Lipoic acid downmodulates CD4 from human T lymphocytes by dissociation of p56Lck
Section snippets
Materials and methods
Reagents. LA was a gift from Integrative Technologies, Inc., (Wilsonville, OR). DHLA was generously provided by Dr. Rolf Winter at the Portland VA Medical Center (Portland, OR). Additional reagents used included: poly-d-Lysine (P-6407), protease inhibitor cocktail (P 8340), and Con A, which were obtained from Sigma; normal goat serum (NGS; 005-00-001), normal rabbit serum (NRS; 011-000-120), secondary antibodies: Rb anti-mouse HRP (315-035-003), Rb anti-goat HRP (305-035-003), Rb anti-mouse HRP
LA modulation of CD4 from PBMC and human T cell lines
Treatment of freshly isolated human PBMC with LA resulted in a concentration-dependent downmodulation of CD4 from the plasma membrane by LA (Fig. 1). DHLA, the reduced form of LA, treatment of PBMC did not induce downmodulation of CD4 at any concentration tested (Fig. 1). Two additional derivatives of LA, lipoamide (LPM; CONH2 replaces the COOH of LA) and dimethyl lipoic acid (DMLA; SCH3 replaces the sulfhydryl groups of DHLA) also had no effect on CD4 expression (data not shown).
Discussion
Herein we describe the novel observation of CD4 downmodulation by the antioxidant LA. Interestingly, its redox partner, DHLA, did not alter surface expression of CD4. Immunofluorescent analyses with monoclonal anti-CD4 antibodies recognizing different Ig-like domains within the CD4 protein support the conclusion that the CD4 is downmodulated by the LA, as opposed to epitope masking. Confocal microscopic analyses revealed the absence of CD4 from the plasma membrane and the cytosol, consistent
Acknowledgments
This work was supported by the Department of Veterans Affairs, National Institutes of Health AT P50 AT00066-01 and the Nancy Davis Center Without Walls. We wish to thank Drs. Rolf Winter and Michael Riscoe for the preparation of DHLA and Integrative Technologies, Inc. (Wilsonville, OR) for providing LA.
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