Effects of cytokines on renovation of acute renal failure in mouse with bone marrow derived mesenchymal stem cell transplantation

AIM: To observe the effects of cytokines on renovation of acute renal failure (ARF) in mouse with bone marrow derived mesenchymal stem cell (MSC) transplantation. METHODS: ARF animal model was induced in mouse by subcutaneous injection of cisplatin. Mice were randomly assigned into 3 groups: normal control group, ARF group and MSC group. After 24 h cisplatin injection, animals were injected intravenously with MSC in MSC group. Animals were sacrificed at 1 d, 4 d, 7 d, 14 d and 28 d after cisplatin injection. The blood urea nitrogen (BUN) and serum creatinine (Scr) were measured. The renal morphologic changes were scored with Paller’s criterion on hematoxylin and eosin (HE) stained sections. The mRNA and protein expressions of HGF, BMP-7, TNF-α and IL-10 were detected by RT-PCR and immunohistochemistry method. RESULTS: After 4 d of cisplatin injection, the BUN and Scr values in MSC group were significantly lower than those in ARF group (P<0.01). After 7 d and 14 d, the values of BUN and Scr in MSC group were still lower than those in ARF group (P<0.01, P<0.05). The renal morphologic scores of MSC group were also lower than those of ARF group. After 7 d, the expressions of HGF, BMP-7 and IL-10 were higher in MSC group than those in ARF group, the expression of TNF-α in MSC group was lower than that in ARF group. CONCLUSION: MSC promotes the recovery of acute renal failure induced by cisplatin. The mechanism may partly depend on paracrine of growth factor and amelioration of inflammatory.     

Role of GPR40 mediates the effects of FFAs on lipoapoptosis in mouse β-cell line NIT-1

AIM: To evaluate the role of G protein-coupled receptor 40 (GPR40) mediates the effects of free fatty acids (FFAs) on lipoapoptosis in mouse β-cell line NIT-1 and the mechanisms involved in this process.METHODS: NIT-1 cells were supplemented with palmitate (500 μmol/L) or oleate (500 μmol/L) for 48 h, then apoptosis of the cells was determined by the methods of Hoechst 33342, TUNEL and flow cytometry (Annexin V/PI). The small interfering RNA technique was used to inhibit the expression of GPR40 in NIT-1 cell. The mock, control siRNA and GPR40 siRNA transfected cells were either supplemented with palmitate (500 μmol/L) or co-incubated with palmitate and oleate (500 μmol/L for each) for 48 h. The percentages of apoptotic cells were quantified. The expression of p-c-Jun, Bcl-2 and Bax were detected by Western blotting.RESULTS: Palmitate induced β cell lipoapoptosis, whereas oleate inhibited NIT-1 cells from palmitate-induced lipoapoptosis. No significant difference of the percentages of apoptotic cells was indicated among the mock, control siRNA and GPR40 siRNA transfected cells treated with palmitate (P>0.05). However, after co-incubated with palmitate and oleate (500 μmol/L for each) for 48 h, the percentage of apoptotic cells in GPR40 siRNA transfected cells was greater than that in mock (P<0.05), while the expression of p-c-Jun was decreased. The expressions of Bcl-2 and Bax were not affected.CONCLUSION: Palmitate induced β cell lipoapoptosis might not be mediated through GPR40, whereas oleate inhibits NIT-1 cells from palmitate-induced lipoapoptosis, which is mediated at least in part through GPR40, the change of c-Jun expression may play an role in this process, suggesting that GPR40 might be implicated in the control of β cell mass plasticity and GPR40 probably provides a link between obesity and type 2 diabetes.     

Overexpression of SSTR2 inhibits growth of SSTR2-positive tumors via multiple signaling pathways

AIM: To study the anti-proliferation effect of overexpressed SSTR2 in experimental cancer with different profiles of endogenous SSTRs expressions and the possible signaling pathways involved. METHODS: In the first experiment, the growth of the tumor xenografts of the inoculated capan-2 cells and A549 cells overexpressing SSTR2 or LacZ was investigated in nude mice. In the second experiment, the adenoviral vector expressing SSTR2 were introduced into experimental capan-2 xenografts by intratumoral injection. The growth inhibition of these experiment tumors was observed and the potential influences on different signaling pathways were analyzed by immunoassays. RESULTS: Overexpression of SSTR2 inhibited the growth of tumors with different profiles of endogenous SSTRs, including experimental capan-2 xenografts that had endogenous SSTR2 expression. Overexpression significantly affected a number of components in apoptotic pathway, MAPK pathway and angiogenesis. CONCLUSION: SSTR2 is a promising candidate for gene therapy in a wide spectrum of cancers.     

Role of A3 adenosine receptor on vascular reactivity of superior mesenteric artery in hemorrhagic shock in rat

AIM: To explore whether A3 adenosine receptor plays a role in the modulation of vascular reactivity after hemorrhagic shock in rat, and to find out the prospective drug target to restore the decreased vascular reactivity following hemorrhagic shock. METHODS: The hemorrhagic shock (40 mmHg) model was established in rat, and the reactivity of superior mesenteric artery (SMA) to norepinephrine (NE) was observed. A3AR expression at protein level and mRNA level were measured by Western blotting and RT-PCR respectively. RESULTS: The vascular reactivity of SMA to NE after hemorrhagic shock (40 mmHg) was decreased significantly in a biphasic response manner. The expression of A3AR mRNA in SMA after hemorrhagic shock decreased without significant difference. The expression of A3AR protein has a slight increase without statistical difference after 30 min of hemorrhagic shock and then has a significant decrease (especially at 2 h and 4 h after hemorrhagic shock). The usage of IB-MECA, a selective A3AR agonist, significantly increased the responsiveness of SMA to NE in hemorrhagic shock in rat. MRS1523, the selective A3AR antagonist, significantly abolished the restoration of the vascular reactivity to NE by IB-MECA in hemorrhagic shock in rat. CONCLUSION: A3AR plays a role in the modulation of vascular responsiveness to NE in hemorrhagic shock in rat, and the selective agonist of A3AR could restore the reactivity of SMA to NE in hemorrhagic shock in rat.     

黄金树花器官发生及发育的形态观察

利用普通光学显微镜和扫描电镜技术对黄金树花器官的发生及发育过程进行了观察.结果显示:(1)黄金树花器官的形态发生及发育过程集中于3月下旬-5月上旬;(2)花原基分化形成花的整个过程符合一般的分化顺序:花萼原基-花冠原基-雄蕊原基-雌蕊原基,且各原基在分化顺序上存在交叉;(3)花药及胚珠的发育与花器官的形态发生之间有明显的连续性,当花蕾直径为3.0 mm左右时花粉母细胞及完整的花粉囊壁形成,直径达到3.5 mm左右时胚珠中出现孢原细胞的分化,它直接起大孢子母细胞的功能.     

Nontypeable haemophilus influenzae induces IL-8 expression in alveolar epithelial cells in a p38 MAPK and NF-κB dependent manner

AIM:To investigate the key molecular mechanism of inflammatory response in alveolar epithelial cells induced by nontypeable haemophilus influenzae (NTHi). METHODS: A549 cells were co-cultured with NTHi (multiplicity of infection, MOI: 10) and harvested 15 min and 30 min after stimulation. The phosphorylation of p38 mitogen activated protein kinase (p38 MAPK) in A549 cells was detected by Western blotting. The intracellular expression of nuclear factor-κB (NF-κB) p65 was examined by flow cytometry 4 h after stimulation. A549 cells were preincubated with p38 inhibitor (SB203580) or NF-κB inhibitor (PDTC) for 1 h and then stimulated with NTHi for 24 h. The level of interleukin 8 (IL-8) in the supernatants was determined by enzyme-linked immunosorbent assay (ELISA). RESULTS: The phosphorylation of p38 MAPK was rapidly induced by NTHi stimulation. The expression of NF-κB p65 in A549 cells after NTHi stimulation was significantly up-regulated compared with control group (P<0.05). The level of IL-8 in the supernatants was increased 24 h after bacterial stimulation compared with control group (P<0.05). Blockage of p38 MAPK or NF-κB remarkably decreased IL-8 secretion in A549 cells (P<0.05). CONCLUSION:NTHi induces inflammatory response in alveolar epithelial cells in a p38 MAPK and NF-κB dependent manner.     

Relationship between endothelial dysfunction and renin-angiotensin-aldosterone system under the condition of fatigue stress and effect of herbs to dredge collaterals

AIM: To investigate the mechanisms and relationship among endothelial dysfunction, renin-angiotensin-aldosterone system (RAAS) and family of interleukins under the condition of excessive fatigue, by using the rats with fatigue stress. METHODS: Healthy male Wistar rats were randomly divided into control group, homocysteine (HCY) group, fatigue stress group, renshen group, shuangshen group and tongxinluo group. Radioimmunoassay was carried out to detect plasma renin activity (PRA), angiotensin II (AngⅡ), aldosterone (ALD), endothelin (ET), thromboxane-2 (TXA2), prostaglandin I2 (PGI2) in plasma and interleukin-1β, 2, 6, 10 (IL -1β, IL -2, IL -6) in sera. ELISA was used to detect NE and IL -10. The content of nitric oxide (NO) in sera was also detected. The bioinformatical analysis was used to determine the relationship between RAAS and endothelial dysfunction. RESULTS: Compared with control group, the levels of ET-1 and TXA2 in fatigue stress group were significantly increased (P<0.01, P<0.05), but the content of PGI2 and NO was significantly decreased (P<0.01, P<0.01). Compared with control group, the renin activity in plasma of animals in fatigue stress group was significantly decreased (P<0.01), the AngⅡ, IL -1β, IL -6 level was significantly increased compared with the control group (P<0.01, P<0.01), and was significantly increased compared with the HCY group (P<0.05, P<0.01, P<0.01). The NE level showed the tendency of decrease in different degree. After the intervention of three kinds of herbs to dredge collaterals, the ET-1, AngⅡ, IL -1β, IL-6 level in plasma was decreased in different degree (P<0.05, P<0.01, P<0.01), and at the same time, the contents of NO and NE level were significantly increased (P<0.05). The ALD level in tongxinluo groups was apparently higher than that in control group and the fatigue stress group (P<0.05). The bioinformatics analysis showed that Ang II and ET, IL-1; PGI2 and ALD; NO and ALD composed of three subsystems and interrelated according to the principle of optimality of complex system, and gradual change regulation was also observed in fatigue stress group. However, in control group, HCY group and tongxinluo group, the same interrelation among subsystems was not existed. CONCLUSION: In a state of long-term excessive fatigue, vascular endothelial dysfunction may be induced, and is related with renin-angiotensin-aldosterone system imbalance and serious turbulence of the autonomic dysfunction. Herbs to dredge collaterals could improve it significantly. The results suggest that bearing excessive fatigue and pressure in long-term may be the potential risk factors to induce vascular endothelial dysfunction and further result in cardio-cerebro vascular diseases, Tongluo therapy may be one of the useful ways to prevent such diseases.     

Mechanism of RhoA on vascular reactivity following hemorrhagic shock in rats

AIM: To observe the mechanisms of RhoA on vascular reactivity following hemorrhagic shock (HS) in rats. METHODS: The superior mesenteric artery (SMA) in rats subjected to hemorrhagic shock was adopted to assay the vascular reactivity via observing the contraction initiated by norepinephrine (NE) with isolated organ perfusion system. Meanwhile, the effects of Rho kinase, myosin light chain phosphatase (MLCP), myosin light chain kinase (MLCK) on RhoA regulating vascular reactivity were observed. The effects of RhoA agonist U-46619 and inhibitor C3 enzyme on the activities of Rho kianse, MLCP, MLCK and phosphorylation of MLC20 in the vascular smooth muscle cells (VSMC) with hypoxia were also measured. RESULTS: As compared to control group, the cumulative dose-response curves of SMA to NE at 2 h after shock shifted to the right, the maximal contractions (Emax) of NE was significantly decreased. RhoA agonist U-46619 increased the vascular reactivity in the late period of shock. C3 enzyme abolished U-46619 induced the increase in the contractile response of SMA to NE. Rho kinase inhibitor Y-27632 decreased U-46619-induced the increase in the vascular reactivity, MLCP inhibitor calyculin further promoted the increase in the vascular reactivity. However, MLCK inhibitor had no effect on the U-46619-induced change of vascular reactivity. After hypoxia, the activities of Rho kinase and MLCK, and the level of MLC20 phosphorylation were decreased, MLCP activity was increased. RhoA agonist U-46619 increased the activity of Rho kinase and phosphorylation of MLC20, decreased the activity of MLCP, but had no effects on MLCK activity. CONCLUSION: RhoA plays an important role in the regulation of vascular reactivity following shock. The mechanism is closely related to regulating the activities of Rho kinase and MLCP, and increasing the phosphorylation of MLC20 in VSMC.     

Adrenomedullin gene transfection enhances the therapeutic effects of transplantation of bone marrow mesenchymal stem cells on cardiac function and ventricle remodeling in acute myocardial infarction rats

AIM: To investigate adrenomedullin gene transfection enhances the therapeutic effects of homogeneous transplantation of bone marrow mesenchymal stem cells (MSCs) on cardiac function and ventricle remodeling in acute myocardial infarction rats. METHODS: MSCs were isolated and expanded using the preplating method. The infection efficiency of adenovirus vector to MSCs was tested by X-gal staining. Ad-ADM expression in MSCs and its secretion in culture medium were measured by ELISA. The left anterior descending branch of rats was ligated to establish a myocardial infarction model. The MSCs were labeled by DAPI, and were directly implanted into the acute infarct site via focal injection. Four weeks later, cardiac function was evaluated using physiological recorder. Hearts were harvested and sliced to be analyzed by immunohistochemistry (factor Ⅷ and ADM) and the DAPI-labeled cells were identified. Sirius red staining was used to identify interstitial collagen on slides. Analysis of collagen type I and III was performed using a polarized filter on sections stained for collagen with Sirius red, and the ratio of collagen type I and III were detected. RESULTS: With X-gal staining, MSCs were effectively transfected by adenovirus in vitro. The transfection efficiency showed the dose-effect relationship with multiplicities of infection (MOI). When MOI was 150, the infection efficiency was 95.4%. The expression of ADM was traced in culture medium and expressed in the time-dependent manner. A maximum production of ADM was observed at 7 d after infection ［(26.53±1.42) ng/L vs (1.34±0.08) ng/L, P<0.05］, and ADM secretion reduced to normal level at 15 d ［(2.20±1.44) ng/L vs (1.52±0.33) ng/L, P>0.05］. DAPI-labeled MSCs transplantation was found in the hearts of the recipients. Immunohistochemical studies demonstrated that intense immunostaining for ADM was higher in Ad-ADM plus MSCs group, compared to other groups. Compared with control, MSCs transplantation significantly increased capillary density in infarct area (P<0.01). A combination of Ad-ADM trensfection and MSCs transplantation demonstrated a further increase in capillary density compared with Ad-ADM or MSCs alone. MSCs transplantation decreased the ratio of collagen type I and III, obviously improved the left ventricular functions. Furthermore the combination treatment resulted in further decrease in the ratio of collagen type I and III, and significantly improved the left ventricular functions. CONCLUSION: Ad-ADM transfection enhances the angiogenic potency of MSCs transplantation and decreases the ratio of collagen type I and III through increasing ADM expression in infarct area, thus contributes to reverse the ventricular remodeling and improves the cardiac function.     

中早熟甘蓝新品种东农611的选育

东农611是以3-1028为母本,以3-1030为父本配制而成的中早熟甘蓝一代杂种。叶球近圆形,外叶灰绿色,表面平滑,中肋细,叶球紧实度高,中心柱小于球高的1/2,叶质脆嫩,VC含量452.3 mg·kg-1,抗TuMV和黑腐病,单球质量2 kg左右,每667 m²产量5 000 kg左右,适于黑龙江、吉林、内蒙古东北部等地春、秋早熟栽培。