Overexpression of Cbfa1 in osteoblasts inhibits osteoblast maturation and causes osteopenia with multiple fractures

Targeted disruption of core binding factor α1 (Cbfa1) showed that Cbfa1 is an essential transcription factor in osteoblast differentiation and bone formation. Furthermore, both in vitro and in vivo studies showed that Cbfa1 plays important roles in matrix production and mineralization. However, it remains to be clarified how Cbfa1 controls osteoblast differentiation, bone formation, and bone remodelling. To understand fully the physiological functions of Cbfa1, we generated transgenic mice that overexpressed Cbfa1 in osteoblasts using type I collagen promoter. Unexpectedly, Cbfa1 transgenic mice showed osteopenia with multiple fractures. Cortical bone, which was thin, porous, and enriched with osteopontin, was invaded by osteoclasts, despite the absence of acceleration of osteoclastogenesis. Although the number of neonatal osteoblasts was increased, their function was impaired in matrix production and mineralization. Furthermore, terminally differentiated osteoblasts, which strongly express osteocalcin, and osteocytes were diminished greatly, whereas less mature osteoblasts expressing osteopontin accumulated in adult bone. These data indicate that immature organization of cortical bone, which was caused by the maturational blockage of osteoblasts, led to osteopenia and fragility in transgenic mice, demonstrating that Cbfa1 inhibits osteoblast differentiation at a late stage

Antiresorptive agent-related osteonecrosis of the jaw: Position Paper 2017 of the Japanese Allied Committee on Osteonecrosis of the Jaw

Antiresorptive agent-related osteonecrosis of the jaw (ARONJ) is an intractable, though rare, complication in cancer patients with bone metastases and patients with osteoporosis who are treated with antiresorptive agents, including bisphosphonates and denosumab. Despite the more than 10 years that have passed since the first cases of bisphosphonate-related osteonecrosis of the jaw (BRONJ) were reported, our understanding of the epidemiology and pathophysiology of ARONJ remains limited, and data supported by evidence-based medicine are still sparse. However, the diagnosis and staging of ARONJ, identification of risk factors, and development of preventive and therapeutic approaches have advanced significantly over the past decade. The Position Paper 2017 is an updated version of the Position Paper 2010 of the Japanese Allied Committee on Osteonecrosis of the Jaw, which now comprises six Japanese academic societies. The Position Paper 2017 describes a new diagnostic definition for ARONJ, as proposed by the American Association of Oral and Maxillofacial Surgeons (AAOMS), summarizes our current understanding of the pathophysiology of ARONJ based on a literature search, and suggests methods for physicians and dentists/oral surgeons to manage the disease. In addition, the appropriateness of discontinuing antiresorptive medications (drug holiday) before, during, and after invasive dental treatments is discussed extensively. More importantly, the manuscript also proposes, for the first time, the importance of interactive communication and cooperation between physicians and dentists/oral surgeons for the successful treatment of ARONJ. The Position Paper 2017 is intended to serve as a guide for improving the management of ARONJ patients in Japan

Comprehensive phenotypic and genomic characterization of venous malformations

Hirose K., Hori Y., Ozeki M., et al. Comprehensive phenotypic and genomic characterization of venous malformations. Human Pathology 145, 48 (2024); https://doi.org/10.1016/j.humpath.2024.02.004.Venous malformations (VMs) are the most common vascular malformations. TEK and PIK3CA are the causal genes of VMs, and may be involved in the PI3K/AKT pathway. However, the downstream mechanisms underlying the TEK or PIK3CA mutations in VMs are not completely understood. This study aimed to identify a possible association between genetic mutations and clinicopathological features. A retrospective clinical, pathological, and genetic study of 114 patients with VMs was performed. TEK, PIK3CA, and combined TEK/PIK3CA mutations were identified in 49 (43%), 13 (11.4%), and 2 (1.75%) patients, respectively. TEK-mutant VMs more commonly occurred in younger patients than TEK and PIK3CA mutation-negative VMs (other-mutant VMs), and showed more frequent skin involvement and no lymphocytic aggregates. No significant differences were observed in sex, location of occurrence, malformed vessel size, vessel density, or thickness of the vascular smooth muscle among the VM genotypes. Immunohistochemical analysis revealed that the expression levels of phosphorylated AKT (p-AKT) were higher in the TEK-mutant VMs than those in PIK3CA-mutant and other-mutant VMs. The expression levels of p-mTOR and its downstream effectors were higher in all the VM genotypes than those in normal vessels. Spatial transcriptomics revealed that the genes involved in “blood vessel development”, “positive regulation of cell migration”, and “extracellular matrix organization” were up-regulated in a TEK-mutant VM. Significant genotype-phenotype correlations in clinical and pathological features were observed among the VM genotypes, indicating gene-specific effects. Detailed analysis of gene-specific effects in VMs may offer insights into the underlying molecular pathways and implications for targeted therapies

Clear Cell Squamous Cell Carcinoma of the Maxillary Gingiva Associated with PIK3CA and HRAS Mutations: Report of a Case and Literature Review

The version of record of this article, first published in Head and Neck Pathology, is available online at Publisher’s website: https://doi.org/10.1007/s12105-023-01580-8.Background: Squamous cell carcinoma (SCC) is the most common oral malignancy, and somatic mutations in some driver genes have been implicated in SCC development. Clear cell SCC (CCSCC) is a rare histological variant of SCC, and various clear cell neoplasms must be considered in the differential diagnosis of CCSCC in the oral cavity. Based on a limited number of CCSCC cases reported in the oral cavity, CCSCC is considered an aggressive variant of SCC with a poor prognosis; however, its genetic characteristics remain unknown. Methods: A maxillary gingival tumor in an 89-year-old female was described and investigated using immunohistochemical staining, special staining, fluorescence in situ hybridization, and next-generation sequencing (NGS) with a custom panel of driver genes, including those associated with SCC and clear cell neoplasm development. Results: Histopathological examination revealed a proliferation of atypical epithelial cells with abundant clear cytoplasm and enlarged and centrally placed round nuclei. The tumor was exophytic with deep, penetrating proliferation. The atypical clear cells were continuous with the conventional SCC cells. Immunohistochemical analysis showed that the clear cells were positive for CK AE1/AE3 and CK5/6 and nuclear-positive for p63. In contrast, the clear cells were negative for αSMA, S100, HMB45, Melan-A, CD10, and p16. p53 immunoreactivity exhibited a wild-type expression pattern. Additionally, the clear cells were positive for periodic acid-Schiff (PAS) and negative for diastase-PAS, mucicarmine, and Alcian blue. Based on these results, the diagnosis of CCSCC was confirmed. Molecular analysis of the clear cells identified PIK3CA p.E542K (c.1624G>A) and HRAS p.G12A (c.35 G>C) somatic mutations classified as oncogenic. No pathogenic variants were identified in TP53, EWSR1, AKT1, PTEN, BRAF, KRAS, NRAS, RASA1, or MAML2. Conclusions: We report a case of CCSCC of the oral cavity with PIK3CA and HRAS mutations. The identification of PIK3CA and/or HRAS mutations is rare in SCC; however, both mutations are important potential targets for antitumor therapy. A detailed analysis of gene mutations in CCSCC may lead to a better understanding of its biological behavior and an improved prognosis, as well as a differential diagnosis from other clear cell neoplasms

Squamous cell carcinoma initially occurring on the tongue dorsum: a case series report with molecular analysis

Ono Sawako, Hirose Katsutoshi, Sukegawa Shintaro, et al. Squamous cell carcinoma initially occurring on the tongue dorsum: a case series report with molecular analysis. Diagnostic Pathology 19, 63 (2024); https://doi.org/10.1186/s13000-024-01487-0.Background: Squamous cell carcinoma (SCC) of the dorsum of the tongue is extremely rare, and it clinically resembles various benign lesions. Somatic mutations in TP53 and some driver genes were implicated in the development of SCC; however, the somatic genetic characteristics of dorsal tongue SCC remain unknown. With a detailed analysis of gene mutations in dorsal tongue SCC, we aimed to better understand its biology. Methods: Four cases of SCC initially occurring on the tongue dorsum were evaluated for clinical and histological findings and immunohistochemical expression of p53 and p16. Gene mutations were analyzed using next-generation sequencing with a custom panel of driver genes.Results: We retrospectively investigated 557 cases of tongue SCC, and only four cases of SCC initially occurred on the tongue dorsum. The four patients (cases 1–4) were one woman and three men with a mean age of 53.75 years (range: 15–74 years). Histological analysis revealed well-differentiated SCC. Through molecular analysis, we identified pathogenic somatic mutations, namely, TP53 p.C176F (c.527G > T) in case 3 and TP53 p.R282W (c.844 C > T) in case 4. No pathogenic variants were identified in the PI3K/AKT or RAS/RAF pathways. The p53 immunohistochemical examination revealed a wild-type expression pattern in cases 1–3 and strong expression in case 4. The results of p16 immunostaining were negative in all cases. Conclusions: We described four previously unreported genetic characteristics of dorsal tongue SCC. Somatic TP53 mutations may contribute to the development of a subset of dorsal tongue SCC; however, more cases with genetic analysis need to be accumulated

SP7 Inhibits Osteoblast Differentiation at a Late Stage in Mice

RUNX2 and SP7 are essential transcription factors for osteoblast differentiation at an early stage. Although RUNX2 inhibits osteoblast differentiation at a late stage, the function of SP7 at the late stage of osteoblast differentiation is not fully elucidated. Thus, we pursued the function of SP7 in osteoblast differentiation. RUNX2 induced Sp7 expression in Runx2−/− calvarial cells. Adenoviral transfer of sh-Sp7 into primary osteoblasts reduced the expression of Alpl, Col1a1, and Bglap2 and mineralization, whereas that of Sp7 reduced Bglap2 expression and mineralization at a late stage of osteoblast differentiation. Sp7 transgenic mice under the control of 2.3 kb Col1a1 promoter showed osteopenia and woven-bone like structure in the cortical bone, which was thin and less mineralized, in a dose-dependent manner. Further, the number of processes in the osteoblasts and osteocytes was reduced. Although the osteoblast density was increased, the bone formation was reduced. The frequency of BrdU incorporation was increased in the osteoblastic cells, while the expression of Col1a1, Spp1, Ibsp, and Bglap2 was reduced. Further, the osteopenia in Sp7 or Runx2 transgenic mice was worsened in Sp7/Runx2 double transgenic mice and the expression of Col1a1 and Bglap2 was reduced. The expression of Sp7 and Runx2 was not increased in Runx2 and Sp7 transgenic mice, respectively. The expression of endogenous Sp7 was increased in Sp7 transgenic mice and Sp7-transduced cells; the introduction of Sp7 activated and sh-Sp7 inhibited Sp7 promoter; and ChIP assay showed the binding of endogenous SP7 in the proximal region of Sp7 promoter. These findings suggest that SP7 and RUNX2 inhibit osteoblast differentiation at a late stage in a manner independent of RUNX2 and SP7, respectively, and SP7 positively regulates its own promoter

Identification and characterization of ameloblastin gene in a reptile

Ameloblastin (AMBN) is one of the enamel sheath proteins which presumably has a role in determining the prismatic structure of growing enamel crystals. There may therefore be a relationship between the molecular evolution of the AMBN gene and the development of enamel prismatic structures. To investigate whether such a relationship exists, it was necessary to identify the homologues of the AMBN gene in a reptile whose teeth lack an enamel prismatic structure. To this end, several clones containing AMBN cDNA were isolated from caiman jaws using the reverse transcription-polymerase chain reaction (RT- PCR) method. Sequence analysis of the AMBN cDNA revealed an open reading frame of 1221 bp encoding a 407-amino-acid protein. Translation of the caiman cDNA starts at the methionine corresponding to the second of two putative start codons conserved in mammalian AMBN genes. The N-terminal part of the caiman AMBN shows high amino acid sequence similarities to human, pig, cattle. rat and mouse AMBN sequences, as well as several other features that have been conserved throughout the evolution of reptiles and mammals. Unexpectedly. the nucleotide sequences of the 3' untranslated region (UTR) are also conserved, not only within mammalian genes but also between reptilian and mammalian genes, The caiman AMBN gene is a single-copy gene, transcribed only in the jaws, presumably in teeth. (C) 2002 Elsevier Science BV. All rights reserved