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T helper 2 cytokines induce preproenkephalin mRNA expression and proenkephalin A in human peripheral blood
S Kamphuis1, A Kavelaars, R Brooimans
1Department of Immunology, University Hospital for Children and Youth, Het Wilhelmina Kinderziekenhuis, Utrecht, The Netherlands. s.kamphuis@wkz.ruu.nl
Abstract:
We investigated the regulatory influence of several cytokines on the expression of preproenkephalin (PPE) mRNA in human peripheral blood mononuclear cells (PBMC). By use of a quantitative reverse-transcriptase-polymerase chain reaction (RT-PCR), we demonstrate that the T helper 2 cytokines IL-4 and IL-10 are more potent in upregulating PPE mRNA expression in human PBMC than the T helper 1 cytokines IL-2 and gamma-IFN. In addition, TGF-beta is also an effective inducer of PPE mRNA. TGF-beta, IL-4 and IL-10 increase the cytoplasmatic concentration of met-enkephalin in PBMC. Secretion of met-enkephalin in the culture supernatant of IL-4- or IL-10-stimulated PBMC could not be observed, but proenkephalin A-derived met-enkephalin containing peptides could be demonstrated. IL-4 and IL-10 do not induce PPE mRNA via the same pathways. We could observe that PKA is involved in IL-4 mediated PPE mRNA induction, whereas IL-10 apparently uses another route.
Insights
T helper 2 cytokines, Interleukin-4 (IL-4) and Interleukin-10 (IL-10), significantly upregulate preproenkephalin (PPE) mRNA in human peripheral blood mononuclear cells (PBMC). These cytokines, along with TGF-beta, increase intracellular met-enkephalin levels.
Area of Science:
- Immunology
- Neuroscience
- Molecular Biology
Background:
- Cytokines play crucial roles in immune regulation and cellular communication.
- Preproenkephalin (PPE) is a precursor to opioid peptides like met-enkephalin, involved in pain modulation and stress response.
- Understanding cytokine-mediated regulation of PPE is vital for exploring neuro-immune interactions.
Purpose of the Study:
- To investigate the differential effects of T helper 1 (Th1) and T helper 2 (Th2) cytokines on PPE mRNA expression in human peripheral blood mononuclear cells (PBMC).
- To determine the role of transforming growth factor-beta (TGF-beta) in regulating PPE mRNA and met-enkephalin levels.
- To elucidate the intracellular signaling pathways involved in IL-4 and IL-10 induced PPE mRNA expression.
Main Methods:
- Quantitative reverse-transcriptase-polymerase chain reaction (RT-PCR) was employed to measure PPE mRNA levels.
- Human peripheral blood mononuclear cells (PBMC) were stimulated with various cytokines, including IL-4, IL-10, IL-2, gamma-interferon (IFN-γ), and TGF-beta.
- Intracellular and secreted met-enkephalin levels were assessed in stimulated PBMC cultures.
Main Results:
- Th2 cytokines (IL-4 and IL-10) were more potent in upregulating PPE mRNA expression than Th1 cytokines (IL-2 and gamma-IFN).
- TGF-beta also effectively induced PPE mRNA expression.
- IL-4, IL-10, and TGF-beta increased intracellular met-enkephalin concentrations in PBMC, with evidence of proenkephalin A-derived peptides in the supernatant.
- Distinct signaling pathways were implicated, with protein kinase A (PKA) involved in IL-4-mediated induction, while IL-10 utilized a different route.
Conclusions:
- Th2-biased cytokine environments, characterized by IL-4 and IL-10, significantly promote PPE mRNA expression in human PBMC.
- TGF-beta also contributes to the upregulation of PPE mRNA and subsequent met-enkephalin production.
- The findings highlight differential cytokine signaling pathways regulating opioid peptide precursor expression, offering insights into neuro-immune system crosstalk.