RESUMEN
CONTEXT: Euphorbia hirta L. (Euphorbiaceae) has been used as a folk remedy in Southeast Asia for the treatment of various ailments. OBJECTIVE: The current study evaluates the cytotoxicity, cell-cycle arrest, and apoptotic induction by E. hirta in MCF-7 breast cancer cells. MATERIALS AND METHODS: Cytotoxic activity of methanol extract of whole part of E. hirta was determined by the MTT assay at various concentrations ranging from 1.96 to 250.00 µg/mL in MCF-7 cells. Cell morphology was assessed by light and fluorescence microscopy. Apoptosis and cell-cycle distribution were determined by annexin V staining and flow cytometry. DNA fragmentation, caspase activity, and reactive oxygen species (ROS) assays were performed using the commercially available kits. To identify the cytotoxic fraction, E. hirta extract was subjected to bioassay-guided fractionation. RESULTS: Euphorbia hirta exhibited significant inhibition of the survival of MCF-7 cells and the half inhibitory concentration (IC50) values was 25.26 µg/mL at 24 h. Microscopic studies showed that E. hirta-treated cells exhibited marked morphological features characteristic of apoptosis. Euphorbia hirta extract also had an ignorable influence on the LDH leakage and generating intracellular ROS. The flow cytometry study confirmed that E. hirta extract induced apoptosis in MCF-7 cells. Euphorbia hirta also resulted in DNA fragmentation in MCF-7 cells. Moreover, E. hirta treatment resulted in the accumulation of cells at the S and G2/M phases as well as apoptosis. The caspase activity study revealed that E. hirta extract induced apoptosis through the caspase-3-independent pathway by the activation of caspase-2, 6, 8, and 9. Euphorbia hirta hexane fraction, namely HFsub4 fraction, demonstrated highest activity among all the fractions tested with an IC50 value of 10.01 µg/mL at 24 h. DISCUSSION AND CONCLUSION: This study revealed that E. hirta induced apoptotic cell death and suggests that E. hirta could be used as an apoptosis-inducing anticancer agent for breast cancer treatment with further detailed studies.
Asunto(s)
Antineoplásicos Fitogénicos/farmacología , Apoptosis/efectos de los fármacos , Neoplasias de la Mama/tratamiento farmacológico , Euphorbia , Puntos de Control de la Fase G2 del Ciclo Celular/efectos de los fármacos , Extractos Vegetales/farmacología , Puntos de Control de la Fase S del Ciclo Celular/efectos de los fármacos , Animales , Antineoplásicos Fitogénicos/aislamiento & purificación , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Caspasas/metabolismo , Supervivencia Celular/efectos de los fármacos , Chlorocebus aethiops , Daño del ADN , Relación Dosis-Respuesta a Droga , Euphorbia/química , Femenino , Células HT29 , Células HeLa , Humanos , Concentración 50 Inhibidora , Células MCF-7 , Estrés Oxidativo/efectos de los fármacos , Fitoterapia , Extractos Vegetales/aislamiento & purificación , Plantas Medicinales , Especies Reactivas de Oxígeno/metabolismo , Factores de Tiempo , Células VeroRESUMEN
Both α- and ß-thalassaemia syndromes are public health problems in the multi-ethnic population of Malaysia. To molecularly characterise the α- and ß-thalassaemia deletions and mutations among Malays from Penang, Gap-PCR and multiplexed amplification refractory mutation systems were used to study 13 α-thalassaemia determinants and 20 ß-thalassaemia mutations in 28 and 40 unrelated Malays, respectively. Four α-thalassaemia deletions and mutations were demonstrated. --SEA deletion and αCSα accounted for more than 70% of the α-thalassaemia alleles. Out of the 20 ß-thalassaemia alleles studied, nine different ß-thalassaemia mutations were identified of which ßE accounted for more than 40%. We concluded that the highest prevalence of (α- and ß-thalassaemia alleles in the Malays from Penang are --SEA deletion and ßE mutation, respectively.
Asunto(s)
Pueblo Asiatico/genética , Talasemia alfa/genética , Talasemia beta/genética , Alelos , Genotipo , Haplotipos , Heterocigoto , Humanos , Malasia , Familia de Multigenes , Polimorfismo de Nucleótido Simple , Globinas alfa/genética , Talasemia alfa/patología , Talasemia beta/patologíaRESUMEN
BACKGROUND: Dengue virus is endemic in peninsular Malaysia. The clinical manifestations vary depending on the incubation period of the virus as well as the immunity of the patients. Glucose-6-phosphate dehydrogenase (G6PD) deficiency is prevalent in Malaysia where the incidence is 3.2%. It has been noted that some G6PD-deficient individuals suffer from more severe clinical presentation of dengue infection. In this study, we aim to investigate the oxidative responses of DENV2-infected monocytes from G6PD-deficient individuals. METHODOLOGY: Monocytes from G6PD-deficient individuals were infected with DENV2 and infection rate, levels of oxidative species, nitric oxide (NO), superoxide anions (O2-), and oxidative stress were determined and compared with normal controls. PRINCIPAL FINDINGS: Monocytes from G6PD-deficient individuals exhibited significantly higher infection rates compared to normal controls. In an effort to explain the reason for this enhanced susceptibility, we investigated the production of NO and O2- in the monocytes of individuals with G6PD deficiency compared with normal controls. We found that levels of NO and O2- were significantly lower in the DENV-infected monocytes from G6PD-deficient individuals compared with normal controls. Furthermore, the overall oxidative stress in DENV-infected monocytes from G6PD-deficient individuals was significantly higher when compared to normal controls. Correlation studies between DENV-infected cells and oxidative state of monocytes further confirmed these findings. CONCLUSIONS/SIGNIFICANCE: Altered redox state of DENV-infected monocytes from G6PD-deficient individuals appears to augment viral replication in these cells. DENV-infected G6PD-deficient individuals may contain higher viral titers, which may be significant in enhanced virus transmission. Furthermore, granulocyte dysfunction and higher viral loads in G6PD-deificient individuals may result in severe form of dengue infection.
Asunto(s)
Virus del Dengue/inmunología , Virus del Dengue/fisiología , Deficiencia de Glucosafosfato Deshidrogenasa , Monocitos/inmunología , Monocitos/virología , Estrés Oxidativo , Adulto , Células Cultivadas , Virus del Dengue/efectos de los fármacos , Granulocitos/inmunología , Humanos , Malasia , Masculino , Óxido Nítrico/análisis , Especies Reactivas de Oxígeno/análisis , Superóxidos/análisis , Carga Viral , Replicación Viral/efectos de los fármacosRESUMEN
New drugs are continuously being developed for the treatment of patients with estrogen receptor-positive breast cancer. Thymoquinone is one of the drugs that exhibits anticancer characteristics based on in vivo and in vitro models. This study further investigates the effects of thymoquinone on human gene expression using cDNA microarray technology. The quantification of RNA samples was carried out using an Agilent 2100 Bioanalyser to determine the RNA integrity number (RIN). The Agilent Low Input Quick Amplification Labelling kit was used to generate cRNA in two-color microarray analysis. Samples with RIN >9.0 were used in this study. The universal human reference RNA was used as the common reference. The samples were labelled with cyanine-3 (cye-3) CTP dye and the universal human reference was labelled with cyanine-5 (cye-5) CTP dye. cRNA was purified with the RNeasy Plus Mini kit and quantified using a NanoDrop 2000c spectrophotometer. The arrays were scanned data analysed using Feature Extraction and GeneSpring software. Two-step qRT-PCR was selected to determine the relative gene expression using the High Capacity RNA-to-cDNA kit. The results from Gene Ontology (GO) analysis, indicated that 8 GO terms were related to biological processes (84%) and molecular functions (16%). A total of 577 entities showed >2-fold change in expression. Of these entities, 45.2% showed an upregulation and 54.7% showed a downregulation in expression. The interpretation of single experiment analysis (SEA) revealed that the cytochrome P450, family 1, subfamily A, polypeptide 1 (CYP1A1) and UDP glucuronosyltransferase 1 family, polypeptide A8 (UGT1A8) genes in the estrogen metabolic pathway were downregulated significantly by 43- and 11fold, respectively. The solute carrier family 7 (anionic amino acid transporter light chain, xc-system), member 11 (SLC7A11) gene in the interferon pathway, reported to be involved in the development of chemoresistance, was downregulated by 15fold. The interferon-induced protein with tetratricopeptide repeats (IFIT)1, IFIT2, IFIT3, interferon, α-inducible protein (IFI)6 (also known as G1P3), interferon regulatory factor 9 (IRF9, ISGF3), 2'-5'-oligoadenylate synthetase 1, 40/46 kDa (OAS1) and signal transducer and activator of transcription 1 (STAT1) genes all showed changes in expression following treatment with thymoquinone. The caspase 10, apoptosis-related cysteine peptidase (CASP10) gene was activated and the protein tyrosine phosphatase, receptor type, R (PTPRR) and myocyte enhancer factor 2C (MEF2C) genes were upregulated in the classical MAPK and p38 MAPK pathways. These findings indicate that thymquinone targets specific genes in the estrogen metabolic and interferon pathways.
Asunto(s)
Antineoplásicos/farmacología , Benzoquinonas/farmacología , Estrógenos/metabolismo , Interferones/metabolismo , Redes y Vías Metabólicas , Análisis por Conglomerados , Regulación hacia Abajo , Femenino , Regulación de la Expresión Génica , Humanos , Células MCF-7 , Análisis de Secuencia por Matrices de Oligonucleótidos , Transducción de Señal , Regulación hacia Arriba , Proteínas Quinasas p38 Activadas por Mitógenos/genética , Proteínas Quinasas p38 Activadas por Mitógenos/metabolismoRESUMEN
BACKGROUND: Nigella sativa or black seed extract has been reported to show various medicinal benefits. Thymoquinone which is an active compound of its seed has been reported to contain anti-cancer properties. OBJECTIVE: The study addressed the anti-cancer efficiency of long-term in vitro treatment with thymoquinone towards human breast cancer cell lines MCF-7. MATERIALS AND METHODS: Cell proliferation was determined with CellTiter 96 Aqueous. Non-Radioactive Cell Proliferation Assay Kit. It was followed with trypan blue exclusion test to determine the percentage of viable cells. The study incorporated cell cycle assay to distinguish cell distribution at various cell cycle phases using Cycletest Plus DNA Reagent Kit. The apoptosis detection kit was used to determine the percentage of apoptotic and necrotic cells using flow cytometry. RESULTS: The 50% inhibitory concentration (IC50) value determined using the proliferation assay was 25 µM thymoquinone. Late apoptotic cell percentage increased rapidly when treatment duration was increased to 24 h with 25 and 100 µM thymoquinone. Further analysis using cell cycle assay showed thymoquinone inhibition of breast cancer cell proliferation at minimal dose 25 µM and led to S phase arrest significantly at 72 h treatment (P = 0.009). It was also noted elevation sub-G1 peak following treatment with 25 µM thymoquinone for 12 h. Increase in thymoquinone to 50 µM caused G2 phase arrest at each time-point studied. CONCLUSION: In general thymoquinone showed sustained inhibition of breast cancer cell proliferation with long-term treatment. Specificity of phase arrest was determined by thymoquinone dose.
RESUMEN
BACKGROUND: Diego blood group antigen, Di(a), is very rare among Caucasians and Blacks, but relatively common among the South American Indians and Asians of Mongolian origin. The antibody to Di(a) is clinically significant to cause hemolytic disease in a new-born or hemolytic transfusion reaction. OBJECTIVES: This study was designed to determine the prevalence of Di(a) antigen among the blood donors from the three major ethnic groups in Klang Valley of Malaysia as well as to find an incidence of an antibody of the Diego antigen, anti-Di(a), in a tertiary care hospital to ascertain the need to include Di(a+) red cells for an antibody screen cell panel. MATERIALS AND METHODS: Serological tests were performed by column agglutination technique using commercial reagents and following instruction as per kit insert. RESULTS: Di(a) antigen was found with a frequency of 2.1% among the Malaysians donors in three ethnic groups viz, Malay, Chinese and Indian. It was present among 1.25% of 401 Malay, 4.01% of Chinese and 0.88% of 114 Indian origin donors. None of the 1442 patients, including 703 antenatal outpatients, had anti-Di(a) in serum. CONCLUSION: The prevalence of Di(a) antigen was found among the donors of all the three ethnic background with varying frequency. Inclusion of Di(a+) red cells in routine antibody screening program would certainly help in detection of this clinically significant antibody and to provide safe blood transfusion in the Klang Valley, though the incidence of antibody appears to be very low in the region.
RESUMEN
OBJECTIVE: To determine the optimum storage temperature and time for prothrombin time and activated partial thromboplastin time at various intervals at both room temperature and refrigerator. STUDY DESIGN: Experimental study. PLACE AND DURATION OF STUDY: Advanced Medical and Dental Institute (AMDI), Laboratory at University Sains Malaysia (USM), from August 2009 to June 2010. METHODOLOGY: After obtaining the consent, 33 blood samples were collected from AMDI staffs and students. Prothrombin time (PT) was measured at 0, 4, 8 and 24 hours (h). Partial thromboplastin time (APTT) was measured at 0, 2, 6 and 8 h both at room temperature (RT) and refrigerator. RESULTS: Thirty three subjects (14 males and 19 females, aged from 20 to 40 years) were involved. PT showed no significant differences at RT at 4 h, while significant differences after 8 h and 24 h at RT and after 4 h, 8 h and 24 h at refrigerator were observed. APTT showed no statistically significant differences at 2 h but showed significant differences at 6 h, 8 h at both RT and refrigerator. CONCLUSION: Samples for PT testing can be accepted only up to 4 h when kept at RT while the samples cannot be accepted when kept at refrigerator for 4 h and above. APTT samples can be accepted up to 2 h only at RT or refrigerator.