Abnormal monocyte chemotaxis in hamsters with human osteosarcoma

ABNORMAL MONOCYTE CHEMOTAXIS

Kwong Y. Tsang,

IN HAMSTERS WITH HUMAN OSTEOSARCOMA

Iqbal Singh, Mark Gnagy and Diane Tanski

Division of Orthopaedic Surgery, Medical College of Ohio C. S. 10008, Toledo, Ohio 43699, U.S.A.

Monocytes from cancer patients, such as carcinoma of the prostate (i), breast (2), malignant melanoma (3) and human osteosarcoma (4) have abnormal chemotaxis in vitro. The present study was undertaken to study the effect of osteosarcoma on monocyte functions in hamsters. The monocyte chemotactic responsiveness (MCR) of thirty normal hamsters and twenty-five hamsters with human osteosarcoma were investigated. Human osteosarcomas were induced in inbred hamsters (LSH/SsLAK) by techniques described by Singh et al (5,6). Laparotomies were performed on pregnant (12-14 days gestation) hamsters. The uterus was exposed and the fetuses (6-10 per hamster identified. A homogenate of 2 x 106 TE-85 cells (derived from a biologically proven human osteosarcoma) in 0. i ml of medium 199 was injected into each fetus through the intact uterine wall. 2 x 106 TE-85-M~ISV cells (TE-85 cells infected with MSV-RD-II4) pseudo type virus) suspended in 0.25 ml of medium 199 were injected adjacent to the midshaft of the femur of 4 day old newborn hamsters. In this study, human osteosarcomas were induced in 4 day old hamsters by injecting 1 x 107 osteosarcoma cells (obtained from the tumored hamsters, induced by a laparotomy procedure) adjacent to the midshaft of the femur without a laparotomy procedure. Tumors developed 12-16 days after injection. Monocytes were isolated from whole heparized blood by the Ficoll-Hypaque density gradient method. The cells were resusDended in serum free RPMI-1640 medium supplemented with penicillin 200 units/ml, streptomycin 200~g/ml. The cell suspension was adjusted to (I x 106 monocytes/ml for use in the chemotaxis assays. The chemotactic factor was generated in fresh hamster serum by incubation with 1 mg of Salmonella typhosa 0901 endotoxin/ml of serum at 37°C for 1 hour followed by 56°C for 30 minutes. Boyden Chemotaxis chambers were used for the assays. Monocytes were separated from the stimulus with a 5.0~ pore size polycarbonate filter. Chemotaxis was quantitated by counting and averaging five randomly selected oil immersion fields and the total number of monocytes that had migrated. The functional chemotactic index (FCI) of the normal and tumored hamsters were compared. The MCR of tumored hamsters was significantly depressed (mean=51.1±7.6) as compared to normal hamsters (mean=68.5±14.1) (Fig. i). Scanning electron microscopy studies on the monocyte chemotaxis of normal and tumored hamsters (Fig. 2,3,4) indicated that more cells migrated onto the bottom surface of the membrane in the control chamber as compared to the experimental chamber. The FCI was 106 for the normal hamsters and 74.5 for the hamsters with human osteosarcoma (p<0.05). The results support the hypothesis that neoplasms suppress monocyte function which may affect cell mediated destruction of malignant cells. REFERENCES Boetcher, D.A., and Leonard, E.T., Abnormal monocyte chemotactic response in cancer patients J. Natl. Cancer Inst. 52, 1091 (1974). Synderman, R., Meadows, L., Holder, W., Abnormal monocyte chemotaxis in patients with breast cancer - evidence for a tumor mediated effect. J. Natl. Cancer Inst. 60,737 (1978). Rub±n, R.H., Cos±m±, A.B. and Goelzd, E.J., Defective human mononuclear leukocyte chemotaxis SN index of host resistance in malignant melanoma. Clin. Im~aunol. ImmunopathoL 6,376 (1976) Tsang, K.Y. and Singh, I., Monocyte chemotaxis in osteosarcoma patients and animals (abstract) Proceedings ASM 60, (1979). Singh, I., Hatheway, T.M. and Tsang, K.Y. et al, An animal model for human osteosarcoma Surgery, 81,168 (1977). Singh, I., Tsang, K.Y. and Blakemore, W.S., A model for human osteosarcoma in hamsters Clin. Ortho. Rel. Res. In Press (1979).

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Fig. 1 Monocyte chemotactic responsiveness of normal hamsters and hamsters with human osteosaromca

Fi~. 2 Scannin~ electron micro~raph of normal hamster monocytes which have migrated to the bottom surface of a po]ycarbonate filter in response to activated serum (X 6]5)

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Fig. 3 High magnification of normal hamster monocytes migrated through a 5.0 ~ pore (X 3300)

Fig. 4 Scannin~ electron micro~raph of MCR of monocytes isolated from tumored hamsters. The ~ICR was significantly depressed when compared to monocytes isolated from normal hamsters

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