梯度低温保存脂肪颗粒作为皮下填充物的可行性
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摘要
背景:在不添加保护剂的情况下,通过适宜冻存方法长期低温保存的多余脂肪组织能否直接二次移植,并达到较好的填充效果,是一项值得研究的课题。目的:应用梯度低温保存的脂肪颗粒作为皮下填充物的移植效果,探讨其临床应用的可行性。方法:将人抽脂获取的脂肪颗粒在不添加保护剂的情况下,经梯度降温置于-80℃冰箱行冷冻保存,实验分组新鲜脂肪、冻存1个月组、冻存3个月组、冻存6个月组。8-10周龄SPF级裸鼠12只,体质量约20 g,由徐州医科大学实验动物中心提供。将各组脂肪颗粒同期植入裸鼠背部皮下,移植后第4,8周取出样本,比较各组移植物大体外观、质量、体积变化;通过细胞提取、MTT检测、苏木精-伊红及免疫组织化学染色,观察比较脂肪组织的成活情况。结果与结论:经梯度低温保存后的脂肪颗粒在移植取出后均有脂肪细胞存活,且胞间可见新生血管生成。其中冻存6个月的脂肪颗粒移植效果甚至优于冻存1、3个月的脂肪。说明通过适宜的冻存步骤,未添加冷冻保护剂的脂肪颗粒经梯度低温保存可安全移植于生物体,并达到较满意的填充效果。
BACKGROUND: In the case of not adding protective agent, whether excessive adipose tissue cryopreserved for long time can be used for the second transplantation and achieve good filling effect is worthy to be investigated.OBJECTIVE: To investigate the transplantation effect of fat particles cryopreserved for different time, and the feasibility of clinical application. METHODS: Without addition of protectant, fat particles were cryopreserved at a final temperature of -80℃. There were four groups: fresh fat, cryopreservation for 1, 3, and 6 months groups. Twelve 8-10-week-old SPF nude mice weighting 20 g provided by the Laboratory Animal Center of Xuzhou Medical University were included in this study. After cryopreserved for different time, fat particles were separately implanted into the back of nude mice. Tissue samples were removed at 4 and 8 weeks after transplantation. Appearance, weight, and volume of graft were compared among groups. The survival of fat tissue was determined by MTT assay, hematoxylin-eosin staining and immunohistochemical staining. RESULTS AND CONCLUSION: After transplantation of cryopreserved fat particles, there were surviving adipocytes, and new blood vessels were observed between cells. The transplantation effect of fat particles cryopreserved for 6 months was superior to that of fat particles cryopreserved for 1 and 3 months. Therefore, fat particles cryopreserved for different time without addition of protectant can be transplanted without safety concern, which can obtain satisfactory filling outcomes.
BACKGROUND: In the case of not adding protective agent, whether excessive adipose tissue cryopreserved for long time can be used for the second transplantation and achieve good filling effect is worthy to be investigated.OBJECTIVE: To investigate the transplantation effect of fat particles cryopreserved for different time, and the feasibility of clinical application. METHODS: Without addition of protectant, fat particles were cryopreserved at a final temperature of -80℃. There were four groups: fresh fat, cryopreservation for 1, 3, and 6 months groups. Twelve 8-10-week-old SPF nude mice weighting 20 g provided by the Laboratory Animal Center of Xuzhou Medical University were included in this study. After cryopreserved for different time, fat particles were separately implanted into the back of nude mice. Tissue samples were removed at 4 and 8 weeks after transplantation. Appearance, weight, and volume of graft were compared among groups. The survival of fat tissue was determined by MTT assay, hematoxylin-eosin staining and immunohistochemical staining. RESULTS AND CONCLUSION: After transplantation of cryopreserved fat particles, there were surviving adipocytes, and new blood vessels were observed between cells. The transplantation effect of fat particles cryopreserved for 6 months was superior to that of fat particles cryopreserved for 1 and 3 months. Therefore, fat particles cryopreserved for different time without addition of protectant can be transplanted without safety concern, which can obtain satisfactory filling outcomes.
引文
[1]Shu Z, Gao D, Pu LL. Update on cryopreservation of adipose tissue and adipose-derived stem cells. Clinics in plastic surgery.2015;42(2):209-218.
[2]张广宇,于冬梅,罗赛,等.不同低温条件保存的大鼠脂肪自体移植后成活率的探究[J].中国美容整形外科杂志,2016,27(3):181-185.
[3]Coleman SR. Structural fat grafts:the ideal filler?. Clinics in plastic surgery.2001;28(1):111.
[4]仇侃敏,吴蒙,梁耀蝉,等.不同冻存液保存的颗粒性脂肪组织活力及移植成活率分析[J].基础医学与临床, 2014,34(4):536-540.
[5]章立群,刘学,任英华,等.大鼠脂肪干细胞冷冻复苏后的生物学特性及成骨细胞分化潜能的研究[J].中国现代医学杂志, 2018(6):33-37.
[6]Moscatello DK, Dougherty M, Narins RS,et al. Cryopreservation of human fat for soft tissue augmentation:viability requires use of cryoprotectant and controlled freezing and storage. Dermatologic Surgery. 2005;31(s4):1506–1510.
[7]谢红炬,邓颖,李明,等.自体纯化冷冻微粒脂肪注射移植修复面部皮肤软组织老化性萎缩凹陷[J].中国组织工程研究,2012,16(5):779-782.
[8]逯岩,项艳,张心霞,等.体外冷冻存储脂肪颗粒在隆乳术中的应用[J].中国美容医学, 2012,21(15):1922-1923.
[9]MD KTMST.长期冷冻脂肪的移植[J].中国美容整形外科杂志,2001,12(6):332-334.
[10]陈静,刘冠兰.冷冻脂肪与新鲜脂肪注射移植的疗效观察[J].中国美容整形外科杂志, 2003,14(3):140-141.
[11]Laschke MW, Karschnia P, Scheuer C,et al. Effects of cryopreservation on adipose tissue-derived microvascular fragments. J Tissue Eng Regen Med. 2018;12(4):1020-1030.
[12]Zanata F, Bowles A, Frazier T, et al. Effect of Cryopreservation on Human Adipose Tissue and Isolated Stromal Vascular Fraction Cells:In Vitro and In Vivo Analyses. Plast Reconstr Surg. 2018;141(2):232e-243e.
[13]Conti G, Jurga M, Benati D,et al. Cryopreserved Subcutaneous Adipose Tissue for Fat Graft. Aesthetic Plastic Surgery. 2015;39(5):800-817.
[14]Hwang SM, Lee JS, Kim HD,et al. Comparison of the viability of cryopreserved fat tissue in accordance with the thawing temperature. Arch Plast Surg. 2015;42(2):143-149.
[15]Shoshani O, Ullmann Y, Shupak A, et al. The role of frozen storage in preserving adipose tissue obtained by suction-assisted lipectomy for repeated fat injection procedures. Dermatol Surg.2001;27(7):645-647.
[16]Zhao G, Liu X, Zhu K,et al. Hydrogel Encapsulation Facilitates Rapid-Cooling Cryopreservation of Stem Cell-Laden Core-Shell Microcapsules as Cell–Biomaterial Constructs. Adv Healthc Mater. 2017 Dec;6(23).
[17]Liu X, Zhao G, Chen Z, et al. Dual Suppression Effect of Magnetic Induction Heating and Microencapsulation on Ice Crystallization Enables Low-Cryoprotectant Vitrification of Stem Cell–Alginate Hydrogel Constructs. ACS Appl Mater Interfaces. 2018;10(19):16822-16835.
[18]刘磊,庸奇,李娜,等.脂肪干细胞-脱钙骨的玻璃化冻存及复苏[J].中国组织工程研究,2018,22(9):1357-63.
[19]Zhang Y, Cai J, Zhou T, et al. Improved Long-Term Volume Retention of Stromal Vascular Fraction Gel Grafting with Enhanced Angiogenesis and Adipogenesis. Plast Reconstr Surg.2018;141(5):676e-686e.
[2]张广宇,于冬梅,罗赛,等.不同低温条件保存的大鼠脂肪自体移植后成活率的探究[J].中国美容整形外科杂志,2016,27(3):181-185.
[3]Coleman SR. Structural fat grafts:the ideal filler?. Clinics in plastic surgery.2001;28(1):111.
[4]仇侃敏,吴蒙,梁耀蝉,等.不同冻存液保存的颗粒性脂肪组织活力及移植成活率分析[J].基础医学与临床, 2014,34(4):536-540.
[5]章立群,刘学,任英华,等.大鼠脂肪干细胞冷冻复苏后的生物学特性及成骨细胞分化潜能的研究[J].中国现代医学杂志, 2018(6):33-37.
[6]Moscatello DK, Dougherty M, Narins RS,et al. Cryopreservation of human fat for soft tissue augmentation:viability requires use of cryoprotectant and controlled freezing and storage. Dermatologic Surgery. 2005;31(s4):1506–1510.
[7]谢红炬,邓颖,李明,等.自体纯化冷冻微粒脂肪注射移植修复面部皮肤软组织老化性萎缩凹陷[J].中国组织工程研究,2012,16(5):779-782.
[8]逯岩,项艳,张心霞,等.体外冷冻存储脂肪颗粒在隆乳术中的应用[J].中国美容医学, 2012,21(15):1922-1923.
[9]MD KTMST.长期冷冻脂肪的移植[J].中国美容整形外科杂志,2001,12(6):332-334.
[10]陈静,刘冠兰.冷冻脂肪与新鲜脂肪注射移植的疗效观察[J].中国美容整形外科杂志, 2003,14(3):140-141.
[11]Laschke MW, Karschnia P, Scheuer C,et al. Effects of cryopreservation on adipose tissue-derived microvascular fragments. J Tissue Eng Regen Med. 2018;12(4):1020-1030.
[12]Zanata F, Bowles A, Frazier T, et al. Effect of Cryopreservation on Human Adipose Tissue and Isolated Stromal Vascular Fraction Cells:In Vitro and In Vivo Analyses. Plast Reconstr Surg. 2018;141(2):232e-243e.
[13]Conti G, Jurga M, Benati D,et al. Cryopreserved Subcutaneous Adipose Tissue for Fat Graft. Aesthetic Plastic Surgery. 2015;39(5):800-817.
[14]Hwang SM, Lee JS, Kim HD,et al. Comparison of the viability of cryopreserved fat tissue in accordance with the thawing temperature. Arch Plast Surg. 2015;42(2):143-149.
[15]Shoshani O, Ullmann Y, Shupak A, et al. The role of frozen storage in preserving adipose tissue obtained by suction-assisted lipectomy for repeated fat injection procedures. Dermatol Surg.2001;27(7):645-647.
[16]Zhao G, Liu X, Zhu K,et al. Hydrogel Encapsulation Facilitates Rapid-Cooling Cryopreservation of Stem Cell-Laden Core-Shell Microcapsules as Cell–Biomaterial Constructs. Adv Healthc Mater. 2017 Dec;6(23).
[17]Liu X, Zhao G, Chen Z, et al. Dual Suppression Effect of Magnetic Induction Heating and Microencapsulation on Ice Crystallization Enables Low-Cryoprotectant Vitrification of Stem Cell–Alginate Hydrogel Constructs. ACS Appl Mater Interfaces. 2018;10(19):16822-16835.
[18]刘磊,庸奇,李娜,等.脂肪干细胞-脱钙骨的玻璃化冻存及复苏[J].中国组织工程研究,2018,22(9):1357-63.
[19]Zhang Y, Cai J, Zhou T, et al. Improved Long-Term Volume Retention of Stromal Vascular Fraction Gel Grafting with Enhanced Angiogenesis and Adipogenesis. Plast Reconstr Surg.2018;141(5):676e-686e.