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摘要:
目的 探讨糖尿病大鼠创面组织能量代谢变化。 方法 该研究为实验研究。将6只8周龄的雄性SD大鼠按照随机数字表法分为造成糖尿病全层皮肤缺损创面的糖尿病组和仅造成全层皮肤缺损创面的对照组,每组3只。计算2组大鼠术后14 d创面愈合率。取2组大鼠术后14 d创面组织,借助色谱-质谱分析进行靶向能量代谢组学检测,筛选2组大鼠创面组织间比较,表达量有显著变化的差异能量代谢物并对差异能量代谢物进行京都基因与基因组百科全书富集分析。 结果 术后14 d,糖尿病组大鼠创面愈合率为（68.3±2.8）%,显著低于对照组的（98.1±1.2）%（t=16.92,P<0.05）。术后14 d,与对照组相比,糖尿病组大鼠创面组织中顺乌头酸、烟酰胺腺嘌呤二核苷酸磷酸、5'-鸟苷二磷酸表达量均显著升高（t值分别为4.74、3.09、3.99,P<0.05）,L-苹果酸、乳酸表达量均显著降低（t值分别为3.45、12.20,P<0.05）；2组大鼠创面组织中另外21种能量代谢物表达量比较,差异均无统计学意义（P>0.05）。5种差异能量代谢物主要在胰高血糖素、Rap1、Ras、缺氧诱导因子1信号通路及三羧酸循环、丙酮酸代谢、线粒体自噬、胞吞作用途径中显著富集。 结论 糖尿病大鼠创面组织中乳酸及L-苹果酸表达量降低,顺乌头酸、烟酰胺腺嘌呤二核苷酸磷酸、5'-鸟苷二磷酸表达量升高；差异能量代谢物主要在氧化应激、能量调控、炎症反应等相关信号通路或途径中富集。 Abstract:Objective To explore the change of energy metabolism in wound tissue of diabetic rats. Methods This study was an experimental study. Six 8-week-old male Sprague-Dawley rats were divided into diabetic group inflicted with a diabetic full-thickness skin defect wound and control group only inflicted with a full-thickness skin defect wound according to the random number table method, with 3 rats in each group. The wound healing rates of rats in the two groups were calculated at 14 d after operation. The wound tissue of two groups of rats was collected at 14 d after operation, the targeted energy metabolomics detection was carried out by chromatography-mass spectrometry analysis, and the differential energy metabolites in the wound tissue were screened with significant change in the expression between the two groups of rats. The Kyoto encyclopedia of genes and genomes enrichment analysis was performed on the differential energy metabolites. Results At 14 d after operation, the wound healing rate of rats in diabetic group was (68.3±2.8)%, which was significantly lower than (98.1±1.2)% in control group (t=16.92, P<0.05). At 14 d after operation, compared with those in control group, the expressions of cis-aconitic acid, nicotinamide adenine dinucleotide phosphate, and 5'-guanosine diphosphate were significantly increased in wound tissue of rats in diabetic group (with t values of 4.74, 3.09, and 3.99, respectively, P<0.05), the expressions of L-malic acid and lactic acid were significantly decreased (with t values of 3.45 and 12.20, respectively, P<0.05), and there were no statistically significant differences in the expression levels of the other 21 energy metabolites in wound tissue of rats in the two groups (P>0.05). The five differential energy metabolites were enriched mainly in the glucagon, Rap1, Ras, hypoxia-inducible factor 1 signaling pathways as well as the tricarboxylic acid cycle, pyruvate metabolism, mitochondrial autophagy,and endocytosis pathways. Conclusions The expressions of lactic acid and L-malic acid in wound tissue of diabetic rats decreased, while the expressions of cis-aconitic acid, nicotinamide adenine dinucleotide phosphate, and 5'-guanosine diphosphate increased. The differential energy metabolites were mainly enriched in oxidative stress, energy regulation, inflammatory reaction, and other related signaling pathways or pathways. -
Key words:
- Wounds and injuries /
- Diabetes mellitus /
- Energy metabolism /
- Citric acid cycle /
- Lactic acid /
- L-malic acid
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王晓阳：酝酿和设计实验、进行实验研究和撰写论文；扈煜婕：整理数据、分析数据；王晓川：统计分析；赵洁、张基勋、姜笃银：研究指导、经费支持所有作者声明不存在利益冲突
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参考文献
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图 1 2组大鼠术后各时间点全层皮肤缺损创面愈合情况及组织病理学情况。1A、1B.分别为对照组术后0（即刻）、14 d创面情况,图1B中创面基本愈合；1C、1D.分别为糖尿病组术后0、14 d创面情况,图1D中仍有较大面积创面未愈合；1E、1F.分别为对照组、糖尿病组术后14 d创面上皮化情况,图1E中创面基本被新生上皮覆盖,图1F中创面尚未完全被新生上皮覆盖 苏木精-伊红×20；1G、1H.分别为对照组、糖尿病组术后14 d创面组织中胶原纤维形成和排列情况,图1G中胶原纤维丰富、排列有序,图1H中胶原纤维稀疏、排列杂乱 Masson×20
注：糖尿病组大鼠诱导糖尿病后通过外科手术制备创面,对照组大鼠仅同前制备创面；红色箭头表示新生上皮,胶原纤维阳性染色为蓝色
图 2 2组大鼠术后14 d全层皮肤缺损创面组织中5种差异能量代谢物的京都基因与基因组百科全书富集分析结果
注：糖尿病组大鼠诱导糖尿病后通过外科手术制备创面,对照组大鼠仅同前制备创面；HIF-1为缺氧诱导因子1
Table 1. 2组大鼠末次注射链脲佐菌素后各时间点一般体征比较（
组别与指标 鼠数（只） 0 d（即刻） 7 d 9 d 11 d 13 d 对照组 3 体重（g） 242.4±2.8 257.6±2.9 260.7±5.0 268.7±2.4 273.7±4.1 饮水量（mL） 17±3 20±3 19±3 22±4 22±6 尿量（g） 17.8±1.7 18.0±3.9 17.5±2.8 22.3±3.1 18.8±2.9 血糖（mmol/L） 5.68±0.32 5.55±0.17 5.82±0.47 5.86±0.10 5.58±0.23 糖尿病组 3 体重（g） 240.3±3.1 213.6±4.3 210.7±5.7 208.9±3.1 204.4±2.3 饮水量（mL） 18±3 55±8 60±3 54±6 57±6 尿量（g） 16.8±1.3 57.8±5.8 54.1±8.4 57.6±9.0 57.2±5.2 血糖（mmol/L） 5.48±0.26 19.27±1.48 20.35±0.95 20.75±0.51 20.03±0.29 t1值 0.85 14.65 11.47 26.05 25.87 P1值 0.441 <0.001 <0.001 <0.001 <0.001 t2值 0.26 6.87 16.72 7.70 7.01 P2值 0.806 0.002 <0.001 0.002 0.002 t3值 0.83 9.83 7.14 6.43 11.10 P3值 0.456 <0.001 0.002 0.003 <0.001 t4值 0.86 15.93 23.81 49.69 66.47 P4值 0.438 <0.001 <0.001 <0.001 <0.001 注：糖尿病组大鼠注射链脲佐菌素构建糖尿病模型,对照组大鼠注射柠檬酸钠缓冲液；体重、饮水量、尿量、血糖时间因素主效应,F值分别为5.03、19.55、18.00、148.80,P值分别为0.059、<0.001、<0.001、<0.001；处理因素主效应,F值分别为367.40、589.00、493.60、4 582.00,P值均<0.001；两者交互作用,F值分别为143.00、14.20、15.51、147.20,P值均<0.001；t1值、P1值,t2值、P2值,t3值、P3值,t4值、P4值分别为2组各时间点体重、饮水量、尿量、血糖比较所得 
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Table 2. 2组大鼠术后14 d全层皮肤缺损创面组织中检测出的能量代谢物的表达量比较（μmol/g,
组别 样本数 乳酸 顺乌头酸 5'-鸟苷二磷酸 L-苹果酸 NADP 硫胺素焦磷酸 5'-腺苷三磷酸 5'-腺苷二磷酸 对照组 3 19 383±876 20.1±2.1 2.115±0.027 225±15 9.9±2.6 3.73±0.39 15.3±1.9 5.12±0.22 糖尿病组 3 10 261±953 26.4±0.8 2.659±0.235 194±5 14.5±0.5 4.32±0.28 18.1±1.3 5.75±0.50 t值 12.20 4.74 3.99 3.45 3.09 2.11 2.10 1.98 P值 <0.001 0.009 0.016 0.026 0.037 0.103 0.103 0.119 注：糖尿病组大鼠诱导糖尿病后通过外科手术制备创面,对照组大鼠仅同前制备创面；NADP为烟酰胺腺嘌呤二核苷酸磷酸,NAD为烟酰胺腺嘌呤二核苷酸,PEP为磷酸烯醇式丙酮酸
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