Gut microbiota regulation of T lymphocyte subsets during systemic lupus erythematosus.

BACKGROUND: Systemic lupus erythematosus (SLE) is an autoimmune disease characterized by disturbance of pro-inflammatory and anti-inflammatory lymphocytes. Growing evidence shown that gut microbiota participated in the occurrence and development of SLE by affecting the differentiation and function of intestinal immune cells. The purpose of this study was to investigate the changes of gut microbiota in SLE and judge its associations with peripheral T lymphocytes. METHODS: A total of 19 SLE patients and 16 HCs were enrolled in this study. Flow cytometry was used to detect the number of peripheral T lymphocyte subsets, and 16 s rRNA was used to detect the relative abundance of gut microbiota. Analyzed the correlation between gut microbiota with SLEDAI, ESR, ds-DNA and complement. SPSS26.0 software was used to analyze the experimental data. Mann-Whitney U test was applied to compare T lymphocyte subsets. Spearman analysis was used for calculating correlation. RESULTS: Compared with HCs, the proportions of Tregs (P = 0.001), Tfh cells (P = 0.018) and Naïve CD4 + T cells (P = 0.004) significantly decreased in SLE patients, and proportions of Th17 cells (P = 0.020) and γδT cells (P = 0.018) increased in SLE. The diversity of SLE patients were significantly decreased. Addition, there were 11 species of flora were discovered to be distinctly different in SLE group (P < 0.05). In the correlation analysis of SLE, Tregs were positively correlated with Ruminococcus2 (P = 0.042), Th17 cells were positively correlated with Megamonas (P = 0.009), γδT cells were positively correlated with Megamonas (P = 0.003) and Streptococcus (P = 0.004), Tfh cells were positively correlated with Bacteroides (P = 0.040), and Th1 cells were negatively correlated with Bifidobacterium (P = 0.005). As for clinical indicators, the level of Tregs was negatively correlated with ESR (P = 0.031), but not with C3 and C4, and the remaining cells were not significantly correlated with ESR, C3 and C4. CONCLUSION: Gut microbiota and T lymphocyte subsets of SLE changed and related to each other, which may break the immune balance and affect the occurrence and development of SLE. Therefore, it is necessary to pay attention to the changes of gut microbiota and provide new ideas for the treatment of SLE.

Identifying and Exploring the Impact Factors for Intraocular Pressure Prediction in Myopic Children with Atropine Control Utilizing Multivariate Adaptive Regression Splines.

PURPOSE: The treatment of childhood myopia often involves the use of topical atropine, which has been demonstrated to be effective in decelerating the progression of myopia. It is crucial to monitor intraocular pressure (IOP) to ensure the safety of topical atropine. This study aims to identify the optimal machine learning IOP-monitoring module and establish a precise baseline IOP as a clinical safety reference for atropine medication. METHODS: Data from 1545 eyes of 1171 children receiving atropine for myopia were retrospectively analyzed. Nineteen variables including patient demographics, medical history, refractive error, and IOP measurements were considered. The data were analyzed using a multivariate adaptive regression spline (MARS) model to analyze the impact of different factors on the End IOP. RESULTS: The MARS model identified age, baseline IOP, End Spherical, duration of previous atropine treatment, and duration of current atropine treatment as the five most significant factors influencing the End IOP. The outcomes revealed that the baseline IOP had the most significant effect on final IOP, exhibiting a notable knot at 14 mmHg. When the baseline IOP was equal to or exceeded 14 mmHg, there was a positive correlation between atropine use and End IOP, suggesting that atropine may increase the End IOP in children with a baseline IOP greater than 14 mmHg. CONCLUSIONS: MARS model demonstrates a better ability to capture nonlinearity than classic multiple linear regression for predicting End IOP. It is crucial to acknowledge that administrating atropine may elevate intraocular pressure when the baseline IOP exceeds 14 mmHg. These findings offer valuable insights into factors affecting IOP in children undergoing atropine treatment for myopia, enabling clinicians to make informed decisions regarding treatment options.

Study on the expression changes of lncRNA in patients with systemic lupus erythematosus and its correlation with Treg cells.

OBJECTIVES: We initially explored the link between the differentially expressed long non-coding RNAs (lncRNAs) and the number of regulatory T (Treg) cells by detecting the lncRNA expression profiles in patients with systemic lupus erythematosus (SLE), then analyzed the correlation between Treg-related lncRNAs and the clinical features of SLE patients, predicting the mechanism by which lncRNAs regulate the differentiation and development of Treg cells, and provided new ideas for the treatment of SLE. METHODS: Peripheral blood of 9 active SLE patients were collected and mononuclear cells (PBMCs) were extracted; the lncRNA expression profiles of PBMCs were analyzed by whole transcriptome sequencing. Nine healthy people were used as controls to screen the differentially expressed lncRNAs, to analyze the correlation between lncRNAs and Treg cell number. Pearson test was used to analyze the correlation between lncRNAs and the number of Treg cell, and the correlation between Treg-associated lncRNA and SLEDAI score, ESR, C3, and C4 in SLE patients. The targeted genes of Treg-associated lncRNAs were predicted with miRcode and Targetscan databases and coexpression network. RESULTS: There were 240 differentially expressed lncRNAs in SLE patients compared with healthy controls, including 134 highly expressed lncRNAs (p < 0.05) and 106 lowly expressed lncRNAs (p < 0.05). The expression of ANKRD44-AS1 (r = 0.7417, p = 0.0222), LINC00200 (r = 0.6960, p = 0.0373), AP001363.2 (r = 0.7766, p = 0.0138), and LINC02824 (r = 0.7893, p = 0.0114) were positively correlated with the number of Treg cell, and the expression of AP000640.1 (r = - 0.7225, p = 0.0279), AC124248.1 (r = - 0.7653, p = 0.0163), LINC00482 (r = - 0.8317, p = 0.0054), and MIR503HG (r = - 0.7617, p < 0.05) were negatively correlated with the number of Treg cell. Among these Treg-associated lncRNAs, the expression of LINC00482 (r = - 0.7348, p < 0.05) and MIR503 HG (r = - 0.7617, p < 0.05) were negatively correlated with C3. LINC00200, ANKRD44 - AS1, and AP000640.1 related to Treg cells regulate the expression of signal transducer and activator of transcription 5 (STAT5), phospholipase D1 (PLD1), homeodomain-only protein X (HOPX), and runt-related transcription factor 3 (RUNX3) through competitive binding of miRNA or trans-regulatory mechanism, thereby regulating the differentiation and development of Treg cell. CONCLUSIONS: The lncRNA expression profiles were changed in SLE patients, the differentially expressed lncRNAs were associated with abnormal number and function of Treg cells in SLE, and Treg-associated lncRNAs were associated with SLE-disease activity, which may affect the expression of STAT5, PLD1, HOPX, RUNX3 and regulate Treg cell function and participate in the pathogenesis and progression of SLE by competitively binding to miRNAs or trans-regulatory mechanism. Key points • Systemic lupus erythematosus (SLE) is an autoimmune disease involving multiple organs and systems. lncRNAs may affect Treg cells function by regulating genes expression, which may be an important pathogenesis of SLE. • This study, taking SLE as an example, preliminarily analyzed the correlation between lncRNA and Treg cells in SLE patients, analyzed the correlation between Treg-related lncRNA and the clinical characteristics of SLE, and speculated that lncRNA could regulate the differentiation and development of Treg cells through competitive combination with miRNA or trans-regulatory mechanisms. • It is possible to target epigenetic therapy for SLE.

Co-augmentation of a transport gene mfsT1 in Mycolicibacterium neoaurum with genome engineering to enhance ergothioneine production.

Ergothioneine (EGT) is a rare thiohistidine derivative with exceptional antioxidant properties. The blood level of EGT is considered highly reliable predictors for cardiovascular diseases and mortality, yet animals lack the ability to synthesize this compound. Free plasmids have been previously used to overexpress genes involved in the EGT biosynthetic pathway of Mycolicibacterium neoaurum. Here, we tentatively introduced a putative transporter gene mfsT1 into high-copy plasmids and sharply increased the ratio of extracellular EGT concentration from 18.7% to 44.9%. Subsequently, an additional copy of egtABCDE, hisG, and mfsT1 was inserted into the genome with a site-specific genomic integration tool of M. neoaurum, leading a 2.7 times increase in EGT production. Co-enhancing the S-adenosyl-L-methionine regeneration pathway, or alternatively, the integration of three copies of egtABCDE, hisG and mfsT1 into the genome further increased the total EGT yield by 16.1% (64.6 mg/L) and 21.7% (67.7 mg/L), respectively. After 168-h cultivation, the highest titer reached 85.9 mg/L in the latter strain with three inserted copies. This study provided a solid foundation for genome engineering to increase the production of EGT in M. neoaurum.

Clinical efficacy of plasma exchange in systemic lupus erythematosus during pregnancy.

OBJECTIVE: To investigate the clinical efficacy of plasma exchange (PE) with or without prednisone and hydroxychloroquine (HCQ) for the treatment of systemic lupus erythematosus (SLE) during pregnancy. METHODS: The clinical characteristics of 14 pregnant women with SLE admitted to our hospital were retrospectively analyzed, including 7 only treated with prednisone and HCQ (non-PE group) as well as 7 combined PE (PE group). The delivery situations of 14 patients were recorded. Data like erythrocyte sedimentation rate (ESR), urine protein, platelet count, and SLEDAI scores were compared between two groups before treatment and 3, 6, and 12 months after delivery. RESULTS: Three patients in the non-PE group ended in miscarriage while all patients in the PE group were delivered successfully. Eleven successfully delivered fetuses in the two groups were healthy, and the Apgar scores were over 8. The ESR of the PE group was significantly lower than that of the non-PE group at 6 and 12 months after delivery, while there was no statistical difference in ESR between the two groups before treatment and 3 months after delivery. The ESR and urine protein were significantly higher in the non-PE group at months 3, 6, and 12 postpartum. There was a significant decrease in disease activity postpartum in the PE group compared to predelivery disease activity. The change in platelet counts between the two groups significantly increased over time in the PE group, while SLEDAI scores decreased. CONCLUSIONS: The combination of PE and oral prednisone and HCQ is possibly a more effective treatment than oral prednisone and HCQ alone for patients with active SLE during pregnancy. This treatment option reduces pregnancy loss and promotes the patients' postpartum condition to a certain extent.

Addition of transarterial chemoembolization improves outcome of tyrosine kinase and immune checkpoint inhibitors regime in patients with unresectable hepatocellular carcinoma.

Background: Transarterial chemoembolization (TACE) is the standard treatment for hepatocellular carcinoma (HCC); the value of its combination with systemic therapy is worthy of further exploration. This study aimed to investigate the efficacy and safety of TACE combined with tyrosine kinase inhibitor (TKI) and immune checkpoint inhibitor (ICI) in the treatment of unresectable HCC. Methods: In this retrospective observational, single-center study, 147 patients with unresectable HCC were divided into a TACE group (n=98) and a non-TACE group (n=49) based on whether TACE was performed during TKI plus ICI therapy. The survival outcomes and adverse events (AEs) of the two groups were compared. Results: Data from patients with unresectable HCC who received TKI plus ICI treatment between July 2017 and April 2020 were collected. The median intrahepatic tumor size was 8.7 cm [interquartile range (IQR), 5.9-12.4 cm]. At data cut-off, overall survival (OS) of the TACE group was significantly longer than that of the non-TACE group (19.5 and 10.8 months, respectively, P=0.005). In the high-risk cohort (with main or contralateral portal vein tumor thrombi and/or bile duct invasion and/or a tumor burden >50% of liver), the OS of the TACE group was still longer than that of the non-TACE group (14.9 and 8.7 months, respectively, P=0.031). Major AEs were tolerated in both groups, and there was no significant difference in their incidence (34.7% and 30.6%, respectively, P=0.621). Conclusions: TACE treatment combined with TKI plus ICI regime resulted in longer OS than treatment with TKI plus ICI alone for patients with unresectable HCC.

Inflammation-related nomogram for predicting survival of patients with unresectable hepatocellular carcinoma received conversion therapy.

BACKGROUND: The efficacy of conversion therapy for patients with unresectable hepatocellular carcinoma (HCC) is a common clinical concern. AIM: To analyse the prognostic factors of overall survival (OS) in patients with unresectable HCC who received conversion therapy. METHODS: One hundred and fifty patients who met the inclusion criteria were enrolled and divided into a training cohort (n = 120) and a validation cohort (n = 30). Using the independent risk factors in the training cohort, a nomogram model was constructed to predict OS for patients treated with transarterial chemoembolization following hepatic resection. The nomogram was internally validated with the bootstrapping method. The predictive performance of nomogram was assessed by Harrell's concordance index (C-index), calibration plot and time-dependent receiver operating characteristic curves and compared with six other conventional HCC staging systems. RESULTS: Multivariate Cox analysis identified that albumin, blood urea nitrogen, gamma-glutamyl transpeptidase to platelet ratio, platelet to lymphocyte ratio, macrovascular invasion and tumour number were the six independent prognostic factors correlated with OS in nomogram model. The C-index in the training cohort and validation cohort were 0.752 and 0.807 for predicting OS, which were higher than those of the six conventional HCC staging systems (0.563 to 0.715 for the training cohort and 0.458 to 0.571 for the validation cohort). The calibration plots showed good consistency between the nomogram prediction of OS and the actual observations of OS. Decision curve analyses indicated satisfactory clinical utility. With a total nomogram score of 196, patients were accurately classified into low-risk and high-risk groups. Furthermore, we have deployed the model into online calculators that can be accessed for free at https://ctmodelforunresectablehcc.shinyapps.io/DynNomapp/. CONCLUSION: The nomogram achieved optimal individualized prognostication of OS in HCC patients who received conversion therapy, which could be a useful clinical tool to help guide postoperative personalized interventions and prognosis judgement.

Sorbitol induces flower bud formation via the MADS-box transcription factor EjCAL in loquat.

Sorbitol is an important signaling molecule in fruit trees. Here, we observed that sorbitol increased during flower bud differentiation (FBD) in loquat (Eriobotrya japonica Lindl.). Transcriptomic analysis suggested that bud formation was associated with the expression of the MADS-box transcription factor (TF) family gene, EjCAL. RNA fluorescence in situ hybridization showed that EjCAL was enriched in flower primordia but hardly detected in the shoot apical meristem. Heterologous expression of EjCAL in Nicotiana benthamiana plants resulted in early FBD. Yeast-one-hybrid analysis identified the ERF12 TF as a binding partner of the EjCAL promoter. Chromatin immunoprecipitation-PCR confirmed that EjERF12 binds to the EjCAL promoter, and ß-glucuronidase activity assays indicated that EjERF12 regulates EjCAL expression. Spraying loquat trees with sorbitol promoted flower bud formation and was associated with increased expression of EjERF12 and EjCAL. Furthermore, we identified EjUF3GaT1 as a target gene of EjCAL and its expression was activated by EjCAL. Function characterization via overexpression and RNAi reveals that EjUF3GaT1 is a biosynthetic gene of flavonoid hyperoside. The concentration of the flavonoid hyperoside mirrored that of sorbitol during FBD and exogenous hyperoside treatment also promoted loquat bud formation. We identified a mechanism whereby EjCAL might regulate hyperoside biosynthesis and confirmed the involvement of EjCAL in flower bud formation in planta. Together, these results provide insight into bud formation in loquat and may be used in efforts to increase yield.

A decreased number of circulating regulatory T cells is associated with adverse pregnancy outcomes in patients with systemic lupus erythematosus.

OBJECTIVE: As an autoimmune disease affecting women of reproductive age, systemic lupus erythematosus (SLE) is linked to adverse fetal and maternal outcomes. However, the status of peripheral lymphocytes in SLE patients with different pregnancy outcomes is unclear. This retrospective cross-sectional study explored the relationship between lymphocyte subpopulations and pregnancy outcomes in married SLE female patients. METHODS: The absolute numbers of peripheral T, helper T (Th)1, Th2, Th17, regulatory T (Treg), B, and natural killer (NK) cell subpopulations from 585 female SLE patients and 91 female healthy controls (HCs) were assessed. We compared the lymphocyte subpopulations in SLE patients with HCs and analyzed the absolute number and ratio of Treg cells according to pregnancy outcome in SLE patients. RESULTS: SLE patients had decreased numbers of T, B, NK, Th1, Th2, Th17, and Treg cells and an imbalance in pro- and anti-inflammatory cells (p < .05), as well as adverse pregnancy outcomes. In abortion patients, the number of Treg cells (p = .008) decreased, leading to an imbalance in effector T and Treg cells. The ratio of Treg cells was higher in SLE patients with nulliparity than in those with one or two parities. CONCLUSIONS: The absolute numbers of lymphocyte subpopulations in SLE patients decreased, which was associated with abortion and parity (p < .05). These results suggest that a loss of immune tolerance mediated by Tregs triggers pregnancy loss.

Construction of an Antibiotic-Free Vector and its Application in the Metabolic Engineering of Escherichia Coli for Polyhydroxybutyrate Production.

An antibiotic- and inducer-free culture condition was proposed for polyhydroxybutyrate (PHB) production in recombinant Escherichia coli. First, antibiotic-free vectors were constructed by installing the plasmid maintenance system, alp7, hok/sok, and the hok/sok and alp7 combination into the pUC19 vector. The plasmid stability test showed that pVEC02, the pUC19 vector containing the hok/sok system, was the most effective in achieving antibiotic-free cultivation in the E. coli B strain but not in the K strain. Second, the putative phaCAB operon derived from Caldimonas manganoxidans was inserted into pVEC02 to yield pPHB01 for PHB production in E. coli BL21 (DE3). The putative phaCAB operon was first shown function properly for PHB production and thus, inducer-free conditions were achieved. However, the maintenance of pPHB01 in E. coli requires antibiotics supplementation. Finally, an efficient E. coli ρ factor-independent terminator, thrLABC (ECK120033737), was inserted between the phaCAB operon and the hok/sok system to avoid possible transcriptional carry-over. The newly constructed plasmid pPHB01-1 facilitates an antibiotic- and inducer-free culture condition and induces the production of PHB with a concentration of 3.0 on0.2 g/L, yield of 0.26 /L0.07 g/g-glucose, and content of 44 /g3%. The PHB production using E. coli BL21 (DE3)/pPHB01-1 has been shown to last 84 and 96 h in the liquid and solid cultures.

Identification of circRNAs involved in the development of hepatocellular carcinoma after insufficient radiofrequency ablation.

Previous studies have reported that circular RNAs (circRNAs) play a key role in the pathogenesis and progression of various diseases. In the present study, we aimed to identify potential circRNAs associated with the progression of hepatocellular carcinoma (HCC) after insufficient radiofrequency ablation (IRFA). A xenograft mouse IRFA model was initially established, and immunohistochemical staining (IHC) and polymerase chain reaction (PCR) were performed to confirm the expression of programmed cell death-ligand 1 (PD-L1) and vascular endothelial growth factor receptor-1 (VEGFR-1). CircRNA expression alterations were screened by next-generation sequencing (RNA-seq). Gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analyses were conducted to predict the function of genes coding differentially expressed circRNAs. The selected circRNAs were validated utilizing PCR and Sanger sequencing. The relationships between circRNAs, microRNAs, PD-L1, and VEGFR-1 were predicted by bioinformatics. Overall, a total of 612 circRNAs were differentially expressed in IRFA-treated subcutaneous tumorigenesis tissue. Among them, 435 circRNAs were significantly upregulated and 177 circRNAs were downregulated. GO and KEGG analyses were employed to predict the functions of these circRNAs. Thereafter, quantitative reverse transcription PCR (qRT-PCR) assays determined that these seven circRNAs were overexpressed in the IRFA group, which was consistent with the RNA-seq results. Based on the bioinformatic analysis, seven circRNAs confirmed by Sanger sequencing were predicted to likely regulate PD-L1 and VEGFR-1 expression levels by functioning as sponges for microRNAs (miRNAs) and forming a circRNA-miRNA-PD-L1/VEGFR-1 regulatory network. Finally, IHC and qRT-PCR of PD-L1 and VEGFR-1 confirmed the activation of this pathway. Taken together, we report that differentially expressed circRNAs might simultaneously regulate PD-L1 and VEGFR-1 in the IRFA tissues, which provides a novel view of circRNAs in HCC progression after the IRFA procedure.

DNA N6-methyladenine involvement and regulation of hepatocellular carcinoma development.

DNA N6-methyladenine (6 mA) is a new type of DNA methylation identified in various eukaryotic cells. However, its alteration and genomic distribution features in hepatocellular carcinoma (HCC) remain elusive. In this study, we found that N6AMT1 overexpression increased HCC cell viability, suppressed apoptosis, and enhanced migration and invasion, whereas ALKBH1 overexpression induced the opposite effects. Further, 23,779 gain-of-6 mA regions and 11,240 loss-of-6 mA regions were differentially identified in HCC tissues. The differential gain and loss of 6 mA regions were considerably enriched in intergenic regions. Moreover, 7% of the differential 6 mA modifications were associated with tumors, with 60 associated with oncogenes and 57 with tumor suppressor genes (TSGs), and 17 were common to oncogenes and TSGs. The candidate genes affected by 6 mA were filtered by gene ontology (GO) and RNA-seq. Using quantitative polymerase chain reaction (qPCR), BCL2 and PARTICL were found to be correlated with DNA 6 mA in certain HCC processes.

Extensive cutaneous metastasis of recurrent gastric cancer: A case report.

BACKGROUND: Cutaneous metastasis is a rare event associated with poor prognosis for gastric cancer and has been rarely reported in the literature. CASE SUMMARY: A 69-year-old male patient who had undergone salvage gastrectomy and a few courses of adjuvant chemotherapy 3 mo earlier for recurrent gastric cancer developed widespread cutaneous metastases. Due to the patient's intolerance to further adjuvant chemotherapy, he was placed in hospice care and expired 1 mo later. In the literature, gastric cancers are rarely reported as the primary malignancies for cutaneous metastasis. We, thus, provide an update on a case review published in 2014 by reviewing 10 more case reports dated from 2014 to 2020. The average age for the new group of patients was 59.4 ± 18.88-years-old. Thirty percent of the patients presented with cutaneous lesions and advanced gastric cancer synchronously while 70% developed cutaneous metastases 1.3 years to 14 years after the initial treatment for primary gastric cancer. Eighty percent of the patients received either local excision or chemo ± radiation therapy to treat their cutaneous metastases. CONCLUSION: This report highlights cutaneous metastasis as a late and untreatable metastasis of gastric cancer.

[Effect of fire needling on mild to moderate knee osteoarthritis and related serum inflammatory cytokines].

OBJECTIVE: To compare the clinical effect of fire needling and filiform needling for mild to moderate knee osteoarthritis (KOA) and observe the influence on related serum inflammatory sytokines. METHODS: A total of 60 patients with mild to moderate KOA were randomly divided into an observation group (30 cases, 4 cases dropped off) and a control group (30 cases, 4 cases dropped off). Both groups were given basic health management, and the acupoints of the two groups were Liangqiu (ST 34), Xuehai (SP 10), Neixiyan (EX-LE 4), Dubi (ST 35), Yanglingquan (GB 34) and ashi point. The observation group was treated with fire needling, while the control group was treated with filiform needling. Both groups were treated once every other day, 3 times a week for 2 weeks. The Western Ontario and McMaster University osteoarthritis index (WOAMC) scores of the two groups were compared before treatment, in 1, 2 weeks of treatment and 2, 6 weeks of follow-up. ELISA method was used to detect the levels of serum interleukin 1α (IL-1α), tumor necrosis factor α (TNF-α) and matrix metalloproteinase 3 (MMP-3) before treatment and in 2 weeks of treatment. The clinical effect of the two groups was evaluated in 1, 2 weeks of treatment and 2, 6 weeks of follow-up. RESULTS: At each time point of treatment and follow-up, the pain, stiffness, difficulty of daily activities scores and WOMAC total scores of the two groups were lower than those before treatment (P<0.05); the stiffness score of the observation group was lower than that of the control group in 1 week of treatment (P<0.05); the various scores and total scores of the WOMAC scale of the observation group were lower than those of the control group in 1, 2 weeks of treatment and 2, 6 weeks of follow-up (P<0.05). In 2 weeks of treatment, the levels of serum MMP-3 and IL-1α in the observation group and IL-1α in the control group were higher than those before treatment (P<0.05). In 1 week of treatment and 2, 6 weeks of follow-up, the total effective rates of the observation group were higher than those in the control group (P<0.05). CONCLUSION: Fire needling can improve the pain, stiffness and joint dysfunction of patients with mild to moderate KOA, and increase serum MMP-3 and IL-1α levels. Its short and long term clinical effects are better than filiform needling.

Exosome-mediated miRNA delivery promotes liver cancer EMT and metastasis.

The deregulation of exosomal microRNAs (miRNAs) plays an important role in the progression of hepatocarcinogenesis. In this study, we highlight exosomes as mediators involved in modulating miRNA profiles in liver cancer cells after induction of the epithelial-mesenchymal transition (EMT) and metastasis. Initially, we induced EMT in a hepatocellular carcinoma cell (HCC) line (Hep3B) by stimulation with transforming growth factor-ß (TGF-ß) and confirmed by western blot detection of EMT markers such as vimentin and E-cadherin. Exosomes were then isolated from the cells and identified by nanoparticle tracking analysis (NTA). The isolated exosomal particles from unstimulated Hep3B cells (Hep3B exo) or TGF-ß-stimulated EMT Hep3B cells (EMT-Hep3B exo) contained higher levels of exosome marker proteins, CD63 and TSG101. After incubation with EMT-Hep3B exo, Hep3B cell proliferation increased. EMT-Hep3B exo promoted the migration and invasion of Hep3B and 7721 cells. High-throughput sequencing of miRNAs and mRNA within the exosomes showed 119 upregulated and 186 downregulated miRNAs and 156 upregulated and 166 downregulated mRNA sequences in the EMT-Hep3B exo compared with the control Hep3B exo. The most differentially expressed miRNAs and target mRNA sequences were validated by RT-qPCR. Based on the known miRNA targets for specific mRNA sequences, we hypothesized that GADD45A was regulated by miR-374a-5p. Inhibition of miR-374a-5p in Hep3B cells resulted in exosomes that inhibited the proliferation, migration, and invasion of HCC cells. These results enhance our understanding of metastatic progression of liver cancer and provide a foundation for the future development of potential biomarkers for diagnosis and prognosis of hepatic cancer.

Fire acupuncture for mild to moderate knee osteoarthritis: a protocol for a randomized controlled pilot trial.

BACKGROUND: Knee osteoarthritis (KOA) is one of the most common bone and joint diseases. As one of the main non-drug therapies, acupuncture is widely used to treat KOA, although the evidence for its efficacy is inconclusive. The objective of this pilot trial is to clarify the clinical efficacy and safety of fire acupuncture in the treatment of mild to moderate KOA and to provide high-quality data for further research. METHODS/DESIGN: This study is a prospective randomized controlled pilot trial in which 120 patients with mild to moderate KOA will be randomly allocated in equal proportions to a fire acupuncture group or a general acupuncture group. They will receive acupuncture for six sessions over 2 weeks. The primary end point is success rate, which will be calculated based on the change from baseline of the pain and function scores in the Western Ontario and McMaster Universities Osteoarthritis Index at 4 weeks. Secondary end points include the proportion of patients achieving clinical improvement based on: (1) the OMERACT-OARSI responder criteria, (2) levels of matrix metalloproteinase 3, interleukin 1ß, and tumor necrosis factor α in blood, and (3) a subjective efficacy evaluation from patients. TRIAL REGISTRATION: Chinese Clinical Trial Registry, ChiCTR1800019162. Registered on 29 October 2018.

Effects of Virtual Reality and Augmented Reality on Induced Anxiety.

To explore the effects of virtual reality (VR) and augmented reality (AR) in the treatment of claustrophobia, the potential effects of VR and AR on induced anxiety were investigated in this paper. During the experiment, 34 subjects were randomly selected and distributed in AR and VR scenes in a sequence. The skin conductance and heart rates of the subjects were measured throughout the entire process, and the anxiety scale was used to assess the subjective anxiety when the task in each scene was completed. The results showed the following: (1) AR and VR scenes led to feelings of discomfort, but the subjective anxiety scores obtained in the two scenes were not significantly different; (2) the skin conductance level of the subjects significantly increased from the baseline when the subjects entered the experimental scene but remained active in the two scenes without showing significant difference between the scenes; and (3) the heart rate index significantly increased from the baseline after the subjects entered the scene and then gradually decreased. The heart rates of the subjects significantly increased again when the anxiety-induced event was triggered. However, no significant difference was observed between AR and VR scenes. AR and VR have induced obvious anxiety, which was reflected in the subjective and objective physiological indicators. However, no significant difference was found in the effects of AR and VR on the induced anxiety. Considering the cost of building two scenes and other factors, AR was more suitable for the treatment of claustrophobia than VR.

Fatal swine acute diarrhoea syndrome caused by an HKU2-related coronavirus of bat origin.

Cross-species transmission of viruses from wildlife animal reservoirs poses a marked threat to human and animal health 1 . Bats have been recognized as one of the most important reservoirs for emerging viruses and the transmission of a coronavirus that originated in bats to humans via intermediate hosts was responsible for the high-impact emerging zoonosis, severe acute respiratory syndrome (SARS) 2-10 . Here we provide virological, epidemiological, evolutionary and experimental evidence that a novel HKU2-related bat coronavirus, swine acute diarrhoea syndrome coronavirus (SADS-CoV), is the aetiological agent that was responsible for a large-scale outbreak of fatal disease in pigs in China that has caused the death of 24,693 piglets across four farms. Notably, the outbreak began in Guangdong province in the vicinity of the origin of the SARS pandemic. Furthermore, we identified SADS-related CoVs with 96-98% sequence identity in 9.8% (58 out of 591) of anal swabs collected from bats in Guangdong province during 2013-2016, predominantly in horseshoe bats (Rhinolophus spp.) that are known reservoirs of SARS-related CoVs. We found that there were striking similarities between the SADS and SARS outbreaks in geographical, temporal, ecological and aetiological settings. This study highlights the importance of identifying coronavirus diversity and distribution in bats to mitigate future outbreaks that could threaten livestock, public health and economic growth.