Adrenal renin: a possible local regulator of aldosterone production

Extrarenal renin has been identified in a number of tissues, including the brain, the submaxillary gland, uterus, ovary, vascular endothelium, testes, pituitary gland, and the adrenal cortex. In some tissues, including the adrenal cortex, all of the components of the renin-angiotensin system have been identified; however, no specific physiologic role has been clearly demonstrated for these extrarenal renin-angiotensin systems. We have studied the role of the renin-angiotensin system in the adrenal cortex of the rat and have found that renin is localized and synthesized in the zona glomerulosa cells. Its production can be influenced by alterations in electrolyte balance, as well as the genetic background of the rat. In adrenal capsular explant cultures, a converting enzyme inhibitor can lower angiotensin II production and reduce the stimulation of aldosterone by potassium, suggesting that this system is involved in the aldosterone response to potassium. In addition to rat adrenals, renin has been identified in human adrenal tissue and human adrenal tumors, including aldosteronomas, and a patient with hypertension has been reported to have an adrenal tumor that appeared to be secreting renin into the circulation.     

Phospholipid composition of human, dog and guinea pig adrenal cortex

The phospholipid composition was studied in human and dog adrenal cortex and in guinea pig adrenal tissue. The major phospholipids of adrenal cortex were phosphatidylcholine and phosphatidylethanolamine whose ratio in the human, dog and guinea pig tissues was 2.16, 2.01, 1.61, respectively. Phosphatidylinositol, phosphatidylserine, sphingomyelin, diphosphatidylglycerin, lysophosphatidylcholine and lysophosphatidyl-ethanolamine were also found in adrenal cortex. A quantitative phospholipid composition of the human adrenal cortex was close to the dog one. The fatty acid composition of phosphatidylcholine from human and dog adrenal cortex was determined and some differences were shown.     

牛肾上腺皮质LDL受体的纯化和抗体的制备

牛肾上腺皮质LDL受体经Triton X-100增溶,DEAE_(32)离子交换柱和LpB Sepharose亲和柱层析,在SDS-PAGE中有三条区带,分别在原点;Mr 160kD;Mr125kD处。进一步用8％SDS-PAGE纯化三个区带的蛋白质分别免疫新西兰大白兔所得的抗体,应用免疫印迹和ECL非同位素标记法可对牛肾上腺皮质和人皮肤纤维细胞膜上的LDL受体进行测定。     

肾上腺皮质束状带特异性表达Cre重组酶转基因小鼠的构建

肾上腺是人体重要的内分泌器官。由于缺乏肾上腺皮质束状带特异性表达Cre酶的工具鼠,目前对肾上腺皮质束状带细胞中特异表达基因的功能缺乏深入的解析。CYP11B1基因编码类固醇11β-羟化酶,该酶是糖皮质激素合成的关键酶,在肾上腺皮质束状带中特异性表达。本研究旨在利用CYP11B1基因在束状带特异性表达的特点,构建在肾上腺皮质束状带中特异性表达Cre重组酶的转基因动物。采用CRISPR/Cas9技术在CYP11B1基因终止密码子位点定点敲入2A-GfpCre表达框,获得CYP11B1-2A-GfpCre同源重组载体,进而构建CYP11B1Cre小鼠,并通过mTmG和LacZ染色确定Cre酶主要表达在小鼠肾上腺皮质束状带。在此基础上,本研究还用该工具鼠与胱硫醚-γ-裂解酶(cystathionineγ-lyase, CTH)条件性敲除鼠交配,获得了肾上腺皮质束状带CTH特异性敲除的小鼠,并证实了该动物肾上腺皮质束状带中CTH表达缺失。以上结果充分说明肾上腺皮质束状带特异性表达Cre重组酶小鼠构建成功。该工具鼠的成功构建,为深入研究肾上腺皮质束状带相关功能提供了有力工具。     

Nestin expression in normal adrenal gland and adrenocortical tumors

Human adrenocortical cells have been shown to express cytokeratins and vimentin. Nestin is an intermediate filament protein that is mainly expressed in the developing nervous system and that has been recently reported in rat adrenal gland as well. Using immunohistochemical and biochemical approaches, the present study demonstrates that nestin is constantly expressed in situ in the cortex of normal human adrenal glands. Nestin expressing cells were prevalently located in the zona reticularis but some positive cells could be spotted in the zona fasciculata as well. Moreover, patches of nestin-positive cells have been constantly detected on sections of cortical adenomas. In contrast, adrenal carcinomas displayed a variable number of nestin-immunoreactive cells that in some cases were virtually absent. Samples of renal clear cell carcinoma metastasis in the adrenals were also examined which did not show nestin-immunoreactivity. We propose that a positive nestin-immunoreaction could be useful in differential diagnosis of clear cell tumors in adrenal glands.     

Effect of ACTH and prolactin on dehydroepiandrosterone, its sulfate ester and cortisol production by normal and tumorous human adrenocortical cells

The effect of ACTH and prolactin on the synthesis of dehydroepiandrosterone (DHEA) and its sulfate ester (DHEAS) was studied in cell suspensions of "normal" and tumorous (adenoma) human adrenal cortex. A stimulation of DHEA and no response of DHEAS production by ACTH in "normal" adrenocortical cell suspension was observed. However ACTH stimulated both DHEA and DHEAS synthesis in tumorous adrenocortical cells. Prolactin did not influence either the basal or the ACTH stimulated DHEA and DHEAS production of adrenocortical cells irrespective of their origin. Our results are compatible with the concept that the biosynthesis of DHEA is under ACTH control, while other factor(s) regulate(s) the sulfate pathway of DHEA secretion under normal conditions. In tumorous adrenocortical cells DHEA may be regulated--at least partly--by ACTH. Prolactin seems to have no direct effect on DHEA and DHEAS synthesis. It is postulated that the relationship between serum prolactin and DHEAS (or DHEA) levels observed by several authors might be an extraadrenal effect of prolactin on adrenal androgens.     

Morphofunctional features of the secretory cells of the adenohypophysis and adrenal glands of the rat during aging

Karyometric and electron microscopic investigation has been performed in the adenohypophyseal secretory cells, in the cortex and medulla of the adrenal glands in mature (6 months) and ageing (25-26 months) white male rats. The diameter of the corticotropic cell nuclei of the adenohypophysis significantly increases with age. In the secretory cells of the adrenal cortex and medulla no important changes in their parameters are revealed. With age destructive changes of the ultrastructures are manifested variously in different types of the secretory cells in the endocrinic glands studied. In the corticotropic, somatotropic, thyreotropic cells of the adenohypophysis, in spongiocytes of the adrenal cortex, compensatory ultrastructural rearrangements develop; they are aimed to preserve functional activity of the cells mentioned. No age changes are revealed in ultrastructure of chromaffin cells of the adrenal medulla.     

The Involvement of Nek2 and Notch in the Proliferation of Rat Adrenal Cortex Triggered by POMC-Derived Peptides

The adrenal gland is a dynamic organ that undergoes constant cell turnover. This allows for rapid organ remodeling in response to the physiological demands of the HPA axis, which is controlled by proopiomelanocortin (POMC)-derived peptides, such as adrenocorticotropic hormone (ACTH) and N-Terminal peptides (N-POMC). In the rat adrenal cortex, POMC-derived peptides trigger a mitogenic effect, and this process increases cyclins D and E, while inhibiting p27Kip1. The goal of the present study was to further explore the mitogenic effect of ACTH and synthetic N-POMC_1–28 peptides by investigating the differences in the expression of key genes involved in the cell cycle of the rat adrenal cortex, following inhibition of the HPA axis. Moreover, we evaluated the differences between the inner and outer fractions of the adrenal cortex (ZF-fraction and ZG-fraction) in terms of their response patterns to different stimuli. In the current study, the inhibition of the HPA axis repressed the expression of Ccnb2, Camk2a, and Nek2 genes throughout the adrenal cortex, while treatments with POMC-derived peptides stimulated Nek2, gene and protein expression, and Notch2 gene expression. Furthermore, Notch1 protein expression was restricted to the subcapsular region of the cortex, an area of the adrenal cortex that is well-known for proliferation. We also showed that different regions of the adrenal cortex respond to HPA-axis inhibition and to induction with POMC-derived peptides at different times. These results suggest that cells in the ZG and ZF fractions could be at different phases of the cell cycle. Our results contribute to the understanding of the mechanisms involved in cell cycle regulation in adrenocortical cells triggered by N-POMC peptides and ACTH, and highlight the involvement of genes such as Nek2 and Notch.     

Morphometric study of the postnatal development of the human adrenal cortex]

Under study was the developmental dynamics of a number of morphometrical parameters of the adrenal cortex of man. The number of glandular cells in the organ was found to considerably decrease at late developmental periods, the size of nuclei in them diminished. These factors are considered to be responsible for the drop of functional activity of the adrenal cortex in elderly and senile age. The increase of the cell size during the growing of the organism is thought to be an important factor of the growth of the adrenal cortex of man.     

Toll-like receptor 2 and Toll-like receptor 4 expression in human adrenals.

Toll-like receptors (TLRs) are key elements in the innate immune response, functioning as pattern-recognition receptors for the detection and response to endotoxins and other microbial ligands. Inflammatory cytokines play an important role in the activation of the hypothalamic-pituitary-adrenal HPA axis during inflammation and sepsis. The newly recognized major role of TLR2 and TLR4 and the adrenal stress response during critical illnesses such as inflammation and sepsis demand comprehensive analysis of their interactions. Therefore, we analyzed TLR2 and TLR4 expression in human adrenal glands. Western blot analysis demonstrated the expression of TLR2 and TLR4 in the human adrenocortical cell line NCI-H295. Immunohistochemical analysis of normal human adrenal glands revealed TLR2 and TLR4 expression in the adrenal cortex, but not in the adrenal medulla. Considering the crucial role of the HPA axis and the innate immune response during acute sepsis or septic shock, elucidating the functional interaction of these systems should be of great clinical relevance.     

Stress-protective effect of the synthetic ACTH-like peptide leucocorticotropin

We found that the tritium-labeled synthetic ACTH-like octapeptide leucocorticotropin corresponding to the 81-88 sequence of the precursor of human interleukin-1alpha ([3H]GKVLKKRR) is bound by the ACTH receptor of rat adrenal cortex with a high affinity and specificity (Kd 2.2 +/- 0.1 nM). This peptide was shown to exert no effect on the adenylate cyclase activity of the membranes of rat adrenal cortex in the concentration range from 1 to 1000 nM. Leucocorticotropin administration three times at doses of 10-20 microg/animal did not change the level of hydroxycorticosteroids (11-HOCS) in the rat adrenal glands in the absence of temperature action. At the same time, the peptide abolishes (at a dose of 20 microg/animal, three times) or significantly decreases (at a dose of 10 microg/animal, three times) the dramatic increase in the 11-HOCS content in the adrenal glands occurring in the case of cold or heat shock. Thus, leucocorticotropin normalizes the 11-HOCS level in the rat adrenal cortex during stress. The stress-protective effect of the peptide is mediated through the ACTH receptor.     

Characterization of the porcine ACTH receptor with the aid of a monoclonal antibody

Monoclonal antibodies were raised against the ACTH receptor by immunization of BALB1 c mice with porcine adrenal cortex cell membranes. Competition and binding experiments confirmed that one of these antibodies, IV/14/9, reacted with the ACTH receptor in competition with ACTH and caused a definite ACTH-like effect. This antibody was used to characterize the hormone receptor of the porcine adrenal cortex. The number of antibody binding sites per cell was calculated from a Scatchard plot to be 124,100 +/- 10,000. The curve was linear indicating the existence of a single class of receptors. The finding that a high concentration of IV/14/9 totally suppressed maximally ACTH-induced steroidogenesis confirms the view that only a single class of ACTH receptors exists. The antibody IV/14/9 neither reacted with intact porcine thymus cells nor with normal porcine lymphocytes but was bound to the mouse adrenal tumor cell line Y1 and to normal rat adrenal cortex cells with a low affinity. Two dimensional electrophoresis of lysates of porcine adrenal cortex cells and subsequent blotting of the proteins on nitrocellulose, using IV/14/9 as a primary and anti-mouse Ig as a secondary reagent, permitted the detection of three forms of the receptor, two of which had an identical apparent molecular mass of about 85,000 Da (isoelectric points: 6.14 and 6.27). The third was somewhat larger (94,000 Da and pI = 6.21).     

ACTH and thyroid hormone regulation of 3 beta-hydroxysteroid dehydrogenase activity in human fetal adrenocortical cells

The regulation of 3 beta-hydroxysteroid dehydrogenase, delta 4,5-isomerase (3 beta-HSD) and hydroxysteroid sulfotransferase (HST) activities by ACTH and thyroid hormones was investigated in cell cultures from the fetal zone or definitive zone of the human fetal adrenal cortex, using a serum-free, defined medium. ACTH alone maximally stimulated the 3 beta-HSD activity several-fold, whereas triiodothyronine (T3) alone had no effect on this enzyme activity in cell cultures from each zone. However, treatment of cultures with maximal concentrations of 10 nM ACTH plus 1 nM T3 significantly increased the 3 beta-HSD activity an additional 59-115% over that for ACTH alone, without alteration of the ACTH ED50 (0.3 nM). The T3 ED50 was 31 pM for this interaction with ACTH. Thyroxine at a reduced sensitivity had the same interaction with ACTH. T3 similarly increased the stimulation of 3 beta-HSD activity by the steroidogenic agents, cholera toxin and a cAMP analog. The HST activity was not affected by T3 alone but was stimulated by ACTH alone. This stimulation was an order of magnitude less than that for the 3 beta-HSD activity in the same cultures. ACTH plus T3 did not have the synergistic effect on HST activity as observed for the 3 beta-HSD activity. These studies show an interaction between ACTH and thyroid hormone for the stimulation of 3 beta-HSD activity in cell cultures of the human fetal adrenal cortex.     

Effect of o,p'-DDD and Li+ on apoptotic DNA fragmentation in conventionally normal and tumour tissues of human adrenal cortex

The actions of 1-(o-chlorophenyl)-1-(p-chlorophenyl)-2,2-dichloroethane (o,p'-DDD), potassium and lithium ions upon apoptotic processes in conventionally normal and tumour tissues of human adrenal cortex were studied. There was no effect of K+ on the apoptosis in tumour tissue. o,p'-DDD--the specific drug for conservative therapy of adrenocortical cancer--enhanced the apoptotic DNA fragmentation in all tested tissues. The conclusion was made that apoptosis may be involved in curative effect of o,p'-DDD in adrenal cortex. Lithium ions, which are used in clinic as antidepressant, inhibited the apoptosis in conventionally normal tissue and in most tumours. On the other hand, lithium enhanced the DNA fragmentation in the postoperative tissue of patients with Cushing disease. The possible mechanisms mediating lithium effects on the adrenal cortex are discussed.     

Formaldehyde-induced appearance of septate junctions between digestive vacuoles

Tissue biopsies from (1) some chronic inflammatory diseases, (2) a necrotic tumoral process, (3) normal human lymphatic ganglia, and (4) two congenital diseases of the adrenal cortex were selected for study. A block from each biopsy was fixed in glutaraldehyde-paraformaldehyde; a second block was fixed in 10% formaldehyde. In all cases septate junctions between digestive vacuoles did occur in phagocytic cells and some adrenal cortex cells fixed in formaldehyde. These junctions were similar to those reported recently for malakoplakia phagocytes. Consistently, they were not found to attach organelles other than lysosomes derivatives. Both phagocytes and adrenal cortex cells in the material fixed in glutaraldehyde-paraformaldehyde did not display adhesive specializations between digestive vacuoles. This suggests that the septate junctions described herein are artifactuous structures induced by formaldehyde. There is, however, a certain degree of specificity of cells having the capability of developing these septate junctions. It is assumed that the coating material of digestive organelles in phogocytes and some other cells would be responsible for both cell specificity and organelle specificity of the formaldehyde-induced septate junctions.