Analysis of potential biomarkers and immune infiltration in autism based on bioinformatics analysis.

Autism spectrum disorder (ASD) is a complex neurodevelopmental disorder caused by both environmental and genetic factors. However, its etiology and pathogenesis remain unclear. The purpose of this study was to establish an immune-related diagnostic model for ASD using bioinformatics methods and to identify ASD biomarkers. Two ASD datasets, GSE18123 and GSE29691, were integrated into the gene expression Database to eliminate batch effects. 41 differentially expressed genes were identified by microarray data linear model (limma package). Based on the results of the immune infiltration analysis, we speculated that neutrophils, B cells naive, CD8+ T cells, and Tregs are potential core immune cells in ASD and participate in the occurrence of ASD. Finally, the differential genes and immune infiltration in ASD and non-ASD patients were compared, and the most relevant genes were selected to construct the first immune correlation prediction model of ASD. After the calculation, the model exhibited better accuracy. The calculations show that the model has good accuracy.

Machine learning predictive models for grading bronchopulmonary dysplasia: umbilical cord blood IL-6 as a biomarker.

Objectives: This study aimed to analyze the predictive value of umbilical cord blood Interleukin-6 (UCB IL-6) for the severity-graded BPD and to establish machine learning (ML) predictive models in a Chinese population based on the 2019 NRN evidence-based guidelines. Methods: In this retrospective analysis, we included infants born with gestational age <32 weeks, who underwent UCB IL-6 testing within 24 h of admission to our NICU between 2020 and 2022. We collected their medical information encompassing the maternal, perinatal, and early neonatal phases. Furthermore, we classified the grade of BPD according to the 2019 NRN evidence-based guidelines. The correlation between UCB IL-6 and the grades of BPD was analyzed. Univariate analysis and ordinal logistic regression were employed to identify risk factors, followed by the development of ML predictive models based on XGBoost, CatBoost, LightGBM, and Random Forest. The AUROC was used to evaluate the diagnostic value of each model. Besides, we generated feature importance distribution plots based on SHAP values to emphasize the significance of UCB IL-6 in the models. Results: The study ultimately enrolled 414 preterm infants, with No BPD group (n = 309), Grade 1 BPD group (n = 73), and Grade 2-3 BPD group (n = 32). The levels of UCB IL-6 increased with the grades of BPD. UCB IL-6 demonstrated clinical significance in predicting various grades of BPD, particularly in distinguishing Grade 2-3 BPD patients, with an AUROC of 0.815 (95% CI: 0.753-0.877). All four ML models, XGBoost, CatBoost, LightGBM, and Random Forest, exhibited Micro-average AUROC values of 0.841, 0.870, 0.851, and 0.878, respectively. Notably, UCB IL-6 consistently appeared as the most prominent feature across the feature importance distribution plots in all four models. Conclusion: UCB IL-6 significantly contributes to predicting severity-graded BPD, especially in grade 2-3 BPD. Through the development of four ML predictive models, we highlighted UCB IL-6's importance.

[Association between platelet-activating factor acetylhydrolase gene polymorphism and intracranial hemorrhage in preterm infants].

OBJECTIVE: To explore whether Val279Phe single nucleotide polymorphisms (SNPs) in the 9th exon of platelet-activating factor acetylhydrolase (PAF-AH) are associated with intracranial hemorrhage in preterm infants. METHODS: A case-control study was performed. Polymerase chain reaction (PCR) was used to test genotype and allele frequencies of the 9th exon Val279Phe SNPs of PAF-AH in 58 preterm infants with intracranial hemorrhage (hemorrhage group) and 65 preterm infants without intracranial hemorrhage (control group). RESULTS: There were significant differences in genotype frequency of Val279Phe SNPs in the 9th exon of PAF-AH between the hemorrhage and control groups (P<0.05). Frequency of normal genotype in the hemorrhage group (63.8%) was significantly lower than in the control group (81.5%). In contrast, frequency of heterozygous genotype (34.5%) in the hemorrhage group was significantly higher than in control group (16.9%). There were also significant differences in allele frequency of Val279Phe SNPs in the 9th exon of PAF-AH between the two groups (P<0.05). T allele frequency in the hemorrhage group (19.0%) was significantly higher than in the control group (10.0%). CONCLUSIONS: Val279Phe SNPs in the 9th exon of PAF-AH may be associated with intracranial hemorrhage in preterm infants.

Establishment of a predictive model for purulent meningitis in preterm infants.

Background: Purulent meningitis (PM) is an important cause of mortality and morbidity in the newborn population throughout the world. The subtle of specific clinical signs and low success rates of lumbar puncture make diagnosis of PM more difficult in preterm than in older children. The objective of this study was to establish a predict model for preterm PM in hopes of helping clinicians develop new diagnostic and treatment strategies. Methods: Premature infants who were admitted to The First Affiliated Hospital of Zhengzhou University from September 2017 to March 2020 were enrolled in this study. All the patients underwent lumbar puncture. We collected data encompassing maternal diseases and neonatal clinical features. Cerebrospinal fluid (CSF) culture is the gold standard for diagnosing meningitis. The PM was diagnosed according to the diagnostic criteria. All statistical analyses were performed using R 3.63 (https://www.r-project.org/). Logistic regression and least absolute shrinkage and selection operator (LASSO) regression analyses were used to establish a risk prediction model of PM. The Brier score, calibration slope, and concordance (C)-index were used to verify the accuracy of prediction model. Results: A total of 168 preterm infants were enrolled in this study, 80 boys and 88 girls, the gestational age (GA) was 26.43-36.86 weeks (32.45±2.79 weeks), the birth weight (BW) was 700-3,400 g (1,814.05±568.84 g). There were 77 preterm infants with PM while 91 without. We identified seven variables as independent risk factors for PM in preterm infants by LASSO analysis [the optimal λ was 0.080960, and log(λ) = -2.5138], including procalcitonin (PCT) on the 1st day after birth, prenatal glucocorticoid use, albumin, the 1-minute Apgar score, the use of non-invasive biphasic positive airway pressure, hemoglobin, and sex. These were used to construct a risk prediction nomogram and verified its accuracy. The Brier score was 0.17, the calibration slope was 0.966, and the concordance index was 0.82018. Conclusions: Our prediction model could predict the risk of PM in preterm infants. Using this prediction model, it may be able to provide reference to determine whether lumbar puncture is performed and whether antibiotics are applied as soon as possible.

The Relationship Between Cord Blood Cytokine Levels and Perinatal Characteristics and Bronchopulmonary Dysplasia: A Case-Control Study.

Objective: To establish the association between serial levels of inflammatory cytokines in cord blood and perinatal characteristics and bronchopulmonary dysplasia (BPD) in preterm infants. Methods: 147 premature infants with gestational age ≤32 weeks who were born and hospitalized in the First Affiliated Hospital of Zhengzhou University between July 2019 and August 2021 were enrolled in this retrospective case-control study. Multiple microsphere flow immunofluorescence was used to detect seven cytokines in cord blood collected within 24 h of birth. Demographics, delivery characteristics, maternal factors, neonatal characteristics, and clinical outcomes were collected for the two groups. An unconditional logistic regression model was used in this study to assess the clinical variables. Results: IL-6 cord blood levels at birth were significantly higher in the BPD group than in the non-BPD group, but the odds ratio (OR) was very small (OR = 1). No differences in other cytokine concentrations were observed between the two groups. Multivariable logistic regression analysis demonstrated that increased maternal white blood cell (WBC) count on admission and lower birth weight increased the risk of BPD progression. Conclusions: Increased IL-6 cord blood levels at birth in preterm infants may have trivial significance for predicting BPD. Furthermore, higher maternal WBC count on admission and lower birth weight increased the risk of BPD.

Development and Validation of a Nomogram for Predicting Bronchopulmonary Dysplasia in Very-Low-Birth-Weight Infants.

Background: Bronchopulmonary dysplasia is a common pulmonary disease in newborns and is one of the main causes of death. The aim of this study was to build a new simple-to-use nomogram to screen high-risk populations. Methods: In this single-center retrospective study performed from January 2017 to December 2020, we reviewed data on very-low-birth-weight infants whose gestational ages were below 32 weeks. LASSO regression was used to select variables for the risk model. Then, we used multivariable logistic regression to build the prediction model incorporating these selected features. Discrimination was assessed by the C-index, and and calibration of the model was assessed by and calibration curve and the Hosmer-Lemeshow test. Results: The LASSO regression identified gestational age, duration of ventilation and serum NT-proBNP in the 1st week as significant predictors of BPD. The nomogram-illustrated model showed good discrimination and calibration. The C-index was 0.853 (95% CI: 0.851-0.854) in the training set and 0.855 (95% CI: 0.77-0.94) in the validation set. The calibration curve and Hosmer-Lemeshow test results showed good calibration between the predictions of the nomogram and the actual observations. Conclusion: We demonstrated a simple-to-use nomogram for predicting BPD in the early stage. It may help clinicians recognize high-risk populations.

A Good Way to Reduce Screening for Retinopathy of Prematurity: Development of the ROP Model in a China Preterm Population.

Importance: Retinopathy of prematurity (ROP) is a preventable cause of blindness in children. Without treatment, more than 45% of eyes may suffer permanent vision loss. Current ROP screening guidelines, which include a range of birth weights (BWs) and gestational ages (GAs), may require screening many low-risk preemies who might develop severe ROP. Method: All high-risk infants in the neonatal intensive care unit (NICU) of the First Affiliated Hospital of Zhengzhou University from 2017 to 2021 were included in this retrospective cohort study. Each of the 27 candidate risk factors was evaluated in univariate analysis and adjusted for known risk factors (i.e., GA and BW). The significant results were analyzed in a backward selection multivariate logistic regression model. Receiver operating characteristic (ROC) curves and a nomogram were drawn. Results: The study included 2,040 infants who underwent ROP screening. The weight gain rate [OR, 2.65; 95% confidence interval (CI), 1.49-1.21 ≤ 12 g/d vs. > 18 g/d; P = 0.001], blood transfusion (OR, 2.03; 95% CI, 1.14-3.64; P = 0.017), invasive mechanical ventilation (OR, 1.74; 95% CI, 1.15-2.66; P = 0.009) and N-terminal segment of pro-B-type natriuretic peptide (NT-proBNP) ≥ 25,000 ng/L (OR, 1.51; 95% CI, 1.00-2.28; P = 0.048) were four new statistically independent risk factors in addition to GA and BW. The area under the curve (AUC) of the final multivariate model was 0.90 (95% CI, 0.88-0.92; P < 0.001). Conclusions and Relevance: These findings add to our understanding of ROP screening because they include all eligible infants rather than only high-risk infants, as in previous studies. Under the control of BW and GA, low weight gain rate, increased number of blood transfusion, invasive mechanical ventilation and NT-proBNP ≥ 25,000 ng/L were "new" statistically independent risk factors for ROP. The ROP risk can be calculated manually or represented by a nomogram for clinical use.

Development of a Nomogram for Moderate-to-Severe Bronchopulmonary Dysplasia or Death: Role of N-Terminal Pro-brain Natriuretic Peptide as a Biomarker.

Objectives: This study aimed to explore the clinical value of N-terminal pro-brain natriuretic peptide (NT-proBNP) in predicting moderate-to-severe bronchopulmonary dysplasia (BPD)/death, and to establish an effective clinical predictive nomogram. Methods: We retrospectively analyzed very low birth weight infants (VLBWs) with gestational age ≤ 32 weeks. The NT-proBNP values were determined on the 1st, 3rd, 7th, 14th, 21st, and 28th days after birth. The correlation between NT-proBNP level and moderate-to-severe BPD/death was evaluated. Receiver operating characteristic (ROC) curve analysis was used to evaluate the prediction ability. Then, we used multivariable logistic regression to build the prediction model and nomogram, and calibration of the model was assessed by calibration curve. Results: In total, 556 VLBWs were involved, among whom 229 developed BPD (mild: n = 109; moderate: n = 68; severe: n = 52) and 18 died. The NT-proBNP level in the moderate-to-severe BPD/death group was significantly higher than that in the no-to-mild BPD group from the 3rd to 28th day (P < 0.001). When the natural logarithm of the serum NT-ProBNP level increased by 1 unit at day 7 (±2 days) of life, the risk of moderate and severe BPD/death was the highest (OR = 3.753; 95% CI: 2.984~4.720), and ROC analysis identified an optimal cutoff point of 3360 ng/L (sensitivity: 80.0%; specificity: 86.2%; AUC: 0.861). After adjusting for confounding factors, the level of NT-proBNP at day 7 (±2 days) of life still had important predictive value for the development of moderate-to-severe BPD/death, significantly improving the predictive ability of the model. Conclusion: The level of NT-proBNP at day 7 (±2 days) of life can be used as an early promising biomarker for VLBWs to develop moderate-to-severe BPD/death. We constructed an early predictive nomogram to help clinicians identify high-risk populations.

Association Between NT-proBNP and Prolonged Length of Stay in Hospital Among Preterm Infants Born at 28-31 Weeks' Gestation.

OBJECTIVE: In the early life of preterm infants, the relationship between heart function and length of hospitalization is unclear. This study aims to examine the association between serum NT-proBNP level on the 7th day (NT-proBNP7) after birth and length of hospitalization among preterm infants. METHODS: A retrospective cohort study was conducted. Patients included 709 preterm infants born at 28-31 weeks' gestational age (GA) admitted to the NICU of the First Affiliated Hospital of Zhengzhou University between December 20, 2016, to April 31, 2021. Main outcome: Late discharge (postmenstrual age at discharge was in the fourth quartile (highest) among infants born at the same GA). Exposure factor: NT-proBNP7. RESULTS: We observed increased prevalence ratios for late discharge among the tertile of logarithm of NT-proBNP7 level (LnNT-proBNP7) which was positive. Compared with the lowest tertile, infants in the highest tertile of LnNT-proBNP7 had an 8.4-fold increased probability of late discharge, and the results were consistent for the subgroups. Next, a non-linear (S-shaped) relationship between LnNT-proBNP7 and late discharge was observed, whose turning points were 7.5 and 9. The effect sizes and the confidence intervals on the left of the first turning point, between two turning points and on the right of the second turning point, were 0.6 (95% CI, 0.2-1.6), 5.0 (95% CI, 2.4-10.6), and 1.1 (95% CI, 0.2-6.1), respectively. In addition, the prevalence of BPD, NEC, nosocomial infection, or any of them was highest in the group of LnNT-proBNP7 ≥ 9, lowest in the group of LnNT-proBNP7 < 7.5. CONCLUSION: Higher NT-proBNP7 levels were associated with longer hospitalization. The relationship between LnNT-proBNP7 and late discharge was S-shaped. LnNT-proBNP7 was positively related with late discharge when LnNT-proBNP7 was between 7.5 and 9.

Congenital Myasthenic Syndrome Caused by a Novel Hemizygous CHAT Mutation.

Congenital myasthenic syndrome (CMS) is a neuromuscular transmission disorder caused by mutations in genes encoding neuromuscular junction proteins. CMS due to choline acetyltransferase (CHAT) gene mutation is characterized by episodic apnoea. To date, 52 cases of CMS caused by CHAT gene mutations have been reported. Here, we report a neonate with the third hemizygous mutation [a 4.9 Mb deletion [10q11.22-10q11.23 (chr10: 46123781-51028772)] containing the whole CHAT gene and c.1976A>T (p.Gln659Leu in the CHAT gene)]. The c.1976A>T (p.Gln659Leu) variant had not been reported in the ExAC or gnomAD databases and was predicted to be pathogenic. The alignment of amino acid sequences revealed that glutamine at codon 659 is highly conserved in different species and causes structural changes in the substrate-binding site. Our female patient with neonate-onset CMS presented with apnoea, dyspnoea, feeding difficulties, weak crying, and seizure-like episodes, and her respiration was ventilator dependent. The prostigmine test was positive. This case may help to further elucidate clinical features and treatment methods in neonate-onset CMS caused by CHAT gene mutations.

Increased risk of intracranial hemorrhage in preterm infants with OPRM1 gene A118G polymorphism.

BACKGROUND: To investigate the relationship between the OPRM1 gene A118G polymorphism and intracranial hemorrhage (ICH) in premature infants and identify the relevant genes in disease occurrence. METHODS: In the present case study analysis, polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) was used to detect the genotype and allele frequencies of the OPRM1 gene All8G single nucleotide polymorphism (SNP) in a case group of premature infants with ICH (n=167) and a control group of premature infants (n=163) without ICH. RESULTS: In the case group, 73 (43.7%) wild type A118 homozygous (A/A), 82 (49.1%) mutant heterozygous (A/G), and 12 (7.2%) mutant G118 homozygous (G/G) individuals were observed. The frequencies of A and G alleles were 68.3% and 31.7% respectively. In the control group, 89 (54.6%) wild type A118 homozygous (A/A), 68 (41.7%) mutant heterozygous (A/G), and 6 (3.7%) mutant G118 homozygous (G/G) individuals were observed. The frequencies of A and G alleles were 75.5% and 24.5% respectively. There was no significant difference in the frequency distribution of the OPRM1 gene A118G polymorphism between the two groups (χ2=4.839, P=0.089). There was a significant difference in the positive rate of wild-type AA and mutant-type (A/G + G/G) between the two groups (χ2=3.913, P=0.048). Carrying the G allele of the individual was 1.5 times more frequent suffering from the risk of ICH than carrying the A allele [odds ratio (OR): 1.549; 95% confidence interval (CI): 1.003-2.391], indicating that the OPRM1 118G allele was positively correlated with ICH and can increase the risk of ICH occurrence. CONCLUSIONS: The OPRM1 gene A118G polymorphism is associated with ICH in premature infants. The OPRM1 gene A118G may play a critical role in the occurrence of ICH.

miR-34a inhibits progression of neuroblastoma by targeting autophagy-related gene 5.

Neuroblastoma (NB) is a common pediatric malignancy with high mortality in childhood. Although many attentions have been gained, novel biomarkers for NB diagnosis and prognosis are still needed. microRNAs (miRNAs) played important roles in NB progression and miR-34a is a tumor suppressor in NB. However, the mechanism that underlies miR-34a regulating proliferation, migration, invasion and autophagy in NB remains poorly understood. In this study, cell proliferation was investigated by MTT and colony assay. Cell apoptosis was measured by caspase 3 activity assay. Cell migration and invasion were detected by trans-well analysis. Autophagy was measured via GFP-LC3 puncta fluorescence assay and western blots (WB). The expression of miR-34a was examined by quantitative real-time PCR (qRT-PCR). The regulatory effect of miR-34a on autophagy-related gene 5 (ATG5) was detected by qRT-PCR and WB. The interaction between miR-34a and ATG5 was probed by luciferase activity and RNA immunoprecipitation (RIP) assay. Results showed that miR-34a expression was inhibited in NB tissues and cells with low survival rate. Addition of miR-34a suppressed cell proliferation, migration, invasion and autophagy but promoted apoptosis in NB cells, whereas miR-34a deficiency played opposite roles in NB progression. Intriguingly, ATG5 was directly targeted by miR-34a. Moreover, ATG5 restoration attenuated miR-34a-mediated inhibitory effect on proliferation, apoptosis, migration, invasion and autophagy. These results indicated miR-34a suppressed proliferation, apoptosis, migration, invasion and autophagy in NB cells by targeting ATG5, providing a novel therapeutic avenue for NB treatment.

Phenotypic and functional characterization of cytokine-induced killer cells derived from preterm and term infant cord blood.

Cord blood has gradually become an important source for hematopoietic stem cell transplantation (HSCT) in the human, particularly in pediatric patients. Adoptive cellular immunotherapy of patients with hematologic malignancies after umbilical cord blood transplant is crucial. Cytokine­induced killer (CIK) cells derived from cord blood are a new type of antitumor immune effector cells in tumor prevention and treatment and have increasingly attracted the attention of researchers. On the other hand, it has been suggested that preterm infant cord blood retains an early differentiation phenotype suitable for immunotherapy. Therefore, we determined the phenotypic and functional characterization of CIK cells derived from preterm infant cord blood (PCB-CIK) compared with CIK cells from term infant cord blood (TCB-CIK). Twenty cord blood samples were collected and classified into two groups based on gestational age. Cord blood mononuclear cells (CBMCs) were isolated, cultured and induced to CIK cells in vitro. We used flow cytometry to detect cell surface markers, FlowJo software to analyze the proliferation profile and intracellular staining to test the secretion of cytokines. Finally, we evaluated the antitumor activity of CIK cells against K562 in vitro. Compared with TCB-CIK, PCB-CIK cells demonstrated faster proliferation and higher expression of activated cell surface markers. The secretion of IL-10 was lower in PCB-CIK cells while the expression of perforin and CD107a had no significant difference between the two cell groups. PCB-CIK cells exhibited a high proliferation rate while the cytotoxic activity had no difference between the PCB-CIK and TCB-CIK cells. Hence preterm infant cord blood may be a potential source for immunotherapy.