Osteogenic Differentiation  in vitro  off Human Osteoblasts is Associated with Only Slight Shift in Their Proteomics Profile

I. A. Khvorova; Хворова И. А.; A. B. Malashicheva; Малашичева А. Б.; V. V. Karelkin; Карелкин В. В.; A. P. Sereda; Середа А. П.; S. A. Bozhkova; Божкова С. А.; R. M. Tikhilov; Тихилов Р. М.; E. S. Gromova; Громова Е. С.; E. A. Fefilova; Фефилова Е. А.; B. R. Zainullina; Зайнуллина Б. Р.; D. A. Kostina; Костина Д. А.; A. A. Lobov; Лобов А. А.

doi:10.31857/S0041377123010066

Osteogenic Differentiation in vitro off Human Osteoblasts is Associated with Only Slight Shift in Their Proteomics Profile

Authors: Khvorova I.A.¹, Malashicheva A.B.¹, Karelkin V.V.², Sereda A.P.², Bozhkova S.A.², Tikhilov R.M.², Gromova E.S.¹, Fefilova E.A.¹, Zainullina B.R.³, Kostina D.A.¹, Lobov A.A.¹
Affiliations:
1. Laboratory of Regenerative Biology, Institute of Cytology of the Russian Academy of Sciences
2. Vreden National Medical Research Center of Traumatology and Orthopedics
3. Resource Centre for Molecular and Cell Technologies, St. Petersburg State University
Issue: Vol 65, No 1 (2023)
Pages: 20-27
Section: Articles
URL: https://journals.rcsi.science/0041-3771/article/view/140064
DOI: https://doi.org/10.31857/S0041377123010066
EDN: https://elibrary.ru/GPCARC
ID: 140064

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Abstract
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Abstract

Fracture healing is a complex process in which the periosteum and endosteum become the main sources of osteoblast progenitor cells. However, cellular mechanisms and signaling cascades underlying the early stages of osteoblast progenitors differentiation in adult bone are still not well understood. Therefore, we performed shotgun proteomics analysis of primary culture of isolated human osteoblasts from femur of adult donors in undifferentiated conditions and on the sixth day of osteogenic differentiation in vitro. This is an early timepoint in which we have observed no extracellular matrix mineralization yet. 1612 proteins identified with at least two unique peptides were included in proteomics analysis. Data are available via ProteomeXchange with identifier PXD033697. Despite the fact, that matrix mineralization starts only after induction of osteogenic differentiation, we revealed unexpectedly weak physiological shift associated with a decrease of cells proliferative activity and changes in proteins inVved in extracellular matrix secretion and organization. We demonstrated that osteoblasts were positive for markers of later osteogenic differentiation stages during standard cultivation: osteopontin, osteocalcin, BMP-2/4 and RUNX2. Therefore, further differentiation required for matrix mineralization needs minimal physiological changes.

Keywords

osteoblasts, osteogenic differentiation, shotgun proteomics, bone, mass spectrometry

References

Bahney C., Zondervan R., Allison P., Theologis A., Ashley J., Ahn J., Miclau T., Marcucio R., Hankenson K. 2019. Cellular biology of fracture healing. J. Orthop. Res. V. 37. P. 35. https://doi.org/10.1002/jor.24170
Blighe K., Sharmila R., Myles L. 2022. EnhancedVcano: publication-ready Vcano plots with enhanced colouring and labeling. https://bioconductor.org/packages/devel/bioc/vignettes/EnhancedVcano/inst/doc/EnhancedVcano.html
Bragdon B., Bahney C. 2018. Origin of reparative stem cells in fracture healing, Curr. Osteoporos. Rep. V. 16. P. 490. https://doi.org/10.1007/s11914-018-0458-4
Cleland T., Vashishth D. 2015. Bone protein extraction without demineralization utilizing principles from hydroxyapatite chromatography. Anal. Biochem. V. 472. P. 62. https://doi.org/10.1016/j.ab.2014.12.006
Cleland T., Voegele K., Schweitzer M. 2012. Empirical evaluation of bone extraction protocols. PLoS One. V. 7. P. e31443. https://doi.org/10.1371/journal.pone.0031443
Florencio-Silva R., Sasso G., Sasso-Cerri E., Simões M., Cerri P. 2015. Biology of bone tissue: structure, function, and factors that influence bone cells. Biomed. Res. Int. V. 2015. P. e421746. https://doi.org/10.1155/2015/421746
Hastie T., Tibshirani R., Narasimhan B., Chu G. 2022. Impute: imputation for microarray data. Bioconductor version: Release (3.14). https://www.bioconductor.org/packages/release/bioc/html/impute.html
Jiang X., Ye M., Jiang X., Liu G., Feng S., Cui L., Zou H. 2007. Method development of efficient protein extraction in bone tissue for proteome analysis. J. Proteome Res. V. 6. P. 2287. https://doi.org/10.1021/pr070056t
Lobov A., Malashicheva A. 2022. Osteogenic differentiation: a universal cell program of heterogeneous mesenchymal cells or a similar extracellular matrix mineralizing phenotype? Bio. Comm. V. 67. P. 32. https://doi.org/10.21638/spbu03.2022.104
Matthews B., Novak S., Sbrana F., Funnell J., Cao Y., Buckels E., Grcevic D., Kalajzic I. 2021. Heterogeneity of murine periosteum progenitors inVved in fracture healing. Elife. V. 10. P. e58534. https://doi.org/10.7554/eLife.58534
Perez-Riverol Y., Bai J.; Bandla C., García-Seisdedos D., Hewapathirana S., Kamatchinathan S., Kundu D.J., Prakash A., Frericks-Zipper A., Eisenacher M., Walzer M., Wang S., Brazma A., Vizcaíno J.A. 2022. The PRIDE database resources in 2022: A hub for mass spectrometry-based proteomics evidences. Nucleic Acids Res. V. 50. D543–D552. https://doi.org/10.1093/nar/gkab1038
Pitkänen S. 2020. In vitro and in vivo osteogenesis and vasculogenesis in synthetic bone grafts. Doctoral dissertation: Tampere University.
Raouf A., Ganss B., McMahon C., Vary C., Roughley P., Seth A. 2002. Lumican is a major proteoglycan component of the bone matrix. Matrix Biol. V. 21. P 361. https://doi.org/10.1016/s0945-053x(02)00027-6
Ritchie M.E., Phipson B., Wu D., Hu Y., Law C.W., Shi W., Smyth G.K. 2015. Limma powers differential expression analyses for RNA-sequencing and microarray studies. Nucleic Acids Res. V. 43. P. e47. https://doi.org/10.1093/nar/gkv007
Rohart F., Gautier B., Singh A., Cao K. 2017. mixOmics: an R package for ‘omics’ feature selection and multiple data integration. PLoS Comput. Biol. V. 13. P. e1005752. https://doi.org/10.1371/journal.pcbi.1005752
Rutkovskiy A., Stensløkken K., Vaage I. 2016. Osteoblast differentiation at a glance. Med. Sci. Monit. Basic Res. V. 22. P. 95. https://doi.org/10.12659/MSMBR.901142
Wickham H. 2016. ggplot2. Cham: Springer Int. Publishing.
Yan L. 2021. ggvenn: Draw Venn Diagram by “ggplot2”. https://cran.r-project.org/web/packages/ggvenn/