本文選題：3D生物打印 + 組織工程表皮模型　； 參考：《南方醫(yī)科大學》2017年碩士論文

【摘要】：背景:皮膚是機體最大的器官,因燒、創(chuàng)傷導致的皮膚大面積損傷會造成機體局部畸形、皮膚功能失調(diào),內(nèi)環(huán)境穩(wěn)態(tài)失衡甚至危及生命。因此,針對創(chuàng)傷后皮膚的修復再生是臨床上亟待解決的問題�；诮M織工程技術制成的皮膚替代物一度成為有效的修復途徑,但這種皮膚替代物缺少汗腺、皮脂腺、毛囊等皮膚附屬器,無法恢復正常皮膚的生理功能。目前,針對皮脂腺、毛囊的再生研究已相對成熟,但針對汗腺再生的研究進展緩慢,有功能的再生汗腺有助于提高機體對外環(huán)境的適應能力和恢復排汗功能。表皮干細胞作為皮膚發(fā)育的祖細胞,在分化為皮膚和皮膚附屬器時有顯著的優(yōu)勢,是汗腺再生研究中首選的干細胞類型。本團隊在前期已證實3D微結構可誘導表皮干細胞分化為汗腺細胞,但不同構架的3D微結構在誘導效應上有差別,這為后續(xù)基于3D生物打印技術的汗腺再生研究在結構選擇上造成了的困難。因此探索不同構架的3D微結構對表皮干細胞行為的影響并建立穩(wěn)定且具良好誘導效應的3D微結構是后續(xù)研究的重要基礎。目的:基于3D生物打印技術探索不同構架的微結構對表皮干細胞行為學變化的影響并建立穩(wěn)定且具良好誘導效應的3D微結構,為創(chuàng)傷后皮膚的修復與再生提供理論基礎。方法:1、表皮干細胞的提取:胚胎期12.5天的GFP-C57BL/6小鼠,剪取其背部皮膚后,采用已成熟的表皮干細胞培養(yǎng)方法進行培養(yǎng)。2、足趾部真皮基質勻漿:選擇出生后0.5天的野生型C57BL/6小鼠,剪取其足趾部研磨成真皮基質勻漿(PD)。3、3D生物打印:選擇3種不同直徑的打印噴頭(210μm、340μm和420μm)構建微結構。4、表皮干細胞行為學檢測:熒光顯微鏡和活/死細胞染色技術用于展示3D微結構中表皮干細胞的增殖能力和細胞活性,免疫熒光染色技術用于檢測3D微結構中表皮干細胞分化為汗腺細胞的能力。5、統(tǒng)計學分析:采用方差分析和樣本t檢驗等方法進行統(tǒng)計學分析。結果:1、三種不同構架的3D微結構均可促進小鼠表皮干細胞的有效增殖。2、在各個觀察時間點上,210μm組3D微結構中的表皮干細胞活性最低,420μm組的細胞活性最高,細胞活性與剪切應力呈反比。3、三種不同構架的3D微結構均可誘導小鼠表皮干細胞分化為汗腺細胞。4、340μm組3D微結構中表皮干細胞可聚集形成類腺樣球形結構。結論:本次研究構建的340μm組3D微結構具有最佳的誘導效應。這為后續(xù)基于3D生物打印技術的汗腺再生研究以及進一步構建具有生物相容性的組織工程表皮模型奠定了基礎。
[Abstract]:Background: skin is the largest organ of the body. Extensive skin damage caused by burn and trauma can cause local malformation, skin dysfunction, homeostasis and even life-threatening. Therefore, the repair and regeneration of post-traumatic skin is an urgent problem to be solved. The skin substitute based on tissue engineering technology was once an effective way to repair the skin, but the skin substitute lacked sweat glands, sebaceous glands, hair follicles and other skin appendages, which could not restore normal skin physiological function. At present, the study of hair follicle regeneration has been relatively mature, but the research on sweat gland regeneration is slow. Functional regenerated sweat gland is helpful to improve the adaptability of the body to external environment and restore the function of sweat excretion. As progenitor cells of skin development, epidermal stem cells have significant advantages in differentiation into skin and skin appendages, and are the preferred stem cell types in the study of sweat gland regeneration. Our team has previously demonstrated that 3D microstructures can induce epidermal stem cells to differentiate into sweat gland cells, but 3D microstructures with different structures have different induction effects. This makes it difficult to select the structure of sweat gland regeneration based on 3D biprint technology. Therefore, it is important to explore the effects of 3D microstructures of different structures on the behavior of epidermal stem cells and to establish stable and well-induced 3D microstructures. Aim: to explore the effects of different structures on the behavioral changes of epidermal stem cells based on 3D biprint technology and to establish a stable and well-induced 3D microstructure which can provide a theoretical basis for the repair and regeneration of post-traumatic skin. Methods: 1. Extraction of epidermal stem cells: GFP-C57BL/6 mice of 12.5 days of embryonic stage were cut off and cultured with mature epidermal stem cells culture method. The dermal matrix homogenate of toe was selected as wild-type C57BL/6 mice 0.5 days after birth. Trimming the toes into dermis matrix homogenate PD-3N 3D biographic printing: selection of three different diameters of print sprayers 210 渭 m, 340 渭 m and 420 渭 m) to construct microstructures .4. Epidermal stem cell behavioral detection: fluorescent microscopy and live / dead cell staining techniques To demonstrate the proliferation and cell activity of epidermal stem cells in 3D microstructures, Immunofluorescence staining was used to detect the ability of epidermal stem cells to differentiate into sweat gland cells in 3D microstructures. Statistical analysis: variance analysis and t-test were used to analyze the ability of differentiation of epidermal stem cells into sweat gland cells. Results three kinds of 3D microstructures with different structures could promote the effective proliferation of mouse epidermal stem cells. In each observation time, the activity of epidermal stem cells was the lowest in 210 渭 m group and the highest in 420 渭 m group. The cell activity was inversely proportional to the shear stress, and three kinds of 3D microstructures with different structures could induce the differentiation of mouse epidermal stem cells into sweat gland cells. 4340 渭 m group could aggregate the epidermal stem cells to form glandular globular structures. Conclusion: the 340 渭 m group of 3D microstructures constructed in this study has the best inductive effect. This will lay a foundation for the further study of sweat gland regeneration based on 3D biprint technology and the further construction of biocompatible tissue engineering epidermis model.
【學位授予單位】：南方醫(yī)科大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R62

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相關期刊論文 前1條

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本文編號：1917918