Protective effect of kaempferol glucoside against lipopolysaccharide-caused acute lung injury via targeting Nrf2/NF-κB/NLRP3/GSDMD: Integrating experimental and computational studies.

The current study explored the protective potential of kaempferol 3-sophoroside-7-glucoside (KSG) against acute lung injury (ALI). Pre-treatment with KSG effectively secured mice from ALI and showed similar efficaciousness to dexamethasone. KSG markedly increased the survival rate and alleviated lung pathological lesions induced by lipopolysaccharide (LPS). Furthermore, KSG attenuated differential and total cell counts in BALF (bronchoalveolar lavage fluid) and MPO (myeloperoxidase) activity. KSG counteracted the NF-κB (nuclear factor-κB) activation and significantly ameliorated the downstream inflammatory cytokine, TNF-α (tumor necrosis factor-α). Simultaneously, KSG suppressed the over-expression of NLRP3 (NOD-like receptor protein 3), caspase-1, and pro-inflammatory cytokine interleukin IL-1ß (interleukine-1ß) and prohibited the elevation of the pyroptotic parameter GSDMD-N (N-terminal domain of gasdermin D) induced by LPS challenge. In addition, KSG significantly enhanced Nrf2 (nuclear-factor erythroid-2-related factor) and HO-1 (heme-oxygenase-1) expression. Meanwhile, KSG mitigated lipid peroxidative markers (malondialdehyde, protein carbonyl and 4-hydroxynonenal) and boosted endogenous antioxidants (superoxide dismutase/reduced glutathione/catalase) in lung tissue. In silico analyses revealed that KSG disrupts Keap1-Nrf2 protein-protein interactions by binding to the KEAP1 domain, consequently activating Nrf2. Specifically, molecular docking demonstrated superior binding affinity of KSG to KEAP1 compared to the reference inhibitor, with docking scores of -9.576 and -6.633 Kcal/mol, respectively. Additionally, the MM-GBSA binding free energy of KSG (-67.25 Kcal/mol) surpassed that of the reference inhibitor (-56.36 Kcal/mol). Furthermore, MD simulation analysis revealed that the KSG-KEAP1 complex exhibits substantial and stable binding interactions with various amino acids over a duration of 100 ns. These findings showed the protective anti-inflammatory and anti-oxidative modulatory efficiencies of KSG that effectively counteracted LPS-induced ALI and encouraged future research and clinical applications of KSG as a protective strategy for ALI.

Modulation of the crosstalk between Keap1/Nrf2/HO-1 and NF-κB signaling pathways by Tomatidine protects against inflammation/oxidative stress-driven fulminant hepatic failure in mice.

Fulminant hepatic failure (FHF) is the terminal phase of acute liver injury, which is characterized by massive hepatocyte necrosis and rapid hepatic dysfunction in patients without preexisting liver disease. There are currently no therapeutic options for such a life-threatening hepatic failure except liver transplantation; therefore, the terminal phase of the underlying acute liver injury should be avoided. Tomatidine (TOM), asteroidal alkaloid, may have different biological activities, including antioxidant and anti-inflammatory effects. Herein, the lipopolysaccharide (LPS)/D-galactosamine (D-GalN)-induced FHF mouse model was established to explore the protective potential of TOM and the underlying mechanisms of action. TOM pretreatment significantly inhibited hepatocyte necrosis and decreased serum aminotransferase activities in LPS/D-GalN-stimulated mice. TOM further increased the level of different antioxidant enzymes while reducing lipid peroxidation biomarkers in the liver. These beneficial effects of TOM were shown to be associated with targeting of NF-κB signaling pathways, where TOM repressed NF-κB activation and decreased LPS/D-GalN-induced TNF-α, IL-6, IL-1ß, and iNOS production. Moreover, TOM prevented LPS/D-GalN-induced upregulation of Keap1 expression and downregulation of Nrf2 and HO-1 expression, leading to increased Nrf2-binding activity and HO-1 levels. Besides, TOM pretreatment repressed LPS/D-GalN-induced upregulation of proliferating cell nuclear antigen (PCNA) expression, which spared the hepatocytes from damage and subsequent repair following the LPS/D-GalN challenge. Collectively, our findings revealed that TOM has a protective effect on LPS/D-GalN-induced FHF in mice, showing powerful antioxidant and anti-inflammatory effects, primarily mediated via modulating Keap1/Nrf2/HO-1 and NF-κB/TNF-α/IL-6/IL-1ß/iNOS signaling pathways.

Development of a highly sensitive fluorescent probe using Delonix regia (Gulmohar) tree pod shell for precise sarcosine detection in human urine samples: advancing prostate cancer diagnosis.

We designed a highly sensitive fluorescent sensor for the early detection of sarcosine, a potential biomarker for prostate cancer. This sensor was based on surface-cobalt-doped fluorescent carbon quantum dots (Co-CD) using a FRET-based photoluminescent sensing platform. Blue luminescent carbon quantum dots (CQD) were synthesised through a hydrothermal approach, utilizing Delonix regia tree pod shells. Cobalt was employed to functionalize the CQD, enhancing the quantum-entrapped effects and minimizing surface flaws. To optimize Co-CD preparation, we employed a Box-Behnken design (BBD), and response surface methodology (RSM) based on single-factor experiments. The Co-CD was then used as a fluorescent probe for selective Cu2+ detection, with Cu2+ quenching Co-CD fluorescence through an energy transfer process, referred to as 'turn-off'. When sarcosine was introduced, the fluorescence intensity of Co-CD was restored, creating a 'turn-on' response. The sensor exhibited a Cu2+ detection limit (LOD) of 2.4 µM with a linear range of 0 µM to 10 µM. The sarcosine detection in phosphate buffer saline (PBS, pH 7.4) resulted in an LOD of 1.54 µM and a linear range of 0 to 10 µM. Importantly, the sensor demonstrated its suitability for clinical analysis by detecting sarcosine in human urine. In summary, our rapid and highly sensitive sensor offers a novel approach for the detection of sarcosine in real samples, facilitating early prostate cancer diagnosis.Communicated by Ramaswamy H. Sarma.

Influence of rosiridin on streptozotocin-induced diabetes in rodents through endogenous antioxidants-inflammatory cytokines pathway and molecular docking study.

The research was undertaken to assess the antidiabetic activity of rosiridin in the streptozotocin (STZ)-induced diabetic model. Type 2 diabetes mellitus was elicited chemically in experimental animals using STZ (60 mg/kg, i.p.). Experimental rats were arbitrarily allocated to normal control, rosiridin perse, diabetic control, and STZ + rosiridin groups. After the confirmation of diabetes, rosiridin (10 mg/kg) was given orally to the experimental animals for 30 days. Various anti-diabetic (blood glucose, insulin), hypolipidemic, anti-inflammatory (Nuclear factor kappa B, tumour necrosis factor-α, interleukin beta (IL-1ß), and IL-6), antioxidant (and malondialdehyde level, hepatic function and others markers (ALT, AST, adiponectin, and FNDC5) and histopathological indices of injury were evaluated. In addition, the rosinidin was docked into the active site of NF-Kß (1SVC), FNDC5 (4LSD) and adiponectin (5LXG) proteins with AutoDock tools. MD simulations were carried out for the complexes of rosiridin with NF-Kß, myokine and human adiponectin receptor 1. Rosiridin treatment restored the biochemical parameters and preserved the histopathological building of the pancreas as compared to the diabetic rats. Histopathological analysis of the pancreas confirmed that rosiridin antidiabetic efficacy in the STZ-induced diabetes mellitus model. The 5LXG_rosinidin showed favourable affinity with the best binding energies at -7.534 kcal/mol. MD simulations were carried out for the complexes of rosiridin with NF-Kß, myokine and human adiponectin receptor 1, the complex of myokine and rosiridin exhibited the most stable complex. Rosiridin may exhibit considerable anti-diabetic activity in the STZ-induced diabetes mellitus model.Communicated by Ramaswamy H. Sarma.

Synthesis and in vitro antitumor activity of novel acylspermidine derivative N-(4-aminobutyl)-N-(3-aminopropyl)-8-hydroxy-dodecanamide (AAHD) against HepG2 cells.

Naturally occurring polyamines like Putrescine, Spermidine, and Spermine are polycations which bind to the DNA, hence stabilizing it and promoting the essential cellular processes. Many synthetic polyamine analogues have been synthesized in the past few years, which have shown cytotoxic effects on different tumours. In the present study, we evaluated the antiproliferative effect of a novel, acylspermidine derivative, (N-(4-aminobutyl)-N-(3-aminopropyl)-8-hydroxy-dodecanamide) (AAHD) on HepG2 cells. Fluorescence staining was performed with nuclear stain (Hoechst 33342) and acridine orange/ethidium bromide double staining. Dose and the time-dependent antiproliferative effect were observed by WST-1 assays, and radical scavenging activity was measured by ROS. Morphological changes such as cell shrinkage & blebbing were analyzed by fluorescent microscopy. It was found that AAHD markedly suppressed the growth of HepG2 cells in a dose- and time-dependent manner. It was also noted that the modulation of ROS levels confirmed the radical scavenging activity. In the near future, AAHD can be a promising drug candidate in chalking out a neoplastic strategy to control the proliferation of tumour cells. This study indicated that AAHD induced anti-proliferative and pro-apoptotic activities on HCC. Since AAHD was active at micromolar concentrations without any adverse effects on the healthy cells (Fibroblasts), it is worthy of further clinical investigations.

Recombinant l-glutaminase obtained from Geobacillus thermodenitrificans DSM-465: characterization and in silico elucidation of conserved structural domains.

Glutaminase (GLS) is an enzyme essential for amino acid metabolism; in particular, it acts as a catalyst in glutaminolysis, a reaction exploited by the malignant cells to meet the nutrient requirements for their accelerated growth and proliferation. Via regulating the initial reaction of the glutaminolysis pathway, glutaminase offers an intriguing target for the development of anticancer drugs. In the present study, we produced a recombinant glutaminase from Geobacillus thermodenitrificans DSM-465 in E. coli. The enzyme was purified to electrophoretic homogeneity, with 40% recovery and 22.36 fold purity. It exhibited a molecular weight of 33 kDa, with an optimum pH and temperature of 9 and 70 °C, respectively. The K M value of the purified enzyme was 104 µM for l-glutamine. A 3D model was built for the enzyme using Swiss-Model and subjected to molecular docking with the substrate and potential inhibitors. Moreover, the subject enzyme was compared with the human kidney type GLS-K by ConSurf and TM-align servers for evolutionary conserved residues and structural domains. Despite having less than 40% amino acid identity, the superimposed monomers of both enzymes exhibited ∼94% structural identity. With a positional difference, the active site residues Ser65, Asn117, Glu162, Asn169, Tyr193, Tyr245, and Val263 found in the bacterial enzyme were also conserved in the human GLS-K. Molecular docking results have shown that CB-839 is the best inhibitor for GLS-GT and UPGL00004 is the best inhibitor for GLS-K, as designated by the binding free energy changes, i.e. ΔG -388.7 kJ mol-1 and ΔG -375 kJ mol-1, respectively. Moreover, six potential inhibitory molecules were ranked according to their binding free energy change values for both enzymes. The information can be used for the in vivo anticancer studies.

Implications of Isoprostanes and Matrix Metalloproteinase-7 Having Potential Role in the Development of Colorectal Cancer in Males.

BACKGROUND: Colorectal cancer (CRC) is the third most common type of cancer and leading cause of death worldwide. Major risk factors involved in the development of CRC are increased dietary sources, genetics, and increasing age. Purpose of the study was to find the role of different variables in the progression of CRC. METHODOLOGY: 50 blood samples from CRC patients and 20 samples from control were collected. Serum was separated from the blood by centrifugation. This serum was assessed for several antioxidants like superoxide dismutase (SOD), glutathione, glutathione peroxidase, glutathione reductase, catalase, vitamin A, C, and E, and pro-oxidants such as malondialdehyde, advanced oxidation protein products (AOPPs), and AGEs according to their respective protocols. Matrix metalloproteinase-7 (MMP-7) and isoprostanes were assessed by ELISA kits. RESULTS: Lower levels of GSH (4.86 ± 0.78 vs 9.65 ± 1.13 µg/dl), SOD (0.08 ± 0.012 vs 0.46 ± 0.017 µg/dl), CAT (2.45 ± 0.03 vs 4.22 ± 0.19 µmol/mol of protein), and GRx (5.16 ± 0.06 vs 7.23 ± 0.36 µmol/ml) in the diseased group were recorded as compared with control. Higher levels of GPx (6.64 ± 0.19 mmol/dl) were observed in the subjects in comparison with control group (1.58 ± 0.30 mmol/dl). Highly significant decreased levels of vitamin A (0.81 ± 0.07 vs 2.37 ± 0.15 mg/ml), vitamin E (15.42 ± 1.26 vs 25.96 ± 2.19 mg/ml), and vitamin C (47.67 ± 7.69 vs 80.37 ± 10.21 mg/ml) were observed in the patients in contrast to control group. The reversal of antioxidants in later stages of CRC may be due to compensatory mechanisms in cancerous cells. The levels of MDA (nmol/ml) were also assessed, which shows significantly increased level in CRC patients as compared with control groups (3.67 ± 0.19 vs 1.31 ± 0.27). The levels of protein oxidation products [AGEs (2.74 ± 0.16 vs 0.84 ± 0.05 IU) and AOPPs (1.32 ± 0.02 vs 0.82 ± 0.07 ng/ml)] were significantly increased in subjects as compared with control. The levels of MMP-7 (64.75 ± 3.03 vs 50.61 ± 4.09 ng/ml) and isoprostanes (0.71 ± 0.03 vs 0.16 ± 0.02 ng/ml) were also analyzed. This shows that the levels of isoprostanes increased due to high lipid peroxidation mediate higher levels of MMP-7, which promotes development of CRC. CONCLUSION: Following study suggested that elevated oxidative and inflammatory status along with lipid peroxidation and matrix metalloproteinases are the chief contributors in the progression of CRC.

Prospects of IL-2 in Cancer Immunotherapy.

IL-2 is a powerful immune growth factor and it plays important role in sustaining T cell response. The potential of IL-2 in expanding T cells without loss of functionality has led to its early use in cancer immunotherapy. IL-2 has been reported to induce complete and durable regressions in cancer patients but immune related adverse effects have been reported (irAE). The present review discusses the prospects of IL-2 in immunotherapy for cancer.

POSSIBLE REGULATION OF LDL-RECEPTOR BY NARINGENIN IN HEPG2 HEPATOMA CELL LINE.

BACKGROUND: High plasma concentration of low-density lipoprotein cholesterol (LDL-c) plays a significant role in the incidence of atherosclerosis and coronary heart diseases (CHD). MATERIALS AND METHODS: The purpose of this study was to investigate the mechanism by which citrus flavonoids, naringenin regulate the LDL receptor (LDLr) gene in human liver using the human hepatoma cell line, HepG2 as a model. RESULTS: Time-course transient transfection of HepG2 cells with luciferase reporter-gene constructs incorporating the promoters of SREBP-1a,-1c, -2 and LDLr, revealed that in lipoprotein-deficient medium (LPDM), only SREBP-1a promoter activity was increased significantly after 4h exposure to 200µM naringenin respectively. However, after 24h incubation with 200µM naringenin the gene expression activities of all the SREBP-1a, -1c, -2 and LDLr promoter-constructs were increased significantly. The effects of both 200µM naringenin on elevating LDLr mRNA are possibly due to regulation of gene transcription by SREBP-la and SREBP-2. However, the suppression effect of 200µM naringenin on hepatic SREBP-1c mRNA expression is likely associated with the reduction in mRNA expression of both acetyl-CoA carboxylase and fatty acid synthase in human hepatoma HepG2 cells. It was found that, 200µM naringenin was likely to stimulate LDLr gene expression via increase phosphorylation of PI3K and ERK1/2 which enhance the transcription factors SREBP-1a and SREBP-2 mRNA levels and increased their protein maturation in human hepatoma HepG2 cell. CONCLUSION: Diets supplemented with naringenin could effectively reduce mortality and morbidity from coronary heart diseases and as cardio-protective effects in humans.