RNA Sequencing and Bioinformatics Analysis to Reveal Potential Biomarkers in Patients with Combined Allergic Rhinitis and Asthma Syndrome.

Introduction: Combined allergic rhinitis and asthma syndrome (CARAS) is a concurrent clinical or subclinical allergic symptom of diseases of the upper and lower respiratory tract. This study is the first to explore the expression profiles of mRNA, lncRNA, and circRNA in CARAS using RNA sequencing, which may provide insight into the mechanisms underlying CARAS. Material and Methods: Whole blood samples from nine participants (three CARAS patients, three AR patients, and three normal control participants) were subjected to perform RNA sequencing, followed by identification of differentially expressed lncRNAs (DElncRNAs), circRNAs (DEcircRNAs) and mRNAs (DEmRNAs). Then, lncRNA/circRNA-mRNA regulatory pairs were constructed, followed by functional analysis, immune infiltration analysis, drug prediction, and expression validation with RT-qPCR and ELISA. Results: The results showed that 61 DEmRNAs, 23 DElncRNAs and 3 DEcircRNAs may be related to the occurrence and development of CARAS. KRT8 may be implicated in the development of AR into CARAS. Three immunity-related mRNAs (IDO1, CYSLTR2, and TEC) and two hypoxia-related mRNAs (TKTL1 and VLDLR) were associated with the occurrence and development of CARAS. TEC may be considered a drug target for Dasatinib in treating CARAS. Several lncRNA/circRNA-mRNA regulatory pairs were identified in CARAS, including LINC00452/MIR4280HG/hsa_circ_0007272/hsa_circ_0070934-CLC, HEATR6-DT/LINC00639/LINC01783/hsa_circ_0008903-TEC, RP11-71L14.3-IDO1/SMPD3, RP11-178F10.2-IDO1/HRH4, and hsa_circ_0008903-CYSLTR2, which may indicate potential regulatory effects of lncRNAs/circRNAs in CARAS. Dysregulated levels of immune cell infiltration may be closely related to CARAS. Conclusion: The regulating effect of lncRNA/circRNA-immunity/hypoxia-related mRNA regulatory pairs may be involved in the occurrence and development of CARAS.

Newcastle Disease Virus (NDV)-based vaccine candidate against SARS-CoV-2 Omicron by intranasal immunization.

Despite global vaccination efforts, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) continues to evolve and spread globally. Currently, the development of affordable vaccine against Omicron variant of concern (VOC) is necessary. Here, we assessed the safety and immunogenicity of a SARS-CoV-2 vaccine consisting of a live Newcastle disease virus vector expressing the spike (S) protein of Omicron BA.1 administrated intranasally (IN) or intramuscularly (IM) in Golden Syrian hamster model. Immunogenicity studies showed that the prime-boost regimen elicited high antibody titers and the modified S antigen (Sm-F) could induce robust antibody response in low dosage immunization through IN route. Sera of the immunized hamsters provided effective cross-neutralizing activity against different Omicron variants, the prototype and delta strains of SARS-CoV-2. Moreover, the vaccine could provide complete immunoprotection in hamsters against the Omicron BA.1 challenge by either intranasal or intramuscular immunization. Overall, our study provides an alternative nasal vaccine against the SARS-CoV-2 Omicron variants.

A high throughput antiviral screening platform for alphaviruses based on Semliki Forest virus expressing eGFP reporter gene.

Alphaviruses, which contain a variety of mosquito-borne pathogens, are important pathogens of emerging/re-emerging infectious diseases and potential biological weapons. Currently, no specific antiviral drugs are available for the treatment of alphaviruses infection. For most highly pathogenic alphaviruses are classified as risk group-3 agents, the requirement of biosafety level 3 (BSL-3) facilities limits the live virus-based antiviral study. To facilitate the antiviral development of alphaviruses, we developed a high throughput screening (HTS) platform based on a recombinant Semliki Forest virus (SFV) which can be manipulated in BSL-2 laboratory. Using the reverse genetics approach, the recombinant SFV and SFV reporter virus expressing eGFP (SFV-eGFP) were successfully rescued. The SFV-eGFP reporter virus exhibited robust eGFP expression and remained relatively stable after four passages in BHK-21 â€‹cells. Using a broad-spectrum alphavirus inhibitor ribavirin, we demonstrated that the SFV-eGFP can be used as an effective tool for antiviral study. The SFV-eGFP reporter virus-based HTS assay in a 96-well format was then established and optimized with a robust Z' score. A section of reference compounds that inhibit highly pathogenic alphaviruses were used to validate that the SFV-eGFP reporter virus-based HTS assay enables rapid screening of potent broad-spectrum inhibitors of alphaviruses. This assay provides a safe and convenient platform for antiviral study of alphaviruses.

[Pollution Characteristics and Source Apportionment of Heavy Metals in Vegetable Field in the Three Gorges Reservoir Area (Chongqing Section)].

To explore the pollution characteristics, ecological risks, and sources of heavy metals, soil surface samples of vegetable fields in 14 typical districts of the Three Gorges Reservoir area (Chongqing section) were collected in October 2021. The contents of seven types of heavy metals (As, Cd, Cr, Cu, Ni, Pb, and Zn) were analyzed. Based on the single-factor pollution index, the Nemerow integrated pollution index and potential ecological risk coefficient of heavy metals were evaluated. Additionally, the effects of different planting years and methods (open-field and greenhouse planting) on soil heavy metal accumulation were analyzed. The results indicated that the mean concentrations of heavy metals in vegetable soils in the area were lower than the national risk screening values for soil contamination of agricultural land (GB 15618-2018) but higher than their background values in Chongqing. According to the single-factor pollution index method, Pb, Zn, Cu, and Cd showed negligible slight hazards. The Nemerow pollution index showed that the study area was at a slight hazard level, and the main factors were Ni and Cd. Heavy metal pollution was found in 91.4% of the soil samples (PN>1) with different degrees, and 9.19% of them were severely polluted. The potential ecological risk coefficient showed that the vegetable lands were polluted slightly, and 9.77% of soil samples polluted by Cd were at moderate ecological risk. According to cluster analysis, the sources of Cd-Cu-Pb-Zn and As-Cr-Ni were similar. The content of heavy metals in the open field and greenhouse showed an increasing trend with the increase in planting years, and the content of heavy metals in greenhouse soil were generally higher than that in open field soil.

Intranasal delivery of replicating mRNA encoding hACE2-targeting antibody against SARS-CoV-2 Omicron infection in the hamster.

The Omicron variant is sweeping the world, which displays striking immune escape potential through mutations at key antigenic sites on the spike protein, making broad-spectrum SARS-CoV-2 prevention or therapeutical strategies urgently needed. Previously, we have reported a hACE2-targeting neutralizing antibody 3E8, which could efficiently block both prototype SARS-CoV-2 and Delta variant infections in prophylactic mouse models, having the potential of broad-spectrum to prevent SARS-CoV-2. However, preparation of monoclonal neutralizing antibodies is severely limited by the time-consuming process and the relative high cost. Here, we utilized a modified VEEV replicon with two subgenomic (sg) promoters engineered to express the light and heavy chains of the 3E8 mAb. The feasibility and protective efficacy of replicating mRNA encoding 3E8 against Omicron infection in the hamster were demonstrated through the lung targeting delivery with the help of VEEV-VRP. Overall, we developed a safe and cost-effective platform of broad-spectrum to prevent SARS-CoV-2 infection.

Diagnostic value of computed tomography-based pulmonary artery to aorta ratio measurement in chronic obstructive pulmonary disease with pulmonary hypertension: A systematic review and meta-analysis.

OBJECTIVE: We conducted a meta-analysis to systematic assess the diagnostic value of computed tomography (CT)-based pulmonary artery to aorta (PA:A) ratio measurement in COPD with pulmonary hypertension (COPD-PH). METHODS: Published studies referring to diagnostic accuracy of PA:A ratio for COPD-PH were screened out from PubMed, Embase, Web of science, China National Knowledge databases (CNKI), Wan fang databases, and VIP databases. We used bivariate random-effects model to estimate pooled sensitivity (SEN), specificity (SPE), positive and negative likelihood ratios (PLR and NLR, respectively), and diagnostic odds ratios (DOR). Summary receiver operating characteristic (SROC) curves and area under the curve (AUC) were also calculated to summarize the aggregate diagnostic performance. RESULTS: Nine eligible studies were included and the pooled SEN was 69% (95% CI: 59 ~ 78), SPE was 85% (95% CI: 77 ~ 90), PLR was 4.5 (95% CI: 2.8 ~ 7.5), and NLR was 0.36 (95% CI: 0.26 ~ 0.51), respectively. DOR reached 13.00 (95% CI: 6.00 ~ 28.00), and value of AUC was 0.84 (95% CI: 0.81 ~ 0.87). Subgroup analysis indicated that when the value of PA:A ratio was equal or greater than one (PA/A ≥ 1), the combined SEN, SPE, AUC, and DOR was 69%, 89%, 0.90, and 19.65, respectively. CONCLUSIONS: PA:A ratio is helpful for appraisal of COPD-PH, and PA/A ≥ 1 possessed prominent diagnostic accuracy.

Long non-coding RNA growth arrest specific-5: a potential biomarker for early diagnosis of severe asthma.

BACKGROUND: The diagnosis of severe asthma (SA) is difficult due to a necessary long-term treatment history currently, while there are few studies on biomarkers in the diagnosis of SA. Long non-coding RNA (lncRNA) growth arrest specific-5 (GAS5) has the potential of playing this role because its binding with glucocorticoid receptor (GR). The purpose of this article is to explore the possibility of lncRNA GAS5 acting as a biomarker for early diagnosis of severe asthma (SA). METHODS: Peripheral blood was obtained from healthy volunteers, patients with non-severe asthma (nSA) and SA, and peripheral blood mononuclear cells (PBMCs) were separated. Twenty-four female BALB/c mice (aged 6 weeks) were randomly and averagely divided into 3 groups, i.e., control group, asthma group and dexamethasone group. The mice were sensitized and challenged with ovalbumin (OVA) and lipopolysaccharide (LPS) to establish a murine model of steroid-insensitive asthma. Human bronchial epithelial cells (HBECs) were cultured, transfected with miR-9 mimics, JNK1 inhibitor and treated with interleukin (IL)-2 + IL-4 and dexamethasone. Western blot was used to detect glucocorticoid receptor phosphorylation at serine 226 (GRser226), and quantitative real-time PCR was used to detect GAS5 level. RESULTS: The level of GAS5 in PBMCs from nSA group elevated 20-fold higher after dexamethasone treatment in vitro, while it reduced 15-fold lower in SA group (P<0.001). The expression of GRser226 in PBMCs from SA group was significantly higher than that from control group and nSA group after dexamethasone treatment (P<0.001). In the lung tissue of mice, the GAS5 level of dexamethasone group was lower than that of asthma group (P<0.001) and control group (P<0.05). Both treatment with IL-2 + IL-4 and transfection of miR-9 mimics could increase the expression of GRser226 in HBECs (P<0.001). The GAS5 level in HBECs after IL-2 + IL-4 + Dexamethasone treatment was lower than that in HBECs only treated with IL-2 + IL-4 (P<0.001). Similarly, dexamethasone treatment also decreased the level of GAS5 in HBECs transfected with miR-9 mimics (P<0.05). Moreover, transfecting with JNK1 inhibitor could reverse the expression of GAS5 in HBECs transfected with miR-9 mimics and treated with dexamethasone. However, the level of GAS5 in HBECs interfered with IL-2 + IL-4 + Dexamethasone was not affected by JNK1 inhibitor. CONCLUSIONS: The expression of GAS5 is different in PBMCs between nSA and SA, and is affected by glucocorticoids treatment, which is due to GRser226 phosphorylation. GAS5 can be used as a potential biomarker for diagnosis of severe asthma by comparing GAS5 level in PBMCs from patients before and after glucocorticoids treatment in vitro.

Comprehensive analysis of miRNA-mRNA-lncRNA networks in severe asthma.

AIM: This study aimed to explore the molecular mechanism of severe asthma. MATERIALS & METHODS: The shared and divergent differentially expressed mRNAs (DEmRNAs), miRNAs (DEmiRNAs) and lncRNAs (DElncRNAs) in asthma and severe asthma were identified by RNA-sequencing. Severe asthma-specific and shared DEmiRNA-DEmRNA-DElncRNA interaction networks were performed. RESULTS: Compared with normal control, 1328 DEmRNAs, 608 DElncRNAs and 63 DEmiRNAs were identified in severe asthma. Compared with asthma, 95 DEmRNAs, 143 DElncRNAs and 96 DEmiRNAs were identified in severe asthma. MiR-133a-3p-EFHD2/CNN2-AC144831.1 interactions and miR-3613-3p-CD44/BCL11B-LINC00158/CTA-217C2.1/AC010976.2/RP11-641A6.2 interactions were speculated to involve with the development of severe asthma. The results of GSE69683 validation were generally consistent with our RNA-sequencing results. CONCLUSION: This study provides clues for understanding the mechanism of severe asthma.

Comprehensive Analysis of miRNA-mRNA-lncRNA Networks in Non-Smoking and Smoking Patients with Chronic Obstructive Pulmonary Disease.

BACKGROUND/AIMS: Chronic obstructive pulmonary disease (COPD) is a major cause of morbidity and mortality worldwide. This study aimed to identify overlapping or diverging dysregulated genes, lncRNAs, miRNAs and signaling pathways in smoking and non-smoking chronic obstructive pulmonary disease (COPD). METHODS: Compared to normal controls, we identified the shared and divergent differentially expressed mRNAs (DEmRNAs), miRNAs (DEmiRNAs) and lncRNAs (DElncRNAs) in smoking and non-smoking COPD by RNA-sequencing and bioinformatics analysis. Functional annotation of DEmRNAs were performed. Both cis and trans-target DEmRNAs of DElncRNAs were identified. The target DEmRNAs of DEmiRNAs were identified as well. The DEmiRNA-DEmRNA-DElncRNA interaction network was constructed. QRT-PCR was performed to validat the selected DEmiRNAs, DEmRNA and DElncRNAs in COPD. RESULTS: Compared to normal control, 1234 DEmRNAs, 96 DElncRNAs and 151 DEmiRNAs were identified in non-smoking patients with COPD; 670 DEmRNAs, 44 DElncRNAs and 63 DEmiRNAs were identified in smoking patients with COPD. Leukocyte transendothelial migration and pathways in cancer were significantly enriched pathways in non-smoking and smoking COPD, respectively. MiR-122-5p-A2M-LINC00987/A2M-AS1/ linc0061 interactions might play key roles in COPD irrespective with the smoking status. Let-7-ADRB1-HLA-DQB1-AS1 might play a key role in the pathogenesis of smoking COPD while miR-218-5p/miR15a-RORA-LOC101928100/LINC00861 and miR-218-5p/miR15a-TGFß3-RORA-AS1 interactions might involve with non-smoking COPD. CONCLUSION: We identified the shared and diverging genes, lncRNAs, miRNAs and their interactions and pathways in smoking and non-smoking COPD which provided clues for understanding the mechanism and developing novel diagnostic and therapeutic strategies for COPD.

MicroRNA-7 inhibits cell proliferation, migration and invasion in human non-small cell lung cancer cells by targeting FAK through ERK/MAPK signaling pathway.

OBJECTIVE: To investigate the effects of microRNA-7 (miR-7) on the proliferation, migration and invasion of non-small cell lung cancer NSCLC) cells by targeting FAK through ERK/MAPK signaling pathway. METHODS: NSCLC tissues and adjacent normal tissues were obtained from 160 NSCLC patients after operation. NSCLC cell lines (A549, H1299 and H1355) and a normal human fetal lung fibroblast cell line (MRC-5) were obtained. NSCLC cells were assigned into miR-7 inhibitors, miR-7 mimics, blank, miR-7 mimics control, miR-7 inhibitors control, FAK siRNA and miR-7 inhibitors + FAK siRNA groups. The expressions of miR-7 and FAK mRNA in tissues and cell lines were detected by qRT-PCR and Western-Blotting. Cell proliferation, migration and invasion were detected by MTT assay, wound scratch assay and Transwell assay. RESULTS: Compared with adjacent normal tissues, miR-7 expression was down-regulated, but the mRNA and protein expressions of FAK, ERK and MAPK were up-regulated. Compared with the blank and mimics control groups, miR-7 significantly increased but FAK, ERK and MAPK expressions decreased in miR-7 mimics and FAK siRNA groups. Cell proliferation, migration and invasion were inhibited in the miR-7 mimics and FAK siRNA groups, while opposite regarding miR-7 inhibitors group. CONCLUSION: The miR-7 can inhibit the activation of ERK/MAPK signaling pathway by down-regulating FAK expression, thereby suppressing the proliferation, migration and invasion of NSCLC cells. The miR-7 and its target gene FAK may be novel targets for the diagnosis and treatment of NSCLC.