Women jù zài yiqi de luxíng shì zhè chang hunyin zhong zuì nánwàng de yibùfèn. 6 Yuèchu, women xian dào liang liàng che qiánwang nándáketa zhou Sisseton de nèizhàn shídài, ránhòu jìnrù bei dáketa zhou, dàodá dí jinsen yixi de huangdì, ránhòu dào nándáketa zhou de heishan, zuìhòu zài huílái jia. Nu'ér lái nà hé ta de nán péngyou jié méi yin bùdé bù shiyòng qízhong yi liàng qìche zaodian huí jia, yinwèi ta bùdé bù jieshòu fayuàn de rènmìng. Nà ba yuè de lìng yigè luxíng shì zài wo de xiaowu fùjìn de subìlì er hú. Women zuìjìn shougòule yi zhi míng jiào luosù de dà gou. Ta hé women zài yiqi, zài shatan shàng hé tíngche chang li de biérén de qìche shàng liáotian, bùjiu zhihòu jiù xiaoshile. Wo hen yíhàn, wo de xiongdì an dí, ránhòu chéngnuò zài an ka ka de xinlíng shèshi, bùnéng péi women qù nà cì luxíng.
Wo hen kuài faxiàn wangzhàn shàng you xuduo wangzhàn mianfèi zhanshì sèqíng nèiróng. Dangrán, zhèxie wangzhàn you shengjí chéngben, dàn yigè rén jiang wúxiàn qí dì baochí zìyóu de dongxi, bìng gandào manyì. Wo bù gan xiangxìn wo suo kàn dào de - luoti nuxìng hé nánzi zhí háng gè zhong xìng xíngwéi de shìpín. Zhèxie fùnu dou shì niánlíng, zhongzú, tizhòng hé shengao, dàn dou xiyin rén. Yóuyú xìng kuàilè zhuyào zài xinlíng, zhè jihu yu zhenshí de dongxi yiyàng hao. Dangrán méiyou duoshao gongzuò. Wo chéngwéi wo yisheng zhong dì yi cì jingcháng xiaofèi sèqíng.
Mar. 2017: I gave a tutorial at ICASSP 2017 together with Prof. Nikos Sidiropouos, Prof. Vagelis Papalexakis (University of California Riverside) and Prof. L. De Lathauwer (KU Leuven). The title is ‘‘ Tensor Decomposition for Signal Processing and Machine Learning ’’ which is based on our IEEE Transactions on Signal Processing overview paper. Check out the slides and the camera-ready paper.

Zhè shì yigè lumang de judòng. Zhulì'an dì èr tian zaoshang jiù líkai mìxixibi zhou, suirán ta huì duanzàn huí dào míngnísudá zhou, dàn ta jìhuà wúxiàn qí de zài nàli zhù. Ciwài, ta shì yigè zhongshí guanxì zhong de nu tóngzhì. Wo méiyou àn ta yào yigè baobao. Dànshì, rúguo wèilái de qíngkuàng shì zhèngquè de, wo quèshí xiwàng ta liaojie wo de yuànwàng. Rán'ér, yigè zhòngyào de yinsù shì, wo xiang kàn kàn wo shìfou you yongqì dàsheng shuo chu wo de xiangfa. You mèilì de nurén yizhí dou dònghè wo. Zhulì'an qíguài de shì you xiyin lì.
Interestingly, in the presence of 50 nM bafilomycin, sporadic ALS iMNs also contained significantly fewer GFP+mRFP+ vesicles than controls (Figure 2, A and C, and Supplemental Figure 2B; 3 controls and 5 sporadic ALS patients), indicating that autophagosome formation was impaired in sporadic ALS iMNs. Thus, both the C9ORF72 and sporadic ALS iMNs display decreased rates of autophagosome formation.
In the past when our Weibo and WeChat accounts were not taken over (by the new media centre of Xiamen Daily)… Journalists running to the spot would make an instant call to our editors, saying how a piece was on WeChat… Nowadays, the journalists no longer have such enthusiasm… Anyway, whatever the achievement is, they are not ours. (Interviewee No. 8)

Over the past two decades, China’s media convergence has undergone three stages (Li, 2017; Yu, 2015). Stage I lasted from the mid-1990s to the late 1990s, during which the content of traditional media was transplanted onto the new media platform. Stage II began in the early years of the 21st century, during which traditional media co-produced content with new media. Stage III started in 2006 and continues to the present, during which traditional media collaborated with new media in many aspects. All the stages of media convergence concur with the conglomeration and capitalisation of Chinese media industries. China’s media reform has constituted the basic context of domestic media convergence. For the past 40 years, the main drive of China’s media reform is the state. Each critical time point of media reform cannot proceed without the propulsion of the national policy from top to bottom.
Suoyou zhèxie yalì hé yalì dou fàng zài huáiyùnle qi gè duo yuè de nurén shenshang, xila hé liang gè nu'ér zài gébì de fángjian dengdài san jian wòshì de fángjian li ban jìn yigè zhuang man xiangzi de xiao fángjian. Tóngshí, xila shuo ta xuyào 1175 meiyuán cáinéng ràng ta huí che; ta céngjing zài mìsuli zhou jiao fù yiliáo fèiyòng. Yigè libài hòu, ta shuo ta de bèibù zhèngzài shanghài ta, yinwèi ta xuyào shuiliáo. Zhè huale 900 meiyuán. Lìngwài, wo xuyào zhifù $ 325 de língjiàn lái xiufù Erika de che.

Zhè shì yigè lumang de judòng. Zhulì'an dì èr tian zaoshang jiù líkai mìxixibi zhou, suirán ta huì duanzàn huí dào míngnísudá zhou, dàn ta jìhuà wúxiàn qí de zài nàli zhù. Ciwài, ta shì yigè zhongshí guanxì zhong de nu tóngzhì. Wo méiyou àn ta yào yigè baobao. Dànshì, rúguo wèilái de qíngkuàng shì zhèngquè de, wo quèshí xiwàng ta liaojie wo de yuànwàng. Rán'ér, yigè zhòngyào de yinsù shì, wo xiang kàn kàn wo shìfou you yongqì dàsheng shuo chu wo de xiangfa. You mèilì de nurén yizhí dou dònghè wo. Zhulì'an qíguài de shì you xiyin lì.
Dangrán, zhè bi qián méiyou zhifù. Lìngwài yibai yuán, wo anpáile yi míng jing zhang de fùshou san tianhòu, a shén lì you zhoumò qingli ta de cáiwù. Xingqí'èr, jing zhang de dàibiao zài gongyù ménkou fale yigè tongzhi, ràng zu kè èrshísì xiaoshí banjia. Yi tianhòu, tamen huí dàole gongyù. Méiyourén zài nàli dàibiaomen liú xiàle yigè shuomíng, shuo zu kè xiànzài yijing héfa quzhúle. Rúguo you rén huí dào gongyù, zhè jiang bèi rènwéi shì rùqin de.
Males and females accounted for 49.6% and 50.4%, respectively, of the total number of respondents. The respondents with a degree of and below junior college accounted for 1.8%, those with bachelor’s degree accounted for 83.5% and those with a master’s degree or above accounted for 14.7%. Among the respondents, 33.2% came from party organs, whilst 66.8% came from metropolis newspapers. The mean age of the respondents was 31 years old (M = 30.63, SD = 6.35) and the mean number of years at work was 8 (M = 7.62, SD = 6.50).
Wénlín ABC EDOC shì zài ABC gǔdài Hànyǔ cíyuán cídiǎn de jīchǔ shàng biānxiě, gāi cídiǎn shì Xiàwēiyí Dàxué Chūbǎnshè rè pěng de ABC Xìliè zhīyī; tā háishi dì-yī kuǎn kě sōusuǒ gǔdài Hànyǔ zìyuán de diànzǐ cídiǎn, qí zhòngdiǎn zàiyú yǔyīn hé Zhōnghuá wénhuà gēnyuán de hányì. Jīngguò jīngxīn shèjì, shìyòng yú fēi zhuānyè rényuán hé zhuānyè rénshì, zhè bù cídiǎn fēicháng yìyú shǐyòng, nèiróng yǐ zìmǔ shùnxù biānpái, yōngyǒu zhòngduō chuàngxīn de císhū gōngnéng. Měi ge cítiáo tígōng yī ge huò duō ge kěnéng de cíyuán, fēnjiě fāyīn hé qítā xiāngguān shùjù. Cídiǎn jiāng cífǎ shàng yǒu guānlián de cízǔ héwéi “cí xì”, yǐbiàn gèng qīngxī de chǎnshì yǔ zhī guānlián de pàishēngcí hé qítā cíyuán biànhuà. Wénlín ABC EDOC cídiǎn diànzǐ bǎn nèihán yī piān jùyǒu zhòngyào yìyì de zuòzhě xù, jièshàole zuòzhě duì Hànyǔ zài Yàzhōu de yǔyánxué dìwèi kànfǎ, xiángshùle Hànyǔ yǔyán jíqí yuánxíng zuìchū de yǔyīn hé cífǎ shǔxìng.

We next wondered if 3K3A-APC could lower DPR levels in vivo. Baloh and colleagues previously generated C9-BAC mice that harbor a human C9ORF72 gene containing 100–1000 GGGGCC repeats and produce DPRs that aggregate in neurons (36). Forty-eight hours after direct injection into the hippocampus, 3K3A-APC–injected hippocampi showed a significant reduction in poly(GR)+, poly(PR)+, and poly(GP)+ punctae (Figure 6, A–D). Thus, in vivo, 3K3A-APC can reduce levels of both nuclear and cytoplasmically localized DPRs, as well as sense- and antisense-transcript-derived DPRs.

47 On 10 and 12 April, media briefings with Foreign Ministry spokesman Qin Gang were convened. See http://www.fmprc.gov.cn, accessed 31 May 2007. For reports see Beijing Youth Daily, 13 April 2008, Jinghua shibao, 13 April 2005, China Daily, 13 April 2005. Similar announcements by the spokesperson of the Public Security Bureau followed. See People's Daily and Beijing Youth Daily, 22 April 2005.

Dànshì, wo de xiezuò xing qù bìng méiyou tíngzhi. Zuòwéi kuàijìshi, wo shòudào qifa, yánjiu láogong tongjì shùzì, bìng jiànlì lìfa lái suoduan gongzuò shíjian. Zhè chéngwéi wo 1981 nián chuban de dì yi ben shu de zhutí. Wo hé wo de qizi zài báixióng hú maile yigè xiao fángzi. Wo yòng yigè pángbian de fáng jiàn zuòwéi wo de gèrén xuéxí. Shujià shàng kàojìn tianhuaban de jiàzi.
“Nuxing yimin de juese: Zhongguo peidu mama zai Xinjiapo” 女性移民的角色:中国陪读妈妈在新加坡 (The Role of Female Migrants: The Role of Study Mamas in Singapore), presented at the conference on “Migration in China and Asia: Experience and Policy,” in Beijing, organized by the Institute of Ethnology and Anthropology, Chinese Academy of Social Sciences, and Metropolis International, 20-21 May 2010.
Wo hen kuài faxiàn wangzhàn shàng you xuduo wangzhàn mianfèi zhanshì sèqíng nèiróng. Dangrán, zhèxie wangzhàn you shengjí chéngben, dàn yigè rén jiang wúxiàn qí dì baochí zìyóu de dongxi, bìng gandào manyì. Wo bù gan xiangxìn wo suo kàn dào de - luoti nuxìng hé nánzi zhí háng gè zhong xìng xíngwéi de shìpín. Zhèxie fùnu dou shì niánlíng, zhongzú, tizhòng hé shengao, dàn dou xiyin rén. Yóuyú xìng kuàilè zhuyào zài xinlíng, zhè jihu yu zhenshí de dongxi yiyàng hao. Dangrán méiyou duoshao gongzuò. Wo chéngwéi wo yisheng zhong dì yi cì jingcháng xiaofèi sèqíng.
Zài 1983 nián xiàtian de ji gè yuè li, wo hen míliàn ma sha. Ta 28 suì rán'ér, ta de zhíyè shengyá bi wo gèngjia xianjìn. Wo wéiyi de míngsheng jiùshì chubanle yi ben jiào duan de gongzuò shíjian de shu. Ma sha you yigè jiyú xiao qiyè hé shougongyè de jingjì de yuànjing. Women zài qúndao hú bian sànbù, zài diànying zhòng kànle “su fei de xuanzé”, qù diàoyú, zài hùnxie zhànqu canjiale yi chang xì, zài kan nóng héshàng dùguòle yigè xiàwu de shíjian, canjiale yicì qiánshui huódòng, yicì zài ta de míng ní abo lì si gongyù li zuò'ài. Wèntí shì wo hàipà shiqù ma sha. Ta ganjué dào zhè yidian, ránhòu la huílái. Women zài 6 yuè hé 7 yuè fàng huan, dàole 8 yuèdi, women de guanxì jiùshì tongguòle. Wo de yuehuì fangshì wo yeshì “zhìxí” de.
(A) Survival of iMNs from 3 control or 1 SOD1A4V ALS patient line in excess glutamate with 10 nM inactive 3K3A-APC or 3K3A-APC. n = 90 iMNs per line per condition, iMNs from all control lines shown in aggregate for clarity. iMNs quantified from 3 biologically independent iMN conversions per line. (B) Survival of iMNs from 2 C9-ALS lines in excess glutamate with inactive 3K3A-APC or different concentrations of 3K3A-APC. n = 90 iMNs per line per condition, iMNs from both lines shown in aggregate for clarity. iMNs quantified from 3 biologically independent iMN conversions per line. (C) C9-ALS iMNs on day 12 of survival in excess glutamate with inactive 3K3A-APC or 3K3A-APC treatment. This experiment was repeated 3 times with similar results. Scale bar: 100 μm. (D) Survival of control iMNs in excess glutamate with 10 nM inactive 3K3A-APC or 3K3A-APC, n = 90 iMNs per line per condition for 3 control and 3 C9-ALS lines, iMNs quantified from 3 biologically independent iMN conversions per line. (E–G) Survival of iMNs from 2 C9-ALS lines in excess glutamate with 3K3A-APC with or without 3 μM PAR1 antagonist treatment (E) or PAR2 antagonist treatment (F). n = 90 iMNs per line per condition, iMNs from both lines shown in aggregate for clarity. iMNs quantified from 3 biologically independent iMN conversions per line. Survival of iMNs from 2 C9-ALS lines in excess glutamate with 3K3A-APC with or without 9 μM PAR1 ASO treatment (G). n = 90 iMNs per line per condition, iMNs from both lines shown in aggregate for clarity. iMNs quantified from 3 biologically independent iMN conversions per line. Each trace includes neurons from 2 donors with the specified genotype. All iMN survival experiments were analyzed by 2-sided log-rank test and corrected for multiple comparisons if applicable. Statistical significance was calculated using the entire survival time course. (H and I) Survival of iMNs from sporadic ALS (sALS) clone 1 in excess glutamate with 10 nM inactive 3K3A-APC or 3K3A-APC (H), or with 3K3A-APC and DMSO or a PAR1 antagonist (I). n = 90 iMNs per condition. iMNs quantified from 3 biologically independent iMN conversions. For all iMN survival experiments, significance was measure by 2-sided log-rank test using the entire survival time course. The day of differentiation stated on each panel indicates the day of differentiation on which the experimental treatment or time course was initiated.
One of the goals of media convergence in the reconstruction process is to modify the journalist–editor relationship by veering from ‘journalist-centred’ to ‘editor-centred’, such that journalists shall mainly undertake the role of information gatherer, whereas editors perform the role of information synthesiser. Regarding real news production, most new media centre editors are recruited from the members of society who are familiar with new media but inexperienced in news gathering and editing; thus, they are mere ‘stevedores’ rather than ‘processors’ of texts. Given that editors lacked the required qualifications, journalists were forced to undertake additional tasks (e.g., verifying information and converting statements from frontline journalists into text) that should have been performed by editors.
Wo de l?shi, ránhòu zàicì xiàng gaodeng fayuàn qisù. Zhè yicì, duìfang rènzhen duìdài shìqíng. Zài wo tóngyì qiaole yiqian meiyuán hòu, wo gei qizi hé wo de cáichan fenpèi shénme, ta tóngyì zhèxie tiáojiàn. Ta baoliúle báixióng hú de cáichan, ér zài 1991 nián zhiqián, wo geile wo $ 32,750 - $ 9,000, yú'é cóng 1991 nián kaishi, yi bai fen zhi qi de lì shuài cóng 1991 nián kaishi. Shìshí shàng, ta ji nián hòu jiù tíngzhi zhifù zhè bi piào, érqie wo bùdé bù ba zhè bi cáichan de liúzhì quán liú gei dangshí dào qí de jin'é. Dang ta zài 2000 nián chushòu fángchan shí, jíshi wo hái qiànle qián, suoyouquán de zhuanràng yeshì rúci.
Lā Qiáolā (La Jolla), Jiālìfúníyàzhōu -- Wénlín Yánjiūsuǒ Shèhuì Mùdì Gōngsī (SPC), Wénlín Hànyǔ Xuéxí Ruǎnjiàn 4.2 bǎn fāxíng gōngsī, xuānbù chénglì dì-yī rèn dǒngshìhuì. Dǒngshìhuì chéngyuán bāokuò: Xiàwēiyí Dàxué Mǎnuò'ā Xiàoqū Zhōngguó Yánjiū Zhōngxīn fùzhǔrèn, Kǒngzǐ Yánjiūsuǒ zhǔrèn Rèn Yǒuméi (Cynthia Ning) bóshì; Shèngdìyàgē Zhōulì Dàxué Zhōngwén Xiàngmù gùwèn, “Měiguó Zhōngwén Jiàoshī Xuéhuì Zázhì” zhǔbiān Zhāng Zhèngshēng (Zheng-sheng Zhang); Richard Cook bóshì (yánjiū yǔ kāifā fùzǒngcái); Elisabeth Nuboer-Ranjhan (zhànlüè guānxì fùzǒngcái); Mark Roblee(yíngxiāo fùzǒngcái) hé Tom Bishop (zǒngcái).
The official statement of the objective of Chinese media convergence lies in coping with the impacts generated by the information technology revolution. Among those impacts, the decline of tax revenue from media industries is one catalyst for Chinese press industry playing the role of precursor of media convergence. In early 2008, the press industry experienced a phase of decline. Until 2015, China’s television industry firstly took on a gliding tendency in the totality of media placement of advertising, with the advertising revenue of the radio industry deemed stable (Cui, 2016: 6–8). The decline of circulation and advertising revenues is often attributed to the effects of new media such as the change in habits of media exposure. Compared with the radio and television industries, the press industry faces more severe challenges from new media. Thus, the press industry has substantially intense impetus to converge with new media.
Hao xiaoxi shì, ta xiànzài zài fùguó yínháng youyi fèn gongzuò, yigè mei xiaoshí zhifù 15 meiyuán. Ta cóng wo nàli toule qián lái zhifù yínháng fèiyòng. Ta ye zhèngzài jìnrù wojia de lóu xià danwèi, bìng you jinjí yuánzhù zhifù yigè yuè de zujin hé sunhui yajin. Wo zuò chule yigè juédìng. Wo keyi xiàng meiguó yínháng bàogào qizhà xíngwéi bìng shouhuí zijin, ye kenéng jiang xila sòng dào jianyù, fouzé wo keyi jieshòu dàoqiè, xiwàng yiqiè shùnlì. Wo xuanzéle hòu zhe.
This decision means that the Chinese media must be dedicated to Party and government propaganda whilst earning profits and seeking for industry development. This institutional framework enabled the metropolis newspapers to emphasise the function of profitability that emerged in the mid-1990s. Differing from the party organs, the metropolis newspapers have consciously catered to the preferences of the audience in terms of content, including considerable emphasis on the timeliness, relevance and interesting aspects of the news, concentration on social and personal stories and a continuous increase in the proportion of entertainment and leisure contents. Zhao (1998: 159) opined that these measures are more about supplement than disobedience to the party organs, thereby extending the official ideology further to the social, personal and even mental domains.
Zài kaichú nu l?shi zhihòu, wo pìnqingle yi wèi zài wo bàngongshì wèi líhun nuzi gongzuò de laonián nán l?shi. Ta shì yigè yúkuài de rén, dàn wo méiyou xianzhe de qinlüè xìng. Wo qizi de dì yi wèi l?shi zìsha; nàme ta de fal? huobàn, yigè zuì'è de rén, jieshoule zhège ànzi. Liang míng l?shi, wo de hé ta, jingguò ji gè yuè de tánpàn jiejuéle ji gè yuè. Wo de qizi jianchí yào ta zài báixióng hú de cáichan. Women de fa xiàn qingqiú bèi hulüè. 1984 Nián 2 yuè de tánpàn zhongduàn shí, zài faguan Roland Faricy xiansheng jìnxíngle shenpàn. Lìng yifang zài shenpàn chéngxù kaishi qián èrshí fenzhong jiang suoyou de fa xiàn ziliào jiao geile xiangzi.

These findings provide an empirical foundation to organise the interview outline and the type of interviewees that should be recruited in the following interviews. More importantly, the results mentioned above indicate that journalists in Fujian do embrace new technologies. In this case, the journalists are not resisting media convergence because they have negative attitudes towards new technologies but due to institutional, organisational and value reasons, which will be discussed thoroughly in the next section.
Understanding why most interviewed journalists also asserted that the assumptions that ‘multi-skilled journalists’ who can write, photograph and edit is unrealistic (if not entirely unreasonable), and claimed that newspaper journalists should be differentiated from specialised new media journalists who will transform into expert-type journalists in the future by delivering objective, rational and in-depth reportage on a certain subdivided domain. In this conception, new media practitioners remain quite distinct from their peers from the traditional media. However, such a difference has undergone slight changes, that is, traditional media practitioners have begun to admit and accept the possibility that their new media counterparts may be professionalised in the domain of news production. Journalists are consulting about their roles in reference to new media, although they are more inclined towards constriction rather than extension when adjusting their professional boundaries.
Rán'ér, ji gè yuè lái, wo dasuàn jià gei ta. Ta huì lái dào wo zài shèngbaoluó dì dìfang, women jingcháng huì zuò'ài. Ránhòu wo huì zài wo de xiezuò xiàngmù shàng gongzuò, ér an baochíle gè zhong gè yàng de fangshì. Women de guanxì fasheng de dì yigè lièfèng fasheng zài an kaishi zài túshu guan yù dào yigè nánrén de chéngshì zhouwéi. Ta zài zhengbiàn shuo wo bùpèi ta. Tamen kànle yixià zài yigè jiào pínqióng de shèngbaoluó fùjìn de ha wéi de fángzi, bìng juédìng bù héshì. an gei wo yigè zuìhòu tong atum, wo bùdé bùbi zhè gèng hao. Zhè sìhu shì duì wo de yiban tàidù. Suoyi jiéhun bèi gezhìle.
Lín dá líkai gai dìqu hòu, wo yùjiànle an, yigè báirén nu, céngjing gen wo de péngyou hé fángdong ha wéi. Ha wéi de qián shìyou ken·kèlisidì ansen (Ken Christianson) yijing kaishile yigè míng wèi “dí huìjiàn rén” de zìyóu liútong bàozhi, wèi you xìngqù huìjiàn qíta danda di danshen rénshì. Zhèyàng jiù hé ha wéi liánxìle. Ta huàn you yìyù zhèng. Ta dí gege yijing shìtú ba ta zhì yú jingshénbìng yuàn, dàn ta yijing táotuole. Zhè shì yigè you xiyin lì de gùshì. Wo chéngle an dì péngyou, hen kuài ta de liànrén hé wèihunfu. Yijiubaliù nián xiàtian, wo dài ta qù dong jie wojia.

B. Wang Lilin. “Xuzhou Han huaxiangshi yanjiuzhong gongren xianxiang de zai renshi” (A Reconsideration of Generally Acknowledged Phenomena in the Study of Han Stone Pictorial Reliefs at Xuzhou) in Xuzhou bowuguan sanshinian jinian wenji (30th Anniversary Commemorative Edition of Collected Works of The Xuzhou Museum). (Beijing: Yanshan chubanshe, 1992), 166.

With either pulsatile glutamate treatment or neurotrophic factor withdrawal, iMNs from all 6 sporadic ALS patients degenerated significantly faster than control iMNs (Figure 1, B–F, and Supplemental Figure 1, M and N). iMNs derived from additional iPSC clones from the same sporadic ALS patients also degenerated faster than control iMNs (Supplemental Figure 1, O–Q). Thus, iMNs derived from these sporadic ALS patients degenerate significantly faster than controls and at a similar rate to C9ORF72 ALS iMNs. These findings strongly suggest that the donor sporadic ALS patients from which these iMNs were derived harbored genetic variants that contributed to their motor neuron disease. Therefore, iMNs from these patients have the potential to provide insights into mechanisms of neurodegeneration in sporadic ALS cases.
Jíshi Sheila bù zài huáiyùn, ta réngrán cóng wo nàli shou dào qián. Rán'ér, ta yuè lái yuè duo de toule qián. Zài mou xie qíngkuàng xià, ta toule wo de zhipiào kòngbái, xiele dà bi jin'é de zhipiào, wèizàole wo de qianmíng. Zài qíta qíngkuàng xià, wo you wo de zhipiào zhànghào huò xìnyòngka xìnxi, ta anpái wo zhifù ta jiatíng de shouji, qìche dàikuan, diànlan huò qíta zhàngdan. Wo zài 2011 nián guanbì bìng chóngxin kaifàngle sancì jianchá zhànghù, bìngqie liang cì shiyòng xìnyòngka, yibiàn baituo wèi jing xuke de fèiyòng. Yinwèi wo de zhàngdan zhong you henduo zhàngdan shì tongguò zhipiào zhànghù zìdòng zhuanzhàng de, suoyi dang wo gaibiàn zhànghù shí, fùkuan fangshì yánzhòng shòu sun.
Hb9::RFP+ iMNs appeared between days 13 and 16 after retroviral transduction. RepSox was removed on day 17 and the survival assay was initiated. For the glutamate treatment condition, 10 μM glutamate was added to the culture medium on day 17 and removed after 12 hours. Cells were then maintained in N3 medium with neurotrophic factors without RepSox. Cultures were treated with 10 nM inactive or active 3K3A-APC after glutamate was removed. The inactive and active 3K3A-APC were maintained for the remainder of the experiment and replenished every other day. For treatment with rapamycin, 10 μM DMSO or rapamycin (Sigma-Aldrich, R8781-200UL) was initiated after the 12-hour glutamate treatment was completed. For PAR1 and PAR2 antagonist treatment, cultures were cotreated with 3 μM PAR1 or PAR2 antagonist starting after glutamate treatment. For PAR1, PAR2, and PAR3 ASO treatment, the cultures were pretreated one time with 9 μM ASOs for 72 hours before the pulse glutamate treatment. PAR1 and PAR2 antagonists were purchased from Tocris (RWJ56110, catalog 2614 and AC55541, catalog 3369, respectively). PAR1, PAR2, and PAR3 ASO gapmers were designed and produced by IDT; they contained 2′-O-Me and phosphorothioate linkage modifications. Longitudinal tracking was performed by imaging neuronal cultures in a Nikon Biostation CT or Molecular Devices ImageExpress once every 24 to 72 hours starting on day 17. Tracking of neuronal survival was performed using SVcell 3.0 (DRVision Technologies) or ImageJ. Neurons were scored as dead when their soma was no longer detectable by RFP fluorescence. All neuron survival assays were performed at least twice, with equal numbers of neurons from 3 individual replicates from one of the trials being used for the quantification shown. All trials quantified were representative of other trials of the same experiment. When iMNs from multiple independent donors were combined into one survival trace in the Kaplan-Meier plots for clarity, this is noted in the figure legend.
We hypothesized that phenotypic analysis of iMNs from these sporadic ALS patients and C9ORF72 ALS/frontotemporal dementia (FTD) patients would enable the identification of degenerative mechanisms that are shared by C9ORF72 and at least a subset of sporadic ALS motor neurons. We further hypothesized that pharmacologic agents capable of normalizing these phenotypes in C9ORF72 and sporadic ALS iMNs might also rescue neurodegeneration and lead to the identification of therapeutic targets with broad efficacy in the ALS population.
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