ABSTRACT
Objective:To explore the role of parental origin verification in chromosomal microarray analysis (CMA) on the determination of the clinical significance of copy number variations (CNVs).Methods:This retrospective study collected clinical information from 73 core families who underwent prenatal diagnosis at Peking University First Hospital from November 2017 to December 2019. Indications for prenatal diagnosis included ultrasound abnormality in 54 cases (including 12 with thickened nuchal translucency (≥2.5 mm), four with fetal growth restriction, seven with abnormal pregnancy history, and 31 with isolated ultrasound abnormality), NIPT indicated high-risk in four cases, advanced age in nine cases, abnormal pregnancy history alone in three cases, intrauterine death in two cases and one with maternal mental retardation. Genomic DNA of amniotic fluid sample, chorionic villi, cord blood, fetal tissues, and fetal heart blood were extracted using genomic DNA extraction kit. The CNVs of prenatal samples in 73 subjects were analyzed using array-based comparative genomic hybridization (array-CGH) analysis and single nucleotide polymorphism array (SNP-array). Peripheral blood DNA of the couples, and relevant families if necessary, were collected and analyzed in the same way. The results of parental origin detection in CMA were summarized.Results:A total of 76 CNVs were detected in these 73 samples, out of which nine were pathogenic and parental origin detection revealed that six were de novo, two were maternally, and one was paternally inherited; six CNVs were likely pathogenic, including three de novo, two maternally inherited and one paternally inherited; 20 CNVs were variants of uncertain significance, including five paternally inherited, three maternally inherited and 12 de novo; 41 CNVs were likely benign, among which 38 were inherited from parents with normal phenotype. Conclusions:Parental origin verification plays an important role in explaining the clinical significance of detected fetal CNVs and thereby can help to analyze its clinical effect and reproductive risk.
ABSTRACT
Objective:To summarize the characteristics of genetic variation and prenatal diagnosis in pedigrees with X-linked adrenoleukodystrophy (X-ALD) and elucidate the value of prenatal diagnosis in preventing the birth of children with X-ALD.Methods:Twenty pedigrees, clinically diagnosed with X-ALD in Peking University First Hospital from November 2012 and March 2019, were included in this retrospective study. Genomic DNA was extracted from peripheral blood and amniotic fluid or chorionic villi samples of probands and their families for detecting variants in ATP-binding cassette subfamily D member 1 ( ABCD1) gene using polymerase chain reaction (PCR)-Sanger sequencing. Linkage analysis was also performed on five microsatellite markers near ABCD1 gene to exclude maternal contamination. Characteristics of ABCD1 gene variants and prenatal diagnosis of X-ALD pedigrees were summarized by descriptive statistics. Results:Twenty ABCD1 gene variants were identified in the 20 pedigrees. The variants in three probands that were not detected by next-generation sequencing were identified by PCR-Sanger sequencing. Among the mothers of the 20 probands, 17 carried ABCD1 variants and three did not. We performed 24 prenatal diagnoses on 20 pregnancies (24 fetuses) and identified eight fetuses with variants who were finally terminated. The 16 cases without variants were born alive. The validation results obtained after termination or delivery were consistent with those performed prenatally. Conclusions:No hotspot variants in ABCD1 gene are detected in these X-ALD patients and most variants are maternally inherited. PCR-Sanger sequencing is an effective method for detecting ABCD1 variants. Prenatal diagnosis for mothers who had a body with X-ALD could prevent another one from birth.
ABSTRACT
Objective:To summarize the prenatal diagnostic characteristics of monogenic global developmental delay/intellectual disability(GDD/ID) pedigrees.Methods:This study retrospectively collected the prenatal molecular diagnostic results of 43 pedigrees that were affected with monogenic GDD/ID in the genetic counseling clinic of Peking University First Hospital from January 2015 to June 2019. The results of prenatal molecular tests were validated after birth or pregnancy termination. Pregnancy outcomes and healthy condition of the offspring were followed up. All data were analyzed by descriptive statistical analysis.Results:Among the 43 pedigrees, 24 were affected with autosomal recessive inheritance (AR) GDD/ID, in which six (25%) fetuses were found to carry two pathogenic variants; 13 (55%) had only one pathogenic variant; five (20%) did not harbor any variant. GDD/ID inherited in an autosomal dominant inheritance (AD) pattern was found in 13 pedigrees, in which 11 fetuses carried no variants while the other two fetuses had the same variants as the proband had (in one pedigree, a low-level variant was detected in the peripheral blood sample of the father while absent in peripheral blood samples of parents in the other pedigree, so it was suspected that the variants of these two affected fetuses were inherited from parental mosaicism). In the other six pedigrees with X-linked inheritance (XL) of GDD/ID, one male fetus was found to harbor the pathogenic variant, while no variants were detected in the others. Maternal contamination was excluded in all prenatal samples using short tandem repeat for linkage analysis. Postnatal validations were consistent with the prenatal tests. All nine affected fetuses were terminated, and the other thirty-four children were delivered and in good health.Conclusions:Prenatal molecular diagnostic test is an effective method to detect pathogenic variants during the first and second trimesters for pedigrees affected by monogenic GDD/ID. For pedigrees affected with AD or XL patterns caused by de novo mutations, potential parental mosaicism should be noted and prenatal diagnostic tests are also recommended.
ABSTRACT
Objective@#To analyze the prenatal clinical characteristics and genetic diagnosis of two fetuses with chromosome 17q12 deletion syndrome mainly manifested by renal structural abnormalities.@*Methods@#Clinical data of two pregnant women admitted to Peking University First Hospital in 2017 due to ultrasound indication of fetal kidney structure abnormality in the second trimester were collected. Results of fetal chromosome karyotype analysis and array-based comparative genomic hybridization (aCGH), and aCGH detection of peripheral blood in the two couples were reviewed.@*Results@#(1) In both pregnancies of case 1 and case 2, no abnormal chromosome karyotype was found. In case 2, the fetal fluorescence in situ hybridization (FISH) results showed no abnormality. (2) During the first pregnancy of case 1, there was a 1.351 Mb of single-copy deletion in chromosome 17q12 (34 817 422-36 168 104) and a 1.187 Mb of single-copy duplication in chromosome 3p26.3 (838 934-2 026 269) extracted from umbilical cord blood. Moreover, a 1.299 Mb of single copy duplication in chromosome 3p26.3 (726 645-2 026 269) extracted from maternal peripheral blood was detected. (3) DNA analysis of the umbilical cord blood of case 2 showed a 1.351 Mb of single copy deletion in 17q12. No abnormal copy number variants (CNVs) were detected in the peripheral DNA of the couple.@*Conclusions@#Invasive prenatal detection of CNVs in cases with abnormal fetal kidney ultrasound findings might help to confirm the diagnosis and guide genetic counseling.
ABSTRACT
Objective To analyze the prenatal clinical characteristics and genetic diagnosis of two fetuses with chromosome 17q12 deletion syndrome mainly manifested by renal structural abnormalities. Methods Clinical data of two pregnant women admitted to Peking University First Hospital in 2017 due to ultrasound indication of fetal kidney structure abnormality in the second trimester were collected. Results of fetal chromosome karyotype analysis and array-based comparative genomic hybridization (aCGH), and aCGH detection of peripheral blood in the two couples were reviewed. Results (1) In both pregnancies of case 1 and case 2, no abnormal chromosome karyotype was found. In case 2, the fetal fluorescence in situ hybridization (FISH) results showed no abnormality. (2) During the first pregnancy of case 1, there was a 1.351 Mb of single-copy deletion in chromosome 17q12 (34 817 422-36 168 104) and a 1.187 Mb of single-copy duplication in chromosome 3p26.3 (838 934-2 026 269) extracted from umbilical cord blood. Moreover, a 1.299 Mb of single copy duplication in chromosome 3p26.3 (726 645-2 026 269) extracted from maternal peripheral blood was detected. (3) DNA analysis of the umbilical cord blood of case 2 showed a 1.351 Mb of single copy deletion in 17q12. No abnormal copy number variants (CNVs) were detected in the peripheral DNA of the couple. Conclusions Invasive prenatal detection of CNVs in cases with abnormal fetal kidney ultrasound findings might help to confirm the diagnosis and guide genetic counseling.
ABSTRACT
Objective To evaluate the effect of fluid resuscitation with early external jugular vein access on prognosis of sepsis patients.Methods One hundred and twenty patients with sepsis,admitted to emergency intensive care unit (EICU) and the general intensive care units (ICU) were randomly divided into two groups,external jugular vein group (n=60) and deep-vein group (e.g.internal jugular vein,subclavian vein,femoral vein,n=60).The time elapsed from admission to initial application of norepinephrine,the time required for getting the early goal directed therapy (EGDT) after standard procedure,the length of time needed for subsequent use of vasoactive agents during the entire course of resuscitation serum lactate level at3 h and 6 h after resuscitation,lactate clearance rate,SOFA scores were documented.The mortality rates of 14 days and 28 days were observed after treatment.Results Compared with deep vein access,the time elapsed from admission to the initial application of norepinephrine and the time required for getting EGDT were significantly shortened [(20.78±5.03) vs.(6.12±2.58),P＜0.01;(6.15±2.03)vs.(5.35±2.21),P＜0.05],and the serum level of lactate was significantly decreased[(6.88±1.71)vs.(6.28±1.51),P＜0.05] at 3 h after resuscitation,and lactate clearance rate in percentage was significantly increased at 3 h after resuscitation,and SOFA at 6 h was decreased[(25.8±9.2) vs.(31.2±13.3),P＜0.05],and SOFA at 6 h was distinctly reduced [(5.78±1.19) vs.(5.38±0.96),P＜0.05],and.the mortality rates of 14 days decreased significantly in the external jugular vein group(33.3％ vs.16.7％,P＜0.05).Conclusions Early external jugular vein access can more significantly save time,improve the effect of fluid resuscitation,promote recovery of important organ.It is helpful for improving prognosis in sepsis patients.
ABSTRACT
Objective To investigate the correlation between sTREM-1 and inflammatory factors expression in critical patients and its effect on severity of disease and clinical prognosis .Methods sTREM-1 ,TNF-α,IL-6 ,IL-10 levels were checked in serum of 54 patients who admitted to intensive care unit (ICU) on the first day and only sTREM-1 was checked again on the third day .At the same time ,APACHEⅡand survival situation in 28 days were recorded .Results sTREM-1 level in critical patients was positive correlated with TNF-α,IL-6 and APACHEⅡ(P<0 .01) .There was positive correlation between APACHE Ⅱ and TNF-α,IL-6 ,but pearson correlation coefficient between sTREM-1 and APACHE Ⅱ was higher than TNF-α,IL-6 .Compared with the survivor group ,the concentrations of serum sTREM-1 was significantly higer in non-survivor group on the first day and the third day after entering ICU(P<0 .05) .Conclusion sTREM-1 level is positive correlated with inflammatory reaction and the severity of disease .It also has prognostic value for outcome in patients with critical illness .
ABSTRACT
<p><b>OBJECTIVE</b>To investigate whether the four boys with delayed motor development and intellectual disability suffer from MECP 2 duplication syndrome.</p><p><b>METHOD</b>Blood specimens and clinical data of four patients and mothers of patient 2 and patient 4 were collected. Genomic DNA was extracted from peripheral blood using DNA extraction kit. At first multiplex ligation-dependent probe amplification (MLPA) was employed in 4 patients, two distinct kits SALSA P036 and P070 for sub-telomere screening, and SALSA P245 for the 22 common microdeletion and microduplication syndromes. Then array-CGH analysis was carried out. Two mothers of patients were tested by array- comparative genomic hybridization (CGH) and X chromosome inactivation analysis.</p><p><b>RESULT</b>All the 4 patients presented with severe hypotonia, delayed motor development, intellectual disability and absent or limited language. Three patients manifested recurrent pneumonia in infancy except patient 2. Four patients had duplication on chromosome Xq28 with MLPA kit SALSA P245. Array-CGH identified the size of each duplication on Xq28. The precise size of each duplication was different in the four patients: patient 1, 14.931 Mb, patient 2, 0.393 Mb, patient 3, 0.482 Mb and patient 4, 0.299 Mb. To compare Xq28 duplications with UCSC database (http://genome.ucsc.edu/) revealed that each duplication harbors the MECP 2 and HCFC 1 gene. Mothers of patient 2 and patient 4 also carried microduplication on Xq28. X chromosome inactivation analysis demonstrated completely skewed inactivation (0: 100) and it is the inactive allele that passed on to the patients.</p><p><b>CONCLUSION</b>For patients that present with delayed motor development, intellectual disability, hypotonia, absent or limited language and recurrent infection, combination of MLPA and array- CGH is effective and specific diagnostic methods of MECP 2 duplication syndrome.</p>
Subject(s)
Humans , Male , Chromosomes, Human, X , Genetics , Comparative Genomic Hybridization , Gene Duplication , Mental Retardation, X-Linked , Diagnosis , Genetics , Methyl-CpG-Binding Protein 2 , Genetics , Multiplex Polymerase Chain Reaction , X Chromosome InactivationABSTRACT
Objective To investigate the role of intestinal fatty acid binding protein(I-FABP)in evaluating intestinal dysfunction of septic rats and the effect of glutamine on I-FABP expression.Methods Rats were divided into 3 groups,control group were only fed with Peptisorb,model group were fed with Peptisorb after intraperitoneal injection with E.coli endotoxin lipopolysaccharidegiven and glutamine group were added glutamine on basis of model group.The correlation between serum I-FABP level and intestinal mucosa damage index were analyzed and the concentrations of serum I-FABP in each group were observed and compared. Results The serum level of I-FABP in rats were correlated with the Chiu’s score of intestinal mucosa,mucosa thickness and villus length(P<0.05 ).Compared with control group,the concentration of serum I-FABP in model group and glutamine group were significantly increased(P<0.05),but which in glutamine group was lower than that in model group(P<0.05).Conclusions Serum I-FABP could be an non-invasive diagnosis index for evaluating acute intestinal dysfunction in septic rats.In addition,dietary glutamine supplementation may ameliorate sepsis-induced intestinal epithelial injury in rats.
ABSTRACT
Objective To study three single nucleotide polymorphisms (SNP), SNP-43, -19 and - 63 of calpain-10 (CAPN10) gene in pregnant women with glucose metabolism disorders and their relationship with the risk of these disorders. Methods Totally, 270 pregnant women including 156 with an abnormal oral glucose tolerance test (study group) and 114 healthy controls were selected among those delivered at the Department of Obstetrics and Gynecology, Peking University First Hospital from Jan. 2005 to Dec. 2006. The SNP of CAPN10 gene at posifons 43, 19, and 63 were analyzed by primer-introduced restriction analysis-polymerase chain reaction (PIRA-PCR). Results (1) The frequencies CAPN10 SNP-19 2R/2R genotype (26.9% ,42/156) and 2R allele (48.9%, 152/312 ) were higher than those in controls (12.3% ,14/114 and 36.8% ,84/228, respectively; P=0.012, 0.006). (2) Stratified analysis according to family history of diabetes mellitus showed the proportion of the CAPN10 SNP-19 2R/2R+2R/3R genotypes (83% ,47/57) in the study group were significantly higher than that of control group (52%,11/21 ; P=0.009), and the proportion of SNP-63 T/T + T/C genotypes(47% ,27/57) in study group were significantly higher than that of control group (14%, 3/21 ; P=0.026) among those with positive family history. (3) The combined effect of CAPN10 SNP-43, -19 and -63 on glucose metabolism disorders was analyzed by comparing with the other haplotypes and showed that the haplotype 121 distribution in study group was significantly higher than those in controls(P=0.036), and 221 haplotype was significantly lower than controls (P=0.042). Conclusions (1) CAPN10 SNP-19 is associated with glucose metabolism disorders in pregnant women. And 2R allele might be the risk factor. CAPN10 SNP-19 2R/2R +2R/3R and SNP-63 T/T + T/C genotypes might increase the risk of glucose metabolism disorders in women with positive family history. Haplotype 121 might increase the risk of glucose metabolism disorders in pregnant women and 221 be a protective factor.
ABSTRACT
Objective To investigate the effects of ischemic preconditioning on pneumocyte apoptosis and the expression of HSFT0 after lung isehemia-reperfusion(I/R) in rats and discuss its possible mechanism of extenu-ating ischemia-repedusion injury. Method Thirtysix male Sprague-Dawley rats were randomly divided into three groups [ sham operation(SO ) group, ischemia-teperfusion(L/R) group, and ischemic preconditioning(IP) group],twelve in each group. Lung croas-clamping was used to build the L/R model. In IP group, three cycles of 5-minute-ischemia + 5-minute-reperfusion were given to the pulmonary artery before the procedure. Sham operation rats had a thoracotomy only. Two hours(or five hours) reperfusion was given to both L/R and IP group. Tenninal-deoxynucleotidyl Transferase Mediated d-UTP Nick End Labeiing(TUNEL) was used to evaluate apoptosis. Expression of HSP/0 in lung was observed by immunohistochemical stain and image analysis. Index of quantitative assessment of histologic lung injury(IQA), wet to dry weight ratio(W/D) were measured. The pathological change of lung tissue was observed under both hght and electron microscopy. Statistical analysis was carried out by One-way Anova. Scheffe test was used for intragroup comparison. Results The apoptosis index and expression of HSP70、W/D,IQA of hng tissue in I/R group were higher than those in the sham operation group (P<0.01). Compared with the L/R group, the apoptosis index and expression of HSP70, W/D, IQA of lung tissue significantly decreased (P<0.01), the levels of expression of HSPTO increased significantly in IP group ( P<0.01 ). The pathological and ultrastructure change of lung tissue was better in IP group than those in I/R group. Condusions Ischemic preconditioning can extenuate lung I/R injury by the possible mechanism of increasing the expression of HSPT0 which inhibits the apoptosis during lung I/R injury.
ABSTRACT
BACKGROUND: The dynamic changes of pneumocyte apoptosis and aspartate-specific cysteine proteases-3 (caspase-3) expression in lung tissue of rats during the process of lung ischemia/reperfusion (I/R) injury and the possible action mechanisms remain unclear.OBJECTIVE: This study was to observe the dynamic changes of pneumocyte apoptosis and caspase-3 expression in the rat lung tissue during the process of lung I/R injury, and to analyze the role of pneumocyte apoptosis and the possible action mechanism.DESIGN: A randomized controlled animal experiment.SETTING: Emergency Center, First Hospital, Nanjing Medical University.MATERIALS: This study was carried out in the Animal Laboratory of the First Hospital of Nanjing Medcial University and Nanjing Center for Radioimmunity between April 2006 and September 2006. Twenty-eight male healthy SD rats of clean grade, with body weight of 250 to 350 g, aged 49 to 76 days, were provided by the Experimental Animal Center of Nanjing Medical University. The involved rats were randomized into experimental group and control group, with 14 rats in each.METHODS: ①Experimental intervention: Rats in the experimental group were created into models of lung I/R injury according to the method of Eppinger et al. They were occluded for 45 minutes at the porta of lung (no systolic and diastolic reactions in lung tissue being considered as successful occlusion), and then they were reperfused (recovery of systolic and diastolic function being considered as successful reperfusion); After that, lung tissues were harvested at 3 and 6 hours after lung I/R injury, 7 rats at each time point. Each rat in the control group was subjected to a thoracotony only, but lung tissues were isolated at the same time point by the same method. ②Experimental evaluation: Apoptotic cells in the lung tissue were detected with a flow cytometer by Annexin-V-PI staining, and apoptosis rate was calculated. Caspase-3 expression in the lung tissue was observed by immunohistochemical method and image analysis. Wet to dry weight ratio(W/D) of lung tissue of rats in the two groups was calculated; the number of injured pulmonary alveoli at I/R 3 hours/that at I/R 6 hours was calculated for quantitative evaluation of injured lung tissue; Patho-morphological changes of lung tissue were observed by haematoxylin & eosin staining under an optical microscope.MAIN OUTCOME MEASURES: ①Pneumocyte apoptosis rate and caspase-3 expression in the lung tissue. ②W/D of lung tissue and quantitative evaluation of injured lung tissue. ③Patho-morphological changes of lung tissue.RESULTS: Twenty-eight rats were involved in the final analysis, without deletion. ①Pneumocyte apoptosis rates in the experimental group at I/R 3 and 6 hours were significantly increased as compared with control group (P＜0.01). In the experimental group, pneumocyte apoptosis rate was decreased a little at I/R 6 hours than at I/R 3 hours (P＜0.05). ②Caspase-3 expression in the lung tissue of rats of experimental group reached its top at I/R 3 hours, and was decreased a little at I/R 6 hours. At each time point, caspase-3 expression in the experimental group was increased as compared with control group (P＜0.01). ③In the experimental group, the number of injured pulmonary alveoli at I/R 3 hours/that at I/R 6 hours and W/D ratios of lung tissues were significantly increased as compared with control group (P＜0.01). In the experimental group, two ratios at I/R 6 hours were higher than those at I/R 3 hours (P＜0.05).④In the experimental group, the structure of pulmonary alveoli was destructed, collapsed and disappeared; lots of inflammatory cell infiltration was found; Patho-morphological changes of injured lung tissue at I/R 6 hours were severer than those at I/R 3 hours. No obvious changes were found in the control group.CONCLUSION: At the early stage of lung I/R injury, the alteration of caspase-3 maybe activate pneumocyte apoptosis and induce the apoptosis of lung tissue, and thereby leads to lung injury.
ABSTRACT
Objective:To investigate the effects of methylprednisolone(MP)on pneumocyte apoptosis during lung ischemia/reperfusion injury in rats and to study the possible role of MP in pneumocyte apoptosis.Methods:Forty-two male Sprague-Dawley rats used for unilateral lung ischemia/reperfusion model were randomly divided into three groups:sham operation group(Sh group),ischemia/reperfusion group(I/R group),and methylprednisolone group(MP group).Each group has two subgroups of three hours and six hours.Apoptosis rate in lung tissue was detected by the way of Annexin-V-PI in flow cytometer.Expression of I?B-? in lung was observed by immunohistochemical stain.The index of quantitative assessment of histological lung injury(IQA),the wet to dry weight ratio(W/D),the pathological and ultrastructure changes of lung tissue were measured.Results:Apoptosis rate,W/D,IQA of lung tissue were significantly higher in I/R group than which in Sh group(P