1~. J. Biochem. Vol 23, No. 9, pp. 819-821, 1991 Printed in Great Britain. All nghts reserved 0020-711X/91 $3.00+0.00 Copyright0 199 I Pergamon Press plc ROLE OF A CALF THYMUS PREPARATION IN THE DEGRADATION OF NATIVE AND REDUCTIVELY METHYLATED LOW DENSITY LIPOPROTEIN PAOLO MONDOLA, MARIAROSARIA SANTILLO, LUCIO CAMMAROTA and FRANCO SANTANGELO Istituto di Scienze Fisiologiche Umane, II Facolta’ di Medicina e Chirurgia, Universita’ di Napoli, via S. Pansini 5, 80131 Napoli, Italia [Tel. (081) 545-30221 (Received 10 September 1990) zyxwvutsrqponmlkjihgfedcbaZYXWVUTSRQ Abstract-l. The clearance of low density lipoprotein (LDL) is mediated by a specific LDL receptor pathway and by an alternative metabolic pathway that is responsible for the receptor-independent LDL catabolism. 2. This alternative catabolism can be studied in vivo using a preparation of chemically modified LDL that are reductively methylated. 3. Recently we showed that a calf thymus protein extract affects the cholesterol metabolism via activation of LDL catabolism. 4. The aim of this study was to investigate whether in vivo the specific LDL receptor pathway and the independent LDL receptor pathway are affected by thymus treatment. 5. The results obtained injecting in rats native and chemically modified ‘r51-LDL to probe the receptor independent pathway, show that the thymus gland decreases serum cholesterol by activation of the specific LDL receptor pathway. 6. This effect is mainly evident in liver and kidney that represent organs in which the specific LDL receptors are widely present. INTRODUCIION Cells have specific receptors for low density lipo- protein (LDL) that are responsible for the catabolism of this lipoprotein, the major cholesterol-carrying vehicle in human plasma (Brown and Goldstein, 1975; Goldstein and Brown, 1977). The number of these receptors is regulated by the cell requirement of cholesterol through the so-called “LDL receptor pathway” (Goldstein and Brown, 1977), which is a finely regulated mechanism. When cells are deprived of cholesterol they synthesize more LDL receptors and therefore more cholesterol is internalized. On the contrary, when the intracellular level of free cholesterol greatly increases, the synthesis of LDL receptors decreases and LDL uptake declines (Brown et al., 1981; Brown and Goldstein, 1983). However, there is an alternative route for LDL catabolism, called “the scavenger pathway” or “receptor-independent LDL catabolism” (Goldstein et al., 1977), which does not seem to be subjected to regulation by intracellular cholesterol (Attie et al., 1982). The methylated LDL does not bind to the classical LDL receptor and therefore provides a measure of non-specific tissue uptake of LDL (Inner- arity et al., 1986). In animals and humans that lack the LDL receptor pathway, the receptor-independent pathway is essentially responsible for all LDL turnover. The receptor-independent pathway plays a prime role in the catabolism of LDL in rat (Koelz et al., 1982; Spady et al., 1985a,b) accounting for the Abbreviations: LDL, low density lipoprotein; FII, thymus fraction; MeLDL, reductively methylated LDL. turnover of about 2@40% of the circulating LDL. In the rat, the spleen exerts an important role, while the liver and endocrine glands have a low clearance of LDL (Spady et al., 1985a,b). Recently, we investigated the mechanism by which the thymus gland is able to affect lipid metabolism in the rat (Mondola et al., 1987) and found that the addition of our calf thymus extract to the culture human fibroblasts and rat liver cells increased the binding and internalization of human and rat ‘251- labeled LDL (Mondola et al., 1989b). These data and those obtained very recently (Mondola et al., 1989a) suggested that the thymus gland affects cholesterol metabolism via activation of the LDL receptor path- way. As part of LDL catabolism in rats is mediated by a mechanism independent of the specific high affinity receptor for LDL (Carew et al., 1982) it is important to investigate which of these two mechanisms is involved in the activation of LDL catabolism and which tissues are mainly affected by thymus treat- ment. MATERIALS AND METHODS Lipoprotein preparation Human LDL, density 1.019-I ,063 g/ml, was prepared from pooled serum of normolipidemic subjects by sequential ultracentrifugation according to the method of Have1 et al. (1955). The radiolabeling of human LDL was performed following the iodine mono-chloride method of McFarlane (1958) modified by Bielheimer et al. (1972). Five milligrams of LDL were incubated with 2mCi of Na’251 in 0.1 N NaOH (Amersham International plc, U.K.) as suggested by Goldstein et al. (1983). The final specific activity of ‘251- labeled human LDL was about 360 cpm/ng of protein. 819