Disorders in blood circulation as a probable cause of death in dogs infected with Babesia canis

Introduction The purpose of the study was to investigate post-mortem changes in dogs infected with Babesia canis and to establish the probable cause of death of the affected animals. Material and Methods Cadavers of six dogs that did not survive babesiosis were collected. Necropsies were performed and samples of various organs were collected for histological examination. Results Necropsies and histological examinations revealed congestion and oedemata in various organs. Most of the dogs had ascites, hydrothorax or hydropericardium, pulmonary oedema, pulmonary, renal, hepatic, and cerebral congestion, and necrosis of cardiomyocytes. Conclusion These results suggested disorders in blood circulation as the most probable cause of death. However, the pulmonary inflammatory response and cerebral babesiosis observed in some of these dogs could also be considered possible causes of death. This study also showed a possible role for renal congestion in the development of renal hypoxia and azotaemia in canine babesiosis.

Mild Attenuation of the Pulmonary Inflammatory Response in a Mouse Model of Hereditary Hemochromatosis Type 4

The respiratory tract is constantly exposed to pathogens that require iron for proliferation and virulence. Pulmonary iron levels are increased in several lung diseases and associated with increased susceptibility to infections. However, regulation of lung iron homeostasis and its cross talk to pulmonary immune responses are largely unexplored. Here we investigated how increased lung iron levels affect the early pulmonary inflammatory response. We induced acute local pulmonary inflammation via aerosolized LPS in a mouse model of hereditary hemochromatosis type 4 (Slc40a1C326S/C326S), which is hallmarked by systemic and pulmonary iron accumulation, specifically in alveolar macrophages. We show that Slc40a1C326S/C326S mice display a mild attenuation in the LPS-induced pulmonary inflammatory response, with a reduced upregulation of some pro-inflammatory cytokines and chemokines. Despite mildly reduced cytokine levels, there is no short-term impairment in the recruitment of neutrophils into the bronchoalveolar space. These data suggest that increased pulmonary iron levels do not strongly alter the acute inflammatory response of the lung.

Dexamethasone can attenuate the pulmonary inflammatory response via regulation of the lncH19/miR-324-3p cascade

Objective To investigate lncRNAs and their roles in regulating the pulmonary inflammatory response under dexamethasone (Dex) treatment. Methods IL-1β (10 ng/mL) and LPS (1 μg/mL) was used to construct inflammatory cell models with A549 cells; IL-1β performed better against LPS. Different concentrations of Dex were used to attenuate the inflammation induced by IL-1β, and its effect was assessed via RT-PCR to detect inflammatory cytokine-related mRNA levels, including those of IKβ-α, IKKβ, IL-6, IL-8, and TNF-α. Furthermore, ELISA was used to detect the levels of the inflammatory cytokines TNF-α, IL-6, and IL-8. RT-PCR was used to quantify the levels of lncRNAs, including lncMALAT1, lncHotair, lncH19, and lncNeat1. LncH19 was most closely associated with the inflammatory response, which was induced by IL-1β and attenuated by Dex. Among the lncRNAs, the level of lncH19 showed the highest increase following treatment with 1 and 10 μM Dex. Therefore, lncH19 was selected for further functional studies. LncH19 expression was inhibited by shRNA transduced with lentivirus. Cell assays for cell proliferation and apoptosis as well as RT-PCR, western blot, and ELISA for inflammatory genes were conducted to confirm the functions of lncH19. The predicted target miRNAs of lncH19 were hsa-miR-346, hsa-miR-324-3p, hsa-miR-18a-3p, hsa-miR-18b-5p, hsa-miR-146b-3p, hsa-miR-19b-3p, and hsa-miR-19a-3p. Following estimation via RT-PCR, hsa-miR-346, hsa-miR-18a-3p, and hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. An miRNA inhibitor was transfected into A549 NC and A549 shlncH19 cells, and the expression levels were determined via RT-PCR. hsa-miR-324-3p was inhibited the most compared with hsa-miR-346 and hsa-miR-18a-3p and was subjected to further functional studies. RT-PCR, ELISA, and western blotting for inflammatory gene detection were conducted to validate the functions of the target hsa-miR-324-3p. Results Treatment with 1 and 10 μM Dex could effectively attenuate the inflammatory response. During this process, lncH19 expression significantly increased (P < 0.05). Therefore, treatment with 1 μM Dex was used for further study. Under IL-1β treatment with or without Dex, lncH19 inhibition led to an increase in cell proliferation; a decrease in cell apoptosis; an increase in the protein levels of inflammatory genes; phosphorylation of P65, ICAM-1, and VCAM-1; and increase inflammatory cytokines. Prediction of the targets of lncH19 and validation via RT-PCR revealed that miR-346, miR-18a-3p, and miR-324-3p negatively correlate with lncH19. Additionally, Dex increased the lncH19 expression but reduced that of the miRNAs. Among the miRNAs, miR-324-3p was the most markedly downregulated miRNA following treatment of miRNA inhibitors. The MTS assay and cell apoptosis assay showed that the miR-324-3p inhibitor inhibited cell proliferation and induced cell apoptosis, thereby significantly attenuating the inflammatory response, which reversed the effect of lncH19 in regulating cell proliferation and the secretion of inflammatory cytokines (P < 0.05). Therefore, lncH19 might regulate miR-324-3p in pulmonary inflammatory response under Dex treatment. Conclusion Dex can attenuate the pulmonary inflammatory response by regulating the lncH19/miR-324-3p cascade.

Modification of the pulmonary MyD88 inflammatory response underlies the role of the Yersinia pestis pigmentation locus in primary pneumonic plague

Pneumonic plague, caused by Yersinia pestis, is a rapidly progressing bronchopneumonia involving focal bacterial growth, neutrophilic congestion, and alveolar necrosis. Within a short time after inhalation of Y. pestis, inflammatory cytokines are expressed via the Toll/IL1 adaptor myeloid differentiation primary response 88 (MyD88), which facilitates the primary lung infection. We previously showed that Y. pestis lacking the 102kb chromosomal pigmentation locus (pgm) are unable to cause inflammatory damage in the lungs, whereas the WT strain induces the toxic MyD88 pulmonary inflammatory response. In this work, we investigated the involvement of the pgm in skewing the inflammatory response during pneumonic plague. We show that the early MyD88-dependent and -independent cytokine responses to pgm- Y. pestis infection of the lungs are similar yet distinct from those that occur during pgm+ infection. Furthermore, we found that MyD88 was necessary to prevent growth of the iron-starved pgm- Y. pestis despite the presence of iron chelators lactoferrin and transferrin. However, while this induced neutrophil recruitment, there was no hyper-inflammatory response and pulmonary disease was mild without MyD88. In contrast, growth in blood and tissues progressed rapidly in the absence of MyD88, due to an almost total loss of serum IFNγ. We further show that the expression of MyD88 by myeloid cells is important to control bacteremia, but not the primary lung infection. The combined data indicate distinct roles for myeloid and non-myeloid MyD88, and suggest that expression of the pgm locus is necessary to skew the inflammatory response in the lungs to cause pneumonic plague.

Dexamethasone can attenuate the pulmonary inflammatory response via regulation of the lncH19/miR-324-3p cascade

Objective: To investigate lncRNAs and their roles in regulating the pulmonary inflammatory response under dexamethasone (Dex) treatmentMethods: IL-1β (10 ng/mL) and LPS (1 μg/mL) was used to construct inflammatory cell models with A549 cells; IL-1β performed better against LPS. Different concentrations of Dex were used to attenuate the inflammation induced by IL-1β, and its effect was assessed via RT-PCR to detect inflammatory cytokine-related mRNA levels, including those of IKβ-α, IKKβ, IL-6, IL-8, and TNF-α. Furthermore, ELISA was used to detect the levels of the inflammatory cytokines TNF-α, IL-6, and IL-8. RT-PCR was used to quantify the levels of lncRNAs, including lncMALAT1, lncHotair, lncH19, and lncNeat1. LncH19 was most closely associated with the inflammatory response, which was induced by IL-1β and attenuated by Dex. Among the lncRNAs, the level of lncH19 showed the highest increase following treatment with 1 and 10 μM Dex. Therefore, lncH19 was selected for further functional studies. LncH19 expression was inhibited by shRNA transduced with lentivirus. Cell assays for cell proliferation and apoptosis as well as RT-PCR, western blot, and ELISA for inflammatory genes were conducted to confirm the functions of lncH19. The predicted target miRNAs of lncH19 were hsa-miR-346, hsa-miR-324-3p, hsa-miR-18a-3p, hsa-miR-18b-5p, hsa-miR-146b-3p, hsa-miR-19b-3p, and hsa-miR-19a-3p. Following estimation via RT-PCR, hsa-miR-346, hsa-miR-18a-3p, and hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. An miRNA inhibitor was transfected into A549 NC and A549 shlncH19 cells, and the expression levels were determined via RT-PCR. hsa-miR-324-3p was inhibited the most compared with hsa-miR-346 and hsa-miR-18a-3p and was subjected to further functional studies. RT-PCR, ELISA, and western blotting for inflammatory gene detection were conducted to validate the functions of the target hsa-miR-324-3p.Results: Treatment with 1 and 10 μM Dex could effectively attenuate the inflammatory response. During this process, lncH19 expression significantly increased (P < 0.05). Therefore, treatment with 1 µM Dex was used for further study. Under IL-1β treatment with or without Dex, lncH19 inhibition led to an increase in cell proliferation; a decrease in cell apoptosis; an increase in the protein levels of inflammatory genes; phosphorylation of P65, ICAM-1, and VCAM-1; and increase inflammatory cytokines. Prediction of the targets of lncH19 and validation via RT-PCR revealed that miR-346, miR-18a-3p, and miR-324-3p negatively correlate with lncH19. Additionally, Dex increased the lncH19 expression but reduced that of the miRNAs. Among the miRNAs, miR-324-3p was the most markedly downregulated miRNA following treatment of miRNA inhibitors. The MTS assay and cell apoptosis assay showed that the miR-324-3p inhibitor inhibited cell proliferation and induced cell apoptosis, thereby significantly attenuating the inflammatory response, which reversed the effect of lncH19 in regulating cell proliferation and the secretion of inflammatory cytokines (P < 0.05). Therefore, lncH19 might regulate miR-324-3p in pulmonary inflammatory response under Dex treatment.Conclusion: Dex can attenuate the pulmonary inflammatory response by regulating the lncH19/miR-324-3p cascade.

Dexamethasone can attenuate the pulmonary inflammatory response via regulation of the lncH19/miR-324-3p cascade

Objective: To investigate lncRNAs and their roles in regulating the pulmonary inflammatory response under treatment of Dexamethasone (Dex).Methods: IL-1β (10 ng/mL) and LPS (1 μg/mL) was used to induce an inflammatory cell model with A549 cells, and the results showed that IL-1β performed better against LPS. Dex with different concentration was used to attenuate inflammation by IL-1β, and its effect was assessed by RT-PCR to detect the inflammatory related mRNA, including IKβ-α, IKKβ, IL-6, IL-8, and TNF-α. And ELISA to detect the inflammatory cytokines TNF-α, IL-6 and IL-8. RT-PCR was used to quantify levels of lncRNAs, including lncMALAT1, lncHotair, lncH19, and lncNeat1. LncH19 was most closely correlated with the inflammatory response, which was induced by IL-1β and attenuated by Dex. Among the lncRNAs, the level of lncH19 exhibited the highest increase following treatment with 1 μM and 10 μM Dex. Therefore, lncH19 was selected for further function study. LncH19 expression was inhibited by shRNA transduced by lentivirus. Cell assays for cell proliferation and apoptosis as well as RT-PCR, western blot, and ELISA for inflammatory related genes were conducted to confirm the functions of lncH19. Predicted target miRNAs of lncH19 included the following: hsa-miR-346, hsa-miR-324-3p, hsa-miR-18a-3p, hsa-miR-18b-5p, hsa-miR-146b-3p, hsa-miR-19b-3p and hsa-miR-19a-3p. Following estimation by RT-PCR, hsa-miR-346, hsa-miR-18a-3p and hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. miRNA inhibitor was transfected into A549 NC and A549 shlncH19cells, and expression levels were determined by RT-PCR. Hsa-miR-324-3p was inhibited the most relative to hsa-miR-346 and hsa-miR-18a-3p and was subjected to further function study. RT-PCR, ELISA and western blotting for inflammatory related genes detection were conducted to validate the functions of the target hsa-miR-324-3p.Results: Dex with 1 μM and 10 μM were shown to be effective in attenuating the inflammatory response. During this process, lncH19 significantly increased in expression (P < 0.05). Dex with 1 μM was for further study. Under IL-1β treatment with or without Dex, the inhibition of lncH19 lead to an increase cell proliferation, a decrease in cell apoptosis, an increase in the protein level of inflammatory-related genes, the phosphorylation of P65, ICAM-1 and VCAM-1, and inflammatory cytokines. Following prediction of the targets of lncH19 and validation by RT-PCR, miR-346, miR-18a-3p and miR-324-3p were found to be negatively correlated to lncH19. Additionally, Dex increased the expression of lncH19, but the expression of the miRNAs was reduced. Among miRNAs, miR-324-3p was the most markedly down-regulated following treatment of miRNA inhibitors. The MTS assay and cell apoptosis assay showed that the miR-324-3p inhibitor inhibited cell proliferation and induced cell apoptosis, thereby significantly attenuating the inflammatory response, which reversed the effect of lncH19 in regulating cell proliferation and the secretion of inflammatory cytokines (P < 0.05). Therefore, lncH19 might regulate miR-324-3p during Dex treatment in pulmonary inflammatory response.Conclusion: Dex can attenuate the pulmonary inflammatory response via regulation of the lncH19/miR-324-3p cascade.