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The E6 and E7 proteins of specific subtypes of human papillomavirus (HPV), including HPV 16 and 18, are highly associated with cervical cancer as they modulate cell cycle regulation. The aim of this study was to investigate the potential antitumor effects of a messenger RNA-HPV therapeutic vaccine (mHTV) containing nononcogenic E6 and E7 proteins. To achieve this, C57BL/6j mice were injected with the vaccine via both intramuscular and subcutaneous routes, and the resulting effects were evaluated. mHTV immunization markedly induced robust T cell-mediated immune responses and significantly suppressed tumor growth in both subcutaneous and orthotopic tumor-implanted mouse model, with a significant infiltration of immune cells into tumor tissues. Tumor retransplantation at day 62 postprimary vaccination completely halted progression in all mHTV-treated mice. Furthermore, tumor expansion was significantly reduced upon TC-1 transplantation 160 days after the last immunization. Immunization of rhesus monkeys with mHTV elicited promising immune responses. The immunogenicity of mHTV in nonhuman primates provides strong evidence for clinical application against HPV-related cancers in humans. All data suggest that mHTV can be used as both a therapeutic and prophylactic vaccine.
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Proteínas Oncogênicas Virais , Infecções por Papillomavirus , Vacinas contra Papillomavirus , Neoplasias do Colo do Útero , Humanos , Feminino , Animais , Camundongos , Papillomavirus Humano , Proteínas Oncogênicas Virais/genética , Infecções por Papillomavirus/prevenção & controle , RNA Mensageiro/genética , Proteínas E7 de Papillomavirus/genética , Camundongos Endogâmicos C57BL , Vacinação/métodos , Imunização , Neoplasias do Colo do Útero/prevenção & controleRESUMO
Chitin is mostly produced from crustaceans, but it is difficult to supply raw materials due to marine pollution, and the commonly used chemical chitin extraction method is not environmentally friendly. Therefore, this study aims to establish a chitin extraction process using enzymes and to develop edible insect-derived chitin as an eco-friendly new material. The response surface methodology (RSM) was used to determine the optimal conditions for enzymatic hydrolysis. The optimal conditions for enzymatic hydrolysis by RSM were determined to be the substrate concentration (7.5%), enzyme concentration (80 µL/g), and reaction time (24 h). The solubility and DDA of the mealworm chitosan were 45% and 37%, respectively, and those of the commercial chitosan were 61% and 57%, respectively. In regard to the thermodynamic properties, the exothermic peak of mealworm chitin was similar to that of commercial chitin. In the FT-IR spectrum, a band was observed in mealworm chitin corresponding to the C=O of the NHCOCH3 group at 1645 cm-1, but this band showed low-intensity C=O in the mealworm chitosan due to deacetylation. Collectively, mealworm chitosan shows almost similar physical and chemical properties to commercial chitosan. Therefore, it is shown that an eco-friendly process can be introduced into chitosan production by using enzyme-extracted mealworms for chitin/chitosan production.
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Quitina , Quitosana , Subtilisinas , Tenebrio , Animais , Acetilação , Varredura Diferencial de Calorimetria , Quitina/química , Quitina/isolamento & purificação , Quitina/metabolismo , Quitosana/química , Quitosana/isolamento & purificação , Quitosana/metabolismo , Crustáceos/química , Insetos Comestíveis/química , Insetos Comestíveis/metabolismo , Hidrólise , Proteólise , Solubilidade , Espectroscopia de Infravermelho com Transformada de Fourier , Subtilisinas/metabolismo , Tenebrio/química , Tenebrio/metabolismo , TermodinâmicaRESUMO
The CopA3 dimer peptide is a coprisin analog that has an anticancer effect against human cancer cells in vitro. In this study, we investigated the anticancer activity of the enantiomeric CopA3 dimer peptide in human gastric cancer cell lines as well as in an in vivo tumor xenograft model. Enantiomeric CopA3 reduced gastric cancer cell viability and exhibited cytotoxicity against cancer cells. Enantiomeric CopA3-induced cell death was mediated by specific interactions with phosphatidylserine and phosphatidylcholine, membrane components that are enriched in cancer cells, in a calcein leakage assay. Moreover, acridine orange/ethidium bromide staining, flow cytometric analysis, and Western blot analysis showed that enantiomeric CopA3 induced apoptotic and necrotic gastric cancer cell death. The antitumor effect was also observed in a mouse tumor xenograft model in which intratumoral inoculation of the peptide resulted in a significant decrease in the SNU-668 gastric cancer tumor volume. In addition, periodic acid-Schiff and hematoxylin staining and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL) assay revealed apoptotic and necrotic cell death in tumor masses treated with greater than 150 µg CopA3. Collectively, these results indicate that the enantiomeric CopA3 dimer peptide induces apoptosis and necrosis of gastric cancer cells in vitro and in vivo, indicating that the peptide is a potential candidate for the treatment of gastric cancer, which is a common cause of cancer and cancer deaths worldwide.
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Peptídeos Catiônicos Antimicrobianos/farmacologia , Proteínas de Insetos/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Ensaios Antitumorais Modelo de Xenoenxerto , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/química , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Western Blotting , Caspases/metabolismo , Linhagem Celular , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Células HeLa , Humanos , Proteínas de Insetos/química , Camundongos Endogâmicos BALB C , Camundongos Nus , Multimerização Proteica , Transdução de Sinais/efeitos dos fármacos , Estereoisomerismo , Neoplasias Gástricas/patologia , Carga Tumoral/efeitos dos fármacosRESUMO
BACKGROUND: No study to our knowledge evaluated the long-term survivorship of the contemporary cemented vs cementless stems more than 25 years of follow-up in the same patients younger than 50 years of age. PURPOSE OF STUDY: The purpose of the study is to determine (1) validated outcome scores, (2) prevalence of polyethylene wear and osteolysis, and (3) survivorship of the cemented vs cementless femoral components in the same young patients at minimum follow-up of 25 years. METHODS: We report 171 patients (mean age, 47.7 years) at the time of the surgery who received simultaneous bilateral total hip arthroplasty with a cemented stem in one hip and a cementless stem in the other (26.1-year follow-up). A cementless acetabular component was used in all hips. There were 125 men and 46 women. The mean age at the time of the arthroplasty was 47.7 ± 10.7 years (range, 21-50 years). RESULTS: The average Harris hip scores were similar between the groups of cemented stem (mean, 91 points) and cementless stem (93 points). Survival rate of the acetabular component (79% vs 78%) and that of the femoral component (96% vs 95%) at 26.1 years were similar between the 2 groups. CONCLUSIONS: The long-term fixation of the acetabular metallic shell and cemented or cementless femoral stem was outstanding. Polyethylene wear and periacetabular osteolysis were contributing factors to reduce the long-term survival of the contemporary total hip arthroplasties with or without cement in young patients. New bearing surfaces such as ceramic on ceramic or ceramic on highly cross-linked polyethylene would improve wear of the bearing surfaces.
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Artroplastia de Quadril/métodos , Fêmur/cirurgia , Prótese de Quadril , Acetábulo/cirurgia , Adulto , Materiais Biocompatíveis , Cimentos Ósseos , Cimentação , Feminino , Seguimentos , Humanos , Masculino , Pessoa de Meia-Idade , Polietileno , Prevalência , Falha de Prótese , Resultado do Tratamento , Adulto JovemRESUMO
The centipede Scolopendra subspinipes mutilans is an environmentally beneficial and medically important arthropod species. Although this species is increasingly applied as a reliable source of new antimicrobial peptides, the transcriptome of this species is a prerequisite for more rational selection of antimicrobial peptides. In this report, we isolated total RNA from the whole body of adult centipedes, S. subspinipes mutilans, that were nonimmunized and immunized against Escherichia coli, and we generated a total of 77,063 pooled contigs and singletons using high-throughput sequencing. To screen putative antimicrobial peptides, in silico analyses of the S. subspinipes mutilans transcriptome were performed based on the physicochemical evidence of length, charge, isoelectric point, and in vitro and in vivo aggregation scores together with the existence of continuous antimicrobial peptide stretches. Moreover, we excluded some transcripts that showed similarity with both previously known antimicrobial peptides and the human proteome, had a proteolytic cleavage site, and had downregulated expression compared with the nonimmunized sample. As a result, we selected 17 transcripts and tested their antimicrobial activity with a radial diffusion assay. Among them, ten synthetic peptides experimentally showed antimicrobial activity against microbes and no toxicity to mouse erythrocytes. Our results provide not only a useful set of antimicrobial peptide candidates and an efficient strategy for novel antimicrobial peptide development but also the transcriptome data of a big centipede as a valuable resource.
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Peptídeos Catiônicos Antimicrobianos/farmacologia , Proteínas de Artrópodes/farmacologia , Artrópodes/genética , Medicamentos de Ervas Chinesas/metabolismo , Transcriptoma , Sequência de Aminoácidos , Animais , Peptídeos Catiônicos Antimicrobianos/síntese química , Peptídeos Catiônicos Antimicrobianos/genética , Proteínas de Artrópodes/biossíntese , Proteínas de Artrópodes/genética , Artrópodes/imunologia , Artrópodes/microbiologia , Candida albicans/efeitos dos fármacos , Candida albicans/crescimento & desenvolvimento , Mapeamento de Sequências Contíguas , Alcaloides Diterpenos , Eritrócitos/citologia , Eritrócitos/efeitos dos fármacos , Escherichia coli/química , Escherichia coli/efeitos dos fármacos , Escherichia coli/crescimento & desenvolvimento , Hemólise/efeitos dos fármacos , Sequenciamento de Nucleotídeos em Larga Escala , Humanos , Imunização , Camundongos , Testes de Sensibilidade Microbiana , Dados de Sequência Molecular , Pseudomonas aeruginosa/efeitos dos fármacos , Pseudomonas aeruginosa/crescimento & desenvolvimento , Alinhamento de Sequência , Técnicas de Síntese em Fase Sólida , Staphylococcus aureus/efeitos dos fármacos , Staphylococcus aureus/crescimento & desenvolvimentoRESUMO
[This corrects the article DOI: 10.5851/kosfa.2024.e15.].
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Oxya chinensis sinuosa (OC) is a well-known edible insect. Several researches on the health benefits of OC consumption have been performed to date; however, their effect on eye health remains largely unknown. This study aimed to assess the protective effects of OC extracts on the oxidative stress on the retinal pigment epithelium (RPE) cells. Oxidative damage has been identified as one of the key regulatory factors in age-related macular degeneration. H2O2-induced reactive oxygen species (ROS) production, a well-known oxidative stress factor, can cause cell death in retinal pigment epithelia cells. In this study, we found that three OC extracts effectively prevented H2O2-induced ROS production and subsequent death of ARPE-19 cells in a dose-dependent manner. In addition, the OC extracts inhibited the phosphorylation of mitogen-activated protein kinases including p38, JNK, and ERK. The OC extracts restored IκBα degradation induced by H2O2, indicating that OC extracts suppressed the activation of nuclear factor-κB. Furthermore, the three OC extracts were shown to have antioxidant effects by up-regulating the intracellular expression of key antioxidant proteins such as SOD, NQO, and HO-1. Here we demonstrated the antioxidant and anti-apoptotic effects of the OC extracts on ARPE-19, indicating their potential role in improving eye health. These results suggest that three OC extracts plays a critical role in oxidative stress-induced cell death protects in ARPE-19 cells.
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The rapid development of messenger RNA (mRNA) vaccines formulated with lipid nanoparticles (LNPs) has contributed to control of the COVID-19 pandemic. However, mRNA vaccines have raised concerns about their potential toxicity and clinical safety, including side effects, such as myocarditis, anaphylaxis, and pericarditis. In this study, we investigated the potential of trehalose glycolipids-containing LNP (LNP S050L) to reduce the risks associated with ionizable lipids. Trehalose glycolipids can form hydrogen bonds with polar biomolecules, allowing the formation of a stable LNP structure by replacing half of the ionizable lipids. The efficacy and safety of LNP S050L were evaluated by encapsulating the mRNA encoding the luciferase reporter gene and measuring gene expression and organ toxicity, respectively. Furthermore, mice immunized with an LNP S050L-formulated mRNA vaccine expressing influenza hemagglutinin exhibited a significant reduction in organ toxicity, including in the heart, spleen, and liver, while sustaining gene expression and immune efficiency, compared to conventional LNPs (Con-LNPs). Our findings suggest that LNP S050L, a trehalose glycolipid-based LNP, could facilitate the development of safe mRNA vaccines with improved clinical safety.
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Modulation of osteoclast formation could be a therapeutic target for inhibiting pathological bone destruction. The receptor activator of nuclear factor (NF)-κB ligand (RANKL) is known to be an essential factor in osteoclast differentiation and activation inducers. However, whether Protaetia brevitarsis seulensis (P. brevitarsis) larvae-a traditional animal-derived medicine used in many Asian countries-can inhibit RANKL-induced osteoclast formation and prevent ovariectomy (OVX)-induced bone loss has not been evaluated. Here, we aimed to investigate the anti-osteoporotic effects of P. brevitarsis larvae ethanol extract (PBE) in RANKL-stimulated RAW264.7 cells and OVX mice. In vitro, PBE (0.1, 0.5, 1, and 2 mg/mL) decreased RANKLinduced tartrate-resistant acid phosphatase (TRAP) activity and expression of osteoclastogenesis-associated genes and proteins. Furthermore, PBE (0.1, 0.5, 1, and 2 mg/mL) significantly inhibited the phosphorylation of p38 and NF-κB. Female C3H/HeN mice were divided into five groups (n = 5 per group), namely, sham-operated, OVX, OVX+PBEL (100 mg/kg, oral gavage), OVX+PBEH (200 mg/kg, oral gavage), and OVX+estradiol (0.03 µg/day, subcutaneous injection). High doses of PBE significantly increased femoral bone mineral density (BMD) and bone volume/tissue volume (BV/TV), whereas femoral bone surface/bone volume (BS/BV) and osteoclastogenesis-associated protein expression decreased compared to those in the OVX group. Moreover, PBE (200 mg/kg) significantly increased estradiol and procollagen type I N-terminal propeptide and decreased N-terminal telopeptide of type I collagen and C-terminal telopeptide of type I collagen compared to those in the OVX group. Our results suggest that PBE can be an effective therapeutic candidate for preventing or treating postmenopausal osteoporosis.
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Doenças Ósseas Metabólicas , Osteoporose , Humanos , Camundongos , Animais , Feminino , Osteogênese , Osteoporose/tratamento farmacológico , Larva/metabolismo , Camundongos Endogâmicos C3H , Osteoclastos , Doenças Ósseas Metabólicas/metabolismo , NF-kappa B/metabolismo , Estradiol/farmacologia , Ovariectomia , Ligante RANK/metabolismoRESUMO
Self-rated health (SRH), affected by sociodemographic and health-related behavioral factors, is related to metabolic syndrome (MetS) and high-sensitivity C-reactive protein (hs-CRP) levels. We hypothesized that SRH would have an independent effect on MetS and high hs-CRP incidence in healthy adults after adjusting for sociodemographic and health-related behavioral confounding factors. Data of 1545 healthy participants (aged 19-65 years; 654 men), selected from the 2015 Korean National Health and Nutritional Examination Survey, were cross-sectionally analyzed. The SRH levels significantly increased with higher income (P < .05) and educational levels (P < .01) and were associated with sex, job, marital status, smoking, physical activity, perceived body image, and weight change (all P < .05). The percentage of participants with "very high or high" perceived stress were significantly lower in the "very healthy" group (18.5%) than in the "unhealthy" group (49.7%). Dietary protein, calcium, and phosphorus intakes were significantly higher with better SRH levels. MetS prevalence was related to sex, educational level, job, marital status, smoking, high-intensity exercise, and aerobic exercise (all P < .05), with a 2.776 times higher risk in the "unhealthy" than in the "very healthy" group after adjusting for these confounders. High hs-CRP levels were related to marital status, smoking, weekly walking, and aerobic exercise (all P < .05), with its risk being 2.093 times higher in the "unhealthy" than in the "very healthy" group after adjusting for these confounders. Thus, SRH may be an independent predictor of MetS and high hs-CRP levels and could be used for the development of a health promotion program in healthy adults.
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Proteína C-Reativa , Síndrome Metabólica , Adulto , Idoso , Proteína C-Reativa/metabolismo , Feminino , Humanos , Masculino , Síndrome Metabólica/epidemiologia , Síndrome Metabólica/etiologia , Pessoa de Meia-Idade , Inquéritos Nutricionais , Prevalência , República da Coreia/epidemiologia , Fatores de Risco , Adulto JovemRESUMO
Aging, and other disease-related muscle disorders are serious health problems. Dexamethasone (DEX), a synthetic glucocorticoid, can trigger skeletal muscle atrophy. This study examined the effects of mealworm (Tenebrio molitor larva) ethanol extract (TME) on C2C12 myoblast differentiation and DEX-induced myotube atrophy. TME induced myotube formation compared to the differentiation medium (DM) group. TME also significantly increased the mRNA expression of muscle creatine kinase (CKm) and myogenic regulatory factors (MRFs), such as myogenin (MyoG), myogenic factor (Myf)5, and MRF4 (Myf6). TME dramatically increased the muscle-specific protein, MyoG, compared to the control, whereas the expression of myogenic differentiation 1 (MyoD) remained unchanged. It also activated the mammalian target of rapamycin (mTOR) signaling pathway. In the DEX-induced muscle atrophy C2C12 model, TME reduced the gene expression of atrogin-1, muscle RING finger protein-1 (MuRF-1), and myostatin, which are involved in protein degradation in skeletal muscles. Furthermore, TME elevated the phosphorylation of forkhead box O3 (FoxO3α) and protein kinase B (Akt). These findings suggest that TME can enhance myotube hypertrophy by regulating the mTOR signaling pathway, and can rescue DEX-induced muscle atrophy by alleviating atrophic muscle markers mediated by Akt activation. Thus, TME can be a potential therapeutic agent for treating muscle weakness and atrophy.
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Repair of a retracted anterior L-shaped rotator cuff tear is challenging because of the lack of mobilization of the torn tendon and destruction of the rotator cable. Restoring the anterior rotator cable decreases gap formation of the repaired tendon, leading to maintenance of the repair integrity of the anterior supraspinatus tendon. This Technical Note introduces the surgical technique of anterior cable reconstruction using the proximal biceps tendon for a retracted anterior L-shaped rotator cuff tear.
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Given the challenges posed by antibiotic resistant microbes and the high mortality rate associated with sepsis, there is an urgent need to develop novel peptide antibiotics that exhibit both antimicrobial and anti-inflammatory activities. Herein, we evaluated antimicrobial activity and anti-inflammatory activity of psacotheasin 2, one of the antimicrobial peptide candidates identified previously using an in silico analysis on the transcriptome of Psacothea hilaris. In addition to exhibiting antimicrobial activities against microorganisms without inducing hemolysis, psacotheasin 2 also decreased the nitric oxide production in lipopolysaccharide (LPS)-induced Raw264.7 cells. Moreover, ELISA and western blot analysis revealed that psacotheasin 2 reduced the expression levels of pro-inflammatory enzymes such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). Further, we found that psacotheasin 2 markedly reduced the expression levels of pro-inflammatory cytokines (IL-6 and IL-1ß) by regulating mitogen-activated protein kinases (MAPKs) and nuclear factor-kB (NF-kB) signaling in LPS-induced Raw264.7 cells. We also confirmed that the binding of psacotheasin 2 to bacterial cell membranes occurs via a specific interaction with LPS. In mouse models of LPS-induced shock, psacotheasin 2 significantly enhanced the survival rate and recovered weight by attenuating pro-inflammatory cytokines. Thus, psacotheasin 2 could be a promising candidate as a peptide antiseptic agent.
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Peptídeos Antimicrobianos/genética , Sepse/metabolismo , Animais , Peptídeos Antimicrobianos/metabolismo , Peptídeos Antimicrobianos/farmacologia , Bactérias/efeitos dos fármacos , Besouros/química , Besouros/genética , Besouros/imunologia , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Imunidade Inata , Mediadores da Inflamação , Camundongos , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Peptídeos/genética , Peptídeos/metabolismo , Células RAW 264.7 , Sepse/imunologia , Transdução de SinaisRESUMO
Cockroaches live in places where various pathogens exist and thus are more likely to use antimicrobial compounds to defend against pathogen intrusions. We previously performed an in silico analysis of the Periplaneta americana transcriptome and detected periplanetasin-5 using an in silico antimicrobial peptide prediction method. In this study, we investigated whether periplanetasin-5 has anticancer activity against the human leukemia cell line K562. Cell growth and survival of K562 cells treated with periplanetasin-5 were decreased in a dose-dependent manner. By using flow cytometric analysis, acridine orange/ethidium bromide (AO/EB) staining and DNA fragmentation, we found that periplanetasin-5 induced apoptotic and necrotic cell death in leukemia cells. In addition, these events were associated with increased levels of the pro-apoptotic proteins Fas and cytochrome c and reduced levels of the anti-apoptotic protein Bcl-2. Periplanetasin-5 induces the cleavage of pro-caspase-9, pro-caspase-8, pro-caspase-3, and poly (ADP-ribose) polymerase (PARP). The above data suggest that periplanetasin-5 induces apoptosis via both the intrinsic and extrinsic pathways. Moreover, caspase-related apoptosis was further confirmed by using the caspase inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]- fluoromethylketone (Z-VAD-FMK), which reversed the periplanetasin-5-induced reduction in cell viability. In conclusion, periplanetasin-5 caused apoptosis in leukemia cells, suggesting its potential utility as an anticancer therapeutic agent.
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Peptídeos Antimicrobianos/farmacologia , Antineoplásicos/farmacologia , Apoptose , Proteínas de Insetos/farmacologia , Periplaneta/química , Animais , Produtos Biológicos/farmacologia , Humanos , Células K562RESUMO
Insect antimicrobial peptides (AMPs) have a wide range of functions and potential applications, and have recently attracted attention as alternative foods and medicines for humans. Our study performed transcriptome analysis to explore the potential of the red-striped golden stink bug (Poecilocoris lewisi), and as a result, we have discovered new features of P. lewisi that have not been identified. Specifically, defensin found in P. lewisi is a well-known AMP and is expressed by various plants, animals and fungi for host defense. Moreover, the discovery of defensin in P. lewisi provides new research and important information. In this study, we identified AMP and related DEG in P. lewisi that are closely related to human disease and immune response. These findings will provide the basis and important information for future research on P. lewisi that has not yet been studied.
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Heterópteros/genética , Animais , Perfilação da Expressão Gênica/métodos , Hemípteros/genética , Sequenciamento de Nucleotídeos em Larga Escala/métodos , Transcriptoma/genéticaRESUMO
Calcific tendinitis is the leading cause of shoulder pain. Among patients with calcific tendinitis, 2.7%-20% are asymptomatic, and 35%-45% of patients whose calcific deposits are inadvertently discovered develop shoulder pain. If symptoms are present, complications such as decreased range of motion of the shoulder joint should be minimized while managing pain. Patients with acute calcific tendinitis respond well to conservative treatment and rarely require surgery. In contrast, patients with chronic calcific tendinitis often do not respond to conservative treatment and do require surgery. Clinical improvement takes time, even after surgical treatment. This review article summarizes the processes related to the diagnosis and treatment of calcific tendinitis with the aim of helping clinicians choose appropriate treatment options for their patients.
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Previously, we performed an in silico analysis of the Periplaneta americana transcriptome. Antimicrobial peptide candidates were selected using an in silico antimicrobial peptide prediction method. It was found that periplanetasin-5 had antimicrobial activity against yeast and grampositive and gram-negative bacteria. In the present study, we demonstrated the anti-inflammatory activities of periplanetasin-5 in mouse macrophage Raw264.7 cells. No cytotoxicity was observed at 60 µg/ml periplanetasin-5, and treatment decreased nitric oxide production in Raw264.7 cells exposed to lipopolysaccharide (LPS). In addition, quantitative RT-PCR and enzyme-linked immunosorbent assay revealed that periplanetasin-5 reduced cytokine (tumor necrosis factor-α, interleukin-6) expression levels in the Raw264.7 cells. Periplanetasin-5 controlled inflammation by inhibiting phosphorylation of MAPKs, an inflammatory signaling element, and reducing the degradation of IκB. Through LAL assay, LPS toxicity was found to decrease in a periplanetasin-5 dose-dependent manner. Collectively, these data showed that periplanetasin-5 had antiinflammatory activities, exemplified in LPS-exposed Raw264.7 cells. Thus, we have provided a potentially useful antibacterial peptide candidate with anti-inflammatory activities.
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Anti-Inflamatórios/farmacologia , Periplaneta/metabolismo , Proteínas Citotóxicas Formadoras de Poros/farmacologia , Animais , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/genética , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Regulação da Expressão Gênica , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Negativas/metabolismo , Inflamação/tratamento farmacológico , Proteínas de Insetos/farmacologia , Lipopolissacarídeos/toxicidade , Camundongos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/antagonistas & inibidores , Óxido Nítrico Sintase Tipo II/genética , Óxido Nítrico Sintase Tipo II/metabolismo , Células RAW 264.7 , Espécies Reativas de Oxigênio/metabolismo , Transdução de SinaisRESUMO
Antimicrobial peptides (AMPs) are the frontline innate defense system evolutionarily preserved in insects to combat invading pathogens. These AMPs could serve as an alternative to classical antibiotics to overcome the burden of treating multidrug resistant bacteria. Psacotheasin, a knottin type AMP was isolated from Psacothea hilaris and shown to exhibit antimicrobial activity, especially against fungi through apoptosis mediated cell death. In this study, we aimed to identify novel probable AMPs from Psacothea hilaris, the yellow spotted longicorn beetle. The beetle was immunized with the two bacterial strains (E. coli and S. aureus), and the yeast strain C. albicans. After immunization, total RNA was isolated and sequenced in Illumina platform. Then, beetle transcriptome was de novo assembled and searched for putative AMPs with the known physiochemical features of the AMPs. A selection of AMP candidates were synthesized and tested for antimicrobial activity. Four peptides showed stronger activity against E. coli than the control AMP, melittin while one peptide showed similar activity against S. aureus. Moreover, four peptides and two peptides showed antifungal activity stronger than and similar to melittin, respectively. Collectively one peptide showed both antibacterial and antifungal activity superior to melittin; thus, it provides a potent antimicrobial peptide. All the peptides showed no hemolysis in all the tested concentrations. These results suggest that in silico mining of insects' transcriptome could be a promising tool to obtain and optimize novel AMPs for human needs.
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In a previous work, we performed de novo RNA sequencing of Allomyrina dichotoma using next generation sequencing and identified several antimicrobial peptide candidates based on transcriptome analysis. Among them, a cationic antimicrobial peptide, allomyrinasin, was selected bioinformatically based on its physicochemical properties. Here, we assessed the antimicrobial and anti-inflammatory activities of allomyrinasin against microorganisms and mouse macrophage Raw264.7 cells. Allomyrinasin showed antimicrobial activities against various microbes and decreased the nitric oxide production of the lipopolysaccharide-induced Raw264.7 cells. Furthermore, quantitative RT-PCR and ELISA revealed that allomyrinasin reduced cytokine expression levels in the Raw264.7 cells. We also identified inducible nitric oxide synthase, cyclooxygenase-2 expression, and PGE2 production through western blot analysis and ELISA. We confirmed that allomyrinasin bound to bacterial cell membranes via a specific interaction with lipopolysaccharides. Taken together, these data indicate that allomyrinasin has antimicrobial and anti-inflammatory activities as exemplified in lipopolysaccharide-induced Raw264.7 cells. We have provided a potentially useful antimicrobial peptide candidate that has both antimicrobial and anti-inflammatory activities.
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Anti-Infecciosos/farmacologia , Anti-Inflamatórios/farmacologia , Peptídeos Catiônicos Antimicrobianos/farmacologia , Besouros/química , Animais , Anti-Infecciosos/química , Anti-Inflamatórios/química , Sobrevivência Celular/efeitos dos fármacos , Ciclo-Oxigenase 2/metabolismo , Citocinas/metabolismo , Lipopolissacarídeos/efeitos adversos , Camundongos , Óxido Nítrico/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Células RAW 264.7/efeitos dos fármacosRESUMO
In a previous study, we analyzed the transcriptome of Scolopendra subspinipes mutilans using next-generation sequencing technology and identified several antimicrobial peptide candidates. One of the peptides, scolopendrasin V, was selected based on the physicochemical properties of antimicrobial peptides using a bioinformatics strategy. In this study, we assessed the antimicrobial activities of scolopendrasin V using the radial diffusion assay and colony count assay. We also investigated the mode of action of scolopendrasin V using flow cytometry. We found that scolopendrasin V's mechanism of action involved binding to the surface of microorganisms via a specific interaction with lipopolysaccharides, lipoteichoic acid, and peptidoglycans, which are components of the bacterial membrane. These results provide a basis for developing peptide antibiotics.