scholarly journals Expression of histidine decarboxylase and cellular histamine-like immunoreactivity in rat embryogenesis.

1995 ◽  
Vol 43 (12) ◽  
pp. 1241-1252 ◽  
Author(s):  
M J Nissinen ◽  
K Karlstedt ◽  
E Castrén ◽  
P Panula
Keyword(s):  
Mast Cells ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Striated Muscle ◽  
Histidine Decarboxylase ◽  
Muscle Cells ◽  
Nerve Fibers ◽  
Developmental Expression ◽  
Mrna Species

In this study we investigated the developmental expression of histidine decarboxylase (HDC) mRNA and the distribution of histamine-immunoreactive (histamine-ir) cells in the rat embryonic tissues. We applied Northern blot analysis, in situ hybridization with synthetic oligonucleotide probes complementary to the rat HDC cDNA, and indirect histamine immunocytochemistry. Northern blot analysis revealed the appearance of a major (2.6 KB) HDC mRNA species in liver on embryonic Day 14. Its hybridization level peaked on Day E18, when two minor (1.6 and 3.5 KB) mRNA species were also present. During the periparturition period, a rapid decrease in HDC RNA was apparent, as the 2.6 KB mRNA species was expressed at a low level on postnatal Day P1. The embryonic liver expressed HDC on days E14-E20. On days E18 and E20, the periosteum and the epiphyseal growth plates of the endochondrally ossificating bones, and some striated muscle cells, showed hybridization signal for HDC. Histamine immunoreactivity was detected in many epithelial and neuronal cell types during embryogenesis. An intense histamine immunoreaction appeared first in essentially all cells of the liver parenchyma on day E12. This parenchymal histamine immunoreactivity disappeared by birth, after which this immunofluorescence in liver was restricted to a few scattered mast cells until adulthood. Some neurons in the peripheral sensory, sympathetic and cranial nerve ganglia were histamine-immunoreactive from day E16 to birth. In addition, many immunoreactive nerve fibers were detected in the gastrointestinal muscularis externa, mesentery, salivary glands, kidney, lung, and muscle tissue. We conclude that during rat embryogenesis histamine is produced and stored transiently by cells in liver, developing bone, and a few striated muscle cells, in addition to previously reported neurons in rat brain. Many peripheral neurons, epithelial cells, and mast cells display histamine immunoreactivity during rat embryogenesis but are devoid of detectable HDC mRNA with the current method. It remains possible that histamine is formed by another enzyme or is taken up from the extracellular space. The results support the concept that a significant proportion of histamine is formed and stored by embryonic cells other than mast cells.

scholarly journals Tissue- and cell-specific expression of Ins(1,4,5)P3 3-kinase isoenzymes

1995 ◽  
Vol 306 (2) ◽  
pp. 429-435 ◽  
Author(s):  
V Vanweyenberg ◽  
D Communi ◽  
C S D'Santos ◽  
C Erneux
Keyword(s):  
Cell Lines ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Biological Data ◽  
Human Neuroblastoma Cell ◽  
Rat Tissues ◽  
Human Neuroblastoma ◽  
Mrna Species ◽  
Kinase A

The phosphorylation of Ins(1,4,5)P3 (InsP3) to Ins(1,3,4,5)P4 (InsP4) is catalysed by InsP3 3-kinase. Molecular-biological data have shown the presence of two human isoenzymes of InsP3 3-kinase, namely InsP3 3-kinases A and B. We have isolated from a rat thymus cDNA library a 2235 bp cDNA (clone B15) encoding rat InsP3 3-kinase B. Northern-blot analysis of mRNA isolated from rat tissues (thymus, testis, brain, spleen, liver, kidney, heart, lung and intestine) revealed that a rat InsP3 3-kinase B probe hybridized to a 6 kb mRNA in lung, thymus, testis, brain and heart. In contrast, Northern-blot analysis of the same tissues probed under stringent conditions with a rat InsP3 3-kinase A probe hybridized to a 2 kb mRNA only in brain and a 1.8-2.0 kb mRNA species in testis. Northern-blot analysis of three human cell lines (HL-60, SH-SY5Y and HTB-138) probed with a human InsP3 3-kinase B probe showed the presence of a 6 kb mRNA in all cell lines, except in the human neuroblastoma cell line (SH-SY5Y), where two mRNA species of 5.7 and 6 kb were detected. Using the same blot, no hybridization signal could be seen with a human InsP3 3-kinase A probe. Altogether, our data are consistent with the notion that the two InsP3 3-kinase isoenzymes, A and B, are specifically expressed in different tissues and cells.

scholarly journals Expression of Cyclooxygenase-2 and Prostanoid Receptors by Human Myometrium*

2000 ◽  
Vol 85 (9) ◽  
pp. 3468-3475 ◽  
Author(s):  
Tiina-Liisa Erkinheimo ◽  
Kirsi Saukkonen ◽  
Kirsi Narko ◽  
Jyrki Jalkanen ◽  
Olavi Ylikorkala ◽  
...  
Keyword(s):  
Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Muscle Cells ◽  
Human Myometrium ◽  
Fetal Membranes ◽  
Prostanoid Receptors ◽  
Cox 2

Abstract Prostanoids play an important role in the regulation of parturition. All reproductive tissues, including fetal membranes, decidua, and myometrium, have the capacity to synthesize prostanoids, and fetal membranes have been shown to express elevated levels of cyclooxygenase-2 (Cox-2) at the onset of labor. We have now investigated the expression of Cox-2 in human myometrium. Myometrial samples collected from women in labor during lower segment cesarean section expressed 15-fold higher levels of Cox-2 messenger ribonucleic acid (mRNA) compared to myometrial specimens collected from women not in labor, as detected by Northern blot analysis. Immunohistochemical detection of Cox-2 protein showed cytoplasmic staining in the smooth muscle cells of the myometrium. Cultured myometrial cells expressed low levels of Cox-2 mRNA under baseline conditions, but interleukin-1β (IL-1β) caused a 17-fold induction of expression of the Cox-2 transcript after incubation for 6 h. IL-1β also induced expression of biologically active Cox-2 protein, as detected by immunofluorescence, Western blot analysis, and measuring the conversion of arachidonic acid to prostanoids in the presence and absence of a Cox-2-selective inhibitor, NS-398. PGE2 receptor subtype EP2 mRNA was expressed in cultured myometrial smooth muscle cells, whereas transcripts for EP1, EP3, EP4, FP, and IP were low or below the detection limit as measured by Northern blot analysis. However, IL-1β stimulated expression of EP4 receptor mRNA. Our data suggest that expression of Cox-2 transcript is elevated at the onset of labor in myometrial smooth muscle cells, which may depend on induction by cytokines. As, in addition to Cox-2, the expression of prostanoid receptors is regulated, not only the production of prostanoids, but also responsiveness to them, may be modulated.

Characterization of neutral endopeptidase in vascular smooth muscle cells of rabbit renal cortex

1993 ◽  
Vol 264 (1) ◽  
pp. F45-F52
Author(s):  
J. C. Dussaule ◽  
A. Stefanski ◽  
M. L. Bea ◽  
P. Ronco ◽  
R. Ardaillou
Keyword(s):  
Smooth Muscle ◽  
Vascular Smooth Muscle ◽  
Smooth Muscle Cells ◽  
Vascular Smooth Muscle Cells ◽  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Renal Cortex ◽  
Muscle Cells ◽  
Neutral Endopeptidase

In addition to biological and clearance receptors for atrial natriuretic factor (ANF), cultured vascular smooth muscle cells (VSMC) from the rabbit renal cortex possess ectoenzymes degrading this hormone. We examined whether neutral endopeptidase (NEP) was implicated in this process. The presence of NEP in VSMC was demonstrated as follows. 1) NEP activity measured from the hydrolysis of a synthetic substrate by intact cells cultured in a medium containing 10% fetal calf serum was 1,609 +/- 65 pmol.min-1 x mg-1 [surface localization of the enzyme was confirmed by low activity (4% of total) in the cytosol; release of NEP activity in the medium was negligible]; 2) a monoclonal antibody directed against rabbit NEP specifically stained VSMC membranes; and 3) mRNA from VSMC hybridized a NEP cDNA probe with a single band as shown by Northern blot analysis. The role of NEP in ANF catabolism was demonstrated by incubating 125I-ANF or unlabeled ANF for increasing periods of time with VSMC in the presence of thiorphan (1-100 microM). Intact hormone estimated by trichloroacetic acid precipitation or radio-immunoassay, respectively, increased markedly compared with control in the presence of this specific inhibitor of NEP. NEP activity was stimulated (x1.6) in quiescent VSMC deprived from serum during 3 days. This effect was dose dependent and was not observed with creatine kinase activity measured as control. NEP expression at the cell surface estimated by sorting of immunostained cells was also increased in the absence of serum. Northern blot analysis showed increases in the mRNA band of NEP with increasing periods of serum deprivation.(ABSTRACT TRUNCATED AT 250 WORDS)

scholarly journals IL-8 and MCP Gene Expression and Production by LPS-Stimulated Human Corneal Stromal Cells

2012 ◽  
Vol 2012 ◽  
pp. 1-4 ◽  
Author(s):  
Roni M. Shtein ◽  
Susan G. Elner ◽  
Zong-Mei Bian ◽  
Victor M. Elner
Keyword(s):  
Gene Expression ◽  
Northern Blot ◽  
Stromal Cells ◽  
Blot Analysis ◽  
Time Course ◽  
Northern Blot Analysis ◽  
Statistical Significance ◽  
Dependent Manner ◽  
Corneal Inflammation ◽  
Chemotactic Protein

Purpose. To determine time course of effect of lipopolysaccharide (LPS) on production of interleukin-8 (IL-8) and monocyte chemotactic protein (MCP) by cultured human corneal stromal cells.Methods. Human corneal stromal cells were harvested from donor corneal specimens, and fourth to sixth passaged cells were used. Cell cultures were stimulated with LPS for 2, 4, 8, and 24 hours. Northern blot analysis of IL-8 and MCP gene expression and ELISA for IL-8 and MCP secretion were performed. ELISA results were analyzed for statistical significance using two-tailed Student'st-test.Results. Northern blot analysis demonstrated significantly increased IL-8 and MCP gene expression after 4 and 8 hours of exposure to LPS. ELISA for secreted IL-8 and MCP demonstrated statistically significant increases (P<0.05) after corneal stromal cell stimulation with LPS.Conclusions. This paper suggests that human corneal stromal cells may participate in corneal inflammation by secreting potent leukocyte chemotactic and activating proteins in a time-dependent manner when exposed to LPS.

scholarly journals A rapid and accurate method for quantitating total RNA transferred during Northern blot analysis

1988 ◽  
Vol 16 (5) ◽  
pp. 2354-2354 ◽  
Author(s):  
Nathalie Denis ◽  
Daniel Corcos ◽  
Jacques Kruh ◽  
Alain Kitzis
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Accurate Method ◽  
Total Rna

The mouse prostaglandin E receptor EP2 subtype: cloning, expression, and Northern blot analysis

FEBS Letters ◽  
1995 ◽  
Vol 372 (2-3) ◽  
pp. 151-156 ◽  
Author(s):  
Masato Katsuyama ◽  
Nobuhiro Nishigaki ◽  
Yukihiko Sugimoto ◽  
Kimiko Morimoto ◽  
Manabu Negishi ◽  
...  
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Prostaglandin E

A procedure for Northern blot analysis of native RNA

1986 ◽  
Vol 159 (1) ◽  
pp. 227-232 ◽  
Author(s):  
Edouard W. Khandjian ◽  
Claude Méric
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis

The expression of NG2 proteoglycan in the developing rat limb

Development ◽  
1991 ◽  
Vol 111 (4) ◽  
pp. 933-944 ◽  
Author(s):  
A. Nishiyama ◽  
K.J. Dahlin ◽  
W.B. Stallcup
Keyword(s):  
Northern Blot ◽  
Blot Analysis ◽  
Northern Blot Analysis ◽  
Core Protein ◽  
Staining Intensity ◽  
Limb Bud ◽  
Protein Size ◽  
Glial Progenitor Cells ◽  
Ng2 Proteoglycan ◽  
Developing Rat

NG2 is a chondroitin sulfate proteoglycan previously found to be expressed by glial progenitor cells of the O2A lineage. We have examined the expression of NG2 in the developing rat limb by immunohistochemistry and northern blot analysis. Staining of embryonic day 14 (E14) rat limb bud sections with polyclonal and monoclonal anti-NG2 antibodies reveals reactivity in the precartilaginous mesenchymal condensation. The staining intensity increases with the differentiation of chondrocytes until E16. NG2 staining is not detected in the mature hypertrophic chondrocytes of E17 and postnatal day 3 (P3) limbs even after treatment of the sections with hyaluronidase or collagenase. Immuno-precipitations with anti-NG2 antibody using 125I-labeled limb cells in culture showed a 400 to 800 × 10(3) Mr proteoglycan species with a core protein size of 300 × 10(3) Mr, comparable to NG2 from O2A cells and neural cell lines. Northern blot analysis reveals the expression of an 8.9 kb mRNA in E16 limbs and at a lower level in P1 cartilage. The northern blot analyses also show that NG2 is distinct from the large aggregating proteoglycan of the cartilage. Our results indicate that in the developing limb cartilage, as in the differentiating oligodendrocytes, NG2 is present on immature cells in the process of differentiating, but its expression is downregulated as terminal differentiation of chondrocytes takes place.

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