James V. Gruber1, Venkat Padyachi1, Lisa Lods1, Michele Benhaim2, Robert Holtz3, Audrey Natalizio4. 1Arch Personal Care Products, South Plainfield, NJ, United States; 2Arch Chemicals, Paris, France; 3BioInnovation Laboratories, Inc., McKinney, TX, United States; 4Dermscan Group, Lyon, France.

This study by researchers at Arch Personal Care Products, Arch Chemicals, Bioinnovation Laboratories, Inc., and Dermscan Group demonstrated how MatTek’s EpiDermFT full-thickness in vitro human skin tissue equivalent can be used to measure the ability of a topically-applied formulation to prevent skin damage by inhibiting the expression of nitric oxide free radicals in human (dermal layer) fibroblasts. Recently it was reported that extracts of soybean symbiosomes (root nodules) might have a controlling effect on the formation of nitric oxide free radicals in human fibroblasts. A molecule of particular interest in this area was a plant-based human myoglobin mimic called leghemoglobin. In this early work, leghemoglobin was shown to bind nitric oxide in a chemical assay employing a nitric oxide donor called Spermine NONOate. In addition, human fibroblasts exposed to the extract from soybean symbiosomes where shown to have reduced amounts of nitric oxide formation under standard growth conditions. Researchers from Arch Personal Care Products, Arch Chemicals, Bioinnovation Laboratories, Inc., and Dermscan Group report here on a more complex in vitro study conducted on a new tissue model available from MatTek Corporation called EpiDermFT EFT-200. This tissue model is reported to have an improved stratum corneum, and a more well-defined dermal and epidermal junction. Methods: Standard in vitro tissue growth conditions were employed in this study. Prior to treatment of the tissue substrate with the test materials, the tissue was irradiated with 2.0 MED of either UVA or UVB irradiation using an EL Series Ultraviolet light source (Ultra Violet Products). The tissue samples were treated for 12 hours with the test materials added immediately after the UV irradiation dose. The resulting tissue specimens were tested for their viability using a standard MTT assay. Nitric oxide concentrations were tested using the Griess reagent. Prostaglandin E2 (PGE2) was tested using a standard ELISA technique. Tumor Necrosis Factor Alpha (TNF-á) was tested using recombinant human TNF-á standards using ELISA techniques. Active Matrix Metalloproteinase-1 (MMP-1) levels were measured using ELISA techniques. In vivo testing was done on five volunteers who had signed consent forms prior to the testing. The subjects were irradiated using a xenon lamp with a short arc (Suntest/Original Hanau) at 1 MED to initiate sub-clinical (actinic) erythema. Test samples were applied a four times: 1) Just prior to irradiation (called Preventative), 2) immediately after irradiation (called Curative), and 2 hours after irradiation (called Curative 2 hours). In addition, one site remained untreated and the additional site was treated with the aqueous vehicle used to apply the test product. Sub-clinical erythema was measured using laser Doppler. Results: Employing this model as a test substrate it appears that the extract from soybean symbiosomes influences the production of nitric oxide arising from both UVA and UVB irradiation. Interestingly, however, the extract appears to influence key enzymatic cascades responsible for production of PGE2 and TNF-á as a result of UVB stimulation, but not UVA stimulation while having little influence on the production of radiation induced IL-1á. In addition, the extract appears to reduce production of UVB-induced active MMP-1, but not UVA-induced MMP-1 production. This in vitro study was then preceded by a five-person subclinical erythema study in which erythema was induced by exposure of the skin to UV radiation in the UVA and UVB region. Conclusion: Based on in vitro tissue culture and in vivo microcirculation results taken using a laser Doppler it appears that the extract has the ability to reduce sub-clinical erythema in vivo.


EFT-200, EpiDerm-FT, EpiDermFT, Free-radical biochemistry, Griess reagent, Irradiation, Laser doppler, Leghemoglobin, MMP, MMP-1, MTT, MTT assay, Matrix metalloproteinase-1, Nitric oxide, PGE2, Soybean symbiosome (root noodles), Spermine NONOate, TNF-a, Tumor necrosis factor alpha, UV-Induced, UVA, UVB

Materials Tested

Soybean symbiosome (root noodles)

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