RESUMEN
The dsRNA-activated protein kinase R (PKR) is one of key antiviral effectors induced by interferons (IFNs), and its functions are largely unknown in tilapia, an important commercial fish species suffering from several viral infectious diseases. In the present study, a PKR gene named On-PKR was identified and cloned from Nile tilapia, Oreochromis niloticus. On-PKR gene was constitutively expressed in all tissues examined, with the highest expression level observed in head kidney and liver, and was rapidly induced in all organs/tissues tested following the stimulation of poly(I:C). Importantly, the expression of On-PKR is induced by group I and group II IFNs with distinct induction kinetics in vivo: group I IFN elicits a relative delayed but sustained induction of On-PKR, whereas group II IFN triggers a rapid and transient expression of On-PKR. Moreover, the overexpression of On-PKR has been proven to inhibit the protein translation and virus replication in fish cells. The present study thus contributes to a better understanding of the functions of antiviral effectors in tilapia, and may provide clues for the prevention and therapy of viral diseases in fish.
Asunto(s)
Cíclidos/genética , Cíclidos/inmunología , Enfermedades de los Peces/inmunología , Regulación de la Expresión Génica/inmunología , eIF-2 Quinasa/genética , eIF-2 Quinasa/inmunología , Secuencia de Aminoácidos , Animales , Enfermedades de los Peces/virología , Proteínas de Peces/química , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Perfilación de la Expresión Génica/veterinaria , Inmunidad/genética , Filogenia , Poli I-C/farmacología , Reoviridae/fisiología , Infecciones por Reoviridae/inmunología , Infecciones por Reoviridae/veterinaria , Infecciones por Reoviridae/virología , Alineación de Secuencia/veterinaria , eIF-2 Quinasa/químicaRESUMEN
Interferons are a family of class II α-helical cytokines playing vital roles in antiviral immune response, and little information is available to date regarding the interferon system of tilapia. In this study, a type I IFN gene, named On-IFNc, was identified in Nile tilapia, Oreochromis niloticus. The predicted protein of On-IFNc contains several structural features known in type I IFNs, and On-IFNc was clustered together with the known IFNc in fish into a separated clade in the phylogenetic tree. On-IFNc gene was constitutively expressed in all tissues examined, with the highest expression level observed in liver, and was rapidly induced in all organs/tissues tested following the stimulation of poly(I:C). In addition, recombinant On-IFNc has been proven to markedly induce the expression of the antiviral effectors, Mx and viperin, the signalling components, STAT1, STAT2, and IRF9, and the transcription factors, IRF3 and IRF7, as well as the tyrosine phosphorylation of STAT1 and STAT2 in fish cells. Furthermore, recombinant On-IFNc has been proven to possess antiviral activity against ISKNV. The present study thus contributes to a better understanding of the functional properties of the type I IFN system in tilapia.
Asunto(s)
Cíclidos/genética , Cíclidos/inmunología , Enfermedades de los Peces/inmunología , Regulación de la Expresión Génica/inmunología , Inmunidad Innata/genética , Interferón gamma/genética , Interferón gamma/inmunología , Secuencia de Aminoácidos , Animales , Proteínas de Peces/química , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Perfilación de la Expresión Génica/veterinaria , Interferón gamma/química , Filogenia , Poli I-C/farmacología , Alineación de Secuencia/veterinaria , Transcriptoma/inmunologíaRESUMEN
Type I interferons are a subset of cytokines playing central roles in host antiviral defense, and their effects depend on the interaction with the heterodimeric receptor complex. Surprisingly, two pairs of the receptor subunits, CRFB1 and CRFB5, and CRFB2 and CRFB5, have been identified in fish, but the studies about preferential receptor usage of different fish IFN subtypes are rather limited. In this study, the three receptor chains of type I IFNs named as On-CRFB1, On-CRFB2 and On-CRFB5 were identified in Nile tilapia, Oreochromis niloticus. These three genes were constitutively expressed in all tissues examined, with the highest expression level observed in muscle and liver, and were rapidly induced in liver following the stimulation of poly(I:C). Interestingly, it is possible that all three subtypes of tilapia IFNs are able to signal through two pairs of the receptor subunits, On-CRFB1 and On-CRFB5, and On-CRFB2 and On-CRFB5. More importantly, tilapia group I IFNs (On-IFNd and On-IFNh) preferentially signal through a receptor complex composed of On-CRFB1 and On-CRFB5, and group II IFNs (On-IFNc) preferentially signal through a receptor complex comprised of On-CRFB2 and On-CRFB5. The present study thus provides new insights into the receptor usage of group I and group II IFNs in fish.
Asunto(s)
Cíclidos/genética , Cíclidos/inmunología , Regulación de la Expresión Génica/inmunología , Inmunidad Innata/genética , Poli I-C/farmacología , Receptores de Interferón/genética , Receptores de Interferón/inmunología , Secuencia de Aminoácidos , Animales , Proteínas de Peces/química , Proteínas de Peces/genética , Proteínas de Peces/inmunología , Perfilación de la Expresión Génica/veterinaria , Filogenia , Receptores de Interferón/química , Alineación de Secuencia/veterinariaRESUMEN
PGRP-SC2, the member of PGRP family, plays an important role in regulation of innate immune response. In this paper, a PGRP-SC2 gene of Nile tilapia, Oreochromis niloticus (designated as On-PGRP-SC2) was cloned and its expression pattern under the infection of Streptococcus agalactiae was investigated. Sequence analysis showed main structural features required for amidase activity were detected in the deduced amino acid sequence of On-PGRP-SC2. In healthy tilapia, the On-PGRP-SC2 transcripts could be detected in all the examined tissues, with the most abundant expression in the muscle. When infected with S. agalactiae, there was a clear time-dependent expression pattern of On-PGRP-SC2 in the spleen, head kidney and brain. The assays for the amidase activity suggested that recombinant On-PGRP-SC2 protein had a Zn(2+)-dependent PGN-degrading activity. Moreover, our works showed that recombinant On-PGRP-SC2 protein could significantly reduce bacterial load in target organs attacked by S. agalactiae. These findings indicated that On-PGRP-SC2 may play important roles in the immune response to S. agalactiae in Nile tilapia.
Asunto(s)
Proteínas Portadoras/genética , Cíclidos/inmunología , Enfermedades de los Peces/genética , Enfermedades de los Peces/inmunología , Proteínas de Peces/genética , Infecciones Estreptocócicas/veterinaria , Secuencia de Aminoácidos , Animales , Proteínas Portadoras/química , Proteínas Portadoras/metabolismo , Clonación Molecular , ADN Complementario/genética , ADN Complementario/metabolismo , Enfermedades de los Peces/microbiología , Proteínas de Peces/química , Proteínas de Peces/metabolismo , Inmunidad Innata/genética , Conformación Molecular , Especificidad de Órganos , Filogenia , ARN Mensajero/genética , ARN Mensajero/metabolismo , Alineación de Secuencia/veterinaria , Infecciones Estreptocócicas/genética , Infecciones Estreptocócicas/inmunología , Infecciones Estreptocócicas/microbiología , Streptococcus agalactiae/fisiologíaRESUMEN
In recent years, there has been a growing focus on the toxicity and mortality induced by nanoplastics (NPs) in aquatic organisms. However, studies investigating mechanisms underlying oxidative stress (OS), apoptosis, and inflammation induced by NPs in fish remain limited. This study observed that polystyrene NPs (PS-NPs) were accumulated into zebrafish larvae and zebrafish embryonic fibroblast (ZF4 cells), accompanied by the occurrence of pathological damage both at the cellular and tissue-organ level. Additionally, the transcriptional up-regulation of NADPH oxidases (NOXs) and subsequent excessive generation of reactive oxygen species (ROS) resulted in notable changes in the relative mRNA and protein expression levels associated with antioxidant oxidase systems in larvae. Furthermore, the study identified the impact of NPs on mitochondrial ultrastructural, resulting in mitochondrial depolarization and downregulation of mRNA expression related to the electron transport chain due to excessive ROS generation. Short-term exposure to NPs also triggered apoptosis and inflammation in zebrafish larvae, evident from significant up-regulation in mRNA expressions of proapoptotic factors and NF-κB proinflammatory signaling pathway, as well as increased transcription and protein levels of pro-inflammatory factors in larvae. Inhibition of intracellular excessive ROS effectively reduced the induction of apoptosis, NF-κB P65 nuclear migration levels, and cytokine secretion, underscoring OS as a pivotal factor throughout the process of apoptosis and inflammatory responses induced by NPs. This research significantly advances our comprehension of biological effects and underlying mechanisms of NPs in freshwater fish.
Asunto(s)
Apoptosis , Inflamación , Larva , Poliestirenos , Especies Reactivas de Oxígeno , Contaminantes Químicos del Agua , Pez Cebra , Animales , Especies Reactivas de Oxígeno/metabolismo , Apoptosis/efectos de los fármacos , Poliestirenos/toxicidad , Inflamación/inducido químicamente , Larva/efectos de los fármacos , Contaminantes Químicos del Agua/toxicidad , Estrés Oxidativo/efectos de los fármacos , Nanopartículas/toxicidadRESUMEN
BACKGROUND AND OBJECTIVE: Acute ischemic stroke (AIS) is a common neurological disorder characterized by the sudden onset of cerebral ischemia, leading to functional impairments. Swift and precise detection of AIS lesions is crucial for stroke diagnosis and treatment but poses a significant challenge. This study aims to leverage multimodal fusion technology to combine complementary information from various modalities, thereby enhancing the detection performance of AIS target detection models. METHODS: In this retrospective study of AIS, we collected data from 316 AIS patients and created a multi-modality magnetic resonance imaging (MRI) dataset. We propose a Multi-Scale Attention-based YOLOv5 (MSA-YOLOv5), targeting challenges such as small lesion size and blurred borders at low resolutions. Specifically, we augment YOLOv5 with a prediction head to detect objects at various scales. Next, we replace the original prediction head with a Multi-Scale Swin Transformer Prediction Head (MS-STPH), which reduces computational complexity to linear levels and enhances the ability to detect small lesions. We incorporate a Second-Order channel attention (SOCA) module to adaptively rescale channel features by employing second-order feature statistics for more discriminative representations. Finally, we further validate the effectiveness of our method using the ISLES 2022 dataset. RESULTS: On our in-house AIS dataset, MSA-YOLOv5 achieves a 79.0% mAP0.5, substantially surpassing other single-stage models. Compared to two-stage models, it maintains a comparable performance level while significantly reducing the number of parameters and resolution. On the ISLES 2022 dataset, MSA-YOLOv5 attains an 80.0% mAP0.5, outperforming other network models by a considerable margin. MS-STPH and SOCA modules can significantly increase mAP0.5 by 2.7% and 1.9%, respectively. Visualization interpretability results show that the proposed MSA-YOLOv5 restricts high attention in the small regions of AIS lesions. CONCLUSIONS: The proposed MSA-YOLOv5 is capable of automatically and effectively detecting acute ischemic stroke lesions in multimodal images, particularly for small lesions and artifacts. Our enhanced model reduces the number of parameters while improving detection accuracy. This model can potentially assist radiologists in providing more accurate diagnosis, and enable clinicians to develop better treatment plans.
Asunto(s)
Accidente Cerebrovascular Isquémico , Accidente Cerebrovascular , Humanos , Accidente Cerebrovascular Isquémico/diagnóstico por imagen , Estudios Retrospectivos , Imagen por Resonancia Magnética , Accidente Cerebrovascular/diagnóstico por imagen , ArtefactosRESUMEN
BACKGROUND AND OBJECTIVE: Stroke is the second most deadly disease globally and seriously endangers people's lives and health. The automatic detection of stroke lesions from diffusion-weighted imaging (DWI) can improve the diagnosis. Recently, automatic detection methods based on YOLOv5 have been utilized in medical images. However, most of them barely capture the stroke lesions because of their small size and fuzzy boundaries. METHODS: To address this problem, a novel method for tracing the edge of the stroke lesion based on YOLOv5 (TE-YOLOv5) is proposed. Specifically, we constantly update the high-level features of the lesion using an aggregate pool (AP) module. Conversely, we feed the extracted feature into the reverse attention (RA) module to trace the edge relationship promptly. Overall, 1681 DWI images of 319 stroke patients have been collected, and experienced radiologists have marked the lesions. DWI images were randomly split into the training and test set at a ratio of 8:2. TE-YOLOv5 has been compared with the related models, and a detailed ablation analysis has been conducted to clarify the role of the RA and AP modules. RESULTS: TE-YOLOv5 outperforms its counterparts and achieves competitive performance with a precision of 81.5%, a recall of 75.8%, and a mAP@0.5 of 80.7% (mean average precision while the intersection over union is 0.5) under the same backbone. At the patient level, the positive finding rate can reach 98.51%, while the confidence is set at 80.0%. After ablating RA, the mAP@0.5 decreases to 79.6%; after ablating RA and AP, the mAP@0.5 decreases to 78.1%. CONCLUSIONS: The proposed TE-YOLOv5 can automatically and effectively detect stroke lesions from DWI images, especially for those with an extremely small size and blurred boundaries. AP and RA modules can aggregate multi-layer high-level features and concurrently track the edge relationship of stroke lesions. These detection methods might help radiologists improve stroke diagnosis and have great application potential in clinical practice.
Asunto(s)
Accidente Cerebrovascular , Humanos , Accidente Cerebrovascular/diagnóstico por imagen , Imagen de Difusión por Resonancia Magnética/métodosRESUMEN
The reduction of disulfide bonds of exogenous antigens is crucial to the MHC-II class antigen processing and presenting pathway and is catalysed by interferon-γ-inducible lysosomal thiol reductase (GILT). In this study, a reptile GILT gene from Chinese soft-shelled turtle, Pelodiscus sinensis (PsGILT), was identified. The full-length cDNA of PsGILT is 1631 nucleotides (nt), including a 5'-untranslated region (UTR) of 3â¯nt, a 3'-UTR of 860â¯nt and an open reading frame (ORF) of 768â¯nt encoding 255 amino acids (aa). The conserved features in known GILTs, such as signal peptide, CXXC motif, GILT signature sequence, N-glycosylation site and conserved cysteines, were all found in the putative PsGILT protein. Genomic analysis revealed that PsGILT kept the "7 exons and 6 introns" structure of vertebrate GILT genes. PsGILT was expressed in all examined organs/tissues and was mainly expressed in spleen and blood. Increased mRNA expression levels of PsIFN-γ and PsGILT in PBLs were observed after induction with LPS, PolyI:C and recombinant IFN-γ (rIFN-γ). We also tested the reductase activity of rGILT in vitro and found that it could reduce intact human IgG into H chains and L chains. These above results implied that PsGILT may play an important role in resisting bacterial and viral infections, like other vertebrate GILTs.
Asunto(s)
Leucocitos Mononucleares/fisiología , Oxidorreductasas actuantes sobre Donantes de Grupos Sulfuro/genética , Proteínas de Reptiles/genética , Bazo/fisiología , Tortugas/inmunología , Animales , Antivirales/metabolismo , Células Cultivadas , Clonación Molecular , Humanos , Inmunidad Innata , Oxidorreductasas actuantes sobre Donantes de Grupos Sulfuro/metabolismo , Filogenia , Poli I-C/inmunología , Proteínas de Reptiles/metabolismo , TranscriptomaRESUMEN
In the present study, interleukin-22 (IL-22) from So-iny mullet (Liza haematocheila) was identified, and its tissue expression in both healthy and Streptococcus dysgalactiae-infected fish was examined. The full length cDNA sequence of mullet IL-22 was 1070bp, containing an open reading frame of 555bp. The deduced amino acid sequence shared high similarity (45.1-67.9%) with IL-22 from other fish species. Mullet IL-22 also contained an IL-10 family signature and four cysteine residues that were well conserved in other vertebrate IL-22 molecules. Mullet IL-22 mRNA was highly expressed in kidney, moderately expressed in liver and gut, and relatively weakly expressed in spleen, and its expression was significantly up-regulated in all the examined tissues following S. dysgalactiae infection. Furthermore, recombinant mullet IL-22 protein was shown to promote the expression of ß-defensin in the four tissues and to increase the survival rate of the fish infected with S. dysgalactiae. Our results suggest mullet IL-22 plays an important role in the immune defense against bacterial infection and has the potential to be used to treat bacterial diseases in fish.