PMC

(a) Laboratory pipeline for simultaneous depletion of rRNA from prokaryotic and eukaryotic RNA mixtures. The enriched mRNA is used to create RNA-Seq libraries. (b) Bioinformatics pipeline for sequential mapping and analysis of simultaneous RNA-Seq data.

(a) Chlamydia 1 hpi versus 24 hpi. Chlamydial genes above the cutoff of RPKM≥0.1 and a minimum of 10 mapped reads are highlighted in blue and red at 1 and 24 hpi respectively; and host cells at (b) 1hpi relative to mock and (c) 24 hpi relative to mock. Significantly differentially expressed host cell transcripts (FDR≤0.05 and LFC≥2.0) between the mock and infected conditions are plotted in red (up-regulated) and blue (down-regulated). Pearson's correlation (R²) between replicates is indicated for each.

(a) Gremlin1 and (b) tenascin-C in Chlamydia-infected cells at 1 and 24 hpi, compared to mock-infected cells. Values are based on fold changes calculated from absolute quantitation of each gene of interest, normalized to human ATP synthase 6. Asterisks indicate statistically significant differences as calculated by Student's t test (***: p<0.0001; **: p<0.002). Error bars represent standard deviation over a minimum of 2 biological replicates.

Infection of epithelial cells by Chlamydia leads to production of proinflammatory cytokines and chemokines that lead to recruitment and activation of immune cells. Recruited immune cells and infected epithelial cells secrete pro-fibrotic matrix metalloproteases (MMPs) that act upon the extracellular matrix (ECM), including collagens. The breakdown products of these proteases are also pro-inflammatory. Infected epithelial cells express the pro-fibrotic molecules TGF-β, Gremlin1 and Tenascin-C; expression of each amplifies the other, creating a series of nested positive feedback loops that increase the deposition of collagens and other ECM components, which in turn further induce immune cell recruitment and activation.