Sources of variability in quantifying circulating thymosin beta-4: literature review and recommendations

ABSTRACT

Introduction: Thymosin beta-4 (TB4) is an endogenous peptide with protective and regenerative effects in models of cellular and organ injury. TB4 is increasingly measured as a potential plasma or serum biomarker in human cardiovascular, liver, infectious, and autoimmune disease.

Areas covered: The focus of this review is the quantification of TB4 in clinical cohort studies and whether reported TB4 concentrations differ with respect to method of sample preparation. We survey current literature for studies measuring TB4 in human serum or plasma and compare reported concentrations in healthy controls.

Expert opinion: We find substantial intra- and inter- study variability in healthy controls, and a lack of protocol standardization. We further highlight three factors that may confound TB4 clinical measurements and should be considered in future study design: 1) residual platelets remaining in suspension after centrifugation, 2) TB4 release following ex vivo platelet activation, and 3) specificity of assays towards posttranslational modifications. Accordingly, we put forth our recommendations to minimize residual and activated platelets during sample collection, and to cross-validate TB4 measurements using both antibody-based and mass spectrometry-based methods.

KEYWORDS:

• Thymosin beta-4
• plasma
• serum
• biomarker
• platelets

Article highlights

• The endogenous peptide thymosin beta-4 is a well-studied protective and regenerative biomolecule.

• Human serum and plasma TB4 has been measured as a biomarker for various diseases including cardiovascular disease, rheumatoid arthritis, sepsis and liver diseases.

• Comparing measurements of controls across studies, there is significant inter- and intra-study variability.

• Platelets contain a reservoir of TB4. Residual platelets in plasma and platelet activation in serum may artificially elevate the apparent TB4 concentration.

• We put forth our recommendation to reduce variability and confounders from ex vivo platelets and suggest that disease-specific posttranslational modifications of TB4 may be important for future study of TB4 as biomarker.

This box summarizes key points contained in the article.

Acknowledgments

We appreciate the assistance of friends and colleagues in proofreading manuscript drafts: Dr Lin Xinyi, Cindy (Duke-NUS Graduate Medical School), Mr Jackson Low (CVRI, NUS), Mr Siddharth Deshpande (NGS, NUS), and Mr Sherman Yee (CVRI, NUS).

Declaration of interest

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties. Peer reviewers on this manuscript have no relevant financial or other relationships to disclose.

Supplemental data

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Additional information

Funding

This paper received funding from the Agency for Science, Technology and Research [SPF2014/001] and the National Medical Research Council of the Singapore Ministry of Health [NMRC/CG/014/2013]. W.K.Y. Tan is financially supported by a PhD scholarship from the NUS Graduate School for Integrative Sciences and Engineering (NGS). This paper has been published as part of a supplement issue covering the proceedings of the Fifth International Symposium on Thymosins in Health and Disease and is funded by SciClone Pharmaceuticals.