世界生命科學(xué)前沿動態(tài)周報（七十九）

（6.10-6.16/2012）
美寶國際集團:陶國新 

　　主要內(nèi)容：再生細(xì)胞能夠恢復(fù)因角膜異常造成的視覺障礙；AMPK促進代謝壓力下腫瘤細(xì)胞的存活；內(nèi)源性逆轉(zhuǎn)錄病毒作用開啟胚胎干細(xì)胞的全能性；炎癥是如何導(dǎo)致癌癥的；多靶點的系統(tǒng)干預(yù)抗癌更有效；發(fā)育時期的內(nèi)臟細(xì)菌數(shù)量調(diào)節(jié)大腦的血清素水平。

　　焦點動態(tài)：內(nèi)源性逆轉(zhuǎn)錄病毒作用開啟胚胎干細(xì)胞的全能性。

1. 再生細(xì)胞能夠恢復(fù)因角膜異常造成的視覺障礙

【動態(tài)】角膜內(nèi)皮是角膜內(nèi)表面的一單層細(xì)胞，維持角膜的透明性。角膜內(nèi)皮功能障礙是嚴(yán)重視覺障礙的一個主要原因。日本科學(xué)家最近發(fā)現(xiàn)在移植培養(yǎng)的角膜內(nèi)皮細(xì)胞時伴以低分子量的Rho關(guān)聯(lián)激酶（ROCK）抑制劑，能夠成功恢復(fù)角膜的透明性。以前的注射到角膜組織上的角膜內(nèi)皮細(xì)胞會被眼房水沖掉，附著很差，而有研究表明ROCK信號途徑干預(yù)此附著過程。因此他們體外培養(yǎng)了兔子的角膜內(nèi)皮細(xì)胞注射到角膜內(nèi)皮損壞的兔子眼前房，結(jié)果與ROCK抑制劑一同注射的兔子的角膜在注射48小時后完全恢復(fù)了透明性，而沒有同時注射ROCK抑制劑的兔子的角膜模糊不清腫脹嚴(yán)重。這種細(xì)胞治療沒有觀察到并發(fā)癥，在有ROCK抑制劑存在下重建的角膜內(nèi)皮呈現(xiàn)出正常的六角形細(xì)胞形態(tài)。之后用與人更相近的猴子所做的同樣試驗也取得了同樣的結(jié)果，恢復(fù)了單層六角形細(xì)胞及角膜長期的透明性。

【點評】該研究表明ROCK抑制劑調(diào)節(jié)下的角膜內(nèi)皮細(xì)胞移植可能對人角膜內(nèi)皮功能障礙有效，鑒于手術(shù)治療替換受傷的角膜內(nèi)皮受制于捐獻角膜的短缺，該種結(jié)合ROCK抑制劑的細(xì)胞療法有可能最終提供醫(yī)生一種新的再生醫(yī)學(xué)的治療方式。但是在真正應(yīng)用于人體取得成功之前，這還只是一種推測。

【參考論文】   
The American Journal of Pathology, 2012 DOI: 10.1016/j.ajpath.2012.03.033
A ROCK Inhibitor Converts Corneal Endothelial Cells into a Phenotype Capable of Regenerating In Vivo Endothelial Tissue
N. Okumura, N. Koizumi, M. Ueno, et al.
Corneal endothelial dysfunction accompanied by visual disturbance is a primary indication for corneal transplantation. We previously reported that the adhesion of corneal endothelial cells (CECs) to a substrate was enhanced by the selective ROCK inhibitor Y-27632. It is hypothesized that the inhibition of ROCK signaling may manipulate cell adhesion properties, thus enabling the transplantation of cultivated CECs as a form of regenerative medicine. In the present study, using a rabbit corneal endothelial dysfunction model, the transplantation of CECs in combination with Y-27632 successfully achieved the recovery of corneal transparency. Complications related to cell injection therapy, such as the abnormal deposition of the injected cells as well as the elevation of intraocular pressure, were not observed. Reconstructed corneal endothelium with Y-27632 exhibited a monolayer hexagonal cell shape with a normal expression of function-related markers, such as ZO-1, and Na+/K+-ATPase, whereas reconstruction without Y-27632 exhibited a stratified fibroblastic phenotype without the expression of markers. Moreover, transplantation of CECs in primates in the presence of the ROCK inhibitor also achieved the recovery of long-term corneal transparency with a monolayer hexagonal cell phenotype at a high cell density. Taken together, these results suggest that the selective ROCK inhibitor Y-27632 enables cultivated CEC-based therapy and that the modulation of Rho-ROCK signaling activity serves to enhance cell engraftment for cell-based regenerative medicine.

 2. AMPK促進代謝壓力下腫瘤細(xì)胞的存活

【動態(tài)】美國科學(xué)家最近發(fā)現(xiàn)AMP激活的蛋白激酶（AMPK）在腫瘤形成時不止會限制細(xì)胞增殖也會幫助癌細(xì)胞存活，對癌細(xì)胞而言AMPK既是“朋友”又是“敵人”。而以前的研究發(fā)現(xiàn)AMPK抑制培養(yǎng)的癌細(xì)胞的生長，使其被看作是開發(fā)新化療藥物的有希望的潛在靶點。但是該新研究發(fā)現(xiàn)當(dāng)細(xì)胞處于代謝壓力下時（葡萄糖攝取不足），會激活A(yù)MPK促進細(xì)胞存活。其機制是AMPK通過調(diào)節(jié)控制脂肪酸合成和氧化的酶而間接調(diào)節(jié)NADPH的抗氧化功能。這也有助于解釋以前的一項意外的發(fā)現(xiàn)，即有AMPK缺陷或激活A(yù)MPK的酶LKB1缺陷的細(xì)胞對癌變有抵抗性。

【點評】 AMPK由于其抑制細(xì)胞增殖的作用可能依然會成為開發(fā)新化療藥物的有希望的潛在靶點，但是該研究說明此種策略若不同時阻斷AMPK對能量短缺時細(xì)胞的保護作用，那么很可能會失敗。

【參考論文】   
Nature, 2012; DOI:10.1038/nature11066
AMPK regulates NADPH homeostasis to promote tumour cell survival during energy stress
Sang-Min Jeon, Navdeep S. Chandel, Nissim Hay. 
Overcoming metabolic stress is a critical step for solid tumour growth. However, the underlying mechanisms of cell death and survival under metabolic stress are not well understood. A key signalling pathway involved in metabolic adaptation is the liver kinase B1 (LKB1)–AMP-activated protein kinase (AMPK) pathway. Energy stress conditions that decrease intracellular ATP levels below a certain level promote AMPK activation by LKB1. Previous studies showed that LKB1-deficient or AMPK-deficient cells are resistant to oncogenic transformation and tumorigenesis, possibly because of the function of AMPK in metabolic adaptation. However, the mechanisms by which AMPK promotes metabolic adaptation in tumour cells are not fully understood. Here we show that AMPK activation, during energy stress, prolongs cell survival by redox regulation. Under these conditions, NADPH generation by the pentose phosphate pathway is impaired, but AMPK induces alternative routes to maintain NADPH and inhibit cell death. The inhibition of the acetyl-CoA carboxylases ACC1 and ACC2 by AMPK maintains NADPH levels by decreasing NADPH consumption in fatty-acid synthesis and increasing NADPH generation by means of fatty-acid oxidation. Knockdown of either ACC1 or ACC2 compensates for AMPK activation and facilitates anchorage-independent growth and solid tumour formation in vivo, whereas the activation of ACC1 or ACC2 attenuates these processes. Thus AMPK, in addition to its function in ATP homeostasis, has a key function in NADPH maintenance, which is critical for cancer cell survival under energy stress conditions, such as glucose limitations, anchorage-independent growth and solid tumour formation in vivo.

 3. 內(nèi)源性逆轉(zhuǎn)錄病毒作用開啟胚胎干細(xì)胞的全能性

【動態(tài)】胚胎干細(xì)胞的治療作用潛力巨大，但是太多的科學(xué)和非科學(xué)方面的阻礙使得科學(xué)家們無法將其用于疾病治療。美國科學(xué)家最近發(fā)現(xiàn)在胚胎發(fā)育早期，胚胎干細(xì)胞周期性的進出一種“神奇狀態(tài)”，在這種狀態(tài)下，一連串的細(xì)胞潛能必需的基因被激活，這種稱作全能性的狀態(tài)使得胚胎干細(xì)胞具有變成體內(nèi)任何類型細(xì)胞的獨特能力，使其成為極具吸引力的治療靶點。在胚胎發(fā)育初期，還只有5-8個細(xì)胞時，這些干細(xì)胞是全能的能夠變成任何類型細(xì)胞。3-5天后胚胎發(fā)育成囊胚，此時的干細(xì)胞是多能的，能夠變成幾乎所有類型細(xì)胞。這些科學(xué)家通過RNA序列檢測，監(jiān)測了老鼠未成熟的卵母細(xì)胞和兩細(xì)胞階段的胚胎，發(fā)現(xiàn)了一系列與全能性緊密相關(guān)的基因以及這些基因是被與這些干細(xì)胞相鄰的逆轉(zhuǎn)錄病毒所激活。人類基因組的近8%是由先祖時期發(fā)生的古代病毒感染的遺跡構(gòu)成，代代相傳但不會產(chǎn)生病毒感染。該研究發(fā)現(xiàn)細(xì)胞利用了某些這類病毒調(diào)節(jié)自身基因的開關(guān)。在胚胎發(fā)育早期的某個很短的特定時間，利用嚴(yán)密控制的古病毒的殘余打開數(shù)百基因使得細(xì)胞有能力發(fā)育成身體的任何組織。

【點評】該研究深化了對于細(xì)胞發(fā)育潛能重要的基因網(wǎng)絡(luò)的了解，發(fā)現(xiàn)了將胚胎干細(xì)胞調(diào)回更年輕狀態(tài)（可塑性更強）的機制。基因中古病毒遺跡的被利用也說明以前被忽視的或被認(rèn)為是垃圾的很多東西可能具有意想不到的關(guān)鍵作用。

【參考論文】            
Nature, 2012; DOI:10.1038/nature11244
Embryonic stem cell potency fluctuates with endogenous retrovirus activity
Todd S. Macfarlan, Wesley D. Gifford, Shawn Driscoll, et al.
Embryonic stem (ES) cells are derived from blastocyst-stage embryos and are thought to be functionally equivalent to the inner cell mass, which lacks the ability to produce all extraembryonic tissues. Here we identify a rare transient cell population within mouse ES and induced pluripotent stem (iPS) cell cultures that expresses high levels of transcripts found in two-cell (2C) embryos in which the blastomeres are totipotent. We genetically tagged these 2C-like ES cells and show that they lack the inner cell mass pluripotency proteins Oct4 (also known as Pou5f1), Sox2 and Nanog, and have acquired the ability to contribute to both embryonic and extraembryonic tissues. We show that nearly all ES cells cycle in and out of this privileged state, which is partially controlled by histone-modifying enzymes. Transcriptome sequencing and bioinformatic analyses showed that many 2C transcripts are initiated from long terminal repeats derived from endogenous retroviruses, suggesting this foreign sequence has helped to drive cell-fate regulation in placental mammals.

 4. 炎癥是如何導(dǎo)致癌癥的

【動態(tài)】肝臟、結(jié)腸或胃等器官發(fā)生癌癥的一個最大的風(fēng)險因素是這些器官因病毒或細(xì)菌感染造成的長期發(fā)炎。美國MIT的一項最新研究追蹤了感染肝螺桿菌的老鼠肝臟和結(jié)腸里一系列基因和化學(xué)變化，給予我們現(xiàn)今關(guān)于這類感染是如何驅(qū)使組織癌變的最完整的了解。發(fā)炎是身體應(yīng)對感染或損傷的正常反應(yīng)，當(dāng)免疫系統(tǒng)監(jiān)測到病原體或細(xì)胞損傷時，它激活巨噬細(xì)胞和中性粒細(xì)胞的匯集，這些細(xì)胞的任務(wù)是吞噬細(xì)菌、死細(xì)胞和碎片（死亡或損傷細(xì)胞釋放的蛋白、核酸等分子），在此期間會產(chǎn)生高活性的化學(xué)物質(zhì)幫助分解細(xì)菌，但同時這些化學(xué)物質(zhì)也會擴散到組織中，長此以往，炎癥很可能引起組織癌變。MIT的實驗對形成慢性肝臟和結(jié)腸感染的老鼠進行了20周的觀察，發(fā)現(xiàn)肝臟和結(jié)腸對感染的反應(yīng)不同。只有在結(jié)腸里，中性粒細(xì)胞會分泌次氯酸，通過氯化嚴(yán)重?fù)p害蛋白、DNA和RNA。本來次氯酸是用于殺死細(xì)菌的，但泄漏到周圍組織里后就損害了結(jié)腸的上皮細(xì)胞。另一個不同是在經(jīng)歷DAN損傷后，肝臟的DNA修復(fù)系統(tǒng)比結(jié)腸的更活躍。 這兩種不同都對結(jié)腸更不利，導(dǎo)致長期炎癥更易于引起結(jié)腸癌。

【點評】 創(chuàng)新的全面的研究會幫助我們更好的認(rèn)識癌癥，該研究是我們對于長期炎癥和癌癥的關(guān)系有了進一步的了解，有助于采取更好的措施來預(yù)防癌癥。

【參考論文】  
Proceedings of the National Academy of Sciences, 2012; DOI:10.1073/pnas.1207829109
PNAS Plus: Infection-induced colitis in mice causes dynamic and tissue-specific changes in stress response and DNA damage leading to colon cancer
A. Mangerich, C. G. Knutson, N. M. Parry, et al. 
Helicobacter hepaticus-infected Rag2-/- mice emulate many aspects of human inflammatory bowel disease, including the development of colitis and colon cancer. To elucidate mechanisms of inflammation-induced carcinogenesis, we undertook a comprehensive analysis of histopathology, molecular damage, and gene expression changes during disease progression in these mice. Infected mice developed severe colitis and hepatitis by 10 wk post-infection, progressing into colon carcinoma by 20 wk post-infection, with pronounced pathology in the cecum and proximal colon marked by infiltration of neutrophils and macrophages. Transcriptional profiling revealed decreased expression of DNA repair and oxidative stress response genes in colon, but not in liver. Mass spectrometric analysis revealed higher levels of DNA and RNA damage products in liver compared to colon and infection-induced increases in 5-chlorocytosine in DNA and RNA and hypoxanthine in DNA. Paradoxically, infection was associated with decreased levels of DNA etheno adducts. Levels of nucleic acid damage from the same chemical class were strongly correlated in both liver and colon. The results support a model of inflammation-mediated carcinogenesis involving infiltration of phagocytes and generation of reactive species that cause local molecular damage leading to cell dysfunction, mutation, and cell death. There are strong correlations among histopathology, phagocyte infiltration, and damage chemistry that suggest a major role for neutrophils in inflammation-associated cancer progression. Further, paradoxical changes in nucleic acid damage were observed in tissue- and chemistry-specific patterns. The results also reveal features of cell stress response that point to microbial pathophysiology and mechanisms of cell senescence as important mechanistic links to cancer.

 5. 多靶點的系統(tǒng)干預(yù)抗癌更有效

【動態(tài)】美國科學(xué)家最近的研究表明相比于特異攻擊某一疾病病理過程中某一基因或蛋白的高度選擇性的特效藥物，在已知化合物庫中找尋那些為數(shù)不多的能夠廣泛破壞整個病理過程的分子可能是更有效的治療策略。他們發(fā)現(xiàn)這種從針對單一靶點轉(zhuǎn)變?yōu)橥瑫r針對一系列靶點的策略使得我們在不造成很多副作用的情況下獲得好得多的阻止癌癥的能力。該策略已產(chǎn)生了兩個潛在的藥物AD80 和 AD81,比現(xiàn)有抗癌藥物凡德他尼（vandetanib）對果蠅的療效更好毒性更低，已被美國FDA批準(zhǔn)用于治療某一特定類型的甲狀腺癌。廣義上講，絕大多數(shù)藥物只是干預(yù)疾病過程中的蛋白和基因的化合物，其破壞關(guān)鍵的疾病過程的能力越強，它就越有效。另一方面，它干擾身體其他部分的作用越大，它的毒性就越大。很多抗癌藥物對癌細(xì)胞和正常細(xì)胞有同樣的殺傷力?，F(xiàn)在看來，選擇性越高越好的觀念也許并不正確。對于癌癥的治療，作用于癌癥病理過程的多個環(huán)節(jié)的低選擇性的化合物或許會帶來更好的效果和更低的毒性。

【點評】 傳統(tǒng)的藥物設(shè)計認(rèn)為化合物的選擇性越高越好，但是該研究的實踐表明，這種觀念也許過時了，尤其是在癌癥治療領(lǐng)域，選擇性低的化合物反而可能更好，意味著特異性干預(yù)疾病單一環(huán)節(jié)的策略向多環(huán)節(jié)系統(tǒng)干預(yù)的策略轉(zhuǎn)移。

【參考論文】   
Nature, 2012; 486 (7401): 80 DOI: 10.1038/nature11127
Chemical genetic discovery of targets and anti-targets for cancer polypharmacology
Arvin C. Dar, Tirtha K. Das, Kevan M. Shokat, Ross L. Cagan. 
The complexity of cancer has led to recent interest in polypharmacological approaches for developing kinase-inhibitor drugs; however, optimal kinase-inhibition profiles remain difficult to predict. Using a Ret-kinase-drivenDrosophila model of multiple endocrine neoplasia type 2 and kinome-wide drug profiling, here we identify that AD57 rescues oncogenic Ret-induced lethality, whereas related Ret inhibitors imparted reduced efficacy and enhanced toxicity. Drosophila genetics and compound profiling defined three pathways accounting for the mechanistic basis of efficacy and dose-limiting toxicity. Inhibition of Ret plus Raf, Src and S6K was required for optimal animal survival, whereas inhibition of the ‘a(chǎn)nti-target’ Tor led to toxicity owing to release of negative feedback. Rational synthetic tailoring to eliminate Tor binding afforded AD80 and AD81, compounds featuring balanced pathway inhibition, improved efficacy and low toxicity in Drosophila and mammalian multiple endocrine neoplasia type 2 models. Combining kinase-focused chemistry, kinome-wide profiling and Drosophila genetics provides a powerful systems pharmacology approach towards developing compounds with a maximal therapeutic index.

 6. 發(fā)育時期的內(nèi)臟細(xì)菌數(shù)量調(diào)節(jié)大腦的血清素水平

【動態(tài)】愛爾蘭科學(xué)家最新研究表明正常成年人的大腦功能依賴于發(fā)育期間內(nèi)臟微生物的存在。血清素（5-羥色胺）是涉及情緒調(diào)節(jié)的主要化學(xué)物質(zhì)，隨壓力、焦慮、抑郁等改變，大多數(shù)臨床有效的抗抑郁藥也是作用于這一神經(jīng)化學(xué)物質(zhì)。該研究用無菌老鼠模型表明生命早期缺乏細(xì)菌顯著影響了成年后大腦的血清素濃度，尤其是對雄性動物影響更大，對大腦功能造成了不可逆的永久印跡。腦-腸-微生物軸的存在是維持正常健康必須的，腸道菌群在內(nèi)臟和大腦的雙向通訊中起了重要作用。

【點評】該研究提供了有趣的機會通過干預(yù)腸道菌群來調(diào)節(jié)大腦的功能障礙，對維護整體健康包括心理健康有多重含意。

【參考論文】   
Molecular Psychiatry, June 12, 2012 DOI:10.1038/mp.2012.77
The microbiome-gut-brain axis during early life regulates the hippocampal serotonergic system in a sex-dependent manner
G Clarke, S Grenham, P Scully,et al.  
Bacterial colonisation of the intestine has a major role in the post-natal development and maturation of the immune and endocrine systems. These processes are key factors underpinning central nervous system (CNS) signalling. Regulation of the microbiome–gut–brain axis is essential for maintaining homeostasis, including that of the CNS. However, there is a paucity of data pertaining to the influence of microbiome on the serotonergic system. Germ-free (GF) animals represent an effective preclinical tool to investigate such phenomena. Here we show that male GF animals have a significant elevation in the hippocampal concentration of 5-hydroxytryptamine and 5-hydroxyindoleacetic acid, its main metabolite, compared with conventionally colonised control animals. Moreover, this alteration is sex specific in contrast with the immunological and neuroendocrine effects which are evident in both sexes. Concentrations of tryptophan, the precursor of serotonin, are increased in the plasma of male GF animals, suggesting a humoral route through which the microbiota can influence CNS serotonergic neurotransmission. Interestingly, colonisation of the GF animals post weaning is insufficient to reverse the CNS neurochemical consequences in adulthood of an absent microbiota in early life despite the peripheral availability of tryptophan being restored to baseline values. In addition, reduced anxiety in GF animals is also normalised following restoration of the intestinal microbiota. These results demonstrate that CNS neurotransmission can be profoundly disturbed by the absence of a normal gut microbiota and that this aberrant neurochemical, but not behavioural, profile is resistant to restoration of a normal gut flora in later life.