LINC01305 recruits basonuclin 1 to act on G-protein pathway suppressor 1 to promote esophageal squamous cell carcinoma.

EsophageaL squamous cell carcinoma (ESCC) is one of the most common and lethal tumors, however, its underlying molecular mechanisms are not completely understood and new therapeutic targets are needed. Here, we found that the transcription factor basonuclin 1 (BNC1) was significantly upregulated and closely related to the differentiation and metastasis of ESCC. Furthermore, BNC1, LINC01305, and G-protein pathway suppressor 1 (GPS1) had significant oncogenic roles in ESCC. In addition, in vivo experiments showed that knockdown of BNC1 indeed significantly inhibited the proliferation and metastasis of ESCC. We also revealed the molecular mechanism by which LINC01305 recruits BNC1 to the promoter of GPS1, and then GPS1 could mediate the JNK signaling pathway to promote the proliferation and metastases of ESCC. Taken together, we discovered the novel molecular mechanism by which LINC01305/BNC1 upregulates GPS1 expression to promote the development of ESCC, providing a new therapeutic target for ESCC.

SCARA5 induced ferroptosis to effect ESCC proliferation and metastasis by combining with Ferritin light chain.

BACKGROUND: Esophageal squamous cell carcinoma (ESCC) remains one of the most lethal cancers worldwide accompany with an extremely poor prognosis. Therefore, this study aims to screen for new molecules affecting ESCC and explore their mechanisms of action to provide ideas for targeted therapies for ESCC. METHODS: Firstly, we screened out the membrane protein SCARA5 by high-throughput sequencing of the ESCC patient tissues, and RT-qPCR and WB were used to verify the differential expression of SCARA5 in esophageal cell lines, and IHC analyzed the expression localization of SCARA5 in ESCC tissue. Then, flow cytometry, wound healing assay, Transwell assay and CCK-8 assay were used to explore the effects of SCARA5 on cell cycle, migration and invasion as well as cell proliferation activity of esophageal squamous carcinoma cells. Meanwhile, transmission electron microscopy was used to detect changes in cellular mitochondrial morphology, and flow cytometry were used to detect changes in intracellular reactive oxygen metabolism, and immunofluorescence and flow cytometry were used to detect changes in intracellular Fe2+. Mechanistically, co-immunoprecipitation was used to detect whether SCARA5 binds to ferritin light chain, and ferroptosis-related protein expression was detected by WB. Finally, the tumor xenograft model was applied to validation the role of SCARA5 tumor growth inhibition in vivo. RESULTS: We found that SCARA5 was aberrantly decreased in ESCC tissues and cell lines. Furthermore, we confirmed that SCARA5 suppressed the cell cycle, metastasis and invasion of ESCC cells. Meanwhile, we also found that overexpression of SCARA5 caused changes in mitochondrial morphology, accumulation of intracellular reactive oxygen species and increased intracellular Fe2+ in ESCC cells, which induced ferroptosis in ESCC cells. Mechanically, we validated that SCARA5 combined with ferritin light chain and increased intracellular Fe2+. As well as, overexpression SCARA5 induced ferroptosis by increasing ferritin light chain in nude mice subcutaneous tumors and inhibited the growth of nude mice subcutaneous tumors. CONCLUSION: Collectively, our findings demonstrated that SCARA5 suppressed the proliferation and metastasis of ESCC by triggering ferroptosis through combining with ferritin light chain.

Systematic profiling identifies PDLIM2 as a novel prognostic predictor for oesophageal squamous cell carcinoma (ESCC).

Till now, no appropriate biomarkers for high-risk population screening and prognosis prediction have been identified for patients with oesophageal squamous cell carcinoma (ESCC). In this study, by the combined use of data from the Gene Expression Omnibus (GEO) datasets and The Cancer Genome Atlas (TCGA)-oesophageal carcinoma (ESCA), we aimed to screen dysregulated genes with prognostic value in ESCC and the genetic and epigenetic alterations underlying the dysregulation. About 222 genes that had at least fourfold change in ESCC compared with adjacent normal tissues were identified using the microarray data in GDS3838. Among these genes, only PDLIM2 was associated with nodal invasion and overall survival (OS) at the same time. The high PDLIM2 expression group had significantly longer OS and its expression was independently associated with better OS (HR: 0.64, 95% CI: 0.43-0.95, P = 0.03), after adjustment for gender and pathologic stages. The expression of its exon 7/8/9/10 had the highest AUC value (0.724) and better prognostic value (HR: 0.43, 95% CI: 0.22-0.83, P = 0.01) than total PDLIM2 expression. PDLIM2 DNA copy deletion was common in ESCC and was associated with decreased gene expression. The methylation status of two CpG sites (cg23696886 and cg20449614) in the proximal promoter region of PDLIM2 showed a moderate negative correlation with the gene expression in PDLIM2 copy neutral/amplification group. In conclusion, we infer that PDLIM2 expression might be a novel prognostic indicator for ESCC patients. Its exon 7/8/9/10 expression had the best prognostic value. Its down-regulation might be associated with gene-level copy deletion and promoter hypermethylation.

ENST00000489707.5 Is a Preferred Alternative Splicing Variant of PTK7 in Adrenocortical Cancer and Shows Potential Prognostic Value.

BACKGROUND This study aimed to explore the transcript preference of PTK7 in adrenocortical cancer (ACC), the prognostic value, and the potential underlying genetic alterations. MATERIAL AND METHODS Data from the Cancer Genome Atlas-Adrenocortical Cancer (TCGA-ACC) and the Genotype-Tissue Expression (GTEx)-normal adrenal gland were used for analysis. RESULTS A non-canonical alternative transcript, ENST00000489707.5, which only encodes an extracellular immunoglobulin (Ig)-like domain and an intracellular kinase domain, is the dominant isoform of PTK7 in both ACC and normal adrenal gland. Its expression percentage was significantly higher in ACC than in normal adrenal gland. ACC tissues showed preferred expression of this transcript compared with other cancers with known PTK7 expression. Prognostic analysis showed that ENST00000489707.5 had independent prognostic value in progression-free survival (PFS) (HR: 1.227, 95%CI: 1.077-1.398, p=0.002) and disease-specific survival (DSS) (HR: 1.419, 95%CI: 1.154-1.745, p=0.001) after adjustment of other risk factors. cg20819617 methylation was negatively correlated with both PTK7 and ENST00000489707.5 expression. CONCLUSIONS ENST00000489707.5 is a preferred alternative splicing product of PTK7, with a significantly increased proportion in ACC compared with other cancers. Its expression shows potential prognostic value in terms of PFS and DSS in ACC patients. The methylation status of cg20819617 might play a critical role in modulating PTK7 transcription and ENST00000489707.5 expression.

PTK7 is a novel oncogenic target for esophageal squamous cell carcinoma.

BACKGROUND: Overexpression of PTK7 has been found in multiple cancers and has been proposed to serve as a prognostic marker for intrahepatic cholangiocarcinoma. Its role in esophageal cancer, however, remains to be clarified. We hypothesize that PTK7 positively regulates tumorigenesis of esophageal cancer. METHODS: We examined PTK7 expression pattern in human esophageal squamous carcinoma by Oncomine expression analysis and by immunohistochemistry (IHC) staining. We knocked down PTK7 in two esophageal squamous cell carcinoma cell lines, TE-5, and TE-9, by siRNA, and evaluated cell proliferation, apoptosis, and migration ofPTK7-defective cells. Expressions of major apoptotic regulators and effectors were also determined by quantitative real-time PCR in PTK7-defective cells. We further overexpressed PTK7 in the cell to evaluate its effects on cell proliferation, apoptosis, and migration. RESULTS: Both Oncomine expression and IHC analyses showed that PTK7 is overexpressed in clinical esophageal squamous cell carcinoma tumors. PTK7 siRNA suppressed cell growth and promoted apoptosis of TE-5 and TE-9. PTK7-defective cells further displayed reduced cellular migration that was concomitant with upregulation of E-cadherin. Conversely, overexpression of PTK7 promotes cell proliferation and invasion, while apoptosis of the PTK7-overexpressing cells is repressed. Notably, major apoptotic regulators, such as p53 and caspases, are significantly upregulated in siPTK7 cells. CONCLUSIONS: PTK7 plays an oncogenic role in tumorigenesis and metastasis of esophageal squamous carcinoma. PTK7 achieves its oncogenic function in esophageal squamous cell carcinoma partially through the negative regulation of apoptosis.

Dose-dependent effect of glucose on GLP-1 secretion involves sweet taste receptor in isolated perfused rat ileum [Rapid Communication].

Luminal glucose is an important stimulus for glucagon-like peptide 1 (GLP-1) secretion from intestinal endocrine cells. However, the effects of luminal glucose concentration on GLP-1 secretion remain unknown. In this study, we investigated the effect of luminal glucose concentrations (3.5, 5, 10, 15, and 20 mmol/L) on GLP-1 secretion from isolated perfused rat ileum. Results showed that the perfusate glucose concentration dose-dependently stimulated GLP-1 secretion from isolated perfused rat ileum, which was eliminated by the sweet taste receptor inhibitor gurmarin (30 µg/mL), but not inhibited by phloridzin (1 mmol/L), a Na+-coupled glucose transporters inhibitor. We conclude that luminal glucose induced GLP-1 secretion from perfused rat ileum in a concentration-dependent manner. This secretion was mediated by sweet taste receptor transducing signal for GLP-1 release on the gut of rat.

The low IGFBP-3 level is associated with esophageal cancer patients: a meta-analysis.

BACKGROUND: Esophageal cancer was a vital cause of cancer-related mortality worldwide, and the insulin-like growth factor-binding proteins (IGFBPs) has been proved to be an important factor of multiple types of tumors. There is a controversy that whether the IGFBP-3 level is associated with the clinical pathological characteristics and overall survival of esophageal cancer patients. Herein, we aimed to comprehensively assess the association between the low IGFBP-3 level and the risk, overall survival and clinical pathological characteristics of esophageal cancer. METHOD: We conducted a meta-analysis using seven eligible studies. The overall odds ratios (OR)/relative risk (RR) and their corresponding 95% confidence interval (CI) were calculated for each parameter. RESULTS: For the risk of esophageal cancer, the OR was 2.342 (p = 0.000), indicating that individuals with lower IGFBP-3 level were more likely to suffer from esophageal cancer, compared to those with relatively high IGFBP-3 level. With respect to the 3-year survival rate, the RR was 2.163 (p = 0.027), which demonstrated that esophageal cancer patients with low IGFBP-3 level had significantly lower 3-year survival rate; in terms of clinical pathological characteristics, significantly lower IGFBP-3 level was found for patients in all categories; for survival status, patients in low IGFBP-3 level are more likely to be in the dead survival status (OR = 4.480, p = 0.000). CONCLUSION: Our meta-analysis suggests that for esophageal cancer, the low IGFBP-3 level is associated with high cancer risk, poor prognosis, and unfavorable tumor stage and metastasis.

A Positive Role of Cytokine-Induced Killer Cell Therapy on Gastric Cancer Therapy in a Chinese Population: A Systematic Meta-Analysis.

BACKGROUND: Chemotherapy and radiation therapy provide limited improvement in survival of gastric cancer patients after tumor resection. It is essential to develop a novel therapeutics for gastric cancer. In the recent years, cytokine-induced killer cells (CIKs)-based adoptive immune therapy has been explored in gastric cancer patients. Due to the small number of patients included in each clinical trial and low-power statistical analysis, the effectiveness of this approach is still unclear. To address this issue, we systemically analyzed the relevant clinical trial data published in recent years by powerful statistical meta-analysis. MATERIAL AND METHODS: Clinical data was searched by multiple electronic databases with a term "gastric cancer" and "cytokine-induced killer cells". Six relevant clinical trials with case-control studies were extracted for our meta-analysis, including 318 patients receiving CIK cell therapy and 369 patients receiving conventional therapy. RESULTS: Overall survival (OS) and odds ratio (OR) were analyzed for patients at 1, 2, 3, and 5 years post-CIK cell therapy and post-conventional therapy. Heterogeneity and publication bias were analyzed for included data quality and publication bias. Our meta-analysis from 6 clinical trials suggests that CIK cell therapy significantly increased 5-year OS from 27±2.44% to 49±7.62% (p<0.05) and 5-year OR up to 1.77 (p<0.05). The increased 5-year survival rate was also highly correlated with the increased CD3+ T cell number and ratio of CD4+/CD8+ in the CIK treated patients. CONCLUSIONS: CIK cell therapy significantly increased 5-year survival rate compared to conventional chemotherapy among gastric cancer patients. The study provides powerful statistical evidence for large-scale clinical trials with CIK cell therapy.

Clinical and genetic analysis of a case of Gitelman syndrome accompanied with Graves disease and adrenocortical adenoma: A case report.

RATIONALE: Gitelman syndrome (GS), also known as familial hypokalemia and hypomagnesemia, is a rare autosomal recessive inherited disease caused by primary renal desalinization caused by impaired reabsorption of sodium and chloride ions in the distal renal tubules. We report a case of clinical and genetic characteristics of GS accompanied with Graves disease and adrenocorticotrophic hormone (ACTH)-independent adrenocortical adenoma. PATIENT CONCERNS: The patient is a 45 year old female, was admitted to our hospital, due to a left adrenal gland occupying lesion as the chief complaint. DIAGNOSIS: The patient was finally diagnosed as GS with Graves disease and adrenocortical adenoma. INTERVENTIONS: Potassium magnesium aspartate (1788 mg/d, taken orally 3 times a day (supplement a few times a day, intake method, treatment duration). Contains 217.2 mg of potassium and 70.8 mg of magnesium, and potassium chloride (4.5 g/d, taken orally 3 times a day (supplement a few times a day, intake method, and treatment duration); Potassium 2356 mg), spironolactone (20 mg/d, taken orally once a day (supplement a few times a day, intake method, treatment duration). After 3 months of treatment, the patient's blood potassium fluctuated between 3.3-3.6 mmol/L, and blood magnesium fluctuated between 0.5-0.7 mmol/L, indicating a relief of fatigue symptoms. OUTCOMES: On the day 6 of hospitalization, the symptoms of dizziness, limb fatigue, fatigue and pain were completely relieved on patient. In the follow-up of the following year, no recurrence of the condition was found. LESSONS: The novel c.1444-10(IVS11)G > A variation may be a splicing mutation. The compound heterozygous mutations of the SLC12A3 gene may be the pathogenic cause of this GS pedigree.

Generation and characterization of mAb 61H9 against junctional adhesion molecule-a with potent antitumor activity.

Junctional adhesion molecule-A (JAM-A) is an adhesion molecule that exists on the surface of certain types of cells, including white blood cells, endothelial cells, and dendritic cells. In this study, the cDNA sequences of JAM-A-Fc were chemically synthesized with optimization for mammalian expression. Afterward, we analyzed JAM-A protein expression through transient transfection in HEK293 cell lines. Mice were immunized with JAM-A-Fc protein, and hybridoma was prepared by fusing myeloma cells and mouse spleen cells. Antibodies were purified from the hybridoma supernatant and four monoclonal strains were obtained and numbered 61H9, 70E5, 71A8, and 74H3 via enzyme-linked immunosorbent assay screening. Immunofluorescence staining assay showed 61H9 was the most suitable cell line for mAb production due to its fluorescence signal being the strongest. Flow cytometric analysis proved that 61H9 possessed high affinity. Moreover, antagonism of JAM-A mAb could attenuate the proliferative, migrative, and invasive abilities of ESCC cells and significantly inhibit tumor growth in mice. By examining hematoxylin-eosin staining mice tumor tissues, we found inflammatory cells infiltrated lightly in the anti-JAM-A group. The expression of BCL-2 and IκBα in the anti-JAM-A group were decreased in mice tumor tissues compared to the control group. Ultimately, a method for preparing high-yield JAM-A-Fc protein was created and a high affinity mAb against JAM-A with an antitumor effect was prepared.

Characterization of risks and pathogenesis of respiratory diseases caused by rural atmospheric PM2.5.

Forty-six percent of the world's population resides in rural areas, the majority of whom belong to vulnerable groups. They mainly use cheap solid fuels for cooking and heating, which release a large amount of PM2.5 and cause adverse effects to human health. PM2.5 exhibits urban-rural differences in its health risk to the respiratory system. However, the majority of research on this issue has focused on respiratory diseases induced by atmospheric PM2.5 in urban areas, while rural areas have been ignored for a long time, especially the pathogenesis of respiratory diseases. This is not helpful for promoting environmental equity to aid vulnerable groups under PM2.5 pollution. Thus, this study focuses on rural atmospheric PM2.5 in terms of its chemical components, toxicological effects, respiratory disease types, and pathogenesis, represented by PM2.5 from rural areas in the Sichuan Basin, China (Rural SC-PM2.5). In this study, organic carbon is the most significant component of Rural SC-PM2.5. Rural SC-PM2.5 significantly induces cytotoxicity, oxidative stress, and inflammatory response. Based on multiomics, bioinformatics, and molecular biology, Rural SC-PM2.5 inhibits ribonucleotide reductase regulatory subunit M2 (RRM2) to disrupt the cell cycle, impede DNA replication, and ultimately inhibit lung cell proliferation. Furthermore, this study supplements and supports the epidemic investigation. Through an analysis of the transcriptome and human disease database, it is found that Rural SC-PM2.5 may mainly involve pulmonary hypertension, sarcoidosis, and interstitial lung diseases; in particular, congenital diseases may be ignored by epidemiological surveys in rural areas, including tracheoesophageal fistula, submucous cleft of the hard palate, and congenital hypoplasia of the lung. This study contributes to a greater scientific understanding of the health risks posed by rural PM2.5, elucidates the pathogenesis of respiratory diseases, clarifies the types of respiratory diseases, and promotes environmental equity.

Covalent binding modes between BMP-2-derived peptides and graphene in 3D scaffolds determine their osteoinductivity and capacity for calvarial defect repair in vivo.

Covalent introduction of bioactive molecules is one of main strategies to significantly enhance the biological activities of bone repair materials. In this study, three most-commonly used chemical groups were respectively introduced on graphene (GP), followed by covalent binding with bone morphogenetic protein-2 (BMP-2) -derived peptides, ensuring that the same molar mass of peptides was bound to different functionalized GP (f-GP). Then the same amount of composites composed of different f-GP and peptides were respectively compounded with poly (lactic-co-glycolic acid) to fabricate 3D scaffolds. In vivo study demonstrated that the scaffolds containing ammonized GP covalently bound with the peptides through amide binding could reach best efficiency of promoting ectopic bone regeneration and repairing calvarial defect probably because the most positive charges on the peptide chain and surface of the ammonized GP could absorb more specific proteins in vivo and have better interactions with them, thereby differentiating most inducible cells into osteogenic cells. Our results indicate that the performances of scaffolds containing covalently bound bioactive molecules can be controlled by the covalent binding mode, and that our prepared scaffold containing ammonized GP covalently bound with the BMP-2-derived peptides through amide binding possess inspiring potential applicable prospects for bone tissue regeneration and engineering.

Nanopore Electrochemistry for Pathogen Detection.

Pathogen infections seriously threaten human health, and there is an urgent demand for rapid and efficient pathogen identification to provide instructions in clinical diagnosis and therapeutic intervention. Recently, nanopore technology, a rapidly maturing technology which delivers ultrasensitive sensing and high throughput in real-time and at low cost, has achieved success in pathogen detection. Furthermore, the remarkable development of nanopore sequencing, for example, the MinION sequencer from Oxford Nanopore Technologies (ONT) as a competitive sequencing technology, has facilitated the rapid analysis of disease-related microbiomes at the whole-genome level and on a large scale. Here, we highlighted recent advances in nanopore approaches for pathogen detection at the single-molecule level. We also overviewed the applications of nanopore sequencing in pathogenic bacteria identification and diagnosis. In the end, we discussed the challenges and future developments of nanopore technology as promising tools for the management of infections, which may be helpful to aid understanding as well as decision-making.

Molecular mechanism of Pulmonary diseases caused by exposure to urban PM2.5 in Chengdu-Chongqing Economic Circle, China.

Chengdu-Chongqing Economic Circle (CD-CQ Economic Circle) is one of China's four major economic circles and five major urban agglomerations located in Southwest China's Sichuan Basin. The CD-CQ Economic Circle, with its strong economic development and dense population, suffers from severe PM2.5 pollution, which is known to cause chronic and acute respiratory ailments. This study examined the lung disease-related hub genes, functions, and pathways that are affected by PM2.5 in summer and winter in the two central megacities of Chengdu and Chongqing. PM2.5 frequently activates lung disease-associated hub genes, most notably the transcription factor TP53. TP53 interacts with the majority of lung disease-related genes and regulates important and commonly occurring biological functions and pathways, including gland development, aging, reactive oxygen species metabolic process, the response to oxygen levels, and fluid shear stress, among others. Thus, PM2.5 has been shown to target TP53 for regulating lung disease genes/functions/pathways, thereby influencing the occurrence and progression of lung illnesses. Notably, PM2.5 may be associated with small cell carcinoma of the lung due to the high number of lung disease genes, hub genes, critical functions, and pathways enriched in this kind of cancer. These findings shed fresh light on the molecular pathophysiology of PM2.5 pollution on the respiratory system in the CD-CQ Economic Circle and aid in the development of novel techniques for mitigating PM2.5 pollution-associated respiratory illness.

Research advances in chimeric antigen receptor-modified T-cell therapy (Review).

Chimeric antigen receptor (CAR)-modified T-cells are T-cells that have been genetically engineered to express CAR molecules to target specific surface antigens on tumor cells. CAR T-cell therapy, a novel cancer immunotherapy, has been attracting increasing attention, since it exhibited notable efficacy in the treatment of hematological tumors in clinical trials. However, for this type of therapy, challenges must be overcome in the treatment of solid tumors. Furthermore, certain side effects associated with CAR T-cell therapy, including cytokine release syndrome, immune effector cell-related neurotoxicity syndrome, tumor lysis syndrome and on-target off-tumor toxicity, must be taken into consideration. The present study provides a systematic review of the principle, clinical application, current challenges, possible solutions and future perspectives for CAR T-cell therapy.

MicroRNA-382-5p inhibits osteosarcoma development and progression by negatively regulating VEZF1 expression.

Human osteosarcoma is the most frequent malignant primary bone tumor that mainly occurs in young adults and children. MicroRNAs (miRNAs/miRs) are abnormally expressed in human osteosarcoma and contribute to osteosarcoma initiation and development. The present study aimed to investigate the role of miR-382-5p in the nosogenesis of osteosarcoma and to identify a novel target for osteosarcoma treatment. miR-382-5p expression was detected in human osteosarcoma clinical tissues and cell lines, including 143B, U2OS and MG63, via reverse transcription-quantitative PCR analysis. Multiple bioinformatic prediction toowe used to identify the potential target genes of miR-382-5p and vascular endothelial zinc finger 1 (VEZF1), which were validated via the dual-luciferase reporter assay. MG63 and U2OS cells were transfected with miR-382-5p mimics. The Cell Counting Kit-8 assay was performed to assess cell proliferation, while the Transwell assay was performed to assess migration and invasion. Cell colony formation was measured via crystal violet staining, and apoptosis was assessed via Annexin V/propidium iodide staining. The wound healing assay was performed to assess the migratory ability of U2OS and MG63 cells. Antitumor effects of miR-382-5p were evaluated in nude mice xenografts using U2OS cells. The results demonstrated that miR-382-5p expression was markedly downregulated in human osteosarcoma tissues and cell lines compared with adjacent normal tissues. Transfection of miR-382-5p mimics into MG63 and U2OS cells significantly inhibited the malignant behaviors of cells, including decreased proliferation, migration, diminished colony formation and invasion, and promoted osteosarcoma cell apoptosis. Bioinformatics prediction indicated that VEZF1 is a direct target gene of miR-382-5p. Overexpression of VEZF1 restored osteosarcoma tumor development inhibited by miR-382-5p in vivo. In addition, overexpression of miR-382-5p restrained the growth of xenograft osteosarcoma in nude mice following co-transfection, and overexpression of VEZF1 attenuated the inhibitory effect of miR-382-5p in nude mice. miR-382-5p acted as a tumor suppressor gene and inhibited the malignant biological behaviors of human osteosarcoma cells and functions associated with directly targeting VEZF1. Taken together, these results suggest that the miR-382-5p/VEZF1 interaction has an important role in osteosarcoma development and progression, and thus may be used as a diagnostic and therapeutic target for osteosarcoma.

Low-Cost and Scalable Platform with Multiplexed Microwell Array Biochip for Rapid Diagnosis of COVID-19.

Sensitive detection of SARS-CoV-2 is of great importance for inhibiting the current pandemic of COVID-19. Here, we report a simple yet efficient platform integrating a portable and low-cost custom-made detector and a novel microwell array biochip for rapid and accurate detection of SARS-CoV-2. The instrument exhibits expedited amplification speed that enables colorimetric read-out within 25 minutes. A polymeric chip with a laser-engraved microwell array was developed to process the reaction between the primers and the respiratory swab RNA extracts, based on reverse transcriptase loop-mediated isothermal amplification (RT-LAMP). To achieve clinically acceptable performance, we synthesized a group of six primers to identify the conserved regions of the ORF1ab gene of SARS-CoV-2. Clinical trials were conducted with 87 PCR-positive and 43 PCR-negative patient samples. The platform demonstrated both high sensitivity (95.40%) and high specificity (95.35%), showing potentials for rapid and user-friendly diagnosis of COVID-19 among many other infectious pathogens.

The structure and unzipping behavior of dumbbell and hairpin DNA revealed by real-time nanopore sensing.

Hairpin structures play an essential role in DNA replication, transcription, and recombination. Single-molecule studies enable the real-time measurement and observation of the energetics and dynamics of hairpin structures, including folding and DNA-protein interactions. Nanopore sensing is emerging as a powerful tool for DNA sensing and sequencing, and previous research into hairpins using an α-hemolysin (α-HL) nanopore suggested that hairpin DNA enters from its stem side. In this work, the translocation and interaction of hairpin and dumbbell DNA samples with varying stems, loops, and toeholds were investigated systematically using a Mycobacterium smegmatis porin A (MspA) nanopore. It was found that these DNA constructs could translocate through the pore under a bias voltage above +80 mV, and blockage events with two conductance states could be observed. The events of the lower blockage were correlated with the loop size of the hairpin or dumbbell DNA (7 nt to 25 nt), which could be attributed to non-specific collisions with the pore, whereas the dwell time of events with the higher blockage were correlated with the stem length, thus indicating effective translocation. Furthermore, dumbbell DNA with and without a stem opening generated different dwell times when driven through the MspA nanopore. Finally, a new strategy based on the dwell time difference was developed to detect single nucleotide polymorphisms (SNPs). These results demonstrated that the unzipping behaviors and DNA-protein interactions of hairpin and dumbbell DNA could be revealed using nanopore technology, and this could be further developed to create sensors for the secondary structures of nucleic acids.

[Preliminary report on partial pressure changes of oxygen and carbon dioxide in umbilical artery and vein before and after the first breath in Chinese neonates II-The personalized analysis of partial pressure of oxygen and carbon dioxide difference between umbilical artery and vein at same time in same newborn].

Objective: In order to explore the mechanism of neonatal spontaneous breathing, the difference of oxygen and carbon dioxide between umbilical cord arteries and veins before the start of spontaneous breathing after birth has been analyzed among people. In this part, the related information is analyzed individually. Methods: After all fetal parents signed the informed consent before birth, and before the newborn was born and did not breathe, the umbilical cord was exposed as quickly as possible, and the heparinized arterial indwelling needle was inserted into the umbilical artery and umbilical vein in the direction of newborn and placenta, and then blood was taken continuously. Although dozens of mothers were selected,but only 3 cases were collected from Pua and Puv blood samplers at the same time for blood gas analysis and determination, and the differences and dynamic changes of umbilical vein and umbilical artery were calculated and analyzed. Results: In all 3 none spontaneous breathing newborns,PuvO2 was significantly higher than PuaO2 at the same time (P<0.01), with an average difference of (24.17±7.09) mmHg; while PuvCO2 was significantly lower than PuaCO2 (all P<0.01), with an average difference of (-7.67±3.70) mmHg.The difference of Puv-uaO2 was significantly higher than those of Puv-uaCO2 (P<0.05). Conclusion: PuaO2 decreases gradually with time (heartbeat frequency) before spontaneous breathing after the delivered fetus as a newborn, and it induces the first inhalation to start spontaneous breathing when it reaches the threshold of triggering breathing.

[Preliminary report on partial pressure changes of oxygen and carbon dioxide in umbilical artery and vein before and after the first breath in Chinese neonates I-The group difference of partial pressure of oxygen and carbon dioxide between umbilical artery and vein in newborns].

Objective: The fetus has no actual respiration, and the newborn begins to breathe after birth. We assume that the first breath dominantly generated by hypoxia. In this study, the changes and lowest limit of blood oxygen partial pressureof umbilical artery (PuaO2) after chemoreceptor were analyzed to explore the mechanism of neonatal spontaneous breathing. Methods: With signed consent form by all fetal parents before birth, 14 newborns successfully completed the umbilical artery or vein catheterization and drawn blood according to the heartbeat. All blood samples analyzed by blood gas analyzer,calculated and analyzed the similarities and differences between umbilical vein(Puv) and umbilical artery(Pua). Results: Although we completed 14 newborns, there were only 9 cases of umbilical artery samples and 8 cases of umbilical vein samples were collected. Only 3 cases collected both Pua and Puv blood samples at the same time (see serial paper II). PuaO2 in gradually decreased with time (heartbeat frequency), until Pua contracted after spontaneous breathing produced about 8~10 heartbeats, and then could not get enough blood samples. Only 3 newborns were able to take blood samples after spontaneous breathing for 8~10 heartbeats, and their PuaO2 were jumped to 186.0, 137.0 and 93.8 mmHg respectively. The mean value of PuaO2 was (25.94±6.79, 18.04~37.51)mmHg, the highest value was (29.11±6.46, 23.00~45.90)mmHg, and the lowest value was (21.34±5.54, 14.00~33.60)mmHg. Although PuvO2 decreased gradually with time (heartbeat) too, most of them also showed the tendency of alternately rising and falling with the regularity of mother's respiration. The mean value of PuvO2 was (53.35±21.35, 32.56~100.73)mmHg, the highest value was (90.38±48.44, 43.40~153.00)mmHg, and the lowest value was (36.96±14.90, 24.80~73.80)mmHg. Although there were large individual differences, the mean, highest and lowest values of PuvO2 were significantly higher than those of PuaO2 (P<0.05); although PuvCO2 slightly lower than PuaCO2, it was no significant difference (P>0.05). Conclusion: PuaO2 decreases gradually with time before spontaneous breathing after the delivered fetus as a newborn, and it induces the first inhalation to start spontaneous breathing when it reaches the threshold of triggering breathing.