Zinc intake, zinc status and expression of zinc transporter genes in younger and older Saudi adults

PhD ThesisBackground: During ageing, there are changes in many physiological systems including the gastrointestinal tract which may result in decreased absorption of micronutrients such as zinc. Decreased dietary absorption efficiency and/or inadequate zinc intake are contributing factors to the decline in plasma zinc concentration that has been reported in older adults. This decline in plasma zinc concentration may have important implications for health, e.g. through reduced immune function. To date, there is limited information on the relationships between zinc intake and plasma zinc concentration during ageing among adults in Saudi Arabia or on the molecular mechanisms responsible for age-related effects. Thus, the aims of this study were to test the hypothesis that changes in expression of zinc transporters including ZnT1, ZnT5 and ZIP4 are associated with changes in zinc status in young and older Saudi adults. Methods: Two hundred and two young and older Saudi adults of both sexes were recruited from Jeddah city in the western region of Saudi Arabia. Intakes of zinc and of zinc absorption modifiers in diet were assessed using an FFQ (designed and validated for this purpose) while plasma zinc concentration was determined by ICP-OES. RNA was extracted from white blood cells and qPCR was used to quantitatively measure expression of the ZnT1, ZnT5 and ZIP4 genes at the mRNA level. Results: Intakes of zinc and of protein were significantly higher in males than in females (P<0.05). A substantial proportion of the participants appeared to have inadequate zinc intake with the risk of inadequacy being greatest for young males (15%) and young females (4%). Plasma zinc concentrations in older adults were significantly higher than in young adults (P<0.05). Older males had significantly higher ZIP4 expression compared with young adults (P<0.05). Although older adults had apparently higher ZnT1 and ZnT5 expression than young adults, these differences were statistically non-significant. Several age and sex-specific correlations between zinc intake, zinc status and expression of zinc transporters were observed. Young and older adults who had apple-shaped fat distribution pattern, had significantly higher expression of ZnT1, ZnT5 and ZIP4 and plasma zinc status compared with participants who had pear-shaped fat distribution pattern (P<0.05). Discussion: Despite the changes in food availability in Saudi Arabia over recent decades, the prevalence of low zinc intake remains substantial. Contrary to reports from elsewhere, the prevalence of low plasma II zinc concentrations was greater in younger than in older Saudi adults. This study also identified relationships between expression of zinc transporters and zinc intake, zinc status and abdominal fatness in Saudi adults

Cell Growth Inhibition, DNA Fragmentation and Apoptosis-Inducing Properties of Household-Processed Leaves and Seeds of Fenugreek (<i>Trigonella Foenum-Graecum</i> Linn.) against HepG2, HCT-116, and MCF-7 Cancerous Cell Lines

Household processing of fenugreek seeds and leaves, including soaking, germination, and boiling of the seeds, and air-drying of the leaves, has improved the levels of human consumption of the bitter seeds and increased the shelf life of fresh leaves, respectively. The potential anticancer activity of either unprocessed or processed fenugreek seeds or leaves and the relative expression of pro-apoptotic and anti-apoptotic genes of the studied cancerous cell lines exposed to IC50 crude extracts was investigated to observe the apoptotic-inducing property of this plant as an anticancer agent. The protein expression of IKK-α and IKK-β, as inhibitors of NF-KB which exhibit a critical function in the regulation of genes involved in chronic inflammatory disorders, were studied in the tested cancerous cell lines. In this study, the anticancer activity of household-processed fenugreek leaves and seeds against HepG2, HCT-116, MCF-7, and VERO cell lines was measured using an MTT assay. DNA fragmentation of both HepG2 and MCF-7 was investigated by using gel electrophoresis. RT-PCR was used to evaluate the relative expression of each p53, caspase-3, Bax, and Bcl-2 genes, whereas ELISA assay determined the expression of caspase-3, TNF-α, and 8-OHDG genes. Western blotting analyzed the protein-expressing levels of IKK-α and IKK-β proteins in each studied cell line. Data showed that at 500 µg mL−1, ADFL had the highest cytotoxicity against the HepG2 and HCT-116 cell lines. Although, each UFS and GFS sample had a more inhibitory effect on MCF-7 cells than ADFL. Gel electrophoresis demonstrated that the IC50 of each ADFL, UFS, and GFS sample induced DNA fragmentation in HepG2 and MCF-7, contrary to untreated cell lines. Gene expression using RT-PCR showed that IC50 doses of each sample induced apoptosis through the up-regulation of the p53, caspase-3, and Bax genes and the down-regulation of the Bcl-2 gene in each studied cell line. The relative expression of TNF-α, 8-OHDG, and caspase-3 genes of each HepG2 and MCF-7 cell line using ELISA assays demonstrated that ADFL, UFS, and GFS samples reduced the expression of TNF-α and 8-OHDG genes but increased the expression of the caspase-3 gene. Protein-expressing levels of IKK-α and IKK-β proteins in each studied cell line, determined using Western blotting, indicated that household treatments decreased IKK-α expression compared to the UFS sample. Moreover, the ADFL and SFS samples had the most activity in the IKK-β expression levels. Among all studied samples, air-dried fenugreek leaves and unprocessed and germinated fenugreek seeds had the most anti-proliferative and apoptotic-inducing properties against human HepG2, MCF-7, and HCT-116 cell lines, as compared to the VERO cell line. So, these crude extracts can be used in the future for developing new effective natural drugs for the treatment of hepatocellular, breast, and colon carcinomas

Cell Growth Inhibition, DNA Fragmentation and Apoptosis-Inducing Properties of Household-Processed Leaves and Seeds of Fenugreek (Trigonella Foenum-Graecum Linn.) against HepG2, HCT-116, and MCF-7 Cancerous Cell Lines

Household processing of fenugreek seeds and leaves, including soaking, germination, and boiling of the seeds, and air-drying of the leaves, has improved the levels of human consumption of the bitter seeds and increased the shelf life of fresh leaves, respectively. The potential anticancer activity of either unprocessed or processed fenugreek seeds or leaves and the relative expression of pro-apoptotic and anti-apoptotic genes of the studied cancerous cell lines exposed to IC50 crude extracts was investigated to observe the apoptotic-inducing property of this plant as an anticancer agent. The protein expression of IKK-&alpha; and IKK-&beta;, as inhibitors of NF-KB which exhibit a critical function in the regulation of genes involved in chronic inflammatory disorders, were studied in the tested cancerous cell lines. In this study, the anticancer activity of household-processed fenugreek leaves and seeds against HepG2, HCT-116, MCF-7, and VERO cell lines was measured using an MTT assay. DNA fragmentation of both HepG2 and MCF-7 was investigated by using gel electrophoresis. RT-PCR was used to evaluate the relative expression of each p53, caspase-3, Bax, and Bcl-2 genes, whereas ELISA assay determined the expression of caspase-3, TNF-&alpha;, and 8-OHDG genes. Western blotting analyzed the protein-expressing levels of IKK-&alpha; and IKK-&beta; proteins in each studied cell line. Data showed that at 500 &micro;g mL&minus;1, ADFL had the highest cytotoxicity against the HepG2 and HCT-116 cell lines. Although, each UFS and GFS sample had a more inhibitory effect on MCF-7 cells than ADFL. Gel electrophoresis demonstrated that the IC50 of each ADFL, UFS, and GFS sample induced DNA fragmentation in HepG2 and MCF-7, contrary to untreated cell lines. Gene expression using RT-PCR showed that IC50 doses of each sample induced apoptosis through the up-regulation of the p53, caspase-3, and Bax genes and the down-regulation of the Bcl-2 gene in each studied cell line. The relative expression of TNF-&alpha;, 8-OHDG, and caspase-3 genes of each HepG2 and MCF-7 cell line using ELISA assays demonstrated that ADFL, UFS, and GFS samples reduced the expression of TNF-&alpha; and 8-OHDG genes but increased the expression of the caspase-3 gene. Protein-expressing levels of IKK-&alpha; and IKK-&beta; proteins in each studied cell line, determined using Western blotting, indicated that household treatments decreased IKK-&alpha; expression compared to the UFS sample. Moreover, the ADFL and SFS samples had the most activity in the IKK-&beta; expression levels. Among all studied samples, air-dried fenugreek leaves and unprocessed and germinated fenugreek seeds had the most anti-proliferative and apoptotic-inducing properties against human HepG2, MCF-7, and HCT-116 cell lines, as compared to the VERO cell line. So, these crude extracts can be used in the future for developing new effective natural drugs for the treatment of hepatocellular, breast, and colon carcinomas