尤其　　郭辉　　孔凡慧　　王冬梅

[摘要] 目的 研究聯氨基姜黄素诱导DC细胞成熟抑制小鼠皮肤鳞癌进展的机制。 方法 梯度浓度（1/5/10/50/100/500 μM）的联氨基姜黄素处理SCC细胞系SCL-1 48 h，MTT检测细胞活性。同时构建自发性SCC小鼠模型，给予药物处理观察肿瘤进展情况，流式检测肿瘤组织树突状成熟状态。Western blot检测联氨基姜黄素处理前后树突状细胞STAT3通路的变化。 结果 不同浓度的联氨基姜黄素在处理SCC细胞系SCL-1的过程中可能会对细胞的活性产生抑制作用，差异有统计学意义（P<0.05）。 半数最大抑制浓度值（IC50）为18.7 μM。正常C57BL/6小鼠在为期2周的肠胃给药处理以后，通过与对照组进行比较发现其体重的变化并不显著。自发性SCC小鼠模型在接受联氨基姜黄素处理后，抗肿瘤的效果较为突出，差异有统计学意义（P<0.05）。流式检测SCC小鼠的肿瘤组织，发现联氨基姜黄素处理可促进树突状细胞成熟（CD80/CD86明显上调），差异有统计学意义（P<0.05）。进一步Western blot检测联氨基姜黄素处理前后树突状细胞 STAT3通路发现处理组 STAT3磷酸化水平显著下调，差异有统计学意义（P<0.05）。 结论 联氨基姜黄素可在体外杀伤SCC细胞，并在体内促进树突状细胞成熟进而抑制SCC进展，其可能的机制是通过抑制 STAT3磷酸化，抑制STAT3通路。

[关键词] 联氨基姜黄素;皮肤鳞状细胞癌;增殖;树突状细胞;STAT3

[中图分类号] R739.5          [文献标识码] A          [文章编号] 1673-9701（2021）12-0039-04

Study on the mechanism of DC maturation induced by hydrazinocurcumin in inhibiting mouse skin squamous cell carcinoma

YOU Qi1   GUO Hui2   KONG Fanhui1   WANG Dongmei3

1.Department of Clinical Laboratory， Zhejiang Provincial Tongde Hospital， Hangzhou   310012， China; 2.Special Service Center of the Air Force in Hangzhou， Hangzhou   310013， China; 3.Department of Dermatology， Zhejiang Provincial Tongde Hospital， Hangzhou   310012， China

[Abstract] Objective To study the mechanism of DC maturation induced by hydrazinocurcumin in inhibiting mouse skin squamous cell carcinoma. Methods SCC cell line SCL-1 was treated with hydrazinocurcumin at gradient concentration（1/5/10/50/100/500 μM） for 48 h， and cell activity was determined by MTT. At the same time， the spontaneous SCC mouse model was constructed， the tumor progression was observed after drug treatment. The dendritic maturation state of tumor tissues was detected by flow. The changes of STAT3 pathway in dendritic cells before and after treatment with hydrazinocurcumin were observed by Western blot. Results Different concentrations of hydrazinocurcumin may inhibit the activity of cells in the treatment of SCC cell line SCL-1， and the difference was significant（P<0.05）. The median maximum inhibitory concentration（IC50） was 18.7 μM. After normal C57BL/6 mice were given gastrointestinal administration of the drug for two weeks， their weight did not change significantly when compared with the control group. After treatment with hydrazinocurcumin， the antitumor effect of spontaneous mouse model was more prominent， and the difference was significant（P<0.05）. Flow cytometry detection of tumor tissues in SCC mice showed that treatment with hydrazinocurcumin could promote dendritic cell maturation（CD80/CD86 was significantly up-regulated）， and the difference was statistically significant（P<0.05）. Further Western blot analysis of the STAT3 pathway in dendritic cells before and after treatment with hydrazinocurcumin showed that the STAT3 phosphorylation level in the treatment group was significantly down-regulated（P<0.05）. Conclusion Hydrazinocurcumin can kill SCC cells in vitro and promote the maturation of dendritic cells in vivo， thus inhibiting the progression of SCC. The possible mechanism is to inhibit the STAT3 pathway by inhibiting the STAT3 phosphorylation.

[Key words] Hydrazinocurcumin; Cutaneous squamous cell carcinoma; Proliferation; Dendritic cell; STAT3

姜黄的活性成分为姜黄素，其为一种被广泛应用于疾病治疗的植物化合物[1，2]。联氨基姜黄素（Hydrazinocurcumin）的药理性更为显著，因此，可大大提升生物的利用程度[3]。前期研究证明HC是STAT3磷酸化的有效抑制剂。它可以下调一系列STAT3下游靶标，从而在体外抑制细胞增殖，集落形成丧失，抑制细胞迁移和侵袭以及诱导细胞凋亡[4-6]。然而HC对于肿瘤免疫微环境的调节少有讨论。本研究探讨HC对于皮肤鳞状细胞癌（Skin squamous cell carcinoma，SCC）小鼠模型成瘤的抑制作用，观察其对肿瘤生长以及肿瘤组织中树突状细胞（Dendritic cell，DC）成熟水平影响，并进一步研究其分子机制。

1 材料与方法

1.1 细胞株和试剂

DMEM培养基、双抗、胎牛血清购自 Gbico（美国），流式抗体 Zombie、PE-CD80、 APC-CD86、FITC-CD11 c购自Becton Dickinson（美国），二抗购自Invitrogen（美国），一抗（anti-STAT3、anti-p-STAT3）、增强型发光试剂盒购自Santa Cruz Biotechnology（美国），内参GAPDH购自 Abclonal（中国），PVDF膜购自Millipore（美国），rmGM-CSF、IL-4、 DMBA、TPA、丙酮溶液均购自Sigma（美国），DMSO、MTT试剂盒从武汉谷歌生物公司购入。酶联免疫检测仪为美国Thermo公司产品（IED0501003M），流式细胞仪为美国Becton Dickinson公司产品（LSRFortessa）。……