Improved diagnostic yield of peripheral pulmonary malignant lesions with emphysema using a combination of radial endobronchial ultrasonography and rapid on-site evaluation.

BACKGROUND: This is a retrospective cohort study from a single center of Chest Medical District of Nanjing Brain Hospital Affiliated to Nanjing Medical University, Jiangsu Province, China. It was aim to evaluate the diagnostic value of radial endobronchial ultrasound (R-EBUS) combination with rapid on-site evaluation (ROSE) guided transbronchial lung biopsy (TBLB) for peripheral pulmonary lesions in patients with emphysema. METHODS: All 170 patients who underwent PPLs with emphysema received an R-EBUS examination with or without the ROSE procedure, and the diagnostic yield, safety, and possible factors influencing diagnosis were analyzed between the two groups by the SPSS 25.0 software. RESULTS: The pooled and benign diagnostic yields were not different in the two groups (P = 0.224, 0.924), but the diagnostic yield of malignant PPLs was significantly higher in the group with ROSE than the group without ROSE (P = 0.042). The sensitivity of ROSE was 79.10%, the specificity, 91.67%, the positive predictive value, 98.15%, and the negative predictive value, 84.62%. The diagnostic accuracy, was 95.52%. In the group of R-EBUS + ROSE, the procedural time and the number of times of biopsy or brushing were both significantly reduced (all P<0.05). The incidence of pneumothorax (1.20%) and bleeding (10.84%) in the group of R-EBUS + ROSE were also less than those in the group of R-EBUS (P<0.05). The lesion's diameter ≥ 2 cm, the distance between the pleura and the lesion ≥ 2 cm, the positive air bronchograms sign, the location of the ultrasound probe within the lesion, and the even echo with clear margin feature of lesion ultrasonic image, these factors are possibly relevant to a higher diagnostic yield. The diagnostic yield of PPLs those were adjacent to emphysema were lower than those PPLs which were away from emphysema (P = 0.048) in the group without ROSE, however, in the group of R-EBUS + ROSE, there was no such difference whether the lesion is adjacent to emphysema or not (P = 0.236). CONCLUSION: Our study found that the combination of R-EBUS and ROSE during bronchoscopy procedure was a safe and effective modality to improve diagnostic yield of PPLs with emphysema, especially for malignant PPLs. The distance between the pleura and the lesion ≥ 2 cm, the positive air bronchograms sign, the location of the ultrasound probe within the lesion, and the even echo with clear margin feature of lesion ultrasonic image, these factors possibly indicated a higher diagnostic yield. Those lesions' position is adjacent to emphysema may reduce diagnostic yield but ROSE may make up for this deficiency.

Preoperative Platelet-Lymphocyte Ratio (PLR) as a prognostic inflammation biomarker in Asian HIV-infected patients with gastric cancer: a single-center study.

BACKGROUND: The serum systemic inflammation biomarkers have been established as predictors of prognosis in gastric cancer (GC) patients, but their prognostic value in human immunodeficiency virus (HIV)-infected patients with GC has not been well studied. This retrospective study aimed to evaluate the prognostic value of preoperative systemic inflammation biomarkers in Asian HIV-infected patients with GC. METHODS: We retrospectively analyzed 41 HIV-infected GC patients who underwent surgery between January 2015 and December 2021 at the Shanghai Public Health Clinical Center. Preoperative systemic inflammation biomarkers were measured and patients were divided into two groups based on the optimal cut-off value. Overall survival (OS) and progression-free survival (PFS) were measured using the Kaplan-Meier method and the log-rank test. Multivariate analysis of variables was performed using the Cox proportional regression model. As a comparison, 127 GC patients without HIV infection were also recruited. RESULTS: The median age of the 41 patients included in the study was 59 years, with 39 males and two females. The follow-up period for OS and PFS ranged from 3 to 94 months. The cumulative three-year OS rate was 46.0%, and the cumulative three-year PFS rate was 44%. HIV-infected GC patients had worse clinical outcomes compared to the normal GC population. The optimal cut-off value for preoperative platelet to lymphocyte ratio (PLR) was 199 in HIV-infected GC patients. Multivariate Cox regression analysis revealed that a low PLR was an independent predictor of better OS and PFS (OS: HR = 0.038, 95% CI: 0.006-0.258, P < 0.001; PFS: HR = 0.027, 95% CI: 0.004-0.201, P < 0.001). Furthermore, higher preoperative PLR in HIV-infected GC was significantly associated with lower BMI, hemoglobin, albumin, CD4 + T, CD8 + T, and CD3 + T cell counts. CONCLUSION: The preoperative PLR is an easily measurable immune biomarker that may provide useful prognostic information in HIV-infected GC patients. Our findings suggest that PLR could be a valuable clinical tool for guiding treatment decisions in this population.

Defining pathogenicity of NOTCH2 variants for diagnosis of Alagille syndrome type 2 using a large cohort of patients.

BACKGROUND AND AIMS: Alagille syndrome (ALGS) type 2 caused by mutations in NOTCH2 has genotypic and phenotypic heterogeneity. Diagnosis in some atypical patients with isolated hepatic presentation could be missed. METHODS: Using 2087 patients with paediatric liver manifestations, NOTCH2 allele frequencies, in-silico prediction, protein domains and clinical features were analysed to define the pathogenicity of NOTCH2 variants for diagnosis of ALGS type 2. RESULTS: Among 2087 patients with paediatric liver manifestations, significantly more NOTCH2 variants were absent in gnomAD in patients with elevated γ-glutamyltransferase (GGT) (p = .041). Significantly more NOTCH2 variants which were absent in gnomAD were located in protein functional domains (p = .038). When missense variants were absent in gnomAD and predicted to be pathogenic by at least three out of seven in-silico tools, they were found to be significantly associated with liver manifestations with elevated GGT (p = .003). Comparing this to patients with likely benign (LB) variants, the patients with likely-pathogenic (LP) variants have significantly more liver manifestations with elevated GGT (p = .0001). Significantly more patients with LP variants had extra-hepatic phenotypes of ALGS compared with those patients with LB variants (p = .0004). CONCLUSION: When NOTCH2 variants are absent in gnomAD, null variants and missense variants which were predicted to be pathogenic by at least three in-silico tools could be considered pathogenic in patients with high GGT chronic liver diseases.

Association of novel TMEM67 variants with mild phenotypes of high gamma-glutamyl transpeptidase cholestasis and congenital hepatic fibrosis.

TMEM67 (mecklin or MKS3) locates in the transition zone of cilia. Dysfunction of TMEM67 disrupts cilia-related signaling and leads to developmental defects of multiple organs in humans. Typical autosomal recessive TMEM67 defects cause partial overlapping phenotypes, including abnormalities in the brain, eyes, liver, kidneys, bones, and so forth. However, emerging reports of isolated nephronophthisis suggest the possibility of a broader phenotype spectrum. In this study, we analyzed the genetic data of cholestasis patients with no obvious extrahepatic involvement but with an unexplained high level of gamma-glutamyl transpeptidase (GGT). We identified five Han Chinese patients from three unrelated families with biallelic nonnull low-frequency TMEM67 variants. All variants were predicted pathogenic in silico, of which p. Arg820Ile and p. Leu144del were previously unreported. In vitro studies revealed that the protein levels of the TMEM67 variants were significantly decreased; however, their interaction with MKS1 remained unaffected. All the patients, aged 7-39 years old, had silently progressive cholestasis with elevated GGT but had normal bilirubin levels. Histological studies of liver biopsy of patients 1, 3, and 5 showed the presence of congenital hepatic fibrosis. We conclude that variants in TMEM67 are associated with a mild phenotype of unexplained, persistent, anicteric, and high GGT cholestasis without typical symptoms of TMEM67 defects; this possibility should be considered by physicians in gastroenterology and hepatology.

Whole-Genome Sequencing Reveals Large ATP8B1 Deletion/Duplications as Second Mutations Missed by Exome-Based Sequencing.

Progressive familial intrahepatic cholestasis type 1 (PFIC1) results from biallelic pathogenic variants in ATP8B1. This study sought second pathogenic variants in ATP8B1 by whole-genome sequencing (WGS) in four unrelated low γ-glutamyl transpeptidase cholestasis patients in whom clinical suspicion of PFIC1 was high and gene-panel or Sanger sequencing had identified only one pathogenic variant in ATP8B1. Sanger sequencing confirmed WGS findings and determined the origin of each variant. Novel nonrecurrent structural variants in three patients (patient 1 to patient 3) were identified in trans: g.55396652_55403080del (6427-bp deletion), g.55335906_55346620dup (10,715-bp duplication), and g.55362063_55364293dup (2231-bp duplication). One synonymous variant in patient 4 was recognized in trans (c.1029G>A, p. Thr343Thr) and demonstrated as deleterious. In conclusion, WGS improves genetic diagnostic yield in PFIC1. These findings expand the gene-variant spectrum associated with familiar intrahepatic cholestasis 1 (FIC1) disease and for the first time report tandem duplication in ATP8B1 associated with cholestasis.

Biallelic loss-of-function ZFYVE19 mutations are associated with congenital hepatic fibrosis, sclerosing cholangiopathy and high-GGT cholestasis.

BACKGROUND: For many children with intrahepatic cholestasis and high-serum gamma-glutamyl transferase (GGT) activity, a genetic aetiology of hepatobiliary disease remains undefined. We sought to identify novel genes mutated in children with idiopathic high-GGT intrahepatic cholestasis, with clinical, histopathological and functional correlations. METHODS: We assembled a cohort of 25 children with undiagnosed high-GGT cholestasis and without clinical features of biliary-tract infection or radiological features of choledochal malformation, sclerosing cholangitis or cholelithiasis. Mutations were identified through whole-exome sequencing and targeted Sanger sequencing. We reviewed histopathological findings and assessed phenotypical effects of ZFYVE19 deficiency in cultured cells by immunofluorescence microscopy. RESULTS: Nine Han Chinese children harboured biallelic, predictedly complete loss-of-function pathogenic mutations in ZFYVE19 (c.314C>G, p.S105X; c.379C>T, p.Q127X; c.514C>T, p.R172X; c.547C>T, p.R183X; c.226A>G, p.M76V). All had portal hypertension and, at liver biopsy, histopathological features of the ductal plate malformation (DPM)/congenital hepatic fibrosis (CHF). Four children required liver transplantation for recurrent gastrointestinal haemorrhage. DPM/CHF was confirmed at hepatectomy, with sclerosing small-duct cholangitis. Immunostaining for two primary-cilium axonemal proteins found expression that was deficient intraluminally and ectopic within cholangiocyte cytoplasm. ZFYVE19 depletion in cultured cells yielded abnormalities of centriole and axoneme. CONCLUSION: Biallelic ZFYVE19 mutations can lead to high-GGT cholestasis and DPM/CHF in vivo. In vitro, they can lead to centriolar and axonemal abnormalities. These observations indicate that mutation in ZFYVE19 results, through as yet undefined mechanisms, in a ciliopathy.

NBAS disease: 14 new patients, a recurrent mutation, and genotype-phenotype correlation among 24 Chinese patients.

AIM: Neuroblastoma amplified sequence (NBAS)-associated disease has a wide phenotypic spectrum, including infantile liver failure syndrome type 2 (ILFS2, OMIM #616483), short stature with optic nerve atrophy and Pelger-Huët anomaly (SOPH) syndrome (OMIM #614800), and a combined phenotype overlapping ILFS2 and SOPH syndrome. The mutation spectra of NBAS and its genotype-phenotype correlation among Chinese were not clear. METHODS: Clinical and genetic data were retrospectively collected from the medical charts of patients with biallelic NBAS mutations, as well as from Chinese patients in previously published reports. RESULTS: Fourteen new patients were identified, including 10 novel mutations: c.648-1G>A, c.2563_c.2577+5del/p.His855_Gln859del, c.3115C>T/p.Gln1039Ter, c.3284G>A/p.Trp1095Ter, c.2570C>T/p.Ala857Val, c.6859G>T/p.Asp2287Tyr, c.1028G>A/p.Ser343Asn, c.1177_1182delinsAGATAGA/p.Val393ArgfsTer2, c.3432_3435dupCAGT/p.Ala1146GlnfsTer14, and c.680_690dupACTGTTTCAGC/p.Phe231ThrfsTer35. All 14 patients presented as fever-triggered liver injury, including nine patients that satisfied the criteria of acute liver failure (ALF) in whom c.3596G>A/p.Cys1199Tyr occurred five times. Nine patients had extrahepatic manifestations including short stature, skeletal abnormalities, intellectual disability, ophthalmic abnormalities, low levels of serum immunoglobulins, facial dysmorphism, and cardiac abnormalities. Ten other Chinese patients were collected through a review of published works. Genotype-phenotype analysis in 24 Chinese patients revealed that the percentage of ALF patients with variants in the Sec39 domain was significantly higher than that in the C-terminal (100% vs. 12.5%, P = 0.000), and the percentage of multi-organ/system involvement in patients with variants in the Sec39 domain was significantly lower than that in the C-terminal (40% vs. 100%, P = 0.0128). CONCLUSIONS: We reported 14 new patients, 10 novel mutations, and a unique recurrent mutation. Correlation analysis indicated that the domain of missense and non-frameshift insertion/deletion mutations in NBAS protein is related to phenotype among Chinese patients.

TJP2 hepatobiliary disorders: Novel variants and clinical diversity.

To assess the spectrum of pediatric clinical phenotypes in TJP2 disease, we reviewed records of our seven patients in whom intrahepatic cholestasis was associated with biallelic TJP2 variants (13; 12 novel) and correlated clinical manifestations with mutation type. The effect of a splicing variant was analyzed with a minigene assay. The effects of three missense variants were analyzed with protein expression in vitro. Our patients had both remitting and persistent cholestasis. Three exhibited growth retardation. Six responded to treatment with cholestyramine, ursodeoxycholic acid, or both. Two had cholecystolithiasis. None required liver transplantation or developed hepatocellular or cholangiocellular malignancy. None manifested extrahepatic disease not attributable to effects of cholestasis. The variant c.2180-5T>G resulted in exon 15 skipping with in-frame deletion of 32 amino acid residues in TJP2. The three missense variants decreased but did not abolish TJP2 expression. Patients with truncating or canonical splice-site variants had clinically more severe disease. TJP2 disease in children includes a full clinical spectrum of severity, with mild or intermittent forms as well as the severe and minimal forms hitherto described. Biallelic TJP2 variants must be considered in children with clinically intermittent or resolved intrahepatic cholestasis.

A Molecular Mechanism Underlying Genotype-Specific Intrahepatic Cholestasis Resulting From MYO5B Mutations.

BACKGROUND AND AIMS: Progressive familial intrahepatic cholestasis (PFIC) 6 has been associated with missense but not biallelic nonsense or frameshift mutations in MYO5B, encoding the motor protein myosin Vb (myoVb). This genotype-phenotype correlation and the mechanism through which MYO5B mutations give rise to PFIC are not understood. The aim of this study was to determine whether the loss of myoVb or expression of patient-specific myoVb mutants can be causally related to defects in canalicular protein localization and, if so, through which mechanism. APPROACH AND RESULTS: We demonstrate that the cholestasis-associated substitution of the proline at amino acid position 600 in the myoVb protein to a leucine (P660L) caused the intracellular accumulation of bile canalicular proteins in vesicular compartments. Remarkably, the knockout of MYO5B in vitro and in vivo produced no canalicular localization defects. In contrast, the expression of myoVb mutants consisting of only the tail domain phenocopied the effects of the Myo5b-P660L mutation. Using additional myoVb and rab11a mutants, we demonstrate that motor domain-deficient myoVb inhibited the formation of specialized apical recycling endosomes and that its disrupting effect on the localization of canalicular proteins was dependent on its interaction with active rab11a and occurred at the trans-Golgi Network/recycling endosome interface. CONCLUSIONS: Our results reveal a mechanism through which MYO5B motor domain mutations can cause the mislocalization of canalicular proteins in hepatocytes which, unexpectedly, does not involve myoVb loss-of-function but, as we propose, a rab11a-mediated gain-of-toxic function. The results explain why biallelic MYO5B mutations that affect the motor domain but not those that eliminate myoVb expression are associated with PFIC6.

Compromised Air Quality and Healthcare Safety from Smoking inside Hospitals in Shantou, China.

Achieving smoke-free healthcare facilities remains a great challenge in countries with a high smoking prevalence and weak regulation. Assessment of the impact of environmental tobacco smoke (ETS) and its constituent PM2.5 on the air quality in Chinese hospitals has not been reported. In this study, we conducted air quality surveys by measuring real-time PM2.5 concentrations with Dylos Air Quality Monitors in five tertiary hospitals in Shantou, China during summer (July-August 2016) and winter (November-February 2017). Twenty-eight-day surveys inside the hospitals showed median PM2.5 concentrations above the China Air Quality Standard in elevator lobbies (51.0 µg/m3, IQR 34.5-91.7), restrooms (40.2, 27.1-70.3), and corridors (36.5, 23.0-77.4). Evidence of tobacco smoking was significantly associated with PM2.5 spikes observed in all the survey locations, contributing to the air quality undesirable for health in 49.1% of total survey hours or 29.3% of summer and 75.4% of winter survey hours inside the buildings, and 33.5%, 25.7%, and 6.8% of survey hours in doctor offices, nurse stations, and patient rooms, respectively. In conclusion, smoking inside hospitals induces PM2.5 spikes that significantly compromise the air quality and impose significant health risk to the hospital inhabitants. Reinforcing comprehensive smoking ban with the vested interest of all stakeholders followed by creative disciplinary actions are suggested to ensure healthcare safety.

A novel mutation in EYA1 in a Chinese family with Branchio-oto-renal syndrome.

BACKGROUND: Branchio-oto-renal (BOR) syndrome is a dominant autosomal disorder characterized by phenotypes such as hearing loss, branchial fistulae, preauricular pits, and renal abnormalities. EYA1, the human homolog of the Drosophila "eye absent" gene on chromosome 8q13.3, is recognized as one of the most important genes associated with BOR syndrome. METHODS: The proposita in this study was a 5-year-old Chinese girl with hearing loss, bilateral otitis media with effusion, microtia, facial hypoplasia, palatoschisis, and bilateral branchial cleft fistulae. The girl's family members, except two who were deceased, agreed to undergo clinical examination. We collected blood samples from 10 family members, including six who were affected by the syndrome. Genomic DNA was extracted and subjected to Sanger sequencing. A minigene assay was performed to confirm whether splicing signals were altered. In addition, we performed western blotting to determine alterations in protein levels of the wild-type and mutant gene. RESULTS: Clinical tests showed that some of the family members met the criteria for BOR syndrome. The affected members harbored a novel heterozygous nonsense variation in exon 11 of EYA1, whereas no unaffected member carried the mutation at this position. Functional experiments did not detect abnormal splicing at the RNA level; however, western blotting showed that the mutated protein was truncated. CONCLUSIONS: This study reports a novel mutation associated with BOR syndrome in a Chinese family. We highlight the usefulness of genetic testing in the diagnosis of BOR syndrome. Thus, we believe that this report would benefit clinicians in this field.

Novel NBAS mutations and fever-related recurrent acute liver failure in Chinese children: a retrospective study.

BACKGROUND: Underlying causes in Chinese children with recurrent acute liver failure (RALF), including liver crises less than full acute liver failure, are incompletely understood. We sought to address this by searching for genes mutated in such children. METHODS: Five unrelated Chinese boys presenting between 2012 and 2015 with RALF of unexplained etiology were studied. Results of whole exome sequencing were screened for mutations in candidate genes. Mutations were verified in patients and their family members by Sanger sequencing. All 5 boys underwent liver biopsy. RESULTS: NBAS was the only candidate gene mutated in more than one patient (biallelic mutations, 3 of 5 patients; 5 separate mutations). All NBAS mutations were novel and predictedly pathogenic (frameshift insertion mutation c.6611_6612insCA, missense mutations c.2407G > A and c.3596G > A, nonsense mutation c.586C > T, and splicing-site mutation c.5389 + 1G > T). Of these mutations, 3 lay in distal (C-terminal) regions of NBAS, a novel distribution. Unlike the 2 patients without NBAS mutations, the 3 patients with confirmed NBAS mutations all suffered from a febrile illness before each episode of liver crisis (fever-related RALF), with markedly elevated alanine aminotransferase and aspartate aminotransferase activities 24-72 h after elevation of body temperature, succeeded by severe coagulopathy and mild to moderate jaundice. CONCLUSIONS: As in other countries, so too in China; NBAS disease is a major cause of fever-related RALF in children. The mutation spectrum of NBAS in Chinese children seems different from that described in other populations.

Defects in myosin VB are associated with a spectrum of previously undiagnosed low γ-glutamyltransferase cholestasis.

Hereditary cholestasis in childhood and infancy with normal serum gamma-glutamyltransferase (GGT) activity is linked to several genes. Many patients, however, remain genetically undiagnosed. Defects in myosin VB (MYO5B; encoded by MYO5B) cause microvillus inclusion disease (MVID; MIM251850) with recurrent watery diarrhea. Cholestasis, reported as an atypical presentation in MVID, has been considered a side effect of parenteral alimentation. Here, however, we report on 10 patients who experienced cholestasis associated with biallelic, or suspected biallelic, mutations in MYO5B and who had neither recurrent diarrhea nor received parenteral alimentation. Seven of them are from two study cohorts, together comprising 31 undiagnosed low-GGT cholestasis patients; 3 are sporadic. Cholestasis in 2 patients was progressive, in 3 recurrent, in 2 transient, and in 3 uncategorized because of insufficient follow-up. Liver biopsy specimens revealed giant-cell change of hepatocytes and intralobular cholestasis with abnormal distribution of bile salt export pump (BSEP) at canaliculi, as well as coarse granular dislocation of MYO5B. Mass spectrometry of plasma demonstrated increased total bile acids, primary bile acids, and conjugated bile acids, with decreased free bile acids, similar to changes in BSEP-deficient patients. Literature review revealed that patients with biallelic mutations predicted to eliminate MYO5B expression were more frequent in typical MVID than in isolated-cholestasis patients (11 of 38 vs. 0 of 13). CONCLUSION: MYO5B deficiency may underlie 20% of previously undiagnosed low-GGT cholestasis. MYO5B deficiency appears to impair targeting of BSEP to the canalicular membrane with hampered bile acid excretion, resulting in a spectrum of cholestasis without diarrhea. (Hepatology 2017;65:1655-1669).

Wilson disease with hepatic presentation in an eight-month-old boy.

Wilson disease is an autosomal recessive disorder of copper metabolism that can cause fatal neurological and hepatic disease if not diagnosed and treated. The youngest child with normal liver function reported so far is an 8-mo-old Japanese boy with low ceruloplasmin levels, and the youngest child with elevated aminotransferase ever reported so far is a 9-mo-old Korean boy with confirmed by genetic testing. Here we report an 8-mo-old Chinese boy presented with elevated liver enzymes, and low serum ceruloplasmin level. Genetic analysis of ATP7B gene detected two heterozygous disease causing mutations (c.2621C>T/p.A874V and c.3809A>G/p.N1270S), and parental origins were determined. Persistent elevation of serum aminotransferase in this infant was normalized after zinc therapy. To our best knowledge, this is the youngest patient with elevated liver enzymes ever reported worldwide. We hope that this will raise awareness among pediatricians, leading to earlier diagnosis, timely treatment, and better clinical outcome.

JAG1 Mutation Spectrum and Origin in Chinese Children with Clinical Features of Alagille Syndrome.

Alagille syndrome is an autosomal dominant disorder that results from defects in the Notch signaling pathway, which is most frequently due to JAG1 mutations. This study investigated the rate, spectrum, and origin of JAG1 mutations in 91 Chinese children presenting with at least two clinical features of Alagille syndrome (cholestasis, heart murmur, skeletal abnormalities, ocular abnormalities, characteristic facial features, and renal abnormalities). Direct sequencing and/or multiplex-ligation-dependent probe amplification were performed in these patients, and segregation analysis was performed using samples available from the parents. JAG1 disease-causing mutations were detected in 70/91 (76.9%) patients, including 29/70 (41.4%) small deletions, 6/70 (8.6%) small insertions, 16/70 (22.9%) nonsense mutations, 8/70 (11.4%) splice-site mutations, 6/70 (9.4%) missense mutations, and 5/70 (7.1%) gross deletions. Of the mutations detected, 45/62 (72.6%) were novel, and almost all were unique, with the exception of c.439C>T, c.439+1G>A, c.703C>T, c.1382_1383delAC, c.2698C>T, and c.2990C>A, which were detected in two cases each; three cases exhibited entire gene deletions. A majority (69.2%) of the point and frameshift mutations could be detected by the sequencing of eleven exons (exons 3, 5, 6, 11, 14, 16, 18, 21, and 23-25). The mutation detection rate was 50.0% (10/20) in atypical cases that only presented with two or three clinical features of Alagille syndrome. Segregation analysis revealed that 81.1% (30/37) of these mutations were de novo. In conclusion, JAG1 mutations are present in the majority of Chinese pediatric patients with clinical features of Alagille syndrome, and the mutations concentrate on different exons from other reports. Genetic study is important for the diagnosis of atypical Alagille syndrome in Chinese patients.

Lysosomal dysfunction and impaired autophagy underlie the pathogenesis of amyloidogenic light chain-mediated cardiotoxicity.

AL amyloidosis is the consequence of clonal production of amyloidogenic immunoglobulin light chain (LC) proteins, often resulting in a rapidly progressive and fatal amyloid cardiomyopathy. Recent work has found that amyloidogenic LC directly initiate a cardio-toxic response underlying the pathogenesis of the cardiomyopathy; however, the mechanisms that contribute to this proteotoxicity remain unknown. Using human amyloidogenic LC isolated from patients with amyloid cardiomyopathy, we reveal that dysregulation of autophagic flux is critical for mediating amyloidogenic LC proteotoxicity. Restoration of autophagic flux by pharmacological intervention using rapamycin protected against amyloidogenic light chain protein-induced pathologies including contractile dysfunction and cell death at the cellular and organ level and also prolonged survival in an in vivo zebrafish model of amyloid cardiotoxicity. Mechanistically, we identify impaired lysosomal function to be the major cause of defective autophagy and amyloidogenic LC-induced proteotoxicity. Collectively, these findings detail the downstream molecular mechanisms underlying AL amyloid cardiomyopathy and highlight potential targeting of autophagy and lysosomal dysfunction in patients with amyloid cardiomyopathy.

Assessment of right ventricular structure and function in mouse model of pulmonary artery constriction by transthoracic echocardiography.

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure(1-3). Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction(4). To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential. A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software. Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC. Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Stanniocalcin1 is a key mediator of amyloidogenic light chain induced cardiotoxicity.

Immunoglobulin light chain (LC) amyloidosis (AL) results from overproduction of circulating amyloidogenic LC proteins and subsequent amyloid fibril deposition in organs. Mortality in AL amyloidosis patients is highly associated with a rapidly progressive AL cardiomyopathy, marked by profound impairment of diastolic and systolic cardiac function and significant early mortality. While myocardial fibril deposition contributes to the severe diastolic dysfunction seen in AL cardiomyopathy patients, the degree of fibril deposition has not been found to correlate with prognosis. Previously, we and others showed a direct cardiotoxic effect of amyloidogenic LC proteins (AL-LC), which may contribute to the pathophysiology and mortality observed in AL cardiomyopathy patients. However, the mechanisms underlying AL-LC related cardiotoxicity remain unknown. Mammalian stanniocalcin1 (STC1) is associated with a number of cellular processes including oxidative stress and cell death. Herein, we find that STC1 expression is elevated in cardiac tissue from AL cardiomyopathy patients, and is induced in isolated cardiomyocytes in response to AL-LC, but not non-amyloidogenic LC. STC1 overexpression in vitro recapitulates the pathophysiology of AL-LC mediated cardiotoxicity, with increased ROS production, contractile dysfunction and cell death. Overexpression of STC1 in vivo results in significant cardiac dysfunction and cell death. Genetic silencing of STC1 prevents AL-LC induced cardiotoxicity in cardiomyocytes and protects against AL-LC induced cell death and early mortality in zebrafish. The cardiotoxic effects of STC1 appears to be mediated via mitochondrial dysfunction as indicated by loss of mitochondrial membrane potential, ROS production and increased mitochondrial calcium levels. Collectively, this work identifies STC1 as a critical determinant of AL-LC cardiotoxicity.

PGC1α plays a critical role in TWEAK-induced cardiac dysfunction.

BACKGROUND: Inflammatory cytokines play an important role in the pathogenesis of heart failure. We have recently found the cytokine TWEAK (tumor necrosis factor (TNF)-like weak inducer of apoptosis), a member of the TNF superfamily, to be increased in patients with cardiomyopathy and result in the development of heart failure when overexpressed in mice. The molecular mechanisms underlying TWEAK-induced cardiac pathology, however, remain unknown. METHODOLOGY AND CRITICAL FINDING: Using mouse models of elevated circulating TWEAK levels, established through intravenous injection of adenovirus expressing TWEAK or recombinant TWEAK protein, we find that TWEAK induces a progressive dilated cardiomyopathy with impaired contractile function in mice. Moreover, TWEAK treatment is associated with decreased expression of peroxisome proliferator-activated receptor gamma coactivator-1α (PGC1α) and genes required for mitochondrial oxidative phosphorylation, which precede the onset of cardiac dysfunction. TWEAK-induced downregulation of PGC1α requires expression of its cell surface receptor, fibroblast growth factor-inducible 14 (Fn14). We further find that TWEAK downregulates PGC1α gene expression via the TNF receptor-associated factor 2 (TRAF2) and NFκB signaling pathways. Maintaining PGC1α levels through adenoviral-mediated gene expression is sufficient to protect against TWEAK-induced cardiomyocyte dysfunction. CONCLUSION: Collectively, our data suggest that TWEAK induces cardiac dysfunction via downregulation of PGC1α, through FN14-TRAF2-NFκB-dependent signaling. Selective targeting of the FN14-TRAF2-NFκB-dependent signaling pathway or augmenting PGC1α levels may serve as novel therapeutic strategies for cardiomyopathy and heart failure.

Intervention of cardiomyocyte death based on the impedance-sensing technique of monitoring cell adhesion.

Cardiomyocyte death is a critical risk factor in a variety of cardiovascular diseases [1, 2]. At the level of individual cardiomyocytes, cell adhesion alters at the early phase of ongoing death. It suggested the possibility of detecting the early sign of cell death through cell detachment and so that we could intervene in the cell response before cell fell into step with the irreversible death processes. In this work, we demonstrated the application of our impedance-sensing technique [3] in the intervention of cell responses to chemical stimuli. The results showed that the cell detachment could be detected with our impedance-sensing technique earlier than conventional biochemical staining techniques, and it could be used to guide the precautionary removal of TNF-alpha from the cell culture to intervene in the cell responses and prevent cardiomyocytes from cell death.