Stingless Bee (Heterotrigona Itama) Honey and Its Phenolic-Rich Extract Ameliorate Oxidant-Antioxidant Balance via KEAP1-NRF2 Signalling Pathway.

Diabetes is associated with an imbalance between oxidants and antioxidants, leading to oxidative stress. This imbalance contributes to the development and progression of diabetic complications. Similarly, renal and liver diseases are characterised by oxidative stress, where an excess of oxidants overwhelms the antioxidant defense mechanisms, causing tissue damage and dysfunction. Restoring the oxidant-antioxidant balance is essential for mitigating oxidative stress-related damage under these conditions. In this current study, the efficacy of stingless bee honey (SBH) and its phenolic-rich extract (PRE) in controlling the oxidant-antioxidant balance in high-fat diet- and streptozotocin/nicotinamide-induced diabetic rats was investigated. The administration of SBH and PRE improved systemic antioxidant defense and oxidative stress-related measures without compromising liver and renal functioning. Analyses of the liver, skeletal muscle and adipose tissues revealed differences in their capacities to scavenge free radicals and halt lipid peroxidation. Transcriptional alterations hypothesised tissue-specific control of KEAP1-NRF2 signalling by upregulation of Nrf2, Ho1 and Sod1 in a tissue-specific manner. In addition, hepatic translational studies demonstrated the stimulation of downstream antioxidant-related protein with upregulated expression of SOD-1 and HOD-1 protein. Overall, the results indicated that PRE and SBH can be exploited to restore the oxidant-antioxidant imbalance generated by diabetes via regulating the KEAP1-NRF2 signalling pathway.

In Vivo Toxicity Assessment of the Probiotic Bacillus amyloliquefaciens HTI-19 Isolated from Stingless Bee (Heterotrigona itama) Honey.

This study evaluated the acute and sub-acute toxicity of B. amyloliquefaciens HTI-19 (isolated from stingless bee honey) in female Sprague Dawley rats. In an acute toxicity study, the rats received a low dosage (1 × 109 CFU·mL-1), medium dosage (3 × 109 CFU·mL-1), or high dosage (1 × 1010 CFU·mL-1) of B. amyloliquefaciens HTI-19 daily orally by syringe-feeding for 14 days. For the subacute toxicity study, rats received a low dosage (1 × 109 CFU·mL-1) or a high dosage (1 × 1010 CFU·mL-1) for 28 days. The probiotic feeding in acute and sub-acute toxicity studies showed no mortality or significant abnormalities in rats throughout the experimental period. In week 2 of the acute study, the body weight of the rats showed a significant increase (p < 0.05) compared to the control. By gross and microscopic examination of organs, no evidently significant changes were observed in the morphology of organs. Serum biochemical tests and blood hematology tests also revealed no treatment-related changes. Overall, these data indicated that oral administration of B. amyloliquefaciens HTI-19 up to 1 × 109 CFU·mL-1 for 28 days can be considered safe.

Essential oils and plant extracts for tropical fruits protection: From farm to table.

The tropical fruit industry in Malaysia makes up a large proportion of the agriculture sector, contributing to the local economy. Due to their high sugar and water content, tropical fruits are prone to pathogenic infections, providing optimal microorganism growth conditions. As one of the largest exporters of these fruits globally, following other Southeast Asian countries such as Thailand, Indonesia and the Philippines, the quality control of exported goods is of great interest to farmers and entrepreneurs. Traditional methods of managing diseases in fruits depend on chemical pesticides, which have attracted much negative perception due to their questionable safety. Therefore, the use of natural products as organic pesticides has been considered a generally safer alternative. The extracts of aromatic plants, known as essential oils or plant extracts, have garnered much interest, especially in Asian regions, due to their historical use in traditional medicine. In addition, the presence of antimicrobial compounds further advocates the assessment of these extracts for use in crop disease prevention and control. Herein, we reviewed the current developments and understanding of the use of essential oils and plant extracts in crop disease management, mainly focusing on tropical fruits. Studies reviewed suggest that essential oils and plant extracts can be effective at preventing fungal and bacterial infections, as well as controlling crop disease progression at the pre and postharvest stages of the tropical fruit supply chain. Positive results from edible coatings and as juice preservatives formulated with essential oils and plant extracts also point towards the potential for commercial use in the industry as more chemically safe and environmentally friendly biopesticides.

Modulation of High-Fat Diet-Induced Brain Oxidative Stress by Ferulate-Rich Germinated Brown Rice Ethyl Acetate Extract.

The oxidative stress resulting from the production of reactive oxygen species plays a vital role in inflammatory processes and is associated with neurodegenerative changes. In view of the ability of germinated brown rice (GBR) to improve learning and memory, this present study aimed to investigate the mechanistic basis of GBR's neuroprotection in a high-fat diet (HFD)-induced oxidative changes in adult Sprague-Dawley rats. Ferulate-rich GBR ethyl acetate extract (GBR-EA; 100 mg/kg and 200 mg/kg body weight) was supplemented orally for the last 3 months of 6 months HFD feeding during the study. GBR-EA supplementation was found to improve lipid profile and serum antioxidant status, when compared to the HFD group. Elevated mRNA expressions of SOD1, SOD2, SOD3, Catalase, and GPX were demonstrated in the frontal cortex and hippocampus of GBR-EA treated animals. The pro-inflammatory changes induced by HFD in the hippocampus were attenuated by GBR-EA through the downregulation of CRP and TNF- α and upregulation of PPAR-γ. GBR also reduced the hippocampal mRNA expression and enzyme level of acetylcholinesterase. In conclusion, this study proposed the possible transcriptomic regulation of antioxidant and inflammation in neurodegenerative processes resulting from high cholesterol consumption, with an emphasis on GBR's potential to ameliorate such changes.

Conditioned media of pancreatic cancer cells and pancreatic stellate cells induce myeloid-derived suppressor cells differentiation and lymphocytes suppression.

As pancreatic cancer cells (PCCs) and pancreatic stellate cells (PSCs) are the two major cell types that comprise the immunosuppressive tumor microenvironment of pancreatic cancer, we aimed to investigate the role of conditioned medium derived from PCCs and PSCs co-culture on the viability of lymphocytes. The conditioned medium (CM) collected from PCCs and/or PSCs was used to treat peripheral blood mononuclear cells (PBMCs) to determine CM ability in reducing lymphocytes population. A proteomic analysis has been done on the CM to investigate the differentially expressed protein (DEP) expressed by two PCC lines established from different stages of tumor. Subsequently, we investigated if the reduction of lymphocytes was directly caused by CM or indirectly via CM-induced MDSCs. This was achieved by isolating lymphocyte subtypes and treating them with CM and CM-induced MDSCs. Both PCCs and PSCs were important in suppressing lymphocytes, and the PCCs derived from a metastatic tumor appeared to have a stronger suppressive effect than the PCCs derived from a primary tumor. According to the proteomic profiles of CM, 416 secreted proteins were detected, and 13 DEPs were identified between PANC10.05 and SW1990. However, CM was found unable to reduce lymphocytes viability through a direct pathway. In contrast, CM that contains proteins secreted by PCC and/or PSC appear immunogenic as they increase the viability of lymphocytes subtypes. Lymphocyte subtype treated with CM-induced MDSCs showed reduced viability in T helper 1 (Th1), T helper 2 (Th2), and T regulatory (Treg) cells, but not in CD8+ T cells, and B cells. As a conclusion, the interplay between PCCs and PSCs is important as their co-culture displays a different trend in lymphocytes suppression, hence, their co-culture should be included in future studies to better mimic the tumor microenvironment.

Theragnostic Applications of Mammal and Plant-Derived Extracellular Vesicles: Latest Findings, Current Technologies, and Prospects.

The way cells communicate is not fully understood. However, it is well-known that extracellular vesicles (EVs) are involved. Researchers initially thought that EVs were used by cells to remove cellular waste. It is now clear that EVs function as signaling molecules released by cells to communicate with one another, carrying a cargo representing the mother cell. Furthermore, these EVs can be found in all biological fluids, making them the perfect non-invasive diagnostic tool, as their cargo causes functional changes in the cells upon receiving, unlike synthetic drug carriers. EVs last longer in circulation and instigate minor immune responses, making them the perfect drug carrier. This review sheds light on the latest development in EVs isolation, characterization and, application as therapeutic cargo, novel drug loading techniques, and diagnostic tools. We also address the advancement in plant-derived EVs, their characteristics, and applications; since plant-derived EVs only recently gained focus, we listed the latest findings. Although there is much more to learn about, EV is a wide field of research; what scientists have discovered so far is fascinating. This paper is suitable for those new to the field seeking to understand EVs and those already familiar with it but wanting to review the latest findings.

Increased fucoxanthin in Chaetoceros calcitrans extract exacerbates apoptosis in liver cancer cells via multiple targeted cellular pathways.

In this study, anti-proliferative effects of C. calcitrans extract and its fucoxanthin rich fraction (FxRF) were assessed on human liver HepG2 cancer cell line. Efficacy from each extract was determined by cytotoxicity assay, morphological observation, and cell cycle analysis. Mechanisms of action observed were evaluated using multiplex gene expression analysis. Results showed that CME and FxRF induced cytotoxicity to HepG2 cells in a dose and time-dependent manner. FxRF (IC50: 18.89 µg.mL-1) was found to be significantly more potent than CME (IC50: 87.5 µg.mL-1) (p < 0.05). Gene expression studies revealed that anti-proliferative effects in treated cells by C. calcitrans extracts were mediated partly through the modulation of numerous genes involved in cell signaling (AKT1, ERK1/2, JNK), apoptosis (BAX, BID, Bcl-2, APAF, CYCS) and oxidative stress (SOD1, SOD2, CAT). Overall, C. calcitrans extracts demonstrated effective intervention against HepG2 cancer cells where enhanced apoptotic activities were observed with increased fucoxanthin content.

Probiotic Properties of Bacillus Strains Isolated from Stingless Bee (Heterotrigona itama) Honey Collected across Malaysia.

This study aimed to isolate, identify, and evaluate the probiotic properties of Bacillus species from honey of the stingless bee Heterotrigona itama. Bacillus spp. were isolated from five different H. itama meliponicultures, and the isolates were characterized through Gram-staining and a catalase test. Tolerance to acidic conditions and bile salt (0.3%), hydrophobicity, and autoaggregation tests were performed to assess the probiotic properties of the selected isolates, B. amyloliquefaciens HTI-19 and B. subtilis HTI-23. Both Bacillus isolates exhibited excellent antimicrobial activity against both Gram-positive and Gram-negative bacteria and possessed significantly high survival rates in 0.3% bile solution for 3 h. Their survival rates in acidic conditions were also comparable to a commercial probiotic strain, Lactobacillus rhamnosus GG. Interestingly, the hydrophobicity and autoaggregation percentage showed no significant difference from L. rhamnosus GG, a commercial probiotic strain. The results from this study suggest that B. amyloliquefaciens HTI-19 and B. subtilis HTI-23 isolated from stingless bee honey have considerably good probiotic properties. Therefore, more studies should be done to investigate the effects of these bacteria cultures on gastrointestinal health.

Therapeutic Properties of Stingless Bee Honey in Comparison with European Bee Honey.

Both honeybees (Apis spp.) and stingless bees (Trigona spp.) produce honeys with high nutritional and therapeutics value. Until recently, the information regarding potential health benefits of stingless bee honey (SBH) in medical databases is still scarce as compared to the common European bee honey (EBH) which is well known for their properties as therapeutic agents. Although there have been very few reports on SBH, empirically these products would have similar therapeutic quality as the EBH. In addition, due to the structure of the nest, few studies reported that the antimicrobial activity of SBH is a little bit stronger than EBH. Therefore, the composition of both the types of honey as well as the traditional uses and clinical applications were compared. The results of various studies on EBH and SBH from tissue culture research to randomised control clinical trials were collated in this review. Interestingly, there are many therapeutic properties that are unique to SBH. Therefore, SBH has a great potential to be developed for modern medicinal uses.

Thymoquinone-rich fraction nanoemulsion (TQRFNE) decreases Aß40 and Aß42 levels by modulating APP processing, up-regulating IDE and LRP1, and down-regulating BACE1 and RAGE in response to high fat/cholesterol diet-induced rats.

Though the causes of Alzheimer's disease (AD) are yet to be understood, much evidence has suggested that excessive amyloid-ß (Aß) accumulation due to abnormal amyloid-ß precursor protein (APP) processing and Aß metabolism are crucial processes towards AD pathogenesis. Hence, approaches aiming at APP processing and Aß metabolism are currently being actively pursued for the management of AD. Studies suggest that high cholesterol and a high fat diet have harmful effects on cognitive function and may instigate the commencement of AD pathogenesis. Despite the neuropharmacological attributes of black cumin seed (Nigella sativa) extracts and its main active compound, thymoquinone (TQ), limited records are available in relation to AD research. Nanoemulsion (NE) is exploited as drug delivery systems due to their capacity of solubilising non-polar active compounds and is widely examined for brain targeting. Herewith, the effects of thymoquinone-rich fraction nanoemulsion (TQRFNE), thymoquinone nanoemulsion (TQNE) and their counterparts' conventional emulsion in response to high fat/cholesterol diet (HFCD)-induced rats were investigated. Particularly, the Aß generation; APP processing, ß-secretase 1 (BACE1), γ-secretases of presenilin 1 (PSEN1) and presenilin 2 (PSEN2), Aß degradation; insulin degrading enzyme (IDE), Aß transportation; low density lipoprotein receptor-related protein 1 (LRP1) and receptor for advanced glycation end products (RAGE) were measured in brain tissues. TQRFNE reduced the brain Aß fragment length 1-40 and 1-42 (Aß40 and Aß42) levels, which would attenuate the AD pathogenesis. This reduction could be due to the modulation of ß- and γ-secretase enzyme activity, and the Aß degradation and transportation in/out of the brain. The findings show the mechanistic actions of TQRFNE in response to high fat and high cholesterol diet associated to Aß generation, degradation and transportation in the rat's brain tissue.

Beneficial effects of TQRF and TQ nano- and conventional emulsions on memory deficit, lipid peroxidation, total antioxidant status, antioxidants genes expression and soluble Aß levels in high fat-cholesterol diet-induced rats.

The study determined the effect of thymoquinone rich fraction (TQRF) and thymoquinone (TQ) in the forms of nano- and conventional emulsions on learning and memory, lipid peroxidation, total antioxidant status, antioxidants genes expression and soluble ß-amyloid (Aß) levels in rats fed with a high fat-cholesterol diet (HFCD). The TQRF was extracted from Nigella sativa seeds using a supercritical fluid extraction system and prepared into nanoemulsion, which later named as TQRF nanoemulsion (TQRFNE). Meanwhile, TQ was acquired commercially and prepared into thymoquinone nanoemulsion (TQNE). The TQRF and TQ conventional emulsions (CE), named as TQRFCE and TQCE, respectively were studied for comparison. Statin (simvastatin) and non-statin (probucol) cholesterol-lowering agents, and a mild-to-severe Alzheimer's disease drug (donepezil) were served as control drugs. The Sprague Dawley rats were fed with HFCD for 6 months, and treated with the intervention groups via oral gavage daily for the last 3 months. As a result, HFCD-fed rats exhibited hypercholesterolaemia, accompanied by memory deficit, increment of lipid peroxidation and soluble Aß levels, decrement of total antioxidant status and down-regulation of antioxidants genes expression levels. TQRFNE demonstrated comparable effects to the other intervention groups and control drugs in serum biomarkers as well as in the learning and memory test. Somehow, TQRFNE was more prominent than those intervention groups and control drugs in brain biomarkers concomitant to gene and protein expression levels. Supplementation of TQRFNE into an HFCD thus could ameliorate memory deficit, lipid peroxidation and soluble Aß levels as well as improving the total antioxidant status and antioxidants genes expression levels.

Dillenia suffruticosa dichloromethane root extract induced apoptosis towards MDA-MB-231 triple-negative breast cancer cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Dillenia suffruticosa is traditionally used for treatment of cancerous growth including breast cancer in Malaysia. AIM OF THE STUDY: Dillenia suffruticosa is a well-known medicinal plant in Malaysia for the treatment of cancer. Nevertheless, no study has been reported the cytotoxicity of this plant towards MDA-MB-231 triple-negative breast cancer cells. The present study was designed to investigate the mode of cell death and signalling pathways of MDA-MB-231 cells treated with dichloromethane Dillenia suffruticosa root extract (DCM-DS). METHODS: Extraction of Dillenia suffruticosa root was performed by the use of sequential solvent procedure. The cytotoxicity of DCM-DS was determined by using MTT assay. The mode of cell death was evaluated by using an inverted light microscope and flow cytometry analysis using Annexin-V/PI. Cell cycle analysis and measurement of reactive oxygen species level were performed by using flow cytometry. The cells were treated with DCM-DS and antioxidants α-tocopherol or ascorbic acid to evaluate the involvement of ROS in the cytotoxicity of DCM-DS. Effect of DCM-DS on the expression of antioxidant, apoptotic, growth, survival genes and proteins were analysed by using GeXP-based multiplex system and Western blot, respectively. The cytotoxicity of compounds isolated from DCM-DS was evaluated towards MDA-MB-231 cells using MTT assay. RESULTS: DCM-DS induced apoptosis, G2/M phase cell cycle arrest and oxidative stress in MDA-MB-231 cells. The induction of apoptosis in MDA-MB-231 cells by DCM-DS is possibly due to the activation of pro-apoptotic JNK1 and down-regulation of anti-apoptotic ERK1, which in turn down-regulates anti-apoptotic BCL-2 to increase the BAX/BCL-2 ratio to initiate the mitochondrial apoptotic pathway. The cell cycle arrest in DCM-DS-treated MDA-MB-231 cells is possibly via p53-independent but p21-dependent pathway. A total of 3 triterpene compounds were isolated from DCM-DS. Betulinic acid appears to be the most major and most cytotoxic compound in DCM-DS. CONCLUSION: The data suggest the potential application of DCM-DS in the treatment of triple-negative breast cancer.

Modulation of Hydrogen Peroxide-Induced Oxidative Stress in Human Neuronal Cells by Thymoquinone-Rich Fraction and Thymoquinone via Transcriptomic Regulation of Antioxidant and Apoptotic Signaling Genes.

Nigella sativa Linn. (N. sativa) and its bioactive constituent Thymoquinone (TQ) have demonstrated numerous pharmacological attributes. In the present study, the neuroprotective properties of Thymoquinone-rich fraction (TQRF) and TQ against hydrogen peroxide- (H2O2-) induced neurotoxicity in differentiated human SH-SY5Y cells were investigated. TQRF was extracted using supercritical fluid extraction while TQ was acquired commercially, and their effects on H2O2 were evaluated using cell viability assay, reactive oxygen species (ROS) assay, morphological observation, and multiplex gene expression. Both TQRF and TQ protected the cells against H2O2 by preserving the mitochondrial metabolic enzymes, reducing intracellular ROS levels, preserving morphological architecture, and modulating the expression of genes related to antioxidants (SOD1, SOD2, and catalase) and signaling genes (p53, AKT1, ERK1/2, p38 MAPK, JNK, and NF-κß). In conclusion, the enhanced efficacy of TQRF over TQ was likely due to the synergism of multiple constituents in TQRF. The efficacy of TQRF was better than that of TQ alone when equal concentrations of TQ in TQRF were compared. In addition, TQRF also showed comparable effects to TQ when the same concentrations were tested. These findings provide further support for the use of TQRF as an alternative to combat oxidative stress insults in neurodegenerative diseases.

N-Acetylneuraminic acid attenuates hypercoagulation on high fat diet-induced hyperlipidemic rats.

BACKGROUND AND OBJECTIVE: N-Acetylneuraminic acid (Neu5Ac), a type of sialic acid, has close links with cholesterol metabolism and is often used as a biomarker in evaluating the risk of cardiovascular diseases. However, most studies on the health implications of Neu5Ac have focused on its effects on the nervous system, while its effects on cardiovascular risk factors have largely been unreported. Thus, the effects of Neu5Ac on coagulation status in high fat diet (HFD)-induced hyperlipidemic rats were evaluated in this study. METHODS: Sprague Dawley male rats were divided into five different groups and fed with HFD alone, HFD low-dose Neu5Ac, HFD high-dose Neu5Ac, HFD simvastatin (10 mg/kg day), and normal pellet alone. Food was given ad libitum while body weight of rats was measured weekly. After 12 weeks of intervention, rats were sacrificed and serum and tissue samples were collected for biochemistry and gene expression analysis, respectively. RESULTS: The results showed that Neu5Ac could improve lipid metabolism and hyperlipidemia-associated coagulation. Neu5Ac exerted comparable or sometimes better physiological effects than simvastatin, at biochemical and gene expression levels. CONCLUSIONS: The data indicated that Neu5Ac prevented HFD-induced hyperlipidemia and associated hypercoagulation in rats through regulation of lipid-related and coagulation-related genes and, by extension, induced metabolite and protein changes. The implications of the present findings are that Neu5Ac may be used to prevent coagulation-related cardiovascular events in hyperlipidemic conditions. These findings are worth studying further.

N-Acetylneuraminic Acid Supplementation Prevents High Fat Diet-Induced Insulin Resistance in Rats through Transcriptional and Nontranscriptional Mechanisms.

N-Acetylneuraminic acid (Neu5Ac) is a biomarker of cardiometabolic diseases. In the present study, we tested the hypothesis that dietary Neu5Ac may improve cardiometabolic indices. A high fat diet (HFD) + Neu5Ac (50 or 400 mg/kg BW/day) was fed to rats and compared with HFD + simvastatin (10 mg/kg BW/day) or HFD alone for 12 weeks. Weights and serum biochemicals (lipid profile, oral glucose tolerance test, leptin, adiponectin, and insulin) were measured, and mRNA levels of insulin signaling genes were determined. The results indicated that low and high doses of sialic acid (SA) improved metabolic indices, although only the oral glucose tolerance test, serum triglycerides, leptin, and adiponectin were significantly better than those in the HFD and HFD + simvastatin groups (P < 0.05). Furthermore, the results showed that only high-dose SA significantly affected the transcription of hepatic and adipose tissue insulin signaling genes. The data suggested that SA prevented HFD-induced insulin resistance in rats after 12 weeks of administration through nontranscriptionally mediated biochemical changes that may have differentially sialylated glycoprotein structures at a low dose. At higher doses, SA induced transcriptional regulation of insulin signaling genes. These effects suggest that low and high doses of SA may produce similar metabolic outcomes in relation to insulin sensitivity through multiple mechanisms. These findings are worth studying further.

High fat diet-induced inflammation and oxidative stress are attenuated by N-acetylneuraminic acid in rats.

BACKGROUND: Serum sialic acid levels are positively correlated with coronary artery disease and inflammation. Although sialic acid is a non-specific marker, it is considered sensitive likely due to its influence in sialylation of glycoprotein structures all over the body. OBJECTIVES: We hypothesized that dietary supplementation with N-acetylneuraminic acid (Neu5Ac), a type of sialic acid, will have profound effects on high fat diet- (HFD-) induced inflammation and oxidative stress in view of the widespread incorporation of sialic acid into glycoprotein structures in the body. METHODS: HFD-fed rats with or without simvastatin or Neu5Ac (50 and 400 mg/kg/day) were followed up for 12 weeks. Lipid profiles, and markers of inflammation (C-reactive protein, interleukin-6, and tumor necrosis factor alpha), insulin resistance (serum insulin and adiponectin, oral glucose tolerance test and homeostatic model of insulin resistance) and oxidative stress (total antioxidant status and thiobarbituric acid reactive species) in the serum and liver were determined, while mRNA levels of hepatic antioxidant and inflammation genes were also quantified. Serum levels of alanine transaminase, aspartate transaminase, alkaline phosphatase, urea, creatinine and uric acid were also assessed. RESULTS: HFD feeding caused hyperlipidemia and insulin resistance, and worsened liver and kidney functions. HFD feeding also potentiated inflammation and oxidative stress, partly through modulation of hepatic gene expression, while Neu5Ac especially at higher doses and simvastatin attenuated HFD-induced changes, although Neu5Ac showed better outcomes. CONCLUSIONS: Based on the present results, we surmised that Neu5Ac can prevent HFD-induced inflammation and oxidative stress, and may in fact be useful in the prevention of hyperlipidemia-associated inflammation and oxidative stress. However, the translational implications of these findings can only be determined after long-term effects are established. Hence, the use of Neu5Ac on obesity-related diseases requires additional attention.

Edible bird's nest attenuates procoagulation effects of high-fat diet in rats.

Edible bird's nest (EBN) is popular in Asia, and has long been used traditionally as a supplement. There are, however, limited evidence-based studies on its efficacy. EBN has been reported to improve dyslipidemia, which is closely linked to hypercoagulation states. In the present study, the effects of EBN on high-fat diet- (HFD-) induced coagulation in rats were evaluated. Rats were fed for 12 weeks with HFD alone or in combination with simvastatin or EBN. Food intake was estimated, and weight measurements were made during the experimental period. After sacrifice, serum oxidized low-density lipoprotein (oxLDL), adiponectin, leptin, von willibrand factor, prostacyclin, thromboxane and lipid profile, and whole blood coagulation indices (bleeding time, prothrombin time, activated partial thromboplastin time, red blood count count, and platelet count) were estimated. Furthermore, hepatic expression of coagulation-related genes was evaluated using multiplex polymerase chain reaction. The results indicated that EBN could attenuate HFD-induced hypercholesterolemia and coagulation similar to simvastatin, partly through transcriptional regulation of coagulation-related genes. The results suggested that EBN has the potential for lowering the risk of cardiovascular disease-related hypercoagulation due to hypercholesterolemia.

Edible Bird's Nest Prevents High Fat Diet-Induced Insulin Resistance in Rats.

Edible bird's nest (EBN) is used traditionally in many parts of Asia to improve wellbeing, but there are limited studies on its efficacy. We explored the potential use of EBN for prevention of high fat diet- (HFD-) induced insulin resistance in rats. HFD was given to rats with or without simvastatin or EBN for 12 weeks. During the intervention period, weight measurements were recorded weekly. Blood samples were collected at the end of the intervention and oral glucose tolerance test conducted, after which the rats were sacrificed and their liver and adipose tissues collected for further studies. Serum adiponectin, leptin, F2-isoprostane, insulin, and lipid profile were estimated, and homeostatic model assessment of insulin resistance computed. Effects of the different interventions on transcriptional regulation of insulin signaling genes were also evaluated. The results showed that HFD worsened metabolic indices and induced insulin resistance partly through transcriptional regulation of the insulin signaling genes. Additionally, simvastatin was able to prevent hypercholesterolemia but promoted insulin resistance similar to HFD. EBN, on the other hand, prevented the worsening of metabolic indices and transcriptional changes in insulin signaling genes due to HFD. The results suggest that EBN may be used as functional food to prevent insulin resistance.

Induction of Apoptosis in MCF-7 Cells via Oxidative Stress Generation, Mitochondria-Dependent and Caspase-Independent Pathway by Ethyl Acetate Extract of Dillenia suffruticosa and Its Chemical Profile.

Dillenia suffruticosa, which is locally known as Simpoh air, has been traditionally used to treat cancerous growth. The ethyl acetate extract of D. suffruticosa (EADs) has been shown to induce apoptosis in MCF-7 breast cancer cells in our previous study. The present study aimed to elucidate the molecular mechanisms involved in EADs-induced apoptosis and to identify the major compounds in the extract. EADs was found to promote oxidative stress in MCF-7 cells that led to cell death because the pre-treatment with antioxidants α-tocopherol and ascorbic acid significantly reduced the cytotoxicity of the extract (P<0.05). DCFH-DA assay revealed that treatment with EADs attenuated the generation of intracellular ROS. Apoptosis induced by EADs was not inhibited by the use of caspase-inhibitor Z-VAD-FMK, suggesting that the cell death is caspase-independent. The use of JC-1 dye reflected that EADs caused disruption in the mitochondrial membrane potential. The related molecular pathways involved in EADs-induced apoptosis were determined by GeXP multiplex system and Western blot analysis. EADs is postulated to induce cell cycle arrest that is p53- and p21-dependent based on the upregulated expression of p53 and p21 (P<0.05). The expression of Bax was upregulated with downregulation of Bcl-2 following treatment with EADs. The elevated Bax/Bcl-2 ratio and the depolarization of mitochondrial membrane potential suggest that EADs-induced apoptosis is mitochondria-dependent. The expression of oxidative stress-related AKT, p-AKT, ERK, and p-ERK was downregulated with upregulation of JNK and p-JNK. The data indicate that induction of oxidative-stress related apoptosis by EADs was mediated by inhibition of AKT and ERK, and activation of JNK. The isolation of compounds in EADs was carried out using column chromatography and elucidated using the nuclear resonance magnetic analysis producing a total of six compounds including 3-epimaslinic acid, kaempferol, kaempferide, protocatechuic acid, gallic acid and ß-sitosterol-3-O-ß-D-glucopyranoside. The cytotoxicity of the isolated compounds was determined using MTT assay. Gallic acid was found to be most cytotoxic against MCF-7 cell line compared to others, with IC50 of 36 ± 1.7 µg/mL (P<0.05). In summary, EADs generated oxidative stress, induced cell cycle arrest and apoptosis in MCF-7 cells by regulating numerous genes and proteins that are involved in the apoptotic signal transduction pathway. Therefore, EADs has the potential to be developed as an anti-cancer agent against breast cancer.