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蛋白质组学分析人类胎盘:为骨关节炎的潜在治疗应用提供新视角。

Proteomic Characterization of Human Placenta: Insights into Potential Therapeutic Applications for Osteoarthritis.

机构信息

Department of Pharmaceutics, Faculty of Pharmacy, MS Ramaiah University of Applied Sciences, University House, Gnanagangothri Campus, New BEL Road, MSR Nagar, Bengaluru, 560054, India.

Cancyte Technologies Pvt Ltd, Sri Shankara Research Centre, Rangadore Memorial Hospital, Shankarapuram, Bengaluru, India.

出版信息

AAPS PharmSciTech. 2024 Jun 18;25(6):139. doi: 10.1208/s12249-024-02851-5.

Abstract

Biologics have become increasingly prominent as therapeutics in recent years due to their innate immune-privileged nature, biocompatibility, and high levels of protein biofactors. The aim of the study is to characterise the biologic, lyophilized human placenta (LHP) and explore its therapeutic potential for osteoarthritis (OA). The presence of six bioactive constituents that regulate cell-extracellular matrix interaction was identified by liquid chromatography coupled to electrospray ionization and quadrupole time-of-flight mass spectrometry (LC-ESI-QTOF/MS). Metalloproteinase inhibitor 3 (TIMP3), alpha-1 anti-trypsin (a1AT), basic fibroblast growth factor (bFGF), and transforming growth factor β1 (TGFβ1) were detected and quantified using ELISA. The total protein content present in LHP by Bradford assay was found to be 409.35 ± 0.005 μg/ml. The analytical techniques such as Attenuated Total Reflectance-Fourier Transform Infrared spectroscopy (ATR-FTIR), solid state carbon-13 Nuclear Magnetic Resonance (ssC NMR) spectroscopy, and Differential Scanning Calorimetry (DSC) revealed the secondary structure and conformational stability of LHP. X-Ray diffraction (XRD) studies showed its amorphous nature. Bioactivity assessment of LHP was performed in human keratinocytes (HaCaT) and human dermal fibroblasts (HDF) by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The LHP was highly proliferative against skin cells and non-toxic, based on the findings of the bioactivity assay. LHP has the potential to be used as a therapeutic agent for OA, as its characterisation unveiled its physical stability, significant concentration of bioactive components that are pertinent to cartilage repair and its conformational stability.

摘要

近年来,生物制剂因其固有免疫特权特性、生物相容性和高水平的蛋白质生物因子而在治疗中变得越来越突出。本研究旨在对生物冻干人胎盘(LHP)进行特征描述,并探索其在骨关节炎(OA)治疗中的潜力。通过液相色谱-电喷雾电离-四极杆飞行时间质谱联用(LC-ESI-QTOF/MS)鉴定了六种调节细胞-细胞外基质相互作用的生物活性成分。采用酶联免疫吸附试验(ELISA)检测和定量金属蛋白酶抑制剂 3(TIMP3)、α-1 抗胰蛋白酶(a1AT)、碱性成纤维细胞生长因子(bFGF)和转化生长因子β1(TGFβ1)。通过 Bradford 测定法发现 LHP 中的总蛋白含量为 409.35±0.005μg/ml。衰减全反射-傅里叶变换红外光谱(ATR-FTIR)、固态碳-13 核磁共振(ssC NMR)光谱和差示扫描量热法(DSC)等分析技术揭示了 LHP 的二级结构和构象稳定性。X 射线衍射(XRD)研究表明其为无定形物质。通过 3-(4,5-二甲基噻唑-2-基)-2,5-二苯基四氮唑溴盐(MTT)测定法在人角质形成细胞(HaCaT)和人真皮成纤维细胞(HDF)中对 LHP 的生物活性进行评估。根据生物活性测定结果,LHP 对皮肤细胞具有高度增殖作用且无毒性。LHP 具有作为 OA 治疗剂的潜力,因为其特性揭示了其物理稳定性、对软骨修复有重要意义的生物活性成分的高浓度及其构象稳定性。

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