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Low temperature and mTOR inhibition favor stem cell maintenance in human keratinocyte cultures [RNA-seq]

ABSTRACT: Purpose: Comparison of genes expression pattern in normal human epidermal keratinocytes (NHEKs) which were cultured at several specific temperatures or with rapamycin. Methods: Basically, NHEKs were cultured with irradiated 3T3J2 feeder cells in cFAD medium (Pr Green's method) for 1 week. NHEKs were cultured at 32, 35, 36, 37 and 38 degrees C from beginning to end of cell culture, respectively. Furthermore, 100 nM rapamycin-added cFAD medium and ed at 37 degrees C. 7days after cell cuture, NHEKs were harvested for RNAseq analysis. Results: The culture condition at 32 degrees C and 37 degrees C with rapamycin maintained human epidermal keratinocyte stem cells. Conclusions: mTORC1 inhibition via temperature changes favors ex vivo maintenance of human epidermal keratinocyte stem cells.

ORGANISM(S): Homo sapiens

PROVIDER: GSE199522 | GEO | 2023/05/29

REPOSITORIES: GEO

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Low temperature and mTOR inhibition favor stem cell maintenance in human keratinocyte cultures [miRNA-seq]

Project description:Purpose: Comparison of genes expression pattern in normal human epidermal keratinocytes (NHEKs) which were cultured at several specific temperatures or with rapamycin. Methods: Basically, NHEKs were cultured with irradiated 3T3J2 feeder cells in cFAD medium (Pr Green's method) for 1 week. NHEKs were cultured at 32, 35, 36, 37 and 38 degrees C from beginning to end of cell culture, respectively. Furthermore, 100 nM rapamycin-added cFAD medium and ed at 37 degrees C. 7days after cell cuture, NHEKs were harvested for RNAseq analysis. Results: The culture condition at 32 degrees C and 37 degrees C with rapamycin maintained human epidermal keratinocyte stem cells. Conclusions: mTORC1 inhibition via temperature changes favors ex vivo maintenance of human epidermal keratinocyte stem cells.

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IL-37 negatively regulates IL-33 expression in human keratinocytes

Project description:Interleukin (IL)-37 suppresses systemic and local inflammation. It is expressed in the epidermis, the external layer of the skin, and is decreased in inflammatory skin diseases including atopic dermatitis (AD) and psoriasis. Therefore, an agent applied topically on the skin that can increase IL-37 could be promising for treating AD and psoriasis; however, the mechanism regulating IL-37 remains largely unknown. Given that IL-37 expression is induced in differentiated keratinocytes, a major component of the epidermis, and that activation of aryl hydrocarbon receptor (AHR), a ligand-activated transcription factor, promotes keratinocyte differentiation, we hypothesized that AHR might be involved in the IL-37 expression in human keratinocytes. We analyzed normal epidermal human keratinocytes (NHEKs) treated with tapinarof and Galactomyces fermentation filtrate (GFF), which are potent AHR modulators. We found that tapinarof and GFF upregulated IL-37 in NHEKs, which was canceled by the knockdown of AHR using siRNA transfection, indicating that AHR mediates IL-37 expression in NHEKs. Furthermore, we found that the knockdown of IL-37 resulted in the upregulation of IL-33, an alarmin cytokine with crucial roles in the pathogenesis of AD and psoriasis. These findings suggest that IL-37 negatively regulates IL-33 expression in NHEKs. Finally, we examined whether tapinarof and GFF treatment modulates IL-33 expression in NHEKs. Such treatment inhibited IL-33 expression, which was partially reversed by the knockdown of either AHR or IL-37. Taken together, our findings provide the first evidence that tapinarof and GFF could have potential to prevent IL-33-overexpressing disorders such as AD and psoriasis via the AHR/IL-37 axis.

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RNA-Seq whole transcriptome analysis of normal human epidermal keratinocytes and cutaneous squamous cell carcinoma cell lines.

Project description:The incidence of keratinocyte-derived skin cancer, cutaneous squamous cell carcinoma (cSCC) is increasing worldwide making it the second most common metastatic skin cancer. In this project we used SOLiD next generation sequencing to characterize gene expression profiles in normal human epidermal keratinocytes (NHEKs) and cSCC cell lines. Total RNAs from normal human epidermal keratinocytes (NHEKs) (n=4) and cSCC cell lines (n=8) were extracted. The samples were sequenced using SOLiD next generation sequencing.

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Profiling of genes expressed in cultured human keratinocytes established from the bulge region of hair follicles.

Project description:We established a culture method of human keratinocytes from the bulge region of a plucked hair follicle, that contains multipotent epithelial stem cells with high proliferative potential. Using our method, keratinocyte cultures were successfully obtained from all subjects without invasive skin biopsies. We compared the gene expression profiles between the cultured keratinocytes derived from human hair-follicle-bulge (bulgeM-bM-^@M-^Sderived keratinocytes; BDKs) and neonatal human epidermal keratinocytes (NHEKs), and between BDKs from donors with atopic dermatitis and non-atopic controls using microarray analysis. Keywords: expressin profiling Two cell cultures, BDK vs. NHEK cells. 18 BDKs; derived from eighteen healthy volunteers , 6 NHEKs; purchased from Kurabo (Osaka, Japan). One replicate per array.

2013-02-05 | E-GEOD-13709 | biostudies-arrayexpress

CRAC of yeast RNA polymerase II in various thermosensitive strains at permissive and non-permissive temperature and anchor-away strains with the addition of rapamycin.

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2018-01-10 | GSE97913 | GEO

Profiling of genes expressed in cultured human keratinocytes established from the bulge region of hair follicles.

Project description:We established a culture method of human keratinocytes from the bulge region of a plucked hair follicle, that contains multipotent epithelial stem cells with high proliferative potential. Using our method, keratinocyte cultures were successfully obtained from all subjects without invasive skin biopsies. We compared the gene expression profiles between the cultured keratinocytes derived from human hair-follicle-bulge (bulge–derived keratinocytes; BDKs) and neonatal human epidermal keratinocytes (NHEKs), and between BDKs from donors with atopic dermatitis and non-atopic controls using microarray analysis. Keywords: expressin profiling

2013-02-05 | GSE13709 | GEO

RNA-Seq whole transcriptome analysis of normal human epidermal keratinocytes and cutaneous squamous cell carcinoma cell lines.

2015-08-27 | GSE66412 | GEO

RNA sequencing of mechanically intact and degraded murine tail tendon fascicles after exposure to differential temperature and oxygen tension

Project description:Tendon fascicles were extracted from tails of freshly euthanized mice and cultured for 6 days ex vivo in serum containing medium (10%) at either 3% oxygen and 29 degrees celsius or 21% oxygen and 37 degrees celsius.

2020-01-31 | E-MTAB-7832 | biostudies-arrayexpress

Epidermal keratinocytes and IFNg, TNFa and IL1 treatment

Project description:Cultured epidermal keratinocyte controls used for IFNg, TNFa and IL1 treatment. Keywords: other

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RNA sequencing of native and ex vivo cultured murine tail tendon fascicles

Project description:Tendon fascicles were analysed directly after isolation from the tail of freshly euthanized mice and after of after ex vivo culture for 6 days. Fascicles were cultured in either serum containing medium (10%) or in serum-free medium at 21% oxygen and 37 degrees celsius.

2020-09-15 | E-MTAB-9540 | biostudies-arrayexpress

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