The Sertoli cells have been identified as the primary locus for creatine synthesis within the seminiferous epithelium. The purpose of the studies reported here was to examine the effect of modulators of Sertoli cell function on creatine secretion by primary cultures of these cells. Sertoli cell-enriched cultures, maintained in a defined medium, secreted creatine into the incubation medium in a manner that was linear with time over at least 6 h, but which had reached a plateau within 24 h. Secretion was stimulated by physiological and toxicological modulators of Sertoli cell function. Incubation of Sertoli cell-enriched cultures in the presence of FSH (> or = 40 mU ml-1), dibutyryl cyclic AMP (> or = 0.1 mmol l-1), mono-(2-ethylhexyl) phthalate (> or = 1 mumol l-1) or cadmium (> or = 3 mumol l-1) increased the secretion of creatine into the incubation medium by at least 85% over 24 h. Creatine secretion by Sertoli cell-enriched cultures, incubated over 4 h in a balanced salt solution, was independent of exogenous L-glutamine. However, the stimulation of secretion induced by 1 mmol dibutyryl cyclic AMP l-1 was dependent on the presence of 4 mmol L-glutamine l-1 in the incubation medium, which suggests that an increase in creatine secretion occurs as a consequence of stimulated glutamine oxidation.

The testis synthesizes creatine from both arginine and glycine precursors, but when rat testicular tissue is separated into seminiferous tubules and interstitial cells, creatine synthesis occurs only in the tubular fraction. The purpose of the work presented here was to define the locus of creatine synthesis within the seminiferous tubules, by using cell separation and culture techniques to examine synthesis in the Sertoli cells and germ cells. The total creatine content, in the cellular compartment and incubation medium, of Sertoli-germ cell co-cultures and of Sertoli cell-enriched cultures, largely free of germ cells, increased by similar amounts over a 24 h incubation period. Sertoli cell-enriched cultures incorporated radioactivity from L-[guanidino-14C]arginine and [1-14C]glycine into both creatine and its biosynthetic precursor, guanidinoacetic acid. Isolated germ cells did not incorporate radioactivity from L-[guanidino-14C]arginine into either creatine or guanidinoacetic acid when incubated at a similar density and protein concentration under similar conditions. It is concluded that the synthesis of creatine observed in isolated rat seminiferous tubules occurs within the Sertoli cells and not the germ cells.

The current work concerns the development and validation of an in vitro reporter gene assay system for the assessment of induction of human CYP3A4. A plasmid containing approximately 1 kb of the CYP3A4 regulatory region (which contains several recognised regulatory elements including glucocorticoid responsive elements) coupled to the reporter gene for human secreted placental alkaline phosphatase (SPAP) was transfected into the human hepatoblastoma cell line HepG2. Calcium phosphate precipitation was the method of choice for transfection. The transfected cells were dosed with known inducers of CYP3A4 and the levels of SPAP in the medium were subsequently measured using a chemiluminescent assay, as an indirect measure of CYP3A4 induction. The inducers used in this study included dexamethasone, phenytoin, triacetyloleandomycin (TAO), rifampicin, carbamazepine, phenylbutazone and sulfinpyrazone. These compounds activated CYP3A4 by between 1.5-4.5-fold thus representing a major advance in assessing the induction of human CYP genes in vitro.

Nephrotoxin-induced damage to cultured kidney cells (LLC-PK1) was investigated using assays of general cytotoxicity (neutral red uptake) and of functional impairment (glucose accumulation). The latter correctly ranked both the cephalosporin and aminoglycoside antibiotics (acute and chronic proximal tubule toxicants respectively) in terms of their relative in vivo toxicities. Functional impairment after low-dose long-term exposure to the aminoglycosides occurred at sublethal drug concentrations. Assessment of general cytotoxicity correctly ranked only the aminoglycoside antibiotics.

Conference Article| June 01 1989 Isolated rat seminiferous tubules synthesize guanidoacetic acid and creatine from arginine NIGEL P. MOORE; NIGEL P. MOORE *Toxicology Unit, The School of Pharmacy, 29/39 Brunswick Square, London WC1N 1AX, U.K. Search for other works by this author on: This Site PubMed Google Scholar JOHN A. TIMBRELL; JOHN A. TIMBRELL *Toxicology Unit, The School of Pharmacy, 29/39 Brunswick Square, London WC1N 1AX, U.K. Search for other works by this author on: This Site PubMed Google Scholar TIM J.B. GRAY TIM J.B. GRAY †British Industrial Biological Research Association, Woodmansterne Road, Carshalton, Surrey SM5 4DS, U.K. Search for other works by this author on: This Site PubMed Google Scholar Biochem Soc Trans (1989) 17 (3): 520–521. https://doi.org/10.1042/bst0170520 Article history Received: November 25 1988 Views Icon Views Article contents Figures & tables Video Audio Supplementary Data Peer Review Share Icon Share Facebook Twitter LinkedIn MailTo Cite Icon Cite Get Permissions Citation NIGEL P. MOORE, JOHN A. TIMBRELL, TIM J.B. GRAY; Isolated rat seminiferous tubules synthesize guanidoacetic acid and creatine from arginine. Biochem Soc Trans 1 June 1989; 17 (3): 520–521. doi: https://doi.org/10.1042/bst0170520 Download citation file: Ris (Zotero) Reference Manager EasyBib Bookends Mendeley Papers EndNote RefWorks BibTex toolbar search Search Dropdown Menu toolbar search search input Search input auto suggest filter your search All ContentAll JournalsBiochemical Society Transactions Search Advanced Search This content is only available as a PDF. © 1989 Biochemical Society1989 Article PDF first page preview Close Modal You do not currently have access to this content.

Many compounds have been shown to produce liver enlargement and hepatic peroxisome proliferation in rodents (Cohen and Grasso 1981; Reddy and Lalwani 1983). Organelle proliferation is accompanied by large increases in certain enzyme activities, particularly in the peroxisomal fatty acid β-oxidation cycle, which is normally measured as cyanide-insensitive palmitoyl-CoA oxidation (Lazarow and DeDuve 1976). Certain peroxisome proliferators have been shown to increase the incidence of liver tumours in rats and/or in mice and it has been suggested that tumour formation is, in some manner, related to organelle proliferation (Reddy and Lalwani 1983; Reddy et al. 1986).

A series of 9 phthalate monoesters were cultured with primary rat hepatocytes for 70 h and their effect on peroxisomal fatty acid β-oxidation (palmitoyl-CoA oxidation) determined Marked quantitative differences in the induction of enzyme activity were observed with both alkyl chain length and position of side chain substitution affecting compound potency. With all 9 compounds a good correlation was observed between electronic structural parameters obtained by molecular orbital calculations and biological activity in the cell culture system. These results demonstrate a relationship between chemical structure and biological activity for a series of phthalate monoesters and indicate the potential usefulness of primary hepatocyte cultures to screen compounds for peroxisome proliferation.

A number of chemicals have been shown to produce liver enlargement and hepatic peroxisome proliferation in rodents and certain of these compounds have also been shown to increase the incidence of liver tumours. Whilst a precise relationship between peroxisome proliferation and liver tumour formation remains to be established, species differences may be important in extrapolating rodent findings to man. In this study the effect of di-(2-ethylhexyl) phthalate, mono-(2-ethylhexyl) phthalate (MEHP) and clofibric acid in Chinese and Syrian hamsters were compared. After oral administration for 14 days all three compounds produced liver enlargement and induction of peroxisomal enzyme activities in both species but the response was more marked in Chinese hamsters. In cell culture studies both MEHP and clofibric acid produced a greater induction of peroxisomal β-oxidation in hepatocytes from Chinese than from Syrian hamsters. These results thus demonstrate a species difference in the magnitude of response to certain peroxisome proliferators after oral administration between Chinese and Syrian hamsters. As this difference was also observed in vitro the results suggest that intrahepatic factor(s), which are retained in cell culture, were primarily responsible for the observed species difference.

In a 90-day toxicity study in rats, di-(2-ethylhexyl)phthalate (DEHP) produced testicular atrophy. To characterise further the testicular toxicity of phthalate esters the effect of age on the induction of testicular atrophy has been examined as well as the reversibility of the lesions and the effects of certain other phthalate esters. Seminiferous tubular atrophy, comprising a loss of spermatids and spermatocytes, resulted when 4-week-old rats were given 10 daily doses of DEHP. In similarly treated 10-week old rats up to 50% of tubules were atrophic while the remainder were unaffected. No testicular damage was produced in 15 week-old rats. The lesion produced in 4-week-old rats was reversible whether treatment was stopped prior to, or continued until after the control rats had reached sexual maturity. Normal testicular weight and histology were restored within 12 and 20 weeks respectively. Of a series of di-n-alkyl phthalates from dimethyl to di-n-octyl, the butyl, pentyl and hexyl esters produced testicular lesions similar to DEHP. The testicular effects of DEHP were not influenced by simultaneous administration of testosterone or follicle stimulating hormone (FSH). The mechanism by which phthalate esters exert their effects is discussed in the context of a possible action on Sertoli cell function.

Groups of 15 male and 15 female rats were given diet containing 0 (control), 0·2, 1·0 or 2·0% di-(2-ethylhexyl) phthalate (DEHP) for 17 wk. At the two higher treatment levels there was a reduced rate of body-weight gain and food intake. A paired-feeding study showed that the reduced food intake did not account fully for the reduced growth rate. There was a decreased packed cell volume in both sexes given 1 or 2% DEHP, and in the male rats given these levels the haemoglobin concentration was also reduced. However, there was no decrease in the total erythrocyte count. Renal concentrating and diluting ability was reduced in the females receiving 2% DEHP. There were no differences between treated and control rats in the results of the serum analyses. With the exception of the liver weights, which were higher, the absolute weights of most organs in the rats given 1 or 2% DEHP were lower than those of the controls, while the relative values were greater. This pattern was considered to be related to the lower body weights at autopsy. At these treatment levels, the relative testis weight was markedly decreased and histopathological examination revealed severe seminiferous tubular atrophy and cessation of spermatogenesis. The testis weight of the rats fed 0·2% DEHP was not reduced, but histologically there was evidence of decreased spermatogenesis. At wk 17, castration cells were observed in the pituitary at all treatment levels, the incidence being dose-related. There were no other histopathological changes attributable to DEHP treatment. In view of the testicular changes and the increase in liver weight at all treatment levels, it was not possible to establish a no-untoward-effect level for DEHP in this study.