动物造模 z-bio 2022-10-24 573

　　Objective Cardiac fibroblasts account for more than 50% of cardiac cells and play an important role in cardiac physiology and pathology. The 7th day after birth is a turning point when the heart loses its ability to regenerate. This study aimed to study the changes in the number and properties of cardiac fibroblasts in the early postnatal period of mice, and to provide basic experimental data for the study of myocardial regeneration mechanism.

　　Methods Apex resection (AR) was performed on C57 mice on the 1st and 7th days after birth. Stereoscopic photography and HE staining were used to observe the incision at the apex. After 21 days, the regeneration was observed. The heart tissue of C57 mice was weighed to compare the weight difference. The pathological sections were stained with HE to observe the pathological structure of the heart at two time points. To accurately quantify the quantitative changes of cardiac fibroblasts, we used the fibroblast surface marker Thymocyte differentiation antigen 1 (Thy1) to separate cardiac tissue cell suspensions of cardiac fibroblasts by flow sorting technology. Selected and compared the proportion of cardiac fibroblasts in the cardiac tissue of mice on 1 d and 7 d; in order to verify whether the cardiac fibroblast characteristics changed, we isolated 1 d and 7 d mouse cardiac fibroblasts, and analyzed the specificity of fibroblasts Sexual markers Thy1, Fibroblast-specific protein 1 (Fsp-1), Periostin, platelet-derived growth factor receptor α (Pdgfrα), type I The expression changes of collagen alpha-1 (Collagen alpha-1, Col1a1) and transcription factor 21 (Transcription factor 21, Tcf21) at the mRNA level.

　　Results The 1-day-old mice could fully regenerate at 21 days after apexectomy (the regeneration rate was 86. 67%), while the 7-day mice could not fully regenerate after the operation (the regeneration rate was 0). The tissue was 13. 13 mg (3. 11 times) heavier than the wet heart weight on 1 d (1 d: (6. 22±0. 19), 7 d: (19. 35±0. 56), P<0. 0001 ;n = 6). Immunofluorescence staining of cardiac fibroblasts showed that the number of cardiac fibroblasts in the 7-day heart was significantly higher than that in the 1-day heart; flow cytometry showed that the proportion of cardiac fibroblasts in the heart tissue of mice at 7 days after birth was higher than that of the total cells at 1 day after birth. 2. 9% (1. 38 times higher) at d (1 d: (7. 7±0. 74), 7 d: (10. 6±0. 95), P= 0. 029; n = 3) ; At the transcriptional level of fibroblasts, the expression of fibroblast marker proteins Thy1, Fsp-1 and Periostin on postnatal day 7 of mice was higher than that of fibroblasts on postnatal day 1 (Thy1: 1 d: (1. 01±0) .12), 7d: (2.71±0.27), P= 0.0288; Fsp-1: 1d: (1.04±0.27), 7d: (5.28±0. 10), P = 0. 0046; Periostin: 1 d: (0. 91 ± 0. 01), 7 d: (1. 13 ± 0. 05), P = 0. 0119; n = 3), while Pdgfrα , Col1a1, Tcf21 three fibroblast marker proteins expression was lower than that of fibroblasts in the mouse heart at 1 d after birth (Pdgfrα: 1 dL: (1. 09±0. 04), 7 d: (0. 62 ± 0. 01), P = 0. 0068; Col1a1: 1 d: (1. 00 ± 0. 09), 7 d: (0. 57 ± 0. 02), P = 0. 0433; Tcf21: 1 d: (1. 00 ± 0. 03), 7 d: (0. 54 ± 0. 02), P = 0. 0054; n = 3), demonstrating cardiac fibrogenesis during postnatal 1 d to 7 d Cell properties have changed.

　　Conclusions The proportion and characters of cardiac fibroblasts in mice in the early postnatal period (1d and 7d) changed significantly, providing some clues for studying the mechanism of mammalian myocardial regeneration loss.