MOLECULAR EVIDENCE FOR THE PRESENCE OF GROWTH HORMONE RECEPTORS IN THE BOVINE MAMMARY GLAND

1990 ◽  
Vol 126 (3) ◽  
pp. R5-R8 ◽  
Author(s):  
D.R. Glimm ◽  
V.E. Baracos ◽  
J.J. Kennelly
Keyword(s):  
In Situ Hybridization ◽  
Mammary Gland ◽  
Messenger Rna ◽  
Alveolar Epithelial Cells ◽  
Mammary Tissue ◽  
Molecular Evidence ◽  
Target Tissue ◽  
Bovine Mammary Gland ◽  
Gh Receptor

ABSTRACT GH receptor messenger RNA (mRNA) was identified and characterized in mammary tissue from normal and GH-treated lactating cows using Northern and in-situ hybridization analyses. One major GH receptor transcript of 4.4 kilobases and a less abundant transcript of 9.2 kilobases were detected in mammary tissue from both normal and GH-treated cows. In-situ hybridization analysis revealed that the GH receptor gene is primarily expressed in the alveolar epithelial cells of mammary tissue. These results are evidence that the lactating mammary gland may synthesize GH receptors. On the basis of these observations it seems likely that the lactating bovine mammary gland is a GH target tissue. This finding challenges the widely accepted view that GH does not directly regulate mammary growth or function.

scholarly journals Cellular localisation of GH receptor in the bovine mammary gland during mammogenesis, lactation and involution

2000 ◽  
Vol 166 (3) ◽  
pp. 503-510 ◽  
Author(s):  
F Sinowatz ◽  
D Schams ◽  
S Kolle ◽  
A Plath ◽  
D Lincoln ◽  
...  
Keyword(s):  
Mammary Gland ◽  
Post Partum ◽  
In Situ Hybridisation ◽  
Alveolar Epithelium ◽  
Mammary Tissue ◽  
Bovine Mammary Gland ◽  
Direct Role ◽  
Gh Receptor ◽  
Gh Receptors

We have used immunohistochemistry and non-radioactive in situ hybridisation to localise the GH receptor and its transcript in the bovine mammary gland during mammogenesis, lactation and involution. We found a characteristic pattern of immunoreactive GH (irGH) receptor distribution in the epithelial and stromal compartments during the different stages of mammary gland development: The ductular epithelium showed a distinct staining for irGH receptor during most stages, whereas the alveolar epithelium contained a modest amount of GH receptor during pregnancy which increased during lactation and galactopoiesis. In dry cows, the immunostaining for GH receptors in the alveolar epithelium was very weak or negative. Curiously, the amount of GH receptor mRNA appeared relatively constant during mammogenesis and lactation. The epithelial cells of the alveoli and ducts as well as the endothelial cells showed a distinct signal in our in situ hy! bridisation studies. The predominant localisation of GH receptors in the epithelium of ducts and alveoli is supportive of a role for GH in epithelial differentiation and maintenance. Furthermore, the increased intensity of immunostaining in bovine mammary tissue post partum suggests a direct role for GH receptor in mediating the effect of GH in milk production and secretion.

scholarly journals Developmental regulation of mammary-derived growth inhibitor expression in bovine mammary tissue.

1990 ◽  
Vol 110 (5) ◽  
pp. 1779-1789 ◽  
Author(s):  
A Kurtz ◽  
F Vogel ◽  
K Funa ◽  
C H Heldin ◽  
R Grosse
Keyword(s):  
In Situ Hybridization ◽  
Mammary Gland ◽  
Epithelial Cells ◽  
Growth Inhibitor ◽  
Mammary Tissue ◽  
Hybridization Technique ◽  
Bovine Mammary Gland ◽  
Experimental Pharmacology ◽  
Plasmid Library

The cDNA for a previously described growth inhibitor, designated as mammary-derived growth inhibitor (MDGI) (Grosse, R., and P. Langen. 1989. In Handbook of Experimental Pharmacology. In press) has been cloned from a plasmid library which was derived from terminally differentiated bovine mammary gland. Sequencing of the cDNA showed an open reading frame coding for a protein of 133 amino acids. In six positions differences were found between the sequence determined from the cDNA and that determined previously by amino acid sequence analysis. Northern blot analysis revealed abundant MDGI mRNA in the terminally differentiated mammary gland, whereas in virgin gland, liver or pancreas transcripts were not expressed. By use of in situ hybridization technique transcription of MDGI in the developing bovine mammary gland was analyzed. Increasing amounts of MDGI mRNA were detected in the epithelial cells of embryonic mammary rudiment, in the epithelium of developing lobules and in terminal parts of ducts and lobuloalveolar epithelial cells of differentiated glands. There was a geographical gradient of MDGI mRNA concentration in bovine mammary gland reaching a maximum in the proximal parts of the tissue. An immunohistochemical analysis with different polyclonal and peptide directed antibodies against MDGI confirmed the in situ hybridization data with respect to the tissue-specific and differentiation-dependent MDGI expression in bovine mammary gland. The results suggest a close relationship between MDGI transcription and developmental processes in the normal bovine mammary gland.

Transcription of prolactin gene in milk secretory cells of the rat mammary gland

1993 ◽  
Vol 136 (2) ◽  
pp. 271-NP ◽  
Author(s):  
R. W. Steinmetz ◽  
A. L. Grant ◽  
P. V. Malven
Keyword(s):  
In Situ Hybridization ◽  
Mammary Gland ◽  
Female Rats ◽  
Northern Analysis ◽  
Mammary Tissue ◽  
Secretory Cells ◽  
Northern Blot Hybridization ◽  
Target Tissue ◽  
Rat Mammary Gland

ABSTRACT In-situ hybridization and Northern blot hybridization were used to identify mRNA for pituitary prolactin in mammary tissue obtained from female rats 1 day before expected parturition, 1 day after parturition and on day 7 of lactation. Prolactin cDNA was labelled with 32P for Northern analysis and with digoxigenin for in-situ hybridization. Total and poly(A)+ RNA from pituitary, mammary and control (fat and kidney) tissues were analysed by agarose gel electrophoresis with transfer to nitrocellulose and hybridization to a cDNA for rat prolactin. Although present in much smaller amounts than the 1·0 kb transcript in pituitary RNA homogenates, mammary RNA homogenates from all three stages contained mRNA of approximately 1·0 kb which hybridized with the prolactin probe. Similar analyses of fat and kidney failed to reveal any hybridization at the 1·0 kb size. When tissue sections were hybridized to the cDNA probe, specific hybridization was observed in the milk secretory cells of the mammary alveoli and the lactotroph cells of the anterior pituitary, but not in liver cells or in RNase-treated sections of mammary tissue. In summary, these results demonstrate that milk secretory cells of the rat mammary gland transcribe the gene for prolactin, and they raise the possibility that a primary target tissue for blood-borne prolactin may also synthesize prolactin. Journal of Endocrinology (1993) 136, 271–276

scholarly journals The expression of the IGF family and GH receptor in the bovine mammary gland

2001 ◽  
Vol 168 (1) ◽  
pp. 39-48 ◽  
Author(s):  
A Plath-Gabler ◽  
C Gabler ◽  
F Sinowatz ◽  
B Berisha ◽  
D Schams
Keyword(s):  
Mammary Gland ◽  
Binding Proteins ◽  
Mammary Development ◽  
Mammary Tissue ◽  
Bovine Mammary Gland ◽  
Igf Binding Proteins ◽  
Gh Receptor ◽  
Igf I ◽  
Temporal Expressions ◽  
Igfbp 3

To study the involvement of the IGFs in mammary development and lactation of the cow, the temporal expressions of IGF-I and -II, its receptor type 1 (IGFR-1), IGF-binding proteins (IGFBPs)-1 to -6 and GH receptor (GHR) mRNA were examined. This was carried out for different stages of mammogenesis, lactogenesis, galactopoiesis and involution in the bovine mammary gland of 26 animals. Furthermore, IGF-I was localised by immunohistochemistry. The highest mRNA concentrations for IGF-I were detected in the mammary tissue of late pregnant heifers (days 255-272) and significantly lower expression was detected during lactogenesis and galactopoiesis. Immunohistochemistry of IGF-I revealed only a weak staining in the epithelium of the ducts during mammogenesis. The epithelium of the alveoli were negative during mammogenesis, lactogenesis and galactopoiesis but displayed distinct IGF-I activity during involution. In the stroma a distinct staining of the cytoplasm of adipocytes and of vascular smooth muscle cells was observed. A certain percentage of fibroblasts (usually 20-30%) were also immunopositive. In contrast, highest expression for IGFR-1 was detected during galactopoiesis and involution. The lowest mRNA concentration for IGFR-1 was found during pregnancy (days 194-213). In general, the expression of IGF-II was not regulated during mammogenesis and lactation, but decreased during involution. The mRNA for the six binding proteins was detected in the bovine mammary gland. The dominant binding proteins were IGFBP-3 and -5. The highest expression of IGFBP-3 was observed during mid-pregnancy and the lowest during late lactation, involution and in non-pregnant heifers. The mRNA for IGFBP-5 increased during late mammogenesis and lactogenesis followed by a decrease thereafter. In general, the mRNA concentrations for IGFBP-2, -4 and -6 were barely detectable during all stages. In contrast, the expression for IGFBP-1 was upregulated in the mammary gland of virgin heifers and increased around the onset of lactation. mRNA for GHR was found during all stages examined without outstanding fluctuations. In conclusion, locally produced IGF-I and -II may mediate mammogenesis. The high mammary IGFR-1 mRNA during lactation suggests a role for peripheral IGF-I in maintenance of lactation. The role of IGFBPs in the mammary gland needs further evaluation.

Expression of transforming growth factors alpha and beta-1 messenger RNA in the bovine mammary gland during different stages of development and lactation

1997 ◽  
Vol 155 (3) ◽  
pp. 501-511 ◽  
Author(s):  
A Plath ◽  
R Einspanier ◽  
F Peters ◽  
F Sinowatz ◽  
D Schams
Keyword(s):  
Mammary Gland ◽  
Growth Factors ◽  
Messenger Rna ◽  
Mammary Tissue ◽  
Ribonuclease Protection Assay ◽  
Transforming Growth Factors ◽  
Bovine Mammary Gland ◽  
Tgf Beta ◽  
Total N ◽  
Prolactin Inhibition

It is now widely accepted that the mammary gland is under interconnected hormonal and local control. Growth factors are involved in the intercellular signalling of the gland. Our aim was the detection of transforming growth factors alpha (TGF-alpha) and beta 1 (TGF-beta 1) messenger RNA during mammogenesis, lactogenesis, galactopoiesis and involution in the bovine mammary gland (total n = 27). During these stages the RNA was assessed by means of ribonuclease protection assay and reverse transcription-polymerase chain reaction (RT-PCR). To study possible influences of oestrogen, progesterone and prolactin on growth factor expression, mammary RNA was obtained from heifers after induced mammogenesis and lactogenesis, with and without additional prolactin inhibition (total n = 20). Very low levels of TGF-alpha and TGF-beta 1 expression were detected during lactogenesis and galactopoiesis, increasing levels during mammogenesis of primigravid heifers, and highest levels during mammogenesis of virgin heifers and during involution. TGF-alpha expression after induced mammogenesis was greater than after induced lactogenesis or physiological mammogenesis during pregnancy. Furthermore, TGF-alpha mRNA contents increased after prolactin inhibition. TGF-beta 1 expression was almost equal after induced mammogenesis and lactogenesis, but greater than during the physiological mammogenesis and lactogenesis. In conclusion, it can be assumed that growth promoting TGF-alpha and growth inhibiting TGF-beta 1 are co-expressed in the bovine mammary gland. Higher mRNA contents of both factors during mammogenesis and involution may indicate autocrine or paracrine functions for these growth factors during proliferation and reorganisation of the mammary tissue.

Transforming growth factor-α messenger RNA localization in the developing adult rat and human mammary gland by in situ hybridization

1990 ◽  
Vol 140 (1) ◽  
pp. 123-131 ◽  
Author(s):  
Daniel S. Liscia ◽  
Giorgio Merlo ◽  
Fortunato Ciardiello ◽  
Nancy Kim ◽  
Gilbert H. Smith ◽  
...  
Keyword(s):  
In Situ Hybridization ◽  
Mammary Gland ◽  
Messenger Rna ◽  
Transforming Growth Factor ◽  
Rna Localization ◽  
Transforming Growth Factor Α ◽  
Adult Rat ◽  
Human Mammary Gland ◽  
Factor Α

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.

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