Severity Of Arthritis In Rats Research Articles

Anti-rheumatoid arthritis potential of Rhododendron molle G. Don leaf extract in adjuvant induced arthritis rats.

The aim of this study was to test the anti-rheumatic arthritis effects of Rhododendron molle G. Don leaf extract in arthritis rats and inflammatory RAW 264.7cells. Preliminary analysis and comparison of potential medicinal components of three polar extracts by HPLC and UHPLC-Q-TOF-MS. SD rats were subcutaneously injected with complete Freund's adjuvant (CFA) to induce inflammation on the right hind paw. RAW 264.7cells were induced by lipopolysaccharide (LPS) to established cell inflammatory model. The volume of rat hind paw was measured with a volume meter to detect swelling, and the weight of rats was measured with an electronic balance. The severity of arthritis in rats was evaluated by arthritis score. The pathological sections of rat hind paw joints were observed by hematoxylin-eosin staining, and the contents of IL-6 and IL-1β in serum were detected. qRT-PCR was used to detect the expression of IL-1β, IL-6, TNF-α and COX-2 genes in RAW 264.7cells. The release of nitric oxide was measured by Griess reaction. The expression levels of IL-6 and IL-1β were detected by Western-Blot. and discussion: The chloroform extract from R. molle leaves (CERL), Ethyl acetate extract from R. molle leaves (EERL), n-butanol extract from R. molle leaves (BERL) could significantly inhibit hind paws swelling and reduce arthritis index in arthritis rats. And it showed dose dependence. Compared with tripterygium glycosides (TG) tablets, an effective drug of RA treatment, CERL have better anti-RA effect after administration. In addition, the three kinds of the polar extracts of Rhododendron molle leaves (PERL) had lower toxicity, with the LD50 279.87, 239.65, 500.08 (mg/kg) respectively, while TG group's LD50 was 96.00 (mg/kg). In vitro experiments showed that the three PERLs can significantly inhibit the level of pro-inflammatory factors and inflammatory mediator, such as TNF-α, IL-1β, IL-6, COX-2 and NO, which were consistent with their anti-RA ability. Among the three kinds of PERLs, CERL showed the best inhibitory activity. The R. molle leaf is a potential medicinal part for the treatment of RA. This study explored the anti-RA and anti-inflammatory activities of CERL, EERL, BERL, which laid a foundation for further promoting the clinical application of R. molle.

Amelioration of oxidative and inflammatory changes by Peganum harmala seeds in experimental arthritis

BackgroundThe present study was designed to investigate the therapeutic effects against oxidative stress and alleviative effects of Peganum harmala seeds (PH) in rats with Complete Freund’s Adjuvant (CFA) induced arthritis.MethodsThe extract was evaluated for its phytoconstituents, antiarthritic and antioxidant properties. The action of chloroform (PHC) and ethanolic (PHE) extracts of PH was evaluated in adult Lewis rats (150-200 g).with CFA induced arthritis. Arthritic rats received PH extracts 100 mg/kg orally for 28 consecutive days (Prophylactic treatment) and from 14th day of CFA injection (Therapeutic treatment).ResultsPHE significantly suppressed the arthritis severity in rats than PHC in 28 days. All complications shown significant reduction (p< 0.05) in arthritic rats including paw volume (63.09%), body weight loss, decreased locomotor activity, erythrocyte sedimentation rate and synovial/hepatic tissues lipid peroxidation and increase in cellular antioxidants superoxide dismutase (U/mg) activities and hemoglobin counts. The results showed the presence of alkaloids and flavonoids in PHE. Histology and radiographic analysis of arthritic ankle joints indicated abnormal changes. Marked reduction in inflammation and arthritic changes were observed after treatment with PHE.ConclusionTherefore, the investigation suggests that PHE at 100 mg/kg will be useful in the management of rheumatoid arthritis complications which may possibly be due to boosting the intracellular antioxidant defense.

Glycogen Metabolism and Rheumatoid Arthritis: The Role of Glycogen Synthase 1 in Regulation of Synovial Inflammation via Blocking AMP-Activated Protein Kinase Activation.

To investigate the role of glycogen metabolism in regulating rheumatoid fibroblast-like synoviocyte (FLS)-mediated synovial inflammation and its underlying mechanism. FLSs were separated from synovial tissues (STs) obtained from rheumatoid arthritis (RA) patients. Glycogen content was determined by periodic acid Schiff staining. Protein expression was analyzed by Western blot or immunohistochemistry. Gene expression of cytokines and matrix metalloproteinases (MMPs) was evaluated by quantitative real-time PCR. FLS proliferation was detected by EdU incorporation. Migration and invasion were measured by Boyden chamber assay. Glycogen levels and glycogen synthase 1 (GYS1) expression were significantly increased in the ST and FLSs of RA patients. TNF-α or hypoxia induced GYS1 expression and glycogen synthesis in RA FLSs. GYS1 knockdown by shRNA decreased the expression of IL-1β, IL-6, CCL-2, MMP-1, and MMP-9 and proliferation and migration by increasing AMP-activated protein kinase (AMPK) activity in RA FLS. AMPK inhibitor or knockdown AMPK could reverse the inhibitory effect of GYS1 knockdown on the inflammatory response in RA FLSs; however, an AMPK agonist blocked RA FLS activity. We further determined that hypoxia-inducible factor-1α mediates TNF-α- or hypoxia-induced GYS1 expression and glycogen levels. Local joint depletion of GYS1 or intraperitoneal administration with an AMPK agonist ameliorated the severity of arthritis in rats with collagen-induced arthritis. Our data demonstrate that GYS1-mediated glycogen accumulation contributes to FLS-mediated synovial inflammation in RA by blocking AMPK activation. In our knowledge, this is a first study linking glycogen metabolism to chronic inflammation. Inhibition of GYS1 might be a novel therapeutic strategy for chronic inflammatory arthritis, including RA.

Conserved 33-kb haplotype in the MHC class III region regulates chronic arthritis

Genome-wide association studies have revealed many genetic loci associated with complex autoimmune diseases. In rheumatoid arthritis (RA), the MHC gene HLA-DRB1 is the strongest candidate predicting disease development. It has been suggested that other immune-regulating genes in the MHC contribute to the disease risk, but this contribution has been difficult to show because of the strong linkage disequilibrium within the MHC. We isolated genomic regions in the form of congenic fragments in rats to test whether there are additional susceptibility loci in the MHC. By both congenic mapping in inbred strains and SNP typing in wild rats, we identified a conserved, 33-kb large haplotype Ltab-Ncr3 in the MHC-III region, which regulates the onset, severity, and chronicity of arthritis. The Ltab-Ncr3 haplotype consists of five polymorphic immunoregulatory genes: Lta (lymphotoxin-α), Tnf, Ltb (lymphotoxin-β), Lst1 (leukocyte-specific transcript 1), and Ncr3 (natural cytotoxicity-triggering receptor 3). Significant correlation in the expression of the Ltab-Ncr3 genes suggests that interaction of these genes may be important in keeping these genes clustered together as a conserved haplotype. We studied the arthritis association and the spliceo-transcriptome of four different Ltab-Ncr3 haplotypes and showed that higher Ltb and Ncr3 expression, lower Lst1 expression, and the expression of a shorter splice variant of Lst1 correlate with reduced arthritis severity in rats. Interestingly, patients with mild RA also showed higher NCR3 expression and lower LST1 expression than patients with severe RA. These data demonstrate the importance of a conserved haplotype in the regulation of complex diseases such as arthritis.

Positioning of a Polymorphic Quantitative Trait Nucleotide in theNcf1Gene Controlling Oxidative Burst Response and Arthritis Severity in Rats

The Ncf1 gene, encoding the P47(PHOX) protein that regulates production of reactive oxygen species (ROS) by the phagocyte NADPH oxidase (NOX2) complex, is associated with autoimmunity and arthritis severity in rats. We have now identified that the single-nucleotide polymorphism (SNP) resulting in an M153T amino acid substitution mediates arthritis resistance and thus explains the molecular polymorphism underlying the earlier identified Ncf1 gene effect. We identified the SNP in position 153 to regulate ROS production using COS(PHOX) cells transfected with mutated Ncf1. To determine the role of this SNP for control of arthritis, we used the Wistar strain, identified to carry only the postulated arthritis resistant SNP in position 153. When this Ncf1 allele was backcrossed to the arthritis susceptible DA strain, both granulocyte ROS production and arthritis resistance were restored. Position 153 is located in the hinge region between the PX and SH3 domains of P47(PHOX). Mutational analysis of this position revealed a need for an -OH group in the side chain but we found no evidence for phosphorylation. The polymorphism did not affect assembly of the P47(PHOX)/P67(PHOX) complex in the cytosol or membrane localization, but is likely to operate downstream of assembly, affecting activity of the membrane NOX2 complex.

Inconsistent susceptibility to autoimmunity in inbred LEW rats is due to genetic crossbreeding involving segregation of the arthritis-regulating gene Ncf1

We recently identified a single-nucleotide polymorphism in the Ncf1 gene, a component of the NADPH oxidase complex, to be the cause of one of the strongest identified loci for arthritis severity in rats. This polymorphism was found to be naturally occurring in a collection of inbred rat strains as well as in wild rats. Among the inbred strains we found that different LEW substrains (LEW/Ztm and LEW/Mol), originating from different breeders, showed an allelic discrepancy in Ncf1, suggesting an impact on arthritis susceptibility between these substrains. In fact, the LEW/Mol strain was completely resistant to pristane-induced arthritis, in contrast to the LEW/Ztm strain, which was susceptible. Moreover, the LEW/Mol strain had higher production of radical oxygen species in peripheral blood leukocytes, a phenomenon most likely regulated by the polymorphisms in the Ncf1 gene. However, the phenotypic difference between LEW/Mol and LEW/Ztm is most likely a combination of several genes, of which Ncf1 is suggested to be the major regulating gene. This has also been confirmed by previous linkage analyses involving the LEW/Ztm strain which shows that a QTL on chromosome 12, most likely caused by polymorphism of Ncf1, is the major regulatory gene but that other loci are contributing. That more genes are likely to contribute was shown by a complete genome comparison of the LEW/Ztm and the LEW/Mol rat strains that uncovered an introduction of approximately 37% non-LEW genome into the LEW/Mol strain, which probably was caused by past crossbreeding. Therefore, the LEW/Mol should be regarded as a recombinant inbred strain.

Positional identification of Ncf1 as a gene that regulates arthritis severity in rats.

The identification of genes underlying quantitative-trait loci (QTL) for complex diseases, such as rheumatoid arthritis, is a challenging and difficult task for the human genome project. Through positional cloning of the Pia4 QTL in rats, we found that a naturally occurring polymorphism of Ncf1 (encoding neutrophil cytosolic factor 1, a component of the NADPH oxidase complex) regulates arthritis severity. The disease-related allele of Ncf1 has reduced oxidative burst response and promotes activation of arthritogenic T cells. Pharmacological treatment with substances that activate the NADPH oxidase complex is shown to ameliorate arthritis. Hence, Ncf1 is associated with a new autoimmune mechanism leading to severe destructive arthritis, notably similar to rheumatoid arthritis in humans.

Effects of ovariectomy and estrogen replacement therapy on arthritis and bone mineral density in rats with collagen-induced arthritis

We investigated the effects of ovariectomy (ovx) and estrogen replacement therapy (ERT) on bone mineral density (BMD) and arthritis severity in rats with collagen-induced arthritis (CIA). Seven-month-old female Sprague-Dawley rats were separated into a sham group (n = 8), CIA group (n = 14), ovx group (n = 10), CIA + ovx group (n = 11), and CIA + ovx + ERT group (n = 14). In these groups, ovx was performed at 7 days, and ERT (17β-estradiol at 20 μg/kg three times per week) was initiated 8 days after sensitization. Every 2 weeks, until 8 weeks after sensitization, arthritis score and hind paw thickness were evaluated, and BMD of the trabecular and cortical bones in the metaphysis and diaphysis of the tibia were measured by peripheral quantitative computed tomography. The arthritis score was highest in the CIA + ovx group at all timepoints after sensitization. The hind paw thickness was significantly higher in the CIA + ovx group than in the CIA group at 8 weeks after sensitization ( p < 0.05). Both the arthritis score and hind paw thickness were lower in the CIA + ovx + ERT group than in the CIA + ovx group. BMD in the metaphysis was significantly decreased in both the trabecular and cortical bones in the CIA + ovx group compared with those in the CIA group at 4, 6, and 8 weeks after sensitization. In the CIA + ovx group, trabecular BMD was changed by −34 ± 11%, and cortical BMD changed by −14 ± 7% in the metaphysis at 8 weeks compared with those at 0 week. In the CIA group, changes of BMD in the metaphysis were −7 ± 11% in trabecular bone and 0 ± 7% in cortical bone. These differences of trabecular and cortical bone loss in the metaphysis were significant (both p < 0.01). BMD reduction was significantly less in the CIA + ovx + ERT group than in the CIA + ovx group at 6 and 8 weeks after sensitization. Although BMD in the diaphysis was also reduced in the groups with CIA, the degree of reduction was smaller than in the metaphysis. We conclude that ovx in CIA rats could enhance the severity of arthritis and bone loss, and that ERT could suppress arthritis and bone loss.

Effect of infection with trypanosomes on the development of experimental allergic neuritis in rabbits.

DATA obtained from hospitals in western Nigeria1 have shown that rheumatoid arthritis is uncommon in that region and, when the disease occurs, it is clinically milder than in Europeans, involving few of the expected immunological changes2,3. Other conditions in which auto-immune processes are thought to be involved were also found to be rare, and Greenwood suggested that repeated exposure to parasitic infections might be responsible, at least in part, for the infrequent occurrence of these auto-immune diseases. Infection with the rodent malaria parasite Plasmodium berghei yoelii delayed the onset of haemolytic disease in 4 week old NZB mice and prevented the development of renal disease in (NZB×NZW) F1 mice at the same age4. The severity of adjuvant arthritis in rats was also reduced by simultaneous inoculation of P. berghei yoelii, which produces a very mild infection, but not by infection with a more virulent strain of P. berghei berghei5. We are investigating the effects of trypanosome parasites on the development in rabbits of experimental allergic neuritis (EAN), a demyelinating paralytic disease6, the genesis of which is primarily a cell-mediated type of immune response (manuscript in preparation), and the preliminary results are reported here.

Effect of Obstructive Jaundice on Polyarthritis in Rats.

SummaryLigation of the common bile duct of rats prior to or 20 hours after the injection of L7 organisms delayed the onset of arthritis, and the subsequent degree of joint involvement up to 12 days' time was markedly diminished. However, when duct ligation was performed after the onset of arthritis there was no effect upon the course of the disease. Since rat arthritis induced by the injection of hemolytic streptococci was not influenced by bile duct ligation, it seems probable that the effect of obstructive jaundice on the development and severity of L7 arthritis in rats was due to the bile solubility of the L7 organisms.