Effect of Wenyang Zhenshuai Granules on autophagy and apoptosis of myocardial cells in septic rats via regulating miR-132-3p/UCP2 expression / 中国中药杂志

This study aimed to investigate the effect of Wenyang Zhenshuai Granules(WYZSG) on autophagy and apoptosis of myocardial cells in rats with sepsis via regulating the expression of microRNA-132-3p(miR-132-3p)/uncoupling protein 2(UCP2). Sixty SD rats were randomly divided into modeling group(n=50) and sham operation group(n=10). The sepsis rat model was constructed by cecal ligation and perforation in the modeling group. The successfully modeled rats were randomly divided into WYZSG low-, medium-and high-dose groups, model group and positive control group. Rats in the sham operation group underwent opening and cecum division but without perforation and ligation. Hematoxylin-eosin(HE) staining was used to observe the pathological changes of rat myocardial tissue. Myocardial cell apoptosis was detected by TdT-mediated dUTP nick end labeling(TUNEL) assay. Real-time quantitative polymerase chain reaction(RT-qPCR) was performed to detect the expression of miR-132-3p and the mRNA expressions of UCP2, microtubule-associated protein light chain 3(LC3-Ⅱ/LC3-Ⅰ), Beclin-1 and caspase-3 in rat myocardial tissue. The protein expressions of UCP2, LC3-Ⅱ/LC3-Ⅰ, Beclin-1 and caspase-3 in myocardial tissue were detected by Western blot. Dual luciferase reporter assay was used to verify the regulatory relationship between miR-132-3p and UCP2. The myocardial fibers of sepsis model rats were disordered, and there were obvious inflammatory cell infiltration as well as myocardial cell edema and necrosis. With the increase of the WYZSG dose, the histopathological changes of myocardium were improved to varying degrees. Compared with the conditions in the sham operation group, the survival rate and left ventricular ejection fraction(LVEF) of rats in the model group, positive control group and WYZSG low-, medium-and high-dose groups were decreased, and the myocardial injury score and apoptosis rate were increased. Compared with the model group, the positive control group and WYZSG low-, medium-and high-dose groups had elevated survival rate and LVEF, and lowered myocardial injury score and apoptosis rate. The expression of miR-132-3p and the mRNA and protein expressions of UCP2 in myocardial tissue in the model group, positive control group and WYZSG low-, medium-and high-dose groups were lower, while the mRNA and protein expressions of LC3-Ⅱ/LC3-Ⅰ, Beclin-1 and caspase-3 were higher than those in the sham operation group. Compared with model group, the positive control group and the WYZSG low-, medium-and high-dose groups had an up-regulation in the expression of miR-132-3p and the mRNA and protein expressions of UCP2, while a down-regulation in the mRNA and protein expressions of LC3-Ⅱ/LC3-Ⅰ, Beclin-1 and caspase-3. WYZSG inhibited excessive autophagy and apoptosis of myocardial cells in septic rats and improved myocardial injury, possibly by regulating the expression of miR-132-3p/UCP2.

Effect of exercise-induced irisin on myocardial fibrosis / 中国组织工程研究

BACKGROUND: Myocardial fibrosis is an important topic in modern medical research, and its development is closely related to common heart diseases such as arrhythmia and chronic heart failure. Exercise intervention can significantly improve myocardial fibrosis, but there is no systematic and comprehensive understanding of the mechanism by which exercise improves myocardial fibrosis as well as the effects of different types of exercises on myocardial fibrosis. To date, it is still unclear about how exercise triggers the production of irisin against myocardial fibrosis. OBJECTIVE: To comprehensively review the exercise-induced production of irisin and its effect on myocardial fibrosis, and reveal its myocardial protection, so as to improve heart function and provide fundamental basis for preventing against common heart diseases, such as arrhythmia and chronic heart failure. METHODS: A search of ELSEVIER, Web of Science, CNKI, WanFanga, VIP and Taiwan Academic Literature Database was performed for articles regarding exercise, irisin, and myocardial fibrosis. The deadline for publication was August 2019. According to the inclusion and exclusion criteria, 58 articles were eligible for review. RESULTS AND CONCLUSION: Long-term and single exercise in human experiments has been shown to improve muscular and circulatory irisin levels, which has been better verified in animal experiments. A few experimental results indicate that long-term exercise has no significant effect on blood irisin levels, which may be due to different research subjects, exercise methods, exercise intensity, and exercise frequency. However, the specific mechanism is still unclear. Exercise can improve myocardial fibrosis by acting on myocardial mitochondrial stabilization, energy metabolism, oxidative stress and inflammatory response. The occurrence of myocardial fibrosis results from the regulation of neuroendocrine and oxidative stress and inflammatory responses. Irisin can influence the processes of oxidative stress and inflammation related to the mechanism of myocardial fibrosis, by inhibiting ROS/p38MAPK/NF-κB signaling pathway, endogenous reactive oxygen species and ROS-NLRP3 inflammation signaling pathway, and regulating the expression of uncoupling protein 2 and mitochondrial homeostasis. Therefore, exercise may improve myocardial fibrosis by upregulating the expression of irisin, thus providing myocardial protection.

Effect of uncoupling protein 2 gene on radiation sensitivity of Siha cells / 中华放射医学与防护杂志

Objective@#To investigate whether silencing UCP2 can sensitize cervical cancer cell line Siha to radiation.@*Methods@#Siha cells were transfected with UCP2 siRNA and then irradiated by X-ray. The radiosensitivity of Siha cells was verified by colony formation, CCK-8, apoptosis and immunofluorescence assays. The mitochondrial membrane potential and the production of reactive oxygen species (ROS) were detected to further explore the related mechanism.@*Results@#RT-PCR and Western blot assays showed that the expression of UCP2 in Siha cells was increased after irradiation and the UCP2 siRNA successfully silenced the expression of UCP in cells. According to the survival curves, the D0, Dq, N and SF2 were 1.54, 1.31, 2.31 Gy and 0.52 for siUCP2 group, 2.50, 3.64, 4.30 Gy and 0.83 for blank control group, and 3.34, 2.16, 1.91 Gy and 0.69, for siNC group, respectively. The radiosensitivity enhancement ratio of silent group was 0.62 and 0.46, compared with blank control group and negative control group, respectively. The proliferative activity of cells in the silent group was lower than that in the control group (t=13.2, P<0.05). Apoptosis levels in the silent group were significantly higher than those in the control group after irradiation(t=3.14, P<0.05). At 4 h after irradiation, the ROS production in the silent group was significantly higher than that in the control group (t=19.10, P<0.05). At 24 h after irradiation, the mitochondrial membrane potential of Siha cells in the silent group was significantly lower than that in the control group (t=4.18, P<0.05).@*Conclusions@#The radiosensitivity of Siha cells is enhanced after UCP2 silencing, and thus UCP2 may applicable as a new target for radiosensitization of cervical cancer cells.

Effect of uncoupling protein 2 gene on radiation sensitivity of Siha cells / 中华放射医学与防护杂志

Objective To investigate whether silencing UCP2 can sensitize cervical cancer cell line Siha to radiation.Methods Siha cells were transfected with UCP2 siRNA and then irradiated by Xray.The radiosensitivity of Siha cells was verified by colony formation,CCK-8,apoptosis and immunofluorescence assays.The mitochondrial membrane potential and the production of reactive oxygen species (ROS) were detected to further explore the related mechanism.Results RT-PCR and Western blot assays showed that the expression of UCP2 in Siha cells was increased after irradiation and the UCP2 siRNA successfully silenced the expression of UCP in cells.According to the survival curves,the D0,Dq,N and SF2 were 1.54,1.31,2.31 Gy and 0.52 for siUCP2 group,2.50,3.64,4.30 Gy and 0.83 for blank control group,and 3.34,2.16,1.91 Gy and 0.69,for siNC group,respectively.The radiosensitivity enhancement ratio of silent group was 0.62 and 0.46,compared with blank control group and negative control group,respectively.The proliferative activity of cells in the silent group was lower than that in the control group (t =13.2,P＜0.05).Apoptosis levels in the silent group were significantly higher than those in the control group after irradiation (t=3.14,P＜0.05).At 4 h after irradiation,the ROS production in the silent group was significantly higher than that in the control group (t=19.10,P＜0.05).At 24 h after irradiation,the mitochondrial membrane potential of Siha cells in the silent group was significantly lower than that in the control group (t =4.18,P ＜ 0.05) Conclusions The radiosensitivity of Siha cells is enhanced after UCP2 silencing,and thus UCP2 may applicable as a new target for radiosensitization of cervical cancer cells.

Effects of uncoupling protein 2 overexpression on myocardial mitochondrial dynamics in sepsis rats / 中华危重病急救医学

Objective@#To investigate the effects of uncoupling protein 2 (UCP2) overexpression on mitochondrial dynamics (mitochondrial division and fusion) of sepsis myocardial injury in rats.@*Methods@#Forty male Sprague-Dawley (SD) rats were randomly divided into four groups (n = 10): sham operation group (Sham group) using normal saline instead of transfection and simulating cecal ligation and perforation (CLP); CLP group using normal saline instead of transfection, performing CLP to induce sepsis; adeno-associated virus (AAV) group using CLP after myocardial transfection with empty virus; UCP2 overexpression group (UCP2 group) CLP was performed 3 weeks after AAV-UCP2 (1×1015 vg/L, a total of 60 μL) myocardial transfection. The rats in each group were examined by echocardiography at 24 hours after the CLP, and then the rats were sacrificed immediately to harvest myocardial tissue. Myocardial ultrastructural changes were observed under the electron microscope, the expression of regulatory proteins related to myocardial mitochondrial dynamics [optic atrophy 1 (Opa1), dynamin-related protein 1 (Drp1) and fission 1 (Fis1)] were detected by Western Blot, and the level of mitochondrial adenosine triphosphate (ATP) production was detected by chemiluminescence.@*Results@#①The echocardiographic results showed that there was no significant difference in left ventricular mass (LVM) and stroke volume (SV). Compared with Sham group, left ventricular diastolic anterior wall thickness (LVAWd), left ventricular systolic anterior wall thickness (LVAWs), left ventricular diastolic posterior wall thickness (LVPWd), left ventricular systolic posterior wall thickness (LVPWs), left ventricular ejection fraction (LVEF) and left ventricular short axis shortening rate (LVFS) were significantly increased in CLP group and AAV group, while left ventricular systolic diameter (LVEDs), left ventricular diastolic diameter (LVEDd), left ventricular end-systolic volume (LVESV), and left ventricular end-diastolic volume (LVEDV) were significantly decreased. Compared with CLP group and AAV group, LVAWs, LVEF, LVFS were significantly decreased in UCP2 group, and LVEDs, LVEDV and LVESV were significantly increased [LVAWs (mm): 3.82±0.42 vs. 4.34±0.30, 4.44±0.12; LVEF: 0.921±0.038 vs. 0.979±0.019, 0.991±0.010; LVFS: (65.33±6.56)% vs. (80.11±8.23)%, (85.31±6.11)%; LVEDs (mm): 1.81±0.36 vs. 0.89±0.54, 0.60±0.17; LVEDV (μL): 137.09±50.05 vs. 89.72±53.04, 85.42±40.99; LVESV (μL): 10.48±4.59 vs. 2.48±3.52, 2.58±2.50, all P < 0.05]. ② Electron microscope showed that the structure of myocardial fibers in the Sham group was clear and aligned with complete intervertebral disc and mitochondrial structure, no damage to mitochondrial membranes, and tight arrangement of cristae. In CLP group and AAV group, muscle fiber breakage, sarcoplasmic reticulum expansion, severe mitochondrial swelling and even cristage structure disorder were observed. In the UCP2 group, only myocardial fiber edema was observed, and the muscle fiber structure was more complete than that of Sham group and AAV group. The mitochondria were slightly swollen and the cristae were intact.③ Western Blot showed that there was no significant difference in the expression of Opa1 and Fis1 in the four groups. The expression of Drp1 in CLP group and AAV group were significantly higher than that in Sham group. The expression of Drp1 in UCP2 group was significantly lower than that in CLP group and AAV group (Drp1/β-actin: 1.01±0.03 vs. 1.39±0.03, 1.49±0.03, both P < 0.05).④ The results of immunofluorescence showed that the ATP content of CLP group and AAV group were significantly lower than that of Sham group; the ATP content of UCP2 group was significantly higher than that of CLP group and AAV group (μmol/L: 1.99±0.15 vs. 1.10±0.17, 1.13±0.19, both P < 0.05).@*Conclusion@#UCP2 overexpression can significantly improve the systemic systolic function of myocardium in sepsis rats, protect myocardial mitochondrial ultrastructure, inhibit mitochondrial division, and improve mitochondrial ATP synthesis.

Effects of uncoupling protein 2 overexpression on myocardial mitochondrial dynamics in sepsis rats / 中华危重病急救医学

Objective To investigate the effects of uncoupling protein 2 (UCP2) overexpression on mitochondrial dynamics (mitochondrial division and fusion) of sepsis myocardial injury in rats. Methods Forty male Sprague-Dawley (SD) rats were randomly divided into four groups (n = 10): sham operation group (Sham group) using normal saline instead of transfection and simulating cecal ligation and perforation (CLP); CLP group using normal saline instead of transfection, performing CLP to induce sepsis; adeno-associated virus (AAV) group using CLP after myocardial transfection with empty virus; UCP2 overexpression group (UCP2 group) CLP was performed 3 weeks after AAV-UCP2 (1×1015 vg/L, a total of 60 μL) myocardial transfection. The rats in each group were examined by echocardiography at 24 hours after the CLP, and then the rats were sacrificed immediately to harvest myocardial tissue. Myocardial ultrastructural changes were observed under the electron microscope, the expression of regulatory proteins related to myocardial mitochondrial dynamics [optic atrophy 1 (Opa1), dynamin-related protein 1 (Drp1) and fission 1 (Fis1)] were detected by Western Blot, and the level of mitochondrial adenosine triphosphate (ATP) production was detected by chemiluminescence. Results ①The echocardiographic results showed that there was no significant difference in left ventricular mass (LVM) and stroke volume (SV). Compared with Sham group, left ventricular diastolic anterior wall thickness (LVAWd), left ventricular systolic anterior wall thickness (LVAWs), left ventricular diastolic posterior wall thickness (LVPWd), left ventricular systolic posterior wall thickness (LVPWs), left ventricular ejection fraction (LVEF) and left ventricular short axis shortening rate (LVFS) were significantly increased in CLP group and AAV group, while left ventricular systolic diameter (LVEDs), left ventricular diastolic diameter (LVEDd), left ventricular end-systolic volume (LVESV), and left ventricular end-diastolic volume (LVEDV) were significantly decreased. Compared with CLP group and AAV group, LVAWs, LVEF, LVFS were significantly decreased in UCP2 group, and LVEDs, LVEDV and LVESV were significantly increased [LVAWs (mm): 3.82±0.42 vs. 4.34±0.30, 4.44±0.12;LVEF: 0.921±0.038 vs. 0.979±0.019, 0.991±0.010; LVFS: (65.33±6.56)% vs. (80.11±8.23)%, (85.31±6.11)%;LVEDs (mm): 1.81±0.36 vs. 0.89±0.54, 0.60±0.17; LVEDV (μL): 137.09±50.05 vs. 89.72±53.04, 85.42±40.99;LVESV (μL): 10.48±4.59 vs. 2.48±3.52, 2.58±2.50, all P < 0.05]. ② Electron microscope showed that the structure of myocardial fibers in the Sham group was clear and aligned with complete intervertebral disc and mitochondrial structure, no damage to mitochondrial membranes, and tight arrangement of cristae. In CLP group and AAV group, muscle fiber breakage, sarcoplasmic reticulum expansion, severe mitochondrial swelling and even cristage structure disorder were observed. In the UCP2 group, only myocardial fiber edema was observed, and the muscle fiber structure was more complete than that of Sham group and AAV group. The mitochondria were slightly swollen and the cristae were intact.③ Western Blot showed that there was no significant difference in the expression of Opa1 and Fis1 in the four groups. The expression of Drp1 in CLP group and AAV group were significantly higher than that in Sham group. The expression of Drp1 in UCP2 group was significantly lower than that in CLP group and AAV group (Drp1/β-actin:1.01±0.03 vs. 1.39±0.03, 1.49±0.03, both P < 0.05).④ The results of immunofluorescence showed that the ATP content of CLP group and AAV group were significantly lower than that of Sham group; the ATP content of UCP2 group was significantly higher than that of CLP group and AAV group (μmol/L: 1.99±0.15 vs. 1.10±0.17, 1.13±0.19, both P < 0.05). Conclusion UCP2 overexpression can significantly improve the systemic systolic function of myocardium in sepsis rats, protect myocardial mitochondrial ultrastructure, inhibit mitochondrial division, and improve mitochondrial ATP synthesis.

Effects of uncoupling protein 2 overexpression on myocardial mitochondrial dynamics in sepsis rats / 中华危重病急救医学

Objective To investigate the effects of uncoupling protein 2 (UCP2) overexpression on mitochondrial dynamics (mitochondrial division and fusion) of sepsis myocardial injury in rats. Methods Forty male Sprague-Dawley (SD) rats were randomly divided into four groups (n = 10): sham operation group (Sham group) using normal saline instead of transfection and simulating cecal ligation and perforation (CLP); CLP group using normal saline instead of transfection, performing CLP to induce sepsis; adeno-associated virus (AAV) group using CLP after myocardial transfection with empty virus; UCP2 overexpression group (UCP2 group) CLP was performed 3 weeks after AAV-UCP2 (1×1015 vg/L, a total of 60 μL) myocardial transfection. The rats in each group were examined by echocardiography at 24 hours after the CLP, and then the rats were sacrificed immediately to harvest myocardial tissue. Myocardial ultrastructural changes were observed under the electron microscope, the expression of regulatory proteins related to myocardial mitochondrial dynamics [optic atrophy 1 (Opa1), dynamin-related protein 1 (Drp1) and fission 1 (Fis1)] were detected by Western Blot, and the level of mitochondrial adenosine triphosphate (ATP) production was detected by chemiluminescence. Results ①The echocardiographic results showed that there was no significant difference in left ventricular mass (LVM) and stroke volume (SV). Compared with Sham group, left ventricular diastolic anterior wall thickness (LVAWd), left ventricular systolic anterior wall thickness (LVAWs), left ventricular diastolic posterior wall thickness (LVPWd), left ventricular systolic posterior wall thickness (LVPWs), left ventricular ejection fraction (LVEF) and left ventricular short axis shortening rate (LVFS) were significantly increased in CLP group and AAV group, while left ventricular systolic diameter (LVEDs), left ventricular diastolic diameter (LVEDd), left ventricular end-systolic volume (LVESV), and left ventricular end-diastolic volume (LVEDV) were significantly decreased. Compared with CLP group and AAV group, LVAWs, LVEF, LVFS were significantly decreased in UCP2 group, and LVEDs, LVEDV and LVESV were significantly increased [LVAWs (mm): 3.82±0.42 vs. 4.34±0.30, 4.44±0.12;LVEF: 0.921±0.038 vs. 0.979±0.019, 0.991±0.010; LVFS: (65.33±6.56)% vs. (80.11±8.23)%, (85.31±6.11)%;LVEDs (mm): 1.81±0.36 vs. 0.89±0.54, 0.60±0.17; LVEDV (μL): 137.09±50.05 vs. 89.72±53.04, 85.42±40.99;LVESV (μL): 10.48±4.59 vs. 2.48±3.52, 2.58±2.50, all P < 0.05]. ② Electron microscope showed that the structure of myocardial fibers in the Sham group was clear and aligned with complete intervertebral disc and mitochondrial structure, no damage to mitochondrial membranes, and tight arrangement of cristae. In CLP group and AAV group, muscle fiber breakage, sarcoplasmic reticulum expansion, severe mitochondrial swelling and even cristage structure disorder were observed. In the UCP2 group, only myocardial fiber edema was observed, and the muscle fiber structure was more complete than that of Sham group and AAV group. The mitochondria were slightly swollen and the cristae were intact.③ Western Blot showed that there was no significant difference in the expression of Opa1 and Fis1 in the four groups. The expression of Drp1 in CLP group and AAV group were significantly higher than that in Sham group. The expression of Drp1 in UCP2 group was significantly lower than that in CLP group and AAV group (Drp1/β-actin:1.01±0.03 vs. 1.39±0.03, 1.49±0.03, both P < 0.05).④ The results of immunofluorescence showed that the ATP content of CLP group and AAV group were significantly lower than that of Sham group; the ATP content of UCP2 group was significantly higher than that of CLP group and AAV group (μmol/L: 1.99±0.15 vs. 1.10±0.17, 1.13±0.19, both P < 0.05). Conclusion UCP2 overexpression can significantly improve the systemic systolic function of myocardium in sepsis rats, protect myocardial mitochondrial ultrastructure, inhibit mitochondrial division, and improve mitochondrial ATP synthesis.

Down-regulated uncoupling protein 2 expression suppresses Fadu cell proliferation / 解剖学报

Objective The effect of uncoupling protein 2(UCP2) on the Fadu cells of the hypopharyngeal cancer was investigated in this study. The siRNA technology was used to study the effect of UCP2 on cell cycle and cell proliferation. iMeanwhile, cisplatin, a commonly chemotherapeutic drug, was researched on the effect of proliferation in Fadu cells after down-regulated UCP2 gene. Methods Flow cytometry/PI staining was used to observe the effect of UCP2 on Fadu cell cycle. Clone formation assay was used to detect the effect of down-regulated UCP2 on the proliferation of Fadu cells. MTT assay and Bnid assay were used to determine whether UCP2 enhanced the sensitivity of Fadu cells to cisplatin and to detect the effect of UCP2 on the proliferation of Fadu. Results The present result showed that down-regulated UCP2 expression inhibited Fadu cell cycle in G,/S phase and cell colony formation, but had no synergistic effect on inhibition of Fadu proliferation by cisplatin; The expression of UCP2 was significantly inhibited by genipin, and then inhibited cell proliferation in a time-concentration-dependent manner. Conclusion The proliferation of Fadu cells was weakened and the cell cycle was inhibited in G,/S phase after down-regulated UCP2, suggesting that UCP2 may play an important adjust role in the development in Fadu of hypopharyngeal cancer.

Correlation between circulating uncoupling protein 2 level and severity of coronary artery lesions in patients with stable coronary artery disease / 实用医学杂志

Objective To investigate the correlation between circulating uncoupling protein 2(UCP2) level and severity of coronary artery disease(including Gensini score and criminal vessel counts)in patients with stable coronary artery disease(SCAD),and to analyze the predictive value of circulating UCP2 and urine acid (UA)for SCAD. Methods Three hundred and thirty patients from June 2015 to June 2017 were enrolled. Two hundred and forty patients with SCAD(SCAD group),90 patients without coronary artery disease(control group) were diagnosed. The circulating UCP2 level was detected by enzyme linked immunosorbent assay (ELISA) sandwich method. Results The levels of circulating UCP2 and UA in SCAD group were higher than those in the control group(UCP2[1.60(0.67,4.60)ng/mL]vs.[0.42(0.28,0.59)ng/mL](P<0.01),UA[(365.74 ± 66.06) μmol/L] vs. [(268.11 ± 45.81)μmol/L],P < 0.01). Multivariate logistic regression analysis showed that UCP2 (OR = 1.010 ,95% CI :1.001 ~ 1.020 ,P = 0.025)and UA(OR = 1.039 ,95% CI :1.007 ~ 1.072 ,P < 0.05)were independently associated with SCAD. Correlation analysis showed that the circulating UCP2 level was positively correlated with Gensini score(r=0.780,P<0.01)and criminal vessel counts(r=0.543,P<0.01). The receiver operating characteristic curve(ROC)showed that the optimal cutoff point of the circulating UCP2 level predicting SCAD was 0.64 ng/mL,and the sensitivity was 0.833 and the specificity was 0.944. No significant difference was observed in area under the curve between circulating UCP2 and UA(ΔAUC). Conclusion The high circulating UCP2 level indicates more severe coronary lesions in patients with SCAD. Circulating UCP2 level may be a new indicator of predicting SCAD,equal to the traditional oxidative stress related indicator of serum UA.

Protective effects and mechanism of insulin on brain in septic rats / 中华实用儿科临床杂志

Objective To investigate the protective effects and mechanism of insulin(INS) on brain in septic rats,and explore the possible role of uncoupling protein 2 (UCP2) in these effects.Methods Fifty male specific pathogen free(SPF) Sprague-Dawley rats were randomly divided into normal control (CN) group(n=10),lipopolysaccharide(LPS) group(n=20) and INS group (n=20) according to random number table.The septic rat model was established through an intraperitoneal injection of 15 mg/kg LPS of gram-negative bacteria.The rats in the INS group received a 1 U/kg INS injection subcutaneously 30 minutes before the injection of LPS,and the rats in the CN group were given equivalent 9 g/L saline in the same way.Eight rats in each group were killed,and their cerebral cortex were collected after the injection of LPS for 24 h.Pathological change of cerebral cortex was detected by Hematoxylin-Eosin(HE) staining.The cerebral cortex mitochondia were extracted for detecting the levels of reactive oxygen species(ROS),malondialdehyde (MDA) and the activity of superoxide dismutase(SOD).Neuronal apoptosis was detected by terminal dexynucleotidyl transferase(TdT)-mediated dUTP nick end labeling staining.UCP2 mRNA expression was detected by quantitative real-time(RT)-PCR.Apoptosis-associated protein B lymphocyte tumor-2(Bcl-2),Bcl-2 associated X protein(Bax),cleaved cysteinyl aspartate specific protease(cleaved Caspase-9) and UCP2 protein expression were determined by Western blot.Results (1)Compared with the CN group,obvious abnormal pathological change was revealed by HE staining in cerebral cortex of rats in the LPS group and the INS group,but the pathological change was attenuated in the INS group compared with the LPS group.(2)Compared with the CN group,the levels of mitochondrial ROS[(210.01±14.09) RFU vs.(49.06±7.28) RFU] and MDA[(2.19±0.18) nmol/mg pro vs.(1.25±0.11)nmol/mg pro]in the LPS group significantly increased,whereas SOD activity significantly decreased [(238.49±35.60) U/g pro vs.(446.66±24.90)U/g pro],and the differences were significant(all P<0.05).Compared with the LPS group,the levels of ROS [(152.69±15.83) RFU vs.(210.01±14.09) RFU] and MDA[(1.55±0.14) nmol/mg pro vs.(2.19±0.18) nmol/mg pro] in the INS group decreased,while SOD activity increased[(327.8±23.26) U/g pro vs.(238.49± 35.60) U/g pro],and the differences were significant(all P<0.05).(3)Compared with the CN group,the neuronal apoptosis index of cortex in the LPS group was elevated[(54.16±6.84)% vs.(5.45±1.43)%],while the expression of Bcl-2 decreased (627±0.018 vs.0.739±0.020),but the expressions of Bax(0.768±0.019 vs.0.520±0.010) and cleaved Caspase-9(0.739±0.016 vs.0.467±0.030) increased,and the differences were significant(all P<0.05).Compared with the LPS group,the neuronal apoptosis index of cortex in the INS group decreased [(33.30±3.07)% vs.(54.16±6.84)%],but the Bcl-2 expression increased (0.743±0.022 vs.0.627±0.018),and Bax (0.687±0.034 vs.0.768±0.019) and cleaved Caspase-9(0.551±0.013 vs.0.739±0.016) were reduced,and the differences were significant (all P<0.05).(4)Compared with the CN group,the mRNA (2.248±0.155 vs.1.000±0.100) and protein expression of UCP2 (0.659±0.016 vs.0.599±0.018) were elevated in the LPS group.Compared with the LPS group,the UCP2 mRNA (2.944±0.117 vs.2.248±0.155) and UCP2 protein (0.719±0.018 vs.0.659±0.016) increased,and the differences were significant(all P<0.05).Conclusions INS can protect the brain of septic rats through alleviating mitochondrial oxidative stress and inhibiting the mitochondrial-initiated apoptotic pathway to reduce neuronal apoptosis.INS upregulates UCP2 expression in the brain of septic rats,which may play a role in the protective effects mentioned above.

Effect and mechanism of uncoupling protein 2 on cardiomyocyte apoptosis induced by high glucose, high lipid and high uric acid / 解放军医学杂志

Objective To investigate the effects of uncoupling protein 2 (UCP2) on the myocardial cells of mice with type 2 diabetes mellitus combined with hyperuricemia (HUA), and clarify the mechanism thereof. Methods The mouse cardiac myocytes (MCM) cultured with 25mmol/L high glucose (HG) medium were divided into two groups: HG plus 300μmol/L sodium palmitate for 18 hours as high glucose and high fat (HG+HF) group, and HG+HF plus 1500μmol/L uric acid (UA) for 18 hours as HG+HF+HUA group. Then the myocardial cells in HG+HF+HUA group, by use or not use UCP2 inhibitor genipin, were further divided into two groups: vehicle group and genipin group. In order to verify the mechanism of UCP2 in myocardial cells injury caused by high glucose, high lipid and high uric acid, the myocardial cells were divided again into genipin group and genipin+N-acetylcysteine (NAC) group. Accordingly, the apoptosis of myocardial cells were measured by flow cytometry at specific time, the mRNA and protein expressions of UCP2 were determined by q-PCR and Western blotting, and the levels of reactive oxygen species (ROS) were detected by DHE staining and ELISA. Results The apoptosis rate of myocardial cells increased obviously, and the expression levels of UCP2 decreased and of ROS elevated significantly in HG+HF+HUA group than in HG+HF group (P<0.05). As the expression levels of UCP2 decreased by genipin intervention, the apoptosis rate of myocardial cells and ROS level in HG+HF+HUA group increased more obviously (P<0.05). In contrast, such an effect was reversed by the application of antioxidants NAC (P<0.05). Conclusion UCP2 can inhibit oxidative stress and alleviate the apoptosis of myocardial cells induced by high glucose, high fat and high uric acid.

Role and bioinformatics analysis of UCP2 in cervical cancer / 实用医学杂志

Objective To investigate the effect of UCP2 on cell proliferation, apoptosis, metastasis in cervical squamous carcinoma. Methods Plasmid?mediated downregulation of UCP2 was obtained in cervical cancer cell lines. MTT, flow cytometry and transwell chamber assay were conducted to detect the ability of proliferation, apoptosis and metastasis. Characteristics of UCP2 protein was analyzed by bioinformatics methods. Results (1)UCP2 was verified to be downregulated by qRT?PCR and Western blotting assay.(2)MTT, apoptosis assay and transwell chamber assay showed that the proliferation of SiHa cell was significantly inhibited, and the apoptosis rate was significantly increased, and metastasis was markedly deduced (P < 0.05). (3) Bioinformatic analysis showed that UCP2 was located in mitochondria with several phosphorylation sites, and UCP2 interacted with other proteins to produce biological effects. Conclusion UCP2 may play an important role in the occurrence and development of cervical cancer, and it is expected to be a new target for the early diagnosis and treatment of cervical cancer.

Antifibrotic effect of total flavonoids of Astmgali Radix on dimethylnitrosamine-induced liver cirrhosis in rats / 中国结合医学杂志

<p><b>OBJECTIVE</b>To study the effect of total flavonoids of Astmgali Radix (TFA) on liver cirrhosis induced with dimethylnitrosamine (DMN) in rats, and the effect on peroxisome proliferator-activated receptor γ (PPARγ), uncoupling protein 2 (UCP2) and farnesoid X receptor (FXR).</p><p><b>METHODS</b>Fifty-three Sprague-Dawley rats were randomly divided into a control group (10 rats) and a DMN group (43 rats). Rats in the DMN group were given DMN for 4 weeks and divided randomly into a model group (14 rats), a low-dosage TFA group (14 rats) and a high-dosage TFA group (15 rats) in the 3rd week. Rats were given TFA for 4 weeks at the dosage of 15 and 30 mg/kg in the low- and high-TFA groups, respectively. At the end of the experiment blood and liver samples were collected. Serum liver function and liver tissue hydroxyproline content were determined. hematoxylin-eosin (HE), Sirus red and immunohistochemical stainings of collagen I, smooth muscle actin (α-SMA) was conducted in paraffinembedded liver tissue slices. Real time polymerase chain reaction (PCR) was adopted to determine PPARγ, UCP2 and FXR mRNA levels. Western blot was adopted to determine protein levels of collagen I, α-SMA, PPARγ, UCP2 and FXR.</p><p><b>RESULTS</b>Compared with the model group, TFA increased the ratio of liver/body weight (low-TFA group P<0.05, high-TFA group P<0.01), improved liver biochemical indices (P<0.01 for ALT, AST, GGT in both groups, P<0.05 for albumin and TBil in the high-TFA group) and reduced liver tissue hydroxproline content (P<0.01 in both groups) in treatment groups significantly. HE staining showed that TFA alleviated liver pathological changes markedly and Sirus red staining showed that TFA reduced collagen deposition, alleviated formation and extent of liver pseudolobule. Collagen I and α-SMA immunohistochemical staining showed that staining area and extent markedly decreased in TFA groups compared with the model group. TFA could increase PPARγ, it regulated target UCP2, and FXR levels significantly compared with the model group (in the low-TFA group all P<0.05, in the high group all P<0.01).</p><p><b>CONCLUSION</b>TFA could improve liver function, alleviate liver pathological changes, and reduce collagen deposition and formation of liver pseudolobule in rats with liver cirrhosis. The antifibrotic effect of TFA was through regulating PPARγ signal pathway and the interaction with FXR.</p>

Effects of Berberine on Hepatic Sirtuin 1-uncoupling Protein 2 Pathway in Non-alcoholic Fatty Liver Disease Rats Induced by High-fat Diet / 中草药·英文版

Objective To investigate the involvement of sirtuin 1 (SIRT1)-uncoupling protein 2 (UCP2) pathway in the development of non-alcoholic fatty liver disease and whether berberine exerts its effects by regulating this pathway. Methods Male SD rats were divided into three groups: normal control group, high-fat diet group, and berberine supplement group. The rats in the normal control group were given normal diet while the rats in the other two groups were fed with high-fat diet. Rats in the berberine supplement group were concurrently given berberine (100 mg/kg body weight) once daily. After 16 weeks, the levels of serum, liver lipids, and serum aminotransferase were measured using an automatic biochemical analyzer. Superoxide dismutase (SOD) activity and malondialdehyde (MDA) content in the liver were measured using commercial kits. Histopathological changes of liver tissues were observed by hematoxylin and eosin (HE) staining and Oil Red O staining. The hepatic mRNA and protein levels of SIRT1 and UCP2 were assayed by reverse transcription polymerase chain reaction (RT-PCR) or Western blotting. Results Berberine supplement could significantly decrease the serum and liver lipid contents in rats fed with high-fat diet. Meanwhile, SOD level was significantly elevated, but MDA level was reduced in the liver. The results of HE and Oil Red O staining showed that the hepatic steatosis was alleviated in berberine supplement group. Furthermore, berberine induced an increase in SIRT1 expression but a decrease in UCP2 expression. Conclusion The regulation of hepatic SIRT1-UCP2 pathway may be an important mechanism by which berberine exerts the beneficial effects in NAFLD rats.

Protective effect of trimetazidine for the rat myocardial tissue after exhaustion exercise / 解放军医学杂志

Objective To explore the protective effect of trimetazidine on the myocardial tissue of rats by observing the oxidative stress, apoptosis and energy metabolism in myocardial tissue of rats after exhausted exercise. Methods Thirty healthy adult male Wistar rats were randomly assigned to three groups (10 each): quiet control group (C), exhausted exercise group (E) and exhausted exercise plus trimetazidine group (TE). Exhausted exercise model was established by forcing the rats swim ceaselessly, rats in group TE received hydrochloric acid trimetazidine (10mg/kg) for intervention. The myocardial tissue of rats was collected after the last exhausted exercise. The myocardial tissue of rats was evaluated by HE staining. The concentrations of superoxide dismutase (SOD), glutathione peroxidase 1 (GSH-Px) and malondialdehyde (MDA) in rats' myocardial cell mitochondria were detected by spectrophotometry. RT-PCR and immunohistochemistry were performed to quantify the mRNA and protein levels of Bax, Bcl-2 and uncoupling protein 2 (UCP2) in myocardial tissue of rats. Terminal-deoxynucleoitidyl Transferase Mediated Nick End Labeling (TUNEL) assay was employed to detect the apoptotic rate of myocardial tissue. Results Compared with group C, the SOD and GSH-Px activities in groups E and TE were reduced (P<0.01), while the MDA activity was increased (P<0.01), and more obvious changes were in group E. The SOD and GSH-Px activities decreased, while the MDA activity increased in group E than in group TE (P<0.01). Compared with group C, the apoptotic rate of myocardial tissue increased in groups E and TE (P<0.01), and the highest apoptotic rate was in group E (P<0.01). Furthermore, the mRNA and protein levels of Bax were significantly elevated in groups E and TE (P<0.01), and the highest expression was in group E (P<0.01). The mRNA and protein levels of BCL-2 were significantly reduced in groups E and TE (P<0.01), and the lowest expression was in group E (P<0.01). Meanwhile, compared with group C, the mRNA and protein expression of UCP2 was significantly increased in groups E and TE (P<0.01). After treatment with hydrochloric acid trimetazidine, the UCP2 expression in group TE was relative decreased than in group E (P<0.01). Conclusion Trimetazidine can reduce the oxidative stress and apoptotic rate of myocardial tissue of rats after exhausted exercise, indicating it plays an important protective role for the myocardial tissue of rats.

Uncoupling protein 2 in regulatory mechanisms and basical experiments / 国际儿科学杂志

Uncoupling protein 2(UCP2)is a proton transporter which presents in the mitochondrial inner membrane. Recent studies have found that UCP2 plays important roles in protecting mitochondria functions,re-ducing mitochondrial ROS generation by inducing proton leak across the inner mitochondrial membrane and pro-moting mild uncoupling which leading to a decrease in the mitochondrial inner membrane′s potential. Because of its antioxidant function,UCP2 has been studied in several domains. This review provides novel insights into the molecular mechanisms,by which the activity of UCP2 is regulated and describe novel findings concerning basical experiments of UCP2.

The role of peroxisome proliferator-activated receptor-γ coactivator-1 in glucose metabolism in the liver of OLETF rat / 实用医学杂志

Objective To investigate the effect of PGC-1 on hepatic glucose metabolism of type 2 diabetes by observing its changes in the liver of OLETF rats. Methods OLETF rats were observed,even-aged LETO rats were controled. Oral glucose tolerance test, Fasting Insulin, triglyceride and total cholesterol were measured and then the protein level of PGC-1 , phosphoenolpyruvate earboxykinase and uncoupling protein 2 of liver tissue were detected by Western blot respectively in 8,18 and 28 weeks. Results (1)OLETF rats had significantly higher levels than LETO rats, in body weight, OGTT2h blood glucose and TG at 18th, 28th week. The levels of Fasting Insulin of OLETF rats were higher while insulin sensitive index were lower than that of LETO rats at 28th week. (2)Protein expressions in the livers: PGC-1 of OLETF rats was lower than that of LETO rats at 18th and 28th week. PEPCK of OLETF rats was more while UCP2 was lower than that of LETO rats at 28th week. Conclusions OLETF rats showed pathological phenotypes of type 2 diabetes. The changes of PGC-1 , PEPCK and UCP2 in 28-weeks OLETF rat suggested that PGC-1 plays an important role in the liver glycometabolism in type 2 diabetes.

Antitumor Effects of Genipin: New and Emerging Insights from Recent Studies

Genipin, an aglycone derived from geniposide found in Gardenia jasminoides, is known to be an excellent natural cross-linker, strong apoptosis inducer, and antiviral agent. Although evidence suggests antiviral activity of genipin in several in vitro viral infection systems, there have been few literatures which review antitumor effects of genipin in a variety of in vitro/in vivo models of cancers yet. In this review, we present some of the latest findings in the studies of genipin focusing on antitumor effects and its mechanisms. In brief, genipin inhibits mitochondrial uncoupling protein 2 to increase accumulation of reactive oxygen species, leading to ROS/c-Jun N-terminal kinase-dependent apoptosis of cancer cells. Genipin also increase tissue inhibitors of metalloproteases (MMP), resulting to decrease activities of MMP-2 which plays a key role in metastasis of cancers. Genipin has shown a biphasic effects on cell death and survival in cancer cells as many other plant-derived phytochemicals do. Finally we discuss the potential of genipin as a promosing novel antitumor agent which could be applicable to chemotherapy and/or chemoprevention for cancers.

PPARγagonist inhibits high glucose-induced production of reactive oxy-gen species by UCP2 up-regulation / 中国病理生理杂志

AIM:To explore the effects of PPARγon the elevated level of reactive oxygen species ( ROS) in-duced by high glucose and its mechanism .METHODS:Human umbilical vein endothelial cells ( HUVECs) were cultured with DMEM containing high glucose (33 mmol/L D-glucose), and DMEM containing lower glucose (5.5 mmol/L D-glu-cose) was used as control .Superoxide anion and nitric oxide fluorescence probes were used to observe the effects of PPAR γagonist on ROS and NO productions in the HUVECs .The uncoupling protein 2 (UCP2) protein level in the HUVECs was detected by Western blotting .RESULTS:PPARγagonist pioglitazone inhibited the ROS generation and prevented the de-crease in NO level under high glucose condition , and these effects were reversed by pretreatment with PPARγantagonist GW9662.The results of Western blotting indicated that PPARγagonist pioglitazone up-regulated the UCP2 expression un-der high glucose condition , and this effect was also blocked by GW 9662.Inhibition of UCP2 by genipin attenuated the effect of pioglotazone on the ROS production .CONCLUSION: Activation of PPARγinhibits ROS generation under high glucose condition , and this effect may mediate by up-regulation of UCP2.

Uncoupling protein 2 gene (UCP2) 45-bp I/D polymorphism is associated with adiposity among Malaysian women.

This study investigated the association of Uncoupling Protein 2 gene (UCP2) 45-bp I/D polymorphism with obesity and adiposity in 926 Malaysian subjects (416 males; 265 obese; 102/672/152 Malays/Chinese/Indians). The overall minor allele frequency (MAF) was 0.14, while MAFs according to Malay/Chinese/Indian were 0.17/0.12/0.21. The polymorphism was associated with ethnicity, obesity and overall adiposity (total body fat percentage, TBF), but not gender and central adiposity (waist–hip ratio, WHR). Gender- and ethnicity-stratified analysis revealed that within males, the polymorphism was not associated with ethnicity and anthropometric classes. However, within females, significantly more Indians, obese and those with high TBF carried I allele. Logistic regression analysis among females further showed the polymorphism was associated with obesity and overall adiposity; however, when adjusted for age and ethnicity, this association was abolished for obesity but remained significant for overall adiposity [Odds Ratio (OR) for ID genotype =2.02 (CI=1.18, 3.45; p=0.01); I allele =1.81 (CI=1.15, 2.84; p=0.01)]. Indeed, covariate analysis controlling for age and ethnicity also showed that those carrying ID genotype or I allele had significantly higher TBF than the rest. In conclusion, UCP2 45-bp I/D polymorphism is associated with overall adiposity among Malaysian women.