Limits...
Mice deficient in involucrin, envoplakin, and periplakin have a defective epidermal barrier.

Sevilla LM, Nachat R, Groot KR, Klement JF, Uitto J, Djian P, Määttä A, Watt FM - J. Cell Biol. (2007)

Bottom Line: Cornified envelopes form but are ultrastructurally abnormal, with reduced lipid content and decreased mechanical integrity.Expression of proteases is reduced and the protease inhibitor, serpina1b, is highly upregulated, resulting in defective filaggrin processing and delayed degradation of desmoglein 1 and corneodesmosin.There is infiltration of CD4+ T cells and a reduction in resident gammadelta+ T cells, reminiscent of atopic dermatitis.

View Article: PubMed Central - PubMed

Affiliation: Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Cambridge CB2 0RE, England, UK.

ABSTRACT
The cornified envelope is assembled from transglutaminase cross-linked proteins and lipids in the outermost epidermal layers and is essential for skin barrier function. Involucrin, envoplakin, and periplakin form the protein scaffold on which the envelope assembles. To examine their combined function, we generated mice deficient in all three genes. The triple knockouts have delayed embryonic barrier formation and postnatal hyperkeratosis (abnormal accumulation of cornified cells) resulting from impaired desquamation. Cornified envelopes form but are ultrastructurally abnormal, with reduced lipid content and decreased mechanical integrity. Expression of proteases is reduced and the protease inhibitor, serpina1b, is highly upregulated, resulting in defective filaggrin processing and delayed degradation of desmoglein 1 and corneodesmosin. There is infiltration of CD4+ T cells and a reduction in resident gammadelta+ T cells, reminiscent of atopic dermatitis. Thus, combined loss of the cornified envelope proteins not only impairs the epidermal barrier, but also changes the composition of T cell subpopulations in the skin.

Show MeSH

Related in: MedlinePlus

Microarray analysis reveals changes in epidermal protease activity. (A) Clustering of microarray data. Note that triple HET samples cluster more closely with WT than with triple KO. (B) Pie graph of genes (grouped according to function) with altered expression in triple KO relative to WT. (C) Semi-quantitative RT-PCR with primers specific for genes identified in the microarray. RT-PCR was performed on RNA isolated from dorsal skin of WT (+/+), triple HET (+/−) and triple KO (−/−) mice. (D) In situ zymography assay demonstrates decreased proteolytic activity, indicated by reduction in green fluorescence, in triple KO epidermis. As a control, addition of the serine protease inhibitor aprotinin reduces fluorescent signal in both WT and triple KO (bottom panels). Note nuclear retention in cornified layers of the triple KO (white arrows). Bar: 50 μm.
© Copyright Policy
Related In: Results  -  Collection


getmorefigures.php?uid=PMC2373502&req=5

fig5: Microarray analysis reveals changes in epidermal protease activity. (A) Clustering of microarray data. Note that triple HET samples cluster more closely with WT than with triple KO. (B) Pie graph of genes (grouped according to function) with altered expression in triple KO relative to WT. (C) Semi-quantitative RT-PCR with primers specific for genes identified in the microarray. RT-PCR was performed on RNA isolated from dorsal skin of WT (+/+), triple HET (+/−) and triple KO (−/−) mice. (D) In situ zymography assay demonstrates decreased proteolytic activity, indicated by reduction in green fluorescence, in triple KO epidermis. As a control, addition of the serine protease inhibitor aprotinin reduces fluorescent signal in both WT and triple KO (bottom panels). Note nuclear retention in cornified layers of the triple KO (white arrows). Bar: 50 μm.

Mentions: Our analysis demonstrated that in the absence of involucrin, envoplakin, and periplakin, the epidermal barrier was abnormal, and suggested that the compensatory mechanism allowing barrier function to be maintained was reduced desquamation. To investigate how this was achieved, microarray analysis was performed using mRNA isolated from dorsal skin of 7-wk-old WT, triple HET, or triple KO female mice (Fig. 5). Skin histology proximal to that used for RNA isolation indicated hyperkeratosis in the triple KO, but not the WT or triple HET (unpublished data). Consistent with their phenotypes, the triple HET samples clustered more closely with those of WT than with triple KO (Fig. 5 A). 207 genes were found to have statistically significant (P < 0.05) changes (twofold or greater; increase or decrease) in expression between WT and triple KO samples (Table S1, available at http://www.jcb.org/cgi/content/full/jcb.200706187/DC1).


Mice deficient in involucrin, envoplakin, and periplakin have a defective epidermal barrier.

Sevilla LM, Nachat R, Groot KR, Klement JF, Uitto J, Djian P, Määttä A, Watt FM - J. Cell Biol. (2007)

Microarray analysis reveals changes in epidermal protease activity. (A) Clustering of microarray data. Note that triple HET samples cluster more closely with WT than with triple KO. (B) Pie graph of genes (grouped according to function) with altered expression in triple KO relative to WT. (C) Semi-quantitative RT-PCR with primers specific for genes identified in the microarray. RT-PCR was performed on RNA isolated from dorsal skin of WT (+/+), triple HET (+/−) and triple KO (−/−) mice. (D) In situ zymography assay demonstrates decreased proteolytic activity, indicated by reduction in green fluorescence, in triple KO epidermis. As a control, addition of the serine protease inhibitor aprotinin reduces fluorescent signal in both WT and triple KO (bottom panels). Note nuclear retention in cornified layers of the triple KO (white arrows). Bar: 50 μm.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2373502&req=5

fig5: Microarray analysis reveals changes in epidermal protease activity. (A) Clustering of microarray data. Note that triple HET samples cluster more closely with WT than with triple KO. (B) Pie graph of genes (grouped according to function) with altered expression in triple KO relative to WT. (C) Semi-quantitative RT-PCR with primers specific for genes identified in the microarray. RT-PCR was performed on RNA isolated from dorsal skin of WT (+/+), triple HET (+/−) and triple KO (−/−) mice. (D) In situ zymography assay demonstrates decreased proteolytic activity, indicated by reduction in green fluorescence, in triple KO epidermis. As a control, addition of the serine protease inhibitor aprotinin reduces fluorescent signal in both WT and triple KO (bottom panels). Note nuclear retention in cornified layers of the triple KO (white arrows). Bar: 50 μm.
Mentions: Our analysis demonstrated that in the absence of involucrin, envoplakin, and periplakin, the epidermal barrier was abnormal, and suggested that the compensatory mechanism allowing barrier function to be maintained was reduced desquamation. To investigate how this was achieved, microarray analysis was performed using mRNA isolated from dorsal skin of 7-wk-old WT, triple HET, or triple KO female mice (Fig. 5). Skin histology proximal to that used for RNA isolation indicated hyperkeratosis in the triple KO, but not the WT or triple HET (unpublished data). Consistent with their phenotypes, the triple HET samples clustered more closely with those of WT than with triple KO (Fig. 5 A). 207 genes were found to have statistically significant (P < 0.05) changes (twofold or greater; increase or decrease) in expression between WT and triple KO samples (Table S1, available at http://www.jcb.org/cgi/content/full/jcb.200706187/DC1).

Bottom Line: Cornified envelopes form but are ultrastructurally abnormal, with reduced lipid content and decreased mechanical integrity.Expression of proteases is reduced and the protease inhibitor, serpina1b, is highly upregulated, resulting in defective filaggrin processing and delayed degradation of desmoglein 1 and corneodesmosin.There is infiltration of CD4+ T cells and a reduction in resident gammadelta+ T cells, reminiscent of atopic dermatitis.

View Article: PubMed Central - PubMed

Affiliation: Cancer Research UK, Cambridge Research Institute, Li Ka Shing Centre, Cambridge CB2 0RE, England, UK.

ABSTRACT
The cornified envelope is assembled from transglutaminase cross-linked proteins and lipids in the outermost epidermal layers and is essential for skin barrier function. Involucrin, envoplakin, and periplakin form the protein scaffold on which the envelope assembles. To examine their combined function, we generated mice deficient in all three genes. The triple knockouts have delayed embryonic barrier formation and postnatal hyperkeratosis (abnormal accumulation of cornified cells) resulting from impaired desquamation. Cornified envelopes form but are ultrastructurally abnormal, with reduced lipid content and decreased mechanical integrity. Expression of proteases is reduced and the protease inhibitor, serpina1b, is highly upregulated, resulting in defective filaggrin processing and delayed degradation of desmoglein 1 and corneodesmosin. There is infiltration of CD4+ T cells and a reduction in resident gammadelta+ T cells, reminiscent of atopic dermatitis. Thus, combined loss of the cornified envelope proteins not only impairs the epidermal barrier, but also changes the composition of T cell subpopulations in the skin.

Show MeSH
Related in: MedlinePlus