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

2021 ◽  
Vol 18 (1) ◽  
Author(s):  
Ye Chen ◽  
Chao Zhang ◽  
Chang-xue Xiao ◽  
Xiao-dong Li ◽  
Zhi-li Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Cell Apoptosis ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Inflammatory Genes ◽  
Functional Studies ◽  
Tnf Α ◽  
Pulmonary Inflammatory Response

Abstract 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.

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

2020 ◽  
Author(s):  
Ye Chen ◽  
Chao Zhang ◽  
Chang-xue Xiao ◽  
Xiao-dong Li ◽  
Zhi-li Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Cell Apoptosis ◽  
Mrna Levels ◽  
Rt Pcr ◽  
Inflammatory Genes ◽  
Functional Studies ◽  
Tnf Α ◽  
Pulmonary Inflammatory Response

Abstract 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.

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

2020 ◽  
Author(s):  
Ye Chen ◽  
Chao Zhang ◽  
Changxue Xiao ◽  
Xiao-dong Li ◽  
Zhi-li Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Cell Apoptosis ◽  
Cell Model ◽  
A549 Cells ◽  
Rt Pcr ◽  
Function Study ◽  
Tnf Α ◽  
Pulmonary Inflammatory Response

Abstract 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.

scholarly journals Dexamethasone can attenuate the inflammatory response in asthma via regulation of the lncH19/miR-324-3p cascade

2020 ◽  
Author(s):  
Ye Chen ◽  
Chao Zhang ◽  
Changxue Xiao ◽  
Xiao-dong Li ◽  
Zhi-li Hu ◽  
...  
Keyword(s):  
Cell Proliferation ◽  
Inflammatory Response ◽  
Western Blot ◽  
Inflammatory Cytokines ◽  
Cell Apoptosis ◽  
Cell Model ◽  
A549 Cells ◽  
Tnf Alpha ◽  
Rt Pcr ◽  
Function Study

Abstract Objective: To investigate lncRNAs and their roles in regulating the inflammatory response under treatment of Dexamethasone (Dex) in asthma.Methods: IL-1 beta (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-1 beta performed better against LPS. Dex with different concentration was used to attenuate inflammation by IL-1 beta, and its effect was assessed by RT-PCR to detect the inflammatory related mRNA, including IKbeta-alpha, IKKbeta, IL-6, IL-8, and TNF-alpha and ELISA to detect the inflammatory cytokines TNF-alpha, 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-1beta 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, hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. miRNA inhibitor was transfected into A549 NC and A549 shlncH19 cells, 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 Blot for inflammatory related genes detection were conducted to validate the functions of the target hsa-miR-324-3p. Results: Dex with 1μM 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 1μM was for further study. Under IL-1 beta 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, and the phosphorylation of P65, ICAM-1, 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 aforementioned 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 inflammatory response of asthma.Conclusion: Dex can attenuate the inflammatory response in asthma via regulation of the lncH19/miR-324-3p cascade.

S100A4 promotes the progression of lipopolysaccharide-induced acute epididymitis in mice†

2020 ◽  
Vol 102 (6) ◽  
pp. 1213-1224 ◽  
Author(s):  
Yingjie Wu ◽  
Haoran Li ◽  
Yinghe Qin
Keyword(s):  
Cell Proliferation ◽  
Cell Apoptosis ◽  
Mrna Levels ◽  
Ki 67 ◽  
Tnf Α ◽  
Proliferation And Apoptosis ◽  
Sperm Membrane ◽  
Marked Cell ◽  
Acute Epididymitis

Abstract S100A4 has been suggested to be a critical regulator of tumor metastasis and is implicated in the progression of inflammation. The aim of this study is to investigate the expression and possible role of S100A4 in epididymitis. Using a mouse model of epididymitis induced by the injection of lipopolysaccharide (LPS) in the deferent duct, we found that LPS administration induced an upregulation of S100a4 transcription (P &lt; 0.05) and a recruitment of S100A4 positive cells in the epididymal interstitium of wild type (WT) mice. Co-immunofluorescence showed that S100A4 was mainly expressed by granulocytes, CD4 lymphocytes, and macrophages. Deficiency of S100A4 reduced epididymal pathological reaction and the mRNA levels of the pro-inflammatory cytokines IL-1β and TNF-α (P &lt; 0.01), suggesting that S100A4 promotes the progression of epididymitis. Furthermore, S100A4 deficiency alleviated the decline of sperm motility and rectified the abnormal expression of sperm membrane protein AMAD3, which suggested that in the progression of epididymitis, S100A4 aggravates the damage to sperm vitality. In addition, both Ki-67 marked cell proliferation and transferase-mediated dUTP-biotin nick end labeling detected cell apoptosis were reduced in S100a4−/− mice compared with WT mice after LPS treatment, indicating that S100A4 promotes both cell proliferation and cell apoptosis in epididymitis. Overall, these results demonstrate that S100A4 promotes the progression of LPS-induced epididymitis and facilitates a decline in sperm vitality, and its function may be related to the process of cell proliferation and apoptosis during inflammation.

scholarly journals Tributyrin Plays an Important Role in Regulating the Growth and Health Status of Juvenile Blunt Snout Bream (Megalobrama amblycephala), as Evidenced by Pathological Examination

2021 ◽  
Vol 12 ◽  
Author(s):  
Hualiang Liang ◽  
Ke Ji ◽  
Xianping Ge ◽  
Bingwen Xi ◽  
Mingchun Ren ◽  
...  
Keyword(s):  
Health Status ◽  
Inflammatory Response ◽  
Antioxidant Capacity ◽  
Inflammatory Cytokines ◽  
Pathological Examination ◽  
Megalobrama Amblycephala ◽  
Intestinal Damage ◽  
Mrna Levels ◽  
Blunt Snout Bream ◽  
Tnf Α

The present study aimed to assess the role of tributyrin (TB) in regulating the growth and health status of juvenile blunt snout bream (Megalobrama amblycephala) through an 8-week feeding experiment. Six groups were fed experimental diets with added TB percentages of 0% (control group), 0.03%, 0.06%, 0.09%, 0.12% and 0.15%. The present results showed that TB supplementation in feed had some positive impacts on FW, WG, FCR and SGR, and the best results were found in the 0.06% TB group (P&lt;0.05). However, TB supplementation in feed had no significant effects on SR, CF, VSI or whole-body composition (P&gt;0.05). TB supplementation in feed increased antioxidant capacity and immunological capacity and attenuated the inflammatory response by increasing the activity of T-SOD, GPx, CAT and the levels of anti-inflammatory cytokines (IL-10 and TGF-β) and decreasing the levels of MDA and anti-inflammatory cytokines (TNF-α) (P&lt;0.05). Furthermore, TB supplementation improved immunity by increasing the levels of immunoglobulins (IgM and IgG), C3 and IFN-γ (P&lt;0.05). Surprisingly, 0.06%-0.12% TB supplementation significantly increased the content of IL-1β (P&lt;0.05). However, TB supplementation in feed had no significant effects on the plasma content of GSH, HSP70, IL-8 and the activity of T-AOC (P&gt;0.05). The possible mechanism was that TB activated PI3K/Akt/Nrf2 and inhibits the NF-κB signaling pathway, further regulating the mRNA levels of key genes with antioxidant capacity and the inflammatory response; for example, it increased the mRNA levels of Nrf2, Cu/Zn-SOD, HO-1, CAT, Akt, PI3K, GPx, IL-10, and TGF-β and decreased the mRNA levels of NF-κB and TNF-α (P&lt;0.05). In addition, 0.06%-0.15% TB supplementation significantly increased the mRNA levels of IL-1β (P&lt;0.05). TB supplementation in feed had no significant effects on the mRNA levels of HSP70, Mn-SOD and IL-8 (P&gt;0.05). Evidence was presented that TB supplementation decreased the mortality rate caused by Aeromonas hydrophila challenge. In pathological examination, TB supplementation prevented hepatic and intestinal damage. Generally, TB supplementation improved the growth performance of juvenile blunt snout bream. Furthermore, TB supplementation activated PI3K/Akt/Nrf2 and inhibited the NF-κB signaling pathway, regulating health status and preventing hepatic and intestinal damage.

scholarly journals Paeoniflorin reduces the inflammatory response of THP-1 cells by up‐regulating microRNA-124

2021 ◽  
Author(s):  
Danyun Huang ◽  
Zhijun Li ◽  
Yue Chen ◽  
Yan Fan ◽  
Tao Yu
Keyword(s):  
Systemic Lupus Erythematosus ◽  
Flow Cytometry ◽  
Inflammatory Response ◽  
Inflammatory Cytokines ◽  
Functional Recovery ◽  
Lupus Erythematosus ◽  
Biological Functions ◽  
Rt Pcr ◽  
Systemic Lupus ◽  
Tnf Α

Abstract Background The activation of macrophages and the release of inflammatory cytokines are the main reasons for the progress of systemic lupus erythematosus (SLE). MicroRNA (miRNA)-124 is involved in the regulation of macrophages and is a key regulator of inflammation and immunity. Objective To explore whether paeoniflorin (PF) regulates the biological functions of macrophages depends on miR-124. Methods RT-PCR, WB, ELISA, CCK-8 and flow cytometry were used to evaluate that PF regulated the biological functions of THP-1 cells through miR-124. Results PF significantly inhibited the proliferation while promotes the apoptosis of THP-1 cells, and inhibited the release of IL-6, TNF-α and IL-1βin THP-1 cells. RT-PCR results shown that PF up-regulated the expression of miR-124 in THP-1 cells. Functional recovery experiments showed that compared with the LPS + mimic-NC group, LPS + miR-124 mimic significantly inhibited the proliferation and the release of IL-6, TNF-α and IL-1β, but promoted the apoptosis of THP-1 cells. In addition, compared with the LPS + PF + inhibitor-NC group, LPS + PF + miR-124 inhibitor significantly promoted the proliferation and the release of IL-6, TNF-α and IL-1β, but inhibited the apoptosis of THP-1 cells. Conclusions By down-regulating miR-124, PF inhibits the proliferation and inflammation of THP-1 cells, and promotes the apoptosis of THP-1 cells.

scholarly journals Protective Effects of MicroRNA-126 on Human Cardiac Microvascular Endothelial Cells Against Hypoxia/Reoxygenation-Induced Injury and Inflammatory Response by Activating PI3K/Akt/eNOS Signaling Pathway

2017 ◽  
Vol 42 (2) ◽  
pp. 506-518 ◽  
Author(s):  
Hong-Hui Yang ◽  
Yan Chen ◽  
Chuan-Yu Gao ◽  
Zhen-Tian Cui ◽  
Jian-Min Yao
Keyword(s):  
Cell Proliferation ◽  
Inflammatory Response ◽  
Signaling Pathway ◽  
Cell Apoptosis ◽  
Control Group ◽  
Protective Effects ◽  
Enos Expression ◽  
Lumen Formation ◽  
Tnf Α ◽  
Cardiac Microvascular Endothelial Cells

Objective: This study explored the protective effects of the microRNA-126 (miR-126)-mediated PI3K/Akt/eNOS signaling pathway on human cardiac microvascular endothelial cells (HCMECs) against hypoxia/reoxygenation (H/R)-induced injury and the inflammatory response. Methods: Untreated HCMECs were selected for the control group. After H/R treatment and cell transfection, the HCMECs were assigned to the H/R, miR-126 mimic, mimic-negative control (NC), miR-126 inhibitor, inhibitor-NC, wortmannin (an inhibitor of PI3K) and miR-126 mimic + wortmannin groups. Super oxide dismutase (SOD), nitric oxide (NO), vascular endothelial growth factor (VEGF) and reactive oxygen species (ROS) were measured utilizing commercial kits. Quantitative real-time polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA) were performed to detect miR-126 expression and the mRNA and protein expression of inflammatory factors. Western blotting was used to determine the expression of key members in the PI3K/Akt/eNOS signaling pathway. ACCK-8 assay and flow cytometry were employed to examine cell proliferation and apoptosis, respectively. The angiogenic ability in each group was detected by the lumen formation test. Results: Compared to the control group, p/t-PI3K, p/t-Akt and p/t-eNOS expression, NO, VEGF and SOD levels, cell proliferation and in vitro lumen formation ability were decreased, while the ROS content, interleukin (IL)-6, IL-10 and tumor necrosis factor (TNF)-α expression and cell apoptosis were significantly increased in the H/R, mimic-NC, miR-126 inhibitor, inhibitor-NC, wortmannin and miR-126 mimic + wortmannin groups. Additionally, in comparison with the H/R group, the miR-126 mimic group had elevated p/t-PI3K, p/t-Akt and p/t-eNOS expression, increased NO, VEGF and SOD contents, and strengthened cell proliferation and lumen formation abilities but also exhibited decreased ROS content, reduced IL-6, IL-10 and TNF-α expressions, and weakened cell apoptosis, while the miR-126 inhibitor and wortmannin group exhibited the opposite results. Furthermore, decreased p/t-PI3K, p/t-Akt and p/t-eNOS expressions, decreased NO, VEGF and SOD contents, cell proliferation and lumen formation abilities, as well as increased ROS content, increased IL-6, IL-10 and TNF-α expression, and increased cell apoptosis were observed in the miR-126 mimic + wortmannin group compared to themiR-126 mimic group. Conclusions: These findings indicated that miR-126 protects HCMECs from H/R-induced injury and inflammatory response by activating the PI3K/Akt/ eNOS signaling pathway.

scholarly journals Dexamethasone can attenuate the inflammatory response in asthma via regulation of the lncH19/miR-324-3p cascade

2020 ◽  
Author(s):  
Ye Chen ◽  
Chao Zhang ◽  
Changxue Xiao ◽  
Xiao-dong Li ◽  
Zhi-li Hu ◽  
...  
Keyword(s):  
Inflammatory Response ◽  
Western Blot ◽  
Cell Viability ◽  
Inflammatory Cytokines ◽  
Cell Apoptosis ◽  
Cell Model ◽  
A549 Cells ◽  
Tnf Alpha ◽  
Rt Pcr ◽  
Cytokine Detection

Abstract Objective To investigate lncRNAs and their roles in regulating the inflammatory response under treatment of Dexamethasone (Dex) in asthma. Methods IL-1beta (10 ng/ml) was used to induce an inflammatory cell model with A549 cells. Dex was used to attenuate inflammation by IL-1beta, and its effect was assessed by RT-PCR to detect the inflammatory cytokines IKbeta-alpha, IKKbeta,, IL-6, IL-8, and TNF-alpha and ELISA to detect the inflammatory cytokines TNF-alpha, 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-1beta and attenuated by Dex, Thus, lncH19 was selected for further study. lncH19 expression was inhibited by shRNA transduced by lentivirus. Cell assays for cell viability and apoptosis as well as RT-PCR, western blot, and ELISA for inflammatory cytokines 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, hsa-miR-324-3p showed consistent patterns in A549 NC and A549 shlncH19. miRNA inhibitor was transfected into A549 cells, 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 study. RT-PCR, ELISA and Western Blot for cytokine detection were conducted to validate the functions of the target hsa-miR-324-3p. Results Dex was shown to be effective in attenuating the inflammatory response. During this process, lncH19 significantly increased in expression ( P < 0.05). Under IL-1beta treatment with or without Dex, the inhibition of lncH19 lead to an increase cell viability, a decrease in cell apoptosis, an increase in the protein level of inflammatory-related genes, and the phosphorylation of P65, ICAM-1, 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 aforementioned 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 viability and induced cell apoptosis, thereby significantly attenuating the inflammatory response, which reversed the effect of lncH19 in regulating cell viability and the secretion of inflammatory cytokines ( P < 0.05). Therefore, lncH19 might regulate miR-324-3p during Dex treatment. Conclusion Dex can attenuate the inflammatory response via regulation of the lncH19/miR-324-3p cascade.

scholarly journals Comparing the protective effects of resveratrol, curcumin and sulforaphane against LPS/IFN-γ-mediated inflammation in doxorubicin-treated macrophages

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Haidy A. Saleh ◽  
Eman Ramdan ◽  
Mohey M. Elmazar ◽  
Hassan M. E. Azzazy ◽  
Anwar Abdelnaser
Keyword(s):  
Nitric Oxide ◽  
Inflammatory Response ◽  
Raw 264.7 Cells ◽  
Inos Mrna ◽  
No Production ◽  
Raw 264.7 ◽  
Mrna Levels ◽  
Protective Effects ◽  
Tnf Α ◽  
Ifn Γ

AbstractDoxorubicin (DOX) chemotherapy is associated with the release of inflammatory cytokines from macrophages. This has been suggested to be, in part, due to DOX-mediated leakage of endotoxins from gut microflora, which activate Toll-like receptor 4 (TLR4) signaling in macrophages, causing severe inflammation. However, the direct function of DOX on macrophages is still unknown. In the present study, we tested the hypothesis that DOX alone is incapable of stimulating inflammatory response in macrophages. Then, we compared the anti-inflammatory effects of curcumin (CUR), resveratrol (RES) and sulforaphane (SFN) against lipopolysaccharide/interferon-gamma (LPS/IFN-γ)-mediated inflammation in the absence or presence of DOX. For this purpose, RAW 264.7 cells were stimulated with LPS/IFN-γ (10 ng/mL/10 U/mL) in the absence or presence of DOX (0.1 µM). Our results showed that DOX alone is incapable of stimulating an inflammatory response in RAW 264.7 macrophages. Furthermore, after 24 h of incubation with LPS/IFN-γ, a significant increase in tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and inducible nitric oxide synthase (iNOS) mRNA levels was observed. Similarly, nitric oxide (NO) production and TNF-α and IL-6 protein levels were significantly upregulated. Moreover, in LPS/IFN-γ-treated macrophages, the microRNAs (miRNAs) miR-146a, miR-155, and miR-21 were significantly overexpressed. Interestingly, upon testing CUR, RES, and SFN against LPS/IFN-γ-mediated inflammation, only SFN was able to significantly reverse the LPS/IFN-γ-mediated induction of iNOS, TNF-α and IL-6 and attenuate miR-146a and miR-155 levels. In conclusion, SFN, at the transcriptional and posttranscriptional levels, exhibits potent immunomodulatory action against LPS/IFN-γ-stimulated macrophages, which may indicate SFN as a potential treatment for DOX-associated inflammation.

scholarly journals Modulation of ACE-2 mRNA by inflammatory cytokines in human thyroid cells: a pilot study

Endocrine ◽  
2021 ◽  
Author(s):  
Francesca Coperchini ◽  
Gianluca Ricci ◽  
Laura Croce ◽  
Marco Denegri ◽  
Rubina Ruggiero ◽  
...  
Keyword(s):  
Inflammatory Cytokines ◽  
Combination Treatment ◽  
Primary Cultures ◽  
Human Thyroid ◽  
Thyroid Cell ◽  
Mrna Levels ◽  
Thyroid Cells ◽  
Tnf Α ◽  
Ifn Γ ◽  
Pro Inflammatory Cytokines

Abstract Introduction Angiotensin-converting-enzyme-2 (ACE-2) was demonstrated to be the receptor for cellular entry of SARS-CoV-2. ACE-2 mRNA was identified in several human tissues and recently also in thyroid cells in vitro. Purpose Aim of the present study was to investigate the effect of pro-inflammatory cytokines on the ACE-2 mRNA levels in human thyroid cells in primary cultures. Methods Primary thyroid cell cultures were treated with IFN-γ and TNF-α alone or in combination for 24 h. ACE-2 mRNA levels were measured by RT-PCR. As a control, the levels of IFN-γ inducible chemokine (CXCL10) were measured in the respective cell culture supernatants. Results The mean levels of ACE-2 mRNA increased after treatment with IFN-γ and TNF-α in all the thyroid cell preparations, while the combination treatment did not consistently synergically increase ACE-2-mRNA. At difference, CXCL10 was consistently increased by IFN-γ and synergically further increased by the combination treatment with IFN-γ + TNF-α, with respect to IFN-γ alone. Conclusions The results of the present study show that IFN-γ and, to a lesser extent TNF-α consistently increase ACE-2 mRNA levels in NHT primary cultures. More interestingly, the combined stimulation (proven to be effective according to the synergic effect registered for CXCL10) produces different responses in terms of ACE-2 mRNA modulation. These results would suggest that elevated levels of pro-inflammatory cytokines could facilitate the entering of the virus in cells by further increasing ACE-2 expression and/or account for the different degree of severity of SARS-COV-2 infection. This hypothesis deserves to be confirmed by further specific studies.

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