We aimed to determine the differences in the growth trajectories of the youngest gestational survivors (<25 weeks’ gestation) up to 6 years of age compared to those of older gestational ages. Preterm infants were divided into two groups: 22–24 weeks’ gestation  16, female (F) 28) and 25–29 weeks’ gestation . Z-scores of body weight (BW), body length (BL), and body mass index (BMI) were derived from Japanese standards at 1, 1.5, 3, and 6 years of corrected age. Comparisons between the two groups by sex were made using the Wilcoxon test and linear regression analysis to examine the longitudinal and time-point associations of anthropometric z-scores, the presence of small for gestational age (SGA), and the two gestational groups. BW, BL, BMI, and z-scores were significantly lower in the 22–24 weeks group at almost all assessment points. However, there were no significant differences in BW, BL, BMI, and z-scores between the two female groups after 3 years. BMI z-scores were significantly associated with the youngest gestational age and the presence of SGA at all ages in males, but not in females. The youngest gestational age had a greater influence in males on the z-score of anthropometric parameters up to 6 years of age. Recent progression in neonatal intensive care has led to dramatic improvements in the survival of very low birth weight  infants and very preterm infants . However3) or body mass index  was calculated. Sex- and GA-independent z-scores and percentiles for anthropometric parameters at birth were calculated according to the Japanese standard charts, which were estimated using 2003–2005 data from the Japan Society of Obstetrics and Gynecology registry database [A retrospective multicenter cohort study was conducted that including preterm infants admitted to neonatal intensive care units (NICUs) in the three affiliated hospitals of Juntendo University  from January 2007 to June 2012. The inclusion criteria for preterm infants were birth weight <1500 g, gestational age (GA) <30 weeks and Japanese parentage. Participants were divided into two subgroups: 22–24 weeks of gestation (22–24 weeks) and 25–29 weeks of gestation (25–29 weeks). GA was determined by last menstrual period and first-trimester ultrasonogram. Body weight (BW) and body length (BL) at birth and 1, 1.5, 3, and 6 years of corrected age  were collected from medical records, and the Ponderal index , Juntendo University Shizuoka Hospital (2017-059) and Juntendo University Urayasu Hospital (rin-541). All methods in this retrospective study were performed in accordance with the relevant guidelines . Because this is a retrospective study, it was very difficult to gain appropriate informed consents from each subject. Therefore, we gained consent using opt-out, which is a way for investigators to give subjects and their guardians an opportunity to refuse to participate in this study by announcing the details of the study in each participating hospital.p < 0.05. The anthropometric parameters were individually plotted between 1 and 6 years of CA to visually inspect the longitudinal pattern. We performed linear regression analysis to determine the association between anthropometric z-scores and the presence of SGA and GA. First, the samples were divided into two GA groups. One group consisted of infants born between 22 and 24 GA, and the other group was born between 25 and 29 GA. We set the group born between 22 and 24 GA and infants with SGA as the reference category and performed linear regression analysis with these variables as independent variables. Dependent variables were z-scores of BW, BL, and BMI at 1, 1.5, 3, and 6 years of CA. Regression analysis was performed separately by sex. Data were analyzed using SAS version 9.4 .Data are presented as the mean and standard deviation. Differences between the groups were identified using the Wilcoxon test. These statistical analyses were conducted using JMP statistical software . Statistical significance was set at During the study period, a total of 529 VLBW infants with GA < 30 weeks were born in the NICUs of three hospitals, and 502 infants met the inclusion criteria at birth. Of these infants, 412 patients were discharged from the NICUs. We could collect the data from 197 children up to 6 years of CA. We excluded ten children who received growth hormone therapy. We analyzed 187 remaining children, of whom 44 were categorized into the 22–24 weeks group  16, Female (F) 28), and 143 in the 25–29 weeks group  .p < 0.05). In females, the z-score of birth weight and BL at birth were also significantly lower in the 22–24 weeks group than those in the 25–29 weeks group, but not in males. BW, BL, and BMI and their z-scores were significantly lower in the 22–24 weeks group than those in the 25–29 weeks group for all assessments. However, there were no significant differences in BW, BL, and BMI and their z-scores between the two female groups in all assessments, but there was a significant difference in BW z-score at 1.5 years of CA.For males, the z-scores of BW, BL, and BMI were significantly lower in the 22–24 weeks group than those in the 25–29 weeks group at all time points except for the BL z-score at CA of 3 years. For females, no significant differences in the z-scores of BW, BL, or BMI were seen between the 22–24 and 25–29 weeks groups at all time points except for BW z-score at CA 1.5 years .Linear regression analysis showed that the association between BW z-scores and both GA and the presence of SGA was seen in all age groups in both sexes, except for 6 years of CA in females . The assThis is, to our best knowledge, the first study to assess the influence of younger gestational age (<25 weeks’ gestation) on anthropometric indices using z-scores during the first 6 years of life among Japanese VLBW infants born at <30 weeks of gestation. We limited preterm infants by GA rather than birth weight e.g., <1500 g) to decrease bias, because more mature children with FGR were included. Our previous study demonstrated that FGR affected the z-scores of BW and BMI in males and of BW and BL in females during 6 years among same population assessed by ANOVA 00 g to d. AnotherA recent systematic review indicated that accelerated weight gain (defined as weight gain velocity during the first two years) significantly increased childhood obesity among preterm infants . SeveralPoor growth attainment in early childhood and early adolescence was reported in extremely preterm survivors ,21,22. IThe previously reported fat-free mass index was lower in males born preterm than that in females, whereas no difference in BMI was detected in either sex . PersistThis study has several limitations. The follow-up rate of participants was low (45%); we analyzed the characteristics of infants with and without follow-up ; and femIn conclusion, a younger gestational age had a greater influence on anthropometric parameters in males up to 6 years of age. A hierarchical analysis for GA and sex is needed to evaluate growth trajectory in VLBW, including periviable births.


There remains controversy regarding whether the growth charts constructed from data of term infants, such as those produced by the World Health Organization (WHO) standards, can appropriately evaluate the postnatal growth of preterm infants. This retrospective cohort study, conducted in the First Affiliated Hospital of Shandong First Medical University in Jinan China, aimed to compare the postnatal growth charts of singleton preterm and term infants using WHO standards at 40–160 weeks postmenstrual age (PMA). A total of 5,459 and 15,185 sets of longitudinal measurements  from birth to 160 weeks PMA were used to construct growth charts for 559 singleton preterm  and 1,596 singleton term infants (born at 40 weeks PMA), respectively, using the Generalized Additive Models for Location, Scale, and Shape (GAMLSS) method. Z-scores (prematurity corrected) were calculated using WHO Anthro software. Compared to WHO standards, all parameters of preterm infants were increased, especially in terms of length/height and weight; the gap between the two almost spanned two adjacent centile curves. Compared to term controls, the length/height, weight, and BMI of preterm infants were higher at 40 weeks PMA, surpassed by term infants at 52–64 weeks PMA, and quite consistent thereafter. The HC of preterm infants at 40–160 weeks PMA was quite consistent with both term controls and the WHO standards. The Z-scores for length/height, weight, and BMI of preterm infants relative to the WHO standards gradually decreased from 1.20, 1.13, and 0.74 at 40–44 weeks PMA to 0.67, 0.42, and 0.03 at 132–160 weeks PMA, respectively; Z-scores for HC of preterm infants rapidly decreased from 0.73 to 0.29 at 40–50 weeks PMA, and then fluctuated in the range of 0.08–0.23 at 50–160 weeks PMA. Preterm infants had higher growth trajectories than the WHO standards and similar but not identical trajectories to term infants during the first 2 years of life. These findings reemphasize the necessity of constructing local growth charts for Chinese singleton preterm infants. Growth impairment during early postnatal life can have permanent detrimental effects in later life, such as short stature, high blood pressure, and impaired neurodevelopment –4. A fulTo clarify the postnatal growth of singleton preterm infants and whether the WHO growth charts can adequately and appropriately evaluate the postnatal growth of preterm infants in Jinan, Shandong Province, China, during the first 2 years of life, we constructed postnatal growth charts using longitudinal data from a specific cohort of singleton preterm infants during the first 2 years of life. These data were compared to data from a cohort of singleton term infants in the same center and the WHO standards .Longitudinal growth data were retrospectively collected from an ongoing cohort study for preterm infants and from routine well-child visits for term infants conducted in the Department of Developmental Pediatrics and Child Health Care, the First Affiliated Hospital of Shandong First Medical University  in Jinan, Shandong Province, China.The inclusion criteria of subjects were as follows: (1) preterm infants [postmenstrual age (PMA) at birth ≤ 36 weeks] and term infants (PMA at birth = 40 weeks) born between January 1, 2014, and June 30, 2018; (2) singleton; (3) no congenital malformations or syndromes; (4) ≥4 follow-up visits before 160 weeks PMA; and (5) PMA at first follow-up visit ≤ 66 weeks  and PMA at last visit ≥118 weeks . The exclusion criteria were as follows: (1) twins; (2) infants with congenital malformations or syndromes; (3) infants with a first follow-up visit after 66 weeks or lost to follow-up before 118 weeks PMA. A flowchart of the sampling process for eligible singleton preterm and term infants is shown in The feeding practices during the early postnatal period for preterm infants were based on the “Nutrition Practice Care Guidelines for Preterm Infants in the Community 2013” and “Chinese Society of Parenteral and Enteral Nutrition (CSPEN) guidelines for nutrition support in neonates” , 12. BriThe parental baseline data collected were as follows: age, education, ethnicity, maternal obstetric history, and mode of delivery. The data on the infants were as follows: sex, PMA at birth (week), and length (cm) and weight (kg) at birth [head circumference (HC) was not measured at birth]. These data were collected from questionnaires filled in by parents at the first follow-up visit. Except for parental education, other information obtained through questionnaires was further verified by birth certificates or medical records. If there were discrepancies, data from birth certificates or medical records were chosen to be documented.It was recommended that infants should be followed up at term (40 weeks PMA) and at 1, 3, 4, 5, 6, 9, 12, 15, 18, and 24 months of age (corrected age for preterm infants). Anthropometric measurements were taken at each follow-up visit by two trained staff members working in pairs. For children ≤ 2 years old (corrected age for preterm infants), the length was measured with the infant lying down and using an infantometer . For children > 2 years old (corrected age for preterm infants), and height was measured with the infant standing upright, using a stadiometer . Weight was measured using an electronic scale . HC was measured with a tape measure , which was replaced once a month. Each staff member independently measured and recorded a complete set of measurements. Thereafter, the two staff members compared their readings and recorded the mean of each pair of readings. The maximum allowable differences were as follows: length/height, 7 mm; weight, 100 g; and HC, 5 mm. If there were greater discrepancies, both staff members independently measured and recorded a second and, if necessary, a third set of readings for the variable (s) in question.The Z-scores of length/height, weight, HC, and BMI  of both the preterm and term infants obtained from each follow-up visit were calculated using WHO Anthro  in R sofThe Z-scores of length and weight at birth according to the INTERGROWTH-21st International Newborn Size References/Standards  were calculated using the International Newborn Size at Birth Standards application .Growth charts  of length/height, weight, HC, and body mass index (BMI) of the preterm and term infants at 40–160 weeks PMA stratified by sex were constructed using Generalized Additive Models for Location, Scale, and Shape (GAMLSS) , 16 in RDifference values for P50 (the 50th percentile) of each growth parameter were calculated between the preterm and term infants, and between the preterm infants and WHO standards.The Z-scores were assigned to 11 PMA clusters based on the PMA at the follow-up visit: PMA ≥ 40 and <44, ≥44 and <50, ≥50 and <56, ≥56 and <62, ≥62 and <68, ≥68 and <76, ≥76 and <84, ≥84 and <94, ≥94 and <112, ≥112 and <132, ≥132 and ≤ 160 weeks. The Z-scores were compared between the preterm and term infants, and we also observed whether the Z-scores were >0 (indicating higher growth levels than the WHO standards).n) and percentage (%). Baseline characteristics were compared between the preterm and term infants, with independent-sample t-tests and chi-square tests being used to compare continuous and categorical variables, respectively. The level of significance was set at P ≤ 0.05.The statistical analysis was conducted using SPSS software version 21 . Continuous variables are presented as mean (SD), while categorical variables are presented as frequency  for 559 eligible preterm infants, and a further 15,185 for 1,596 term infants, from birth to 160 weeks PMA. That was, the mean of the follow-up visits was 9.77 for preterm infants and 9.51 for term infants (including that at birth). The comparison of the baseline characteristics between the preterm and term infants is shown in The mean PMA at birth for the preterm infants was 33.84 weeks ; 71.0% were late preterm infants, and almost half of them had normal birth weight (NBW). The general baseline characteristics of the preterm and term infants were similar. Almost all infants were of Han ethnicity (>98%), and most had well-educated parents (>85% had at least one parent with higher education). The proportions of boys and infants born by cesarean section were significantly higher among the preterm infants than the term infants. The parents of the preterm infants were older than those of the term infants, and there were fewer mothers experiencing their first gestation and first birth in the preterm infant cohort. The HAZ and WAZ at birth of both the preterm and term infants according to the INSR/S  were >0,Growth charts  of length/height, weight, HC, and BMI for the preterm and term infants stratified by sex at 40–160 weeks PMA are shown in The difference values for P50 of each growth parameter between the preterm and term infants, and the preterm infants and WHO standards, are shown in As shown in 2. After 144 weeks PMA, the weight of the preterm infants was rapidly approaching the WHO standards, the BMI became smaller than the WHO standards, while the height tended to even larger than the WHO standards.In contrast, at 40–144 weeks PMA, the preterm infants had a greater length, weight, and BMI than the WHO standards . The BMIAs shown in The Z-scores of each growth parameter for the preterm and term infants according to the WHO standards are shown in For preterm infants, the largest mean values of HAZ, WAZ, HCZ, and BAZ were 1.20, 1.13, 0.73, and 0.74, respectively, at 40–44 weeks PMA. HAZ, WAZ, and BAZ then gradually decreased. At 132–160 weeks PMA, HAZ, WAZ, and BAZ were at their lowest values . HCZ rapidly decreased from 0.73 at 40–44 weeks PMA to 0.29 at 44–50 weeks PMA, and it then fluctuated in the range of 0.08–0.23 at 50–160 weeks PMA.Although there was a difference in the mean PMA between the preterm and term infants in each PMA cluster, we still compared the Z-scores of the preterm and term infants as, unlike the linear relationship between growth parameters and PMA, there is no such direct linear relationship between the Z-scores of growth parameters and PMA.Before 50 weeks PMA, the HAZ, WAZ, and BAZ were all significantly larger for the preterm infants than the term infants. At 50–62 weeks PMA, the HAZ, WAZ, and BAZ were similar between the preterm and term infants, but thereafter, the preterm infants mostly had smaller values (except for BAZ at 132–160 weeks PMA) than the term infants .At 44–50 weeks PMA, the HCZ of the preterm infants was similar to that of the term infants. Subsequently, the term infants had larger HCZ than the preterm infants .Our retrospective cohort study benefits from the stringent inclusion and exclusion criteria used for the preterm infants and their control term infants. For example, all subjects were singletons born during the same period of time at the same center and had a sufficient follow-up frequency and duration. Furthermore, the selection of control subjects who were born at strictly 40 weeks PMA took into account the fact that term infants born at 37–41 weeks PMA might have varying growth trajectories. Thus, the growth level of the selected term infants represented the ideal growth level of healthy local children. In addition, we selected PMA as the age variable, which allowed for accurate and specific comparisons among the preterm infants, term infants, and the WHO standards.The study demonstrated that the postnatal growth trajectories of the preterm infants were different from the WHO standards and similar but not identical to those of their term counterparts. Overall, the length/height, weight, HC, and BMI of the preterm infants were larger than the WHO standards, especially length/height and weight, of which the gap between the preterm infants and the WHO standards during the corrected term to 2 years old could span two adjacent centile curves. The growth trajectories of the preterm infants were more consistent with their term counterparts than the WHO standards, especially for length/height and weight after around 50 weeks PMA.Kang et al. lately published a multi-center cohort study conducted in Sichuan, China, which also made a comparison of the postnatal growth between preterm and term infants using longitudinal growth data during 40–88 weeks PMA . DiffereThe conclusion that preterm infants had higher postnatal growth than international growth charts and consistent growth with term counterparts, was different from most previous studies, which concluded that children born preterm tended to exhibit less growth than children born full-term and that these children had an increased risk of growth retardation –22. ThesThese findings indicated that Chinese preterm infants might also have higher growth levels than international growth standards during the first 2 years of life, and these findings include some that were found in previous population-based studies with term infants in China. For example, both the Chinese National Growth Survey (CNGS) 2005 and 2015, which constructed the growth charts of Chinese children under 7 years of age with cross-sectional data of urban singleton term-born children from nine cities in China, had concluded the higher growth levels than the WHO standards , 25. TheGiven the concern of the relationship between excessive catch-up growth and subsequent development of metabolic problems later in life , this hiAs we have known, there is a trend of constructing and using international references/standards of optimal physiological growth in the evaluation of growth level and health status of children worldwide over the past decades, the most prominent of which is the WHO standards for term infants , 27, FenThere are some limitations to this study. First, our study was conducted in a single center in Jinan, China, although the First Affiliated Hospital of Shandong First Medical University is a tertiary public hospital that covers the prevention and treatment of maternal and child diseases in Shandong province, China. Second, given that the mean gestational age among the preterm infants was 33.84 weeks and the fact that 71% of the preterm infants were late preterm infants, our study might not be sufficiently powered to evaluate the specific growth of extremely preterm, moderate preterm, and ELBW/VLBW infants. Third, infants aged <4 months in the WHO cohort were exclusively breastfed , while igenetically different than that of the WHO cohort, which was thought to represent the ideal growth pattern.In a future study, we will establish a birth cohort of preterm infants with a larger sample size, more detailed documentation of growth data and potential bias factors, and increased duration of follow-up, and we will construct stable and reliable growth charts stratified by sex and PMA at birth.In conclusion, the singleton preterm infants in a single-center in China had specific growth trajectories that were obviously higher than the WHO standards and similar but not identical to their term counterparts during the first 2 years of life, which reemphasizes the necessity of constructing growth charts especially for a local Chinese population of singleton preterm infants.The original contributions presented in the study are included in the article/The studies involving human participants were reviewed and approved by Medical Ethics Committee of the First Affiliated Hospital of Shandong First Medical University. Written informed consent to participate in this study was provided by the participants' legal guardian/next of kin.LZ made a major contribution to the conception and design of the study, the collection, analysis, and interpretation of the data, and the writing and revision of the manuscript. J-GL made a substantial contribution to the data analysis. SL revised the manuscript for important intellectual content. JS, N-NG, QW, H-YZ, H-JL, X-DC, and YC contributed to the acquisition of data. YL made a substantial contribution to the supervision and project administration of the study and the writing and revision of the manuscript. All authors read and approved the final manuscript.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


Scd-1, Fas) and transcription factors . In addition, alterations in the gene expression of cytochrome P450 proteins, particularly Cyp2e1 and Cyp4a, together with increased Nrf2, contributed to the improvement of hepatic lipid metabolism after empagliflozin administration. Decreased circulating levels of fetuin-A improved lipid metabolism and attenuated insulin resistance in the liver and in peripheral tissues. Our results highlight the beneficial effect of empagliflozin on hepatic lipid metabolism and lipid accumulation independent of obesity, with the mechanisms understood to involve decreased lipogenesis, alterations in cytochrome P450 proteins, and decreased fetuin-A. These changes help to alleviate NAFLD symptoms in the early phase of the disease and before the onset of diabetes.Recent studies suggest that treatment with SGLT-2 inhibitors can reduce hepatic lipid storage and ameliorate non-alcoholic fatty liver disease (NAFLD) development beyond their glycemic benefits. However, the exact mechanism involved is still unclear. We investigated the hepatic metabolic effect of empagliflozin (10 mg/kg/day for eight weeks) on the development of NAFLD and its complications using HHTg rats as a non-obese prediabetic rat model. Empagliflozin treatment reduced neutral triacylglycerols and lipotoxic diacylglycerols in the liver and was accompanied by significant changes in relative mRNA expression of lipogenic enzymes ( Non-alcoholic fatty liver disease (NAFLD) is a hepatic manifestation of metabolic syndrome, obesity, and various prediabetic states and is characterized by increased hepatic lipid accumulation (˃5%). Considered an independent risk factor for cardiovascular events ,2, NAFLDAlthough approximately 50% of obese individuals and up to 70% of diabetic patients exhibit characteristic features of fatty liver, NAFLD can also occur in non-obese individuals. The prevalence of non-obese NAFLD ranges widely from 3 to 30% .Despite the high prevalence of NAFLD, an effective therapy has yet to be developed. However, drugs with antidiabetic properties, such as thiazolidinediones, glucagon-like peptide-1 (GLP-1) analogs, and metformin, can attenuate the development and progression of NAFLD. Sodium-glucose cotransporter-2 (SGLT-2) inhibitors are a class of drugs that exert pleiotropic effects on NAFLD. One of the most promising is empagliflozin ,6. AccorIn animal studies, empagliflozin administration has been shown to decrease hepatic lipid accumulation and ameliorate signs of NAFLD in models of diet-induced obesity and diet-induced NAFLD followed by T2DM onset ,12,13. IHowever, the potential advantageous effects of empagliflozin on NAFLD have not yet been studied independently of obesity or before the onset of hyperglycemia. To address this deficit, our study investigated the metabolic effect of empagliflozin on hepatic lipid and glucose metabolism in hereditary hypertriglyceridemic (HHTg) rats, a non-obese prediabetic rat model ,15. Thesp ˂ 0.05) but had no effect on drink intake.Prior to empagliflozin administration, there were no differences in initial body weight, glucose or serum triacylglycerols (TAG) between the control Wistar and Wistar + empa groups or between the prediabetic HHTg and HHTg + empa groups. Empagliflozin administration slightly increased food intake . Empagliflozin treatment decreased glycemia and improved glucose tolerance, manifesting in significantly reduced serum fasting glucose, non-fasting glucose, and AUC in the empagliflozin-treated groups of rats . Empagliglycogen . In Wistglycogen . HoweverFas and Scd1 and the lipogenic transcription factor Srebf1 in the liver decreased significantly after empagliflozin treatment  and fatty acid metabolism (Pparγ), it had no effect on genes related to lipid oxidation or transport . Inhibition of de novo lipogenesis in the liver is understood to be an additional benefit of empagliflozin treatment, helping to reduce hepatic lipid accumulation. In our study, empagliflozin administration affected gene expression of the main lipogenic enzymes  and transcription factors . Suppression of Scd1 after empagliflozin was observed in another study with obese mice [(−/−) mice demonstrated that elevated expression of Cyp1a1 protects against NAFLD development, while dysregulation of the Cyp1a1 gene is reportedly involved in the transport and metabolism of lipids [Srebp1 and Pparγ plays a potential role in reducing hepatic lipid accumulation via suppression of lipogenesis, in turn contributing to improved hepatic insulin sensitivity. Reduced SREBP1 after empagliflozin treatment has also been demonstrated in studies of OLETF rats [In our study, although empagliflozin treatment modulated genes related to lipid synthesis  may also be involved in improving hepatic lipid metabolism following empagliflozin administration, thus alleviating symptoms of NAFLD. Some CYP enzymes are effective in the pathogenesis of NAFLD. Recent studies suggest that CYP2E1 plays an indispensable role in the development and progression of NAFLD , adaptinf CYP2E1 . Therefof CYP2E1 , as wellf CYP2E1 . Inhibitf CYP2E1 . Our resAlterations in CYP enzymes after empagliflozin treatment can affect hepatic metabolites and oxidative stress products, thus modulating their pharmacotherapeutic effects. To the best of our knowledge, the effect of empagliflozin on cytochrome P450 proteins has yet to be investigated.The beneficial effect of empagliflozin on NAFLD and its metabolic complications can be measured based on the levels of secreted hepatokines, which directly modulate glucose and lipid metabolism in the liver and establish crosstalk with adipose and muscle tissues. In the present study, empagliflozin administration markedly decreased circulating levels of fetuin-A in both strains. Surprisingly, however, it only slightly decreased levels of FGF21. Alterations in hepatokine secretion play a role in the pathogenesis of NAFLD and its complications . EpidemiFgf21 decreased in association with elevated fat utilization [Compared to fetuin-A, FGF21 is a protective factor that regulates lipid metabolism and reduces lipid accumulation in hepatocytes through an insulin-independent pathway . FGF21 alization . During −/− mice showed that empagliflozin decreases liver mRNA expression of Mcp-1 [Mcp-1 was unchanged. In addition, we observed decreased MCP-1 concentration in the liver, while the ApoE−/− mice study did not measure it. According to our results, decreased hepatic MCP-1 levels after empagliflozin treatment can be supported by improved oxidative stress and increased mRNA expression of Nrf2. Another study involving diet-induced obese mice found that the anti-inflammatory action of empagliflozin led to the suppression of Nfκb and Tnfα genes in the liver [In the present study, empagliflozin treatment in prediabetic rats decreased pro-inflammatory MCP-1, attenuating the hepatic pro-inflammatory state. Decreased MCP-1 in the liver can help to reduce macrophage infiltration, leading to alleviation of hepatic steatosis and inflammation. In addition, MCP-1 regulates the expression of other pro-inflammatory cytokines, such as TNFα and IL-6. A recent study of diet-induced ApoEof Mcp-1 ; howeverhe liver . A variehe liver . A crucial regulator of the cellular defense against oxidative stress is transcription factor Nrf2 . Nrf2 alEmpagliflozin regulates oxidative stress by stimulating antioxidant enzyme activity, SOD, and GPx via upregulation of Nrf2 rather than the direct inactivation of free radicals. In addition, increased activity of SOD and GPx may play a role in decreasing lipid peroxidation in the liver by participating in the removal of lipoperoxidation products. Increased activity of GPx after empagliflozin treatment was also linked to elevated glutathione levels and a suitable reduced/oxidized glutathione ratio. Glutathione is a sensitive marker of hepatic oxidative damage. However, in our study, we observed a trend for increased hepatic levels of glutathione following empagliflozin treatment. As compared to other experimental studies, empagliflozin suppresses hepatic oxidative stress via the activation of Nrf2/PPARγ crosstalk in mice, but it is about alcohol-induced liver damage . Future research directions of experimental studies should be focused on investigating the circulating markers—hepatokines, adipocytokines, inflammatory parameters, or others, that would well reflect hepatic metabolic disorders in relation to the NAFLD development. A reliable parameter that would well reflect the stages of NAFLD development is still lacking in the human field of medicine.All experiments were performed in agreement with the Animal Protection Law of the Czech Republic (311/1997), which is in compliance with the European Community Council recommendations (86/609/ECC) on the use of laboratory animals and approved by the Ethics Committee of the Institute for Clinical and Experimental Medicine . The study was performed on 6-month-old male Wistar rats  as the control group and 6-month-old male HHTg rats  as the non-obese prediabetic model. Rats were kept under temperature (22 °C) and humidity-controlled conditions under a 12/12 h light/dark cycle with free access to food  and drinking water. Wistar and HHTg rats were randomized into groups with or without empagliflozin treatment mixed as part of a standard diet at a dose of 10 mg/kg b.wt. for 8 weeks. At the end of the experiment, rats were sacrificed after light anesthesia (zoletil 5 mg/kg b.wt.) in a postprandial state. Aliquots of serum and tissue samples were collected and stored at −80 °C for further analysis. Serum levels of triacylglycerols (TAG), glucose, NEFA, ALT, and AST, as well as total and HDL cholesterol, were measured using commercially available kits . Serum insulin, glucagon, leptin, and HMW adiponectin concentrations were determined using the Rat Insulin ELISA kit  and the Rat Adiponectin ELISA kit . Serum MCP-1, TNFα, IL-6, and hsCRP were also measured using rat ELISA kits . Serum β-hydroxybutyrate concentrations were measured using a colorimetric assay kit . Serum levels of the hepatokines FGF21 and fetuin-A were determined by rat ELISA kits .For the oral glucose tolerance test (OGTT), blood glucose was determined after a glucose load (300 mg/100g b.wt.) administered intragastrically after overnight fasting. Blood was drawn from the tail before the glucose load at 0 min and at 30, 60, and 120 min thereafter. musculus soleus by measuring the incorporation of 14C-U glucose into glycogen as described previously [For the determination of TAG and cholesterol in tissues, samples were extracted in chloroform/methanol. The resulting pellet was dissolved in isopropyl alcohol, with TAG content determined by enzymatic assay . To determine DAG in the liver, samples were extracted in dichloromethane/methanol. The resulting pellet was dissolved in isopropyl alcohol and isolated by thin-layer chromatography. The content of separated DAG was determined by enzymatic assay . As a marker of skeletal muscle insulin sensitivity, basal and insulin-stimulated glycogen synthesis was determined ex vivo in the isolated eviously . 2O. Aliquots were used to measure glycogen by glucose oxidase assay .For the determination of hepatic glycogen concentration, liver tissues were hydrolyzed in 30% potassium hydroxide (KOH) and precipitated by 96% ethanol overnight. The precipitate was then centrifuged, washed with ethanol, and dissolved in HThe levels of reduced (GSH) and oxidized (GSSG) forms of glutathione were determined by the HPLC method with fluorescence detection using an HPLC diagnostic kit . The activity of antioxidant enzymes superoxide dismutase and glutathione peroxidase were analyzed using Cayman Chemicals assay kits . MDA, a parameter of lipid peroxidation, was assessed by the HPLC method with fluorescence detection; 4-HNE, a sensitive product of lipoperoxidation, was determined using a rat ELISA kit .TM 7 Real-Time PCR System . Relative expressions were determined after normalization against β-actin and Hprt1 as an internal reference and calculated using the 2−ΔΔCt method. Results were run in triplicate.Total RNA was isolated from tissues using RNA Blue . Reverse transcription and quantitative real-time PCR analysis were performed using the TaqMan RNA-to-CT 1-Step Kit, TaqMan Gene Expression Assays , and the ViiAMiddle plane sagittal hepatic slices were fixed in formaldehyde for 48 h and processed using standard techniques into paraffin blocks. Sections were 3 μm thick and stained using routine hematoxylin and eosin staining and evaluated by a histopathologist.p < 0.05. All results are expressed as mean ± SEM. Statistical analysis was performed using StatSoft Statistica 14 software .Two-way ANOVA was used to analyze the individual and combined effects of treatment and strain for treatment-vs.-strain interactions. All data analyzed were of normal distribution. Fisher’s LSD post-hoc test was used for variables showing evidence of treatment-vs.-strain interactions. The test was adjusted for multiple comparisons to determine whether empagliflozin treatment would significantly influence hepatic metabolic parameters in HHTg and Wistar strains. Statistical significance was set at a value of Scd1, Fas, Srebp1, and Pparγ. However, the genes involved in lipid oxidation were not affected. We speculate that increased expression of the transcription factor Nrf2 contributed to the improvement of hepatic lipid metabolism and the alleviation of oxidative stress. Changes in the expression of the cytochrome P450 family proteins, in particular Cyp2e1, Cyp1a1, Cyp4a1, and Cyp4a2, which are involved in the pathogenesis of NAFLD, had a beneficial effect on lipid metabolism and insulin signaling after empagliflozin administration. In addition, markedly reduced levels of the important hepatokine fetuin-A were associated with improved insulin sensitivity and low-grade inflammation.Our results show that empagliflozin treatment in prediabetic rats can mitigate hepatic steatosis by reducing hepatic triacylglycerols as well as lipotoxic intermediates. In the liver, empagliflozin modulated the expression of genes involved in lipogenesis and lipid storage, including Our results highlight the therapeutic potential of empagliflozin in the treatment of NAFLD and its complications in the early phase of development independent of obesity and before the onset of diabetes. Thus, empagliflozin could ameliorate or delay NAFLD development in prediabetic patients; however, more clinical evidence is needed.


To investigate predictors of low birth satisfaction in a sample of Iranian postpartum women.This was a cross‐sectional study.This study was conducted on 767 postpartum women using a convenience sampling method between June and September 2019. We collected data on socio‐demographic variables, maternal well‐being, fear of childbirth and birth satisfaction. We used multiple linear regression analyses to determine predictors of low birth satisfaction.The women who gave birth by elective caesarean, emergency caesarean and vaginal birth were 13.2%, 19.06% and 67.8%, respectively. Predictors of low birth satisfaction were primiparity , low level of well‐being , low [−0.193 ] and moderate [−0.143 ] satisfaction with pregnancy, moderate satisfaction with spouse's emotional/financial support [−0.150 ], emergency caesarean [−0.086 ], severe fear of childbirth [−0.315 ] and long interval between admission to hospital and giving birth [−0.188 ]. Childbirth dissatisfaction reduces maternal desire for subsequent pregnancies , developed in 2014 by Martin and Martin, has been recommended as the measure of choice for assessing women's experiences of childbirth by the International Consortium for Health Outcome Measures (ICHOM)  . The women also rated their level of satisfaction with pregnancy, spouse's emotional/financial support and conjugal and sexual satisfaction on a five‐point Likert scale ranging from 1–5 (1 = not satisfied to 5 = very satisfied). With regard to the satisfaction with pregnancy, we asked the following question: Given the health problems you faced during your pregnancy, how satisfied are you with your pregnancy? Data .3.2.2The Birth Satisfaction Scale – Revised (BSS‐R) consists of ten items that participants respond to on a four‐point Likert scale which ranges from 0–4 (0 = strongly disagree to 4 = strongly agree). The minimum and maximum scores of the questionnaire are 0 and 40, respectively. The reliability of BSS‐R has been confirmed . The scale was translated into Persian (Persian BSS‐R) in 2019, and its validity and reliability were investigated. The results of the confirmatory factor analysis on the scale confirmed the structure proposed by the scale developers. The reliability of the Persian BSS‐R has been confirmed  . The scale's total score ranges from 0–165, with larger scores corresponding to greater levels of fear. The W‐DEQ was translated into Persian. The validity assessment of the Persian W‐DEQ indicates that the scale consists of six factors. It showed moderate correlations with the Childbirth Attitude Questionnaire and the State‐Trait Anxiety Inventory. The Cronbach's alpha coefficients of the scale and its factors were in the acceptable range (between 0.633–0.919) . Each question is scored from 0 (having good feelings at no time)–5 . The scale's total score ranges from 0–25 which is converted to a scale of 0–100. A score of 50 has been used as the cut‐off score in screening for depression. In the case of persons with scores less than 50, referral for further assessment has been recommended. Mortazavi and colleagues investigated the validity and reliability of the Persian WHO‐5 Well‐Being Index in pregnant women. The results confirmed the reliability and unidimensionality of the scale  . Sample characteristics are presented in Table The mean values of age, education (in years), gestational age, birth weight and admission to delivery duration were 28.5 ± 6.1, 11.2 ± 3.7, 39.0 ± 1.3, 3170.8 ± 491.8 and 7.2 ± 7.5, respectively. The mean Wijma scores, WHO‐5 well‐being scores and birth satisfaction scores were 73.9 ± 23.0, 53.2 ± 25.2 and 23.0 ± 7.1, respectively. Fifty‐nine point six percent (59.6%) of the women were multiparous, and of these, 34.2% and 25.4% were para 2 and para 3, respectively. The correlation between birth satisfaction total scores and the duration between admissions to hospital and giving birth was −0.305 (In Table p = .012). Women who had emergency caesarean accompanied by a doula reported a lower level of birth satisfaction than those without a doula (p = .045)  Table .5In this study, we investigated the predictors of low birth satisfaction in early postpartum. Our results indicate that primiparity, emergency caesarean, long duration from admission to delivery, low level of well‐being, low and moderate satisfaction with pregnancy, moderate satisfaction with spouse's emotional/financial support and severe fear of childbirth are predictors of low birth satisfaction. The proportion of the variance in birth satisfaction explained by the demographic and psychological variables was 14.5% and 28.8%, respectively. The total explained variance by all the independent variables was 36.8%. Our results can be compared to those obtained by Nahaee and colleagues in a study in Tehran which investigated, pre‐ and during‐labour predictors of low birth satisfaction. They found that the proportion of the variance explained by during‐labour, pre‐labour and overall variables was 75%, 14% and 76%, respectively .The authors have no conflict of interest.MM collected the data and wrote the first draft of the manuscript. FM analysed the data and wrote the final draft of the manuscript. The authors have read and approved the manuscript.The Ethics Committee of Sabzevar University of Medical Sciences has reviewed and approved this study . All procedures were performed in accordance with the guidelines of Sabzevar University of Medical Sciences, which is in accordance with the Declaration of Helsinki. Women who consented to participate in the study signed an informed consent form and were assured about the confidentiality of their information.Supplementary MaterialClick here for additional data file.


Early essential newborn care (EENC) was introduced to medical practice in China in 2016, but the number of medical institutions that have put EENC into practice remains low due to insufficient clinical evidence and the absence of awareness among health professionals. This study aimed to explore the effect of EENC on physiological variables and sleep state among newborn infants and to provide evidence to support the implementation of EENC.A quasi-experimental design was conducted among 182 newborn infants in a tertiary maternity hospital in China from May 2020 to January 2021. A total of 91 newborn infants were included in the intervention group, and 91 were included in the control group to receive EENC or routine birth care, respectively.p < 0.05). The time of first breathing after birth in the intervention group was earlier than that in the control group , and the infants had a better sleep state at 30 min, 60 min, 90 min, and 120 min after birth (p < 0.05).The newborn infants in the intervention group had a lower incidence of hypothermia than those in the control group at 75 min, 90 min, 105 min, and 120 min after birth , registration number:  Reducing the mortality rate of newborn infants is a major challenge worldwide, especially in the Western Pacific Region, where a newborn infant dies every 2 minutes . There hAction plan for healthy newborn infants in the Western Pacific Region (2014–2020) was formulated by the World Health Organization Western Pacific Regional Office (WHO/WPRO) in 2013 [The Region 204–2020 waRegion 204–2020 wa in 2013 . The WHO in 2013 .Routine birth care has many outdated newborn care practices, such as unnecessary suctioning and immediate cord cutting, while drying and first breastfeeding are delayed, thereby exposing newborn infants to hypothermia, breathing and circulatory problems, infection and brain haemorrhage .ComparedEENC has been introduced and implemented in over 100 medical institutions in China since 2015 . TherefoA quasi-experimental design was implemented with the aim of evaluating the effectiveness of EENC (intervention) compared with routine birth care (control). This study was conducted in a tertiary maternity hospital in Sichuan Province in China from May 2020 to January 2021. This hospital has two labour wards, which were randomly assigned to the intervention group and control group using a table of random numbers. As these two wards are in different hospital areas, any contamination can be excluded. Before the current study, neither ward had implemented EENC or received EENC coaching. In addition, the composition of health professionals was comparable between the two groups, and the facilities of both groups were identical.All pregnant women who were eligible for this study and were willing to provide informed consent were recruited from the maternity ward before birth, and their babies were recruited into the study after birth. In this hospital, the pregnant women chose the labour ward to give birth based on the hospital areas where they received antenatal care. The choice of which hospital area to receive antenatal care depended entirely on the preference of the pregnant woman, such as the consideration for the distance to the hospital area from home. Hence, the participants were nonrandomly assigned into the two groups. However, neither the pregnant women nor their obstetricians knew the group assignment in advance.The inclusion criteria were as follows: (1) female age ≥ 18 years, (2) gestational age ≥ 37 weeks and < 42 weeks, (3) singleton pregnancy, (4) vaginal delivery, (5) the birth weight of newborn infants was between 2500 g and 4000 g estimated by obstetric ultrasound, and (5) provision of informed consent.The exclusion criteria were as follows: (1) women with severe pregnancy complications and/or underlying disease, (2) women with medical indications against breastfeeding, such as acquired immunodeficiency syndrome, syphilis, and history of breast surgery, (3) women with no intention to breastfeed, (4) newborn infants with deformities, and (5) newborn infants who needed to be transferred to the neonatal intensive care unit (NICU) immediately after birth. In addition, if the actual birth weight of newborn infants was < 2500 g or > 4000 g, they would be excluded from the data analysis.The current study is part of a larger trial and focused only on the effect of EENC on the physiological variables and sleep state of newborn infants. The association of EENC with other outcomes will be published separately.Coaching for the First Embrace: Facilitator’s Guide : Module 2 [Participants in the intervention group received EENC interventions supported by trained midwives from November 2020 to January 2021. These midwives were trained by national and provincial facilitators from May 2020 to October 2020, following the guidance of Module 2 . Before The core EENC interventions for all newborn infants are as follows: (1) drying the newborn infant immediately and thoroughly within 5 s after birth, (2) immediate and continuous skin-to-skin contact between the mother and newborn infant for at least 90 min, (3) appropriately timed clamping and cutting of the cord, (4) early exclusive breastfeeding, and (5) routine care, namely, eye care, vitamin K, immunizations, weighing and examinations.Participants in the control group received routine birth care supported by the midwives. The flow of routine birth care was as follows: (1) drying of the newborn infant, (2) placing of the newborn infant on the radiant warmer to keep warm for 20 min, during which the umbilical cord was clamped and the weight and length were measured, (3) vaccination of the infant, and (4) skin-to-skin contact between the mother and newborn infant and breastfeeding of the infant after the third stage of labour.The current study was a single-blinded trial. It was impossible to blind the midwives and data collectors in the labour wards. Hence, only participants were blinded.This study used a self-designed form to collect the sociodemographic information of participants, including women and their newborn infants. Data on the age, educational level, gestational age, previous obstetric history, height, and weight of the women were collected by data collectors before birth. In addition, data on the sex, length, and birthweight of infants were collected by data collectors after birth.The primary outcome of the current study was the incidence of hypothermia among newborn infants within the first 2 h after birth. The body temperature of newborn infants was measured with an ear thermometer by the data collectors every 15 min within the first 2 h after birth. If the body temperature of the newborn infant was lower than 36.5 °C, hypothermia was considered . Both boThe secondary outcomes were the time of first breathing after birth, the incidence of low oxygen saturation, the incidence of abnormal neonatal heart rate, and sleep state in newborn infants. The time of first breathing after birth in the newborn infants was measured with the stopwatch and then recorded by the data collectors. In addition, low oxygen saturation and abnormal heart rates within the first 2 h after birth were measured with the disposable probe cover on pulse oximetry and recorded by data collectors every 10 min. According to the newborn resuscitation guidelines of China , the defAll data collectors in this study were women with at least a medical-related undergraduate degree. They obtained permission to enter the labour ward and collect data from both the ethics committee of the hospital and the participants. The data collectors received training on methods for collecting data by researchers.α = 0.05, β = 0.1). Considering the 10% drop-out rate, the minimum sample size was 182 mother–newborn pairs, with 91 pairs in each group. To reduce sampling error [PASS 15 software was used to calculate the sample size. The sample size was estimated according to the primary outcome of this study, which is the incidence of hypothermia within 2 h after birth in the intervention group and the control group. The pilot study showed that the incidences of hypothermia within 2 h after birth were 11.1 and 33.3% in the intervention group and control group, respectively, so the total sample size was 164 mother–newborn pairs , and a t test was then used to analyse significant differences between the two groups. Continuous data with a skewed distribution are presented as the median , and the Mann–Whitney U test was used to analyse differences between these two groups. Pearson’s chi-square or Fisher’s exact test was used to compare categorical variables. Repeated measures analysis of variance was used to compare the trends of changes in body temperature among newborn infants in both groups across the 2-h post-delivery period. A p > 0.05), the details of which are listed in Table Between November 2020 and January 2021, 203 pregnant women who met the inclusion criteria were recruited for this study. After exclusions, 182 mother–newborn pairs who received either EENC (91 pairs) or routine birth care (91 pairs) were included at data analysis stage. The flow diagram of participants in this study is shown in Fig. p = 0.045), 90 min , 105 min , and 120 min  after birth, while it was higher at 15 min  after birth. There was no significant difference in the incidence of hypothermia of newborn infants at 30 min, 45 min, or 60 min after birth between the two groups (p > 0.05) . The infant body temperature in the intervention group was lower than that in the control group at 15 min  and 30 min  after birth but higher than that of the control group at 75 min , 90 min , 105 min , and 120 min  after birth. Moreover, the newborn infant body temperature was different at the different time points (p < 0.001), and the interaction effect was significant (p = 0.034), indicating that newborn infant body temperature was influenced by time and the intervention. The trend for changes in body temperature over time was also different between the two groups . There were 10 (11.0%) and 12 (13.2%) newborn infants who had abnormal heart rates in the intervention group and control group, respectively, within 30 min after birth. Additionally, 5 (5.5%) and 8 (8.8%) newborn infants had low oxygen saturation in these two groups, but no significant difference was found  . However, at 30 min, 60 min, 90 min, and 120 min after birth, more newborn infants in the intervention group had drowsy, light sleep, and deep sleep states than those in the control group, and the difference was statistically significant (p < 0.001), which indicates that the newborn infants in the intervention group had better sleep states than those in the control group after birth.Table This study compared the effect of EENC and routine birth care on the physiological variables and sleep states of newborn infants in a tertiary maternity hospital in Sichuan Province, China, and the results showed that EENC can reduce the incidence of hypothermia, promote breathing initiation, and improve the sleep state of newborn infants.’s study [Neonatal hypothermia is associated with the major causes of neonatal deaths, including severe infections, prematurity, and birth asphyxia . EENC sh’s study  in the i’s study .Previous studies showed that approximately 8 to 12% of newborn infants initiated breathing after being stimulated , 33. If The levels of corticosteroids and catecholamines in newborn infants increase after birth and lead to stress responses, which could increase the consumption of energy . SleepinTo our knowledge, the current study is the first study to be designed as a controlled study to explore the effect of EENC on physiological variables and the sleep states of newborn infants in China, but there were some limitations that need to be addressed in future work. First, the follow-up period was relatively short, so the long-term effect of the implementation of EENC on newborn infants remains unknown. Second, although the sample size of this study was calculated according to pretrial estimates, the sample size was not large. Third, only one hospital was selected to carry out the study, and the results may not be representative. Last, the results of nonrandomized studies are more likely to be affected by confounding bias, although many attempts were made to avoid them. Therefore, a long-term, multi-centre, and large-sample randomized controlled trial should be considered in the future to explore the clinical effect of EENC and better ensure the health of newborn infants.Compared to routine birth care, EENC can better maintain the body temperature of the newborn infant within the normal range, reduce the incidence of hypothermia, promote the establishment of breathing, and make their sleep state more stable within 2 h after birth. To provide a healthy start for every newborn infant and improve newborn health, implementing high-quality EENC is recommended to policy makers, hospital managers and medical professionals in China.


Birth triggers a large fetal neuroendocrine response, which is more pronounced in infants born vaginally than in those born by elective cesarean section (ECS). The two related peptides arginine vasopressin (AVP) and oxytocin (OT) play an essential role in peripheral and central stress adaptation and have a shared receptor mediating their function. Elevated cord blood levels of AVP and its surrogate marker copeptin, the C-terminal part of AVP prohormone, have been found after vaginal delivery (VD) as compared to ECS, while release of OT in response to birth is controversial. Moreover, AVP, copeptin and OT have not yet been measured simultaneously at birth.To test the hypothesis that AVP but not OT levels are increased in infants arterial umbilical cord blood in response to birth stress and to characterize AVP secretion in direct comparison with plasma copeptin.In a prospective single-center cross-sectional study, we recruited healthy women with a singleton pregnancy and more than 36 completed weeks of gestation delivering via VD or ECS (cesarean without prior uterine contractions or rupture of membranes). Arterial umbilical cord blood samples were collected directly after birth, centrifuged immediately and plasma samples were frozen. Concentrations of AVP and OT were determined by radioimmunoassay and that of copeptin by ultrasensitive immunofluorescence assay.n = 29 from VD and n = 24 from ECS. Ten venous blood samples from pregnant women without stress were collected as controls. AVP and copeptin concentrations were significantly higher in the VD group than in the ECS group (both p < 0.001), median (range) AVP 4.78 (2.38–8.66) vs. 2.38 (1.79–3.88) (pmol/L), copeptin 1692 (72.1–4094) vs. 5.78 (3.14–17.97), respectively, (pmol/L). In contrast, there was no difference in OT concentrations (pmol/L) between VD and ECS, 6.00 (2.71–7.69) vs. 6.14 (4.26–9.93), respectively. AVP and copeptin concentrations were closely related  while OT did not show any correlation to either AVP or copeptin. In linear regression models, vaginal delivery and biochemical stress indicators, base deficit and pH, were independent predictors for both AVP and copeptin. OT was not linked to base deficit or pH.A total of 53 arterial umbilical cord blood samples were collected, Vaginal birth causes a profound secretion of AVP and copeptin in infants. Whereas AVP indicates acute stress events, copeptin provides information on cumulative stress events over a longer period. In contrast, fetal OT is unaffected by birth stress. Thus, AVP signaling but not OT mediates birth stress response in infants. This unique hormonal activation in early life may impact neurobehavioral development in whole life. Childbirth involves a complex hormonal action including the peptides arginine vasopressin (AVP) and oxytocin (OT) to facilitate fetal adaptation from intra- to extra-uterine life, involving cardiovascular and pulmonary adaptation, thermoregulation, metabolic homeostasis and behavior .AVP, also known as antidiuretic hormone, is produced as a larger precursor AVP prohormone in the hypothalamus, cleaved in different peptides, including the bioactive non-apeptide AVP and copeptin, the C-terminal part of AVP prohormone. These are eventually secreted in equimolar amounts by the posterior pituitary gland . The phySimilar to AVP, OT is also synthesized in the hypothalamus and differs in only two amino acids from its evolutionary older sister peptide AVP and exerts its action via the OT receptor (OTR), including patterns of growth, resilience, and healing with stress-coping, anti-inflammatory, and antioxidant functions . HoweverAt the end of pregnancy, the maternal level of oxytocin rises, and a noticeable surge of maternal OT occurs at parturition in the mothers blood . HoweverOn top of their broad peripheral actions, both peptides AVP and OT are well known for their distinct neurobehavioral actions . InteresIn contrast to OT, which ideally is determined in blood by OT itself, AVP concentrations in blood can be deduced from analyzing copeptin concentrations. In fact, copeptin measurement achieved wide clinical utility as copeptin is much more stable than AVP and can be determined quantitatively in an automated fashion . DespiteAVP, its surrogate marker copeptin and OT were measured each in many studies on fetal response to childbirth . There iThe present study was designed to directly compare for the first time the three peptides, AVP, copeptin, and oxytocin, in arterial umbilical cord blood samples in response to birth stress. The two aims of our study were first, to assess whether fetal OT level respond to birth stress by comparing fetal arterial umbilical cord blood OT levels after vaginal delivery (VD) and elective cesarean section (ECS) and, second, to assess AVP and copeptin in parallel.This prospective, cross-sectional study was conducted in a University women’s and children’s hospital in Germany. The study was approved by the Ethics Committee of the University of Regensburg (file number: 20-1876-101) and written consent was obtained prior to enrollment. Inclusion criteria for both birth modes, VD and ECS, were singleton pregnancy and a gestational age of 37-42 weeks. ECS was defined as cesarean section done either on maternal request or due to medical indications, such as breech presentation or repeated cesarean. Hence, no onset of labor, meaning no uterine contractions or rupture of membranes before ECS, was mandatory. In contrast, instrumental VD and secondary cesarean  was an exclusion criterion. Further maternal exclusion criteria were infections, hypertension, preeclampsia, and substance abuse. Fetal exclusion criteria were fetal malformation, chromosomal aberration, fetal growth restriction and infections. Infants requiring advanced medical support and admitted to the neonatal intensive care unit were also excluded from the study.In addition, venous blood samples from 10 healthy pregnant women (37-40 weeks of gestation) at regular pregnancy control visits and before the onset of uterine contractions or rupture of membranes, were used as controls. The inclusion and exclusion criteria for this group were the same as above.Arterial umbilical blood samples (0.5 mL) were collected directly after birth into EDTA tubes and plasma was frozen at −20°C (maximum within 30 min after delivery). All samples were centrifuged by 4°C up to 10 min with a relative centrifugal force of 1,300 g according to a previously published protocol . AVP andMeasurements of copeptin were done by using an ultrasensitive immunofluorescence assay . The labInformation concerning maternal body mass index prior to pregnancy, gestational age at delivery, delivery parameters  and newborn characteristics at birth  were collected.U-test and categorical variables  with the chi-square test. Spearman’s rank-order correlation was applied to assess the correlation among AVP, copeptin, and OT. Univariable linear regression analysis was used to explore the effect of delivery mode, sex, gestational age, birth weight, umbilical blood pH and base deficit (independent variables) on the log-transformed AVP, copeptin, and OT levels (dependent variables). Parameters with P < 0.1 in the univariable analysis were included in multivariable models. Statistical analyses were performed using the IBM SPSS version 25 .Continuous variables (median with IQR) were compared with the Mann-Whitney In total, 29 subjects were included in the VD group and 24 in the ECS group. There were no differences in mothers somatometric data and infants weight, head circumference, sex and postnatal adaptation characteristics between the two groups . In contp < 0.001 for both comparisons). In contrast, OT levels were similar between the two   as compared to the ECS group  .p < 0.001) but not between AVP and OT  or between copeptin and OT . AVP and copeptin were negatively correlated with the pH  and the BE  . There was no correlation between OT and pH or between OT and BE (data not shown).There was a strong correlation between AVP and copeptin  with those of the pregnant women before the onset of labor , all three peptides were significantly higher in infants. More specifically, AVP was 2.93 pmol/L  in infants vs. 2.001 pmol/L (1.74–2.77 pmol/L) in controls (P = 0.042), copeptin 174.4 pmol/L (5.99–1831.5 pmol/L) vs. 7.44 pmol/L (5.52–9.68 pmol/L) (p < 0.001), and OT 6.06 pmol/L (5.59–7.16 pmol/L) vs. 4.00 pmol/L (3.69–4.26 pmol/L) (p < 0.001), respectively.When comparing AVP, copeptin and OT concentrations in all infants (The peptides AVP and OT are key hormones in the peripheral system and in the brain with largely opposite roles in modulating stress, anxiety, and social behaviors . HoweverTo clarify the effect of birth stress on the secretion of these peptides, the present study aimed to assess for the first time simultaneously the levels of AVP, OT, and the AVP surrogate marker copeptin in fetal arterial umbilical cord blood at birth. The main finding was that in contrast to OT, both AVP and copeptin are significantly increased after VD as compared to deliveries without preceding birth stress, namely ECS. Thus, OT secretion in infants is not subject to birth stress.Determination of AVP and OT levels in clinical settings is challenging, as half time of both peptides is short and in particular their stability in sampling matrices is very low . In contSince we strictly followed previously published protocols on sampling and post-sampling handling with immediate cooling down to preserve stability of all peptides , we concIn the present analysis, oxytocin levels were completely unaffected by the mode of delivery and did not show any association to clinical and biochemical stress indicators nor to AVP or copeptin. This confirms previous findings by AVP mRNA levels and AVP protein levels rise significantly at birth (Animal models support the findings of a perinatal surge in AVP and copeptin with higher levels in plasma after a vaginal than a Cesarean section . In paraat birth . Hypoxicat birth . Whereasat birth , centralat birth  and suppat birth . To whatat birth .Strengths of our study include the prospective design, the strict adherence to sampling and post-sampling protocols as well as state of the art measurement of peptides. Limitations include the lack of maternal blood samples in parallel to that from fetuses and the lack of additional measurements within the first hours of life to determine the half-life of AVP and copeptin in the peripheral circulation of neonates.In conclusion, AVP and copeptin reflect both birth stress but whereas AVP indicates acute stress events, copeptin provides information on cumulative stress events over a longer period. In contrast to AVP and copeptin, OT level in infants at birth are not associated with any clinical or biochemical stress indicator, thus, OT is not subject to fetal stress response during birth.The raw data supporting the conclusions of this article will be made available by the authors, without undue reservation.The studies involving human participants were reviewed and approved by the Ethics Committee of the University of Regensburg. Written informed consent to participate in this study was provided by the participants’ legal guardian/next of kin.SFM, EB-Z, LB, and HM performed clinical data acquisition, biomarker analyses, and coordination and check. SF performed statistical analysis and contributed to the manuscript. SFM, CB, and SW supervised the whole project. All authors made substantial contributions to conception and design, analyses and interpretation of data, and revising the article, read the manuscript, and approved its submission.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


How do serum levels of the brain injury biomarkers neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) change after preterm birth, and are these biomarkers associated with neonatal morbidities?In this cohort study of 221 infants born preterm, infants experienced a postnatal increase in NfL that continued for at least 1 month after birth. High serum levels of NfL were associated with the retinal neurovascular disease retinopathy of prematurity (ROP) and, in an exploratory analysis, associated with an unfavorable neurodevelopmental outcome at 2 years of age.These findings suggest that serum levels of the neuronal injury biomarker NfL might be used as an early clinical biomarker of ROP development and neurodevelopmental outcome; however, this should be investigated further. This cohort study examines postnatal longitudinal serum levels of neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) in preterm infants during the first 15 weeks of life and explores possible associations between these biomarkers, neonatal morbidities, and neurodevelopmental outcomes at 2 years. Circulating levels of neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) are important in the course of brain injury in adults, but longitudinal postnatal circulating levels in preterm infants have not been investigated.To examine postnatal longitudinal serum levels of NfL and GFAP in preterm infants during the first 15 weeks of life and to explore possible associations between these biomarkers, neonatal morbidities, and neurodevelopmental outcomes at 2 years.This cohort study used data from 3 clinical studies, including 221 infants born before 32 weeks gestational age (GA) from 1999 to 2015; neurodevelopmental outcomes were evaluated in 120 infants. Data were collected at tertiary-level neonatal intensive care units in Gothenburg, Lund, and Uppsala, Sweden. Data analysis was conducted from January to October 2020.Preterm birth.Serum NfL and GFAP levels, retinopathy of prematurity (ROP), intraventricular hemorrhage, and Bayley Scales of Infant Development II and III at 2 years of age, analyzed by multivariate logistic regression measured by odds ratio (OR), and receiver operating characteristic curve (ROC) analysis. Area under the curve (AUC) was also measured.P < .001). In an exploratory analysis adjusted for GA at birth and sex, NfL AUC at weeks 2 to 4 was independently associated with unfavorable neurodevelopmental outcomes at 2 years corrected age . Longitudinal GFAP levels were not significantly associated with neonatal morbidity or neurodevelopmental outcome.The 221 included infants (108 [48.9%] girls) had a mean (SD) GA at birth of 26.5 (2.1) weeks and a mean (SD) birth weight of 896 (301) grams. NfL levels increased after birth, remaining high during the first 4 weeks of life before declining to continuously low levels by postnatal age 12 weeks . In a binary logistic regression model adjusted for GA at birth, birth weight SD score, Apgar status at 5 minutes, and mode of delivery, the NfL AUC at weeks 2 to 4 was independently associated with any ROP (OR, 4.79; 95% CI, 2.17-10.56; In this study, high NfL levels during the first weeks of life were associated with ROP and poor neurodevelopmental outcomes at 2 years of age. Associations between NfL and later neurovascular development in infants born prematurely should be investigated further. The protein has historically been measured in cerebrospinal fluid (CSF), but methodological advances now allow quantification of NfL in serum using single-molecule array (Simoa) technology, which strongly correlates with levels measured in the CSF.6 The brain-specific GFAP is located in the astroglia and is released after cell injury and death.7 GFAP exhibits clinical prognostic potential in adult brain injury, including traumatic injury and stroke,9 and neurodegenerative disorders.10 Elevated GFAP concentrations in neonates, in both serum and CSF, have been described in association with periventricular white matter injury, brain injury after extracorporeal membrane oxygenation, and in full-term infants with abnormal brain magnetic resonance imaging (MRI) and adverse neurodevelopmental outcome following asphyxia.15Brain injury biomarkers neurofilament light (NfL) and glial fibrillary acidic protein (GFAP) are specific peptides reflecting neuro-axonal injury and astroglia cell injury in the brain. NfL constitutes a part of the neuronal cytoskeleton and shows promise in evaluating acute and chronic severity and the course of neurodegenerative diseases, such as Alzheimer disease, Parkinson disease, and multiple sclerosis,16 Due to rapid maternal and neonatal care improvements, the number of surviving preterm infants is increasing. Preterm infants are at high risk of brain injury with subsequent neurodevelopmental impairment. This is due, in part, to macrostructural brain injury intraventricular hemorrhage (IVH). However, the dominant underlying pathology is white matter damage (WMD) with affected glia accompanied by the neuronal-axonal unit anomalies.17 WMD is associated with reduced brain volumes in preterm infants, linked to gray matter disturbances,19 resulting in poor neurodevelopment.20 Furthermore, prematurity itself is associated with disturbances in both white matter and gray matter.23 In addition, preterm infants are at particular risk of other morbidities affecting the neurovascular unit, such as the prematurity-related disorder retinopathy of prematurity (ROP). ROP is a leading cause of childhood blindness associated with later neurodevelopmental impairment25 and reduced brain volumes,26 although this link is not fully understood. Studies have also highlighted the possibility of using the retina as a proxy for cerebral integrity.27 In unique longitudinal multicenter cohorts of preterm infants, we could describe, for the first time that we are aware of, the longitudinal levels of brain biomarkers NfL and GFAP following preterm birth and investigate the possible association with morbidities in the neurovascular unit as well as the eye and the brain, which are also key organs determining long-term outcome.An estimated 15 million babies are born before 37 weeks gestation annually.31Infants from 3 Swedish cohorts of infants born before 32 weeks’ gestational age (GA) between 1999 and 2015 were included. The cohorts were earlier described separately.NCT02760472). Infants were included after oral and written informed consent from parents or legal guardians. This study followed the Strengthening the Reporting of Observational Studies in Epidemiology (STROBE) reporting guideline.The ethics committees of the respective institutions approved the clinical studies in cohort 1 at the University of Gothenburg and Uppsala University. Approval was obtained from the regional ethical review board in Lund for cohort 2. Cohort 3 was approved by the Regional Ethical Board in Gothenburg and registered at ClinicalTrials.gov  at Queen Silvia Children’s Hospital (n = 70) and Uppsala University Hospital (n = 14) between December 1999 and April 2002. Blood samples were collected on postnatal day (PND) 1 and then once every week until 40 weeks postmenstrual age (PMA).Cohort number 2 included 64 infants born at less than 31 weeks gestation and admitted to the NICU at Lund University Hospital between January 2005 and May 2007. Blood was collected at PND 3, 7, and 14, and then once every week until 40 weeks’ PMA.Cohort number 3 included 90 infants born at less than 28 weeks gestation in the NICU at Queen Silvia Children’s Hospital between April 2013 and September 2015. Cohort 3 was a randomized clinical interventional study in which participants were randomized to different parenteral nutrition strategies regarding lipids, either an omega-3-enriched lipid emulsion or standard care. Blood samples were collected from the umbilical cord and at PND 1, 7, 14, and 28 and at PMA 32, 36, and 40 weeks.32 and Consolidated Standards of Reporting Trials (CONSORT) reporting guideline33 in Cohorts are summarized according to STROBENfL and GFAP were measured simultaneously with all individuals randomized across the included assays using the same reagent batch of SIMOA NF-Light Advantage kit and SIMOA GFAP Discovery Kit, respectively, on the Simoa HD-X platform (Quanterix Corp). Detailed descriptions of the protocol for analytical measurements, interassay and intra-assay variability, calibration limits, and data on the number of samples analyzed are described in the eAppendix in the Longitudinal data on NfL and GFAP were categorized in subgroups based on the PNDs of sampling as follows: at birth, PND 0-3; week 1, PND 4-10; week 2, PND 11-17; week 3, PND 18-24; week 4, PND 25-31; week 5, PND 32-38; week 6, PND 39-45; week 7, PND 46-52; week 8, PND 53-59; week 9, PND 60-66; week 10, PND 67-73; week 11, PND 74-80; week 12, PND 81-87; week 13, PND 88-94; week 14, PND 95-101; and week 15, PND 102-108. Multiple data points during 1 of these periods from a single individual were transformed into a mean data point for the corresponding period. GFAP results are presented in the eAppendix in the Diagnoses of IVH and ROP were retrieved from clinical records. ROP was diagnosed by repeated ophthalmological examinations and defined as non-ROP or ROP . ROP evaluations were initiated according to national guidelines not earlier than at PMA 31 weeks. IVH included any stage of IVH diagnosed by cerebral ultrasound.34 The thresholds used for BSID III closely resembled the suggested cutoff levels for moderate to severe neurodevelopmental delay in BSID III compared with BSID II.36 The categorization of neurodevelopment was based on BSID findings.Neurodevelopmental outcomes were assessed by Bayley Scales of Infant Development (BSID) at 2 years of corrected age in cohorts 2 (n = 48) and 3 (n = 72). Due to updated test routines between study periods, cohort 2 was evaluated by BSID II and cohort 3 by BSID III. Moderate to severe neurodevelopmental impairment in BSID II was defined as either a psychomotor developmental index (PDI) or mental development index (MDI) score of less than 70 and in BSID III as a composite cognitive, composite language, or composite motor scale score below 2 SD compared with healthy full-term infants, as retrieved from the Swedish population-based cohort Express Study.P < .05 was considered significant, unadjusted for multiple testing.The identification of postnatal period of interest for further investigation was based on visual analysis of postnatal development of brain biomarkers. Statistical analyses were performed using SPSS Statistics version 26 (IBM Corp). Nonparametric statistical methods were used because NfL concentrations did not follow the normal distribution. Correlation between GA and cord blood NfL levels were investigated by Spearman correlation coefficient. The correlation between BSID scores and longitudinal NfL levels was also assessed by the Spearman correlation coefficient. To investigate the development of NfL levels after birth, the Mann-Whitney U test was used to compare concentrations at group levels with Bonferroni-Holm correction for multiple testing, and the Wilcoxon signed-rank test was used to investigate the difference in concentrations from birth  to PND 1 for infants with repeated samples. In all analyses, a 2-sided 37 biweekly, based on NfL concentrations or linear interpolation if the value at the first or last point was not obtainable and at least 2 adjacent values were available. Cognitive and motor outcomes were used as the dependent variables in binary regression models to investigate the association between NfL AUC at weeks 2 to 4 with the neurodevelopmental outcome.A measure of longitudinal NfL concentrations was calculated using the area under the curve (AUC). AUC was calculated accordingly the trapezoidal rule, as described by Fekedulegn et al,P < .20 for each morbidity based on the Mann-Whitney U test were considered in the models, with birth characteristics, longitudinal NfL levels, and other morbidities. Included variables were entered stepwise using the forward method. Variables significantly contributing to the models were included in the final logistic regression model with clinically relevant variables. For neurodevelopmental outcomes, the model was adjusted for GA at birth and sex.Binary logistic regression models based on significant variables were used to calculate the probability values for morbidities as odds ratios (ORs) reported with 95% CIs. In models with ROP as the outcome, the natural logarithm of NfL AUC (ln NfL AUC) was studied because of its stronger association with the outcome than that of NfL AUC (based on lower Akaike information criterion). Variables with an association of The models were validated for linearity between the included variables and ln-odds, the independence between observations, multicollinearity, and outliers. The models did not yield any severe diagnostics when investigated for multicollinearity (variance inflation factor between 1 and 2). Goodness-of-fit for the logistic regression models was tested by the Hosmer-Lemeshow test and was found satisfactory.Receiver operating characteristic (ROC) curves were used to evaluate the statistical associations of disease and possible cutoff values with the highest explanatory contributions to morbidities. The cutoff was determined from a table with all possible cutoff values for the coordinate with the highest sensitivity and specificity. Based on selected cutoffs, positive predictive values (PPVs) and negative predicted values (NPVs) were computed and reported with 95% CIs. The orthogonal projections to latent structures (OPLS) model is described in the eAppendix in the A total of 221 infants with a mean (SD) GA at birth of 26.5 (2.1) weeks and a mean (SD) birth weight (BW) of 896.5 (301.4) grams were included in the study population. The clinical characteristics of the included infants are shown in P = .009) (P < .001) (P < .001).Concentrations of serum NfL at birth (cord blood) and PND 1 were compared in a subpopulation (cohort 3). NfL cord blood levels were negatively correlated with GA  A. The me < .001) B. Among Longitudinal serum levels of NfL from birth to 15 weeks’ PMA are illustrated in P < .001; among 92 infants with vs 62 infants without ROP: 124.9 [23.3-832.2] ng/L vs 43.3 [12.2-368.0] ng/L; P < .001).The biweekly period with the strongest association with IVH and ROP as well as a high number of available AUC values (n = 155) was weeks 2 to 4, and it was used for further analyses. Unadjusted, the NfL AUC at weeks 2 to 4 was higher in infants with IVH or ROP compared with infants without the respective morbidity .Longitudinal levels of NfL in infants with or without ROP according to GA at birth are shown in P < .001)  at 5 minutes. The OR for ln NfL AUC at weeks 2 to 4 for 150 infants was 4.79 . The explanatory contribution to BSID outcome was excellent  GA at birth of 25.9 (1.7) weeks. Among 81 infants with available NfL AUC, the NfL AUC at weeks 2 to 4 was independently associated with moderate to severe neurodevelopmental impairment at 2 years, adjusted for GA at birth and sex .We observed an association between NfL during the first PN weeks and long-term neurodevelopmental outcomes at 2 years, as determined by an unfavorable BSID score. This association was independent of GA at birth, previously established as a factor associated with neurocognitive outcome. A reduced BSID score at 2 years of age is associated with later adverse neurodevelopmental outcomes.The discovery and validation of neonatal biomarkers of brain injury is a critical step in the evolution of neonatal neuroprotection. The possibility of detecting and monitoring the effects of axonal damage and brain development in peripheral blood samples would be of tremendous benefit in clinical decision-making in neonatal intensive care.51This study has limitations, including its retrospective study design. In addition, the prolonged recruitment period may be a possible shortcoming, given that there have been significant developments of new clinical practices. Thus, different tests  were used when assessing neurodevelopmental outcomes in cohorts 2 and 3. BSID III has been shown to result in higher scale points compared with BSID II, which has been the reason for attempts at conversion factors and different thresholds when combining the 2 tests.53 Nevertheless, the storage time is an uncertain factor that might affect the quality of the samples and may affect the measurements’ results. The fact that measurements were made on singleton samples due to the limited sample volumes increases the uncertainty in the results.Another limitation is the prolonged time between sampling and biomarker measurement. NfL has been reported to be stable for as many as 5 freeze-thaw cycles and delayed freezing after blood sampling for as long as 24 hours.In this study, preterm birth was associated with persistently increased serum levels of the brain injury biomarker NfL, corresponding to levels observed after adult brain injury. Furthermore, early postnatal increases in NfL levels were strongly associated with the neurovascular disease ROP and long-term neurodevelopmental outcomes at 2 years of age, possibly illustrating WMD with associated axonal injury. The discovery and validation of a neonatal biomarker for brain injury are crucial in neonatal neuroprotection. The promising role of NfL as a biomarker for ROP and the development of the neurovascular unit should be investigated further.


Cord blood leptin and adiponectin are adipokines known to be associated with birth weight and overall infant adiposity. However, few studies have investigated their associations with abdominal adiposity in neonates. We examined maternal factors associated with cord blood leptin and adiponectin, and the association of these adipokines with neonatal adiposity and abdominal fat distribution measured by magnetic resonance imaging (MRI) in an Asian mother–offspring cohort.Growing Up in Singapore Towards healthy Outcomes (GUSTO), is a prospective mother–offspring birth cohort study in Singapore. Cord blood plasma leptin and adiponectin concentrations were measured using Luminex and Enzyme-Linked Immunosorbent Assay respectively in 816 infants. A total of 271 neonates underwent MRI within the first 2-weeks after delivery. Abdominal superficial (sSAT), deep subcutaneous (dSAT), and intra-abdominal (IAT) adipose tissue compartment volumes were quantified from MRI images. Multivariable regression analyses were performed.Indian or Malay ethnicity, female sex, and gestational age were positively associated with cord blood leptin and adiponectin concentrations. Maternal gestational diabetes (GDM) positively associated with cord blood leptin concentrations but inversely associated with cord blood adiponectin concentrations. Maternal pre-pregnancy body mass index (BMI) showed a positive relationship with cord blood leptin but not with adiponectin concentrations. Each SD increase in cord blood leptin was associated with higher neonatal sSAT, dSAT and IAT; differences in SD (95% CI): 0.258 , 0.386  and 0.250 , respectively. Similarly, each SD increase in cord blood adiponectin was associated with higher neonatal sSAT and dSAT; differences in SD (95% CI): 0.185  and 0.173 , respectively. The association between cord blood adiponectin and neonatal adiposity was observed in neonates of obese mothers only.Cord blood leptin and adiponectin concentrations were associated with ethnicity, maternal BMI and GDM, sex and gestational age. Both adipokines showed positive association with neonatal abdominal adiposity. Adipokines are hormones secreted by adipocytes and suggested to play important roles in energy homeostasis and cell metabolism . Leptin,Adiponectin, one of the adipokines produced by adipocytes, mediates communication of the adipose tissue with other metabolic tissues such as the liver and skeletal muscle . AdiponeIncreased abdominal adiposity (AA) is a known independent risk factor for adverse cardio-metabolic outcomes in adults . IncreasBoth cord blood leptin and adiponectin have been reported to be positively associated with birth weight and overall adiposity at birth measured mostly by simple anthropometry such as BMI and skinfold thickness , 25, 28.The study was based on mother–offspring pairs from the Growing Up in Singapore Towards healthy Outcomes (GUSTO) study, a prospective cohort study in Singapore . PregnanDemographic data, lifestyle, obstetric and medical history were collected at multiple study visits using interviewer administered questionnaires and from hospital records. Self-reported pre-pregnancy weights of mothers were recorded. Pre-pregnancy body mass index (ppBMI) was calculated from the self-reported pre-pregnancy weight and measured height at booking. ppBMI categories were defined using WHO recommended BMI cutoff points for Asians . PregnanGUSTO = −0.022 + (0.307 × weight) − (0.077 × gender) + (0.028 × subscapular skinfolds) − , where gender = 1 for male, 0 for female) was used to calculate fat mass [® Infant Body Composition System Version 3.1.0  was used to measure body composition, i.e., fat mass and fat free mass [® measurement, and those that had <2.5 kg birthweight, <5% percent body fat in PEA POD® assessment [® – adipokine association. Clothing was removed from the infant and the infant was placed on the scale for body mass measurement and inside the chamber for body volume measurement. Percent body fat was computed by the PEA POD system. Age and sex-specific densities of free fat mass based on multi-compartment studies are used by the PEA POD® [Information on birth weight, birth length, and head circumference were obtained from medical records. Triceps and subscapular skinfolds were measured on the right side in triplicates to the nearest 0.2 mm using Holtain skinfold calipers . The validated GUSTO equation for estimating fat mass  of a pooled plasma control was 17.3%. Umbilical cord blood total adiponectin was measured in EDTA plasma from venous cord blood using the Adiponectin Human in vitro ELISA (Enzyme-Linked Immunosorbent Assay) kit (Abcam). The CV of a pooled plasma control was 13.3%. Adjustment for plate effect was performed using median centering where the median of each plate was shifted to the global median for all plates.333 healthy neonates born ≥34 weeks gestation with birth weight (BW) ≥ 2000g had abdominal adipose tissue compartment (AAT) volumes data available from MRI scans performed within two weeks post-birth. A detailed participant selection flow chart can be accessed from Tint MT et al. . Of thes® subsets using independent sample T-tests for continuous variables and chi square tests for categorical variables. Cord blood leptin and adiponectin concentrations were not normally distributed thus Mann–Whitney U tests were used for group comparison.The characteristics of mothers and offspring were compared among the whole cohort vs. MRI and PEAPODMultivariable regression analyses were performed to determine maternal and neonatal factors associated with cord blood adipokines. Outcome variables i.e., adipokine concentrations were transformed into standardized scores so that the strength of associations were comparable in regression model. Models were mutually adjusted for ethnicity, maternal education, tobacco exposure, parity, maternal age, maternal pre-pregnancy BMI, GWG categories, GDM status, gestational age at delivery and child’s sex.P values were corrected using Benjamini–Hockberg method with false discovery rate of 0.05(25) [Multivariable regression analyses were performed with cord blood adipokines (leptin and adiponectin) as the main exposures, birth weight, skinfold thickness measurement as total adiposity measure and AAT compartment volumes as outcomes of interest. All exposures and outcome variables were transformed into standardized scores so that observed strength of associations were comparable. Covariates were controlled for based on prior knowledge from the literature about factors that might confound the associations between maternal and cord adipokines and neonatal adiposity. Models were adjusted for ethnicity, pre-pregnancy BMI, GDM status, gestational age at delivery, and child’s sex and birth length. When studying neonatal adiposity by MRI, models were additionally adjusted for age on MRI day. 0.05(25) . All star = 0.103, p = 0.003).A flow chart of this study is shown in Supplementary Fig. Factors associated with cord blood leptin and adiponectin are shown in Table Table Similarly, each SD increase in cord blood adiponectin was associated with a 0.110  SD increase in birth weight and a 0.136  SD increase in fat mass by PEAPOD in neonates. Each SD increase in cord blood adiponectin was associated with 0.185 , 0.173  and 0.092  SD increases in neonatal sSAT, dSAT and IAT, respectively.2 showed statistically significant positive association between cord adiponectin concentrations and birthweight as well as measures of neonatal adiposity. The interaction term was significant (p < 0.05) for the interaction between obese maternal BMI category and the cord adiponectin concentrations in the association with predicted fat mass and sSAT  covering around 50% of the global population. While most previous studies used total adiposity of offspring in association with adipokine concentrations, we used AAT quantified from MRI, an accurate method without radiation to quantify AA, which has been shown to be related to metabolic health. The timing of MRI scans for neonates was within 2-weeks after delivery thus the observation would largely reflect the developmental influences on the offspring before any postnatal environmental exposures.Our study has some limitations. One is that only a subset of eligible neonates whose parents gave consent for their MRI were included in the study on AA, so care therefore should be taken not to generalize our findings. Future studies are warranted to confirm these findings in larger study populations. Furthermore, as with any other observational studies, we cannot completely exclude bias due to residual confounding although major potential confounders were adjusted for in the study.In summary, we show that  fetal adSupplementary data file