Human umbilical cord blood plasma as an alternative to animal sera for mesenchymal stromal cells in vitro expansion - A multicomponent metabolomic analysis

PLoS One. 2018 Oct 10;13(10):e0203936. doi: 10.1371/journal.pone.0203936. eCollection 2018.

Abstract

Mesenchymal Stromal cells (MSCs) have a potential role in cell-based therapies. Foetal bovine serum (FBS) is used to supplement the basal cell culture medium but presents several disadvantages and risks. Other alternatives have been studied, including human umbilical cord blood plasma (hUCBP), aiming at the development of xeno-free culturing protocols. A comparative characterization of multicomponent metabolic composition of hUCBP and commercial FBS based on Nuclear Magnetic Resonance (NMR) spectroscopy and multivariate statistical analysis was performed. The analysis of 1H-NMR spectra revealed both similarities and differences between the two proposed supplements. Similar metabolites (amino acids, glucose, lipids and nucleotides) were found in the hUCBP and FBS NMR spectra. The results show that the major difference between the metabolic profiles of the two proposed supplements are due to the significantly higher levels of glucose and lower levels of lactate, glutamate, alanine and branched chain amino acids in hUCBP. Similar or slightly different levels of important proteinogenic amino acids, as well as of nucleotides, lipids were found in the hUCBP and FBS. In order to validate it's suitability for cell culture, umbilical cord-MSCs (UC-MSCs) and dental pulp stem cells (DPSCs) were expanded using hUCBP. In both hMSCs, in vitro culture with hUCBP supplementation presented similar to improved metabolic performances when compared to FBS. The two cell types tested expressed different optimum hUCBP percentage content. For DPSCs, the optimum hUCBP content was 6% and for UC-MSCs, 4%. Cultured hMSCs displayed no changes in senescence indicators, as well as maintained characteristic surface marker's expression. FBS substitution was associated with an increase in early apoptosis events, in a dose dependent manner, as well as to slight up- and down-regulation of targeted gene's expression. Tri-lineage differentiation capacity was also influenced by the substitution of FBS by hUCBP.

Publication types

  • Comparative Study
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Cell Culture Techniques / methods*
  • Cell Differentiation
  • Cells, Cultured
  • Culture Media / chemistry
  • Dental Pulp / cytology
  • Humans
  • In Vitro Techniques
  • Mesenchymal Stem Cells / cytology*
  • Metabolomics / methods*
  • Proton Magnetic Resonance Spectroscopy
  • Serum / chemistry*
  • Stem Cells / cytology
  • Umbilical Cord / chemistry*

Substances

  • Culture Media

Grants and funding

This research was supported by Programa Operacional Regional do Norte (ON.2 – O Novo Norte), QREN, FEDER with the project “iBone Therapies: Terapias inovadoras para a regeneração óssea”, ref. NORTE-01-0247-FEDER-003262, and by the program COMPETE – Programa Operacional Factores de Competitividade, Projects PEst-OE/AGR/UI0211/2011 and PEst-C/EME/UI0285/2013 funding from FCT. This research was also supported by Programa Operacional Competitividade e Internacionalização (P2020), Fundos Europeus Estruturais e de Investimento (FEEI) and FCT with the project “BioMate – A novel bio-manufacturing system to produce bioactive scaffolds for tissue engineering” with reference PTDC/EMS-SIS/7032/2014 and by COMPETE 2020, from ANI – Projectos ID&T Empresas em Copromoção, Programas Operacionais POCI, by the project “insitu.Biomas - Reinvent biomanufacturing systems by using an usability approach for in situ clinic temporary implants fabrication” with the reference POCI-01-0247-FEDER-017771. Ana Rita Caseiro (SFRH/BD/101174/2014) acknowledges FCT, for financial support. This work received financial support from the framework of QREN through Project NORTE-07-0124-FEDER-000066. The Bruker Avance III 600 HD spectrometer was purchased under the framework of QREN, through Project NORTE-07-0162-FEDER-000048, and is part of the Portuguese NMR Network created with support of FCT through Contract REDE/1517/RMN/2005, with funds from POCI 2010 (FEDER). PP Barbosa is an employee of Biosckin, Molecular and Cell Therapies, S.A., and she played a role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. Biosckin, Molecular and Cell Therapies, S.A., provided support in the form of salaries for authors [PP Barbosa], but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. The specific roles of these authors are articulated in the ‘author contributions’ section.