生物交联剂京尼平交联牛心包生物支架材料的性能

脱细胞牛心包膜因其具有优良特性而成为组织工程生物瓣膜中理想的支架材料.采用冻融+表面活性剂法对牛心包组织进行脱细胞处理后,用戊二醛或京尼平对其进行表面修饰固定.通过HE染色及扫描电镜观察脱细胞效果,并对交联组织进行厚度检测、含水量和接触角测试、力学检测、交联指数和差示扫描量热(DSC)测定、体外降解、溶血试验以及细胞毒性试验,判断两种交联方法对脱细胞牛心包基质的影响,从而选择出交联脱细胞组织的最好方法.结果显示,冻融+表面活性剂法脱除细胞彻底,戊二醛或京尼平交联后组织厚度明显增加分别约20%,25%,亲水性良好,力学特性稳定,交联指数都高达90%,且28 d降解率分别为5%和3%,交联后组织稳定性高.但京尼平组溶血率(0.37%)远小于戊二醛组溶血率(13.77%),且细胞毒性很低.作为瓣膜组织工程支架材料,京尼平交联脱细胞牛心包膜比戊二醛交联效果具有更好生物相容性,交联效果好,是较好的交联方法.     

Study on the Preparation of Decellularized Scaffold of Bovine Pericardium

Objective: To search for the best procedure on preparation of acellular bovine pericardium,so to provide scaffolds for constructing tissue-engineering Methods:The bovine pericardiums were treated with 5 methods,which were divided into 6 groups.Group A:Fresh bovinepericardium;GroupB:Trypsin-detergentgroup;GroupC:Freeze-thaw-detergent24 h group;Group D:.Freeze-thaw-detergent 48 h group;Group E:Freeze-thaw-nuclease group;Group F:Detergent-nuclease group.Then,by HE staining and scanning electron microscope to observe the effects of decellularization and fibrous changes among the 6 groups;by water content testingmechanical testing to observe the changes in physical properties of the matrix;by detecting the DNA content of each group to determine the effect of decellularization qualitatively;by cytotoxicity test to detect the biocompatibility of bovine pericardium in each group.Results:The 5 methods can all remove the cellular components effectively,compared with the fresh bovine pericardium,the water content of each decellularized group were increased (P＜0.05),while the DNA content decreased (P＜0.05),with statistically significant differences.Of group E,the fibers were a little disorder,with the largest tension and the elastic modulus increased,while the rupture tensile rate decreased.Compared with fresh bovine pericardium,the largest tension of the other decellularization groups were all decreased (P＜0.05).The fibers of group B,group D were irregularly arranged and also with ruptures,both the elastic modulus and the rupture tensile rate decreased(P＜0.05).In group C and F,the fibers were dense and their direction was normal,the elastic modulus and the rupture tensile rate were similar to the fresh bovine pericardium (P＞0.05).Cytotoxicity results showed that the cell toxicity of group B,group C,group D,group E and group F were respectively 0.9,0.6,1.0,1.0 and 0.5,each group were qualified toxicity test,in which group C and group F were with the lowest cytotoxicity.Conclusion:Group C and group F can remove the cell components of bovine pericardium successfully,while maintaining the major structural components and the histological and biological properties of bovine pericardium,and with low cytotoxicity.However,group C is more economical than group F,and easier to operate.So the method on freeze-thaw-detergent 24 h can be the best choice to produce a decellularized bovine pericardium.     

交联剂对脱细胞膀胱基质生物力学性能的影响

目的 研究不同交联剂对猪脱细胞膀胱基质的组织结构影响,并比较其生物力学性能,为盆底修复替代材料的选择提供依据.方法 采用表面活性剂+酶消化法去除新鲜猪膀胱的细胞成分,将脱细胞膀胱基质随机分为3组,A组经0.25％戊二醛交联,B组经0.625％京尼平交联,C组未交联.对各组材料进行HE染色,观察纤维的变化情况.使用生物力学性能测试系统检测抗拉强度、断裂伸长率、弹性模量,并进行统计学分析.结果 京尼平交联脱细胞膀胱基质后呈深蓝色,保持了天然组织构架的完整,纤维更加致密.戊二醛交联脱细胞膀胱基质后呈浅黄色,纤维排列紊乱且有断裂.新鲜猪膀胱经上述三种方法处理后,其弹性模量增大、断裂伸长率减小,而其中京尼平交联处理的脱细胞膀胱基质力学性能与新鲜膀胱组织更为相近.结论 京尼平交联的脱细胞膀胱基质组织结构的形态佳,同时较大限度地保留膀胱组织的力学性能,可能是较理想的盆底重建材料.     

心包纤维取向对压缩性能的影响

目的探究不同纤维取向心包组织压缩后的力学性能,评估对经导管瓣膜耐久性的影响。方法将心包条分为横、纵、斜向的心包条各12组,每组中包含3条,其中1条不压缩,另外两条分别在12.5、25 N下压缩1 h。利用单轴拉伸方法对压缩后的心包进行测试,分析其力学性能的变化。结果不同纤维取向心包在压缩前后均表现出明显的各向异性。压缩前后的割线模量、弹性模量以及极限拉伸强度,总体上都表现为纵向斜向横向。纵、斜向牛心包在25 N压缩后的平均拉伸强度比值分别为0.92和0.87,压缩前后的拉伸强度比值均有显著性差异(P0.05)。横向牛心包25 N压缩后的平均拉伸强度比值为0.97,压缩前后的拉伸强度比值无显著性差异(P0.05)。结论不同纤维取向心包在低应力、短时间下压缩损伤都很小。在经导管瓣膜的设计和制造过程中,应该充分考虑心包纤维取向和压缩后强度变化对瓣膜性能的影响。     

环氧改性牦牛心包材料的力学性能

本文研究了经新型生物材料交联剂——乙二醇二缩水甘油醚(ethylene diglycidyl ether)处理牦牛心包材料的粘弹性,并与新鲜样品及戊二醛处理样品进行比较。环氧交联剂主要在胶原纤维的分子链间形成交联键,材料的应力松驰速率下降;通过对应力——应变曲线,材料的模量和断裂强度等分析,证明环氧交联剂处理的牦牛心包较用戊二醛处理的有较高的抗张强度和断裂伸长率,并具有一定弹性,作为人工心瓣的瓣叶,在力学性能上是有利的。     

Effect of suturing on the mechanical properties of bovine pericardium — implications for cardiac valve bioprosthesis

In the fabrication of a bioprosthetic heart valve from bovine pericardial tissues, the tissues are subjected to suturing. The stress-strain response of sutured as well as unsutured strips of this tissue were examined. The stress-strain response was determined using a tensile-testing machine. It was found that suturing weakens the tissue in that sutured strips are more extensible, exhibit a lower stress at rupture and a lower final elastic modulus. In addition, it was also found that the bigger the suture/needle size used the greater the decrease in tissue strength. In all, tissue strength was observed to decrease by 22 to 59% in this study. The weakening of the tissue is attributed to the puncture holes created by the surgeon's needle which create regions of weakness. This response of bovine pericardial tissue to suturing should be given due consideration in the fabrication of a bioprosthetic heart valve using this tissue.     

Orthotropic Mechanical Properties of Chemically Treated Bovine Pericardium

To facilitate bioprosthetic heart valve design, especially in the use of novel antimineralization chemical technologies, a thorough understanding of the multiaxial mechanical properties of chemically treated bovine pericardium (BP) is needed. In this study, we utilized a small angle light scattering based tissue pre-sorting procedure to select BP specimens with a high degree of structural uniformity. Both conventional glutaraldehyde (GL) and photo-oxidation (PO) chemical treatment groups were studied, with untreated tissue used as the control group. A second set of GL and PO groups was prepared by prestretching them along the preferred fiber direction during the chemical treatment. An extensive biaxial test protocol was used and the resulting stress-strain data fitted to an exponential strain energy function. The high structural uniformity resulted in both a consistent mechanical response and low variability in the material constants. For free fixed tissues, the strain energy per unit volume for GL treated BP was 2.8 times that of PO treated BP at an equibiaxial Greens strain level of 0.16. Prestretched tissues exhibited a profound increase in both stiffness and the degree of anisotropy, with the GL treatment demonstrating a greater effect. Thus, structural control leads to an improved understanding of chemically treated BP mechanical properties. Judicious use of this knowledge can facilitate the design and enhanced long-term performance of bioprosthetic heart valves. © 1998 Biomedical Engineering Society. PAC98: 8790+y, 8745Bp, 8780+s     

脱细胞肠系膜制备生物膜支架的实验研究

目的应用肠系膜脱细胞基质制备生物膜支架,探讨其理化特性和生物学特性。方法应用反复冻融肠系膜组织后,将肠系膜置于胰蛋白酶消化脱肠系膜脱细胞,并分为肠系膜基质组(A组)和去细胞肠系膜基质组(B组),通过HE染色、电镜、DNA检测、细胞毒性实验、拉伸力学测试,检测两组肠系膜基质的理化特性;制备覆膜支架,植入兔髂血管,术后1周、1月、2月超声检测血管血流情况,血管取材病理检测。结果 HE染色、电镜检测结果表明,B组基质组织疏松,纤维排列较整齐,未见细胞存留; DNA检测结果表明,B组DNA表达水平低,脱细胞较彻底; CCK-8细胞毒性实验检测结果表明,两组均无细胞毒性; FDA-PI荧光染色结果表明,两组细胞存活良好,未见死亡细胞;拉伸力学测试结果表明,两组最大拉伸力、最大力伸长率、屈服强度、屈服点伸长率差异无统计学意义;脱细胞肠系膜支架植入兔血管后超声检测两组早期通畅性良好,支架植入2月内皮增生较明显。结论冻融和酶消化法脱肠系膜细胞,肠系膜基质去除细胞彻底,无细胞毒性,力学特征良好;肠系膜支架植入血管后早期通畅性良好,2月后内皮增生明显。     

不同方法处理的牛心包生物相容性和钙化结果分析

目的 观察脱细胞处理对牛心包体内生物相容性及钙化的影响并与戊二醛处理的牛心包进行比较；材料和方法对新鲜牛心包随机分为三组，A、戊二醛处理组：新鲜牛心包采用0．5％的戊二醛进行处理；B、新鲜牛心包组：新鲜牛心包保存于四联抗生素液中；C、脱细胞组：新鲜牛心包酶．去污剂联合脱细胞后保存于四联抗生素液中。上述处理后的牛心包经细菌培养，显示无细菌生长后，植入雄性昆明小鼠皮下三周观察炎性浸润及钙化情况。采用von Kossa钙染色检查进行定性分析；原子吸收光谱检查进行钙含量定量分析。同时体外对三组牛心包进行力学和热皱缩温度进行测试。结果 三组间炎性浸润程度明显不同，0．5％的戊二醛处理组钙化程度明显增高，每克干重牛心包平均含钙量71．2mg，较其他两组有明显差异（P〈0．001），新鲜牛心包组（平均2．12mg／g干重）和脱细胞组（1．41mg／g干重）间钙化程度方面亦有显著统计学意义（P〈0．001）。组织学钙染色光镜下检查显示三组间黑色颗粒有明显差异，支持三组间钙含量定量结果的差异。戊二醛处理组的力学性能与优于新鲜组和脱细胞处理组，热皱缩温度高于新鲜组和脱细胞处理组。结论 新鲜牛心包脱细胞处理后钙化显著降低，免疫源性降低，有很好的生物相容性。     

应用Triton-x100加丹参酚酸B制备脱细胞血管支架及其血液相容性研究

应用传统的Triton-x100法与应用Triton-x100和丹参酚酸B法制备脱细胞血管支架,比较这两种支架的理化性质及其血液相容性,展望应用丹参酚酸B后处理脱细胞血管支架在临床上治疗心脑血管疾病方面的应用前景,以及在组织工程血管材料方面的开发空间。选择SD大鼠16只,雄性,鼠龄7周,体质量250 g左右。解剖大鼠,取腹主动脉;随机分为2组,每组8个。用Triton-x100法(Tx组)、Triton-x100加入丹参酚酸B法(Tx-sal 组),加工制成两种脱细胞血管支架。观察脱细胞血管支架的宏观形貌;亲水性实验测定亲水性;体外溶血实验计算体外溶血率;复钙化凝血时间实验测定复钙化凝血时间;血小板黏附实验测定动态凝血时间,以及体外血小板的黏附;补体激活实验测定补体激活程度情况;比较两组脱细胞血管支架的血液相容性。Tx组接触角为76.36°±4.65°,Tx-sal组接触角为71.26°±3.55°,Tx组接触角度数略高于Tx-sal 组,差异无显著统计学意义。Tx-sal 组溶血率为1.5%,Tx组溶血率为2.1%。Tx组复钙化凝血时间为(212±11.32)s,Tx-sal 组复钙化凝血时间为(231±13.53)s,Tx组与Tx-sal 组的复钙化凝血时间差异有显著统计学意义。Tx-sal组相比Tx组延缓了凝血时间,抗凝血性更加优良。Tx-sal 组黏附的血小板数目低于Tx组的相应数目。Tx-sal 组补体激活水平低。与传统的Triton-x100法制备的脱细胞血管支架相比,加入丹参酚酸B后处理的脱细胞血管支架具有更加优良的血液相容性,更适合作为组织工程材料应用于心脑血管疾病的治疗,在临床医学上的应用前景更加广阔。     

机械应力对牛心包瓣原发性损坏的影响

引起牛心包瓣原发性损坏的生理学和病理学因素与机械学因素同等重要。从我院再次手术中收集到的22只失效瓣膜来看,在瓣架尖端区和侧边区的损坏发生率分别占30.3％和36％,撕裂、磨损和钙化的发生率分别占25.8％、18％和31.6％。牛心包片的弯曲试验和牛心包瓣的显微镜观察表明,由于经戊二醛处理的牛心包片的拉伸弹性模量是压缩模量的25倍,因此当牛心包片弯曲变形时,弯曲内侧必然出现挤压屈服,并逐渐发展到楔状挤压。牛心包瓣临床失效的形式及其分布提示,瓣叶损坏主要是由于瓣叶在启闭活动过程中的这种弯曲挤压引起的。因此在改进牛心包瓣时,应考虑机械应力,特别是弯曲变形的应力对瓣膜损坏的影响。     

Effect of Biodegradable Epoxidized Castor Oil on Physicochemical and Mechanical Properties of Epoxy Resins

Summary: Biobased epoxy materials were prepared from diglycidyl ether of bisphenol A (DGEBA) and epoxidized castor oil (ECO) initiated by a latent thermal catalyst. The physicochemical and mechanical interfacial properties of the DGEBA/ECO blends were investigated. As a result, the thermal stability of the cured epoxy blends showed a maximum value in the presence of 10 wt.‐% ECO content, which was attributed to the excellent network structure in the DGEBA/ECO blends. The storage modulus and glass transition temperature of the blends were lower than those of neat epoxy resins. The mechanical interfacial properties of the cured specimens were significantly increased with increasing the ECO content. This could be interpreted in terms of the addition of larger soft segments of ECO into the epoxy resins and thus reducing the crosslinking density of the epoxy network, which results in increasing toughness in the blends.

K_IC values of the DGEBA/ECO blends as a function of ECO content.     

不同预处理脱细胞牛骨基质复合成骨化骨髓间充质干细胞生物相容性研究

目的将经成骨化诱导后大鼠骨髓间充质干细胞(MSC)与脱细胞牛骨基质(ABECM)复合,构建组织工程骨。实验着重观察不同预处理方法对构建的组织工程骨的过程——细胞与ABECM复合和生长的影响,为有效快速构建组织工程化骨奠定理论基础。方法用Wistar大鼠2只,从股骨取MSC在体外扩增、纯化、诱导成骨化,做苏木精-伊红染色鉴定。实验组用胎牛血清(FBS)、多聚赖氨酸(PLL),对照组用磷酸盐缓冲溶液预处理的ABECM复合构建组织工程骨,体外培养。用流式细胞仪计数并计算细胞黏附率、测定碱性磷酸酶活性,倒置光学显微镜、扫描电子显微镜观察细胞生长、增殖情况。结果接种后6、12h实验组细胞黏附率高于对照组(P0.05)。扫描电子显微镜观察:复合培养第3天,实验组细胞在材料表面已完全伸展,附着在材料表面;对照组细胞大多呈不规则形,未完全伸展。碱性磷酸酶活性测定:复合培养第3、6天,实验组的碱性磷酸酶活性均明显高于对照组(P0.05)。结论用FBS和PLL分别预处理,可以改善ABECM材料表面特性,利于提高细胞接种效率,从而能更有效、更快速地构建组织工程骨。     

种植施万细胞的脱细胞同种异体神经移植物对大鼠坐骨神经缺损的修复作用

目的观察种植施万细胞的脱细胞同种异体神经移植物,桥接大鼠坐骨神经缺损后的神经再生。方法应用酶反复消化法与差速贴壁法体外分离培养乳鼠施万细胞;显微注射法将施万细胞种植到脱细胞同种异体神经移植物内;再应用种植施万细胞的脱细胞同种异体神经移植物桥接大鼠坐骨神经10 mm缺损。光镜、透射电镜和扫描电镜观察再生神经的形态结构、有髓神经纤维数量、平均髓鞘厚度并进行统计学分析。结果光、电镜观察到实验组(SCs+ARSN)的施万细胞在再生神经纤维中互相连结纵行排列成类似Büngner带样细胞链,对照组(ARSN)未见到施万细胞的链状排列。实验组再生有髓神经纤维的髓鞘厚度较对照组均匀且较厚,有髓神经纤维数量和平均髓鞘厚度明显多于对照组(P<0.05)。结论种植施万细胞的脱细胞同种异体神经移植物对缺损的坐骨神经再生有更加有效的促进作用。     

微囊牛嗜铬细胞移植镇痛作用的实验研究

目的 观察微囊牛嗜铬细胞（BCC）的镇痛作用；方法 应用胶原酶消化法分离、提取牛嗜铬细胞，进行APA微囊化；制作大鼠坐骨神经结扎和福尔马林实验疼痛动物模型，模型大鼠各分为4组（微囊BCC组、裸细胞组、空囊组和生理盐水组），分别在蛛网膜下腔植入微囊化BCC、未包囊BCC、空囊和生理盐水，观察坐骨神经结扎疼痛模型动物冷、热刺激后的行为学变化及福尔马林疼痛模型动物福尔马林刺激后的行为学变化；结果 坐骨神经结扎实验动物冷、热刺激后，徵中BCC组冷刺激收缩时间、次数和热刺激潜伏期差异分数明显下降，裸细胞组下降只维持了2周，而空囊组和生理盐水组无变化；福尔马林实验动物刺激后，微囊BCC组收缩次数下降了13周以上，裸细胞组下降只维持了2周，而空囊组和生理盐水组无变化；结论 微囊牛嗜铬细胞对异种动物疼痛具有镇痛作用。     

单宁酸与牛颈静脉的交联机制研究

为从化学角度探索单宁酸与牛颈静脉的交联机制,使用5种含有单宁酸不同特征基团的模式分子,分别对戊二醛固定后的牛颈静脉带瓣血管进行交联处理。体内实验采用20只SD大鼠皮下植入处理后的带瓣血管,21和60 d后用原子吸收光谱法测定抗钙化效果,用Masson和EVG染色测定交联程度。体外采用热稳定性和酶解实验检测交联程度,并使用傅里叶变换红外光谱检测带瓣血管化学结构的变化,推测单宁酸与带瓣血管交联作用的类型;进一步在确定交联类型的体系中加入系列浓度梯度(0.1、0.3、0.5、1 M)的尿素,对交联类型使用同样方法进行验证。结果显示,含多酚羟基的单宁酸组带瓣血管交联程度最高,21和60 d钙含量分别为2.25、8.26 mg/g,抗钙化能力最优(P<0.05);傅里叶变换红外光谱图中,单宁酸组带瓣血管有氢键生成。浓度梯度尿素实验表明,随着尿素浓度的增加,单宁酸与牛颈静脉组织交联程度下降(P<0.05),60 d时单宁酸组钙含量为8.10 mg/g,远低于加入0.1 M尿素组的16.83 mg/g和0.5 M尿素组的50.76 mg/g(P < 0.001)。研究表明,单宁酸对牛颈静脉带瓣血管的交联作用是由其所含的多酚羟基与组织中蛋白质基团间通过氢键而产生的,为进一步优化单宁酸抗钙化处理流程提供依据。     

杆状和片状三周期极小曲面模型孔隙特征与力学性能对比

目的 分析对比杆状与片状三周期极小曲面(triply periodic minimal surface,TPMS)模型孔隙特征与力学性能,构建高比表面积、低刚度和高强度的多孔结构.方法 构建相同孔隙率的D、G、P 3种单元杆状TPMS与片状TPMS模型,对比模型的孔径、杆径、比表面积等孔隙特征;利用有限元方法分析模型的...     

大学生双性化人格特征研究

目的：探讨中国双性化大学生的人格特点。方法：使用BSRI和CPAI-2量表对云南省昆明地区976名大学生被试进行集体测试，并运用方差分析方法统计所得数据。结果：中国大学生四种性别类型的分布差异显著；双性化人格特征中明显带有男性化和女性化的典型特征，但又与中性化的人格特征差异较大。结论：中国双性化大学生具有“领导性”和“人际取向”的人格特征。     

硅胶材料穿刺力学性能的实验研究

目的 通过实验研究获取静脉穿刺机器人较为合适的穿刺方法。方法 对弹性材料采用不同的穿刺速度、角度进行穿刺,从而获得各种穿刺方法过程中的力-时间曲线。结果 在穿刺过程中,随着穿刺角度增大,所需的穿刺力越小;穿刺速度越快,穿刺力越大。结论 在进行静脉穿刺机器人穿刺方法设计时,宜采用 40°～45°穿刺角度和120～300 mm/min穿刺速度进行穿刺。研究结果为静脉穿刺机器人穿刺角度及速度的选取提供参考。     

The Influence of RAFT on the Microstructure and the Mechanical Properties of Photopolymerized Poly(butyl acrylate) Networks

Statistical networks of butyl acrylate and 1,4‐butanediol diacrylate are photopolymerized in bulk, both in the presence and absence of the RAFT agent EPHT. The amounts of crosslinker and RAFT agent are varied. The Young's moduli of the RAFT networks are always lower than those of conventionally obtained networks. With increasing amounts of RAFT agent, the Young's modulus decreases and the strain at break values increase, respectively. The network density, the apparent molar mass of the network chains and the swelling show significant differences between RAFT‐ and conventionally obtained networks. The degree of swelling and the amount of extractable polymer increase largely with the amount of RAFT agent. Kinetic DSC measurements show that the gel point is delayed with increasing amounts of RAFT agent. In addition, a rate retardation effect with increasing RAFT agent concentration is observed.