MATERIALS AND METHODS: In this study, DET (0.625. 1.25 and 2.5 mg/kg, i.p.) was administered in rats for 21 days and those animals were challenged with single injection of LPS (250 μg/kg, i.p.) for 7 days. Cognitive and behavioral assessment was carried out for 7 days followed by molecular assessment on brain hippocampus. Statistical significance was analyzed with one-way analysis of variance followed by Dunnett's test to compare the treatment groups with the control group.
KEY FINDINGS: DET ameliorated LPS-induced neuroinflammation by suppressing major pro-inflammatory mediators such as iNOS and COX-2. Furthermore, DET enhanced the anti-inflammatory cytokines and concomitantly suppressed the pro-inflammatory cytokines and chemokine production. DET treatment also reversed LPS-induced behavioral and memory deficits and attenuated LPS-induced elevation of the expression of AD markers. DET improved synaptic-functionality via enhancing the activity of pre- and post-synaptic markers, like PSD-95 and SYP. DET also prevented LPS-induced apoptotic neurodegeneration via inhibition of PARP-1, caspase-3 and cleaved caspase-3.
SIGNIFICANCE: Overall, our studies suggest DET can prevent neuroinflammation-associated memory impairment and neurodegeneration and it could be developed as a therapeutic agent for the treatment of neuroinflammation-mediated and neurodegenerative disorders, such as AD.
AIM OF STUDY: To investigate the potential protective effects of L. flavescens in pancreatic β cells through inhibition of apoptosis and autophagy cell death mechanisms in in vitro and in vivo models.
MATERIALS AND METHODS: L. flavescens leaves were extracted using solvent in increasing polarities: hexane, ethyl acetate, methanol and water. All extracts were tested for INS-1 β cells viability stimulated by streptozotocin (STZ). The extract which promotes the highest cell protective activity was further evaluated for insulin secretion, apoptosis and autophagy signaling pathways. Then, the acute toxicity of extract was carried out in SD rats according to OECD 423 guideline. The active extract was tested in diabetic rats where the pancreatic β islets were evaluated for insulin, apoptosis and autophagy protein.
RESULTS: The methanolic extract of L. flavescens (MELF) was found to increase INS-1 β cells viability and insulin secretion against STZ. In addition, MELF has been shown to inhibit INS-1 β cells apoptosis and autophagy activity. Notably, there was no toxicity observed in SD rats when administered with MELF. Furthermore, MELF exhibited anti-hyperglycemic activity in diabetic rats where apoptosis and autophagy protein expression was found to be suppressed in pancreatic β islets.
CONCLUSION: MELF was found to protect pancreatic β cells function from STZ-induced apoptosis and autophagy in in vitro and in vivo.
AIM OF THE REVIEW: This review aims to provide a comprehensive report on the ethnomedicinal use, phytochemistry, pharmacological activities, molecular mechanisms, and nutritional values of C. nutans. The present review will open new avenues for further in-depth pharmacological studies of C. nutans for it to be developed as a potential nutraceutical and to improve the available products in the market.
MATERIAL AND METHODS: All the available information on C. nutans was collected using the key words "Clinacanthus nutans" and/or "ethnomedicine" and/or "phytochemicals" and/or "anticancer" and/or "anti-inflammatory" and/or "antiviral" through an electronic search of the following databases: PubMed, Web of Science, EMBASE, Cochrane Library, Clinical Trials.org, SciFinder Scholar, Scopus, and Google Scholar. In addition, unpublished materials, Ph.D. and M.Sc. dissertations, conference papers, and ethnobotanical textbooks were used. The Plant List (www.theplantlist.org) and International Plant Name Index databases were used to validate the scientific name of the plant.
RESULTS: The literature supported the ethnomedicinal uses of C. nutans as recorded in Thailand, Indonesia, and Malaysia for various purposes. Bioactivities experimentally proven for C. nutans include cytotoxic, anticancer, antiviral, anti-inflammatory, immunomodulatory, antidiabetic, antioxidant, antihyperlipidemic, antimicrobial, and chemotherapeutic (in aquaculture) activities. Most of these activities have so far only been investigated in chemical, cell-based, and animal assays. Various groups of phytochemicals including five sulfur-containing glycosides, eight chlorophyll derivatives, nine cerebrosides, and a monoacylmonogalactosyl glycerol are present in C. nutans. The presence of two glycerolipids, four sulfur-containing compounds, six known flavones, a flavanol, four flavonols, two phytosterols, one polypeptide, and various phenolics and fatty acids largely influences its diverse bioactivities. Numerous reports justify the ethnomedicinal use of C. nutans as an antiviral agent in treating herpes simplex virus and varicella-zoster virus infections and as part of a traditional anticancer anti-inflammatory concoction agent for various inflammatory diseases. C. nutans tea was reported to have a good percentage of carbohydrate, crude protein, minerals, essential amino acids, nonessential amino acids, and essential fatty acids. Acute, subacute, and subchronic toxicity studies demonstrated that oral administration of ethanol and methanol extracts of C. nutans to male Swiss albino mice and male Sprague-Dawley (SD) rats, respectively, did not lead to any toxicity or adverse effects on the animal behavior and organs when used in amounts as high as 2g/kg.
CONCLUSION: The collected literatures demonstrated that, as an important traditional medicine, C. nutans is a promising ethnomedicinal plant with various extracts and bioactive compounds exhibiting multifarious bioactivities. However, it is important for future studies to conduct further in vitro and in vivo bioactivity evaluations systematically, following the standard pharmacology guidelines. It is crucial to elucidate in-depth molecular mechanisms, structure-activity relationships, and potential synergistic and antagonistic effects of multi-component extracts and bioactive constituents derived from C. nutans. Further studies should also focus on comprehensive toxicity that includes long-term effects and adverse effects on target organs of C. nutans and bioactive compounds in correlation with the specific pharmacological effects.
METHODS: The effect of AMEAE on cell proliferation of different cell lines was analyzed by MTT assay. High content screening (HCS) was applied to investigate the suppression of NF-κB translocation, cell membrane permeability, mitochondrial membrane potential (MMP) and cytochrome c translocation from mitochondria to cytosol. Reactive oxygen species (ROS) formation, lactate dehydrogenase (LDH) release and activation of caspase-3/7, -8 and -9 were measured while treatment. The western blot analysis also carried out to determine the protein expression of cleaved caspase-3 and -9. Flow cytometry analysis was used to determine the cell cycle distribution and phosphatidylserine externalization. Quantitative PCR analysis was performed to measure the gene expression of Bax and Bcl-2 proteins.
RESULTS: Cell viability analysis revealed the selective cytotoxic effect of AMEAE towards lung cancer cells, A549, with an IC50 value of 5.09 ± 0.41 μg/mL after 72 h of treatment. Significant LDH leakage and phosphatidylserine externalization were observed in AMEAE treated cells by fluorescence analysis. Treatment of A549 cells with AMEAE significantly elevated ROS formation, followed by attenuation of MMP via upregulation of Bax and downregulation of Bcl-2, accompanied by cytochrome c release to the cytosol. The incubation of A549 cells with superoxide dismutase and catalase significantly attenuated the cytotoxicity caused by AMEAE, indicating that intracellular ROS plays a pivotal role in cell death. The released cytochrome c triggered the activation of caspase-9 followed by caspase-3. In addition, AMEAE-induced apoptosis was accompanied by cell cycle arrest at G0/G1 phase. Moreover, AMEAE suppressed the induced translocation of NF-κB from cytoplasm to nucleus.
CONCLUSIONS: Our data showed for the first time that the ethyl acetate extract of Annona muricata inhibited the proliferation of A549 cells, leading to cell cycle arrest and programmed cell death through activation of the mitochondrial-mediated signaling pathway with the involvement of the NF-kB signalling pathway.