scholarly journals A Study on the Transcriptional Profile of NOS2 and IFN-γ Genes in River Buffalo with Endometritis

2021 ◽  
pp. 32-38
Author(s):  
Othman E. Othman ◽  
Noha M. Osman ◽  
Nadia A. Abo El-Maaty ◽  
Eman R. Mahfouz
Keyword(s):  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
River Buffalo ◽  
Quantitative Real Time Pcr ◽  
Uterine Tissue ◽  
Uterine Lumen ◽  
Ifn Γ ◽  
Nos2 Expression ◽  
Apparently Healthy

Background and Aim: Uterine lumen contamination with bacteria is ubiquitous in buffalo after parturition. Nearly one-third of these infected animals develop endometritis which leads to reduced fertility. The present study aimed to evaluate the expressions of IFN-γ and NOS2 genes in uterine tissue of buffaloes with endometritis and comparing them with those in healthy animals using RT-qPCR Materials and Methods: Uterine samples were collected from 50 apparently healthy and 50 clinically infected buffaloes. RNA was extracted from the collected buffalo's uteri and cDNA was synthesized from extracted RNA. Quantitative Real Time PCR technique was performed using this synthesized cDNA. Results: Apparent up-regulation of both genes mRNA expression was recorded in endometritis-infected animals with 8.3-folds for IFN-γ and 9.99-folds for NOS2 (P<0.001). Conclusion: The upregulation of IFN-γ and NOS2 expression in the uterine tissue of endometritis-infected buffaloes can be used as a scale for measuring the efficiency of drugs used for endometritis treatment.

p29ING4 Regulates the Production of Pro-Angiogenic Molecules by Human Myeloma Cells in Normoxic and Hypoxic Conditions Being Involved in Myeloma-Induced Angiogenesis.

Blood ◽  
2006 ◽  
Vol 108 (11) ◽  
pp. 515-515
Author(s):  
Sara Tagliaferri ◽  
Francesca Morandi ◽  
Paolo Lunghi ◽  
Simona Colla ◽  
Mirca Lazzaretti ◽  
...  
Keyword(s):  
Tumor Suppressor ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Interleukin 8 ◽  
Mrna Levels ◽  
Quantitative Real Time Pcr ◽  
Angiogenic Switch ◽  
Hypoxic Conditions

Abstract Multiple myeloma (MM) cells produce several angiogenic molecules as VEGF, Angiopoietin-1 (Ang-1), interleukin-8 (IL-8) and osteopontin (OPN), however the molecular mechanisms underlying the angiogenic switch are not completely elucidated. The candidate tumor suppressor gene inhibitor of growth family member 4 (p29ING4) has been recently implicated in solid tumors as a repressor of angiogenesis and tumor growth through the suppression of angiogenic related molecules including interleukin-8 (IL-8) and the hypoxia inducible factor (HIF)-1 alpha. In this study we investigate the potential involvement of p29ING4 in the angiogenic switch in MM. First using quantitative real time PCR we compared p29ING4 with VEGF, Ang-1, IL-8 and OPN mRNA levels in eight human myeloma cell lines (HMCLs). A significantly negative correlation was observed between ING4 and IL-8 and a trend of correlation with OPN. Following we transfected HMCLs JJN3, OPM-2 and RPMI-8226 with specific siRNA to completely block the expression of p29ING4 checking the effect on the expression and production of the myeloma-related angiogenic molecules VEGF, Ang-1, IL-8 and OPN by quantitative real time PCR and ELISA assay. p29ING4 suppression in HMCLs did not affect VEGF and Ang-1 production but induced a strong up-regulation of IL-8 mRNA and IL-8 protein secretion. Similarly an induction of OPN mRNA expression as well as OPN secretion was induced by siRNA anti-p29ING4. Moreover conditioned media of HMCLs transfected with siRNA anti p29ING4 significantly increased vessel formation in an experimental in vitro model of angiogenesis (ANGIO kit) as compared to controls. Further we investigate the role of p29ING4 in the production of angiogenic molecule by MM cells in hypoxic condition compared to normoxic one as well as its potential relationship with HIF-1alpha system. Hypoxia induced HIF-1alpha expression at nuclear level and its activity in HMCLs and p29ING4 suppression by siRNA further induced HIF-1alpha transcriptional activity with an increase of its target gene Nip-3 in HMCLs. In turn the block of HIF1-alpha by specific siRNA up-regulated p29ING4 and suppressed IL-8 and OPN mRNA expression by HMCLs suggesting a relationship between p29ING4 and HIF-1alpha activity. These in vitro observations have been extended in vivo by the finding of a significant correlation between bone marrow (BM) plasma IL-8 levels and p29ING4 mRNA expression in purified MM cells obtained from 40 newly diagnosed MM patients (R=−0.58 Spearman’s 2-tailed test: p=0.04), consistently MM patients with higher BM IL-8 levels have a significantly lower p29ING4 mRNA levels. Similarly MM patients positive for OPN expression with high OPN BM levels had a significant lower p29ING4 levels (p=0.02). Finally we found that MM patients with high microvalscular density (MVD&gt;30) have significant lower p29ING4 levels as compared to those with low MVD (&lt;30) (p=0.04) and that MM patients with histological high grade had significant lower p29ING4 mRNA levels (Mann-Whitney 2-tailed: p=0.05). In conclusion, our data indicate that the tumor suppressor p29ING4 regulate the production of angiogenic molecules by MM cells both in normoxic and hypoxic conditions being involved in MM-induced angiogenesis and potentially in tumor progression.

Quantification of C13orf19/P38IP mRNA expression by quantitative real-time PCR in patients with urological malignancies

2005 ◽  
Vol 225 (2) ◽  
pp. 253-260 ◽  
Author(s):  
Uta Schmidt ◽  
Susanne Fuessel ◽  
Michael Haase ◽  
Kai Kraemer ◽  
Axel Meye ◽  
...  
Keyword(s):  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Real Time Pcr ◽  
Urological Malignancies

scholarly journals Biological evaluation of transdichloridoplatinum( II) complexes with 3- and 4-acetylpyridine in comparison to cisplatin

2013 ◽  
Vol 47 (4) ◽  
pp. 346-357 ◽  
Author(s):  
Lana Filipovic ◽  
Sandra Arandelovic ◽  
Nevenka Gligorijevic ◽  
Ana Krivokuca ◽  
Radmila Jankovic ◽  
...  
Keyword(s):  
Western Blot ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Inductively Coupled Plasma ◽  
Gelatin Zymography ◽  
Biological Evaluation ◽  
Emission Spectrometry ◽  
Quantitative Real Time Pcr

Abstract Background. In our previous study we reported the synthesis and cytotoxicity of two trans-platinum(II) complexes: trans-[PtCl2(3-acetylpyridine)2] (1) and trans-[PtCl2(4-acetylpyridine)2] (2), revealing significant cytotoxic potential of 2. In order to evaluate the mechanism underlying biological activity of both trans-Pt(II) isomers, comparative studies versus cisplatin were performed in HeLa, MRC-5 and MS1 cells. Materials and methods. The cytotoxic activity of the investigated complexes was determined using SRB assay. The colagenolytic activity was determined using gelatin zymography, while the effect of platinum complexes on matrix metalloproteinases 2 and 9 mRNA expression was evaluated by quantitative real-time PCR. Apoptotic potential and cell cycle alterations were determined by FACS analyses. Western blot analysis was used to evaluate the effect on expression of DNA-repair enzyme ERCC1, and quantitative real-time PCR was used for the ERCC1 mRNA expression analysis. In vitro antiangiogenic potential was determined by tube formation assay. Platinum content in intracellular DNA and proteins was determined by inductively coupled plasma-optical emission spectrometry. Results. Compound 2 displayed an apparent cytoselective profile, and flow cytometry analysis in HeLa cells indicated that 2 exerted antiproliferative effect through apoptosis induction, while 1 induced both apoptosis and necrosis. Action of 1 and 2, as analyzed by quantitative real-time PCR and Western blot, was associated with down-regulation of ERCC1. Both trans-complexes inhibited MMP-9 mRNA expression in HeLa, while 2 significantly abrogated in vitro tubulogenesis in MS1 cells. Conclusions. The ability of 2 to induce multiple and selective in vitro cytotoxic effects encourages further investigations of trans-platinum(II) complexes with substituted pyridines.

scholarly journals Simultaneous Quantitative Detection of Relevant Biomarkers in Breast Cancer by Quantitative Real-Time PCR

2006 ◽  
Vol 21 (1) ◽  
pp. 30-39 ◽  
Author(s):  
M. Labuhn ◽  
V. Vuaroqueaux ◽  
F. Fina ◽  
A. Schaller ◽  
I. Nanni-Metellus ◽  
...  
Keyword(s):  
Breast Cancer ◽  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Detection ◽  
Mrna Levels ◽  
Quantitative Real Time Pcr ◽  
Expression Levels ◽  
Qrt Pcr ◽  
Mrna Expression Levels

The assessment of ERα, PgR and HER2 status is routinely performed today to determine the endocrine responsiveness of breast cancer samples. Such determination is usually accomplished by means of immunohistochemistry and in case of HER2 amplification by means of fluorescent in situ hybridization (FISH). The analysis of these markers can be improved by simultaneous measurements using quantitative real-time PCR (Qrt-PCR). In this study we compared Qrt-PCR results for the assessment of mRNA levels of ERα, PgR, and the members of the human epidermal growth factor receptor family, HER1, HER2, HER3 and HER4. The results were obtained in two independent laboratories using two different methods, SYBR Green I and TaqMan probes, and different primers. By linear regression we demonstrated a good concordance for all six markers. The quantitative mRNA expression levels of ERα, PgR and HER2 also strongly correlated with the respective quantitative protein expression levels prospectively detected by EIA in both laboratories. In addition, HER2 mRNA expression levels correlated well with gene amplification detected by FISH in the same biopsies. Our results indicate that both Qrt-PCR methods were robust and sensitive tools for routine diagnostics and consistent with standard methodologies. The developed simultaneous assessment of several biomarkers is fast and labor effective and allows optimization of the clinical decision-making process in breast cancer tissue and/or core biopsies.

scholarly journals Leptin in Whales: Validation and Measurement of mRNA Expression by Absolute Quantitative Real-Time PCR

PLoS ONE ◽  
2013 ◽  
Vol 8 (1) ◽  
pp. e54277 ◽  
Author(s):  
Hope C. Ball ◽  
Robert K. Holmes ◽  
Richard L. Londraville ◽  
Johannes G. M. Thewissen ◽  
Robert Joel Duff
Keyword(s):  
Mrna Expression ◽  
Real Time ◽  
Real Time Pcr ◽  
Quantitative Real Time Pcr

scholarly journals KLK11 promotes the activation of mTOR and protein synthesis to facilitate cardiac hypertrophy

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Yi Wang ◽  
Hongjuan Liao ◽  
Yueheng Wang ◽  
Jinlin Zhou ◽  
Feng Wang ◽  
...  
Keyword(s):  
Protein Synthesis ◽  
Angiotensin Ii ◽  
Cardiac Hypertrophy ◽  
Western Blot ◽  
Real Time ◽  
Real Time Pcr ◽  
Mtor Signaling ◽  
Heart Weight ◽  
Cardiomyocyte Hypertrophy ◽  
Quantitative Real Time Pcr

Abstract Background Cardiovascular diseases have become the leading cause of death worldwide, and cardiac hypertrophy is the core mechanism underlying cardiac defect and heart failure. However, the underlying mechanisms of cardiac hypertrophy are not fully understood. Here we investigated the roles of Kallikrein 11 (KLK11) in cardiac hypertrophy. Methods Human and mouse hypertrophic heart tissues were used to determine the expression of KLK11 with quantitative real-time PCR and western blot. Mouse cardiac hypertrophy was induced by transverse aortic constriction (TAC), and cardiomyocyte hypertrophy was induced by angiotensin II. Cardiac function was analyzed by echocardiography. The signaling pathway was analyzed by western blot. Protein synthesis was monitored by the incorporation of [3H]-leucine. Gene expression was analyzed by quantitative real-time PCR. Results The mRNA and protein levels of KLK11 were upregulated in human hypertrophic hearts. We also induced cardiac hypertrophy in mice and observed the upregulation of KLK11 in hypertrophic hearts. Our in vitro experiments demonstrated that KLK11 overexpression promoted whereas KLK11 knockdown repressed cardiomyocytes hypertrophy induced by angiotensin II, as evidenced by cardiomyocyte size and the expression of hypertrophy-related fetal genes. Besides, we knocked down KLK11 expression in mouse hearts with adeno-associated virus 9. Knockdown of KLK11 in mouse hearts inhibited TAC-induced decline in fraction shortening and ejection fraction, reduced the increase in heart weight, cardiomyocyte size, and expression of hypertrophic fetal genes. We also observed that KLK11 promoted protein synthesis, the key feature of cardiomyocyte hypertrophy, by regulating the pivotal machines S6K1 and 4EBP1. Mechanism study demonstrated that KLK11 promoted the activation of AKT-mTOR signaling to promote S6K1 and 4EBP1 pathway and protein synthesis. Repression of mTOR with rapamycin blocked the effects of KLK11 on S6K1 and 4EBP1 as well as protein synthesis. Besides, rapamycin treatment blocked the roles of KLK11 in the regulation of cardiomyocyte hypertrophy. Conclusions Our findings demonstrated that KLK11 promoted cardiomyocyte hypertrophy by activating AKT-mTOR signaling to promote protein synthesis.

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