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Met-enkephalin-containing peptides encoded by proenkephalin A mRNA expressed in activated murine thymocytes inhibit
K M Linner1, H E Quist, B M Sharp
1Minneapolis Medical Research Foundation, MN 55404, USA.
Abstract:
Murine thymocytes activated with the mitogen Con A express proenkephalin A mRNA (PEA mRNA) and met-enkephalin and/or met-enkephalin-containing peptides ("enkephalins"). This Con A-induced expression of PEA mRNA is modulated by the delta opioid receptor agonist, deltorphin I, in a biphasic, dose-dependent manner. That is, 10(-13) M to 10(-11) M deltorphin enhanced PEA mRNA expression 3- to 3.5-fold over the level induced by Con A alone, and 10(-9) M to 10(-7) M deltorphin inhibited it 40 to 70%. delta opioid receptor antagonists recognizing the delta-2 (naltrindole (NTI) and naltriben (NTB)), but not the delta-1 (7-benzylidenenaltrexone (BNTX)), subtype of opioid receptor described in brain, reversed both the enhancing and inhibiting effects of deltorphin on Con A-induced PEA mRNA expression. In addition, the delta-2 receptor-specific antagonists, NTI and NTB, directly inhibited Con A-induced PEA mRNA expression. The function of the enkephalins expressed by thymocytes was examined by using 1) delta opioid receptor antagonists, 2) PEA mRNA-specific antisense cDNA, and 3) Ab to met-enkephalin, and measuring cell proliferation. All three reagents caused enhancement of Con A-induced proliferation, with effects ranging from two- to fourfold over the response to Con A alone. Again, the delta-2 subtype-specific antagonists, NTI and NTB, were functional and the delta-1 subtype-specific antagonist, BNTX, was not. The PEA mRNA-specific antisense cDNA blocked translation but not transcription of PEA mRNA. The data suggest that 1) endogenous enkephalins induced in thymocytes modulate their own expression through delta-2-like opioid receptors, and 2) these endogenous enkephalins function to inhibit the proliferation of activated thymocytes.
Insights
Activated mouse thymocytes produce enkephalins, which are opioid peptides. These enkephalins regulate their own production via delta-2 opioid receptors and inhibit thymocyte proliferation.
Area of Science:
- Immunology
- Neuroendocrinology
- Molecular Biology
Background:
- Murine thymocytes, upon activation with Concanavalin A (Con A), express proenkephalin A mRNA (PEA mRNA) and resulting enkephalins.
- The delta opioid receptor system plays a role in immune cell function and peptide expression.
Purpose of the Study:
- To investigate the modulation of Con A-induced PEA mRNA expression and thymocyte proliferation by opioid receptor agonists and antagonists.
- To elucidate the role of endogenous enkephalins in regulating thymocyte activation and proliferation.
Main Methods:
- Treatment of murine thymocytes with Con A and varying concentrations of deltorphin I (a delta opioid receptor agonist).
- Assessment of PEA mRNA levels using molecular techniques.
- Utilizing delta opioid receptor antagonists (naltrindole, naltriben, 7-benzylidenenaltrexone) specific for delta-1 and delta-2 subtypes.
- Measuring thymocyte proliferation in response to Con A and enkephalin-modulating agents.
Main Results:
- Deltorphin I exhibited biphasic modulation of PEA mRNA expression: enhancement at low concentrations and inhibition at higher concentrations.
- Delta-2 selective antagonists (naltrindole, naltriben) reversed deltorphin's effects and directly inhibited Con A-induced PEA mRNA expression.
- Blocking enkephalin function via antagonists, antisense cDNA, or antibodies enhanced Con A-induced thymocyte proliferation.
Conclusions:
- Endogenous enkephalins in thymocytes modulate their own expression through delta-2-like opioid receptors.
- These endogenous enkephalins function as inhibitors of activated thymocyte proliferation, suggesting an autocrine regulatory mechanism.