壳聚糖修饰海藻酸钙微球构建肿瘤-内皮细胞共培养模型

Chitosan-Modified Calcium Alginate Microspheres for Constructing Tumor-Endothelial Cell Co-culture Models

摘要

To establish a tumor-endothelial cell co-culture model with a spatially partitioned structure, this study employed a chitosan electrostatic adsorption coating strategy to surface-modify calcium alginate microspheres. Breast tumor cells (MCF-7) were encapsulated within the microspheres, while human umbilical vein endothelial cells (HUVECs) were seeded on the microsphere surface, mimicking the spatial relationship between tumor cell clusters and vascular endothelium in solid tumors. By systematically investigating the effects of varying chitosan mass concentrations (1, 3, 5, 7 g/L) on microsphere particle size, surface charge, dimensional stability, and HUVECs adhesion, the optimal coating conditions were identified, and the proliferation behavior of tumor cells in the co-culture model was evaluated. Results indicated that microspheres modified with 5 g/L chitosan exhibited optimal particle size stability, a surface zeta potential of (9.00±0.57) mV, the highest HUVECs adhesion density with good spreading, and no significant cytotoxicity toward internal MCF-7 cells. In the co-culture model constructed using this optimized coating, HUVECs formed a continuous cell layer on the microsphere surface, connecting dispersed microspheres into tissue-like aggregates. MCF-7 cell activity in the co-culture group was enhanced compared to the monoculture group (reaching 115% on day 7). This model showed potential applications in studying tumor-endothelial cell interaction mechanisms, drug evaluation, and personalized medicine, providing a novelin vitro platform for simulating the tumor microenvironment.

Key words

chitosan / calcium alginate microsphere / tumor-endothelial cell co-culture / compartmental model / surface modification

参考文献

[1] HAN Y, ZHU B, MENG S. Endothelial cell in tumor angiogenesis: Origins, mechanisms, and therapeutic implication[J].Genes & Diseases, 2025, 12(6):101611.

[2] KANE K, EDWARDS D, CHEN J. The influence of endothelial metabolic reprogramming on the tumor microenvironment[J].Oncogene, 2025, 44(2):51-63.

[3] BAHCECEIOGLU G, BASARA G, ELLIS B W, et al. Breast cancer models: Engineering the tumor microenvironment[J].Acta Biomaterialia, 2020, 106:1-21.

[4] WU K Z, ZENG B, SEAH D H, et al. Advances in in-vitro modeling of cancer-stromal interactions: From fundamental insights to translational applications[J].Advanced Materials, 2025, 37(50):e12252.

[5] XIE Y, YANG X, PAN R, et al. Co-culture of natural killer cells and tumor spheroids on a heterogeneous multilayer paper stack[J].Journal of Zhejiang University. Science. B, 2024, 25(12):1097-1107.

[6] FRANCO M, ROSWALL P, CORTEZ E, et al. Pericytes promote endothelial cell survival through induction of autocrine VEGF-A signaling and Bcl-w expression[J].Blood, 2011, 118(10):2906-2917.

[7] RO J, KIM J, PARK J, et al. ODSEI chip: An open 3D microfluidic platform for studying tumor spheroid-endothelial interactions[J].Advanced Science, 2025, 12(13):2410659.

[8] YANG M, HONG M, WANG G, et al. Preparation of 3D zonal and interactional glomerular models based on composite core-shell hydrogel microspheres[J].ACS Materials Letters, 2024, 6(11):5154-5162.

[9] [ 9 ] NING L, SHIM J, TOMOV M L, et al. A 3D bioprinted in vitro model of neuroblastoma recapitulates dynamic tumor-endothelial cell interactions contributing to solid tumor aggressive behavior[J].Advanced Science, 2022, 9(23):2200244.

[10] LEE K Y, MOONEY D J. Alginate: Properties and biomedical applications[J].Progress in Polymer Science, 2012, 37(1):106-126.

[11] YU Y, WANG X, ZHU Y, et al. Is polydopamine beneficial for cells on the modified surface?[J].Regenerative Biomaterials, 2022, 9:rbac078.

[12] 胡萍, 张双英, 李洁, 等. 壳聚糖/海藻酸钙/自组装多肽微凝胶纤维的制备及应用[J].高分子通报, 2020, 33(8):49-54.

[13] 贺庆, 敖强, 韩大庆, 等. 基于琼脂糖/壳聚糖共混凝胶模型的壳聚糖生物相容性的机理研究 [J].国际生物医学工程杂志, 2012, 35(2):65-69.

[14] PARISI L, TOFFOLI A, BIANCHI M G, et al. Functional fibronectin adsorption on aptamer-doped chitosan modulates cell morphology by integrin-mediated pathway[J].Materials, 2019, 12(5):812.

[15] GONG J, SCHUURMANS C, GENDEREN A, et al. Complexation-induced resolution enhancement of 3D-printed hydrogel constructs[J].Nature Communications, 2020, 11(1):1267.

[16] CIARLANTINI C, FRANCOLINI I, SILVESTRO I, et al. Design of bioactive and biomimetic scaffolds based on chitosan-alginate polyelectrolyte complexes for tissue engineering[J].Carbohydrate Polymers, 2024, 327:121684.

[17] CHEN K Y, ZENG S Y. Preparation and characterization of quaternized chitosan coated alginate microspheres for blue dextran delivery[J].Polymers, 2017, 9(6):210.

[18] STOLL S, CHODANOWSKI P. Polyelectrolyte adsorption on an oppositely charged spherical particle: Chain rigidity effects[J].Macromolecules, 2002, 35(25):9556-9562.

[19] OUYANG L, SHAIK R, XU R, et al.Mapping surface charge distribution of single-cell via charged nanoparticle[J].Cells, 2021, 10(6):1519.

[20] VARGAS F F, CAVIEDES P F, GRANT D S, et al. Electrophysiological characteristics of cultured human umbilical vein endothelial cells[J].Microvascular Research, 1994, 47(2):153-165.

[21] AMARAL I F, SOUSA S R, NEIVA I, et al. Kinetics and isotherm of fibronectin adsorption to three-dimensional porous chitosan scaffolds explored by¹²⁵I-radiolabelling[J].Biomatter, 2013, 3(2):e24791.

[22] DAMAGHI M, GILLIES R. Phenotypic changes of acid-adapted cancer cells push them toward aggressiveness in their evolution in the tumor microenvironment[J].Cell Cycle, 2017, 16(19):1739-1743.

[23] PENG X, JANICIGEVIC Z, LEMM S, et al. Shell engineering in soft alginate-based capsules for culturing liver spheroids[J].Biotechnology Journal, 2023, 18(6):2200365.

[24] GHIABI P, JIANG J, PASQUIER J, et al. Endothelial cells provide a notch-dependent pro-tumoral niche for enhancing breast cancer survival, stemness and pro-metastatic properties[J].Plos One, 2014, 9(11):e112424.

[25] CHIANG K H, CHANG W, LIU Y, et al. Quantification of oxygen tension variation in an in vitro tumor microenvironment model based on co-culture of tumor spheroid and endothelial cells in a microfluidic device[J].Sensors and Actuators Reports, 2025, 10:100396.

[26] KABIR A U, SUBRAMANIAN M, KWON Y, et al. Linking tumour angiogenesis and tumour immunity[J].Nature Reviews Immunology, 2026, 26(1):35-51.

[27] WANG J, PAN X, ZHANG J, et al. Active molecule-based theranostic agents for tumor vasculature normalization and antitumor efficacy[J].Chinese Chemical Letters, 2024, 35(8):109187.

基金资助

新疆医科大学附属肿瘤医院重点实验室开放项目(XJKLO-2025S006)

新疆维吾尔自治区科技厅科技援疆计划项目(2026E02074)