RESUMEN
Cancer therapy, from malignant tumor inhibition to cellular eradication treatment, remains a challenge, especially regarding reduced side effects and low energy consumption during treatment. Hence, phytochemicals as cytotoxic sensitizers or photosensitizers deserve special attention. The dark and photo-response of Yemenite 'Etrog' leaf extracts applied to prostate PC3 cancer cells is reported here. An XTT cell viability assay along with light microscope observations revealed pronounced cytotoxic activity of the extract for long exposure times of 72 h upon concentrations of 175 µg/mL and 87.5 µg/mL, while phototoxic effect was obtained even at low concentration of 10.93 µg/mL and a short introduction period of 1.5 h. For the longest time incubation of 72 h and for the highest extract concentration of 175 µg/mL, relative cell survival decreased by up to 60% (below the IC50). In combined phyto-photodynamic therapy, a reduction of 63% compared to unirradiated controls was obtained. The concentration of extract in cells versus the accumulation time was inversely related to fluorescence emission intensity readings. Extracellular ROS production was also shown. Based on an ATR-FTIR analysis of the powdered leaves and their liquid ethanolic extract, biochemical fingerprints of both polar and non-polar phyto-constituents were identified, thereby suggesting their implementation as phyto-medicine and phyto-photomedicine.
Asunto(s)
Supervivencia Celular , Fotoquimioterapia , Extractos Vegetales , Hojas de la Planta , Neoplasias de la Próstata , Humanos , Masculino , Extractos Vegetales/farmacología , Fotoquimioterapia/métodos , Neoplasias de la Próstata/tratamiento farmacológico , Hojas de la Planta/química , Supervivencia Celular/efectos de los fármacos , Fármacos Fotosensibilizantes/farmacología , Células PC-3 , Especies Reactivas de Oxígeno/metabolismo , Yemen , Línea Celular Tumoral , Antineoplásicos Fitogénicos/farmacologíaRESUMEN
BACKGROUND: Citrus plants are prone to infection by different viroids which deteriorate their vigor and production. Citrus viroid V (CVd-V) is among the six citrus viroids, belongs to genus Apscaviroid (family Pospiviroidae) which induces symptoms of mild necrotic lesions on branches and cracks on trunk portion. METHODS AND RESULTS: A survey was conducted to evaluate the prevalence of CVd-V in core and non-core citrus cultivated areas of Punjab, Pakistan. A total of 154 samples from different citrus cultivars were tested for CVd-V infection by RT-PCR. The results revealed 66.66% disease incidence of CVd-V. Citrus cultivars Palestinia Sweet lime, Roy Ruby, Olinda Valencia, Kaghzi lime, and Dancy were identified as new citrus hosts of CVd-V for the first time from Pakistan. The viroid infection was confirmed by biological indexing on indicator host Etrog citron. The reported primers used for the detection of CVd-V did not amplify, rather showed non-specific amplification, which led to the designing of new primers. Whereas, new back-to-back designed primers (CVd-V AF1/CVd-V AR1) detected CVd-V successfully and obtained an expected amplified product of CVd-V with 294 bp. Sequencing analysis confirmed the new host of CVd-V showing 98-100% nucleotide sequence homology with those reported previously from other countries while 100% sequence homology to the isolates reported from Pakistan. Based on phylogenetic analysis using all CVd-V sequences in GenBank, two main CVd-V groups (I and II) were identified, and newly identified isolates during this study fall in the group I. CONCLUSION: The study revealed that there are some changes in the nucleotide sequences of CVd-V which made difficult for their detection using reported primers. All isolates of Pakistan showed high sequence homology with other isolates of CVd-V from Iran and USA whereas; the isolates from China, Japan, Tunisia, and Africa are distantly related. It is evident that CVd-V is spreading in all citrus cultivars in Pakistan.
Asunto(s)
Citrus , Viroides , Citrus/virología , Pakistán , Filogenia , Enfermedades de las Plantas , Túnez , Viroides/genéticaRESUMEN
Citrus exocortis viroid (CEVd) is the causal agent of citrus exocortis disease. We employed CEVd-infected 'Etrog' citron as a system to study the feedback regulation mechanism using transcriptome analysis in this study. Three months after CEVd infection, the transcriptome of fresh leaves was analyzed, and 1530 differentially expressed genes were detected. The replication of CEVd in citron induced upregulation of genes encoding key proteins that were involved in the RNA silencing pathway such as Dicer-like 2, RNA-dependent RNA polymerase 1, argonaute 2, argonaute 7, and silencing defective 3, as well as those genes encoding proteins that are related to basic defense responses. Many genes involved in secondary metabolite biosynthesis and chitinase activity were upregulated, whereas other genes related to cell wall and phytohormone signal transduction were downregulated. Moreover, genes encoding disease resistance proteins, pathogenicity-related proteins, and heat shock cognate 70 kDa proteins were also upregulated in response to CEVd infection. These results suggest that basic defense and RNA silencing mechanisms are activated by CEVd infection, and this information improves our understanding of the pathogenesis of viroids in woody plants.