Jun Dimerization Protein 2 (JDP2), a Member of the AP-1 Family of Transcription Factor, Mediates Osteoclast Differentiation Induced by RANKL

Osteoclasts are multinucleated cells that resorb bones, and are derived from hematopoietic cells of the monocyte/macrophage lineage. The receptor activator of NF-κB ligand (RANKL, also called ODF/TRANCE/OPGL) stimulates both osteoclast differentiation from osteoclast progenitors and activation of mature osteoclasts. To identify genes responsible for osteoclast differentiation, we used a molecular indexing technique. Here, we report a clone of one of these genes whose transcription is induced by soluble RANKL (sRANKL) in both the RAW264.7 cells of the mouse macrophage cell line and the mouse primary bone marrow cells. The predicted protein was found to be a mouse homologue of Jun dimerization protein 2 (JDP2), a member of the AP-1 family of transcription factors, containing a basic region-leucine zipper motif. Transient transfection experiments revealed that overexpression of JDP2 leads to activation of both tartrate-resistant acid phosphatase (TRAP) and cathepsin K gene promoters in RAW264.7 cells. Infection of mouse primary bone marrow cells with retroviruses expressing JDP2-facilitated sRANKL-mediated formation of TRAP-positive multinuclear osteoclasts. Importantly, antisense oligonucleotide to JDP2 strongly suppressed sRANKL-induced osteoclast formation of RAW264.7 cells. Our findings suggest that JDP2 may play an important role in the RANK-mediated signal transduction system, especially in osteoclast differentiation

Pain following COVID-19 vaccination

Pain at the injection site is the most frequent reaction among COVID-19 vaccine recipients, but its characteristics were not fully described yet. The purpose of this study was to investigate multiple domains of pain following BNT162b2 mRNA vaccination. We included 107 subjects undergoing primary shot of the vaccination twice into deltoid muscle with a 3-week interval. They completed 6 sessions of pain assessments, one before the first and second dose (1-0, 2-0), and 1st / 7th day after the first and second dose (1-1 / 1-7, 2-1 / 2-7). Pain visual analog scale (VAS), pain distribution, and pressure pain threshold (PPT) on deltoid muscle were evaluated in each session. The mean VAS (at rest / shoulder motion) was 6.0 / 27.6 mm at 1-1, and 12.8 / 34.0 mm at 2-1. Approximately, 90% of recipients showed localized pain within the upper arm. Percentage change of PPTs at 1-1 and 2-1 was bilaterally (ipsilateral / contralateral) decreased to 87.4 / 89.4% and 80.6 / 91.0%, which was recovered to the baseline level at 1-7 and 2-7. Temporary, mild-to-moderate intensity, localized distribution, concomitant with bilateral mechanical hyperalgesia on the deltoid muscle, were typical pain characteristics following this vaccination. These findings provide a rationale that will be informative for future recipients

Effects of drug discontinuation after short-term daily alendronate administration on osteoblasts and osteocytes in mice

In order to determine whether osteoclastic bone resorption is restarted after withdrawn of bisphosphonates, we conducted histological examinations on murine osteoclasts, osteoblasts and osteocytes after discontinuation of a daily regimen of alendronate (ALN) with a dosage of 1 mg/kg/day for 10 days. After drug discontinuation, metaphyseal trabecular number and bone volume remained unaltered for the first 4 days. Osteoclast number did not increase, while the number of apoptotic osteoclasts was elevated. On the other hand, tissue non-specific alkaline phosphatase-immunoreactive area was markedly reduced after ALN discontinuation. In addition, osteocytes showed an atrophic profile with empty lacunar areas during and after ALN treatment. Interestingly, as early as 36 h after a single ALN injection, osteocytes show signs of atrophy despite the presence of active osteoblasts. Structured illumination microscopy system showed shortening of osteocytic cytoplasmic processes after drug cessation, suggesting a possible morphological and functional disconnection between osteocytes and osteoblasts. Taken together, it appears that osteoclastic bone resorption is not resumed after ALN discontinuation; also, osteoblasts and osteocytes hardly seem to recover once they are inactivated and atrophied by ALN. In summary, it seems that one must pay more attention to the responses of osteoblasts and osteocytes, rather focusing on the resuming of osteoclastic bone resorption after the ALN discontinuation

Patched1 Haploinsufficiency Increases Adult Bone Mass and Modulates Gli3 Repressor Activity

SummaryHedgehog (Hh)-Patched1 (Ptch1) signaling plays essential roles in various developmental processes, but little is known about its role in postnatal homeostasis. Here, we demonstrate regulation of postnatal bone homeostasis by Hh-Ptch1 signaling. Ptch1-deficient (Ptch1+/−) mice and patients with nevoid basal cell carcinoma syndrome showed high bone mass in adults. In culture, Ptch1+/− cells showed accelerated osteoblast differentiation, enhanced responsiveness to the runt-related transcription factor 2 (Runx2), and reduced generation of the repressor form of Gli3 (Gli3rep). Gli3rep inhibited DNA binding by Runx2 in vitro, suggesting a mechanism that could contribute to the bone phenotypes seen in the Ptch1 heterozygotes. Moreover, systemic administration of the Hh signaling inhibitor cyclopamine decreased bone mass in adult mice. These data provide evidence that Hh-Ptch1 signaling plays a crucial role in postnatal bone homeostasis and point to Hh-Ptch1 signaling as a potential molecular target for the treatment of osteoporosis

A complex rearrangement between APC and TP63 associated with familial adenomatous polyposis identified by multimodal genomic analysis: a case report

Genetic testing of the APC gene by sequencing analysis and MLPA is available across commercial laboratories for the definitive genetic diagnosis of familial adenomatous polyposis (FAP). However, some genetic alterations are difficult to detect using conventional analyses. Here, we report a case of a complex genomic APC-TP63 rearrangement, which was identified in a patient with FAP by a series of genomic analyses, including multigene panel testing, chromosomal analyses, and long-read sequencing. A woman in her thirties was diagnosed with FAP due to multiple polyps in her colon and underwent total colectomy. Subsequent examination revealed fundic gland polyposis. No family history suggesting FAP was noted except for a first-degree relative with desmoid fibromatosis. The conventional APC gene testing was performed by her former doctor, but no pathogenic variant was detected, except for 2 variants of unknown significance. The patient was referred to our hospital for further genetic analysis. After obtaining informed consent in genetic counseling, we conducted a multigene panel analysis. As insertion of a part of the TP63 sequence was detected within exon16 of APC, further analyses, including chromosomal analysis and long-read sequencing, were performed and a complex translocation between chromosomes 3 and 5 containing several breakpoints in TP63 and APC was identified. No phenotype associated with TP63 pathogenic variants, such as split-hand/foot malformation (SHFM) or ectrodactyly, ectodermal dysplasia, or cleft lip/palate syndrome (EEC) was identified in the patient or her relatives. Multimodal genomic analyses should be considered in cases where no pathogenic germline variants are detected by conventional genetic testing despite an evident medical or family history of hereditary cancer syndromes

Deterioration of high-resolution computed tomography findings predicts disease progression after initial decline in forced vital capacity in idiopathic pulmonary fibrosis patients treated with pirfenidone

Background Pirfenidone suppresses the decline of forced vital capacity (FVC) in patients with idiopathic pulmonary fibrosis (IPF). However, IPF progresses in some patients despite treatment. We analyzed patients with meaningful FVC declines during pirfenidone treatment and explored the factors predictive of disease progression after FVC decline. Methods This study was a retrospective, multicenter, observational study conducted by the Okayama Respiratory Disease Study Group. We defined initial decline in %FVC as 5% or greater per 6-month period during pirfenidone treatment. IPF patients who were treated with pirfenidone and experienced an initial decline from December 2008 to September 2017 were enrolled. Results We analyzed 21 patients with IPF. After the initial decline, 4 (19.0%) patients showed improvement in disease, 11 (52.4%) showed stable disease, and 6 (28.6%) showed progressive disease. There was no significant correlation between %FVC reduction on initial decline and subsequent %FVC change (p = 0.475). Deterioration of high-resolution computed tomography (HRCT) findings on initial decline was observed significantly more often in the progressive versus improved/stable disease groups (100% vs 20.0%, p = 0.009). Conclusions We revealed that deterioration of HRCT findings may predict disease progression after the initial decline in %FVC in IPF patients treated with pirfenidone

Detection of epidermal growth factor receptor mutations in exhaled breath condensate using droplet digital polymerase chain reaction

The detection of certain oncogenic driver mutations, including those of epidermal growth factor receptor (EGFR), is essential for determining treatment strategies for advanced non‑small cell lung cancer (NSCLC). The current study assessed the feasibility of testing exhaled breath condensate (EBC) for EGFR mutations by droplet digital PCR (ddPCR). Samples were collected from 12 patients with NSCLC harboring EGFR mutations that were admitted to Okayama University Hospital between June 1, 2014 and December 31, 2017. A total of 21 EBC samples were collected using the RTube™ method and EGFR mutations (L858R, exon 19 deletions or T790M) were assessed through ddPCR analysis (EBC‑ddPCR). A total of 3 healthy volunteer samples were also tested to determine a threshold value for each mutation. Various patient characteristics were determined, including sex (3 males and 9 females), age (range 54‑81 years; median, 66 years), smoking history (10 had never smoked; 2 were former smokers), histology (12 patients exhibited adenocarcinoma), clinical stage (9 patients were stage IV; 3 exhibited post‑operative recurrence) and EGFR mutation type (4 had L858R; 8 had exon 19 deletions; 8 had T790M). EBC‑ddPCR demonstrated positive droplets in 8 of the 12 patients. The sensitivity and specificity of each mutation was as follows: 27.3 and 80.0% for EGFR L858R, 30.0 and 90.9% for EGFR Ex19del, and 22.2 and 100% for EGFR T790M. EBC‑ddPCR analysis of EGFR mutations exhibited modest sensitivity and acceptable specificity. EBC‑ddPCR is a minimally invasive and replicable procedure and may be a complementary method for EGFR testing in patients where blood or tissue sampling proves difficult