scholarly journals Development of Adrenal Zonation in Fetal Rats Defined by Expression of Aldosterone Synthase and 11β-Hydroxylase

Endocrinology ◽  
1998 ◽  
Vol 139 (10) ◽  
pp. 4397-4403 ◽  
Author(s):  
Cheryl Wotus ◽  
Brett K. Levay-Young ◽  
Lisa M. Rogers ◽  
Celso E. Gomez-Sanchez ◽  
William C. Engeland
Keyword(s):  
In Situ Hybridization ◽  
Adrenal Cortex ◽  
Messenger Rna ◽  
Zona Glomerulosa ◽  
Immunofluorescent Staining ◽  
Cell Phenotype ◽  
Zona Fasciculata ◽  
Protein Distribution ◽  
Aldosterone Synthase

Abstract The adult rat adrenal cortex is comprised of three concentric steroidogenic zones that are morphologically and functionally distinguishable: the zona glomerulosa, zona intermedia, and the zona fasciculata/reticularis. Expression of the zone-specific steroidogenic enzymes, cytochrome P450 aldosterone synthase (P450aldo), and P450 11β hydroxylase (P45011β), produced by the zona glomerulosa and zona fasciculata/reticularis, respectively, can be used to define the adrenal cortical cell phenotype of these two zones. In this study, immunohistochemistry and in situ hybridization were used to determine the ontogeny of expression of P450aldo and P45011β to monitor the pattern of development of the rat adrenal cortex. RIA was used to measure adrenal content of aldosterone and corticosterone, the resulting products of the two enzymatic pathways. Double immunofluorescent staining for both enzymes at gestational day 16 (E16) showed P45011β protein expressed in cells distributed throughout most of the adrenal intermixed with a separate, but smaller, population of cells expressing P450aldo protein. Whereas expression of P45011β protein retained a similar pattern of distribution from E16 to adulthood (ignoring distribution of SA-1 positive, presumptive medullary cells), P450aldo protein changed its pattern of distribution by E19, becoming localized in a discontinuous ring of cells adjacent to the capsule. By postnatal day 1, P450aldo protein distribution was similar to that observed in adult glands; P450aldo-positive cells formed a continuous zone underlying the capsule. In situ hybridization showed that the pattern of P45011β messenger RNA expression paralleled protein expression at all times, whereas P450aldo messenger RNA paralleled protein at E19 and after, but was undetectable before E19. However, adrenal aldosterone and corticosterone, as measured by RIA, were detected by E16, supporting the functional capacity of both phenotypes for all ages studied. These data suggest that the development of the adrenal zona glomerulosa occurs in two distinct phases; initial expression of the glomerulosa phenotype in scattered cells of the inner cortex before E17, followed by a change in distribution to the outer cortex between E17 and E19. It is hypothesized that this change in distribution occurs via cell differentiation, rather than cell migration, and that a possible regulator of these events is the fetal renin-angiotensin system.

Localization of the gene transcripts of 11 ?-hydroxylase and aldosterone synthase in the rat adrenal cortex by in situ hybridization

1991 ◽  
Vol 96 (5) ◽  
pp. 391-394 ◽  
Author(s):  
M. Yabu ◽  
T. Senda ◽  
Y. Nonaka ◽  
N. Matsukawa ◽  
M. Okamoto ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Adrenal Cortex ◽  
Aldosterone Synthase ◽  
Gene Transcripts

Endocrine control of the distribution of basic fibroblast growth factor, insulin-like growth factor-I and transforming growth factor-β1 mRNAs in adult rat adrenals using non-radioactive in situ hybridization

1995 ◽  
Vol 144 (2) ◽  
pp. 379-387 ◽  
Author(s):  
M M Ho ◽  
G P Vinson
Keyword(s):  
Growth Factor ◽  
Adrenal Cortex ◽  
Transforming Growth Factor ◽  
Zona Glomerulosa ◽  
Dietary Sodium ◽  
Zona Fasciculata ◽  
Sodium Restriction ◽  
Igf I ◽  
Rat Adrenals

Abstract This study located the particular cell types involved in the synthesis of growth factors in adult female rat adrenal glands. Non-isotopic in situ hybridization was used and the cellular localizations of the mRNAs of basic fibroblast growth factor (bFGF), IGF-I, and transforming growth factor-β1 (TGF-β1) were studied in adrenals from control animals and from those treated with ACTH or subjected to dietary sodium restriction. The adrenal medulla was the richest source of both bFGF and IGF-1 mRNA in both control and experimental rat adrenals. In the cortex, bFGF and IGF-I mRNAs were found mainly in the zona fasciculata in control animals, although some transcription was also detected in the zona reticularis and zona glomerulosa. Both ACTH and sodium restriction activated bFGF and IGF-I gene expression in the zona glomerulosa. Since cellular proliferation and differentiation occur primarily in the outer cortex, the data are consistent with the view that bFGF and IGF-I act as an autocrine/paracrine mitogen and differentiation regulator respectively in the rat adrenal cortex. Very small amounts of TGF-β1 mRNA were detected, predominantly in the zona fasciculata of control rats. There were no observable differences in amounts and localization of TGF-β1 mRNA between the adrenals of control rats and those treated with ACTH for 1 day. TGF-β1 mRNA was very weak or undetectable in the adrenals from rats treated with ACTH for three and five days or from sodium-restricted rats. Although TGF-β1 immunoreactive protein has been shown to be present in the zonae fasciculata and reticularis and to modulate negatively the steroidogenic activities in the adrenal cortex of other species, its gene is not actively expressed in rat adrenals. The present results showed that ACTH administration or dietary sodium restriction, both important adrenal mitogens in vivo, significantly altered the spatial patterns of the distribution of bFGF and IGF-I mRNAs and also increased the amount of bFGF mRNA in the adrenal cortex. This suggests that growth and differentiation of the adrenal cortex are partly mediated by bFGF and IGF-I. Journal of Endocrinology (1995) 144, 379–387

Changes in the glomerulosa cell phenotype during adrenal regeneration in rats

1999 ◽  
Vol 276 (5) ◽  
pp. R1374-R1382 ◽  
Author(s):  
W. C. Engeland ◽  
B. K. Levay-Young
Keyword(s):  
Cellular Differentiation ◽  
Zona Glomerulosa ◽  
Cell Phenotype ◽  
Zona Fasciculata ◽  
Elevated Plasma ◽  
Plasma Acth ◽  
Acth Treatment ◽  
Glomerulosa Cells ◽  
Cytochrome P 450

In situ hybridization was used to examine cellular differentiation during rat adrenal regeneration, defining zona glomerulosa [cytochrome P-450 aldosterone synthase ( P-450aldo) mRNA positive], zona fasciculata [cytochrome P-450 11β-hydroxylase ( P-45011β) mRNA positive], or zona intermedia [negative for both but 3β-hydroxysteroid dehydrogenase (3β-HSD) mRNA positive]. After unilateral adrenal enucleation with contralateral adrenalectomy (ULE/ULA), the expression of all mRNA was reduced at 2 days. From 5 to 10 days, P-45011β and 3β-HSD mRNA increased while P-450aldo remained low; at 20 days, all mRNA were increased. From 2 to 10 days, cells adjacent to the capsule showed intermedia cell differentiation; by 20 days, the subcapsular glomerulosa cells reappeared. This suggests that after enucleation the glomerulosa dedifferentiates to zona intermedia. The experiment was repeated in rats where the postenucleation ACTH rise was prevented. Rats underwent ULE with sham ULA (ULE/SULA) or ULE/SULA with ACTH treatment. Adrenals from ULE/SULA rats expressed increased P-450aldo mRNA at 10 days and reduced P-45011β mRNA and adrenal weight at 30 days. ACTH treatment reversed the pattern toward that seen in ULE/ULA. These findings show that the enucleation-induced dedifferentitation of the glomerulosa cell may result in part from elevated plasma ACTH and that prevention of dedifferentiation may result in impaired regeneration.

11β-Hydroxylase gene expression in the rat adrenal cortex

1993 ◽  
Vol 139 (2) ◽  
pp. 301-NP ◽  
Author(s):  
M. M. Ho ◽  
G. P. Vinson
Keyword(s):  
Adrenal Cortex ◽  
Zona Glomerulosa ◽  
Dietary Sodium ◽  
Zona Fasciculata ◽  
Hybridization Technique ◽  
Sodium Restriction ◽  
Zonal Distribution ◽  
Aldosterone Synthase ◽  
Acth Treatment ◽  
Dietary Sodium Restriction

ABSTRACT It is now known that in the rat there are two distinct species of cytochrome P45011β/18, namely aldosterone synthase and 11β-hydroxylase. Whereas aldosterone synthase is located exclusively in the zona glomerulosa, the zonal distribution and site of production of 11 β-hydroxylase is not entirely clear. In the present study we examined the zonal expression of 11β-hydroxylase mRNA in adrenals from control rats and animals subjected to ACTH treatment and dietary sodium restriction using a non-isotopic in-situ hybridization technique. The results were compared with those obtained using an inner zone specific antigenic (IZAg) marker to give unequivocal identification of the adrenocortical cell types. 11 β-Hydroxylase mRNA was clearly shown to be expressed in the inner zones of the control rat adrenal cortex, and none was detected in the zona glomerulosa and medulla. The message was more abundant in the outer zona fasciculata. A similar pattern of distribution of 11β-hydroxylase mRNA was observed in adrenals from rats subjected to dietary sodium restriction, although the width of the negatively staining layer of zona glomerulosa was significantly increased. In rats treated with 100 μg ACTH for 1 day, the number of cells expressing 11β-hydroxylase mRNA was increased, especially in the zona reticularis. With continued ACTH treatment, 11β-hydroxylase mRNA was found in the region of the zona glomerulosa, and after 3 and 5 days of ACTH treatment cells expressing 11β-hydroxylase mRNA extended to the connective tissue capsule. At this time there was a significant reduction in the total expression of the message compared with the controls. These results suggest that the presence of 11β-hydroxylase in the zona glomerulosa cells is not essential for the late pathway for aldosterone biosynthesis from deoxycorticosterone. Like IZAg, the distribution of 11β-hydroxylase mRNA after prolonged ACTH treatment provides further evidence to support the hypothesis that ACTH increases the conversion of zona glomerulosa to zona fasciculata-like cells. Journal of Endocrinology (1993) 139, 301–306

scholarly journals Ultrastructural visualization of cytoskeletal mRNAs and their associated proteins using double-label in situ hybridization.

1989 ◽  
Vol 108 (6) ◽  
pp. 2343-2353 ◽  
Author(s):  
R H Singer ◽  
G L Langevin ◽  
J B Lawrence
Keyword(s):  
In Situ Hybridization ◽  
High Resolution ◽  
Colloidal Gold ◽  
Messenger Rna ◽  
Signal To Noise Ratio ◽  
Simultaneous Detection ◽  
Gold Particles ◽  
Rna Molecules ◽  
Nucleic Acid Molecule

We have been able to visualize cytoskeletal messenger RNA molecules at high resolution using nonisotopic in situ hybridization followed by whole-mount electron microscopy. Biotinated cDNA probes for actin, tubulin, or vimentin mRNAs were hybridized to Triton-extracted chicken embryo fibroblasts and myoblasts. The cells were then exposed to antibodies against biotin followed by colloidal gold-conjugated antibodies and then critical-point dried. Identification of mRNA was possible using a probe fragmented to small sizes such that hybridization of several probe fragments along the mRNA was detected as a string of colloidal gold particles qualitatively and quantitatively distinguishable from nonspecific background. Extensive analysis showed that when eight gold particles were seen in this iterated array, the signal to noise ratio was greater than 30:1. Furthermore, these gold particles were colinear, often spiral, or circular suggesting detection of a single nucleic acid molecule. Antibodies against actin, vimentin, or tubulin proteins were used after in situ hybridization, allowing simultaneous detection of the protein and its cognate message on the same sample. This revealed that cytoskeletal mRNAs are likely to be extremely close to actin protein (5 nm or less) and unlikely to be within 20 nm of vimentin or tubulin filaments. Actin mRNA was found to be more predominant in lamellipodia of motile cells, confirming previous results. These results indicate that this high resolution in situ hybridization approach is a powerful tool by which to investigate the association of mRNA with the cytoskeleton.

scholarly journals Human Adrenocortical Remodeling Leading to Aldosterone-Producing Cell Cluster Generation

2016 ◽  
Vol 2016 ◽  
pp. 1-6 ◽  
Author(s):  
Koshiro Nishimoto ◽  
Tsugio Seki ◽  
Yuichiro Hayashi ◽  
Shuji Mikami ◽  
Ghaith Al-Eyd ◽  
...  
Keyword(s):  
Adrenal Cortex ◽  
Zona Glomerulosa ◽  
Statistical Analyses ◽  
Cell Cluster ◽  
Zona Fasciculata ◽  
Undifferentiated Cell ◽  
Immunohistochemical Detection ◽  
Cluster Generation ◽  
Over Time ◽  
Cell Zone

Background. The immunohistochemical detection of aldosterone synthase (CYP11B2) and steroid 11β-hydroxylase (CYP11B1) has enabled the identification of aldosterone-producing cell clusters (APCCs) in the subcapsular portion of the human adult adrenal cortex. We hypothesized that adrenals have layered zonation in early postnatal stages and are remodeled to possess APCCs over time.Purposes. To investigate changes in human adrenocortical zonation with age.Methods. We retrospectively analyzed adrenal tissues prepared from 33 autopsied patients aged between 0 and 50 years. They were immunostained for CYP11B2 and CYP11B1. The percentage of APCC areas over the whole adrenal area (AA/WAA, %) and the number of APCCs (NOA, APCCs/mm2) were calculated by four examiners. Average values were used in statistical analyses.Results. Adrenals under 11 years old had layered zona glomerulosa (ZG) and zona fasciculata (ZF) without apparent APCCs. Some adrenals had an unstained (CYP11B2/CYP11B1-negative) layer between ZG and ZF, resembling the rat undifferentiated cell zone. Average AA/WAA and NOA correlated with age, suggesting that APCC development is associated with aging. Possible APCC-to-APA transitional lesions were incidentally identified in two adult adrenals.Conclusions. The adrenal cortex with layered zonation remodels to possess APCCs over time. APCC generation may be associated with hypertension in adults.

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