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. 2020 Oct 20;40(10):1432–1438. [Article in Chinese] doi: 10.12122/j.issn.1673-4254.2020.10.08

薄荷醇增强白介素-13诱导的人支气管上皮细胞粘液蛋白5AC的合成与分泌

Menthol enhances interleukin-13-induced synthesis and secretion of mucin 5AC in human bronchial epithelial cells

Mingyang ZHANG 1, Jing WANG 1, Minchao LI 1,*
PMCID: PMC7606232  PMID: 33118512

Abstract

目的

研究白细胞介素-13（IL-13）联合薄荷醇对人支气管上皮细胞（16HBE）黏液蛋白（MUC）5AC合成及分泌的影响，并探索瞬时受体电位通道（TRP）melastatin 8（TRPM8）和抗凋亡因子B淋巴细胞瘤-2（Bcl-2）在此环节中所起的作用。

方法

将16HBE细胞分为：对照组、IL-13组（培养基加入终浓度10 ng/mL的IL-13连续刺激）、薄荷醇组（于第6天加入1 mmol/L薄荷醇刺激）、IL-13+薄荷醇组（IL-13连续刺激6 d后加入1 mmol/L薄荷醇刺激），观察各组MUC5AC表达量、胞内Ca2+浓度和Bcl-2表达；予以Bcl-2抑制剂ABT-263、TRPM8离子通道特异性抑制剂BCTC干预，观察两者对各组MUC5AC的影响及BCTC对胞内Ca2+浓度、Bcl-2表达的影响。CCK-8法检测细胞活力，流式细胞术检测各组细胞内Ca2+荧光强度，实时荧光定量PCR检测MUC5AC及Bcl-2mRNA水平，酶联免疫吸附法检测培养液中MUC5AC含量。

结果

IL-13组、薄荷醇组、IL-13+薄荷醇组MUC5AC mRNA和蛋白的表达均明显高于对照组（P < 0.05），且IL-13+薄荷醇组最高并于24 h达到最高峰（P < 0.01）；薄荷醇组、IL-13组、IL-13+薄荷醇组胞内Ca2+荧光强度、Bcl-2mRNA表达均明显高于对照组（P < 0.05），且IL-13+薄荷醇组最高（P < 0.01）；予以BCTC干预后，薄荷醇组、IL-13+薄荷醇组胞内Ca2+荧光强度、Bcl-2mRNA、MUC5AC mRNA和蛋白表达水平均显著低于相应的未干预组（P < 0.05），而IL-13组未发生明显变化（P>0.05）；予以ABT-263干预后，各ABT-263干预组MUC5ACmRNA和蛋白表达水平均显著低于相应的未干预组（P < 0.05）。

结论

IL-13联合薄荷醇对于16HBE细胞MUC5AC的合成及分泌产生协同效应，其机制可能与TRPM8受体诱导的Bcl-2协同性增强有关。

Keywords: 支气管哮喘, 白细胞介素-13, 薄荷醇, 黏液蛋白5AC, 抗凋亡蛋白, 瞬时受体电位melastatin 8

正常的黏液分泌是气道防御的屏障，但对于支气管哮喘（简称哮喘）患者而言，黏液的过度分泌导致黏液纤毛清除系统功能失常和局部防御功能损害，从而引起呼吸道反复感染。感染和黏液高分泌互为因果，加速了疾病的发展，加重了疾病的病残率和病死率[1-2]。而冷空气刺激是哮喘的一个重要急性加重因素，受到冷空气刺激时表现为气道痉挛阻塞、炎性因子浸润及中性粒细胞募集、黏液蛋白分泌增加，从而导致病情急剧加重[3-4]。许多研究表明，多数哮喘内因型系经典的TH2高型，细胞因子白细胞介素（IL）-4、IL-5及IL-13与TH2型哮喘的发病有着紧密联系，其中IL-13是目前最重要的细胞因子，它可以引起气道炎症、黏液过度分泌、杯状细胞增生和黏蛋白（MUC）5AC高表达等多种改变[5-6]，而抗凋亡蛋白B淋巴细胞瘤-2（Bcl-2）可能参与该病理过程[7-8]。有研究表明Bcl-2和MUC5AC在气道上皮细胞中共表达，并且Bcl-2是MUC5AC上游重要调控蛋白[9-10]。近些年研究表明瞬时受体电位通道（TRP）melastatin 8（TRPM8）是Ca2+可渗透的非选择性阳离子通道，在神经组织和非神经组织中均有广泛表达，是机体感知冷刺激的重要受体。研究证实在人支气管上皮细胞存在TRPM8异构体的表达，冷刺激和薄荷醇均能激活TRPM8从而诱导炎性因子产生及黏液高分泌[1, 11-12]。薄荷醇不仅具有和冷刺激相同的激活TRPM8通道并产生类似的病理生理学效应的功能，同时薄荷醇还可以发挥一定的抗凋亡活性[13-14]，但是其对气道上皮细胞是否具有抗凋亡活性仍然未知。因此，我们采用IL-13连续作用于16HBE细胞，用于模拟哮喘的气道炎性环境，用薄荷醇模拟冷空气刺激环境，试图明确冷刺激是否能加重哮喘患者气道黏液高分泌，并试图探索TRPM8和Bcl-2在此过程中发挥的作用及可能涉及的机制研究。

1. 材料和方法

1.1. 试剂和材料

人支气管上皮细胞系16HBE、KM无血清培养基（富衡）；RPMI 1640培养基、胎牛血清（Gibco）；薄荷醇（阿拉丁）；RNA提取试剂（北京天根）；逆转录试剂盒ReverTra Ace-α（-上海东洋纺）；酶联免疫吸附测定试剂盒（武汉基因美）；Fluo-4AM（碧云天）；Bcl-2抑制剂ABT-263（MCE）；TRPM8特异性阻断剂BCTC（MCE）。

1.2. 细胞的培养与分组处理

细胞以5×105/孔的密度接种于6孔板，于37 ℃、5% CO2的培养箱中培养，每隔48 h换液。细胞融合至80% ~90%时给予无血清培养基，将细胞分为：①对照组；② IL-13组：培养基加入IL-13（终浓度10 ng/mL）连续刺激；③薄荷醇组：于第6天细胞中加入1 mmol/L薄荷醇刺激；④IL-13+薄荷醇组（IL-13连续刺激6 d后加入1 mmol/L薄荷醇刺激）；上述4组培养8、16、24、36 h后收集各个时间点的细胞和培养上清液，测定细胞MUC5AC mRNA及培养液中MUC5AC蛋白含量。选择MUC5AC mRNA及蛋白含量最高的时间点，予以Bcl-2抑制剂ABT-263以及TRPM8抑制剂BCTC干预处理，检测其干预效应，并依次分为：⑤IL-13+ABT-263组：细胞在最后1次换液前给予1 μmol/L ABT-263干预2 h处理；⑥薄荷醇+ABT-263组：在加入薄荷醇之前予以1 μmol/L ABT-263干预2 h处理；⑦IL-13+薄荷醇+ ABT-263组：IL-13连续刺激，并在薄荷醇之前予以1 μmol/L ABT-263干预2 h处理，同理，在加入薄荷醇4 h前予以BCTC 15 μmol/L干预处理，依次分组为：⑧ IL-13+BCTC组；⑨薄荷醇+BCTC组；⑩IL-13+薄荷醇+BCTC组。各组均设3个复孔。

1.3. CCK-8法检测细胞活力

取对数生长期16HBE细胞，2.5 g/L胰酶消化，以1×104/mL细胞的密度，每孔加入100 μL的细胞悬液接种于96孔板，按前述①~④组方法进行培养，培养至8、16、24、36 h时，吸弃上清液，每孔加入100 μL培养基及10 μL CCK8溶液继续孵育培养1 h，酶标仪上450 nm处检测吸光度A。细胞存活率（%）=[（A实验－A空白）（/A对照－ A空白）]×100%，每组5个副孔。

1.4. 流式细胞术检测各组细胞内Ca2+荧光强度

在1.2小节实验分组的基础上，增设一组正常对照组，其相较于空白对照组，需负载与各实验组等量的Fluo-4AM细胞。在用薄荷醇处理10 min后，将除空白对照组外的各组细胞重悬于无Ca2+缓冲液中，加入终浓度为5 μmol/L Fluo-4AM，37 ℃避光孵育30 min，而后用无Ca2+缓冲液洗涤3次，离心5 min后，用无Ca2+缓冲液1 mL重悬。而空白对照组消化重悬1 mL无Ca2+缓冲液中。

1.5. 实时荧光定量PCR法检测MUC5AC、Bcl-2mRNA的表达

各组细胞培养至设定时间，按Trizol试剂说明书分别提取各组细胞总RNA，按高效率逆转录试剂盒ReverTraAce-α-逆转录合成cDNA。PCR仪进行Real-time PCR扩增：95 ℃预变性30 s，95 ℃变性5 s，60 ℃退火30 s，共40个循环。以β-actin为参照基因，采用2－△△Ct相对定量表示各组MUC5AC和Bcl-2mRNA的表达量。

1.6. ELISA法检测细胞培养液中MUC5AC蛋白表达

用无菌离心管收集培养8、16、24、36 h后的细胞上清液，2000~3000 r/min离心20 min左右，仔细收集上清。用酶联免疫吸附测定试剂盒检测胞外MUC5AC水平。

1.7. 统计学分析

采用SPSS 23.0统计软件处理实验数据，实验结果以均数±标准差表示，多样本均数间比较采用单因素方差分析，P < 0.05表示差异有统计学意义。

2. 结果

2.1. 细胞活性检测结果

CCK8检测结果显示，在8、16、24、36 h时，不同细胞组细胞之间的细胞活力无显著差异（P > 0.05），这表明IL-13或/和薄荷醇对细胞活力没有显著影响。

1.

Real-time PCR引物序列

Sequences of primers for real-time PCR

Primer name Primer sequence
MUC5AC-F 5'-CCCAAACCTGCTTCTGCAAC-3'
MUC5AC-R 5'-GGGCTCGGAGGTGGATATTG-3'
Bcl-2-F 5'-GTGGCCTTCTTTGAGTTCGG-3'
Bcl-2-R 5'-CGGTTCAGGTACTCAGTCATCC-3'
β-actin-F 5'-CCTGTACGCCAACACAGTGC-3'
β-actin-R 5'-ATACTCCTGCTTGCTGATCC-3'
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2.2. 薄荷醇和或IL-13对MUC5AC合成及分泌水平的影响

实时荧光定量PCR结果显示，在8、16、24、36 h，IL-13组MUC5AC mRNA的表达量均约为对照组的1.8倍（P < 0.05），而薄荷醇组MUC5AC mRNA的表达于8 h后明显升高，并于24 h达到峰值（P < 0.01）。IL-13+薄荷醇组MUC5AC mRNA的表达于16 h后开始明显升高，峰值出现在薄荷醇处理后24 h（图 1A）。

1.

1

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不同处理时间胞内及胞外MUC5AC的相对表达量

Relative expression of intracellular and extracellular MUC5AC at different points of treatment detected using qPCR (A) and ELISA (B). *P < 0.05, **P < 0.01 vs control group, #P < 0.05 vs IL-13 group, △P < 0.05 vs menthol group.

ELISA结果显示，在8、16、24、36 h，IL-13组胞外MUC5AC蛋白的表达量均约为对照组的1.5倍（P < 0.05），薄荷醇组胞外MUC5AC蛋白的表达于8 h后明显升高，并于24 h达到峰值（P < 0.01）。IL-13+薄荷醇组胞外MUC5AC蛋白的表达于16 h后开始明显升高，峰值出现在薄荷醇处理24 h后（图 1B）。

2.3. TRPM8抑制剂对薄荷醇和/或IL-13诱导的胞内Ca2+升高的影响

流式细胞检测细胞内Ca2+荧光强度结果显示，与空白对照组相比，正常对照组胞内Ca2+荧光强度升高（P < 0.05），表明正常对照组中成功装载Fluo-4 AM钙离子探针。与正常对照组相比，IL-13组、薄荷醇组、IL-13+薄荷醇组胞内Ca2+荧光强度均升高（P < 0.05），并且IL- 13+薄荷醇组对细胞内Ca2+荧光强度的升高更加明显（P < 0.01）。与IL-13组相比，IL-13+BCTC组细胞内Ca2+荧光强度无明显差异（P > 0.05），薄荷醇+BCTC组、IL-13+薄荷醇+BCTC组细胞内Ca2+荧光强度分别低于薄荷醇组、IL-13+薄荷醇组（P < 0.05），并且IL-13+薄荷醇+BCTC组降低到接近IL-13组水平（图 2）。

2.

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流式细胞术检测各组细胞内Ca2+荧光强度

Flow cytometry detection of intracellular Ca2+ fluorescence intensity. #P < 0.05 υs blank control group, *P < 0.05, **P < 0.01 υs normal control group, △P < 0.05 υs menthol group, ▼P < 0.05 υs IL-13 group, ▲P < 0.05 υs IL-13+menthol group. A: Blank control group; B: Normal control group; C: IL-13 group; D: Menthol group; E: IL-13+menthol group; F: IL-13+ BCTC group; G: menthol+BCTC group; H: IL-13+menthol+BCTC group.

2.4. Bcl-2和TRPM8抑制剂对于MUC5AC合成及分泌水平的影响

基于MUC5AC mRNA和蛋白相对表达量于24 h显著增强，于24 h后获取细胞行实时荧光定量PCR及ELISA检测ABT-263和BCTC的干预作用。

实时荧光定量PCR结果显示：IL-13组与IL-13+ BCTC组比较，MUC5AC mRNA的表达未发生明显变化（P > 0.05），薄荷醇+BCTC组、IL-13+薄荷醇+BCTC组MUC5AC mRNA的表达分别低于薄荷醇组、IL-13+薄荷醇组（P < 0.05），并且IL-13+薄荷醇+BCTC组降低到接近IL-13组水平（图 3A）；IL-13+ABT-263组（1.43± 0.09）、薄荷醇+ABT-263组（1.76±0.11）、IL-13+薄荷醇+ ABT-263组（3.71±0.1）MUC5AC mRNA的表达分别低于IL-13组（2.13±0.05）、薄荷醇组（2.52±0.29）、IL-13+薄荷醇组（5.75±0.4）（P < 0.05，图 3B）。

3.

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在24 h时间点各组中胞内及胞外MUC5AC的相对表达量

Relative expression of intracellular and extracellular MUC5AC in each group at 24 h detected using qPCR (A, B) and ELISA (C, D). *P < 0.05, **P < 0.01 vs control group, #P < 0.05 vs IL-13 group, △P < 0.05 vs menthol group, ▲P < 0.05 vs IL-13+menthol group.

ELISA结果显示：IL-13组与IL-13+BCTC组比较，MUC5AC分泌蛋白水平未发生明显变化（P > 0.05），薄荷醇+BCTC组、IL-13+薄荷醇+BCTC组MUC5AC分泌蛋白水平分别低于薄荷醇组、IL-13+薄荷醇组（P < 0.05），并且IL-13+薄荷醇组+BCTC组降低到接近IL-13组分泌水平（图 3C）；IL-13+ABT-263组（1.25±0.07）、薄荷醇+ABT-263组（1.44±0.09）、IL-13+薄荷醇+ABT-263组（2.06±0.04）MUC5AC分泌蛋白水平分别低于IL-13组（1.55±0.07）、薄荷醇组（1.97±0.07）、IL-13+薄荷醇组（3.72±0.06）（P < 0.05，图 3D）。

2.5. TRPM8对Bcl-2表达的影响

实时荧光定量PCR结果显示：IL-13组和薄荷醇组16HBE细胞Bcl-2 mRNA表达量与空白对照组相比明显上升（P < 0.05），IL-13+薄荷醇组16HBE细胞Bcl-2 mRNA表达高于IL-13及薄荷醇组（P < 0.05）。予以BCTC干预处理后，薄荷醇+BCTC组（1.42±0.08）、IL-13+薄荷醇+BCTC组（1.74±0.09）的Bcl-2 mRNA水平分别低于薄荷醇组（2.13±0.11）、IL-13+薄荷醇组（3.26± 0.26）（P < 0.05），IL-13 +BCTC组（1.63±0.08）与IL-13组（1.73±0.09）相比无明显差异（P > 0.05，图 4）。

4.

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TRPM8在薄荷醇对IL-13诱导的Bcl-2mRNA表达的影响

Effect of TRPM8 on IL-13-induced Bcl-2 mRNA expression in menthol-treated cells. *P < 0.05, **P < 0.01 vs control group, #P < 0.05 vs IL-13 group, △P < 0.05 vs menthol group, ▲P < 0.05 vs IL-13+ menthol group.

3. 讨论

黏液高分泌是慢性气道炎症性疾病包括COPD、支气管哮喘、囊性纤维化等共同而重要的病理生理特征之一。MUC5AC是人气道上皮细胞在对TH2/IL-13刺激的体外应答中增加的主要黏蛋白，IL-13是由T型辅助2型（TH2）细胞产生的细胞因子，在动物模型和人气道上皮中均显示IL-13会增加上皮细胞黏液[15-16]。虽然既往有研究报道过薄荷醇和IL-13均会引起MUC5AC合成及分泌的增加[1, 17]，但是本研究结果首次证明了经过IL-13预处理过的细胞，薄荷醇对于MUC5AC合成及分泌有显著的增强效应。在临床上对于哮喘患者而言，冷刺激作用于气道上皮细胞后，因其气道内大量炎性因子IL-13的浸润，冷刺激与IL-13产生协同效应，势必会导致黏液的过度分泌，从而引起疾病的急性发作或加重。

目前对于冷刺激诱发哮喘急性发作或加重的病理生理机制研究主要集中在TRPM8冷敏感通道。TRPM8被低温或冷物质模拟剂激活后可引起细胞内Ca2+浓度升高，进一步导致IL-4、IL-8、IL-13及肿瘤坏死因子等多种炎症因子的产生[18-19]。抗凋亡蛋白Bcl-2作为Bcl-2家族中的重要一员，通过抑制线粒体细胞色素C的释放而发挥存活功能，在细胞凋亡过程中发挥着重要作用[20]。本实验发现薄荷醇和IL-13都能引起细胞内Ca2+浓度的升高，TRPM8阻断剂BCTC抑制了薄荷醇引起的Ca2+浓度的升高，然而BCTC并未阻止IL-13引起的Ca2+浓度的升高，说明薄荷醇能够激活TRPM8受体，而IL-13引起的Ca2+浓度的升高与TRPM8受体无关。进一步实验发现，TRPM8阻断剂BCTC显著抑制薄荷醇诱导黏液蛋白的表达，说明薄荷醇诱导黏液蛋白的表达与TRPM8通道蛋白有关，这与前期实验结果一致[21]；并且薄荷醇能上调16HBE细胞Bcl-2的表达，当运用BCTC干预处理后，Bcl-2表达明显下调，说明薄荷醇上调Bcl-2的表达与TRPM8通道蛋白有关；IL-13和薄荷醇均能升高Bcl-2的表达，且二者具有协同性升高Bcl-2的作用，Bcl-2抑制剂ABT-263可使IL-13和薄荷醇组诱导的MUC5AC合成及分泌水平显著降低，并且能够减弱IL-13联合薄荷醇对MUC5AC合成及分泌的协同增强效应，说明薄荷醇协同性增强IL-13诱导MUC5AC合成及分泌的作用与二者协同性上调Bcl-2的作用有关。

薄荷醇抗凋亡作用在非呼吸系统疾病的研究中有零星报道，该实验发现薄荷醇能够通过激活TRPM8受体通道来上调Bcl-2的表达，发挥一定的抗凋亡活性，且和IL-13协同性地诱导MUC5AC的合成及分泌。其机制可能基于TRPM8通路依赖途径：①TRPM8通路激活后引起细胞内Ca2+浓度升高进一步导致IL-8、IL-13产生。有研究报道，炎性因子IL-8在前列腺癌以及肝癌细胞中上调Bcl-2、Bcl-xL的表达抑制细胞的凋亡，从而增加肿瘤细胞的存活率[22-24]；IL-13水平的升高可进一步上调Bcl-2进而抑制凋亡，从而体现出薄荷醇与IL-13的协同作用；②前期研究工作表明，冷刺激或薄荷醇激活TRPM8后经由Ca2+-磷脂酶C-磷脂酰肌醇二磷酸-蛋白激酶C（PKC）通路参与了MUC5AC产生的过程。有研究表明，PKC的激活可以通过上调Bcl-2的表达抑制细胞凋亡[25-26]，因此推测冷刺激激活TRPM8后PKC被活化，通过上调Bcl-2的表达在16HBE细胞中发挥抗凋亡的作用，进而对下游MUC5AC的合成及分泌产生影响。另有研究表明，IL-13亦能够激活PKC[27-28]，薄荷醇与IL-13的协同作用可能与二者共同激活PKC有关。但目前PKC对细胞凋亡的影响仍存在着一些争议[29-30]，冷刺激激活后的PKC是否能上调Bcl-2的表达从而发挥抗凋亡作用还需要相关实验进一步验证。

综上所述，本研究建立细胞体外哮喘模型，初步证实冷空气刺激模拟物薄荷醇可协同性上调IL-13对HBE细胞MUC5AC的合成与分泌作用，并阐明了其中潜在的细胞机制。从临床角度讲，冷空气刺激会导致哮喘患者黏液分泌进一步增加从而诱发哮喘的急性加重，故该实验为冷空气刺激诱发哮喘的急性发作或加重的潜在机制提供了理论基础，为哮喘的防控提供了一个新的视角。

Biography

张明洋，在读硕士研究生，E-mail: 974902710@qq.com

Funding Statement

国家自然科学面上基金（81270102）；重庆市自然科学基金（cstc2012jjA10050）；重庆市教委科学技术研究项目（KJ120301）

Supported by National Natural Science Foundation of China (81270102)

Contributor Information

张 明洋 (Mingyang ZHANG), Email: 974902710@qq.com.

李 敏超 (Minchao LI), Email: liminc66@aliyun.com.

References

  • 1.Li MC, Li Q, Yang G, et al. Cold temperature induces mucin hypersecretion from normal human bronchial epithelial cells in vitro through a transient receptor potential melastatin 8 (TRPM8)- mediated mechanism. J Allergy Clin Immunol. 2011;128(3):626–34. doi: 10.1016/j.jaci.2011.04.032. [Li MC, Li Q, Yang G, et al. Cold temperature induces mucin hypersecretion from normal human bronchial epithelial cells in vitro through a transient receptor potential melastatin 8 (TRPM8)- mediated mechanism[J]. J Allergy Clin Immunol, 2011, 128(3): 626-34.e1-5.] [DOI] [PubMed] [Google Scholar]
  • 2.D'Amato M, Molino A, Calabrese G, et al. The impact of cold on the respiratory tract and its consequences to respiratory health. Clin Transl Allergy. 2018;8:20. doi: 10.1186/s13601-018-0208-9. [D'Amato M, Molino A, Calabrese G, et al. The impact of cold on the respiratory tract and its consequences to respiratory health[J]. Clin Transl Allergy, 2018, 8: 20.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Koskela HO. Cold air-provoked respiratory symptoms: the mechanisms and management. Int J Circumpolar Health. 2007;66(2):91–100. doi: 10.3402/ijch.v66i2.18237. [Koskela HO. Cold air-provoked respiratory symptoms: the mechanisms and management[J]. Int J Circumpolar Health, 2007, 66(2): 91-100.] [DOI] [PubMed] [Google Scholar]
  • 4.Hyrkäs H, Ikäheimo TM, Jaakkola JJ, et al. Asthma control and cold weather-related respiratory symptoms. Respir Med. 2016;113:1–7. doi: 10.1016/j.rmed.2016.02.005. [Hyrkäs H, Ikäheimo TM, Jaakkola JJ, et al. Asthma control and cold weather-related respiratory symptoms[J]. Respir Med, 2016, 113: 1-7.] [DOI] [PubMed] [Google Scholar]
  • 5.Zhao JM, Minami Y, Etling E, et al. Preferential generation of 15- HETE-PE induced by IL-13 regulates goblet cell differentiation in human airway epithelial cells. Am J Respir Cell Mol Biol. 2017;57(6):692–701. doi: 10.1165/rcmb.2017-0031OC. [Zhao JM, Minami Y, Etling E, et al. Preferential generation of 15- HETE-PE induced by IL-13 regulates goblet cell differentiation in human airway epithelial cells[J]. Am J Respir Cell Mol Biol, 2017, 57(6): 692-701.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 6.Yu HM, Li Q, Kolosov VP, et al. Interleukin-13 induces mucin 5AC production involving STAT6/SPDEF in human airway epithelial cells. http://www.wanfangdata.com.cn/details/detail.do?_type=perio&id=a36d86e4d7ff25a454285f780ecccfe1. Cell Commun Adhes. 2010;17(4/5/6):83–92. doi: 10.3109/15419061.2010.551682. [Yu HM, Li Q, Kolosov VP, et al. Interleukin-13 induces mucin 5AC production involving STAT6/SPDEF in human airway epithelial cells[J]. Cell Commun Adhes, 2010, 17(4/5/6): 83-92.] [DOI] [PubMed] [Google Scholar]
  • 7.Chand HS, Harris JF, Tesfaigzi Y. IL-13 in LPS-induced inflammation causes bcl-2 expression to sustain hyperplastic mucous cells. Sci Rep. 2018;8(1):436. doi: 10.1038/s41598-017-18884-9. [Chand HS, Harris JF, Tesfaigzi Y. IL-13 in LPS-induced inflammation causes bcl-2 expression to sustain hyperplastic mucous cells[J]. Sci Rep, 2018, 8(1): 436.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 8.Chand HS, Mebratu YA, Kuehl PJ, et al. Blocking Bcl-2 resolves IL- 13-mediated mucous cell hyperplasia in a Bik-dependent manner. J Allergy Clin Immunol. 2017;140(5):1456–9. doi: 10.1016/j.jaci.2017.05.038. [Chand HS, Mebratu YA, Kuehl PJ, et al. Blocking Bcl-2 resolves IL- 13-mediated mucous cell hyperplasia in a Bik-dependent manner [J]. J Allergy Clin Immunol, 2017, 140(5): 1456-9.e9.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Chand HS, Woldegiorgis Z, Schwalm K, et al. Acute inflammation induces insulin-like growth factor-1 to mediate Bcl-2 and Muc5ac expression in airway epithelial cells. Am J Respir Cell Mol Biol. 2012;47(6):784–91. doi: 10.1165/rcmb.2012-0079OC. [Chand HS, Woldegiorgis Z, Schwalm K, et al. Acute inflammation induces insulin-like growth factor-1 to mediate Bcl-2 and Muc5ac expression in airway epithelial cells[J]. Am J Respir Cell Mol Biol, 2012, 47(6): 784-91.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 10.Takeyama K, Tamaoki J, Kondo M, et al. Role of epidermal growth factor receptor in maintaining airway goblet cell hyperplasia in rats sensitized to allergen. Clin Exp Allergy. 2008;38(5):857–65. doi: 10.1111/j.1365-2222.2008.02951.x. [Takeyama K, Tamaoki J, Kondo M, et al. Role of epidermal growth factor receptor in maintaining airway goblet cell hyperplasia in rats sensitized to allergen[J]. Clin Exp Allergy, 2008, 38(5): 857-65.] [DOI] [PubMed] [Google Scholar]
  • 11.Kim JH, Jang YS, Kim HI, et al. Activation of transient receptor potential melastatin family member 8 (TRPM8) receptors induces proinflammatory cytokine expressions in bronchial epithelial cells. Allergy Asthma Immunol Res. 2020;12(4):684–700. doi: 10.4168/aair.2020.12.4.684. [Kim JH, Jang YS, Kim HI, et al. Activation of transient receptor potential melastatin family member 8 (TRPM8) receptors induces proinflammatory cytokine expressions in bronchial epithelial cells [J]. Allergy Asthma Immunol Res, 2020, 12(4): 684-700.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Sabnis AS, Shadid M, Yost GS, et al. Human lung epithelial cells express a functional cold-sensing TRPM8 variant. Am J Respir Cell Mol Biol. 2008;39(4):466–74. doi: 10.1165/rcmb.2007-0440OC. [Sabnis AS, Shadid M, Yost GS, et al. Human lung epithelial cells express a functional cold-sensing TRPM8 variant[J]. Am J Respir Cell Mol Biol, 2008, 39(4): 466-74.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 13.Rozza AL, Meira de Faria F, Souza Brito AR, et al. The gastroprotective effect of menthol: involvement of anti-apoptotic, antioxidant and anti-inflammatory activities. PLoS One. 2014;9(1):e86686. doi: 10.1371/journal.pone.0086686. [Rozza AL, Meira de Faria F, Souza Brito AR, et al. The gastroprotective effect of menthol: involvement of anti-apoptotic, antioxidant and anti-inflammatory activities[J]. PLoS One, 2014, 9(1): e86686.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Muruganathan U, Srinivasan S, Vinothkumar V. Antidiabetogenic efficiency of menthol, improves glucose homeostasis and attenuates pancreatic β-cell apoptosis in streptozotocin-nicotinamide induced experimental rats through ameliorating glucose metabolic enzymes. Biomedecine Pharmacother. 2017;92:229–39. doi: 10.1016/j.biopha.2017.05.068. [Muruganathan U, Srinivasan S, Vinothkumar V. Antidiabetogenic efficiency of menthol, improves glucose homeostasis and attenuates pancreatic β-cell apoptosis in streptozotocin-nicotinamide induced experimental rats through ameliorating glucose metabolic enzymes [J]. Biomedecine Pharmacother, 2017, 92: 229-39.] [DOI] [PubMed] [Google Scholar]
  • 15.Fujisawa T, Ide K, Holtzman MJ, et al. Involvement of the p38 MAPK pathway in IL-13-induced mucous cell Metaplasia in mouse tracheal epithelial cells. Respirology. 2008;13(2):191–202. doi: 10.1111/j.1440-1843.2008.01237.x. [Fujisawa T, Ide K, Holtzman MJ, et al. Involvement of the p38 MAPK pathway in IL-13-induced mucous cell Metaplasia in mouse tracheal epithelial cells[J]. Respirology, 2008, 13(2): 191-202.] [DOI] [PubMed] [Google Scholar]
  • 16.Kondo M, Tamaoki J, Takeyama K, et al. Elimination of IL-13 reverses established goblet cell Metaplasia into ciliated epithelia in airway epithelial cell culture. Allergol Int. 2006;55(3):329–36. doi: 10.2332/allergolint.55.329. [Kondo M, Tamaoki J, Takeyama K, et al. Elimination of IL-13 reverses established goblet cell Metaplasia into ciliated epithelia in airway epithelial cell culture[J]. Allergol Int, 2006, 55(3): 329-36.] [DOI] [PubMed] [Google Scholar]
  • 17.Hao WM, Wang J, Zhang Y, et al. Leptin positively regulates MUC5AC production and secretion induced by interleukin-13 in human bronchial epithelial cells. Biochem Biophys Res Commun. 2017;493(2):979–84. doi: 10.1016/j.bbrc.2017.09.106. [Hao WM, Wang J, Zhang Y, et al. Leptin positively regulates MUC5AC production and secretion induced by interleukin-13 in human bronchial epithelial cells[J]. Biochem Biophys Res Commun, 2017, 493(2): 979-84.] [DOI] [PubMed] [Google Scholar]
  • 18.Liu HP, Hua L, Liu QH, et al. Cold stimuli facilitate inflammatory responses through transient receptor potential melastatin 8 (TRPM8) in primary airway epithelial cells of asthmatic mice. Inflammation. 2018;41(4):1266–75. doi: 10.1007/s10753-018-0774-y. [Liu HP, Hua L, Liu QH, et al. Cold stimuli facilitate inflammatory responses through transient receptor potential melastatin 8 (TRPM8) in primary airway epithelial cells of asthmatic mice[J]. Inflammation, 2018, 41(4): 1266-75.] [DOI] [PubMed] [Google Scholar]
  • 19.李 敏超, PERELMAN JM, KOLOSOV VP, et al. 瞬时受体电位M8离子通道对冷刺激诱导气道上皮细胞炎症反应的影响. http://www.wanfangdata.com.cn/details/detail.do?_type=perio&id=zhjhhhx201110010. 中华结核和呼吸杂志. 2011;34(10):1–5. [李敏超, PERELMAN JM, KOLOSOV VP, 等.瞬时受体电位M8离子通道对冷刺激诱导气道上皮细胞炎症反应的影响[J].中华结核和呼吸杂志, 2011, 34(10): 1-5.] [Google Scholar]
  • 20.Lewis A, Hayashi T, Su TP, et al. Bcl-2 family in inter-organelle modulation of calcium signaling; roles in bioenergetics and cell survival. http://www.wanfangdata.com.cn/details/detail.do?_type=perio&id=1ce77b3d4ea2ddfe8396590b1105cab9. J Bioenerg Biomembr. 2014;46(1):1–15. doi: 10.1007/s10863-013-9527-7. [Lewis A, Hayashi T, Su TP, et al. Bcl-2 family in inter-organelle modulation of calcium signaling; roles in bioenergetics and cell survival[J]. J Bioenerg Biomembr, 2014, 46(1): 1-15.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Liu SC, Lu HH, Fan HC, et al. The identification of the TRPM8 channel on primary culture of human nasal epithelial cells and its response to cooling. Medicine (Baltimore) 2017;96(31):e7640. doi: 10.1097/MD.0000000000007640. [Liu SC, Lu HH, Fan HC, et al. The identification of the TRPM8 channel on primary culture of human nasal epithelial cells and its response to cooling[J]. Medicine (Baltimore), 2017, 96(31): e7640.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Choi SH, Park JY, Kang W, et al. Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells. Apoptosis. 2016;21(1):85–95. doi: 10.1007/s10495-015-1185-2. [Choi SH, Park JY, Kang W, et al. Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells[J]. Apoptosis, 2016, 21(1): 85-95.] [DOI] [PubMed] [Google Scholar]
  • 23.Singh RK, Lokeshwar BL. Depletion of intrinsic expression of Interleukin-8 in prostate cancer cells causes cell cycle arrest, spontaneous apoptosis and increases the efficacy of chemotherapeutic drugs. Mol Cancer. 2009;8:57. doi: 10.1186/1476-4598-8-57. [Singh RK, Lokeshwar BL. Depletion of intrinsic expression of Interleukin-8 in prostate cancer cells causes cell cycle arrest, spontaneous apoptosis and increases the efficacy of chemotherapeutic drugs[J]. Mol Cancer, 2009, 8: 57.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Stronach EA, Cunnea P, Turner C, et al. The role of interleukin-8 (IL-8) and IL-8 receptors in platinum response in high grade serous ovarian carcinoma. Oncotarget. 2015;6(31):31593–603. doi: 10.18632/oncotarget.3415. [Stronach EA, Cunnea P, Turner C, et al. The role of interleukin-8 (IL-8) and IL-8 receptors in platinum response in high grade serous ovarian carcinoma[J]. Oncotarget, 2015, 6(31): 31593-603.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Ge XW, Chen JF, Li L, et al. Midostaurin potentiates rituximab antitumor activity in Burkitt's lymphoma by inducing apoptosis. http://www.nature.com/articles/s41419-018-1259-5. Cell Death Dis. 2018;10(1):8. doi: 10.1038/s41419-018-1259-5. [Ge XW, Chen JF, Li L, et al. Midostaurin potentiates rituximab antitumor activity in Burkitt's lymphoma by inducing apoptosis[J]. Cell Death Dis, 2018, 10(1): 8.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 26.Weinreb O, Amit T, Bar-Am O, et al. Novel neuroprotective mechanism of action of rasagiline is associated with its propargyl moiety: interaction of Bcl-2 family members with PKC pathway. Ann N Y Acad Sci. 2005;1053:348–55. doi: 10.1196/annals.1344.030. [Weinreb O, Amit T, Bar-Am O, et al. Novel neuroprotective mechanism of action of rasagiline is associated with its propargyl moiety: interaction of Bcl-2 family members with PKC pathway[J]. Ann N Y Acad Sci, 2005, 1053: 348-55.] [DOI] [PubMed] [Google Scholar]
  • 27.Moriya C, Jinnin M, Yamane K, et al. Expression of matrix metalloproteinase-13 is controlled by IL-13 via PI3K/Akt3 and PKC-δ in normal human dermal fibroblasts. J Investig Dermatol. 2011;131(3):655–61. doi: 10.1038/jid.2010.361. [Moriya C, Jinnin M, Yamane K, et al. Expression of matrix metalloproteinase-13 is controlled by IL-13 via PI3K/Akt3 and PKC-δ in normal human dermal fibroblasts[J]. J Investig Dermatol, 2011, 131(3): 655-61.] [DOI] [PubMed] [Google Scholar]
  • 28.Sui GD, Cheng G, Yuan JJ, et al. Interleukin (IL)-13, prostaglandin E2 (PGE2), and prostacyclin 2 (PGI2) activate hepatic stellate cells via protein kinase C (PKC) pathway in hepatic fibrosis. Med Sci Monit. 2018;24:2134–41. doi: 10.12659/MSM.906442. [Sui GD, Cheng G, Yuan JJ, et al. Interleukin (IL)-13, prostaglandin E2 (PGE2), and prostacyclin 2 (PGI2) activate hepatic stellate cells via protein kinase C (PKC) pathway in hepatic fibrosis[J]. Med Sci Monit, 2018, 24: 2134-41.] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 29.Lu PW, Gu YT, Li L, et al. Belinostat suppresses cell proliferation by inactivating Wnt/β-catenin pathway and promotes apoptosis through regulating PKC pathway in breast cancer. Artif Cells Nanomed Biotechnol. 2019;47(1):3955–60. doi: 10.1080/21691401.2019.1671855. [Lu PW, Gu YT, Li L, et al. Belinostat suppresses cell proliferation by inactivating Wnt/β-catenin pathway and promotes apoptosis through regulating PKC pathway in breast cancer[J]. Artif Cells Nanomed Biotechnol, 2019, 47(1): 3955-60.] [DOI] [PubMed] [Google Scholar]
  • 30.Lu QB, Du Q, Wang HP, et al. Salusin-β mediates tubular cell apoptosis in acute kidney injury: Involvement of the PKC/ROS signaling pathway. Redox Biol. 2020;30:101411. doi: 10.1016/j.redox.2019.101411. [Lu QB, Du Q, Wang HP, et al. Salusin-β mediates tubular cell apoptosis in acute kidney injury: Involvement of the PKC/ROS signaling pathway[J]. Redox Biol, 2020, 30: 101411.] [DOI] [PMC free article] [PubMed] [Google Scholar]

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