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世界生命科学前沿动态周报(二)
（03.29--04.04 / 2010）
美宝国际集团:陶国新
本周的进展大多是关于基因治疗方面的。 其中发现控制涡虫再生能力“总开关” 的报道比较有意义。对于我们了解动物的再生有很好的启示。另外两篇有关多巴胺D2受体的文章在基因层面上看到了其对吃垃圾食品和睡眠的影响，对于养生保健或许有所帮助。其余是有关基因疗法的一些进展和基因运载的新技术，基因与衰老、免疫的关系等。
1. 发现控制涡虫再生能力“总开关”
【摘要】
一种名叫“涡虫”的扁形虫即使被切成百段，一两周后每段都会再生出完整的涡虫。涡虫这种超强再生能力一直是科学家感兴趣的研究课题。近年来对涡虫最感兴趣的是从事干细胞研究的科学家，因为研究发现，涡虫再生的奥秘在于其体内有一种散布全身的全能干细胞，其分化能力类似人类胚胎干细胞。有所不同的是，涡虫这种干细胞能在任何时间分化成其他任何种类的细胞。涡虫的身体被切断后，它体内散布在各处的这些干细胞能转变成神经、肌肉、肠等各种组织细胞，重新长出那些失去的部分。
德国科学家最近发现了一种对涡虫的再生能力有关键调节作用的蛋白质。他们希望这一发现有助于人类干细胞研究。 德国马克斯•普朗克分子生物研究所科学家用核糖核酸（RNA）干扰抑制基因表达的方式抑制了涡虫体内蛋白质“Smed—SmB”的合成，结果发现这导致涡虫体内的全能干细胞均不能分裂，涡虫因此失去了再生能力。参与研究的科学家说，这相当于发现了影响涡虫干细胞分裂的“总开关” ，这一发现可能有助于人们深入了解组织缺损修复的机理。由于涡虫细胞中四分之三的基因与人类基因相似，科学家还希望他们的研究成果有助于人类干细胞研究。新华网 发布时间：2010-4-1 16:12:39
【点评】
如果能证实关闭其它基因涡虫依然能再生的话，那么Smed-Smb蛋白就很可能是个涡虫全效生长因子。如果能在哺乳动物体内发现类似功能的蛋白，那将是非常惊人的。或许美宝再生物质里有类似功能的成分！？ 结合上期的“发现哺乳动物控制再生能力基因”关闭p21，开启再生的报道，一对掌控再生能力的基因开关就出现了: p21抑制再生，表达Smed-Smb类似蛋白的基因促进再生。 而发展有效的 RNAi 技术来按需要关闭对应基因可以达到促进或抑制再生的目的。不过看起来离这个结果还很远。但是对于进一步了解动物的再生机理是有帮助的。
【原文摘录】
doi: 10.1242/dev.042564 April 1, 2010 Development 137,1055-1065.
Smed-SmB, a member of the LSm protein superfamily, is essential for chromatoid body organization and planarian stem cell proliferation
Enrique Fernandéz-Taboada1, Sören Moritz2, Dagmar Zeuschner2, Martin Stehling2, Hans R. Schöler2,3, Emili Saló1,* and Luca Gentile2,*
Planarians are an ideal model system to study in vivo the dynamics of adult pluripotent stem cells. However, our knowledge of the factors necessary for regulating the ‘stemness’ of the neoblasts, the adult stem cells of planarians, is sparse. Here, we report on the characterization of the first planarian member of the LSm protein superfamily, Smed-SmB, which is expressed in stem cells and neurons in Schmidtea mediterranea. LSm proteins are highly conserved key players of the splicing machinery. Our study shows that Smed-SmB protein, which is localized in the nucleus and the chromatoid body of stem cells, is required to safeguard the proliferative ability of the neoblasts. The chromatoid body, a cytoplasmatic ribonucleoprotein complex, is an essential regulator of the RNA metabolism required for the maintenance of metazoan germ cells. However, planarian neoblasts and neurons also rely on its functions. Remarkably, Smed-SmB dsRNA-mediated knockdown results in a rapid loss of organization of the chromatoid body, an impairment of the ability to post-transcriptionally process the transcripts of Smed-CycB, and a severe proliferative failure of the neoblasts. This chain of events leads to a quick depletion of the neoblast pool, resulting in a lethal phenotype for both regenerating and intact animals. In summary, our results suggest that Smed-SmB is an essential component of the chromatoid body, crucial to ensure a proper RNA metabolism and essential for stem cell proliferation.
2. 垃圾食品成瘾症或确实存在
【摘要】
美国最新研究显示，肥胖人群无法拒绝美食诱惑的解释可能并不是给自己找借口，垃圾食品成瘾症似乎确实存在。 这一发现是通过老鼠研究得出的。在研究人员无限制地为老鼠提供熏肉、磅饼、糖块以及其它垃圾食品等高热量食物之后，老鼠体重快速增加。随着身体越来越胖，吃东西变成一种强迫，即使这么做双脚会遭受电击，它们也不愿意放下爪子，继续享用美食。相比之下，享用健康食品的老鼠并未增加太多体重，在意识到吃的过多会遭到电击之后，它们便停止进食。研究人员指出，更令人感到吃惊的是，在拿走肥胖老鼠的垃圾食品并换上健康食品之后，这些家伙居然选择绝食。在长达两周时间里，它们拒绝吃任何东西。研究人员尚无法确定研究结果是否也适用于人类。
在对胖老鼠的大脑进行分析时，研究人员发现多巴胺D2受体减少。根据此前进行的研究，这种受体与可卡因和海洛因成瘾有关。肯尼说：“毒瘾的一个标志就是导致大脑奖赏系统工作机制发生变化。”在人工抑制其他老鼠脑中的这种受体之后，这些老鼠也开始情不自禁地转向垃圾食品。波士顿大学医学院成瘾症实验室助理教授皮埃特罗•科特纳表示，不断堆积的脂肪中的一些物质也会改变大脑的奖赏阈限，进而形成一个恶性循环——只有吃得更多，才能获得满足感。科特纳说：“回到正常状态的唯一方式就是长期节食、减少体重同时不再吃垃圾食品。”他与同事此前进行的研究显示，让老鼠摆脱高热量食品可能导致大脑出现与戒毒和戒酒类似的变化。
【点评】
多巴胺相关的大脑奖赏系统工作机制因为成瘾行为而发生改变，多巴胺D2受体减少。该文老鼠实验显示，现在这类成瘾行为行列里很可能又加入了吃垃圾食品。再生养生的健康食谱排除了这种成瘾行为，预防大脑奖赏系统工作机制的异常改变，维持健康的大脑功能。结合下篇的多巴胺D2受体剔除小鼠的睡眠研究，可以推测，垃圾食品成瘾的人很可能的会越吃越多，越睡越多，越长越胖，任期发展下去，最后身体各器官会不堪重负，出现各种病理状况乃至衰竭。可见科学饮食对健康是多么的重要。
【原文摘录】
Nature Neuroscience | Article
Dopamine D2 receptors in addiction-like reward dysfunction and compulsive eating in obese rats
Published online 28 March 2010 Paul M Johnson & Paul J Kenny
We found that development of obesity was coupled with emergence of a progressively worsening deficit in neural reward responses. Similar changes in reward homeostasis induced by cocaine or heroin are considered to be crucial in triggering the transition from casual to compulsive drug-taking. Accordingly, we detected compulsive-like feeding behavior in obese but not lean rats, measured as palatable food consumption that was resistant to disruption by an aversive conditioned stimulus. Striatal dopamine D2 receptors (D2Rs) were downregulated in obese rats, as has been reported in humans addicted to drugs. Moreover, lentivirus-mediated knockdown of striatal D2Rs rapidly accelerated the development of addiction-like reward deficits and the onset of compulsive-like food seeking in rats with extended access to palatable high-fat food. These data demonstrate that overconsumption of palatable food triggers addiction-like neuroadaptive responses in brain reward circuits and drives the development of compulsive eating. Common hedonic mechanisms may therefore underlie obesity and drug addiction.
3. “多巴胺D2受体”研究为治疗“第一晚效应”提供新思路
【摘要】
医学上把一些人换床后无法入睡的现象称为“第一晚效应”。资料显示，随着工业化进程的加快，社会竞争、工作压力、不良夜生活习惯及人口老龄化等原因，全球三分之一的人存在睡眠问题，其中不少人因经常出差不能在习惯的床上睡眠或入睡前情绪改变、精神亢奋或紧张而难以入眠，深陷“第一晚效应”的痛苦之中不能自拔，严重影响到次日工作效率和身体健康。复旦大学医学神经生物学国家重点实验室黄志力课题组研究人员曲卫敏副教授、徐昕红博士等运用高度自动化睡眠觉醒解析系统，记录已经基因剔除“多巴胺D2受体”小鼠的睡眠过程，并结合药理学等手段，从基因到行为解析了多巴胺D2受体在睡眠觉醒调控中的作用。结果发现，与正常小鼠（野生型小鼠）相比，剔除了多巴胺D2受体的小鼠，活动期维持觉醒困难，睡眠量增加。 为模拟人在新环境下如出差住宿等，出现换床后失眠现象，即“第一晚效应”，研究人员更换动物居住环境，考察小鼠在新环境中的睡眠行为。结果显示，多巴胺D2受体正常的小鼠面对新环境刺激，极为不习惯、入睡困难，而剔除了多巴胺D2受体的小鼠则“高枕无忧”，迅速入睡。
【点评】
多巴胺D2受体删除的老鼠实验中不出现”第一晚效应”, 在新环境中能迅速入睡，且睡眠量增加。 这个现象是否与生物钟有关，能否在改善睡眠上有利用价值，值得研究一下，看是否可能用在生物钟饮食疗法上。不过需要注意的是，通过基因敲除技术研究的结论只是单个（或某几个）基因的作用，无法考虑或排除整体的调控在其中的影响，因此结论往往不是那么十分确定的。
【原文摘录】
The Journal of Neuroscience, March 24, 2010, 30(12):4382-4389; doi:10.1523
Essential Role of Dopamine D2 Receptor in the Maintenance of Wakefulness, But Not in Homeostatic Regulation of Sleep, in Mice
Wei-Min Qu,1 Xin-Hong Xu,1 Ming-Ming Yan,1 Yi-Qun Wang,1 Yoshihiro Urade,2 and Zhi-Li Huang1
Dopamine (DA) and its D2 receptor (R) are involved in cognition, reward processing, and drug addiction. However, their roles in sleep–wake regulation remain unclear. Herein we investigated the role of D2R in sleep–wake regulation by using D2R knock-out (KO) mice and pharmacological manipulation. Compared with WT mice, D2R KO mice exhibited a significant decrease in wakefulness, with a concomitant increase in non-rapid eye movement (non-REM, NREM) and REM sleep and a drastic decrease in the low-frequency (0.75–2 Hz) electroencephalogram delta power of NREM sleep, especially during the first 4 h after lights off. The KO mice had decreased mean episode duration and increased episode numbers of wake and NREM sleep, many stage transitions between wakefulness and NREM sleep during the dark period, suggesting the instability of the wake stage in these D2R KO mice. When the KO mice were subjected to a cage change or an intraperitoneal saline injection, the latency to sleep in the KO mice decreased to half of the level for WT mice. The D2R antagonist raclopride mimicked these effects in WT mice. When GBR12909, a dopamine transport inhibitor, was administered intraperitoneally, it induced wakefulness in WT mice in a dose-dependent manner, but its arousal effect was attenuated to one-third in the D2R KO mice. However, these 2 genotypes showed an identical response in terms of sleep rebound after 2, 4, and 6 h of sleep deprivation. These results indicate that D2R plays an essential role in the maintenance of wakefulness, but not in homeostatic regulation of NREM sleep.
4. 番茄基因加抗艾滋药物的新基因疗法治疗癌症
【摘要】
瑞典研究人员最新发现，一种番茄基因与药物组合后能破坏癌细胞，这一发现将有助于用基因疗法治疗癌症。瑞典隆德大学研究人员日前发表公报说，这种番茄基因在帮助建立和修复番茄基因组方面“非常活跃”，但它本身并不足以破坏癌细胞。在先后测试了不同药物后，研究人员最终发现，这种番茄基因与抗艾滋病药物AZT组合后，能更有效地打击癌细胞。研究人员指出，很多人对基因疗法心存疑虑，担心病人的基因在接受治疗后发生改变，引发更多的不良反应。然而，上述研究并不存在这种风险 ，因为番茄基因仅仅被注入癌细胞内，并不影响其他细胞。 新华网 2010-3-29 12:11:30
【点评】
细胞培养以及裸鼠实验显示 西红柿TK1基因与抗艾滋药物逆转录酶抑制剂AZT组合的自杀基因疗法结合干细胞介导的基因注入 显著提高了癌细胞对药物的敏感度，实质性的抑制了肿瘤生长，可以说在针对癌细胞的攻击方面这的确是个很好的策略，只是一方面还仅仅在实验动物身上看到效果，另一方面，它并未有显示出可以治愈癌症的潜力，而且将异种基因转入动物体内哪怕只是体内的癌细胞中会产生什么后果尚未可知，也就存在着未知的风险。最基本的，这种策略依然是治标不治本。
【原文摘录】
Neuro Oncol. 2010 Feb 13 PMID: 20154339
Plant thymidine kinase 1: a novel efficient suicide gene for malignant glioma therapy.
Khan Z, Knecht W, Willer M, Rozpedowska E, Kristoffersen P, Clausen AR, Munch-Petersen B, Almqvist PM, Gojkovic Z, Piskur J, Ekstr?m TJ.
The prognosis for malignant gliomas remains poor, and new treatments are urgently needed. Targeted suicide gene therapy exploits the enzymatic conversion of a prodrug, such as a nucleoside analog, into a cytotoxic compound. Although this therapeutic strategy has been considered a promising regimen for central nervous system (CNS) tumors, several obstacles have been encountered such as inefficient gene transfer to the tumor cells, limited prodrug penetration into the CNS, and inefficient enzymatic activity of the suicide gene. We report here the cloning and successful application of a novel thymidine kinase 1 (TK1) from the tomato plant, with favorable characteristics in vitro and in vivo. This enzyme (toTK1) is highly specific for the nucleoside analog prodrug zidovudine (azidothymidine, AZT), which is known to penetrate the blood-brain barrier. An important feature of toTK1 is that it efficiently phosphorylates its substrate AZT not only to AZT monophosphate, but also to AZT diphosphate, with excellent kinetics. The efficiency of the toTK1/AZT system was confirmed when toTK1-transduced human glioblastoma (GBM) cells displayed a 500-fold increased sensitivity to AZT compared with wild-type cells. In addition, when neural progenitor cells were used as delivery vectors for toTK1 in intracranial GBM xenografts in nude rats, substantial attenuation of tumor growth was achieved in animals exposed to AZT, and survival of the animals was significantly improved compared with controls. The novel toTK1/AZT suicide gene therapy system in combination with stem cell-mediated gene delivery promises new treatment of malignant gliomas.
5. 一种可使药物直接攻击癌细胞的全新方法
【摘要】
加拿大蒙特利尔大学和拉瓦尔大学的科学家发现了一种可使药物直接攻击癌细胞的全新方法，其可为急性骨髓白血病患者等癌症病人带来福音。据科学家称，这种新方法目前已接近于临床试验。相关文章发表在最新出版的《生物化学杂志》上。 研究负责人、蒙特利尔大学药学系教授丁戴尔•拉门塔尔表示，他们发现人体中部分类型的细胞存在一个“门口”，如源自骨髓的细胞就存在一个允许博来霉素等抗癌药物进入的“门”，找到并打开这扇“门”就可让药物直接攻击引发白血病的癌细胞。该成果为癌症治疗开辟了一条新途径。
拉门塔尔教授介绍，他在十年前开始将该理论付诸实践，在与人体细胞十分接近的发酵用酵母上进行了试验。目前所获发现正是基于酵母实验的成果，新方法可被应用于人体细胞，并能很快进入临床治疗。
据介绍，新方法对于癌症患者特别是急性骨髓白血病患者实属福音。急性骨髓白血病影响人的白细胞，这种癌症非常难治疗，绝大部分患者对各种抗癌药物没有反应。拉门塔尔教授表示，新方法可以将抗癌药剂以束流的形式治疗急性骨髓白血病。他说：“例如我们发现博来霉素等抗癌药剂对来自患者身上的淋巴瘤细胞具有正面结果，但同时还要依靠‘门口’的存在。”由于博来霉素不表现为免疫抑制剂，他认为这对患者来说是一个十分利好的消息。
拉门塔尔教授还提醒到，新找到的“门口”只存在于部分细胞类型，比如那些来自于骨髓的细胞，但对于乳腺癌等就不起作用，这样就很难使用博来霉素等来治疗乳腺癌患者。因此，他认为目前应着手寻找能够刺激“门口”产生的方式，这样才能够使用博来霉素等药物治疗更多类型的癌症
【点评】
寻找能够刺激“门口”产生的方式是没准的事情，要寻找只刺激癌细胞“门口”产生的方式，不影响正常细胞，更难。总之不能解决药物对癌细胞和正常细胞的相似作用，化疗的前景就无法看好。如果能像再生营养物质那样在有利于正常细胞的同时消灭癌细胞，把看似矛盾的两个方面统一起来做到一举两得。这才是癌症治疗的最好结果。
【原文摘录】
JBC doi: 10.1074/jbc.M109.046151
The Human Carnitine Transporter SLC22A16 Mediates High Affinity Uptake of the Anticancer Polyamine Analogue Bleomycin-A5*
Mustapha Aouida,1, Richard Poulin§ and Dindial Ramotar,2
Bleomycin is used in combination with other antineoplastic agents to effectively treat lymphomas, testicular carcinomas, and squamous cell carcinomas of the cervix, head, and neck. However, resistance to bleomycin remains a persistent limitation in exploiting the full therapeutic benefit of the drug with other types of cancers. Previously, we documented that the Saccharomyces cerevisiae L-carnitine transporter Agp2 is responsible for the high affinity uptake of polyamines and of the polyamine analogue bleomycin-A5. Herein, we document that the human L-carnitine transporter hCT2 encoded by the SLC22A16 gene is involved in bleomycin-A5 uptake, as well as polyamines. We show that NT2/D1 human testicular cancer cells, which highly express hCT2, are extremely sensitive to bleomycin-A5, whereas HCT116 human colon carcinoma cells devoid of detectable hCT2 expression or MCF-7 human breast cancer cells that only weakly express the permease showed striking resistance to the drug. NT2/D1 cells accumulated fluorescein-labeled bleomycin-A5 to substantially higher levels than HCT116 cells. Moreover, L-carnitine protected NT2/D1 cells from the lethal effects of bleomycin-A5 by preventing its influx, and siRNA targeted to hCT2 induced resistance to bleomycin-A5-dependent genotoxicity. Furthermore, hCT2 overexpression induced by transient transfection of a functional hCT2-GFP fusion protein sensitized HCT116 cells to bleomycin-A5. Collectively, our data strongly suggest that hCT2 can mediate bleomycin-A5 and polyamine uptake, and that the rate of bleomycin-A5 accumulation may account for the differential response to the drug in patients.
6. C60运载基因技术为糖尿病患者送福音
【摘要】
日本东京大学的研究人员首次开发出了利用超小球形碳分子C60（富勒烯）导入基因的新技术。该技术有望为糖尿病患者带来福音。C60是60个碳原子结合在一起形成的直径不足1纳米的球状微小粒子。东京大学副教授野入英世和教授中村荣一率领的研究小组让C60携带4个氨基，制造出了水溶性C60，使其与基因结合成为可能。
研究人员将结合了绿色荧光蛋白基因的水溶性C60注射到实验鼠体内。结果发现实验鼠的肺、肝和脾都出现了该基因，证实了水溶性C60具有强大的基因运载能力 。在随后的实验中，研究人员让水溶性C60携带指导合成胰岛素的基因进入实验鼠体内，结果实验鼠体内的胰岛素水平增加到平常的1.5倍，血糖值也降低了20%以上。研究人员介绍说，与基因结合的水溶性C60穿过细胞膜以后就会与基因分离，随尿液排出体外，不会在体内堆积。
目前，治疗糖尿病的手段主要是通过给患者直接注射胰岛素来降低血糖值。日本研究人员认为，此次开发的新技术达到实用化水平后，降低血糖值效果的持续时间将比直接注射还要长，由此将大大减轻患者的负担。另外，这项新技术还有可能促成安全性更高的基因治疗糖尿病方法的出现。
【点评】
该技术若能发展成熟，可能会为RNAi技术更好的用于需基因治疗的疾病提供很大帮助。总之，这看上去是一项很好的基因运载技术。
【原文摘录】
PNAS doi: 10.1073/pnas.0909223107
In vivo gene delivery by cationic tetraamino fullerene
Rui Maeda-Mamiyaa,b, Eisei Noirib,1, Hiroyuki Isobec, Waka Nakanishic, Koji Okamotob, Kent Doib, Takeshi Sugayad, Tetsuro Izumie, Tatsuya Hommaa, and Eiichi Nakamuraa,1
Application of nanotechnology to medical biology has brought remarkable success. Water-soluble fullerenes are molecules with great potential for biological use because they can endow unique characteristics of amphipathic property and form a self-assembled structure by chemical modification. Effective gene delivery in vitro with tetra(piperazino)fullerene epoxide (TPFE) and its superiority to Lipofectin have been described in a previous report. For this study, we evaluated the efficacy of in vivo gene delivery by TPFE. Delivery of enhanced green fluorescent protein gene (EGFP) by TPFE on pregnant female ICR mice showed distinct organ selectivity compared with Lipofectin; moreover, higher gene expression by TPFE was found in liver and spleen, but not in the lung. No acute toxicity of TPFE was found for the liver and kidney, although Lipofectin significantly increased liver enzymes and blood urea nitrogen. In fetal tissues, neither TPFE nor Lipofectin induced EGFP gene expression. Delivery of insulin 2 gene to female C57/BL6 mice increased plasma insulin levels and reduced blood glucose concentrations, indicating the potential of TPFE-based gene delivery for clinical application. In conclusion, this study demonstrated effective gene delivery in vivo for the first time using a water-soluble fullerene.
7. 基因疗法恢复患眼疾小鼠视力
【摘要】
据国外媒体报道，来自美国纽约州布法罗市、俄亥俄州克利夫兰市和俄克拉何马州的科学家使用基因疗法，改善具有视网膜色素变性疾病的老鼠视力。这一研究结果表明，科学家在使盲人恢复视力的道路上取得了长足的进步。据悉，《美国实验生物学学会联合会杂志》2010年4月刊上发表的一篇研究报告中，科学家详细阐述了利用合成的纳米颗粒，改善具有视网膜色素变性疾病老鼠视力的过程。视网膜色素变性是视网膜光感受器细胞和色素上皮细胞变性，从而导致夜盲和进行性视野缺损的一组具有临床亚型的基因遗传性致盲眼病。
研究小组成员，俄克拉何马州奥克拉荷马大学健康科学中心细胞生物学系莱西博士和她的同事一起，研究了一组带有视网膜缓慢变性基因的老鼠。莱西和她的同事对这些老鼠进行了三种不同类型的治疗方法：一种方法是用包含Rds基因的纳米颗粒来治疗，一种方法是用正常基因来治疗，还有一种方法是通过生理盐水来治疗。 实施三种不同类型的治疗方法后，研究人员将实验老鼠和其它具有视网膜色素变性或视网膜缓慢变性疾病老鼠进行比较，从而分析得出实验老鼠视网膜的功能和结构。研究人员发现，接受纳米颗粒基因疗法的老鼠，其视觉功能得到改善，具有明显愈合的迹象，而且这种效果到实验结束都还保持完好，而接受正常基因和生理盐水治疗的老鼠，其视力不断下降。上述实验结果表明，纳米颗粒是耐受性良好，并且是安全无副作用的治疗方法。
研究人员称，他们希望此研究结果可帮助治愈那些和视网膜色素变性、遗传性疾病和后天视网膜疾病等导致失明的疾病。 《美国实验生物学学会联合会杂志》杂志主编，杰拉尔• 德韦斯曼说：“使盲人恢复视力曾经被称为奇迹。随着我们对进化、遗传学和纳米技术理解的加深，这种神奇的治疗方法将变得非常普遍。”
【点评】
盲人复明是圣经中的神迹，是医学上的难题，上述基因疗法还只是在老鼠实验中显示了效力，不过也能给盲人患者带来一丝期望，是基因治疗的进步之一，虽然基因疗法在临床应用上还很不成熟，也不确定到底能否成熟起来，毕竟这是在干预人体自身的遗传信息，会造成多大的影响，什么样的影响，我们并不清楚。
【原文摘录】
Published as doi: 10.1096/fj.09-139147. (The FASEB Journal. 2010;24:1178-1191.)
Gene delivery to mitotic and postmitotic photoreceptors via compacted DNA nanoparticles results in improved phenotype in a mouse model of retinitis pigmentosa
Xue Cai*, Shannon M. Conley*, Zack Nash*, Steven J. Fliesler , , , Mark J. Cooper|| and Muna I. Naash*,1
The purpose of the present study was to test the therapeutic efficiency and safety of compacted-DNA nanoparticle-mediated gene delivery into the subretinal space of a juvenile mouse model of retinitis pigmentosa. Nanoparticles containing the mouse opsin promoter and wild-type mouse Rds gene were injected subretinally into mice carrying a haploinsufficiency mutation in the retinal degeneration slow (rds+/–) gene at postnatal day (P)5 and 22. Control mice were either injected with saline, injected with uncompacted naked plasmid DNA carrying the Rds gene, or remained untreated. Rds mRNA levels peaked at postinjection day 2 to 7 (PI-2 to PI-7) for P5 injections, stabilized at levels 2-fold higher than in uninjected controls for both P5 and P22 injections, and remained elevated at the latest time point examined (PI-120). Rod function (measured by electroretinography) showed modest but statistically significant improvement compared with controls after both P5 and P22 injections. Cone function in nanoparticle-injected eyes reached wild-type levels for both ages of injections, indicating full prevention of cone degeneration. Ultrastructural examination at PI-120 revealed significant improvement in outer segment structures in P5 nanoparticle-injected eyes, while P22 injection had a modest structural improvement. There was no evidence of macrophage activation or induction of IL-6 or TNF- mRNA in P5 or P22 nanoparticle-dosed eyes at either PI-2 or PI-30. Thus, compacted-DNA nanoparticles can efficiently and safely drive gene expression in both mitotic and postmitotic photoreceptors and retard degeneration in this model. These findings, using a clinically relevant treatment paradigm, illustrate the potential for application of nanoparticle-based gene replacement therapy for treatment of human retinal degenerations.—Cai, X., Conley, S. M., Nash, Z., Fliesler, S. J., Cooper, M. J., Naash, M. I. Gene delivery to mitotic and postmitotic photoreceptors via compacted DNA nanoparticles results in improved phenotype in a mouse model of retinitis pigmentosa.
8. DAF-16基因同寿命密切相关
【摘要】
从遗传学角度研究衰老机制的一组英国科学家4月1日在《公共科学图书馆•综合》网站撰文指出，他们针对实验室蠕虫进行的研究表明，DAF-16基因同寿命、免疫力密切相关。由于很多动物和人体内都拥有DAF-16基因，该发现有助于更好地了解影响人类衰老和免疫功能的原因。全球各地的人们正在大踏步迈向衰老，给健康和社会保障体系提出了巨大挑战。丹麦科学家去年进行的一项研究发现，富裕国家出生的婴儿中，有一半将可以庆祝其百岁诞辰。科学家急切渴望能够找到令人衰老的原因，据此研发出药物帮助人们尽可能长寿，并在有生之年保持健康。英国伯明翰大学的罗宾•梅尔领导了这项研究。梅尔团队比较了4种关系密切的蠕虫的寿命、抗药性以及免疫力情况，他们发现，这4种蠕虫体内的DAF-16基因的活性存在巨大的差异。更重要的是，DAF-16活性的差异同寿命、抵抗力和免疫力相辅相成：DAF-16的活性越高，蠕虫的寿命越长，抗感染的免疫力越好。梅尔表示，这表明，免疫力和衰老密切相关。物种之间的DAF-16基因的活性的差异对衰老和健康具有非常重要的影响，这或许可以解释人与人之间的寿命为何不同。
梅尔称，DAF-16在体内大多数细胞中都很活跃，它们同人体内的FoxO家族调节基因非常类似，科学家认为FOXO家族在动物细胞的分化、生长、增殖、代谢、免疫及衰老调节方面具有多样性功能。英国生物技术和生物科学研究协会负责人道格拉斯•凯尔表示，这个发现将帮助科学家理解决定人类衰老的相关机制。
【点评】
在分子机制上研究衰老和免疫，期望能够找到令人衰老的原因，据此研发出药物帮助人们尽可能长寿，并在有生之年保持健康。在复杂的分子调控网络层次是做这项研究，即使有希望，也还有很漫长的路要走。而在细胞水平上的研究，人体再生复原科学则已经找到了预防衰老的途径并正在用于人体养生。
【原文摘录】
Phenotypic Covariance of Longevity, Immunity and Stress Resistance in the Caenorhabditis Nematodes
Francis R. G. Amrit, Claudia M. L. Boehnisch, Robin C. May*
Abstract
Background
Ageing, immunity and stresstolerance are inherent characteristics of all organisms. In animals, these traits are regulated, at least in part, by forkhead transcription factors in response to upstream signals from the Insulin/Insulin–like growth factor signalling (IIS) pathway. In the nematode Caenorhabditis elegans, these phenotypes are molecularly linked such that activation of the forkhead transcription factor DAF-16 both extends lifespan and simultaneously increases immunity and stress resistance. It is known that lifespan varies significantly among the Caenorhabditis species but, although DAF-16 signalling is highly conserved, it is unclear whether this phenotypic linkage occurs in other species. Here we investigate this phenotypic covariance by comparing longevity, stress resistance and immunity in four Caenorhabditis species.
Methodology/Principal Findings
We show using phenotypic analysis of DAF-16 influenced phenotypes that among four closely related Caenorhabditis nematodes, the gonochoristic species (Caenorhabditis remanei and Caenorhabditis brenneri) have diverged significantly with a longer lifespan, improved stress resistance and higher immunity than the hermaphroditic species (C. elegans and Caenorhabditis briggsae). Interestingly, we also observe significant differences in expression levels between the daf-16 homologues in these species using Real-Time PCR, which positively correlate with the observed phenotypes. Finally, we provide additional evidence in support of a role for DAF-16 in regulating phenotypic coupling by using a combination of wildtype isolates, constitutively active daf-16 mutants and bioinformatic analysis.
Conclusions
The gonochoristic species display a significantly longer lifespan (p<0.0001) and more robust immune and stress response (p<0.0001, thermal stress; p<0.01, heavy metal stress; p<0.0001, pathogenic stress) than the hermaphroditic species. Our data suggests that divergence in DAF-16 mediated phenotypes may underlie many of the differences observed between these four species of Caenorhabditis nematodes. These findings are further supported by the correlative higher daf-16 expression levels among the gonochoristic species and significantly higher lifespan, immunity and stress tolerance in the constitutively active daf-16 hermaphroditic mutants。