D. Szabó

Correlation of lipid droplet content and steroidogenic capacity in zona glomerulosa and fasciculata cells from lipoprotein-deficient rats

Lipoprotein deficiency was maintained for 48 h by applying 4-aminopyrazolo(3,4-d)pyrimidine in male albino rats and the effect on the structure and function of isolated adrenocortical cells was investigated. Neither the morphology nor the steroidogenic response of zona glomerulosa cells changed; however, in the zona fasciculata cells a marked reduction in the quantity of lipid droplets was associated with a decreased steroidogenic response to ACTH. The results indicate that despite lipoprotein deficiency, zona glomerulosa cells contain a reserve of intracellular cholesterol, thus retaining their responsiveness to stimulation. The fact that the lipoprotein deficiency does not affect lipid droplets located in the glomerulosa cells and in the fasciculata cells in a similar manner is probably due to the differences in their physical state and chemical composition.

Lipoproteins, lipid droplets, lysosomes, and adrenocortical steroid hormone synthesis: Morphological studies

Recent studies concerning cellular cholesterol homeostasis suggest that there is a relationship between the serum lipoproteins (low density and high density lipoproteins: LDL and HDL), the intracellular storage of cholesterol (lipid droplets), lysosomes, and the steroidogenic activity of adrenocortical cells. This review surveys the current knowledge on cholesterol import from LDL/HDL by adrenocortical cells, its regulation, and the participation of lipid droplets and lysosomes in this process. The possible role of adrenocortical cell microvilli in the uptake of LDL/HDL is discussed. Under certain physiological, experimental, and pathological circumstances lysosomes accumulate unesterified and/or esterified cholesterol in the form of lipid‐lyososome complexes. As suggested by the data presented in this review, lipid‐lysosome complexes appear to be involved in cholesterol homeostasis, via altering lipid compartmentalization. Since previous reports do not clearly demonstrate a positive correlation between the volume of lipid‐ and lysosome‐ compartments and the rate of steroid hormone synthesis [for review, see Nussdorfer (1986) Int. Rev. Cytol., 98:1–405], the objective of this review is to provide a better understanding of the interactions of plasma lipoproteins, lipid droplets, lysosomes, and steroidogenesis. Microsc. Res. Tech. 36:480–492, 1997.

Changes in the fine structure and function of a hormone-secreting adrenocortical tumour investigated in tissue culture

Tissue cultures of a surgically removed adrenocortical tumour causing Cushings syndrome, and tissue cultures from the attached, tumour-free adrenal were studied. There were two cell types characteristic of tumour tissue. The cell type occurring most frequently had pronounced hypertrophied agranular endoplasmic reticulum. A fewer number of lipid-rich cells containing many electron-dense granules could also be found. The ratio of cells changed during cultivation. In the 17 days tumour culture, a higher percentage of lipid-rich cells could be observed. In spite of continuous ACTH treatment, the initially high hydrocortisone level decreased, gradually. It may be assumed that the lipid-rich cells are of reduced ability as regards hydrocortisone production.

Effect of histamine on corticosteroid secretion of isolated human and rat adrenocortical cells

Slices of human adrenal cortex (obtained from renal transplant donors) and adrenals obtained from male CFY (Wistar) rats weighing 200-250g were used. The rat adrenocortical zona glomerulosa and fasciculata [1] and human adrenocortical [2] cell preparation used in our laboratory were described previously. All types of collagenase-dispersed cells were prepared in Krebs-Ringer bicarbonate buffer (pH 7.4) containing 2g/1 glucose and 40g/1 human serum albumin (HSA). Approximately 2 x l0 s cells/ml were incubated in one session, in a shaking water-bath at 37 °C under an atmosphere of 95% 02 and 5% CO2 for 2 hours. Histamine dihydrochloride (Fluka Chemie AG) was added to the samples in 10 -9 to 10 -3 M concentrations. The concentrations of aldosterone, corticosterone, cortisol, dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulphate (DHEAS) in the incubation media were determined by radioimmunoassay [2]. All data are presented as means ~z S.E.M. For statistical evaluation, Dunnetts test was applied after analysis of variance.

Morphological evidence of lysosomal uptake of high-density lipoproteins by rat adrenocortical cells in vitro

The high-density lipoprotein (HDL) pathway in rat adrenocortical cells was studied at the electron microscopic level in vitro via colloidal gold labelling. Steroid hormone assays were performed to confirm that the cells remained intact, viable, responsive to ACTH under the applied conditions, and to reveal the steroidogenic effect of HDL. The gold-labelled HDL particles (HDL-Au) were observed on the surface of the parenchymal cells, often attached to the membranes of the microvilli, but rarely in coated pits and coated vesicles. HDL-Au was accumulated by non-coated vesicles, multivesicular bodies and lysosomes. The lysosomes were identified by means of a non-specific esterase reaction. It is concluded that HDL particles are internalized by both zona glomerulosa and zona fasciculata cells. HDL is required for the enhanced functional activity of these cells in long-term incubation, and the lysosomes are involved in the process.

Interrelationship between corticosteroid production and fine structure in the fetal adrenal cortex.

Abstract The cat fetal adrenal is capable of corticosteroid production from an early stage of ontogenesis (crown-rump length: 2–3 cm) and reacts to ACTH by corticosteroid production. Its hormone-producing ability and responsiveness to ACTH operate before the fine structure characterizing the adult zona fasciculata cells is recognizable. In tissue culture of human fetal adrenal, corticosteroids are produced in response to ACTH even if the fine structure of the endoplasmic reticulum and mitochondria, on which corticosteroid production is currently believed to depend, shows marked deviation from the histological picture characterizing the zona fasciculata cells.

Viscosity of rat adrenocortical lipids in different functional states: Morphological characteristics

Impaired adrenocortical steroidogenic activity is often concomitant with morphologically and physiologically altered lipids in the cells of the adrenal cortex. The physical state of these lipid droplets and the morphological characteristics of crystal-shaped bodies were studied in different functional states of adrenocortical cells. In the perinatal period when steroidogenesis is suppressed by a negative feedback mechanism, crystal-shaped bodies (i.e. rectangular, electron-lucent formations, either alone or in clusters, surrounded by lysosomal matrix or in close proximity of lysosomes) were frequently observed in the inner zona fasciculata and zona reticularis. In experimentally suppressed adrenocortical activity, following the administration of dexamethasone, aminoglutethimide or cycloheximide, almost identical crystal-shaped bodies were frequently observed in adrenocortical cells. These crystal-shaped bodies appear to be cholesterol, as revealed by the digitonin reaction at the electron microscopic level. Following stimulation of the zona fasciculata by ACTH treatment for 14 days, a marked increase in the fluidity of the lipid droplets was observed in the thermotropic phase transitions with the polarizing microscope. In contrast, following aminoglutethimide treatment, the fluidity of the lipid droplets decreased. The thermotropic phase transitions of normal and neoplastic human adrenal cells, namely adrenocortical tumours causing Conns or Cushings syndrome, were also investigated. When hormone biosynthesis was enhanced, the appearance of birefringence and multiple phase transitions of lipid droplets was demonstrable in the low-temperature range.

Colloidal gold-labeled lipoprotein binding and internalization in adrenocortical cells in vitro.

Human (normal and adenomatous) and rat (normal) adrenocortical cells were incubated in vitro with colloidal gold labeled low-density (LDL-Au) and high-density (HDL-Au) lipoproteins, respectively, in order to visualize lipoprotein binding and internalization at an electron microscopic level. Both normal and adenomatous human adrenocortical cells accumulated LDL-Au by receptor-mediated endocytosis via coated pits, coated vesicles, noncoated vesicles, and lysosomes. HDL-Au was not internalized. In rat adrenocortical cells, both HDL-Au and to a lesser extent LDL-Au were internalized. It is concluded that LDL-Au and HDL-Au conjugates can be used to identify lipoprotein receptors and to follow lipoprotein internalization in adrenocortical cells.

Effects of joining peptide (1-18) and histamine on dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulphate (DHEAS) production of human adrenocortical cells in vitro

Joining peptide 1-18 (JP 1-18), added alone in concentrations of 10(-13)-10(-7) M to collagenase-dispersed human adrenocortical cells, did not affect the basal production of corticosterone, cortisol, aldosterone, dehydroepiandrosterone (DHEA) and dehydroepiandrosterone sulphate (DHEAS). JP 1-18 potentiated the ACTH-stimulated production of steroids. When administered in combination with histamine (10(-8)-10(-3) M), JP 1-18 (10(-8) or 10(-10) M), enhanced the synthesis of DHEA and DHEAS. JP 1-18, together with histamine, may play a role in the regulation of DHEA and DHEAS production.

Effect of chronic ACTH treatment on the physical state of lipid droplets in rat adrenocortical cells

A histophysical method has been adapted to determine the thermotropic phase transitions of adrenocortical lipid droplets using a polarizing microscope equipped with a cold/hot stage. Cryosections of freshly-removed, unfixed adrenals, derived from control (untreated), and 14 days ACTH-treated rats were examined. The lipid droplets in the zona fasciculata and zona reticularis of the untreated rats were birefringent at room temperature (22 degrees C). The birefringence of zona glomerulosa lipids selectively increased in the temperature range from -10 to -15 degrees C. In cryosections prepared from ACTH-treated rats, thermotropic phase transitions of the lipid droplets appeared at a temperature range between -30 and -40 degrees C in each cortical zone. The chemical analysis of the isolated lipids revealed that the relative amount of triglycerides in the zona fasciculata lipids increased, while that of free and esterified cholesterol decreased after chronic ACTH treatment. Present data suggest that the increased fluidity of lipid droplets promotes lipid mobilization in response to the enhanced demand of the chronically stimulated adrenocortical cells. Viscosity-dependent mobilization of free cholesterol from lipid droplets is not a rate-limiting process in adrenal steroidogenesis, but rather may represent an important control of the availability of precursor from lipid stores.