Clinical characteristics and severity of beta and delta variants of SARS-CoV-2 and the effect of vaccine on delta variants.

Background: The Delta variant of concern (VOC) is rapidly becoming the dominant strain globally. We report the clinical characteristics and severity of hospitalized patients infected with Delta and Beta VOCs during the local outbreak in Harbin, Heilongjiang Province, China, and the effect of vaccines on the Delta variant. Methods: We collected a total of 735 COVID-19 patients from the First Affiliated Hospital of Harbin Medical University, including 96 cases infected with the Delta VOC and 639 cases infected with the Beta VOC. Demographic, clinical characteristic and laboratory findings were collected and compared. Results: Differences in viral shedding, IgG and IgM levels, and the neutrophil-to-lymphocyte ratio were noted between the Delta and Beta VOCs (p < 0.05). Survival analysis of the two groups revealed longer viral shedding of the Delta VOC (p < 0.05). For the Delta VOC, the longer the vaccination period, the lower the IgG and IgM levels. IgM levels were higher in the convalescent plasma group, whereas lymphocyte counts were lower. Conclusions: Delta VOC virus shedding was longer compared with Beta VOC shedding. Vaccination with inactivated vaccines can reduce the severe illness rate of the Delta VOC. IgG and IgM levels are reduced as the time period between the first and second vaccine doses increases.

Pyoluteorin Induces Apoptosis and Autophagy in NSCLC Cells.

Pyoluteorin is a natural occurring antibiotic and its anti-tumor activity has rarely been reported. This study aims to investigate the anti-tumor effects of pyoluteorin on human non-small cell lung cancer (NSCLC) cells. The cell proliferation was measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was determined through caspase3 activity assay and immunoblotting. Autophagy was measured by transmission electron microscope (TEM) and immunostaining. The autophagy-related proteins were detected through immunoblotting. We found that pyoluteorin showed significant anti-tumor effects on human NSCLC cell lines H1299 (IC50 = 1.57 µM) and H2030 (IC50 = 1.94 µM). Moreover, pyoluteorin could induce apoptosis and autophagy as evidence by the upregulation of caspase3 activity, the accumulation of LC3 and expression of apoptosis or autophagy related proteins. In addition, pyoluteorin induced autophagy through c-Jun N-terminal kinase/B-cell lymphoma-2 (JNK/Bcl-2) signal pathway. Blocking JNK/Bcl-2 pathway significantly attenuated pyoluteorin-induced autophagy. Moreover, inhibition of autophagy by 3-methyladenine (3-MA) or Beclin1 knockout greatly promoted pyoluteorin-induced apoptosis and cell death. Our results showed that pyoluteorin could induce both apoptosis and autophagy in human NSCLC cells. Combination of pyoluteorin with autophagy inhibitior significantly promoted pyoluteorin-induced apoptosis and may be a potential anticancer strategy in the NSCLC therapy.

Low Expression of Rasal2 Promotes Non-small Cell Lung Cancer Metastasis through Ras/ERK Pathway.

The RAS protein activator like 2 (Rasal2) has been reported to be a tumor suppressor in variety of cancers; while an oncogenic protein in ovarian cancer and triple negative breast cancer (TNBC). However, the exact role of Rasal2 in non-small cell lung cancer (NSCLC) is lacking. This study aimed to investigate the role of Rasal2 in NSCLC and the underlying mechanisms. Rasal2 expression level was measured in NSCLC tissue and cells by using quantitative (q)-PCR and immunoblotting analysis. The clinical implication of Rasal2 in NSCLC patients was also analyzed. The function role of Rasal2 in NSCLC cells were measured by small interfering RNA (si-RNA), immunostaining, transwell assay and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Low Rasal2 expression level was observed in human NSCLC tissue and cell lines and significantly related to tumor thickness, ulceration and TNM staging in NSCLC patients. Rasal2 knockdown significantly increased NSCLC cell invasion and migration. Mechanistically, we showed that Rasal2 knockdown significantly increased the phosphorylation level of extracellular signal-regulated kinase (ERK)/Raf1/mitogen-activated protein extracellular kinase (MEK) thus activated Ras/ERK signal pathway. Thus, our data showed that Rasal2 is downregulated in NSCLC cells and act as an epithelial-mesenchymal transition (EMT) and metastasis suppressor through the Ras/ERK pathway. Rasal2 may be a prognostic biomarker for NSCLC in the future.

Accumulated Clinical Experiences from Successful Treatment of 1377 Severe and Critically Ill COVID-19 Cases.

In late December 2019, COVID-19 was firstly recognized in Wuhan, China and spread rapidly to all of the provinces of China. The West Campus of Wuhan Union Hospital, the designated hospital to admit and treat the severe and critically ill COVID-19 cases, has treated a large number of such patients with great success and obtained lots of valuable experiences based on the Chinese guideline (V7.0). To standardize and share the treatment procedures of severe and critically ill cases, Wuhan Union Hospital has established a working group and formulated an operational recommendation, including the monitoring, early warning indicators, and several treatment principles for severe and critically ill cases. The treatment experiences may provide some constructive suggestions for treating the severe and critically ill COVID-19 cases all over the world.

Lung myofibroblast transition and fibrosis is regulated by circ0044226.

BACKGROUND AND PURPOSE: Idiopathic pulmonary fibrosis (IPF) is a life-threatening progressive disease characterized by aberrant fibroblast activation. This study aims to explore the role of the circ0044226 on fibroblast-to-myofibroblast transition (FMT). METHODS: Bleomycin and TGF-ß1 were respectively used to induce the IPF mice model and human lung fibroblasts to myofibroblast differentiation. The mRNA and protein levels were examined by qRT-PCR and western blot. Localization of α-SMA was evaluated by immunofluorescence staining. Cell viability and proliferation were evaluated by CCK8 and EDU test. Dual-luciferase reporter assay was used to analyze the interaction between miR-7 and circ0044226 or sp1. Fluorescence in situ hybridization (FISH) assay was used for the identification of sub-location of circ0044226 and miR-7 in cells. The IPF model mice received intratracheal injection of AAV-sh-NC and AAV-sh- circ0044226, and lung fibrosis was detected by HE staining, Masson staining and immunohistochemistry assay. RESULTS: The circ0044226 was upregulated while miR-7 was downregulated in IPF mice model and FMT-derived myofibroblasts. miR-7 was a target of circ0044226 and sp1 was a target of miR-7. circ0044226 was distributed mostly in the cytoplasm and functioned as a miR-7 sponge to positively regulate the expression of sp1. Intervention of circ0044226 could ameliorate FMT and suppress fibroblast viability and proliferation by functioning as an endogenous miR-7 sponge. CONCLUSION: Circ0044226 knockdown alleviates fibroblast proliferation and FMT by functioning as a competing endogenous RNA, which may represent a promising therapy for pulmonary fibrosis.

LncRNA PICART1 suppressed non-small cell lung cancer cells proliferation and invasion by targeting AKT1 signaling pathway.

LncRNAs play significant roles in various cell biological processes. In the present study, we demonstrated that PICART1 expression was down-regulated in non-small cell lung cancer (NSCLC) tissues. Lower expression level of PICART1 was associated with advanced stage. In addition, PICART1 expression was down-regulated in NSCLC cell lines. Overexpression of PICART1 inhibited NSCLC cell growth and induced cell cycle arrest at G2/M phase. Elevated expression of PICART1 suppressed NSCLC cell colony formation and cell invasion. Ectopic expression of PICART1 promoted the expression of epithelial marker E-cadherin while suppressed the mesenchymal marker expression such as N-cadherin and Snail and Vimentin. Furthermore, PICART1 overexpression suppressed AKT phosphorylation and c-Myc expression while inhibited the p21 expression in NSCLC cell. AKT phosphorylation was involved in PICART1 mediated suppression of cell growth and invasion. These results suggested that overexpression of PICART1 suppressed cell growth and invasion partly through regulating AKT signaling pathway in NSCLC.

Inhibition of bleomycin-induced pulmonary fibrosis by bone marrow-derived mesenchymal stem cells might be mediated by decreasing MMP9, TIMP-1, INF-γ and TGF-ß.

The study was aimed to investigate the mechanism and administration timing of bone marrow-derived mesenchymal stem cells (BMSCs) in bleomycin (BLM)-induced pulmonary fibrosis mice. Thirty-six mice were divided into six groups: control group (saline), model group (intratracheal administration of BLM), day 1, day 3 and day 6 BMSCs treatment groups and hormone group (hydrocortisone after BLM treatment). BMSCs treatment groups received BMSCs at day 1, 3 or 6 following BLM treatment, respectively. Haematoxylin and eosin and Masson staining were conducted to measure lung injury and fibrosis, respectively. Matrix metalloproteinase (MMP9), tissue inhibitor of metalloproteinase-1 (TIMP-1), γ-interferon (INF-γ) and transforming growth factor ß1 (TGF-ß) were detected in both lung tissue and serum. Histologically, the model group had pronounced lung injury, increased inflammatory cells and collagenous fibres and up-regulated MMP9, TIMP-1, INF-γ and TGF-ß compared with control group. The histological appearance of lung inflammation and fibrosis and elevation of these parameters were inhibited in BMSCs treatment groups, among which, day 3 and day 6 treatment groups had less inflammatory cells and collagenous fibres than day 1 treatment group. BMSCs might suppress lung fibrosis and inflammation through down-regulating MMP9, TIMP-1, INF-γ and TGF-ß. Delayed BMSCs treatment might exhibit a better therapeutic effect. Highlights are as follows: 1. BMSCs repair lung injury induced by BLM. 2. BMSCs attenuate pulmonary fibrosis induced by BLM. 3. BMSCs transplantation down-regulates MMP9 and TIMP-1. 4. BMSCs transplantation down-regulates INF-γ and TGF-ß. 5. Delayed transplantation timing of BMSCs might exhibit a better effect against BLM.

miR-99a suppresses the metastasis of human non-small cell lung cancer cells by targeting AKT1 signaling pathway.

MicroRNAs (miRNAs) play an important role in the development and progression of non-small cell lung cancer (NSCLC). Recently, several studies have shown that miR-99a is downregulated in various cancers, which can affect tumor initiation and maintenance. Herein, we found that miR-99a was downregulated in NSCLC tissues and suppressed tumor metastasis of NSCLC cells. Down-regulation of miR-99a is significantly associated with last-stage and tumor metastasis in NSCLC patients. Further functional experiments found that overexpression of miR-99a inhibit cell proliferation, migration, and invasion of NSCLC cells in vitro and tumor metastasis of NSCLC in vivo. In addition, we also found that AKT1 is directly involved in miR-99a-mediated tumor suppression. Restored the expression of AKT1 partially abolished the suppressive effects miR-99a on proliferation and invasion of NSCLC cells. Collectively, our data suggest that miR-99a plays an important role in the tumorigenesis and metastasis of NSCLC and may serve as a therapeutic target to avoid dissemination of NSCLC cells.

Identification of 4438 novel lincRNAs involved in mouse pre-implantation embryonic development.

Long intergenic non-coding RNAs (lincRNAs) as a key group of non-coding RNAs have gained substantial attention. Though lincRNAs have been systematically explored in various mouse tissues and cell lines, large-scale identification of lincRNAs in mouse pre-implantation embryonic development (PED) process has not be documented previously. Therefore, it is important to identify and characterize novel lincRNAs that may be involved in PED. In this paper, we performed transcriptome assembly based on published single-cell RNA-seq data during mouse PED and identified 4,438 putative lincRNAs. Combining these with Ensembl lincRNAs, we established a reference catalog of 5,808 transcribed lincRNAs in PED. We then systematically analyzed the lincRNAs in this reference catalog and revealed that the identified novel PED lincRNAs are generally comparable with known Ensembl lincRNAs in genomic aspects. In addition, the global expression patterns can be separated by zygote first cleavage division in clustering analysis and we further identified and analyzed differentially expressed lincRNAs involved in this process. The expression of lincRNAs involved in the process is negatively correlated with promoter methylation in trend. The identified lincRNAs involved in zygote first cleavage division could have important roles in mouse early embryonic development and need further functional studies. Altogether, a novel reference catalog of mouse PED lincRNAs is provided and characterized, which would be a valuable resource for further functional analyses and may help elucidate the pre-implantation regulatory mechanism.

DevMouse, the mouse developmental methylome database and analysis tools.

DNA methylation undergoes dynamic changes during mouse development and plays crucial roles in embryogenesis, cell-lineage determination and genomic imprinting. Bisulfite sequencing enables profiling of mouse developmental methylomes on an unprecedented scale; however, integrating and mining these data are challenges for experimental biologists. Therefore, we developed DevMouse, which focuses on the efficient storage of DNA methylomes in temporal order and quantitative analysis of methylation dynamics during mouse development. The latest release of DevMouse incorporates 32 normalized and temporally ordered methylomes across 15 developmental stages and related genome information. A flexible query engine is developed for acquisition of methylation profiles for genes, microRNAs, long non-coding RNAs and genomic intervals of interest across selected developmental stages. To facilitate in-depth mining of these profiles, DevMouse offers online analysis tools for the quantification of methylation variation, identification of differentially methylated genes, hierarchical clustering, gene function annotation and enrichment. Moreover, a configurable MethyBrowser is provided to view the base-resolution methylomes under a genomic context. In brief, DevMouse hosts comprehensive mouse developmental methylome data and provides online tools to explore the relationships of DNA methylation and development. Database URL: http://www.devmouse.org/

Spatiotemporal expression of retrogene-host pair Mcts2/H13 in mouse embryo, and Mcts2 has no influence on H13 transcription pattern in NIH/3T3 cells.

Mcts2 and H13 comprise an imprinted retrogene-host gene pair. Imprinted genes have been proved to be closely related with embryo development. In order to understand its expression relationship during embryo development and influence of the retrogene on the host gene, we studied expression patterns in mouse embryos and transcriptional interference in a cell culture system. The present study determined the spatio-temporal expression pattern of Mcts2 and H13 from embryonic day 9.5 to 15.5. A similar expression pattern between Mcts2 and H13 was observed in mouse embryogenesis by in situ hybridization and real-time PCR, these two genes were extensively expressed in the neural tissues at mid-embryonic stages. As the embryo development proceeded, H13 and Mcts2 were widely detected throughout the developing organism, especially highly expressed in brain. Moreover, neither over expression nor knockdown of Mcts2 has any significant detectable effect on H13 expression in NIH/3T3 cells. In addition, transcriptional up-regulation of Mcts2 caused by demethylation of DMR in the Mcts2 promoter was not directly associated with the H13 transcription in NIH/3T3 cells treated by 5-Aza-cdR. The regulatory relationship between H13 transcripts and the promoter methylation status of Mcts2 was complex, demonstrating host/retrogene relationship may not be limited to the imprinted locus.

Effect of thoracic epidural blockade on hypoxia-induced pulmonary arterial hypertension in rats.

OBJECTIVES: The present study was aimed to investigate the influence of thoracic epidural blockade on hypoxia-induced pulmonary hypertension in rats. MATERIALS AND METHODS: Forty eight Wistar rats were randomly divided into 4 equal groups, named normoxia hypoxia hypoxia/ ropivacaine and hypoxia/saline. Animals were placed in a hypoxia chamber and instrumented with epidural catheters at the thoracic level. Rats were injected with saline or ropivacaine. Haemodynamic measurements included pulmonary artery pressure and right ventricular hypertrophy. Degree of pulmonary vascular remodeling was determined by Hematoxylin and Eosin (HE) staining. Serum cyclic GMP (cGMP) and TNF-α were measured using radioimmuno assay. Real-time PCR and western boltting were employed to examine the expression of cAMP responding-element binding protein (CREB). RESULTS: We found that the thoracic epidural blockade significantly decreased chronic hypoxia-induced pulmonary hypertension and vascular remodeling in rats. Ropivacaine-treated rats exhibited significantly lower mean pulmonary artery pressure (mPAP), ratio of right ventricular weight to left ventricular plus septal weight (RV/(LV+S)) and wall thickness of pulmonary artery compared with those of control rats. Hypoxia-induced increase in levels of serum cGMP and TNF-α was reversed by thoracic epidural blockade. Moreover, hypoxia increased expression of CREB at mRNA and protein levels which could be suppressed by thoracic epidural blockade. CONCLUSION: Thoracic epidural blockade reduced mPAP and serum level of TNF-α and increased cGMP. The treatment reversed upregulated expression of CREB at mRNA and protein production.