title: Targeted Deletion of Autophagy Genes Atg5 or Atg7 in the Chondrocytes Promotes Caspase-Dependent Cell Death and Leads to Mild Growth Retardation.
abstract: Longitudinal bone growth takes place in epiphyseal growth plates located in the ends of long bones. The growth plate consists of chondrocytes traversing from the undifferentiated (resting zone) to the terminally differentiated (hypertrophic zone) stage. Autophagy is an intracellular catabolic process of lysosome-dependent recycling of intracellular organelles and protein complexes. Autophagy is activated during nutritionally depleted or hypoxic conditions in order to facilitate cell survival. Chondrocytes in the middle of the growth plate are hypoxic and nutritionally depleted owing to the avascular nature of the growth plate. Accordingly, autophagy may facilitate their survival. To explore the role of autophagy in chondrocyte survival and constitutional bone growth, we generated mice with cartilage-specific ablation of either Atg5 (Atg5cKO) or Atg7 (Atg7cKO) by crossing Atg5 or Atg7 floxed mice with cartilage-specific collagen type 2 promoter-driven Cre. Both Atg5cKO and Atg7cKO mice showed growth retardation associated with enhanced chondrocyte cell death and decreased cell proliferation. Similarly, inhibition of autophagy by Bafilomycin A1 (Baf) or 3-methyladenine (3MA) promoted cell death in cultured slices of human growth plate tissue. To delineate the underlying mechanisms we employed ex vivo cultures of mouse metatarsal bones and RCJ3.IC5.18 rat chondrogenic cell line. Baf or 3MA impaired metatarsal bone growth associated with processing of caspase-3 and massive cell death. Similarly, treatment of RCJ3.IC5.18 chondrogenic cells by Baf also showed massive cell death and caspase-3 cleavage. This was associated with activation of caspase-9 and cytochrome C release. Altogether, our data suggest that autophagy is important for chondrocyte survival, and inhibition of this process leads to stunted growth and caspase-dependent death of chondrocytes.
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Karuna K
authors.initials: KK
authors.lastname: Vuppalapati
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Thibault
authors.initials: T
authors.lastname: Bouderlique
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.|Department of Women's and Children's Health, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Phillip T
authors.initials: PT
authors.lastname: Newton
authors.affiliation: Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Vitaliy O
authors.initials: VO
authors.lastname: Kaminskyy
authors.affiliation: Pediatric Orthopedic Clinic, Karolinska University Hospital, Stockholm, Sweden.
authors.forename: Henrik
authors.initials: H
authors.lastname: Wehtje
authors.affiliation: Centre for Bone and Arthritis Research, Institute of Medicine, Sahlgrenska Academy, University of Gothenburg, Gothenburg, Sweden.
authors.forename: Claes
authors.initials: C
authors.lastname: Ohlsson
authors.affiliation: Institute of Environmental Medicine, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Boris
authors.initials: B
authors.lastname: Zhivotovsky
authors.affiliation: Department of Physiology and Pharmacology, Karolinska Institutet, Stockholm, Sweden.
authors.forename: Andrei S
authors.initials: AS
authors.lastname: Chagin
mesh_terms: D000225:Adenine / Q000031:analogs & derivatives / Q000378:metabolism; D000818:Animals; D017209:Apoptosis; D001343:Autophagy; D000071187:Autophagy-Related Protein 5; D000071193:Autophagy-Related Protein 7; D020169:Caspases / Q000378:metabolism*; D016923:Cell Death; D002460:Cell Line; D049109:Cell Proliferation; D002470:Cell Survival; D019902:Chondrocytes / Q000166:cytology*; D024043:Collagen Type II / Q000378:metabolism; D045304:Cytochromes c / Q000378:metabolism; D017353:Gene Deletion; D006132:Growth Plate / Q000254:growth & development; D006801:Humans; D000860:Hypoxia; D007150:Immunohistochemistry; D017403:In Situ Hybridization; D018942:Macrolides / Q000378:metabolism; D008682:Metatarsal Bones / Q000378:metabolism; D051379:Mice; D018345:Mice, Knockout; D008869:Microtubule-Associated Proteins / Q000235:genetics* / Q000502:physiology; D011506:Proteins / Q000235:genetics; D051381:Rats; D044764:Ubiquitin-Activating Enzymes / Q000235:genetics
chemical_list: C495773:Atg5 protein, mouse; C545865:Atg5 protein, rat; C502661:Atg7 protein, mouse; C561242:Atg7 protein, rat; D000071187:Autophagy-Related Protein 5; D024043:Collagen Type II; D018942:Macrolides; D008869:Microtubule-Associated Proteins; D011506:Proteins; C025946:3-methyladenine; C040929:bafilomycin A1; D045304:Cytochromes c; D020169:Caspases; D000071193:Autophagy-Related Protein 7; D044764:Ubiquitin-Activating Enzymes; D000225:Adenine
keywords: APOPTOSIS; AUTOPHAGY; GROWTH PLATE; BONE; CHONDROCYTE