肝细胞癌细胞粘弹性的改变及细胞骨架干扰剂的影响

目的 从细胞骨架角度探讨肝癌细胞粘弹性改变的基础 ,及细胞骨架结构干扰剂对其改变的影响。方法 采用微管吸吮技术测定正常肝细胞和肝癌细胞的粘弹性 ,三参数标准线性固体模型拟合实验结果。结果 肝癌细胞的弹性系数较肝细胞增高 ,秋水仙素、长春花碱使癌细胞粘弹性系数与肝细胞相比呈现不同的效应方式和强度 ,细胞松驰素D则使 2种细胞粘弹性系数均呈现相似的显著下降趋势。结论 细胞骨架结构的完整性是维持肝细胞和肝癌细胞粘弹性及被动变形机能的必要条件 ,2种细胞的粘弹性对细胞骨架干扰剂的效应差异 ,反映了肝癌细胞在细胞骨架结构和机能状态中的变化 ,这种变化可能影响癌细胞的浸润、转移特性以及与微观力学环境的相互作用。

QCM实时监测五价砷胁迫对水稻悬浮细胞粘弹性的影响

本文通过QCM技术实时监测水稻悬浮细胞在不同浓度五价砷连续及单独胁迫下的粘弹性变化,从细胞整体水平评估了重金属对水稻悬浮细胞力学性质的影响。结果表明:1)在2~10 mmol/L范围内,五价砷对水稻悬浮细胞的连续和单独胁迫的影响均随着浓度增加而增强;2)在连续胁迫和单独胁迫下,随着五价砷浓度的增加,水稻悬浮细胞粘弹性指数均下降,细胞软硬度减小即细胞变软;3)在五价砷连续梯度浓度胁迫下,水稻悬浮细胞粘弹性指数呈梯度下降;4)在五价砷不同浓度单独胁迫下,随着五价砷浓度的增加,水稻悬浮细胞粘弹性指数变化逐渐平缓,即水稻悬浮细胞随着五价砷浓度升高,敏感程度降低。以期建立一种用来衡量重金属胁迫对植物影响的一种新方法。

大鼠肺微血管内皮细胞培养及其粘弹性研究

为了建立肺微血管内皮细胞培养方法 ,研究肺微血管内皮细胞粘弹性。我们取大鼠肺周边组织 (宽度不应大于 1.5 mm) ,将组织剪成 1.5 mm× 1mm× 1mm的组织块 ,贴入无菌的 2 5 cm3培养瓶 ,每瓶 10～ 15块 ,同时加入含 2 0胎牛血清、肝素 90 U/ml、L-谷氨酰胺 4mmol、青霉素 10 0 U/ml和链霉素 10 0 μg/ml的 DMEM培养基 3ml,放入 37℃二氧化碳培养箱中静置培养 ;8h后翻转培养瓶 ,6 0 h后取出肺组织块 ,接着继续培养 2～ 4d后进行传代。最后消化分离肺微血管内皮细胞 ,用微管吸吮系统研究肺微血管内皮细胞粘弹性。结果显示 :肺微血管内皮细胞通过倒置相差显微镜观察 ,细胞呈鹅卵石镶嵌状排列 ,状如梭形或多角形 ,大小均匀 ,胞核清晰 ,呈卵圆形 ,胞浆丰富 ; 因子相关抗原免疫荧光染色呈阳性 ;肺微血管内皮细胞弹性模量 K1 =49.3± 9.2 Pa、K2 =73.2±2 4.8Pa、粘性系数 μ=19.2± 7.2 Pa.s。这些结果表明用组织块法培养肺微血管内皮细胞是可行的 。

烧伤血清对人肝细胞粘弹性的影响

目的烧伤血清影响人肝细胞亚细胞水平的细胞骨架变化的力学研究．方法应用体外人肝细胞的培养和细胞力学（微管吸吮技术）动态观察了烧伤患者（５０％以上，Ⅱ～Ⅲ度）血清对肝细胞损伤的酶学变化和粘弹性的变化．结果烧伤血清对肝细胞有直接损伤作用，且随培养时间的延长损伤加重（从１ｈ到２４ｈ四个时相点，ＡＳＴ（Ｕ／Ｌ）分别为１２０±４１，１４６±３３，１６７±２１，２１７±１２，Ｐ＜００１；ＡＬＴ（Ｕ／Ｌ）分别为２２±０４，３０±０５，３７±０３，４１±０６，Ｐ＜００５和Ｐ＜００１；ＬＤＨ（Ｕ／Ｌ）分别为３２６±７３，４０１±１０７，６０４±１６６，７０３±１８３，Ｐ＜００１）．肝细胞粘弹性的变化，烧伤血清作用２ｈ粘弹性明显增加［Ｋ１（Ｎ／ｍ２）：３８４±４６，Ｋ２（Ｎ／ｍ２）：２８２±３２，Ｕ（Ｎ．ｓ／ｍ２）：９８±１１，Ｐ＜００１］，从６ｈ开始到２４ｈ肝细胞粘弹性降低，刚性增加，这和肝细胞损伤后酶的变化一致．结论烧伤血清直接损伤肝细胞，引起酶释放并随作用时间的延长而增加；生物力学技术检测烧伤血清对肝细胞的损伤过程有一个先激活后损伤的过程，这个现象用酶的变化是无法反?

QCM监测不同浓度肌力药物作用下乳鼠原代心肌细胞的粘弹性响应

采用石英晶体微天平(QCM)非侵入式实时监测乳鼠原代心肌细胞于金晶体电极上的动态黏附响应与不同浓度肌力药物下的粘弹性响应。在乳鼠原代心肌细胞黏附铺展于金晶体电极表面后,加入不同浓度的正性肌力药物异丙肾上腺素与负性肌力药物维拉帕米,监测QCM频率(F)以及动态电阻(R)的实时变化,采用粘弹性指数CVI(CVI=ΔR/ΔF)表征细胞的粘弹性变化,并通过光学显微镜观察药物作用下细胞的形态变化。结果表明:随着异丙肾上腺素浓度的增加,药物引起的QCM频率下降、电阻增加与CVI增大的幅度均增大;光学显微镜下观测到细胞收缩,符合CVI变大、细胞变硬的响应;随着维拉帕米浓度的增加,药物引起的QCM频率升高、电阻下降与CVI减小的幅度均增大;光学显微镜下观测到细胞舒张,符合CVI变小、细胞变软的响应。说明QCM与原代心肌细胞结合作为心血管药物筛选细胞模型与工具有很大的应用前景。

脑动脉粥样硬化患者红细胞膜粘弹性与宏观血液流变学相关性的初步研究

本文采用微管吸吮技术研究了脑动脉粥样硬化患者与正常健康人红细胞在吸进微管的变形过程，采用二维Ｋｅｌｉｖｉｎ模型拟合实验结果，定量地比较患者与正常人红细胞膜的粘弹性，并采用新定义的对数还原复粘度法运用Ｌｏｗｓｈｅａｒ３０等仪器对患者与正常人宏观血液流变学参量之间相关性进行了研究，结果表明：脑动脉粥样硬化患者红细胞弹性系数（（５．７４±１．７７）×１０－３ｄｙｎ／ｃｍ）略高于正常人（（５．３６±０５６）×１０－３ｄｙｎ／ｃｍ），而粘性系数（（４．１１±１．６８）×１０－４ｄｙｎ·ｓ／ｃｍ）明显高于正常（（１．９１±０．９６）×１０－４ｄｙｎ·ｓ／ｃｍ），全血粘度及对数还原粘弹性Ｌ１及Ｌ２在由低到高各剪切率下，患者明显高于正常人。红细胞粘滞性增大，变形能力降低，这正是宏观血液流变学指标增高主要原因之一，也可能是造成患者脑硬塞的主要原因。

Mechanical properties of hepatocellular carcinoma cells

AIM: To study the viscoelastic properties of human hepatocytes and hepatocellular carcinoma (HCC) cells under cytoskeletal perturbation, and to further to study the viscoelastic properties and the adhesive properties of mouse hepatoma cells (HTC) in different cell cycle. METHODS: Micropipette aspiration technique was adopted to measure viscoelastic coefficients and adhesion force to collagen coated surface of the cells. Three kinds of cytoskeleton perturbing agents, colchicines (Col), cytochalasin D (CD) and vinblastine (VBL), were used to treat HCC cells and hepatocytes and the effects of these treatment on cell viscoelastic coefficients were investigated. The experimental results were analyzed with a three-element standard linear solid. Further, the viscoelastic properties of HTC cells and the adhesion force of different cycle HTC cells were also investigated. The synchronous G(1) and S phase cells were achieved through thymine-2-desoryriboside and colchicines sequential blockage method and thymine-2-desoryriboside blockage method respectively. RESULTS: The elastic coefficients, but not viscous coefficient of HCC cells (K(1)=103.6+/-12.6N.m(-2), K(2)=42.5 +/ 10.4N.m(-2), mu=4.5 +/- 1.9Pa.s), were significantly higher than the corresponding value for hepatocytes (K(1)=87.5 +/- 12.1N.m(-2), K(2)=33.3+/-10.3N.m(-2), mu=5.9+/-3.0Pa.s, P【0.01). Upon treatment with CD, the viscoelastic coefficients of both hepatocytes and HCC cells decreased consistently, with magnitudes for the decrease in elastic coefficients of HCC cells (K(1): 68.7 N.m(-2) to 81.7N.m(-2), 66.3% to 78.9%; K(2): 34.5N.m(-2) to 37.1N.m(-2), 81.2% to 87.3%, P【0.001) larger than those for normal hepatocytes (K(1): 42.6N.m(-2) to 49.8N.m(-2), 48.7% to 56.9%; K(2): 17.2N.m(-2) to 20.4N.m(-2), 51.7% to 61.3%, P【0.001). There was a little decrease in the viscous coefficient of HCC cells (2.0 to 3.4Pa.s, 44.4 to 75.6%, P【0.001) than that for hepatocytes (3.0 to 3.9Pa.s, 50.8 to 66.1% P【0.001). Upon treatment with Col and VBL, the elastic coefficients of hepatocytes generally increased or tended to increase while those of HCC cells decreased. HTC cells with 72.1% of G(1) phase and 98.9% of S phase were achieved and high K(1), K(2) value and low mu value were the general characteristics of HTC cells. G(1) phase cells had higher K(1) value and lower mu value than S phase cells had, and G(1) phase HTC cells had stronger adhesive forces ((275.9 +/- 232.8) x 10(-10)N) than S phase cells ((161.2 +/- 120.4) x 10(-10)N, P【0.001). CONCLUSION: The difference in both the pattern and the magnitude of the effect of cytoskeletal perturbing agent on the viscoelastic properties between HCC cells and hepatocytes may reflect differences in the state of the cytoskeleton structure and function and in the sensitivity to perturbing agent treatment between these two types of cells. Change in the viscoelastic properties of cancer cells may affect significantly tumor cell invasion and metastasis as well as interactions between tumor cells and their micro-mechanical environments.

Atomic force microscopy studies on cellular elastic and viscoelastic properties

In this work, a method based on atomic force microscopy (AFM) approach-reside-retract experiments was established to simultaneously quantify the elastic and viscoelastic properties of single cells. First, the elastic and viscoelastic properties of normal breast cells and cancerous breast cells were measured, showing significant differences in Young’s modulus and relaxation times between normal and cancerous breast cells. Remarkable differences in cellular topography between normal and cancerous breast cells were also revealed by AFM imaging. Next, the elastic and viscoelasitc properties of three other types of cell lines and primary normal B lymphocytes were measured; results demonstrated the potential of cellular viscoelastic properties in complementing cellular Young’s modulus for discerning different states of cells. This research provides a novel way to quantify the mechanical properties of cells by AFM, which allows investigation of the biomechanical behaviors of single cells from multiple aspects.