【Biomaterial】天大姬晓元团队、烟台大学张竹红团队联手|双模式自供电敷料，实现190 mV微电压与98%愈合率，高效解决糖尿病感染创面修复#

Fig. 2. Preparation and Characterization of PTG/TEG/AST Devices. (A) 3D Printing Schematic Diagram. (B) Photograph of PTG/TEG/AST Device Based on 3D Printing. (C) PTG; (D) PVA/AST Hydrogel; (E) SEM Images of Pure PEDOT: PSS and (F) Bi2Te3 Doped TEG. (G) The viscosity of the PVA/AST hydrogel-shear strain (H) The storage modulus (G′) and loss modulus (G″). (I) Young’s Modulus and (J) Ultimate Stress of Different Parts in PTG/TEG/AST.#

Fig. 3. Thermoelectric performance characterization of PTG/TEG/AST Devices. (A) Schematic diagram of optical-to-thermal conversion and (B) thermoelectric conversion. (C) Image of PTG’s thermal rise under light. (D) Thermoelectric voltage generated by TEG device at different temperatures. (E) Actual temperature difference image recorded by infrared thermal imager of TEG device at different temperatures. (F) Thermal rise curve of PTG under light. (G) Variation of thermoelectric voltage with temperature. (H) Calculation of Seebeck coefficient by linear fitting of ΔV and ΔT curves.#

器件的物理微观结构与力学性能展现出优异的生物适配性。如图2所示，通过电纺与3D打印技术，PTG呈现出均匀的网状孔隙结构，TEG在基底上实现了P-N型热电电极阵列的精准排列，而底层的PVA水凝胶表现出良好的水合状态与粘附力。流变学与力学测试表明，水凝胶具有典型的剪切稀化特性与强附着力，各组分集成后展现出良好的拉伸强度与柔韧性，能够无缝贴合不规则的创面几何形状，并在使用过程中保持结构完整性。

光热转化与温电发电性能在体外测试中得到了系统验证。如图3所示，在808 nm近红外光照射5分钟内，PTG表面温度迅速提升至45°C。温度梯度的建立显著激活了TEG，随着热端温度升高，输出电压由50 mV提升至225 mV，在模拟生理条件下可稳定输出190 mV电压，线性拟合计算出的塞贝克系数高达11.56 mV K⁻¹。同时，45°C的热效应减弱了PVA水凝胶的物理交叉氢键网络，使得虾青素的累积释放率由25°C时的25%大幅提升至90%，实现了受控按需给药。

Fig. 4. In vitro antibacterial performance characterization. (A) The killing effects of photothermal effect and thermoelectric effect on bacteria. (B) Images of S. aureus and E. coli grown on agar plates after different treatments. (C) Images of inhibition rings and quantitative determination of inhibition ring areas for S. aureus and E. coli after different treatments. (D) The absorbance of bacterial solutions measured at different times after different treatments for S. aureus and (F) E. coli. (E) Survival rate curves of S. aureus and (G) E. coli after different treatments. (H) SEM images of S. aureus and E. coli after different treatments. (I) S. aureus; (J) E. coli. Each experiment was independently repeated three times with similar results.#

体外抗菌实验展示了光热与电刺激协同作用的强大威力。如图4所示，单独的PTG光热作用或TEG电刺激作用对细菌的抑制相对有限，但在近红外光照射下，PTG与TEG结合产生的协同效应使金黄色葡萄球菌与大肠杆菌的存活率分别降低了97% 和98%。扫描电镜图显现出非常直观的物理破坏景象，受杀杀菌处理后的细菌细胞膜严重皱缩破裂，胞浆大量外漏，证实了电场与热能协同打孔的抑菌机制。

Fig. 5. The in vitro antioxidant and anti-inflammatory performance of PTG/TEG/AST device. (A) Using CLSM and (B) using FCM to detect the ROS clearance within cells after different treatments. (C) Analyzing the polarization ratio of macrophages after different treatments using FCM. (D) Quantitative analysis of the ratio of M1 and M2 macrophages in different treatment groups. (E) CLSM images of macrophages after different treatments. (F) (G) Quantitative analysis of the fluorescence intensity of M1 and M2 macrophages in different treatment groups. (H) Schematic diagram of the anti-inflammatory effect of PTG/TEG/AST. Each experiment was repeated independently three times with similar results.#

在细胞水平的抗氧化与巨噬细胞表型调控方面，敷料展现出卓越的微环境重塑能力。如图5所示，水凝胶释放的虾青素能够清除细胞内约90%的过量ROS。流式细胞术与免疫荧光观察表明，在氧化应激条件下，PTG/TEG/AST联合处理能够显著下调表达CD86的促炎M1型巨噬细胞比例，同时大幅上调表达CD206的抗炎M2型巨噬细胞比例，RT-qPCR结果进一步证实促炎因子iNOS被抑制而促修复因子TGF-β显著增加，证明该系统成功将创面免疫环境由促炎转变为促进再生的状态。

Fig. 6. PTG/TEG/AST device promote the in vitro proliferation ability of cells. (A) The CCK8 assay measures the cell viability of L929 cells and (B) HUVEC cells on the 1st, 3rd, and 5th days after different treatments. (C) the cell migration images and (H) quantitative analysis of L929 cells after different treatments. (D) the angiogenesis images and (I) quantitative analysis of HUVEC cells after different treatments. (E) CLSM images of F-actin in L929 and HUVEC cells. (F) Representative images and (J) quantification of VEGFA in HUVEC. (G) CLSM images of the cell viability after different treatment, with red representing dead cells and green representing live cells. (K) Observe the cell apoptosis after different treatments through FCM. Each experiment was repeated independently three times with similar results. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)#

电刺激与抗氧化的双重作用显著加速了修复相关细胞的生物学行为。如图6所示，在H2O2诱导的氧化应激损伤模型中，PTG/TEG/AST敷料不仅保护了L929成纤维细胞与HUVEC内皮细胞免于凋亡，还显著增强了成纤维细胞的划痕迁移速率。在Matrigel成管实验中，受电刺激激活的内皮细胞形成了极为密集的血管样管状网络，细胞内血管内皮生长因子A（VEGFA）的表达量大幅增加，荧光染色观察到细胞骨架F-actin形成明显的伪足延伸，表明微电场有效引导了细胞的定向迁移与血管化进程。

Fig. 7. PTG/TEG/AST device promotes healing of infected diabetic wounds. (A) Schematic diagram of PTG/TEG/AST for treating diabetic infected mice. (B) Temperature rise images and thermoelectric voltage generation of PTG/TEG/AST in vivo. (C) Representative wound photos of each treatment group on different days. (D) Wound closure at different time points. (E) Quantitative analysis of wound area. (F) HE staining and (G) MASSON staining of wounds on the 3rd and 9th days. Each experiment was repeated independently three times with similar results.#

为了验证临床应用潜力，研究人员建立了小鼠糖尿病金黄色葡萄球菌感染全厚皮损模型。如图7所示，敷料贴合于创面后在光照下升温至45°C，并在光照结束后依靠体表温差持续维持106 mV的输出电压。治疗至第9天时，PTG/TEG/AST组的创面愈合率达到了98%，远高于其他对照组。组织学切片（HE与Masson染色）显示，该组创面实现了完整的再上皮化，再生表皮层厚度适宜，真皮层中胶原蛋白沉积丰富且呈平行有序排列，同时主要器官切片未见任何病理损伤，证实了良好的全身生物安全性。

Fig. 8. Immune analysis of the wound healing process in different treatment groups. (A) Representative fluorescence images of CD31/α-MSA staining, HIF-α staining, EGF staining, CD206/CD86 staining, IL-6 staining and IL-10 staining in the wound tissue on the 6th day after different treatments, and quantitative analysis (C) CD31; (D) α-MSA; (E) HIF-α; (F) EGF; (G) M2/M1 type macrophages (H) IL-6; (I) IL-10. (B) FCM analysis of the polarization of macrophages in the wound tissue and (J) quantitative analysis. Each experiment was repeated independently three times with similar results.#

创面组织的免疫荧光与流式细胞深入检测进一步揭示了组织再生的微观机制。如图8所示，PTG/TEG/AST治疗组创面组织中的CD31与α-SMA血管标记物表达最强，成熟新生血管密度显著增加。随着血流灌注的改善，创面缺氧诱导因子HIF-α的表达显著下降，说明缺氧应激得到有效缓解。此外，创面组织中表皮生长因子（EGF）与抗炎因子IL-10表达显著升高，促炎因子IL-6受到强烈抑制，巨噬细胞全面向M2表型转化，形成了高度协调的组织再生微环境。

Fig. 9. Single-cell RNA sequencing analysis. (A) Principal component analysis (PCA) of gene expression profiles among the Control, PTG/AST, and PTG/TEG treatment groups. (B) Co-expression Venn diagram of differentially expressed genes among the three groups. (C) Volcano plot showing differentially expressed genes between the Control and PTG/AST groups. (D) Volcano plot showing differentially expressed genes between the Control and PTG/TEG groups. (E) Schematic illustration of the signaling pathways involved in PTG/TEG-mediated wound healing and anti-inflammatory effects. (F) GO and (G) KEGG enrichment analysis of differentially expressed genes between the Control and PTG/AST groups. (H) GO and (I) KEGG enrichment analysis of differentially expressed genes between the Control and PTG/TEG groups. Each experiment was independently repeated three times with consistent results.#

最后，转录组学与单细胞RNA测序从基因层面阐明了作用通路。如图9所示，差异基因富集分析表明，PTG/TEG产生的电刺激能够改变细胞膜电位，打开电压门控钙通道，促进钙离子内流并激活钙调蛋白依赖性信号途径，从而调控细胞代谢、骨架重组与血管生成；而虾青素则主要通过激活PI3K/Akt信号通路，抑制NF-κB炎症途径，协同实现促炎基因下调与组织修复基因上调，两者从物理电信号与化学药物两条路径实现了对糖尿病创面修复过程的精准分子干预。