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Objective:To evaluate the effects of human umbilical cord mesenchymal stem cells-derived exosomes (hUCMSCs-ex) injection on cardiac function and myocardial fibrosis in dilated cardiomyopathy (DCM) rats induced by Adriamycin(ADR).Methods:One hundred male SD rats were randomly divided into the normal group (20 rats) and the DCM group (80 rats). The rats in DCM group were treated with ADR by intravenous injection to induce DCM.DCM rats were randomly divided equally into DCM group, low-dose group, medium-dose group and high-dose group which were received intravenous injection 1 mL/kg Dulbecco′s modified eagle medium(DMEM), 20 μg/kg, 100 μg/kg and 250 μg/kg exosomes.After modeling, 10 rats in normal group and 30 rats in DCM group were randomly selected to receive echocardiography to evaluate the cardiac function.After exosomes treatment, 10 rats were randomly selected form each group for echocardiography to evaluate the cardiac function.The morphological changes in myocardial cells were observed by using Masson staining in each group; Western blot detection between groups of rats was used to analyze the expression of myocardial collagen Ⅰ type(COLⅠ), Smad2 and alpha smooth muscle actin (α-SMA).Results:Left ventricular ejection fraction(LVEF) and left ventricular fraction shortening (LVFS)in the DCM group [(64.30±3.51)% and (38.70±2.85)%] were significantly lower than those of the normal group [(78.80±1.52)% and (50.60±1.50)%], and the differences were statistically significant ( t=20.518, 22.311, all P<0.01). The left ventricular end-diastolic diameter(LVEDD) and left ventricular end-systolic diameter (LVESD) [(4.62±0.13) mm and (3.40±0.12) mm] of the DCM group were significantly higher than those of the normal group[(3.29±0.24) mm and (3.16±0.33) mm], and the differences were statistically significant( t=2.854, 3.800, all P<0.01). After exosomes treatment, LVEF[(84.3±2.6)% and (83.4±3.2)%] in the medium-dose and high-dose groups were significantly higher than that in the DCM group [(79.2±2.4)%], and the diffe-rences were statistically significant(all P<0.01). Masson staining found that collagen fibers were less in exosomes treating group than those in the DCM group; Western blot test showed that high-dose exosomes can reduce the expression of α-SMA and Smad2, high-dose and low-dose exosomes can both significantly reduce the expression of COLⅠ. Conclusions:It suggests that exosomes intravenous injection from hUCMSCs-ex can significantly improve myocardial fibrosis in DCM rats induced by ADR and cardiac function.
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Objective To evaluate the effect of human umbilical cord mesenchymal stem cells (hUCMSCs) treatment through intramuscular administration on the heart function and angiogenesis of the myocardium in dilated car-diomyopathy (DCM)rats induced by Adriamycin(ADR). Methods One hundred male SD rats were randomly divided into the normal group and the DCM group. Rats in the DCM group were treated with ADR by intraperitoneal injection of 2. 0 mg/ kg dose per week for 8 weeks in order to induce DCM. Sixty modeled surviving rats with DCM were randomly divided equally into 3 groups,and they were treated with hUCMSCs or DMEM by intramuscular injection. Rats in the DMEM group (20 cases)received intramuscular infusion 2 mL DMEM alone;rats in the low - dose group (20 cases) underwent intramuscular infusion of 1 × 106 hUCMSCs/ 2 mL in DMEM;rats in the high dose group (20 cases)under-went intramuscular infusion of 10 × 106 hUCMSCs/ 2 mL in DMEM. Echocardiography and plasma brain natriuretic pep-tide(BNP)were used to assess cardiac function in modeled rats. The morphological changes in myocardial cells were observed by using HE and Masson staining after ADR injection stopped for one week. Four weeks after administration of hUCMSCs,echocardiography was performed to evaluate the cardiac function,and plasma BNP level was detected by en-zyme immunoassay kit. Western blot was used to analyze the expression of vascular endothelial growth factor(VEGF)in myocardium of rats in each group. Myocardial microvessel density was detected by using anti - CD34 monoclonal antibody and transmission electron microscopy (TEM)were performed to observe the ultrastructure of microvessel. Results Left ventricular ejection (LVEF)and left ventricular fractional shortening (LVFS)in the DCM groups [(66. 17 ± 3. 54)％,(31. 33 ± 3. 20)％]were significantly decreased compared to those in the normal group [(77. 25 ± 3. 40)％,(41. 00 ± 2. 94)％],and the differences were statistically significant(t = 10. 620,10. 328,all P < 0. 05);the morphological changes in myocardial cells was observed by using HE and Masson staining. Pit - induced typical his-tological lesion of myocardial tissue was observed in the DCM group,such as congestion,edema,a disorganization of myocytes and focal necrosis and myocardial tissue with wispy,broad collagen fibers predominating in the matrix. Four weeks after administration of hUCMSCs,LVEF in the low dose group or the high dose group were significantly higher compared with those in the DMEM group[(72. 27 ± 2. 44)％ or (70. 92 ± 2. 68)％ vs. (62. 89 ± 2. 54)％],and the differences were statistically significant(t = 2. 145,2. 131,all P < 0. 05);and LVFS were significantly higher compared with that in the DMEM group [(34. 96 ± 2. 08)％ or (33. 49 ± 2. 19)％ vs. (30. 98 ± 2. 22)％],and the differences were statistically significant (t = 2. 491,4. 086,all P < 0. 05). The plasma level of BNP was significantly declined in the hUCMSCs treated rats as compared to those before treatment [low dose group (352. 68 ± 41. 25)ng/ L vs. (202. 68 ± 20. 38)ng/ L,t = 2. 052,P < 0. 05;high dose group (355. 79 ± 48. 32)ng/ L vs. (193. 62 ± 15. 41)ng/ L,t = 2. 074,P < 0. 05]. Quantitative analysis demonstrated that microvessel density was significantly hi-gher in low - dose and high - dose hUCMSCs treated DCM rats than that in the DMEM treated DCM rats [(84. 00 ± 19. 18)/ mm2 or (86. 67 ± 20. 88)/ mm2 vs. (27. 14 ± 13. 97)/ mm2 ,t = 2. 109,2. 101,all P < 0. 05];Western blot test showed that there had high expression of VEGF in myocardium and TEM in the high dose group,and vessel injury in DMEM treated rats were more serious than that of hUCMSCs treated rats. Conclusion It suggests that hUCMSCs in-tramuscular injection may improve heart function and angiogenesis of myocardium in DCM rats induced by adriamycin.
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Objective To explore the effects of intramuscular injection human umbilical cord mesenchymal stem cells(hUCMSC)intramuscular injection on the cardiac function and myocardial ultrastructure in rats with Adria-mycin -induced dilated cardiomyopathy (DCM)rats.Methods One hundred and sixty rats were randomly divided in-to a normal group (20 cases)and DCMgroups (1 40 cases),rats in DCMgroups receiving Adriamycin (2 mg/kg)in-traperitoneally once a week for 8 weeks to establish DCM models.The DCM rats were randomly divided into a model control group (served as model group),the supernatant of hUCMSC group (served as supernatant group),the low -dose hUCMSC group(served as low -dose group),the medial -dose hUCMSC group(served as medial -dose group), and the high -dose hUCMSC group(served as high -dose group).Echocardiography was performed to evaluate the car-diac function,plasma brain natriuretic peptide (BNP)level and serum cardiac troponin I (cTnI)level were detected by enzyme -linked immunosorbent assay kit;light microscope and transmission electron microscopy (TEM)were used to observe the ultrastructure of myocardium.Results Rats in the DCM group showed low spirit,declining food intake, progressive emaciation,slow growth,hair loss and ascites.After the intramuscular injection of hUCMSC,the above symp-toms of rats in the low -dose and the medial -dose hUCMSC groups were improved significantly.Before the administra-tion of hUCMSC,the left ventricular ejection fraction (LVEF)[(64.53 ±2.61 )%]and the left ventricular fractional shortening (LVFS)[(30.80 ±2.1 1 )%]were significantly decreased in the DCMgroup compared to those of the con-trol group[(79.67 ±3.02 )%,(43.08 ±3.1 5 )%,all P <0.01 ].After the administration of hUCMSC,LVEF [(75.5 ±7.4)%,(74.0 ±6.1 )%]and LVFS[(40.8 ±3.8)%,(40.2 ±5.0)%]were significantly increased in the low -dose and the medial -dose group compared with those of the model group [(65.8 ±4.5)%,(30.2 ± 2.9)%,all P <0.01 ].The concentration of plasma BNP level [(438.3 ±82.2)ng/L,(341 .7 ±68.9)ng/L]and serum cTnI level [(375.9 ±1 1 0.9)ng/L,(355.9 ±55.6)ng/L]were significantly decreased compared with those of the model group [(449.9 ±91 .8)ng/L,(425.9 ±42.6)ng/L,all P <0.05].The findings of HE staining showed that cardiomyocytes were orderly arranged,edema decreased and cell nucleus homogeneously stained in the low -dose and the medial -dose group.The outcomes of TEM revealed that the ultrastructure of cardiomyocytes was improved in the low -dose and the medial -dose group compared with that of model group,and the cardiomyocyte sarcolemma re-mained intact,and the swelling of mitochondria ameliorated and the cristae of mitochondria remained clear.Conclusions Intramuscular injection of hUCMSC could significantly increase LVEF and LVFS in the Adriamycin -induced DCM rats,and decrease the plasma BNP levels and the serum cTnI levels,attenuate the myocardial pathological damage and improve myocardial ultrastructure.
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BACKGROUND:To date, it is stil unclear whether the intramuscular injection of heterogeneous umbilical cord mesenchymal stem cel s (UC-MSC) can cause cardiac ectopic pathological angiogenesis as wel as increase col agen synthesis to promote myocardial fibrosis. OBJECTIVE:To explore the effects of intramuscular injection of human UC-MSCs on myocardial micrangium and col agen expression in normal Wistar rats. METHODS:After 2 weeks of feeding, 60 male SPF Wistar rats were randomly assigned to receive intramuscular injection of PBS (normal group), DMEM (culture medium group), human UC-MSCs supernatant (supernatant group), 0.25×105, 1.0×105, 4.0×105 human UC-MSCs (low-, moderate-and high-dose groups), respectively (n=10 per group). Al the rats were subjected to second injection (same dose) at 4 weeks after first intramuscular injection. Then, the rats were kil ed under anesthesia at 4 weeks after second injection, to take heart tissues from the left ventricle for pathological observation, immunohistochemical examination and Masson staining. RESULTS AND CONCLUSION:No alteration of the response, activity, victualage, faeces, weight growth, and fur was found, and there was no death in rats during the experiment. Al the rats had no symptoms of molt, inflammation, skin ulcer, scleroma. Strong positive expression of CD34 for the micrangium in the myocardial tissue was observed, and positive expression of the col agen in the myocardial tissue observed by Masson staining. There were no significant differences in the microvessel density and col agen expression in the myocardium among the groups (F=0.110 and 0.585, P>0.05). To conclude, hUC-MSCs or its supernatant via intramuscular injection has no effect on the micrangium and col agen expression in normal rats.
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Objective The 3243A ＞ G mutation in mitochondrial DNA is a common cause of the classical mitochondrial diseases characterized by neuro-muscular disorders.This study reports a rare case with the main manifestations of mitochondrial disease in children of mitochondrial cardiomyopathy and respiratory muscle damage.Methods The clinical characteristics,diagnosis and treatment,biochemical,pathological and genetic features of a 10-year-old girl were studied.Results The girl was admitted because of heart failure and respiratory failure at the age of 10.Ragged red fibers in skeletal muscles had been noticed.On her mitochondrial gene,3243A ＞ G mutation,Leu tRNA (UUR),was detected.The mutation rate in the peripheral blood cells was 94％.After the treatment with a high dose of creatine phosphate sodium,coenzyme Q10 and L-carnitine with assisted ventilation,the patient improved rapidly.The child was followed up for 2 years without recurrence.Meanwhile the growth,development and daily life were normal.Conclusions Cardiac and respiratory muscle impairments that appeared at the same time as the first manifestations of the children's mitochondrial disease is not common,and it is rare to have cardiomyopathy based mitochondrial gene 3243A ＞ G mutation is seldom seen clinically.Skeletal muscle biopsy and genetic test is the key for accurate diagnosis.Improving mitochondrial metabolism and assisted ventilation appear to be helpful treatments.
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Objective To investigate the immunological pathogenesis of Kawasaki disease( KD)through examination of changes in the expression of suppressors of cytokine signaling 1(SOCS1)and SOCS3,helper T cells and CD4 + CD25 + regulatory T cells(CD4 + CD25 + Treg)in peripheral blood from children with acute KD. Methods Six-teen children[10 boys,6 girls,aged 1 - 2 years old,averaged(1. 6 ± 0. 3)years old]in the acute phase of KD(KD group),16 children[9 boys,7 girls,aged 1 - 3 years old,averaged(1. 5 ± 1. 1)years old]with pneumonia(pneumo-nia group)and 8 normal children[5 boys,3 girls,aged 1 - 5 years old,averaged(2. 0 ± 1. 1)years old]of the same age(normal control group)from the Affiliated Hospital of Qingdao University who were admitted from October 2012 to March 2013 were recruited. The mRNA levels of SOCS1 and SOCS3 in the T cells from peripheral blood were examined by way of reverse transcription - polymerase chain reaction(RT - PCR). Interferon - γ( IFN - γ),interleukin - 4 (IL - 4)and CD4 + CD25 + Treg were quantified by means of fluorescence activated cell sorting(FACS). Results The expressions of SOCS1 and SOCS3,the percentage of IL - 4 T cells observed in the peripheral blood of the pneumonia group were similar to the normal control group(P ﹥ 0. 05),but significantly decreased in the percentage of INF - γ and the level of CD4 + CD25 + Treg(t = 3. 71,12. 81,all P ﹤ 0. 05). Compared to the normal control group and the pneumo-nia group,the expressions of SOCS1 and SOCS3,the percentage of INF - γ and IL - 4 T cells decreased significantly in the peripheral blood of the KD group(t = 2. 27,4. 48,17. 64,2. 73,2. 74,1. 25,2. 36,2. 59,all P ﹤0. 05 ). On the other hand,the level of CD4 + CD25 + Treg in the peripheral blood of the KD group was markedly lower than that in the normal control group(t =7. 70,P ﹤0. 05),but similar to the pneumonia group(P ﹥0. 05). Conclusions The function of helper T cells is inhibited in acute KD. The CD4 + CD25 + Treg may be involved in the immunological pathogenesis of KD.
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BACKGROUND:Studies have shown that intramuscular transplantation of xenogeneic umbilical cord mesenchymal stem cel s in a certain dose range is safe and reliable, and it also confirm that this approach is equal y safe and effective for heart failure in rats with dilated cardiomyopathy. OBJECTIVE:To explore the effect of human umbilical cord mesenchymal stem cel s through intramuscular injection on the cytokine expression in adriamycin-induced dilated cardiomyopathy (DCM) rats. METHODS:Total y 160 rats were randomly divided into control group (n=20) and DCM group (n=140). Rats in the DCM group were administered adriamycin intraperitoneal y to establish DCM model. The DCM rats were randomly subdivided into model control group (served as model group), cel supernatant group, the low-dose mesenchymal stem cel group (served as low-dose group), the middle-dose mesenchymal stem cel group (served as middle-dose group), and the high-dose mesenchymal stem cel s group (served as high-dose group). Secondary injection was performed at 4 weeks after first injection. RESULTS AND CONCLUSION:The ELISA test showed that the serum levels of vascular endothelial growth factor (VEGF), hepatocyte growth factor (HGF), leukemia inhibitor factor (LIF) and granulocyte macrophage colony stimulating factor (GM-CSF) were higher in the model group than the control group before and after intramuscular injection (P0.05). Both the immunohistochemical and RT-PCR results showed that the expressions of insulin-like growth factor-1, VEGF and HGF were increased in al the DCM rats as compared with the control group, which were increased most in the middle-dose group. These findings indicate that low-dose and middle-dose human umbilical cord mesenchymal stem cel s intramuscular injection can increase the serum levels of HGF, LIF, GM-CSF, VEGF and the expressions of IGF-1, HGF and VEGF in the myocardium of DCM rats.
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Objective To investigate the expressions of Livin ? and Livin ? in marrow mononuclear cells(MMNCs)of childhood acute leukemia(CAL)and explore the clinical significance.Methods Real-time quantitative PCR and Western blot were used to detect the mRNA and protein expressions of Livin ? and Livin ? in MMNCs of CAL,respectively.Results Both Livin? and Livin? expression rates and expression levels were higher in preliminary diagnosis group(n=51)of CAL,including acute lymphoblastic leukemia(n=39)and acute myeloid leukemia(n=12),compared with the control group(P5%)to induction chemotherapy of ALL than in the patients without chemotherapy(P