New aspects of a small GTPase RAB35 in brain development and function

In eukaryotic cells,organelles in the secretory,lysosomal,and endocytic pathways actively exchange biological materials with each other through intracellular membrane trafficking,which is the process of transporting the cargo of proteins,lipids,and other molecules to appropriate compartments via transport vesicles or intermediates.These processes are strictly regulated by various small GTPases such as the RAS-like in rat brain(RAB)protein family,which is the largest subfamily of the RAS superfamily.Dysfunction of membrane trafficking affects tissue homeostasis and leads to a wide range of diseases,including neurological disorders and neurodegenerative diseases.Therefore,it is important to understand the physiological and pathological roles of RAB proteins in brain function.RAB35,a member of the RAB family,is an evolutionarily conserved protein in metazoans.A wide range of studies using cultured mammalian cells and model organisms have revealed that RAB35 mediates various processes such as cytokinesis,endocytic recycling,actin bundling,and cell migration.RAB35 is also involved in neurite outgrowth and turnover of synaptic vesicles.We generated brain-specific Rab35 knockout mice to study the physiological roles of RAB35 in brain development and function.These mice exhibited defects in anxiety-related behaviors and spatial memory.Strikingly,RAB35 is required for the precise positioning of pyramidal neurons during hippocampal development,and thereby for normal hippocampal lamination.In contrast,layer formation in the cerebral cortex occurred superficially,even in the absence of RAB35,suggesting a predominant role for RAB35 in hippocampal development rather than in cerebral cortex development.Recent studies have suggested an association between RAB35 and neurodegenerative diseases,including Parkinson's disease and Alzheimer's disease.In this review,we provide an overview of the current understanding of subcellular functions of RAB35.We also provide insights into the physiological role of RAB35 in mammalian brain development and function,and discuss the involvement of RAB35 dysfunction in neurodegenerative diseases.

Glucagon-like peptide-1 analogue prevents nonalcoholic steatohepatitis in non-obese mice

AIM: To investigate whether a glucagon-like peptide-1(GLP-1) analogue inhibits nonalcoholic steatohepatitis(NASH), which is being increasingly recognized in Asia, in non-obese mice. METHODS: A methionine-choline-deficient diet(MCD) along with exendin-4(20 μg/kg per day, ip), a GLP-1 analogue, or saline was administered to male db/db mice(non-obese NASH model). Four or eight weeks after commencement of the diet, the mice were sacrificed and their livers were excised. The excised livers were examined by histochemistry for evidence of hepatic steatosis and inflammation. Hepatic triglyceride(TG) and free fatty acid(FFA) content was measured, and the expression of hepatic fat metabolism- and inflammation-related genes was evaluated. Oxidative stress-related parameters and macrophage recruitment were also examined using immunohistochemistry.RESULTS: Four weeks of MCD feeding induced hepatic steatosis and inflammation and increased the hepatic TG and FFA content. The expression of fattyacid transport protein 4(FATP4), a hepatic FFA influxrelated gene; macrophage recruitment; and the level of malondialdehyde(MDA), an oxidative stress marker, were significantly augmented by a 4-wk MCD. The levels of hepatic sterol regulatory element-binding protein-1c(SREBP-1c) m RNA(lipogenesis-related gene) and acyl-coenzyme A oxidase 1(ACOX1) m RNA(β-oxidation-related gene) had decreased at 4 wk and further decreased at 8 wk. However, the level of microsomal triglyceride transfer protein m RNA(a lipid excretion-related gene) remained unchanged. The administration of exendin-4 significantly attenuated the MCD-induced increase in hepatic steatosis, hepatic TG and FFA content, and FATP4 expression as well as the MCD-induced augmentation of hepatic inflammation, macrophage recruitment, and MDA levels. Additionally, it further decreased the hepatic SREBP-1c level and alleviated the MCD-mediated inhibition of the ACOX1 m RNA level. CONCLUSION: These results suggest that GLP-1 inhibits hepatic steatosis and inflammation through the inhibition of hepatic FFA influx and oxidative stress in a non-obese NASH model.

Spontaneous regression of hepatic inflammatory pseudotumor with primary biliary cirrhosis:Case report and literature review

Hepatic inflammatory pseudotumor （IPT） is a rare benign non-neoplastic lesion characterized by proliferating fibrous tissue infiltrated by inflammatory cells. The exact etiology of IPT remains unclear. Although the association of IPT with systemic inflammatory disorders has been well established, a specific relationship with cholangitis is distinctly rare. We report a case of spontaneous regression of hepatic IPT with primary biliary cirrhosis （PBC）. To date, only two cases of IPT with PBC have been reported. In our case, however, IPT developed during the course of improvement of cholangitis of PBC induced by effective treatment, differing from two previously reported cases. Our case indicates that the development of IPT does not also relate to the activity of cholangitis and/or hyper gamma-globulinemia, since our case was confirmed radiologically to be free of IPT when biliary enzymes and immunoglobulins were much higher than the corresponding values on admission. Comparison of our case with the two previously reported cases suggests that IPT occurring with PBC does not represent the same disease entity or be a bystander for PBC.

Hepatocellular carcinoma in extremely elderly patients:An analysis of clinical characteristics,prognosis and patient survival

AIM： To identify the clinical and prognostic features of patients with hepatocellular carcinoma （HCC） aged 80 years or more. METHODS： A total of 1310 patients with HCC were included in this study. Ninety-one patients aged 80 years or more at the time of diagnosis of HCC were defined as the extremely elderly group. Two hundred and thirty-four patients aged 〉/ 50 years but less than 60 years were regarded as the non-elderly group. RESULTS： The sex ratio （male to female） was significantly lower in the extremely elderly group （0.90：1） than in the non-elderly group （3.9：1, P〈 0.001）. The positive rate for HBsAg was significantly lower in the extremely elderly group and the proportion of patients negative for HBsAg and HCVAb obviously increased in the extremely elderly group （P〈 0.001）. There were no significant differences in the following parameters： diameter and number of tumors, Child-Pugh grading, tumor staging, presence of portal thrombosis or ascites, and positive rate for HCVAb. Extremely elderly patients did not often receive surgical treatment （P 〈 0.001） and they were more likely to receive conservative treatment （P〈 0.01）. There were no significant differences in survival curves based on the Kaplan-Meier methods in comparison with the overall patients between the two groups. However, the survival curves were significantly worse in the extremely elderly patients with stage Ⅰ/Ⅱ, stage Ⅰ/Ⅱ and Child-Pugh grade A cirrhosis in comparison with the non-elderly group. The causes of death did not differ among the patients, and most cases died of liverrelated diseases even in the extremely elderly patients. CONCLUSION： In the patients with good liver functions and good performance status, aggressive treatment for HCC might improve the survival rate, even in extremely elderly patients.

Advanced hepatocellular carcinoma responds to MK615, a compound extract from the Japanese apricot “Prunus mume ”

MK615, a compound extracted from the Japanese apricot "Prunus mume " has been reported to have in vitro anti-tumor activities against several cancer cell lines,including hepatocellular carcinoma(HCC). However, the clinical effects and feasibility of administering MK615for patients with HCC were unknown. We experienced a case with advanced HCC for which MK615 was effective against both lymph node and pulmonary metastases. A 60-year-old female underwent surgical resection of a 9 cm HCC in the right lobe. The pathological diagnosis was moderately differentiated HCC with vascular invasion. The HCC recurred in the liver 8 mo after the surgery. Radiofrequency ablation and transarterial infusion chemotherapy were performed, but the recurrence was not controlled. One year after the intrahepatic recurrence, pulmonary and lymph metastasis appeared.Sorafenib was administered, but was not effective.Then, MK615 was administered as a final alternative therapy after informed consent was obtained from the patient. Three months later, her alpha-fetoprotein level decrease and both the lymph node and pulmonary metastases decreased in size. The patient has survived for more than 17 mo after the MK615 administration, and was in good condition. Although further investigations are necessary to clarify its safety and efficacy in humans, MK615 may be useful for the treatment of HCC,without serious adverse effects.

MK-0626,a selective DPP-4 inhibitor,attenuates hepatic steatosis in ob/ob mice

AIM: To investigate the mechanism and in vivo effects of MK-0626, a dipeptidyl peptidase-4 inhibitor, on hepatic steatosis using ob/ob mice.

Challenge to overcome: Nonstructural protein 5A-P32 deletion in direct-acting antiviral-based therapy for hepatitis C virus

Interferon(IFN)-based therapy for hepatitis C virus(HCV) infection has recently been replaced by IFNfree direct-acting antiviral(DAA)-based therapy, which has been established as a 1^(st) line therapy with high efficacy and tolerability due to its reasonable safety profile. Resistance-associated substitutions(RASs) have been a weakness of DAA-based therapy. For example, combination therapy with daclatasvir and asunaprevir(DCV/ASV) is less effective for HCV genotype 1-infected patients with Y93H as a nonstructural protein 5A RAS. However, the problem regarding RASs has been gradually overcome with the advent of recently developed DAAs, such as sofosbuvir-based regimens or combination therapy with glecaprevir and pibrentasvir. Despite the high efficiency of DAA-based therapy, some cases fail to achieve viral eradication. P32 deletion, an NS5A RAS, has been gradually noticed in patients with DCV/ASV failure. P32 deletion has been sporadically reported and the prevalence of this RAS has been considered to be low in patients with DCV/ASV failure. Thus, the picture of P32 deletion has not been fully evaluated. Importantly, currently-commercialized DAA-based combination therapy was not likely to be effective for patients with P32 deletion. Exploring and overcoming this RAS is essential for antiviral therapy for chronic hepatitis C.

Overexpression of NK2 inhibits liver regeneration after partial hepatectomy in mice

AIM： To investigate the in vivo effects of NK2 on liver regeneration after partial hepatectomy （PH）. METHODS： Survival after PH was observed with 21 NK2 transgenic mice and 23 wild-type （WT） mice over 10 d. Liver regeneration was analyzed using histology and immunohistochemistry. Expressions of genes were analyzed using Northern blot analysis, immunoprecipitation and immunoblotting, and reverse transcriptase polymerase chain reaction assay. Kaplan Meier method and the log-rank test were used for analyzing the survival after PH. Differences in the results of immunohistochemistry and percentage of liver regeneration was determined by the Student＇s t-test. RESULTS： More than half of NK2 transgenic mice died within 48 h after PH. After PH, increased deposition of small lipid droplets in hepatocytes was evident and hepatic proliferation was inhibited in NK2 transgenic mice. The hepatic expression and kinase activity of HGF receptor, c-Met, were unchanged among WT mice and NK2 transgenic mice after PH. The expression of tumor necrosis factor-co （TNF-c0 and interleukin-6 （IL-6） in liver tissues were prolonged in NK2 transgenic mice that died after PH. CONCLUSION： Our findings indicate that overexpression of NK2 inhibits liver regeneration after PH.

Clinical characteristics of null responders to Peg-IFNα2b/ ribavirin therapy for chronic hepatitis C

AIM: To predict which chronic hepatitis C patients are likely to be late-responders, we herein investigated the clinical characteristics of null-responders at 36 wk with hepatitis C virus (HCV) genotype Ib and a high viral load during the course of pegylated interferon (Peg-IFN)/ ribavirin ther apy. METHODS: One hundred forty-two patients with genotype Ib HCV and a high viral load were included in this study. Peg-IFNα2b (1.5 μg/kg once a week) and ribavirin (600-1000 mg per day according to body weight) were administered for 48 wk. We def ined nullresponders as the cases that never cleared serum HCV RNA as determined using RT-PCR until 36 wk. Other patients were def ined as responders. We compared the clinical characteristics (age, gender, body mass index, previous treatment) and HCV RNA titer during the therapy between null-responders and responders.RESULTS: The HCV RNA clearance rate was 17.9% (24/134), 46.3% (62/134), 60.6% (86/142), 86.6% (123/142), and 88.0% (125/142) at 4, 8, 12, 24, and 36 wk, respectively. There were 17 patients (12.0%) who were still null-responders at 36 wk. There were no differences in the clinical characteristics between the responders and null-responders except for the titer and decline rates of HCV RNA at 1 wk and 4 wk. The HCV RNA titers at 1 wk and after 4 wk of treatment were significantly higher in the null-responders in comp arison to the responders (P <0.01). The serum HCV RNA titers of the responders decreased by 1.3 log after 1 wk of treatment, and 1.6 log after 4 wk of treatm ent, respectively. On the other hand, the titers of the null responders decreased by only 0.5 log after 1 wk, and 0.7 log after 4 wk of treatment, respectively. The decrease rates of HCV RNA after 1 and 4 wk of treatm ent were signif icantly worse for null responders than for the responders (P <0.01). CONCLUSION: The HCV RNA titer at 1 wk and 4 wk after initiating treatment may be useful for predicting null-responders to Peg-IFNα2b/ribavirin therapy. However, further investigation is needed to determine the optimal time at which the decision to discontinue the Peg-IFNα2b/ribavirin therapy for null-responders can be made.

Non-invasive prediction of non-alcoholic steatohepatitis in Japanese patients with morbid obesity by artificial intelligence using rule extraction technology

AIM To construct a non-invasive prediction algorithm for predicting non-alcoholic steatohepatitis(NASH), we investigated Japanese morbidly obese patients using artificial intelligence with rule extraction technology.METHODS Consecutive patients who required bariatric surgery underwent a liver biopsy during the operation. Standard clinical, anthropometric, biochemical measurements were used as parameters to predict NASH and were analyzed using rule extraction technology. One hundred and two patients, including 79 NASH and 23 non-NASH patients were analyzed in order to create the predictionmodel, another cohort with 77 patients including 65 NASH and 12 non-NASH patients were analyzed to validate the algorithm.RESULTS Alanine aminotransferase, C-reactive protein, homeostasis model assessment insulin resistance, albumin were extracted as predictors of NASH using a recursive-rule extraction algorithm. When we adopted the extracted rules for the validation cohort using a highly accurate rule extraction algorithm, the predictive accuracy was 79.2%. The positive predictive value, negative predictive value,sensitivity and specificity were 88.9%, 35.7%, 86.2% and 41.7%, respectively.CONCLUSION We successfully generated a useful model for predicting NASH in Japanese morbidly obese patients based on their biochemical profile using a rule extraction algorithm.

Combination therapy with daclatasvir and asunaprevir for dialysis patients infected with hepatitis C virus

The standard antiviral therapy for dialysis patients infected with hepatitis C virus(HCV) is(pegylated) interferon monotherapy, but its efficacy is insufficient. Oral direct-acting antiviral agents(DAAs) have recently been developed for chronic hepatitis C patients. However, some DAAs have contraindications for chronic renal failure(CRF). Daclatasvir and asunaprevir are metabolized largely in the liver and are not contraindicated in CRF. Combination therapy with daclatasvir and asunaprevir was used for 4 dialysis patients infected with genotype 1b HCV. One patient had viral breakthrough, and the 3 others had sustained virological response 12. One patient was admitted for heart failure and percutaneous coronary intervention due to concomitant ischemic disease. Heart failure was unlikely to be caused by the combination therapy, as it was probably due to water overload. The patient continued to receive the combination therapy after the remission of the heart failure. The combination therapy was well tolerated in the other patients.

Strategy for the control of drug-induced liver injury due to investigational treatments/drugs for COVID-19

Investigational treatments/drugs for coronavirus disease 2019(COVID-19)have been applied,with repurposed or newly developed drugs,and their effectiveness has been evaluated.Some of these drugs may be hepatotoxic,and each monotherapy or combination therapy may increase the risk of drug-induced liver injury(DILI).We should aim to control dysregulation of liver function,as well as the progression of COVID-19,as much as possible.We discussed the potential risks of investigational treatments/drugs and promising drugs for both COVID-19 and DILI due to investigational treatments/drugs.

Probable case of drug reaction with eosinophilia and systemic symptom syndrome due to combination therapy with daclatasvir and asunaprevir

A 66-year-old, interferon-ineligible, treatment-naive man who was diagnosed with chronic hepatitis C due to hepatitis C virus genotype 1b began combination therapy with daclatasvir and asunaprevir. On day 14 of treatment, hepatic reserve and renal function deterioration was observed, while his transaminase levels were normal. Both daclatasvir and asunaprevir were discontinued on day 18 of treatment, because the patient complained of dark urine and a rash on his trunk and four limbs. After discontinuing antiviral therapy, the abnormal laboratory finding and clinical manifestations gradually improved, without recurrence. Our case fulfilled the diagnostic criteria of probable drug reaction with eosinophilia and systemic symptom(DRESS) syndrome. Despite the 18-d treatment, sustained virological response 12 was achieved. Based on the clinical course, we concluded that there was a clear cause-and-effect relationship between the treatment and adverse events. To our knowledge, this patient represents the first case of probable DRESS syndrome that includes concomitant deterioration of hepatic reserve and renal function due to combination therapy with daclatasvir and asunaprevir, regardless of normalization of transaminase levels. Our case suggests that we should pay attention not only to the transaminase levels but also to allergic symptoms associated with organ involvement during combination therapy with daclatasvir and asunaprevir.

Acute esophageal necrosis caused by alcohol abuse

Acute esophageal necrosis (AEN) is extremely rare and the pathogenesis of this is still unknown. We report a case of AEN caused by alcohol abuse. In our case, the main pathogenesis could be accounted for low systemic perfusion caused by severe alcoholic lactic acidosis. After the healing of AEN, balloon dilatation was effective to manage the stricture.

Living-related liver transplantation for multiple liver metastases from rectal carcinoid tumor: A case report

A 42-year-old woman was admitted to our hospital because of multiple liver tumors detected by ultrasonography. Colonoscopy revealed submucosal tumor in the rectum, which was considered the primary lesion. Endoscopic mucosal resection followed by histopathological examination revealed that the tumor was carcinoid. The resected margin of the tumor was positive for malignant cells. Two courses to transcatheter arterial chemotherapy for liver metastasis were ineffective. Accordingly, the rectal tumor and metastatic lymph nodes were surgically resected. One month after the operation, she received liver transplantation （left lateral segment and caudate lobe） from her son. No recurrent lesion has been observed at two years after the liver transplantation. Liver transplantation should be considered as a treatment option even in advanced case of carcinoid metastasis to the liver. We also discuss the literature on liver transplantation for metastatic carcinoid tumor.

Multimodality Treatment for Thymic Carcinoma: Review of 11 Cases at a Single Institute

Background: We reported our experience with thymic carcinomas and review their clinical features, treatment strategies, and prognoses. Methods: From April 1998 to November 2012, 11 patients pathologically diagnosed with thymic carcinoma and treated in our hospital were investigated. Results: There were 7 men and 4 women, with a median age of 62 years (range, 35 - 72). According to the Masaoka staging system, 3 patients had stage II, 1 stage III disease, 3 stage IVa disease and 4 stage IVb disease. Ten patients had squamous cell carcinoma, whereas 1 had large cell neuroendocrine carcinoma (LCNEC). We performed surgery or multimodality therapy including surgery as the initial therapy for 8 patients. Of the non-surgical cases, 1 patient received chemoradiotherapy and survived for over 6 years without recurrence, whereas 2 received palliative care. Three of 4 patients who underwent complete resection survived without disease recurrence, whereas only 1 patient with LCNEC survived in the incomplete resection group. Multimodality therapy with cisplatin and docetaxel was provided to 3 patients, and recurrence has not been observed in any of the cases. Conclusions: Favorable outcomes could be achieved in patients with thymic carcinoma who underwent intensive treatment. In particular, surgery combined with cisplatin and docetaxel plus thoracic irradiation may be an attractive approach for thymic carcinoma.

Extended therapy duration for therapy-refractory hepatitis C patients with genotype 2

We devised an extended 72-wk peginterferon--2a/ribavirin therapy regimen for the retreatment of highly intractable cases,i.e.,48-wk peginterferon--2b/ribavirin therapy-intractable cases.Although 2 cases achieved a rapid virological response to 72-wk peginterferon--2a/ribavirin therapy,1 case failed to achieve a sustained virological response.Although the reason for this difference in the effectiveness of 72-wk peginterferon--2a/ribavirin therapy between the cases was unclear,the rebound phenomenon of serum transaminase after48-wk peginterferon--2b/ribavirin therapy and the resultant lower viral load compared to that before 48-wk peginterferon--2b/ribavirin therapy might have influenced the treatment outcome.Thus,it may be beneficial to consider the rebound phenomenon of serum transaminase and the changes in viral load resulting from previous interferon-based therapy and then cau-tiously determine the indication and the timing of the administration of 72-wk peginterferon--2a/ribavirin in highly intractable cases.Further studies should be performed to confirm this strategy.

Sofosbuvir/Ribavirin therapy for patients experiencing failure of ombitasvir/paritaprevir/ritonavir + ribavirin therapy: Two cases report and review of literature

BACKGROUND The effectiveness of sofosbuvir/ribavirin(SOF/RBV) combination therapy,which is one of the 1 st-choice therapeutic options for patients with hepatitis C virus(HCV) genotype 2(HCV-G2) in Japan according to the most recent version of the Japan Society of Hepatology guideline, for patients who experienced failure of the ombitasvir/paritaprevir/ritonavir plus ribavirin(OBV/PTV/r+RBV) combination therapy, which was another option for patients with HCV-G2, is unknown.CASE SUMMARY We evaluated the effects of SOF/RBV combination therapy in two patients with genotype 2 a who could not achieve a sustained virological response(SVR) by OBV/PTV/r+RBV combination therapy. One patient was complicated with VogtKoyanagi-Harada(VKH) disease. Resistance-associated variations before SOF/RBV combination therapy were not detected in two patients. Both patients had an SVR at 12 wk after the treatment(SVR12). Regarding adverse events(AEs), itching, chill, a dull feeling in the throat and cough as well as increase of alanine transaminase level were shown in one patient, while a headache and deterioration of light aversion probably due to the recurrence of VKH disease were shown in the other patients. In addition, the latter patient developed arthralgia and morning stiffness approximately 7 wk after the therapy and turned out to be diagnosed with rheumatoid arthralgia.CONCLUSION SOF/RBV therapy might be effective for patients experiencing failure of OBV/PTV/r+RBV therapy, but caution should be taken regarding the AEs.

Triple therapy of interferon and ribavirin with zinc supplementation for patients with chronic hepatitis C:A randomized controlled clinical trial

AIM： To study the therapeutic effect of interferon （IFN） and ribavirin with zinc supplement on patients with chronic hepatitis C viral （HCV） infection. METHODS： A total of 102 patients confirmed histologically to have chronic HCV infection with genotype lb and more than 100 KIU/mL of HCV were randomly assigned to each arm of the study and each received 10 million units of pegylated interferon （IFN- alpha-2b） daily for 4 wk followed by the same dose every other day for 20 wk plus ribavirin （600 or 800 mg/d depending on body weight）, with or without polaprezinc （150 mg/d） orally for 24 wk. The primary endpoint was sustained virological response （SVR） defined as negative HCV-RNA in the serum 6 mo after treatment. RESULTS： There were no differences in the clinical background between the two groups except for more females in the dual therapy group than in the other group （P〈 0.05）. SVR was observed in 33.3% of the triple therapy group and 33.3% of the dual therapy group. The side effects were almost the same in both groups except for gastrointestinal symptoms, which were less in the triple therapy group （P= 0.019）. CONCLUSION： Considered together, triple therapy of zinc plus IFN and ribavirin for HCV infection patients with genotype lb and high viral load is not better than dual therapy except for lower incidence of gastrointestinal side effects.

Different outcomes of nosocomial infection with hepatitis C virus from the same origin

The outcome of infection with hepatitis C virus （HCV） varies substantially from self-limiting infection to chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma among the individuals. The mechanisms that determine the clearance or the persistence of HCV have not yet been clarified. Here, we experienced two cases of hospital-related infection with HCV from the same origin but with quite different outcomes. One case resolved after an episode of acute hepatitis, while the other case developed a chronic hepatitis although they were infected with HCV of the same origin. Although infected with the virus of the same origin, the clinical and virological courses were completely different. This suggests that host factors play a major role in conditioning the outcome of acute HCV infection.