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Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids

Background and Objectives: Centipeda minima (L.) is a well-known and traditional pharmaceutical that has been utilized to treat different conditions controlling rhinitis, soothe pain, and decrease swelling. We assessed the impacts of Centipeda minima (L.) extricates (CMTs) on the osteogenic differen...

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Autores principales: Lee, Hyun-Jin, Na, Kyung-Hwan, Uddin, Md. Salah, Park, Jun-Beom
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9863519/
https://www.ncbi.nlm.nih.gov/pubmed/36676667
http://dx.doi.org/10.3390/medicina59010043
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author Lee, Hyun-Jin
Na, Kyung-Hwan
Uddin, Md. Salah
Park, Jun-Beom
author_facet Lee, Hyun-Jin
Na, Kyung-Hwan
Uddin, Md. Salah
Park, Jun-Beom
author_sort Lee, Hyun-Jin
collection PubMed
description Background and Objectives: Centipeda minima (L.) is a well-known and traditional pharmaceutical that has been utilized to treat different conditions controlling rhinitis, soothe pain, and decrease swelling. We assessed the impacts of Centipeda minima (L.) extricates (CMTs) on the osteogenic differentiation of cell spheroids made of human-bone-marrow-derived mesenchymal stem cells. Materials and Methods: Mesenchymal stem cells (MSCs) in spheroid 3D culture were generated and propagated in the presence of CMTs ranging from 0 to 1 μg/mL. Cell morphology was measured on Days 1, 3, 5, and 7. The quantitative cellular viability was evaluated on Days 1, 3, 5, and 7. Alkaline phosphatase activity assays were designed to measure the osteogenic differentiation of mesenchymal stem cell spheroids on Day 7. Alizarin Red S staining was performed to investigate the mineralization of cell spheroids on Days 7 and 14. Real-time polymerase chain reactions were used to measure the expression levels of RUNX2 and COL1A1 on Day 14. Western blot techniques were performed to identify the protein expression of Runt-related transcription factor 2 and type I collagen. Results: The control group’s mesenchymal stem cells displayed a spheroid shape. There was no noticeable change in morphology with the addition of CMTs at final concentrations of 0.001, 0.01, 0.1, and 1 μg/mL compared with the untreated (control) group. The application of CMTs did not induce a significant change in cell viability. The relative alkaline phosphatase activity values in the 0.001, 0.01, 0.1, and 1 μg/mL CMT groups were 114.4% ± 8.2%, 130.6% ± 25.3%, 87.8% ± 3.4%, and 92.1% ± 6.8%, respectively, considering a control of 100% (100.0% ± 17.9%). On Day 14, calcium deposits were clearly observed in each group. The relative values of Alizarin Red S staining in the 0.001, 0.01, 0.1, and 1 μg/mL CMT groups were 100.1% ± 8.9%, 105.9% ± 0.0%, 109.7% ± 19.1%, and 87.0% ± 40.9%, respectively, considering a control of 100% (100.0% ± 28.7%). The addition of CMT significantly increased RUNX2 expression in the 0.01 μg/mL group and COL1A1 in the 0.001 and 0.01 μg/mL groups. Normalization of protein expression showed that the addition of CMTs significantly increased type I collagen expression in the 0.001, 0.01, and 1 μg/mL groups. Conclusions: In conclusion, CMTs influence the osteogenic differentiation of bone-marrow-derived mesenchymal stem cells and the use of CMTs may positively influence the osteogenic differentiation of cell spheroids.
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spelling pubmed-98635192023-01-22 Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids Lee, Hyun-Jin Na, Kyung-Hwan Uddin, Md. Salah Park, Jun-Beom Medicina (Kaunas) Article Background and Objectives: Centipeda minima (L.) is a well-known and traditional pharmaceutical that has been utilized to treat different conditions controlling rhinitis, soothe pain, and decrease swelling. We assessed the impacts of Centipeda minima (L.) extricates (CMTs) on the osteogenic differentiation of cell spheroids made of human-bone-marrow-derived mesenchymal stem cells. Materials and Methods: Mesenchymal stem cells (MSCs) in spheroid 3D culture were generated and propagated in the presence of CMTs ranging from 0 to 1 μg/mL. Cell morphology was measured on Days 1, 3, 5, and 7. The quantitative cellular viability was evaluated on Days 1, 3, 5, and 7. Alkaline phosphatase activity assays were designed to measure the osteogenic differentiation of mesenchymal stem cell spheroids on Day 7. Alizarin Red S staining was performed to investigate the mineralization of cell spheroids on Days 7 and 14. Real-time polymerase chain reactions were used to measure the expression levels of RUNX2 and COL1A1 on Day 14. Western blot techniques were performed to identify the protein expression of Runt-related transcription factor 2 and type I collagen. Results: The control group’s mesenchymal stem cells displayed a spheroid shape. There was no noticeable change in morphology with the addition of CMTs at final concentrations of 0.001, 0.01, 0.1, and 1 μg/mL compared with the untreated (control) group. The application of CMTs did not induce a significant change in cell viability. The relative alkaline phosphatase activity values in the 0.001, 0.01, 0.1, and 1 μg/mL CMT groups were 114.4% ± 8.2%, 130.6% ± 25.3%, 87.8% ± 3.4%, and 92.1% ± 6.8%, respectively, considering a control of 100% (100.0% ± 17.9%). On Day 14, calcium deposits were clearly observed in each group. The relative values of Alizarin Red S staining in the 0.001, 0.01, 0.1, and 1 μg/mL CMT groups were 100.1% ± 8.9%, 105.9% ± 0.0%, 109.7% ± 19.1%, and 87.0% ± 40.9%, respectively, considering a control of 100% (100.0% ± 28.7%). The addition of CMT significantly increased RUNX2 expression in the 0.01 μg/mL group and COL1A1 in the 0.001 and 0.01 μg/mL groups. Normalization of protein expression showed that the addition of CMTs significantly increased type I collagen expression in the 0.001, 0.01, and 1 μg/mL groups. Conclusions: In conclusion, CMTs influence the osteogenic differentiation of bone-marrow-derived mesenchymal stem cells and the use of CMTs may positively influence the osteogenic differentiation of cell spheroids. MDPI 2022-12-26 /pmc/articles/PMC9863519/ /pubmed/36676667 http://dx.doi.org/10.3390/medicina59010043 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Lee, Hyun-Jin
Na, Kyung-Hwan
Uddin, Md. Salah
Park, Jun-Beom
Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title_full Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title_fullStr Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title_full_unstemmed Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title_short Assessment of the Impacts of Centipeda minima (L.) on Cell Viability, and Osteogenic Differentiation of Mesenchymal Stem Cell Spheroids
title_sort assessment of the impacts of centipeda minima (l.) on cell viability, and osteogenic differentiation of mesenchymal stem cell spheroids
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9863519/
https://www.ncbi.nlm.nih.gov/pubmed/36676667
http://dx.doi.org/10.3390/medicina59010043
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