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Although postbiotics have demonstrated potential in alleviating constipation, their specific mechanism of action and bioactive components remain unclear. This study aimed to investigate the ameliorative effects and potential mechanisms of postbiotics derived from Bifidobacterium animalis subsp. lactis BL-99 (BL-99) on FC using both in vivo and in vitro models. The findings revealed that both BL-99 and its postbiotics significantly mitigated FC symptoms, as evidenced by enhanced intestinal motility, and elevated fecal water content. Additionally, treatment with BL-99 postbiotics was associated with an increase in the thickness of the intestinal muscular layer and a reduction in apoptosis of intestinal smooth muscle cells (SMCs). Mechanistically, BL-99 postbiotics were found to enhance the contractile response and promote the proliferation of intestinal SMCs. Furthermore, untargeted metabolomics analysis identified two key bioactive peptides, Glu-Val and Glu-Leu, as the active components in BL-99 responsible for regulating SMC function. Collectively, these findings highlight the potential of BL-99 postbiotics as a promising functional food ingredient for alleviating FC, providing a novel and effective strategy for the developing dietary interventions targeting this condition.<\/jats:p>","DOI":"10.3390\/foods14193441","type":"journal-article","created":{"date-parts":[[2025,10,8]],"date-time":"2025-10-08T12:04:52Z","timestamp":1759925092000},"page":"3441","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":7,"title":["Modulation of Intestinal Smooth Muscle Cell Function by BL-99 Postbiotics in Functional Constipation"],"prefix":"10.3390","volume":"14","author":[{"ORCID":"https:\/\/orcid.org\/0000-0002-0094-711X","authenticated-orcid":false,"given":"Wen","family":"Zhao","sequence":"first","affiliation":[{"name":"Key Laboratory of Functional Dairy, Department of Nutrition and Health, China Agricultural University, Beijing 100190, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Mingkun","family":"Liu","sequence":"additional","affiliation":[{"name":"School of Food and Biological Engineering, Hefei University of Technology, Hefei 230009, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Hanglian","family":"Lan","sequence":"additional","affiliation":[{"name":"National Center of Technology Innovation for Dairy, Hohhot 100118, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ran","family":"Wang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Functional Dairy, Department of Nutrition and Health, China Agricultural University, Beijing 100190, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Wei-Lian","family":"Hung","sequence":"additional","affiliation":[{"name":"National Center of Technology Innovation for Dairy, Hohhot 100118, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jian","family":"He","sequence":"additional","affiliation":[{"name":"National Center of Technology Innovation for Dairy, Hohhot 100118, China"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-4818-9079","authenticated-orcid":false,"given":"Bing","family":"Fang","sequence":"additional","affiliation":[{"name":"Key Laboratory of Functional Dairy, Department of Nutrition and Health, China Agricultural University, Beijing 100190, China"}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"1968","published-online":{"date-parts":[[2025,10,8]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"86","DOI":"10.5694\/mja18.00241","article-title":"Chronic idiopathic constipation in adults: Epidemiology, pathophysiology, diagnosis and clinical management","volume":"209","author":"Black","year":"2018","journal-title":"Med. 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