IDH突变和1p/19q共缺失型少突胶质细胞瘤临床病理特征和预后分析

目的探讨I D H突变和1p/19q共缺失型少突胶质细胞瘤的临床病理特征及预后相关影响因素。方法收集54例IDH突变和1p/19q共缺失型少突胶质细胞瘤病例,分析其临床病理特点,包括年龄、组织学分级和肿瘤部位等因素对无进展生存期和总生存期的影响。结果 54例患者中,肿瘤发生于1个脑叶者46例,发生于2个脑叶以上者8例。肿瘤组织学WHO分级2级12例,3级42例。FISH检测显示54例均为1p/19q共缺失;免疫组织化学检测显示Olig2均为弥漫强阳性;GFAP均为阳性;p53有6例强阳性;48例患者ATRX未缺失;Ki-67增殖指数5%~60%。Sanger测序显示54例均发生IDH基因突变(40例为IDH1突变,14例为IDH2突变),33例发生TERT启动子突变。16例在治疗过程中发生复发及转移。单因素分析显示,手术后复发转移间隔时间超过2年可以延长患者无进展生存和总生存期。54例患者平均无进展生存期33.5个月,平均总生存期40.7个月。结论 IDH突变和1p/19q共缺失型少突胶质细胞瘤术后联合精准放化疗降低了进展风险,手术后复发转移间隔时间与该型患者预后相关。

Screening of Differently Expressed Genes in Human Prostate Cancer Cell Lines with Different Metastasis Potentials

In order to screen the genes differentially expressed in two human prostate cancer cells with different metastasis potentials, suppression subtractive hybridization （SSH） was done twice on human prostate cancer cell line with high potential of metastasis PC3M-IE8 and its synogenetic cell line PC3M-2B4 with low metastasis potential. In the first subtraction PC3M-2B4 was used as tester and PC3M-1E8 as driver and the forward subtractive library was constructed. In the second one the tester and driver were interchanged and the reverse subtractive library was constructed. The screened clones of both libraries were sequenced and Gene Bank homology search was performed. Some clones were confirmed by quantitative real-time PCR. The results showed that two subtractive libraries containing 238 positive clones were constructed. Analysis of 16 sequenced clones randomly picked from two libraries showed that 4 differentially expressed gene fragments were identified as new EST with unknown functions. It was concluded that two subtractive libraries of human prostate cancer cell lines with different metastasis potentials were constructed successfully.

Identification of Novel Epithelial Ovarian Cancer Biomarkers by Cross-laboratory Microarray Analysis

The purpose of this study was to pool information in epithelial ovarian cancer by combining studies using Affymetrix expression microarray datasets made at different laboratories to identify novel biomarkers.Epithelial microarray expression information across laboratories was screened and combined after preprocessing raw microarray data,then ANOVA and unpaired T test statistical analysis was performed for identifying differentially expressed genes(DEGs),followed by clustering and pathway analysis for these DEGs.In this work,we performed a combination analysis on microarrays from three different laboratories using gene expression data on ovarian cancer and obtained a list of differential expression profiles identified as potential candidate in aggressiveness of ovarian cancer.The clustering and pathway analysis explored the different molecular basis of different ovarian cancer stages and potential important regulatory pathways in ovarian cancer development.Our results showed that combination of microarray data from different laboratories in the same platforms may overcome biases derived from probe design and technical features,thereby accelerating the identification of trustworthy DEGs,and demonstrating the advantage of integrative analysis in gene expression studies on epithelial ovarian cancer research.

Expression of Pinl and Ki67 in Cervical Cancer and Their Significance

In order to investigate the expression levels of Pinl mRNA and protein in cervical cancer and its association with Ki67 and their clinical significance, amplification of Pinl gene was examined by RT-PCR, and the expression of both Pinl and Ki67 protein was detected by immunohistochemistry in cervical cancer tissues. It was shown that the expression levels of Pinl were higher in cervical cancer than in normal cervical tissues （P〈0.05）. The expression of Pinl protein was increased progressively along with the disease process from normal cervix to CIN and to cervical cancer （P〈0.05）. No significant difference in the Pinl expression was found between disease stages （FIGO）, pathological grades or pelvic lymph node metastasis status （P〉0. 05）. The expression of Pin1 was significantly higher in adenocarcinoma than in squamous carcinoma of the uterine cervix （P〈0.05）. In cervical cancer, the overexpression of Pinl was positively correlated with that of Ki67 （P〈 0. 05）. These results suggested that the overexpression of Pinl was closely related with cancer cell proliferation or progression of cervical cancer and contributed to oncogenesis. Pinl may serve as a potential marker for cervical cancer diagnosis.

环保型组织样本制备液在不同脂肪细胞检测中的应用价值

背景:当前使用的传统试剂(甲醛溶液固定、乙醇脱水、二甲苯透明脱蜡)对组织前期处理制作的常规石蜡切片质量较低,已成为制约脂肪细胞性肿瘤病理诊断的瓶颈。而且,传统试剂容易对实验室环境造成污染,对实验室人员造成多种健康危害。目的:探讨环保型组织样本制备液在脂肪细胞肿瘤苏木精-伊红染色及非典型性脂肪瘤性肿瘤/高分化脂肪肉瘤MDM2基因检测中的应用价值。方法:选取2016年2月至2022年7月期间592例脂肪细胞肿瘤标本为研究对象,同一标本对半切开,按照所使用的标本前期处理试剂的不同,随机分为2组:传统组和环保组。传统组使用传统试剂甲醛溶液固定、乙醇脱水、二甲苯透明脱蜡制作常规石蜡切片592张;环保组使用环保型组织样本制备液制作切片592张。依据切片苏木精-伊红染色质量的不同等级,比较两组切片的苏木精-伊红染色优良率;进一步对其中病理确诊为非典型性脂肪瘤性肿瘤/高分化脂肪肉瘤的33例标本再次切片后,使用荧光原位杂交法检测MDM2基因,比较两组切片MDM2基因扩增率的差异。结果与结论:(1)相对于传统组,环保组的脂肪细胞肿瘤的组织切片更舒展、更完整,细胞无折叠,染色更清晰,红蓝对比度更佳,细胞结构更致密;(2)环保组的组织切片苏木精-伊红染色优良率高于传统组,差异有显著性意义(P=0.000);(3)相对于传统组,环保组的MDM2探针和CSP12探针仅在标本染色体特定的区域内结合,更具特异性;(4)环保组杂交成功细胞数高于传统组,差异有显著性意义(P=0.000);(5)传统组和环保组的MDM2基因信号数、MDM2/细胞值、CSP12信号数、CSP12/细胞值、MDM2/CSP12值之间的差异无显著性意义(P>0.05);(6)两组MDM2基因扩增率比较,差异无显著性意义(P=0.31);(7)结果表明,环保型组织样本制备液有利于提高脂肪细胞肿瘤的苏木精-伊红染色优良率,保证了非典型性脂肪瘤性肿瘤/高分化脂肪肉瘤MDM2基因的检测质量,有临床推广使用的价值。

Assessment of thiol/disulfide homeostasis for prognostic follow-up of cholangiocarcinoma patients

Background:Cholangiocarcinoma(CCA)is a cancer type that builds in the bile ducts that carry digestive fluid,bile.These ducts connect the liver to the gallbladder and the small intestine.The disease is often diagnosed at an advanced stage,resulting in a low 5-year survival rate.This study aims to evaluate the concentrations of serum thiol/disulfide in CCA patients and healthy volunteers and investigate the association between oxidative activity and clinical and pathological characteristics in CCA patients.By examining the relationship between reduced thiol/disulfide measures and tumorigenesis of the disease,we can potentially identify an unfavourable prognosis in CCA patients.Methods:To assess the status of thiol/disulfide in the blood of Cholangiocarcinoma patients using a novel automated homeostasis assay,we recruited 55 individuals for this study(CCA patients,n=27;healthy volunteers,n=28).We measured the levels of serum total thiol(TT)(–SH+–S-S–),native thiol(NT)(–SH),and disulfide(DD)(–S-S–)in both the CCA group and the control group.Additionally,we calculated the ratio of thiol/disulfide(–SH/–S-S–).Results:We explored the relationship between oxidative activity and clinical and pathological characteristics in CCA patients.The study found no significant differences in age and sex between CCA patients and controls.NT and TT levels were higher in the control group compared to the CCA group.However,the disulfide level did not significantly differ between the two groups.Pearson's correlation matrix analysis revealed positive correlations between TT and NT levels and negative correlations between NT/TT%,DD level,and DD/TT%.Conclusion:Our findings suggest a statistical difference in serum thiol/disulfide parameters.As a marker of total oxidant status,thiol and disulfide levels have decreased in patients with CCA.However,no correlation was found between the tumour markers CEA and CA19-9 and thiol levels.A decrease in blood thiol and disulfide concentrations or the ratio of thiol to disulfide may indicate an increase in oxidative stress and decreased antioxidant capacity.As an indication of the prognosis of cholangiocarcinoma disease in the early period,it may be used to determine the effectiveness of medical interventions.The NT/TT%,disulfide/native thiol percentage,and DD/TT%ratios did not significantly change between the groups.Therefore,measuring serum thiol levels could be a helpful marker for assessing prognosis in the early stages of CCA.Nevertheless,further studies must validate these results and investigate the underlying mechanisms.

Inhibitory Effects of Anti-sense PTTG on Malignant Phenotype of Human Ovarian Carcinoma Cell Line SK-OV-3

To construct eukaryotic expression vector expressing full length anti-sense pituitary tumor transforming gene (PTTG) mRNA and observe its blocking effect on the potential invasion of human ovarian carcinoma cell line SK-OV-3. PCR primers containing designed enzyme cut sites were used for cloning full-length PTTG gene fragment, and the resulting PCR product was inserted into the eukaryotic vector pcDNA3.1 in the antisense direction. The recombinant vector was then transfected into SK-OV-3 by Lipofectamine. The positive cell clone was screened by G418, PTTG and bFGF at protein level expression were detected by Western blot. The biological behavior change of transfection positive cells was observed by colony formation in soft agar assay. Our results showed that SK-OV-3 clones stably expressing full-length recombinant pcDNA3.1-PTTGas were obtained. The expressions of PTTG and bFGF protein in transfected cells were decreased by 61.5 % and 52.3%, respectively as compared with non-transfected ones. The number of colony formation was reduced significantly in transfected cells as compared with empty vector transfected and non-transfected cells. It is concluded that the recombinant vector pcDNA3.1-PTTGas is a novel tool and provides an alternative anti-sense gene therapy targeted at PTTG in human carcinoma.

Construction of Antisense RNA Expression Plasmid for u-PAR and Its Transfection to Highly Invasive PC-3M Cell Subclones

To evaluate the specific inhibition of antisense u PAR on the u PAR expressions in highly invasive cell subclones and to determine its blocking function in the invasion by those cells, a cDNA fragment of u PAR obtained by RT PCR was inserted into a plasmid vector named pcDNA3 in antisense orientation. Then the antisense u PAR recombinant was transfected into highly invasive cell subclones. The u PAR expression in neo resistant cells was examined by RT PCR and immunohistochemical assay. Compared to the control cells, the content of mRNA and protein of u PAR in transfected cells decreased sharply, and the rate of inhibition was 53 % and 73 %, respectively, indicating that an antisense u PAR might have played a specific inhibitory role in its expression in the cells, which may provide a good cell model for making further investigation of the inhibitory effects of the antisense u PAR on invasion in highly invasive cell subclones of human prostate carcinoma.

C-kit:PDGFRα的表达与卵巢浆液性癌顺铂耐药的临床研究(英文)

Objective:To investigate the expression of C-Kit and PDGFRα and their correlation with chemotherapy resis-tance in ovarian serous carcinoma.Methods:We undertook SP immunohistochemical technique to examine the expression of C-Kit and PDGFRα in 59 cases with ovarian serous carcinomas,using archival paraffin-embedded specimens.Then we observed the correlation with chemotherapy resistance.Results:C-Kit and PDGFRα immunostainings were observed posi-tively expressed in 57.63% and 66.10% cases.C-Kit expression was statistically correlated with the progression of disease after first-line chemotherapy(P < 0.05),but PDGFRα was not statistically significant(P > 0.05).There were great difference between of C-Kit and PDGFRα expressions in samples of different differentiated and clinical stages of ovarian serous carci-nomas(P < 0.01 or P < 0.05).Conclusion:C-Kit is statistically correlated with chemotherapy resistance,while PDGFRα is not correlated.

Clinical Grade of Gerneration of Dendritic Cells for Immunotherapy

In order to develop a protocol for clinical grade generation of dendritic cells (DCs) for cancer immumotherapy, aphereses were performed with the continuous flow cell separator and mate- rials were derived from 10 leukemia patients that had achieved complete remission. Peripheral blood monocytes were cultured in vitro with GM-CSF, IL-4 for 6 days, then TNF-α (the TNF-α group) or TNF-α, IL-1β, IL-6, PGE2 (the cytokine mixture group) were added to promote maturation. Cell number was counted by hematology analyzer, and phenotype study (CD1a, CD14, CD83) was carried out by flow cytometry, and the function of DCs was examined by mixed lymphocyte reaction. The results showed that (0.70±0.13)×107/mL (the TNF-α group) and (0.79±0.04)×107/mL (the cytokine mixture group) DCs were generated respectively in peripheral blood obtained by leucapheresis. The phenotypes were as follows: CD1a+ (74.65±4.45)%, CD83+ (39.50±4.16)%, CD14+ (2.90±1.76)% in TNF-α group, and CD1a+ (81.86±5.87)%, CD83+ (81.65±6.36)%, CD14+ (2.46±1.68)% in the cyto- kine mixture group. It was concluded that leucapheresis may be a feasible way to provide large num- ber of peripheral blood monocytes for DC generation, and combined administration of TNF-α, IL-1β, IL-6, and PGE2 may greatly promote maturity.

The Inhibitory Effects of an Antisense u-PAR Vector on Invasion of Highly Invasive Human Prostate Carcinoma PC-3M Cell Subclones

To observe the inhibitory effects of an antisense u-PAR vector on invasion of highly invasive PC-3M cell subclones, the effects of the antisense u-PAR on activity of MMP-9 in those highly invasive cell subclones were detected by a quantitative RT-PCR and zymography. The monolayer invasion assay and colony formation assay in soft agar were used. And tumorigenesis rate and invasions by the cell subclones with or without the antisense u-PAR were observed in nude mice. It was found that in vitro growth of highly invasive PC-3M cell subclones transfected with the antisense u-PAR was declined, and the ability of anchorage-independent growth of those cell subclones was found decreased sharply, with the inhibiting rate becoming 79%and 60% , respectively. Although the anti-sense u-PAR didn't change MMP-9 gene transcription, they could inhibit the activation of MMP-9 of highly invasive PC-3M cell subclones. Moreover, the tumorigenesis rate of the cell subclones with the antisense u-PAR decreased and the growth of a neoplasm also slowed down. The t tests showed the difference between experimental and control groups was statistically significant (P<0.01). The anti-sense u-PAR vector could not only inhibit the invasion ability of highly invasive PC-3M cell subclones in vitro but also restrain the growth of those cell subclones in vivo.

Nanoparticle(NP)-mediated APOC1 silencing to inhibit MAPK/ERK and NF-κB pathway and suppress breast cancer growth and metastasis

Breast cancer is one of the most common malignant tumors with high mortality and poor prognosis in women.There is an urgent need to discover new therapeutic targets for breast cancer metastasis.Herein,we identified that Apolipoprotein C1(APOC1)was up-regulated in primary tumor of breast cancer patient that recurrence and metastasis by immunohistochemistry(IHC).Kaplan-Meier Plotter database showed that high levels of APOC1 in breast cancer patients were strongly associated with worse overall survival(OS)and relapse-free survival(RFS).Mechanistically,APOC1 silencing significantly inhibits MAPK/ERK kinase pathway and restrains the NF-κB to decrease the transcription of target genes related to growth and metastasis in vitro.Based on this regulatory mechanism,we developed these findings into potential therapeutic drugs,glutathione(GSH)responsive nanoparticles(NPs)were used for systemic APOC1 siRNA delivery,NPs(siAPOC1)silenced APOC1 expression,and subsequently resulted in positive anti-tumor effects in orthotopic and liver metastasis models in vivo.Taken together,GSH responsive NPmediated siAPOC1 delivery was proved to be effective in regulating growth and metastasis in multiple tumor models.These findings show that APOC1 could be a potential biomarker to predict the prognosis of breast cancer patients and NP-mediated APOC1 silencing could be new strategies for exploration of new treatments for breast cancer metastasis.

骨巨细胞瘤患者预后影响因素分析

目的:回顾性分析骨巨细胞瘤(giant cell tumor of bone,GCTB)患者的预后复发影响因素。方法:收集2010年至2023年中山大学肿瘤防治中心分子诊断科90例GCTB患者病例资料,分析其临床特点、治疗方式、无进展生存期(progression-free survival,PFS),并对影响预后的因素进行单因素及多因素分析。结果:90例GCTB患者临床资料中,单因素及多因素分析显示:在PFS中,Ki-67的蛋白表达(P=0.013)和手术方式(P=0.013)为PFS预后影响因素(P<0.05);H3F3A、性别、年龄、肿瘤部位、术后是否接受地舒单抗治疗、P53及P63的蛋白表达与预后不相关(P>0.05)。多因素结果分析显示Ki-67的蛋白表达和手术方式是PFS的独立预后因素。结论:在GCTB中,性别、年龄、肿瘤部位、肿瘤直径大小、术后是否接受地舒单抗治疗、H3F3A、P53及P63的蛋白表达等指标均与预后不相关,而Ki-67的蛋白表达及手术方式为预后有价值的参考指标。

CMKLR1 senses chemerin/resolvin E1 to control adipose thermogenesis and modulate metabolic homeostasis

Induction of beige fat for thermogenesis is a potential therapy to improve homeostasis against obesity.β3-adrenoceptor(β3-AR),a type of G protein-coupled receptor(GPCR),is believed to mediate the thermogenesis of brown fat in mice.However,β3-AR has low expression in human adipose tissue,precluding its activation as a standalone clinical modality.This study aimed at identifying a potential GPCR target to induce beige fat.We found that chemerin chemokine-like receptor 1(CMKLR1),one of the novel GPCRs,mediated the development of beige fat via its two ligands,chemerin and resolvin E1(RvE1).The RvE1 levels were decreased in the obese mice,and RvE1 treatment led to a substantial improvement in obese features and augmented beige fat markers.Inversely,despite sharing the same receptor as RvE1,the chemerin levels were increased in obesogenic conditions,and chemerin treatment led to an augmented obese phenotype and a decline of beige fat markers.Moreover,RvE1 and chemerin induced or restrained the development of beige fat,respectively,via the mechanistic target of rapamycin complex 1(mTORC1)signaling pathway.We further showed that RvE1 and chemerin regulated mTORC1 signaling differentially by forming hydrogen bonds with different binding sites of CMKLR1.In conclusion,our study showed that RvE1 and chemerin affected metabolic homeostasis differentially,suggesting that selectively modulating CMKLR1 may be a potential therapeutic target for restoring metabolic homeostasis.

Biomarkers for predicting efficacy of chimeric antigen receptor T cell therapy and their detection methods

Cancer immunotherapy has emerged as the fourth most prevalent approach to tumor treatment,alongside surgery,radiotherapy,and chemotherapy.After several decades of development,chimeric antigen receptor T(CAR-T)cell therapy,a promising branch of adoptive T-cell therapy,has demonstrated superior efficacy and safety in comparison to other cell therapies in the treatment of cancer.At present,CAR-T cells are predominantly used to treat hematological malignancies,although their application in solid tumors is being readily investigated.Although numerous studies have examined the biomarkers associated with the safety of CAR-T cell therapy,few have evaluated predictors of CAR-T cell therapeutic efficacy.Thus,the primary objective of this review article was to provide a comprehensive overview of the factors predicting the efficacy of CAR-T cell therapy,with a particular focus on biomarkers and their detection methods.

Cellular vesicles expressing PD-1-blocking scFv reinvigorate T cell immunity against cancer

Cancer cells aberrantly express immunosuppressive checkpoint ligands and produce certain metabolites that lead to T cell exhaustion.Immune checkpoint blockade(ICB)therapy that reinvigorates exhausted T cells have achieved impressive response in clinical cancer treatment.However,the limited clinical response rate and off-tumor toxicities restrict ICB therapy.Herein,cellular vesicles displaying anti-programmed cell death-1(PD-1)single-chain variable fragment antibody(aPD-1-scFv)were prepared to reinvigorate T cell immunity to counteract cancer.The nanovesicles displaying aPD-1-scFv(aPD-1-scFv NVs)could enhance the anti-tumor activation of T cells through PD-1 blockade.Furthermore,NVs loading the A_(2a)adenosine receptor(A_(2a)R)antagonist CPI-444 assisted T cells to antagonize adenosine,an immunosuppressive metabolite produced by cancer cells.Hence,CPI-444 loaded aPD-1-scFv NVs could intensively increase the density and activity of tumor infiltrating T cells,directly restraining tumor progress and metastasis.

Hepatocyte nuclear factor 1A suppresses innate immune response by inducing degradation of TBK1 to inhibit steatohepatitis

Non-alcoholic steatohepatitis(NASH),a progressive form of non-alcoholic fatty liver disease(NAFLD),is characterised by chronic liver inflammation,which can further prog-ress into complications such as liver cirrhosis and NASH-associated hepatocellular carcinoma(HCC)and therefore has become a growing health problem worldwide.The type I interferon(IFN)signaling pathway plays a pivotal role in chronic inflammation;however,the molecular mechanisms underlying NAFLD/NASH from the perspective of innate immune response has not yet been fully explored.In this study,we elucidated the mechanisms of how innate im-mune response modulates NAFLD/NASH pathogenesis,and demonstrated that hepatocyte nu-clear factor-1alpha(HNF1A)was suppressed and the typeⅠIFN production pathway was activated in liver tissues of patients with NAFLD/NASH.Further experiments suggested that HNF1A negatively regulates the TBK1-IRF3 signaling pathway by promoting autophagic degra-dation of phosphorylated-TBK1,which constrains IFN production,thereby inhibiting the activa-tion of type I IFN signaling.Mechanistically,HNF1A interacts with the phagophore membrane protein LC3 through its LIR-docking sites,and mutations of LIRs(LIR2,LIR3,LIR4,and LIRs)block the HNF1A-LC3 interaction.In addition,HNF1A was identified not only as a novel autop-hagic cargo receptor but also to specifically induce K33-linked ubiquitin chains on TBK1 at Lys670,thereby resulting in autophagic degradation of TBK1.Collectively,our study illustrates the crucial function of the HNF1A-TBK1 signaling axis in NAFLD/NASH pathogenesis via cross-talk between autophagy and innate immunity.

一个常染色体显性遗传型地中海热家系的临床及遗传学分析

目的分析1个由MEFV基因变异所致的常染色体显性遗传型地中海热(autosomal dominant-familial Mediterranean fever,AD-FMF)家系的临床及遗传学特点。方法先证者为男性,5岁,以"发作性无菌性脑膜炎"作为首发表现,症状表现为发作性发热伴头痛呕吐;其母亲、胞兄及胞姐均存在类似发作性发热。提取该家系成员外周血DNA,应用全外显子基因组测序法检测相关基因变异,并通过Sanger测序法验证变异。对可疑变异进行生物信息学分析。结果经全外显子基因组测序分析发现,先证者MEFV基因上第10外显子存在一个c.2229C>G(p.Phe743Leu)错义变异,该变异在其母亲、胞兄及胞姐中均有检出。c.2229C>G虽为国内外未报道的错义变异,但其导致的氨基酸水平的改变(p.Phe743Leu)既往在阿拉伯发病人群中有报道(PS1);该变异在常见正常人群数据库中不存在(PM2),且经Mutationtaster、PROVEAN、PolyPhen-2等变异预测软件预测结果均提示为有害变异;经PubMed BLAST系统分析MEFV基因编码蛋白pyrin第743位氨基酸的改变可影响SPRY_PRY_TRIM20及SPRY_superfamily结构域的完整性;同时经计算机模拟对蛋白3D结构建模分析发现,该氨基酸的改变可导致pyrin蛋白原有空间结构发生改变,原有功能丧失(PP3)。根据美国医学遗传学与基因组学学会序列变异解释指南判定该家系检出变异为可能致病性变异(PS1+PM2+PP3)。结论MEFV基因c.2229C>G(p.Phe743Leu)可能为该家系罹患AD-FMF的致病原因,以"发作性无菌性脑膜炎"作为主要临床表现在AD-FMF患者中较罕见,国内尚未见报道,分析该家系成员的临床表型及基因变异特点有助于提高国内医师对该病的认识。

一例晚期婴儿型异染性脑白质营养不良病患儿的ARSA基因变异分析

目的分析1例晚期婴儿型异染性脑白质营养不良病(metachromatic leukodystrophy,MLD)患儿芳基硫酸酯酶A(arylsulfatase A,ARSA)编码基因的变异情况。方法应用Sanger测序方法检测ARSA基因第1~8共8个外显子序列。用PubMed Protein BLAST分析ARSA的跨种属保守性;应用Ucsf chimera软件对正常结构及变异结构的ARSA进行蛋白质3D结构建模及比对来分析变异所致的蛋白二级结构的丧失及蛋白空间结构的改变;应用PolyPhen-2、Mutation Taster及SIFT软件对新变异进行功能预测。结果患儿携带ARSA基因第3外显子c.467G>A(p.Gly156Asp)和第5外显子c.960G>A(p.Trp320*)复合杂合变异。c.467G>A(p.Gly156Asp)变异经PolyPhen-2、Mutation Taster、SIFT及PROVEAN预测软件预测为可能有害变异,同时经PubMed Protein BLAST分析ARSA第156位Gly在各种属间均高度保守,该氨基酸改变可导致编码的ARSA功能发生障碍;c.960G>A(p.Trp320*)变异经Ucsf chimera软件进行蛋白3D结构建模分析发现,该变异可导致编码蛋白空间结构严重变形,原有功能丧失。结论ARSA基因c.467G>A(p.Gly156Asp)和c.960G>A(p.Trp320*)复合杂合变异可能为该患儿罹患MLD的致病原因,基因变异检测结果可以为家系的遗传咨询和产前诊断提供依据。

一例重型Cornelia de Lange综合征患者的NIPBL基因变异分析

目的分析1例重型Cornelia de Lange综合征(Cornelia de Lange syndrome,CdLS)患儿的NIPBL基因变异,明确其遗传学病因。方法提取患儿及其父母外周血中DNA物质,应用全外显子基因组测序法检测相关基因变异,应用Sanger测序法验证变异。对可疑变异进行生物信息学预测。结果经全外显子基因组测序分析并经Sanger测序验证,发现患儿NIPBL基因第9外显子存在c.1507A>G(p.Lys503Glu)杂合错义变异,该变异为新发变异,且为未报道过的新变异。经PolyPhen-2、Mutation Taster、SIFT预测软件预测c.1507A>G(p.Lys503Glu)变异为可能有害变异,并经HomoloGene系统分析NIPBL蛋白第503位Lys在各种属间均高度保守,该位点氨基酸改变可导致编码的NIPBL原有蛋白功能发生障碍。而经过PubMed BLAST系统进一步分析发现该位点氨基酸的改变可通过影响Neuromodulin_N superfamily结构域的形成来导致NIPBL蛋白功能发生障碍的。结论NIPBL基因c.1507A>G(p.Lys503Glu)错义变异可能为该患儿罹患重型CdLS的致病原因,基因变异检测结果可以为家系的遗传咨询和产前诊断提供依据。