RESUMO
We have developed a reflection-mode switchable subwavelength Bessel-beam (BB) and Gaussian-beam (GB) photoacoustic microscopy (PAM) system. To achieve both reflection-mode and high resolution, we tightly attached a very small ultrasound transducer to an optical objective lens with numerical aperture of 1.0 and working distance of 2.5 mm. We used axicon and an achromatic doublet in our system to obtain the extended depth of field (DOF) of the BB. To compare the DOF performance achieved with our BB-PAM system against GB-PAM system, we designed our system so that the GB can be easily generated by simply removing the lenses. Using a 532 nm pulse laser, we achieved the lateral resolutions of 300 and 270 nm for BB-PAM and GB-PAM, respectively. The measured DOF of BB-PAM was approximately 229 µm, which was about 7× better than that of GB-PAM. We imaged the vasculature of a mouse ear using BB-PAM and GB-PAM and confirmed that the DOF of BB-PAM is much better than the DOF of GB-PAM. Thus, we believe that the high resolution achieved at the extended DOF by our system is very practical for wide range of biomedical research including red blood cell (RBC) migration in blood vessels at various depths and observation of cell migration or cell culture.
Assuntos
Microscopia/métodos , Fenômenos Ópticos , Técnicas Fotoacústicas/métodos , Animais , Orelha/diagnóstico por imagem , Desenho de Equipamento , Estudos de Viabilidade , Camundongos , Microscopia/instrumentação , Distribuição Normal , Técnicas Fotoacústicas/instrumentaçãoRESUMO
Lipotropes (methyl group containing nutrients, including methionine, choline, folate, and vitamin B(12)) are dietary methyl donors and cofactors that are involved in one-carbon metabolism, which is important for genomic DNA methylation reactions and nucleic acid synthesis. One-carbon metabolism provides methyl groups for all biological methylation pathways and is highly dependent on dietary supplementation of methyl nutrients. Nutrition is an important determinant of breast cancer risk and tumor behavior, and dietary intervention may be an effective approach to prevent breast cancer. Apoptosis is important for the regulation of homeostasis and tumorigenesis. The anti-apoptotic protein Bcl-2 may be a regulatory target in cancer therapy; controlling or modulating its expression may be a therapeutic strategy against breast cancer. In this study, the effects of lipotrope supplementation on the growth and death of human breast cancer cell lines T47D and MCF-7 were examined and found to inhibit growth of both T47D and MCF-7 cells. Furthermore, the ratios of apoptotic cells to the total number of cells were approximately 44% and 34% higher in the lipotrope-supplemented treatments of T47D and MCF-7 cancer cells, respectively, compared with the control treatments. More importantly, Bcl-2 protein expression was decreased by approximately 25% from lipotrope supplementation in T47D cells, suggesting that lipotropes can induce breast cancer cell death by direct downregulation of Bcl-2 protein expression. Cancer treatment failure is often correlated with Bcl-2 protein upregulation. These data may be useful in the development of effective nutritional strategies to prevent and reduce breast cancer in humans.
Assuntos
Neoplasias da Mama/patologia , Alimentos , Lipotrópicos/farmacologia , Linhagem Celular Tumoral , Núcleo Celular/efeitos dos fármacos , Núcleo Celular/metabolismo , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Fatores de TempoRESUMO
Genetic evidence has implicated several genes as being critical for the development of cardiomyocytes. Whereas a few of the targets of these genes and the pathways they constitute are known the majority of targets and the interrelationships of the pathways involved still remains largely unknown. The power of high-throughput analytical techniques like microarrays and real-time RT-PCR combined with the ability to selectively silence specific mRNA in model tissue culture systems can begin to fill in these gaps and increase our understanding of the molecular mechanisms of cell commitment and terminal differentiation. We have used microarray analysis and siRNA directed against the cardiac-specific transcription factor Nkx2.5 and one of its targets Cripto in P19 clone 6 (P19Cl6) cells to identify potential targets for these genes. We demonstrate Nkx2.5 affects genes that have been shown to be controlled by the canonical Wnt or TGFbeta/BMP signaling pathways. We also show that Cripto can regulate the critical stem cell gene Nanog and two Oct 4-regulated genes: Dppa2 and 4. Cripto also affects the formation of nitric oxide, a small signaling molecule that has been reported to be important for growth and development of cardiac and smooth muscle. It affects the nitric oxide system by regulating genes that control the levels of nitric oxide synthase mRNA concentration as well as the activation and bioavailability of the protein.
Assuntos
Diferenciação Celular/genética , Fator de Crescimento Epidérmico/genética , Proteínas de Homeodomínio/genética , Glicoproteínas de Membrana/genética , Miocárdio/citologia , Proteínas de Neoplasias/genética , Fatores de Transcrição/genética , Animais , Sequência de Bases , Western Blotting , Linhagem Celular , Primers do DNA , Etiquetas de Sequências Expressas , Proteína Homeobox Nkx-2.5 , Camundongos , Análise de Sequência com Séries de Oligonucleotídeos , RNA Interferente Pequeno , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
The proper nutritional status during the hormone-sensitive growth phases preceding first parturition can affect mammary development and subsequent lactation performance. We developed a compensatory nutrition regimen (CNR), which is designed to stimulate mammary growth by exploiting the biological characteristics of the energy restriction and compensatory growth phenomenon. In the present study, we examined the effect of compensatory growth induced only once during late gestation upon mammary development and subsequent lactation potential over 2 lactation cycles. Female rats were mated and randomly assigned to either the control or the CNR group. Control rats were offered the control diet (AIN-93G) throughout the experiment. CNR rats were subjected to 40% energy restriction during the first 10 d of gestation followed by free access to the control diet for the remainder of the experiment. Dams on the CNR produced 14% more milk than control dams (P = 0.12). Mammary cell proliferation rates were approximately 46% (P < 0.05) and 27% (P = 0.07) higher in the CNR group than in the control during late gestation and early lactation of the first lactation cycle, respectively. Caspase-3 enzyme activity was decreased 15% (P < 0.05) and 22% (P = 0.11) in mammary tissues from the CNR group compared with that from the controls during the first and second lactation cycles, respectively. These results indicate that compensatory growth induced only once during late gestation increases mammary cell proliferation and differentiation and decreases regression of mammary cells throughout consecutive lactation cycles.
Assuntos
Fenômenos Fisiológicos da Nutrição Animal , Lactação , Glândulas Mamárias Animais/crescimento & desenvolvimento , Animais , Peso Corporal , Caseínas/genética , Caspase 3 , Caspases/metabolismo , Divisão Celular , Dieta , Ingestão de Energia , Feminino , Idade Gestacional , Tamanho da Ninhada de Vivíparos , Glândulas Mamárias Animais/citologia , Glândulas Mamárias Animais/enzimologia , Ornitina Descarboxilase/genética , Ornitina Descarboxilase/metabolismo , Gravidez , RNA Mensageiro/análise , RatosRESUMO
Among the first nutrients to be linked to cancer were methyl group containing nutrients including methionine. Methionine and its metabolic derivatives are essential components in several indispensable biological reactions including protein synthesis, polyamine synthesis, and many transmethylation reactions. The purpose of this study was to determine the extent to which methionine excess affects the proliferation and gene expression of the human breast cancer cell line MCF-7. Cells were first grown in control medium; the medium was then replaced with either control or methionine-supplemented treatment media. We found that 5 and 10 g/L methionine significantly suppressed cell growth on day 1, and no further growth was detected after 3 d of treatment. Cell proliferation in the methionine treated group was significantly lower than that of the control group. Northern analysis revealed that expression of p53 in methionine-treated MCF-7 cells was approximately 70% lower than that of control cells. p53 is a key cell cycle regulatory protein that has been implicated in tumorigenesis and cancer progression. Alteration of the p53 tumor suppressor gene is the most common genetic change found in a wide variety of malignancies, including cancer. This study shows that excess methionine (5 g/L) inhibited proliferation of MCF-7 breast cancer cells, and down regulation of p53 is correlated with this inhibition. These findings may aid in the development of nutritional strategies for breast cancer therapy.
Assuntos
Neoplasias da Mama/patologia , Sobrevivência Celular/efeitos dos fármacos , Metionina/toxicidade , Divisão Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Feminino , Humanos , RNA Ribossômico 28S/efeitos dos fármacos , RNA Ribossômico 28S/genéticaAssuntos
Dermatite Herpetiforme/diagnóstico , Complicações na Gravidez/diagnóstico , Psoríase/diagnóstico , Aborto Induzido , Adulto , Dermatite Herpetiforme/tratamento farmacológico , Dermatite Herpetiforme/patologia , Diagnóstico Diferencial , Feminino , Humanos , Isotretinoína/uso terapêutico , Ceratolíticos/uso terapêutico , Fotoquimioterapia , Prednisolona/uso terapêutico , Gravidez , Complicações na Gravidez/tratamento farmacológico , Complicações na Gravidez/patologia , Segundo Trimestre da Gravidez , Psoríase/tratamento farmacológico , Psoríase/patologiaRESUMO
Lipotropes, a methyl group containing nutrients, including choline, methionine, folic acid, and vitamin B(12), are essential nutrients for humans. They are important methyl donors that interact in the metabolism of one-carbon units and are essential for the synthesis and methylation of deoxyribonucleic acid. The purpose of this study was to examine the effects of excess lipotropes on the growth of a human breast cancer cell line, MCF-7, and normal mammary cells, MCF-10A, in culture. Both cell lines were grown in basal culture medium for 24 h and then switched to medium supplemented with 50 times the amount of each lipotrope as basal culture medium (control). Although there were no significant differences in growth between treatments in either cell line, gene array and Northern analysis revealed that expression of bcl-2 was decreased in lipotrope-treated MCF-7 cells. The ability to induce tumor cell death could have many uses in the prevention and treatment of cancer. Bcl-2 regulates apoptosis and has been shown to directly affect the sensitivity of cancer cells to chemotherapy agents, and it is suggested that strategies designed to block Bcl-2 might prove useful in sensitizing tumor cells to chemotherapy-induced apoptosis. This study shows that although excess lipotropes do not inhibit the growth of breast cancer cells, they can down-regulate the bcl-2 gene, suggesting that lipotropes may increase the susceptibility of breast cancer cells to anticancer drugs.