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Artikel ini membicarakan pentingnya pemuridan dilakukan seorang ayah terhadap anak-anak dalam keluarga. Bagaimana kedudukan dan peranan ayah sebagai imam yang melaksanakan pemuridan kepada anak-anak dalam keluarga. Dalam artikel ini akan dibicarakan kualitas keluarga yang disebabkan oleh seorang ayah yang melakukan perannya yakni memuridkan anak-anak dalam keluarga. Tujuan dari artikel ini ialah: pertama, untuk mengingatkan para ayah agar mereka peduli terhadap pendidikan rohani anak-anak mereka yang dapat dimulai dari anak yang baru lahir hingga dewasa. Kedua, agar pendidikan rohani anak-anak dalam keluarga dilakukan secara merata baik kepada anak-anak. Seorang ayah bertanggung jawab untuk menjaga kualitas pendidikan rohani anak selama mereka masih berada di rumah. Ketiadaan pemuridan seorang ayah terhadap anak-anaknya akan mengakibatkan lemahnya keimaman dalam keluarga dan mudahnya meninggalkan imannya. Penelitian ini menggunakan pendekatan kualitatif dengan metode deskriptif analisis. Di akhir pembahasan dikemukakan model pemuridan seorang ayah dalam keluarga sebagai usulan.

Amyloid aggregation is a ubiquitous form of protein misfolding underlying the pathologies of Alzheimer’s disease (AD), Parkinson’s disease (PD), and type 2 diabetes (T2D), three primary forms of human amyloid diseases. While much has been learned about the origin, diagnosis, and management of these neurological and metabolic disorders, no cure is currently available due in part to the dynamic and heterogeneous nature of the toxic oligomers induced by amyloid aggregation. Here, we synthesized β-casein-coated iron oxide nanoparticles (βCas IONPs) via a 2-(bis-((phosphonic acid)methyl)amino)ethyl-(poly(oligo ethylene glycol)methyl ether acrylate)-b-(poly(N-ethyl-2,3-dibromomaleimide)acrylate)-((butylthio)-carbonothioyl)thio propionate (BPA-P(OEGA-b-DBM)) block copolymer linker. Using a thioflavin T kinetic assay, transmission electron microscopy, Fourier transform infrared spectroscopy, discrete molecular dynamics simulations, and cell viability assays, we examined the Janus characteristics and the inhibition potential of βCas IONPs against the aggregation of amyloid β (Aβ), α synuclein (αS), and human islet amyloid polypeptide (IAPP), which are implicated in the pathologies of AD, PD, and T2D. Incubation of zebrafish embryos with the amyloid proteins largely inhibited hatching and elicited reactive oxygen species, which were effectively rescued by the inhibitor. Furthermore, Aβ-induced damage to the mouse brain was mitigated in vivo with the inhibitor. This study revealed the potential of Janus nanoparticles as a new nanomedicine against a diverse range of amyloid diseases.

Mucopolysaccharidoses are lysosomal storage diseases that collectively represent a major cause of lethal, treatment-refractory childhood dementias 1–7 Clinically-useful interventions are hampered due to an incomplete understanding of their neuropathological origins. Using the zebrafish sgsh model of mucopolysaccharidosis IIIA 8 (MPS IIIA, Sanfilippo syndrome A), we conducted several ‘omics-based analyses, and developed and benchmarked a novel bioinformatic feature classification and ranking model for high-throughput datasets – ExIR – to prioritise important features in the progression of neurological manifestations of the disease. We find that the massive endolysosomal burden resulting from increased lysosomal storage of heparan sulfate and other secondarily accumulating substrates, such as sphingolipids, induces abnormal microtubule organisation and vesicle trafficking in neurons. This results in a gradual impairment of synaptic vesicle localisation at the presynaptic terminal and consequently impaired neuronal activity. Importantly, the endolysosomal phenotype in MPS IIIA zebrafish well-precedes the onset of neural pathology, though the larval MPS IIIA brain was found to be more susceptible to perturbation than wild type siblings. Collectively, these analyses demonstrate the presence of a progressive ‘functional neurodegenerative’ phenotype underpinning neurological disease in MPS IIIA. Our findings provide direct mechanistic evidence linking the well-described lysosomal storage basis for MPS IIIA to its disproportionately severe neural clinical involvement, enabling development and refinement of future therapeutic interventions for this currently untreatable disorder.HighlightsMPS IIIA represents one of the most common causes of broadly fatal childhood dementia, but the mechanisms underlying disease progression are poorly understood.The first systems-level analyses of disease state and progression in the CNS of an MPS IIIA animal model were performed.Experimental data-based Integrative Ranking (ExIR) was developed to provide unbiased prioritisation and classification of biological data as drivers, biomarkers and mediators of biological processes from high-throughput data at a systems level.Application of ExIR to a transcriptomic and proteomic analyses of a zebrafish model of MPS IIIA implies progressive deficiencies in synaptic activity as a key driver of disease progression correlating with progressive neuronal endolysosomal burden and secondary storage diseases.A novel unifying explanation of pathobiology and progression of MPS IIIA facilitates identification of clinically targetable features and may be generalised to other neuronopathic storage disorders.

Purpose: This retrospective audit aimed to evaluate the impact of combined iStent®Inject (iSI) and phacoemulsification on medication number in Australians with open-angle glaucoma. Secondary outcomes included intraocular pressure (IOP), best-corrected visual acuity, refraction and visual fields. Patients and Methods: Patients with glaucoma that received combined iSI and phacoemulsification by the same surgeon between 1 February 2016 and 1 February 2018 were audited for postoperative medication number, pressures after 1 day, 1 week, 4 weeks and 6, 12, 18 and 24 months, visual acuity, refraction and visual fields. These parameters were compared to baseline levels and with those from a separate cohort of patients without glaucoma that received standalone phacoemulsification. Results: Forty-one patients (63 eyes) received the combined procedure. Thirty-four patients (59 eyes) received standalone phacoemulsification. Four weeks after receiving combined iSI and phacoemulsification the mean medication number was significantly reduced by 1.3 (p < 0.001) for those on medication at baseline and by 0.5 (p = 0.002) overall. Mean IOP was significantly reduced from baseline after 6 months (-16%; p = 0.012; n = 35) and 12 months (-29%; p = 0.004; n = 16). Patients receiving standalone phacoemulsification had short-term reductions in IOP at 4 weeks (-8%; p < 0.001; n = 57) and 6 months (-16%; p < 0.001; n = 32). These patients without glaucoma had lower pressures overall compared to those with glaucoma that received the combined procedure (p = 0.019). There were no differences in final visual acuity or refractive outcomes between groups. Conclusion: This audit suggests that iSI and phacoemulsification are at least as effective in controlling IOP as medical therapy. It may have an important role in reducing the medication burden in Australians with cataract and glaucoma. This study is one of the first to confirm refractive stability in concomitant iSI and phacoemulsification.

Mucopolysaccharidosis IIIA (MPS IIIA, Sanfilippo syndrome type A), a paediatric neurological lysosomal storage disease, is caused by impaired function of the enzyme N-sulfoglucosamine sulfohydrolase (SGSH) resulting in impaired catabolism of heparan sulfate glycosaminoglycan (HS GAG) and its accumulation in tissues. MPS IIIA represents a significant proportion of childhood dementias. This condition generally leads to patient death in the teenage years, yet no effective therapy exists for MPS IIIA and a complete understanding of the mechanisms of MPS IIIA pathogenesis is lacking. Here, we employ targeted CRISPR/Cas9 mutagenesis to generate a model of MPS IIIA in the zebrafish, a model organism with strong genetic tractability and amenity for high-throughput screening. The sgshΔex5−6 zebrafish mutant exhibits a complete absence of Sgsh enzymatic activity, leading to progressive accumulation of HS degradation products with age. sgshΔex5−6 zebrafish faithfully recapitulate diverse CNS-specific features of MPS IIIA, including neuronal lysosomal overabundance, complex behavioural phenotypes, and profound, lifelong neuroinflammation. We further demonstrate that neuroinflammation in sgshΔex5−6 zebrafish is largely dependent on interleukin-1β and can be attenuated via the pharmacological inhibition of Caspase-1, which partially rescues behavioural abnormalities in sgshΔex5−6 mutant larvae in a context-dependent manner. We expect the sgshΔex5−6 zebrafish mutant to be a valuable resource in gaining a better understanding of MPS IIIA pathobiology towards the development of timely and effective therapeutic interventions.

Cell adhesion molecules are membrane-bound proteins predominantly expressed in the central nervous system along principal axonal pathways with key roles in nervous system development, neural cell differentiation and migration, axonal growth and guidance, myelination, and synapse formation. Here, we describe ten affected individuals with bi-allelic variants in the neuronal cell adhesion molecule NRCAM that lead to a neurodevelopmental syndrome of varying severity; the individuals are from eight families. This syndrome is characterized by developmental delay/intellectual disability, hypotonia, peripheral neuropathy, and/or spasticity. Computational analyses of NRCAM variants, many of which cluster in the third fibronectin type III (Fn-III) domain, strongly suggest a deleterious effect on NRCAM structure and function, including possible disruption of its interactions with other proteins. These findings are corroborated by previous in vitro studies of murine Nrcam-deficient cells, revealing abnormal neurite outgrowth, synaptogenesis, and formation of nodes of Ranvier on myelinated axons. Our studies on zebrafish nrcama(Delta) mutants lacking the third Fn-III domain revealed that mutant larvae displayed significantly altered swimming behavior compared to wild-type larvae (p < 0.03). Moreover, nrcama(Delta) mutants displayed a trend toward increased amounts of alpha-tubulin fibers in the dorsal telencephalon, demonstrating an alteration in white matter tracts and projections. Taken together, our study provides evidence that NRCAM disruption causes a variable form of a neurodevelopmental disorder and broadens the knowledge on the growing role of the cell adhesion molecule family in the nervous system.

BackgroundCell therapy of diabetes aims at restoring the physiological control of blood glucose by transplantation of functional pancreatic islet cells. A potentially unlimited source of cells for such transplantations would be islet cells derived from an in vitro differentiation of human pluripotent stem cells (hESC/hiPSC). The islet-like clusters (ILC) produced by the known differentiation protocols contain various cell populations. Among these, the β-cells that express both insulin and the transcription factor Nkx6.1 seem to be the most efficient to restore normoglycemia in diabetes animal models. Our aim was to find markers allowing selection of these efficient cells.MethodsFunctional Cell-Capture Screening (FCCS) was used to identify markers that preferentially capture the cells expressing both insulin and Nkx6.1, from hESC-derived ILC cells. In order to test whether selection for such markers could improve cell therapy in diabetic mouse models, we used ILC produced from a clinical-grade line of hESC by a refined differentiation protocol adapted to up-scalable bioreactors. Re-aggregated MACS sorted cells were encapsulated in microspheres made of alginate modified to reduce foreign body reaction. Implantation was done intraperitoneally in STZ-treated C57BL/6 immuno-competent mice.ResultsCD49A (integrin alpha1) was identified by FCCS as a marker for cells that express insulin (or C-peptide) as well as Nkx6.1 in ILC derived by hESC differentiation. The ILC fraction enriched in CD49A+ cells rapidly reduced glycemia when implanted in diabetic mice, whereas mice receiving the CD49A depleted population remained highly diabetic. CD49A-enriched ILC cells also produced higher levels of human C-peptide in the blood of transplanted mice. However, the difference between CD49A-enriched and total ILC cells remained small. Another marker, CD26 (DPP4), was identified by FCCS as binding insulin-expressing cells which are Nkx6.1 negative. Depletion of CD26+ cells followed by enrichment for CD49A+ cells increased insulin+/Nkx6.1+ cells fraction to ~70%. The CD26-/CD49A+ enriched ILC exhibited improved function over non-sorted ILC or CD49A+ cells in diabetic mice and maintain prolonged blood C-peptide levels.ConclusionsRefining the composition of ILC differentiated from hPSC by negative selection to remove cells expressing CD26 and positive selection for CD49A expressing cells could enable more effective cell therapy of diabetes.

Cyclin-dependent kinase-like-5 (CDKL5) deficiency disorder (CDD) is a severe X-linked neurodegenerative disease characterised by early-onset epileptic seizures, low muscle tone, progressive intellectual disability and severe motor function. CDD affects ∼1 in 60,000 live births, with many patients experiencing a reduced quality of life due to the severity of their neurological symptoms and functional impairment. There are no effective therapies for CDD, with current treatments focusing on improving symptoms rather than addressing the underlying causes of the disorder. Zebrafish offer many unique advantages for high-throughput preclinical evaluation of potential therapies for neurological diseases, including CDD. In particular, the large number of offspring produced, together with the possibilities for in vivo imaging and genetic manipulation, allows for the detailed assessment of disease pathogenesis and therapeutic discovery. We have characterised a loss-of-function zebrafish model for CDD, containing a nonsense mutation in cdkl5. cdkl5 mutant zebrafish display defects in neuronal patterning, seizures, microcephaly, and reduced muscle function caused by impaired muscle innervation. This study provides a powerful vertebrate model for investigating CDD disease pathophysiology and allowing high-throughput screening for effective therapies. This article has an associated First Person interview with the first author of the paper.

How diverse adult stem and progenitor populations regenerate tissue following damage to the brain is poorly understood. In highly regenerative vertebrates, such as zebrafish, radial-glia (RG) and neuro-epithelial-like (NE) stem/progenitor cells contribute to neuronal repair after injury. However, not all RG act as neural stem/progenitor cells during homeostasis in the zebrafish brain, questioning the role of quiescent RG (qRG) post-injury. To understand the function of qRG during regeneration, we performed a stab lesion in the adult midbrain tectum to target a population of homeostatic qRG, and investigated their proliferative behaviour, differentiation potential, and Wnt/β-catenin signalling. EdU-labelling showed a small number of proliferating qRG after injury (pRG) but that progeny are restricted to RG. However, injury promoted proliferation of NE progenitors in the internal tectal marginal zone (TMZi) resulting in amplified regenerative neurogenesis. Increased Wnt/β-catenin signalling was detected in TMZi after injury whereas homeostatic levels of Wnt/β-catenin signalling persisted in qRG/pRG. Attenuation of Wnt signalling suggested that the proliferative response post-injury was Wnt/β-catenin-independent. Our results demonstrate that qRG in the tectum have restricted capability in neuronal repair, highlighting that RG have diverse functions in the zebrafish brain. Furthermore, these findings suggest that endogenous stem cell compartments compensate lost tissue by amplifying homeostatic growth.

Zebrafish larvae are suitable in vivo models for toxicological and pharmacological screens due to their transparency, small size, ex utero development, and genetic and physiological similarity to humans. Using modern imaging techniques, cells and tissues can be dynamically visualized over several days in multiple zebrafish larvae. However, precise specimen immobilization and maintenance of homeostatic conditions remain a challenge for longitudinal studies. A highly customizable mounting configuration with inbuilt means of controlling temperature and media flow would therefore be a valuable tool to facilitate long-term imaging of a large number of specimens. Using three-dimensional printing, we have developed a millifluidic, modular homeostatic imaging plate (HIP), which consists of a customizable sample insert and a temperature-controlled incubation chamber that is continuously perfused, providing an ideal environment for long-term experiments where homeostatic conditions are desired. The HIP is cheap to produce, has a standard microtiter well plate format, and can be fitted to most microscopes. We used the device to image dynamic regeneration of spinal cord neurons. The flexibility and adaptability of the HIP facilitate long-term in vivo imaging of many samples, and can be easily adapted to suit a broad range of specimens.

Mucopolysaccharidosis IIIA (MPS IIIA, Sanfilippo syndrome type A), a paediatric neurological lysosomal storage disease, is caused by impaired function of the enzyme N-sulfoglucosamine sulfohydrolase (SGSH) resulting in impaired catabolism of heparan sulfate glycosaminoglycan (HS GAG) and its accumulation in tissues. MPS IIIA represents a significant proportion of childhood dementias. This condition generally leads to patient death in the teenage years, yet no effective therapy exists for MPS IIIA and a complete understanding of the mechanisms of MPS IIIA pathogenesis is lacking. Here, we employ targeted CRISPR/Cas9 mutagenesis to generate a model of MPS IIIA in the zebrafish, a model organism with strong genetic tractability and amenity for high-throughput screening. The sgshΔex5−6 zebrafish mutant exhibits a complete absence of Sgsh enzymatic activity, leading to progressive accumulation of HS degradation products with age. sgshΔex5−6 zebrafish faithfully recapitulate diverse CNS-specific features of MPS IIIA, including neuronal lysosomal overabundance, complex behavioural phenotypes, and profound, lifelong neuroinflammation. We further demonstrate that neuroinflammation in sgshΔex5−6 zebrafish is largely dependent on interleukin-1β and can be attenuated via the pharmacological inhibition of Caspase-1, which partially rescues behavioural abnormalities in sgshΔex5−6 mutant larvae in a context-dependent manner. We expect the sgshΔex5−6 zebrafish mutant to be a valuable resource in gaining a better understanding of MPS IIIA pathobiology towards the development of timely and effective therapeutic interventions.

The Zebrafish Embryo Genotyper (ZEG) device provides a promising tool for genotyping live embryos. Although the gross morphology and survival of embryos after the use of ZEG are unaffected, the cellular and molecular effects of the ZEG protocol remain unknown. To address this, we have examined the integrity of specific tissues, and evaluated the expression of stress-responsive genes to determine the impact of the ZEG protocol. Our analyses reveal that although ZEG results in a low-level acute stress response, no long-lasting effects are evident, supporting its utilization for a variety of downstream assays.

Alon M Salby,1 Simon E Skalicky1,2 1Department of Ophthalmology, Royal Melbourne Hospital, Melbourne, VIC, Australia; 2Glaucoma Unit, Royal Victorian Eye and Ear Hospital, Melbourne, VIC, AustraliaCorrespondence: Alon M Salby Email amicahmd@gmail.comPurpose: This retrospective audit aimed to evaluate the impact of combined iStent® Inject (iSI) and phacoemulsification on medication number in Australians with open-angle glaucoma. Secondary outcomes included intraocular pressure (IOP), best-corrected visual acuity, refraction and visual fields.Patients and Methods: Patients with glaucoma that received combined iSI and phacoemulsification by the same surgeon between 1 February 2016 and 1 February 2018 were audited for postoperative medication number, pressures after 1 day, 1 week, 4 weeks and 6, 12, 18 and 24 months, visual acuity, refraction and visual fields. These parameters were compared to baseline levels and with those from a separate cohort of patients without glaucoma that received standalone phacoemulsification.Results: Forty-one patients (63 eyes) received the combined procedure. Thirty-four patients (59 eyes) received standalone phacoemulsification. Four weeks after receiving combined iSI and phacoemulsification the mean medication number was significantly reduced by 1.3 (p < 0.001) for those on medication at baseline and by 0.5 (p = 0.002) overall. Mean IOP was significantly reduced from baseline after 6 months (– 16%; p = 0.012; n = 35) and 12 months (– 29%; p = 0.004; n = 16). Patients receiving standalone phacoemulsification had short-term reductions in IOP at 4 weeks (– 8%; p < 0.001; n = 57) and 6 months (– 16%; p < 0.001; n = 32). These patients without glaucoma had lower pressures overall compared to those with glaucoma that received the combined procedure (p = 0.019). There were no differences in final visual acuity or refractive outcomes between groups.Conclusion: This audit suggests that iSI and phacoemulsification are at least as effective in controlling IOP as medical therapy. It may have an important role in reducing the medication burden in Australians with cataract and glaucoma. This study is one of the first to confirm refractive stability in concomitant iSI and phacoemulsification.Keywords: glaucoma, iStent Inject, micro-invasive glaucoma surgery, trabecular micro-bypass, phacoemulsification

An innovative drug delivery vehicle based on polynorepinephrine (PNE) with controllable size modification, high delivery efficacy and low cytotoxicity is presented. Highly monodisperse PNE nanoparticles are fabricated by the autoxidation of norepinephrine monomers in an alkaline water/ethanol mixture via stirring at room temperature. We demonstrated the facile optimization of particle size to enhance particle stability and biocompatibility by varying solvent and monomer dosage. To demonstrate the suitability and potential application of PNE particles in cancer therapy, we show that these particles are biocompatible in vitro with HeLa cells and in vivo in zebrafish embryos. After loading the anti-cancer chemotherapy drug doxorubicin (DOX) into the PNE nanoparticles, a consistent and pH responsive drug release profile of DOX was achieved in different environmental conditions. It was found that DOX loaded PNE nanoparticles (PNE/DOX) exhibit much higher pharmaceutical cytotoxicity than free DOX on HeLa cells. Furthermore, the amount of drug released was significantly enhanced in acidic environments that mimic the pH of extracellular tumour microenvironments. Taken together, the PNE nanoparticles represent a new class of melanin particles with promising potential in drug delivery and as a therapeutic platform for cancer treatment.

Purpose This retrospective audit aimed to evaluate the impact of combined iStent® Inject (iSI) and phacoemulsification on medication number in Australians with open-angle glaucoma. Secondary outcomes included intraocular pressure (IOP), best-corrected visual acuity, refraction and visual fields. Patients and Methods Patients with glaucoma that received combined iSI and phacoemulsification by the same surgeon between 1 February 2016 and 1 February 2018 were audited for postoperative medication number, pressures after 1 day, 1 week, 4 weeks and 6, 12, 18 and 24 months, visual acuity, refraction and visual fields. These parameters were compared to baseline levels and with those from a separate cohort of patients without glaucoma that received standalone phacoemulsification. Results Forty-one patients (63 eyes) received the combined procedure. Thirty-four patients (59 eyes) received standalone phacoemulsification. Four weeks after receiving combined iSI and phacoemulsification the mean medication number was significantly reduced by 1.3 (p < 0.001) for those on medication at baseline and by 0.5 (p = 0.002) overall. Mean IOP was significantly reduced from baseline after 6 months (–16%; p = 0.012; n = 35) and 12 months (–29%; p = 0.004; n = 16). Patients receiving standalone phacoemulsification had short-term reductions in IOP at 4 weeks (–8%; p < 0.001; n = 57) and 6 months (–16%; p < 0.001; n = 32). These patients without glaucoma had lower pressures overall compared to those with glaucoma that received the combined procedure (p = 0.019). There were no differences in final visual acuity or refractive outcomes between groups. Conclusion This audit suggests that iSI and phacoemulsification are at least as effective in controlling IOP as medical therapy. It may have an important role in reducing the medication burden in Australians with cataract and glaucoma. This study is one of the first to confirm refractive stability in concomitant iSI and phacoemulsification.

Tissue regeneration and functional restoration after injury are considered as stem- and progenitor-cell-driven processes. In the central nervous system, stem cell-driven repair is slow and problematic because function needs to be restored rapidly for vital tasks. In highly regenerative vertebrates, such as zebrafish, functional recovery is rapid, suggesting a capability for fast cell production and functional integration. Surprisingly, we found that migration of dormant "precursor neurons" to the injury site pioneers functional circuit regeneration after spinal cord injury and controls the subsequent stem-cell-driven repair response. Thus, the precursor neurons make do before the stem cells make new. Furthermore, RNA released from the dying or damaged cells at the site of injury acts as a signal to attract precursor neurons for repair. Taken together, our data demonstrate an unanticipated role of neuronal migration and RNA as drivers of neural repair.

Background: The molecular changes involved in Alzheimer’s disease (AD) progression remain unclear since we cannot easily access antemortem human brains. Some non-mammalian vertebrates such as the zebrafish preserve AD-relevant transcript isoforms of the PRESENILIN genes lost from mice and rats. One example is PS2V, the alternative transcript isoform of the PSEN2 gene. PS2V is induced by hypoxia/oxidative stress and shows increased expression in late onset, sporadic AD brains. A unique, early onset familial AD mutation of PSEN2, K115fs, mimics the PS2V coding sequence suggesting that forced, early expression of PS2V-like isoforms may contribute to AD pathogenesis. Here we use zebra fish to model the K115fs mutation to investigate the effects of forced PS2V-like expression on the transcriptomes of young adult and aged adult brains. Methods: We edited the zebra fish genome to model the K115fs mutation. To explore its effects at the molecular level, we analysed the brain transcriptome and proteome of young (6-month-old) and aged (24-month-old) wild type and heterozygous mutant female sibling zebra fish. Finally, we used gene co-expression network analysis (WGCNA) to compare molecular changes in the brains of these fish to human AD. Results: Young heterozygous mutant fish show transcriptional changes suggesting accelerated brain aging and increased glucocorticoid signalling. These early changes precede a transcriptional ‘inversion’ that leads to glucocorticoid resistance and other likely pathological changes in aged heterozygous mutant fish. Notably, microglia-associated immune responses regulated by the ETS transcription factor family are altered in both our zebra fish mutant model and in human AD. The molecular changes we observe in aged heterozygous mutant fish occur without obvious histopathology and possibly in the absence of Aβ. Conclusions: Our results suggest that forced expression of a PS2V-like isoform contributes to immune and stress responses favouring AD pathogenesis. This highlights the value of our zebra fish genetic model for exploring molecular mechanisms involved in AD pathogenesis. Refereed/Peer-reviewed

Human amyloids and plaques uncovered post mortem are highly heterogeneous in structure and composition, yet literature concerning the hetero-aggregation of amyloid proteins is extremely scarce. This knowledge deficiency is further exacerbated by the fact that peptide delivery is a major therapeutic strategy for targeting their full-length counterparts associated with the pathologies of a range of human diseases, including dementia and type 2 diabetes (T2D). Accordingly, here we examined the co-aggregation of full-length human islet amyloid polypeptide (IAPP), a peptide associated with type 2 diabetes, with its primary and secondary amyloidogenic fragments 19–29 S20G and 8–20. Single-molecular aggregation dynamics was obtained by high-speed atomic force microscopy, augmented by transmission electron microscopy, X-ray diffraction and super-resolution stimulated emission depletion microscopy. The co-aggregation significantly prolonged the pause phase of fibril elongation, increasing its dwell time by threefold. Surprisingly, unidirectional elongation of mature fibrils, instead of protofilaments, was observed for the co-aggregation, indicating a new form of tertiary protein aggregation unknown to existing theoretical models. Further in vivo zebrafish embryonic assay indicated improved survival and hatching, as well as decreased frequency and severity of developmental abnormalities for embryos treated with the hetero-aggregates of IAPP with 19–29 S20G, but not with 8–20, compared to the control, indicating the therapeutic potential of 19–29 S20G against T2D.

The environmental impact of exposure to 3D-printed plastics as well as potential migration of toxic chemicals from 3D-printed plastics remains largely unexplored. In this work we applied leachates from plastics fabricated using a stereolithography (SLA) process to early developmental stages of zebrafish (Danio rerio) to investigate developmental toxicity and neurotoxicity. Migration of unpolymerized photoinitiator, 1-hydroxycyclohexyl phenyl ketone (1−HCHPK) from a plastic solid phase to aqueous media at up to 200 mg/L in the first 24 h was detected using gas chromatography–mass spectrometry. Both plastic extracts (LC50 22.25% v/v) and 1−HCHPK (LC50 60 mg/L) induced mortality and teratogenicity within 48 h of exposure. Developmental toxicity correlated with in situ generation of reactive oxygen species (ROS), an increase in lipid peroxidation and protein carbonylation markers and enhanced activity of superoxide dismutase (SOD) and glutathione-S-transferase (GST) in embryos exposed to concentrations as low as 20% v/v for plastic extracts and 16 mg/L for 1−HCHPK. ROS-induced cellular damage led to induction of caspase-dependent apoptosis which could be pharmacologically inhibited with both antioxidant ascorbic acid and a pan-caspase inhibitor. Neuro-behavioral analysis showed that exposure to plastic leachates reduced spontaneous embryonic movement in 24–36 hpf embryos. Plastic extracts in concentrations above 20% v/v induced rapid retardation of locomotion, changes in photomotor response and habituation to photic stimuli with progressive paralysis in 120 hpf larvae. Significantly decreased acetylcholinesterase (AChE) activity with lack of any CNS-specific apoptotic phenotypes as well as lack of changes in motor neuron density, axonal growth, muscle segment integrity or presence of myoseptal defects were detected upon exposure to plastic extracts during embryogenesis. Considering implications of the results for environmental risk assessment and the growing usage of 3D-printing technologies, we speculate that some 3D-printed plastic waste may represent a significant and yet very poorly uncharacterized environmental hazard that merits further investigation on a range of aquatic and terrestrial species.