This study determined Native American population, aged 17-30 years, at Fort Lewis
College shows a higher likelihood for developing insulin resistance than the Caucasian population. High levels of molecular markers such as IL-6, TNF-α, and erythrocyte sedimentation rate are indicative of inflammation which can be correlated to insulin resistance. In order to determine IL-6 concentrations, serum was extracted from whole blood and a sandwich ELISA was performed. The concentration of IL-6 was determined by absorbance at 405 nm. The mean concentration of IL-6 in the Native American group was 1.7885 pg/mL + 0.84 whereas mean concentration for the Caucasian group was 1.3465 pg/mL + 0.15. This indicates a significant difference among the populations studied.
There were moderately significant correlations for IL-6 with TNF-α and erythrocyte sedimentation rate shown by r values equaling 0.43 and 0.50 respectively. For continued research a larger sample population would be required to further correlate data.
Insulin resistance (IR) is a growing problem for Native Americans that carries into a high propensity for developing diabetes leading to a shortened lifespan (Carter et al.,
2000). Chronic inflammation has been identified as a contributor to IR and metabolic syndrome (Fernandez-Real and Ricart, 2003; Dandona et al., 2005; Petersen and Pedersen,
2005; Andreozzi et al., 2006). Inflammatory markers indicate the presence of IR which progresses into type II diabetes (Fernandez-Real and Ricart, 2003). Adipocytes in visceral fat are responsible for releasing proinflammatory cytokines such as Interleukin-6 (IL-6) and Tumor Necrosis Factor-α (TNF- α) (Pedersen et al., 2001; Shoelson et al., 2006), thus increased adiposity correlates with increased inflammation.
IL-6 is a pro-inflammatory cytokine (Coppack, 2001; Andreozzi et al., 2006) involved in the inflammation process and hypothesized to lead to IR (Kopp et al., 2003). IR is positively correlated with high levels of IL-6 in the bloodstream (Kern et al., 2001;
Fernandez-Real and Ricart, 2003; Kershaw and Flier, 2004).
TNF- α is found to have a direct role in metabolic syndrome (Petersen and Pedersen, 2005) through blocking insulin function (Fernandez-Real and Ricart, 2003) and generating IR
(Coppack, 2001; Kershaw and Flier, 2004).
IL-6 and TNF- α share a mechanistic pathway in which they block the expression of glucose transporter-4 (Glut-4) and insulin receptor substrate-1 (IRS-1). Glut-4 has been found to aid in the insulin stimulated glucose transport system while IRS-1 is a protein that aids in the binding of insulin to its receptors on cells (Rotter et al., 2003). Through inhibition of integral protein function, the effectiveness of insulin to bind is decreased
(Rotter et al., 2003).
Volunteers were randomly selected through the Native American and Caucasian student population at Fort Lewis College who were aged 17-30. The only consideration was that they could not have been diagnosed with diabetes or insulin resistance. Subject participation was voluntary receiving only $10 Durango dollars in return for participation.
Patient volunteers excluded only by presence of known diabetes condition.
Ten mL of blood were collected in EDTA tubes (Becton Dickinson; Franklin Lakes, NJ) by venipuncture at Fort Lewis College using standard protocols by a licensed phlebotomist.
Immediately after collection, specimens were processed per variable protocol. All specimens were centrifuged at 1000g within 30 minutes of collection, and serum separated from whole cells. All serum samples were stored at -20°C until subsequent ELISA analysis.
IL-6 was measured by a quantitative sandwich ELISA kit (R&D Systems Inc.;
Minneapolis, MN) in duplicate.
TNF-α was measured by a quantitative sandwich ELISA kit (R&D Systems Inc.;
Minneapolis, MN) in duplicate.
One mL of whole blood was added to a 1mL serological pipette with a 0.3cm width, coated with a 1.25M solution of sodium citrate. After one hour the sedimentation rate was determined by recording the distance the erythrocytes have fallen in mm per hour.
2,5
2
1,5
1
0,5
0
Mean IL-6 Concentration per
Population Group
Native American Caucasian
Population Group
Figure 1. The average plasma IL-6 concentration was significantly higher in NA population (p=0.037).
50
45
40
35
30
25
20
15
10
5
0
0
Comparison of IL-6 to
Sedimentation Rate in Native
American Subjects y = 5.3076x + 6.4361
R² = 0.2563
1 2 3
IL-6 Concentration (pg/mL)
4 5
50
45
40
35
30
25
20
15
10
5
0
1
Comparison of IL-6 to
Sedimentation Rate in Caucasian
Subjects
1,2 1,4 1,6
IL-6 Concentration (pg/mL) y = 15.731x - 3.2891
R² = 0.0491
1,8 2
Figure 2. Comparison of IL-6 plasma concentrations to erythrocyte sedimentation rates. For Native Americans, the r-value was 0.508, which is moderately correlated. For Caucasians the r-value was 0.2945, which is a low correlation.
20
10
0
0
50
40
30
60
Comparison of IL-6 to TNF-α in
Native American Subjects y = 2.1394x + 36.109
R² = 0.1814
1 2 3
IL-6 Concentration (pg/mL)
4 5
60
Comparison of IL-6 to TNF-α in
Caucasian Subjects
50
40
30
20
10
0
1 y = -11.913x + 54.159
R² = 0.1072
1,2 1,4 1,6
IL-6 Concentration (pg/mL)
1,8 2
Figure 3. Comparison of IL-6 plasma concentrations to TNF-α plasma concentrations. Has shown to be moderately correlated in Native American subjects with an r-value of 0.425. For Caucasians the r-value was found to be -0.181, which is a low correlation.
Table 1. Anthropometric and biochemical characteristics of the study subjects.
Characteristics
Sex (M/F)
Waist-to-hip ratio
BMI (kg/m 2 )
% Body Fat
Fasting Glucose (mg/dL)
Native American
3/11
0.81 ± 0.08 (0.69-0.96)
24.94 ± 3.58 (19.8-32.9)
22.29 ± 5.16 (16-29.8)
78.43 ± 20.46 (33-109)
Caucasian
3/11
0.79 ± 0.07 (0.90-.68)
23.87 ± 3.50 (19.2-32.1)
19.31 ± 5.56 (13-27.9)
80.71 ± 10.74 (61-102)
Sedimentation Rate (mm/hr) 15.93 ± 8.82 (5-34.5)
TNF-α (pg/mL)
Adiponectin (pg/mL)
39.94 ± 4.23 (32.29-49.42)
2265.43 ± 1384.47 (915-
6000)
17.89 ± 10.59 (4-41)
38.12 ± 5.43 (27.87-47.37)
2261.79 ± 1389.96 (915-
6000)
IL-6 (pg/mL) 1.80 ± 0.84 (1.22-3.05) 1.34 ± 0.15 (1.12-1.69)
Data are means ± SD (range) and median (range).
Figure 4. Interleukin-6 protein structure.
INFLAMMATION
ENDOTHELIAL
DYSFUNCTION
IL-6/TNF-α
SERINE
PHOPHORYLATION
OF IRS-1
DECREASED
INSULIN SIGNAL
TRANSDUCTION
INSULIN
RESISTANCE
Figure 5. Possible pathway of the proinflammatory cytokines IL-6 and TNF-alpha leading to insulin resistance.
Figure 6. 2010 Diabetes Research Team. From left to right: Leon Clah, Katie Zortman, Gabe
Thom, Chelsea Bonfiglio, Samantha Johnson,
Brittany Walters, Hannah Meinking, Heather
Dahm, Dr. Sherell Byrd, Edlin Jara-Molinar.

Explore the relationships between inflammatory cytokines IL-6 and TNF-alpha

Explore the data within the research project, in order to find new correlations between different indicators of type II diabetes

Discover whether or not there is a higher propensity for developing insulin resistance among the Native American population versus the Caucasian population at Fort
Lewis College

Implement an awareness campaign if Native Americans are found to be at greater risk of developing diabetes

Add on to the current knowledge of Native American risk for developing type II diabetes
Evidence for the link between inflammation and IR is provided in this study which builds upon current research. Systemic inflammation, as measured by serum IL-6 concentration was significantly higher in Native American as compared to their matched Caucasian peers.
Furthermore, IL-6 positively correlated with other physiological indicators of inflammation, TNF- α and erythrocyte sedimentation rate. TNF- α locally identifies inflammation and moderately correlates ESR in the Native American population.
Sedimentation rate positively correlates with IL-6 collectively identifying inflammation as prevalent in the Native American population. The goal of our study was to identify cellular markers of IR in Native Americans. A high correlation in inflammatory cellular markers
IL-6, TNF- α, and sedimentation rate was present in the Native American population. This indicates that underlying inflammation that is often a precursor to insulin resistance, is already present in this college-aged population.
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Thank you to Fort Lewis College for assisting us in our research through a research grant from the Department of Natural and Behavioral Sciences. We acknowledge the Student
Health Center staff for doing all of the blood draws.