HOXA 11 and 13 expression in women with pelvic floor disorder

Abstract

Pelvic floor dysfunction (PFD), including stress urinary incontinence (SUI) and pelvic organ prolapse (POP), is a major health problem for elderly women. Although PFD is a highly prevalent disease, the underlying mechanism and basic pathophysiology are poorly understood. Uterosacral ligaments (USLs) are the key structures of the uterus and vagina and are fragile in patients with PFD. Endopelvic fascias in the vagina compose the supportive tissue of the pelvic floor and often are diminished in PFD.Homeobox (HOX) genes are transcriptional regulators that orchestrate embryonic development. The HOXA11 gene controls the development of the lower uterine cervix and vagina. The HOXA13 gene is also responsible for the development of the vagina and regulates extracellular matrix constituents. We hypothesized that expressions of HOXA11 in USLs and HOXA13 in the vagina might be decreased in women with PFD. In addition, we investigated quantitative mRNA expression of matrix metalloproteinases (MMPs), collagen І, and collagen Ш, from PFD and non-PFD patients.Sixty-eight subjects were enrolled for the experimental procedure. Biopsy specimens were obtained from the anterior and posterior apex of the vagina and bilateral USLs from women with no PFD, SUI combined with POP, and POP; from patients with SUI, we took only anterior vaginal specimens by biopsies. Specimens were divided in half. One section was prepared for real-time polymerase chain reaction (RT-PCR), and the other half was saved for western blot analysis. RT-PCR and western blots were used to determine HOXA11 and HOXA13 expression and the protein levels. In addition, we performed RT-PCR to compare the expression of MMP2, MMP9, collagen І, and collagen Ш genes in the vaginal wall and USLs of all enrolled patients. Quantitative data are expressed as mean + SEM and as percentages. The results of relative mRNA expression analysis were compared using the Kruskal–Wallis test for independent ‘k’ samples, complemented when necessary by Dunnett’s multiple comparisons test, to evaluate possible differences between patient groups, for both vaginal wall and USL samples. Statistical analyses were performed with SPSS 19 (SPSS Inc. Chicago, IL, USA) and Graph-Pad Prism 5.0 (GraphPad Software Inc., San Diego, CA, USA). A P value less than 0.05 was considered statistically significant for all data analyzed.We found that the relative HOXA13 mRNA expression in the anterior vaginal wall of SUI patients was 4.3-fold lower compared with that of controls (P = 0.006). In addition, the anterior vaginal wall specimens from patients in the SUI combined POP and POP groups had lower expression of HOXA13 compared with the controls (4.3-fold, P = 0.007; 6.3-fold, P < 0.001, respectively). In the posterior vaginal wall, the relative mRNA expression of HOXA13 of SUI combined POP and POP patients was lower (3.1-fold, P < 0.001; 2.7-fold, P < 0.001, respectively) than that of control patients. There were no differences in vaginal expression of HOXA13 between SUI and POP specimens or between specimens obtained from patients with SUI combined POP and patients with only POP. Expression of HOXA11 was 2.3-fold lower in women with POP compared with controls (P = 0.004). HOXA 11 expression in USLs was also 2.7-fold lower in SUI combined POP patients compared with controls (P = 0.004); however, there was no statistically significant difference between POP patients and women with SUI combined POP (P = 0.08). Western blot analysis demonstrated the presence of HOXA11 and 13 in the USLs and vagina, although the levels were lower in the POP and/or SUI group compared with controls (P < 0.001). Collagen І and collagen Ш expressions were lower in groups of PFD patients than controls. The expression of MMP 2 mRNA in USLs was elevated in POP patients with/without SUI compared to controls, although this alteration was not statistically significant for women with POP. In addition, the change in expression detected for both MMP2 and MMP9 was not statistically significant in samples from the anterior and posterior vaginal wall.Expression of HOXA13 and both collagens in the vagina was reduced in women with SUI, SUI combined POP, and POP compared with control women. Expression of HOXA11 and both collagens in USLs was also reduced in all PFD patients. These reduced levels may contribute to alterations in the biomechanical strength of the pelvic supportive tissue, leading to prolapse or incontinence. A better understanding of the influence of these genes may prove beneficial in defining the underlying etiologies of PFD development and aid in the development of new treatment options