Analysis of hair follicle- and keratinocyte differentiation-associated transcripts in ILK-deficient epidermis.

<p>(<b>A</b>) Paraffin sections of skin isolated from 3 day-old mice of the indicated genotype were processed for histological analysis, and stained with hematoxylin and eosin. Bar, 40 µm (<b>B, C</b>) Expression levels of selected transcripts encoding proteins present in the hair follicle (panel B) or in differentiated keratinocytes (panel C) in ILK-deficient <i>K14Cre-Ilk^f/f</i> epidermis, determined by qPCR. The results are expressed as the mean+SEM (n = 5), and asterisks indicate <i>P</i><0.001 (Student's t test) relative to the corresponding transcript levels in ILK-expressing <i>K14Cre-Ilk^f/+</i> epidermis (set to 1). (<b>D</b>) Protein lysates prepared from the skin of 3 day-old mice of the indicated genotypes were resolved by denaturing gel electrophoresis, followed by immunoblot analysis using antibodies against anti-S100A3. The blots were also probed with antibodies against GAPDH, used as loading control.</p

Targeted inactivation of <i>Ilk</i> and loss of ILK transcripts and protein in mouse epidermis.

<p>(<b>A</b>) The skin of 3 day-old <i>K14Cre-Ilk^f/+</i> and <i>K14Cre-Ilk^f/f</i> mice (4 each, from two different litters) was isolated and treated with dispase to separate the epidermis from the dermis. Epidermal lysates were prepared and analyzed by immunoblot with antibodies against ILK or γ-tubulin, used as loading control. (<b>B</b>) Total RNA from <i>K14Cre-Ilk^f/+</i> and <i>K14Cre-Ilk^f/f</i> mice (5 each) was used to interrogate GeneChip Mouse Gene 1.0 ST Arrays, and the signal intensity corresponding to individual <i>Ilk</i> exons was analyzed. The average intensity values obtained for each exon in RNA from <i>K14Cre-Ilk^f/f</i> epidermis are shown relative to those in <i>K14Cre-Ilk^f/+</i> tissues, which have been set to 1.0.</p

Validation of the differential expression of selected genes in ILK-deficient epidermis.

<p>Validation of the differential expression of selected genes in ILK-deficient epidermis.</p

Altered expression of genes encoding growth factors and developmental regulators in ILK-deficient epidermis.

<p>(<b>A, B</b>) Expression levels of selected transcripts encoding proteins associated with growth factors or pigmentation (panel A), or key factors in developmental epidermal pathways (panel B) in ILK-deficient <i>K14Cre-Ilk^f/f</i> epidermis, determined by qPCR. The results are expressed as the mean+SEM (n = 5). The differences in expression in ILK-deficient epidermis for each of transcripts shown reached statistical significance (<i>P</i><0.001, Student's t test) relative to the corresponding transcript levels in ILK-expressing <i>K14Cre-Ilk^f/+</i> epidermis (set to 1). (<b>C</b>) Protein lysates prepared from the skin of 3 day-old <i>K14Cre-Ilk^f/f</i> or <i>K14Cre-Ilk^f/+</i> mice (two animals of each genotype) were resolved by denaturing gel electrophoresis, followed by immunoblot analysis using antibodies against LGR5, or γ-tubulin, used as loading control. Lysates prepared from wild type mouse brain were used as a positive control for LGR5 expression.</p