Ceramides and sphingosine-1-phosphate mediate the distinct effects of M1/M2-macrophage infusion on liver recovery after hepatectomy.

Post-hepatectomy liver dysfunction is a life-threatening morbidity that lacks efficient therapy. Bioactive lipids involved in macrophage polarization crucially regulate tissue injury and regeneration. Herein, we investigate the key bioactive lipids that mediate the cytotherapeutic potential of polarized-macrophage for post-hepatectomy liver dysfunction. Untargeted lipidomics identified elevation of ceramide (CER) metabolites as signature lipid species relevant to M1/M2 polarization in mouse bone-marrow-derived-macrophages (BMDMs). M1 BMDMs expressed a CER-generation-metabolic pattern, leading to elevation of CER; M2 BMDMs expressed a CER-breakdown-metabolic pattern, resulting in upregulation of sphingosine-1-phosphate (S1P). After infusing M1- or M2-polarized BMDMs into the mouse liver after hepatectomy, we found that M1-BMDM infusion increased M1 polarization and CER accumulation, resulting in exaggeration of hepatocyte apoptosis and liver dysfunction. Conversely, M2-BMDM infusion enhanced M2 polarization and S1P generation, leading to alleviation of liver dysfunction with improved hepatocyte proliferation. Treatment of exogenous CER and S1P or inhibition CER and S1P synthesis by siRNA targeting relevant enzymes further revealed that CER induced apoptosis while S1P promoted proliferation in post-hepatectomy primary hepatocytes. In conclusion, CER and S1P are uncovered as critical lipid mediators for M1- and M2-polarized BMDMs to promote injury and regeneration in the liver after hepatectomy, respectively. Notably, the upregulation of hepatic S1P induced by M2-BMDM infusion may have therapeutic potential for post-hepatectomy liver dysfunction.

Electrolyte disorders are ERAS-associated in patients undergoing hepato-pancreato-biliary surgery.

PURPOSE: Emerging evidences have raised concerns about electrolyte disorders caused by restrictive fluid management in the enhanced recovery after surgery (ERAS) protocol. This study aims to investigate the morbidity and treatment of electrolyte disorders associated with ERAS in patients undergoing hepato-pancreato-biliary (HPB) surgery. METHODS: Clinical data from 157 patients under the ERAS program and 166 patients under the traditional (Non-ERAS) program after HPB surgery were retrospectively analyzed. Risk factors and predictive factors of postoperative electrolyte disorders were analyzed by logistic regression analysis and receiver operator characteristic (ROC) curve analysis, respectively. RESULTS: The average of intravenous fluid, sodium, chloride, and potassium supplementation after surgery were significantly lower in the ERAS group. Hypokalemia was the most common type of electrolyte disorders in the ERAS group, whose incidence was substantially increased compared to that in the Non-ERAS group [28.77% vs. 8.97%, p < 0.001, on postoperative (POD) 5]. Logistic regression analysis identified the ERAS program and age as independent risk factors of hypokalemia. ROC curve analysis identified serum potassium levels below 3.76 mmol/L on POD 3 (area under curve 0.731, sensitivity 58.54%, specificity 82.69%) as a predictive factor for postoperative hypokalemia in ERAS patients. Oral supplementation at an average of 35.41 mmol potassium per day was effective in restoring the ERAS-associated hypokalemia. CONCLUSIONS: ERAS procedures were particularly associated with a lower supplementation of potassium and a higher incidence of hypokalemia in patients after HPB surgery. Oral potassium supplementation could be an adopted ERAS program for the elderly undergoing HPB surgery.

Correction: Targeting alkaline ceramidase 3 alleviates the severity of nonalcoholic steatohepatitis by reducing oxidative stress.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Lipid oversupply induces CD36 sarcolemmal translocation via dual modulation of PKCζ and TBC1D1: an early event prior to insulin resistance.

Lipid oversupply may induce CD36 sarcolemmal translocation to facilitate fatty acid transport, which in turn causes dyslipidemia and type 2 diabetes. However, the underlying mechanisms of CD36 redistribution are still yet to be unraveled. Methods: High fat diet fed mice and palmitate/oleic acid-treated L6 cells were used to investigate the initial events of subcellular CD36 recycling prior to insulin resistance. The regulation of CD36 sarcolemmal translocation by lipid oversupply was assessed by insulin tolerance test (ITT), oral glucose tolerance test (OGTT), glucose/fatty acid uptake assay, surface CD36 and GLUT4 detection, and ELISA assays. To elucidate the underlying mechanisms, specific gene knockout, gene overexpression and/or gene inhibition were employed, followed by Western blot, co-immunoprecipitation, immunostaining, and kinase activity assay. Results: Upon lipid/fatty acid overload, PKCζ activity and TBC1D1 phosphorylation were enhanced along with increased sarcolemmal CD36. The inhibition of PKCζ or TBC1D1 was shown to block fatty acid-induced CD36 translocation and was synergistic in impairing CD36 redistribution. Mechanically, we revealed that AMPK was located upstream of PKCζ to control its activity whereas Rac1 facilitated PKCζ translocation to the dorsal surface of the cell to cause actin remodeling. Furthermore, AMPK phosphorylated TBC1D1 to release retained cytosolic CD36. The activated PKCζ and phosphorylated TBC1D1 resulted in a positive feedback regulation of CD36 sarcolemmal translocation. Conclusion: Collectively, our study demonstrated exclusively that lipid oversupply induced CD36 sarcolemmal translocation via dual modulation of PKCζ and TBC1D1, which was as an early event prior to insulin resistance. The acquired data may provide potential therapy targets to prevent lipid oversupply-induced insulin resistance.

Targeting alkaline ceramidase 3 alleviates the severity of nonalcoholic steatohepatitis by reducing oxidative stress.

Overload of palmitic acids is linked to the dysregulation of ceramide metabolism in nonalcoholic steatohepatitis (NASH), and ceramides are important bioactive lipids mediating the lipotoxicity of palmitic acid in NASH. However, much remains unclear about the role of ceramidases that catalyze the hydrolysis of ceramides in NASH. By analyzing the National Center for Biotechnology Information (NCBI) Gene Expression Omnibus (GEO) database, we found that alkaline ceramidase 3 (ACER3) is upregulated in livers of patients with NASH. Consistently, we found that Acer3 mRNA levels and its enzymatic activity were also upregulated in mouse livers with NASH induced by a palmitate-enriched Western diet (PEWD). Moreover, we demonstrated that palmitate treatment also elevated Acer3 mRNA levels and its enzymatic activity in mouse primary hepatocytes. In order to investigate the function of Acer3 in NASH, Acer3 null mice and their wild-type littermates were fed a PEWD to induce NASH. Knocking out Acer3 was found to augment PEWD-induced elevation of C18:1-ceramide and alleviate early inflammation and fibrosis but not steatosis in mouse livers with NASH. In addition, Acer3 deficiency attenuated hepatocyte apoptosis in livers with NASH. These protective effects of Acer3 deficiency were found to be associated with suppression of hepatocellular oxidative stress in NASH liver. In vitro studies further revealed that loss of ACER3/Acer3 increased C18:1-ceramide and inhibited apoptosis and oxidative stress in mouse primary hepatocytes and immortalized human hepatocytes induced by palmitic-acid treatment. These results suggest that ACER3 plays an important pathological role in NASH by mediating palmitic-acid-induced oxidative stress.

The preventive effect of Chinese herbal preparation Xuebijing against hyperactive inflammation after hepato-pancreato-biliary surgery.

BACKGROUND: Hepato-pancreato-biliary (HPB) surgery is a primary treatment for benign and malignant diseases of the liver, biliary tract, and pancreas. Hyperactive inflammation has been indicated as a critical risk factor of post-operation death after HPB surgery. Xuebijing is an anti-inflammatory intravenous herbal preparation made from traditional Chinese medicines. Emerging evidence has implicated a protective role of Xuebijing against hyperactive inflammation. METHODS: A retrospective cohort study was conducted. We analyzed a total of 638 cases of HPB surgery, including hepatectomy, Whipple's surgery, and surgeries for cholelithiasis, which were divided into a Xuebijing treatment group and a conventional treatment group according to whether they were treated with Xuebijing injection or not. Clinical data related to liver function and inflammation were compared between the two groups after operation, including liver function index, white blood cell (WBC) count, neutrophil percentage (NE%), C-reactive protein (CRP), serum interleukin-6 (IL-6), body temperature, mortality, incidence of adverse reaction, length of postoperative hospital stay, and hospitalization cost. RESULTS: Xuebijing injection was found to decrease the levels of inflammatory markers in the blood significantly, including WBC, NE%, CRP, IL-6, and reduce the incidence of postoperative fever without prolonging in-hospital length or increasing cost compared to the conventional treatment group. Moreover, our data demonstrated that Xuebijing injection did not impact liver function after hepatectomy. CONCLUSIONS: These results suggest that Xuebijing injection alleviates hyperactive inflammation caused by HPB surgery, and support the application of Xuebijing injection as a safe therapeutic approach against hyperactive inflammation in patients with HPB surgery.

miR-942 promotes proliferation and metastasis of hepatocellular carcinoma cells by inhibiting RRM2B.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the leading causes of cancer-related death. MicroRNA-942 (miR-942) plays a critical role in promoting proliferation and metastasis of cancer cells and is associated with poor prognosis in some types of cancers. However, the prognostic value of miR-942 and its functional role in HCC remain unclear. MATERIALS AND METHODS: Real-time PCR (RT-PCR) was used to detect the expression of miR-942 in HCC tissues and adjacent normal liver tissues. Next, the correlations between miR-942 expression and clinicopathological parameters including the survival rate were analyzed. Interaction between miR-942 and ribonucleotide reductase regulatory TP53 inducible subunit M2B (RRM2B) was determined by RT-PCR, Western blot and luciferase assay. The biological influence of miR-942 on HCC cell lines was studied using CCK-8 assay, colony formation assay and transwell assay in vitro. Western blot and RT-PCR were used to analyze the change of downstream genes after miR-942 mimics transfection. RESULTS: miR-942 was significantly up-regulated in HCC. Its high expression was associated with serum alanine transaminase level (P=0.0350), tumor size (P=0.0195), T stage (P=0.0045) and lymphatic metastasis (P=0.0013). High expression of miR-942 was associated with shorter overall survival and disease-free survival time of HCC patients. RRM2B was validated as a target gene of miR-942. miR-942 mimics markedly promoted the malignant phenotypes of Huh7 and MHCC97H cell lines, while its inhibitor had the opposite effect. miR-942 can regulate the downstream genes of RRM2B including Egr-1 and PTEN, markers of epithelial-mesenchymal transition and matrix metalloproteinases. CONCLUSION: miR-942 may serve as a potential biomarker for HCC and its inhibitor may be a therapeutic agent for the treatment of this deadly disease.

Stabilized ß-Hairpin Peptide Inhibits Insulin Degrading Enzyme.

Insulin-degrading enzyme (IDE) plays a critical role in both the proteolytic degradation and inactivation of insulin. The exploration of novel IDE inhibitors could aid in the study of novel therapeutics for type-2 diabetes. Herein, we report a hypothesized stabilized ß-hairpin peptide that can efficiently inhibit the enzymatic activity of IDE. The resulting stabilized peptide B35 is demonstrated to activate the AKT phosphorylation pathway in skeletal muscle cells and is shown to slow insulin degradation. An 80 mg kg-1 intraperitoneal (i.p.) injection of the stabilized ß-hairpin peptide B35 is demonstrated to improve glucose tolerance during an oral glucose tolerance test in obese mouse model. We note that this stabilized peptide exhibited negligible cytotoxicity in both in vitro and in vivo assays, even at high concentrations (300 µM). This study suggests that IDE peptide inhibitors could function as potentially meaningful candidates for the development of type-2 diabetes therapeutics.

Protective effect of gadolinium chloride on early warm ischemia/reperfusion injury in rat bile duct during liver transplantation.

BACKGROUND: Activation of Kupffer cell (KC) is acknowledged as a key event in the initiation and perpetuation of bile duct warm ischemia/reperfusion injury. The inhibitory effect of gadolinium chloride (GdCl(3)) on KC activation shows potential as a protective intervention in liver injury, but there is less research with regard to bile duct injury. METHODS: Sixty-five male Sprague-Dawley rats (200-250 g) were randomly divided into three experimental groups: a sham group (n = 15), a control group (n = 25), and a GdCl(3) group (n = 25). Specimen was collected at 0.5, 2, 6, 12 and 24 h after operation. Alanine aminotransferase (ALT), alkaline phosphatase (ALP) and total bilirubin (TBIL) of serum were measured. Tumor necrosis factor-α (TNF-α), Capase-3 activity and soluble Fas (sFas) were detected. The pathologic changes of bile duct were observed. Immunochemistry for bile duct Fas was performed. Apoptosis of bile duct cells was evaluated by the terminal UDP nick end labeling assay. RESULTS: GdCl(3) significantly decreased the levels of ALT, ALP and TBIL at 2, 6, 12, and 24 h, and increased serum sFas at 2, 6 and 12 h (P<0.05). TNF-α was lower in the GdCl(3) group than in the control group at 2, 6, 12 and 24 h (P<0.05). Preadministration of GdCl(3) significantly reduced the Caspase-3 activity and bile duct cell apoptosis at 2, 6, 12 and 24 h. After operation for 2, 6 and 12 h, the expression of Fas protein was lower in the GdCl(3) group than in the control group (P<0.05). CONCLUSIONS: GdCl(3) plays an important role in suppressing bile duct cell apoptosis, including decreasing ALT, ALP, TBIL and TNF-α; suppressing Fas-FasL-Caspase signal transduction during transplantation.