TAURINE INFLUX IN CULTURED RAT CARDIOMYOCYTES AND CHANGES AFTER CVB3 INFECTION

Monolayer culture of spontaneously contracting rat heart cells was obtained from newborn SD rats andseeded intO culture plates. 1OOTCID50 of coxsackie virus B3 (CVB3) (Nancy strain )was added as the infectedgroup. The dynamics of taurine transport across membrane of normal heart cells and changes after infectionwere examined by using radioactive isotope tracing techniques. The results demonstrated that: (1)two uptake systems of taurine with different affinities and capacities were present in the Plasma membrane of cul-tured rat cardiOmyocytes,the K. of high and low affinity were 9. 5 X 10-5mol/l, and l. 3 X 1 O 2 mol/l,,respectively; (2)taurine transport across membrane was sodium-dependent with a positive linear relationshipbetween amount of taurine influx and extracellular concentration of sOdium;in addition,it was alsO regulat-ed by osmOlarity; (3) fyalanine,a carboxylic analog of taurine,could inhibit the uptake of taurine in dosedependently; (4)taurine influx was decreased in cultured rat cardiomyocytes after infected with CVB3. (5)the lnhibition ratio of taurine transport was enhanced as the infected time prolonged. This experiment pro-vides a basis fOr further investigation of kinetic test of taurine transport and more study should be carriedout to learn the effect of taurine in viral myocarditis.

Effects of L-THP on Ca^(2+) Overload of Cultured Rat Cardiomyocytes during Hypoxia and Reoxygenation

The effects of L-tetrahydropalmatine (L-THP) on the cultured rat cardiomyocytes during hypoxia and reoxygenation and the mechanism of L-THP treating reperfusion-arrythmias were stud- ied. The concentration of intracellular free calcium ([Ca2+]i) of single cultured ventricular myocyte was determined by using EPC-9 light-electricity measurement system. It was found that L-THP (100μmol/L) could reduce the [Ca2+]i augmentation in single cultured ventricular myocyte during hypoxia and reoxygenation. Verapamil (10 μmol/L ) had the similar effect. It was concluded that L- THP could inhibit the Ca2+ overload of cultured rat cardiomyocytes during hypoxia and reoxygena- tion.

The antiapoptotic effect of insulin against anoxia/reoxygenation injury in cultured cardiomyocyte of neonatal rat

Objective： To study protective effect of insulin against cardiomyocyte apoptosis in anoxia/reoxygenation （A/R） injury of neonatal rat. Methods： The model of A/R injury was finished through receiving anoxia for 2h and reoxygenation for 4h in cultured cardiomyocytes of neonatal rat. The cardiomyocytes were divided randomly into 3 groups： control group （CON）, anoxia/reoxygenation group （A/R） and insulin-treated group （INS）. At the end of reoxygenation of 4 hours, activities of lactate dehydrogenase （LDH）, contents of malondiaidehyde （MDA）, were assessed through spectrophotometric procedures, myocyte apoptosis were detected through TUNEL and DNA Ladder. Results： MDA, LDH, and Apoptosis Index were significantly decreased in INS group compared with A/R group （P〈0.01）. Conclusion： Insulin has a protective effect against A/R injury in cultured cardiomyocyte of neonatal rat; the protective mechanism may contribute to antiapoptosis of insulin.

Effects of docosahexaenoic acid or arachidonic acid supplementation on gene expression and contractile force of rat cardiomyocytes in primary culture

While fatty acids play essential roles in the physiology of the myocardium,conventional culture media contain little lipid.We previously revealed that rat neonatal myocardium mainly contains docosahexaenoic(DHA),linoleic(LA),and arachidonic(AA)acids as polyunsaturated fatty acids(PUFAs),and these contents in cultured cardiomyocytes derived from fetal rats were markedly lower than those in the neonatal myocardium.In this study,we first assessed the effects of supplementation of DHA,LA,or AA on the fatty acid contents and the percentage change of contractile area in primarily cultured rat cardiomyocytes.Based on this assessment,we then evaluated the effects of DHA or AA supplementation on mRNA expression and further directly measured the contractile force of cardiomyocytes with the supplementations.This study revealed that percentage change of contractile area was maximized under 20μM DHA or 50μM AA supplementation while LA supplementation did not affect this contraction index,and that a widespread upregulation tendency of the mRNA expression related to differentiation,maturity,fatty acid metabolism,and cell adhesion was seen in the cultured cardiomyocytes with supplementation of DHA or AA.In particular,upregulation of the gene expression of cellular adhesion molecules connexin43 and N-cadherin were remarkable,whereas the effects on differentiation and maturation were less pronounced.Correspondingly,the increase of the percentage change of the contractile area of cardiomyocyte clusters in culture dishes with the supplementations was significant,whereas the enhancement of the contractile force was modest.These results suggest that supplementation of DHA or AA to the fetal cardiomyocyte culture may play effective roles in preventing the de-differentiation of the cardiomyocytes in culture and that the enhancement of the contractile performance may be mainly attributed to the improvement of intercellular connection.

A Comparative Study on the Therapeutic Effect of Astragaloside (黄芪总皂甙) and Perindopril in Treating CVB_3-Infected Cardiomyocytes in Rats

Objective: To compare the therapeutic effects of Astragaloside and Perindopril on myocardial sarco/endoplasmic reticulum Ca 2+ ATPase (SERCA) activity and the SERCA type 2 mRNA level in Coxsackievirus B 3 (CVB 3) infected cardiomyocytes. Methods: Cultured cardiomyocytes of rats were divided into normal, model, Astragaloside and Perindopril groups. The model, Astragaloside and Perindopril groups were infected with CVB 3. Meanwhile, the Astragaloside and the Perindopril groups were treated with Astragaloside (10 μg/ml) and Perindopril (1.3 μg/ml) respectively. Cytopathic effect (CPE), cardiac troponin I ( cTnI) , the SERCA activity and mRNA level of the SERCA type 2 were observed after 96 hours. Results: The CPE and cTnI of model group were significantly higher than those of normal, Astragaloside and Perindopril groups ( P <0.01). The activity and the mRNA expression of myocardial SERCA of model group were significantly lower than those of normal, Astragaloside and Perindopril groups ( P <0.01-0.05). Compared with Astragaloside group, the CPE, cTnI of Perindopril group were higher and the activity and mRNA level of Perindopril group were lower. But there were no significant difference between the two groups ( P >0.05). Conclusion: Astragaloside and Perindopril were able to reverse the down regulations of cardiac SERCA activity and mRNA expression caused by virus infection to alleviate the cardiomyocyte injury.