Tocotrienol-rich Fraction Modulates Cardiac Metabolic Profile changes in Isoprenaline-Induced Myocardial Infarction rats

In myocardial infarction (MI), the occurrence of energy depletion, oxidative stress, and decreased amino acids metabolism alter tissue metabolites. Evidence has shown that tocotrienol-rich fraction (TRF) prevents myocardial injury in MI. However, the protective mechanism at the metabolite level is unknown. Male Sprague-Dawley rats were grouped into control, isoprenaline (ISO)-induced MI (MI), healthy rats receiving 200 mg/kg TRF (200TRF), and MI rats receiving 200 mg/kg TRF (200TRF+MI) groups. TRF was administered via oral gavage daily for 12 weeks followed by intraperitoneal ISO injection (85 mg/kg) for two consecutive days at a 24-hour interval to induce MI. High-performance liquid chromatography was performed to analyze serum α-tocopherol and tocotrienol concentration whereas ultrahigh-performance liquid chromatography-mass spectrometry was used for the untargeted metabolomic study. Serum α-tocopherol but not tocotrienol was increased in the 200TRF (p=0.121) and 200TRF+MI (p<0.05) following TRF supplementation. Multivariate analysis by Orthogonal Projections to Latent Structures Discriminant Analysis showed high predictability of the group comparison models for MI vs control and 200TRF+MI vs MI (cross-validation: Q2 >0.7, R2 Y>0.8, p<0.05). A total of 84 and 37 metabolites [when covariance of p≥|0.05| (magnitude) and p(corr)≥|0.5| (reliability)] were significantly different in the myocardial homogenates of MI vs control and 200TRF+MI vs MI, respectively. MI rats had reduced S-adenosylmethionine and L-cystathionine that might worsen MI by disturbing glutathione metabolism; decreased phosphoribosyl-pyrophosphate and purine salvage process that might impair DNA synthesis, and elevated glucose-6-phosphate suggesting enhanced anaerobic glycolysis possibly for rapid production of energy. Conversely, TRF supplementation reversed the impaired metabolic pathways caused by MI

Tocotrienol-rich fraction modulates cardiac metabolic profile changes in isoprenaline-induced myocardial infarction rats

In myocardial infarction (MI), the occurrence of energy depletion, oxidative stress, and decreased amino acids metabolism alter tissue metabolites. Evidence has shown that tocotrienol-rich fraction (TRF) prevents myocardial injury in MI. However, the protective mechanism at the metabolite level is unknown. Male Sprague-Dawley rats were grouped into control, isoprenaline (ISO)-induced MI (MI), healthy rats receiving 200 mg/kg TRF (200TRF), and MI rats receiving 200 mg/kg TRF (200TRF+MI) groups. TRF was administered via oral gavage daily for 12 weeks followed by intraperitoneal ISO injection (85 mg/kg) for two consecutive days at a 24-hour interval to induce MI. High-performance liquid chromatography was performed to analyze serum α-tocopherol and tocotrienol concentration whereas ultra-high-performance liquid chromatography-mass spectrometry was used for the untargeted metabolomic study. Serum α-tocopherol but not tocotrienol was increased in the 200TRF (p=0.121) and 200TRF+MI (p0.7, R2Y>0.8, p<0.05). A total of 84 and 37 metabolites [when covariance of p≥|0.05| (magnitude) and p(corr)≥|0.5| (reliability)] were significantly different in the myocardial homogenates of MI vs control and 200TRF+MI vs MI, respectively. MI rats had reduced S-adenosylmethionine and L-cystathionine that might worsen MI by disturbing glutathione metabolism; decreased phosphoribosyl-pyrophosphate and purine salvage process that might impair DNA synthesis, and elevated glucose-6-phosphate suggesting enhanced anaerobic glycolysis possibly for rapid production of energy. Conversely, TRF supplementation reversed the impaired metabolic pathways caused by MI

Preventative and therapeutic potential of tocotrienols on musculoskeletal diseases in ageing

Musculoskeletal health is paramount in an ageing population susceptible to conditions such as osteoporosis, arthritis and fractures. Age-related changes in bone, muscle, and joint function result in declining musculoskeletal health, reduced mobility, increased risk of falls, and persistent discomfort. Preserving musculoskeletal wellbeing is essential for maintaining independence and enhancing the overall quality of life for the elderly. The global burden of musculoskeletal disorders is significant, impacting 1.71 billion individuals worldwide, with age-related muscle atrophy being a well-established phenomenon. Tocotrienols, a unique type of vitamin E found in various sources, demonstrate exceptional antioxidant capabilities compared to tocopherols. This characteristic positions them as promising candidates for addressing musculoskeletal challenges, particularly in mitigating inflammation and oxidative stress underlying musculoskeletal disorders. This review paper comprehensively examines existing research into the preventive and therapeutic potential of tocotrienols in addressing age-related musculoskeletal issues. It sheds light on the promising role of tocotrienols in enhancing musculoskeletal health and overall wellbeing, emphasizing their significance within the broader context of age-related health concerns

Characterization of keratinocytes, fibroblasts and melanocytes isolated from human skin using gene markers

Cells isolated from skin have wide applications in studies of the pathogenesis of skin-related diseases and the construction of 3D skin equivalents. This study aimed to isolate keratinocytes, fibroblasts, and melanocytes from human foreskin and characterize the purity of the cell types. Keratinocytes, fibroblasts, and melanocytes from human foreskin were isolated by differential trypsinization and media selection. The purity of the cell types was characterized based on the expression of gene markers. The assessment of gene marker expression involved RNA extraction, primer design, quantitative polymerase chain reaction (qPCR) and immunocytochemical staining. Our results showed that in cocultures of keratinocytes and fibroblasts isolated from the dermis, fibroblasts could be separated from keratinocytes by quick trypsinization and culture in Dulbecco’s modified Eagle’s medium. The remaining keratinocytes are cultured in Epilife medium. Melanocytes in cocultures of melanocytes and keratinocytes isolated from the epidermis could be selected by changing Epilife medium to M254 medium. Gene marker results suggested that cytokeratin 14 (CK14) is a suitable marker for keratinocytes, elastin (ELN) is a suitable marker for fibroblasts, and tyrosinase (TYR) and tyrosinase-related proteins 1 and 2 (TYRP1 and TYRP2) are suitable markers for melanocytes. In conclusion, keratinocytes, fibroblasts, and melanocytes can be isolated from the same human foreskin sample by differential trypsinization and media selection techniques and characterized by suitable gene markers. This finding will aid in the isolation of pure skin cell types for various applications in regenerative medicine and toxicity studies

Gamma-tocotrienol acts as a BH3 mimetic to induce apoptosis in neuroblastoma SH-SY5Y cells

Bcl-2 family proteins are crucial regulators of apoptosis. Both pro- and antiapoptotic members exist, and overexpression of the latter facilitates evasion of apoptosis in many cancer types. Bcl-2 homology domain 3 (BH3) mimetics are small molecule inhibitors of antiapoptotic Bcl-2 family members, and these inhibitors are promising anticancer agents. In this study, we report that gamma-tocotrienol (γT3), an isomer of vitamin E, can inhibit Bcl-2 to induce apoptosis. We demonstrate that γT3 induces cell death in human neuroblastoma SH-SY5Y cells by depolarising the mitochondrial membrane potential, enabling release of cytochrome c to the cytosol and increasing the activities of caspases-9 and -3. Treatment of cells with inhibitors of Bax or caspase-9 attenuated the cell death induced by γT3. Simulated docking analysis suggested that γT3 binds at the hydrophobic groove of Bcl-2, while a binding assay showed that γT3 competed with a fluorescent probe to bind at the hydrophobic groove. Our data suggest that γT3 mimics the action of BH3-only protein by binding to the hydrophobic groove of Bcl-2 and inducing apoptosis via the intrinsic pathway in a Bax- and caspase-9-dependent manner

Comparison of the antioxidant activity of Malaysian ginger (Zingiber officinale Roscoe) extracts with that of selected natural products and its effect on the viability of myoblast cells in culture

Ginger has been proven to possess various therapeutic effects, including antibacterial, anticancer, anti-inflammatory, and antioxidant effects. However, data on the comparison of ginger antioxidant activity with that of other natural products are still lacking. This study aimed to analyse and compare the antioxidant properties of two types of Malaysian ginger extracts (GE1 and GE2) with that of selected natural products. The antioxidant activities were measured by 2,2-diphenyl-1-picryl-hydrazyl-hydrate (DPPH) and ferric reducing antioxidant power (FRAP) assays, while cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfonyl)-2H-tetrazolium (MTS) assay. The order of the DPPH scavenging activities was as follows: vitamin C > palm tocotrienol-rich fraction (TRF) > a-tocopherol > N-acetylcysteine (NAC) > Ficus deltoidea > butylated hydroxytoluene (BHT) > Centella asiatica > GE2 > GE1 > Moringa oleifera > Kelulut honey; the order of the mean FRAP value was as follows: NAC > a-tocopherol > BHT > TRF > Ficus deltoidea > Moringa oleifera > GE2 = GE1 > Centella asiatica > Kelulut honey. The viability assays showed that both ginger extracts significantly increased the percentage of viable cells (p < 0.05). In conclusion, neither of the ginger extracts was cytotoxic toward cells and both possessed comparable antioxidant properties, indicating their potential for ameliorating oxidative stress

Effects of age and tocotrienol-rich fraction on mitochondrial respiratory complexes in the hippocampus of rats

Mitochondrial dysfunction is common in the brain with age. Prevention of mitochondrial dysfunction at an early age may protect the brain against neurodegeneration in later life. Tocotrienol-rich fraction (TRF) has been reported to be neuroprotective in old rats, but its effect remains unknown for middle-aged animals. This study aimed to determine the effect of TRF on activities of mitochondrial respiratory chain complexes in the hippocampus of middle-aged rats. Male Sprague Dawley rats were divided into 4 groups: young control (3 months old), adult control (12 months old), adult rats supplemented with palm kernel oil (PKO) as the vehicle, and adult rats supplemented with TRF by gavage at 200 mg/kg body weight/day for 3 months. At the end of the supplementation, activities of complex I, I+III, II, II+III, III, IV, and citrate synthase in the isolated mitochondria of the hippocampus were measured by spectrophotometry. Complex II activity was higher, while citrate synthase activity was lower in adult rats than in young rats. A decrease of citrate synthase activity suggests loss of mitochondrial mass and intactness in the hippocampus at middle age. Interestingly, PKO-treated adult rats had lower complex I and IV activities, but higher complex I+III activity than adult control rats. These findings indicate PKO modulated activities of the complexes. In TRF-treated adult rats, the complex I activity was higher, while the complex IV activity was lower than PKO-treated adult rats. TRF restored the complex I activity and may have the potential to reverse complex I deficiency

LC-MS/MS and GC-MS Analysis for the Identification of Bioactive Metabolites Responsible for the Antioxidant and Antibacterial Activities of Lygodium microphyllum (Cav.) R. Br

Natural products serve as a valuable source of antioxidants with potential health benefits for various conditions. Lygodium microphyllum (Cav.) R. Br., also known as Old World climbing fern, is an invasive climbing fern native to Southeast Asia, Africa, South America, Australia, and Melanesia. It has been reported to possess interesting pharmacological properties including hepatoprotective and anti-inflammatory mechanisms. This study analyzed the potential bioactive metabolites that contribute to the antioxidant and antimicrobial effects of L. microphyllum (LM) by profiling the crude extract using high-resolution LC-MS/MS and GC-MS systems. Several classes of compounds such as phenolics, flavonoids, terpenoids, steroids, macrolides, vitamins, lipids, and other hydrocarbons were found in the crude extract of LM through non-targeted analysis. A total of 74 compounds were detected in LC-MS/MS, whereas a total of nine compounds were identified in GC-MS. Out of the 74 compounds detected in LC-MS/MS, 34 compounds, primarily quercetin, kaempferol, trifolin, pyroglutamic acid, arachidonic acid, and rutin were reported with antioxidant, antimicrobial, antiinflammatory, and hepatoprotective activities. The presence of phenolic and flavonoid compounds with reported bioactivities in the crude extract of LM evidence its pharmacological properties

Antimicrobial Potential of Aqueous Extract of Giant Sword Fern and Ultra-High-Performance Liquid Chromatography–High-Resolution Mass Spectrometry Analysis

Vibriosis and parasitic leech infestations cause the death of various farmed fish, such as groupers, hybrid groupers, sea bass, etc., in Malaysia and other Southeast Asian countries. In the absence of natural control agents, aquaculture operators rely on toxic chemicals to control Vibrio infections and parasitic leeches, which can have a negative impact on the environment and health. In the present study, we investigated the antivibrio and antiparasitic activities of the aqueous extract of giant sword fern (GSF) (Nephrolepis biserrata, Nephrolepidaceae, locally known as “Paku Pedang”) against four Vibrio spp. and the parasitic leech Zeylanicobdella arugamensis, as well as its metabolic composition using the ultra-high-performance liquid chromatography–high-resolution mass spectrometry system (UHPLC-HRMS). The data show that the aqueous extract of GSF at a concentration of 100 mg/mL exhibits potent bactericidal activity against V. parahaemolyticus with a zone of inhibition of 19.5 mm. In addition, the extract showed dose-dependent activity against leeches, resulting in the complete killing of the parasitic leeches within a short period of 11–43 min when tested at concentrations ranging from 100 to 25 mg/mL. The UHPLC-HRMS analysis detected 118 metabolites in the aqueous extract of GSF. Flavonoids were the primary metabolites, followed by phenolic, aromatic, fatty acyl, terpenoid, vitamin and steroidal compounds. Notably, several of these metabolites possess antibacterial and antiparasitic properties, including cinnamaldehyde, cinnamic acid, apigenin, quercetin, cynaroside, luteolin, naringenin, wogonin, 6-gingerol, nicotinamide, abscisic acid, daidzein, salvianolic acid B, etc. Overall, our study shows the significant antibacterial and antiparasitic potential of the GSF aqueous extract, which demonstrates the presence of valuable secondary metabolites. Consequently, the aqueous extract is a promising natural alternative for the effective control of Vibrio infections and the treatment of parasitic leeches in aquaculture systems

Zingiber officinale Roscoe prevents DNA damage and improves muscle performance and bone integrity in old Sprague Dawley rats

Age-related loss of skeletal muscle mass and strength or sarcopenia is attributed to the high level of oxidative stress and inadequate nutritional intake. The imbalance in oxidative status with increased production of free radicals results in damage to the DNA which leads to cell dysfunction. This study aimed to determine the effect of Zingiber officinale Roscoe (ginger) on muscle performance and bone integrity in Sprague Dawley (SD) rats. SD rats aged three (young), nine (adult), and twenty-one (old) months old were treated with either distilled water or ginger extract at a concentration of 200 mg/kg body weight (BW) daily for 3 months via oral gavage. Muscle performance was assessed at 0, 1, 2, and 3 months of treatment by measuring muscle strength, muscle function, and bone integrity while DNA damage was determined by comet assay. Muscle cell histology was analyzed by hematoxylin and eosin (H&E) staining. Young and adult ginger-treated rats showed a significant improvement in muscle strength after 3 months of supplementation. Bone mineral density (BMD) and bone mineral content (BMC) were increased while fat free mass (FMM) was decreased after 3 months of ginger supplementation in young rats but not changed in adult and old ginger supplemented groups. Interestingly, supplementation of ginger for 3 months to the old rats decreased the level of damaged DNA. Histological findings showed reduction in the size of muscle fibre and fascicles with heterogenous morphology of the muscle fibres indicating sarcopenia was evident in old rats. Treatment with ginger extract improved the histological changes even though there was evidence of cellular infiltration (mild inflammation) and dilated blood vessels. In conclusion, Z. officinale Roscoe prevents DNA damage and improves muscle performance and bone integrity in SD rats indicating its potential in alleviating oxidative stress in ageing and thus delaying sarcopenia progression