Objective:To investigate the screening, clinical and genetic characteristics and prognosis of hereditary tyrosinemia type Ⅰ (HT-Ⅰ) in some areas of Shanghai, and to summarize the relevant characteristics of Chinese cases reported at home and abroad.Methods:From December 2010 to May 2023, the clinical data of children diagnosed with HT-Ⅰ by tandem mass spectrometry combined with genetic detection in Neonatal Screening Center of Shanghai Children′s Hospital were retrospectively analyzed, and the relevant literature was reviewed.Results:A total of 282 149 neonates were screened for genetic metabolic disease by tandem mass spectrometry, and 1 case of HT-Ⅰ was diagnosed, with an incidence of 1∶282 149. Complex heterozygous mutations of FAH genes c. 974C>T and c. 22G>T were found by genetic testing. c. 22G>T was not reported as a new mutation. Diet and drug therapy were given immediately after diagnosis. At present, the follow-up was up to 8 months, and the physical and intellectual development were normal. A total of 32 literatures meeting the inclusion criteria were obtained through database search, and 46 cases of HT-Ⅰ Chinese children were reported. Most of the clinical manifestations were abdominal distension, poor appetite, jaundice, etc., accompanied by different degrees of abnormal coagulation function, hepatosplenomegalysis, cirrhosis and even liver failure. A total of 25 alleles were reported, and the variation of c. 455G>A was the most common. Conclusions:HT-Ⅰ is rare in the population of Shanghai, China, and new mutations enrich the variation spectrum of HT-Ⅰ, which provides basis for family genetic counseling and prenatal diagnosis of children.

Objective:To analyze the differences in screening neonatal glucose-6-phosphate dehydrogenase (G6PD) deficiency in different birth seasons, establish screening thresholds for G6PD concentration in each season using indirect methods, and verify the reliability of the results.Methods:This was a cross-sectional study.A total of 140 823 newborns were collected from the Neonatal Screening Center of Shanghai Children′s Hospital from January 2020 to December 2023, including 41 029 cases, 35 796 cases, 33 969 cases and 30 029 cases in spring, summer, autumn and winter groups, respectively.The concentration of G6PD on the dried blood filter paper was determined using an automatic fluorescence analyzer.The distribution and statistical index of concentration values in four seasons were analyzed.The Kolmogorov-Smirnov test was used for normal distribution.The skewed distribution data was converted into approximately normal distribution using Box-Cox.Outliers were eliminated using the interquartile range (Turkey) method.The cumulative frequency distribution map was drawn through R language programming.The linear regression equation Y=B X+ A was fitted.The 0.5th percentile ( P0.5) was used as the screening threshold, which was compared with the reference value given by the manufacturer or laboratory and with the reference change value (RCV). Results:In the spring group, the positive rate was 4.02‰, 91 cases were confirmed, and the incidence was 1∶451.In the summer group, the positive rate was 7.18‰, 90 cases were confirmed, and the incidence was 1∶398.In the autumn group, the positive rate was 3.21‰, 86 cases were confirmed, and the incidence was 1∶395.In the winter group, the positive rate was 2.26‰, 61 cases were confirmed, and the incidence was 1∶492.The incidence rate did not change significantly in the four seasons ( P>0.05).The G6PD concentrations in the four seasons were compared in pairs, and the result was winter>autumn>spring>summer.The thresholds for G6PD screening were established indirectly: 25.08 U/dL, 22.83 U/dL, 26.63 U/dL and 38.01 U/dL in spring, summer, autumn and winter groups, respectively.The relative deviation in the threshold between the summer group and the laboratory was lower than RCV, while that between the other groups was higher than RCV.According to the screening threshold, the negative and positive conformity rates of 12 batches of 120 samples in the inter-laboratory evaluation program of Chinese Taiwan Preventive Medicine Foundation of China reached 100%. Conclusions:There is no difference in the incidence of G6PD deficiency between birth seasons.It is feasible to establish the screening threshold in each season using indirect methods, which is conducive to improving the efficiency of screening.

Objective:To analyze the differences in screening neonatal glucose-6-phosphate dehydrogenase (G6PD) deficiency in different birth seasons, establish screening thresholds for G6PD concentration in each season using indirect methods, and verify the reliability of the results.Methods:This was a cross-sectional study.A total of 140 823 newborns were collected from the Neonatal Screening Center of Shanghai Children′s Hospital from January 2020 to December 2023, including 41 029 cases, 35 796 cases, 33 969 cases and 30 029 cases in spring, summer, autumn and winter groups, respectively.The concentration of G6PD on the dried blood filter paper was determined using an automatic fluorescence analyzer.The distribution and statistical index of concentration values in four seasons were analyzed.The Kolmogorov-Smirnov test was used for normal distribution.The skewed distribution data was converted into approximately normal distribution using Box-Cox.Outliers were eliminated using the interquartile range (Turkey) method.The cumulative frequency distribution map was drawn through R language programming.The linear regression equation Y=B X+ A was fitted.The 0.5th percentile ( P0.5) was used as the screening threshold, which was compared with the reference value given by the manufacturer or laboratory and with the reference change value (RCV). Results:In the spring group, the positive rate was 4.02‰, 91 cases were confirmed, and the incidence was 1∶451.In the summer group, the positive rate was 7.18‰, 90 cases were confirmed, and the incidence was 1∶398.In the autumn group, the positive rate was 3.21‰, 86 cases were confirmed, and the incidence was 1∶395.In the winter group, the positive rate was 2.26‰, 61 cases were confirmed, and the incidence was 1∶492.The incidence rate did not change significantly in the four seasons ( P>0.05).The G6PD concentrations in the four seasons were compared in pairs, and the result was winter>autumn>spring>summer.The thresholds for G6PD screening were established indirectly: 25.08 U/dL, 22.83 U/dL, 26.63 U/dL and 38.01 U/dL in spring, summer, autumn and winter groups, respectively.The relative deviation in the threshold between the summer group and the laboratory was lower than RCV, while that between the other groups was higher than RCV.According to the screening threshold, the negative and positive conformity rates of 12 batches of 120 samples in the inter-laboratory evaluation program of Chinese Taiwan Preventive Medicine Foundation of China reached 100%. Conclusions:There is no difference in the incidence of G6PD deficiency between birth seasons.It is feasible to establish the screening threshold in each season using indirect methods, which is conducive to improving the efficiency of screening.

Objective:To investigate the screening, clinical and genetic characteristics and prognosis of hereditary tyrosinemia type Ⅰ (HT-Ⅰ) in some areas of Shanghai, and to summarize the relevant characteristics of Chinese cases reported at home and abroad.Methods:From December 2010 to May 2023, the clinical data of children diagnosed with HT-Ⅰ by tandem mass spectrometry combined with genetic detection in Neonatal Screening Center of Shanghai Children′s Hospital were retrospectively analyzed, and the relevant literature was reviewed.Results:A total of 282 149 neonates were screened for genetic metabolic disease by tandem mass spectrometry, and 1 case of HT-Ⅰ was diagnosed, with an incidence of 1∶282 149. Complex heterozygous mutations of FAH genes c. 974C>T and c. 22G>T were found by genetic testing. c. 22G>T was not reported as a new mutation. Diet and drug therapy were given immediately after diagnosis. At present, the follow-up was up to 8 months, and the physical and intellectual development were normal. A total of 32 literatures meeting the inclusion criteria were obtained through database search, and 46 cases of HT-Ⅰ Chinese children were reported. Most of the clinical manifestations were abdominal distension, poor appetite, jaundice, etc., accompanied by different degrees of abnormal coagulation function, hepatosplenomegalysis, cirrhosis and even liver failure. A total of 25 alleles were reported, and the variation of c. 455G>A was the most common. Conclusions:HT-Ⅰ is rare in the population of Shanghai, China, and new mutations enrich the variation spectrum of HT-Ⅰ, which provides basis for family genetic counseling and prenatal diagnosis of children.

Objective:To analyze the difference and reliability of blood 17-hydroxyprogesterone (17-OHP), an indirect screening index for congenital adrenal hyperplasia (CAH), between preterm and full-term infants.Methods:In this retrospective cross-sectional study, a total of 210 285 newborns who underwent CAH screening at the Neonatal Screening Center of Shanghai Children′s Hospital from January 2019 to December 2022 were collected, including 14 312 premature infants and 195 973 full-term infants.The concentration of 17-OHP in dried blood spots on filter paper was determined by an automatic fluorescence analyzer.The distribution of 17-OHP levels in preterm and full-term infants and its statistical index were analyzed.The Kolmogorov-Smirnov test was used for normal distribution.The skewed distribution data was converted into approximately normal distribution using Box-Cox.Outliers were eliminated by the interquartile range method.The cumulative frequency distribution map was drawn by R language programming.The 99.5 th percentile value was used as the screening threshold and compared with the reference value given by the manufacturer or laboratory and with the reference change value (RCV). Results:According to the threshold provided by the laboratory, 26.76‰ of premature infants were tested positive in preliminary screening, and 4 were confirmed with an incidence of 1∶3 578, while 0.79‰ of full-term infants were tested positive in preliminary screening, and 11 were confirmed with an incidence of 1∶17 816.The thresholds for CAH screening established indirectly were 20.35 nmol/L in preterm infants and 10.78 nmol/L in full-term infants.The relative deviations between the indirect CAH screening thresholds and the manufacturer′s or laboratory′s CAH screening thresholds were higher than the RCV, respectively.According to the indirect CAH screening thresholds, the negative and positive coincidence rates of 65 samples in 13 batches from the Centers for Disease Control and Prevention interlaboratory quality assessment program in the United States reached 100%.A retrospective analysis of 210 285 neonates showed that 17-OHP concentration was higher than the screening threshold in all CAH-positive neonates.The application of this screening threshold reduced the false positive rate of preterm infants by 59.79%.Conclusions:It is feasible to establish the CAH screening thresholds for premature and full-term infants by an indirect method, which can improve the efficiency of screening and provide better diagnostic basis for clinical practice.

Objective:To explore the clinical, biochemical and genetic characteristics of three children with Isoleucine metabolic disorders due to variants of HSD17B10 and ACAT1 genes. Methods:Two children with 17β hydroxysteroid dehydrogenase 10 (HSD17B10) deficiency and a child with β-ketothiolase deficiency (BKD) diagnosed at Shanghai Children′s Hospital between 2014 and 2021 were selected as the study subjects. Clinical data of the children were collected. The children were subjected to blood acylcarnitine, urinary organic acid and genetic testing, and candidate variants were analyzed with bioinformatic tools.Results:The main symptoms of the three children had included epilepsy, developmental delay, hypotonia and acidosis. Their blood acylcarnitine methylcrotonyl carnitine (C5: 1), 3-hydroxyisovalerylcarnitine (C5-OH) and 3-hydroxybutylcarnitine (C4OH) were increased to various extents, and urine organic acids including methyl crotonylglycine and 2-methyl-3-hydroxybutyric acid were significantly increased. Child 1 and child 2 were respectively found to harbor a c. 347G>A (p.R116Q) variant and a c. 274G>A (p.A92T) variant of the HSD17B10 gene, and child 3 was found to harbor compound heterozygous variants of the ACAT1 gene, namely c. 547G>A (p.G183R) and a c. 331G>C (p.A111P). Among these, the c. 274G>A (p.A92T) and c. 331G>C (p.A111P) variants were unreported previously. Based on the guidelines from the American College of Medical Genetics and Genomics (ACMG), they were respectively classified as variant of unknown significance (PP3_Strong+ PM2_supporting) and likely pathogenic (PM3+ PM2_Supporting+ PP3_Moderate+ PP4). Conclusion:Both the HSD17B10 deficiency and BKD can lead to Isoleucine metabolism disorders, which may be difficult to distinguish clinically. Genetic testing can further confirm the diagnosis. Discoveries of the HSD17B10: c. 274G>A (p.A92T) variant and the ACAT1: c. 331G>C (p.A111P) variant have enriched the mutational spectrum of the two diseases.

Objective:To investigate the diagnostic value of fluorescence quantitative method and G6PD/6PGD ratio method in glucose-6-phosphate dehydrogenase (G6PD) deficiency and the type of gene mutation.Methods:A total of 1 201 patients (711 males and 490 females) with suspected G6PD deficiency in Shanghai Children′s Hospital were collected from June 2018 to March 2021. Fluorescence quantification method, G6PD/6PGD ratio method and multicolor melting curve were used to detects enzyme activity, ratio and gene mutation type. Comparison of each index and evaluation of its diagnostic efficiency were performed.Results:Among 1 201 suspicious samples, 163 cases (135 males and 28 females) were finally diagnosed. 156 cases were diagnosed by fluorescence quantitative method with a detection rate of 95.71%, and 140 cases were diagnosed by G6PD/6PGD ratio method with a detection rate of 85.89%. enzymatic activity of G6PD and ratio of G6PD/6PGD in male were significantly lower than female, and the differences were statistically significant ( U=642.5, 734.5, P<0.001). 112 cases received G6PD gene mutation detection and 92 cases were diagnosed, 74 were hemizygous mutations, 1 were homozygous mutations, 15 were heterozygous mutations, and 2 were compound heterozygous mutations. Among 15 cases of heterozygous mutations, 11 cases were diagnosed by fluorescence quantitative method, the diagnosed rate was 73.33%, 4 cases were diagnosed by G6PD/6PGD ratio method, and the diagnosed rate was 26.67%. A total of 7 mutation sites were detected and the proportions were c.1388G>A (32.22%), c.1376G>T (30.00%), c.871G>A (13.33%), c.1024C>T (11.11%). c.95A>G (7.78%), c.487G>A (4.44%), c.392G>T (1.11%). The enzymatic activities of c.1376G>T and c.1024C>T, c.487G>A were statistically significant ( P<0.001,0.015); the G6PD/6PGD ratios of c.1024C>T and c.1388G>A, c.1376G>T were statistically significant ( P=0.017,0.002,0.011,0.013). Fluorescence quantitative method had sensitivity of 100%, specificity of 95.65%, and the area under the curve (AUC) is 0.972. The sensitivity of the G6PD/6PGD ratio method was 100%, the specificity was 94.57%, and the AUC was 0.979. The sensitivity of fluorescence quantitative method combined with G6PD/6PGD ratio was 96.7%, the specificity was 100%, and the AUC was 0.992. Conclusions:Compared with fluorescence quantification, the G6PD/6PGD ratio method might not be able to diagnose female heterozygotes effectively; The panel of G6PD fluorescence quantification and G6PD/6PGD ratio was helpful to reduce the missed diagnosis. Combined with gene mutation analysis, it could improve the diagnosis rate of G6PD deficiency in the children.

Objective:To establish a disease risk prediction model for the newborn screening system of inherited metabolic diseases by artificial intelligence technology.Methods:This was a retrospectively study. Newborn screening data ( n=5 907 547) from February 2010 to May 2019 from 31 hospitals in China and verified data ( n=3 028) from 34 hospitals of the same period were collected to establish the artificial intelligence model for the prediction of inherited metabolic diseases in neonates. The validity of the artificial intelligence disease risk prediction model was verified by 360 814 newborns ' screening data from January 2018 to September 2018 through a single-blind experiment. The effectiveness of the artificial intelligence disease risk prediction model was verified by comparing the detection rate of clinically confirmed cases, the positive rate of initial screening and the positive predictive value between the clinicians and the artificial intelligence prediction model of inherited metabolic diseases. Results:A total of 3 665 697 newborns ' screening data were collected including 3 019 cases ' positive data to establish the 16 artificial intelligence models for 32 inherited metabolic diseases. The single-blind experiment ( n=360 814) showed that 45 clinically diagnosed infants were detected by both artificial intelligence model and clinicians. A total of 2 684 cases were positive in tandem mass spectrometry screening and 1 694 cases were with high risk in artificial intelligence prediction model of inherited metabolic diseases, with the positive rates of tandem 0.74% (2 684/360 814)and 0.46% (1 694/360 814), respectively. Compared to clinicians, the positive rate of newborns was reduced by 36.89% (990/2 684) after the application of the artificial intelligence model, and the positive predictive values of clinicians and artificial intelligence prediction model of inherited metabolic diseases were 1.68% (45/2 684) and 2.66% (45/1 694) respectively. Conclusion:An accurate, fast, and the lower false positive rate auxiliary diagnosis system for neonatal inherited metabolic diseases by artificial intelligence technology has been established, which may have an important clinical value.

BACKGROUND:To improve local microenvironment and reduce local scars is conducive to peripheral nerve regeneration that promotes nerve function recovery.OBJECTIVE:To evaluate the effect of fresh amniotic membrane on the regeneration of tinjured peripheral nerve.METHODS:Sixty healthy Sprague-Dawley rats were randomly divided into three groups (n=20 per group) after constructing a model of sciatic nerve injury of the unilateral leg. In group A, the nerve was wrapped with fresh human amnion at the anastomosis end after the repair of nerve. In group B, the nerve was wrapped with biofilm at the anastomosis end after the repair of nerve. In group C, no treatment was conducted after the repair of nerve (blank control). The effects were evaluated by anatomical observation, light microscope observation, immunohistochemical detection (2, 4, 8, 12 weeks after surgery), transmission electron microscope observation, axon imaging analysis, action potential detection, and sciatic nerve function index (4, 8, 12 weeks after surgery).RESULTS AND CONCLUSION: (1) Gross observation. The amniotic membrane and biofilm were absorbed partialy at postoperative 2 weeks, mostly at postoperative 4 weeks and completely at postoperative 8 weeks. In the groups A and B, the nerve was adhered slightly and loosely to the surrounding tissues, with a fair range of motion. In the group C, the nerve was tightly adhered to the surrounding tissues, with a poor range of motion. (2) Observation under light microscope. The nerve regeneration was better in the groups A and B than group C at 2, 4, 8, 12 weeks postoperatively. (3) Observation under electron microscope. Regenerated nerve fibers were rarely seen and lamelar structures were unclear in the three groups at 4 weeks postoperatively. Then, increased regenerated nerve fibers, thickened myelin sheath, clear lamelar structure and enlarged axon diameter were found in the groups A and B compared with the group C at 8 and 12 weeks postoperatively. (4) Immunohistochemical detection. The expression and distribution of S-100 protein in the groups A and B were better than those in the group C. (5) Axon image analysis. Groups A and B were superior to the group C in the diameter of myelinated nerve fibers, thickness of myelin sheath and number of regenerated nerve fibers. There was a significant difference by statistical analysis (P < 0.05). (6) Electrophysiological examination. Shorter latency period, higher amplitude and faster nerve conduction velocities were observed in the groups A and B compared with the group C (P < 0.05). (7) The sciatic function index. The sciatic function index in group A or B was significantly higher than that in group C (P < 0.05). To conclude, the human amniotic membrane can reduce adhesion between the damaged nerve and surrounding tissues, and prevent scarring at the anastomosis end. In addition, it promotes the regeneration of nerve fibers, increase axon diameter and myelin sheath thickness, and ease inflammatory and immune responses at the neural incision.

Objective To evaluate the feasibility of PerkinElmer Genetic Screening Processor(GSP) in the application of newborn screening for congenital hypothyroidism (CH) and congenital adrenal hyperplasia (CAH) by detecting thyroid-stimulating hormone (TSH) and 17-OH-progesterone(17-OHP).Methods The dried-blood spots specimens from Centers for Disease Control and Prevention(CDC) and the quality control in the reagent kit were detected and the accuracy,precision and linearity were calculated.A total of 1 012 samples of TSH(60 of positive and 952 negative samples) and 991 samples of 17-0HP(34 positive and 957 negative samples)were detected.The initial cut-off value was determined by ROC curve determined.The consistency between the results from GSP and clinical diagnosis was analyzed.Results The average of within-run coefficient of variation(CV) of TSH and 17-OHP were 6.69％ to 12.6％ and 7.52％ to 9.29％,and the average of between-run CV were 6.91％ to 10.96％ and 6.86％ to 12.36％,respectively.The average of bias of TSH and 170HP were-14.28％ to-0.74％ and-0.45％ to 12.54％.The linearity of GSP detection was fine.The initial cut-off values were 23.43 U/mL(TSH) and 21.42 ng/mL(17-OHP).The sensitivity of GSP detection was 100％ and the specificity of TSH and 17-OHP were 98.11％ and 99.58 ％ respectively.The results of GSP detection showed good consistency with clinical diagnosis.Conclusion As the first real automatic fluorescence immunoassay analyzer,GSP could be used in routine clinical diagnosis for CH and CAH.