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Effective Polarization of Human Th17 Cells with Biologically Relevant HumanKine® Recombinant Human TGFβ1, TGFβ2, TGFβ3

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Introduction

Transforming growth factors beta (TGFß) are highly pleiotropic cytokines that act as cellular switches and regulate immune function, proliferation and epithelial-mesenchymal transition. These proteins are produced as precursors then a furin-like convertase processes the proprotein to generate an N-terminal latency-associated peptide (LAP) and a C-terminal mature TGFß. Disulfide-linked homodimers of LAP and TGFß remain non-covalently associated after secretion, forming the small latent TGFß complex. Covalent linkage of LAP to latent TGFß binding proteins creates a large latent complex that may interact with the extracellular matrix. Commercially available TGFß proteins are produced as a recombinant protein expressed in CHO cells or as purified native protein from human platelets. Due to complex post-proteolytic modifications, TGFß yield is low and the products are not available in economic bulk quantity. HumanZyme has developed an efficient human-cell based technology, HumaXpress®, for scalable production of human cytokines and produces TGFß1, ß2, and ß3 from engineered human 293 cells. The proteins are highly purified disulfide-linked dimers of 25kD that can be cost-effectively produced in large scale.

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TH17 Polarization

TGFßs, which are important for the polarization of murine Th17 cells, are reported not required, and are even inhibitory, for human Th17 polarization. In this study, whole CD4+ cells isolated from a healthy donor were stimulated with 10 µg/ml plate bound aniti-CD3 and 10 µg/ml soluble anti-CD28 in the presence of Th17 polarizing cytokines from HumanZyme and another commercial vendor. After 5 days, supernatants were harvested for measurement of IL-17 by ELISA. The results show that all the HumanKine TGFßs are effective at inducing IL-17 secretion with an optimal concentration of 0.1 ng/ml TGFß.

In contrast, TGFß1 from insect cells showed only marginal or even inhibitory effects. The results indicate that using biologically relevant cytokines can more effectively induce Th17 cell polarization and lead to a more accurate scientific understanding of the human biological process.

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