可注射透明质酸/羟基磷灰石复合材料: 制备、理化性能和细胞相容性

doi:10.15541/jim20210020

无机材料学报 ›› 2021, Vol. 36 ›› Issue (9): 981-990.DOI: 10.15541/jim20210020

所属专题：【虚拟专辑】抗菌材料(2020~2021)

可注射透明质酸/羟基磷灰石复合材料: 制备、理化性能和细胞相容性

朱雨桐(), 谭佩洁, 林海(), 朱向东, 张兴栋

四川大学国家生物医学材料工程技术研究中心, 成都 610064

收稿日期:2021-01-11 修回日期:2021-01-22 出版日期:2021-09-20 网络出版日期:2021-03-01
通讯作者: 林海, 副研究员. E-mail: linhai028@scu.edu.cn
作者简介:朱雨桐(1997-), 女, 硕士. E-mail: 876073900@qq.com
基金资助:
国家自然科学基金面上项目(32071330)

Injectable Hyaluronan/Hydroxyapatite Composite: Preparation, Physicochemical Property and Biocompatibility

ZHU Yutong(), TAN Peijie, LIN Hai(), ZHU Xiangdong, ZHANG Xingdong

National Engineering Research Center for Biomaterials, Sichuan University, Chengdu 610064, China

Received:2021-01-11 Revised:2021-01-22 Published:2021-09-20 Online:2021-03-01
Contact: LIN Hai, associate professor. E-mail: linhai028@scu.edu.cn
About author:ZHU Yutong (1997-), female, master. E-mail: 876073900@qq.com
Supported by:
National Natural Science Foundation of China(32071330)

摘要/Abstract

摘要：

羟基磷灰石(HAP)具有优异的生物相容性、生物活性和缓慢的降解速率等特性, 有可能弥补透明质酸(HA)用作软组织填充材料的不足。本研究采用一步反应法制备均匀负载HAP颗粒的HA-HAP复合水凝胶, 分析了制备工艺参数对复合水凝胶理化性能的影响及调控方法, 评价了其细胞相容性。结果表明, 采用该方法制备的复合水凝胶含1.5%~3.0%HA、2.38%~15.8% HAP, 具有良好的灭菌稳定性。通过改变HAP含量和交联剂的用量可以调控复合水凝胶的力学性能、溶胀行为等理化性能。利用一阶指数衰减方程可以较好地拟合复合水凝胶的溶胀行为, 质量溶胀率在1 h时已达到平衡溶胀率的90%以上。复合水凝胶具有良好的HA酶降解性, 1500 U/mL的酶作用2 min后HA降解率超过95%, 复合水凝胶可以快速溃散; 加入HAP颗粒不仅使复合水凝胶有更好的可塑性, 而且更有利于面部软组织填充和轮廓修正。优化制备的可注射HA-HAP复合水凝胶具有可调的理化性能和良好的细胞相容性, 可以较长时间地保留在体内, 兼具促进组织修复或重建的潜力, 有望成为优良的软组织填充材料。

关键词: 可注射材料, 透明质酸, 羟基磷灰石, 理化性能, 细胞相容性

Abstract:

With good cytocompatibility, bioactivity, and slow degradation rate, hydroxyapatite (HAP) may remedy drawbacks of hyaluronan (HA) as soft tissue filler. Here, we prepared HA-HAP composite hydrogels by one-step reaction method, investigated various processing parameters on their physicochemical properties, and evaluated their biological performance. As-prepared HA-HAP composite hydrogels contained 1.5%-3.0% HA and 2.38%-15.8% HAP with good sterilization stability. Swelling-ratio and mechanical performance of the composite hydrogels could be adjusted by controlling HAP content and crosslinking reagent. The equilibrium swelling ratio achieved over 90% only after 1 h swell, and the swelling properties fitted well with 1^st-order exponential decay equations. By 1500 U/mL HAase solutions, the composites degraded 95% after only 2 min and dispersed rapidly, showing good enzymatic degradability. HAP was demonstrated not only endowing the composite hydrogels with better plasticity for facial soft tissue filler and contour modification, but also providing good microenvironment for attachment and proliferation of fibroblasts. Therefore, the as-prepared HA-HAP composite could be used as a long-acting filler for remodeling extracellular matrix, and thus potentially applied in cosmetic medicine.

Key words: injectable composite, hyaluronan, hydroxyapatite, physicochemical property, cytocompatibility

中图分类号:

TQ174

朱雨桐, 谭佩洁, 林海, 朱向东, 张兴栋. 可注射透明质酸/羟基磷灰石复合材料: 制备、理化性能和细胞相容性[J]. 无机材料学报, 2021, 36(9): 981-990.

ZHU Yutong, TAN Peijie, LIN Hai, ZHU Xiangdong, ZHANG Xingdong. Injectable Hyaluronan/Hydroxyapatite Composite: Preparation, Physicochemical Property and Biocompatibility[J]. Journal of Inorganic Materials, 2021, 36(9): 981-990.

图/表 13

图1 HA与BDDE的反应示意图

Fig. 1 Diagram of reaction between HA and BDDE

表1 各复合水凝胶样品制备时HA、HAP与BDDE的用量

Table 1 Amount of HA, HAP and BDDE in preparing composite hydrogels

Sample	HA/wt%	HAP/wt%	BDDE equivalent
HA-1.0	10	0	1.0
HAP15-0.5	10	15	0.5
HAP30-0.5	10	30	0.5
HAP30-1.0	10	30	1.0
HAP45-1.0	10	45	1.0

表2 复合水凝胶透析前后的HA与HAP含量

Table 2 Contents of HA and HAP in different composites before and after dialysis

Sample	Before dialysis		After dialysis
Sample	HA/%	HAP/%	HA/%	HAP/%
HA-1.0	10	0	2.01	-
HAP15-0.5	10	15	1.58	2.38
HAP30-0.5	10	30	1.54	4.63
HAP30-1.0	10	30	2.19	7.73
HAP45-1.0	10	45	3.00	15.8

图2 复合水凝胶灭菌前后的形态

Fig. 2 Appearance of composite hydrogels before and after autoclaving (A) Fully dialyzed; (B) Not dialyzed. From left to right: HA-1.0, HAP15-0.5, HAP30-0.5, HAP30-1.0, and HAP45-1.0

图3 含残留交联剂的复合水凝胶HAP45-1.0灭菌后的形态

Fig. 3 Appearance of composite hydrogels HAP45-1.0 with remained crosslinking reagents From left to right: fully dialyzed, alkali-contained, cross linker-contained

图4 水凝胶材料灭菌前后储能模量(a)、损耗模量(b)及复合黏度(c)的变化

Fig. 4 Storage modulus (a), loss modulus (b) and composite viscosity (c) of hydrogels before and after sterilization

图5 水凝胶的扫描电镜照片

Fig. 5 SEM morphologies of hydrogels

图6 水凝胶的推挤力-行程变化曲线

Fig. 6 Push-out strain-stress curves of hydrogels

图7 复合水凝胶HAP45-1.0的溶胀行为及稳定性

Fig. 7 Swelling behavior of composite hydrogel HAP45-1.0 and stabilization in vitro (A) Mass and volume swelling rate; (B) Sizes of samples before and after swelling for 8 h; (C) Stability of the samples immersed in PBS for different periods

表3 复合水凝胶HAP45-1.0溶胀率的一阶指数衰减拟合参数

Table 3 The 1st order exponential decay equation fitting results of hydrogel HAP45-1.0 swelling behavior

	A	y₀	R²
Mass swelling rate	-51.64	51.64	0.9992
Volume swelling rate	-78.81	78.66	0.9984

图8 在120 U/mL的HAase作用下, 复合水凝胶HAP45-1.0的体外酶降解

Fig. 8 Enzymatic degradation of composite hydrogel HAP45-1.0 in vitro by HAase at 120 U/mL

图9 成纤维细胞在水凝胶材料表面的粘附观察

Fig. 9 Fibroblast adhesion after being seeded on the surface of hydrogels observed by confocal fluorescence microscope

图10 成纤维细胞在水凝胶表面的增殖 (*p<0.05)

Fig. 10 Proliferation of fibroblasts on surface of hydrogels (*p<0.05)

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可注射透明质酸/羟基磷灰石复合材料: 制备、理化性能和细胞相容性

Injectable Hyaluronan/Hydroxyapatite Composite: Preparation, Physicochemical Property and Biocompatibility

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