Insulin Requirement for Gestational Diabetes Control Is Related to Higher Vitamin D Levels up to 1 Year Postpartum: A Prospective Cohort Study.

Vitamin D deficiency is highly prevalent in pregnant women and has been related to a higher risk of gestational diabetes mellitus (GDM). The aim of this study is to analyze vitamin D status evolution in a population of pregnant women with and without GDM. Two-hundred women were included from January 2019 to February 2022 as follows: Control group -CG-, Lifestyle group -LG- (GDM not requiring insulin), and Insulin group -IG- (GDM requiring insulin). Visits were carried out at baseline, antenatal, postpartum, and 1 year after birth. Vitamin D levels, weight, and insulin resistance were measured at every visit. Data about the season, vitamin D supplementation, Mediterranean diet adherence, and physical activity were included. In the three groups, 134 women were included in the CG, 43 in the LG, and 23 in the IG. Vitamin D levels were similar among the groups at baseline, but they were significantly higher in the LG and IG in comparison with the CG at the antenatal visit and significantly higher in the IG vs. CG and LG at the postpartum and 1 year after birth visits. Vitamin D levels were independently related to vitamin D supplementation and the season at baseline, to the season and belonging to the LG or IG at the antenatal visit, and were only independently associated with belonging to the IG at postpartum and 1 year after birth visits. In conclusion, in our population, women with GDM requiring insulin had higher levels of vitamin D in comparison with those not requiring insulin and healthy controls at postpartum and 1 year after pregnancy. Requiring insulin during pregnancy seems to be a factor that independently determines the levels of vitamin D until 1 year after birth. More studies are required to reproduce these data in other populations and to elucidate the mechanisms underlying these findings.

Impact of Immunosuppressive Drugs on the Metabolism of T Cells.

Energetic metabolism supports rapid cell growth and proliferation, differentiation, polarization, and effector functions of T cells. T lymphocytes have the remarkable plasticity that allows them to shape their metabolism to adapt to extracellular and intracellular cues, a process that involves molecular modules referred to as "metabolic checkpoints" that sense metabolic signals and transduce effector messages. These metabolic checkpoints may represent a novel therapeutic strategy for immune modulation. Chemical immunosuppressive drugs including mammalian target of rapamycin inhibitors (sirolimus and everolimus), calcineurin inhibitors (tacrolimus and cyclosporine), and purine and pyrimidine synthesis inhibitors (6-mercaptopurine, mycophenolic acid, and methotrexate) are widely prescribed for the treatment of autoimmune and inflammatory diseases and for controlling alloimmunity in interfering with the signals that activate and allow T cells to proliferate. Emerging evidence indicates that these drugs also target T-cell metabolism and metabolic checkpoints, which, as a consequence, could contribute to their immunosuppressive effects. These examples raise the issue of how the modulation of these metabolic checkpoints can regulate T-cell activation, differentiation, and function. In this review we highlight emerging concepts about the modulation of metabolic reprogramming in T-cell responses by immunosuppressive drugs and how potential therapeutic interventions influence T-cell fate and effector function.

A comprehensive characterization of the impact of mycophenolic acid on the metabolism of Jurkat T cells.

Metabolic reprogramming is critical for T cell fate and polarization and is regulated by metabolic checkpoints, including Myc, HIF-1α, AMPK and mTORC1. Our objective was to determine the impact of mycophenolic acid (MPA) in comparison with rapamycin (Rapa), an inhibitor of mTORC1, on the metabolism of Jurkat T cells. We identified a drug-specific transcriptome signature consisting of the key enzymes and transporters involved in glycolysis, glutaminolysis or nucleotide synthesis. MPA produced an early and transient drop in the intracellular ATP content related to the inhibition of de novo synthesis of purines, leading to the activation of the energy sensor AMPK. MPA decreases glycolytic flux, consistent with a reduction in glucose uptake, but also in the oxidation of glutamine. Additionally, both drugs reduce aerobic glycolysis. The expression of HIF-1α and Myc, promoting the activation of glycolysis and glutaminolysis, was inhibited by MPA and Rapa. In conclusion, we report that MPA profoundly impacts the cellular metabolism of Jurkat T cells by generating an energetic distress, decreasing the glycolytic and glutaminolytic fluxes and by targeting HIF-1α and Myc. These findings open interesting perspectives for novel combinatorial therapeutic strategies targeting metabolic checkpoints to block the proliferation of T cells.

6-mercaptopurine promotes energetic failure in proliferating T cells.

The anticancer drug 6-mercaptopurine (6-MP) inhibits de novo purine synthesis and acts as an antiproliferative agent by interfering with protein, DNA and RNA synthesis and promoting apoptosis. Metabolic reprogramming is crucial for tumor progression to foster cancer cells growth and proliferation, and is regulated by mechanistic target of rapamycin (mTOR) and AMP-activated protein kinase (AMPK) as well as the oncogenes Myc and hypoxia inducible factor 1α (HIF-1α). We hypothesized that 6-MP impacts metabolic remodeling through its action on nucleotide synthesis. The aim of our study is to provide a comprehensive characterization of the metabolic changes induced by 6-MP in leukemic T cells. Our results indicate that exposition to 6-MP rapidly reduces intracellular ATP concentration, leading to the activation of AMPK. In turn, mTOR, an AMPK target, was inhibited, and the expression of HIF-1α and Myc was reduced upon 6-MP incubation. As a consequence of these inhibitions, glucose and glutamine fluxes were strongly decreased. Notably, no difference was observed on glucose uptake upon exposition to 6-MP. In conclusion, our findings provide new insights into how 6-MP profoundly impacts cellular energetic metabolism by reducing ATP production and decreasing glycolytic and glutaminolytic fluxes, and how 6-MP modifies human leukemic T cells metabolism with potential antiproliferative effects.

The effect of immunosuppressive molecules on T-cell metabolic reprogramming.

T lymphocytes undergo metabolic reprogramming to adapt to extracellular and intracellular cues. Specifically, T-cell metabolism results into ATP production, anabolism and catabolism pathways that not only support rapid cell growth and proliferation, but also differentiation and effector functions, recently referred as "immunometabolism". Quiescent naïve T cells rely on oxidative phosphorylation whereas aerobic glycolysis (Warburg effect) occurs in activated T cells (effector CD4(+) and CD8(+)). The molecular mechanisms that sense metabolic status and influence T-cell function require metabolic checkpoints including sensors of metabolic signals and transducers (Myc, HIF-1α, AMPK and mTOR). These metabolic checkpoints represent a novel therapeutic strategy for immune modulation. Interestingly, many immunosuppressive drugs including mTOR inhibitors (rapamycin), calcineurin inhibitors (tacrolimus, cyclosporine A) and inhibitors of de novo purine synthesis (6-mercaptopurine, mycophenolic acid and methotrexate) provide examples into how modulating these metabolic checkpoints can regulate T-cell activation, differentiation and function. In this Review we highlight emerging concepts about metabolic reprogramming in T-cell responses and we discuss the potential therapeutic interventions to influence T-cell fate and effector function.

Molecular insight into thiopurine resistance: transcriptomic signature in lymphoblastoid cell lines.

BACKGROUND: There has been considerable progress in the management of acute lymphoblastic leukemia (ALL) but further improvement is needed to increase long-term survival. The thiopurine agent 6-mercaptopurine (6-MP) used for ALL maintenance therapy has a key influence on clinical outcomes and relapse prevention. Genetic inheritance in thiopurine metabolism plays a major role in interindividual clinical response variability to thiopurines; however, most cases of thiopurine resistance remain unexplained. METHODS: We used lymphoblastoid cell lines (LCLs) from healthy donors, selected for their extreme thiopurine susceptibility. Thiopurine metabolism was characterized by the determination of TPMT and HPRT activity. We performed genome-wide expression profiling in resistant and sensitive cell lines with the goal of elucidating the mechanisms of thiopurine resistance. RESULTS: We determined a higher TPMT activity (+44%; P = 0.024) in resistant compared to sensitive cell lines, although there was no difference in HPRT activity. We identified a 32-gene transcriptomic signature that predicts thiopurine resistance. This signature includes the GTPBP4 gene coding for a GTP-binding protein that interacts with p53. A comprehensive pathway analysis of the genes differentially expressed between resistant and sensitive cell lines indicated a role for cell cycle and DNA mismatch repair system in thiopurine resistance. It also revealed overexpression of the ATM/p53/p21 pathway, which is activated in response to DNA damage and induces cell cycle arrest in thiopurine resistant LCLs. Furthermore, overexpression of the p53 target gene TNFRSF10D or the negative cell cycle regulator CCNG2 induces cell cycle arrest and may also contribute to thiopurine resistance. ARHGDIA under-expression in resistant cell lines may constitute a novel molecular mechanism contributing to thiopurine resistance based on Rac1 inhibition induced apoptosis and in relation with thiopurine pharmacodynamics. CONCLUSION: Our study provides new insights into the molecular mechanisms underlying thiopurine resistance and suggests a potential research focus for developing tailored medicine.

Effect of CPAP on oxidative stress and circulating progenitor cell levels in sleep patients with apnea-hypopnea syndrome.

BACKGROUND: The sleep apnea-hypopnea syndrome is associated with elevated oxidative stress, which is associated with reduced levels and functional impairment of progenitor cells. OBJECTIVE: To evaluate whether one month of CPAP treatment affects circulating-progenitor-cell levels and oxidative stress in patients with sleep apnea-hypopnea syndrome. METHODS: We enrolled 13 patients with sleep apnea-hypopnea syndrome who required nasal CPAP. We evaluated white-blood-cell oxidative stress and CD45-, CD34+, KDR+, and CD133+ cell levels via flow-cytometry, before and one month after CPAP treatment. RESULTS: Superoxide anion and hydrogen peroxide were reduced, and markers of protection against oxidative stress were increased after CPAP. Progenitor-cell levels increased significantly after CPAP. There was a significant negative correlation between CD45-, CD34+, KDR+, and CD133+ cell levels and the severity of sleep apnea-hypopnea syndrome and superoxide anion. CONCLUSIONS: CD45-, CD34+, KDR+, and CD133+ cell levels rose significantly and reached values close to those in the control group after one month of CPAP. This change was accompanied by a significant decrease in oxidative stress, and no change in anthropometric or metabolic variables, including insulin resistance, weight, blood pressure, or lipid levels; consequently, the increase in progenitor-cell levels might be attributable to reduced oxidative stress.

Continuous positive airway pressure therapy reduces oxidative stress markers and blood pressure in sleep apnea-hypopnea syndrome patients.

Sleep apnea-hypopnea syndrome (SAHS) is characterized by recurrent episodes of hypoxia/reoxygenation, which seems to promote oxidative stress. SAHS patients experience increases in hypertension, obesity and insulin resistance (IR). The purpose was to evaluate in SAHS patients the effects of 1 month of treatment with continuous positive airway pressure (CPAP) on oxidative stress and the association between oxidative stress and insulin resistance and blood pressure (BP). Twenty-six SAHS patients requiring CPAP were enrolled. Measurements were recorded before and 1 month after treatment. Cellular oxidative stress parameters were notably decreased after CPAP. Intracellular glutathione and mitochondrial membrane potential increased significantly. Also, total antioxidant capacity and most of the plasma antioxidant activities increased significantly. Significant decreases were seen in BP. Negative correlations were observed between SAHS severity and markers of protection against oxidative stress. BP correlated with oxidative stress markers. In conclusion, we observed an obvious improvement in oxidative stress and found that it was accompanied by an evident decrease in BP with no modification in IR. Consequently, we believe that the decrease in oxidative stress after 1 month of CPAP treatment in these patients is not contributing much to IR genesis, though it could be related to the hypertension etiology.

Evaluación de un método POCT (pruebas a la cabecera del paciente) para la determinación de hemoglobina glucosilada / Evaluation of a POCT (point of care testing) assay for haemoglobin A1c

Introducción. La hemoglobina glucosilada (HbA1C) presenta un papel fundamental en el control y prevención de complicaciones en el paciente diabético, puesto que permite la monitorización de sus valores de glucemia en los 3 meses previos al análisis. Introducción. Debido a la importancia de su determinación han aparecido diferentes métodos de cuantificación. El objeto de este trabajo es evaluar el método POCT (point of care testing ‘pruebas a la cabecera del paciente’) mediante el aparato In2it de Bio-Rad. Material y métodos. Se realizó una comparación entre 2 métodos de cuantificación: el HPLC (high–performance liquid chromatography ‘cromatografía líquida de alta resolución o de alta presión’) (ADAMS-A1c HA-8160, Menarini Diagnostics, Florencia, Italia) y el POCT (In2it, Bio-Rad, California, EE.UU.). Para esto, se compararon los resultados de 60 muestras, además de determinar la imprecisión intraserial y la imprecisión interdiaria de ambos aparatos. Resultados. La comparación entre ambos métodos mostró una buena correlación entre el HPLC y el POCT: Y=0,4000+1,000 X. Los intervalos de confianza del 95% para la coordenada en el origen y la pendiente fueron de −0,1500 a 0,6500 y de 0,9565 a 1,0769, respectivamente. Resultados. La imprecisión intraserial se situó entre el 2,5 y el 3% (del 2,20 al 3,04%) y la imprecisión interdiaria se situó entre el 3,53 y el 2,95% para valores altos y bajos, respectivamente. Discusión. El método POCT ha demostrado cumplir todos los requisitos técnicos necesarios para su validación como método para la determinación de la HbA1C, ya que presenta igualdad de prestaciones analíticas con el método HPLC. Además, es apropiado para el seguimiento de pacientes diabéticos en atención primaria, dada su facilidad y sencillez de manejo (AU)

Introduction. The HbA1c is related to the control and prevention of complications in diabetics. This parameter enables glucose control to be monitored in the three months previous to the determination. Introduction. Due to the significance of its determination, different measurement methods have been developed. The aim of this work is to evaluate the POCT (point of care testing) method used by the In2it BioRad. Material and methods. We report on a comparison between two methods: a HPLC (high–performance liquid chromatography) (ADAMS-A1c HA-8160, Menarini) and the POCT test (In2it, BioRad). To determine the within and between day imprecision, 60 samples were analysed and compared. Results. Both methods showed good correlations (Y=0.4000+1.000 X). The 95% confidence intervals for the coordinate at the origin and the slope were (−0.1500 to 0.6500) and (0.9565 to 1.0769), respectively. Within day imprecision was 2.5–3% (2.20–3.04%) and between day imprecision was 3.53% and 2.95% for high and low values, respectively. Discussion. The POCT test (BioRad System) has demonstrated that it fulfils all the technical requirements needed for its validation as an HbA1c analysis method as it has a similar performance to the HPLC method The POCT method has shown to be suitable for the follow-up of diabetic patients in primary care due its ease and simplicity of use (AU)

Evaluación de un método inmunonefelométrico competitivo para la cuantificación de la homocisteína / Evaluation of a competitive immunonephelometric method for the measurement of homocysteine

Introducción y objetivos. La homocisteína se relaciona con enfermedad vascular, alteraciones del estado nutricional y detección de homocistinuria en neonatos, entre otras enfermedades. Debido a la importancia de su determinación, han aparecido diferentes métodos de cuantificación; el objeto de este trabajo es evaluar el método inmunonefelométrico del aparato BN II (Dade Behring). Material y método. Se realizó una comparación entre 2 métodos de cuantificación: el inmunoanálisis competitivo (IMMULITE 2000, DPC) y el análisis nefelométrico (BN II, Dade Behring), para lo cual se compararon los resultados de 74 muestras, además de determinar la imprecisión intraserial, imprecisión interdiaria, el límite de detección, el límite de cuantificación y el valor crítico del método inmunonefelométrico del BN II de Dade Behring. Resultados. La comparación entre ambos métodos mostró una buena correlación entre el inmunoanálisis competitivo, IMMULITE 2000 de DPC y el análisis nefelométrico, BN II de Dade Behring (Y = 1,4825 + 0,8342X). El valor crítico obtenido fue de 5,46 mmol/l y el límite de detección, de 5,77 mmol/l. La imprecisión intraserial fue inferior al 5% (3,65-4,66%). Conclusiones. El análisis nefelométrico (BN II, Dade Behring) ha demostrado cumplir todos los requisitos técnicos necesarios para su validación como método para la determinación de la homocisteína(AU)

Introduction. The homocysteine is associated with vascular diseases, alterations in the nutritional states, homocystinuria detection in neonates, as well as other diseases. Due to the importance of its determination, different measurement methods have been developed. The aim of this work is to evaluate the imunonephelometric method used in the Dade Behring BN II Nephelometer system. Material and method. We present a comparison between 2 methods: a competitive inmunoassay (IMMULITE 2000, DPC) and the nephelometric test (BN II, Dade Behring). For the determination of within batch and between-day imprecision, 74 samples were analysed and compared. Results. The detection and quantification limits, and the critical value of the inmunonephelometric method, were also determined. Both methods showed good correlations (Y = 1.4825 + 0.8342X). We also obtained a critical value of 5.46 mmol/L and the detection limit was 5.77 mmol/L. Within batch imprecision was below 5 % (3.65-4.66%). Conclusions. The nephelometric test (Dade Behring BN II System) has demonstrated to fulfill all the technical requirements needed for its validation as a method for the determination of homocysteine(AU)