Cl-在鼻咽癌细胞调节性容积回缩中的作用

目的:研究鼻咽癌细胞的调节性容积回缩(RVD)及Cl^-在其中所起的作用。方法:活细胞图像分析低分化鼻咽癌细胞(CNE-2Z)在细胞外低渗刺激时的细胞容积改变,用离子置换法和通道阻断法阐明Cl^-在RVD中所起的作用。结果:细胞外低渗刺激使细胞膨胀并诱发RVD。在160-230mOsmol/L范围内,RVD与细胞膨胀程度正相关。用葡萄糖酸取代灌流液Cl^-时RVD增强,细胞内Cl^-耗竭时RVD消失。氯通道阻断剂阻抑RVD,但不同的阻断剂对RVD的阻抑率有明显差异。结论:Cl^-是CNE-2Z细胞RVD的关键离子,Cl^-通道激活,Cl^-外流是RVD的主要机制。     

辛伐他汀抑制胎牛血清及PDGF-BB诱导的血管平滑肌细胞增殖及对抑癌基因PTEN表达的影响

目的:观察辛伐他汀(simvastatin)对血清及血小板源生长因子(PDGF-BB)诱导的血管平滑肌细胞(VSMCs)增殖的抑制作用及simvastatin对细胞周期和G₁/S周期转换重要调节基因PTEN蛋白表达的影响。方法:[³H]-胸腺嘧啶核苷酸掺入测定VSMCsDNA合成,流式细胞仪检测细胞周期情况,Western印迹杂交法检测PTEN蛋白表达。结果:Simvastatin以剂量依赖关系抑制血清及PDGF-BB诱导的VSMCs[³H]-胸腺嘧啶核苷酸掺入,使G₀/G₁期细胞比例明显增多,S期细胞比例显著减少,并上调PTEN蛋白表达,而3羟3甲戊二酰辅酶A代谢产物甲羟戊酸能抑制simvastatin上调PTEN的表达。结论:Simvastatin抑制血清及PDGF-BB诱导的VSMCs增殖阻滞细胞周期可能与上调PTEN表达有关,且simvastatin调节PTEN的表达可能通过抑制甲羟戊酸的合成而实现。     

NS-398对肝癌细胞HepG2增殖和凋亡的影响

目的:探讨选择性环氧合酶II(COX-2)抑制剂NS-398对人肝癌细胞HepG2增殖和凋亡的影响。方法: 应用MTT法研究不同浓度NS-398对HepG2细胞增殖的影响,DNA梯状电泳(DNA ladder)检测凋亡的发生,流式细胞仪检测细胞周期的改变及凋亡百分率的变化,竞争性RT-PCR检测环氧合酶II COX-2 mRNA及抑凋亡基因bcl-2 mRNA表达的改变。结果: NS-398呈剂量依赖性抑制HepG2细胞增殖,并诱导其凋亡,细胞周期分析表明随着浓度增大S期细胞明显减少,有G₀/G₁期细胞累积现象,并伴有Bcl-2 mRNA表达的下调,而对COX-2 mRNA表达改变无明显影响,且COX-2表达改变与NS-398引起的HepG2细胞的增殖和凋亡均无相关性(相关系数分别为:r=0.056,P>0.05和r=0.119,P>0.05)。结论: NS-398能明显抑制HepG2细胞增殖并诱导其凋亡,与细胞G₀/G₁期阻滞以及bcl-2基因表达下调有关,而非依赖于抑制COX-2基因的表达。     

丹参酮ⅡA对U251胶质瘤细胞的增殖抑制和原癌基因表达的影响

目的:探讨丹参酮ⅡA对人胶质瘤细胞株U251的增殖抑制作用及其作用机理。方法:应用MTT比色法检测不同浓度丹参酮ⅡA对U251细胞的增殖抑制,应用流式细胞仪观察丹参酮ⅡA对U251细胞周期的影响,应用RT-PCR观察相关基因C-myc表达的变化。结果:丹参酮ⅡA对U251细胞增殖抑制作用呈剂量依赖性,当浓度为0.10 g·L^-1时,对U251细胞增殖的抑制率达到(53.7±6.0)%。流式细胞仪分析显示,用0.10 g·L^-1丹参酮ⅡA培养3 d,细胞周期中G₀/G₁期所占比例增高,S期占细胞周期的比例降低。RT-PCR结果显示,随着丹参酮ⅡA作用剂量的增加,c-myc基因的表达被明显抑制。结论:丹参酮ⅡA对人胶质瘤细胞系U251的增殖具有明显的抑制作用,并对原癌基因c-myc的表达具有抑制作用。     

c-sis反义寡核苷酸对肺血管平滑肌细胞增殖和细胞周期的影响

目的:探讨从血小板源生长因子-B链(PDGF-B)的基因水平阻断对肺血管平滑肌细胞增殖的影响。方法:给肺动脉平滑肌细胞(VSMC)分别施加不同剂量的c-sis癌基因反义寡核苷酸(ASON), 通过四甲基偶氮唑蓝(MTT)法观察细胞不同时相的增殖曲线;采用流式细胞术观察不同干预条件下VSMC细胞周期的变化。结果:中、大浓度ASON对细胞增殖有明显抑制作用。在中浓度的ASON作用下, 加入PDGF-BB能促进VSMC的增殖。ASON干预前后, VSMC增殖周期中各期细胞构成发生显著的变化, 以G₀＋G₁期细胞增多、S+G₂＋M期细胞减少为特征。各组的G₀＋G₁期细胞均显著多于对照组(P＜0.05)。小剂量与中剂量、中剂量与大剂量间G₀＋G₁期细胞有显著差异(P＜0.05)。结论:中、大剂量ASON能明显抑制肺VSMC的增殖, 可使G₀＋G₁期细胞数明显增多, 且与ASON有剂量依赖关系。     

人参单体皂甙Rh2对前列腺癌细胞株PC-3M细胞移行周期的影响

目的:探讨Rh₂对体外培养的PC-3M细胞移行周期的影响。方法:应用[³H]-TdR掺入法和流式细胞仪测定DNA含量及进行细胞周期的分析。结果:PC-3M细胞的[³H]-TdR掺入量明显少于对照组(P＜0.01);流式细胞仪测定G₁期细胞数百分比明显高于对照组。结论:Rh₂可使PC-3M细胞增殖周期阻滞在G₁期,肿瘤细胞增殖减慢。     

食药用真菌粗提物促血管内皮活性的筛选及对高糖损伤内皮的保护作用

目的: 观察古尼虫草、花脸香菇、冬虫夏草和巴西革耳4种食药用真菌粗提物对高糖诱导的人脐静脉内皮细胞(HUVECs)增殖活性的作用,筛选具有保护内皮细胞活性的有效菌菇并探讨其对HUVECs细胞周期及活性氧簇(ROS)水平的影响。方法: 体外培养HUVECs,分为对照组 (M199正常培养液)、高糖组(33 mmol/L高糖培养液)、4种食药用真菌(古尼虫草、花脸香菇、冬虫夏草和巴西革耳)粗提物干预组 (每种粗提物以终浓度12.5、25、50、100 mg/L+高糖培养液),以四甲基偶氮唑蓝(MTT)比色法检测细胞增殖活性,流式细胞术检测细胞周期及细胞内ROS水平。结果: 与对照组相比,高糖组HUVECs 的MTT吸光度值显著下降,G₀/G₁期的细胞百分率明显增加,S+G₂/M期的细胞百分率明显下降,且ROS水平亦增加(均P<0.05)。与高糖组相比,花脸香菇和巴西革耳粗提物干预降低了细胞吸光度值(P<0.05),且抑制效应呈剂量依赖性增强,而古尼虫草粗提物对其无影响。冬虫夏草粗提物(12.5~50 mg/L)显著提高了HUVECs的增殖活性(P<0,05),减少了G₀/G₁期细胞百分率,提高了S+G₂/M期的细胞百分率(P<0.05),降低了细胞内ROS水平(均P<0.05),且效应呈剂量依赖性增强。结论: 4种食药用真菌粗提物经MTT比色法筛选后,仅冬虫夏草粗提物对高糖诱导的HUVECs有显著保护作用,其可能通过促进内皮细胞增殖、进入细胞周期及降低细胞内活性氧水平途径。     

1,25-二羟维生素D3抑制被动致敏人气道平滑肌细胞的增殖

目的: 检测1,25-二羟维生素D₃ 对被动致敏的人气道平滑肌细胞(HASMCs)增殖的调节作用,探讨其对哮喘气道重塑的可能作用。方法: 离体培养HASMCs,并将细胞分为对照组、哮喘组及1,25-(OH)₂D₃组。MTT法检测细胞增殖活力并确定1,25-(OH)₂D₃最佳作用浓度;用最佳作用浓度刺激HASMCs 48 h后以流式细胞术测定细胞周期,免疫细胞化学染色(SABC法)检测增殖细胞核抗原(PCNA)的表达。结果: 1,25-(OH)₂D₃在10^-9-10^-7 mol/L范围内能显著抑制哮喘血清被动致敏的HASMCs增殖(P<0.05),且10^-7 mol/L时抑制作用最强;流式细胞术检测显示这一最佳作用浓度的1,25-(OH)₂D₃能显著抑制被动致敏HASMCs由G₀/G₁期向S期的转化;此外,1,25-(OH)₂D₃能显著抑制被动致敏HASMCs中PCNA的表达。结论: 1,25-(OH)₂D₃能抑制哮喘血清被动致敏的HASMCs增殖,有助于哮喘气道重塑的防治。     

反义多重耐药性基因抑制眼睫状体非色素上皮细胞容积激活性氯电流

目的:研究多重耐药性(MDR1)基因及其产物P糖蛋白(P-gp)与容积激活性Cl^-电流的关系。方法:用膜片钳全细胞记录技术记录牛眼睫状体非色素上皮(NPCE)细胞容积激活性Cl^-电流,反义寡核苷酸阻抑细胞MDR1基因表达,在激光共聚焦显微镜下检测细胞P-gp免疫荧光。结果:P-gp免疫荧光与人源反义MDR1呈剂量依赖性减弱关系,容积激活性Cl^-电流被人源反义MDR1特异性地部分阻抑,电流潜伏期延长,峰电流值减少,电流抑制率与人源反义MDR1呈现剂量依赖性增强关系,(r=0.99, P＜0.01)。 P-gp表达抑制率和容积激活性Cl^-电流抑制率高度正相关(r=0.99, P＜0.01)。结论: MDR1基因及其产物P-gp参与NPCE细胞容积激活性Cl^-电流的形成。     

Bmi-1 基因过表达对人正常胃黏膜上皮细胞株GES-1增殖的影响

目的: 探讨原癌基因B淋巴瘤莫洛尼鼠白血病病毒插入区1( Bmi-1 )过表达对人正常胃黏膜上皮细胞株GES-1增殖的影响。方法: 采用逆转录病毒介导转染方法将携带原癌基因 Bmi-1 的质粒或空质粒稳定转染GES-1细胞,通过real-time PCR及Western blotting在mRNA及蛋白水平鉴定转染效果。流式细胞术检测过表达 Bmi-1 对GES-1细胞周期的影响。应用CCK-8(Cell Counting Kit-8)试剂盒检测稳定转染 Bmi-1 对GES-1细胞增殖的影响。结果: Real-time PCR及Western blotting结果均表明成功建立稳定转染 Bmi-1 基因的GES-1细胞株。流式细胞术结果表明,过表达 Bmi-1 基因使GES-1细胞G₀/G₁期减少,G₂/M期和S期细胞增多。生长曲线显示,过表达 Bmi-1 基因使GES-1细胞增殖速度明显提高。结论: 过表达 Bmi-1 基因能调控GES-1细胞的细胞周期,促进GES-1细胞的增殖。     

Activation of basolateral Cl− channels in the rat colonic epithelium during regulatory volume decrease

Exposure to a hypotonic medium caused an increase in the diameter of isolated crypts from the rat colon. The increase in cell volume was only transient and lasted about 7 min. Despite of the continuous presence of the hypotonic medium, cell volume decreased again. This regulatory volume decrease (RVD) was inhibited by 5-nitro-2-(3-phenylpropylamino)-benzoate (NPPB), a Cl^– channel blocker, and by Ba²⁺, a K⁺ channel blocker. Cell-attached patch-clamp recordings revealed that the RVD was associated with the activation of previously silent basolateral channels. These channels were identified after excision of the patch as Cl^– channels (28 pS) and as K⁺ channels (45–60 pS). The RVD was dependent on the presence of external Ca²⁺. The phospholipase A₂ inhibitor, quinacrine, and the lipoxygenase blocker, nordihydroguaiaretic acid, inhibited RVD, while indomethacin had no effect. In Ussing chamber experiments an exposure to hypotonic media caused an initial, transient increase in tissue conductance (G _t), followed by a prolonged decrease in short-circuit current (I _sc) and the potential difference (V). The height of the electrical response was dependent on the decrease in the osmolarity in a range from 20 mosmol l^–1 to 90 mosmol l^–1. The increase in G _t was blocked by NPPB and Ba²⁺, whereas the decrease in I _sc or V was inhibited by NPPB but enhanced by Ba²⁺. This suggests that in the later phase the osmotically induced Cl^– conductance exceeds the K⁺ conductance leading to an electrogenic response, while the initial response of the RVD is an opening of Cl^– and K⁺ channels in a ratio of about 11. With respect to the inhibitory efficacy of nordihydroguaiaretic acid and the inefficacy of indomethacin, leukotrienes seem to be involved in the mediation of this response.     

Regulatory volume decrease in a renal distal tubular cell line (A6) I. Role of K+ and Cl−

Changes in volume of A6 epithelial cells were monitored by recording cell thickness (T _c). The response ofT _c to a reduction of the basolateral osmolality from 260 to 140 mosmol/kg was recorded while transepithelial Na⁺ transport was inhibited by 20 M amiloride. With Cl^–-containing bathing media, this osmotic challenge elicited a rapid rise inT _c followed by a regulatory volume decrease (RVD). Substitution of SO₄ ^2– or gluconate for Cl^– markedly reduced the RVD, whereas cells completely maintained their ability to regulate their volume after replacing Cl^– by NO₃ ^–. A conductive pathway for Cl^– excretion is suggested, which is insensitive to NPPB [5-nitro-2-(3-phenylpropylamino)benzoic acid], an inhibitor of some types of Cl^– channels. Ba²⁺ (5 or 20 mM) reduced the RVD. A more pronounced inhibition of the RVD was obtained with 500 M quinine, a potent blocker of volume-activated K⁺ channels. K⁺-induced depolarization of the basolateral membranes of tissues incubated with SO₄ ^2–-containing solutions completely abolished the RVD. Noise analysis in the presence of Ba²⁺ showed the activation of an apical K⁺ conductive pathway. These results demonstrate that cell volume regulation is controlled by processes involving Cl^– and K⁺ excretion through conductive pathways.     

Effects of chloride transport inhibitors on intestinal fluid and ion transport in vivo and in vitro

The hypothesis that intestinal fluid secretion is driven by Cl^- has been tested by investigating the effects of NPPB (5-nitro-2-(3-phenyl-propylamino)-benzoate), a blocker of Cl^- channels in nephrons, and the loop diuretic bumetanide, an inhibitor of the Na⁺, K⁺, 2 Cl^--co-transporter. Both NPPB (IC₅₀ (inhibitory concentration) ～ 100 μM) and bumetanide (IC₅₀～μM) inhibited stimulated short-circuit current (I_sc) in monolayers of a colonic cell line T84. NPPB also inhibited ^3aCl^- uptake by these cells, indicating that NPPB acts as a CI^- channel blocker in the T84 cells. NPPB (300μM) and bumetanide (10 μM) abolished both stimulated /_sc and CI^- secretion in isolated rat colonic mucosa. As judged by autoradiography, [³H]NPPB was found both in the crypts and at the surface after exposure of either side to the compound. In line with these results, NPPB and bumetanide reduced stimulated fluid secretion in everted colon sacs from the rat. In the anaesthetized rat model, neither bumetanide nor NPPB affected the net fluid transport. After luminal administration of [³H]NPPB to the rat, radioactivity was found mainly in the villus tips, whereas no labelling was found in the crypts. NPPB was bound to plasma protein (99%), and the inhibitory effects of both NPPB and bumetanide on I_sc in T84 cells and fluid secretion in the colonic sacs decreased in the presence of albumin, again indicating that the compounds might not reach the in vivo target, or that the mechanism for fluid secretion in vivo may not be explained solely by the secretion of CI^-     

Actions of the CI- channel blocker NPPB on absorptive and secretory transport processes of Na+ and CI- in rat descending colon

The effects of the Cl^- channel blocker, NPPB (5-nitro-2-(3-phenylpropylamino)-benzoate), on the transport of Na⁺ and Cl^- in the descending colon of the rat were studied in the Ussing chamber. In control tissue, NPPB administered at the mucosal side of the epithelium increased the short-circuit current (I_sc) and inhibited the unidirectional mucosa-to-serosa fluxes of Na⁺ and CI^-. In HCO^-₃-_- or Cl^--free media or in the presence of SITS (4-acetamido-4′-isothiocyanato-stilbene-2,2′-disulphonic acid), this increase in I_sc caused by mucosal NPPB was not observed. The serosal administration of NPPB was without effect. Mhdcosal NPPB (10^--⁴ mol l^-1hd decreased the forskolin-induced increase in I_sc by only about 60%. However, the activation of the serosa-to-mucosa flux of Cl^- caused by forskolin was inhibited completely. NPPB decreased the mucosa-to-serosa fluxes of Na⁺ and Cl^- reduced additionally by forskolin. Serosal NPPB decreased I_sc and J^Na_sm, but had no effect on J^Na_sm or J^Na_sm. In HCO^-₃-free buffer the increase in I_sc induced by forskolin was inhibited completely by NPPB. The inhibition of Cl^- secretion by NPPB fits well with the capacity of the drug to block Cl^- channels. For the inhibition of neutral NaCl absorption two sites of action are discussed: an interaction with the Cl^-/HCO^-₃ exchanger or an interference with the extrusion of Cl^- through the basolateral membrane.,     

Modulation of volume-sensitive Cl – channels and cell volume by actin filaments and microtubules in human cervical cancer HT-3 cells

Hypotonicity activates volume-sensitive Cl^– currents, which are implicated in the regulatory volume decrease (RVD) responses and transport of taurine in human cervical cancer HT-3 cells. In this study, the role of cytoskeleton in the regulation of volume-sensitive Cl^– channels and RVD responses in HT-3 cells was studied. Cells were incubated with various compounds, which depolymerized or polymerized cytoskeletal elements, i.e. actin filaments and microtubules. The hypotonicity-induced changes in Cl^– conductance and in cell volume were measured by whole-cell voltage clamping and cell size monitoring, respectively. Our results show that in HT-3 cells hypotonicity activated an outward rectified Cl^– current that was abrogated by Cl^– channel blockers. Cytochalasin B, an actin-depolymerizing compound, induced a substantial increase in Cl^– conductance under isotonic condition and potentiated the expression of Cl^– currents in hypotonic stress. Phorbol 12-myristate 13-acetate (PMA) significantly inhibited the cytochalasin B-induced activation of Cl^– conductance under isotonic condition. On the other hand, treatment with cytochalasin B significantly prolonged the RVD responses. Phalloidin, a stabilizer of actin polymerization, did not change the basal currents under isotonic condition, but completely abolished the increase in whole-cell Cl^– conductance elicited by hypotonicity and retarded the cell volume recovery. Colchicine, a microtubule-assembly inhibitor, had no effect on either basal Cl^– conductance or volume-sensitive Cl^– current and was unable to inhibit the RVD responses. Taxol, a microtubule-stabilizing compound, did not alter the basal Cl^– conductance, but inhibited the activation of volume-sensitive Cl^– channels as well as the process of RVD in a dose-dependent manner. These data support the notion that functional integrity of actin filaments and microtubules plays critical roles in maintaining the RVD responses and activation of Cl^– channels in human cervical cancer HT-3 cells.     

A role for the volume regulated anion channel in volume regulation in the murine CNS cell line,CAD

Aim: The role of the volume regulated anion channel (VRAC) in a model CNS neuronal cell line, CAD, was investigated. Methods: Changes in cell volume following hypotonic challenges were measured using a video-imaging technique. The effect of the Cl⁻ channel antagonists tamoxifen (10 μm ) and 4,4′-diisothiocyanatostilbene-2,2′-disulphonic acid (DIDS; 100 μm ) on regulatory volume decrease (RVD) were measured. The whole-cell voltage-clamp technique was used to characterize ICl_swell, the current underlying the VRAC. Results: Using the video-imaging technique, CAD cells were found to swell and subsequently exhibit RVD when subjected to a sustained hypotonic challenge from 300 mOsmol kg⁻¹ H₂O to 210 mOsmol kg⁻¹ H₂O. In the presence of tamoxifen (10 μm ) or DIDS (100 μm ) RVD was abolished, suggesting a role for the VRAC. A hypotonic solution (230 mOsmol kg⁻¹ H₂O) evoked ICl_swell, an outwardly rectifying current displaying time-independent activation, which reversed upon return to isotonic conditions. The reversal potential (E_rev) for ICl_swell was −14.7 ± 1.4 mV, similar to the theoretical E_rev for a selective Cl⁻ conductance. ICl_swell was inhibited in the presence of DIDS (100 μm ) and tamoxifen (10 μm ), the DIDS inhibition being voltage dependent. Conclusions: Osmotic swelling elicits an outwardly rectifying Cl⁻ conductance in CAD cells. The ICl_swell observed in these cells is similar to that observed in other cells, and is likely to provide a pathway for the loss of Cl⁻ which leads to water loss and RVD. As ischaemia, brain trauma, hypoxia and other brain pathologies can cause cell swelling, CAD cells represent a model cell line for the study of neuronal cell volume regulation.     

Dual roles of plasmalemmal chloride channels in induction of cell death

Even under anisotonic conditions, most cells can regulate their volume by mechanisms called regulatory volume decrease (RVD) and increase (RVI) after osmotic swelling or shrinkage, respectively. In contrast, the initial processes of necrosis and apoptosis are associated with persistent swelling and shrinkage. Necrotic volume increase (NVI) is initiated by uptake of osmolytes, such as Na⁺, Cl^– and lactate, under conditions of injury, hypoxia, ischaemia, acidosis or lactacidosis. Persistence of NVI is caused by dysfunction of RVD due to impairment of volume-sensitive Cl^– channels under conditions of ATP deficiency or lactacidosis. Both lactacidosis-induced RVD dysfunction and necrotic cell death are prevented by pretreatment of cells with the vacuolating cytotoxin-A (VacA) toxin protein purified from Helicobacter pylori, which forms a lactacidosis-resistant anion channel. Apoptotic volume decrease (AVD) is triggered by activation of K⁺ and Cl^– conductances following stimulation with a mitochondrion-mediated or death receptor-mediated apoptosis inducer. Apoptotic cell death can be prevented by blocking the Cl^– channels but not the K⁺-Cl^– cotransporters. Thus, the volume regulatory anion channel plays, unless impaired, a cell-rescuing role in the necrotic process by ensuring RVD after swelling induced by necrotic insults, whereas normotonic activation of the anion channel plays a cell-killing role in the apoptotic process by triggering AVD following stimulation with apoptosis inducers.     

Sodium-dependent activity of aquaporin-1 in rat glioma cells: a new mechanism of cell volume regulation

Cell volume controls many functions and is itself regulated. To study cell volume regulations, the mean volume of C6-BU-1 rat glioma cells was electronically measured under isotonic and anisotonic conditions. Two isotonic solutions were used containing either normal (solution 1) or low (solution 2) NaCl. Anisotonicity was induced by changing NaCl or sucrose concentrations in solutions 1 and 2, respectively. The cells behaved like perfect osmometers when the tonicity was increased. In contrast, just after hypotonic challenges, the cell volume was smaller than that predicted by a perfect osmometer. This deviation reveals a new mechanism, which we call the volume increase limitation (VIL). When hypotonicity was induced by decreasing NaCl, a classical slow regulatory volume decrease (RVD) was also observed in addition to VIL. The cells expressed aquaporin-1 sensitive to HgCl₂ and decreased by siRNA, which both reduced fast volume changes. In this study, we show that: (1) RVD is proportional to the change in external Cl⁻ concentration and is inhibited by Cl⁻ channel and K⁺–Cl⁻ cotransporter blockers; (2) cell swelling due to the influx of H₂O through aquaporins shows rectification with decreasing osmolarity and is sensitive to the internal Na⁺ concentration; (3) VIL is linearly related with hypotonicity and is abolished in solutions 1 and 2 by the Na⁺ ionophore monensin and in solution 1 by the Na⁺–K⁺ ATPase inhibitor ouabain. These results suggest that VIL is triggered by the decrease in internal Na⁺ caused by hyponatrema and cell swelling. In addition to RVD, VIL should protect cells during hyposmotic stress.     

188铼辐射抑制血管平滑肌细胞增殖研究

目的：探讨放射性核素¹⁸⁸铼（¹⁸⁸Re）对培养平滑肌细胞增殖的作用及其机制。方法：应用¹⁸⁸Re β射线进行培养平滑肌细胞的内辐射。通过细胞计数、再增殖试验、[³H]-TdR掺入试验、流式细胞术细胞周期分析、免疫细胞化学及细胞存活率检测等方法,观察辐射抑制平滑肌细胞增殖的有效及最佳剂量、有效抑制时间、细胞增殖率变化、细胞周期分布、细胞存活率及其相关基因表达。结果：5.2 Gy剂量辐射,50%以上平滑肌细胞增殖受抑,细胞增殖率为46%;20.6 Gy剂量辐射,DNA合成抑制率达92%,增殖期细胞占3%;辐射后2周未见细胞再增殖,<20.6 Gy辐射未见细胞存活力下降;辐射后P53表达升高,PCNA表达降低。结论：¹⁸⁸Re β辐射对平滑肌细胞增殖抑制有效且持久半数有效剂量为5 Gy最佳辐射剂量为20 Gy。低剂量辐射抑制平滑肌细胞增殖的主要机制为损伤细胞分裂增殖能力。P53及PCNA调控辐射抑制细胞增殖过程。     

Volume-activated Cl− currents in different mammalian non-excitable cell types

The existence and properties of volume-activated Cl^– currents were studied in 15 different cell types (endothelium: human umbilical vein, human aorta, bovine pulmonary artery; fibroblasts: Swiss 3T3, L, C3H 10T1/2 and COS-1; epithelium: KB3, HeLa and A6; blood cells: RBL-2H3 and Jurkat; endothelioma cells derived from both subcutaneous and thymic hemangiomas; skin: IGR1 melanoma). Volume-activated Cl^– currents with common characteristics, i.e. small conductance, outward rectification, higher permeability for iodide than for chloride and sensitivity to block by 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB) could be elicited in all cells. The block of this current by tamoxifen and dideoxyforskolin is different for the various cell types, as well as the time course and the amplitude of the responses induced by repetitive applications of hypotonicity. Volume-activated Cl^– channels with similar biophysical properties are therefore widespread among mammalian cells. This may reflect either a single Cl^– channel that is ubiquitously expressed or a family of functionally related Cl^– channels with cell specific expression patterns.