MyD88 But Not TRIF Is Essential for Osteoclastogenesis Induced by Lipopolysaccharide, Diacyl Lipopeptide, and IL-1α

Abstract

Myeloid differentiation factor 88 (MyD88) plays essential roles in the signaling of the Toll/interleukin (IL)-1 receptor family. Toll–IL-1 receptor domain-containing adaptor inducing interferon-β (TRIF)-mediated signals are involved in lipopolysaccharide (LPS)-induced MyD88-independent pathways. Using MyD88-deficient (MyD88−/−) mice and TRIF-deficient (TRIF−/−) mice, we examined roles of MyD88 and TRIF in osteoclast differentiation and function. LPS, diacyl lipopeptide, and IL-1α stimulated osteoclastogenesis in cocultures of osteoblasts and hemopoietic cells obtained from TRIF−/− mice, but not MyD88−/− mice. These factors stimulated receptor activator of nuclear factor-κB ligand mRNA expression in TRIF−/− osteoblasts, but not MyD88−/− osteoblasts. LPS stimulated IL-6 production in TRIF−/− osteoblasts, but not TRIF−/− macrophages. LPS and IL-1α enhanced the survival of TRIF−/− osteoclasts, but not MyD88−/− osteoclasts. Diacyl lipopeptide did not support the survival of osteoclasts because of the lack of Toll-like receptor (TLR)6 in osteoclasts. Macrophages expressed both TRIF and TRIF-related adaptor molecule (TRAM) mRNA, whereas osteoblasts and osteoclasts expressed only TRIF mRNA. Bone histomorphometry showed that MyD88−/− mice exhibited osteopenia with reduced bone resorption and formation. These results suggest that the MyD88-mediated signal is essential for the osteoclastogenesis and function induced by IL-1 and TLR ligands, and that MyD88 is physiologically involved in bone turnover.

Rab27a and Rab27b are involved in stimulation-dependent RANKL release from secretory lysosomes in osteoblastic cells

“…This is in marked contrast to the mechanism of lytic granule secretion in hematopoietic cells, in which Slp1, Slp2-a, and Munc13-4, but not Slp4-a and Slp5, have been reported to be involved. (28,35) As mentioned earlier, there are functional differences between Rab27a and Rab27b. There are reports suggesting that the Rab27 isoforms are functionally redundant and can compensate for each other.…”

“…Primary osteoblastic cells (POBs) were collected from the calvaria of 2-to 3-day-old wild-type C57BL6 or Jinx mice using previously reported methods. (35) RNA interference RNA interference was used to suppress Rab27a, Rab27b, and their effector molecules by transient transfection with specific siRNAs directed against each gene using Lipofectamine 2000 (Invitrogen) following the manufacturer's protocol. The sequences selected as targets for each gene were designed by BLOCK-iT RNAi Designer on the Web (Invitrogen) and are as follows: 5'-GCU UAA CCA CUG CAU UCU U-3' for Rab27a, 5'-CCA GUC AAC AGA GCU UCU U-3' for Rab27b, 5'-CCA UCU CAG GAG AGG CUU U-3' for Slp1, 5'-GCG UUU ACA GUG GAG ACU U-3' for Slp2-a, 5'-GCU UGC UUC CCA GGU GAA U-3' for Slp4-a, 5'-GCG UUU CAA GCA AGU CAA U-3' for Slp5, and 5'-GGA GAA CUU CAG CAG CCU U-3' for Munc13-4.…”

Function of OPG as a traffic regulator for RANKL is crucial for controlled osteoclastogenesis

“…(20) Bone marrow cells obtained from the tibias of 6-to 8-week-old male C57/BL6 mice were cultured in a-MEM containing 10% FBS in the presence of macrophage colony-stimulating factor (M-CSF; 5 ng/mL; Cosmo Bio, Tokyo, Japan) using a previously described method with some modifications. (21) Nonadherent cells were harvested and used in the coculture assay with ST2 or primary osteoblastic cells.…”

Lipopolysaccharide-Induced Osteoclastogenesis from Mononuclear Precursors: A Mechanism for Osteolysis in Chronic Otitis

“…2E). LPS is well known to elicit TNF-α (Abu-Amer et al 1997;Zou and Bar-Shavit 2002;Itoh et al 2003), IL-6 (Itoh et al 2003;Sato et al 2004), IL-1α (Hong et al 2004), andIL-1β (Itoh et al 2003) release from monocytes. It is also well established that these cytokines promote Oc formation (Jimi et al 1999;Azuma et al 2000;Zhang et al 2001;Zou et al 2001;Kudo et al 2003;Wei et al 2005).…”