Liquiritin reduces lipopolysaccharide-aroused HaCaT cell inflammation damage via regulation of microRNA-31/MyD88

AbstractChronic arsenic exposure causes skin cancer, although the underlying molecular mechanisms are not well defined. Altered microRNA and mRNA expression likely play a pivotal role in carcinogenesis. Changes in genome-wide differential expression of miRNA and mRNA at 3 strategic time points upon chronic sodium arsenite (As3+) exposure were investigated in a well-validated HaCaT cell line model of arsenic-induced cutaneous squamous cell carcinoma (cSCC). Quadruplicate independent HaCaT cell cultures were exposed to 0 or 100 nM As3+ for up to 28-weeks (wk). Cell growth was monitored throughout the course of exposure and epithelial-mesenchymal transition (EMT) was examined employing immunoblot. Differentially expressed miRNA and mRNA profiles were generated at 7, 19, and 28-wk by RNA-seq, followed by identification of differentially expressed mRNA targets of differentially expressed miRNAs through expression pairing at each time point. Pathway analyses were performed for total differentially expressed mRNAs and for the miRNA targeted mRNAs at each time point. RNA-seq predictions were validated by immunoblot of selected target proteins. While the As3+-exposed cells grew slower initially, growth was equal to that of unexposed cells by 19-wk (transformation initiation), and exposed cells subsequently grew faster than passage-matched unexposed cells. As3+-exposed cells had undergone EMT at 28-wk. Pathway analyses demonstrate dysregulation of carcinogenesis-related pathways and networks in a complex coordinated manner at each time point. Immunoblot data largely corroborate RNA-seq predictions in the endoplasmic reticulum stress (ER stress) pathway. This study provides a detailed molecular picture of changes occurring during the arsenic-induced transformation of human keratinocytes.

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Normal keratinization in a spontaneously immortalized aneuploid human keratinocyte cell line.

The Journal of Cell Biology ◽

10.1083/jcb.106.3.761 ◽

1988 ◽

Vol 106 (3) ◽

pp. 761-771 ◽

Cited By ~ 2754

Author(s):

P Boukamp ◽

R T Petrussevska ◽

D Breitkreutz ◽

J Hornung ◽

A Markham ◽

...

Keyword(s):

Epithelial Cell ◽

Cell Line ◽

Human Skin ◽

Cell Lines ◽

Hacat Cell ◽

Epithelial Cell Line ◽

Spontaneous Transformation ◽

Chromosomal Alterations ◽

Hacat Cell Line

In contrast to mouse epidermal cells, human skin keratinocytes are rather resistant to transformation in vitro. Immortalization has been achieved by SV40 but has resulted in cell lines with altered differentiation. We have established a spontaneously transformed human epithelial cell line from adult skin, which maintains full epidermal differentiation capacity. This HaCaT cell line is obviously immortal (greater than 140 passages), has a transformed phenotype in vitro (clonogenic on plastic and in agar) but remains nontumorigenic. Despite the altered and unlimited growth potential, HaCaT cells, similar to normal keratinocytes, reform an orderly structured and differentiated epidermal tissue when transplanted onto nude mice. Differentiation-specific keratins (Nos. 1 and 10) and other markers (involucrin and filaggrin) are expressed and regularly located. Thus, HaCaT is the first permanent epithelial cell line from adult human skin that exhibits normal differentiation and provides a promising tool for studying regulation of keratinization in human cells. On karyotyping this line is aneuploid (initially hypodiploid) with unique stable marker chromosomes indicating monoclonal origin. The identity of the HaCaT line with the tissue of origin was proven by DNA fingerprinting using hypervariable minisatellite probes. This is the first demonstration that the DNA fingerprint pattern is unaffected by long-term cultivation, transformation, and multiple chromosomal alterations, thereby offering a unique possibility for unequivocal identification of human cell lines. The characteristics of the HaCaT cell line clearly document that spontaneous transformation of human adult keratinocytes can occur in vitro and is associated with sequential chromosomal alterations, though not obligatorily linked to major defects in differentiation.

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Achyrocline satureioides (Lam.) DC (Asteraceae) Extract-Loaded Nanoemulsions as a Promising Topical Wound Healing Delivery System: In Vitro Assessments in Human Keratinocytes (HaCaT) and HET-CAM Irritant Potential

Pharmaceutics ◽

10.3390/pharmaceutics13081241 ◽

2021 ◽

Vol 13 (8) ◽

pp. 1241

Author(s):

Lucélia Albarello Balestrin ◽

Tainá Kreutz ◽

Flávia Nathiely Silveira Fachel ◽

Juliana Bidone ◽

Nicolly Espindola Gelsleichter ◽

...

Keyword(s):

Wound Healing ◽

Hacat Cell ◽

Chorioallantoic Membrane ◽

Human Keratinocytes ◽

Scratch Assay ◽

Hen’S Egg ◽

And Migration ◽

Achyrocline Satureioides ◽

Proliferation And Migration

Achyrocline satureioides (Lam.) DC Asteraceae extracts (ASEs) have been investigated for the treatment of various skin disorders. This study reports the effects of ASE-loaded nanoemulsions (NEASE) on the cellular viability, death by necrosis, and migration of immortalized human keratinocytes (HaCaT cell line), as well as the irritant potential through the hen’s egg chorioallantoic membrane test (HET-CAM). NEASE exhibited a polydispersity index above 0.12, with a droplet size of 300 nm, ζ-potential of −40 mV, and content of flavonoids close to 1 mg/mL. No cytotoxicity of the ASE was observed on HaCaT by MTT assay (up to 10 µg/mL). A significant increase of HaCaT viability was observed to NEASE (up to 5 μg/mL of flavonoids), compared to treatment with the ASE. The necrosis death evaluation demonstrated that only NEASE did not lead to cell death at all the tested concentrations. The scratch assay demonstrated that NEASE was able to increase the cell migration at low flavonoid concentrations. Finally, the HET-CAM test proved the non-irritative potential of NEASE. Overall, the results indicate the potential of the proposed formulations for topical use in wound healing, in view of their promising effects on proliferation and migration in keratinocytes, combined with an indication of the absence of cytotoxicity and non-irritating potential.

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Co(II) Complex of Quercetin–Spectral, Anti-/Pro-Oxidant and Cytotoxic Activity in HaCaT Cell Lines

Applied Sciences ◽

10.3390/app11199244 ◽

2021 ◽

Vol 11 (19) ◽

pp. 9244

Author(s):

Monika Kalinowska ◽

Hanna Lewandowska ◽

Marek Pruszyński ◽

Grzegorz Świderski ◽

Ewelina Gołębiewska ◽

...

Keyword(s):

Antioxidant Activity ◽

Solid State ◽

Cytotoxic Activity ◽

General Formula ◽

Cell Lines ◽

Elemental Analysis ◽

Hacat Cell ◽

Water Solution ◽

Ft Ir ◽

Hacat Cell Lines

In this study a cobalt(II) complex of quercetin was synthetized in the solid state with the general formula Co(C15H9O7)2∙2H2O. The FT-IR, elemental analysis, and UV/Vis methods were used to study the composition of the complex in a solid state and in a water solution. The anti-/pro-oxidant activity of quercetin and the Co(II) complex was studied by means of spectrophotometric DPPH (2,2-diphenyl-1-picrylhydrazyl), FRAP (ferric reducing antioxidant activity) and Trolox oxidation assays. The cytotoxicity of quercetin and Co(II)-quercetin complex in HaCat cell lines was then established.

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Bambusae caulisin Liquamen Suppresses the Expression of Thymus and Activation-Regulated Chemokine and Macrophage-Derived Chemokine in Human Keratinocytes due to Antioxidant Effect

Evidence-based Complementary and Alternative Medicine ◽

10.1155/2012/617494 ◽

2012 ◽

Vol 2012 ◽

pp. 1-9 ◽

Cited By ~ 5

Author(s):

Xu-Feng Qi ◽

Dong-Heui Kim ◽

Yang-Suk Yoon ◽

Soon-Bong Song ◽

Yung-Chien Teng ◽

...

Keyword(s):

Atopic Dermatitis ◽

Hacat Cell ◽

Therapeutic Potential ◽

Human Keratinocytes ◽

Antioxidant Effect ◽

Traditional Herbal Medicine ◽

Th2 Chemokines ◽

Ifn Γ ◽

Significant Block ◽

Antioxidant Effects

Bambusae caulisin Liquamen (BCL), traditional herbal medicine used in East Asia, is known to have antioxidative and immune-regulating properties. We hypothesized that the potential antioxidant effects of BCL might suppress the production of thymus and activation-regulated chemokine (TARC) and macrophage-derived chemokine (MDC) in human keratinocytes (HaCaT cell). The immune-regulating effect of BCL was demonstrated by antioxidant capacity using DPPH analysis and DCFH-DA analysis. We found that BCL had strong ROS scavenge effect in HaCaT cell. BCL also showed suppression of IFN-γ-induced expression of TARC and MDC, activation of NF-κB, and, moreover, significant block of IFN-γ-induced degradation and phosphorylation of IκB. However, it had no effects on phosphorylation of p38 MAPK. Collectively, these results suggest that BCL may have a therapeutic potential on skin disease such as atopic dermatitis by inhibiting Th2 chemokines which is due, at least in part, to its antioxidant capacities.

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Bioactivity Determination of a Therapeutic Recombinant Human Keratinocyte Growth Factor by a Validated Cell-based Bioassay

Molecules ◽

10.3390/molecules24040699 ◽

2019 ◽

Vol 24 (4) ◽

pp. 699 ◽

Cited By ~ 2

Author(s):

Wenrong Yao ◽

Ying Guo ◽

Xi Qin ◽

Lei Yu ◽

Xinchang Shi ◽

...

Keyword(s):

Growth Factor ◽

Cell Lines ◽

Hacat Cell ◽

Keratinocyte Growth Factor ◽

Growth Factor Receptor ◽

Luciferase Reporter ◽

Hematopoietic Stem ◽

Human Keratinocyte ◽

Serum Response ◽

Hacat Cell Lines

The therapeutic recombinant human keratinocyte growth factor 1 (rhKGF-1) was approved by the FDA for oral mucositis resulting from hematopoietic stem cell transplantation for hematological malignancies in 2004. However, no recommended bioassay for rhKGF-1 bioactivity has been recorded in the U.S. Pharmacopoeia. In this study, we developed an rhKGF-1-dependent bioassay for determining rhKGF-1 bioactivity based on HEK293 and HaCat cell lines that stably expressed the luciferase reporter driven by the serum response element (SRE) and human fibroblast growth factor receptor (FGFR2) IIIb. A good responsiveness to rhKGF-1 and rhKGF-2 shared by target HEK293/HaCat cell lines was demonstrated. Our stringent validation was completely focused on specificity, linearity, accuracy, precision, and robustness according to the International Council for Harmonization (ICH) Q2 (R1) guidelines, AAPS/FDA Bioanalytical Workshop and the Chinese Pharmacopoeia. We confirmed the reliability of the method in determining rhKGF bioactivity. The validated method is highly timesaving, sensitive, and simple, and is especially valuable for providing information for quality control during the manufacture, research, and development of therapeutic rhKGF.

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