RESUMO
The Kv1.3 channel has been widely demonstrated to play crucial roles in the activation and proliferation of T cells, which suggests that selective blockers could serve as potential therapeutics for autoimmune diseases mediated by T cells. We previously described that the toxin mimic FS48 from salivary gland of Xenopsylla cheopis downregulates the secretion of proinflammatory factors by Raw 264.7 cells by blocking the Kv1.3 channel and the subsequent inactivation of the proinflammatory MAPK/NF-κB pathways. However, the effects of FS48 on human T cells and autoimmune diseases are unclear. Here, we described its immunomodulatory effects on human T cells derived from suppression of Kv1.3 channel. Kv1.3 currents in Jurkat T cells were recorded by whole-cell patch-clamp, and Ca2+ influx, cell proliferation, and TNF-α and IL-2 secretion were measured using Fluo-4, CCK-8, and ELISA assays, respectively. The in vivo immunosuppressive activity of FS48 was evaluated with a rat DTH model. We found that FS48 reduced Kv1.3 currents in Jurkat T cells in a concentration-dependent manner with an IC50 value of about 1.42 µM. FS48 also significantly suppressed Kv1.3 protein expression, Ca2+ influx, MAPK/NF-κB/NFATc1 pathway activation, and TNF-α and IL-2 production in activated Jurkat T cells. Finally, we show that FS48 relieved the DTH response in rats. We therefore conclude that FS48 can block the Kv1.3 channel and inhibit human T cell activation, which most likely contributes to its immunomodulatory actions and highlights the great potential of this evolutionary-guided peptide as a drug template in future studies.
Assuntos
Doenças Autoimunes , Canal de Potássio Kv1.3 , Venenos de Escorpião , Linfócitos T , Xenopsylla , Adjuvantes Imunológicos/farmacologia , Animais , Doenças Autoimunes/tratamento farmacológico , Doenças Autoimunes/imunologia , Humanos , Fatores Imunológicos/farmacologia , Interleucina-2/metabolismo , Canal de Potássio Kv1.3/imunologia , Ativação Linfocitária/efeitos dos fármacos , NF-kappa B/metabolismo , Bloqueadores dos Canais de Potássio/imunologia , Ratos , Glândulas Salivares/química , Venenos de Escorpião/farmacologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/imunologia , Xenopsylla/químicaRESUMO
Talaromycosis, namely Talaromyces marneffei infection, is increasing gradually and has a high mortality rate even under antifungal therapy. Although autophagy acts differently on different pathogens, it is a promising therapeutic strategy. However, information on autophagy in macrophages and animals upon infection by T. marneffei is still limited. Therefore, several models were employed here to investigate the role of autophagy in host defense against T. marneffei, including RAW264.7 macrophages as in vitro models, different types of Caenorhabditis elegans and BALB/c mice as in vivo models. We applied the clinical T. marneffei isolate SUMS0152 in this study. T. marneffei-infected macrophages exhibit increased formation of autophagosomes. Further, macrophage autophagy promoted by rapamycin or Earle's balanced salt solution (EBSS) inhibited the viability of intracellular T. marneffei. In vivo, compared with uninfected Caenorhabditis elegans, the wild-type nematodes upregulated the expression of the autophagy-related gene lgg-1 and atg-18, and nematodes carrying GFP reporter were induced to form autophagosomes (GFP::LGG-1) after T. marneffei infection. Furthermore, the knockdown of lgg-1 significantly reduced the survival rate of T. marneffei-infected nematodes. Likewise, the autophagy activator rapamycin reduced the fungal burden and suppressed lung inflammation in a mouse model of infection. In conclusion, autophagy is essential for host defense against T. marneffei in vitro and in vivo. Therefore, autophagy may be an attractive target for developing new therapeutics to treat talaromycosis.
Assuntos
Caenorhabditis elegans , Talaromyces , Animais , Camundongos , Autofagia , Sirolimo/farmacologiaRESUMO
The voltage-gated potassium (Kv) 1.3 channel plays a crucial role in the immune responsiveness of T-lymphocytes and macrophages, presenting a potential target for treatment of immune- and inflammation related-diseases. FS48, a protein from the rodent flea Xenopsylla cheopis, shares the three disulfide bond feature of scorpion toxins. However, its three-dimensional structure and biological function are still unclear. In the present study, the structure of FS48 was evaluated by circular dichroism and homology modeling. We also described its in vitro ion channel activity using patch clamp recording and investigated its anti-inflammatory activity in LPS-induced Raw 264.7 macrophage cells and carrageenan-induced paw edema in mice. FS48 was found to adopt a common αßß structure and contain an atypical dyad motif. It dose-dependently exhibited the Kv1.3 channel in Raw 264.7 and HEK 293T cells, and its ability to block the channel pore was demonstrated by the kinetics of activation and competition binding with tetraethylammonium. FS48 also downregulated the secretion of proinflammatory molecules NO, IL-1ß, TNF-α, and IL-6 by Raw 264.7 cells in a manner dependent on Kv1.3 channel blockage and the subsequent inactivation of the MAPK/NF-κB pathways. Finally, we observed that FS48 inhibited the paw edema formation, tissue myeloperoxidase activity, and inflammatory cell infiltrations in carrageenan-treated mice. We therefore conclude that FS48 identified from the flea saliva is a novel potassium channel inhibitor displaying anti-inflammatory activity. This discovery will promote understanding of the bloodsucking mechanism of the flea and provide a new template molecule for the design of Kv1.3 channel blockers.
Assuntos
Anti-Inflamatórios/farmacologia , Edema/tratamento farmacológico , Inflamação/tratamento farmacológico , Canal de Potássio Kv1.3/antagonistas & inibidores , Macrófagos/efeitos dos fármacos , Glândulas Salivares/metabolismo , Venenos de Escorpião/química , Animais , Edema/imunologia , Edema/metabolismo , Edema/patologia , Feminino , Humanos , Inflamação/imunologia , Inflamação/metabolismo , Inflamação/patologia , Macrófagos/imunologia , Macrófagos/metabolismo , Macrófagos/patologia , Masculino , Camundongos , NF-kappa B/metabolismo , Transdução de Sinais , Fator de Necrose Tumoral alfa/metabolismo , XenopsyllaRESUMO
Smp43, a cationic antimicrobial peptide identified from the venom gland of the Egyptian scorpion Scorpio maurus palmatus, shows cytotoxicity toward hepatoma cell line HepG2 by membrane disruption. However, its underlying detailed mechanisms still remain to be further clarified. In the present study, we evaluated the cellular internalization of Smp43 and explored its effects on cell viability, cell cycle, apoptosis, autophagy, necrosis, and factor expression related to these cellular processes in human HepG2. Smp43 was found to suppress the growth of HepG2, Huh7, and human primary hepatocellular carcinoma cells while showing low toxicity to normal LO2 cells. Furthermore, Smp43 could interact with the cell membrane and be internalized into HepG2 cells via endocytosis and pore formation, which caused a ROS production increase, mitochondrial membrane potential decline, cytoskeleton disorganization, dysregulation of cyclin expression, mitochondrial apoptotic pathway activation, and alteration of MAPK as well as PI3K/Akt/mTOR signaling pathways. Finally, Smp43 showed effective antitumor protection in the HepG2 xenograft mice model. Overall, these findings indicate that Smp43 significantly exerts antitumor effects via induction of apoptosis, autophagy, necrosis, and cell cycle arrest due to its induction of mitochondrial dysfunction and membrane disruption. This discovery will extend the antitumor mechanisms of antimicrobial peptides and contribute to the development of antitumor agents against hepatocellular carcinoma.
Assuntos
Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Mitocôndrias Hepáticas/efeitos dos fármacos , Peptídeos/farmacologia , Venenos de Escorpião/química , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Endocitose/efeitos dos fármacos , Humanos , Camundongos , Membranas Mitocondriais/efeitos dos fármacos , Peptídeos/química , Peptídeos/uso terapêutico , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
The inflammatory response plays important roles in acne vulgaris and pain pathogenesis. In previous study, Esc-1GN with anti-inflammatory, antimicrobial, and lipopolysacchride (LPS) binding activity was identified from the skin of the frog Hylarana guentheri. Here, we report its therapeutic potentials for acne vulgaris and inflammatory pain. Esc-1GN destroyed the cell membrane of Propionibacteria acnes in the membrane permeability assays. In addition, bacterial agglutination test suggested that Esc-1GN triggered the agglutination of P. acnes, which was affected by LPS and Ca2+ . Meanwhile, in vivo anti-P. acnes and anti-inflammatory effects of Esc-1GN were confirmed by reducing the counts of P. acnes in mice ear, relieving P. acnes-induced mice ear swelling, decreasing mRNA expression and the production of pro-inflammatory cytokines, and attenuating the infiltration of inflammatory cells. Moreover, Esc-1GN also displayed antinociceptive effect in mice induced by acetic acid and formalin. Therefore, Esc-1GN is a promising candidate drug for treatment of acne vulgaris and inflammatory pain.
Assuntos
Acne Vulgar/tratamento farmacológico , Antibacterianos/farmacologia , Anti-Inflamatórios não Esteroides/farmacologia , Inflamação/tratamento farmacológico , Dor/tratamento farmacológico , Proteínas Citotóxicas Formadoras de Poros/metabolismo , Propionibacterium acnes/efeitos dos fármacos , Acne Vulgar/microbiologia , Animais , Antibacterianos/síntese química , Antibacterianos/química , Anti-Inflamatórios não Esteroides/síntese química , Anti-Inflamatórios não Esteroides/química , Camundongos , Testes de Sensibilidade Microbiana , Proteínas Citotóxicas Formadoras de Poros/síntese química , Proteínas Citotóxicas Formadoras de Poros/químicaRESUMO
Scorpion venom is a potent natural source for antitumor drug development due to the multiple action modes of anticancer components. Although the sequence of Androcin 18-1 has been identified from the transcriptome profile of the scorpion venom Androctonus bicolor, its bioactivity remains unclear. In this study, we described the antitumor mechanism whereby Androcin 18-1 inhibits the proliferation and induces apoptosis by inducing cell membrane disruption, ROS accumulation, and mitochondrial dysfunction in human U87 glioblastoma cells. Moreover, Androcin 18-1 could suppress cell migration via the mechanisms associated with cytoskeleton disorganization and MMPs/TIMPs expression regulation. The discovery of this work highlights the potential application of Androcin 18-1 in drug development for glioblastoma treatment.
Assuntos
Antineoplásicos , Mitocôndrias , Venenos de Escorpião , Humanos , Venenos de Escorpião/farmacologia , Venenos de Escorpião/química , Linhagem Celular Tumoral , Mitocôndrias/metabolismo , Mitocôndrias/efeitos dos fármacos , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Animais , Proliferação de Células/efeitos dos fármacos , Glioblastoma/tratamento farmacológico , Glioblastoma/metabolismo , Movimento Celular/efeitos dos fármacos , Escorpiões , Peptídeos/farmacologiaRESUMO
Parkinson's disease (PD) is a neurodegenerative condition that results in dyskinesia, with oxidative stress playing a pivotal role in its progression. Antioxidant peptides may thus present therapeutic potential for PD. In this study, a novel cathelicidin peptide (Cath-KP; GCSGRFCNLFNNRRPGRLTLIHRPGGDKRTSTGLIYV) was identified from the skin of the Asiatic painted frog ( Kaloula pulchra). Structural analysis using circular dichroism and homology modeling revealed a unique αßß conformation for Cath-KP. In vitro experiments, including free radical scavenging and ferric-reducing antioxidant analyses, confirmed its antioxidant properties. Using the 1-methyl-4-phenylpyridinium ion (MPP +)-induced dopamine cell line and 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced PD mice, Cath-KP was found to penetrate cells and reach deep brain tissues, resulting in improved MPP +-induced cell viability and reduced oxidative stress-induced damage by promoting antioxidant enzyme expression and alleviating mitochondrial and intracellular reactive oxygen species accumulation through Sirtuin-1 (Sirt1)/Nuclear factor erythroid 2-related factor 2 (Nrf2) pathway activation. Both focal adhesion kinase (FAK) and p38 were also identified as regulatory elements. In the MPTP-induced PD mice, Cath-KP administration increased the number of tyrosine hydroxylase (TH)-positive neurons, restored TH content, and ameliorated dyskinesia. To the best of our knowledge, this study is the first to report on a cathelicidin peptide demonstrating potent antioxidant and neuroprotective properties in a PD model by targeting oxidative stress. These findings expand the known functions of cathelicidins, and hold promise for the development of therapeutic agents for PD.
Assuntos
Discinesias , Fármacos Neuroprotetores , Doença de Parkinson , Animais , Camundongos , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/farmacologia , 1-Metil-4-Fenil-1,2,3,6-Tetra-Hidropiridina/uso terapêutico , 1-Metil-4-fenilpiridínio/farmacologia , 1-Metil-4-fenilpiridínio/uso terapêutico , Antioxidantes/farmacologia , Antioxidantes/metabolismo , Catelicidinas/metabolismo , Discinesias/tratamento farmacológico , Discinesias/veterinária , Fármacos Neuroprotetores/farmacologia , Fármacos Neuroprotetores/uso terapêutico , Estresse Oxidativo , Doença de Parkinson/veterináriaRESUMO
FS145, a protein containing a WGD motif, was previously described from the salivary transcriptome of the flea Xenopsylla cheopis. Nevertheless, its biological function and complete structure are still uncertain. Herein, FS145 was confirmed to adopt a common αßß structure with the WGD motif exposed on its surface and located right at the top of a loop composed of residues 72-81. Furthermore, FS145 dose-dependently inhibited the proliferation, adhesion, migration, and tube formation of HUVECs by not only binding to integrin αvß3 but also by subsequently inactivating the FAK/Src/MAPK pathway along with the reduction of the expression of MMP-2, MMP-9, VEGFA, bFGF, Ang2, Tie2, HIF-1α, and FAK. Moreover, FS145 also inhibited aortic vessel sprout and showed strong anti-angiogenic activities as assessed ex vivo, by employing the rat aortic ring assay, chick embryo chorioallantoic membrane, and zebrafish embryo models. Altogether, our results suggest that FS145 suppresses angiogenesis ex vivo and in vitro by blocking integrin αvß3. The current study reveals the first anti-angiogenesis disintegrin with WGD motif from invertebrates and provides a beneficial pharmacological activity to inhibit abnormal angiogenesis.
Assuntos
Desintegrinas , Sifonápteros , Embrião de Galinha , Ratos , Animais , Desintegrinas/farmacologia , Desintegrinas/química , Integrina alfaVbeta3/metabolismo , Sifonápteros/metabolismo , Angiogênese , Peixe-Zebra/metabolismo , Células Cultivadas , Neovascularização Fisiológica , Movimento Celular , Inibidores da Angiogênese/farmacologia , Inibidores da Angiogênese/químicaRESUMO
Protease inhibitors regulate various biological processes and prevent host tissue/organ damage. Specific inhibition/regulation of proteases is clinically valuable for treating several diseases. Psoriasis affects the skin in the limbs and scalp of the body, and the contribution of cysteine and serine proteases to the development of skin inflammation is well documented. Cysteine protease inhibitors from ticks have high specificity, selectivity, and affinity to their target proteases and are efficient immunomodulators. However, their potential therapeutic effect on psoriasis pathogenesis remains to be determined. Therefore, we tested four tick cystatins (Sialostatin L, Sialostatin L2, Iristatin, and Mialostatin) in the recently developed, innate immunity-dependent mannan-induced psoriasis model. We explored the effects of protease inhibitors on clinical symptoms and histological features. In addition, the number and percentage of immune cells (dendritic cells, neutrophils, macrophages, and γδT cells) by flow cytometry, immunofluorescence/immunohistochemistry and, the expression of pro-inflammatory cytokines (TNF-a, IL-6, IL-22, IL-23, and IL-17 family) by qPCR were analyzed using skin, spleen, and lymph node samples. Tick protease inhibitors have significantly decreased psoriasis symptoms and disease manifestations but had differential effects on inflammatory responses and immune cell populations, suggesting different modes of action of these inhibitors on psoriasis-like inflammation. Thus, our study demonstrates, for the first time, the usefulness of tick-derived protease inhibitors for treating skin inflammation in patients.
Assuntos
Dermatite , Psoríase , Humanos , Inibidores de Cisteína Proteinase , Mananas , Psoríase/induzido quimicamente , Psoríase/tratamento farmacológico , Inflamação/tratamento farmacológico , Inibidores de Proteases , Imunidade Inata , Endopeptidases , Peptídeo HidrolasesRESUMO
Platelet activation contributes to sepsis development, leading to microthrombosis and increased inflammation, which results in disseminated intravascular coagulation and multiple organ dysfunction. Although Cathelicidin can alleviate sepsis, its role in sepsis regulation remains largely unexplored. In this study, we identified Cath-HG, a novel Cathelicidin from Hylarana guentheri skin, and analyzed its structure using nuclear magnetic resonance spectroscopy. The modulatory effect of Cath-HG on the symptoms of mice with sepsis induced by cecal ligation and puncture was evaluated in vivo, and the platelet count, degree of organ damage, and microthrombosis were measured. The antiplatelet aggregation activity of Cath-HG was studied in vitro, and its target was verified. Finally, we further investigated whether Cath-HG could regulate thrombosis in vivo in a FeCl3 injury-induced carotid artery model. The results showed that Cath-HG exhibited an α-helical structure in sodium dodecyl sulfate solution and effectively reduced organ inflammation and damage, improving survival in septic mice. It alleviated sepsis-induced thrombocytopenia and microthrombosis. In vitro, Cath-HG specifically inhibited collagen-induced platelet aggregation and modulated glycoprotein VI (GPVI) signaling pathways. Dot blotting, enzyme-linked immunosorbent assay, and pull-down experiments confirmed GPVI as the target of Cath-HG. Molecular docking and amino acid residue truncations/mutations identified crucial sites of Cath-HG. These findings suggest that GPVI represents a promising therapeutic target for sepsis, and Cath-HG may serve as a potential treatment for sepsis-related thrombocytopenia and thrombotic events. Additionally, identifying Cath-HG as a GPVI inhibitor provides insights for developing novel antithrombotic therapies targeting platelet activation mediated by GPVI.
RESUMO
Antimicrobial peptide is one important component of the first protective barrier of organisms. They not only have potent antimicrobial activity which can protect the body from the invading pathogens, but also participate in the immune regulation of the body. In this study, a Brevinin-1 peptide named by Brevinin-1GHd was identified from Hoplobatrachus rugulosus, and the similarity of mature peptide sequence among Brevinin-1GHd, Brevinin-1HL and Brevinin-1GHa supported the close species relationship between H. rugulosus, Hylarana latouchii and Hylarana guertheri. Moreover, the secondary structure of Brevinin-1GHd was found to possess α-helical characteristics and high thermal stability. In addition, Brevinin-1GHd could bind to LPS with a Kd value of 6.49 ± 5.40 mM and suppress the release of TNF-α, NO, IL-6 and IL-1ß by inactivation of MAPK signaling pathway in RAW 264.7 cells induced by LPS. Furtherly, Brevinin-1GHd had a significant inhibitory effect on acute edema development in the right paw of mice injected by carrageenan. Thus, the significant LPS-neutralizing and anti-inflammatory activities of Brevinin-1GHd were demonstrated in this study, which made it become the first Brevinin-1 family peptide with anti-inflammatory activity reported so far, and the biological activity of Brevinin-1GHd made it promising to be a novel therapeutic drug for infectious inflammation.
RESUMO
Septic shock caused by Gram-positive bacteria continues to be a major cause of morbidity and mortality in intensive care units globally. Most Temporins are excellent growth inhibitors of gram-positive bacteria and candidates for developing antimicrobial treatments due to their biological action and small molecular weight. In this study, a novel Temporin peptide from the skin of Fejervarya limnocharis frog, named as Temporin-FL, was characterized. Temporin-FL was found to adopt typical α-helical conformation in SDS solution and to exhibit selective antibacterial activity against Gram-positive bacteria through a membrane destruction mechanism. Accordingly, Temporin-FL showed protective effects against Staphylococcus aureus-induced sepsis in mice. Finally, Temporin-FL was demonstrated to exert anti-inflammatory effects by neutralizing the action of LPS/LTA and by inhibiting MAPK pathway activation. Therefore, Temporin-FL represents a novel candidate for moleculartherapy of Gram-positive bacterial sepsis.
Assuntos
Anti-Infecciosos , Choque Séptico , Animais , Camundongos , Lipopolissacarídeos/toxicidade , Sequência de Aminoácidos , Proteínas de Anfíbios/farmacologia , Proteínas de Anfíbios/uso terapêutico , Proteínas de Anfíbios/química , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/uso terapêutico , Anti-Infecciosos/farmacologia , Anti-Infecciosos/uso terapêutico , Anti-Infecciosos/química , Antibacterianos/farmacologia , Antibacterianos/uso terapêutico , Antibacterianos/metabolismo , Ranidae/metabolismo , Pele , Bactérias Gram-Positivas , Choque Séptico/metabolismo , Testes de Sensibilidade MicrobianaRESUMO
Wound healing is a complex process and remains a considerable challenge in clinical trials due to the lack of ideal therapeutic drugs. Here, a new peptide TK-HR identified from the skin of the frog Hoplobatrachus rugulosus was tested for its ability to heal cutaneous wounds in mice. Topical application of TK-HR at doses of 50-200 µg/mL significantly accelerated wound closure without causing any adverse effects in the animals. In vitro and in vivo investigations proved the regulatory role of the peptide on neutrophils, macrophages, keratinocytes, and vein endothelial cells involved in the inflammatory, proliferative, and remodeling phases of wound healing. Notably, TK-HR activated the MAPK and TGF-ß-Smad signaling pathways by acting on NK1R in RAW264.7 cells and mice. The current work has identified that TK-HR is a potent wound healing regulator that can be applied for the treatment of wounds, including diabetic foot ulcers and infected wounds, in the future.
Assuntos
Células Endoteliais , Receptores da Neurocinina-1 , Camundongos , Animais , Receptores da Neurocinina-1/metabolismo , Pele/metabolismo , Cicatrização , Peptídeos/farmacologia , Medicina TradicionalRESUMO
Chansu, a mixture extracted from Duttaphrynus melanostictus or Bufo gargarizans Cantor, is a traditional Chinese medicine with a broad range of medical applications. However, the peptides/proteins in it have not received adequate attention. Herein, a Cathelicidin-DM-derived peptide named Cath-DM-NT was identified from the skin of D. melanostictus. Previous studies have shown that Cathelicidin-DM has significant antibacterial activity, while Cath-DM-NT has no antibacterial activity. In this study, Cath-DM-NT is found to have lectin-like activity which can agglutinate erythrocytes and bacteria, and bind to lipopolysaccharide (LPS). In addition, Cath-DM-NT has antioxidant activity, which can scavenge 2,2'-azino-bis(3-ethylbenzothiazoline-6-sulphonic acid) (ABTS), 1,1-diphenyl-2-picrylhydrazyl (DPPH), and nitric oxide (NO) radicals and reduce Fe3+. Consistently, Cath-DM-NT can protect PC12 cells from H2O2-induced oxidative damage and carrageenan-induced paw edema, reduce malondialdehyde (MDA) and reactive oxygen species (ROS) accumulation, and restore superoxide dismutase (SOD) and glutathione (GSH) levels. Our study suggests that Cath-DM-NT can serve as a lead compound for the development of drugs with dual lectin and antioxidant effects.
Assuntos
Antioxidantes , Catelicidinas , Ratos , Animais , Antioxidantes/farmacologia , Antioxidantes/química , Lectinas/farmacologia , Peróxido de Hidrogênio/farmacologia , Glutationa , BufonidaeRESUMO
Research has been conducted to investigate the potential application of scorpion venom-derived peptides in cancer therapy. Smp43, a cationic antimicrobial peptide from Scorpio maurus palmatus venom, has been found to exhibit suppressive activity against the proliferation of multiple cancer cell lines. However, its impact on non-small-cell lung cancer (NSCLC) cell lines has not been previously investigated. This study aimed to determine the cytotoxicity of Smp43 towards various NSCLC cell lines, particularly A549 cells with an IC50 value of 2.58 µM. The results indicated that Smp43 was internalized into A549 cells through membranolysis and endocytosis, which caused cytoskeleton disorganization, a loss of mitochondrial membrane potential, an accumulation of reactive oxygen species (ROS), and abnormal apoptosis, cell cycle distribution, and autophagy due to mitochondrial dysfunction. Additionally, the study explored the in vivo protective effect of Smp43 in xenograft mice. The findings suggest that Smp43 has potential anticarcinoma properties exerted via the inducement of cellular processes related to cell membrane disruption and mitochondrial dysfunction.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Neoplasias Pulmonares , Humanos , Animais , Camundongos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/metabolismo , Carcinoma Pulmonar de Células não Pequenas/patologia , Neoplasias Pulmonares/metabolismo , Células A549 , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/uso terapêutico , Peptídeos Catiônicos Antimicrobianos/metabolismo , Mitocôndrias/metabolismo , Apoptose , Espécies Reativas de Oxigênio/metabolismo , Proliferação de Células , Linhagem Celular Tumoral , Potencial da Membrana MitocondrialRESUMO
OBJECTIVE: Acute lung injury (ALI) is an acute clinical syndrome characterized by uncontrolled inflammation response, which causes high mortality and poor prognosis. The present study determined the protective effect and underlying mechanism of Periplaneta americana extract (PAE) against lipopolysaccharide (LPS)-induced ALI. METHODS: The viability of MH-S cells was measured by MTT. ALI was induced in BALB/c mice by intranasal administration of LPS (5 mg/kg), and the pathological changes, oxidative stress, myeloperoxidase activity, lactate dehydrogenase activity, inflammatory cytokine expression, edema formation, and signal pathway activation in lung tissues and bronchoalveolar lavage fluid (BALF) were examined by H&E staining, MDA, SOD and CAT assays, MPO assay, ELISA, wet/dry analysis, immunofluorescence staining and Western blotting, respectively. RESULTS: The results revealed that PAE obviously inhibited the release of proinflammatory TNF-α, IL-6 and IL-1ß by suppressing the activation of MAPK/Akt/NF-κB signaling pathways in LPS-treated MH-S cells. Furthermore, PAE suppressed the neutrophil infiltration, permeability increase, pathological changes, cellular damage and death, pro-inflammatory cytokines expression, and oxidative stress upregulation, which was associated with its blockage of the MAPK/Akt/NF-κB pathway in lung tissues of ALI mice. CONCLUSION: PAE may serve as a potential agent for ALI treatment due to its anti-inflammatory and anti-oxidative properties, which correlate to the blockage of the MAPK/NF-κB and AKT signaling pathways.
Assuntos
Lesão Pulmonar Aguda , Periplaneta , Camundongos , Animais , Lipopolissacarídeos/toxicidade , NF-kappa B/metabolismo , Periplaneta/metabolismo , Proteínas Proto-Oncogênicas c-akt/metabolismo , Lesão Pulmonar Aguda/induzido quimicamente , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Inflamação/induzido quimicamente , Inflamação/tratamento farmacológico , Inflamação/metabolismo , Citocinas/metabolismo , Estresse Oxidativo , Camundongos Endogâmicos BALB CRESUMO
Acute pancreatitis (AP) is a serious inflammatory disorder and still lacks effective therapy globally. In this study, a novel Ranacyclin peptide, Ranacin, was identified from the skin of Pelophylax nigromaculatus frog. Ranacin adopted a compact ß-hairpin conformation with a disulfide bond (Cys5-Cys15). Ranacin was also demonstrated effectively to inhibit trypsin and have anticoagulant and antioxidant activities in vitro. Furthermore, the severity of pancreatitis was significantly alleviated in l-Arg-induced AP mice after treatment with Ranacin. In addition, structure-activity studies of Ranacin analogues confirmed that the sequences outside the trypsin inhibitory loop (TIL), especially at the C-terminal side, might be closely associated with the efficacy of its trypsin inhibitory activity. In conclusion, our data suggest that Ranacin can improve pancreatic injury in mice with severe AP through its multi-activity. Therefore, Ranacin is considered a potential drug candidate in AP therapy.
Assuntos
Pancreatite , Animais , Camundongos , Pancreatite/induzido quimicamente , Pancreatite/tratamento farmacológico , Antioxidantes/farmacologia , Antioxidantes/uso terapêutico , Doença Aguda , Tripsina , Anfíbios , Anticoagulantes/farmacologia , Anticoagulantes/uso terapêuticoRESUMO
Smp24, a cationic antimicrobial peptide identified from the venom gland of the Egyptian scorpion Scorpio maurus palmatus, shows variable cytotoxicity on various tumor (KG1a, CCRF-CEM and HepG2) and non-tumor (CD34+, HRECs, HACAT) cell lines. However, the effects of Smp24 and its mode of action on lung cancer cell lines remain unknown. Herein, the effect of Smp24 on the viability, membrane disruption, cytoskeleton, migration and invasion, and MMP-2/-9 and TIMP-1/-2 expression of human lung cancer cells have been evaluated. In addition, its in vivo antitumor role and acute toxicity were also assessed. In our study, Smp24 was found to suppress the growth of A549, H3122, PC-9, and H460 with IC50 values from about 4.06 to 7.07 µM and show low toxicity to normal cells (MRC-5) with 14.68 µM of IC50. Furthermore, Smp24 could induce necrosis of A549 cells via destroying the integrity of the cell membrane and mitochondrial and nuclear membranes. Additionally, Smp24 suppressed cell motility by damaging the cytoskeleton and altering MMP-2/-9 and TIMP-1/-2 expression. Finally, Smp24 showed effective anticancer protection in a A549 xenograft mice model and low acute toxicity. Overall, these findings indicate that Smp24 significantly exerts an antitumor effect due to its induction of membrane defects and cytoskeleton disruption. Accordingly, our findings will open an avenue for developing scorpion venom peptides into chemotherapeutic agents targeting lung cancer cells.
Assuntos
Neoplasias Pulmonares , Venenos de Escorpião , Animais , Peptídeos Catiônicos Antimicrobianos/farmacologia , Linhagem Celular Tumoral , Citoesqueleto , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Metaloproteinase 2 da Matriz , Camundongos , Venenos de Escorpião/farmacologia , Escorpiões , Inibidor Tecidual de Metaloproteinase-1RESUMO
BACKGROUND: Bacterial resistance to all currently available conventional antibiotics has caused a global public health crisis and led to an imperative search for new agents. Antimicrobial peptides (AMPs) are essential components of host innate immune defense against microbial invasions. OBJECTIVES: The objective of this study was to report a novel AMP, brevinin-2KP, from the skin of the black Kaloula pulchra frog and describe its structural and biological characterization. MATERIALS AND METHODS: The physical and chemical parameters of brevinin-2KP were predicted with the ExPASy Bioinformatics Resource Portal. The assembled sequences were aligned with ClustalW, and the phylogenetic tree was constructed using Mega. Circular dichroism (CD) experiments were carried out to identify the secondary structure and the stability of peptide in different solvent environments. The cytotoxicity of brevinin-2KP was evaluated by the MTT test. To determine antibacterial activity of brevinin- 2KP, a standard two-fold broth dilution method was used. SEM was carried out to observe the morphological change in the bacterial treated by brevinin-2KP. The live/dead bacterial viability was measured with a LIVE/DEAD® BacLight kit. Histamine release and mast cell degranulation assays were performed. RESULTS: The precursor of brevinin-2KP contains 72 amino acid residues, including a conserved signal peptide, acidic propeptide with KR residues, and mature peptide with a sequence of GVITDALKGAAKTVAAELLKKAHCKLTNSC. Phylogenetic analysis based on the amino acid sequences of 34 brevinin-2 peptides from 30 anuran species demonstrates that K. pulchra is genetically closely related to the genus Hylarana. The CD spectra analysis indicates that brevinin-2KP adopts random coil in the water and an organized α-helical conformation in SDS solution. Further, this secondary structure is stable under high salt and high-temperature conditions. Brevinin-2KP is weakly active towards the tested Gram-positive and Gram-negative bacteria as well as fungi due to its membranolytic action. Moreover, brevinin-2KP inhibits the proliferation of several mammal cells with IC50 values ranging from 3.27 to 59.75 µM. In addition, brevinin-2KP promotes degranulation and histamine release of mast cells, indicating that it is involved in the inflammatory response. CONCLUSION: This is the first report on AMP identified from the skin of K. pulchra. Brevinin-2KP adopts a typical amphipathic α-helix conformation in membrane mimic environment and shows antimicrobial and antitumor activities by potential membranolytic mechanism. In addition, brevinin-2KP can promote degranulation and histamine release of mast cells. Brevinin-2KP is expected to become a good drug temple molecule.
Assuntos
Anti-Infecciosos , Peptídeos Catiônicos Antimicrobianos , Animais , Aminoácidos , Antibacterianos/metabolismo , Anti-Infecciosos/farmacologia , Peptídeos Catiônicos Antimicrobianos/genética , Peptídeos Catiônicos Antimicrobianos/farmacologia , Peptídeos Catiônicos Antimicrobianos/química , Peptídeos Antimicrobianos , Anuros/metabolismo , Clonagem Molecular , Bactérias Gram-Negativas/metabolismo , Bactérias Gram-Positivas/metabolismo , Mamíferos/metabolismo , Testes de Sensibilidade Microbiana , Filogenia , Sinais Direcionadores de Proteínas , Pele/metabolismo , Solventes , ÁguaRESUMO
Scorpion-venom-derived peptides have become a promising anticancer agent due to their cytotoxicity against tumor cells via multiple mechanisms. The suppressive effect of the cationic antimicrobial peptide Smp24, which is derived from the venom of Scorpio Maurus palmatus, on the proliferation of the hepatoma cell line HepG2 has been reported earlier. However, its mode of action against HepG2 hepatoma cells remains unclear. In the current research, Smp24 was discovered to suppress the viability of HepG2 cells while having a minor effect on normal LO2 cells. Moreover, endocytosis and pore formation were demonstrated to be involved in the uptake of Smp24 into HepG2 cells, which subsequently interacted with the mitochondrial membrane and caused the decrease in its potential, cytoskeleton reorganization, ROS accumulation, mitochondrial dysfunction, and alteration of apoptosis- and autophagy-related signaling pathways. The protecting activity of Smp24 in the HepG2 xenograft mice model was also demonstrated. Therefore, our data suggest that the antitumor effect of Smp24 is closely related to the induction of cell apoptosis, cycle arrest, and autophagy via cell membrane disruption and mitochondrial dysfunction, suggesting a potential alternative in hepatocellular carcinoma treatment.