Efficacy and safety of gastric per-oral endoscopic myotomy (GPOEM) in lung transplant patients with refractory gastroparesis: a systematic review and meta-analysis.

BACKGROUND: Post-lung transplant gastroparesis is a frequent debilitating complication of lung transplant recipients, as it can increase the risk for gastro-esophageal reflux disease and subsequent graft dysfunction. We performed a systematic review and meta-analysis to evaluate the efficacy and safety of GPOEM in lung transplant patients with refractory gastroparesis. METHODS: The present systematic review and meta-analysis wer performed according to the Preferred Reporting Items for Systematic reviews and Meta-Analyses guidelines. We selected studies that analyzed the gastroparesis cardinal symptom index (GCSI) before and after the procedure to verify the efficacy of GPOEM. Random-effects model was used and the analysis was performed with STATA 17. RESULTS: Four observational studies (one conference abstract) with 104 patients were included in the meta-analysis. Prior treatments for gastroparesis included prokinetic agents and botulinum toxin in 78% (78/104) and 66.7% (66/99), respectively. Pooled estimate for clinical efficacy of GPOEM was 83% (95% CI 76%-90%). The pooled mean reduction in GCSI following the procedure was - 2.01 (- 2.35, - 1.65, p = 0.014). Three studies reported statistically significant improvement of gastro-esophageal retention or emptying in the post-GPOEM period. 30-day post-operative complications included minor or major bleeding (11.6%), severe reflux (1.2%), and pyloric stenosis (1.2%) requiring re-intervention. 90-day all-cause mortality was 2.6% with one patient dying from severe allograft rejection. CONCLUSION: Our study showed that GPOEM is an effective and safe strategy for lung transplant patients with refractory gastroparesis and should be considered as a therapeutic strategy in this population. Larger multicenter trials are needed in the future to further evaluate the effect of GPOEM on allograft function and rates of rejection.

Impaired Regulatory Volume Decrease and Characterization of Underlying Volume-Activated Currents in Cystic Fibrosis Human Cholangiocyte Cell Line.

The volume-activated chloride channel (VACC) serves vital cellular functions in secretion and cell volume regulation via regulatory volume decrease (RVD) in various epithelia. Previously, we have shown that RVD in primary CF mouse cholangiocytes is impaired. Thus, the effect of CFTR defect on VACC and RVD in CF human immortalized cholangiocyte cell (HBDC) was examined in comparison with those in normal HBDC by using cell volume measurement and whole-cell patch clamp techniques, respectively. The CF HBDC had an impaired RVD, which was not further inhibited by removing the extracellular calcium or administering BAPTA-AM, NPPB, or DIDS. When exposed to a hypotonic solution, CF HBDC exhibited large, outwardly rectified currents with time-dependent inactivation at a positive potential. The amplitude of the outward currents was about three times that of the inward currents. The amplitude and reversal potential of VACC was dependent on chloride concentration. The VACC was significantly inhibited by replacing chloride with gluconate, glutamate, sucrose, or acetate in the hypotonic solution as well as by an administration of NPPB or tamoxifen, classical VACC inhibitors. Surprisingly, the VACC amplitude is greater in CF HBDC than in normal HBDC, suggesting that the channel density or open probability of VACC is increased, thus CFTR may have inhibitory effects on VACC. On the contrary, the amplitude of the volume-activated potassium current is lower in CF HBDC, suggesting the potassium channel density or open probability is decreased in CF cholangiocytes and/or CFTR may have regulatory effects on volume-activated potassium current. In conclusion, RVD is impaired in CF human cholangiocytes. The VACC of CF human cholangiocytes has similar electrophysiological characteristics as that of normal cholangiocytes but its activity is augmented in CF cholangiocytes, while volume-activated potassium current is decreased in CF human cholangiocytes, providing a fundamental underlying pathophysiologic mechanism for the impaired RVD in CF cholangiocytes.

Patients Receiving Prebiotics and Probiotics Before Liver Transplantation Develop Fewer Infections Than Controls: A Systematic Review and Meta-Analysis.

BACKGROUND & AIMS: Among patients who have received liver transplants, infections increase morbidity and mortality and prolong hospital stays. Administration of antibiotics and surgical trauma create intestinal barrier dysfunction and microbial imbalances that allow enteric bacteria to translocate to the blood. Probiotics are believed to prevent bacterial translocation by stabilizing the intestinal barrier and stimulating proliferation of the intestinal epithelium, mucus secretion, and motility. We performed a meta-analysis to determine the effects of probiotics on infections in patients receiving liver transplants. METHODS: We searched PubMed and EMBASE for controlled trials that evaluated the effects of prebiotics and probiotics on infections in patients who underwent liver transplantation. Heterogeneity was analyzed by the Cochran Q statistic. Pooled Mantel-Haenszel relative risks were calculated with a fixed-effects model. RESULTS: We identified 4 controlled studies, comprising 246 participants (123 received probiotics, 123 served as controls), for inclusion in the meta-analysis. In these studies, the intervention groups received enteric nutrition and fiber (prebiotics) with probiotics, and the control groups received only enteric nutrition and fiber without probiotics. The infection rate was 7% in groups that received probiotics vs 35% in control groups (relative risk [RR], 0.21; 95% confidence interval [CI], 0.11-0.41; P = .001). The number needed to treat to prevent 1 infection was 3.6. In subgroup analyses, only 2% of subjects in the probiotic groups developed urinary tract infections, compared with 16% of controls (RR, 0.14; 95% CI, 0.04-0.47; P < .001); only 2% of subjects in the probiotic groups developed intra-abdominal infections, compared with 11% of controls (RR, 0.27; 95% CI, 0.09-0.78; P = .02). Subjects receiving probiotics also had shorter stays in the hospital than controls (mean difference, 1.41 d; P < .001), as well as in the intensive care unit (mean difference, 1.41 d; P < .001), and duration of antibiotic use (mean difference, 3.89 d; P < .001). There was no difference in mortality between groups (RR, 0.97; 95% CI, 0.21-4.47). There was no significant heterogeneity among studies. CONCLUSIONS: Based on the meta-analysis, giving patients a combination of probiotics and prebiotics before, or on the day of, liver transplantation reduces the rate of infection after surgery. These agents also reduced the amount of time spent in the hospital or intensive care unit and the duration of antibiotic use.

Decreasing trend of upper gastrointestinal bleeding mortality risk over three decades.

BACKGROUND: Upper gastrointestinal bleeding (UGIB) causes over $1 billion in medical expenses annually. AIMS: The purpose of this study was to examine changes of UGIB mortality risks and trends over the last three decades. METHODS: We analyzed the National Hospital Discharge Sample from 1979 to 2009. Patients with primary ICD-9 code representing a diagnosis of UGIB were included. The UGIB mortality risks and trends in each decade by anatomical sites, bleeding causes, comorbidities, and other important variables were analyzed. RESULTS: UGIB mortality risk decreased by 35.4 % from 4.8 % in the first decade to 3.1 % in the third decade (P < 0.001). Age and number of hospitalization days were significant risk factors in all decades. Most significant decreases were observed in patients over 65 years and during the first day of admission. Gastric (P < 0.001) and esophageal (P = 0.018) bleedings showed significant decreasing mortality risk trends. Duodenal bleeding mortality risk was stable in three decades. Mortality risk declined significantly among patients with renal failure (from 50.0 to 4.0 %) and heart failure (from 17.9 to 5.2 %; both P < 0.001) while changes in cases with ischemic heart disease, cancer, and liver failure were less significant. CONCLUSION: UGIB morality risks, especially of the first hospital day and geriatric patients, significantly decreased over the last three decades, presumably from recent advances in emergency medical care. Mortality risk of gastric, but not duodenal, bleeding had the most significant reduction. Critical care improvements in patients with various comorbidities may explain significant UGIB mortality risk reductions. This study provides invaluable insight into the causes and trends of UGIB mortality risks for future studies.

Esophageal squamous papilloma: Literature review and case-control retrospective study with histopathological exam of human papillomavirus.

Background and Aim: Esophageal squamous papilloma (ESP) is a benign growth in the esophagus with unknown malignant potential. The mechanism underlying ESP formation is unknown, but human papillomavirus (HPV) infection has been proposed as a potential etiology. We sought to investigate the clinical characteristic of ESP in our population, review the current literature, and highlight the role of HPV. Methods: This is a retrospective case-control study conducted at two referral centers. We selected the ESP population by free-text search in the pathology department database and selected controls randomly from the general endoscopy population. Immunostains were used to evaluate ESP tissue for HPV. Results: Between January 2016 and December 2021, we identified 66 patients with ESP, with a prevalence of 0.72%. ESP patients were younger, with a median age of 52 years (P = 0.021), and more likely African American (34.4 vs 7.5%, P < 0.001) compared to controls. On endoscopy images, the growth was predominantly solitary (92.5%) in the middle of the esophagus (39.4%), with sizes ranging from 0.2 to 2.3 cm. A total of 62 patients had available tissue for HPV immune staining, and none tested positive for HPV. Eighteen patients had a follow-up endoscopy with an average of 504.5 days follow-up period. One patient developed esophageal squamous cell carcinoma during follow-up. Conclusions: We observed a higher prevalence of ESP compared to previous studies. The formation of ESP is multifactorial and partially explained by HPV infection in selected populations. The malignant potential of ESP is low but not negligible.

Predictors of colorectal carcinoma and inflammatory bowel disease in patients with colonic wall thickening.

Background and Aim: Colonic wall thickening (CWT) is commonly associated with clinically significant pathologies, but predictive factors of such pathologies are not well known. This study aims to identify the predictors of clinically significant pathologies, such as colorectal carcinoma (CRC) and inflammatory bowel disease (IBD), in patients with CWT. Methods: Subjects with an abnormal abdominal computed tomography (CT) and a follow-up colonoscopy between 2010 and 2020 were retrospectively reviewed. Patients with CWT in the CT were included and examined in this study. A multivariable logistic regression analysis was performed to assess for factors independently associated with CRC or IBD in these subjects. Receiver operating characteristic (ROC) curve analysis was used to further examine significant parameters in multivariable logistic regression analysis. Results: Among 403 patients with CWT on CT scans who underwent a colonoscopy, 269 subjects who met the inclusion criteria were identified and studied. On multivariable logistic regression models, elevated platelet count, low hematocrit, and localized CWT were found to be independently associated with CRC, while elevated platelet count and younger age were independently associated with IBD. On ROC curve analysis for CRC, area under the curve (AUC) for hematocrit, platelets, and localized CWT was 0.76, 0.75, and 0.61, respectively. On ROC curve analysis for IBD, AUC for age and platelets was 0.90 and 0.69, respectively. Conclusion: Elevated platelet count, low hematocrit, and localized CWT can be potentially used as predictors of CRC in patients with CWT. Elevated platelet count and young age can be used to predict IBD in these patients.

Ethanol metabolism by HeLa cells transduced with human alcohol dehydrogenase isoenzymes: control of the pathway by acetaldehyde concentration.

BACKGROUND: Human class I alcohol dehydrogenase 2 isoenzymes (encoded by the ADH1B locus) have large differences in kinetic properties; however, individuals inheriting the alleles for the different isoenzymes exhibit only small differences in alcohol elimination rates. This suggests that other cellular factors must regulate the activity of the isoenzymes. METHODS: The activity of the isoenzymes expressed from ADH1B*1, ADH1B*2, and ADH1B*3 cDNAs was examined in stably transduced HeLa cell lines, including lines which expressed human low K(m) aldehyde dehydrogenase (ALDH2). The ability of the cells to metabolize ethanol was compared with that of HeLa cells expressing rat class I alcohol dehydrogenase (ADH) (HeLa-rat ADH cells), rat hepatoma (H4IIEC3) cells, and rat hepatocytes. RESULTS: The isoenzymes had similar protein half-lives in the HeLa cells. Rat hepatocytes, H4IIEC3 cells, and HeLa-rat ADH cells oxidized ethanol much faster than the cells expressing the ADH1B isoenzymes. This was not explained by high cellular NADH levels or endogenous inhibitors; but rather because the activity of the ß1 and ß2 ADHs was constrained by the accumulation of acetaldehyde, as shown by the increased rate of ethanol oxidation by cell lines expressing ß2 ADH plus ALDH2. CONCLUSION: The activity of the human ß2 ADH isoenzyme is sensitive to inhibition by acetaldehyde, which likely limits its activity in vivo. This study emphasizes the importance of maintaining a low steady-state acetaldehyde concentration in hepatocytes during ethanol metabolism.

Characterization of volume-activated chloride currents in regulatory volume decrease of human cholangiocyte.

Volume-activated chloride channel (VACC) plays vital roles in many physiological functions. In bile duct epithelium, VACC actively participates in biliary secretion and cell volume regulation, and it mediates regulatory volume decrease (RVD). Recently, we have shown that mouse cholangiocytes have an intact RVD via VACC and K(+) conductance. However, such cell volume regulation was not studied in the normal human cholangiocyte. Volume measurement by Coulter counter and whole-cell patch clamp technique were used to characterize the RVD and VACC in human cholangiocyte cell line (HBDC). When exposed to hypotonic solution, HBDC exhibited an intact RVD, which was inhibited by 1,2-Bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetrakis(acetoxymethyl ester) (BAPTA-AM), NPPB (5-nitro-2'- (3-phenylpropylamino)-benzoate), DIDS (4,4'-diisothiocyanatostilbene-2-disulfonic acid), and tamoxifen, but was not affected by the removal of extracellular calcium. During RVD, HBDC exhibited large, outwardly rectifying currents and time-dependent inactivation at positive potential. The amplitude of the outward current was approximately 3 times of that of the inward current, and this volume-activated current returned to the baseline when switched to isotonic solution. The amplitude and reversal potential of the volume-activated current was dependent on Cl(-) concentration, and the VACC was significantly inhibited by replacing chloride with gluconate, glutamate, sucrose, and acetate in the hypotonic solution. In addition, classical VACC inhibitors, such as NPPB or tamoxifen, inhibited the VACC. These inhibitory effects were reversible with washing out the inhibitors from the bath solution. The present study is the first to characterize and show that HBDC has an intact RVD, mediated by VACC, which has similar electrophysiological characteristics as that in mouse cholangiocytes.

Role of calcium in volume-activated chloride currents in a mouse cholangiocyte cell line.

Volume-activated Cl(-) channels (VACCs) play vital roles in many cells including cholangiocytes. Previously, we characterized the VACCs in mouse cholangiocytes. Since calcium plays an important role in VACC regulation in many cells, we have studied the effect of calcium modulation on the regulatory volume decrease (RVD) and VACC currents in mouse bile duct cells (MBDCs). Cell volume measurements were assessed by a Coulter counter with cell sizer, and conventional whole-cell patch-clamp techniques were used to study the role of calcium on RVD and VACC currents. Cell volume study indicated that MBDCs exhibited RVD, which was inhibited by 5-nitro-2'-(3-phenylpropylamino)-benzoate (NPPB), 4,4'-diisothiocyanostilbene-2,2'-disulfonate (DIDS) and 1,2-bis(o-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid tetra-acetoxymethyl ester (BAPTA-AM) but not by removal of extracellular calcium. During hypotonic challenge, MBDCs exhibited an outwardly rectified current, which was significantly inhibited by administration of classical chloride channel inhibitors such as NPPB and tamoxifen. Chelation of the intracellular calcium with BAPTA-AM or removal of extracellular calcium and calcium channel blocker had no significant effect on VACC currents during hypotonic challenge. In addition to VACC, MBDC had a calcium-activated chloride channel, which was inhibited by NPPB. The present study is the first to systemically study the role of calcium on the VACC and RVD in mouse cholangiocytes and demonstrates that a certain level of intracellular calcium is necessary for RVD but the activation of VACC during RVD does not require calcium. These findings suggest that calcium does not have a direct regulatory role on VACC but has a permissive role on RVD in cholangiocytes.

Permissive role of calcium on regulatory volume decrease in freshly isolated mouse cholangiocytes.

Calcium (Ca2+) pathways are important in cell volume regulation in many cells, but its role in volume regulatory processes in cholangiocytes is unclear. Thus, we have investigated the role of Ca2+ in regulatory volume decrease (RVD) in cholangiocytes using freshly isolated bile duct cell clusters (BDCCs) from normal mouse. No significant increase in [Ca2+]i was observed during RVD, while ionomycin and ATP showed significant increases. Confocal imaging also showed no significant changes in the levels or distributions of intracellular Ca2+ during RVD. Cell volume study by quantitative videomicroscopy indicated that removal and chelation of extracellular Ca2+ by ethylene glycol-bis (beta-aminoethyl ether)-N,N,N-tetraacetic acid (EGTA) or administration of nifedipine did not affect RVD but verapamil significantly inhibited the RVD. Moreover, Ca2+ agonists or inhibitors of Ca2+ release from intracellular stores had no significant effect on RVD. However, 1,2-bis (2-aminophenoxy) ethane-N,N,N'N'-tetraacetic acid-AM (BAPTA-AM) showed significant decreases in [Ca2+]i and significantly inhibited RVD, which was reversed with coadministration of valinomycin, suggesting that BAPTA-AM-induced inhibition is due to potassium conductance or other cellular processes requiring permissive [Ca2+](i. These findings indicate that an increase in [Ca2+]i or extracellular Ca2+ is not required for RVD but Ca2+ has a permissive role in RVD of mouse cholangiocytes.

Glycosylphosphatidylinositol-specific phospholipase D influences triglyceride-rich lipoprotein metabolism.

Glycosylphosphatidylinositol-specific phospholipase D (GPI-PLD) is a minor HDL-associated protein. Because many minor HDL-associated proteins exchange between different lipoprotein classes during the postprandial state and are also involved in triglyceride (TG) metabolism, we hypothesized that GPI-PLD may play a role in the metabolism of TG-rich lipoproteins. To test this hypothesis, we examined the distribution of GPI-PLD among lipoprotein classes during a fat tolerance test in C57BL/6 and LDL receptor-deficient (LDLR(-/-)) mice fed either a chow or high-fructose diet. In the fasting state in wild-type mice fed a chow diet, GPI-PLD was only present in HDL, whereas in LDLR(-/-) mice GPI-PLD was present in HDL and intermediate-density lipoproteins (IDL)/LDL. During the fat tolerance test, there was no change in total serum GPI-PLD levels in either model; however, a significant amount of GPI-PLD appeared in both VLDL (0.5-1% of total GPI-PLD) and IDL/LDL (5-10% of total GPI-PLD) in both models. The high-fructose diet increased both fasting and postprandial TG and serum GPI-PLD levels in both strains as well as the amount of GPI-PLD in VLDL. To determine whether GPI-PLD plays a direct role in TG metabolism, we increased liver GPI-PLD expression in C57BL/6 mice by adenovirus-mediated gene transfer, which resulted in a sevenfold increase in serum GPI-PLD levels. This change was associated with an increase in fasting (30%) and postprandial TG (50%) and a twofold reduction in TG-rich lipoprotein catabolism compared with saline or control adenovirus-treated mice. These studies demonstrate that GPI-PLD affects serum TG levels by altering catabolism of TG-rich lipoproteins.

Hepatocellular carcinoma occurring in a patient with Crohn's disease treated with both azathioprine and infliximab.

Hepatocellular carcinoma is known to be associated with underlying liver diseases, such as cirrhosis, hemochromatosis, and chronic viral hepatitis. All reported cases of hepatocellular carcinoma in association with Crohn's disease involve patients treated previously with azathioprine or both azathioprine and steroids. However, hepatocellular carcinoma associated with the use of azathioprine and infliximab has not been reported. In this report, we describe an unusual case of hepatocellular carcinoma and focal hepatic glycogenosis (FHG) occurring in a non-cirrhotic Crohn's disease patient who has been treated with both azathioprine and infliximab.

Characterization of regulatory volume decrease in freshly isolated mouse cholangiocytes.

Cell volume regulation plays a vital role in many cell functions. Recent study indicates that both K(+) and Cl(-) channels are important for the regulatory volume decrease (RVD) of cholangiocarcinoma cells, but its physiological significance is unclear due to the tumorous nature of the cells used. This present study reports the RVD of normal mouse cholangiocytes by using freshly isolated bile duct cell clusters (BDCC). A relatively simple and practical method of measuring the cross-sectional area of BDCCs by quantitative videomicroscopy was used to indirectly measure their volumes. Mouse cholangiocytes exhibited RVD, which was inhibited by 5-nitro-2'-(3-phenylpropylamino)-benzoate, DIDS, and glibenclamide, suggesting its dependence on certain chloride channels, such as volume-activated chloride channels. It is also inhibited by barium chloride but not by tetraethylammonium chloride, indicating its dependence on certain potassium channels. However, cAMP agonists had no significant effect on the RVD of BDCCs. This indirect method described can be used to study the RVD of cholangiocytes from normal as well as genetically altered mouse livers.

Electrophysiological characterization of volume-activated chloride currents in mouse cholangiocyte cell line.

Recent electrophysiological and radioisotope efflux studies have demonstrated various Cl(-) channels in cholangiocytes including volume-activated Cl(-) channels (VACC). Because VACCs play prominent roles in many vital cellular functions and physiology in cholangiocytes, we have examined their electrophysiological characteristics in mouse cholangiocytes to provide an important framework for studying in the future. The present study is to characterize VACCs expressed in the mouse bile duct cell (MBDC) line, conditionally immortalized by SV40 virus. Conventional whole cell patch-clamp techniques were used to study the electrophysiological characteristics of VACC in MBDC. When the MBDCs were exposed to hypotonic solution, they exhibited an outwardly rectified current, which was significantly inhibited by replacing chloride in the bath solution with gluconate or glutamate and by administration of classic chloride channel inhibitors 5-nitro-2-(3-phenylpropylamino)-benzoate, glybenclamide, DIDS, and tamoxifen. These inhibitory effects were reversible with washing them out from the bath solution. Moreover, the ion selectivity of the volume-activated channel to different anions indicates that it is more permeable to SCN(-) > I(-) >/= Cl(-) > F(-) >/= acetate >/= glutamate >/= gluconate. These electrophysiological characteristics demonstrate that the volume-activated current observed is a VACC. In addition, the VACC in MBDC has electrophysiological characteristics similar to those of the VACC in human cholangiocarcinoma cell line. The present study is the first to characterize the VACC in mouse cholangiocyte and will provide an important framework for further studies to examine and understand the role of the VACC in biliary secretion and ion-transport physiology.

Impaired regulatory volume decrease in freshly isolated cholangiocytes from cystic fibrosis mice: implications for cystic fibrosis transmembrane conductance regulator effect on potassium conductance.

Various K(+) and Cl(-) channels are important in cell volume regulation and biliary secretion, but the specific role of cystic fibrosis transmembrane conductance regulator in cholangiocyte cell volume regulation is not known. The goal of this research was to study regulatory volume decrease (RVD) in bile duct cell clusters (BDCCs) from normal and cystic fibrosis (CF) mouse livers. Mouse BDCCs without an enclosed lumen were prepared as described (Cho, W. K. (2002) Am. J. Physiol. 283, G1320-G1327). The isotonic solution consisted of HEPES buffer with 40% of the NaCl replaced with isomolar amounts of sucrose, whereas hypotonic solution was the same as isotonic solution without sucrose. The cell volume changes were indirectly assessed by measuring cross-sectional area (CSA) changes of the BDCCs using quantitative videomicroscopy. Exposure to hypotonic solutions increased relative CSAs of normal BDCCs to 1.20 +/- 0.01 (mean +/- S.E., n = 50) in 10 min, followed by RVD to 1.07 +/- 0.01 by 40 min. Hypotonic challenge in CF mouse BDCCs also increased relative CSA to 1.20 +/- 0.01 (n = 53) in 10 min but without significant recovery. Coadministration of the K(+)-selective ionophore valinomycin restored RVD in CF mouse BDCCs, suggesting that the impaired RVD was likely from a defect in K(+) conductance. Moreover, this valinomycin-induced RVD in CF mice was inhibited by 5-nitro-2'-(3-phenylpropylamino)-benzoate, indicating that it is not from nonspecific effects. Neither cAMP nor calcium agonists could reverse the impaired RVD seen in CF cholangiocytes. Our conclusion is that CF mouse cholangiocytes have defective RVD from an impaired K(+) efflux pathway, which could not be reversed by cAMP nor calcium agonists.

Transcriptional control of the human aldehyde dehydrogenase 2 promoter by hepatocyte nuclear factor 4: inhibition by cyclic AMP and COUP transcription factors.

An important regulatory element (designated FP330-3') of the ALDH2 promoter mediates activation by hepatocyte nuclear factor 4 (HNF4). This activation of promoter constructs containing this element by HNF4 was reduced by nearly half by 8-Br-cAMP in H4IIEC3 cells, an effect that was blocked by inhibitors of protein kinase A (PKA). Cotransfection assays showed that COUP-TF I, ARP-1, or PPARdelta suppressed the ability of HNF4 to activate the reporter. The repression was potentiated by 8-Br-cAMP. Electrophoretic mobility shift assays revealed that treatment of hepatoma cells or cultured rat hepatocytes with 1 mM 8-Br-cAMP or glucagon reduced binding of FP330-3' by HNF4 by half. In vitro phosphorylation of HNF4 by PKA decreased binding to FP330-3'. Fasting reduced the ALDH2 protein level in liver and kidney, two tissues expressing HNF4, but not heart. These data suggest that ALDH2 expression can be suppressed by cAMP, most likely through phosphorylation of HNF4 by PKA, and this may account for the reduction in enzyme protein during fasting.

The role of AMP-activated protein kinase in the action of ethanol in the liver.

BACKGROUND & AIMS: Our previous work has shown that ethanol induces the fatty acid synthesis pathway by activation of sterol regulatory element-binding protein 1 (SREBP-1). In the present study, we studied the mechanisms of this activation by identifying a new target of ethanol, AMP-activated protein kinase (AMPK). METHODS: The effects of ethanol on AMPK, acetyl-CoA carboxylase (ACC), and SREBP-1 were assessed in rat hepatic cells and in the livers of ethanol-fed mice. RESULTS: In rat hepatoma H4IIEC3 or McA-RH 7777 cell lines, ethanol-induced transcription of an SREBP-regulated promoter was suppressed by the presence of 5-aminoimidazole-4-carboxamide ribonucleoside (AICAR) or metformin, 2 known AMPK activators. Consistent with this, over expression of a constitutively active form of AMPK blocked the effect of ethanol, whereas coexpression of a dominant-negative form of AMPK augmented the effect. Moreover, activation of AMPK by metformin or AICAR largely blocked the ability of ethanol to increase levels of mature SREBP-1 protein. These findings suggest that the effect of ethanol on SREBP-regulated promoter activation was partially mediated through AMPK inhibition. We further demonstrated that AMPK was inhibited by ethanol in hepatic cells. In parallel, ethanol increased the activity of ACC and suppressed the rate of palmitic acid oxidation. Finally, feeding mice a low-fat diet with ethanol resulted in significantly reduced hepatic AMPK activity, increased ACC activity, and enhanced malonyl CoA content. CONCLUSIONS: Taken together, our findings suggest that AMPK may play a key role in regulating the effects of ethanol on SREBP-1 activation, fatty acid metabolism, and development of alcoholic fatty liver.

Identification of a retinoid receptor response element in the human aldehyde dehydrogenase-2 promoter.

BACKGROUND: The human aldehyde dehydrogenase-2 promoter contains sites that bind members of the nuclear receptor family, and one (designated FP330-3') is predicted to bind retinoic acid receptors. METHODS: Binding of retinoid receptors to the FP330-3' oligonucleotide duplex and point mutations thereof was assayed using electrophoretic mobility shift assays. The function of the promoter element was determined in transfection assays. RESULTS: Heterodimers of retinoic acid receptor (RAR)alpha, beta, and gamma with retinoid X receptor (RXR)alpha bound the FP330-3' site. Mutagenesis of the FP330-3' site suggested that either the upstream DR-5 or downstream DR-1 could mediate binding of RAR/RXR. FP330-3' oligonucleotide duplexes were not bound by in vitro translated RXR homodimers but weakly competed with a synthetic DR-1 oligonucleotide duplex for binding by RXR. A reporter construct carrying four copies of the FP330-3' element was induced by cotransfection of rat hepatoma cells with a construct encoding RARalpha, when the RAR-specific ligand AM580 was present. Each of the three RXR isoforms alpha, beta, and gamma stimulated the expression of reporter constructs containing the FP330-3' sites in a 9-cis retinoic acid-dependent fashion in cells in culture. This was confirmed in the case of RXRalpha using the RXR-specific ligand methoprene. CONCLUSION: The human aldehyde dehydrogenase-2 promoter contains a retinoid response element, which may contribute to regulation of the gene.