Application of tandem mass spectrometry in the screening and diagnosis of mucopolysaccharidoses.

Mucopolysaccharidoses (MPSs) are caused by a deficiency in the enzymes needed to degrade glycosaminoglycans (GAGs) in the lysosome. The storage of GAGs leads to the involvement of several systems and even to the death of the patient. In recent years, an increasing number of therapies have increased the treatment options available to patients. Early treatment is beneficial in improving the prognosis, but children with MPSs are often delayed in their diagnosis. Therefore, there is an urgent need to develop a method for early screening and diagnosis of the disease. Tandem mass spectrometry (MS/MS) is an analytical method that can detect multiple substrates or enzymes simultaneously. GAGs are reliable markers of MPSs. MS/MS can be used to screen children at an early stage of the disease, to improve prognosis by treating them before symptoms appear, to evaluate the effectiveness of treatment, and for metabolomic analysis or to find suitable biomarkers. In the future, MS/MS could be used to further identify suitable biomarkers for MPSs for early diagnosis and to detect efficacy.

Gene variants and clinical characteristics of children with sitosterolemia.

OBJECTIVE: To enhance the detection, management and monitoring of Chinese children afflicted with sitosterolemia by examining the physical characteristics and genetic makeup of pediatric patients. METHODS: In this group, 26 children were diagnosed with sitosterolemia, 24 of whom underwent genetic analysis. Patient family medical history, physical symptoms, tests for liver function, lipid levels, standard blood tests, phytosterol levels, cardiac/carotid artery ultrasounds, fundus examinations, and treatment were collected. RESULTS: The majority (19, 73.1%) of the 26 patients exhibited xanthomas as the most prevalent manifestation. The second most common symptoms were joint pain (7, 26.9%) and stunted growth (4, 15.4%). Among the 24 (92.3%) patients whose genetics were analyzed, 16 (66.7%) harbored ABCG5 variants (type 2 sitosterolemia), and nearly one-third (8, 33.3%) harbored ABCG8 variants (type 1 sitosterolemia). Additionally, the most common pathogenic ABCG5 variant was c.1166G > A (p.Arg389His), which was found in 10 patients (66.7%). Further analysis did not indicate any significant differences in pathological traits among those carrying ABCG5 and ABCG8 variations (P > 0.05). Interestingly, there was a greater abundance of nonsense variations in ABCG5 than in ABCG8 (P = 0.09), and a greater frequency of splicing variations in ABCG8 than ABCG5 (P = 0.01). Following a change in diet or a combination of ezetimibe, the levels of cholesterol and low-density lipoprotein were markedly decreased compared to the levels reported before treatment. CONCLUSION: Sitosterolemia should be considered for individuals presenting with xanthomas and increased cholesterol levels. Phytosterol testing and genetic analysis are important for early detection. Managing one's diet and taking ezetimibe can well control blood lipids.

Using sno-lncRNAs as potential markers for Prader-Willi syndrome diagnosis.

The genetic disorder Prader-Willi syndrome (PWS) is mainly caused by the loss of multiple paternally expressed genes in chromosome 15q11-q13 (the PWS region). Early diagnosis of PWS is essential for timely treatment, leading to effectively easing some clinical symptoms. Molecular approaches for PWS diagnosis at the DNA level are available, but the diagnosis of PWS at the RNA level has been limited. Here, we show that a cluster of paternally transcribed snoRNA-ended long noncoding RNAs (sno-lncRNAs, sno-lncRNA1-5) derived from the SNORD116 locus in the PWS region can serve as diagnostic markers. In particular, quantification analysis has revealed that 6,000 copies of sno-lncRNA3 are present in 1 µL whole blood samples from non-PWS individuals. sno-lncRNA3 is absent in all examined whole blood samples of 8 PWS individuals compared to 42 non-PWS individuals and dried blood samples of 35 PWS individuals compared to 24 non-PWS individuals. Further developing a new CRISPR-MhdCas13c system for RNA detection with a sensitivity of 10 molecules per µL has ensured sno-lncRNA3 detection in non-PWS, but not PWS individuals. Together, we suggest that the absence of sno-lncRNA3 represents a potential marker for PWS diagnosis that can be detected by both RT-qPCR and CRISPR-MhdCas13c systems with only microlitre amount of blood samples. Such an RNA-based sensitive and convenient approach may facilitate the early detection of PWS.

Effects of early recombinant human growth hormone treatment in young Chinese children with Prader-Willi syndrome.

BACKGROUND: Prader-Willi syndrome (PWS) is a rare and multisystemic genetic disorder that is characterized by severe hypotonia, hyperphagia, short stature, and global developmental delay. Although early recombinant human growth hormone (rhGH) treatment has been proven to rescue some symptoms and bring additional benefits to PWS patients, studies in patients under 2 years old are scarce. Thus, this study aims to investigate the effectiveness and safety of rhGH treatment for young children. METHODS: A total of 96 genetically confirmed Chinese PWS infants or toddlers (47 males) followed between 2013 and 2022 were retrospectively analyzed. Sixty-five infants (early treatment group) started rhGH treatment during their first year, and 31 toddlers (later treatment group) started at the age of 1-2 years. Auxological parameters, carbohydrate metabolism parameters, thyroid function, liver function, insulin-like growth factor-1 (IGF-1), and radiographs were acquired before the initiation of the treatment and every 3-6 months thereafter. Height/length, weight, and weight for height were expressed as standard deviation scores (SDSs) according to WHO child growth standards. RESULTS: The mean SDS of length/height in the early treatment group was significantly higher than that in the later treatment group throughout the observation period (all P < 0.001). The change in the length SDS between the two groups at 1 year old and 4 years old was 1.50 (95% CI, 0.88-2.13) and 0.63 (95% CI, 0.16-1.10), respectively. Compared to the later treatment group, the weight SDS in the early treatment group increased by 0.94 (95% CI, 0.37-1.52) at 1 year old and 0.84 (95% CI, 0.28-1.39) at 2 years old. No statistical significance was found after 2.5 years of age. No significant differences were observed in IGF-1, incidence of liver dysfunction, hypothyroidism or spinal deformity between the two groups. CONCLUSIONS: rhGH treatment improved growth and body composition in infants and toddlers. Furthermore, an early start of rhGH treatment is expected to have more efficacy than the later treatment group without an increase in adverse effects.

Detection of BCR-ABL Fusion Gene in Chronic Myeloid Leukemia by Novel Digital PCR / 中国实验血液学杂志

OBJECTIVE@#To establish a new digital polymerase chain reaction (dPCR) system for the detection of BCR-ABL fusion gene in patients with chronic myeloid leukemia (CML), and explore its analytical performance and clinical applicability in the detection of BCR-ABLp190/210/230.@*METHODS@#A new dPCR system for detecting BCR-ABLp190/210/230 was successfully developed, and its sensitivity difference with qPCR and improvement of drug side effects in patients with CML during drug reduction or withdrawal were compared.@*RESULTS@#Among 176 samples, qPCR and dPCR showed high consistency in the sensitivity of detecting BCR-ABL (82.39%), and the positive rate of dPCR was about 5 times higher that of qPCR (20.45% vs 3.98%). During follow-up, blood routine (25% vs 10%), kidney/liver/stomach (25% vs 20%) and cardiac function (10% vs 0) were significantly improved after drug reduction or withdrawal in patients with initial dPCR negative compared with before drug reduction or withdrawal.@*CONCLUSIONS@#This new dPCR detection system can be applied to the detection of BCR-ABLp190/210/230. It has better consistency and higher positive detection rate than qPCR. Drug withdrawal or dose reduction guided by dPCR has a certain effect on improving drug side effects.

Case Report: Mucolipidosis II and III Alpha/Beta Caused by Pathogenic Variants in the GNPTAB Gene (Mucolipidosis).

Objective: This study aimed to improve the cognition of mucolipidosis (ML) II and III alpha/beta by analyzing the clinical manifestations of two patients. Methods: The clinical, biochemical, and molecular data of two clinical cases associated with ML II and III alpha/beta were analyzed and compared with other case reports of ML II and III alpha/beta. Results: The first patient was a 14-month-old girl who was hospitalized because of abnormal postnatal coarse facial features. The child had no abnormal birth history, but developed multiple abnormalities such as psychomotor retardation, abnormal facial features, bilateral limb muscle hypotonia, and genital abnormalities. The X-ray of the spine revealed multiple bone malformations. Brain magnetic resonance imaging (MRI) showed delayed myelination. Genetic testing showed the presence of two compound heterozygous pathogenic variants (c.1364C>T and c.1284+1G>T) in the GNPTAB gene. The second patient was an 18-month-old boy who was hospitalized for recurrent respiratory tract infections. The patient was a high-risk preterm infant with postnatal psychomotor retardation, language development retardation, intellectual disability, and coarse facial features. X-ray showed multiple bone malformations. Craniocerebral ultrasound showed bilateral ventricle widening. Genetic testing showed the presence of two compound heterozygous pathogenic variants (c.1284+1G>T and c.483delT) in the same gene. Conclusions: ML II and III alpha/beta are rare autosomal-recessive lysosomal storage diseases that are attributed to GNPTAB variants that cause N-acetylglucosamine-1-phosphotransferase deficiency, finally leading to multiple clinical signs and symptoms. A proper ML II and/or III alpha/beta diagnosis requires a combined analysis of a patient's clinical manifestations, imaging examination, enzymatic analysis, and genetic testing results. Ultimately, genetic counseling is essential for this disease.

Gitelman syndrome combined with growth hormone deficiency: Three cases report.

RATIONALE: Gitelman syndrome (GS) is a rare autosomal recessive hereditary salt-losing tubulopathy caused by loss-of-function mutations in the SLC12A3 gene. It is usually characterized by hypokalemia, metabolic alkalosis, hypomagnesemia, and hypocalciuria. There are only a few reports on GS combined with growth hormone deficiency (GHD). PATIENT CONCERNS: Three patients presented with weakness, spasm, and growth retardation, respectively. DIAGNOSES: GS was diagnosed based on the clinical symptoms, laboratory test results, and genetic analysis. GH stimulation tests were performed when the magnesium level returned to normal under magnesium oxide (MgO) therapy. INTERVENTIONS: Initially, all patients received oral replacement of MgO and potassium chloride, and 2 of them received simultaneous spironolactone therapy. Recombinant human growth hormone (rhGH) therapy was initiated after they were diagnosed with GHD. OUTCOMES: All 3 patients exhibited satisfactory growth velocity and normal serum magnesium level, although the potassium level was still slightly lower than normal. LESSONS: We suggest that all GS patients should undergo genetic evaluation, especially regarding SLC12A3 gene mutation. GHD should be considered if these patients have short stature. rhGH therapy is useful for stimulating the patients' growth, and it may increase the serum magnesium level.

WT1 mutation as a cause of 46 XY DSD and Wilm's tumour: a case report and literature review.

AIM: The Wilms' Tumour gene is thought to have tumour suppressor activity and to play an important role in nephrogenesis, genitourinary development, haematopoiesis and sex determination. WT1 mutations will impair gonadal and urinary tract development and have been demonstrated to cause syndromes of WAGR, Denys-Drash and Fraiser. METHODS: To elucidate the role of constitutional mutations of WT1, in the expression of the different clinical feature, we describe a 14-year-9-month nonmosaic XY sex-reversed woman with pure gonadal dysgenesis (46, XY karyotype, completely female external genitalia, normal Mullerian ducts, absence of Wolffian ducts, streak gonads) who had right kidney removed at 7 months of age because of Wilms' tumour and was diagnosed as secondary thrombocytopenia (Plt 60-80 × 10(9) /L) since she was 4 years old. We sequenced the genomic DNA of all the 10 exons of the WT1 in which mutations may occur in proposita. RESULTS: A new de novo insertion mutation in the first exon was found. A 'GCCGCCTCACTCC' is inserted between codon 138 and 139, resulting in the creation of a stop codon and a truncated protein. CONCLUSION: The present data provide further evidence to support the role of WT1 in diverse cellular functions.

[A 10-year review of childhood type 1 diabetes mellitus and the clinical value of interleukin-10 in diabetic ketoacidosis].

OBJECTIVE: To review the incident status of childhood type 1 diabetes mellitus hospitalized in the Children's Hospital of Zhejiang University School of Medicine from 1999 to 2009 and to explore the clinical value of IL-10 in diabetic ketoacidosis. METHODS: The clinical data of 263 children with type 1 diabetes mellitus hospitalized in the Children's Hospital of Zhejiang University School of Medicine from January 1999 to February 2009 were retrospectively reviewed. Serum lipid levels were measured in 48 children with type 1 diabetes mellitus and in 24 healthy children. The diabetic children were classified into two subgroups, with or without ketoacidosis. Serum lipid and cytokines levels were compared. RESULTS: Childhood type 1 diabetes mellitus was common in females (56.3%). The peak incident age of the disease was between 6 and 11.9 years. Diabetic ketoacidosis was as the presenting symptom for the first visit in 86 cases (32.7%). The levels of serum lipid, blood glucose and HbA1c in diabetic children with ketoacidosis were significantly higher than those without ketoacidosis (P<0.05). Logistic analysis demonstrated that the increased levels of blood glucose, serum lipid and HbA1c were risk factors for diabetic ketoacidosis. The level of serum IL-10 in diabetic children with ketoacidosis was significantly higher than that in patients without ketoacidosis (P<0.01), while there were no differences in serum levels IL-2, IL4, IL-6, TNF-α and IFN-γ between them. Serum levels IL-2, IL-4, IL-6, IL-10, TNF-α and IFN-γ in diabetic children were significantly higher than those in healthy children (P<0.01). CONCLUSIONS: Ketoacidosis is a common acute complication of type 1 diabetes mellitus. The disorders of glucose and lipid metabolism are the risk factors for ketoacidosis in diabetic children. IL-10 may be a sensitive index of diabetic ketoacidosis in children with type 1 diabetes mellitus.