Fig 2: Effect of the PAR-2 inhibitor, GB83, on PG-LPS-induced MMP expression, IRAK1/TAK1 activation, and proinflammatory cytokine expression in HGFs. (a) Cells were pretreated with GB83 for 1 h and then stimulated with PG-LPS for 24 and 48 h. MMP1 mRNA levels were analyzed via the quantitative real-time polymerase chain reaction (qRT-PCR) using GAPDH as an internal control. (b) Cells were pretreated with GB83 for 1 h and then stimulated with PG-LPS for 0, 2, 4, and 6 h. Total cellular mRNA was extracted, and the mRNA levels of IL6, IL8, TNFα, and IL1β were determined via the qRT-PCR. (c) Cells were pretreated with GB83 for 1 h and then stimulated with PG-LPS for 5 min. Cell lysates were subjected to western blotting to determine the levels of total and phosphorylated IRAK1 and TAK1 proteins. Values are presented as means ± standard error of mean of three independent experiments. ∗p < 0.05 vs. untreated control; ∗∗p < 0.01 vs. PG-LPS. MMP: matrix metalloproteinase; PG-LPS, Porphyromonas gingivalis-derived lipopolysaccharide; HGFs: human gingival fibroblasts; IRAK1: interleukin-1 receptor-associated kinase 1; TAK1: transforming growth factor β-activated kinase 1; IL6: interleukin6; IL8: interleukin8; TNFα: tumor necrosis factor α; IL1β: interleukin 1β; GAPDH: glyceraldehyde 3-phosphate dehydrogenase.

Fig 3: Effects of hsa-miR-100-5p transfection on the downstream target molecule expression in NESCs. (a) hsa-miR-100-5p expression after precursor miRNA transfection. Note that the vertical axis is expressed as a logarithmic scale. (b) SMARCD1 mRNA expression. (c) MMP1 mRNA expression. (d) Active MMP1 protein expression. *p < 0.05, **p < 0.005, #p < 0.0005 vs. controls (Student’s t-test). MMP1, matrix metallopeptidase 1; NESCs, normal endometrial stromal cells; SMARCD1, SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1

Fig 4: Effects of SMARCD1 siRNA transfection on the MMP1 mRNA expression in NESCs. (a) SMARCD1 mRNA expression after SMARCD1 siRNA transfection. (b) MMP1 mRNA expression. *p < 0.05, **p < 0.005 vs. controls (Student’s t-test). MMP1, matrix metallopeptidase 1; NESCs, normal endometrial stromal cells; SMARCD1, SWItch/sucrose non-fermentable-related matrix-associated actin-dependent regulator of chromatin subfamily D member 1

Fig 5: Validation of and NMLSMR targets identified by the high throughput assay. A, To test for reproducibility of the assay, 293T cells were transfected with the MMP‐1/pGL3 reporter plasmid. Cells were seeded in three 96‐well plates using an interleaved format and treated with 5% CSE media (high‐signal) 2% CSE (medium‐signal) no CSE (low‐signal). After a 24‐h incubation, the luciferase activity was measured in each well. Representative data from days 1–3 is shown. The calculated Z' factor was 0.71 ± 0.05 SEM, n = 5. B, As expected, the ERK inhibitor PD98059 inhibited luciferase expression in a dose‐dependent fashion in positive controls exposed to 5% CSE (*** p ≤ 0.001, significantly different from control; ### p ≤ 0.001, significantly different from promoter + CSE; ## p≤0.01, significantly different from promoter + CSE). C, Data from the high throughout screen displaying each compound tested as a block dot. The y‐axis illustrates the change in MMP1 expression compared with expression levels in CSE exposed in HEK293 cells. The drugs in the −100% range (red shaded area) are capable of nullifying all CSE induced MMP1 transcriptional activity