Long non-coding RNA H19 facilitates bone marrow mesenchymal stem cell survival and vascularization in hypoxic-ischemic conditions in vitro / 中国组织工程研究

BACKGROUND: Our previous study demonstrated that bone marrow mesenchymal stem cells (MSCs) presented with a low survival rate and newly formed vascular-like structures was sparsely distributed in the local infarct tissues after cell transplantation, which certainly impaired the therapeutic efficacy. Long non-coding RNA-H19 (lncRNA-H19) has been confirmed to be associated with MSCs differentiation and mediate vascularization. OBJECTIVE:To observe the influence of lncRNA-H19 on the survival and vascularization potential of MSCs in vitro and to explore the possible mechanism. METHODS:MSCs were obtained and cultured in vitro.Cells were divided into five groups:MSCs+H19,MSCs+H19 negative control (MSCs+H19 NC), MSCs+si-H19, MSCs+si-H19 negative control (MSCs+si-H19 NC) and MSCs groups. MSCs+H19 and MSCs+H19 NC groups were transfected with lncRNA-H19 and lncRNA-H19 scramble RNA respectively, while MSCs+siH19 and MSCs+si-H19 NC groups were transfected with lncRNA-H19 siRNA and lncRNA-H19 siRNA scramble respectively. Cells were cultured under hypoxic-ischemic condition (serum-free medium, 1% O2) for 24 hours. Then, cell proliferation and apoptosis were evaluated using MTS and TUNEL, respectively. Cell supernatant from each experimental group was further co-cultured with human umbilical vein endothelial cells to induce vascularization. The expression of vascular endothelial growth factor A (VEGFA) was thereafter detected using western blot assay. RESULTS AND CONCLUSION: Compared with MSCs+H19 NC and MSCs groups, MSCs+H19 group presented with significantly higher proliferation rate, lower apoptosis percentage and a larger number of vascular branches on matrigel (P < 0.01). There was a significantly higher expression of VEGFA in the MSCs+H19 group than MSCs+H19 NC and MSCs groups. Compared with the MSCs and MSCs+si-H19 NC groups, MSCs+H19 group presented with significantly lower proliferation rate, higher apoptosis percentage and a less number of vascular branches on matrigel (P < 0.01). In addition, VEGFA expression was distinctly downregulated in the MSCs+si-H19 group in comparison with the MSCs+si-H19 NC and MSCs groups. These findings indicate that lncRNA-H19 effectively promotes MSCs survival and vascularization under hypoxic-ischemic condition in vitro,and this effect may be associated with the upregulation of VEGFA.

LncRNA-Braveheart promotes the differentiation of bone marrow mesenchymal stem cells in vitro into cardiomyocyte-like cells / 中国组织工程研究

BACKGROUND:Our previous work demonstrated that bone marrow mesenchymal stem cells (BMSCs) transplantation could improve cardiac function in rats with myocardial infarction.However,the overall efficacy was unsatisfactory,and there was a low efficiency of BMSCs differentiating into cardiomyocytes in the local infarct myocardium.OBJECTIVE:To transfect long non-coding RNA-Braveheart (IncRNA-Bvht) into BMSCs in order to observe whether it could promote cardiomyocyte differentiation of BMSCs in vitro.METHODS:pLVX-IRES-ZsGreen1-IncRNA-Bvht vector was constructed and applied to transfect IncRNA-Bvht into bMSCs,and then,the transfection efficiency was detected.BMSCs were obtained from C57BL/6 mice and cultured ir vitro.Passage 3 cells were divided into three groups:BMSCs group,null vector group and IncRNA-Bvht group.All cells in the three groups were cultured in the normal condition for 48 hours and cardiomyocytes differentiation was induced by 5-azacytidine for another 24 hours followed by 2-week culture under normal conditions.Cardiomyocyte differentiation of BMSCs was observed under fluorescence microscopy and expression of cardiac specific cell markers including troponin T and myosin were examined using immunofluorescent staining,western blot assay,and qRT-PCR.RESULTS AND CONCLUSION:Cell morphological changes could be observed in all the groups 2 weeks after the induction.Interconnected cells arranged consistently in all the three groups.Immunofiuorescent staining results showed that the expression of troponin T and myosin was notably positive,and the proportion of troponin T positive cells was significantly increased.qRT-PCR and western blot assay results indicated that there were significantly increased levels of troponin T and myosin in the IncRNA-Bvht group as compared with the BMSCs and null vector groups (P ＜ 0.01),suggesting that IncRNA-Bvht could efficiently promote cardiomyocyte differentiation of BMSCs in vitro.

Throat infection, neck spinal disease, chest pain and cardiac response: a new clinical syndrome? / 中华心血管病杂志

<p><b>OBJECTIVE</b>To analyze the characteristics of a new clinical syndrome, including throat infection, neck spinal disease, chest pain and cardiac response.</p><p><b>METHODS</b>A total of 165 patients with above mentioned symptoms admitted to Tongji hospital from 2003 to 2005 were included in this study and underwent further medical history inquiry, physical examination and laboratory tests. Eighty-five healthy subjects served as controls. Serum myocardial auto-antibodies against beta(1)-adrenoceptor, alpha-myosin heavy chain, M(2)-muscarinic receptor and adenine-nucleotide translocator were detected, inflammatory cytokines, high sensitivity C-reaction protein, serum antibodies against Coxsackie virus-B, cytomegalovirus, Mycoplasma pneumoniae and Chlamydia pneumoniae were determined and lymphocyte subclasses were assayed by flow cytometry.</p><p><b>RESULTS</b>All patients had a complex of four symptoms or tetralogy: (1) persistent throat or upper respiratory tract infection; (2) neck pain; (3) chest pain; (4) chest depression or dyspnea, some of them with anxiety. Anti-myocardial auto-antibodies (AMCA) were present in all patients vs. 8% in controls. TNF-alpha, IL-1 and IL-6 were significantly higher in patients than controls (P < 0.01). CD3(+) and CD4(-)CD8(+) lymphocytes were significantly higher and CD56(+) lymphocytes lower in patients than those in controls (P < 0.01). The ratios of serum pathogen antibodies positive against Coxsackie virus-B, cytomegalovirus, Mycoplasma pneumoniae and Chlamydia pneumoniae were all significantly higher in patients than in controls.</p><p><b>CONCLUSIONS</b>These data led to identification of a persistent respiratory infection-related clinical syndrome, including persistent throat infection, neck spinal lesion, rib cartilage inflammation, symptoms of cardiac depression and dyspnea with or without anxiety.</p>

Effects of telmisartan on endoplasmic reticulum stress induced cardiomyocyte apoptosis in abdominal aortic banded rats / 中华心血管病杂志

<p><b>OBJECTIVE</b>To investigate the effects of telmisartan on endoplasm reticulum (ER) stress signal pathways and cardiomyocyte apoptosis in abdominal aortic banded rats.</p><p><b>METHODS</b>Male SD rats were randomly divided into sham-operated group, abdominal aortic banding group (AAB) and AAB + telmisartan (5 mgxkg(-1)xd(-1) per gavage, beginning at 1 week before AAB for 8 weeks, n = 10 each). Ten weeks post AAB, hemodynamic measurements were performed, whole heart and left ventricular weights were obtained, cardiomyocyte apoptosis was measured by TUNEL method. Myocardial GRP78 and CHOP protein expressions were detected by Western blot and immunohistochemistry.</p><p><b>RESULTS</b>The ratio of left ventricular weight to body weight, the ratio of heart weight to body weight, left ventricular end diastolic pressure and the apoptosis index were significantly increased while left ventricular end systolic pressure and +/- dp/dt(max) were significantly decreased in AAB group than those in sham-operated group (all P < 0.01), these changes could be significantly attenuated by telmisartan (all P < 0.01). Moreover, myocardial GRP78 and CHOP expressions were significantly upregulated in AAB group than those in sham-operated group and telmisartan could significantly downregulate the increased GRP78, CHOP expressions (all P < 0.01).</p><p><b>CONCLUSIONS</b>Increased ER stress might be responsible for enhanced cardiomyocyte apoptosis in AAB rats. Telmisartan effectively attenuated the cardiomyocyte apoptosis and cardiac hypertrophy in AAB rats possibly through reducing ER stress.</p>