Deacetyl Ganoderic Acid F Inhibits LPS-Induced Neural Inflammation via NF-κB Pathway Both In Vitro and In Vivo

Microglia mediated neuronal inflammation has been widely reported to be responsible for neurodegenerative disease. Deacetyl ganoderic acid F (DeGA F) is a triterpenoid isolated from Ganoderma lucidum, which is a famous edible and medicinal mushroom used for treatment of dizziness and insomnia in traditional medicine for a long time. In this study the inhibitory effects and mechanisms of DeGA F against lipopolysaccharide (LPS)-induced inflammation both in vitro and in vivo were investigated. On murine microglial cell line BV-2 cells, DeGA F treatment inhibited LPS-triggered NO production and iNOS expression and affected the secretion and mRNA levels of relative inflammatory cytokines. DeGA F inhibited LPS-induced activation of the NF-κB pathway, as evidenced by decreased phosphorylation of IKK and IκB and the nuclear translocation of P65. In vivo, DeGA F treatment effectively inhibited NO production in zebrafish embryos. Moreover, DeGA F suppressed the serum levels of pro-inflammatory cytokines, including TNF-α and IL-6 in LPS-stimulated mice model. DeGA F reduced inflammatory response by suppressing microglia and astrocytes activation and also suppressed LPS-induced NF-κB activation in mice brains. Taken together, DeGA F exhibited remarkable anti-inflammatory effects and promising therapeutic potential for neural inflammation associated diseases.

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Bullatine a Exerted Anti-Inflammatory Effects by Inhibiting JNK/ROS/NF-κB Pathway and Attenuates Systemic Inflammatory Response in LPS-Challenged Mice

10.21203/rs.3.rs-710437/v1 ◽

2021 ◽

Author(s):

Shuhan Liu ◽

Meichen Yan ◽

Yajin Liao ◽

Yong Cheng

Keyword(s):

Inflammatory Response ◽

Inflammatory Cytokines ◽

Nuclear Translocation ◽

Ros Generation ◽

Mrna Levels ◽

Iκb Kinase ◽

Inflammatory Mechanism ◽

Anti Inflammatory

Abstract Background: The genus Aconitum has rich pharmacological characteristics. Aconiti brachypodi Radix (Xue-shang-yi-zhi-hao) is a dried root of aconitum, which is considered to be analgesic and anti-inflammatory in modern medical and pharmaceutical clinical studies. Bullatine A (BA), a major active ingredient of this plant, has been reported for its significant anti-analgesic effect in previous studies. However, the role of BA in inflammation is unknown. In the current study, we aimed to explore the effect of BA on lipopolysaccharide (LPS)-induced inflammatory response both in vitro and in vivo and its potential anti-inflammatory mechanism.Materials and Methods: The anti-inflammatory effect of BA was evaluated in two different types of LPS-induced macrophages, including BV-2 microglial cells and immortalized murine bone marrow-derived macrophages (iBMDMs), and in acute inflammation mouse models induced by LPS. Immunofluorescence, flow cytometry, quantitative RT-PCR, western blot and Hematoxylin-Eosin staining were used to determine the anti-inflammatory properties of BA.Results: The results showed that BA significantly reduced the mRNA levels of several pro-inflammatory cytokines induced by LPS both in BV-2 cells and iBMDMs. Inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) in response to LPS were also decreased by BA. Further investigations indicated BA significantly blocked the phosphorylation of IκB kinase, degradation of the inhibitor IκBa and the nuclear translocation of nuclear factor-κB (NF-κB) p65. BA also reduced c-Jun N-terminal kinases (JNK) phosphorylation and ROS generation in iBMDMs activated with LPS, but had no effect on other mitogen-activated protein kinases (MAPKs) family proteins such as extracellular signal-regulated kinase (ERK) or p38. Furthermore, BA treatment alleviate liver and lung tissue damage, reduce inflammatory cell infiltration, and inhibit the expression of inflammatory cytokines in LPS-challenged mice.Conclusions: This study illustrated that BA has obvious anti-inflammatory effects both in vitro and in vivo, and its underlying anti-inflammatory mechanism may be via inactivating JNK/ROS/NF-κB pathway. Therefore, BA may have a certain therapeutic potential for inflammatory-related diseases.

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Inhibition of RANKL-Induced Osteoclastogenesis by Novel Mutant RANKL

International Journal of Molecular Sciences ◽

10.3390/ijms22010434 ◽

2021 ◽

Vol 22 (1) ◽

pp. 434

Author(s):

Yuria Jang ◽

Hong Moon Sohn ◽

Young Jong Ko ◽

Hoon Hyun ◽

Wonbong Lim

Keyword(s):

Nuclear Translocation ◽

Critical Role ◽

Osteoclast Differentiation ◽

Point Mutations ◽

Tartrate Resistant Acid Phosphatase ◽

Mice Model ◽

Gsk 3Β ◽

Rank Signaling

Background: Recently, it was reported that leucine-rich repeat-containing G-protein-coupled receptor 4 (LGR4, also called GPR48) is another receptor for RANKL and was shown to compete with RANK to bind RANKL and suppress canonical RANK signaling during osteoclast differentiation. The critical role of the protein triad RANK–RANKL in osteoclastogenesis has made their binding an important target for the development of drugs against osteoporosis. In this study, point-mutations were introduced in the RANKL protein based on the crystal structure of the RANKL complex and its counterpart receptor RANK, and we investigated whether LGR4 signaling in the absence of the RANK signal could lead to the inhibition of osteoclastogenesis.; Methods: The effects of point-mutated RANKL (mRANKL-MT) on osteoclastogenesis were assessed by tartrate-resistant acid phosphatase (TRAP), resorption pit formation, quantitative real-time polymerase chain reaction (qPCR), western blot, NFATc1 nuclear translocation, micro-CT and histomorphological assay in wild type RANKL (mRANKL-WT)-induced in vitro and in vivo experimental mice model. Results: As a proof of concept, treatment with the mutant RANKL led to the stimulation of GSK-3β phosphorylation, as well as the inhibition of NFATc1 translocation, mRNA expression of TRAP and OSCAR, TRAP activity, and bone resorption, in RANKL-induced mouse models; and Conclusions: The results of our study demonstrate that the mutant RANKL can be used as a therapeutic agent for osteoporosis by inhibiting RANKL-induced osteoclastogenesis via comparative inhibition of RANKL. Moreover, the mutant RANKL was found to lack the toxic side effects of most osteoporosis treatments.

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LINGO-1 shRNA protects the brain against ischemia/reperfusion injury by inhibiting the activation of NF-κB and JAK2/STAT3

Human Cell ◽

10.1007/s13577-021-00527-x ◽

2021 ◽

Author(s):

Jiaying Zhu ◽

Zhu Zhu ◽

Yipin Ren ◽

Yukang Dong ◽

Yaqi Li ◽

...

Keyword(s):

Cerebral Ischemia ◽

Nuclear Translocation ◽

Ischemia Reperfusion Injury ◽

Ischemia Reperfusion ◽

Glucose Deprivation ◽

Artery Occlusion ◽

Mice Model ◽

Down Regulation

AbstractLINGO-1 may be involved in the pathogenesis of cerebral ischemia. However, its biological function and underlying molecular mechanism in cerebral ischemia remain to be further defined. In our study, middle cerebral artery occlusion/reperfusion (MACO/R) mice model and HT22 cell oxygen–glucose deprivation/reperfusion (OGD/R) were established to simulate the pathological process of cerebral ischemia in vivo and in vitro and to detect the relevant mechanism. We found that LINGO-1 mRNA and protein were upregulated in mice and cell models. Down-regulation LINGO-1 improved the neurological symptoms and reduced pathological changes and the infarct size of the mice after MACO/R. In addition, LINGO-1 interference alleviated apoptosis and promoted cell proliferation in HT22 of OGD/R. Moreover, down-regulation of LINGO-1 proved to inhibit nuclear translocation of p-NF-κB and reduce the expression level of p-JAK2 and p-STAT3. In conclusion, our data suggest that shLINGO-1 attenuated ischemic injury by negatively regulating NF-KB and JAK2/STAT3 pathways, highlighting a novel therapeutic target for ischemic stroke.

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In Vitro Evaluation of the Anti-Inflammatory Effect of KMUP-1 and in Vivo Analysis of Its Therapeutic Potential in Osteoarthritis

Biomedicines ◽

10.3390/biomedicines9060615 ◽

2021 ◽

Vol 9 (6) ◽

pp. 615

Author(s):

Shang-En Huang ◽

Erna Sulistyowati ◽

Yu-Ying Chao ◽

Bin-Nan Wu ◽

Zen-Kong Dai ◽

...

Keyword(s):

Articular Cartilage ◽

Inflammatory Responses ◽

Therapeutic Potential ◽

Antioxidant Properties ◽

Serum Levels ◽

Gene Expressions ◽

Tnf Α ◽

Anti Inflammatory

Osteoarthritis is a degenerative arthropathy that is mainly characterized by dysregulation of inflammatory responses. KMUP-1, a derived chemical synthetic of xanthine, has been shown to have anti-inflammatory and antioxidant properties. Here, we aimed to investigate the in vitro anti-inflammatory and in vivo anti-osteoarthritis effects of KMUP-1. Protein and gene expressions of inflammation markers were determined by ELISA, Western blotting and microarray, respectively. RAW264.7 mouse macrophages were cultured and pretreated with KMUP-1 (1, 5, 10 μM). The productions of TNF-α, IL-6, MMP-2 and MMP- 9 were reduced by KMUP-1 pretreatment in LPS-induced inflammation of RAW264.7 cells. The expressions of iNOS, TNF-α, COX-2, MMP-2 and MMP-9 were also inhibited by KMUP-1 pretreatment. The gene expression levels of TNF and COX families were also downregulated. In addition, KMUP-1 suppressed the activations of ERK, JNK and p38 as well as phosphorylation of IκBα/NF-κB signaling pathways. Furthermore, SIRT1 inhibitor attenuated the inhibitory effect of KMUP-1 in LPS-induced NF-κB activation. In vivo study showed that KMUP-1 reduced mechanical hyperalgesia in monoiodoacetic acid (MIA)-induced rats OA. Additionally, KMUP-1 pretreatment reduced the serum levels of TNF-α and IL-6 in MIA-injected rats. Moreover, macroscopic and histological observation showed that KMUP-1 reduced articular cartilage erosion in rats. Our results demonstrated that KMUP-1 inhibited the inflammatory responses and restored SIRT1 in vitro, alleviated joint-related pain and cartilage destruction in vivo. Taken together, KMUP-1 has the potential to improve MIA-induced articular cartilage degradation by inhibiting the levels and expression of inflammatory mediators suggesting that KMUP-1 might be a potential therapeutic agent for OA.

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Hexarelin exerts neuroprotective and antioxidant effects against hydrogen peroxide-induced toxicity through the modulation of MAPK and PI3K/Akt patways in Neuro-2A cells

10.1101/2020.11.16.384321 ◽

2020 ◽

Author(s):

Ramona Meanti ◽

Laura Rizzi ◽

Elena Bresciani ◽

Laura Molteni ◽

Vittorio Locatelli ◽

...

Keyword(s):

Oxidative Stress ◽

Hydrogen Peroxide ◽

Apoptotic Cell ◽

Neuroblastoma Cell ◽

Neuroblastoma Cell Line ◽

Inhibitory Influence ◽

No Production ◽

Mrna Levels

AbstractHexarelin, a synthetic hexapeptide, protects cardiac and skeletal muscles by inhibiting apoptosis, both in vitro and in vivo. Moreover, evidence suggests that hexarelin could have important neuroprotective bioactivity.Oxidative stress and the generation of free radicals has been implicated in the etiologies of several neurodegenerative diseases, including amyotrophic lateral sclerosis, Parkinson’s disease, Alzheimer’s disease, Huntington’s disease and multiple sclerosis. In addition to direct oxidative stress, exogenous hydrogen peroxide (H2O2) can penetrate biological membranes and enhance the formation of other reactive oxygen species.The aim of this study was to examine the inhibitory influence of hexarelin on H2O2-induced apoptosis in Neuro-2A cells, a mouse neuroblastoma cell line. Our results indicate that H2O2 reduced the viability of Neuro-2A cells in a dose-related fashion. Furthermore, H2O2 induced significant changes in the morphology of Neuro-2A cells, reflected in the formation of apoptotic cell bodies, and an increase of nitric oxide (NO) production. Hexarelin effectively antagonized H2O2 oxidative damage to Neuro-2A cells as indicated by improved cell viability, normal morphology and reduced nitrite (NO2−) release. Hexarelin treatment of Neuro-2A cells also reduced mRNA levels of caspases−3 and −7 and those of the pro-apoptotic molecule Bax; by contrast, hexarelin treatment increased anti-apoptotic Bcl-2 mRNA levels. Hexarelin also reduced MAPKs phosphorylation induced by H2O2 and concurrently increased p-Akt protein expression.In conclusion, our results identify several neuroprotective and anti-apoptotic effects of hexarelin. These properties suggest that further investigation of hexarelin as a neuroprotective agent in an investigational and therapeutic context are merited.

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Targeting RNF8 Effectively Reverse Cisplatin and Doxorubicin Resistance in Endometrial Cancer

10.21203/rs.3.rs-57263/v1 ◽

2020 ◽

Author(s):

Ben Yang ◽

Wang Ke ◽

Yingchun Wan ◽

Tao Li

Keyword(s):

Endometrial Cancer ◽

Cell Lines ◽

Quantitative Polymerase Chain Reaction ◽

Mrna Levels ◽

Mice Model ◽

Expression Levels ◽

Gynecological Malignancy ◽

Ec Cells

Abstract Background Endometrial cancer (EC) is one of the most frequent gynecological malignancy worldwide. However, resistance to chemotherapy remains one of the major difficulties in the treatment of EC. Thus, there is an urgent requirement to understand mechanisms of chemoresistance and identify novel regimens for patients with EC. Methods Cisplatin and doxorubicin resistant cell lines were acquired by continuous exposing parental EC cells to cisplatin or doxorubicin for 3 months. Cell viability was determined by using MTT assay. Protein Expression levels of protein were examined by western blotting assay. mRNA levels were measured by quantitative polymerase chain reaction (qPCR) assay. Ring finger protein 8 (RNF8) knockout cell lines were generated by clustered regularly interspaced short palindromic repeats (CRISPR)–Cas9 gene editing assay. Nonhomologous end joining (NHEJ) efficiency were quantified by plasmid based NHEJ assay. DNA double strand breaks (DSB) were generated using laser micro-irradiation. Protein recruitment to DSB was analyzed by immunofluorescent assay. Tumor growth was examined by AN3CA xenograft mice model. Results We found that protein and mRNA expression levels of RNF8 were significantly increased in both cisplatin and doxorubicin resistant EC cells. Cell survival assay showed that RNF deficiency significantly enhanced the sensitivity of resistant EC cells to cisplatin and doxorubicin (P < 0.01). In addition, chemoresistant EC cells exhibited increased NHEJ efficiency. Knockout of RNF8 in chemoresistant EC cells significantly reduced NHEJ efficiency and prolonged Ku80 retention on DSB. Moreover, cisplatin resistant AN3CA xenograft showed that RNF8 deficiency overcame cisplatin resistance. Conclusions Our in vitro and in vivo assays provide evidence for RNF8, which is a NHEJ factor, serving as a promising, novel target in EC chemotherapy.

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Diesel particles increase phosphatidylcholine release through a NO pathway in alveolar type II cells

AJP Lung Cellular and Molecular Physiology ◽

10.1152/ajplung.00213.2001 ◽

2002 ◽

Vol 282 (5) ◽

pp. L1075-L1081 ◽

Cited By ~ 10

Author(s):

Philippe Juvin ◽

Thierry Fournier ◽

Martine Grandsaigne ◽

Jean-Marie Desmonts ◽

Michel Aubier

Keyword(s):

Diesel Exhaust Particles ◽

Major Constituent ◽

Alveolar Type ◽

No Production ◽

Mrna Levels ◽

Type Ii ◽

Diesel Particles ◽

Nos Inhibitor

Diesel exhaust particles (DEPs) have been shown in vivo as well as in vitro to affect the respiratory function and in particular the immune response to infection and allergens. In the current study, we investigated the effect of DEPs on the production of phosphatidylcholine (PC), a major constituent of surfactant, by rat alveolar type II (ATII) primary cells in vitro. Our results demonstrate that incubation of ATII cells with DEPs lead to a time- and dose-dependent increase in labeled PC release. This effect was mimicked by nitric oxide (NO) donors and cGMP and was abolished by inhibitors of NO synthase (NOS). In addition, a NOS inhibitor inhibits by itself the basal secretion of PC. We next examined the effects of DEPs on NOS gene expression and showed that DEPs increase NO production and upregulate both protein content and mRNA levels of the inducible NOS (NOS II). Together our data demonstrate that DEPs alter the production of surfactant by ATII cells through a NO-dependent signaling pathway.

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Nerolidol Mitigates Colonic Inflammation: An Experimental Study Using both In Vivo and In Vitro Models

Nutrients ◽

10.3390/nu12072032 ◽

2020 ◽

Vol 12 (7) ◽

pp. 2032

Author(s):

Vishnu Raj ◽

Balaji Venkataraman ◽

Saeeda Almarzooqi ◽

Sanjana Chandran ◽

Shreesh K. Ojha ◽

...

Keyword(s):

Nuclear Translocation ◽

In Vitro Models ◽

Mrna Levels ◽

Colonic Inflammation ◽

Tnf Α ◽

Cox 2 ◽

Anti Inflammatory ◽

Ht 29

Nerolidol (NED) is a naturally occurring sesquiterpene alcohol present in various plants with potent anti-inflammatory effects. In the current study, we investigated NED as a putative anti-inflammatory compound in an experimental model of colonic inflammation. C57BL/6J male black mice (C57BL/6J) were administered 3% dextran sodium sulfate (DSS) in drinking water for 7 days to induce colitis. Six groups received either vehicle alone or DSS alone or DSS with oral NED (50, 100, and 150 mg/kg body weight/day by oral gavage) or DSS with sulfasalazine. Disease activity index (DAI), colonic histology, and biochemical parameters were measured. TNF-α-treated HT-29 cells were used as in vitro model of colonic inflammation to study NED (25 µM and 50 µM). NED significantly decreased the DAI and reduced the inflammation-associated changes in colon length as well as macroscopic and microscopic architecture of the colon. Changes in tissue Myeloperoxidase (MPO) concentrations, neutrophil and macrophage mRNA expression (CXCL2 and CCL2), and proinflammatory cytokine content (IL-1β, IL-6, and TNF-α) both at the protein and mRNA level were significantly reduced by NED. The increase in content of the proinflammatory enzymes, COX-2 and iNOS induced by DSS were also significantly inhibited by NED along with tissue nitrate levels. NED promoted Nrf2 nuclear translocation dose dependently. NED significantly increased antioxidant enzymes activity (Superoxide dismutase (SOD) and Catalase (CAT)), Hemeoxygenase-1 (HO-1), and SOD3 mRNA levels. NED treatment in TNF-α-challenged HT-29 cells significantly decreased proinflammatory chemokines (CXCL1, IL-8, CCL2) and COX-2 mRNA levels. NED supplementation attenuates colon inflammation through its potent antioxidant and anti-inflammatory activity both in in vivo and in vitro models of colonic inflammation.

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A Limulus Antilipopolysaccharide Factor-Derived Peptide Exhibits a New Immunological Activity with Potential Applicability in Infectious Diseases

Clinical and Diagnostic Laboratory Immunology ◽

10.1128/cdli.7.4.669-675.2000 ◽

2000 ◽

Vol 7 (4) ◽

pp. 669-675 ◽

Cited By ~ 28

Author(s):

Maribel G. Vallespi ◽

Luis A. Glaria ◽

Osvaldo Reyes ◽

Hilda E. Garay ◽

Joel Ferrero ◽

...

Keyword(s):

Infectious Diseases ◽

Cyclic Peptides ◽

Mononuclear Cells ◽

Therapeutic Potential ◽

Peritoneal Macrophages ◽

Induced Response ◽

No Production ◽

Human Mononuclear Cells

ABSTRACT Previous studies have shown that cyclic peptides corresponding to residues 35 to 52 of the Limulus antilipopolysaccharide (anti-LPS) factor (LALF) bind and neutralize LPS-mediated in vitro and in vivo activities. Therapeutic approaches based on agents which bind and neutralize LPS activities are particularly attractive because these substances directly block the primary stimulus for the entire proinflammatory cytokine cascade. Here we describe new activities of the LALF31–52 peptide, other than its LPS binding ability. Surprisingly, supernatants from human mononuclear cells stimulated with the LALF peptide are able to induce in vitro antiviral effects on the Hep-2 cell line mediated by gamma interferon (IFN-γ) and IFN-α. Analysis of the effect of LALF31–52 on tumor necrosis factor (TNF) and nitric oxide (NO) production by LPS-stimulated peritoneal macrophages revealed that a pretreatment with the peptide decreased LPS-induced TNF production but did not affect NO generation. This indicates that the LALF peptide modifies the LPS-induced response. In a model in mice with peritoneal fulminating sepsis, LALF31–52 protected the mice when administered prophylactically, and this effect is related to reduced systemic TNF-α levels. This study demonstrates, for the first time, the anti-inflammatory properties of the LALF-derived peptide. These properties widen the spectrum of the therapeutic potential for this LALF-derived peptide and the molecules derived from it. These agents may be useful in the prophylaxis and therapy of viral and bacterial infectious diseases, as well as for septic shock.

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Wogonin induces G1 phase arrest through inhibiting Cdk4 and cyclin D1 concomitant with an elevation in p21Cip1 in human cervical carcinoma HeLa cells

Biochemistry and Cell Biology ◽

10.1139/o09-060 ◽

2009 ◽

Vol 87 (6) ◽

pp. 933-942 ◽

Cited By ~ 34

Author(s):

Li Yang ◽

Hai-wei Zhang ◽

Rong Hu ◽

Yong Yang ◽

Qi Qi ◽

...

Keyword(s):

Cyclin D1 ◽

Cervical Carcinoma ◽

Hela Cells ◽

Therapeutic Potential ◽

P53 Gene ◽

Mrna Levels ◽

Human Cervical Carcinoma ◽

Phase Arrest

Wogonin, a naturally occurring flavonoid, has been shown to have tumor therapeutic potential both in vitro and in vivo. To better understand its anticancer mechanism, we examined the effect of wogonin on human cervical carcinoma HeLa cells. In this study, we observed that G1 phase arrest was involved in wogonin-induced growth inhibition in HeLa cells. Over a 24 h exposure of HeLa cells to 90 µmol·L–1 wogonin, the promoters of G1–S transition, including cyclin D1/Cdk4 and pRb, decreased within 12 h and E2F-1 depleted in the nucleus at the same time. As the G1 phase arrest developed, p53 and the Cdk inhibitor p21Cip1 elevated both at protein and mRNA levels. Furthermore, the up-regulation of p21Cip1 induced by wogonin was dramatically inhibited by siRNA-mediated p53 gene silencing. Collectively, our data suggested that wogonin induced G1 phase arrest in HeLa cells by modulating several key G1 regulatory proteins, such as Cdk4 and cyclin D1, as well as up-regulation of a p53-midiated p21Cip1 expression. This mechanism of wogonin may play an important role in the killing of cancerous cells and offer a potential mechanism for its anticancer action in vivo.

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