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Steroidogenesis occurs locally in peripheral tissues and via adrenal and gonadal glands' biosynthesis. The C2C12 mouse myoblast cell line and rat skeletal muscles harbor a local steroidogenesis pathway for glucocorticoids, and corticosterone is biosynthesized from skeletal muscle cells. However, Cyp11a1 and StAR protein expressions are not observed in C2C12 cells or rat muscular tissues. In this context, this study investigated the relationship between DNA methylation and key steroidogenic genes. Bioinformatics analysis of methylated DNA immune precipitation showed that C2C12 myoblasts and myotubes did not have remarkable DNA methylated regions in the gene-body of Cyp11a1. However, a highly methylated region in the CpG island was detected in the intronic enhancer of Ad4BP/SF-1, known as the transcriptional factor for steroidogenic genes. After C2C12 myoblasts treatment with 5-aza-2-deoxycytidine, the gene expressions of Ad4BP/SF-1, Cyp11a1, and StAR were significantly time- and concentration-dependent upregulated. To clarify the contribution of Ad4BP/SF-1 on Cyp11a1 and StAR transcripts, we silenced Ad4BP/SF-1 during the 5-aza-2-deoxycytidine treatment in C2C12 myoblasts, resulting in significant suppression of both Cyp11a1 and StAR. Additionally, pregnenolone levels in the supernatants of C2C12 cells were enhanced by 5-aza-2-deoxycytidine treatment, whereas pregnenolone production by C2C12 myoblasts was significantly suppressed by Ad4BP/SF-1 knockdown. These results indicate that DNA methylation of Ad4BP/SF-1 might be involved in the downregulation of steroidogenic genes, such as Cyp11a1 and StAR in C2C12 myoblasts.
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Phage therapy has re-emerged in modern medicine as a robust antimicrobial strategy in response to the increasing prevalence of antimicrobial-resistant bacteria. However, bacterial resistance to phages can also arise via a variety of molecular mechanisms. In fact, several clinical studies on phage therapy have reported the occurrence of phage-resistant variants, representing a significant concern for the successful development of phage-based therapies. In this context, the fitness trade-offs between phage and antibiotic resistance have revealed new avenues in the field of phage therapy as a countermeasure against phage resistance. This strategy forces to restore the antibiotic susceptibility of antimicrobial-resistant bacteria as compensation for the development of phage resistance. Here, we present the key achievements of these fitness trade-offs, notably focusing on the enhancement of antibiotic sensitivity through the induction of large chromosomal deletions by bacteriophage infection. We also describe the challenges of this strategy that need to be overcome to promote favorable therapeutic outcomes and discuss future directions. The insights gained from the trade-offs between phage and antibiotic sensitivity will help maximize the potential of phage therapy for the treatment of infectious diseases.
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Mycoplasma bovis mastitisis highly contagious and disrupts lactation, posing a significant threat to the dairy industry. While the mammary gland's defence mechanism involves epithelial cells and mononuclear cells (MNC), their interaction with M. bovis remains incompletely understood. In this study, we assessed the immunological reactivity of bovine mammary epithelial cells (bMEC) to M. bovis through co-culture with MNC. Upon co-culture with MNC, the mRNA expression levels of interleukin (IL)-1ß, IL-6, IL-8 and tumor necrosis factor (TNF)-α in bMEC stimulated by M. bovis showed a significant increase compared to monoculture. Additionally, when stimulated with M. bovis, the culture supernatant exhibited significantly higher concentrations of IL-6 and interferon (IFN)-γ, while IL-1ß concentration tended to be higher in co-culture with MNC than in monoculture. Furthermore, the mRNA expression levels of toll-like receptor (TLR) 2 in bMEC stimulated with M. bovis tended to increase, and TLR4 significantly increased when co-cultured with MNC compared to monocultures. However, the surface expression levels in bMEC did not exhibit significant changes between co-culture and monoculture. Overall, our research indicates that the inflammatory response of bMEC is increased during co-culture with MNC, suggesting that the interaction between bMEC and MNC in the mammary gland amplifies the immune response to M. bovis in cows affected by M. bovis mastitis.
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Técnicas de Cocultivo , Células Epiteliales , Inmunidad Innata , Glándulas Mamarias Animales , Mastitis Bovina , Infecciones por Mycoplasma , Mycoplasma bovis , Animales , Bovinos , Mycoplasma bovis/inmunología , Mastitis Bovina/inmunología , Mastitis Bovina/microbiología , Femenino , Células Epiteliales/inmunología , Células Epiteliales/microbiología , Infecciones por Mycoplasma/veterinaria , Infecciones por Mycoplasma/inmunología , Infecciones por Mycoplasma/microbiología , Técnicas de Cocultivo/veterinaria , Glándulas Mamarias Animales/inmunología , Glándulas Mamarias Animales/microbiología , Citocinas/metabolismo , Citocinas/genética , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/microbiología , Células CultivadasRESUMEN
The transcription factor hypoxia-inducible factor 1α (HIF-1α) drives metabolic reprogramming in gliomas (GLs) under hypoxic conditions, promoting glycolysis for tumor development. Evofosfamide (EVO) releases a DNA-alkylating agent within hypoxic regions, indicating that it may serve as a hypoxia-targeted therapy. The aim of this study was to investigate the glycolytic metabolism and antitumor effects of EVO in a canine GL model. Our clinical data showed that overall survival was significantly decreased in GL dog patients with higher HIF-1α expression compared to that of those with lower HIF-1α expression, and there was a positive correlation between HIF-1α and pyruvate dehydrogenase kinase 1 (PDK1) expression, suggesting that glycolytic activity under hypoxia conditions may contribute to poor outcomes in canine GL. Our glycolysis assay tests showed that the glycolytic ATP level was higher than the mitochondrial ATP level in three types of canine GL cell lines by activating the HIF-1 signal pathway under hypoxia conditions, resulting in an overall increase in total cellular ATP production. However, treatment with EVO inhibited the glycolytic ATP level in the GL cell lines under hypoxia conditions by targeting HIF-1α-positive cells, leading to decrease in total cellular ATP production. Our in vivo tests showed that EVO significantly reduced tumor development compared to controls and temozolomide in murine GL models. A metabolic analysis demonstrated that EVO effectively suppressed glycolytic metabolism by eliminating HIF-1α-positive cells, suggesting that it may restore metabolism in canine GLs. The evidence presented here supports the favorable preclinical evaluation of EVO as a potential improvement in cancer metabolism.
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Bacteriophages are expected to be therapeutic agents against infectious diseases. Streptococcus mutans are involved in dental plaque formation related to dental caries and periodontitis. In S. mutans, lytic bacteriophages have been isolated previously, but the isolation of temperate bacteriophage has not been reported although their presence in the genome has been confirmed. Here, we report the isolation of temperate bacteriophage, φKSM96, from S. mutans. φKSM96 has a circular DNA 39,820 bp long and reveals Siphoviridae morphology. φKSM96 shows a broad range of susceptibility against S. mutans strains with different serotypes. By the addition of φKSM96, S. mutans growth and biofilm formation were significantly inhibited. In cocultures of S. mutans with other bacterial species, the proportion of S. mutans significantly decreased in the presence of φKSM96. In summary, φKSM96 shows selective anti-S. mutans activity. The isolation of temperate bacteriophage is important for future genetic manipulation to create more efficient bacteriophages.
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In recent decades, phage therapy has been overshadowed by the widespread use of antibiotics in Western countries. However, it has been revitalized as a powerful approach due to the increasing prevalence of antimicrobial-resistant bacteria. Although bacterial resistance to phages has been reported in clinical cases, recent studies on the fitness trade-offs between phage and antibiotic resistance have revealed new avenues in the field of phage therapy. This strategy aims to restore the antibiotic susceptibility of antimicrobial-resistant bacteria, even if phage-resistant variants develop. Here, we summarize the basic virological properties of phages and their applications within the context of antimicrobial resistance. In addition, we review the occurrence of phage resistance in clinical cases, and examine fitness trade-offs between phage and antibiotic sensitivity, exploring the potential of an evolutionary fitness cost as a countermeasure against phage resistance in therapy. Finally, we discuss future strategies and directions for phage-based therapy from the aspect of fitness trade-offs. This approach is expected to provide robust options when combined with antibiotics in this era of phage 're'-discovery.
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Infecciones Bacterianas , Bacteriófagos , Terapia de Fagos , Humanos , Bacteriófagos/genética , Antibacterianos/farmacología , Antibacterianos/uso terapéutico , Terapia de Fagos/métodos , Infecciones Bacterianas/terapia , BacteriasRESUMEN
Fasting induces metabolic changes in muscles, which are differentiated by muscle fiber type. In this study, the mechanism of fasting-induced muscle atrophy in rats was examined to determine the differences between muscle fiber types in energy production. Fasting for 96 h did not alter the weight of the soleus (SOL), a fiber type I muscle, but did significantly reduce the weight of gastrocnemius (GM), a fiber type II muscle. GM, SOL and blood pregnenolone and testosterone levels decreased under fasting, which induced energy deprivation, whereas corticosterone (CORT) levels significantly increased. However, the expression of 3ß-HSD and P45011ß in GM was unaffected by fasting. The decrease in GM weight may be due to decreased levels of testosterone and reduced synthesis of mammalian target of rapamycin (mTOR). Significant increases in CORT both GM and SOL were associated with increases in the amount of branched-chain amino acids available for energy production. However, decreased levels of mTOR and IGF1 and increased levels of CORT and IL-6 in SOL suggest that GM proteolysis was followed by SOL proteolysis for additional energy production. In conclusion, IGF1 levels decreased significantly in SOL, whereas those of IL-6 significantly increased in SOL and blood but decreased in GM. Blood branched-chain amino acids (BCAA) levels were unaffected due to fasting, whereas an increase was noted in the levels of BCAA in GM and SOL. These results show that fasting for 96 h restricts energy supply, producing fast-twitch muscle atrophy followed by slow-twitch muscle atrophy.
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Interleucina-6 , Fibras Musculares Esqueléticas , Ratas , Masculino , Animales , Interleucina-6/metabolismo , Fibras Musculares Esqueléticas/metabolismo , Atrofia Muscular/metabolismo , Músculo Esquelético/metabolismo , Ayuno , Aminoácidos de Cadena Ramificada/metabolismo , Testosterona/metabolismo , Serina-Treonina Quinasas TOR/metabolismo , Mamíferos/metabolismoRESUMEN
The rapid identification of specific bacterial pathogens in bovine mastitis is crucial for appropriate antimicrobial treatment. Sequencing of 16S rRNA gene amplicons is a proven, useful strategy for diagnosing bacterial infections. In this study, the use of 16S rRNA analysis with nanopore sequencer for the rapid identification of causative bacteria in bovine mastitis, was evaluated. DNA was extracted from 122 milk samples from cattle with suspected mastitis based on clinical symptoms. 16S rRNA gene amplicon sequencing was conducted using a nanopore sequencer. The efficacy of bacterial identification was verified by comparison with conventional culture methods. Nanopore sequencing identified the causative bacteria with high accuracy within approximately 6 h from the time of sample collection. When the major causative bacteria of bovine mastitis (Escherichia coli, Streptcoccus uberis, Klebsiella pneumoniae, and Staphylococcus aureus) were detected by nanopore sequencing, 98.3% of the results were consistent with identification through conventional culturing methods. 16S rRNA gene analysis using a nanopore sequencer enabled the rapid and accurate identification of bacterial species in bovine mastitis.
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Enfermedades de los Bovinos , Mastitis Bovina , Secuenciación de Nanoporos , Nanoporos , Infecciones Estafilocócicas , Femenino , Animales , Bovinos , ARN Ribosómico 16S/genética , Mastitis Bovina/diagnóstico , Mastitis Bovina/microbiología , Secuenciación de Nanoporos/veterinaria , Bacterias/genética , Infecciones Estafilocócicas/veterinaria , Leche/microbiologíaRESUMEN
This study aimed to examine the antimicrobial susceptibility of bovine mastitis pathogens in Japan and develop criteria for testing antimicrobial susceptibility using the simplified agar disk diffusion (ADD) method that is currently being used in clinical practice. Milk samples from 1,349 dairy cows with clinical mastitis were collected and cultured. The minimum inhibitory concentrations (MICs) of the antimicrobials were determined for 504 strains of 28 bacteria. Of the gram-positive bacteria, most Staphylococcus spp. were susceptible to penicillin G (PCG), kanamycin (KM), oxytetracycline (OTC), cefazolin (CEZ), pirlimycin, enrofloxacin, and marbofloxacin. Streptococcus spp. and Trueperella pyogenes showed resistance to OTC and KM. Most gram-negative bacteria were resistant to OTC and CEZ and particularly susceptible to fluoroquinolones. To develop the criteria for a disk diffusion test of the simplified ADD method, the relationships between MICs and diameters of inhibition zones (DIZs) were analyzed and compared with the conventional method. The susceptibility breakpoints of several antimicrobials were lower for both gram-positive and gram-negative bacteria. Particularly for gram-positive bacteria, the application of the new criteria lowers the breakpoint for PCG, suggesting that the use of PCG instead of CEZ may increase. The results suggest that use of these criteria for the simplified ADD method may lead to appropriate antimicrobial choice and consequently the appropriate use of antimicrobials in clinical practice.
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Antiinfecciosos , Enfermedades de los Bovinos , Mastitis Bovina , Femenino , Animales , Bovinos , Antibacterianos/farmacología , Agar , Mastitis Bovina/microbiología , Japón , Bacterias Grampositivas , Bacterias Gramnegativas , Cefazolina , Antiinfecciosos/farmacología , Pruebas de Sensibilidad Microbiana/veterinaria , Farmacorresistencia BacterianaRESUMEN
Staphylococcus virus ΦSA012 has a wide host range and efficient lytic activity. Here, we assessed the biological stability of ΦSA012 against temperature, freeze-thawing, and pH to clinically apply the phage. In addition, inoculation of ΦSA012 through i.p. and i.v. injections into mice revealed that phages were reached the limit of detection in serum and accumulated notably spleens without inflammation at 48 h post-inoculation. Furthermore, inoculation of ΦSA012 through s.c. injections in mice significantly induced IgG, which possesses neutralizing activity against ΦSA012 and other Staphylococcus viruses, ΦSA039 and ΦMR003, but not Pseudomonas viruses ΦS12-3 and ΦR18 or Escherichia viruses T1, T4, and T7 in vitro. Immunoelectron microscopic analysis showed that purified anti-phage IgG recognizes the long-tail fiber of staphylococcus viruses. Although S. aureus inoculation resulted in a 25% survival rate in a mouse i.p. model, ΦSA012 inoculation (i.p.) improved the survival rate to 75%; however, the survival rate of ΦSA012-immunized mice decreased to less than non-immunized mice with phage i.v. injection at a MOI of 100. These results indicated that ΦSA012 possesses promise for use against staphylococcal infections but we should carefully address the appropriate dose and periods of phage administration. Our findings facilitate understandings of staphylococcus viruses for phage therapy.
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Terapia de Fagos , Infecciones Estafilocócicas , Ratones , Animales , Terapia de Fagos/métodos , Fagos de Staphylococcus/ultraestructura , Staphylococcus aureus , Staphylococcus , Infecciones Estafilocócicas/terapia , Myoviridae/ultraestructura , Inmunoglobulina GRESUMEN
Bovine pneumonia is a disease that causes significant economic losses in livestock industries and is vital for animal welfare. The whole-genome sequence of Pasteurella multocida strain Pm1, isolated from a calf suffering from pneumonia in Japan, is reported here.
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Bisphenol A (BPA) has been shown to exhibit various toxic effects, including the induction of reproductive disorders. Generally, BPA is converted to conjugated metabolites, leading to bio-inactivation. On the other hand, the toxicity of conjugated metabolites is not fully understood. Notably, the placenta develops the sulfate-sulfatase pathway, which transports and reactivates sulfated steroids. Therefore, we investigated the potential adverse effects of the BPA-sulfate conjugate (BPA-S) on human placenta-derived BeWo cytotrophoblasts. In the present study, high-concentration BPA-S (100 µM) induced significant inhibition of BeWo growth, with effects similar to those seen with unconjugated BPA (100 µM and 100 nM). This growth inhibition was restored by treatment of the cells with an inhibitor of the organic anion-transporting peptides (OATPs) (bromosulphophthalein) or with a sulfatase (STS) inhibitor (STX64). BeWo exhibits expression of the genes encoding OATP1A2 and OATP4A1 as known sulfated steroid transporters and STS, suggesting that BPA-S suppresses cell growth activity via the sulfate-sulfatase pathway. In addition, cell cycle analysis revealed that BPA-S (100 µM) increased the fraction of cytotrophoblasts in the G2/M phases and significantly decreased the accumulation of the transcript encoding Aurora kinase A (AURKA), which is a critical regulator of cellular division. These results suggested that BPA-S triggers cell cycle arrest and inhibits proliferation of BeWo cytotrophoblasts by decreased AURKA, an effect that is mediated by the sulfate-sulfatase pathway. Overall, these findings provide insights into the reactivation of sulfated endocrine-disrupting chemicals and subsequent adverse effects.
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Aurora Quinasa A , Trofoblastos , Compuestos de Bencidrilo/toxicidad , Ciclo Celular , División Celular , Femenino , Humanos , Fenoles , Embarazo , Sulfatos/metabolismo , Sulfatos/farmacología , Trofoblastos/metabolismoRESUMEN
Although Escherichia coli is a commensal bacterium of the bovine vaginal microbiota, it is an important pathogenic bacterium that causes diseases of the reproductive tract and sub-fertility. Recent studies have focused on virulence factors (VFs) of intrauterine E. coli; however, actual endometrial VFs have not been clearly identified. The purpose of this study was to identify the VFs of E. coli associated with clinical metritis and endometritis. Thirty-two strains of E. coli and four mixed Trueperella pyogenes (TP) strains were detected in the uterus of 19 Holstein dairy cows with obvious clinical signs (between 8 and 66 days postpartum). The presence of six E. coli VFs (fimH, fyuA, kpsMTII, hra1, csgA, and astA) was examined by PCR, and clinical signs and reproductive performance (mixed TP, the percentage of polymorphonuclear neutrophils [PMN%], days to uterine involution, etc.) were evaluated. Four VFs (fimH, hra1, csgA, and astA) were detected in all E. coli strains, whereas fyuA and kpsMTII were detected in 94% and 50% of strains, respectively. Cows with E. coli strains harboring kpsMTII exhibited significantly severe clinical scores (vaginal discharge score, PMN%, uterine involution), suggesting that kpsMTII is a key VF for progression of clinical metritis and endometritis. In the present study, we clearly identified six VFs associated with clinical metritis and endometritis. In addition, E. coli strains with kpsMTII probably play a crucial role in the progression of clinical metritis and endometritis.
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Enfermedades de los Bovinos , Endometritis , Infecciones por Escherichia coli , Animales , Bovinos , Enfermedades de los Bovinos/patología , Endometritis/microbiología , Endometritis/veterinaria , Escherichia coli , Infecciones por Escherichia coli/microbiología , Infecciones por Escherichia coli/veterinaria , Femenino , Periodo Posparto , Útero/patología , Factores de VirulenciaRESUMEN
Bacteriophages are viruses that specifically infect bacteria and are classified as either virulent phages or temperate phages. Despite virulent phages being promising antimicrobial agents due to their bactericidal effects, the implementation of phage therapy depends on the availability of virulent phages against target bacteria. Notably, virulent phages of Streptococcus gordonii, which resides in the oral cavity and is an opportunistic pathogen that can cause periodontitis and endocarditis have previously never been found. We thus attempted to isolate virulent phages against S. gordonii. In the present study, we report for the first time a virulent bacteriophage against S. gordonii, ΦSG005, discovered from drainage water. ΦSG005 is composed of a short, non-contractile tail and a long head, revealing Podoviridae characteristics via electron microscopic analysis. In turbidity reduction assays, ΦSG005 showed efficient bactericidal effects on S. gordonii. Whole-genome sequencing showed that the virus has a DNA genome of 16,127 bp with 21 coding sequences. We identified no prophage-related elements such as integrase in the ΦSG005 genome, demonstrating that the virus is a virulent phage. Phylogenetic analysis indicated that ΦSG005 forms a distinct clade among the streptococcus viruses and is positioned next to streptococcus virus C1. Molecular characterization revealed the presence of an anti-CRISPR (Acr) IIA5-like protein in the ΦSG005 genome. These findings facilitate our understanding of streptococcus viruses and advance the development of phage therapy against S. gordonii infection.
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Genoma Viral , Filogenia , Fagos de Streptococcus/genética , Fagos de Streptococcus/patogenicidad , Streptococcus gordonii/virología , Repeticiones Palindrómicas Cortas Agrupadas y Regularmente Espaciadas , Terapia de Fagos , Fagos de Streptococcus/clasificación , Virulencia , Secuenciación Completa del GenomaRESUMEN
Pseudomonas aeruginosa, which causes chronic infections, has demonstrated rapid acquisition of antimicrobial resistance (AMR). Therefore, bacteriophages have received significant attention as promising antimicrobial agents; however, previous trials have reported the occurrence of phage-resistant variants. P. aeruginosa has lost large chromosomal fragments via evolutionary selection by MutL. Mutants lacking galU and hmgA, located in close proximity, exhibit phage resistance and brown color phenotype since hmgA encodes a homogentisic acid metabolic enzyme and deletion of galU results in a lack of O-antigen polysaccharide and absence of the phage receptor. In the present study, we evaluated this mechanism for controlling phage resistance in P. aeruginosa veterinary isolate Pa12. Phage-resistant Pa12 brown mutants (brmts) with galU and hmgA deletions were isolated. Whole-genome sequencing of the brmts revealed that regions 148-27 kbp upstream and 261-110 kbp downstream of galU were largely deleted from the Pa12 parental chromosome. Furthermore, all of these fluctuating deleted sequences in Pa12 brmts, tentatively designated bacteriophage-induced galU deficiency (BigD) regions, harbor multi-drug efflux system genes (mexXY). Minimum inhibitory concentration (MIC) assays demonstrated that brmts altered sensitivity to antibiotics and exhibited increased levofloxacin sensitivity compared with the Pa12 parent. Orbifloxacin and enrofloxacin also effectively suppressed growth of the Pa12 brmts, suggesting that MexXY, which mediates quinolone efflux and is located in the BigD region, might be associated with restoration of fluoroquinolone sensitivity. Our findings indicate that AMR-related genes in the BigD region could produce trade-off effects between phages and drug sensitivity and thereby contribute to a potential strategy to control and prevent phage-resistant variants in phage therapy.
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Bacteriófagos , Proteínas HMGA , Terapia de Fagos , Antibacterianos/farmacología , Bacteriófagos/genética , Fluoroquinolonas/metabolismo , Fluoroquinolonas/farmacología , Proteínas HMGA/metabolismo , Pseudomonas aeruginosa/genéticaRESUMEN
Pseudomonas aeruginosa causes various opportunistic infections in animals. Here, we report the complete genome sequence of P. aeruginosa strain Pa12, a fluoroquinolone-resistant isolate from a canine skin lesion. To expand the molecular antimicrobial characteristics of the isolate, the whole Pa12 genome was sequenced and assembled via long- and short-read platforms.
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Mycoplasma bovis (M. bovis) is a significant worldwide pathogen of cattle. Neutrophils have an important role in the innate immune response during infection with M. bovis. However, even though neutrophils accumulate in M. bovis infection, the interaction of M. bovis and neutrophils has not been fully elucidated. We attempted to elucidate the innate immune response of neutrophils stimulated with M. bovis and evaluate the transcriptome and functional analysis of bovine neutrophils stimulated with M. bovis. Proinflammatory cytokines, such as inducible nitric oxide (iNOS), which was the most increased gene in transcriptome analysis, were increased in quantitative polymerase chain reaction analysis of bovine neutrophils stimulated with live or heat-killed M. bovis. Nitric oxide and intracellular reactive oxygen species production of neutrophils stimulated with M. bovis was significantly increased. Neutrophils stimulated with M. bovis showed an increased ratio of nonapoptotic cell death compared to unstimulated controls. We demonstrated that neutrophil extracellular traps (NETs) formation was not recognized in neutrophils stimulated with live M. bovis. However, heat-killed M. bovis induced NETs formation. We also showed the interaction with M. bovis and bovine neutrophils regarding proinflammatory cytokine gene expression and functional expression related to NETs formation. Live and killed M. bovis induced innate immune responses in neutrophils and had the potential to induce NETs formation, but live M. bovis escaped NETs.
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Enfermedades de los Bovinos/inmunología , Trampas Extracelulares/metabolismo , Expresión Génica/inmunología , Inmunidad Innata , Infecciones por Mycoplasma/veterinaria , Mycoplasma bovis/fisiología , Neutrófilos/inmunología , Animales , Bovinos , Enfermedades de los Bovinos/genética , Enfermedades de los Bovinos/microbiología , Trampas Extracelulares/microbiología , Infecciones por Mycoplasma/genética , Infecciones por Mycoplasma/inmunología , Infecciones por Mycoplasma/microbiologíaRESUMEN
The immune response during the onset of coliform mastitis in vaccinated cows was investigated by measuring lactoferrin (LF), interleukin-8 (IL-8), and interleukin-1ß (IL-1ß) concentrations and somatic cell counts in 28 milk samples at the onset of acute coliform mastitis (ACM) and 73 milk samples at the onset of peracute coliform mastitis (PCM). Vaccinated ACM, unvaccinated ACM, and vaccinated PCM showed significantly higher values for LF and IL-1ß levels than unvaccinated PCM (p < .01). The IL-8 concentration was lower in vaccinated PCM than in unvaccinated PCM (p < .05). There was no significant difference in somatic cell counts for each parameter. There were no significant differences in the parameters between vaccinated and unvaccinated ACM cows, or vaccinated ACM and PCM cows. From the above results, it is suggested that mastitis vaccination improved the early immune response, particularly at the onset of PCM, and played a large role in host defense against the initial infection.
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Vacunas Bacterianas/inmunología , Infecciones por Enterobacteriaceae , Enterobacteriaceae/inmunología , Mastitis Bovina/inmunología , Mastitis Bovina/microbiología , Vacunación/veterinaria , Animales , Bovinos , Recuento de Células , Femenino , Interleucina-1beta/metabolismo , Interleucina-8/metabolismo , Lactoferrina/metabolismo , Leche/citología , Leche/inmunología , Leche/metabolismoRESUMEN
Mycoplasma bovis causes chronic arthritis in cattle, accompanied by a severe inflammatory reaction of the joints. Recent studies demonstrated that M. bovis can invade bovine non-phagocytic cells, but the mechanism of M. bovis internalization in the cells remains unclear. In this study, we examined the mechanism by which M. bovis invades synovial cells, including the pathway of cell invasion. Using fluorescence and electron microscopy, multiple M. bovis were observed to adhere to and be internalized in cultured bovine synovial cells. The number of M. bovis colocalized with clathrin heavy chain (CLTC) per cell was significantly higher than the number of M. bovis colocalized with caveolin-1 (Cav-1). The internalized ratio of M. bovis in synovial cells treated with clathrin-dependent endocytosis inhibitor and small interfering RNA (siRNA) against CLTC was significantly lower than that in control cells. In contrast, the internalized ratio of M. bovis in synovial cells was unaffected by siRNA against Cav-1. These findings provide the first evidence that clathrin-dependent endocytosis is one of the major pathways by which M. bovis invades into synovial cells.