Objective To observe the skin ultrastructure change of electric shock death rats and to test the expression changes of hypoxia-inducible factor-2α （HIF-2α） and heart type-fatty acid-binding protein （H-FABP） of myocardial cells, in order to provide basis for forensic identification of electric shock death. Methods The electric shock model of rats was established. The 72 rats were randomly divided into control group, electric shock death group and postmortem electric shock group. Each group was divided into three subgroups, immediate （0 min）, 30 min and 60 min after death. The skin changes of rats were observed by HE staining, the changes of skin ultrastructure were observed by scanning electron microscopy, and the expression of HIF-2α and H-FABP in rats myocardium was tested by immunohistochemical staining. Results The skin in the electric shock death group and postmortem electric shock group had no significant difference through the naked eye or by HE staining. Under the scanning electron microscope, a large number of cellular debris, cells with unclear boundaries, withered cracks, circular or elliptical holes scattered on the cell surface and irregular edges were observed. A large number of spherical foreign body particles were observed. Compared with the control group, the expression of HIF-2α in all electric shock death subgroups increased, reaching the peak immediately after death. In the postmortem electric shock group, HIF-2α expression only increased immediately after death, but was lower than that of electric shock death group （P<0.05）. Compared with the control group, the expression of H-FABP in all subgroups of electric shock death group and postmortem electric shock group significantly decreased. The expression of H-FABP in all subgroups of electric shock death group was lower than that of the postmortem electric shock group （P<0.05）. Conclusion Electric shock can increase HIF-2α expression and decrease H-FABP expression in the myocardium, which may be of forensic significance for the determination of electric shock death and identification of antemortem and postmortem electric shock.
Animals ; Autopsy ; Basic Helix-Loop-Helix Transcription Factors/metabolism* ; Fatty Acid Binding Protein 3/metabolism* ; Myocardium/metabolism* ; Myocytes, Cardiac/metabolism* ; Rats ; Skin/ultrastructure*

OBJECTIVETo investigate the relationship between serum high-sensitivity C-reaction protein (hs-CRP) and heart fatty acid binding protein (h-FABP) on cardiac accidents in patients with unstable angina pectoris (UAP).

METHODSerum levels of hs-CRP, h-FABP, cardiac troponin-I (cTn-I) and creatine kinase MB isoenzyme (CK-MB) were measured and cardiac accidents within 2 weeks after the test were observed in 74 patients (male 45) with stable AP (SAP) and 56 patients (male 29) with UAP.

RESULTSThe incidence of cardiac accidents was significantly higher in UAP group (26.8%) than that in SAP group (10.53%, P < 0.001). Serum hs-CRP [(7.64 +/- 2.18) mg/L vs. (1.78 +/- 0.62) mg/L, P < 0.001], h-FABP [(16.46 +/- 5.28) microg/L vs. (3.15 +/- 2.61) microg/L, P < 0.001] and cTn-I [(1.28 +/- 0.43) microg/L vs. (0.67 +/- 0.09) microg/L, P < 0.001] levels were also significantly higher in UAP group than those in SAP group. The serum hs-CRP and h-FABP levels for patients with cardiac accidents in the SAP group were (6.32 +/- 2.06) microg/L and (8.76 +/- 3.83) microg/L respectively, which were higher than those for the patients having no cardiac accidents in the control (P < 0.01). The serum hs-CRP, h-FABP, cTn-I and CK-MB levels in patients with cardiac accidents were significantly higher than those in patients without cardiac accidents in both SAP and UAP groups.

CONCLUSIONMeasuring traditional parameters for myocardial damage (cTn-I and CK-MB) in combination with hs-CRP and h-FABP is valuable for predicting the risk of recent cardiac accidents for AP patients.

Adult ; Aged ; Aged, 80 and over ; Angina, Unstable ; blood ; diagnostic imaging ; C-Reactive Protein ; metabolism ; Case-Control Studies ; Coronary Angiography ; Fatty Acid Binding Protein 3 ; Fatty Acid-Binding Proteins ; blood ; Female ; Humans ; Male ; Middle Aged

OBJECTIVE: To investigate the changes in myocardial calcium currents in rats subjected to forced running exercise during acute hypoxia and their association with myocardial injury. METHODS: Forty SD rats were randomized into quiescent group and running group either in normal oxygen (NQ and NR groups, respectively) or in acute hypoxia (HQ and HR groups, respectively). Hypoxia was induced by keeping the rats in a hypobaric oxygen chamber (PaO₂=61.6kpa) for 4 h a day; the rats in the two running groups were forced to run on running wheels for 4 h each day. Rat ventricular myocytes was isolated by enzymatic digestion for recording action potentials and currents using patch clamp technique, and confocal Ca²⁺ imaging was used to monitor intracellular Ca²⁺ levels. The expressions of Cav1.2 channel and the cardiac ryanodine receptor (RyR2) were determined using Western blotting. RESULTS: Compared with those in NQ group, the rats in HR group showed significantly decreased SOD activity (P < 0.01), increased h-FABP, hs-CRP and IMA levels (P < 0.05 or 0.01), obvious myocardial pathology, and prolonged APD50 and APD90 (P < 0.05). Of the different stress conditions, forced running in acute hypoxia resulted in the most prominent increase of the densities of ICa, L currents, causing also a significant left shift of the steady state activation curve and a significant right shift of the steady state inactivation curve. Compared with those in NQ group, the rats in NR, HQ and HR groups all exhibited higher rates of spontaneous calcium wave events in the cardiac myocytes, increased frequency of calcium sparks with lowered amplitude, enhanced calcium release amplitude in the ventricular myocytes, and delayed calcium ion reabsorption; in particular, these changes were the most conspicuous in HR group (P < 0.05 or 0.01). There was also a significant increase in the protein levels of Cav1.2 channel and RyR2 receptor in HR group (P < 0.05 or 0.01). CONCLUSIONS The mechanism of myocardial injury in rats subjected to forced running in acute hypoxia may involve the increase of oxidative stress and calcium current and intracellular calcium overload.
Animals ; C-Reactive Protein/metabolism* ; Calcium/metabolism* ; Calcium Signaling ; Fatty Acid Binding Protein 3/metabolism* ; Heart Injuries/metabolism* ; Hypoxia/metabolism* ; Myocytes, Cardiac/metabolism* ; Oxygen/metabolism* ; Rats ; Rats, Sprague-Dawley ; Ryanodine Receptor Calcium Release Channel/metabolism* ; Superoxide Dismutase/metabolism*

OBJECTIVE: To observe the protective effect of heart-fatty acid binding protein (H-FABP) on lipopolysaccharide (LPS)-induced cardiomyocyte damage. METHODS: The cardiomyocytes were isolated and cultured from 1-3 days old neonatal rats. The specific siRNA or plasmid of H-FABP were transfected into cells to alter H-FABP expression, which was evaluated by Western blot and quantitative-PCR. LPS-induced cardiomyocyte damage and inflammation were estimated by detecting the contents of lactate dehydrogenase(LDH), TNF-α, and IL-1β as well as cell viability. RESULTS: LPS treatment induced inflammation and cell damage indicated by a decrease in cell viability and an increase in LDH, TNF-α and IL-1β in the medium. When H-FABP was downregulated by siRNA transfection, the LPS-induced inflammation and cell damage were augmented. In contrast, when H-FABP was overexpressed by pcDNA3.1-H-FABP transfection, the LPS-induced inflammation and cell damage were suppressed. CONCLUSION H-FABP protects cardiomyocytes from LPS-induced inflammation and cell injury.
Animals ; Animals, Newborn ; Cell Line ; Cell Survival ; Down-Regulation ; Fatty Acid Binding Protein 3 ; Fatty Acid-Binding Proteins ; metabolism ; Inflammation ; metabolism ; Interleukin-1beta ; metabolism ; L-Lactate Dehydrogenase ; metabolism ; Lipopolysaccharides ; adverse effects ; Myocytes, Cardiac ; cytology ; drug effects ; RNA, Small Interfering ; genetics ; Rats ; Transfection ; Tumor Necrosis Factor-alpha ; metabolism

Escherichia coli (E. coli) FadR regulator plays dual roles in fatty acid metabolism, which not only represses the fatty acid degradation (fad) system, but also activates the unsaturated fatty acid synthesis pathway. Earlier structural and biochemical studies of FadR protein have provided insights into interplay between FadR protein with its DNA target and/or ligand, while the missing knowledge gap (esp. residues with indirect roles in DNA binding) remains unclear. Here we report this case through deep mapping of old E. coli fadR mutants accumulated. Molecular dissection of E. coli K113 strain, a fadR mutant that can grow on decanoic acid (C10) as sole carbon sources unexpectedly revealed a single point mutation of T178G in fadR locus (W60G in FadRk113). We also observed that a single genetically-recessive mutation of W60G in FadR regulatory protein can lead to loss of its DNA-binding activity, and thereby impair all the regulatory roles in fatty acid metabolisms. Structural analyses of FadR protein indicated that the hydrophobic interaction amongst the three amino acids (W60, F74 and W75) is critical for its DNA-binding ability by maintaining the configuration of its neighboring two β-sheets. Further site-directed mutagenesis analyses demonstrated that the FadR mutants (F74G and/or W75G) do not exhibit the detected DNA-binding activity, validating above structural reasoning.
3-Oxoacyl-(Acyl-Carrier-Protein) Synthase ; genetics ; metabolism ; Amino Acid Sequence ; Bacterial Proteins ; chemistry ; genetics ; metabolism ; DNA, Bacterial ; chemistry ; metabolism ; Escherichia coli ; genetics ; metabolism ; Escherichia coli Proteins ; genetics ; metabolism ; Fatty Acid Synthase, Type II ; genetics ; metabolism ; Fatty Acids ; metabolism ; Gene Expression Regulation, Bacterial ; Hydro-Lyases ; genetics ; metabolism ; Hydrophobic and Hydrophilic Interactions ; Lipid Metabolism ; Models, Molecular ; Molecular Sequence Data ; Mutation ; Protein Binding ; Protein Structure, Secondary ; Repressor Proteins ; chemistry ; genetics ; metabolism ; Sequence Alignment ; Sequence Homology, Amino Acid ; Signal Transduction

OBJECTIVETo identify the underlying mechanisms of the protective effects of Dingxin Recipe (: , DXR), a Chinese compound prescription that has been used clinically in China for more than 20 years, on ischemia/reperfusion (I/R)-induced arrhythmias in rat model.

METHODSA total of 30 rats were randomly divided into three groups: sham group, I/R group, and DXR-pretreated I/R (DXR-I/R) group. Rats in the DXR-DXRI/R group were intragastrically administrated with DXR (12.5 g/kg per day) for consecutive 7 days, while rats I/in the sham and I/R groups were administrated with normal saline. Arrhythmias were introduced by I/R and electrocardiograms (ECG) were recorded. Two-dimensional (2-D) polyacrylamide gel electrophoresis and matrix-matrix assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF MS) were used to identify assisted differentially expressed proteins. Immunohistochemistry, real-time quantitative polymerase chain reaction (RQ-RQPCR), Western blot, and enzyme-linked immunosorbent assay (ELISA) were performed to analyze proteins PCR), obtained in the above experiments.

RESULTSDXR significantly reduced the incidence and mean duration of ventricular tachycardia and ventricular fibrillation and dramatically decreased the mortality, as well as arrhythmia score, compared with those of the I/R group. Among successfully identified proteins, prohibitin (PHB) and heart fatty acid binding protein (hFABP) were up-regulated in DXR-pretreated I/R rats compared with those of the I/R rats. In addition, compared with the I/R group, the level of glutathione (GSH) was elevated accompanied by reduced expressions of interleukin-6 (IL-6) and neutrophil infiltration in I/R rats with DXR pretreatment.

CONCLUSIONSDXR could alleviate I/R-induced arrhythmias, which might be related to increased expression of PHB. The enhanced expression of PHB prevented against the depletion of GSH and consequently inhibited apoptosis of cardiomyocytes. Furthermore, up-regulation of PHB might ameliorate I/R-induced cell death and leakage of hFABP by suppressing neutrophil infiltration and IL-6 expressions.

Animals ; Arrhythmias, Cardiac ; etiology ; prevention & control ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Electrophoresis, Gel, Two-Dimensional ; Fatty Acid Binding Protein 3 ; Fatty Acid-Binding Proteins ; metabolism ; Glutathione ; metabolism ; Heart Ventricles ; drug effects ; metabolism ; pathology ; Immunohistochemistry ; Inflammation ; complications ; metabolism ; pathology ; Interleukin-6 ; metabolism ; Male ; Myocardial Infarction ; complications ; drug therapy ; pathology ; Neutrophil Infiltration ; drug effects ; Peptide Mapping ; Proteomics ; Rats ; Rats, Wistar ; Reperfusion Injury ; complications ; Repressor Proteins ; metabolism ; Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization ; Spectrophotometry ; Up-Regulation ; drug effects

Serum palmitic acid (PA), a type of saturated fatty acid, causes lipid accumulation and induces toxicity in hepatocytes. Ethanol (EtOH) is metabolized by the liver and induces hepatic injury and inflammation. Herein, we analyzed the effects of EtOH on PA-induced lipotoxicity in the liver. Our results indicated that EtOH aggravated PA-induced apoptosis and lipid accumulation in primary rat hepatocytes in dose-dependent manner. EtOH intensified PA-caused endoplasmic reticulum (ER) stress response in vitro and in vivo, and the expressions of CHOP, ATF4, and XBP-1 in nucleus were significantly increased. EtOH also increased PA-caused cleaved caspase-3 in cytoplasm. In wild type and CHOP(-/-) mice treated with EtOH and high fat diet (HFD), EtOH worsened the HFD-induced liver injury and dyslipidemia, while CHOP knockout blocked toxic effects of EtOH and PA. Our study suggested that targeting UPR-signaling pathways is a promising, novel approach to reducing EtOH and saturated fatty acid-induced metabolic complications.
Activating Transcription Factor 4 ; drug effects ; metabolism ; Animals ; Apoptosis ; drug effects ; Caspase 3 ; drug effects ; Chemical and Drug Induced Liver Injury ; metabolism ; DNA-Binding Proteins ; drug effects ; metabolism ; Diet, High-Fat ; adverse effects ; Dose-Response Relationship, Drug ; Dyslipidemias ; chemically induced ; metabolism ; Endoplasmic Reticulum Stress ; drug effects ; Ethanol ; metabolism ; toxicity ; Fatty Liver ; chemically induced ; metabolism ; Gene Knockout Techniques ; Hepatocytes ; drug effects ; metabolism ; Lipid Metabolism ; drug effects ; Liver ; metabolism ; Male ; Mice ; Palmitic Acid ; toxicity ; Rats ; Rats, Sprague-Dawley ; Regulatory Factor X Transcription Factors ; Signal Transduction ; drug effects ; Transcription Factor CHOP ; drug effects ; genetics ; metabolism ; Transcription Factors ; drug effects ; metabolism ; Unfolded Protein Response ; drug effects ; X-Box Binding Protein 1