Analysis of mineral apposition rates during alveolar bone regeneration over three weeks following transfer of BMP-2/7 gene via in vivo electroporation

Alveolar bone is not spontaneously regenerated following trauma or periodontitis. We previously proposed an animal model for new alveolar bone regeneration therapy based on the non-viral BMP-2/7 gene expression vector and in vivo electroporation, which induced the formation of new alveolar bone over the course of a week. Here, we analysed alveolar bone during a period of three weeks following gene transfer to periodontal tissue. Non-viral plasmid vector pCAGGS-BMP-2/7 or pCAGGS control was injected into palatal periodontal tissue of the first molar of the rat maxilla and immediately electroporated with 32 pulses of 50 V for 50 msec. Over the following three weeks, rats were double bone-stained by calcein and tetracycline every three days and mineral apposition rates (MAR) were measured. Double bone-staining revealed that MAR of alveolar bone was as similar level three days before BMP-2/7 gene transfer as three days after gene transfer. However, from 3 to 6 days, 6 to 9 days, 9 to 12 days, 12 to 15 days, 15 to 18 days, and 18 to 20 days after, MARs were significantly higher than prior to gene transfer. Our proposed gene therapy for alveolar bone regeneration combining non-viral BMP-2/7 gene expression vector and in vivo electroporation could increase alveolar bone regeneration potential in the targeted area for up to three weeks

The whole blood transcriptional regulation landscape in 465 COVID-19 infected samples from Japan COVID-19 Task Force

「コロナ制圧タスクフォース」COVID-19患者由来の血液細胞における遺伝子発現の網羅的解析 --重症度に応じた遺伝子発現の変化には、ヒトゲノム配列の個人差が影響する--. 京都大学プレスリリース. 2022-08-23.Coronavirus disease 2019 (COVID-19) is a recently-emerged infectious disease that has caused millions of deaths, where comprehensive understanding of disease mechanisms is still unestablished. In particular, studies of gene expression dynamics and regulation landscape in COVID-19 infected individuals are limited. Here, we report on a thorough analysis of whole blood RNA-seq data from 465 genotyped samples from the Japan COVID-19 Task Force, including 359 severe and 106 non-severe COVID-19 cases. We discover 1169 putative causal expression quantitative trait loci (eQTLs) including 34 possible colocalizations with biobank fine-mapping results of hematopoietic traits in a Japanese population, 1549 putative causal splice QTLs (sQTLs; e.g. two independent sQTLs at TOR1AIP1), as well as biologically interpretable trans-eQTL examples (e.g., REST and STING1), all fine-mapped at single variant resolution. We perform differential gene expression analysis to elucidate 198 genes with increased expression in severe COVID-19 cases and enriched for innate immune-related functions. Finally, we evaluate the limited but non-zero effect of COVID-19 phenotype on eQTL discovery, and highlight the presence of COVID-19 severity-interaction eQTLs (ieQTLs; e.g., CLEC4C and MYBL2). Our study provides a comprehensive catalog of whole blood regulatory variants in Japanese, as well as a reference for transcriptional landscapes in response to COVID-19 infection

DOCK2 is involved in the host genetics and biology of severe COVID-19

「コロナ制圧タスクフォース」COVID-19疾患感受性遺伝子DOCK2の重症化機序を解明 --アジア最大のバイオレポジトリーでCOVID-19の治療標的を発見--. 京都大学プレスリリース. 2022-08-10.Identifying the host genetic factors underlying severe COVID-19 is an emerging challenge. Here we conducted a genome-wide association study (GWAS) involving 2, 393 cases of COVID-19 in a cohort of Japanese individuals collected during the initial waves of the pandemic, with 3, 289 unaffected controls. We identified a variant on chromosome 5 at 5q35 (rs60200309-A), close to the dedicator of cytokinesis 2 gene (DOCK2), which was associated with severe COVID-19 in patients less than 65 years of age. This risk allele was prevalent in East Asian individuals but rare in Europeans, highlighting the value of genome-wide association studies in non-European populations. RNA-sequencing analysis of 473 bulk peripheral blood samples identified decreased expression of DOCK2 associated with the risk allele in these younger patients. DOCK2 expression was suppressed in patients with severe cases of COVID-19. Single-cell RNA-sequencing analysis (n = 61 individuals) identified cell-type-specific downregulation of DOCK2 and a COVID-19-specific decreasing effect of the risk allele on DOCK2 expression in non-classical monocytes. Immunohistochemistry of lung specimens from patients with severe COVID-19 pneumonia showed suppressed DOCK2 expression. Moreover, inhibition of DOCK2 function with CPYPP increased the severity of pneumonia in a Syrian hamster model of SARS-CoV-2 infection, characterized by weight loss, lung oedema, enhanced viral loads, impaired macrophage recruitment and dysregulated type I interferon responses. We conclude that DOCK2 has an important role in the host immune response to SARS-CoV-2 infection and the development of severe COVID-19, and could be further explored as a potential biomarker and/or therapeutic target

FK506 ニ ヨル メンエキ ヨクセイカ デノ BMP-2 ハツゲン アデノウイルス ベクター ニ ヨル ラット ノ コツ ユウドウ ニ カンスル ケンキュウ

京都大学0048新制・論文博士博士(医学)乙第11157号論医博第1805号新制||医||833(附属図書館)UT51-2003-H819(主査)教授 中村 孝志, 教授 開 祐司, 教授 飯塚 忠彦学位規則第4条第2項該当Doctor of Medical ScienceKyoto UniversityDA

Complement Factor H Is an Early Predictive Biomarker of the Therapeutic Efficacy of Sublingual Immunotherapy for Japanese Cedar Pollinosis

Sublingual immunotherapy for Japanese cedar pollinosis can improve the symptoms of allergic rhinitis and modify its natural course. However, sublingual immunotherapy requires a long treatment period and some patients do not respond to treatment. In this study, we aimed to identify biomarkers that could predict the efficacy of sublingual immunotherapy at an early stage. In this study, 40 patients from phase III trials were recruited and divided into good and poor response groups. Using peripheral blood mononuclear cells from before and two months after the start of medication, microarray, discriminant analysis, and real-time polymerase chain reaction were performed to extract candidate genes that could be biomarkers. Furthermore, these genes were validated in 30 patients in general clinical practice. Complement factor H was upregulated in the good response group and downregulated in the poor response group. Complement factor H may be a useful biomarker for predicting the efficacy of sublingual immunotherapy for Japanese cedar pollinosis at early time points after treatment initiation

The impact of postoperative inclination of the joint line on clinical outcomes in total knee arthroplasty using a prosthesis with anatomical geometry

Abstract The goal of this study was to investigate the impact of postoperative inclination of the joint line on clinical results after total knee arthroplasty (TKA) using a prosthesis with anatomical geometry. This study included 145 primary cruciate-retaining type of knee prosthesis with anatomical geometry. Three years postoperatively, clinical outcomes including the patient-reported outcomes (PROs) were recorded. Limb alignment was evaluated by the hip-knee-ankle (HKA) axis and inclination of the joint line was assessed by the joint line orientation angle (JLOA). Knees were divided into two groups according to the HKA: in-range (− 3 to 3°) and outlier group ( 3°) or the JLOA: in-range (2–4°) and outlier group ( 4°), and clinical outcomes were compared between the groups. Postoperative Knee Society Function Score (KS-FS) was significantly higher in the HKA in-range group than the outlier group (p = 0.01). The Knee Society Knee Score and all subscales of the Knee injury Osteoarthritis Outcome Score were comparable between the groups. A multivariate analysis revealed a significant association between age at operation and postoperative KS-FS > of 80 points. Neither HKA in-range nor JLOA in-range were associated with the higher knee function. In conclusion, TKA-postoperative inclination of the joint line was not relevant to the short-term PROs. Treatment strategies that attempt to make joint line inclination in order to improve postoperative PROs should be avoided, and alignment goals such as kinematic alignment should be considered carefully

Determination of cell fate in skeletal muscle following BMP gene transfer by in vivo electroporation

We previously developed a novel method for gene transfer, which combined a non-viral gene expression vector with transcutaneous in vivo electroporation. We applied this method to transfer the bone morphogenetic protein (BMP) gene and induce ectopic bone formation in rat skeletal muscles. At present, it remains unclear which types of cells can differentiate into osteogenic cells after BMP gene transfer by in vivo electroporation. Two types of stem cells in skeletal muscle can differentiate into osteogenic cells: muscle-derived stem cells, and bone marrow-derived stem cells in the blood. In the present study, we transferred the BMP gene into rat skeletal muscles. We then stained tissues for several muscle-derived stem cell markers (e.g., Pax7, M-cadherin), muscle regeneration-related markers (e.g., Myod1, myogenin), and an inflammatory cell marker (CD68) to follow cell differentiation over time. Our results indicate that, in the absence of BMP, the cell population undergoes muscle regeneration, whereas in its presence, it can differentiate into osteogenic cells. Commitment towards either muscle regeneration or induction of ectopic bone formation appears to occur five to seven days after BMP gene transfer.</p

Unilateral laminectomy for bilateral decompression improves low back pain while standing equally on both sides in patients with lumbar canal stenosis: analysis using a detailed visual analogue scale

Abstract Background Unilateral laminectomy for bilateral decompression (ULBD) for lumbar spinal stenosis (LSS) is a less invasive technique compared to conventional laminectomy. Recently, several authors have reported favorable results of low back pain (LBP) in patients of LSS treated with ULBD. However, the detailed changes and localization of LBP before and after ULBD for LSS remain unclear. Furthermore, unsymmetrical invasion to para-spinal muscle and facet joint may result in the residual unsymmetrical symptoms. To clarify these points, we conducted an observational study and used detailed visual analog scale (VAS) scores to evaluate the characteristics and bilateral changes of LBP and lower extremity symptoms. Methods We included 50 patients with LSS treated with ULBD. A detailed visual analogue scale (VAS; 100 mm) score of LBP in three different postural positions: motion, standing, and sitting, and bilateral VAS score (approached side versus opposite side) of LBP, lower extremity pain (LEP), and lower extremity numbness (LEN) were measured. Oswestry Disability Index (ODI) was used to quantify the clinical improvement. Results Detailed LBP VAS score before surgery was 51.5 ± 32.5 in motion, 63.0 ± 30.1 while standing, and 37.8 ± 31.8 while sitting; and showed LBP while standing was significantly greater than LBP while sitting (p < 0.01). After surgery, LBP while standing was significantly improved relative to that while sitting (p < 0.05), and levels of LBP in the three postures became almost the same with ODI improvement. Bilateral VAS scores showed significant improvement equally on both sides (p < 0.01). Conclusions ULBD improves LBP while standing equally on both sides in patients with LCS. The improvement of LBP by the ULBD surgery suggests radicular LBP improved because of decompression surgery. Furthermore, the symmetric improvement of LBP by the ULBD surgery suggests unsymmetrical invasion of the paraspinal muscles and facet joints is unrelated to residual LBP