基于介孔纳米碳球接枝聚胍类化合物协同光热抗菌策略的研究

doi:10.6023/A21120548

摘要/Abstract

细菌感染是严重威胁着人们生命健康的主要问题之一. 与传统抗生素疗法相比, 新型抗菌策略光热疗法(PTT)展现出可控、微创及不易使细菌产生耐药性的优良性能. 然而单模形式下PTT疗法并不理想, 且高温下通常伴随着炎性反应等副作用, 联合抗菌策略可有效解决这一问题. 本研究制备了富含羧基的氧化介孔纳米碳球(OMCN), 通过酰胺化反应, 共价接枝聚六亚甲基双胍(PHMB), 得到OMCN-PHMB纳米平台. 实验结果表明, OMCN的光热性能具有良好的浓度与近红外光照功率依赖性. 在808 nm激光照射下, OMCN-PHMB的体内外联合光热治疗效果显著优于单一模式下的其它治疗组, 且组织学分析结果表明, 该纳米平台对小鼠的主要器官没有产生明显的毒性, 具有较高的生物相容性.

关键词: 细菌感染, 光热疗法(PTT), 聚六亚甲基双胍(PHMB), 联合治疗, 抗菌

Bacterial infection is one of the major problems that seriously threaten people's life and health. In recent years, photothermal therapy (PTT), which uses photothermal conversion nanomaterials to convert optical energy into thermal energy for sterilization under specific wavelength laser irradiation, has aroused wide interest of researchers. Compared with traditional antibiotic therapy, the new anti-bacterial strategy photothermal therapy shows the excellent performance of controllable, minimally invasive and less bacterial resistance. However, monomodal PTT therapy is not ideal because it is often associated with side effects such as inflammatory reaction. Therefore, it is necessary to develop novel photothermal antibacterial system with high biocompatibility and safety to fight bacterial infection. Combined antibacterial strategy can effectively solve this problem. In this work, mesoporous carbon nanospheres (MCN) were prepared and oxidized by the mixed acid to obtain carboxyl-rich oxidized mesoporous carbon nanospheres (OMCN) with high biocompatibility and photothermal properties. Then, OMCN- PHMB nano-antibacterial platform was obtained by grafting the antimicrobial agent poly(hexamethylene biguanide)hydrochlo- ride (PHMB) onto the surface of OMCN with amide covalently. The photothermal properties of the system were evaluated and the results showed that the photothermal performance of OMCN had a good dependence on the concentration and power density of near-infrared light. Similar to the OMCN, the obtained OMCN-PHMB exhibited excellent performance of photothermal controllability and photothermal stability. In vivo and in vitro antibacterial experiments showed that the therapeutic effect of OMCN-PHMB under 808 nm laser irradiation was significantly better than that of other treatment groups under the single mode, which confirmed the excellent antibacterial effect of OMCN-PHMB combined with photothermal therapy. In addition, histological analysis showed that the nanoplatform had no significant toxicity to the major organs of mice, indicating OMCN-PHMB had a high biocompatibility. To sum up, the photothermal synergistic nano-antibacterial platform constructed in this study is expected to serve as a safe and controllable biomedical platform to combat various diseases caused by bacterial infection, providing a new antibacterial strategy for clinical treatment of bacterial infection diseases.

Key words: bacterial infection, photothermal therapy (PTT), poly(hexamethylene biguanide)hydrochloride (PHMB), combination therapy, antibacterial

引用此文

张审, 冯闪, 马陇豫, 杨莹莹, 刘超群, 宋宁宁, 杨彦伟. 基于介孔纳米碳球接枝聚胍类化合物协同光热抗菌策略的研究[J]. 化学学报, 2022, 80(3): 265-271.

Shen Zhang, Shan Feng, Longyu Ma, Yingying Yang, Chaoqun Liu, Ningning Song, Yanwei Yang. Research of Synergistic Photothermal Antibacterial Strategy Based on Polymeric Guanidine Derivative Grafted on Mesoporous Carbon Nanospheres[J]. Acta Chimica Sinica, 2022, 80(3): 265-271.

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图/表 7

图式1. OMCN-PHMB制备工艺示意图及其在协同光热抗菌治疗中的应用

Scheme 1. Schematic illustration of preparation process of OMCN- PHMB and its application in synergistic photothermal antibacterial therapy

图1. (a) MCN和(b) OMCN的SEM图像、(c) MCN和(d) CMCN的XRD图谱、(e) MCN、OMCN和OMCN-PHMB的FTIR图谱、(f) MCN和OMCN的粒径大小分布图以及(g) MCN、OMCN和OMCN-PHMB的Zeta电位分布图

Figure 1. SEM images of (a) MCN and (b) OMCN, XRD patterns of (c) MCN and (d) OMCN, (e) FTIR spectra of MCN, OMCN and OMCN-PHMB, (f) size distribution of MCN and OMCN, and (g) Zeta potential of MCN, OMCN and OMCN-PHMB

图2. (a) MCN和OMCN的紫外-可见-近红外吸收光谱(插图为对应的水分散性图片)、(b)不同浓度OMCN在808 nm激光(1.5 W/cm2)照射下温度的变化和(c)相应的热红外图、(d) OMCN (55 µg/mL)在不同功率强度808 nm激光照射下温度的变化和(e)相应的热红外图以及(f) OMCN (55 µg/mL)经过五次循环激光照射(1.5 W/cm2)的光热稳定性

Figure 2. (a) UV-Vis-NIR spectrum of MCN and OMCN (Inset: photographs of MCN and OMCN dispersed in deionized water), (b) temperature changes of OMCN at different concentrations illustrated with an 808 nm laser (1.5 W/cm2) and (c) corresponding infrared thermal images, (d) temperature changes of OMCN (55 µg/mL) after irradiated by an 808 nm laser with different power intensities and (e) corresponding infrared thermal images, and (f) recycling-heating profiles of OMCN dispersion (55 µg/mL) irradiated for five on/off cycles (1.5 W/cm2)

图3. (a) PBS (pH 7.4)、OMCN (55 μg/mL)、PHMB (2.86 μg/mL)和OMCN-PHMB (55 μg/mL)在有无近红外激光(808 nm, 1.5 W/cm2, 10 min)照射处理后的金黄色葡萄球菌在LB琼脂平板上的菌落照片、(b)有无激光照射下(808 nm, 1.5 W/cm2, 10 min)的金黄色葡萄球菌存活率[通过测量600 nm处的光密度值确定; 误差条表示标准差(n=3), 星号表示有统计学意义的差异($\overline{x}$±s, *P<0.05, **P<0.01, ***P<0.001]以及(c) PBS (pH 7.4)、OMCN (55 μg/mL)、PHMB (2.86 μg/mL)和OMCN- PHMB (55 μg/mL)在有无近红外(808 nm, 1.5 W/cm2, 10 min)照射下处理金黄色葡萄球菌的活/死荧光染色结果

Figure 3. (a) Photographs of S. aureus colonies on an LB agar plate after treated with PBS (pH 7.4), OMCN (55 µg/mL), PHMB (2.86 μg/mL) and OMCN-PHMB (55 µg/mL) with or without NIR irradiation (808 nm, 1.5 W/cm2, 10 min), (b) S. aureus urvival ratio for the different groups with or without laser irradiation (808 nm, 1.5 W/cm2, 10 min) [determined by measuring the optical density at 600 nm; Error bars represent the standard deviation (n=3). Asterisks indicate statistically significant differences ($\overline{x}$±s, *P<0.05, **P<0.01, ***P<0.001)], and (c) LIVE/DEAD fluorescent staining results of S. aureus after treated with PBS (pH 7.4), OMCN (55 µg/mL), PHMB (2.86 μg/mL) and OMCN-PHMB (55 µg/mL) with or without NIR irradiation (808 nm, 1.5 W/cm2, 10 min)

图4. HL-7702细胞分别与不同浓度OMCN和OMCN-PHMB孵育后的细胞活力[误差条表示标准差(n=3)]

Figure 4. Cell viability of HL-7702 cells after incubation with different concentration of OMCN and OMCN-PHMB, respectively [Error bars represent the standard deviation (n=3)]

图5. (a)小鼠感染创面的形成及后续处理的示意图、(b)不同组治疗1、3、5 d后的伤口感染照片、(c)第5 d从小鼠创面分离细菌(置于LB琼脂平板培养)、(d)分别于第5 d对各组创面组织切片进行H&E染色、(e)不同治疗组组织处菌落的定量分析、(f)第5 d不同治疗组创口相对面积的定量分析以及(g)不同组治疗1、3、5 d后的小鼠体重变化情况[误差条表示标准差(n=3), 星号表示有统计学意义的差异($\overline{x}$±s, *P<0.05, **P<0.01, ***P<0.001)]

Figure 5. (a) Schematic diagram of infected wound formation in mice and the following treatments, (b) photographs of infected wounds after different treatments for 1, 3 and 5 d, (c) bacteria isolated from the mice wounds on day 5 and cultured on LB agar plates, (d) H&E-stained tissue slices from the wound areas of different groups on day 5, (e) quantitative analysis of bacterial colonies from tissues of different treatment groups, (f) quantitative analysis of relative wound area of different treatment groups on day 5, and (g) body weight variation among infected mice after different treatments for 1, 3 and 5 d [Error bars represent the standard deviation (n=3). Asterisks indicate statistically significant differences ($\overline{x}$ ±s, *P<0.05, **P<0.01, ***P<0.001)]

图6. 对各组小鼠主要脏器(心、肝、脾、肾和肺)H&E染色进行组织学分析

Figure 6. Histology analysis of major organs (heart, liver, spleen, kidney and lung) in different groups by H&E staining

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