Delineating a Metabolomic Signature for the Transition from Gestational Diabetes to Type 2 Diabetes

Although gestational diabetes (GDM) is of a transient nature, up to 50% of women with GDM develop type 2 diabetes (T2D) within 5 years. Despite this extremely high risk, post-partum screening remains low. Here, we delineated a predictive metabolomic signature of disease transition. The study patients were part of the SWIFT Cohort, which follows 1010 GDM women, 109 of which developed T2D two years post-partum. Future cases were matched to future controls based on ethnicity, age and pre-pregnancy BMI. Hexoses, specific amino acids, specific sphingomyelin and specific phosphatidylcholine species associated with T2D. Through a J48 decision tree predictive modelling in the training set, future T2D incidence was predicted in an independent testing set with discriminative power 0.769 (PM.Sc.2017-01-12 00:00:0

Glucose transporter expression in an avian nectarivore: the ruby-throated hummingbird (Archilochus colubris).

Glucose transporter (GLUT) proteins play a key role in the transport of monosaccharides across cellular membranes, and thus, blood sugar regulation and tissue metabolism. Patterns of GLUT expression, including the insulin-responsive GLUT4, have been well characterized in mammals. However, relatively little is known about patterns of GLUT expression in birds with existing data limited to the granivorous or herbivorous chicken, duck and sparrow. The smallest avian taxa, hummingbirds, exhibit some of the highest fasted and fed blood glucose levels and display an unusual ability to switch rapidly and completely between endogenous fat and exogenous sugar to fuel energetically expensive hovering flight. Despite this, nothing is known about the GLUT transporters that enable observed rapid rates of carbohydrate flux. We examined GLUT (GLUT1, 2, 3, & 4) expression in pectoralis, leg muscle, heart, liver, kidney, intestine and brain from both zebra finches (Taeniopygia guttata) and ruby-throated hummingbirds (Archilochus colubris). mRNA expression of all four transporters was probed using reverse-transcription PCR (RT-PCR). In addition, GLUT1 and 4 protein expression were assayed by western blot and immunostaining. Patterns of RNA and protein expression of GLUT1-3 in both species agree closely with published reports from other birds and mammals. As in other birds, and unlike in mammals, we did not detect GLUT4. A lack of GLUT4 correlates with hyperglycemia and an uncoupling of exercise intensity and relative oxidation of carbohydrates in hummingbirds. The function of GLUTs present in hummingbird muscle tissue (e.g. GLUT1 and 3) remain undescribed. Thus, further work is necessary to determine if high capillary density, and thus surface area across which cellular-mediated transport of sugars into active tissues (e.g. muscle) occurs, rather than taxon-specific differences in GLUT density or kinetics, can account for observed rapid rates of sugar flux into these tissues

Glucose transporter staining in hummingbird and mouse liver.

<p>Sections of hummingbird (a) and mouse (b) liver tissue stained with GLUT1 primary antibody. Staining was visualized with a FITC-conjugated secondary antibody (green). Sections were counterstained with DAPI to visualize nuclei (blue).</p

GLUT amino acid sequence identities and similarities.

<p>Sequence identity and, in parentheses, similarity are listed for each paired comparison. Sequences for ruby-throated hummingbirds are extrapolated from an optimal alignment based on RT-PCR products of the partial cDNA sequence (see text). Sequences from all other species are those published in GendBank.</p

GLUT cDNA sequence identities.

<p>Sequences for ruby-throated hummingbird are based on partial gene products of RT-PCR reactions. Partial sequences based on zebra finch RT-PCR products showed >99% identity with published sequences in GenBank. Thus, sequences from zebra finches and other species listed used for comparison are taken from GenBank.</p

Glucose transporter staining in hummingbird and mouse skeletal muscle.

<p>Immunohistochemically-stained cross-sections of hummingbird pectoralis (a, c) and mouse gastrocnemius (b, d) muscle. Panels a and b) Immunostaining of tissues with GLUT1 primary antibody, visualized with a FITC-conjugated secondary antibody (green). Panels c and d) Immunostaining of tissues with GLUT4 primary antibody, visualized with a FITC-conjugated secondary antibody (green). Note, GLUT1 staining of the hummingbird pectoralis (a) is homogenous, and fiber sizes are all similar, reflecting the homogeneity of fiber type (type IIa; Fast oxidative-glycolytic). GLUT1 (and GLUT4) staining in the mouse gastrocnemius (b, d) is heteogenous and fiber diameters are varied, reflecting the diverse fiber type makeup of this muscle. Hummingbird pectoralis exhibited no staining using the GLUT4 antibody (intensity similar to use of secondary antibody alone; data not shown). Tissues in each panel were counterstained with DAPI in order to visualize nuclei (blue).</p

Glucose transporter mRNA expression in hummingbird tissues.

<p>Agarose gels (1.5%) of RT-PCR products for a) GLUT1 (340 bp), b) GLUT2 (305 bp), c) GLUT3 (543 bp), and d) GLUT4 (449 bp, expected product size from mouse), and e) GAPDH (585 bp). A 100 bp ladder was run in lane 1 of each gel. PCR reactions were performed on cDNA from hummingbird pectoralis (P), brain (B), heart (H), liver (L), ankle-extensor group muscles (G; e.g. gastrocnemius and soleus), wrist-extensor group muscle (E; e.g. extensor digitorum longus), kidney (K), and intestine (I), as well as cDNA from mouse cardiac tissue (MH; GLUT4 gel only) and samples of the reaction mixture were run in other lanes. Identical patterns of expression were observed using samples isolated from tissues of zebra finches (data not shown). Due to insufficient numbers of lanes per gel or because small tissue masses necessitated pooling of samples from 2 individuals, reaction products from some samples had to be run on separate gels. These are indicated by breaks in the image and by asterisks next to the lane headings (e.g. E*).</p

Glucose transporter protein expression in hummingbird tissues.

<p>Western blots using primary antibodies against a) GLUT1, b) GLUT4, and c) GAPDH. Samples were included from hummingbird pectoralis (P), brain (B), heart (H), liver (L), and for GLUT4 only, intestine (I) and kidney (K). Blots for GLU1 (a) and GAPDH (c) include samples from two different individual hummingbirds (e.g. P1 and P2). Samples from mouse (M) soleus (a, c) and cardiac (b) tissue are included as positive controls.</p

Glucose transporter staining in hummingbird heart and brain.

<p>Sections of hummingbird (a) heart and (b) brain tissue stained with GLUT1 primary antibody. Staining was visualized with a FITC-conjugated secondary antibody (green). Sections were counterstained with DAPI to visualize nuclei (blue).</p

Intensive lactation among women with recent gestational diabetes significantly alters the early postpartum circulating lipid profile: the SWIFT study

Abstract Background Women with a history of gestational diabetes mellitus (GDM) have a 7-fold higher risk of developing type 2 diabetes (T2D). It is estimated that 20-50% of women with GDM history will progress to T2D within 10 years after delivery. Intensive lactation could be negatively associated with this risk, but the mechanisms behind a protective effect remain unknown. Methods In this study, we utilized a prospective GDM cohort of 1010 women without T2D at 6-9 weeks postpartum (study baseline) and tested for T2D onset up to 8 years post-baseline (n=980). Targeted metabolic profiling was performed on fasting plasma samples collected at both baseline and follow-up (1-2 years post-baseline) during research exams in a subset of 350 women (216 intensive breastfeeding, IBF vs. 134 intensive formula feeding or mixed feeding, IFF/Mixed). The relationship between lactation intensity and circulating metabolites at both baseline and follow-up were evaluated to discover underlying metabolic responses of lactation and to explore the link between these metabolites and T2D risk. Results We observed that lactation intensity was strongly associated with decreased glycerolipids (TAGs/DAGs) and increased phospholipids/sphingolipids at baseline. This lipid profile suggested decreased lipogenesis caused by a shift away from the glycerolipid metabolism pathway towards the phospholipid/sphingolipid metabolism pathway as a component of the mechanism underlying the benefits of lactation. Longitudinal analysis demonstrated that this favorable lipid profile was transient and diminished at 1-2 years postpartum, coinciding with the cessation of lactation. Importantly, when stratifying these 350 women by future T2D status during the follow-up (171 future T2D vs. 179 no T2D), we discovered that lactation induced robust lipid changes only in women who did not develop incident T2D. Subsequently, we identified a cluster of metabolites that strongly associated with future T2D risk from which we developed a predictive metabolic signature with a discriminating power (AUC) of 0.78, superior to common clinical variables (i.e., fasting glucose, AUC 0.56 or 2-h glucose, AUC 0.62). Conclusions In this study, we show that intensive lactation significantly alters the circulating lipid profile at early postpartum and that women who do not respond metabolically to lactation are more likely to develop T2D. We also discovered a 10-analyte metabolic signature capable of predicting future onset of T2D in IBF women. Our findings provide novel insight into how lactation affects maternal metabolism and its link to future diabetes onset. Trial registration ClinicalTrials.gov NCT01967030