Diabetes type two is a debilitating disease that leads to chronic morbidity such as
accelerated microvascular disease. Accelerated microvascular disease may produce blindness,
end stage renal disease, myocardial infarction, stroke, and limb ischemia. Strategies to
prevent or delay microvascular disease have the potential to improve the lives of millions
and prevent catastrophic illness. The major focus of prevention of microvascular disease in
diabetes has been on the endothelium and its role in protection of blood vessels. An
unexpected means to prevent microvascular disease in diabetes may be coupled to the function
of vitamin C in red blood cells (RBCs) of diabetic subjects. Based on new and emerging data,
vitamin C concentrations in RBCs may be inversely related to glucose concentrations found in
diabetes. Based on animal data, we hypothesize that RBCs with low vitamin C levels may have
decreased deformability, leading to slower flow in capillaries and microvascular hypoxia, the
hallmark of diabetic microangiopathy. Low vitamin C concentrations in RBCs of diabetic
subjects may be able to be increased, by using vitamin C supplements. Findings in animals may
not accurately reflect effects in humans because of species differences in mechanisms of
vitamin C entry into RBCs. Therefore, clinical research is essential to characterize vitamin
C physiology in RBCs of diabetic subjects. In this protocol we will investigate physiology of
vitamin C in RBCs of diabetic subjects as a function of glycemia, without vitamin C
supplementation (arm 1) and with vitamin C supplementation (arm 2). We will screen type II
diabetic subjects on insulin and/or oral hypoglycemic medication(s) and select those with
hemoglobin A1C concentrations of less than or equal to 12%. Selected subjects may be
hospitalized twice, each time for approximately one week. The primary objective of the first
hospitalization (arm 1) will be to evaluate the effect of hyperglycemia on vitamin C RBC
physiology regardless of baseline vitamin C concentrations (without any vitamin C
supplementation). The second hospitalization (arm 2) investigates the effect (if any) of
vitamin C supplementation to changes in RBC physiology during euglycemic and hyperglycemic
states. As inpatients, subjects will have two venous sampling periods each of approximately
24 hours. On admission, subjects may be fitted with continuous glucose monitors (CGMs), oral
hypoglycemic agents will be discontinued, and basal-bolus insulin regimen initiated. Insulin
doses will be clinically determined and titrated to achieve euglycemia (fasting and pre-meal
glucoses <140mg/dl) prior to the first sampling period (euglycemic sampling). The first
sampling period will be performed under conditions of euglycemic control for 24 hours. The
second sampling period will be performed under controlled hyperglycemia induced by
withholding basal and bolus insulin and providing a high carbohydrate load diet (70-75%
carbohydrate). Correction-scale insulin will be provided for glucoses >350-400mg/dl.
Hyperglycemia will not exceed 9 hours, and will be reversed by reinstituting insulin.
During the two sampling periods, samples will be withdrawn via venous catheter for RBC
deformability, vitamin C concentrations and other related research studies. Following
completion of arm 1, subjects considered for participation in arm 2 will be provided a
prescription for vitamin C 500mg twice daily. Given that vitamin C and vitamin E are related
antioxidants, and that both vitamins appear to be associated with RBC rigidity, diabetic
subjects may also be given a prescription for 400 international units (IU) of vitamin E (RRR
alpha tocopherol) daily. Subjects will continue vitamin C and E supplementation for a minimum
of 8 weeks depending on RBC vitamin C concentrations. To evaluate any effect of vitamin E
supplementation, plasma and RBC vitamin E levels may be measured concurrently with vitamin C
levels during various phases of arms 1 and 2. All subjects will be seen as outpatients at
biweekly or monthly intervals with regular measurement of plasma and RBC vitamin C
concentrations. Target RBC vitamin C concentration >30uM is required prior to arm 2 inpatient
sampling studies. Vitamins C and E supplementation will be discontinued upon inpatient
admission for arm 2. Risk of both vitamin supplements are minimal as both supplementation
doses are safe. Outcomes are to measure RBC rigidity and vitamin concentrations before and
after supplementation. After a minimum of 8 weeks (depending on RBC vitamin C levels),
subjects will be hospitalized again, and sampling repeated as described. In this manner, each
subject serves as his/her own control, and deformability of red blood cells can be determined
in relation to glycemia and to vitamin C concentrations in RBCs and plasma.
Subjects will be required to consume standardized meals during inpatient stays. All meals
will be prepared by the NIH Clinical C...
- Vitamin E is an antioxidant that reduces the damaging effects of oxygen in the body.
Most American men (90%) and women (96%) do not get enough vitamin E from their diets;
however, the amount of vitamin E needed by the body has been studied only in men, not
women. In addition, it is unknown whether another antioxidant, vitamin C, helps vitamin
E in protecting the body. Because vitamin E is a fat-soluble vitamin, how much body fat
a person has could affect the amount of vitamin E needed for protection.
Objectives: This study has three arms to examine vitamin E requirements:
- To determine the amount of fat required to get the best vitamin E absorption from a
- To determine the amount (i.e., best dose) of vitamin E that must be consumed before it
can be measured in the blood.
- To examine how vitamin E and vitamin C work together in the body, in conjunction with
diet and vitamin supplements.
- Arms 1 and 2: Women between the ages of 18 and 40 years who have a normal weight and
body mass index (BMI) of 27 or less.
- Arm 3: Women between the ages of 18 and 40 years who have a normal weight (BMI 27), who
are overweight (BMI > 27), or who are overweight (BMI > 27) and have non
- Arm 1: Five studies, each lasting 1 month with 1 month off between studies (total study
= 10 months). Participants will take 500 1,000 mg of vitamin C twice daily for 2 weeks
before admission to the clinical center for 1 week.
- Study 1: Participants will eat breakfast containing a known amount of fat, after which
they will take a vitamin E pill as well as receive an IV injection of vitamin E. Other
foods contain only negligible amounts of vitamin E. Blood and urine samples will measure
levels of vitamin E and other substances.
- Studies 2 5: Outpatient visits will consist of the same tests as in Study 1; however,
the amount of fat in the breakfast will range from 0% to 40% in random order. During one
of the studies, an adipose tissue biopsy will be collected to determine how much vitamin
E is in the tissues.
- Arm 2: Five studies, each lasting 1 month with 1 month off between studies (total study
= 10 months). Preparation for Arm 2 is the same as in Arm 1. The proportion of fat,
muscle, and water in the body will also be measured.
- Study 1: Participants will eat breakfast containing 30% fat, after which they will take
a vitamin E pill as well as receive an IV injection of vitamin E. Conditions and
procedures are the same as in Arm 1.
- Studies 2 5: Outpatient visits will consist of the same tests as in Study 1; however,
the amount of vitamin E in the breakfast will range from 2 to 30 mg in random order.
- Arm 3: Outpatient (2 to 6 weeks) and inpatient studies (4 to 6 weeks).
- Outpatient study: Participants will take 500 1,000 mg of vitamin C daily and provide
blood and urine samples, as well as an adipose tissue sample.
- Inpatient studies: Two vitamin E inpatient studies. Before these begin, participants
vitamin C blood levels will be reduced by means of a diet low in vitamin C. Blood tests
will determine how quickly vitamin C leaves the body. Once the vitamin C level is
reduced, the first vitamin E study will begin.
Study A: The procedure for this study is the same as in Arm 2, Study 1.
Study B: The procedure for this study is the same as in Study A, except that the participants
blood vitamin C levels will be higher.
Several studies have reported that diabetic subjects have lower plasma vitamin C
concentrations than non-diabetic subjects. Although urinary vitamin C loss in diabetic
subjects was reported to be increased in two studies, these are difficult to interpret due to
lack of controlled vitamin C intake, inadequate sampling, lack of control subjects, or
methodology uncertainties in vitamin C assay and sample processing. Consequently, it is
unclear whether diabetic subjects truly have both low plasma and high urine vitamin C
concentrations. We propose that low plasma vitamin C concentrations in diabetic subjects are
due in part to inappropriate renal loss of vitamin C in these subjects but not in healthy
controls. We will study vitamin C concentrations in patients with type 1 and type 2 diabetes
and in matched healthy research subjects. Vitamin C concentrations in plasma, neutrophils (as
a proxy for tissue concentrations) and in urine will be measured in outpatients. In those
willing to be admitted to the Clinical Center, we will measure 24-hour urinary excretion of
vitamin C while on a vitamin C free diet, and creatinine clearance, a measure of glomerular
filtration rate. On day 2 of the inpatient study, subjects will receive a single 200mg dose
of oral vitamin C and we will measure vitamin C concentrations in frequent blood and urine
samples to determine the renal threshold and relative bioavailability for vitamin C. Single
nucleotide polymorphisms (SNPs) will be determined in genomic DNA responsible for the two
proteins mediating sodium-dependent vitamin C transport, SVCT1 and SVCT2. If low plasma and
high urine vitamin C concentrations are found in diabetic subjects, further studies will be
needed to explore mechanisms and to determine recommended dietary allowances for this patient