| Objectives:Aging is one of the important risk fectors for cardiovascular disease,but the changes in aging-related cardiac structure and its mechanisms are not fully understood.In this experiment,we constructed a natural aging mouse model to study the changes in cardiac structure of natural aging mice and explore its possible mechanisms.Methods:Ten 7-month-old C57BL/6 mice were routinely maintained for 15 months in the animal room of the 920th hospital of the Union Support Force.A aging mouse model(22 months old)was constructed and named as the elderly group(aged)Another 10 2-month-old C57BL/6 mice were used as a young control group(young).After the model was successfully constructed,the weight of the mice was routinely weighed and sacrificed.The weight and body weight of the heart samples were weighed,and the body weight ratio was calculated.Hematoxylin Eosin(HE)staining was used to observe the gross sections of myocardial tissue,inflammatory cell infiltration and myocardial necrosis and hypertrophy;Masson staining to observe the degree of myocardial fibrosis,fluorescence Tunel to detect the degree of myocardial apoptosis;ELISA method for detecting myocardial fibers Expression level of the relative faector type Ⅰ pre-faro collagen peptide and type Ⅲ procollagen aninso torminsal peptide.The nuclear factor-related factor-2 and heme oxygenase-1 in the myocardium were detected by qPCR and Western blot.Results:1.Compared with the young group,the mice in the aged group responded slowly,exercised decreased,and the fur is dull,the skin is alopecia,and the intake of drinking water is reduced;2.Compared with the young group,the weight and heart weight of the aged group were significantly increased(P<0.001),but there was no significant difference in heart weight ratio(P>0.05);3.myocardial HE staining results show:compared with the young group,the aged group of mice increased heart cavity,gap widening,fiber thickening,disordered arrangement;Masson staining showed increased myocardial collagen fibers in the aged group;fluorescent Tunel results There was no significant difference in myocardial apoptosis rate between the young group and the aged group(P>0.05);4.The results of ELISA showed that the expression levels of type Ⅰ and type Ⅲcollagen fibers in the aged group were higher than those in the young group,but there was no significant difference between them(P>0.05);5.qPCR and Western blot showed that the mRNA and protein expressions of Nrf2 and Ho-1 were found in the myocardium of the two groups,and the expression level of myocardium in the young group was significantly higher than that in the aged group(P<0.05);6.Correlation analysis showed that the protein expression levels of Nrf2 and Ho-1 in myocardial tissue were negatively correlated with the expression levels of type Ⅰ and type Ⅲ procollagen peptides(P<0.05).Conclusions:1.Aging can cause heart cavity enlargement,myocardial fiber thickening,disordered arrangement,myocardial gap widening,collagen deposition and other changes;ELISA detection found that myocardial collagen increased,suggesting that early heart aging in mice may be mainly related to fibrosis;2.Aging-related changes in cardiac structure The increase in myocardial collagen may be associated with decreased expression of Nrt2 and Ho-1,which may be involved in the aging of mouse hearts.Objectives:To explored the effects of UCMSCs transplantation and Klotho protein treatment on aging-related cardiac structural changes.Methods:Human UCMSCs were expanded in vitro,UCMSCs were identified by cell morphology and cell surfece antigen,and the 5th gereration UCMSCs were identified.32 22-month-old C57BL/6 mice were randomly divided into four groups,name ly:aged+UCMSCs group(on the basis of natural aging model,tail vein injection of mouse umbilical cord mesenchymal stem cells,1x105/0.5ml),aged+Klotho protein group(Klotho protein,10ug/kg in the tail vein based on natural aging model),aged+saline group(based on the natural aging model,the tail vein was injected with the same volume of normal saline),the aged group(without any treatment).Four weeks after transplantation of UCMSCs and Klotho protein treatment,the mice were weighed and sacrificed.The wet weight of the heart was weighed and the weight ratio of the heart was calculated.Hematoxylin Eosin(HE)staining to observe gross myocardial appearance,inflammatory cell infiltration and myocardial necrosis,hypertrophy;Masson staining observation Myocardial fibrosis and fluorescence TUNEL were used to detect the degree of myocardial apoptosis.The expression levels of type Ⅰ and Ⅲ procollagen peptides in myocardium were detected by ELISA.The nuclear foctor-related factor-2(Nrf2)in the myocardium was detected by qPCR and Western blot.And heme oxygenase-1(Ho-1)messenger RNA and protein levels;Results:1.Cell identification and culture results:UCMSCs cultured in adherent method were expanded into fusiform and polygonal shapes,and the cells grew vigorously.Flow cytometry showed that the 5th generatkion UCMSCs expressed CD73,CD90 and CD105,but did not express CD34,CD45.Comply with UCMSCs characteristics;2.Compared with the aged group and the aged+saline group,UCMSCs and Klotho were treated with thick coat and alopecia areata after 4 weeks of treatment.The activity of the mice increased,and the intake and drinking increased.3.There were no significant differences in body weight,cardiac wet weight and heart weight ratio between the aged group,the aged+saline group,the aged+UCMSCs group and the aged+Klotho group(P>0.05).4.HE staining,Masson staining and fluorescent Tunel staining showed that there was no significant difference in myocardial inflammatory cell infiltration,degeneration and necrosis,myocardial fibrosis and myocardial apoptosis in the four groups.5.ELISA results showed that compared with the aged group and the aged+saline group,the expression of type I and type III procollagen peptides in myo tissue of aged+UCMSCs group and aged+Klotho group was decreased,but the four groups were No significant difference(P>0.05);6.The results of qPCR and Western blot showed that the mRNA and protein expressions of Nrf2 and Ho-1 in myocardial tissue of aged+UCMSCs group and aged+Klotho group were significantly higher than those of age group and aged+saline group.(P<0.05).7.Correlation analysis showed that the protein expression of Nrf2 and Ho-1 in myocardial tissue was negatively correlated with the expression of type I and tytpe III procollagen peptides(P<0.05).Conclusions:1.UCMSCs and Klotho protein therapy can significantly increase the expression of Nrf2 and Ho-1 in aging myocardium,attenuate oxidative stress and improve the aging of heart at the molecular level.2.UCMSCs and Klotho protein treatment attenuated skin and hair aging. |
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