Ivy Hau-Yee Chan

Middle East Respiratory Syndrome Coronavirus Efficiently Infects Human Primary T Lymphocytes and Activates the Extrinsic and Intrinsic Apoptosis Pathways.

Abstract Middle East respiratory syndrome (MERS) is associated with a mortality rate of >35%. We previously showed that MERS coronavirus (MERS-CoV) could infect human macrophages and dendritic cells and induce cytokine dysregulation. Here, we further investigated the interplay between human primary T cells and MERS-CoV in disease pathogenesis. Importantly, our results suggested that MERS-CoV efficiently infected T cells from the peripheral blood and from human lymphoid organs, including the spleen and the tonsil. We further demonstrated that MERS-CoV infection induced apoptosis in T cells, which involved the activation of both the extrinsic and intrinsic apoptosis pathways. Remarkably, immunostaining of spleen sections from MERS-CoV–infected common marmosets demonstrated the presence of viral nucleoprotein in their CD3+ T cells. Overall, our results suggested that the unusual capacity of MERS-CoV to infect T cells and induce apoptosis might partly contribute to the high pathogenicity of the virus.

Fine Mapping of the NRG1 Hirschsprung's Disease Locus

The primary pathology of Hirschsprungs disease (HSCR, colon aganglionosis) is the absence of ganglia in variable lengths of the hindgut, resulting in functional obstruction. HSCR is attributed to a failure of migration of the enteric ganglion precursors along the developing gut. RET is a key regulator of the development of the enteric nervous system (ENS) and the major HSCR-causing gene. Yet the reduced penetrance of RET DNA HSCR-associated variants together with the phenotypic variability suggest the involvement of additional genes in the disease. Through a genome-wide association study, we uncovered a ∼350 kb HSCR-associated region encompassing part of the neuregulin-1 gene (NRG1). To identify the causal NRG1 variants contributing to HSCR, we genotyped 243 SNPs variants on 343 ethnic Chinese HSCR patients and 359 controls. Genotype analysis coupled with imputation narrowed down the HSCR-associated region to 21 kb, with four of the most associated SNPs (rs10088313, rs10094655, rs4624987, and rs3884552) mapping to the NRG1 promoter. We investigated whether there was correlation between the genotype at the rs10088313 locus and the amount of NRG1 expressed in human gut tissues (40 patients and 21 controls) and found differences in expression as a function of genotype. We also found significant differences in NRG1 expression levels between diseased and control individuals bearing the same rs10088313 risk genotype. This indicates that the effects of NRG1 common variants are likely to depend on other alleles or epigenetic factors present in the patients and would account for the variability in the genetic predisposition to HSCR.

Evaluation of defecative function 5 years or longer after laparoscopic-assisted pull-through for imperforate anus

BACKGROUND Laparoscopic anorectoplasty was introduced in 2000, but the procedure has not yet gained universal acceptance. Previous studies, including ours, reported satisfactory early postoperative outcome as compared with posterior sagittal anorectoplasty (PSARP), but mid- to long-term results are not available. Here, we aim to evaluate the mid- to long-term defecative function in these patients. MATERIALS AND METHODS A retrospective study was carried out and included all patients who underwent laparoscopic-assisted anorectoplasty for high-/intermediate-type imperforate anus between 2001 and 2005. Their degree of continence was graded according to the Krickenbeck classification and compared with historical controls treated with PSARP. The results were compared using χ(2) test, and P < .05 was considered statistically significant. RESULTS There were 18 patients who underwent laparoscopic anorectoplasty (LAR) in the study period. They were compared with 20 historical PSARP patients. For defecation sensation, 16 of the 18 LAR patients were positive, whereas there were 16 of 20 PSARP patients. Eight LAR patients were clean without any fecal soiling or incontinence (11/20 PSARP). Only 3 of 18 LAR had constipation as compared with 7 of 20 PSARP. The need for rectal enema for evacuation was seen in 1 of 20 LAR patients and 2 of 20 PSARP patients (for all categories, P > .05). CONCLUSIONS Mid- to long-term follow-up study revealed satisfactory defecative function for patients with high-/intermediate-type imperforate anus after LAR. The outcome is at least as good as PSARP. Longer term follow-up with larger sample size is necessary to demonstrate the benefits of LAR over PSARP.

Genome-wide copy number variation study in anorectal malformations

Anorectal malformations (ARMs, congenital obstruction of the anal opening) are among the most common birth defects requiring surgical treatment (2-5/10 000 live-births) and carry significant chronic morbidity. ARMs present either as isolated or as part of the phenotypic spectrum of some chromosomal abnormalities or monogenic syndromes. The etiology is unknown. To assess the genetic contribution to ARMs, we investigated single-nucleotide polymorphisms and copy number variations (CNVs) at genome-wide scale. A total of 363 Han Chinese sporadic ARM patients and 4006 Han Chinese controls were included. Overall, we detected a 1.3-fold significant excess of rare CNVs in patients. Stratification of patients by presence/absence of other congenital anomalies showed that while syndromic ARM patients carried significantly longer rare duplications than controls (P = 0.049), non-syndromic patients were enriched with both rare deletions and duplications when compared with controls (P = 0.00031). Twelve chromosomal aberrations and 114 rare CNVs were observed in patients but not in 868 controls nor 11 943 healthy individuals from the Database of Genomic Variants. Importantly, these aberrations were observed in isolated ARM patients. Gene-based analysis revealed 79 genes interfered by CNVs in patients only. In particular, we identified a de novo DKK4 duplication. DKK4 is a member of the WNT signaling pathway which is involved in the development of the anorectal region. In mice, Wnt disruption results in ARMs. Our data suggest a role for rare CNVs not only in syndromic but also in isolated ARM patients and provide a list of plausible candidate genes for the disorder.

RET Mutational Spectrum in Hirschsprung Disease: Evaluation of 601 Chinese Patients

Rare (RVs) and common variants of the RET gene contribute to Hirschsprung disease (HSCR; congenital aganglionosis). While RET common variants are strongly associated with the commonest manifestation of the disease (males; short-segment aganglionosis; sporadic), rare coding sequence (CDS) variants are more frequently found in the lesser common and more severe forms of the disease (females; long/total colonic aganglionosis; familial). Here we present the screening for RVs in the RET CDS and intron/exon boundaries of 601 Chinese HSCR patients, the largest number of patients ever reported. We identified 61 different heterozygous RVs (50 novel) distributed among 100 patients (16.64%). Those include 14 silent, 29 missense, 5 nonsense, 4 frame-shifts, and one in-frame amino-acid deletion in the CDS, two splice-site deletions, 4 nucleotide substitutions and a 22-bp deletion in the intron/exon boundaries and 1 single-nucleotide substitution in the 5′ untranslated region. Exonic variants were mainly clustered in RET the extracellular domain. RET RVs were more frequent among patients with the most severe phenotype (24% vs. 15% in short-HSCR). Phasing RVs with the RET HSCR-associated haplotype suggests that RVs do not underlie the undisputable association of RET common variants with HSCR. None of the variants were found in 250 Chinese controls.

Thoracoscopic repair of oesophageal atresia: experience of 33 patients from two tertiary referral centres.

BACKGROUND With advances in minimally invasive surgery, thoracoscopic repair of oesophageal atresia has become popular in many centres worldwide and indeed has been described as the pinnacle of neonatal surgery. Here, we report our experience in two tertiary referral centres. METHODS Thoracoscopic technique was introduced in 2007. Thus, a retrospective review of all patients diagnosed with oesophageal atresia was carried out. Patients who had thoracoscopic repair were included, and those who had open repair due to co-morbidities were excluded. Patient demographics, operative data, complications, and associated anomalies were noted. RESULTS A total of thirty-three patients underwent thoracoscopic repair during the time period. Thirty-one were successfully repaired thoracoscopically. Two patients had conversions due to intra-operative instability. The mean body weight of the neonates was 2.58 kg. The mean operative time was 146 min. Three patients suffered from minor anastomotic leaks, which healed on conservative management. Seven patients had anastomotic strictures, which responded successfully to endoscopic dilatation. Two patients died in the post-operative period due to pneumonia. One patient had a recurrent fistula 3 months after the primary repair, and he subsequently underwent a successful second repair. CONCLUSIONS In experienced hands, thoracoscopic repair of oesophageal atresia is at least as good as open surgery but with less surgical trauma. Standard of post-operative care contributes significantly to post-operative outcome. Thoracoscopic technique is now our preferred approach.

Advances in paediatric urology

Paediatric urological surgery is often required for managing congenital and acquired disorders of the genitourinary system. In this Series paper, we highlight advances in the surgical management of six paediatric urological disorders. The management of vesicoureteral reflux is evolving, with advocacy ranging from a less interventional assessment and antimicrobial prophylaxis to surgery including endoscopic injection of a bulking agent and minimally invasive ureteric reimplantation. Evidence supports early orchidopexy to improve fertility and reduce malignancy in boys with undescended testes. A variety of surgical techniques have been developed for hypospadias, with excellent outcomes for distal but not proximal hypospadias. Pelvi-ureteric junction obstruction is mostly detected prenatally; indications for surgery have been refined with evidence, and minimally invasive pyeloplasty is now standard. The outlook for patients with neurogenic bladder has been transformed by a combination of clean intermittent catheterisation, algorithms of diagnostic investigations, and innovative medical and surgical therapies. Posterior urethral valves are associated with considerable mortality; fetal diagnosis allows stratification of candidates for intervention, but ongoing bladder dysfunction in patients after valve ablation remains a cause of long-term morbidity.

Human intestinal tract serves as an alternative infection route for Middle East respiratory syndrome coronavirus

Human intestinal tract as an alternative route to acquire MERS-CoV infection. Middle East respiratory syndrome coronavirus (MERS-CoV) has caused human respiratory infections with a high case fatality rate since 2012. However, the mode of virus transmission is not well understood. The findings of epidemiological and virological studies prompted us to hypothesize that the human gastrointestinal tract could serve as an alternative route to acquire MERS-CoV infection. We demonstrated that human primary intestinal epithelial cells, small intestine explants, and intestinal organoids were highly susceptible to MERS-CoV and can sustain robust viral replication. We also identified the evidence of enteric MERS-CoV infection in the stool specimen of a clinical patient. MERS-CoV was considerably resistant to fed-state gastrointestinal fluids but less tolerant to highly acidic fasted-state gastric fluid. In polarized Caco-2 cells cultured in Transwell inserts, apical MERS-CoV inoculation was more effective in establishing infection than basolateral inoculation. Notably, direct intragastric inoculation of MERS-CoV caused a lethal infection in human DPP4 transgenic mice. Histological examination revealed MERS-CoV enteric infection in all inoculated mice, as shown by the presence of virus-positive cells, progressive inflammation, and epithelial degeneration in small intestines, which were exaggerated in the mice pretreated with the proton pump inhibitor pantoprazole. With the progression of the enteric infection, inflammation, virus-positive cells, and live viruses emerged in the lung tissues, indicating the development of sequential respiratory infection. Taken together, these data suggest that the human intestinal tract may serve as an alternative infection route for MERS-CoV.

Endoscopic injection for primary vesicoureteric reflux: Predictors of resolution and long term efficacy

AIM We investigated the efficacy of endoscopic-Deflux-injection in treating primary-vesicoureteric-reflux (VUR) and identified factors to predict resolution. MATERIALS AND METHODS Records of children treated with Deflux for primary-VUR from 1995 to 2016 were reviewed, and outcomes were investigated. RESULTS Eighty-eight ureters (35 bilateral, 18 unilateral) in 53 children underwent 124 injections. Thirty-five (66%) patients had single injection (13 unilateral, 22 bilateral). Fifteen (28%), two (37%), and one (2%) patients had two, three, and four injections, respectively. Overall success rate by ureters was 57% after single injection. Complete resolution occurred in 65% of ureters with VUR below grade III, 63% of grade III, 40% of grade IV, and 70% of grade V VUR. Four patients had reimplantation. The median follow up duration was 60months (range 20-216months). Univariate analysis showed that lower VUR grade (p=0.03) and absent renal scars (p=0.04) were statistically significant predictors of resolution. In multivariate analysis, absent renal scars were statistically significant (p=0.01). CONCLUSION We demonstrated efficacy of endoscopic-Deflux-injection as the first line treatment for primary-VUR. Absent renal scar and lower VUR grade were statistically significant predictors of resolution after single injection. TYPE OF STUDY Case-Control / Retrospective Comparative Study. LEVEL OF EVIDENCE Level III.

Long-term implications and global impact of paediatric surgery

1012 www.thelancet.com Vol 390 September 9, 2017 Irrespective of variations across geography, culture, and socioeconomic status, paediatric surgery differs from other surgical subspecialties. Children are not small adults. Surgery for infants and children is typically undertaken for congenital, rare, and complex conditions and the consequences of both the condition and its treatment can affect that individual for life. Above all, the surgical outcome needs to stand the test of time. Our Lancet Series papers in this issue on advances in paediatric urology and gastroenterology highlight the transition of paediatric surgery from a focus on shortterm patient survival to evidence and innovations for long-term quality of life globally. We believe the lessons and challenges have wide implications and offer insights into how knowledge gaps can be filled and new treatments designed. Most paediatric surgery requires general anaesthesia and concern about anaesthetic agents harming the developing brain has been raised by some experimental animal studies. The 2016 US Food and Drug Administration’s warning about possible neuro developmental effects of anaesthetic and sedative drugs in infants younger than 3 years and in the fetus during the third trimester should, however, be interpreted with caution. Evidence of harm in human beings is inconclusive and, reassuringly, a randomised controlled trial has shown no mediumterm adverse neurodevelopmental effects after a single general anaesthetic during infancy. Nevertheless, the long-term outcome of repeated or lengthy anaesthetic exposure, especially in more vulnerable subgroups such as premature infants, remains unknown. This potential risk of anaesthesia in infancy has created a dilemma about whether to delay the timing of surgery until after the age 3 years for elective conditions such as hypospadias and craniofacial anomalies in which early correction can confer functional and psychocosmetic benefits. Surgical decisions made in infancy and childhood have even more profound effects on adult life in the management of some disorders of sexual development (DSD). In the past, decisions about genital surgery and assignation of gender were made during infancy and remained a compact between the surgical team and the parents and did not account for underlying ethical implications related to the exclusion of patient’s choice. Surgeons are now more aware of the problems arising from surgery for DSD in infancy. For example, early reduction clitoroplasty for enlarged clitoris in female DSD can compromise later sexual function, and often leads to major concerns about gender identity, body habitus, and psychosexual functioning. Although the exact timing of reconstructive surgery remains controversial, common practice follows the Chicago DSD Consensus guidelines. Ethical considerations also affect surgical management for other conditions such as total intestinal aganglionosis in which surgery results in a major health burden of life-long total parenteral nutrition. When parental wishes and surgical opinions differ, for example in end-of-life decisions for multiple anomalies, the medical community has to be prepared for ethical, legal, and public debate. Quality of life in adults with DSD is also affected by hormonal replacement, issues with fertility, an increased risk of malignancy, and personal, family, and cultural perceptions of “normal” sex development. Many adults with complex anorectal malformations similarly experience sexual dysfunction and more than 50% of children with such congenital malformations need special education. In terms of increased risk of malignancy, there can be a predisposition to laterlife malignancy inherent in the condition (eg, DSD, undescended testes, or choledochal cysts), or the actual nature of the surgical reconstruction can predispose Long-term implications and global impact of paediatric surgery 6 NSABP B-39, RTOG 0413: a randomized phase III study of conventional whole breast irradiation versus partial breast irradiation for women with stage 0, I, or II breast cancer. Clin Adv Hematol Oncol 2006; 4: 719–21. 7 Livi L, Meattini I, Marrazzo L, et al. Accelerated partial breast irradiation using intensity-modulated radiotherapy versus whole breast irradiation: 5-year survival analysis of a phase 3 randomised controlled trial. Eur J Cancer 2015; 51: 451–63. 8 Hughes KS, Schnaper LA, Berry D, et al, for the Cancer and Leukemia Group B, Radiation Therapy Oncology Group, and Eastern Cooperative Oncology Group. Lumpectomy plus tamoxifen with or without irradiation in women 70 years of age or older with early breast cancer. N Engl J Med 2004; 351: 971–77. 9 Kunkler IH, Williams LJ, Jack WJ, Cameron DA, Dixon JM, on behalf of the PRIME II investigators. Breast-conserving surgery with or without irradiation in women aged 65 years or older with early breast cancer (PRIME II): a randomised controlled trial. Lancet Oncol 2015; 16: 266–273. 10 Gradishar WJ, Anderson BO, Balassanian R, et al. NCCN guidelines insights: breast cancer, version 1·2017. J Natl Compr Canc Netw 2017; 15: 433–51.