SRY-box 9 (SOX9) is a master transcription factor that regulates cartilage development. SOX9 haploinsufficiency resulting from breakpoints in a ∼1-Mb region upstream of SOX9 was reported in acampomelic campomelic dysplasia (ACD) patients, suggesting that essential enhancer regions of SOX9 for cartilage development are located in this long non-coding sequence. However, the cis-acting enhancer region regulating cartilage-specific SOX9 expression remains to be identified. To identify distant cartilage Sox9 enhancers, we utilized the combination of multiple CRISPR/Cas9 technologies including enrichment of the promoter-enhancer complex followed by next-generation sequencing and mass spectrometry (MS), SIN3A-dCas9-mediated epigenetic silencing, and generation of enhancer deletion mice. As a result, we could identify a critical far-upstream cis-element and Stat3 as a trans-acting factor, regulating cartilage-specific Sox9 expression and subsequent skeletal development. Our strategy could facilitate definitive ACD diagnosis and should be useful to reveal the detailed chromatin conformation and regulation.

Muscle atrophy (loss of skeletal muscle mass) causes progressive deterioration of skeletal function. Recently, excessive intake of fats was suggested to induce insulin resistance, followed by muscle atrophy. Green tea extracts (GTEs), which contain polyphenols such as epigallocatechin gallate, have beneficial effects on obesity, hyperglycemia, and insulin resistance, but their effects against muscle atrophy are still unclear. Here, we found that GTEs prevented high-fat (HF) diet–induced muscle weight loss in senescence-accelerated mouse prone-8 (SAMP8), a murine model of senescence. SAMP8 mice were fed a control diet, an HF diet, or HF with 0.5% GTEs (HFGT) diet for 4 months. The HF diet induced muscle weight loss with aging (measured as quadriceps muscle weight), whereas GTEs prevented this loss. In HF diet–fed mice, blood glucose and plasma insulin concentrations increased in comparison with the control group, and these mice had insulin resistance as determined by homeostasis model assessment of insulin resistance (HOMA-IR). In these mice, serum concentrations of leukocyte cell–derived chemotaxin 2 (LECT2), which is known to induce insulin resistance in skeletal muscle, were elevated, and insulin signaling in muscle, as determined by the phosphorylation levels of Akt and p70 S6 kinases, tended to be decreased. In HFGT diet–fed mice, these signs of insulin resistance and elevation of serum LECT2 were not observed. Although our study did not directly show the effect of serum LECT2 on muscle weight, insulin resistance examined using HOMA-IR indicated an intervention effect of serum LECT2 on muscle weight, as revealed by partial correlation analysis. Accordingly, GTEs might have beneficial effects on age-related and HF diet–induced muscle weight loss, which correlates with insulin resistance and is accompanied by a change in serum LECT2.

We clearly found a change of D-dimer during pregnancy. When D-dimer was higher than 3.2 microg/mL, the percentage of ultrasonographically positive women was high. We propose that women with D-dimer higher than 3.2 microg/mL are closely monitored for prevention of pulmonary thromboembolism.

BackgroundPlate fixation is one of the standard surgical treatments for distal femoral fractures. There are few reports on the relationship between the screw position and bone union when fixing by the bridging plate (relative stability) method.MethodsThis retrospective study included 71 distal femoral fractures of 70 patients who were treated with the locking compression plate for distal femur (DePuy Synthes Co., Ltd, New Brunswick, CA, USA). The following measurements were evaluated and analyzed: (1) bone union rate, (2) bridge span length (distance between screws across the fracture), (3) plate span ratio (plate length/bone fracture length), (4) number of empty holes (number of screw holes not inserted around the fracture), and (5) medial fracture distance (bone fracture distance on the medial side of the distal femur). Patient demographics (age), comorbidities (smoking, diabetes, chronic steroid use, dialysis), and injury characteristics (AO type, open fracture, infection) were obtained for all participants. Univariate analysis was performed on them.ResultsOf 71 fractures, 26 fractures were simple fractures, 45 fractures were comminuted fractures, and 7 fractures resulted in non-union. Non-union rate was significantly higher in comminuted fractures with bone medial fracture distance exceeding 5 mm.Non-union was founded in simple fractures with bone medial fracture distance exceeding 2 mm, but not significant (p = 0.06). In cases with simple fractures, one non-union case had one empty hole and one non-union case had four empty holes, whereas in cases with comminuted fractures, five non-union cases had two more empty holes.ConclusionsWe concluded that bone fragment distance between fracture fragments is more important than bridge span length of the fracture site and the number of empty holes. Smoking and medial fracture distance are prognostic risk factors of nonunion in distal femoral fractures treated with LCP as bridging plate.

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Infrapatellar fat pad has been implicated in knee osteoarthritis. We examined whether infrapatellar fat pad volume is associated with quantitative cartilage changes using magnetic resonance imaging T1ρ mapping. Fifty‐eight knees of knee pain patients (19 men, 39 women, mean age 57.0 [range 29–85] years) who were conservatively managed and had >1 T1ρ mapping images taken over time were evaluated. We used three slices from the medial femoral and tibial cartilage; areas showing T1ρ values <130 ms and >50 ms were designated as having cartilage degeneration. Cases were categorized into three groups: Improvement, No Change, and Deterioration. Fat‐suppressed T2‐weighted sagittal magnetic resonance images were used for measuring infrapatellar fat pad volume. Percent change in infrapatellar fat pad volume was −5.01 ± 5.66%, −2.06 ± 4.92%, and 0.05 ± 6.09% in the Improvement (n = 22), No Change (n = 22), and Deterioration (n = 14) groups, respectively, demonstrating significantly reduced infrapatellar fat pad volume in the Improvement group (p < 0.05). Multivariate regression analyses revealed that the percent change in infrapatellar fat pad volume significantly affected T1ρ change category independent of age, sex, follow‐up period, baseline infrapatellar fat pad volume, and Kellgren‐Lawrence grade. Infrapatellar fat pad volume and obesity or body weight change showed no correlation. Infrapatellar fat pad volume was reduced in patients with improved quantitative cartilage assessment on magnetic resonance imaging T1ρ mapping. This is the first study demonstrating associations between quantitative cartilage changes and infrapatellar fat pad morphological changes, suggesting a detrimental role of infrapatellar fat pad volume in articular cartilage degeneration. © 2018 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res

We report the complete nucleotide sequence of plasmid pMR68, isolated from Pseudomonas strain K-62, two plasmids contribute to broad-spectrum mercury resistance and that the mer operon from one of them (pMR26) has been previously characterized. The plasmid was 71,020 bp in length and contained 75 coding regions. Three mer gene clusters were identified. The first comprised merR-orf4-orf5-merT1-merP1-merF-merA-merB1, which confers bacterial resistance to mercuric ions and organomercury. The second and third clusters comprised merT2-merP2, which encodes a mercury transport system, and merB2, which encodes an organomercurial lyase, respectively. The deduced amino acid sequences for the proteins encoded by each of the mer genes identified in pMR68 bore greater similarity to sequences from Methylobacterium extorquens AM1 than to those from pMR26, a second mercury-resistance plasmid from Pseudomonas strain K-62. Escherichia coli cells carrying pMKY12 (containing merR-orf4-orf5-merT1-merP1-merF-merA-merB1 cloned from pMR68) and cells carrying pMRA114 (containing merR-merT-merP-merA-merG-merB1 cloned from plasmid pMR26) were more resistant to, and volatilized more, mercury from mercuric ions and phenylmercury than the control cells. The present results, together with our earlier findings, indicate that the high phenylmercury resistance noted for Pseudomonas strain K-62 seems to be achieved by multiple genes, particularly by the multiple merB encoding organomercurial lyase and one merG encoding cellular permeability to phenylmercury. The novel mer gene identified in pMR68 may help us to design new strategies aimed at the bioremediation of mercurials.