Expression of COX-2 and Nrf2/GPx3 in the anterior vaginal wall tissues of women with pelvic organ prolapse.

PURPOSE: To evaluate COX-2 and Nrf2/GPx3 expressions in the lamina propria of the anterior vaginal wall tissues of women with and without pelvic organ prolapse (POP). METHODS: Tissue samples of anterior vaginal wall were examined using HE staining, immuohistochemical staining and Western blot for the expressions of COX-2/PGE2, Nrf2/GPx3, MMP2, TIMP1, collagen I and collagen III (n = 35, per group). RESULTS: Compared with control group, collagen fibers of the anterior vaginal wall were disorganized and discontinuous. Expressions of Nrf2, GPx3, TIMP1, collagen I and collagen III were found significantly lower in POP group (P < 0.05); while, expressions of COX-2, PGE2, and MMP2 were found significantly higher in POP group (P < 0.05). Statistically significant correlations of COX-2 and Nrf2/GPx3 were showed (P < 0.01). CONCLUSION: We found that the interaction between inflammation and oxidative stress was closely related to the development of POP. This study demonstrates that COX-2 and Nrf2 pathways may be involved in pathogenesis of POP, as promising potential therapeutic targets and agents.

Expression of Heparanase in uterosacral ligaments of women with or without uterine prolapse.

OBJECTIVE: Pelvic organ prolapse (POP) is a global health problem for which the pathophysiological mechanism remains unclear. The loss of extracellular matrix proteins is considered an important molecular basis for this pathology. Heparanase is a heparin sulfate degrading endoglycosidase that has an important role in various biological processes and is a key component of extracellular matrix. The aim of this study was to compare expression of Heparanase in connective tissue of uterosacral ligaments in women with or without uterine prolapse. STUDY DESIGN: Thirty-nine women who underwent hysterectomy for benign reasons were enrolled in the study. Twenty-three women with uterine prolapse (stage ≥3) who underwent vaginal hysterectomy (VH) - POP group, were compared to sixteen women without uterine prolapse who underwent abdominal hysterectomy (stage <2) - control group. Uterosacral ligaments (USL) biopsies were obtained from all uterine specimens near their origin. All tissue samples were analyzed by immunohistochemistry and tested for the presence of Heparanase using antiheparanse antibody 733. RESULTS: Heparanse positive staining was more common in the connective tissue of uterosacral ligaments in women with uterine prolapse. Positive staining was seen in 17/23 (73.9 %) women with uterine prolapse compared to 4/16 (25 %) without uterine prolapse (p = 0.003). On multivariate logistic regression analysis, positive staining displayed a trend for an independent association with POP, after controlling for menopausal status and parity (OR 13.57, 95 %CI 0.82-224.4, p = 0.06). CONCLUSION: Heparanase expression is more common in the connective tissue of uterosacral ligaments in women with uterine prolapse compared to women with no prolapse.

Vaginal Expression of LOXL1 in Premenopausal and Postmenopausal Women With Pelvic Organ Prolapse.

OBJECTIVES: This study aimed to compare cellular expression of lysyl oxidase-like 1 (LOXL1), a key enzyme in elastin metabolism, of premenopausal women with pelvic organ prolapse (POP) compared with premenopausal controls without POP and postmenopausal women with POP. In addition, we examined whether variation of LOXL1 expression was dependent on biopsy site. METHODS: A standardized protocol was utilized to obtain vaginal biopsies from 30 women (10 premenopausal POP, 10 postmenopausal POP, and 10 premenopausal non-POP). Expression levels of messenger RNA (mRNA) and protein of LOXL1 were determined using real-time quantitative polymerase chain reactions and enzyme-linked immunosorbant assays. Analysis was performed to determine if there were differences between group or biopsy site. RESULTS: Significant differences in LOXL1 mRNA expression were found between patient groups (P = 0.0033). LOXL1 mRNA expression (relative to 18S) was upregulated in the postmenopausal POP group (54.5 ± 14.7) compared with the premenopausal POP group (5.2 ± 14.7, P = 0.0034) and the premenopausal non-POP group (23 ± 18, P = 0.0359). No significant differences in LOXL1 protein expression (nanogram/milliliter per microgram total protein) were seen between groups (premenopausal POP, 3.2 × 10 ± 6.3 × 10; postmenopausal POP, 4.3 × 10 ± 6.3 × 10; premenopausal non-POP, 5.0 × 10 ± 7.7 × 10; P = 0.15). No differences in mRNA expression were seen between sites (P = 0.74), but significant variation was noted in protein expression (P = 0.001). CONCLUSIONS: Premenopausal and postmenopausal women with POP exhibit differential expression of LOXL1 suggesting different pathways in the pathogenesis of POP. The role of biopsy location on LOXL1 expression requires further investigation.

Dysregulation of protease and protease inhibitors in a mouse model of human pelvic organ prolapse.

Mice deficient for the fibulin-5 gene (Fbln5(-/-)) develop pelvic organ prolapse (POP) due to compromised elastic fibers and upregulation of matrix metalloprotease (MMP)-9. Here, we used casein zymography, inhibitor profiling, affinity pull-down, and mass spectrometry to discover additional protease upregulated in the vaginal wall of Fbln5(-/-) mice, herein named V1 (25 kDa). V1 was a serine protease with trypsin-like activity similar to protease, serine (PRSS) 3, a major extrapancreatic trypsinogen, was optimum at pH 8.0, and predominantly detected in estrogenized vaginal epithelium of Fbln5(-/-) mice. PRSS3 was (a) localized in epithelial secretions, (b) detected in media of vaginal organ culture from both Fbln5(-/-) and wild type mice, and (c) cleaved fibulin-5 in vitro. Expression of two serine protease inhibitors [Serpina1a (α1-antitrypsin) and Elafin] was dysregulated in Fbln5(-/-) epithelium. Finally, we confirmed that PRSS3 was expressed in human vaginal epithelium and that SERPINA1 and Elafin were downregulated in vaginal tissues from women with POP. These data collectively suggest that the balance between proteases and their inhibitors contributes to support of the pelvic organs in humans and mice.

Increased matrix metalloproteinases-1,-9 in the uterosacral ligaments and vaginal tissue from women with pelvic organ prolapse.

OBJECTIVES: To investigate the possible association of increased matrix metalloproteinases (MMPs)-1,-9 with pelvic organ prolapse (POP) and to evaluate whether inflammatory processes contribute to its development. STUDY DESIGN: Forty women who underwent hysterectomy, 20 with POP grade 2 and above, and 20 without POP, participated in the study. Biopsies from the uterosacral ligaments and vaginal mucosa were obtained from each woman. Each biopsy was sectioned and stained for MMP-1 and MMP-9 by immunohistochemical methods and with hematoxylin and eosin (H&E). MMP-1,-9 expressions were evaluated on the immunostained slides. H&E stained sections were examined for possible inflammatory changes. RESULTS: A higher stromal (extra-cellular) expression of MMPs-1,-9 was found in POP cases compared with controls in vaginal biopsies (MMP-1: p=0.004; MMP-9: p=0.042) as well as in uterosacral ligament biopsies (MMP-1: p=0.011; MMP-9: p=0.015). Increased intracellular expression of both MMPs was also demonstrated in fibroblasts in biopsies of women with POP (p<0.001 for all). Most of these differences persisted after controlling for age. The degree of inflammatory changes reflected by the number of lymphocytes, plasma cells and capillary-sized blood vessels per 10 high power fields, was similar in specimens obtained from women with and without POP. CONCLUSIONS: The expression of MMPs-1,-9 appears to be increased in tissues from women with POP. This supports an association, although not a causal relation, between increased MMPs-1,-9 and POP. Inflammation does not seem to play an important role in the pathogenesis of POP.

LOX family enzymes expression in vaginal tissue of premenopausal women with severe pelvic organ prolapse.

INTRODUCTION AND HYPOTHESIS: The extracellular matrix proteins collagen and elastin provide tissue strength and resilience, whereas lysyl oxidase enzymes play a major role in their stabilization. This study examines the expression and tissue localization of lysyl oxidase family proteins in the anterior vaginal wall of premenopausal women with advanced pelvic organ prolapse (POP, n = 15) and asymptomatic controls (n = 11). All women were in the proliferative phase of menstrual cycle. METHODS: Total mRNAs and proteins extracted from the vaginal tissue were examined by real-time polymerase chain reaction and immunoblotting, and tissue specimens were analyzed by immunohistochemistry. RESULTS: The expression of LOX, LOXL1, and LOXL3 genes as well as LOX and LOXL3 proteins were significantly reduced in POP patients (P < 0.05). Immunolocalization of LOX family proteins was confirmed in all vaginal specimens. CONCLUSION: We proposed that reduced expression of LOX enzymes may result in defective assembly of pelvic tissues and development of POP.

Expression of matrix metalloproteinase-1 in uterosacral ligaments tissue of women with genital prolapse.

Collagen metabolism is altered in the pelvic organ tissues of women with genital prolapse. The aim of this study was to compare collagen metabolism by measuring matrix metalloproteinase-1 (MMP-1) expression in uterosacral ligament tissues of postmenopausal women with and without genital prolapse. Uterosacral ligament tissues were obtained at the time of abdominal or vaginal surgery from twenty-four patients with pelvic organ prolapse (POP) and 21 women who underwent gynecologic surgery for benign indications. The tissue samples were analyzed by immunohistochemistry. There were no differences in age, BMI and parity between two groups. The patients with genital prolapse demonstrated significantly higher occurences of MMP-1 expression compared to controls. These findings indicate that increased MMP-1 expression in uterosacral ligaments is associated with genital prolapse. Our data are consistent with the theory that increased collagen breakdown may play an important role in the onset and development of pelvic organ prolapse (POP).

Is lysyl oxidase-like protein-1, alpha-1 antitrypsin, and neutrophil elastase site specific in pelvic organ prolapse?

INTRODUCTION AND HYPOTHESIS: We investigated whether the expression of alpha-1 antitrypsin (ATT), neutrophil elastase (NE), and lysyl oxidase-like protein 1 (LOXL-1) vary within the vagina in subjects with pelvic organ prolapse (POP). METHODS: Biopsies were obtained from the anterior and posterior vaginal wall of 22 women with POP (> or =stage 2 by POP-Q). The subjects were grouped by the most prominent defect: cystocele, cystocele plus uterine prolapse, and rectocele. Comparative real-time PCR, Western blotting, and NE enzyme activity assay were performed. RESULTS: The ratio of anterior and posterior vaginal wall ATT, NE, and LOXL-1 expression varied between individuals within the same defect group. CONCLUSIONS: ATT, NE, and LOXl-1 expression was variable among different biopsy sites in the vagina. No consistent pattern was present when the subjects were grouped by the most prominent defect. We recommend careful consideration of biopsy sites in future studies on POP to enhance reproducibility of data.