Kai Xu

MicroRNA-31 controls phenotypic modulation of human vascular smooth muscle cells by regulating its target gene cellular repressor of E1A-stimulated genes

Phenotypic modulation of vascular smooth muscle cells (VSMCs) plays a critical role in the pathogenesis of a variety of proliferative vascular diseases. The cellular repressor of E1A-stimulated genes (CREG) has been shown to play an important role in phenotypic modulation of VSMCs. However, the mechanism regulating CREG upstream signaling remains unclear. MicroRNAs (miRNAs) have recently been found to play a critical role in cell differentiation via target-gene regulation. This study aimed to identify a miRNA that binds directly to CREG, and may thus be involved in CREG-mediated VSMC phenotypic modulation. Computational analysis indicated that miR-31 bound to the CREG mRNA 3 untranslated region (3-UTR). miR-31 was upregulated in quiescent differentiated VSMCs and downregulated in proliferative cells stimulated by platelet-derived growth factor and serum starvation, demonstrating a negative relationship with the VSMC differentiation marker genes, smooth muscle α-actin, calponin and CREG. Using gain-of-function and loss-of-function approaches, CREG and VSMC differentiation marker gene expression levels were shown to be suppressed by a miR-31 mimic, but increased by a miR-31 inhibitor at both protein and mRNA levels. Notably, miR-31 overexpression or inhibition affected luciferase expression driven by the CREG 3-UTR containing the miR-31 binding site. Furthermore, miR-31-mediated VSMC phenotypic modulation was inhibited in CREG-knockdown human VSMCs. We also determined miR-31 levels in the serum of patients with coronary artery disease (CAD), with or without in stent restenosis and in healthy controls. miR-31 levels were higher in the serum of CAD patients with restenosis compared to CAD patients without restenosis and in healthy controls. In summary, these data demonstrate that miR-31 not only directly binds to its target gene CREG and modulates the VSMC phenotype through this interaction, but also can be an important biomarker in diseases involving VSMC phenotypic modulation. These novel findings may have extensive implications for the diagnosis and therapy of a variety of proliferative vascular diseases.

First-in-man study evaluating the safety and efficacy of a second generation biodegradable polymer sirolimus-eluting stent in the treatment of patients with de novo coronary lesions: Clinical, Angiographic, and OCT outcomes of CREDIT-1

To evaluate the preliminary safety and efficacy of the EXCEL II stent system.

Estrogen-eluting stent implantation inhibits neointimal formation and extracellular signal-regulated kinase activation.

Objectives: The goal of this study was to evaluate the efficacy and the mechanism of 17β‐estradiol‐eluting stents on inhibiting neointimal formation of abdominal aortas in rabbits fed with high fat diet. Background: Animal experiments have shown that local delivery of estrogen reduced neointimal formation after coronary angioplasty. Preliminary trial of estrogen‐eluting stent implantation in patients with coronary disease also suggests a reduction in restenosis. Methods: We implanted 17β‐estradiol‐eluting stents, or control phosphorylcholine‐ coated stents, or bare metal stents of abdominal aortas in rabbits fed with high fat diet. Histology, immunohistochemistry, and Western blot analysis were used to assess the efficacy and mechanism of inhibiting neointimal formation of 17β‐estradiol‐eluting stents. Results: Western blot analysis revealed marked increase in ERK phosphorylation in 30 min after deployment of phosphorylcholine‐coated or bare metal stents, indicating activation of MAP kinase pathway. Immunohistochemistry showed intense staining of phospho‐ERK in the medial smooth muscle cells in stent‐implanted region. Extensive neointimal hyperplasia developed 12 weeks after stenting. In contrast, we observed significant inhibition of ERK phosphorylation and neointimal thickening in estrogen‐eluting stent‐implanted animals. Immunohistochemistry of factor VIII‐related antigen demonstrated an accelerated reendothelialization as compared with the bare metal stent or phosphorylcholine‐coated stent‐implanted controls. Conclusions: Current study suggests that estrogen‐eluting stents reduce neointimal formation and hence prevent restenosis after angioplasty possibly by inhibiting ERK activation in smooth muscle cells and promoting reendothelialization.

One-Time versus Staged Multivessel Intervention in Intermediate to Very High-Risk Patients with Non-ST-Segment Elevation Acute Coronary Syndromes

Background and Objectives To compare clinical outcomes of staged versus one-time percutaneous coronary intervention (PCI) in intermediate to very high-risk patients with non–ST-segment elevation acute coronary syndromes (NSTE-ACS) and multivessel coronary disease (MVD). Subjects and Methods 1531 NSTE-ACS patients with multivessel PCI and meeting the criteria of intermediate to very high risk were screened from a prospectively registered database obtained from General Hospital of Shenyang Military Region between 2008 and 2012. They were categorized into one-time PCI (n=859) and staged PCI (n=672) according to intervention strategy. The primary outcomes included a 3-year major adverse cardiac event (MACE), a composite of cardiac death, myocardial infarction (MI), and target vessel revascularization. Results At 3 years, no significant differences in MACE (20.8% vs. 19.7%, p=0.608) and cardiac death/MI (7.1% vs. 9.1%, p=0.129) were observed between the two groups. After propensity score matching, there was no statistical significance in MACE (18.9% vs. 21.8%, p=0.249); whereas cardiac death/MI was significantly lower in the staged PCI group (7.0% vs.11.1%, p=0.033). Ninety-day landmark analysis showed that the staged PCI group had a lower 90-day incidence of MACE (1.2% vs. 3.3%, p= 0.037) and cardiac death/MI (0.7% vs. 2.6%, p=0.031). For the 90-day to 3-year follow-up period, the incidences of MACE (17.9% vs. 19.1%, p=0.641) and cardiac death/MI (6.3% vs. 8.7%, p=0.191) were similar in both groups. Conclusion In intermediate- to very high-risk NSTE-ACS patients with MVD, staged PCI is superior to one-time PCI in terms of cardiac death/MI.

Comparison of the efficacy of drug-eluting stents versus bare-metal stents for the treatment of left main coronary artery disease.

Background:Recent studies reported that percutaneous coronary intervention with stent implantation was safe and feasible for the treatment of left main coronary artery (LMCA) disease in select patients. However, it is unclear whether drug-eluting stents (DESs) have better outcomes in patients with LMCA disease compared with bare-metal stent (BMS) during long-term follow-up in Chinese populations. Methods:From a perspective multicenter registry, 1136 consecutive patients, who underwent BMS or DES implantation for unprotected LMCA stenosis, were divided into two groups: 1007 underwent DES implantation, and 129 underwent BMS implantation. The primary outcome was the rate of major adverse cardiac events (MACEs), including cardiovascular (CV) death, myocardial infarction (MI), and target lesion revascularization (TLR) at 5 years postimplantation. Results:Patients in the DES group were older and more likely to have hyperlipidemia and bifurcation lesions. They had smaller vessels and longer lesions than patients in the BMS group. In the adjusted cohort of patients, the DES group had significantly lower 5 years rates of MACE (19.4% vs. 31.8%, P = 0.022), CV death (7.0% vs. 14.7%, P = 0.045), and MI (5.4% vs. 12.4%, P = 0.049) than the BMS group. There were no significant differences in the rate of TLR (10.9% vs. 17.8%, P = 0.110) and stent thrombosis (4.7% vs. 3.9%, P = 0.758). The rates of MACE (80.6% vs. 68.2%, P = 0.023), CV death (93.0% vs. 85.3%, P = 0.045), TLR (84.5% vs. 72.1%, P = 0.014), and MI (89.9% vs. 80.6%, P = 0.029) free survival were significantly higher in the DES group than in the BMS group. When the propensity score was included as a covariate in the Cox model, the adjusted hazard ratios for the risk of CV death and MI were 0.41 (95% confidence interval [CI]: 0.21–0.63, P = 0.029) and 0.29 (95% CI: 0.08–0.92, P = 0.037), respectively. Conclusions:DES implantation was associated with more favorable clinical outcomes than BMS implantation for the treatment of LMCA disease even though there was no significant difference in the rate of TLR between the two groups.

Simultaneous Determination of Two Epimeric Furofuran Lignans (Sesamin and Asarinin) of Asarum heterotropoides Extract in Rat Plasma by LC/MS/MS: Application to Pharmacokinetic Study

A rapid, sensitive and selective liquid chromatography-tandem mass spectrometry was developed to determine two epimeric furofuran lignans (sesamin and asarinin) simultaneously from Asarum heterotropoides extract in rat plasma. Simple protein precipitation with acetonitrile was performed to extract analytes by using alantolactone as an internal standard. Chromatographic separation was achieved using a DIKMA Diamonsil C18 analytical column (4.6 mm × 150 mm, i.d., 5 µm) by isocratically eluting with a mobile phase consisting of methanol/5 mM ammonium acetate/formic acid (75:25:0.1, v/v/v) at a flow rate of 0.8 mL/min. Tandem mass spectrometric detection with an electrospray ionization interface was performed by multiple reaction monitoring in positive ionization mode. This method was validated according to specificity, sensitivity, linearity, intra- and inter-day precision (<10.7%) and accuracy (<2.3%) and recovery and stability in a concentration range of 25.0-15 000 ng/mL for sesamin and 5.00-3 000 ng/mL for asarinin. This method has been successfully applied in a pharmacokinetic study of A. heterotropoides extract containing sesamin and asarinin after this extract was orally administrated in rats.

Purification and functional assessment of smooth muscle cells derived from mouse embryonic stem cells

Objective To obtain a pure population of smooth muscle cells (SMC) derived from mouse embryonic stem cells (ESC) and further assess their functions. Methods A vector, expressing both puromycin resistance gene (puror) and enhanced green fluorescent protein (EGFP) gene driven by smooth muscle 22α (SM22α) promoter, named pSM22α-puror-IRES2-EGFP was constructed and used to transfect ESC. Transgenic ESC (Tg-ESC) clones were selected by G418 and identified by PCR amplification of puror gene. The characteristics of Tg-ESC were detected by alkaline phosphatase (ALP) staining, SSEA-1 immunofluorescence and teratoma formation test in vivo. After induction of SMC differentiation by all-trans retinoic acid, differentiated Tg-ESC were treated with 10 µg/mL puromycin for three days to obtain purified SMC (P-SMC). Percentage of EGFP+ cells in P-SMC was assessed by flow cytometer. Expressions of smooth muscle specific markers were detected by immunostaining and Western blotting. Proliferation, migration and contractility of P-SMC were analyzed by growth curve, trans-well migration assay, and carbachol treatment, respectively. Finally, both P-SMC and unpurified SMC (unP-SMC) were injected into syngeneic mouse to see teratoma development. Results Tg-ESC clone was successfully established and confirmed by PCR detection of puror gene in its genomic DNA. The Tg-ESC was positive for ALP staining, SSEA-1 staining and formed teratoma containing tissues derived from three germ layers. After retinoic acid induction, large amount of EGFP positive cells outgrew from differentiated Tg-ESC. Three days of puromycin treatment produced a population of P-SMC with an EGFP+ percentage as high as 98.2% in contrast to 29.47% of unP-SMC. Compared with primary mouse vascular smooth muscle cells (VSMC), P-SMC displayed positive, but lowered expression of SMC-specific markers including SM α-actin and myosin heavy chain (SM-MHC) detected either, by immunostaining, or immunoblotting, accelerated proliferation, improved migration (99.33 ± 2.04 vs. 44.00 ± 2.08 migrated cells/field, P < 0.05), and decreased contractility in response to carbachol (7.75 ± 1.19 % vs. 16.50 ± 3.76 % in cell area reduction, P < 0.05). In vivo injection of unP-SMC developed apparent teratoma while P-SMC did not. Conclusions We obtained a pure population of ESC derived SMC with less mature (differentiated) phenotypes, which will be of great use in research of vascular diseases and in bio-engineered vascular grafts for regenerative medicine.

The efficacy and safety of pharmacoinvasive therapy with prourokinase for acute ST-segment elevation myocardial infarction patients with expected long percutaneous coronary intervention-related delay.

OBJECTIVESnTo elucidate the efficacy and safety of pharmacoinvasive therapy by using prourokinase (prouk) in patients with ST-segment elevation myocardial infarction (STEMI).nnnBACKGROUNDnPatients with STEMI often have long percutaneous coronary intervention (PCI)-related delays due to various reasons, which are associated with poor outcomes.nnnMETHODSnA randomized study which enrolled patients from four centers in China was conducted. Patients were randomly assigned to accept routine primary PCI or prouk-PCI. The primary end points were the angiographic parameters, including thrombolysis in myocardial infarction (TIMI) flow grade, TIMI frame count, and myocardial blush grade. Secondary endpoints were incidence of major adverse cardiac events (MACE, defined as death from all causes, reinfarction, revascularization, or rehospitalization due to new or worsening congestive heart failure) at 30 days and 1 year.nnnRESULTSnOne hundred and ninety-seven eligible patients were enrolled, of whom 100 were randomized to the prouk-PCI group. Significantly more patients in the prouk-PCI group than in the PCI group had an opened infarct-related artery on arrival in the catheterization laboratory (48% vs. 21%, P = 0.0002) and better TIMI frame count after PCI (33 ± 6 vs. 40 ± 10, P < 0.001). At 1-year follow-up, there was a trend that patients in the prouk-PCI group had less chances to have MACE (7.0% vs. 12.6%, P = 0.235) or be readmitted to hospital due to new or worsening congestive heart failure (1.0% vs. 4.1%, P = 0.209).nnnCONCLUSIONnA strategy of emergent PCI preceded by fibrinolysis with prouk results in a better myocardial perfusion in infarct-related artery compared with primary PCI alone in patients with STEMI and long PCI-related delay.

Translational research on novel drug-eluting stents in percutaneous coronary intervention

Although first-generation drug-eluting stents (DES) have markedly reduced restenosis, complications of late and very late in-stent thrombosis have emerged as prime limitations to this technology. The development of new DES is a key process to prevent these complications. Translational research plays a very important role in experiments which determine the safety and efficacy of DES before human clinical trials. The present review focuses on translational research of novel DES, including drug discovery, creation of preclinical research models, planning and conducting of first-in-man studies, and developing next-generation DES systems.

Short-term rosuvastatin treatment for the prevention of contrast-induced acute kidney injury in patients receiving moderate or high volumes of contrast media: a sub-analysis of the TRACK-D study.

Background:Current randomized trials have demonstrated the effects of short-term rosuvastatin therapy in preventing contrast-induced acute kidney injury (CIAKI). However, the consistency of these effects on patients administered different volumes of contrast media is unknown. Methods:In the TRACK-D trial, 2998 patients with type 2 diabetes and concomitant chronic kidney disease (CKD) who underwent coronary/peripheral arterial angiography with or without percutaneous intervention were randomized to short-term (2 days before and 3 days after procedure) rosuvastatin therapy or standard-of-care. This prespecified analysis compared the effects of rosuvastatin versus standard therapy in patients exposed to (moderate contrast volume [MCV], 200–300 ml, n = 712) or (high contrast volume [HCV], ≥300 ml, n = 220). The primary outcome was the incidence of CIAKI. The secondary outcome was a composite of death, dialysis/hemofiltration or worsened heart failure at 30 days. Results:Rosuvastatin treatment was associated with a significant reduction in CIAKI compared with the controls (2.1% vs. 4.4%, P = 0.050) in the overall cohort and in patients with MCV (1.7% vs. 4.5%, P = 0.029), whereas no benefit was observed in patients with HCV (3.4% vs. 3.9%, P = 0.834). The incidence of secondary outcomes was significantly lower in the rosuvastatin group compared with control group (2.7% vs. 5.3%, P = 0.049) in the overall cohort, but it was similar between the patients with MCV (2.0% vs. 4.2%, P = 0.081) or HCV (5.1% vs. 8.8%, P = 0.273). Conclusions:Periprocedural short-term rosuvastatin treatment is effective in reducing CIAKI and adverse clinical events for patients with diabetes and CKD after their exposure to a moderate volume of contrast medium.