Hepatocellular carcinoma mouse models:Hepatitis B virusassociatedhepatocarcinogenesis and haploinsufficienttumor suppressor genes

The multifactorial and multistage pathogenesis of hepatocellular carcinoma(HCC)has fascinated a wide spectrum of scientists for decades.While a number of major risk factors have been identified,their mechanistic roles in hepatocarcinogenesis still need to be elucidated.Many tumor suppressor genes(TSGs)have been identified as being involved in HCC.These TSGs can be classified into two groups depending on the situation with respect to allelic mutation/loss in the tumors:the recessive TSGs with two required mutated alleles and the haploinsufficient TSGs with one required mutated allele.Hepatitis B virus(HBV)is one of the most important risk factors associated with HCC.Although mice cannot be infected with HBV due to the narrow host range of HBV and the lack of a proper receptor,one advantage of mouse models for HBV/HCC research is the numerous and powerfulgenetic tools that help investigate the phenotypic effects of viral proteins and allow the dissection of the dose-dependent action of TSGs.Here,we mainly focus on the application of mouse models in relation to HBV-associated HCC and on TSGs that act either in a recessive or in a haploinsufficient manner.Discoveries obtained using mouse models will have a great impact on HCC translational medicine.

Mouse models for the discovery of colorectal cancer drivergenes

Colorectal cancer(CRC) constitutes a major publichealth problem as the third most commonly diagnosed and third most lethal malignancy worldwide. The prevalence and the physical accessibility to colorectal tumors have made CRC an ideal model for the study of tumor genetics. Early research efforts using patient derived CRC samples led to the discovery of several highly penetrant mutations(e.g., APC, KRAS, MMR genes) in both hereditary and sporadic CRC tumors. This knowledge has enabled researchers to develop genetically engineered and chemically induced tumor models of CRC, both of which have had a substantial impact on our understanding of the molecular basis of CRC. Despite these advances, the morbidity and mortality of CRC remains a cause for concern and highlight the need to uncover novel genetic drivers of CRC. This review focuses on mouse models of CRC with particular emphasis on a newly developed cancer gene discovery tool, the Sleeping Beauty transposon-based mutagenesis model of CRC.

CMT1A current gene therapy approaches and promising biomarkers

Charcot-Marie-Tooth neuropathies(CMT)constitute a group of common but highly heterogeneous,non-syndromic genetic disorders affecting predominantly the peripheral nervous system.CMT type 1A(CMT1A)is the most frequent type and accounts for almost~50%of all diagnosed CMT cases.CMT1A results from the duplication of the peripheral myelin protein 22(PMP22)gene.Overexpression of PMP22 protein overloads the protein folding apparatus in Schwann cells and activates the unfolded protein response.This leads to Schwann cell apoptosis,dys-and de-myelination and secondary axonal degeneration,ultimately causing neurological disabilities.During the last decades,several different gene therapies have been developed to treat CMT1A.Almost all of them remain at the pre-clinical stage using CMT1A animal models overexpressing PMP22.The therapeutic goal is to achieve gene silencing,directly or indirectly,thereby reversing the CMT1A genetic mechanism allowing the recovery of myelination and prevention of axonal loss.As promising treatments are rapidly emerging,treatment-responsive and clinically relevant biomarkers are becoming necessary.These biomarkers and sensitive clinical evaluation tools will facilitate the design and successful completion of future clinical trials for CMT1A.

Analysis of Proteotranscriptomics Landscape Reveals Differentially Regulated Pathways in Toxoplasma gondii Infected Mouse Liver

Toxoplasma gondii (T. gondii) an intracellular protozoan parasite, infects mammals including human population world-wide. Upon primary infection, the parasite contributes to mild flu like symptoms in immune competent host, but life threatening complication is seen in immune compromised patients and in pregnant women. Understanding the host-parasite interaction is critical for understanding the pathogenesis and biology parasite reactivation in the host. In this study, we used proteotrasncriptomics analyses by integrating the transcriptomics and proteomics data of T. gondii infected mouse liver to uncover the effector molecules responsible for disease pathogenesis that can be used as candidate markers for diagnosis and drug target. With this aim, we systematically integrated transcriptomicand proteomic data, representing the parasite infected mouse liver. Out of 2758 differentially expressed genes (DEGs) and 301 differentially expressed proteins (DEPs), 159 overlapping genes were identified. Among them, 86 genes were upregulated and 72 were downregulated in their respective mRNA and protein levels in the infected condition. Gene Ontology (GO) analysis revealed that the upregulated genes were mostly associated with immune system processes whereas the downregulated genes were involved in oxidation-reduction process and metabolism of lipid, and fatty acids. Protein-protein interaction (PPI) network analysis uncovered an interaction-hub including, Psmb8, Psmb9 and Tap1 for upregulated proteins and Cyp1A2, Cyp4A10 and Cyp3A11 for down-regulated proteins. Further studies are needed to validating these effector molecules. These molecules are likely to play a vital role in disease pathogenesis, as well as can be used as potential diagnostic marker and drug target candidates.

两种肌少症小鼠模型的功能表型、肌肉质量及力量特点比较

背景:地塞米松和后肢悬吊是常用的动物肌少症造模方法,具有造模时间短、操作简便、成本低等特点。目的:比较地塞米松和后肢悬吊诱导小鼠肌少症模型在肌肉质量、力量及功能表型以及分子机制表型方面的差异。方法:采用随机抽样法将30只C57BL/6小鼠随机分成3组,每组10只:正常对照组不进行任何干预;地塞米松组小鼠腹腔注射地塞米松磷酸钠溶液1 mg/(kg•d),连续注射6周,建立肌少症模型;后肢悬吊组采用鼠尾套悬吊小鼠后肢16 h,1次/d,连续悬吊6周,建立肌少症模型。造模6周内,监测小鼠体质量变化;造模6周后,检测小鼠四肢抓力、活动能力(游泳实验)、骨骼肌湿质量、骨骼肌病理形态,采用RT-PCR与Western blot检测骨骼肌蛋白质合成与分解代谢指标、AMPK/FoXO3α信号通路的表达。结果与结论:①从造模后第2周开始,地塞米松组、后肢悬吊组小鼠体质量均低于正常对照组(P<0.05)。②造模6周后,与正常对照组相比,造模两组小鼠四肢抓力均下降(P<0.05),腓肠肌湿质量、趾长伸肌湿质量、腓肠肌与比目鱼肌横截面积均下降(P<0.05);地塞米松组小鼠腓肠肌横截面积明显小于后肢悬吊组(P<0.05),比目鱼肌横截面积大于后肢悬吊组(P<0.05);与正常对照组、后肢悬吊组相比,地塞米松组小鼠活动能力降低(P<0.05)。③与正常对照组相比,造模两组PI3K、mTOR、AMPK、PGC-1αmRNA表达与p-AMPK/AMPK蛋白均降低(P<0.05),FoXO3αmRNA表达与PGC-1α、FoXO3蛋白表达均升高(P<0.05);地塞米松组Akt1 mRNA表达降低(P<0.05),Atrogin-1、MuRF-1 mRNA表达升高(P<0.05);后肢悬吊组Akt1 mRNA表达升高(P<0.05)。④与地塞米松组相比,后肢悬吊组mTOR、Akt1、FoXO3αmRNA表达升高(P<0.05),Atrogin-1、MuRF-1 mRNA表达降低(P<0.05)。⑤结果表明,两种造模方法均会导致骨骼肌线粒体能量代谢水平下降,地塞米松组通过泛素蛋白酶体和能量代谢途径的双重作用介导骨骼肌发生萎缩,后肢悬吊组通过AMPK/FoXO3α信号通路介导能量代谢途径引起骨骼肌发生萎缩,从而引起骨骼肌的质量、力量及功能下降。

Ⅰ型毛-肝-肠综合征小鼠模型构建及表型分析

目的 建立基于Ttc37(Tetratricopeptide Repeat Domain 37)基因缺失的Ⅰ型毛-肝-肠综合征(trichohepato-enteric syndrome,THES)的小鼠疾病模型。方法 利用CRISPR/CAS9技术在小鼠Ttc37基因中插入loxP序列构建Ttc37flox品系,通过与全身表达的CAG-Cre品系交配产生全身敲除小鼠Ⅰ型THES动物模型(Ttc37flox/flox;CAG-Cre),利用荧光定量PCR和Western blot确认敲除效果。选取8周龄小鼠,对皮肤、脾脏、肝脏、膀胱和胃肠道等主要组织进行苏木精-依红染色和病理分析,对血清中的谷草转氨酶(aspartate aminotransferase,AST)和谷丙转氨酶(alanine aminotransferase,ALT)进行酶学检测,对血清中的血红蛋白进行检测,注射抗原后利用ELISA检测免疫球蛋白IgM和IgG水平。结果 Ttc37flox/flox;CAG-Cre表现出与Ⅰ型THES相似的毛发、皮肤、B细胞和眼睛发育异常表型;常规饲养条件下未表现出肝脏、胃肠道、膀胱和血红蛋白异常。结论 Ⅰ型THES的小鼠疾病模型构建成功,可用于病理机制研究。

Aqp5-C末端截断小鼠的构建及其潜在应用

目的 Aqp5蛋白的C末端结构对其功能的发挥起着重要作用。本研究旨在通过构建Aqp5-C末端6个氨基酸缺失的突变小鼠,验证Aqp5-C末端6个氨基酸缺失对该蛋白亚细胞定位的影响,并初步比较研究此突变小鼠的肺脏组织结构和转录组的改变,探讨该突变小鼠用于Aqp5功能研究的潜在应用价值。方法 采用基于gRNA的CRISPER Cas 9技术构建Aqp5末端6个氨基酸缺失的突变小鼠(Aqp5-p.I260*)并进行序列鉴定,采用组织切片、荧光染色和显微镜观察等手段明确突变蛋白在肺泡上皮细胞的亚细胞定位和突变小鼠肺组织结构,采用转录组测序研究突变小鼠肺脏基因差异表达。结果 本研究成功构建了Aqp5蛋白C末端截断小鼠,发现p.I260*改变了亚细胞定位,Aqp5-p.I260*鼠肺泡数目少于野生鼠;Aqp5-p.I260*小鼠肺组织的转录组上调基因主要富集于发育和造血细胞谱系等通路,其Aqp5功能缺陷后诱发炎症相关通路基因(Hspa1a、IL-1β、Col3a1等)的表达水平改变。结论 本研究在体验证了小鼠Aqp5蛋白C末端6个氨基酸缺失突变对该蛋白细胞膜定位的关键作用,表明了以该小鼠作为Aqp5功能缺陷模型探讨Aqp5蛋白在肺发育和骨髓造血、炎症和免疫调节等生物学功能的潜在应用价值。

Efficient generation of the mouse model with a defined point mutation through haploid cell-mediated gene editing

Generation of mouse models carrying a defined point mutation,especially disease-related point mutations,is of considerable interest for research in biology and medicine.The standard method based on embryonic stem cell（ESC）-mediated homologous recombination（HR）is time-and labor-consuming.

Mapping novel genetic loci associated with female liver weight variations using Collaborative Cross mice

Background: Liver weight is a complex trait, controlled by polygenic factors and differs within populations. Dissecting the genetic architecture underlying these variations will facilitate the search for key role candidate genes involved directly in the hepatomegaly process and indirectly involved in related diseases etiology.Methods: Liver weight of 506 mice generated from 39 different Collaborative Cross(CC) lines with both sexes at age 20 weeks old was determined using an electronic balance. Genomic DNA of the CC lines was genotyped with high-density single nucleotide polymorphic markers.Results: Statistical analysis revealed a significant(P < 0.05) variation of liver weight between the CC lines, with broad sense heritability(H^2) of 0.32 and genetic coefficient of variation(CV_G) of 0.28. Subsequently, quantitative trait locus(QTL) mapping was performed, and results showed a significant QTL only for females on chromosome 8 at genomic interval 88.61-93.38 Mb(4.77 Mb). Three suggestive QTL were mapped at chromosomes 4, 12 and 13. The four QTL were designated as LWL1-LWL4 referring to liver weight loci 1-4 on chromosomes 8, 4, 12 and 13,respectively.Conclusion: To our knowledge, this report presents, for the first time, the utilization of the CC for mapping QTL associated with baseline liver weight in mice. Our findings demonstrate that liver weight is a complex trait controlled by multiple genetic factors that differ significantly between sexes.

Identification in Chinese patients with GLIALCAM mutations of megalencephalic leukoencephalopathy with subcortical cysts and brain pathological study on Glialcam knock-in mouse models

Background Megalencephalic leukoencephalopathy with subcortical cysts(MLC)is a rare neurological degenerative disorder caused by the mutations of MLC1 or GLIALCAM with autosomal recessive or autosomal dominant inheritance and a different prognosis,characterized by macrocephaly,delayed motor and cognitive development,and bilateral abnormal signals in cerebral white matter(WM)with or without cysts on magnetic resonance imaging(MRI).This study aimed to reveal the clinical and genetic features of MLC patients with GLIALCAM mutations and to explore the brain pathological characteristics and prognosis of mouse models with different modes of inheritance.Methods Clinical information and peripheral venous blood were collected from six families.Genetic analysis was performed by Sanger sequencing of GLIALCAM.Glialcam^(Arg92Trp/+)and Glialcam^(Lys68Met/Thr132Asn)mouse models were generated based on mutations from patients(c.274C>T(p.Arg92Trp)(c.203A>T(p.Lys68Met),and c.395C>A(p.Thr132Asn))).Brain pathologies of the mouse models at different time points were analyzed.Results Six patients were clinically diagnosed with MLC.Of the six patients,five(Pt1-Pt5)presented with a heterozygous mutation in GLIALCAM(c.274C>T(p.Arg92Trp)or c.275G>C(p.Arg92Pro))and were diagnosed with MLC2B;the remaining patient(Pt6)with two compound heterozygous mutations in GLIALCAM(c.203A>T(p.Lys68Met)and c.395C>A(p.Thr132Asn))was diagnosed with MLC2A.The mutation c.275C>G(p.Arg92Pro)has not been reported before.Clinical manifestations of the patient with MLC2A(Pt6)progressed with regression,whereas the course of the five MLC2B patients remained stable or improved.The Glialcam^(Arg92Trp/+)and Glialcam^(Lys68Met/Thr132Asn)mouse models showed vacuolization in the anterior commissural WM at 1 month of age and vacuolization in the cerebellar WM at 3 and 6 months,respectively.At 9 months,the vacuolization of the GlialcamiLys68Met/Thr132Asn mouse model was heavier than that of the Glialcam^(Arg92Trp/+)mouse model.Decreased expression of Glialcam in Glialcam^(Arg92Trp/+)and Glialcam^(Lys68Met/Thr132Asn)mice may contribute to the vacuolization.Conclusions Clinical and genetic characterization of patients with MLC and GLIALCAM mutations revealed a novel mutation,expanding the spectrum of GLIALCAM mutations.The first Glialcam mouse model with autosomal recessive inheritance and a new Glialcam mouse model with autosomal dominant inheritance were generated.The two mouse models with different modes of inheritance showed different degrees of brain pathological features,which were consistent with the patients'phenotype and further confirmed the pathogenicity of the corresponding mutations.

LRRK2 mutant knock-in mouse models: therapeutic relevance in Parkinson’s disease

Mutations in the leucine-rich repeat kinase 2 gene (LRRK2) are one of the most frequent genetic causes of both familial and sporadic Parkinson’s disease (PD). Mounting evidence has demonstrated pathological similarities between LRRK2-associated PD (LRRK2-PD) and sporadic PD, suggesting that LRRK2 is a potential disease modulator and a thera-peutic target in PD. LRRK2 mutant knock-in (KI) mouse models display subtle alterations in pathological aspects that mirror early-stage PD, including increased susceptibility of nigrostriatal neurotransmission, development of motor and non-motor symptoms, mitochondrial and autophagy-lysosomal defects and synucleinopathies. This review provides a rationale for the use of LRRK2 KI mice to investigate the LRRK2-mediated pathogenesis of PD and implications from current findings from different LRRK2 KI mouse models, and ultimately discusses the therapeutic potentials against LRRK2-associated pathologies in PD.

Organ-specific alterations in circadian genes by vertical sleeve gastrectomy in an obese diabetic mouse model

Current therapies for obesity and related complications have been shown to have limited benefits,including unsatisfactory weight loss and poor metabolic improvement.With recent developments in bariatric surgery,promising advancements have been made in clinical and scientific research,particularly in the management of obesity and diabetes.Vertical sleeve gastrectomy(VSG)has become increasingly popular due to its safety,simplicity,

CRISPR/Cas9 mediated somatic gene therapy for insertional mutations:the vibrator mouse model

Somatic gene therapy remains technically challenging,especially in the central nervous system(CNS).Efficiency of gene delivery,efficacy in recipient cells,and proportion of cells required for overall benefit are the key points needed to be considered in any therapeutic approach.Recent efforts have demonstrated the efficacy of RNA-guided nucleases such as CRISPR/Cas9 in correcting point mutations or removing dominant mutations.Here we used viral delivered Cas9 plasmid and two guide RNAs to remove a recessive insertional mutation,vibrator(vb),in the mouse brain.The vb mice expressed∼20%of normal levels of phosphatidylinositol transfer protein,α(PITPα)RNA and protein due to an endogenous retrovirus inserted in intron 4,resulting in early-onset tremor,degeneration of brainstem and spinal cord neurons,and juvenile death.The in situ CRISPR/Cas9 viral treatment effectively delayed neurodegeneration,attenuated tremor,and bypassed juvenile death.Our studies demonstrate the potential of CRISPR/Cas9-mediated gene therapy for insertional mutations in the postnatal brain.

Using Huntingtin Knock-In Minipigs to Fill the Gap Between Mouse Models and Patients with Huntington's Disease

Huntington＇s disease （HD） is an inherited autosomal dominant neurodegenerative disease characterized by pro- gressive motor deficits, cognitive decline, and psychiatric symptoms. It is caused by a pathological expansion of CAG trinucleotide repeats in exon 1 of the HD gene, resulting in the translation of a mutant form of huntingtin protein （mutant Htt） with an expanded polyglutamine domain in the N-terminal region [1 ]. Despite great progress in understanding the pathogenesis of HD using multiple mouse models, the exact mechanisms by which mutant Htt induces neuronal dysfunction and death are still not completely clear, and there is no curative treatment for this disease. An important reason is that the mouse, which is the most widely used animal model in HD research, differs from the human in many aspects, including the physiology, drug metabolism, blood-brain barrier, life span, brain volume, and neuroanatomical organization [2]. Thus, it is necessary to establish HD models with higher species than rodents, such as the dog, pig, and non- human primate, so as to bridge the gap between preclinical mouse models and clinical studies.

Generation and characterization of Men1 mutant mouse models for studying MEN1 disease

Patients with multiple endocrine neoplasia type 1(MEN1)mutations are predisposed to MEN1 syndrome affecting various endocrine cell lineages.Following its identification in the late 1990s,laboratories around the world,including our own,used gene-targeting approaches in murine models to study the MEN1 gene and its related diseases.Subsequently,this field of research witnessed an upsurge in the use of Men1 mutant mouse models to dissect MEN1 functions.These studies led to unraveling the natural history of MEN disease,and highlighted cellular and molecular mechanisms underlying the development of the disease.In this review,we present the currently available data concerning the generation and characterization of Men1 mutant mouse models in connection with MEN1 syndrome.

吸入气管内雾化脂多糖致急性肺损伤小鼠肺组织转录组分析

背景急性肺损伤(acute lung injury,ALI)/急性呼吸窘迫综合征(acute respiratory distress syndrome,ARDS)是危重症医学和基础医学研究重要的关注焦点。明确ALI时关键损伤机制及差异性基因的变化,对探究ALI的发生发展至关重要。目的研究气管内雾化脂多糖(lipopolysaccharides,LPS)致ALI小鼠肺组织的转录组基因谱变化,探索ALI潜在损伤机制及治疗靶点。方法36只小鼠随机分为对照组和气管内雾化LPS组(时间分为致伤后6 h、12 h、24 h,每组各9只)。实验组给予气管内雾化50μL的LPS 15 mg/kg,对照组给予气管内雾化等体积0.9%氯化钠注射液。比较组间组织病理学、病理损伤评分及肺系数变化,检测肺泡灌洗液(bronchoalveolar lavage fluid,BALF)及血清中肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)、白细胞介素-6(interleukin-6,IL-6)、单核细胞趋化蛋白-1(monocyte chemoattractant protein-1,MCP-1)等水平。肺组织行转录组测序,寻找差异基因,并对上调、下调基因分别进行GO和KEGG富集分析,对持续性改变的差异基因进行GO功能富集整合及共表达网络分析。结果与对照组相比,吸入气管内雾化LPS后小鼠肺组织炎症细胞浸润、水肿加重伴出血,肺损伤评分及肺系数评分逐渐增高,24 h时损伤最严重;TNF-α、IL-6、MCP-1的表达均显著升高(P<0.01),BALF与血浆中表达存在一定差异;转录组学提示12 h组上调基因1545个、下调基因1670个,24 h组上调基因1312个、下调基因1139个。KEGG富集分析显示差异基因涉及炎症反应、遗传信息调控、信号转导等通路,GO富集分析显示差异基因集中在细胞因子生成及免疫反应调控、白细胞的免疫介导及免疫调节、白细胞迁移(上调基因),以及血液循环、信号传导、神经突触、肌肉收缩等(下调基因)。PPI分析提示持续性上调的关键基因包括Tnf、IL-1β、Ifn-γ、Ccl2、Ccl5、Nod2、Tlr2、Lgals9。而持续性下调的关键基因包括Cav1、Sulf1、Sox4、Tgf、Apoe、Tek。结论气管内雾化LPS 12 h建立稳定、可靠的小鼠急性肺损伤模型。Tnf、IL-1β、Ifn-γ、Ccl2、Ccl5、Nod2、Tlr2、Lgals9等关键基因可能是ALI中免疫与炎症反应中的关键调节因子和靶基因。

CRISPR/Cas9技术构建MMACHC基因c.80A>G(p.Q27R)纯合突变甲基丙二酸血症小鼠模型

目的利用CRISPR/Cas9技术构建MMACHC基因c.80A>G(p.Q27R)纯合突变的cblC型甲基丙二酸血症合并同型半胱氨酸血症小鼠模型。方法利用CRIPSR/Cas9基因编辑技术,将小鼠MMACHC基因第80位碱基由碱基A突变为碱基G,在靶位点区域设计单链导向RNA(single-guide RNA,sgRNA),构建打靶载体,将Cas9/sgRNA及打靶载体显微注射到小鼠受精卵中,培育获得F0代小鼠,以小鼠鼠尾鉴定基因型,将点突变阳性F0代小鼠与野生型小鼠交配获得具有稳定基因型的F1代小鼠,F1代小鼠交配繁育获得MMACHC基因c.80A>G纯合突变型小鼠,对纯合型小鼠、杂合型小鼠、野生型小鼠进行血代谢产物甲基丙二酸和总同型半胱氨酸检测。结果获得12只MMACHC基因c.80A>G点突变阳性的F1代小鼠,并进行培育繁殖扩大种群,获得纯合突变型小鼠,纯合型小鼠无早期死亡,其血甲基丙二酸、总同型半胱氨酸水平显著高于杂合型和野生型小鼠(P<0.05),杂合型和野生型小鼠血甲基丙二酸、总同型半胱氨酸水平无差异(P>0.05)。结论利用CRISPR/Cas9技术成功构建MMACHC基因c.80A>G(p.Q27R)纯合突变的cblC型甲基丙二酸血症合并同型半胱氨酸血症小鼠模型。

裸鼠脾脏移植瘤模型在NGX6抗结肠癌转移作用研究中的应用

目的:建立结肠癌肝转移模型,为抑瘤基因NGX6功能研究提供体内试验平台。方法:将低分化结肠癌细胞系HT-29组(HT-29)、空白质粒转染组[pcDNA3.1(+)/HT-29]、NGX6转染组[(pcDNA3.1(+)/NGX6/HT-29)]细胞分别接种于裸鼠脾脏下极包膜内,每天称量裸鼠体质量并观察裸鼠摄食、活动及精神情况,45d后处死裸鼠,分别从大体水平以及镜下观察肝、脾、肺、肾、脑等器官肿瘤转移情况。结果:pcDNA3.1(+)/NGX6/HT-29组裸鼠较其他两组裸鼠肝脏转移灶数目明显减少、脾脏上种植瘤形成明显减少。结论:抑瘤基因NGX6抑制结肠癌HT-29细胞的转移,裸鼠脾内种植转移模型可作为新基因体内功能研究的理想模型。

基因敲除鼠疾病模型的研究进展

基因敲除是研究生物体基因功能的有效手段。通过基因敲除建立的鼠疾病模型,在研究基因功能及人类疑难病症致病机制等方面发挥着前所未有的作用。本文对目前已获得的基因敲除鼠疾病模型进行了分类和总结,为相关研究的展开奠定了基础。

采用活体成像技术监测肿瘤生长及转移模型的建立

目的采用活体成像技术监测稳定高表达荧光素酶报告基因的肿瘤细胞在小鼠体内生长及转移情况,为肿瘤治疗的药物研发提供新的有用工具。方法采用lipofectamine2000介导的基因转染方法,将pcDNA3·1/Luc载体转染小鼠高转移乳腺癌细胞株4T1、EMT-6及结肠癌细胞株CT26,经G418抗性筛选及有限稀释法获得可稳定高表达荧光素酶的单克隆细胞;MTT法测定各转染细胞对不同化疗药物的抗性,并采用活体成像的方法检测各转染细胞在小鼠体内的成瘤和转移。结果获得了可稳定高表达荧光素酶基因的单克隆细胞株,该单克隆细胞株具有与亲本细胞系相同的对化疗药物的敏感性;将单克隆细胞株植入小鼠皮下,可采用活体成像技术准确监测肿瘤细胞体内生长及转移。结论采用活体成像技术构建的肿瘤动物模型是拓展肿瘤体内生长、转移及治疗相关研究的理想模型。