Effects of Growth Conditions on the Barrier Properties of a Human Skin Equivalent

Batheja, Priya; Song, Yifan; Wertz, Philip W.; Michniak‐Kohn, Bozena

doi:10.1007/s11095-009-9879-1

Cited by 49 publications

(38 citation statements)

References 31 publications

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“…We also assessed penetration and knockdown efficacy of SNA-NCs in human skin equivalents, given that human skin is known to be thicker and more difficult to penetrate than mouse skin. Human skin equivalents (3D organotypic raft cultures) simulate in vivo human epidermis, including an intact lipid and well-differentiated protein epidermal barrier (29). Given the excellent penetration and knockdown in mouse skin with 50 nM EGFR SNA-NCs, the applied concentration was reduced to 25 nM for testing in human skin equivalents.…”

Section: Resultsmentioning

confidence: 99%

Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation

Zheng

Giljohann

Chen

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

368

288

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Topical application of nucleic acids offers many potential therapeutic advantages for suppressing genes in the skin, and potentially for systemic gene delivery. However, the epidermal barrier typically precludes entry of gene-suppressing therapy unless the barrier is disrupted. We now show that spherical nucleic acid nanoparticle conjugates (SNA-NCs), gold cores surrounded by a dense shell of highly oriented, covalently immobilized siRNA, freely penetrate almost 100% of keratinocytes in vitro, mouse skin, and human epidermis within hours after application. Significantly, these structures can be delivered in a commercial moisturizer or phosphate-buffered saline, and do not require barrier disruption or transfection agents, such as liposomes, peptides, or viruses. SNANCs targeting epidermal growth factor receptor (EGFR), an important gene for epidermal homeostasis, are >100-fold more potent and suppress longer than siRNA delivered with commercial lipid agents in cultured keratinocytes. Topical delivery of 1.5 uM EGFR siRNA (50 nM SNA-NCs) for 3 wk to hairless mouse skin almost completely abolishes EGFR expression, suppresses downstream ERK phosphorylation, and reduces epidermal thickness by almost 40%. Similarly, EGFR mRNA in human skin equivalents is reduced by 52% after 60 h of treatment with 25 nM EGFR SNA-NCs. Treated skin shows no clinical or histological evidence of toxicity. No cytokine activation in mouse blood or tissue samples is observed, and after 3 wk of topical skin treatment, the SNA structures are virtually undetectable in internal organs. SNA conjugates may be promising agents for personalized, topically delivered gene therapy of cutaneous tumors, skin inflammation, and dominant negative genetic skin disorders.T he recent development of small molecule inhibitors and antibodies that target components of signaling pathways has revolutionized the treatment of cancers, inflammatory diseases, and genetic disorders, including those that largely manifest in skin (1-3). These protein-based therapeutics, however, are costly, have limited targeting ability, and, when delivered through traditional intravenous or gastrointestinal routes, can lead to systemic toxicity (4, 5). Topical delivery is particularly attractive for the therapy of skin disorders. However, proteins larger than a few hundred daltons cannot easily enter the skin, and high concentrations of proteins must be applied for a cutaneous effect (6). An alternative to protein-based pathway inhibition involves the blocking and/or degradation of precursor mRNA before translation into protein. Gene silencing leads to down-regulation of protein expression and functions with greater specificity than inhibitors of protein function (7). In fact, targeted gene suppression by antisense DNA and siRNA has shown promising preclinical results, and/or is currently in clinical trials for a variety of diseases, including many forms of cancer (e.g., melanoma, neuroblastoma, and pancreatic adenocarcinoma), genetic disorders, and macular degeneration (8). For gene su...

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Section: Resultsmentioning

confidence: 99%

Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation

Zheng

Giljohann

Chen

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

368

288

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“…The growth conditions of CHE influenced the SC barrier functions. 22) Moreover, skin has a species difference and the skin penetration flux also differs among species. 23) We have to improve the SC barrier function of CHE if we will use the CHE for the skin permeability screening of drugs.…”

Section: Resultsmentioning

confidence: 99%

Prediction of Percutaneous Absorption in Human Using Three-Dimensional Human Cultured Epidermis LabCyte EPI-MODEL

Hikima

Kaneda

Matsuo

et al. 2012

Biological & Pharmaceutical Bulletin

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The objective of this study is to establish a relationship of the skin penetration parameters between the three-dimensional cultured human epidermis LabCyte EPI-MODEL (LabCyte) and hairless mouse (HLM) skin penetration in vitro and to predict the skin penetration and plasma concentration profile in human. The skin penetration experiments through LabCyte and HLM skin were investigated using 19 drugs that have a different molecular weight and lipophilicity. The penetration flux for LabCyte reached 30 times larger at maximum than that for HLM skin. The human data can be estimated from the in silico approach with the diffusion coefficient (D), the partition coefficient (K) and the skin surface concentration (C) of drugs by assuming the bi-layer skin model for both LabCyte and HLM skin. The human skin penetration of β-estradiol, prednisolone, testosterone and ethynylestradiol was well agreed between the simulated profiles and in vitro experimental data. Plasma concentration profiles of β-estradiol in human were also simulated and well agreed with the clinical data. The present alternative method may decrease human or animal skin experiment for in vitro skin penetration.Key words in silico correlation; reconstructed epidermis; LabCyte EPI-MODEL; hairless mouse Human skin permeability of drugs in vivo can be estimated from in vitro experiment using human skin. However, some researchers have ethical and cultural difficulty to get human skins. Thus, animal skins have been widely used for pharmaceutical, toxicological and cosmetic R&D although the skins have a species difference between human. Recently, the 7th Amendment of the EC Cosmetic Directive prohibits the sale of cosmetics and ingredients validated their safety by animal testing with an alternative method. 1) Thus, researchers utilize reconstructed human skin instead of animal skin. The experiments used reconstructed epidermis is approved by the European Centre for the Validation of Alternative Methods (ECVAM) as the scientifically validated methods for skin corrosion and irritation. 2,3) Permeability studies with Episkin ® , EpiDerm TM and SkinEthic ® comparison to human and animal skin were investigated and the authors reported that the penetration flux through the reconstructed skin was higher than that of skin. [4][5][6] The OECD regulations accepted the results from the reconstructed skin 7) but don't approve it as an alternative method for in vitro human skin penetration experiment yet.Skin penetration is widely expressed by the mathematical models on the basis of the process of diffusion and dissolution. Penetration parameters, diffusion coefficient D [cm 2 /s] and partition coefficient K [-], through stratum corneum (SC) and viable skin (VS) of hairless mouse skin are found to well agree with that of human skin. 8) Not only the skin penetration profile but the blood concentration profile in human is well simulated using the penetration parameters determined from hairless mouse skin, the thickness of human SC and VS, the skin enzymatic parameters (first-ord...

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“…For data presentation, mean flux, lag time, and cumulative amount after 24 h were calculated. The flux values of MTZ permeated through the skin membranes into the receptor medium were determined from slopes of plots of concentration in the receptor phase as a function of time and expressed as micrograms per square centimeter per hour using linear regression (Microsoft Excel) (21). The degree of penetration retardation is defined as the retardant ratio (RR), which is calculated from the following equation (22): RR ¼ Flux for the formulation containing retardants=…”

Section: Data and Statistical Analysismentioning

confidence: 99%

Effect of Hydrocarbon Chain Length in 1,2-Alkanediols on Percutaneous Absorption of Metronidazole: Toward Development of a General Vehicle for Controlled Release

Jia

Zhang

et al. 2014

AAPS PharmSciTech

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Abstract. The objective of the present study is to investigate the effect of hydrocarbon chain length in 1,2-alkanediols on percutaneous absorption of metronidazole (MTZ). Twelve formulations (1,2-propanediol, 1,2-butanediol, 1,2-pentanediol, 1,2-hexanediol in 4% concentration, 1,2-hexanediol, and 1,2-heptanediol in 1% concentration, in the absence and presence of 1,4-cyclohexanediol, respectively) were studied in an in vitro hairless mouse skin model using Franz diffusion cell. Based on the flux values and retardation ratios (RR), a penetration retardation effect on percutaneous absorption of MTZ was observed for the formulations containing 1,2-diols having six-to seven-carbon chain in the presence of 1,4-cyclohexanediol (1,2-hexanediol with chain length of six hydrocarbons, RRs are 0.69 and 0.76 in the concentration of 4% and 1%, respectively; 1,2-heptanediol with chain length of seven hydrocarbons, RR is 0.78 in the concentration of 1%). On the other hand, no retardation effect was observed in formulations containing short alkyl chains (RRs of 1,2-propanediol, 1,2-butanediol, and 1,2-pentanediol are 0.99, 1.61, and 0.96, respectively). Instead, a penetration enhancement effect was observed for 1,2-diols having four and five carbons. In other words, effect of 1,2-alkanediols on percutaneous absorption of MTZ can be systematically modulated by simply varying number of -CH 2 groups in the hydrocarbon chain-from being a penetration enhancer to retardant. These observations shed light on mechanism of the penetration enhancement and retardation effect and provide insight into rational design of penetration enhancers and retardants. Furthermore, the combination of 1,2-alkanediols and 1,4-cyclohexanediol could become a general vehicle for controlled release of pharmaceutical and cosmetic active ingredients.

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Effects of Growth Conditions on the Barrier Properties of a Human Skin Equivalent

Abstract: Culture with the growth media additives combination produced a pronounced effect on the permeability barrier of the HSEs. Further validation of permeability with additional agents could comprise the first step toward their use in skin permeability screening.

Cited by 49 publications

References 31 publications

Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation

Topical delivery of siRNA-based spherical nucleic acid nanoparticle conjugates for gene regulation

Prediction of Percutaneous Absorption in Human Using Three-Dimensional Human Cultured Epidermis LabCyte EPI-MODEL

Effect of Hydrocarbon Chain Length in 1,2-Alkanediols on Percutaneous Absorption of Metronidazole: Toward Development of a General Vehicle for Controlled Release

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