Figure 4.

Neonatal maternal separation (NMS) and vaginal balloon distension (VBD) significantly disrupted gene and protein expression within the vagina. RT-PCR (A) and Western blot (B–C) was performed on vagina from naïve and NMS21 mice that either did not (Naïve-ND and NMS21-ND) or did undergo VBD (Naïve-VBD and NMS21-VBD). A) NMS and VBD had a significant interaction effect on IL10 mRNA expression with NMS21-VBD expressing significantly more IL10 mRNA than NMS21-ND or naïve-VBD. NMS and VBD also had a significant interaction effect on TNFα mRNA expression with NMS21-VBD expressing significantly more TNFα mRNA than naïve-VBD. VBD significantly increased Artemin (Art) mRNA expression with NMS21-VBD expression significantly more Art than NMS21-ND. IL6 and nerve growth factor (NGF) transcripts were not affected by either NMS or VBD. B) Representative Western blots are shown for CRF₁, CRF₂, and corresponding GAPDH protein expression with bands at 55, 47, and 35kD, respectively. NMS and VBD had a significant interaction effect on CRF₁ protein expression in the vagina with NMS21-VBD expressing significantly less CRF₁ protein than naïve-VBD. CRF₂ protein expression in the vagina was unaffected by NMS or VBD. C) Representative Western blots are shown for TRPV1, TRPA1, and corresponding GAPDH protein expression with bands at 85, 127, and 35kD, respectively. VBD significantly decreased the protein expression of both TRPV1 and TRPA1 in the vagina. Naïve-VBD and NMS21-VBD both expressed significantly less TRPV1 than their non-distended counterparts. NMS21-VBD also expressed significantly less TRPA1 protein than NMS21-ND. Housekeeping gene: GAPDH (A). Brackets indicate significant impact of VBD (^#,####p < 0.05, 0.0001) or an NMS/VBD interaction (^+,++p < 0.05, 0.01), two-way ANOVA; *,**p < 0.05, 0.01 vs. naïve, ^#,### p < 0.05, 0.001 vs. baseline, Bonferroni posttest.