Ryo Uemura1, Daisuke Tachibana2, Masayuki Shiota3, Kayo Yoshida4, Kohei Kitada1, Akihiro Hamuro1, Takuya Misugi1, Masayasu Koyama1. 1. Department of Obstetrics and Gynecology, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno-ku, Osaka, 545-8585, Japan. 2. Department of Obstetrics and Gynecology, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno-ku, Osaka, 545-8585, Japan. dtachibana@med.osaka-cu.ac.jp. 3. Research Support Platform, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno-ku, Osaka, 545-8585, Japan. 4. Department of Laboratory Animal Science, Osaka City University Graduate School of Medicine, 1-4-3 Asahimachi, Abeno-ku, Osaka, 545-8585, Japan.
Abstract
INTRODUCTION AND HYPOTHESIS: Pelvic organ prolapse (POP) in women is associated with deficiency of elastic fibers, and fibulin-5 is known to be a critical protein in the synthesis of elastin. The purpose of this study is to investigate the related pathway for the synthesis of elastin via fibulin-5 using fibulin-5 knockout mice. METHODS: Fibulin-5 knockout mice were generated using the CRISPR/Cas9 system, and vaginal dilatation was used to mimic vaginal delivery. We divided the mice into three groups: Fbln5+/+ mice immediately after dilatation (Fbln5+/+ day0), Fbln5+/+ mice 3 days after dilatation (Fbln5+/+ day3) and Fbln5-/- mice 3 days after dilatation (Fbln5-/- day3). Proteins related to elastogenesis in the vaginal wall were measured by liquid chromatography mass spectrometry (LC-MS/MS) analysis, and differences in the expression of these proteins between the Fbln5-/- mice and the Fbln5+/+ mice were analyzed using western blotting. RESULTS: In the LC-MS/MS analysis, protein tyrosine kinase 7 (PTK7) was not detected in the Fbln5-/- day3 group, although the expression increased by > 1.5 times between the Fbln5+/+ day0 and day3 groups. PTK7 and β-catenin are known to act in the Wnt/β-catenin pathway, and both were upregulated after dilatation in the Fbln5+/+ mice, though not in the Fbln5-/- mice. CONCLUSION: Our findings suggest that these proteins are involved in elastogenesis via fibulin-5, and the impairment of these proteins might be the underlying cause of POP manifestation.
INTRODUCTION AND HYPOTHESIS: Pelvic organ prolapse (POP) in women is associated with deficiency of elastic fibers, and fibulin-5 is known to be a critical protein in the synthesis of elastin. The purpose of this study is to investigate the related pathway for the synthesis of elastin via fibulin-5 using fibulin-5 knockout mice. METHODS: Fibulin-5 knockout mice were generated using the CRISPR/Cas9 system, and vaginal dilatation was used to mimic vaginal delivery. We divided the mice into three groups: Fbln5+/+ mice immediately after dilatation (Fbln5+/+ day0), Fbln5+/+ mice 3 days after dilatation (Fbln5+/+ day3) and Fbln5-/- mice 3 days after dilatation (Fbln5-/- day3). Proteins related to elastogenesis in the vaginal wall were measured by liquid chromatography mass spectrometry (LC-MS/MS) analysis, and differences in the expression of these proteins between the Fbln5-/- mice and the Fbln5+/+ mice were analyzed using western blotting. RESULTS: In the LC-MS/MS analysis, protein tyrosine kinase 7 (PTK7) was not detected in the Fbln5-/- day3 group, although the expression increased by > 1.5 times between the Fbln5+/+ day0 and day3 groups. PTK7 and β-catenin are known to act in the Wnt/β-catenin pathway, and both were upregulated after dilatation in the Fbln5+/+ mice, though not in the Fbln5-/- mice. CONCLUSION: Our findings suggest that these proteins are involved in elastogenesis via fibulin-5, and the impairment of these proteins might be the underlying cause of POP manifestation.
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