Deactivation of the Unfolded Protein Response Aggravated Renal AA Amyloidosis in HSF1 Deficiency Mice.

Systemic amyloid A (AA) amyloidosis, which is considered the second most common form of systemic amyloidosis usually takes place several years prior to the occurrence of chronic inflammation, generally involving the kidney. Activated HSF1, which alleviated unfolded protein response (UPR) or enhanced HSR, is the potential therapeutic target of many diseases. However, the effect of HSF1 on AA amyloidosis remains unclear. This study focused on evaluating effect of HSF1 on AA amyloidosis based on HSF1 knockout mice. As a result, aggravated amyloid deposits and renal dysfunction have been found in HSF1 knockout mice. In progressive AA amyloidosis, HSF1 deficiency enhances serum amyloid A production might to lead to severe AA amyloid deposition in mice, which may be related to deactivated unfolded protein response as well as enhanced inflammation. Thus, HSF1 plays a significant role on UPR related pathway impacting AA amyloid deposition, which can mitigate amyloidogenic proteins from aggregation pathologically and is the possible way for intervening with the pathology of systemic amyloid disorder. In conclusion, HSF1 could not only serve as a new target for AA amyloidosis treatment in the future, but HSF1 knockout mice also can be considered as a valuable novel animal model for renal AA amyloidosis.

miR-4645-3p attenuates podocyte injury and mitochondrial dysfunction in diabetic kidney disease by targeting Cdk5.

Podocyte injury plays a critical role in the progression of diabetic kidney disease (DKD), but the underlying cellular and molecular mechanisms remain poorly understanding. MicroRNAs (miRNAs) can disrupt gene expression by inducing translation inhibition and mRNA degradation, and recent evidence has shown that miRNAs may play a key role in many kidney diseases. In this study, we identified miR-4645-3p by global transcriptome expression profiling as one of the major downregulated miRNAs in high glucose-cultured podocytes. Moreover, whether DKD patients or STZ-induced diabetic mice, expression of miR-4645-3p was also significantly decreased in kidney. In the podocytes cultured by normal glucose, inhibition of miR-4645-3p expression promoted mitochondrial damage and podocyte apoptosis. In the podocytes cultured by high glucose (30 mM glucose), overexpression of miR-4645-3p significantly attenuated mitochondrial dysfunction and podocyte apoptosis induced by high glucose. Furthermore, we found that miR-4645-3p exerted protective roles by targeting Cdk5 inhibition. In vitro, miR-4645-3p obviously antagonized podocyte injury by inhibiting overexpression of Cdk5. In vivo of diabetic mice, podocyte injury, proteinuria, and impaired renal function were all effectively ameliorated by treatment with exogenous miR-4645-3p. Collectively, these findings demonstrate that miR-4645-3p can attenuate podocyte injury and mitochondrial dysfunction in DKD by targeting Cdk5. Sustaining the expression of miR-4645-3p in podocytes may be a novel strategy to treat DKD.

Activation of IGFBP4 via unconventional mechanism of miRNA attenuates metastasis of intrahepatic cholangiocarcinoma.

BACKGROUND: Intrahepatic cholangiocarcinoma (ICC) is the second most common primary liver malignancy. Although its incidence is lower than that of hepatocellular carcinoma (HCC), ICC has a worse prognosis, and it is more prone to recur and metastasize, resulting in a far greater level of malignancy. METHODS: Bioinformatics analysis and qRT-PCR were applied to assess the level of miR-122-5p and IGFBP4. Western blot, transwell assays, wound-healing assays, real-time cellular invasion monitoring, in vivo study were applied to explore the function of miR-122-5p and IGFBP4. Dual luciferase reporter assays and chromatin isolation by RNA purification (ChiRP) were applied to explore the regulation of IGFBP4 by miR-122-5p. RESULTS: Using The Cancer Genome Atlas (TCGA) data set, Sir Run Run Shaw hospital data set and bioinformatics analyses, we identified miR-122-5p as a potential tumor suppressor in ICC and validated its suppressive effect in metastasis and invasion of ICC. Transcriptome sequencing, rescue and complement experiments were used to identify insulin-like growth factor binding protein 4 (IGFBP4) as a target of miR-122-5p. The mechanism by which miR-122-5p regulates IGFBP4 was clarified by chromatin separation RNA purification technology, and dual-luciferase reporter assays. We discovered a rare novel mechanism by which miR-122-5p promotes IGFBP4 mRNA transcription by binding to its promoter region. Furthermore, in mouse orthotopic metastasis model, miR-122-5p inhibited the invasion of ICC. CONCLUSION: In summary, our study revealed a novel mechanism of miR-122-5p and function of the miR-122-5p/IGFBP4 axis in the metastasis of ICC. We also highlighted the clinical value of miR-122-5p and IGFBP4 in inhibiting ICC invasion and metastasis.

Development of an Aerobic Glycolysis Index for Predicting the Sorafenib Sensitivity and Prognosis of Hepatocellular Carcinoma.

Hepatocellular carcinoma (HCC) is a deadly tumor with high heterogeneity. Aerobic glycolysis is a common indicator of tumor growth and plays a key role in tumorigenesis. Heterogeneity in distinct metabolic pathways can be used to stratify HCC into clinically relevant subgroups, but these have not yet been well-established. In this study, we constructed a model called aerobic glycolysis index (AGI) as a marker of aerobic glycolysis using genomic data of hepatocellular carcinoma from The Cancer Genome Atlas (TCGA) project. Our results showed that this parameter inferred enhanced aerobic glycolysis activity in tumor tissues. Furthermore, high AGI is associated with poor tumor differentiation and advanced stages and could predict poor prognosis including reduced overall survival and disease-free survival. More importantly, the AGI could accurately predict tumor sensitivity to Sorafenib therapy. Therefore, the AGI may be a promising biomarker that can accurately stratify patients and improve their treatment efficacy.

Efficacy of Laparoscopic Hepatectomy versus Open Surgery for Hepatocellular Carcinoma With Cirrhosis: A Meta-analysis of Case-Matched Studies.

BACKGROUND: The role of laparoscopic hepatectomy (LH) in hepatocellular carcinoma (HCC) with cirrhosis remains controversial and needs to be further assessed. The present meta-analysis aimed to compare the surgical and oncological outcomes of LH with those of open hepatectomy (OH) for HCC with cirrhosis. METHODS: The PubMed, Embase, and Cochrane Library databases were searched for studies comparing LH and OH until Mar 2021. Weighted mean differences (WMDs), odds ratios (ORs), and hazard ratios (HRs) were calculated for continuous, dichotomous, and long-term variables, respectively, with 95% confidence intervals (CIs). Subgroup analysis was performed according to different resection types: major resection and minor resection. The meta-analysis was performed using the STATA 12.0. RESULTS: A total of 16 case-matched studies (784 patients in the LH group and 1,191 patients in the OH group.) were included in this meta-analysis. In terms of primary outcomes, LH was associated with decreased overall complication rate (OR 0.57; 95% CI 0.46 to 0.71; P <0.01), major complication rate (OR 0.52; 95% CI 0.33 to 0.82; P < 0.01), postoperative mortality (OR 0.27; 95% CI 0.11 to 0.66; P <0.01), 1-y overall survival (OS) rate (HR 0.48; 95% CI 0.31 to 0.73; P <0.01), 2-y OS (HR 0.61; 95% CI 0.45 to 0.83; P < 0.01), and 5-y OS (0.67; 95% CI 0.53 to 0.85; P < 0.01). With respect to secondary outcomes, blood loss (WMD -69.16; 95% CI -101.72 to -36.61; P < 0.01), length of hospitalization (LOH) (WMD -2.65; 95% CI -3.41 to -1.89; P < 0.01), minor complication rate (OR 0.70; 95% CI 0.53 to 0.94; P = 0.02), postoperative liver failure (OR 0.60; 95% CI 0.38 to 0.95; P = 0.03), and postoperative ascites (OR 0.44; 95% CI 0.28 to 0.72; P < 0.01) was lower in LH than in OH. No significant differences in operation time (P = 0.07), transfusion rate (P = 0.05), 1-, 2-, and 5-year DFS rate (1-year, P = 0.08; 2-year, P = 0.08; 5-year, P = 0.23) were noted between LH and OH. Subgroup analysis based on minor resection revealed that LH had similar favored outcomes in comparison with those in the overall pooled analysis. However, LH had a longer operation time than OH in the setting of major resection (P < 0.01). CONCLUSION: LH is technically feasible and safe for selected HCC patients with cirrhosis. LH can achieve favored short-term and long-term oncological outcomes in minor liver resection. Laparoscopic major hepatectomy (LMH) seems to offer some advantages over the open approach; however concerns about surgical and oncological safety remain. More evidence on LMH is warranted before expanding its indication to patients with cirrhosis.

TAK1 Is a Novel Target in Hepatocellular Carcinoma and Contributes to Sorafenib Resistance.

BACKGROUND & AIMS: Identifying novel and actionable targets in hepatocellular carcinoma (HCC) remains an unmet medical need. TAK1 was originally identified as a transforming growth factor-ß-activated kinase and was further proved to phosphorylate and activate numerous downstream targets and promote cancer progression. However, the role of TAK1 in developed HCC progression and targeted therapy resistance is poorly understood. METHODS: The expression of TAK1 or MTDH in HCC cell lines, tumor tissues, and sorafenib-resistant models was analyzed by in silico analysis, quantitative real-time polymerase chain reaction, Western blotting, and immunohistochemistry. In vivo and in vitro experiments were introduced to examine the function of TAK1 or MTDH in HCC and sorafenib resistance using small interfering RNA and pharmacologic inhibitors in combination with or without sorafenib. Co-immunoprecipitation and RNA immunoprecipitation were carried out to determine the binding between TAK1 and FBXW2 or between MTDH and FBXW2 mRNA. Protein half-life and in vitro ubiquitination experiment was performed to validate whether FBXW2 regulates TAK1 degradation. RESULTS: Our findings unraveled the clinical significance of TAK1 in promoting HCC and sorafenib resistance. We identified a novel E3 ubiquitin ligase, FBXW2, targeting TAK1 for K48-linked polyubiquitylation and subsequent degradation. We also found that MTDH contributes to TAK1 up-regulation in HCC and sorafenib resistance through binding to FBXW2 mRNA and accelerates its degradation. Moreover, combination of TAK1 inhibitor and sorafenib suppressed the growth of sorafenib-resistant HCCLM3 xenograft in mouse models. CONCLUSIONS: These results revealed novel mechanism underlying TAK1 protein degradation and highlighted the therapeutic value of targeting TAK1 in suppressing HCC and overcoming sorafenib resistance.

UBQLN1 mediates sorafenib resistance through regulating mitochondrial biogenesis and ROS homeostasis by targeting PGC1ß in hepatocellular carcinoma.

The treatment for hepatocellular carcinoma (HCC) is promising in recent years, but still facing critical challenges. The first targeted therapy, sorafenib, prolonged the overall survival by months. However, resistance often occurs, largely limits its efficacy. Sorafenib was found to target the electron transport chain complexes, which results in the generation of reactive oxygen species (ROS). To maintain sorafenib resistance and further facilitate tumor progression, cancer cells develop strategies to overcome excessive ROS production and obtain resistance to oxidative stress-induced cell death. In the present study, we investigated the roles of ROS in sorafenib resistance, and found suppressed ROS levels and reductive redox states in sorafenib-resistant HCC cells. Mitochondria in sorafenib-resistant cells maintained greater functional and morphological integrity under the treatment of sorafenib. However, cellular oxygen consumption rate and mitochondria DNA content analyses revealed fewer numbers of mitochondria in sorafenib-resistant cells. Further investigation attributed this finding to decreased mitochondrial biogenesis, likely caused by the accelerated degradation of peroxisome proliferator-activated receptor γ coactivator 1ß (PGC1ß). Mechanistic dissection showed that upregulated UBQLN1 induced PGC1ß degradation in a ubiquitination-independent manner to attenuate mitochondrial biogenesis and ROS production in sorafenib-resistant cells under sorafenib treatment. Furthermore, clinical investigations further indicated that the patients with higher UBQLN1 levels experienced worse recurrence-free survival. In conclusion, we propose a novel mechanism involving mitochondrial biogenesis and ROS homeostasis in sorafenib resistance, which may offer new therapeutic targets and strategies for HCC patients.

Cdk5-Mediated Phosphorylation of Sirt1 Contributes to Podocyte Mitochondrial Dysfunction in Diabetic Nephropathy.

Aims: Mitochondrial dysfunction contributes to podocyte injury, which is the leading cause of proteinuria in diabetic nephropathy (DN). In this study, we explored the role of cyclin-dependent kinase 5 (Cdk5) in mitochondrial dysfunction of podocytes under diabetic conditions. Results: Our results showed that the expression and activity of Cdk5 were significantly upregulated in vivo and in vitro under diabetic conditions, accompanied by the downregulation of synaptopodin and nephrin, as well as structural and functional mitochondrial dysfunction. Inhibition of Cdk5 with roscovitine or dominant-negative Cdk5 led to the attenuation of podocyte injury by upregulating synaptopodin and nephrin. The inhibition of Cdk5 also ameliorated mitochondrial dysfunction by decreasing reactive oxygen species levels and cytochrome c release, while increasing adenosine triphosphate production. Sirt1, an NAD+-dependent deacetylase, was decreased in podocytes with high glucose (HG) treatment; however, its phosphorylation level at S47 was significantly upregulated. We demonstrated that HG levels cause overactive Cdk5 to phosphorylate Sirt1 at S47. Suppression of Cdk5 reduced Sirt1 phosphorylation levels and mutation of S47 to nonphosphorable alanine (S47A), significantly attenuated podocyte injury and mitochondrial dysfunction in diabetic condition in vivo and in vitro. Innovation and Conclusion: Our study has demonstrated the role of Cdk5 in regulating mitochondrial function through Sirt1 phosphorylation and thus can potentially be a new therapeutic target for DN treatment. IRB number: 20190040. Antioxid. Redox Signal. 34, 171-190.

N6-methyladenosine-modified CircRNA-SORE sustains sorafenib resistance in hepatocellular carcinoma by regulating ß-catenin signaling.

BACKGROUND AND AIMS: Accumulating evidence suggests that the primary and acquired resistance of hepatocellular carcinoma (HCC) to sorafenib is mediated by multiple molecular, cellular, and microenvironmental mechanisms. Understanding these mechanisms will enhance the likelihood of effective sorafenib therapy. METHODS: In vitro and in vivo experiments were performed and clinical samples and online databases were acquired for clinical investigation. RESULTS: In this study, we found that a circular RNA, circRNA-SORE, which is up-regulated in sorafenib-resistant HCC cells, was necessary for the maintenance of sorafenib resistance, and that silencing circRNA-SORE substantially increased the efficacy of sorafenib-induced apoptosis. Mechanistic studies determined that circRNA-SORE sequestered miR-103a-2-5p and miR-660-3p by acting as a microRNA sponge, thereby competitively activating the Wnt/ß-catenin pathway and inducing sorafenib resistance. The increased level of circRNA-SORE in sorafenib-resistant cells resulted from increased RNA stability. This was caused by an increased level of N6-methyladenosine (m6A) at a specific adenosine in circRNA-SORE. In vivo delivery of circRNA-SORE interfering RNA by local short hairpin RNA lentivirus injection substantially enhanced sorafenib efficacy in animal models. CONCLUSIONS: This work indicates a novel mechanism for maintaining sorafenib resistance and is a proof-of-concept study for targeting circRNA-SORE in sorafenib-treated HCC patients as a novel pharmaceutical intervention for advanced HCC.

LXR activation potentiates sorafenib sensitivity in HCC by activating microRNA-378a transcription.

Sorafenib resistance is a major obstacle to the treatment of advanced hepatocellular carcinoma (HCC). MicroRNAs (miRNAs) are multifunctional regulators of gene expression with profound impact for human disease. Therefore, better understanding of the biological mechanisms of abnormally expressed miRNAs is critical to discovering novel, promising therapeutic targets for HCC treatment. This study aimed to investigate the role of miR-378a-3p in the sorafenib resistance of HCC and elucidate the underlying molecular mechanisms. Methods: A novel hub miR-378a-3p was identified based on miRNA microarray and bioinformatics analysis. The abnormal expression of miR-378-3p was validated in different HCC patient cohorts and sorafenib-resistant (SR) HCC cell lines. The functional role of miR-378a-3p and its downstream and upstream regulatory machinery were investigated by gain-of-function and loss-of-function assays in vitro and in vivo. Interactions among miR-378a-3p, LXRα, and IGF1R were examined by a series of molecular biology experiments. Then, the clinical relevance of miR-378a-3p and its targets were evaluated in HCC samples. HCC patient-derived xenograft (PDX) model was used to assess the therapeutic value of LXRα and its downstream miR-378a-3p. Results: miR-378a-3p expression was frequently reduced in established sorafenib-resistant HCC cell lines. The decreased miR-378a-3p levels correlated with poor overall survival of HCC patients following sorafenib treatment. miR-378a-3p overexpression induced apoptosis in SR HCC cells, whereas miR-378a-3p silencing exerted the opposite effects. IGF1R was identified as a novel target of miR-378a-3p. Furthermore, the primary miR-378 level was not consistent with its precursor miRNA level in SR HCC cells, which was attributed to the downregulation of exportin5 (XPO5) and subsequently reduced nuclear export of precursor miR-378 and restrained maturation of miR-378-3p. In this context, we combined an agonist GW3965 of liver X receptor alpha (LXRα), which functioned as a transcription activator of miRNA-378a, and its activation re-sensitized sorafenib-resistant cells to sorafenib treatment in vitro and in vivo. Conclusions: Our finding suggested decreased expression of XPO5 prevents maturation of miR-378a-3p, which leaded to the overexpression of IGF-1R and counteracted the effects of sorafenib-induced apoptosis. LXRα was able to activate miRNA-378a-3p transcription in HCC cells and could be a potential combinable treatment strategy with sorafenib to suppress HCC progression.

miR-486-3p mediates hepatocellular carcinoma sorafenib resistance by targeting FGFR4 and EGFR.

HCC is a common malignancy worldwide and surgery or reginal treatments are deemed insufficient for advanced-stage disease. Sorafenib is an inhibitor of many kinases and was shown to benefit advanced HCC patients. However, resistance emerges soon after initial treatment, limiting the clinical benefit of sorafenib, and the mechanisms still remain elusive. Thus, this study aims to investigate the mechanisms of sorafenib resistance and to provide possible targets for combination therapies. Through miRNA sequencing, we found that miR-486-3p was downregulated in sorafenib resistant HCC cell lines. Cell viability experiments showed increased miR-486-3p expression could induce cell apoptosis while miR-486-3p knockdown by CRISPR-CAS9 technique could reduce cell apoptosis in sorafenib treatment. Clinical data also indicated that miR-486-3p level was downregulated in tumor tissue compared with adjacent normal tissue in HCC patients. Mechanism dissections showed that FGFR4 and EGFR were the targets of miR-486-3p, which was verified by luciferase reporter assay. Importantly, FGFR4 or EGFR selective inhibitor could enhance sorafenib efficacy in the resistant cells. Moreover, in vivo sorafenib resistant model identified that over-expressing miR-486-3p by lentivirus injection could overcome sorafenib resistance by significantly suppressing tumor growth in combination with the treatment of sorafenib. In conclusion, we found miR-486-3p was an important mediator regulating sorafenib resistance by targeting FGFR4 and EGFR, thus offering a potential target for HCC treatment.

The microenvironmental and metabolic aspects of sorafenib resistance in hepatocellular carcinoma.

In most cases, sorafenib-resistant HCC cells exhibit significant mesenchymal phenotype and stemness features. In this context, tumor cells might undergo cell fate transition in response to sorafenib or other targeted drugs in the presence or absence of genetic mutations. Therefore, understanding the major characteristics of drug-resistant cells state helps to discover new treatments that overcome drug resistance. To note, little is known about the metabolic or microenvironmental aspects of the certain tumor cell states beyond the genome. This review mainly focuses on the underlying mechanisms of acquired sorafenib resistance based on CSCs and EMT models, which explain tumor heterogeneity and have been considered the major cause of secondary sorafenib resistance. In particular, it discusses how the tumor microenvironment and tumor metabolism regulate cell stemness, mesenchymal state, and sorafenib resistance through epigenetic regulations, and provides reliable targets that might have synergetic effect with sorafenib.

Laparoscopic Versus Open Resection of Small Bowel Gastrointestinal Stromal Tumors: Systematic Review and Meta-Analysis.

BACKGROUND: Laparoscopic resection (LAP) for small bowel gastrointestinal stromal tumors (GISTs) is not as common as for stomach. This study aimed to evaluate the safety and efficacy of LAP for small bowel GISTs with systematic review and meta-analysis. METHODS: The Web of Science, Cochrane Library, Embase, and PubMed databases before December 2016 were comprehensively searched to retrieve comparative trials of LAP and conventional open resection (OPEN) for GISTs of small bowel with a relevance of review object. These researches reported intraoperative and postoperative clinical course (operation time, blood loss, time to first flatus and oral intake, hospital stay, morbidity, and mortality), oncologic outcomes, and long-term survival status. RESULTS: Six studies involving 391 patients were identified. Compared to OPEN, LAP had associated with a shorter operation time (weighted mean difference [WMD] = -27.97 min, 95% confidence interval [CI]: -49.40--6.54, P < 0.01); less intraoperative blood loss (WMD = -0.72 ml; 95% CI: -1.30--0.13, P = 0.02); earlier time to flatus (WMD = -0.83 day; 95% CI: -1.44--0.22, P < 0.01); earlier time to restart oral intake (WMD = -1.95 days; 95% CI: -3.31--0.60, P < 0.01); shorter hospital stay (WMD = -3.00 days; 95% CI: -4.87--1.13, P < 0.01); and a decrease in overall complications (risk ratio = 0.56, 95% CI: 0.33-0.97, P = 0.04). In addition, the tumor recurrence and long-term survival rate showed that there was no significant difference between the two groups of patients. CONCLUSIONS: LAP for small bowel GISTs is a safe and feasible procedure with shorter operation time, less blood loss, less overall complications, and quicker recovery. Besides, tumor recurrence and the long-term survival rate are similar to open approach. Because of the limitations of this study, methodologically high-quality studies are needed for certain appraisal.