Rare coding variants in NOX4 link high ROS levels to psoriatic arthritis mutilans.

Psoriatic arthritis mutilans (PAM) is the rarest and most severe form of psoriatic arthritis, characterized by erosions of the small joints and osteolysis leading to joint disruption. Despite its severity, the underlying mechanisms are unknown, and no susceptibility genes have hitherto been identified. We aimed to investigate the genetic basis of PAM by performing massive parallel sequencing in sixty-one patients from the PAM Nordic cohort. We found rare variants in the NADPH oxidase 4 (NOX4) in four patients. In silico predictions show that the identified variants are potentially damaging. NOXs are the only enzymes producing reactive oxygen species (ROS). NOX4 is specifically involved in the differentiation of osteoclasts, the cells implicated in bone resorption. Functional follow-up studies using cell culture, zebrafish models, and measurement of ROS in patients uncovered that these NOX4 variants increase ROS levels both in vitro and in vivo. We propose NOX4 as the first candidate susceptibility gene for PAM. Our study links high levels of ROS caused by NOX4 variants to the development of PAM, offering a potential therapeutic target.

Loss of ctnnd2b affects neuronal differentiation and behavior in zebrafish.

Delta-catenin (CTNND2) is an adhesive junction associated protein belonging to the family of p120 catenins. The human gene is located on the short arm of chromosome 5, the region deleted in Cri-du-chat syndrome (OMIM #123450). Heterozygous loss of CTNND2 has been linked to a wide spectrum of neurodevelopmental disorders such as autism, schizophrenia, and intellectual disability. Here we studied how heterozygous loss of ctnnd2b affects zebrafish embryonic development, and larvae and adult behavior. First, we observed a disorganization of neuronal subtypes in the developing forebrain, namely the presence of ectopic isl1-expressing cells and a local reduction of GABA-positive neurons in the optic recess region. Next, using time-lapse analysis, we found that the disorganized distribution of is1l-expressing forebrain neurons resulted from an increased specification of Isl1:GFP neurons. Finally, we studied the swimming patterns of both larval and adult heterozygous zebrafish and observed an increased activity compared to wildtype animals. Overall, this data suggests a role for ctnnd2b in the differentiation cascade of neuronal subtypes in specific regions of the vertebrate brain, with repercussions in the animal's behavior.

The Beauty and the Beast: Bariatric Surgery and a Case of Oxalate Nephropathy.

Oxalate nephropathy is a rare cause of kidney failure. Roux-en-Y gastric bypass surgery is a technique used for surgical treatment of obesity as well as for the treatment of gastric carcinoma. We report the case of a 46-year-old male who was admitted to the nephrology department due to kidney dysfunction eight months after bariatric surgery.

A Missense Variant in PDK1 Associated with Severe Neurodevelopmental Delay and Epilepsy.

The pyruvate dehydrogenase complex (PDC) is responsible for the conversion of pyruvate into acetyl-CoA, which is used for energy conversion in cells. PDC activity is regulated by phosphorylation via kinases and phosphatases (PDK/PDP). Variants in all subunits of the PDC and in PDK3 have been reported, with varying phenotypes including lactic acidosis, neurodevelopmental delay, peripheral neuropathy, or seizures. Here, we report a de novo heterozygous missense variant in PDK1 (c.1139G > A; p.G380D) in a girl with developmental delay and early onset severe epilepsy. To investigate the role of PDK1G380D in energy metabolism and neuronal development, we used a zebrafish model. In zebrafish embryos we show a reduced number of cells with mitochondria with membrane potential, reduced movements, and a delay in neuronal development. Furthermore, we observe a reduction in the phosphorylation of PDH-E1α by PDKG380D, which suggests a disruption in the regulation of PDC activity. Finally, in patient fibroblasts, a mild reduction in the ratio of phosphorylated PDH over total PDH-E1α was detected. In summary, our findings support the notion that this aberrant PDK1 activity is the cause of clinical symptoms in the patient.

A complex DICER1 syndrome phenotype associated with a germline pathogenic variant affecting the RNase IIIa domain of DICER1.

BACKGROUND: Germline pathogenic variants in DICER1 cause DICER1 syndrome, an autosomal dominant, pleiotropic tumour predisposition syndrome with variable expressivity and reduced penetrance for specific dysplastic and neoplastic lesions. Recently, a syndrome with the acronym GLOW (Global developmental delay, Lung cysts, Overgrowth, Wilms tumour) was described in two children with mosaic missense mutations in hotspot residues of the DICER1 RNase IIIb domain. METHODS: Whole genome sequencing, exome sequencing, Sanger sequencing, digital PCR and a review of Wilms tumours with DICER1 RNase III domain mutations were performed. RESULTS: A de novo heterozygous c.4031C>T (p.S1344L) variant in the sequence encoding the RNase IIIa domain of DICER1 was detected. Clinical investigations revealed a phenotype that resembles the GLOW subphenotype of DICER1 syndrome. CONCLUSION: The phenotypic overlap between patients with p.S1344L mutation and GLOW syndrome provide clinical support for recent discoveries that RNase IIIa-Ser1344 site mutations impede miRNA-5p biogenesis analogous to DICER1 hotspot mutations in the RNase IIIb domain. We show that an individual with a heterozygous germline p.S1344L mutation has a severe form of DICER1 syndrome ('DICER1 syndrome plus'), with notable features of intellectual disability, macrocephaly, physical abnormalities, Wilms tumour and a well-differentiated fetal adenocarcinoma of the lung.

Emerging Optical Materials in Sensing and Discovery of Bioactive Compounds.

Optical biosensors are used in numerous applications and analytical fields. Advances in these sensor platforms offer high sensitivity, selectivity, miniaturization, and real-time analysis, among many other advantages. Research into bioactive natural products serves both to protect against potentially dangerous toxic compounds and to promote pharmacological innovation in drug discovery, as these compounds have unique chemical compositions that may be characterized by greater safety and efficacy. However, conventional methods for detecting these biomolecules have drawbacks, as they are time-consuming and expensive. As an alternative, optical biosensors offer a faster, simpler, and less expensive means of detecting various biomolecules of clinical interest. In this review, an overview of recent developments in optical biosensors for the detection and monitoring of aquatic biotoxins to prevent public health risks is first provided. In addition, the advantages and applicability of these biosensors in the field of drug discovery, including high-throughput screening, are discussed. The contribution of the investigated technological advances in the timely and sensitive detection of biotoxins while deciphering the pathways to discover bioactive compounds with great health-promoting prospects is envisaged to meet the increasing demands of healthcare systems.

ANCA-associated vasculitis in a patient with enteropathic spondylarthritis: a case report and literature review.

Coexistence of antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) and inflammatory bowel disease (IBD) is rare (Sy et al. in Semin Arthritis Rheum 45:475-482, 2016). Nevertheless, we present a case of an AAV in a 53-year-old female with enteropathic spondylarthritis previously treated with tumor necrosis factor α inhibitors (TNFi). Management of vasculitis in a patient with IBD may be problematic due to the difficulty in distinguishing if the vasculitis is an extraintestinal manifestation of the IBD or a new coexistent entity. Moreover, in our report, the previous treatment with TNFi is a possible confounding factor due to the paradoxical effects induced by TNFi, including vasculitis (Ramos-Casals et al. in Curr Rheumatol Rep 10:442-448, 2008). The reported case alerts to the complexity in the management of patients with enteropathic spondylarthritis and vasculitis, as well as discusses the diversity of differential diagnosis in this particular clinical scenario.

FLNC Expression Level Influences the Activity of TEAD-YAP/TAZ Signaling.

Filamin C (FLNC), being one of the major actin-binding proteins, is involved in the maintenance of key muscle cell functions. Inherited skeletal muscle and cardiac disorders linked to genetic variants in FLNC have attracted attention because of their high clinical importance and possibility of genotype-phenotype correlations. To further expand on the role of FLNC in muscle cells, we focused on detailed alterations of muscle cell properties developed after the loss of FLNC. Using the CRISPR/Cas9 method we generated a C2C12 murine myoblast cell line with stably suppressed Flnc expression. FLNC-deficient myoblasts have a significantly higher proliferation rate combined with an impaired cell migration capacity. The suppression of Flnc expression leads to inability to complete myogenic differentiation, diminished expression of Myh1 and Myh4, alteration of transcriptional dynamics of myogenic factors, such as Mymk and Myog, and deregulation of Hippo signaling pathway. Specifically, we identified elevated basal levels of Hippo activity in myoblasts with loss of FLNC, and ineffective reduction of Hippo signaling activity during myogenic differentiation. The latter was restored by Flnc overexpression. In summary, we confirmed the role of FLNC in muscle cell proliferation, migration and differentiation, and demonstrated for the first time the direct link between Flnc expression and activity of TEAD-YAP\TAZ signaling. These findings support a role of FLNC in regulation of essential muscle processes relying on mechanical as well as signaling mechanisms.

Photonics in nature and bioinspired designs: sustainable approaches for a colourful world.

Biological systems possess nanoarchitectures that have evolved for specific purposes and whose ability to modulate the flow of light creates an extraordinary diversity of natural photonic structures. In particular, the striking beauty of the structural colouration observed in nature has inspired technological innovation in many fields. Intense research has been devoted to mimicking the unique vivid colours with newly designed photonic structures presenting stimuli-responsive properties, with remarkable applications in health care, safety and security. This review highlights bioinspired photonic approaches in this context, starting by presenting many appealing examples of structural colours in nature, followed by describing the versatility of fabrication methods and designed coloured structures. A particular focus is given to optical sensing for medical diagnosis, food control and environmental monitoring, which has experienced a significant growth, especially considering the advances in obtaining inexpensive miniaturized systems, more reliability, fast responses, and the use of label-free layouts. Additionally, naturally derived biomaterials and synthetic polymers are versatile and fit many different structural designs that are underlined. Progress in bioinspired photonic polymers and their integration in novel devices is discussed since recent developments have emerged to lift the expectations of smart, flexible, wearable and portable sensors. The discussion is expanded to give emphasis on additional functionalities offered to related biomedical applications and the use of structural colours in new sustainable strategies that could meet the needs of technological development.

Truncating Variant in Myof Gene Is Associated With Limb-Girdle Type Muscular Dystrophy and Cardiomyopathy.

Even though genetic studies of individuals with neuromuscular diseases have uncovered the molecular background of many cardiac disorders such as cardiomyopathies and inherited arrhythmic syndromes, the genetic cause of a proportion of cardiomyopathies associated with neuromuscular phenotype still remains unknown. Here, we present an individual with a combination of cardiomyopathy and limb-girdle type muscular dystrophy where whole exome sequencing identified myoferlin (MYOF)-a member of the Ferlin protein family and close homolog of DYSF-as the most likely candidate gene. The disease-causative role of the identified variant c.[2576delG; 2575G>C], p.G859QfsTer8 is supported by functional studies in vitro using the primary patient's skeletal muscle mesenchymal progenitor cells, including both RNA sequencing and morphological studies, as well as recapitulating the muscle phenotype in vivo in zebrafish. We provide the first evidence supporting a role of MYOF in human muscle disease.

Zebrafish Models of Neurodevelopmental Disorders: Limitations and Benefits of Current Tools and Techniques.

For the past few years there has been an exponential increase in the use of animal models to confirm the pathogenicity of candidate disease-causing genetic variants found in patients. One such animal model is the zebrafish. Despite being a non-mammalian animal, the zebrafish model has proven its potential in recapitulating the phenotypes of many different human genetic disorders. This review will focus on recent advances in the modeling of neurodevelopmental disorders in zebrafish, covering aspects from early brain development to techniques used for modulating gene expression, as well as how to best characterize the resulting phenotypes. We also review other existing models of neurodevelopmental disorders, and the current efforts in developing and testing compounds with potential therapeutic value.

Avaliação das práticas de higienização das mãos em três unidades de terapia intensiva / Evaluation of hand hygiene practices in three intensive care units / Evaluación de lãs prácticas de higienización de las manos em la unidades de terapia intensiva

Justificativa e Objetivos: Higienização das Mãos (HM) é uma medida simples e eficaz no combate às Infecções Relacionadas à Assistência à Saúde (IRAS). Apesar do esforço feito pelos profissionais do Serviço de Controle de Infecção Hospitalar (SCIH), o cumprimento das práticas ainda permanece baixo, geralmente inferior a 50%. Por esse motivo, este estudo objetivou avaliar as práticas de HM em três Unidades de Terapia Intensiva de um hospital privado de Belo Horizonte, MG, Brasil. Método: Estudo transversal, descritivo de natureza quantitativa, realizado em três UTIs de um hospital privado de Belo Horizonte, MG, Brasil. No período de julho/2016 a julho/2017 foram avaliados todos os documentos de HM do hospital de estudo, totalizando 1.100 observações in loco realizadas pelos enfermeiros do SCIH. Para análise dos dados adotou-se estatística descritiva e regressão linear utilizando o coeficiente de correlação de Pearson. Resultados: A taxa de adesão à HM variou entre 27,3 e 88,2%, com média de 47% (±16,8). O consumo de preparação alcoólica e sabonete líquido obtiveram, respectivamente, uma média de 32 e 47 ml/paciente-dia. Houve correlação positiva entre o consumo de preparação alcoólica e a taxa de adesão (r2=0,49, p=0,01). Conclusão: Os profissionais de saúde ainda necessitam de sensibilização acerca da importância da higiene de mãos para redução das IRAS.(AU)

Background and Objectives: Hand Hygiene (HH) is a simple and effective measure in the fight against Healthcare-associated Infections (HAIs). Despite the effort made by professionals of the Infection Control Service, compliance with practices remains low, generally below 50%. For this reason, this study aimed to evaluate HH practices in three Intensive Care Units of a private hospital in Belo Horizonte, MG, Brazil. Methods: Cross-sectional, descriptive study performed in three ICUs of a private hospital in Belo Horizonte, MG, Brazil. From July 2016 to July 2017, all HH documents of the study hospital were evaluated, totaling 1,100 observations in loco performed by the nurses of the Infection Control Service. Data analysis was performed using descriptive statistics and linear regression using the Pearson correlation coefficient. Results: The rate of adhesion ranged from 27.3 to 88.2%, with an average of 47% (±16.8). Consumption of alcohol preparation and liquid soap obtained, respectively, an average of 32 and 47 ml/patient-day. Pearson's coefficient showed a positive correlation between alcohol consumption and adhesion rate (r=0.49, p=0.01). Conclusion: Healthcare workers still need to be aware of the importance of HH for reducing HAIs.(AU)

Justificación y objetivos: La Higienización de las Manos (HM) es una medida simple y eficaz en el combate a las Infecciones Relacionadas a la Asistencia Sanitaria (IRAS). A pesar del esfuerzo realizado por los profesionales del Servicio de Control de Infección Hospitalaria, el cumplimiento de las prácticas sigue siendo bajo, generalmente inferior al 50%. Por este motivo, este estudio objetivó evaluar las prácticas de HM en tres Unidades de Terapia Intensiva de un hospital privado de Belo Horizonte, MG, Brasil. Método: Estudio transversal, descriptivo de naturaleza cuantitativa, realizado en tres UTIs de un hospital privado de Belo Horizonte, MG, Brasil. En el período de julio/2016 a julio/2017 se evaluaron todos los documentos de HM del hospital de estudio, totalizando 1.100 observaciones in loco realizadas por los enfermeros del Servicio de Control de Infección. Para el análisis de los datos se adoptó estadística descriptiva y regresión lineal utilizando el coeficiente de correlación de Pearson. Resultados: La tasa de adhesión las prácticas de HM varió entre el 27,3 y el 88,2%, con una media del 47% (±16,8). El consumo de preparación alcohólica y jabón líquido obtuvieron, respectivamente, una media de 32 y 47 ml / paciente-día. Se observó una correlación positiva entre el consumo de alcohol y la tasa de adhesión (r2=0,49, p=0,01). Conclusiones: Los profesionales de la salud todavía necesitan sensibilización acerca de la importancia de la HM para reducir las IRAS.(AU)

Alu-Alu mediated intragenic duplications in IFT81 and MATN3 are associated with skeletal dysplasias.

Skeletal dysplasias are a diverse group of rare Mendelian disorders with clinical and genetic heterogeneity. Here, we used targeted copy number variant (CNV) screening and identified intragenic exonic duplications, formed through Alu-Alu fusion events, in two individuals with skeletal dysplasia and negative exome sequencing results. First, we detected a homozygous tandem duplication of exon 9 and 10 in IFT81 in a boy with Jeune syndrome, or short-rib thoracic dysplasia (SRTD) (MIM# 208500). Western blot analysis did not detect any wild-type IFT81 protein in fibroblasts from the patient with the IFT81 duplication, but only a shorter isoform of IFT81 that was also present in the normal control samples. Complementary zebrafish studies suggested that loss of full-length IFT81 protein but expression of a shorter form of IFT81 protein affects the phenotype while being compatible with life. Second, a de novo tandem duplication of exons 2 to 5 in MATN3 was identified in a girl with multiple epiphyseal dysplasia (MED) type 5 (MIM# 607078). Our data highlights the importance of detection and careful characterization of intragenic duplication CNVs, presenting them as a novel and very rare genetic mechanism in IFT81-related Jeune syndrome and MATN3-related MED.

De novo mutations in FLNC leading to early-onset restrictive cardiomyopathy and congenital myopathy.

Mutations in FLNC for a long time are known in connection to neuromuscular disorders and only recently were described in association with various cardiomyopathies. Here, we report a new clinical phenotype of filaminopathy in four unrelated patients with early-onset restrictive cardiomyopathy (RCM) in combination with congenital myopathy due to FLNC mutations (NM_001458.4:c.3557C>T, p.A1186V, rs1114167361 in three probands and c.[3547G>C; 3548C>T], p.A1183L, rs1131692185 in one proband). In all cases, concurrent myopathy was confirmed by neurological examination, electromyography, and morphological studies. Three of the patients also presented with arthrogryposis. The pathogenicity of the described missense variants was verified by cellular and morphological studies and by in vivo modeling in zebrafish. Combination of in silico and experimental approaches revealed that FLNC missense variants localized in Ig-loop segments often lead to development of RCM. The described FLNC mutations associated with early-onset RCMP extend cardiac spectrum of filaminopathies and facilitate the differential diagnosis of restrictive cardiac phenotype associated with neuromuscular involvement in children.

A Multicenter Cohort Study of Histologic Findings and Long-Term Outcomes of Kidney Disease in Women Who Have Been Pregnant.

BACKGROUND AND OBJECTIVES: For many women pregnancy is the first contact with health services, thus providing an opportunity to identify renal disease. This study compares causes and long-term renal outcomes of biopsy-proven renal disease identified during pregnancy or within 1 year postpartum, with nonpregnant women. DESIGN, SETTING, PARTICIPANTS, & MEASUREMENTS: Native renal biopsies (1997-2012), in women of childbearing age (16 to <50 years), from 21 hospitals were studied. The pregnancy-related diagnosis group included those women with abnormal urinalysis/raised creatinine identified during pregnancy or within 1 year postpartum. Pregnancy-related and control biopsies were matched for age and ethnicity (black versus nonblack). RESULTS: One hundred and seventy-three pregnancy-related biopsies (19 antenatal, 154 postpregnancy) were identified and matched with 1000 controls. FSGS was more common in pregnancy-related biopsies (32.4%) than controls (9.7%) (P<0.001) but there were no differences in Columbia classification. Women with a pregnancy-related diagnosis were younger (32.1 versus 34.2 years; P=0.004) and more likely to be black (26.0% versus 13.3%; P<0.001) than controls, although there were no differences in ethnicities in women with FSGS. The pregnancy-related group (excluding antenatal biopsies) was more likely to have a decline in Chronic Kidney Disease Epidemiology Collaboration eGFR in the follow-up period than the control group (odds ratio, 1.67; 95% confidence interval, 1.03 to 2.71; P=0.04), and this decline appeared to be more rapid (-1.33 versus -0.56 ml/min per 1.73 m2 per year, respectively; P=0.045). However, there were no differences between groups in those who required RRT or who died. CONCLUSIONS: Pregnancy is an opportunity to detect kidney disease. FSGS is more common in women who have been pregnant than in controls, and disease identified in pregnancy or within 1 year postpartum is more likely to show a subsequent decline in renal function. Further work is required to determine whether pregnancy initiates, exacerbates, or reveals renal disease.

Trends of teenage pregnancy in Brazil, 2000-2011.

OBJECTIVE: To evaluate the frequency of teenage pregnancy in Brazil, from 2000 to 2011, in all five Brazilian macroregions and age groups (10-14 and 15-19 years), correlating it with the human development index (HDI). METHOD: Descriptive epidemiological study, with cross-sectional design, performed by searching the database of the National Health System (Datasus), using information from the Information System (Sinasc). RESULTS: There was a decrease in the percentage of live births (LB) from teenage mothers (10-19 years) in Brazil (23.5 % in 2000 to 19.2 % in 2011). This reduction was observed in all Brazilian macroregions in the group of mothers aged 15 to 19 years. The number of LB increased by 5.0% among mothers aged 10-14 years (increase in the North and Northeast and decline in the other macroregions). The proportion of LB shows an inversely proportional trend to HDI score, with the Southeast having the highest HDI and the lowest proportion of LB to teenage mothers in the country. CONCLUSION: Brazil shows a decline in the percentage of LB to adolescent mothers, tending to be inversely related to HDI score. It is important to empower strategies to address the problem, so that teenage pregnancy is seen as a personal decision rather than the result of a lack of policies targeting adolescent health.

Trends of teenage pregnancy in Brazil, 2000-2011 / Tendências da gravidez na adolescência no Brasil, 2000-2011

Summary Objective: To evaluate the frequency of teenage pregnancy in Brazil, from 2000 to 2011, in all five Brazilian macroregions and age groups (10-14 and 15-19 years), correlating it with the human development index (HDI). Method: Descriptive epidemiological study, with cross-sectional design, performed by searching the database of the National Health System (Datasus), using information from the Information System (Sinasc). Results: There was a decrease in the percentage of live births (LB) from teenage mothers (10-19 years) in Brazil (23.5 % in 2000 to 19.2 % in 2011). This reduction was observed in all Brazilian macroregions in the group of mothers aged 15 to 19 years. The number of LB increased by 5.0% among mothers aged 10-14 years (increase in the North and Northeast and decline in the other macroregions). The proportion of LB shows an inversely proportional trend to HDI score, with the Southeast having the highest HDI and the lowest proportion of LB to teenage mothers in the country. Conclusion: Brazil shows a decline in the percentage of LB to adolescent mothers, tending to be inversely related to HDI score. It is important to empower strategies to address the problem, so that teenage pregnancy is seen as a personal decision rather than the result of a lack of policies targeting adolescent health.

Resumo Objetivo: avaliar a frequência da gravidez na adolescência no Brasil, no período entre 2000 e 2011, nas cinco diferentes regiões brasileiras e por faixas de idade (10 a 14 e 15 a 19 anos), correlacionando com o índice de desenvolvimento humano (IDH). Método: estudo epidemiológico, descritivo, com desenho transversal, realizado por busca no banco de dados do sistema único de saúde (Datasus), utilizando informações do Sistema de Informação sobre Nascidos Vivos (Sinasc). Resultados: ocorreu queda do percentual de nascidos vivos (NV) de mães adolescentes (10 a 19 anos) no Brasil (23,5% em 2000 para 19,2% em 2011). Essa redução foi notada em todas as regiões brasileiras na parcela de mães entre 15 e 19 anos. O número de NV aumentou 5,0% entre mães de 10 a 14 anos (incremento no Norte e Nordeste e redução nas demais regiões). A proporção de NV mostra tendência inversamente proporcional ao IDH, tendo o Sudeste o maior IDH e a menor proporção de NV de mães adolescentes no país. Conclusão: o Brasil apresenta declínio do percentual de NV de mães adolescentes, com tendência a estar inversamente relacionado ao IDH. É importante intensificar as estratégias de abordagem do problema, a fim de que a gravidez na adolescência seja uma decisão própria e não consequência da falta de políticas públicas direcionadas ao adolescente.

Bone morphogenetic protein/retinoic acid inducible neural-specific protein (brinp) expression during Danio rerio development.

Prototype Membrane Attack Complex/Perforin (MACPF) superfamily proteins such as complement and perforin play crucial roles in immune defense where they drive lytic pore formation. However, it is evident that other MACPF family members are important in the central nervous system. For example, three bone morphogenetic protein/retinoic acid inducible neural-specific proteins (Brinp1, Brinp2 and Brinp3) are present in developing and mature mammalian neurons, but their molecular function is unknown. In this study we have identified and cloned full-length orthologues of all three human brinps from Danio rerio (zebrafish). Zebrafish and human brinps show very high sequence conservation, and the chromosomal loci are syntenic. We also identified two additional brinp3 paralogues at a separate locus in the zebrafish genome. The spatiotemporal expression of all five zebrafish brinps was determined by RT-PCR and whole mount RNA in situ hybridisation. Each brinp is expressed broadly in the developing nervous system at early stages (24 hours post fertilisation), but localises to specific structures in older embryos (48-72 hpf), as has been reported in mice. The conserved structures and spatiotemporal expression patterns of brinps reported in this study suggest that zebrafish will be useful for generating loss of function phenotypes to assist in determining the molecular role of these proteins.

Comparison of different numerical treatments for x-ray phase tomography of soft tissue from differential phase projections.

X-ray imaging of soft tissue is made difficult by their low absorbance. The use of x-ray phase imaging and tomography can significantly enhance the detection of these tissues and several approaches have been proposed to this end. Methods such as analyzer-based imaging or grating interferometry produce differential phase projections that can be used to reconstruct the 3D distribution of the sample refractive index. We report on the quantitative comparison of three different methods to obtain x-ray phase tomography with filtered back-projection from differential phase projections in the presence of noise. The three procedures represent different numerical approaches to solve the same mathematical problem, namely phase retrieval and filtered back-projection. It is found that obtaining individual phase projections and subsequently applying a conventional filtered back-projection algorithm produces the best results for noisy experimental data, when compared with other procedures based on the Hilbert transform. The algorithms are tested on simulated phantom data with added noise and the predictions are confirmed by experimental data acquired using a grating interferometer. The experiment is performed on unstained adult zebrafish, an important model organism for biomedical studies. The method optimization described here allows resolution of weak soft tissue features, such as muscle fibers.

Zebrafish models of BAG3 myofibrillar myopathy suggest a toxic gain of function leading to BAG3 insufficiency.

Mutations in the co-chaperone Bcl2-associated athanogene 3 (BAG3) can cause myofibrillar myopathy (MFM), a childhood-onset progressive muscle disease, characterized by the formation of protein aggregates and myofibrillar disintegration. In contrast to other MFM-causing proteins, BAG3 has no direct structural role, but regulates autophagy and the degradation of misfolded proteins. To investigate the mechanism of disease in BAG3-related MFM, we expressed wild-type BAG3 or the dominant MFM-causing BAG3 (BAG3(P209L)) in zebrafish. Expression of the mutant protein results in the formation of aggregates that contain wild-type BAG3. Through the stimulation and inhibition of autophagy, we tested the prevailing hypothesis that impaired autophagic function is responsible for the formation of protein aggregates. Contrary to the existing theory, our studies reveal that inhibition of autophagy is not sufficient to induce protein aggregation. Expression of the mutant protein, however, did not induce myofibrillar disintegration and we therefore examined the effect of knocking down Bag3 function. Loss of Bag3 resulted in myofibrillar disintegration, but not in the formation of protein aggregates. Remarkably, BAG3(P209L) is able to rescue the myofibrillar disintegration phenotype, further demonstrating that its function is not impaired. Together, our knockdown and overexpression experiments identify a mechanism whereby BAG3(P209L) aggregates form, gradually reducing the pool of available BAG3, which eventually results in BAG3 insufficiency and myofibrillar disintegration. This mechanism is consistent with the childhood onset and progressive nature of MFM and suggests that reducing aggregation through enhanced degradation or inhibition of nucleation would be an effective therapy for this disease.