Identification and Validation of Hub Genes for Predicting Treatment Targets and Immune Landscape in Rheumatoid Arthritis.

Background: Rheumatoid arthritis (RA) is recognized as a chronic inflammatory disease featured by pathological synovial inflammation. Currently, the underlying pathophysiological mechanisms of RA remain unclear. In the study, we attempted to explore the underlying mechanisms of RA and provide potential targets for the therapy of RA via bioinformatics analysis. Methods: We downloaded four microarray datasets (GSE77298, GSE55235, GSE12021, and GSE55457) from the GEO database. Firstly, GSE77298 and GSE55457 were identified DEGs by the "limma" and "sva" packages of R software. Then, we performed GO, KEGG, and GSEA enrichment analyses to further analyze the function of DEGs. Hub genes were screened using LASSO analysis and SVM-RFE analysis. To further explore the differences of the expression of hub genes in healthy control and RA patient synovial tissues, we calculated the ROC curves and AUC. The expression levels of hub genes were verified in synovial tissues of normal and RA rats by qRT-PCR and western blot. Furthermore, the CIBERSORTx was implemented to assess the differences of infiltration in 22 immune cells between normal and RA synovial tissues. We explored the association between hub genes and infiltrating immune cells. Results: CRTAM, CXCL13, and LRRC15 were identified as RA's potential hub genes by machine learning and LASSO algorithms. In addition, we verified the expression levels of three hub genes in the synovial tissue of normal and RA rats by PCR and western blot. Moreover, immune cell infiltration analysis showed that plasma cells, T follicular helper cells, M0 macrophages, M1 macrophages, and gamma delta T cells may be engaged in the development and progression of RA. Conclusions: In brief, our study identified and validated that three hub genes CRTAM, CXCL13, and LRRC15 might involve in the pathological development of RA, which could provide novel perspectives for the diagnosis and treatment with RA.

Effect of chrono-moxibustion and its influence on circadian rhythm for rheumatoid arthritis: A study protocol for a randomized controlled trial.

BACKGROUND: Rheumatoid arthritis (RA) is a chronic disease which is characterized by a circadian variation of key clinical symptoms and findings, with prominent joint swelling, stiffness and pain occurring in the early morning and light clinical symptoms during the day. Chrono-moxibustion is carried out at different time, which could result in dissimilar therapeutic effects. However, its efficacy has seldom been systematically demonstrated and few studies have reported that Chrono-moxibustion may regulate the circadian rhythm of RA. We therefore designed a randomized trial to explore the effective difference of Chrono-moxibustion in RA treatment, as well as to study its influence on circadian rhythm of RA patients. METHODS: This study is a randomized controlled trial involving 120 participants, and a total of 90 eligible RA patients will be randomly allocated to three groups in a 1:1:1 ratio as moxibustion at 7 to 9 am, moxibustion at 5 to 7 pm, and waiting list group, meanwhile, 30 healthy people will be divided into the control group. Patients in moxibustion groups will be treated for 30 minutes per session, 3 times a week, lasting 6 weeks. All of RA patients will be evaluated with questionnaires and laboratory tests before treatment, as well as 3 weeks, 6 weeks, and 3 months after treatment. One way analysis of variance (ANOVA) with multiple comparisons will be applied to identify differences more than two groups. Halberg cosiner software will be used to analysis the circadian rhythm. RESULTS: The results of this study will be published in a peer-reviewed journal. CONCLUSION: This study will provide evidence-based evidence for the effective difference of Chrono-moxibustion in RA treatment and its influence on circadian rhythm of RA patients.

Identification and validation of potential hub genes in rheumatoid arthritis by bioinformatics analysis.

OBJECTIVE: Rheumatoid arthritis (RA) is considered to be a chronic immune disease pathologically characterized by synovial inflammation and bone destruction. At present, the potential pathogenesis of RA is still unclear. Hub genes are recognized to play a pivotal role in the occurrence and progression of RA. METHODS: Firstly, we attempted to screen hub genes that are associated with RA, to clarify the underlying pathological mechanisms of RA, and to offer potential treatment methods for RA. We acquired these datasets (GSE12021, GSE55235, and GSE55457) of RA patients and healthy samples from the Gene Expression Omnibus (GEO) database. Differentially expressed genes (DEGs) were recognized via R software. Then, Gene ontology (GO) functional analysis and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment analysis were utilized to deeply explore the underlying biological functions and pathways closely associated with RA. In addition, a protein-protein interaction (PPI) network was built to further evaluate and screen for hub genes. Finally, on the basis of the results of PPI analysis, we confirmed the mRNA expression levels of five hub genes in the synovial tissue of rats modeled with RA. RESULTS: In the human microarray datasets, LCK, JAK2, SOCS3, STAT1, and EGFR were identified as hub genes associated with RA by bioinformatics analysis. Furthermore, we verified the differential expression levels of hub genes in rat synovial tissues via qRT-PCR (P < 0.05). CONCLUSIONS: Our findings suggest that the hub genes LCK, JAK2, SOCS3, STAT1, and EGFR might have vital roles in the progression of RA and may offer novel therapeutic treatments for RA.

Chrono-moxibustion adjusts circadian rhythm of CLOCK and BMAL1 in adjuvant-induced arthritic rats.

OBJECTIVE: The clinical symptoms of rheumatoid arthritis (RA) have significant circadian rhythms, with morning stiffness and joint pain. Moxibustion is effective in the treatment of RA, while the underlying therapeutic mechanisms remain limited. Thus, we explored whether moxibustion could adjust the circadian rhythm of RA by modulating the core clock genes CLOCK and BMAL1 at the molecular level. METHODS: 144 Sprague Dawley rats were randomly divided into four groups: control group (group A), model group (group B), 7-9 am moxibustion treatment group (group C), and 5-7 pm moxibustion treatment group (group D). Each group was divided into 6 time points (0 am, 4 am, 8 am, 12 N, 6 pm, and 8 pm) with an equal number of rats at each time point. Except for group A, all rats were injected with Freund's Complete Adjuvant (FCA) 0.15 ml on the right foot pad to establish the RA model. The rats of the two moxibustion treatment groups were respectively subjected to moxibustion at 7-9 am and 5-7 pm. After 3 weeks of treatment, the tissues were collected at 6 time points during the next 24 hours. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) was used to test the mRNA expression of CLOCK and BMAL1 in the hypothalamus and synovial tissues. CLOCK and BMAL1 protein expression in synovial tissues were detected with western blot. RESULTS: Compared to group A, group B showed significantly down-regulated expression levels of CLOCK and BMLA1 at synovial tissue (P < 0.05), while no statistically significant difference was found in the hypothalamus (P > 0.05). The expression levels of CLOCK and BMLA1 were up-regulated in the moxibustion treatment groups in different tissues, especially in synovial tissue (P < 0.05) compared to group B. Nevertheless, no difference was observed between groups C and D (P > 0.05). CONCLUSIONS: Moxibustion could treat RA by modulating clock core genes CLOCK and BMAL1 to regulate the circadian rhythm. However, there was no significant difference between the 7-9 am moxibustion treatment group and the 5-7 pm moxibustion treatment group. This study provides a basis for research on moxibustion in the treatment of RA.

MiR-221/222 Ameliorates Deoxynivalenol-Induced Apoptosis and Proliferation Inhibition in Intestinal Epithelial Cells by Targeting PTEN.

Intestinal epithelial cells are critical for nutrient absorption and defending against pathogen infection. Deoxynivalenol (Don), the most common mycotoxin, contaminates cereals and food throughout the world, causes serious damage to mammal intestinal mucosa, and appears as intestinal epithelial cell apoptosis and proliferation inhibition. Our previous study has found that milk-derived exosome ameliorates Don-induced intestinal damage, but the mechanism is still not fully understood. In this study, we demonstrated that Don downregulated the expression of miR-221/222 in intestinal epithelial cells, and exosome treatment reversed the inhibitory effect of Don on miR-221/222. Through immunofluorescence and flow cytometry analysis, we identified that miR-221/222 ameliorates Don-induced apoptosis and proliferation inhibition in intestinal epithelial cells. Through bioinformatics analyses and RNA immunoprecipitation analysis, we identified Phosphatase and tensin homolog (PTEN) is the target of miR-221/222. Through the PTEN interfering experiment, we found Don-induced apoptosis and proliferation inhibition relied on PTEN. Finally, through adenovirus to overexpress miR-221/222 in mice intestinal epithelial cells specifically, our results showed that miR-221/222 ameliorated Don-induced apoptosis and proliferation inhibition in intestinal epithelial cells by targeting PTEN. This study not only expands our understanding of how miR-221/222 and the host gene PTEN regulate intestinal epithelial cells defending against Don-induced damage, but also provides a new way to protect the development of the intestine.

Linking the low-density lipoprotein receptor-binding segment enables the therapeutic 5-YHEDA peptide to cross the blood-brain barrier and scavenge excess iron and radicals in the brain of senescent mice.

INTRODUCTION: Iron accumulates in the brain during aging, which catalyzes radical formation, causing neuronal impairment, and is thus considered a pathogenic factor in Alzheimer's disease (AD). To scavenge excess iron-catalyzed radicals and thereby protect the brain and decrease the incidence of AD, we synthesized a soluble pro-iron 5-YHEDA peptide. However, the blood-brain barrier (BBB) blocks large drug molecules from entering the brain and thus strongly reduces their therapeutic effects. However, alternative receptor- or transporter-mediated approaches are possible. METHODS: A low-density lipoprotein receptor (LDLR)-binding segment of Apolipoprotein B-100 was linked to the 5-YHEDA peptide (bs-5-YHEDA) and intracardially injected into senescent (SN) mice that displayed symptoms of cognitive impairment similar to those of people with AD. RESULTS: We successfully delivered 5-YHEDA across the BBB into the brains of the SN mice via vascular epithelium LDLR-mediated endocytosis. The data showed that excess brain iron and radical-induced neuronal necrosis were reduced after the bs-5-YHEDA treatment, together with cognitive amelioration in the SN mouse, and that the senescence-associated ferritin and transferrin increase, anemia and inflammation reversed without kidney or liver injury. DISCUSSION: bs-5-YHEDA may be a mild and safe iron remover that can cross the BBB and enter the brain to relieve excessive iron- and radical-induced cognitive disorders.

[The application of variable theoretical alpha influence coefficent to the matrix correction in Cr-Fe-Ni stainless steel].

X-ray fluorescence analysis software Spectra Plus was used to calculate theoretical alpha influence coefficents of other elements to Cr in seven stainless steel standard samples, theoretical alpha influence coefficients of elements, by which Cr signal was enhanced, varied largely with the change of elements content. Variable theoretical alpha influence coefficients, which varied with elements content, were used to correct the matrix effects in stainless steel, the secondary excitation of Cr by other elements were corrected, and Cr (0.3%-20.8%) in stainless steel and low alloy steel could be analysed in accordance with one calibration curve. The matrix effects in samples can be corrected by variable theoretical alpha influence coefficients, so the measurable content range of calibration curve was enlarged. The contents of fifteen elements Al, Si, P, S, Ti, Cr, Mn, Co, Ni, Cu, As, Mo, Sn, W and Pb in stainless steel were measured by X-ray fluorescence spectrometer, variable theoretical alpha influence coefficients were used to correct the matrix effects, and the analysis results are comparable to those obtained by wet chemical method.