Therapeutic potential of berberine in attenuating cholestatic liver injury: insights from a PSC mouse model.

BACKGROUND AND AIMS: Primary sclerosing cholangitis (PSC) is a chronic liver disease characterized by progressive biliary inflammation and bile duct injury. Berberine (BBR) is a bioactive isoquinoline alkaloid found in various herbs and has multiple beneficial effects on metabolic and inflammatory diseases, including liver diseases. This study aimed to examine the therapeutic effect of BBR on cholestatic liver injury in a PSC mouse model (Mdr2-/- mice) and elucidate the underlying mechanisms. METHODS: Mdr2-/-mice (12-14 weeks old, both sexes) received either BBR (50 mg/kg) or control solution daily for eight weeks via oral gavage. Histological and serum biochemical analyses were used to assess fibrotic liver injury severity. Total RNAseq and pathway analyses were used to identify the potential signaling pathways modulated by BBR in the liver. The expression levels of key genes involved in regulating hepatic fibrosis, bile duct proliferation, inflammation, and bile acid metabolism were validated by qRT-PCR or Western blot analysis. The bile acid composition and levels in the serum, liver, small intestine, and feces and tissue distribution of BBR were measured by LC-MS/MS. Intestinal inflammation and injury were assessed by gene expression profiling and histological analysis. The impact on the gut microbiome was assessed using 16S rRNA gene sequencing. RESULTS: BBR treatment significantly ameliorated cholestatic liver injury, evidenced by decreased serum levels of AST, ALT, and ALP, and reduced bile duct proliferation and hepatic fibrosis, as shown by H&E, Picro-Sirius Red, and CK19 IHC staining. RNAseq and qRT-PCR analyses indicated a substantial inhibition of fibrotic and inflammatory gene expression. BBR also mitigated ER stress by downregulating Chop, Atf4 and Xbp-1 expression. In addition, BBR modulated bile acid metabolism by altering key gene expressions in the liver and small intestine, resulting in restored bile acid homeostasis characterized by reduced total bile acids in serum, liver, and small intestine and increased fecal excretion. Furthermore, BBR significantly improved intestinal barrier function and reduced bacterial translocation by modulating the gut microbiota. CONCLUSION: BBR effectively attenuates cholestatic liver injury, suggesting its potential as a therapeutic agent for PSC and other cholestatic liver diseases.

Sex- and reproductive status-specific relationships between body composition and non-alcoholic fatty liver disease.

BACKGROUND: Sex and reproductive status differences exist in both non-alcoholic fatty liver disease (NAFLD) and body composition. Our purpose was to investigate the relationship between body composition and the severity of liver steatosis and fibrosis in NAFLD in different sex and reproductive status populations. METHODS: This cross-sectional study included 880 patients (355 men, 417 pre-menopausal women, 108 post-menopausal women). Liver steatosis and fibrosis and body composition data were measured using FibroScan and a bioelectrical impedance body composition analyzer (BIA), respectively, and the following parameters were obtained: liver stiffness measurement (LSM), controlled attenuation parameter (CAP), waist circumference (WC), body mass index (BMI), percent body fat (PBF), visceral fat area (VFA), appendicular skeletal muscle mass (ASM), appendicular skeletal muscle mass index (ASMI), fat mass (FM), fat free mass (FFM), and FFM to FM ratio (FFM/FM). Multiple ordinal logistic regression (MOLR) was used to analyze the independent correlation between body composition indicators and liver steatosis grade and fibrosis stage in different sex and menopausal status populations. RESULTS: Men had higher WC, ASM, ASMI, FFM, and FFM/FM than pre- or post-menopausal women, while pre-menopausal women had higher PBF, VFA, and FM than the other two groups (p < 0.001). Besides, men had greater CAP and LSM values (p < 0.001). For MOLR, after adjusting for confounding factors, WC (OR, 1.07; 95% CI, 1.02-1.12; P = 0.011) and FFM/FM (OR, 0.52; 95% CI, 0.31-0.89; P = 0.017) in men and visceral obesity (OR, 4.16; 95% CI, 1.09-15.90; P = 0.037) in post-menopausal women were independently associated with liver steatosis grade. WC and visceral obesity were independently associated with liver fibrosis stage in men (OR, 1.05; 95% CI, 1.01-1.09, P = 0.013; OR, 3.92; 95% CI, 1.97-7.81; P < 0.001, respectively). CONCLUSIONS: Increased WC and low FFM/FM in men and visceral obesity in post-menopausal women were independent correlates of more severe liver steatosis. In addition, increased WC and visceral obesity were independent correlates of worse liver fibrosis in men. These data support the sex- and reproductive status-specific management of NAFLD.

Corrigendum: Platelet abnormalities in autoimmune thyroid diseases: A systematic review and meta-analysis.

[This corrects the article DOI: 10.3389/fimmu.2022.1089469.].

Accuracy of controlled attenuation parameter (CAP) and liver stiffness measurement (LSM) for assessing steatosis and fibrosis in non-alcoholic fatty liver disease: A systematic review and meta-analysis.

Background: Non-alcoholic fatty liver disease (NAFLD) is a common chronic liver disease, and among the non-invasive tests, controlled attenuation parameter (CAP) and liver stiffness measurement (LSM) have shown better diagnostic performance in NAFLD. This meta-analysis aimed to evaluate the performance of CAP and LSM for assessing steatosis and fibrosis in NAFLD. Methods: We searched the PubMed, Web of Science, Cochrane Library, and Embase databases for relevant articles published up to February 13th, 2022, and selected studies that met the inclusion and exclusion criteria, and evaluated the quality of evidence. Then we pooled sensitivity (SE), specificity (SP), and area under receiver operating characteristic (AUROC) curves. A random effect model was applied regardless of heterogeneity. Meta-regression analysis and subgroup analysis were performed to explore heterogeneity, and Fagan plot analysis was used to evaluate clinical utility. This meta-analysis was completed in Nanjing, Jiangsu and registered on PROSPERO (CRD42022309965). Findings: A total of 10537 patients from 61 studies were included in our meta-analysis. The AUROC of CAP were 0·924, 0·794 and 0·778 for steatosis grades ≥ S1, ≥ S2 and = S3, respectively, and the AUROC of LSM for detecting fibrosis stages ≥ F1, ≥ F2, ≥ F3, and = F4 were 0·851, 0·830, 0·897 and 0·925, respectively. Subgroup analysis revealed that BMI ≥ 30 kg/m² had lower accuracy for diagnosing S ≥ S1, ≥ S2 than BMI<30 kg/m². For the mean cut-off values, significant differences were found in CAP values among different body mass index (BMI) populations and LSM values among different regions. For diagnosing S ≥ S1, ≥ S2 and = S3, the mean CAP cut-off values for BMI ≥ 30 kg/m² were 30·7, 28·2, and 27·9 dB/m higher than for BMI < 30 kg/m² (P = 0·001, 0·001 and 0·018, respectively). For diagnosing F ≥ F2 and = F4, the mean cut-off values of Europe and America were 0·96 and 2·03 kPa higher than Asia (P = 0·027, P = 0·034), respectively. In addition, the results did not change significantly after sensitivity analysis and the trim and fill method to correct for publication bias, proving that the conclusions are robust. Interpretation: The good performance of CAP and LSM for the diagnosis of mild steatosis (S ≥ S1), advanced liver fibrosis (F ≥ F3), and cirrhosis (F = F4) can be used to screen for NAFLD in high-risk populations. Of note, the accuracy of CAP for the detection of steatosis in patients with obesity is reduced and requires specific diagnostic values. For LSM, the same diagnostic values can be used when the appropriate probes are selected based on BMI and the automated probe selection tool. The performance of CAP and LSM in assessing steatosis in patients with obesity, moderate to severe steatosis, and low-grade fibrosis should be further validated and improved in the future. Funding: The study was funded by the Priority Academic Program Development of Jiangsu Higher Education Institutions (PAPD).

Iptakalim improves cerebral microcirculation in mice after ischemic stroke by inhibiting pericyte contraction.

Pericytes are present tight around the intervals of capillaries, play an essential role in stabilizing the blood-brain barrier, regulating blood flow and immunomodulation, and persistent contraction of pericytes eventually leads to impaired blood flow and poor clinical outcomes in ischemic stroke. We previously show that iptakalim, an ATP-sensitive potassium (K-ATP) channel opener, exerts protective effects in neurons, and glia against ischemia-induced injury. In this study we investigated the impacts of iptakalim on pericytes contraction in stroke. Mice were subjected to cerebral artery occlusion (MCAO), then administered iptakalim (10 mg/kg, ip). We showed that iptakalim administration significantly promoted recovery of cerebral blood flow after cerebral ischemia and reperfusion. Furthermore, we found that iptakalim significantly inhibited pericytes contraction, decreased the number of obstructed capillaries, and improved cerebral microcirculation. Using a collagen gel contraction assay, we demonstrated that cultured pericytes subjected to oxygen-glucose deprivation (OGD) consistently contracted from 3 h till 24 h during reoxygenation, whereas iptakalim treatment (10 µM) notably restrained pericyte contraction from 6 h during reoxygenation. We further showed that iptakalim treatment promoted K-ATP channel opening via suppressing SUR2/EPAC1 complex formation. Consequently, it reduced calcium influx and ET-1 release. Taken together, our results demonstrate that iptakalim, targeted K-ATP channels, can improve microvascular disturbance by inhibiting pericyte contraction after ischemic stroke. Our work reveals that iptakalim might be developed as a promising pericyte regulator for treatment of stroke.

Platelet abnormalities in autoimmune thyroid diseases: A systematic review and meta-analysis.

Background: Some degree of platelet index abnormality has been found clinically in the autoimmune thyroid disease (AITD), but the findings are not uniform. Methods: The PubMed, Web of Science, Cochrane Library, and Embase databases were searched for relevant articles published up to August 16th, 2022, with no restrictions on the language of the articles. Reference lists of eligible articles were also searched. A random effect model was used to pool the standardized mean difference (SMD) and 95% confidence interval (95% CI) of platelet count (PLT), mean platelet volume (MPV), and platelet distribution width (PDW) between AITD patients and healthy controls, and subgroup analyses were performed. Results: A total of 19 articles with 6173 people (3824 AITD patients and 2349 healthy people) were included in the meta-analysis. The results showed that PLT and MPV values were significantly increased in AITD patients when compared with healthy people (SMD: 0.164, 95% CI: 0.044 to 0.285; SMD: 0.256, 95% CI: 0.013 to 0.500), while no significant difference was found in PDW between the AITD group and the control group (SMD: 0.060, 95% CI: -0.164 to 0.284). Subgroup analysis according to disease type and thyroid function revealed that for PLT, this difference was only found in the Hashimoto's thyroiditis (HT) and hypothyroid groups, but not in the Graves' disease (GD) and hyperthyroid groups. For MPV, the results were the opposite of those for PLT: MPV was significantly higher in the GD, hyperthyroid, and euthyroid groups than in the control group, but not in the HT and hypothyroid groups. Sensitivity analysis showed that the stability of the pooled MPV was not good. No publication bias was found. Conclusions: PLT and MPV are significantly elevated in patients with AITD, with PLT being more significantly elevated in HT and hypothyroidism, and MPV being more significantly increased in GD and hyperthyroidism. Appropriate clinical attention can be paid to the thyroid function of patients when abnormal platelet indices are found, and conversely, the consequences of abnormal platelet parameters such as elevated MPV lead to an increased occurrence of cardiovascular events, which should also be addressed in the AITD population. Systematic review registration: https://www.crd.york.ac.uk/PROSPERO/, identifier CRD42022341823.

Prognostic significance of ERCC1 expression in postoperative patients with gastric cancer.

AIM: This study explored the correlation between the expression of excision repair cross-complementation group 1 (ERCC1) and the prognosis of gastric cancer patients. METHODS: From January 2005 to December 2008, 605 patients who underwent radical surgery in The First Affiliated Hospital of Nanjing Medical University were enrolled. We conducted the follow-up every 6 months and its contents included a comprehensive medical history, tumor markers and abdominal ultrasound or CT and other imaging findings. Deadline was April 30, 2013 and follow-up time between 51 to 91 months. Survival time is calculated from the date of diagnosis to death or last follow-up date. Immunohistochemistry (IHC) was used to assess the expression of ERCC1 in resected samples. The relationship between ERCC1 expression and survival of patients was investigated. The comparison of count data were analyzed by Chi-square test. Median survival time (MST) and the 5-year survival rate were calculated by life table analysis. The Kaplan-Meier curves were used for survival analysis. RESULTS: ERCC1 expression was positive in 412 patients (68.1%). There is no significant difference between ERCC1-positive group and ERCC1-negative group in terms of the MST and 5-year survival rate (P=0.455). The MST and 5-year survival rate have no significant difference (P=0.162) between group with chemotherapy and group with no chemotherapy in patients with ERCC1-positive expression. However, the MST and 5-year survival rate in patients with ERCC1-negative expression benefited more from with chemotherapy (P=0.019). The ERCC1-positive patients survived longer than those ERCC1-negative patients (P=0.183) in subgroup with no adjuvant chemotherapy. In the subgroup analysis, ERCC1 expression had no significant relationship with overall survival in patients with stage II or III gastric cancer (P>0.05). CONCLUSIONS: ERCC1 might be a good prognostic factor for the patients of gastric cancer after radical resection. Patients with ERCC1-negative expression could benefit more from adjuvant chemotherapy.

[Toll-like receptor gene polymorphisms and susceptibility to inflammatory bowel disease in Chinese Han and Caucasian populations].

OBJECTIVE: To explore the distribution of Toll-like receptors gene polymorphisms in inflammatory bowel disease (IBD) in Chinese Han patients and Caucasians. METHODS: The toll-like receptor 2 (TLR2) genes Arg677Trp and Arg753Glu, TLR4 genes Asp299Gly and Thr399Ile, and TLR9 gene 1237T/C polymorphisms were genotypes in 113 patients with IBD and 120 age and gender-matched healthy controls by the analyses of polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP). A meta-analysis was performed to test whether TLR4 Asp299Gly and Thr399Ile polymorphisms were associated with ulcerative colitis (UC) or Crohn's disease (CD) susceptibility and whether 299Gly carriage was associated with phenotypes of CD patients in the Caucasian population. RESULTS: We found two carriers of TLR9 1237C in UC patients, one carrier in CD patients and one in healthy controls respectively (CD: P = 0.361; UC: P = 0.569). There was no statistically significant difference in both allelic and genotypic frequencies. The mutant genotypes of TLR2 gene Arg677Trp and Arg753Glu, TLR4 gene Asp299Gly and Thr399Ile were not found in either the IBD patients or the healthy controls. The TLR4 299G allele showed a significant association with CD and UC in Caucasian population (OR = 1.29, 95%CI: 1.08 - 1.54, P = 0.004 and OR = 1.28, 95%CI: 1.08 - 1.51, P = 0.004 respectively). Similar association was detected between T399I polymorphism and susceptibility to IBD (OR = 1.37, 95%CI: 1.12 - 1.68, P = 0.002 and OR = 1.46, 95%CI: 1.13 - 1.88, P = 0.003 respectively). However, no significant association was identified between CD phenotypes and 299Gly carriage. CONCLUSION: TLR2 genes Arg677Trp and Arg753Glu, TLR4 genes Asp299Gly and Thr399Ile and TLR9 gene 1237T/C polymorphisms are not associated with IBD in Chinese Han patients. In Caucasians, both TLR4 299G and 399I confer a significant risk for developing CD and UC. The contribution of genetic determinants may differ significantly between ethnicities.

Iptakalim ameliorates MPP+-induced astrocyte mitochondrial dysfunction by increasing mitochondrial complex activity besides opening mitoK(ATP) channels.

In addition to the established role of the mitochondrion in energy metabolism, regulation of cell death has been regarded as a major function of this organelle. Our previous studies have demonstrated that iptakalim (IPT), a novel ATP-sensitive potassium channel (K(ATP) channel) opener, protects against 1-methyl-4-phenyl-pyridinium ion (MPP+)-induced astrocyte apoptosis via mitochondria and mitogen-activated protein kinase signal pathways. The present study aimed to investigate whether IPT can protect astrocyte mitochondria against MPP+-induced mitochondrial dysfunction. We showed that treatment with IPT could ameliorate the inhibitory effect of MPP+ on mitochondrial respiration and ATP production by using mitochondrial complex I-supported substrates. IPT could also inhibit the increased production of mitochondrial reactive oxygen species (ROS) and the release of cytochrome c from mitochondria induced by MPP+. However, mitochondrial ATP-sensitive potassium (mitoK(ATP)) channel blocker 5-hydroxydecanoate (5-HD) could partly abolish all of the above effects of IPT. Because mitochondrial complex dysfunction impairs mitochondrial respiration and ATP production, a further experiment was undertaken to study the effects of IPT on the activity of mitochondrial complex (COX) I and COX IV. It was found that IPT inhibited the decrease in mitochondrial COX I and COX IV activity induced by MPP+, but 5-HD failed to abolish these effects. Taken together, these findings suggest that IPT may protect astrocyte mitochondrial function by regulating complex activity in addition to opening mitoK(ATP) channels.

Neuroprotective effects of berberine on stroke models in vitro and in vivo.

Berberine is an alkaloid derived from herb medicine Coptidis Rhizom. Although there are increasing evidences that berberine exhibits neuroprotective effects against ischemic brain damage, little is known about the mechanism. In this study, we investigated the effect of berberine on ischemic injury in a middle cerebral artery occlusion (MCAO) model. We found that berberine improved neurological outcome and reduced ischemia/reperfusion (I/R)-induced cerebral infarction 48h after MCAO. The protective effect of berberine was confirmed in in vitro study. Berberine protected PC12 cells against oxygen-glucose deprivation (OGD)-induced injury. The results showed that berberine inhibited reactive oxygen species (ROS) generation, and subsequent release of pro-apoptotic factor cytochrome c and apoptosis-inducing factors (AIFs) evoked by OGD. Findings of this study suggest that berberine protects against ischemic brain injury by decreasing the intracellular ROS level and subsequently inhibiting mitochondrial apoptotic pathway.

ATP-sensitive potassium channel opener iptakalim protects against MPP-induced astrocytic apoptosis via mitochondria and mitogen-activated protein kinase signal pathways.

Inhibition of astrocytic apoptosis has been regarded as a novel prospective strategy for treating neurodegenerative disorders such as Parkinson's disease. In the present study, we demonstrated that iptakalim (IPT), an ATP-sensitive potassium channel (K(ATP) channel) opener, exerted protective effect on MPP(+)-induced astrocytic apoptosis, which was reversed by selective mitochondrial K(ATP) channel blocker 5-hydroxydecanoate. Further study revealed that IPT inhibited glutathione (GSH) depletion, mitochondrial membrane potential loss and subsequent release of pro-apoptotic factors (cytochrome c and apoptosis-inducing factor (AIF), and c-Jun NH(2)-terminal kinase/mitogen-activated protein kinases (MAPK) phosphorylation induced by MPP(+). Meanwhile, extracellular signal-regulated kinase (ERK) 1/2 inhibitor PD98059 inhibited the protective effect of IPT on MPP(+)-induced astrocytic apoptosis. Furthermore, IPT could also activate ERK/MAPK and maintain increased phospho-ERK1/2 level after MPP(+) exposure. Taken together, these findings reveal for the first time that IPT protects against MPP(+)-induced astrocytic apoptosis via inhibition of mitochondria apoptotic pathway and regulating the MAPK signal transduction pathways by opening mitochondrial ATP-sensitive potassium (mitoK(ATP)) channels in astrocytes. And targeting K(ATP) channels expressed in astrocytes may provide a novel therapeutic strategy for neurodegenerative disorders.

Alterations of striatal neurotransmitter release in aquaporin-4 deficient mice: An in vivo microdialysis study.

Our previous investigation has demonstrated that the lack of aquaporin-4 (AQP4) expressions in mice is paralleled by sex- and region-specific abnormalities in neurotransmission. In the present study, we examined the effect of AQP4 deficiency on release of neurotransmitters in mouse striatum using in vivo microdialysis after high K(+) stimulus. The results showed that neurotransmitter releases under the basal and K(+)-stimulated conditions in the striatum of wildtype mice were similar to AQP4 knockout mice, except for taurine, when measured at 24h after microdialysis surgery. However, the basal extracellular levels of dopamine and its metabolites were significantly increased in knockout mice, followed by reduced or no response to depolarizing stimuli when measured at 7 d after surgery. In addition, it was found that there were higher responses of amino acids to high K(+) stimulus in knockout mice. This experiment provides the in vivo evidence that AQP4 participates in the regulation of neurotransmitter release induced by depolarizing stimuli.