Targeted Gene Sanger Sequencing Should Remain the First-Tier Genetic Test for Children Suspected to Have the Five Common X-Linked Inborn Errors of Immunity.

To address inborn errors of immunity (IEI) which were underdiagnosed in resource-limited regions, our centre developed and offered free genetic testing for the most common IEI by Sanger sequencing (SS) since 2001. With the establishment of The Asian Primary Immunodeficiency (APID) Network in 2009, the awareness and definitive diagnosis of IEI were further improved with collaboration among centres caring for IEI patients from East and Southeast Asia. We also started to use whole exome sequencing (WES) for undiagnosed cases and further extended our collaboration with centres from South Asia and Africa. With the increased use of Next Generation Sequencing (NGS), we have shifted our diagnostic practice from SS to WES. However, SS was still one of the key diagnostic tools for IEI for the past two decades. Our centre has performed 2,024 IEI SS genetic tests, with in-house protocol designed specifically for 84 genes, in 1,376 patients with 744 identified to have disease-causing mutations (54.1%). The high diagnostic rate after just one round of targeted gene SS for each of the 5 common IEI (X-linked agammaglobulinemia (XLA) 77.4%, Wiskott-Aldrich syndrome (WAS) 69.2%, X-linked chronic granulomatous disease (XCGD) 59.5%, X-linked severe combined immunodeficiency (XSCID) 51.1%, and X-linked hyper-IgM syndrome (HIGM1) 58.1%) demonstrated targeted gene SS should remain the first-tier genetic test for the 5 common X-linked IEI.

Sonographic Findings in Superficial Angiomyxoma of the Vulva in a Perimenopausal Female.

Superficial angiomyxoma (SAM) is an extremely rare soft tissue tumor. It is especially rare in the vulva, with only a few such cases reported in the medical literature. Here, we report a case of SAM of the vulva that was initially suspected to be a Bartholin gland cyst. The patient underwent local excision of the vulvar cyst under lumbar anesthesia. Clinical manifestations and B-scan ultrasonographic features are similar between SAM and cysts. Echoes in the mass are uneven and exhibit low echoes and punctate hyperechoic floating. Thus, increasing sonographers' awareness of the high-frequency ultrasonography findings associated with this rare tumor could broaden their knowledge base.

The epidemiology of paediatric Mycoplasma pneumoniae pneumonia in North China: 2006 to 2016.

Paediatric Mycoplasma pneumoniae pneumonia (MPP) is a major cause of community-acquired pneumonia in China. Data on epidemiology of paediatric MPP from China are little known. This study retrospectively collected data from June 2006 to June 2016 in Beijing Children's Hospital, Capital Medical University of North China and aims to explore the epidemiological features of paediatric MPP and severe MPP (SMPP) in North China during the past 10 years. A total of 27 498 paediatric patients with pneumonia were enrolled. Among them, 37.5% of paediatric patients had MPP. In this area, an epidemic took place every 2-3 years at the peak, and the positive rate of MPP increased during these peak years over time. The peak age of MPP was between the ages of 6 and 10 years, accounting for 75.2%, significantly more compared with other age groups (χ2 = 1384.1, P < 0.0001). The epidemics peaked in September, October and November (χ2 = 904.9, P < 0.0001). Additionally, 13.0% of MPP paediatric patients were SMPP, but over time, the rate of SMPP increased, reaching 42.6% in 2016. The mean age of paediatric patients with SMPP (6.7 ± 3.0 years old) was younger than that of patients with non-SMPP (7.4 ± 3.2 years old) (t = 3.60, P = 0.0001). The prevalence of MPP and SMPP is common in China, especially in children from 6 to 10 years old. Paediatric patients with SMPP tend to be younger than those with non-SMPP. MPP outbreaks occur every 2-3 years in North China. September, October and November are the peak months, unlike in South China. Understanding the epidemiological characteristics of paediatric MPP can contribute to timely treatment and diagnosis, and may improve the prognosis of children with SMPP.

A cascade oxidation/[4 + 1] annulation of sulfonium salts for synthesis of polyfunctional furans: DMSO as one carbon source.

A novel DMSO-involved cascade reaction of stabilized sulfonium salts has been established for direct construction of polyfunctional furans. This one-pot sequential reaction involving in situ generated α-methylene sulfonium salts was followed by [4 + 1] annulation with sulfur ylides. Notably, DMSO plays a very important role in this transformation, not only as a solvent but also as one carbon source.

Clinical and genetic characteristics of Chinese pediatric patients with chronic granulomatous disease.

BACKGROUND: Chronic granulomatous disease (CGD) is a rare disease in China, and very little large-scale studies have been conducted to date. We aimed to investigate the clinical and genetic features of CGD in Chinese pediatric patients. METHODS: Pediatric patients with CGD from Beijing Children's Hospital, Capital Medical University, China, were enrolled from January 2006 to December 2016. RESULTS: A total of 159 pediatric patients with CGD were enrolled. The median age of clinical onset was 1.4 months, and 73% (116/159) had clinical onset symptoms before the 1 year of age. The most common site of invasion was the lungs. The lymph nodes, liver, and skin were more frequently invaded in X-linked (XL) CGD patients than in autosomal recessive (AR) CGD patients (P < 0.05). Approximately 64% (92/144) of the pediatric patients suffered from abnormal response to BCG vaccination. The most frequent pathogens were Aspergillus and Mycobacterium tuberculosis. Gene analysis indicated that 132 cases (89%, 132/147) harbored CYBB pathogenic variants, 7 (5%, 7/147) carried CYBA pathogenic variants, 4 (3%, 4/147) had NCF1 pathogenic variants, and 4 (3%, 4/147) had NCF2 pathogenic variants. The overall mortality rate in this study was 43%, particularly the patients were males, with CYBB mutant and did not receive HSCT treatment. CONCLUSIONS: Chronic granulomatous disease is a rare disease affecting Chinese children; however, it is often diagnosed at a later age, and thus, the mortality rate is relatively high. The prevalence and the severity of disease in XL-CGD are higher than AR-CGD.

TFA-Mediated DMSO-Participant Sequential Oxidation/1,3-Dipolar Cycloaddition Cascade of Pyridinium Ylides for the Assembly of Indolizines.

A trifluoroacetic acid (TFA)-mediated cascade oxidation/1,3-dipolar cycloaddition reaction of stabilized pyridinium salts with dimethyl sulfoxide (DMSO) has been developed in the presence of K2S2O8 and trimethylethylenediamine (TMEDA). In this transition-metal-free reaction, DMSO acts as a one-carbon source, thus providing a convenient method for the efficient and direct synthesis of various indolizine derivatives.

Sequential σ-Bond Insertion/Benzannulation Involving Arynes: Selective Synthesis of Polysubstituted Naphthalenes.

An interesting σ-bond insertion/benzannulation reaction for the synthesis of polysubstituted naphthalene derivatives has been developed from readily accessible ketones, arynes, and alkynoates. This practical and transition-metal-free method provides a novel route to diverse naphthalenes through a substrate-controlled rearrangement reaction with the cleavage of C-C bonds.

Family History of Early Infant Death Correlates with Earlier Age at Diagnosis But Not Shorter Time to Diagnosis for Severe Combined Immunodeficiency.

BACKGROUND: Severe combined immunodeficiency (SCID) is fatal unless treated with hematopoietic stem cell transplant. Delay in diagnosis is common without newborn screening. Family history of infant death due to infection or known SCID (FH) has been associated with earlier diagnosis. OBJECTIVE: The aim of this study was to identify the clinical features that affect age at diagnosis (AD) and time to the diagnosis of SCID. METHODS: From 2005 to 2016, 147 SCID patients were referred to the Asian Primary Immunodeficiency Network. Patients with genetic diagnosis, age at presentation (AP), and AD were selected for study. RESULTS: A total of 88 different SCID gene mutations were identified in 94 patients, including 49 IL2RG mutations, 12 RAG1 mutations, 8 RAG2 mutations, 7 JAK3 mutations, 4 DCLRE1C mutations, 4 IL7R mutations, 2 RFXANK mutations, and 2 ADA mutations. A total of 29 mutations were previously unreported. Eighty-three of the 94 patients fulfilled the selection criteria. Their median AD was 4 months, and the time to diagnosis was 2 months. The commonest SCID was X-linked (n = 57). A total of 29 patients had a positive FH. Candidiasis (n = 27) and bacillus Calmette-Guérin (BCG) vaccine infection (n = 19) were the commonest infections. The median age for candidiasis and BCG infection documented were 3 months and 4 months, respectively. The median absolute lymphocyte count (ALC) was 1.05 × 109/L with over 88% patients below 3 × 109/L. Positive FH was associated with earlier AP by 1 month (p = 0.002) and diagnosis by 2 months (p = 0.008), but not shorter time to diagnosis (p = 0.494). Candidiasis was associated with later AD by 2 months (p = 0.008) and longer time to diagnosis by 0.55 months (p = 0.003). BCG infections were not associated with age or time to diagnosis. CONCLUSION: FH was useful to aid earlier diagnosis but was overlooked by clinicians and not by parents. Similarly, typical clinical features of SCID were not recognized by clinicians to shorten the time to diagnosis. We suggest that lymphocyte subset should be performed for any infant with one or more of the following four clinical features: FH, candidiasis, BCG infections, and ALC below 3 × 109/L.

Matrine reduces proliferation of human lung cancer cells by inducing apoptosis and changing miRNA expression profiles.

Matrine, a main active component extracted from dry roots of Sophora flavecens , has been reported to exert antitumor effects on A549 human non-small lung cancer cells, but its mechanisms of action remain unclear. To determine effects of matrine on proliferation of A549 cells and assess possible mechanisms, MTT assays were employed to detect cytotoxicity, along with o flow cytometric analysis of DNA content of nuclei of cells following staining with propidium iodide to analyze cell cycle distribution. Western blotting was performed to determined expression levels of Bax, Bcl-2, VEGF and HDAC1, while a microarray was used to assessed changes of miRNA profiles. In the MTT assay, matrine suppressed growth of human lung cancer cell A549 in a dose- and time- dependent manner at doses of 0.25-2.5 mg/ml for 24h, 48h or 72h. Matrine induced cell cycle arrest in G0/G1 phase and decreased the G2/M phase, while down-regulating the expression of Bcl2 protein, leading to a reduction in the Bcl-2/Bax ratio. In addition, matrine down regulated the expression level of VEGF and HDAC1 of A549 cells. Microarray analysis demonstrated that matrine altered the expression level of miRNAs compared with untreated control A549 cells. In conclusion, matrine could inhibit proliferation of A549 cells, providing useful information for understanding anticancer mechanisms.

[Clinical manifestations and gene analysis of 2 Chinese children with cystic fibrosis].

OBJECTIVE: Cystic fibrosis (CF) is rare in Chinese mainland. We present two cases of CF patients diagnosed by gene analysis. Their clinical manifestations and genetic mutation features are analyzed in this article. It will be of special interest to pediatricians in recognition of CF. METHOD: The clinical material of two CF patients who were diagnosed by gene analysis was retrospectively analyzed. RESULT: The first patient is a 13-year-old girl. She had a complaint of recurrent fever and cough for 6 months, expectoration for 2 months and hemoptysis for 20 days. After 3 months of her birth, she was operated on for bullae of lung. She was susceptible to upper respiratory tract infection. There was no family history of recurrent wheeze and other special diseases. Aspergillus fumigatus specific IgE was at grade 3 and aspergillus fumigatus IgG was high. Pseudomonas aeruginosa was positive in sputum culture. Sweat testing was performed and Na+ was higher. Pulmonary CT indicated bronchiectasis. Nasal sinus CT showed optical density of soft tissue within maxillary sinus and chronic bilateral sinusitis. The electron microscopy of cilia suggested immobile cilia syndrome. A heterozygotic mutation (263T > G, 2909G > A) was found after CFTR genetic mutation analysis. Both her parents were carriers. She was treated with inhalation of nebulized hypertonic saline and postural drainage for a long time. And she got better during a follow up period of 1 year. The second patient was a 10-year-old girl who complained of recurrent expectoration for 3 years and shortness of breath for half a year. She had a history of sinusitis and steatorrhea. The family history was normal. Both the lipase and insulin level in blood serum was lower.Pseudomonas aerugino and Aspergillus fumigatus were both positive in sputum culture. Aspergillus fumigatus IgE was normal. Pulmonary CT indicated bronchiolitis and bronchiectasis. Nasal sinus CT showed bilateral maxillary sinusitis. CFTR genetic mutation analysis revealed a homozygous mutation (3196C > T). Her parents and relatives did not participate in this study. Unfortunately, this child died of respiratory failure 3 months after discharge. CONCLUSION: CFTR gene mutation was a main cause of CF. Common symptoms are those of bronchiectasis, pancreatitis and sinusitis. The two Chinese patients were diagnosed by gene analysis. One had a heterozygous mutation (263T > G, 2909G > A) and the other had a homozygous mutation (3196C > T), not ΔF508 which is common in western countries.

[Clinical features and molecular analysis of 2 Chinese children with autosomal recessive chronic granulomatous disease caused by CYBA mutations].

OBJECTIVE: To summarize clinical and molecular features of two children with autosomal recessive chronic granulomatous disease caused by CYBA mutations. METHOD: The clinical records and CYBA mutations were reviewed for analysis of infections and inflammatory complications. RESULT: The first case was a girl diagnosed with "liver and spleen abscess" in our hospital when she was 2.9 years old, with past history of neonatal impetigo and recurrent purulent lymphadenitis and positive family history. The results of DHR123 flow-cytometry showed that positive phagocytes after phorbol ester (PMA) stimulation was 84.63%. CYBA mutation analysis showed that she had heterozygous 35C > T, Q3X and IVS-2A > G. The second case was a boy diagnosed with "sepsis (salmonella D)" when he was 4 years old with a past history of impetigo, sepsis, perianal abscess, skin infection and positive family history. The results of flow cytometry showed that positive phagocytes after PMA stimulation was 96.13%. CYBA mutation analysis showed that he had homozygous 35C > T, Q3X and his parents were all carriers. All of them had BCG related axillary lymphnode calcification. CONCLUSION: A22CGD cases had recurrent purulent infections (skin, lymphnode, liver and spleen, lung, blood), DHR123 flow cytometric analysis helped the diagnosis of CGD, CYBA mutation analysis ascertained the diagnosis of A22CGD.

[Screening and application of prokaryotic enhancer-like sequence 3A].

OBJECTIVE: To screen enhancer-like sequences from Escherichia coli strain C600 genome, to construct an expression vector harboring prokaryotic enhancer-like sequence and study the effect of interferon gene expression. METHODS: Enhancer-like element from Escherichia coli strain C600 genome was obtained by using the chloramphenicol acetyl-transferase (CAT) gene as reporter gene. An expression vector harboring prokaryotic enhancer-like sequence from Escherichia coli strain C600 was constructed. Interferon was expressed and assayed. RESULTS: An enhancer-like sequences with distance and orientation independence property were screened and named 3A. Quantification test showed that the direct and reverse orientation of 3A could increase the activity of beta-galactosidase with 7.11 and 2.93 times. The enhancing activity of the element was on transcription level. An expression vector harboring the prokaryotic enhancer-like sequence 3P3 which was enhancing function region of sequence 3A was constructed. Using this vector the antiviral activity of interferon alpha-2b was increased by 3.7 times in comparison with the original expression plasmid. CONCLUSION: 3A enhancer-like sequence was screened from Escherichia coli strain C600 genome. Interferon gene was highly expressed by using an expression vector harboring enhancer-like sequences.

[Clinical analysis of 18 children with disseminated Bacille Calmette-Guérin infection].

OBJECTIVE: To explore the clinical manifestation, immune abnormality and outcome of disseminated Bacille Calmette-Guérin (BCG) infection in children. METHOD: The clinical data of 18 children with disseminated BCG infection seen from January 2000 to December 2007 were analyzed retrospectively. RESULT: Thirteen of the children were male among 18 patients. Disseminated infection first appeared in armpit lymph nodes ipsilateral to the vaccination site, then spread to lungs in 15, lymphnodes of mediastinum or abdominal cavity in 18, skin and soft tissues in 5, skeletons in 4, liver in 4, spleen in 8, kidney, adrenal gland or meninges in 3. Twelve children were diagnosed to have primary immunodeficiency; 3 had severe combined immunodeficiency (SCID); 7 had chronic granulomatous disease (CGD), 2 had IL-12/IFN-gamma passageway deficiency. Eleven of the 18 patients died, and the remaining 7 patients were followed up from 1 to 9 years and are alive at present, but presented recurrent skin and bone tuberculosis in 4 and recurrent other infection in 3. CONCLUSION: Most Children with disseminated BCG infection had primary immunodeficiency. CGD and IL-12/IFN-gamma passageway deficiency accounted for considerable proportion, so special immune function should be detected in these patients. The prognosis was poor. The type of the immunodeficiency diseases should be identified in early stage and the specific immune treatment should be given to the patients.

[The clinical application of the emergency remedial transthoracic cardiac pacing used by a steel wire loop electrode].

OBJECTIVE: To explore an effective method of emergency remedial cardiac pacing and evaluate its clinical application. METHODS: The transthoracic cardiac pacing was used by a steel wire loop electrode to 18 patients with cardiac arrest or serious bradycardia, after a routine cardiac pacing was failed by way of jugular or subclavian vein. RESULTS: It is a simple procedure used by steel wire loop. All the pacing had been successfully captured the hearts except 5 cases without capturing. There were 3 cases of cardiac arrest, 2 cases of cardiac arrest after electric defibrillation, 1 case of cardiac arrest after electric shock because of ventricular running and all the 5 cases of severe cardiac bradycardia due to serious atrioventricular block caused by high blood potassium were survived. Eight cases of cardiac arrest were successful capture, that was due to the short period of time from cardiac arrest to ventricle puncture than that of 5 cases of ventricular pacing which was not successful [(17.6+/-4.6) minutes vs. (26.4+/-5.4) minutes, P<0.05]. CONCLUSION: The steel wire loop electrode is a safe and reliable emergency cardiac pacing method, which contribute to the rapid establishment of an effective emergency cardiac pacing. When the conventional intravenous cardiac pacing was not successful, the method of emergency transthoracic cardiac pacing used by steel wire loop electrode should be bold and use to improve the successful rate of resuscitation.

[Expression of B7-related protein-1 on human coronary artery endothelial cells and its clinical significance].

OBJECTIVE: To study the effect of oxidized low density lipoprotein (ox-LDL) on the expression of B7-related protein-1 (B7RP-1) on human coronary artery endothelial cells (HCAECs). METHODS: HCAECs were incubated in the presence of 100 mg/L ox-LDL for 24 h, and B7RP-1 expression levels were determined using fluorescence reverse transcription PCR (RT-PCR) and Western blotting. RESULTS: B7RP-1 expression was detected HCAECs, with spots of fluorescence signals distributing on the cell membrane as observed under fluorescence microscope. RT-PCR with B7RP-1 specific primers yielded products of expected size (496 bp). Western blotting identified B7RP-1 expression in the HCAECs as a cell-associated protein with an apparent molecular mass of 70,000. Treatment of the cells with ox-LDL significantly increased B7RP-1 expression at both the mRNA and protein levels (P<0.05). CONCLUSION: B7RP-1 is expressed on the membrane of HCAECs. ox-LDL can promote up-regulate the expression of B7RP-1, which might be one of the immunopathogenesis of atherosclerosis.