Dihydroartemisinin inhibits restenosis after balloon angioplasty via circHSPA4/miR-19a-5p axis.

Dihydroartemisinin (DHA) inhibits restenosis following balloon angioplasty. However, data on the mechanisms of DHA activity in restenosis remains scant. Here, we investigated the role of circRNAs in mediating the inhibitory activity of DHA in neointimal formation. We used total RNA sequencing data to profile the expression of mRNA, circRNA and small RNA in sham, vascular balloon injury (VBI) and DHA-treated groups. CCK8 and EdU assays were employed to analyze cell proliferation, while qRT-PCR and Western blot were used to analyze the RNA or protein expression. In addition, we used RNA immunoprecipitation and luciferase reporter assay to assess the binding of circHSPA4 with miR-19a-5p. RNA sequencing demonstrated that circHSPA4 was upregulated in VBI. Treatment with DHA effectively suppressed the upregulation of the circHSPA4. In addition, analysis of platelet-derived growth family factor bb (PDGFbb)-induced HA-VSMCs showed upregulation of circHSPA4, which was associated with cell proliferation and differentiation. CircHSPA4 was shown to induce dedifferentiation and proliferation of smooth muscle cells. PDGFBB-induced overexpression of CircHSPA4 in HA-VSMCs led to suppression of miR-19a-5p, a phenomenon that was reversed by DHA, in concentration-dependent fashion. In addition, miR-19a-5p reduced the dedifferentiation and proliferation of the smooth muscle cells. Our data demonstrated that CircHSPA4 regulates proliferation and differentiation of smooth muscle cells. DHA and miR-19a-5p modulates CircHSPA4 and can be used as coated drugs on balloon catheter to improve the success rate of vascular remodeling.

Dihydroartemisinin ameliorates balloon injury-induced neointimal formation through suppressing autophagy in vascular smooth muscle cells.

The present study was designed to investigate the therapeutic effects of injection of dihydroartemisinin (DHA) into the balloon-injured carotid arteries on balloon injury-induced neointimal formation and to explore whether autophagy is involved in the action of DHA. Percutaneous transluminal balloon angioplasty was performed in Sprague-Dawley rats to induce neointimal formation, immediately after which DHA (100 µmol/l × 1 ml) and/or Rapamycin (1 mg/100 µl), were injected into the balloon-injured carotid arteries. After 14 days, the serum samples and carotid artery tissues were harvested for analysis. Rat aortic vascular smooth muscle cells (VSMCs) were pretreated with DMSO (vehicle), DHA (1, 10, and 100 µmol/l), or 3-methyladenine (3-MA; 10 mM) for 1 h and then stimulated with platelet-derived growth factor-BB (PDGF-BB; 10 ng/ml) for another 24 h. Animal experiments showed that DHA attenuated the balloon injury-induced neointimal formation, inflammation and VSMC phenotypic transition by inhibiting the balloon injury-induced autophagy activation. In vitro results showed that DHA attenuated the PDGF-BB-induced VSMC phenotypic transition, proliferation, and migration by inhibiting the PDGF-BB-induced autophagy activation. Taken together, DHA ameliorates balloon injury-induced neointimal formation through suppressing autophagy. This study provides insights into the development of a drug-eluting stent using DHA.

Dihydroartemisinin Inhibits TGF-ß-Induced Fibrosis in Human Tenon Fibroblasts via Inducing Autophagy.

BACKGROUND: The formation of hypertrophic scars (HS) can result in the failure of glaucoma surgery, and fibrosis is known to be closely associated with the progression of HS. Dihydroartemisinin (DHA) has been reported to inhibit the progression of fibrosis; however, whether DHA can alleviate the formation of HS remains unclear. METHODS: In the present study, in order to examine the effects of DHA on the progression of HS, human Tenon's capsule fibroblasts (HTFs) were isolated from patients who underwent glaucoma surgery. In addition, Western blot analysis, microtubule associated protein 1 light chain 3 α staining and reverse transcription-quantitative PCR were performed to detect protein and mRNA expression levels in the HTFs, respectively. Cell proliferation was detected by Ki67 staining. Flow cytometry was used to examine apoptosis and reactive oxygen species (ROS) levels in the HTFs. RESULTS: The results revealed that TGF-ß promoted the proliferation and fibrosis of HTFs; however, DHA significantly reversed the effects of TGF-ß by increasing cell autophagy. In addition, DHA notably induced the apoptosis of TGF-ß-stimulated HTFs by increasing the ROS levels, while these increases were partially reversed by 3-methyladenine. Furthermore, DHA notably increased the expression of microRNA (miR)-145-5p in HTFs in a dose-dependent manner. CONCLUSION: The present study demonstrated that DHA inhibits the TGF-ß-induced fibrosis of HTFs by inducing autophagy. These findings may aid in the development of novel agents for the prevention of the formation of HS following glaucoma surgery.

Rapid PCR-free meat species mitochondrial DNA identification using Electric Field Induced Release and Measurement (EFIRM®).

This work details the usage of EFIRM® (Electric Field Induced Release and Measurement) for PCR-free rapid electrochemical detection of mitochondrial DNA. EFIRM® was able to perform highly sensitive detection of animal species for meat contamination testing without multistep sample lysis, DNA extraction, or PCR amplification steps, demonstrating the capability to detect the presence of foreign meat species that only constituted 0.1% of the total mass of a food sample (achieving sensitivity equivalent to that of PCR). The EFIRM® strategy utilizes surface immobilized nucleic acid probes that complement to mitochondrial sequence of Ovis Aries, Sus Scrofa, and Bos Taurus and are immobilized in a polypyrrole matrix on a 96-electrode array. Quantification was performed through amperometric measurement of oxidation-reduction reactions on a streptavidin-peroxidase enzyme chain that completes the nucleic acid complex. All electrochemical procedures were performed using a high-throughput potentiostat system that allows parallelized electrochemical measurement and interfacing to the 96-electrode array.

Dihydroartemisinin ameliorates balloon injury-induced neointimal formation in rats.

OBJECTIVES: This study aims to evaluate the effects of dihydroartemisinin (DHA) on the balloon injury-induced neointimal formation in rats and to investigate the underlying mechanism. METHODS: The balloon-induced carotid artery injury model was established in male Sprague-Dawley rats, immediately after which the DHA solution was injected into the tail vein of rats. In in vitro assays, primary rat vascular smooth muscle cells (VSMCs) were pretreated with DHA and then coincubated with LPS. RESULTS: DHA ameliorated the induced neointimal formation and fibrosis but enhanced apoptosis in rat carotid artery after balloon injury. Furthermore, DHA suppressed migration and enhanced apoptosis of the lipopolysaccharide (LPS)-treated primary VSMCs in vitro. Moreover, in both the balloon injury-induced rat sera and the LPS-treated VSMCs, DHA significantly inhibited proinflammatory cytokines, including interleukin-1ß, tumor necrosis factor-ɑ, and matrix metalloproteinase-1. Importantly, DHA significantly decreased the balloon injury-increased expression of nuclear factor kappa B (NF-κB) subunit NF-κB p65 expression, and increased the balloon injury-reduced expression of inhibitor of NF-κB-alpha, indicating the inhibition of the IκB/NF-κB pathway. CONCLUSION: DHA significantly inhibited neointimal formation in balloon-induced rat carotid artery injury and the mechanism may be related to the inhibition of IκB/NF-κB signaling, which alleviates the inflammatory response.

Bioinformatic Analysis Identifies Three Potentially Key Differentially Expressed Genes in Peripheral Blood Mononuclear Cells of Patients with Takayasu's Arteritis.

OBJECTIVES: This study aimed to identify several potentially key genes associated with the pathogenesis of Takayasu's arteritis (TA). This identification may lead to a deeper mechanistic understanding of TA etiology and pave the way for potential therapeutic approaches. MATERIALS AND METHODS: In this experimental study, the microarray dataset GSE33910, which includes expression data for peripheral blood mononuclear cell (PBMC) samples isolated from TA patients and normal volunteers, was downloaded from the publicly accessible Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were identified in PBMCs of TA patients compared with normal controls. Gene ontology (GO) enrichment analysis of DEGs and analysis of protein-protein interaction (PPI) network were carried out. Several hub proteins were extracted from the PPI network based on node degrees and random walk algorithm. Additionally, transcription factors (TFs) were predicted and the corresponding regulatory network was constructed. RESULTS: A total of 932 DEGs (372 up- and 560 down-regulated genes) were identified in PBMCs from TA patients. Interestingly, up-regulated and down-regulated genes were involved in different GO terms and pathways. A PPI network of proteins encoded by DEGs was constructed and RHOA, FOS, EGR1, and GNB1 were considered to be hub proteins with both higher random walk score and node degree. A total of 13 TFs were predicted to be differentially expressed. A total of 49 DEGs had been reported to be associated with TA in the Comparative Toxicogenomics Database (CTD). The only TA marker gene in the CTD database was NOS2, confirmed by three studies. However, NOS2 was not significantly altered in the analyzed microarray dataset. Nevertheless, NOS3 was a significantly down-regulated gene and was involved in the platelet activation pathway. CONCLUSIONS: RHOA, FOS, and EGR1 are potential candidate genes for the diagnosis and therapy of TA.

Risk factors and medico-economic effect of pancreatic fistula after pancreaticoduodenectomy.

The study aimed to uncover the risk factors for the new defined pancreatic fistula (PF) and clinical related PF (CR-PF) after pancreaticoduodenectomy (PD) surgery and to evaluate the medico-economic effect of patients. A total of 412 patients were classified into two groups according to different criteria, PF and NOPF according to PF occurrence: CR-PF (grades B and C) and NOCR-PF (grade A) based on PF severity. A total of 28 factors were evaluated by univariate and multivariate logistic regression test. Hospital charges and stays of these patients were assessed. The results showed that more hospital stages and charges are needed for patients in PF and CR-PF groups than in NOPF and NOCR-PF groups (P < 0.05). The excessive drinking, soft remnant pancreas, preoperative albumin, and intraoperative blood transfusion are risk factors affecting both PF and CR-PF incidence. More professional surgeons can effectively reduce the PF and CR-PF incidence. Patients with PF and CR-PF need more hospital costs and stages than that in NOPF and NOCR-PF groups. It is critical that surgeons know the risk factors related to PF and CR-PF so as to take corresponding therapeutic regimens for each patient.

The effects of artemisinin on the proliferation and apoptosis of vascular smooth muscle cells of rats.

OBJECTIVE: The aim of this study was to evaluate the effect of artemisinin on the proliferation and apoptosis of rat vascular smooth muscle cells (VSMCs). METHOD: Primary rat VSMCs were treated with various doses of artemisinin. Cell proliferation was evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, and the messenger RNA and protein expressions of proliferating cell nuclear antigen were determined by reverse-transcription polymerase chain reaction and immunohistochemistry. Apoptosis was measured using annexin V and propidium iodide double staining evaluated by flow cytometry. Protein expression of Bax, Bcl2, and cyclin-dependent kinase 4 was determined by Western blot. RESULTS: After 72 h of treatment, artemisinin significantly inhibited VSMC proliferation in a dose-dependent manner. Treatment with 1 mM artemisinin for 72 h significantly reduced the expression of proliferating cell nuclear antigen messenger RNA. On the other hand, the same treatment increased the apoptosis of VSMCs, the activation of caspase-3, the Bax protein expression, and the Bax/Bcl2 ratio. CONCLUSION: The results suggest that artemisinin can effectively inhibit VSMC proliferation and induce VSMC apoptosis.

Serum bilirubin level is negatively correlated with disease progression of peripheral arterial disease: an observational cohort study.

We determined whether low bilirubin level is a risk factor for peripheral arterial disease (PAD). We recruited 318 patients with PAD and 100 healthy volunteers. Patients were divided into 4 groups by the Fontaine classification for PAD, namely, group 1 (grade 1, n = 4); group 2 (grade 2, n = 114), group 3 (grade 3, n = 164), and group 4 (grade 4, n = 36). Total bilirubin (T-BIL), direct bilirubin (D-BIL), and indirect bilirubin (I-BIL) levels were compared using stepwise multiple regressions adjusted for selected factors. After adjusting for gender, age, smoking, and diastolic blood pressure, serum levels of T-BIL, D-BIL, and I-BIL were significantly lower in the PAD group (P < .05). Patients with grade 4 PAD showed significantly (P < .05) lower levels of T-BIL when compared with grade 2 patients. We concluded that serum bilirubin levels are negatively correlated with the severity and progression of PAD.

The three-dimensional nanofiber scaffold culture condition improves viability and function of islets.

Significant problems existing in the islet transplantation include a poor survival ability of the islet cells cultured under static conditions in vitro, decreased secretion function, and limited transplantation efficiency. In this study, we cocultured the three-dimensional (3D) self-assembling peptide nanofiber hydrogel scaffold with the islets from adult Wistar rats. The nanofiber scaffold constructed a 3D environment for the islets culture. The results of DTZ staining showed that the purity of the islets in the scaffold was >80%. The result of the fluorescent staining with AO-PI demonstrated that the viability of the islets in the 3D culture environment (within scaffold) was greater than those in the two-dimensional (2D) culture environment (without scaffold). The islets encapsulated in the 3D peptide nanofiber scaffold exhibited better secretion function. The insulin releasing index in the 3D group was remarkably higher than that in the 2D group. By scanning electron microscopy, it was observed that the 3D self-assembling peptide nanofiber hydrogel scaffold formed a nano scale fiber with a geometric form and the islets were encapsulated in this scaffold. Our research demonstrated that this nanofiber scaffold provided a favorable 3D environment for the islets to be cultured in vitro and then improve the secretion function and prolong the survival time of the islet in vitro.