091 Detection of androgen receptor, type I and type II 5α-reductase mRNAs in human dermal papilla cells by in situ hybridization

During limb development, type I collagen disappears from the region where cartilage develops and synthesis of type II collagen, which is characteristic of cartilage, begins. In situ hybridization using antisense RNA probes was used to investigate the spatial localization of type I and type II collagen mRNAs. The distribution of the mRNA for type II collagen corresponded well with the pattern of type II collagen synthesis, suggesting control at the level of transcription and mRNA accumulation. In contrast, the pattern of mRNA for type I collagen remained more or less uniform and did not correspond with the synthesis of the protein, suggesting control primarily at the level of translation or of RNA processing.

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Assignment1 of interleukin-1 receptor, type I (IL1R1) to bovine chromosome band 11q12 by in situ hybridization

Cytogenetic and Genome Research ◽

10.1159/000015605 ◽

2000 ◽

Vol 89 (3-4) ◽

pp. 166-167

Author(s):

B. Castiglioni ◽

S. Comincini ◽

M.G. Foti ◽

L. Ferretti

Keyword(s):

In Situ Hybridization ◽

Interleukin 1 ◽

Bovine Chromosome ◽

Receptor Type ◽

Chromosome Band ◽

Type I

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21 Expression of androgen receptor, type I and type II 5 α-reductase in human hair follicle cells

Journal of Dermatological Science ◽

10.1016/0923-1811(96)83591-2 ◽

1996 ◽

Vol 12 (1) ◽

pp. 86

Author(s):

S. Itami ◽

J. Nakanishi ◽

K. Yoshikawa ◽

S. Takayasu

Keyword(s):

Androgen Receptor ◽

Hair Follicle ◽

Human Hair ◽

Receptor Type ◽

Follicle Cells ◽

Type I ◽

Type Ii ◽

Human Hair Follicle

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Detection and Enumeration of Methanotrophs in Acidic Sphagnum Peat by 16S rRNA Fluorescence In Situ Hybridization, Including the Use of Newly Developed Oligonucleotide Probes for Methylocella palustris

Applied and Environmental Microbiology ◽

10.1128/aem.67.10.4850-4857.2001 ◽

2001 ◽

Vol 67 (10) ◽

pp. 4850-4857 ◽

Cited By ~ 113

Author(s):

Svetlana N. Dedysh ◽

Manigee Derakshani ◽

Werner Liesack

Keyword(s):

In Situ Hybridization ◽

16S Rrna ◽

Fluorescence In Situ Hybridization ◽

Fluorescence Signal ◽

Type I ◽

Oligonucleotide Probes ◽

Type Ii ◽

Cell Counts ◽

Wet Weight

ABSTRACT Two 16S rRNA-targeted oligonucleotide probes, Mcell-1026 and Mcell-181, were developed for specific detection of the acidophilic methanotroph Methylocella palustris using fluorescence in situ hybridization (FISH). The fluorescence signal of probe Mcell-181 was enhanced by its combined application with the oligonucleotide helper probe H158. Mcell-1026 and Mcell-181, as well as 16S rRNA oligonucleotide probes with reported group specificity for either type I methanotrophs (probes M-84 and M-705) or theMethylosinus/Methylocystis group of type II methanotrophs (probes MA-221 and M-450), were used in FISH to determine the abundance of distinct methanotroph groups in aSphagnum peat sample of pH 4.2. M. palustris was enumerated at greater than 106 cells per g of peat (wet weight), while the detectable population size of type I methanotrophs was three orders of magnitude below the population level of M. palustris. The cell counts with probe MA-221 suggested that only 104 type II methanotrophs per g of peat (wet weight) were present, while the use of probe M-450 revealed more than 106 type II methanotroph cells per g of the same samples. This discrepancy was due to the fact that probe M-450 targets almost all currently known strains of Methylosinus andMethylocystis, whereas probe MA-221, originally described as group specific, does not detect a large proportion ofMethylocystis strains. The total number of methanotrophic bacteria detected by FISH was 3.0 (±0.2) × 106 cells per g (wet weight) of peat. This was about 0.8% of the total bacterial cell number. Thus, our study clearly suggests that M. palustris and a defined population ofMethylocystis spp. were the predominant methanotrophs detectable by FISH in an acidic Sphagnum peat bog.

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Differential localization of mRNAs of collagen types I and II in chick fibroblasts, chondrocytes, and corneal cells by in situ hybridization using cDNA probes.

The Journal of Cell Biology ◽

10.1083/jcb.102.6.2302 ◽

1986 ◽

Vol 102 (6) ◽

pp. 2302-2309 ◽

Cited By ~ 79

Author(s):

M Hayashi ◽

Y Ninomiya ◽

J Parsons ◽

K Hayashi ◽

B R Olsen ◽

...

Keyword(s):

Endothelial Cells ◽

In Situ Hybridization ◽

Epithelial Cells ◽

Type Ii Collagen ◽

Type I ◽

Type Ii ◽

Collagen Types ◽

Paraffin Sections ◽

Extracellular Matrix Components

We have employed a highly specific in situ hybridization protocol that allows differential detection of mRNAs of collagen types I and II in paraffin sections from chick embryo tissues. All probes were cDNA restriction fragments encoding portions of the C-propeptide region of the pro alpha-chain, and some of the fragments also encoded the 3'-untranslated region of mRNAs of either type I or type II collagen. Smears of tendon fibroblasts and those of sternal chondrocytes from 17-d-old chick embryos as well as paraffin sections of 10-d-old whole embryos and of the cornea of 6.5-d-old embryos were hybridized with 3H-labeled probes for either type I or type II collagen mRNA. Autoradiographs revealed that the labeling was prominent in tendon fibroblasts with the type I collagen probe and in sternal chondrocytes with the type II collagen probe; that in the cartilage of sclera and limbs from 10-d-old embryos, the type I probe showed strong labeling of fibroblast sheets surrounding the cartilage and of a few chondrocytes in the cartilage, whereas the type II probe labeled chondrocytes intensely and only a few fibroblasts; and that in the cornea of 6.5-d-old embryos, the type I probe labeled the epithelial cells and fibroblasts in the stroma heavily, and the endothelial cells slightly, whereas the type II probe labeled almost exclusively the epithelial cells except for a slight labeling in the endothelial cells. These data indicate that embryonic tissues express these two collagen genes separately and/or simultaneously and offer new approaches to the study of the cellular regulation of extracellular matrix components.

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Localization of types I, II, and III collagen mRNAs in developing human skeletal tissues by in situ hybridization

The Journal of Cell Biology ◽

10.1083/jcb.104.4.1077 ◽

1987 ◽

Vol 104 (4) ◽

pp. 1077-1084 ◽

Cited By ~ 210

Author(s):

M Sandberg ◽

E Vuorio

Keyword(s):

In Situ Hybridization ◽

Type I Collagen ◽

Cdna Probe ◽

Type Ii Collagen ◽

Northern Hybridization ◽

Cdna Clones ◽

Sequence Information ◽

Type I ◽

Type Ii

Paraffin sections of human skeletal tissues were studied in order to identify cells responsible for production of types I, II, and III collagens by in situ hybridization. Northern hybridization and sequence information were used to select restriction fragments of cDNA clones for the corresponding mRNAs to obtain probes with a minimum of cross-hybridization. The specificity of the probes was proven in hybridizations to sections of developing fingers: osteoblasts and chondrocytes, known to produce only one type of fibrillar collagen each (I and II, respectively) were only recognized by the corresponding cDNA probes. Smooth connective tissues exhibited variable hybridization intensities with types I and III collagen cDNA probes. The technique was used to localize the activity of type II collagen production in the different zones of cartilage during the growth of long bones. Visual inspection and grain counting revealed the highest levels of pro alpha 1(II) collagen mRNAs in chondrocytes of the lower proliferative and upper hypertrophic zones of the growth plate cartilage. This finding was confirmed by Northern blotting of RNAs isolated from epiphyseal (resting) cartilage and from growth zone cartilage. Analysis of the osseochondral junction revealed virtually no overlap between hybridization patterns obtained with probes specific for type I and type II collagen mRNAs. Only a fraction of the chondrocytes in the degenerative zone were recognized by the pro alpha 1(II) collagen cDNA probe, and none by the type I collagen cDNA probe. In the mineralizing zone virtually all cells were recognized by the type I collagen cDNA probe, but only very few scattered cells appeared to contain type II collagen mRNA. These data indicate that in situ hybridization is a valuable tool for identification of connective tissue cells which are actively producing different types of collagens at the various stages of development, differentiation, and growth.

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Differential localization of distinct keratin mRNA-species in mouse tongue epithelium by in situ hybridization with specific cDNA probes.

The Journal of Cell Biology ◽

10.1083/jcb.103.6.2583 ◽

1986 ◽

Vol 103 (6) ◽

pp. 2583-2591 ◽

Cited By ~ 35

Author(s):

M Rentrop ◽

B Knapp ◽

H Winter ◽

J Schweizer

Keyword(s):

In Situ Hybridization ◽

Basal Layer ◽

Hybridization Technique ◽

Type I ◽

Type Ii ◽

Basal Cells ◽

Fungiform Papillae ◽

Filiform Papillae ◽

Tongue Epithelium

The tongue of the adult mouse is covered by a multilayered squamous epithelium which is continuous on the ventral surface, however interrupted on the dorsal surface by many filiform and few fungiform papillae. The filiform papillae themselves are subdivided into an anterior and posterior unit exhibiting different forms of keratinization. Thus, the entire epithelium shows a pronounced morphological diversity of well recognizable tissue units. We have used a highly sensitive in situ hybridization technique to investigate the differential expression of keratin mRNAs in the tongue epithelium. The hybridization probes used were cDNA restriction fragments complementary to the most specific 3'-regions of any given keratin mRNA. We could show that independent of the morphologically different tongue regions, all basal cells uniformly express the mRNA of a type I 52-kD keratin, typical also for basal cells of the epidermis. Immediately above the homogenous basal layer a vertically oriented specialization of the keratin expression occurs within the morphological tissue units. Thus the dorsal interpapillary and ventral epithelium express the mRNAs of a type II 57-kD and a type I 47-kD keratin pair. In contrast, in the anterior unit of the filiform papillae, only the 47-kD mRNA is present, indicating that this keratin may be coexpressed in tongue epithelium with different type II partners. In suprabasal cells of both, the fungiform papillae and the posterior unit of the filiform papillae, a mRNA of a type I 59-kD keratin could be detected; however, its type II 67-kD epidermal counterpart seems not to be present in these cells. Most surprisingly, in distinct cells of both types of papillae, a type I 50-kD keratin mRNA could be localized which usually is associated with epidermal hyperproliferation. In conclusion, the in situ hybridization technique applied has been proved to be a powerful method for detailed studies of differentiation processes, especially in morphologically complex epithelia.

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Fluorescence In Situ Hybridization-Flow Cytometry-Cell Sorting-Based Method for Separation and Enrichment of Type I and Type II Methanotroph Populations

Applied and Environmental Microbiology ◽

10.1128/aem.00161-06 ◽

2006 ◽

Vol 72 (6) ◽

pp. 4293-4301 ◽

Cited By ~ 78

Author(s):

Marina G. Kalyuzhnaya ◽

Rebecca Zabinsky ◽

Sarah Bowerman ◽

David R. Baker ◽

Mary E. Lidstrom ◽

...

Keyword(s):

Flow Cytometry ◽

In Situ Hybridization ◽

16S Rrna ◽

Fluorescence In Situ Hybridization ◽

Cell Sorting ◽

Type I ◽

Type Ii ◽

Specific Probe ◽

Artificial Mixtures

ABSTRACT A fluorescence in situ hybridization-flow cytometry (FISH/FC)-based method was optimized using artificial mixtures of pure cultures of methanotrophic bacteria. Traditional oligonucleotide probes targeting 16S rRNAs of type I (MG84/705 probe) and type II (MA450 probe) methanotrophs were labeled with fluorescein or Alexa fluor and used for FISH, followed by fluorescence-activated FC analysis and cell sorting (FACS). The method resulted in efficient separation of target cells (type I or type II methanotrophs) from the artificial mixtures. The method was then applied for detection and enrichment of type I and type II methanotroph populations from a natural sample, Lake Washington sediment. Cells were extracted from the sediment, fixed, and subjected to FISH/FC/FACS. The resulting subpopulations were analyzed by reverse transcriptase PCR surveys of 16S rRNA, pmoA (encoding a subunit of particulate methane monooxygenase), and fae (encoding formaldehyde-activating enzyme) genes. The functional gene analysis indicated specific separation of the type I and type II methanotroph populations. 16S rRNA gene analysis revealed that type I methanotrophs comprised 59% of the subpopulation separated using the type I-specific probe and that type II methanotrophs comprised 47.5% of the subpopulation separated using the type II-specific probe. Our data indicate that the FISH/FC/FACS protocol described can provide significant enrichment of microbial populations of interest from complex natural communities and that these can be used for genetic tests. We further tested the possibility of direct whole-genome amplification (WGA) from limited numbers of sorted cells, using artificial mixtures of microbes whose genome sequences are known. We demonstrated that efficient WGA can be achieved using 104 or more cells separated by 16S rRNA-specific FISH/FC/FACS, while fewer cells resulted in less specific WGA.

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Enhanced expression of TGF-beta and c-fos mRNAs in the growth plates of developing human long bones

Development ◽

10.1242/dev.102.3.461 ◽

1988 ◽

Vol 102 (3) ◽

pp. 461-470 ◽

Cited By ~ 8

Author(s):

M. Sandberg ◽

T. Vuorio ◽

H. Hirvonen ◽

K. Alitalo ◽

E. Vuorio

Keyword(s):

In Situ Hybridization ◽

Expression Patterns ◽

Northern Blotting ◽

Northern Hybridization ◽

Type I ◽

Long Bones ◽

Type Ii ◽

Tgf Beta ◽

Growth Plates

The expression of mRNAs for type I and type II procollagens, transforming growth factor-beta (TGF-beta) and c-fos was studied in developing human long bones by Northern blotting and in situ hybridization. The cells producing bone and cartilage matrix were identified by hybridizations using cDNA probes for types I and II collagen, respectively. Northern blotting revealed that the highest levels of TGF-beta mRNA were associated with the growth plates. By in situ hybridization, this mRNA was localized predominantly in the osteoblasts and osteoclasts of the developing bone, in periosteal fibroblasts and in individual bone marrow cells. These findings are consistent with the view that TGF-beta may have a role in stimulation of type I collagen production and bone formation. Only a low level of TGF-beta mRNA was detected in cartilage where type II collagen mRNA is abundant. In Northern hybridization, the highest levels of c-fos mRNA were detected in epiphyseal cartilage. In situ hybridization revealed two cell types with high levels of c-fos expression: the chondrocytes bordering the joint space and the osteoclasts of developing bone. These differential expression patterns suggest specific roles for TGF-beta and c-fos in osseochondral development.

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