Morphology of LSMCs changes on different collagen matrices.

<p><b>Equal numbers of</b> LSMCs were cultured on plastic (A), monomeric (B) or fibrillar (C) collagen-coated dishes in serum containing medium for 48 hours and then imaged. Three hours after seeding, the fibrillar collagen coating of a dish was partially removed to expose LSMCs to the plastic matrix (D) (Bars 200 µm in main figures; 50 µm in inserts).</p

Activation of MAPK signaling pathway is influenced by the interaction of LSMCs with ECM.

<p>(A & B) LSMCs were cultured on plastic, monomeric or fibrillar collagen-coated dishes and then treated with 10 ng/ml PDGF for various time points. Lysates were blotted for (A) pERK1/2 and ERK1/2, and (B) pPDGFR and PDGFR. Densitometry values were normalized to total protein expression for each phosphorylated protein. (C) LSMCs cultured on plastic dishes were pre-treated with 10 µM of AG1296 or 25 µM of PD98059 in serum-free medium for two hours followed by treatment with 10 ng/ml PDGF for 10 minutes. Lysates were blotted for pERK1/2 and ERK1/2 Statistical significance within each matrix and compared to non-treated sample is indicated by asterisks. Letters indicate statistical differences across all samples on different matrices (n =3, p<0.05).</p

Growth, but not attachment of LSMCs is different on collagen matrices.

<p>(A) LSMCs were cultured on plastic, monomeric, or fibrillar collagen-coated dishes in serum-containing medium for 6 days. At the end of each day cells were trypsinized and counted using a hemacytometer. Rate of growth on each matrix was calculated using linear regression. (B) LSMCs cultured on different matrices were gently washed three hours after seeding and imaged to count the number of attached cells to each matrix. Statistical significance between matrices is indicated by asterisk (n=3, p<0.05).</p

Basal and PDGF-stimulated proliferation of LSMCs are different on collagen matrices.

<p>Cell-cycle synchronized LSMCs were cultured on 96-well plates coated with different forms of collagen in medium with or without PDGF(10 ng/ml) for 24 hours. At the end of the treatment, cell proliferation was measured with thymidine incorporation assays. Statistical significance of the cellular growth rate between matrices within each treatment group as well as between stimulated and non-stimulated cells are indicated by asterisks (n=3, p<0.05).</p

Effect of ECM collagen on cell cycle progression in LSMCs.

<p>Leiomyoma cells were treated with serum free (SF) medium or PDGF (10ng/ml) for 24 hours on either plastic or monomeric collagen matrices and then assayed by FACs analysis for cell cycle phase. 10,000 events were counted per cell line and the percent of cells in each phase is expressed as total number of diploid cells. Statistical analysis by orthogonal contrast statements was performed between treatments in the S phase of the cellular cycle to address changes in cellular proliferation. Asterisks denote statistical significance of the indicated comparisons (n=6; p<0.05).</p

Activation of focal adhesion complexes and F-actin stress fibers in LSCMs is altered on different collagen matrices.

<p>LSMCs were cultured on different matrices for 24 hour and then fixed and stained with DAPI and specific antibodies against FAK pY397, α2 integrin, vinculin, and F-actin for imaging (Bar 50µm).</p

Effect of ECM collagen on p21 expression in LSMCs.

<p>Leiomyoma cells were treated with serum free (SF) or PDGF (10ng/ml) for 24 hours on either plastic, monomeric or fibrillar collagen matrices. Cell lysates were subjected to SDS-PAGE and immunoblotted for p21and GAPDH expression. Densitometry values were normalized to GAPDH expression and compared amongst all treatments and matrices. Statistical analysis by orthogonal contrast statements was performed comparing each treatment within each matrix and also comparing each treatment across all matrices. Statistical significance for all comparisons is indicated by asterisks (n=4; p<0.05).</p

PCoA plots of distances between pig urogenital bacterial communities.

<p>Variances explained are shown on the axes. Plots include all vaginal swab and cervical flush samples taken of each pig throughout the course of the study. (A) Comparison of lean and obese animals throughout the whole study. (B) Comparison of all samples from both obese and lean animals during the induction versus the maintenance phase of the diet. Each point represents one sample. The color key provides information to determine the influence of features on spatial placement.</p

Alpha-diversity on number of OTUs observed.

<p>Results are expressed as least squared mean ± SEM (control, <i>n</i> = 3; obese, <i>n</i> = 2).</p

Taxon summary of relative abundance within urogenital microbial communities of control and obese pigs shown as an average of all animals from each treatment group.

<p>Taxa with the highest abundance is presented in the key. (A) Phylum-level relative abundance. (B) Genus-level relative abundance.</p