Background::Bladder cancer, characterized by a high potential of tumor recurrence, has high lifelong monitoring and treatment costs. To date, tumor cells with intrinsic softness have been identified to function as cancer stem cells in several cancer types. Nonetheless, the existence of soft tumor cells in bladder tumors remains elusive. Thus, our study aimed to develop a microbarrier microfluidic chip to efficiently isolate deformable tumor cells from distinct types of bladder cancer cells.Methods::The stiffness of bladder cancer cells was determined by atomic force microscopy (AFM). The modified microfluidic chip was utilized to separate soft cells, and the 3D Matrigel culture system was to maintain the softness of tumor cells. Expression patterns of integrin β8 (ITGB8), protein kinase B (AKT), and mammalian target of rapamycin (mTOR) were determined by Western blotting. Double immunostaining was conducted to examine the interaction between F-actin and tripartite motif containing 59 (TRIM59). The stem-cell-like characteristics of soft cells were explored by colony formation assay and in vivo studies upon xenografted tumor models. Results::Using our newly designed microfluidic approach, we identified a small fraction of soft tumor cells in bladder cancer cells. More importantly, the existence of soft tumor cells was confirmed in clinical human bladder cancer specimens, in which the number of soft tumor cells was associated with tumor relapse. Furthermore, we demonstrated that the biomechanical stimuli arising from 3D Matrigel activated the F-actin/ITGB8/TRIM59/AKT/mTOR/glycolysis pathways to enhance the softness and tumorigenic capacity of tumor cells. Simultaneously, we detected a remarkable up-regulation in ITGB8, TRIM59, and phospho-AKT in clinical bladder recurrent tumors compared with their non-recurrent counterparts.Conclusions::The ITGB8/TRIM59/AKT/mTOR/glycolysis axis plays a crucial role in modulating tumor softness and stemness. Meanwhile, the soft tumor cells become more sensitive to chemotherapy after stiffening, that offers new insights for hampering tumor progression and recurrence.

Objective:To analyze the molecular mechanism of Zhenqi Fuzheng Capsules in the adjuvant treatment of AIDS by network pharmacology method and molecular docking technology.Methods:The active components and targets of Zhenqi Fuzheng Capsules were obtained through TCMSP, and the AIDS-related targets were obtained through GeneCards, OMIM and DrugBank databases. The intersection target PPI network was constructed through STRING 11.5 database, and Cytoscape 3.9.1 software was used for network topology analysis; Metascape database was used for GO function and KEGG pathway enrichment analysis of core targets; Cytoscape 3.9.1 was used to construct Zhenqi Fuzheng Capsules component-target-pathway network; Autodock Tools software was used to carry out molecular docking of core targets and active components.Results:Totally 31 active components and 180 targets of Zhenqi Fuzheng Capsules were screened out. TNF, IL6, AKT1, IL1B, TP53, VEGFA, RELA, EGFR and CASP3 were identified as the core targets. GO functional enrichment analysis obtained 1 436 biological processes, 53 cellular components, and 117 molecular functions. KEGG pathway enrichment analysis obtained 167 pathways, which were related to pathways in cancer, AGE-RAGE signaling pathway in diabetic complications, and IL-17 signaling pathway was closely related. Molecular docking results showed that core targets such as AKT1 and TNF had good binding activity to quercetin, kaempferol, and luteolin.Conclusion:The main active components of Zhenqi Fuzheng Capsules in the adjuvant treatment of AIDS are quercetin, kaempferol and luteolin, which may treat AIDS through the IL-17 signaling pathway.

Postzygotic mutations are acquired in normal tissues throughout an individual's lifetime and hold clues for identifying mutagenic factors.Here,we investigated postzygotic mutation spectra of healthy individuals using optimized ultra-deep exome sequencing of the time-series samples from the same volunteer as well as the samples from different individuals.In blood,sperm,and muscle cells,we resolved three common types of mutational signatures.Signatures A and B represent clock-like mutational processes,and the polymorphisms of epigenetic regulation genes influence the pro-portion of signature B in mutation profiles.Notably,signature C,characterized by C＞T transitions at GpCpN sites,tends to be a feature of diverse normal tissues.Mutations of this type are likely to occur early during embryonic development,supported by their relatively high allelic frequencies,presence in multiple tissues,and decrease in occurrence with age.Almost none of the public datasets for tumors feature this signature,except for 19.6％of samples of clear cell renal cell carcinoma with increased activation of the hypoxia-inducible factor 1(HIF-1)signaling pathway.Moreover,the accumulation of signature C in the mutation profile was accelerated in a human embryonic stem cell line with drug-induced activation of HIF-1α.Thus,embryonic hypoxia may explain this novel signature across multiple normal tissues.Our study suggests that hypoxic condition in an early stage of embryonic development is a crucial factor inducing C＞T transitions at GpCpN sites;and indi-viduals'genetic background may also influence their postzygotic mutation profiles.

OBJECTIVE: To carry out genome-wide copy number variations (CNVs) analysis for a boy with autism by using single nucleotide polymorphism array (SNP array). METHODS: SNP array analysis was conducted for the boy and his parents, and the data was validated by real-time PCR. Correlation between the deleted genes and the phenotype was analyzed by reviewing the literature. RESULTS: The patient was found to carry a terminal deletion of 18q22.3q23 (7.1 Mb), which involved FBXO15, ZNF407, ZADH2, TSHZ1, MBP and ADNP2 genes. No pathogenic CNVs were found in the parents. Comparison of the patient with cases reported in the literature suggested that the ZNF407 gene probably accounts for the autistic phenotype in these patients. CONCLUSION The autistic phenotype of the patient may be attributed to the 18q deletion, for which ZNF407 may be a critical candidate. SNP array has provided an useful tool for the study of molecular mechanism underlying autism.
Autistic Disorder ; genetics ; DNA Copy Number Variations ; Humans ; Male ; Microarray Analysis ; Polymorphism, Single Nucleotide

Objective: To investigate whether miR-140 could increase the sensitivity of cervical cancer (CC) to oxaliplatin by downregulating the expression of programmed death-1 (PD-L1). Methods: qPCR was used to analyze miR-140 expression in normal human cervical cells, CC cells and oxaliplatin-resistant CC cells. Cells were transfected with miR-140 mimic, and then, the proliferation of CC cells and oxaliplatin-resistant CC cells was detected by using CCK-8 assay, and the colony formation rate of CC cells was obtained by using colony formation assay. Starbase and TargetScan were used to predict the targeted binding site of miR-140 and PD-L1, and the influence of miR-140 on the expression of PD-L1 was validated by dual luciferase reporter gene assay.Annexin V FITC/PI double staining and Wb assays were used to detect the effect of over-expression of miR-140 or both over-expression of PD-L1 and miR140 on the apoptosis, migration and expression of apoptosis-related proteins in CC cells after treatment with oxaliplatin. Moreover, transplantation tumor of CC cell lines was established in nude mice to assess the effects of miR-140 on enhancing the sensitivity of tumors to oxaliplatin. Results: The expression of miR-140 was significantly decreased in oxaliplatin-resistant CC cells (P<0.01). Over-expression of miR140 could significantly increase the sensitivity of oxaliplatin-resistant CC cells to oxaliplatin (P<0.05), and inhibit the CC cells proliferation and colony formation (P<0.01). miR-140 showed targeted binding to PD-L1 3'-UTR and inhibited its expression. Over-expression of miR-140 significantly promoted CC cell migration and apoptosis (P<0.01). However, co-transfection of PD-L1 counteracts the effects of miR-140 on cell metastasis and apoptosis (all P<0.05). In addition, xenograft tumor model in mice also verified that miR-140 could promote the sensitivity of tumors to oxaliplatin. Conclusion: miR-140 increases the sensitivity of CC to oxaliplatin through inhibition of PD-L1 expression. Therefore, up-regulation of miR-140 or down-regulation of PD-L1 in combination with oxaliplatin may be a novel strategy for the treatment of Oxaliplatin-resistant CC.

OBJECTIVETo explore the clinical features and genomic abnormality of a patient with Coffin-Siris syndrome.

METHODSMicrodeletion and microduplication were detected with chromosomal microarray analysis (CMA) and verified with real-time quantitative PCR.

RESULTSThe patient, a 6-month-old boy, featured global development delay, thick eyebrows, low frontal hairline, long eyelash, flat nasal bridge, hypotonia, difficulty in turning over, over stretching of head, and hypoplatic nails. He could not stand stability or actively grasp. He also has characteristics of rickets. Chromosome karyotype of the patient was normal. Genomic analysis has detected a 1.3 Mb deletion in 6q25.3 region encompassing the ARID1B gene. Neither of his parents was found to harbor the same deletion.

CONCLUSIONThe 6q25.3 microdeletion probably underlies the Coffin-Siris syndrome in this patient, and rickets may be part of its clinical spectrum.

Objectives To explore the association between the single nucleotide polymorphism (SNP) rs2239669 in makorin ring-finger protein 3 (MKRN3) gene and the susceptibility to central precocious puberty (CPP). Methods A case-control study including 246 children with CPP and 269 healthy children was performed.The genotype and MKRN3 expression levels of patients were analyzed by PCR-HRM and RT-PCR,respectively. Results SNP rs2239669 genotype (TT,TC,CC) and allele frequencies (T and C) were different between cases and controls,with higher CC genotype in CPP patients. Under recessive model (CC/TT+TC),CC genotype was higher in CPP group and associated with higher risk of CPP (95%CI:1.062-2.143,P=0.021). MKRN3 expression levels were different among patients with different genotypes,of which TT genotype had the highest level followed by TC and CC (0.376±0.094, 0.330±0.068, 0.250±0.072, P=0.041). Conclusions MKRN3 SNP rs2239669 was associated with increased risk of CPP, and patients with TT genotype had higher MKRN3 levels.

Objective To investigate the clinical manifestations and motor neuron and pancreas homeobox 1 (MNX1) gene mutation features of Currarino syndrome.Methods Microdeletion and microduplication of the patients were detected by chromosomal microarray analysis (CMA),and literature review was performed for the clinical syndrome of Currarino syndrome with similar genotype.Results Two patients with Currarino syndrome were recruited in this study.Patient 1,a 7-day girl,came to hospital because of recurrent vomiting.Physical examinations showed coarse facial features,vision problems,serious abdominal flatulence and anal stenosis.Bowel imaging revealed malrotation of the midgut;and the magnetic resonance imaging (MRI) showed tethered spinal cord and malformation of sacrococcygeal vertebra.A 7.89 Mb deletion in chromosome 7 q36.lq36.3 region including MNX1 gene and a 2.20 Mb duplication in 14q32.33 area was found by using CMA.Patient 2,a 1 year and 3 months girl,came to hospital with global development delay.Clinical examination showed facial dysmorphic,growth retardation,intellectural disability,ptosis in right eye and anal stenosis.This patient had developmental retardation in language and movement.MRI showed spina bifida occulta.And a 15.00 Mb deletion in chromosome 7 q35q36.3 region was found including MNX1 gene.Literature review revealed that deletions in MNX1 gene led to Currarino syndrome with coarse facial features,growth retardation and intellectural disability,and this type of Currarino syndrome had not been reported in China.Conclusions Two cases of Currarino syndrome caused by microdeletion in 7q36 are reported for the first time in China,and this study can help clinicians to have a better understanding of this disease.

Objective To learn the cognitive status for newborns medical pain in medical workers of neonatal intensive care unit of children′s hospitals and provide clinical basis for optimizing neonatal pain management. Methods Totally 320 medical staff in neonatal intensive care units of six children′s hospital in Wuhan, Shenzhen, Shanghai, Nanjing, Zhejiang, Chongqing were investigated using medical pain management survey questionnaires by means of convenience sampling method. Results The knowledge on neonatal pain were at quite high level, accuracy rate was (80.5 ± 18.1)%;but on pain feel, pain relief rate and anodyne-use, the accuracy rate was relatively low; chest drainage, lumbar puncture and PICC placement were generally considered to cause severe pain with scores of 8.03±2.20, 7.17±2.17, 7.09 ± 2.00, respectively. Medical staff with different degrees, having children or not, different titles and with different years of working at NICU had impact on neonatal pain assessment on heel prick and venipuncture among full-term newborns. The difference was statistically significant (χ2=6.208-23.314, P<0.05). Conclusions Neonatal pain management is gradually emphasized by clinical medical staff, but there exist a certain bias on the perception and assessment of pain. So, it needs to strengthen pain-related knowledge training on health care for newborns to prevent and reduce neonatal pain, improving neonatal pain management.

Objective To analyze the clinical features and gene mutation in mthylmalonic acidemia (MMA) accompanied by homocysteinemia (cblC), and review the relevant literatures. Methods The clinical features of 3 cases of MMA diagnosed by gene detection were retrospectively analyzed, and meanwhile the pertinent literatures of pathogenesis of MMA, especially combined with late-onset cblC and its gene detection, were reviewed. Results Patient 1 (26 days old) suffered from intermittent convulsions for 3 days, with isosuccinic acid 175.8 μmol/L, C3/C2 rate 1.363, homocysteine >?65 μmol/L and abnormal EEG. MMACHC gene detection found an exon deficiency (delEXON1), which has not been reported. Patient 2 ( 12 year old) was hospitalized for limb shaking, hyperspasmia and vomiting. His isosuccinic acid level was 334.3 μmol/L, C3/?C2 rate was 0.37, homocysteine >?65 μmol/L, and had abnormal EEG. MMACHC gene detection found the mutations of c.482G?>?A and c.609G?>?A. Patient 3 was hospitalized for intermittent convulsions for 20 days, whose isosuccinic acid, C3/?C2 rate, and homocysteine were increased. MMACHC gene detection found the mutations of c.394C?>?T and c.540del8 and c.540del8 had not been reported. Review of literatures discovered that MMA was combined with epileptic seizure in some patents, which further validate that the mutation in MMACHC gene c.482G?>?A may be related to the late-onset of cblC. Conclusions Gene detection contributes to the diagnosis of MMA; the mutation of MMACHC gene c.482G>A may be related to the late-onset of cblC; delEXON1 and c.540del8 are new mutations which have not been reported.