UCP2 overexpression activates SIRT3 to regulate oxidative stress and mitochondrial dynamics induced by myocardial injury.

OBJECTIVE: Our previous study found that overexpression of uncoupling protein-2 (UCP2) had a protective effect on lipopolysaccharide (LPS)-induced sepsis cardiomyocytes. The aim of this study was to explore the effect and mechanism of uncoupling protein-2 (UCP2) on myocardial ischemia-reperfusion injury. METHODS: In this study, we established hypoxia-reoxygenation (HR) injury model in rats and isolated cardiomyocytes of newborn rats. We also carried out following methods which include virus transfection technology, cell counting Kit-8 (CCK8), flow cytometry, enzyme linked immunosorbent assay (ELISA), Western blot (WB), quantitative reverse transcription PCR (RT qPCR), transmission electron microscopy, fluorescence colocalization and immunoprecipitation. MAIN RESULTS: The results of this study showed that hypoxia-reoxygenation treatment in cardiomyocytes increased UCP2, myocardial enzyme and myocardial apoptosis and weakened cardiomyocyte viability. We observed increased cardiomyocyte viability and mitochondrial membrane potential, decreased myocardial enzyme and myocardial apoptosis, Inhibition of oxidative stress when UCP2 was overexpressed in cardiomyocytes. It also can Increase ATP and stabilize mitochondrial dynamics. Further studies founded that Sirtuin-3(SIRT3) changed with the expression of UCP2, which was confirmed by fluorescence co-localization and immunoprecipitation. CONCLUSIONS: Our findings revealed that UCP2 and SIRT3 were important targets of anti-myocardial injury by inhibiting cellular oxidative stress and stabilizing mitochondrial dynamics.

IMPLICATIONS OF YWHAH GENE EXPRESSION IN THE EARLY DETECTION OF SEPSIS.

ABSTRACT: Sepsis, a complex and multifaceted condition, is a common occurrence with serious implications for critically ill patients in the intensive care unit (ICU). The YWHAH gene encodes the 14-3-3n protein, a member of the 14-3-3 protein family. While existing research primarily focuses on the role of 14-3-3n in conditions such as schizophrenia and various cancers, our study revealed that the expression of the YWHAH gene remained relatively stable in both infected individuals and healthy controls. Through Venn plot analysis following weighted gene correlation network analysis, we observed a potential association between elevated YWHAH expression and the transition from infection to sepsis. In a comprehensive analysis of public single-cell transcriptome databases, the expression of YWHAH was found to be distinctive in cases of sepsis and infection. These findings were corroborated through an in vitro analysis utilizing real-time polymerase chain reaction. This study represents the initial identification of variations in YWHAH gene expression between patients with infection and sepsis, potentially offering insights for the development of early detection and treatment strategies for sepsis.

Graves' hyperthyroidism induced pancytopenia, epilepsia and muscle weakness: A case report.

RATIONALE: Pancytopenia and epilepsia are rare complications of Graves' disease (GD). Muscle weakness is a physical sign of GD. It is extremely rare for GD patients to present 3 symptoms at the same time. PATIENT CONCERNS: A 35-year-old female was admitted to hospital for dizziness for 1 day. The results of laboratory examination on admission showed pancytopenia and hypothyroidism. Her clinical manifestations include pancytopenia, epilepsy, and muscle weakness. DIAGNOSIS: Graves' hyperthyroidism. INTERVENTIONS: She received endotracheal intubation, ventilator, antithyroid drugs, and hormone therapy. OUTCOME: The patient was discharged after treatment. LESSON: Severe complications caused by GD are rare and require antithyroid therapy. Although glucocorticoid is not recommended by the guidelines, it can effectively improve thrombocytopenia.

[Characteristics of abdominal hemorrhage in patients with severe acute pancreatitis and its influence on outcomes].

OBJECTIVE: To explore the risk factors of abdominal hemorrhage (AH) in patients with severe acute pancreatitis (SAP) and its impact on outcome. METHODS: The clinical data of 231 SAP patients admitted to Diagnosis and Treatment Center for SAP of Guizhou Province from January 1, 2015 to December 31, 2019 were retrospectively analyzed. These patients were divided into AH group and non-AH group. The general information, etiology, acute physiology and chronic health evaluation II (APACHE II) score, sequential organ failure assessment (SOFA) score, organ failure, complications, interventions, bleeding time, bleeding site and outcome were compared between the two groups. Binary multivariate Logistic regression analysis was used to explore the risk factors of AH in SAP patients and whether the time and location of AH were risk factors affecting the outcome. RESULTS: A total of 231 patients were enrolled in the analysis, including 198 patients without AH and 33 with AH (14.3%). There was no significant difference in gender, age or etiology between the two groups. The scores of APACHE II and SOFA in AH group were significantly higher than those in non-AH group [APACHE II score: 18 (12, 24) vs. 13 (9, 19), SOFA score: 9 (5, 15) vs. 5 (4, 11), both P < 0.01]. The incidences of acute kidney injury (AKI), gastrointestinal dysfunction, coagulation disorders, necrotic infection, pseudocyst and gastrointestinal fistula in AH group were significantly higher than those in non-AH group (66.7% vs. 47.0%, 36.4% vs. 7.1%, 18.2% vs. 6.6%, 66.7% vs. 9.1%, 66.7% vs. 34.3%, 9.1% vs. 1.5%, all P < 0.05). The proportions of requiring mechanical ventilation (MV) and surgical intervention in AH group were significantly higher than those in non-AH group (69.7% vs. 43.4, 48.5% vs. 14.6%, both P < 0.01). The length of intensive care unit (ICU) stay and hospital stay in AH group were significantly longer than those in non-AH group [length of ICU stay (days): 13 (8, 19) vs. 7 (3, 16), length of hospital stay: 24 (13, 40) vs. 17 (12, 24), both P < 0.01], and the hospital mortality was significantly higher (60.6% vs. 9.6%, P < 0.01). Multivariate Logistic regression analysis showed that APACHE II score [odds ratio (OR) = 1.157, 95% confidence interval (95%CI) was 1.030-1.299, P = 0.014], infectious necrosis (OR = 12.211, 95%CI was 4.063-36.697, P < 0.01), pseudocyst (OR = 3.568, 95%CI was 1.238-10.283, P = 0.019) and requiring MV (OR = 0.089, 95%CI was 1.354-6.625, P = 0.007) were the risk factors of AH in SAP patients. In 33 AH patients, there was no significant difference in hospital mortality between early hemorrhage (occurred within 2 weeks of onset) and late hemorrhage (occurred 2 weeks after onset) groups [66.7% (8/12) vs. 57.1% (12/21), P > 0.05]. All 4 patients in the unspecified bleeding site group died during hospitalization; half or more patients died in the pseudocyst/abscess bleeding (14 cases), mesenteric/intestinal bleeding (13 cases) and gastric variceal bleeding (2 cases) groups (7 cases, 8 cases and 1 case respectively), and there were significant differences among the groups (P < 0.05). Multivariate Logistic regression analysis showed that neither bleeding time (OR = 0.989, 95%CI was 0.951-1.028, P = 0.574) nor bleeding site (OR = 2.009, 95%CI was 0.822-4.907, P = 0.126) was the risk factor of death in patients with SAP combined with AH. CONCLUSIONS: Both early and late bleeding significantly increased the length of hospital stay and mortality of SAP patients. APACHE II score, infectious necrosis and pseudocyst were the risk factors of AH in SAP patients. Neither bleeding time nor bleeding site was the risk factors of death in patients with SAP combined with AH. However, it still needed to be confirmed by a large sample clinical study.

[Uncoupling protein 2 overexpression alleviates sepsis-induced myocardial injury via inhibiting reactive oxygen species production and inflammation].

OBJECTIVE: To investigate whether the overexpression of uncoupling protein 2 (UCP2) can protect myocardium from sepsis by inhibiting the production of reactive oxygen species (ROS) and inflammatory response. METHODS: Forty Sprague-Dawley rats were divided into four groups according to random number table method (n = 10): sham transfection and sham surgery group (Sham group), sham transfection and cecal ligation and perforation (CLP) group (CLP group), simple adeno-associated virus (AAV) transfection surgery group (AAV group), and UCP2 overexpression surgery group (UCP2 group). In UCP2 group, UCP2 adeno-associated virus (AAV-UCP2; titer 1×1012 v.g/mL, 10 µL per site, 60 µL in total) was injected into myocardium, and CLP was performed 3 weeks later. In AAV group, the myocardium was transfected with AAV virus and CLP was performed 3 weeks later. Twenty-four hours after modeling, whether the model was successfully prepared was evaluated. The transfection effect of AAV virus on the frozen sections of myocardial tissue was observed under fluorescence microscope, the expression of UCP2 protein was detected by Western blotting, ROS production was detected by dihydroethidine (DHE) staining, and serum myocardial markers and inflammatory cytokines were detected by enzyme linked immunosorbent assay (ELISA). RESULTS: Twenty-four hours after CLP, the rats showed stiff hair, increased secretions from eyes, nose and mouth, and symptoms of pyuria, loose stools, and dyspnea. After laparotomy, the cecum showed purple and black, and there was purulent exudation around the intestinal cavity. The virus was successfully transfected on frozen section under the fluorescence microscope (the site of the transfection was green fluorescence), and further Western blotting revealed that the expression of UCP2 in the CLP group was higher than that in the Sham group (UCP2/ß-tubulin: 1.53±0.06 vs. 1, P < 0.01). Compared with the AAV group, UCP2 expression was further increased in the UCP2 group (UCP2/ß-tubulin: 1.96±0.22 vs. 1.59±0.07, P < 0.01). Under the fluorescence microscope, ROS production in the CLP and AAV groups were found significantly increased compared with that in the Sham group; when UCP2 was overexpressed, ROS production were significantly decreased compared with the CLP and AAV groups (A value: 1.03±0.10 vs. 1.81±0.13, 1.67±0.08, both P < 0.01). ELISA showed that compared with the Sham group, the levels of lactate dehydrogenase (LDH), creatine kinase (CK), cardiac troponin I (cTnI), tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6) were significantly increased in the CLP and AAV groups; when UCP2 was overexpressed, the above myocardial enzymes and inflammatory cytokines secretion were significantly decreased compared with the CLP group and AAV group [LDH (ng/L): 48.97±1.04 vs. 56.85±1.36, 57.08±1.54; CK (ng/L): 235.23±20.33 vs. 306.34±25.93, 304.76±25.29; cTnI (ng/L): 199.79±18.27 vs. 241.88±14.32, 243.33±23.79; TNF-α (ng/L): 385.71±20.09 vs. 488.92±26.92, 489.03±33.37; IL-6 (ng/L): 121.12±7.61 vs. 159.07±17.65, 157.61±15.13; all P < 0.01]. Kaplan-Meier survival curve showed that the survival rate of rats 36 hours after CLP was only 30.0%. When UCP2 overexpressed, the survival rate was significantly higher than that of the CLP group and AAV group (60.0% vs. 30.0%, 30.0%, both P < 0.05). There was no significant difference between the AAV group and CLP group. CONCLUSIONS: UCP2 overexpression can reduce myocardial injury and improve the survival rate of septic rats by reducing ROS production and inhibiting inflammatory reaction in septic myocardium.

[Effects of uncoupling protein 2 overexpression on myocardial mitochondrial dynamics in sepsis rats].

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: (1) The 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/ß-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). CONCLUSIONS: 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.