Differences in outcomes of different scraping therapy schemes for lumbar disc herniation based on metabonomics

doi:10.3969/j.issn.2095-4344.0556

Abstract

Abstract:

BACKGROUND: Scraping therapy has been widely used to treat lumbar disc herniation. The underlying mechanism can be studied through multiple links, pathways and targets using metabonomics.
OBJECTIVE: To analyze the serum metabolic profile changes in lumbar disc herniation rats interfered with scraping therapy and differences in efficacy of different regimens by gas chromatography-mass spectrometry (GC-MS), and to explore the underlying mechanism to provide new ideas and basis for clinical treatment, and promote the normalization and standardization of scraping therapy.
METHODS: Forty-eight Sprague-Dawley rats from Zhejiang Provincial Animal Center were randomly divided into Scraping therapy groups (corresponding channels with ecchymosis, corresponding channels without ecchymosis, acupoints with ecchymosis, acupoints without ecchymosis), model group, and blank control group (n=8 per group). A rat model of noncompressive lumbar disc herniation induced by autologous nucleus pulposus was established in the model and scraping therapy groups. Different scraping therapy programs were given to scraping therapy groups, once every other day, for 3 courses (3 days a course). The mechanical pain threshold before and after modeling was detected by electronic pain threshold detector. The serum endogenous differential metabolites were analyzed by GC-MS.
RESULTS AND CONCLUSION: After modeling, the mechanical pain threshold in each group was significantly lower than before modeling (P < 0.01). According to the metabonomics, 14 potential biomarkers such as glycine, pyruvate acid and ornithine were screened out, and these potential biomarkers were involved in seven metabolic pathways, including glycine, serine and threonine metabolism, pyruvate metabolism and citric cycle. Scraping therapy played an anti-inflammatory effect on lumbar disc herniation rats by regulating the urea content, glycine, serine and threonine metabolism, pyruvate, and achieved analgesic effect through the adjustment of arginine and proline metabolism pathways. After comparing the contents of glycine, pyruvate acid, ornithine and urea in four groups of scraping therapy, the corresponding channels with ecchymosis group showed the best therapeutic effect on the abnormal metabolites. In the future, large sampled clinical trials are needed for further research.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

Key words: Systems Biology, Spinal Diseases, Lumbar Vertebrae, Intervertebral Disk Displacement, Tissue Engineering

CLC Number:

R318

Chen Li-hong1, Yue Rong-zhao1, Zhang Yong-yi1, Zhang Qin1, Wang Yan-jun1, Gui Qian1, Yang Min1, Xu Gui-hua1, Xie Tong2, Peng Lin-xiu2. Differences in outcomes of different scraping therapy schemes for lumbar disc herniation based on metabonomics [J]. Chinese Journal of Tissue Engineering Research, 2018, 22(32): 5110-5116.

Figures/Tables 2

2.1 实验动物数量分析纳入48只大鼠，共分为6组，每组8只。营养状况良好，建造自体髓核移植非压迫性腰椎间盘突出症模型和进行后续实验时，均无死亡，所有大鼠全部进入结果分析。 2.2 行为学观察刮痧组和模型组大鼠出现烦躁、撕咬肢体、反复舔后爪、食量降低等表现[26]；刮痧组大鼠干预后期症状呈好转趋势。整个实验过程中未出现死亡例数，未出现脊髓损伤相关症状，造模创口愈合良好，未出现明显炎症反应及瘢痕组织。 2.3 刮痧组、模型组机械痛阈的比较刮痧组和模型组造模后的机械痛阈值均较各组造模前显著降低(P < 0.01)，详见表1。 2.4 实验质量控制每走10针正式样本进一针QC样本，利用所有样本(包括QC样本)的主成分分析和各物质在QC样本中的变异度来检测仪器的可靠性和重复性[27]。计算所有鉴定出来的代谢物在总QC样本中峰高的RSD值，RSD分布见图1。同时，将QC样本与实验样本进行主成分分析，见图2。从主成分分析图可以看出，QC样本相对集中，表明此次实验过程质控较好。因此，实验操作和仪器条件相对稳定。 2.5 GC-MS结果分析采用GC-MS技术对6组大鼠血清进行分析，得到总离子流图，见图3。 2.6 正交偏最小二乘判别分析正交偏最小二乘判别分析是偏最小二乘判别分析的一种衍生运算分析方法，主要结合了正交矫正信号(OSC)和PLS两种方法，通过将X轴矩阵信息分解成与Y相关和不相关的2类信息，能够排除与分组不相关的一些变量，更准确的筛选出有价值的差异性变量从而使判别能力更优[28]。正交偏最小二乘判别分析模型参数中，R2Y表示模型解释能力，Q2表示模型预测能力，以此考察模型的可靠程度[29]。使用正交偏最小二乘判别分析处理6组血清代谢物谱数据，模型相关参数显示为：R2X=0.616，Q2=0.302，R2Y=0.673，说明模型对样本的解释性较好，但由于样本量较小，该模型的预测性一般。从正交偏最小二乘判别分析得分图(图4)中可以看出空白组和模型组、刮痧4组区分显著，无重叠交叉。模型组和刮痧4组呈现出较为明显的区分。 2.7 潜在生物标志物鉴定根据主成分分析与正交偏最小二乘判别分析，并通过fold change>1.2及单因素方差分析(P < 0.05)筛选，共找到14种潜在的生物标志物，见表2。14种潜在生物标志物在各组血清中含量见图5。 2.8 代谢通路分析使用MetaboAnalyst 3.6平台，将筛选出来的差异代谢物导入并进行代谢通路分析。代谢通路重要值(pathway impact，PI)的临界值设置为0.02，高于这个值将被选择作为潜在的靶标路径。共得到7条相关代谢通路信息(图6)：①甘氨酸、丝氨酸和苏氨酸代谢；②精氨酸和脯氨酸代谢；③柠檬酸循环；④丙酮酸代谢；⑤糖酵解或糖异生；⑥半胱氨酸和蛋氨酸代谢；⑦原代胆汁酸生物合成。其中，甘氨酸、丝氨酸和苏氨酸代谢，丙酮酸代谢，精氨酸和脯氨酸代谢，柠檬酸循环这4条代谢途径在此次实验腰椎间盘突出症大鼠内源性代谢中起到重要作用。"

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