Effect of sodium selenite on proliferation of endometrial cancer cells

AIM: To investigate the effect and mechanism of sodium selenite (Na₂SeO₃) on the proliferation of endometrial cancer cells. METHODS: Endometrial cancer Ishikawa cells and HEC-1A cells were treated with Na₂SeO₃. The effect of Na₂SeO₃ on cell proliferation was determined by MTT assay. The effects of Na₂SeO₃ on cell cycle distribution and apoptosis were tested by flow cytometric analysis. The expression of cyclin A was detected by Western blotting. RESULTS: Na₂SeO₃ inhibited the proliferation of Ishikawa cells and HEC-1A cells. For Ishikawa cells, IC₅₀ was 3.26 μmol/L, and for HEC-1A cells, IC₅₀ was 4.77 μmol/L. After treated with Na₂SeO₃, the cells in G₀/G₁ phase were reduced and the cells in S phase and G₂/M phase were increased. Na₂SeO₃ also increased the percentage of apoptosis cells. The result of Western blotting showed that the expression of cyclin A was increased. CONCLUSION: Na₂SeO₃ inhibits the proliferation of endometrial cancer Ishikawa cells and HEC-1A cells via up-regulating the expression of cyclin A, arresting cell cycle and inducing apoptosis.     

Rapid preparation of hypoxic Krebs-Henseleit solution with sodium sulfite for in vitro model of hypoxic pulmonary vasoconstriction

AIM: To investigate the feasibility of using sodium sulfite (Na₂SO₃) to prepare hypoxic Krebs-Henseleit (KH) solution for hypoxic pulmonary vasoconstriction (HPV) model in vitro. METHODS: Different doses of Na₂SO₃ were added into 0.5 L KH solution at 37°C. An i-STAT portable clinical analyzer was used to measure the oxygen partial pressure (PO₂), carbon dioxide partial pressure (PCO₂), pH value and the concentration of sodium (Na⁺) in these KH-Na₂SO₃ solutions 1 min after administration. Then the dose of Na₂SO₃ suitable for HPV model was dissolved in 0.5 L KH solution and the above indexes in the solution were monitored at various time points at 37°C under atmospheric pressure. RESULTS: More than 0.2 g (including 0.2 g) Na₂SO₃ reduced the PO₂ of 0.5 L KH solution in a dose-dependent manner (P<0.01). In addition, 1.5 g Na₂SO₃ reduced the PO₂ of 0.5 L KH solution to 20~40 mmHg and maintained the hypoxic state for at least 90 min (suitable for HPV model in vitro), but had nearly no effect on the PCO₂, pH and Na⁺ levels. CONCLUSION: The hypoxia solution for HPV model could be reached by Na₂SO₃ in open air and the method is simple, easily feasible and stable.     

Influence of maxadilan on human adipose-derived stem cells

AIM: To investigate the effect of maxadilan, which specifically activates pituitary adenylate cyclase-activating polypeptide type I receptor (PAC1 receptor), on the proliferation, apoptosis and differentiation potential of human adipose-derived stem cells (ASCs). METHODS: ASCs from human adipose tissue were isolated by enzymatic digestion and cultured. ASCs were confirmed by the analysis of the markers for cell phenotypes by flow cytometry (FCM) and adipogenic/osteogenic induction. The effect of maxadilan on ASCs viability was analyzed by CCK-8 assay and FCM. ASCs were irradiated by ultraviolet C (UVC) at 254 nm and the absorbance of apoptotic ASCs induced by various doses of UVC was measured by CCK-8 assay. ASCs were exposed to 702 J/m² UVC for 24 h to induce apoptosis. The effect of maxadilan on ASC apoptosis was analyzed by FCM and the determination of caspase 3 and caspase 9 levels. RESULTS: Adipose-derived stem cells were confirmed by the detection of the positive expression of cell phenotypes including CD29, CD44, CD59 and CD105 by FCM. The data of CCK-8 assay revealed that ASCs treated with maxadilan (80 nmol/L) had the strongest ability of proliferation. The data of FCM also demonstrated that the addition of 80 nmol/L maxadilan to ASCs in experimental group markedly improved the proliferation capacity of the cells compared with control group (P<0.05). The apoptosis of ASCs exposed to 702 J/m² UVC was dramatically inhibited by the treatment with maxadilan (80 nmol/L). Such process involved the caspase signaling pathway including caspase 3 and caspase 9. There was statistical significance (P<0.05) between experiment group (ASCs irradiated by UVC and supplemented with maxadilan) and control group (ASCs only irradiated by UVC). Meanwhile, adipogenic and osteogenic differentiation potentials were both positive in experiment group and control group. CONCLUSION: Maxadilan promotes proliferation and inhibits apoptosis of the ASCs. The differentiation potential of ASCs toward adipogenic and osteogenic lineages wouldn't be altered by maxadilan. Maxadilan would benefit to growth and expansion of ASCs in vitro.     

碳酸钠和碳酸氢钠对番茄壮苗和灰霉病的防治作用

以番茄品种L402幼苗为试材,研究外源喷施碳酸钠和碳酸氢钠对番茄生长和灰霉病的防治作用。结果表明:25mmol·L~(-1)和50 mmol·L~(-1)的碳酸钠与碳酸氢钠对番茄幼苗具有显著的壮苗作用;30～60 mmol·L~(-1)的碳酸钠和碳酸氢钠对灰霉菌菌丝生长具有不同程度的抑制作用,且抑制作用通常随着浓度的升高而加强;30～40 mmol·L~(-1)的碳酸钠和50～60mmol·L~(-1)的碳酸氢钠对番茄灰霉病具有良好的预防作用;50 mmol·L~(-1)的碳酸钠和50～60 mmol·L~(-1)的碳酸氢钠对番茄灰霉病具有良好的抑制作用。生产上可应用适宜浓度的碳酸钠和碳酸氢钠防治番茄灰霉病。     

The role of TF-1 cell apoptosis-related gene 19 in systemic lupus erythematosus

AIM:To clarify the role of new apoptosis-related gene, TF-1 cell apoptosis-related gene 19(TFAR19), in the pathogenesis of systemic lupus erythematosis (SLE) and the relationship between TFAR19 and SLE.METHODS:DNA Ladder detection, Western blotting, immunological fluorescence method, ELISA and so on were used to test if ultraviolet B(UVB) could induce HaCaT cell apoptosis and TFAR19 expression.RESULTS:HaCaT cell apoptosis could be detected after 24 hours of 30 mj/cm² UVB irradiation. Also, we found that in active SLE patients, the TFAR19 antibody was increased, but not significant compared to the normal control.CONCLUSION:TFAR 19 is involved in the process of UVB induced ketatinocyte line HaCaT apoptosis and SLE pathogenesis.     

Role of potassium channels in the regulation of intracellular free Ca2+ concentration of pulmonary artery smooth muscle cells in rats

AIM: To investigate the role of potassium channels in the regulation of intracellular free calcium concentration ( [Ca²⁺]_i) of pulmonary artery smooth muscle cells (PASMCs) in rats. METHODS: The fluorescence Ca²⁺ indicator Fura-2/AM was used to observe [Ca²⁺]_i of rat PASMCs in normal and chronic hypoxic condition. The influences of potassium channels on PASMCs proliferation were assessed by MTT assay. RESULTS: 1. In normoxic condition, [Ca²⁺]_i was (156.91±8.60) nmol/L, and in hypoxic condition, [Ca²⁺]_i was (294.01±16.81) nmol/L. 2. In normoxic condition, the voltage-dependent K⁺-channel antagonist 4-aminopyridine (4AP), but not the Ca²⁺-activated K⁺-channel antagonist tetraethylammonium (TEA) and the ATP-sensitive K⁺-channel antagonist glibenclamide (Glib) increased [Ca²⁺]_i. 3. In hypoxic condition, 4AP and TEA caused the rise in [Ca²⁺]_i , but Glib had no effect on [Ca²⁺]_i. 4. MTT assay showed that 4AP increased the value of absorbing light degree (A value) in normoxic and hypoxic condition (0.582±0.062,0.873±0.043,respectively, P＜0.01), TEA increased A value only in hypoxic condition, and Glib had no effect on the proliferation of PASMCs. CONCLUSIONS: K_V plays an important role in the regulation of [Ca²⁺]_i and proliferation of PASMCs. K_Ca serves as distinct responsive roles in the regulation of proliferation of PASMCs in hypoxic condition. K_ATP has no effect on [Ca²⁺]_i and proliferation of PASMCs in normoxic and hypoxic conditions.     

C*HSDGIC* from cyclization of PACAP (1-5) attenuates UVB-induced apoptosis of human retinal pigment epithelial cells

AIM: To investigate the influence of C^*HSDGIC^* (CHC), a cyclopeptide from the cyclization of with disulfide, on the proliferation and apoptosis of cultured human retinal pigment epithelial (RPE) cells induced by ultraviolet B(UVB). METHODS: The expression of PACAP type 1 (PAC1) receptor in human RPE cells was identified by Western blotting. The cells were exposed to UVB irradiation and cultured in fresh medium with or without gradient concentrations (1 nmol/L to 1 mmol/L) of CHC. The viability of the cells was determined by CCK-8 assay. The early apoptosis of the cells was detected by flow cytometry with annexin V and propidium iodide staining.The mitochondrial menbrane potential was detected by flow cytometry with JC-1 staining. RESULTS: The PAC1 receptor in human RPE cells was identified by Western blotting. The best results of CHC on the proliferation and anti-apoptosis of human RPE cells were achieved at the concentration of 100 μmol/L, which increased the viability by (34.23±3.39)% and (20.10±1.48)%, respectively. The percentage of apoptotic cells was decreased by (5.63±1.49)% with CHC treatment (100 μmol/L) after UVB irradiation,and the percentage of mitochondrium-depolarizing cells was decreased by (5.2±0.5)%. CONCLUSION: PAC1 receptor exists in human RPE cells. C^*HSDGIC^* increases the viability of RPE cells and attenuates UVB-induced apoptosis.     

Effect of progesterone on SH-SY5Y cells injured by ATP

AIM: To investigate the neuroprotective effect of progesterone against adenosine triphosphate (ATP)-injured human neuroblastoma SH-SY5Y cells.METHODS: The SH-SY5Y cells in the logarithmic phase were divided into different groups according to the progesterone and ATP concentrations. The cell viability was measured by CCK-8 assay. The membrane permeability was detected using fluorescent dye YO-PRO-1. Cytosolic Ca²⁺ concentration was measured with fluorescent dye Fluo-3/AM. The expression of purinergic P2X₇ receptor was assessed by Western blot.RESULTS: The viability of the SH-SY5Y cells was significantly decreased (P<0.05) and YO-PRO-1 uptake was obviously increased (P<0.05) in a concentration-dependent manner compared with control group when SH-SY5Y cells were treated with ATP at 1, 3, 5 and 7 mmol/L for 2 h. The viability reduction of the SH-SY5Y cells induced by ATP was obviously counteracted by treatment with progesterone at 3, 10 and 30 nmol/L for 30 min (P<0.05) as compared with ATP group. YO-PRO-1 fluorescence enhancement induced by ATP in SH-SY5Y cells was significantly reduced (P<0.05) by progesterone (30 nmol/L) or P2X₇ receptor antagonist KN-62 (500 nmol/L) pretreatment for 30 min, and no obvious difference between treatments with progesterone and KN-62 was observed. Cytosolic Ca²⁺ fluorescence intensity in normal group was a little, but that in ATP group was increased (P<0.05). Progesterone or KN-62 pretreatment significantly decreased the cytosolic fluorescence intensity of Ca²⁺ induced by ATP (P<0.05). However, no obvious difference between treatments with progesterone and KN-62 was found. The expression of P2X₇ receptor in ATP group was significantly higher than that in control group (P<0.05), and progesterone inhibited ATP-induced P2X₇ receptor expression (P<0.05).CONCLUSION: Progesterone inhibits P2X₇ receptor expression, membrane pore formation, intracellular Ca²⁺ increase and cell death induced by ATP, so progesterone may protect SH-SY5Y cells against ATP-induced injuries.     

Se-containing spirulina phycocyanin attenuated liver injury induced by carbon tetrachloride in mice

AIM:To investigate the effect of Se-containing spirulina phycocyanin (Se-SPC) on liver injury of mice induced by carbon tetrachloride (CCl₄). METHODS:The mouse model was conducted by intragastric feeding with 2% CCl₄ oil for three times, meanwhile Se-SPC, spirulina phycocyanin (SPC) and Na₂SeO₃ were injected (ip) to various groups for 7 days. Then selenium (Se), glutathione peroxidase (GPx), superoxide dismutase (SOD), alanine aminotransferase (ALT), malondiaoldehyde (MDA) and nitric oxide (NO) levels in blood and liver were measured. RESULTS:The level of Se,GPx and SOD activities were obviously higher(P<0.05)but ALT activity,MDA and NO₂^-/NO₃^- levels were remarkably lower(P<0.05)in Se-SPC treated groups than those in CCl₄ groups,and effects of high dose Se-SPC on Se,GPx,MDA and NO₂^-/NO₃^- were even more significant(P<0.01).Under the same dose of Se or protein,effects of all selected targets in Se-SPC groups were more efficient than those in SPC groups and inorganic-Se groups.Furthermore,Se levels had a positive correlation with GPx activity(r=01705),which had negative correlation with levels of MDA,NO₂^-/NO₃^- and ALT(r=-0.629,r=-0.336,r=-0.457,respectively), and positive correlations between ALT activity and MDA or NO₂^-/NO₃^- level were found (r=0.519,r=0.641). CONCLUSION:These results indicated that Se-SPC may attenuate liver injury of mice induced by CCl₄ through its anti-inflammatory action and enhancing selenoenzyme expression.     

《科技导报》（半月刊）欢迎订阅、欢迎投稿、欢迎刊登广告

AIM: To study the effects of ultraviolet (UV) on mitochondrial functions and apoptosis in HaCaT cells. METHODS: After irradiation by UV at low dose (UVA 2 J/cm2，UVB 10 mJ/cm2) and high dose (UVA 6 J/cm2，UVB 30 mJ/cm2), HaCaT cells were cultured for 15 hours. Flow cytometry was used to measure mitochondrial membrane potential, mitochondrial mass and apoptotic rate. Annexin V-FITC/PI staining of apoptotic cells was analyzed by laser confocal microscopy. RESULTS: After UV irradiation, cell proportion with low mitochondrial membrane potential increased with irradiation doses. The proportion of control group, low dose group and high dose group were 7.94%±1.02%, 25.87%±4.55% and 39.27%±5.32%, respectively. Cells proportion with low mitochondrial mass increased with irradiation doses. The proportion of control group, low dose group and high dose group were 15.19%±1.58%, 40.36%±4.41% and 68.79%±5.46%, respectively. The hypodiploid peaks of DNA content analysis represented the apoptotic rate of HaCaT cells. The apoptotic rate of control group, low dose group and high dose group were 1.82%±0.51%, 30.16%±5.47% and 58.49%±5.98%, respectively. To analyze the cells apoptosis by staining with annexin V-FITC and PI, the results were consistent with those of DNA content analysis. Cells in control group showed almost no positive staining cells. Single annexin V-FITC positive cells in low dose group and double positive cells in high dose group were predominant, respectively. CONCLUSION: UV irradiation induces HaCaT cell mitochondrial depolarization, as well as mitochondrial mass loss. These changes are related to cell apoptosis.     

Inhibitory effect of 1,25-(OH)2D3 on proliferation of airway smooth muscle cells from passively-sensitized patients

AIM: To investigate the effects of 1,25-dihydroxyvitamin D₃ on the proliferation of passively-sensitized human airway smooth muscle cells (HASMCs), and to explore its potential role in asthmatic airway remodeling.METHODS: HASMCs were passively sensitized with 10% serum from asthmatic patients.1,25-(OH)₂D₃ was used as the interventor.The effect of 1,25-(OH)₂D₃ on the cell proliferation and its optimal concentration were determined by MTT colorimetric assay.The cell cycle analysis was performed by flow cytometry.The expression of proliferating cell nuclear antigen (PCNA) was measured by the method of immunocytochemical staining.RESULTS: 1,25-(OH)₂D₃ at the concentrations of 10^-9-10^-7 mol/L markedly inhibited the cell proliferation and the maximum effect was observed at the concentration of 10^-7 mol/L.This concentration of 1,25-(OH)₂D₃ markedly suppressed the PCNA-positive rate and hampered the G₁/S transition in HASMCs passively-sensitized by asthmatic serum.CONCLUSION: 1,25-(OH)₂D₃ has direct inhibitory effects on the proliferation of passively-sensitized HASMCs in vitro, which may be concerned with the beneficial role of 1,25-(OH)₂D₃ on the prevention and therapy of asthmatic airway remodeling.     

Calcitonin gene-related peptide triggers proliferation in HaCaT keratinocytes by ERK1/2 signaling pathway

AIM: To investigate the effect of calcitonin gene-related peptide (CGRP) on the proliferative potential of HaCaT keratinocytes and whether CGRPR and ERK1/2 pathway is involved in this progress.METHODS: ［³H］^-TdR test was used to estimate the CGRP-induced proliferative potential of HaCaT keratinocytes and the influence of CGRP8-37 (CGRP receptor 1 antagonist) and PD98059 (ERK1/2 inhibitor) on this effect.Western blotting was used to test the activation of ERK1/2 pathway.RESULTS: Exposure of HaCaT keratinocytes to CGRP induced proliferation through the CGRP receptor and ERK1/2 pathway.CGRP 8-37 and PD98059 inhibited CGRP-induced proliferation of HaCaT keratinocytes.Phosphorylation of ERK1/2 was activated by CGRP in a time-dependent manner,which was inhibited by CGRP 8-37 and PD98059.CONCLUSION: This study indicates that CGRP triggers the proliferation of HaCaT keratinocytes by CGRP receptor and ERK1/2 signaling pathway.     

硒提高马铃薯幼苗抗寒性生理研究

低温胁迫是影响我国南方冬闲田马铃薯种植的一大阻碍因素,发生在马铃薯植株生长发育前期。以马铃薯品种金 湘为试材,研究不同硒浓度（1、3、5 mg·L^-1）和适宜浓度的喷硒次数（1、2、3 次）对马铃薯幼苗抗寒性的影响。结果表 明：外源喷施1、3、5 mg·L^-1 亚硒酸钠处理均可以提高低温胁迫下马铃薯叶片中叶绿素含量,降低叶片相对电导率以及丙 二醛（MDA）、H2O2 含量,且3 mg·L^-1 的喷施效果最佳。低温胁迫下喷施3 mg·L^-1 亚硒酸钠溶液1～3 次的幼苗叶绿素含 量高于未喷硒对照,MDA 和H2O2 含量均降低,且喷施2 次时效果最好。综上,适宜的喷硒浓度（3 mg·L^-1）和次数（2 次） 能在一定程度上降低低温胁迫所带来的伤害,从而提高马铃薯幼苗的抗寒性。     

Heterogeneity of basal intracellular calcium concentration and its relations to the reactivity in mouse peritoneal macrophages

AIM: To investigate the heterogeneity of basal intracellular free calcium concentration([Ca²⁺]_i) in peritoneal macrophages(PM) and whether it is relative to the reactivity of PM at the single cell level. METHODS:[Ca²⁺]_iimplicated stimulated were measured by fluorescent microscopic imaging system after loading with fluorescent probe fura-2/AM. Superoxide(O₂^-)produced by single PM was determined by modified NBT test. RESULTS: The values of basal[Ca²⁺]_idetermined in 392 PMs of 7 mice showed normal distribution [(54±24) nmol/L, n=392] with wide range(less than 20 nmol/L to more than 100 nmol/L), among which about 50% were in the range of 40-60 nmol/L. When stimulated with PMA or fMLP,[Ca²⁺]_iwas increased, the peak values were positively correlated with the basal[Ca²⁺]_iin one mouse(PMA stimulated cells: r=0.52, P<0.01, n=58; fMLP stimulated cells:r=0.59, P<0.01, n=44. Both of the experiments were repeated in 3 mice, the results in the other 2 mice were similar). The O₂^- in PMA stimulated PMs were also positively correlated with the basal i(r=0.42, P<0.01, n=43, repeated in 4 mice, the results in the other 3 mice were similar). CONCLUSION: Basal[Ca²⁺]_iin murine PM is heterogeneous, and the value of basal[Ca²⁺]_iis tightly correlated to the reactivity of PM stimulated by proinflammatory factors.     

High glucose induces H9c2 cardiomyocyte hypertrophy through Ca2+-CaN-NFAT3 signaling pathway

AIM: To study the morphological changes of cardiac H9c2 cells during the developmental process of fetal rat. METHODS: Embryonic rat heart-derived H9c2 cells were maintained in DMEM supplemented with 10% fetal bovine serum. The H9c2 cells were plated at a density of 6000 cells/cm and divided into 5 groups: H9c2 cells were treated with 5 mmol/L glucose, 25 mmol/L glucose, 50 mmol/L glucose, Norvasc (25 nmol/L)+25 mmol/L glucose, or Norvasc (25 nmol/L)+50 mmol/L glucose for 48 h. The morphology of H9c2 cells was observed. The cell surface area was measured by Image-Pro Plus 6.1 software. Fluorescence spectrophotometry was used to detect the concentration of intracellular calcium ion ([Ca²⁺]_i)in the cardiomyocytes. The concentration of CaN in the cell was measured by ELISA. The mRNA expression of CaNAβ, NFAT3 and β-MHC in the cells was detected by real-time PCR. The protein levels of CaNAβ, NFAT3 and β-MHC in cultural H9c2 cells were detected by Western blot. RESULTS: The mean area of the cells, the mean fluorescence value of [Ca²⁺]_i and the concentration of CaN in 25 mmol/L glucose group were higher than those in 5 mmol/L glucose group, and those were lower than those in 50 mmol/L glucose group. After treated with Norvasc, those results decreased significantly. The expression of CaNAβ, NFAT3 and β-MHC at mRNA and protein levels in 25 mmol/L glucose group was higher than those in 5 mmol/L glucose group, but was lower than those in 50 mmol/L glucose group. The expression of CaNAβ, NFAT3 and β-MHC at mRNA and protein levels decreased significantly in Norvasc treatment group. CONCLUSION: Ca²⁺-CaN-NFAT3 signaling pathway is perhaps involved in high glucose-induced H9c2 cardiomyocyte hypertrophy.     

Effects of Bene Jones protein and TGF-β1 on the proliferation of rat renal proximal tubular cells in vitro

AIM:To study the effect of Bene Jones protein (BJP) from multiple myeloma(MM) patient and TGF-β₁ on cultured renal proximal tubular cell(PTC) proliferation.METHODS:[H³]TdR incorporation was used to study the effect of λBJP and TGF-β₁ on cultured rat NRK.52E PTC proliferation, the expression of TGF-β₁ in the supernatant of PTC cultured with BJP was assessed with ELISA.RESULTS:① [H³]TdR incorporation of PTC was inhibited by BJP in a dose-dependent manner, when co-cultured with 100-800 μmol/L BJP and 2.0 μg/L TGF-β₁, the [H³]TdR incorporation was lower than that of BJP alone, especially when BJP≥400 μmol/L;②The expression of TGF-β₁ in the supernatant of PTC cultured with BJP was increased, especially when BJP≥400 μmol/L(P＜0.05);③ The [H³]TdR incorporation of PTC was also inhibited by exogenous TGF-β₁ in a dose-dependent manner.CONCLUSION:λBJP has antiproliferative effect on rat PTC in vitro, The effect is related with stimulating the PTC to produce excessive TGF-β₁, which also has antiproliferative effect on PTC in some degree.     

Effect of urotensin II on cultured pulmonary arterial smooth muscle cells of rabbits

AIM: The effect of urotensin II (U-II) on proliferation of cultured pulmonary arterial smooth muscle cells (PASMCs) of rabbits and its mechanism are investigated. METHODS: PASMCs were isolated using explant technique. RPASMCs were incubated in serum-free medium with different concentrations of nicardipine, calcimodulin antagonist W₇, PKC inhibitor H₇ or MAPK inhibitor (PD₉₈₀₅₉), with or without U-II. RPASMC proliferation was examined by MTT assay and by the increase in [³H]-thymidine incorporation into DNA. RESULTS: U-II (10^-9 mol/L-10^-7 mol/L) increased A value of PASMCs by MTT assay and [³H]-thymidine incorporation in PASMCs in a dose-dependent manner. U-II induced a maximal effect at a concentration of 10^-7 mol/L. A value and [³H]-thymidine incorporation rose 42.9% and 68.5% (P<0.05), respectively. U-II had no effect at a concentration of 10^-10 mol/L. Nicardipine, W₇, H₇, PD₉₈₀₅₉ (10^-7 mol/L-10^-5 mol/L) inhibited the effect of U-II in inducing increase of A value and -thymidine incorporation in a dose-dependent manner, with the maximal inhibitory rate of 42.3%, 19.6%, 23.2%, 10.5% (P<0.05) in A value and 46.6%, 9.8%, 21.7%, 14.7% (P<0.05) in [³H]-thymidine incorporation at concentration of 10^-5 mol/L, respectively. CONCLUSION: Our results suggest that U-II may induce proliferation of PASMCs of rabbits by Ca²⁺, CaM, PKC and MAPK signal transduction pathway.     

Effect of miR-21 over-expression on proliferation,migration and diffe-rentiation of c-Kit+ cardiac stem cells

AIM: To explore the effect of microRNA (miR)-21 on proliferation, migration and differentiation abilities of c-Kit⁺ cardiac stem cells (CSCs). METHODS: c-Kit⁺ CSCs were cultured and selected by the methods of enzyme digestion and magnetic bead separation. miR-21 mimics (50 nmol/L) and mimics negative control (MNC) were transfected into c-Kit⁺ CSCs with Lipofectamine^® 2000. The cells was divided into 3 groups:control group:c-Kit⁺ CSCs without any pretreatment; MNC group:the cells were transfected with MNC for 48 h; mimics group:the cells were transfected with miR-21 mimics for 48 h. qPCR was used to assess the expression of miR-21 in each group. CCK-8 and EdU assays were used to determine the cell proliferation. qPCR and immunofluorescence were used to detect the differentiation in each group. Scratch assay was adopted to explore the migration ability of the cells. RESULTS: The expression of c-Kit in the c-Kit⁺ CSCs were 90.8%, with 0.6% of CD45 and 0.5% of CD34. A significant increase in miR-21 expression was observed when the cells were transfected with miR-21 mimics for 48 h (P<0.05). CCK-8 and EdU assays showed that miR-21 significantly increased cell proliferation as compared with MNC group and control group (P<0.05). No difference in the expression of Nkx2.5, CD31 and α-SMA at mRNA and protein levels was observed, and no difference of the migration ability in 3 groups of the c-Kit⁺ CSCs was found. CONCLUSION: Over-expression of miR-21 significantly promotes the proliferation of c-Kit⁺ CSCs, without any effect on the cell migration and differentiation.     

Hypoxic preconditioning increases tolerant ability of old human bone marrow mesenchymal stem cells to oxidative stress injury through AKT pathway

AIM: To investigate the protective effect of hypoxic preconditioning on human bone marrow mesenchymal stem cells (hBM-MSCs), and to provide basic experimental support for more effective autologous stem cell transplantation in aged patients. METHODS: The old hBM-MSCs were subjected to hypoxic preconditioning using a hypoxia incubator chamber for 24 h. The cells were divided into young group, old group and old+hypoxia group (with 24 h hypoxic preconditioning). Hydrogen peroxide (H₂O₂, 300 μmol/L) was applied to simulate the oxidative stress. The cells were treated with 50 μmol/L LY294002 for 2 h to inhibit PI3K/AKT pathway. BrdU incorporation and CCK-8 assay were used for analyzing the cell proliferation and viability. The protein levels of Bax, Bcl-2 and p-AKT were measured by Western blot. RESULTS: BrdU-positive cells, which represented the cell proliferation, and the cell viability were significantly increased in old+hypoxia group compared with old group (P<0.05). The protein level of Bax decreased (P<0.05) and Bcl-2 increased (P<0.05) in old+hypoxia group compared with old group after using 300 μmol/L H₂O₂ simulate. the oxidative stress. The phosphorylation of AKT was enhanced by hypoxic preconditioning in old group (P<0.05). The protective effect of hypoxic preconditioning on the cell survival was decreased after treated with LY294002 (inhibitor of the PI3K/AKT pathway) (P<0.05). CONCLUSION: Hypoxic preconditioning increases the survival and proliferation of old hBM-MSCs by activation of AKT pathway.