[Research Progress on Localization of Pulmonary Nodules].

With the widespread adoption of low-dose computed tomography (LDCT) and advancements in computed tomography image resolution, the detection rate of pulmonary nodules, especially smaller ones, has significantly improved. The risk of developing malignant tumors increases with the pulmonary nodule diameter. Video-assisted thoracoscopic surgery (VATS) stands out as the preferred surgical method. The accurate localization of pulmonary nodules is crucial for the success of VATS and remains a significant challenge for thoracic surgeons. Currently, commonly employed localization methods include CT-guided percutaneous positioning, bronchoscope-guided positioning, intraoperative ultrasound positioning, augmented reality (AR), and 3D print-assisted positioning. This review explores recent research progress, highlights the strengths and weaknesses of various pulmonary nodule localization methods. The aim is to provide valuable insights for clinical applications and guide future developments in this field.

Mutations in ARHGEF15 cause autosomal dominant hereditary cerebral small vessel disease and osteoporotic fracture.

Cerebral small vessel disease (CSVD) is a prominent cause of ischemic and hemorrhagic stroke and a leading cause of vascular dementia, affecting small penetrating vessels of the brain. Despite current advances in genetic susceptibility studies, challenges remain in defining the causative genes and the underlying pathophysiological mechanisms. Here, we reported that the ARHGEF15 gene was a causal gene linked to autosomal dominant inherited CSVD. We identified one heterozygous nonsynonymous mutation of the ARHGEF15 gene that cosegregated completely in two families with CSVD, and a heterozygous nonsynonymous mutation and a stop-gain mutation in two individuals with sporadic CSVD, respectively. Intriguingly, clinical imaging and pathological findings displayed severe osteoporosis and even osteoporotic fractures in all the ARHGEF15 mutation carriers. In vitro experiments indicated that ARHGEF15 mutations resulted in RhoA/ROCK2 inactivation-induced F-actin cytoskeleton disorganization in vascular smooth muscle cells and endothelial cells and osteoblast dysfunction by inhibiting the Wnt/ß-catenin signaling pathway in osteoblast cells. Furthermore, Arhgef15-e(V368M)1 transgenic mice developed CSVD-like pathological and behavioral phenotypes, accompanied by severe osteoporosis. Taken together, our findings provide strong evidence that loss-of-function mutations of the ARHGEF15 gene cause CSVD accompanied by osteoporotic fracture.

Co-treatment of betulin and gefitinib is effective against EGFR wild-type/KRAS-mutant non-small cell lung cancer by inducing ferroptosis.

Clinical trials suggest that non-small-cell lung cancer (NSCLC) patients with KRAS mutations and wild-type EGFR have reduced benefits from gefitinib treatment. Ferroptosis is a new form of cell death that plays an important role in mediating the sensitivity of EGFR-TIKs. Here, we explored the antitumor ability of gefitinib in combination with betulin to overcome drug resistance through ferroptosis in wild-type EGFR/KRAS-mutant NSCLC cells. A549 and H460 cells were treated with gefitinib and betulin, and cell viability, apoptosis, and migration ability were assessed using the CCK-8 assay, flow cytometry, and wound-healing assay, respectively. Several cell death inhibitors were used to study the form of cell death. Ferroptosis-related events were detected by performing reactive oxygen species (ROS) and iron level detection, malondialdehyde (MDA) assay, and glutathione (GSH) assay. EMT-associated proteins and ferroptosis-related proteins were detected by using western blotting. A xenograft model was constructed in vivo to investigate the role of the combination treatment of betulin and gefitinib in NSCLC tumor growth. Gefitinib in combination with betulin exhibited antagonistic effects on cellular viability and induced cell apoptosis. It also induced ROS accumulation, lipid peroxidation, and GSH depletion and induced ferroptosis-related gene expression. Moreover, ferroptosis inhibitors, but not inhibitors of other forms of cell death, abrogated the effect of gefitinib in combination with betulin. Moreover, it also inhibited the tumor growth of NSCLC in vivo. Our findings suggest that gefitinib in combination with betulin is a novel therapeutic approach to overcome gefitinib resistance in EGFR wild-type/KRAS-mutant NSCLC cells by inducing ferroptosis.

Long-read sequencing identified intronic repeat expansions in SAMD12 from Chinese pedigrees affected with familial cortical myoclonic tremor with epilepsy.

BACKGROUND: The locus for familial cortical myoclonic tremor with epilepsy (FCMTE) has long been mapped to 8q24 in linkage studies, but the causative mutations remain unclear. Recently, expansions of intronic TTTCA and TTTTA repeat motifs within SAMD12 were found to be involved in the pathogenesis of FCMTE in Japanese pedigrees. We aim to identify the causative mutations of FCMTE in Chinese pedigrees. METHODS: We performed genetic linkage analysis by microsatellite markers in a five-generation Chinese pedigree with 55 members. We also used array-comparative genomic hybridisation (CGH) and next-generation sequencing (NGS) technologies (whole-exome sequencing, capture region deep sequencing and whole-genome sequencing) to identify the causative mutations in the disease locus. Recently, we used low-coverage (~10×) long-read genome sequencing (LRS) on the PacBio Sequel and Oxford Nanopore platforms to identify the causative mutations, and used repeat-primed PCR for validation of the repeat expansions. RESULTS: Linkage analysis mapped the disease locus to 8q23.3-24.23. Array-CGH and NGS failed to identify causative mutations in this locus. LRS identified the intronic TTTCA and TTTTA repeat expansions in SAMD12 as the causative mutations, thus corroborating the recently published results in Japanese pedigrees. CONCLUSIONS: We identified the pentanucleotide repeat expansion in SAMD12 as the causative mutation in Chinese FCMTE pedigrees. Our study also suggested that LRS is an effective tool for molecular diagnosis of genetic disorders, especially for neurological diseases that cannot be positively diagnosed by conventional clinical microarray and NGS technologies.

Retrovirus-Mediated Transfection of the Tissue-type Plasminogen Activator Gene Results in Increased Thrombolysis of Blood Clots.

Tissue-type plasminogen activator (tPA) is involved in the lysis of blood clots. In this study, we attempted to target thrombolysis and enhance blood clot lysis by generating a construct (pLEGFP-N1-tPA) to integrate tPA gene into the genome of different cell lines. pLEGFP-N1-tPA construct was generated and used to target the tPA gene in different cell lines. The thrombolytic effects mediated by the supernatant from transfected HeLa cells and Linx cells were assessed using plasma thrombus plates. Furthermore, enhanced green fluorescent protein (EGFP), which was fused to the tPA gene in the pLEGFP-N1-tPA construct, was analyzed under the fluorescent microscope to assess tPA localization. We also monitored tPA activity and expression in the transfected cell lines. As part of the study, we successfully generated the pLEGFP-N1-tPA construct. The sequence of this construct was verified and the construct was subsequently used to generate the PT67/pLEGFP-N1-tPA cell line. The pLEGFP-N1-tPA construct was also used to transfect HeLa cells and Linx cells. We observed that supernatants from transfected cells were capable of lysing thrombi. In addition, tPA activity and tPA concentration were elevated in the latter supernatants and tPA was rapidly and stably expressed in the transfected cell lines. These results reveal a potentially important thrombolytic role for tPA-targeted gene therapy following cardiac valve replacement.

Pretreatment of glial cell-derived neurotrophic factor and geranylgeranylacetone ameliorates brain injury in Parkinson's disease by its anti-apoptotic and anti-oxidative property.

The aim of the present study was to determine the combined effects of glial cell-derived neurotrophic factor (GDNF) and geranylgeranylacetone (GGA) on neuron apoptosis and oxidative stress in Parkinson's disease (PD). A mouse MPTP model of PD and cellular models of H2 O2 and MPP+ -treated PC12 cells were established. Swimming, pole, and traction tests were used to detect behavioral impairment. Tyrosine hydroxylase (TH) immunohistochemistry was used to evaluate the neuron loss. TUNEL and flow cytometer method were used to identify the neuron apoptosis. MDA levels and activities of antioxidant enzymes were used to detect the oxidative damage. The PD model of mice received GDNF and GGA exhibited a significant recovery in their swim, pole, and traction scores. Moreover, the combined treatment significantly reduced the neuron apoptosis in the substantia nigra (SN) of PD mice or in H2 O2 or MPP+ -induced PC12 cells compared with the single drug group. In addition, significant reduction of MDA levels and improvement of activities of CAT, SOD, and GSH-px were observed after GDNF and GGA treatment in the PD model and H2 O2 or MPP+ -induced PC12 cells. The combination of GDNF and GGA ameliorated neural apoptosis and oxidative damage in PD.

Ultrasound-triggered effects of the microbubbles coupled to GDNF- and Nurr1-loaded PEGylated liposomes in a rat model of Parkinson's disease.

The purpose of this study was to investigate ultrasound-triggered effects of the glial cell line-derived neurotrophic factor (GDNF) + nuclear receptor-related factor 1 (Nurr1)-polyethylene glycol (PEG)ylated liposomes-coupled microbubbles (PLs-GDNF + Nurr1-MBs) on behavioral impairment and neuron loss in a rat model of Parkinson's disease (PD). The unloaded PEGylated liposomes-coupled microbubbles (PLs-MBs) were characterized for zeta potential, particle size, and concentration. 6-hydroxydopamine (6-OHDA) was used to establish the PD rat model. Rotational, climbing pole, and suspension tests were used to detect behavioral impairment. The immunohistochemical staining of tyrosine hydroxylase (TH) and dopamine transporter (DAT) was used to assess the neuron loss. Western blot and quantitative real-time PCR (qRT-PCR) analysis were used to measure the expression levels of GDNF and Nurr1. The particle size of PLs-MBs was gradually increased, while the concentration and absolute zeta potential were gradually decreased as the time prolongs. 6-OHDA increased amphetamine-induced rotations and loss of dopaminergic neurons as compared to sham group. Interestingly, PLs-GDNF-MBs or PLs-Nurr1-MBs decreased rotations and increased the TH and DAT immunoreactivity. Combined of both genes resulted in a robust reduction in the rotations and a greater increase of the dopaminergic neurons. The delivery of PLs-GDNF + Nurr1-MBs into the brains using magnetic resonance imaging (MRI)-guided focused ultrasound may be more efficacious for the treatment of PD than the single treatment.

[Genetic diagnosis of a child with aortic stenosis and thumb aplasia].

OBJECTIVE: To analyze the molecular mechanism and prognosis of a child with aortic stenosis and thumb aplasia. METHODS: The karotypes of the child and his parents were analyzed with routine G-banding. Their genomic DNA was also analyzed with array comparative genomic hybridization(aCGH) for chromosomal duplications/deletions. RESULTS: No karyotypic abnormality was detected at cytogenetic level for the child and his parents. aCGH identified a de novo 5.86 Mb deletion at 2q22.3-q23.3 in the child. CONCLUSION: The child was diagnosed with 2q23.1 microdeletion syndrome. MBD5 may be the key gene for the 2q23.1 microdeletion syndrome.

Intranasal Administration of GDNF Protects Against Neural Apoptosis in a Rat Model of Parkinson's Disease Through PI3K/Akt/GSK3ß Pathway.

Glial cell line-derived neurotrophic factor (GDNF) plays important roles in protecting the damaged or dying dopamine neurons in the animal models of Parkinson's disease (PD). This study was to determine the effect and mechanisms of GDNF on the apoptosis of neurons in 6-hydroxydopamine (6-OHDA) induced Parkinson's disease model of rats. Healthy male Sprague-Dawley rats (220-240 g) were randomly divided into six groups (n = 10). 6-OHDA was used to establish the PD rat model. Tyrosine hydroxylase (TH) immunohistochemistry was used to assess the neuron loss in 6-OHDA-lesioned rats. TUNEL and western blot were used to identify the effects and mechanisms of GDNF in the rat model of PD. The numbers of TH-positive neurons in the 6-OHDA-injected lesioned substantia nigra (SN) decreased significantly compared with the Sham group. GDNF treatment effectively ameliorated the apoptosis of neuronal cells in SN induced by 6-OHDA. In addition, GDNF significantly increased serine protein kinase B (Akt) and glycogen synthase kinase 3 beta (GSK3ß) phosphorylation induced by 6-OHDA. In contrast, application of LY294002 or triciribine reversed the roles of GDNF in PD models. The results implicated that the anti-apoptosis effects of GDNF in neurons might be mediated through PI3K/Akt/GSK3ß pathway. Therefore, GDNF may be a promising agent for PD treatment.

[Genetic and prenatal diagnosis for a haemophilia A family with two novel mutations of F8 gene].

OBJECTIVE: To conduct genetic diagnosis for a family affected with hamophilia A. METHODS: Potential mutations of the F8 gene were analyzed with PCR and Sanger sequencing. Carriers of the mutation were identified through linkage analysis using short tandem repeat (STR) markers. Suspected mutations were verified among 100 healthy controls to rule out genetic polymorphism. Prenatal diagnosis was provided based on the above results. RESULTS: Sequencing analysis has identified two mutations, c.1 A>T and c.4 C>T, which have replaced the start codon (ATG) with leucine (TTG) and glutamine (GAA) with the stop codon (TAA), respectively. The same mutations were not found among the 100 healthy controls. The patient's mother and sister were heterozygous for the same mutations. Upon prenatal diagnosis, the fetus was determined as a male and did not harbor the above mutations. Linkage analysis also confirmed that the fetus has inherited the non-risk X chromosome from his maternal grandfather. CONCLUSION: Detection of pathogenic mutations can enable prenatal diagnosis for the disease.

Numb Protects Human Renal Tubular Epithelial Cells From Bovine Serum Albumin-Induced Apoptosis Through Antagonizing CHOP/PERK Pathway.

In recent studies, we found that Numb is involved in oxidative stress-induced apoptosis of renal proximal tubular cells; however, its function on ER stress-induced apoptosis in proteinuric kidney disease remains unknown. The objective of the present study is to explore the role of Numb in urinary albumin-induced apoptosis of human renal tubular epithelial cells (HKCs). In this study, we demonstrate that incubation of HKCs with bovine serum albumin (BSA) resulted in caspase three-dependent cell death. Numb expression was down-regulated by BSA in a time- and dose-dependent manner. Knockdown of Numb by siRNA sensitized HKCs to BSA-induced apoptosis, whereas overexpression of Numb protected HKCs from BSA-induced apoptosis. Moreover, BSA activated CHOP/PERK signaling pathway in a time- and dose-dependent manner as indicated by increased expression of CHOP, PERK, and P-PERK. Furthermore, knockdown of CHOP or PERK significantly attenuated the promoting effect of Numb on BSA-induced apoptosis, while overexpression of CHOP impaired the protective effect of Numb on BSA-induced apoptosis. Taken together, our findings demonstrate that Numb plays a protective role on BSA-induced apoptosis through inhibiting CHOP/PERK signaling pathway in human renal tubular epithelial cells. Therefore, the results from this study provides evidence that Numb is a new target of ER-associated apoptotic signaling networks and Numb may serve as a promising therapeutic target for proteinuric diseases.

[Genetic diagnosis for a family without exonic deletions and duplications of dystrophin gene].

OBJECTIVE: To conduct genetic diagnosis for a family in which no exonic deletions and duplications of the dystrophin gene were detected. METHODS: Potential exonic deletions and duplications of the dystrophin gene were initially analyzed with using multiplex ligation-dependent probe amplification (MLPA). Subsequently, all of the 79 exons of the dystrophin gene of the proband and a pregnant woman from the family were analyzed with PCR amplification and DNA sequencing. Following identification of the causative mutation, prenatal diagnosis was provided. RESULTS: MLPA analysis had detected no exonic deletions and duplications of the dystrophin gene. Sequence analysis has identified a C>T mutation on the 22nd nucleotide position of the 70th exon of the dystrophin gene (c.10108 C>T), which has replaced the codon CGA to a stop codon (TGA). The patient's mother and sister were both heterozygous for the same mutation. Upon prenatal diagnosis, the fetus was found to be positive for the Y chromosome sex-determining gene (SRY) and has carried above mutation. The result of short tandem repeat linkage analysis also confirmed that the fetus has inherited the mutant X chromosome. CONCLUSION: The causative mutation of the dystrophin gene has been discovered in an affected family, which has enabled prenatal diagnosis of the disease.

[Association of polymorphisms of HLA-DRB1 gene with unexplained recurrent spontaneous abortion in ethnic Hans from Henan].

OBJECTIVE: To assess the association of polymorphisms of human leukocyte antigen DRB1 gene (HLA-DRB1) with susceptibility to unexplained recurrent spontaneous abortion (URSA). METHODS: The HLA-DRB1 gene was typed with polymerase chain reaction-specific sequence primers (PCR-SSP) method in 200 couples with URSA and 200 couples with a normal pregnancy history. RESULTS: The frequencies of DRB1*09 and DRB1*13 alleles were significantly greater in the URSA group compared with the control group (14.50% vs. 9.50%, and 7.00% vs. 4.38%, both P<0.05), whilst the frequencies of DRB1*04 and DRB1*12 alleles were significantly lower (7.13% vs. 10.75%, and 8.63% vs. 14.38%, both P<0.05). For females from the URSA group, the frequency of DRB1*09 allele (14.00%) was significantly higher compared with the controls (9.25%) (P=0.036), whilst the frequency of DRB1*12(8.50%) allele was significantly lower (14.00%) (P=0.014). For males in the URSA group, the frequencies of DRB1*09 and DRB1*13 alleles were significantly higher than those of the controls (15.00% vs. 9.75%, and 9.25% vs. 4.00%, both P<0.05), whilst the frequencies of DRB1*04 and DRB1*12 alleles were significantly lower (5.75% vs. 12.25%, and 8.75% vs. 14.75%, P<0.05). CONCLUSION: The DRB1*09 and DRB1*13 alleles may contribute to the susceptibility of URSA, while DRB1*04 and DRB1*12 alleles may confer a protective effect factors. For females, however, no significant association of DRB1*13 and DRB1*04 alleles with URSA was found.

[Predictive value of HLA-DRB1 gene for the treatment of unexplained recurrent spontaneous abortion with paternal lymphocyte alloimmunization therapy in Henan Hans].

OBJECTIVE: To explore the value of HLA-DRB1 gene in predicting the outcome of unexplained recurrent spontaneous abortion (URSA) treated with paternal lymphocyte alloimmunization therapy (PLAT) in Henan Hans. METHODS: Three hundred URSA patients were recruited. Following PLAT treatment, they were divided into two groups according to the outcome of pregnancy. Polymerase chain reaction sequence specific primer (PCR-SSP) were conducted to analyze the HLA-DRB1 gene. RESULTS: For those who have received PLAT treatment, the frequency of HLA-DRB1*11 was significantly lower in successfully treated cases than those with abortion (0.052 vs. 0.110, P < 0.05, OR=0448), whilst the frequency of HLA-DRB1*15 was significantly greater in the former (0.207 vs. 0.100, P < 0.05, OR=2.352). CONCLUSION: For patients who have received PLAT treatment, those with HLA-DRB1*15 are more likely to conceive that those with HLA-DRB1*11.

[Notch signaling pathway in 1-methyl-4-phenylpyridinium-induced apoptosis of SH-SY5Y cell].

OBJECTIVE: To explore the role of Notch signaling pathway and the effect of γ-secretase inhibitor (DAPT) on the apoptosis induced by 1-methyl-4-phenylpyridinium (MPP(+)) in differentiated SH-SY5Y cell. METHODS: SH-SY5Y cell were incubated with various concentrations (0, 0.5, 1, 1.5, 2 mmol/L) of MPP(+) for 0, 24, 48 and 72 h respectively. Flow cytometry with Annexin V-FITC/PI double staining was used for apoptotic analysis. The protein expressions of Notch-1, Jagged-1 and Hes-1 were detected by Western blot. SH-SY5Y cell were preincubated with 10 µmol/L DAPT for 15 min before 1.5 mmol/L MPP(+) treatment for 48 h. Flow cytometry and Western blot were performed to analyze the cell apoptosis and protein expressions of Notch-1, Jagged-1 and Hes-1. RESULTS: MPP(+) induced the apoptosis of SH-SY5Y cell in a dose (3.20% ± 0.19% vs 10.00% ± 1.72%, 20.60% ± 3.76%, 32.80% ± 5.12%, 46.00% ± 5.06%, all P < 0.05) and time- (2.80% ± 0.21% vs 12.30% ± 1.82%, 19.60% ± 2.89%, 35.00% ± 4.78%, all P < 0.05) dependent manner. MPP(+) up-regulated the expressions of Notch-1, Jagged-1 and Hes-1 in a dose- and time-dependent manner in SH-SY5Y cell. DAPT treatment decreased MPP(+)-induced apoptosis (3.10% ± 0.21% vs 35.50% ± 4.98%, 19.20% ± 2.98%, both P < 0.05) and the expressions of Notch-1, Jagged-1 and Hes-1 in SH-SY5Y cell. CONCLUSION: The activation of Notch signaling pathway plays an important role in MPP(+)-induced apoptosis of SH-SY5Y cell. DAPT inhibits Notch signaling pathway and protects SH-SY5Y cell from MPP(+)-induced apoptosis.

Enlarged perivascular spaces in alcohol-related brain damage induced by dyslipidemia.

Perivascular spaces (PVSs) as the anatomical basis of the glymphatic system, are increasingly recognized as potential imaging biomarkers of neurological conditions. However, it is not clear whether enlarged PVSs are associated with alcohol-related brain damage (ARBD). We aimed to investigate the effect of long-term alcohol exposure on dyslipidemia and the glymphatic system in ARBD. We found that patients with ARBD exhibited significantly enlargement of PVSs in the frontal cortex and basal ganglia, as well as a notable increased levels of total cholesterol (TC) and triglycerides (TG). The anatomical changes of the glymphatic drainage system mentioned above were positively associated with TC and TG. To further explore whether enlarged PVSs affects the function of the glymphatic system in ARBD, we constructed long alcohol exposure and high fat diet mice models. The mouse model of long alcohol exposure exhibited increased levels of TC and TG, enlarged PVSs, the loss of aquaporin-4 polarity caused by reactive astrocytes and impaired glymphatic drainage function which ultimately caused cognitive deficits, in a similar way as high fat diet leading to impairment in glymphatic drainage. Our study highlights the contribution of dyslipidemia due to long-term alcohol abuse in the impairment of the glymphatic drainage system.

Numb/Notch signaling plays an important role in cerebral ischemia-induced apoptosis.

Numb has been shown to play diverse roles in the central nervous system of adult mammals, and accumulating evidence indicates a role for Numb in apoptosis. In this study, we characterize the role of Numb in ischemia-induced apoptosis, and investigate the underlying pathway involved in this process. In vivo, exposure of pheochromocytoma (PC12) cells to glucose deprivation (GD) resulted in caspase-3-dependent apoptosis. Numb expression was upregulated by GD in a time-dependent manner, while Notch expression was down regulated. Knocking down endogenous Numb expression via siRNA protected PC12 cells from GD-induced apoptosis, whereas Numb overexpression sensitized PC12 cells to GD-induced apoptosis. In vivo, significantly increased Numb expression levels, together with activation of apoptosis, can be observed in the ischemic penumbra following cerebral ischemia. Taken together, our data show that Numb promotes ischemia-induced apoptosis. Based on these results, we conclude that inhibition of Numb could be a novel therapeutic approach for inhibiting apoptosis in the ischemic penumbra.

[Genetic analysis and prenatal diagnosis for non-syndromic hearing impairment].

OBJECTIVE: To detect genetic mutations underlying non-syndromic hearing impairment (NSHI) and establish a method for prenatal diagnosis. METHODS: Sixty six NSHI patients were included in this study. DNA was extracted from peripheral blood. Genetic mutations were detected by gene chip analysis and direct sequencing of GJB2 gene. For 7 pregnant women at high risk, prenatal genetic diagnosis was provided. RESULTS: Fourteen cases (21.21%) were found to have GJB2 mutations by both methods (homozygous 235delC mutation in 3 cases, homozygous 176del16 mutation in 2 cases, 235delC and 299delAT compound heterozygous mutation in 2 cases, 299delAT and 176del16 compound heterozygous mutation in 1 case, c.339T > G and 313del12bp compound heterozygous mutation 1 case, and 235delC heterozygous mutation in 5 cases). 13 (19.70%) had SLC26A4 mutations (IVS7-2 A >G homozygous mutation in 2 cases, IVS7-2 A > G homozygous mutation in 2 cases, IVS7-2 A > G and 2168A > G compound heterozygous mutation in 3 cases, 2168A>G heterozygous mutation in 3 cases, and IVS7-2 heterozygous mutation in 3 cases); and 3 had mtDNA12S rRNA mutation (1555A > G mutation in 2 cases, 1494C > T mutation in 1 case). Prenatal diagnosis suggested that 3 fetuses have carried a heterozygous mutation. Two fetuses were detected as normal and confirmed to have normal hearing after birth. Two fetuses were found to have carried compound mutations of GJB2. CONCLUSION: Gene chip combined with GJB2 gene analysis is an accurate and effective method for the diagnosis of NSHI. The results can facilitate accurate prenatal diagnosis.

[Efficiency of multiplex ligation-dependent probe amplification combined with short tandem repeat linkage analysis for the prenatal diagnosis for Duchenne muscular dystrophy].

OBJECTIVE: To investigate the efficiency of multiplex ligation-dependent probe amplification (MLPA) combined with short tandem repeat (STR) linkage analysis for the prenatal diagnosis for Duchenne muscular dystrophy (DMD). METHODS: Gender of the fetus was first determined by the presence of Y chromosome sex-determining gene (SRY). Subsequently, combined MLPA and STR linkage analysis were applied for the probands, pregnant women and fetuses in 45 affected families. RESULTS: Among the 45 families, 31 SRY-positive fetuses were identified, among whom six were diagnosed with DMD. For 14 SRY-negative fetuses, four were diagnosed as carriers. The remainders were normal. CONCLUSION: MLPA can detect mutations in the exons of dystrophin gene, whilst STR linkage analysis can determine whether the fetus has inherited the maternal X chromosome bearing the mutant gene. As the result, the method can detect affected fetuses in which no exonic mutations are detected with MLPA. By combining the two methods, the diagnostic rate for DMD can be greatly improved.

Microbiota-ear-brain interaction is associated with generalized anxiety disorder through activation of inflammatory cytokine responses.

Introduction: Generalized anxiety disorder (GAD) is one of the most enduring anxiety disorders, being associated with increased systemic inflammation. However, the trigger and mechanisms underlying the activation of inflammatory cytokine responses in GAD remain poorly understood. Materials and methods: We characterized the ear canal microbiome in GAD patients through 16S rRNA gene sequencing and metagenomic sequencing and identified the serum inflammatory markers in GAD patients. Spearman correlations were applied to test the relationship between the microbiota changes and systemic inflammation. Results: Our findings showed the higher microbial diversity, accompanied with the significantly increased abundance of Proteobacteria, and decreased abundance of Firmicutes in the ear canal of GAD participants compared to that of the age- and sex-matched healthy controls (HC). Metagenomic sequencing showed that Pseudomonas aeruginosa were significantly increased at species-level in GAD patients. Furthermore, we observed the relative abundance of Pseudomonas aeruginosa was positively associated with elevated systemic inflammatory markers and the severity of disease, suggesting that these ear canal microbiota alterations might be correlated with GAD by activating the inflammatory response. Conclusions: These findings indicate that microbiota-ear-brain interaction via upregulating inflammatory reaction involve in the development of GAD, as well as suggest that ear canal bacterial communities may be a target for therapeutic intervention.