Umeå University

Umeå International School of Public Health

Department of Public Health & Clinical Medicine

Association of severe periodontitis with microalbuminuria and

chronic kidney disease

Author: Shabnam Salimi

Umeå, 2010

Supervisors:

Nawi Ng: Umeå University, Sweden

Afshin Parsa: University of Maryland, USA

I

Abbreviation:

AAP American Academy Periodontology

ACR Albumin to Creatinine Ratio

ARIC Atherosclerosis Risk in Communities

BOP Bleeding on Probing

CAL Clinical Attachment Loss

CDC Centers for Disease Control and Prevention

CEJ Cementoenamel junction

CHF Cardiac Heart Failure

CKD Chronic Kidney Disease

COPD Chronic obstructive pulmonary disease

CRP C-Reactive Protein

CVD Cardio Vascular Disease

eGFR Estimated Glomerular Filtration Rate

ESRD End Stage Renal Disease

FGM Free Gingival Margin

HbA1c Hemoglobin type A1c

HTN Hypertension

MEC Mobile Examination Centers

NHANESIII The National Health and Nutrition Examination

Surveys phase III

NIDR National Institute of Dental Research

PD Pocket Depth

PSU Primary Sampling Unit

PPS Probability Proportional to Size

Standardized Serum creatinine Serum creatinine -0.23

USA United State of America

WHR Waist to Hip Ratio

II

Abstract

Background: Chronic kidney disease (CKD) is a major global health burden because of its significant morbidity

and mortality. A significant association between periodontitis, chronic inflammation, and cardiovascular diseases

(CVD) has been demonstrated. The chronic inflammation may be causal in promoting endothelial dysfunction and

associated CVD outcomes. Inflammation and endothelial dysfunction in kidneys can cause damage to the glomerulus,

initially causing the leaking of proteins into the urine, which is clinically identified as microalbuminuria, the early

manifestation of kidney endothelial cell dysfunction which is potentially reversible. This will subsequently lead to the

impairment of kidney function noted by a decrease in the renal glomerular filtration rate (GFR). As such, we

hypothesized that severe periodontal disease, as a common chronic infectious and inflammatory state, is associated

with microalbuminuria, which can be reversible and impaired kidney function.

Objective: To determine the association of severe periodontitis with microalbuminuria and chronic kidney disease

Method: A total of 13,958 participants 20 years old or above were included in this study from NHANESIII 1988-

1994 population based database. An adapted CDC definition was used to define periodontitis. Severe periodontitis was

defined as two or more interproximal sites with ≥6-mm clinical attachment loss (CAL), not on the same tooth, and one

or more interproximal sites with ≥5-mm pocket depth (PD) or complete edentulous. Moderate periodontitis was

defined as two or more interproximal sites with ≥4mm CAL, not on the same tooth, or two or more interproximal sites

with ≥ 5mm PD, not on the same tooth. The association between severe periodontitis and microalbuminuria (ACR 30-

300 mg/g) and CKD (estimated GFR<60ml/min/1,73m2) was analyzed using multivariate logistic regression,

adjusting for demographic, socioeconomic, health behavior, HbA1c, hypertension, serum lead, lipid profiles, and BMI.

Results: Proportion of severe periodontitis among women and men was 10.5% and 9.3 %, respectively. Overall

prevalence of microalbuminuria (30≤ACR<300mg/g) and CKD was 6.7% (7.8% in women and 5.4% in men) and 4.1%,

respectively. There was significant association between CRP as inflammatory factor and microalbuminuria after

adjustment for hypertension, HbA1c and other risk factors. In the multivariate analysis, there was a significant

association between severe periodontitis and both microalbuminuria (OR: 1.32; CI: 1.03-1.70) and CKD (OR=1.62;

CI:1.31-2.01).

Conclusion: Severe periodontitis is, independent of established risk factors, significantly associated with

inflammatory markers, microalbuminuria and CKD. These findings suggest that periodontal disease, potentially

through an inflammatory pathway, may relate to microalbuminuria and progression to CKD. These findings further

add to the plausibility of a biological association between periodontal disease and vascular related disease stated such

as CVD and CKD. Although our findings overall consistent with previous studies and have strong biological

plausibility and evidence of dose response, as a cross sectional analysis, we cannot demonstrate causality regarding

temporality. Considering the high prevalence of periodontal disease and the significant disease burden of CKD, a

longitudinal study to define temporality and prove causality are well warranted. Also, in public health point of view,

considering oral health in primary health care with allocating insurance and treatment of periodontitis is of

paramount importance to prevent microalbuminuria and later chronic kidney disease.

III

Contents

1. Introduction........................................................................ .................. ....................................1

1.1 Anatomy and physiology of Kidney.......................................................................1

1.2 Chronic kidney disease definition ..................................................................... ...1

1.3 Epidemiology of CKD.............................................................................................1

1.4 Pathology of CKD...................................................................................................2

1.5 Indicators of kidney function................................................................................2

1.6 Periodontal disease as a non-traditional risk factor for CKD...............................3

1.6.1 Periodontal disease definition...............................................................................3

1.6.2 Case definition of periodontitis in epidemiologic studies....................................4

1.7 Risk factors for progression of periodontitis........................................................5

1.7.1. Putative risk factors…………………………………………………………………………………..5

1.7.2 Established risk factors.........................................................................................5

1.8 Periodontitis as cause of peripheral vascular disease...........................................6

1.9 Justification of Study.............................................................................................6

2. Hypothesis and Objectives....................................................................................................8

3. Methods......................................................................................................................................9

3.1 Methods applied in NHANES III..........................................................................9

3.2 Definition of applied variables ................................................................. ..........11

3.3 Exclusion criteria and sample size .................................................................... .13

3.4 Analysis and statistical methods ....................................................................... .13

4. Results.......................................................................................................................................15

4.1 Descriptive summary ........................................................................................ .15

4.2 Univariate analysis ............................................................................................ .17

4.3 Multivariate Analysis...........................................................................................22

5. Discussion.................................................................................................................................27

6. Conclusion................................................................................................................................30

7. Acknowledgment.....................................................................................................................31

8. Reference .................................................................................................................................32

1

1. Introduction

Chronic kidney disease (CKD) is a leading of cause of morbidity and mortality in the USA and

the epidemic global increase in end stage renal disease (ESRD) resulted from CKD, along with

the marked increase in associated CVD is recognized as a one of the global public health chal-

lenges. As a result, primary and secondary prevention is crucial and searching for more

comprehensive nontraditional risk factor for CKD can help in more efficient stratification of the

individuals at risk, the development of preventative measures and treatment modalities.1

1.1 Anatomy and physiology of Kidney Kidneys are one of the important organs of the body which are constituted from segmented

nephrons, blood vessels, and filtering capillaries. Kidneys have multiple functions with diverse

mechanisms as excretion of waste products and foreign chemicals, control of blood pressure and

keeping the electrolyte and water balance, regulation of acid-base balance, controlling osmolarity

of body fluid and concentration of electrolytes, secretion, excretion and metabolism of

hormones. 2

The most significant function is filtering plasma and eliminating substances and materials that

are not needed for body. Indeed, the substances contain urea due to amino acids metabolism,

creatinine from muscle creatine, uric acid from nucleic acids and products due to hemoglobin

breakdown like bilirubin and metabolites of the hormones. Eventually, kidneys should remove

these waste materials from the filtrate and blood. 2

The nephrons which are functional unit of kidney produce urine and are not regenerated. Each

nephron is structured by afferent arteriole, bunch of glomerular capillaries named glomerulus

which filter large amounts of fluid from the blood and long tubule that converts the fluid to urine

on the way to collecting duct, pelvis of the kidney efferent arteriole, proximal and distal tubules.

The anatomy of the fenestrated endothelial cells in the glomerular capillary and epithelial cells in

tubular systems are developed to filtrate the renal blood. Afferent arteriole conducts blood to

nephron and then it enters into a glomerular capillary for filtration of most amounts of fluid. The

ratio of glomerular filtration rate (GFR) to renal plasma flow determines the filtration fraction.2

1.2 Chronic kidney disease definition Chronic kidney disease (CKD) is associated with diminished kidney function and progressive

decline in glomerular filtration rate (GFR). In many studies the CKD is defined as estimated GFR

<60ml/min/1.73m2. 2-5

1.3 Epidemiology of CKD Chronic kidney disease is a worldwide major public health problem with its fast-growing number

of patients with end stage renal disease and enhance in mortality and morbidity. It is, also, one

of the main health burdens in the United State. Coresh et al reported that 4.2% of individuals

above 20 year-old had GFR 15 to 59 ml/min per 1.73 m2 in NHANESIII. The prevalence of end-

2

stage renal disease is higher in Japan followed by Taiwan and USA. In USA, the rate of patients

who need treatment is estimated to enhance up to 60% by 2010.3 Likewise, premature death

among people with CKD is three-fold higher than general population. In the USA, a number of

106,912 patients tolerated ESRD during 2005. CKD and following ESRD lead to congestive heart

failure (CHF) and cardiovascular disease (CVD) as a result of increased cardio vascular risk

factors. Some evidence shows that adverse outcomes can be prevented by early diagnosis and

intervention.3, 6, 7

1.4 Pathology of CKD Diabetic mellitus especially type 2, leads to diabetic nephropathy that is the most common cause

of CKD. In addition, hypertensive nephropathy is another common reason of CKD among old

population. The basis of nephrosclerosis caused by vascular disease is the same as which makes

the coronary heart disease and cerebrovascular disease. Atherosclerotic vascular disease among

the old patients is the manifestation of generalized vascular disease. The majority of this

population does not show the last stages of CKD as they die due to cardiovascular and

cerebrovascular diseases. The early stage of CKD is revealed by microalbuminuria and minor

reduction in GFR which in recent studies are shown as a risk factor for CVD.3, 6

Beside all causes, chronic inflammation has been proposed as one of the probable pathologies for

CKD. Regarding some common causes leading to CVD and CKD; some epidemiological studies

demonstrated the chronic inflammation as another potential predictor for CKD and CVD.

Although causal relation between CVD and chronic systemic inflammation has not been proved,

some cohort, nested case control and Meta analysis investigations revealed the link between

inflammation and CVD. Different mechanisms have been suggested about the impact of

inflammation in pathogenesis of CVD. Chronic systematic infection leads to elevated

inflammatory markers level including C reactive protein (CRP), interleukin-6, haptoglobin and

fibrinogen.5 In some studies it is reported that elevated CRP damaged vascular endothelium

leading to microalbuminuria. Thus, microalbuminuria is recognized as the laboratory indicator

of damage to the endothelial cells. In some epidemiologic studies microalbuminuria is a risk

factor for cardiovascular disease. Based on these evidences, microalbuminuria is the early

manifestation of renal damage due to endothelial dysfunction before decrease in GFR which

later leads to macroalbuminuria and more advanced kidney failure manifested by decreased

GFR.6-14

1.5 Indicators of kidney function CKD at the early stage is manifested with microalbuminuria which later in extended stages leads

to macroalbuminuria. Albuminuria is indicated by urinary albumin-to-creatinine ratio (ACR)

which is reported in mg/g. Microalbuminuria is defined as ACR > 30 mg/g to less than 300

mg/g and individuals with ACR ≥300mg/g are classified as having macroalbuminuria.2,3

3

The recommendation of National Kidney Foundation and American Society of Nephrology is to

use creatinine to estimate GFR (eGFR), within clinical settings. Among the young the normal

range of GFR is 120-130 mL/min/1.73m2 which decreases with age. Most common sequels of

CKD occurs with a GFR <60 mL/min/1.73m2. 2-4

1.6 Periodontal disease as a non-traditional risk factor for CKD For long time, diabetic mellitus, hypertension and atherosclerosis diseases has been known as

risk factors of kidney dysfunction. Based on new studies, chronic inflammation is speculated as

risk factor for impairment of kidney function. Periodontitis, as longtime inflammation is posed as

one of the non-traditional risk factors.

1.6.1 Periodontal disease definition

Periodontal disease is the chronic inflammation of the tissues supporting tooth and one of the

most prevalent oral diseases. It is occurs due to the interaction of gram negative bacteria and the

host inflammatory response.15,16 With the extension of inflammation and chronic irritation, there

would be loss of connective tissue fibers and ligaments that attach gum to the teeth and alveolar

bone near to the tooth. With the progression of inflammation to deeper structure, there would

be destruction of the collagen and connective tissue and loss of alveolar bone (attachment loss)

which eventually lead to loss of tooth. 9, 17

With degeneration of junctional epithelium, a “pocket” epithelium which proliferate apically and

laterally and ultimately a true pocket is formed which is suitable for opportunistic bacteria.17

Pocket depth (PD), the distance from the margin of gingival, is an index to find current

periodontitis. In practice, clinical attachment loss (CAL) is one of the methods to define previous

and long term periodontitis. CAL is the distance from cement-enamel junction to the base of the

sulcus or periodontal pocket.9, 17

The plaque produced by bacteria in the oral cavity it finally calcifies and alters to calculus (Fig1).

Calculus is a trap which enhances the growth of the constructed plaque and irritates gingival

more. Then it is known as secondary aetiology of periodontitis as primary one is the plaque

produced by bacteria. 17

Calculus can develop along the gum line or within the narrow sulcus between the teeth and the

gingival, which are called supragingival and subgingival respectively. Calculus can result in

chronic gingivitis and eventually periodontitis.

Almost all individuals with periodontitis demonstrate substantial subgingival calculus. So,

Calculus is another index to diagnose periodontitis. 17

4

Fig 1. Calculus: Adapted from color atlas of dental hygiene17

A.

B. Fig2. Periodontal tissues and disease destruction. (A) A tooth with the periodontal tissues (cementum, connective tissue, and alveolar bone), healthy on the left side and on the right side compromised due to periodontal disease. The arrows indicate the corresponding areas. (B) Radiograph of molar teeth. Local destruction supporting alveolar bone due to periodontal disease is demonstrated by the right arrow, while bone levels are shown to be optimal by the left arrow9

1.6.2 Case definition of periodontitis in epidemiologic studies

There are various definitions based on different criteria to define periodontitis, yet there is no

unique agreement on a standard definition which is required as case definition implementing in

population-based studies to allow data interpretation in diverse global epidemiologic studies.18, 19

It is important to know that periodontitis which has acute and chronic components cannot be

measured merely by a single variable. When trying to capture the lifetime periodontitis burden,

attachment loss (CAL), appears to be the most reliable. As a result, this measure should be a

primary variable utilized in studies of risk factors for chronic periodontitis. To evaluate a risk

factor on current status of periodontitis, combination of attachment loss and bleeding on

probing depth (BOP) and Pocket Depth is acceptable. 11 In 2003, the Division of Oral Health at

the Centers for Disease Control and Prevention (CDC) in collaboration with American Academy

Periodontology (AAP) reviewed various conducted studies to develop a standardized clinical

definition for periodontitis in epidemiologic studies.19-21

Based on the Page R C and Eke P I study, the definition applied to NHANESIII is ≥ 1 tooth with

CAL ≥3mm + PD≥ 4 mm at the same site. In NHANES 1999-2000 the case definition of

periodontitis is ≥3 sites with CAL ≥4mm +≥ 2sites with PD ≥3mm. 18

Likewise, several definitions of periodontitis have been utilized in national survey in USA. Two

definitions are provided based on CDC, one for severe periodontitis and another for moderate

periodontitis. The case definition for severe periodontitis requires two or more interproximal

sites with ≥6-mm CAL, not on the same tooth, and one or more interproximal sites with ≥5-mm

PD. Moderate periodontitis was defined as two or more interproximal sites with ≥4-mm CAL,

not on the same tooth, or two or more interproximal sites with ≥5-mm PD, not on the same

5

tooth. Also, the radiographic pattern and extent of alveolar bone loss and gingival inflammation

measured as bleeding on probing, or a combination of these measures is applied for the

definition. Although, some of these measures have been used in surveys (for example, PD and

CAL), others are not suitable in population based surveys, for example, radiographic

measurements.22

Measurement at interproximal sites is important because, unlike buccal or lingual sites,

periodontitis usually commence at these sites and is most severe at interproximal sites.

Moreover, the effects of gingival recession on accuracy of the PD measurements are minimized.

At least two sites with CAL ≥6 mm, not on the same tooth, are demanded since it is probable to

have abnormal CAL without having periodontitis. For many reasons both CAL and PD indexes

are necessary. CAL is considered as the gold standard for diagnosing the severity of the disease

and reasonably considered a more accurate measure than PD. Utilizing only CAL, however, could

mistakenly include some periodontal healthy sites since attachment loss can come along with

gingival recession which is non-inflammatory status. Gingival recession can take place when a

tooth with periodontitis is treated successfully or the disease resolved by itself without returning

to normal status. Furthermore, attachment loss can be due to non-inflammatory causes.22, 23

In ARIC study CAL was measured in 6 tooth sites to increase the accuracy of measurement. The

periodontitis was determined in three various levels regarding the degree of periodontitis as: 1)

severe periodontitis: 2 or more interproximal sites (not on the same tooth) with a clinical

attachment level of 6 mm or greater and 1 or more pocket depths of 5 mm or greater; 2) initial

periodontitis: 2 or more interproximal sites with a 4-mm or greater clinical attachment level (not

on the same tooth); and 3) healthy/gingivitis: individuals who did not meet criteria 1 and 2. 24

1.7 Risk factors for progression of periodontitis

1.7.1 Putative risk factors

Periodontitis is often associated with age, gender, obesity, metabolic syndrome, dental hygiene,

socioeconomic status, race/ethnicity and probable genetic factors.

1.7.2 Established risk factors

- Specific Plaque, bacteria

- Environmental factors

There is excessive inflammatory response resulting in more severe periodontitis in smokers.

Tobacco smoking is a strong predicting factor for periodontitis in some longitudinal studies.

Moreover, it is manifested that quitting smoking shows less tooth loss and lower incidence of

periodontitis. Cigarettes smoke impact on attachment to root surface of tooth.19

6

Diabetes mellitus is another risk factor for progression of periodontitis. In addition, high

association between poor metabolic control (HbA1c ≥7%) and periodontitis among patients with

diabetic mellitus is reported.19

1.8 Periodontitis as cause of peripheral vascular disease In recent decade many studies manifested the association between periodontitis and systemic

diseases. Especially, some studies showed significant link between cardiovascular disease (CVD)

and periodontal disease.24 Likewise, some investigations showed the role of periodontitis in

other systemic disease enclosing stroke, diabetes mellitus and pre-term pregnancy.25, 26

1.9. Justification of Study Kidney disease is associated with significant economic and health burden and is increasing in

prevalence and chronic kidney disease (CKD) has a global prevalence of 7% which is increasing

at an alarming rate.

microalbuminuria as an early marker of renal dysfunction is similarly associated with CVD

morbidity and mortality. On the other hand, inflammation appears to be a common mediator

between renal disease progression and CVD outcomes. Also, periodontal disease is the most

prevalent cause of chronic inflammation and has been associated with CVD outcomes and with

CKD.

Inflammation has a vital role in the pathologic pathway inducing endothelial damage, and

vascular dysfunction. In some studies even mild systemic inflammation is associated with

cardiovascular disease, although the causality has not been established so far. Periodontitis as a

chronic systemic inflammation affects on the tissues around teeth leading to loss of attachment

and increasing both local and systemic inflammation and associated markers such as serum

levels of CRP and fibrinogen. Such inflammation can also cause systemic endothelial

dysfunction. In Kidney tissue, inflammatory factors impact on the glomerular endothelium

leading to micro and then macroalbuminuria, which are early manifestation of kidney

dysfunction, compared to decreased GFR.

Some studies evaluated the predictor values of such inflammatory disease as periodontal disease

in GFR and also macroalbuminuria in last stages of kidney function. Since microalbuminuria, is

an early manifestation that is readily measurable and can be reversible by early treatment, as

shown in previous studies in diabetic nephropathy, it is paramount of usefulness in early

diagnosis of nephropathy. As such, we decided to evaluate the association between periodontal

diseases and microalbuminuria in NHANESIII. We also wish to further explore the association

between periodontal disease and established CKD, as a marker of sever disease.

Specifically, using the NHANESIII survey, we plan to examine the association between CDC

classification schemes for periodontal disease with established measures of renal function. These

measures will include microalbuminuria and creatinine based measures of estimated glomerular

7

filtration rate (eGFR). For our evaluation of periodontal disease we used measures

encompassing, chronic exposure and disease burden, as opposed to measures of acute

periodontitis, since they may not be as relevant to the long-term development of vascular related

disease, such as CKD.

Also in the previous studies in NHANESIII, the applied definitions for periodontitis did not

include pocket depth, in definition of periodontitis. Our primary interest is accumulated disease

burden. Despite bleeding, gingivitis and probing depth indicating current disease, attachment

loss measurement manifests the accumulated disease burden and not accurately current disease

activity.23 Measurement at interproximal sites is vital because, periodontitis usually begins at

these sites and is most severe at interproximal sites. Moreover, the effects of gingival recession

on accuracy of the PD measurements are minimized. Yet, in previous studies minimizing the

influence of gingival recession in accuracy of measurement was not taken into account. Thus, in

this study, we include only interproximal site which is mesiobuccal, as only mesiobuccal sites

were measured in NHANESIII.

Since there is no unique definition for population-based surveillance of periodontitis, we decided

to choose the employed epidemiologic definition by CDC-AAP for periodontitis and include

edentulous status as a criteria of sever periodontitis.

We will also contrast the effect of periodontal disease classification and case definition on the

related association with the existing established renal disease classifications, which include

microalbuminuria as early manifestation of kidney function and eGFR <60 mL/min/1.73m2 as

the standard measure of CKD case definition.

8

2. Hypothesis and Objectives

We hypothesized that severe periodontal disease is associated with microalbuminuria and

chronic kidney disease.

The study objectives are:

Objective 1: To study the association between severe periodontal disease and

microalbuminuria.

Objective 2: To study the association between severe periodontal disease and established chronic kidney disease.

9

3. Methods 3.1 Methods applied in NHANESIII

Study design

The cross sectional National Health and Nutrition Examination Survey (NHANESIII) carried out

by National Center for Health Statistics from 1988 to 1994. The study was conducted in multi-

stage, stratified clustered probability sample of non-institutionalized civilian US residents of US.

This multi-stage cluster sampling was conducted in various stages:

The sample represented the total civilian, noninstitutionalized population, in the 50 states and

the District of Columbia of US. The first stage of the design comprised of selecting a sample of

81 PSU’s (primary sampling unit) that were mostly individual counties. In a few cases,

adjacent counties were combined to keep PSU's above a minimum population size. Then, the

PSU's were stratified and selected with probability proportional to size (PPS). Thirteen large

counties (strata) were chosen with certainty (probability of one). For practical reasons, these

13 certainty PSU's were divided into 21 survey locations. After the 13 certainty strata were

selected, the remaining PSU's in the United States were grouped into 34 strata, and two PSU's

were selected per stratum (68 survey locations). The selection was done with PPS and without

replacement. Thus, the NHANES III sample is composed of 81 PSU's or 89 locations.

The 89 locations were divided into two groups randomly, one for each phase. The first group

included 44 and the other 45 locations. One set of PSU's was designated to the first three-year

survey period (1988-91) and the other allocated to the second three-year period (1991-94).

For most of the sample, the second stage of the design included area segments composed of city

or suburban blocks, combinations of blocks, or other area segments in places where block

statistics were not produced in the 1980 Census. In the first phase of NHANES III, the area

segments were used only for a sample of persons who lived in housing units built before 1980.

For units built in 1980 and later, the second stage consisted of sets of addresses selected from

building permits issued in 1980 or later. These are referred to as "new construction segments."

In the second phase, 1990 Census data and maps were used to define the area segments.

The third stage of sample design comprised of households and particular types of group

quarters, such as dormitories. All households and eligible group quarters in the sample segments

were listed, and a subsample was designated for screening to identify potential sample persons.

The subsampling rates enabled production of a national, approximately equal-probability

sample of households in most of the United States with higher rates for the geographic strata

with high Mexican-American populations. Within each geographic stratum, there was a nearly

equal-probability sample of households across all 89 stands.

The fourth stage of sample selection included persons within the sample of households or group

quarters. All eligible individuals within a household were listed, and a subsample of them was

selected based on sex, age, and race or ethnicity. The definitions of the sex, age, race or ethnic

10

classes, subsampling rates, and selection of potential sample persons within screened

households were developed to provide approximately self-weighting samples for each sub

domain within geographic strata and at the same time to maximize the average number of

sample persons per sample household. Based on previous NHANES studies, this increased the

overall participation rate.

Data Collection in NHANESIII

Data for NHANES III were collected through two phases. The data was collected through

interview, examination and laboratory tests.

In household interviews the individuals are asked about demographic, socioeconomic, and

health-related questions; and this followed by the extensive physical examinations, dental

examinations, health and dietary interviews, and laboratory tests which were conducted in

mobile examination centers (MECs).

Household Interviews

The interviewers performed all interviews in household level and then scheduled certain

appointments for examinations which were conducted in the MEC.

Advance Letter, Confidentiality and informed consent form

Before interview, a letter was sent to the household which explained the purpose of study and

why it is important and the participant’s right comprising confidentiality of information taken

and informs them that the interviewer will come to the household. In the first interview only

verbal consent was enough but to be included in examination in MEC signed consent form was

necessary to show the willingness of the participant in the survey otherwise the participant was

excluded due to refusal.27

Household Interviews and examination in Mobile Examination Center (MEC)

Interview

During interview demographic, socioeconomic, environmental data and history of diseases were

assessed by a designed questionnaire.

Exam Team Responsibilities

One physician reviewed the participant’s medical history, managed the medical examination

and recorded the results of examination and blood tests.

For measuring blood pressure, a mercury sphygmomanometer was utilized based on the

standardized blood pressure measurement protocols recommended by the American Heart

Association.28

One expert dentist conducted the dental exam and a health technician recorded the dentist’s

exam findings. Three medical technicians carried out urine and blood tests then recorded the

results and prepared and shipped specimens to different laboratories for more specific tests.

11

Dental Examination

In NHANESIII, not everybody received dental examination. In oral examination those who had

medical problems examination were excluded. The list of these health problems asked at the first

questionnaire at the beginning of oral exam was heart problems like congenital heart murmurs,

heart valve problems, congenital heart disease, and bacterial endocarditis, Rheumatic fever,

kidney disease requiring dialysis, hemophilia, having pacemaker or other artificial material in

heart veins or arteries, hip, joint or bone replacement.29 Also participants with age less than 20

years were excluded.

In NHANES III, only two quadrants were examined randomly and a computer program selected

the random quadrant to evaluate the periodontal status. Buccal and mesiobuccal sites of two

quadrants were assessed.

Periodontal Destruction

Examination of the periodontal status included loss of attachment and periodontal pocket depth.

The examination included only permanent full erupted teeth. Only two sites were assessed:

buccal and mesiobuccal (interproximal ).

- Clinical Attachment Loss (CAL): Clinical attachment loss is the distance from the

cementoenamel junction (CEJ) to bottom of the pocket in millimeters (mm).

- Pocket depth (PD): Pocket depth is the distance from the free gingival margin (FGM)

to the bottom of the sulcus

The NIDR periodontal probe was used to measure the buccal and mesiobuccal sites. For each

site, first, the distance from the free gingival margin (FGM) to the CEJ, and second, the distance

from the FGM to the bottom of the pocket was measured.

3.2 Definition of applied variables The designated variables encompassing outcome of interest, independent variables including

sociodemographic status, life styles, health behaviour and laboratory assays are summarized in

table 1.

12

Table 1. Studied variables including main outcomes and independent variables.

Variables Definitions and categories

Outcomes variables

Albumin Creatinine Ratio (ACR) Normal: ACR<30 mg/g(Reference),Microalbuminuria:Albumin Creatinine Ratio ≥30 mg/g and <300mg/g, macroalbuminuria: ACR≥300

Chronic Kidney Disease status Normal:eGFR≥60ml/min/1.73m2(Reference);CKD=eGFR<60

ml/min/1.73m2

Independent variables

Main risk factors of interest

Periodontitis* (non-traditional risk

factor)

Severe periodontitis: Two or more interproximal sites with ≥6-mm

CAL, not on the same tooth, and one or more interproximal sites with ≥5-mm PD or complete edentulous, Moderate: Two or more interproximal sites with ≥4-mm CAL, not on the same tooth, or two or more interproximal sites with ≥5-mm PD, not on the same

tooth. Mild or healthy: Non severe nor moderate (Reference)

Edentulous Loss of all natural teeth

Socio-demographic variables

Age 20-59 (Reference) ; ≥60 years

Gender Man(Reference) , Women

Marital Status Married(Reference), Widowed, divorced, single

Race ethnicity Non Hispanic white, Non Hispanic Black, Mexican-American or other(Reference)

Country of Birth US(Reference), Non US

Education >12 years (Reference) , ≤12 years

Last visit by doctor ≤1year (Reference), >1 year

Last visit by dentist ≤1 years (Reference), >1 years

Annual income ≥20000$ (Reference), <20000$

Smoking** (traditional risk factor) Current smoker1, Former smoker2, non-smoker3 (Reference)

Health variables

Hypertension(traditional risk factor )

Average Systolic blood pressure≥140mmHg or Diastolic blood pressure ≥90 or currently taking prescription to control hypertension

Heart disease History of heart attack, congestive heart failure, angina or stroke

WHR ≤ 0.8 for women and ≤ 1.0 for men (Reference);≥0.8 if men ,≥1.0

if women

Laboratory assays

CRP (C- Reaction Protein) CRP: <0.22 mg/dl(Reference) ,0.22≤CRP≤1.0mg/dl, >1mg/dl

Fibrinogen <450 mg/dl (Reference); ≥450 mg/dl

HbA1c(GlycosylatedHemoglubine) <7% (Reference); ≥7%

Total Cholesterol <240mg/dl(Reference); ≥240mg/dl

HDL** ≥35mg/dl(Reference); <35mg/dl

Serum Lead <10 ug/dl(Reference); ≥10 ug/dl

Standardizedserum creatinine Serum creatinine-0.23 (mg/dl) to calculate GFR27,29,30

*Severe periodontitis in this study is determined regarding CDC definition for severe periodontitis or

complete edentulous 1. Positive answer to “Have you smoked at least 100 cigarettes in lifetime” and

“Do you smoke now” or serum cotinine ≥15 ng/ml 2. Positive answer to “Have you smoked at least

100 cigarettes in lifetime” 3. Neither former nor current smoker, ** High density lipoprotein

13

3.3 Exclusion criteria and sample size

The following exclusion criteria were used in the selection of study subjects from the NHANES

III database:

1. Age less than 20 year-old

2. Currently pregnant

3. Currently in menstruation period

4. Individuals with no serum creatinine measurement

5. Medical problems with contradiction to having oral examination.

After considering exclusion criteria, 13,958 participant’s data were included in this study.

3.4 Analysis and statistical methods NHANESIII is a complex multistage, stratified and clustered study. Due to the non-random

selection of subjects, sample weights were applied in the analysis. Sample weights in NHANES

III were utilized to achieve the following objectives:

- To compensate for different probabilities of selection among subgroups (age-sex-race-

ethnicity subdomains; persons living in different geographic strata sampled at different

rates)

- To reduce biases arising from the fact that nonrespondents may be different from the

participants.

Weighting took into account several aspects of the study: the specific probabilities of selection

for the oversampled individual domains, as well as nonresponse and differences between the

sample and the total population. Differences between the sample and the population may appear

due to sampling variability, different coverage of demographic groups, and perhaps other types

of response errors, such as differential response rates or misclassification errors.

According to the NHANESIII analysis guideline, MEC and home examination weights was more

suitable as weighting variable .27 The weighting was based on the probability of selection of each

case which was depends on three factors: 1) the person’s age-sex-race/ethnicity domain; 2) the

density stratum; and 3) the PSU. The following provides a brief description of each of the three

components.

Older persons, children, Mexican-Americans, and black individuals were oversampled to insure

a minimum sample size for each analytic domain so that estimates of the health status of the

participants in each domain could be made with satisfactory precision. The oversampling in

NHANES III was part of a pattern established in the sample design. The population was

decomposed into 52 subdomains: 7 age groups by sex for black and Mexican-American persons

and 12 age groups by sex for white persons and other racial groups combined. After defining age-

sex-race/ethnicity subdomains, variable sampling rates were extracted to ensure the appropriate

sample sizes to allow analyses of the data for each subdomain.

14

Also, the density strata were considered by dividing the census blocks (or counting districts) in

each sampled PSU into six classes with each class having a different level of concentration of

Mexican-American individuals. Blocks with high concentrations of Mexican-American persons

were oversampled to increase the sample produced for this group. In addition, for the PSU

factor, it was introduced to adjust the basic weights to reflect the effect of the relatively fixed

sample size within each PSU in NHANES III on the sample weights. The reason for the relatively

fixed sample size by PSU was to have a manageable and efficient field procedure. However, the

use of nearly a fixed number of examinations per PSU implied that NHANES III would not

consist of exact self-weighting samples. The more details of weighting is explained in

NHANESIII guide line.27

The raw data from NHANESIII data base transferred to STATA 8 with STAT transfer program 9.

To test the hypothesis of association between periodontitis and traditional and non- traditional

risk factors with microalbuminuria and CKD, univariable analysis was employed. Afterwards,

backward stepwise multivariable logistic regression model was deployed to take into account the

statistically significatnt risk factors recognized in the univariable analysis. The 95% confidence

interval (CI) was used to assess statistical significance. We identified the significant risk factors

for microalbuminuria and CKD in univariable model. The OR derived in the multivariable

logistic regression model were applied to estimate the odds of decreased eGFR(CKD) or

microalbuminuria among those with risk factors. As there was strong association between

HbA1c and history of diabetes, just HbA1c was included in analysis to prevent collinearity.

Furthermore, HbA1c data was well captured whereas there was more missing data related to the

“history of diabetes” variable.

15

4. Results

4.1 Descriptive summary Table1 shows the main characteristics of the men and women who participated in the study. One

fourth of the women were over the age of 60. Almost one third of women had a lower level of

education, while in men higher education than 12 years had the same proportion as lower level

(p<0.005). Regarding marital status, the majority were married in both men and women

(p<0.005). Both sex had equal distribution with respect to country of birth. (Table 2) Table2. Description of demographic data

Men n=6,878(51%)

Women n=7080(49%)

Age

20-59

60+

81.2%

18.8%

74.8%

25.2%

Ethnicity Mexican- American &others Non-Hispanic white

Non-Hispanic black

13.3% 77.2%

9.5%

12.8% 75.7%

11.5%

Marital status Married Widowed Divorced Single

71.0% 2.3% 7.7% 19.0%

61.0% 12.0% 13.7% 13.3%

Education >12 years ≤12 years

55.5% 44.5%

62.0% 38.0%

Country of Birth USA Non USA

85.7% 14.3%

86.3% 13.7%

Income ≥20000$

<20000$

71.0%

29.0%

63.4%

36.6%

About 8% of women showed microalbuminuria but above 5% of men experienced

microalbuminuria while macroalbuminuria had higher proportion among men compared to

women (p<0.005). Moreover, chronic kidney disease was 5.1% among women Vs 3.2% in men

(p<0.005). Although moderate periodontitis had higher proportion among men, more women

endured severe periodontitis (p<0.005). Almost half of women never smoked in their life time

while 35% of men were current smoker. Furthermore, distribution of hypertension, control of

diabetes and history of cardiovascular disease had similar pattern in both sex (p>0.05). Less

than one fifth of men experienced serum lead more than 10 ug/dl. Also, three forth of women

had WHR≥0.8 which was almost three times more than WHR≥1.0 in men (p<0.005). The

proportion of other risk factors is summarized in table 3. Among the participants without CKD

(eGFR>60), 23% of the study subject experienced microalbuminuria and 8.6% had

macroalbuminuria.

16

Table.3 Distribution of outcomes and risk factors in men and women Men¥ Women¥

Albuminuriaᶲ

No(ACR<30) Micro (30≤ACR<300) Macro(≥300)

93.4% 5.4%

1.2%

91.3% 7.8%

0.9%

CKDᶲ

eGFR≥60 eGFR<60

96.8% 3.2%

94.9% 5.1%

Periodontitisᶲ

No/Mild moderate Severe or Edentulous

81.0% 9.7% 9.3%

83.0% 6.5% 10.5%

Smoking ᶲ

Never smoked

Former smoker

Current smoker

35.6%

28.7% 35.7%

53.6%

19.3% 27.1%

Hypertension No Yes

76.7% 23.3%

76.0% 24.0%

CVDᶲ

No Yes

90.0% 10.0%

91.7% 8.3%

BMI <25 25-30 >30

40.6% 40.0% 19.4%

48.4% 26.2% 25.4%

WHRᶲ

<0.8if men ,<1.0 if women

≥0.8 if men ,≥1.0 if women

73.0%

27.0%

24.0%

76.0%

Last visit by dentistᶲ

< 1 years ≥1year

62.8% 38.2%

68.0% 32.0%

Last visit by doctor ᶲ

<1 years ≥1year

73.0% 27.0%

87.8% 12.2%

HbA1c HbA1c<7 % HbA1c≥7 %

96.3% 3.7%

96.0% 4.0%

CRPᶲ

<0.22 ≥0.22 &<1.0 ≥1.0

78.0%

17.5% 4.5%

65.7%

25.4% 8.9%

Fibrinogen

<450 ≥450

95.0% 5.0%

94.4% 5.7%

Total Cholesterolᶲ (mg/dl)

<240 ≥240

83.4% 16.6%

78.7% 21.3%

HDLᶲ (mg/dl)

≥35 <35

81.8%

18.2%

94.0%

6.0%

Leadᶲ (ug/dl)

<10 ≥10

94.8% 5.2%

98.9% 1.1%

¥ .Weighted percent, ᶲp<0.05

17

The prevalence of macroalbuminuria was higher in people with poor kidney function (eGFR<60)

as compared to people with better kidney function eGFR >60 (p<0.05). As microalbuminuria is

the early manifestation of kidney endothelial cell dysfunction which is potentially reversible,

hence, in univariate and multivariate analysis, microalbuminuria (30≤ACR<300) was compared

against no albuminuria (ACR<30 mg/g).

4.2 Univariate analysis Univariable logistic regression models (OR crude), illustrated in Tables 3 and 4, demonstrate the

unadjusted association between single predictor variables and the outcome binary ACR and CKD

categories. The percentages in the second and third columns in Table 3 and 4 indicate the

weighted proportion.

The prevalence of microalbuminuria among women was significantly higher than men while

CKD had higher proportion in men compared to women. The likelihood of having CKD in Non-

Hispanic white was 2.47 times more than Mexican-American and other ethnicities. (Table 5)

Among widowed group less than one fifth had microalbuminuria and proportion of CKD was the

same in this group while single individuals were less probable to have microalbuminuria or CKD.

Country of birth was not significant risk factor for microalbuminuria whereas those who were

born in non USA countries were less likely to suffer from CKD. The odds of microalbuminuria

were 50% higher among those for whom it took more than one year to be visited by a dentist. In

contrast, microalbuminuria was more frequent among those visited a doctor in less than one

year compared to those who so do not. (Table 4 and Table 5)

About one third of the individuals with HbA1c >7% had microalbuminuria while it was 15%

among hypertensive individuals. In addition, 15% of the participants with severe periodontitis

manifested microalbuminuria. The prevalence of CKD in the hypertensive individuals, those

with HbA1c>7% and the participants with sever periodontitis was above one tenth, 12.2%, 14.05

and 13.7%, respectively. The prevalence of microalbuminuria and CKD regarding other risk

factors is summarized in Table 4 and 5.

In the univariate logistic regression model, considering periodontitis as non-traditional risk

factor of microalbuminuria, severe periodontitis or edentulous had almost three times higher

probability of experiencing microalbuminuria compared to no/mild periodontitis while those

with moderate periodontitis had about two times higher likelihood to have microalbuminuria.

The probability of having microalbuminuria among women was higher by 48% compared to

men. Similarly, in the elder, microalbuminuria was 3.51 times more common than the younger.

There was no significant association between non-Hispanic black and microalbuminuria.

Although being Non-Hispanic white sounds protective factor against microalbuminuria, it was

not significant. Regarding marital status, single condition looked protective factor against

microalbuminuria while widowed individuals were more probable to have microalbuminuria

compared to married status.

18

Table4: The Univariate association between the other risk factors and microalbuminuria

Risk Factors ACR <30 mg/g (93.3%)

30≤ACR<300mg/g (6.7%)

OR Crude*

(95% CI)

Periodontitis No/Mild

Moderate Severe or Edentulous

94.5%

90.2% 85.3%

5.5%

9.8% 14.7%

1.00

1.90(1.50-2.50) 3.00(2.30-3.97)

Sex Men Women

94.5% 92.1%

5.5% 7.9%

1.00

1.48 (1.15- 1.93)

Age 20-59 60+

95.4% 85.4%

4.6% 14.6%

1.00

3.51(2.86- 4.31)

Ethnicity Mexican-American or others

Non-Hispanic white Non-Hispanic black

92.9%

93.7% 91.2%

7.1%

6.3% 8.8%

1.00

0.87(0.61- 1.25) 1.25(0.88- 1.76)

Marital status Married Widowed

Divorced Single

94.0% 80.3%

92.2% 95.2%

6.0% 19.7%

7.8% 4.8%

1.00

3.81(2.98- 4.90)

1.20(0.84- 1.65) 0.67(0.50-0.90)

Education >12years ≤12 years

95.2% 92.0%

4.8% 8.0%

1.00

1.70(1.35- 2.16)

Country of Birth USA Non USA

93.4% 93.1 %

6.6% 6.9%

1.00

1.04(0.72- 1.50)

Income

≥20000$

<20000$

94.6%

90.9%

5.4%

9.1%

1.00

1.74(1.43-2.12)

Smoking Never smoked Former smoker Current smoker

93.9% 91.8% 93.6%

6.1% 8.2% 6.4%

1.00

1.36(1.08- 1.73) 1.04(0.79- 1.36)

Hypertension No Yes

95.9% 84.5%

4.1% 15.5%

1.00

4.28(3.42- 5.36)

CVD No

Yes

93.7%

88.8%

6.3%

11.2%

1.00

1.88(1.47- 2.41)

BMI <25 25-30

>30

94.6% 93.0%

89.3%

5.4% 7.0%

10.7%

1.00

1.32(1.02- 1.72)

1.70(1.34- 2.17)

WHR <0.8if men ,<1.0 if women ≥0.8 if men ,≥1.0 if women

95.8% 91.4%

4.2% 8.6%

1.00

2.17(1.74- 2.70)

Last visit by dentist <1 years ≥1year

94.3% 91.6%

5.7% 8.4%

1.00

1.51(1.17- 1.95)

Last visit by doctor <1 years

≥1year

92.8%

95.5%

7.2%

4.5%

1.00

0.61 (0.46-0 .81)

19

Table 4(continue)

* OR Crude = Unadjusted odds ratio for the association between chronic kidney disease and the

suspected/recognized risk factors in sample

The likelihood of microalbuminuria was 1.70 times higher in individuals with lower level of

education, as compared to those with higher level of education, and 1.74 times higher in

individuals with low income, as compared to those with higher income.

Shorter period till being visited by dentist were protective factors to have microalbuminuria

whereas longer time to see a doctor or health professional was posed as a protective factor.

Individuals with hypertension, HbA1c>7% and CVD as traditional risk factors had significant

association with microalbuminuria.

Likewise, inflammatory factors encompassing CRP and fibrinogen were significant risk factors

for having microalbuminuria. That is, the likelihood of having microalbuminuria in the group

with higher CRP was almost four times more than the lowest one and odds of having

microalbuminuria in the individual with higher serum fibrinogen was 2.16 (CI: 1.48- 3.15). The

more exposure to lead, the higher the likelihood was to have microalbuminuria. Also, among

lipid profiles, higher HDL had 71 % less likely to have microalbuminuria (Table 4).

ACR <30 mg/g 30≤ACR<300 mg/g

OR Crude* (95% CI)

HbA1c HbA1c<7 % HbA1c≥7 %

94.2% 67.8%

5.8% 32.2%

1

7.77(5.91-10.22)

CRP

<0.22 ≥0.22 &<1.0 >1.0

94.9% 90.8% 89.9%

5.1% 9.2% 10.1%

1.00 1.90(1.46- 2.48) 3.77(2.91- 4.90)

Fibrinogen <450 ≥450

91.4% 83.1%

8.6% 16.9%

1.00

2.16(1.48- 3.15)

Total Cholesterol (mg/dl) <240

≥240

96.9% 91.8%

3.1% 8.2%

1.00 1.93(1.56- 2.40)

HDL(mg/dl) ≥35

<35

94.0%

90.2%

6.0%

9.8%

1.00

1.71(1.34- 2.18)

Lead(ug/dl) <10 ≥10

93.5% 86.1%

6.5% 13.9%

1.00

2.32(1.65- 3.28)

20

Table5. Proportion and univariate association between the risk factors and CKD Risk Factors No CKD (95.8%) CKD (4.2%) OR Crude

* (95% CI)

Periodontitis No/Mild Moderate

Severe or Edentulous

97.8% 95.4%

87.85%

2.2% 4.6%

12.2%

1.00

2.11(1.51-2.95)

6.05(4.66-7.86)

Sex Men Women

94.9% 96.8%

5.1% 3.2%

1.00

1.64 (1.26-2.13)

Age 20-59 60+

99.0% 84.4%

9.0% 15.6%

1.00

20.65(15.16- 28.13)

Ethnicity Mexican- American &others Non-Hispanic white

Non-Hispanic black

98.0% 95.4%

96.7%

2.0% 4.6%

3.3%

1.00

2.47( 1.32-4.60)

1.72(0.89- 3.29)

Marital status Married Widowed Divorced

Single

96.5% 81.3% 97.0%

98.8%

3.5% 18.7% 3.0%

1.2%

1.00

6.42(5.07- 8.13) 0.87(0.59- 1.28)

0.33(0.20-0.54)

Education >12years ≤12 years

97.4% 94.8%

2.6% 5.2%

1.00

2.04(1.56- 2.67)

Country of Birth

USA Non USA

95.7% 97.0%

4.3% 3.0%

1.00 0.66(0.74- 0.96)

Income ≥20000$

<20000$

97.8%

94.3%

2.2%

5.7%

1.00

2.66(2.06-3.43)

Smoking Never smoked Former smoker Current smoker

96.0% 93.2% 97.8%

4.0% 6.8% 2.2%

1.00

1.72(1.41- 2.11) 0.51(0.35- 0.75)

Hypertension

No Yes

98.5% 87.8%

1.5% 12.2%

1.00 9.55(7.59- 12.02)

CVD No Yes

96.8% 86.4%

3.2% 13.6%

1.00

4.80(3.87- 5.97)

BMI <25 25-30 >30

96.6% 95.9% 94.5%

3.4% 4.1% 5.6%

1.00

1.21(0.96- 1.52) 1.67(1.33- 2.10)

WHR

<0.8if men ,<1.0 if women ≥0.8 if men ,≥1.0 if women

98.4% 94.5%

1.6% 5.5%

1.00 3.53(2.70- 4.63)

Last visit by dentist <1 years ≥1year

97.0% 94.5%

3.0% 5.5%

1.00

1.70(1.34- 2.14)

Last visit by doctor <1 years ≥1year

95.3% 98.5%

4.7% 1.5%

1.00

0.31 (0.21-0 .47)

21

Table5. (Continued)

* OR Crude = Unadjusted odds ratio for the association between chronic kidney disease and the

suspected/recognized risk factors in sample

Table 5 illustrates the association between various risk factors and CKD using univariate

analysis. In the individual with moderate peritonitis and severe periodontitis or edentulous, the

probability of CKD was raised with increase in severity.

In women odds of having CKD was 64% more than men. The older individuals were almost 20

times more likely to have CKD compared to the younger. In addition, the widowed group was

more probable to have CKD. The less the education, the more likely they have CKD.

Moreover, those who experienced longer period till being visited by a doctor were 69% less

probable to develop CKD. Likewise, CKD is less common among those who were born in non-US

countries. The odds of having CKD were significantly higher in the participants with

hypertension (OR=9.55), higher level of glycosylated hemoglobin (OR=4.2) and CVD

(OR=4.80). The association between other risk factors with CKD is summarized in Table 5.

Risk Factors No CKD (95.8%) CKD (4.2%) OR Crude * (95% CI)

HbA1c HbA1c<7 % HbA1c≥7 %

96.3% 86.0%

3.7% 14.0%

1.00

4.20(3.16- 5.59)

CRP <0.22 ≥0.22 &<1.0

>1.0

66.9% 24.7%

8.4%

50.8% 33.3%

15.9%

1.00

2.13(1.71- 2.65)

3.67(2.73- 4.94)

Fibrinogen <450 ≥450

93.3% 6.71%

86.1% 13.9%

1.00

2.86(2.04- 3.99)

Total Cholesterol(mg/dl) <240 ≥240

80.9% 19.1%

64.7% 35.3%

1.00

2.77(2.13- 3.60)

HDL (mg/dl) ≥35 <35

89.1% 10.95

82.3% 17.75

1.00

1.74(1.31- 2.30)

Lead (ug/dl) <10 ≥10

95.3% 4.7%

92.1% 7.9%

1.00

2.22(1.45- 3.41)

22

4.3 Multivariate Analysis At the first step of multivariable analysis for both microalbuminuria and CKD outcomes, all

significant and insignificant risk factors from univariate analysis were taken into account to

evaluate the effects of severe periodontitis for microalbuminuria and CKD, adjusted

simultaneously adjusted association for all variables. Although macroalbuminuria was excluded

while analyzing for microalbuminuria, it was taken into account when analyzing CKD.

Afterwards, in each step of back-ward stepwise logistic regression, the non-significant from each

group of risk factors were excluded. The result including the first and last models is summarized

in Table 5 and 6. Since there was high collinearity between CRP and fibrinogen, fibrinogen was

not taken into account in the multivariate logistic regression analysis.

The crude OR for having microalbuminuria among people with severe periodontitis was 3.00

(95% CI: 2.30-3.97). Sex, ethnicity, education, country of birth and income were not significant

which excluded from second model. (Second model is not illustrated in the table 6). In the third

model, last visit by dentist or doctor, smoking, CVD and BMI were not taken into account in the

analysis as the result of no significant association with microalbuminuria. In the last model

encompassing only the significant risk factors such as age, marital status, hypertension, HbA1c,

CRP, HDL and serum Lead, the odds of having microalbuminuria in individuals with severe

periodontitis remained significant (OR: 1.32;CI:1.03-1.7). (Table 6) In the last model, those with

severe periodontitis status were 32% more likely to have microalbuminuria compared to

individuals without periodontitis. On the other hand, moderate periodontitis was not significant

in association with microalbuminuria while adjusted for other risk factors although OR crude

was significant. (Table 4 and Table 6) The widowed individuals remained in higher risk by 83%

to have microalbuminuria when adjusted for other variables whereas the being single was not

significant any more. Moreover, the higher the CRP, the higher likelihood of enduring

microalbuminuria when adjusted for other risk factors encompassing traditional ones. In

addition, those with higher HDL were less probable to have microalbuminuria by 35% while total

cholesterol was not risk factor in the last model. Likewise, serum lead remained a significant risk

factor for microalbuminuria. (Table 4 and Table 6)

23

Table 6. Multivariable logistic regression analysis of risk factors of microalbuminuria

Risk Factors Model 1 ORadj *(95%CI)

(last model) ORadj * (95%CI)

Periodontitis No/Mild Moderate Severe or Edentulous

1.00

1.06(0.77-1.46) 1.33(1.04-1.69)

1.00

1.11(0.82-1.50) 1.32(1.03-1.70)

Sex

Men Women

1.00 1.33(0.97-1.81)

Age 20-59 60+

1.00 1.55(1.24-1.95)

1.00

1.60(1.28-2.00)

Ethnicity Mexican- American &others

Non-Hispanic white Non-Hispanic black

1.00

0.73(0.48-1.10) 0.92(0.63-1.35)

Marital status

Married Widowed Divorced Single

1.00 1.67(1.19-2.35) 1.10(0.77-1.56) 1.07(0.76-1.50)

1.00 1.83(1.35-2.48) 1.16(0.83-1.63) 1.06(0.78-1.44)

Education >12years

≤12 years

1.00

1.00(0.98-1.02)

Country of Birth USA Non USA

1.00 1.15(0.75-1.76)

Income ≥20000$

<20000$

1.00

1.05(0.86-1.29)

Last visit by dentist <1 years ≥1year

1.00

1.16(0.9-1.5)

Last visit by doctor <1 years ≥1year

1.00

0.79(0.58-1.06)

Smoking

Never smoked Former smoker Current smoker

1.00 1.19(0.9-1.58) 1.15(0.85-1.56)

Hypertension No Yes

1.00

2.59(2.09-3.2)

1.00

2.54(2.08-3.11)

CVD No Yes

1.00

0.91(0.69-1.2)

BMI

<25 ≥25 & ≤30 >30

1.00 0.98(0.77-1.23) 0.90(0.66-1.24)

HbA1c HbA1c<7 % HbA1c≥7 %

1.00

4.12(2.95-5.75)

1.00

4.21(3.09-5.75)

24

Table6. (Continued)

*OR adj = adjusted odds ratio for the association between chronic kidney disease and the

suspected/recognized risk factors in sample

Model 1: Goodness of fit: F-adjusted test statistic = 5.251, p= 0.000

Model final: Goodness of fit: F-adjusted test statistic = 1.251, p = 0.291

F-adjusted mean residual goodness-of-fit test was applied and suggested no evidence of lack of

fit in the last model (F-adjusted test statistic = 1.251, p = 0.291). (Table 6)

Table 7 illustrates the multivariate logistic regression analysis of traditional and non-traditional

risk factors of chronic kidney disease in this study.

In the first model adjusted for all the risk factors, the significant and insignificant ones which

were input in the univariate analysis, odds of having CKD in severe periodontitis group was 49%

more than those with no periodontitis or mild but moderate periodontitis was not significant risk

factor.

In the last model, socio demographic, health behavior and laboratory assays with a p-value

<0.05 were excluded stepwise. Eventually, adjusted for significant risk factors including age,

marital status (widowed), since last time visited by a doctor, hypertension, CVD, CRP, total

cholesterol and HDL, odds of having CKD in severe periodontitis group remained significant

(OR: 1.62;CI: 1.31-2.01). (Table 7)

Risk Factors Model 1 ORadj(95%CI)

(last model) ORadj(95%CI)

CRP <0.22 ≥0.22 &<1.0 >1.0

1.00

1.33(1.01-1.74) 2.23(1.57-3.16)

1.00

1.35(1.04-1.76) 2.35(1.73-3.19)

Total Cholesterol(mg/dl)

<240 ≥240

1.00 1.02(0.80-1.31)

HDL(mg/dl) ≥35 <35

1.00

1.51(1.17-1.94)

1.00

1.35(1.04-1.75)

Lead (ug/dl) <10

≥10

1.00

1.86(1.33-2.61)

1.00

1.92(1.40-2.62)

25

Table 7. Multivariable regression analysis of risk factors of CKD

Risk Factors Model 1 ORadj* (95%CI)

(last model) ORadj* (95%CI)

Periodontitis No/Mild Moderate Severe or Edentulous

1.00

0.84(0.59-1.19) 1.49(1.19-1.86)

1.00

0.83(0.59-1.15) 1.62(1.31-2.01)

Sex Men Women

1.00 1.28(0.92-1.77)

Age 20-59 60+

1.00 7.40(5.32-10.30)

1.00

8.04(5.90-10.96)

Ethnicity Mexican- American &others

Non-Hispanic white Non-Hispanic black

1.00

1.48(0.8-2.75) 1.05(0.53-2.09)

Marital status Married Widowed Divorced Single

1.00

1.36(1.03-1.81) 1.02(0.61-1.70) 0.79(0.44-1.41)

1.00

1.58(1.22-2.05) 1.07(0.71-1.63) 0.87(0.51-1.49)

Education

>12years ≤12 years

1.00 1(0.99-1.01)

Country of Birth USA Non USA

1.00

1.15(0.76-1.72)

Income ≥20000$

<20000$

1.00

1.33(0.99-1.78)

Last visit by dentist <1 years

≥1year

1.00

1.12(0.90-1.40)

Last visit by doctor <1 years ≥1year

1.00

0.56(0.37-0.85)

1.00

0.54(0.35-0.84)

Smoking Never smoked Former smoker Current smoker

1.00

1.17(0.89-1.54) 0.73(0.53-0.99)

Hypertension No

Yes

1.00

2.68(2.07-3.46)

1.00

2.85(2.2-3.69)

CVD No Yes

1.00

1.77(1.41-2.22)

1.00

1.92(1.52-2.42)

BMI <25 ≥25 & ≤30 >30

1.00

0.81(0.61-1.08) 1.00(0.74-1.36)

HbA1c HbA1c<7 %

HbA1c≥7 %

1.00

1.18(0.85-1.63)

26

Table7. (Continued)

*OR adj = adjusted odds ratio for the association between chronic kidney disease and the

suspected/recognized risk factors in sample

Model 1: Goodness of fit: F-adjusted test statistic = 236.88, p= 2.029 Last model: Goodness of fit: F-adjusted test statistic= 170675.36, p=8.370

The likelihood of having CKD among the older was almost 8 times as much as the younger.

Those who were widowed, had CVD, higher total cholesterol or lower HDL were more likely to

manifest chronic kidney disease.

In individuals with more than one year since to visit doctor, CKD was less common by 46.

(Table7)

Among those who had hypertension and severe periodontitis, the probability of getting CKD

was 4.62 times as much as those with neither severe periodontitis nor hypertension (CI:3.31-

6.45) (analysis is not depicted in the table 7).

Despite the result for having microalbuminuria, lipid profile (higher total cholesterol and lower

HDL) remained risk factor to have CKD after adjustment for other risks while serum lead was

not significant risk factor for CKD.

Risk Factors Model 1 ORadj (95%CI)

(last model) ORadj (95%CI)

CRP <0.22 ≥0.22 &<1.0 >1.0

1.00

1.42(1.08-1.87) 1.72(1.17-2.55)

1.00

1.44(1.12-1.85) 1.95(1.33-2.85)

Total Cholesterol (mg/ml) <240 ≥240

1.00

1.59(1.21-2.08)

1.00

1.63(1.22-2.18)

HDL (mg/dl) ≥35

<35

1.00

1.84(1.26-2.69)

1.00

1.71(1.22-2.40)

Lead(ug/dl) <10

≥10

1.00

1.81(1.04-3.15)

27

5. Discussion Chronic kidney disease (CKD) is posed as a significant global challenge and burden of public

health. Evaluation of predictors of CKD is a complex issue due to existence of association

between CKD and age, race and other chronic diseases such as diabetes, hypertension and

cardiovascular disease (CVD). CKD is a crucial public health burden regarding its morbidity and

mortality. 4, 24

Albumin to creatinine ratio above 30 and less than 300 mg/g is an early manifestation of renal

cell damage, which can lead to decreased GFR and CKD.1 Also, periodontitis which can be

controlled is known as the non-traditional risk factor for some systemic diseases such as CVD by

effect on the endothelial cells.4

In addition to the traditional risk factors encompassing diabetes and hypertension, the non-

traditional risks factors as periodontitis are speculated with their influence on endothelium

structure and function through infection-driven inflammatory markers in the kidney. In many

studies, the association between periodontitis and cardiovascular disease was revealed through

inflammatory mechanism .7, 9,11,24,25,31-38

In a study conducted by Tonetti et al, they showed intensive periodontitis treatment leads to

improvement in endothelial function via decrease in inflammatory factors. 11 In the research

conducted by Salzberg et al in 2006 , it is proved that patients with severe periodontitis had

statistically significant elevated serum CRP compared to those without periodontitis.39 Likewise,

Paraskeva et al disclosed that there is strong evidence that serum CRP is highly associated with

periodontitis.40 In a study was performed by Navarro et al in 2003, the association between

urinary albumin excretion and inflammatory factors in the patients with diabetes type2 was

depicted.41 In our study there was significant association between higher increased CRP and

microalbuminuria (OR: 2.35, CI: 1.73-3.19) indicating significant association between

microalbuminuria and the inflammatory factor.

Kshirsagar et al in ARIC study (Atherosclerotic Risk in Community) concluded the existence of

association between periodontal disease and prevalent renal insufficiency.13 They studied 2,276

individuals with initial periodontitis and 947 with severe periodontitis. In their study an

association between periodontitis and CKD was also remarked (OR: 2.00; 95% CI: 1.23 to 3.24).

But, in this study edentulousness, for which periodontitis is the primary cause was excluded 41.

Also microalbuminuria was not taken into account as one of the main outcomes. Moreover, the

definition of periodontitis was not based on new CDC definition, albeit measured 6

interproximal Sites which increase the accuracy of measurement.22

In our study we extended the findings by demonstrating an association with CRP and

microalbuminuria, and employed an updated definition of periodontitis while focusing on

chronic and severity of periodontal disease.

28

There are challenges regarding case definition to determine periodontitis across researches.

Although many studies assessed the association between periodontitis and chronic kidney

disease or CVD, there is dearth of uniformity in the definition of periodontitis in epidemiologic

studies.18-23

Fisher et al42 in 2009 studied a prediction model for CKD including periodontitis. In their study

they considered one or more sites with CAL ≥4mm and bleeding on the same tooth as definition

of periodontitis. Also they defined the edentulous status as alternative indicator of severe

periodontitis. The main outcome was CKD (eGFR<60ml/min/1.73m2). The result after

adjustment for various risk factors were OR: 1.60, CI: 1.07 to 2.39 and OR: 2.03, CI: 1-31 to 3.14

in order for periodontitis and edentulous states. The CDC definition adapted in our study 17

combines severe status with edentulous as indicator of severe periodontitis and the odds of

developing CKD was 62% (OR adjust=1.62, CI: 1.31-2.01). In previous study, inflammatory factors

or microalbuminuria were not taken into account except t macroalbuminuria (OR: 3.41, CI: 1.82

to 6.41). Yet, in the current survey microalbuminuria was one of the main outcome as early

manifestation of kidney dysfunction which can potentially be reversible.

Although nephrotoxic effect of lead have been known, in non of the former studies serum lead

was controlled as a risk factor in the analysis43 . In the current study, there was significant

association between higher serum lead and microalbuminuria adjusted for other risk factors

(OR: 1.92, CI: 1.40-2.62).

In a similar study44 Fisher et al took into consideration CRP and other serologic markers but not

microalbuminuria as early manifestation of kidney dysfunction which can be potentially reversed

by treatment of periodontal disease as common source of chronic inflammation.

On the other hand, gingival recession which can be the result of periodontitis and even after

treatment leads to false positive in measurement of CAL and periodontitis was not considered

not in the former neither in the latter studies. While, in the present study we considered

interproximal site (mesiobuccal) to measure CAL plus pocket depth which diminish the

likelihood of this bias since accuracy of periodontitis measurement by utilizing these indices

together are less influenced by gingival recession. In another survey, Fisher et al proved the

association between edentulous as a single risk factor and chronic kidney disease. Similarly,

microalbuminuria was not studied. 45

In 2009, Garcez et al in a case-control study assessed the oral health status of individuals with

mild decrease in eGFR (60-89 ml/min). They considered various oral health status as number of

decayed, missing or filled teeth, supra gingival inflammation, pocket depth, CAL and dental

mobility. There was no alteration of the oral health between case and control groups which can

be justified as the consequence of small sample size. 46

In current survey, the relationship between smoking status and microalbuminuria was not

statistically significant albeit reverse association with CKD which is counter to some but not all

29

other studies.44, 45 This result can be due to the method of definition for smoking since those who

were known as former smoker or never smoker but with high serum cotinine were indicated as

current smoker, yet cotinine can be higher in passive smokers or in those who snuff nicotine

instead of smoking while in Fisher et al study, there was significant association between both

former and current smokers with CKD. 44, 46

WHR and BMI had no significant association neither with microalbuminuria nor CKD. With

respect to various studies which reviewed traditional and non-traditional risk factors for CKD

and also microalbuminuria, subsequent risk factors were taken in to account in the model: age,

sex, marital status, HbA1c, HDL, serum lead.

As the limitation, this study utilized NHANESIII data the number of interproximal sites

measured is restricted to only mesiobuccal site while if were measured in six interproximal sites

the result could be altered probably toward stronger association.

Another important limitation of this study is its design which is cross-sectional which cannot

assess temporality. To measure causality of periodontitis as exposure, it is necessary that it

precedes the progression of microalbuminuria or CKD. Also, there is no uniform definition for

periodontitis to be able to compare the result with all other conducted studies.

Despite all limitations in this study, there is some evidence that warrant considering severe

periodontitis as risk factor of microalbuminuria and CKD (Table 5 and 6). Our findings overall

consistent with previous studies and have strong biological plausibility. Moreover, severe

periodontitis has stronger association with microalbuminuria or CKD compared to no

periodontitis, on the other hand, no association between moderate periodontitis and

microalbuminuria or CKD, which strengthen the probable causality regarding dose response.

Also, in NHANES data questionnaire, laboratory tests and examination were conducted in

unbiased methods.

30

6. Conclusion

Severe periodontitis had association with microalbuminuria as early indicator of endothelia

damage in kidney glomerular system which can be reversible at the early stage.

From a public health perspective, we suggest that the association between periodontitis with

inflammatory markers and both microalbuminuria and CKD have reasonable probability of

being causal. If this link is indeed true, then the treatment of periodontitis may in addition to

improving oral health may impede the development of microalbuminuria and CKD. Thus, oral

health and dental insurance in a public health system should be one of the priorities and we

believe that the current evidence is strong enough to warrant longitudinal studies to help further

elucidate causality and that clinical trials of periodontal treatment, which are beneficial to

participants and plausible, should also be evaluated in respect to potential reduction in

microalbuminuria.

31

7. Acknowledgment

This research was performed by utilizing data from NHANESIII conducted by National Center

for Health Statistics. My gratefulness goes to Nawi Ng, my supervisor from Umea University,

department of public health and epidemiology that always kept me on track by his significant

and scientific guidance and his generosity of sharing knowledge.

Particular thanks to Afshin Parsa, the second supervisor that his Nephrology knowledge and

advice lightened the way of the work. Also, deep appreciation goes for Public Health and

Epidemiology Department that provided me with the great opportunity to raise my knowledge

embedded in this research.

Eventually, especial thanks to my parents that were always my best encourager. Their spirits

may in peace.

32

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