Proceedings from a clinical roundtable.
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Humans↗
Biomedical subjects
Ken Marenus
Publications and source records attributed to Ken Marenus.
The cytoprotective effects of exogenous DNA fragments.
Ultraviolet irradiation of normal human keratinocytes induces a cytotoxic effect. The chromophore for this effect is believed to be genomic DNA. However, DNA damage is known to be repaired in UVB irradiated keratinocytes. The trigger for this DNA repair is potentially damaged DNA itself. To test the hypothesis that damaged DNA can induce the host cell's own DNA repair mechanism, we treated the keratinocytes with the damaged DNA and evaluated its cytoprotective effects. We have observed that fragmented calf thymus DNA irradiated and damaged with a UVC light can induce a protective effect in cultured human keratinocytes. Keratinocytes treated with UVC damaged DNA fragments are less susceptible to UVB irradiation-induced cell death as measured by neutral red uptake. Unirradiated exogenous DNA did not induce this protective effect. Similar protective effects can be seen with irradiated salmon sperm DNA. UVC damaged DNA fragments induced 60% increase in protection in human HaCaT keratinocyte in culture to the cell death induced by UVB. Similar protection was observed with UVC irradiated oligothymidylic acid (dT3-dT5) which increased the survival of human HaCaT keratinocytes after UVB irradiated by 50%. Isolated mononucleotides, irradiated or not, do not increase UVB survivability. Cellular DNA synthesis was greatly inhibited by UVB, becoming undetectable at 40 mJ/cm(2). Exogenous treatment with damaged fragments causes immediate and significant inhibition of total cellular DNA synthesis. This inhibition was dose dependent. Cells that undergo damage to their DNA are known to inhibit endogenous DNA synthesis via p53 suppressor gene activation. This is believed to allow them sufficient time to repair the host DNA. The cellular response to exogenous damaged DNA may be a similar mechanism.
Animals↗
Evidence and utility of melanin degrading enzymes.
The biochemical synthesis of human melanin is understood in some detail. However, little is known about melanin degradation and catabolism of melanin. We hypothesize that human skin contains enzymes that degrade melanin and these enzymes can be used to reduce skin color. To test this hypothesis, HaCaT keratinocytes and normal human keratinocytes in culture were pulse labeled for one hour with radiolabeled synthetic melanin. This melanin was synthesized in vitro using tyrosinase enzyme from mushrooms and using radiolabeled [14C]3,4-dihydroxyphenylalanine (DOPA) as a substrate. After the initial pulse labeling, samples of both the cells and media were taken at 2, 4, 6 and 18 hours. Over these time periods the counts remaining in the media and cell fraction were significantly decreased. This data suggests the need for new protein synthesis and the lysosome organelle function for the degradation. Melanin degrading extracts isolated from Aspergillus fumigatus and Saccharomyces cerevisiae were applied to human skin. These extracts cause significant reduction in UVB induced pigmentation. These extracts may be useful in developing new whitening products to even skin color and tone.
Aspergillus fumigatus↗
Normal human epidermal keratinocytes treated with 7-dehydrocholesterol express increased levels of heat shock protein.
Human skin, and its isolated cells, respond to insults with a variety of repair and protective mechanisms. One such mechanism is the production of heat shock proteins (HSPs). Heat shock proteins help the other cellular proteins fold correctly into their active three-dimensional structures. Therefore, they can enhance the survival of cells under harsh, denaturing conditions. In order to develop a means of promoting the heat shock response to prepare the skin to withstand insult, we are investigating materials that appear to protect the skin biologically. One such material is vitamin D3 and its precursors. We have observed that keratinocytes treated with 7-dehydrocholesterol (7-DHC), a precursor of vitamin D3, have increased levels of protein and mRNA for heat shock proteins. In addition, we observed that topically applied 7-DHC increases the minimal dose of UVB required to induce erythema. These data suggest that 7-DHC can induce heat shock proteins in skin keratinocytes and that they will be more resistant to UVB insult.
Cells, Cultured↗