Advances in the pathogenesis and biomarker of macrophage activation syndrome in children / 国际儿科学杂志

Macrophage activation syndrome is a life-threatening syndrome of multiple causes secondary to rheumatic immune diseases.It is characterized by the continuous activation and proliferation of T lymphocytes and macrophages that leads to overwhelming immune response and excessive release of pro-inflammatory mediators, which eventually causes cytokine storm and multiple organ failure.The main clinical manifestations and laboratory abnormalities include fever, hemocytopenia, hepatomegaly, splenomegaly, lymph node enlargement, coagulation disorders, liver function damage, hyperferritinemia, hypertriglyceridemia and the phenomenon of phagocytosis of blood cells in bone marrow.This article reviews the progress of epidemiology, pathogenesis and biomarkers in macrophage activation syndrome to provide new insights for early diagnosis and identification of the complication which has a rapid progress and high fatality rate.

Novel compound heterozygous mutations in the PCDH15 gene in a family affected with Usher syndrome type 1F with retinitis pigmentosa sine pigmento / 中华眼底病杂志

Objective:To identify the causative gene in a family affected with Usher syndrome (USH) with retinitis pigmentosa sine pigmento (RPSP) and to analyze the genotype-phenotype correlation.Methods:A retrospective clinical study. A 9-year-old girl with RPSP type 1F USH diagnosed in the ophthalmology clinic of Henan Provincial People's Hospital in November 2019 and her parents were included in the study. The patient had bilateral night blindness for more than 4 years, she suffered from hearing loss 7 years, and is currently binaural sensorineural deafness. The best corrected visual acuity in both eyes was 0.5 +. There was showed no obvious pigmentation on the fundus. The visual acuity of the peripheral field of vision decreased. Optical coherence tomography showed that the outer layer of the peripheral retina became thinner and the ellipsoid band disappeared. On electroretinogram examination, the rod and cone system response was severely decreased. The clinical phenotype of the parents of the child were normal. The peripheral venous blood of the child and his parents were extracted, the whole genome DNA was extracted, the custom developed targeted capture kit (PS400) was used, and the next-generation sequencing technology was used to detect genetic mutations. The suspected pathogenic mutation sites were verified by Sanger; co-segregation was performed among family members. The pathogenicity of variants were evaluated according to the interpretation standards and guidelines of sequence variants. Bioinformatics techniques were used to assess the impact of variants on encoded proteins. Results:The results of genetic testing showed that the proband detected the PCDH15 gene c.4109dupA (p.K1370fs) (M1), c.17dupA (p.Y6_L7delinsX) (M2) compound heterozygous mutation sites, verified by Sanger sequencing, the mutations were in the family in a state of co-segregation. According to the evaluation of sequence variation interpretation standards and guidelines, M1 and M2 were pathogenic variants of the PCDH15 gene. M1 led to a complete change in the transmembrane structure of the encoded protein, and M2 caused the gene to only translate 6 amino acids, which predicted that the PCDH15 protein cannot be synthesized. According to the clinical phenotype, gene mutation pathogenicity and protein structure prediction, the final clinical diagnosis was PCDH15-related type 1F. Conclusions:PCDH15 genes c.4109dupA and c.17dupA are the pathogenic mutation sites of USH in this family. These compound heterozygous new mutations lead to the failure of normal synthesis of PCDH15 protein, which leads to ocular and ear manifestations.

Mutation analysis and phenotypic investigation in patients with primary distal renal tubular acidosis / 中华肾脏病杂志

Objective:To analyze the gene variants in patients with primary distal renal tubular acidosis (dRTA), and explore the correlation between the genotype and phenotype.Methods:The Sanger direct sequencing or whole-exome sequencing was used to identify causal variants and the variation pathogenicity was evaluated according to 2015 American College of Medical Genetics and Genomics (ACMG) standards and guidelines in 44 dRTA patients (37 families) diagnosed in the Affiliated Qingdao Municipal Hospital of Qingdao University and the Affiliated Hospital of Qingdao University from April 2010 to September 2020. The clinical features of the patients were summarized, and the correlation between the genotype and phenotype was investigated.Results:Seven variants of SLC4A1 gene, 17 variants of ATP6V0A4 gene, and 15 variants of ATP6V1B1 gene were identified in 44 patients with dRTA, and of which 11 variants were new ones. According to ACMG guidelines, the pathogenic, likely pathogenic, benign variants among the 39 variants were 22, 16 and 1, respectively. Nine patients were autosomal dominant hereditary dRTA caused by SLC4A1 gene mutation, 4 patients with autosomal recessive hereditary dRTA complicated with Southeast Asian ovalocytosis and anemia were caused by SLC4A1 gene mutation, and 14 patients caused by ATP6V0A4 gene mutation and 8 patients caused by ATP6V1B1 gene mutation were autosomal recessive hereditary dRTA; Two children with dRTA were found to carry one monoallelic defect in ATP6V1B1, and no causal gene mutation was identified in 7 patients. One patient showed incomplete dRTA, and the other 43 patients showed complete dRTA. The prevalence of sensory neural hearing loss caused by ATP6V0A4 and ATP6V1B1 mutation were 2/14 and 6/10 respectively. The frequency of chronic kidney disease in adults, children and infants were 4/4, 2/4, and 1/36, separately. After the drug treatment based on potassium citrate and sodium citrate, the growth and development (28/40) and electrolyte disturbance (41/44) of most patients were significantly improved. Conclusions:The present study has identified 39 variants of SLC4A1, ATP6V0A4 and ATP6V1B1 genes in 44 patients with dRTA, including 11 novel ones. There is a close relationship between genotype and phenotype in dRTA patients and most patients' conditions were improved after proper treatment. This study enriches the human gene mutation database and provides valuable references for diagnosis, treatment and genetic counseling in patients with dRTA.

Mutation analysis of KCNJ1 gene and investigation of phenotype in 5 Chinese patients with Bartter syndrome type 2 / 中华肾脏病杂志

Objective@#To identify and analyze the variants of the KCNJ1 gene in five Chinese patients with Bartter syndrome type 2 (BS2), and to describe their clinical features as well as treatment results.@*Methods@#Data and blood samples of five BS2 patients and their relatives confirmed by Qingdao Municipal Hospital from June 2012 to January 2019 were collected. Whole-exome-sequencing (WES) based on the second generation high throughput sequencing was performed to detect variants. The 2015 American College of Medical Genetics and Genomics Standards and Guidelines were applied to analyze the pathogenicity of the variants. The clinical features and laboratory results were retrospectively studied. The response to treatment and follow-up data were reviewed.@*Results@#Ten variants including six novel ones of KCNJ1 gene were identified through WES and verified by Sanger dideoxy sequencing. Missense variants accounted for the highest proportion. The common symptoms and signs of five BS2 patients from high to low incidence were polydipsia and polyuria (5/5), one of them (1/5) presented with diabetes insipidus; maternal polyhydramnios and premature delivery (4/5); growth retardation (3/5). Initially, two patients presented with hypochloremic metabolic alkalosis and hypokalemia, whereas the acid-base disturbance was absent in the others. One patient experienced hyperkalemia. In terms of calcium-phosphorus metabolism, one patient had evident parathyroid hormone (PTH) resistance (hypocalcemia, hyperphosphatemia and markedly elevated serum intact PTH levels), three presented with PTH overacting (hypercalcemia, hypophosphatemia and mild elevated serum intact PTH levels), and one showed normal blood calcium and phosphorus concentrations with high-normal serum intact PTH levels. All patients had nephrocalcinosis or hypercalciuria, and one of them complicated with nephrolithiasis. Indomethacin helped to correct the growth retardation, halt polydipsia polyuria, decrease the elevated urinary calcium excretion, and normalize electrolyte disturbance as well as PTH parameters in some patients.@*Conclusions@#This investigation identifies ten variants of KCNJ1 gene, including six ones that have not been previously reported, which will enrich the human gene mutation database (HGMD). These patients in our study have atypical BS phenotype, so that careful differentiation from other parathyroid diseases will be required for clinicians.

Mutation analysis of KCNJ1 gene and investigation of phenotype in 5 Chinese patients with Bartter syndrome type 2 / 中华肾脏病杂志

Objective:To identify and analyze the variants of the KCNJ1 gene in five Chinese patients with Bartter syndrome type 2 (BS2), and to describe their clinical features as well as treatment results. Methods:Data and blood samples of five BS2 patients and their relatives confirmed by Qingdao Municipal Hospital from June 2012 to January 2019 were collected. Whole-exome-sequencing (WES) based on the second generation high throughput sequencing was performed to detect variants. The 2015 American College of Medical Genetics and Genomics Standards and Guidelines were applied to analyze the pathogenicity of the variants. The clinical features and laboratory results were retrospectively studied. The response to treatment and follow-up data were reviewed.Results:Ten variants including six novel ones of KCNJ1 gene were identified through WES and verified by Sanger dideoxy sequencing. Missense variants accounted for the highest proportion. The common symptoms and signs of five BS2 patients from high to low incidence were polydipsia and polyuria (5/5), one of them (1/5) presented with diabetes insipidus; maternal polyhydramnios and premature delivery (4/5); growth retardation (3/5). Initially, two patients presented with hypochloremic metabolic alkalosis and hypokalemia, whereas the acid-base disturbance was absent in the others. One patient experienced hyperkalemia. In terms of calcium-phosphorus metabolism, one patient had evident parathyroid hormone (PTH) resistance (hypocalcemia, hyperphosphatemia and markedly elevated serum intact PTH levels), three presented with PTH overacting (hypercalcemia, hypophosphatemia and mild elevated serum intact PTH levels), and one showed normal blood calcium and phosphorus concentrations with high-normal serum intact PTH levels. All patients had nephrocalcinosis or hypercalciuria, and one of them complicated with nephrolithiasis. Indomethacin helped to correct the growth retardation, halt polydipsia polyuria, decrease the elevated urinary calcium excretion, and normalize electrolyte disturbance as well as PTH parameters in some patients. Conclusions:This investigation identifies ten variants of KCNJ1 gene, including six ones that have not been previously reported, which will enrich the human gene mutation database (HGMD). These patients in our study have atypical BS phenotype, so that careful differentiation from other parathyroid diseases will be required for clinicians.

X linked rod-cone dystrophy associated with a novel splicing variation in the intron of OPN1LW gene / 中华实验眼科杂志

Objective:Mutations in the OPN1LW gene located in X chromosome usually lead to blue cone monochromacy.Variations in OPN1LW gene usually occur in the exon region, but was rare in the intron region.This study was to report a Chinese family with X-linked rod-cone dystrophy associated with a novel OPN1LW gene hemizygotic splicing variation and analyze the clinical phenotype and gene mutation characteristics Methods:A pedigree investigation was performed.The family members clinically diagnosed as rod-cone dystrophy with myopia were enrolled in Henan Provincial People's Hospital on January 9, 2020.Detailed ophthalmological examination was carried out, and the periphery venous blood was collected for DNA extraction.The target gene sequencing panel PS400 developed by Henan Clinical Research Center for Eye Diseases and whole exon sequencing were used to detect pathogenic mutations.Sanger sequencing and pedigree co-segregation were used to verify variations.The pathogenicity of the novel variation was analyzed based on American College of Medical Genetics (ACMG) Guidelines and online tools SIFT, Polyphen2, Mutation Taster.This study adhered to the Declaration of Helsinki, and the study protocol was approved by the Medical Ethics Committee of Henan Eye Hospital (HNEECKY-2019 [15]). Written informed consent was obtained from each family member before any medical examination.Results:The proband was a 5-year-old boy with poor vision, red-green blindness and nystagmus in both eyes.No obvious abnormality in ocular anterior segment was found.The boundary of optic disc was clear and the color was reddish, and the reflection of macular fovea was clearly visible.OCT image showed indistinct reflection of some ellipsoids in macular area of both eyes.The amplitudes of a and b waves of full-field ERG were not recorded in scotopic 0.01 scale and significantly reduced in scotopic 3.0 and photopic 3.0 ERG.The uncle of the proband had a more severe clinical phenotype.Wide-angle fundus photography showed high myopia findings, peripheral retinal atrophy and sporadic black lesions, and autofluorescence examination showed attenuated fluorescence in peripheral retina.No obvious abnormality was found in the middle-peripheral retinal region.The results of two kinds next generation sequencing showed a novel hemizygotic splicing variation c. 112+ 2T>G in the intron of OPN1LW gene and an unreported heterozygous variation c. 1913A>C (p.Y638S) in the SEMA4A gene.The c. 112+ 2T>G mutation further leaded to the sequence change of classic splice donor of intron 1.According to the ACMG guidelines, the pathogenicity score was PVS1+ PM2+ PP1, which was considered as a pathogenic level. Conclusions:This is the first report of X-linked rod-cone dystrophy associated with OPN1LW gene variation, and this novel variant c. 112+ 2T>G locates in the intron region.This result is different from past knowledge that variations of OPN1LW gene primarily occur in exon.This study expands the mutational spectrum of OPN1LW gene inducing retinal degeneration and the spectrum of clinical phenotype.

Analysis of USH2A gene mutation and clinical phenotype in families with Usher syndrome type 2 and retinitis pigmentosa / 中华眼底病杂志

Objective:To observe the gene mutations and clinical phenotypes in patients with Usher syndrome type 2 (USH2) and retinitis pigmentosa (RP).Methods:From August 2018 to January 2019, 4 patients and 11 normal family members from 3 families of USH2 and RP who visited Henan Eye Hospital were enrolled in the study. Detailed medical history was obtained and visual acuity, fundus color photography, OCT, visual field, full field ERG examination were performed. Among the three families, pedigree 1 was diagnosed with USH2, pedigree 2 and pedigree 3 were diagnosed with RP. The peripheral venous blood of patients and their family members were collected, and the whole genomic DNA was extracted. Targeted capture next generation sequencing analysis was performed on these members, and Sanger sequencing and family cosegregation were verified.Results:In the family F1, the proband had symptoms of RP and sensorineural deafness. Sequencing revealed two heterozygous frameshift variants: c.13877-13880 del AGAC (p. Q4626P) in exon 64 and c.798 del T (p. F266L) in exon 5 of USH2A. Both patients of family 2 and 3 showed RP signs without deafness. Two heterozygous variants c.15178T> C (p. S5060 P) in exon 70 and c.6986C> A (p. P2329H) in exon 37, and a pathogenic heterozygous variant c.5836C> T (p. R1946X) in exon 29 of USH2A were identified in family F2. A heterozygous missense variant c.14951C> T (p. P4984L) in exon 68 and a variant c.11156G> A (p. R3719H) in exon 57 of USH2A were found in family F3. The results of conservation analysis showed that the corresponding amino acid sites of USH2A p.Q4626P, p.F266L, p.S5060P, p.P2329H and p.P4984L were highly conserved in many species. Among these 7 pathogenic variants detected, M1-M4 and M6 were novel.Conclusions:Mutation USH2A gene are the main cause of USH2 and non-syndromic RP. Different variants affect protein translation and synthesis, consequently causing different clinical phenotypes.

Optimization of Flash Extraction Technology for Guizhi Fuling Capsule by Box-Behnken Design-response Surface Method / 中国药房

OBJECTIVE:To optimize the flash extraction technology for Guizhi fuling capsule. METHODS:Using blade speed,liquid-solid ratio,extraction time as investigation factors,using the comprehensive evaluation values consisting of transfer rates of gallic acid,paeoniflorin,benzoic acid,cinnamic acid,benzoyl paeoniflorin,amygdalin as investigation index,single fac-tor test was combined with Box-Behnken design-response surface method to optimize the flash extraction technology for Guizhi ful-ing capsule. Verification test was conducted and compared with conventional decoction extraction method(extracting twice,totally 240 min). RESULTS:The optimal flash extraction technology for Guizhi fuling capsule was using water as extraction solvent with blade speed of 5600 r/min and liquid-solid ratio of 10.5:1,extracting for 60 s. The average comprehensive evaluation value of veri-fication test was 85.40%(n=3),with relative error of 0.15% to the predicted value(85.55%),higher than decoction extraction method(72.65%). CONCLUSIONS:Optimized flash extraction technology for Guizhi fuling capsule is efficient and rapid.

Curcumin protects against the intestinal ischemia-reperfusion injury: involvement of the tight junction protein ZO-1 and TNF-alpha related mechanism

Present study aimed to investigate the eff ect of curcumin-pretreatment on intestinal I/R injury and on intestinal mucosa barrier. Thirty Wistar rats were randomly divided into: sham, I/R, and curcumin groups (n=10). Animals in curcumin group were pretreated with curcumin by gastric gavage (200 mg/kg) for 2 days before I/R. Small intestine tissues were prepared for Haematoxylin & Eosin (H&E) staining. Serum diamine oxidase (DAO) and tumor necrosis factor (TNF)-alpha levels were measured. Expression of intestinal TNF-alpha and tight junction protein (ZO-1) proteins was detected by Western blot and/or immunohistochemistry. Serum DAO level and serum and intestinal TNF-alpha leves were signifi cantly increased after I/R, and the values were markedly reduced by curcumin pretreatment although still higher than that of sham group (p<0.05 or p<0.001). H&E staining showed the significant injury to intestinal mucosa following I/R, and curcumin pretreatment signifi cantly improved the histological structure of intestinal mucosa. I/R insult also induced significantly down-regulated expression of ZO-1, and the eff ect was dramatically attenuated by curcumin-pretreatment. Curcumin may protect the intestine from I/R injury through restoration of the epithelial structure, promotion of the recovery of intestinal permeability, as well as enhancement of ZO-1 protein expression, and this eff ect may be partly attributed to the TNF-alpha related pathway.