Primary care diabetes assessment when HbA1c and other measures of glycemia disagree.

AIMS: Although diabetes management decisions in primary care are typically based largely on HbA1c, mismatches between HbA1c and other measures of glycemia that are increasingly more available present challenges to optimal management. This study aimed to assess a systematic approach to identify the frequency of mismatches of potential clinical significance amongst various measures of glycemia in a primary care setting. METHODS: Following screening to exclude conditions known to affect HbA1c interpretation, HbA1c, and fructosamine were obtained and repeated after ∼90 days on 53 adults with prediabetes or type 2 diabetes. A subset of 13 participants with repeat labs wore continuous glucose monitoring (CGM) for 10 days. RESULTS: As expected, HbA1c and fructosamine only modestly correlated (initial R2 = 0.768/repeat R2 = 0.655). The HbA1c/fructosamine mismatch frequency of ± 0.5% (using the following regression HbA1c = 0.015 *fructosamine + 2.994 calculated from the initial sample) was 27.0%. Of the 13 participants with CGM data, HbA1c and CGM-based Glucose Management Indicator correlated at R2 = 0.786 with a mismatch frequency of ± 0.5% at 46.2% compared to a HbA1c/fructosamine mismatch frequency of ± 0.5% at 30.8%. CONCLUSIONS: HbA1c is frequently mismatched with fructosamine and CGM data. As each of the measures has strengths and weaknesses, the utilization of multiple different measures of glycemia may be informative for diabetes assessment in the clinical setting.

Skeletal muscle quality, measured via phase angle, and cardiorespiratory fitness in patients with obesity and heart failure with preserved ejection fraction.

OBJECTIVES: Cardiorespiratory fitness (CRF) is influenced by body composition quantity and quality in heart failure with preserved ejection fraction (HFpEF) and obesity. Bioelectrical impedance analysis (BIA) provides a noninvasive quantitative and qualitative body composition assessment. The aim of this study was to determine the role of phase angle (PhA), a BIA-measure of skeletal muscle quality and body cell mass, on CRF in patients with obesity and HFpEF. METHODS: Fifty-nine consecutive outpatients with HFpEF underwent cardiopulmonary exercise testing to measure CRF. Single-frequency segmental BIA was used to measure PhA and body composition quantity. Resting Doppler echocardiography and biomarkers were measured to assess cardiac function and systemic inflammation. RESULTS: Compared with patients with lower PhA, patients with higher PhA (above mean 5.8°) presented a greater absolute peak oxygen consumption (VO2; 1.83 [1.3-2.1] versus 1.39 [1.1-1.6] L/min, P = 0.003), VO2 peak adjusted for body weight (17.5 [12.3-18.1] versus 13.3 [12.7-15.2] mL/kg/min, P = 0.040), and a lower edema index (48.7 [2.9] versus 51.4% [2.7], P < 0.001) and N-terminal pro-B-type natriuretic peptide (NT-proBNP; 64 [50-121] versus 183 [68-343.5] pg/dL, P < 0.001). In the overall sample, PhA was correlated with absolute VO2 peak (r = 0.468, P < 0.001), VO2 peak adjusted for body weight (r = 0.368, P = 0.004), VO2 peak adjusted for fat-free mass (r = 0.315, P = 0.015), edema index (r = -0.508, P < 0.001), and NT-proBNP (r = -0.579, P < 0.001). PhA remained a significant predictor for CRF even after adjustment for potential confounders and HFpEF severity. CONCLUSION: In patients with obesity and HFpEF, a greater PhA is an independent predictor for favorable CRF.

Antibodies against biotin-labeled red blood cells can shorten posttransfusion survival.

BACKGROUND: In hematologic and transfusion medicine research, measurement of red blood cell (RBC) in vivo kinetics must be safe and accurate. Recent reports indicate use of biotin-labeled RBC (BioRBC) to determine red cell survival (RCS) offers substantial advantages over 51 Cr and other labeling methods. Occasional induction of BioRBC antibodies has been reported. STUDY DESIGN AND METHODS: To investigate the causes and consequences of BioRBC immunization, we reexposed three previously immunized adults to BioRBC and evaluated the safety, antibody emergence, and RCS of BioRBC. RESULTS: BioRBC re-exposure caused an anamnestic increase of plasma BioRBC antibodies at 5-7 days; all were subclass IgG1 and neutralized by biotinylated albumin, thus indicating structural specificity for the biotin epitope. Concurrently, specific antibody binding to BioRBC was observed in each subject. As biotin label density increased, the proportion of BioRBC that bound increased antibody also increased; the latter was associated with proportional accelerated removal of BioRBC labeled at density 6 µg/mL. In contrast, only one of three subjects exhibited accelerated removal of BioRBC density 2 µg/mL. No adverse clinical or laboratory events were observed. Among three control subjects who did not develop BioRBC antibodies following initial BioRBC exposure, re-exposure induced neither antibody emergence nor accelerated BioRBC removal. DISCUSSION: We conclude re-exposure of immunized subjects to BioRBC can induce anamnestic antibody response that can cause an underestimation of RCS. To minimize chances of antibody induction and underestimation of RCS, we recommend an initial BioRBC exposure volume of ≤10 mL and label densities of ≤18 µg/mL.

Exercise intolerance in kidney diseases: physiological contributors and therapeutic strategies.

Exertional fatigue, defined as the overwhelming and debilitating sense of sustained exhaustion that impacts the ability to perform activities of daily living, is highly prevalent in chronic kidney disease (CKD) and end-stage renal disease (ESRD). Subjective reports of exertional fatigue are paralleled by objective measurements of exercise intolerance throughout the spectrum of the disease. The prevalence of exercise intolerance is clinically noteworthy, as it leads to increased frailty, worsened quality of life, and an increased risk of mortality. The physiological underpinnings of exercise intolerance are multifaceted and still not fully understood. This review aims to provide a comprehensive outline of the potential physiological contributors, both central and peripheral, to kidney disease-related exercise intolerance and highlight current and prospective interventions to target this symptom. In this review, the CKD-related metabolic derangements, cardiac and pulmonary dysfunction, altered physiological responses to oxygen consumption, vascular derangements, and sarcopenia are discussed in the context of exercise intolerance. Lifestyle interventions to improve exertional fatigue, such as aerobic and resistance exercise training, are discussed, and the lack of dietary interventions to improve exercise tolerance is highlighted. Current and prospective pharmaceutical and nutraceutical strategies to improve exertional fatigue are also broached. An extensive understanding of the pathophysiological mechanisms of exercise intolerance will allow for the development of more targeted therapeutic approached to improve exertional fatigue and health-related quality of life in CKD and ESRD.

Physical activity, health-related quality of life, and adjuvant endocrine therapy-related symptoms in women with hormone receptor-positive breast cancer.

BACKGROUND: Physical activity (PA) is recommended for women with breast cancer (BC); however, data are sparse on the association of PA with quality of life (QOL) and patient-reported symptoms for women on adjuvant endocrine therapy (AET). METHODS: Women with hormone receptor-positive BC who were taking AET completed standardized surveys about their health-related QOL, AET-related symptoms, and levels of PA using validated measures. A Wald chi-square test and an analysis of variance were used to assess associations with PA and independent variables. Generalized linear regression analyses assessed associations between PA, QOL, and AET-related symptoms. RESULTS: The analytic cohort included 485 Black and White women. Black race, a high body mass index (BMI), and being on aromatase inhibitors (vs tamoxifen) were associated with lower PA in a bivariate analysis. In a multivariate analysis, lower self-reported PA was associated with a high BMI (P = .02) and chemotherapy uptake (P = .006). Better health-related QOL (P = .01), less severe overall AET-related symptoms (P = .02), and less severe gynecological symptoms (P = .03) were associated with increasing levels of moderate PA. CONCLUSIONS: Among women taking AET, moderate levels of PA may be associated with fewer medication-related symptoms and overall better ratings of health-related QOL. Because of the low levels of PA observed in the sample overall and particularly for Black women, identifying successful strategies to promote PA are needed.

Enhanced Efficacy of Combination of Gemcitabine and Phosphatidylserine-Targeted Nanovesicles against Pancreatic Cancer.

Phosphatidylserine (PS) is often externalized in viable pancreatic cancer cells and is therapeutically targetable using PS-selective drugs. One of the first-line treatments for advanced pancreatic cancer disease, gemcitabine (GEM), provides only marginal benefit to patients. We therefore investigated the therapeutic benefits of combining GEM and the PS-targeting drug, saposin C-dioleoylphosphatidylserine (SapC-DOPS), for treating pancreatic ductal adenocarcinoma (PDAC). Using cell-cycle analyses and a cell surface PS-based sorting method in vitro, we observed an increase in surface PS as cells progress through the cell cycle from G1 to G2/M. We also observed that GEM treatment preferentially targets G1 phase cells that have low surface PS, resulting in an increased median surface PS level of PDAC cells. Inversely, SapC-DOPS preferentially targets high surface PS cells that are predominantly in the G2/M phase. Finally, combination therapy in subcutaneous and orthotopic PDAC tumors in vivo with SapC-DOPS and GEM or Abraxane (Abr)/GEM (one of the current standards of care) significantly inhibits tumor growth and increases survival compared with individual treatments. Our studies confirm a surface PS and cell cycle-based enhancement of cancer cytotoxicity following SapC-DOPS treatment in combination with GEM or Abr/GEM. Thus, PDAC patients treated with Abr/GEM may benefit from concurrent administration of SapC-DOPS.

Enhanced phosphatidylserine-selective cancer therapy with irradiation and SapC-DOPS nanovesicles.

Normal living cells exhibit phosphatidylserine (PS) primarily within the intracellular leaflet of the plasma membrane. In contrast, viable cancer cells have high levels of PS on the external surface, and exhibit a broad range of surface PS, even within specific types of cancer. Agents that target surface PS have recently been developed to treat tumors and are expected to be more effective with higher surface PS levels. In this context, we examined whether surface PS is increased with irradiation. In vitro irradiation of cancer cell lines selected surviving cells that had higher surface PS in a dose- and time-dependent manner. This was more pronounced if surface PS was initially in the lower range for cancer cells. Radiation also increased the surface PS of tumor cells in subcutaneous xenografts in nude mice. We found an inverse relationship between steady state surface PS level of cancer cell lines and their sensitivity to radiation-induced cell death. In addition, serial irradiation, which selected surviving cells with higher surface PS, also increased resistance to radiation and to some chemotherapeutic drugs, suggesting a PS-dependent mechanism for development of resistance to therapy. On the other hand, fractionated radiation enhanced the effect of a novel anti-cancer, PS-targeting drug, SapC-DOPS, in some cancer cell lines. Our data suggest that we can group cancer cells into cells with low surface PS, which are sensitive to radiation, and high surface PS, which are sensitive to SapC-DOPS. Combination of these interventions may provide a potential new combination therapy.

Development, validation, and potential applications of biotinylated red blood cells for posttransfusion kinetics and other physiological studies: evidenced-based analysis and recommendations.

The current reference method in the United States for measuring in vivo population red blood cell (RBC) kinetics utilizes chromium-51 (51 Cr) RBC labeling for determining RBC volume, 24-hour posttransfusion RBC recovery, and long-term RBC survival. Here we provide evidence supporting adoption of a method for kinetics that uses the biotin-labeled RBCs (BioRBCs) as a superior, versatile method for both regulatory and investigational purposes. RBC kinetic analysis using BioRBCs has important methodologic, analytical, and safety advantages over 51 Cr-labeled RBCs. We critically review recent advances in labeling human RBCs at multiple and progressively lower biotin label densities for concurrent, accurate, and sensitive determination of both autologous and allogeneic RBC population kinetics. BioRBC methods valid for RBC kinetic studies, including successful variations used by the authors, are presented along with pharmacokinetic modeling approaches for the accurate determination of RBC pharmacokinetic variables in health and disease. The advantages and limitations of the BioRBC method-including its capability of determining multiple BioRBC densities simultaneously in the same individual throughout the entire RBC life span-are presented and compared with the 51 Cr method. Finally, potential applications and limitations of kinetic BioRBC determinations are discussed.

Antibodies to biotinylated red blood cells in adults and infants: improved detection, partial characterization, and dependence on red blood cell-biotin dose.

BACKGROUND: Biotin-labeled red blood cells (BioRBCs) are used for in vivo kinetic studies. Because BioRBC dosing occasionally induces antibodies, a sensitive and specific anti-BioRBC detection assay is needed. STUDY DESIGN AND METHODS: Aims were to 1) develop a gel card assay to evaluate existing, naturally occurring and BioRBC-induced plasma antibodies, 2) compare gel card and tube agglutination detection results, and 3) test for a relationship of antibody induction and BioRBC dose. Reagent BioRBCs were prepared using sulfo-NHS biotin ranging from densities 18 (BioRBC-18) to 1458 (BioRBC-1458) µg/mL RBCs. RESULTS: Among BioRBC-exposed subjects, gel card and tube agglutination results were concordant in 21 of 22 adults and all 19 infant plasma samples. Gel card antibody detection sensitivity was more than 10-fold greater than tube agglutination. Twelve to 16 weeks after BioRBC exposure, induced anti-antibodies were detected by gel card in three of 26 adults (12%) at reagent densities BioRBC-256 or less, but in none of 41 infants. Importantly, induced anti-BioRBC antibodies were associated with higher BioRBC dose (p = 0.008); no antibodies were detected in 18 subjects who received BioRBC doses less than or equal to BioRBC-18. For noninduced BioRBC antibodies, six of 1125 naïve adults (0.3%) and none of 46 naïve infants demonstrated existing anti-BioRBC antibodies using reagent BioRBC-140 or -162. Existing anti-BioRBCs were all neutralized by biotin compounds, while induced antibodies were not. CONCLUSIONS: The gel card assay is more sensitive than the tube agglutination assay. We recommend reagent BioRBC-256 for identifying anti-BioRBCs. Use of a low total RBC biotin label dose (≤ BioRBC-18) may minimize antibody induction.

Assessment of Cardiopulmonary Responses to Treadmill Walking Following Gastric Bypass Surgery.

BACKGROUND: Studies that have evaluated cardiopulmonary responses to exercise within the first few months of bariatric surgery have utilized cycle ergometry. However, walking is the most commonly reported mode of both pre- and post-operative PA. The divergent cardiopulmonary responses and metabolic costs of weight-bearing (walking) and non-weight-bearing (cycling) exercises warrant examination of the effects of bariatric surgery on cardiopulmonary responses during walking. METHODS: Nine women completed a maximal cardiopulmonary exercise test on a treadmill 2 weeks before and 3 months after gastric bypass surgery (GBS). Heart rate (HR), oxygen uptake (VO2), oxygen pulse (O2-p), and time to fatigue were compared before and after surgery and between the GBS group and a comparison group of 12 normal-weight (NW) women who completed the same exercise testing protocol. RESULTS: Time to fatigue increased by ~140 s following GBS (p = 0.018). No other parameter improved during maximal exercise from pre- to post-surgery. Body weight- and fat-free mass-corrected VO2 and O2-p at peak exercise differed between the GBS and NW groups before surgery, while only weight-corrected values were different following surgery. These differences disappeared after controlling for body fat percentage. CONCLUSION: We have demonstrated that weight loss alone was not sufficient to improve select cardiopulmonary fitness measures during treadmill walking in obese females 3 months after GBS. However, we did observe a significant overall improvement in exercise capacity as the GBS group was able to exercise longer, presumably due to significant reductions in body mass and a subsequent reduced metabolic cost of walking.

Biochemical surrogate markers of hemolysis do not correlate with directly measured erythrocyte survival in sickle cell anemia.

Hemolysis is a key feature of sickle cell anemia (HbSS). Direct quantitation of hemolysis could be used as an objective outcome in clinical trials of new therapeutics for HbSS and would also enable better human studies of the pathogenesis of complications of HbSS that are ostensibly hemolysis-related, such as pulmonary hypertension. However, contemporary human studies in HbSS have used only surrogate markers of hemolysis rather than direct measurements of RBC survival. We directly quantified hemolysis in HbSS by measuring survival of an age cohort of RBCs labeled with a stable isotope, administered orally as 15 N-glycine, a metabolic precursor of heme. The atomic excess of 15 N in heme extracted from blood was monitored by mass spectrometry over time. We performed 13 labeling experiments in 11 individuals with HbSS. Mean RBC survival was 31.9 days (range 14.1-53.6). Both HbF level, a known determinant of hemolysis, and absolute reticulocyte count (ARC), an index of the marrow's response to hemolysis, correlated with directly measured RBC survival (r = 0.61, P < 0.002; r = -0.84, P < 0.001). However, commonly used biochemical surrogates of hemolysis (LDH, AST, bilirubin, and plasma free hemoglobin) did not correlate with directly measured RBC survival. These biochemical surrogates should be interpreted cautiously, at best, in clinical trials and human physiologic studies in HbSS. ARC was the best correlate of total hemolysis, but only 70% of the variation in RBC survival was reflected in this marker. If greater accuracy is required in human studies, 15 N-glycine RBC labeling can directly and accurately quantify hemolysis. Am. J. Hematol. 91:1195-1201, 2016. © 2016 Wiley Periodicals, Inc.

Changes in Resting Energy Expenditure in Relation to Body Weight and Composition Following Gastric Restriction: A Systematic Review.

In comparison to gastric bypass surgery, gastric restriction without malabsorption more closely simulates dietary adherence while still producing durable weight loss. The latter is achieved despite considerable reductions in resting energy expenditure (REE), and whether REE is adjusted for body weight/composition using ratio- or regression-based methods could influence understanding of how these procedures affect energy balance. This systematic review identified studies that reported REE before and after gastric restriction in order to compare changes using each method. Ratio assessments revealed increases and decreases when REE was expressed per kilogram of body weight and per kilogram of fat-free mass, respectively. In comparison, measured REE tended to be less than predicted from linear regression after surgery. Explanations for these seemingly disparate findings and future directions are discussed.

Red Blood Cell Dysfunction Induced by High-Fat Diet: Potential Implications for Obesity-Related Atherosclerosis.

BACKGROUND: High-fat diet (HFD) promotes endothelial dysfunction and proinflammatory monocyte activation, which contribute to atherosclerosis in obesity. We investigated whether HFD also induces the dysfunction of red blood cells (RBCs), which serve as a reservoir for chemokines via binding to Duffy antigen receptor for chemokines (DARC). METHODS AND RESULTS: A 60% HFD for 12 weeks, which produced only minor changes in lipid profile in C57/BL6 mice, markedly augmented the levels of monocyte chemoattractant protein-1 bound to RBCs, which in turn stimulated macrophage migration through an endothelial monolayer. Levels of RBC-bound KC were also increased by HFD. These effects of HFD were abolished in DARC(-/-) mice. In RBCs from HFD-fed wild-type and DARC(-/-) mice, levels of membrane cholesterol and phosphatidylserine externalization were increased, fostering RBC-macrophage inflammatory interactions and promoting macrophage phagocytosis in vitro. When labeled ex vivo and injected into wild-type mice, RBCs from HFD-fed mice exhibited ≈3-fold increase in splenic uptake. Finally, RBCs from HFD-fed mice induced increased macrophage adhesion to the endothelium when they were incubated with isolated aortic segments, indicating endothelial activation. CONCLUSIONS: RBC dysfunction, analogous to endothelial dysfunction, occurs early during diet-induced obesity and may serve as a mediator of atherosclerosis. These findings may have implications for the pathogenesis of atherosclerosis in obesity, a worldwide epidemic.

Variation in human cancer cell external phosphatidylserine is regulated by flippase activity and intracellular calcium.

Viable cancer cells expose elevated levels of phosphatidylserine (PS) on the exoplasmic face of the plasma membrane. However, the mechanisms leading to elevated PS exposure in viable cancer cells have not been defined. We previously showed that externalized PS may be used to monitor, target and kill tumor cells. In addition, PS on tumor cells is recognized by macrophages and has implications in antitumor immunity. Therefore, it is important to understand the molecular details of PS exposure on cancer cells in order to improve therapeutic targeting. Here we explored the mechanisms regulating the surface PS exposure in human cancer cells and found that differential flippase activity and intracellular calcium are the major regulators of surface PS exposure in viable human cancer cells. In general, cancer cell lines with high surface PS exhibited low flippase activity and high intracellular calcium, whereas cancer cells with low surface PS exhibited high flippase activity and low intracellular calcium. High surface PS cancer cells also had higher total cellular PS than low surface PS cells. Together, our results indicate that the amount of external PS in cancer cells is regulated by calcium dependent flippase activity and may also be influenced by total cellular PS.

SapC-DOPS nanovesicles induce Smac- and Bax-dependent apoptosis through mitochondrial activation in neuroblastomas.

BACKGROUND: High toxicity, morbidity and secondary malignancy render chemotherapy of neuroblastoma inefficient, prompting the search for novel compounds. Nanovesicles offer great promise in imaging and treatment of cancer. SapC-DOPS, a stable nanovesicle formed from the lysosomal protein saposin C and dioleoylphosphatidylserine possess strong affinity for abundantly exposed surface phosphatidylserine on cancer cells. Here, we show that SapC-DOPS effectively targets and suppresses neuroblastoma growth and elucidate the molecular mechanism of SapC-DOPS action in neuroblastoma in vitro. METHODS: In vivo targeting of neuroblastoma was assessed in xenograft mice injected intravenously with fluorescently-labeled SapC-DOPS. Xenografted tumors were also used to demonstrate its therapeutic efficacy. Apoptosis induction in vivo was evaluated in tumor sections using the TUNEL assay. The mechanisms underlying the induction of apoptosis by SapC-DOPS were addressed through measurements of cell viability, mitochondrial membrane potential (ΔΨM), flow cytometric DNA fragmentation assays and by immunoblot analysis of second mitochondria-derived activator of caspases (Smac), Bax, Cytochrome c (Cyto c) and Caspase-3 in the cytosol or in mitochondrial fractions of cultured neuroblastoma cells. RESULTS: SapC-DOPS showed specific targeting and prevented the growth of human neuroblastoma xenografts in mice. In neuroblastoma cells in vitro, apoptosis occurred via a series of steps that included: (1) loss of ΔΨM and increased mitochondrial superoxide formation; (2) cytosolic release of Smac, Cyto c, AIF; and (3) mitochondrial translocation and polymerization of Bax. ShRNA-mediated Smac knockdown and V5 peptide-mediated Bax inhibition decreased cytosolic Smac and Cyto c release along with caspase activation and abrogated apoptosis, indicating that Smac and Bax are critical mediators of SapC-DOPS action. Similarly, pretreatment with the mitochondria-stabilizing agent bongkrekic acid decreased apoptosis indicating that loss of ΔΨM is critical for SapC-DOPS activity. Apoptosis induction was not critically dependent on reactive oxygen species (ROS) production and Cyclophilin D, since pretreatment with N-acetyl cysteine and cyclosporine A, respectively, did not prevent Smac or Cyto c release. CONCLUSIONS: Taken together, our results indicate that SapC-DOPS acts through a mitochondria-mediated pathway accompanied by an early release of Smac and Bax. Specific tumor-targeting capacity and anticancer efficacy of SapC-DOPS supports its potential as a dual imaging and therapeutic agent in neuroblastoma therapy.

Activation of protein kinase C by phorbol ester increases red blood cell scramblase activity and external phosphatidylserine.

Externalization of phosphatidylserine (PS) is thought to contribute to sickle cell disease (SCD) pathophysiology. The red blood cell (RBC) aminophospholipid translocase (APLT) mediates the transport of PS from the outer to the inner RBC membrane leaflet to maintain an asymmetric distribution of PL, while phospholipid scramblase (PLSCR) equilibrates PL across the RBC membrane, promoting PS externalization. We previously identified an association between PS externalization level and PLSCR activity in sickle RBC under basal conditions. Other studies showed that activation of protein kinase C (PKC) by PMA (phorbol-12-myristate-13-acetate) causes increased external PS on RBC. Therefore, we hypothesized that PMA-activated PKC stimulates PLSCR activity in RBC and thereby contributes to increased PS externalization. In the current studies, we show that PMA treatment causes immediate and variable PLSCR activation and subsequent PS externalization in control and sickle RBC. While TfR+ sickle reticulocytes display some endogenous PLSCR activity, we observed a robust activation of PLSCR in sickle reticulocytes treated with PMA. The PKC inhibitor, chelerythrine (Chel), significantly inhibited PMA-dependent PLSCR activation and PS externalization. Chel also inhibited endogenous PLSCR activity in sickle reticulocytes. These data provide evidence that PKC mediates PS externalization in RBC through activation of PLSCR.

Use of an oral stable isotope label to confirm variation in red blood cell mean age that influences HbA1c interpretation.

HbA1c is commonly used to monitor glycemic control. However, there is growing evidence that the relationship between HbA1c and mean blood glucose (MBG) is influenced by variation in red blood cell (RBC) lifespan in hematologically normal individuals. Correction of HbA1c for mean RBC age (MRBC ) requires a noninvasive, accurate, and affordable method to measure RBC survival. In this study, we evaluated whether a stable isotope approach would satisfy these requirements. RBC lifespan and MRBC were determined in a group of nine hematologically normal diabetic and nondiabetic subjects using oral (15) N-glycine to label heme in an age cohort of RBC. The MRBC was 58.7 ± 9.1 (2SD) days and RBC lifespan was 106 ± 21 (2SD) days. This degree of variation (±15-20%) is consistent with previous studies using other techniques. In a subset of seven subjects, MRBC determined with the biotin label technique were available from approximately five years prior, and strongly correlated with the stable isotope values (R(2) = 0.79). This study suggests that the MRBC is stable over time but varies substantially among individuals, and supports the importance of its variation in HbA1c interpretation. The characteristics of the stable isotope method support its suitability for studies to directly evaluate the impact of variation in MRBC on the interpretation of HbA1c.