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Summary: Point mutations in KIF22 have been linked to spondyloepimetaphyseal dysplasia with joint laxity, type 2 (SEMDJL2). Skeletal features of SEMDJL2 include short stature and joint laxity. Mechanisms underlying these limb abnormalities are unknown. Here in this manuscript, we have investigated the function of KIF22 in chondrocytes. Quantitative PCR and immunostaining revealed that Kif22 was highly expressed in proliferating-zone growth-plate chondrocytes. Kif22 knockdown resulted in defective mitotic spindle formation and reduced cell proliferation. Forced expression of SEMDJL-associated mutant Kif22 constructs likewise induced abnormal mitotic spindle morphology and reduced proliferation. Mice expressing a KIF22 truncation mutant had shorter growth plates and shorter tibial bones compared to wild-type mice. These results suggest that KIF22 regulates mitotic spindle formation in proliferating chondrocytes thereby linking the stunted longitudinal bone growth observed in SEMDJL2 to failures of chondrocyte division.

Abstract Salivary glands develop through epithelial-mesenchymal interactions and are formed through repeated branching. The Crk-associated substrate protein (p130Cas) serves as an adapter that forms a complex with various proteins via integrin and growth factor signaling, with important regulatory roles in several essential cellular processes. We found that p130Cas is expressed in ductal epithelial cells of the submandibular gland (SMG). We generated epithelial tissue-specific p130Cas-deficient (p130Cas Δepi– ) mice and aimed to investigate the physiological role of p130Cas in the postnatal development of salivary glands. Histological analysis showed immature development of granular convoluted tubules (GCT) of the SMG in male p130Cas Δepi– mice. Immunofluorescence staining showed that nuclear-localized androgen receptors (AR) were specifically decreased in GCT cells in p130Cas Δepi– mice. Furthermore, epidermal growth factor-positive secretory granules contained in GCT cells were significantly reduced in p130Cas Δepi– mice with downregulated AR signaling. GCTs lacking p130Cas showed reduced numbers and size of secretory granules, disrupted subcellular localization of the cis-Golgi matrix protein GM130, and sparse endoplasmic reticulum membranes in GCT cells. These results suggest that p130Cas plays a crucial role in androgen-dependent GCT development accompanied with ER-Golgi network formation in SMG by regulating the AR signaling.

Abstract Texture has enormous effects on food preferences. The materials used to study texture discrimination also have tastes that experimental animal can detect; therefore, such studies must be designed to exclude taste differences. In this study, to minimize the effects of material tastes, we utilized high- and low-viscosity forms of carboxymethyl cellulose (CMC-H and CMC-L, respectively) at the same concentrations (0.1–3%) for viscosity discrimination tests in rats. In two-bottle preference tests of water and CMC, rats avoided CMC-H solutions above 1% (63 mPa·s) but did not avoid less viscous CMC-L solutions with equivalent taste magnitudes, suggesting that rats spontaneously avoided high viscosity. To evaluate low-viscosity discrimination, we performed conditioned aversion tests to 0.1% CMC, which initially showed a comparable preference ratio to water in the two-bottle preference tests. Conditioning with 0.1% CMC-L (1.5 mPa·s) did not induce aversion to 0.1% CMC-L or CMC-H. However, rats acquired a conditioned aversion to 0.1% CMC-H (3.6 mPa·s) even after latent inhibition to CMC taste by pre-exposure to 0.1% CMC-L. These results suggest that rats can discriminate considerably low viscosity independent of CMC taste. This novel approach for viscosity discrimination can be used to investigate the mechanisms of texture perception in mammals.

Severe intraoral pain induces difficulty in eating and speaking, leading to a decline in the quality of life. However, the molecular mechanisms underlying intraoral pain remain unclear. Here, we investigated gene modulation in the trigeminal ganglion and intraoral pain-related behavior in a rat model of acetic acid-induced oral ulcerative mucositis. Oral ulceration was observed on day 2 after acetic acid treatment to the oral mucosa of male Wistar rats, causing spontaneous pain and mechanical allodynia. Deoxyribonucleic acid microarray analysis of trigeminal ganglion tissue indicated that Hamp (a hepcidin gene that regulates cellular iron transport) was the most upregulated gene. In the oral ulcerative mucositis model, the upregulation of Hamp was also induced in the ulcer region but not in the liver, with no increase in hepcidin levels in the plasma and saliva, indicating that hepcidin was produced locally in the ulcer region in the model. Systemic antibiotic pretreatment did not increase the mRNA levels of Hamp in the trigeminal ganglion and ulcer regions. Hepcidin injection into the oral mucosa enhanced neuronal excitability in response to noxious mechanical stimulation of the oral mucosa in trigeminal spinal subnucleus interpolaris/caudalis neurons. These results imply that oral ulcerative mucositis induces oral mucosal pain because of infectious inflammation of the ulcerative area and potentiates Hamp, which represents anti-bacterial and anti-peptidase gene expression in the ulcer region and trigeminal ganglion. The regulation of cellular iron transport by hepcidin is likely involved in oral ulcerative mucositis-induced pain.

Despite the long history of use of steroid ointments for oral mucositis, the analgesic mechanism has not been fully elucidated. In this study, we examined the effects of triamcinolone acetonide (Tmc) on oral ulcerative mucositis-induced pain in conscious rats by our proprietary assay system. Based on evaluations of the physical properties and retention periods in the oral mucosa of human volunteers and rats, we selected TRAFUL® ointment as a long-lasting base. In oral ulcerative mucositis model rats, TRAFUL® with Tmc suppressed cyclooxygenase-dependent inflammatory responses with upregulations of glucocorticoid receptor-induced anti-inflammatory genes and inhibited spontaneous nociceptive behavior. When an ointment with a shorter residual period was used, the effects of Tmc were not elicited or were induced to a lesser extent. Importantly, TRAFUL® with Tmc also improved oral ulcerative mucositis-induced mechanical allodynia, which has been reported to be independent of cyclooxygenase. Ca2+ imaging in dissociated trigeminal ganglion neurons showed that long-term preincubation with Tmc inhibited the hypertonic stimulation-induced Ca2+ response. These results suggest that the representative steroid Tmc suppresses oral ulcerative mucositis-induced pain by general anti-inflammatory actions and inhibits mechanical sensitivity in peripheral nerves. For drug delivery, long-lasting ointments such as TRAFUL® are needed to sufficiently induce the therapeutic effects.

Osteoclasts resorb bone by attaching on the bone matrix and forming a sealing zone. In Src-deficient mice, osteoclasts cannot form the actin ring, a characteristic actin structure that seals the resorbed area, and resorb hardly any bone as a result. However, the molecular mechanism underlying the role of Src in the regulation and organization of the actin ring is still unclear. We identified an actin-regulatory protein, protein phosphatase 1 regulatory subunit 18 (PPP1r18), as an Src-binding protein in an Src-, Yes-, and Fyn-deficient fibroblast (SYF) cell line overexpressing a constitutively active form of Src. PPP1r18 was localized in the nucleus and actin ring. PPP1r18 overexpression in osteoclasts inhibited terminal differentiation, actin ring formation, and bone-resorbing activity. A mutation of the protein phosphatase 1 (PP1)-binding domain of PPP1r18 rescued these phenotypes. In contrast, PPP1r18 knockdown promoted terminal differentiation and actin ring formation. In summary, we showed that PPP1r18 likely plays a role in podosome organization and bone resorption. [ABSTRACT FROM AUTHOR]

Satellite cells are skeletal muscle stem cells that provide myonuclei for postnatal muscle growth, maintenance, and repair/regeneration in adults. Normally, satellite cells are mitotically quiescent, but they are activated in response to muscle injury, in which case they proliferate extensively and exhibit up-regulated expression of the transcription factor MyoD, a master regulator of myogenesis. MyoD forms a heterodimer with E proteins through their basic helix-loop-helix domain, binds to E boxes in the genome and thereby activates transcription at muscle-specific promoters. The central role of MyoD in muscle differentiation has increased interest in finding potential MyoD regulators. Here we identified transducin-like enhancer of split (TLE3), one of the Groucho/TLE family members, as a regulator of MyoD function during myogenesis. TLE3 was expressed in activated and proliferative satellite cells in which increased TLE3 levels suppressed myogenic differentiation, and, conversely, reduced TLE3 levels promoted myogenesis with a concomitant increase in proliferation. We found that, via its glutamine- and serine/proline-rich domains, TLE3 interferes with MyoD function by disrupting the association between the basic helix-loop-helix domain of MyoD and E proteins. Our findings indicate that TLE3 participates in skeletal muscle homeostasis by dampening satellite cell differentiation via repression of MyoD transcriptional activity. [ABSTRACT FROM AUTHOR]