Abnormal Maturation Research Articles

Changes in the Expression of DNMTs Before and After Treatment with Methotrexate (MTX)/Mercaptopurine (6-MP) in B-Cell ALL Children

Background: DNA methylation is catalyzed by DNA methyltransferases (DNMTs) which are encoded by DNMT1, DNMT3A, and DNMT3B. DNMTs play a major role in the abnormal methylation of tumor suppressors and cancer-related genes. Herein, this study explored the expression profile of DNMTs in pediatric patients with B-cell acute lymphoblastic leukemia (ALL), before and after methotrexate (MTX)/mercaptopurine (6-MP) treatment.
 Materials and Methods: This before-after prospective study included 30 matched children in sex and age (20 children with B-cell ALL and 10 healthy children used as a control or calibrator group). The expression profile of DNMTs was assessed at two-time points; at the diagnosis time and after MTX/6-MP treatment in the consolidation-maintenance phase of therapy. Notable, all pediatric patients included in this study continued the therapy without adverse events, except two children who were excluded from the study.
 Results: The average age of the patient group was 7.1 ± 1.3 years (in the range of 4-9 years), and the average age of the control group was 8.3 ± 1.7 years (6-10 years). The expression profile of DNMTs in B-cell ALL children was obtained completely different from that in the healthy group. After MTX/6-MP treatment of B-cell ALL children, the expression levels of DNMT1 and 3A were increased (p <0.01 & 0.04, respectively), and the expression level of DNMT3B was decreased (p <0.01), significantly.
 Conclusions: In ALL, the expression profile of DNMTs would be changed whereby contribute to abnormal growth and maturation capacity of leukemic stem cells and MTX/6-MP treatment could reverse this profile from a cancerous phenotype to the normal one.

HSPA9/mortalin inhibition disrupts erythroid maturation through a TP53-dependent mechanism in human CD34+ hematopoietic progenitor cells.

Myelodysplastic syndromes (MDS) are a heterogeneous group of clonal hematopoietic stem cell malignancies characterized by abnormal hematopoietic cell maturation, increased apoptosis of bone marrow cells, and anemia. They are the most common myeloid blood cancers in American adults. The full complement of gene mutations that contribute to the phenotypes or clinical symptoms in MDS is not fully understood. Around 10%-25% of MDS patients harbor an interstitial heterozygous deletion on the long arm of chromosome 5 [del(5q)], creating haploinsufficiency for a large set of genes, including HSPA9. The HSPA9 gene encodes for the protein mortalin, a highly conserved heat shock protein predominantly localized in mitochondria. Our prior study showed that knockdown of HSPA9 induces TP53-dependent apoptosis in human CD34+hematopoietic progenitor cells. In this study, we explored the role of HSPA9 in regulating erythroid maturation using human CD34+cells. We inhibited the expression of HSPA9 using gene knockdown and pharmacological inhibition and found that inhibition of HSPA9 disrupted erythroid maturation as well as increased expression of p53 in CD34+cells. To test whether the molecular mechanism of HSPA9 regulating erythroid maturation is TP53-dependent, we knocked downHSPA9 and TP53 individually or in combination in human CD34+cells. We found that the knockdown of TP53 partially rescued the erythroid maturation defect induced by HSPA9 knockdown, suggesting that the defect in cells with reduced HSPA9 expression is TP53-dependent. Collectively, these findings indicate that reduced levels of HSPA9 may contribute to the anemia observed in del(5q)-associated MDS patients due to the activation of TP53.

Identification novel mutations and phenotypic spectrum expanding in PATL2 in infertile women with IVF/ICSI failure.

Abnormalities in oocyte maturation, fertilization, and early embryonic development are major causes of primary infertility in women who are undergoing IVF/ICSI attempts. Although many genetic factors responsible for these abnormal phenotypes have been identified, there are more additional pathogenic genes and variants yet to be discovered. Previous studies confirmed that bi-allelic PATL2 deficiency is an important factor for female infertility. In this study, 935 infertile patients with IVF/ICSI failure were selected for whole-exome sequencing, and 18 probands carrying PATL2 variants with a recessive inheritance pattern were identified. We estimated that the prevalence contributed by PATL2 was 1.93% (18/935) in our study cohort. 15 novel variants were found in those families, including c.1093C > T, c.1609dupA, c.1204C > T, c.643dupG, c.877-2A > G, c.1228C > G, c.925G > A, c.958G > A, c.4A > G, c.1258T > C, c.1337G > A, c.1264dupA, c.88G > T, c.1065-2A > G, and c.1271T > C. The amino acids altered by the corresponding variants were highly conserved in mammals, and in silico analysis and 3D molecular modeling suggested that the PATL2 mutants impaired the physiologic function of the resulting proteins. Diverse clinical phenotypes, including oocyte maturation defect, fertilization failure, and early embryonic arrest might result from different variants of PATL2. These results expand the spectrum of PATL2 variants and provide an important reference for genetic counseling for female infertility, and they increase our understanding of the mechanisms of oocyte maturation arrest caused by PATL2 deficiency.

Mechanistic interplay of dual environmental stressors: Bisphenol-A and cadmium-induced ovarian follicular damage and hepatocyte dysfunction in vivo

This study investigates the individual and combined toxic effects of Bisphenol A (BPA) and Cadmium (Cd) in zebrafish, recognizing the complex mixture of pollutants organisms encounter in their natural environment. Examining developmental, neurobehavioral, reproductive, and physiological aspects, the study reveals significant adverse effects, particularly in combined exposures. Zebrafish embryos exposed to BPA + Cd exhibit synergistically increased mortality, delayed hatching, and morphological abnormalities, emphasizing the heightened toxicity of the combination. Prolonged exposure until 10 days post-fertilization underscores enduring effects on embryonic development. BPA and Cd induce oxidative stress, as evidenced by increased production of reactive oxygen species and lipid peroxidation. This oxidative stress disrupts cellular functions, affecting lipid metabolism and immune response. Adult zebrafish exposed to BPA and Cd for 40 days display compromised neurobehavioral functions, altered antioxidant defenses, and increased oxidative stress, suggesting potential neurotoxicity. Additionally, disruptions in ovarian follicle maturation and skeletal abnormalities indicate reproductive and skeletal impacts. Histological analysis reveals significant liver damage, emphasizing the synergistic hepatotoxicity of BPA and Cd. Molecular assessments further demonstrate compromised cellular defense mechanisms, synaptic function, and elevated cellular stress and inflammation-related gene expression in response to combined exposures. Bioaccumulation analysis highlights differential tissue accumulation patterns. In conclusion, this study provides comprehensive insights into the multifaceted toxicological effects of BPA and Cd in zebrafish, raising concerns about potential adverse impacts on environmental ecosystems and human health.

Inhibition of HSP90AA1 induces abnormalities in bovine oocyte maturation and embryonic development.

HSP90AA1 is a ubiquitous molecular chaperone that can resist cellular stress, such as oxidative stress and apoptosis, and mediate the efficacy and protein folding of normal cells during heat stress, as well as many other functions. This study further reveals the role of HSP90AA1 in bovine oocyte maturation and early embryonic development. HSP90AA1, a highly abundant and ubiquitous molecular chaperone, plays important roles in various cellular processes including cell cycle control, cell survival, and hormone signaling pathways. In this study, we investigated the functions of HSP90AA1 in bovine oocyte and early embryo development. We found that HSP90AA1 was expressed at all stages of development, but was mainly located in the cytoplasm, with a small amount distributed in the nucleus. We then evaluated the effect of HSP90AA1 on the in vitro maturation of bovine oocytes using tanespimycin (17-AAG), a highly selective inhibitor of HSP90AA1. The results showed that inhibition of HSP90AA1 decreased nuclear and cytoplasmic maturation of oocytes, disrupted spindle assembly and chromosome distribution, significantly increased acetylation levels of α-tubulin in oocytes and affected epigenetic modifications (H3K27me3 and H3K27ac). In addition, H3K9me3 was increased at various stages during early embryo development. Finally, the impact of HSP90AA1 on early embryo development was explored. The results showed that inhibition of HSP90AA1 reduced the cleavage and blastocyst formation rates, while increasing the fragmentation rate and decreasing blastocyst quality. In conclusion, HSP90AA1 plays a crucial role in bovine oocyte maturation as well as early embryo development.

Brain Volumes in Opsoclonus-Myoclonus Ataxia Syndrome: A Longitudinal Study.

Little is known about the longitudinal trajectory of brain growth in children with opsoclonus-myoclonus ataxia syndrome. We performed a longitudinal evaluation of brain volumes in pediatric opsoclonus-myoclonus ataxia syndrome patients compared with age- and sex-matched healthy children. This longitudinal case-control study included brain magnetic resonance imaging (MRI) scans from consecutive pediatric opsoclonus-myoclonus ataxia syndrome patients (2009-2020) and age- and sex-matched healthy control children. FreeSurfer analysis provided automatic volumetry of the brain. Paired t tests were performed on the curvature of growth trajectories, with Bonferroni correction. A total of 14 opsoclonus-myoclonus ataxia syndrome patients (12 female) and 474 healthy control children (406 female) were included. Curvature of the growth trajectories of the cerebral white and gray matter, cerebellar white and gray matter, and brainstem differed significantly between opsoclonus-myoclonus ataxia syndrome patients and healthy control children (cerebral white matter, P = .01; cerebral gray matter, P = .01; cerebellar white matter, P < .001; cerebellar gray matter, P = .049; brainstem, P < .01). We found abnormal brain maturation in the supratentorial brain, brainstem, and cerebellum in children with opsoclonus-myoclonus ataxia syndrome.

Application of neuroimaging in diagnosis of focal cortical dysplasia: A survey of computational techniques

Focal Cortical Dysplasia (FCD) is a neurodevelopmental disorder characterized by abnormal neuronal migration, differentiation, and maturation, resulting in a range of clinical symptoms including drug-resistant epilepsy. Accurate and timely diagnosis of FCD is essential for effective treatment and management of patients with this condition. In recent years, there have been significant advances in neuroimaging and machine learning techniques, enabling automated detection of FCD lesions with increasing accuracy and efficiency.In this survey article, we provide an overview of the current state-of-the-art in computational techniques for the automated detection of FCD lesions using neuroimaging. We first introduce some common imaging findings that radiologists look for in FCD lesions, then review the automatic detection techniques studied in the last decade. These techniques have shown promising results in detecting and localizing FCD lesions, improving diagnostic accuracy and reducing misdiagnoses, diagnosis time, and cost of care.We suggest that the use of these types of quantitative image analysis tools and algorithms can play a key role in improving the overall management and outcome of patients with FCD. However, further studies are needed to validate and optimize the performance of these techniques in clinical practice. In summary, the combination of advanced neuroimaging and computational techniques has the potential to revolutionize the diagnosis and treatment of FCD, leading to better outcomes for patients with this condition.

Depletion of SMN protein in mesenchymal progenitors impairs the development of bone and neuromuscular junction in spinal muscular atrophy.

Spinal muscular atrophy (SMA) is a neuromuscular disorder characterized by the deficiency of the survival motor neuron (SMN) protein, which leads to motor neuron dysfunction and muscle atrophy. In addition to the requirement for SMN in motor neurons, recent studies suggest that SMN deficiency in peripheral tissues plays a key role in the pathogenesis of SMA. Using limb mesenchymal progenitor cell (MPC)-specific SMN-depleted mouse models, we reveal that SMN reduction in limb MPCs causes defects in the development of bone and neuromuscular junction (NMJ). Specifically, these mice exhibited impaired growth plate homeostasis and reduced insulin-like growth factor (IGF) signaling from chondrocytes, rather than from the liver. Furthermore, the reduction of SMN in fibro-adipogenic progenitors (FAPs) resulted in abnormal NMJ maturation, altered release of neurotransmitters, and NMJ morphological defects. Transplantation of healthy FAPs rescued the morphological deterioration. Our findings highlight the significance of mesenchymal SMN in neuromusculoskeletal pathogenesis of SMA and provide insights into potential therapeutic strategies targeting mesenchymal cells for the treatment of SMA.

Depletion of SMN protein in mesenchymal progenitors impairs the development of bone and neuromuscular junction in spinal muscular atrophy

Spinal muscular atrophy (SMA) is a neuromuscular disorder characterized by the deficiency of the survival motor neuron (SMN) protein, which leads to motor neuron dysfunction and muscle atrophy. In addition to the requirement for SMN in motor neurons, recent studies suggest that SMN deficiency in peripheral tissues plays a key role in the pathogenesis of SMA. Using limb mesenchymal progenitor cell (MPC)-specific SMN-depleted mouse models, we reveal that SMN reduction in limb MPCs causes defects in the development of bone and neuromuscular junction (NMJ). Specifically, these mice exhibited impaired growth plate homeostasis and reduced insulin-like growth factor (IGF) signaling from chondrocytes, rather than from the liver. Furthermore, the reduction of SMN in fibro-adipogenic progenitors (FAPs) resulted in abnormal NMJ maturation, altered release of neurotransmitters, and NMJ morphological defects. Transplantation of healthy FAPs rescued the morphological deterioration. Our findings highlight the significance of mesenchymal SMN in neuromusculoskeletal pathogenesis of SMA and provide insights into potential therapeutic strategies targeting mesenchymal cells for the treatment of SMA.

Development of trofinetide for the treatment of Rett syndrome: from bench to bedside.

Rett syndrome (RTT) is rare neurodevelopmental disorder caused by mutations in the MECP2 gene that encodes methyl-CpG-binding protein 2 (MeCP2), a DNA-binding protein with roles in epigenetic regulation of gene expression. Functional loss of MeCP2 results in abnormal neuronal maturation and plasticity, characterized by loss of verbal communication and loss of fine and gross motor function, among others. Trofinetide, a synthetic analog of glycine-proline-glutamate, was approved by the US Food and Drug Administration for the treatment of RTT in adult and pediatric patients aged 2years and older. Here, we present the development of trofinetide from bench research to clinical studies and emphasize how the collaboration between academia, the pharmaceutical industry, and patient advocacy led to the recent approval. The bench-to-bedside development of trofinetide underscores the value of collaboration between these groups in the development and approval of treatments for rare diseases.

Assessing cortical features in early stage ASD children.

Autism Spectrum Disorder (ASD) is defined as a neurodevelopmental disorder largely investigated in the neurologic field. Recently, neuroimaging studies have been conducted in order to investigate cerebral morphologic alterations in ASD patients, demonstrating an atypical brain development before the clinical manifestations of the disorder. Cortical Thickness (CT) and Local Gyrification Index (LGI) distribution for ASD children were investigated in this study, with the aim to evaluate possible relationship between brain measures and individual characteristics (i.e., IQ and verbal ability). 3D T1-w sequences from 129 ASD and 58 age-matched Healthy Controls (HC) were acquired and processed in order to assess CT and LGI for each subject. Intergroup differences between ASD and HC were investigated, including analyses of 2 ASD subgroups, split according to patient verbal ability and IQ. When compared to HC, ASD showed increased CT and LGI within several brain areas, both as an overall group and as verbal ability an IQ subgroups. Moreover, when comparing language characteristics of the ASD subjects, those patients with verbal ability exhibit significant CT and LGI increase was found within the occipital lobe of right hemisphere. No significant results occurred when comparing ASD patients according to their IQ value. These results support the hypothesis of abnormal brain maturation in ASD since early childhood with differences among clinical subgroups suggesting different anatomical substrates underlying an aberrant connectivity.

Cfp1 Controls Cardiomyocyte Maturation by Modifying Histone H3K4me3 of Structural, Metabolic, and Contractile Related Genes.

Cardiomyocyte maturation is the final stage of heart development, and abnormal cardiomyocyte maturation will lead to serious heart diseases. CXXC zinc finger protein 1 (Cfp1), a key epigenetic factor in multi-lineage cell development, remains underexplored in its influence on cardiomyocyte maturation. This study investigates the role and mechanisms of Cfp1 in this context. Cardiomyocyte-specific Cfp1 knockout (Cfp1-cKO) mice died within 4 weeks of birth. Cardiomyocytes derived from Cfp1-cKO mice showed an inhibited maturation phenotype, characterized by structural, metabolic, contractile, and cell cycle abnormalities. In contrast, cardiomyocyte-specific Cfp1 transgenic (Cfp1-TG) mice and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) overexpressing Cfp1 displayed a more mature phenotype. Mechanistically, deficiency of Cfp1 led to a reduction in trimethylation on lysine 4 of histone H3 (H3K4me3) modification, accompanied by the formation of ectopic H3K4me3. Furthermore, Cfp1 deletion decreased the level of H3K4me3 modification in adult genes and increased the level of H3K4me3 modification in fetal genes. Collectively, Cfp1 modulates the expression of genes crucial to cardiomyocyte maturation by regulating histone H3K4me3 modification, thereby intricately influencing the maturation process. This study implicates Cfp1 as an important molecule regulating cardiomyocyte maturation, with its dysfunction strongly linked to cardiac disease.

Update on Adrenarche-Still a Mystery.

Adrenarche marks the timepoint of human adrenal development when the cortex starts secreting androgens in increasing amounts, in healthy children at age 8-9 years, with premature adrenarche (PA) earlier. Because the molecular regulation and significance of adrenarche are unknown, this prepubertal event is characterized descriptively, and PA is a diagnosis by exclusion with unclear long-term consequences. We searched the literature of the past 5 years, including original articles, reviews, and meta-analyses from PubMed, ScienceDirect, Web of Science, Embase, and Scopus, using search terms adrenarche, pubarche, DHEAS, steroidogenesis, adrenal, and zona reticularis. Numerous studies addressed different topics of adrenarche and PA. Although basic studies on human adrenal development, zonation, and zona reticularis function enhanced our knowledge, the exact mechanism leading to adrenarche remains unsolved. Many regulators seem involved. A promising marker of adrenarche (11-ketotestosterone) was found in the 11-oxy androgen pathway. By current definition, the prevalence of PA can be as high as 9% to 23% in girls and 2% to 10% in boys, but only a subset of these children might face related adverse health outcomes. New criteria for defining adrenarche and PA are needed to identify children at risk for later disease and to spare children with a normal variation. Further research is therefore required to understand adrenarche. Prospective, long-term studies should characterize prenatal or early postnatal developmental pathways that modulate trajectories of birth size, early postnatal growth, childhood overweight/obesity, adrenarche and puberty onset, and lead to abnormal sexual maturation, fertility, and other adverse outcomes.

Cypher/ZASP drives cardiomyocyte maturation via actin-mediated MRTFA-SRF signalling.

Rationale: Cardiomyocytes (CMs) undergo dramatic structural and functional changes in postnatal maturation; however, the regulatory mechanisms remain greatly unclear. Cypher/Z-band alternatively spliced PDZ-motif protein (ZASP) is an essential sarcomere component maintaining Z-disc stability. Deletion of mouse Cypher and mutation in human ZASP result in dilated cardiomyopathy (DCM). Whether Cypher/ZASP participates in CM maturation and thereby affects cardiac function has not been answered. Methods: Immunofluorescence, transmission electron microscopy, real-time quantitative PCR, and Western blot were utilized to identify the role of Cypher in CM maturation. Subsequently, RNA sequencing and bioinformatics analysis predicted serum response factor (SRF) as the key regulator. Rescue experiments were conducted using adenovirus or adeno-associated viruses encoding SRF, both in vitro and in vivo. The molecular mechanisms were elucidated through G-actin/F-actin fractionation, nuclear-cytoplasmic extraction, actin disassembly assays, and co-sedimentation assays. Results: Cypher deletion led to impaired sarcomere isoform switch and morphological abnormalities in mitochondria, transverse-tubules, and intercalated discs. RNA-sequencing analysis revealed significant dysregulation of crucial genes related to sarcomere assembly, mitochondrial metabolism, and electrophysiology in the absence of Cypher. Furthermore, SRF was predicted as key transcription factor mediating the transcriptional differences. Subsequent rescue experiments showed that SRF re-expression during the critical postnatal period effectively rectified CM maturation defects and notably improved cardiac function in Cypher-depleted mice. Mechanistically, Cypher deficiency resulted in the destabilization of F-actin and a notable increase in G-actin levels, thereby impeding the nuclear localisation of myocardin-related transcription factor A (MRTFA) and subsequently initiating SRF transcription. Conclusion: Cypher/ZASP plays a crucial role in CM maturation through actin-mediated MRTFA-SRF signalling. The linkage between CM maturation abnormalities and the late-onset of DCM is suggested, providing further insights into the pathogenesis of DCM and potential treatment strategies.

ZP1-Y262C mutation causes abnormal zona pellucida formation and female infertility in humans.

Defective oocyte maturation is a common cause of female infertility. The loss of the zona pellucida (ZP) represents a specific condition of impaired oocyte maturation. The extracellular matrix known as the ZP envelops mammalian oocytes and preimplantation embryos, exerting significant influence on oogenesis, fertilization, and embryo implantation. However, the genetic factors leading to the loss of the ZP in oocytes are not well understood. This study focused on patients who underwent oocyte retrieval surgery after ovarian stimulation and were found to have abnormal oocyte maturation without the presence of the ZP. Ultrasonography was performed during the surgical procedure to evaluate follicle development. Peripheral blood samples from the patient were subjected to exome sequencing. Here, a novel, previously unreported heterozygous mutation in the ZP1 gene was identified. Within the ZP1 gene, we discovered a novel heterozygous mutation (ZP1 NM_207341.4:c.785A>G (p.Y262C)), specifically located in the trefoil domain. Bioinformatics comparisons further revealed conservation of the ZP1-Y262C mutation across different species. Model predictions of amino acid mutations on protein structure and cell immunofluorescence/western blot experiments collectively confirmed the detrimental effects of the ZP1-Y262C mutation on the function and expression of the ZP1 protein. The ZP1-Y262C mutation represents the novel mutation in the trefoil domain of the ZP1 protein, which is associated with defective oocyte maturation in humans. Our report enhances comprehension regarding the involvement of ZP-associated genes in female infertility and offers enriched understanding for the genetic diagnosis of this condition.

Spinal epidural abscess mimicking chloroma

Background: Spinal epidural abscess (SEA) is a rare and generally challenging condition. It represents a challenge in terms of diagnosis and treatment due to the low incidence, vague presentation, and rapid neurologic deterioration. Case Presentation: A 40-year-old gentleman who was previously healthy was presented with a 2-week history of decreased vision associated with fever and cough one week before the eye symptoms. The patient was admitted as a case of acute promyelocytic leukemia. And was started on chemotherapy, which was complicated by septicemia and was shifted to the intensive care unit. Three weeks after admission he started to have severe non-radicular neck pain. He also had bilateral lower limb pain spasm-like in nature. Patient denies any history of trauma, weakness, numbness, saddle anesthesia or loss of sphincter control. Labs showed leukopenia, anemia and thrombocytopenia. Bone marrow result showed hypercellularity with active trilineage hematopoiesis and no immunomorphological evidence of abnormal promyelocytes or maturation arrest. He underwent C3-C4 laminectomy and evacuation of epidural pus collection. Intra-operative course was free of complication. Upon post-operative assessment he was hemodynamically stable, his neck pain was better with morphine doses and he was able to mobilize it. Conclusion: Spinal epidural abscess can mimic chloroma clinically and radiologically, however, diffusion with the images and enhancement is essential for proper assessment since a proper medical and surgical intervention is needed.

Cadmium-induced Carcinogenesis in Respiratory Organs and the Prostate: Insights from Three Perspectives on Toxicogenomic Approach.

Cadmium (Cd) exposure primarily occurs through inhalation, either by smoking or occupational exposure to contaminated air. Upon inhalation, Cd ultimately reaches the prostate through the bloodstream. In this review, we investigate the carcinogenic potential of Cd in both respiratory organs and the prostate. Specifically, this review examines cellular metabolism, comprehensive toxicity, and carcinogenic mechanisms by exploring gene ontology, biological networks, and adverse outcome pathways. In the respiratory organs, Cd induces lung cancer by altering the expression of IL1B and FGF2, causing DNA damage, reducing cell junction integrity, and promoting apoptosis. In the prostate, Cd induces prostate cancer by modifying the expression of EDN1 and HMOX1, leading to abnormal protein activities and maturation, suppressing tumor suppressors, and inducing apoptosis. Collectively, this review provides a comprehensive understanding of the carcinogenic mechanisms of Cd in two different organs by adopting toxicogenomic approaches. These insights can serve as a foundation for further research on cadmium-induced cancer, contributing to the establishment of future cancer prevention strategies.

Abnormal tooth maturation associated with port wine stains.

This study aims to evaluate the potential impact of port wine stains on the development of permanent teeth in mixed dentition and provide insights for managing tooth abnormalities in patients with port wine stains. A retrospective analysis was conducted on 21 patients with mixed dentition and unilateral maxillary port wine stains. Two researchers concurrently utilized Nolla Analysis to assess the developmental stage of bilateral maxillary and permanent mandibular teeth based on panoramic radiographs. The cumulative developmental values of upper and lower permanent teeth on both sides were calculated, and a comparison was made between the developmental stages of the upper and lower jaws. Port wine stains can influence the maturity of permanent upper teeth, within the unilateral maxillary port wine stains range, with an early developmental completion observed on the affected side compared to the unaffected side. While the developmental stages of the lower teeth on both sides showed similarities. Port wine stains can accelerate the maturity of teeth on the affected side, leading to alterations in the order of tooth eruption and subsequent abnormal occlusion in children with mixed dentition. These findings provide a basis for developing appropriate management strategies for addressing tooth abnormalities in patients with port wine stains.

HDAC1-Mediated lncRNA Stimulatory Factor of Follicular Development to Inhibit the Apoptosis of Granulosa Cells and Regulate Sexual Maturity through miR-202-3p-COX1 Axis.

Abnormal sexual maturity exhibits significant detrimental effects on adult health outcomes, and previous studies have indicated that targeting histone acetylation might serve as a potential therapeutic approach to regulate sexual maturity. However, the mechanisms that account for it remain to be further elucidated. Using the mouse model, we showed that Trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, downregulated the protein level of Hdac1 in ovaries to promote the apoptosis of granulosa cells (GCs), and thus arrested follicular development and delayed sexual maturity. Using porcine GCs as a cell model, a novel sexual maturity-associated lncRNA, which was named as the stimulatory factor of follicular development (SFFD), transcribed from mitochondrion and mediated by HDAC1, was identified using RNA sequencing. Mechanistically, HDAC1 knockdown significantly reduced the H3K27ac level at the -953/-661 region of SFFD to epigenetically inhibit its transcription. SFFD knockdown released miR-202-3p to reduce the expression of cyclooxygenase 1 (COX1), an essential rate-limited enzyme involved in prostaglandin synthesis. This reduction inhibited the proliferation and secretion of 17β-estradiol (E2) while promoting the apoptosis of GCs. Consequently, follicular development was arrested and sexual maturity was delayed. Taken together, HDAC1 knockdown-mediated SFFD downregulation promoted the apoptosis of GCs through the miR-202-3p-COX1 axis and lead to delayed sexual maturity. Our findings reveal a novel regulatory network modulated by HDAC1, and HDAC1-mediated SFFD may be a promising new therapeutic target to treat delayed sexual maturity.

Modeling TCIRG1 Neutropenia By Utilizing Patient Derived Induced Pluripotent Stem Cells

Background: Severe congenital neutropenia (SCN) is a hereditary hematopoietic disorder characterized by recurrent infections and leukemic transformation. We previously identified a novel heterozygous missense mutation (R736S) in TCIRG1 causing SCN in a large multigenerational family (PMID: 24753205). We also identified a statistically significant correlation in individuals with lower ANC with rare missense variants in TCIRG1 in a large cohort of population-based genome screening (PMID: 27229898). Recently, we have discovered another TCIRG1 heterozygousvariant: R736C in two unrelated families with congenital neutropenia. Independently, Shinwari et al identified a novel TCIRG1 mutation, V52L, which was correlated with the neutropenic phenotype (PMID: 35573728). Homozygous mutations of TCIRG1 are causing autosomal recessive osteopetrosis (ARO), which is characterized by osteopetrosis and secondary hematological and neurological abnormalities. The Genome Aggregation Database (gnomAD) has no data about TCIRG1 R736S and R736C variants accruing in the healthy population. This is in contrast of numerous healthy individuals with a various heterozygous variants throughout the TCIRG1 gene. Aims: We aim to generate in-vitro model of TCIRG1 neutropenia by utilizing patient derived induced pluripotent stem cells (iPSCs) and characterize the hematological abnormalities caused by mutant TCIRG1 expression. We also aim to characterize the incidence of TCIRG1-associated neutropenia and the prevalence of TCIRG1 variants by sharing this new information within the scientific community through identifying more cases/families with this disease. Methods: Medical history, differential WBC counts and whole exome sequencing (WES) was used to identify the genetic basis of neutropenia in the family with R736S mutation. The R736C mutation in the new families also were identified by WES of the index cases. Medical history, blood counts and Sanger sequencing identified their affected relatives. iPSC lines of 4 affected individuals from 2 families harboring TCIRG1 R736S and R736C mutations and healthy volunteer were generated using standard methodology. S732F benign heterozygous TCIRG1 variant was generated by utilizing CRISPR/Cas9 knock-In gene editing using a healthy volunteer iPSC line. iPSCs hematopoietic differentiation was achieved by using STEMdiff Hematopoietic Kit. Resultant CD34+ cells were differentiated towards neutrophils using a published protocol (PMID: 35795780). Cell proliferation, survival and differentiation characteristics were measured by automated cell counter and flow cytometry, cell viability staining and surface markers analysis for granulocytic differentiation. Results: The discovery of these families came through the reputation and work of the Severe Congenital Neutropenia International Registry (SCNIR) and the expanding use of WES to identify the causes of hereditary disorders. With the identification of the index cases, we have identified several affected family members in each family. In all three families the severity of neutropenia is variable and correlates with the incidence of recurrent bacterial infections. Myeloid differentiation of CD34+ cells derived from iPSCs of 4 patients from two families with R736S and R736C mutations revealed maturation, survival and proliferation abnormalities compared to volunteer cells and cell line harboring benign S732F heterozygous TCIRG1 mutation. All patient cells had from 2 to 3-fold drop in cell proliferation and cell viability drop by over 70% for the patients with R736S and up to 90% with R736C mutation compared to the controls. The myeloid differentiation in the patients' cells, measured by flow cytometry and assessed by CD66b+/CD14+ and CD11b+/CD15+ markers was impaired by 38% and 23% respectively for the patients with R736S and 43% and 24% for the patients harboring R736C mutation. Conclusions: The pathogenesis of TCIRG1-associated neutropenia is currently unknown. Carriers of pathogenic variants from these families appear to have neutropenia with variable severity. Our preliminary studies show that patient iPSC derived hematopoietic cells closely recapitulate the neutropenic phenotype. Population based genomic studies; careful family histories and WES will identify new patients with this disorder.