TY - JOUR
T1 - Localization of the NO-cGMP signaling pathway molecules, NOS III-phosphorylation sites, ERK1/2, and Akt/PKB in osteoclasts
AU - Korkmaz, Yüksel
AU - Baumann, Michael A
AU - Schröder, Hannsjörg
AU - Behrends, Sönke
AU - Addicks, Klaus
AU - Raab, Wolfgang H M
AU - Bloch, Wilhelm
PY - 2004/8/1
Y1 - 2004/8/1
N2 - BACKGROUND: Nitric oxide (NO) mediates different cellular functions by activating soluble guanylate cyclase (sGC) that converts guanosine-5'-triphosphate (GTP) to cyclic guanosine-3',5'-monophosphate (cGMP). Membrane-bound GCs produce cGMP in response to natriuretic peptides in osteoblasts, but neither the NO-target enzyme sGC, nor the phosphorylation sites of NOS III, nor their regulation by extracellular signal-regulated kinases 1 and 2 (ERK1/2) and Akt/protein kinase B (Akt/PKB) in osteoclasts have been established.METHODS: Rat molars with periodontium were perfusion- and post-fixed, decalcified, and frozen-sectioned. Free-floating sections were stained using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and tartrate-resistant acid phosphatase (TRAP) histochemical techniques and immunoreacted with antisera against NO-synthase (NOS) I-III, NOS III phoshorylated at Thr495, NOS III phoshorylated at Serine1177 (Ser1177), ERK1/2, phosphorylated ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2/beta1), and cGMP.RESULTS: NADPH-d staining and immunostaining of NOS I-III, NOS III phosphorylated at Ser1177, ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2 and beta1-subunits), and cGMP were detected in osteoclasts. Immunohistochemical reaction products for NOS III phosphorylated at threonine495 (Thr495) and phosphorylated ERK1/2 could not be identified in osteoclasts. Comparison of TRAP activity and immunostaining for sGC beta1-subunit revealed that sGC beta1-subunit is only expressed in a sub-population of osteoclasts.CONCLUSIONS: NO is likely to be generated by NOS I and NOS III in osteoclasts. The inconstant expression of NOS II in some osteoclasts may be explained with inducible expression of NOS II upon physiological cell activation. Localization of the sGC alpha2- and beta1-subunits and cGMP in osteoclasts is compatible with an involvement of NO-sGC signaling in the homeostasis of osteoclasts. The phosphorylation site of NOS III at Ser1177 and phosphorylated Akt/PKB are involved in regulation of NO production by NOS III in osteoclasts under basal conditions.
AB - BACKGROUND: Nitric oxide (NO) mediates different cellular functions by activating soluble guanylate cyclase (sGC) that converts guanosine-5'-triphosphate (GTP) to cyclic guanosine-3',5'-monophosphate (cGMP). Membrane-bound GCs produce cGMP in response to natriuretic peptides in osteoblasts, but neither the NO-target enzyme sGC, nor the phosphorylation sites of NOS III, nor their regulation by extracellular signal-regulated kinases 1 and 2 (ERK1/2) and Akt/protein kinase B (Akt/PKB) in osteoclasts have been established.METHODS: Rat molars with periodontium were perfusion- and post-fixed, decalcified, and frozen-sectioned. Free-floating sections were stained using nicotinamide adenine dinucleotide phosphate-diaphorase (NADPH-d) and tartrate-resistant acid phosphatase (TRAP) histochemical techniques and immunoreacted with antisera against NO-synthase (NOS) I-III, NOS III phoshorylated at Thr495, NOS III phoshorylated at Serine1177 (Ser1177), ERK1/2, phosphorylated ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2/beta1), and cGMP.RESULTS: NADPH-d staining and immunostaining of NOS I-III, NOS III phosphorylated at Ser1177, ERK1/2, Akt/PKB, phosphorylated Akt/PKB, sGC (alpha2 and beta1-subunits), and cGMP were detected in osteoclasts. Immunohistochemical reaction products for NOS III phosphorylated at threonine495 (Thr495) and phosphorylated ERK1/2 could not be identified in osteoclasts. Comparison of TRAP activity and immunostaining for sGC beta1-subunit revealed that sGC beta1-subunit is only expressed in a sub-population of osteoclasts.CONCLUSIONS: NO is likely to be generated by NOS I and NOS III in osteoclasts. The inconstant expression of NOS II in some osteoclasts may be explained with inducible expression of NOS II upon physiological cell activation. Localization of the sGC alpha2- and beta1-subunits and cGMP in osteoclasts is compatible with an involvement of NO-sGC signaling in the homeostasis of osteoclasts. The phosphorylation site of NOS III at Ser1177 and phosphorylated Akt/PKB are involved in regulation of NO production by NOS III in osteoclasts under basal conditions.
KW - Alveolar Process
KW - Animals
KW - Cyclic GMP
KW - Guanylate Cyclase
KW - Immunoenzyme Techniques
KW - Male
KW - Nitric Oxide
KW - Nitric Oxide Synthase
KW - Osteoclasts
KW - Phosphorylation
KW - Protein-Serine-Threonine Kinases
KW - Rats
KW - Rats, Wistar
KW - Second Messenger Systems
U2 - 10.1902/jop.2004.75.8.1119
DO - 10.1902/jop.2004.75.8.1119
M3 - Journal articles
C2 - 15455741
SN - 0022-3492
VL - 75
SP - 1119
EP - 1125
JO - Journal of periodontology
JF - Journal of periodontology
IS - 8
ER -