Shaoyu Yan

Shear Stress Induces Endothelial Differentiation From a Murine Embryonic Mesenchymal Progenitor Cell Line

Objective—Recent studies have illustrated that mesenchymal stem cells possess the potential to differentiate along an endothelial lineage, but the effect of shear on mesenchymal differentiation is unknown. Thus, we developed an in vitro shear stress system to examine the relationship between shear stress and the endothelial differentiation of a murine embryonic mesenchymal progenitor cell line, C3H/10T1/2. Methods and Results—The parallel plate system of fluid shear stress was used. Shear stress significantly induced expression of mature endothelial cell–specific markers in CH3H/10T1/2 cells such as CD31, von Willebrand factor, and vascular endothelial–cadherin at both the mRNA and protein levels with real-time polymerase chain reaction and immunofluorescence analyses, respectively. In addition, shear-induced augmentation of functional markers of the mature endothelial phenotype such as uptake of acetylated low-density lipoproteins and formation of capillary-like structures on Matrigel. Furthermore, shear stress significantly upregulated angiogenic growth factors while downregulating growth factors associated with smooth muscle cell differentiation. Conclusions—This study demonstrates, for the fist time, endothelial differentiation in a mesenchymal progenitor CH3H/10T1/2 cell line resulting from shear exposure. Thus, this analysis may serve as a basis for further understanding the effect of shear on mesenchymal and vascular cell differentiation.

Human Stanniocalcin-1 Blocks TNF-α–Induced Monolayer Permeability in Human Coronary Artery Endothelial Cells

Objective—Our previous studies revealed upregulation of stanniocalcin-1 (STC1) in cardiac vessels in dilated cardiomyopathy. However, the functional significance of STC1 is unknown. The objective of this study was to determine the effects of STC1 on TNF-&agr;–induced monolayer permeability of human coronary artery endothelial cells (HCAECs). Methods and Results—Cells were pretreated with STC1 for 30 minutes followed by treatment with TNF-&agr; (2 ng/mL) for 24 hours. Monolayer permeability was studied using a transwell system. STC1 pretreatment significantly blocked TNF-&agr;–induced monolayer permeability in a concentration- and time-dependent manner. STC1 effectively blocked TNF-&agr;–induced downregulation of endothelial tight junction proteins zonula occluden-1 and claudin-1 at both mRNA and protein levels. STC1 also significantly decreased TNF-&agr;–induced superoxide anion production. The inhibitory effect of STC1 was specific to TNF-&agr;, as it failed to inhibit VEGF-induced endothelial permeability. Furthermore, STC1 partially blocked NF-&kgr;B and JNK activation in TNF-&agr;–treated endothelial cells. JNK inhibitor and antioxidant also effectively blocked TNF-&agr;–induced NF-&kgr;B activation and monolayer permeability in HCAECs. Conclusions—STC1 maintains endothelial permeability in TNF-&agr;–treated HCAECs through preservation of tight junction protein expression, suppression of superoxide anion production, and inhibition of the activation of NF&kgr;B and JNK, suggesting an important role for STC1 in regulating endothelial functions during cardiovascular inflammation.

Effects of 5 HIV protease inhibitors on vasomotor function and superoxide anion production in porcine coronary arteries

HIV protease inhibitors (PIs) have been implicated to cause cardiovascular complications. Previous studies demonstrated that the PI ritonavir (RTV) caused endothelial dysfunction in porcine arteries. This study investigated and compared the effects of 5 commonly used PIs on vasomotor function, endothelial nitric oxide synthase (eNOS) expression, and oxidative stress in porcine coronary arteries. Vessel rings were incubated with 15 μM of RTV, amprenavir (APV), saquinavir (SQV), indinavir (IDV), or nelfinavir (NFV) for 24 hours. Vasomotor function was studied using a myograph system. The contractility of the rings was significantly reduced for RTV and SQV. In response to bradykinin at 10−5 M, the endothelium-dependent relaxation was significantly reduced for RTV, APV, and SQV. The eNOS mRNA levels were significantly reduced for RTV, APV, and SQV. Furthermore, the superoxide anion (O2−) levels of the vessels were significantly increased for RTV and APV. It was found that nitric oxide production was decreased, whereas the level of nitrotyrosine proteins was increased in RTV-treated vessels. Furthermore, antioxidant seleno-L-methionine (SeMet) reversed RTV-induced O2− production and vasomotor dysfunction. Thus, the HIV PIs RTV, APV, and SQV at 15 μM have more potent in vitro effects on vasomotor dysfunction, eNOS downregulation, and O2− production than IDV and NFV. The antioxidant SeMet can block these adverse effects of RTV. The results suggest that antioxidant therapy may have applications for controlling PI-associated cardiovascular complications.

Resistin Increases Monolayer Permeability of Human Coronary Artery Endothelial Cells

Resistin has been linked to obesity, insulin resistance, atherosclerosis, and the development of cardiovascular disease. Nevertheless, the effects and the molecular mechanisms of resistin on endothelial permeability, a key event in the development of atherosclerosis, inflammation, and vascular disease, are largely unknown. In order to determine the effect of resistin on endothelial permeability, human coronary artery endothelial cells (HCAECs) were treated with clinically relevant concentrations of resistin and the endothelial permeability was measured using the Transwell system with a Texas-Red-labeled dextran tracer. The permeability of HCAEC monolayers treated with resistin (80 ng/mL) was 51% higher than the permeability of control monolayers (P<0.05). The mRNA levels of tight junction proteins zonula occludens-1 (ZO-1) and occludin in resistin-treated cells were 37% and 42% lower, respectively, than the corresponding levels in untreated cells. The protein levels of these molecules in resistin-treated cells were significantly reduced by 35% and 37%, respectively (P<0.05), as shown by flow cytometry and Western blot analysis. Superoxide dismutase (SOD) mimetic MnTBAP effectively blocked the resistin-mediated reduction of ZO-1 and occludin levels in HCAECs. In addition, superoxide anion production was increased from 21% (untreated cells) to 55% (cells treated with 40 ng/mL resistin), and 64% (resistin, 80 mg/mL) (P<0.05). The natural antioxidant Ginkgolide A effectively inhibited resistin-induced increase in permeability and the increase in superoxide anion production in HCAECs. Furthermore, resistin treatment significantly activated p38 MAPK, but not ERK1/2. Pretreatment of HCAECs with a p38 inhibitor effectively blocked resistin-induced permeability. These results provide new evidence that resistin may contribute to the vascular lesion formation via increasing endothelial permeability through the mechanism of oxidative stress and the activation of p38 MAPK.

Ginkgolic acid inhibits HIV protease activity and HIV infection in vitro

Summary Background Several HIV protease mutations, which are resistant to clinical HIV protease inhibitors (PIs), have been identified. There is a great need for second-generation PIs with different chemical structures and/or with an alternative mode of inhibition. Ginkgolic acid is a natural herbal substance and a major component of the lipid fraction in the nutshells of the Ginkgo biloba tree. The objective of this study was to determine whether ginkgolic acid could inhibit HIV protease activity in a cell free system and HIV infection in human cells. Material/Methods Purified ginkgolic acid and recombinant HIV-1 HXB2 KIIA protease were used for the HIV protease activity assay. Human peripheral blood mononuclear cells (PBMCs) were used for HIV infection (HIV-1SF162 virus), determined by a p24gag ELISA. Cytotoxicity was also determined. Results Ginkgolic acid (31.2 μg/ml) inhibited HIV protease activity by 60%, compared with the negative control, and the effect was concentration-dependent. In addition, ginkgolic acid treatment (50 and 100 μg/ml) effectively inhibited the HIV infection at day 7 in a concentration-dependent manner. Ginkgolic acid at a concentration of up to 150 μg/ml demonstrated very limited cytotoxicity. Conclusions Ginkgolic acid effectively inhibits HIV protease activity in a cell free system and HIV infection in PBMCs without significant cytotoxicity. Ginkgolic acid may inhibit HIV protease through different mechanisms than current FDA-approved HIV PI drugs. These properties of ginkgolic acid make it a promising therapy for HIV infection, especially as the clinical problem of viral resistance to HIV PIs continues to grow.

New Roles of a Neuropeptide Cortistatin in the Immune System and Cancer

Cortistatin (CST) is a neuropeptide that strongly resembles somatostatin (SS) structurally and functionally. CST binds to all five SS receptors (SSTR1-SSTR5) with high affinity and exerts its function mainly through SSTRs. Despite many similar functions between these two neuropeptides, they are products of different genes. Recently, some distinct functions and receptor usage of CST have been reported. Some of the interesting functions of CST were not found with SS. Therefore CST could have potential new roles in an ex-neuronal system that regulates immune responses as well as other cellular functions in the body. In this review, we discuss the new functions of CST in the immune system, cancer pathogenesis, and possible CST-specific receptors.