A cleft lip and palate gene, Irf6, is involved in osteoblast differentiation of craniofacial bone

Abstract

Background: Interferon regulatory factor 6 (IRF6) plays a critical role in embryonic tissue development, including differentiation of epithelial cells. Besides orofacial clefting due to haploinsufficiency of IRF6, recent human genetic studies indicated that mutations in IRF6 are linked to small mandible and digit abnormalities. The function of IRF6 has been well studied in oral epithelium; however, its role in craniofacial skeletal formation remains unknown. In this study, we investigated the role of Irf6 in craniofacial bone development using comparative analyses between wild‐type (WT) and Irf6‐null littermate mice. Results: Immunostaining revealed the expression of IRF6 in hypertrophic chondrocytes, osteocytes, and bone matrix of craniofacial tissues. Histological analysis of Irf6‐null mice showed a remarkable reduction in the number of lacunae, embedded osteocytes in matrices, and a reduction in mineralization during bone formation. These abnormalities may explain the decreased craniofacial bone density detected by micro‐CT, loss of incisors, and mandibular bone abnormality of Irf6‐null mice. To validate the autonomous role of IRF6 in bone, extracted primary osteoblasts from calvarial bone of WT and Irf6‐null pups showed no effect on osteoblastic viability and proliferation. However, a reduction in mineralization was detected in Irf6‐null cells. Conclusions: Altogether, these findings suggest an autonomous role of Irf6 in regulating bone differentiation and mineralization. Developmental Dynamics 248:221‐232, 2019. © 2019 Wiley Periodicals, Inc.

New Surgical Model for Bone–Muscle Injury Reveals Age and Gender-Related Healing Patterns in the 5 Lipoxygenase (5LO) Knockout Mouse

“…Then, specimens were processed for histological embedding in paraffin blocks. Semi-serial 5-μm histological transversal slices were obtained from the central area of the bone defect and were used for Hematoxylin and Eosin (H&E), modified Goldner's Trichrome/Alcian Blue ( 58 ), and Picrosirius red staining for birefringence and immunohistochemistry for 5LO, Runx2, and TRAP.…”

“…Two histological fields (two biological replicates) of each femur defects stained with Picrosirius Red were captured with a 10× objective under polarizing lens coupled to a binocular inverted microscope (Leica DM IRB/E) and analyzed as previously described (21,61). Green birefringence color indicates thin fibers; yellow and red colors at birefringence analysis indicate thick and organized collagen fibers (21,58). Briefly, spectra of green, yellow, and red colors were defined by RGB values, and the quantity of pixels-squared was calculated for each field by using the AxioVision 4.8 software (CarlZeiss).…”

Interferon regulatory factor 6 is required for proper wound healing in vivo

“…Interferon regulatory factor (IRF) 6 is a transcription factor that belongs to the larger family of IRFs mediating the interferon response following viral infection . IRF6 is expressed in many cell types including cutaneous and oral keratinocytes, innate immune cells, and occasional myocytes and osteocytes . IRF6 belongs to a feedback loop with TP63 that regulates epidermal proliferation and differentiation .…”

Medication-related osteonecrosis of the jaws after tooth extraction in senescent female mice treated with zoledronic acid: Microtomographic, histological and immunohistochemical characterization

“…For control staining of the antibodies, serial sections were treated only with the Universal immuno-enzyme polymer, in a separate preparation. Immunofluorescence was performed following protocols previously described [56, 57]. For detection of cell proliferation in the oral mucosa, sections from the initial portions of sockets were incubated with primary antibody for Proliferating Cell Nuclear Antigen (PCNA) (#MAB424, Merck Millipore, Darmstadt, Germany) diluted to 1:150 in blocking solution and incubated at 4°C overnight.…”