世界生命科學前沿動態(tài)周報（五十六）

（9.5-9.11/2011）
美寶國際集團:陶國新 

　　主要內容：全面比較胚胎干細胞和誘導多能干細胞的蛋白質組；耐力鍛煉能夠促進骨髓造血作用；解決抗癌藥物抗藥性的新策略；自吞噬通過溶酶體水解調節(jié)泡沫細胞中膽固醇外流；衰老的系統(tǒng)環(huán)境降低了神經形成和認知功能；T細胞急性淋巴細胞白血病與胰島素樣生長因子受體1過高表達有關。

　　焦點動態(tài)：解決抗癌藥物抗藥性的新策略。

1.  全面比較胚胎干細胞和誘導多能干細胞的蛋白質組

【動態(tài)】    
　　美國科學家利用高準度質譜、同位素標記和多元化大規(guī)模定量分析蛋白信息的軟件相結合的技術，三重測定了四種胚胎干細胞和四種誘導多能干細胞的非常全面的蛋白質組。這24個多能細胞樣本的比較產生了一大套鑒定的蛋白質和磷酸化位點。其統(tǒng)計分析顯示胚胎干細胞和誘導多能干細胞在蛋白表達和蛋白磷酸化方面存在微小但可重復的差異。將這些數(shù)據(jù)和RNA序列分析數(shù)據(jù)合并，他們發(fā)現(xiàn)在各個調節(jié)層面上存在與功能相關的差異。文章中也介紹了干細胞組學庫（SCOR）,一個資源庫用于核對和顯示多層面測量的定量信息，包括mRNA、蛋白質和翻譯后修飾。這是第一次對蛋白質組進行了如此細致全面的比較，得益于蛋白質質譜精度的提高和一次比較多達8種細胞系的技術。但是由于用于臨床的話需要多能細胞分化成具有特定功能的體細胞，所以還需要進一步研究干細胞分化后的蛋白生產情況。

【點評】
　　目前的技術進步使得科學家們可以更深入的研究細胞之間所含物質成分的差異，對于研究細胞的生長變化過程很有幫助。但是對于多能干細胞的臨床應用推動不大，這是細胞移植的替代療法固有缺陷所決定的。

【參考論文】
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.耐力鍛煉能夠促進骨髓造血作用
【動態(tài)】
　　耐力鍛煉能夠促進骨髓造血作用， 加拿大科學家研究了耐力訓練對造血作用的直接影響及其可能的作用機制。在跑步機上訓練4周大的雄性C57Bl/6老鼠十個星期，速度逐步提高，最后一次訓練兩天后收集組織。用流式細胞儀、鵝卵石區(qū)域形成細胞實驗、甲基纖維素菌落形成實驗評價骨髓中被動員的造血干細胞和祖細胞。用實時定量PCR和蛋白質印跡實驗測定造血細胞因子的產生。用組化實驗評價骨髓微環(huán)境對訓練的適應性變化。對于不同類型細胞，耐力訓練能夠增加骨髓中被動員的造血干細胞和祖細胞50%到800%。訓練同時減少了78%的骨髓腔脂肪，增加了至少60%骨骼肌造血因子的表達。 不運動的老鼠作為以上實驗的對照組。結論是，耐力訓練大大促進了造血作用，機制是通過改善骨髓微環(huán)境結構和增加骨骼肌造血因子的表達。一周三次，每次跑不到半小時，已足夠對實驗鼠的骨髓造血作用產生顯著影響。

【點評】
　　間充質干細胞最可能變成脂肪或骨細胞，取決于所走路線。利用跑步機訓練老鼠，該研究表明有氧鍛煉觸發(fā)這些細胞更多變成骨細胞而不是脂肪，而不運動的老鼠的相同干細胞更多變成脂肪。

【參考論文】
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.  解決抗癌藥物抗藥性的新策略

【動態(tài)】
　　西妥昔單抗是針對表皮生長因子受體（EGFR）的抗體，臨床能有效治療結腸直腸癌、頭頸癌和非小細胞肺癌，特別是有野生型致癌基因KRAS 和BRAF的癌癥。但最終都因為逐漸產生的抗藥性而限制了其治療效果，而抗藥性原因還不清楚。美國科學家及其國際合作者的最新研究顯示激活細胞中ERBB2信號，不管是通過擴增ERBB2還是上調heregulin，都會產生持久的細胞外信號調節(jié)的激酶1/2信號，結果導致西妥昔單抗抗藥性。抑制ERBB2或破壞ERBB2/ERBB3異二聚體能夠恢復體內外對西妥昔單抗的敏感性。有一組表現(xiàn)出新生的或獲得性的西妥昔單抗抗藥性的結腸直腸癌患者出現(xiàn)ERBB2擴增或高水平人血液heregulin。合在一起看，這些發(fā)現(xiàn)確定了兩種不同的抗藥性機制，都促進了介導西妥昔單抗抗藥性的異常ERBB2信號。而且，這些結果提示ERBB2抑制劑與西妥昔單抗聯(lián)合使用可能是合理的治療策略，應該在西妥昔單抗抗藥的患者中進行評價。

【點評】
　　該研究發(fā)現(xiàn)了西妥昔單抗抗藥的患者中出現(xiàn)的替代EGFR的ERBB2信號使得癌細胞得以規(guī)避西妥昔單抗的傷害，堵住ERBB2信號途徑可以恢復西妥昔單抗的的抗癌效果。只是這終究是被動的應對措施，不知道什么時候癌細胞又發(fā)展出另一種抗藥機制。尋找癌癥發(fā)病的最根本原因，開發(fā)主動預防和治療的途徑才是根本解決癌癥的方式。

【參考論文】
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.  自吞噬通過溶酶體水解調節(jié)泡沫細胞中膽固醇外流

【動態(tài)】
　　脂滴是巨噬細胞形成的泡沫細胞中儲存膽固醇的主要地方，也是治療動脈粥樣硬化的潛在靶點。以膽甾醇酯形式儲存的膽固醇從這里釋放出來轉運到膽固醇接受體?，F(xiàn)有理論認為細胞質內膽甾醇酯的水解都是中性的膽甾醇酯水解酶的作用。而美國和加拿大的科學家最近發(fā)現(xiàn)在吞入膽固醇的巨噬細胞中，除了中性的膽甾醇酯水解酶，溶酶體在水解脂滴膽甾醇酯中也起重要作用。此外，他們還發(fā)現(xiàn)脂滴是通過自吞噬進入溶酶體，其中的溶酶體酸性脂肪酶水解脂滴膽甾醇酯產生游離膽固醇，主要是為了ABCA1依賴的外流。這一過程是被巨噬細胞吞入膽固醇所特異誘導。他們的結論是巨噬細胞形成的泡沫細胞中，溶酶體的水解作用參與了動員脂滴中膽固醇進行逆行轉運。

【點評】
　　膽固醇在動脈壁上積累導致動脈粥樣硬化或動脈狹窄致使堵塞減少心臟血流，常常最終產生中風和心臟病發(fā)作。該研究發(fā)現(xiàn)的自吞噬在水解脂滴膽固醇中起作用，可以促使膽固醇從泡沫細胞中向外轉運而不是內流積累，從而減輕膽固醇在動脈壁上的積累。

【參考論文】
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.  衰老的系統(tǒng)環(huán)境降低了神經形成和認知功能

【動態(tài)】
　　在中樞神經系統(tǒng)，衰老導致成體神經干細胞/祖細胞以及神經形成的迅速減少，同時伴隨認知功能的損害。有趣的是，這種損壞可以通過系統(tǒng)干預如鍛煉身體而減輕。美國科學家利用異時異種共生表明全身存在的血源性因子能夠以年齡依賴性的方式抑制或促進老鼠的成體神經形成，相應的，年輕老鼠置于老的全身環(huán)境或接觸老齡老鼠的血漿會降低突觸可塑性，損害對恐懼因果關系和對空間的認知和記憶。他們確定了包括CCL11（即嗜酸細胞激活趨化因子） 在內的細胞因子血漿水平與異時異種共生和衰老老鼠中的神經形成減少有關，在健康老年人的血漿和腦脊液中這些細胞因子水平增高。最后，在年輕老鼠體內增加外周CCL11趨化因子水平減少了成體神經形成和損害了學習記憶能力。所有的數(shù)據(jù)合在一起說明了衰老過程中觀察到的神經形成減少和認知功能損害部分是由于血源性細胞因子的變化。

【點評】
　　該研究所發(fā)現(xiàn)的血液細胞因子的變化與衰老表征之間有關聯(lián)，它們是相互促進還是互為因果并不十分清楚。但是如果不從根本上解決衰老問題，這些現(xiàn)象總會發(fā)生，至于它們之間的相互關系就不重要了。

【參考論文】
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細胞急性淋巴細胞白血病與胰島素樣生長因子受體1過高表達有關

【動態(tài)】
T細胞急性淋巴細胞白血?。═-ALL）是未成熟T細胞的惡性腫瘤，經常表現(xiàn)出異常激活Notch1 和 PI3K–Akt信號途徑。雖然激活PI3K–Akt信號途徑的基因突變已被確認，相關的生長因子依賴的激活所起的作用還不清楚。美加德法科學家的國際合作研究發(fā)現(xiàn)了白血病干細胞的停止信號，顯示藥物抑制或基因刪除胰島素樣生長因子受體1（IGF1R）阻礙了T-ALL細胞的生長和活力，而適度減少IGF1R信號可以中和由在同基因/同源異基因的次級接受體中的可移植性所定義的白血病起始細胞的活性。IGF1R是Notch1的一個作用靶點，而Notch1信號途徑是維持T-ALL細胞高水平表達IGF1R所必須的。這些發(fā)現(xiàn)提示Notch對白血病起始細胞活性的作用可能部分受增強T-ALL細胞對周圍環(huán)境中生長因子的反應性所調節(jié)，并為用IGF1R抑制劑提高治療的起始反應和長期治愈T-ALL患者提供了很強的理論基礎。

【點評】
該研究表明激素類作用的胰島素樣生長因子受體1功能異常升高可能會導致細胞癌變促進腫瘤起始細胞的增殖分化。這也提示了某些激素類藥物的使用可能存在的潛在致癌風險。

【參考論文】
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.