Porphyromonas gingivalis as a promotor in the development of the alcoholic liver disease via ferroptosis.

Alcoholic liver disease (ALD) is a liver disease caused by heavy drinking. Porphyromonas gingivalis (P.g), a major cause of periodontitis, whose antibodies are elevated in severe ALD patients in the plasma. The purpose of this study is to further study the role and the molecular mechanism of P.g in the progress of ALD. In this study, saliva of patients with ALD was collected. Then, an animal model of ALD with oral P.g administration was established, pathology of liver and spleen, intestinal microorganisms and metabolites were analyzed. The molecular mechanism of P.g on ALD was analyzed in vitro. ALD and intestinal microflora and metabolite changes were observed more serious in the alcohol and P.g groups than the alcohol group. Moreover, ferroptosis was aggravated by P.g in the liver. Meanwhile, P.g promoted ferroptosis accomplication with alcohol in vitro, which can be reversed by ferroptosis inhibitors. In conclusion, P.g aggravates ALD through exacerbation gut microbial metabolic disorder in mice with alcohol, which maybe depend on ferroptosis activation in hepatocytes. The study provides a new strategy for prevention and treatment of ALD by improving the oral micro-environment.

Prognostic Value and the Immune Microenvironment-associated Role of SMAD4 in Pancreatic Adenocarcinoma.

BACKGROUND/AIM: SMAD4 is a well-known cancer suppressor gene that regulates cell proliferation, differentiation, autoimmunity, pluripotency, and immune responses in pancreatic adenocarcinoma (PAAD). We herein investigated a novel involvement of SMAD4 within the PAAD microenvironment. MATERIALS AND METHODS: Transcriptome data, derived from The Cancer Genome Atlas and Genotype-Tissue Expression Using TIMER 2.0, CIBERSORT, and ImmuCellAI, were used to identify the immune cell infiltration pattern of PAAD. We then knocked-down SMAD4 in the PANC-1 cell line and acquired RNA-seq data through the Illumina microarray technology. Kyoto Encyclopedia of Genes and Genomes was utilized along with Gene Ontology enrichment analyses, and protein-protein interaction network analysis to screen for genes that were differentially expressed. We constructed a miRNA-mRNA regulatory network and analyzed SMAD4 copy number variation (CNV) data. RESULTS: In PAAD, decreased levels of SMAD4 expression were found to be connected to an unfavorable prognosis. There was a significantly higher infiltration level of DC cells, CD8+ T cells, TgD, Tc, and Tex cells and a lower level of B cells and Th2 in the SMAD4-high group. The expression of SMAD4 and immune cell infiltration including CD8+ T cells, myeloid dendritic cells, neutrophils, and macrophages are significantly positively correlated. DEGs were found enriched in the hypoxia response pathway. The six hypoxia-related genes exhibited a significant correlation with immune cell infiltration and survival rates. SMAD4 CNV levels were associated with MSI, stemness, infiltration of immune cells, and survival rates. CONCLUSION: A statistically significant correlation was found between SMAD4 expression and immune cell infiltration. SMAD4 could mediate hypoxia response in pancreatic cancer. The CNV levels of SMAD4 were associated with prognosis. SMAD4 has potential as a prognostic biomarker and provides a new orientation for the immunotherapy of PAAD.

Proanthocyanidin Alleviates Liver Ischemia/Reperfusion Injury by Suppressing Autophagy and Apoptosis via the PPARα/PGC1α Signaling Pathway.

Background and Aims: Hepatic ischemia-reperfusion injury (IRI) is a common pathophysiological phenomenon in clinical practice, which usually occurs in liver transplantation, liver resection, severe trauma, and hemorrhagic shock. Proanthocyanidin (PC), exerted from various plants with antioxidant, antitumor, and antiaging activity, were administrated in our study to investigate the underlying mechanism of its protective function on IRI. Methods: Two doses of PC (50 mg/kg, 100 mg/kg) were given to BALB/c mice by intragastric administration for 7 days before partial (70%) warm IR surgery. Serum and liver tissues were collected 2, 8, and 24 h after reperfusion for relevant experiments. Results: The results of transaminase and hematoxylin and eosin staining indicated that PC pretreatment significantly alleviated IRI in mice. Serum total superoxide dismutase increased and malondialdehyde decreased in PC pretreatment groups. Enzyme-linked immunosorbent assays, western blotting, quantitative real-time polymerase chain reaction, and immunohistochemistry showed that inflammation, apoptosis, and autophagy in PC preprocessing groups were significantly inhibited and were dose-dependent. The protein, mRNA expression, and immunohistochemical staining results of peroxisome proliferator-activated receptor alpha (PPARα) and peroxisome proliferator-activated receptor gamma coactivator 1-alpha (PGC1α) in the PC pretreatment groups were significantly upregulated compared with the IR group in a dose-dependent manner. Conclusions: PC pretreatment suppressed inflammation, apoptosis, and autophagy via the PPAR-α signaling pathway to protect against IRI of the liver in mice.

Identification and Characterization of a Novel Nanobody Against Human CTGF to Reveal Its Antifibrotic Effect in an in vitro Model of Liver Fibrosis.

Background: No agents are currently available for the treatment or reversal of liver fibrosis. Novel antifibrotic therapies for chronic liver diseases are thus urgently needed. Connective tissue growth factor (CTGF) has been shown to contributes profoundly to liver fibrogenesis, which makes CTGF as a promising target for developing antifibrotic agents. Methods: In this study, we identified a novel nanobody (Nb) against human CTGF (anti-CTGF Nb) by phage display using an immunized camel, which showed high affinity and specificity in vitro. LX-2 cells, the immortalized human hepatic stellate cells, were induced by transforming growth factor beta1 (TGFß1) as an in vitro model of liver fibrosis to verify the antifibrotic activity of the anti-CTGF Nb. Results: Our data demonstrated that anti-CTGF Nb effectively alleviated TGFß1-induced LX-2 cell proliferation, activation, and migration, and promoted the apoptosis of activated LX-2 cells in response to TGFß1. Moreover, the anti-CTGF Nb remarkably reduced the levels of TGFß1, Smad2, and Smad3 expression in LX-2 stellate cells stimulated by TGFß1. Conclusion: Taken together, we successfully identified a novel Nb against human CTGF, which exhibited antifibrotic effects in vitro by regulating the biological functions of human stellate cells LX-2.

The Role of miR-1183: A Potential Suppressor in Hepatocellular Carcinoma via Regulating Splicing Factor SRSF1.

Purpose: Hepatocellular carcinoma (HCC) is a severe global health problem, causing many deaths of patients all over the world. Serine and arginine-rich splicing factor 1 (SRSF1) functions as an important oncogenic role in tumorigenesis and progression in HCC. Therefore, therapies targeting SRSF1 may provide promising therapeutic approaches. MiRNAs are virtually involved at the post-transcriptional level and bind to 3' untranslated region (3'-UTR) of their target messenger RNA (mRNA) to suppress expression. Methods: Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to detect the expression of SRSF1 and miR-1183 in HCC cell lines. CCK8 assay, colony formation assay and wound healing assay were used to detect the function of miR-1183 in HCC cell lines in vitro. Luciferase reporter assay and Western blot were applied to detect the regulation of particular molecules. Xenograft tumor assay was used to detect the function of miR-1183 in HCC cell lines in vivo. Immunohistochemistry (IHC) was used to detect the expression of SRSF1 in HCC tissues and Xenograft tumors. Results: In this study, we identified that miR-1183 was downregulated in HCC cell lines. Functional assays indicated that miR-1183-upregulation cells show weakened proliferation ability and migration ability in vitro and inhibit subcutaneous tumor formation in vivo. With respect to the underlying mechanism, we found that miR-1183 function as a tumor suppressor by specifically binding to SRSF1. Conclusion: This study is the first to demonstrate that miR-1183 function as an important tumor-suppressing role by binding to the 3'-UTR of SRSF1 mRNA and suppressing its protein level in HCC cells in vitro and in vivo. Further, miR-1183 may be a potential target in the prognosis and treatment of HCC.

Promotion of colorectal cancer cell death by ezetimibe via mTOR signaling-dependent mitochondrial dysfunction.

Introduction: Colorectal cancer (CRC) is the fourth most common cancer worldwide, with high morbidity and mortality rates. In recent years, high-fat diet has been shown to increase CRC morbidity, highlighting the possibility of the application of hypolipidemic drugs for CRC treatment. In this study, we preliminarily evaluated the effects and mechnisms of ezetimibe against CRC through the blockage of lipid absorption in small intesine. Methods: In this study, CRC cell proliferation, invasion, apoptosis, and autophagy were evaluated using cellular and molecular assays. Fluorescent microscopy, and a flow cytometric assay were used to assess mitochondrial activity in vitro. A subcutaneous xenograft mouse model was used to evaluate the effects of ezetimibe in vivo. Results: We found that ezetimibe inhibited CRC cell proliferation, and migration, and facilitated autophage-associated apoptosis in HCT116 and Caco2 cells. Ezetimibe-induced mitochondrial dysfunction in CRC cells was found to be correlated with mTOR signaling activity. Discussion: Ezetimibe exhibits effects against CRC through the promotion of cancer cell death via mTOR signaling-dependent mitochondrial dysfunction, highlighting its potential value in CRC therapy.

Hepatic ER stress suppresses adipose browning through ATF4-CIRP-ANGPTL3 cascade.

Hepatic endoplasmic reticulum (ER) stress is a hallmark of obesity-induced liver steatosis and contributes to the progress of steatosis and insulin resistance in liver. However, its influence on adipose function is still unclear. Here, we identify a hepatic ER stress-induced activating transcription factor 4 (ATF4)-cold-inducible RNA-binding protein (CIRP)-angiopoietin-related protein3 (ANGPTL3) cascade critical for the regulation of adipose browning. We find that obesity increases CIRP expression in liver through ER stress-induced ATF4. CIRP in turn binds to the 3' UTR and increases mRNA stability of ANGPTL3. ANGPTL3 secreted from liver suppresses uncoupling protein 1 expression through integrin αvß3 and c-Jun N-terminal kinase in adipose tissue. While hepatic expression of either ATF4, CIRP, or ANGPTL3 suppresses adipose browning, knockdown of CIRP and ANGPTL3 in liver or administration of integrin αvß3 inhibitor cilengitide increases adipose browning process. Taken together, we identify a communication mechanism to link hepatic ER stress and adipose browning that may imply a reciprocal regulation of obesity and liver steatosis.

Luteolin Pretreatment Attenuates Hepatic Ischemia-Reperfusion Injury in Mice by Inhibiting Inflammation, Autophagy, and Apoptosis via the ERK/PPARα Pathway.

Hepatic ischemia-reperfusion (IR) injury is a clinically significant process that frequently occurs in liver transplantation, partial hepatectomy, and hemorrhagic shock. The aim of this study was to explore the effectiveness of luteolin in hepatic IR injury and the underlying mechanism. BALB/c mice were randomly divided into six groups, including normal controls (NC), luteolin (50 mg/kg), sham procedure, IR+25 mg/kg luteolin, and IR+50 mg/kg luteolin group. Serum and tissue samples were collected at 6 and 24 h after reperfusion to assay liver enzymes, inflammatory factors, expression of proteins associated with apoptosis and autophagy, and factors associated with the extracellular signal-regulated kinase/peroxisome proliferator-activated receptor alpha (ERK/PPARα) pathway. Luteolin preconditioning decreased hepatocyte injury caused by ischemia-reperfusion, downregulated inflammatory factors, and inhibited apoptosis and autophagy. Luteolin also inhibited ERK phosphorylation and activated PPARα.

Gastrodin ameliorates Concanavalin A-induced acute hepatitis via the IL6/JAK2/STAT3 pathway.

AIMS: Gastrodin, the main active ingredient of Gastrodia elata Blume, has been shown to protect against many inflammatory diseases. Our study aimed to investigate the anti-inflammatory role of gastrodin in concanavalin A (ConA)-induced acute hepatitis in mice and to explore its precise mechanism. METHODS: C57BL/6 mice were administered with gastrodin (50 or 100mg/kg) for 3 days prior to intravenous injection of ConA to induce acute autoimmune hepatitis (AIH). Serum aminotransferases levels and cytokine levels were measured. Liver tissue histology was conducted to assess the degree of liver injury. Splenocytes pretreated with gastrodin were stimulated with ConA to observe splenocyte proliferation. RESULTS: Gastrodin greatly reduced the level of serum aminotransferases, inflammatory cytokine such as IL-6 and TNF-α and histopathological damage in ConA-induced hepatitis. Besides, gastrodin had an inhibitory effect on liver apoptosis, and autophagy. Furthermore, gastrodin inhibited the proliferation of splenocytes in vitro. The protein expression of p-JAK2 and p-STAT3 was markedly affected by gastrodin pretreatment. CONCLUSIONS: The present study indicated that gastrodin pretreatment exerted protective effects against ConA-induced acute hepatitis, partly through the inhibition of the IL6/JAK2/STAT3 pathway. Further studies are recommended to determine the potential therapeutic role of gastrodin in acute AIH.

Role of eIF3C Overexpression in Predicting Prognosis of Intrahepatic Cholangiocarcinoma.

BACKGROUND: Elevated expression of eukaryotic initiation factor 3c (eIF3C) was recently uncovered to promote several types of cancer progression by inducing cell proliferation. Here, we aimed to assess the expression and prognostic value of eIF3C in intrahepatic cholangiocarcinoma (ICC) patients. METHODS: Expression of eIF3C was analyzed by immunohistochemistry in tissue microarrays (TMAs) containing 138 ICC and paired peritumoral tissues from ICC patients. Then, the roles of eIF3C in ICC cells were investigated by RNA interference, and the relationship between the eIF3C and KI67 expression was explored in ICC cells and tissues. Finally, the relation between the eIF3C level and clinicopathologic features of ICC was probed, and Kaplan-Meier and Cox's analyses were performed to assess the prognostic merit of eIF3C and KI67 in ICC patients. RESULTS: The expression of eIF3C was elevated in ICC tissues compared to paired peritumoral tissues, which was consistent with the result from the GEPIA database. The downregulation of eIF3C in ICC cells impaired the cellular invasion, metastasis, colony formation, and proliferation. Moreover, we further found a positive relationship between the eIF3C and KI67 expression in ICC cells and tissues. The expression of eIF3C in ICC tissues was positively correlated with lymphatic metastasis (p = 0.049), and the high level of KI67 was frequently found in ICC patients with the large tumor (p = 0.028), high serum AFP (p = 0.019), or lymphatic metastasis (p = 0.039). Notably, patients with the eIF3C or KI67 overexpression had shorter overall survival and higher disease-free survival rates than those with low expression of eIF3C or KI67, and the combination of eIF3C or KI67 expression was an independent parameter for predicting the prognosis and recurrence of ICC patients. CONCLUSIONS: Elevated eIF3C expression promotes ICC development, and combination of eIF3C and KI67 is a valuable predictor of the survival and recurrence of ICC patient.

Social skill training for children with autistic spectrum disorder on the uncertainty of the disease from mothers / 中国学校卫生

Objective@#To improve the social skills of children with ASD by using Program for the Education and Enrichment of Relational Skills(PEERS ), and to reduce the uncertainty towards ASD and negative emotions for mothers of ASD children.@*Methods@#From September to October 2017, 30 dyads of autistic mother and child were recruited and divided into intervention group and control group (15 mother child dyads each). Based on the content of PEERS social skill, cognitive behavior therapy was delivered in group format, through demonstration, role play and group exercise. At the same time, mother child dyads were trained using parallel social technology. Mothers and children with ASD were investigated using Parents Perception of Uncertainty Scale (PPUS), Patient Health Questionnaire 9 (PHQ-9), Chinese Version of the Beck Depression Inventory II(BDI-Ⅱ-C), Beck Anxiety Inventory (BAI), State Trait Anxiety Inventory(STAI-Form Y), and Autism Behavior Checklist (ABC), Cildhood Autism Rating Scale (CARS), and Social Communication Questionnaire (SCQ).@*Results@#Changes in ASD symptom score in children and emotional score of mothers in the intervention group were less than 0. The total score of mother disease uncertainty(74.93±13.58, 90.40± 9.21 ), ambiguity(31.13±7.07, 38.93±4.73), lack of clarity information(11.93±2.09, 13.80±2.54), unpredictability(9.60±1.99, 12.07±2.89), significantly changed after intervention( t =-3.65, -3.55, -2.20, -2.72, P <0.05).@*Conclusion@#Social PEERS group intervention can enhance the social skills of children with ASD, reduce uncertainty of illness among mother of ASD children. Timely disease related information, guidance for mothers to actively participate in child care and training, might help to reduce cognitive bias, depressive and anxiety symptoms among mothers.

circHMGCS1-016 reshapes immune environment by sponging miR-1236-3p to regulate CD73 and GAL-8 expression in intrahepatic cholangiocarcinoma.

BACKGROUND: Accumulating evidence indicates that circRNAs may serve as essential regulators in the progression of several human cancers, but the function and mechanism of circRNAs in intrahepatic cholangiocarcinoma (ICC) are largely unknown. METHODS: RNA-seq was used to assess differentially expressed circRNAs between 4 ICC and peritumor tissues. Quantitative RT-PCR and in situ hybridization were used to determine the circHMGCS1-016 expression in ICC tissues. The function and mechanism of circHMGCS1-016 were further identified via in vivo experiments. The clinical characteristics and prognostic significance of circHMGCS1-016 were analyzed by a retrospective study. The functions of circHMGCS1-016 were assessed via modifying circRNA expression in ICC cells. Moreover, the molecular mechanisms of circHMGCS1-016 in ICC cells were explored by circRNA precipitation, miRNA immunoprecipitation, SILAC and luciferase reporter assays. RESULTS: We identified that compared with peritumor tissues, ICC tissues expressed hsa_circ_0008621 (circHMGCS1-016) high by RNA-seq, which was further identified by qRT-PCR and in situ hybridization. Moreover, the expression of circHMGCS1-016 was revealed to be associated with survival and recurrence of ICC patients. By regulating circHMGCS1-016 expression, we found that elevated circHMGCS1-016 promoted ICC development both in vitro and in vivo. By SILAC and circRNA-pull down, we demonstrated that circHMGCS1-016 induced ICC cell invasion and reshaped the tumor immune microenvironment via the miR-1236-3p/CD73 and GAL-8 axis. In ICC tissues, we uncovered that a high level of circHMGCS1-016 was positively associated with CD73 and GAL-8 expression and negatively related to the CD8+ T cells infiltration, which was further validated by establishing a humanized mouse tumor model. Importantly, we displayed that ICC patients with high levels of circHMGCS1-016 in tumor tissues benefited less from anti-PD1 treatment compared to those with low levels of circHMGCS1-016. CONCLUSIONS: CircHMGCS1-016 is a forceful contributor in ICC development and immune tolerance via miR-1236-3p/CD73 and GAL-8 axis. CircHMGCS1-016 can be explored as a new potential biomarker and therapeutic target for PD1-resistant ICC.

Cellular based immunotherapy for primary liver cancer.

Primary liver cancer (PLC) is a common malignancy with high morbidity and mortality. Poor prognosis and easy recurrence on PLC patients calls for optimizations of the current conventional treatments and the exploration of novel therapeutic strategies. For most malignancies, including PLC, immune cells play crucial roles in regulating tumor microenvironments and specifically recognizing tumor cells. Therefore, cellular based immunotherapy has its instinctive advantages in PLC therapy as a novel therapeutic strategy. From the active and passive immune perspectives, we introduced the cellular based immunotherapies for PLC in this review, covering both the lymphoid and myeloid cells. Then we briefly review the combined cellular immunotherapeutic approaches and the existing obstacles for PLC treatment.

High level of CD73 predicts poor prognosis of intrahepatic cholangiocarcinoma.

Background: Despite recent improvements in the diagnosis and therapy of intrahepatic cholangiocarcinoma (ICC), the prognosis for ICC patients remains poor. Therefore, it is needed to identify new biological indicators for ICC progression. Methods: Immunohistochemistry was engaged to inspect the ecto-5'-nucleotidase (CD73) and CD8 expressions in tissue microarrays including tissues from 140 ICC patients. Then, the association between the level of CD73/CD8 and clinicopathologic characteristics of ICC was analysed. Finally, the prognostic value of CD73 and CD8 levels in ICC patients was assessed by Kaplan-Meier and multivariate and univariate analyses. Results: The CD73 expression was evidently upregulated in ICC tissues compared to the corresponding peritumoral tissues. The elevated CD73 expression was positively related to the lymphatic metastasis (p=0.049). While the level of tumour-infiltrating CD8 T+ cells in tumour tissues was negatively associated with serum AFP (p=0.019), tumor size (p=0.028), and lymphatic metastasis (p=0.039). Additionally, patients with elevated CD73 expression or low tumour-infiltrating CD8+ T cells exhibited shorter overall survival (OS) and higher disease-free survival (DFS) rates than patients with low CD73 expression and/or high tumour-infiltrating CD8+ T cells. Notably, the overexpression of CD73 or low tumour-infiltrating CD8+ T cells was an independent indicator for predicting the OS and DFS of ICC patients. Conclusions: We revealed that CD73 expression and low tumour-infiltrating CD8+T cells are valuable predictors of survival and recurrence in patients with ICC.

Rapid and precise diagnosis of pneumonia coinfected by Pneumocystis jirovecii and Aspergillus fumigatus assisted by next-generation sequencing in a patient with systemic lupus erythematosus: a case report.

BACKGROUND: Pneumocystis jirovecii and Aspergillus fumigatus, are opportunistic pathogenic fungus that has a major impact on mortality in patients with systemic lupus erythematosus. With the potential to invade multiple organs, early and accurate diagnosis is essential to the survival of SLE patients, establishing an early diagnosis of the infection, especially coinfection by Pneumocystis jirovecii and Aspergillus fumigatus, still remains a great challenge. CASE PRESENTATION: In this case, we reported that the application of next -generation sequencing in diagnosing Pneumocystis jirovecii and Aspergillus fumigatus coinfection in a Chinese girl with systemic lupus erythematosus (SLE). Voriconazole was used to treat pulmonary aspergillosis, besides sulfamethoxazole and trimethoprim (SMZ-TMP), and caspofungin acetate to treat Pneumocystis jirovecii infection for 6 days. On Day 10 of admission, her chest radiograph displayed obvious absorption of bilateral lung inflammation though the circumstance of repeated fever had not improved. Unfortunately, the patient discharged from the hospital since the financial burden, and during the follow-up, it was documented the patient died within one week after discharge. CONCLUSIONS: This successful application of the next generation sequencing assisting the rapid diagnosis of Pneumocystis jirovecii and Aspergillus fumigatus coinfection provides a new perspective in the clinical approach against the systematic fungi infections and highlights the potential of this technique in rapid etiological diagnosis.

Rox8 promotes microRNA-dependent yki messenger RNA decay.

The Hippo pathway is an evolutionarily conserved regulator of organ growth and tumorigenesis. In Drosophila, oncogenic RasV12 cooperates with loss-of-cell polarity to promote Hippo pathway-dependent tumor growth. To identify additional factors that modulate this signaling, we performed a genetic screen utilizing the Drosophila RasV12/lgl-/- in vivo tumor model and identified Rox8, a RNA-binding protein (RBP), as a positive regulator of the Hippo pathway. We found that Rox8 overexpression suppresses whereas Rox8 depletion potentiates Hippo-dependent tissue overgrowth, accompanied by altered Yki protein level and target gene expression. Mechanistically, Rox8 directly binds to a target site located in the yki 3' UTR, recruits and stabilizes the targeting of miR-8-loaded RISC, which accelerates the decay of yki messenger RNA (mRNA). Moreover, TIAR, the human ortholog of Rox8, is able to promote the degradation of yki mRNA when introduced into Drosophila and destabilizes YAP mRNA in human cells. Thus, our study provides in vivo evidence that the Hippo pathway is posttranscriptionally regulated by the collaborative action of RBP and microRNA (miRNA), which may provide an approach for modulating Hippo pathway-mediated tumorigenesis.

Role of bile acids in the diagnosis and progression of liver cirrhosis: A prospective observational study.

The accumulation of toxic bile acids (BAs) is closely related to liver injury, inflammation and tumorigenesis. The aim of the present study was to determine the role of the serum BA spectrum in the diagnosis and progression of liver cirrhosis. This was a prospective observational study involving patients with chronic hepatitis (n=23), liver cirrhosis (n=101), and cirrhosis complicated with hepatocellular carcinoma (CC-HCC; n=56). The 6-month survival of cirrhotic patients was recorded after blood collection. Comparisons of serum total BAs and individual BAs between different groups were performed using the Mann-Whitney U or Kruskal-Wallis tests. Correlation analysis was conducted by Spearman's correlation. Diagnosis and prediction analyses were performed using receiver operating characteristic curves. Survival was analyzed using the Kaplan-Meier method and multivariable Cox regression analysis. The concentrations of total BAs, glycocholic acid (GCA), glycochenodeoxycholic acid (GCDCA), taurocholic acid (TCA), taurochenoxycholic acid and tauroursodeoxycholic acid (TUDCA) were increased significantly in patients with early cirrhosis compared to patients with chronic hepatitis (P<0.05) and were associated with the diagnosis of cirrhosis (P=0.049, 0.004, 0.002, 0.003, 0.010 and 0.009, respectively). The levels of total BAs, primary conjugated BAs, and TUDCA increased as liver cirrhosis progressed (P<0.05). Serum total BAs, GCA, GCDCA, and TCA predicted the 6-month survival of patients with liver cirrhosis (P=0.0003, 0.005, 0.002, and 0.010 respectively). Based on multivariate Cox regression analysis, the level of total BAs was an independent predictor of mortality in cirrhotic patients (hazard ratios, 4.046; 95% CI, 1.620-10.108; P=0.003). In the early-stage cirrhosis group, the concentrations of total BAs and primary conjugated BAs were significantly elevated in patients with CC-HCC compared with patients with cirrhosis alone. In conclusion, total and individual BAs, especially primary conjugated BAs, are effective non-invasive markers in the diagnosis and prognosis of liver cirrhosis, and may be potential indicators in the occurrence of hepatocellular carcinoma in patients with early cirrhosis.

Isorhamnetin Inhibits Liver Fibrosis by Reducing Autophagy and Inhibiting Extracellular Matrix Formation via the TGF-ß1/Smad3 and TGF-ß1/p38 MAPK Pathways.

OBJECTIVE: Liver fibrosis is a consequence of wound-healing responses to chronic liver insult and may progress to liver cirrhosis if not controlled. This study investigated the protection against liver fibrosis by isorhamnetin. METHODS: Mouse models of hepatic fibrosis were established by intraperitoneal injection of carbon tetrachloride (CCl4) or bile duct ligation (BDL). Isorhamnetin 10 or 30 mg/kg was administered by gavage 5 days per week for 8 weeks in the CCl4 model and for 2 weeks in the BDL model. Protein and mRNA expressions were assayed by western blotting, immunohistochemistry, and quantitative real-time polymerase chain reaction. RESULTS: Isorhamnetin significantly inhibited liver fibrosis in both models, inhibiting hepatic stellate cell (HSC) activation, extracellular matrix (ECM) deposition, and autophagy. The effects were associated with downregulation of transforming growth factor ß1 (TGF-ß1) mediation of Smad3 and p38 mitogen-activated protein kinase (MAPK) signaling pathways. CONCLUSION: Isorhamnetin protected against liver fibrosis by reducing ECM formation and autophagy via inhibition of TGF-ß1-mediated Smad3 and p38 MAPK signaling pathways.

Alleviation of hepatic fibrosis and autophagy via inhibition of transforming growth factor-ß1/Smads pathway through shikonin.

BACKGROUND AND AIM: Liver fibrosis is a worldwide clinical challenge during the progression of chronic liver disease to liver cirrhosis. Shikonin is extracted from the root of Lithospermum erythrorhizon with antioxidant, anti-inflammatory, anticancer, and wound-healing properties. The study aims to investigate the protective effect of shikonin on liver fibrosis and its underlying mechanism. METHODS: Two liver fibrosis models were established in male C57 mice by intraperitoneal injection of CCl4 or bile duct ligation. Shikonin was administered orally three times weekly at a dose of 2.5 or 5 mg/kg. Protein and mRNA expressions were assayed by quantitative real-time polymerase chain reaction, Western blotting, and immunohistochemical staining. RESULTS: Shikonin significantly inhibited activation of hepatic stellate cells and extracellular matrix formation by downregulating the transforming growth factor-ß1 expression and maintaining the normal balance between metalloproteinase-2 and tissue inhibitor of metalloproteinase-1. Shikonin also decreased hepatic stellate cell energy production by inhibiting autophagy. CONCLUSIONS: The results confirmed that shikonin attenuated liver fibrosis by downregulating the transforming growth factor-ß1/Smads pathway and inhibiting autophagy.

Beraprost sodium preconditioning prevents inflammation, apoptosis, and autophagy during hepatic ischemia-reperfusion injury in mice via the P38 and JNK pathways.

OBJECTIVE: The goal of this study was to determine the effects of beraprost sodium (BPS) preconditioning on hepatic ischemia-reperfusion (IR) injury and its underlying mechanisms of action. MATERIALS AND METHODS: Mice were randomly divided into sham, IR, IR+BPS (50 µg/kg), and IR+BPS (100 µg/kg) groups. Saline or BPS was given to the mice by daily gavage for 1 week before the hepatic IR model was established. Liver tissues and orbital blood were collected at 2, 8, and 24 hours after reperfusion for the determination of liver enzymes, inflammatory mediators, apoptosis- and autophagy-related proteins, key proteins in P38 and c-Jun N-terminal kinase (JNK) cascades, and evaluation of liver histopathology. RESULTS: BPS preconditioning effectively reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels, improved pathological damage, ameliorated production of tumor necrosis factor-α (TNF-α) and interleukin-1ß (IL-1ß), and affected expressions of Bax, Bcl-2, Caspase-3, Caspase-8, and Caspase-9, microtubule-associated protein 1 light chain 3 (LC3), Beclin-1, and P62. The protective effects of BPS preconditioning were associated with reduced P38 and JNK phosphorylation. CONCLUSION: BPS preconditioning ameliorated hepatic IR injury by suppressing inflammation, apoptosis, and autophagy, partially via inhibiting activation of the P38 and JNK cascades.