to patients of unresectable pancreas cancer,start taking orally of Temozolomide Pancrelipase capsules at the same time,and continue for 8 weeks. Key Word(s): 1. pancreas cancer; 2. exocrine dysfunction; 3. Pancrelipase; 4. nutritional ; Presenting Author: BEOMYONG YOON Additional Authors: HYEJIN KIM, SEWOONG HWANG, SEYOUNG PARK, SUNHYUNG KNAG, HEESEOK MOON, SEOKHYUN KIM, JAEKYU SEONG, EAUMSEOK LEE, BYUNGSEOK LEE, HYUNYONG JEONG, HEONYOUNG LEE Corresponding Author: BEOMYONG YOON Affiliations: Chungnam Nat. Univ. Hospital Objective: Introduction: Pseudomyxoma peritonei (PMP) is a clinical condition in which the abdominal cavity becomes filled with gelatinous collections

from mucinous implants. It is well recognized that PMP arises predominantly from an appendiceal mucinous neoplasm. However, PMP can occasionally arise from mucinous neoplasms of other organs, such as the ovary, colorectum, stomach, gallbladder, and pancreas. An intraductal papillary mucinous neoplasm (IPMN) is a type of neoplasm composed of mucin-producing cells that arise in the pancreatic duct. Although the clinical manifestation of an IPMN has been gradually clarified, it is not well accepted whether an IPMN is associated with PMP. Here we report a case of PMP combined with an IPMN spontaneously ruptured causing mucinous materials to spill into the free abdominal cavity. Methods: Case description: A 59-year-old man was admitted to our

hospital due to severe epigastric pain in July 2012. Computed 2-hydroxyphytanoyl-CoA lyase tomography (CT) showed cystic lesion (2.8 cm) with intracystic papillary growing lesions in pancreatic head; severe dilatation of main pancreatic duct with multiple small papillary growing lesions, suggesting IPMN (combined main and branch duct type). Results: The patient received neoadjuvant systemic chemotherapy with Gemcitabine monotherapy among planned sessions in other hospital. On October 2012, he referred to our hospital with severe abdominal pain and distension. An emergent computed tomography revealed a focal rupture of main pancreatic duct of body of pancreas with diffuse smudgelike infiltration in omentum and small amount of complicated fluid collection, suggesting peritonitis. And less than 11cm sized amorphic pseudocysts were shown in the gastropancreatic space, probably due to rupture of pancreatic duct.

These impairments occurred after as early as one week of alcohol

These impairments occurred after as early as one week of alcohol administration, when the presence of a fatty liver is first identified. Fatty liver, both non-alcoholic (NAFL) and alcoholic (AFL), affects nearly one-fourth of the U.S. population. Patients with either NAFL (seen in up to 85% of obese individuals) or AFL (seen in the majority

of alcoholics) can progress to hepatitis, fibrosis, and cirrhosis, and fatty liver is no longer considered benign. While we have established that AFL leads to impaired RME, endocytosis in NAFL has LY2835219 price not been studied. Here, we investigated RME and the changes caused in RME in rats exhibiting AFL or NAFL to see if endocytosis defects were a result of alcohol administration, or were seen in all fatty livers.

Methods: Wistar rats were fed liquid control or alcohol diet (Lieber DeCarli, 35% of BGB324 cost calories as ethanol; 35% calories as fat, groups 1 & 2), or lean or high-fat pellets (12% fat or 60% fat, respectively, groups 3 & 4). Carbohydrate (maltose/ dextrin) was similar in all groups. Animals were sacrificed after 8-10 weeks of feeding, and serum, isolated hepatocytes, and intact liver obtained for determination of serum enzymes, histology, fat content, protein content, and measurement of endo-cytosis. Results: Histologically, the AFL and NAFL rats were indistinguishable, showing fatty liver but no signs of steato-hepatitis. Serum ALT and AST were significantly increased in both the AFL

and NAFL rats, as was triglyceride content in hepatocytes and whole liver, compared to Glutathione peroxidase controls. Binding and internalization of 125I- ASOR was determined in isolated hepatocytes, and significant impairments of both processes were found in hepatocytes from alcohol fed animals compared to controls. No difference in binding or internalization, however was found in the hepatocytes isolated from the lean and high fat diets. Western Blot analysis of ASGPR and two Rab proteins known to be important in vesicle trafficking (Rab 3D and Rab 18) showed significant decreases in the AFL, but not the NAFL rats. Conclusions: Our findings suggest that the impairment in endocytosis and vesicle trafficking protein content in the ethanol fed animals is a direct result of the alcohol administration, and not a result of a fatty liver. Future studies examining the activation status of Rabs and content of Rab effector proteins in the AFL model may aid in the development of therapeutic targets. Disclosures: The following people have nothing to disclose: Karuna Rasineni, Daniel Pen-rice, Edward N. Harris, Cliff I. Stains, Jon Beck, Benita L. McVicker, Mark A. McNiven, Carol A. Casey Background: Epidemiologic data link alcoholic liver disease (ALD) to binge drinking and cigarette smoking. Previous studies showed that heavy alcohol or low-level dietary nitrosamine exposures cause steatohepatitis with insulin resistance and oxi-dative stress in experimental animals.

“Dream no small dreams for they have no power to move the

“Dream no small dreams for they have no power to move the hearts of men. Goethe I would like to thank Dr. Lindor and other members of the editorial

board of HEPATOLOGY for honoring me by asking me to contribute to the “Masters of Hepatology” series. My background is very different from that of other renowned hepatologists who have already contributed to this series. For this reason, I will go back to the beginning of my medical career to explain why I left Argentina selleck chemicals llc to continue my medical career in this country and why, once in the United States, I decided to pursue an academic career. I hope that this brief memoir will inspire potential researchers in this generation with the excitement of scientific discovery that has sustained me through my career. FHVP, free hepatic venous pressure; HVPG, hepatic venous pressure gradient; NSBB, nonselective beta blocker; VA, Veterans Administration; WHVP, wedged hepatic venous pressure. I was born in Buenos Aires, Argentina, where my father was a Jewish Hungarian immigrant and my mother was Argentinian of Jewish Hungarian descent. My father arrived in Argentina in the 1920s fleeing an already convulsed Europe. I attended public school and then studied medicine

at the University of Buenos Aires School of Medicine, which was also public and free. At the time, the best schools were public and funded by the government so tuition was free. In order to limit class sizes, EPZ-6438 research buy to matriculate at the School of Medicine, it was necessary to pass a rigorous academic exam that excluded many candidates. Nevertheless, the medical students’ classes were very large (approximately

4000 students the first year) and impersonal and did not allow for any close contact between students and professors. The intensely interesting study of medicine in Buenos Aires in the early 1960s took place against a dramatic backdrop of inescapable political unrest and violence in a country that was alternately governed by short-lived democratic government and military Non-specific serine/threonine protein kinase dictators. Dr. Bernardo Houssay, one of three Nobel Prize winners from the University of Buenos Aires, had brought the Department of Physiology at Buenos Aires School of Medicine to its highest prestige. Dr. Houssay and his collaborators had written a seminal textbook of physiology from which we studied at that time and which was also widely used in translation at U.S. medical schools. The tradition in Physiology was to teach through practical demonstrations in classical experimental models. Most experimental demonstrations were performed in a toad (Bufo arenarum Hensel, a species commonly found in Argentina). I found this part of my medical education especially riveting. From these experiences, I developed a great respect for the power of well-designed experimental models to translate science from the realm of ideas to the laboratory bench and from there to the clinical bedside.

Three weeks of DDC treatment produced the same findings despite c

Three weeks of DDC treatment produced the same findings despite clear expression of these mesenchymal proteins by other cells in the surrounding stroma (Figs. 6, 7). Therefore, we conclude that EMT does not occur in this model of biliary fibrosis characterized by bipotential progenitor cell proliferation. We report here a broad-based approach to the study of EMT in liver fibrosis. Employing a heritable marker expressed in nearly all bipotential progenitor this website cells,

cholangiocytes, and hepatocytes, we found no evidence in three different fibrosis models that epithelial cells in the adult liver undergo transition to fibrogenic myofibroblasts or other mesenchymal cells during hepatic injury. Our labeling strategy click here enabled us to bypass potential problems associated with the use of the cholangiocyte marker K19 for fate mapping. Several studies have suggested that K19 is expressed in only a subset of biliary epithelial cells.47 Tan et al.27 demonstrated in diseased human livers that K19 is absent in some keratin 7-positive biliary cells, and additionally, studies with human and rodent livers affected by biliary pathology found that a significant number of putative hepatic progenitor cells are K19-negative, even in the ducts.48, 49 Unlike K19, AFP is expressed in mice beginning at embryonic day (E)8.25-8.5. In the Alfp-Cre strain, Cre recombinase

activity is detectible by E9.5, and its efficient recombination results in labeling of more than 98% of cholangiocytes and hepatocytes (Figs. 1, 2B; Supporting

Information Fig. 1B).28.29,32 A significant advantage of this approach is the marking of K19-negative cholangiocyte progenitors. These potentially include hepatocytes, which stop expressing K19 after maturation but may transdifferentiate into cholangiocytes,19-23 and AFP-positive/K19-negative progenitor cells.24-26, 50 An important part of our study was the use of the DDC model, which permits a more complete assessment of bipotential progenitor cell fate during the ductular reaction than is possible with the CCl4 and BDL models. Bipotential progenitor cell activation is central to the “ductular reaction” that characterizes biliary Roflumilast fibrosis.43, 51 Recent studies have demonstrated a direct association between the degree of the ductular reaction and the severity of fibrosis in diseases including biliary atresia and hepatitis C.52 We have previously shown that costaining of epithelial and mesenchymal markers primarily occurs in diseases with a marked ductular reaction. Notably, in livers from patients with primary biliary cirrhosis, marker costaining was limited to epithelial cells of the ductular reaction and was not seen in mature ducts.4 Using the DDC model, in which bipotential progenitor cell expansion is associated with fibrosis, we observed no evidence of EMT.

Confocal fluorescence images were collected (excitation 488 nm, e

Confocal fluorescence images were collected (excitation 488 nm, emission 505-550 nm) at a rate of one image / 2.5 seconds in a horizontal plane through

the hepatocytes to visualize the canalicular spaces. After establishing a baseline level of fluorescence, 1 μM CGamF was perfused through the chamber Y-27632 and images were collected for 10 minutes. CGamF is a bile acid conjugate that relies on basolateral uptake before being transported across the canalicular membrane into the sealed canalicular vacuole of cultured hepatocytes.24, 25 The increase in fluorescence intensity in the canalicular space over time relative to baseline fluorescence served as a measure of Bsep activity. For LPS-induced cholestasis, rats were anesthetized with isoflurane and injected intravenously with 1 mg/kg LPS, or 0.9% saline as control. After 16 hours, animals were anesthetized with pentobarbital sodium Roxadustat nmr (50 mg/kg) and their livers were harvested and snap-frozen in liquid nitrogen for further analysis.26 For estrogen-induced cholestasis, 17 alpha-ethinylestradiol (EE) dissolved in 1,2-propanediol (5 mg/mL) was administered to rats subcutaneously (5 mg/kg/day) for 5 consecutive days as

described.26 Control animals received equivalent amounts of vehicle alone. After 5 days animals were anesthetized and livers harvested as above. Hepatocytes on glass coverslips were washed and then fixed in ice-cold methanol for 5 minutes; rat liver was snap-frozen DOK2 under liquid nitrogen, cryosectioned, and then fixed in 4% paraformaldehyde for 10 minutes. Samples were then permeabilized with 1% Triton X-100, blocked in 2% bovine serum albumin, incubated with primary antibodies for 1 hour at room temperature, washed with phosphate-buffered saline (PBS), incubated with fluorophore-conjugated secondary antibodies for 1 hour at room temperature, washed again with PBS, and then mounted. Negative controls were incubated with secondary antibodies alone. Double- and triple-labeled specimens were examined with a Zeiss LSM 510 Confocal Microscope (Thornwood, NY).23 Liver was homogenized in protein extraction reagent from

Thermo (Rockford, IL). Protein was extracted from isolated hepatocytes in the same buffer. Proteins were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) electrophoresis on a 4%-20% gel. After transferring to PVDF using a semidry system, the membrane was blocked, incubated with primary antibodies overnight at 4°C, washed, incubated with horseradish peroxidase conjugated secondary antibodies for 1 hour, washed, and then revealed by enhanced chemiluminescence.27 Values listed are mean ± standard error of the mean (SEM). Comparisons were made using Student’s t test or analysis of variance (ANOVA). P < 0.05 was considered significant. Like hepatocytes in intact rat liver, hepatocytes in collagen sandwich culture have a well-developed canalicular network to which canalicular membrane proteins appropriately traffic.