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

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.

scholarly journals Intrarenal signaling mediated by CCK plays a role in salt intake-induced natriuresis

2017 ◽  
Vol 313 (1) ◽  
pp. F20-F29 ◽  
Author(s):  
Hiromi Takahashi-Iwanaga ◽  
Shunsuke Kimura ◽  
Kohtarou Konno ◽  
Masahiko Watanabe ◽  
Toshihiko Iwanaga
Keyword(s):  
In Situ Hybridization ◽  
Sodium Excretion ◽  
Interstitial Cells ◽  
Hybridization Technique ◽  
Renal Tubules ◽  
Outer Medulla ◽  
Natriuretic Hormone ◽  
High Sodium ◽  
Gene Transcripts

The natriuretic hormone CCK exhibits its gene transcripts in total kidney extracts. To test the possibility of CCK acting as an intrarenal mediator of sodium excretion, we examined mouse kidneys by 1) an in situ hybridization technique for CCK mRNA in animals fed a normal- or a high-sodium diet; 2) immuno-electron microscopy for the CCK peptide, 3) an in situ hybridization method and immunohistochemistry for the CCK-specific receptor CCKAR; 4) confocal image analysis of receptor-mediated Ca2+ responses in isolated renal tubules; and 5) metabolic cage experiments for the measurement of urinary sodium excretion in high-salt-fed mice either treated or untreated with the CCKAR antagonist lorglumide. Results showed the CCK gene to be expressed intensely in the inner medulla and moderately in the inner stripe of the outer medulla, with the expression in the latter being enhanced by high sodium intake. Immunoreactivity for the CCK peptide was localized to the rough endoplasmic reticulum of the medullary interstitial cells in corresponding renal regions, confirming it to be a secretory protein. Gene transcripts, protein products, and the functional activity for CCKAR were consistently localized to the late proximal tubule segments (S2 and S3) in the medullary rays, and the outer stripe of the outer medulla. Lorglumide significantly diminished natriuretic responses of mice to a dietary sodium load without altering the glomerular filtration rate. These findings suggest that the medullary interstitial cells respond to body fluid expansion by CCK release for feedback regulation of the late proximal tubular reabsorption.

Localization of CYP2D16 in the guinea pig adrenal cortex by immunohistochemistry and in situ hybridization

1997 ◽  
Vol 134 (2) ◽  
pp. 139-146 ◽  
Author(s):  
Bing-Bing Yuan ◽  
Ruy Tchao ◽  
Jeffrey M Voigt ◽  
Howard D Colby
Keyword(s):  
In Situ Hybridization ◽  
Guinea Pig ◽  
Adrenal Cortex

scholarly journals Myosin functional domains encoded by alternative exons are expressed in specific thoracic muscles of Drosophila.

1991 ◽  
Vol 114 (2) ◽  
pp. 263-276 ◽  
Author(s):  
G A Hastings ◽  
C P Emerson
Keyword(s):  
In Situ Hybridization ◽  
Direct Flight ◽  
Alternatively Spliced ◽  
Gene Transcripts ◽  
Flight Muscles ◽  
Thoracic Muscles ◽  
Muscle Myosin ◽  
Muscle Specificity ◽  
Esophageal Muscle

The Drosophila 36B muscle myosin heavy chain (MHC) gene has five sets of alternatively spliced exons that encode functionally important domains of the MHC protein and provide a combinatorial potential for expression of as many as 480 MHC isoforms. In this study, in situ hybridization analysis has been used to examine the complexity and muscle specificity of MHC isoform expression in the fibrillar indirect flight muscle (IFM), the tubular direct flight muscles (DFM) and tubular tergal depressor of the trochanter muscle (TDT), and the visceral esophageal muscle in the adult thorax. Our results show that alternative splicing of the MHC gene transcripts is precisely regulated in these thoracic muscles, which express three MHC isoforms. Individual thoracic muscles each express transcripts of only one isoform, as detectable by in situ hybridization. An apparently novel fourth MHC isoform, with sequence homology to the rod but not to the head domain of the 36B MHC, is expressed in two direct flight muscles. These findings form a basis for transgenic experiments designed to analyze the muscle-specific functions of MHC domains encoded by alternative exons.

scholarly journals Transcriptional pattern of 21-hydroxylase gene (P-450C21) during embryonic development, before, and after birth in mice as determined by in situ hybridization.

1989 ◽  
Vol 37 (5) ◽  
pp. 751-756 ◽  
Author(s):  
G Raschellà ◽  
G Smets ◽  
A Claeys ◽  
P Verdood ◽  
A Romeo ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Adrenal Cortex ◽  
Steady Increase ◽  
Computer Assisted ◽  
Adult Age ◽  
Stage Of Development ◽  
Transcriptional Pattern ◽  
Before And After ◽  
Embryonic Life

21-Hydroxylase is a member of the P-450 superfamily of genes involved in the biosynthesis of cortisol and aldosterone in the adrenal cortex. Congenital adrenal hyperplasia, a well-characterized disease, originates from a lack of this enzyme. We present in this report an in situ hybridization study aimed at detecting 21-hydroxylase activity during murine development, from mid gestation to adulthood. Our results demonstrate that even during the embryonic period the adrenal cortex is the only major site of transcription of this enzyme, which is detectable beginning at embryonic day 14. In addition, a peculiar topographical pattern of transcriptional activity, characteristic of the stage of differentiation of the gland, could be drawn. Using a computer-assisted method, we were able to quantitate the relative transcription level at each stage of development. A steady increase in the level of transcription was demonstrated throughout embryonic life to birth, with a drop during the prepubertal period and a final rise at adult age. The possible physiological significance of our findings is discussed.

scholarly journals Lymphocyte Subsets in the Adrenal Glands of Dogs With Primary Hypoadrenocorticism

2016 ◽  
Vol 55 (1) ◽  
pp. 177-181 ◽  
Author(s):  
S. G. Friedenberg ◽  
D. L. Brown ◽  
K. M. Meurs ◽  
J. McHugh Law
Keyword(s):  
T Cells ◽  
In Situ Hybridization ◽  
T Cell ◽  
Adrenal Cortex ◽  
Genetic Polymorphisms ◽  
Adrenal Glands ◽  
Lymphocyte Subsets ◽  
Clinical Syndrome ◽  
Autoimmune Condition

Primary hypoadrenocorticism, or Addison’s disease, is an autoimmune condition common in certain dog breeds that leads to the destruction of the adrenal cortex and a clinical syndrome involving anorexia, gastrointestinal upset, and electrolyte imbalances. Previous studies have demonstrated that this destruction is strongly associated with lymphocytic-plasmacytic inflammation and that the lymphocytes are primarily T cells. In this study, we used both immunohistochemistry and in situ hybridization to characterize the T-cell subtypes involved. We collected postmortem specimens of 5 dogs with primary hypoadrenocorticism and 2 control dogs and, using the aforementioned techniques, showed that the lymphocytes are primarily CD4+ rather than CD8+. These findings have important implications for improving our understanding of the pathogenesis and in searching for the underlying causative genetic polymorphisms.

scholarly journals Saponin pre-treatment in pre-embedding electron microscopic in situ hybridization for detection of specific RNA sequences in cultured cells: a methodological study.

1995 ◽  
Vol 43 (10) ◽  
pp. 1005-1018 ◽  
Author(s):  
M V Macville ◽  
K C Wiesmeijer ◽  
R W Dirks ◽  
J A Fransen ◽  
A K Raap
Keyword(s):  
In Situ Hybridization ◽  
Cultured Cells ◽  
Elongation Factor ◽  
Housekeeping Gene ◽  
28S Rrna ◽  
Electron Microscopic ◽  
Rna Sequences ◽  
Gene Transcripts ◽  
Pre Treatment

We describe a method for detection of specific RNA targets in cultured cells at the electron microscopic (EM) level using pre-embedding in situ hybridization (ISH). The specimens were monitored by reflection-contrast microscopy (RCM) before processing for EM. A good balance between preservation of ultrastructure and intensity of hybridization signals was obtained by using mild aldehyde fixation followed by saponin permeabilization. Digoxigenin-labeled probes were used for detection of human elongation factor (HEF) mRNA in HeLa cells, immediate early (IE) mRNA in rat 9G cells, and 28S rRNA in both cell lines. The hybrids were detected immunocytochemically by the peroxidase/diaminobenzidine (DAB) method or by ultra-small gold with silver enhancement. Comparison of these methods favored the peroxidase/DAB system. The accessibility of RNA in the different cell compartments was dependent on the extent of cross-linking during primary fixation even after permeabilization with saponin. By using the most optimal ISH protocol and the peroxidase/DAB system, we detected 28S rRNA over all ribosomes in the cytoplasm but not in the nucleoli, and IE mRNA in a large spot with many smaller spots around it in the nucleoplasm as well as in speckles over the cytoplasm. The sensitivity of the method is such that HEF housekeeping gene transcripts were detected in the cytoplasm.

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