The Distinguishing Factor in Soccer Players is Aerobic Performance, not the ACTN3 Gene / El Factor Distintivo en los Jugadores de Fútbol es el Rendimiento Aeróbico y no el gen ACTN3

SUMMARY: The purpose of this study was to reveal the differences between ACTN3 genotype (RR, RX, XX) and aerobic performance [Yo-Yo IRT1 (m), VO2 max (ml/kg/min)] in professional and regional amateur league soccer players and to reveal which of these parameters was a distinctive factor in these athletes.71 professional soccer players (age: 23.66 ± 4.11 years; body height: 1.79 ± 6.99 m; body weight: 76.02 ± 6.76 kg; body fat: 11.59±3.11 %) and 62 regional amateur soccer players (age: 23.63 ±3.77 years; body height: 1.81 ± 5.77 m; body weight: 76.36 ± 7.53 kg; body fat: 15.60±4.65 %) volunteered for the study. After DNA extraction from buccal epithelial cells via a commercial kit was performed for the genetic background of the athletes, Real-Time PCR was carried out for genotyping. Furthermore, Yo-Yo IRT1 test was performed to determine the aerobic performance of the soccer players. SPSS 23 (SPSS Inc., Chicago, IL, USA) package program was used for the statistical analysis of the data obtained in the tests. Shapiro-Wilk test for normality and Levene's test for homogeneity of variance were performed. Chi-Square, Independent Sample T Test and One Way ANOVA test were used in the analysis of the parameters. Statistical significance was set as p0.05); however, there was a statistical significance in favor of professional soccer players in terms of aerobic parameters (p<0.05). Consequently, it can be said that aerobic performance is the distinguishing factor, not the ACTN3 gene, in soccer players.

El objetivo de este estudio fue revelar las diferencias entre el genotipo ACTN3 (RR, RX, XX) y el rendimiento aeróbico [Yo-Yo IRT1 (m), VO2 max (ml/kg/min)] en jugadores de fútbol de ligas profesionales y amateurs regionales y determinar cuál de estos parámetros es un factor distintivo en estos deportistas. 71 futbolistas profesionales (edad: 23,66 ±4,11 años; altura corporal: 1,79 ± 6,99 m; peso corporal: 76,02 ± 6,76 kg; grasa corporal: 11,59±3,11 %) y 62 jugadores de fútbol amateur regionales (edad: 23,63 ± 3,77 años; altura corporal: 1,81 ± 5,77 m; peso corporal: 76,36 ± 7,53 kg; grasa corporal: 15,60 ± 4,65 %) se ofrecieron como voluntarios para el estudio. Después de realizar la extracción de ADN de las células epiteliales orales mediante un kit comercial para obtener los antecedentes genéticos de los atletas, se llevó a cabo una PCR en tiempo real para el genotipado. Además, se realizó la prueba Yo-Yo IRT1 para determinar el rendimiento aeróbico de los futbolistas. Para el análisis estadístico de los datos obtenidos en las pruebas se utilizó el programa SPSS 23 (SPSS Inc., Chicago, IL, EE. UU.). Se realizó la prueba de normalidad de Shapiro- Wilk y la prueba de homogeneidad de la varianza de Levene. En el análisis de los parámetros se utilizaron Chi-cuadrado, prueba T para muestra independiente y prueba ANOVA unidireccional. La significancia estadística se estableció en p0,05); sin embargo, hubo significación estadística a favor de los futbolistas profesionales en cuanto a los parámetros aeróbicos (p<0,05). En consecuencia, se puede decir que el rendimiento aeróbico es el factor distintivo, no el gen ACTN3, en los jugadores de fútbol.

Advance in research on genetic polymorphisms associated with athletic ability / 中华医学遗传学杂志

Genetic factors play a key role in human athletic ability, and endurance quality and explosive power quality are the important components of athletic ability. In this review, we aimed to reveal the biological genetic mechanism of human athletic ability at the molecular level through summarizing the relationship between genetic variants and human athletic ability, including endurance quality related genetic markers angiotensin converting enzyme (ACE) gene, creatine kinase MM (CKMM) gene and explosive power quality related genetic markers alpha actinin 3 (ACTN3) gene, angiotensinogen (AGT) gene and interleukin6 (IL6) gene. Meanwhile, we also summarized the distribution of allele frequencies among various populations.

The Role of Zyxin in Regulating Platelet Cytoskeleton Distribution / 中国实验血液学杂志

OBJECTIVE@#To investigate the regulatory effect of zyxin on the distribution of platelet cytoskeleton.@*METHODS@#Platelets were isolated from zyxin-knockout (Zyx@*RESULTS@#After zyxin gene was knockout, the expressions of cytoskeleton proteins β-actin, α-actinin, filamin A, and myosin Ⅱ A in resting and Jas-induced platelets were significantly increased. In the platelet spreading on fibrinogen surface, F-actin was increased in Zyx@*CONCLUSION@#Zyxin significantly regulates the distribution of platelet cytoskeleton, which plays an important role in maintaining platelet cytoskeleton homeostasis.

Frecuencia del polimorfismo genético ACTN3 R577X y ECA I/D en atletas ciegos de fútbol 5 / Frequency of genetic polymorphism ACTN3 R577X and ACE I/D in blind athletes of 5-a-side football

RESUMEN: El objetivo de este estudio fue describir la frecuencia genotípica y alélica del ACTN3 R577X y ECA I/D en atletas ciegos de fútbol 5. Se incluyó una metodología descriptiva con una muestra de 63 deportistas ciegos (28,0±5,8 años), todos varones, de equipos de fútbol 5 de alto rendimiento. El polimorfismo se determinó mediante la reacción en cadena de la polimerasa en tiempo real (RT-PCR). La estadística fue descriptiva realizada a partir de las medidas de frecuencia de genotipos y alelos. La frecuencia genotípica de la ACTN3 en los deportistas presentó la siguiente distribución: el 28,6 % con genotipo RR, el 54 % con RX y el 17,4 % con XX y frecuencia alélica del 55,6 % para el alelo R y del 44,4 % para el alelo X. En cuanto a la ECA I/D, la frecuencia genotípica fue del 63,5 % para el genotipo ID, del 22,2 % para el DD y del 14,3 % para el II. La frecuencia alélica presentó prevalencia del alelo D con el 53,9 %. El estudio constató una predominancia de los genotipos y alelos representativos de las modalidades de fuerza y velocidad para ACTN3 R577X y ECA I/D de atletas de fútbol 5.

SUMMARY: The aim of this study was to describe the genotypic and allele frequency of ACTN3 R577X and ACE I/D in blind athletes of 5-a-side football performance. A descriptive methodology was included with a sample of 63 blind male athletes (28.0 ±5.8 years) of football teams with a 5-a-side performance rating. The polymorphism was determined by means by of real-time Polymerase Chain Reaction (rt-PCR). Statistics were descriptive based on the measures of frequency of genotypes and alleles. The genotypic frequency of ACTN3 by the athletes presented the following distribution: 28.6 % with RR genotype, 54 % with RX and 17.4 % XX and allele frequency of 55.6 % for the R allele and 44.4 % for the X allele. As for ACE I/D, the genotype frequency was 63.5 % for genotype ID, 22.2 % for DD and 14.3 % for II. The allele frequency showed a predominance of the D allele with 53.9 %. The study found for ACTN3 R577X and ACE I/ D of blind athletes of 5-a-side football, a predominance of genotypes and alleles representative of strength and speed modalities.

Comparison of the Cardiomyogenic Potency of Human Amniotic Fluid and Bone Marrow Mesenchymal Stem Cells

BACKGROUND AND OBJECTIVES: Most studies in cardiac regeneration have explored bone marrow mesenchymal stem cells (BM-MSC) with variable therapeutic effects. Amniotic fluid MSC (AF-MSC) having extended self-renewal and multi-potent properties may be superior to bone marrow MSC (BM-MSC). However, a comparison of their cardiomyogenic potency has not been studied yet.METHODS: The 5-azacytidine (5-aza) treated AF-MSC and BM-MSC were evaluated for the expression of GATA-4, Nkx2.5 and ISL-1 transcripts and proteins by quantitative RT-PCR and Western blotting, respectively as well as for the expression of cardiomyogenic differentiation markers cardiac troponin-T (cTNT), beta myosin heavy chain (βMHC) and alpha sarcomeric actinin (ASA) by immunocytochemistry.RESULTS: The AF-MSC as compared to BM-MSC had significantly higher expression of GATA-4 (183.06±29.85 vs. 9.80±0.05; p<0.01), Nkx2.5 (8.3±1.4 vs. 1.82±0.32; p<0.05), and ISL-1 (39.59±4.05 vs. 4.36±0.39; p<0.01) genes as well as GATA-4 (2.01±0.5 vs. 0.6±0.1; p<0.05), NKx2.5 (1.9±0.14 vs. 0.8±0.2; p<0.01) and ISL-1 (1.7±0.3 vs. 0.9±0.1; p<0.05) proteins. The AF-MSC also had significantly elevated expression of cTNT (5.0×10⁴±0.6×10⁴ vs. 3.5 ×10⁴±0.8×10⁴; p<0.01), β-MHC (15.7×10⁴±0.9×10⁴ vs. 8.2×10⁴±0.6×10⁴; p<0.01) and ASA (18.6×10⁴±4.9×10⁴ vs. 13.1×10⁴±3.0×10⁴; p<0.05) than BM-MSC.CONCLUSIONS: Our data suggest that AF-MSC have greater cardiomyogenic potency than BM-MSC, and thus may be a better source of MSC for therapeutic applications in cardiac regenerative medicine.

Polimorfismo del gen ACTN3 y ECA en seleccionados de gimnasia de Brasil y Japón / Gene polymorphism ACTN3 and ACE in selected gymnasts athletes in Brazil and Japan

En el rendimiento deportivo, el estudio de la genética ha cobrado un papel fundamental al momento de estudiar el rendimiento deportivo. El objetivo del estudio fue analizar la frecuencia de genotipo y alelo de a - actinina 3 (ACTN3) R577X y enzima convertidora de angiotensina (ECA) I/D polimorfismo en un grupo de gimnastas de Brasil y Japón. Se considero una muestra de 73 gimnastas (14 de Japón y 59 de Brasil), todos los sujetos firmaron de consentimiento informado. Para la obtención de ACTN3 y ECA se recogió una muestra la saliva y analizaron los genotipos mediante el empleo de cadena de la polimerasa en tiempo real a partir del iQ5 Thermal Cycler, BioRad. Los resultados muestran una prevalencia del genotipo RX ACTN3, prevalencia del alelo R, prevalencia del genotipo DI ECA y prevalencia de alelo D en el total del grupo. Se concluye que el predominio RX del ACTN3, prevalencia del alelo R y predominio del genotipo DI en ECA pueden ofrecer una ventaja genética en cuanto a niveles de fuerza y potencia muscular, posiblemente facilitando la práctica y el éxito competitivo en gimnastas.

In the field of competitive sport, genetics has gained a fundamental role in the study of sport performance. The aim of this study was to analyze the prevalence of polymorphisms R577X and insertion/deletion (I/D), occurring in a-actinin-3 (ACTN3) and angiotensin converting enzyme (ACE) genes, respectively, in Brazilian and Japanese gymnasts. A suitable non-probabilistic sample of 73 gymnasts (14 from Japan) was recruited and signed an informed consent. To measure ACTN3 and ECA saliva samples were obtained by means of real time polymerase chain reaction (iQ5 Thermal Cycler, BioRad). A high prevalence of RX ACTN3 genotype, R allele, ACE I/D genotype, and D allele were observed in Brazilian and Japanese gymnasts. In conclusion a high prevalence of RX ACTN3 genotype, R allele and ACE I/D genotype would allow a genetic advantage regarding muscle strength and power, possibly facilitating competitive success in gymnastics.

Expression patterns of sarcomeric α-actin, α-actinin and UCP2 in the myocardium of Kunming mice after exposure to c-terminal polypeptide of cardiotrophin-1 / 华中科技大学学报（医学）（英德文版）

Cardiotrophin-1 (CT-1) activates a distinct form of cardiac muscle cell hypertrophy in which the sarcomeric units are assembled in series. The aim of the study was to determine the expression pattern of sarcomeric contractile protein α-actin, specialized cytoskeletal protein α-actinin and mitochondrial uncoupling protein-2 (UCP2) in myocardial remodeling induced by chronic exposure to CT-1. Kunming mice were intraperitoneally injected with carboxy-terminal polypeptide (CP) of CT-1 (CT-1-CP, 500 μg·kg(-1)· day(-1)) for 1, 2, 3 and 4 week (s), respectively (4 groups obtained according to the injection time, n=10 each, with 5 males and 5 females in each group). Those injected with physiological saline for 4 weeks served as controls (n=10, with 5 males and 5 females). The heart tissues of mice were harvested at 1, 2, 3 or 4 week (s). Immunohistochemistry (IHC) and Western blotting (WB) were used to detect the distribution and expression of sarcomeric α-actin, α-actinin and mitochondrial UCP2 in myocardial tissues. IHC showed that α-actin was mainly distributed around the nuclei of cardiomyocytes, α-actinin concentrated around the striae and UCP2 scattered rather evenly in the plasma. The expression of α-actin was slightly greater than that of α-actinin and UCP2 in the control group (IHC: χ(2)=6.125; WB: F=0.249, P>0.05) and it gradually decreased after exposure to CT-1-CP. There was no significant difference in the expression of α-actin between the control group and the CT-1-CP-treated groups (χ (2)=7.386, P>0.05). But Western blotting revealed significant difference in the expression of α-actin between the control group and the 4-week CT-1-CP-treated group (F=2.912; q=4.203, P<0.05). Moreover, it was found that the expression of α-actinin increased stepwise with the exposure time in CT-1-CP-treated groups and differed significantly between CT-1-CP-treated groups and the control group (ICH: χ (2)=21.977; WB: F=50.388; P<0.01). The expression of UCP2 was initially increased (WB: control group vs. 1- or 2-week group, q values: 5.603 and 9.995, respectively, P<0.01) and then decreased (WB: control group vs. 3-week group, q=4.742, P<0.01; control group vs. 4-week group, q=0.558, P>0.05). It was suggested that long-term exposure to CT-1-CP could lead to the alteration in the expression of sarcomeric α-actin, α-actinin and mitochondrial UCP2. The different expressions of sarcomeric structure proteins and mitochondrial UCP2 may be involved in myocardial remodeling.

Polimorfismo do gene da eca e da a-actinina 3 na escoliose idiopática do adolescente / Polymorphism of the ace gene and the a-actinin-3 gene in adolescent idiopathic scoliosis

Objetivo: O polimorfismo da enzima conversora de angiotensina (ECA) I/D e da α-actinina 3 (ACTN3) R577X está relacionado a variações na função do musculoesquelético. O objetivo deste trabalho foi avaliar a distribuição destes polimorfismos em uma família com múltiplos membros com escoliose idiopática do adolescente. Métodos: avaliação de 25 indivíduos de uma família, com múltiplos membros com escoliose idiopática, por meio da coleta de 10 ml de sangue para extração de DNA. A genotipagem do polimorfismo I/D do gene da ECA e R577X do gene da ACTN3 foi realizada utilizando sistema de 2 iniciadores (‘‘primers’’) específicos, para classificar os indivíduos em homozigotos ou heterozigotos. Resultados: Em relação ao polimorfismo da ECA encontrou-se 19 indivíduos DD (76%) e 6 ID (24%). A prevalência do alelo D foi de 88% e do alelo I foi de 12%. Quanto ao polimorfismo da ACTN3 observou-se seis indivíduos RR (24%), 11 RX (44%) e 8 XX (32%). A prevalência do alelo R foi 23 (46%) e do alelo X foi 27 (54%). Conclusão: observou-se diferença entre a distribuição do polimorfismo da ECA e da ACTN3 na família estudada. Ao avaliar o polimorfismo da ECA notou-se maior prevalência do alelo D em relação ao alelo I. Nível de Evidência III, Estudo Clinico, Transversal.

Objective: The I/D polymorphism of angiotensin-converting enzyme (ACE) and R577X of the α-actinin-3 (ACTN3) is related to changes in skeletal muscle function. The aim of this study was to evaluate the distribution of these polymorphisms in a family with multiple members with adolescent idiopathic scoliosis (AIS). Methods: evaluated 25 subjects from a family with multiple members with AIS, by collecting 10mL of blood for DNA isolation. The genotyping of the I/D polymorphism of the ACE gene and the R577X of the ACTN3 gene was performed using two specific primers to classify individuals as homozygous or heterozygous. Results: regarding the ACE polymorphism it was found that 19 (76%) subjects were DD and 6 (24%) ID. The prevalence of the D allele was 88% and the I allele was 12%. Regarding the ACTN3 polymorphism there were 6 subjects RR (24%), 11 RX (44%) and 8 XX (32%). The prevalence of the R allele was 23 (46%) and the X allele was 27 (54%). Conclusion: there was a difference between the distribution of the polymorphism of ACE and ACTN3 in the family studied. When assessing the ACE polymorphism a higher prevalence of the D allele was observed as compared with the I allele. Level of Evidence III, Cross-sectional, Clinical Trial.

Expression of Junctophilin 1 during cardiogenesis of mouse embryonic stem cells and rat embryos / 浙江大学学报·医学版

<p><b>OBJECTIVE</b>To investigate the expression of Junctophilin 1 (JP1) in cardiogenesis of mammalian.</p><p><b>METHODS</b>Cardiac differentiation of embryonic stem cells (ESCs) was generated by hanging drop method. Fetal heart was obtained from the rats aged d 14-20 of gestation. The expression of JP1 and JP2 during cardiogenesis of ESCs and rat embryos was analyzed by RT-PCR or Western blotting. Immunofluorescence staining was employed to reveal the distribution of JP1 and JP2 in embryoid body (EB), probing for merging of JP1 and JP2 and cardiac sarcomeric α-Actinin or Troponin-T. Percentage of JP1 and JP2-positive staining cells was analyzed quantitatively by FCS on d17.</p><p><b>RESULTS</b>JP1 mRNA was up-regulated at the early stage (d 5-11) and then decreased. The expression of JP1 protein was up-regulated at the early stage (d 7-9), then decreased gradually and disappeared after d 15. While JP2 gene and protein expression increased in a time-dependent manner during cardiogenesis of rat embryos. The results of immunofluorescence staining showed that there was a parallel co-localization of JP2 with Troponin-T or α-Actinin on d17, while JP1 failed to express in the sarcomeric positive area at the same time point. Furthermore, FCS analysis showed that about 16.59% of cells were JP2-positive, while no cells were stained positively for JP1 in d17 EBs.</p><p><b>CONCLUSION</b>JP1 gene is expressed during the whole process of cardiogenesis, while JP1 protein only appears on the early stage. The expression of JP1 in cardiogenesis of ESCs is consistent with that of rat embryos.</p>

Genetic doping and health damages

Use of genetic doping or gene transfer technology will be the newest and the lethal method of doping in future and have some unpleasant consequences for sports, athletes, and outcomes of competitions. The World Anti-Doping Agency [WADA] defines genetic doping as "the non-therapeutic use of genes, genetic elements, and/or cells that have the capacity to enhance athletic performance". The purpose of this review is to consider genetic doping, health damages and risks of new genes if delivered in athletes. This review, which is carried out by reviewing relevant publications, is primarily based on the journals available in GOOGLE, ELSEVIER, PUBMED in fields of genetic technology, and health using a combination of keywords [e.g., genetic doping, genes, exercise, performance, athletes] until July 2010. There are several genes related to sport performance and if they are used, they will have health risks and sever damages such as cancer, autoimmunization, and heart attack

Relationship between α-actinin and cardiac function in rats with myocardial ischemia-reperfusion / 南方医科大学学报

<p><b>OBJECTIVE</b>To explore the relationship between α-actinin content and cardiac function in rats during myocardial ischemia-reperfusion.</p><p><b>METHODS</b>Thirty-two rats were randomized equally into sham-operated group, 30 min ischemia group, 1 h ischemia group, and 1 h ischemia with 2 h reperfusion group. Acute myocardial ischemia was induced in the 3 ischemia groups by ligation of the left anterior descending coronary artery, and the cardiac functions were evaluated. The myocardial contents of α-actinin was measured by immunohistochemistry, and phospholipase C (PLC) and phosphatidylinositol-3-kinase (PI3K) contents were determined by ELISA after the operations.</p><p><b>RESULTS</b>The left ventricular systolic pressure (LVSP), +dp/dt max, and -dp/dt max tended to decrease during myocardial ischemia, and increased after reperfusion, and the left ventricular end-diastolic pressure (LVEDP) showed reverse changes. The levels of α-actinin decreased with prolonged ischemia, showing a significant difference in 1 h ischemia group from those in the other 3 groups. PI3K and PLC contents were significantly increased with prolonged myocardial ischemia. Stimulation by LY-294002 and U-73122 caused enhanced contraction of single cardiomyocytes, and also increased the fluorescence intensity of α-actinin in the cardiomyocytes compared with that in 1 h ischemia group.</p><p><b>CONCLUSIONS</b>The cardiac dysfunction during acute ischemia-reperfusion in rats may be related with the changes of myocardial α-actinin content, which are probably a result of increased PI3K and PLC contents in the ischemic myocardium.</p>

The effects of pulsed electromagnetic fields on the induction of rat bone marrow mesenchymal stem cells to differentiate into cardiomyocytes-like cells in vitro / 生物医学工程学杂志

The aim of this study is to investigate the effects of pulsed electromagnetic fields (PEMFs) on the induction of rat bone marrow mesenchymal stem cells (rBMSCs) to differentiate into cardiomyocytes-like cells in vitro. rBMSCs were randomly divided into PEMFs groups, 5-Azacytidine (5-Aza) groups and control groups. PEMFs groups were exposed to 50 Hz, 1 mT PEMFs for 30 min every day, lasting for 10 d, 15 d and 20 d, respectively. 5-Aza groups were induced by 10 micromol/L 5-Aza for 1 day, then the medium was changed to complete medium without 5-Aza. And control groups were only cultured with complete medium, rBMSCs growth status and morphological features were observed by inverted phase microscope every day. The mRNA expressions of cardiac troponin T (TNNT2) and alpha-actinin (ACTN2) were determined by Real-Time PCR. The results showed that rBMSCs were spindle, polygon or fusiform in control groups. The cells gradually got longer and grew close together after being stimulated by PEMFs and 5-Aza, and with the extension of induction time, the tendency became obvious. At 20th day after PEMFs or 5-Aza treatment, rBMSCs gathered like a long chain, got much longer obviously at the high magnification, and some of them even fused with their neighbors. Compared with control groups, the levels of TNNT2 mRNA expression in 5-Aza groups were 19.40 fold (P < 0.01), 21.02 fold (P < 0.01) and 2.38 fold at 10 d, 15 d, 20 d and the levels of ACTN2 mRNA expression in 5-Aza groups were 6.64 fold (P < 0.01), 6.67 fold (P < 0.01) and 0.76 fold at 10 d, 15 d, 20 d. However, the levels of TNNT2 mRNA expression in PEMFs groups were 15.78 fold (P < 0.01), 6.73 fold (P < 0.05) and 2.73 fold (P < 0.01) of control groups at 10 d, 15 d, 20 d and the levels of ACTN2 mRNA expression in PEMFs groups were 4.93 fold (P < 0.01), 1.89 fold and 0.64 fold, respectively. Compared with 5-Aza groups, the levels of TNNT2 mRNA expression in PEMFs groups were 0.81 fold, 0.32 fold (P < 0.01) and 1.15 fold at 10 d, 15 d, 20 d and the levels of ACTN2 mRNA expression in PEMFs groups were 0.74 fold, 0.28 fold (P < 0.01) and 0.83 fold at 10 d, 15 d, 20 d. PEMFs could contribute to the induction of rat marrow rBMSCs to differentiate into cardiomyocytes-like cells in vitro, and the best exposure time might be 10 days, but further investigation is still needed.

Effects of puromycin aminonucleoside on the cytoskeletal changes of glomerular epithelial cells / 소아과

PURPOSE:This study was designed to clarify the mechanism of proteinuria in nephrotic syndrome patients by using puromycin aminonucleoside (PAN) nephrosis model. METHODS:Following administration of various concentrations of PAN and antioxidants we observed the changes of podocyte cytoskeletons in cultured rat glomerular epithelial cells (GEpC) by method of scanning electron microscope, reactive oxyten species (ROS) analysis, permeability assay, confocal microscope, and Western blot assay. RESULTS:PAN not only induced the ultrastructural changes of GEpC, such as shortening and fusion of microvilli, but also separated the intercellular gaps and linear ZO-1. PAN induced oxidative stresses in time and dose dependent manners and increases of intercellular permeability in anti-oxidants inhibitable manners. High concentration of PAN induced not only actin polymerization and disorganization, but also the conglomerulation and internal dislocation of alpha-actinin protein. The intensities of fluorescences of ZO-1 protein were diminished and internalized by PAN in a dose-dependent manner, which were inhibited by anti anti-oxidants. CONCLUSION:PAN induced the changes of podocytes cytoskeleton and junctional barriers by way of increasing ROS in GEpC that resulted in increasing their permeability in a antioxidatn-inhibitable manner. Glomerular hyperpermeability induced by PAN mediateing through oxidative stresses is thought to take part in the mechanism of proteinuria in nephrotic syndrome. (Korean J Pediatr 2008;51:54-61)

MicroRNAs can be expressed in cardiomyocyte-like cells differentiated from human mesenchymal stem cells / 南方医科大学学报

<p><b>OBJECTIVE</b>To investigate the expression of representative heart-specific primary microRNAs (pri-miRNAs) in the cardiomyocyte-like cells differentiated from human mesenchymal stem cells (hMSCs).</p><p><b>METHODS</b>The phenotype of hMSCs isolated was identified by flow cytometry using monoclonal antibodies against FITC-conjugated CD29, CD34, and CD11b. The third-passage hMSCs were induced to differentiate into cardiomyocyte-like cells by 5-azacytidine and indirect coculture with neonatal rat myocytes, respectively. Immunocytochemical analysis was performed to detect the expression of the cardiac-specific proteins, namely cardiac troponin I (cTnI) and sarcomeric alpha-actinin, in the cardiomyocyte-like cells differentiated from hMSCs. RT-PCR and DNA sequencing were used to identify the expression of the 5 representative heart-specific pri-miRNAs.</p><p><b>RESULTS</b>High hMSC marker CD29 expression rate (98.87%) and low hematopoietic cell markers CD34 (5%) and CD11b (0.4%) expression rates were identified in the hMSCs isolated. cTnI and sarcomeric alpha-actinin expression occurred in the hMSCs following induction with the 2 differentiation-inducing methods. miRNA-143 and -181 expressions were induced in the hMSCs by 5-azacytidine and miRNA-143, -181, -206, and -208 expressions were induced by indirect coculture with neonatal rat myocytes, but pri-miRNA-1-2 expression failed to be induced by these two induction methods.</p><p><b>CONCLUSION</b>Expressions of the representative heart-specific pri-miRNAs in different patterns can be induced in cardiomyocyte-like cells differentiated from hMSCs by 5-azacytidine and indirect coculture with neonatal rat myocytes.</p>

Identification of Oocyte-Specific Diva-Associated Proteins using Mass Spectrometry / 대한불임학회지

OBJECTIVE: We previously described that Diva is highly expressed in matured metaphase II (MII) oocytes compared to immature germinal vesicle (GV) oocytes in mouse.1 We report here that the expression of Diva transcript as well as protein is oocyte-specific. To elucidate its physiological role in oocyte, the binding partner(s) of Diva has been identified by using immunoprecipitation (IP) followed by Mass Spectrometry. METHODS: NIH/3T3 cells were transiently transfected for 24 h with either empty vector for control or FLAG-tagged mouse Diva construct, and IP was performed with anti-FLAG antibody. The immuno-isolated complexes were resolved by SDS-PAGE on a 12% gel followed by Coomassie Blue staining. For in-gel digestion, 15 bands of interest were excised manually and digested with trypsin. All mass spectra were acquired at a positive reflector mode by a 4700 Proteomics Analyzer (Applied Biosystems, Framingham, MA). Proteins were identified by searching the NCBI nonredundant database using MASCOT Peptide Mass Fingerprint software (Matrixscience, London). RESULTS: Diva-associated complexes were formed in FLAG-tagged mouse Diva-overexpressed NIH/3T3 cells via IP using anti-FLAG-conjugated beads. Among the excised 15 bands, actin and actin-binding proteins such as tropomyosin, tropomodulin 3, and alpha-actinin were identified. Binding between Diva and actin or tropomyosin was confirmed by IP followed by Western blot analysis. Both bindings were also detected endogenously in mouse ovaries, indicating that Diva works with actin and tropomyosin. CONCLUSIONS: This is the first report that immuno-isolated Diva-associated complexes are related to actin filament of the cytoskeletal system. When we consider the association of Diva with actin and tropomyosin, oocyte-specific Diva may play a role in modulating the cytoskeletal system during oocyte maturation.

Suppressive effects of GTW treatment on mesangial lesions in experimental irreversible glomerulosclerosis / 中国中药杂志

<p><b>OBJECTIVE</b>To examine suppressive effects of multi-glycoside of Tripterygium wilfordii Hook. f. (GTW)on mesangial injury induced by two-injecti on of anti-Thy1. 1 monoclonal antibody(mAb) 1-22-3 in vitro.</p><p><b>METHOD</b>We established the irreversible model of glomerulosclerosis with anti-Thy1. 1 mAb 1-22-3. After 42 days of oral treatment with GTW (50 mg x kg(-1) BW)and vehicle (distilled water), to observe effects of GTW on proteinuria, renal function, mesangial morphological change, and mRNA expressions of collagen type I and TGF-beta by light microscope (LM), immunofluorescence (IF), and Reverse Transcription Polymerase Chain Reaction (RT-PCR).</p><p><b>RESULT</b>GTW ameliorated proteinuria (from day24 to day 42) and mesangial proliferation [total cell number, GTW group 65.67+/-3.43 vs. control group 87.02+/-2.41, P < 0.05; matrix expansion, GTW group 1.20+/-0.06 vs. control group 2.77+/-0.23, P < 0.05; alpha-smooth muscle actin(alpha-SMA) expression, GTW group 1.75+/-0.33 vs. control group 2.62+/-0.15, P < 0.05; collagen type I expression, GTW group 1.68+/-0.31 vs. control group 2.06+/-0.24, P < 0.05], moreover, significantly reduced the glomerular expression of mRNA for collagen type 1(53.5% to the control group, P < 0.05)and TGF-beta(14.7% to the control group, P < 0.05)on day 42day.</p><p><b>CONCLUSION</b>GTW can not only decrease proteinuria, but also ameliorate mesangial alterations probably by the reduction of cytokines. GTW may be a promising agent for the prevention of progressive and irreversible glomerulosclerosis.</p>

Cytoplasmic Domain of Intercellular Adhesion Molecule-1(ICAM-1)is Important for the ICAM-1 Mediated Membrane Projection of Endothelial Cells / 대한해부학회지

Intercellular adhesion molecule-1 (ICAM-1)has been shown to enhance leukocyte adhesion, thereby inducing migration through blood endothelial cells. However, the molecular event during the process of adhesion is largely unknown. To examine the role of ICAM-1 cytoplasmic domain in SDF-1 alpha-induced T lymphocyte migration and adhesion, mutant human ICAM-1 molecules were expressed in COS-7 cell line. COS-7 cells expressing ICAM-1_GFP mutant without alpha-actinin revealed no association with the actin cytoskeleton, while wild-type ICAM-showed clear association with the actin, as observed by confocal microscopy, suggesting that actinin binding motif in the cytoplasmic domain of ICAM-1 is important for the proper localization of ICAM-1 on the cell membrane. However, based on adhesion assay, we found that the cytoplasmic domain of ICAM-1 is not essential for the binding of lymphocytes which were activated by SDF-1alpha. On the other hand, ICAM-1-mediated receptor-ligand clustering event was significantly inhibited in the cells expressing ICAM-1 mutants without alpha-actinin or whole cytoplasmic domain. Taken together, these results suggest that ICAM-1 cytoplasmic domain is not essential for the adhesion but important for the ligand-receptor-mediated membrane projection of endothelial cells before trans-endothelial migration of lymphocytes.

Cellular Cardiomyoplasty Using Bone Marrow Derived Mesenchymal Stem Cells Transplantation in Post Myocardial Infarction Heart Failure

BACKGROUND AND OBJECTIVES: Cellular cardiomyoplasty (CCM) is considered to be a novel therapeutic approach for post-myocardial infarction (MI) heart failure. In this study, the functional effects of cultured mesenchymal stem cells (MSCs) transplantation and the associated histopathologic changes were evaluated in a rat model of MI. MATERIALS AND METHODS: Rats were subjected to 5 hours of coronary ligation followed by reperfusion, and 10 days after MI, animals were randomized into either the MSCs transplantation (MI-MSC, n=8) group or the control (n=8) group. Allogeneic MSCs (3x10(6) cells) or media were epicardially injected into the center and the border area of the infarct scar. RESULTS: Four weeks after the MSCs transplantation, the echocardiogram showed preserved anterior regional wall motion and increases in fractional shortening in the MI-MSC heart relative to the control heart. Left ventricular (LV) end diastolic pressure was smaller in the MI-MSC than in the control group. Implanted MSCs formed islands of cell clusters on the border of the infarct scar, and the cells were positively immunostained by sarcomeric alpha-actinin and cardiac troponin T. In addition, the number of microvessels on the border area of the infarct scar was greater in the MI-MSC than in the control group. CONCLUSION: Allogeneic MSCs transplanted into the MI scar formed clusters of cell grafts on the border of the infarct, expressed cardiac muscle proteins, increased microvessel formation, and improved regional and global LV function. Our data indicate that CCM using MSCs may have a significant role in the treatment of post-MI heart failure.

A novel mutation of NPHS2 identified in a Chinese family with steroid-resistant nephrotic syndrome / 中华儿科杂志

<p><b>OBJECTIVE</b>Autosomal recessive steroid-resistant nephrotic syndrome (SRNS) is a subgroup of familial nephrotic syndrome. A causative gene has been identified, that is NPHS2, in chromosome 1q25-31, which encodes podocin. This study aimed to detect NPHS2 mutation in a Chinese family with SRNS.</p><p><b>METHODS</b>Renal biopsy was performed on the proband and her sibling for routine histologic and immunohistochemical investigation and electron microscopic examination. The expressions of podocin, nephrin, alpha-actinin and WT1 in glomeruli of the proband were detected by indirect immunofluorescence. Peripheral blood samples were collected for genetic analysis from the proband and her parents, and 53 adults with normal urinalysis. Genomic DNA was isolated from peripheral blood leucocytes. Eight exons of NPHS2 were amplified by polymerase chain reaction. Mutational analysis was performed using denaturing high-performance liquid chromatography (DHPLC) and DNA fragments with aberrant elution profiles of both strands revealed by DHPLC were re-amplified and sequenced directly.</p><p><b>RESULTS</b>The histologic findings on kidney biopsies were focal segmental glomerulosclerosis. In controls, the distribution of staining with P35, rabbit against a human podocin recombinant protein (amino acids 135 - 383 = all the C-terminal part of the protein downstream the transmembrane domain), and P21, rabbit against a human podocin recombinant protein (amino acids 15 - 89 = all the N-terminal part of the protein upstream the transmembrane domain) showed a linear pattern along glomerular capillary walls on glomeruli, and the fluorescent intensity of the staining with P35 was intensely positive. The fluorescent intensity of the staining with P21 was positive. In the proband, the distribution of the staining with P35 showed uneven and nonlinear, and the fluorescent intensity of the staining with P35 was weakly positive. The staining with P21 was negative. The area, location, distribution and fluorescent intensity of the staining with nephrin, alpha-actinin and WT1 on glomeruli of the proband were the same as those in the controls. The DHPLC elution profiles of exon 4 of NPHS2 from the proband and her parent were aberrant. The chromatograms by sequencing detected in the exon 4 of NPHS2 showed a composite heterozygous mutation of both 467_468insT and 503G > A in the proband, a heterozygous mutation of 503G > A in her father, and a heterozygous mutation of 467_468insT in her mother, respectively.</p><p><b>CONCLUSION</b>The study demonstrated for the first time a novel mutation, 503G > A, of NPHS2 in Chinese kindred with autosomal recessive SRNS. A significantly decreased or negative expression was also revealed in glomeruli of the proband stained with two kinds of anti-podocin antibodies.</p>

Morphological Observations on the Changes of Membrane Permeability by Immobilization Stress in the Rat Atrial Myocyte / 대한해부학회지

This study was performed to investigate the effect of immobilization stress on the ultrastructural changes and membrane permeability in rat atrial myocyte using immunohistochemical and lanthanum tracer techniques. Male Sprague-Dawley rats, body weight 160~200 g, were used for all immobilization stress group. Rats were immobilized in small round plastic tube for 6, 12 or 24 hours, except for the control group. Alterations of myocardial myoglobin and alpha-actinin as well as membrane permeability after immobilization stress were examined by immunohistochemistry, and lanthanum permeability of the rat atrial myocyte were observed by electron microscopy. In the control group, there was no loss of myoglobin or alpha-actinin from the atrial myocytes. After 6 and 12 hours immobilization stress, the loss of myoglobin and alpha-actinin could be identified the atrial myocytes. In the 24 hour immobilization groups, the content of the myoglobin and alpha-actinin recovered partially. Lanthanum was deposited only in the intercellular space of the atrial myocardium in the control group. In the 6 hour immobilization group, the atrial myocytes showed severe ultrastructural changes during immobilization stress. Lanthanum deposited in the sarcoplasm, myofibrils, adjacent of mitochondria, and mitochondrial matrix. In the 12 or 24 hour immobilization groups, the morphological alteration of atrial myocytes appeared weekly. In the 12 hour group, lanthanum deposited in myofibrils, adjacent of mitochondria and in the mitochondrial matrix. In the 24 hour group, lanthanum deposited mainly in intercellular space of atrial myocardium, and rarely in the sarcoplasm of myocytes. These results suggest that the immobilization stress may induce the alteration of cardiac cell membrane permeability and the ultrastructures of atrial myocardium.