Gestational changes in fetal renal and hepatic angiotensinogen mRNA and protein

1998 ◽  
Vol 10 (5) ◽  
pp. 399 ◽  
Author(s):  
David Y. Zhang ◽  
Eugenie R. Lumbers ◽  
June J. Wu
Keyword(s):  
Fetal Liver ◽  
Rnase Protection Assay ◽  
Protein Product ◽  
Late Gestation ◽  
Protection Assay ◽  
Rnase Protection ◽  
Fetal Sheep ◽  
Protein Levels ◽  
Adult Kidney ◽  
Liver And Kidney

The aim of the study was to determine the amount of angiotensinogen expression and its protein product in fetal sheep liver and kidney in the last third of gestation. Angiotensinogen mRNA was measured by RNase protection assay and its protein levels were measured by radioimmunoassay. Levels were measured at 80, 95, 111, 125 and 139 days. Angiotensinogen mRNA was present in all fetal liver and kidney samples tested. The ratio of hepatic angiotensinogen mRNA/18 S rRNA increased by 100% (P<0.001) and angiotensinogen levels increased by 33% (P<0.001) in fetal sheep from 80 to 139 d. Over the same period the ratio of renal angiotensinogen mRNA/18 S rRNA increased by 170% (P<0.001) and renal angiotensinogen protein increased by 41% (P<0.001). The levels of angiotensinogen mRNA and its protein in the adult kidney were less than in kidneys of 139 d old fetuses (P<0.01). There was a direct relationship between levels of angiotensinogen mRNA and its protein in the liver (r = 0.53, P<0.01, n = 25) and in the kidney (r = 0.75, P<0.0001, n = 24). These findings demonstrate that there is a significant increase in both hepatic and renal angiotensinogen gene expression in the last third of gestation in the fetal sheep and that this increase is associated with an increase of angiotensinogen levels in both tissues. This increase in angiotensinogen in late gestation could influence the activity of both the intrarenal and circulating renin angiotensin systems.

A new quantitative solution hybridisation-RNase protection assay for APP and APLP2 mRNA

1996 ◽  
Vol 43 (1-2) ◽  
pp. 77-84 ◽  
Author(s):  
Janet A Johnston ◽  
Svante Norgren ◽  
Göran Annerén ◽  
Richard F Cowburn ◽  
Lars Lannfelt
Keyword(s):  
Rnase Protection Assay ◽  
Protection Assay ◽  
Rnase Protection

scholarly journals A novel transcript encoding an N-terminally truncated AML1/PEBP2 alphaB protein interferes with transactivation and blocks granulocytic differentiation of 32Dcl3 myeloid cells.

1997 ◽  
Vol 17 (7) ◽  
pp. 4133-4145 ◽  
Author(s):  
Y W Zhang ◽  
S C Bae ◽  
G Huang ◽  
Y X Fu ◽  
J Lu ◽  
...  
Keyword(s):  
Fetal Liver ◽  
Myeloid Cells ◽  
Myeloid Cell ◽  
Protein Product ◽  
Terminal Region ◽  
Human Leukemia ◽  
Granulocytic Differentiation ◽  
Runt Domain ◽  
Rnase Protection ◽  
Novel Transcript

The gene AML1/PEBP2 alphaB encodes the alpha subunit of transcription factor PEBP2/CBF and is essential for the establishment of fetal liver hematopoiesis. Rearrangements of AML1 are frequently associated with several types of human leukemia. Three types of AML1 cDNA isoforms have been described to date; they have been designated AML1a, AML1b, and AML1c. All of these isoforms encode the conserved-Runt domain, which harbors the DNA binding and heterodimerization activities. We have identified a new isoform of the AML1 transcript, termed AML1 deltaN, in which exon 1 is directly connected to exon 4 by alternative splicing. The AML1 deltaN transcript was detected in various hematopoietic cell lines of lymphoid to myeloid cell origin, as revealed by RNase protection and reverse transcriptase PCR analyses. The protein product of AML1 deltaN lacks the N-terminal region of AML1, including half of the Runt domain, and neither binds to DNA nor heterodimerizes with the beta subunit. However, AML1 deltaN was found to interfere with the transactivation activity of PEBP2, and the molecular region responsible for this activity was identified. Stable expression of AML1 deltaN in 32Dcl3 myeloid cells blocked granulocytic differentiation in response to granulocyte colony-stimulating factor. These results suggest that AML1 deltaN acts as a modulator of AML1 function and serves as a useful tool to dissect the functional domains in the C-terminal region of AML1.

Abnormal myocyte Ca2+homeostasis in rabbits with pacing-induced heart failure

1998 ◽  
Vol 275 (4) ◽  
pp. H1441-H1448 ◽  
Author(s):  
Atsushi Yao ◽  
Zhi Su ◽  
Akihiko Nonaka ◽  
Iram Zubair ◽  
Kenneth W. Spitzer ◽  
...  
Keyword(s):  
Heart Failure ◽  
Ventricular Myocytes ◽  
Rnase Protection Assay ◽  
Protection Assay ◽  
Rnase Protection ◽  
Contraction Coupling ◽  
Intracellular Ca ◽  
Rapid Pacing ◽  
Sodium Binding ◽  
The Times

To determine whether there are abnormalities in myocyte excitation-contraction coupling and intracellular Ca2+concentration ([Ca2+]i) homeostasis in pacing-induced heart failure (PF), we measured L-type Ca2+ current ( I Ca,L) and Na+/Ca2+exchanger current ( I Na/Ca) with voltage clamp and measured intracellular Na+ concentration ([Na+]i) and [Ca2+]iwith the use of sodium-binding benzofuran isophthalate (SBFI) and fluo 3 in ventricular myocytes isolated from control and paced rabbits. The peak systolic and diastolic levels and the amplitude of electrically stimulated [Ca2+]itransients (0.25 Hz, extracellular Ca2+ concentration = 1.08 mM) were significantly less in PF myocytes. Also, there was prolongation of the times to peak and decline of [Ca2+]itransients. I Ca,Ldensity was markedly decreased in PF myocytes. I Na/Ca at −40 mV elicited by rapid exposure to 0 Na+ solution with a rapid solution switcher was significantly reduced in PF myocytes, suggesting that the function of the Na+/Ca2+exchanger is impaired in these myocytes. In PF myocytes the decline of the [Ca2+]itransient when the Na+/Ca2+exchanger was abruptly disabled was markedly prolonged compared with the decline in control myocytes, consistent with depressed sarcoplasmic reticulum (SR) Ca2+-ATPase function. RNase protection assay showed decreased levels of Na+/Ca2+exchanger and SR Ca2+-ATPase mRNA in PF hearts, consistent with the function studies. We conclude that the functions of L-type Ca2+channels, Na+/Ca2+exchanger, and SR Ca2+-ATPase are impaired in myocytes from rabbit hearts with failure induced by rapid pacing. These abnormalities result in reduced [Ca2+]itransients and systolic and diastolic dysfunction and appear to account for the abnormal ventricular function observed.

scholarly journals Coordinated changes in hepatic amino acid metabolism and endocrine signals support hepatic glucose production during fetal hypoglycemia

2015 ◽  
Vol 308 (4) ◽  
pp. E306-E314 ◽  
Author(s):  
Satya S. Houin ◽  
Paul J. Rozance ◽  
Laura D. Brown ◽  
William W. Hay ◽  
Randall B. Wilkening ◽  
...  
Keyword(s):  
Fetal Liver ◽  
Hepatic Glucose Production ◽  
Plasma Concentrations ◽  
Glucose Production ◽  
Late Gestation ◽  
Fetal Sheep ◽  
Fetal Plasma ◽  
Hepatic Glucose ◽  
Molar Percent ◽  
Glucose Output

Reduced fetal glucose supply, induced experimentally or as a result of placental insufficiency, produces an early activation of fetal glucose production. The mechanisms and substrates used to fuel this increased glucose production rate remain unknown. We hypothesized that in response to hypoglycemia, induced experimentally with maternal insulin infusion, the fetal liver would increase uptake of lactate and amino acids (AA), which would combine with hormonal signals to support hepatic glucose production. To test this hypothesis, metabolic studies were done in six late gestation fetal sheep to measure hepatic glucose and substrate flux before (basal) and after [days (d)1 and 4] the start of hypoglycemia. Maternal and fetal glucose concentrations decreased by 50% on d1 and d4 ( P < 0.05). The liver transitioned from net glucose uptake (basal, 5.1 ± 1.5 μmol/min) to output by d4 (2.8 ± 1.4 μmol/min; P < 0.05 vs. basal). The [U-13C]glucose tracer molar percent excess ratio across the liver decreased over the same period (basal: 0.98 ± 0.01, vs. d4: 0.89 ± 0.01, P < 0.05). Total hepatic AA uptake, but not lactate or pyruvate uptake, increased by threefold on d1 ( P < 0.05) and remained elevated throughout the study. This AA uptake was driven largely by decreased glutamate output and increased glycine uptake. Fetal plasma concentrations of insulin were 50% lower, while cortisol and glucagon concentrations increased 56 and 86% during hypoglycemia ( P < 0.05 for basal vs. d4). Thus increased hepatic AA uptake, rather than pyruvate or lactate uptake, and decreased fetal plasma insulin and increased cortisol and glucagon concentrations occur simultaneously with increased fetal hepatic glucose output in response to fetal hypoglycemia.

RNase Protection Assay

2019 ◽  
pp. 109-116
Author(s):  
Jianzhu Zhao ◽  
Jun Tang ◽  
Justin Elfman ◽  
Hui Li
Keyword(s):  
Rnase Protection Assay ◽  
Protection Assay ◽  
Rnase Protection

The mCK-5 Multiprobe RNase Protection Assay Kit Can Yield Erroneous Results for the Murine Chemokines IP-10 and MCP-1

2000 ◽  
Vol 286 (2) ◽  
pp. 193-197 ◽  
Author(s):  
Bruno Luckow ◽  
Holger Maier ◽  
Silvia Chilla ◽  
Guillermo Pérez de Lema
Keyword(s):  
Rnase Protection Assay ◽  
Protection Assay ◽  
Rnase Protection

scholarly journals Amniotic IGF-I supplementation of growth-restricted fetal sheep alters IGF-I and IGF receptor type 1 mRNA and protein levels in placental and fetal tissues

2005 ◽  
Vol 186 (1) ◽  
pp. 145-155 ◽  
Author(s):  
S Shaikh ◽  
F H Bloomfield ◽  
M K Bauer ◽  
H H Phua ◽  
R S Gilmour ◽  
...  
Keyword(s):  
Late Gestation ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Igf Binding Proteins ◽  
Fetal Sheep ◽  
Hepatic Expression ◽  
Protein Levels ◽  
Igf I ◽  
Igf Receptor

We have previously reported that chronic intra-amniotic supplementation of the late gestation growth-restricted (IUGR) ovine fetus with IGF-I (20 μg/day) increased gut growth but reduced liver weight and circulating IGF-I concentrations. Here we report mRNA and protein levels of IGF-I, the type 1 IGF receptor (IGF-1R) and IGF-binding proteins (IGFBP)-1, -2 and -3 in fetal gut, liver, muscle and placenta from fetuses in that earlier study in an attempt to explain these contrasting results. mRNA and protein were extracted from tissues obtained at post mortem at 131 days of gestation (term, 145 days) from three groups of fetuses (control, IUGR+saline and IUGR+IGF-I, n=9 per group). Control fetuses were unembolised and untreated. In the IUGR groups, growth restriction was induced from 113 to 120 days by placental embolisation; from 120 to 130 days fetuses were treated with daily intra-amniotic injections of either saline or 20 μg IGF-I. mRNA was measured by RT-PCR or real-time RT-PCR, and protein by Western blot. In liver, muscle and placenta, IGF-I mRNA and protein levels were reduced by between 8 and 30% in IGF-I-treated fetuses compared with saline-treated fetuses and controls with no change in IGF-1R mRNA or protein levels. In contrast, in the gut, IGF-I mRNA and protein levels were not significantly altered with IGF-I treatment, but IGF-1R levels were increased, especially in the jejunum. Immunolocalisation demonstrated that IGF-1R expression was confined to the luminal aspect of the gut. mRNA levels of all three IGFBPs were reduced in the gut of IGF-I-treated fetuses, but hepatic expression was significantly increased. These data demonstrated tissue-specific regulation of IGF-I, IGF-1R and IGFBPs-1, -2 and -3 in response to intra-amniotic IGF-I supplementation, though the underlying mechanisms remain obscure.

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