软骨多糖诱导小鼠S180细胞凋亡的作用机制

目的研究软骨多糖对S180荷瘤小鼠的作用,并探讨其抑瘤作用机制。方法采用小鼠肉瘤S180细胞建立动物腹水瘤模型,通过腹腔注射软骨多糖进行治疗,治疗期间抽取腹水瘤细胞进行细胞生物学分析。通过HE染色,流式细胞术、TUNEL法检测细胞形态学方面、细胞周期及凋亡率的变化情况;通过免疫荧光方法检测Fas、增殖细胞核抗原(PCNA)的表达情况。结果软骨多糖可以明显提高S180荷瘤小鼠的生存率,细胞形态学观察可见细胞出现细胞质浓缩、核固缩及凋亡小体等现象。软骨多糖作用后的S180细胞,其细胞周期被阻遏于G2/M期,Fas蛋白的表达水平于给药24 h后升高,增殖细胞核抗原PCNA表达下降。结论软骨多糖可能通过影响肿瘤细胞周期和Fas、PCNA蛋白的表达来诱导S180细胞凋亡,并显著抑制肿瘤细胞的生长,延长S180荷瘤小鼠的生存时间,研究证实动物软骨多糖具有潜在的药用价值。     

盐酸洛拉曲克在体内、外对S-180细胞株的抗增殖作用

研究合合成新型胸苷合成酶抑制剂盐酸洛拉曲克在体内、外对S－180细胞株及正常人胚肾HEK293细胞的抗增殖作用;使用MTT法测定抑制率,以提高荷腹水瘤小鼠存活时间及荷实体瘤瘤重减轻情况为指标考察盐酸洛拉曲克对S－180所致肿瘤的治疗作用。结果表明：在体外盐酸洛拉曲克对S－180肿瘤细胞株有较强的细胞毒作用,对正常细胞HEK293抑制作用较弱（P＜0.05）;体内实验显示盐酸洛拉曲克可明显提高荷腹水瘤小鼠的存活时间,减轻荷实体瘤小鼠的瘤重,疗效与氟尿嘧啶（10mg/kg）相当（P＞0.05）或更好（P＜0.05）。可见,盐酸洛拉曲克在体内、外对S－180肿瘤细胞有显著的抗增殖作用,在体外对肿瘤细胞 具有选择性的抗增殖作用。     

软骨多糖对荷瘤小鼠免疫功能影响的初步研究

目的研究软骨多糖对荷瘤小鼠的作用,并探讨其对免疫功能的影响。方法采用小鼠肉瘤S180细胞建立动物腹水瘤模型,然后随机将小鼠分为生理盐水对照组和软骨多糖给药组,连续腹腔注射生理盐水或软骨多糖,分别测量ConA和LPS刺激下小鼠脾细胞淋巴增殖情况、外周血NK细胞的活性,及外周血单个核细胞E花环形成率。结果软骨多糖能刺激淋巴细胞增殖,明显提高NK细胞的活性,提高E花环形成率。结论软骨多糖能通过增强S180荷瘤小鼠的免疫功能而抑制肿瘤的生长。     

细胞生物学实验课教学改革初探

实验教学是细胞生物学课程学习的重要组成部分,本文结合当前国内高校细胞生物学实验课的教学模式,分析了细胞生物学实验课教学过程中存在的一些主要问题,从实验内容的选择、教学方法和考核评价体系等方面提出了实验课教学改革的几点建议,为增强生物专业学生的实验课能力提供方案。     

细胞生物学实验课教学改革初探

实验教学是细胞生物学课程学习的重要组成部分,本文结合当前国内高校细胞生物学实验课的教学模式,分析了细胞生物学实验课微学过程中存在的一些主要问题,从实验内容的选择、教学方法和考核评价体系等方面提出了实验课教学改革的几点建议,为增强生物专业学生的实验课能力提供方案。     

小鼠肿瘤细胞染色体核仁组织者的细胞化学观察

本文报道用Ag-AS染色技术对几种小鼠肿瘤细胞(Ehrlich腹水瘤,肉瘤180,淋巴瘤1号)核仁组织者(NORs)的观察结果,发现肿瘤细胞的NORs即18 S+28 S rDNA或核糖体基因的位置和大小与正常细胞的不同。正常小鼠细胞有3—6条染色体带有银染色的核仁组织者(Ag-NORs),分布位置都紧靠在着丝点下方;而三种小鼠肿瘤细胞都有一个中等大小的近端着丝点染色体,其Ag-NOR的位置移至长臂的中部。小鼠淋巴瘤1号     

细胞大小测定及细胞计数实验的教法改进及注意事项

细胞大小测定及细胞计数实验是细胞生物学的基础实验,在实验过程中经常出现仪器损耗、学生对血球计数板了解不深入的问题。为了解决这些问题,通过长期的实验教学,从实验前、中、后3个阶段提出改进方法,并对实验过程中的注意事项进行了阐述,达到充分利用实验仪器资源,提高学生学习主动性的目的。     

一种新的制备多克隆抗体的方法

用亲和色谱纯化的rhEPO作为抗原免疫KM小鼠和Bal b/c小鼠,然后腹腔注射S180细胞或SP2/0细胞。S180细胞可刺激KM小鼠产生腹水,腹水量大,抗体效价高。而直接注射SP2/0细胞既未能使KM小鼠产生腹水,也未能使Bal b/c小鼠产生腹水。实验提供了一种新的简便、快速、经济地制备高效价多克隆抗体的方法。     

细胞生物学实验教学改革探索

细胞生物学是当代生命科学的基础学科,也是发展迅猛的前沿学科之一。为了促进学生的实践动手能力和实践创新能力,配合细胞生物学理论课教学改革的不断深化,细胞生物学实验教学改革已是大势所趋。该教研组经过多年的探索,从教学体系、教学内容和教学手段等方面对细胞生物学实验教学进行了改革,旨在充分调动学生学习的主动性和积极性,培养学生的独立思考、综合运用知识和科研创新的能力。     

超声激活血卟啉对S180细胞杀伤作用及形态学研究

利用频率为1.1 MHz及不同强度超声激活血卟啉, 对小鼠腹水型S180肿瘤细胞和诱发的在体肉瘤进行不同的实验处理. 通过光镜、电子显微镜技术、荧光标记、细胞化学等方法, 探讨超声激活血卟啉对S180细胞杀伤作用及形态学变化. 研究表明: 超声激活血卟啉对腹水型S180肿瘤细胞显微结构及表面超微结构破坏随声照强度增加而加剧; 声照后细胞显微结构、超微结构、酶活性的变化及DNA的降解丢失是癌细胞生长抑制、死亡的重要因素, 实验中发现的一定超声强度激活血卟啉可诱导癌细胞凋亡现象提示声动力疗法有促细胞死亡和诱导细胞凋亡两种模式. 通过细胞形态学变化, 以及辐照处理后细胞存在的由原发性损伤到继发性损伤直至死亡的动态变化过程, 探讨超声激活血卟啉对S180细胞杀伤作用, 以期为声动力疗法杀伤机制的研究积累资料.     

激光联合环磷酰胺对荷瘤鼠脾重和外周免疫细胞数量影响的试验研究

目的:试验以小鼠S180腹水瘤为动物实验模型,对He-Ne激光照射与化疗药物联合应用对S180荷瘤鼠脾脏重量和免疫细胞数量影响作了系统性研究。方法:应用11.00J/cm2,14.67J/cm2和22.00J/cm2三种剂量He-Ne激光照射荷瘤鼠内眼角,同时联合应用化疗药物环磷酰胺,以观察其对荷瘤鼠脾脏重量和免疫细胞数量的影响。结果:健康小鼠脾脏随日龄增加呈持续上升趋势,肿瘤对照组及CYT对照组在第4d.显著升高。此后便持续下降,尤其是CYT对照组,第8d.荷瘤鼠脾脏萎缩极为严重。而CYT/He-Ne激光联合应用组小鼠脾脏在第4d.轻微下降后,便呈持续上升趋势;CYT对外周血白细胞总数、淋巴细胞数和中性幼稚细胞数有着显著的抑制作用(P<0.01,P<0.05)。CYT/He-Ne激光联合应用组白细胞总数和中性幼稚细胞数的变化较单纯CYT应用组为轻微。结论:He-Ne激光对CYT及肿瘤本身所致免疫抑制具有明显的改善效应。     

Ribosomal RNA on the surfaces of nonleukemic mouse ascites tumor cells

RNAs on the cell surfaces of two nonleukemic and two leukemic strains of mouse ascites tumor cells were studied by fractionating the RNAs released from the cell surface by gentle pronase treatment after sucrose density gradient centrifugation, by indirect membrane immunofluorescence that used anti-RNA antibody and by cell electrophoresis. RNA was extracted from the cell supernatants of Ehrlich ascites tumor and sarcoma 180 cells (nonleukemic) that had been treated or not treated with pronase (1 microgram/ml, 37 degrees C, 20 min) followed by sucrose density gradient centrifugation. It was clearly demonstrated that the amounts of ribosomal RNA (18S and 28S) released after pronase treatment were approximately 80% greater than that of nonpronase-treated cells. Ehrlich ascites tumor cells that had been treated with actinomycin D (100 micrograms/kg body weight of mouse, 16 h) in vivo released an amount of ribosomal RNA after pronase treatment only 20% greater than the value for untreated cells. Actinomycin D treatment greatly reduced both the cell surface negative charge and the cell surface immunofluorescence when rabbit anti-RNA antibody was used. Under the same experimental conditions with actinomycin D, only ribosomal RNA synthesis showed preferential inhibition, not the syntheses of poly A-containing messenger RNA, 4S or other small-size RNAs. In contrast, L1210 and C1498 cells (leukemic) showed no change in the amounts of ribosomal RNA released after pronase treatment. L1210 cells also showed no change in the surface negative charge after being treated with actinomycin D.(ABSTRACT TRUNCATED AT 250 WORDS)     

Anthracycline resistance and consequences of the in situ-in vitro transfer

Adriamycin-resistant and normal cells of the sarcoma 180 of the mouse undergo qualitatively different deflections from the in situ state when prepared for an experiment. Resistant cells perform a fast reactive decline in the proliferative activity. They are capable of quiescence as defined by the time needed for the induction of the proliferation. Sensitive cells seem to be unable to quiesce and are only slowed down. These facts must be taken into account in interpretation of similar results. Differences in experiments need not necessarily imply differences in situ. Such in vitro appearing differences between sensitive and adriamycin-resistant cells of the murine sarcoma 180 include the retention of the mitochondria-specific stain rhodamine 123 and the uptake of anthracyclines, both being reduced in resistant cells. After labeling sensitive cells with thymidine in vivo and sorting them according to their rhodamine 123-derived fluorescence, the label was only found in the major, highly fluorescing fraction. A small low-fluorescing fraction remained unlabeled. We were able to demonstrate similar results with labeled anthracyclines applied to both the sensitive and the resistant cells in a short period between the removal of the cells from the ascites and the cell sorting. The adriamycin resistance seems to be joined with the ability of the cells to reduce their proliferative activity following changes to unfavorable conditions in vitro. Quiescent cells of the resistant line demonstrate the "anthracycline pump." Substances which are known to increase the sensitivity of anthracycline-resistant cells (TWEEN, verapamil) also shift the cells from low to high rhodamine 123-fluorescence.     

De novo synthesis of NGF subunits in S-180 mouse sarcoma cell line

It is an accepted hypothesis that the nerve growth factor protein (NGF) plays an important role in the development of vertebrate sympathetic and sensory ganglia and has effects on some central neurons. The best known NGF species is that isolated from the mouse submaxillary gland, MSG-NGF. MSG-NGF can be isolated as a subunit containing protein, 7S-NGF, made up of three dissimilar subunits called alpha-, beta-, and gamma-NGF. Beta-NGF is the biologically active subunit and its synthesis in vivo and in vitro has been demonstrated. Less is known about the synthesis of the alpha- and gamma-NGF or the assembly of the subunits into the 7S complex. In order to develop a clonal model system for the study of NGF synthesis, processing and secretion, affinity chromatography techniques were applied to cell extracts of S180 mouse sarcoma, a cell line known to synthesize NGF. After incubating S180 cells in³⁵S-Methionine, cell extracts were exposed to antibody directed against alpha-NGF, gamma-NGF or beta-NGF covalently bound to Sepharose beads in order to elute and characterize the desired NGF subunits. Parallel experiments using immunoabsorbed [³⁵S]Methionine-beta-NGF were carried out in the presence or absence of excess NGF, in order to demonstrate the specificity of this procedure. Affinity chromatography with a substrate analogue to arginine ester bound to Sepharose beads was also used to isolate de novo synthesized gamma-NGF. We were able to show that the S180 line synthesized alpha-, beta-, and gamma-NGF indistiguishable from alpha-, beta-, and gamma-NGF isolated from mouse submaxillary gland in terms of antigenic and physicochemical properties, and biological and enzymatic activities. These results are consistent with the hypothesis that NGF is synthesized, assembled and secreted by a single cell type.Special Issue dedicated to Dr. E. M. Shooter and Dr. S. Varon.     

用尖吻蝮蛇毒、蝗蛇毒及抗蛇毒血清处理小鼠S_(180)、EAC腹水癌的初步观察

本文报道了尖吻蝮蛇毒、蝮蛇毒及抗蛇毒血清能使接种S_(180)、EAC腹水癌的小鼠明显延长存活时间、降低接种率,但不能完全阻止痛细胞生长。体外具有较明显的导致,菡细胞肿胀、膜破裂,核纤维化,坏死等。从腹水酶活力测定及抗血清初步研究结果表明,癌细胞病变中产生的某些抗原物质可能与蛇毒中的酶和毒蛋白相近。因此注射蛇毒后可在体内产生相关抗体,中和癌细胞产生的毒素以达到治疗目的。     

金环蛇蛇毒对S180,EAC腹水癌细胞的细胞毒性作用

目的　探讨金环蛇毒对 S1 80 ,EAC腹水癌细胞的细胞毒性作用。　方法　采用小白鼠腹腔和皮下接种 S1 80 ,EAC腹水癌细胞造成小白鼠腹水模型后腹腔注射金环蛇毒。　结果　腹腔注射金环蛇毒 ,能抑制肿瘤细胞的生长 ,降低接种率。但不能完全控制腹水和癌细胞的生长。体外试验表明有明显的细胞毒作用。台酚蓝染色镜检可见死细胞显著增加 ,腹水图片检查给药后细胞膜破裂 ,纤维化坏死明显。　结论　能延长小白鼠存活时间。     

C_(60)对小鼠S_(180)肉瘤光动力学作用模型的建立

为验证Ｃ６０对活体肿瘤的光动力学损伤作用,我们从两方面进行实验：Ｃ６０对荷瘤小鼠的Ｓ１８０实体瘤的光动力学杀伤作用和Ｃ８０对离体Ｓ１８０肿瘤细胞的光动力学杀伤作用,在小鼠的瘤体上注射Ｃ８０光敏剂,在５１１ｎｍ、５７８ｎｍ混合黄录色激光照射正派主发Ｃ６０,产生大量的单线态氧,杀伤活性肿瘤。激光光强为５００ｍＷ,Ｃ６０浓度为３０μｇ／ｍｌ时,荷瘤小鼠寿命平均延长５天,瘤径减小１ｃｍ,瘤重减轻０．８克,     

Superoxide dismutase induces G1‐phase cell cycle arrest by down‐regulated expression of Cdk‐2 and cyclin‐E in murine sarcoma S180 tumor cells

As an efficient reactive oxygen species–scavenging enzyme, superoxide dismutase (SOD) has been shown to inhibit tumor growth and interfere with motility and invasiveness of cancer cells. In this study, the molecular mechanisms of cell cycle arrest when S180 tumor cells were exposed to high levels of SOD were investigated. Here, both murine sarcoma S180 tumor cells and NIH‐3T3 mouse fibroblasts were respectively treated with varying concentrations of Cu/Zn‐SOD for 24, 48 and 72 h to determine optimal dose of SOD, which was a concentration of 800 U/ml SOD for 48 h. It is found that SOD induced S180 cell cycle arrest at G1‐phase with decreasing level of superoxide production, whereas SOD had less effect on proliferation of NIH‐3T3 cells. Moreover, the expression rate of Proliferating Cell Nuclear Antigen (PCNA) in S180 tumor cells was suppressed after SOD treatment, which indicated the inhibition of DNA synthesis in S180 cells. Besides, there were significant down‐regulations of cyclin‐E and Cdk‐2 in S180 cells after SOD treatment, which contributed to the blockage of G1/S transition in S180 cell cycle. Together, our data confirmed that SOD could notably inhibit proliferation of S180 tumor cell and induce cell cycle arrest at G1‐phase by down‐regulating expressions of cyclin‐E and Cdk‐2. Copyright © 2012 John Wiley & Sons, Ltd.     

cDNAs of cell adhesion molecules of different specificity induce changes in cell shape and border formation in cultured S180 cells

The liver cell adhesion molecule (L-CAM) and N-cadherin or adherens junction-specific CAM (A-CAM) are structurally related cell surface glycoproteins that mediate calcium-dependent adhesion in different tissues. We have isolated and characterized a full-length cDNA clone for chicken N-cadherin and used this clone to transfect S180 mouse sarcoma cells that do not normally express N-cadherin. The transfected cells (S180cadN cells) expressed N-cadherin on their surfaces and resembled S180 cells transfected with L-CAM (S180L cells) in that at confluence they formed an epithelioid sheet and displayed a large increase in the number of adherens and gap junctions. In addition, N-cadherin in S180cadN cells, like L-CAM in S180L cells, accumulated at cellular boundaries where it was colocalized with cortical actin. In S180L cells and S180cadN cells, L-CAM and N-cadherin were seen at sites of adherens junctions but were not restricted to these areas. Adhesion mediated by either CAM was inhibited by treatment with cytochalasin D that disrupted the actin network of the transfected cells. Despite their known structural similarities, there was no evidence of interaction between L-CAM and N-cadherin. Doubly transfected cells (S180L/cadN) also formed epithelioid sheets. In these cells, both N-cadherin and L-CAM colocalized at areas of cell contact and the presence of antibodies to both CAMs was required to disrupt the sheets of cells. Studies using divalent antibodies to localize each CAM at the cell surface or to perturb their distributions indicated that in the same cell there were no interactions between L-CAM and N-cadherin molecules. These data suggest that the Ca(++)-dependent CAMs are likely to play a critical role in the maintenance of epithelial structures and support a model for the segregation of CAM mediated binding. They also provide further support for the so-called precedence hypothesis that proposes that expression and homophilic binding of CAMs are necessary for formation of junctional structures in epithelia.     

橙盖鹅膏多糖(AC-1)体外免疫调节活性、细胞毒性和抗肿瘤活性的研究

为研究橙盖鹅膏多糖的体外免疫调节活性、细胞毒性和抗肿瘤活性,本研究运用细胞学技术探究AC-1对T细胞、B细胞和RAW264.7三种免疫细胞的体外免疫调节活性;研究AC-1对小鼠成纤维细胞(L929)的细胞毒性以及对小鼠胃癌细胞(MFC)、小鼠肉瘤细胞(S180)的体外抗肿瘤活性。结果表明,AC-1能在体外显著刺激三种免疫细胞的增殖及RAW264.7细胞的吞噬,表明AC-1在增强机体免疫方面具有重要作用,进一步研究发现AC-1主要通过显著促进IgE和IgG的分泌来增强体液免疫。在浓度为5~20μg/mL时AC-1对L929细胞无显著的促进及抑制作用,说明AC-1对正常细胞无毒性;在浓度为10~20μg/mL时AC-1能显著抑制小鼠胃癌细胞(MFC)的生长,但对小鼠肉瘤细胞(S180)的抑制效果较弱,说明AC-1在体外能够直接抑制肿瘤细胞的生长,但对不同的肿瘤细胞具有不同的抑制效果。综上所述,橙盖鹅膏多糖(AC-1)在体外具有良好的免疫调节活性、抗肿瘤活性,但对不同的肿瘤细胞具有不同的抑制效果并且对正常细胞无毒性。