檞皮素抑制人脑胶质瘤SHG-44细胞增殖的实验研究

目的探讨中草药檞皮素抑制人胶质瘤SHG-44细胞的增殖作用。方法运用MTT法检测檞皮素对SHG-44细胞增殖的抑制作用,流式细胞仪技术分析檞皮素对SHG-44细胞增殖周期的影响。结果50～200μmol/L浓度的檞皮素均能抑制SHG-44细胞的生长,且呈明显的时间,剂量依赖性关系,细胞周期分析显示檞皮素组G1期细胞比例明显增多,呈量效关系。结论檞皮素抑制SHG-44细胞增殖,可能通过使SHG-44细胞停滞在G1期而诱导其凋亡。     

蛋白酶体抑制剂Bortezomib对人脑胶质瘤增殖抑制及诱导凋亡作用

目的 探讨Bortezomib对体外SHG-44胶质瘤细胞株形态的影响及诱导凋亡作用.方法 Bortezomib体外处理培养的SHG-44胶质瘤细胞后,采用MTT法检测细胞增殖、HE染色光镜观察、Hochest33342染色荧光显微镜观察以及锇铀铅染色透射电镜观察胶质瘤细胞的形态变化,流式细胞术分析细胞凋亡率及细胞周期.结果 MTT显示Bortezomib抑制SHG-44胶质瘤的增殖与浓度和时间呈相关性;6.0 μmol/L Bortezomib处理SHG-44胶质瘤细胞24 h后,光镜、荧光显微镜、透射电镜下均可见凋亡形态学改变.流式细胞术分析细胞凋亡率为(20.31±2.84)% ( P＜0.01);细胞周期变化显示S期细胞减少到(17.65±6.38)% (P＜0.01),细胞周期被阻滞于G_0/G_1期和G_2/M期,分别为(68.51±5.62) % (P＜0.01)和(14.75±2.48)% (P＜0.01).结论 Bortezomib抑制C6胶质瘤细胞的增殖并诱导其凋亡.     

紫杉醇对胶质瘤SHG-44细胞的增殖抑制和辐射增敏作用

目的观察紫杉醇对胶质瘤SHG-44细胞的抑制及放射增敏作用,并探讨其作用机制。方法不同浓度紫杉醇(0、0.062 5、1.25、2.5、5和10μg/mL)作用于SHG-44细胞,MTT法检测紫杉醇对SHG-44细胞的增殖抑制作用;克隆形成法检测细胞辐射敏感性变化,流式细胞术检测细胞凋亡百分率变化。结果紫杉醇有明显的抑制SHG-44细胞增殖和提高其放射敏感性的作用,紫杉醇可提高3、6 Gy照射24 h后SHG-44细胞的凋亡率。结论紫杉醇对人胶质瘤SHG-44细胞具有杀伤及放射增敏作用。其可能机制是加强射线对肿瘤细胞的诱导凋亡效应,增强其杀伤肿瘤作用。     

姜黄素对人脑胶质瘤SHG-44细胞抑制及凋亡的实验研究

目的探讨姜黄素（erueumin）对人脑胶质瘤SHG-44细胞体外抑制及凋亡调控作用。方法20-80μmol／L姜黄素分别处理SHG-44细胞48h后，MTT法测定姜黄素对SHG-44细胞的细胞抑制作用及各种caspase抑制剂对姜黄素作用的影响；通过流式细胞仪AnnexinV—FITC／PI凋亡检测试剂盒检测细胞凋亡；透射电镜观察细胞形态学变化；荧光分光光度法检测easpase-3活性的改变。结果姜黄素明显抑制SHG-44细胞的增殖；诱导SHG-44细胞凋亡；caspase-3活性增强，各种caspase抑制剂可抑制姜黄素诱导的SHG-44细胞凋亡。结论姜黄素可明显抑制并诱导SHG-44细胞凋亡。     

榄香烯聚氰基丙烯酸正丁酯纳米粒子对SHG-44脑胶质瘤的抑制作用

目的探究榄香烯聚氰基丙烯酸正丁酯纳米粒子(Gd-PBCA-NP)对SHG-44脑胶质瘤的抑制作用。方法提取大鼠的脑胶质瘤细胞SHG-44,并对其进行体外培养实验,将这些脑胶质瘤细胞SHG-44按照随机的原则分为干预组与对照组,对照组为空白,干预组则严格按照Gd-PBCA-NP浓度进行分组,然后采用MTT比色法,对榄香烯不同浓度体外培养下的脑胶质瘤细胞SHG-44的抑制作用给予有效的检测,然后借助流式细胞仪测定干预组的脑胶质瘤细胞SHG-44中的细胞凋亡情况。结果榄香烯的浓度不同,其对SHG-44脑胶质瘤体的抑制作用也有着一定的区别。10.0 mol/L浓度的Gd-PBCA-NP对SHG-44脑胶质瘤的抑制作用明显优于2.50 mol/L浓度,且干预组的脑胶质瘤细胞SHG-44的存活率明显低于对照组(P0.05);Gd-PBCA-NP浓度越高,其对脑胶质瘤细胞SHG-44的抑制作用就越强。结论 Gd-PBCA-NP对SHG-44脑胶质瘤有着明显的抑制作用,能够在一定程度上抑制脑胶质瘤SHG-44细胞的凋亡与增殖,而且能够增强细胞抗肿瘤活性,具有一定的安全性。     

蛋白酶体抑制剂对肾间质成纤维细胞凋亡和增殖的影响

目的:探讨泛素蛋白酶体抑制剂MG-132对大鼠肾间质成纤维细胞(NRK-49F)及其在转化生长因子β1(TGF-β1)诱导下的细胞凋亡和增殖情况,并初步探索其机制。方法:采用5ng/ml的TGF-β1作用于NRK-49F,不同浓度(0~5μM)的蛋白酶体抑制剂MG-132预处理NRK-49F细胞,应用MTT法检测其增殖情况,LDH法检测细胞毒性,应用流式细胞仪检测细胞周期和细胞凋亡率,Hoechst 33258染色观察细胞凋亡形态学变化,Western blot检测p-IκB,IκB蛋白的变化。结果:TGF-β1(5ng/ml)能促进NRK-49F的增殖(0.661±0.04vs0.495±0.06),MG-132(0.25~5μM)呈剂量依赖型抑制这种效应;MG-132(0.1~5μM)对NRK-49F有一定的细胞毒性作用,在0.5μM时达高峰;TGF-β1单独不能促使NRK-49F的凋亡[(3.8±0.4)%vs(4.7±1.6)%],但加用MG-132(0.1~2.5μM)干预后呈剂量依赖关系促进细胞凋亡,MG-132单独也有促进细胞凋亡作用;MG-132(0~1μM)作用于TGF-β1诱导的NRK-49F后,其S期比例呈剂量依赖关系显著减少,G1期细胞比例增加;Hoechst 33258染色显示MG-132(0.5μM) TGF-β1组及MG-132组细胞核均出现核固缩、核碎裂、凋亡小体等凋亡特异形态学改变;Western blot结果显示,TGF-β1及MG-132(2.5μM) TGF-β1均能以时间依赖方式诱导p-IκB、IκB蛋白表达增加,且MG-132能上调TGF-β1诱导下的p-IκB,IκB蛋白表达。结论:泛素蛋白酶体抑制剂MG-132对正常及病理状态下的NRK-49F都有促凋亡作用,并可能通过G1期阻滞过程来抑制其增殖作用。其机制可能与MG-132阻断IκB蛋白的泛素化降解有关。     

RNAi沉默survivin基因对胰腺癌细胞增殖、凋亡和周期的影响

目的 应用RNAi技术沉默胰腺癌细胞株中survivin基因对肿瘤细胞凋亡、细胞周期和增殖的变化.方法 靶向survivin的RNAi载体转染胰腺癌细胞,Western印迹检测基因沉默效果,MTT法和流式细胞术检测细胞增殖、凋亡及细胞周期.结果 survivin基因沉默效率可达85%;MTT检测显示,干扰组survivin沉默细胞增殖抑制与非特异性质粒对照组相比显著增高(P<0.01).流式细胞术显示,干扰组细胞凋亡百分率显著高于非特异性质粒对照组(P<0.01);G_2/M期显著高于非特异性质粒对照组(P<0.01或P<0.05),而S期则显著降低(P<0.05).结论 survivin基因沉默可引起胰腺癌细胞周期阻滞,促进细胞凋亡,抑制细胞增殖.     

五味子粗多糖与顺铂抑制脑胶质瘤细胞增殖的作用

目的研究五味子粗多糖(FSP)单独或与顺铂联合抑制脑胶质瘤细胞增殖的作用。方法培养脑胶质瘤(SHG-44)细胞,应用四甲基偶氮唑(MTT)法检测FSP单独及联合顺铂抑制细胞增殖的情况。结果脑胶质瘤SHG-44细胞在FSP浓度200、400、800μg/ml时,均可以抑制细胞的增殖(P<0.05或P<0.01),且有浓度依赖性。FSP与顺铂联合作用时可以明显地抑制细胞的生长(P<0.01)。结论 FSP可以抑制胶质瘤SHG-44细胞的增殖,联合顺铂时可发挥协同作用。     

MG-132抑制大鼠缺血再灌注心肌细胞凋亡

目的观察蛋白酶体抑制剂MG-132对急性缺血再灌注大鼠心肌细胞凋亡的影响。方法建立大鼠心肌缺血再灌注模型,治疗组及治疗对照组于再灌注前5min静脉注射MG-132(0.75mg/kg),缺血再灌注组及假手术组注射生理盐水,观察各组心肌组织炎症细胞浸润及心肌细胞凋亡情况。结果MG-132能显著抑制心肌梗死周围组织嗜中性粒细胞的浸润;与缺血再灌注组相比,治疗组核因子κBmRNA水平和蛋白水平显著降低(P<0.05);治疗组再灌注2h、6h及24h亚组凋亡指数较缺血再灌注组同时间点显著下降;与缺血再灌注组相比,Bax的积分光密度值降低(P<0.05),Bcl-2蛋白水平明显上调,Bcl-2/Bax比值显著增加。结论蛋白酶体抑制剂能够抑制急性缺血再灌注心肌的凋亡,具有心肌保护作用。     

siRNA抑制PTTG表达对人结肠癌LoVo细胞增殖、凋亡及细胞周期的影响

目的 探讨siRNA抑制人垂体瘤转化基因PTTG表达对人结肠癌LoVo细胞增殖、凋亡及细胞周期的影响.方法 LoVo细胞分为3组:正常细胞对照组、Lipo2000+ pGenesil-2.1 HK组(HK阴性对照组)、Lipo2000+ pGenesil-2.1-PTTG siRNA组.应用Western印迹法检测转染48 h后各组细胞PTTG蛋白表达,MTT法检测转染24、48和72 h后各组细胞增殖情况,流式细胞术检测转染48 h后各组LoVo细胞凋亡和细胞周期情况.结果 pGenesil-2.1-PTTG siRNA质粒转染LoVo细胞后PTTG蛋白表达明显低于正常和HK阴性对照组.MTT比色分析法显示,pGenesil-2.1-PTTGsiRNA质粒转染LoVo细胞后,在24、48和72 h细胞增殖能力均显著低于正常对照组和阴性对照HK组(均P＜0.05).流式细胞术检测结果显示,与对照组相比,转染pGenesil-2.1 PTTG siRNA质粒后LoVo细胞Go/G1期细胞百分率均高于正常和阴性HK对照组(均P＜0.01),而S期细胞百分率均低于正常和阴性HK对照组(均P＜0.01);Annexin V-PI双染结果显示,转染pGenesil-2.1 PTTG siRNA质粒后LoVo细胞凋亡百分率高于正常和阴性HK对照组(均P＜0.01).结论 pGenesil-2.1-PTTG siRNA质粒可明显抑制LoVo细胞PTTG蛋白表达,并进而抑制LoVo细胞增殖,促进其凋亡,使细胞阻滞于G0/G1期.     

Three layer functional model and energy exchange concept of aging process

Relying on a certain degree of abstraction, we can propose that no particular distinction exists between animate or living matter and inanimate matter. While focusing attention on some specifics, the dividing line between the two can be drawn. The most apparent distinction is in the level of structural and functional organization with the dissimilar streams of ‘energy flow’ between the observed entity and the surrounding environment. In essence, living matter is created from inanimate matter which is organized to contain internal intense energy processes and maintain lower intensity energy exchange processes with the environment. Taking internal and external energy processes into account, we contend in this paper that living matter can be referred to as matter of dissipative structure, with this structure assumed to be a common quality of all living creatures and living matter in general. Interruption of internal energy conversion processes and terminating the controlled energy exchange with the environment leads to degeneration of dissipative structure and reduction of the same to inanimate matter, (gas, liquid and/or solid inanimate substances), and ultimately what can be called ‘death.’ This concept of what we call dissipative nature can be extended from living organisms to social groups of animals, to mankind. An analogy based on the organization of matter provides a basis for a functional model of living entities. The models relies on the parallels among the three central structures of any cell (nucleus, cytoplasm and outer membrane) and the human body (central organs, body fluids along with the connective tissues, and external skin integument). This three-part structural organization may be observed almost universally in nature. It can be observed from the atomic structure to the planetary and intergalactic organizations. This similarity is corroborated by the membrane theory applied to living organisms. According to the energy nature of living matter and the proposed functional model, the decreased integrity of a human body's external envelope membrane is a first cause of the structural degradation and aging of the entire organism. The aging process than progresses externally to internally, as in single cell organisms, suggesting that much of the efforts towards the restoration and maintenance of the mechanisms responsible for structural development should be focused accordingly, on the membrane, i.e., the skin. Numerous reports indicate that all parts of the human body, like: bones, blood with blood vessels, muscles, skin, and so on, have some ability for restoration. Therefore, actual revival of not only aging tissue of the human body's membrane, but the entire human body enclosed within, with all internal organs, might be expected. We assess several aging theories within the context of our model and provide suggestions on how to activate the body's own anti-aging mechanisms and increase longevity. This paper presents some analogies and some distinctions that exist between the living dissipative structure matter and inanimate matter, discusses the aging process and proposes certain aging reversal solutions.     

Unusual responses of nocturnal pineal melatonin synthesis and secretion to swimming: Attempts to define mechanisms

Abstract: The effect of swimming at night on rat pineal melatonin synthesis was compared with that of light exposure at night. Rats were forced to swim at 0030 hr (lights out at 2000 hr) and sacrificed by decapitation 15 and 30 min later, immediately after swimming. Other groups of animals were exposed to white light (650μW/cm²) for 15 and 30 min at same time. Swimming caused a rapid and highly significant drop in the melatonin content in the pineal gland; however, the activity of N-acetyltransferase (NAT), the supposed rate limiting enzyme in the melatonin production, was not changed. Despite the drop in pineal melatonin levels, serum concentrations of the indole remained elevated in the rats that swam. In contrast, melatonin levels in the pineal and serum of light exposed rats fell precipitously, accompanied by a significant suppression of NAT activity. Since we anticipated that the strenuous exercise associated with swimming may induce release of artrial natriuretic peptide (ANP) from the heart, which in turn could cause the release of pineal melatonin, in a second study we injected physiological saline intravenously to stretch the cardiac muscle and release ANP. Three milliliters of normal saline was injected during the day into the jugular vein of anesthetized rats that were pretreated with isoproterenol to stimulate pineal melatonin production. Animals were killed 15 min after the saline injection, and pineal NAT activity and pineal melatonin levels were measured. The saline injections caused no alteration in the elevated levels of either NAT or melatonin. These data suggest that the disparity in pineal NAT activity (which was high) and pineal melatonin (which was low), in animals swum at night, may not be caused by ANP which is released during strenuous exercise such as swimming.     

Melatonin and photoperiodic time measurement: Seasonal breeding in the sheep

Abstract: Well-established circadian physiology supports the view that photoperiodic time measurement utilizes the coincidence between the presence of light and a photosensitive phase of a 'biological clock' to alter reproductive status—the so-called external coincidence model of seasonal breeding. In this review, we examine the mechanism whereby photoperiod interacts with presumed suprachiasmatic nuclei activity to allow endogenous melatonin to normally synchronize reproductive activity to the optimal time of year. The Romney Marsh sheep is particularly explored as an experimental model. It is suggested that the on/off activity of seasonal reproduction may be a robust mechanism able to be predictably manipulated by the judicious use of the light/dark cycle and exogenous melatonin, but firmly based on circadian principles.     

The immunoneuroendocrine role of melatonin

Abstract: A tight, physiological link between the pineal gland and the immune system is emerging from a series of experimental studies. This link might reflect the evolutionary connection between self-recognition and reproduction. Pinealectomy or other experimental methods which inhibit melatonin synthesis and secretion induce a state of immunodepression which is counteracted by melatonin. In general, melatonin seems to have an immunoenhancing effect that is particularly apparent in immunodepressive states. The negative effect of acute stress or immunosuppressive pharmacological treatments on various immune parameters are counteracted by melatonin. It seems important to note that one of the main targets of melatonin is the thymus, i.e., the central organ of the immune system. The clinical use of melatonin as an immunotherapeutic agent seems promising in primary and secondary immunodeficiencies as well as in cancer immunotherapy. The immunoenhancing action of melatonin seems to be mediated by T-helper cell-derived opioid peptides as well as by lymphokines and, perhaps, by pituitary hormones. Melatonin-induced-immuno-opioids (MHO) and lymphokines imply the presence of specific binding sites or melatonin receptors on cells of the immune system. On the other hand, lymphokines such as -γ-interferon and interleukin-2 as well as thymic hormones can modulate the synthesis of melatonin in the pineal gland. The pineal gland might thus be viewed as the crux of a sophisticated immunoneuroendocrine network which functions as an unconscious, diffuse sensory organ.     

Serological survey of HIV and syphilis in pregnant women in Madagascar

Objectives Peripartal transmission of human immunodeficiency virus (HIV) and Treponema pallidum, the causative agent of syphilis, leads to severe consequences for newborns. Preventive measures require awareness of the maternal infection. Although HIV and syphilis testing in Madagascar could be theoretically carried out within the framework of the national pregnancy follow‐up scheme, the required test kits are rarely available at peripheral health centres. In this study, we screened blood samples of pregnant Madagascan women for HIV and syphilis seroprevalence to estimate the demand for systemic screening in pregnancy. Methods Retrospective anonymous serological analysis for HIV and syphilis was performed in plasma samples from 1232 pregnant women that were taken between May and July 2010 in Ambositra, Ifanadiana, Manakara, Mananjary, Moramanga and Tsiroanomandidy (Madagascar) during pregnancy follow‐up. Screening was based on Treponema pallidum haemagglutination tests for syphilis and rapid tests for HIV, with confirmation of positive screening results on line assays. Results Out of 1232 pregnant women, none were seropositive for HIV and 37 (3%) were seropositive for Treponema pallidum. Conclusions Our findings are in line with previous studies that describe considerable syphilis prevalence in the rural Madagascan population. The results suggest a need for screening to prevent peripartal Treponema pallidum transmission, while HIV is still rare. If they are known, Treponema pallidum infections can be easily, safely and inexpensively treated even in pregnancy to reduce the risk of transmission.     

Response of rat pineal melatonin to calcium, magnesium, and lithium is circadian stage dependent

Abstract: Herein we documented the response of pineal melatonin production to electrolytes known to be effective on pineal function in view of a possible circadian stage dependence. We studied the release of melatonin by perifused rat pineal glands at 2 different circadian stages corresponding to the middle of the light and dark periods, i.e., respectively, 7 and 19 HALO (Hours After Light Onset, L:D = 12:12). The initial efflux rates were, as expected, much higher in the perifusates of glands removed from rats sacrificed during the dark phase than of those removed during the light phase. After 3 hr of perifusion, melatonin release reached similar levels which were found constant up to the 8th hr of perifusion, whatever the circadian stage. Perifusion of the glands with physiological concentrations for the rat of calcium (5.2 mmol/1) and magnesium (1.34 mmol/1) resulted in a stimulatory effect on the pineal glands removed from rats sacrificed in the middle of the dark period (19 HALO), whereas no effects were observed on the pineal glands removed from rats sacrificed during the light (7 HALO). Lithium (0.28 and 0.55 mmol/1) was ineffective on melatonin release in pineal glands removed 7 and 19 HALO. Our results show differences in the initial efflux rates of melatonin and in the response of perifused pineal glands to calcium and magnesium according to the circadian stage.     

Variabilité des concentrations plasmatiques de l’angiotensinogène et risque d’hypertension artérielle chez le rat transgénique

Aim

Genetic polymorphisms of the human angiotensinogen gene are frequent and may induce up to 30% increase of plasma angiotensinogen concentrations with a blood pressure increase of up to 5 mmHg. Their role for the pathogenesis of human arterial hypertension remains unclear. High plasma angiotensinogen levels could increase the sensitivity to other blood pressure stressors.

Methods

Male transgenic rats with a 9-fold increase of plasma angiotensinogen concentrations and male non-transgenic rats aged 10 weeks were treated or not with NG-Nitro-L-arginine-methyl ester for 3 weeks in their drinking water (n = 3/group). Systolic blood pressure and body weight were measured at baseline and at the end of the study when left ventricular weight and ventricular expression of angiotensin I-converting enzyme and procollagen Iα1 were determined (polymerase chain reaction).

Results

At baseline, transgenic rats had +18 mmHg higher bood pressure and –8% lower body weight compared to non-transgenic rats (P < 0.05) without significant changes for the vehicle groups throughout the study (P > 0.05). NG-Nitro-L-arginine-methyl ester increased blood pressure, left ventricular weight and left ventricular weight indexed for body weight by +41%, +17.6% and +18.6% (P < 0.05) in transgenic and +25%, +5.3% and +6.7% (P > 0.05) in non-transgenic rats compared to untreated animals, respectively. Cardiac gene expression showed no differences between groups (P > 0.05).

Conclusion

Increased plasma angiotensinogen levels may sensitize to additional blood pressure stressors. Our preliminary results point towards an independent role of angiotensinogen in the pathogenesis of human hypertension and associated end-organ damage.