There have been a few studies using cultured cardiac myocytes in the experimental field of cardiac surgery. In this paper, we demonstrated the effectiveness and advantage of our cultured myocyte model for the evaluation of the myocardial damages during cardiac surgery, in which the heart is exposed to ischemic and postischemic reoxygenation (reperfusion) conditions.
In our study, the ventricles removed from neonatal rats were minced into fine fragments, which were digested by 0.1% collagenase and passed through a wire-mesh screen to remove large aggregates and debris. The cellular filtrate was suspended in culture medium containing 5% fetal calf serum (FCS) to separate cardiac myocytes from fibroblasts by differential adherence protocol. After incubation, unattached cells were resuspended in culture medium containing 2% FCS, transferrin and insulin (>95% myocytes). On the 4th day of culture, the myocytes were incubated under various conditions according to experimental protocols to evaluate the cellular injury. The functional effects were evaluated by myocyte beating rate recovery and the biochemical effects were evaluated by creatine kinase and lactate dehydrogenase release into incubation media.
The results may not contribute directly to in vivo conditions. However, this procedure provides a well-defined myocyte-culture system that is a useful model for evaluating the direct effects on myocyte injury by various environmental or chemical stimuli, including hypothermia or ischemia.

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We have established a novel measurement instrument to measure the chronotropy and the contractility of cultured neonatal rat cardiac myocytes. We used a Fotonic Sensor^TM, a fiber optic displacement measurement instrument. The principle of the measurement is to detect changes in the distance between the probe and myocytes vertically extruded by the contraction. Our results indicate that the Fotonic Sensor ^TM is an appropriate instrument for evaluating the chronotropy and the contractility of cultured myocytes. Although cultured neonatal rat cardiac myocyte models have both merits and demerits, they are useful for elucidation of the pharmacological research and for development of drugs and therapies.

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Myocardium consists of cardiac myocytes, fibroblastic cells, and vascular cells, mainly. However, the clinical significance of cell-cell interaction between these cells is controversial. Cardiac myocytes contain fibroblast growth factors (FGF-1,2) and endothelin1(ET-1). On the other hand, cardiac fibroblastic cells also synthesize ET-1. The synthesis of FGFs and ET-1 in the myocardium are reported to be stimulated in the ischemic lesions. FGFs stimulate the growth of vascular cells and fibroblastic cells, therefore, FGFs may contribute to the formation of vascularization or fibrosis. Because ET-1 stimulates cardiac contraction, ET-1 may contribute the maintenance of contractile ability in the ischemic myocardium. Pathophysiological roles of FGFs and ET-1 in heart are discussed at the point of view of cell-cell interaction between cardiac myocytes and cardiac fibroblastic cells.

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Infectious virus contamination is an important issue for security of researchers in the field of cell and tissue culture. To the exclude the possibility of accidental infection of HIV-1 to cell culture workers, we examined 254 human and monkey cell lines by a PCR protocol for detection of latent HIV-1 DNA in cultured cell lines. With this protocol, the partial HIV-I gag DNA is detected with high sensitivity; a sample containing only 55 copies in a plasmid DNA or in 10 HIV-1-infected cells showed as positive in the test. So far, none of the cell lines examined showed evidence of latent infection with HIV-1. The results show that, at present, primate cell lines in common use in Japan can be considered to be HIV-I virus free.

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We have developed a new skin model which was composed of two types of collagen sponge and two types of human cells. This skin model has two layers system, dermis and epidermis, like a real skin. In immunohistochemical study, differentiation markers of epidermis, high-molecularweight cytokeratin and involucrin, were detected in the skin model.
Furthermore, we performed an alternative study of animal testing using the skin model. Ten test chemicals were evaluated their irritancy with M I I assay which evaluates cell viability by measurement of its mitochondria's dehydrogenase activity. In the result, the skin model could distinguish non- and severe irritant.

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