Metabolism human fibroblasts

Imidazole antimycotics, ketoconazole, clotrimazole, and miconazole are potent inhibitors of various cytochrome P450-isoenzymes that also affect the metabolism of retinoids. They were fust shown to inhibit the metabolism of RA in F9 embryonal carcinoma cells. When tested in vitm liarazole, a potent CYP-inhibitor, suppressed neoplastic transformation and upregulated gap junctional communication in murine and human fibroblasts, which appeared to be due to the presence of retinoids in the serum component of the cell culture medium. Furthermore, liarazole magnified the cancer chemopreventive activity of RA and (3-carotene in these experiments by inhibiting RA-catabolism as demonstrated by absence of a decrease in RA-levels in the culture medium in the presence of liarazole over 48 h, whereas without liarazole 99% of RA was catabolized. In vivo, treatment with liarazole and ketoconazole reduced the accelerated catabolism of retinoids and increased the mean plasma all-irans-RA-concentration in patients with acute promyelocytic leukemia and other cancels. 

These interrelationships may occur in the plasma compartment, on the surface of cells, or within the cell. Our purpose here will be to review briefly some of the previous work in the above area and to present some of our recent, preliminary data on GSL and lipoprotein metabolism. Our approach has been to study simultaneously cultured human fibroblasts derived from both normal subjects and those heterozygous or homozygous for familial hypercholesterolemia (FH), a relatively common disorder of cholesterol and low density lipoprotein (LDL) metabolism. 

The metabolism of HDL probably involves interaction with both hepatic and peripheral cells, as well as with other lipoproteins. HDL may remove cholesterol from tissues, the "scavenger hypothesis (11,12). The cholesterol may then be esterifed by the action of lecithin cholesterol acyl transferase. HDL may provide cholesterol to the liver for bile acid synthesis (13) and some HDL may be catabolized by the liver in the process. HDL has not been found to interfere with the binding of LDL in cultured human fibroblasts (6). However, in cultured human arterial cells, porcine or rat hepatocytes, and rat adrenal gland, there appears to be some competition of HDL with LDL binding sites, suggesting the presence of a "lipoprotein-binding" site (14). 

The metabolism of GSLs has been studied in cultured human fibroblasts from normal subjects, patients with lipid storage diseases, and those with FH. The content of the GSLs, as well as activities of the biosynthetic enzymes, the glycosyltransferases and the lysosomal GSL hydrolases,have been studied. Complex gangliosides, such as M1, GDla, have been found in this cell system to serve as receptors for cholera toxin and thyrotropin, respectively (24-26). More recently, GT1 and GDla have been postulated to be receptors for fibronectin in cultured fibroblasts... 

One possible explanation for the alteration of GSL metabolism in T.B. is that the abnormality is related to or dependent upon the characteristics of cell growth in culture. To address this possibility, we co-cultured the cells from T.B. along with normal human fibroblasts. For comparative purposes, we also... 

Grider, A., and Young, E. M. (1996). The acrodermatitis enteropathica mutation transiently affects ziiK metabolism in human fibroblasts. /. Muir. 126, 219-224. 

Russell, J.D., Russell, S.B. and Trupin, K.M. (1978). Differential effects of hydrocortisone on both growth and collagen metabolism of human fibroblasts from normal and keloid tissue. J. Cell. Physiol. 97, 221-223. 

The feeding of an EPA-free source of supplementary DHA (from algae) to human volunteers indicated a metabolic retroconversion of DHA to EPA (Conquer and Holub, 1996, 1997). Previous animal and in vitro studies in isolated rat liver cells have demonstrated that DHA can be retroconverted to EPA, and that this retroconversion is a peroxisomal function (Schlenk et al., 1969 Gronn et al., 1991). Studies in isolated rat liver cells by Schlenk et al., 1969 have also indicated that the resultant EPA can be chain-elongated to DPA (22 5n-3) for subsequent esterification into cellular lipids. An acyl-CoA oxidase has been identified as the enzyme responsible for the chain shortening of DHA in the peroxisomal beta-oxidation of PUFA in human fibroblasts (Christensen et al., 1993). The aforementioned in vivo human studies have estimated the extent of retroconversion of DHA to EPA to be approximately 10% (Conquer and Holub, 1996, 1997). 

TCII plays a major role in transport of cobalamins to tissues. A receptor for the TCII-B12 complex has been tentatively identified in vitro on HeLa cells,Ehrlich ascites cells,and human fibroblasts. The bound complex is transferred into the cell, where the cobalamin is released and the TCII is degraded in lysosomes (Figure 38-18). An inborn error of vitamin Bj2 metabolism has been attributed to a defect in vitamin B12 release from lysosomes. A congenital... 

At high concentrations, sodium selenite induces unscheduled DNA synthesis and chromosome aberrations in cultured human fibroblasts (Lo et al. 1978). The addition of a metabolic activator (S9 fraction) or glutathione increased both the number of aberrations and the toxicity of sodium selenite (Whiting et al. 1980) and sodium selenate (Lo et al. 1978 Whiting et al. 1980). 

Poot, M., Verkerk, A., Koster, J. F and Jongkind, J. F. (1986) De novo synthesis of glutathione in human fibroblasts during in vitro ageing and in some metabolic diseases as measured by a flow cytometric method. Biochim. Biophys. Acta 883(3), 580-584. 

RDX was shown to be inactive in the UDS in WI-38 human fibroblasts, with and without S9 metabolic activation [47], Reported test concentrations were in the range of 250 to 4000 pg ml-1, well over the aqueous solubility of the compound (approximately 70 pg ml-1) [48], Similarly, in the HGPRT-V79 assay with or without metabolic activation, both RDX and HMX were inefficient in inducing 6-thioguanine resistant cells. The test concentrations used were up to 40 pg ml1 for RDX and 10 pg ml-1 for HMX, close to the solubility limit of these compounds in water [2], Another confirmation of the lack of mutagenic activity of RDX was provided recently when the compound was found inactive in the mouse lymphoma forward mutation assay using the L5178Y cell line [49],... 

Fig. 5. Cyclic changes in cholesterol metabolism that occur in cultured human fibroblasts when LDL is removed from the culture medium (-LDL) and is subsequently returned to the medium (+LDL). The relative level of each constituent is indicated by the size of the square. Reproduced, with permission of the authors and publishers, from Goldstein and Brown [10].

Rizzo W.B., Craft D.A., Dammann A.L., Phillips M.W., Fatty alcohol metabolism in cultured human fibroblasts. Evidence for a fatty alcohol cycle. The Journal of biological chemistry 262 (1987) 17412-17419. 

The LDL Pathway in Human Fibroblasts A Receptor-Mediated Mechanism for the Regulation of Cholesterol Metabolism Joseh L. Goldstein and Michael S. Brown... 

Jacobson EL, Antol KM, Juarez-Salinas H, Jacobson MK (1983) Poly(ADP-ribose) metabolism in ultraviolet irradiated human fibroblasts. J Biol Chem 258 103-107... 

The cellular content of NAD the substrate for poly(ADP-ribose) polymerase is probably one of the important factors regulating the enzyme activity. Several studies relate the significance of NAD metabolism to the poly(ADP-ribosyl)ation status of the acceptor proteins under a variety of experimental conditions (13). We find here (Table 3) that there is an inverse relationship between the levels of poly(ADP-ribose) polymerase activity and NAD levels of Ewing s sarcoma and laryngeal squamous cell carcinoma and lung carcinoma as compared to normal human fibroblasts. 

Joseph J, Ranganathan S, Mehta JL. Low density lipoproteins modulate collagen metabolism in fibroblasts. J Cardiovasc Pharmacol Ther 2003 8 161-166. Lemaitre V, O Byme TK, Borczuk AC, et al. ApoE knockout mice expressing human matrix metalloproteinase-1 in macrophages have less advanced atherosclerosis. J Clin Invest 2001 107 1227-1234. 

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