[1]朱然,梁舒,钱晶晶,等.高糖和糖浓度波动对人角膜上皮细胞中MMP-9表达及真菌黏附的影响[J].眼科新进展,2021,41(6):534-539.[doi:10.13389/j.cnki.rao.2021.0111]
　ZHU Ran,LIANG Shu,QIAN Jingjing,et al.Effects of high glucose and fluctuating glucose on the expression of MMP-9 in human corneal epithelium cells and the adhesion of Fungi[J].Recent Advances in Ophthalmology,2021,41(6):534-539.[doi:10.13389/j.cnki.rao.2021.0111]
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高糖和糖浓度波动对人角膜上皮细胞中MMP-9表达及真菌黏附的影响/HTML
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《眼科新进展》[ISSN:1003-5141/CN:41-1105/R]

卷:
41卷
期数:
2021年6期
页码:
534-539
栏目:
实验研究
出版日期:
2021-06-05

文章信息/Info

Title:
Effects of high glucose and fluctuating glucose on the expression of MMP-9 in human corneal epithelium cells and the adhesion of Fungi
作者:
朱然; 梁舒; 钱晶晶; 李豪亮; 张希熹
226001　江苏省南通市，南通大学附属医院眼科研究所
Author(s):
ZHU Ran; LIANG Shu; QIAN Jingjing; LI Haoliang; ZHANG Xixi
Department of Ophthalmology,Affiliated Hospital of Nantong University,Nantong 226001,Jiangsu Province,China
关键词:
茄病镰刀菌; 尖端赛多孢子菌; 人角膜上皮细胞; 高糖; 糖浓度波动
Keywords:
Fusarium solani;  Scedosporium apiospermum;  human corneal epithelium cells;  high glucose;  fluctuating glucose
分类号:
R772.2
DOI:
10.13389/j.cnki.rao.2021.0111
文献标志码:
A
摘要:
目的　探讨不同浓度葡萄糖环境下茄病镰刀菌和尖端赛多孢子菌对人角膜上皮细胞（HCEC）的黏附能力和HCEC 中MMP-9表达的影响。方法　将处于对数生长期的HCEC分为3组：低糖组（5.5 mmol·L-1葡萄糖培养基培养）、高糖组（25.0 mmol·L-1葡萄糖培养基培养）和糖波动组（5.5 mmol·L-1和50.0 mmol·L-1葡萄糖培养基交替培养），根据干预真菌的不同共分9小组（低糖-对照组、高糖-对照组、糖波动-对照组、低糖-茄病镰刀菌组、高糖-茄病镰刀菌组、糖波动-茄病镰刀菌组、低糖-尖端赛多孢子菌组、高糖-尖端赛多孢子菌组、糖波动-尖端赛多孢子菌组）。真菌干预组均加入相应真菌孢子悬液，且孢子与HCEC的比例为10∶1，对照组加入等量的无菌生理盐水。观察真菌在培养板上的生长状况，倒置显微镜下观察每组HCEC的形态，荧光白染色法和平板稀释涂布法用于HCEC的黏附真菌计数，利用qRT-PCR检测各组HCEC中MMP-9 mRNA的表达、Western blot检测各组HCEC 中MMP-9蛋白的表达。结果　倒置显微镜下可见与真菌共培养的HCEC出现不同程度的形变和脱落，尤以高糖-茄病镰刀菌组和糖波动-茄病镰刀菌组最为显著。荧光白染色法和平板稀释涂布法计数结果显示，两种真菌高糖组和糖波动组对HCEC的黏附量均大于低糖组，差异均有统计学意义（均为P<0.05）；高糖和糖波动条件下，尖端赛多孢子菌黏附量大于茄病镰刀菌，差异均有统计学意义（均为P<0.05）。qRT-PCR和Western blot检测结果显示，MMP-9 mRNA和蛋白在各组HCEC中均有表达，与低糖组相比，高糖组和糖波动组HCEC中MMP-9 mRNA和蛋白表达均增多；高糖和糖波动条件下，真菌干预组HCEC中MMP-9 mRNA和蛋白表达均高于对照组，差异均有统计学意义（均为P<0.05），而两种真菌干预组之间差异均无统计学意义（均为P>0.05）。结论　高糖和糖波动环境能够促进HCEC分泌更多的MMP-9，并增加真菌对HCEC的黏附。
Abstract:
Objective　To investigate the adhesion ability of Fusarium solani (F.solani) and Scedosporium apiospermum (S.apiospermum) to human corneal epithelium cells (HCEC) and their effects on the expression of matrix metalloproteinase-9 (MMP-9) in HCEC under different concentrations of glucose.Methods　The cells in logarithmic growth were divided into 3 groups: low glucose group, in which cells were cultured with 5.5 mmol·L-1 glucose concentration medium, high glucose group, in which cells were cultured with 25 mmol·L-1 glucose concentration medium and fluctuating glucose group, in which cells were cultured with 5.5 mmol·L-1 and 50 mmol·L-1 glucose concentration medium alternately. Then the cells were divided into 9 groups according to different fungal interventions, and they were low glucose-control group, high glucose-control group, fluctuating glucose-control group, low glucose-F.solani intervention group, high glucose-F.solani intervention group, fluctuating glucose-F.solani intervention group, low glucose-S.apiospermum intervention group, high glucose-S.apiospermum intervention group and fluctuating glucose-S.apiospermum intervention group. The fungal intervention groups were treated with corresponding fungal spores suspension and the ratio of spores to HCEC was 10∶1. The control groups were given the sterile normal saline of equal volume. We observed the growth of fungi on the agar plate. The morphology of HCEC in each group was observed by the inverted microscope. Calcofluor white fluorescent staining and spread plate methods were used to count the adherent fungal spores of HCEC. Quantitative real-time PCR was used to determine the relative expression of MMP-9 mRNA and Western blot was used to determine the relative expression of MMP-9 proteins in HCEC. Results　We observed that the HCEC co-cultured with fungi showed varying degrees of deformation and abscission, especially in the high glucose-F.solani intervention group and fluctuating glucose-F.solani intervention group. The results of Calcofluor white fluorescent staining and spread plate methods showed that the adhesion population of fungi to HCEC was higher in high glucose groups and fluctuating glucose groups than that in the low glucose groups (all P<0.05). The adhesion population of S.apiospermum was higher than that of F.solani under the conditions of high glucose and fluctuating glucose, and the differences were statistically significant (all P<0.05). The result of Quantitative real-time PCR and Western blot showed that both MMP-9 mRNA and protein were expressed in HCEC of each group. The expression of MMP-9 mRNA and protein in HCEC was higher in high glucose groups and fluctuating glucose groups than that in low glucose groups. under the conditions of high glucose and fluctuating glucose, the expression levels of MMP-9 mRNA and protein in HCEC of fungal intervention groups was higher than those of the control groups (all P<0.05), but there was no statistically significant difference between the F.solani intervention group and S.apiospermum intervention group (P>0.05).Conclusion　High glucose and fluctuating glucose can promote MMP-9 secretion and increase the fungal adherence to the HCEC.

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相似文献/References:

[1]李妍,李丽红,刘海,等.树鼩茄病镰刀菌性角膜炎房水中Th1/Th2炎性因子的变化[J].眼科新进展,2017,37(9):824.[doi:10.13389/j.cnki.rao.2017.0208]
　LI Yan,LI Li-Hong,LIU Hai,et al.Changes of Th1/Th2 inflammatory factors in the aqueous humor of tree shrews with fusarium solani keratitis[J].Recent Advances in Ophthalmology,2017,37(6):824.[doi:10.13389/j.cnki.rao.2017.0208]
[2]李豪亮,朱然,张希熹,等.茄病镰刀菌对人角膜上皮细胞NOD2表达及相关炎症因子分泌水平的影响[J].眼科新进展,2021,41(7):633.[doi:10.13389/j.cnki.rao.2021.0130]
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备注/Memo

备注/Memo:
江苏省卫健委医学科研面上项目（编号：H2019083）；南通市市级科技计划项目（编号：MSZ19054）
更新日期/Last Update: 2021-06-05