Anti-Inflammatory Effects of Rhamnetin on Bradykinin-Induced Matrix Metalloproteinase-9 Expression and Cell Migration in Rat Brain Astrocytes

Bradykinin (BK) has been shown to induce matrix metalloproteinase (MMP)-9 expression and participate in neuroinflammation. The BK/MMP-9 axis can be a target for managing neuroinflammation. Our previous reports have indicated that reactive oxygen species (ROS)-mediated nuclear factor-kappaB (NF-κB) activity is involved in BK-induced MMP-9 expression in rat brain astrocytes (RBA-1). Rhamnetin (RNT), a flavonoid compound, possesses antioxidant and anti-inflammatory effects. Thus, we proposed RNT could attenuate BK-induced response in RBA-1. This study aims to approach mechanisms underlying RNT regulating BK-stimulated MMP-9 expression, especially ROS and NF-κB. We used pharmacological inhibitors and siRNAs to dissect molecular mechanisms. Western blotting and gelatin zymography were used to evaluate protein and MMP-9 expression. Real-time PCR was used for gene expression. Wound healing assay was applied for cell migration. 2ʹ,7ʹ-dichlorodihydrofluorescein diacetate (H2DCF-DA) and nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) were used for ROS generation and NOX activity, respectively. Promoter luciferase assay and chromatin immunoprecipitation (ChIP) assay were applied to detect gene transcription. Our results showed that RNT inhibits BK-induced MMP-9 protein and mRNA expression, promoter activity, and cell migration in RBA-1 cells. Besides, the levels of phospho-PKCδ, NOX activity, ROS, phospho-ERK1/2, phospho-p65, and NF-κB p65 binding to MMP-9 promoter were attenuated by RNT. In summary, RNT attenuates BK-enhanced MMP-9 upregulation through inhibiting PKCδ/NOX/ROS/ERK1/2-dependent NF-κB activity in RBA-1.

Comparative Study on Phenolic Compounds and Antioxidant Activities of Hop (Humulus lupulus L.) Strobile Extracts

In this study, we investigated the phenolic compounds in hop strobile extracts and evaluated their antioxidant property using DPPH and ABTS assay. The total phenolic compound (TPC) and total flavonoid compound (TFC) estimated in two different solvent extracts considerably varied depending on the extraction solvent. The most abundant phenolic compound in hop strobile was humulones (α-acid) with levels ranging from 50.44 to 193.25 µg/g. El Dorado accession revealed higher antioxidant activity in ethanol extracts (DPPH: IC50 124.3 µg/mL; ABTS: IC50 95.4 µg/mL) when compared with that of the other accessions. Correlations between DPPH (IC50) scavenging TFC in ethanol extract (TFC_E, −0.941), and TPC_E (−0.901), and between ABTS (IC50) scavenging TFC_E (−0.853), and TPC_E (−0.826), were statistically significant at p < 0.01 level, whereas no significant correlation was observed between antioxidant activities, TPC and TFC in water extract. This study is the first to report that variations in the level of phenolic contents and antioxidant activity of various hop cultivars depended on the type of extraction solvent used and the cultivation regions. These results could provide valuable information on developing hop products.

Tanshinone I Mitigates Steroid-Induced Osteonecrosis of the Femoral Head and Activates the Nrf2 Signaling Pathway in Rats

Steroid-induced osteonecrosis of the femoral head (SIONFH) is a frequent orthopedic disease caused by long-term or high-dose administration of corticosteroids. Tanshinone I (TsI), a flavonoid compound isolated from Salvia miltiorrhiza Bunge, has been reported to inhibit osteoclastic differentiation in vitro. This study aimed to investigate whether TsI can ameliorate SIONFH. Herein, SIONFH was induced by intraperitoneal injection of 20 μg/kg lipopolysaccharide every 24 h for 2 days, followed by an intramuscular injection of 40 mg/kg methylprednisolone every 24 h for 3 days. Four weeks after the final injection of methylprednisolone, the rats were intraperitoneally administrated with low-dose (5 mg/kg) and high-dose (10 mg/kg) TsI once daily for 4 weeks. Results showed that TsI significantly alleviated osteonecrotic lesions of the femoral heads as determined by micro-CT analysis. Furthermore, TsI increased alkaline phosphatase activity and expressions of osteoblastic markers including osteocalcin, type I collagen, osteopontin, and Runt-related transcription factor 2 and decreased tartrate-resistant acid phosphatase activity and expressions of osteoclastic markers including cathepsin K and acid phosphatase 5. TsI also reduced inflammatory response and oxidative stress and activated the nuclear factor erythroid 2-related factor 2 (Nrf2) signaling pathway in the femoral heads. Taken together, our findings show that TsI can relieve SIONFH, indicating that it may be a candidate for preventing SIONFH.

Preclinical toxicity test results of a new antiviral–immune-modulator compound consisting of flavonoid molecules (COVID-19 clinical trial preliminary data)

Aim: Isolated specific glycone–aglycone conjugated flavonoids which are investigated for their effect of bioavailability and molecular concentrations. The specific formula is then tested via in vitro and in vivo cytotoxicity tests. Methods: Considering the higher affinity for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), quercetin, quercetin 3-sambubioside-3’-glucoside, luteolin, apigenin-7-4’alloside, kaempferol-7-O-glucoside, epicatechin-epigallocatechin-3-O-gallate, and hesperetin were selected to investigate the effects of a new combination of the formula. Specific chemical analyses, such as high-performance liquid chromatography (HPLC), liquid chromatography–mass spectrometry (LC–MS), quadrupole time of flight mass spectrometry (QTOF–MS) analysis and ultraviolet–visible (UV–VIS) spectrophotometry, were performed for molecular qualification and quantification. Results: In silico molecular docking analyses have shown that flavonoids can bind strongly to the spike protein and main protease of the SARS-CoV-2 virus. Flavonoids also have anti-inflammatory and immune-modulating activity by inhibiting cytokines. Although flavonoids may be a treatment alternative for coronavirus disease 2019 (COVID-19), an effective flavonoid compound has yet to be developed. The main problem here is that the absorption rate of flavonoids is very low (2–10%) in the intestines, and these compounds are metabolized rapidly. In contrast, according to recent literature, a conjugated flavonoid mixture is better absorbed in the small intestine, and its toxic effects are relatively fewer. Conclusions: It is found that the new formula has no cytotoxic or genotoxic effects. Furthermore, oral administrations of the new compound did not produce any toxicity symptoms or any mortality in male and female rats. The pre-clinical in vitro and in vivo toxicity test results indicated that the new flavonoid formula can be safely used for clinical trials.

A Flavonoid compound of Turbinaria decurrens Bory with The Potential Antioxidant and Anticancer Activity

Isolation and characterization of quercetin flavonoid compound from Dutungan Island, South Sulawesi Province, Indonesia has been successfully done from ethyl acetate extract. Extraction method used maceration, isolation used chromatography, anticancer activity with MTT method and antioxidant test used DPPH radical. Structure was discussed with the FT-IR, NMR spectrophotometer and compared with the literature. Total flavonoids from ethyl acetate extract were 4.8 mgEQ/g, IC50 value of antioxidant activity was 4.23 μg/ml using the DPPH method (2,2-diphenyl-1-picrylhydrazyl), and anticancer activity of H460 cells IC50 value was 10.95 μg/ml. The quercetin compound is potential as an anticancer and was first report in the T. decurrens Bory species.

A Multidrug Approach to Modulate the Mitochondrial Metabolism Impairment and Relative Oxidative Stress in Fanconi Anemia Complementation Group A

Fanconi Anemia (FA) is a rare recessive genetic disorder characterized by aplastic anemia due to a defective DNA repair system. In addition, dysfunctional energy metabolism, lipid droplets accumulation, and unbalanced oxidative stress are involved in FA pathogenesis. Thus, to modulate the altered metabolism, Fanc-A lymphoblast cell lines were treated with quercetin, a flavonoid compound, C75 (4-Methylene-2-octyl-5-oxotetrahydrofuran-3-carboxylic acid), a fatty acid synthesis inhibitor, and rapamycin, an mTOR inhibitor, alone or in combination. As a control, isogenic FA cell lines corrected with the functional Fanc-A gene were used. Results showed that: (i) quercetin recovered the energy metabolism efficiency, reducing oxidative stress; (ii) C75 caused the lipid accumulation decrement and a slight oxidative stress reduction, without improving the energy metabolism; (iii) rapamycin reduced the aerobic metabolism and the oxidative stress, without increasing the energy status. In addition, all molecules reduce the accumulation of DNA double-strand breaks. Two-by-two combinations of the three drugs showed an additive effect compared with the action of the single molecule. Specifically, the quercetin/C75 combination appeared the most efficient in the mitochondrial and lipid metabolism improvement and in oxidative stress production reduction, while the quercetin/rapamycin combination seemed the most efficient in the DNA breaks decrement. Thus, data reported herein suggest that FA is a complex and multifactorial disease, and a multidrug strategy is necessary to correct the metabolic alterations.

Bioactive compounds derived from Streptomyces sp. SA32: antibacterial activity, chemical profile, and their related genes

Microbes which are resistant to drugs and antibiotics as well as multi-drug resistant (MDR) microbes have developed due to the improper use of antibiotics and led to explore the microbial isolates as the sources of new antibiotics or those with highly effective and stabile attack. Streptomyces sp. SA32 was selected to inhibit the growth of MDR bacteria Escherichia coli, Staphylococcus aureus, Klebsiella pneumonia, Pseudomonas aeruginosa, Enterobacter cloaceae, and Enterococcus sp. E. coli was sensitive to crude extracts of Streptomyces sp. SA32 at the concentration of 19 g.mL-1 with moderate strength against E. cloacae. The bioactive compounds analyzed using thin layer chromatography and phytochemical methods showed that the spot with Rf 0.63 and 0.68 was polyketide compound and that with Rf 0.74 was flavonoid compound. The bioautography assay on the TLC plate confirmed the absence of MDR bacterial growth on polyketide and flavonoid spots. The synthesis of antibacterial compounds was also confirmed by the successful analysis on both non-ribosomal peptide synthetase (NRPS) and polyketide synthase (PKS) gene sequences.

Trilobatin Alleviates Cognitive Deficits and Pathologies in an Alzheimer’s Disease Mouse Model

Alzheimer’s disease (AD) is the most common neurodegenerative disease nowadays that causes memory impairments. It is characterized by extracellular aggregates of amyloid-beta (Aβ), intracellular aggregates of hyperphosphorylated Tau (p-Tau), and other pathological features. Trilobatin (TLB), a natural flavonoid compound isolated from Lithocarpuspolystachyus Rehd., has emerged as a neuroprotective agent. However, the effects and mechanisms of TLB on Alzheimer’s disease (AD) remain unclear. In this research, different doses of TLB were orally introduced to 3×FAD AD model mice. The pathology, memory performance, and Toll-like receptor 4- (TLR4-) dependent inflammatory pathway protein level were assessed. Here, we show that TLB oral treatment protected 3×FAD AD model mice against the Aβ burden, neuroinflammation, Tau hyperphosphorylation, synaptic degeneration, hippocampal neuronal loss, and memory impairment. The TLR4, a pattern recognition immune receptor, has been implicated in neurodegenerative disease-related neuroinflammation. We found that TLB suppressed glial activation by inhibiting the TLR4-MYD88-NFκB pathway, which leads to the inflammatory factor TNF-α, IL-1β, and IL-6 reduction. Our study shows that TLR4 might be a key target of TLB in AD treatment and suggests a multifaceted target of TLB in halting AD. Taken together, our findings suggest a potential therapeutic effect of TLB in AD treatment.