Prevalence and effect on prognosis of sarcopenia in patients with primary biliary cholangitis.

Background: Sarcopenia adversely affects the treatment outcomes in Cirrhosis and NAFLD. However, such research is limited in primary biliary cholangitis (PBC) patients. This study was performed to examine the prevalence of sarcopenia and its impact on PBC patients' prognoses. Methods: This study enrolled confirmed PBC patients who had an abdominal CT scan. Sarcopenia was determined by the L3-skeletal muscle index with a Chinese population-based cut-off value. Laboratory test values and liver stiffness measurements values were obtained from the electronic medical records. Results: In total, 174 PBC patients with a median age of 54 (IQR, 48, 62) years old, were enrolled. 45 (25.9%) patients among them were diagnosed with sarcopenia. Univariate and multivariate logistic regression results illustrated that male gender (OR = 9.152, 95%CI = 3.131-26.751, p < 0.001) and LSM ≥ 12.8 kPa (OR = 4.539, 95%CI = 1.651, 12.478, p = 0.003) were the independent risk factors of sarcopenia in PBC patients. In the prognosis analysis, sarcopenia was determined as a risk factor for indicating adverse events in PBC patients (HR = 4.058, 95%CI = 1.955-8.424, p < 0.001) by Cox proportional hazards regression. Conclusion: The current findings illustrate that comprehensive evaluation and management of sarcopenia may contribute to the improvement of treatment outcomes and life quality of PBC patients.

Native and engineered extracellular vesicles: novel tools for treating liver disease.

Liver diseases are classified as acute liver damage and chronic liver disease, with recurring liver damage causing liver fibrosis and progression to cirrhosis and hepatoma. Liver transplantation is the only effective treatment for end-stage liver diseases; therefore, novel therapies are required. Extracellular vesicles (EVs) are endogenous nanocarriers involved in cell-to-cell communication that play important roles in immune regulation, tissue repair and regeneration. Native EVs can potentially be used for various liver diseases owing to their high biocompatibility, low immunogenicity and tissue permeability and engineered EVs with surface modification or cargo loading could further optimize therapeutic effects. In this review, we firstly introduced the mechanisms and effects of native EVs derived from different cells and tissues to treat liver diseases of different etiologies. Additionally, we summarized the possible methods to facilitate liver targeting and improve cargo-loading efficiency. In the treatment of liver disease, the detailed engineered methods and the latest delivery strategies were also discussed. Finally, we pointed out the limitations and challenges of EVs for future development and applications. We hope that this review could provide a useful reference for the development of EVs and promote the clinical translation.

Hypercholesterolemia Is Associated With Dysregulation of Lipid Metabolism and Poor Prognosis in Primary Biliary Cholangitis.

BACKGROUND & AIMS: Hypercholesterolemia is frequently diagnosed in patients with primary biliary cholangitis (PBC). However, its association with the prognosis and lipid metabolism is unknown. In this study, we aimed to investigate the prognostic value of baseline total cholesterol (TC) levels in PBC and characterized the associated lipid metabolism. METHODS: Five hundred and thirty-one patients with PBC without prior cirrhosis-related complications were randomly divided into the derivation and validation cohorts at a ratio of 7:3. Complete clinical data were obtained and analyzed. The endpoints were defined as liver-related death, liver transplantation, and cirrhosis-related complications. Lipidomics was performed in 89 patients and 28 healthy controls. RESULTS: Baseline TC was independently associated with poor liver-related outcomes, and adjusted C-statistics were 0.80 (95% confidence interval [CI]: 0.74-0.85) and 0.88 (95% CI: 0.78-0.91) in the derivation and validation cohorts, respectively. The predictive ability of TC for disease outcomes was stable over time and comparable with the Globe score. The 200 mg/dL cut-off optimally divided patients into low- and high-TC groups. A combination of TC and Globe score provided a more accurate stratification of patients into risk subgroups. Lipidomics indicated an up-regulation of lipid families in high-TC patients. Pathway analysis of 66 up-regulated lipids revealed the dysregulation of glycerophospholipid and sphingolipid metabolism in high-TC patients, which were associated with poor liver-related outcomes. CONCLUSIONS: Our results indicate that patients with PBC having baseline TC levels above 200 mg/dL have unique lipidome characteristics and are at a higher risk of poor liver-related outcomes.

Identifying optimal candidates for autologous peripheral blood stem cell therapy in patients with decompensated liver cirrhosis: a prognostic scoring system.

BACKGROUND: Stem cell transplantation shows great potential to improve the long-term survival of cirrhosis patients. However, therapeutic effects may not be homogeneous across the whole study population. This study constructed an easy-to-use nomogram to improve prognostic prediction and aid in treatment decision making for cirrhotic patients. METHODS: From August 2005 to April 2019, 315 patients with decompensated cirrhosis receiving autologous peripheral blood stem cell (PBSC) transplantation were enrolled in this study. They were randomly classified into training (2/3) and validation (1/3) groups. A predictive model was developed using Cox proportional hazard models and subsequently validated. The predictive performance of the model was evaluated and also compared with other prognostic models. RESULTS: Age, creatinine, neutrophil-to-lymphocyte ratio, and Child-Turcotte-Pugh class were included in the nomogram as prognostic variables. The nomogram showed high discrimination power concerning the area under receiver operating characteristic curves (3/5-year AUC: 0.742/0.698) and good consistency suggested by calibration plots. Patients could be accurately stratified into poor- and good-outcome groups regarding liver-transplantation free survival after receiving PBSC therapy (P < 0.001). Compared with poor-outcome group, the liver function of patients listed for liver transplantation in the good-outcome group was significantly improved (P < 0.001). Besides, our nomogram achieved a higher C-index (0.685, 95% CI 0.633-0.738) and better clinical utility compared with other conventional prognostic models. CONCLUSIONS: The proposed nomogram facilitated an accurate prognostic prediction for patients with decompensated cirrhosis receiving PBSC transplantation. Moreover, it also held the promise to stratify patients in clinical trials or practice to implement optimal treatment regimens for individuals.

Prognostic significance of liver stiffness in patients with primary biliary cholangitis: validation of Baveno VII criteria.

BACKGROUND: The role of liver stiffness measurements (LSM) in patients with primary biliary cholangitis (PBC) remains to be further elucidated. AIMS: To clarify the prognostic role of LSM and to validate the "novel concepts" proposed by the Baveno VII Working Group. METHODS: An analysis of the prognostic significance of LSM was performed involving 672 patients. RESULTS: LSM and ΔLSM/ΔT were independent risk factors for liver decompensation, liver transplantation, or liver-related death (primary outcomes, p < 0.001, both). A rule of 5 kPa for LSM (10-15-20 kPa) could be used to denote progressively higher relative risks of primary outcomes. Patients with LSM < 10 kPa have a negligible 3-year risk of primary outcomes (< 1%). Cut-off values of 10 and 15 kPa can be used to classify PBC patients into low-, medium-, and high-risk groups. A clinically significant decrease in LSM, evaluated at 6, 12, or 24 months elastography tests, was associated with a substantially reduced risk of primary outcomes (p < 0.05, all), which can be defined as a decrease in LSM of > - 20% associated with LSM < 20 kPa or any decrease to LSM < 10 kPa. A clinically significant increase in LSM, evaluated at 6, 12, or 24 months elastography tests, was associated with a substantially raised risk of primary outcomes (p < 0.05, all), which can be defined as an increase in LSM of ≥ + 20% or any increase to LSM ≥ 15 kPa. CONCLUSIONS: LSM can be used to monitor disease progression and predict long-term prognosis in patients with PBC.

Integrative bioinformatics analysis and experimental validation of key biomarkers for risk stratification in primary biliary cholangitis.

BACKGROUND: Primary biliary cholangitis (PBC) is an autoimmune liver disease, whose etiology is yet to be fully elucidated. Currently, ursodeoxycholic acid (UDCA) is the only first-line drug. However, 40% of PBC patients respond poorly to it and carry a potential risk of disease progression. So, in this study, we aimed to explore new biomarkers for risk stratification in PBC patients to enhance treatment. METHODS: We first downloaded the clinical characteristics and microarray datasets of PBC patients from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified and subjected to enrichment analysis. Hub genes were further validated in multiple public datasets and PBC mouse model. Furthermore, we also verified the expression of the hub genes and developed a predictive model in our clinical specimens. RESULTS: A total of 166 DEGs were identified in the GSE79850 dataset, including 95 upregulated and 71 downregulated genes. Enrichment analysis indicated that DEGs were significantly enriched in inflammatory or immune-related process. Among these DEGs, 15 risk-related genes were recognized and further validated in the GSE119600 cohort. Then, TXNIP, CD44, ENTPD1, and PDGFRB were identified as candidate hub genes. Finally, we proceeded to the next screening with these four genes in our serum samples and developed a three-gene panel. The gene panel could effectively identify those patients at risk of disease progression, yielding an AUC of 0.777 (95% CI, 0.657-0.870). CONCLUSIONS: In summary, combining bioinformatics analysis and experiment validation, we identified TXNIP, CD44, and ENTPD1 as promising biomarkers for risk stratification in PBC patients.

Integrated analysis and validation of the TRIM28-H2AX-CDK4 diagnostic model assists to predict the progression of HCC.

Hepatocellular carcinoma (HCC) is the second leading cause of cancer-related mortality in the world. However, identifying key genes that can be exploited for the effective diagnosis and management of HCC remains difficult. The study aims to examine the prognostic and diagnostic value of TRIM28-H2AX-CDK4 axis in HCC. Analysis in TCGA, GSEA and Gene expression profiling interactive analysis online tools were performed to explore the expression profiles of TRIM28, H2AX and CDK4. Data demonstrating the correlation between TRIM28 expression levels and immune infiltration states or the expression of genes associated with immune checkpoints genes were exacted from TCGA and TIMER. Genetic alteration and enrichment analysis were performed using the cBioPortal and GEPIA2 tools. Finally, the expression of these proteins in HCC was then examined and validated in an independent cohort using immunohistochemistry. TRIM28 alteration exhibited co-occurrence instead of mutual exclusivity with a large number of immune checkpoint components and tumor-infiltrating immune cells, especially B cells, were found to serve roles in patients with HCC with different TRIM28 expression levels. Higher expression levels of TRIM28, H2AX and CDK4 were associated with a poorer prognosis and recurrence in patients with HCC according to TCGA, which was validated further in an independent cohort of patients with HCC. Area under curve revealed the superior predictive power of applying this three-gene signatures in this validation cohort. The diagnostic model based on this TRIM28-H2AX-CDK4 signature is efficient and provides a novel strategy for the clinical management of HCC.

Cancer associated fibroblast derived SLIT2 drives gastric cancer cell metastasis by activating NEK9.

The secretory properties of cancer-associated fibroblasts (CAFs) play predominant roles in shaping a pro-metastatic tumor microenvironment. The present study demonstrated that SLIT2, an axon guidance protein, produced by CAFs and promoted gastric cancer (GC) metastasis in two gastric cancer cell lines (AGS and MKN45) by binding to roundabout guidance receptor 1 (ROBO1). Mass-spectrometry analysis revealed that ROBO1 could interact with NEK9, a serine/threonine kinase. And their mutual binding activities were further enhanced by SLIT2. Domain analysis revealed the kinase domain of NEK9 was critical in its interaction with the intracellular domain (ICD) of ROBO1, and it also directly phosphorylated tripartite motif containing 28 (TRIM28) and cortactin (CTTN) in AGS and MKN45 cells. TRIM28 function as a transcriptional elongation factor, which directly facilitate CTTN activation. In addition, Bioinformatics analysis and experimental validation identified transcriptional regulation of STAT3 and NF-κB p100 by TRIM28, and a synergetic transcription of CTTN by STAT3 and NF-κB p100 was also observed in AGS and MKN45. Therefore, CAF-derived SLIT2 increased the expression and phosphorylation levels of CTTN, which induced cytoskeletal reorganization and GC cells metastasis. A simultaneous increase in the expression levels of NEK9, TRIM28 and CTTN was found in metastatic GC lesions compared with paired non-cancerous tissues and primary cancer lesions via IHC and Multiplex IHC. The analysis of the data from a cohort of patients with GC revealed that increased levels of NEK9, TRIM28 and CTTN were associated with a decreased overall survival rate. On the whole, these findings revealed the connections of CAFs and cancer cells through SLIT2/ROBO1 and inflammatory signaling, and the key molecules involved in this process may serve as potential biomarkers and therapeutic targets for GC.

N6-methyladenosine RNA methylation in liver diseases: from mechanism to treatment.

Epigenetic modification occurring in RNA has become the hotspot of the field. N6-methyladenosine (m6A) methylation is the most abundant RNA internal modification mainly occurring at the consensus motif DR (m6A) CH (D = A/G/U, R = A/G, H = A/C/U) in the 3'-UTR particularly the region near stop codons. The life cycle of m6A methylation includes "writers," "erasers," and "readers", which are responsible for the addition, removal, and recognition of m6A, respectively. m6A modification has been reported changing RNA secondary structure or modulating the stability, localization, transport, and translation of mRNAs to play crucial roles in various physiological and pathological conditions. Liver, as the largest metabolic and digestive organ, modulates vital physiological functions, and its dysfunction gives rise to the occurrence of various diseases. Despite the advanced intervening measures, mortality due to liver diseases is continuously high. Recent studies have explored the roles of m6A RNA methylation in the pathogenesis of liver diseases, providing new insights for studying the molecular mechanism of liver diseases. In the review, we extensively summarize the life cycle of m6A methylation, as well as its function and relevant mechanisms in liver fibrosis (LF), nonalcoholic fatty liver disease (NAFLD), non-alcoholic steatohepatitis (NASH), hepatitis virus infection, and hepatocellular carcinoma (HCC), and eventually we explore the potential of m6A as a treatment option for these liver diseases.

Effectiveness of Fenofibrate in Treatment-Naive Patients With Primary Biliary Cholangitis: A Randomized Clinical Trial.

INTRODUCTION: Primary biliary cholangitis (PBC) is a progressive autoimmune liver disease, and patients with inadequate response to ursodeoxycholic acid (UDCA) treatment show reduced long-term survival. Recent studies have shown that fenofibrate is an effective off-label therapy for PBC. However, prospective studies on biochemical response including the timing of fenofibrate administration are lacking. This study is aimed to evaluate the efficacy and safety of fenofibrate in UDCA treatment-naive patients with PBC. METHODS: A total of 117 treatment-naive patients with PBC were recruited from the Xijing Hospital for a 12-month randomized, parallel, and open-label clinical trial. Study participants were assigned to receive either UDCA standard dose (UDCA-only group) or fenofibrate at a daily dose of 200 mg in addition to UDCA (UDCA-Fenofibrate group). RESULTS: The primary outcome was biochemical response percentage in patients according to the Barcelona criterion at 12 months. In the UDCA-Fenofibrate group, 81.4% (69.9%-92.9%) of patients achieved the primary outcome and 64.3% (51.9%-76.8%) in the UDCA-only group achieved the primary outcome ( P = 0.048). There was no difference between the 2 groups in noninvasive measures of liver fibrosis and biochemical markers other than alkaline phosphatase at 12 months. Creatinine and transaminases levels in the UDCA-Fenofibrate group increased within the first month, then returned to normal, and remained stable thereafter until the end of the study, even in patients with cirrhosis. DISCUSSION: In this randomized clinical trial in treatment-naive patients with PBC, the combination of fenofibrate and UDCA resulted in a significantly higher biochemical response rate. Fenofibrate seemed to be well-tolerated in patients.

Fenofibrate normalizes alkaline phosphatase and improves long-term outcomes in patients with advanced primary biliary cholangitis refractory to ursodeoxycholic acid.

BACKGROUND: Although patients with advanced liver disease have been included in studies evaluating fibrates for the treatment of primary biliary cholangitis (PBC), the frequency of biochemical responses and adverse effects for this group of patients was not reported separately and comprehensively. AIMS: to evaluate the efficacy and safety of additional fenofibrate therapy in patients with advanced and ursodeoxycholic acid (UDCA)-refractory PBC. METHODS: Patients were analyzed retrospectively to determine the clinical therapeutic effects of UDCA with additional fenofibrate therapy versus continued UDCA monotherapy. The liver transplantation (LT)-free survival and the alkaline phosphatase (ALP) normalization rates were estimated using Cox regression analyses and Kaplan-Meier plots with inverse probability of treatment weighting (IPTW). RESULTS: A total of 118 patients were included: 54 received UDCA alone and 64 received UDCA in combination with fenofibrate therapy. In the fenofibrate and UDCA groups, 37% and 11% of patients with advanced and UDCA-refractory PBC, respectively, achieved ALP normalization (P=0.001). Additional fenofibrate therapy improved both LT-free survival and ALP normalization rate after IPTW (hazard ratio [HR]: 0.23, 95% confidence interval [CI]: 0.07-0.75, P=0.015; and HR: 11.66, 95% CI: 5.02-27.06, P=0.001, respectively). These effects were supported by parallel changes in the rates of liver decompensation and histologic progression, and the United Kingdom (UK)-PBC and Globe risk scores. During the follow-up period, serum levels of ALP and aminotransferase decreased significantly, while total bilirubin, albumin, platelet, serum creatinine, and estimated glomerular filtration rate remained stable in fenofibrate-treated participants. No fenofibrate-related significant adverse events were observed in our cohort. CONCLUSIONS: Additional fenofibrate therapy significantly improved LT-free survival and ALP normalization in patients with advanced and UDCA-refractory PBC. Furthermore, adding-on fenofibrate therapy appeared to be safe and well tolerated in this population.

Prediction and evaluation of high-risk patients with primary biliary cholangitis receiving ursodeoxycholic acid therapy: an early criterion.

BACKGROUND AND AIMS: Current treatment guidelines recommend ursodeoxycholic acid (UDCA) as the first-line treatment for new-diagnosed primary biliary cholangitis (PBC) patients. However, up to 40% patients are insensitive to UDCA monotherapy, and evaluation of UDCA response at 12 months may result in long period of ineffective treatment. We aimed to develop a new criterion to reliably identify non-response patients much earlier. METHODS: Five hundred sixty-nine patients with an average of 59 months (Median: 53; IQR:32-79) follow-up periods were randomly divided into either the training (70%) or the validation cohort (30%). The efficiency of different combinations of total bilirubin (TBIL), alkaline phosphatase (ALP), and aspartate aminotransferase (AST) threshold values to predict outcomes was assessed at 1, 3 or 6 month after the initiation of UDCA therapy. The endpoints were defined as adverse outcomes, including liver-related death, liver transplantation and complications of cirrhosis. Adverse outcome-free survival was compared using various published criteria and a proposed new criterion. RESULTS: A new criterion of evaluating UDCA responses at 1 month was established as: ALP ≤ 2.5 × upper limit of normal (ULN) and AST ≤ 2 × ULN, and TBIL ≤ 1 × ULN (Xi'an criterion). The 5 year adverse outcome-free survival rate of UDCA responders, defined by Xi'an criterion, was 97%, which was significantly higher than that of those non-responders (64%). An accurate distinguishing high-risk patients' capacity of Xi'an criterion was confirmed in both early and late-stage PBC. CONCLUSIONS: Xi'an criterion has a similar or even higher ability to distinguish high-risk PBC patients than other published criteria. Xi'an criterion can facilitate early identification of patients requiring new therapeutic approaches.

A Novel Gene Signature Associated with Inflammatory Responses and Immune Status Assists in Prognosis and Intervention for Patients with HCC.

Background: Tumor growth depends on tumor cells and the tumor microenvironment, which are regulated by inflammation and immune responses. However, the roles of inflammation and immune status in hepatocellular carcinoma (HCC) remain unclear. The aim of this study was to evaluate the prognostic value of an inflammatory response- related gene signature associated with immune status, which may provide insight into new treatment options for HCC patients. Materials and Methods: Differentially expressed genes associated with inflammation were obtained from The Cancer Genome Atlas (TCGA), the Gene Expression Omnibus, and the Molecular Signatures Database. An inflammation-associated prognostic gene signature was constructed and validated using TCGA and the International Cancer Genome Consortium datasets, respectively, using LASSO Cox regression analysis. Log-rank was performed to compare the overall survival of low- and high-risk score cohorts. Immune cell infiltration and immune-related functions were analyzed using single-sample gene enrichment analysis. The structures of the drugs identified by the prognostic model were predicted using PubChem. The drugs sensitivity of bleomycin, simvastatin and zoledronate detected by CCK8 colorimetric assay. The mRNA levels of 7 genes in HCC after drug treatment analyzed via qRT-PCR. Results: Inflammation-associated genes, including ITGA5, MEP1A, P2RX4, RIPK2, SLC7A1 and SRI, were identified and found to be associated with the prognosis of HCC. We further found that the high-risk patients experienced poor prognosis, which was observed to be an independent and significant risk factor for prognosis. Moreover, we observed elevated expression levels in multiple immune cell types and immune function. Lastly, we validated that bleomycin, simvastatin and zoledronate could regulate these genes in HCC. Conclusion: The inflammatory-response-associated gene signature could predict the prognosis and the immunological status of HCC patients. Additionally, bleomycin, simvastatin and zoledronate may represent potential drug candidates that could inhibit these genes. This may constitute a new approach for the treatment of HCC.

Efficacy and safety of fenofibrate addition therapy in patients with cirrhotic primary biliary cholangitis with incomplete response to ursodeoxycholic acid.

Fenofibrate (FF) has shown potential benefits in patients with primary biliary cholangitis (PBC) who have an incomplete response to ursodeoxycholic acid (UDCA). However, the efficacy and safety of FF in patients with cirrhosis remain unclear. To evaluate the efficacy and safety of additional FF therapy in patients with PBC-related cirrhosis with an incomplete response to UDCA, we conducted a retrospective analysis comparing the clinical results of additional FF therapy and continued UDCA monotherapy. A total of 59 patients were included; 27 cases underwent UDCA monotherapy and 32 cases underwent UDCA combined with FF therapy. A significant difference in alkaline phosphatase (ALP) normalization was achieved in the FF group compared to the UDCA group (37% vs. 11%, respectively; p = 0.020). Additional FF therapy was an independent risk factor for ALP normalization (hazard ratio, 7.679; 95% confidence interval, 2.059-28.633; p = 0.003). Hepatic deterioration was experienced by 40% versus 48% (p = 0.562) while 11% vs. 37% (p = 0.111) experienced liver-related mortality or liver transplantation in the FF and UDCA groups, respectively. Compared to UDCA monotherapy, additional FF therapy was associated with lower United Kingdom (UK)-PBC risk score and surrogate serum indices of liver fibrosis. After 12 months of add-on FF therapy, median ALP level and UK-PBC risk score decreased 35% and 52% from baseline (p = 0.001 and 0.210, respectively). Serum aminotransferase, triglyceride, and cholesterol decreased progressively, while total bilirubin, serum creatinine, blood urea, estimated glomerular filtration rate, aspartate aminotransferase-to-platelet ratio index, and fibrosis-4 index remained stable in FF-treated cirrhotic cases during follow-up. No significant adverse effects associated with additional FF therapy were observed in our cohort. Conclusion: Additional FF therapy was associated with higher ALP normalization rates and lower UK-PBC risk scores in patients with cirrhotic PBC with an incomplete response to UDCA. In addition, FF therapy seemed safe and well tolerated with a low frequency of adverse effects in patients with cirrhosis.

Mesenchymal stem cell-derived exosomes protect against liver fibrosis via delivering miR-148a to target KLF6/STAT3 pathway in macrophages.

BACKGROUND: Despite emerging evidence on the therapeutic potential of mesenchymal stem cells (MSCs) for liver fibrosis, the underlying mechanisms remain unclear. At present, MSC-derived exosomes (MSC-EXOs) are widely accepted as crucial messengers for intercellular communication. This study aimed to explore the therapeutic effects of MSC-EXOs on liver fibrosis and identify the mechanisms underlying the action of MSC-EXOs. METHODS: Carbon tetrachloride was used to induce a liver fibrosis model, which was intravenously administered with MSCs or MSC-EXOs to assess treatment efficacy. The resulting histopathology, fibrosis degree, inflammation and macrophage polarization were analyzed. RAW264.7 and BMDM cells were used to explore the regulatory effects of MSC-EXOs on macrophage polarization. Then, the critical miRNA mediating the therapeutic effects of MSC-EXOs was screened via RNA sequencing and validated experimentally. Furthermore, the target mRNA and downstream signaling pathways were elucidated by luciferase reporter assay, bioinformatics analysis and western blot. RESULTS: MSCs alleviated liver fibrosis largely depended on their secreted exosomes, which were visualized to circulate into liver after transplantation. In addition, MSC-EXOs were found to modulate macrophage phenotype to regulate inflammatory microenvironment in liver and repair the injury. Mechanically, RNA-sequencing illustrates that miR-148a, enriched in the MSC-EXOs, targets Kruppel-like factor 6 (KLF6) to suppress pro-inflammatory macrophages and promote anti-inflammatory macrophages by inhibiting the STAT3 pathway. Administration of miR-148a-enriched MSC-EXOs or miR-148a agomir shows potent ameliorative effects on liver fibrosis. CONCLUSIONS: These findings suggest that MSC-EXOs protect against liver fibrosis via delivering miR-148a that regulates intrahepatic macrophage functions through KLF6/STAT3 signaling and provide a potential therapeutic target for liver fibrosis.

Rho GTPase-activating protein 35 suppresses gastric cancer metastasis by regulating cytoskeleton reorganization and epithelial-to-mesenchymal transition.

Cytoskeletal reorganization and epithelial-to-mesenchymal transition (EMT) are key processes and typical characteristics of metastatic cancer cells. Rho GTPase­activating protein 35 (ARHGAP35) is a GTPase-activating protein, which has a significant effect on cell motility. However, the particular function of ARHGAP35 in gastric cancer (GC) remains unknown. In the present study, the role of ARHGAP35 in GC was investigated by in vitro loss-of-function and gain-of-function experiments. Cytoskeletal reorganization in GC cells was evaluated using immunofluorescence staining and the protein expression levels of key molecules and active RhoA were detected by western blot analysis. Additionally, the clinical evaluation of proteins in human GC tissues was assessed by immunohistochemistry. The results showed that ARHGAP35, a tumor suppressor, was downregulated in GC tissues and its decreased expression was associated with the metastatic status of GC. Additionally, Transwell and wound healing assays demonstrated that ARHGAP35 knockdown promoted cell motility in vitro. However, the above effects were abrogated following ectopic ARHGAP35 expression. Furthermore, ARHGAP35 could affect cytoskeletal reorganization via directly regulating RhoA activation. In addition, ARHGAP35 upregulated E-cadherin and attenuated EMT in GC cells. Both ARHGAP35 and E-cadherin were associated with overall survival in patients with GC, while their combination allowed for an even greater capacity for distinguishing GC patients with different prognosis. Overall, the results of the current study suggested that ARHGAP35 could directly regulate cell morphology and motility via affecting cytoskeletal reorganization and EMT via targeting RhoA and E-cadherin, respectively. Targeting the ARHGAP35/RhoA/E-cadherin pathway could be a potential approach for treating GC.

Efficacy and safety of apatinib as second or later-line therapy in extensive-stage small cell lung cancer: a prospective, exploratory, single-arm, multi-center clinical trial.

Background: A paucity of strategies exist for extensive-stage small cell lung cancer (ES-SCLC) patients who fail the first-line chemotherapy. Apatinib is a tyrosine kinase inhibitor (TKI) that selectively inhibits vascular endothelial growth factor receptor-2 (VEGFR-2), which has been demonstrated to have active anti-tumor activity in ES-SCLC when used only or combined with PD-1 inhibitors or chemotherapy with good tolerance. However, the efficacy and safety of apatinib monotherapy is unclear in second-line or beyond treatment of ES-SCLC. Methods: In this prospective, exploratory, single-arm, multi-center study, eligible patients were aged 18 years or older with histologically confirmed ES-SCLC, and had progressed on, or were intolerant to previous systemic treatment. Patients received apatinib 500 mg (orally qd, every 4 weeks a cycle). The efficacy was assessed after 1 cycle and then every 2 cycles based on computed tomography imaging per the Response Evaluation Criteria in Solid Tumors (RECIST, version 1.1). The primary endpoint was progression-free survival (PFS). The adverse events (AEs) were assessed per the National Cancer Institute Common Terminology Criteria for Adverse Events 4.0 (NCI-CTCAE 4.0). This study is registered in the Chinese Clinical Trial Registry, number ChiCTR-OPC-17013964. Results: From 28 July 2017 to 21 June 2019, 62 patients were screened for eligibility, among whom 57 patients were available for efficacy and safety analysis. The objective response rate (ORR) was 14.3% and disease control rate (DCR) was 79.6%. The median PFS was 5.6 months [95% confidence interval (CI): 3.3-8.0 months] and the median overall survival (OS) was 11.2 months (95% CI: 7.5-24.0 months). Among the participants who received apatinib as second-line treatment, the median PFS and OS were 6.1 months (95% CI: 2.6-7.6 months) and 12.0 months (95% CI: 7.9 months to not reached), respectively. The most common AEs of all grades were anemia (36.8%), hypertension (33.3%), fatigue (31.6%), blood bilirubin increased (22.8%), elevated transaminase (19.3%), and hand-foot syndrome (17.54%). Grade 3 AEs included 2 (3.5%) cases of hypertension and 1 (1.8%) case of fatigue. No grade 4/5 AEs were observed. Conclusions: Apatinib showed encouraging anti-tumor activity in pretreated ES-SCLC patients with tolerable toxicities. Further larger scale studies are warranted to demonstrate the efficacy of apatinib.

Comprehensive evaluation of the effects of long-term cryopreservation on peripheral blood mononuclear cells using flow cytometry.

Human peripheral blood mononuclear cells (PBMCs) originate from hematopoietic stem cells in the bone marrow, which mainly includes lymphocytes (T cells, B cells, and natural killer cells) and monocytes. Cryopreserved PBMCs providing biobank resources are crucial for clinical application or scientific research. Here, we used flow cytometry to explore the influence of long-term cryopreservation on the quality of PBMCs with the aim of providing important evidence for the effective utilization of biobank resources. The PBMCs were isolated from the peripheral blood, which was collected from volunteers in the hospital. After long-term cryopreservation in liquid nitrogen, we analyzed the changes in cell numbers, viability, and multiple subtypes of PBMCs and studied the apoptosis, proliferation, activation, function, and status of T cells in comparison with freshly isolated PBMCs by flow cytometry, and then further tracked the effects of long-term cryopreservation on the same sample. Although the different cell types in the PBMCs dynamically changed compared with those in the freshly isolated samples, PBMC recovery and viability remained stable after long-term cryopreservation, and the number of most innate immune cells (e.g., monocytes and B cells) was significantly reduced compared to that of the freshly isolated PBMCs or long-term cryopreserved PBMCs; more importantly, the proportion of T cell subtypes, apoptosis, proliferation, and functional T cells, except for Tregs, were not affected by long-term cryopreservation. However, the proportions of activated T, naïve T, central memory T, effector T, and effector memory T cells dynamically changed after long-term cryopreservation. This article provides important evidence for the effective utilization of biobank resources. Long-term cryopreserved PBMCs can be partly used as biological resources for clinical research or basic studies, but the effect of cryopreservation on PBMCs should be considered when selecting cell samples, especially in research relating to activating or inhibiting function.

A novel immune checkpoint-related gene signature for hepatocellular carcinoma to predict clinical outcomes and therapeutic response.

Immune checkpoint genes (ICGs) have recently been proven to perform instrumental functions in the maintenance of immune homeostasis and represent a promising therapeutic strategy; however, their expression patterns and prognostic values are not fully elucidated in hepatocellular carcinoma (HCC). In this investigation, we focused on establishing and validating a prognostic gene signature to facilitate decision-making in clinical practice. Clinical information, as well as transcriptome data, was obtained from the Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) database. Univariate Cox regression and least absolute shrinkage and selection operator (LASSO) Cox method were employed to build a multi-gene signature in the TCGA database, while the ICGC database was used for validation. Subsequently, utilizing the six-gene signature, we were able to categorize patients into high- and low-risk groups. In two cohorts, survival analysis findings revealed a dismal outlook for the high-risk group. The receiver operating characteristic curves were utilized to estimate the gene signature's prediction ability. Moreover, correlation analysis showed high-risk group was linked to advanced pathological stage, infiltration of immune cells and therapeutic response. In summary, this unique gene profile might serve not only as a useful prognostic indicator but also as a marker of therapy responsiveness in HCC.