scholarly journals TRPM7-Mediated Calcium Transport in HAT-7 Ameloblasts

2021 ◽  
Vol 22 (8) ◽  
pp. 3992
Author(s):  
Kristóf Kádár ◽  
Viktória Juhász ◽  
Anna Földes ◽  
Róbert Rácz ◽  
Yan Zhang ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cell Line ◽  
Calcium Transport ◽  
Potential Role ◽  
Dental Enamel ◽  
Entry Pathway ◽  
Mrna And Protein Expression ◽  
Expression Characteristic ◽  
Transmembrane Currents

TRPM7 plays an important role in cellular Ca2+, Zn2+ and Mg2+ homeostasis. TRPM7 channels are abundantly expressed in ameloblasts and, in the absence of TRPM7, dental enamel is hypomineralized. The potential role of TRPM7 channels in Ca2+ transport during amelogenesis was investigated in the HAT-7 rat ameloblast cell line. The cells showed strong TRPM7 mRNA and protein expression. Characteristic TRPM7 transmembrane currents were observed, which increased in the absence of intracellular Mg2+ ([Mg2+]i), were reduced by elevated [Mg2+]i, and were inhibited by the TRPM7 inhibitors NS8593 and FTY720. Mibefradil evoked similar currents, which were suppressed by elevated [Mg2+]i, reducing extracellular pH stimulated transmembrane currents, which were inhibited by FTY720. Naltriben and mibefradil both evoked Ca2+ influx, which was further enhanced by the acidic intracellular conditions. The SOCE inhibitor BTP2 blocked Ca2+ entry induced by naltriben but not by mibefradil. Thus, in HAT-7 cells, TRPM7 may serves both as a potential modulator of Orai-dependent Ca2+ uptake and as an independent Ca2+ entry pathway sensitive to pH. Therefore, TRPM7 may contribute directly to transepithelial Ca2+ transport in amelogenesis.

scholarly journals 3-Iodothyronamine and 3,5,3′-triiodo-L-thyronine reduce SIRT1 protein expression in the HepG2 cell line

2020 ◽  
Vol 41 (1) ◽  
Author(s):  
Ginevra Sacripanti ◽  
Leonardo Lorenzini ◽  
Lavinia Bandini ◽  
Sabina Frascarelli ◽  
Riccardo Zucchi ◽  
...  
Keyword(s):  
Protein Expression ◽  
Cell Viability ◽  
Cell Line ◽  
Potential Role ◽  
Colorimetric Assay ◽  
Tumor Cell Line ◽  
Rat Hepatocytes ◽  
Carcinoma Cells ◽  
Primary Rat Hepatocytes

AbstractBackground3-Iodothyronamine (T1AM) is an endogenous messenger chemically related to thyroid hormone. Recent results indicate significant transcriptional effects of chronic T1AM administration involving the protein family of sirtuins, which regulate important metabolic pathways and tumor progression. Therefore, the aim of this work was to compare the effect of exogenous T1AM and 3,5,3′-triiodo-L-thyronine (T3) chronic treatment on mammalian sirtuin expression in hepatocellular carcinoma cells (HepG2) and in primary rat hepatocytes at micromolar concentrations.Materials and methodsSirtuin (SIRT) activity and expression were determined using a colorimetric assay and Western blot analysis, respectively, in cells treated for 24 h with 1–20 μM T1AM or T3. In addition, cell viability was evaluated by the MTTtest upon 24 h of treatment with 0.1–20 μM T1AM or T3.ResultsIn HepG2, T1AM significantly reduced SIRT 1 (20 μM) and SIRT4 (10–20 μM) protein expression, while T3 strongly decreased the expression of SIRT1 (20 μM) and SIRT2 (any tested concentration). In primary rat hepatocytes, T3 decreased SIRT2 expression and cellular nicotinamide adenine dinucleotide (NAD) concentration, while on sirtuin activity it showed opposite effects, depending on the evaluated cell fraction. The extent of MTT staining was moderately but significantly reduced by T1AM, particularly in HepG2 cells, whereas T3 reduced cell viability only in the tumor cell line.ConclusionsT1AM and T3 downregulated the expression of sirtuins, mainly SIRT1, in hepatocytes, albeit in different ways. Differences in mechanisms are only observational, and further investigations are required to highlight the potential role of T1AM and T3 in modulating sirtuin expression and, therefore, in regulating cell cycle or tumorigenesis.

GCM2 Silencing in Parathyroid Adenoma is associated with Promoter Hypermethylation and Gain of Methylation on Histone 3

2021 ◽  
Author(s):  
Priyanka Singh ◽  
Sanjay Kumar Bhadada ◽  
Divya Dahiya ◽  
Uma Nahar Saikia ◽  
Ashutosh Kumar Arya ◽  
...  
Keyword(s):  
Protein Expression ◽  
Parathyroid Adenoma ◽  
Dna Methyltransferase ◽  
Promoter Hypermethylation ◽  
Epigenetic Alterations ◽  
Protein Levels ◽  
Histone 3 ◽  
Parathyroid Adenomas ◽  
Mrna And Protein Expression

Abstract Purpose Glial cells missing 2 (GCM2), a zinc finger-transcription factor, is essentially required for the development of parathyroid glands. We sought to identify if the epigenetic alterations in the GCM2 transcription are involved in the pathogenesis of sporadic parathyroid adenoma. In addition, we examined the association between promoter methylation and histone modifications with disease indices. Experimental design mRNA and protein expression of GCM2 were analyzed by RT-qPCR and immunohistochemistry in 33 adenomatous and 10 control parathyroid tissues. DNA methylation and histone methylation/acetylation of GCM2 promoter were measured by bisulfite sequencing and ChIP-qPCR. Additionally, we investigated the role of epigenetic modifications on GCM2 and DNA methyltransferase 1 (DNMT1) expression in PTH-C1 cells by treating with 5-aza 2’deoxycytidine (DAC) and BRD4770 and assessed for GCM2 mRNA and DNMT1 protein levels. Results mRNA and protein expression of GCM2 were lower in sporadic adenomatous than in control parathyroid tissues. This reduction correlated with hypermethylation (P<0.001) and higher H3K9me3 levels in GCM2 promoter (P<0.04) in adenomas. In PTH-C1 cells, DAC treatment resulted in increased GCM2 transcription and decreased DNMT1 protein expression, while cells treated with the BRD4770 showed reduced H3K9me3 levels but a non-significant change in GCM2 transcription. Conclusion These findings suggest the concurrent association of promoter hypermethylation and higher H3K9me3 with the repression of GCM2 expression in parathyroid adenomas. Treatment with DAC restored GCM2 expression in PTH-C1 cells. Our results showed a possible epigenetic landscape in the tumorigenesis of parathyroid adenoma and also that DAC may be promising avenues of research for parathyroid adenoma therapeutics.

Function and repair of dental enamel – Potential role of epithelial transport processes of ameloblasts

Pancreatology ◽  
2015 ◽  
Vol 15 (4) ◽  
pp. S55-S60 ◽  
Author(s):  
Gábor Varga ◽  
Beáta Kerémi ◽  
Erzsébet Bori ◽  
Anna Földes
Keyword(s):  
Potential Role ◽  
Epithelial Transport ◽  
Dental Enamel ◽  
Transport Processes

148 EXPRESSION OF OVIDUCT-SPECIFIC GLYCOPROTEIN IN THE CANINE OVIDUCT DURING THE PERIOVULATORY PERIOD

2014 ◽  
Vol 26 (1) ◽  
pp. 187 ◽  
Author(s):  
C. Marnier ◽  
M. Saint-Dizier ◽  
M. Z. Tahir ◽  
S. Chastant-Maillard ◽  
K. Reynaud
Keyword(s):  
Protein Expression ◽  
Western Blot ◽  
Potential Role ◽  
Germinal Vesicle ◽  
Blastocyst Stage ◽  
Santa Cruz ◽  
Immuno Histochemistry ◽  
Imagej Software ◽  
Mouse Antibody

In the canine species, the oocyte is ovulated at the immature germinal vesicle (GV) stage and will reach metaphase II stage after 3 to 4 days spent in the oviduct. Fertilization and embryonic development to the blastocyst stage also take place in the oviduct. In a previous study (Tahir et al. 2012 Reprod. Domest. Anim. 47, 487), we reported the expression of oviductin (oviduct-specific glycoprotein) mRNA in the oviduct. The present study aimed to describe the oviductin protein expression (immunolocalization and Western blot quantification) and the effect of the oviducal region and the ovarian cycle. Beagle bitches were ovariectomized at 6 stages (6 bitches/stage): anestrus, after the onset of proestrus and before the LH peak (Pre-LH), after the LH peak and before ovulation (Pre-ov), 1 day (Day 1), 4 days (Day 4), and 7 days (Day 7) after ovulation. Three oviducal regions were collected [i.e. ampulla, isthmus, and ampulla-isthmus junction (AIJ)]. Ampulla and isthmus were fixed in paraformaldehyde, embedded in paraffin, and 7-μm sections were used for immuno-histochemistry using a goat polyclonal anti-human oviductin (N20; Santa Cruz Biotechnology, Santa Cruz, CA, USA) and the ImmPress kit (Vector Laboratories, Burlingame, CA, USA). Total protein from the AIJ was extracted and used for Western Blot using a mouse monoclonal anti-mouse antibody (H8; Santa Cruz Biotechnology). The expression of oviductin in AIJ was quantified in duplicate on blots using ImageJ software and normalized with actin levels. Relative amounts of oviductin were compared between stages by ANOVA followed by a Tukey test. Immuno-histochemistry revealed that oviductin was specifically expressed in the nonciliated cells of the oviducal epithelium from Pre-LH to Day 7, with a stronger staining in the isthmus than in the ampulla at all stages. Furthermore, the expression of oviduct-specific glycoprotein, detected by Western Blot, varied significantly with the stage (P < 0.0001). The oviductin protein expression was at its lowest level at anestrus, then increased significantly at Pre-LH and Pre-ov (35- and 41-fold higher levels than anestrus, respectively), reached a maximal level at Day 1 (66-fold higher than anestrus), then decreased at Days 4 and 7 (47- and 20-fold higher than anestrus, respectively). In conclusion, this is the first report of oviductin protein expression in the canine oviduct. The region-specific higher expression of oviductin at Day 1 post-ovulation suggests a potential role of this glycoprotein in gamete maturation and fertilization in the bitch.

scholarly journals Cathepsin V suppresses GATA3 protein expression in luminal A breast cancer

2020 ◽  
Vol 22 (1) ◽  
Author(s):  
Naphannop Sereesongsaeng ◽  
Sara H. McDowell ◽  
James F. Burrows ◽  
Christopher J. Scott ◽  
Roberta E. Burden
Keyword(s):  
Breast Cancer ◽  
Protein Expression ◽  
Cell Line ◽  
Cysteine Protease ◽  
Breast Cancers ◽  
Gata3 Expression ◽  
Er Positive ◽  
Cathepsin V ◽  
Gata3 Protein

Abstract Background Lysosomal cysteine protease cathepsin V has previously been shown to exhibit elevated expression in breast cancer tissue and be associated with distant metastasis. Research has also identified that cathepsin V expression is elevated in tumour tissues from numerous other malignancies, but despite this, there has been limited examination of the function of this protease in cancer. Here we investigate the role of cathepsin V in breast cancer in order to delineate the molecular mechanisms by which this protease contributes to tumourigenesis. Methods Lentiviral transductions were used to generate shRNA cell line models, with cell line validation undertaken using RQ-PCR and Western blotting. Phenotypic changes of tumour cell biology were examined using clonogenic and invasion assays. The relationship between GATA3 expression and cathepsin V was primarily analysed using Western blotting. Site-directed mutagenesis was used to generate catalytic mutant and shRNA-resistant constructs to confirm the role of cathepsin V in regulating GATA3 expression. Results We have identified that elevated cathepsin V expression is associated with reduced survival in ER-positive breast cancers. Cathepsin V regulates the expression of GATA3 in ER-positive breast cancers, through promoting its degradation via the proteasome. We have determined that depletion of cathepsin V results in elevated pAkt-1 and reduced GSK-3β expression, which rescues GATA3 from proteasomal degradation. Conclusions In this study, we have identified that cysteine protease cathepsin V can suppress GATA3 expression in ER-positive breast cancers by facilitating its turnover via the proteasome. Therefore, targeting cathepsin V may represent a potential therapeutic strategy in ER-positive breast cancers, by restoring GATA3 protein expression, which is associated with a more favourable clinical outcome.

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