diabetes of long duration results in irreversible functional and structural damage leading to several complications .
diabetes is often associated with severe periodontal disease and is considered a risk factor for periodontal disease progression .
analysis of the national health and nutrition examination survey ( nhanes ) iii data set showed that the prevalence of diabetes in patients with periodontal disease is twice of that seen in non - periodontal disease patients ( 12.5% versus 6.3% ) . in india , up to 70% of the adult population appears to have periodontal disease and in the diabetic , this is estimated to be 82.7% .
periodontal disease is caused by periodontal pathogens such as porphyromonas gingivalis , actinobacillus actinomycetemcomitans , campylobacter rectus etc .
pro - inflammatory cytokines and chemical mediators are significantly increased in gingival inflammation and in periodontal disease progression .
monocyte chemoattractant protein-1 , a chemokine , is implicated in the pathogenesis of numerous complications of diabetes .
the role of mcp-1 has been studied in the pathogenesis of interstitial lung disease , breast cancer cells , rheumatoid arthritis , cytomegalovirus encephalitis , and small cell vasculitis .
the over expression of mcp-1 is found to increase insulin resistance by initiating macrophage infiltration of the adipose tissue and thus predisposing to the onset of diabetes mellitus .
mcp-1 may play a role in the development of numerous complications of diabetes in angiopathy and cause infiltration and activation of monocytes in the glomerulus , which leads to the diabetic nephropathy .
the increased level of mcp-1 in microglia and macrophages leads to the development of diabetic retinopathy .
the role of serum mcp-1 as a biomarker of periodontitis is well documented ; however , its role in diabetic patients with periodontitis is unknown .
this study was conducted to determine the presence and concentration of serum mcp-1 in diabetic patients with and without periodontitis and to correlate it with glycemic status with periodontitis .
a prospective exploratory study was conducted in diabetic patients with and without periodontal disease , attending the out - patient clinic in a tertiary care hospital in sub - urban chennai .
the study protocol was approved by the institution ethics committee , and informed consent was obtained .
sample size was determined based on the number of patients to be enrolled to obtain a meaningful data and was done in consultation with a statistician using convenient sampling .
adult diabetic patients with ( 18 yrs ) with type ii dm who had not received any kind of dental treatment or prophylaxis in the past six months were included and the patients with hypertension , cardiovascular disorders , abnormal hepatic function , hemoglobinopathies , renal failure , retinopathy , pregnancy , patients on current antibiotic treatment or prophylaxis and other endocrine disorders such as hypothyroidism , hypo - parathyroidism , addison 's disease , cushing 's syndrome , and thyroid cancer , which would influence serum mcp-1 levels , were excluded from the study .
patients who were smokers , pan chewers , as well as habitual consumers of alcohol were also excluded .
subjects with hba1c < 7% were considered to have well controlled diabetes and poorly controlled / uncontrolled diabetes if hba1c 7% . fasting blood sugar ( fbs ) level , post - prandial blood sugar ( ppbs ) level , mpg , lipid profile , and routine urine analysis were also recorded . periodontal status of the study population was assessed by oral examination by a qualified dentist .
patients were enrolled for oral hygiene index ( ohi - s ) , evaluation of bleeding on probing ( bop ) , periodontal probing depths ( ppd ) , and clinical attachment loss ( cal ) .
presence of bop , ppd > 5 mm , and cal > 3 mm were considered diagnostic criteria for periodontal disease .
subsequently , the demographic details , general health conditions , oral hygiene behavior , and periodontal evaluation were analyzed .
group i : consists of diabetic patients with good glycemic control with no signs of periodontal disease group ii : consists of well - controlled diabetic patients with evidence of periodontal disease group iii : consists of diabetic patients with poor glycemic control and showed evidence of periodontal disease .
four milliliter of blood was collected in a sterile vacutainer ( bd diagnostics , franklin lakes , usa ) and immediately transferred to the laboratory .
the samples were centrifuged at 3500 rpm for 15 min and were aliquotted and stored at -70c until the time of assay .
serum mcp-1 levels were determined using a quantitative sandwich enzyme - linked immunosorbent assay ( human ccl2 ( mcp-1 ) elisa kit , bioscience , inc . ,
the sensitivity of the assay was 7 pg / ml-1000 pg / ml for mcp-1 .
the kit has been calibrated with national institute for biological standards and control ( nibsc ) ; the co - efficient of variation of this assay is 3.3% .
the absorbance ( od ) was measured at 450 nm by the elisa plate reader ( multiskan ex primary eia v.2.3 ) .
simple linear regression and correlation tests were performed using r statistical software ( version 2.11.0 ) , medcalc software ( version 12.6.1 ) to observe correlation between the serum mcp-1 and glycated hemoglobin level . the age and gender analysis were analyzed only after the completion of the study .
adult diabetic patients with ( 18 yrs ) with type ii dm who had not received any kind of dental treatment or prophylaxis in the past six months were included and the patients with hypertension , cardiovascular disorders , abnormal hepatic function , hemoglobinopathies , renal failure , retinopathy , pregnancy , patients on current antibiotic treatment or prophylaxis and other endocrine disorders such as hypothyroidism , hypo - parathyroidism , addison 's disease , cushing 's syndrome , and thyroid cancer , which would influence serum mcp-1 levels , were excluded from the study .
patients who were smokers , pan chewers , as well as habitual consumers of alcohol were also excluded .
subjects with hba1c < 7% were considered to have well controlled diabetes and poorly controlled / uncontrolled diabetes if hba1c 7% . fasting blood sugar ( fbs ) level , post - prandial blood sugar ( ppbs ) level , mpg , lipid profile , and routine urine analysis were also recorded .
periodontal status of the study population was assessed by oral examination by a qualified dentist .
patients were enrolled for oral hygiene index ( ohi - s ) , evaluation of bleeding on probing ( bop ) , periodontal probing depths ( ppd ) , and clinical attachment loss ( cal ) .
presence of bop , ppd > 5 mm , and cal > 3 mm were considered diagnostic criteria for periodontal disease .
subsequently , the demographic details , general health conditions , oral hygiene behavior , and periodontal evaluation were analyzed .
group i : consists of diabetic patients with good glycemic control with no signs of periodontal disease group ii : consists of well - controlled diabetic patients with evidence of periodontal disease group iii : consists of diabetic patients with poor glycemic control and showed evidence of periodontal disease .
four milliliter of blood was collected in a sterile vacutainer ( bd diagnostics , franklin lakes , usa ) and immediately transferred to the laboratory .
the samples were centrifuged at 3500 rpm for 15 min and were aliquotted and stored at -70c until the time of assay .
serum mcp-1 levels were determined using a quantitative sandwich enzyme - linked immunosorbent assay ( human ccl2 ( mcp-1 ) elisa kit , bioscience , inc . , catalogue number : 88 - 7399 ) .
the sensitivity of the assay was 7 pg / ml-1000 pg / ml for mcp-1 .
the kit has been calibrated with national institute for biological standards and control ( nibsc ) ; the co - efficient of variation of this assay is 3.3% .
the absorbance ( od ) was measured at 450 nm by the elisa plate reader ( multiskan ex primary eia v.2.3 ) .
simple linear regression and correlation tests were performed using r statistical software ( version 2.11.0 ) , medcalc software ( version 12.6.1 ) to observe correlation between the serum mcp-1 and glycated hemoglobin level .
the study group comprised of 37 adult diabetic patients ; majority ( 62% ) were females ( n = 21 ) and 18.9% were males ( n = 16 ) .
both group i and group ii consisted of well - controlled diabetic patients with a mean hba1c of 6.2% and a range of 5.2 - 6.9% . in diabetic patients with poor glycemic control ( group iii ) ,
[ table 1 ] group i patients were younger with a mean age of 26.86 years [ table 1 ] .
descriptive statistics of the study groups showing mean , median , standard deviation , and range for age , glycated hemoglobin ( hba1c ) , and serum mcp-1 levels all 37 diabetic patients serum samples tested positive for the presence of mcp-1 , with concentrations that ranged from 135.6 - 543.5 pg / ml . in diabetics with good glycemic control and no evidence of periodontal disease ( group i ) , the mean serum mcp-1 concentration was 149.3 pg / ml ( range : 135.6 - 160.4 pg / ml ) . in group
ii ( well - controlled diabetes with periodontal disease ) , the mean mcp-1 concentration was 398.8 pg / ml ( range : 330.4 - 435.1 pg / ml ) . in diabetics with poor glycemic control and periodontal disease ( group iii ) , the mean serum mcp-1 concentration was 482.3 pg / ml ( range : 460.9 - 543.5 pg / ml ) .
the mean serum mcp-1 concentration was highest ( 482.3 pg / ml ) in group iii and lowest ( 149.3 pg / ml ) in group i [ table 1 ] .
simple linear regression and correlation analysis performed to observe the relation between glycated hemoglobin ( hba1c ) levels and serum mcp-1 concentrations suggests a linear progression [ figures 1b and c ] in group ii and group iii .
the linear progression was more marked in group iii [ figure 1c ] , and the correlation was highly significant with a p < 0.001 , suggesting that serum mcp-1 levels are markedly elevated as the glycated hemoglobin levels increase in poorly - controlled diabetic patients ( hba1c 7% ) with periodontal disease . in group iii , in three patients with serum mcp-1 concentrations above 530 pg / ml , the corresponding hba1c values were found to be above 12.1% .
( a - c ) correlations of glycated hemoglobin levels ( hba1c ) with serum mcp-1 in groups i , ii , and iii respectively .
the values are not significant in group i and group ii , whereas in group iii , it is significant in group ii [ figure 1b ] , serum mcp-1 concentrations of 320 - 380 pg / ml clustered around hba1c of 6% ; serum mcp-1 was higher > 410 pg / ml when the hba1c levels were in the range of 6.5 - 6.9% .
two subjects with good glycemic control ( hba1c < 6% ) in group ii had elevated levels of serum mcp-1 ( > 410 pg / ml ) , which may be due to the severity of the periodontal disease .
the correlation between hba1c and serum mcp-1 was not significant in group i ( diabetic patients with good glycemic control and no periodontal disease ) [ figure 1a ] .
from the linear regression analysis performed on all study subjects [ figure 2 ] , it was found that a 1% rise of hba1c resulted in 37.278 pg / ml increase of serum mcp-1 concentration , which was statistically significant .
linear regression analysis between hba1c ( % ) and serum mcp-1 ( pg / ml ) in all the study subjects shows a significant intercept value of 37.278 when we analyzed the serum mcp-1 concentrations of each patient in the different groups , we found that the mcp-1 values tend to cluster based on hba1c levels [ figure 3 ] .
mcp-1 values in the range of 350 - 450 pg / ml clustered around group ii ( hba1c < 7% ) . in group iii , the values clustered around 460 - 550 pg / ml .
projection of serum mcp-1 levels in all the three study groups the serum mcp-1 concentrations in diabetic patients with periodontal disease were above 300 pg / ml irrespective of their glycemic status ( group ii and group iii ) .
however , the serum mcp-1 levels were above 460 pg / ml in diabetic patients with poor glycemic control and periodontal disease ( group iii ) , whereas it was only below 450 pg / ml in group ii ( well - controlled diabetic patients with periodontal disease ) .
simple linear regression and correlation analysis performed to observe the relation between glycated hemoglobin ( hba1c ) levels and serum mcp-1 concentrations suggests a linear progression [ figures 1b and c ] in group ii and group iii .
the linear progression was more marked in group iii [ figure 1c ] , and the correlation was highly significant with a p < 0.001 , suggesting that serum mcp-1 levels are markedly elevated as the glycated hemoglobin levels increase in poorly - controlled diabetic patients ( hba1c 7% ) with periodontal disease . in group iii , in three patients with serum mcp-1 concentrations above 530 pg
( a - c ) correlations of glycated hemoglobin levels ( hba1c ) with serum mcp-1 in groups i , ii , and iii respectively .
the values are not significant in group i and group ii , whereas in group iii , it is significant in group ii [ figure 1b ] , serum mcp-1 concentrations of 320 - 380 pg / ml clustered around hba1c of 6% ; serum mcp-1 was higher > 410 pg / ml when the hba1c levels were in the range of 6.5 - 6.9% .
two subjects with good glycemic control ( hba1c < 6% ) in group ii had elevated levels of serum mcp-1 ( > 410 pg / ml ) , which may be due to the severity of the periodontal disease .
the correlation between hba1c and serum mcp-1 was not significant in group i ( diabetic patients with good glycemic control and no periodontal disease ) [ figure 1a ] . from the linear regression analysis performed on all study subjects [ figure 2 ]
, it was found that a 1% rise of hba1c resulted in 37.278 pg / ml increase of serum mcp-1 concentration , which was statistically significant .
( p < 0.001 ) . linear regression analysis between hba1c ( % ) and serum mcp-1 ( pg / ml ) in all the study subjects shows a significant intercept value of 37.278
when we analyzed the serum mcp-1 concentrations of each patient in the different groups , we found that the mcp-1 values tend to cluster based on hba1c levels [ figure 3 ] .
for example , in group i , the serum mcp-1 values clustered below 200 pg / ml . mcp-1 values in the range of 350 - 450 pg / ml clustered around group ii ( hba1c < 7% ) . in group iii ,
projection of serum mcp-1 levels in all the three study groups the serum mcp-1 concentrations in diabetic patients with periodontal disease were above 300 pg / ml irrespective of their glycemic status ( group ii and group iii ) .
however , the serum mcp-1 levels were above 460 pg / ml in diabetic patients with poor glycemic control and periodontal disease ( group iii ) , whereas it was only below 450 pg / ml in group ii ( well - controlled diabetic patients with periodontal disease ) .
the process of tissue destruction ( connective tissue and alveolar bone ) in periodontitis results from the interaction of bacteria or bacterial substances with host cell as well the host response to bacterial invasion .
bacterial plaque is responsible for inducing host inflammatory processes followed by secondary colonization of periodontal pathogens such as porphyromonas gingivalis , actinobacillus actinomycetemcomitans , prevotella intermedia , and others .
the number of monocytes / macrophages are more in progressing periodontal lesions as compared to non - progressing periodontal lesions and may be closely related to the pathogenesis and progression of periodontitis .
mcp-1 is synthesized in inflamed gingiva by mononuclear phagocytes , endothelial cells , and osteoblasts . increased levels of mcp-1
have been reported in gingival biopsies of patients with periodontal disease and in the gingival crevicular fluid ( gcf ) of periodontal patients .
the recovery of periodontal pathogens has been studied in saliva and varying rates of detection tanerella forsythensis ( 56.9% ) , treponema denticola ( 38.2% ) , and porphyromonas gingivalis 35.4% reported .
periodontal disease is considered as potential risk factor for progression of various systemic diseases , also increased levels of serum mcp-1 due to periodontal disease may be a risk factor for systemic diseases .
the association between periodontal disease and cardiovascular diseases is well established ; the role of serum mcp-1 has also been assessed . however ,
the role of mcp-1 in periodontal disease and uncontrolled diabetes mellitus has not been studied . in our study ,
serum mcp-1 levels were raised in diabetic patients with good glycemic control having periodontal disease ( group ii ) when compared with well - controlled diabetic patients without periodontal disease ( group i ) , with even higher levels of serum mcp-1 in diabetic with poor glycemic control having periodontal disease ( group iii ) .
circulating levels of mcp-1 are known to be elevated in diabetic patients and may contribute to the development of complications in diabetics .
diabetic patients with good glycemic control and no periodontal disease ( group i ) were much younger ( mean age of 26.86 yrs ) when compared with group ii and group iii .
the patients were enrolled in to the different group based on their glycemic status , and it was incidental finding that patient with good glycemic controls were younger . aging does increase circulating levels of mcp-1 .
experimental studies on rat model have stated that aging induces expression of mcp-1/ccl-2 receptors leading to age - associated arterial remodeling .
however , age - related mcp changes are gradual and do not increase dramatically as seen in our study .
we believe that mcp-1 differences can not be dismissed because of age alone . when the glycated hemoglobin levels were in a range of 5.2 - 6.9% indicating good glycemic control , the serum mcp-1 concentrations were found to be within acceptable limits ( < 165 pg / ml in group i ) .
the concentration of mcp-1 in serum was increased by 37.278 pg / ml for every 1% rise in hba1c .
regression analysis performed for group iii was found to be significant , suggesting that poor glycemic control contributes to increase in serum mcp-1 levels . even in the absence of any complications such as renal failure , retinopathy , and cardiovascular diseases among the patients enrolled in the study ,
the serum mcp-1 levels clustered into specific ranges even without knowledge of hba1c and periodontal status . based on their glycemic status ,
the mcp-1 values were increased in group i ( 135 -160 pg / ml ) , group ii ( 330 - 435 pg / ml ) , and in group iii .
( 461 - 543 pg / ml ) thus , the determination of serum mcp-1 may be used to assess both the glycemic and periodontal status .
the clustering of serum mcp-1 values among the different groups of patients suggests that there may be threshold levels related to age , glycemic status , and periodontal disease .
if indeed threshold values are established , it will definitely alert the clinician to take measures to pre - empt the progression of diabetes to various complications . to our knowledge , this is the first study to determine serum mcp-1 levels in diabetic patients with periodontal disease and correlate with glycemic control .
this is a prospective exploratory study conducted in 37 diabetic patients , and mcp-1 levels were estimated as a onetime assessment .
prospective studies need to be undertaken with serial estimation of mcp-1 to determine the progression to complications in larger group of patients .
a significant positive correlation between serum mcp-1 and glycemic status ( hba1c ) has been documented .
it has been found that 1% rise of hba1c resulted in 37.278 pg / ml increase of serum mcp-1 concentration .
our findings suggest that glycemic status influences serum mcp-1 levels in diabetic patients with periodontal disease . elevated serum mcp-1 levels could contribute to onset and progression of several complications in diabetes .
thus , serum mcp-1 may serve as a biomarker of inflammatory activity and helps in early detection and intervention of diabetic complications .
 introduction : the role of serum monocyte chemoattractant protein-1 ( mcp-1 ) as a biomarker of periodontitis is well documented ; however , its role in diabetic patients with periodontitis is unknown.aim:this study was conducted to determine the presence and concentration of serum mcp-1 in diabetic patients with and without periodontitis and correlate it glycemic status with periodontitis.materials and methods : adult diabetic patients were enrolled and grouped into group i , ii , and iii based on their glycemic status and serum mcp-1 estimated by elisa . 
 linear regression and correlation tests were performed using r statistical software , medcalc software to observe correlation between the serum mcp-1 and glycated hemoglobin level among different groups.results:serum samples obtained from 37 patients tested positive for mcp-1 . mean serum mcp-1 concentration was highest ( 482.3 pg / ml ) in group iii , lowest ( 149.3 pg / ml ) in group i , and intermediate 398.8 pg / ml in group ii . correlation and regression analysis was done between hba1c and serum mcp-1 . a significant positive correlation ( p < 0.001 ) was observed . 
 serum mcp-1 increased by 37.278 pg / ml for every 1% rise in hba1c , and the levels were raised in group ii and group iii than in group i irrespective of their glycemic status . with an hba1c range of 6.5 - 6.9% ( group ii ) 
 , the serum mcp-1 values cluster around 380 - 410 pg / ml . 
 elevated levels of serum mcp-1 ( > 500 pg / ml ) in three subjects corresponded to hba1c values more than 12.2% ( group iii).conclusion : to our knowledge , this is the first study to document serum mcp-1 levels in diabetic patients with periodontitis . 
 glycemic status influences serum mcp-1 , and lack of glycemic control contributes to increased serum mcp-1 levels . 
 serum mcp-1 may thus serve as a biomarker of inflammation and disease progression in diabetes with periodontitis . 
I
M
Inclusion and exclusion criteria
Assessment of glycemic status
Oral health assessment
Categorization of the study population
Collection of clinical specimens
Determination of MCP-1 concentration in serum
R
Correlation between serum MCP-1 and glycated hemoglobin
Serum MCP-1 levels in different study groups
D
C