Acinar Cells Are Target Cells for Androgens in Mouse Submandibular Glands

The variable coding sequence (VCS) multigene family encodes diverse salivary proteins, such as the SMR1 prohormone and the PR-VB1 proline-rich protein in the rat. In situ hybridization was used to study the cell-specific expression of two new mouse VCS genes, Vcs1 and Vcs2. We show that the Vcs1 transcripts, which code for a proline-rich protein, MSG1, are highly abundant in male and female parotid glands, in which they are specifically detected in acinar cells. No expression was seen in the submandibular or sublingual glands. In contrast, Vcs2 transcripts were found only in the acinar cells of the submandibular glands (SMGs) of male mice, in which they are expressed in response to androgens. Expression was found to be heterogeneous within acinar structures. No Vcs2 transcripts were detected in the SMGs of females or castrated males by Northern blot, RNase protection, or in situ hybridization. Androgen administration to females or castrated males induced expression at a level comparable to that of intact males. The Vcs2 gene is the first example of a mouse androgen-regulated gene that is expressed in SMG acinar cells. This result, in addition to our previous observation on SMR1 expression in rats, demonstrates that both acinar cells and granular convoluted tubule (GCT) cells are target cells for androgen action in rodent SMG.

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IL-1β and IL-6 in mouse parotid acinar cells: characterization of synthesis, storage, and release

AJP Gastrointestinal and Liver Physiology ◽

10.1152/ajpgi.1998.274.1.g147 ◽

1998 ◽

Vol 274 (1) ◽

pp. G147-G156 ◽

Cited By ~ 3

Author(s):

Naoko Tanda ◽

Hiroe Ohyama ◽

Midori Yamakawa ◽

Maria Ericsson ◽

Takanori Tsuji ◽

...

Keyword(s):

In Situ Hybridization ◽

Time Course ◽

Light And Electron Microscopy ◽

Acinar Cells ◽

Secretory Cells ◽

Parotid Glands ◽

Adrenergic Stimulation ◽

Rt Pcr ◽

Amylase Release

Synthesis, storage, and secretion of the proinflammatory cytokine interleukin-1β (IL-1β) and the anti-inflammatory cytokine IL-6 have not been established in normal exocrine gland secretory cells. Parotid glands and isolated acinar cells prepared from BALB/c mice were homogenized for RNA isolation and reverse transcription-polymerase chain reaction (RT-PCR). IL-1β and IL-6 enzyme-linked immunosorbent assays (ELISAs) were done on supernatants prepared from mouse parotid acinar cell (MPAC) preparations unstimulated or stimulated between 0 and 10 min with 10−5M norepinephrine at 37°C. MPACs were fixed in paraformaldehyde, frozen sectioned for light and electron microscopy, and labeled with antibodies to IL-1β and IL-6. Mouse specific riboprobes to IL-1 and IL-6 were used for in situ hybridization. RT-PCR yielded the expected IL-1 (336-bp) and IL-6 (614-bp) mRNA products. By ELISA, stimulated MPACs showed a significant increase in IL-1β ( P < 0.03) and IL-6 ( P < 0.01) release into supernatants by 10 min that paralleled the time course of amylase release. In situ hybridization showed the presence of transcripts for IL-1 and IL-6 in glandular epithelial cells. Gold-labeled IL-1β and IL-6 were significantly higher ( P < 0.01) in granules than in the nucleus and cytoplasm. This study shows that MPACs synthesize IL-1β and IL-6 and release these cytokines from their granules after α- and β-adrenergic stimulation.

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Expression of the mRNAs for the Kv3.1 potassium channel gene in the adult and developing rat brain

Journal of Neurophysiology ◽

10.1152/jn.1992.68.3.756 ◽

1992 ◽

Vol 68 (3) ◽

pp. 756-766 ◽

Cited By ~ 158

Author(s):

T. M. Perney ◽

J. Marshall ◽

K. A. Martin ◽

S. Hockfield ◽

L. K. Kaczmarek

Keyword(s):

In Situ Hybridization ◽

Rat Brain ◽

K Channel ◽

Rnase Protection ◽

Hybridization Histochemistry ◽

Adult Rat ◽

In Situ Hybridization Histochemistry ◽

Adult Rat Brain ◽

Developing Rat

1. The gene for a mammalian Shaw K+ channel has recently been cloned and has been shown, by alternative splicing, to give rise to two different transcripts, Kv3.1 alpha and Kv3.1 beta. To determine whether these channels are associated with specific types of neurons and to determine whether or not the alternately spliced K+ channel variants are differentially expressed, we used ribonuclease (RNase) protection assays and in situ hybridization histochemistry to localize the specific subsets of neurons containing Kv3.1 alpha and Kv3.1 beta mRNAs in the adult and developing rat brain. 2. In situ hybridization histochemistry revealed a heterogeneous expression pattern of Kv3.1 alpha mRNA in the adult rat brain. Highest Kv3.1 alpha mRNA levels were expressed in the cerebellum. High levels of hybridization were also detected in the globus pallidus, subthalamus, and substantia nigra reticulata. Many thalamic nuclei, but in particular the reticular thalamic nucleus, hybridized well to Kv3.1 alpha-specific probes. A subpopulation of cells in the cortex and hippocampus, which by their distribution and number may represent interneurons, were also found to contain high levels of Kv3.1 alpha mRNA. In the brain stem, many nuclei, including the inferior colliculus and the cochlear and vestibular nuclei, also express Kv3.1 alpha mRNA. Low or undetectable levels of Kv3.1 alpha mRNA were found in the caudate-putamen, olfactory tubercle, amygdala, and hypothalamus. 3. Kv3.1 beta mRNA was also detected in the adult rat brain by both RNase protection assays and by in situ hybridization experiments. Although the beta splice variant is expressed at lower levels than the alpha species, the overall expression pattern for both mRNAs is similar, indicating that both splice variants co-expressed in the same neurons. 4. The expression of Kv3.1 alpha and Kv3.1 beta transcripts was examined throughout development. Kv3.1 alpha mRNA is detected as early as embryonic day 17 and then increases gradually until approximately postnatal day 10, when there is a large increase in the amount of Kv3.1 alpha mRNA. Interestingly, the expression of Kv3.1 beta mRNA only increases gradually during the developmental time frame examined. Densitometric measurements indicated that Kv3.1 alpha is the predominant splice variant found in neurons of the adult brain, whereas Kv3.1 beta appears to be the predominant species in embryonic and perinatal neurons. 5. Most of the neurons that express the Kv3.1 transcripts have been characterized electrophysiologically to have narrow action potentials and display high-frequency firing rates with little or no spike adaptation.(ABSTRACT TRUNCATED AT 400 WORDS)

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Localization of type 1 17beta-hydroxysteroid dehydrogenase mRNA and protein in syncytiotrophoblasts and invasive cytotrophoblasts in the human term villi

Journal of Endocrinology ◽

10.1677/joe.0.1650217 ◽

2000 ◽

Vol 165 (2) ◽

pp. 217-222 ◽

Cited By ~ 18

Author(s):

M Bonenfant ◽

PR Provost ◽

R Drolet ◽

Y Tremblay

Keyword(s):

In Situ Hybridization ◽

Hydroxysteroid Dehydrogenase ◽

Binding Activity ◽

Placental Lactogen ◽

Specific Expression ◽

P450 Aromatase ◽

Chain Cleavage ◽

Extravillous Cytotrophoblasts

The 17beta-hydroxysteroid dehydrogenases (17beta-HSDs) play a key role in the synthesis of sex steroids. The hallmark of this family of enzymes is the interconversion, through their oxydoreductive reactivity at position C17, of 17-keto- and 17beta-hydroxy-steroids. Because this reaction essentially transforms steroids having low binding activity for the steroid receptor to their more potent 17beta-hydroxysteroids isoforms, it is crucial to the control of the physiological activities of both estrogens and androgens. The human placenta produces large amounts of progesterone and estrogens throughout pregnancy. The placental type 1 17beta-HSD enzyme (E17beta-HSD) catalyzes the reduction of the low activity estrogen, estrone, into the potent estrogen, estradiol. We studied the cell-specific expression of type 1 17beta-HSD in human term placental villous tissue by combining in situ hybridization to localize type 1 17beta-HSD mRNA with immunohistochemistry using an antibody against human placental lactogen, a trophoblast marker. Immunolocalization of E17beta-HSD was also performed. To ascertain whether other steroidogenic enzymes are present in the same cell type, cytochrome P450 cholesterol side-chain cleavage (P450scc), P450 aromatase, and type 1 3beta-hydroxysteroid dehydrogenase (3beta-HSD) were also localized by immunostaining. Our results showed that the syncytium is the major steroidogenic unit of the fetal term villi. In fact, type 1 17beta-HSD mRNA and protein, as well as P450scc, P450 aromatase, and 3beta-HSD immunoreactivities were found in these cells. In addition, our results revealed undoubtedly that extravillous cytotrophoblasts (CTBs), e.g. those from which cell columns of anchoring villous originate, also express the type 1 17beta-HSD gene. However, CTBs lying beneath the syncytial layer, e.g. those from which syncytiotrophoblasts develop, contained barely detectable amounts of type 1 17beta-HSD mRNA as determined by in situ hybridization. These findings, along with those from other laboratories confirm the primordial role of the syncytium in the synthesis of steroids during pregnancy. In addition, our results indicate for the first time that CTBs differentiating along the invasive pathway contain type 1 17beta-HSD mRNA.

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Stage-Specific expression of rat transition protein 2 mrna and possible localization to the chromatoid body of step 7 spermatids by in situ hybridization using a nonradioactive riboprobe

Molecular Reproduction and Development ◽

10.1002/mrd.1080330404 ◽

1992 ◽

Vol 33 (4) ◽

pp. 385-391 ◽

Cited By ~ 62

Author(s):

P. T. K. Saunders ◽

M. R. Millar ◽

S. M. Maguire ◽

R. M. Sharpe

Keyword(s):

In Situ Hybridization ◽

Chromatoid Body ◽

Specific Expression ◽

Transition Protein

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Tissue-specific expression of type 1 angiotensin II receptor subtypes. An in situ hybridization study.

Hypertension ◽

10.1161/01.hyp.24.5.531 ◽

1994 ◽

Vol 24 (5) ◽

pp. 531-537 ◽

Cited By ~ 124

Author(s):

J M Gasc ◽

S Shanmugam ◽

M Sibony ◽

P Corvol

Keyword(s):

In Situ Hybridization ◽

Angiotensin Ii ◽

Receptor Subtypes ◽

Hybridization Study ◽

Angiotensin Ii Receptor ◽

Specific Expression ◽

Tissue Specific Expression ◽

Angiotensin Ii Receptor Subtypes

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Integrin alpha subunit mRNAs are differentially expressed in early Xenopus embryos

Development ◽

10.1242/dev.117.4.1239 ◽

1993 ◽

Vol 117 (4) ◽

pp. 1239-1249 ◽

Cited By ~ 7

Author(s):

C.A. Whittaker ◽

D.W. DeSimone

Keyword(s):

In Situ Hybridization ◽

Rnase Protection Assay ◽

Early Embryo ◽

Mrna Levels ◽

Rnase Protection ◽

Dorsal Midline ◽

Transmembrane Receptors ◽

Early Embryos ◽

Whole Mount

Adhesion of cells to extracellular matrix proteins is mediated, in large part, by transmembrane receptors of the integrin family. The identification of specific integrins expressed in early embryos is an important first step to understanding the roles of these receptors in developmental processes. We have used polymerase chain reaction methods and degenerate oligodeoxynucleotide primers to identify and clone Xenopus integrin alpha subunits from neurula-stage (stage 17) cDNA. Partial cDNAs encoding integrin subunits alpha 2, alpha 3, alpha 4, alpha 5, alpha 6 and an alpha IIb-related subunit were cloned and used to investigate integrin mRNA expression in early embryos by RNase protection assay and whole-mount in situ hybridization methods. Considerable integrin diversity is apparent early in development with integrins alpha 2, alpha 3, alpha 4, alpha 5 and alpha 6 each expressed by the end of gastrulation. Both alpha 3 and alpha 5 are expressed as maternal mRNAs. Zygotic expression of alpha 2, alpha 3, alpha 4 and alpha 6 transcripts begins during gastrulation. Integrin alpha 5 is expressed at relatively high levels during cleavage, blastula and gastrula stages suggesting that it may represent the major integrin expressed in the early embryo. We demonstrated previously that integrin beta 1 protein synthesis remains constant following induction of stage 8 animal cap cells with activin (Smith, J. C., Symes, K., Hynes, R. O. and DeSimone, D. W. (1990) Development 108, 289–298.). Here we report that integrin alpha 3, alpha 4 and alpha 6 mRNA levels increase following induction with 10 U/ml activin-A whereas alpha 5, beta 1 and beta 3 mRNA levels remain unchanged. Whole-mount in situ hybridization reveals that alpha 3 mRNAs are expressed by cells of the involuting mesoderm in the dorsal lip region of early gastrulae. As gastrulation proceeds, alpha 3 expression is localized to a stripe of presumptive notochordal cells along the dorsal midline. In neurulae, alpha 3 mRNA is highly expressed in the notochord but becomes progressively more restricted to the caudalmost portion of this tissue as development proceeds from tailbud to tadpole stages. In addition, alpha 3 is expressed in the forebrain region of later stage embryos. These data suggest that integrin-mediated adhesion may be involved in the process of mesoderm involution at gastrulation and the organization of tissues during embryogenesis.

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Spatial- and temporal-restricted pattern for amelogenin gene expression during mouse molar tooth organogenesis

Development ◽

10.1242/dev.104.1.77 ◽

1988 ◽

Vol 104 (1) ◽

pp. 77-85 ◽

Cited By ~ 2

Author(s):

M.L. Snead ◽

W. Luo ◽

E.C. Lau ◽

H.C. Slavkin

Keyword(s):

Gene Expression ◽

In Situ Hybridization ◽

Gene Transcription ◽

Developmental Stages ◽

Three Dimensional ◽

Molar Tooth ◽

Specific Expression ◽

Amelogenin Gene ◽

Stage Of Development

Position- and time-restricted amelogenin gene transcription was analysed in developing tooth organs using in situ hybridization with asymmetric complementary RNA probes produced from a cDNA specific to the mouse 26 × 10(3) Mr amelogenin. In situ analysis was performed on developmentally staged fetal and neonatal mouse mandibular first (M1) and maxillary first (M1) molar tooth organs using serial sections and three-dimensional reconstruction. Amelogenin mRNA was first detected in a cluster of ameloblasts along one cusp of the M1 molar at the newborn stage of development. In subsequent developmental stages, amelogenin transcripts were detected within foci of ameloblasts lining each of the five cusps comprising the molar crown form. The number of amelogenin transcripts appeared to be position-dependent, being more abundant on one cusp surface while reduced along the opposite surface. Amelogenin gene transcription was found to be bilaterally symmetric between the developing right and left M1 molars, and complementary between the M1 and M1 developing molars; indicating position-restricted gene expression resulting in organ stereoisomerism. The application of in situ hybridization to forming tooth organ geometry provides a novel strategy to define epithelial-mesenchymal signal(s) which are believed to be responsible for organ morphogenesis, as well as for temporal- and spatial-restricted tissue-specific expression of enamel extracellular matrix.

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