Weight History Correlates of Resting Energy Expenditure in Women With Bulimia Nervosa.

OBJECTIVE: Among those with bulimia nervosa, weight suppression has been associated with illness severity and treatment prognosis. Although significant weight loss is known to reduce metabolic rate, the relation between weight suppression and resting energy expenditure (REE) in bulimia nervosa has not been examined. This study tested the hypothesis of an inverse relation between weight suppression and REE in a sample of women with bulimia nervosa (N = 84). METHODS: In primary analyses, linear regressions were conducted between weight suppression and REE, corrected for fat-free mass. In follow-up, exploratory analyses, stepwise linear regressions were conducted to explore the main and interaction effects of weight history and weight suppression on REE. RESULTS: Neither traditional (TWS) nor developmental weight suppression (DWS) correlated with REE. Results from exploratory analyses, however, revealed a medium-to-large inverse relation between several weight history variables and REE (highest past weight, sr2 = 0.05; lowest postmorbid weight, sr2 = 0.07; current weight, sr2 = 0.05). Additionally, DWS interacted with current (sr2 = 0.08) and highest premorbid (sr2 = 0.05) z-BMI to influence REE with a medium-to-large effect. For individuals low in current and premorbid z-BMIs, higher DWS associated with lower REE levels. However, for individuals at higher premorbid z-BMIs, higher DWS unexpectedly associated with greater REE levels. DISCUSSION: In this sample of women with bulimia nervosa, reduced REE associated with higher weights across all timepoints. If the interaction effect between DWS and z-BMI history persists in future studies, this may indicate unique challenges faced by individuals low in z-BMI and high in DWS related to weight gain and normalization of eating.

Physiological, body composition, and body mass measures show that a developmental measure of weight suppression is more valid than the traditional measure.

OBJECTIVE: The traditional measure of weight suppression (TWS; the difference between an individual's highest past weight at adult height and current weight), has been associated with many psychological, behavioral and biological variables in those with eating disorders. A new measure of weight suppression, called developmental weight suppression (DWS), corrects two major problems in the original measure. Initial research indicates that DWS represents a superior operationalization of the construct weight suppression was originally designed to measure (Lowe [1993, Psychol Bull, 114: 100]). This study is the first to examine the relation between both WS measures and weight history, body composition and a variety of metabolic hormones. METHODS: Data were collected in 91 women with bulimia nervosa (BN) or BN-spectrum disorders. RESULTS: Both weight suppression indices were related to multiple hormones. However, multiple regression analyses showed that the independent effects of DWS differed from the independent effects of TWS in that only DWS was negatively related to: (1) current z-BMI, (2) body fat percentage, and (3) insulin, leptin, T3 free, and TSH. This differential pattern also occurred when results were corrected for multiple comparisons. DISCUSSION: Findings provide stronger biological support for the construct validity of DWS than TWS and suggest that: (1) from the perspective of individuals with BN, high DWS embodies success at food restriction and weight loss, (2) elevated DWS may trap individuals with BN in a powerful biobehavioral bind, and (3) DWS is the preferred measure of weight suppression in future research on eating disorders. PUBLIC SIGNIFICANCE: Most individuals with bulimia nervosa lose substantial weight in the process of developing their disorder. Such weight suppression is related to many characteristics of those with the eating disorder bulimia nervosa. This study shows why a new measure of weight suppression, based on an individual's growth during development, is more biologically valid than the traditional measure of weight suppression.

Testing associations between assessments of cognitive flexibility and eating disorder symptoms in adolescent bulimia nervosa.

OBJECTIVE: Cognitive rigidity, or difficulty adapting to changing demands, is commonly observed in anorexia nervosa. Less is known, however, about cognitive flexibility (CF) in bulimia nervosa (BN) and, particularly, adolescence. Clarifying this relation and best assessment practices may guide informed clinical decision-making. The current study compared how two measures of CF (i.e., Wisconsin Card Sort Task [WCST] and Trail Making Task [TMT]) relate to BN symptoms among adolescents. METHODS: Data from a subsample (n = 78) of adolescents with BN were analyzed. Linear and hurdle regressions were used to compare the effects of WCST perseverative errors and TMT performance on Eating Disorder Examination Global Scores, objective binge episodes, and self-induced vomiting episodes (SVEs) at baseline and end-of-treatment (EOT). RESULTS: Neither CF measure associated with baseline BN symptoms. TMT performance positively associated with the likelihood of engaging in SVEs at EOT (𝛽 = 0.47, p = 0.01, 95% confidence interval [CI] = [0.11-0.84]) and, among adolescents who endorsed ≥1 SVE at EOT, WCST perseverative errors (𝛽 = 0.05, p = 0.005, 95% CI = [0.01-0.08]) positively associated with SVE frequency at EOT. DISCUSSION: The overall lack of associations between CF and outcomes suggests that cognitive rigidity may not be as relevant to the clinical profile of adolescent BN as for anorexia nervosa. In the few significant associations that emerged, the WCST and TMT uniquely predicted the severity of vomiting at EOT in this sample. Given the lack of CF deficits, future work should aim to test the role of other executive functions (e.g., impulsivity), in addition to CF, to determine which deficits are present in adolescent BN and may predict outcomes. PUBLIC SIGNIFICANCE: Patients with eating disorders often have difficulties thinking flexibly, which may interfere with their recovery. We tested two ways of measuring flexible thinking in adolescents with BN. Overall, flexible thinking was not associated with symptom-level outcomes. However, less flexible thinking at the start of treatment predicted self-induced vomiting at EOT. If findings are replicable, then assessing and addressing flexible thinking could improve outcomes for adolescents with BN.

A comparison of traditional and developmental measures of weight suppression in residential patients with bulimia nervosa.

OBJECTIVE: Weight suppression (WS) is associated with many eating disorder (ED)-related symptoms. However, traditional calculations of WS do not consider the age or height at which one's highest past weight was reached. Lowe et al. (2022) found that developmental WS (DWS) was associated with a wider variety of ED-related symptoms compared with traditional WS (TWS). This study replicated and extended these findings in a larger sample of individuals with bulimia nervosa (BN) at a residential ED treatment center. METHODS: Participants were 1051 female patients with BN. We examined the relations between each WS measure and ED symptoms, emotional symptoms, and weight history variables. RESULTS: TWS and DWS showed a similar number of relations with ED-related symptoms. DWS was positively related to behavioral symptoms (e.g., vomiting), and negatively related to cognitive symptoms (e.g., weight/eating concern). TWS was positively related to highest premorbid, highest postmorbid, and lowest postmorbid weights. DWS was also positively related to highest premorbid z-scored body mass index (zBMI), but negatively related to lowest and highest postmorbid zBMI. CONCLUSIONS: DWS, relative to TWS, may better capture the psychobiological impact of the weight discrepancy that a measure of WS is meant to reflect. PUBLIC SIGNIFICANCE: Weight suppression, the difference between an individual's past highest weight and current weight, is significantly related to many ED-related symptoms. This study found that a new weight suppression measure, based on expected weight-for-height during physical development, relates to ED characteristics in a different manner from the traditional measure of weight suppression, showing positive associations with behavioral symptoms and negative associations with cognitive symptoms.

Association between maternal eating and young child feeding in a community sample.

BACKGROUND: Early childhood is a pivotal period for the development of healthy eating practices. One way to promote child health is to identify early modifiable factors that affect child eating and weight. Given the intergenerational transmission of eating behaviors, this study examined how mothers' eating behaviors were associated with child feeding practices, and whether child weight-for-length (z-WFL) moderated this relation, in a community sample. METHODS: Participants were 72 mother-child dyads. Maternal eating behaviors-emotional, external and restrained-were assessed 9-months postpartum, using the Dutch Eating Behavior Questionnaire. Child feeding-restrictive, pressure, and concern about overeating/overweight or undereating/underweight-was measured using the Infant Feeding Questionnaire, and child z-WFL were assessed 18-months postpartum. Linear regressions were used to test the main effect of maternal eating and the interaction effect of maternal eating and child z-WFL, on child feeding practices. RESULTS: Maternal restrained eating was associated with child pressure feeding, and contrarily with concerns about overeating/overweight. However, a significant interaction between child z-WFL and both maternal emotional and external eating were found with regard to concern about child undereating/underweight. Paradoxically, among children who weighed more, greater maternal emotional and greater external eating were associated with greater concern about child undereating/underweight. CONCLUSIONS: In this community sample, mothers were more likely to report contradictory feeding practices and concerns, suggesting complicated relations among a mother's own eating behavior, her child's weight, and her perceptions of child eating and weight. This may indicate a need for better communication and support of infant feeding practices. TRIAL REGISTRATION: Data was collected as part of two grants (MAMAS Grant ID: HL097973-01; SEED Grant ID: HL116511-02) conducted at the University of California, San Francisco (UCSF). All subjects gave their informed consent for inclusion before they participated in the study. The study was conducted in accordance with the Declaration of Helsinki, and the protocol was approved by institutional review board at UCSF.

Bruton's tyrosine kinase inhibition induces rewiring of proximal and distal B-cell receptor signaling in mice.

Bruton's tyrosine kinase (Btk) is a crucial signaling molecule in BCR signaling and a key regulator of B- cell differentiation and function. Btk inhibition has shown impressive clinical efficacy in various B-cell malignancies. However, it remains unknown whether inhibition additionally induces changes in BCR signaling due to feedback mechanisms, a phenomenon referred to as BCR rewiring. In this report, we studied the impact of Btk activity on major components of the BCR signaling pathway in mice. As expected, NF-κB and Akt/S6 signaling was decreased in Btk-deficient B cells. Unexpectedly, phosphorylation of several proximal signaling molecules, including CD79a, Syk, and PI3K, as well as the key Btk-effector PLCγ2 and the more downstream kinase Erk, were significantly increased. This pattern of BCR rewiring was essentially opposite in B cells from transgenic mice overexpressing Btk. Importantly, prolonged Btk inhibitor treatment of WT mice or mice engrafted with leukemic B cells also resulted in increased phosho-CD79a and phospho-PLCγ2 in B cells. Our findings show that Btk enzymatic function determines phosphorylation of proximal and distal BCR signaling molecules in B cells. We conclude that Btk inhibitor treatment results in rewiring of BCR signaling, which may affect both malignant and healthy B cells.

Overexpression of SH2-Containing Inositol Phosphatase Contributes to Chronic Lymphocytic Leukemia Survival.

Balanced activity of kinases and phosphatases downstream of the BCR is essential for B cell differentiation and function and is disturbed in chronic lymphocytic leukemia (CLL). In this study, we employed IgH.TEµ mice, which spontaneously develop CLL, and stable EMC CLL cell lines derived from these mice to explore the role of phosphatases in CLL. Genome-wide expression profiling comparing IgH.TEµ CLL cells with wild-type splenic B cells identified 96 differentially expressed phosphatase genes, including SH2-containing inositol phosphatase (Ship2). We found that B cell-specific deletion of Ship2, but not of its close homolog Ship1, significantly reduced CLL formation in IgH.TEµ mice. Treatment of EMC cell lines with Ship1/2 small molecule inhibitors resulted in the induction of caspase-dependent apoptosis. Using flow cytometry and Western blot analysis, we observed that blocking Ship1/2 abrogated EMC cell survival by exerting dual effects on the BCR signaling cascade. On one hand, specific Ship1 inhibition enhanced calcium signaling and thereby abrogated an anergic response to BCR stimulation in CLL cells. On the other hand, concomitant Ship1/Ship2 inhibition or specific Ship2 inhibition reduced constitutive activation of the mTORC1/ribosomal protein S6 pathway and downregulated constitutive expression of the antiapoptotic protein Mcl-1, in both EMC cell lines and primary IgH.TEµ CLL cells. Importantly, also in human CLL, we found overexpression of many phosphatases including SHIP2. Inhibition of SHIP1/SHIP2 reduced cellular survival and S6 phosphorylation and enhanced basal calcium levels in human CLL cells. Taken together, we provide evidence that SHIP2 contributes to CLL pathogenesis in mouse and human CLL.

Microscope-Integrated Optical Coherence Tomography-Guided Autologous Full-Thickness Neurosensory Retinal Autograft for Large Macular Hole-Related Total Retinal Detachment.

PURPOSE: To evaluate the feasibility and utility of microscope-integrated optical coherence tomography in patients undergoing full-thickness neurosensory retinal autograft for refractory macular hole (MH)-associated retinal detachment. METHODS: We analyzed two eyes of two patients who had undergone a neurosensory retinal autograft for large MH associated retinal detachment. Both cases had microscope-integrated optical coherence tomography-guided placement and sizing of the retinal autograft. Time taken for obtaining microscope-integrated optical coherence tomography images, morphology of the retinal autograft (intraoperative and postoperative), and anatomic and functional outcomes were noted. RESULTS: The first case had optic disc pit-related maculopathy with a large MH and total retinal detachment. She had undergone a vitrectomy with internal limiting membrane peeling elsewhere. The second patient had a treatment-naive large MH with total retinal detachment. Both patients underwent vitrectomy with microscope-integrated optical coherence tomography-guided autologous neurosensory retinal autograft placement and silicone oil tamponade. At 6 months and 3 months follow-up, respectively, both patients had closed MHs, attached retinas, and improvement in visual acuity. CONCLUSION: Microscope-integrated optical coherence tomography provides intraoperative visualization of MHs and provides real-time feedback regarding dimensions of the retinal autograft, thus aiding in accurate sizing of the graft. This ensures that the autograft fits snugly in the MH, thereby restoring macular structure and improving visual acuity.

MULTICOLOR CONFOCAL SCANNING LASER OPHTHALMOSCOPE IMAGING IN POSTERIOR UVEITIS.

PURPOSE: To study the utility of MultiColor confocal scanning laser ophthalmoscope imaging (MCI) in identifying the morphology of uveitic lesions compared with conventional color fundus photography (CFP) in patients with posterior uveitis. METHODS: In this prospective observational study, subjects with posterior uveitis underwent MCI and CFP. The images obtained by the two modalities were analyzed by two independent reviewers for vitreoretinal surface abnormalities, retinal fluid and hemorrhages, and depth/location of lesions. These findings were compared with the clinical findings and other imaging techniques. RESULTS: Sixty-nine eyes of 43 patients (25 men) with mean age of 33.5 ± 13.9 years were studied. MultiColor imaging had better sensitivity and specificity in detecting vitreoretinal interface abnormalities, such as epiretinal membrane and inner retinal striae, compared with CFP. MultiColor imaging failed to detect retinochoroiditis lesions in 5 of 6 eyes (83%) and choroiditis in 9 46 eyes (20%), which were detected on CFP and clinical examination. Also, MCI showed a high false-positive rate of 34% in detecting intraretinal hemorrhages. CONCLUSION: Retinochoroidal lesions in posterior uveitis may be poorly identified on MCI compared with CFP and clinical examination. One must exercise caution in commenting on disease morphology based on MCI alone.

Plentiful melanin pigment containing histiocyte-like cells in Coats disease: Awareness avoids diagnostic pitfall.

Coats disease is an exudative retinal vasculopathy characterised by presence of yellow-golden deposits in the retina and retinal detachment. Subretinal fluid drainage performed as a part of therapeutic management makes the fluid amenable to cytological examination. Infection by Toxoplasma may closely simulate the ocular symptoms seen in Coats disease. Awareness of the cytological findings in Coats disease helps to clinch accurate diagnosis. We herein present a case of Coats disease with many histiocyte-like cells with plentiful intracytoplasmic melanin pigment in cytology smears from subretinal fluid, where cytological diagnosis was challenging and a correct diagnosis was made with the aid of ancillary techniques.

Traditional versus developmental measures of weight suppression: Exploring their relationships with bulimic psychopathology.

OBJECTIVE: Weight suppression (WS) is related to a wide variety of eating disorder characteristics. However, individuals with eating disorders usually reach their highest premorbid weight while still developing physically. Therefore, a more sensitive index of individual differences in highest premorbid weight may be one that compares highest premorbid z-BMI to current z-BMI (called developmental weight suppression [DWS] here). METHOD: In this exploratory study, we compared the relationships between traditional weight suppression (TWS) and DWS and a variety of measures related to bulimic psychopathology in 91 females (M age, 25.2; 60.5% White), with clinical or sub-clinical bulimia nervosa. RESULTS: TWS and DWS were correlated (r = 0.40, p < 0.001). TWS was only significantly related to a measure of physical activity whereas DWS was related to 14 outcomes. DWS showed consistent positive relations with behavioural outcomes (e.g., binge eating) but consistent negative relations with cognitive/affective outcomes (e.g., weight concerns). CONCLUSIONS: Findings indicated much more consistent relationships between the novel DWS measure and bulimic characteristics than with the TWS measure. DWS showed both positive and negative relations with bulimic symptoms, though these findings require replication to confirm their validity. Consistent evidence indicated that the two WS measures served as mutual suppressor variables.

Within-subject weight variability in bulimia nervosa: Correlates and consequences.

OBJECTIVE: The oscillations between binge eating, purging, and dieting in bulimia nervosa (BN) may produce substantial within-subject weight variability. Although weight variability has been predictive of eating- and weight-related variables in community samples, it has not been empirically examined in eating disorders. The current study examined cross-sectional and prospective associations between weight variability and BN pathology. METHOD: Four weights were collected over an average of 42.02 days, and weight variability was calculated as the root mean square error around each individual's weight trajectory regression line. Linear regressions were performed to examine the association between weight variability and eating disorder psychopathology, cross-sectionally at baseline and prospectively at 6-month follow-up, adjusting for baseline BMI. RESULTS: Weight variability was cross-sectionally associated with eating pathology, but these relationships became non-significant after adjusting for BMI. However, at 6-month follow-up, greater baseline weight variability predicted increases in body dissatisfaction, shape and weight concerns, and global eating pathology, even after adjusting for baseline BMI. DISCUSSION: These findings demonstrate, for the first time, that within-subject weight variability predicts greater eating disorder pathology over time in BN. The results add to evidence that weight history variables contribute to BN psychopathology above and beyond well-documented psychological dysfunction in BN.

The impact of comorbid posttraumatic stress disorder on eating disorder treatment outcomes: Investigating the unified treatment model.

PURPOSE: Many women with eating disorders (EDs) have comorbid posttraumatic stress disorder (PTSD). However, there have been few studies on how comorbid PTSD may impact ED treatment outcomes. METHOD: Participants were 2,809 patients from residential ED treatment facilities who were treated using the Unified Treatment Model (UTM). We investigated whether PTSD diagnosis at admission was associated with changes in Eating Disorder Examination-Questionnaire (EDE-Q) scores, binge eating, self-induced vomiting, and restriction, across three time points, as well as clinically significant improvement and treatment drop-out. RESULTS: Using latent growth models, with time modeled as a second-order polynomial, we found that EDE-Q scores and behavioral symptoms decreased from admission to discharge, but increased from discharge to 6-month follow-up. PTSD diagnosis was associated with higher baseline EDE-Q scores and restriction, and lower binge-eating frequency. PTSD diagnosis was not associated with symptom change over time, treatment dropout, or clinically significant change. DISCUSSION: Although PTSD diagnoses were associated with higher ED symptom levels at admission, PTSD was not associated with worse treatment outcomes, suggesting the UTM is a promising treatment for patients with and without PTSD. Future studies should investigate the impact of ED treatment on PTSD symptoms in order to determine the need for integrated treatments for these comorbid conditions.

A new, developmentally-sensitive measure of weight suppression.

OBJECTIVE: Weight suppression (WS) has demonstrated associations with numerous indices of eating behavior, psychopathology and eating disorder prognosis. However, because WS has traditionally been measured as a simple subtraction of current weight from highest past weight at adult height, this calculation is problematic for most individuals with disordered eating, who usually reach their highest past weight during adolescence. Here we propose a new method for computing WS to address this shortcoming, termed "developmental weight suppression" (DWS), and provide a web-based tool for ease of calculation. METHOD: DWS is calculated as the difference between one's highest premorbid z-BMI (i.e., BMI z-score), and current z-BMI. z-BMIs were calculated using Cole's lambda-mu-sigma (LMS) approach, in accordance with LMS parameters publicly available from the Center for Disease Control (2010). A web-based user interface is available at https://niuxin.shinyapps.io/devws/, making its computation easier and its adoption by researchers simpler. DISCUSSION: By using z-BMIs in place of weights, DWS is more sensitive to the developmentally-relevant factors of age, height, and sex. Preliminary findings suggest that DWS is more strongly related to measures of eating pathology and biological reactions to weight loss than traditionally-computed WS, although more research is needed to test this hypothesis.

Correction to: Role of Bruton's tyrosine kinase in B cells and malignancies.

Following publication of the original article [1], the authors reported an error in Table 1.

Predictors of early response in conjoint and separated models of family-based treatment for adolescent anorexia nervosa.

OBJECTIVE: Early response, as indicated by early weight gain, in family-based treatment (FBT) for adolescent anorexia nervosa (AN) predicts remission at end of treatment. However, little is known about what factors contribute to early response. Further, no previous studies have examined early response to separated forms of FBT. METHOD: Data from a randomised clinical trial of conjoint FBT and separated FBT (parent-focused treatment, PFT) were analysed to examine the timing and amount of early weight gain that predicted remission and identify factors associated with early response. RESULTS: Weight gain of at least 2.80 kg in FBT (N = 55) and 2.28 kg in PFT (N = 51), by Session 5, were the best predictors of remission at end of treatment. Early response in FBT was predicted by greater paternal therapeutic alliance and lower paternal criticism. Early response in PFT was predicted by less severe eating-disorder symptoms and negative affect at baseline, lower maternal criticism, and greater adolescent therapeutic alliance. CONCLUSIONS: The results confirm that early weight gain is an important prognostic indicator in both conjoint FBT and PFT and suggest that addressing negative emotion, parental criticism, and therapeutic alliance early in treatment could improve remission rates.

Role of Bruton's tyrosine kinase in B cells and malignancies.

Bruton's tyrosine kinase (BTK) is a non-receptor kinase that plays a crucial role in oncogenic signaling that is critical for proliferation and survival of leukemic cells in many B cell malignancies. BTK was initially shown to be defective in the primary immunodeficiency X-linked agammaglobulinemia (XLA) and is essential both for B cell development and function of mature B cells. Shortly after its discovery, BTK was placed in the signal transduction pathway downstream of the B cell antigen receptor (BCR). More recently, small-molecule inhibitors of this kinase have shown excellent anti-tumor activity, first in animal models and subsequently in clinical studies. In particular, the orally administered irreversible BTK inhibitor ibrutinib is associated with high response rates in patients with relapsed/refractory chronic lymphocytic leukemia (CLL) and mantle-cell lymphoma (MCL), including patients with high-risk genetic lesions. Because ibrutinib is generally well tolerated and shows durable single-agent efficacy, it was rapidly approved for first-line treatment of patients with CLL in 2016. To date, evidence is accumulating for efficacy of ibrutinib in various other B cell malignancies. BTK inhibition has molecular effects beyond its classic role in BCR signaling. These involve B cell-intrinsic signaling pathways central to cellular survival, proliferation or retention in supportive lymphoid niches. Moreover, BTK functions in several myeloid cell populations representing important components of the tumor microenvironment. As a result, there is currently a considerable interest in BTK inhibition as an anti-cancer therapy, not only in B cell malignancies but also in solid tumors. Efficacy of BTK inhibition as a single agent therapy is strong, but resistance may develop, fueling the development of combination therapies that improve clinical responses. In this review, we discuss the role of BTK in B cell differentiation and B cell malignancies and highlight the importance of BTK inhibition in cancer therapy.

Outcome parameters associated with perceived helpfulness of family-based treatment for adolescent eating disorders.

OBJECTIVE: Family-based treatment (FBT) is an efficacious treatment for adolescent eating disorders, yet it is not routinely implemented in clinical practice. Given that consumers play a role in treatment selection, this study sought to examine families' perspectives on FBT and remission markers associated with increased treatment satisfaction across families. METHOD: Participants were 40 adolescents and 43 caregivers who received outpatient FBT. FBT helpfulness was assessed using a treatment follow-up questionnaire, and eating disorder symptomatology was assessed using percent expected body weight (%EBW) and the eating disorder examination (EDE). Regression analyses were used to assess whether changes in symptoms from baseline to end-of-treatment (EOT) were significantly associated with helpfulness reports. RESULTS: On average, patients and their parents perceived FBT as "quite helpful" and "extremely helpful," respectively. Improvements in all EDE subscales, with the exception of restraint, were significantly associated with adolescent report of helpfulness (all p < .05); increase in %EBW was significantly associated with maternal report of helpfulness (p = .03). There were no significant findings for paternal report. DISCUSSION: Both patients and their parents perceived FBT as helpful, but patients seemed to prioritize cognitive improvements while mothers prioritized physical improvements in rating their satisfaction with FBT.

Metabolic shift in density-dependent stem cell differentiation.

BACKGROUND: Vascular progenitor cells (VPCs) derived from embryonic stem cells (ESCs) are a valuable source for cell- and tissue-based therapeutic strategies. During the optimization of endothelial cell (EC) inductions from mouse ESCs using our staged and chemically-defined induction methods, we found that cell seeding density but not VEGF treatment between 10 ng/mL and 40 ng/mL was a significant variable directing ESCs into FLK1+ VPCs during stage 1 induction. Here, we examine potential contributions from cell-to-cell signaling or cellular metabolism in the production of VPCs from ESCs seeded at different cell densities. METHODS: Using 1D 1H-NMR spectroscopy, transcriptomic arrays, and flow cytometry, we observed that the density-dependent differentiation of ESCs into FLK1+ VPCs positively correlated with a shift in metabolism and cellular growth. RESULTS: Specifically, cell differentiation correlated with an earlier plateauing of exhaustive glycolysis, decreased lactate production, lower metabolite consumption, decreased cellular proliferation and an increase in cell size. In contrast, cells seeded at a lower density of 1,000 cells/cm2 exhibited increased rates of glycolysis, lactate secretion, metabolite utilization, and proliferation over the same induction period. Gene expression analysis indicated that high cell seeding density correlated with up-regulation of several genes including cell adhesion molecules of the notch family (NOTCH1 and NOTCH4) and cadherin family (CDH5) related to vascular development. CONCLUSIONS: These results confirm that a distinct metabolic phenotype correlates with cell differentiation of VPCs.