Background/Objectives: Lifestyle Modification Programs (LMPs) based on exercise and nutrition aim to prevent/manage chronic diseases and foster well-being. However, moving LMPs from research to medical practice can be challenging, as programs must be both effective and feasible. The primary goal of this study was to assess cardiorespiratory fitness (CRF) changes according to an LMP, measured through VO2max, as a key indicator of health outcomes and intervention efficacy. Methods: In this single-arm intervention study, 100 subjects were enrolled; per-protocol analysis of main parameters was performed on 85 participants (15 were excluded due to medical/technical reasons). A feasible intervention program (of low resource intensity with only two physician/patient encounters) provided personalized exercise prescription, optimized nutritional habits based on the Mediterranean diet and Healthy Eating Plate principles, and supported behaviour change. We assessed CRF through VO2max, a key indicator of health outcomes and intervention efficacy. We also analyzed, using regression analysis, the relationship between VO2max (the gold-standard measure of CRF) and METSpeak, a simpler, feasible parameter of CRF derived from Exercise Stress Testing. Body composition (BC) and AHA diet score were also measured at baseline and post-6-month intervention. Statistical analyses included paired comparisons and multivariable regression to explore factors influencing CRF changes. Results: Analysis on the primary outcome, VO2max, was performed according to the intention-to-treat principle and per-protocol. This feasible protocol resulted in a significant increase in VO2max, improvements in fat-free mass, and a reduction in fat mass. Overall, 42.4% of participants achieved an improvement of ≥1 MET, a change previously associated with reduced mortality risk. Older participants tend to experience smaller improvements in VO2max. Conclusions: Although observing an improvement in CRF and BC following an LMP is not surprising, the strength of the study is to show the feasibility of implementing an effective, feasible LMP into clinical routine, supporting the integration of such programs into clinical practice.

Background/Objectives: Maternal nutrition and pregestational BMI are critical determinants of pregnancy outcomes. This prospective multicenter observational study investigated the interplay between prepregnancy BMI, dietary patterns, and oxidative/inflammatory status in 153 Italian healthy pregnant women with normal weight (NW), overweight (OW), or obesity (OB). Methods: Detailed clinical, biochemical, placental, and neonatal data were measured at third trimester and delivery. Dietary intake was assessed via a validated questionnaire, and dietary patterns were derived using principal component analysis. Results: OW and OB women had significantly higher levels of inflammatory (CRP, hepcidin) and oxidative stress biomarkers (DNA/RNA damage, catalase activity) than NW. Multivariate models confirmed independent associations between BMI and these biomarkers (CRP: β = 0.297, p = 0.000; hepcidin: β = 1.419, p = 0.006; DNA/RNA damage: β = 409.9, p = 0.000; catalase activity: β = 1.536, p = 0.000). Superoxide dismutase activity and total antioxidant capacity were not associated with BMI. Nutritional intake across BMI groups was largely suboptimal relative to national recommendations, with insufficient levels of polyunsaturated fats and key micronutrients. Four dietary patterns were identified, with adherence varying by BMI. A “prudent-style” pattern (high plant, low animal) was positively associated with gestational age (β = 0.243, p = 0.033) and inversely with neonatal head circumference (β = −0.414, p = 0.050). A “Western-like” pattern (high sugars, snacks, animal fats) was linked to reduced maternal ferritin (β = −2.093, p = 0.036) and increased neonatal head circumference (β = 0.403, p = 0.036). However, not all deviations from the “prudent-style” pattern were metabolically equivalent: while Pattern 3 (high-protein, carbohydrate) may offer partial protective effects, Pattern 4 (moderate protein/plant/sugar) displayed elements of nutritional imbalance with signs of placental inefficiency (β = −0.384, p = 0.023). Conclusions: These findings underscore the dual impact of maternal BMI and diet quality on oxidative-inflammatory balance and perinatal outcomes, supporting the need for early, individualized nutritional strategies in pregnancy. This is further emphasized by the variability in dietary adherence across BMI categories.

Long-chain polyunsaturated fatty acids (LC-PUFAs) are essential nutrients for feto-placental development. We aimed to evaluate the associations between maternal pregestational BMI, mid-pregnancy LC-PUFA status, and delivery outcomes in non-obese pregnancies. This was a secondary analysis of two Italian cohorts including healthy non-obese women with singleton spontaneous pregnancies previously studied for maternal nutritional habits, multivitamin supplementation, blood biomarkers and infant biometry/measures. In the present analysis, included women were stratified according to pregestational BMI (normal weight (NW) versus overweight (OW) groups). Fasting venous blood samples were collected between 24 and 34 gestational weeks for fatty acid (FA) analysis. Pregnancy outcomes were recorded at delivery. Multi-adjusted generalized linear models were applied to first assess the associations between BMI-based groups and mid-pregnancy LC-PUFA concentrations, and second to evaluate the associations between the LC-PUFA profile and pregnancy outcomes. 283 pregnancies were included. The OW group showed lower eicosapentaenoic acid (EPA) levels (β= -0.09; 95

BACKGROUND:Placental development, maternal adaptations to pregnancy, and birth outcomes appear to differ based on fetal sex. The aim of this prospective cohort study was to investigate first trimester markers of feto-placental development and birth outcomes in a large population of healthy women with autologous pregnancies according to fetal sex. METHODS:Healthy women with singleton pregnancies were enrolled at 11-13+6 weeks. At enrollment, maternal blood sample was collected for placental marker (Pregnancy Associated Plasma Protein A, free β-Human Chorionic Gonadotropin (HCG)), and ultrasound assessed mean pulsatility index of uterine arteries (UtA PI), placental volume and crown-rump length (CRL). Multi-adjusted generalized linear models were used to evaluate the associations between fetal sex, first trimester feto-placental markers, and pregnancy outcomes. RESULTS:Among 1052 pregnancies (523 male, 529 female), female fetuses showed lower first trimester mean UtA PI (β = -0.10 (95 % CI -0.16; 0.03), p < 0.01) and CRL (β = -0.49 (95 % IC -0.96; 0-04), p < 0.05), and higher free-β-hCG (β = 19.8 (95 % IC 13.8-25.8), p < 0.001) in multi-adjusted models. Female sex was associated with a 69 % lower risk of preterm delivery. Male fetuses showed higher birth weight (p < 0.001). After excluding women undergoing a planned cesarean section, male fetuses showed lower arterial cord pH values regardless of the delivery mode (β = -0.21; 95 %IC: -0.40; -0-01, p < 0.05). CONCLUSIONS:The present study highlights crucial differences in prenatal development and placentation between male and female fetuses, leading to significant differences in pregnancy duration and response to labor stress.

Periodontal disease (PD) during pregnancy may trigger systemic inflammation, increasing the risk of developing cardiometabolic disease (CMD). As a consequence, PD may result in the activation of cellular and molecular pathways, affecting the disease course and pregnancy outcome. Although microRNAs (miRNAs) are considered ideal biomarkers for many diseases, few studies have investigated salivary miRNAs and their role in pregnancy or neonatal outcomes. In this study, we sought to investigate the associations between salivary miRNAs of pregnant women with oral diseases and their effects on neonatal outcomes. Eleven (n = 11) salivary miRNAs from a cohort of pregnant women with oral diseases (n = 32; oral health, H; gingivitis, G; and periodontitis, P) were detected using a previous profiling analysis with an FDR < 0.20 and a fold change (FC) < 0.5 or FC > 2 for the most highly expressed miRNAs. Spearman correlations were performed for 11 salivary microRNAs associated with oral-derived inflammation, which could affect neonatal outcomes during pregnancies at risk for cardiometabolic disease (CMD), defined by the presence of a high pregestational BMI. In addition, ROC curves demonstrated the diagnostic accuracy of the markers used. Upregulation of miR-423-5p expression and a decrease in miR-27b-3p expression were detected in the P-group (p < 0.05), and ROC analysis revealed the diagnostic accuracy of miR-423-5p for discriminating oral diseases, such as gingivitis versus periodontitis (P vs. G, AUC = 0.78, p < 0.05), and for discriminating it from the healthy oral cavity (P vs. H, AUC = 0.9, p < 0.01). In addition, miR-27b-3p and miR-622 were also able to discriminate the healthy group from the P-group (AUC = 0.8, p < 0.05; AUC = 0.8, p < 0.05). miR-483-5p was able to discriminate between the G-group (AUC = 0.9, p < 0.01) and the P-group (AUC = 0.8, p < 0.05). These data support the role of salivary miRNAs as early biomarkers for neonatal outcomes in pregnant women with periodontal disease at high risk for CMD and suggest that there is cross-talk between salivary miRNAs and subclinical systemic inflammation.

Inflamm-aging refers to the chronic low-grade inflammation that occurs with aging and cellular senescence, and it is linked to various diseases. Understanding the markers involved in inflammation and aging, as well as their interaction with environmental factors and bodily control mechanisms, can provide crucial tools for assessing the resilience (i.e. the ability to adapt and improve) of the human body, particularly in the presence of chronic degenerative conditions or vulnerable life stages, that place the individual and the community to which he belongs in a state of potential fragility. HEBE focuses on physical exercise, along with nutritional and lifestyle recommendations, to reduce systemic inflammation and promote healthy aging. HEBE encompasses multiple research lines (LR). In the ongoing LR1 (“proof of concept”), healthy lifestyle recommendations were provided to University of Milan employees, and changes in quality of life and well-being were assessed using a specialized questionnaire. The first 100 eligible subjects, who expressed their willingness to participate, underwent a personalized physical exercise protocol based on clinical and objective assessments. Biomedical samples were collected at baseline (T0) and follow-up (T1) to establish a shared biobank and identify non-invasive biomarkers that monitor the impact of physical exercise on individual characteristics such as cardiovascular and metabolic health. Subsequently (LR2-LR10), the proof of concept findings will be expanded to include various conditions of vulnerability such as obesity, cancer, endocrine disorders, cardiovascular diseases, infertility, functional syndromes, respiratory disorders, neurodegenerative diseases, and autoimmune conditions. The research lines will leverage the expertise of the 94 participating investigators to form a collaborative network that maximizes the potential for investigation and knowledge exchange. This approach fosters a culture of health promotion and disease prevention. ClinicalTrials.gov Identifier: NCT05815732.

The HEBE project, "Healthy Aging versus Inflamm-Aging: The Role of Physical Exercise in Modulating the Biomarkers of Age-Associated and Environmentally Determined Chronic Diseases," investigates the phenomenon of inflamm-aging, a chronic low-grade inflammatory process that accelerates cellular decline and contributes to age-related diseases. This longitudinal study examined 100 participants at two time points—before and after a personalized physical exercise intervention—to analyze dynamic changes in cytokines and other biomarkers associated with aging. Using advanced multivariate statistical approaches such as Principal Component Analysis (PCA) and its extensions, along with Repeated Measures Analysis of Variance-Simultaneous Component Analysis (RM-ASCA), the study explored the interplay among inflammatory and non-inflammatory markers, including cytokines, telomere length, and the oxidative stress indicator 8-OHdG. These methods enabled researchers to uncover the intricate interactions driving inflamm-aging and identify modifiable factors that could mitigate its effects. The findings highlight that physical exercise, combined with lifestyle changes, significantly influences key biomarkers, fostering resilience against age-related inflammation and decreasing the risk of non-communicable diseases. The integrative multivariate approach provided an in-depth understanding of temporal trends and complex biomarker interrelationships, revealing latent patterns and clustering effects that traditional univariate analyses might overlook. This research emphasizes the crucial role of controlling inflammation in aging and positions physical exercise as a pivotal intervention to counteract its negative impacts. The results offer fresh perspectives on the systemic responses induced by exercise and propose actionable strategies for preventing age-related decline through targeted lifestyle modifications.

Maternal obesity has been associated with short- and long-term risks of pregnancy-perinatal adverse events, possibly due to alterations of placental mitochondrial bioenergetics. However, several detrimental mechanisms occurring in the placentas of women with obesity still need to be clarified. Here, we analyzed placental mitochondrial features and oxidative environment of 46 pregnancies in relation to pre-pregnancy BMI. Seventeen Caucasian normal-weight (NW) and twenty-nine women who were obese (OB) were enrolled. The protein expression of mitochondrial CypD and electron transfer chain complexes (C) I–V were measured, as well as ATP production and oxygen consumption rates (OCRs). The protein levels of the pro/anti-oxidant enzymes TXNIP, SOD2, and PON2 were also analyzed. Despite no differences in CypD expression, OCRs were significantly lower in OB vs. NW women. Accordingly, ATP synthase (CV) levels and ATP content were decreased in OB women, positively correlating with placental efficiency, suggesting a link between ATP deficiency and placental dysfunction. SOD2 expression negatively correlated with maternal BMI, indicating a possible impairment of antioxidant defenses with increasing BMI. These changes were worsened in 10 OB women presenting with gestational diabetes mellitus. Overall, these results suggest alterations of placental bioenergetics in pregnancies of women with obesity, possibly leading to placental dysfunction and altered fetal development and programming.

Introduction: Monochorionic twins (MC) develop under unique intrauterine conditions and show a high risk of compromise during fetal life. Here we describe umbilical vein blood flow (UVBF) and fetal oxygen and glucose utilization in uncomplicated MC twins and investigate possible differences within twin-pairs according to birth -order.Methods: Prospective single-center study on 48 uncomplicated MC twins enrolled at the time of elective cesarean delivery. Ultrasound measurements of UVBF for Twin 1 and Twin 2 labelled according to birth-order were performed before spinal anesthesia. Umbilical arterial and venous blood samples were collected for each twin after fetal delivery, and fetal oxygen and glucose deliveries and uptakes were computed.Results: All twins were delivered within 2 min from one-another under steady-state conditions at 36.4 weeks of median gestational age (IQR 36.0-37.0). Birthweight and umbilical cord gas analyses were within physiological ranges for all twins. Second-born twins showed significantly lower UVBF, measured before delivery, and lower median birthweight compared to first-borns. Moreover, median values of estimated fetal oxygen and glucose consumption were lower in second compared to first MC twins.Discussion: Uncomplicated MC twins show different birthweight, oxygenation and metabolic rates based on their position in utero, hinting at pre-existing conditions possibly deriving by uneven vascular and metabolic distri-bution of the two placental territories. The innovative findings of this study emphasize the biological uniqueness of these pregnancies and prompt further physiological studies on MC twins and placenta metabolism.

Maternal obesity (MO) is expanding worldwide, contributing to the onset of Gestational Diabetes Mellitus (GDM). MO and GDM are associated with adverse maternal and foetal outcomes, with short- and long-term complications. Growing evidence suggests that MO and GDM are characterized by epigenetic alterations contributing to the pathogenesis of metabolic diseases. In this pilot study, plasma microRNAs (miRNAs) of obese pregnant women with/without GDM were profiled at delivery. Nineteen women with spontaneous singleton pregnancies delivering by elective Caesarean section were enrolled: seven normal-weight (NW), six obese without comorbidities (OB/GDM(−)), and six obese with GDM (OB/GDM(+)). miRNA profiling with miRCURY LNA PCR Panel allowed the analysis of the 179 most expressed circulating miRNAs in humans. Data acquisition and statistics (GeneGlobe and SPSS software) and Pathway Enrichment Analysis (PEA) were performed. Data analysis highlighted patterns of significantly differentially expressed miRNAs between groups: OB/GDM(−) vs. NW: n = 4 miRNAs, OB/GDM(+) vs. NW: n = 1, and OB/GDM(+) vs. OB/GDM(−): n = 14. For each comparison, PEA revealed pathways associated with oxidative stress and inflammation, as well as with nutrients and hormones metabolism. Indeed, miRNAs analysis may help to shed light on the complex epigenetic network regulating metabolic pathways in both the mother and the foeto-placental unit. Future investigations are needed to deepen the pregnancy epigenetic landscape in MO and GDM.

Abstract Background High-resolution respirometry (HRR) of human biopsies can provide useful metabolic, diagnostic, and mechanistic insights for clinical research and comparative medical studies. Fresh tissues analysis offers the potential best condition, the drawback being the need to use them shortly after dissection for mitochondrial respiratory experiments. The development of effective long-term storage protocols for biopsies that allow the assessment of key Electron Transport System (ETS) parameters at later stages is thus a major need. Methods We optimised a cryopreservation protocol that preserves mitochondrial membranes intactness, otherwise affected by direct tissue freezing. The protocol is based on a gradual freezing step from on-ice to liquid nitrogen and − 80 °C storage using a specific DMSO-based buffer. Results Placenta is a suitable tissue to design and test the effectiveness of long-term storage protocols being metabolically active foetal tissue with mitochondrial dysfunctions contributing to placental disease and gestational disorders. Here we designed and tested the effectiveness of the cryopreservation protocol using human placenta biopsies; we measured the ETS activity by HRR of placenta specimens comparing fresh, cryopreserved, and snap frozen conditions. Conclusions By this protocol, Oxygen Consumption Rate (OCR) measurements of fresh and cryopreserved placental specimens are comparable whereas snap frozen procedure impairs mitochondrial activity. Graphical Abstract

Background: Nutritional quality during pregnancy is crucial for mother and child health and their short/long-term outcomes. The aim of this study is to evaluate the adherence to nutritional recommendations in Italy during the three pregnancy trimesters in Normal Weight (NW) and Over Weight (OW) women. Methods: Data from a multicenter randomized controlled trial included 176 women (NW = 133; OW = 43) with healthy singleton pregnancies enrolled within 13 + 6 weeks of gestation. Dietary intake was assessed every trimester by a Food Frequency Questionnaire. Results: OW and NW had similar gestational weight gain. However, as Institute of Medicine (IOM) recommend lower gestational weight gain (GWG) for OW, they exceeded the suggested range. In both groups, caloric intake during the three trimesters never met recommendations. Protein intake in first and second trimester was higher than recommendations, as was sugars percentage. Dietary fiber intake was lower in OW. Polyunsaturated fatty acids, calcium, iron and folic acid requirements were never satisfied, while sodium intake exceeded recommendations. Conclusions: NW and OW women in Italy do not adhere to nutritional recommendations during pregnancy, with lower caloric intake, protein and sugars excess and inadequacies in micronutrients intake. Pregnant women in Italy should be provided with an adequate counseling and educational intervention as well as supplementation when indicated.

Alterations in intrauterine fetal growth increase the risk of adverse perinatal and neonatal outcomes. In this retrospective study, we analyzed data of 906 pregnancies collected in our maternal fetal medicine center, with different patterns of growth: 655 AGA (Appropriate for Gestational Age), 62 SGA (Small for Gestational Age: fetuses born with a weight less than 10° centile, not diagnosed before delivery), 189 FGR (Fetal Growth Restriction, classified in early and late according to gestational week at diagnosis). For each group, we compared maternal characteristics, gestational age at delivery, and perinatal and neonatal outcomes. Risk factors for fetal growth alterations were advanced age, being primiparous, and a lower pregestational BMI. FGR fetuses were born at earlier gestational ages (32 [IQR 29–38] early-FGR and 38 [IQR 36–39] late-FGR), with blood gas values comparable to the AGA group but worse neonatal outcomes related to prematurity. Unexpected SGA fetuses born by vaginal delivery, managed as AGA, were more hyperlactacidemic (4.4 [IQR 2.7–5.5]) and hypoxemic (−5.0 [IQR −7.1–2.8]) at birth than both AGA and FGR. However, neonatal outcomes (accesses and days of hospitalization in NICU) were better than FGR, likely due to gestational age and birthweight similar to AGA.