The present study investigated the mechanism of artesunate in the treatment of bone destruction in experimental rheumatoid arthritis(RA) based on transcriptomics and network pharmacology. The transcriptome sequencing data of artesunate in the inhibition of osteoclast differentiation were analyzed to obtain differentially expressed genes(DEGs). GraphPad Prism 8 software was used to plot volcano maps and heat maps were plotted through the website of bioinformatics. GeneCards and OMIM were used to collect information on key targets of bone destruction in RA. The DEGs of artesunate in inhibiting osteoclast differentiation and key target genes of bone destruction in RA were intersected by the Venny 2.1.0 platform, and the intersection target genes were analyzed by Gene Ontology(GO)/Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment. Finally, the receptor activator of nuclear factor-κB(RANKL)-induced osteoclast differentiation model and collagen-induced arthritis(CIA) model were established. Quantitative real time polymerase chain reaction(q-PCR), immunofluorescence, and immunohistochemistry were used to verify the pharmacological effect and molecular mechanism of artesunate in the treatment of bone destruction in RA. In this study, the RANKL-induced osteoclast differentiation model in vitro was established and intervened with artesunate, and transcriptome sequencing data were analyzed to obtain 744 DEGs of artesunate in inhibiting osteoclast differentiation. A total of 1 291 major target genes of bone destruction in RA were obtained from GeneCards and OMIM. The target genes of artesunate in inhibiting osteoclast differentiation and the target genes of bone destruction in RA were intersected to obtain 61 target genes of artesunate against bone destruction in RA. The intersected target genes were analyzed by GO/KEGG enrichment. According to the results previously reported, the cytokine-cytokine receptor interaction signaling pathway was selected for experimental verification. Artesunate intervention in the RANKL-induced osteoclast differentiation model showed that artesunate inhibited CC chemokine receptor 3(CCR3), CC chemokine receptor 1(CCR1) and leukemia inhibitory factor(LIF) mRNA expression in osteoclasts in a dose-dependent manner compared with the RANKL-induced group. Meanwhile, the results of immunofluorescence and immunohistochemistry showed that artesunate could dose-dependently reduce the expression of CCR3 in osteoclasts and joint tissues of the CIA rat model in vitro. This study indicated that artesunate regulated the CCR3 in the cytokine-cytokine receptor interaction signaling pathway in the treatment of bone destruction in RA and provided a new target gene for the treatment of bone destruction in RA.
Rats ; Animals ; Arthritis, Experimental/drug therapy* ; Artesunate/therapeutic use* ; Arthritis, Rheumatoid/genetics* ; Transcriptome ; Network Pharmacology ; Osteoclasts ; Receptors, Cytokine/therapeutic use*

BACKGROUNDAlcohol dependence (AD) is a complex disorder characterized by impaired control over drinking. It is determined by both genetic and environmental factors. The recent approach of genome-wide association study (GWAS) is a powerful tool for identifying complex disease-associated susceptibility alleles, however, a few GWASs have been conducted for AD, and their results are largely inconsistent. The present study aimed to screen the loci associated with alcohol-related phenotypes using GWAS technology.

METHODSA genome-wide association study with the behavior of regular alcohol drinking and alcohol consumption was performed to identify susceptibility genes associated with AD, using the Affymetrix 500K SNP array in an initial sample consisting of 904 unrelated Caucasian subjects. Then, the initial results in GWAS were replicated in three independent samples: 1972 Caucasians in 593 nuclear families, 761 unrelated Caucasian subjects, and 2955 unrelated Chinese Hans.

RESULTSSeveral genes were associated with the alcohol-related phenotypes at the genome-wide significance level, with the ankyrin repeat domain 7 gene (ANKRD7) showing the strongest statistical evidence for regular alcohol drinking and suggestive statistical evidence for alcohol consumption. In addition, certain haplotypes within the ANKRD7 and cytokine-like1 (CYTL1) genes were significantly associated with regular drinking behavior, such as one ANKRD7 block composed of the SNPs rs6466686-rs4295599-rs12531086 (P = 6.51 × 10(-8)). The association of alcohol consumption was successfully replicated with rs4295599 in ANKRD7 gene in independent Caucasian nuclear families and independent unrelated Chinese Hans, and with rs16836497 in CYTL1 gene in independent unrelated Caucasians. Meta-analyses based on both the GWAS and replication samples further supported the observed significant associations between the ANKRD7 or CYTL1 gene and alcohol consumption.

CONCLUSIONThe evidence suggests that ANKRD7 and CYTL1 genes may play an important role in the variance in AD risk.

Adult ; Aged ; Alcohol Drinking ; genetics ; Blood Proteins ; Cytokines ; Female ; Genome-Wide Association Study ; Haplotypes ; Humans ; Male ; Middle Aged ; Polymorphism, Single Nucleotide ; Proteins ; genetics ; Receptors, Cytokine ; genetics

OBJECTIVE: To investigate the clinical features and prognostic factors of acute lymphoblastic leukemia (ALL) children with P2RY8-CRLF2 gene rearrangement. METHODS: A total of 108 children with B-cell ALL (B-ALL) were diagnosed and systematically treated according to Chinese Children's Leukemia Group (CCLG) -ALL 2008 in our hospital from January 2016 to December 2016. The 108 patients were divided into two groups according to the result of mutiplex polymerase chain reaction: group with P2RY8-CRLF2 gene rearrangement and group without P2RY8-CRLF2 gene rearrangement. The ALL children with P2RY8-CRLF2 gene rearrangement were all treated by CCLG-ALL 2008 high-risk group (HR) regimens, and the ALL children in group without P2RY8-CRLF2 gene rearrangement received different intensity chemotherapy according to clinical risk classification. RESULTS: Five (4 male and 1 female) out of 108 patients with B-ALL had P2RY8-CRLF2 gene rearrangement. In the 5 B-ALL patients with P2RY8-CRLF2 gene rearrangement, the median age of the was 4 (2-6) years old and the median WBC count was 26.2 (2.46-525.1)×10 CONCLUSION The early treatment response and prognosis of ALL children with P2RY8-CRLF2 gene rearrangement are worse, and more effective protocol is needed for this subtype patients.
Child ; Child, Preschool ; Disease-Free Survival ; Female ; Gene Rearrangement ; Humans ; Male ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma/genetics* ; Precursor Cell Lymphoblastic Leukemia-Lymphoma/genetics* ; Prognosis ; Receptors, Cytokine/genetics* ; Receptors, Purinergic P2Y/genetics*

OBJECTIVETo investigate the expression of the granulocyte-macrophage colony-stimulating factor (GM-CSF) and GM-CSF/IL-3/IL-5 receptor common beta chain (beta c receptor) in an adult patient with idiopathic pulmonary alveolar proteinosis (PAP), so as to demonstrate the possible association of the GM-CSF and beta c receptor with the pathogenesis of human PAP.

METHODSThe GM-CSF levels were measured with a commercial ELISA kit (sensitivity 5 pg/ml) and the beta c receptor expression on the cell surface was detected by flow cytometry analysis. Reverse transcription polymerase chain reaction (RT-PCR) analysis was employed to detect the expression of the GM-CSF mRNA and the beta c receptor mRNA in peripheral blood mononuclear cells and alveolar macrophages. The entire coding regions of the GM-CSF cDNA and the beta c receptor cDNA were sequenced by the Sanger dideoxy-mediated chain termination method to detect possible mutations.

RESULTSThe patient with PAP failed to release the GM-CSF protein either from circulating mononuclear cells or from alveolar macrophages. The expression of the GM-CSF mRNA was normal after the stimulation of lipopolysaccharide, whereas a point mutation at position 382 of the GM-CSF cDNA from "T" to "C" was revealed by cDNA sequencing, which caused a change in amino acid 117 of the protein from isoleucine to threonine. The beta c receptor expression on the cell surface was normal, and the beta c receptor mRNA expression and the sequence of the entire coding region of the beta c receptor were also normal.

CONCLUSIONSThe decreased GM-CSF production is associated with the pathogenesis of human PAP. A point mutation of the GM-CSF cDNA may contribute to the decreased GM-CSF production in our adult PAP patient. The mutation of the beta c receptor in some of paediatric patients with PAP may not be a common problem in adult patients.

DNA, Complementary ; chemistry ; Granulocyte-Macrophage Colony-Stimulating Factor ; analysis ; biosynthesis ; Humans ; Male ; Middle Aged ; Pulmonary Alveolar Proteinosis ; etiology ; metabolism ; RNA, Messenger ; analysis ; Receptors, Cytokine ; biosynthesis ; genetics

OBJECTIVE: To investigate the expression of EBI3 and p28 mRNA (the 2 subunits of IL-27) in the brain and spinal cord of the model of experimental autoimmune encephalomyelitis (EAE), and to explore their effect on EAE. METHODS: Seventy-two adult female SPF C57BL/6J mice (inbred strain) were randomly divided into a control group, an adjuvant group, and an EAE group. RT-PCR was performed to detect the expression of EBI3 mRNA and p28 mRNA in the brain and spinal cord. RESULTS: The expression of EBI3 mRNA and p28 mRNA was up-regulated at onset in the EAE group, which increased quickly during peak phase and maintained at a high level in the chronic phase. There was significant difference in the expression of EBI3 and p28 mRNA between the EAE group and the control/adjuvant group (P<0.01). Additionally, there was no remarkable difference in the expression of EBI3 and p28 mRNA in the brain and spinal cord between the control group and the adjuvant group (P>0.05). CONCLUSION IL-27 may play a role of promoting the morbility of EAE in the early stage, and sustain the inflammatory response in endgame.
Animals ; Brain ; metabolism ; Chronic Disease ; Encephalomyelitis, Autoimmune, Experimental ; metabolism ; Interleukins ; genetics ; metabolism ; Male ; Mice ; Mice, Inbred C57BL ; Minor Histocompatibility Antigens ; Protein Subunits ; genetics ; metabolism ; RNA, Messenger ; genetics ; metabolism ; Receptors, Cytokine ; genetics ; metabolism ; Spinal Cord ; metabolism

IL-28RA is one of the important candidate genes for complex trait of genetic diseases, but there is no published information of the genetic variation in this gene. We scanned the seven exons and their boundary introns sequence of IL-28RA including the promoter regions to analyze genetic variation sites, and identified eighteen single nucleotide polymorphisms (SNPs) and two variation sites. We chose seven SNPs (g.-1193 A>C, g.-30 C>T, g.17654 C>T, g.27798 A>G, g.31265 C>T, g.31911 C>T and g.32349 G>A) of them for large sample size genotyping, and assessed the association of genotype and allele frequencies of these SNPs between allergic rhinitis patients and non-allergic rhinitis controls. We also compared the genotype frequencies between Korean controls and Han Chinese control or Korean Chinese control. We investigated the frequencies of haplotype constructed by these SNPs between allergic rhinitis patients and non-allergic rhinitis controls. Our results suggested that the g.32349 G>A polymorphism of IL-28RA might be associated with susceptibility to allergic rhinitis (P=0.032), but seems to have no relationship with serum total IgE levels. The haplotype frequencies by these SNPs also show significant association between controls and allergic rhinitis patients.
Variation (Genetics) ; Rhinitis, Allergic, Seasonal/blood/*genetics ; Rhinitis, Allergic, Perennial/blood/*genetics ; Receptors, Cytokine/*genetics ; Promoter Regions (Genetics)/genetics ; Polymorphism, Single Nucleotide/*genetics ; Male ; Immunoglobulin E/blood ; Humans ; Haplotypes ; Genotype ; Genetic Predisposition to Disease/genetics ; Gene Frequency ; Female ; Exons/genetics ; Case-Control Studies ; Alleles ; Adult

OBJECTIVETo investigate the expression of fractalkine (FKN) and its receptor CX3CR1 in cardiac allografts and the effect of Cyclosporin A (CsA).

METHODSThree groups of rats underwent heterotopic cardiac transplantation, 45 cases in each group and 5 cases in control group: SD to SD regarded as isograft group (group A), Wistar to SD divided into CsA untreated allograft group (group B) and CsA treated allograft group (group C), normal SD rats as control group. The FKN mRNA expression was detected by one-step RT-PCR method and the expression of FKN and CX3CR1 protein was detected by standard ABC immunohistochemical technique.

RESULTSThe expression of FKN mRNA and protein was weak in both isografts and normal heart specimens. The changes of FKN mRNA expression were correlated with the process of acute allograft rejection. The peak of FKN mRNA expression (0.8 +/- 0.26) appeared on the seventh day after transplantation, which could be down-regulated by CsA significantly (t = 2.390, P < 0.05). FKN protein was located in endothelia cells and its receptor CX3CR1 was located in infiltrating mononuclear cells in allografts.

CONCLUSIONSUpregulation of FKN and its receptor was significantly correlated with the trafficking of mononuclear cells which play an important role in acute allograft rejection. It may be one of the important mechanisms of CsA to intervene the acute rejection by inhibiting the activation of the FKN-CX3CR1 pathway.

Acute Disease ; Animals ; CX3C Chemokine Receptor 1 ; Chemokine CX3CL1 ; Chemokines, CX3C ; genetics ; metabolism ; Cyclosporine ; pharmacology ; Graft Rejection ; immunology ; pathology ; prevention & control ; Heart Transplantation ; immunology ; pathology ; Immunohistochemistry ; Male ; Membrane Proteins ; genetics ; metabolism ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Rats, Wistar ; Receptors, Cytokine ; genetics ; metabolism ; Receptors, HIV ; genetics ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction ; Transplantation, Homologous

OBJECTIVETo observe the effect of Chinese herbs for supplementing Shen and strengthening bone (HB) on myelogenic osteoclasts formation, and gene expression of interleukin-6 (IL-6), IL-6 receptor (IL-6R) and gp130 in bone marrow.

METHODSSeventy-two healthy female SD rats of 3 months, were randomly divided into three groups, 24 in the sham-operated group (A), 24 in the ovariectomized group (B) and 24 in the after ovariectomy HB treated group (C). Bone marrow cells of 6 rats from each group were respectively collected and cultured at four time points (2nd, 4th, 6th and 12th weeks after operation). After 6 days of culture, the bone marrow cells were differentiated by Wright-Giemsa stain and TRAP stain, and total RNA in them was extracted by TRIZOL.

RESULTSBeginning from the 2nd week, the osteoclasts formation in Group B was higher than that in Group A (P < 0.05), and IL-6, IL-6R gene expression significantly increased in Group B (P < 0.05 or P < 0.01). These changes reached the peak in the 4th to 6th week, with the level maintained to the 12th week. As for comparison of Group B and C, the above-mentioned changes were significantly weakened in the latter (P < 0.05 or P < 0.01). No significant change of gp130 gene expression revealed in the whole course in either group.

CONCLUSIONHB could inhibit the myelogenic osteoclasts formation in ovariectomized rats, this effect may be correlated with, partially at least, its inhibitory effect on the over-expressed IL-6 and IL-6R gene expression in myelocytes after ovariectomy.

Animals ; Antigens, CD ; biosynthesis ; genetics ; Bone Marrow ; metabolism ; Cell Division ; drug effects ; Cells, Cultured ; Cytokine Receptor gp130 ; Drugs, Chinese Herbal ; chemistry ; pharmacology ; Female ; Granulocyte Precursor Cells ; metabolism ; Interleukin-6 ; biosynthesis ; genetics ; Isoflavones ; pharmacology ; Membrane Glycoproteins ; biosynthesis ; genetics ; Osteoblasts ; pathology ; Osteoporosis ; metabolism ; pathology ; Ovariectomy ; RNA ; biosynthesis ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Interleukin-6 ; biosynthesis ; genetics

OBJECTIVETo investigate the genetic linkage between several cytokine and cytokine-related receptor gene loci and essential hypertension (EH) in Chinese.

METHODSLinkage between seven genetic markers and EH in 95 Chinese nuclear families with EH (including 477 subjects) was analyzed using a technique of fluorescence-based gene scan with DNA short tandem repeat loci. These markers were selected from the chromosomal regions nearby eight cytokines and their receptor genes. The two-point non-parametric linkage analysis (NPL), maximum Lod score and transmission/disequilibrium test (TDT) with GENEHUNTER software package were used in this study.

RESULTSResult of TDT showed significant transmission disequilibrium between D14S61 and EH (Chi square 14.29,P=0.00016) although NPL and Lod score revealed no significant linkage (Z=0.78, P>0.05 and Lod score =0.72 respectively) at this locus. No linkage between other loci typed and EH was found by the three genetic analysis methods (P>0.05 or Lod score<-1).

CONCLUSIONAlleles at D14S61 were of significant transmission disequilibrium in affected siblings. Transforming growth factor beta 3 is 0.1 cM away from D14S61, which suggests that the relationship between genes at or near this regions and EH needs to be further explored.

Adult ; Alleles ; Blood Glucose ; metabolism ; Blood Pressure ; physiology ; Body Mass Index ; China ; Cholesterol ; blood ; Cholesterol, HDL ; blood ; Cholesterol, LDL ; blood ; Cytokines ; genetics ; DNA ; genetics ; Family Health ; Female ; Genetic Linkage ; Humans ; Hypertension ; blood ; genetics ; physiopathology ; Linkage Disequilibrium ; Lod Score ; Male ; Microsatellite Repeats ; Middle Aged ; Receptors, Cytokine ; genetics ; Triglycerides ; blood

OBJECTIVETo investigate the mechanism of growth hormone insensitivity of rats under the endotoxemic condition.

METHODSSprague Dawley rats (n = 180) were injected endotoxin, TNF-alpha, and IL-6 respectively. Part of endotoxin injected rats were treated with exogenous somatotropin simultaneously, and all rats were killed at different time points. Liver expression of IGF I, GHR and SOCS-3 mRNA was detected by RT-PCR, the levels of growth hormone (GH) were measured by radioimmunoassay, and the levels of TNF-alpha and IL-6 were detected by ELISA.

RESULTSSerum GH levels showed no significant change after endotoxin injection; however, liver IGF I and GHR mRNA expressions were obviously down-regulated in endotoxemic rats, with the lowest decrease of 53% and 89% respectively. Although SOCS-3 mRNA was weakly expressed in control rats, it was strongly up-regulated in endotoxemic rats and the marked increase was 7.84 folds. The higher LPS dosage induced marked GHR mRNA down-regulation and marked SOCS-3 mRNA up-regulation. Exogenous GH made IGFI mRNA expression increase 25% in the control rats, but it did fail to prevent the decline in IGFI mRNA expression in endotoxemic rats. Endotoxin stimulated the production of TNF-alpha and IL-6, and the elevated IL-6 levels showed a positive correlation with increased SOCS-3 mRNA expression. Liver GHR mRNA expression was obviously down-regulated after TNF-alpha i.v. injection, but for IL-6, it mainly up-regulated the liver SOCS-3 mRNA expression.

CONCLUSIONThe growth hormone insensitivity could be induced by LPS injection, which might be associated with down-regulated GHR mRNA expression and up-regulated SOCS-3 mRNA expression. The in vivo biological activities of LPS may be partially mediated by TNF-alpha and IL-6 at different aspects.

Animals ; Down-Regulation ; Growth Hormone ; metabolism ; pharmacology ; Insulin-Like Growth Factor I ; genetics ; metabolism ; Interleukin-6 ; pharmacology ; physiology ; Lipopolysaccharides ; toxicity ; Liver ; drug effects ; metabolism ; Male ; RNA, Messenger ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Somatotropin ; genetics ; Suppressor of Cytokine Signaling Proteins ; genetics ; metabolism ; Tumor Necrosis Factor-alpha ; pharmacology ; physiology