- 作者列表："Ambrosi TH","Goodnough LH","Steininger HM","Hoover MY","Kim E","Koepke LS","Marecic O","Zhao L","Seita J","Bishop JA","Gardner MJ","Chan CKF
:Fragility fractures have a limited capacity to regenerate, and impaired fracture healing is a leading cause of morbidity in the elderly. The recent identification of a highly purified bona fide human skeletal stem cell (hSSC) and its committed downstream progenitor cell populations provides an opportunity for understanding the mechanism of age-related compromised fracture healing from the stem cell perspective. In this study, we tested whether hSSCs isolated from geriatric fractures demonstrate intrinsic functional defects that drive impaired healing. Using flow cytometry, we analyzed and isolated hSSCs from callus tissue of five different skeletal sites (n = 61) of patients ranging from 13 to 94 years of age for functional and molecular studies. We observed that fracture-activated amplification of hSSC populations was comparable at all ages. However, functional analysis of isolated stem cells revealed that advanced age significantly correlated with reduced osteochondrogenic potential but was not associated with decreased in vitro clonogenicity. hSSCs derived from women displayed an exacerbated functional decline with age relative to those of aged men. Transcriptomic comparisons revealed downregulation of skeletogenic pathways such as WNT and upregulation of senescence-related pathways in young versus older hSSCs. Strikingly, loss of Sirtuin1 expression played a major role in hSSC dysfunction but re-activation by trans-resveratrol or a small molecule compound restored in vitro differentiation potential. These are the first findings that characterize age-related defects in purified hSSCs from geriatric fractures. Our results provide a foundation for future investigations into the mechanism and reversibility of skeletal stem cell aging in humans.
: 脆性骨折的再生能力有限，骨折愈合障碍是老年人发病的主要原因。一种高度纯化的真正的人类骨骼干细胞 (hSSC) 的最新鉴定其承诺的下游祖细胞群为从干细胞角度理解年龄相关的骨折愈合障碍的机制提供了机会。在本研究中，我们检测了从老年骨折中分离的 hSSCs 是否表现出驱动愈合受损的内在功能缺陷。使用流式细胞术，我们分析并分离了 13 ~ 94 岁患者 5 个不同骨骼部位 (n = 61) 愈伤组织的 hSSCs，进行功能和分子研究。我们观察到 hSSC 人群的骨折激活扩增在所有年龄都具有可比性。然而，分离干细胞的功能分析发现，高龄与骨软骨病潜能降低显著相关，但与体外克隆形成能力降低无关。相对于老年男性，来源于女性的 hSSCs 显示出随年龄增长而加剧的功能下降。转录组学比较揭示了年轻与老年 hSSCs 中 WNT 等骨骼生成途径的下调和衰老相关途径的上调。引人注目的是，Sirtuin1 表达缺失在 hSSC 功能障碍中起主要作用，但反式白藜芦醇或小分子化合物的再激活恢复了体外分化潜能。这些是首次发现老年骨折纯化 hSSCs 中年龄相关缺陷的特征。我们的结果为将来研究人类骨骼干细胞衰老的机制和可逆性提供了基础。
METHODS::A major challenge in translating findings from genome-wide association studies (GWAS) to biological mechanisms is pinpointing functional variants because only a very small percentage of variants associated with a given trait actually impact the trait. We used an extensive epigenetics, transcriptomics, and genetics analysis of the TBX15/WARS2 neighbourhood to prioritize this region's best-candidate causal variants for the genetic risk of osteoporosis (estimated bone density, eBMD) and obesity (waist-hip ratio or waist circumference adjusted for body mass index). TBX15 encodes a transcription factor that is important in bone development and adipose biology. Manual curation of 692 GWAS-derived variants gave eight strong candidates for causal SNPs that modulate TBX15 transcription in subcutaneous adipose tissue (SAT) or osteoblasts, which highly and specifically express this gene. None of these SNPs were prioritized by Bayesian fine-mapping. The eight regulatory causal SNPs were in enhancer or promoter chromatin seen preferentially in SAT or osteoblasts at TBX15 intron-1 or upstream. They overlap strongly predicted, allele-specific transcription factor binding sites. Our analysis suggests that these SNPs act independently of two missense SNPs in TBX15. Remarkably, five of the regulatory SNPs were associated with eBMD and obesity and had the same trait-increasing allele for both. We found that WARS2 obesity-related SNPs can be ascribed to high linkage disequilibrium with TBX15 intron-1 SNPs. Our findings from GWAS index, proxy, and imputed SNPs suggest that a few SNPs, including three in a 0.7-kb cluster, act as causal regulatory variants to fine-tune TBX15 expression and, thereby, affect both obesity and osteoporosis risk.
METHODS:PURPOSE:Systemic mastocytosis (SM) is characterized by the expansion of clonal mast cells that infiltrate various organ systems. The extent of organ infiltration and subsequent organ damage distinguishes between indolent SM (ISM) defined by a nearly normal life expectancy and advanced SM (AdvSM) defined by poor prognosis. In ISM, measurement of the bone mineral density (BMD) frequently reveals osteoporosis. In contrast, the clinical implication of an increased BMD and osteosclerosis remains unclear. METHODS:BMD was evaluated in 61 patients with mastocytosis (ISM, n = 29, 48%; AdvSM, n = 32, 52%). We correlated the prevalence of osteoporosis, increased BMD and osteosclerosis with clinical parameters, disease variant and prognosis. RESULTS:Osteoporosis was detected in 11/29 (38%) patients with ISM but only in 2/32 (6%) patients with AdvSM (p = 0.004). An increased BMD was detected in 1/29 (3%) patients with ISM and 24/32 (75%) patients with AdvSM (p < 0.001) while osteosclerosis was only detected in AdvSM patients (16/32, 50%). AdvSM patients with increased BMD had higher levels of bone marrow mast cell infiltration, higher serum tryptase and alkaline phosphatase levels compared to ISM as well as higher number of high-molecular risk mutations (p < 0.05). In addition, we found that the prognosis of AdvSM patients with increased BMD is inferior compared to those without increased BMD (median overall survival 3.6 years versus not reached, p = 0.031). CONCLUSIONS:Osteoporosis is a common feature in ISM but not in AdvSM. An increased BMD is frequently present in AdvSM but not in ISM and is associated with more advanced disease and inferior outcome.
METHODS::We present salmon calcitonin (SCT) loaded Hydroxyapatite nanoparticles (HAP-NPs) for sublingual osteoporosis therapy. Surface stabilized HAP-NPs were prepared by aqueous precipitation. SCT was loaded by ionic complexation, as confirmed by FTIR. SCT-HAP-NPs exhibited high loading efficiency (~85%), average size (~100nm), and zeta potential (~-25 mv). Stability of SCT was established by circular dichroism spectroscopy and HPLC analysis. Confocal laser scanning microscopy confirmed deep penetration of SCT-HAP-NPs into the mucosa with >4-fold enhancement in permeability relative to SCT solution. Sublingual SCT-HAP-NPs revealed relative bioavailability of ~15% compared to the subcutaneous injection in rabbits (200iu). Significant and comparable improvement in serum biomarkers with increase in bone mass and mechanical strength and decreased bone erosion compared to subcutaneous SCT was confirmed in ovariectomized (OVX) osteoporosis rat model. Such comparable pharmacodynamic effect at the same dose suggested targeted bone delivery and promise of sublingual SCT-HAP-NPs as a non-invasive alternative to the injection.