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Different Roles for Colony Stimulating Factor 1 Isoforms in Anabolic Therapy for Low Bone Mass

集落刺激因子 1 同工型在低骨量合成代谢治疗中的不同作用

基本信息

批准号：
10585240
负责人：
KARL Leonard INSOGNA
金额：
$ 65.93万
依托单位：
YALE UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2023
资助国家：
美国
起止时间：
2023-07-10 至 2028-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10585240
关键词：
Affinity Agonist Anabolism Animal Model Animals Attenuated B-Lymphocytes Binding Bone Resorption C57BL/6 Mouse CSF1 gene CSF1R gene Cells Coculture Techniques Colony-Stimulating Factors Data Dose Enzymes Estrogen deficiency Event Exposure to Immunophenotyping Impairment In Vitro Kinetics Macrophage Colony-Stimulating Factor Membrane Modeling Molecular Mus Osteoblasts Osteoclasts Osteogenesis Osteoporosis Paracrine Communication Patients Phospholipids Population Study Postmenopausal Osteoporosis Production Protein Isoforms Regimen Role Serum Signal Transduction T-Lymphocyte Testing Therapeutic Time Treatment Efficacy Treatment Protocols Wild Type Mouse bone bone loss bone mass density experimental study in vivo in vivo Model inhibitor insight mouse model new therapeutic target novel osteoclastogenesis paracrine pharmacologic pre-clinical receptor response skeletal skeletal disorder sphingosine 1-phosphate sphingosine kinase transcriptome transcriptome sequencing transcriptomic profiling

项目摘要

CSF1 is the principal colony stimulating activity released by osteoblasts in response to PTH treatment. Its receptor, c-fms, is more highly expressed on mature osteoclasts than any other cell in bone. We found that deleting c-fms in osteoclasts attenuates the anabolic response to PTH. This indicates that part of PTH's anabolic actions could be via a paracrine loop in which PTH stimulates expression of CSF1 in osteoblasts, which then acts on osteoclasts to induce anabolic clastokines that augment bone formation. There are two major isoforms of CSF1: soluble (sCSF1) and membrane-associated (mCSF1). We found that the anabolic response to PTH is augmented in animals only expressing the sCSF1 isoform due to a greater increase in osteoblast number in these mice compared to PTH-treated controls. In striking contrast, animals only expressing mCSF1 had no increase in bone mass in response to an anabolic PTH regimen. Importantly, sCSF1 and mCSF1 differ in the kinetics and extent to which they activate the CSF1 receptor, c-fms, suggesting that their divergent in vivo actions may be due in part, to intrinsic differences in cell-signaling. Based on these data, we hypothesize that mCSF1 inhibits PTH anabolism by opposing the actions of sCSF1 on osteoclasts. When unopposed, sCSF1 contributes to PTH anabolism by inducing production of anabolic clastokines. Specifically, single daily doses of PTH induce transient increases in sCSF1 in osteoblasts that cause bursts of anabolic clastokine production. Consistent with this, we found that sCSF1 stimulates production of the anabolic clastokine, sphingosine-1-phosphate (S-1-P) in osteoclasts. To test these hypotheses, we will, in SA1, determine if adding intermittent dosing of sCSF1 to an anabolic PTH treatment regimen augments the skeletal response to PTH in wild type animals, while adding mCSF1 to that regimen attenuates the response. We will then try to restore the anabolic response to PTH in the sCSF1-/- mice by treating with PTH plus sCSF1. These experiments will provide pharmacologic evidence that the very different response to PTH in sCSF1-/- and mCSF1-/- mice is directly due to differing actions of the two CSF1 isoforms in bone. In SA 2, we will treat wild type mice that were ovariectomized 5 months earlier, with PTH plus sCSF1 to determine if it restores bone mass to pre-OVX levels as a model of therapy for established post-menopausal osteoporosis. In SA 3, we will determine if sCSF1 and mCSF1 induce production of different clastokine profiles in mature osteoclasts. We will use osteoblast/osteoclast cocultures as a model of in vivo paracrine signaling in bone and profile the transcriptome of osteoclasts exposed to PTH-treated osteoblasts expressing only sCSF1 or only mCSF1 to identify differences in the types of clastokines produced. We will also determine differences in the transcriptomes of osteoblasts in the cocultures stimulated by these different clastokine profiles. Finally, we will examine binding kinetics of sCSF1 and mCSF1 to c-fms, and differences in downstream signaling, to gain molecular insight into the divergent in vivo and in vitro actions of these two isoforms.

CSF 1是成骨细胞响应PTH处理释放的主要集落刺激活性。其受体c-fms在成熟破骨细胞上的表达高于骨中的任何其它细胞。我们发现在破骨细胞中删除c-fms减弱了对PTH的合成代谢反应。这表明部分PTH的合成代谢作用可能是通过旁分泌回路，其中PTH刺激成骨细胞中CSF 1的表达，作用于破骨细胞以诱导增加骨形成的合成代谢断裂因子。有两种主要的同种型 CSF 1：可溶性（sCSF 1）和膜相关（mCSF 1）。我们发现对甲状旁腺素的合成代谢反应在只表达sCSF 1亚型的动物中，由于成骨细胞的大量增加，与PTH处理的对照相比，这些小鼠中的数量。与此形成鲜明对比的是， mCSF 1对合成代谢PTH方案的骨量没有增加。重要的是，sCSF 1和 mCSF 1在激活CSF 1受体c-fms的动力学和程度上不同，这表明它们的不同的体内作用可能部分是由于细胞信号传导的内在差异。根据这些数据，我们假设mCSF 1通过对抗sCSF 1对破骨细胞的作用来抑制PTH拮抗剂。当 sCSF 1通过诱导合成代谢断裂因子的产生而有助于PTH的抑制。具体而言，每日单剂量PTH诱导成骨细胞中sCSF 1的瞬时增加，合成代谢断裂因子的爆发与此相一致，我们发现sCSF 1刺激产生破骨细胞中的合成代谢断裂因子，鞘氨醇-1-磷酸（S-1-P）。为了验证这些假设，我们将在 SA 1，确定在合成代谢PTH治疗方案中添加sCSF 1间歇给药是否会增加在野生型动物中对PTH骨骼反应，而向该方案中加入mCSF 1减弱了反应然后，我们将尝试通过用PTH治疗来恢复sCSF 1-/-小鼠对PTH的合成代谢反应。加上sCSF 1。这些实验将提供药理学证据，表明在不同的人中， sCSF 1-/-和mCSF 1-/-小鼠的差异直接归因于两种CSF 1同种型在骨中的不同作用。在SA 2中，我们将用PTH加sCSF 1治疗5个月前切除卵巢的野生型小鼠，以确定它可以将骨量恢复到OVX前的水平，作为治疗绝经后骨质疏松症的模型。骨质疏松在SA 3中，我们将确定sCSF 1和mCSF 1是否诱导产生不同的断裂因子，在成熟破骨细胞中的分布。我们将使用成骨细胞/破骨细胞共培养作为体内旁分泌的模型骨中的信号传导，并分析暴露于PTH处理的成骨细胞的破骨细胞的转录组，仅sCSF 1或仅mCSF 1以鉴定产生的断裂因子类型的差异。我们还将确定这些不同的断裂因子谱刺激的共培养物中成骨细胞转录组的差异。最后，我们将研究sCSF 1和mCSF 1与c-fms的结合动力学，以及下游信号传导的差异，以获得对这两种异构体在体内和体外的不同作用的分子洞察。