Pseudoepidermis, Constructed in vitro, for Use in Toxicological and Pharmacological Studies

Scavarelli-Karantsavelos, R M; Saroya, S Zaman; Vaughan, F. L.; Bernstein, Isadore A.

doi:10.1159/000210858

Cited by 8 publications

(4 citation statements)

References 16 publications

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“…The doses used in this study were 0.01, 1.0 and 10 nmol SM/cm 2 of culture surface. The fact that exposure was indeed topical and not happening via spillage into the underlying medium was confirmed in another study by autoradiographic observation of cultures treated with [14C]SM [11]. The SM was then removed after 30 min by washing with EBSS, following which the cultures were transferred to another petri dish containing fresh growth medium and incubated for the selected post-exposure periods.…”

Section: Methodsmentioning

confidence: 83%

“…In contrast, protein synthesis was unaffected by an exposure of 10 nmol/cm 2 but was significantly inhibited by 50 nmol/cm 2 [3]. Similarly, cells in the germinative basal layer showed slight pathology at 10 nmol/cm 2 and nearly complete destruction after an exposure to 50 nmol/cm 2 [11].…”

Section: Discussionmentioning

confidence: 92%

“…The inhibitory effect on DNA synthesis, presumably a result of alkylation of the nucleic acid by SM [5,15,17], was seen at a lower concentration and sooner than on RNA or protein synthesis. However, each of these synthetic processes was affected at much lower exposure than was necessary to obtain irreversible necrosis of the germinative cell layer in the culture [11] suggesting that inhibition of s.c. DNA synthesis is a specific effect of SM and not a result of general cytotoxicity. Destruction of the epidermal basal layer of cells is a major concomitant of the vesicant response in skin exposed to SM [6,20].…”

Section: Discussionmentioning

confidence: 99%

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The effect of 2,2′-dichlorodiethyl sulfide on DNA synthesis of a murine stratified keratinocyte culture system

Zaman-Saroya

Vaughan

Bernstein

1992

Chemico-Biological Interactions

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A primary stratified keratinocyte culture resembling the epidermis in situ was used as a model for studying the effects of exposure to 2,2'-dichlorodiethyl sulfide, or sulfur mustard (SM), on DNA synthesis. A method that distinguishes between semi-conservative (s.c.) DNA synthesis and repair synthesis was used to determine if the former was inhibited following treatment with SM. In this method the density of the newly synthesized DNA was increased by incorporation of 5-bromo-2-deoxyuridine. Density gradient centrifugation was then used to isolate the heavy DNA for quantification. It was demonstrated that topically applied SM in the dose range of 1-10 nmole/cm2 inhibited s.c. DNA synthesis (replication) in a dose and time related manner. Inhibition of DNA replication by SM would result in inhibition of cell division which must be preceded by s.c. DNA synthesis. This failure to replace damaged germinative cells may lead to the destruction of the basal layer which is observed in vivo and in our epidermal culture following exposure to SM. This may also be related to development of vesication observed in exposed intact human skin.

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Section: Methodsmentioning

confidence: 83%

Section: Discussionmentioning

confidence: 92%

Section: Discussionmentioning

confidence: 99%

See 1 more Smart Citation

The effect of 2,2′-dichlorodiethyl sulfide on DNA synthesis of a murine stratified keratinocyte culture system

Zaman-Saroya

Vaughan

Bernstein

1992

Chemico-Biological Interactions

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“…24 As the cultured dermal layer develops, the fibroblasts and collagen fibrils begin to interact and the dermal layer begins to uniformly contract as a result of collagen polymerization (Figure 1). Cultured dermis can be prepared one of two ways, either as de-epidermized dermis (DED) in which the living epidermis has been removed or as a cultured equivalent consisting of fibroblast cells mixed with collagen.…”

Section: Discussionmentioning

confidence: 99%

Evaluation of Transdermal Penetration Enhancers Using A Novel Skin Alternative

Godwin

Michniak

Creek

1997

Journal of Pharmaceutical Sciences

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A novel alternative to animal skin models was developed in order to aid in the screening of transdermal penetration enhancers. The skin alternative consists of a dermal layer containing human fibroblasts dispersed in a collagen matrix and an epidermal layer of differentiated and stratified human keratinocytes. Skin alternatives were placed in modified Franz diffusion cells (receptor volume 12 mL, donor area 3.14 cm2, n = 5) and enhancer solution (0.4 M in propylene glycol (PG)) was applied. Following 1 h of pretreatment, 10 microL of saturated hydrocortisone (HC) solution in PG was applied, and the cells were occluded with Parafilm. Samples were removed from the receptor compartment over 24 h, replaced with fresh receptor solution, and analyzed for steroid content using HPLC. Skin HC content was also determined. Receptor concentration at 24 h (Q24) for full-thickness skin alternative (control) was 28.6 +/- 13.7 microM and permeability (P) was 8.3 x 10(-4) +/- 5.5 x 10(-4) cm h-1. Azone (1) produced a Q24 of 105.0 +/- 36.1 microM and a P of 11.3 x 10(-4) +/- 1.8 x 10(-4) cm h-1, while the novel penetration enhancer 1-dodecyl-2-pyrrolidinone (2) produced a Q24 of 164.8 +/- 61.2 microM and a P of 33.3 x 10(-4) +/- 6.6 x 10(-4) cm h-1. Compound 3 produced the highest values for all permeability parameters tested with a P of 48.0 +/- 36.8 cm h-1 and a Q24 of 186.1 +/- 45.1 microM. When compared to the control, compound 1 gave an enhancement ratio (ER) of 3.7 for Q24 and 1.4 for P. Compound 2 gives an ER 5.8 for Q24 and 4.0 for P. These enhancement ratios are similar to those found using HC and human skin.

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Assessment of the role of DNA damage and repair in the survival of primary cultures of rat cutaneous keratinocytes exposed to bis(2-chloroethyl)sulfide

Ribeiro

Mitra

Bernstein

1991

Toxicology and Applied Pharmacology

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Toxicity manifests itself as vesication in human skin exposed topically to bis(2-chloroethyl)sulfide (BCES). The destruction of the proliferating population of epidermal cells is a major component of the pathogenic process. Available data strongly suggest that damage to cellular DNA is a critical factor in the loss of these cells. However, the influence of DNA repair on this toxic response has not been adequately studied. Therefore, a study was undertaken to ascertain the influence of DNA repair on the survival of primary monolayer cultures of rat cutaneous keratinocytes exposed to BCES. The sensitive nucleoid sedimentation assay was employed for the determination of DNA damage in cultures exposed to very low levels of BCES. Initial experiments demonstrated that within 1 hr of exposure to as little as 0.1 microM BCES the structural integrity of cellular DNA was compromised, presumably resulting from the appearance of single-strand breaks in the nucleic acid. This same effect was demonstrated in basal cells derived from a stratified, cornified culture grown at the air-liquid interface and exposed topically to the vesicant. Further studies with the monolayer culture demonstrated that the gross structural integrity of the DNA in cells exposed to as much as 5 microM BCES was completely restored within the first 22 hr following the exposure. However, this repair process appeared to be inefficient since a depression of thymidine incorporation into DNA and a significant loss of DNA were exhibited in exposed cultures as long as 72 hr after the initial exposure.

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Pseudoepidermis, Constructed in vitro, for Use in Toxicological and Pharmacological Studies

Cited by 8 publications

References 16 publications

The effect of 2,2′-dichlorodiethyl sulfide on DNA synthesis of a murine stratified keratinocyte culture system

The effect of 2,2′-dichlorodiethyl sulfide on DNA synthesis of a murine stratified keratinocyte culture system

Evaluation of Transdermal Penetration Enhancers Using A Novel Skin Alternative

Assessment of the role of DNA damage and repair in the survival of primary cultures of rat cutaneous keratinocytes exposed to bis(2-chloroethyl)sulfide

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