Inhibition of protein kinase-D promotes cartilage repair at injured growth plate in rats Rosa Chung a,b , Bruce K. Foster c , Cory J. Xian a,b, * a Sansom Institute for Health Research, School of Pharmacy and Medical Sciences, University of South Australia, Adelaide, SA 5001, Australia b Discipline of Physiology, School of Medical Sciences, University of Adelaide, Adelaide, SA 5005, Australia c Department of Orthopaedic Surgery, Women’s and Children’s Hospital, North Adelaide, SA 5006, Australia Introduction Being a large cartilaginous structure at the ends of all long bones in children, the growth plate is the most fragile region in the bone itself. Hence, growth plate related injuries are common in children, and, depending on the severity and location of the injuries, up to 30% of them are repaired by bony tissue that can result in orthopaedic problems such as limb length discrepancy and bone angulation deformities. 1 Out of the possible five clinically recognised Salter-Harris injury types, long term complications arise often from those with Type III-V growth plate injuries. 2,3 Currently, highly invasive and sometime ineffective surgical techniques are implemented as corrective procedures for growth plate injury–induced bone defects, and there are no known biological therapies for preventing these bone growth defects. 4,5 At present, molecular mechanisms for the undesirable bony repair of the injured growth plate remain unclear, and under- standing the molecular events occurring during the bony repair would be invaluable towards the development of a potential biological therapy. Using a full thickness rat tibial growth plate injury model, previous studies have identified four distinct phases of growth plate repair responses which are similar to repair characteristics clinically reported following a Salter-Harris Type III-V growth plate injury, 6,7 namely the initial inflammatory phase, the fibrogenic, the osteogenic, and the bone remodelling phases. After the inflammatory event and during the fibrogenic phase, there is an influx of mesenchymal stromal cells entering the injury Injury, Int. J. Care Injured 44 (2013) 914–922 A R T I C L E I N F O Article history: Accepted 27 January 2013 Keywords: Growth plate Injury repair Osteoblasts Stromal progenitor cells Differentiation Protein kinase-D Osterix A B S T R A C T Introduction: Injured growth plate cartilage is often repaired by bony tissue, causing bone growth defects in children. Currently, mechanisms for the undesirable repair remain unclear and there are no biological treatments available to prevent the associated bone growth defects. Osterix is known as a vital transcription factor for osteoblast differentiation which is critical for normal bone formation and bone repair, and osterix is known to be regulated by protein kinase-D; however it is unknown whether protein kinase-D–osterix signalling plays any roles in the bony repair of injured growth plate. Methods: Using a rat model, this study investigated potential roles of protein kinase-D (PKD) in regulating expression of osteogenic transcription factor osterix and the growth plate bony repair. 4 days post injury at the proximal tibial growth plate, rats received four once-daily injections of vehicle or 2.35 mg/kg go ¨ 6976 (a PKD inhibitor), and growth plate tissues collected at day 10 were examined histologically and molecularly. In addition, effects of PKD inhibition on osteogenic and chondrogenic differentiation were examined in vitro using rat bone marrow mesenchymal stromal cells. Results: Compared to vehicle control, PKD inhibition caused a decrease in bone volume (p < 0.05), an increase in % of mesenchymal tissue (p < 0.01), and an increase in cartilaginous tissue within the injury site. Consistently, go ¨ 6976 treatment tended to decrease expression of bone-related genes (osterix, osteocalcin) and increase levels of cartilage-related genes (Sox9, collagen-2a, collagen-10a1). In support, in vitro experiments showed that go ¨ 6976 presence in the primary rat marrow stromal cell culture resulted in a decrease of alkaline phosphatase + CFU-f colonies formed (p < 0.05) and an increase in collagen-2a expression in chondrogenic pellet culture (p < 0.05). Conclusion: These studies suggest that PKD is important for growth plate bony repair and its inhibition after growth plate injury may result in less bone formation and potentially more cartilage repair. ß 2013 Elsevier Ltd. All rights reserved. * Corresponding author at: Sansom Institute for Health Research, University of South Australia, City East Campus, GPO Box 2471, Adelaide 5001, Australia. Tel.: +61 8 83021944; fax: +61 8 8302 1087. E-mail addresses: rosa.chung@unisa.edu.au (R. Chung), bruce.foster@health.sa.gov.au (B.K. Foster), cory.xian@unisa.edu.au (C.J. Xian). Contents lists available at SciVerse ScienceDirect Injury jo ur n al ho m epag e: ww w.els evier .c om /lo cat e/inju r y 0020–1383/$ – see front matter ß 2013 Elsevier Ltd. All rights reserved. http://dx.doi.org/10.1016/j.injury.2013.01.038