ulcerative colitis ( uc ) is an inflammatory bowel disease ( ibd ) whose etiology is only partially known . it is characterized by diffuse inflammation of the colonic mucosa , which starts in the rectum and extends continuously along the colon . it is presented in variable clinical course with periods of remission or activity . in general , patients with uc have increased body fat and decreased lean body mass , increased production of inflammatory cytokines and they maintain a chronic low - grade inflammation [ 24 ] . cardiovascular diseases ( cvd ) are also chronic disorders associated with ( low - grade ) inflammation . atherosclerosis is the main cause of cvd and is associated with endothelial dysfunction , increased accumulation of oxidized ldl ( oxldl ) , leukocytes , and smooth muscle cells in the intima of the artery , with release of inflammatory mediators ( tnf , il-6 , ccl-2 ) , resulting in the development and expansion of the atheroma plaque . risk factors for cvd include smoking , alcohol , sedentary , diet , hypertension , and diabetes mellitus ( dm ) and dyslipidemia , as well as emerging factors such as total leukocytes count , levels of hemoglobin , and reactive c protein , il-6 , and fibrinogen [ 58 ] . several studies have demonstrated that patients with chronic inflammatory diseases such as rheumatoid arthritis and systemic lupus erythematosus have higher incidence of cardiovascular disease ( cvd ) due to chronic and systemic inflammation [ 912 ] . several studies have suggested that patients with ibd also have higher risk of atherosclerosis and cardiovascular events compared the healthy population [ 7,1319 ] . however , there are no studies evaluating this risk in uc patients when the inflammatory process is controlled . the aim of this study was to evaluate the risk of cvd in women with uc , in clinical remission , using of aminosalicylates or azathioprine and aminosalicylates and to compare the results to those obtained in healthy matched volunteers and suggest a mechanism explaining the relationship between uc and cvd . the patients were selected at the reference center for inflammatory bowel disease of the alfa institute of gastroenterology , clinical hospital , federal university of minas gerais ( ufmg ) . inclusion criteria included women aged 1860 years that did not present hypertension , dyslipidemia , diabetes mellitus , hormonal disorders , or cardiovascular disease . those presenting symptoms and signs of disease activity or using glucocorticoids or biologic agents were excluded . from 144 patients with uc monitored in the reference center , only 21 met the inclusion criteria , showing the high prevalence of risk factors for cardiovascular disease in this population and the presence of active disease phase ( supplementary figure 1 ) . all participants were in clinical remission [ do not fit the truelove and witts   criteria ] by the use of aminosalicylates ( sulfasalazine or mesalazine ) ( 12 patients ) or azathioprine combined with aminosalicylate ( 9 patients ) . because the introduction of azathioprine could modify the immunological and inflammatory status of patients , we initially divided patients in an asa group ( aminosalicylates ) and an aza+asa group ( azathioprine + aminosalicylates ) to determine the inflammatory profile and cardiovascular risk ; later , we compared both treatments for remission to observe possible differences . the study was approved by the ethics committee in human research of the federal university of minas gerais ( protocol 342/11 ) . blood pressure and anthropometric data such as weight , height , body mass index ( bmi ) , waist circumference , and body composition ( body fat and body fat - free ) were also obtained . after 12 h of fasting , 20 ml blood was collected for biochemical and inflammatory markers analysis . total cholesterol , hdl - cholesterol , glycemia , total protein , albumin , hemoglobin , fibrinogen , and high - sensitive c reactive protein ( hs crp ) were assessed using commercial kits ( labstest and bioclin , belo horizonte , brazil ) . total and differential leukocyte count was obtained using a neubauer chamber , and a differential count was performed on slides using a rapid kit panoptic kit ( laborclin , paran , brazil ) . il-6 , tnf , il-1 , il-10 , tgf- , ccl-2 , and anti - oxldl antibodies were determined by elisa ( bd opteia  , united states ) . the 10- and 30-year framingham risk scores ( frs ) were determined using online calculators ( http://www.framinghamheartstudy.org/ ) , considering lipid profile or bmi . unpaired mann - whitney , wilcoxon , or student s t - tests were used for continuous variables . statistical analysis was performed using spss software ( statistical package for social sciences ) version 19.0 . body weight , bmi , waist circumference , and traditional risk factors for cvd such as glycemia , triglyceridemia , total cholesterol , and its fractions were similar between groups ( tables 1 , 2 ) , while body fat was increased and fat - free mass was reduced in asa group ( table 1 ) . asa patients also showed increased total proteins due to increase in globulin , with the concomitant decreased on albumin fraction ( figure 1 ) . diastolic and systolic blood pressures another important risk factor for cvd , and hscrp and esr were higher in asa patients ( table 1 , figure 1 ) . inflammatory markers such as il-6 and ccl2 were also increased in the asa group ( figure 2 ) . nitrite concentration ( indicating nitric oxide status ) and igg anti - oxldl ( an indicator of oxldl concentration ) were increased in women in the asa group ( figure 3 ) , suggesting an increased risk of atherosclerosis in these patients . the framingham scores showed that asa patients have a higher 10-year risk for cvd , considering lipid profile or bmi . similar results were obtained when assessing the 30-year risk of cvd , showing higher relative risk of cvd in the asa group ( figure 4 ) . together , changes in blood pressure and inflammation markers show that patients with uc in clinical remission and treated with aminosalicylates have higher risk of cvd when compared to healthy subjects . patients using of aza+asa had higher bmi ( p=0.06 ) , increased waist circumference , fat mass , and blood pressure , but there was no relation with physical activity in their routines ( table 1 ) . lipid profile , glycemia , total protein , globulin , albumin , hemoglobin , esr , and fibrinogen were similar between groups ( figure 1 , table 2 ) and the crp was higher in the aza+asa group ( figure 1 ) . levels of il-6 , tnf , ccl-2 , il-10 , tgf- ( figure 2 ) , nitrite , and igg anti - oxldl ( figure 3 ) were increased in the aza+asa group , showing higher risk of atherosclerosis and cvd in this group . simultaneously , adding azathioprine treatment promotes the increase of anti - inflammatory mediators in attempting to regulate the inflammatory process . the relative risk for cvd assessed by 10- and 30-year framingham scores was higher in the aza+asa group ( figure 4 ) , suggesting increased relative risk of cvd in these patients . after comparing the patients with their respective controls , the remission treatments ( asa and aza+asa ) were compared . women in the aza+asa group presented decreased levels of total protein , globulin , esr , and lymphocytes ( figure 1 , table 2 ) compared to the asa group , suggesting better regulation of the inflammatory process characteristic of the disease . patients in the aza+asa group had higher levels of tnf , il-6 , il-10 , and tgf- compared to the asa group . others parameters were similar between groups ( tables 1 , 2 , figure 3 ) . however , the presence of these differences between both treated uc groups and the 10 and 30-year framingham risk scores were similar between groups ( figure 4 ) . body weight , bmi , waist circumference , and traditional risk factors for cvd such as glycemia , triglyceridemia , total cholesterol , and its fractions were similar between groups ( tables 1 , 2 ) , while body fat was increased and fat - free mass was reduced in asa group ( table 1 ) . asa patients also showed increased total proteins due to increase in globulin , with the concomitant decreased on albumin fraction ( figure 1 ) . diastolic and systolic blood pressures another important risk factor for cvd , and hscrp and esr were higher in asa patients ( table 1 , figure 1 ) . inflammatory markers such as il-6 and ccl2 were also increased in the asa group ( figure 2 ) . nitrite concentration ( indicating nitric oxide status ) and igg anti - oxldl ( an indicator of oxldl concentration ) were increased in women in the asa group ( figure 3 ) , suggesting an increased risk of atherosclerosis in these patients . the framingham scores showed that asa patients have a higher 10-year risk for cvd , considering lipid profile or bmi . similar results were obtained when assessing the 30-year risk of cvd , showing higher relative risk of cvd in the asa group ( figure 4 ) . together , changes in blood pressure and inflammation markers show that patients with uc in clinical remission and treated with aminosalicylates have higher risk of cvd when compared to healthy subjects . patients using of aza+asa had higher bmi ( p=0.06 ) , increased waist circumference , fat mass , and blood pressure , but there was no relation with physical activity in their routines ( table 1 ) . lipid profile , glycemia , total protein , globulin , albumin , hemoglobin , esr , and fibrinogen were similar between groups ( figure 1 , table 2 ) and the crp was higher in the aza+asa group ( figure 1 ) . levels of il-6 , tnf , ccl-2 , il-10 , tgf- ( figure 2 ) , nitrite , and igg anti - oxldl ( figure 3 ) were increased in the aza+asa group , showing higher risk of atherosclerosis and cvd in this group . simultaneously , adding azathioprine treatment promotes the increase of anti - inflammatory mediators in attempting to regulate the inflammatory process . the relative risk for cvd assessed by 10- and 30-year framingham scores was higher in the aza+asa group ( figure 4 ) , suggesting increased relative risk of cvd in these patients . after comparing the patients with their respective controls , the remission treatments ( asa and aza+asa ) were compared . women in the aza+asa group presented decreased levels of total protein , globulin , esr , and lymphocytes ( figure 1 , table 2 ) compared to the asa group , suggesting better regulation of the inflammatory process characteristic of the disease . patients in the aza+asa group had higher levels of tnf , il-6 , il-10 , and tgf- compared to the asa group . others parameters were similar between groups ( tables 1 , 2 , figure 3 ) . however , the presence of these differences between both treated uc groups and the 10 and 30-year framingham risk scores were similar between groups ( figure 4 ) . in this paper we highlight the mechanisms by which patients with uc in clinical remission are at higher risk for cvd and how drug treatment influences this . this is the first work performed in patients with uc in clinical remission that associates markers of inflammation , risk of cvd , and drug treatment . compared with healthy women , women with uc in clinical remission showed increased body fat with reduced fat - free mass . the aza+asa group still had an increase in bmi and waist circumference and in risk markers for cvd , including diseases such as diabetes mellitus and dyslipidemia . changes in body composition have been reported in ibd patients   and could be related to several factors such as poor eating habits , physical inactivity , and previous use of steroids . furthermore , patients with crohn s disease have increased number of adipocytes , with reduced volume and increased inflammatory infiltrate of mononuclear cells , favoring expansion and inflammatory capacity . adipose tissue is an endocrine organ that produces and secretes inflammatory cytokines and nitric oxide , leading to a chronic low - grade systemic inflammation , increasing the risk for cvd . the expansion of adipose tissue may contribute to increased blood pressure , another risk factor for cvd that has been observed in ibd   and in this study . even in remission , patients with uc have a subclinical inflammatory response in the colon , which may also contribute to the increase of these inflammatory mediators . the reduction in fat - free mass in the uc group can indicate changes in muscle mass , bone mass , and hydration status and appears to be due to poor dietary habits , physical inactivity , previous use of corticosteroids , or even due to catabolic process in common chronic diseases . the increased levels of circulating cytokines and globulins ( igg anti - oxldl and anti - cytokine antibodies ) favors the diversion of amino acids for production of inflammatory mediators affecting protein synthesis and body composition . the increase of total protein seen in the asa group is the result of increased globulins . albumin synthesis may also be affected , leading to a reduction in its level , as seen in the asa group . previous studies suggest the increase of anti - immunoglobulins cytokines in patients with chronic inflammatory diseases in an attempt to combat inflammatory process . in the aza+asa group , there were no changes in these parameters , which might be associated with better immunological and inflammatory regulation and nutritional status , which were not observed in the asa group , suggesting the effect of azathioprine in uc remission . even so , the crp levels were higher in uc groups compared to controls , while esr was higher only in the asa group , suggesting better regulation of the inflammatory process in the aza+asa group and the effect of azathioprine in this group . inflammatory cytokines such as tnf and il-6 induce hepatic production of crp , which induces expression of adhesion molecules and oxldl uptake by macrophages , favoring the development of atherosclerosis plaque and cardiovascular events . serum nitrite concentration was higher in the uc group compared to matched controls , possibly due to higher inflammatory cytokine levels that also induce inos expression , generating no and peroxynitrite , triggering changes in vascular endothelium , and increasing expression of adhesion molecules and vascular permeability , leading to increased oxidative stress and increased blood pressure [ 3234 ] , which are risk factors for cvd . although the lipid profile was similar , changes in the vascular endothelium in the uc group triggered by the increase of no , crp , and inflammatory cytokines may favor the migration of ldl in the arterial intima and its oxidation , contributing to formation atherosclerosis plaque . the increase of circulating igg anti - oxldl is related to risk of atherosclerosis and suggests that patients with uc have higher levels of oxldl than controls and higher risk of cvd . compared to the control group , the asa group showed increased serum levels of il-6 and ccl-2 , while the aza+asa group has higher levels of tnf , il-6 , ccl-2 , il-10 , and tgf- circulating . the increases in these cytokines may be derived from adipose tissue , colonic mucosa affected by uc , or of circulating inflammatory cells as mononuclear cells [ 3638 ] . il-6 plays an important role in the maintenance of chronic inflammation , preventing apoptosis of lamina propria t cells . il-6 usually binds to its membrane receptor , interacting with the gp80 and gp130 subunits , and activating the stat3 pathway , but sometimes il-6 binds to its soluble receptor and that interact with gp130 on the surface of cells ( especially those not expressing il-6 receptors ) , leading to activation of stat3 . in turn , stat3 promotes upregulation of anti - apoptotic genes , avoiding leukocytes elimination in intestinal mucosa , thus contributing to maintenance of inflammation and release of these mediators to the circulation , even during the remission phase [ 3941 ] . these processes , in turn , may interfere in the cardiovascular system , favoring changes in vascular permeability and ldl oxidation , and leading to development of atherogenesis and its complications . in this regard , we question whether il-6 could interfere with leukocyte survival in atherosclerotic plaque , contributing to its maintenance and progression . il-6 is also capable of reducing the production of no via enos , resulting in endothelial dysfunction as well as increased expression of adhesion molecules and the scavenger receptors cd36 and sr - a1 [ 38,4245 ] . vascular permeability can be altered by these factors , favoring the migration of monocytes to the intima of arteries and the up - take of oxldl , contributing once again to the formation of the foam cells and progression of atherosclerosis . ccl-2 , in turn promotes the recruitment of monocytes , thereby facilitating the migration to different locations , including adipose tissue , colon , and intima of arteries , thereby contributing to the formation of atheroma . although the levels of circulating leukocytes , especially monocytes , were similar between groups , the increased concentration of ccl-2 could induce migration of monocytes , perpetuating inflammation , but keeping monocyte concentration in the normal range in the systemic circulation . tnf were increased only in the aza+asa group , demonstrating higher severity of the disease , which can justify the use of azathioprine in this group . in general , the addition of azathioprine occurs when the patient does not respond to corticosteroids , when the patient is dependent on corticosteroids , or when the patient does not respond to conventional treatment with aminosalicylates , requiring the addition of the drug . tnf induces the secretion of other inflammatory mediators such as il-1 , pcr , and no . in addition , it also increases adhesion molecules and causes endothelial dysfunction , increasing risk of dcv [ 41,4648 ] . only the aza+asa group showed increased production of il-10 and tgf- , which are anti - inflammatory cytokines . il-10 reduces the antigen presentation and activation of inflammatory cells and reduced the activation of nf-b as well as production of inflammatory mediators , contributing to immunology homeostasis , while tgf- participates of the modulation of the activation of endothelial cells and th1 cells , inducing the formation of regulatory t cells   and promoting mucosal intestinal healing . increased levels of these cytokines in patients using azathioprine compared to controls suggest a mechanism of immunoregulation , which was not seen when the asa group was compared to its matched controls . when both uc groups were compared , we noted that the aza+asa group showed increased il-6 , ccl-2 , tnf , il-10 , and tgf- , indicating that although there is increase of inflammatory cytokines , the association of azathioprine and aminosalicylate induces increase of the anti - inflammatory cytokines ( il-10 and tgf- ) , suggesting better immunoregulation in this group , and consequently reducing the risk of atherosclerosis . azathioprine reduces colonic inflammation by blocking rac1 via , resulting in apoptosis of tcd4 + lymphocytes and regulating colonic inflammation . thus , azathioprine appears to modulate t cell repertoire , reducing tcd4 + lymphocytes and preserving the il-10 and tgf- producer regulatory t cells , and promoting immunoregulation of the inflammation related to uc and cvd in this group . although aminosalicylates are associated with reduction of inflammatory cytokines and oxidative stress , the asa group was not able to induce anti - inflammatory mediators , suggesting that this group has a more inflammatory profile that leads to a more susceptible cvd status compared to the aza+asa group . the combination of azathioprine and aminosalicylates changed the profile of immune - inflammatory response in the studied patients , regulating the inflammatory process and reducing the risk of cvd , probably because the drugs have different mechanisms of action that block different inflammatory pathways . however , it is not possible to know whether the isolated use of azathioprine induces immunoregulatory response similar to that observed or whether this effect is only seen when the drugs are combined . the relative risk for cvd , estimated by the framingham risk scores at 10 and 30 years , showed that asa and aza+asa groups had higher cvd risk compared to their controls . although most patients did not show traditional risk factor for cvd , higher risk was probably due to the increase in blood pressure in uc groups . although the framingham risk scores were similar in both uc groups , the aza+asa group showed a better inflammatory profile , which is not included in the framingham score , and which could benefit these patients , improving cvd risk and its complications . we believe that previous use of glucocorticoids associated with sedentary lifestyle and poor dietary habits favor the expansion of adipose tissue . adipose tissue expansion and the inflamed bowel mucosa are factors related to higher production of cytokines and oxidative stress . increased inflammation markers result in changes in the vascular permeability , including increased expression of adhesion molecules and endothelial dysfunction , favoring the development of complications such as increased blood pressure and migration of monocytes to the arterial intima . increased inflammatory cytokines , rcp , and no also induce ldl oxidation , which was indirectly seen by anti - oxldl antibodies . together , this profile favors the development of atherosclerotic plaque , resulting in a higher risk for cvd . in turn , therapy for maintenance of clinical remission affects the inflammatory response , suggesting that the addition of azathioprine to conventional treatment with aminosalicylates decreases the inflammatory process , induces anti - inflammatory mediators , and reduces the risk of cvd in this group ( figure 5 ) . women with uc in clinical remission maintain a more inflammatory profile compared to their healthy controls , suggesting increased risk of atherosclerosis and cvd . the addition of azathioprine to treatment with aminosalicylates reduces inflammation and increases anti - inflammatory cytokines , evidencing better regulation of inflammatory processes and reducing the risk of cvd in this group when compared to that treated with aminosalicylates alone . patients with uc treated in the reference center for inflammatory bowel disease of the alfa institute of gastroenterology . of the total of 144 patients with uc treated at the reference center , 93 were women and 51 were men . among men , 48 were excluded for drug treatment ( use of corticosteroids or biology therapy ) ; 23 ( 24.7% ) were excluded for risk factors or diagnosis of cvd or other diseases ; 8 ( 8.6% ) were excluded by age ; 10 ( 10.7% ) abandoned the treatment , were not found , or did not want to participate in the study , and only 21 ( 22.6% ) women met the study criteria and wanted to participate .