Human thymus in health and disease: Recent advances in diagnosis and biology

The thymus is the crucial tissue where thymocytes develop from hematopoietic precursors that originate from the bone marrow and differentiate to generate a repertoire of mature T cells able to respond to foreign antigens while remaining tolerant to self-antigens. Until recently, most of the knowledge on thymus biology and its cellular and molecular complexity have been obtained through studies in animal models, because of the difficulty to gain access to thymic tissue in humans and the lack of in vitro models able to faithfully recapitulate the thymic microenvironment. This review focuses on recent advances in the understanding of human thymus biology in health and disease obtained through the use of innovative experimental techniques (eg. single cell RNA sequencing, scRNAseq), diagnostic tools (eg. next generation sequencing), and in vitro models of T-cell differentiation (artificial thymic organoids) and thymus development (eg. thymic epithelial cell differentiation from embryonic stem cells or induced pluripotent stem cells).     

先天性遗传代谢病的早期诊断

目的提高儿科医生对新生儿期遗传代谢病的认识，做到早期诊断、早期治疗。方法自2003年9月至2004年9月，根据临床表现确定18名遗传代谢病高危患儿，用“滤纸片代”将采集的尿标本外寄进行气相色谱．质谱(GC／MS)分析，筛查遗传代谢病。结果18例高危儿中确诊为遗传代谢病5例，分别为戊二酸尿症Ⅱ型1例(46h，男)，鸟氨酸氨甲酰转移酶缺陷1例(66h，男)，枫糖尿病1例(8d，男)，甲基丙二酸血症1例(13d，男)，丙酸血症1例(21d，女)，并对其临床特点进行归纳总结。结论掌握新生儿遗传代谢病临床特点，对高危儿早期进行尿GC／MS分析，可以早期诊断遗传代谢病，有利于优生优育。     

Identification of five novel inactivating mutations in the human thyroid peroxidase gene by denaturing gradient gel electrophoresis

Thyroid peroxidase (TPO) is the key enzyme in the synthesis of thyroid hormones. Defects in the TPOgene are reported to be the cause of congenital hypothyroidism due to a Total Iodide Organification Defect (TIOD). This type of defect, where iodide taken up by the thyroid gland cannot be oxidized and bound to protein, is the most common hereditary inborn error causing congenital hypothyroidism in the Netherlands. Denaturing Gradient Gel Electrophoresis (DGGE) of PCR amplified genomic DNA was used to screen for mutation in the TPO gene of TIOD patients from nine apparently unrelated families, and seven different mutations were detected. Three frameshift mutations were found: a 20 bp duplication in exon 2, a 4 bp duplication in exon 8, and an insertion of a single nucleotide (C) at pos. 2505 in exon 14. In addition, four single nucleotide substitutions were identified: one single-base, mutation resulted in a premature termination codon (C → T at pos. 1708 in exon 10), two single-base substitutions changed an amino acid in highly conserved regions of the gene (Tyr → Asp in exon 9 and Glu → Lys in exon 14). The fourth single-base mutation located at the exon 10/intron 10 border altered a conserved Gly into Ser and could also affect splicing. Nine TIOD patients from five families were compound heterozygotes and six patients from four families were homozygous for one of the mentioned mutations in the TPO gene. © 1995 Wiley-Liss, Inc.     

Mild inborn errors of metabolism in commonly used inbred mouse strains

Inbred mouse strains are a cornerstone of translational research but paradoxically many strains carry mild inborn errors of metabolism. For example, α-aminoadipic acidemia and branched-chain ketoacid dehydrogenase deficiency are known in C57BL/6J mice. Using RNA sequencing, we now reveal the causal variants in Dhtkd1 and Bckdhb, and the molecular mechanism underlying these metabolic defects. C57BL/6J mice have decreased Dhtkd1 mRNA expression due to a solitary long terminal repeat (LTR) in intron 4 of Dhtkd1. This LTR harbors an alternate splice donor site leading to a partial splicing defect and as a consequence decreased total and functional Dhtkd1 mRNA, decreased DHTKD1 protein and α-aminoadipic acidemia. Similarly, C57BL/6J mice have decreased Bckdhb mRNA expression due to an LTR retrotransposon in intron 1 of Bckdhb. This transposable element encodes an alternative exon 1 causing aberrant splicing, decreased total and functional Bckdhb mRNA and decreased BCKDHB protein. Using a targeted metabolomics screen, we also reveal elevated plasma C5-carnitine in 129 substrains. This biochemical phenotype resembles isovaleric acidemia and is caused by an exonic splice mutation in Ivd leading to partial skipping of exon 10 and IVD protein deficiency. In summary, this study identifies three causal variants underlying mild inborn errors of metabolism in commonly used inbred mouse strains.     

Pharmacological evidence that α-ketoisovaleric acid induces convulsions through GABAergic and glutamatergic mechanisms in rats

Neurological dysfunction is common in patients with maple syrup urine disease (MSUD). However, the mechanisms underlying the pathophysiology of this disorder are poorly known. In the present study we investigated the effect of intrastriatal administration of the α-keto acids accumulating in MSUD on the behavior of adult rats. After cannula placing, rats received unilateral intrastriatal injections of α-ketoisocaproic acid (KIC, 8 μmol), α-ketoisovaleric acid (KIV, 8 μmol), α-keto-β-methylvaleric acid (KMV, 6 μmol) or NaCl. KIV elicited clonic convulsions in a dose–response manner, whereas KIC and KMV did not induce seizure-like behavior. Convulsions provoked by KIV were prevented by intrastriatal preadministration of muscimol (46 pmol) and MK-801 (3 nmol), but not by the preadministration of DNQX (8 nmol). These results indicate that among the keto acids that accumulate in MSUD, KIV is the only metabolite capable of causing convulsions in the present animal model and indicates that KIV is an important excitatory metabolite. Moreover, the participation of GABAergic and glutamatergic NMDA mechanisms in the KIV-induced convulsant behavior is suggested, since KIV-induced convulsions are attenuated by muscimol and MK-801. The authors suggest that KIV may play an important role in the convulsions observed in MSUD, and highlight its relevance to the understanding of the pathophysiology of the neurological dysfunction of MSUD patients.     

经高危筛查发现的遗传性代谢病15例分析

目的：提高临床医生对非特异性临床表现的遗传性代谢病的认识,并通过实验室检查早期诊断、早期治疗,减少后遗症。方法：对2003年6月1日至2006年9月30日期间入住上海交通大学医学院附属新华医院儿内科病房的132例非特异性临床表现的高危儿,在常规进行临床生化检查的同时行血串联质谱和尿气相质谱检测。结果：132例中诊断遗传性代谢病15例（11.5%）。其中甲基丙二酸血症(MMA)6例(40％);丙酸血症2例（13.3%）;瓜氨酸血症-II型2例（13.3%）;生物素酶缺乏症1例（6.7%）;酪氨酸血症1例（6.7%）;枫糖尿病1例（6.7%）;鸟氨酸氨甲酰转移酶缺乏症1例（6.7%）;极长链酰基肉碱辅酶A脱氢酶缺乏症1例（6.7%）。结论：对非特异性临床表现疑似遗传性代谢病的高危儿应及时进行串联质谱及气相质谱检查有助于遗传性代谢病的检出。     

Scavenger treatment of free radical injury in familial amyloidotic polyneuropathy: a study on Swedish transplanted and non-transplanted patients

Since oxidative stress has been implicated in amyloid diseases, a study of scavenger treatment of hereditary transthyretin amyloidosis was undertaken on 23 familial amyloidotic polyneuropathy (FAP) patients. Nine patients had undergone a liver transplantation for the disease. Twenty patients completed the 6-month study period of scavenger treatment (vitamin C, 1 g, three times daily, vitamin E, 0.1 g, three times daily and acetylcysteine, 0.2 g three times daily). They were evaluated clinically and by immunohistochemical measurement of hydroxynonenal (HNE), a product of lipid peroxidation, in biopsy specimens. For non-transplanted patients, no improvement was found for HNE in relation to the amyloid content in biopsy specimens, whereas a tendency to a decreased amount was noted for transplanted patients. Clinically, no differences were found for non-transplanted patients, but an increased nutritional status, measured by a modified body mass index (mBMI) was noted for transplanted patients. In summary, scavenger treatment with the drugs and doses used in the present study appears to be unable to decrease lipid peroxidation in amyloid-rich tissue in non-transplanted FAP patients. For transplanted patients, lipid peroxidation tended to decrease, and the nutritional status measured by mBMI improved, even though the findings may be explained by liver transplantation alone, scavenger treatment may facilitate recovery after transplantation.     

Methylmalonic acidemia: brain imaging findings in 52 children and a review of the literature

BACKGROUND: Methylmalonic acidemia (MMA) is an autosomal-recessive inborn error of metabolism. OBJECTIVE: To recognize the CT and MR brain sectional imaging findings in children with MMA. MATERIALS AND METHODS: Brain imaging studies (47 MR and 5 CT studies) from 52 children were reviewed and reported by a neuroradiologist. The clinical data were collected for each patient. RESULTS: The most common findings were ventricular dilation (17 studies), cortical atrophy (15), periventricular white matter abnormality (12), thinning of the corpus callosum (8), subcortical white matter abnormality (6), cerebellar atrophy (4), basal ganglionic calcification (3), and myelination delay (3). The brain images in 14 patients were normal. CONCLUSION: Radiological findings of MMA are nonspecific. A constellation of common clinical and radiological findings should raise the suspicion of MMA.     

Reliability of urine collection pads for routine and metabolic biochemistry in infants and young children

The aim of our study was to evaluate whether specifically designed urine collection pads give reliable results for routine and metabolic biochemistry tests in paediatric urine. Urine collected by bag or clean-catch from infants and children <2 yrs without metabolic disorders was divided into two aliquots, one of which was added to a collection pad, incubated for 15 min at 37°C (simulating in vivo collection conditions), then recovered by aspiration. Urine from adults with phaeochromocytoma and aqueous solutions of catecholamines were similarly treated. Routine, catecholamine, and metabolic analyses were performed on pad/non-pad aliquots. Selected metabolic analyses were also performed on pad/non-pad urine from patients with diagnosed inborn errors and urine containing added metabolites to simulate metabolic disorders. Routine tests (urea, electrolytes, creatinine, osmolality, calcium:creatinine, phosphate:creatinine, magnesium:creatinine, urate:creatinine [n = 32], oxalate:creatinine [n = 10]), and catecholamines (n = 12) showed good or acceptable concordance with no clinically significant pad/non-pad differences. Metabolic tests in infants and children without metabolic disorders all showed good pad/non-pad concordance for amino acids (n = 10), organic acids (n = 12), and glycosaminoglycans (n = 8). In patients with metabolic disorders (phenylketonuria [n = 1], homozygous/heterozygous cystinuria [n = 3], mucopolysaccharidoses II [n = 2] and III [n = 1], organic acid disorders [n = 6]) and urine containing added orotic acid to simulate urea cycle disorders, there was also good pad/non-pad concordance for diagnostic urinary metabolites. No extraneous organic acids were eluted from the pads. Sugar chromatography showed identical staining intensity in pad/non-pad samples. In conclusion, urine collection pads give reliable results for a wide range of routine and metabolic biochemistry tests in urine from paediatric patients.