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1.
Malar J ; 18(1): 356, 2019 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-31703583

RESUMO

BACKGROUND: Control and elimination of malaria can be accelerated by transmission-blocking interventions such as vaccines. A surface antigen of Plasmodium falciparum gametocytes, Pfs230, is a leading vaccine target antigen, and has recently progressed to experimental clinical trials. To support vaccine product development, an N-terminal Pfs230 antigen was designed to increase yield, as well as to improve antigen quality, integrity, and homogeneity. METHODS: A scalable baculovirus expression system was used to express the Pfs230D1+ construct (aa 552-731), which was subsequently purified and analysed. Pfs230D1+ was designed to avoid glycosylation and protease digestion, thereby potentially increasing homogeneity and stability. The resulting Pfs230D1+ protein was compared to a previous iteration of the Pfs230 N-terminal domain, Pfs230C1 (aa 443-731), through physiochemical characterization and in vivo analysis. The induction of functional antibody responses was confirmed via the standard membrane feeding assay (SMFA). RESULTS: Pfs230D1+ was produced and purified to an overall yield of 23 mg/L culture supernatant, a twofold yield increase over Pfs230C1. The Pfs230D1+ protein migrated as a single band via SDS-PAGE and was detected by anti-Pfs230C1 monoclonal antibodies. Evaluation by SDS-PAGE, chromatography (size-exclusion and reversed phase) and capillary isoelectric focusing demonstrated the molecule had improved homogeneity in terms of size, conformation, and charge. Intact mass spectrometry confirmed its molecular weight and that it was free of glycosylation, a key difference to the prior Pfs230C1 protein. The correct formation of the two intramolecular disulfide bonds was initially inferred by binding of a conformation specific monoclonal antibody and directly confirmed by LC/MS and peptide mapping. When injected into mice the Pfs230D1+ protein elicited antibodies that demonstrated transmission-reducing activity, via SMFA, comparable to Pfs230C1. CONCLUSION: By elimination of an O-glycosylation site, a potential N-glycosylation site, and two proteolytic cleavage sites, an improved N-terminal Pfs230 fragment was produced, termed D1+, which is non-glycosylated, homogeneous, and biologically active. An intact protein at higher yield than that previously observed for the Pfs230C1 fragment was achieved. The results indicate that Pfs230D1+ protein produced in the baculovirus expression system is an attractive antigen for transmission-blocking vaccine development.


Assuntos
Antígenos de Protozoários/genética , Expressão Gênica/imunologia , Vacinas Antimaláricas/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/genética , Animais , Antígenos de Protozoários/imunologia , Camundongos , Proteínas de Protozoários/imunologia
2.
BMC Infect Dis ; 19(1): 999, 2019 Nov 27.
Artigo em Inglês | MEDLINE | ID: mdl-31775660

RESUMO

BACKGROUND: Recent studies have shown that CD103 is an important marker for tissue-resident memory T cells (TRM) which plays an important role in anti-infection. However, the role of CD103+ TRM was not elucidated in the progress of S. japonicum infection induced disease. METHODS: 6-8 weeks old C57BL/6 mice were infected by S. japonicum. Mice were sacrificed and the lungs were removed 5-6 weeks after infection. Immunofluorescent staining and Q-PCR were performed to identify the expression of CD103 molecule. Single cellular populations were made, percentages of CD103 on both CD4+ and CD8+ T lymphocytes were dynamical observed by flow cytometry (FCM). Moreover, the expression of memory T cells related molecules CD69 and CD62L, T cell function associated molecules CD107a, IFN-γ, IL-4, IL-9, and IL-10 were compared between CD103+ CD4+ and CD8+ T cells by FCM. RESULTS: CD103+ cells were emerged in the lung of both naive and S. japonicum infected mice. Both the percentage and the absolute numbers of pulmonary CD4+ and CD8+ cells were increased after S. japonicum infection (P < 0.05). The percentage of CD103+ cells in CD8+ T cells decreased significantly at the early stage of S. japonicum infection (P < 0.05). Increased CD69, decreased CD62L and CD107a expressions were detected on both CD4+ and CD8+ CD103+ T cells in the lungs of infected mice (P < 0.05). Compared to CD8+ CD103+ T cells, CD4+ CD103+ T cells from infected mice expressed higher level of CD69 and lower level CD62L molecules (P < 0.05). Moreover, higher percentage of IL-4+, IL-9+ and IL-10+ cells on CD4+ CD103+ pulmonary T cells was found in infected mice (P < 0.05). Significantly increased IL-4 and IL-9, and decreased IFN-γ expressing cells were detected in CD8+CD103+ cells of infected mice (P < 0.05). CONCLUSIONS: CD103-expressing pulmonary CD4+ and CD8+ T cells play important roles in mediating S. japonicum infection induced granulomatous inflammation in the lung.


Assuntos
Antígenos CD/genética , Antígenos CD/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Cadeias alfa de Integrinas/genética , Cadeias alfa de Integrinas/metabolismo , Schistosoma japonicum , Esquistossomose Japônica/metabolismo , Animais , Biomarcadores/metabolismo , Citocinas/metabolismo , Feminino , Expressão Gênica/imunologia , Memória Imunológica , Pulmão/metabolismo , Pulmão/parasitologia , Camundongos , Camundongos Endogâmicos C57BL , Reação em Cadeia da Polimerase em Tempo Real , Esquistossomose Japônica/microbiologia
3.
Fish Shellfish Immunol ; 94: 769-779, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31580935

RESUMO

Although viruses represent a major threat for cultured fish worldwide, the commercialization of vaccines capable of providing effective and long-lasting protection is still lacking for most of these viral diseases. In this situation, the use of supplemented diets could be a suitable strategy to increase the immune status of the fish and reduce the impact of viral pathogens. Among possible immunostimulants that could be included in these functional feeds, some studies have previously shown that certain ß-glucans can significantly increase certain immune parameters of fish and reduce the impact of viral diseases. However, the mechanisms through which ß-glucans exert their activity have not been fully elucidated yet. In the current study, we have studied the immune response of different tissues to viral haemorrhagic septicaemia virus (VHSV) in rainbow trout fed with a non-supplemented control diet as well as in fish fed a commercial functional aquafeed (Protec™, Skretting) containing ß-glucans, vitamin C, vitamin E and zinc. For this, after 30 days of feeding the fish with one of the two diets, they were subsequently infected with VHSV by bath or mock-infected. After 2 or 6 days post-infection, fish were sacrificed and the levels of transcription of different immune genes such as IgM, IgT, IgD, Mx, interferon γ (IFN γ) and perforin studied in different tissues (kidney, gut and gills). Additionally, the levels of natural IgMs in serum were also determined. Our results demonstrate that fish fed the functional diet were capable of mounting an increased IgM, IgT, IgD and Mx transcriptional response to the virus. Additionally, these fish also showed increased levels of natural IgMs in serum. These results reveal a previously undescribed effect of functional diets on fish Ig production and point to Protec™ as an adequate diet to be incorporated in holistic programs aimed at mitigating the effect of viral diseases.


Assuntos
Proteínas de Peixes/genética , Expressão Gênica/imunologia , Septicemia Hemorrágica Viral/imunologia , Novirhabdovirus/fisiologia , Oncorhynchus mykiss/imunologia , Transcrição Genética/imunologia , Ração Animal/análise , Animais , Ácido Ascórbico/administração & dosagem , Ácido Ascórbico/metabolismo , Dieta/veterinária , Suplementos Nutricionais/análise , Proteínas de Peixes/metabolismo , Glucanos/administração & dosagem , Glucanos/metabolismo , Septicemia Hemorrágica Viral/genética , Vitamina E/administração & dosagem , Vitamina E/metabolismo , Vitaminas/administração & dosagem , Vitaminas/metabolismo , Zinco/administração & dosagem , Zinco/metabolismo
4.
Cytogenet Genome Res ; 159(2): 55-65, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31630146

RESUMO

Klinefelter syndrome (KS) is one of the most common congenital disorders of male infertility. Given its high heterogeneity in clinical and genetic presentation, the relationship between transcriptome, clinical phenotype, and associated co-morbidities seen in KS has not been fully clarified. Here, we report a 47,XXY Chinese male with infertility and analyzed the differences in gene expression patterns of peripheral blood mononuclear cells (PBMCs) with regard to a Chinese male and a female control with normal karyotype by single-cell sequencing. A total of 24,439 cells were analyzed and divided into 5 immune cell types (including B cells, T cells, macrophage cells, dendritic cells, and natural killer cells) according to marker genes. Using unsupervised dimensionality reduction and clustering algorithms, we identified molecularly distinct subpopulations of cells between the KS patient and both controls. Gene ontology enrichment analyses yielded terms associated with well-known comorbidities seen in KS as well as an affected immune system and type I diabetes mellitus. Based on our data, we identified several candidate genes which may be implicated in regulating the phenotype of KS. Overall, this analysis provides a comprehensive map of the cell types of PBMCs in a KS patient at the single-cell level, which will contribute to the prevention of comorbidity and improvement of the life quality of KS patients.


Assuntos
Síndrome de Klinefelter/genética , Feminino , Expressão Gênica/genética , Expressão Gênica/imunologia , Genótipo , Humanos , Sistema Imunitário/imunologia , Infertilidade Masculina/genética , Infertilidade Masculina/imunologia , Síndrome de Klinefelter/imunologia , Leucócitos Mononucleares/imunologia , Leucócitos Mononucleares/fisiologia , Masculino , Fenótipo , Análise de Célula Única/métodos , Transcriptoma/genética , Transcriptoma/imunologia
5.
Fish Shellfish Immunol ; 94: 548-557, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31539573

RESUMO

A 56-day growth trial was conducted to investigate the effects of dietary yeast hydrolysate on the growth performance, antioxidation, immune response and resistance against Aeromonas hydrophila in largemouth bass. Four experimental diets were prepared with yeast hydrolysate levels of 0% (Y0), 1.5% (Y1.5), 3.0% (Y3.0) and 4.5% (Y4.5). Each diet was randomly assigned to triplicate 150-L tanks and each tank was stocked with 30 largemouth bass (initial body weight, IBW = 7.71 ±â€¯0.02 g). A challenge test was carried out after the feeding trial by injecting A. hydrophila intraperitoneally for 4-day observation. The results showed that the FBW and WGR in Y1.5 group were significantly higher than those in Y0 group (P < 0.05) and the feed conversion ratio (FCR) got the lowest value in Y1.5 group. And the hydrolysate supplement significantly increased the 4-day cumulative survival rate after the bacterial challenge (P < 0.05). The plasma malondialdehyde was lower in the yeast hydrolysate supplement groups in both pre- and post-challenge test (P < 0.05), while the plasma C3 increased (P < 0.05). In post-challenge test, the plasma superoxide dismutase (SOD) and catalase (CAT) activities increased in the Y1.5 and Y3.0 groups respectively (P < 0.05), and plasma lysozyme in Y1.5 group and the plasma IgM in Y3.0 group were higher than those in others respectively (P < 0.05). For the q-PCR results, in post-challenge test, the hepatic hep2 expression level in Y1.5 and Y4.5 groups were both significantly higher than those in others (P < 0.05), as well as il-8 in Y3.0 group. The spleen hif-1alpha and tgf-beta1 expression levels in Y4.5 group were all significantly lower than those in others (P < 0.05), while the gilt was significantly higher (P < 0.05) in the post-challenge test. And the expression levels of spleen tnf-alpah1 in Y1.5 and Y3.0 groups and il-8 in Y3.0 group were all significantly higher than those in other groups (P < 0.05) in the post-challenge test. The head kidney gilt expression level was significantly higher in the yeast hydrolysate supplement groups compared with the Y0 group (P < 0.05), and the head kidney il-8 expression level in Y1.5 group was significant higher than those in other groups in post-challenge test (P < 0.05). The present results indicated dietary yeast hydrolysate improved the antioxidant ability and enhanced the immune response of largemouth bass without negative effect on growth. And 1.5% or 3.0% of dietary yeast hydrolysate was recommended for largemouth bass based on the present results.


Assuntos
Bass , Resistência à Doença/efeitos dos fármacos , Doenças dos Peixes/imunologia , Expressão Gênica/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Fermento Seco/metabolismo , Aeromonas hydrophila/fisiologia , Ração Animal/análise , Animais , Bass/crescimento & desenvolvimento , Dieta/veterinária , Suplementos Nutricionais/análise , Doenças dos Peixes/genética , Doenças dos Peixes/metabolismo , Infecções por Bactérias Gram-Negativas/genética , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/metabolismo , Fermento Seco/administração & dosagem
6.
Fish Shellfish Immunol ; 94: 705-710, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31505247

RESUMO

The present study investigates the effect of dietary Ziziphus jujube fruit extract [ZJFE] on skin mucus non-specific immune parameters as well as mRNA levels of immune related gene in the skin of common carp fingerlings. An eight-week feeding trial was performed using different levels of dietary ZJFE (0, 0.25, 0.5 and 1%). At the end of the trial skin mucus immune parameters (total Ig, lysozyme and protease activity), cytokines genes (il1b, il8, il10 and tnf-alpha) expression in skin and growth performance were studied. The result showed highest and lowest skin mucus total Ig were observed in fish fed diet containing 0.5% ZJFE and control group, respectively (P < 0.05). There were no significant difference among treatments regarding skin mucus lysozyme activity (P >0.05). It should be noted that, feeding on 0.5 and 1% ZJFE significantly increased skin mucus protease activity (P <0.05). Likewise, gene expression studies in skin showed significant increase of il1b expression in fish fed 0.5% ZJFE compared other treatments (P <0.05). Also, il8 gene was noticeably up-regulated in 0.5 and 1% treatments compared to the control group (P <0.05). While there were no significant difference between 0.25% JFE treatment and control in case of relative il10 gene expression (P >0.05), feeding on diets containing 0.5% or 1% ZJFE significantly down-regulated il10 gene (P <0.05). Our study indicated that relative expression of tnf-alpha gene significantly increased in treated groups (P <0.05). Also, feeding on ZJFE supplemented diet improved growth performance parameters. Overall, this experiment demonstrated the potentially useful effects of ZJFE on skin mucosal immunity and performance of common carp fingerlings.


Assuntos
Carpas/imunologia , Expressão Gênica/imunologia , Imunidade nas Mucosas/efeitos dos fármacos , Extratos Vegetais/metabolismo , Ração Animal/análise , Animais , Carpas/genética , Carpas/crescimento & desenvolvimento , Dieta/veterinária , Suplementos Nutricionais/análise , Frutas/química , Extratos Vegetais/administração & dosagem , Pele/imunologia , Pele/metabolismo , Ziziphus/química
7.
Fish Shellfish Immunol ; 93: 631-640, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31377431

RESUMO

Fish aquaculture is the world's fastest growing food production industry and infectious diseases are a major limiting factor. Vaccination is the most appropriate method for controlling infectious diseases and a key reason for the success of salmonid cultivation and has reduced the use of antibiotics. The development of fish vaccines requires the use of a great number of experimental animals that are challenged with virulent pathogens. In vitro cell culture systems have the potential to replace in vivo pathogen exposure for initial screening and testing of novel vaccine candidates/preparations, and for batch potency and safety tests. PBL contain major immune cells that enable the detection of both innate and adaptive immune responses in vitro. Fish PBL can be easily prepared using a hypotonic method and is the only way to obtain large numbers of immune cells non-lethally. Distinct gene expression profiles of innate and adaptive immunity have been observed between bacterins prepared from different bacterial species, as well as from different strains or culturing conditions of the same bacterial species. Distinct immune pathways are activated by pathogens or vaccines in vivo that can be detected in PBL in vitro. Immune gene expression in PBL after stimulation with vaccine candidates may shed light on the immune pathways involved that lead to vaccine-mediated protection. This study suggests that PBL are a suitable platform for initial screening of vaccine candidates, for evaluation of vaccine-induced immune responses, and a cheap alternative for potency testing to reduce animal use in aquaculture vaccine development.


Assuntos
Aquicultura/métodos , Vacinas Bacterianas/imunologia , Doenças dos Peixes/prevenção & controle , Expressão Gênica/imunologia , Infecções por Bactérias Gram-Negativas/veterinária , Oncorhynchus mykiss/genética , Oncorhynchus mykiss/imunologia , Aeromonas salmonicida/imunologia , Animais , Vacinas Bacterianas/administração & dosagem , Doenças dos Peixes/imunologia , Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Negativas/prevenção & controle , Técnicas In Vitro/métodos , Leucócitos/imunologia , Yersinia ruckeri/imunologia
8.
Fish Shellfish Immunol ; 94: 27-37, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31465876

RESUMO

In teleost fish, myelopoiesis is maintained both in the head (HK) and trunk kidney (TK), but only the HK holds the endocrine cells that produce the stress hormone cortisol. We now compared the effects of prolonged restraint stress (in vivo) and cortisol (in vitro) on the polarization of HK and TK-derived carp macrophages. Monocytes/macrophages from both sources were treated in vitro with cortisol, lipopolysaccharide or with both factors combined. In vivo, fish were challenged by a prolonged restraint stress. Gene expression of several markers typical for classical M1 and alternative M2 macrophage polarization, as well as glucocorticoid receptors, were measured. Cells from both sources did not differ in the constitutive gene expression of glucocorticoid receptors, whereas they significantly differed in their response to cortisol and stress. In the LPS-stimulated HK monocytes/macrophages, cortisol in vitro counteracted the action of LPS while the effects of cortisol on the activity of TK monocytes/macrophages were less explicit. In vivo, restraint stress up-regulated gene expression of M2 markers in freshly isolated HK monocytes/macrophages, while at the same time it did not affect TK monocytes/macrophages. Moreover, LPS-stimulated HK monocytes/macrophages from stressed animals showed only minor differences in the gene expression of M1 and M2 markers, compared to LPS-treated monocytes/macrophages from control fish. In contrast, stress-induced changes in TK-derived LPS-treated cells were more pronounced. However, these changes did not clearly indicate whether in TK monocytes/macrophages stress will stimulate classical or alternative polarization. Altogether, our results imply that cortisol in vitro and stress in vivo direct HK, but not TK, monocytes/macrophages to the path of alternative polarization. These findings reveal that like in mammals, also in fish the glucocorticoids form important stimulators of alternative macrophage polarization.


Assuntos
Anexina A1/administração & dosagem , Carpas/fisiologia , Proteínas de Peixes/administração & dosagem , Expressão Gênica/imunologia , Hidrocortisona/administração & dosagem , Macrófagos/imunologia , Peptídeos/administração & dosagem , Estresse Fisiológico/imunologia , Animais , Carpas/imunologia , Inflamação/imunologia , Inflamação/veterinária , Macrófagos/metabolismo
9.
Fish Shellfish Immunol ; 94: 132-141, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31461659

RESUMO

A 12-week feeding trial was conducted to investigate the effect of dietary daidzein on the intestinal mucosal barrier function and the intestinal microbiota profile of juvenile turbot (Scophthalmus maximus L.). Three isonitrogenous and isolipidic experimental diets were formulated to contain 0 (FM), 40 (D.40) and 400 (D.400) mg kg-1 daidzein, respectively. Fish fed D.400 had significantly lower growth performance than fish fed D.40. Dietary daidzein significantly increased the feed efficiency, while significantly decreased the feed intake. Daidzein supplementation increased the activity of total anti-oxidative capacity and the gene expression of anti-inflammatory cytokine transforming growth factor-ß1, Mucin-2 and tight junction proteins (Tricellulin, Zonula occludens-1 transcript variant 1, Zonula occludens-1 transcript variant 2 and Claudin-like and Occludin), and down-regulated the gene expression of pro-inflammatory cytokines interleukin-1ß and tumor necrosis factor-α in the intestine of turbot. Dietary daidzein increased intestinal microbial diversities, the abundance of several short chain fatty acids producers, and decreased the abundance of some potential pathogenic bacteria. However, D.400 had dual effects on lactic acid bacteria and increased the abundance of potential harmful bacterium Prevotella copri. Collectively, dietary daidzein at the levels of 40 and 400 mg kg-1 could enhance the intestinal mucosal barrier function and alter the intestinal microbiota of turbot. However, high dose of daidzein must be treated with caution for its unclear effects on intestinal microbiota of turbot in the present study.


Assuntos
Linguados/imunologia , Linguados/microbiologia , Microbioma Gastrointestinal/efeitos dos fármacos , Expressão Gênica/imunologia , Mucosa Intestinal/efeitos dos fármacos , Isoflavonas/metabolismo , Ração Animal/análise , Animais , Dieta/veterinária , Suplementos Nutricionais/análise , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Linguados/genética , Microbioma Gastrointestinal/fisiologia , Mucosa Intestinal/metabolismo , Isoflavonas/administração & dosagem
10.
Fish Shellfish Immunol ; 93: 986-996, 2019 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-31422176

RESUMO

Evolutionary development has increased the diversity of genotypes and the complexity of gene functions in fish. TLR22 has been identified as a teleost-specific gene, but its functions are tremendously different among different fish species. Whether the functional diversity relates to the difference of genotypes remains poorly understand. In this study, we cloned and identified three TLR22 molecules from Schizothorax prenanti (S. prenanti), named as spTLR22-1, spTLR22-2 and spTLR22-3. The full-length coding regions of spTLR22s are 2841 bp, 2805 bp and 2868 bp and coding 946 aa, 934 aa and 955 aa, respectively. All spTLR22s are composed of multiple leucine-rich repeat (LRR) domains, a transmembrane structure and a Toll/IL-1 receptor (TIR) region. The phylogenetic analysis showed that three spTLR22s were close to Cyprinus carpio TLR22-1, TLR22-2 and TLR22-3, respectively. Among the spTLR22s, they presented not close relationship but remained to belong to TLR22 subfamily. All spTLR22s were ubiquitously expressed in all tested tissues, but the expression levels of spTLR22s were dominant in immune-related tissues, such as gill and spleen. The expression levels of spTLR22-1 and spTLR22-3 were significantly increased after treatment with bacteria, LPS and Poly(I:C). However, spTLR22-2 seems like no response to these treatments. The luciferase reporter assay demonstrated that all spTLR22s could activate NF-κB signaling pathway, but only spTLR22-1 and spTLR22-2 could activate IFN-ß signaling pathway. Interestingly, in the ligand recognition analysis, spTLR22-1 and spTLR22-3 but not spTLR22-2 had the recognized potential to Poly(I:C), and all spTLR22s could not recognize LPS. Both spTLR22-1 and spTLR22-3 significantly up-regulated the expression of anti-viral-related genes (Mx, IFN and ISG15) and down-regulated the expression of anti-inflammatory factor IL-10 after the overexpression in carp EPC cell line, but spTLR22-2 failed to impact the expression of these genes. Moreover, we found that all spTLR22s localized to the intracellular region. Taken together, our results reveal that spTLR22-1 and spTLR22-3 but not spTLR22-2 may be involved into the anti-viral immune response via IFN-ß signaling pathway, and all spTLR22s can activate NF-κB signaling pathway but only spTLR22-1 and spTLR22-3 response to the stimulation of bacteria and LPS.


Assuntos
Cyprinidae/genética , Cyprinidae/imunologia , Proteínas de Peixes/genética , Expressão Gênica/imunologia , Receptores Toll-Like/genética , Animais , Fenômenos Fisiológicos Bacterianos , Linhagem Celular , Cyprinidae/metabolismo , Citocinas/metabolismo , Proteínas de Peixes/metabolismo , Perfilação da Expressão Gênica/veterinária , Lipopolissacarídeos/farmacologia , Luciferases/metabolismo , Filogenia , Poli I-C/farmacologia , Análise de Sequência de Proteína/veterinária , Receptores Toll-Like/metabolismo
11.
Fish Shellfish Immunol ; 92: 756-764, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31288098

RESUMO

Tiger puffer (Takifugu rubripes) is one of the major aquaculture fish species in China due to its high economic value. In this study, the transcriptions of hepatic antioxidant enzyme, stress, apoptosis, and immune-related genes of sub-adult tiger puffers (Takifugu rubripes) were evaluated under two different rearing systems [offshore sea cage aquaculture system (OSCS) and recirculating aquaculture system (RAS)]. Results showed that the mRNA expression levels of the antioxidant enzyme (mn-sod, cu/zn-sod, gpx, and gr) and stress-related (hsp70 and hsp90) genes of male tiger puffers reared in the OSCS were significantly higher than female fish reared in the OSCS and fish reared in the RAS. The anti-apoptotic gene bcl2 exhibited the similar results. By contrast, the mRNAs of the pro-apoptotic genes (p53, caspase8, caspase9, and caspase3) of male tiger puffers reared in the OSCS were significantly lower than female fish reared in the OSCS and fish reared in the RAS. Male tiger puffers reared in the OSCS displayed significantly higher complement components (c3) and inflammatory cytokine (il-6) mRNAs, whereas B-cell activating factor (baf) and tumor necrosis factor α (tnf-α) mRNAs remained unchanged. Meanwhile, the mRNA levels of pro-apoptotic (bax, caspase8) and immunity-related (c3, il-6 and il-7) genes of female tiger puffers reared in the OSCS were significantly lower and higher than female fish reared in the RAS, respectively. In conclusion, the hepatic antioxidant, anti-apoptosis, and innate immunity of tiger puffers reared in the OSCS were better than fish in the RAS, male tiger puffer obtained the best values. These results expand the knowledge on the combined RAS and OSCS alternative aquaculture model for tiger puffers and aid in their management in captive.


Assuntos
Apoptose/genética , Aquicultura/métodos , Expressão Gênica/imunologia , Imunidade Inata/genética , Estresse Oxidativo/genética , Takifugu/genética , Animais , Feminino , Perfilação da Expressão Gênica , Masculino , RNA Mensageiro/genética , Takifugu/imunologia
12.
BMC Biotechnol ; 19(1): 44, 2019 07 03.
Artigo em Inglês | MEDLINE | ID: mdl-31269942

RESUMO

BACKGROUND: Engineered therapeutic cells have attracted a great deal of interest due to their potential applications in treating a wide range of diseases, including cancer and autoimmunity. Chimeric antigen receptor (CAR) T-cells are designed to detect and kill tumor cells that present a specific, predefined antigen. The rapid expansion of targeted antigen beyond CD19, has highlighted new challenges, such as autoactivation and T-cell fratricide, that could impact the capacity to manufacture engineered CAR T-cells. Therefore, the development of strategies to control CAR expression at the surface of T-cells and their functions is under intense investigations. RESULTS: Here, we report the development and evaluation of an off-switch directly embedded within a CAR construct (SWIFF-CAR). The incorporation of a self-cleaving degradation moiety controlled by a protease/protease inhibitor pair allowed the ex vivo tight and reversible control of the CAR surface presentation and the subsequent CAR-induced signaling and cytolytic functions of the engineered T-cells using the cell permeable Asunaprevir (ASN) small molecule. CONCLUSIONS: The strategy described in this study could, in principle, be broadly adapted to CAR T-cells development to circumvent some of the possible hurdle of CAR T-cell manufacturing. This system essentially creates a CAR T-cell with an integrated functional rheostat.


Assuntos
Antígenos CD19/imunologia , Expressão Gênica/imunologia , Receptores de Antígenos Quiméricos/imunologia , Linfócitos T/imunologia , Antígenos CD19/genética , Antígenos CD19/metabolismo , Linhagem Celular Tumoral , Membrana Celular/efeitos dos fármacos , Membrana Celular/metabolismo , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/genética , Humanos , Isoquinolinas/farmacologia , Inibidores de Proteases/farmacologia , Receptores de Antígenos Quiméricos/genética , Receptores de Antígenos Quiméricos/metabolismo , Sulfonamidas/farmacologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismo
13.
Gynecol Oncol ; 154(3): 524-530, 2019 09.
Artigo em Inglês | MEDLINE | ID: mdl-31353053

RESUMO

OBJECTIVE: To determine the feasibility of pharmacologic beta-adrenergic blockade in women with newly diagnosed stage II-IV epithelial ovarian cancer (EOC) throughout primary treatment. METHODS: Patients initiated propranolol prior to beginning chemotherapy or surgery. Feasibility was assessed as proportion able to complete 6 chemotherapy cycles while on adrenergic suppression. Descriptive statistics summarized surveys, and paired changes were analyzed using signed rank tests. Random-intercept Tobit models examined immune response. RESULTS: Median age was 59.9; 88.5% were stage IIIC/IV; and 38.5% underwent primary debulking. Thirty-two patients were enrolled; 3 excluded because they never took propranolol; an additional 3 didn't meet inclusion criteria, leaving 26 evaluable. Eighteen of 26 (69%), 90% credible interval (CI) of 53-81%, completed 6 chemotherapy cycles plus propranolol (an 82% posterior probability that the true proportion of success is ≥60%). Among the 23 patients with baseline and six month follow up data, overall QOL, anxiety, and depression improved (P < 0.05) and leukocyte expression of pro-inflammatory genes declined (P = 0.03) after completion of therapy. Decrease from baseline of serum IL-6 and IL-8 preceded response to chemotherapy (P < 0.0014). Change from baseline IL-10 preceded complete response. CONCLUSION: Use of propranolol during primary treatment of EOC is feasible and treatment resulted in decrease in markers of adrenergic stress response. In combination with chemotherapy, propranolol potentially results in improved QOL over baseline.


Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/administração & dosagem , Carcinoma Epitelial do Ovário/tratamento farmacológico , Neoplasias Ovarianas/tratamento farmacológico , Propranolol/administração & dosagem , Antagonistas Adrenérgicos beta/administração & dosagem , Idoso , Carboplatina/administração & dosagem , Carcinoma Epitelial do Ovário/genética , Carcinoma Epitelial do Ovário/imunologia , Carcinoma Epitelial do Ovário/cirurgia , Quimioterapia Adjuvante , Citocinas/sangue , Citocinas/genética , Citocinas/imunologia , Estudos de Viabilidade , Feminino , Expressão Gênica/efeitos dos fármacos , Expressão Gênica/imunologia , Humanos , Leucócitos/efeitos dos fármacos , Leucócitos/imunologia , Estudos Longitudinais , Pessoa de Meia-Idade , Terapia Neoadjuvante , Neoplasias Ovarianas/genética , Neoplasias Ovarianas/imunologia , Neoplasias Ovarianas/cirurgia , Paclitaxel/administração & dosagem , Projetos Piloto , Estudos Prospectivos , Qualidade de Vida
14.
Fish Shellfish Immunol ; 92: 552-569, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31252043

RESUMO

This study investigated the effects of bile acid (BA) supplementation on growth performance, intestinal immune function and the mRNA expression of the related signalling molecules in on-growing grass carp (Ctenopharyngodon idella). A total of 540 healthy grass carp (mean weight 179.85 ±â€¯1.34 g) were fed a normal protein and lipid (NPNL) diet containing 29% crude protein (CP) and 5% ether extract (EE), and five low-protein and high-lipid (LPHL) diets (26% CP, 6% EE) with graded levels of BA (0-320 mg/kg diet) for 50 days. The fish were then challenged with Aeromonas hydrophila for 14 days. The results indicated that compared with the NPNL diet, the LPHL diet (unsupplemented BA) suppressed the growth performance, intestinal development and enteritis resistance capability and impaired the partial intestinal immune function of on-growing grass carp. Whereas in the LPHL diet, optimal BA supplementation significantly improved fish growth performance (percent weight gain, specific growth rate, feed intake and feed efficiency) and intestinal growth and function (intestine weight, intestine length and intestosomatic index), increased beneficial bacteria Lactobacillus and Bifidobacterium amounts, decreased harmful bacteria Aeromonas and Escherichia coli amounts, elevated lysozyme and acid phosphatase activities, increased complement (C3 and C4) and immunoglobulin M contents, and upregulated ß-defensin-1, hepcidin, liver expressed antimicrobial peptide 2A (LEAP-2A), LEAP-2B, Mucin2, interleukin 10 (IL-10), IL-11, transforming growth factor (TGF)-ß1, TGF-ß2, IL-4/13A (not IL-4/13B), TOR, S6K1 and inhibitor of κBα (IκBα) mRNA levels. In addition, optimal BA supplementation in the LPHL diet downregulated tumour necrosis factor α (TNF-α), interferon γ2 (IFN-γ2), IL-1ß, IL-6, IL-8, IL-15, IL-17D, IL-12p35, IL-12p40 (rather than proximal intestine (PI) or mid intestine (MI), nuclear factor kappa B p65 (NF-κB p65) (except NF-κB p52), c-Rel, IκB kinase ß (IKKß), IKKγ (except IKKα), eIF4E-binding proteins (4E-BP)1 and 4E-BP2 mRNA levels in all three intestinal segments of on-growing grass carp (P < 0.05). These findings suggest that BA supplementation in the LPHL diet improves growth and intestinal immune function of fish. Furthermore, 240 mg/kg BA supplementation in the LPHL diet was superior to the NPNL diet in improving growth and enhancing intestinal immune function of fish. Finally, based on percent weight gain, feed intake, protecting fish against enteritis, lysozyme activity in MI and acid phosphatase activity in distal intestine (DI), the optimal BA supplementation for on-growing grass carp were estimated to be 168.98, 170.23, 166.67, 176.50 and 191.97 mg/kg diet, respectively.


Assuntos
Ácidos e Sais Biliares/metabolismo , Carpas/imunologia , Metabolismo dos Lipídeos , NF-kappa B/genética , Serina-Treonina Quinases TOR/genética , Ração Animal/análise , Animais , Ácidos e Sais Biliares/administração & dosagem , Carpas/genética , Carpas/crescimento & desenvolvimento , Carpas/metabolismo , Dieta/veterinária , Suplementos Nutricionais/análise , Expressão Gênica/imunologia , Intestinos/imunologia , NF-kappa B/imunologia , RNA Mensageiro/genética , Transdução de Sinais , Serina-Treonina Quinases TOR/imunologia
15.
Mol Biol Rep ; 46(4): 3617-3623, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31201676

RESUMO

The callitrichids are non-human primates that feed on insects and plant matter in nature, but in captivity, they are fed mostly an artificial diet containing amounts of gluten, in their toxic forms in items such as wheat, barley and rye. The aim of this research was to estimate the blood ß-defensin and Toll like receptor 5 (TLR5) gene expressions and to analyze the stool consistency (firm, soft, diarrheic) in Leontocebus fuscicollis raised in captivity. Blood samples of animals under gluten-free and gluten diets were collected and their fecal output quality was periodically monitored and classified during the course of the study. Gene expression was evaluated using real-time PCR. The stool consistencies of individuals fed a gluten diet were most frequently soft or diarrheic, while it was mostly normal in individuals fed a gluten-free diet. ß-Defensin expression increased in individuals fed a gluten diet, but decreased after 15 days. Expression normalized between 30 and 45 days on a gluten-free diet. However, expression of the TLR5 gene did not change under a gluten diet. A gluten diet affects stool quality, and brings about an immediate increase in blood ß-defensin expression in the beginning but decreases after 15 days.


Assuntos
Dieta Livre de Glúten , Expressão Gênica/imunologia , Glutens/metabolismo , Animais , Callitrichinae , Diarreia , Fezes , Imunidade Inata , Inflamação , Receptor 5 Toll-Like/sangue , beta-Defensinas/sangue
16.
Dev Comp Immunol ; 99: 103404, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31152761

RESUMO

Scavenger receptor (SR) class B (SR-B) is a transmembrane protein that belongs to the SR family with a wide range of functions in innate immunity. Here, an SR-B homologue, designated as AjSR-B, was cloned from the sea cucumber Apostichopus japonicus. AjSR-B comprised 2519 nucleotides with a 5'-untranslated region (UTR) of 153 bp, an open reading frame of 1581 bp encoding a 526 amino acid protein, and a 3'-UTR of 785 bp. SMART analysis indicated that AjSR-B has two transmembrane regions and a cluster determinant 36 domain. Multiple alignments and phylogenetic analysis supported that AjSR-B is a novel member of the SR-B protein family. Moreover, AjSR-B was constitutively expressed in all detected tissues, with the highest levels recorded in the intestine. Both were significantly induced in coelomocytes and the intestine after Vibrio splendidus challenge. Functionally, the recombinant rAjSR-B that corresponds to a large extracellular loop can bind pathogen-associated molecular patterns (PAMPs), including lipopolysaccharide (LPS), peptidoglycan, and mannan, with a high binding affinity to LPS. Bacterial agglutination assay showed that rAjSR-B can agglutinate the four tested bacteria (Gram-negative and Gram-positive bacteria) with calcium dependence. However, the agglutination ability for Gram-negative bacteria completely disappeared in the presence of PAMPs but a weak ability to bind Gram-positive bacteria (Micrococcus luteus) was still exhibited, suggesting there might exist a competition between Gram-positive bacteria and PAMPs under same condition. Our current study indicated that AjSR-B is a PAMP that plays important roles in the innate immune process of sea cucumbers.


Assuntos
Receptores Depuradores Classe B/genética , Receptores Depuradores Classe B/imunologia , Stichopus/genética , Testes de Aglutinação , Sequência de Aminoácidos , Animais , Bactérias/imunologia , Sequência de Bases , Expressão Gênica/imunologia , Imunidade Inata/genética , Fases de Leitura Aberta , Padrões Moleculares Associados a Patógenos/metabolismo , Filogenia , Domínios Proteicos , Receptores de Reconhecimento de Padrão/química , Receptores de Reconhecimento de Padrão/genética , Receptores de Reconhecimento de Padrão/imunologia , Receptores Depuradores Classe B/química , Alinhamento de Sequência , Stichopus/imunologia , Distribuição Tecidual
17.
J Immunol ; 203(2): 329-337, 2019 07 15.
Artigo em Inglês | MEDLINE | ID: mdl-31175163

RESUMO

Despite recent advances in asthma management with anti-IL-5 therapies, many patients have eosinophilic asthma that remains poorly controlled. IL-3 shares a common ß subunit receptor with both IL-5 and GM-CSF but, through α-subunit-specific properties, uniquely influences eosinophil biology and may serve as a potential therapeutic target. We aimed to globally characterize the transcriptomic profiles of GM-CSF, IL-3, and IL-5 stimulation on human circulating eosinophils and identify differences in gene expression using advanced statistical modeling. Human eosinophils were isolated from the peripheral blood of healthy volunteers and stimulated with either GM-CSF, IL-3, or IL-5 for 48 h. RNA was then extracted and bulk sequencing performed. DESeq analysis identified differentially expressed genes and weighted gene coexpression network analysis independently defined modules of genes that are highly coexpressed. GM-CSF, IL-3, and IL-5 commonly upregulated 252 genes and downregulated 553 genes, producing a proinflammatory and survival phenotype that was predominantly mediated through TWEAK signaling. IL-3 stimulation yielded the most numbers of differentially expressed genes that were also highly coexpressed (n = 119). These genes were enriched in pathways involving JAK/STAT signaling. GM-CSF and IL-5 stimulation demonstrated redundancy in eosinophil gene expression. In conclusion, IL-3 produces a distinct eosinophil gene expression program among the ß-chain receptor cytokines. IL-3-upregulated genes may provide a foundation for research into therapeutics for patients with eosinophilic asthma who do not respond to anti-IL-5 therapies.


Assuntos
Citocinas/imunologia , Eosinófilos/imunologia , Expressão Gênica/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/imunologia , Interleucina-3/imunologia , Interleucina-5/imunologia , Asma/imunologia , Regulação para Baixo/imunologia , Humanos , Transdução de Sinais/imunologia , Regulação para Cima/imunologia
18.
Immunol Invest ; 48(8): 835-843, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31122084

RESUMO

MicroRNAs are small non-coding molecules playing a significant regulatory role in several allergic diseases. However their role in tolerance induction remains unclear. The aim of this study was to determine the expression of selected microRNAs during the first three months of wasp venom immunotherapy (VIT). 5 adult patients with a history of severe systemic reactions after stinging by wasps and confirmed sensitization were included. Venous blood samples were collected before VIT, 24 hours after completing its initial phase and after 3 months of the maintenance therapy. A control group was comprised of 5 healthy individuals with no history of allergy. In the blood samples expression of 96 microRNAs was determined with the use of microfluidic cards. In a statistical analysis the expression was compared between the study groups as well as between the pre- and post-VIT samples. Significant differences were found between the patients with wasp venom allergy and the healthy controls in the expression of miR-601 and miR-1201 upregulated in allergic patients at every time point (p = 0.04; p = 0.015, respectively). During VIT profile of microRNA was changing with lower expression of 6 microRNAs (including miR-182, miR-342, miR-375) and higher of 11 microRNAs (including let-7d, miR-34b, miR-143). To conclude, VIT has led to some changes in the expression of microRNA associated with Th2-type inflammation and tolerance induction.


Assuntos
Mordeduras e Picadas/imunologia , Expressão Gênica/imunologia , Imunoterapia/métodos , MicroRNAs/genética , Venenos de Vespas/imunologia , Vespas/imunologia , Adulto , Animais , Dessensibilização Imunológica/métodos , Feminino , Perfilação da Expressão Gênica/métodos , Humanos , Masculino , Pessoa de Meia-Idade , Fatores de Tempo , Venenos de Vespas/administração & dosagem , Adulto Jovem
19.
Fish Shellfish Immunol ; 92: 40-44, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31132466

RESUMO

Stocking density is a crucial factor in shellfish aquaculture that affects overall growth performance and health status. Present study analyzes the effects of stocking densities on growth, survival and hemolymph immune status of noble scallop Chlamys nobilis. The scallops with the same size were separately placed in the lantern cages (10 layers per cage) using high stocking density (500 scallops per cage) and low stocking density (100 scallops per cage) and cultivated in the same location for 60 days. The results indicated that the scallops cultivated at high stocking density had significantly higher mortality and slower growth than those cultivated at low stocking density. Moreover, the hemolymph of scallops cultivated at high density showed significantly higher bacterial load, higher reactive oxygen species (ROS), higher expression level of Nrf2 and lower expression level of Keap1, as well as lower antibacterial ability of Vibrio parahemolyticus than that of scallops cultivated at low density. The present results demonstrated that long-term overcrowding is detrimental for the scallops, which can not only lead to high mortality and slow growth, but also cause more vulnerable to pathogenic bacteria. Therefore, we speculated that high stocking density culture practice of scallops in China might be the root of infectious bacteria outbreaks.


Assuntos
Antioxidantes/metabolismo , Carga Bacteriana/fisiologia , Expressão Gênica/imunologia , Hemolinfa/imunologia , Pectinidae/imunologia , Espécies Reativas de Oxigênio/metabolismo , Vibrio parahaemolyticus/fisiologia , Animais , Aquicultura , Pectinidae/crescimento & desenvolvimento , Pectinidae/microbiologia , Densidade Demográfica , Distribuição Aleatória
20.
Mol Immunol ; 111: 209-219, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31096062

RESUMO

We have previously reported Israa, immune-system-released activating agent, as a novel gene nested in intron 8 of the mouse Zmiz1 gene. We have also shown that Israa encodes for a novel FYN-binding protein and might be involved in the regulation of T-cell activation. In this report, we demonstrate that Israa gene product regulates the expression of a pool of genes involved in T-cell activation and signaling. Real time PCR and GFP knock-in expression analysis showed that Israa is transcribed and expressed in the spleen mainly by CD3+CD8+ cells as well as in the thymus by CD3+ (DP and DN), CD4+SP and CD8+SP cells at different developmental stages. We also showed that Israa is downregulated in T-cells following activation of T-cell receptor. Using yeast two-hybrid analysis, we identified ELF1, a transcription factor involved in T-cell regulation, as an ISRAA-binding partner. Transcriptomic analysis of an EL4 cell line overexpressing ISRAA revealed differential expression of several genes involved in T-cell signaling, activation and development. Among these genes, Prkcb, Mib2, Fos, Ndfip2, Cxxc5, B2m, Gata3 and Cd247 were upregulated whereas Itk, Socs3, Tigit, Ifng, Il2ra and FoxJ1 were downregulated. Our findings support the existence in mouse of a novel FYN-related T-cell regulation pathway involving the product of an intron-nested gene.


Assuntos
Íntrons/imunologia , Ativação Linfocitária/imunologia , Linfócitos/imunologia , Linfocinas/imunologia , Genes Inseridos/imunologia , Transdução de Sinais/imunologia , Linfócitos T/imunologia , Animais , Complexo CD3/imunologia , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Linhagem Celular , Regulação para Baixo/imunologia , Feminino , Expressão Gênica/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Receptores de Antígenos de Linfócitos T/imunologia , Fatores de Transcrição/imunologia , Regulação para Cima/imunologia
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