Specific binding of soluble peptidoglycan and muramyldipeptide to CD14 on human monocytes.
Previously, we were able to show that soluble peptidoglycan (sPG)-induced monokine production in human peripheral monocytes is inhibited by anti-CD14 monoclonal antibodies and by lipid A partial structures. This suggested but did not prove that monocytic surface protein CD14 is involved in the activation of human monocytes not only by cell wall components of gram-negative bacteria such as lipopolysaccharide (LPS) but also by cell wall components of gram-positive bacteria such as sPG. In the present study, we provide experimental evidence that CD14 indeed constitutes a binding site for sPG recognition and activation of human monocytes. The results show that fluorescein isothiocyanate-sPG (FITC-sPG) binds to human monocytes in a saturable, dose-dependent, and specific manner. For maximal binding, 2 to 3 microg of FITC-sPG per ml was sufficient, and this binding is completed within 90 min; about 40% of the binding is completed within the first 3 min. The FITC-sPG binding is considered specific because unlabeled sPG and also muramyldipeptide (MDP), the minimal bioactive structure of sPG, inhibit the binding of sPG to monocytes in a dose-dependent manner. This specific binding was also inhibited by an anti-CD14 monoclonal antibody, LPS, and lipid A partial structure compound 406. Direct evidence for an interaction of sPG with CD14 is provided by experiments involving native polyacrylamide gel electrophoresis that showed a shift of the electrophoretic mobility of CD14 by LPS as well as by sPG. These results allow the conclusion that sPG binds directly to CD14, that MDP represents the active substructure of sPG, and that CD14 may be a lectin-like receptor which plays a key role in cellular stimulation by bioactive components of not only gram-negative but also gram-positive bacteria.
Weidemann
B
B
Research Center Borstel, Center for Medicine and Biosciences, Borstel, Germany.
Schletter
J
J
Dziarski
R
R
Kusumoto
S
S
Stelter
F
F
Rietschel
E T
ET
Flad
H D
HD
Ulmer
A J
AJ
D000119:Acetylmuramyl-Alanyl-Isoglutamine / Q000378:metabolism*; D000818:Animals; D001665:Binding Sites; D016650:Fluorescein-5-isothiocyanate; D006801:Humans; D015850:Interleukin-6 / Q000096:biosynthesis; D018950:Lipopolysaccharide Receptors / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D051379:Mice; D009000:Monocytes / Q000378:metabolism*; D010457:Peptidoglycan / Q000378:metabolism*; D011112:Polymyxin B / Q000494:pharmacology
D015850:Interleukin-6; D018950:Lipopolysaccharide Receptors; D008070:Lipopolysaccharides; D010457:Peptidoglycan; D000119:Acetylmuramyl-Alanyl-Isoglutamine; D016650:Fluorescein-5-isothiocyanate; D011112:Polymyxin B