RESUMO
Diabetic peripheral neuropathy (DPN) is a common diabetes complication (DM). Aldose reductase -2 (ALR-2) is an oxidoreductase enzyme that is most extensively studied therapeutic target for diabetes-related complications that can be inhibited by epalrestat, which has severe adverse effects; hence the discovery of potent natural inhibitors is desired. In response, a pharmacophore model based on the properties of eplarestat was generated. The specified pharmacophore model searched the NuBBEDB database of natural compounds for prospective lead candidates. To assess the drug-likeness and ADMET profile of the compounds, a series of in silico filtering procedures were applied. The compounds were then put through molecular docking and interaction analysis. In comparison to the reference drug, four compounds showed increased binding affinity and demonstrated critical residue interactions with greater stability and specificity. As a result, we have identified four potent inhibitors: ZINC000002895847, ZINC000002566593, ZINC000012447255, and ZINC000065074786, that could be used as pharmacological niches to develop novel ALR-2 inhibitors.
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Spices and herbs are recognized as sources of natural antioxidants and thus play an important role in the chemoprevention of diseases and aging. Piper cubeba is one among them and known for its medicinal properties for decades. Various biological activities are associated with its extract and phytocompounds. However, the anti-mutagenic activity of antioxidant rich extract is less explored. In this study, we performed the fraction-based antioxidant activity of P. cubeba using four different assays and evaluated the anti-mutagenic activity of most potent antioxidant fraction using Salmonella typhimurium tester strains against four mutagens (methyl methanesulfonate [MMS], sodium azide [SA], benzo(a)pyrene, and 2-aminoflourene) respectively. Among all tested fractions at 25-200 µg/ml, ethanolic extract revealed highest antioxidant activity and significant anti-mutagenicity against both direct and indirect acting mutagens at least one tester strain. Phytochemical analysis by gas chromatography-mass spectrometry (GC/MS) revealed the presence of various phytocompounds including copaene, isocaryophyllene, α-cubebene, etc. Molecular docking studies on DNA binding interactions of GC/MS detected phytocompounds highlight the possible mode of binding. In summary, these in vitro studies have provided the scientific basis for validation of using this plant in the traditional system of medicine and highlighted the need for exploring the role of various compounds for therapeutic efficacy. On the other hand, synergistic interaction among phytocompounds is to be explored to optimize or standardize the extracts for the exploitation in modern phytomedicine.
Assuntos
Antimutagênicos/farmacologia , Antioxidantes/farmacologia , Piper , Extratos Vegetais/farmacologia , Frutas , Cromatografia Gasosa-Espectrometria de Massas , Simulação de Acoplamento Molecular , Piper/químicaRESUMO
Previous studies of naturally occurring mouse papillomavirus (PV) MmuPV1-induced tumors in B6.Cg-Foxn1nu/nu mice suggest that T cell deficiency is necessary and sufficient for the development of such tumors. To confirm this, MmuPV1-induced tumors were transplanted from T cell-deficient mice into immunocompetent congenic mice. Consequently, the tumors regressed and eventually disappeared. The elimination of MmuPV1-infected skin/tumors in immunocompetent mice was consistent with the induction of antitumor T cell immunity. This was confirmed by adoptive cell experiments using hyperimmune splenocytes collected from graft-recipient mice. In the present study, such splenocytes were injected into T cell-deficient mice infected with MmuPV1, and they eliminated both early-stage and fully formed tumors. We clearly show that anti-tumor T cell immunity activated during tumor regression in immunocompetent mice effectively eliminates tumors developing in T cell-deficient congenic mice. The results corroborate the notion that PV-induced tumors are strongly linked to the immune status of the host, and that PV antigens are major anti-tumor antigens. Successful anti-PV T cell responses should, therefore, lead to effective anti-tumor immune therapy in human PV-infected patients.
Assuntos
Modelos Animais de Doenças , Imunidade Celular/imunologia , Papillomaviridae/imunologia , Infecções por Papillomavirus/complicações , Neoplasias Cutâneas/prevenção & controle , Linfócitos T/imunologia , Animais , Feminino , Camundongos , Camundongos Congênicos , Camundongos Endogâmicos C57BL , Camundongos Nus , Infecções por Papillomavirus/virologia , Neoplasias Cutâneas/etiologia , Neoplasias Cutâneas/patologiaRESUMO
Fruits, vegetables and medicinal herbs rich in phenolics antioxidants contribute toward reduced risk of age-related diseases and cancer. In this study, Psidium guajava leaf extract was fractionated in various organic solvents viz. petroleum ether, benzene, ethyl acetate, ethanl and methanol and tested for their antioxidant and antimutagenic properties. Methanolic fraction showed maximum antioxidant activity comparable to ascorbic acid and butylated hydroxyl toluene (BHT) as tested by DPPH free radical scavenging, phosphomolybdenum, FRAP (Fe3 + reducing power) and CUPRAC (cupric ions (Cu2+) reducing ability) assays. The fraction was analyzed for antimutagenic activities against sodium azide (NaN3), methylmethane sulfonate (MMS), 2-aminofluorene (2AF) and benzo(a)pyrene (BP) in Ames Salmonella tester strains. The methanol extracted fraction at 80 µg/ml concentration inhibited above 70% mutagenicity. Further, phytochemical analysis of methanol fraction that was found to be most active revealed the presence of nine major compounds by gas chromatography-mass spectrometry (GC-MS). This data suggests that guava contains high amount of phenolics responsible for broad-spectrum antimutagenic and antioxidant properties in vitro and could be potential candidates to be explored as modern phytomedicine.
Assuntos
Antimutagênicos/farmacologia , Antioxidantes/farmacologia , Mutação/efeitos dos fármacos , Extratos Vegetais/farmacologia , Folhas de Planta/química , Psidium/química , Antimutagênicos/química , Antimutagênicos/isolamento & purificação , Antioxidantes/química , Antioxidantes/isolamento & purificação , Compostos de Bifenilo/química , Cobre/química , DNA Bacteriano/efeitos dos fármacos , DNA Bacteriano/genética , Relação Dose-Resposta a Droga , Ferricianetos/química , Cromatografia Gasosa-Espectrometria de Massas , Molibdênio/química , Testes de Mutagenicidade , Mutagênicos/toxicidade , Oxirredução , Fenóis/isolamento & purificação , Fenóis/farmacologia , Fitoterapia , Picratos/química , Extratos Vegetais/química , Extratos Vegetais/isolamento & purificação , Plantas Medicinais , Salmonella/efeitos dos fármacos , Salmonella/genética , Solventes/químicaRESUMO
We isolated two new manatee papillomavirus (PV) types, TmPV3 and TmPV4, from a Florida manatee (Trichechus manatus latirostris). Two PV types were previously isolated from this species. TmPV1 is widely dispersed amongst manatees and a close-to-root PV; not much is known about TmPV2. The genomes of TmPV3 and TmPV4 were 7622 and 7771 bp in size, respectively. Both PVs had a genomic organization characteristic of all PVs, with one non-coding region and seven ORFs, including the E7 ORF that is absent in other cetacean PVs. Although these PVs were isolated from separate genital lesions of the same manatee, an enlarged E2/E4 ORF was found only in the TmPV4 genome. The full genome and L1 sequence similarities between TmPV3 and TmPV4 were 63.2 and 70.3 %, respectively. These genomes shared only 49.1 and 50.2 % similarity with TmPV1. The pairwise alignment of L1 nucleotide sequences indicated that the two new PVs nested in a monophyletic group of the genus Rhopapillomavirus, together with the cutaneotropic TmPV1 and TmPV2.
Assuntos
Clonagem Molecular , Genoma Viral , Papillomaviridae/genética , Infecções por Papillomavirus/veterinária , Trichechus manatus/virologia , Animais , DNA Viral/genética , Papillomaviridae/classificação , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/virologia , FilogeniaRESUMO
BACKGROUND: Current vaccines against Human Papillomavirus (HPV) are highly effective and based on recombinant virus-like particles (VLPs) of the major capsid protein L1. Since these vaccines are HPV type-specific and expensive for global implementation, an alternative, broader-spectrum immunogen would be the N-terminus of the minor capsid protein L2 that induces low titered broadly cross-neutralizing antibodies. Here we analyzed the reactivity of different synthetic L2 peptides containing N-terminus amino acids 17-36 in order to test their antigenicity. METHODS: Different synthetic peptides were designed to target the 17-36 amino acid sequences, present in highly antigenic amino-terminus of L2 protein. Six different peptides including Cys22-Cys28 disulfide bonded cyclized L2 peptide were examined for their antigenicity against mouse monoclonal antibody RG-1 and rabbit polyclonal antisera to HPV L2 by enzyme-linked immunosorbent assay (ELISA). RESULTS: Here we report that the cyclized form of synthetic L2 peptide, which is formed through Cys22-Cys28 disulfide bridges, has the highest reactivity to antibodies than other synthetic L2 peptides. CONCLUSION: A cyclized L2 peptide has potential to be an excellent candidate to formulate a low-cost, broadly protective pan-oncogenic HPV vaccine.
Assuntos
Proteínas do Capsídeo/imunologia , Dissulfetos/química , Epitopos/química , Epitopos/imunologia , Proteínas Oncogênicas Virais/imunologia , Infecções por Papillomavirus/prevenção & controle , Fragmentos de Peptídeos/farmacologia , Vacinas Sintéticas/imunologia , Sequência de Aminoácidos , Animais , Anticorpos Neutralizantes/imunologia , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Dissulfetos/imunologia , Ensaio de Imunoadsorção Enzimática , Humanos , Camundongos , Dados de Sequência Molecular , Papillomaviridae/imunologia , Infecções por Papillomavirus/imunologia , Infecções por Papillomavirus/virologia , Vacinas contra Papillomavirus/imunologia , Vacinas contra Papillomavirus/uso terapêutico , Fragmentos de Peptídeos/imunologia , Coelhos , Homologia de Sequência de AminoácidosRESUMO
BACKGROUND: Human papillomavirus (HPV)-positive oropharyngeal cancer is associated with improved survival and treatment response as compared to HPV-negative cancers. P16 overexpression is widely accepted as a surrogate marker for HPV positivity. METHODS: A total of 92 serum samples from 75 head and neck squamous cell carcinoma (HNSCC) patients were examined for HPV16 and 18 E7 antibodies by ELISA. Available tissue was tested for HPV-DNA by PCR, and p16 immunohistochemistry was obtained from a deidentified database. RESULTS: Of 75 HNSCC patients, 25 were HPV E7 seropositive. Seropositivity was strongly associated with cancers of the oropharynx, and correlated with positive p16 immunohistochemistry (IHC) and HPV-DNA. Post-treatment serum was available in a limited subset of patients, revealing a decrease in antibody titers following response to treatment. CONCLUSIONS: HPV E7 seropositivity correlated with positive tumor HPV-DNA and p16 expression, and was strongly associated with cancers of the oropharynx. E7 serology warrants further study as a potential biomarker in HPV-positive HNSCC.
Assuntos
Carcinoma de Células Escamosas/metabolismo , Inibidor p16 de Quinase Dependente de Ciclina/metabolismo , Neoplasias de Cabeça e Pescoço/metabolismo , Proteínas E7 de Papillomavirus/sangue , Infecções por Papillomavirus/virologia , Adulto , Idoso , Carcinoma de Células Escamosas/patologia , Carcinoma de Células Escamosas/virologia , DNA Viral/genética , Ensaio de Imunoadsorção Enzimática , Feminino , Neoplasias de Cabeça e Pescoço/patologia , Neoplasias de Cabeça e Pescoço/virologia , Papillomavirus Humano 16/genética , Papillomavirus Humano 16/isolamento & purificação , Humanos , Técnicas Imunoenzimáticas , Masculino , Pessoa de Meia-Idade , Estadiamento de Neoplasias , Infecções por Papillomavirus/metabolismo , Infecções por Papillomavirus/patologia , Reação em Cadeia da Polimerase , Prognóstico , Estudos SoroepidemiológicosRESUMO
OBJECTIVES: Patient experiences are critical when determining the acceptability of novel interventional pharmaceuticals. Here, we report the development and validation of a product acceptability questionnaire (SPRAY PAL) assessing feasibility, acceptability and tolerability of an intranasal Q-Griffithsin (Q-GRFT) drug product designed for COVID-19 prophylaxis. DESIGN: SPRAY PAL validation was undertaken as part of an ongoing phase 1 clinical trial designed to test the safety, pharmacokinetics and tolerability of intranasally administered Q-GRFT for the prevention of SARS-CoV-2 infection. SETTING: The phase 1 clinical trial took place at a University Outpatient Clinical Trials Unit from November 2021 to September 2023. PARTICIPANTS: The initial SPRAY PAL questionnaire was piloted among healthy volunteers ages 25 to 55 in phase 1a of the clinical trial (N=18) and revised for administration in phase 1b for participants ages 24-59 (N=22). RESULTS: Spearman correlations tested convergent and discriminant validity. Internal consistency was assessed using Cronbach's alpha, and test-retest reliability was assessed using intraclass correlation coefficients of responses collected from three repeated questionnaire administrations. The initial version demonstrated excellent internal consistency. The revised version demonstrated very good internal consistency after removal of one item (alpha=0.739). Excellent test-retest reliability (intraclass coefficient=0.927) and adequate convergent (r's=0.208-0.774) and discriminant (r's=0.123-0.392) validity were achieved. Subscales adequately distinguished between the constructs of acceptability, feasibility and tolerability. CONCLUSIONS: The SPRAY PAL product acceptability questionnaire is a valid and reliable patient-reported outcomes measure that can be considered a credible tool for assessing patient-reported information about product acceptability, feasibility of use, tolerability of product and side effects and cost of product for novel intranasal drug formulations. The SPRAY PAL is generalisable, and items may be readily adapted to assess other intranasal formulations. TRIAL REGISTRATION NUMBERS: NCT05122260 and NCT05437029.
Assuntos
COVID-19 , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos , Humanos , COVID-19/prevenção & controle , Reprodutibilidade dos Testes , SARS-CoV-2RESUMO
Multidrug resistance (MDR) in microorganisms is a cause of major concern for clinicians and pharmaceutical industries. Continuous development of new antimicrobial drugs with multiple targets and potentials is expected to efficiently combat MDR in these microorganisms. In a continued exploration of new antimicrobial drug leads, 11 marine natural products, semisynthetic, or related synthetic analogs (1-11) and two tobacco cembranoids (12 and 13) were screened for their antimicrobial, antioxidant, and antimutagenic activities. Eight compounds showed varying levels of both antibacterial and antifungal activities. Compounds such as 17-O-methyllatrunculin-A, verongiaquinol, (1S,2E,4R,6R,7E,11E)-2,7,11-cembratriene-4,6-diol), and manzamine-A showed a broad spectrum of activity, inhibiting six of seven tested bacteria with zone of inhibition diameter from 9 to 30 mm. Four of these active compounds also showed antifungal activity. The findings of the in vitro time-kill assay of the most active compound, verongiaquinol, against Staphylococcus aureus indicated its subinhibitory effect at the level lower than the minimal inhibitory concentration (MIC) values (i.e., 2 and 4 µg/mL). At the MIC (8 µg/mL), bacterial cells were completely killed within 18 hours of incubation. DPPH free radical scavenging activity was demonstrated by five compounds in the range of 89.65-36.19% decolorization. Further, four compounds evaluated for their antimutagenic activity against the directly acting mutagens, methyl methanesulfonate and sodium azide, in Salmonella typhimurium strains, interestingly, showed no sign of mutagenicity. Verongiaquinol and manzamine A, in fact, reduced the mutagenicity by 50-75% at a dose of 5 µg/plate in different test strains. Our study seems to provide some novel antimicrobial leads with strong antioxidant potential and the associated ability of antimutagenicity.
Assuntos
Anti-Infecciosos/farmacologia , Antimutagênicos/farmacologia , Antioxidantes/farmacologia , Diterpenos/farmacologia , Biologia Marinha , Nicotiana/química , Anti-Infecciosos/química , Anti-Inflamatórios , Antimutagênicos/química , Antioxidantes/química , Compostos de Bifenilo/química , Radicais Livres/química , Fungos/efeitos dos fármacos , Testes de Sensibilidade Microbiana , Picratos/química , Extratos Vegetais/farmacologia , Salmonella typhimurium/efeitos dos fármacos , Salmonella typhimurium/genética , Extratos de Tecidos/farmacologiaRESUMO
Piper nigrum is a widely used plant in traditional remedies and known for its numerous biological properties. However, fraction-based antioxidant activity and their antimutagenic potential are not yet fully investigated. Different extracts of the seeds P. nigrum were obtained by sequential extraction in different solvents. All extracts were evaluated for antibacterial and antioxidant activities using different methods. The most active fraction was analyzed for antimutagenic activity using the Ames Salmonella test. The antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) was found to be more prominent compared to ESßL producing Klebsiella pneumoniae isolates. The MIC values were found to be lower against MRSA than K. pneumoniae. The extract showing highest antioxidant activity (methanol extract) was further tested for antimutagenic activity both against direct and indirect-acting mutagens. A varying level of antimutagenic activity was shown by methanol extract at highest tested concentration (200⯵g/plate). Alkaloids, phenols, and flavonoids were detected as major class of compounds in methanol extract. Gas chromatography-mass spectrometry (GC-MS) analysis showed the presence of various phytocompounds. Based on molecular docking of two major active phytocompounds (piperine and copaene), they were found to interact at the minor groove of DNA. Molecular dynamics (MD) simulation revealed that both the ligands were quite stable with DNA under physiological conditions. The ability of phytocompounds to interact with DNA might be reducing the interaction of mutagens and could be one of the possible mechanism of anti-mutagenic activity of P. nigrum extract. This study highlights the antioxidant and antimutagenic potential of Piper nigrum. The role of phytocompounds present in the bioactive extract is needed to be explored further for herbal drug research.
RESUMO
Over the past few decades, scientific research has indicated a credible basis for some of the traditional ethnomedicinal uses of pomegranate. This study aims to evaluate the broad spectrum antioxidant and antimutagenic activities of peel extracts of pomegranate. The sequentially extracted Punica granatum peel fractions were tested for their antioxidant activity by DPPH free radical scavenging, phosphomolybdenum, FRAP (Fe(3+) reducing power) and CUPRAC (cupric ions (Cu(2+)) reducing ability) assays. The methanol fraction showed highest antioxidant activity by all the four in vitro assays comparable to ascorbic acid and butylated hydroxy toluene (BHT) followed by activity in ethanol, acetone, and ethyl acetate fractions. Based on the promising antioxidant activities, the methanol fraction was evaluated for antimutagenic activity by Ames Salmonella/microsome assay against sodium azide (NaN(3)), methyl methane sulphonate (MMS), 2-aminofluorene (2-AF) and benzo(a)pyrene (B(a)P) induced mutagenicity in Salmonella typhimurium (TA97a, TA98, TA100 and TA102) tester strains. The methanol fraction showed no sign of mutagenicity at tested concentration of 10-80µg/mL. This fraction showed antimutagenic activity against NaN(3) and MMS with percent inhibition of mutagenicity ranging from 66.76% to 91.86% in a concentration-dependent manner. Similar trend of inhibition of mutagenicity (81.2-88.58%) against indirect mutagens (2-AF and B(a)P) was also recorded. Phytochemical analysis by HPLC, LC-MS and total phenolic content revealed high content of ellagitannins which might be responsible for promising antioxidant and antimutagenic activities of P. granatum peel extract. Further, contribution of bioactive compounds detected in this study is to be explored to understand the exact mechanism of action as well as their therapeutic efficacy.
Assuntos
Antimutagênicos/farmacologia , Antioxidantes/farmacologia , Lythraceae/química , Extratos Vegetais/farmacologia , Testes de Mutagenicidade , Extratos Vegetais/química , Salmonella typhimurium/genéticaRESUMO
Discovery of quorum sensing (QS) system to coordinate virulence and biofilm formation in bacterial pathogens has triggered search for safe, stable and non-toxic anti-QS compounds from natural products. Ethanolic extracts of 24 Indian medicinal plants were tested by agar well and disc diffusion assay for anti-QS activity using Chromobacterium violaceum (CV12472 and CVO26) reporter strains. AHL from C. violaceum CV31532 was isolated and partially purified for its use in CVO26 based bioassay. Effect on swarming-motility of Pseudomonas aeruginosa (PAO1) was also recorded at sub-MIC concentrations of extracts. Of the 24 medicinal plants screened Hemidesmus indicus (L.) Schult (root), Holarrhena antidysenterica (Roth) A.DC. (bark), Mangifera indica L. (seed) Punica granatum L. (pericarp) and Psoralea corylifolia L. (seed) demonstrated varying level of inhibition of violacein production in the reporter strains. Moreover, a significant reduction in swarms was recorded over control. The inhibition of violacein production and swarming motility may be due to direct or indirect interference on QS by active constituents or the interactive effect of different phytocompounds present in the extracts. These plant extracts may be selected for activity guided fractionation to identify and characterize the active principle.
Assuntos
Chromobacterium/efeitos dos fármacos , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Pseudomonas aeruginosa/efeitos dos fármacos , Percepção de Quorum/efeitos dos fármacos , Animais , Índia , Indóis/metabolismo , Testes de Sensibilidade MicrobianaRESUMO
Effect of growing seedling, seeds and seedlings extracts from seven leguminous plants (Pisum sativum, Vigna radiata, Vigna mungo, Cajanus cajan, Lentil culinaris, Cicer arietinum and Trigonella foenum graecum) were screened for their ability to influence quorum sensing controlled pigment production in Chromobacterium violaceum indicator strains (CV12472 and CVO26). Germinating seedling and seedling extracts of only P. sativum (pea) showed inhibition of violacein production. Interestingly, the T. foenum graecum (fenugreek) seed extracts enhances the pigment production. Quorum sensing regulated swarming motility in Pseudomonas aerugionsa PAO1 was reduced by pea seedling extract while enhanced by the fenugreek seed extracts. These findings suggest that plant metabolites of some legumes interact actively with bacterial quorum sensing and could modulate its associated functions.
RESUMO
The COVID-19 pandemic is responsible for an unprecedented disruption to the healthcare systems and economies of countries around the world. Developing novel therapeutics and a vaccine against SARS-CoV-2 requires an understanding of the similarities and differences between the various human coronaviruses with regards to their phylogenic relationships, transmission, and management. Phylogenetic analysis indicates that humans were first infected with SARS-CoV-2 in late 2019 and the virus rapidly spread from the outbreak epicenter in Wuhan, China to various parts of the world. Multiple variants of SARS-CoV-2 have now been identified in particular regions. It is apparent that MERS, SARS-CoV, and SARS-CoV-2 present with several common symptoms including fever, cough, and dyspnea in mild cases, but can also progress to pneumonia and acute respiratory distress syndrome. Understanding the molecular steps leading to SARS-CoV-2 entry into cells and the viral replication cycle can illuminate crucial targets for testing several potential therapeutics. Genomic and structural details of SARS-CoV-2 and previous attempts to generate vaccines against SARS-CoV and MERS have provided vaccine targets to manage future outbreaks more effectively. The coordinated global response against this emerging infectious disease is unique and has helped address the need for urgent therapeutics and vaccines in a remarkably short time.
RESUMO
A rapid and efficient one-pot condensation reaction of long-chain alkyl and alkenyl acid hydrazides and nitriles was carried out to afford 3,5-disubstituted-1H-1,2,4-triazoles. The compounds 5a-o were screened for in-vitro antibacterial activity against the representative panel of two Gram-positive and two Gram-negative bacteria. All the synthesized compounds were also tested for their inhibitory action against five strains of fungi. The various compounds show potent inhibitory action against test organisms. The compounds 5a-o were characterized on the basis of elemental analysis and spectral data.
Assuntos
Antibacterianos/síntese química , Antifúngicos/síntese química , Triazóis/síntese química , Antibacterianos/farmacologia , Antifúngicos/farmacologia , Fungos/efeitos dos fármacos , Bactérias Gram-Negativas/efeitos dos fármacos , Bactérias Gram-Positivas/efeitos dos fármacos , Hidrazinas/química , Testes de Sensibilidade Microbiana , Nitrilas/química , Relação Estrutura-Atividade , Triazóis/farmacologiaRESUMO
Methanolic extracts of Acorus calamus (Rhizome), Hemidesmus indicus (Stem), Holarrhena antidysenterica (Bark) and Plumbago zeylanica (Root), were tested for their antimutagenic potential. These extracts, at tested concentrations, showed no sign of mutagenicity to Salmonella typhimurium tester strains. The extracts of the plants exhibited varying level of antimutagenicity. At a dose of 100 microg/plate, the extracts exhibited the inhibition of His+ revertants from 18.51% to 82.66% against direct acting mutagens, methyl methanesulphonate (MMS) and sodium azide (NaN3) induced mutagenicity in Salmonella tester strains TA 97a, TA 100, TA 102 and TA 104. However, at lower concentrations (25 and 50 mcirog/plate) of the plant extracts, a decrease in antimutagenic activity was recorded. Dose dependent antimutagenic activity of the extracts is also evident from linear regression analysis of the data. The over all antimutagenic potential of above four extracts was found to be in order of A. calamus > H. indicus > H. antidysenterica > P. zeylanica. Further, total phenolic content of these extracts did not correlate with its antimutagenic activity in A. calamus and P. zeylanica.
Assuntos
Antimutagênicos/farmacologia , Ayurveda , Metanol , Extratos Vegetais/química , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Antimutagênicos/química , Salmonella/efeitos dos fármacos , Salmonella/genéticaRESUMO
This study evaluated the integration and methlyation of human papillomavirus type 16 (HPV16) in head and neck squamous cell carcinoma (HNSCC) and its oral precursor, high-grade oral epithelial dysplasia (hgOED). Archival samples of HPV16-positive hgOED (N = 19) and HNSCC (N = 15) were evaluated, along with three HNSCC (UMSCC-1, -47 and -104) and two cervical cancer (SiHa and CaSki) cell lines. HgOED cases were stratified into three groups with increasing degrees of cytologic changes (mitosis, karyorrhexis and apoptosis). The viral load was higher and the E2/E6 ratio lower (indicating a greater tendency toward viral integration) in group 3 than in groups 1 or 2 (p = 0.002, 0.03). Methylation was not observed in hgOED cases and occurred variably in only three HNSCC cases (26.67%, 60.0% and 93.3%). In HNSCC cell lines, lower E7 expression correlated with higher levels of methylation. HgOED with increased cytologic change, now termed HPV-associated oral epithelial dysplasia (HPV-OED), exhibited an increased viral load and a tendency toward DNA integration, suggesting a potentially increased risk for malignant transformation. More detailed characterization and clinical follow-up of HPV-OED patients is needed to determine whether HPV-OED is a true precursor to HPV-associated HNSCC and to clarify the involvement of HPV in HNSCC carcinogenesis.
RESUMO
The Florida manatee (Trichechus manatus latirotris) is a threatened aquatic mammal in United States coastal waters. Over the past decade, the appearance of papillomavirus-induced lesions and viral papillomatosis in manatees has been a concern for those involved in the management and rehabilitation of this species. To date, three manatee papillomaviruses (TmPVs) have been identified in Florida manatees, one forming cutaneous lesions (TmPV1) and two forming genital lesions (TmPV3 and TmPV4). We identified DNA sequences with the potential to form G-quadruplex structures (G4) across the three genomes. G4 were located on both DNA strands and across coding and non-coding regions on all TmPVs, offering multiple targets for viral control. Although G4 have been identified in several viral genomes, including human PVs, most research has focused on canonical structures comprised of three G-tetrads. In contrast, the vast majority of sequences we identified would allow the formation of non-canonical structures with only two G-tetrads. Our biophysical analysis confirmed the formation of G4 with parallel topology in three such sequences from the E2 region. Two of the structures appear comprised of multiple stacked two G-tetrad structures, perhaps serving to increase structural stability. Computational analysis demonstrated enrichment of G4 sequences on all TmPVs on the reverse strand in the E2/E4 region and on both strands in the L2 region. Several G4 sequences occurred at similar regional locations on all PVs, most notably on the reverse strand in the E2 region. In other cases, G4 were identified at similar regional locations only on PVs forming genital lesions. On all TmPVs, G4 sequences were located in the non-coding region near putative E2 binding sites. Together, these findings suggest that G4 are possible regulatory elements in TmPVs.
Assuntos
DNA Viral/química , DNA Viral/genética , Quadruplex G , Papillomaviridae/genética , Infecções por Papillomavirus/veterinária , Trichechus manatus/virologia , Animais , Sequência de Bases , Fenômenos Biofísicos , Florida , Genoma Viral , Humanos , Simulação de Dinâmica Molecular , Papillomaviridae/química , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/virologiaRESUMO
OBJECTIVES: Reported cytologic alterations associated with high-risk human papillomavirus (HR-HPV) in oral epithelial dysplasia (HPV-OED) need further characterization. STUDY DESIGN: Archival cases of high-grade oral epithelial dysplasia (hgOED) (N = 38) were assigned a cytologic score (CS) based on the average number of mitotic, karyorrhectic, and apoptotic cells per high-power field. Three groups were then generated on the basis of increasing CS: Focal (group 1, N = 14), Intermediate (group 2, N = 12), and Diffuse (group 3, N = 12). Polymerase chain reaction-based HPV genotyping and p16 immunohistochemistry were performed. RESULTS: HR-HPV was found significantly more in group 3 (83.3%) compared with groups 1 and 2 (group 1&2; 42.9% and 41.7%, respectively; P = .047). HPV16 predominated in HR-HPV-positive cases (90.5%). By location, the tongue or the floor of mouth was associated with all groups (P = .04). Increasing CS was associated with a slightly younger age (P = .04) and increased expression of p16 (P = .005). CS and p16 expression were not sensitive but were highly specific predictors for HR-HPV presence. Based on limited follow-up information, HPV-OED does not differ in clinical aggressiveness compared with conventional OED. CONCLUSIONS: Increased CS in hgOED is strongly associated with HR-HPV (mostly HPV16) and p16 expression. CS and p16 expression are specific predictors of HR-HPV presence. Further molecular study and long-term follow-up of HPV-OED are needed.
Assuntos
Doenças da Boca/virologia , Neoplasias Bucais/virologia , Infecções por Papillomavirus/patologia , Lesões Pré-Cancerosas/virologia , Adulto , Apoptose , Carcinoma in Situ/virologia , Diferenciação Celular , Proliferação de Células , Inibidor p16 de Quinase Dependente de Ciclina/análise , Feminino , Humanos , Imuno-Histoquímica , Masculino , Pessoa de Meia-Idade , Reação em Cadeia da Polimerase , Fatores de RiscoRESUMO
Cervical cancer is the most common malignancy among women particularly in developing countries, with human papillomavirus (HPV) 16 causing 50% of invasive cervical cancers. A plant-based HPV vaccine is an alternative to the currently available virus-like particle (VLP) vaccines, and would be much less expensive. We optimized methods to express HPV16 L1 protein and purify VLPs from tobacco (Nicotiana benthamiana) leaves transfected with the magnICON deconstructed viral vector expression system. L1 proteins were extracted from agro-infiltrated leaves using a series of pH and salt mediated buffers. Expression levels of L1 proteins and VLPs were verified by immunoblot and ELISA, which confirmed the presence of sequential and conformational epitopes, respectively. Among three constructs tested (16L1d22, TPL1d22, and TPL1F), TPL1F, containing a full-length L1 and chloroplast transit peptide, was best. Extraction of HPV16 L1 from leaf tissue was most efficient (> 2.5% of total soluble protein) with a low-salt phosphate buffer. VLPs were purified using both cesium chloride (CsCl) density gradient and size exclusion chromatography. Electron microscopy studies confirmed the presence of assembled forms of HPV16 L1 VLPs. Collectively; our results indicated that chloroplast-targeted transient expression in tobacco plants is promising for the production of a cheap, efficacious HPV16 L1 VLP vaccine. Studies are underway to develop plant VLPs for the production of a cervical cancer vaccine.