Moyer Hunter R, Kinney Ramsey C, Singh Kimberly A, Williams Joseph K, Schwartz Zvi, Boyan Barbara D
Division of Plastic and Reconstructive Surgery, Emory University, Atlanta, GA 30322, USA.
Ann Plast Surg. 2010 Nov;65(5):497-503. doi: 10.1097/SAP.0b013e3181d37713.
Autologous fat is the ideal soft-tissue filler; however, its widespread application is limited because of variable clinical results and poor survival. Engineered fillers have the potential to maximize survival. Alginate is a hydrogel copolymer that can be engineered into spheres of <200 μm, thus facilitating mass transfer, allowing for subcutaneous injection, and protecting cells from shearing forces.
Alginate powder was dissolved in saline, and adipose-derived stem cells (ADSCs) were encapsulated (1 million cells/mL) in alginate using an electrostatic bead generator. To assess effects of injection on cell viability, microspheres containing ADSCs were separated into 2 groups: the control group was decanted into culture wells and the injection group was mixed with basal media and injected through a 21-gauge needle into culture wells. Microbeads were cultured for 3 weeks, and cell number and viability were measured weekly using electron and confocal microscopy. To assess effects of percutaneous injection in vivo, twenty-four male nude mice were randomly separated into 2 groups and injected with either empty microcapsules or ADSC-laden microcapsules. Mice were harvested at 1 and 3 months, and the implants were examined microscopically to assess bead and cell viability.
A flow rate of 5 mL/h and an electrostatic potential of 7 kV produced viable ADSC-laden microbeads of <200 μm. There were no differences in bead morphology and ADSC viability between microcapsules placed versus injected into tissue culture plates for up to 3 weeks. Microspheres implanted in a nude mouse model show durability up to 3 months with a host response around each individual sphere. ADSCs remained viable and showed signs of mitosis.
ADSCs can be readily cultured, encapsulated, and injected in alginate microspheres. Stem cells suspended in alginate microspheres survive in vivo and are seen to replicate in vitro.
自体脂肪是理想的软组织填充剂；然而，由于临床效果不一且存活率低，其广泛应用受到限制。工程填充剂有提高存活率的潜力。藻酸盐是一种水凝胶共聚物，可加工成直径小于200μm的球体，从而促进物质传递，便于皮下注射，并保护细胞免受剪切力影响。
将藻酸盐粉末溶解于盐水中，使用静电微珠发生器将脂肪来源干细胞（ADSCs）包裹于藻酸盐中（100万个细胞/毫升）。为评估注射对细胞活力的影响，将含有ADSCs的微球分为两组：对照组倾入培养孔，注射组与基础培养基混合后通过21号针头注入培养孔。将微珠培养3周，每周使用电子显微镜和共聚焦显微镜测量细胞数量和活力。为评估体内经皮注射的效果，将24只雄性裸鼠随机分为两组，分别注射空微胶囊或载有ADSCs的微胶囊。在1个月和3个月时处死小鼠，对植入物进行显微镜检查以评估微珠和细胞活力。
流速为5毫升/小时、静电势为7千伏时可产生直径小于200μm且有活力的载有ADSCs的微珠。置于组织培养板与注入组织培养板的微胶囊在长达3周的时间里，微珠形态和ADSCs活力并无差异。植入裸鼠模型的微球在长达3个月的时间里保持完好，每个微球周围有宿主反应。ADSCs保持活力并显示有丝分裂迹象。
ADSCs可轻松培养、包裹并注射到藻酸盐微球中。悬浮于藻酸盐微球中的干细胞在体内存活，并在体外可见其复制。
