USP51/ZEB1/ACTA2 axis promotes mesenchymal phenotype in gastric cancer and is associated with low cohesion characteristics.

poorly cohesive (PC) gastric cancer (GC) (PC-GC) is a distinct histological subtype of GC and is defined as a tumor consisting of isolated or small clusters of tumor cells with poorly differentiated and metastatic characteristics. According to multiple studies, PC-GC is intrinsically heterogeneous, with mesenchymal variants being the most aggressive. However, to date, the molecular mechanisms associated with PC-GC are still not fully understood. This study investigated the role of the USP51/ZEB1/ACTA2 axis in promoting GC metastasis. Single-cell sequencing revealed that E-box binding homeobox 1 (ZEB1) expression was significantly increased in a subpopulation of low-adherent cells and was an independent prognostic factor in GC patients. Furthermore, the bulk transcriptome analysis revealed a significant positive correlation between Ubiquitin Specific Peptidase 51 (USP51), ZEB1, and Actin Alpha 2 (ACTA2), and our data further confirmed that all three were highly co-localized in PC-GC tissues. According to the findings of in vitro and in vivo experiments, USP51 was able to maintain ZEB1 expression to promote ACTA2 transcription, thereby activating the mesenchymal phenotype of GC cells and promoting tumor metastasis. Moreover, USP51 could recruit and activate stromal cells, including M2-like macrophages and fibroblasts, through cancer cells. Clinical data suggested that overexpression of USP51 predicts that patients have difficulty benefiting from immunotherapy and is associated with immune-exclusion tumor characteristics. Collectively, the findings of this study shed light on a key mechanism by which elevated USP51 expression induces Epithelial-mesenchymal transition (EMT) in GC cells, hence facilitating GC cell proliferation, survival, and dissemination. In this view, USP51/ZEB1/ACTA2 may serve as a candidate therapeutic target against GC metastasis.

[Prenatal diagnosis and genetic analysis of a rare case with 8p deletion and duplication].

OBJECTIVE: To explore the genetic etiology for a child featuring mental retardation, language delay and autism. METHODS: G-banding chromosomal karyotyping and single nucleotide polymorphism array (SNP-array) were carried out for the child and her parents. RESULTS: The child was found to have a 46,XX,dup(8p?) karyotype, for which both of her parents were normal. SNP-array revealed that the child has harbored a 6.8 Mb deletion in 8p23.3p23.1 and a 21.8 Mb duplication in 8p23.1p12, both of which were verified as de novo pathogenic copy number variants. CONCLUSION: The clinical features of the child may be attributed to the 8p deletion and duplication. SNP-array can facilitate genetic diagnosis for children featuring mental retardation in conjunct with other developmental anomalies.

Identification of COX4I2 as a hypoxia-associated gene acting through FGF1 to promote EMT and angiogenesis in CRC.

BACKGROUND: Current evidence suggests that the hypoxic tumor microenvironment further aggravates tumor progression, leading to poor therapeutic outcomes. There is as yet no biomarker capable of evaluating the hypoxic state of the tumor. The cytochrome c oxidase (COX) subunit is crucial to the mitochondrial respiratory chain. METHODS: We investigated the potential oncogenic role of COX subunit 4 isoform 2 gene (COX4I2) in colorectal cancer (CRC) by least absolute shrinkage and selection operator (LASSO) and COX regression analysis to examine whether COX4I2 overexpression can predict colorectal cancer (CRC) prognosis. The association of COX4I2 levels with clinical features and its biological actions were evaluated both in vitro and in vivo. RESULTS: Our analysis showed that elevated COX4I2 levels were correlated with poor clinical outcomes. We also observed that that COX4I2 may be involved in epithelial-mesenchymal transition, activation of cancer-related fibroblasts and angiogenesis in relation to fibroblast growth factor 1. CONCLUSIONS: The COX4I2 level may be a predictor of outcome in CRC and may represent a novel target for treatment development.

[Clinical and genetic analysis of a patient with 10q26.3 microdeletion in conjunct with 18q22.3q23 microduplication].

OBJECTIVE: To explore the genetic etiology for a patient featuring intellectual disability and torticollis. METHODS: Peripheral blood sample was collected from the patient and subjected to G-banded karyotyping analysis and single nucleotide polymorphism array (SNP-array) assay. RESULTS: The patient was found to have a chromosomal karyotype of 46,XX. SNP-array revealed that she has harbored a 3.8 Mb microdeletion at 10q26.3 which has encompassed 21 OMIM genes including EBF3 and ECHS1, and a 7.3 Mb duplication at 18q22.3q23 which has encompassed 19 OMIM genes including TSHZ1 and TXNL4A. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), the 10q26.3 deletion was predicted to be pathogenic, whilst the 18q22.3q23 duplication was predicted to be variation of unknown significance. CONCLUSION: The clinical phenotype of the patient may be mainly attributed to the 10q26.3 microdeletion, and haploinsufficiency of the EBF3 gene may account for her intellectual deficiency. Above finding has provided a basis for genetic counseling for the patient.

[Application of chromosomal microarray analysis in the diagnosis of genetic etiology of spontaneous abortions].

OBJECTIVE: To explore the genetic etiology of spontaneous abortions by using chromosomal microarray analysis (CMA). METHODS: Fetal tissues derived from 106 spontaneous abortion samples were subjected to CMA assay to detect genome copy number variants (CNVs). RESULTS: The test was successful in 94 cases (88.68%). Fifty four chromosomal abnormalities were detected, which included 44 numerical chromosomal aberrations mainly consisting of aneuploidies, triploidies and mosaicisms. Four pathogenic CNVs were detected, and two of which involved the Cri-du-chat syndrome regions. In addition, 6 chromosomal mosaicism were detected. CONCLUSION: Numerical chromosomal aberrations and CNVs are the main causes for early spontaneous abortions. CMA can effectively reveal the genetic etiology of spontaneous abortions. Spontaneous abortions at gestational weeks 10 to 11+6 has the highest rate for chromosomal abnormalities, which may provide valuable information for clinical counseling.

Application of the BACs-on-Beads assay for the prenatal diagnosis of chromosomal abnormalities in Quanzhou, China.

BACKGROUND: An increasing number of techniques have been used for prenatal diagnosis of genetic abnormalities. Our initial objective was to explore the value of the BACs-on-Beads (BoBs) assay for the prenatal diagnosis of aneuploidies and microdeletion/microduplication syndromes in Quanzhou, Southeast China. METHODS: A total of 1409 pregnant women with high-risk factors for chromosomal abnormalities admitted to Quanzhou Women's and Children's Hospital were enrolled in this study. BoBs assays and karyotype analyses were conducted for all subjects. Subsequently, chromosome microarray analysis (CMA) or fluorescence in situ hybridization (FISH) was performed to validate the findings. RESULTS: In this study, karyotype analysis and BoBs assay failed in 4 cases, and 2 cases, respectively. A total of 1403 cases were successfully analyzed, with success rates of 99.72% (1405/1409) and 99.85% (1407/1409) for karyotype analysis and Bobs assay, respectively. BoBs assay rapidly detected chromosomal aneuploidies in line with the karyotyping data. Additionally, 23 cases of microdeletions/microduplications were detected by BoBs assay but missed by karyotyping, including 22q11.2 microdeletions/microduplications, 5p15.32p15.33 microdeletion, Xp22.31 microdeletions/microduplications, Xq27.3 microdeletion, and Yp11.2 and Yq11.22q11.222 microduplication. In comparison with karyotyping, fewer mosaicisms were identified by BoBs assay. A high detection rate of chromosomal abnormalities was observed in the high-risk group during noninvasive prenatal testing (NIPT) (41.72%) and the abnormal ultrasound group (13.43%). CONCLUSIONS: BoBs assay can be used for the rapid and efficient prenatal diagnosis of common aneuploidies and microdeletion/microduplication syndromes. Moreover, the combined use of BoBs assay and karyotyping in prenatal diagnosis may allow for a more effective detection of chromosomal abnormalities.

[Molecular genetic analysis of a child with de novo 16p11.2 microdeletion].

OBJECTIVE: To explore the genetic basis for a child featuring developmental delay, intelligent disability and language deficit. METHODS: Peripheral blood samples of the child and her parents were collected for routine G-banding karyotyping analysis and single nucleotide polymorphism array (SNP array) detection. Amniotic fluid was also sampled from the mother for karyotyping analysis and SNP array detection. RESULTS: No karyotypic abnormality was found with the child and her parents. SNP array showed that the child has carried a 761.4 kb microdeletion at 16p11.2, while her mother has carried a 444.4 kb microduplication at 15q13.3. Her father's result was negative. Further analysis showed that the 15q13.3 microduplication was inherited from her maternal grandfather who was phenotypically normal. Prenatal diagnosis showed that the fetus has inherited the15q13.3 microduplication from its mother. CONCLUSION: The child has carried a de novo 16p11.2 microdeletion, which overlaps with 16p11.2 microdeletion syndrome region, in addition with similar clinical phenotypes. The 16p11.2 microdeletion probably underlies her abnormal phenotype.

[Clinical and genetic study of a child with 15q11.2 microduplication].

OBJECTIVE: To explore the genetic basis of a child with developmental delay and intellectual disability. METHODS: Peripheral blood samples of the child and his parents were collected for routine G-band karyotyping analysis and single nucleotide polymorphism array (SNP array) assay. Amniotic fluid sample was collected during the next pregnancy for prenatal diagnosis. RESULTS: No karyotypic abnormality was found in the child and his parents. SNP array showed that the child has carried a 855.3 kb microduplication in 15q11.2. His mother carried the same duplication but had no phenotypic anomaly. No microdeletion/microduplication was found in his father. Upon prenatal diagnosis, no abnormalities was found with the chromosomal karyotype and SNP array result of the fetus. CONCLUSION: 15q11.2 microduplication may result in developmental delay and intellectual disability, for which CYFIP1 may be a candidate gene. However, the duplication may increase the risk but with a low penetrance. This should attract attention during clinical consultation.

Molecular analysis of a large novel deletion causing α+-thalassemia.

BACKGROUND: α-thalassaemia is an inherited blood disorder caused by mutations in the α-globin gene cluster. Recognizing the pathogenic α-globin gene mutations associated with α-Thalassemia is of significant importance to thalassaemia's diagnosis and management. METHODS: A family with α-thalassaemia from Fujian, China was recruited for this study. The phenotype was confirmed through haematological analysis. Commercially available Gap-PCR genotypic methods were employed to identify the known deletions causing α-thalassemia. MLPA analysis was used to study the novel mutations; this was then confirmed through DNA sequencing and bioinformatics analysis. RESULTS: The proband of the family belonged to Southeast Asian type (--SEA) thalassaemia. None of the known mutations associated with α-thalassaemia were detected in this family's genetics, whereas a novel 6.9 kb deletion (16p13.3 g.29,785-36,746) covering the α2 gene on the globin gene cluster was identified with MLPA and confirmed through Sanger Sequencing. This data led us to propose a novel pathogenic deletion associated with α-thalassemia: -α6.9 /--SEA. CONCLUSIONS: A novel α-thalassaemia deletion was identified in members of a Chinese family and subsequently analyzed. This finding has helped broaden the spectrum of pathogenic mutations leading to the development of α-thalassaemia, paving the way for improved disease diagnosis and management.

[Analysis a family with partial Xq deletion].

OBJECTIVE: To analyze partial deletion of the long arm of X chromosome in a family and explore the mechanism underlying its phenotypes. METHODS: G-banding technique was employed to analyze the karyotypes of the subjects, and fluorescence in situ hybridization (FISH) was used to analyze their X chromosomes with Xpter, Xqter and WCPX probes. RESULTS: The karyotypes of the proband, her mother and her fetus were all 46,X,del(X)(q24). Combined FISH and karyotyping analysis suggested that the proband and her fetus both carried a Xq24q27.3 deletion. CONCLUSION: The Xq24q27.3 deletion carried by the family is closely related with premature ovarian failure but not with short stature, gonadal dysgenesis and primary amenorrhea.

Prenatal diagnosis of fetuses with absent/hypoplastic nasal bone in second-trimester using chromosomal microarray analysis.

BACKGROUND: Pathogenic copy number variants (pCNVs) are associated with fetal ultrasound anomalies, which can be efficiently identified through chromosomal microarray analysis (CMA). The primary objective of the present study was to enhance understanding of the genotype-phenotype correlation in fetuses exhibiting absent or hypoplastic nasal bones using CMA. METHODS: Enrolled in the present study were 94 cases of fetuses with absent/hypoplastic nasal bone, which were divided into an isolated absent/hypoplastic nasal bone group (n = 49) and a non-isolated group (n = 45). All pregnant women enrolled in the study underwent karyotype analysis and CMA to assess chromosomal abnormalities in the fetuses. RESULTS: Karyotype analysis and CMA detection were successfully performed in all cases. The results of karyotype and CMA indicate the presence of 11 cases of chromosome aneuploidy, with trisomy 21 being the most prevalent among them. A small supernumerary marker chromosome (sSMC) detected by karyotype analysis was further interpreted as a pCNV by CMA. Additionally, CMA detection elicited three cases of pCNVs, despite normal findings in their karyotype analysis results. Among them, one case of Roche translocation was identified to be a UPD in chromosome 15 with a low proportion of trisomy 15. Further, a significant difference in the detection rate of pCNVs was observed between non-isolated and isolated absent/hypoplastic nasal bone (24.44% vs. 8.16%, p < .05). CONCLUSION: The present study enhances the utility of CMA in diagnosing the etiology of absent or hypoplastic nasal bone in fetuses. Further, isolated cases of absent or hypoplastic nasal bone strongly suggest the presence of chromosomal abnormalities, necessitating genetic evaluation through CMA.

A First Clinical and Molecular Study of Rare IVS-II-806 (G > C) (HBB:c.316-45G > C) Variant in the ß-globin Gene: A Possibly Benign Variant.

Introduction: ß-thalassemia is a common genetic disease affecting a single gene, disease with a high incidence in South China. We hereby, aim to provide the clinical and hematological features of a rare ß-globin gene variant in the Chinese population. Methods: Ten subjects from three unrelated Chinese families were enrolled in this study. Hematological analysis and thalassemia gene testing were preformed to screen for common α and ß-thalassemia variants. Gap-polymerase chain reaction (Gap-PCR) and DNA sequencing were utilized to examine the rare or novel thalassemia variants. Results: Six cases were identified carrying the rare IVS-II-806 (G > C) (HBB:c.316-45G > C) variant in the ß-globin gene. The proband in family 1 carry three rare ß-globin gene mutations including CD39 (C > T), IVS-II-81 (C > T) and IVS-II-806 (G > C) combined with a --SEA/αα deletion, exhibiting the ß-thalassemia trait. Further pedigree investigation indicated that the genotype of the proband in family 1 was --SEA/αα, ßCD39 (C>T), IVS-II-81(C>T)/ßIVS-II-806(G>C). Meanwhile, the twin girls in family 1 carrying the IVS-II-806 (G > C) mutation demonstrated a normal hematological phenotype. In family 2, the proband and his sister carry the IVS-II-806 (G > C) mutation, eliciting high levels of Hb A2 and slightly low levels of MCV and MCH. Moreover, the proband in family 3 carrying the same mutation exhibited a slightly low MCV level as well. Conclusions: In this study, clinical and hematological analysis of the IVS-II-806 (G > C) mutation was first conducted within the Chinese population, with results indicating that it may be a benign variant.

Third-Generation Sequencing as a New Comprehensive Technology for Identifying Rare α- and ß-Globin Gene Variants in Thalassemia Alleles in the Chinese Population.

CONTEXT.­: Identification of rare thalassemia variants requires a combination of multiple diagnostic technologies. OBJECTIVE.­: To investigate a new approach of comprehensive analysis of thalassemia alleles based on third-generation sequencing (TGS) for identification of α- and ß-globin gene variants. DESIGN.­: Enrolled in this study were 70 suspected carriers of rare thalassemia variants. Routine gap-polymerase chain reaction and DNA sequencing were used to detect rare thalassemia variants, and TGS technology was performed to identify α- and ß-globin gene variants. RESULTS.­: Twenty-three cases that carried rare variants in α- and ß-globin genes were identified by the routine detection methods. TGS technology yielded a 7.14% (5 of 70) increment of rare α- and ß-globin gene variants as compared with the routine methods. Among them, the rare deletional genotype of -THAI was the most common variant. In addition, rare variants of CD15 (G>A) (HBA2:c.46G>A), CD117/118(+TCA) (HBA1:c.354_355insTCA), and ß-thalassemia 3.5-kilobase gene deletion were first identified in Fujian Province, China; to the best of our knowledge, this is the second report in the Chinese population. Moreover, HBA1:c.-24C>G, IVS-II-55 (G>T) (HBA1:c.300+55G>T) and hemoglobin (Hb) Maranon (HBA2:c.94A>G) were first identified in the Chinese population. We also identified rare Hb variants of HbC, HbG-Honolulu, Hb Miyashiro, and HbG-Coushatta in this study. CONCLUSIONS.­: TGS technology can effectively and accurately detect deletional and nondeletional thalassemia variants simultaneously in one experiment. Our study also demonstrated the application value of TGS-based comprehensive analysis of thalassemia alleles in the detection of rare thalassemia gene variants.

A de novo PAK1 likely pathogenic variant and a de novo terminal 1q microdeletion in a Chinese girl with global developmental delay, severe intellectual disability, and seizures.

BACKGROUND: Pathogenic PAK1 variants were described to be causative of neurodevelopmental disorder with macrocephaly, seizures, and speech delay. Herein, we present a de novo PAK1 variant combine with a de novo terminal 1q microdeletion in a Chinese pediatric patient, aiming to provide more insights into the underlying genotype-phenotype relationship. METHODS: Enrolled in this study was a 6-year-old girl with clinical features of global developmental delay, severe intellectual disability, speech delay, and seizures from Quanzhou region of China. Karyotype and chromosomal microarray analysis (CMA) were performed to detect chromosome abnormalities in this family. Whole exome sequencing (WES) was performed to investigate additional genetic variants in this family. RESULTS: No chromosomal abnormalities were elicited from the entire family by karyotype analysis. Further familial CMA results revealed that the patient had a de novo 2.7-Mb microdeletion (arr[GRCh37] 1q44(246,454,321_249,224,684) × 1]) in 1q44 region, which contains 14 OMIM genes, but did not overlap the reported smallest region of overlap (SRO) responsible for the clinical features in 1q43q44 deletion syndrome. In addition, WES result demonstrated a de novo NM_002576: c.251C > G (p.T84R) variant in PAK1 gene in the patient, which was interpreted as a likely pathogenic variant. CONCLUSION: In this study, we identify a novel PAK1 variant associated with a terminal 1q microdeletion in a patient with neurodevelopmental disorder. In addition, we believe that the main clinical features may ascribe to the pathogenic variant in PAK1 gene in the patient.

Etiological identification of recurrent male fatality due to a novel NSDHL gene mutation using trio whole-exome sequencing: A rare case report and literature review.

BACKGROUND: Congenital hemidysplasia with ichthyosiform nevus and limb defects (CHILD) syndrome is a rare X-linked dominant, lethal male disorder caused by mutations to the NSDHL (NAD(P)H steroid dehydrogenase-like protein) gene. It primarily exhibits strictly unilateral congenital hemidysplasia with ichthyosiform erythroderma and ipsilateral limb defects in female individuals. METHODS: A Chinese couple suffering from recurrent spontaneous abortion in male fetuses was enrolled in this study. Chromosomal microarray analysis and whole-exome sequencing were performed for genetic etiological diagnosis. RESULTS: A 33-year-old pregnant woman with recurrent spontaneous abortion was experiencing her third pregnancy with a male embryo. In this pregnancy, a miscarriage occurred at a gestational age of 10+6  weeks with no copy number variants. However, a novel mutation c.790-6C>T in the NSDHL gene was observed in the fetus through whole-exome sequencing (WES). Parental verification indicated that the NSDHL gene variant was inherited from the mother. Additionally, the variant in the NSDHL gene was absent in her subsequent pregnancy with a female fetus. CONCLUSION: In this study, we detected c.790-6C>T, a novel variant in the NSDHL gene that results in recurrent miscarriage in males. Our study may broaden the scope of research on the NSDHL gene in CHILD syndrome and strengthens the application value of WES for the genetic etiological identification of recurrent miscarriage.

Molecular cytogenetic analysis of partial monosomy 10p and trisomy 10q resulting from familial pericentric inversion (10): a first case report in Chinese population.

BACKGROUND: Chromosome aberrations of 10p monosomy and 10q trisomy resulting from parental pericentric inversion 10 are extremely rare, and to date, very few reports have been published on the matter. CASE PRESENTATION: A 30-year-old pregnant woman with recurrent pregnancy loss is enrolled in this research. In this pregnancy, spontaneous abortion occurred in the first trimester of her pregnancy. Chromosomal microarray analysis of the abortion tissue showed a partial 10p monosomy (arr[GRCh37] 10p15.3p11.21(100,047_34,848,853) × 1) and a duplication of 10q (arr[GRCh37] 10q26.13q26.3(126,093,990_135,426,386) × 3). Further parental karyotype analysis indicated that the chromosomal abnormalities in the fetus was resulted from paternal pericenric inversion inv(10)(p11.21q26.13). This study presents the first case of a large deletion of 10p combined with 10q trisomy, resulting in pregnancy loss. Of these two manifestations, the large deletion of chromosome 10p may be the primary reason for spontaneous abortion in this subject. CONCLUSIONS: This study presents the first case of partial 10p monosomy associated with 10q trisomy in Chinese population. It provides more information on the chromosome aberration of 10p monosomy and 10q trisomy and further strengthens the application value of microarray in the molecular etiological diagnosis of recurrent spontaneous abortion.

USP35 is a Potential Immunosuppressive Factor in Skin Cutaneous Melanoma.

Background: As one of the most immunogenic malignancies, skin cutaneous melanoma (SKCM) is mainly characterized by a high prevalence in immune-compromised patients and a brisk lymphocyte infiltration in the tumor microenvironment (TME). However, to date, studies on deubiquitination in SKCM are still very limited. Methods: Public data with regard to this study in SKCM patients were acquired from The Cancer Genome Atlas (TCGA) and the Gene-Expression Omnibus (GEO) databases. We stratified TCGA-SKCM cases using consensus clustering and identified independent prognostic factors in deubiquitinating enzymes encoding genes (DECGs) by LASSO-Cox analysis. USP35 transcriptome level was examined using public data and validated by Immunohistochemical (IHC) staining at the protein level. Enrichment analysis was used to explore the potential functions of USP35, and the TISCH database, providing further evidence at the single-cell level. The CIBERSORT algorithm was used to assess the relationship between USP35 and the immune microenvironment, and IHC was used to further evaluate the relationship between USP35 and immunotherapy response. Finally, we used the cBioPortal and the Methsurv database to analyze the significance of genomic alterations of USP35 in melanoma. Results: Our results showed that DECGs can be effectively used to stratify SKCM patients, suggesting their potential significance in the development of SKCM. Furthermore, USP35 overexpression was significantly associated with an unfavorable prognosis. We further revealed that USP35 may be involved in the activation of TORC1 signaling. Most importantly, USP35 was found to be significantly associated with an immunosuppressive TME, both in terms of negative correlation with the abundance of infiltrating CD8+ T cells and in terms of the fact that patients with high USP35 expression may benefit less from immunotherapy than those with low USP35 expression. Conclusion: Deubiquitinating enzymes are of great importance in the diagnosis and treatment of SKCM, and USP35 is an extremely promising target for immunotherapy.

Identification of a Rare Variant of c.1777G>A (p.G593S) in the COL1A1 Gene as the Etiology of Recurrent Osteogenesis Imperfecta by Whole-Exome Sequencing.

Background: Osteogenesis imperfecta (OI) is a rare heterogeneous disorder typically featured by fragile bones and susceptibility to fracture. The aim of the present study was to explore the genetic etiology of familial recurrent OI and the genotype-phenotype correlation. Methods: Karyotyping, chromosomal microarray analysis, and whole-exome sequencing (WES) were performed to determine the genetic etiology of OI in the enrolled family. Western blotting analysis was carried out using the fetal skin tissue for type I collagen production analysis. Results: At the first pregnancy, a c.1777G>A mutation in the COL1A1 gene was detected in the fetus who exhibited skeletal dysplasia. In this second pregnancy, severe fetal skeletal dysplasia was also presented without significant chromosomal abnormality detected by karyotype and chromosomal microarray analysis in the fetus. Further WES results demonstrated a de novo missense mutation of c.1777G>A (p.G593S) in the fetus, which was classified as a pathogenic variant according to the ACMG guidelines. The recurrent mutation in the two fetuses hinted at the possible existence of gonadal mosaicism in the parents, while no mutation in the COL1A1 gene was identified in the DNA from the father's sperm. In addition, Western blot results demonstrated no reduced type I procollagen production in the affected fetus compared with the age-matched controls. Conclusions: To the best of our knowledge, this is the first study that identified a rare variant of c.1777G>A in the COL1A1 gene that led to recurrent OI in the Chinese population. Additionally, we believe that this rare variant of c.1777G>A in the COL1A1 gene will lead to OI type II. The results of the present study further verify the application value of WES in identifying fetuses with ultrasound anomalies.

An Analysis Regarding the Association Between Connexins and Colorectal Cancer (CRC) Tumor Microenvironment.

Background: Gap junctions, as one of the major ways to maintain social connections between cells, are now considered as one of the potential regulators of tumor metastasis. However, to date, studies on the relationship between gap junctions and colorectal cancer (CRC) are limited. Methods: We synthesized connexins-coding gene expression data from public Gene Expression Omnibus (GEO) and The Cancer Genome Atlas (TCGA) databases. Bioinformatics analysis was performed using R software and several database resources such as MEXPRESS database, Gene Set Cancer Analysis (GSCA) database, Human Protein Atlas (HPA) database, Tumor Immune Single Cell Hub (TISCH) database, Search Tool for Retrieval of Gene Interaction Relationships (STRING), and Cytoscape software, etc., to investigate the biological mechanisms that may be involved in connexins. Immunofluorescence and immunohistochemical staining were used to validate the expression and localization of GJA4. Results: We found that CRC patients can be divided into two connexin clusters and that patients in cluster C1 had shorter survival than in cluster C2. The infiltration of M1 macrophages and NK cells was lower in cluster C1, while the levels of M2 macrophages and immune checkpoints were higher, indicating an immunosuppressed state in cluster C1. In addition, the epithelial-mesenchymal transition (EMT) phenotype was significantly activated in cluster C1. We observed that GJA4 was up-regulated in colorectal cancer tissues, which was related to poor prognosis. It was mainly expressed in fibroblasts, but the expression levels in normal intestinal epithelial cells were low. Finally, we found that GJA4 was associated with M2 macrophages and may be a potential immunosuppressive factor. Conclusion: We found that there is a significant correlation between abnormal connexins expression and patients' prognosis, and connexins play an important role in stromal-tumor interactions. Connexins, especially GJA4, can help enhance our understanding of tumor microenvironment (TME) and may guide more effective immunotherapeutic strategies.

Case Report: Prenatal Whole-Exome Sequencing Identified a Novel Nonsense Mutation of the KCNH2 Gene in a Fetus With Familial 2q14.2 Duplication.

Background: Pathogenic mutations in the KCNH2 gene were associated with long QT syndrome 2 (LQT2), which typically manifest in a prolonged QT interval and may lead to recurrent syncopes, seizure, or sudden death. Limited reports indicated that the KCNH2 mutations would result in LQT2 combined with tetralogy of fallot. Our goal was to present an additional case of LQT2 combined with the tetralogy of fallot in a fetus with a novel KCNH2 mutation. Case presentation: Enrolled in this study was a 23-year-old pregnant woman from Quanzhou Fujian province, China. In her pregnancy, fetal ultrasound anomalies were identified, including tetralogy of fallot, coronary sinus enlargement, and persistent left superior vena cava. No chromosomal abnormality was detected by fetal karyotype analysis. However, 238.1-kb duplication in the 2q14.2 region containing the GLI2 gene was observed in the fetus by chromosomal array analysis, which was inherited from the mother with normal clinical features and interpreted as a variant of uncertain significance (VOUS). Furthermore, whole-exome sequencing (WES) detection identified a novel nonsense c.1907C > G (p.S636*) mutation in the KCNH2 gene in the fetus, and it was classified as a likely pathogenic variant, according to the ACMG guidelines. Parental verification analysis indicated that c.1907C > G (p.S636*) mutation was inherited from the mother. Conclusion: In this study, we believe that 2q14.2 duplication may not be the reason for fetal heart defects; moreover, we described an additional case with KCNH2 gene mutation, which may lead to LQTS and be associated with congenital heart defects. In addition, our study further confirms the application value of the WES technology in prenatal genetic etiology diagnosis of fetuses with structural anomalies and unexplained structural variants.