Effect of Embryo Extract on Growth of Fibroblasts Previously Irradiated with X-rays

Abstract

Embryo extract is a powerful growth-stimulant for cell colonies in vitro. It is generally accepted that its influence is manifested in, and limited to, the activation of cell-division. Our observations on the effect of X-rays on cell cultures make it necessary to revise this view. The investigations reported here demonstrate that cell cultures deprived of mitotic capacity by previous irradiation with X-rays will also grow at a considerably increased rate after the addition of embryo extract.

The experiments were performed on standardized cultures of chicken fibroblasts derived from the hearts of 7-days-old embryo The cell colonies were cultivated in hanging drops according to the method of Carrel. After the irradiation, they were planted into Carrel flasks. Growth curves of the culture were constructed from planimetric measurements of outline drawings of the surface areas made every 24 hours. Irradiation was carried out using a demountable X-ray tube which worked at a tension of 35 KV on currents 20 MA. The tube had a copper anticathode and a window of aluminium foil, 30 μ thick. Absorption analysis showed that the rays penetrating through the window foil and the mica coverglass of the cultures were mainly copper K-rays.

The following procedure was adopted: Preliminary studies having shown that the X-ray dose necessary to inhibit completely cell multiplication is 10,000 r units, cultures of fibroblasts were irradiated with 25,000 r units, i.e., a dose 2 1/2 times greater than that required to arrest all mitoses. The cultures irradiated were divided into two equal parts: one fragment was planted into a medium which consisted of chicken plasma diluted with Tyrode's solution in the proportion 1:2; the other half was cultured in a medium of the same composition to which 30% embryo extract was added as a fluid phase.