Lentiviral transduction of primary mouse bone marrow monocytes using ViroMag R/L

Mouse primary non-adherent bone marrow monocytes (BMM) were transduced with 10 µL of Lentiviral particles (5x10.7/mL) combined to 2 µL of ViroMag R/L.

This paper shows the efficiency of ViroMag R/L from OZ Biosciences to transduce non primary mouse adherent bone marrow monocytes with lentivirus.

article reference: J Bone Miner Res. 2013 Jun 21. 

TBK1 mediates critical effects of measles virus nucleocapsid protein (MVNP) on pagetic osteoclast formation.
Sun Q, Sammut B, Wang FM, Kurihara N, Windle JJ, Roodman GD, Galson DL.

Abstract

Paget's disease of bone (PDB) is characterized by abnormal osteoclasts with unique characteristics that include: increased sensitivity of osteoclast progenitors to 1,25(OH)₂ D₃ , RANKL and TNF-α, increased osteoclast numbers, increased expression of IL-6 and several transcription factors. We recently reported that measles virus nucleocapsid protein (MVNP) plays a key role in the development of these abnormal osteoclasts. MVNP can induce the pagetic osteoclast phenotype in vitro and in vivo in TRAP-MVNP transgenic mice. However, the molecular mechanisms by which MVNP generates pagetic osteoclasts have not been determined. TANK-binding kinase 1 (TBK1) and IκB kinase-ϵ (IKKϵ) are IKK family members which complex with MVNP and activate both IRF3 and NF-κB pathways. MVNP increases the amount of TBK1 protein in bone marrow monocytes (BMM). Interestingly, we found that RANKL increased TBK1 and IKKϵ early in osteoclast differentiation, suggesting a possible role in normal osteoclastogenesis. However, only TBK1 is further increased in osteoclasts formed by TRAP-MVNP BMM due to increased TBK1 protein stability. TBK1 over-expression induced IL6 promoter reporter activity, and elevated endogenous IL6 mRNA and p65 NF-κB, TAF12 and ATF7 proteins in several cell lines. Over-expression of TBK1 was insufficient to induce pagetic osteoclasts from WT BMM, but synergized with MVNP to increase pagetic osteoclast formation from TRAP-MVNP BMM. BX795 inhibition of TBK1 impaired MVNP-induced IL-6 expression in both NIH3T3 cells and BMM, and shRNA knockdown of Tbk1 in NIH3T3 cells impaired IL-6 secretion induced by MVNP and decreased TAF12 and ATF7, factors involved in 1,25(OH)₂ D₃ hypersensitivity of pagetic osteoclasts. Similarly, Tbk1 knockdown in BMM from TRAP-MVNP and WT mice specifically impaired development of the MVNP-induced osteoclast pagetic phenotype. These results demonstrate that TBK1 plays a critical role in mediating the effects of MVNP on osteoclast differentiation, and on the expression of IL-6, a key contributor to the pagetic osteoclast phenotype.

ViroMag R/L from OZ Biosciences is a magnetic nanoparticles formulation optimized for increasing any kind of virus infection and transduction both in vitro and in vivo.