Cytotoxicity of HBD3 for dendritic cells, normal human epidermal keratinocytes, hTERT keratinocytes, and primary oral gingival epithelial keratinocytes in cell culture conditions

Nattawut Leelakanok; Carol L Fischer; Amber M Bates; Janet M Guthmiller; Georgia K Johnson; Aliasger K Salem; Kim A Brogden; Nicole K Brogden

doi:10.1016/j.toxlet.2015.09.006

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Cytotoxicity of HBD3 for dendritic cells, normal human epidermal keratinocytes, hTERT keratinocytes, and primary oral gingival epithelial keratinocytes in cell culture conditions

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Cytotoxicity of HBD3 for dendritic cells, normal human epidermal keratinocytes, hTERT keratinocytes, and primary oral gingival epithelial keratinocytes in cell culture conditions

Nattawut Leelakanok, Carol L Fischer, Amber M Bates, Janet M Guthmiller, Georgia K Johnson, Aliasger K Salem, Kim A Brogden and Nicole K Brogden

Toxicology letters, Vol.239(2), pp.90-96

12/03/2015

DOI: 10.1016/j.toxlet.2015.09.006

PMCID: PMC4600674

PMID: 26367466

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Abstract

•HBD3 is a very versatile host defense peptide.•It has antimicrobial activities, regulates innate immune mechanisms, and regulates chemokine and cytokine responses.•HBD3 also has cytotoxicity against eukaryotic cells.•We show that the cytotoxicity of HBD3 for DCs, NHEKs, hTERT keratinocytes, and GE keratinocytes varied with the cell type and the culture conditions.•Thus HBD3 has some cytotoxicity, which needs to be considered in future studies of HBD3-modulated chemokine and cytokine responses. Human β-defensin 3 (HBD3) is a prominent host defense peptide. In our recent work, we observed that HBD3 modulates pro-inflammatory agonist-induced chemokine and cytokine responses in human myeloid dendritic cells (DCs), often at 20.0μM concentrations. Since HBD3 can be cytotoxic in some circumstances, it is necessary to assess its cytotoxicity for DCs, normal human epidermal keratinocytes (NHEKs), human telomerase reverse transcriptase (hTERT) keratinocytes, and primary oral gingival epithelial (GE) keratinocytes in different cell culture conditions. Cells, in serum free media with resazurin and in complete media with 10% fetal bovine serum and resazurin, were incubated with 5, 10, 20, and 40μM HBD3. Cytotoxicity was determined by measuring metabolic conversion of resazurin to resorufin. The lethal dose 50 (LD50, mean μM±Std Err) values were determined from the median fluorescent intensities of test concentrations compared to live and killed cell controls. The LD50 value range of HBD3 was 18.2–35.9μM in serum-free media for DCs, NHEKs, hTERT keratinocytes, and GE keratinocytes, and >40.0μM in complete media. Thus, HBD3 was cytotoxic at higher concentrations, which must be considered in future studies of HBD3-modulated chemokine and cytokine responses in vitro.

Defensins

Dendritic cells

HBD3

Cytotoxicity

Epidermal Keratinocytes

hTERT Keratinocytes

Gingival epithelial GE keratinocytes

Details

Title: Subtitle: Cytotoxicity of HBD3 for dendritic cells, normal human epidermal keratinocytes, hTERT keratinocytes, and primary oral gingival epithelial keratinocytes in cell culture conditions
Creators: Nattawut Leelakanok - Division of Pharmaceutics and Translational Therapeutics, Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, The University of Iowa, Iowa City, IA 52242, USA
Carol L Fischer - Dows Institute for Dental Research, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA
Amber M Bates - Dows Institute for Dental Research, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA
Janet M Guthmiller - College of Dentistry, The University of Nebraska Medical Center, Lincoln, NE 68583, USA
Georgia K Johnson - Department of Periodontics, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA
Aliasger K Salem - Division of Pharmaceutics and Translational Therapeutics, Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, The University of Iowa, Iowa City, IA 52242, USA
Kim A Brogden - Dows Institute for Dental Research, College of Dentistry, The University of Iowa, Iowa City, IA 52242, USA
Nicole K Brogden - Division of Pharmaceutics and Translational Therapeutics, Department of Pharmaceutical Sciences and Experimental Therapeutics, College of Pharmacy, The University of Iowa, Iowa City, IA 52242, USA
Resource Type: Journal article
Publication Details: Toxicology letters, Vol.239(2), pp.90-96
DOI: 10.1016/j.toxlet.2015.09.006
PMID: 26367466
PMCID: PMC4600674
NLM abbreviation: Toxicol Lett
ISSN: 0378-4274
eISSN: 1879-3169
Publisher: Elsevier Ireland Ltd
Grant note: DOI: 10.13039/100000072, name: NIH NIDCR, award: R01 DE014390, NIH NIDCR T90 DE02350; DOI: 10.13039/100015515, name: Carver College of Medicine; DOI: 10.13039/100011343, name: Holden Comprehensive Cancer Center; name: Iowa City Veterans Administration Medical Center
Language: English
Date published: 12/03/2015
Academic Unit: Roy J. Carver Department of Biomedical Engineering; Dermatology; Pharmaceutical Sciences and Experimental Therapeutics; Craniofacial Anomalies Research Center; Dental Research; Chemical and Biochemical Engineering; Periodontics
Record Identifier: 9983985832302771

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