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Relevant research data shows that there is a certain degree of energy metabolism imbalance in highland residents. Protein phosphatase 4 (PP4) has been found as a new factor in the regulation of sugar and lipid metabolism. Here, we investigate the differential expression of PP4 at a simulated altitude of 4,500 m in the heart, lung, and brain tissues of rats. A hypoxic plateau rat model was established using an animal decompression chamber. A blood routine test was performed by an animal blood cell analyzer on rats cultured for different hypoxia periods at 4,500 m above sea level. Quantitative polymerase chain reaction (qPCR) and western blot were used to detect the changes of protein phosphatase 4 catalytic subunit (PP4C) gene and protein in heart, lung, and brain tissues. The PP4C gene with the highest expression level found in rats slowly entering the high altitude area (20 m-2200 m-7 d-4500 m-3 d) was about twice as high as the low elevation group (20 m above sea level). The simulated high-altitude hypoxia induced an increase of PP4C expression level in all tissues, and the expression in the lung tissue was twice as expressed as heart and brain tissue at high altitude (P < 0.05). These results suggest that the PP4 phosphatase complex is ubiquitously expressed in rat tissues and likely involved in adaptation to or disease associated with high-altitude hypoxia.
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Objective:To explore the basic biological characteristics of lncRNA B230352I09 and its role in the process of myocardial injury.Methods:We analyzed the biological characteristics of lncRNA B230352I09 on the UCSC website and predicted the possible binding protein of lncRNA B230352I09 by the catRAPID. Real-time fluorescence quantitative (RT) PCR method was applied to detect the expression of lncRNA B230352I09 in heart tissues at different time points (0, 1, 3, 7d) within 7 days after birth, the organs distribution and expression of lncRNA B230352I09 in neonatal mouse and the expression pattern of lncRNA B230352I09 in the heart of mice with myocardial injury. In addition, we constructed hypoxia model by culturing primary cardiomyocytes to detect the effect of lncRNA 230352I09 overexpression on hypoxic cardiomyocyte apoptosis by Hoechst staining kit, the effect of lncRNA B230352I09 overexpression on ROS content of hypoxic cardiomyocyte by DCFDA probe and changes in mitochondrial membrane potential of hypoxic cardiomyocytes by JC-1 Fluorescent probes.Results:Full-length of mouse B230352I09 was 663bp, located in the chr7:123031415-123066439 forward strand. RBBP6 gene was adjacent to B230352I09, which may be the target of lncRNA B230352I09 by catrapid prediction analysis. With the development of the heart, the expression level of lncRNA B230352I09 showed a gradual downward trend. The main expression organs of lncRNA B230352I09 in 1-day-old mice were heart, brain, kidney and liver. In heart tissue, lncRNA B230352I09 expression in non-cardiomyocytes was significantly less than in cardiomyocytes [ (1.0± 0.03) vs. (9.2± 3.29), P=0.013]. After myocardial injury, the expression level of lncRNA B230352I09 showed an increasing trend compared with the normal developing mice, but there was no statistical significance. Hoechst staining showed that lncRNA B230352I09 could inhibit the apoptosis of hypoxic cardiomyocytes. Detecting the content of ROS in cardiomyocytes showed that compared with the hypoxia group, the generation of ROS was significantly reduced in the lncRNA B230352I09 overexpression group ([(3.8±0.71) vs. (1.65±0.56), P=0.015]). JC-1 fluorescent probe was used to detect the mitochondrial membrane potential, and the results showed that the mitochondrial membrane potential of cardiomyocytes in the lncRNA B230352I09 overexpression group was significantly higher than that in the hypoxia group. Conclusions:In heart tissue, lncRNA B230352I09 was mainly expressed in cardiomyocytes. LncRNA B230352I09 has a protective effect in the process of myocardial injury in mice, mainly by inhibiting apoptosis of cardiomyocytes, reducing ROS production, and protecting mitochondrial membrane potential of cardiomyocytes.
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BACKGROUND: Bone marrow mesenchymal stem cells can be induced into myocardial tissue-like structure in vitro. Myocardial tissue lysates from different parts of the myocardium can be used to construct differential microenvironments. Few studies have investigated the targeted induction efficiency of bone marrow mesenchymal stem cells and the expression of related genes. OBJECTIVE: To investigate the effects of lysates from different parts of the myocardium on the differentiation of bone marrow mesenchymal stem cells into myocardial tissue-like structures and to assess the correlation between the expressions of related genes during induction and myocardial tissue engineering. METHODS: Passage 3 bone marrow mesenchymal stem cells from Sprague-Dawley rats were cultured. Whole heart tissue lysate, atrial muscle tissue lysate, and ventricular muscle tissue lysate were used to construct different microenvironments to induce bone marrow mesenchymal stem cells to differentiate into cardiomyocyte-like cells. The morphological changes and ultrastructure of cells were observed with an inverted phase contrast microscope and transmission electron microscopy. Expression of α-actin and cTnI was detected by immunofluorescence staining. qRT-PCR was used to detect the expression of upstream molecules ANP, HCN4 and downstream molecules MLC-2v in the cAMP/PKA signaling pathway after induction. RESULTS AND CONCLUSION: (1) Each induction cell group followed similar morphological changes: Cardiomyocyte-like cells were capable of autonomic beats, rhythmic contractions and relaxations; under the transmission electron microscope, there were a large number of arranged myofilaments; immunofluorescent staining showed positive expression of α-actin and cTnI. (2) The whole heart tissue lysate was able to induce the differentiation of bone marrow mesenchymal stem cells into rice grain-sized myocardial tissue-like structures with collagen fibers. (3) Atrial muscle tissue lysate could induce bone marrow mesenchymal stem cells to differentiate into atrial muscle-like cells. Autonomic beats appeared earlier, but the beat frequency and duration were shorter. ANP and HCN4 were highly expressed in atrial myocytes. (4) Ventricular muscle tissue lysate induced bone marrow mesenchymal stem cells to differentiate into ventricular muscle-like cells with no secretory granules, and MLC-2v was highly expressed in ventricular muscle-like cells. (5) To conclude, the whole heart tissue lysate can induce bone marrow mesenchymal stem cells to form myocardial tissue-like structures. Atrial muscle tissue lysate can induce bone marrow mesenchymal stem cells to differentiate into atrial muscle-like cells. Ventricular muscle tissue lysate can induce bone marrow mesenchymal stem cells to differentiate into ventricular muscle-like cells. By constructing a specific microenvironment, bone marrow mesenchymal stem cells can be differentiated to cardiomyocytes in different parts, providing laboratory data for the construction of myocardial tissue engineering.
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@#The engineered heart tissues (EHTs) is regarded as a hope for myocardial repair and regeneration. But a series of “bottleneck” problems, such as vascularization, hinder their clinical translation. This review focuses on the strategies to vascularization of EHTs and encourages the emergence of novel EHTs that can meet clinic requirement properly.
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Objective To observe the expression changes in microRNA (miR)-379 in the developmental process of the mouse heart and during the differentiation of P19 cells into cardiac myocytes,and to explore the possible relationship between miR-379 and the differentiation of cardiacmyocytes.Methods Heart tissues were collected from fetal mice in pregnant ones at their gestational age (8.5,11.5,14.5 and 18.5 days) respectively.Heart tissue sections of the fatal mice were obtained to observe the heart development process.Then total RNA was isolated from heart tissues by using the TRIzol method.Complementary DNA was synthesized from 1 μg total RNA by using a Reverse Transcriptase Kit.Finally,real-time PCR (RT-PCR) was employed to detect the expression of miR-379.At the same time,P19 cells were cultured with 10 mL/L Dimethyl sulfoxide in suspension for 4 days to form cell aggregation,and these aggregations were transferred into 6-wells plate for culturing by adherence.Beating cells were detected with microscopy on the 10th day after induction.Afterwards,total RNA was extracted from cultured P19 cells at different time points.Reverse transcription was executed to get DNA.At last,RT-PCR was used to explore the expression of miR-379 on 0,4,6,10 days after aggregation.Results The expression level of miR-379 was down-regulated gradually in the developing heart (at gestational age of 8.5,11.5,14.5,16.5 days,respectively),and there were significant differences on the different days (F =21.13,P ＜ 0.05).On the other hand,myocardial markers of troponin T represented an increasing trend during the process of P19 cells induction,which demonstrated that P19 cells were successfully induced into cardiomyocyte-like cells.Meanwhile,miR-379 showed a low expression on day 0 of P19 cells aggregation.On day 4,miR-379 demonstrated a higher level.Afterwards,miR-379 proved to be down-regulated gradually.Conclusions miR-379 plays a role in the process of the heart development,but the specific mechanisms need further research.
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10.3969/j.issn.2095-4344.2013.24.006
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10.3969/j.issn.2095-4344.2013.24.007
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The crude buffer (Tris Buffer Saline-I) extracts of muscles, liver, kidney and heart of goat and chicken (White leghorn) were screened against 16 clinical isolates. Among the five tissues, the heart tissue of each animal showed significant bactericidal activities on many isolates. The acid extracted crude proteins of both heart tissues also showed significant antibacterial activities against many bacterial isolates. The crude proteins of goat heart tissues displayed strong bactericidal activities against Salmonella paratyphi ‘A’ and Salmonella typhimurium (MIC: 16 g/ml) whereas the crude proteins of chicken heart tissues displayed strong bactericidal activities against Escherichia coli ATCC and Pseudomonas aeruginosa at 16 and 63 g/ml concentrations respectively. The peptides of low molecular weight (< 30 kDa) were also separated from the acid extracted crude proteins of goat and chicken heart tissues by SDS-PAGE after staining with silver nitrate solution.
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Objective To investigate the feasibility of transplantation of human fetal liver-delivered mesenchymal stem cells(HFMSCs) and porous microcarries into normal heart tissue and whether it can improve heart function and regeneration of heart tissue.Methods SD rats were divided into HFMSCs injection group(n=9),microcarrier injection group(n=9) and control group(n=4),in which 80-100 ?l Perfadex with HFMSCs or gelatin porous microcarriers or pure Perfadex was injected into the wall of left ventricle.Heart function was evaluated by UCG before and 7 d after transplantation.On day 7,14,the survival of HFMSCs was tested by fluorescent in situ hybridization(FISH),regeneration or cardiac differentiation by immunohistological staining against desmin,tropomyosin and lectin,cellular immune response by the infiltration of macrophages,and lymphocyte reaction to HFMSCs by mixed lymphocyte culture(MLC) in vitro.Results Seven days after injection,the HFMSCs survived and improved the heart function,though no sign of differentiation into cardiomyocytes was seen.On day 14,a large amount of macrophages infiltrated into injection sites,and MLC showed prominent enhancement of proliferation of lymphocytes,when no transplanted cells were detected in the myocardium.On day 7,14,the microcarriers retained their round shape at the injection sites and were attatched by a large quantity of cells which were proven not cardiomyocytes or capilaries by immunohistological staining.Conclusion Transplantation of HFMSCs into normal heart improves heart function by short-period survival without differentiation,but the transplanted cells disappeared because of immune reaction.Transplantation of porous microcarriers into normal heart could not improve heart function either by regeneration of heart tissue or capilaries.