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Knowledge-embedded spatio-temporal analysis for euploidy embryos identification in couples with chromosomal rearrangements

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Chinese Medical Journal 2024年第6期137卷 694-703页

作者： Fangying Chen Xiang Xie Du Cai Pengxiang Yan Chenhui Ding Yangxing Wen Yanwen Xu Feng Gao canquan zhou Guanbin Li Qingyun Mai Reproductive Medicine Center The First Affiliated Hospital of Sun Yat-sen UniversityGuangzhouGuangdong 510080China Guangdong Provincial Key Laboratory of Reproductive Medicine GuangzhouGuangdong 510080China School of Computer Science and Engineering Sun Yat-sen UniversityGuangzhouGuangdong 510006China Department of General Surgery(Colorectal Surgery) The Sixth Affiliated Hospital of Sun Yat-sen UniversityGuangzhouGuangdong 510655China Guangdong Provincial Key Laboratory of Colorectal and Pelvic Floor Diseases The Sixth Affiliated HospitalSun Yat-sen UniversityGuangzhouGuangdong 510655China Biomedical Innovation Center The Sixth Affiliated HospitalSun Yat-sen UniversityGuangzhouGuangdong 510655China Shanghai Artificial Intelligence Laboratory Shanghai 200232China

Background:The goal of the assisted reproductive treatment is to transfer one euploid blastocyst and to help infertile women giving birth one healthy *** algorithms have been used to assess the ploidy status of embryos derived from couples with normal chromosome,who subjected to preimplantation genetic testing for aneuploidy(PGT-A)***,it is currently unknown whether artificial intelligence model can be used to assess the euploidy status of blastocyst derived from populations with chromosomal ***:From February 2020 to May 2021,we collected the whole raw time-lapse videos at multiple focal planes from in vitro cultured embryos,the clinical information of couples,and the comprehensive chromosome screening results of those blastocysts that had received PGT ***,we developed a novel deep learning model called the Attentive Multi-Focus Selection Network(AMSNet)to analyze time-lapse videos in real time and predict blastocyst *** upon AMSNet,we integrated additional clinically predictive variables and created a second deep learning model,the Attentive Multi-Focus Video and Clinical Information Fusion Network(AMCFNet),to assess the euploidy status of *** efficacy of the AMCFNet was further tested in embryos with parental chromosomal *** receiver operating characteristic curve(ROC)was used to evaluate the superiority of the ***:A total of 4112 embryos with complete time-lapse videos were enrolled for the blastocyst formation prediction task,and 1422 qualified blastocysts received PGT-A(n=589)or PGT for chromosomal structural rearrangement(PGT-SR,n=833)were enrolled for the euploidy assessment task in this *** AMSNet model using seven focal raw time-lapse videos has the best real-time *** real-time accuracy for AMSNet to predict blastocyst formation reached above 70%on the day 2 of embryo culture,and then increased to 80%on the day 4 of embryo ***

关键词： Euploidy status Embryo Time-lapse imaging Deep learning Spatio-temporal analysis Preimplantation genetic testing for chromosomal structural rearrangement

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Correction of β-thalassemia mutant by base editor in human embryos

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Protein & Cell 2017年第11期8卷 811-822页

作者： Puping Liang Chenhui Ding Hongwei Sun Xiaowei Xie Yanwen Xu Xiya Zhang Ying Sun Yuanyan Xiong Wenbin Ma Yongxiang Liu Yali Wang Jianpei Fang Dan Liu zhou Songyang canquan zhou Junjiu Huang Key Laboratory of Gene Engineering of the Ministry of Education Guangzhou Key Laboratory of Healthy Aging Research and State Key Laboratory of Biocontrol SYSU-BCM Joint Research Center School of Life Sciences Sun Yat-sen University Guangzhou 510275 China Key Laboratory of Reproductive Medicine of Guangdong Province School of Life Sciences and the First Affiliated Hospital Sun Yat-sen University Guangzhou 510275 China Department of Pediatrics Second Affiliated Hospital Sun Yat-sen University Guangzhou 510120 China Verna and Marrs Mclean Department of Biochemistry and Molecular Biology Baylor College of Medicine One Baylor Plaza Houston. TX 77030. USA

β-Thalassemia is a global health issue, caused by mutations in the HBB gene. Among these mutations, HBB -28 （A〉G） mutations is one of the three most common mutations in China and Southeast Asia patients with β-thalassemia. Correcting this mutation in human embryos may prevent the disease being passed onto future generations and cure anemia. Here we report the first study using base editor （BE） system to correct disease mutant in human embryos. Firstly, we produced a 293T cell line with an exogenous HBB -28 （A〉G） mutant fragment for gRNAs and targeting efficiency evaluation. Then we collected primary skin fibroblast cells from a β-thalassemia patient with HBB -28 （A〉G） homozygous mutation. Data showed that base editor could precisely correct HBB -28 （A〉G） mutation in the patient＇s primary cells. To model homozygous mutation disease embryos, we consb＇ucted nuclear transfer embryos by fusing the lymphocyte or skin fibroblast cells with enucleated in vitro matured （IVM） ***, the gene correction efficiency was over 23.0% in these embryos by base editor. Although these embryos were still mosaic, the percentage of repaired blastomeres was over 20.0%. In addition, we found that base editor variants, with narrowed deamination window, could promote G-to-A conversion at HBB -28 site precisely in human embryos. Collectively, this study demonstrated the feasibility of curing genetic disease in human somatic cells and embryos by base editor system.

关键词： β-thalassemia HBB -28 (A〉G) baseeditor human embryo

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CRISPR/Cas9-mediated gene editing in human tripronuclear zygotes

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Protein & Cell 2015年第5期6卷 363-372页

作者： Puping Liang Yanwen Xu Xiya Zhang Chenhui Ding Rui Huang Zhen Zhang Jie Lv Xiaowei Xie Yuxi Chen Yujing Li Ying Sun Yaofu Bai zhou Songyang Wenbin Ma canquan zhou Junjiu Huang Guangdong Province Key Laboratory of Reproductive Medicine the First Affiliated Hospital and Key Laboratory of GeneEngineering of the Ministry of Education School of Life Sciences Sun Yat-sen University Guangzhou 510275 China

Genome editing tools such as the clustered regularly interspaced short palindromic repeat （CRISPR）-associated system （Cas） have been widely used to modify genes in model systems including animal zygotes and human cells, and hold tremendous promise for both basic research and clinical applications. To date, a serious knowledge gap remains in our understanding of DNA repair mechanisms in human early embryos, and in the efficiency and potential off-target effects of using technologies such as CRISPR/Cas9 in human pre-implantation embryos. In this report, we used tripronuclear （3PN） zygotes to further investigate CRISPR/Cas9-mediated gene editing in human cells. We found that CRISPR/Cas9 could effectively cleave the endogenous β-globin gene （HBB）. However, the efficiency of homologous recombination directed repair （HDR） of HBB was low and the edited embryos were mosaic. Off-target cleavage was also apparent in these 3PN zygotes as revealed by the T7E1 assay and whole-exome sequencing. Furthermore, the endogenous delta-globin gene （HBD）, which is homologous to HBB, competed with exogenous donor oligos to act as the repair template, leading to untoward mutations. Our data also indicated that repair of the HBB locus in these embryos occurred preferentially through the non-crossover HDR pathway. Taken together, our work highlights the pressing need to further improve the fidelity and specificity of the CRISPR/Cas9 platform, a prerequisite for any clinical applications of CRSIPR/Cas9-mediated editing.

关键词： CRISPR/Cas9 β-thalassemia humantripronuclear zygotes gene editing homologousrecombination whole-exome sequencing

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The Clinic Analysis of Diclofenac Suppository for Oocyte Retrieval Analgesia in IVF-ET Cycles

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International Journal of Clinical Medicine 2013年第8期4卷 350-354页

作者： Yubin Li Qingyun Mai Tao Li Yiping Zhong canquan zhou The Reproductive Center The First Affiliated Hospital of Sun Yat-sen University Guangzhou China

Objective: To study the effect of diclofenac suppository in oocyte retrieval of IVF-ET. Study Design: 1176 patients with informed consents were enrolled into this prospective randomized controlled study. The setting was an IVF-ET program at the First Affiliated Hospital of Sun Yat-sen University, Guangzhou, China. According to the analgesic drug use, the patients were randomly divided into pethidine group (573 cases) and diclofenac suppository group (603 cases). The data of vital signs, common adverse reactions, severe adverse events and pain degree in oocyte retrieval were collected. The IVF-ET outcomes were also compared. Results: The post-operation pressure and pulse were lower in pethidine group than in diclofenac suppository group (both P0.001).The rest vital signs were not statistically different (all P>0.05). Common adverse reactions in diclofenac suppository group were relative less (all P0.05). Pain degree between the two groups was not statistically different (P=0.304). IVF-ET outcomes were also not statistically different (all P>0.05). There were 3 cases serious abdominal bleeding with shock in the diclofenac suppository group. Conclusion: Using diclofenac suppository in oocyte retrieval analgesic had a good effect. And there was no adverse effect in the IVF-ET outcome. But we should pay close attention to the probability of serious abdominal bleeding.

关键词： Diclofenac Suppository IVF-ET Oocyte Retrieval Analgesia

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HBB-deficient Macaca fascicularis monkey presents with human β-thalassemia

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Protein & Cell 2019年第7期10卷 538-542页

作者： Yan Huang Chenhui Ding Puping Liang Duanduan Li Yu Tang Wei Meng Hongwei Sun Hongyu Lu Yu Chen Xueying Chen Qunshan Huang Jianpei Fang canquan zhou Shihua Yang Junjiu Huang College of Veterinary Medicine Key Laboratory of Comprehensive Prevention and Control for Severe Clinical Animal Diseases of Guangdong Province South China Agricultural University Guangzhou 510642 China Key Laboratory of Reproductive Medicine of Guangdong Province The First Affiliated Hospital and School of Life Sciences Sun Yatsen University Guangzhou 510275 China MOE Key Laboratory of Gene Function and Regulation State Key Laboratory of Biocontrol School of Life Sciences SunYat-sen University Guangzhou 510275 China Department of Pediatrics Second Affiliated Hospital Sun Yat-sen University Guangzhou 510120 China

Dear Editor,β-Thalassemia is a common severe genetic disease caused by mutations in HBB and affects approximately 1.5% of the global population (Origa, 2017). In southern China, the carrier rate of β-thalassemia is as high as 6.43%, creating a high socio-economic burden (Xiong et al., 2010). In adult humans, there are three types of hemoglobin: HbA1 (~97%), HbA2 (~2%) and HbF (~1%). HbA1 (α2β2) is composed of two a-globin and two β-globi n sub units en coded by HBA and HBB, respectively;HbF (α2β2)is made up of two α-globin subunits and two β-globin sub units en coded by HBG. Mutations in the coding region or regulatory region of HBB are involved in β-thalassemia pathogenesis. Except for some rare dominant mutations, most HBB mutations are recessive (Origa, 2017). Depending on the mutation type, the β-globin level will either be reduced or completely depleted, resulting in α-globin accumulation and precipitation. These α-globin precipitates lead to red blood cell death, resulting in anemia and tissue damage, and even death in thalassemia major patients. Blood transfusions can help slow disease progression but lead to iron overload, ultimately resulting in iron toxicity. Bone marrow transfer is the only cure in the clinic and is available only to a small percentage of patients with human leukocyte antigervmatched donors. Recently, gene therapy and gene editing therapy have shown great promise in curing β-thalassemia (Glaser et al., 2015;Thompson et al., 2018). However, no appropriate animal models are available for evaluating the safety and efficacy of such advanced therapeutic strategies in vivo.β-thalassemia mice are the sole animal model available for research. However, substantial differences have been reported between the types and expressi on patter ns of human and mouse globins (McColl and Vadolas, 2016). Moreover, mice contain no fetal globin gene equivalent, and homozygous mutations of HBB in mouse for early models of β-thalassemia major or Cooley anemia are

关键词： mutation mouse Editor mice most help null sub

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Preimplantation gender diagnosis by fluorescence in situ hybridization

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Chinese Medical Journal 2002年第6期115卷 874-877,152-153页

作者：徐艳文庄广伦舒益民李满周灿权张敏芳中山医科大学附属第一医院妇产科生殖医学研究中心广州510080

Objective To describe the clinical application of fluorescence in situ hybridization (FISH) in preimplantation gender *** Preimplantation gender diagnosis was performed in 2 female hemophilia A carriers, 1 male patient with glucose-6-phosphate dehydrogenase (G-6-PD) deficiency and 2 male patients with Y chromosome abnormality. Embryo sex was identified by FISH in total of 6 treatment *** A total of 123 cumulus-oocytes were retrieved in 6 treatment cycles. Sixty-one embryos were available for embryo biopsy. The success rate of biopsy was 86.9% (53/61), with a further cleavage rate of 62.3% (33/53). In the FISH procedure, one cell was lost during fixation, leading to a 98.1% (52/53) fixation rate. Totally, 16 female embryos and 3 male embryos were transferred to 5 patients in 6 cycles. Three healthy babies were born. The diagnosis was confirmed by subsequent analysis of amniocytes and embryonic buds after embryo *** FISH is an efficient and reliable technique for determining the sex of human preimplantation embryos. Selective abortion and births of affected children can be avoided by preimplantation gender diagnosis.

关键词： Blastocyst In Situ Hybridization, Fluorescence Sex Determination (Analysis) Adult Amniocentesis Biopsy Female Humans Male Pregnancy Research Support, Non-U.S. Gov't

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Genome editing of PAR2 through targeted delivery of CRISPR-Cas9 system for alleviating acute lung inflammation via ERK/NLRP3/IL-1β and NO/iNOS signalling

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Acta Pharmaceutica Sinica B 2024年第3期14卷 1441-1456页

作者： Xin Zhuo Yue Wu Xiujuan Fu Jianbin Li Yuxin Xiang Xiaoyu Liang canquan Mao Yuhong Jiang Sichuan Engineering Research Center for Biomimetic Synthesis of Natural Drugs School of Life Science and EngineeringSouthwest Jiaotong UniversityChengdu 610031China

Excessive and uncontrollable inflammatory responses in alveoli can dramatically exacerbate pulmonary disease progressions through vigorous cytokine releases,immune cell infiltration and protease-driven tissue *** is an urgent need to explore potential drug strategies for mitigating lung ***-activated receptor 2(PAR2)as a vital molecular target principally participates in various inflammatory diseases via intracellular signal ***,it has been rarely reported about the role of PAR2 in lung *** study applied CRISPR-Cas9 system encoding Cas9 and sg RNA(p Cas9-PAR2)for PAR2 knockout and fabricated an anionic human serum albuminbased nanoparticles to deliver p Cas9-PAR2 with superior inflammation-targeting efficiency and stability(TAP/p Cas9-PAR2).TAP/p Cas9-PAR2 robustly facilitated p Cas9-PAR2 to enter and transfect inflammatory cells,eliciting precise gene editing of PAR2 in vitro and in ***,PAR2 deficiency by TAP/p Cas9-PAR2 effectively and safely promoted macrophage polarization,suppressed proinflammatory cytokine releases and alleviated acute lung inflammation,uncovering a novel value of *** also revealed that PAR2-mediated pulmonary inflammation prevented by TAP/p Cas9-PAR2was mainly dependent on ERK-mediated NLRP3/IL-1β and NO/i NOS ***,this work indicated PAR2 as a novel target for lung inflammation and provided a potential nanodrug strategy for PAR2 deficiency in treating inflammatory diseases.

关键词： Protease-activated receptor 2(PAR2) CRISPR-Cas9 Gene editing Inflammation Acute lung inflammation NLRP3 Nanoparticles Drug delivery

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辅助生育技术的新进展

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中国实用妇科与产科杂志 2006年第1期22卷 25-28页

作者：徐艳文周灿权中山大学附属第一医院生殖医学中心广东广州510080

自1978年世界首例试管婴儿诞生以来，全世界范围内已经有超过100万个经体外受精与胚胎移植（in vitro fenilization and embryo transfer，IVF—ET）及其相关技术即辅助生殖生育技术（assisted reproductive technique，ART）出生的婴儿... 详细信息

自1978年世界首例试管婴儿诞生以来，全世界范围内已经有超过100万个经体外受精与胚胎移植（in vitro fenilization and embryo transfer，IVF—ET）及其相关技术即辅助生殖生育技术（assisted reproductive technique，ART）出生的婴儿。ART各方面的进步，已完全改变了人类对自身生殖活动的调控。

关键词：辅助生育技术试管婴儿世界范围内世界首例胚胎移植体外受精辅助生殖相关技术生殖活动 and

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高度重视辅助生殖技术实施中出现的问题

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中国实用妇科与产科杂志 2006年第12期22卷 884-885页

作者：周灿权徐艳文中山大学附属第一医院生殖医学中心广东广州510080

人类辅助生殖技术(assisted reproductive technology,ART)是20世纪最伟大的医学成就之一.迄今为止,通过体外受精和胚胎移植技术(in vitro fertilization-embryo transfer,IVF-ET)分娩的婴儿已超过100000个,在某些发达国家占出生总人数... 详细信息

关键词：人类辅助生殖技术技术实施胚胎移植技术 ART技术医学成就体外受精发达国家科学技术

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Effect of surgical intervention on expression of Leukemia inhibitory factor and L-selectin ligand in endometrium of hydrosalpinx patients in implantation window

Effect of surgical intervention on expression of Leukemia in...

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中华医学会第十次全国妇产科学术会议妇科内分泌会场（妇科内分泌学组、绝经学组、计划生育学组）

作者： Yiping Zhong Jin Li Haitao Wu Ying Ying Yafeng Liu canquan zhou Qiong Wang Yanwen Xu Xiaoting Shen Reproductive Medicine Center The First Affiliated HospitalSun Yat-Sun University Reproductive Medicine Center Guangdong No.2 Provincial People's Hospital

Objective To investigate the effect of surgical intervention on the expression of leukemia inhibitory factor (LIF) and L - selectin ligand in the endometrium of patients with hydrosalpinx in implantation *** A total of 60 patients with hydrosalpinx and 30 patients with tubal obstruction were recruited,and immunohistochemsitry was done to detect the expression of LIF and L - selectin ligand in the endometrium of hydrosalpinx patients before and after surgery and of patients with tubal *** The expression of LIF and L-selectin ligand in the endometrium of hydrosalpinx patients before surgery was markedly lower than that in patients with tubal obstruction(PP > 0.05).In addition,there was pronounced difference in the expression of LIF and L - selectin ligand in the endometrium before and after surgery in patients with hydrosalpinx(PConclusion: Hydrosalpinx reduces the expression of LIF and L - selectin ligand in the endometrium in the implantation window. LIF and L - selectin ligand might be important factors affecting the endometrial receptivity of hydrosalpinx patients and surgery can improve the expression of LIF and L - selectin ligand in the endometrium in the implantation window.

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