Data_Sheet_1_Causal effect of early life adiposity on gestational diabetes mellitus and mediating roles of lipidomic biomarkers.docx

ObjectiveThe causal relationship between early life adiposity and gestational diabetes mellitus (GDM) and the underlying mechanisms remains unclear. This study aimed to investigate the independent causal association between early life adiposity and GDM and identify potential metabolic mediators and their mediating effects on this relationship.MethodsUsing genome-wide association study (GWAS) summary statistics from the publicly available database of early life adiposity (5,530 cases and 8,318 controls) and GDM (11,279 cases and 179,600 controls), a two-step, two-sample Mendelian randomization (MR) was conducted to estimate the causal mediation effects of lipidomic biomarkers including low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), triglyceride, apolipoprotein A-Ι, and apolipoprotein B on the relationship between early life adiposity and GDM.ResultsGenetically predicted childhood adiposity was positively associated with risk of GDM (OR: 1.21, 95%CI: 1.09–1.34, p = 4.58 × 10−4). This causal relationship remained after accounting for adult adiposity traits in the multivariable MR analyses. Two-step MR identified three candidate mediators that partially mediated the effect of early life adiposity on GDM, including HDL-C (5.81, 95%CI: 3.05–8.57%), apolipoprotein A-Ι (4.16, 95%CI: 1.64–6.69%), and triglyceride (2.20, 95%CI: 0.48–3.92%).ConclusionThis MR study demonstrated that the causal effect of childhood obesity on future GDM risk was independent of adult adiposity. We identified three mediators, including HDL-C, apolipoprotein A-Ι, and triglyceride, in this association pathway. Our results provide insights into the pathogenesis of GDM and suggest additional prevention and treatment targets for GDM related to early life adiposity.</p

Suberitine A–D, Four New Cytotoxic Dimeric Aaptamine Alkaloids from the Marine Sponge <i>Aaptos suberitoides</i>

Suberitine A–D (<b>1</b>–<b>4</b>), four new bis-aaptamine alkaloids with two aaptamine skeleton units, 8,9,9-trimethoxy-9<i>H</i>-benzo[<i>de</i>][1,6]-naphthyridine and demethyl(oxy)-aaptamine, linked through a rare C-3–C-3′ or C-3–C-6′ <b>σ</b>-bond between the 1,6-naphthyridine rings, together with two known monomers <b>5</b> and <b>6</b>, were isolated from the marine sponge <i>Aaptos suberitoides</i>. Their structures were elucidated using NMR spectroscopy. Compounds <b>2</b> and <b>4</b> showed potent cytotoxicity against P388 cell lines, with IC₅₀ values of 1.8 and 3.5 μM, respectively

ALDH+ beta cells are not in M or S phase.

<p>(A–B) To determine precisely at which phase of a cell cycle, beta cells activate ALDH activity and thus become ALDH+, we co-stained ALDH (in green), insulin (in red) with PHH3 (in blue) and BrdU (in blue). BrdU was given to the mice two hours prior to sacrifice. (A) Representative images show essentially no PHH+ALDH+ beta cells, suggesting that beta cells lose ALDH activity when they enter M phase of a cell cycle. (B) Representative images show essentially no BrdU+ALDH+ beta cells, suggesting that beta cells lose ALDH activity when they enter S phase of a cell cycle. Scale bar is 50 µm.</p

ALDH Expression Characterizes G1-Phase Proliferating Beta Cells during Pregnancy

<div><p>High levels of aldehyde dehydrogenase (ALDH) activity have been detected in various progenitor and stem cells. Thus, Aldefluor fluorescence, which represents precisely the ALDH activity, has been widely used for the identification, evaluation, and isolation of stem and progenitor cells. Recently, ALDH activity was detected in embryonic and adult mouse pancreas, specifically in adult centroacinar and terminal duct cells supposed to harbor endocrine and exocrine progenitor cells in the adult pancreas. Nevertheless, ALDH activity and aldeflour fluorescence have not been examined in beta cells. Here, we report a dynamic increase in the number of aldeflour+ beta cells during pregnancy. Interestingly, nearly all these aldeflour+ beta cells are positive for Ki-67, suggesting that they are in an active cell cycle (G1, S and M phases). To determine precisely at which phase beta cells activate ALDH activity and thus become aldeflour+, we co-stained insulin with additional proliferation markers, phosphohistone3 (PHH3, a marker for M-phase proliferating cells) and Bromodeoxyuridine (BrdU, a marker for S-phase proliferating cells). Our data show little aldeflour+ beta cells that were positive for either PHH3, or BrdU, suggesting that beta cells activate ALDH and become Aldefluor+ when they enter G1-phase of active cell cycle, but may downregulate ALDH when they leave G1-phase and enter S phase. Our data thus reveal a potential change in ALDH activity of proliferating beta cells during pregnancy, which provides a novel method for isolation and analysis of proliferating beta cells. Moreover, our data also suggest that caution needs to be taken on interpretation of Aldefluor lineage-tracing data in pancreas.</p></div

Clinical characteristics of pregnant women with early and late onset preeclampsia and the matched control group (Control A and B).

<p>Values are shown as mean ± SD.</p><p>PE, preeclampsia; SBP, maximal systolic blood pressure; DBP, maximal diastolic blood pressure; Control A and Control B are gestational age-matched controls of normal pregnancies for ePE and lPE, respectively.</p>*<p><i>P</i><0.05, ePE versus Control A.</p>$<p><i>P</i><0.05, lPE versus Control B.</p><p>There were no significant differences in maternal age and delivery age between early and late-onset preeclampsia group and the matched control groups (<i>p</i>≥0.05). The blood pressure, systolic and diastolic blood pressures were significantly higher in preeclampsia groups than that in the matched control group (<i>p</i><0.05).</p

KiSS-1 and GPR54 mRNA expression in early-onset and late-onset pre-eclamptic placenta.

<p>Quantitative RT-PCR analysis of KiSS-1 and GPR54 mRNA expressions in preeclamptic (early-onset and late-onset) pregnancies compared with the respective matched control groups. The results demonstrated a significant up-regulation of KiSS-1 transcripts in the placenta of the early-onset preeclamptic pregnancies (A) and not significant difference between the late-onset preeclamptic (B) and control pregnancies. The transcript level of the KiSS-1 was significantly higher in early-onset placenta compared with late-onset preeclampsia (C). GPR54 revealed no significant difference in expression between early-onset/late-onset preeclamptic placenta and controls. Data represent means±SD after normalization to β-actin. *P<0.05, significantly decreased/increased compared with the respective control.</p

Beta cells increase ALDH activity in G1 phase during proliferation.

<p>Our finding was summarized and illustrated. Beta cells activate ALDH and become aldefluor+ when they enter G1-phase of an active cell cycle, but may downregulate ALDH and become Aldefluor- when they leave G1-phase and enter S phase.</p

Aldefluor+ cells are detected in the islets of pregnant mice.

<p>(A) Representative flow cytometry analysis of aldefluor fluorescence in the bone marrow cells (+/− treatment with a specific ALDH inhibitor DEAB) and isolated islets from pregnant mice 9 days after pregnancy (G9), compared with non-pregnant mice (G0). While no aldefluor+ cells were detected in G0 islets, aldefluor+ cells (circled) were readily detected in the islets from G9 islets. (B) Isolated G9 aldeflouor+ cells were immunostained positive for ALDH. (C) Quantification of aldefluor+ cells in the islet fraction from pregnant mice at 3, 6, 9, 12, 15 and 18 days after pregnancy (G3, G6, G9, G12, G15 and G18, respectively). These data show that in the mouse pancreas, islet cells upregulate ALDH activity during pregnancy. SSC: side-light scatter. *: p<0.05.</p

ALDH+ cells in the islets are predominantly beta cells in active cell cycle.

<p>(A–B) Representative triple staining for ALDH (in green), insulin (in red) and Ki-67 (in blue), together with nuclei staining with DAPI are shown in G9 (A) and G0 (B) pancreas. Each channel was shown independently. A merged image for ALDH, insulin and Ki-67 was also shown. The result suggests that aldefluor+ cells in the islets are predominantly proliferating beta cells. Arrows point to ALDH+ terminal cells. Scale bar is 50 µm.</p

IHC of metastin expression in the placentas of control A (A), ePE (B), control B (C), lPE (D), isotype control (E), positive control (F), and negative control (G).

<p>Panel H shows metastin immunity intensity score semiquantitation. Metastin staining is present in both syncytiotrophoblasts and cytotrophoblasts from Control A, ePE, Control B and lPE (A–D). Trophoblasts from ePE had higher expression of metastin as compared to control A (A, B). lPE had the same intensity expression of metastin as compared with control B (C,D). ePE had significantly higher expression of metastin as compared with control A and lPE (H). Red arrows indicate the syncytiotrophoblast; Black arrows indicate the cytotrophoblast.</p