ObjectiveTo investigate the effect of pharmacologic delay with pioglitazone, a peroxisome proliferator-activated receptor γ (PPAR-γ) agonist, on extended perforator flap survival in a rat model. MethodsSeventy male Sprague Dawley rats, weighing 250-300 g, were randomly divided into control group (n=35) and experimental group (n=35). A three-territory flap was made, including two choke zones. Pioglitazone was dissolved in 1.5 mL saline. Oral doses of pioglitazone[10 mg/(kg·d)] was given by gavaged for 5 days in the experimental group, while the same volume of saline was given in the control group at same time point. After 7 days, the flap survival area was measured and angiographic diagnosis was made. The tissue samples were harvested from choke zone Ⅱ for histological study and vascular endothelial growth factor (VEGF) expression detection by immunohistochemical staining. The content of nitric oxide (NO) in choke zones I and Ⅱ was measured at immediate, 1, 3, 5, and 7 days after operation. ResultsThe flap general change of 2 groups was similar. Varying degrees of necrosis occurred with the extension of time in 2 groups. At 7 days after operation, the flap survival rate was 87.73%±3.25% in the experimental group and 76.07%±2.92% in the control group, showing a significant difference (t=-10.338, P=0.000). The number of true anastomosis in choke zones I and Ⅱ was 5.40±1.14 and 3.00±0.71 in the experimental group, and was 3.20±0.84 and 0.80±0.84 in the control group respectively, showing significant differences between the 2 groups (t=-3.479, P=0.008;t=-4.491, P=0.002). The microvessel density and the expression of VEGF in choke zone Ⅱ of experimental group were (33.16±7.73)/mm2 and 4 368.80±458.23, respectively, which were significantly higher than those of control group[(23.29±5.91)/mm2 and 2 241.24±554.43] (t=5.073, P=0.000;t=-14.789, P=0.000). The content of NO in the experimental group were significantly higher than those in the control group at other time points (P<0.05) except for at immediate after operation. ConclusionPharmacologic delay with pioglitazone can improve extended perforator flap viability through increasing ischemia-induced angiogenesis and choke vessels vasodilation in rat models.
ObjectiveTo evaluate the correlation of activated partial thromboplastin time (aPTT), activated clotting time (ACT) and the activity of anti-factor Ⅹa activity with the concentration of unfractionated heparin (UFH) during extracorporeal membrane oxygenation (ECMO) in children after cardiac surgery.MethodsThe clinical data of children (aged 6 months to 6 years) who received ECMO support after cardiac surgery in Fuwai Hospital from January 2010 to October 2020 were retrospectively collected. And the aPTT value, ACT value, anti-Ⅹa activity and the corresponding UFH dose measured simultaneously during ECMO were recorded. According to the Extracorporeal Life Support Organization anticoagulation guideline, the bleeding events of children during ECMO support were defined, and the children were divided into a bleeding group and a non-bleeding group according to whether bleeding events occurred. Pearson correlation was used to evaluate the correlation between ACT, aPTT or anti-Ⅹa activity and UFH in the same patient.ResultsA total of 58 children, including 33 males and 25 females, aged 27.31±34.17 months, were enrolled and divided into the bleeding group (n=39) and the non-bleeding group (n=19). Univariate analysis showed that compared with children in the non-bleeding group, children in the bleeding group had lower red blood cell counts (P=0.049), hemoglobin concentration (P=0.010), and hematocrit (P=0.046) on the day of ECMO installation. In addition, the transfusion volume of fresh frozen plasma (P=0.034) and fibrinogen (P=0.033) in the bleeding group was relatively more, and the proportion of exploratory thoracotomy for hemostasis was high (P=0.000); there was a moderate degree of correlation between anti-Ⅹa and UFH (r=0.418, P=0.013) but there was no correlation between ACT or aPTT and UFH.ConclusionThe aPTT value and ACT value are poorly correlated with the concentration of UFH transfused during ECMO in children after cardiac surgery, while the anti-Ⅹa activity is moderately correlated with it.
Objective To investigate the role and mechanism of peroxisome proliferator-activated receptor gamma coactivator 1α (PGC-1α) in the activation of aortic valve interstitial cells (AVICs) in aortic stenosis. Methods Isolating primary AVICs and stimulating their activation with transforming growth factor β1 (TGF-β1, 30 ng/mL), the expression of PGC-1α was detected. The activation of AVICs induced by TGF-β1 was observed after overexpression of PGC-1α by adenovirus or inhibition of PGC-1α function by GW9662. The possible downstream molecular mechanism of PGC-1α in AVICs activation was screened. Finally, the phenotype was further verified in primary human AVICs. Results The expression of PGC-1α decreased after the activation of AVICs induced by TGF-β1 (control group: 1.00±0.18; 24 h: 0.31±0.10; 48 h: 0.32±0.06; 72 h: 0.20±0.07; P<0.05). Specific overexpression of PGC-1α by adenovirus inhibited the activation of AVICs induced by TGF-β1 stimulation (periostin: 3.17±0.64 vs. 1.45±0.54, P<0.05; α-smooth muscle actin: 0.77±0.11 vs. 0.28±0.06, P<0.05). On the contrary, inhibition of PGC-1α function by GW9662 promoted the activation of AVICs (periostin: 2.20±0.68 vs. 7.99±2.50, P<0.05). Subsequently, it was found that PGC-1α might inhibit the activation of AVICs through downregulating the expression of calcium/calmodulin-dependent protein kinase (CAMK1δ) (0.97±0.04 vs. 0.74±0.11, P<0.05), and downregulating the expression of CAMK1δ alleviated the activation of AVICs (periostin: 1.76±0.11 vs. 0.99±0.20, P<0.05). The possible mechanism was that the activation of mammalian target of rapamycin (mTOR) signaling pathway was inhibited by reducing the accumulation of reactive oxygen species (ROS) (778.3±139.4 vs. 159.3±43.2, P<0.05). Finally, the protective effect of PGC-1α overexpression was verified in the activated phenotype of human AVICs (periostin: 2.73±0.53 vs. 1.63±0.14, P<0.05; connective tissue growth factor: 1.27±0.04 vs. 0.48±0.09, P<0.05). Conclusions The expression of PGC-1α significantly decreases during the activation of AVICs induced by TGF-β1. The overexpression of PGC-1α significantly inhibites the activation of AVICs, suggesting that PGC-1α plays a protective role in the activation of AVICs. The possible mechanism is that PGC-1α can inhibit the activation of CAMK1δ-ROS-mTOR pathway. In conclusion, interventions based on PGC-1α expression levels are new potential therapeutic targets for aortic stenosis.
Objective To study the effect of p38MAPK activity on tumor necrosis factor-α （TNF-α） mRNA and intercellular adhesion molecule 1 （ICAM1） mRNA expressions of isolated rabbit liver during early stage of cold preservation and reperfusion period. Methods Based on the cold preservation and reperfusion model of isolated rabbit liver, the animals were divided into inhibition group （n=12） with 3 μmol/L SB202190 (p38MAPK specificity inhibitor) in perfusate and control group （n=12） with no SB202190 in perfusate. Liver tissue samples were harvested at the time points of before resection, end of cold preservation, and different reperfusion period (10, 30, 60 and 120 min). Protein expression and activity of p38MAPK were detected by Western blot and immunoprecipitation respectively, expression of TNF-α mRNA was detected by RT-PCR, and expression of ICAM1 mRNA was detected by in situ hybridization. Results There was no obvious change of expression of p38MAPK protein in liver tissue both in two groups during the total period （P＞0.05）， and there was no statistically significant difference between two groups （P＞0.05）. At time points of end of cold preservation, 10, 30 and 60 min of reperfusion, the activity of p38MAPK in control group was significantly higher than that at the time points of before resection and 120 min of reperfusion (P＜0.01), and was also significantly higher than that in inhibition group at the same time points (P＜0.01). There was no significant difference in activity of p38MAPK among all time points in inhibition group (P＞0.05). The expressions of TNF-α mRNA and ICAM1 mRNA at the time points of before resection, end of cold preservation, and 10 and 30 min of reperfusion were significantly lower than those in 60 and 120 min of reperfusion in both two groups (P＜0.05, P＜0.01); The expressions of TNF-α mRNA and ICAM1 mRNA in inhibition group were significantly lower than those in control group at the time points of 60 and 120 min of reperfusion (P＜0.01). The activity of p38MAPK of liver tissue during cold preservation and reperfusion period was significantly correlated with the level of TNF-α mRNA and level of ICAM1 mRNA expression （r=0.996, P＜0.01； r=0.985, P＜0.01）. Conclusions These results suggest that p38MAPK pathway may regulate the expressions of TNF-α and ICAM1 at the level of transcription and the activation of p38MAPK can up-regulate TNF-α and ICAM1 expressions, which may be one of the important mechanisms to cause ischemia-reperfusion injury of isolated liver during cold preservation and reperfusion period.
Objective To investigate the role of peroxisome proliferator activated receptor α （PPARα） in the pathogenesis of colorectal cancer. Method The literatures about PPARα and the pathogenesis of colorectal cancer were reviewed and analyzed. Result The relationships of PPARα to the proliferation， apoptosis， and differentiation of colorectal cancer cells in the pathogenesis of colorectal cancer were controversial. Conclusions PPARα might be involved in the regulation of proliferation， differentiation， apoptosis of colorectal cancer cells， but the pathogenesis and the up- and down-stream signal pathways are not elucidated. In additional， PPARα might partly be involved in the mechanism of drug resistance of chemotherapy drugs for colorectal cancer， but the role is not very clear yet. So more research works need to be done about the relationship of PPARα to pathogenesis of colorectal cancer.
【 Abstract 】 Objective To investigate the protective effect of peroxisome proliferator-activated receptor γ (PPAR γ ) activator 15-deoxyprostaglandin J2 (15d-PGJ2) in rat hepatic ischemia-reperfusion injury and its mechanism. Methods The models of 70% warm ischemia-reperfusion injury were established in SD rats, rats were randomly divided into 4 groups: sham operation group, ischemia-reperfusion group, 15d-PGJ2 group and 15d-PGJ2+GW9662 group. After reperfusion, serum AST and ALT levels were determined; the liver tissues were removed for measurement of activity of NF-κB and myeloperoxidase (MPO), TNF-α content and expression of ICAM-1. Results Compared with sham operation group, the serum levels of ALT and AST, and the activities of MPO and NF- κ B, TNF- α content and expression of ICAM-1 in ischemia-reperfusion group, 15d-PGJ2 group and 15d-PGJ2+GW9662 group were greatly improved (P ＜ 0.05). Compared with ischemia-reperfusion group, the serum levels of ALT and AST and the activities of MPO and NF- κ B, TNF- α content and expression of ICAM-1 in 15d-PGJ2 group were significantly decreased (P ＜ 0.05). Compared with 15d-PGJ2 group, the serum levels of ALT and AST, and the activities of MPO and NF- κ B, TNF- α content and the expression of ICAM-1 in 15d-PGJ2+GW9662 group were obviously increased (P ＜ 0.05). Conclusion PPAR γ activator 15d-PGJ2 could protect against ischemia-reperfusion injury in rats, with its possible mechanism of inhibiting NF-κB activation and down-regulating TNF-α content and ICAM-1 expression in a PPARγ dependent fashion.
【Abstract】ObjectiveTo explore the effects of p38 mitogenactivated protein kinase (MAPK) on apoptosis of small intestinal epithelial cells after transplantation in rats. MethodsSmall intestinal transplantation was performed in SD and Wistar rats. The recipients were divided into three groups: isograft group (Wistar→Wistar group), allograft group (SD→Wistar group) and allograft+cyclosporine A group (SD→Wistar+CsA group). The grafts were harvested on day 1, 3, 5 and 7 after operation. All graft samples were subjected to histological examination. The apoptosis of graft epithelial cells was detected by TUNEL method. p38 MAPK was measured by Westernblotting method and serum TNFα was determined by ELISA. ResultsMild, moderate and severe rejection reaction occurred in the SD→Wistar group, it was showed that the number of apoptotic cells increased with the severity of the rejection reaction by TUNEL. In SD→Wistar group, the numbers of apoptotic cells were significantly higher than those of the other two groups （P＜0.01）. The severity of rejection reaction in SD→Wistar+CsA group was less than that of SD→Wistar group and the number of apoptotic cells increased with the severity of the rejection reaction (P＜0.01). The level of serum TNFα varied with the apoptotic degree of small intestinal epithelial cells in SD→Wistar group and SD→Wistar+CsA group （P＜0.01）. The expression of p38 MAPK increased with the number of the apoptotic cells in SD→Wistar group and SD→Wistar+CsA group （P＜0.01）, but there was no evident change in Wistar→Wistar group （Pgt;0.05）. The expression of p38 MAPK and the level of serum TNFα were positively correlated with apoptosis in small intestinal rejection after transplantation （r=0.875， P＜0.01； r=0.837， P＜0.01）. p38 MAPK and TNFα were also positively correlated （r=0.826，P＜0.01）. ConclusionApoptosis plays an important role in small intestinal rejection. p38 MAPK is involved in apoptosis and is an important regulator in signal pathway of cell apoptosis.
ObjectiveTo observe the effects of p21 activated kinase 4 (PAK4) on the mitochondrial function and biological behavior in retinal vascular endothelial cells. MethodsThe experimental study was divided into two parts: in vivo animal experiment and in vitro cell experiment. In vivo animal experiments: 12 healthy C57BL/6J male mice were randomly divided into normal control group and diabetes group, with 6 mice in each group. Diabetes mice were induced by streptozotocin to establish diabetes model. Eight weeks after modeling, quantitative real-time polymerase chain reaction and Western blots were performed to detect the expression of PAK4 in diabetic retinas. In vitro cell experiments: the human retinal microvascular endothelial cells (hRMEC) were divided into three groups: conventional cultured cells group (N group), empty vector transfected (Vector group); pcDNA-PAK4 eukaryotic expression plasmid transfected group (PAK4 group). WB and qPCR were used to detect transfection efficiency, while scratching assay, cell scratch test was used to detect cell migration in hRMEC of each group. In vitro white blood cell adhesion experiment combined with 4 ', 6-diamino-2-phenylindole staining was used to detect the number of white blood cells adhering to hRMEC in each group. The Seahorse XFe96 cell energy metabolism analyzer measures intracellular mitochondrial basal respiration, adenosine triphosphate (ATP) production, maximum respiration, and reserve respiration capacity. The t-test was used for comparison between the two groups. Single factor analysis of variance was used for comparison among the three groups. ResultsIn vivo animal experiments: compared with normal control group, the relative expression levels of PAK4 mRNA and protein in retina of diabetic mice were significantly increased, with statistical significance (t=25.372, 22.419, 25.372; P＜0.05). In vitro cell experiment: compared with the N group and Vector group, the PAK4 protein, mRNA relative expression and cell mobility in the hRMEC of PAK4 group were significantly increased, with statistical significance (F=36.821, 38.692, 29.421; P＜0.05). Flow cytometry showed that the adhesion number of leukocytes on hRMEC in PAK4 group was significantly increased, and the difference was statistically significant (F=39.649, P＜0.01). Mitochondrial pressure measurement results showed that the capacity of mitochondrial basic respiration, ATP production, maximum respiration and reserve respiration in hRMEC in PAK4 group was significantly decreased, with statistical significance (F=27.472, 22.315, 31.147, 27.472; P＜0.05). ConclusionOver-expression of PAK4 impairs mitochondrial function and significantly promotes leukocyte adhesion and migration in retinal vascular endothelial cells.
Objective To explore the regulation of peroxisome proliferator-activated receptor γ coactivator 1α( PGC-1α) and NF-E2-related factor 2( Nrf2) on expression of γ-glutamylcysteine synthetase ( γ-GCS) , and their roles in chronic obstructive pulmonary disease( COPD) . Methods Twenty-four SD rats were randomly divided into a COPD group and a normal control group. COPD model was established by intratracheal instillation of lipopolysaccharide ( LPS) and daily exposure to cigarette smog in the COPD group. The lung function was measured and the pathological changes were observed. The protein and mRNA expressions of PGC-1α, Nrf2, and γ-GCS in lung tissue were measured by immunohistochemistry, Western blot, in site hybridization ( ISH) , and reverse transcription-polymerase chain reaction ( RT-PCR ) ,respectively. Results In the COPD group, the pulmonary function ( FEV0. 3, FEV0. 3 /FVC, PEF) damage and lung pathological changes were conformed as morphological characteristics of COPD. The mRNA of PGC-1α and Nrf2 expressed in lung tissues of two group rats in the region consistent with γ-GCS mRNA. The protein and mRNA expressions of PGC-1αand γ-GCS were markedly increased in the COPD group( all P lt;0. 05) ,and the protein expression of Nrf2 was obviously up-regulated ( P lt; 0. 01) , while Nrf2 mRNA had no significant difference between the two groups( P gt;0. 05 ) . Linear correlation analysis showed that the level ofPGC-1αprotein was positively correlated with the levels of Nrf2 protein and mRNA ( r = 0. 775, 0. 515, all P lt; 0. 01) , and the levels of PGC-1αand Nrf2 protein were positively correlated with the levels of γ-GCS protein ( r = 0. 531, 0. 575, all P lt; 0. 01) and mRNA ( r = 0. 616, 0. 634, all P lt; 0. 01) . Conclusions PGC-1α, which may serve as a co-activator of Nrf2, can up-regulate the expression of γ-GCS gene cooperatively with Nrf2 through a common pathway, which might involve in the oxidative and antioxidative mechanism in the pathogenesis of COPD.
Epilepsy is a disorder of the brain in which sudden abnormal discharges of neurons cause transient dysfunction and is a common disorder of the nervous system. Although most patients experience remission of symptoms with medication, about 20 ~ 30% of patients still have poor outcomes with medication and progress to refractory epilepsy. The etiology of epilepsy is complex and the exact pathogenesis is not yet clear. Current research has explored the pathophysiological mechanisms underlying epileptogenesis, thus providing a basis for identifying potential therapeutic targets for epilepsy and advancing the precision treatment of epilepsy. p38 Mitogen-activated protein kinase (MAPK) signalling pathway is a conserved class of kinases involved in many physiological/pathological processes by regulating intracellular gene expression levels, cell division, differentiation and apoptosis in response to various extracellular stimuli in order to mediate intracellular signalling cascades. The p38 MAPK signalling pathway is one of the subfamilies of MAPK that mediates inflammatory responses, apoptosis, tissue edema and other biological processes involved in the development of central nervous system diseases. The p38 MAPK signalling pathway is now reviewed for its involvement in the development of epilepsy through unused pathways, in order to identify new potential targets for epilepsy treatment and provide clinical precision.