Fibronectin and Integrin α5 play overlapping and independent roles in regulating the development of pharyngeal endoderm and cartilage.

Craniofacial skeletal elements are derived from cranial neural crest cells (CNCCs), which migrate along discrete paths and populate distinct pharyngeal arches, structures that are separated by the neighboring endodermal pouches (EPs). Interactions between the CNCCs and the endoderm are critical for proper craniofacial development. In zebrafish, integrin α5 (Itga5) functions in the endoderm to regulate formation of specifically the first EP (EP1) and the development of the hyoid cartilage. Here we show that fibronectin (Fn), a major component of the extracellular matrix (ECM), is also required for these developmental processes, and that the penetrance of defects in mutants is temperature-dependent. fn1a-/- embryos exhibited defects that are similar to, but much more severe than, those of itga5-/- embryos, and a loss of integrin av (itgav) function enhanced both endoderm and cartilage defects in itga5-/- embryos, suggesting that Itga5 and Itgav cooperate to transmit signals from Fn to regulate the development of endoderm and cartilage. Whereas the endodermal defects in itga5; itga5v-/- double mutant embryos were comparable to those of fn1a-/- mutants, the cartilage defects were much milder. Furthermore, Fn assembly was detected in migrating CNCCs, and the epithelial organization and differentiation of CNCC-derived arches were impaired in fn1a-/- embryos, indicating that Fn1 exerts functions in arch development that are independent of Itga5 and Itgav. Additionally, reduction of itga5 function in fn1a-/- embryos led to profound defects in body axis elongation, as well as in endoderm and cartilage formation, suggesting that other ECM proteins signal through Itga5 to regulate development of the endoderm and cartilage. Thus, our studies reveal that Fn1a and Itga5 have both overlapping and independent functions in regulating development of the pharyngeal endoderm and cartilage.

Comprehensive Analysis of Sideroflexin 4 in Hepatocellular Carcinoma by Bioinformatics and Experiments.

Background: Sideroflexins (SFXNs) are a family of highly conserved mitochondrial transporters which regulate iron homeostasis and mitochondrial respiratory chain. However, the roles and mechanisms of SFXNs in HCC remain unknown. Methods: SFXNs expression and prognostic value in HCC was comprehensively analyzed. Proteins interacting with SFXN4 were analyzed in STRING database. The co-expression genes of SFXN4 were analyzed in cBioPortal database, and function of SFXN4 co-expression genes were annotated. The putative transcription factors and miRNA targeting SFXN4 were analyzed in NetworkAnalyst. The correlation between SFXN4 expression and immune infiltration was analyzed by ssGSEA. Cancer pathway activity and drug sensitivity related to SFXN4 were explored in GSCALite. The roles of SFXN4 in proliferation, migration and invasion of HCC were assessed in vitro and in vivo. Results: SFXN4 was consistently elevated in HCC, positively correlated with clinicopathological characteristics and predicted poor outcome. Functional enrichment showed SFXN4 was mainly related to oxidative phosphorylation, reactive oxygen species and metabolic pathways. SFXN4 expression was regulated by multiple transcription factors and miRNAs, and SFXN4 expression in HCC was associated with several cancer pathways and drug sensitivity. SFXN4 expression correlated with immune infiltration in HCC. In vitro, knockdown of SFXN4 inhibited HCC proliferation, migration and invasion, and decreased the expression of cyclin D1 and MMP2. In vivo, knockdown of SFXN4 inhibited the growth of tumor xenografts in mice. Conclusion: SFXN4 was upregulated in HCC, predicted poor prognosis, and may facilitate HCC development and progression via various mechanisms. For HCC, SFXN4 may provide both prognostic information and therapeutic potential.

Transcriptomics and Phenotypic Analysis of gpr56 Knockout in Zebrafish.

The adhesion G-protein-coupled receptor is a seven-transmembrane receptor protein with a complex structure. Impaired GPR56 has been found to cause developmental damage to the human brain, resulting in intellectual disability and motor dysfunction. To date, studies on gpr56 deficiency in zebrafish have been limited to the nervous system, and there have been no reports of its systemic effects on juvenile fish at developmental stages. In order to explore the function of gpr56 in zebrafish, the CRISPR/Cas9 gene-editing system was used to construct a gpr56-knockout zebrafish. Subsequently, the differentially expressed genes (DEGs) at the transcriptional level between the 3 days post fertilization (dpf) homozygotes of the gpr56 mutation and the wildtype zebrafish were analyzed via RNA-seq. The results of the clustering analysis, quantitative PCR (qPCR), and in situ hybridization demonstrated that the expression of innate immunity-related genes in the mutant was disordered, and multiple genes encoding digestive enzymes of the pancreatic exocrine glands were significantly downregulated in the mutant. Motor ability tests demonstrated that the gpr56-/- zebrafish were more active, and this change was more pronounced in the presence of cold and additional stimuli. In conclusion, our results revealed the effect of gpr56 deletion on the gene expression of juvenile zebrafish and found that the gpr56 mutant was extremely active, providing an important clue for studying the mechanism of gpr56 in the development of juvenile zebrafish.

Clinical benefit of tunnel endoscopic submucosal dissection for esophageal squamous cancer: a multicenter, randomized controlled trial.

BACKGROUND AND AIMS: Endoscopic submucosal dissection (ESD) is widely accepted as a primary treatment modality for dysplastic and early cancerous lesions of the GI tract. However, prolonged procedure time and life-threatening adverse events remain obstacles to the successful treatment of esophageal cancer. This study aimed to compare the efficacy and safety of tunnel ESD (T-ESD) with conventional ESD (C-ESD) for superficial esophageal squamous neoplasms. METHODS: A prospective, multicenter trial was conducted at 5 hospitals in China. Patients with esophageal squamous neoplasms were enrolled and randomly assigned to undergo C-ESD or T-ESD. Randomization was stratified by tumor location and circumference extent (<1/2 or ≥1/2). The primary endpoint was procedure time. RESULTS: Between January and July 2018, 160 patients were enrolled. One hundred fifty-two patients (76 in the C-ESD group and 76 in the T-ESD group) were included in the final analysis. The median procedure time was 47.3 minutes (interquartile range, 31.7-81.3) for C-ESD and 40.0 minutes (interquartile range, 30.0-60.0) for T-ESD (P = .095). However, T-ESD specifically reduced the median procedure time 34.5% (29.5 minutes) compared with C-ESD for lesions ≥1/2 circumference (P < .001). Among the multiple secondary outcomes, muscular injury was less frequent in the T-ESD group compared with the C-ESD group (18.4% vs 38.2%, P = .007), but complete healing of artificial mucosal defect in 1-month follow-up was more common in the T-ESD group than the C-ESD group (95.9% vs 84.7%, P =.026). CONCLUSIONS: Our study suggests that T-ESD results in shorter procedure time, specifically for lesions ≥1/2 circumference of the esophagus. In addition, T-ESD has a better safety profile indicated by less frequent muscular injury and improved healing of artificial mucosal defects caused by ESD procedures. (Clinical trial registration number: NCT03404921.).

Screening and Mechanism of Novel Angiotensin-I-Converting Enzyme Inhibitory Peptides in X. sorbifolia Seed Meal: A Computer-Assisted Experimental Study Method.

Angiotensin-I-converting enzyme (ACE) inhibitors are used extensively to control hypertension. In this study, a computer-assisted experimental approach was used to screen ACE-inhibiting peptides from X. sorbifolum seed meal (XSM). The process conditions for XSM hydrolysis were optimized through the orthogonal experimental method combined with a database. The optimal conditions for ACE inhibition included an alkaline protease dose of 5%, 45 °C, 15 min and pH 9.5. The hydrolysate was analyzed by LC-MS/MS, and 10 optimal peptides were screened. Molecular docking results revealed four peptides (GGLPGFDPA, IMAVLAIVL, ETYFIVR, and INPILLPK) with ACE inhibitory potential. At 0.1 mg/mL, the synthetic peptides GGLPGFDPA, ETYFIVR, and INPILLPK provided ACE inhibition rates of 24.89%, 67.02%, and 4.19%, respectively. GGLPGFDPA and ETYFIVR maintained high inhibitory activities during in vitro digestions. Therefore, the XSM protein may be a suitable material for preparing ACE inhibitory peptides, and computer-assisted experimental screening is an effective, accurate and promising method for discovering new active peptides.

Evaluation of the effects of an artificial intelligence system on endoscopy quality and preliminary testing of its performance in detecting early gastric cancer: a randomized controlled trial.

BACKGROUND: Esophagogastroduodenoscopy (EGD) is a prerequisite for detecting upper gastrointestinal lesions especially early gastric cancer (EGC). An artificial intelligence system has been shown to monitor blind spots during EGD. In this study, we updated the system (ENDOANGEL), verified its effectiveness in improving endoscopy quality, and pretested its performance in detecting EGC in a multicenter randomized controlled trial. METHODS: ENDOANGEL was developed using deep convolutional neural networks and deep reinforcement learning. Patients undergoing EGD in five hospitals were randomly assigned to the ENDOANGEL-assisted group or to a control group without use of ENDOANGEL. The primary outcome was the number of blind spots. Secondary outcomes included performance of ENDOANGEL in predicting EGC in a clinical setting. RESULTS: 1050 patients were randomized, and 498 and 504 patients in the ENDOANGEL and control groups, respectively, were analyzed. Compared with the control group, the ENDOANGEL group had fewer blind spots (mean 5.38 [standard deviation (SD) 4.32] vs. 9.82 [SD 4.98]; P < 0.001) and longer inspection time (5.40 [SD 3.82] vs. 4.38 [SD 3.91] minutes; P < 0.001). In the ENDOANGEL group, 196 gastric lesions with pathological results were identified. ENDOANGEL correctly predicted all three EGCs (one mucosal carcinoma and two high grade neoplasias) and two advanced gastric cancers, with a per-lesion accuracy of 84.7 %, sensitivity of 100 %, and specificity of 84.3 % for detecting gastric cancer. CONCLUSIONS: In this multicenter study, ENDOANGEL was an effective and robust system to improve the quality of EGD and has the potential to detect EGC in real time.

Norovirus Outbreak Surveillance, China, 2016-2018.

CaliciNet China, a network of provincial, county, and city laboratories coordinated by the Chinese Centers for Disease Control and Prevention, was launched in October 2016 to monitor the epidemiology and genotype distribution of norovirus outbreaks in China. During October 2016-September 2018, a total of 556 norovirus outbreaks were reported, and positive fecal samples from 470 (84.5%) outbreaks were genotyped. Most of these outbreaks were associated with person-to-person transmission (95.1%), occurred in childcare centers or schools (78.2%), and were reported during November-March of each year (63.5%). During the 2-year study period, 81.2% of all norovirus outbreaks were typed as GII.2[P16]. In China, most norovirus outbreaks are reported by childcare centers or schools; GII.2[P16] is the predominant genotype. Ongoing surveillance by CaliciNet China will provide information about the evolving norovirus genotype distribution and outbreak characteristics important for the development of effective interventions, including vaccines.

Endoderm convergence controls subduction of the myocardial precursors during heart-tube formation.

Coordination between the endoderm and adjacent cardiac mesoderm is crucial for heart development. We previously showed that myocardial migration is promoted by convergent movement of the endoderm, which itself is controlled by the S1pr2/Gα13 signaling pathway, but it remains unclear how the movements of the two tissues is coordinated. Here, we image live and fixed embryos to follow these movements, revealing previously unappreciated details of strikingly complex and dynamic associations between the endoderm and myocardial precursors. We found that during segmentation the endoderm underwent three distinct phases of movement relative to the midline: rapid convergence, little convergence and slight expansion. During these periods, the myocardial cells exhibited different stage-dependent migratory modes: co-migration with the endoderm, movement from the dorsal to the ventral side of the endoderm (subduction) and migration independent of endoderm convergence. We also found that defects in S1pr2/Gα13-mediated endodermal convergence affected all three modes of myocardial cell migration, probably due to the disruption of fibronectin assembly around the myocardial cells and consequent disorganization of the myocardial epithelium. Moreover, we found that additional cell types within the anterior lateral plate mesoderm (ALPM) also underwent subduction, and that this movement likewise depended on endoderm convergence. Our study delineates for the first time the details of the intricate interplay between the endoderm and ALPM during embryogenesis, highlighting why endoderm movement is essential for heart development, and thus potential underpinnings of congenital heart disease.

S1pr2/Gα13 signaling regulates the migration of endocardial precursors by controlling endoderm convergence.

Formation of the heart tube requires synchronized migration of endocardial and myocardial precursors. Our previous studies indicated that in S1pr2/Gα13-deficient embryos, impaired endoderm convergence disrupted the medial migration of myocardial precursors, resulting in the formation of two myocardial populations. Here we show that endoderm convergence also regulates endocardial migration. In embryos defective for S1pr2/Gα13 signaling, endocardial precursors failed to migrate towards the midline, and the presumptive endocardium surrounded the bilaterally-located myocardial cells rather than being encompassed by them. In vivo imaging of control embryos revealed that, like their myocardial counterparts, endocardial precursors migrated with the converging endoderm, though from a more anterior point, then moved from the dorsal to the ventral side of the endoderm (subduction), and finally migrated posteriorly towards myocardial precursors, ultimately forming the inner layer of the heart tube. In embryos defective for endoderm convergence due to an S1pr2/Gα13 deficiency, both the medial migration and the subduction of endocardial precursors were impaired, and their posterior migration towards the myocardial precursors was premature. This placed them medial to the myocardial populations, physically blocking the medial migration of the myocardial precursors. Furthermore, contact between the endocardial and myocardial precursor populations disrupted the epithelial architecture of the myocardial precursors, and thus their medial migration; in embryos depleted of endocardial cells, the myocardial migration defect was partially rescued. Our data indicate that endoderm convergence regulates the medial migration of endocardial precursors, and that premature association of the endocardial and myocardial populations contributes to myocardial migration defects observed in S1pr2/Gα13-deficient embryos. The demonstration that endoderm convergence regulates the synchronized migration of endocardial and myocardial precursors reveals a new role of the endoderm in heart development.

Characterization of the new GII.17 norovirus variant that emerged recently as the predominant strain in China.

Human noroviruses are the most important viral pathogens causing epidemic acute gastroenteritis, in which the GII.4 viruses have been predominant worldwide for the past decades. During 2014-2015 winter season, a new GII.17 variant emerged as the predominant virus in China surpassing the GII.4 virus in causing significantly increased acute gastroenteritis outbreaks. Genome sequences of the new GII.17 variant was determined and compared with other GII.17 noroviruses, revealing residue substitutions at specific locations, including the histo-blood group antigen-binding site and the putative antigenic epitopes. Further study of GII.17 outbreaks focusing on host susceptibility showed that the new GII.17 variant infected secretor individuals of A, B, O and Lewis types. Accordingly, the P particles of the new GII.17 variant bound secretor saliva samples of A, B, O and Lewis types with significantly higher binding signals than those of the P particles of the previous GII.17 variants. In addition, human sera collected from the outbreaks exhibited stronger blockade against the binding of the new GII.17 P particles to saliva samples than those against the binding between the P particles of previous GII.17 variants and saliva samples. Taken together, our data strongly suggested that the new GII.17 variant gained new histo-blood group antigen-binding ability and antigenic features, which may contribute to its predominance in causing human norovirus epidemics.

Circulation of Coxsackievirus A6 in hand-foot-mouth disease in Guangzhou, 2010-2012.

BACKGROUND: Hand, foot and mouth disease (HFMD) is usually caused by Enterovirus 71(EV71), and Coxsackievirus A16 (CV-A16) in Guangzhou, the biggest city of South China. However, Coxsackievirus A6 (CV-A6) were observed increased dramatically from 2010-2012. METHODS: In order to understand and to describe the epidemiologic and genetic characteristics of CV-A6, specimens of 5482 suspected HFMD cases were collected and examined by real-time fluorescence PCR. All samples positive for enteroviruses were analyzed by descriptive statistics. Phylogenetic analysis of CV-A6 based on the VP1 sequences was performed to investigate molecular and evolutionary characteristics. RESULTS: Coxsackievirus A6 increased dramatically from 9.04% in 2010 to 23.21% in 2012 and became one of the main causative agents of HFMD in Guangzhou. CV-A6 attack rates were highest in one to two year olds (33.14%). Typical clinic symptoms of CV-A6 HFMD include fever (589/720, 81.81%), maculopopular rash and vesicular exanthema around the perioral area (408/720, 56.66%), intraoral (545/720, 75.69%), the buttock (395/720, 54.86%), the trunk (244/720, 33.89%), the knee (188/720, 26.11%), and the dorsal aspects of hands (437/720, 60.69%). Phylogenetic analysis showed the CV-A6 isolates in this study belonged to Cluster A1 and were similar to those found in Shanghai in 2011 and 2012 (JX495148, KC414735), Shenzhen in 2011 (JX473394), Japan in 2011 (AB649243, AB649246), France in 2010(HE572928), Thailand in 2012(JX556564) and Israel in 2012 and 2013(.KF991010, KF991012).

[Sequence analysis of VP1 region of coxsackievirus A4 and coxsackievirus A10 in Guangzhou city, 2010-2012].

OBJECTIVE: To identify the enterovirus from stool samples of patients with hand, foot and mouth disease(HFMD) in Guangzhou from 2010 to 2012 and to perform phylogenetic analysis of the VP1 gene sequences of coxsackievirus A4 and coxsackievirus A10. METHODS: A total of 5 484 samples of suspected cases of HFMD which Guangzhou Center for Disease Control received from 2010 to 2012 were collected.Virus RNA was tested by nested RT-PCR method as human enterovirus 71, coxsackievirus A16, coxsackievirus A4, coxsackievirus A10 and other enteroviruses positive, and 4 111 samples were positive. Phylogenetic tree was constructed by partial VP1 gene sequences of coxsackievirus A4 and coxsackievirus A10 to perform phylogenetic analysis. RESULTS: In 4 111 enterovirus-positive samples, the positive rate of EV71, CoxA16, CoxA10 and CoxA4 was 35.1% (1 443/4 111) , 30.7% (1 261/4 111) , 2.0% (82/4 111),0.8% (31/4 111) respectively. Different enterovirus-positive rate was statistically significant (χ(2) = 148.34, P < 0.05) .Incidences of coxsackievirus A4 positive was highest in 3-year old children as 1.3% (7/534) , and that of coxsackievirus A10 positive was highest in 0-year old children as 3.7% (34/914) . The highest positive rate of diagnosed coxsackievirus A4 positive cases was admitted in April(2.6%, 12/460) , and the highest positive rate of diagnosed coxsackievirus A10 positive cases was admitted in August 4.3% (12/278). Phylogenetic analysis indicated that all the CoxA4 stains were divided into subtype A and subtype B, and the CoxA10 stains were divided into subtypes A, subtype B and subtype C. The VP1 gene nucleotide sequences of CoxA4 and CoxA10 this study measured both belonged to subtype A. CONCLUSIONS: The VP1 gene nucleotide sequences of CoxA4 and CoxA10 in Guangzhou from 2010 to 2012 both belonged to subtype A.

Zebrafish cobll1a regulates lipid homeostasis via the RA signaling pathway.

Background: The COBLL1 gene has been implicated in human central obesity, fasting insulin levels, type 2 diabetes, and blood lipid profiles. However, its molecular mechanisms remain largely unexplored. Methods: In this study, we established cobll1a mutant lines using the CRISPR/Cas9-mediated gene knockout technique. To further dissect the molecular underpinnings of cobll1a during early development, transcriptome sequencing and bioinformatics analysis was employed. Results: Our study showed that compared to the control, cobll1a -/- zebrafish embryos exhibited impaired development of digestive organs, including the liver, intestine, and pancreas, at 4 days post-fertilization (dpf). Transcriptome sequencing and bioinformatics analysis results showed that in cobll1a knockout group, the expression level of genes in the Retinoic Acid (RA) signaling pathway was affected, and the expression level of lipid metabolism-related genes (fasn, scd, elovl2, elovl6, dgat1a, srebf1 and srebf2) were significantly changed (p < 0.01), leading to increased lipid synthesis and decreased lipid catabolism. The expression level of apolipoprotein genes (apoa1a, apoa1b, apoa2, apoa4a, apoa4b, and apoea) genes were downregulated. Conclusion: Our study suggest that the loss of cobll1a resulted in disrupted RA metabolism, reduced lipoprotein expression, and abnormal lipid transport, therefore contributing to lipid accumulation and deleterious effects on early liver development.

[The molecular epidemiology characteristics of norovirus in environment and clinical samples in Guangzhou from 2009 to 2011].

OBJECTIVE: To investigate the molecular epidemiological characteristics of norovirus in Guangzhou from 2009 to 2011. METHODS: A total of 183 water samples, 1162 seafood samples and 1066 diarrhea stool specimens were collected from January 2010 to May 2011, June 2009 to June 2011 and July 2009 to December 2010 respectively in Guangzhou. Norovirus was detected by real time reverse transcript-PCR (qRT-PCR). The partial polymerase gene was amplified from norovirus positive samples, then sequenced and compared with the sequences of norovirus in GenBank. The phylogenetic tree was created. RESULTS: The positive rate was 19.67% (36/183), 8.26% (96/1162) and 37.05% (395/1066) in water samples, seafood and diarrhea patients respectively. Noroviruses from positive samples could be divided into 10 representative strains, in which 7 representative strains of genotype of 208 samples was type G2-4. The sequences from water, seafood and stool specimens were highly homologous with the similarity of 94% - 100%. CONCLUSION: In Guangzhou, the predominant Norovirus genotype was G2-4 and the positive rate of samples was high.

Roles of miR-196a and miR-196b in Zebrafish Motor Function.

BACKGROUND: The exertion of motor function depends on various tissues, such as bones and muscles. miR-196 has been widely studied in cancer and other fields, but its effect on bone and skeletal muscle is rarely reported. In order to explore the role of miR-196 family in bone and skeletal muscle, we used the previously successfully constructed miR-196a-1 and miR-196b gene knockout zebrafish animal models for research. METHODS: The behavioral trajectories of zebrafish from 4 days post-fertilization (dpf) to 7 dpf were detected to analyze the effect of miR-196a-1 and miR-196b on motor ability. Hematoxylin-eosin (HE) staining and transmission electron microscopy (TEM) were used to detect the dorsal muscle tissue of zebrafish. The bone tissue of zebrafish was detected by microcomputed tomography (micro-CT). Real-time PCR was used to detect the expression levels of related genes, including vcp, dpm1, acta1b, mylpfb, col1a1a, bmp8a, gdf6a, and fgfr3. RESULTS: The behavioral test showed that the total behavioral trajectory, movement time, and movement speed of zebrafish larvae were decreased in the miR-196a-1 and miR-196b gene knockout lines. Muscle tissue analysis showed that the structure of muscle fibers in the zebrafish lacking miR-196a-1 and miR-196b was abnormal and was characterized by vacuolar degeneration of muscle fibers, intranuclear migration, melanin deposition, and inflammatory cell infiltration. Bone CT examination revealed decreased bone mineral density and trabecular bone number. The real-time PCR results showed that the expression levels of vcp, dpm1, gdf6a, fgfr3, and col1a1a were decreased in the miR-196b gene knockout group. The expression levels of dpm1, acta1b, mylpfb, gdf6a, and col1a1a were decreased, and the expression level of fgfr3 was increased in the miR-196b gene knockout group compared with the wild-type group. CONCLUSIONS: miR-196a-1 and miR-196b play an important role in muscle fiber structure, bone mineral density, and bone trabecular quantity by affecting the expression of vcp, dpm1, acta1b, mylpfb, gdf6a, fgfr3, and col1a1a and then affect the function of the motor system.

Soybean protein isolate treated with transglutaminase (TGase) enhances the heat tolerance of selected lactic acid bacteria strains to spray drying.

TGase treated soy protein isolate (SPI) was confirmed to improve the survival of Lb. bulgaricus CICC 6047 subjected to spray drying. The viability of this strain increased from 70.69 % to 86.01 % due to elevating thermal stability. The smoother bacterial cell surface was observed by TGase treated SPI. TGase treatment changed the secondary conformation of SPI, having α-helical structure increased. Chemical bonds (hydrogen bond, CN bond, NH bond) proved the enzyme-modified SPI to have enhanced resistance to heat. The addition of TGase treated SPI allowed other LAB strains to tolerate the spray drying better. Their survival percentage after spray drying was 83.44 % for Le. mesenteroides CICC 6055, 80.64 % for Lb. acidophilus NCFM, 87.79 % for B. animalis BI-03 and 87.02 % for Lb. plantarum CICC 6009, respectively. The good survival of the selected LAB strains from TGase treated SPI powders was observed during storage of 8 weeks at 4 °C.

MTAP-ANRIL gene fusion promotes melanoma epithelial-mesenchymal transition-like process by activating the JNK and p38 signaling pathways.

Gene fusions caused by cytogenetic aberrations play important roles in the initiation and progression of cancers. The recurrent MTAP-ANRIL fusion gene was reported to have a frequency of greater than 7% in melanoma in our previous study. However, its functions remain unclear. Truncated MTAP proteins resulting from point mutations in the last three exons of MTAP can physically interact with the wild-type MTAP protein, a tumor suppressor in several human cancers. Similarly, MTAP-ANRIL, which is translated into a truncated MTAP protein, would influence wild-type MTAP to act as an oncogene. Here, we found that MTAP-ANRIL gene fusion downregulated the expression of wild-type MTAP and promoted epithelial-mesenchymal transition-like process through the activation of JNK and p38 MAPKs in vitro and in vivo. Our results suggest that MTAP-ANRIL is a potential molecular prognostic biomarker and therapeutic target for melanoma.

cdh23 affects congenital hearing loss through regulating purine metabolism.

Introduction: The pathogenic gene CDH23 plays a pivotal role in tip links, which is indispensable for mechanoelectrical transduction in the hair cells. However, the underlying molecular mechanism and signal regulatory networks that influence deafness is still largely unknown. Methods: In this study, a congenital deafness family, whole exome sequencing revealed a new mutation in the pathogenic gene CDH23, subsequently; the mutation has been validated using Sanger sequencing method. Then CRISPR/Cas9 technology was employed to knockout zebrafish cdh23 gene. Startle response experiment was used to compare with wide-type, the response to sound stimulation between wide-type and cdh23-/-. To further illustrate the molecular mechanisms underlying congenital deafness, comparative transcriptomic profiling and multiple bioinformatics analyses were performed. Results: The YO-PRO-1 assay result showed that in cdh23 deficient embryos, the YO-PRO-1 signal in inner ear and lateral line neuromast hair cells were completely lost. Startle response experiment showed that compared with wide-type, the response to sound stimulation decreased significantly in cdh23 mutant larvae. Comparative transcriptomic showed that the candidate genes such as atp1b2b and myof could affect hearing by regulating ATP production and purine metabolism in a synergetic way with cdh23. RT-qPCR results further confirmed the transcriptomics results. Further compensatory experiment showed that ATP treated cdh23-/- embryos can partially recover the mutant phenotype. Conclusion: In conclusion, our study may shed light on deciphering the principal mechanism and provide a potential therapeutic method for congenital hearing loss under the condition of CDH23 mutation.

A Serotype-Specific and Multiplex PCR Method for Whole-Genome Sequencing of Dengue Virus Directly from Clinical Samples.

Dengue virus (DENV) is the most globally prevalent member of the genus Flavivirus in the family Flaviviridae, which can be classified into four serotypes. Historically, molecular epidemiological studies of DENV depended on E gene sequencing. The development of next-generation sequencing (NGS) allowed its application to viral whole-genome sequencing (WGS). In this study, we report the improvement of the existing WGS process for DENV by optimizing the primer design procedure, designing serotype-specific primer panels and reducing the sizes of amplicons. A total of 31 DENV-positive serum samples belonging to 4 serotypes and 9 genotypes of DENV were involved in the validation of the primer panels. The threshold cycle (CT) values of these samples ranged from 23.91 to 35.11. The validation results showed that the length of consensus sequences generated at a coverage depth of 20× or more ranged from 10,370 to 10,672 bp, with 100.00% coverage of the open reading frames and 97.34% to 99.52% coverage of the DENV genome. The amplification efficiency varied across amplicons, genotypes, and serotypes of DENVs. These results indicate that the serotype-specific primer panels allow users to obtain the whole genome of DENV directly from clinical samples, providing a universal, rapid, and effective tool for the integration of genomics with dengue surveillance. IMPORTANCE Dengue virus (DENV) is becoming the most globally prevalent arbovirus. The number of people living under the threat of DENV is increasing year by year. With the development of next-generation sequencing (NGS) technology, whole-genome sequencing (WGS) has been more and more widely used in infectious disease surveillance and molecular epidemiological studies. DENV population sequencing by NGS can increase our understanding of the changing epidemiology and evolution of the DENV genome at the molecular level, which demands universal primer panels and combination with NGS platforms. Multiplex PCR with a short-amplicon approach proved superior for amplifying viral genomes from clinical samples, particularly when the viral RNA was present at low concentrations. Additionally, DENV are known for their genetic diversity within serotype groups and geographical regions, so the primer panels we designed focused on universality, which would be useful in future local DENV outbreaks.

Contact tracing period and epidemiological characteristics of an outbreak of the SARS-CoV-2 Delta variant in Guangzhou.

OBJECTIVES: An outbreak of the SARS-CoV-2 Delta variant occurred in Guangzhou in 2021. This study aimed to identify the transmission dynamics and epidemiological characteristics of the Delta variant outbreak to formulate an effective prevention strategy. METHODS: A total of 13102 close contacts and 69 index cases were collected. The incubation period, serial interval, and time interval from the exposure of close contacts to the symptom onset of cases were estimated. Transmission risks based on the exposure time and various characteristics were also assessed. RESULTS: The mean time from exposure to symptom onset among non-household presymptomatic transmission was 3.83 ± 2.29 days, the incubation period was 5 days, and the serial interval was 3 days. The secondary attack rate was high within 4 days before onset and 4-10 days after symptom onset. Compared with other contact types, household contact had a higher transmission risk. The transmission risk increased with the number and frequency of contact with index cases. Cycle threshold (Ct) values were associated with lower transmission risk (adjusted odds ratio [OR] 0.93 [95% CI 0.88-0.99] for ORF 1ab gene; adjusted OR 0.91 [95% CI 0.86-0.97] for N gene). CONCLUSION: The contact tracing period may need to be extended to 4 days before symptom onset. The low Ct value of index cases, the high number and frequency of contact with index cases, and household contacts were associated with a higher transmission risk of SARS-CoV-2 Delta.