Search

Research Highlights: MG-HS regulates the expression of transcription factor STAT5.Transcription factor STAT5 can target miR-33-5p promoter element.MG-influenced STAT5 regulates miR-33-5p and its target gene expression.

Chick embryos are a valuable model for studying immunity and vaccines. Therefore, it is crucial to investigate the molecular mechanism of the Mycoplasma gallisepticum (MG)-induced immune response in chick embryos for the prevention and control of MG. In this study, we screened for downregulated let-7d microRNA in MG-infected chicken embryonic lungs to explore its involvement in the innate immune mechanism against MG. Here, we demonstrated that low levels of let-7d are a protective mechanism for chicken embryo primary type II pneumocytes (CP-II) in the presence of MG. Specifically, we found that depressed levels of let-7 in CP-II cells reduced the adhesion capacity of MG. This suppressive effect was achieved through the activated mitogen-activated protein kinase phosphatase 1 (MKP1) target gene and the inactivated mitogen-activated protein kinase (MAPK) pathway. Furthermore, MG-induced hyperinflammation and cell death were both alleviated by downregulation of let-7d. In conclusion, chick embryos protect themselves against MG infection through the innate immune molecule let-7d, which may result from its function as an inhibitor of the MAPK pathway to effectively mitigate MG adhesion, the inflammatory response and cell apoptosis. This study may provide new insight into the development of vaccines against MG., (© 2023. The Author(s).)

A disruption in the expression of gga-miR-365-3p was confirmed in the Mycoplasma gallisepticum (MG)-infected Chicken primary alveolar type II epithelial (CP-II) cells based on previous sequencing results, but the role it plays in the infection was unclear. In the present study, we demonstrate that MG evaded cellular host immunity via a gga-miR-365-3p/SOCS5-JAK/STATs negative feedback loop. Specifically, we found that at the initial stage of MG infection in cells, gga-miR-365-3p was rapidly increased and activated the JAK/STAT signaling pathway by inhibiting SOCS5, which induced the secretion of inflammatory factors and triggered immune response against MG infection. Over time, though, the infection progressed, MG gradually destroyed the immune defences of CP-II cells. In late stages of infection, MG escaped host immunity by reducing intracellular gga-miR-365-3p and inhibiting the JAK/STAT pathway to suppress the secretion of inflammatory factors and promote MG adhesion or invasion. These results revealed the game between MG and host cell interactions, providing a new perspective to gain insight into the pathogenic mechanisms of MG or other pathogens. Meanwhile, they also contributed to novel thoughts on the prevention and control of MG and other pathogenic infections, shedding light on the immune modulating response triggered by pathogen invasion and their molecular targeting., (© 2022. The Author(s).)

Mycoplasma gallisepticum (MG) is a pathogenic microorganism that causes chronic respiratory disease (CRD). MG infection has a serious negative impact on the poultry industry. Andrographolide (AG) is known to regulate immune responses, antimicrobial infections, and anti-inflammatory responses. However, the underlying molecular mechanisms of AG action in MG-infected chickens remain unclear. Hence, we constructed models of MG infection by using chickens and chicken macrophage-like (HD11) cells in vivo and in vitro, respectively. The results showed that AG significantly inhibited the mRNA and protein expression of the toxic adhesion protein pMGA1.2 in vivo and in vitro. Meanwhile, AG treatment significantly decreased the mRNA expression of pro-inflammatory such as interleukin-6 (IL-6) and interleukin- 1β (IL-1β), and increased the mRNA expression of an anti-inflammatory such as interleukin-10 (IL-10) and transforming growth factor beta (TGF-β) in vivo and in vitro. Furthermore, AG treatment down-regulated inflammasome NLRP3 and apoptosis genes caspase3 and caspase9, and up-regulated autophagy protein light chain 3 (LC3) by regulating the PI3K/Akt signaling pathway in vitro. Our results suggest that AG can reduce the expression of NLRP3 and alleviate the inflammatory response from MG infection by inducing autophagy, probably by modulating PI3K/Akt signaling pathway. This study demonstrates that AG can be used as a specific target to prevent and treat MG infection effectively., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Mycoplasma gallisepticum (MG) is the main pathogen of chronic respiratory disease (CRD), an infectious disease in chickens with high morbidity. Exosomal miRNAs are emerging as important regulators in host immune response to microbial invasion. Previously, we found that gga-miR-193a was significantly up-regulated in exosomes from MG-infected primary chicken type II pneumocytes (CP-IIs). Therefore, the purpose of this study was to investigate the role of exosomal gga-miR-193a in MG infection. Exosomes were isolated and identified via ultracentrifugation, transmission electron microscopy, and nanoparticle-tracking analysis. Real-time quantitative PCR and Western blot were used to detect the gene expression. Enzyme-linked immunosorbent assay was used to examine the levels of the inflammatory cytokines. CCK-8 and flow cytometry assays were applied to analyze the cell functions. The results showed that MG infection induced high expression of gga-miR-193a in exosomes from CP-IIs. Moreover, exosomes secreted by MG-infected CP-IIs could selectively transport gga-miR-193a into DF-1 cells. Exosomal gga-miR-193a internalized by DF-1 cells inhibited cell proliferation, promoted apoptosis, and increased interleukin-1β and tumor necrosis factor-α secretions by targeting the RAS/ERK signaling pathway. These results suggest that MG induced the secretion of gga-miR-193a by exosomes to damage the life activities of normal cells, which partially interpreted the mechanism of MG establishing systemic chronic infection in the body., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

High-mobility group box 1 (HMGB1), a member of damage-associated molecular patterns (DAMPs), is involved in the immune regulation of several infectious diseases. Mycoplasma gallisepticum (MG) infection is proved to cause an abnormal immune response, but the role of HMGB1 in MG-induced chronic respiratory disease (CRD) is unclear. In this study, we found that HMGB1 was released from the nucleus to the extracellular in macrophages upon infection with MG. Extracellular HMGB1 bound to TLR2 activating the NF-κB pathway triggering a severe inflammatory storm and promoting the progression of MG infection. More importantly, TLR4 could be activated by HMGB1 to trigger immune disorders after TLR2 was silenced. This disease process could be interrupted by ethyl pyruvate (EP) inhibition of HMGB1 release or glycyrrhizic acid (GA). Furthermore, treatment of MG-infected chickens with GA significantly alleviated immune organ damage. In conclusion, we demonstrate that HMGB1 is secreted extracellularly to form an inflammatory environment upon MG infection, triggering a further cellular inflammatory storm in a positive feedback approach. Blocking MG-induced HMGB1 release or suppression downstream of the HMGB1-TLR2/TLR4 axis may be a promising novel strategy for the treatment of CRD. Furthermore, this study may provide a theoretical reference for understanding non-LPS-activated TLR4 events.

Mycoplasma gallisepticum (MG) is the primary etiologic agent of chronic respiratory disease (CRD) in chickens. Respiratory tract inflammation and apoptosis are the main features of CRD. Andrographolide (Andro), a natural small molecule compound, is known for its excellent anti-pathogenic and anti-inflammatory properties. Hence, this study was to evaluate the anti-inflammation and anti-apoptosis effects of Andro as well as the underlying mechanism in the chicken lungs and primary alveolar type II epithelial cells (AEC II). Results showed Andro had no side effects on AEC II viability at concentrations below 200 μg/ml. Compared with the model group, terminal deoxynucleotidyl transferase-mediated dUTP nick endlabeling (TUNEL), western blot (WB), quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assays (ELISA) results showed Andro treatment significantly reduced apoptosis in the chicken lungs and AEC II, and down-regulated the expression levels of the protein of MG adhesin 1.2 (pMGA1.2), IL-1β, TNF-α, IL-6, Bax, Caspase 9 and Caspase 3, and up-regulated the expression levels of Bcl-2 and Bcl-xL in the chicken lungs, serum and AEC II (P ≤ 0.05). Moreover, Andro inhibited the MG-induced JAK/PI3K/AKT signal pathway activation in the chicken lungs and AEC II. Inhibiting of the JAK/PI3K/AKT signal pathway significantly alleviated MG-induced inflammation and apoptosis in the AEC II. Andro may exert an anti-inflammatory and anti-apoptotic effect by inhibiting the JAK/PI3K/AKT signal pathway in the chicken lungs and AEC II. In conclusion, Andro could act as a potential agent against MG infection by inhibiting the JAK/PI3K/AKT signal pathway and pMGA1.2 expression in the chickens., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Mycoplasma gallisepticum (MG) is one of the most important pathogens, that causes chronic respiratory disease (CRD) in chickens. Long non-coding RNAs (lncRNAs) are emerging as new regulators for many diseases and some lncRNAs can function as competing endogenous RNAs (ceRNAs) to regulate mRNAs by competitively binding to miRNAs. Here, we found that miR-33-5p was significantly up-regulated both in MG-infected chicken embryonic lungs and chicken embryo fibroblast cells (DF-1), and Lnc90386 negatively correlated with miR-33-5p. miR-33-5p, as a new regulator for MG infection, repressed apoptosis, inflammatory factors in DF-1 cells by targeting JNK1. Further analyses showed that Lnc90386 sponged miR-33-5p to weaken its inhibitory effect on JNK1, forming the ceRNA regulatory network. Furthermore, knockdown of Lnc90386 significantly inhibited apoptosis and inflammatory factors, and promoted DF-1 cells proliferation. However, co-treatment with miR-33-5p inhibitor and Lnc90386 siRNA showed that knockdown of Lnc90386 could partially eliminate the inhibiting effect of miR-33-5p inhibitor on inflammation, cell apoptosis and proliferation. In conclusion, Lnc90386 sponges miR-33-5p to defend against MG infection by inhibiting the JNK signaling pathway., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Sun, Wang, Zou, Wang, Wang and Peng.)

Mycoplasma gallisepticum (MG) is the primary pathogen of chronic respiratory diseases (CRDs) in chickens. In poultry production, antibiotics are mostly used to prevent and control MG infection, but the drug resistance and residue problems caused by them cannot be ignored. Glycyrrhizic acid (GA) is derived from licorice, a herb traditionally used to treat various respiratory diseases. Our study results showed that GA significantly inhibited the mRNA and protein expression of pMGA1.2 and GapA in vitro and in vivo. Furthermore, the network pharmacology study revealed that GA most probably resisted MG infection through the MAPK signaling pathway. Our results demonstrated that GA inhibited MG-induced expression of MMP2/MMP9 and inflammatory factors through the p38 and JUN signaling pathways, but not the ERK pathway in vitro. Besides, histopathological sections showed that GA treatment obviously attenuated tracheal and lung damage caused by MG invasion. In conclusion, GA can inhibit MG-triggered inflammation and apoptosis by suppressing the expression of MMP2/MMP9 through the JNK and p38 pathways and inhibit the expression of virulence genes to resist MG. Our results suggest that GA might serve as one of the antibiotic alternatives to prevent MG infection.

Mycoplasma gallisepticum (MG) is one of the most important pathogens that causes chronic respiratory disease (CRD) in chickens. Exosomes secreted from cells have been well demonstrated to deliver miRNAs to recipient cells to modulate cellular functions. The purpose of this study is to explore the underlying functions and mechanisms of exosomal miR-181a-5p in MG-HS infection. In this study, we found that miR-181a-5p expression in vivo and in vitro was significantly up-regulated after MG-HS infection. It was also upregulated in exosomes, which were derived from MG-HS-infected type-II pneumocytes cells (CP-II). In addition, exosomes secreted by MG-HS-infected CP-II were able to transfer miR-181a-5p to recipient chicken embryo fibroblast cells (DF-1), resulting in a significant upregulation of miR-181a-5p expression in recipient DF-1 cells. We further identified that Mg 2+ /Mn 2+ -dependent protein phosphatase 1B (PPM1B) was the target gene of miR-181a-5p. Overexpression of miR-181a-5p or knockdown of PPM1B activated the nuclear factor-κB (NF-κB) signaling pathway, whereas inhibition of miR-181a-5p and overexpression of PPM1B led to the opposite results. Besides, up-regulation of miR-181a-5p significantly increased the expression of toll-like receptor 2 (TLR2), myeloid differentiation factor 88 (MyD88), tumor necrosis factors alpha (TNF-α) and interleukin-1β (IL-1β), whereas inhibition of miR-181a-5p showed a contrary result. Up-regulation of miR-181a-5p promoted cell proliferation, cell cycle progression and inhibited apoptosis to resist MG-HS infection. Moreover, overexpression of miR-181a-5p significantly negative regulated the expression of Mycoplasma gallisepticum adhesin protein (pMGA1.2) by directly inhibiting PPM1B. Thus, we concluded that exosomal miR-181a-5p from CP-II cells activated the TLR2-mediated MyD88/NF-κB signaling pathways by directly targeting PPM1B to promote the expression of pro-inflammatory cytokines for defending against MG-HS infection in recipient DF-1 cells., (Copyright © 2021. Published by Elsevier Ltd.)

Mycoplasma gallisepticum (MG) is a major poultry pathogen that can induce Chronic Respiratory Disease (CRD) in chickens, causing serious economic losses in the poultry industry worldwide. Increasing evidence suggests that microRNAs (miRNAs) act as a vital role in resisting microbial pathogenesis and maintaining cellular mechanism. Our previous miRNAs sequencing data showed that gga-miR-223 expression level significantly decreased in MG-infected chicken lungs. The aim of this study was to reveal the role of gga-miR-223 in MG-induced CRD progression. We found that gga-miR-223 was remarkably down regulated and forkhead box O3 (FOXO3) was up-regulated in both MG-infected chicken embryos lungs and the chicken embryonic fibroblast cell line (DF-1) by qPCR. FOXO3 was verified as the target gene of gga-miR-223 through bioinformatics analysis and dual-luciferase reporter assay. Further studies showed that overexpressed gga-miR-223 could promote cell proliferation, cell cycle, and inhibit cell apoptosis by notably promoting the expression of cell cycle marker genes cyclin-dependent kinase 1 (CDK1), cyclin-dependent kinase 6 (CDK6) and Cyclin D1 (CCND1) and inhibiting the expression of apoptosis markers Bcl-2-like 11(BIM), FAS ligand (FASLG) and TNF-related apoptosis-inducing ligand (TRAIL). As expected, FOXO3 knockdown group got similar results. Overexpression of gga-miR-223 observably promoted cell multiplication, cell cycle progression, and inhibited apoptosis of MG-infected DF-1 cells, while inhibited gga-miR-223 had the opposite effect. Taken together, upon MG-infection, downregulated gga-miR-223 could decrease proliferation, cycle progression, and increase apoptosis through directly targeting FOXO3 to exert an aggravating MG-infectious effect., (Copyright © 2021. Published by Elsevier Ltd.)

Mycoplasma gallisepticum (MG) infection is the main cause of chronic respiratory disease (CRD) characterized by severe respiratory inflammation in chickens. Polydatin (PD) is a resveratrol glycoside isolated from Polygonum cuspidatum, which has prominent anti-inflammatory effect. The purpose of this study was to investigate the therapeutic effect of PD against MG-induced inflammation in chicken and its underlying mechanism. Histopathological analysis showed that PD treatment (15, 30, and 45 mg/kg) apparently alleviated MG-induced pathological changes of chicken embryonic lung. In chicken embryo fibroblast (DF-1) cells, PD treatment (15, 30, and 60 μg/mL) could effectively suppress MG propagation, promote MG-infected cell proliferation and cell cycle progress, and inhibit MG-induced cell apoptosis. ELISA and qPCR assays showed that PD treatment significantly suppressed the expression of interleukin-6 (IL-6), IL-1β and tumor necrosis factor-α (TNF-α) induced by MG both in vivo and in vitro. Besides, molecular studies indicated that the MG-induced levels of toll-like receptor-6 TLR6, myeloid differentiation-88 (MyD88) and nuclear factor κB (NF-κB) were significantly decreased by PD treatment. Moreover, immunofluorescence analysis showed that PD treatment restrained the MG-induced NF-κB-p65 nuclear translocation. Taken together, these results indicate the protective effects of PD against MG-induced inflammation injury in chicken were mainly by inhibiting the TLR6/MyD88/NF-κB pathway., (Copyright © 2020. Published by Elsevier Ltd.)

Mycoplasma gallisepticum (MG) is the primary etiological agent of chicken chronic respiratory disease (CRD), which mainly causes inflammatory damage of the host respiratory system. Previous studies suggest that puerarin (PUE) plays a pivotal regulatory role in inflammatory diseases, whereas the impacts of PUE on MG-induced inflammation remain unclear. This study investigated the effects of PUE on MG-HS infection in vitro and in vivo and indicated its potential therapeutic and preventive value. Experimental results showed that PUE significantly suppressed pMGA1.2 expression, promoted MG-infected cell proliferation and cell cycle process by reducing apoptosis. Histopathological examination of lung tissue showed severe histopathological lesions including thickened alveolar walls, narrowed alveolar cavity, and inflammatory cell infiltration in the MG-infected chicken group. However, PUE treatment significantly ameliorated MG-induced pathological damage in lung. Compared to the MG-infected group, PUE effectively inhibited the expression of MG-induced inflammatory genes, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), cytokines interleukin-6 (IL-6), toll-like receptor 6 (TLR6), myeloid differentiation primary response gene 88 (MyD88) and nuclear factor κB (NF-κB). Moreover, PUE dose-dependently inhibited MG-induced NF-κB p65 to enter the cell nucleus. In conclusion, our findings indicate that PUE treatment can efficiently inhibit MG-induced inflammatory response and apoptosis, and protect the lung from MG infection-induced damage by inhibiting the TLR6/MyD88/NF-κB signaling pathway activation. The study suggests that PUE may be a potential anti-inflammatory agent defense againstMGinfection in chicken., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Mycoplasma gallisepticum (MG) can target host cells and cause chronic respiratory disease (CRD) in chickens that is characterized by pMGA and concomitant. Although microRNAs (miRNAs) have been manifested are crucial regulatory noncoding RNAs with important effects on microbial pathogenesis and inflammatory response, how miRNAs regulate MG-induced inflammation remains to be discovered. The results showed that gga-miR-21 was up-regulated in MG-infected chicken embryonic lungs and MG infection of chicken embryo fibroblast cells (DF-1) compared with the control group. Overexpression of gga-miR-21 increased the inflammatory cytokines production, including tumor necrosis factor alpha (TNF-α), interleukin-1β (IL-1β), interleukin-6 (IL-6), and interleukin-8 (IL-8) after MG infection, knockdown of gga-miR-21 had thoroughly inverse effects. Gene expression data combined with bioinformatics analysis and luciferase reporter assays demonstrated that mitogen-activated protein kinase kinase kinase 1(MAP3K1) was a novel target of gga-miR-21. The inhibition of MAP3K1 by gga-miR-21 resulted in the accumulation of NF-κB in the nucleus, which in turn generate higher inflammatory cytokines. Furthermore, upregulation of gga-miR-21 significantly inhibited MG propagation and promoted MG-infected DF-1 cells proliferation by increasing the cell cycle progression and suppressing cell apoptosis. Our study provides evidence for proinflammatory effects of gga-miR-21 which is mediated at least in part by targeting MAP3K1 in the MG-infected DF-1 cells. gga-miR-21 activates MAPKs and NF-κB signaling pathways via targeting MAP3K1, and then promotes the production of inflammatory cytokines and cell proliferation by increasing the cell cycle progression and suppressing cell apoptosis to defend against MG infection., (Copyright © 2019 Elsevier B.V. All rights reserved.)

MicroRNAs (miRNAs) have been determined to be important regulators for pathogenic microorganism infection. However, it is largely unclear how miRNAs are triggered during pathogen infection. We previously reported that the up-regulation of gga-miR-451 negatively regulates the Mycoplasma gallisepticum (MG)-induced production of inflammatory cytokines via targeting tyrosine3-monooxygenase/tryptophan5-monooxygenase activation protein zeta (YWHAZ). The aim of this study was to investigate the mechanism regulating gga-miR-451 in MG infection in chickens. Analysis of gga-miR-451 precursor, pri-miR-451, and pre-miR-451 indicated that the regulation occurred transcriptionally. We also identified the transcriptional regulatory region of gga-miR-451 that contained consensus-binding motif for aryl hydrocarbon receptor (AhR) and aryl hydrocarbon receptor nuclear translocator (Arnt) complex, which is known as the transcription factor that regulates gene expression. Luciferase reporter assays combined with chromatin immunoprecipitation (ChIP) demonstrated that AhR:Arnt bound directly to the promoter elements of gga-miR-451, which were responsible for gga-miR-451 transcription in the context of MG infection. Furthermore, upregulation of AhR:Arnt significantly induced gga-miR-451 and inhibited YWHAZ expression, suggesting that AhR:Arnt may play an anti-inflammatory role in MG infection. This discovery suggests that induced gga-miR-451 expression is modulated by AhR:Arnt in response to MG infection.

Mycoplasma gallisepticum ( MG ) mainly infects chickens to initiate chronic respiratory disease (CRD). microRNAs (miRNAs) play vital roles according to previously reported studies. Our previous study showed that gga-miR-16-5p, in MG -infected lungs of chicken embryo, was upregulated by Illumina sequencing. The study aimed to reveal what role gga-miR-16-5p plays in CRD progression. gga-miR-16-5p was upregulated in MG -infected fibroblast cells (DF-1). Phosphoinositide-3-kinase regulatory subunit 1 (PIK3R1) was demonstrated as the target gene of gga-miR-16-5p. Furthermore, PIK3R1 expression was lower in MG -infected groups than it in noninfected controls measured by qPCR. Additionally, overexpressed gga-miR-16-5p could downregulate PIK3R1 and phosphorylated serine/threonine kinase (p-Akt) to express protein, whereas there is an opposite effect on inhibition. Overexpressed gga-miR-16-5p resulted in decreased activity of tumor necrosis factor alpha (TNF-α) and the nuclear factor-kappaB (NF-κB) by qPCR. Furthermore, overexpressed gga-miR-16-5p restricted cell multiplication, cycle progression, and increased apoptosis of MG -infected DF-1 cells, whereas inhibited gga-miR-16-5p led to the opposite effect. Collectively, upregulated gga-miR-16-5p could decrease multiplication, cycle progression, and increase apoptosis of MG -infected DF-1 cells, at least partly through directly targeting PIK3R1 and inhibiting PI3K/Akt/NF-κB pathway to exert an anti-inflammatory effect. Our results will provide more experimental evidence to bring pathogenesis of MG infection to light.

Mycoplasma gallisepticum ( MG ) is the pathogen of chronic respiratory disease (CRD), hallmarked by vigorous inflammation in chickens, causing the poultry industry enormous losses. miRNAs have emerged as important regulators of animal diseases. Previous miRNA sequencing data has demonstrated that miR-130b-3p is up-regulated in MG -infected chicken embryo lungs. Therefore, we aimed to investigate the function of miR-130b-3p in MG infection of chickens. RT-qPCR results confirmed that miR-130b-3p was up-regulated both in MG -infected chicken embryo lungs and chicken embryonic fibroblast cells (DF-1 cells). Furthermore, functional studies showed that overexpression of miR-130b-3p promoted MG -infected DF-1 cell proliferation and cell cycle, whereas inhibition of miR-130b-3p weakened these cellular processes. Luciferase reporter assay combined with gene expression data supported that phosphatase and tensin homolog deleted on chromosome ten (PTEN) was a direct target of miR-130b-3p. Additionally, overexpression of miR-130b-3p resulted in up-regulations of phosphatidylinositol-3 kinase (PI3K), serine/threonine kinase (AKT), and nuclear factor-κB (NF-κB), whereas inhibition of miR-130b-3p led to the opposite results. Altogether, upon MG infection, up-regulation of miR-130b-3p activates the PI3K/AKT/NF-κB pathway, facilitates cell proliferation and cell cycle via down-regulating PTEN. This study helps to understand the mechanism of host response to MG infection.

Mycoplasma gallisepticum (MG) is the most economically significant mycoplasma pathogen of poultry that causes chronic respiratory disease (CRD) in chickens. Although miRNAs have been identified as a major regulator effect on inflammatory response, it is largely unclear how they regulate MG-induced inflammation. The aim of this study was to investigate the functional roles of gga-miR-451 and identify downstream targets regulated by gga-miR-451 in MG infection of chicken. We found that the expression of gga-miR-451 was significantly up-regulated during MG infection of chicken embryo fibroblast cells (DF-1) and chicken embryonic lungs. Overexpression of gga-miR-451 decreased the MG-induced inflammatory cytokine production, including tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and interleukin-6 (IL-6), whereas inhibition of gga-miR-451 had the opposite effect. Gene expression data combined with luciferase reporter assays demonstrated that tyrosine3-monooxygenase/tryptophan5-monooxygenase activation protein zeta (YWHAZ) was identified as a direct target of gga-miR-451 in the context of MG infection. Furthermore, upregulation of gga-miR-451 significantly inhibited the MG-infected DF-1 cells proliferation, induced cell-cycle arrest, and promoted apoptosis. Collectively, our results demonstrate that gga-miR-451 negatively regulates the MG-induced production of inflammatory cytokines via targeting YWHAZ, inhibits the cell cycle progression and cell proliferation, and promotes cell apoptosis. This study provides a better understanding of the molecular mechanisms of MG infection., Competing Interests: The authors declare no conflict of interest.

Simple Summary: Mycoplasma gallisepticum causes respiratory disease in chickens. This study used RNA-seq to investigate the immune response of chicken embryos and chicks to infection. Weight loss and immune damage were observed in infected chickens of both ages. Transcriptome analysis revealed differentially expressed genes related to innate immunity and inflammation, with toll-like receptors and cytokine pathways playing a dominant role in the immune response. The immune response was stronger in embryos than in chicks. These findings provide valuable insights for developing disease control strategies. Mycoplasma gallisepticum (MG) is a major cause of chronic respiratory diseases in chickens, with both horizontal and vertical transmission modes and varying degrees of impact on different ages. The innate immune response is crucial in resisting MG infection. Therefore, this study aimed to investigate the innate immune response of chicken embryos and newly hatched chicks to MG infection using comparative RNA-seq analysis. We found that MG infection caused weight loss and immune damage in both chicken embryos and chicks. Transcriptome sequencing analysis revealed that infected chicken embryos had a stronger immune response than chicks, as evidenced by the higher number of differentially expressed genes associated with innate immunity and inflammation. Toll-like receptor and cytokine-mediated pathways were the primary immune response pathways in both embryos and chicks. Furthermore, TLR7 signaling may play an essential role in the innate immune response to MG infection. Overall, this study sheds light on the development of innate immunity to MG infection in chickens and can help in devising disease control strategies. [ABSTRACT FROM AUTHOR]

Simple Summary: Mycoplasma gallisepticum (MG) can induce chronic respiratory disease (CRD) in chickens. Glycyrrhizic acid (GA) is the major ingredient of licorice and shows excellent anti-pathogenic microorganism and anti-inflammatory properties. Based on above, this study has evaluated the therapeutic effects and safety of GA in MG-infected broilers. Study results showed that GA could inhibit the proliferation of MG in vitro and expression of MG adhesion protein (pMGA1.2) in broiler lungs. GA restored production performances and attenuated MG-induced organ damage and abnormal biochemical indicator changes in MG-infected broilers. In conclusion, GA displayed significant therapeutic efficacy regarding MG infection and had no adverse effects on broilers (100 mg/kg/d). This study was conducted to evaluate the therapeutic effects and safety of GA in MG-infected broilers. Our results showed that the minimum inhibitory concentration of GA was 31.25 μg/mL. Moreover, GA inhibited the expression of MG adhesion protein (pMGA1.2) in the broilers' lungs. GA treatment clearly decreased the morbidity of CRD and mortality in the MG-infected broilers. Compared with the model group, GA treatment significantly decreased gross air sac lesion scores and increased average weight gain and feed conversion rate in the MG-infected broilers. Histopathological examination showed GA treatment attenuated MG-induced trachea, immune organ and liver damage in the broilers. Moreover, GA treatment alone did not induce abnormal morphological changes in these organs in the healthy broilers. Compared with the model group, serum biochemical results showed GA treatment significantly decreased the content of total protein, albumin, globulin, alanine aminotransferase, aspartate aminotransferase, total bilirubin, creatinine, uric acid, total cholesterol, and increased the content of albumin/globulin, alkaline phosphatase, apolipoprotein B and apolipoprotein A-I. In conclusion, GA displayed a significant therapeutic efficacy regarding MG infection and had no adverse effects on the broilers (100 mg/kg/d). [ABSTRACT FROM AUTHOR]

Mycoplasma gallisepticum (MG) can cause chronic respiratory disease (CRD) in chickens. While several studies have reported the inflammatory functions of microRNAs during MG infection, the mechanism by which exosomal miRNAs regulate MG‐induced inflammation remains to be elucidated. The expression of exosome‐microRNA derived from MG‐infected chicken type II pneumocytes (CP‐II) was screened, and the target genes and function of differentially expressed miRNAs (DEGs) were predicted. To verify the role of exosomal gga‐miR‐451, Western blot, ELISA and RT‐qPCR were used in this study. The results showed that a total of 722 miRNAs were identified from the two exosomal small RNA (sRNA) libraries, and 30 miRNAs (9 up‐regulated and 21 down‐regulated) were significantly differentially expressed. The target miRNAs were significantly enriched in the treatment group, such as cell cycle, Toll‐like receptor signalling pathway and MAPK signalling pathway. The results have also confirmed that gga‐miR‐451‐absent exosomes derived from MG‐infected CP‐II cells increased inflammatory cytokine production in chicken fibroblast cells (DF‐1), and wild‐type CP‐II cell–derived exosomes displayed protective effects. Collectively, our work suggests that exosomes from MG‐infected CP‐II cells alter the dynamics of the DF‐1 cells, and may contribute to pathology of the MG infection via exosomal gga‐miR‐451 targeting YWHAZ involving in inflammation. [ABSTRACT FROM AUTHOR]