世界生命科学前沿动态周报（五十六）

（9.5-9.11/2011）
美宝国际集团:陶国新 

　　主要内容：全面比较胚胎干细胞和诱导多能干细胞的蛋白质组；耐力锻炼能够促进骨髓造血作用；解决抗癌药物抗药性的新策略；自吞噬通过溶酶体水解调节泡沫细胞中胆固醇外流；衰老的系统环境降低了神经形成和认知功能；T细胞急性淋巴细胞白血病与胰岛素样生长因子受体1过高表达有关。

　　焦点动态：解决抗癌药物抗药性的新策略。

1.  全面比较胚胎干细胞和诱导多能干细胞的蛋白质组

【动态】    
　　美国科学家利用高准度质谱、同位素标记和多元化大规模定量分析蛋白信息的软件相结合的技术，三重测定了四种胚胎干细胞和四种诱导多能干细胞的非常全面的蛋白质组。这24个多能细胞样本的比较产生了一大套鉴定的蛋白质和磷酸化位点。其统计分析显示胚胎干细胞和诱导多能干细胞在蛋白表达和蛋白磷酸化方面存在微小但可重复的差异。将这些数据和RNA序列分析数据合并，他们发现在各个调节层面上存在与功能相关的差异。文章中也介绍了干细胞组学库（SCOR）,一个资源库用于核对和显示多层面测量的定量信息，包括mRNA、蛋白质和翻译后修饰。这是第一次对蛋白质组进行了如此细致全面的比较，得益于蛋白质质谱精度的提高和一次比较多达8种细胞系的技术。但是由于用于临床的话需要多能细胞分化成具有特定功能的体细胞，所以还需要进一步研究干细胞分化后的蛋白生产情况。

【点评】
　　目前的技术进步使得科学家们可以更深入的研究细胞之间所含物质成分的差异，对于研究细胞的生长变化过程很有帮助。但是对于多能干细胞的临床应用推动不大，这是细胞移植的替代疗法固有缺陷所决定的。

【参考论文】
Nature Methods, 2011; DOI:10.1038/nmeth.1699
Proteomic and phosphoproteomic comparison of human ES and iPS cells
Douglas H Phanstiel, Justin Brumbaugh, Craig D Wenger, et al.
Combining high-mass-accuracy mass spectrometry, isobaric tagging and software for multiplexed, large-scale protein quantification, we report deep proteomic coverage of four human embryonic stem cell and four induced pluripotent stem cell lines in biological triplicate. This 24-sample comparison resulted in a very large set of identified proteins and phosphorylation sites in pluripotent cells. The statistical analysis afforded by our approach revealed subtle but reproducible differences in protein expression and protein phosphorylation between embryonic stem cells and induced pluripotent cells. Merging these results with RNA-seq analysis data, we found functionally related differences across each tier of regulation. We also introduce the Stem Cell–Omics Repository (SCOR), a resource to collate and display quantitative information across multiple planes of measurement, including mRNA, protein and post-translational modifications.

1.耐力锻炼能够促进骨髓造血作用
【动态】
　　耐力锻炼能够促进骨髓造血作用， 加拿大科学家研究了耐力训练对造血作用的直接影响及其可能的作用机制。在跑步机上训练4周大的雄性C57Bl/6老鼠十个星期，速度逐步提高，最后一次训练两天后收集组织。用流式细胞仪、鹅卵石区域形成细胞实验、甲基纤维素菌落形成实验评价骨髓中被动员的造血干细胞和祖细胞。用实时定量PCR和蛋白质印迹实验测定造血细胞因子的产生。用组化实验评价骨髓微环境对训练的适应性变化。对于不同类型细胞，耐力训练能够增加骨髓中被动员的造血干细胞和祖细胞50%到800%。训练同时减少了78%的骨髓腔脂肪，增加了至少60%骨骼肌造血因子的表达。 不运动的老鼠作为以上实验的对照组。结论是，耐力训练大大促进了造血作用，机制是通过改善骨髓微环境结构和增加骨骼肌造血因子的表达。一周三次，每次跑不到半小时，已足够对实验鼠的骨髓造血作用产生显著影响。

【点评】
　　间充质干细胞最可能变成脂肪或骨细胞，取决于所走路线。利用跑步机训练老鼠，该研究表明有氧锻炼触发这些细胞更多变成骨细胞而不是脂肪，而不运动的老鼠的相同干细胞更多变成脂肪。

【参考论文】
The FASEB Journal, 2011; DOI: 10.1096/fj.11-189043
Endurance exercise training promotes medullary hematopoiesis
J. M. Baker, M. De Lisio, G. Parise.
Endurance exercise is a poorly defined yet powerful mediator of hematopoiesis. The purpose of this study was to directly investigate the effects of endurance exercise training on hematopoiesis and to identify potential mechanisms responsible for any observed changes. Four-week-old male C57Bl/6 mice were trained on a treadmill at progressive speeds over a 10-wk period. Tissues were harvested 2 d following the final training session. Flow cytometry, the cobblestone area-forming cell assay, and the methycellulose colony-forming unit assay were used to assess medullary and mobilized hematopoietic stem and progenitor cells. Quantitative real-time PCR and Western blots were used to measure hematopoietic cytokine production. Histochemistry was also used to assess adaptations to exercise in the bone marrow niche. Depending on the cell type, endurance training increased medullary and mobilized hematopoietic stem and progenitor cell content from 50 to 800%. Training also reduced marrow cavity fat by 78%. Skeletal muscle hematopoietic cytokine expression was also increased at least 60% by training. Sedentary mice served as controls for the above experiments. In conclusion, endurance exercise training greatly promotes hematopoiesis and does so through improvements in medullary niche architecture as well as increased skeletal muscle hematopoietic cytokine production.-Baker, J. M., De Lisio, M., Parise, G. Endurance exercise training promotes medullary hematopoiesis.

3.  解决抗癌药物抗药性的新策略

【动态】
　　西妥昔单抗是针对表皮生长因子受体（EGFR）的抗体，临床能有效治疗结肠直肠癌、头颈癌和非小细胞肺癌，特别是有野生型致癌基因KRAS 和BRAF的癌症。但最终都因为逐渐产生的抗药性而限制了其治疗效果，而抗药性原因还不清楚。美国科学家及其国际合作者的最新研究显示激活细胞中ERBB2信号，不管是通过扩增ERBB2还是上调heregulin，都会产生持久的细胞外信号调节的激酶1/2信号，结果导致西妥昔单抗抗药性。抑制ERBB2或破坏ERBB2/ERBB3异二聚体能够恢复体内外对西妥昔单抗的敏感性。有一组表现出新生的或获得性的西妥昔单抗抗药性的结肠直肠癌患者出现ERBB2扩增或高水平人血液heregulin。合在一起看，这些发现确定了两种不同的抗药性机制，都促进了介导西妥昔单抗抗药性的异常ERBB2信号。而且，这些结果提示ERBB2抑制剂与西妥昔单抗联合使用可能是合理的治疗策略，应该在西妥昔单抗抗药的患者中进行评价。

【点评】
　　该研究发现了西妥昔单抗抗药的患者中出现的替代EGFR的ERBB2信号使得癌细胞得以规避西妥昔单抗的伤害，堵住ERBB2信号途径可以恢复西妥昔单抗的的抗癌效果。只是这终究是被动的应对措施，不知道什么时候癌细胞又发展出另一种抗药机制。寻找癌症发病的最根本原因，开发主动预防和治疗的途径才是根本解决癌症的方式。

【参考论文】
Science Translational Medicine, 2011; 3 (99): 99ra86 DOI:10.1126/scitranslmed.3002442
Activation of ERBB2 Signaling Causes Resistance to the EGFR-Directed Therapeutic Antibody Cetuximab
Kimio Yonesaka, Kreshnik Zejnullahu, Isamu Okamoto, et al.
Cetuximab, an antibody directed against the epidermal growth factor receptor, is an effective clinical therapy for patients with colorectal, head and neck, and non-small cell lung cancer, particularly for those with KRAS and BRAF wild-type cancers. Treatment in all patients is limited eventually by the development of acquired resistance, but little is known about the underlying mechanism. Here, we show that activation of ERBB2 signaling in cell lines, either through ERBB2 amplification or through heregulin up-regulation, leads to persistent extracellular signal-regulated kinase 1/2 signaling and consequently to cetuximab resistance. Inhibition of ERBB2 or disruption of ERBB2/ERBB3 heterodimerization restores cetuximab sensitivity in vitro and in vivo. A subset of colorectal cancer patients who exhibit either de novo or acquired resistance to cetuximab-based therapy has ERBB2 amplification or high levels of circulating heregulin. Collectively, these findings identify two distinct resistance mechanisms, both of which promote aberrant ERBB2 signaling, that mediate cetuximab resistance. Moreover, these results suggest that ERBB2 inhibitors, in combination with cetuximab, represent a rational therapeutic strategy that should be assessed in patients with cetuximab-resistant cancers.

4.  自吞噬通过溶酶体水解调节泡沫细胞中胆固醇外流

【动态】
　　脂滴是巨噬细胞形成的泡沫细胞中储存胆固醇的主要地方，也是治疗动脉粥样硬化的潜在靶点。以胆甾醇酯形式储存的胆固醇从这里释放出来转运到胆固醇接受体。现有理论认为细胞质内胆甾醇酯的水解都是中性的胆甾醇酯水解酶的作用。而美国和加拿大的科学家最近发现在吞入胆固醇的巨噬细胞中，除了中性的胆甾醇酯水解酶，溶酶体在水解脂滴胆甾醇酯中也起重要作用。此外，他们还发现脂滴是通过自吞噬进入溶酶体，其中的溶酶体酸性脂肪酶水解脂滴胆甾醇酯产生游离胆固醇，主要是为了ABCA1依赖的外流。这一过程是被巨噬细胞吞入胆固醇所特异诱导。他们的结论是巨噬细胞形成的泡沫细胞中，溶酶体的水解作用参与了动员脂滴中胆固醇进行逆行转运。

【点评】
　　胆固醇在动脉壁上积累导致动脉粥样硬化或动脉狭窄致使堵塞减少心脏血流，常常最终产生中风和心脏病发作。该研究发现的自吞噬在水解脂滴胆固醇中起作用，可以促使胆固醇从泡沫细胞中向外转运而不是内流积累，从而减轻胆固醇在动脉壁上的积累。

【参考论文】
Cell Metabolism, 2011; 13 (6): 655 DOI: 10.1016/j.cmet.2011.03.023
Autophagy Regulates Cholesterol Efflux from Macrophage Foam Cells via Lysosomal Acid Lipase
Mireille Ouimet, Vivian Franklin, Esther Mak, et al.
The lipid droplet (LD) is the major site of cholesterol storage in macrophage foam cells and is a potential therapeutic target for the treatment of atherosclerosis. Cholesterol, stored as cholesteryl esters (CEs), is liberated from this organelle and delivered to cholesterol acceptors. The current paradigm attributes all cytoplasmic CE hydrolysis to the action of neutral CE hydrolases. Here, we demonstrate an important role for lysosomes in LD CE hydrolysis in cholesterol-loaded macrophages, in addition to that mediated by neutral hydrolases. Furthermore, we demonstrate that LDs are delivered to lysosomes via autophagy, where lysosomal acid lipase (LAL) acts to hydrolyze LD CE to generate free cholesterol mainly for ABCA1-dependent efflux; this process is specifically induced upon macrophage cholesterol loading. We conclude that, in macrophage foam cells, lysosomal hydrolysis contributes to the mobilization of LD-associated cholesterol for reverse cholesterol transport.

5.  衰老的系统环境降低了神经形成和认知功能

【动态】
　　在中枢神经系统，衰老导致成体神经干细胞/祖细胞以及神经形成的迅速减少，同时伴随认知功能的损害。有趣的是，这种损坏可以通过系统干预如锻炼身体而减轻。美国科学家利用异时异种共生表明全身存在的血源性因子能够以年龄依赖性的方式抑制或促进老鼠的成体神经形成，相应的，年轻老鼠置于老的全身环境或接触老龄老鼠的血浆会降低突触可塑性，损害对恐惧因果关系和对空间的认知和记忆。他们确定了包括CCL11（即嗜酸细胞激活趋化因子） 在内的细胞因子血浆水平与异时异种共生和衰老老鼠中的神经形成减少有关，在健康老年人的血浆和脑脊液中这些细胞因子水平增高。最后，在年轻老鼠体内增加外周CCL11趋化因子水平减少了成体神经形成和损害了学习记忆能力。所有的数据合在一起说明了衰老过程中观察到的神经形成减少和认知功能损害部分是由于血源性细胞因子的变化。

【点评】
　　该研究所发现的血液细胞因子的变化与衰老表征之间有关联，它们是相互促进还是互为因果并不十分清楚。但是如果不从根本上解决衰老问题，这些现象总会发生，至于它们之间的相互关系就不重要了。

【参考论文】
Nature, 2011; 477 (7362): 90 DOI:10.1038/nature10357
The ageing systemic milieu negatively regulates neurogenesis and cognitive function
Saul A. Villeda, Jian Luo, Kira I. Mosher, et al. 
In the central nervous system, ageing results in a precipitous decline in adult neural stem/progenitor cells and neurogenesis, with concomitant impairments in cognitive functions. Interestingly, such impairments can be ameliorated through systemic perturbations such as exercise. Here, using heterochronic parabiosis we show that blood-borne factors present in the systemic milieu can inhibit or promote adult neurogenesis in an age-dependent fashion in mice. Accordingly, exposing a young mouse to an old systemic environment or to plasma from old mice decreased synaptic plasticity, and impaired contextual fear conditioning and spatial learning and memory. We identify chemokines—including CCL11 (also known as eotaxin)—the plasma levels of which correlate with reduced neurogenesis in heterochronic parabionts and aged mice, and the levels of which are increased in the plasma and cerebrospinal fluid of healthy ageing humans. Lastly, increasing peripheral CCL11 chemokine levels in vivo in young mice decreased adult neurogenesis and impaired learning and memory. Together our data indicate that the decline in neurogenesis and cognitive impairments observed during ageing can be in part attributed to changes in blood-borne factors.

6.  T细胞急性淋巴细胞白血病与胰岛素样生长因子受体1过高表达有关

【动态】
T细胞急性淋巴细胞白血病（T-ALL）是未成熟T细胞的恶性肿瘤，经常表现出异常激活Notch1 和 PI3K–Akt信号途径。虽然激活PI3K–Akt信号途径的基因突变已被确认，相关的生长因子依赖的激活所起的作用还不清楚。美加德法科学家的国际合作研究发现了白血病干细胞的停止信号，显示药物抑制或基因删除胰岛素样生长因子受体1（IGF1R）阻碍了T-ALL细胞的生长和活力，而适度减少IGF1R信号可以中和由在同基因/同源异基因的次级接受体中的可移植性所定义的白血病起始细胞的活性。IGF1R是Notch1的一个作用靶点，而Notch1信号途径是维持T-ALL细胞高水平表达IGF1R所必须的。这些发现提示Notch对白血病起始细胞活性的作用可能部分受增强T-ALL细胞对周围环境中生长因子的反应性所调节，并为用IGF1R抑制剂提高治疗的起始反应和长期治愈T-ALL患者提供了很强的理论基础。

【点评】
该研究表明激素类作用的胰岛素样生长因子受体1功能异常升高可能会导致细胞癌变促进肿瘤起始细胞的增殖分化。这也提示了某些激素类药物的使用可能存在的潜在致癌风险。

【参考论文】
Journal of Experimental Medicine, 2011; DOI: 10.1084/jem.20110121
High-level IGF1R expression is required for leukemia-initiating cell activity in T-ALL and is supported by Notch signaling
H. Medyouf, S. Gusscott, H. Wang, et al.  
T cell acute lymphoblastic leukemia (T-ALL) is an aggressive cancer of immature T cells that often shows aberrant activation of Notch1 and PI3K–Akt pathways. Although mutations that activate PI3K–Akt signaling have previously been identified, the relative contribution of growth factor-dependent activation is unclear. We show here that pharmacologic inhibition or genetic deletion of insulin-like growth factor 1 receptor (IGF1R) blocks the growth and viability of T-ALL cells, whereas moderate diminution of IGF1R signaling compromises leukemia-initiating cell (LIC) activity as defined by transplantability in syngeneic/congenic secondary recipients. Furthermore, IGF1R is a Notch1 target, and Notch1 signaling is required to maintain IGF1R expression at high levels in T-ALL cells. These findings suggest effects of Notch on LIC activity may be mediated in part by enhancing the responsiveness of T-ALL cells to ambient growth factors, and provide strong rationale for use of IGF1R inhibitors to improve initial response to therapy and to achieve long-term cure of patients with T-ALL.