Dong Wei-Chong, Guo Jia-Liang, Jiang Xin-Hui, Xu Lei, Wang Huan, Ni Xiao-Yu, Zhang Ying-Ze, Zhang Zhi-Qing, Jiang Ye
Department of Pharmacy, The Second Hospital of Hebei Medical University 215# Heping West Road Shijiazhuang Hebei Province 050051 China
Department of Pharmaceutical Analysis, School of Pharmacy, Hebei Medical University 361# East Zhongshan Road Shijiazhuang Hebei Province 050017 China
RSC Adv. 2023 Jan 5;13(2):1267-1277. doi: 10.1039/d2ra05572d. eCollection 2023 Jan 3.
: oxidative stress is linked to various human diseases which developed into the idea of "disrupted redox signaling". Osteoporosis (OP) is a chronic skeletal disorder characterized by low bone mineral density and deterioration of bone microarchitecture among which estrogen deficiency is the main cause. Lack of estrogen leads to the imbalance between oxidation and anti-oxidation in patients, and oxidative stress is an important link in the pathogenesis of OP. The ratio of the reduced to the oxidized thiols can characterize the redox status. However, few methods have been reported for the simultaneous determination of reduced forms and their oxidized forms of thiols in plasma. : we developed a hollow fiber centrifugal ultrafiltration (HFCF-UF) method for sample preparation and validated a high-performance liquid chromatography tandem mass spectrometry (HPLC-MS/MS) method to determine two reduced forms of thiols-homocysteine (Hcy), cysteine (Cys) levels and their respective oxidized compounds, homocystine (HHcy) and cystine (Cyss) in rat plasma simultaneously for the first time. Thirty-six female rats were randomly divided into three groups: normal control (NC), oxidative stress (ovariectomy, OVX) and ovariectomy with hydrogen-rich saline administration (OVX + HRS). : the validation parameters for the methodological results were within the acceptance criteria. There were both significant differences of Hcy/HHcy (Hcy reduced/oxidized) and Cys/Cyss (Cys reduced/oxidized) in rat plasma between three groups with both < 0.05 and meanwhile, the values of malondialdehyde, superoxide dismutase and glutathione peroxidase were all less than 0.01. The value of both Hcy/HHcy and Cys/Cyss were significantly decreased with the change of Micro-CT scan result of femoral neck in OVX group (both the trabecular thickness and trabecular number significantly decreased with a significant increase of trabecular separation) which demonstrate OP occurs. The change of Hcy/HHcy is more obvious and prominent than Cys/Cyss. : the Hcy/HHcy and Cys/Cyss could be suitable biomarkers for oxidative stress and especially Hcy/HHcy is more sensitive. The developed method is simple and accurate. It can be easily applied in clinical research to further evaluate the oxidative stress indicator for disease risk factors.
氧化应激与多种人类疾病相关，由此发展出了“氧化还原信号紊乱”的概念。骨质疏松症（OP）是一种慢性骨骼疾病，其特征是骨矿物质密度低和骨微结构恶化，其中雌激素缺乏是主要原因。雌激素缺乏导致患者体内氧化与抗氧化失衡，氧化应激是OP发病机制中的一个重要环节。还原型硫醇与氧化型硫醇的比例可以表征氧化还原状态。然而，同时测定血浆中硫醇的还原形式及其氧化形式的方法鲜有报道。我们开发了一种中空纤维离心超滤（HFCF-UF）方法用于样品制备，并首次验证了一种高效液相色谱串联质谱（HPLC-MS/MS）方法，可同时测定大鼠血浆中两种还原型硫醇——同型半胱氨酸（Hcy）、半胱氨酸（Cys）的水平及其各自的氧化产物，即同型胱氨酸（HHcy）和胱氨酸（Cyss）。36只雌性大鼠随机分为三组：正常对照组（NC）、氧化应激组（卵巢切除，OVX）和卵巢切除并给予富氢盐水组（OVX + HRS）。方法学结果的验证参数在可接受标准范围内。三组大鼠血浆中Hcy/HHcy（Hcy还原型/氧化型）和Cys/Cyss（Cys还原型/氧化型）均存在显著差异（均P < 0.05），同时，丙二醛、超氧化物歧化酶和谷胱甘肽过氧化物酶的值均小于0.01。OVX组中，随着股骨颈Micro-CT扫描结果的变化（骨小梁厚度和骨小梁数量显著降低，骨小梁间距显著增加），Hcy/HHcy和Cys/Cyss的值均显著降低，这表明发生了OP。Hcy/HHcy的变化比Cys/Cyss更明显和突出。Hcy/HHcy和Cys/Cyss可能是氧化应激的合适生物标志物，尤其是Hcy/HHcy更敏感。所开发的方法简单准确。它可以很容易地应用于临床研究，以进一步评估疾病危险因素的氧化应激指标。
