Hytakerol-Induced Metastatic Calcification in Trypanosomiases.

Heavy infections with Trypanosoma equiperdum and T. equinum reduced significantly the overall incidence of hytakerol-induced metastatic calcification, while infections with T. cruzi had no comparable effect. Differences between T. cruzi infections and infections with the African trypanosomes were also apparent in respect to calcification incidence of certain organs, e.g., the liver was definitely more calcified in the former than in the latter, while an opposite relationship was found in regard to the mosaic type of calcification of the stomach mucosa. No calcification around T. cruzi foci was found in any of the invaded tissues. Cold stress and cortisone administration increased rather than decreased the calcification susceptibility of uninfected rats. The question as to what role, if any, nonspecific stresses produced by the infection play in modifying the calcification response to hytakerol administration, cannot be decided unequivocally at present. In previous investigations of metastatic calcification induced by hytakerol in animals infected with malaria (Mercado and von Brand, 1962, 1964), significant differences were observed between two species of malaria, i.e., Plasmodium berghei in rats and P. gallinaceum in chickens. In earlier work von Brand and Holtz (1933) observed that the response of canaries infected with P. praecox to the calcifying effects of another ergosterol derivative, 7-dehydrocholesterol, also differed from that of the infected animals mentioned above. The differences involved mainly the overall incidence of calcification as compared to similarly treated control specimens, but differences in the susceptibilities of individual organs also occurred. Von Brand and Mercado (1956) and Mercado and von Brand (1960) reported that various parasitic infections such as malaria and trypanosomiasis induced different changes in liver glycogen and lipid. It was thought of interest, therefore, to extend the calcification studies to trypanosomiasis and to investigate whether or not the response of the host to the calcifying effect of hytakerol would be modified in this infection also. It was considered that such a comparative approach might shed light on the difficult question whether changes in physiological reactions in parasitized animals are due primarily to nonspecific stress produced by the presence of parasites or to specific influences from the parasites (e.g., toxins). Received for publication 18 September 1965. MATERIALS AND METHODS The United States Department of Agriculture strain of Trypanosoma equiperdum, the Wellcome strain of T. equinum, and the Tulahuen strain of T. cruzi were used. The T. equiperdum and T. equinum infections were developed in 6-week and 8-week-old Osborn-Mendel female rats, respectively. Six-week-old CFW mice were used for the T. cruzi infection. Generally 5,000 to 10,000 trypanosomes were inoculated intraperitoneally. Hemocytometer counting of the trypanosomes was done in glucose saline (1.6 g glucose in 800 ml normal saline, pH 7). This medium prolonged the survival of the flagellates and made counting possible for as long as 1 hr after withdrawal of the blood. Treatment with hytakerol (dihydrotachysterol concentrate, 20 X in sesame oil, with an activity equivalent to about 5 mg U.S.P. XVI dihydrotachysterol crystals per ml, Winthrop Laboratories) was initiated on the same day the rats were inoculated with T. equiperdum, 4 to 6 days after inoculation with T. equinum, and 6 days after the T. cruzi inoculations. The younger rats received one daily orally administered dose of 0.1 ml drug per 100 g body weight. The older rats and the 6-week-old mice received 0.08 ml and 0.3 ml, respectively. The drug was given for 4 days and the animals were killed on the 5th day. The main experimental series consisted of infected and uninfected animals which were treated with hytakerol, but a few shorter series consisting of uninfected and infected animals which did not receive hytakerol were completed also. In addition, two other series were done to test the possible influence of stress, induced by cold exposure and cortisone, on the calcifying effects of hytakerol in uninfected rats. In the first series rats were exposed to a temperature of 4 C for periods of 5, 10, and 17 days, corresponding controls being maintained at room temperature (22 C). Both groups were treated with hytakerol for 4 days as follows: The animals of the 5-day group received the drug starting on the 1st day of exposure and