Objective:To investigate the possible mechanism of DiI labeled bone marrow mesenchymal stem cells (BMSCs) in contact co-cultured with neonatal rat cardiomyocytes (CMs) on polycaprolactone (PCL) film to make myocardial patch.Methods:BMSCs from Sprague Dawley rats (aged 5-6 weeks) were isolated, cultured, and characterized for surface marker expression using flow cytometry. CMs from 15 neonatal rats were isolated and cultured. After cultured for 3 generations, BMSCs were labeled with DiI dye. On PCL film, DiI labeled BMSCs were co-cultured with CMs as the experimental group, and CMs were replaced with the same amount of unlabeled BMSCs in the control group. After 24 h of co-culture, the cell growth was observed under fluorescence microscope and the co-culture was observed under scanning electron microscope. Immunofluorescence staining was performed after 7 days to detect myocardial markers, including cardiac troponin T (cTnT) and α-actinin. BMSC differentiation on the PCL film was observed under a fluorescence microscope. The differentiation efficiency of BMSCs into cardiomyoid cells was analyzed by flow cytometry on days 1 and 7 of co-culture. Intercellular dye transfer was observed by staining CMs with calcein and co-culturing them with DiI-labeled BMSCs on PCL film. The cells were stained with immunofluorescence to detect the expression of connexin 43 (Cx43) and observe the relationship between gap junction and contact co-culture.Results:Flow cytometry showed strong positivity for CD90 and CD44 and negativity for CD11b/c and CD45 on BMSCs. After 24 h of co-culture, DiI labeled BMSCs glowed red on the PCL film, while unlabeled CMs did not; the number of cells on PCL film was large and cell morphology appeared normal under scanning electron microscope. On the 7th day of co-culture, some DiI labeled BMSCs expressed cTnT and α-actinin. Flow cytometry showed a higher differentiation rate of stem cells in the experimental group on day 7 compared to the control group ((20.12±0.15)% vs. (3.49±0.20)%, P<0.05). From the second day of co-culture, some BMSCs exhibited green dot fluorescence in Cx43 immunofluorescence staining; and by the third day, dye transfer test showed green fluorescence emission from some BMSCs. Conclusion:Contact co-culture of DiI labeled BMSCs and CMs on PCL film can make myocardial patch. The mechanism of contact co-culture promoting the differentiation and formation of myocardial patch may be associated with gap junctions and intercellular signal pathways mediated by gap junctions.

Objective    To provide experimental data and theoretical support for further studying the maturity of cardiac patches in other in vitro experiments and the safety in other in vivo animal experiments, through standard chemically defined and small molecule-based induction protocol (CDM3) for promoting the differentiation of human induced pluripotent stem cells (hiPSCs) into myocardium, and preliminarily preparing cardiac patches. Methods    After resuscitation, culture and identification of hiPSCs, they were inoculated on the matrigel-coated polycaprolactone (PCL). After 24 hours, the cell growth was observed by DAPI fluorescence under a fluorescence microscope, and the stemness of hiPSCs was identified by OCT4 fluorescence. After fixation, electron microscope scanning was performed to observe the cell morphology on the surface of the patch. On the 1st, 3rd, 5th, and 7th days of culture, the cell viability was determined by CCK-8 method, and the growth curve was drawn to observe the cell growth and proliferation. After co-cultured with matrigel-coated PCL for 24 hours, hiPSCs were divided into a control group and a CDM3 group, and continued to culture for 6 days. On the 8th day, the cell growth was observed by DAPI fluorescence under a fluorescence microscope, and hiPSCs stemness was identified by OCT4 fluorescence, and cTnT and α-actin for cardiomyocyte marker identification. Results    Immunofluorescence of hiPSCs co-cultured with matrigel-coated PCL for 24 hours showed that OCT4 emitted green fluorescence, and hiPSCs remained stemness on matrigel-coated PCL scaffolds. DAPI emitted blue fluorescence: cells grew clonally with uniform cell morphology. Scanning electron microscope showed that hiPSCs adhered and grew on matrigel-coated PCL, the cell outline was clearly visible, and the morphology was normal. The cell viability assay by CCK-8 method showed that hiPSCs proliferated and grew on PCL scaffolds coated with matrigel. After 6 days of culture in the control group and the CDM3 group, immunofluorescence showed that the hiPSCs in the control group highly expressed the stem cell stemness marker OCT4, but did not express the cardiac markers cTnT and α-actin. The CDM3 group obviously expressed the cardiac markers cTnT and α-actin, but did not express the stem cell stemness marker OCT4. Conclusion    hiPSCs can proliferate and grow on matrigel-coated PCL. Under the influence of CDM3, hiPSCs can be differentiated into cardiomyocyte-like cells, and the preliminary preparation of cardiac patch can provide a better treatment method for further clinical treatment of cardiac infarction.

Objective     To analyze the efficacy of off-pump coronary artery bypass grafting (OPCABG) in elderly patients with coronary artery disease complicated with moderate ischemic mitral regurgitation. Methods     The clinical data of patients aged≥70 years with coronary artery disease complicated with moderate mitral regurgitation, and undergoing OPCABG from January 2009 to January 2020 in Beijing Anzhen Hospital were retrospectively analyzed. The echocardiographic indicators of the patients were compared preoperatively, postoperatively before discharge and during the follow-up. Results     Finally 239 patients were enrolled. There were 136 males and 103 females, aged 74.1±3.2 years. Before postoperative discharge, 49 (20.5%) patients had no mitral regurgitation, 144 (60.3%) mild regurgitation, 46 (19.2%) moderate regurgitation, and 0 severe regurgitation. The area of mitral regurgitation was significantly improved (2.5±1.8 cm2 vs. 5.6±1.0 cm2, P<0.001). There were 10 (4.2%) patients of hospital death, 23 (9.6%) of low cardiac output, 3 (1.3%) of myocardial infarction, and 8 (3.3%) of nervous system injury after operation. As a result, 208 (90.8%) patients were followed up and the mean follow-up time was 3.4 years (range 1-9 years). The cumulative survival rates at postoperative 2, 4, 6, and 8 years were 95.8%, 88.0%, 78.4%, and 73.1%, respectively. Postoperative follow-up showed significant improvements compared with those before surgery in the area of mitral regurgitation, left ventricular ejection fraction, left ventricular end-diastolic and left ventricular end-systolic diameters (all P<0.05). Duirng the follow-up, the major adverse cardiac and cerebrovascular events were all cause death in 22 (10.6%) patients, including cardiac death in 17 (8.2%) patients, myocardial infarction in 7 (3.4%) patients, heart failure in 24 (11.5%) patients, cerebrovascular events in 11 (5.3%) patients, re-hospitalization due to heart disease in 23 (11.1%) patients, and none of the patients with myocardial infarction were revascularized. Conclusion     The mid- and long-term outcomes of OPCABG in the treatment for elderly patients with coronary artery disease complicated with moderate ischemic mitral regurgitation is good.

Objective:To analysis the risk factors of early death and long-term outcomes of myocardial infarction complicated with ventricular septal rupture.Methods:A total of 135 patients with myocardial infarction complicated with ventricular septal rupture in Beijing Anzhen Hospital from January 2008 to December 2020 were retrospectively analyzed.According to the survival or death within 30 days after ventricular septal rupture, the patients were divided into the early survival group(n=71)and the early death group(n=64). The clinical characteristics of the two groups were observed, and the risk factors for early death group were analyzed.The long-term outcomes of the surgery group(n=69)and the non-surgery group(n=66)was analyzed.Results:The early mortality rate of patients with myocardial infarction complicated with ventricular septal rupture was 47.4%(64/135). Univariate analysis showed that age, sex, white blood cell count, platelet count, C-reactive protein level, left ventricular end-diastolic diameter, abnormal liver function, pulmonary infection, no surgery repair and Killip grade ≥3 were associated with early death as compared with the early survival group(all P<0.05). Multivariate regression analysis showed that no surgery repair( OR=16.103, 95% CI: 4.400-58.930, P<0.001)and Killip≥3 grade( OR=9.014, 95% CI: 2.506-32.428, P=0.001)and abnormal liver function( OR=5.171, 95% CI: 1.388-19.264, P=0.014)were independent risk factors for early death in patients with myocardial infarction complicated with ventricular septal rupture.During follow-up of 1.0 to 11.8(median 3.2)years, the 2-year and 10-year cumulative survival rates were significantly higher in the surgery group than in the non-surgery group(76.7% vs.16.7%, P<0.001; 73.1% vs.16.7%, P<0.001). Conclusions:No surgical repair, Killip grade ≥3 and abnormal liver function are independent risk factors for early death in patients with myocardial infarction complicated with ventricular septal rupture.The long-term outcomes of surgical treatment for myocardial infarction complicated with ventricular septal rupture is good.

Objective:To compare the efficacy of off-pump coronary artery bypass grafting (CABG) or CABG plus mitral valve plasty (MVP) in patients with coronary heart disease complicated with moderate ischemic mitral insufficiency.Methods:The clinical data of 1 050 patients with coronary heart disease complicated with moderate ischemic mitral insufficiency who underwent surgical procedures from January 2009 to December 2020 were analyzed retrospectively. There were 733 males and 317 females, aging (63.3±9.0) years (range: 31 to 83 years). Patients were divided into CABG+MVP group and CABG group according to surgical methods, and the two groups of patients were matched for 1∶4 by the propensity score matching method. There were 107 patients in the CABG+MVP group and 406 patients in the CABG group after matching. The t test, Mann-Whitney U test, χ 2 test, Fisher′s exact probability method and repeated measures anova were used to compare the surgical outcomes and overall survival in the two groups. Results:There were no significant differences in perioperative death and postoperative complications between the two groups (all P>0.05). Compared with CABG group, CABG+MVP group had longer operation time ((5.6±1.2) hours vs. (4.2±1.0) hours, t=11.528, P<0.01), ICU stay( M(IQR))(43.0(47.3) hours vs. 25.0(33.6) hours, Z=2.483, P=0.013), and postoperative hospital stay (8(4) days vs. 7(5) days, Z=2.143, P=0.032). The amount of erythrocyte and platelet used in CABG+MVP group was significantly increased (2.0(6.5) U vs. 0(2.0) U, Z=7.084, P<0.01; 0(0.5) U vs. 0(0) U, Z=5.210, P<0.01). A total of 463 cases (93.9%) were followed up. Median follow-up was 32(31) months (range: 3 to 105 months). There was no significant difference in overall survival and no major adverse cardic and cerebrovascular events survival between CABG group and CABG+MVP group ( P=0.196, P=0.305). Echocardiography showed that there was no significant difference in ejection fraction left ventricular end-diastolic diameter between the two groups ( F=0.322, P=0.571; F=0.681, P=0.410). However, CABG+MVP improved mitral regurgitation better than CABG ( F=160.222, P<0.01). Conclusions:For patients with coronary heart disease with moderate ischemic mitral insufficiency, the rates of all-cause mortality and major adverse cardiac and cerebrovascular events are similar between the two surgeries. Although CABG+MVP improves mitral regurgitation better than CABG, it increases the duration of surgery, ICU stay, postoperative hospital stay, and blood transfusion requirement.

Objective:To compare the efficacy of off-pump coronary artery bypass grafting (CABG) or CABG plus mitral valve plasty (MVP) in patients with coronary heart disease complicated with moderate ischemic mitral insufficiency.Methods:The clinical data of 1 050 patients with coronary heart disease complicated with moderate ischemic mitral insufficiency who underwent surgical procedures from January 2009 to December 2020 were analyzed retrospectively. There were 733 males and 317 females, aging (63.3±9.0) years (range: 31 to 83 years). Patients were divided into CABG+MVP group and CABG group according to surgical methods, and the two groups of patients were matched for 1∶4 by the propensity score matching method. There were 107 patients in the CABG+MVP group and 406 patients in the CABG group after matching. The t test, Mann-Whitney U test, χ 2 test, Fisher′s exact probability method and repeated measures anova were used to compare the surgical outcomes and overall survival in the two groups. Results:There were no significant differences in perioperative death and postoperative complications between the two groups (all P>0.05). Compared with CABG group, CABG+MVP group had longer operation time ((5.6±1.2) hours vs. (4.2±1.0) hours, t=11.528, P<0.01), ICU stay( M(IQR))(43.0(47.3) hours vs. 25.0(33.6) hours, Z=2.483, P=0.013), and postoperative hospital stay (8(4) days vs. 7(5) days, Z=2.143, P=0.032). The amount of erythrocyte and platelet used in CABG+MVP group was significantly increased (2.0(6.5) U vs. 0(2.0) U, Z=7.084, P<0.01; 0(0.5) U vs. 0(0) U, Z=5.210, P<0.01). A total of 463 cases (93.9%) were followed up. Median follow-up was 32(31) months (range: 3 to 105 months). There was no significant difference in overall survival and no major adverse cardic and cerebrovascular events survival between CABG group and CABG+MVP group ( P=0.196, P=0.305). Echocardiography showed that there was no significant difference in ejection fraction left ventricular end-diastolic diameter between the two groups ( F=0.322, P=0.571; F=0.681, P=0.410). However, CABG+MVP improved mitral regurgitation better than CABG ( F=160.222, P<0.01). Conclusions:For patients with coronary heart disease with moderate ischemic mitral insufficiency, the rates of all-cause mortality and major adverse cardiac and cerebrovascular events are similar between the two surgeries. Although CABG+MVP improves mitral regurgitation better than CABG, it increases the duration of surgery, ICU stay, postoperative hospital stay, and blood transfusion requirement.

Objective: To investigate the clinical efficacy of reduced port laparoscopic radical resection on colorectal cancer. Methods: Clinical data of 46 patients with colorectal cancer undergoing reduced port laparoscopic radical resection were retrospectively analyzed. Results: All of 46 patients underwent laparoscopic surgery, with an average operation time of 206 minutes, an average intraoperative bleeding of 56 ml, an average number of lymph nodes removement of 12/case (ranged from 6 to 21). One case had incision infection, 2 cases had anastomotic leakage, and they all recovered and discharged after treatment. Conclusions Reduced port laparoscopic radical resection of colorectal cancer is safe and feasible, reduces labor costs, and has a good clinical efficacy, which is worthy of clinical promotion.

Objective To investigate the effects of hypoxic three-dimensional culture microenvironment on the proliferation of bone marrow mesenchymal stem cells and its mechanism. Methods P5 generation mouse bone marrow mesenchymal stem cells and P (3HB-co-4HB) were co-cultured under normoxic three-dimensional (20%) and hypoxic three-dimensional microenvironment (4%) respectively. After 24 hours, the proliferation of the two groups was determined by CCK-8 method. The expression of HIF-1α gene was detected by real-time quantitative PCR after 12 hours. Western blotting was used to detect the expression of HIF-1α protein after 24 hours. Results After 24 hours, the CCK-8 method showed that the OD value of the hypoxia group was significantly higher than that of the normoxia group (0.455±0.027 vs. 0.352±0.090, n=12, P<0.05). After 12 hours of hypoxic culture, the expression level of HIF-1α mRNA in the hypoxia group was significantly higher than that in the normoxia group (P<0.05). Compared with the normoxia group (0.47± 0.05), the relative expression level of HIF-1α protein in the hypoxia group (0.63±0.06) significantly increased in the Western blotting after 24 hours (n=3, P<0.05). Conclusion The hypoxic three-dimensional microenvironment can promote the proliferation of bone marrow mesenchymal stem cells, which may be related to the activation of HIF-1α signaling pathway.

Microenvironment has a major impact on stem-cell differentiation. Stem-cell differentiation achieved by controlling the microenvironment is more similar to physiological characteristic than the traditional chemical induction, which will undoubtedly have a broader clinical application prospect. The impact of microenvironment on stem cell to cardiomyocyte differentiation is reviewed in this paper.