Comment on, "Increased expression of ephrin A1 in brain arteriovenous malformation: DNA microarray analysis".

This study by Sasahara et al. explores the role of ephrin A1 in brain arteriovenous malformations (AVM) using DNA microarray analysis, quantitative real-time RT-PCR, and immunohistochemistry. The research identifies significant upregulation of ephrin A1 in AVM, suggesting its potential involvement in the abnormal vascular architecture characteristic of this condition. The study's innovative methodology and thorough exploration of gene expression patterns contribute valuable insights into AVM pathogenesis, highlighting ephrin A1 as a potential therapeutic target. However, the study's limitations include clinical variability among patient samples and the use of draining veins as controls, which may affect the robustness of the findings. Future research should address these limitations by using more homogeneous samples and expanding the investigation to include other ephrin family members. This could provide a broader understanding of ephrin signaling in AVM and guide the development of targeted therapies.

Dysregulation of metallothionein MT1 sub-types in TCF3::PBX1 pre-B-cell acute lymphoblastic leukemia.

The translocation between chromosomes 1 and 19 t(1;19) produces the TCF3::PBX1 fusion protein, which leads to childhood pre-B-cell acute lymphoblastic leukemia (ALL). The molecular mechanism of oncogenesis, however, remains obscure. This study aims to identify the genes specifically dysregulated in TCF3::PBX1 translocation. The publicly available expression microarray datasets on ALL were used for weighted gene co-expression network analysis (WGCNA) to identify modules associated with TCF3::PBX1. The available knockdown and ChIP-Seq datasets were used to assess the direct targets of TCF3::PBX1. The WGCNA revealed a module enriched in genes involved in the metal ion stress to be positively correlated with TCF3::PBX1, with metallothionein isoform MT1 subtypes MT1E, MT1F, MT1G, MT1H, and MT1X as the hub genes. Of the 145 positively correlated genes, 19 were downregulated upon TCF3::PBX1 knockdown. Eleven of these 19 genes including MT1G, showed TCF3::PBX1 occupancy at the promoter. The Metallothionein 1 family has been implicated in various cancers; however, their role in t(1;19) pre-B-cell ALL has not been previously demonstrated. Our analysis effectively accounts for the cellular and population-level heterogeneity and identifies a novel mechanism for the TCF3::PBX1 action.

Genetic Insights Into Early Pregnancy Loss: A Case Study of Trisomy 22 and an Enlarged Yolk Sac.

The first trimester of pregnancy is crucial for organ development but also carries a high risk of complications, with early pregnancy loss being the most common. Anomalies in the yolk sac, the first extra-embryonic structure seen by ultrasonography, can indicate severe fetal growth abnormalities and are linked to higher rates of first-trimester loss. This case report details a 38-year-old woman with a history of recurrent pregnancy loss (RPL) who presented with per vaginal bleeding and mild abdominal pain. Transvaginal ultrasonography revealed a yolk sac larger than 10 mm, prompting further genetic investigation. Chromosomal microarray analysis confirmed Trisomy 22. The presence of an enlarged yolk sac, correlated with Trisomy 22, highlights the importance of early detection through sonography and genetic testing. This approach aids in managing RPL by identifying genetic causes, thereby informing pre-conception counseling and future pregnancy management. An abnormal yolk sac size necessitates thorough evaluation, including cytogenetic microarray testing and quantitative fluorescent-polymerase chain reaction analysis, to guide clinical decisions and improve pregnancy outcomes.

1,2,4-trihydroxybenzene induces non-apoptotic cell death via the structural damage of intracellular organelles.

Benzene occurs naturally and is widely applied in the production process of petrochemical products. It is mainly exposed through the respiratory tract and dermal and metabolized in the liver, leading to systemic health effects, and 1,2,4-trihydroxybenzene (THB) is a benzene metabolite used as a hair dye ingredient in some countries. In an effort to identify a toxic mechanism of THB, we first analyzed the hair of consumers who used a shampoo containing THB, and contrary to our expectations, THB was not persistent in the hair. Following, we treated THB to human keratinocytes and HeLa Chang liver cells. Membrane damage was observed in both cell lines, which was more notable in HeLa Chang liver cells than in keratinocytes. Thus, we decided on HeLa Chang liver cells as target cells for further study. Cell viability decreased sharply between 20 µg/ml and 40 µg/mL, inducing G2/M phase arrest and non-apoptotic cell death. The expression of carcinogenesis-, DNA damage-, and transcriptional dysregulation-related genes were notably up-regulated, and the structure and function of mitochondria were disrupted. The volume of the ER and acidic compartments decreased, and intracellular ROS and calcium ion levels increased. More interestingly, we found that THB formed unique structures within the cells, especially around the nuclear membrane, and that those structures seemed to dig into the nucleus over time. A reverse docking analysis also showed that SULT1A1, CYP2E1, and CAT, known to play a significant role in protecting cells from harmful factors, might be potential target proteins for THB. Taken together, we suggest that THB induces non-apoptotic cell death via structural damage of intracellular organelles, especially the nuclear membrane.

Lectin-Based Fluorescent Comparison of Glycan Profile-FDA Validation to Expedite Approval of Biosimilars.

Glycan profile comparisons are one of the most tedious analytical exercises for establishing compliance with recombinant therapeutic protein batches. Based on its intensive research, the FDA has confirmed that lectin array binding with fluorescent monitoring is the fastest and most reliable method for profile comparisons. Using a database of over 150 biological products expressed in nine diverse mammalian cell systems, the FDA immobilized 74 lectins to study their binding using fluorescently labeled glycoproteins. The FDA identified nine distinct lectins from a custom-designed lectin microarray: rPhoSL, rOTH3, RCA120, rMan2, MAL_I, rPSL1a, PHAE, rMOA, and PHALs, which detect core fucose, terminal GlcNAc, terminal ß-galactose, high mannose, α-2,3-linked sialic acids, α-2,6-linked sialic acids, bisecting GlcNAc, terminal α-galactose, and triantennary structures, respectively. This method can be used for screening and routine testing and to monitor batch-to-batch variability of therapeutic proteins, including establishing analytical similarity as a crucial part of biosimilar development.

Novel Role of the ALPI Gene Associated with Constipation Caused by Complement Component 3 Deficiency.

Complement component 3 (C3) deficiency has recently been reported as one of the novel causes of constipation. To identify a unique gene specific to constipation caused by C3 deficiency, the total RNA extracted from the mid colon of C3 knockout (C3 KO) mice was hybridized to oligonucleotide microarrays, and the function of the candidate gene was verified in in vitro and in vivo models. C3 KO mice used for microarrays showed definite phenotypes of constipation. Overall, compared to the wild type (WT), 1237 genes were upregulated, and 1292 genes were downregulated in the C3 KO mice. Of these, the major genes included were lysine (K)-specific demethylase 5D (KDM5D), olfactory receptor 870 (Olfr870), pancreatic lipase (PNLIP), and alkaline phosphatase intestinal (ALPI). Specifically, the ALPI gene was selected as a novel gene candidate based on alterations during loperamide (Lop)-induced constipation and intestinal bowel disease (IBD). The upregulation of ALPI expression treated with acetate recovered the expression level of mucin-related genes in primary epithelial cells of C3 KO mice as well as most phenotypes of constipation in C3 KO mice. These results indicate that ALPI plays an important role as the novel gene associated with C3 deficiency-induced constipation.

Profiling of autoantibodies in the sera of glioblastoma patients.

Aim: This study aimed to determine the expression pattern of autoantibody proteins from the serum of grade IV glioblastoma patients.Materials & methods: We performed high throughput antibody profiling via the Sengenics i-Ome® Protein Array to determine the differentially expressed autoantibodies.Results: The results portrayed that anti-COL4A3BP and anti-HSP90AA1 were among the upregulated autoantibodies in glioblastoma sera.Conclusion: The selected autoantibodies offer promising targets for future glioblastoma pathogenesis. However, further validation is required to elucidate the autoantibody signature in glioblastoma patients.

Glioblastoma is a disease affecting many people. Herein we have identified several proteins called autoantibodies that are abundant in the serum of glioblastoma patients and can be used as targets to treat or diagnose this disease.

Novel long-acting treatment for schizophrenia based on paliperidone dissolving and implantable microarray patches.

Schizophrenia is a severe mental disorder that affects millions of people worldwide. Several atypical antipsychotic medications, including paliperidone (PPD), has been developed and proven effective in treating it. To date, four PPD extended-release products have been launched commercially, providing up to six months of therapeutic effect with a single administration. However, the need for hospital injections by professional healthcare workers not only lead to poor patients' adherence, but also put additional pressure on the healthcare system. Therefore, three PPD microarray patch (PPD MAP) systems based on dissolving microneedle technology and implantable microneedle technology were developed in this work. The two dissolving microarray patch systems contained either PPD crude drug (PPD DMAP-CD) or PPD nanocrystal (PPD DMAP-NC) and the implantable MAP contained PPD crude drug (PPD IMAP). All three types of PPD MAPs showed excellent mechanical and insertion properties as they achieved over 256 µm insertion depth in skin model. In vitro release study showed that PPD released from IMAP in a much more sustained manner (up to 14 days) than PPD did from DMAPs (7 days), with only 20 % initial burst release from IMAP compared with 43-71 % from DMAPs. The MAP dissolution study showed that both DMAPs can be immediately dissolved within less than 3 min once inserted into the skin, indicating a faster action potential compared with IMAP. Ex vivo delivery study showed that 1.68 ±â€¯0.23 mg, 1.39 ±â€¯0.07 mg, and 1.18 ±â€¯0.12 mg were delivered from DMAP-CD, DMAP-NC and IMAP, respectively, demonstrating that over 50 % and up to 70 % of PPD in the MAPs can be delivered into the skin. The IMAP offers most sustained release of PPD whereas DMAP-NC exhibits fastest PPD release (11.19 % vs 20.01 % into Franz cell receiver compartment over 24 h). This work presents a promising alternative for the sustained delivery of antipsychotic drugs, allowing for patient self-administration and extended release concurrently. Patients may potentially use both DMAP and IMAP to achieve a sustained release of PPD while also avoid having an initial therapeutic lag.

Profiling of B cells and their subsets by whole blood gene expression analysis versus flow cytometry in multiple sclerosis.

We investigated if differentially expressed mRNA targets could be used as surrogate markers for circulating B cells and subsets. In paired blood samples from patients with untreated, anti-CD20-treated, fingolimod-treated, and natalizumab-treated multiple sclerosis, whole blood expression of CD19 correlated with B cell counts determined by flow cytometry, ROR1 with transitional B cells, TCL1A and ZNF727 with naïve B cells, NEXMIF with memory B cells and BCMA with plasmablasts. CD19 expression distinguished patients with B cell repletion and may be used as an alternative to flow cytometry, but NEXMIF was unsuitable for memory B cell monitoring in rituximab-treated patients.

Molecular cytogenetic characterization of isolated recurrent 4q35.2 microduplication in Chinese population: a seven-year single-center retrospective study.

BACKGROUND: With the extensive use of chromosomal microarray analysis (CMA), an increasing number of variants of uncertain significance (VOUS) have been detected. The objective of the present study was to elucidate the pathogenicity and clinical variability associated with isolated recurrent 4q35.2 microduplications within the Chinese population. METHODS: The present study involved 14 cases of isolated recurrent 4q35.2 microduplication (including 12 fetuses and 2 cases of pediatric patients) out of 5,188 subjects who sought genetic consultation at our hospital and received CMA detection. WES technology was subsequently utilized to identify additional sequence variants in a patient with multiple clinical anomalies. RESULTS: All 14 cases exhibited isolated recurrent 4q35.2 microduplications spanning a 1.0-Mb region encompassing the ZFP42 gene. Among the 12 fetuses, 11 displayed normal clinical features, while one was born with renal duplication and hydronephrosis. Additionally, in the two pediatric patients, WES was performed for Case 1, who presented with congenital cataracts, severe intellectual disability, and seizures. This patient inherited the 4q35.2 microduplication from his phenotypically normal mother. WES identified a novel NM_000276:c.2042G > T (p.G681V) variant in the OCRL gene, which is associated with Lowe syndrome and may account for the observed phenotypic variability within this family. CONCLUSION: A series of 14 cases with isolated recurrent 4q35.2 microduplications were investigated, highlighting a potential association with increased susceptibility to renal abnormalities. Further, the present findings may expand the mutation spectrum of the OCRL gene associated with Lowe syndrome and provide valuable insights for the genetic etiological diagnosis of patients with unexplained copy number variants.

Clinical and genetic analysis of two phenotypically normal families carrying 4p16.1 microduplications.

OBJECTIVE: To help determine the pathogenicity of 4p16.1 microduplications, we reported two asymptomatic families carrying this variation. CASE REPORT: We present the prenatal diagnosis and genetic analysis of two normal families with 4p16.1 microduplications. CONCLUSION: This paper highlights two families with clinically asymptomatic 4p16.1 microduplications that assisted in determining the pathogenicity of this fragment. The findings can be used as a reference for genetic counseling in cases of similar abnormalities encountered during future prenatal diagnosis.

Insights into Glycobiology and the Protein-Glycan Interactome Using Glycan Microarray Technologies.

Glycans linked to proteins and lipids and also occurring in free forms have many functions, and these are partly elicited through specific interactions with glycan-binding proteins (GBPs). These include lectins, adhesins, toxins, hemagglutinins, growth factors, enzymes, but antibodies can also bind glycans. While humans and other animals generate a vast repertoire of GBPs and different glycans in their glycomes, other organisms, including phage, microbes, protozoans, fungi, and plants also express glycans and GBPs, and these can also interact with their host glycans. This can be termed the protein-glycan interactome, and in nature is likely to be vast, but is so far very poorly described. Understanding the breadth of the protein-glycan interactome is also a key to unlocking our understanding of infectious diseases involving glycans, and immunology associated with antibodies binding to glycans. A key technological advance in this area has been the development of glycan microarrays. This is a display technology in which minute quantities of glycans are attached to the surfaces of slides or beads. This allows the arrayed glycans to be interrogated by GBPs and antibodies in a relatively high throughput approach, in which a protein may bind to one or more distinct glycans. Such binding can lead to novel insights and hypotheses regarding both the function of the GBP, the specificity of an antibody, and the function of the glycan within the context of the protein-glycan interactome. This article focuses on the types of glycan microarray technologies currently available to study animal glycobiology and examples of breakthroughs aided by these technologies. (254 words).

The storage time of cryopreserved human spermatozoa does not affect pathways involved in fertility.

BACKGROUND: Cryopreservation of human spermatozoa is a widely used technique in the assisted reproduction technology laboratory for the storage of gametes for later use, for the fertility preservation and for sperm donation programs. Cryopreservation can cause damage to membrane, cytoskeletal, acrosome and increased oxidative stress, sperm DNA damage and transcriptome changes. To assess the impact of storage time on the transcriptome of frozen human spermatozoa, semen samples were collected from 24 normospermic donors of whom 13 had cryostored semen for a short-time (1 week) and 11 had cryostored semen for a long-time (median 9 years). RESULTS: RNA was extracted from each frozen-thawed sperm sample, randomized in pools, and analyzed by microarrays. Five transcripts were in higher abundance in the long-time respect to the short-time storage group. Functional annotation enrichment disclosed that that the length of cryostorage has no effect on critical pathways involved in sperm physiology and function. CONCLUSIONS: The storage time of cryopreserved human spermatozoa does not affect pathways involved in fertility.

RéSUMé: CONTEXTE: La cryoconservation des spermatozoïdes humains est une technique largement utilisée, dans les laboratoires de procréation médicalement assistée, pour le stockage des gamètes en vue d'une utilisation ultérieure, dans le cadre d'une préservation de la fertilité et dans les programmes de don de sperme. La cryoconservation peut altérer la membrane, le cytosquelette, l'acrosome, et augmenter le stress oxydatif des spermatozoïdes, endommager l'ADN et modifier le transcriptome. Pour évaluer l'impact du temps de stockage sur le transcriptome de spermatozoïdes humains congelés, des échantillons de sperme ont été prélevés auprès de 24 donneurs normozoospermiques, dont 13 avaient cryoconservé du sperme pendant une courte période (1 semaine) et 11 avaient cryoconservé du sperme pendant une longue période (médiane de 9 ans). RéSULTATS: L'ARN a été extrait de chaque échantillon de sperme congelé-décongelé, randomisé dans des pools et analysé par microarrays. Cinq transcrits étaient en plus grande abondance dans le groupe de stockage à long terme que dans le groupe de stockage de courte durée. L'enrichissement en annotation fonctionnelle a révélé que la durée de la cryoconservation n'a aucun effet sur les voies critiques impliquées dans la physiologie et la fonction des spermatozoïdes. CONCLUSIONS: Le temps de stockage des spermatozoïdes humains cryoconservés n'affecte pas les voies impliquées dans la fertilité.

Inter-individual and inter-regional variability of breast milk antibody reactivity to bacterial lipopolysaccharides.

Breast milk is a vital source of nutrients, prebiotics, probiotics, and protective factors, including antibodies, immune cells and antimicrobial proteins. Using bacterial lipopolysaccharide arrays, we investigated the reactivity and specificity of breast milk antibodies towards microbial antigens, comparing samples from rural Kenya and urban Switzerland. Results showed considerable variability in antibody reactivity both within and between these locations. Kenyan breast milk demonstrated broad reactivity to bacterial lipopolysaccharides, likely due to increased microbial exposure. Antibodies primarily recognized the O-antigens of lipopolysaccharides and showed strong binding to specific carbohydrate motifs. Notably, antibodies against specific Escherichia coli O-antigens showed cross-reactivity with parasitic pathogens like Leishmania major and Plasmodium falciparum, thus showing that antibodies reacting against lipopolysaccharide O-antigens can recognize a wide range of antigens beyond bacteria. The observed diversity in antigen recognition highlights the significance of breast milk in safeguarding infants from infections, particularly those prevalent in specific geographic regions. The findings also offer insights for potential immunobiotic strategies to augment natural antibody-mediated defense against diverse pathogens.

Comment on, "Gene expression in a canine basilar artery vasospasm model: a genome-wide network-based analysis".

The study by Sasahara et al. (2008) offers a comprehensive exploration of the molecular mechanisms underlying cerebral vasospasm following subarachnoid hemorrhage, utilizing genome-wide microarray technology and network-based analysis in a canine model. Their work identifies significant gene expression changes, particularly in IL-6, IL-8, and CCL2, which are implicated in cell signaling, host-pathogen interactions, and immune responses. Despite the study's methodological rigor, it is limited by a single time-point analysis and the use of non-injected controls, which may not fully account for procedural effects. Future studies should include a time-course analysis and more appropriate controls, as well as isolate specific cell types to enhance the relevance of the findings. Further research could explore therapeutic interventions targeting the identified pathways, particularly those involved in calcium signaling and inflammation, to develop more effective treatments for cerebral vasospasm.

Identification and functional characterization of differentially expressed circRNAs in high glucose treated endothelial cells: Construction of circRNA-miRNA-mRNA network.

Background: Endothelial dysfunction is a complication of diabetes mellitus (DM), characterized by impaired endothelial function in both microvessels and macrovessels, closely linked to atherosclerosis (AS). Endothelial dysfunction, characterized by impaired endothelial cell (EC) function, is a pivotal factor in AS and DM. Circular RNAs (circRNAs) are endogenous non-coding RNAs that can act as competing endogenous RNAs (ceRNAs) and regulate gene expression. However, the role of circRNAs in ECs dysfunction and AS under high glucose (HG) condition remains elusive. Methods: We performed high-throughput sequencing to identify differentially expressed (DE) circRNAs in human umbilical vein endothelial cells (HUVEC) exposed to HG, one risk factors of endothelial dysfunction and AS. We then validated eight candidate circRNAs by qRT-PCR and functional analysis, directing our attention to hsa_circ_0122319. Moreover, microarray analysis identified the differential expression profiles of miRNAs and mRNAs regulated by hsa_circ_0122319. Subsequently, the construction of the ceRNAs network employed bioinformatic analysis and Cytoscape software. Furthermore, the role of the PI3K-Akt signaling pathway in regulating ceRNAs was evaluated. Results: We detected 917 DE circRNAs in HG treated HUVEC. The parental genes of these circRNAs were enriched in cell cycle, cellular senescence and endocytosis related pathways. The differential expression of hsa_circ_0122319 was confirmed to be most obvious at the cellular level and in clinical samples by qPCR experiments. After overexpression of hsa_circ_0122319, 49 DE miRNAs and 459 DE mRNAs were identified using microarray analysis. Subsequently, a ceRNAs network was constructed, comprising hsa_circ_0122319, 8 miRNAs, and 41 mRNAs. Conclusion: In summary, our study delves into the role of circRNAs in endothelial dysfunction associated with DM and AS. Through high-throughput sequencing and validation, we identified hsa_circ_0122319 as a pivotal regulator of ECs function under HG conditions. It also showed that hsa_circ_0123319 has the potential to serve as a biomarker for DM and its vascular complications, and provides new evidence for future exploration of the intricate molecular mechanisms of endothelial dysfunction in the progression of DM and AS.

Prenatal diagnosis of 17q12 copy number variants in fetuses via chromosomal microarray analysis - A retrospective cohort study and literature review.

Purpose: 17q12 copy number variants (CNVs) have variable presentations and incomplete penetrance, challenging prenatal counseling and management. This study aims to investigate the intrauterine phenotype. Methods: We included 48 fetuses diagnosed with 17q12 microdeletion or microduplication by chromosomal microarray analysis. Results: For 17q12 deletion, renal anomalies were found in 35 fetuses (35/37, 94.6 %), with hyperechogenic kidneys (HEK, 28/37, 75.7 %) and multicystic dysplastic kidneys (17/37, 45.9 %) being the most common findings. Duodenal obstruction (DO) was most frequently combined in 17q12 duplication fetuses. In addition, cardiac abnormalities were the first reported prenatal phenotype in 17q12 duplication fetuses. Conclusion: Our study shows that HEK and DO are the most predominant presentations of 17q12 deletion and duplication, respectively, and cardiac structural abnormalities may be associated with the latter. Although 17q12 CNVs have incomplete penetrance and variable expressivity and may be mainly involved in neurodevelopmental disorders, their short-term prognosis appears positive.

Unveiling therapeutic biomarkers and druggable targets in ALS: An integrative microarray analysis, molecular docking, and structural dynamic studies.

Amyotrophic lateral sclerosis (ALS), commonly known as Lou Gehrig's disease, is a debilitating neurodegenerative disorder characterized by the progressive degeneration of nerve cells in the brain and spinal cord. Despite extensive research, its precise etiology remains elusive, and early diagnosis is challenging due to the absence of specific tests. This study aimed to identify potential blood-based biomarkers for early ALS detection and monitoring using datasets from whole blood samples (GSE112680) and oligodendrocytes, astrocytes, and fibroblasts (GSE87385) obtained from the NCBI-GEO repository. Through bioinformatics analysis, including protein-protein interactions and molecular pathway analyses, we identified differentially expressed genes (DEGs) associated with ALS. Notably, ALS2, ADH7, ALDH8A1, ALDH3B1, ABHD2, ABHD17B, ABHD12, ABHD13, PGAM2, AURKB, ANAPC11, VAPA, UNC45B, and TNNT2 emerged as top-ranked DEGs, implicated in drug metabolism, protein depalmytilation, and the AKT/mTOR signaling pathways. Among these, AurKB established as a potential therapeutic biomarker with relevance to various neurological conditions. Consequently, AurKB was selected for identifying potential therapeutic molecules and utilized for in silico structural characterization studies. Exploration of the IMPATT database led to the discovery of a lead compound similar to Fostamatinib, currently used for AurKB. Initial molecular docking and MMGBSA-based binding energy analysis were followed by molecular dynamics simulation (MDS) and free energy landscape (FEL) analysis to validate the ligand's binding efficacy and understand dynamic processes within the biological system. The identified potential biomarkers and lead molecule provide novel insights into the correlation between blood cell transcripts and ALS pathology, paving the way for blood-based diagnostic tools for early ALS detection and ongoing disease monitoring.

Diagnostic Utility of Whole Genome Sequencing After Negative Karyotyping/Chromosomal Microarray in Infants Born With Multiple Congenital Anomalies.

BACKGROUND: Achieving a definitive genetic diagnosis of unexplained multiple congenital anomalies (MCAs) in neonatal intensive care units (NICUs) infants is challenging because of the limited diagnostic capabilities of conventional genetic tests. Although the implementation of whole genome sequencing (WGS) has commenced for diagnosing MCAs, due to constraints in resources and faculty, many NICUs continue to utilize chromosomal microarray (CMA) and/or karyotyping as the initial diagnostic approach. We aimed to evaluate the diagnostic efficacy of WGS in infants with MCAs who have received negative results from karyotyping and/or CMA. METHODS: In this prospective study, we enrolled 80 infants with MCAs who were admitted to a NICU at a single center and had received negative results from CMA and/or karyotyping. The phenotypic characteristics were classified according to the International Classification of Diseases and the Human Phenotype Ontology. We assessed the diagnostic yield of trio-WGS in infants with normal chromosomal result and explored the process of diagnosing by analyzing both phenotype and genotype. Also, we compared the phenotype and clinical outcomes between the groups diagnosed with WGS and the undiagnosed group. RESULTS: The diagnostic yield of WGS was 26% (21/80), of which 76% were novel variants. There was a higher diagnostic yield in cases of craniofacial abnormalities, including those of the eye and ear, and a lower diagnostic yield in cases of gastrointestinal and genitourinary abnormalities. In addition, higher rates of rehabilitation therapy and gastrostomy were observed in WGS-diagnosed infants than in undiagnosed infants. CONCLUSION: This prospective cohort study assessed the usefulness of trio-WGS following chromosomal analysis for diagnosing MCAs in the NICU and revealed improvements in the diagnostic yield and clinical utility of WGS.