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There have been few reports on investigation of the cell kinetics of brain tumors using a laser scanning cytometer (LSC). We compared DNA analysis using a flow cytometer (FCM) with that by LSC using established cell lines of brain tumors, and we demonstrated the similarity of both analytical results. The DNA-index (DI) and proliferating index (PI) of various brain tumors were determined, and the results showed that these indices reflected the malignancy of the tumors. The DI values were higher for astrocytoma grade IV than for astrocytoma grade II, frequently showing an aneuploid pattern. The determination of DNA-ploidy using LSC, together with cryopathological diagnosis, was considered to be useful for deciding the area to be resected for glioma perioperatively and to be worth applying clinically.

Cyclins are key components of the cell cycle progression machinery. Immunohistochemical detection of cyclins in relation to cell position, DNA content, by multiparameter flow cytometry has provided new cell cycle studies. T98G, U373MG, and U251MG were scheduled expression of cyclin A, B1 which are maximal in late S and G2M phase cells. T98G was scheduled expression of cyclin E. being maximal in G1 and S phase cells, but U251MG, U373MG were unscheduled expression of cyclin E. The expression of cyclin D1 were not expressed a pattern. These result may mean that cyclin E in an important protein of cell cycle machinery of malignant brain tumor.

Meningioma is one of the popular benign brain tumors. However, the recurrence of this tumor is not infrequently encountered. In an attempt to establish the useful adjuvant therapy for the recurrent meningioma, in vitro chemosensitivity study for meningioma was conducted. Among various chemotherapeutic agents tested here, cisplatin showed highest cytotoxicity on cultured meningioma cells. In conclusion, cisplatin may be useful in adjuvant chemotherapy for the recurrent meningioma.

Antineoplastic effects of interferons (IFNs) on brain tumors have often been reported in the literature, however, so far as we know, there are no reports of the study on the antineoplastic effect of IFNs (alpha, beta, and gamma) labelled with fluorescein isothiocyanate (FITC) using flow cytometry (FCM). Three established glioma cell lines and 11 cultured cells of brain tumor from surgical specimens were exposed to IFN-alpha, beta, and gamma at the concentrations of 10(2)-10(5) IU/ml for 24 h, respectively. Using FCM, the viability of the cells was evaluated with fluorescein diacetate stain and the cell cycle was analyzed from the DNA-histogram with propidium iodide stain. Furthermore, FITC-labelled IFN-alpha, beta and gamma were also contacted with each cell to calculate respective positive cells. The viability decreased about 60% on day 1 and day 3, indicating the effect of IFN-alpha and beta on U373MG cells and on some cultured glioma cells from surgical materials, whereas, IFN-gamma had no effects. Antineoplastic effect of each IFN well correlated with FITC-positive rates, demonstrating S phase block in the cell cycle. IFN-gamma had no antineoplastic effects, whereas IFN-alpha and beta showed antineoplastic effects, which fact suggested that IFN-gamma receptor be different from those of IFN-alpha and beta. The method of FITC-labelling for IFNs with the aid of FCM has the advantages as follows: 1) Antineoplasticity of IFN can be simply evaluated with FCM; 2) It is easy to analyze the action mechanism of IFN; 3) Information on the receptor is obtainable; and 4) Sensitivity can be evaluated prior to administration of IFN, suggesting possibilities of clinical application of this method.

For the purpose of FCM, we have developed a new method of sterile sorting of NK cells by negative selection using FITC-labelled monoclonal antibodies. In this method, over 95% lymphocytes with large granular lymphocytes (NK cells) were obtained by exclusion of Leu-1 and HLA-DR positive cells from human peripheral blood. From the results of cytotoxicity test of NK cells against 4 types of glioma cell lines and 6 kinds of cultured cells from surgical materials, the antineoplastic effect was recognized in 4 out of 6 surgical materials. Administration of NK cells through an Ommaya tube in 2 brain tumor patients caused no side effects. This technique is clinically applicable because 1) NK cells are easy to obtain, 2) NK cells show antineoplastic effect on brain tumors, and 3) the sensitivity can be evaluated in individual patients.

With the aid of flow cytometry (FCM), distribution of DNA content in 40 cases of brain tumour, primary culture cell, and secondary culture cell can be determined and chronological change after subculture is studied from the analysis of their cell cycle. In most primary cultures, proliferating index (PI) is likely to decrease, which suggests that environmental change might affect the growth activity. In comparison with that of the original sample, DNA-histogram of the secondary culture can be divided into the following 3 types: the type recovering to the original pattern ("adapting type"), in which astrocytoma, ependymoma, glioblastoma and medulloblastoma are included, 2) the type increasing more at G2 + M phase than the original ("proliferating type"), in which meningioma and some of glioblastoma are included, and 3) the type decreasing so far as to induce degeneration or death ("degenerating type"), in which pituitary adenoma and neurinoma are included. FCM is of great usefulness for the study of cell kinetics of a tumour cell undergoing culture and the present method will be available for the respective study of biological characteristics of the cultured cell, established cell line or sensitivity test for antineoplastic agents.