Lutein ameliorates high-fat diet-induced obesity, fatty liver, and glucose intolerance in C57BL/6J mice.

Obesity is a multi-factorial metabolic syndrome that increases the risk of cardiovascular diseases, diabetes, and cancer. We recently demonstrated the antiadipogenic efficacy of lutein using a 3 T3-L1 cell culture model. This study aimed to examine the antiobesity efficacy of lutein on high-fat (60% kcal fat) diet-induced C57BL/6J obese mice model. Lutein (300 and 500 µM), Orlistat (30 mg/kg body weight - positive control), and its combination (orlistat, 15 mg/kg body weight+lutein, 300 µM) were administered in high-fat diet (HFD)-fed mice every other day for 24 weeks. The effect on serum and hepatic lipid parameters was estimated using biochemical assay kits. The adipose tissue expression of adipocyte differentiation markers at gene and protein levels was analyzed by RT-PCR and western blotting, respectively. The results showed that lutein administration and drug significantly reduced epididymal and abdominal adipose tissue weights. Further, lutein reduced the serum cholesterol and LDL-C concentration compared to the HFD group. The HFD-induced elevation in the hepatic triglycerides and cholesterol levels were significantly blocked by lutein and its combination with the drug. Similarly, lutein and its drug combination efficiently lowered the HFD-mediated elevated blood glucose levels. Lutein downregulated the expression of CEBP-α, PPAR-γ, and FAS in the epididymal adipose tissue. Thus, supplementation of lutein may control diet-induced obesity and associated complications in the human population.

Lutein inhibits breast cancer cell growth by suppressing antioxidant and cell survival signals and induces apoptosis.

Reduced risk of breast cancer upon intake of lutein-rich food supplements creates an interest to investigate the molecular mechanism underlying the growth inhibitory potential of lutein in MCF-7 and MDA-MB-231 cells. Lutein purified from Spinacia oleracea was identified by high-performance liquid chromatography and liquid chromatography-mass spectrometry. The cell viability was measured by water-soluble tetrazolium-1 assay. The intracellular reactive oxygen species level was examined by 2',7'-dichlorofluorescein assay. The protein expression of the markers of antioxidant defense, cell survival, and apoptosis was analyzed by western blot analysis. The induction of apoptosis by lutein was measured by 4',6-diamidino-2-phenylindole staining and caspase-3 activity assay. The purified lutein inhibited the viability of MCF-7 and MDA-MB-231 cells. The growth inhibitory effect of lutein was associated with suppressed protein expression of superoxide dismutase-2 and heme oxygenase-1, and its transcription factor nuclear factor erythroid 2-related factor-2. Lutein treatment subsequently blocked the expression of intracellular cell survival proteins, phosphorylated protein kinase B, phosphorylated extracellular-regulated kinase 1/2, and nuclear factor-kB. Suppression of antioxidant defense and cell survival markers by lutein was further linked to apoptosis induction with elevated caspase-3 activity and downregulated expression of Bcl-2 and poly-ADP ribose polymerase. Our results emphasize a significant role of lutein as an effective inhibitor of human breast cancer cell growth that activates cell death partly through the modulation of antioxidant defense response-linked cell survival signaling markers.

Phenolic Extract of Seagrass, Halophila ovalis Activates Intrinsic Pathway of Apoptosis in Human Breast Cancer (MCF-7) Cells.

The marine ecosystem is considered as a treasure of numerous novel biologically active molecules. We investigated the anticancer potential of the phenolic extract of Halophila ovalis in breast cancer (MCF-7) cells and characterized the possible underlying molecular mechanism. The phenolic extract (5 µl) of H. ovalis effectively inhibited the growth of MCF-7 cells. The results of DAPI staining indicated that this phenolic extract potently induces apoptosis in MCF-7 cells which was observed by increased chromatin condensation in the treated cells. An increased expression of the active fragments of an executioner caspase, caspase 3 in phenolic extract-treated MCF-7 cells further confirms this apoptosis induction. In consequence, the loss of mitochondrial membrane potential was noticed in treated cells. The protein expression analyzes show decreased expression of the anti-apoptotic protein, Bcl-2, and DNA repair enzyme, PARP in treated cells indicating the probable molecular targets of apoptosis. Further, the phenolic extract of H. ovalis blocked the antioxidant defense system in MCF-7 cells by down-regulating the protein expression of a major transcription factor, Nrf-2 and regulatory antioxidant enzymes, SOD-2 and HO-1. These results show the presence of chemopreventive compound(s) in the phenolic extract, which offers a platform for future studies to identify the active principles.

An in vitro protocol to study the effect of hyperglycemia on intracellular redox signaling in human retinal pigment epithelial (ARPE-19) cells.

DMEM/F12 nutrient mixture, a recommended media for ARPE-19 culture, contains glucose concentration of 17.5 mM. But, several recent studies employed normal glucose media (5.5 mM) that was shown to affect the growth and function of ARPE-19 cells. Here, we set a protocol to study the effect of hyperglycemia on intracellular oxidative stress and redox status in ARPE-19 using DMEM/F12 as control. The WST-1 assay was performed to analyze the viability of ARPE-19 upon glucose treatment. The intracellular oxidative stress was measured by a dichlorofluorescein assay. The mitochondrial membrane potential (MMP) was monitored by using a JC-10 MMP assay kit. The expression of antioxidant marker proteins was analyzed by western blotting. Exogenous addition of glucose (7.5 and 12.5 mM) for 24 and 48 h did not change the viability and morphology of ARPE-19 cells. Hyperglycemia increased intracellular ROS level and decreased MMP in a dose-dependent manner. High-glucose treatment for 24 h down-regulated the protein expression of redox-specific transcription factors Nrf-2, XBP-1 and NF-κB, and subsequently decreased the expression of HO-1, catalase, and SOD-2. This study offers baseline information for the subsequent use of DMEM/F12 nutrient mixture to study glucose-mediated changes in intracellular oxidative stress and redox status of ARPE-19 without affecting its basic functions.

Effective inhibition of adipogenesis-mediated inflammation by a macular carotenoid, lutein in vitro.

An absolute interlinks between inflammation and obesity with scarce investigations on the role of lutein in inflammation-induced obesity motivated us to explore the protective mechanism of lutein on adipogenesis-mediated inflammation in vitro by culturing RAW264.7 macrophages in adipocyte conditioned medium. The RAW264 macrophage cells were cultured with adipocyte-conditioned media, and the potency of lutein on the expression of adipocyte inflammation-associated protein markers (IL-1ß, MCP-1, TNF-α, IL-6, NF-κB, and IKKα/ß) were analyzed by western blotting. The data revealed that lutein effectively reduces the protein levels of major inflammatory markers such as NF-κB, IL-1ß, MCP-1, and TNF-α in differentiated adipocytes. Interestingly, lutein hampered inflammation in the RAW264 cells that were cultured in adipocyte-conditioned media by lowering the protein expression of IL-1ß, MCP-1, and TNF-α. The blockage of inflammation by lutein in both differentiated adipocytes, and adipogenesis-induced macrophages is associated with suppression of IKK α/ß phosphorylation. These data suggest that lutein potentially alters adipocyte differentiation-mediated inflammation by regulating the NF-κB signaling pathway. Thus, lutein could be utilized as a potent nutraceutical agent in the management of obesity and associated inflammation. PRACTICAL APPLICATIONS: Lutein isolated from a dietary source exhibited an inhibitory effect in adipogenesis-induced inflammations. The findings of this study authenticate the diversified prospective of lutein in regulating obesity and other inflammation-related diseases. Thus, it is understood that continuous intake of lutein-rich food or dietary intervention of lutein may reduce the risk of developing obesity.

Inhibitory efficacy of lutein on adipogenesis is associated with blockage of early phase regulators of adipocyte differentiation.

A comprehensive molecular mechanistic role of lutein on adipogenesis is not well understood. The present study focused to evaluate the effect of lutein at the early and late phase of adipocyte differentiation in vitro using a 3T3-L1 cell model. The effect of purified carotenoid on the viability of normal and differentiated 3T3-L1 cells was analyzed by WST-1 assay. Oil Red O and Nile red staining were employed to observe lipid droplets in mature adipocytes. The effect of lutein on gene and protein expression of major transcription factors and adipogenic markers was analyzed by RT-PCR and western blotting, respectively. The role of lutein on mitotic clonal expansion was analyzed by flow cytometry. The results showed a significant reduction (p < 0.05) in the accumulation of lipid droplets in lutein-treated (5 µM) cells. Inhibition in lipid accumulation was associated with down-regulated expression of CEBP-α and PPAR-γ at gene and protein levels. Subsequently, lutein repressed gene expression of FAS, FABP4, and SCD1 in mature adipocytes. Interestingly, it blocks the protein expression of CEBP-α and PPAR-γ in the initial stages of adipocyte differentiation. This early-stage inhibition of adipocyte differentiation is linked with repressed phosphorylation AKT and ERK. Further, upregulated cyclin D and down-regulated CDK4 and CDK2 in lutein treated adipocytes enumerate its role in delaying the cell cycle progression at the G0/G1 phase. Our results emphasize that adipogenesis inhibitory efficacy of lutein is potentiated by halting early phase regulators of adipocyte differentiation, which strengthens the competency of lutein besides its inevitable presence in the human body.

Lactucaxanthin - a potential anti-diabetic carotenoid from lettuce (Lactuca sativa) inhibits α-amylase and α-glucosidase activity in vitro and in diabetic rats.

Intestinal and pancreatic α-amylase and α-glucosidase inhibitors offer an approach to lower the levels of post-prandial hyperglycemia through the control of dietary starch breakdown in digestion. This study hypothesized that lactucaxanthin (Lxn) in lettuce (Lactuca sativa) inhibits the activity of α-amylase and α-glucosidase. In this study, the interaction of Lxn with α-amylase and α-glucosidase in silico and its inhibitory effect on these enzymes were studied using in vitro and STZ-induced diabetic rat models. Lxn was isolated from lettuce with 96% purity confirmed by HPLC and LCMS. The in silico analysis showed that Lxn has a lower binding energy (-6.05 and -6.34 kcal mol-1) with α-amylase and α-glucosidase compared to their synthetic inhibitors, acarbose (-0.21 kcal mol-1) and miglitol (-2.78 kcal mol-1), respectively. In vitro α-amylase and α-glucosidase inhibition assays revealed that Lxn had IC50 values of 435.5 µg mL-1 and 1.84 mg mL-1, but acarbose has values of 2.5 and 16.19 µg mL-1. The in vivo results showed an increased activity for α-amylase and α-glucosidase in the intestine (4.7 and 1.30 fold, p < 0.05) and pancreas (1.3 and 1.48 fold, p < 0.05) of STZ induced diabetic rats compared to normal rats. Whereas the activity decreased (p < 0.05) in the Lxn fed diabetic rats, except for the intestinal α-glucosidase activity (1.69 ± 0.12 PNP per min per mg protein). This was confirmed by the low blood glucose level (239.4 ± 18.2 mg dL-1) in diabetic rats fed Lxn compared to the diabetic group (572.2 ± 30.5 mg dL-1, p < 0.05). Lxn significantly inhibited (p < 0.05) the activity of α-amylase and α-glucosidase and could be of medical and nutritional relevance in the treatment of diabetes.