Shreesha K Bhat, Manjunath B Joshi, Sampara Vasishta, Rajesh N Jagadale, Setlur G Biligiri, Monika A Coronado, Raghuvir K Arni, Kapaettu Satyamoorthy
{"title":"两栖类的 P-I 金属蛋白酶和 L- 氨基酸氧化酶可抑制血管生成。","authors":"Shreesha K Bhat, Manjunath B Joshi, Sampara Vasishta, Rajesh N Jagadale, Setlur G Biligiri, Monika A Coronado, Raghuvir K Arni, Kapaettu Satyamoorthy","doi":"10.1590/1678-9199-JVATITD-2020-0180","DOIUrl":null,"url":null,"abstract":"<p><strong>Background: </strong>Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, <i>inter alia</i>. In the present study, we have examined the effects of P-I metalloproteinases isolated from <i>Bothrops moojeni</i> (BmMP-1) and <i>Bothrops atrox</i> (BaMP-1) and L-amino acid oxidases (LAAO) isolated from <i>B. moojeni</i> (BmLAAO) and <i>B. atrox</i> (BaLAAO) on biochemical and functional aspects of angiogenesis.</p><p><strong>Methods: </strong>P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by <i>in vitro</i> 3D spheroid assay, Matrigel tube forming assay, and <i>in vivo</i> agarose plug transformation in mice.</p><p><strong>Results: </strong>P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. <i>In vitro,</i> P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery.</p><p><strong>Conclusion: </strong>Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties <i>in vitro</i> and <i>in vivo</i>.</p>","PeriodicalId":1,"journal":{"name":"Accounts of Chemical Research","volume":" ","pages":"e20200180"},"PeriodicalIF":16.4000,"publicationDate":"2021-08-18","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8381740/pdf/","citationCount":"0","resultStr":"{\"title\":\"P-I metalloproteinases and L-amino acid oxidases from <i>Bothrops</i> species inhibit angiogenesis.\",\"authors\":\"Shreesha K Bhat, Manjunath B Joshi, Sampara Vasishta, Rajesh N Jagadale, Setlur G Biligiri, Monika A Coronado, Raghuvir K Arni, Kapaettu Satyamoorthy\",\"doi\":\"10.1590/1678-9199-JVATITD-2020-0180\",\"DOIUrl\":null,\"url\":null,\"abstract\":\"<p><strong>Background: </strong>Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, <i>inter alia</i>. In the present study, we have examined the effects of P-I metalloproteinases isolated from <i>Bothrops moojeni</i> (BmMP-1) and <i>Bothrops atrox</i> (BaMP-1) and L-amino acid oxidases (LAAO) isolated from <i>B. moojeni</i> (BmLAAO) and <i>B. atrox</i> (BaLAAO) on biochemical and functional aspects of angiogenesis.</p><p><strong>Methods: </strong>P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by <i>in vitro</i> 3D spheroid assay, Matrigel tube forming assay, and <i>in vivo</i> agarose plug transformation in mice.</p><p><strong>Results: </strong>P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. <i>In vitro,</i> P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery.</p><p><strong>Conclusion: </strong>Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties <i>in vitro</i> and <i>in vivo</i>.</p>\",\"PeriodicalId\":1,\"journal\":{\"name\":\"Accounts of Chemical Research\",\"volume\":\" \",\"pages\":\"e20200180\"},\"PeriodicalIF\":16.4000,\"publicationDate\":\"2021-08-18\",\"publicationTypes\":\"Journal Article\",\"fieldsOfStudy\":null,\"isOpenAccess\":false,\"openAccessPdf\":\"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC8381740/pdf/\",\"citationCount\":\"0\",\"resultStr\":null,\"platform\":\"Semanticscholar\",\"paperid\":null,\"PeriodicalName\":\"Accounts of Chemical Research\",\"FirstCategoryId\":\"3\",\"ListUrlMain\":\"https://doi.org/10.1590/1678-9199-JVATITD-2020-0180\",\"RegionNum\":1,\"RegionCategory\":\"化学\",\"ArticlePicture\":[],\"TitleCN\":null,\"AbstractTextCN\":null,\"PMCID\":null,\"EPubDate\":\"2021/1/1 0:00:00\",\"PubModel\":\"eCollection\",\"JCR\":\"Q1\",\"JCRName\":\"CHEMISTRY, MULTIDISCIPLINARY\",\"Score\":null,\"Total\":0}","platform":"Semanticscholar","paperid":null,"PeriodicalName":"Accounts of Chemical Research","FirstCategoryId":"3","ListUrlMain":"https://doi.org/10.1590/1678-9199-JVATITD-2020-0180","RegionNum":1,"RegionCategory":"化学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":null,"EPubDate":"2021/1/1 0:00:00","PubModel":"eCollection","JCR":"Q1","JCRName":"CHEMISTRY, MULTIDISCIPLINARY","Score":null,"Total":0}
P-I metalloproteinases and L-amino acid oxidases from Bothrops species inhibit angiogenesis.
Background: Snake venoms are composed of pharmacologically active proteins that are evolutionarily diverse, stable and specific to targets. Hence, venoms have been explored as a source of bioactive molecules in treating numerous diseases. Recent evidences suggest that snake venom proteins may affect the formation of new blood vessels. Excessive angiogenesis has been implicated in several pathologies including tumours, diabetic retinopathy, arthritis, inter alia. In the present study, we have examined the effects of P-I metalloproteinases isolated from Bothrops moojeni (BmMP-1) and Bothrops atrox (BaMP-1) and L-amino acid oxidases (LAAO) isolated from B. moojeni (BmLAAO) and B. atrox (BaLAAO) on biochemical and functional aspects of angiogenesis.
Methods: P-I metalloproteinases and LAAO were purified from venom by molecular size exclusion and ion-exchange chromatography and subsequently confirmed using mass spectrometry. The P-I metalloproteinases were characterized by azocaseinolytic, fibrinogenolytic and gelatinase activity and LAAO activity was assessed by enzyme activity on L-amino acids. Influence of these proteins on apoptosis and cell cycle in endothelial cells was analysed by flow cytometry. The angiogenic activity was determined by in vitro 3D spheroid assay, Matrigel tube forming assay, and in vivo agarose plug transformation in mice.
Results: P-I metalloproteinases exhibited azocaseinolytic activity, cleaved α and partially β chain of fibrinogen, and displayed catalytic activity on gelatin. LAAO showed differential activity on L-amino acids. Flow cytometry analysis indicated that both P-I metalloproteinases and LAAO arrested the cells in G0/G1 phase and further induced both necrosis and apoptosis in endothelial cells. In vitro, P-I metalloproteinases and LAAO exhibited significant anti-angiogenic properties in 3D spheroid and Matrigel models by reducing sprout outgrowth and tube formation. Using agarose plug transplants in mice harbouring P-I metalloproteinases and LAAO we demonstrated a marked disruption of vasculature at the periphery.
Conclusion: Our research suggests that P-I metalloproteinases and LAAO exhibit anti-angiogenic properties in vitro and in vivo.
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Accounts of Chemical Research presents short, concise and critical articles offering easy-to-read overviews of basic research and applications in all areas of chemistry and biochemistry. These short reviews focus on research from the author’s own laboratory and are designed to teach the reader about a research project. In addition, Accounts of Chemical Research publishes commentaries that give an informed opinion on a current research problem. Special Issues online are devoted to a single topic of unusual activity and significance.
Accounts of Chemical Research replaces the traditional article abstract with an article "Conspectus." These entries synopsize the research affording the reader a closer look at the content and significance of an article. Through this provision of a more detailed description of the article contents, the Conspectus enhances the article's discoverability by search engines and the exposure for the research.
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