In-vitro Augmentation of Mesenchymal Stem Cells by Using Adult Bovine Serum


Cite item

Full Text

Abstract

Background:Umbilical cord mesenchymal stem cells (UC-MSCs) are increasingly being utilized for immune-related disease therapies due to their low immunogenicity. However, the primary culture of UC-MSCs requires the supplementation of serum in the growth medium, which has posed a challenge due to ethical issues related to the collection method of the fetal bovine serum (FBS) that is routinely used in cell culture.

Aim:In order to address this, the purpose of this research was to assess the effectiveness of adult bovine serum (ABS) as a different and more affordable source of serum for the in-vitro cultivation of UC-MSCs. UC-MSCs were isolated from the umbilical cord of Wharton’s jelly of cow immediately after birth, by digestion with Collagenase type I.

Method:ABS was collected from fresh bovine sources and heat-inactivated. The morphology of UC-MSCs was observed under an inverted microscope, and growth patterns, proliferative index, and doubling time were calculated every two days to compare the efficacy of ABS with FBS. Immunocytochemistry for specific markers was also conducted on the MSCs.

Result:The results showed a notable difference in morphology, growth rate, population doubling, and proliferative index between ABS and FBS.

Conclusion:Intriguingly, ABS proved to be an effective supplement in the growth medium for expanding UC-MSCs in vitro, providing a viable alternative to FBS.

About the authors

Somia Shehzadi

University Institute of Medical Laboratory Technology, The University of Lahore

Author for correspondence.
Email: info@benthamscience.net

Maryam Javed

Institute of Biochemistry and Biotechnology,, University of Veterinary and Animal Sciences

Email: info@benthamscience.net

Asmat Ullah

Clinical Research Institute,, Zhejiang Provincial People’s Hospital

Author for correspondence.
Email: info@benthamscience.net

Ahmad Bilal Waqar

University Institute of Medical Laboratory Technology, The University of Lahore, 54000, Pakistan, The University of Lahore

Email: info@benthamscience.net

Fatima Iftikhar Shah

University Institute of Medical Laboratory Technology, The University of Lahore

Email: info@benthamscience.net

Sajjad Ullah

University Institute of Medical Laboratory Technology,, The University of Lahore

Email: info@benthamscience.net

References

  1. Pittenger MF, Mackay AM, Beck SC, et al. Multilineage potential of adult human mesenchymal stem cells. Science 1999; 284(5411): 143-7. doi: 10.1126/science.284.5411.143 PMID: 10102814
  2. Dominici M, Le Blanc K, Mueller I, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy 2006; 8(4): 315-7. doi: 10.1080/14653240600855905 PMID: 16923606
  3. Prockop DJ. Marrow stromal cells as stem cells for nonhematopoietic tissues. Science 1997; 276(5309): 71-4. doi: 10.1126/science.276.5309.71 PMID: 9082988
  4. Le Blanc K, Rasmusson I, Sundberg B, et al. Treatment of severe acute graft-versus-host disease with third party haploidentical mesenchymal stem cells. Lancet 2004; 363(9419): 1439-41. doi: 10.1016/S0140-6736(04)16104-7 PMID: 15121408
  5. Lazarus HM, Koc ON, Devine SM, et al. Cotransplantation of HLA-identical sibling culture-expanded mesenchymal stem cells and hematopoietic stem cells in hematologic malignancy patients. Biol Blood Marrow Transplant 2005; 11(5): 389-98. doi: 10.1016/j.bbmt.2005.02.001 PMID: 15846293
  6. Consentius C, Reinke P, Volk HD. Immunogenicity of allogeneic mesenchymal stromal cells: What has been seen in vitro and in vivo? Regen Med 2015; 10(3): 305-15. doi: 10.2217/rme.15.14 PMID: 25933239
  7. Hachisuga T, Fukuda K, Hayashi Y, Iwasaka T, Sugimori H. Immunohistochemical demonstration of histiocytes in normal ectocervical epithelium and epithelial lesions of the uterine cervix. Gynecol Oncol 1989; 33(3): 273-8. doi: 10.1016/0090-8258(89)90511-8 PMID: 2656425
  8. Chase LG, Yang S, Zachar V, et al. Development and characterization of a clinically compliant xeno-free culture medium in good manufacturing practice for human multipotent mesenchymal stem cells. Stem Cells Transl Med 2012; 1(10): 750-8. doi: 10.5966/sctm.2012-0072 PMID: 23197667
  9. Chase LG, Lakshmipathy U, Solchaga LA, Rao MS, Vemuri MC. A novel serum-free medium for the expansion of human mesenchymal stem cells. Stem Cell Res Ther 2010; 1(1): 8. doi: 10.1186/scrt8 PMID: 20504289
  10. Bakopoulou A, Apatzidou D, Aggelidou E, et al. Isolation and prolonged expansion of oral mesenchymal stem cells under clinical- grade, GMP-compliant conditions differentially affects "stemness" properties. Stem Cell Res Ther 2017; 8(1): 247. doi: 10.1186/s13287-017-0705-0 PMID: 29096714
  11. Gottipamula S, Muttigi MS, Chaansa S, et al. Large-scale expansion of pre-isolated bone marrow mesenchymal stromal cells in serum-free conditions. J Tissue Eng Regen Med 2016; 10(2): 108-19. doi: 10.1002/term.1713 PMID: 23495227
  12. Kim J, Kim M, Nahm SS, Lee DM, Pokharel S, Choi I. Characterization of gender-specific bovine serum. Anim Cells Syst 2011; 15(2): 147-54. doi: 10.1080/19768354.2011.577584
  13. Witzeneder K, Lindenmair A, Gabriel C, et al. Human-derived alternatives to fetal bovine serum in cell culture. Transfus Med Hemother 2013; 40(6): 417-23. doi: 10.1159/000356236 PMID: 24474892
  14. Barberini DJ, Freitas NPP, Magnoni MS, et al. Equine mesenchymal stem cells from bone marrow, adipose tissue and umbilical cord: Immunophenotypic characterization and differentiation potential. Stem Cell Res Ther 2014; 5(1): 25. doi: 10.1186/scrt414 PMID: 24559797
  15. Majore I, Moretti P, Hass R, Kasper C. Identification of subpopulations in mesenchymal stem cell-like cultures from human umbilical cord. Cell Commun Signal 2009; 7(1): 6. doi: 10.1186/1478-811X-7-6 PMID: 19302702
  16. Thaweesapphithak S, Tantrawatpan C, Kheolamai P, Tantikanlayaporn D, Roytrakul S, Manochantr S. Human serum enhances the proliferative capacity and immunomodulatory property of MSCs derived from human placenta and umbilical cord. Stem Cell Res Ther 2019; 10(1): 79. doi: 10.1186/s13287-019-1175-3 PMID: 30845980
  17. Maqbool T, Awan SJ, Malik S, Hadi F, Shehzadi S, Tariq K. In-Vitro Anti-Proliferative, Apoptotic and Antioxidative Activities of Medicinal Herb Kalonji (Nigella sativa). Curr Pharm Biotechnol 2019; 20(15): 1288-308. doi: 10.2174/1389201020666190821144633 PMID: 31433749
  18. Shahdadfar A, Frønsdal K, Haug T, Reinholt FP, Brinchmann JE. In vitro expansion of human mesenchymal stem cells: Choice of serum is a determinant of cell proliferation, differentiation, gene expression, and transcriptome stability. Stem Cells 2005; 23(9): 1357-66. doi: 10.1634/stemcells.2005-0094 PMID: 16081661
  19. Bieback K, Hecker A, Kocaömer A, et al. Human alternatives to fetal bovine serum for the expansion of mesenchymal stromal cells from bone marrow. Stem Cells 2009; 27(9): 2331-41. doi: 10.1002/stem.139 PMID: 19544413
  20. Lindroos B, Boucher S, Chase L, et al. Serum-free, xeno-free culture media maintain the proliferation rate and multipotentiality of adipose stem cells in vitro. Cytotherapy 2009; 11(7): 958-72. doi: 10.3109/14653240903233081 PMID: 19903107
  21. Patrikoski M, Juntunen M, Boucher S, et al. Development of fully defined xeno-free culture system for the preparation and propagation of cell therapy-compliant human adipose stem cells. Stem Cell Res Ther 2013; 4(2): 27. doi: 10.1186/scrt175 PMID: 23497764
  22. Gstraunthaler G, Lindl T, van der Valk J. A plea to reduce or replace fetal bovine serum in cell culture media. Cytotechnology 2013; 65(5): 791-3. doi: 10.1007/s10616-013-9633-8 PMID: 23975256
  23. Gstraunthaler G. Alternatives to the use of fetal bovine serum: Serum-free cell culture. Altern Anim Exp 2003; 20(4): 257-81. doi: 10.14573/altex.2003.4.257 PMID: 14671707
  24. van der Valk J, Mellor D, Brands R, et al. The humane collection of fetal bovine serum and possibilities for serum-free cell and tissue culture. Toxicol In Vitro 2004; 18(1): 1-12. doi: 10.1016/j.tiv.2003.08.009 PMID: 14630056
  25. Zamansky GB, Arundel C, Nagasawa H, Little JB. Adaptation of human diploid fibroblasts in vitro to serum from different sources. J Cell Sci 1983; 61(1): 289-97. doi: 10.1242/jcs.61.1.289 PMID: 6885940
  26. Ziegler A, Everett H, Hamza E, et al. Equine dendritic cells generated with horse serum have enhanced functionality in comparison to dendritic cells generated with fetal bovine serum. BMC Vet Res 2016; 12(1): 254. doi: 10.1186/s12917-016-0880-8 PMID: 27846835
  27. Shanskii YD, Sergeeva NS, Sviridova IK, et al. Human platelet lysate as a promising growth-stimulating additive for culturing of stem cells and other cell types. Bull Exp Biol Med 2013; 156(1): 146-51. doi: 10.1007/s10517-013-2298-7 PMID: 24319712
  28. Franke J, Abs V, Zizzadoro C, Abraham G. Comparative study of the effects of fetal bovine serum versus horse serum on growth and differentiation of primary equine bronchial fibroblasts. BMC Vet Res 2014; 10(1): 119. doi: 10.1186/1746-6148-10-119 PMID: 24886635
  29. Fernyhough ME, Hausman GJ, Dodson MV. Progeny from dedifferentiated bovine adipocytes display protracted adipogenesis. Cells Tissues Organs 2008; 188(4): 359-72. doi: 10.1159/000134007 PMID: 18493114
  30. Wu H, Ren Y, Li S, et al. In vitro culture and induced differentiation of sheep skeletal muscle satellite cells. Cell Biol Int 2012; 36(6): 579-87. doi: 10.1042/CBI20110487 PMID: 22233500
  31. Lee DM, Bajracharya P, Lee EJ, et al. Effects of gender-specific adult bovine serum on myogenic satellite cell proliferation, differentiation and lipid accumulation. In Vitro Cell Dev Biol Anim 2011; 47(7): 438-44. doi: 10.1007/s11626-011-9427-2 PMID: 21614650
  32. Yu JE, Kim M, Pokharel S, et al. Potential use of adult bovine serum obtained during the slaughtering process as a biological reagent. Anim Cells Syst 2013; 17(2): 106-12. doi: 10.1080/19768354.2013.772073
  33. Kleeberger C, Shore D, Gunter E, Sandler DP, Weinberg CR. The Effects of Long-term Storage on Commonly Measured Serum Analyte Levels. Epidemiology 2018; 29(3): 448-52. doi: 10.1097/EDE.0000000000000810 PMID: 29384792
  34. Ulfman LH, Leusen JHW, Savelkoul HFJ, Warner JO, van Neerven RJJ. Effects of Bovine Immunoglobulins on Immune Function, Allergy, and Infection. Front Nutr 2018; 5: 52. doi: 10.3389/fnut.2018.00052 PMID: 29988421

Supplementary files

Supplementary Files
Action
1. JATS XML
Download

Copyright (c) 2024 Bentham Science Publishers