Alpha B-Crystallin Protects Rat Articular Chondrocytes against Casein Kinase II Inhibition-Induced Apoptosis

PLoS One. 2016 Nov 16;11(11):e0166450. doi: 10.1371/journal.pone.0166450. eCollection 2016.

Abstract

Although alpha (α)B-crystallin is expressed in articular chondrocytes, little is known about its role in these cells. Protein kinase casein kinase 2 (CK2) inhibition induces articular chondrocyte death. The present study examines whether αB-crystallin exerts anti-apoptotic activity in articular chondrocytes. Primary rat articular chondrocytes were isolated from knee joint slices. Cells were treated with CK2 inhibitors with or without αB-crystallin siRNA. To examine whether the silencing of αB-crystallin sensitizes rat articular chondrocytes to CK2 inhibition-induced apoptosis, we assessed apoptosis by performing viability assays, mitochondrial membrane potential measurements, flow cytometry, nuclear morphology observations, and western blot analysis. To investigate the mechanism by which αB-crystallin modulates the extent of CK2 inhibition-mediated chondrocyte death, we utilized confocal microscopy to observe the subcellular location of αB-crystallin and its phosphorylated forms and performed a co-immunoprecipitation assay to observe the interaction between αB-crystallin and CK2. Immunochemistry was employed to examine αB-crystallin expression in cartilage obtained from rats with experimentally induced osteoarthritis (OA). Our results demonstrated that silencing of αB-crystallin sensitized rat articular chondrocytes to CK2 inhibitor-induced apoptosis. Furthermore, CK2 inhibition modulated the expression and subcellular localization of αB-crystallin and its phosphorylated forms and dissociated αB-crystallin from CK2. The population of rat articular chondrocytes expressing αB-crystallin and its phosphorylated forms was reduced in an experimentally induced rat model of OA. In summary, αB-crystallin protects rat articular chondrocytes against CK2 inhibition-induced apoptosis. αB-crystallin may represent a suitable target for pharmacological interventions to prevent OA.

MeSH terms

  • Animals
  • Apigenin / pharmacology
  • Apoptosis* / drug effects
  • Benzimidazoles / pharmacology
  • Cartilage, Articular / cytology*
  • Casein Kinase II / antagonists & inhibitors*
  • Casein Kinase II / metabolism
  • Chondrocytes / cytology*
  • Chondrocytes / drug effects
  • Chondrocytes / metabolism
  • Cytoprotection* / drug effects
  • Disease Models, Animal
  • Gene Knockdown Techniques
  • Gene Silencing / drug effects
  • Male
  • Osteoarthritis / chemically induced
  • Osteoarthritis / metabolism
  • Osteoarthritis / pathology
  • Phenotype
  • Phosphorylation / drug effects
  • Protein Transport / drug effects
  • Rats, Sprague-Dawley
  • Subcellular Fractions / drug effects
  • Subcellular Fractions / metabolism
  • Triazoles / pharmacology
  • alpha-Crystallin B Chain / metabolism*

Substances

  • 4,5,6,7-tetrabromobenzotriazole
  • 5,6-dichlorobenzimidazole
  • Benzimidazoles
  • Triazoles
  • alpha-Crystallin B Chain
  • Apigenin
  • Casein Kinase II

Grant support

This work was supported by grants from the National Research Foundation of Korea (NRF) funded by the Korean government (MSIP) (NRF-2015R1A2A1A10051603, NRF-2013R1A1A4A01006046, and 2016R1A5A2007009).