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Preliminary Results of BÂRRX Ablation Trial in Patients with Non-Dysplastic Intestinal Metaplasia Versus Low Or High Grade Dysplasia Franklin Tsai, Mehrdad Vosoghi, Reza Khoshini, Sonya Reicher, David Chung, Samuel French, Viktor Eysselein Aims: Assess the response associated with BÂRRX ablation of Barrett’s esophagus (BE) in patients with no dysplasia (ND) vs low grade dysplasia (LGD) or high grade dysplasia (HGD), as well as short segment (SSBE, !3 cm) vs long segment (LSBE, O3 cm). Methods: Prospective trial in patients with BE confirmed by bx on prior EGD. Patients receive 2 ablations at 10 J/cm in the ND group or 12 J/cm in the LGD group during each treatment. Tolerability is recorded using a 7-day, 10-pt scale patient symptom diary. All patients receive lansoprazole 30 mg bid and have 3 & 12 mo follow-up EGD with 4-quadrant bx every 1 cm from original BE region. Any patients with all bx negative are deemed to have complete response (CR). Patients with intestinal metaplasia present on any bx or incomplete response (IR) return at 6 mo for repeat treatment. Patients with known HGD who are not surgical candidates are evaluated with EUS to rule out invasion. Those with HGD limited to the mucosa will undergo EMR if not already done, then return in 2 mo to proceed with the BÂRRX ablation at 12 J/cm. Results: 11 of 75 planned patients (5 male, mean age 57.6, 9 ND, 1 LGD, 1 HGD, 8 LSBE, mean length 5.7 cm, range 1 to 11 cm, 3 SSBE) have been treated. One patient refused follow-up EGD & dropped out of the study. 3-mo follow-up EGD is pending for 4 patients. Amongst the 6 patients (5 ND, 1 HGD, 4 LSBE) who have already returned for 3-mo follow-up EGD, all had complete healing with no evidence of BE on EGD, except 2 patients with !2 cm patches of residual BE. In the ND group, all 4 LSBE patients had IR while the one SSBE patient had CR. The patient with HGD & SSBE had CR after one treatment. The mean percentage of negative bx at 3 mo was 54% (range 20 to 88%) for LSBE versus 100% for SSBE, with a mean of 69.3% for all 6 patients. Amongst all 11 treated patients, the procedure was better tolerated by those with SSBE (mean 2/10 symptom score) vs LSBE (6.6/10). The most frequently reported symptoms were odynophagia & nausea. There were no strictures, perforations, or other serious complications. Conclusions: Preliminary data suggests that BÂRRX ablation is safe, well-tolerated, & effective at clearing SSBE (!3 cm). Our first patient with HGD had CR with ablation at 12 J/cm. Data are pending for patients with LGD. LSBE was most associated with IR, but given the high percentage of negative bx (54%), we expect significant rates of CR after these patients return for repeat ablation.

Background BÂRRX Medical has developed a promising new balloon-based electrode device for ablation of Barrett9s esophagus (BE) using rapid circumferential radiofrequency energy. Aims To assess the safety, tolerability, and response associated with BÂRRX ablation of short-segment (SSBE) versus long-segment Barrett9s esophagus (LSBE). Methods Prospective trial in patients followed at Harbor-UCLA Medical Center with non-dysplastic intestinal metaplasia confirmed by biopsy on prior EGD within the last year. Patients are assigned to either the SSBE ( Results 7 of 50 planned patients (4 male, mean age 53.4, 5 with LSBE, mean length 5.7 cm, range 2-10 cm) have been enrolled and treated. One patient refused follow-up EGD and dropped out of the study. Three-month follow-up EGD is pending for two patients. Among the four patients (1 SSBE and 3 LSBE) who have already returned for 3-month follow-up EGD, none had endoscopic evidence of residual BE, except for one patient with LSBE (5 cm). Only the patient with SSBE (2 cm) had CR, while all three of the patients with LSBE had IR. The percentage of negative biopsies at 3 months after one treatment was 100% for the SSBE group and 61% for the LSBE group, with a mean of 71% for all four patients. Among all seven patients enrolled, the procedure was better tolerated by the patients with SSBE (mean 2/10 symptom score) versus patients with LSBE (mean 7/10 symptom score). The most frequently reported post-procedure symptoms were odynophagia and nausea. There were no strictures, perforations, or other serious complications. Conclusions Preliminary data suggest that BÂRRX ablation is safe, well tolerated, and effective at clearing SSBE (

The Utility of Immunohistochemical Staining with CEA, CA19-9, P53, and MIB to Improve the Diagnostic Yield of Endoscopic Ultrasonography Guided Fine Needle Pancreatic Aspirates Alia Dadabhai, Stephen L. Chen, Steve Tsai, Rose Venegas, Samuel French, Sartaj Arora, Mehrdad Vasdoghi, Ben Garrett, David Chung, Shahid H. Sial, Viktor Eysselein In a study of 100 specimens obtained by EUS-FNA, P53, MIB, CA19-9 and CEA were used in an analysis of 71 samples to determine if tumor markers increased the diagnostic yield of cytologically atypical or benign samples. A prospective analysis was performed of all patients undergoing EUS-FNA of pancreatic lesions. Methods: Four antibodies were used for immunohistochemical labeling: p53 (DO-7)Zymed and MIB (Ki-67)Zymed, CEA (monoclonal ab) – Zymed and CA19-9. Cytology smears and cell blocks were reviewed by two cytopathologists who were blinded to the tumor markers and the final diagnoses. The specimens were categorized as tumor marker positive if there was O 5% nuclear staining by MIB ab, and if there was positive labeling for p53, CEA and CA19-9 in one or more cells. Follow-up data for each patient was determined by surgical pathology or clinical follow-up. Results: In 100 consecutive pancreatic EUS-FNAs, an adequate amount of aspirate was obtained for cytology smear in 89% (89/100) of patients, and for immunohistochemical labeling in 71% (71/100) of specimens. The cytology smears showed 24 benign lesions, 23 malignant lesions, 24 with atypical cells, with 11 hypocellular aspirates. Of the adequate cytology smears were 34% (24/71) were atypical. 58% of the atypical aspirates (14/24), 29% (7/24) of the benign aspirates, and 100% of the malignant smears, were malignant on clinical or surgical follow-up. Combined tumor markers and cytology increased the sensitivity of aspirates to 89% and increased the number of patients detected with malignant disease from 23 to 39 out of the total 44 patients with cancer. Sensitivity of cytologic diagnosis alone was 52%. This sensitivity increased to 89% when combined tumor markers were used. The specificities of the individual tumor markers ranged from 80-100%. Conclusions: The addition of immunohistochemical labeling with p53, MIB, CEA and CA19-9 to cytology enhances the diagnostic yield and sensitivity for finding pancreatic malignancy by EUS-FNA by 31%. Immunohistochemical labeling may be useful to detect well-differentiated tumors when diagnosis by cytology is nonconclusive or benign and may reduce the need for repeat EUS-FNA.

For polysiloxanes to be used as a protective coating it is important that proton transfer, a trigger to polymerisation, is a facile process. Here we investigate the initial stages of polycondensation and compare different silanol tail groups and the effect of solvent (isopropanol). In the case of (3-aminopropyl)trihydroxysilane we see the potential for self catalysis as the tail group is a proton acceptor, while thiolpropyltrihydroxysilane and isopropanol do not promote proton transfer.

The ethanol inducible isoform of cytochrome P450, CYP2E1, may play a role in ethanol-induced liver injury. Therefore, the factors which govern CYP2E1 degradation and turnover were investigated. These factors include cAMP, ubiquitin, proteasomal enzymes and CYP2E1 mRNA. Rats fed ethanol or pair-fed isocaloric dextrose were pair-fed with rats fed ethanol or dextrose treated with cAMP for 2 months. The liver pathology, regenerative activity, fatty acid composition, NFkappaB activation, ubiquitin conjugates and proteasomal enzymes were measured as were the apoprotein levels of CYP2E1, CYP3A, CYP4A and mRNA levels for CYP2E1 and ubiquitin expression. The results showed, that the cAMP treatment ameliorated the increase liver fat storage and changes in the fatty acid composition in the livers of ethanol fed rats. Other histologic features of alcoholic liver disease were not changed. Western blot quantitation showed that the amount of ubiquitin and ubiquitin conjugates were markedly reduced by ethanol treatment. Similarly, ethanol decreased the level of ubiquitin mRNA. cAMP ameliorated the inhibition of the proteasomal enzyme proteolysis caused by ethanol feeding. The ethanol-induced increase in the CYP2E1 protein was partially inhibited by cAMP treatment. cAMP treatment decreased CYP2E1 mRNA levels in both ethanol-fed and pair fed control rats. Likewise NFkappaB activation was not increased by ethanol but cAMP reduced the level of NFkappaB activation. CAMP treatment also reduced CYP4A but not CYP3A. The results support the concept that cAMP treatment partially protects the liver from ethanol-induced fatty liver by reducing CYP2E1 induction through cAMP's effects on CYP2E1 synthesis.

Monooxygenase enzymatic activities were measured in liver and kidney microsomes of control and ethanol-treated rats. Animals were administered alcohol by using a model for alcoholic liver injury. Several in vitro approaches were used to compare the laurate metabolism in liver and kidney microsomes: correlation studies between specific P-450 catalytic activities, immunoblot analysis, and chemical and immunoinhibitions. Ethanol treatment increased the liver and renal hydroxylations of chlorzoxazone and 4-nitrophenol. Moreover, lauric acid (ω-1)-hydroxylation was found to be significantly increased (∼6-fold) after ethanol treatment in liver, but not in kidney microsomes. The laurate ω-1/ω ratio increased from 1.52 ± 0.49 to 4.11 ± 1.01 in liver microsomes of control and ethanol-treated rats, and from 0.29 ± 0.06 to 0.44 < 0.07 in kidney microsomes. Immunoblot analysis using polyclonal anti-cytochrome P-450 (CYP) 2E1 or CYP4A antibodies showed an increase of CYP2E1 and CYP4A contents in both organs, but the increase was higher in liver than in kidney microsomes. Chemical inhibitions using CYP2E1 competitive inhibitors (such as chlorzoxazone and ethanol) led to a nonsignificant inhibition of the renal (ω-1)-hydroxylation of lauric acid. In contrast, 17-octadecynoic acid (a mechanism-based inhibitor of ω-hydroxylase) was able to inhibit both ω- and (ω-1)-hydroxylations of lauric acid in kidney microsomes. Immunoinhibitions specific to CYP2E1 significantly decreased the (ω-1)-hydroxylation of lauric acid in liver, but not in kidney microsomes, whereas the polyclonal anti-CYP4A1 antibody inhibited ω- and (ω-1)-hydroxylations of lauric acid in kidney microsomes. All of these results show that lauric acid hydroxylations in liver and kidney respond in different manners to ethanol treatment. Lauric acid (ω-1)-hydroxylation, a highly specific probe for CYP2E1 in rat and human liver microsomes, is mediated by a CYP4A isoform in rat kidney microsomes.

A new method of visualizing the three-dimensional architecture of the cytokeratin filaments of the intact rat hepatocyte in situ has been achieved. Frozen sections of liver cut 10 micron thick were serially extracted to remove all elements of the cells except the intermediate filaments. Parallel sections were stained with monoclonal antibodies to the two main cytokeratins found in bile duct and liver cells. Immunofluorescent antibody and immunogold electron microscopy techniques were used to identify the proteins morphologically. Several new observations resulted from these studies. The pericanalicular sheath of intermediate filaments was visualized using steropairs as an uninterrupted branching tubular structure composed of cytokeratins located in the cell cortex of adjacent hepatocytes. Intermediate filaments in the cell cortex formed a distinct sheet of matted filaments which enveloped the entire hepatocyte. The cortical intermediate filaments were in continuity with the pericanalicular sheath and the filaments located within the cytoplasm. The intermediate filaments are attached to the centrioles and appeared to tent the nuclear lamina-pore complex at points of contact. Monoclonal antibodies to rat liver intermediate filament cytokeratins (CK49 and CK55) each stained intermediate filaments located in the cell cortex, within the cytoplasm and at the nucleus. By immunogold staining, some of the intermediate filament filaments were shown to contain both cytokeratins. Filaments which did not stain were thought to be either actin at the cell periphery or nuclear lamins around the nucleus. It is concluded that the cytokeratins form a specialized framework for the cell cortex, canaliculus, centrioles and the nucleus of hepatocytes. The filaments run continuously throughout the cytoplasm without terminating.

Mouse and human extracted liver tissue were examined by indirect immunofluorescent staining and transmission electron microscopy in order to study the alteration of cytokeratin intermediate filaments associated with Mallory body formation. Frozen sections of griseofulvin-fed mouse liver and human liver of primary biliary cirrhosis and primary sclerosing cholangitis were extracted by Triton X-100 and nuclease. Indirect immunofluorescent staining was performed by using anticow hoof keratin antibody for mouse liver and anti-human epidermal keratin antibody (AE1 and AE3) for human liver. Transmission electron microscopy was also performed on extracted and critical point-dried samples. The griseofulvin-fed mouse hepatoma cells showed four different types of altered staining pattern based on the indirect immunofluorescent staining of the cytoplasm and Mallory bodies: Type I—cytoplasm(+), Mallory body(−); Type II—cytoplasm(+), Mallory body(+); Type III—cytoplasm(−), Mallory body(+), and Type IV—cytoplasm(−), Mallory body(−). Types I and III were predominant, however, some hepatoma cells which contain Mallory bodies revealed bright cytoplasmic staining (Type II). The nuclear rims were strongly stained. In human liver, AE1 stained Mallory bodies and the bile duct epithelium intensely, but did not stain normal hepatocytes. AE3 mainly stained Mallory bodies and normal hepatocytes, but also stained bile duct epithelium weakly. Indirect immunofluorescent staining for human liver showed the same staining patterns as found in mouse liver, except that Type IV was not observed. Although many hepatocytes which contained Mallory bodies did not react with either of these two antibodies (Type III), some of the hepatocytes were stained, not only with AE3, but also with AE1 (Type II). Transmission electron microscopy revealed that Mallory bodies were often surrounded by numerous intermediate filaments, which is equivalent to the Type II pattern in indirect immunofluorescent staining. This was true of mouse liver frozen sections and primary cultures, as well as human liver studied. The results clearly show that cells associated with Mallory bodies show changes in the expression of cytokeratins as well as rearrangement of intermediate filaments within the cell. Hepatocytes forming Mallory bodies often contain a variable density of intermediate filaments which fail to stain with cytokeratin antibodies, indicating that the antigenic determinants of the intermediate filaments were lost or masked.

The effect of ethanol withdrawal on the cAMP response of cerebral cortical brain slices was studied. The cAMP response was evoked in vitro by various neurotransmitters including norepinephrine (NE), histamine, serotonin, dopamine, acetylcholine, and γ-aminobutyric acid (GABA). The cAMP response to NE and histamine was enhanced by ethanol withdrawal. Serotonin evoked a cAMP response in the brain slices from ethanol-withdrawal rats but not in pair-fed controls. The histamine and serotonin evoked responses were blocked by chlortripolon and methysergide, respectively. The responses to histamine and serotonin were also blocked by a- and β-adrenergic antagonists, possibly because of the nonspecific membrane stabilizing effect of these antagonists. GABA inhibited the NE stimulated cAMP response possibly through the hyperpolarizing action of GABA. The results support the hypothesis that ethanol withdrawal induces a nonspecific postjunctional supersensitivity. It is postulated that the supersensitivity involves a partial depolarization of the receptor membrane. Alternative hypotheses are reviewed.

The uptake, transport, and secretion processes of hepatocytes probably depends on a contractile system of fibrous proteins similar to those found in other cells. However, the contractile system in in situ liver cells has not been thoroughly investigated. In this report, a network of actin-like filaments was identified in liver cells using the heavy meromyosin binding technique. Actin-like filaments were found in the ectoplasm of the liver cells. These filaments appeared more numerous and thickened when they bound heavy meromyosin. Although they were found over the entire cell periphery they were conspicuous where the plasma membrane faced the space of Disse or the bile canaliculus. Gaps in the filament network were encountered. The filaments appeared to have attachment points to the plasma membrane and the organelles. It is postulated that the actin-like filament network attached to the plasma membrane could provide the motive force for endocytosis, transport of vesicles, and flow of bile.

We investigated the possibility that organs other than liver in women were more susceptible to alcohol than in men. Female:male cirrhosis mortality ratio in different countries was used as an index of female susceptibility to alcohol. The F:M cirrhosis ratio correlated significantly with F:M ratios for cerebrovascular disease and cancers of the esophagus, liver and lung. This preliminary evidence suggests that other organs and/or disease processes in women may be affected more by alcohol than in men.

A case of multiple (59) gastric polyps in a patient with pernicious anemia is reported. Examination of the stomach revealed hyperplasia of Kulchitsky cells, as well as frank carcinoid neoplasia with regional lymph node metastasis. Results of light and electron microscopy, immunofluorescence and histochemistry are presented, and the possible pathogenetic role of chronic hypergastrinism is discussed.

The clinical and histologic significance of cytokeratin antigen expression in various intrahepatic locations was assessed in 57 patients with alcoholic liver disease as part of a large Veterans Administration Cooperative Study of Alcoholic Hepatitis. Cytokeratin antigen was demonstrated in fixed, paraffin-embedded liver tissue by an avidin-biotin peroxidase method using a mixture of two different monoclonal antibodies, AE1 (acidic; 48, 50 and 56.5 kD) and AE3 (basic; 52, 56, 58 and 65 to 67 kD). In contrast to the normal liver, in which only bile duct epithelium was positive, this antibody mixture stained both bile ducts and hepatocytes in pathologic livers. Serum levels of vitamin A showed a significant inverse correlation with the amount of cytokeratin antigen (scale: 0 to 3) in hepatocytes without Mallory bodies (p = 0.001), in Mallory body-containing hepatocytes (p < 0.0001) and in bile ducts (p = 0.0074). Increased amount of cytokeratin antigen in each of these locations, in turn, correlated directly with the histologic severity of the liver disease. Histologic severity (fibrosis, parenchymal degeneration/necrosis, hepatocyte regeneration and inflammation) was significantly higher in patients when either Mallory bodies (p < 0.0001) or cytokeratin antigen (p = 0.0021) was present in hepatocytes. Demonstration of cytokeratin antigen in hepatocytes which contained Mallory bodies correlated positively (p = 0.03) with clinical severity of the liver disease as determined by high serum bilirubin and prolonged prothrombin time (Maddrey's discriminant function). This study suggests that (i) vitamin A deficiency may be a triggering factor in aggregation of cytokeratin filaments, and (ii) the degree of aggregation of cytokeratin may determine the histologic severity and thereby the prognosis of patients with alcoholic liver disease.

The case of a patient who had renal cell carcinoma with cytoplasmic eosinophilic globular hyaline bodies antigenically similar to Mallory bodies, highly suggestive of globular type III alcoholic hyalin, is presented. Mallory bodies are not found exclusively in the liver, having also been demonstrated in the lung. The renal tumor in this case represents an additional extrahepatic site.

Blood alcohol levels (BAL) were maintained at high levels (overall mean +/- S.D. achieved in 14 alcoholic rats was 216.0 +/- 120.1 mg%) in male Wistar rats for 15 to 85 days by continuous intragastric infusion of ethanol and nutritionally defined low fat liquid diet. The ethanol intake was progressively increased from 32% of total calories up to 41.4% in order to maintain high BAL. Pair-fed animals received isocaloric glucose solution and the liquid diet. Despite the low level of dietary fat (4.9% of total calories), histopathological evaluation of the liver revealed severe and progressive fatty infiltration in the alcoholic rats. In addition, following 30 days of intoxication, one third of the animals showed focal necrosis with mononuclear cell infiltration in centrilobular areas of the livers. This was correlated with the markedly elevated levels of SGOT and SGPT in these animals. Pair-fed controls showed no abnormality in the morphology of liver or blood chemistry. Chemical quantitation of liver triglycerides confirmed the histological observation, with triglyceride levels of 61.51 +/- 16.45 and 89.61 +/- 5.94 mg per gm at 30 and 85 days, respectively. Most importantly, the degree of steatosis was tightly and significantly correlated with the mean BAL achieved (r = 0.80, p less than 0.001). These data represent the first confirmation of the hypothesis that continuously high BAL correlate with the severity of alcohol-induced liver pathology.

Previous work in our laboratory has shown that chronic ethanol ingestion induced a change in rat liver mitochondria which we termed increased fragility. This change was detected biochemically by measuring an increase in oxygen consumption during the succinic dehydrogenase assay. It was indicated ultrastructurally by swelling of the matrix compartment and loss of the outer membrane. This change was postulated to represent an acceleration of the aging process resulting from an increase in the rate of membrane phospholipid hydrolysis. In this study, the biochemical and ultrastructural changes caused by exogenous phospholipase, oleate, and lysolecithin were studied using normal liver mitochondria. The protective effects of heparin and albumin on mitochondria exposed to these agents were also studied. As was the case with the ethanol-induced fragility phenomenon, the phospholipase A induced fragility was prevented by heparin. Neither heparin nor albumin protected the mitochondria by inhibiting phospholipid hydrolysis. The effects of exogenous oleate simulated those of phospholipase A but only albumin protected the mitochondria from oleate. The evidence supports the hypothesis that the loss of mitochondrial phospholipid by hydrolysis may be the cause of the ethanol-induced fragility phenomenon and that albumin may prevent fragility by binding released fatty acids. Heparin prevents fragility by still unknown means. Studies with high amplitude phosphate swelling and contraction of the inner membrane and matrix indicated that the mitochondrial fragility phenomenon was partially reversible.

SummaryThe concentrations of ATP in rat liver were determined at 2, 8, and 24 hours after stomach-tube feeding of ethanol. ATP concentration nearly doubled 8 hours after feeding but returned to control levels by 24 hours. This acute effect of ethanol ingestion was not sex dependent. When male rats were fed ethanol for 2 months or longer, the liver ATP content decreased. This depression in ATP persisted for as long as 16 months but returned to control levels 2 days after ethanol was withdrawn. These results of chronic ethanol feeding were found only in the experiments with male rats, not females. The results with males were duplicated by chronic acetaldehyde feeding.

The chorioallantoic membranes and embryos of developing hen's eggs infected with a strain variant canary pox virus were examined histologically in an effort to determine the cause of death of the embryo. The presence of a markedly thickened chorioallantois, in which the ectoderm and mesoderm are principally involved, and the conspicuous simultaneous absence of specific embryo degenerative or hyperplastic changes suggest that the embryo dies of hypoxia resulting from impaired oxygen diffusion through the thickened membrane. The evidence supporting this hypothesis is discussed.

Summary Changes in the liver mitochondria of rats after chronic ethanol feeding were studied by manometric assay of succinic dehydrogenase (SD) activity and by electron microscopy of isolated liver mitochondrial fractions. Abnormalities were not detectable until the assay of SD activity had been allowed to progress for 30 min. By 60 min of assay, an increase in permeability to phenazine methosulf ate was demonstrated. This effect was obliterated by eliminating the membrane barrier to the phenazine by adding calcium to the assay media. The electron micrographs of assayed mitochondria showed that the chronic ethanol feeding induced an increase in the number of mitochondria which had lost their outer membranes and cristae, compared with the mitochondria in pair-fed control animals. This loss of outer membranes was correlated with the loss of the permeability barrier to phenazine methosulf ate, probably because the SD, located in the inner mitochondrial membrane, was made more accessible to the phenazine. Indirect evidence indicated that neither microsomal contamination nor an increase of lysosomes in the mitochondrial fraction accounted for the ethanol-induced mitochondrial alteration in the ethanol-fed rat liver. In the case of microsomal contamination, purification of the mitochondrial fraction did not alter the ethanol effect. In the case of lysosomal contamination of the mitochondria, no increase in acid phosphatase was observed in the mitochondrial fractions obtained from the ethanol-fed rats. It is concluded that the fragility of mitochondria induced by ethanol feeding is probably due to an abnormality in the mitochondrial membrane.

Non-membrane bound cytoplasmic crystalline inclusions were found in the liver cells of two alcoholics. The inclusions varied in size, shape, and internal structure. They were found in cells which also contained mitochondria with crystalline inclusions. The cytoplasmic crystals found in alcoholic liver cells are similar to inclusions previously observed in liver and kidney cells under a variety of conditions. Morphological evidence suggests that the alcoholic inclusions are composed of phospholipid and/or protein. The significance of these crystalline inclusions is not yet apparent.

Physiologic and anatomic changes following unilateral pulmonary irradiation up to 6,000 rads in dogs were studied. Fractionate irradiation was given to the right lung of 11 dogs divided into three groups which received 2,000, 4,000 and 6,000 rads respectively. Differential bronchospirometry was serially performed at approximately four week intervals for one year. There was no significant change in the relative distribution of ventilation and oxygen uptake between the irradiated and the contralateral control lungs. Upon diversion of the entire cardiac output through the irradiated lung by temporary unilateral pulmonary artery occlusion, pulmonary artery pressure and left atrial pressure remained within normal limits. Thus, there was no suggestion of increased pulmonary vascular resistance as a consequence of irradiation. Neither adhesions nor abnormalities of the bronchial vessels were observed. The microscopic findings consisted of focal organized pneumonia which was found only in the 4,000 and 6,000 rads groups, and mild focal interstitial fibrosis plus focal vascular sclerosis found in representatives of each of the three groups. We conclude that no significant physiologic abnormalities following unilateral pulmonary irradiation up to 6,000 rads in dogs were demonstrated.

Pluripotent stem cells are being actively studied as a cell source for regenerating damaged liver. For long-term survival of engrafting cells in the body, not only do the cells have to execute liver-specific function but also withstand the physical strains and invading pathogens. The cellular innate immune system orchestrated by the interferon (IFN) pathway provides the first line of defense against pathogens. The objective of this study is to assess the innate immune function as well as to systematically profile the IFN-induced genes during hepatic differentiation of pluripotent stem cells. To address this objective, we derived endodermal cells (day 5 post-differentiation), hepatoblast (day 15) and hepatocyte-like cells (day 21) from human embryonic stem cells (hESCs). Day 5, 15 and 21 cells were stimulated with IFN-α and subjected to IFN pathway analysis. Transcriptome analysis was carried out by RNA sequencing. The results showed that the IFN-α treatment activated STAT–JAK pathway in differentiating cells. Transcriptome analysis indicated stage specific expression of classical and non-classical IFN-stimulated genes (ISGs). Subsequent validation confirmed the expression of novel ISGs including RASGRP3, CLMP and TRANK1 by differentiated hepatic cells upon IFN treatment. Hepatitis C virus replication in hESC-derived hepatic cells induced the expression of ISGs — LAMP3, ETV7, RASGRP3, and TRANK1. The hESC-derived hepatic cells contain intact innate system and can recognize invading pathogens. Besides assessing the tissue-specific functions for cell therapy applications, it may also be important to test the innate immune function of engrafting cells to ensure adequate defense against infections and improve graft survival.

We have previously demonstrated that quercetin, a bioflavonoid, blocks hepatitis C virus (HCV) proliferation by inhibiting NS5A-driven internal ribosomal entry site (IRES)-mediated translation of the viral genome. Here, we investigate the mechanisms of antiviral activity of quercetin and six additional bioflavonoids. We demonstrate that catechin, naringenin, and quercetin possess significant antiviral activity, with no associated cytotoxicity. Infectious virion secretion was not significantly altered by these bioflavonoids. Catechin and naringenin demonstrated stronger inhibition of infectious virion assembly compared to quercetin. Quercetin markedly blocked viral translation whereas catechin and naringenin demonstrated mild activity. Similarly quercetin completely blocked NS5A-augmented IRES-mediated translation in an IRES reporter assay, whereas catechin and naringenin had only a mild effect. Moreover, quercetin differentially inhibited HSP70 induction compared to catechin and naringenin. Thus, the antiviral activity of these bioflavonoids is mediated through different mechanisms. Therefore combination of these bioflavonoids may act synergistically against HCV.

We previously identified HSP70 and HSC70 in complex with NS5A in a proteomic screen. Here, coimmunoprecipitation studies confirmed NS5A/HSC70 complex formation during infection, and immunofluorescence studies showed NS5A and HSC70 to colocalize. Unlike HSP70, HSC70 knockdown did not decrease viral protein levels. Rather, intracellular infectious virion assembly was significantly impaired by HSC70 knockdown. We also discovered that both HSC70 nucleotide binding and substrate binding domains directly bind NS5A whereas only the HSP70 nucleotide binding domain does. Knockdown of both HSC70 and HSP70 demonstrated an additive reduction in virus production. This data suggests that HSC70 and HSP70 play discrete roles in the viral life cycle. Investigation of these different functions may facilitate developing of novel strategies that target host proteins to treat HCV infection.

Mortality from cirrhosis in many countries deviates markedly from that expected for a given per capita alcohol intake. Since lipid peroxidation is thought to be one of the mechanisms involved in the pathogenesis of liver disease and since uric acid is a natural antioxidant, we investigated the possibility that deviation in mortality rates from cirrhosis in different countries might be related to serum uric acid levels in the populations studied. Deviations from expected cirrhosis mortality were calculated as a percentage and correlated with the serum uric acid levels in nine countries. Since animal protein is a major source of dietary purines, the percentage deviation was also correlated with animal protein intake (g/capita/day) for 15 countries. Significant Pearson r correlations were obtained between the percentage deviation in cirrhosis mortality and serum uric acid (-.70, p less than .05) and animal protein ingestion (-.67, p less than .02). This suggests that higher levels of serum uric acid or higher intake of protein may protect against the development of cirrhosis.

A case of globular amyloid in the liver is reported. The amyloid nature of the unusual deposits was established by special stains and electron microscopy. The significance of globular amyloid in the liver is unknown. Of the three cases reported in the world literature, none was clinically manifest. Two occurred in patients who had diabetes mellitus.

Among 50 postoperative stomachs examined endoscopically, there were 13 cases showing a pattern of linear erythematous streaks radiating from the stoma. Biopsy sections obtained under direct vision from the erythematous streaks disclosed more significant histologic changes than those taken from the intervening pale areas. The finding of reparative changes was the most reliable criterion for differentiating the degree of change in the 2 areas. The topography of gastritis in the postoperative stomach suggests that mechanical factors may be important in the pathogenesis of reflux gastritis.

Evidence has been accumulating that Mallory body (MB) is a heritable phenotype of hepatocellular carcinoma (HCC) and is closely related to hepatocarcinogenesis in some carcinogen-fed animals. This prompted us to try to evaluate the MB phenomenon in human HCCs, in which its significance remains unclear to date. Whole liver slices were obtained from 28 autopsied livers in which MB-positive HCC was found, and distribution of MBs within the HCC was examined. There were three distribution patterns: clustering, diffuse, and sparse types. The clustering type was the most frequent and was found in 21 of the 28 cases. In this type, there was a formation of groups of the MB-containing cells within the HCC, and the cluster of MB-positive cells was variable in number, size, and shape. There was often a sharp border line between MB-positive clusters and the surrounding MB-negative HCC cell areas where a collision line could occasionally be seen. Some of the MB-containing HCC cells were positive for γ-glutamyl transpeptidase and α-fetoprotein. These data led the authors to speculate that MB-positive HCC cells in human livers are capable of proliferation to form the clusters and that therefore, MBs are expressed as a kind of heritable phenotype in such clusters.

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An improved methodology is described for long-term venous and gastric cannulation in the rat. The long-term efficacy of the cannulas for blood sampling and intragastric infusion of liquid diet was determined in 18 rats. No animals died after surgery, and weight gain was normal. During the first 6 wk, blood could be drawn repetitively in 94% of the rats, and continuous intragastric infusion of diet was maintained in all the animals. For an extended period of 11 wk, success rates for blood sampling and dietary infusion were 75 and 100%, respectively. Standard hematological, histopathological, and clinical laboratory tests showed no abnormal changes. Furthermore, plasma corticosterone levels were consistently low (0.5 +/- 0.11 to 1.52 +/- 0.55 micrograms/dl) from the third postoperative day throughout a period of 6 wk, indicating that animals were under minimal stress.