Cholesterol efflux regulator ABCA1 exerts protective role against high shear stress-induced injury of HBMECs via regulating PI3K/Akt/eNOS signaling.

BACKGROUND: In brain, microvascular endothelial cells are exposed to various forces, including shear stress (SS). However, little is known about the effects of high shear stress (HSS) on human brain microvascular endothelial cells (HBMECs) and the underlying mechanism. The cholesterol efflux regulator ATP-binding cassette subfamily A member 1 (ABCA1) has been demonstrated to exert protective effect on HBMECs. However, whether ABCA1 is involved in the mechanism underneath the effect of HSS on HBMECs remains obscure. In the present study, a series of experiments were performed to better understand the effect of HSS on cellular processes of HBMECs and the possible involvement of ABCA1 and PI3K/Akt/eNOS in the underlying mechanisms. RESULTS: HBMECs were subjected to physiological SS (PSS) or high SS (HSS). Cell migration was evaluated using Transwell assay. Apoptotic HBMECs were detected by flow cytometry or caspase3/7 activity. IL-1ß, IL-6, MCP-1 and TNF-α levels were measured by ELISA. RT-qPCR and western blotting were used for mRNA and protein expression detection, respectively. ROS and NO levels were detected using specific detection kits. Compared to PSS, HBMECs exhibited decreased cell viability and migration and increased cell apoptosis, increased levels of inflammatory cytokines, and improved ROS and NO productions after HSS treatment. Moreover, HSS downregulated ABCA1 but upregulated the cholesterol efflux-related proteins MMP9, AQP4, and CYP46 and activated PI3K/Akt/eNOS pathway. Overexpression of ABCA1 in HBMECS inhibited PI3K/Akt/eNOS pathway and counteracted the deleterious effects of HSS. Contrary effects were observed by ABCA1 silencing. Inhibiting PI3K/Akt/eNOS pathway mimicked ABCA1 effects, suggesting that ABCA1 protects HBMECs from HSS via PI3K/Akt/eNOS signaling. CONCLUSION: These results advanced our understanding on the mechanisms of HSS on HBMECs and potentiated ABCA1/PI3K/Akt/eNOS pathway as therapeutic target for cerebrovascular diseases.

[Geniposide inhibits hepatic fibrosis and hepatic stellate cell activation through blocking the TGF-ß1/Smad signaling pathway].

The purpose of this study was to investigate the effect of Geniposide on hepatic fibrosis and activation of hepatic stellate cells (HSCs) and to explore possible underlying mechanism. Human HSCs (LX-2) were treated with 5 ng/mL transforming growth factor-ß1 (TGF-ß1), followed by co-culture with Geniposide at various concentrations (0, 1, 2.5, 5, 10, 20, 40, 60, 80, 100 µmol/L). Cell viability was determined by MTT assay. Then, LX-2 cells were divided into control, TGF-ß1 (5 ng/mL) and TGF-ß1 + Geniposide (20 µmol/L) groups, and the gene and protein expression of collagen I, fibronectin, α-smooth muscle actin (α-SMA), p-Smad2 and p-Smad3 was detected by qPCR and Western blot, respectively. BALB/c mice were treated with CCl4 (25%, 1 mL/kg) to generate a model of hepatic fibrosis (CCl4 group), and the control group and CCl4 + Geniposide group were administered with olive oil and CCl4 + 40 mg/kg Geniposide, respectively. After 4 weeks of treatment, the liver function and serum hepatic fibrosis indexes of mice were detected, histological observation was performed by HE and Masson staining, and α-SMA expression in the tissue was analyzed by immunohistochemistry. Western blot was utilized for the determination of the protein expression of α-SMA, TGF-ß1, p-Smad2 and p-Smad3. The results showed that Geniposide inhibited LX-2 cell proliferation. In addition, Geniposide significantly downregulated the gene and protein expression of collagen I, fibronectin and α-SMA and the expression of TGF-ß1/Smad signaling-related proteins induced by TGF-ß1 in vitro. Histological observations showed that Geniposide significantly inhibited CCl4-induced hepatic fibrosis, HSC activation and expression of TGF-ß1/Smad signaling-related proteins in mice. In summary, Geniposide prevents the hepatic fibrosis and HSC activation possibly through the inhibition of the TGF-ß1/Smad signaling pathway.

Experimental study of fat derived pellets promoting wound healing in rats.

To observe the effect of fat-derived pellets (FDP) on wound healing in rats, the inguinal fat of rats was obtained, and the FDP were obtained after centrifugation. The cell activity and growth factor secretion of FDP were measured. The wounds in rats were created, and FDP was used to treat the wounds of rats. The phenotype of macrophages and the expression of angiogenic factors expression in wounds were measured. The cell viability in FDP remains in high level after centrifugation and the expression of vascular endothelial growth factor (VEGF) and Basic Fibroblast Growth Factor (bFGF) from FDP was observed in vitro. The FDP significantly promoted the wound healing of rats compared with that in control groups. Moreover, the expression of M2 macrophages and VEGF in FDP group were significantly higher than that in the control group. FDP is a kind of stem cell product, which can be obtained from adipose tissue by physical centrifugation. The cytotherapeutic effect of FDP makes it a promising product for wound healing in clinics.

miR-25-5p regulates endothelial progenitor cell differentiation in response to shear stress through targeting ABCA1.

The importance of flow shear stress (SS) on the differentiation of endothelial progenitor cells (EPCs) has been demonstrated in various studies. Cholesterol retention and microRNA regulation have been also proposed as relevant factors involved in this process, though evidence regarding their regulatory roles in the differentiation of EPCs is currently lacking. In the present study on high shear stress (HSS)-induced differentiation of EPCs, we investigated the importance of ATP-binding cassette transporter 1 (ABCA1), an important regulator in cholesterol efflux, and miR-25-5p, a potential regulator of endothelial reconstruction. We first revealed an inverse correlation between miR-25-5p and ABCA1 expression levels in EPCs under HSS treatment; their direct interaction was subsequently validated by a dual-luciferase reporter assay. Further studies using flow cytometry and quantitative polymerase chain reaction demonstrated that both miR-25-5p overexpression and ABCA1 inhibition led to elevated levels of specific markers of endothelial cells, with concomitant downregulation of smooth muscle cell markers. Finally, knockdown of ABCA1 in EPCs significantly promoted tube formation, which confirmed our conjecture. Our current results suggest that miR-25-5p might regulate the differentiation of EPCs partially through targeting ABCA1, and such a mechanism might account for HSS-induced differentiation of EPCs.

[The Thyroid Hormone Receptor Mediated Luciferase Reporter Gene Assays Screening for EDCs].

OBJECTIVE: This study in order to use report gene assay based on the thyroid hormone receptor (TR) α/ß from human origin for screening endocrine disruptors chemicals (EDCs), evaluating the thyroid hormone activity of Bisphenol (BPA), 1-Naphthaleny methyl carbamate and 1-naphthol (1-NAP). METHODS: Using Rhesus monkey kidney cells (LLC-MK2) as transfection cell to establish the gene report assay system based on pGL-3-promega and pGL4.27 of TRα/ß through the method of transient transfection. Using T3 and T4 as positive subjects to evaluation the effectiveness of two detection systems and detect the thyroid hormone activity of BPA, 1-Naphthaleny methyl carbamate, 1-NAP. RESULTS: The TRß LLC-MK2 report gene assay based on pGL3-promega, the minimum detectable limit of T3 is 1.216×10-11 mol/L, the largest induction multiple was shown at 7.482×10-6 mol/L, the expression multiple of induced Lucifrerase was 5.98-fold that of the vehicle control, the EC50 was 3.327×10-8 mol/L; The minimum detectable limit of T4 was 1.622×10-8 mol/L, the largest induction Luc expression was 3.4-fold of vehicle control, the EC50 was 2.213×10-7 mol/L. The TRß LLC-MK2 report gene assay based on pGL4.27, the minimum detectable limit of T3 was 9.863×10-12 mol/L, the largest induction Luc expression as shown at 1.671×10-6 mol/L, resulting in 8.57-fold of vehicle control, the EC50 is 3.327×10-8 mol/L. The minimum detectable limit of T4 was 1.349×10-9 mol/L, the largest induction Luc expression was 4.6-fold of vehicle control, the EC50 is 4.074×10-7 mol/L. There was no thyroid hormone activity by using TRß report gene assay to evaluate BPA, 1-Naphthaleny methyl carbamate or 1-NAP, but 1-Naphthaleny methyl carbamate and 1-NAP have some degree receptor antagonism. CONCLUSIONS: The TRß LLC-MK2 report gene assay based on pGL3- promega and pGL4.27 show highly sensitive (pGL4.27 relatively higher), can be used to screen for EDCS and test chemical thyroid hormone activity effectively.

Prognostic significance of neutrophil to lymphocyte ratio in patients with gastrointestinal stromal tumors: A meta-analysis.

BACKGROUND: The neutrophil to lymphocyte ratio (NLR) is reported to be a prognostic factor in multiple malignancies. However, its prognostic value in patients with gastrointestinal stromal tumors (GISTs) remains controversial. This study aims to evaluate the prognostic value of preoperative NLR in GISTs. METHODS: MEDLINE, EMBASE, and, Cochrane databases were searched until February 2017. Eligible articles were defined as studies assessing the prognostic role of preoperative NLR in GISTs. The end points were overall survival (OS), disease-free survival (DFS), recurrence-free survival (RFS), and clinicopathological parameters. Pooled hazard ratios (HRs) or odds ratios (ORs) with 95% confidence intervals (CIs) were calculated using fixed-effects/random-effects models. RESULTS: A total of eight studies comprising 1676 patients with GISTs were included. Elevated NLR had an association with decreased DFS/RFS (HR: 2.18, 95% CI: 1.30-3.67, P=0.003), but not OS (HR: 1.74, 95% CI: 0.63-4.84, P=0.29). The findings from most subgroup analyses were consistent with those from the overall analysis. Moreover, high NLR was significantly correlated with male, stomach lesion, tumor size (>5cm), tumor rupture (+), tumor recurrence (+), mitotic index (>5/50HPF), and NIH risk category (high/intermediate). CONCLUSIONS: Elevated preoperative NLR may be an unfavorable prognostic biomarker in patients with GISTs.

Platelet to lymphocyte ratio in biliary tract cancer: Review and meta-analysis.

BACKGROUND: The platelet to lymphocyte ratio (PLR) has been found to predict clinical outcomes in multiple malignancies. The aim of this study was to assess the prognostic value of pretreatment PLR in biliary tract cancer (BTC). METHODS: We searched the MEDLINE, EMBASE, and Cochrane databases to identify studies evaluating the prognostic significance of pretreatment PLR in BTC. The end points were overall survival (OS), recurrence-free survival (RFS). Pooled hazard ratios (HRs) or odds ratios (ORs) with 95% confidence intervals (CIs) were calculated using fixed-effects/random-effects models. RESULTS: A total of eleven studies comprising 2392 patients were included in the study. Pooled results showed that elevated PLR was significantly associated with decreased overall survival (HR: 1.59, 95% confidence interval [CI]: 1.42-1.78, p<0.001) and recurrence-free survival (HR: 1.53, 95% CI: 1.16-2.00, p=0.002). Subgroup analyses suggested that a high PLR predicted decreased OS in patient with BTC, regardless of sample size (<200 or ≥200), treatment methods (surgery, mixed, or chemotherapy), tumor stage (mixed or metastatic), analysis methods (univariate or multivariate), cut-off values (<150 or ≥150), and NOS score (<7 or ≥7). CONCLUSIONS: Elevated pretreatment PLR may be an unfavorable prognostic factor for clinical outcomes in patients with biliary tract cancer.

Binding pancreaticojejunostomy: 150 consecutive cases without leakage.

The objective of this study was to verify the safety of a new technique termed "binding pancreaticojejunostomy" in a prospective cohort study. Pancreaticojejunal anastomostic leakage is a major cause of morbidity and mortality after pancreaticoduodenectomy. To prevent the development of pancreatic fistulas, we designed a special technique that we termed binding pancreaticojejunostomy. Binding pancreaticojejunostomy entails binding 3 cm of the serosamuscular sheath of the jejunum to the intussuscepted pancreatic stump. From January 1996 to May 2001, a total of 150 consecutive patients were treated with this type of pancreaticojejunostomy, including typical pancreaticoduodenectomy in 120, hepatopancreaticoduodenectomy in 17, pylorus-preserving pancreaticoduodenectomy in 10, and duodenal-preserving resection of the head of the pancreas in three. None of the patients developed pancreatic fistulas. The overall morbidity was 31.3%. The following complications occurred: gastrointestinal bleeding in six, pulmonary infection in 12, wound infection in 20, delayed gastric emptying in three, incision dehiscence in four, and hepatic insufficiency in two. The mean postoperative hospital stay was 19.8 +/- 5 days. Binding pancreaticojejunostomy is a safe, simple, and effective technique.