RESUMO
BACKGROUND: This study aimed to investigate the interactions among three core elements of respiratory infection-pathogen, lung microbiome, and host response-and their avocation with the severity and outcomes of Mycoplasma pneumoniae pneumonia (MPP) in children. METHODS: We prospectively collected bronchoalveolar lavage fluid from a cohort of 41 children with MPP, including general MPP (GMPP) and complicated MPP (CMPP), followed by microbiome and transcriptomic analyses to characterize the association among pathogen, lung microbiome, and host response and correlate it with the clinical features and outcomes. RESULTS: The lung microbiome of patients with CMPP had an increased relative abundance of Mycoplasma pneumoniae (MP) and reduced alpha diversity, with 76 differentially expressed species. Host gene analysis revealed a key module associated with neutrophil function and several inflammatory response pathways. Patients with a high relative abundance of MP, manifested by a specific lung microbiome and host response type, were more prone to CMPP and had a long imaging recovery time. CONCLUSION: Patients with CMPP have a more disrupted lung microbiome than those with GMPP. MP, lung microbiome, and host response interacts with each other and are closely related to disease severity and outcomes in children with MPP.
Assuntos
Mycoplasma pneumoniae , Nitrobenzenos , Compostos Organofosforados , Pneumonia por Mycoplasma , Criança , Humanos , Mycoplasma pneumoniae/genética , Transcriptoma , Pneumonia por Mycoplasma/diagnóstico , Pneumonia por Mycoplasma/genética , PulmãoRESUMO
Mycoplasma pneumoniae (MP) is primarily recognized as a respiratory pathogen that causes community-acquired pneumonia, which can lead to acute upper and lower airway inflammation and extrapulmonary syndrome. Refractory pneumonia caused by MP can cause severe complications and even be life-threatening, particularly in infants and the elderly. It is well-known that non-coding RNAs (ncRNAs) represented by miRNAs, lncRNAs and circRNAs have been manifested to be widely involved in the regulation of gene expression. Growing evidence indicates that these ncRNAs have distinct differentiated expression in MP infection and affect multiple biological processes, playing an indispensable role in the initiation and promotion of MP infection. However, the epigenetic mechanisms involved in the development of MP infection remain unclear. This article reviews the mechanisms by which miRNAs, lncRNAs, and circRNAs mediate MP infection, such as inflammatory responses, apoptosis and pulmonary fibrosis. Focusing on miRNAs, lncRNAs and circRNAs associated with MP infection could provide new insights into this disease's early diagnosis and therapeutic approaches.
Assuntos
MicroRNAs , Pneumonia por Mycoplasma , RNA Longo não Codificante , Lactente , Humanos , Idoso , Pneumonia por Mycoplasma/tratamento farmacológico , Pneumonia por Mycoplasma/genética , MicroRNAs/genética , RNA Circular/genética , RNA Circular/uso terapêutico , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , RNA Longo não Codificante/uso terapêutico , Mycoplasma pneumoniae/genéticaRESUMO
Screening of acute respiratory infections causes serious challenges in urgent point-of-care scenarios where conventional methods are impractical and alternative techniques suffer from low accuracy, poor robustness, and reliance on sophisticated instruments. As an improvement to this paradigm, we report a point-of-care lateral flow biosensor (LFB) based on the recognition property of clustered regularly interspaced short palindromic repeats (CRISPR)/associated protein 9 (Cas9) and apply it to the detection of Mycoplasma pneumoniae (M. pneumoniae). The designed biosensor employs CRISPR/Cas9 for secondary recognition after preamplification of target gene using specific primer set, avoiding false positives caused by nontarget factors. The high amplification efficiency and low applicable temperatures of recombinase polymerase amplification brings the detection limit of the biosensor to 3 copies even at a preamplification temperature of 25 °C. Its practical application is further demonstrated with 100% accuracy by testing with 43 M. pneumoniae-infected specimens and 80 uninfected specimens. Additionally, the entire detection, including pretreatment, preamplification, CRISPR/Cas9 recognition, and visual analysis, can be completed in 30 min. Featured with the combination of CRISPR/Cas9 and LFB, the biosensor we developed herein ensures excellent convenience, accuracy, and robustness, which endows promising point-of-care screening potential for infectious pathogens.
Assuntos
Técnicas Biossensoriais , Pneumonia por Mycoplasma , Humanos , Sistemas CRISPR-Cas , Pneumonia por Mycoplasma/diagnóstico , Pneumonia por Mycoplasma/genética , Sistemas Automatizados de Assistência Junto ao Leito , Técnicas Biossensoriais/métodosRESUMO
Background: The functional causal single-nucleotide polymorphisms (SNPs) associated with susceptibility to Mycoplasma pneumoniae Pneumonia (MPP) have scarcely been identified. In this study, we aimed to analyze the association between the functional expression quantitative trait locus (eQTL)-SNPs and the risk of MPP. Methods: First, we identified reported genes associated with MPP from the human disease database, MalaCards. After investigating multiple databases, we systematically selected seven functional eQTL-SNPs (rs2070874, rs360720, rs8032531, rs4316, rs4353, rs7258241, and rs2250656). Finally, the selected eQTL-SNPs were genotyped using the TaqMan genotyping technology, and compared between 100 children with MPP and 178 healthy controls. Results: We found that three eQTL-SNPs (rs8032531 in CD276 and rs4316 and rs4353 in ACE) were significantly associated with susceptibility to MPP. Joint analysis of the three eQTL-SNPs revealed that the risk of MPP increased with an increase in the number of risk alleles present. Plasma protein expression levels of CD276 and ACE were distinctively higher in children with MPP than in healthy children (CD276: P < 0.001; ACE: P = 0.001). Conclusion: Functional eQTL-SNPs in CD276 and ACE may affect the susceptibility to MPP. The risk of developing MPP is higher in patients harboring a greater number of unfavorable alleles of the aforementioned SNPs.
Assuntos
Mycoplasma pneumoniae , Pneumonia por Mycoplasma , Antígenos B7/genética , Criança , Humanos , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/genética , Polimorfismo de Nucleotídeo Único , Locos de Características Quantitativas/genéticaRESUMO
Mycoplasma pneumoniae pneumonia (MPP) represents a common respiratory disease in children patients. Kukoamine A (KuA) is a spermine alkaloid found in the Chinese herb Cortex Lycii radices, which has a variety of pharmacological properties. However, no study has been reported on the role of KuA in MPP. Exosomes, a type of lipid bilayer-enclosed extracellular vesicles, can be delivered to the target cells, where they regulate function and physiology. With the use of human alveolar basal epithelial cells (HABECs) as an in vitro model, in this study, we sought to characterize the changes in levels of superoxide dismutase 2 (SOD2) and proinflammatory cytokines including IL-6 and TNF-α in HABECs in response to exosomes, which were isolated from peripheral blood serum of MPP patients. We found that, compared to normal, MPP patients exhibited a significant up-regulated miR-222-3p. Further, exosomal miR-222-3p downregulated SOD2 activity but promoted nuclear NF-κB activity and expression of IL-6 and TNF-α in HABECs, ultimately leading to an oxidative stress condition. Interestingly, such stimulating effects were attenuated by the pretreatment of KuA. This study suggests a critical role possessed by KuA in MPP by regulating the miR-222-3p/SOD2 axis, which represents a promising strategy for the treatment of MPP.
Assuntos
MicroRNAs , Mycoplasma pneumoniae , Pneumonia por Mycoplasma , Espermina , Criança , Humanos , MicroRNAs/genética , MicroRNAs/metabolismo , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/tratamento farmacológico , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/metabolismo , Espermina/análogos & derivados , Espermina/farmacologia , Superóxido Dismutase , Fator de Necrose Tumoral alfa/metabolismoRESUMO
OBJECTIVE: This study aimed to investigate the role of p50-associated cyclooxygenase- (COX-2) extragenic RNA (PACER) on the inflammation of airway epithelium caused by Mycoplasma pneumoniae (MP) infection. METHODS: A549 cells and MP strain were cultured respectively. The expressions of PACER, IL-8, TNF-α and COX-2 in MP-infected cells were detected by qRT-PCR, the concentration of IL-8 and TNF-α in the supernatant of the cells were detected by ELISA, and the expression of COX-2 protein in the cells was detected by western-blot. After knockdown of PACER, the expression of IL-8, TNF-α and COX-2 in MP infected cells were observed. The activity of NF-κB in cells was detected by fluorescence reporter assay, and the interaction between PACER and NF-κB was verified by RNA immunoprecipitation. RESULTS: First, we observed that PACER was upregulated in MP infected A549 cells. Knockdown of PACER suppressed the production of inflammatory cytokines as well as the expression of COX-2 in A549 cells after MP infection. By performing luciferase reporter assay, we found PACER knockdown inhibited NF-κB activation induced by MP. Furthermore, RNA immunoprecipitation showed that PACER could physically bind to NF-κB p50 in MP-treated A549 cells. CONCLUSION: Collectively, our data demonstrated that attenuation of PACER reduces the inflammatory response of MP-infected epithelial cells via regulating NF-κB.
Assuntos
Mycoplasma pneumoniae , NF-kappa B , Pneumonia por Mycoplasma , RNA Longo não Codificante , Células A549 , Humanos , Inflamação/genética , Inflamação/metabolismo , Inflamação/microbiologia , NF-kappa B/metabolismo , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/metabolismo , Pneumonia por Mycoplasma/microbiologia , RNA Longo não Codificante/genética , RNA Longo não Codificante/metabolismo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Pneumonia caused by Mycoplasma pneumoniae (M. pneumoniae) is a major cause of communityacquired pneumonia in children. In some cases, M. pneumoniae pneumonia (MPP) can develop into refractory MPP (RMPP), which shows no clinical or radiological response to macrolides, and can progress to severe and complicated pneumonia. However, the pathogenesis of RMPP remains poorly understood. The present study aimed to identify target genes that could be used as biomarkers for the clinical diagnosis of earlystage RMPP through highthroughput sequencing technology. The differences in long noncoding (lnc)RNAs, mRNAs and circular (circ)RNAs were examined between wholeblood samples from two patients with nonrefractory MPP (NRMPP), two patients with RMPP and three healthy children using ribosomal (r)RNAdepleted RNAsequencing techniques and an integrated mRNA/circRNA analysis. A total of 17 lncRNAs (four upregulated and 13 downregulated), 18 mRNAs (six upregulated and 12 downregulated) and 24 circRNAs (12 upregulated and 12 downregulated) were the most significantly differentially expressed (P<0.05) between the NRMPP and RMPP groups. Upon functional analysis, the significantly differentially expressed genes encoded by the targeting mRNAs (prostaglandinendoperoxide synthase 2, IL8 and foslike antigen 1) were screened and identified to be enriched in the 'IL17 signaling pathway'. Furthermore, the key circRNAs in the NRMPP and RMPP comparative groups were primarily enriched in 'herpes simplex virus 1 infection', 'viral carcinogenesis' and 'RNA transport'. In the present study, a comprehensive analysis of the differences between the NRMPP and RMPP cases was performed based on rRNAdepleted RNAsequencing techniques, and the selected genes and circRNAs may be closely associated with the complex pathogenesis of RMPP.
Assuntos
Antibacterianos/farmacologia , Infecções Comunitárias Adquiridas/tratamento farmacológico , Farmacorresistência Bacteriana/genética , Mycoplasma pneumoniae/patogenicidade , Pneumonia por Mycoplasma/tratamento farmacológico , Antibacterianos/uso terapêutico , Biomarcadores/análise , Criança , Pré-Escolar , Infecções Comunitárias Adquiridas/genética , Infecções Comunitárias Adquiridas/imunologia , Infecções Comunitárias Adquiridas/microbiologia , Feminino , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Humanos , Lactente , Macrolídeos/farmacologia , Macrolídeos/uso terapêutico , Masculino , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Pneumonia por Mycoplasma/microbiologia , RNA Circular/análise , RNA Longo não Codificante/análise , RNA Mensageiro/análise , RNA Ribossômico , Análise de Sequência de RNA/métodosRESUMO
The secretory function of airway epithelial cells is important in the pathogenesis of Mycoplasma pneumoniae pneumonia (MPP). To investigate the regulatory function of NKILA (nuclear factor-κB (NF-κB) interacting long noncoding RNA (lncRNA)) in MPP, we first detected NKILA as well as the concentration of interleukin 8 (IL-8) and tumor necrosis factor-α (TNF-α) in bronchoalveolar lavage fluid of children with MPP. Then, NKILA was knocked down in epithelial cells to investigate its effect on their secretory function. The results suggested that NKILA was downregulated in children with MPP, while IL-8 and TNF-α levels increased. Knockdown of NKILA in vitro promoted the inflammatory effects of Mycoplasma pneumoniae (MP) in epithelial A549 and BEAS-2B cells. Knockdown of NKILA promoted inhibitor of κBα (IκBα) phosphorylation and degradation, and NF-κB p65 nuclear translocation. Furthermore, RNA immunoprecipitation showed that NKILA could physically bind to IκBα in MP-treated A549 cells. Collectively, our data demonstrated that attenuation of NKILA enhances the effects of MP-stimulated secretory functions of epithelial cells via regulation of NF-κB signaling.
Assuntos
Células Epiteliais/metabolismo , Células Epiteliais/microbiologia , Pulmão/patologia , Mycoplasma pneumoniae/fisiologia , NF-kappa B/metabolismo , RNA Longo não Codificante/metabolismo , Células A549 , Líquido da Lavagem Broncoalveolar , Pré-Escolar , Citocinas/genética , Citocinas/metabolismo , Regulação para Baixo/genética , Células Epiteliais/patologia , Feminino , Humanos , Mediadores da Inflamação/metabolismo , Masculino , Inibidor de NF-kappaB alfa/metabolismo , Fosforilação , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/microbiologia , Ligação Proteica , Transporte Proteico , RNA Longo não Codificante/genética , Fator de Transcrição RelA/metabolismoRESUMO
Mycoplasma pneumoniae pneumonia (MPP) is a type of pneumonia induced by M. pneumoniae (MP) infection. The present study investigated the effect of long noncoding RNA growth arrestspecific 5 (GAS5) in MPP and the underlying molecular mechanism of this. The expression of GAS5, microRNA2223p, (miR2223p) and tissue inhibitor of metalloproteinases3 (TIMP3) in MPP was investigated using reverse transcriptionquantitative PCR. Lipidassociated membrane protein (LAMP)induced THP1 cells were used to model MPP. The viability of LAMPinduced THP1 cells was analyzed using an MTT assay. Expression levels of interleukin (IL)1ß, IL6 and tumor necrosis factorα (TNFα) proinflammatory cytokines, and the antiinflammatory cytokine heme oxygenase1 (HO1) in LAMPinduced THP1 cells were measured by ELISA. A dualluciferase reporter assay assessed the associations among GAS5, miR2223p and TIMP3. The expression of GAS5 and TIMP3 was downregulated in MPP. Expression of miR2223p was upregulated. GAS5overexpression increased the viability of LAMPinduced THP1 cells. GAS5 upregulation decreased the levels of IL1ß, IL6, TNFα and HO1 levels in LAMPinduced THP1 cells. GAS5 directly interacted with miR2223p. TIMP3 was a target of miR2223p. miR2223p upregulation or TIMP3knockdown reversed the promotion effect on cell viability as well as the inhibitory effect on inflammation caused by GAS5overexpression in LAMPinduced THP1 cells. GAS5overexpression increased the viability and decreased the inflammation of LAMPinduced THP1 cells by regulating the miR2223p/TIMP3 axis. These results demonstrated a potential therapeutic target for MPP treatment.
Assuntos
Inflamação/genética , MicroRNAs/genética , Pneumonia por Mycoplasma/genética , RNA Longo não Codificante/genética , Inibidor Tecidual de Metaloproteinase-3/genética , Apoptose/genética , Citocinas/genética , Regulação da Expressão Gênica/genética , Humanos , Inflamação/microbiologia , Inflamação/patologia , Mycoplasma pneumoniae/genética , Mycoplasma pneumoniae/patogenicidade , Pneumonia por Mycoplasma/microbiologia , Pneumonia por Mycoplasma/patologiaRESUMO
Mycoplasma ovipneumoniae infects both sheep and goats causing pneumonia resulting in considerable economic losses worldwide. Current diagnosis methods such as bacteriological culture, serology, and PCR are time consuming and require sophisticated laboratory setups. Here we report the development of two rapid, specific and sensitive assays; an isothermal DNA amplification using recombinase polymerase amplification (RPA) and a real-time PCR for the detection of M. ovipneumoniae. The target for both assays is a specific region of gene WP_069098309.1, which encodes a hypothetical protein and is conserved in the genome sequences of ten publicly available M. ovipneumoniae strains. The RPA assay performed well at 39°C for 20 min and was combined with a lateral flow dipstick (RPA-LFD) for easy visualization of the amplicons. The detection limit of the RPA-LFD assay was nine genome copies of M. ovipneumoniae per reaction and was comparable to sensitivity of the real-time PCR assay. Both assays showed no cross-reaction with 38 other ovine and caprine pathogenic microorganisms and two parasites of ruminants, demonstrating a high degree of specificity. The assays were validated using bronchoalveolar lavage fluid and nasal swab samples collected from sheep. The positive rate of RPA-LFD (97.4%) was higher than the real-time PCR (95.8%) with DNA as a template purified from the clinical samples. The RPA assay was significantly better at detecting M. ovipneumoniae in clinical samples compared to the real-time PCR when DNA extraction was omitted (50% and 34.4% positive rate for RPA-LFD and real-time PCR respectively). The RPA-LFD developed here allows easy and rapid detection of M. ovipneumoniae infection without DNA extraction, suggesting its potential as a point-of-care test for field settings.
Assuntos
Mycoplasma ovipneumoniae/patogenicidade , Pneumonia por Mycoplasma/microbiologia , Recombinases/metabolismo , Animais , Mycoplasma ovipneumoniae/genética , Mycoplasma ovipneumoniae/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico , Plasmídeos/genética , Pneumonia por Mycoplasma/genética , Reação em Cadeia da Polimerase em Tempo RealRESUMO
Mannose-binding lectin (MBL) glycoproteins in blood can selectively recognise lectins on the surface of bacteria, and play an important role in natural immunity. Micro RNAs (miRNAs) are key molecules that regulate gene expression at the post-transcriptional level in vivo, and their pathways are specific and effective. Previous studies indicate that small RNAs such as miRNAs perform regulatory roles in immunology. Herein, we investigated differential expression of miRNAs during MBL protein immunotherapy in sheep following treatment with different MBL genotypes (resistant and susceptible), and identified miRNAs linked to different target genes and pathways. RNA was extracted from liver tissue of resistant and susceptible sheep, miRNAs were identified by high-throughput sequencing, and differentially expressed miRNAs were analysed by SOAP to predict target genes and biological pathways. Results: Some miRNAs (oar-mir-143, oar-mir-10b, oar-mir-382, oar-mir-432 and oar-mir-379) were up-regulated, while others were down-regulated. GPATCH3 and DNAJC5 were predicted target genes of oar-mir-379, DMRT1 and GATA4 were linked to oar-mir-382, and oar-mir-432 was associated with STAT2, DMRT1 and ATG16L1. Identification of miRNAs differentially expressed in resistant and susceptible sheep may expand our understanding of miRNAs in immune regulation, and the role of MBL in innate immunity.
Assuntos
Lectinas de Ligação a Manose/genética , MicroRNAs/fisiologia , Pneumonia por Mycoplasma/veterinária , Doenças dos Ovinos/genética , Animais , Resistência à Doença/genética , Genótipo , Sequenciamento de Nucleotídeos em Larga Escala/veterinária , MicroRNAs/genética , MicroRNAs/metabolismo , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Fator de Transcrição STAT2/metabolismo , Ovinos , Doenças dos Ovinos/imunologiaRESUMO
Herein, we found that serum concentration of superoxide dismutase 3 (SOD3) was significantly reduced in children with mycoplasma pneumonia (MP) infection. To study the roles of SOD3 in inflammatory regulation of MP infection, human A549 type II alveolar epithelial cells were stimulated with 107 CCU/ml of MP to build MP infection in vitro. Secretion of pro-inflammatory cytokine interleukin (IL)-8 and tumor necrosis factor (TNF)-α were measured via enzyme-linked immunosorbent assay (ELISA) to assess the inflammatory response of A549 cells. Levofloxacin (LVFX) was used as an anti-inflammatory drug while recombinant TNF-α was used as an inflammatory promotor in MP-infected cells. Transcriptional activity of nuclear factor (NF)-κB was assessed by detecting protein levels of nuclear NF-κB and cytoplasm NF-κB using Western blot analysis. Our data suggested that the expression of SOD3 mRNA and protein, as well as content of SOD3 in cultured supernatant, were time-dependently inhibited in MP-infected A549 cells. However, lentiviruses-mediated SOD3 overexpression alleviated inflammatory response of MP-infected A549 cells, and prevented the unclear translocation of NF-κB, as evidenced by obviously reducing the production of IL-8 and TNF-α in cell cultured supernatant, as well as decreasing nuclear NF-κB while increasing cytoplasm NF-κB. Inspiringly, SOD3 overexpression induced anti-inflammatory effect and the inactivation of NF-κB was similar to that of 2 lg/ml of LVFX, but reversed by additional TNF-α treatment. Therefore, we can conclude that transcriptional activity of NF-jB was the underlying mechanism, by which SOD3 regulated inflammatory response in MP infection in vitro.
Assuntos
Inflamação/genética , Interleucina-8/genética , Pneumonia por Mycoplasma/genética , Superóxido Dismutase/genética , Fator de Necrose Tumoral alfa/genética , Células A549 , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/genética , Criança , Humanos , Inflamação/tratamento farmacológico , Inflamação/microbiologia , Levofloxacino/farmacologia , Lipopolissacarídeos/farmacologia , Mycoplasma pneumoniae/patogenicidade , Pneumonia por Mycoplasma/microbiologia , Pneumonia por Mycoplasma/patologia , RNA Mensageiro/genéticaRESUMO
The Bashbay sheep (Ovis aries), an indigenous breed of Xinjiang, China, has many excellent characteristics. It is resistant to Mycoplasma ovipneumoniae infection, the causative agent of mycoplasma ovipneumonia, a chronic respiratory disease that is harmful to the sheep industry. To date, knowledge regarding the mechanisms responsible for M. ovipneumoniae pathogenesis in scant. Herein, we report the results of transcriptome profiling of lung tissues from Bashbay sheep experimentally infected with an M. ovipneumoniae strain at 4 and 14 days post-infection, in comparison to mock-infected animals (0 d). Transcriptome profiling was performed by deep RNA sequencing, using the Illumina platform. The analysis of differentially expressed genes was performed to determine concomitant gene-specific temporal patterns of mRNA expression in the lungs after M. ovipneumoniae infection. We found 1048 differentially expressed genes (575 up-regulated, 473 down-regulated) when comparing transcriptomic data at 4 and 0 days post-infection, and 2823 (1362 up-regulated, 1461 down-regulated) when comparing 14 versus 0 days post-infection. Kyoto Encyclopedia of Genes and Genomes pathway analysis showed that the differentially expressed genes at 4 and 14 versus 0 days post-infection were enriched in 245 and 287 pathways, respectively, and the Toll-like receptor (TLR) signaling pathway was considered most closely related to MO infection (p < 0.01). Two pathways (LAMP-TLR2/TLR6-MyD88-MKK6-AP1-IL1B and LAMP-TLR8MyD88-IRF5-RANTES) were identified based on the TLR signaling pathway from differentially expressed genes related M. ovipneumoniae infection. Gene Ontology analysis showed that differentially expressed genes in different groups were enriched for 1580 and 4561 terms, where those most closely related to M. ovipneumoniae infection are positive regulators of inflammatory responses (p < 0.01). These results could aid in understanding how M. ovipneumoniae infection progresses in the lungs and may provide useful information regarding key regulatory pathways.
Assuntos
Pulmão/metabolismo , Pneumonia por Mycoplasma/patologia , Análise de Sequência de RNA/métodos , Doenças dos Ovinos/patologia , Transcriptoma , Animais , Regulação para Baixo , Mycoplasma ovipneumoniae/isolamento & purificação , Mycoplasma ovipneumoniae/patogenicidade , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/veterinária , RNA Mensageiro/química , RNA Mensageiro/metabolismo , Ovinos , Doenças dos Ovinos/genética , Doenças dos Ovinos/metabolismo , Transdução de Sinais/genética , Fatores de Tempo , Receptores Toll-Like/genética , Receptores Toll-Like/metabolismo , Regulação para CimaRESUMO
Human surfactant protein-A2 (hSP-A2) is a component of pulmonary surfactant that plays an important role in the lung's immune system by interacting with viruses, bacteria, and fungi to facilitate pathogen clearance and by downregulating inflammatory responses after an allergic challenge. Genetic variation in SP-A2 at position Gln223Lys is present in up to â¼30% of the population and has been associated with several lung diseases, such as asthma, pulmonary fibrosis, and lung cancer (M. M. Pettigrew, J. F. Gent, Y. Zhu, E. W. Triche, et al., BMC Med Genet 8:15, 2007, https://bmcmedgenet.biomedcentral.com/articles/10.1186/1471-2350-8-15; Y. Wang, P. J. Kuan, C. Zing, J. T. Cronkhite, et al., Am J Hum Genet 84:52-59, 2009, https://www.cell.com/ajhg/fulltext/S0002-9297(08)00595-8). Previous work performed by our group showed differences in levels of SP-A binding to non-live mycoplasma membrane fractions that were dependent on the presence of a lysine (K) or a glutamine (Q) at amino acid position 223 in the carbohydrate region of SP-A2. On the basis of these differences, we have derived 20-amino-acid peptides flanking this region of interest in order to test the ability of each to regulate various immune responses to live Mycoplasma pneumoniae in SP-A knockout mice and RAW 264.7 cells. In both models, the 20-mer containing 223Q significantly decreased both tumor necrosis factor alpha (TNF-α) mRNA levels and protein levels in comparison to the 20-mer containing 223K during M. pneumoniae infection. While neither of the 20-mer peptides (223Q and 223K) had an effect on p38 phosphorylation during M. pneumoniae infection, the 223Q-20mer peptide significantly reduced NF-κB p65 phosphorylation in both models. Taken together, our data suggest that small peptides derived from the lectin domain of SP-A2 that contain the major allelic variant (223Q) maintain activity in reducing TNF-α induction during M. pneumoniae infection.
Assuntos
Anti-Inflamatórios/farmacologia , Interações entre Hospedeiro e Microrganismos/imunologia , Mycoplasma pneumoniae/imunologia , Peptídeos/farmacologia , Pneumonia por Mycoplasma/tratamento farmacológico , Proteína A Associada a Surfactante Pulmonar/imunologia , Fator de Necrose Tumoral alfa/imunologia , Adesinas Bacterianas/genética , Adesinas Bacterianas/imunologia , Animais , Anti-Inflamatórios/síntese química , Modelos Animais de Doenças , Regulação da Expressão Gênica , Interações entre Hospedeiro e Microrganismos/genética , Humanos , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Mycoplasma pneumoniae/efeitos dos fármacos , Mycoplasma pneumoniae/patogenicidade , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/microbiologia , Peptídeos/síntese química , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Pneumonia por Mycoplasma/microbiologia , Domínios Proteicos , Proteína A Associada a Surfactante Pulmonar/química , Proteína A Associada a Surfactante Pulmonar/deficiência , Proteína A Associada a Surfactante Pulmonar/genética , Células RAW 264.7 , RNA Mensageiro/genética , RNA Mensageiro/imunologia , Transdução de Sinais , Fator de Transcrição RelA/genética , Fator de Transcrição RelA/imunologia , Fator de Necrose Tumoral alfa/antagonistas & inibidores , Fator de Necrose Tumoral alfa/genética , Proteínas Quinases p38 Ativadas por Mitógeno/genética , Proteínas Quinases p38 Ativadas por Mitógeno/imunologiaRESUMO
Major histocompatibility complex (MHC) polymorphisms are associated with animal and human diseases. However, only a few studies have reported an association between MHC polymorphisms and mycoplasma ovipneumonia (MO). In the present study, three resistance/susceptibility genotypes associated with MO were identified by polymerase chain reaction-restriction fragment length polymorphism genotyping, assessing the clinical and pathological features, and examining the immune factors. The current results showed that MvaI bb and HaeIII ee were dominant genotypes in the susceptible Hu population, while MO-resistant populations, Dorper and D 9 H hybrids, were dominated by the MvaI cc and HaeIII dd genotypes, suggesting that MvaI cc and HaeIII dd genotypes might be associated with the trait of MO resistance. Further, the clinical symptoms and pathological morphology in the susceptibility group infected with MO were more severe than those in the resistant groups infected similarly. The data on the changes in the immune factor responses were utilized to deduce the molecular mechanism underlying the MO resistance/susceptibility. The results showed that the susceptible genotypes promote the inflammatory responses by inducing a high expression of TNFa, IFNc, IL-4, IL-6, and IL-1b, while the resistant genotypes inhibit the inflammatory response by increasing the expression of IL-2 and IL-10 significantly. This finding would provide the theoretical guidance for propagating sheep breeds that are highly resistant to MO.
Assuntos
Complexo Principal de Histocompatibilidade/genética , Mycoplasma ovipneumoniae , Pneumonia por Mycoplasma/veterinária , Doenças dos Ovinos/genética , Doenças dos Ovinos/imunologia , Animais , Citocinas/metabolismo , Suscetibilidade a Doenças/veterinária , Éxons , Frequência do Gene , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Pulmão/patologia , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Pneumonia por Mycoplasma/patologia , Polimorfismo de Fragmento de Restrição/imunologia , Ovinos , Doenças dos Ovinos/microbiologia , Doenças dos Ovinos/patologiaRESUMO
BACKGROUND: Extensive studies have focused on the diagnosis and treatment of Mycoplasma pneumoniae infection; however, rare studies investigated the posttreatment conditions. We analyzed the carrying status of M. pneumoniae in the respiratory tract of children before and after treatment. METHODS: Ninety-two children with M. pneumoniae pneumonia were included in this study. Clinical data were obtained from each patient, and pharyngeal swab sampling was performed at preliminary diagnosis and discharge. Real-time PCR and dilution quantitative culture were utilized to determine the DNA quantification and number of viable M. pneumoniae from samples collected upon preliminary diagnosis and discharge. RESULTS: All the 92 cases showed DNA positivity upon preliminary diagnosis, serum IgM antibody was detected in 80 patients, and positivity of M. pneumoniae culture was observed in 82 cases. Upon discharge, the M. pneumoniae nucleotide and culture positivity were detected in 87 and 49 cases, respectively. The content of viable M. pneumoniae was 10-104 CCU/mL and 10-102 CCU/mL in the preliminary diagnosis samples and discharge samples, respectively. CONCLUSIONS: Real-time PCR was rapid and effective for the qualitative diagnosis of M. pneumoniae at the early stage, but it cannot be used to evaluate the prognosis of patients with M. pneumoniae infection. Quantitative analysis for M. pneumoniae DNA could not directly reflex the viable strain content.
Assuntos
Anticorpos Antibacterianos/sangue , Imunoglobulina M/sangue , Mycoplasma pneumoniae , Faringe/microbiologia , Pneumonia por Mycoplasma , Reação em Cadeia da Polimerase em Tempo Real , Adolescente , Criança , Pré-Escolar , DNA Bacteriano/genética , DNA Bacteriano/metabolismo , Feminino , Humanos , Masculino , Mycoplasma pneumoniae/genética , Mycoplasma pneumoniae/crescimento & desenvolvimento , Mycoplasma pneumoniae/isolamento & purificação , Pneumonia por Mycoplasma/sangue , Pneumonia por Mycoplasma/diagnóstico , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/microbiologiaRESUMO
In the present study, we examined the function of microRNA (miR)146a5p in patients with refractory Mycoplasma pneumoniae pneumonia. In brief, the expression of miR146a5p was reduced in patients with refractory Mycoplasma pneumoniae pneumonia. Downregulation of miR146a5p reduced inflammation in an in vitro model of refractory Mycoplasma pneumoniae pneumonia, whilst overexpression of miR146a5p promoted inflammation. Downregulation of miR146a5p induced the protein expression of ATPbinding cassette subfamily G member 1 (ABCG1) and interleukin 1 receptorassociated kinase 1 (IRAK1), while suppressed expression was observed of the aforementioned proteins following overexpression of miR146a5p in an in vitro model of refractory Mycoplasma pneumoniae pneumonia. The administration of small interfering RNA against RXR or IRAK1 attenuated the effects of miR146a5p on inflammation in an in vitro model of refractory Mycoplasma pneumoniae pneumonia. Collectively, these results suggested that miR146a5p reduced ABCG1 expression in refractory Mycoplasma pneumoniae pneumonia via downregulation of IRAK1.
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Membro 1 da Subfamília G de Transportadores de Cassetes de Ligação de ATP/genética , Quinases Associadas a Receptores de Interleucina-1/genética , MicroRNAs/genética , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/genética , Células A549 , Criança , Pré-Escolar , Feminino , Regulação da Expressão Gênica/genética , Humanos , Lactente , Inflamação/genética , Inflamação/microbiologia , Inflamação/patologia , Masculino , MicroRNAs/farmacologia , Mycoplasma pneumoniae/patogenicidade , Pneumonia por Mycoplasma/microbiologia , Pneumonia por Mycoplasma/patologia , RNA Interferente Pequeno/genética , Receptores X de Retinoides/antagonistas & inibidores , Receptores X de Retinoides/genéticaRESUMO
Interleukin 1 receptor-like 1 (IL1RL1), also known as suppression of tumorigenicity 2 (ST2), is the receptor for interleukin 33 (IL-33) and has been increasingly studied in type 2 inflammation. An increase in airway IL-33/ST2 signaling in asthma has been associated with eosinophilic inflammation, but little is known about the role of ST2 in neutrophilic inflammation. Airway Mycoplasma pneumoniae and human rhinovirus (HRV) infections are linked to neutrophilic inflammation during acute exacerbations of asthma. However, whether ST2 contributes to M. pneumoniae- and HRV-mediated airway inflammation is poorly understood. The current study sought to determine the functions of ST2 during airway M. pneumoniae or HRV infection. In cultured normal human primary airway epithelial cells, ST2 overexpression (OE) increased the production of neutrophilic chemoattractant IL-8 in the absence or presence of M. pneumoniae or HRV1B infection. ST2 OE also enhanced HRV1B-induced IP-10, a chemokine involved in asthma exacerbations. In the M. pneumoniae-infected mouse model, ST2 deficiency, in contrast to sufficiency, significantly reduced the levels of neutrophils following acute (≤24 h) infection, while in the HRV1B-infected mouse model, ST2 deficiency significantly reduced the levels of proinflammatory cytokines KC, IP-10, and IL-33 in bronchoalveolar lavage (BAL) fluid. Overall, ST2 overexpression in human epithelial cells and ST2 sufficiency in mice increased the M. pneumoniae and HRV loads in cell supernatants and BAL fluid. After pathogen infection, ST2-deficient mice showed a higher level of the host defense protein lactotransferrin in BAL fluid. Our data suggest that ST2 promotes proinflammatory responses (e.g., neutrophils) to airway bacterial and viral infection and that blocking ST2 signaling may broadly attenuate airway infection and inflammation.
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Infecções por Enterovirus/imunologia , Enterovirus/fisiologia , Proteína 1 Semelhante a Receptor de Interleucina-1/imunologia , Mycoplasma pneumoniae/fisiologia , Pneumonia por Mycoplasma/microbiologia , Sistema Respiratório/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Líquido da Lavagem Broncoalveolar/microbiologia , Líquido da Lavagem Broncoalveolar/virologia , Enterovirus/genética , Infecções por Enterovirus/genética , Infecções por Enterovirus/virologia , Células Epiteliais/imunologia , Células Epiteliais/microbiologia , Células Epiteliais/virologia , Humanos , Proteína 1 Semelhante a Receptor de Interleucina-1/genética , Interleucina-33/genética , Interleucina-33/imunologia , Interleucina-8/genética , Interleucina-8/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Mycoplasma pneumoniae/genética , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/imunologia , Sistema Respiratório/microbiologia , Sistema Respiratório/virologiaRESUMO
BACKGROUND: Mycoplasma pneumoniae is a common cause of community-acquired pneumonia (CAP). This cross-sectional study aimed to determine the prevalence of macrolide-resistant M. pneumoniae strains in a convenience series of 234 adult hospitalised and nonhospitalised subjects with a diagnosis of CAP in January 2013 to April 2015 in South Italy. METHODS: Respiratory samples were subjected to real-time PCR. In M. pneumoniae-positive samples, domain V of 23S rRNA was sequenced to detect resistance-conferring point mutations. P1 major adhesion protein typing and multiple loci variable-number tandem repeat analysis (MLVA) were also performed. RESULTS: Of the 234 samples, 15 (6.4%) were positive for M. pneumoniae. Three of these had a macrolide-resistant genotype: two and one had A2063G and A2064G mutations, respectively. Fourteen of the 15 strains were subtyped: half had subtype 1 and half had subtype 2. Eight strains underwent MLVA profiling: one each had the J, A, and Z type. The remainder was unclassifiable. CONCLUSIONS: This novel discovery of macrolide-resistant M. pneumoniae strains in adults with CAP in Italy suggests that there may be increasing circulation of these strains in the population. To facilitate rapid optimization of the antibiotic strategy in Italy, macrolide resistance should be monitored by a surveillance system that is based on molecular methods.
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Infecções Comunitárias Adquiridas/tratamento farmacológico , Farmacorresistência Bacteriana/genética , Macrolídeos/uso terapêutico , Pneumonia por Mycoplasma/tratamento farmacológico , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Antibacterianos/efeitos adversos , Antibacterianos/uso terapêutico , Infecções Comunitárias Adquiridas/epidemiologia , Infecções Comunitárias Adquiridas/genética , Infecções Comunitárias Adquiridas/microbiologia , Feminino , Genótipo , Humanos , Itália/epidemiologia , Macrolídeos/efeitos adversos , Masculino , Testes de Sensibilidade Microbiana , Pessoa de Meia-Idade , Repetições Minissatélites/genética , Mutação , Mycoplasma pneumoniae/efeitos dos fármacos , Mycoplasma pneumoniae/genética , Mycoplasma pneumoniae/patogenicidade , Pneumonia por Mycoplasma/epidemiologia , Pneumonia por Mycoplasma/genética , Pneumonia por Mycoplasma/microbiologia , Adulto JovemRESUMO
We characterized 419 Mycoplasma pneumoniae isolates collected between 2011 and 2017 in Osaka prefecture of Japan. This analysis revealed high prevalence of macrolide-resistant M. pneumoniae (MRMP) in Osaka during 2011 and 2014 with annual detection rates of MRMP strains between 71.4% and 81.8%. However, in 2015 and after, the detection rate of MRMP decreased significantly and did not exceed 50%. Genotyping of the p1 gene of these isolates showed that most of MRMP strains harbored type 1 p1 gene. In contrast, strains expressing p1 gene type 2 or its variant were largely macrolide-susceptible M. pneumoniae (MSMP) strains. There was a strong correlation between p1 gene genotype and the presence of mutations conferring macrolide resistance in M. pneumoniae isolated in Osaka. These results indicate that lower incidence of MRMP strains in Osaka from 2015 was associated with the relative increase of p1 gene type 2 lineage strains. During these experiments, we also isolated three M. pneumoniae strains that showed irregular typing pattern in the polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the p1 gene. Two of these strains harbored new variants of type 2 p1 gene and were designated as type 2f and 2g. The remaining strain with an irregular typing pattern had a large deletion in the p1 operon.