Unique kinetics of Oct3/4 microlocalization following dissociation of human embryonic stem cell colonies

Ichikawa, Hinako; Kanoh, Yoshiya; Shirasawa, Sakiko; Yokoyama, Tetsuji; Yue, Fengming; Tomotsune, Daihachiro; Sasaki, Katsunori

doi:10.1016/j.aanat.2012.05.004

Cited by 5 publications

(2 citation statements)

References 14 publications

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“…Until now, nonviral delivery methods have suffered from poor transfection efficiency with high cell viability or high transfection efficiency with poor cell viability. RhoA GTP signaling pathways are critical for inducing several apoptotic mechanisms in response to unfavorable environmental changes (Ichikawa et al, 2013;Ohgushi et al, 2010). Inhibiting ROCK can reduce apoptosis associated with detachment and freezing protocols (Harb et al, 2008;Ichikawa et al, 2012;Koyanagi et al, 2008;Kurosawa, 2012;Olson, 2008).…”

Section: Resultsmentioning

confidence: 99%

Improving Viability and Transfection Efficiency with Human Umbilical Cord Wharton's Jelly Cells Through Use of a ROCK Inhibitor

Mellott

Godsey²,

Shinogle³

et al. 2014

Cellular Reprogramming

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Differentiating stem cells using gene delivery is a key strategy in tissue engineering and regenerative medicine applications. Nonviral gene delivery bypasses several safety concerns associated with viral gene delivery; however, leading nonviral techniques, such as electroporation, subject cells to high stress and can result in poor cell viabilities. Inhibition of Rho-associated coiled-coil kinase (ROCK) has been shown to mitigate apoptotic mechanisms associated with detachment and freezing of induced pluripotent stem cells and embryonic stem cells; however, inhibiting ROCK in mesenchymal stromal cells (MSCs) for improving gene delivery applications has not been reported previously. In this study, we hypothesized that ROCK Inhibitor (RI) would improve cell viability and gene expression in primary human umbilical cord mesenchymal stromal cells (hUCMSCs) when transfected via Nucleofection™. As hypothesized, the pre-treatment and post-treatment of hUCMSCs transfected via nucleofection with Y-27632-RI significantly improved survival rates of hUCMSCs and gene expression as measured by green fluorescent protein intensity. This study provides the first comparative look at the effect of Y-27632-RI on hUCMSCs that underwent transfection via nucleofection and shows that using Y-27632-RI in concert with nucleofection could greatly enhance the utility of differentiating and reprogramming hUCMSCs for tissue engineering applications.

show abstract

Section: Resultsmentioning

confidence: 99%

Improving Viability and Transfection Efficiency with Human Umbilical Cord Wharton's Jelly Cells Through Use of a ROCK Inhibitor

Mellott

Godsey²,

Shinogle³

et al. 2014

Cellular Reprogramming

View full text Add to dashboard Cite

show abstract

“…For pharmacological interventions, HK-2 cells were pretreated with 1 μM SB203580 (MCE, HY-10256), 1 μM TAK-715 (MCE, HY-10456), or 10 μM tamsulosin 16,17 (Cayman, CAS No. 106463-17-6) for 16 hours before NE/ TGF-β1 incubation for an additional 24 and 48 hours for quantitative reverse transcription polymerase chain reaction (RT-PCR) 18 and western blotting, 19 respectively.…”

Section: Cell Culturementioning

confidence: 99%

Inhibition of α1‐adrenoceptor reduces TGF‐β1‐induced epithelial‐to‐mesenchymal transition and attenuates UUO‐induced renal fibrosis in mice

et al. 2020

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Renal fibrosis is a common pathological hallmark of chronic kidney disease (CKD). Renal sympathetic nerve activity is elevated in patients and experimental animals with CKD and contributes to renal interstitial fibrosis in obstructive nephropathy. However, the mechanisms underlying sympathetic overactivation in renal fibrosis remain unknown. Norepinephrine (NE), the main sympathetic neurotransmitter, was found to promote TGF-β1-induced epithelial-mesenchymal transition (EMT) and fibrotic gene expression in the human renal proximal epithelial cell line HK-2. Using both genetic and pharmacological approaches, we identified that NE binds Gαqcoupled α1-adrenoceptor (α1-AR) to enhance EMT of HK-2 cells by activating p38/ Smad3 signaling. Inhibition of p38 diminished the NE-exaggerated EMT process and increased the fibrotic gene expression in TGF-β1-treated HK-2 cells. Moreover, the pharmacological blockade of α1-AR reduced the kidney injury and renal fibrosis in a unilateral ureteral obstruction mouse model by suppressing EMT in the kidneys. Thus, sympathetic overactivation facilitates EMT of renal epithelial cells and fibrosis via the α1-AR/p38/Smad3 signaling pathway, and α1-AR inhibition may be a promising approach toward treating renal fibrosis.

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Pathway of Maternal Serotonin to the Human Embryo and Fetus

et al. 2018

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Serotonin [5-hydroxytryptamine (5-HT)] is essential to intrauterine development, but its source is debated. We used immunocytochemistry to gauge 5-HT, its biosynthetic enzyme tryptophan hydroxylase 1 (TPH1); an importer (serotonin transporter, 5-HTT/SERT/SLC6A); other transporters [P-glycoprotein 1 (P-gp/ABCB1), OCT3/SLC22A3, and gap junction connexin-43]; and the 5-HT degradative enzyme monoamine oxidase A (MAOA) in sections of placentas. In humans, 5-HT was faintly stained only in first-trimester trophoblasts, whereas TPH1 was not seen at any stage. SERT was expressed in syncytiotrophoblasts and, more strongly, in cytotrophoblasts. MAOA was prominent in syncytiotrophoblasts, OCT3 and gap junctions were stained in cytotrophoblasts, and P-gp was present at the apical surfaces of both epithelia. 5-HT added to cultured placental explants accumulated in the trophoblast epithelium and reached the villus core vessels. Trophoblast uptake was blocked by the SERT inhibitor escitalopram. Inhibition of gap junctions with heptanol prevented the accumulation of 5-HT in cytotrophoblasts, whereas blocking OCT3 with decynium-22 and P-gp with mitotane led to its accumulation in cytotrophoblasts. Reducing 5-HT destruction by inhibiting MAOA with clorgyline increased the accumulation of 5-HT throughout the villus. In the mouse fetus, intravascular platelets stained prominently for 5-HT at day 13.5, whereas the placenta and yolk sac endoderm were both negative. TPH1 was not detected, but SERT was prominent in these mouse tissues. We conclude that serotonin is conveyed from the maternal blood stream through syncytiotrophoblasts, cytotrophoblasts and the villus core to the fetus through a physiological pathway that involves at least SERT, gap junctions, P-gp, OCT3, and MAOA.

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Unique kinetics of Oct3/4 microlocalization following dissociation of human embryonic stem cell colonies

Cited by 5 publications

References 14 publications

Improving Viability and Transfection Efficiency with Human Umbilical Cord Wharton's Jelly Cells Through Use of a ROCK Inhibitor

Improving Viability and Transfection Efficiency with Human Umbilical Cord Wharton's Jelly Cells Through Use of a ROCK Inhibitor

Inhibition of α1‐adrenoceptor reduces TGF‐β1‐induced epithelial‐to‐mesenchymal transition and attenuates UUO‐induced renal fibrosis in mice

Pathway of Maternal Serotonin to the Human Embryo and Fetus

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