Author Correction: Mir-21 Mediates the Inhibitory Effect of Ang (1-7) on AngII-induced NLRP3 Inflammasome Activation by Targeting Spry1 in lung fibroblasts.

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Mir-21 Mediates the Inhibitory Effect of Ang (1-7) on AngII-induced NLRP3 Inflammasome Activation by Targeting Spry1 in lung fibroblasts.

MicroRNA-21 (mir-21) induced by angiotensin II (AngII) plays a vital role in the development of pulmonary fibrosis, and the NLRP3 inflammasome is known to be involved in fibrogenesis. However, whether there is a link between mir-21 and the NLRP3 inflammasome in pulmonary fibrosis is unknown. Angiotensin-converting enzyme 2/angiotensin(1-7) [ACE2/Ang(1-7)] has been shown to attenuate AngII-induced pulmonary fibrosis, but it is not clear whether ACE2/Ang(1-7) protects against pulmonary fibrosis by inhibiting AngII-induced mir-21 expression. This study's aim was to investigate whether mir-21 activates the NLRP3 inflammasome and mediates the different effects of AngII and ACE2/Ang(1-7) on lung fibroblast apoptosis and collagen synthesis. In vivo, AngII exacerbated bleomycin (BLM)-induced lung fibrosis in rats, and elevated mir-21 and the NLRP3 inflammasome. In contrast, ACE2/Ang(1-7) attenuated BLM-induced lung fibrosis, and decreased mir-21 and the NLRP3 inflammasome. In vitro, AngII activated the NLRP3 inflammasome by up-regulating mir-21, and ACE2/Ang(1-7) inhibited NLRP3 inflammasome activation by down-regulating AngII-induced mir-21. Over-expression of mir-21 activated the NLRP3 inflammasome via the ERK/NF-κB pathway by targeting Spry1, resulting in apoptosis resistance and collagen synthesis in lung fibroblasts. These results indicate that mir-21 mediates the inhibitory effect of ACE2/Ang(1-7) on AngII-induced activation of the NLRP3 inflammasome by targeting Spry1 in lung fibroblasts.

[Receptor for advanced glycation end products upregulates MUC5AC expression and promotes mucus overproduction in mice with toluene diisocyanate-induced asthma].

OBJECTIVE: To explore the role of the receptor for advanced glycation end products (RAGE) in regulating the expression of MUC5AC and mucus production in a mouse model of toluene diisocyanate (TDI)?induced asthma. METHODS: BALB/c mice were randomly divided into control group, vehicle (AOO) group, TDI?induced asthma group and RAGE inhibitor (FPS?ZM1) group. PAS staining, Western blotting, and immunohistochemistry were used to analyze the changes in mucus production and MUC5AC expression in the airway of the mice, and the expression of p?ERK was detected with Western blotting. In vitro cultured human bronchial epithelial cell line 16HBE was transfected with lentiviral vector carrying short hairpin RNA targeting RAGE (shRNA?RAGE) and subsequently challenged with a TDI?human serum albumin (TDI-HSA) conjugate, and the changes in cellular MUC5AC mRNA expression as detected using RT-PCR; the protein expressions of ERK and p?ERK in the cells were examined with Western blotting. The effect of ERK inhibitor U0126 pretreatment on MUC5AC mRNA expression was also analyzed in the cells. RESULTS: Compared with the control mice, TDI-induced asthmatic mice showed significantly higher rates of PAS positivity and increased MUC5AC and p?ERK expressions in the airway (P<0.05). Treatment with FPS?ZM1 significantly decreased PAS positivity and lowered MUC5AC and p?ERK expressions in the airway of the asthmatic mice (P<0.05). Exposure of 16HBE cells to TDI?HSA caused a significant increase in MUC5AC mRNA expression and p?ERK protein expression (P<0.05), while RAGE knockdown obviously suppressed TDI?HSA-induced upregulation of p-ERK and MUC5AC mRNA (P<0.05). Treatment with the ERK inhibitor U0126 also lowered TDI?HSA?induced up?regulation of MUC5AC mRNA in the cells (P<0.05). CONCLUSION: RAGE signaling induces MUC5AC expression via extracellular signal-regulated kinase pathway to promote mucus overproduction in mice with TDI-induced asthma.

Characteristics of the mitochondrial genome of four native goats in China (Capra hircus).

Here, we describe the complete mitochondrial genome sequences of Jining Gray goat, Fushun black goat, Youzhou black-skin goat, and Hechuan white goat. The mitogenome of those four goats consisted of 16,640 nt, consisting of 13 protein-coding genes, 22 transfer RNA (tRNA) genes, 2 ribosomal RNA (rRNA) genes and a control region. As in other mammals, most mitochondrial genes are encoded on the heavy strand, except for ND6 and eight tRNA genes, which are encoded on the light strand. The complete mitogenome of these four local breeds of Chinese native goats could provide an important data to further breed improvement and animal genetics resource conservation in China.

The angiotensin-converting enzyme 2/angiotensin (1-7)/Mas axis protects against lung fibroblast migration and lung fibrosis by inhibiting the NOX4-derived ROS-mediated RhoA/Rho kinase pathway.

UNLABELLED: Reactive oxygen species (ROS) generated by NADPH oxidase-4 (NOX4) have been shown to initiate lung fibrosis. The migration of lung fibroblasts to the injured area is a crucial early step in lung fibrosis. The angiotensin-converting enzyme 2 (ACE2)/angiotensin (1-7) [Ang(1-7)]/Mas axis, which counteracts the ACE/angiotensin II (AngII)/angiotensin II type 1 receptor (AT1R) axis, has been shown to attenuate pulmonary fibrosis. Nevertheless, the exact molecular mechanism remains unclear. AIMS: To investigate the different effects of the two axes of the renin-angiotensin system (RAS) on lung fibroblast migration and extracellular matrix accumulation by regulating the NOX4-derived ROS-mediated RhoA/Rho kinase (Rock) pathway. RESULTS: In vitro, AngII significantly increased the NOX4 level and ROS production in lung fibroblasts, which stimulated cell migration and α-collagen I synthesis through the RhoA/Rock pathway. These effects were attenuated by N-acetylcysteine (NAC), diphenylene iodonium, and NOX4 RNA interference. Moreover, Ang(1-7) and lentivirus-mediated ACE2 (lentiACE2) suppressed AngII-induced migration and α-collagen I synthesis by inhibiting the NOX4-derived ROS-mediated RhoA/Rock pathway. However, Ang(1-7) alone exerted analogous effects on AngII. In vivo, constant infusion with Ang(1-7) or intratracheal instillation with lenti-ACE2 shifted the RAS balance toward the ACE2/Ang(1-7)/Mas axis, alleviated bleomycin-induced lung fibrosis, and inhibited the RhoA/Rock pathway by reducing NOX4-derived ROS. INNOVATION: This study suggests that the ACE2/Ang(1-7)/Mas axis may be targeted by novel pharmacological antioxidant strategies to treat lung fibrosis induced by AngII-mediated ROS. CONCLUSION: The ACE2/Ang(1-7)/Mas axis protects against lung fibroblast migration and lung fibrosis by inhibiting the NOX4-derived ROS-mediated RhoA/Rock pathway.

Angiotensin-converting enzyme 2/angiotensin-(1-7)/Mas axis protects against lung fibrosis by inhibiting the MAPK/NF-κB pathway.

Accumulating evidence has demonstrated that up-regulation of the angiotensin (Ang)-converting enzyme (ACE)/AngII/AngII type 1 receptor (AT1R) axis aggravates pulmonary fibrosis. The recently discovered ACE2/Ang-(1-7)/Mas axis, which counteracts the activity of the ACE/AngII/AT1R axis, has been shown to protect against pulmonary fibrosis. However, the mechanisms by which ACE2 and Ang-(1-7) attenuate pulmonary fibrosis remain unclear. We hypothesized that up-regulation of the ACE2/Ang-(1-7)/Mas axis protects against bleomycin (BLM)-induced pulmonary fibrosis by inhibiting the mitogen-activated protein kinase (MAPK)/NF-κB pathway. In vivo, Ang-(1-7) was continuously infused into Wistar rats that had received BLM or AngII. In vitro, human fetal lung-1 cells were pretreated with compounds that block the activities of AT1R, Mas (A-779), and MAPKs before exposure to AngII or Ang-(1-7). The human fetal lung-1 cells were infected with lentivirus-mediated ACE2 before exposure to AngII. In vivo, Ang-(1-7) prevented BLM-induced lung fibrosis and AngII-induced lung inflammation by inhibiting the MAPK phosphorylation and NF-κB signaling cascades. However, exogenous Ang-(1-7) alone clearly promoted lung inflammation. In vitro, Ang-(1-7) and lentivirus-mediated ACE2 inhibited the AngII-induced MAPK/NF-κB pathway, thereby attenuating inflammation and α-collagen I production, which could be reversed by the Mas inhibitor, A-779. Ang-(1-7) inhibited AngII-induced lung fibroblast apoptotic resistance via inhibition of the MAPK/NF-κB pathway and activation of the BCL-2-associated X protein/caspase-dependent mitochondrial apoptotic pathway. Ang-(1-7) alone markedly stimulated extracellular signal-regulated protein kinase 1/2 phosphorylation and the NF-κB cascade. Up-regulation of the ACE2/Ang-(1-7)/Mas axis protected against pulmonary fibrosis by inhibiting the MAPK/NF-κB pathway. However, close attention should be paid to the proinflammatory effects of Ang-(1-7).

[Expression and significance of Toll-like receptor-4 in rats lung established by passive smoking or associated with intratracheal instillation of lipopolysaccharide].

OBJECTIVE: To explore the expression and effect of Toll-like receptor 4 (TLR-4) in the lung tissue of rats established by passive smoking or intratracheal instillation of lipopolysaccharide (LPS). METHODS: Eight-week-old male Sprague-Dawley rats (n = 15) were randomly divided into 3 groups, including: (1) group A: conventional breeding; (2) group B: the rats were placed into a 120-L organic glass box with twice-daily exposure to cigarette smoking plus an intratracheal instillation of water at Day 1 and 14; (3) Group C: exposure to cigarette smoking the same as group B plus intratracheal instillation of lipopolysaccharide (1 mg/kg) at Day 1 and 14. Four weeks later, general status, arterial blood gas, pulmonary function and histopathology were analyzed. The expressions of TLR-4 and nuclear factor kappa-B (NF-κB) were determined by immunohistochemistry. Western blot was used to measure the protein contents of TLR-4, NF-κB, p-Iκ-Kα/ß, Iκ-Kα/ß, IκB-α. And real-time polymerase chain reaction (PCR) was employed to detect the mRNA levels of tumor necrosis factor α (TNF-α) and interleukin-6 (IL-6). RESULTS: Rats in Groups B and C were marantic with intermittent cough and dyspnea. Peak expiratory flow (PEF) and 50% expiratory flow-volume (EP50) were much lower in Group C ((10.6 ± 1.4), (0.77 ± 0.14) ml/s) than that in Groups A ((13.5 ± 2.0), (1.01 ± 0.08) and B (12.3 ± 0.9), (0.91 ± 0.10) ml/s) (all P < 0.05). Accumulated volume (AV) and carbon dioxide pressure (PCO2) were much higher in Groups B ((4358 ± 1501) ml, (52.77 ± 1.97) mm Hg) (1 mm Hg = 0.133 kPa) and C ((10 077 ± 1866) ml, (51.03 ± 4.96) mm Hg) than that in Group A ((1735 ± 798) ml, (39.57 ± 1.43) mm Hg) (all P < 0.05). Hematoxylin and eosin stain showed chronic bronchitis and emphysema in Groups B and C. Besides, quantitative analysis demonstrated that in unit area, mean lining interval (MLI) and destruction index (DI) in Group B ((84 ± 13) µm, 0.228 ± 0.047) and Group C ((86 ± 10) µm, 0.294 ± 0.060) significantly increased versus Group A ((65 ± 6) µm, 0.036 ± 0.012) (all P < 0.05). Immunohistochemical staining indicated that the expression of TLR-4 in cytoplasm and cytomembrane and NF-κB in nucleus markedly increased in Groups B and C versus Group A. Relative expressions of TLR-4 and NF-κB assayed by Western blot increased in Group B (0.68 ± 0.03, 0.21 ± 0.08) and Group C (1.12 ± 0.11, 0.59 ± 0.06) than that in Group A (1.36 ± 0.07, 1.04 ± 0.08). Compared with Group A, the expression levels of TLR-4, NF-κB and IκB-α and the phosphorylation levels of Iκ-Kα/ß in Group B and C significantly increased (all P < 0.05). The mRNA levels of TNF-α and IL-6 increased in Group B (3.95 ± 0.29, 5.04 ± 0.28) and C (5.33 ± 0.26, 7.23 ± 0.39) versus that in Group C (1.00 ± 0.37, 1.00 ± 0.25) (all P < 0.05). CONCLUSIONS: Both passive smoking and intratracheal instillation of LPS may cause lung injury analogous to chronic obstructive pulmonary disease via NF-κB signaling pathway. And TLR4 plays an important role in this process.

[Anti-fibrotic effects of angiotensin1-7 on bleomycin-induced pulmonary fibrosis in rats].

OBJECTIVE: To explore the anti-fibrotic effects of angiotensin (Ang) 1-7 on bleomycin (BLM) -induced pulmonary fibrosis in rats. METHODS: Eighteen Wistar male rats were randomly divided into 3 groups, including control group (intratracheal instillation with physiological saline and subcutaneous micro-pump with bi-distilled water at the rate of 0.29 µl/h), BLM group (intratracheal instillation with bleomycin and subcutaneous micro-pump with bi-distilled water at the same rate) and BLM+Ang1-7 group (intratracheal instillation with bleomycin and subcutaneous micro-pump with Ang1-7 at a dose of 25 µg·kg(-1) · h(-1) at the same rate). At Day 28, lung tissues were collected. Histological changes of lungs were evaluated by hematoxylin and eosin and Masson's trichrome stains. Collagen content of lung tissues was assessed by hydroxyprolin concentration. Then the products of protein and RNA were collected. And Western blot and realtime polymerase chain reaction (RT-PCR) were used to detect the protein or mRNA of TGF-ß1 and α-collagenI. Human embryonic lung fibroblast (HFL-1) was divided into 5 groups: (1) control group: no stimulation; (2) AngII group: stimulation of AngII (10(-7)mol/L) ; (3) Ang1-7 group: stimulation of Ang1-7 (10(-7)mol/L); (4) Ang1-7 plus AngII group: stimulation by AngII (10(-7)mol/L) with Ang1-7 (10(-7)mol/L) pre-treatment; (5) Ang1-7+AngII+A-779 group: stimulation by AngII and Ang1-7 (10(-7) mol/L) with Mas receptor inhibitor A-779 (10(-6)mol/L) pre-treatment. Then the products of protein and RNA were collected. And QuantiGene and RT-PCR were used to detect the activation of TGF-ß1, and α-collagenI mRNA. RESULTS: Compared with control group, fibrosis score and hydroxyproline concentrations increased significantly in BLM group, but declined in BLM+Ang1-7 group. The difference was statistically significant (P < 0.05). TGF-ß1 mRNA, α-collagenI mRNA and α-collagenI protein level were up-regulated by BLM (4.45 ± 0.45 vs 1.00 ± 0.20, 5.14 ± 0.55 vs 1.00 ± 0.08, 1.48 ± 0.34 vs 0.23 ± 0.11) (all P < 0.05); while compared with BLM group, those of BLM+Ang1-7 group were down-regulated (2.80 ± 0.35, 3.10 ± 0.52, 0.49 ± 0.11) (all P < 0.05). In vitro: TGF-ß1 mRNA and α-collagen I mRNA level were up-regulated by AngII (1.67 ± 0.26 vs 1.00 ± 0.10, 4.86 ± 1.36 vs 1.46 ± 0.54) (all P < 0.05); while those of AngII+Ang1-7 group were down-regulated (0.91 ± 0.30, 1.57 ± 0.27) compared with AngII group (all P < 0.05); no significant difference existed between the AngII+Ang1-7+A-779 group (1.25 ± 0.14, 1.29 ± 0.49) and AngII+Ang1-7 group (P > 0.05). CONCLUSION: Ang1-7 has anti-fibrous effect upon bleomycin-induced pulmonary fibrosis in rats and such an effect of Ang1-7 may be associated with AngII-induced expression of TGF-ß1.

Spironolactone lowers portal hypertension by inhibiting liver fibrosis, ROCK-2 activity and activating NO/PKG pathway in the bile-duct-ligated rat.

OBJECTIVE: Aldosterone, one of the main peptides in renin angiotensin aldosterone system (RAAS), has been suggested to mediate liver fibrosis and portal hypertension. Spironolactone, an aldosterone antagonist, has beneficial effect on hyperdynamic circulation in clinical practice. However, the mechanisms remain unclear. The present study aimed to investigate the role of spionolactone on liver cirrhosis and portal hypertension. METHODS: Liver cirrhosis was induced by bile duct ligation (BDL). Spironolactone was administered orally (20 mg/kg/d) after bile duct ligation was performed. Liver fibrosis was assessed by histology, Masson's trichrome staining, and the measurement of hydroxyproline and type I collagen content. The activation of HSC was determined by analysis of alpha smooth muscle actin (α-SMA) expression. Protein expressions and protein phosphorylation were determined by immunohistochemical staining and Western blot analysis, Messenger RNA levels by quantitative real time polymerase chain reaction (Q-PCR). Portal pressure and intrahepatic resistance were examined in vivo. RESULTS: Treatment with spironolactone significantly lowered portal pressure. This was associated with attenuation of liver fibrosis, intrahepatic resistance and inhibition of HSC activation. In BDL rat liver, spironolactone suppressed up-regulation of proinflammatory cytokines (TNFα and IL-6). Additionally, spironolactone significantly decreased ROCK-2 activity without affecting expression of RhoA and Ras. Moreover, spironolactone markedly increased the levels of endothelial nitric oxide synthase (eNOS), phosphorylated eNOS and the activity of NO effector-protein kinase G (PKG) in the liver. CONCLUSION: Spironolactone lowers portal hypertension by improvement of liver fibrosis and inhibition of intrahepatic vasoconstriction via down-regulating ROCK-2 activity and activating NO/PKG pathway. Thus, early spironolactone therapy might be the optional therapy in cirrhosis and portal hypertension.

[Effect of partial splenic embolization in prevention of gastroesophageal variceal rebleeding].

OBJECTIVE: To evaluate the effect of partial splenic embolization (PSE) in prevention of gastroesophageal variceal rebleeding. METHODS: Sixty-two patients with recent gastroesophageal variceal bleeding were treated by PSE with Seldinger technique. All the patients were followed-up for 12 months. The data including peripheral blood cell count, liver function, plasma prothrombin time (PT), portal vein diameter, and appearance of gastroesophageal varices under gastroscopy were collected before and after embolization for statistical analysis. RESULTS: Five days after the operation, the numbers of leucocytes and platelets were significantly increased (P<0.05), and PT was significantly shortened (P<0.05). All the patients showed a good response after PSE with reduced internal diameter of the portal vein and blood flow (P<0.05). Gastroesophageal varices were relieved in all the patients. Rebleeding occurred in 11 patients during the follow-up. CONCLUSION: PSE can be effective in preventing gastroesophageal variceal rebleeding.