Practice of clinical application of noninvasive prenatal testing based on cell-free fetal DNA / 中华医学遗传学杂志

OBJECTIVE@#To assess the application value of noninvasive prenatal testing (NIPT) based on cell-free fetal DNA.@*METHODS@#The results of 2777 cases of basic and extended NIPT were retrospectively analyzed. The clinical data and outcome of pregnancy were analyzed, in addition with the diagnosis rate and testing efficiency.@*RESULTS@#Among the 2777 pregnant women, 1192 (42.9%) had accepted basic NIPT and 1585 (57.1%) accepted extended NIPT. With a failure rate of 0.1%, 8 and 6 cases were reported respectively as high-risk pregnancies for trisomy 21 and sex chromosomal abnormalities. Other genetic abnormalities were detected in 32 cases. The positive predictive value for trisomy 21 was 85.7%, and one case of 47,XXX was diagnosed among 3 women with high risks for sex chromosomal abnormalities. For those with a high risk for other genetic abnormalities, pregnant diagnosis rates of basic and extended NIPT were 71.4% (5/7) and 68.2% (15/22), respectively. Seven copy number variations (CNVs) were confirmed, including one pathogenic CNV, one likely pathogenic CNV and 5 variants of unknown significance. Among 6 cases with high-risk of maternal CNVs, 5 fetuses and the mothers were confirmed to be carriers. No CNV was detected in the remainder fetus by chromosomal microarray analysis, while its mother was a carrier of the corresponding CNV.@*CONCLUSION@#NIPT has shown a relatively high positive predictive value for the screening of trisomy 21 and maternal CNVs but with a limited efficiency for the discovery of fetal CNVs. For other genetic abnormalities signaled by NIPT, informed choice by the pregnant women during pre-testing consultation is recommended. Invasive prenatal diagnosis should be considered in the combination of NIPT reports and fetal ultrasound, while the residual risks should be fully informed.

Study on deoxygenation of polymerized bovine hemoglobin and its effects on the molecular stability / 中国输血杂志

【Objective】 To identify the key factors that regulate the physical vacuum stirring deoxygenation process of bovine hemoglobin and evaluate the stabilities of polyHb solution with different oxygenation degree. 【Methods】 The effects of several factors including gas liquid specific phase boundary area, pH values and polyHb concentrations on deoxygenation process were investigated through monitoring the oxygen saturation of polyHb solution. Furthermore, the MetHb contents of the studied polyHb solutions were measured at regular intervals during the experiments to evaluate the molecular stability of the polyHb through monitoring the oxidation degrees of the experimental Hb solutions under the mimic pasteurevirus inactivation conditions. 【Results】 The results showed that the more gas liquid specific phase boundary area could accelerate deoxygenation rate, the better deoxygenation results could be yielded. On the other hand, lower pH value could promote the deoxygenation process of the polyHb solution due to the decreasing of oxygen affinity of the polyhemoglobin molecules. Furthermore, results of investigation about the Hb concentration presented that Hb concentration had no remarkable effects on the deoxygenation of polyHb. In addition, the results from the stability experiments presented that higher degree of deoxygenation could decrease the oxidation process and thus increase the heat tolerance of the polyhemoglobin. 【Conclusion】 The deoxygenation of polyhemglonbin could improve the stability of the polyhemoglobin and the deoxygenation process could be regulated by changing the gas liquid specific phase boundary area and oxygen affinity. This study could provide reference and preliminary basis for the industrialization and development of HBOCs in the future.

Genetic analysis and prenatal diagnosis of a pedigree with oculo-facio-cardio-dental syndrome: a case report and literature review / 中华围产医学杂志

Objective:To analyze the clinical phenotypes and prenatal diagnosis of a pedigree with oculo-facio-cardio-dental (OFCD) syndrome.Methods:A pregnant woman at 17 gestational weeks was admitted to the Nanjing Drum Tower Hospital, the Affiliated Hospital of Nanjing University Medical School in 2017 for genetic counseling. Genetic tests as performed for the proband (the pregnant woman), her husband, and the induced fetus of last pregnancy genetic test and the detected variants were analyzed and verified by chromosomal microarray analysis (CMA), multiplex ligation-dependent probe amplification (MLPA) and quantitative real time-polymerase chain reaction (Q-PCR). The detection platform established by MLPA and Q-PCR technology was used to perform prenatal diagnosis of the present pregnancy. Other family members were screened for BCOR gene mutation. Related mutation types were retrieved from ClinVar database with term of " BCOR", and related literature from CNKI and PubMed with terms of "OFCD syndrome", " BCOR gene", and "oculo facio cardiac dental syndrome" to summarize the clinical manifestations, mutation type and pathogenesis of this disease. Results:The proband has congenital cataracts, long face, congenital atrial septal defect, and severe dental malformations, which were consistent with the clinical features of OFCD syndrome. WES suggested that the proband and her induced fetus were suspected of having a large submicroscopic deletion of the exons of BCOR gene, which was confirmed by CMA, MLPA and Q-PCR, with a 105 kb deletion containing BCOR exons 1-15. The amniotic fluid genetic analysis of the present pregnancy showed that the fetus has a normal female karyotype, and did not carry the same BCOR gene copy number abnormality as the proband. The child grew and normally developed without any characteristic manifestations of OFCD syndrome during follow-up. Other families of the proband did not show clinical features of OFCD syndrone, and no BCOR gene copy number abnormality was detected. A total of 35 cases of BCOR gene mutation types related to OFCD syndrome were retrieved from ClinVar database. The data analysis revealed that the differences in clinical manifestations between Lenz microphthalmos syndrome and OFCD syndrome were mainly caused by different mutation types of BCOR gene. Among the 90 retrieved cases of OFCD syndrome obtained through literature, only one case was reported in China. Analysis of these 90 cases showed that the characteristic manifestations of OFCD syndrome, involving the eye, face, heart, teeth, and skeletal system. OFCD syndrome were confirmed in the proband and her induced fetus according to the clinical manifestation and the mutation type of BCOR gene. Conclusions:The clinical manifestations of OFCD syndrome are complicated, caused by various mutation types of BCOR. Systematic molecular genetic technology can be effectively applied for gene and prenatal diagnosis of OFCD syndrome.

Analysis to the failure rate and causes of noninvasive prenatal testing based on high-throughput sequencing / 中华医学遗传学杂志

OBJECTIVE@#To analyze the cause and pregnancy outcome for non-reportable cell-free DNA (cfDNA) results during non-invasive prenatal testing (NIPT).@*METHODS@#cfDNA was extracted from maternal plasma from 5898 singleton pregnancies at 12 to 22 gestational weeks and underwent NIPT with strict quality control standards. For those with sub-standard results, redraw or invasive prenatal procedures were recommended.@*RESULTS@#Among the 5898 cases, 32 have failed for the initial NIPT, including 17 cases with substandard cffDNA%, 10 cases with data fluctuation after twice library constructing and sequencing, and 5 cases with unidentifiable sex chromosome abnormalities. For these 32 cases, 2 directly underwent amniocentesis, and karyotyping analysis showed both were normal. Six of the 30 redrawn cases finally turned out to be nonreportable. The final nonreportable rate was therefore 0.1% (8/5898). Of the redrawn cases, 1 trisomy 21, 1 trisomy 18 and 1 trisomy 13 high risk-cases were identified, which were all confirmed to be false positive. Among the 6 nonreportable cases, 2 women underwent invasive prenatal testing, and 1 was found to have a normal fetal karyotype, while another was found to have an abnormal karyotype of mos45,X[32]/46,XY[18]. The other 4 nonreportable cases who did not accept invasive prenatal testing have all reported normal child development at follow-up.@*CONCLUSION@#The main reason for nonreportable NIPT results was low cffDNA%. The high success rate of the redrawn cases has effectively increased the overall NIPT success rate and reduced the number of the cases necessitating invasive prenatal diagnosis. The initially nonreportable women may consider retesting after careful counseling with informed consent.

Clinical application of noninvasive high-frequency ventilation in neonatal respiratory support / 中华实用儿科临床杂志

Noninvasive high-frequency ventilation (NIHFV) is a new form of noninvasive ventilation which distends positive pressure continuously with oscillations superimposed on spontaneous tidal breathing, through a noninvasive interface.NIHFV, which provides the advantages of nasal continuous positive airway pressure and high frequency ventilation, can rapidly improve oxygenation, effectively remove carbon dioxide, and does not require synchronous techno-logy.It is considered as a new and effective noninvasive ventilation mode.In this paper, the application of NIHFV to neonates were reviewed, including mechanics of gas exchange, research advances and approach to the use of NIHFV.

Experimental studies on the repair and restitution of cartilage by cartilage acellular extracellular matrix and adipose tissue-derived stem cells / 中华耳鼻咽喉头颈外科杂志

Objective@#To investigate the effects of the repair and restitution of ear-shaped cartilage by adipose tissue-derived stem cells(ADSC) and cartilage acellular extracellular matrix.@*Methods@#ADSC were extracted by digesting with collagenase type II from the adipose tissue from 32 patients with adiposity whose fats were drawn, and were cultured and subcultured in vitro. The natural biological scaffolds were prepared by acellular method using porcine ear cartilage, and then the second generation ADSC(5.0×107/ml) were inoculated on the preformed natural bio-scaffold scaffold by culturing in vitro for 3 days to form a cell scaffold complex. 32 New Zealand white rabbits were randomly divided into the experimental groups, the control group A, the control group B and the control group C. All New Zealand white rabbits were modeled by ear cartilage defects. The cell scaffolds composite was implanted into the experimental group of the ear cartilage defects of rabbits, the ADSC were implanted into the control group A, the cartilage acellular extracellular matrix scaffold was implanted into the control group B and the control group C was modeled only by ear cartilage defects. After 16 weeks, the animals were sacrificed and the repair effect was observed by gross appearance and histological examinations including HE, Toluidine blue staining, Safranin O and typeⅡ collagen staining. Its were quantitatively analyzed by positive staining results of type Ⅱ collagen. Ear cartilage tissue elasticity was detected. SPSS 17.0 software was used to analyze the data.@*Results@#The cartilage defects in the experimental group were repaired well by general shape observation and those in the control group was filled in with granulation tissue. There were significant differences between the experimental group and the control group in the wet weight(P<0.05). HE staining showed that cartilage cavities formed in articular cartilage defects, and only the fibrous tissue was filled with the ear cartilage defect in the control groups. In the repair area, Toluidine blue staining, Safranin O and type Ⅱ collagen staining were positive in the experimental group, and negative in the control groups. There was no significant difference between the experimental group and the normal ear cartilage in the ear cartilage elastic constant detection(P>0.05).@*Conclusions@#The mechanics and histology of rabbit ear neonatal cartilage constructed by ADSC combined with cartilage acellular matrix are close to normal ear cartilage. Cartilage acellular matrix material combined with adipose-derived stem cells has good repair and reconstruction ability for ear cartilage defects, which possesses potential clinical application value.

Clinical application of noninvasive high_frequency ventilation in neonatal respiratory support / 中华实用儿科临床杂志

Noninvasive high_frequency ventilation( NIHFV)is a new form of noninvasive ventilation which distends positive pressure continuously with oscillations superimposed on spontaneous tidal breathing,through a nonin_vasive interface. NIHFV,which provides the advantages of nasal continuous positive airway pressure and high frequency ventilation,can rapidly improve oxygenation,effectively remove carbon dioxide,and does not require synchronous techno_logy. It is considered as a new and effective noninvasive ventilation mode. In this paper,the application of NIHFV to neo_nates were reviewed,including mechanics of gas exchange,research advances and approach to the use of NIHFV.

Label-Free Detection of Immunoglobulin G Based On Silicon-on-Insulator Microring Resonator Analysis System Integrated with Microfluidic Channels / 分析化学

The microfluidic channels integrated with microring resonator were designed. The salt coalescence on chip surface caused by liquid volatilization in open environment was avoided and only 30 μL of reaction solution was consumed. These channels significantly reduced the experiment cost. The design, fabrication and characterization of a highly sensitive and label-free silicon-on-insulator (SOI) microring optical resonator integrated with the microfluidic channels were demonstrated. The radius of the microring was 5 μm and the straight waveguide with a width of 450 nm was employed in the microring resonator. The microring resonator device had many advantages such as high sensitivity, label-free and real-time detection. Using different concentrations of ethanol solution with known refractive indices, the refractive index detection sensitivity was 76.09 nm/RIU and the volume refractive index detection limit was 5.25×10Symbolm_4 RIU. We also demonstrated the label-free quantitative specific detections of human immunoglobulin G (IgG) solutions using an antibody-modified microring resonator by measuring the resonance wavelength shift resulting from refraction index changes causing by the immobilization of antibodies and specific recognition between antibodies and antigens, respectively. The results showed that the microring optical resonator could real-time monitor the reaction between biological molecules, the resonator could be used in the quantitative detection and biological sensing.

Prenatal diagnosis and follow-up of a case with Lowe syndrome caused by interstitial deletion of Xq25-26 / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To report on a sporadic case of Lowe syndrome diagnosed prenatally with ultrasound examination and genetic testing.</p><p><b>METHODS</b>Detailed sonographic fetal screening was performed by an experienced sonographer at 32 weeks of gestation. Fetal cranial magnetic resonance imaging (MRI) was applied to detect potential brain abnormality. Chromosomal microarray analysis (CMA) was conducted on amniotic fluid sample from the fetus and peripheral blood sample from the mother.</p><p><b>RESULTS</b>Congenital cataract and enlarged posterior fossa were detected by fetal ultrasound screening. Fetal cranial MRI found hypoplasia of the gyrus. CMA revealed that the fetus has carried a 633 kb deletion at Xq25-26.1 which encompassed the OCRL gene. The mother was a carrier of the same deletion. Clinical examination after birth confirmed that the neonate was affected with Lowe syndrome in addition with an atrial septal defect.</p><p><b>CONCLUSION</b>Prenatal diagnosis of Lowe syndrome without a family history largely depends on fetal imaging. Should cataract be found by ultrasound screening, fetal MRI may be considered to rule out central nervous system anomalies. CMA assay should also be considered to facilitate the diagnosis.</p>

Detection of a recurrent de novo mutation in a Chinese family affected with Duchenne muscular dystrophy / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To provide genetic analysis for a family affected with Duchenne muscular dystrophy (DMD) with a recurrent de novo mutation.</p><p><b>METHODS</b>Multiplex ligation dependent probe amplification (MLPA) was used to detect potential deletion and duplication of the DMD gene, and the DNA products were sequenced on a Genetic Analyzer 3130 sequencer. Haplotype analysis was performed using four short tandem repeat polymorphism loci (44C/A, 45C/A, 49C/A and 63C/A) of the DMD gene for the family.</p><p><b>RESULTS</b>A same deletional mutation (Del 48-50) of the DMD gene was detected in the proband and fetus, but not in their mother. The proband and fetus have inherited the same haplotype of the DMD gene from their mother. The fetus was predicted to be affected by the disease.</p><p><b>CONCLUSION</b>Above findings suggested that the mother was very likely to have a germline mosaicism for the DMD gene mutation. For the de novo DMD mutation, although genetic analysis of peripheral blood DNA has indicated that the proband's mother was not a carrier, germline mosaicism could still not be ruled out, and prenatal gene diagnosis should be provided for subsequent pregnancies.</p>

Application of different technologies for distinguishing true and pseudo mosaicisms during prenatal diagnosis / 中华医学遗传学杂志

<p><b>OBJECTIVE</b>To use different technologies to distinguish true and pseudo mosaicisms among cultured amniocytes in order to attain more accurate diagnosis.</p><p><b>METHODS</b>With informed consent, 20 mL of amniotic fluid was obtained from pregnant women at between 18 to 24 gestational week. Each amniotic fluid sample was processed as two separate lines for the culturing, observation, harvesting and analysis. All procedures were conducted conforming to the Technology Standards of Cytogenetic Prenatal Diagnosis of Fetal Chromosome Abnormalities issued by the Ministry of Health in 2010. Umbilical cord blood, fluorescence in situ hybridization (FISH), single nucleotide polymorphism array (SNP-array) and flow cytometer were applied when necessary.</p><p><b>RESULTS</b>Among 3910 cases, 128(3.3%) were detected as mosaicisms. Further analysis with the above technologies has verified 6 cases as true mosaicisms and the remaining 120 as pseudomosaicisms. For one case detected by karyotype analysis as 47, XXY/46, XY, the ratio of different cell lines was confirmed by FISH as 1:2. Another case, detected by karyotype analysis as 47, XX,+mar/46, XX (1:1), was verified by SNP-array as 18p duplication. A suspected polyploidy mosaicism was rejected by flow cytometry and cord blood karyotyping.</p><p><b>CONCLUSION</b>Two separate cell cultures are important for distinguishing true and pseudo mosaicisms. Combined FISH, SNP-array and flow cytometry can attain more reliable and accurate diagnosis for mosaicisms.</p>

Stereotaxic transplantation of bone marrow mesenchymal stem cells in treatment of spinal cord injury in rats / 中国组织工程研究

BACKGROUND:The key of stem cells for treating nervous tissue injury is the transplantation of stem coils that have regeneration capacity.The structure and function of central nervous system were re-established by multiple action mechanisms.OBJECTIVE:To explore the effects and mechanisms of bone marrow mesenchymal stem cells(BMSCs)locally transplanted into rats with spinal cord injury on neurological recovery.METHODS:BMSCs were separated with density gradient centrifugation and cell attachment.10 mg/L BrdU was used for labeling before cell transplantation.Adult female Wistar rats were used to establish spinal cord injury models using an aneurysm clip,and they were then randomly divided into control group,saline group and transplantation group.In the transplantation group,BMSCs were transplanted into the damaged spinal cord by stereotaxis at day 7 following damage.In the saline group,an equal volume of saline was utilized.In the control group,the rats were left intact.Basso,Beattie and Bresnahan(BBB)locomotor rating scale was used before and at 7,14,30,60 and 90 days following damage.Rats were sacrificed at day 90.BrdU-positive cells,Brdu+neuron specific enolase,Brdu+glial fibrillary acidic protein(GFAP),Brdu+basic fibroblast growth factor(bFGF),and Brdu+brain-derived nerve growth factor(BDNF)immunohistochemistry double-staining cells and simple staining positive calls were observed.RESULTS AND CONCLUSION:The recovery of BBB function score was better in the transplantation group than in the control group(P＜0.05).The recovery speed of BBB function score was slower in the saline group than in the control group at 30 days following damage(P＜0.05).No significant difference was determined at day 90 compared with the control group(P＞0.05).BrdU-positive cells and double-staining cells of immunohistochemistry could be found at the center of damage site and 1 cm from caudal end to damaged site in rats of the transplantation group.The number of NSE,GFAP,bFGF and BDNF simple staining cells was significantly greater in the transplantation group than in the control and saline groups(P＜0.05).Results indicated that BMSC transplantation can improve the recovery of nervous function of rats with spinal cord injury.Its mechanism may be correlated with the differentiation of transplanted cells into neuron-like and glial cell-like cells,secretion or promoting secretion of neurotrophic factors in host.