1.Comparative Study in DNA-mediated Vaccination Efficaency Among the Plasmids with Different Promoters.
Sun Hwa CHANG ; Koo Nam YANG ; Yong Suk JANG
Korean Journal of Immunology 1998;20(4):375-379
Plasmid vectors with either RSV or CMV promoter are frequently used for DNA- mediated immunization due to the availability in commercial. Consequently, influence of the vector constituents, such as promoter, enhancer and transcription termination signal etc. on vaccination efficiency is not studied extensively. As an initial attempt to develop an efficient vector system for DNA-rnediated immunization, influence of promoter for antigen gene expression on vaccination efficiency has been analyzed. Initially, plasmids with either B-actin or muscle creatine kinase (MCK) promoter were constructed from the plasmid with prototype CMV promoter. In addition, ovalbumin (OVA) antigen gene has been cloned into each vectors to generate the plasmid vectors with different promoters for induction of the anti-OVA immune responses. Antigen protein expression in antigen gene transfected mouse muscle myoblast cells showed that the level from MCK promoter containing plasmid was slightly higher than those from either CMV or B-actin promoter containing plasmids. Also, the same plasmid turned out to be slightly more efficient than other plasmids in antibody imrnune response induction in vivo, when they were applied both through intramuscularly and intradermally. These results suggest that the commonly used CMV promoter containing plasmid vector could be further modified to develop an efficient vector for DNA-mediated immunization.
Animals
;
Clone Cells
;
Creatine Kinase, MM Form
;
Gene Expression
;
Immunization
;
Mice
;
Myoblasts
;
Ovalbumin
;
Plasmids*
;
Vaccination*
2.Hybrid retroviral vector with MCK enhancers inserted in LTR for stable and specific expression of human factor IX in skeletal muscle.
Jian-min WANG ; Jun HOU ; Xin-fang QIU ; Kotoku KURACHI ; Jing-lun XUE
Chinese Medical Journal 2004;117(6):893-898
BACKGROUNDRetroviral vectors have been widely used to introduce foreign into various target cells in vitro, thus showing relatively high systemic delivery efficiency of various transgene products. The authors investigated the stability and efficiency of skeletal muscle-specific hybrid retroviral vectors in expression of human factor IX (FIX) in vitro and iv vivo.
METHODSFIX cDNA in LIXSN vector was replaced with a FIX minigene containing splicing donor and splicing acceptor sequence of first intron of human FIX gene. Two copies of muscle creatine kinase enhancer (MCK, Me2) were inserted in forward or reverse orientation at NheI site of 3' long terminal repeat (LTR), resulting in two hybrid vectors, which were designated as LMe2IXm2SN(F) and LMe2IXm2SN(R), respectively. The vectors were tested in vitro and in vivo for stability and muscle-specificity of factor IX expression with SCID mice.
RESULTSMuscle cells carrying vector with Me2 expressed significantly higher levels of FIX (up to 1800 ng/106.24 h) than those without Me2, thus suggesting that Me2 could specifically increase expression level of FIX in muscle cells. Myoblasts transduced with LMe2IXm2SN(R) produced much less FIX in vivo in SCID mice than LMe2IXm2SN(F). One or two copies of Me2 sequence were deleted in myoblasts transduced with LMe2IXm2SN(R) without changing the orientation of Me2.
CONCLUSIONSLTR inserted with MCK enhancers can specifically increase human FIX expression in skeletal muscle cells in vitro and in vivo, and MCK enhancer should be positioned in the same orientation as that of LTR promoter.
Animals ; Creatine Kinase ; genetics ; Creatine Kinase, MM Form ; Enhancer Elements, Genetic ; Factor IX ; analysis ; genetics ; Gene Expression ; physiology ; Genetic Techniques ; Genetic Vectors ; Hybridization, Genetic ; Isoenzymes ; genetics ; Mice ; Mice, SCID ; Retroviridae ; genetics ; Terminal Repeat Sequences
3.Association of CKMM gene A/G polymorphism and athletic performance of uyghurnationality.
En-peng HE ; Yan-hong LI ; Jian-dong QIAN ; Hua-wei YAN
Chinese Journal of Applied Physiology 2016;32(1):82-86
OBJECTIVEDiscusses the distributive characters of the Creatine Kinase MM (CKMM) gene A/G Polymorphism in XinjiangUyghur, One hundred and fourtheen athletes and 441 general population of Uyghur were involved in the study.
METHODSPolymerase chain reaction-restriction fragment length polymorphism was used.
RESULTS(1) The CKMM gene A/G frequency in Uyghur general population was(AA, AG and GG) 0.497, 0.392 and 0.111, the result test by Hardy-Weinberg (H-W) equilibrium and x² = 2.72, P = 0.1, df = 2, indicated that the control group had representative. (2) AA, AG and GG genotype frequency of power-oriented athlete respectively was 0.442,0.302 and 0.256, frequency of GG genotype and G allele was higher than the control group, there were significant differences compared to thecontrol( P < 0.05, df = 2); (3) A/G genotype frequency of Endurance-oriented athletere spectively was 0.571, 0.400 and 0.029, there were nosignificant differences compared to the controls ( P > 0. 05, df = 2). (4) A/G genotype frequency of Uyghur soccer athletes respectively was0.472, 0.361 and 0.167, G allele was higher than the Endurance-oriented athlete and lower than the power-oriented athletes. and no significant differences compared to the controls( P > 0.05, df = 2).
CONCLUSIONThe results indicate that the CKMM gene GG genotype and G alleleare represented in power-oriented athletes, but don't find A/G polymorphism correlation with endurance and the football sport performance.
Alleles ; Asian Continental Ancestry Group ; genetics ; Athletes ; Athletic Performance ; China ; Creatine Kinase, MM Form ; genetics ; Gene Frequency ; Genotype ; Humans ; Physical Endurance ; genetics ; Polymorphism, Genetic ; Polymorphism, Restriction Fragment Length
4.Sumoylation of histone deacetylase 1 regulates MyoD signaling during myogenesis
Hosouk JOUNG ; Sehee KWON ; Kyoung Hoon KIM ; Yun Gyeong LEE ; Sera SHIN ; Duk Hwa KWON ; Yeong Un LEE ; Taewon KOOK ; Nakwon CHOE ; Jeong Chul KIM ; Young Kook KIM ; Gwang Hyeon EOM ; Hyun KOOK
Experimental & Molecular Medicine 2018;50(1):e427-
Sumoylation, the conjugation of a small ubiquitin-like modifier (SUMO) protein to a target, has diverse cellular effects. However, the functional roles of the SUMO modification during myogenesis have not been fully elucidated. Here, we report that basal sumoylation of histone deacetylase 1 (HDAC1) enhances the deacetylation of MyoD in undifferentiated myoblasts, whereas further sumoylation of HDAC1 contributes to switching its binding partners from MyoD to Rb to induce myocyte differentiation. Differentiation in C2C12 skeletal myoblasts induced new immunoblot bands above HDAC1 that were gradually enhanced during differentiation. Using SUMO inhibitors and sumoylation assays, we showed that the upper band was caused by sumoylation of HDAC1 during differentiation. Basal deacetylase activity was not altered in the SUMO modification-resistant mutant HDAC1 K444/476R (HDAC1 2R). Either differentiation or transfection of SUMO1 increased HDAC1 activity that was attenuated in HDAC1 2R. Furthermore, HDAC1 2R failed to deacetylate MyoD. Binding of HDAC1 to MyoD was attenuated by K444/476R. Binding of HDAC1 to MyoD was gradually reduced after 2 days of differentiation. Transfection of SUMO1 induced dissociation of HDAC1 from MyoD but potentiated its binding to Rb. SUMO1 transfection further attenuated HDAC1-induced inhibition of muscle creatine kinase luciferase activity that was reversed in HDAC1 2R. HDAC1 2R failed to inhibit myogenesis and muscle gene expression. In conclusion, HDAC1 sumoylation plays a dual role in MyoD signaling: enhancement of HDAC1 deacetylation of MyoD in the basally sumoylated state of undifferentiated myoblasts and dissociation of HDAC1 from MyoD during myogenesis.
Creatine Kinase, MM Form
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Gene Expression
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Histone Deacetylase 1
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Histone Deacetylases
;
Histones
;
Luciferases
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Muscle Cells
;
Muscle Development
;
Myoblasts
;
Myoblasts, Skeletal
;
Sumoylation
;
Transfection
5.The preliminary study on change of serum enzyme level in crush syndrome rat.
Shui Ping LIU ; Yu Chuan CHEN ; Wei GUO ; Jian Ding CHENG
Journal of Forensic Medicine 2001;17(4):205-206
OBJECTIVE:
To study the changes of serum creatinine kinase(CK) and its cardiac-specific isoenzyme compound(CK-MB) levels in crush injury rats.
METHODS:
Crush injury was produced in SD rats, the serum levels of CK and CK-MB were studied by automated biochemical analyzer.
RESULTS:
The levels of plasma CK and CK-MB were much higher in crush injury rats than those of the control group.
CONCLUSIONS
Cardiomyocyte injury may be induced in the early stage of crush injury rats.
Animals
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Creatine Kinase/blood*
;
Creatine Kinase, MB Form
;
Crush Syndrome/enzymology*
;
Female
;
Isoenzymes/blood*
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Male
;
Rats
;
Rats, Sprague-Dawley
6.Advances in biomarkers of myocardial injury in sepsis.
Junhai ZHEN ; Li LI ; Jing YAN
Chinese Critical Care Medicine 2018;30(7):699-702
Sepsis is a common disease in critical patients, which may lead to myocardial damage, thereby aggravating the severity of the patients' condition, and causing adverse prognosis. How to detect sepsis with myocardial injury as early as possible, and use corresponding treatment measures on time are essential. Cardiac troponin I (cTnI), brain natriuretic peptide (BNP), myoglobin (Mb), MB isoenzyme of creatine kinase (CK-MB) and other traditional cardiac markers are easily affected by the complications of other critical diseases, thus the diagnostic value of those markers for myocardial injury of sepsis is reduced. In recent years, there have been some studies on heart-type fatty acid binding protein (H-FABP), microRNA (miRNA), soluble triggering receptor expressed on myeloid cell-1 (sTREM-1), high mobility group protein B1 (HMGB1), neutrophil gelatinase-associated lipocalin (NGAL), histone and other new biomarkers of myocardial injury in septic patients. This article reviewed the value of these unconventional cardiac markers in the diagnosis of sepsis-induced myocardial injury, with the hope to provide some help for clinic.
Biomarkers
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Creatine Kinase, MB Form
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Humans
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Myocardial Infarction
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Myoglobin
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Natriuretic Peptide, Brain
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Sepsis
;
Troponin I
9.Ischemic preconditioning in immature hearts: mechanism and compatibility with cardioplegia.
Bin ZHU ; Su MIN ; Cun LONG ; Tiehu YE
Chinese Medical Journal 2003;116(2):253-257
OBJECTIVETo investigate (1) whether ischemic preconditioning (IPC) could protect immature rabbit hearts against ischemia-reperfusion injury and (2) the role of K(ATP) channel in the mechanism of myocardial protection. Since cardioplegia is a traditional and effective cardioprotective measure in clinic, our study is also designed to probe the compatibility between IPC and cardioplegia.
METHODSNew Zealand rabbits aged 14 - 21 days weighing 220 - 280 g were used. The animals were anesthetized and heparinized. The chest was opened and the heart was quickly removed for connection of the aorta via Langendorff's method within 30 s after excision. All hearts were perfused with Krebs-Henseleit buffer balanced with gas mixture (O(2):CO(2) = 95%:5%) at 60 cm H(2)O (perfusion pressure). IPC consisted of 5 min global ischemia plus 10 min reperfusion. Glibenclamide was used as the K(ATP) channel blocker at a concentration of 10 micro mol/L before IPC. Cardiac arrest was induced with 4 degrees C St. Thomas cardioplegic solution, at which point the heart was made globally ischemic by withholding perfusion for 45 min followed by 40 min reperfusion. Thirty immature rabbit hearts were randomly divided into four groups: CON (n = 9) was subjected to ischemia-reperfusion only; IPC (n = 9) underwent IPC and ischemia-reperfusion; Gli (n = 6) was given glibenclamide and ischemia-reperfusion; and Gli + IPC (n = 6) underwent glibenclamide, IPC and ischemia-reperfusion. Coronary flow (CF), HR, left ventricle developed pressure (LVDP), and +/- dp/dt(max) were monitored at equilibration (baseline value) and 5, 10, 20, 30 and 40 min after reperfusion. The values resulting from reperfusion were expressed as a percentage of their baseline values. Arrhythmia quantification, myocardial enzyme in the coronary effluent and myocardial energy metabolism were also determined.
RESULTSThe recovery of CF, HR, LVDP and +/- dp/dt(max) in preconditioned hearts was best among the four groups. The incidence of arrhythmia was low and less CK-MB leaked out in the IPC group. Myocardial ATP content was better preserved by IPC. Pretreatment with glibenclamide completely abolished the myocardial protection provided by IPC, but did not affect ischemia-reperfusion injury.
CONCLUSIONSWhile applying cardioplegia, IPC provides significant cardioprotective effects. Activation of K(ATP) channels is involved in the mechanism of IPC-produced cardioprotection.
Adenosine Triphosphate ; analysis ; Animals ; Creatine Kinase ; secretion ; Creatine Kinase, MB Form ; Heart Arrest, Induced ; Hemodynamics ; Ischemic Preconditioning, Myocardial ; Isoenzymes ; secretion ; Potassium Channels ; physiology ; Rabbits
10.Characteristics of myocardial enzymes of critical illness children with non-cardiac diseases.
Jian-Ling HUANG ; Hai-Yan YING ; Qing YANG
Chinese Journal of Contemporary Pediatrics 2006;8(5):431-432
Adolescent
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Alanine Transaminase
;
blood
;
Child
;
Child, Preschool
;
Creatine Kinase
;
blood
;
Creatine Kinase, MB Form
;
blood
;
Critical Illness
;
Female
;
Humans
;
Infant
;
L-Lactate Dehydrogenase
;
blood
;
Male
;
Myocardium
;
enzymology