Continuous age- and sex-specific reference ranges of liver enzymes in Chinese children and application in pediatric non-alcoholic fatty liver disease.

BACKGROUND: Alanine aminotransferase (ALT) is widely used to screen patients with hepatic diseases. However, the current reference ranges (< 50 U/L) were developed by laboratories and have not been validated in populations with a large number of healthy individuals. METHODS: This study collected venous blood and anthropometric data from a total of 13,287 healthy children aged 3 months to 18 years who underwent routine physical examinations in the Department of Pediatric Healthcare. We applied the least mean square algorithm to establish age- and sex-related reference percentiles of serum levels of transaminases. For validation, we recruited 4276 children and adolescents with obesity/overweight who underwent evaluation and metabolic tests in the hospital. Using receiver operating characteristic curves, we determined age- and sex-specific upper limit percentiles of liver enzymes for fatty liver diseases. RESULTS: This study revealed a significant correlation between serum transaminase levels and age and sex (P < 0.01). These transaminase levels exhibited age- and sex-specific patterns. Among individuals in the non-alcoholic fatty liver disease (NAFLD) cohort, elevated ALT levels displayed a positive association with clinical markers of disease severity, including homeostatic model assessment of insulin resistance, waist-hip ratio, and serum uric acid levels (P < 0.01). According to the receiver operating characteristic curves, ALT levels at the 92.58th percentile for boys and the 92.07th percentile for girls yielded the highest accuracy and specificity. CONCLUSIONS: This study provides age- and sex-specific reference ranges for ALT, aspartate aminotransferase, and γ-glutamyltransferase in Chinese children and adolescents, making it the largest population study to date. Furthermore, the study establishes a precise upper limit for ALT levels, facilitating their use in NAFLD screening. Video Abstract.

Gender differences in non-alcoholic fatty liver disease in obese children and adolescents: a large cross-sectional study.

OBJECTIVE: Previous studies have reported sex differences in non-alcoholic fatty liver disease (NAFLD) among adults; however, little is known about its occurrence in children and adolescents. This study aims to examine the prevalence of NAFLD among them and investigate the relationship between sex hormones and NAFLD. METHOD: This study included 2999 obese Chinese children aged 2-18 years. We examined the prevalence of NAFLD by sex, age, and Tanner stage. The regression model and principal component analysis were used to analyze the relationship between sex hormones and NAFLD. RESULTS: The prevalence of NAFLD increased with age in both sexes, and the gender difference appeared before puberty. The prevalence in boys tended to stabilize at the age of 11 years, whereas girls reached their peak temporarily. NAFLD prevalence was positively associated with estradiol in boys (p = 0.011), but the opposite trend was observed in girls (p = 0.031). Testosterone levels decreased with the increase of NAFLD prevalence in boys (p < 0.001). Luteinizing hormone and prolactin were inversely associated with NAFLD prevalence in boys and girls, respectively. Results from the principal component analysis showed that sex hormone levels and fat distribution were important risk factors for the prevalence of NAFLD in obese children (p < 0.001). CONCLUSION: The significant difference in NAFLD prevalence between genders in obese children begins in early childhood. This distinction emerges long before puberty onset and tends to stabilize during late adolescence. Sex hormones are associated with NAFLD prevalence and are influenced by the Tanner stages and fat distribution.

The relative contributions of soft tissue mass components as risk or protective factors of non-alcoholic fatty liver disease in children.

BACKGROUND/OBJECTIVE: Several body components are known to be associated with non-alcoholic fatty liver disease (NAFLD) in children. However, the relative contributions of soft tissue mass components as risk or protective factors of NAFLD are largely unknown because measurements of these components are often highly correlated. Therefore, we aimed to estimate levels of association between soft tissue mass components and NAFLD. SUBJECTS/METHODS: We collected the medical records of 555 Chinese children (aged 3-18 years). Five mutually exclusive and exhaustive components of soft tissue mass were measured using dual energy X-ray absorptiometry. NAFLD was diagnosed with abdominal B-ultrasound scan. We fit Dirichlet regression and multivariate linear regression models wherein age and NAFLD were used as predictors of the proportional measurements of soft tissue mass components. RESULTS: The proportion of android fat was significantly higher in children with NAFLD than in those without NAFLD (ratio of proportions ranged from 1.18 to 1.30), whereas proportions of trunk lean and limb lean were significantly lower (ratio of proportions ranged from 0.87 to 0.92 for trunk lean and from 0.82 to 0.91 for limb lean). The proportion of gynoid fat was slightly higher in boys with NAFLD than in those without NAFLD (ratio = 1.05), but this proportion was not significantly higher in girls. The association between the proportion of android fat and NAFLD appeared to be somewhat greater than the associations between proportions of trunk lean or limb lean components and NAFLD. CONCLUSION: Our findings suggest that lowering fat mass and increasing lean mass can both be used to combat NAFLD in children and that more studies are needed to determine the association between gynoid fat and NAFLD.

Cytoskeleton remodeling mediated by circRNA-YBX1 phase separation suppresses the metastasis of liver cancer.

Metastasis, especially intrahepatic, is a major challenge for hepatocellular carcinoma (HCC) treatment. Cytoskeleton remodeling has been identified as a vital process mediating intrahepatic spreading. Previously, we reported that HCC tumor adhesion and invasion were modulated by circular RNA (circRNA), which has emerged as an important regulator of various cellular processes and has been implicated in cancer progression. Here, we uncovered a nuclear circRNA, circASH2, which is preferentially lost in HCC tissues and inhibits HCC metastasis by altering tumor cytoskeleton structure. Tropomyosin 4 (TPM4), a critical binding protein of actin, turned out to be the major target of circASH2 and was posttranscriptionally suppressed. Such regulation is based on messenger RNA (mRNA)/precursormRNA splicing and degradation process. Furthermore, liquid-liquid phase separation of nuclear Y-box binding protein 1 (YBX1) enhanced by circASH2 augments TPM4 transcripts decay. Together, our data have revealed a tumor-suppressive circRNA and, more importantly, uncovered a fine regulation mechanism for HCC progression.

The potential of mecciRNA in hepatic stellate cell to regulate progression of nonalcoholic hepatitis.

BACKGROUND: Nonalcoholic steatohepatitis (NASH) occupies a substantial proportion of chronic liver disease worldwide, of which pathogenesis needs further research. Recent studies have demonstrated the significant roles of circular RNAs (circRNAs) in NASH, while the function of a novel type of circRNAs, namely mitochondria-encoded circRNAs (mecciRNAs), remains elusive. Therefore, we aimed to investigate their potential to regulate the progression of NASH in this study. METHODS: GSE134146 was used to screen for differentially expressed mecciRNAs in NASH, while GSE46300 was used to identify NASH-related genes. To establish the mecciRNA-miRNA-mRNA networks, circMINE and miRNet databases were used for predicting downstream targets. Then, consensus clustering analysis was used to determine immune subtypes of NASH. Finally, we successfully validated our findings in vitro (LPS-treated hepatic stellate cells [HSCs]) and in vivo (MCD-diet mice) NASH models. RESULTS: We confirmed that circRNomics balance is disrupted in HSCs of NASH, while two mecciRNAs (hsa_circ_0089761 and hsa_circ_0089763) could function as competing for endogenous RNAs (ceRNAs) to regulate fibrosis-related signals. Furthermore, we constructed two ceRNA networks based on mecciRNAs for the first time. Cell and animal NASH models validated our findings that c-MYC and SMAD2/3 were upregulated in HSCs, while THBS1 and p-STAT3 were upregulated in hepatocytes. Moreover, we identified 21 core genes by overlapping the differentially expressed genes (NASH vs. Normal) with mecciRNA-targeted genes. According to their expression profiles, NASH patients could be divided in 2 different clusters, in which proinflammatory signals (TNF and IL-17 pathways) are significantly activated in Cluster 1. CONCLUSION: We successfully established two novel mecciRNA-miRNA-mRNA networks in HSCs and hepatocytes, which were further confirmed by in vitro and in vivo models. Meanwhile, the novel immunotyping model revealed the heterogeneity of NASH, thereby might guiding treatment options. Altogether, our study brought a distinct perspective on the relationship between mecciRNAs and NASH.

High Prevalence of Cardiometabolic Comorbidities Among Children and Adolescents With Severe Obesity From a Large Metropolitan Centre (Hangzhou, China).

Objective: This study aimed to describe the clinical characteristics of children and adolescents with obesity, and the prevalence of cardiometabolic comorbidities over 10 years in this population from a large metropolitan centre in China. Methods: This was a cross-sectional study (2008-2017) of patients aged <18 years with obesity [body mass index (BMI) ≥ 95th percentile for age and sex] enrolled at the Department of Endocrinology, Children's Hospital of Zhejiang University School of Medicine (Hangzhou, Zhejiang Province). Clinical assessments included anthropometry, blood pressure, liver ultrasound, lipid profile, oral glucose tolerance test, and uric acid. For examination of outcomes, our study cohort was stratified by sex and age bands (<10 vs. ≥10 years), with the study period also split into two strata (2008-2012 and 2013-2017). Results: A total of 2,916 patients (1,954 boys and 962 girls) were assessed at a mean age of 10.5 years. Patients almost invariably presented severe obesity (median BMI SDS = 2.98; Q1 = 2.60, Q3 = 3.39). Obesity-related comorbidities were common among boys and girls, including type 2 diabetes mellitus (2.6% and 3.6%, respectively), abnormal glycaemia (33.6% and 35.5%, respectively), hypertension (33.9% and 32.0%, respectively), dyslipidaemia (35.2% and 39.6%, respectively), hyperuricaemia (16.2% and 8.3%, respectively), acanthosis nigricans (71.9% and 64.0%, respectively), abnormal liver function (66.9% and 47.0%, respectively), and non-alcoholic fatty liver disease (NAFLD) (63.8% and 45.1%, respectively); 38.7% of boys and 44.4% of girls aged ≥10 years had metabolic syndrome. Notably, the incidence of many cardiometabolic comorbidities was in 2013-2017 compared to 2008-2012. For example, rates of hypertension among boys aged <10 years and aged ≥10 years rose from 28.4% and 26.5% to 48.0% and 35.8%, respectively, and in girls from 20.3% and 20.8% to 41.7% and 39.6%, respectively. In 2013-2017, 9.5% of girls in the older group had metabolic syndrome compared to 2.2% in 2008-2013. Conclusions: We observed a high incidence of obesity-related cardiometabolic comorbidities among Chinese children and adolescents with severe obesity over 10 years. It was particularly concerning that rates of several comorbidities rose markedly over the study period, highlighting the need to address the obesity epidemic early in life (in China and elsewhere) to prevent the development of obesity-related comorbidities and, subsequently, of overt disease.

10-Year Incidence of Diabetic Ketoacidosis at Type 1 Diabetes Diagnosis in Children Aged Less Than 16 Years From a Large Regional Center (Hangzhou, China).

Background: Diabetic ketoacidosis (DKA) is a potentially life-threatening complication of type 1 diabetes (T1D), and a leading cause of death in children aged <15 years with new-onset T1D. Aims: i) to assess the incidence of DKA in children and adolescents newly diagnosed with T1D over a 10-year period at a large regional center in China; and ii) to examine the clinical symptoms and demographic factors associated with DKA and its severity at diagnosis. Methods: We carried out a retrospective audit of a regional center, encompassing all youth aged <16 years diagnosed with T1D in 2009-2018 at the Children's Hospital, Zhejiang University School of Medicine (Hangzhou, China). DKA and its severity were classified according to ISPAD 2018 guidelines. Results: 681 children were diagnosed with T1D, 50.1% having DKA at presentation (36.0% mild, 30.0% moderate, and 33.9% severe DKA). The number of patients diagnosed with T1D progressively rose from approximately 39 cases/year in 2009-2010 to 95 cases/year in 2017-2018 (≈2.5-fold increase), rising primarily among children aged 5-9 years. DKA incidence was unchanged but variable (44.8% to 56.8%). At T1D diagnosis, 89% of patients reported polyuria and 91% polydipsia. Children presenting with DKA were more likely to report vomiting, abdominal pain, and particularly fatigue. DKA was most common among the youngest children, affecting 4 in 5 children aged <2 years (81.4%), in comparison to 53.3%, 42.7%, and 49.3% of patients aged 2-4, 5-9, and ≥10 years, respectively. Children with severe DKA were more likely to report vomiting, fatigue, and abdominal pain, but less likely to report polyuria, polydipsia, and polyphagia than those with mild/moderate DKA. Rates of severe DKA were highest in children aged <2 years (51.1%). Conclusions: The number of children diagnosed with T1D at our regional center increased over the study period, but DKA rates were unchanged. With 9 of 10 children reporting polyuria and polydipsia prior to T1D diagnosis, increasing awareness of this condition in the community and among primary care physicians could lead to earlier diagnosis, and thus potentially reduce rates of DKA at presentation.

Abdominal Adiposity and Total Body Fat as Predictors of Cardiometabolic Health in Children and Adolescents With Obesity.

Objective: We aimed to assess the role of adipose tissue distribution in cardiometabolic risk (in particular insulin sensitivity) in a population of children and adolescents with obesity. Methods: In this cross-sectional study, participants were 479 children and adolescents with obesity (322 boys and 157 girls) aged 3 to 18 years attending the Children's Hospital at Zhejiang University School of Medicine (Hangzhou, China). Clinical assessments included anthropometry, body composition (DXA scans), carotid artery ultrasounds, and OGTT. Insulin sensitivity was assessed using the Matsuda index. Participants were stratified into groups by sex and pubertal stage. Key predictors were DXA-derived android-to-gynoid-fat ratio (A/G) and total body fat percentage (TBF%). Results: Irrespective of sex and pubertal stage, there was a strong association between increasing A/G (i.e., greater abdominal adiposity) and lower insulin sensitivity. In multivariable models, every 0.1 increase in A/G was associated with a reduction in insulin sensitivity in prepubertal boys [-29% (95% CI -36%, -20%); p < 0.0001], pubertal boys [-13% (95% CI -21%, -6%); p = 0.001], and pubertal girls [-16% (95% CI -24%, -6%); p = 0.002]. In contrast, TBF% was not associated with insulin sensitivity when A/G was adjusted for, irrespective of pubertal stage or sex. In addition, every 0.1 increase in A/G was associated with increased likelihood of dyslipidemia in prepubertal boys [adjusted odds ratio (aOR) 1.62 (95% CI 1.05, 2.49)], impaired glucose tolerance in pubertal boys [aOR 1.64 (95% CI 1.07, 2.51)] and pubertal girls [aOR 1.81 (95% CI 1.10, 2.98)], and odds of NAFLD in both prepubertal [aOR 2.57 (95% CI 1.56, 4.21)] and pubertal [aOR 1.69 (95% CI 1.18, 2.40)] boys. In contrast, higher TBF% was only associated with higher fasting insulin and ALT in pubertal boys, being also predictive of NAFLD in this group [aOR 1.15 per percentage point (95% CI 1.06, 1.26)], but was not associated with the likelihood of other cardiometabolic outcomes assessed in any group. Conclusions: A/G is a much stronger independent predictor of cardiometabolic risk factors in children and adolescents with obesity in China, particularly glucose metabolism.