Whole paternal uniparental disomy of chromosome 4 with a novel homozygous IDUA splicing variant, c.159-9T>A, in a Chinese patient with mucopolysaccharidosis type I.

BACKGROUND: Mucopolysaccharidosis type I (MPS-I) is a rare autosomal recessive genetic lysosomal storage disorder that is caused by pathogenic variants of the α-L-iduronidase (IDUA) gene. This study aimed to identify the genetic causes of MPS-I in a Chinese patient and construct a minigene of IDUA to analyze its variants upon splicing. METHODS: Whole-exome sequencing (WES) and Sanger sequencing were used to confirm the potential causative variants. Single-nucleotide polymorphism (SNP) array was subsequently performed to confirm uniparental disomy (UPD). Minigene assay was performed to analyze the effect on splicing of mRNA. We meanwhile explored the conservative analysis and protein homology simulation. RESULTS: A novel homozygous splicing mutation of IDUA, c.159-9T>A, was identified in an individual presenting with overlapping features of MPS-I. Interestingly, only the father and sisters, but not the mother, carried the variant in a heterozygous state. WES and SNP array analyses validated paternal UPD on chromosome 4. Minigene splicing revealed two aberrant splicing events: exon 2 skipping and intron 1 retention. Moreover, the specific structure of the mutant protein obviously changed according to the results of the homologous model. CONCLUSIONS: This study describes a rare autosomal recessive disorder with paternal UPD of chromosome 4 leading to the homozygosity of the IDUA splicing variant in patients with MPS-I for the first time. This study expands the variant spectrum of IDUA and provides insights into the splicing system, facilitating its enhanced diagnosis and treatment.

[Clinical characteristics and genetic analysis of children and adolescents with monogenic diabetes].

OBJECTIVE: To explore the clinical characteristics and molecular basis for children and adolescents with monogenic diabetes. METHODS: A retrospective analysis was carried out for the clinical manifestations and laboratory data of 116 children and adolescents diagnosed with diabetes at Ningbo Women and Children's Hospital from January 2020 to March 2023. Whole exome sequencing and mitochondrial gene sequencing were carried out on 21 children with suspected monogenic diabetes. RESULTS: A total of 10 cases of monogenic diabetes were diagnosed, all of which were Maturity-onset Diabetes Of the Young (MODY). Six cases of MODY2 were due to GCK gene mutations, 1 case of MODY3 was due to HNF1A gene mutation, 2 cases of MODY12 were due to ABCC8 gene mutations, and 1 case of MODY13 was due to KCNJ11 gene mutation. Nine of the 10 patients with MODY had no typical symptoms of diabetes. A family history of diabetes was significantly more common in the MODY group compared with the T1DM and T2DM groups (P < 0.05). The BMI of the MODY group was higher than that of the T1DM group (P < 0.05). The initial blood glucose level was lower than that of the T1DM group (P < 0.05), and there was no significant difference compared with the T2DM group. The fasting C-peptide level of the MODY group was higher than that of the T1DM group (P < 0.05), and there was no significant difference compared with the T2DM group. Glycosylated hemoglobin of the MODY group was lower than both the T1DM and T2DM groups (P < 0.05). CONCLUSION: In this study, MODY has accounted for the majority of monogenic diabetes among children and adolescents, and the common mutations were those of the GCK gene in association with MODY2. Blood glucose and glycosylated hemoglobin of children with MODY were slightly increased, whilst the islet cell function had remained, and the clinical manifestations and laboratory tests had overlapped with those of type 2 diabetes. WES and mitochondrial gene sequencing can clarify the etiology of monogenic diabetes and facilitate precise treatment.

Predictive Care Improves Recovery Status and Complications after Ureteral Calculi Surgery: A Retrospective Study.

OBJECTIVE: Predictive care in patients undergoing ureteroscopic stone surgery has emerged as a promising approach. Thus, this study aims to enhance personalised nursing plans and reduce the risk of complications by conducting predictive analysis of possible risks early in the treatment and nursing process. METHODS: Clinical data were collected from 108 patients who underwent ureteroscopic stone surgery and were admitted to our hospital between January 2020 and January 2023. Patients were divided into a control group (conventional nursing, n = 53) and an observation group (predictive care, n = 55) based on the nursing method, and various clinical indicators were compared between the two groups of surgical patients. RESULTS: No significant difference in general data was found between the two groups (p > 0.05). Compared with the control group, the first time to exhaust gas (p < 0.05), the first time to get out of bed (p < 0.05), the time to exhaust stone (p < 0.05), the first time to defecate (p < 0.05) and the length of hospital stay (p < 0.05) in the observation group were shorter; 1 day after surgery, no significant differences in all dimensions of the general comfort questionnaire (GCQ) score were found; 2 days after surgery, the GCQ score in all dimensions of the observation group was significantly higher than that of the control group (p < 0.05). The incidence of postoperative complications in the observation group was significantly lower than that in the control group (p < 0.05). CONCLUSIONS: Predictive nursing can effectively improve the patients with ureteral calculi, accelerate the process of postoperative recovery and reduce the occurrence of complications; Thus, this process is worthy of widespread clinical promotion.

Case Report: Identification of a novel PRR12 variant in a Chinese boy with developmental delay and short stature.

Proline Rich 12 (PRR12) protein is primarily expressed in the brain and localized in the nucleus. The variants in the PRR12 gene were reported to be related to neuroocular syndrome. Patients with PRR12 gene presented with intellectual disability (ID), neuropsychiatric disorders, some congenital anomalies, and with or without eye abnormalities. Here, we report an 11-year-old boy with a novel PRR12 variant c.1549_1568del, p.(Pro517Alafs*35). He was the first PRR12 deficiency patient in China and presented with ID, short stature, and mild scoliosis. He could not concentrate on his studies and was diagnosed with attention deficit hyperactivity disorder (ADHD). The insulin-like growth factor 1 (IGH-1) was low in our patient, which may be the cause of his short stature. Patients with neuroocular syndrome are rare, and further exploration is needed to understand the reason for neurodevelopmental abnormalities caused by PRR12 variants. Our study further expands on the PRR12 variants and presents a new case involving PPR12 variants.

The significance of machine learning in neonatal screening for inherited metabolic diseases.

Background: Neonatal screening for inherited metabolic diseases (IMDs) has been revolutionized by tandem mass spectrometry (MS/MS). This study aimed to enhance neonatal screening for IMDs using machine learning (ML) techniques. Methods: The study involved the analysis of a comprehensive dataset comprising 309,102 neonatal screening records collected in the Ningbo region, China. An advanced ML system model, encompassing nine distinct algorithms, was employed for the purpose of predicting the presence of 31 different IMDs. The model was compared with traditional cutoff schemes to assess its diagnostic efficacy. Additionally, 180 suspected positive cases underwent further evaluation. Results: The ML system exhibited a significantly reduced positive rate, from 1.17% to 0.33%, compared to cutoff schemes in the initial screening, minimizing unnecessary recalls and associated stress. In suspected positive cases, the ML system identified 142 true positives with high sensitivity (93.42%) and improved specificity (78.57%) compared to the cutoff scheme. While false negatives emerged, particularly in heterozygous carriers, our study revealed the potential of the ML system to detect asymptomatic cases. Conclusion: This research provides valuable insights into the potential of ML in pediatric medicine for IMD diagnosis through neonatal screening, emphasizing the need for accurate carrier detection and further research in this domain.

Metformin plus L-carnitine enhances brown/beige adipose tissue activity via Nrf2/HO-1 signaling to reduce lipid accumulation and inflammation in murine obesity.

This study investigated how Metformin (Met) combined with L-carnitine (L-car) modulates brown adipose tissue (BAT) to affect obesity. High-fat-induced obese rats received daily oral gavage with Met and/or L-car, followed by serum biochemical analysis, histopathological observation on adipose tissues, and immunochemistry test for the abdominal expression of BAT-specific uncoupling protein 1 (UCP1). Mouse-embryonic-fibroblast cells were induced into adipocytes, during which Met plus L-car was added with/without saturated fatty acid (SFA). The role of nuclear factor erythroid 2-related factor 2 (Nrf2) in adipocyte browning was investigated by gene silencing. Mitochondria biogenesis in adipocytes was inspected by Mitotracker staining. Nrf2/heme oxygenase-1 (HO-1)/BAT-related genes/proinflammatory marker expressions in adipose tissues and/or adipocytes were analyzed by Western blot, qRT-PCR, and/or immunofluorescence test. Met or L-car improved metabolic disorders, reduced adipocyte vacuolization and swelling, upregulated levels of BAT-related genes including UCP1 and downregulated proinflammatory marker expressions, and activated the Nrf2/HO-1 pathway in adipose tissues of obese rats. Met and L-car functioned more strongly than alone. In adipocytes, Met plus L-car upregulated BAT-related gene levels and protected against SFA-caused inflammation promotion and mitochondria degeneration, which yet was attenuated by Nrf2 silencing. Met plus L-car enhances BAT activity and white adipose tissue browning via the Nrf2/HO-1 pathway to reduce lipid accumulation and inflammation in obese rats.

Ovotesticular Disorder of Sex Development Presenting as a Scrotal Emergency.

Ovotesticular (OT) disorder of sex development (DSD) is a rare condition that affects the development of reproductive organs and manifests in a wide range of phenotypic presentations. The clinical diagnosis of this condition is challenging because of its atypical nature, and the variability of presentation in 46,XX OT-DSD cases makes it a complex issue in medical practice. We report a case of a 13-year-old boy who presented with left scrotal pain. Further exploration revealed a tunica rupture without testicular torsion of the left testis, whereas the histopathological analysis of a nodule excised from the right testis indicated the presence of ovotestis tissues. A second nonemergent surgery preserved the testicular tissues as the ovarian tissue in both gonads was excised. After 22 months of follow-up, the patient's testes produced normal testosterone levels sustained over time without any exogenous supplementation. This case reveals that, in male children who present with an acute scrotal disease as adolescents, the gonads should be retained until the etiology is confirmed, and the possibility of OT-DSD should be considered.

[A prospective study of genetic screening of 2 060 neonates by high-throughput sequencing].

OBJECTIVE: To assess the value of genetic screening by high-throughput sequencing (HTS) for the early diagnosis of neonatal diseases. METHODS: A total of 2 060 neonates born at Ningbo Women and Children's Hospital from March to September 2021 were selected as the study subjects. All neonates had undergone conventional tandem mass spectrometry metabolite analysis and fluorescent immunoassay analysis. HTS was carried out to detect the definite pathogenic variant sites with high-frequency of 135 disease-related genes. Candidate variants were verified by Sanger sequencing or multiplex ligation-dependent probe amplification (MLPA). RESULTS: Among the 2 060 newborns, 31 were diagnosed with genetic diseases, 557 were found to be carriers, and 1 472 were negative. Among the 31 neonates, 5 had G6PD, 19 had hereditary non-syndromic deafness due to variants of GJB2, GJB3 and MT-RNR1 genes, 2 had PAH gene variants, 1 had GAA gene variants, 1 had SMN1 gene variants, 2 had MTTL1 gene variants, and 1 had GH1 gene variants. Clinically, 1 child had Spinal muscular atrophy (SMA), 1 had Glycogen storage disease II, 2 had congenital deafness, and 5 had G6PD deficiency. One mother was diagnosed with SMA. No patient was detected by conventional tandem mass spectrometry. Conventional fluorescence immunoassay had revealed 5 cases of G6PD deficiency (all positive by genetic screening) and 2 cases of hypothyroidism (identified as carriers). The most common variants identified in this region have involved DUOX2 (3.93%), ATP7B (2.48%), SLC26A4 (2.38%), GJB2 (2.33%), PAH (2.09%) and SLC22A5 genes (2.09%). CONCLUSION: Neonatal genetic screening has a wide range of detection and high detection rate, which can significantly improve the efficacy of newborn screening when combined with conventional screening and facilitate secondary prevention for the affected children, diagnosis of family members and genetic counseling for the carriers.

[Analysis of a Chinese pedigree affected with familial short stature due to 15q25.3q26.1 deletion involving the ACAN gene].

OBJECTIVE: To analyze the genetic etiology of a Chinese pedigree affected with short stature. METHODS: A child with familial short stature (FSS) who had presented at the Ningbo Women and Children's Hospital in July 2020 and his parents and paternal and maternal grandparents were selected as the study subject. Clinical data of the pedigree was collected, and the proband was subjected to routine growth and development assessment. Peripheral blood samples were collected. The proband was subjected to whole exome sequencing (WES), and the proband, his parents and grandparents were subjected to chromosomal microarray analysis (CMA). RESULTS: The height of the proband and his father was 87.7cm (-3 s) and 152 cm (-3.39 s) respectively. Both of them were found to harbor a 15q25.3-q26.1 microdeletion, which has encompassed the whole of the ACAN gene which is closely associated with short stature. The CMA results of his mother and grandparents were all negative, and above deletion has not been included in population database and related literature, and was rated as pathogenic based on the guidelines from the American College of Medical Genetics and Genomics (ACMG). After 14 months of rhGH treatment, the height of the proband has increased to 98.5 cm (-2.07 s). CONCLUSION: The 15q25.3-q26.1 microdeletion probably underlay the FSS, in this pedigree. Short-term rhGH treatment can effectively improve the height of the affected individuals.

[Clinical and genetic analysis of a child with Culler-Jones syndrome due to variant of GLI2 gene].

OBJECTIVE: To explore the genetic basis for a child featuring short stature and postaxial polydactyly. METHODS: A child who presented at Ningbo Women & Children's Hospital in May 2021 due to the"discovery of growth retardation for more than two years" was selected as the subject. Peripheral blood samples of the child and his parents were collected for the extraction of genomic DNA. Whole exome sequencing was carried out for the child, and candidate variant was verified by Sanger sequencing of his family members. RESULTS: The child was found to harbor a heterozygous c.3670C>T (p.Q1224) variant of the GLI2 gene, which may lead to premature termination of protein translation. The variant was not detected in either parent. CONCLUSION: The child was diagnosed with Culler-Jones syndrome. The c.3670C>T (p.Q1224*) variant of the GLI2 gene probably underlay the disease in this child.

KISS1/KISS1R mediates Sertoli cell apoptosis via the PI3K/AKT signalling pathway in a high­glucose environment.

In male patients with diabetes, reduced sperm motility and fertility are observed. KiSS­1 metastasis suppressor (KISS1)/KISS1 receptor (KISS1R) serves an important role in regulating adolescent sexual maturity and reproductive system development in mammals; however, the mechanism underlying KISS1/KISS1R in reproductive dysfunction in male patients with diabetes is not completely understood. The aim of the present study was to examine the role of KISS1/KISS1R in Sertoli cells. High glucose (HG)­induced mouse Sertoli cells were used to model diabetes in vitro. KISS1/KISS1R overexpression and knockdown were established in mouse Sertoli cells. Reverse transcription­quantitative PCR and western blotting were performed to measure the expression levels of KISS1/KISS1R and apoptosis­related proteins. Cell viability and apoptosis was assessed by performing Cell Counting Kit­8, TUNEL staining and flow cytometry assays, respectively. Western blotting was performed to assess the expression levels of PI3K/AKT signalling­related proteins. KISS1/KISS1R expression levels were downregulated in HG­induced mouse Sertoli cells compared with control cells. KISS1/KISS1R overexpression significantly suppressed HG­induced apoptosis and decrease of viability in mouse Sertoli cells. Moreover, the western blotting results indicated that KISS1/KISS1R activated PI3K/AKT signalling. Treatment with PI3K/AKT pathway inhibitor significantly reversed KISS1/KISS1R­mediated protective effects. Collectively, the results of the present study suggested that KISS1/KISS1R mediated Sertoli cell apoptosis via the PI3K/AKT signalling pathway under HG conditions, which provided reliable targets for the treatment of reproductive dysfunction in male patients with diabetes.

Identification of Six Novel Variants of ACAD8 in Isobutyryl-CoA Dehydrogenase Deficiency With Increased C4 Carnitine Using Tandem Mass Spectrometry and NGS Sequencing.

Isobutyryl-CoA dehydrogenase deficiency (IBDHD, MIM: #611283) is a rare autosomal recessive hereditary disease, which is caused by genetic mutations of acyl-CoA dehydrogenase (ACAD) 8 and associated with valine catabolism. Here, tandem mass spectrometry (MS/MS) was applied to screen 302,993 neonates for inherited metabolic diseases (IMD) in Ningbo of China from 2017 to 2020. The results suggest that 198 newborns (0.7‰) were initially screened positive for IBDHD with C4-Carnitine, and 27 cases (0.1‰) were re-screened positive. Genetic diagnosis was performed on 21 of the 27 cases. Seven compound heterozygous variations, three biallelic variations, and one heterozygous variation of ACAD8 were found with a pathogenicity rate of 33.3% (7/21). In addition, seven biallelic variations, one heterozygous variation of acyl-CoA dehydrogenase short chain (ACADS), and one biallelic variation of acyl-CoA dehydrogenase short/branched chain (ACADSB) was detected. Further research showed that ACAD8 mutations of 11 IBDHD cases distributed in six different exons with total 14 mutation sites. Five of which were known suspected pathogenic sites (c.286G > A, c.553C > T, c.1000C > T, c.409G > A, c.500del) and six were novel mutation sites: c.911A > T, c.904C > T, c.826G > A, c.995T > C, c.1166G > A, c.1165C > T. This finding enriched the mutation spectrum of ACAD8 in IBDHD.

Curcumin Inhibited Podocyte Cell Apoptosis and Accelerated Cell Autophagy in Diabetic Nephropathy via Regulating Beclin1/UVRAG/Bcl2.

INTRODUCTION: Curcumin has various biological properties including being anti-inflammatory and antidiabetic. Podocyte apoptosis and autophagy dysfunction have been found to be responsible for the development of diabetic nephropathy (DN). Thus, the aim of the study was to investigate the effects of curcumin on the podocyte apoptosis and autophagy in DN and clarify its potential mechanisms. METHODS: The mice with DN induced by injection of streptozotocin were treated with curcumin by gavage at a dose of 200 mg/kg/day for 8 weeks. The serum lipid levels were detected by total cholesterol (TC) and triglyceride (TG) kits at different time points. Renal damage was assessed by detecting urine albumin, serum creatinine (Scr), HE staining and PAS staining. The renal impairment was detected by immunohistochemical staining and TUNEL staining. Western blot assay tested the expression of autophagy-related and apoptotic-related proteins in vivo and vitro. The viabilities and apoptosis of MPC5 cells exposed to high glucose (HG) or curcumin were respectively detected by CCK-8 assay and flow cytometry. RESULTS: The results showed that curcumin significantly decreased the progress of DN possibly via increasing autophagy and inhibiting apoptosis of renal cell in DN mice. Besides, podocyte marker proteins (podocalyxin and nephrin) were markedly increased in DN mice by curcumin treatment. The autophagy-related proteins LC3, p62, Beclin1, UVRAG and ATG5 were significantly affected in DN mice by curcumin, along with reducing expression of pro-apoptotic protein Bax and caspase-3 and increasing anti-apoptotic protein Bcl-2. In vitro, curcumin increased the viabilities and inhibited apoptosis of MPC5 cells exposed to high glucose (HG). In addition, the podocyte autophagy was enhanced partly via regulating beclin1/UVRAG. DISCUSSION: Together, the results showed that curcumin inhibited podocyte apoptosis and accelerated cell autophagy via regulating Beclin1/UVRAG/Bcl2. Thus, the study showed that curcumin exerted significantly protective effects in DN.