{"title":"Botox-A Induced Apoptosis and Suppressed Cell Proliferation in Fibroblasts Pre-Treated with Breast Cancer Exosomes.","authors":"Hossein Sayaf, Niloufar Salimian, Mahnaz Mohammadi, Parisa Ahmadi, Amir Gholamzad, Sadegh Babashah, Maliheh Entezari, Najma Farahani, Maryam Montazeri, Mehrdad Hashemi","doi":"10.1016/j.mcp.2024.102007","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>breast cancer-associated fibroblast (CAF) is linked to metastasis and is poor for breast cancer prognosis. Since Clostridium Toxin A (Botox-A) had represented a cytotoxic effect on fibroblasts, this study aims to assess Botox-A cytotoxicity in both normal fibroblasts and exosome-induced CAFs.</p><p><strong>Material and method: </strong>the serum exosomes of 40 BC patients and 30 healthy individuals were isolated and lncRNA H19 (lnch19) levels were assessed by qRT-PCR method. After that, Breast Cancer (BC) exosomes co-cultured with Human foreskin fibroblasts (HFF) and qRT-PCR were applied to evaluate α-SMA, Vimentin, BCL-2, and BAX expression. Both Normal and malignant HFFs co-cultured with Botox-A, and Botox-A loaded exosome for 24 and 48 hours and their apoptosis, Cell proliferation, and viability were monitored by MTT assay, Annexin V-FITC and PI staining and qRT-PCR for BCL-2, BAX, and cyclin D1 mRNAs.</p><p><strong>Results: </strong>Serum exosomes of BC patients had significantly higher levels of lncRNA H19 than healthy individuals. MTT assay results showed Botox-A decreased vital Human foreskin fibroblasts in a dose-dependent manner. BC exosomes significantly increased α-SMA, Vimentin, and BCL-2 mRNA levels in Human foreskin fibroblasts, on the other hand, BAX decreased meaningfully. Co-culture of exosome-treated HFF cells with both Botox-A and Botox-A loaded exosomes significantly boosted BCL-2 mRNA levels, completely contrary to BAX and cyclid d1 expression. Meanwhile, flow cytometry results confirmed a high rate of apoptosis in malignant Human foreskin fibroblasts treated with Botox-A loaded exosome.</p><p><strong>Conclusion: </strong>The findings of this study indicate that exosomal lncRNA H19 could be a diagnostic marker for Breast Cancer and these Breast cancer exosomes can induce malignant phenotype in fibroblasts and turn them into CAFs. Botox-A could be toxic for both normal fibroblasts and CAFs, inducing apoptosis and suppressing cell proliferation among them.</p>","PeriodicalId":49799,"journal":{"name":"Molecular and Cellular Probes","volume":" ","pages":"102007"},"PeriodicalIF":2.3000,"publicationDate":"2024-12-26","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"","citationCount":"0","resultStr":null,"platform":"Semanticscholar","paperid":null,"PeriodicalName":"Molecular and Cellular Probes","FirstCategoryId":"99","ListUrlMain":"https://doi.org/10.1016/j.mcp.2024.102007","RegionNum":3,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"","PubModel":"","JCR":"Q3","JCRName":"BIOCHEMICAL RESEARCH METHODS","Score":null,"Total":0}
引用次数: 0
Abstract
Background: breast cancer-associated fibroblast (CAF) is linked to metastasis and is poor for breast cancer prognosis. Since Clostridium Toxin A (Botox-A) had represented a cytotoxic effect on fibroblasts, this study aims to assess Botox-A cytotoxicity in both normal fibroblasts and exosome-induced CAFs.
Material and method: the serum exosomes of 40 BC patients and 30 healthy individuals were isolated and lncRNA H19 (lnch19) levels were assessed by qRT-PCR method. After that, Breast Cancer (BC) exosomes co-cultured with Human foreskin fibroblasts (HFF) and qRT-PCR were applied to evaluate α-SMA, Vimentin, BCL-2, and BAX expression. Both Normal and malignant HFFs co-cultured with Botox-A, and Botox-A loaded exosome for 24 and 48 hours and their apoptosis, Cell proliferation, and viability were monitored by MTT assay, Annexin V-FITC and PI staining and qRT-PCR for BCL-2, BAX, and cyclin D1 mRNAs.
Results: Serum exosomes of BC patients had significantly higher levels of lncRNA H19 than healthy individuals. MTT assay results showed Botox-A decreased vital Human foreskin fibroblasts in a dose-dependent manner. BC exosomes significantly increased α-SMA, Vimentin, and BCL-2 mRNA levels in Human foreskin fibroblasts, on the other hand, BAX decreased meaningfully. Co-culture of exosome-treated HFF cells with both Botox-A and Botox-A loaded exosomes significantly boosted BCL-2 mRNA levels, completely contrary to BAX and cyclid d1 expression. Meanwhile, flow cytometry results confirmed a high rate of apoptosis in malignant Human foreskin fibroblasts treated with Botox-A loaded exosome.
Conclusion: The findings of this study indicate that exosomal lncRNA H19 could be a diagnostic marker for Breast Cancer and these Breast cancer exosomes can induce malignant phenotype in fibroblasts and turn them into CAFs. Botox-A could be toxic for both normal fibroblasts and CAFs, inducing apoptosis and suppressing cell proliferation among them.
期刊介绍：
MCP - Advancing biology through–omics and bioinformatic technologies wants to capture outcomes from the current revolution in molecular technologies and sciences. The journal has broadened its scope and embraces any high quality research papers, reviews and opinions in areas including, but not limited to, molecular biology, cell biology, biochemistry, immunology, physiology, epidemiology, ecology, virology, microbiology, parasitology, genetics, evolutionary biology, genomics (including metagenomics), bioinformatics, proteomics, metabolomics, glycomics, and lipidomics. Submissions with a technology-driven focus on understanding normal biological or disease processes as well as conceptual advances and paradigm shifts are particularly encouraged. The Editors welcome fundamental or applied research areas; pre-submission enquiries about advanced draft manuscripts are welcomed. Top quality research and manuscripts will be fast-tracked.
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