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1.
Clin Exp Ophthalmol ; 48(4): 500-511, 2020 05.
Artigo em Inglês | MEDLINE | ID: mdl-31872542

RESUMO

BACKGROUND: Retinoblastoma is a rare intraocular malignancy in children. Current treatments have many adverse effects. New therapeutic approaches like intravitreal injections of chemotherapies are currently being developed but their toxicities need to be evaluated on animal models. This study compares the efficacy and toxicity of intravitreal melphalan, topotecan and carboplatin, alone or in combination (sequential administration), in the LHBetaTag retinoblastoma mice. METHODS: Mice were divided into nine groups: control, carboplatin 1.5 and 4 µg, melphalan 0.1 and 1 µg, topotecan 0.1 and 1 µg, carboplatin 4 µg/topotecan 0.1 µg and melphalan 1 µg/topotecan 0.1 µg. The follow-up was performed using fundus imaging and optical coherence tomography combined with histopathological analysis. Absence of tumour and presence of calcified tumours were the criteria for therapeutic response assessment. Ocular complications were assessed after four weekly injections. Retinal toxicity was defined by the decrease of retinal thickness and of the number of retinal layers. RESULTS: Topotecan was inactive on retinal tumours. Melphalan (1 µg) led to a complete tumour control in 91.7% of eyes. Carboplatin strongly decreased the tumour burden (85.7-93.8% of eyes without retinal tumour). The intravitreal injection itself led to ocular complications (25% of media opacities and 45.7% of retinal detachment). Only melphalan at 1 µg showed a strong retinal toxicity. The two combinations showed a good efficacy in reducing the number of eyes with retinal tumours with a reduced retinal toxicity. CONCLUSIONS: This preclinical study suggests that intravitreal injection of carboplatin has a low toxicity and could be evaluated in clinical practice to treat patients suffering from retinoblastoma.


Assuntos
Neoplasias da Retina , Retinoblastoma , Animais , Carboplatina/uso terapêutico , Humanos , Injeções Intravítreas , Melfalan/uso terapêutico , Melfalan/toxicidade , Camundongos , Retina , Neoplasias da Retina/tratamento farmacológico , Retinoblastoma/tratamento farmacológico , Estudos Retrospectivos
2.
Ophthalmic Res ; 59(3): 164-169, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-29587271

RESUMO

BACKGROUND: Due to the presence of speckle Poisson noise, the interpretation of spectral domain-optical coherence tomography (SD-OCT) images frequently requires the use of data averaging to improve the signal-to-noise ratio. This implies long acquisition times and requires patient sedation in some cases. Iterative variance stabilizing transformation (VST) is a possible approach by which to remove speckle Poisson noise on single images. METHODS: We used SD-OCT images of human and murine (LH Beta-Tag mouse model) retinas with and without retinoblastoma acquired with 2 different imaging devices (Bioptigen and Micron IV). These images were processed using a denoising workflow implemented in Matlab. RESULTS: We demonstrated the presence of speckle Poisson noise, which can be removed by a VST-based approach. This approach is robust as it works in all used imaging devices and in both human and mouse retinas, independently of the tumor status. The implemented algorithm is freely available from the authors on demand. CONCLUSIONS: On a single denoised image, the proposed method provides results similar to those expected from the SD-OCT averaging. Because of the friendly user interface, it can be easily used by clinicians and researchers in ophthalmology.


Assuntos
Algoritmos , Processamento de Imagem Assistida por Computador , Retina/patologia , Retinoblastoma/patologia , Tomografia de Coerência Óptica/métodos , Animais , Humanos , Camundongos , Neoplasias Experimentais/patologia , Razão Sinal-Ruído
3.
Vet Res ; 44: 43, 2013 Jun 19.
Artigo em Inglês | MEDLINE | ID: mdl-23783173

RESUMO

Bovine Viral Diarrhoea Virus (BVDV) is a pestivirus which infects cattle populations worldwide and is recognised as a significant source of economic loss through its impact on health and productivity. Studies investigating the molecular epidemiology of BVDV can give invaluable information about the diversity of viral strains present in a population and this, in turn, can inform control programs, drive vaccine development and determine likely infection sources. The current study investigated 104 viral isolates from forty farms across the UK. Through phylogenetic and nucleotide sequence analysis of the 5'UTR and Npro regions of the isolates investigated, it was determined that BVDV 1a was the predominant sub-genotype. However, BVDV 1b, 1e and 1i were also identified and, for the first time in the UK, BVDV 1d. Through analysis of animal movement data alongside the phylogenetic analysis of these BVD isolates, it was possible to link animal movements to the viral isolates present on several premises and, for the first time, begin to elucidate the routes of viral transmission. With further work, this type of analysis would enable accurate determination and quantification of the true biosecurity risk factors associated with BVDV transmission.


Assuntos
Doença das Mucosas por Vírus da Diarreia Viral Bovina/epidemiologia , Vírus da Diarreia Viral Bovina Tipo 1/genética , Variação Genética , Criação de Animais Domésticos , Animais , Doença das Mucosas por Vírus da Diarreia Viral Bovina/virologia , Bovinos , Vírus da Diarreia Viral Bovina Tipo 1/classificação , Vírus da Diarreia Viral Bovina Tipo 1/isolamento & purificação , Inglaterra/epidemiologia , Epidemiologia Molecular , Dados de Sequência Molecular , Filogenia , RNA Viral/genética , Escócia/epidemiologia , Análise de Sequência de DNA/veterinária , Meios de Transporte , País de Gales/epidemiologia
4.
Artigo em Inglês | MEDLINE | ID: mdl-23295482

RESUMO

Bovine viral diarrhoea virus (BVDV) is an economically important animal pathogen which is closely related to Hepatitis C virus. Of the structural proteins, the envelope glycoprotein E2 of BVDV is the major antigen which induces neutralizing antibodies; thus, BVDV E2 is considered as an ideal target for use in subunit vaccines. Here, the expression, purification of wild-type and mutant forms of the ectodomain of BVDV E2 and subsequent crystallization and data collection of two crystal forms grown at low and neutral pH are reported. Native and multiple-wavelength anomalous dispersion (MAD) data sets have been collected and structure determination is in progress.


Assuntos
Vírus da Diarreia Viral Bovina Tipo 1/química , Proteínas do Envelope Viral/química , Proteínas do Envelope Viral/isolamento & purificação , Sequência de Bases , Clonagem Molecular , Cristalização/métodos , Cristalografia por Raios X/métodos , Dados de Sequência Molecular , Conformação Proteica , Proteínas do Envelope Viral/genética
5.
PLoS Pathog ; 6(9): e1001119, 2010 Sep 23.
Artigo em Inglês | MEDLINE | ID: mdl-20886105

RESUMO

Plasmodesmata (PD) are essential but poorly understood structures in plant cell walls that provide symplastic continuity and intercellular communication pathways between adjacent cells and thus play fundamental roles in development and pathogenesis. Viruses encode movement proteins (MPs) that modify these tightly regulated pores to facilitate their spread from cell to cell. The most striking of these modifications is observed for groups of viruses whose MPs form tubules that assemble in PDs and through which virions are transported to neighbouring cells. The nature of the molecular interactions between viral MPs and PD components and their role in viral movement has remained essentially unknown. Here, we show that the family of PD-located proteins (PDLPs) promotes the movement of viruses that use tubule-guided movement by interacting redundantly with tubule-forming MPs within PDs. Genetic disruption of this interaction leads to reduced tubule formation, delayed infection and attenuated symptoms. Our results implicate PDLPs as PD proteins with receptor-like properties involved the assembly of viral MPs into tubules to promote viral movement.


Assuntos
Doenças das Plantas/virologia , Proteínas do Movimento Viral em Plantas/metabolismo , Vírus de Plantas/fisiologia , Plasmodesmos/metabolismo , Plasmodesmos/virologia , Receptores de Superfície Celular/metabolismo , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Arabidopsis/virologia , Comunicação Celular , Parede Celular/metabolismo , Chenopodium quinoa/crescimento & desenvolvimento , Chenopodium quinoa/metabolismo , Chenopodium quinoa/virologia , Immunoblotting , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Folhas de Planta/virologia , Transporte Proteico , RNA Viral/genética , Nicotiana/crescimento & desenvolvimento , Nicotiana/metabolismo , Nicotiana/virologia
6.
Proc Natl Acad Sci U S A ; 106(9): 3615-20, 2009 Mar 03.
Artigo em Inglês | MEDLINE | ID: mdl-19218459

RESUMO

Cell-to-cell transport in plants occurs through cytoplasmic channels called "plasmodesmata" and is regulated by developmental and environmental factors. Callose deposition modulates plasmodesmal transport in vivo, but little is known about the mechanisms that regulate this process. Here we report a genetic approach to identify mutants affecting plasmodesmal transport. We isolated 5 mutants, named gfp arrested trafficking (gat), affected in GFP unloading from the phloem into the meristem. gat1 mutants were seedling lethal and carried lesions in an m-type thioredoxin that is expressed in non-green plastids of meristems and organ primordia. Callose and hydrogen peroxide accumulated in gat1 mutants, and WT plants subjected to oxidative conditions phenocopied the gat1 trafficking defects. Ectopic expression of GAT1 in mature leaves increased plasmodesmal permeability and led to a delay in senescence and flowering time. We propose a role for the GAT1 thioredoxin in the redox regulation of callose deposition and symplastic permeability that is essential for meristem maintenance in Arabidopsis.


Assuntos
Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Meristema/crescimento & desenvolvimento , Meristema/metabolismo , Tiorredoxinas/metabolismo , Arabidopsis/genética , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Transporte Biológico , Regulação da Expressão Gênica de Plantas , Proteínas de Membrana Transportadoras/genética , Proteínas de Membrana Transportadoras/metabolismo , Meristema/genética , Fenótipo , Plantas Geneticamente Modificadas , Plântula/genética , Plântula/crescimento & desenvolvimento , Plântula/metabolismo , Tiorredoxinas/genética
7.
Pharmaceuticals (Basel) ; 15(8)2022 Aug 10.
Artigo em Inglês | MEDLINE | ID: mdl-36015132

RESUMO

PURPOSES: The aim of the study was to assess the efficacy of a treatment protocol that combines photodynamic therapy (PDT) and nitroglycerin (NG) on human retinoblastoma tumors xenografted on mice. We aimed to increase the PDT efficiency (in our least treatment-responsive retinoblastoma line) with better PS delivery to the tumor generated by NG, which is known to dilate vessels and enhance the permeability and retention of macromolecules in solid tumors. METHODS: In vivo follow-up of the therapeutic effects was performed by sodium MRI, which directly monitors variations in sodium concentrations non-invasively and can be used to track the tumor response to therapy. NG ointment was applied one hour before PDT. The PDT protocol involves double-tumor targeting, i.e., cellular and vascular. The first PS dose was injected followed by a second one, separated by a 3 h interval. The timelapse allowed the PS molecules to penetrate tumor cells. Ten minutes after the second dose, the PS was red-light-activated. RESULTS: In this study, we observed that the PDT effect was enhanced by applying nitroglycerin ointment to the tumor-bearing animal's skin. PDT initiates the bystander effect on retinoblastomas, and NG increases this effect by increasing the intratumoral concentration of PS, which induces a higher production of ROS in the illuminated region and thus increases the propagation of the cell death signal deeper into the tumor (bystander effect).

8.
PLoS Biol ; 6(1): e7, 2008 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-18215111

RESUMO

Plasmodesmata provide the cytoplasmic conduits for cell-to-cell communication throughout plant tissues and participate in a diverse set of non-cell-autonomous functions. Despite their central role in growth and development and defence, resolving their modus operandi remains a major challenge in plant biology. Features of protein sequences and/or structure that determine protein targeting to plasmodesmata were previously unknown. We identify here a novel family of plasmodesmata-located proteins (called PDLP1) whose members have the features of type I membrane receptor-like proteins. We focus our studies on the first identified type member (namely At5g43980, or PDLP1a) and show that, following its altered expression, it is effective in modulating cell-to-cell trafficking. PDLP1a is targeted to plasmodesmata via the secretory pathway in a Brefeldin A-sensitive and COPII-dependent manner, and resides at plasmodesmata with its C-terminus in the cytoplasmic domain and its N-terminus in the apoplast. Using a deletion analysis, we show that the single transmembrane domain (TMD) of PDLP1a contains all the information necessary for intracellular targeting of this type I membrane protein to plasmodesmata, such that the TMD can be used to target heterologous proteins to this location. These studies identify a new family of plasmodesmal proteins that affect cell-to-cell communication. They exhibit a mode of intracellular trafficking and targeting novel for plant biology and provide technological opportunities for targeting different proteins to plasmodesmata to aid in plasmodesmal characterisation.


Assuntos
Proteínas de Arabidopsis/metabolismo , Proteínas de Transporte/metabolismo , Comunicação Celular , Plasmodesmos/metabolismo , Sequência de Aminoácidos , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Arabidopsis/metabolismo , Proteínas de Arabidopsis/química , Proteínas de Arabidopsis/classificação , Proteínas de Arabidopsis/genética , Proteínas de Transporte/química , Proteínas de Transporte/classificação , Proteínas de Transporte/genética , Matriz Extracelular/metabolismo , Peptídeos e Proteínas de Sinalização Intracelular , Dados de Sequência Molecular , Mutação/genética , Filogenia , Transporte Proteico , Alinhamento de Sequência , Transdução de Sinais
9.
Front Vet Sci ; 8: 659330, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33898551

RESUMO

Bovine Viral Diarrhea virus (BVDV) is a pestivirus with a single-stranded, positive sense RNA genome. It is endemic in many cattle populations, causing major economic losses in part due to reduced fertility. BVDV exhibits great genetic diversity and is classified as type 1 or 2 (BVDV-1, BVDV-2) with either non-cytopathogenic (ncp) or cytopathogenic (cp) biotypes. Differing strains of ncpBVDV differ in virulence, affecting clinical outcome. BVDV replicates in the reproductive tract, affecting host immunity and embryo survival. This study used an in vitro model of primary bovine endometrial cell cultures to compare the effects of two BVDV ncp type 1a strains of differing virulence (termed HO and KY) on endometrial transcription of candidate interferon stimulated genes (ISG) using qPCR. Half the cultures were stimulated with interferon tau (IFNT, the conceptus produced pregnancy recognition factor) in the presence or absence of viral infection. Cultures were replicated on cells from 10 BVDV-free cows. IFNT treatment stimulated transcription of 10 candidate ISGs, whereas both ncpBVDV-1 strains alone inhibited transcription of 8/10 ISGs. In combined BVDV-1+IFNT cultures, the stimulatory effect of IFNT on expression of GBP4, ISG15, HERC5, RSAD2, IFIH1, IFIT3, and MX1 was significantly inhibited by HO, but only ISG15, RSAD2, IFI27, and IFIT3 were decreased by KY. Inhibition by HO was generally greater. The IFNT-induced expression of TRIM56 was, however, increased by HO. These data show that HO, the more virulent ncpBVDV-1 strain, has a greater capacity to inhibit key antiviral pathways. These differences need confirmation at the protein level but may influence immune tolerance of the host. They could also reduce fertility by increasing uterine susceptibility to bacterial infection and disrupting IFNT-mediated pregnancy recognition.

10.
Plant J ; 59(3): 426-36, 2009 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-19392692

RESUMO

In Arabidopsis thaliana, auxin is a key regulator of tissue patterning in the developing embryo. We have identified a group of proteins that act downstream of auxin accumulation in auxin-mediated root and vascular development in the embryo. Combined mutations in OBERON1 (OBE1) and OBERON2 (OBE2) give rise to obe1 obe2 double mutant seedlings that closely phenocopy the monopteros (mp) mutant phenotype, with an absence of roots and defective development of the vasculature. We show that, in contrast to the situation in mp mutants, obe1 obe2 double mutant embryos show auxin maxima at the root pole and in the provascular region, and that the SCF(TIR1) pathway, which translates auxin accumulation into transcriptional activation of auxin-responsive genes, remains intact. Although we focus on the impact of obe mutations on aspects of embryo development, the effect of such mutations on a broad range of auxin-related gene expression and the tissue expression patterns of OBE genes in seedlings suggest that OBE proteins have a wider role to play in growth and development. We suggest that OBE1 and OBE2 most likely control the transcription of genes required for auxin responses through the action of their PHD finger domains.


Assuntos
Proteínas de Arabidopsis/metabolismo , Arabidopsis/metabolismo , Proteínas de Homeodomínio/metabolismo , Ácidos Indolacéticos/metabolismo , Meristema/crescimento & desenvolvimento , Raízes de Plantas/crescimento & desenvolvimento , Arabidopsis/genética , Arabidopsis/crescimento & desenvolvimento , Proteínas de Arabidopsis/genética , Regulação da Expressão Gênica de Plantas , Proteínas de Homeodomínio/genética , Mutação , Fenótipo , Plantas Geneticamente Modificadas/genética , Plantas Geneticamente Modificadas/crescimento & desenvolvimento , Plantas Geneticamente Modificadas/metabolismo , Transdução de Sinais
11.
Engineering (Beijing) ; 6(1): 26-33, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-32288965

RESUMO

Many viral diseases are endemic in cattle populations worldwide. The ability of many viruses to cross the placenta and cause abortions and fetal malformations is well understood. There is also significant evidence that viral infections have additional actions in dairy cows, which are reflected in reduced conception rates. These effects are, however, highly dependent on the time at which an individual animal first contracts the disease and are less easy to quantify. This paper reviews the evidence relating to five viruses that can affect fertility, together with their potential mechanisms of action. Acute infection with non-cytopathic bovine viral diarrhea virus (BVDV) in mid-gestation increases abortion rates or causes the birth of persistently infected calves. BVDV infections closer to the time of breeding can have direct effects on the ovaries and uterine endometrium, which cause estrous cycle irregularities and early embryo mortality. Fertility may also be reduced by BVDV-induced immunosuppression, which increases the susceptibility to bacterial infections. Bovine herpesvirus (BHV)-1 is most common in pre-pubertal heifers, and can slow their growth, delay breeding, and increase the age at first calving. Previously infected animals subsequently show reduced fertility. Although this may be associated with lung damage, ovarian lesions have also been reported. Both BHV-1 and BHV-4 remain latent in the host following initial infection and may be reactivated later by stress, for example associated with calving and early lactation. While BHV-4 infection alone may not reduce fertility, it appears to act as a co-factor with established bacterial pathogens such as Escherichia coli and Trueperella pyogenes to promote the development of endometritis and delay uterine repair mechanisms after calving. Both Schmallenberg virus (SBV) and bluetongue virus (BTV) are transmitted by insect vectors and lead to increased abortion rates and congenital malformations. BTV-8 also impairs the development of hatched blastocysts; furthermore, infection around the time of breeding with either virus appears to reduce conception rates. Although the reductions in conception rates are often difficult to quantify, they are nevertheless sufficient to cause economic losses, which help to justify the benefits of vaccination and eradication schemes.

12.
Vet Rec ; 187(7): e47, 2020 Oct 03.
Artigo em Inglês | MEDLINE | ID: mdl-32054720

RESUMO

BACKGROUND: Bovine viral diarrhoea (BVD) is a production disease commonly found in British cattle herds. Species other than cattle have been shown to be infected with the virus, thereby providing a potential source of infection for livestock. This study surveyed serum samples taken from 596 culled wild deer from England and Wales, between 2009 and 2010, for the presence of BVD antibodies. METHODS: 596 samples were tested with the SVANOVIR BVDV p80-Ab ELISA and a subset of 64 were tested with the IDEXX BVDV p80-Ab ELISA. ELISA results were confirmed using serum neutralisation tests. RESULTS: 2/596 samples (0.35 per cent) tested positive for BVD antibodies using the Svanova test and one of these tested positive and the other inconclusive using the IDEXX test; both were confirmed positive with serum neutralisation tests. These were both red deer stags, one from Devon and the other from East Anglia. CONCLUSIONS: The results indicate that it is unlikely that BVD virus is widely circulating within the wild deer population and particularly unlikely that persistently infected deer are present in the populations surveyed. These results suggest that wild deer are unlikely to be a significant reservoir of BVD infection in cattle.


Assuntos
Animais Selvagens/virologia , Cervos/virologia , Vírus da Diarreia Viral Bovina/isolamento & purificação , Animais , Anticorpos Antivirais/sangue , Vírus da Diarreia Viral Bovina/imunologia , Inglaterra , Ensaio de Imunoadsorção Enzimática/veterinária , País de Gales
13.
Anticancer Res ; 29(6): 1933-41, 2009 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-19528450

RESUMO

BACKGROUND: Although well-acknowledged in vivo, spontaneous death of cancer cells in vitro is less widely appreciated. MATERIALS AND METHODS: Colony formation was studied in untreated control plates of standard clonogenic assays and measurements of actual and potential doubling times performed in asynchronous cultures of human cancer cells lines. Western blotting of lung large cell carcinoma, COR-L23 cells actively undergoing spontaneous cell death was also carried out. RESULTS: Catastrophic disintegration of mature colonies could be seen in the untreated plates of lung large cell carcinoma, H460 and colon adenocarcinoma, SW620 human cancer cell lines and a significant cell loss factor was present in the cell lines growing as adherent cells in continuous culture. Western blotting demonstrated alterations of relative cyclin dependent kinase (Cdk)1 to Cdk4 protein expression in dying COR-L23 cells. CONCLUSION: The phenomenon of spontaneous cell death should be considered a hallmark of cancer and may be the result of failure to stabilise unstable, fully developed cancer cells due to the disruption of Cdk1/Cdk4 co-expression in those cells.


Assuntos
Proteína Quinase CDC2/metabolismo , Carcinoma de Células Grandes/patologia , Quinase 4 Dependente de Ciclina/metabolismo , Neoplasias Pulmonares/patologia , Regressão Neoplásica Espontânea/patologia , Western Blotting , Carcinoma de Células Grandes/metabolismo , Sobrevivência Celular , Ensaio de Unidades Formadoras de Colônias , Fibroblastos/metabolismo , Citometria de Fluxo , Humanos , Neoplasias Pulmonares/metabolismo , Pele/citologia , Pele/metabolismo , Células Tumorais Cultivadas
14.
Anim Health Res Rev ; 20(1): 72-85, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-31895016

RESUMO

Bovine viral diarrhea virus (BVDV) is an important infectious disease agent that causes significant reproductive and economic losses in the cattle industry worldwide. Although BVDV infection is known to cause poor fertility in cattle, a greater part of the underlying mechanisms particularly associated with early reproductive losses are not clearly understood. Previous studies reported viral compromise of reproductive function in infected bulls. In females, BVDV infection is thought to be capable of killing the oocyte, embryo or fetus directly, or to induce lesions that result in fetal abortion or malformation. BVDV infections may also induce immune dysfunction, and predispose cattle to other diseases that cause poor health and fertility. Other reports also suggested BVDV-induced disruption of the reproductive endocrine system, and a disruption of leukocyte and cytokine functions in the reproductive organs. More recent studies have provided evidence of viral-induced suppression of endometrial innate immunity that may predispose to uterine disease. Furthermore, there is new evidence that BVDV may potentially disrupt the maternal recognition of pregnancy or the immune protection of the conceptus. This review brings together the previous reports with the more recent findings, and attempts to explain some of the mechanisms linking this important virus to infertility in cattle.


Assuntos
Aborto Animal/virologia , Doença das Mucosas por Vírus da Diarreia Viral Bovina/complicações , Infertilidade/veterinária , Complicações Infecciosas na Gravidez/veterinária , Animais , Doença das Mucosas por Vírus da Diarreia Viral Bovina/imunologia , Bovinos , Vírus da Diarreia Viral Bovina/imunologia , Feminino , Infertilidade/virologia , Gravidez , Complicações Infecciosas na Gravidez/virologia
15.
Int J Oncol ; 32(4): 895-907, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18360717

RESUMO

Despite major advances in the molecular biology of the cancer cell over the past two decades, the great majority of patients are still treated by conventional cytotoxic drugs. The chemotherapy regimens employed frequently include platinating agents, taxanes, intercalating agents and topoisomerase inhibitors. Attempts to predict the therapeutic efficacy of such drugs by molecular profiling (theranostics) have up to the present time had limited success. Genes responsible for the control of cell division, senescence and apoptosis whose normal functions become corrupted during carcinogenesis, might potentially play a part in determining chemotherapeutic response. Here we have examined the relationships between the chemoresponsiveness of 18 human in vitro cancer cell lines and proteomic expression of Ras, cyclins B1 and D1 and cyclin-dependent kinases Cdk1 and Cdk4. When all 18 cell lines were examined as a single group, proteomic expression did not provide any helpful theranostic predictors. Clear relationships between proteomic expression and drug efficacy emerged, however, when Ras, cyclin B1, cyclin D1, Cdk1 and Cdk4 were examined separately in p53 wild-type and p53 mutant cell subsets. We suggest that the theranostic relationships we have detected in vitro may have potential relevance in vivo and should prompt clinical theranostic studies which take account of p53 mutational status.


Assuntos
Quinases Ciclina-Dependentes/análise , Ciclinas/análise , Genes p53 , Mutação , Neoplasias/tratamento farmacológico , Proteômica , Proteínas ras/análise , Linhagem Celular Tumoral , Cisplatino/farmacologia , Humanos , Modelos Lineares , Neoplasias/química , Neoplasias/genética
16.
J Exp Ther Oncol ; 7(3): 237-54, 2008.
Artigo em Inglês | MEDLINE | ID: mdl-19066132

RESUMO

Tumour heterogeneity is becoming increasingly important as an obstacle to genomic and proteomic technologies designed to improve the diagnosis and treatment of human cancer. In a panel of 19 human in-vitro cancer cell lines, we show marked heterogeneity of proteomic expression of key genes responsible for the control of cell division and death. Patterns of expression of these proteins were unique for each cell line. In addition, dynamic heterogeneity of proteomic expression of Cyclin D1, Cdk1, Cdk4 and even actin was detected. The relative levels of each protein fluctuated independently from experiment to experiment separated only by short passages in tissue culture. Cdk1 and Cdk4 proteomic co-expression (Seabra, 2007) was not, however, affected by dynamic heterogeneity, or, in 4 cell lines, by treatment with D0.1 doses of CDDP. Cdk1/Cdk4 may thus provide a complex molecular target for anti-cancer drug development which is unaffected by tumour heterogeneity and is not disrupted by conventional chemotherapy.


Assuntos
Proteína Quinase CDC2/metabolismo , Quinase 4 Dependente de Ciclina/metabolismo , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Neoplasias/enzimologia , Antineoplásicos/farmacologia , Western Blotting , Proliferação de Células/efeitos dos fármacos , Ciclina D1/metabolismo , Humanos , Neoplasias/patologia , Células Tumorais Cultivadas , Ensaio Tumoral de Célula-Tronco
17.
Invest Ophthalmol Vis Sci ; 58(7): 3055-3064, 2017 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-28622397

RESUMO

Purpose: Because retinoblastoma therapies have many adverse effects, new approaches must be developed and evaluated on animal models. We describe orthotopic xenograft models of retinoblastoma using different strains of mice, suitable for this purpose. Methods: Human retinoblastoma tumors were established on immunodeficient mice by subcutaneous engraftment of tumors from enucleated eyes. The orthotopic model was obtained by subretinal injections of suspension cells into the right eye of immunodeficient (Swiss-nude, severe combined immunodeficiency [SCID]) and immunocompetent mice (C57BL/6N, B6Albino). In vivo tumor growth was monitored by fundus and spectral-domain optical coherence tomography (SD-OCT) imaging and compared with histology. Results: Retinal and vitreal tumor growth was achieved both in immunocompetent and immunodeficient strains after the subretinal injection of tumor cells. The best tumor engraftment rate was obtained in the SCID mice (68.8%). No tumor growth was observed in the C57BL/6N strain. Chronic retinal detachment may occur in most strains after the subretinal injection, in particular the Swiss-nude strain, which exhibits retinal degeneration. Conclusions: The setting up of an orthotopic mouse model depends mainly on the choice of the engrafted cells (cell lines or patient-derived xenografts) but it can also depend on the xenografted mouse strain. Severe combined immunodeficiency mice (an immunodeficient strain) achieved the best tumor engraftment rate (68.8%). However, intraocular tumor growth was also satisfactory (50%) in the immunocompetent strain B6Albino, and this strain will allow to exploit the immune response after a tumor treatment. Both of these strains may therefore be recommended when setting up orthotopic retinoblastoma xenografts.


Assuntos
Neoplasias Experimentais , Retina/patologia , Neoplasias da Retina/diagnóstico , Retinoblastoma/diagnóstico , Animais , Linhagem Celular Tumoral , Progressão da Doença , Feminino , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos SCID , Tomografia de Coerência Óptica
18.
J Neuroimaging ; 15(1): 92-6, 2005 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-15574583

RESUMO

A 54-year-old woman presented for cardiac evaluation of atypical chest pain. Workup included coronary angiography and a left ventriculogram, during which air was inadvertently injected, resulting in the development of an acute right hemisphere syndrome. Right carotid angiography was immediately performed, yielding only a delayed diffuse venous phase without focal vessel cutoffs. Within 60 minutes, the patient underwent hyperbaric oxygen therapy for the suspected cerebral air emboli. After removal from the chamber for technical reasons, she had a generalized tonic-clonic seizure, and further hyperbaric oxygen therapy was withheld. Initial computed tomography imaging obtained approximately 8 hours after symptom onset showed signs of early right hemispheric edema. Subsequent magnetic resonance imaging studies were markedly abnormal and suggestive of diffuse bilateral but predominantly right-sided parietal lobe edema with mildly positive diffusion-weighted imaging. Follow-up magnetic resonance imaging at 6 months was normal, and the patient's neurological examination returned to normal.


Assuntos
Embolia Aérea/terapia , Oxigenoterapia Hiperbárica , Embolia Intracraniana/terapia , Embolia Aérea/diagnóstico por imagem , Feminino , Humanos , Embolia Intracraniana/diagnóstico por imagem , Pessoa de Meia-Idade , Tomografia Computadorizada por Raios X
19.
Photodiagnosis Photodyn Ther ; 12(2): 267-75, 2015 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-25638484

RESUMO

BACKGROUND: PDT represents a very localized and non-mutagen antitumoral treatment using a photosensitive molecule (porphyrin family) light activated. The first way of cell damage is a direct one, active on the very site where ROSs have been produced. The second one is indirect by activating and transmitting the processes of cellular death signaling. In order to seek for a better characterization of the photo-biology involved in in vivo PDT and to better understand the differences on the treatment outcome, we have used three different human retinoblastomas xenografted on mice. METHODS: Mice were treated according to the double targeting protocol exposed in a previous paper. One i.v. dose (0.6 mg/kg) of PS was followed by a second dose, separated by a 3 h interval (double targeting PDT). As a consequence both cancer cells and blood vessels were targeted. The treatment was repeated two times, at 4 days interval. RESULTS: First of all, sodium MRI revealed qualitative differences in the sodium average content of the three retinoblastoma lines before treatment. After the PDT treatments the tumor responses were different between the lines as revealed by sodium MRI and later on by histology. CONCLUSIONS: We have put into evidence that PDT is accompanied by a bystander effect that may propagate the cellular death triggered by the initial photoreaction. This effect is highly dependent on the cellular density of the tissue; therefore this factor is to be taken into account in clinical PDT protocols.


Assuntos
Morte Celular/efeitos dos fármacos , Neovascularização Patológica/tratamento farmacológico , Fotoquimioterapia/métodos , Fármacos Fotossensibilizantes/farmacologia , Retinoblastoma/tratamento farmacológico , Animais , Contagem de Células , Linhagem Celular Tumoral , Humanos , Imageamento por Ressonância Magnética , Camundongos , Camundongos Nus , Espécies Reativas de Oxigênio/metabolismo
20.
Vaccine ; 33(32): 4004-12, 2015 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-26079613

RESUMO

DNA vaccination is effective in inducing potent immunity in mice; however it appears to be less so in large animals. Increasing the dose of DNA plasmid to activate innate immunity has been shown to improve DNA vaccine adaptive immunity. Retinoic acid-inducible gene I (RIG-I) is a critical cytoplasmic double-stranded RNA pattern receptor required for innate immune activation in response to viral infection. RIG-I recognise viral RNA and trigger antiviral response, resulting in type I interferon (IFN) and inflammatory cytokine production. In an attempt to enhance the antibody response induced by BVDV DNA in cattle, we expressed BVDV truncated E2 (E2t) and NS3 codon optimised antigens from antibiotic free-plasmid vectors expressing a RIG-I agonist and designated either NTC E2t(co) and NTC NS3(co). To evaluate vaccine efficacy, groups of five BVDV-free calves were intramuscularly injected three times with NTC E2t(co) and NTC NS3(co) vaccine plasmids individually or in combination. Animals vaccinated with our (previously published) conventional DNA vaccines pSecTag/E2 and pTriExNS3 and plasmids expressing RIG-I agonist only presented both the positive and mock-vaccine groups. Our results showed that vaccines coexpressing E2t with a RIG-I agonist induced significantly higher E2 antigen specific antibody response (p<0.05). Additionally, E2t augmented the immune response to NS3 when the two vaccines were delivered in combination. Despite the lack of complete protection, on challenge day 4/5 calves vaccinated with NTC E2t(co) alone or NTC E2t(co) plus NTC NS3(co) had neutralising antibody titres exceeding 1/240 compared to 1/5 in the mock vaccine control group. Based on our results we conclude that co-expression of a RIG-I agonist with viral antigen could enhance DNA vaccine potency in cattle.


Assuntos
Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , Doença das Mucosas por Vírus da Diarreia Viral Bovina/prevenção & controle , Vírus da Diarreia Viral Bovina/imunologia , Vacinas de DNA/imunologia , Vacinas Virais/imunologia , Animais , Bovinos , Vírus da Diarreia Viral Bovina/genética , Modelos Animais de Doenças , Expressão Gênica , Vetores Genéticos , Injeções Intramusculares , Peptídeo Hidrolases/genética , Peptídeo Hidrolases/imunologia , Plasmídeos , RNA Helicases/genética , RNA Helicases/imunologia , Resultado do Tratamento , Vacinas de DNA/administração & dosagem , Vacinas de DNA/genética , Proteínas do Envelope Viral/genética , Proteínas do Envelope Viral/imunologia , Proteínas não Estruturais Virais/genética , Proteínas não Estruturais Virais/imunologia , Vacinas Virais/administração & dosagem , Vacinas Virais/genética
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