氯化锂增加获得性耐药人肺癌细胞系H460R对TRAIL的敏感性

目的:通过人工诱导建立肿瘤坏死因子相关凋亡诱导配体(TRAIL)获得性耐药的肺癌细胞系,氯化锂联合rhTRAIL作用细胞,以期了解氯化锂增敏TRAIL的现象及机制。方法:通过低剂量rhTRAIL诱导人肺大细胞癌细胞系H460,建立TRAIL耐药细胞株H460R并鉴定,应用MTT和流式细胞术分析氯化锂和rhTRAIL联合给药后亲本株与耐药株细胞增殖和凋亡差异,应用RT-PCR和Western blot检测死亡受体表达。结果:rhTRAIL处理亲本株H460和耐药株H460R后细胞存活率存在显著差异(P<0.01),IC50分别为59.2ng/ml和294.8ng/ml。60ng/ml rhTRAIL处理耐药株及亲本株后平均细胞凋亡比例存在显著差异(10.5%vs 19.4%,P<0.01),耐药株表现出显著的TRAIL耐受现象。但联合20mmol/L氯化锂后,MTT及流式细胞术检测耐药株细胞增殖及凋亡发现H460R对rhTRAIL的敏感性显著增加。进一步研究发现死亡受体DR4和DR5的mRNA及蛋白水平升高,这可能是药物增敏的机制之一。结论:氯化锂能够增加获得性耐药细胞系H460R对TRAIL的敏感性,死亡受体表达增加是可能的增敏机制之一。     

MEK/ERKs signaling is essential for lithium-induced neurite outgrowth in N2a cells

Lithium, a drug used for the treatment of bipolar disorder, has been shown to affect different aspects of neuronal development such as neuritogenesis, neurogenesis and survival. The underlying mechanism responsible for lithium's influence on neuronal development, however, still remains to be elucidated. In the present study, we demonstrate that lithium increases the phosphorylation of extracellular-signal regulated kinases (ERKs) and protein kinase B (Akt) and promotes neurite outgrowth in mouse N2a neuroblastoma cells (N2a). The inactivation of mitogen-activated protein kinase kinase (MEK)/ERKs signaling with a MEK inhibitor inhibits neurite outgrowth, but it enhances Akt activation in lithium-treated N2a cells. Furthermore, the inactivation of phosphoinositide-3-kinase (PI3K)/Akt signaling with a PI3K inhibitor increases both lithium-induced ERKs activation and lithium-induced neurite outgrowth. Taken together, our study suggests that lithium-induced neurite outgrowth in N2a cells is regulated by cross-talk between the MEK/ERKs and PI3K/Akt pathways and requires the activation of the MEK/ERKs signaling.     

GSK-3β抑制剂LiCl干预大鼠重症急性胰腺炎的量效关系

目的 探讨糖原合成酶激酶-3β(GSK-3β)抑制剂氯化锂(LiCl)干预大鼠重症急性胰腺炎(SAP)的量效关系.方法 50只SPF级雄性Wistar大鼠,按数字表法随机分为五组(n=10):假手术组(SO组)、重症急性胰腺炎组(SAP组)、LiCl 40 mg/kg组、60 mg/kg组和80 mg/kg预处理组(LiCl组).胆胰管逆行注射5%牛磺胆酸钠溶液(0.1 mL/100 g)制备SAP大鼠模型;SO组在胆胰管内注射等量生理盐水替代牛磺胆酸钠.LiCl组造模前30 min经股静脉注射相应浓度LiCl.术后12 h分批处死大鼠,检测血清淀粉酶(AMY)、谷丙转氨酶(ALT)、肌酐(Cr)、腹水量和肺湿干比.光镜下观察胰腺组织病理学变化并进行病理学评分.结果 SAP组大鼠AMY、ALT、Cr、腹水量、肺湿干比、胰腺病理评分均较SO组升高[(7224.14±1872.41)U/L vs(1780.21±231.12)U/L、(250.56±51.56)U/L vs(98.45±10.36)U/L、(85±19)U/L vs(36±4)U/L、(11.05±2.76)g vs(0.33±0.04)g、(3.21±0.61)vs(1.60±0.20)、(11.52±1.23)分vs(0.54±0.02)分],差异均具有统计学意义(P<0.05);LiCl 60 mg/kg组上述指标与SAP组相比较均有下降[(4215.36±940.24)U/L vs(7224.14±1872.41)U/L、(160.25±35.39)U/L vs(250.56±51.56)U/L、(50±11)U/L vs(85±19)U/L、(6.04±1.03)g vs(11.05±2.76)g、(2.70±0.54)vs(3.21±0.61)、(9.03±1.09)分vs(11.52±1.23)分],差异均有统计学意义(P<0.05);而LiCl 40 mg/kg组AMY、腹水量、肺湿干比以及胰腺病理评分与SAP组比较差异均无统计学意义(P>0.05);LiCl-80 mg/kg组大鼠的ALT升高大于SAP组[(312.47±69.41)U/L vs(250.56±51.56)U/L],差异有统计学意义(P<0.05),Cr水平与SAP组比较[(79±17)U/L vs(85±19)U/L]差异无统计学意义(P>0.05).结论 LiCl-60 mg/kg是干预重症急性胰腺炎大鼠模型最佳有效剂量.     

Increased and decreased preference for saccharin immediately following injections of various agents

Rats pretrained for preference testing and injected with LiCl showed a reduction of saccharin preference in a test with a novel saccharin solution paired with water given 15 min following the injection. Previous exposure to the saccharin solution precluded this selective depression of drinking. In a second experiment, using rats naive to saccharin, insulin injection produced an increase, 2-DG and hypertonic NaCl produced a decrease, and saccharin injection produced no change in saccharin preference.     

Effects of single albino gene substitutions on the performance of mice in a compound avoidance-discrimination task

Congenic C57BL/6J mice of the +/+, +/c and c/c genotypes were tested for motoric activity, active avoidance and visual discrimination performance in a variant of the Thompson-Bryant box. The three genotypes were observed to be identical in terms of motor activity, but differed greatly with regard to both avoidance and discrimination performance. On the avoidance portion of the task the c gene appeared to act in an additive fashion, with the +/+ mice being superior to the +/c mice, which were in turn superior to the c/c animals. Visual discrimination performance, however, indicated that the + allele operated in a dominant manner; here c/c mice were found to be inferior in performance when compared to either +/+ or +/c animals while no differences were observed between the two pigmented genotypes. Therefore, under the conditions of the present experiment, the albino gene shows incomplete dominance for active avoidance behavior and recessiveness for visual discrimination performance.     

Re-establishment of gap junctional intercellular communication (GJIC) between human endometrial carcinomas by prostaglandin E2

Reduced intercellular communication via gap junctions is correlated with carcinogenesis. Gap junctional intercellular communication (GJIC), between normal human endometrial epithelial cells is enhanced when endometrial stromal cells were present in culture. This enhancement of GJIC between normal epithelial cells also occurs when they are cultured in medium conditioned by stromal cells. This observation indicated that a soluble compound (or compounds) produced and secreted by stromal cells mediates GJIC in epithelial cells. Previous studies have shown that endometrial stromal cells release prostaglandin E₂ (PGE₂) and prostaglandin F_2α (PGF_2α) under physiological conditions. When we evaluated the response of normal endometrial epithelial cells to various concentrations of PGE_2, we found enhanced GJIC with 1 nM PGE₂. This is a smaller increase in GJIC than that induced by medium conditioned by stromal cells. When the extracellular concentration of PGE₂ was measured after incubation with stromal cells, it was found to be similar to the concentrations showing maximal GJIC between the normal epithelial cells. When indomethacin was used to inhibit prostaglandin synthesis by stromal cells, GJIC was reduced but not eliminated between normal endometrial epithelial cells. These observations suggest that although PGE₂ secreted by stromal cells is an important mediator of GJIC between the epithelial cells, it is not the sole mediator. Transformed endometrial epithelial cells did not demonstrate GJIC even in the presence of stromal cells. However, we were able to re-establish GJIC in transformed epithelial cells when we added PGE₂ to the cells. Our findings show that PGE₂ may serve as an intercellular mediator between stromal and epithelial cells that regulates GJIC in normal and malignant epithelial cells. This suggests that maintenance of GJIC by preserving or replacing PGE₂ secretion by endometrial stromal cells may have the potential to suppress carcinogenesis in endometrial epithelial cells.     

Drug Development Targeting the Glycogen Synthase Kinase-3β (GSK-3β)-Mediated Signal Transduction Pathway: The Role of GSK-3β in the Maintenance of Steady-State Levels of Insulin Receptor Signaling Molecules and Nav1.7 Sodium Channel in Adrenal Chromaffin Cells

Glycogen synthase kinase-3 (GSK-3) is constitutively active in nonstimulated cells, where the majority of its substrates undergo inactivation/proteolysis by phosphorylation. Extracellular stimuli (e.g., insulin) catalyze inhibitory Ser⁹-phosphorylation of GSK-3β, turning on signaling and causing other biological consequences otherwise constitutively suppressed by GSK-3β. Regulated and dysregulated activities of GSK-3β are pivotal to health, disease, and therapeutics (e.g., insulin resistance, neurodegeneration, tumorigenesis, inflammation); however, the underlying mechanisms of multifunctional GSK-3β remain elusive. In cultured bovine adrenal chromaffin cells, 1) constitutive and negatively-regulated activities of GSK-3β up- and down-regulated insulin receptor, insulin receptor substrate-1 (IRS-1), IRS-2, and Akt levels via controlling proteasomal degradation and protein synthesis; 2) nicotinic receptor/protein kinase C-α (PKC-α) / extracellular signal-regulated kinase (ERK) pathway up-regulated IRS-1 and IRS-2 levels, enhancing insulin-induced the phosphoiNOSitide 3-kinase (PI3K) / Akt / GSK-3β pathway; 3) inhibition of calcineurin by cyclosporin A or FK506 down-regulated IRS-2 level, attenuating insulin-like growth factor-I (IGF-I)-induced ERK and GSK-3β pathways; and 4) insulin, IGF-I or therapeutics (e.g., lithium) up-regulated the voltage-dependent Na_v1.7 sodium channel.     

Thalamic taste nuclei lesions and taste aversion

Rats with thalamic taste nuclei lesions were adapted to a 23 hr 50 min deprivation schedule and then presented with 0.125 percent saccharin followed by an injection of LiCl or saline. When retested with saccharin, animals with lesions showed a marked attenuation in taste aversion as compared to controls.     

Control of polydipsic drinking by a taste aversion procedure

Rats were given daily sessions with free access to food and saccharin flavored water. After fluid consumption had stabilized food was delivered once every minute. Water was always available in the home cage. All rats showed the marked increase in fluid consumption known as schedule-induced polydipsia. The rats were then poisoned with lithium chloride after each of three sessions in an attempt to condition a taste aversion to the saccharin. On recovery from the toxicosis all rats showed first a reduction and then a recovery in saccharin intake. To establish the nature of this effect, the rats were poisoned after saccharin consumption in the home cage. Again there was a marked reduction in polydipsic drinking in the experimental chamber. These results indicate that common incentive mechanisms govern normal and polydipsic drinking and stand in contrast to published results pointing to different drive systems in the brain mediating normal and polydipsic drinking.     

Using profiles of saccharin and water drinking to detect and discriminate actions of drugs and toxicants

Experiments were conducted to investigate the feasibility of using the pattern of saccharin and water drinking to detect acute and chronic administration of drugs and toxicants. Procedural variables were found to be crucial. When rats were naive for the saccharin drinking fluid, a single injection of LiCl or 2-deoxyglucose produced persistant saccharin aversion. Hypertonic saline produced only a transient saccharin aversion. If rats were pre-exposed to saccharin, the 2-deoxyglucose injection and hypertonic NaCl produced an increase in saccharin drinking but LiCl was without effect. Several types of chronic treatment were given to saccharin-experienced rats. Chronic 2-deoxyglucose, LiCl, and Pb administration produced gradually developing saccharin aversion and qualitatively different patterns of saccharin and water drinking. Chronic administration of hypertonic NaCl or insulin or chronic food deprivation had no impact on saccharin preference. It was concluded that patterns of saccharin and water drinking can be used to detect the administration of a drug or toxicant and perhaps even the time course of action, but may not detect a substance given previous to saccharin, perhaps because the animal cannot associate these now familiar perturbations with the novel saccharin solution. This means that existing toxic states may not be detected by using saccharin preference as a probe.