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MicroRNA-199a-5p (miR-199a-5p) and -3p are enriched in the myocardium, but it is unknown whether miR-199a-5p and -3p are co-expressed in cardiac remodeling and what roles they have in cardiac hypertrophy and fibrosis. We show that miR-199a-5p and -3p are co-upregulated in the mouse and human myocardium with cardiac remodeling and in Ang-II-treated neonatal mouse ventricular cardiomyocytes (NMVCs) and cardiac fibroblasts (CFs). miR-199a-5p and -3p could aggravate cardiac hypertrophy and fibrosis in vivo and in vitro. PPAR gamma coactivator 1 alpha (Ppargc1a) and sirtuin 1 (Sirt1) were identified as target genes to mediate miR-199a-5p in promoting both cardiac hypertrophy and fibrosis. However, miR-199a-3p aggravated cardiac hypertrophy and fibrosis through targeting RB transcriptional corepressor 1 (Rb1) and Smad1, respectively. Serum response factor and nuclear factor κB p65 participated in the upregulation of miR-199a-5p and -3p in Ang-II-treated NMVCs and mouse CFs, and could be conversely elevated by miR-199a-5p and -3p. Together, Ppargc1a and Sirt1, Rb1 and Smad1 mediated the pathological effect of miR-199a-5p and -3p by promoting cardiac hypertrophy and fibrosis, respectively. This study suggests a possible new strategy for cardiac remodeling therapy by inhibiting miR-199a-5p and -3p., Graphical abstract, Transcription factor SRF and NF-κB p65 regulate the co-expression of miR-199a-5p and -3p by positive feedback in the pathological cardiac remodeling. Ppargc1a and Sirt1, Rb1 and Smad1 mediate the pathological role of miR-199a-5p and -3p by promoting cardiac hypertrophy and fibrosis, respectively.

Atrial fibrosis induced by aging is one of the main causes of atrial fibrillation (AF), but the potential molecular mechanism is not clear. Acetyltransferase p300 participates in the cellular senescence and fibrosis, which might be involved in the age-related atrial fibrosis. Four microarray datasets generated from atrial tissue of AF patients and sinus rhythm (SR) controls were analyzed to find the possible relationship of p300 (EP300) with senescence and fibrosis. And then, biochemical assays and in vivo electrophysiological examination were performed on older AF patients, aging mice, and senescent atrial fibroblasts. The results showed that (1) the left atrial tissues of older AF patients, aging mouse, and senescence human atrial fibroblasts had more severe atrial fibrosis and higher protein expression levels of p300, p53/acetylated p53 (ac-p53)/p21, Smad3/p-Smads, and fibrosis-related factors. (2) p300 inhibitor curcumin and p300 knockdown treated aging mouse and senescence human atrial fibroblasts reduced the senescence ratio of atrial fibroblasts, ameliorated the atrial fibrosis, and decreased the AF inducibility. In contrast, over-expression of p300 can lead to the senescence of atrial fibroblasts and atrial fibrosis. (3) p53 knockdown decreased the expression of aging and fibrosis-related proteins. (4) Co-immunoprecipitation and immunofluorescence showed that p53 forms a complex with smad3 and directly regulates the expression of smad3 in atrial fibroblasts. Our findings suggest that the mechanism of atrial fibrosis induced by aging is, at least, partially dependent on the regulation of p300, which provides new sights into the AF treatment, especially for the elderly.

Vascular dysfunction is a common pathological basis for complications in individuals affected by diabetes. Previous studies have established that endothelial dysfunction is the primary contributor to vascular complications in type 2 diabetes (T2DM). However, the role of vascular smooth muscle cells (VSMCs) in vascular complications associated with T2DM is still not completely understood. The aim of this study is to explore the potential mechanisms associated with Ca2+ handling dysfunction and how this dysfunction contributes to diabetic vascular smooth muscle impairment. The results indicated that endothelium-dependent vasodilation was impaired in diabetic aortae, but endothelium-independent vasodilation was not altered. Various vasoconstrictors such as phenylephrine, U46619 and 5-HT could induce vasoconstriction in a concentration-dependent manner, such that the dose-response curve was parallel shifted to the right in diabetic aortae, compared to the control. Vasoconstrictions mediated by L-type calcium (Cav1.2) channels were attenuated in diabetic aortae, but effects mediated by store-operated calcium (SOC) channels were enhanced. Intracellular Ca2+ concentration ([Ca2+]i) in VSMCs was detected by Fluo-4 calcium fluorescent probes, and demonstrated that SOC-mediated Ca2+ entry was increased in diabetic VSMCs. VSMC-specific knockout of STIM1 genes decreased SOC-mediated and phenylephrine-induced vasoconstrictive response in mice aortae. Additionally, Orai1 expression was up-regulated, Cav1.2 expression was downregulated, and the phenotypic transformation of diabetic VSMCs was determined in diabetic aortae. The overexpression of Orai1 markedly promoted the OPN expression of VSMCs, whereas SKF96365 (SOC channel blocker) reversed the phenotypic transformation of diabetic VSMCs. Our results demonstrated that the vasoconstriction response of aortic smooth muscle was weakened in type 2 diabetic rats, which was related to the downregulation of the Cav1.2 channel and the up-regulation of the SOC channel signaling pathway.

We collected remote sensing images of lower reaches of the Shiyang River Basin in 1995, 2000, 2005, 2010, 2015, and 2018. We analyzed the dynamics of desertification in Minqin County by constructing desertification difference index, using RS and GIS theories and methods. We established the Albedo-NDVI feature space based on the normalized vegetation index (NDVI) and surface albedo (Albedo). The results showed that the desertification area in the lower reaches of Shi-yang River Basin accounted for more than 90% of the total area of the Minqin. Oasis scattered in large areas of desertified land. Moderate desertification accounted for more than 70% of the total area of the county. The area of desertification in Minqin showed an decreasing trend from 1995 to 2018, with an average annual reduction area of 22.06 km以石羊河流域下游甘肃省民勤县为研究区,选取1995、2000、2005、2010、2015、2018年6期遥感影像作为数据源,利用RS和GIS理论和方法,基于归一化植被指数(NDVI)和地表反照率(Albedo)建立Albedo-NDVI特征空间,构建荒漠化差值指数,对民勤县荒漠化动态变化进行分析。结果表明: 石羊河流域下游地区荒漠化面积占民勤县总面积的90%以上,绿洲零星分布在大面积荒漠化土地中,以中度荒漠化为主,占全县总面积的70%以上;1995—2018年间,民勤县荒漠化面积整体呈减少趋势,年均减少面积为22.06 km

Background and Aim. The senescence of human bone marrow mesenchymal stem cells (hBMSCs) can be induced by oxidative stress, but the mechanism by which it occurs is not yet clear. Here, we performed an investigation on the pathogenesis of hypoxia-induced senescence through proteomic analyses and aimed to explore the mechanisms of stem cell senescence. Methods. Hypoxia in hBMSCs was induced for 0, 4, and 12 hours, and cellular senescence was evaluated by senescence-associated β-galactosidase (SA-β-gal) staining. Tandem mass tag (TMT) labeling was combined with liquid chromatography-tandem mass spectrometry (LC-MS/MS) for differential proteomic analysis of hypoxia in hBMSCs. Parallel reaction monitoring (PRM) analysis was used to validate the candidate proteins. Verifications of signaling pathways were evaluated by western blotting. Cell apoptosis was evaluated using Annexin V/7-AAD staining by flow cytometry. The production of reactive oxygen species (ROS) was detected by the fluorescent probe 2,7-dichlorodihydrofluorescein diacetate (DCFH-DA). Results. Cell senescence detected by SA-β-gal activity was higher in the 12-hour hypoxia-induced group. TMT analysis of 12-hour hypoxia-induced cells identified over 6000 proteins, including 686 differentially expressed proteins. Based on biological pathway analysis, we found that the senescence-associated proteins were predominantly enriched in the cancer pathways, PI3K-Akt pathway, and cellular senescence signaling pathways. CDK1, CDK2, and CCND1 were important nodes in PPI analyses. Moreover, the CCND1, UQCRH, and COX7C expressions were verified by PRM. Hypoxia induction for 12 hours in hBMSCs reduced CCND1 expression but promoted ROS production and cell apoptosis. Such effects were markedly reduced by the PI3K agonist, 740 Y-P, and attenuated by LY294002. Conclusions. Hypoxia of hBMSCs inhibited CCND1 expression but promoted ROS production and cell apoptosis through activating the PI3K-dependent signaling pathway. These findings provided a detailed characterization of the proteomic profiles related to hypoxia-induced senescence of hBMSCs and facilitated our understanding of the molecular mechanisms leading to stem cell senescence.

Background: Ca2+ plays an important role in the regulation of vasoconstriction. Ca2+ signaling is regulated by a number of Ca2+-handling proteins. However, whether differences in Ca2+ handling affect the regulation of vasoconstriction in different arteries remains elusive. Objective: To determine whether differences in Ca2+ handling affect the response to vasoconstrictors in different arteries. Methods: Arterial ring contraction was measured using a Multi Myograph System. Vascular smooth muscle cells (VSMCs) were digested with type 2 collagenase in DMEM, then intracellular calcium concentration was measured with the Ca2+ probe fluo-4/AM in the isolated cells. Calcium-related proteins were assayed by Western blotting. Results: Phenylephrine did not induce ­coronary arterial contraction. There were differences in ­5-hydroxytryptamine, 9,11-dideoxy-11a,9a-epoxymethano-prostaglandin F2a, and endothelin 1-induced vasoconstriction in different solutions between coronary and renal arteries. Vasoconstrictions in the presence of Bay K8644 were stronger in coronary than in renal arteries. Store-operated calcium (SOC) channels could mediate Ca2+ influx in VSMCs of both groups. SOC channels did not participate in the contraction of coronary arteries. In addition, there were significant differences in the expressions of receptors and ion channels between the two groups. Conclusions: Ca2+ handling contributed to the different responses to vasoconstrictors between coronary and renal arteries.

Hypertension is a main risk factor for atrial fibrillation, but the direct effects of hydrostatic pressure on the atrial fibrosis are still unknown. The present study investigated whether hydrostatic pressure is responsible for atrial fibrosis, and addressed a potential role of the Smad pathway in this pathology. Biochemical assays were used to study regulation and expression of fibrotic factors in spontaneously hypertensive rats (SHRs) and Wistar rats, and in cardiac fibroblasts (CFs) cultured under standard (0 mmHg) and elevated (20, 40 mmHg) hydrostatic pressure. Levels of atrial fibrosis and protein expression of fibrotic factors Col-1A1/-3A1, TGF-β1, and MMP-2 in SHRs' left atrial tissues were higher than those in Wistar rats. Exposure to elevated pressure was associated with the proliferation of CFs. The protein expression of Col-1A1/-3A1, TGF-β1, and MMP-2 in CFs was also up-regulated in a pressure-dependent manner. The proliferation of CFs and increased expressions of fibrotic markers induced by elevated hydrostatic pressure could be reversed by the Smad3 inhibitor naringenin. The activation of Smad3 pathway was also stimulated by elevated hydrostatic pressure. These results demonstrate that CF secretory function and proliferation can be up-regulated by exposure to elevated pressure, and that Smad3 may modulate CF activation induced by high hydrostatic pressure.

Thromboxane A2 (TXA2) has been implicated in the pathogenesis of diabetic vascular complications, although the underlying mechanism remains unclear. The present study investigated the alterations in TXA2 receptor signal transduction in type 2 diabetic renal arteries. The contraction of renal arterial rings in control (db/m+) mice and type 2 diabetic (db/db) mice was measured by a Multi Myograph System. Intracellular calcium concentration ([Ca2+]i) in vascular smooth muscle cells was measured by Fluo-4/AM dye and confocal laser scanning microscopy. Quantitative real-time PCR and Western blot analysis were used to determine gene and protein expression levels, respectively. A stable TXA2 mimic U46619 caused markedly stronger dose-dependent contractions in the renal arteries of db/db mice than in those of db/m+ mice. This response was completely blocked by a TXA2 receptor antagonist GR32191 and significantly inhibited by U73122. U46619-induced vasoconstriction was increased in the presence of nifedipine in db/db mice compared with that in db/m+ mice, whereas the response to U46619 did not differ between the two groups in the presence of SKF96365. Sarcoplasmic reticulum Ca2+ release-mediated and CaCl2-induced contractions did not differ between the two groups. In db/db mice, store-operated Ca2+(SOC) entry-mediated contraction in the renal arteries and SOC entry-mediated Ca2+ influx in smooth muscle cells were significantly increased. And the gene and protein expressions of TXA2 receptors, Orai1 and Stim1 were upregulated in the diabetic renal arteries. Therefore the enhancement of U46619-induced contraction was mediated by the upregulation of TXA2 receptors and downstream signaling in the diabetic renal arteries.

Vascular dysfunction is a common pathological basis for complications in individuals affected by diabetes. Previous studies have established that endothelial dysfunction is the primary contributor to vascular complications in type 2 diabetes (T2DM). However, the role of vascular smooth muscle cells (VSMCs) in vascular complications associated with T2DM is still not completely understood. The aim of this study is to explore the potential mechanisms associated with Ca

Aims Hypertension is an independent risk factor for atrial fibrillation (AF). However, the direct effect of hydrostatic pressure on atrial electrical remodeling is unclear. The present study investigated whether hydrostatic pressure is responsible for atrial electrical remodeling and addressed a potential role of inflammation in this pathology. Main methods Whole-cell patch-clamp recordings and biochemical assays were used to study the regulation and expression of ion channels in left atrial appendages in patients with AF, spontaneously hypertensive rats (SHRs), and atrium-derived cells (HL-1 cells) exposed to standard (0 mmHg) and elevated (20, 40 mmHg) hydrostatic pressure. Key findings Both TNF-α and MIF were highly expressed in patients with AF and SHRs. AF inducibility in SHRs was higher after atrial burst pacing, accompanied by a decrease in the L-type calcium current (ICa,L), an increase in the transient outward K+ current (Ito) and ultra-rapid delayed rectifier K+ current (IKur), and a shortened action potential duration (APD), which could be inhibited by atorvastatin. Furthermore, exposure to elevated pressure was associated with electrical remodeling of the HL-1 cells. The peak current density of ICa,L was reduced, while Ito and IKur were increased. Moreover, the expression levels of Kv4.3, Kv1.5, TNF-α, and MIF were upregulated, while the expression of Cav1.2 was downregulated in HL-1 cells after treatment with high hydrostatic pressure (40 mmHg). Atorvastatin alleviated the electrical remodeling and increased inflammatory markers in HL-1 cells induced by high hydrostatic pressure. Significance Elevated hydrostatic pressure led to atrial electrical remodeling and increased AF susceptibility by upregulating inflammation.

Hypertension is an independent risk factor for atrial fibrillation (AF), although its specific mechanisms remain unclear. Previous research has been focused on cyclic stretch, ignoring the role of high hydrostatic pressure. The present study aimed to explore the effect of high hydrostatic pressure stimulation on electrical remodeling in atrial myocytes and its potential signaling pathways. Experiments were performed on left atrial appendages from patients with chronic AF or sinus rhythm, spontaneously hypertensive rats (SHRs) treated with or without valsartan (10 mg/kg/day) and HL-1 cells were exposed to high hydrostatic pressure using a self-developed device. Whole-cell patch-clamp recordings and western blots demonstrated that the amplitudes of ICa,L, Ito, and IKur were reduced in AF patients with corresponding changes in protein expression. Angiotensin protein levels increased and Ang1–7 decreased, while focal adhesion kinase (FAK) and Src kinase were enhanced in atrial tissue from AF patients and SHRs. After rapid atrial pacing, AF inducibility in SHR was significantly higher, accompanied by a decrease in ICa,L, upregulation of Ito and IKur, and a shortened action potential duration. Angiotensin upregulation and FAK/Src activation in SHR were inhibited by angiotensin type 1 receptor inhibitor valsartan, thus, preventing electrical remodeling and reducing AF susceptibility. These results were verified in HL-1 cells treated with high hydrostatic pressure, and demonstrated that electrical remodeling regulated by the FAK-Src pathway could be modulated by valsartan. The present study indicated that high hydrostatic pressure stimulation increases AF susceptibility by activating the renin-angiotensin system and FAK-Src pathway in atrial myocytes.

Constitutive activation of Hedgehog (Hh) pathway is intimately related with the occurrence and development of several malignancies, such as medulloblastoma (MB) and other tumors. Therefore, small molecular inhibitors of Hh pathway are urgently needed. In this study, three new steroidal alkaloids, ⊿5 (20R, 24R) 23-oxo-24-methylsolacongetidine, ⊿5 (20S, 24R) 23-oxo-24-methylsolacongetidine and veralinine 3-O-α- l -rhamnopyranosyl-(1 → 2)-β-D-glucopyranoside, together with six known alkaloids, 20-epi-verazine, verazine, protoverine 15-(l)-2′-methylbutyrate, jervine, veramarine and β1-chaconine, were isolated and determined from Veratrum grandiflorum Loes. The dual-luciferase bioassay indicated that all compounds exhibited significant inhibitions of Hh pathway with IC50 values of 0.72–14.31 μM against Shh-LIGHT 2 cells. To determine whether these Hh pathway inhibitors act with the Smoothened (Smo) protein, which is an important oncoprotein and target for this pathway, BODIPY-cyclopamine (BC) competitive binding assay was preferentially performed. Compared with BC alone, all compounds obviously reduced the fluorescence intensities of BC binding with Smo in Smo-overexpression HEK293T cells through fluorescence microscope and flow cytometer. By directly interacting with Smo, it revealed that they were actually novel natural Smo inhibitors. Then, their anti-tumor effects were investigated against the human MB cell line DAOY, which is a typical pediatric brain tumor cells line with highly expressed Hh pathway. Interestingly, most of compounds had slight proliferation inhibitions on DAOY cells after treatment for 24 h same as vismodegib, while β1-chaconine showed the strongest inhibitory effect on the growth of DAOY with IC50 value of 5.35 μM. In conclusion, our studies valuably provide several novel natural Smo inhibitors for potential targeting treatment of Hh-dependent tumors.

Aim: To investigate whether plasma miRNAs targeting CYP3A4/5 have an impact on the variance of pharmacokinetics of clopidogrel. Materials & methods: The contribution of 13 miRNAs to the CYP3A4/5 gene expression and activity was investigated in 55 liver tissues. The association between plasma miRNAs targeting CYP3A4/5 mRNA and clopidogrel pharmacokinetics was analyzed in 31 patients with coronary heart disease who received 300 mg loading dose of clopidogrel. Results: Among 13 miRNAs, miR-142 was accounting for 12.2% (p = 0.002) CYP3A4 mRNA variance and 9.4% (p = 0.005) CYP3A5 mRNA variance, respectively. Plasma miR-142 was negatively associated with H4 Cmax (r = -0.5269; p = 0.0040) and associated with H4 AUC0–4h (r = -0.4986; p = 0.0069) after 300 mg loading dose of clopidogrel in coronary heart disease patients. Conclusion: miR-142 could account for a part of missing heritability of CYP3A4/5 functionality related to clopidogrel activation.

Ca2+ plays an important role in the regulation of vasoconstriction. Ca2+ signaling is regulated by a number of Ca2+-handling proteins. However, whether differences in Ca2+ handling affect the regulation of vasoconstriction in different arteries remains elusive.To determine whether differences in Ca2+ handling affect the response to vasoconstrictors in different arteries.Arterial ring contraction was measured using a Multi Myograph System. Vascular smooth muscle cells (VSMCs) were digested with type 2 collagenase in DMEM, then intracellular calcium concentration was measured with the Ca2+ probe fluo-4/AM in the isolated cells. Calcium-related proteins were assayed by Western blotting.Phenylephrine did not induce -coronary arterial contraction. There were differences in -5-hydroxytryptamine, 9,11-dideoxy-11a,9a-epoxymethano-prostaglandin F2a, and endothelin 1-induced vasoconstriction in different solutions between coronary and renal arteries. Vasoconstrictions in the presence of Bay K8644 were stronger in coronary than in renal arteries. Store-operated calcium (SOC) channels could mediate Ca2+ influx in VSMCs of both groups. SOC channels did not participate in the contraction of coronary arteries. In addition, there were significant differences in the expressions of receptors and ion channels between the two groups.Ca2+ handling contributed to the different responses to vasoconstrictors between coronary and renal arteries.

Cyclins/retinoblastoma protein (pRb) pathway participates in cardiomyocyte hypertrophy. MicroRNAs (miRNAs), the endogenous small non-coding RNAs, were recognized to play significant roles in cardiac hypertrophy. But, it remains unknown whether cyclin/Rb pathway is modulated by miRNAs during cardiac hypertrophy. This study investigates the potential role of microRNA-16 (miR-16) in modulating cyclin/Rb pathway during cardiomyocyte hypertrophy. An animal model of hypertrophy was established in a rat with abdominal aortic constriction (AAC), and in a mouse with transverse aortic constriction (TAC) and in a mouse with subcutaneous injection of phenylephrine (PE) respectively. In addition, a cell model of hypertrophy was also achieved based on PE-promoted neonatal rat ventricular cardiomyocyte and based on Ang-II-induced neonatal mouse ventricular cardiomyocyte respectively. We demonstrated that miR-16 expression was markedly decreased in hypertrophic myocardium and hypertrophic cardiomyocytes in rats and mice. Overexpression of miR-16 suppressed rat cardiac hypertrophy and hypertrophic phenotype of cultured cardiomyocytes, and inhibition of miR-16 induced a hypertrophic phenotype in cardiomyocytes. Expressions of cyclins D1, D2 and E1, and the phosphorylated pRb were increased in hypertrophic myocardium and hypertrophic cardiomyocytes, but could be reversed by enforced expression of miR-16. Cyclins D1, D2 and E1, not pRb, were further validated to be modulated post-transcriptionally by miR-16. In addition, the signal transducer and activator of transcription-3 and c-Myc were activated during myocardial hypertrophy, and inhibitions of them prevented miR-16 attenuation. Therefore, attenuation of miR-16 provoke cardiomyocyte hypertrophy via derepressing the cyclins D1, D2 and E1, and activating cyclin/Rb pathway, revealing that miR-16 might be a target to manage cardiac hypertrophy.

The relationship between either paraoxonase 1 (PON1) gene promoter DNA methylation or genetic variations and bleeding or major adverse cardiac events after dual antiplatelet therapy has been incompletely characterized. We aimed to systematically investigate the role of genetic variations and DNA methylation of the PON1 CpG island promoter on the clinical outcomes of dual antiplatelet therapy for patients with coronary artery disease (CAD) who underwent percutaneous coronary intervention (PCI). This study included 653 patients with CAD undergoing PCI and receiving dual antiplatelet therapy. Genomic DNAs were isolated from whole blood and were genotyped for the three single nucleotide polymorphisms (SNPs) of the PON1 gene. The DNA methylation levels in the PON1 promoter region were determined by bisulfite sequencing or pyrosequencing at five CpG sites (positions −142, −161, −163, −170, and −184 from the transcription start site). Clopidogrel and its metabolites in plasma were examined using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS), and platelet function analysis was performed using the VerifyNow assay. Statistically significant associations between methylation levels at five PON1 CpG sites and bleeding were observed: −184 [odds ratio (OR) 0.98, 95% confidence interval (CI) 0.96–1.00, p = 0.028]; −170 (OR 0.99, 95% CI 0.97–1.00, p = 0.048); −163 (OR 0.98, 95% CI 0.96–1.00, p = 0.029); −161 (OR 0.98, 95% CI 0.97–1.00, p = 0.026); and −142 (OR 0.98, 95% CI 0.97–1.00, p = 0.042) at a false discovery rate of

Thromboxane A

Background In the early stage of diabetes, the cardiac ejection fraction is preserved, despite the existence of the subclinical cardiac dysfunction to some extent. However, the detailed phenotype of this dysfunction and the underlying mechanism remain unclear. To improve our understanding of this issue, we used low-dose STZ and high-fat diet to induce type 2 diabetic models in rats. The effects and the mechanism associated with the early stages of the disease were analyzed. Methods The type 2 diabetic mellitus (T2DM) in SD rats were induced through 30 mg/kg STZ and high-fat diet. Two-dimensional spackle-tracking echocardiography (STE) and the dobutamine test were performed to examine the cardiac function. Calcium transients of left ventricular myocytes were detected and the related intracellular signalling factors were analyzed by western blotting. Results After 6-weeks, T2DM rats in left ventricular (LV) diastole showed decreased global and segment strain(S) levels (P

Results In the 74 malignant effusion specimens, 57 (77.02%) were positive for seprase, while only 2 (22.22%) of the control group were positively stained (P=0.001). In the malignant effusions, 17 (22.97%), 22 (29.73%), 22 (29.73%), 13 (17.57%) had negative, weak, moderate and strong seprase protein expression, respectively. The expression of seprase protein was predominant in cytoplasm of carcinoma cells. Increased seprase protein was negatively associated with the overall survival rate of the patients (P=0.03). However, there was no significant correlation between protein expression and FIGO stage, age, histology, and histological grade. By Western blotting, 27 of the 34 effusions showed the presence of both 170-kD dimeric form and 97-KD monomeric form of seprase while only 1 of the 34 had 170-KD dimeric form, which was consistent with the results of immunohistochemistry (P＝0.05). Conclusions Seprase may be involved in the development of ovarian cancer, and is a potential predictive marker for the disease. Chin Med J 2007;120(8):663-668

The reprogramming of fibroblasts to induced pluripotent stem cells raises the possibility that somatic cells could be directly reprogrammed to cardiac progenitor cells (CPCs). The present study aimed to assess highly efficient protein-based approaches to reduce or eliminate the genetic manipulations to generate CPCs for cardiac regeneration therapy. A combination of QQ-reagent-modified Gata4, Hand2, Mef2c, and Tbx5 and three cytokines rapidly and efficiently reprogrammed human dermal fibroblasts (HDFs) into CPCs. This reprogramming process enriched trimethylated histone H3 lysine 4, monoacetylated histone H3 lysine 9, and Baf60c at the Nkx2.5 cardiac enhancer region by the chromatin immunoprecipitation quantitative polymerase chain reaction assay. Protein-induced CPCs transplanted into rat hearts after myocardial infarction improved cardiac function, and this was related to differentiation into cardiomyocyte-like cells. These findings demonstrate that the highly efficient protein-transduction method can directly reprogram HDFs into CPCs. This protein reprogramming strategy lays the foundation for future refinements both in vitro and in vivo and might provide a source of CPCs for regenerative approaches. Significance The findings from the present study have demonstrated an efficient protein-transduction method of directly reprogramming fibroblasts into cardiac progenitor cells. These results have great potential in cell-based therapy for cardiovascular diseases.

Cardiac progenitor cells (CPCs) potentially offer a novel approach to cardiovascular regeneration. The reprogramming of fibroblasts to induced pluripotent stem cells (iPSCs) raises the possibility that a somatic cell could be directly reprogrammed to a CPC. However, practical application has been

Cast lightweight composite insulation wall is a new, energy-saving, environmentally friendly wall materials, this article from cast lightweight composite wall construction technology, quality standards and other aspects of the construction of lightweight composite wall cast a more comprehensive exposition, to clarify the key construction technology, materials properties, development advantages and other aspects of cast lightweight composite wall. It has certain significance on future construction of such walls.

Through the foam concrete block cement type, water reducer type, content and tailings using water reducing powder content of the different experimental groups, analyzed the relationship between raw material composition and amount due to compressive strength of foam concrete and different effects of moisture content and compressive strength, and thus presumably the ideal mix. It can be used for future theoretical study reference.

Carvedilol, a nonselective β-adrenoreceptor antagonist, protects against myocardial injury induced by acute myocardium infarction (AMI). The mechanisms underlying the anti-fibrotic effects of carvedilol are unknown. Recent studies have revealed the critical role of microRNAs (miRNAs) in a variety of cardiovascular diseases. This study investigated whether miR-29b is involved in the cardioprotective effect of carvedilol against AMI-induced myocardial fibrosis. Male SD rats were randomized into several groups: the sham surgery control, left anterior descending (LAD) surgery-AMI model, AMI plus low-dose carvedilol treatment (1 mg/kg per day, CAR-L), AMI plus medium-dose carvedilol treatment (5 mg/kg per day, CAR-M) and AMI plus high-dose carvedilol treatment (10 mg/kg per day, CAR-H). Cardiac remodeling and impaired heart function were observed 4 weeks after LAD surgery treatment; the observed cardiac remodeling, decreased ejection fraction, and fractional shortening were rescued in the CAR-M and CAR-H groups. The upregulated expression of Col1a1, Col3a1, and α-SMA mRNA was significantly reduced in the CAR-M and CAR-H groups. Moreover, the downregulated miR-29b was elevated in the CAR-M and CAR-H groups. The in vitro study showed that Col1a1, Col3a1, and α-SMA were downregulated and miR-29b was upregulated by carvedilol in a dose-dependent manner in rat cardiac fibroblasts. Inhibition of ROS-induced Smad3 activation by carvedilol resulted in downregulation of Col1a1, Col3a1, and α-SMA and upregulation of miR-29b derived from the miR-29b-2 precursor. Enforced expression of miR-29b significantly suppressed Col1a1, Col3a1, and α-SMA expression. Taken together, we found that smad3 inactivation and miR-29b upregulation contributed to the cardioprotective activity of carvedilol against AMI-induced myocardial fibrosis.

Aim of this study The panax notoginseng saponins (PNS) have been clinically used for the treatment of cardiovascular diseases and stroke in China. Evidences demonstrated that PNS could protect cardiomyocytes from injury induced by ischemia, but the underlying molecular mechanisms of this protective effect are still unclear. This study was aimed to investigate the protective effect and potential molecular mechanisms of PNS on apoptosis in H9c2 cells in vitro and rat myocardial ischemia injury model in vivo. Materials and methods H9c2 cells subjected to serum, glucose and oxygen deprivation (SGOD) were used as in vitro models and SD rats subjected to left anterior descending (LAD) coronary artery ligation were used as in vivo models. The anti-apoptotic effect of PNS was evaluated by Annexin V/PI analysis or TUNEL assay. Mitochondrial membrane potential (Δψm) was detected by JC-1 analysis. The expression of Akt and phosphorylated Akt (p-Akt) were detected by western blot assay. Results PNS exhibited anti-apoptotic effect both in H9c2 cells and in ischemic myocardial tissues. However, the effect was blocked in vitro by LY294002, a specific PI3K inhibitor. The anti-apoptotic effect of PNS was mediated by stabilizing Δψm in H9c2 cells. Furthermore the indices of the left ventricular ejection fractions (EF), left ventricular fractional shortening (FS), left ventricular dimensions at end diastole (LVDd) and left ventricular dimensions at end systole (LVDs) suggested that PNS improved rats cardiac function. PNS significantly increased p-Akt both in H9c2 cells and in ischemic myocardial tissues and this effect was also blocked by LY294002 in H9c2 cells. Conclusion Results of this study suggested that PNS could protect myocardial cells from apoptosis induced by ischemia in both the in vitro and in vivo models through activating PI3K/Akt signaling pathway.

To investigate the expression of dipeptidyl peptidase IV (DPPIV) in patients with epithelial ovarian carcinoma (EOC) and its clinical significance.Immunohistochemistry (IHC) was used to detect the expression of DPPIV protein in 378 formalin-fixed paraffin-embedded EOC tissue samples. The expression of DPPIV mRNA in 86 EOC tissue samples were examined by in situ hybridization (ISH) using specific FITC-labelled RNA probes. Forty-two samples of normal ovarian tissues were used as control. Statistical analyses were carried out by Chi-square test, Spearman rank correlation and Kaplan-Meier method.Among the 378 epithelial ovarian carcinomas, 351 (92.9%) showed a positive expression of DPPIV protein, while only 25/42 (59.5%) of normal ovaries had a positive expression by semi-quantitative IHC analysis. The expression level of DPPIV protein was significantly lower in the normal ovaries than that in ovarian carcinomas (chi(2) = 18.4, P = 0.001). There was no significant correlation between the expression of DPPIV protein and age, FIGO stage and histological grade (P0.05). However, the expression of DPPIV protein was significantly associated with histological type (chi(2) = 28.5, P = 0.005). The patients with high level expression of DPPIV protein likely had a poor prognosis in terms of overall survival (P = 0.02). Of the 86 patients, 84 (97.7%)showed positive expression of DPPIV mRNA, also higher than that in normal ovarian tissues (P0.05). A statistically significant correlation between DPPIV mRNA and protein expression was observed (r(s) = 0.66, P = 0.001).DPPIV may be involved in the carcinogenesis of ovarian cancer, and may become a potential prognostic marker for epithelial ovarian carcinoma.

To construct the recombinant eukaryotic expression vector pRNAT-U6.1-siEdg4 which carries small interfering RNA (siRNA) of Edg4 and observe the silencing effect of Edg4 gene targeted siRNA in ovarian cancer cell line SKOV3.The Edg4 gene-targeted hairpin siRNA sequence was designed according to the Edg4 sequence in Genbank, and the two complementary oligo nucleotide strands were synthesized and annealed and inserted into the pRNAT-U6.1 plasmid to build a recombinant Edg4 siRNA eukaryotic expression vector, which was sequenced and identified to contain the correct Edg4 siRNA sequence. The human ovarian carcinoma cell lines SKOV3 were transfected with the vector using lipofectamine method. The efficiency of transfecting cells was observed with fluorescent microscope and the mRNA expression level of Edg4 gene was detected by real time quantitative PCR. The LPA levels in cell supernatants were detected using a biochemical method. And the apoptosis of SKOV3 cells induced by the vector was evaluated by flow cytometry.The recombinant eukaryotic expression vector was confirmed to contain correct Edg4 siRNA sequence by PCR and sequencing. After transfection large amounts of green fluorescence were seen in plasma and nuclei of SKOV3 cells and the positive cell rates were 64%. The expression level of Edg4 mRNA in transfected SKOV3 cell line was significantly decreased (0.05 +/- 0.01vs 0.29 +/- 0.04, P0.05). The decrease in LPA level in the cell supernatants was revealed [(3.0 +/- 1.0) vs (7.5 +/- 2.2)micromol/L, P0.05]. The apoptosis rate of transfected SKOV3 was increased obviously (53.38% vs 0.51%, P0.05).We have successfully constructed the recombinant eukaryotic expression vector containing Edg4 gene targeted siRNA (pRNAT-U6.1-siEdg4). The vector could effectively transfect SKOV3 cell line, and obviously suppress the Edg4 mRNA expression and induce cell apoptosis in ovarian cancer cell line SKOV3.

Seprase plays an important role in malignant cell invasion and metastasis by degrading the extracellular matrix. However, its clinical significance remains largely unknown. The objective of the current study was to evaluate the expression of seprase in effusions from patients with epithelial ovarian carcinoma and its clinical values.Immunohistochemistry was used to examine the expression of seprase protein in a series of 74 malignant peritoneal (n = 64) and pleural (n = 10) effusions from Norwegian patients with epithelial ovarian carcinoma. Additionally, 34 effusions were evaluated using the Western blotting. Nine reactive effusions, obtained from patients with benign lesions, served as a control group. Statistical analyses were carried out by Chi-square test and Kaplan-Meier method.In the 74 malignant effusion specimens, 57 (77.02%) were positive for seprase, while only 2 (22.22%) of the control group were positively stained (P = 0.001). In the malignant effusions, 17 (22.97%), 22 (29.73%), 22 (29.73%), 13 (17.57%) had negative, weak, moderate and strong seprase protein expression, respectively. The expression of seprase protein was predominant in cytoplasm of carcinoma cells. Increased seprase protein was negatively associated with the overall survival rate of the patients (P = 0.03). However, there was no significant correlation between protein expression and FIGO stage, age, histology, and histological grade. By Western blotting, 27 of the 34 effusions showed the presence of both 170-kD dimeric form and 97-KD monomeric form of seprase while only 1 of the 34 had 170-KD dimeric form, which was consistent with the results of immunohistochemistry (P = 0.05).Seprase may be involved in the development of ovarian cancer, and is a potential predictive marker for the disease.

Among the many proteases associated with human cancer, seprase or fibroblast activation protein α, a type II transmembrane glycoprotein, has two types of EDTA-resistant protease activities: dipeptidyl peptidase and a 170-kDa gelatinase activity. To test if activation of gelatinases associated with seprase could be involved in malignant tumors, we used a mammalian expression system to generate a soluble recombinant seprase (r-seprase). In the presence of putative EDTA-sensitive activators, r-seprase was converted into 70- to 50-kDa shortened forms of seprase (s-seprase), which exhibited a 7-fold increase in gelatinase activity, whereas levels of dipeptidyl peptidase activity remained unchanged. In malignant human tumors, seprase is expressed predominantly in tumor cells as shown by in situ hybridization and immunohistochemistry. Proteins purified from experimental xenografts and malignant tumors using antibody- or lectin-affinity columns in the presence of 5 mmol/L EDTA were assayed for seprase activation in vivo. Seprase expression and activation occur most prevalently in ovarian carcinoma but were also detected in four other malignant tumor types, including adenocarcinoma of the colon and stomach, invasive ductal carcinoma of the breast, and malignant melanoma. Together, these data show that, in malignant tumors, seprase is proteolytically activated to confer its substrate specificity in collagen proteolysis and tumor invasion. (Cancer Res 2006; 66(20): 9977-85)

Studies on the relevance of the N-nitrosamines to esophageal cancer in China are reviewed. Esophageal cancer is a complex and multifactorial problem. Although a causal association between nitrosamines exposure and esophageal cancer in China has not yet been rigorously established, exposure of Lin-Xian subjects to nitrosamines either directly or as a result of their in vivo formation has been detected in our study. Several N-nitrosamines (NDMA, NDEA, NMBzA, NPyr, NPip, and NSAR) in gastric juice collected from Lin-Xian inhabitants have been detected. A correlation was found between the lesions of esophageal epithelium and the amount of nitrosamines present. In addition, the amounts of N-nitrosamino acids (N-nitrosoproline, N-nitrosothiazolidine 4-carboxylic acid, NSAR, and nitrates) excreted in 24-hr urine of subjects in Lin-Xian were significantly higher than those in Fan-Xian, indicating a higher exposure to N-nitroso compound and their precursors of the inhabitants in the high-risk area. The effect of nitrosamines on human esophagus has been investigated at the cellular levels. The amounts of O6-MedG in DNA of esophageal or stomach mucosa of patients from Lin-Xian were higher than that from Europe (Lyon and Essen). The presence of O6-MedG in the human fetal esophagus cultured with NMBzA was also detected. These findings indicate that the elevated levels of O6-MedG in esophageal DNA could be the result of a recent exposure to N-nitroso compounds or a genetically determined reduced cellular capacity for repair of O6-MedG from DNA. The hyperplasia was induced in the esophagus of human fetus that cultured with NMBzA for 2 weeks to 2 months. The intervention studies of esophageal cancer in Lin-Xian have been pursued. Intake of moderate doses of ascorbic acids by Lin-Xian subjects effectively reduced the urinary levels of N-nitrosamino acids to those found in un-dosed subjects in the low-risk area. If N-nitroso compounds are formed in vivo and are among the causative factors of esophageal cancer in Lin-Xian, ascorbic acid appears to be effective in lowering the body burden of these carcinogenic compounds. Thus, the plan of chemoprevention is carried out in Lin-Xian.