Fructose-induced Hepatic De Novo Lipogenesis in Adolescents With Obesity

Hypothesis: Hepatic DNL in response to the ingestion of fructose:glucose 1:1 is significantly
greater in adolescents with obesity and prediabetes compared to adolescents with obesity who
are metabolically healthy.

Thirty obese adolescents, ages 12-21 years, will be recruited to each of two groups: 1)
prediabetes (n=15) and 2) metabolically healthy (n=15). An attempt will be made to achieve
similar age, sex and BMI distributions in the two groups.

Visit 1- Screening: Eligible subjects will be enrolled after Informed consent/assent are
obtained by a physician investigator. The medical history will be reviewed and a physical
exam performed, including an assessment of pubertal status. Blood will be sampled after at
least a 12 hour fast (except water) for complete blood count, serum glutamate pyruvate,
glucose, HbA1C, and lipid panel, if not previously documented in the medical record within
three months of the screening visit. These tests may be done without fasting, but if the
glucose or lipid results are not normal, these two tests must be repeated after fasting.

Visit 2 - Sugar Challenge Test with Blood Sampling: Within three months of the screening
visit, eligible participants will return after at least a 12 hour fast (except water).
General health, family history and recent diet will be reviewed. The height, weight, waist
circumference at the umbilicus, and blood pressure will be recorded. A urine sample will be
obtained for microalbuminuria. After placement of an intravenous catheter and withdrawal of
the baseline blood samples, participants will drink a sweet beverage prepared by a dietician
(fructose:glucose 1:1, 3g/kg, in water). Blood sampling will continue every hour for three
hours( ~12 mL per time point).

Percent body fat will be measured by bioimpedance. The percent body fat will be calculated by
computer software with prediction equations validated in the pediatric and adult age ranges.

Plasma samples will be obtained at 4 time points (0, 1, 2 and 3 hours), the fatty acid
composition of plasma very low density lipoprotein triglyceride (VLDL TG), including
%palmitate (16:0), will be measured. A lipid panel and VLDL triglycerides and apolipoprotein
B levels will also be measured at each time point. Other assays to be performed include
insulin and and glucose, HbA1C and high sensitivity C reactive protein (hsCRP, a marker of
inflammation). The urine sample will be assayed for microalbuminuria (albumin/creatinine >30
mg/g).

An interim analysis of the primary endpoint (increase in VLDL TG %16:0) will be performed in
the first three subjects of each group to determine whether the dose of sugar and sampling
duration of 3 hours are sufficient to produce a measurable increase in VLDL TG %16:0.

Inclusion Criteria:

1. Age: 12-21 years old.

2. BMI percentile >95th and <140th percentile of the 95th, maximum weight 110 kg.

3. Group 1: HbA1C <5.7% and no history of fasting hyperglycemia (glucose greater or equal
to100 mg/dL and/or HbA1C greater or equal to 5.7%), sustained hypertension (>95th
percentile) or abnormal lipids (LDL cholesterol <130 mg/dL, TG <130 mg/dL, HDL-C >40
mg/dL).

Group 2: HbA1C greater or equal to 5.7 to 6.49%

4. Pubertal or post- pubertal stage (pre-pubertal excluded).

5. Willing and able to stop fish oil, fiber supplement, other non-prescribed
vitamins/supplements for 1 week prior to study visit.

6. Willing to not drink alcohol for at least 48 hours before the study visit.

7. Willing to refrain from vigorous exercise the day before the study visit.

Exclusion Criteria:

1. Diabetes (fasting blood glucose greater or equal to126 mg/dL, and/or HbA1C >6.5%
and/or history of a 2 hour oral glucose tolerance test (OGTT) >200 mg/dL).

2. Triglycerides >500 mg/dL, LDL cholesterol >160 mg/dL.

3. Medications affecting lipid or glucose metabolism, including hormonal contraceptives.

4. Liver disease or intestinal dysfunction. Serum glutamate oxaloacetate and/or serum
glutamate pyruvate < 2x upper limit normal is permitted if there is no diagnosis of
liver disease by a gastroenterologist.

5. Weight loss >10% in past 6 months.

6. Cigarette smoking.

7. Alcohol or drug abuse.

8. Systemic illness, including polycystic ovary disease and acute infections within two
weeks of study visits.

9. Anemia.

10. Pregnancy or breast feeding.

11. Psychiatric illness.

12. Extreme diet or level of physical activity.

13. Fructose intolerance.

14. Implanted electronic devices (bioimpedance measurement).

15. Any other condition, which in the opinion of the principal investigator, should
prohibit participation in the study.