Inhibitory effects and mechanisms of the anti-covid-19 traditional Chinese prescription, Keguan-1, on acute lung injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Keguan-1, a new traditional Chinese medicine (TCM) prescription contained seven Chinese herbs, is developed to treat coronavirus disease 19 (COVID-19). The first internationally registered COVID-19 randomised clinical trial on integrated therapy demonstrated that Keguan-1 significantly reduced the incidence of ARDS and inhibited the severe progression of COVID-19. AIM OF THE STUDY: To investigate the protective mechanism of Keguan-1 on ARDS, a lipopolysaccharide (LPS)-induced acute lung injury (ALI) model was used to simulate the pathological state of ARDS in patients with COVID-19, focusing on its effect and mechanism on ALI. MATERIALS AND METHODS: Mice were challenged with LPS (2 mg/kg) by intratracheal instillation (i.t.) and were orally administered Keguan-1 (low dose, 1.25 g/kg; medium dose, 2.5 g/kg; high dose, 5 g/kg) after 2 h. Bronchoalveolar lavage fluid (BALF) and lung tissue were collected 6 h and 24 h after i.t. administration of LPS. The levels of inflammatory factors tumour necrosis factor alpha (TNF-α), interleukin (IL)-6, IL-1ß, keratinocyte-derived chemokine (KC or mCXCL1), macrophage inflammatory protein 2 (MIP2 or mCXCL2), angiotensin II (Ang II), and endothelial cell junction-associated proteins were analysed using ELISA or western blotting. RESULTS: Keguan-1 improved the survival rate, respiratory condition, and pathological lung injury; decreased the production of proinflammatory factors (TNF-α, IL-6, IL-1ß, KC, and MIP2) in BALF and the number of neutrophils in the lung tissues; and ameliorated inflammatory injury in the lung tissues of the mice with LPS-induced ALI. Keguan-1 also reduced the expression of Ang II and the adhesion molecule ICAM-1; increased tight junction proteins (JAM-1 and claudin-5) and VE-cadherin expression; and alleviated pulmonary vascular endothelial injury in LPS-induced ALI. CONCLUSION: These results demonstrate that Keguan-1 can improve LPS-induced ALI by reducing inflammation and pulmonary vascular endothelial injury, providing scientific support for the clinical treatment of patients with COVID-19. Moreover, it also provides a theoretical basis and technical support for the scientific use of TCMs in emerging infectious diseases.

Remote Inflammatory Preconditioning Alleviates Lipopolysaccharide-Induced Acute Lung Injury via Inhibition of Intrinsic Apoptosis in Rats.

BACKGROUND: Acute lung injury (ALI) always leads to severe inflammation. As inflammation and oxidative stress are the common pathological basis of endotoxin-induced inflammatory injury and ischemic reperfusion injury (IRI), we speculate that remote ischemic preconditioning (RIPC) can be protective for ALI when used as remote inflammatory preconditioning (RInPC). METHOD: A total of 21 Sprague-Dawley rats were used for the animal experiments. Eighteen rats were equally and randomly divided into the control (NS injection), LPS (LPS injection), and RInPC groups. The RInPC was performed prior to the LPS injection via tourniquet blockage of blood flow to the right hind limb and adopted three cycles of 5 min tying followed by 5 min untying. Animals were sacrificed 24 hours later. There were 2 rats in the LPS group and 1 in the RInPC group who died before the end of the experiment. Supplementary experiments in the LPS and RInPC groups were conducted to ensure that 6 animals in each group reached the end of the experiment. RESULTS: In the present study, we demonstrated that the RInPC significantly attenuated the LPS-induced ALI in rats. Apoptotic cells were reduced significantly by the RInPC, with the simultaneous improvement of apoptosis-related proteins. Reduction of MPO and MDA and increasing of SOD activity were found significantly improved by the RInPC. Increasing of TNF-α, IL-1ß, and IL-6 induced by the LPS was inhibited, while IL-10 was significantly increased by RInPC, compared to the LPS group. CONCLUSION: RInPC could inhibit inflammation and attenuate oxidative stress, thereby reducing intrinsic apoptosis and providing lung protection in the LPS-induced ALI in rats.

Baicalin Magnesium Salt Attenuates Lipopolysaccharide-Induced Acute Lung Injury via Inhibiting of TLR4/NF-κB Signaling Pathway.

Baicalin (BA) magnesium salt (BA-Mg) is a good water-soluble ingredient extracted from Scutellaria baicalensis Georgi, a commonly used traditional Chinese medicine. This study is aimed at investigating whether BA-Mg could exert a better protective effect on lipopolysaccharide- (LPS-) induced acute lung injury (ALI) in mice and illuminate the underlying mechanisms in vivo and in vitro. Mice were intraperitoneally administrated with equimolar BA-Mg, BA, and MgSO4 before LPS inducing ALI. Lung tissues and bronchoalveolar lavage fluid were collected for lung wet/dry ratio, histological examinations, cell counts, and biochemical analyses at 48 h post-LPS exposure. Meanwhile, the protein expressions of TLR4/NF-κB signaling pathway and proinflammatory cytokines in lung tissues and lung bronchial epithelial cells (BEAS-2B) were detected. The results showed BA-Mg pronouncedly ameliorated LPS-induced inflammatory response and histopathological damages, elevated antioxidant enzyme activity (SOD), and downregulated myeloperoxidase (MPO) and malonaldehyde (MDA) levels through the inhibition of TLR4/NF-κB signaling pathway activation. Moreover, the effect of BA-Mg was significantly better than that of BA and MgSO4 in ameliorating symptoms. Overall, BA-Mg can effectively relieve inflammatory response and oxidative stress triggered by LPS, indicating it may be a potential therapeutic candidate for treating ALI.

Oligosaccharides from Polygonatum Cyrtonema Hua: Structural characterization and treatment of LPS-induced peritonitis in mice.

Fructooligosaccharide was isolated from Polygonatum Cyrtonema Hua (PFOS) for the first time. Structure characterized using FT-IR, MALDI-TOF-MS, NMR, AFM, and TEM, indicated that PFOS was graminan-type fructan with a degree of polymerization ranging from 5 to 10. A murine model of lipopolysaccharide (LPS)-induced peritonitis was used to evaluate the in vivo anti-inflammatory and lung protective efficacy of PFOS. The result shown that pretreatment with PFOS (1.0 mg/mL) in peritonitis-induced mice could significantly inhibit the level of pro-inflammatory cytokines (TNF-α, IL-1ß) in serum (P < 0.001), increase mice survival rate from 12.5 % to 54 % (P < 0.05), and alleviated lung injury through ameliorating the damage of the pulmonary cellular architecture and reducing inflammatory monocyte accumulation in lung tissue. This effect of oligosaccharides could explain the traditional usage of P. cyrtonema as a tonic medicine for respiratory problems and it could be used as a potential natural ingredient with anti-inflammatory activity.

Carvacrol inhibits the excessive immune response induced by influenza virus A via suppressing viral replication and TLR/RLR pattern recognition.

ETHNOPHARMACOLOGICAL RELEVANCE: Carvacrol, a monoterpene phenol from Mosla chinensis Maxim, which is a commonly Chinese herbal medicine. The most important pharmacology of it is dispelling exogenous evils by increasing perspiration. And it is the gentleman medicine in the Chinese herbal compound prescription of Xin-Jia-Xiang-Ru-Yin, mainly for the treatment of summer colds with dampness including influenza virus A infection. AIM OF THE STUDY: Our preliminary study verified that the Xin-Jia-Xiang-Ru-Yin could inhibit acute lung injury of mice with influenza virus A infection. And there have been some reports implicating the high antimicrobial activity of carvacrol for a wide range of product preservation, but little research including the effects of it on viral infection. The aim of this study was to reveal the antiviral effects of carvacrol, the main constituent in Mosla chinensis Maxim. MATERIALS AND METHODS: Initially, C57BL/6 mice were grouped and intranasally administered FM1 virus to construct viral infection models. After treatment with ribavirin and carvacrol for 5 days, all mice were euthanized, and specimens were immediately obtained. Histology, flow cytometry and Meso Scale Discovery (MSD) analysis were used to analyze pathological changes in lung tissue, the expression levels of cytokines and the differentiation and proportion of CD4+ T cells subsets, while Western blot and qRT-PCR were used to detect the expression of related proteins and mRNA. RESULTS: Carvacrol attenuated lung tissue damage, the proportions of Th1, Th2, Th17 and Treg in CD4+ T cells and the relative proportions of Th1/Th2 and Th17/Treg cells. Carvacrol inhibited the expression of inflammation-associated cytokines including IFN-γ, IL-2, IL-4, IL-5, IL-12 and TNF-ɑ, IL-1, IL-10, IL-6. Decreased levels of TLR7, MyD88, IRAK4, TRAK6, NF-κB, RIG-I, IPS-I and IRF mRNA in carvacrol-treated mice were observed comparing to the mice in VC group. Further, the total expression of RIG-I, MyD88 and NF-κB proteins had increased significantly in the VC group but reduced obviously in the group treated with ribavirin or carvacrol. CONCLUSIONS: These results indicate that carvacrol is a potential alternative treatment for the excessive immune response induced by influenza virus A infection, the cold-fighting effect of Mosla chinensis Maxim may depend on the anti-virus of carvacrol.

Electroacupuncture pre­conditioning protects from lung injury induced by limb ischemia/reperfusion through TLR4 and NF­κB in rats.

Limb ischemia/reperfusion (I/R) can induce inflammation, causing acute lung injury. The Toll­like receptor 4 (TLR4)/NF­κB pathway plays an important role in acute and chronic inflammatory disorders. Several studies have demonstrated the efficacy of acupuncture in lung inflammatory injury. The aim of the present study was to elucidate the mechanism underlying the protective effect of electroacupuncture (EA) against lung injury induced by limb I/R. EA applied at the Zusanli and Sanyinjiao acupoints attenuated lung injury and decreased the secretion of inflammatory factors such as tumor necrosis factor­α, interleukin (IL)­1, IL­6 and myeloperoxidase. Moreover, the expression levels of TLR4 and NF­κB were suppressed by EA. Thus, the present findings suggested that EA can reduce pulmonary inflammation induced by limb I/R injury, possibly via the inhibition of the TLR4/NF­κB pathway.

Rugosic acid A, derived from Rosa rugosa Thunb., is novel inhibitory agent for NF-κB and IL-6/STAT3 axis in acute lung injury model.

Rosa rugosa Thunb., is as a medicinal plant known for anti-diabetic, and anti-inflammatory activities. However, the specific active compounds responsible for the individual pharmacological effects of in R. rugosa extract (95% EtOH) remain unknown. Here, we hypothesized that terpenoid structure, the most abundant constituents in R. rugosa extract, are responsible for its anti-inflammatory activity. We investigated the phytochemical substituents (compounds 1-13) and newly purified 11-methoxy polisin A, and 13-methoxy bisaborosaol F using NMR and ESI-MS and to screened their effects on NO production in LPS-induced macrophages. Rugosic acid A (RA) induced to ameliorate NO production, iNOS, and pro-inflammatory cytokines associated with the NF-κB. And, RA suppressed IL-6 secretion and IL-6-mediated STAT3 activation in LPS-mediated inflammation. In addition, RA was evaluated in LPS-mediated acute lung injury (ALI) model similar to acute pneumonia. Our results suggested that RA was suppressed to translocate nuclear NF-κB and IL-6-mediated STAT3 activation. Finally, RA led to amelioration of ALI by decreasing myeloperoxidase (MPO) and inhibiting phosphorylation of NF-κB and STAT3. Our group originally found that R. rugosa extract had new methoxy compounds and RA may be alternative natural agent for acute pneumonia similar to severe acute respiratory syndrome by coronavirus.

Inhibition of a triggering receptor expressed on myeloid cells-1 (TREM-1) with an extracellular cold-inducible RNA-binding protein (eCIRP)-derived peptide protects mice from intestinal ischemia-reperfusion injury.

BACKGROUND: Intestinal ischemia-reperfusion injury results in morbidity and mortality from both local injury and systemic inflammation and acute lung injury. Extracellular cold-inducible RNA-binding protein is a damage associated molecular pattern that fuels systemic inflammation and potentiates acute lung injury. We recently discovered a triggering receptor expressed on myeloid cells-1 serves as a novel receptor for extracellular cold-inducible RNA-binding protein. We developed a 7-aa peptide, named M3, derived from the cold-inducible RNA-binding protein, which interferes with cold-inducible RNA-binding protein's binding to a triggering receptor expressed on myeloid cells-1. Here, we hypothesized that M3 protects mice against intestinal ischemia-reperfusion injury. METHODS: Intestinal ischemia was induced in C57BL/6 mice via clamping of the superior mesenteric artery for 60 minutes. At reperfusion, mice were treated intraperitoneally with M3 (10 mg/kg body weight) or normal saline vehicle. Mice were killed 4 hours after reperfusion and blood and lungs were collected for various analysis. A 24-hours survival after intestinal ischemia-reperfusion was assessed. RESULTS: Serum levels of organ injury markers aspartate aminotransferase, alanine aminotransferase, lactate dehydrogenase, and lactate were increased with intestinal ischemia-reperfusion, while treatment with M3 significantly decreased their levels. Serum, intestinal, and lung levels of proinflammatory cytokines and chemokines were also increased by intestinal ischemia-reperfusion, and treatment with M3 significantly reduced these values. Intestinal ischemia-reperfusion caused significant histological intestinal and lung injuries, which were mitigated by M3. Treatment with M3 improved the survival from 40% to 80% after intestinal ischemia-reperfusion. CONCLUSION: Inhibition of triggering receptor expressed on myeloid cells-1 by an extracellular cold-inducible RNA-binding protein-derived small peptide (M3) decreased inflammation, reduced lung injury, and improved survival in intestinal ischemia-reperfusion injury. Thus, blocking the extracellular cold-inducible RNA-binding protein-triggering receptor expressed on myeloid cells-1 interaction is a promising therapeutic avenue for mitigating intestinal ischemia-reperfusion injury.

Isofraxidin ameliorated influenza viral inflammation in rodents via inhibiting platelet aggregation.

Platelets have been proved to exacerbate influenza infection and its complications. Inhibition of platelet activation may be a feasible method for preventing severe infection and secondary acute lung injury (ALI). Isofraxidin (IFD) is a natural coumarin isolated from the plants Sarcandra glabra and Siberian ginseng, and exerts anticancer, antioxidant and antiinflammatory effects. In the present study, we examined the therapeutic effects of IFD in ADP- or arachidonic acid (AA)-induced platelet aggregation model and in influenza A virus (IAV)-induced ALI mouse model. The results showed that IFD significantly inhibited platelet aggregation induced by ADP and AA in vitro in a concentration-dependent manner as well as the release of soluble P-selectin and platelet factor 4. Moreover, IFD significantly relieved IAV-induced lung inflammation, reduced the expressions of platelet activation biomarkers (P-selectin and CD61), decreased the serum levels of TNF-α, IL-1ß, IL-6 and MIP-2, suppressed peripheral platelet aggregation and prolonged the survival time of infected mice. The western blotting results also demonstrated that IFD reduced the phosphorylation levels of PI3K, AKT and p38 in the activated platelets stimulated by ADP and IAV infection. But IFD did not have any effects on IAV replication. It indicated that IFD ameliorated IAV-induced severe lung damage and lethal infection by suppressing platelet aggregation via regulating PI3K/AKT and MAPK pathways.

An untargeted metabolomics approach to investigate the wine-processed mechanism of Scutellariae radix in acute lung injury.

ETHNOPHARMACOLOGICAL RELEVANCE: Scutellariae radix (SR) is one of the most popular traditional Chinese medicines (TCM). Crude SR (CSR) and wine-processed SR (WSR) are the two most common commercial specifications. According to the theories of TCM, wine-processing increases the inclination and direction of SR's actions, thereby strengthening its efficacy in clearing the upper-energizer lung damp heat. The pharmacological mechanism-related research on WSR for the treatment of lung disease is limited and needs to be expanded. AIM OF THE STUDY: The aim of this report was to identify the relevant biological pathways by assessing changes in plasma metabolites between CSR and WSR in a lipopolysaccharide (LPS)-induced acute lung injury (ALI) model, and thus, revealed the potential mechanism of wine processing in SR. MATERIALS AND METHODS: Rats with LPS-induced ALI were treated with CSR and WSR. The contents of inflammatory cytokines and histopathological examination were determined to explore the effects of CSR and WSR. Next, the metabolic profiling of rat plasma samples was performed by ultra-performance liquid chromatography-quadrupole time-of-flight mass spectrometry (UPLC-QTOF-MS). Then, principal component analysis (PCA) were used to provide an overview for all of the groups and orthogonal partial least squares-discriminant analysis (OPLS-DA) was utilized to maximize the discrimination and present the differences in the metabolite between all of the groups. RESULTS: WSR exhibited a more remarkable effect on improving ALI than CSR by reducing the levels of inflammatory factors, including nitric oxide (NO), tumor necrosis factor alpha (TNF-α), interleukin-6 (IL-6), and interleukin-8 (IL-8). On the basis of UPLC-QTOF-MS technology, an unequal curative effect was revealed by nontargeting metabolomics. Sixteen biomarkers were discovered in the plasma of LPS-induced rats. Pathway analysis indicated that CSR acted on ALI by regulating the abnormal sphingolipid metabolism pathways; however, an WSR-mediated cure of ALI was linked primarily to reversing the abnormality of retinol metabolism pathways and tryptophan metabolism pathways. CONCLUSIONS: This report examined the underlying wine-processing mechanism of SR from the perspective of plasma metabolites. In addition, this work provided a novel and valuable insight into interpretation of the processing mechanisms of TCM in a holistic way.

Electroacupuncture Attenuates Limb Ischemia-Reperfusion-Induced Lung Injury Via p38 Mitogen-Activated Protein Kinase-Nuclear Factor Erythroid-2-Related Factor-2/Heme Oxygenase Pathway.

BACKGROUND: Electroacupuncture has been reported to protect the body from organ damages, but its mechanisms remain to be explored. This research was designed to investigate the function of electroacupuncture in lung injury resulted from hind limb ischemia-reperfusion (LIR) and whether p38 mitogen-activated protein kinase (p38 MAPK)-mediated nuclear factor erythroid-2-related factor-2 (Nrf2)/heme oxygenase (HO)-1 pathway contributes to the protective effect of electroacupuncture on LIR-originated lung damage. MATERIALS AND METHODS: Rabbits were subjected to occluding femoral artery for 2 h. Then they received reperfusion for 4 h to establish lung injury model. Electroacupuncture stimulation was performed bilaterally at Feishu and Zusanli acupoints for 15 min once a day for 5 d before the experiment and throughout the hind LIR model performing in the experimental day. Blood samples and lung tissues were collected to examine the role of electroacupuncture treatment in inflammatory response, oxidative stress, and lung injury. Both the protein expression and the messenger RNA level of Nrf2 and HO-1 were detected. RESULTS: The results showed that electroacupuncture treatment remarkably alleviated lung injury, decreased inflammatory cytokines secretion, attenuated lung oxidative stress, increased the amount of Nrf2 and HO-1, and increased the ratio of phospho-p38 MAPK to p38 MAPK after LIR. However, the protective effects exerted by electroacupuncture were reversed to some extent by the preconditioning with SB203580, a p38 MAPK-specific inhibitor. CONCLUSIONS: These results suggested that electroacupuncture could attenuate lung injury in rabbits subjected to LIR by inhibition of proinflammatory cytokine response and oxidative stress through activating p38 MAPK-mediated Nrf2/HO-1 pathway.

Oral administration of Ulmus davidiana extract suppresses interleukin-1ß expression in LPS-induced immune responses and lung injury.

BACKGROUND: Ulmus davidiana (UD) is a traditional Korean herb medicine that is used to treat inflammatory disorders. UD has been shown to modulate a number of inflammatory processes in vitro or in vivo studies. However, the molecular mechanisms of UD on lipopolysaccharide (LPS)-induced acute lung injury remain to be understood. OBJECTIVE: The primary objective of this study is to determine the effect of UD bark water extract on LPS-induced immune responses and lung injury using both in vitro and in vivo models. METHODS: RAW 264.7 cells and a rat model of acute lung injury (ALI) were used to study the effects of UD on several parameters. Nitrite level, lactate dehydrogenase (LDH) level, and superoxide dismutase (SOD) activities were measured. Tumor necrosis factor-α (TNF-α), interleukin-1ß (IL-1ß), and plasma transaminase activities in blood were also determined. Pathological investigations were also performed. RESULTS: LPS infusion resulted in elevated IL-1ß mRNA expression, nitrite levels, TNF-α expression, and IL-1ß expression in RAW 264.7 cells. LPS infusion also increased levels of nitrite/nitrate, total protein, LDH, and TNF-α in bronchoalveolar lavage fluid, but reduced SOD levels in ex vivo and in vivo models. UD administration ameliorated all these inflammatory markers. In particular, treatment with UD reduced LPS-induced nitrite production in RAW 264.7 cells in a dose-dependent manner. UD treatment also counteracted the LPS-induced increase in alanine aminotransferase (ALT) and aspartate transaminase (AST) activity in rat plasma, leading to a significant reduction in ALT and AST activity. CONCLUSIONS: The results revealed that UD treatment reduces LPS-induced nitrite production, IL-1ß mRNA expression, and TNF-α expression. In addition, LPS-induced decrease in SOD level is significantly elevated by UD administration. These results indicate that UD extract merits consideration as a potential drug for treating and/or preventing ALI.

p-Coumaric-Acid-Containing Adenostemma lavenia Ameliorates Acute Lung Injury by Activating AMPK/Nrf2/HO-1 Signaling and Improving the Anti-oxidant Response.

Adenostemma lavenia is a perennial herb belonging to the Compositae family and is widely distributed in the tropical parts of Asia. It has been widely used as medicine in Taiwan with the whole plant used to treat pulmonary congestion, pneumonia, bacterial infections of the respiratory tract, edema, and inflammation. This study sought to investigate the anti-inflammatory effects of A. lavenia in vitro and in animal models. The anti-inflammatory effects of ethyl acetate fractions of A. lavenia (EAAL) were stimulated with lipopolysaccharide (LPS) murine macrophages (RAW 264.7) and lung injury in mice. EAAL reduced proinflammatory cytokine responses. Preoral EAAL alleviated LPS-induced histological alterations in lung tissue and inhibited the infiltration of inflammatory cells and protein concentrations in bronchoalveolar lavage fluid (BALF). EAAL prevented protein expression of inducible NO synthase (iNOS) and cyclooxygenase-2 (COX-2); phosphorylation of IκB-α, MAPKs, and AMP-activated protein kinase (AMPK); and activated anti-oxidant enzymes (catalase, SOD, and GPx), heme oxygenase-1 (HO-1), and nuclear factor E2-related factor 2 (Nrf2) in LPS-stimulated cells and lung tissues. Fingerprinting of EAAL was performed with HPLC to control its quality, and p-coumaric acid was found to be a major constituent. This study suggests that EAAL is a potential therapeutic agent to treat inflammatory disorders.

Galloyl-Hexahydroxydiphenoyl (HHDP)-Glucose Isolated From Punica granatum L. Leaves Protects Against Lipopolysaccharide (LPS)-Induced Acute Lung Injury in BALB/c Mice.

The hydroalcoholic extract and ethyl acetate fraction of Punica granatum leaves have been known to exhibit anti-inflammatory activities. In this study, we investigated the therapeutic effects of galloyl-hexahydroxydiphenoyl (HHDP)-glucose isolated from pomegranate leaves on lipopolysaccharide (LPS)-induced acute lung injury (ALI) in mice. Male BALB/c mice were treated with different doses of galloyl-HHDP-glucose (5, 50, and 100 mg/Kg) or dexamethasone at 5 mg/Kg (per os) 6 h after intra-tracheal instillation of LPS. Vehicle-treated mice were used as controls. Twenty-four hours after LPS challenge, bronchoalveolar lavage fluid (BALF), and lung samples were collected for analyses. They were evaluated by monitoring the expression of NF-κB, JNK, and cytokine genes and proteins, as well as cell migration and lung function. All doses of galloyl-HHDP-glucose inhibited LPS-induced JNK and NF-κB activation. Likewise, the galloyl-HHDP-glucose-treated animals presented reduced expression of the TNF-α, IL-6, and IL-1ß genes in the lungs and reduced TNF-α, IL-6, IL-1ß, and IL-8 protein levels when compared with the vehicle-treated LPS-challenged mice. In addition, the ALI mice treated with galloyl-HHDP-glucose also presented reduced lung inflammatory cell accumulation, especially that of neutrophils, in their BALF and lungs. In addition, galloyl-HHDP-glucose treatment markedly ameliorated the LPS-induced pulmonary mechanism complications and attenuated weight loss. Overall, we showed for the first time that galloyl-HHDP-glucose protects against ALI, and may be useful for treating ALI and other inflammatory disorders.

Bee Pollen Extracts Modulate Serum Metabolism in Lipopolysaccharide-Induced Acute Lung Injury Mice with Anti-Inflammatory Effects.

Bee pollen (BP) collected from different floras possesses various potential bioactivities, but the mechanism-related research on anti-inflammatory effects is limited. Here, three types of BP originating from Camellia sinensis L. (BP-Cs), Nelumbo nucifera Gaertn. (BP-Nn), and Brassica campestris L. (BP-Bc) were assessed using molecular and metabolomics methods to determine their anti-inflammatory effects. The differences in polyphenolic abundance of three types of BP extracts were determined by HPLC-DAD/Q-TOF-MS. In vitro anti-inflammatory effects of three BP extracts were evaluated in a lipopolysaccharide (LPS)-induced RAW 264.7 cells model. BP-Cs extract with the most abundant polyphenols was found to be the most effective in reducing inflammation by downregulating inflammatory-related genes expression and blocking the activation of MAPK and NF-κB signaling pathways. Polyphenol-rich BP-Cs was further evaluated for their in vivo anti-inflammatory effect in a LPS-induced acute lung injury mouse model. An UPLC-Q-TOF/MS-based metabolomics approach was applied to analyze metabolite changes in mouse serum. Weshowed that the pretreated BP-Cs extract alleviated inflammation and regulated glycerophospholipid metabolism significantly. Our findings provide a foundation for developing and justifying BP as a potential anti-inflammatory ingredient in functional foods or nutraceutical formulations.

Protective effect of Jie-Geng-Tang against Staphylococcus aureus induced acute lung injury in mice and discovery of its effective constituents.

ETHNOPHARMACOLOGICAL RELEVANCE: Jie-Geng-Tang (JGT), a famous traditional Chinese medicine prescription, consists of Platycodonis Radix and Glycyrrhizae Radix et Rhizoma. According to traditional medicinal theory, JGT exerts various effects, including apocenosis, detoxifying, moisturizing the lung and relieving sore throat. It is often used to treat throat inflammation and lung diseases. AIM OF THE STUDY: To determine the protective effect of JGT on Staphylococcus aureus (S. aureus)-induced acute lung injury (ALI) in mice and to identify the compounds in the prescription that may be responsible for antibacterial activity. MATERIALS AND METHODS: The protective effect of JGT was assessed using S. aureus-induced ALI mice (i.g., 2.7 g/kg/day). Bacterial burden, pathological morphology, cytokine levels of TNF-α, IL-1ß, KC, and MIP-2 were evaluated in the lung and bronchoalveolar lavage fluid at 24 h post-infection, respectively. Twenty three compounds in the prescription were evaluated for their minimum inhibitory concentration (MIC) in vitro by means of microbroth dilution method against S. aureus. The antibacterial effects in vitro of licochalcone A and isoliquiritigenin were also investigated by transmission electron microscopy. In vivo antibacterial activities of licochalcone A and isoliquiritigenin were evaluated by survival rates, bacterial burden, and pathological morphology of lung tissues on S. aureus-induced ALI in mice (i.p., 160 mg/kg/day). RESULTS: Pretreatment with JGT significantly improved the pathological morphology of lung tissues on S. aureus-induced ALI in mice, accompanied with the reduced bacterial burden in the lungs and inhibited expression of inflammatory cytokine levels at 24 h post-infection. Five compounds, namely licochalcone A, licoisoflavone B, glyasperin A, isoliquiritigenin, and licochalcone B from Jie-Geng-Tang displayed good antibacterial activities against S. aureus (MIC < 128 µg/mL). Furthermore, applications of licochalcone A and isoliquiritigenin resulted in the increased survival rates, reduced bacterial burden in the lungs, and improved pathological morphology of lung tissues in S. aureus infected mice. CONCLUSION: The study demonstrated that Jie-Geng-Tang presented protective role of acute lung injury, which supported its traditional use for the treatment of lung diseases. Licochalcone A, isoliquiritigenin, licoisoflavone B, glyasperin A, and licochalcone B might contribute to the antibacterial activity of JGT on S. aureus-induced acute lung injury. The anti-S. aureus activity of licoisoflavone B, glyasperin A, and licochalcone B in vitro, as well as the anti-S. aureus activity of licochalcone A in vivo, were first reported in this study.

Dachengqi decoction alleviates acute lung injury and inhibits inflammatory cytokines production through TLR4/NF-κB signaling pathway in vivo and in vitro.

BACKGROUND AND OBJECTIVES: Sepsis that arises from uncontrolled pulmonary inflammation could induce acute lung injury (ALI), leading to the high death rate. Dachengqi decoction (DCQD) is a common traditional Chinese herbal medicine with strong anti-inflammatory effects. The current study aimed to explore the effect of DCQD on the inflammatory cytokines production, the aquaporin-1 (AQP-1) and AQP-5 protein expression in lipopolysaccharide (LPS)-induced ALI models, and the potential mechanisms underlying its effects. METHODS: Sprague-Dawley rats and HULEC-5a cells were used as study models in the research. To detect related molecules in the study, the real-time polymerase chain reaction analysis, cell counting kit-8 assay, Western blot analysis, and enzyme-linked immunosorbent assay were performed. RESULTS: DCQD could inhibit the expression of LPS-induced inflammatory cytokines, including interleukin-6 (IL-6), IL-8, and tumor necrosis factor-α (TNF-α), in lung tissues and could reduce pulmonary edema by upregulating the expression of AQP-1 and AQP-5 in rats with LPS-induced ALI. Moreover, the results suggested that the toll-like receptor 4 (TLR4)/NF-κB signaling is indispensable for DCQD to increase the expression of AQP-1 and AQP-5 and inhibits the production of IL-6, IL-8, and TNF-α in LPS-induced HULEC-5a cells. CONCLUSION: The results of our study suggested that DCQD suppresses the TLR4/NF-κB signaling pathway, increases the protein expression of AQP-1 and AQP-5, and inhibits the production of inflammatory cytokines, by which it may alleviate the inflammatory reactions in ALI and benefit the treatments.

Neutrophil elastase inhibitory effects of pentacyclic triterpenoids from Eriobotrya japonica (loquat leaves).

ETHNOPHARMACOLOGICAL RELEVANCE: Eriobotrya japonica, a traditional herbal medicine in China and Japan, has long been used to treat chronic bronchitis and coughs. AIM OF THE STUDY: Pentacyclic triterpenoids (PTs), especially ursolic acid (UA), have been found as reversibly and competitively human neutrophil elastase (HNE) inhibitors. However, the limited solubility and poor bioavailability of PTs hinder their clinical use. Crude plant extracts may have a greater activity than isolated constituents of the equivalent dosage. In this study, an Eriobotrya japonica (loquat leaves) extract (triterpenoid composition of loquat leaves, TCLL) with enriched PTs such as UA was prepared. The study aims to compare the HNE inhibitory (HNEI) effect in vitro and the therapeutic effect on acute lung injury (ALI) in vivo between TCLL and UA. MATERIALS AND METHODS: An HNEI activity bioassay was performed with Sivelestat sodium hydrate as a positive control. A lipopolysaccharide (LPS)-induced lung inflammatory model was established to evaluate TCLL's therapeutic effect on ALI in vivo. The absorption of UA in TCLL and in UA alone was determined using a Caco-2 cell uptake model and LC-MS. RESULTS: The IC50 values of TCLL and UA for the HNEI effect were 3.26 ±â€¯0.56 µg/mL and 8.49 ±â€¯0.42 µg/mL (P < 0.01), respectively. TCLL significantly improved the inflammatory cells and inflammatory cytokine production in mice compared with the LPS group (P < 0.05). Additionally, it performed better than the UA alone group (P < 0.05). Moreover, the uptake by Caco-2 cells of UA in TCLL was higher than that in UA alone (P < 0.05). CONCLUSION: TCLL has a significant HNEI effect in vitro and a therapeutic effect on LPS-induced inflammation in a mouse model. Both the effects are more efficient than UA. Improved absorption of PTs in TCLL may be one explanation for these results.

TRAF1 meditates lipopolysaccharide-induced acute lung injury by up regulating JNK activation.

Acute lung injury (ALI) is served as a severe life-threatening disease. However, the pathogenesis that contributes to ALI has not been fully understood. Tumor necrosis factor receptor-associated factor 1 (TRAF1) interacts with multiple regulators, performing its diverse role in biological functions. However, the effects of TRAF1 on ALI remain unknown. In this study, we attempted to explore the role of TRAF1 in ALI progression. The findings suggested that TRAF1-knockout (KO) markedly attenuated LPS-induced severe mortality rate in murine animals. LPS-elicited histological alterations in pulmonary tissues were significantly alleviated by TRAF1-deletion. Additionally, TRAF1 knockout effectively attenuated lung injury, as evidenced by the reduced lung wet/dry (W/D) weight ratio, as well as decreased bronchoalveolar lavage fluid (BALF) protein levels and neutrophil infiltration. Meanwhile, TRAF1 deletion markedly lessened inflammation, oxidative stress and apoptosis in BALF and/or lung tissues. The levels of pro-inflammatory cytokines stimulated by LPS were down-regulated by TRAF1 ablation, along with the inactivation of nuclear factor κB (NF-κB). LPS-promoted reactive oxygen species (ROS) generation was decreased in TRAF1-KO mice, partly through the improvement of anti-oxidants. Apoptosis was also inhibited by TRAF1 deletion in lung tissues of LPS-challenged mice through the suppression of cleaved Caspase-3. Moreover, TRAF1 knockout significantly decreased c-Jun N-terminal kinase (JNK) activation and its down-streaming signal of c-Jun in pulmonary samples of LPS-induced mice. Importantly, the in vitro study suggested that promoting JNK activation markedly abrogated TRAF1 knockdown-attenuated inflammation, ROS production and apoptosis in LPS-exposed A549 cells. Therefore, our experimental results provided evidence that TRAF1 suppression effectively protected LPS-induced ALI against inflammation, oxidative stress and apoptosis through the suppression of JNK activity.

The Nox1/Nox4 inhibitor attenuates acute lung injury induced by ischemia-reperfusion in mice.

Lung ischemia and reperfusion injury (LIRI) were mediated by several processes including over-production of reactive oxygen species (ROS) and inflammatory activation. ROS generated by nicotinamide adenine dinucletide phosphate (NADPH) oxidase (Nox) may play a pivotal role in pathophysiological changes in a range of disease. However, it was poorly understood in LIRI. Thus, the purpose of our study was to explore whether GKT137831, as a special dual inhibitor of Nox1 and 4, could alleviate LIRI in mice model and explore the minimal dose. According to the protocol, this study was divided into two parts. The first part was to determine the minimal dose of Nox1/4 inhibitor in attenuating LIRI via histopathology and apoptosis analysis. Eighteen C57BL/6J male wild-type mice were randomly divided in to sham, 2.5Nox+sham, 5.0Nox+sham, IR, 2.5Nox+IR and 5.0Nox+IR groups. According to the different group, mice were pretreated with corresponding dose of Nox1/4 inhibitors or normal saline. After LIRI, the results showed 5.0mg/kg Nox1/4 inhibitor could be considered as the minimal dose to alleviate injury by decreasing of lung injury score and the number of TUNEL-positive cells. The second part was to further verify the benefit of 5.0mg/kg Nox1/4 inhibitor in lung protective effects. Thirty-seven C57BL/6J male wild-type mice were divided in to sham, IR and 5.0Nox+IR groups randomly. The results showed that expressions of inflammatory, autophagy cytokines were markedly elevated and PH value was declined after LIRI. However, 5.0 mg/kg Nox1/4 inhibitor significantly attenuated cytokine production as reflected by immunohistochemistry, western blotting and Q-PCR analysis. In conclusion, our findings suggested that 5.0mg/kg Nox1/4 inhibitor contributed to protect lung tissue damage after LIRI via the suppression of inflammatory and autophagy activation.