OBJECTIVE: To explore the mechanism of BNIP3-mediated mitophagy in m.3635G>A related Leber's hereditary optic neuropathy (LHON). METHODS: A trans-mitochondrial cybrid cell line derived from a Chinese LHON patient carrying the m.3635G>A, diagnosed at the Eye Hospital of Wenzhou Medical University in September 2013, was selected as the study subject. A trans-mitochondrial cybrid cell line from a healthy control with an identical mitochondrial background was included as a control. Immunofluorescence, real-time quantitative PCR (RT-qPCR), and Western blotting were employed to assess the expression of autophagy-related proteins, aiming to explore the role of BNIP3-mediated mitophagy in m.3635G>A related LHON. This study was approved by the Medical Ethics Committee of the Eye Hospital of Wenzhou Medical University (Ethics No. 2023-J-096). RESULTS: Compared with the control group, the protein expression levels of autophagy-related markers LC3 (LC3-II/LC3-I) and LAMP1 were significantly reduced in the variant group (P < 0.05). Additionally, the protein levels of macroautophagy-related proteins ATG12, ATG7, and ATG5 were also significantly decreased (P < 0.05). Compared with the control cells, the mRNA and protein expression levels of mitophagy-associated protein BNIP3 were significantly reduced in the cells of the variant group (P < 0.05). Compared with the control group, both mRNA and protein expression levels of the mitophagy-related protein BNIP3 were significantly reduced in the variant group (P < 0.05). CONCLUSION The m.3635G>A inhibits BNIP3-mediated mitophagy, thereby contributing to the pathogenesis of LHON.
Humans ; Proto-Oncogene Proteins/metabolism* ; Mitophagy/genetics* ; Membrane Proteins/metabolism* ; Optic Atrophy, Hereditary, Leber/metabolism* ; Mitochondria/metabolism* ; Cell Line ; Male

OBJECTIVE: To explore the genotype-phenotype relationship in a child with Opitz G/BBB syndrome (OS) with mild clinical phenotype. METHODS: A child with motor developmental delay as the initial symptom admitted to Xi'an Children's Hospital on June 10, 2021 was selected for this study. Clinical data were collected, and peripheral blood samples were obtained from the child and his mother. Whole exome sequencing (WES) was performed to identify genetic variant in the child. Candidate variant were verified by Sanger sequencing to assess inheritance patterns and pathogenicity. Real-time fluorescence quantitative PCR (RT-qPCR) and Western blot (WB) analyses were conducted to evaluate the effects of the variant on mRNA and protein expression, respectively, using recombinant expression plasmids generated in vitro. This study was approved by the Medical Ethics Committee of Xi'an Children's Hospital (Ethics No. 20240045). RESULTS: The child, a 9-month-and-7-day-old boy, presented with a low nasal bridge, hypertelorism, and difficulty sitting independently. Echocardiography revealed an atrial septal defect. WES identified a homozygous variant in the MIDI gene, c.1483C>T (p.R495X), which was confirmed by Sanger sequencing and found to be inherited from the mother.Recombinant expression plasmids were successfully constructed. RT-qPCR analysis showed that the variant significantly reduced MIDI gene mRNA expression, while WB results indicated that the variant led to the production of a truncated protein. CONCLUSION The mild clinical phenotype of OS in this child may be attributed to the mRNA degradation escape mechanism induced by the nonsense variant c.1483C>T (p.R495X) in the MIDI gene. These findings provide valuable diagnostic insights for this pedigree and contribute to the understanding of the genotype-phenotype correlation in OS.
Humans ; Male ; Infant ; Transcription Factors/metabolism* ; Microtubule Proteins/genetics* ; Craniosynostoses/genetics* ; Hypospadias/genetics* ; Codon, Nonsense/genetics* ; RNA, Messenger/metabolism* ; Female ; RNA Stability/genetics* ; Phenotype ; Nuclear Proteins/genetics* ; Ubiquitin-Protein Ligases ; Esophagus/abnormalities* ; Hypertelorism

Global developmental delay (GDD) and intellectual disability (ID) refer to deficits in cognitive and adaptive functioning that arise during the developmental period. GDD/ID is often accompanied by complex developmental abnormalities, with congenital craniofacial malformations being among the most common, such as craniosynostosis, cleft lip and palate, and congenital tooth agenesis. However, the underlying mechanisms of GDD/ID associated with congenital craniofacial malformations remain unclear. With the increasing number of reported genetic syndromes, genetic factors are emerging as key contributors to the concurrent abnormalities in brain and craniofacial development. Studies have identified Wnt, SHH, FGF, and BMP as classical regulatory molecules in craniofacial development, and their roles have also been closely linked to various stages of brain development. This review focuses on the regulatory roles of Wnt, SHH, FGF, and BMP signaling pathways in brain and craniofacial development, as well as the pathogenic mechanisms underlying their association with GDD/ID and craniofacial malformations. The aim is to provide new insights into the etiology of GDD/ID combined with congenital craniofacial malformations.
Humans ; Craniofacial Abnormalities/complications* ; Signal Transduction ; Developmental Disabilities/metabolism* ; Intellectual Disability/complications* ; Animals ; Hedgehog Proteins/genetics* ; Fibroblast Growth Factors/genetics*

OBJECTIVE: To elucidate the clinical and genetic characteristics of familial cases with Glucose transporter type 1 deficiency syndrome (Glut1DS). METHODS: A survey of family history was conducted on children (proband) with Glut1DS who had visited Peking University First Hospital between November 2008 and April 2024 by focusing on the clinical manifestations of family members. Peripheral venous blood (2 mL) was collected from the pediatric patients and their parents. Genomic DNA was extracted and sequenced subsequently. Sanger sequencing was performed to validate the identified variant sites of the SLC2A1 gene in the probands and their family members. The pathogenicity of suspected variants was analyzed according to the 2015 American College of Medical Genetics and Genomics (ACMG) Standards and Guidelines for the Interpretation of Sequence Variants. The clinical features, auxiliary examinations, and mutational characteristics of family members with SLC2A1 variants were analyzed. This study has been approved by the Clinical Research Ethics Committee of Peking University First Hospital (Ethics No. 2021 Research 332). RESULTS: Among 87 cases with Glut1DS, 10 families with autosomal dominate inherited cases were identified, accounting for 11.0% of the cases. Of the 11 children, 8 were boys and 3 were girls. The onset of the disease had ranged from 3 months to 120 months (median 6 months), with 4 cases of early-onset classic type, 2 cases of late-onset classic type, and 5 cases of non-classic type. Six children had seizures, and 7 exhibited movement disorders. Seven children underwent developmental assessment, of which 3 had mild developmental delay, 2 were borderline, and 2 were normal. Nine children underwent lumbar puncture. The cerebrospinal fluid glucose levels ranged from 1.45 to 2.25 mmol/L (median 1.86 mmol/L), and the cerebrospinal fluid to blood glucose ratios ranged from 0.29 to 0.44 (median 0.35). Among the 8 fathers with SLC2A1 gene variants, 4 were asymptomatic, 2 developed paroxysmal exercise-induced movement disorders (PED) in childhood and adulthood, respectively. 1 had poor memory since childhood, 1 developed migraines during adolescence, and his sister was an asymptomatic carrier. The father with childhood-onset PED had a cerebrospinal fluid test with CSF glucose of 1.85 mmol/L. Of the 3 mothers with SLC2A1 gene mutations, 1 was an asymptomatic carrier; 2 developed PED in childhood and after the age of 20, respectively. The mother who developed PED in childhood also had psychomotor developmental delay. Genetic testing results revealed that among 10 families, 8 carried missense variants, 1 carried a nonsense variant, and 1 carried a small fragment insertion leading to a frameshift variant. Among the 11 cases, SLC2A1 gene variants in 8 children were inherited from their fathers, while in 3 cases, the variants were inherited from their mothers. The pathogenicity of the genetic variants was evaluated according to the Standards and Guidelines for the Interpretation of Sequence Variants published by the ACMG. Among the 8 variants identified in the 10 families, 4 were classified as pathogenic variants, 1 as likely pathogenic, and 3 as variants of uncertain significance (VUS). Four variant sites, including c.204_205insTCTC (p.V69fs), c.412G>C (p.G138R), c.431T>G (p.V144G), and c.875A>G (p.Y292C), were not previously reported in the literature. Among these, the latter three were categorized as VUS. CONCLUSION Familial Glut1DS account for 11.0% of the cases in China, with the majority of SLC2A1 gene variants inherited from the fathers, predominantly missense mutations, and with an autosomal dominant inheritance pattern. Probands tend to have earlier onset and more severe symptoms than their parents, who often present with mild or no symptoms.
Humans ; Male ; Female ; Glucose Transporter Type 1/deficiency* ; Monosaccharide Transport Proteins/deficiency* ; Child ; Child, Preschool ; Carbohydrate Metabolism, Inborn Errors/genetics* ; Mutation ; Infant ; Pedigree ; Adolescent ; Adult

The SMPX (small muscle protein X-linked) gene encodes a small-molecular-weight protein that is mainly expressed in skeletal and cardiac muscles and is involved in cytoskeletal dynamics and mechanical stress responses. In recent years, missense variants of the SMPX gene have been identified as the cause of a novel X-linked distal myopathy (Distal myopathy 7). This article has systematically reviewed the molecular functions, variant types, and pathological mechanisms of the SMPX gene by integrating its clinical classification, molecular pathological evidence, and experimental model data, and revealed its pathgenetic mechanism through protein aggregation, dynamic dysregulation of stress granules, abnormal Rac1/p38 signaling pathways, and future research directions.
Humans ; Mutation ; Muscle Proteins/metabolism* ; Deafness/genetics* ; Animals ; Muscular Diseases/genetics* ; Genes, X-Linked

OBJECTIVE: To explore the clinical phenotypes and genetic characteristics of a child with Acid-labile subunit deficiency (ALS). METHODS: A male child diagnosed with ALS at Dongguan Maternal and Child Health Care Hospital in March 2021 was selected as the study subject. Clinical data of his family was collected. Peripheral blood samples were collected from the child and his parents. Following extraction of genomic DNA, whole-exome sequencing (WES) was carried out, and Sanger sequencing was used for family verification of candidate variants. Based on guidelines from the American College of Medical Genetics and Genomics (ACMG), the pathogenicity of the candidate variant was classified. This study was approved by the Medical Ethics Committee of the hospital (Ethics No.: 2020-6). RESULTS: The patient, a 5-year-and-7-month-old boy, presented with short stature and delayed bone age. Endocrine examinations showed decreased serum concentrations of insulin-like growth factor-1 (IGF-1) and IGF binding protein-3 (IGFBP3). WES revealed that he has harbored compound heterozygous variants of the IGFALS gene, namely c.741_742del, p.Y248Pfs83 and c.272del, p.P91Rfs31. Sanger sequencing verified that the variants were inherited from his father and mother, respectively. According to the ACMG guidelines, c.741_742del, p.Y248Pfs83 and c.272del, p.P91Rfs31 variants were classified as likely pathogenic (PVS1+PM2_supporting). Based on the pre-set literature search strategy, 11 research literature on ALS were retrieved, which involved a total of 33 families and 62 patients. Combined with the patient in this study, 31 IGFALS gene variants were identified among the 63 patients, which mainly consisted of missense variants (20 types), with variant sites concentrated in exon 2. The main clinical features were short stature in conjunct with delayed puberty, with a significant genotype-phenotype correlation. CONCLUSION The IGFALS gene variants NM_004970.2: c.741_742del, p.Y248Pfs83 and c.272del, p.P91Rfs31 may be the genetic etiology in this family. This study has expanded the variant spectrum of the IGFALS gene and provided valuable information for the diagnosis, genetic counseling and clinical treatment of the disease.
Humans ; Male ; Phenotype ; Child, Preschool ; Carrier Proteins/genetics* ; Glycoproteins/deficiency* ; Exome Sequencing ; Female ; Mutation ; Insulin-Like Growth Factor I/metabolism* ; Growth Disorders/genetics*

OBJECTIVES: The neurotoxicity of carbon monoxide (CO) to the central nervous system is a key pathogenesis of delayed encephalopathy after acute carbon monoxide poisoning (DEACMP). Our previous study found that retinoic acid (RA) can suppress the neurotoxic effects of CO. This study further explores, in vivo and in vitro, the molecular mechanisms by which RA alleviates CO-induced central nervous system damage. METHODS: A cytotoxic model was established using the mouse hippocampal neuronal cell line HT22 and primary oligodendrocytes exposed to CO, and a DEACMP animal model was established in adult Kunming mice. Cell viability and apoptosis of hippocampal neurons and oligodendrocytes were assessed using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and Annexin V/propidium iodide (PI) double staining. The transcriptional and protein expression of each gene was detected using real-time fluorescence quantitative PCR (RT-qPCR) and Western blotting. Long noncoding RNA (lncRNA) SNHG15 and LINGO-1 were knocked down or overexpressed to observe changes in neurons and oligodendrocytes. In DEACMP mice, SNHG15 or LINGO-1 were knocked down to assess changes in central nervous tissue and downstream protein expression. RESULTS: RA at 10 and 20 μmol/L significantly reversed CO-induced apoptosis of hippocampal neurons and oligodendrocytes, downregulation of SNHG15 and LINGO-1, and upregulation of brain-derived neurotrophic factor (BDNF) and tyrosine kinase receptor B (TrkB) (all P<0.05). Overexpression of SNHG15 or LINGO-1 weakened the protective effect of RA against CO-induced cytotoxicity (all P<0.05). Knockdown of SNHG15 or LINGO-1 alleviated CO-induced apoptosis of hippocampal neurons and oligodendrocytes and upregulated BDNF and TrkB expression levels (all P<0.05). Experiments in DEACMP model mice showed that knockdown of SNHG15 or LINGO-1 mitigated central nervous system injury in DEACMP (all P<0.05). CONCLUSIONS RA alleviates CO-induced apoptosis of hippocampal neurons and oligodendrocytes, thereby reducing central nervous system injury and exerting neuroprotective effects. LncRNA SNHG15 and LINGO-1 are key molecules mediating RA-induced inhibition of neuronal apoptosis and are associated with the BDNF/TrkB pathway. These findings provide a theoretical framework for optimizing the clinical treatment of DEACMP and lay an experimental foundation for elucidating its molecular mechanisms.
Animals ; RNA, Long Noncoding/physiology* ; Brain-Derived Neurotrophic Factor/genetics* ; Carbon Monoxide Poisoning/complications* ; Mice ; Tretinoin/pharmacology* ; Nerve Tissue Proteins/metabolism* ; Membrane Proteins/metabolism* ; Apoptosis/drug effects* ; Hippocampus/cytology* ; Receptor, trkB/metabolism* ; Neurons/drug effects* ; Male ; Brain Diseases/etiology* ; Oligodendroglia/drug effects* ; Signal Transduction ; Cell Line

BACKGROUND: T-cell lymphoblastic lymphoma/acute lymphoblastic leukemia (T-LBL/ALL) is an aggressive form of hematological malignancy associated with poor prognosis in adult patients. Histone deacetylases (HDACs) are aberrantly expressed in T-LBL/ALL and are considered potential therapeutic targets. Here, we investigated the antitumor effect of a novel HDAC inhibitor, chidamide, on T-LBL/ALL. METHODS: HDAC1, HDAC2 and HDAC3 levels in T-LBL/ALL cell lines and patient samples were compared with those in normal controls. Flow cytometry, transmission electron microscopy, and lactate dehydrogenase release assays were conducted in Jurkat and MOLT-4 cells to assess apoptosis and pyroptosis. A specific forkhead box O1 (FOXO1) inhibitor was used to rescue pyroptosis and upregulated gasdermin E (GSDME) expression caused by chidamide treatment. The role of the FOXO1 transcription factor was evaluated by dual-luciferase reporter and chromatin immunoprecipitation assays. The efficacy of chidamide in vivo was evaluated in a xenograft mouse. RESULTS: The expression of HDAC1, HDAC2 and HDAC3 was significantly upregulated in T-LBL/ALL. Cell viability was obviously inhibited after chidamide treatment. Pyroptosis, characterized by cell swelling, pore formation on the plasma membrane and lactate dehydrogenase leakage, was identified as a new mechanism of chidamide treatment. Chidamide triggered pyroptosis through caspase 3 activation and GSDME transcriptional upregulation. Chromatin immunoprecipitation assays confirmed that chidamide led to the increased transcription of GSDME through a more relaxed chromatin structure at the promoter and the upregulation of FOXO1 expression. Moreover, we identified the therapeutic effect of chidamide in vivo . CONCLUSIONS This study suggested that chidamide exerts an antitumor effect on T-LBL/ALL and promotes a more inflammatory form of cell death via the FOXO1/GSDME axis, which provides a novel choice of targeted therapy for patients with T-LBL/ALL.
Humans ; Pyroptosis/drug effects* ; Forkhead Box Protein O1/genetics* ; Aminopyridines/pharmacology* ; Animals ; Mice ; Benzamides/pharmacology* ; Cell Line, Tumor ; Precursor T-Cell Lymphoblastic Leukemia-Lymphoma/drug therapy* ; Phosphate-Binding Proteins/metabolism* ; Histone Deacetylase Inhibitors/pharmacology* ; Jurkat Cells ; Histone Deacetylases/metabolism* ; Apoptosis/drug effects* ; Gasdermins

BACKGROUND: Enzalutamide, a second-generation androgen receptor (AR) pathway inhibitor, is widely used in the treatment of castration-resistant prostate cancer. However, after a period of enzalutamide treatment, patients inevitably develop drug resistance. In this study, we characterized leucine-rich repeated G-protein-coupled receptor 5 (LGR5) and explored its potential therapeutic value in prostate cancer. METHODS: A total of 142 pairs of tumor and adjacent formalin-fixed paraf-fin-embedded tissue samples from patients with prostate cancer were collected from the Pathology Department at Sun Yat-sen Memorial Hos-pital. LGR5 was screened by sequencing data of enzalutamide-resistant cell lines combined with sequencing data of lesions with different Gleason scores from the same patients. The biological function of LGR5 and its effect on enzalutamide resistance were investigated in vitro and in vivo . Glutathione-S-transferase (GST) pull-down, coimmunoprecipitation, Western blotting, and immunofluorescence assays were used to explore the specific binding mechanism of LGR5 and related pathway changes. RESULTS: LGR5 was significantly upregulated in prostate cancer and negatively correlated with poor patient prognosis. Overexpression of LGR5 promoted the malignant progression of prostate cancer and reduced sensitivity to enzalutamide in vitro and in vivo . LGR5 promoted the phosphorylation of glycogen synthase kinase-3β (GSK-3β) by binding heat shock protein 90,000 alpha B1 (HSP90AB1) and mediated the activation of the Wingless/integrated (WNT)/β-catenin signaling pathway. The increased β-catenin in the cytoplasm entered the nucleus and bound to the nuclear AR, promoting the transcription level of AR, which led to the enhanced tolerance of prostate cancer to enzalutamide. Reducing HSP90AB1 binding to LGR5 significantly enhanced sensitivity to enzalutamide. CONCLUSIONS LGR5 directly binds to HSP90AB1 and mediates GSK-3β phosphorylation, promoting AR expression by regulating the WNT/β-catenin signaling pathway, thereby conferring resistance to enzalutamide treatment in prostate cancer.
Male ; Humans ; Phenylthiohydantoin/pharmacology* ; Benzamides ; Receptors, G-Protein-Coupled/genetics* ; Nitriles ; Cell Line, Tumor ; HSP90 Heat-Shock Proteins/metabolism* ; Drug Resistance, Neoplasm/genetics* ; Prostatic Neoplasms/drug therapy* ; beta Catenin/metabolism* ; Receptors, Androgen/genetics* ; Animals ; Mice ; Wnt Signaling Pathway/physiology*

BACKGROUND: Temozolomide (TMZ) resistance is a significant challenge in treating glioblastoma (GBM). Collagen remodeling has been shown to be a critical factor for therapy resistance in other cancers. This study aimed to investigate the mechanism of TMZ chemoresistance by GBM cells reprogramming collagens. METHODS: Key extracellular matrix components, including collagens, were examined in paired primary and recurrent GBM samples as well as in TMZ-treated spontaneous and grafted GBM murine models. Human GBM cell lines (U251, TS667) and mouse primary GBM cells were used for in vitro studies. RNA-sequencing analysis, chromatin immunoprecipitation, immunoprecipitation-mass spectrometry, and co-immunoprecipitation assays were conducted to explore the mechanisms involved in collagen accumulation. A series of in vitro and in vivo experiments were designed to assess the role of the collagen regulators prolyl 4-hydroxylase subunit alpha 1 (P4HA1) and yes-associated protein (YAP) in sensitizing GBM cells to TMZ. RESULTS: This study revealed that TMZ exposure significantly elevated collagen type I (COL I) expression in both GBM patients and murine models. Collagen accumulation sustained GBM cell survival under TMZ-induced stress, contributing to enhanced TMZ resistance. Mechanistically, P4HA1 directly binded to and hydroxylated YAP, preventing ubiquitination-mediated YAP degradation. Stabilized YAP robustly drove collagen type I alpha 1 ( COL1A1) transcription, leading to increased collagen deposition. Disruption of the P4HA1-YAP axis effectively reduced COL I deposition, sensitized GBM cells to TMZ, and significantly improved mouse survival. CONCLUSION P4HA1 maintained YAP-mediated COL1A1 transcription, leading to collagen accumulation and promoting chemoresistance in GBM.
Temozolomide ; Humans ; Glioblastoma/drug therapy* ; Animals ; Mice ; Cell Line, Tumor ; Drug Resistance, Neoplasm/genetics* ; YAP-Signaling Proteins ; Hydroxylation ; Dacarbazine/pharmacology* ; Adaptor Proteins, Signal Transducing/metabolism* ; Transcription Factors/metabolism* ; Collagen/biosynthesis* ; Collagen Type I/metabolism* ; Prolyl Hydroxylases/metabolism* ; Antineoplastic Agents, Alkylating/therapeutic use*