中国组织工程研究与临床康复
中國組織工程研究與臨床康複
중국조직공정연구여림상강복
JOURNAL OF CLINICAL REHABILITATIVE TISSUE ENGINEERING RESEARCH
2008年
1期
166-169
,共4页
陈冀衡%杨茂元%李继勇%林献忠%毕好生
陳冀衡%楊茂元%李繼勇%林獻忠%畢好生
진기형%양무원%리계용%림헌충%필호생
微囊化%嗜铬细胞%体外培养%免疫隔离%生物材料%组织工程
微囊化%嗜鉻細胞%體外培養%免疫隔離%生物材料%組織工程
미낭화%기락세포%체외배양%면역격리%생물재료%조직공정
背景:课题组前期在建立海藻酸钠-多聚赖氨酸-海藻酸钠微囊包裹活细胞制备技术的基础上,已证明微囊化嗜铬细胞有良好的镇痛效果,而该微囊包被材料的免疫隔离作用尚需明确.目的:观察海藻酸钠-多聚赖氨酸-海藻酸钠微囊化嗜铬细胞移植到大鼠眼前房和足胝部的免疫排斥反应,评价微囊化技术的免疫隔离作用.设计:随机对照动物实验.单位:华中科技大学同济医学院附属同济医院麻醉学教研室.材料:选用雌性 SD大鼠48只,鼠龄3个月,由华中科技大学同济医学院实验动物部提供.实验过程中对动物处置符合动物伦理学标准.实验所用海藻酸钠、多聚赖氨酸为美国Sigma公司产品,微囊发生器为德国赠送.方法:实验于2002-09/2003-09在华中科技大学同济医学院附属同济医院麻醉学实验室完成.①取6名脑死亡健康成人的肾上腺髓质,经分离、消化、培养后,制备成人嗜铬细胞悬液.供者家属对实验知情同意,实验方案通过医院伦理委员会批准.采用海藻酸钠-聚赖氨酸-海藻酸钠法制作空微囊和微囊化细胞.②48只大鼠被随机分为3组:人嗜铬细胞移植组、空微囊移植组、微囊化人嗜铬细胞移植组,每组分眼前房和足胝部两个部位进行移植,每个部位8只.人嗜铬细胞移植组分别将2×1010 L-1细胞悬液注入大鼠右眼前房和左足胝部.空微囊移植组和微囊化人嗜铬细胞组分别吸取空微囊(100个微囊)或ME-HCC(100个微囊,每个微囊包裹400～500个细胞)注入大鼠右眼前房和左足胝部.主要观察指标:于移植术后第7天采用ELISA法测定血清白细胞介素2水平.采用激光散射比浊仪测定血清IgG和IgM水平.移植术后第28天取大鼠右侧眼球及左侧足组织作常规切片,苏木精-伊红染色,40倍光镜下观察组织形态.结果:大鼠48只均进入结果分析.①血清白细胞介素2,IgG,IgM水平:空微囊移植组和微囊化人嗜铬细胞移植组均低于人嗜铬细胞移植组,差异有显著性意义(t=8.544～21.64,P < 0.01).②大鼠眼前房和足胝部组织形态:人嗜铬细胞移植组大鼠的眼前房内和足胝部可见大量淋巴细胞和中性粒细胞浸润.空微囊移植组和微囊化人嗜铬细胞移植组大鼠眼前房和足胝部仅见少量淋巴细胞和中性粒细胞.结论:海藻酸钠-多聚赖氨酸-海藻酸钠微囊化所产生的良好生物相容性及其机械稳定性,使之有效地发挥了免疫排斥隔离作用.
揹景:課題組前期在建立海藻痠鈉-多聚賴氨痠-海藻痠鈉微囊包裹活細胞製備技術的基礎上,已證明微囊化嗜鉻細胞有良好的鎮痛效果,而該微囊包被材料的免疫隔離作用尚需明確.目的:觀察海藻痠鈉-多聚賴氨痠-海藻痠鈉微囊化嗜鉻細胞移植到大鼠眼前房和足胝部的免疫排斥反應,評價微囊化技術的免疫隔離作用.設計:隨機對照動物實驗.單位:華中科技大學同濟醫學院附屬同濟醫院痳醉學教研室.材料:選用雌性 SD大鼠48隻,鼠齡3箇月,由華中科技大學同濟醫學院實驗動物部提供.實驗過程中對動物處置符閤動物倫理學標準.實驗所用海藻痠鈉、多聚賴氨痠為美國Sigma公司產品,微囊髮生器為德國贈送.方法:實驗于2002-09/2003-09在華中科技大學同濟醫學院附屬同濟醫院痳醉學實驗室完成.①取6名腦死亡健康成人的腎上腺髓質,經分離、消化、培養後,製備成人嗜鉻細胞懸液.供者傢屬對實驗知情同意,實驗方案通過醫院倫理委員會批準.採用海藻痠鈉-聚賴氨痠-海藻痠鈉法製作空微囊和微囊化細胞.②48隻大鼠被隨機分為3組:人嗜鉻細胞移植組、空微囊移植組、微囊化人嗜鉻細胞移植組,每組分眼前房和足胝部兩箇部位進行移植,每箇部位8隻.人嗜鉻細胞移植組分彆將2×1010 L-1細胞懸液註入大鼠右眼前房和左足胝部.空微囊移植組和微囊化人嗜鉻細胞組分彆吸取空微囊(100箇微囊)或ME-HCC(100箇微囊,每箇微囊包裹400～500箇細胞)註入大鼠右眼前房和左足胝部.主要觀察指標:于移植術後第7天採用ELISA法測定血清白細胞介素2水平.採用激光散射比濁儀測定血清IgG和IgM水平.移植術後第28天取大鼠右側眼毬及左側足組織作常規切片,囌木精-伊紅染色,40倍光鏡下觀察組織形態.結果:大鼠48隻均進入結果分析.①血清白細胞介素2,IgG,IgM水平:空微囊移植組和微囊化人嗜鉻細胞移植組均低于人嗜鉻細胞移植組,差異有顯著性意義(t=8.544～21.64,P < 0.01).②大鼠眼前房和足胝部組織形態:人嗜鉻細胞移植組大鼠的眼前房內和足胝部可見大量淋巴細胞和中性粒細胞浸潤.空微囊移植組和微囊化人嗜鉻細胞移植組大鼠眼前房和足胝部僅見少量淋巴細胞和中性粒細胞.結論:海藻痠鈉-多聚賴氨痠-海藻痠鈉微囊化所產生的良好生物相容性及其機械穩定性,使之有效地髮揮瞭免疫排斥隔離作用.
배경:과제조전기재건립해조산납-다취뢰안산-해조산납미낭포과활세포제비기술적기출상,이증명미낭화기락세포유량호적진통효과,이해미낭포피재료적면역격리작용상수명학.목적:관찰해조산납-다취뢰안산-해조산납미낭화기락세포이식도대서안전방화족지부적면역배척반응,평개미낭화기술적면역격리작용.설계:수궤대조동물실험.단위:화중과기대학동제의학원부속동제의원마취학교연실.재료:선용자성 SD대서48지,서령3개월,유화중과기대학동제의학원실험동물부제공.실험과정중대동물처치부합동물윤리학표준.실험소용해조산납、다취뢰안산위미국Sigma공사산품,미낭발생기위덕국증송.방법:실험우2002-09/2003-09재화중과기대학동제의학원부속동제의원마취학실험실완성.①취6명뇌사망건강성인적신상선수질,경분리、소화、배양후,제비성인기락세포현액.공자가속대실험지정동의,실험방안통과의원윤리위원회비준.채용해조산납-취뢰안산-해조산납법제작공미낭화미낭화세포.②48지대서피수궤분위3조:인기락세포이식조、공미낭이식조、미낭화인기락세포이식조,매조분안전방화족지부량개부위진행이식,매개부위8지.인기락세포이식조분별장2×1010 L-1세포현액주입대서우안전방화좌족지부.공미낭이식조화미낭화인기락세포조분별흡취공미낭(100개미낭)혹ME-HCC(100개미낭,매개미낭포과400～500개세포)주입대서우안전방화좌족지부.주요관찰지표:우이식술후제7천채용ELISA법측정혈청백세포개소2수평.채용격광산사비탁의측정혈청IgG화IgM수평.이식술후제28천취대서우측안구급좌측족조직작상규절편,소목정-이홍염색,40배광경하관찰조직형태.결과:대서48지균진입결과분석.①혈청백세포개소2,IgG,IgM수평:공미낭이식조화미낭화인기락세포이식조균저우인기락세포이식조,차이유현저성의의(t=8.544～21.64,P < 0.01).②대서안전방화족지부조직형태:인기락세포이식조대서적안전방내화족지부가견대량림파세포화중성립세포침윤.공미낭이식조화미낭화인기락세포이식조대서안전방화족지부부견소량림파세포화중성립세포.결론:해조산납-다취뢰안산-해조산납미낭화소산생적량호생물상용성급기궤계은정성,사지유효지발휘료면역배척격리작용.
BACKGROUND: Based on previous technique prepared for encapsulating living cells with alginate-polysine- alginate (APA) microcapsules, it has been confirmed that microencapsulated chromaffin cells have good analgesic effects. The immunoisolated effects of such microcapsule materials need to be evaluated. OBJECTIVE: This study aimed to investigate the immunological rejections of APA microencapsulated chromaffin cells transplanted into rat anterior chamber of eyes and tendon of feet, and to evaluate the immunoisolated effect of microencapsulation.DESIGN: A randomized controlled animal experiment. SETTING: Department of Anesthesiology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology.MATERIALS: Forty-eight female SD rats, with the age of 3 months, were provided by the Laboratory Animal Center, Tongji Medical College, Huazhong University of Science and Technology. The protocol was carried out in accordance with ethical guidelines for the use and care of animals. Alginate and polylysine used in the experiment were the products of Sigma Company, USA. Microcapsule generator was gifted by Germany. METHODS: This study was performed at the Department of Anesthesiology, Tongji Hospital of Tongji Medical College, Huazhong University of Science and Technology from September 2002 to September 2003. Suprarenal medulla was taken from 6 healthy adult cadavers of brain death. After isolated, digested and cultured, suprarenal medulla was prepared into chromaffin cell suspension. Written informed consents were obtained from the family members of donors, and the protocol was given approval by the Ethics Committee of the hospital. Empty microcapsules and microencapsulated cells were prepared by APA. The 48 rats were randomly divided into the human chromaffin cell (HCC) group, the empty microcapsule group and the microencapsulated HCC (ME-HCC) group. In each group, there were two transplanted regions of anterior chamber of eyes and tendon of feet, with 8 rats used for each region. Each rat in the HCC group was perfused 2×1010 L-1 cell suspension into the anterior chamber of eyes and tendon of feet. Those in the empty microcapsule group and the ME-HCC group were perfused 100 empty capsules and ME-HCCs (100 microcapsules, 400-500 HCCs per microcapsule) into the same regions, respectively. MAIN OUTCOME MEASURES: On day 7 after transplantation, serum interleukin (IL)-2 level was determined by ELISA. Serum IgG and IgM levels were determined with a laser turbidimeter. On day 28 after transplantation, rat right eyeball and left feet were harvested, routinely sliced and stained by haematoxylin-eosin (HE). Histo-morphological structure was observed under a 40×light microscope. RESULTS: Forty-eight rats were included in the final analysis. Serum IL-2, IgG and IgM levels were significantly lower in the empty microcapsule group and ME-HCC group than in the HCC group (t=8.544-21.64, P < 0.01). A lot of lymphocyte and neutrophile infiltration could be found in the anterior chamber of eyes and tendon of feet of rats in the HCC group, but a little seen in that of the empty microcapsule group and ME-HCC group. CONCLUSION: APA microencapsulation has an effective immunoisolated effect on immunological rejection due to its good biocompatibility and mechanical stability.
