rheumatoid arthritis ( ra ) is a chronic disease characterized by synovitis and autoantibody such as rheumatoid factor ( rf ) and anti - cyclic citrullinated peptide antibody ( anti - ccp ) [ 1 , 2 ] . besides the destruction of cartilage and bone , ra also leads to system damage including cardiovascular , pulmonary , and psychological systems [ 3 , 4 ] . blockage of proinflammatory cytokines activity involved in the chronic inflammation of ra , such as tumor necrosis factor - alpha ( tnf- ) and interleukin-6 ( il-6 ) , was applied in clinical therapy . il-6 is a pleiotropic proinflammatory cytokine produced by synovial cells and endothelial cells [ 6 , 7 ] . il-6 favors t - cell activation , b - cell proliferation , and immunoglobulin secretion . moreover , il-6 participates in the synthesis of acute phase proteins in the liver and proliferation and differentiation of hematopoietic precursor cells . previous studies have defined the relationship between inflammatory factors and insulin resistance [ 1012 ] . increased levels of serum inflammatory marker in addition , il-6 treatment induces beta - cell apoptosis via stat-3-mediated nitric oxide production [ 1315 ] . recent reports have shown that the prevalence of diabetes mellitus ( dm ) in patients with ra is about 15%19% , which is significantly higher than that of the global dm [ 16 , 17 ] . the fasting plasma glucose ( fbg ) in ra patients with type 2 diabetes mellitus ( t2 dm ) is higher than that of t2 dm patients , suggesting that the islet -cell damage in ra patients complicated with t2 dm is more serious , and these may be related to their long - term systemic inflammatory status [ 18 , 19 ] . however , the mechanism of ra with -cell damage is still uncertain . in this study , the model of cia was established to observe the expressions of fbg , fins , il-6 , and islet caspase-3 . furthermore , in line with increased il-6 expression , the apoptosis related enzyme caspase-3 was also markedly increased . these data showed that the possible mechanism of ra complicated with abnormal glucose metabolism might result from the -cell apoptosis induced by il-6 . female wistar rats , weighing 150180  g , were purchased from kaifu district , changsha city , east laboratory animal service department , and were bred in a specific pathogen - free ( spf ) environment . all rats were maintained ( 2024c and 5560% humidity ) under a standard 12-hour light / dark cycle . il-6 elisa kits were purchased from raybiotech company ( norcross , usa ) ; insulin elisa kits were purchased from wuhan elabscience biotechnology co. , ltd . ( wuhan , china ) ; bovine type ii collagen ( c7806 ) and complete freund 's adjuvant were purchased from sigma company ( shanghai , china ) ; rabbit anti - rat caspase-3 was provided by abcam company ( cambridge , usa ) . two - step immunohistochemical kits and diaminobenzidine ( dab ) color liquid were purchased from beijing zhong shan - golden bridge biological technology co. , ltd . ( beijing , china ) . bovine type ii collagen ( 10  mg ) was dissolved in 5  ml 0.1  mol / l acetic acid solution , maintained at 4c overnight , and set aside . an equal dose of collagen solution and complete freund 's adjuvant was thoroughly mixed and completely emulsified ; then , a cii / ifa emulsion ( 1  mg / ml ) was freshly manufactured . rats were injected with emulsion at the base of the tail and back within three points ( day 1 ) ; animals were boosted with the dose of 0.2  ml cii - ifa emulsion at day 7 by the same method . the control group were treated with complete freund 's adjuvant according to the same protocol . after initial immunization , the rats ' arthritis index ( ai ) was scored as in a previous report . blood samples were collected from the intraocular adjoined veins of the rats on the 17th day after initial immunization . the pancreatic tissue was fixed in 4% paraformaldehyde ( ph 7.4 ) for histological analysis . to detect the expression of caspase-3 , the slides were incubated with antibody diluted at 1  :  80 overnight at 4c , followed by dab coloration , and stained with hematoxylin dye and dehydrated ( transparent ) . image - pro plus 6.0 was used to analyze and carefully delineate the scope of each field in the islets , to measure the mean absorbance of the islets , as an index for expression of islets caspase-3 .   the protein extracted from the pancreatic tissue antibodies against cleaved caspase-3 were obtained from cell signaling technology ( ma , usa ) . after incubating with secondary antibody conjugated horseradish peroxidase , the immunoreactivity was developed by an ecl system ( pierce ) . paired student 's t - test was used for paired data . a value of p < 0.05 was considered as statistically significant . in recent studies , there has been increased evidence of the relationship between glucose metabolism dysfunction and human autoimmune disease . to explore the mechanism of this observation in rheumatoid arthritis , rat collagen - induced arthritis model was established . in this study , on day 17 after the primary immunization , the cia rats showed mild edema in one or both sides of the hind limbs . furthermore , histological analysis showed hyperplasia of the synovial membrane and the infiltration of inflammatory cells in cia rats ( figures 1(a ) and 1(b ) ) . as expected , a markedly increased level of fbg was observed in cia rats when compared with the control group . meanwhile , a decreased level of fins was observed in cia rats when compared with the control group ( figure 2 ) . il-6 is a critical inflammatory cytokine in the pathogenesis of ra , and blockage of its activity by il-6r antibody was applied in clinical treatment . here , we also observed an increased expression of il-6 in cia rats when compared with control ( figure 3 ) . next , to determine the role of il-6 in the regulation of glucose metabolism in cia rats , the correlation between il-6 and fbg or fins was analyzed . the result met our expectations ; a significantly positive correlation was observed between il-6 and fbg ( figure 4(a ) ) . in addition , a negative correlation was depicted in il-6 and fins ( figure 4(b ) ) . a critical role of -cell apoptosis has been demonstrated in the development of diabetes . in line with an increased expression of il-6 ( figure 3 ) , the apoptosis related enzyme caspase-3 was also markedly increased in cia rats ( figures 5(a)5(c ) ) . furthermore , the cleaved caspase-3 was also detected , as expected , and the cia rats showed an increased level when compared with control rats . previous studies have shown that il-6 treatment induces beta - cell apoptosis via stat-3-mediated nitric oxide production . accordingly , a positive correlation between caspase-3 and il-6 was observed in our study . taken together , these results suggest that the abnormal glucose metabolism might be due to the -cell apoptosis induced by inflammatory cytokine il-6 ( figure 5(d ) ) . recent studies have shown that the prevalence of ra patients with abnormal glucose metabolism was significantly increased [ 11 , 16 ] , and the decreased insulin sensitivity and islet -cell function due to islet -cell excessive apoptosis in these patients and the mechanism might be related to overexpressed proinflammatory cytokines . in this experiment , the expression of il-6 in group cia was significantly higher than that of control group , and the abnormal glucose metabolism was accompanied by the increased il-6 expression . furthermore , in line with increased il-6 expression , the apoptosis related enzyme caspase-3 was also markedly increased . these data showed that the abnormal glucose metabolism in ra might result from the -cell apoptosis induced by il-6 . il-6 , an important proinflammatory cytokine , could promote insulin secretion at low concentration of il-6 . the cytotoxicity of high il-6 concentrations could upregulate the expression of cleaved caspase-3 and phosphorylated p38 mitogen - activated protein kinase and phosphorylated nuclear factor-b , in part via signal transducers and activators of transcription-3-mediated no production . in addition , il-6 can increase lipolysis in adipocytes as well as the release of free fatty acids , thereby damaging the mitochondria and glucose transporter 2 ( glut2 ) function and insulin sensitivity . here , in our study , we also observed elevated il-6 expression in cia , which positively correlated with glucose . furthermore , il-6 can also induce the production of il-1 , il-2 , and tnf- and enhance the biological effects when combined with these cytokines . the previous studies have also found that tnf- produced by adipose tissue can induce insulin resistance ( ir ) ; these data showed the relationship between inflammatory factors and insulin resistance . not only were il-6 levels in cia rats significantly increased , but also other proinflammatory cytokines like tnf- and ifn- were overexpressed . il-6 can also be combined with tnf- , il-1 , and induced classical caspase - dependent apoptosis pathway , thereby increasing the apoptosis of -cells and causing a reduction of insulin secretion . these might be the reason for the high prevalence of diabetes mellitus ( dm ) in patients with ra when compared with healthy controls . insulin is one of the most important hormones in an organism that regulates blood sugar levels , which is secreted by pancreatic -cells ; insulin deficiency can lead to elevated blood glucose levels . as expected , we observed that fasting serum insulin levels were significantly decreased in cia . when the cells are stimulated by the proapoptotic factors , caspase-3 can be activated and then induce their apoptosis [ 24 , 25 ] . in addition , il-6 was positively correlated with caspase-3 production which leads -cell to apoptosis in our study . the elevated levels of il-6 inhibiting the insulin secretion of islet -cells might be through promoting -cells apoptosis . this study suggests the significantly increased fbg and decreased fins in cia rats , which might be a result from autoimmune inflammatory mediators such as il-6 . il-6 induced abnormal glucose metabolism might be through upregulating caspase-3 expression , because of the positive correlation between il-6 and apoptosis related enzyme caspase-3 . taken together , these data showed that the elevated fbg and reduced fins level in cia may be induced by the inflammatory cytokines generated by arthritis , which is significantly different from the traditional t2 dm . the treatment of the primary disease in ra combined with dm can improve glucose metabolism .
the aim of this study was to explore the possible mechanism of rheumatoid arthritis- ( ra- ) related abnormal glucose metabolism . 
 the model of collagen - induced arthritis ( cia ) was established by intradermal injection of type ii collagen into wistar rats ; complete freund 's adjuvant injections were used as the control group . fasting plasma glucose ( fbg ) 
 was measured by the glucose oxidase method . 
 fasting insulin ( fins ) and the expressions of il-6 were detected by elisa . 
 islet caspase-3 was examined by immunohistochemistry . on day 17 after immunization , fbg of the cia group showed an elevated fbg value compared with the control group . meanwhile , the fins of group cia was lower when compared with the control group . 
 interestingly , the inflammatory cytokine il-6 expression was significantly increased when compared with the control group . 
 as expected , the abnormal glucose metabolism was accompanied by the increased il-6 expression . 
 furthermore , in line with the upregulated il-6 expression , the apoptosis related enzyme caspase-3 was also markedly increased . 
 these data showed that the elevated fbg in cia may be associated with the reduced fins level secondary to the overapoptosis of pancreas islet cells induced by il-6 .