RESUMO
The brain regulates multiple metabolic processes, such as food intake, energy expenditure, insulin secretion, hepatic glucose production, and glucose and fatty acid metabolism in adipose tissue, which are fundamental for the maintenance of energy and glucose homeostasis during lactation and pregnancy. In addition, brain expression has a fundamental impact on the development of maternal behavior. Although brain functions are partly regulated by long noncoding RNAs (lncRNAs), their expression profiles have not been characterized in depth in any ruminant species. We have sequenced the transcriptome of 12 brain tissues from 3 goats that were 1 mo pregnant and 4 nonpregnant goats to investigate their lncRNA expression patterns. Between 4,363 (adenohypophysis) and 4,604 (olfactory bulb) lncRNAs were expressed in brain tissues, leading us to establish a set of 794 already annotated lncRNAs and 5,098 novel lncRNA candidates. The detected lncRNAs shared features with those of other mammals, and tissue-specific lncRNAs were enriched in brain development-related terms. Differential expression analyses between goats that were 1 mo pregnant and nonpregnant goats showed that the lncRNA expression profiles of certain brain regions experience substantial changes associated with early pregnancy (238 lncRNAs are differentially expressed in the olfactory bulb), but others do not. Enrichment analysis showed that differentially expressed lncRNAs from the olfactory bulb are co-expressed with genes previously linked to behavioral changes related to pregnancy. These findings provide a first characterization of the landscape of lncRNA expression in the goat brain and provides valuable clues to understand the molecular events triggered by early pregnancy in the central nervous system.
Assuntos
Encéfalo , Cabras , RNA Longo não Codificante , Animais , Cabras/genética , Cabras/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Feminino , Encéfalo/metabolismo , Gravidez , Transcriptoma , Lactação/genéticaRESUMO
OBJECTIVES: This study aimed to evaluate the pain perception and several aspects of disrupted body schema, in a sample of patients suffering from fibromyalgia (FM) syndrome. METHODS: Twenty-six patients were organised into two groups: the tactile discrimination group and control group (exposed to tactile stimulation alone). Outcome measures were the pain intensity in body regions commonly described as painful (visual analogue scale) and clinical status, body esteem scale (BES), interoceptive awareness. Tactile acuity was measured by the two-point discrimination test (TPD), hits in the location of the stimulus, the probe size discrimination and the graphesthesia task. RESULTS: The group exposed to tactile discrimination experienced a significant improvement in all tactile acuity outcome measures. The decrease of the Fibromyalgia Impact Questionnaire variable was relevant (81.58, SEM 3.29 vs. 72.91, SEM 6.43; p=0.07). Likewise, pain perception was lower in all of the body regions evaluated (reduction of 12.2% in the stimulated body region (cervical VAS) with a large effect size, a pain reduction of 11.3% in the wrists and 9.2% in the knees. The correlation index showed association between the cervical VAS and TPD (ρ=0.53; p<0.05). CONCLUSIONS: There was no improvement in pain scores in the control group but the TPD was decreased also. The BES scores did not show differences between groups. However, interoceptive awareness showed a slight reduction in the group exposed to tactile discrimination (3.68, SEM 0.15 vs. 3.35, SEM 0.19; p=0.01). After short-term tactile discrimination protocol, the group exposed to tactile discrimination experienced a significant improvement in all tactile acuity outcome measures: pain perception, tactile acuity and body perception, compatible with adjustments in the body schema. The tactile stimulation alone group did not show the same improvement.
Assuntos
Fibromialgia , Percepção do Tato , Imagem Corporal , Fibromialgia/diagnóstico , Fibromialgia/terapia , Humanos , Dor , Percepção da Dor , Percepção do Tato/fisiologiaRESUMO
BACKGROUND: Long non-coding RNAs (lncRNAs) are involved in several immune processes, including the immune response to vaccination, but most of them remain uncharacterised in livestock species. The mechanism of action of aluminium adjuvants as vaccine components is neither not fully understood. RESULTS: We built a transcriptome from sheep PBMCs RNA-seq data in order to identify unannotated lncRNAs and analysed their expression patterns along protein coding genes. We found 2284 novel lncRNAs and assessed their conservation in terms of sequence and synteny. Differential expression analysis performed between animals inoculated with commercial vaccines or aluminium adjuvant alone and the co-expression analysis revealed lncRNAs related to the immune response to vaccines and adjuvants. A group of co-expressed genes enriched in cytokine signalling and production highlighted the differences between different treatments. A number of differentially expressed lncRNAs were correlated with a divergently located protein-coding gene, such as the OSM cytokine. Other lncRNAs were predicted to act as sponges of miRNAs involved in immune response regulation. CONCLUSIONS: This work enlarges the lncRNA catalogue in sheep and puts an accent on their involvement in the immune response to repetitive vaccination, providing a basis for further characterisation of the non-coding sheep transcriptome within different immune cells.
Assuntos
RNA Longo não Codificante , Vacinas , Alumínio , Animais , Perfilação da Expressão Gênica , Imunidade/genética , RNA Longo não Codificante/genética , Ovinos , TranscriptomaRESUMO
BACKGROUND: MicroRNAs (miRNAs) are short endogenous, single-stranded, noncoding small RNA molecules of approximately 22 nucleotides in length. They regulate gene expression posttranscriptionally by silencing mRNA expression, thus orchestrating many physiological processes. The Small Ruminant Lentiviruses (SRLV) group includes the Visna Maedi Virus (VMV) and Caprine Arthritis Encephalitis (CAEV) viruses, which cause a disease in sheep and goats characterized by pneumonia, mastitis, arthritis and encephalitis. Their main target cells are from the monocyte/macrophage lineage. To date, there are no studies on the role of miRNAs in this viral disease. RESULTS: Using RNA-seq technology and bioinformatics analysis, the expression levels of miRNAs during different clinical stages of infection were studied. A total of 212 miRNAs were identified, of which 46 were conserved sequences in other species but found for the first time in sheep, and 12 were completely novel. Differential expression analysis comparing the uninfected and seropositive groups showed changes in several miRNAs; however, no significant differences were detected between seropositive asymptomatic and diseased sheep. The robust increase in the expression level of oar-miR-21 is consistent with its increased expression in other viral diseases. Furthermore, the target prediction of the dysregulated miRNAs revealed that they control genes involved in proliferation-related signalling pathways, such as the PI3K-Akt, AMPK and ErbB pathways. CONCLUSIONS: To the best of our knowledge, this is the first study reporting miRNA profiling in sheep in response to SRLV infection. The known functions of oar-miR-21 as a regulator of inflammation and proliferation appear to be a possible cause of the lesions caused in the sheep's lungs. This miRNA could be an indicator for the severity of the lung lesions, or a putative target for therapeutic intervention.
Assuntos
Infecções por Lentivirus/veterinária , Pulmão/metabolismo , MicroRNAs/genética , Análise de Sequência de RNA/métodos , Doenças dos Ovinos/genética , Animais , Vírus da Artrite-Encefalite Caprina/fisiologia , Análise por Conglomerados , Feminino , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , Interações Hospedeiro-Patógeno , Infecções por Lentivirus/genética , Infecções por Lentivirus/virologia , Pulmão/patologia , Pulmão/virologia , Ovinos , Doenças dos Ovinos/virologia , Vírus Visna-Maedi/fisiologiaRESUMO
Aluminum hydroxide has long been employed as a vaccine adjuvant for its safety profile, although its precise mechanism of action remains elusive. In this study, we investigated the transcriptomic responses in sheep spleen following repetitive vaccination with aluminum adjuvanted vaccines and aluminum hydroxide alone. Notably, this work represents the first exploration of the sheep spleen transcriptome in such conditions. Animals were splitted in 3 treatment groups: vaccine group, adjuvant alone group and control group. A total of 18 high-depth RNA-seq libraries were sequenced, resulting in a rich dataset which also allowed isoform-level analysis. The comparisons between vaccine-treated and control groups (V vs C) as well as between vaccine-treated and adjuvant-alone groups (V vs A) revealed significant alterations in gene expression profiles, including protein coding genes and long non-coding RNAs. Among the differentially expressed genes, many were associated with processes such as endoplasmic reticulum (ER) stress, immune response and cell cycle. The analysis of co-expression modules further indicated a correlation between vaccine treatment and genes related to ER stress and unfolded protein response. Surprisingly, adjuvant-alone treatment had little impact on the spleen transcriptome. Additionally, the role of alternative splicing in the immune response was explored. We identified isoform switches in genes associated with immune regulation and inflammation, potentially influencing protein function. In conclusion, this study provides valuable insights into the transcriptomic changes in sheep spleen following vaccination with aluminum adjuvanted vaccines and aluminum hydroxide alone. These findings shed light on the molecular mechanisms underlying vaccine-induced immune responses and emphasize the significance of antigenic components in aluminum adjuvant mechanism of action. Furthermore, the analysis of alternative splicing revealed an additional layer of complexity in the immune response to vaccination in a livestock species.
Assuntos
Adjuvantes Imunológicos , Baço , Transcriptoma , Vacinação , Animais , Baço/imunologia , Baço/metabolismo , Ovinos , Perfilação da Expressão Gênica , Vacinas/imunologia , Hidróxido de Alumínio/imunologia , Processamento AlternativoRESUMO
MicroRNAs (miRNAs) are evolutionarily conserved small non-coding RNAs that regulate gene expression. In livestock species, miRNAs also show great potential as biomarkers for animal health and product quality. In sheep, few miRNAs have been described in comparison with other livestock species or model organisms. We uniformly analyzed 172 public ovine small RNA sequencing datasets from 21 different tissues in order to predict conserved and novel miRNA precursors and profile their expression patterns. In addition to the 106 annotated sheep miRNAs, 1047 previously unannotated miRNA precursor sequences were detected and 41 % of them were assigned an orthologue from other close species. Considering expression levels, a set of miRNAs with high sequence conservation were detected in all tissues, while 733 mature miRNAs were robustly expressed in at least one tissue. 270 miRNAs showed high tissue specificity index values. Brain, male reproductive tissues and PBMCs showed the most distinct expression patterns. Strikingly, over one hundred precursors from the ruminant specific family of mir-2284/mir-2285 miRNAs were found, which were enriched in immune related tissues. This work supports the known high conservation of many miRNAs, but also highlights the potential of clade-specific innovations in ruminant evolution.
Assuntos
MicroRNAs , Ovinos/genética , Animais , Masculino , MicroRNAs/genética , MicroRNAs/metabolismo , Análise de Sequência de RNA , Especificidade de Órgãos , Perfilação da Expressão GênicaRESUMO
Accumulative evidence has shown that short non-coding RNAs such as miRNAs can regulate the innate and adaptive immune responses. Aluminium hydroxide is a commonly used adjuvant in human and veterinary vaccines. Despite its extended use, its mechanism of action is not fully understood and very few in vivo studies have been done to enhance understanding at the molecular level. In this work, we took advantage of a previous long-term experiment in which lambs were exposed to three different treatments by parallel subcutaneous inoculations with aluminium-containing commercial vaccines, an equivalent dose of aluminium or mock injections. Spleen samples were used for miRNA-seq. A total of 46 and 16 miRNAs were found differentially expressed when animals inoculated with commercial vaccines or the adjuvant alone were compared with control animals, respectively. Some miRNAs previously related to macrophage polarization were found dysregulated exclusively by the commercial vaccine treatment but not in the aluminium inoculated animals. The dysregulated miRNAs in vaccine group let-7b-5p, miR-29a-3p, miR-27a and miR-101-3p are candidates for further research, since they may play key roles in the immune response induced by aluminium adjuvants added to vaccines. Finally, protein-protein interaction network analysis points towards leucocyte transendothelial migration as a specific mechanism in animals receiving adjuvant only.
Assuntos
MicroRNAs , Vacinas , Ovinos , Humanos , Animais , MicroRNAs/genética , Baço , Alumínio , Adjuvantes Imunológicos/farmacologia , VacinaçãoRESUMO
Circular RNAs (circRNAs) are covalently closed circular non-coding RNAs. Due to their structure, circRNAs are more stable and have longer half-lives than linear RNAs making them good candidates for disease biomarkers. Despite the scientific relevance of these molecules, the study of circRNAs in non-model organisms is still in its infancy. Here, we analyse total RNA-seq data to identify circRNAs in sheep from peripheral blood mononuclear cells (PBMCs) and parietal lobe cortex. Out of 2510 and 3403 circRNAs detected in parietal lobe cortex and in PBMCs, a total of 1379 novel circRNAs were discovered. Remarkably, around 63% of all detected circRNAs were found to be completely homologous to a circRNA annotated in human. Functional enrichment analysis was conducted for both tissues based on GO terms and KEGG pathways. The enriched terms suggest an important role of circRNAs from encephalon in synaptic functions and the involvement of circRNAs from PBMCs in basic immune system functions. In addition to this, we investigated the role of circRNAs in repetitive vaccination experiments via differential expression analysis and did not detect any significant relationship. At last, our results support both the miRNA sponge and the miRNA shuttle functions of CDR1-AS in sheep brain. To our knowledge, this is the first study on circRNA annotation in sheep PBMCs or parietal lobe cortex samples.
Assuntos
Splicing de RNA/genética , RNA Circular/genética , Ovinos/genética , Animais , Sequência Conservada , Perfilação da Expressão Gênica , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Redes Reguladoras de Genes , Estudos de Associação Genética , Sequenciamento de Nucleotídeos em Larga Escala , Leucócitos Mononucleares/metabolismo , MicroRNAs/genética , Lobo Parietal/metabolismo , Sítios de Splice de RNA/genética , RNA Circular/isolamento & purificação , RNA Mensageiro/genética , Ovinos/sangue , Vacinação/veterinária , Vacinas/farmacologiaRESUMO
Despite its potential clinical relevance, the product of the DMWD (dystrophia myotonica, WD repeat containing) gene is a largely uncharacterized protein. The DMWD amino acid sequence is similar to that of WDR20, a known regulator of the USP12 and USP46 deubiquitinases (DUBs). Here, we apply a combination of in silico and experimental methods to investigate several aspects of DMWD biology. Molecular evolution and phylogenetic analyses reveal that WDR20 and DMWD, similar to USP12 and USP46, arose by duplication of a common ancestor during the whole genome duplication event in the vertebrate ancestor lineage. The analysis of public human gene expression datasets suggests that DMWD expression is positively correlated with USP12 expression in normal tissues and negatively correlated with WDR20 expression in tumors. Strikingly, a survey of the annotated interactome for DMWD and WDR20 reveals a largely nonoverlapping set of interactors for these proteins. Experimentally, we first confirmed that DMWD binds both USP12 and USP46 through direct coimmunoprecipitation of epitope-tagged proteins. We found that DMWD and WDR20 share the same binding interface in USP12, suggesting that their interaction with the DUB may be mutually exclusive. Finally, we show that both DMWD and WDR20 promote USP12 enzymatic activity, but they differentially modulate the subcellular localization of the DUB. Altogether, our findings suggest a model whereby mutually exclusive binding of DMWD and WDR20 to USP12 may lead to formation of deubiquitinase complexes with distinct subcellular localization, potentially targeting different substrate repertoires.
Assuntos
Proteínas de Transporte/metabolismo , Endopeptidases/metabolismo , Regulação da Expressão Gênica , Distrofia Miotônica/patologia , Proteínas/metabolismo , Ubiquitina Tiolesterase/metabolismo , Repetições WD40 , Sequência de Aminoácidos , Proteínas de Transporte/genética , Endopeptidases/genética , Evolução Molecular , Perfilação da Expressão Gênica , Células HEK293 , Células HeLa , Humanos , Distrofia Miotônica/genética , Distrofia Miotônica/metabolismo , Filogenia , Ligação Proteica , Proteínas/genética , Homologia de Sequência , Ubiquitina Tiolesterase/genéticaRESUMO
Aluminium hydroxide adjuvants are crucial for livestock and human vaccines. Few studies have analysed their effect on the central nervous system in vivo. In this work, lambs received three different treatments of parallel subcutaneous inoculations during 16 months with aluminium-containing commercial vaccines, an equivalent dose of aluminium hydroxide or mock injections. Brain samples were sequenced by RNA-seq and miRNA-seq for the expression analysis of mRNAs, long non-coding RNAs and microRNAs and three expression comparisons were made. Although few differentially expressed genes were identified, some dysregulated genes by aluminium hydroxide alone were linked to neurological functions, the lncRNA TUNA among them, or were enriched in mitochondrial energy metabolism related functions. In the same way, the miRNA expression was mainly disrupted by the adjuvant alone treatment. Some differentially expressed miRNAs had been previously linked to neurological diseases, oxidative stress and apoptosis. In brief, in this study aluminium hydroxide alone altered the transcriptome of the encephalon to a higher degree than commercial vaccines that present a milder effect. The expression changes in the animals inoculated with aluminium hydroxide suggest mitochondrial disfunction. Further research is needed to elucidate to which extent these changes could have pathological consequences.
Assuntos
Adjuvantes Imunológicos/administração & dosagem , Hidróxido de Alumínio/administração & dosagem , Encéfalo/efeitos dos fármacos , Encéfalo/imunologia , Carneiro Doméstico/imunologia , Animais , Encéfalo/metabolismo , Perfilação da Expressão Gênica , Redes Reguladoras de Genes , MicroRNAs/genética , MicroRNAs/metabolismo , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , RNA-Seq , Carneiro Doméstico/genética , Carneiro Doméstico/metabolismo , Transcriptoma/efeitos dos fármacos , Transcriptoma/imunologia , Vacinas/administração & dosagemRESUMO
OBJECTIVE: Evidence suggests that there is an association between chronic pain and disruption of the body schema. We tested the hypothesis in fibromyalgia syndrome (FMS). MATERIALS AND METHODS: We investigated distinct perceptual aspects of the body schema both in a sample of patients with FMS and in pain-free controls. Performances on the left/right judgment task were measured; tactile acuity was assessed by using the 2-point discrimination test. Furthermore, we evaluated sensations evoked by tactile stimulation with von Frey filaments to body parts that were experiencing pain. Anomalous sensations elicited by sensory-motor conflict (SMC) were also investigated. RESULTS: Patients with FMS showed inferior performance on the right/left judgment task, both in terms of correct matches (75.38% vs. 89.67%, respectively; P<0.05) and response time (2.58 s vs. 1.89 s, respectively; P<0.05). Effect sizes were large and very large, respectively. Two-point discrimination thresholds were significantly higher (P<0.05) in participants from the FMS sample (mean of 49.71 mm, SD: 12.09 mm) relative to controls (mean of 37.36 mm, SD: 7.81 mm). Nine of 14 participants with FMS, but no control participants, reported referred sensations upon tactile stimulation, including tingling, pins and needles, weight, and cramps. Referral sites included regions both adjacent and remote to stimulated sites. Patients with FMS scored across all items within the administered questionnaire addressing anomalous sensations on the mirror setup (Cohen d=1.04 to 2.42 across all items), and FMS patients perceived pain during the SMC (the required statistical power for it to be statistically significant was 96% and for it to be recognized as a difference of means in pain item). CONCLUSION: Our present findings suggest a disrupted body schema and propensity to experiencing anomalous somatosensory sensations during SMC in people with FMS.
Assuntos
Imagem Corporal/psicologia , Discriminação Psicológica/fisiologia , Fibromialgia/psicologia , Percepção do Tato/fisiologia , Tato/fisiologia , Adulto , Feminino , Fibromialgia/fisiopatologia , Humanos , Julgamento/fisiologia , Pessoa de Meia-Idade , Medição da Dor , Limiar da Dor/fisiologiaRESUMO
Paratuberculosis is chronic granulomatous enteritis of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). Whole RNA-sequencing (RNA-Seq) is a promising source of novel biomarkers for early MAP infection and disease progression in cattle. Since the blood transcriptome is widely used as a source of biomarkers, we analyzed whether it recapitulates, at least in part, the transcriptome of the ileocecal valve (ICV), the primary site of MAP colonization. Total RNA was prepared from peripheral blood (PB) and ICV samples, and RNA-Seq was used to compare gene expression between animals with focal or diffuse histopathological lesions in gut tissues versus control animals with no detectable signs of infection. Our results demonstrated both shared, and PB and ICV-specific gene expression in response to a natural MAP infection. As expected, the number of differentially expressed (DE) genes was larger in the ICV than in the PB samples. Among the DE genes in the PB and ICV samples, there were some common genes irrespective of the type of lesion including the C-X-C motif chemokine ligand 8 (CXCL8/IL8), apolipoprotein L (APOLD1), and the interferon inducible protein 27 (IFI27). The biological processes (BP) enriched in the PB gene expression profiles from the cows with diffuse lesions included the killing of cells of other organism, defense response, immune response and the regulation of neutrophil chemotaxis. Two of these BP, the defense and immune response, were also enriched in the ICV from the cows with diffuse lesions. Metabolic analysis of the DE genes revealed that the N-glycan biosynthesis, bile secretion, one-carbon pool by folate and purine metabolism were significantly enriched in the ICV from the cows with focal lesions. In the ICV from cows with diffuse lesions; the valine, leucine and isoleucine degradation route, purine metabolism, vitamin digestion and absorption and the cholesterol routes were enriched. Some of the identified DE genes, BP and metabolic pathways will be studied further to develop novel diagnostic tools, vaccines and immunotherapeutics.
Assuntos
Doenças dos Bovinos/imunologia , Valva Ileocecal/imunologia , Valva Ileocecal/metabolismo , Interleucina-8 , Paratuberculose/imunologia , Animais , Biomarcadores/sangue , Bovinos , Feminino , Valva Ileocecal/patologia , Interleucina-8/sangue , Interleucina-8/imunologia , Mycobacterium avium subsp. paratuberculosis/imunologia , RNA-Seq , Transdução de Sinais , TranscriptomaRESUMO
Fibromyalgia syndrome (FMS) is a highly prevalent, chronic musculoskeletal condition characterized by widespread pain and evoked pain at tender points. This study evaluated various aspects of body awareness in a sample of 14 women with FMS and 13 healthy controls, such as plasticity of the body schema, body esteem, and interoceptive awareness. To this end, the Rubber Hand Illusion (RHI), the Body Esteem Scale (BES), and the Body Perception Questionnaire (BPQ) were used, respectively. Consistent with increased plasticity of the body schema, FMS patients scored higher, with large or very large effect sizes, across all three domains evaluated in the RHI paradigm, namely proprioceptive drift and perceived ownership and motor control over the rubber hand. Scores on all items addressed by the BES were consistently lower among FMS subjects (2.52, SEM .19 vs 3.89, SEM .16, respectively, p < .01, Cohen's d = .38-.66). In the FMS sample, BES scores assigned to most painful regions also were lower than those assigned to the remaining body sites (1.58, SEM .19 vs 2.87, SEM .18, respectively, p < .01). Significantly higher scores (p < .01, Cohen's d = .51-.87) were found in the FMS sample across awareness (3.57 SEM .15 vs 1.87 SEM .11), stress response (3.76 SEM .11 vs 1.78 SEM .11), autonomic nervous system reactivity (2.59 SEM .17 vs 1.35 SEM .07), and stress style 2 (2.73 SEM .27 vs 1.13 SEM .04) subscales of the BPQ. Intensity of ongoing clinical pain was found to be strongly correlated with interoceptive awareness (r = .75, p = .002). The results suggest a disturbed embodiment in FMS, characterized by instability of the body schema, negatively biased cognitions regarding one's own body, and increased vigilance to internal bodily cues. These manifestations may be interpreted as related with the inability of incoming sensory inputs to adequately update negatively biased off-line somatorepresentations stored as long-term memory.
Assuntos
Fibromialgia/fisiopatologia , Adulto , Idoso , Conscientização , Imagem Corporal , Estudos de Casos e Controles , Feminino , Fibromialgia/diagnóstico , Humanos , Interocepção , Pessoa de Meia-Idade , Inquéritos e QuestionáriosRESUMO
There have been few in vivo studies on the effect of aluminum hydroxide adjuvant and its influence on the immune response to vaccination. In this study, lambs received a parallel subcutaneous treatment with either commercial vaccines containing aluminum hydroxide or an equivalent dose of this compound only with the aim of identifying the activated molecular signature. Blood samples were taken from each animal at the beginning and at the end of the experiment and PBMCs isolated. Total RNA and miRNA libraries were prepared and sequenced. After alignment to the Oar3.1 reference genome and differential expression with 3 programs, gene enrichment modeling was performed. For miRNAs, miRBase and RNAcentral databases were used for detection and characterization. Three expression comparisons were made: vaccinated animals at the beginning and at the end of the treatment, adjuvanted animals at the same times, and animals of both treatments at the end of the experiment. After exposure to both treatments, a total of 2,473; 2,980 and 429 differentially expressed genes were identified in vaccinated animals, adjuvanted animals and animals at the end of both treatments, respectively. In both adjuvant and vaccine treated animals the NF-κB signaling pathway was enriched. On the other hand, it can be observed a downregulation of cytokines and cytokine receptors in the adjuvanted group compared to the vaccinated group at the final time, suggesting a milder induction of the immune response when the adjuvant is alone. As for the miRNA analysis, 95 miRNAs were detected: 64 previously annotated in Ovis aries, 11 annotated in Bos taurus and 20 newly described. Interestingly, 6 miRNAs were differentially expressed in adjuvant treated animals, and 3 and 1 in the other two comparisons. Lastly, an integrated miRNA-mRNA expression profile was developed, in which a miRNA-mediated regulation of genes related to DNA damage stimulus was observed. In brief, it seems that aluminum-containing adjuvants are not simple delivery vehicles for antigens, but also induce endogenous danger signals that can stimulate the immune system. Whether this contributes to long-lasting immune activation or to the overstimulation of the immune system remains to be elucidated.
Assuntos
Adjuvantes Imunológicos , Hidróxido de Alumínio/imunologia , Sequenciamento do Exoma/métodos , Leucócitos Mononucleares/fisiologia , MicroRNAs/genética , RNA Mensageiro/genética , Carneiro Doméstico/genética , Animais , Bovinos , Dano ao DNA/genética , Humanos , Imunidade , Carneiro Doméstico/imunologia , Transcriptoma , Vacinação , Vacinas/imunologiaRESUMO
Activation of the N-methyl-d-aspartate receptor (NMDAR) in dorsal horn neurons is recognized as a fundamental mechanism of central sensitization and pathologic pain. This study assessed the influence of dopaminergic, D1-like receptor-mediated input to the spinal dorsal horn on NMDAR function. Spinal superfusion with selective NMDAR agonist cis-ACPD significantly increased C-fiber-evoked field potentials in rats subjected to spinal nerve ligation (SNL), but not in sham-operated rats. Simultaneous application of D1LR antagonist SCH 23390 dramatically reduced hyperexcitability induced by cis-ACPD. Furthermore, cis-ACPD-induced hyperexcitability seen in nerve-ligated rats could be mimicked in unin-jured rats during stimulation of D1LRs by agonist SKF 38393 at subthreshold concentration. Phosphorylation of NMDAR subunit NR1 at serine 889 at postsynaptic sites was found to be increased in dorsal horn neurons 90 min after SNL, as assessed by increased co-localization with postsynaptic marker PSD-95. Increased NR1 phosphorylation was attenuated in the presence of SCH 23390 in the spinal superfusate. The present results support that D1LRs regulate most basic determinants of NMDAR function in dorsal horn neurons, suggesting a potential mechanism whereby dopaminergic input to the dorsal horn can modulate central sensitization and pathologic pain.