Phytol from Scoparia dulcis prevents NF-κB-mediated inflammatory responses during macrophage polarization.

Macrophages are primary immune cells that mediate a wide range of inflammatory diseases through their polarization potential. In this study, phytol isolated from Scoparia dulcis has been explored against 7-ketocholesterol and bacterial lipopolysaccharide-induced macrophage polarization in IC-21 cells. Isolated phytol has been characterized using GC-MS, TLC, HPTLC, FTIR, 1H-NMR, and HPLC analyses. The immunomodulatory effects of viable concentrations of phytol were tested on oxidative stress, arginase activity, nuclear and mitochondrial membrane potentials in IC-21 cells in addition to the modulation of calcium and lipids. Further, gene and protein expression of atherogenic markers were studied. Results showed that the isolated phytol at a viable concentration of 400 µg/ml effectively reduced the production of nitric oxide, superoxide anion (ROS generation), calcium and lipid accumulation, stabilized nuclear and mitochondrial membranes, and increased arginase activity. The atherogenic markers including iNOS, COX-2, IL-6, IL-1ß, MMP-9, CD36, and NF-κB were significantly downregulated at the levels of gene and protein expression, while macrophage surface and nuclear receptor markers (CD206, CD163, and PPAR-γ) were significantly upregulated by phytol pre-treatment in macrophages. Therefore, the present pharmacognostic study supports the role of phytol isolated from Scoparia dulcis in preventing M2-M1 macrophage polarization under inflammatory conditions, making it a promising compound. Supplementary Information: The online version contains supplementary material available at 10.1007/s13205-024-03924-9.

Interactions between macrophage membrane and lipid mediators during cardiovascular diseases with the implications of scavenger receptors.

The onset and progression of cardiovascular diseases with the major underlying cause being atherosclerosis, occur during chronic inflammatory persistence in the vascular system, especially within the arterial wall. Such prolonged maladaptive inflammation is driven by macrophages and their key mediators are generally attributed to a disparity in lipid metabolism. Macrophages are the primary cells of innate immunity, endowed with expansive membrane domains involved in immune responses with their signalling systems. During atherosclerosis, the membrane domains and receptors control various active organisations of macrophages. Their scavenger/endocytic receptors regulate the trafficking of intracellular and extracellular cargo. Corresponding influence on lipid metabolism is mediated by their dynamic interaction with scavenger membrane receptors and their integrated mechanisms such as pinocytosis, phagocytosis, cholesterol export/import, etc. This interaction not only results in the functional differentiation of macrophages but also modifies their structural configurations. Here, we reviewed the association of macrophage membrane biomechanics and their scavenger receptor families with lipid metabolites during the event of atherogenesis. In addition, the membrane structure of macrophages and the signalling pathways involved in endocytosis integrated with lipid metabolism are detailed. This article establishes future insights into the scavenger receptors as potential targets for cardiovascular disease prevention and treatment.

Sitosterol-rich Digera muricata against 7-ketocholesterol and lipopolysaccharide-mediated atherogenic responses by modulating NF-ΚB/iNOS signalling pathway in macrophages.

Digera muricata L., commonly known as Tartara, is an edible herb used as traditional medicine in many countries of Africa and Asia. This study aimed to elucidate the effect of a phytosterol-rich extract of D. muricata on 7-ketocholesterol-mediated atherosclerosis in macrophages. The extract was examined by phytochemical analyses, GC-MS, TLC, DPPH scavenging and hRBC membrane stabilization assays. Macrophage polarization was studied with experimental groups framed based on alamar blue cell viability and griess assays. Regulations of arginase enzyme activity, ROS generation, mitochondrial membrane potential, cell membrane integrity, pinocytosis, lipid uptake and peroxidation, as well as, intracellular calcium deposition were determined. In addition, expressions of atherogenic mediators were analysed using PCR, ELISA and immunocytochemistry techniques. Diverse phytochemicals with higher free radical scavenging activity and anti-inflammatory potential have been detected in the D. muricata. Co-treatment with D. muricata markedly reduced the atherogenic responses induced by 7KCh in the presence of LPS such as ROS, especially, NO and O2- along with lipid peroxidation. Furthermore, D. muricata significantly normalized mitochondrial membrane potential, cell membrane integrity, pinocytic activity, intracellular lipid accumulation and calcium deposition. These results provided us with the potentiality of D. muricata in ameliorating atherogenesis. Additionally, it decreased the expression of pro-atherogenic mediators (iNOS, COX-2, MMP9, IL-6, IL-1ß, CD36, CD163 and TGFß1) and increased anti-atherogenic mediators (MRC1 and PPARγ) with high cellular expressions of NF-κB and iNOS. Results showed the potential of sitosterol-rich D. muricata as a versatile biomedical therapeutic agent against abnormal macrophage polarization and its associated pathologies.

Dynamic Role of Macrophage Sub Types on Development of Atherosclerosis and Potential Use of Herbal Immunomodulators as Imminent Therapeutic Strategy.

Atherosclerosis, a major contributor to cardiovascular disease, is a global alarm causing mortality worldwide. Being a progressive disease in the arteries, it mainly causes the recruitment of monocytes to the inflammatory sites and subsides pathological conditions. Monocyte-derived macrophage mainly acts in foam cell formation by engorging the LDL molecules, oxidizes it into Ox-LDL and leads to plaque deposit development. Macrophages in general differentiate, proliferate and undergo apoptosis at the inflammatory site. Frequently two subtypes of macrophages M1 and M2 have to act crucially in balancing the micro-environmental conditions of endothelial cells in arteries. The productions of pro-inflammatory mediators like IL-1, IL-6, TNF-α by M1 macrophage have atherogenic properties majorly produced during the early progression of atherosclerotic plaques. To counteract cytokine productions and M1-M2 balance, secondary metabolites (phytochemicals) from plants act as a therapeutic agent in alleviating atherosclerosis progression. This review summarizes the fundamental role of the macrophage in atherosclerotic lesion formation along with its plasticity characteristic as well as recent therapeutic strategies using herbal components and anti-inflammatory cytokines as potential immunomodulators.

An insight on 7- ketocholesterol mediated inflammation in atherosclerosis and potential therapeutics.

7-ketocholesterol, a toxic oxidative product of oxysterol is a causative agent of several diseases and disabilities concomitant to aging including cardiovascular diseases like atherosclerosis. Auto-oxidation of cholesterol esters present in low-density lipoprotein (LDL) deposits lead to the formation of oxidized LDL (Ox-LDL) along with its byproducts, namely 7KCh. It is predominantly found in atherosclerotic plaque and also found to be more atherogenic than cholesterol by being cytotoxic, interfering with cellular homeostasis. This makes it a serious threat by being the foremost cause of morbidity and mortality worldwide and is likely to become more serious during forth coming years. It involves in mediating inflammatory mechanisms characterized by the advancement of fibroatheroma plaques. The atherosclerotic lesion is composed of Ox-LDL along with fibrotic mass consisting of immune cells and molecules. Macrophages being the specialized phagocytic cells, contribute to removal of detrimental contents of the lesion along with accumulated lipids leading to alteration of its biology and functionality due to its plasticity. Here, we have explored the known as well as proposed mechanisms involved with 7KCh associated atherogenesis along with potential therapeutic strategies for targeting 7KCh as a diagnostic and target in medicine.

Amarogentin, a secoiridoid glycoside, activates AMP- activated protein kinase (AMPK) to exert beneficial vasculo-metabolic effects.

INTRODUCTION: AMP-activated protein kinase (AMPK) is a drug target for treatment of metabolic and cardiovascular complications. Extracts of Gentianaceace plants exhibit anti-diabetic and anti-atherosclerotic effects, however, whether their phyto-constitutents activate AMPK remains to be determined. METHODS: Molecular docking of Gentiana lutea constituents was performed with crystal structure of human α2ß1γ1 trimeric AMPK (PDB ID: 4CFE). Binding of Amarogentin (AG) to α2 subunit was confirmed through isothermal titration calorimetry (ITC) and in vitro kinase assays were performed. L6 myotube, HUH7 and endothelial cell cultures were employed to validate in silico and in vitro observations. Lipid lowering and anti-atherosclerotic effects were confirmed in streptozotocin induced diabetic mice via biochemical measurements and through heamatoxylin and eosin, Masson's trichrome and Oil Red O staining. RESULTS: AG interacts with the α2 subunit of AMPK and activates the trimeric kinase with an EC50 value of 277 pM. In cell culture experiments, AG induced phosphorylation of AMPK as well as its downstream targets, acetyl-coA-carboxylase (ACC) and endothelial nitric oxide synthase (eNOS). Additionally, it enhanced glucose uptake in myotubes and blocked TNF-α induced endothelial inflammation. Oral supplementation of AG significantly attenuated diabetes-mediated neointimal thickening, and collagen and lipid deposition in the aorta. It also improved circulating levels of lipids and liver function in diabetic mice. CONCLUSION: In conclusion, AG exerts beneficial vasculo-metabolic effects by activating AMPK. GENERAL SIGNIFICANCE: Amarogentin, a naturally occurring secoiridoid glycoside, is a promising lead for design and synthesis of novel drugs for treatment and management of dyslipidemia and cardiovascular diseases.

Galangin, a dietary flavonol inhibits tumor initiation during experimental pulmonary tumorigenesis by modulating xenobiotic enzymes and antioxidant status.

The aim of present study was to elucidate anti-initiating efficacy of galangin against benzo(a)pyrene (B(a)P)-induced lung carcinogenesis in male Swiss albino mice. Therefore, the activities of xenobiotic metabolic enzymes such as phase I and II were examined in lung as well as liver tissues (to compare the effects between target and non-target organs). Besides, the activities/levels of tissue marker enzymes, antioxidants, lipid peroxidation (LPO), cytochrome P450 1A1 (CYP1A1) expressions and histological observation of lungs were also analyzed. B(a)P (50 mg/kg body weight) was administered to male Swiss albino mice (20-25 g) to experimentally induce lung cancer. B(a)P-induced animals showed increased activity of phase I (Cytochrome P450, Cytochrome b5, NADPH Cytochrome P450 redcutase and NADH Cytochrome b5 reductase) drug metabolic enzymes, LPO levels, tissue marker enzymes and decreased activity of phase II metabolic enzymes (glutathione-S-transferase, DT-diaphorase and UDP-glucuronyl transferase) as well as antioxidant levels. Histological examination of lungs revealed severe alveolar and bronchiolar damages in B(a)P-induced mice. Immunohistochemical and western blot analysis of CYP1A1 increased significantly in lung tissues of B(a)P-induced animals. Treatment with galangin (20 mg/kg body weight) efficiently counteracted all the above anomalies and restored cellular homeostasis. Our results demonstrate that galangin can modify xenobiotic enzymes in murine model of pulmonary tumorigenesis.

ASK2 Bioactive Compound Inhibits MDR Klebsiella pneumoniae by Antibiofilm Activity, Modulating Macrophage Cytokines and Opsonophagocytosis.

The emergence and spread of pathogens harboring extended spectrum beta-lactamase (ESBL) like carbapenem resistant Gram negative bacteria are the major emerging threat to public health. Of particular concern Klebsiella pneumoniae carbapenamase- producing strains have been recorded worldwide. Catheter associated urinary tract infections (CAUTI) caused by K. pneumoniae are significantly associated with morbidity and mortality. Hence the present work was aimed to develop a strategy for addressing these issues through an innovative approach of antibiofilm and immunomodulation. These two independent activities were analyzed in a Streptomyces derived ASK2 bioactive compound. While analysing the effect of sub-minimum inhibitory concentrations (sub-MICs), 0.5x of Minimum Inhibitory Concentration (MIC) was found to be more effective in preventing biofilm formation on coverslip and silicone catheter. The minimum biofilm eradication concentration (MBEC) was found to be 15-fold higher MIC with eradication of 75% of 3 day old biofilm. Apart from its antibiofilm potential, ASK2 also acts as an opsonin and enhances phagocytic response of macrophages against multidrug resistant K. pneumoniae. In addition, ASK2 resulted in elevated levels of nitric oxide generation by the macrophages and has a stimulating effect on IL-12, IFN-γ, and TNF-α proinflammatory cytokines. The opsonic role of ASK2 and its potential in modulating proinflammatory cytokines secreted by macrophages implies the importance of ASK2 in modulating cellular immune response of macrophages against MDR K. pneumoniae. The present study proposes ASK2 as a promising candidate for treating MDR K. pneumoniae infections with its dual properties of antibiofilm and immunomodulatory activities.

Purification and characterisation of a pronase-inducible lectin isolated from human serum.

A new lectin was purified to electrophoretic homogeneity from pronase treated human serum by a single-step of affinity chromatography on concanavalin A-Sepharose 4B. The isolated lectin agglutinated five types of vertebrate RBC, with highest titer against hen RBC. This activity was independent of divalent cations, insensitive to EDTA and specific to mannosamine, glucosamine as well as galactosamine. Purified lectin gave a single symmetrical peak in its native form with a molecular mass estimate of 6kDa in FPLC analysis and 6.5kDa by MALDI-TOF MS. SDS-PAGE analysis of the lectin revealed that it is a homo-oligomer of a 3kDa subunit protein. Isolated lectin did possess both, hemagglutinating and phenoloxidase activities, but did not exhibit any antibacterial or antifungal activities. In addition, this lectin could oxidize all nine different phenolic substrates tested, with hydroquinone proving to be the best among them. Phenoloxidase inhibitors namely, phenylthiourea and tropolone inhibited this oxidation activity.

Cytoprotective mechanism of action of curcumin against cataract.

This review discusses the relationship between oxidative stress and cataract formation, molecular mechanism of curcumin action and potential benefits of treatment with the antioxidant curcumin. The first section deals with curcumin and endogenous antioxidants. The second section focuses on the action of curcumin on lipid peroxidation. Calcium homeostasis and curcumin will be discussed in the third section. The fourth section discusses the role of crystallin proteins that are responsible for maintaining lens transparency and the role of curcumin in regulating crystallin expression. The interaction of curcumin with transcription factors will be dealt in the fifth section. The final section will focus on the effect of curcumin on aldose reductase, which is associated with hyperglycemia and cataract. One of the strongest antioxidants is curcumin which has been shown to be very effective against cataract. This compound is better than other antioxidants in preventing cataract but its limited bioavailability can be addressed by employing nanotechnology.

Synthesis of silver nanoparticles using Solanum trilobatum fruits extract and its antibacterial, cytotoxic activity against human breast cancer cell line MCF 7.

In the present study, we have synthesized silver nanoparticles by a simple and eco-friendly method using unripe fruits of Solanum trilobatum. The aqueous silver ions when exposed to unripe fruits extract were reduced and stabilized over long time resulting in biosynthesis of surface functionalized silver nanoparticles. The bio-reduced silver nanoparticles were characterized by UV-visible spectroscopy, Fourier transform infrared spectroscopy (FTIR), scanning electron microscopy (SEM), transmission electron microscopy (TEM), energy-dispersive spectroscopy (EDX) and X-ray diffraction (XRD). These biologically synthesized silver nanoparticles were tested for its antibacterial activity against few human pathogenic bacteria including Gram-positive (Streptococcus mutans, Enterococcus faecalis) and Gram-negative (Escherichia coli, Klebsiella pneumoniae) bacteria. In addition, we also demonstrated anticancer activity of these nanoparticles in vitro against human breast cancer cell line (MCF 7) using MTT, nuclear morphology assay, Western blot and RT-PCR expression. These results taken together show the potential applications of biosynthesized silver nanoparticles using S. trilobatum fruits.

Cytoprotective and anti-inflammatory effects of kernel extract from adenanthera pavonina on lipopolysaccharide-stimulated rat peritoneal macrophages

Objective: To investigate mechanism of anti-inflammatory activity of Adenanthera pavonina (A. pavonina) extracts. Methods: Rat peritoneal macrophages were treated with different concentrations of lipopolysaccharide and H2O2 in the presence and absence of kernel extract from A. pavonina. Nitric oxide, Superoxide anion generation, cell viability and nuclear fragmentation were investigated. Results: The pre-treatment of kernel extract from A. pavonina suppressed nitric oxide, superoxide anion, cell death, nuclear fragmentation in lipopolysaccharide and H2O2 stimulated or induced macrophages, respectively. Conclusions: These results suggest that A. pavonina extract suppresses the intra cellular peroxide production.

Cytoprotective and anti-inflammatory effects of kernel extract from Adenanthera pavonina on lipopolysaccharide-stimulated rat peritoneal macrophages

OBJECTIVE@#To investigate mechanism of anti-inflammatory activity of Adenanthera pavonina (A. pavonina) extracts.@*METHODS@#Rat peritoneal macrophages were treated with different concentrations of lipopolysaccharide and H2O2 in the presence and absence of kernel extract from A. pavonina. Nitric oxide, Superoxide anion generation, cell viability and nuclear fragmentation were investigated.@*RESULTS@#The pre-treatment of kernel extract from A. pavonina suppressed nitric oxide, superoxide anion, cell death, nuclear fragmentation in lipopolysaccharide and H2O2 stimulated or induced macrophages, respectively.@*CONCLUSIONS@#These results suggest that A. pavonina extract suppresses the intra cellular peroxide production.

Synthesis and characterization of silver nanoparticles using crystal compound of sodium para-hydroxybenzoate tetrahydrate isolated from Vitex negundo. L leaves and its apoptotic effect on human colon cancer cell lines.

Metallic nanoparticles are major concern, particularly silver nanoparticles (AgNPs) are used in various applications. In the present investigation, we report a novel strategy with biological approach for synthesis of AgNPs using sodium para-hydroxybenzoate tetrahydrate (SPHT) isolated from Vitex negundo leaves. The synthesized SPHT-AgNPs were characterized by UV-vis spectroscopy, high resolution transmission electron microscopy (HRTEM) with selected area electron diffraction (SAED) pattern, field emission scanning electron microscopy (FESEM) with energy-dispersive X-ray spectroscopy (EDX), zeta potential and Fourier transform infrared spectroscopy (FT-IR) analysis. The various pH and temperature were evaluated to find their stability effects on SPHT-AgNPs synthesis peak at 430 nm. The size of SPHT-AgNPs were ranging from 26 to 39 nm and were spherical in shape. The hydroxyl and carboxylic functional groups from bio-reducing mediators of SPHT have a stronger ability towards synthesis of AgNPs, which was confirmed using FT-IR spectrum. In addition, anticancer activity were determined by MTT assay, Annexin V-FITC/PI and cell cycle analysis.

Identification of serum component involved in generation of neo-lectin with agglutinating and phenoloxidase activities in human serum.

Human serum albumin (HSA) was identified as the component involved in generation of neo-lectin molecules with both lectin and phenoloxidase activities. Pronase treated HSA was able to agglutinate hen RBC and oxidize hydroquinone. Sodium dodecyl sulphate (SDS) treated HSA agglutinated both hen and sheep RBC as well as oxidized dopamine. The hemagglutinating activities of pronase/SDS treated HSA observed against hen RBC were dosimetric. The oxidation of pronase/SDS treated HSA with hydroquinone/dopamine, respectively, was inhibitable by inhibitors of phenoloxidase, namely, phenylthiourea and tropolone. Very low concentrations of HSA could generate these humoral neo-lectin molecules.

Luteolin induces growth arrest in colon cancer cells through involvement of Wnt/ß-catenin/GSK-3ß signaling.

Cancer is a multistep process that typically occurrs over an extended period of time, beginning with initiation followed by promotion and progression. Colon cancer is the leading cause of morbidity and mortality worldwide. For a variety of reasons, patients prefer naturally occurring dietary substances over synthetic agents to prevent cancer. Luteolin, a bioflavonoid, possesses antioxidant, anti-inflammatory, and antiproliferative effects. We analyzed the in vitro anticancer and apoptosis-inducing property of luteolin using HCT-15 colon adenocarcinoma cells. Cell viability was assessed using trypan blue assay at different concentrations. Luteolin at a concentration of 100 µM (IC50) decreased the expressions of non-P-ß-catenin, phosphorylated (inactive) glycogen synthase kinase-3ß, and cyclin D1 expressions in HCT-15 cells, which were confirmed by Western blot analysis. Luteolin also promoted substantial cell cycle arrest at the G2/M phase in HCT-15 cells, and it induces apoptosis in HCT-15 cells, as revealed by flow cytometric analysis. Furthermore, Western blot analysis showed that luteolin treatment enhanced the expression of Bax and caspase-3, whereas the expression of Bcl-2 was suppressed. Together, the results of this study revealed that luteolin can act as a potent inhibitor of HCT-15 proliferation and can be used as an agent against colon cancer.

Detection of natural and induced phenoloxidase activities in human serum.

Natural and induced phenoloxidase activities were detected in human serum using nine different phenolic substrates, namely, tyrosine, tyramine, L-DOPA, DL-DOPA, dopamine, catechol, hydroquinone, protocatechuic acid and pyrogallol. Phenoloxidase activity was induced anew in serum using exogenous elicitors, such as proteases or detergents. Among the proteases and detergents tested, pronase, SDS and Tween 20 were the best elicitors of phenoloxidase activities in serum, wherein, hydroquinone was the best phenolic substrate for both untreated as well as pronase treated serum and SDS or Tween 20 treated serum resulted in highest oxidation of dopamine or tyrosine, respectively. In the present study, all these oxidative reactions were inhibited by phenoloxidase inhibitors, namely, PTU and tropolone, thereby, confirming the role of phenoloxidase in human serum.

Protective effect of ferulic acid and resveratrol against alloxan-induced diabetes in mice.

The present study was to investigate the effect of ferulic acid and resveratrol on alloxan-induced diabetic mice, through analysis of basic biochemical parameters, enzymic as well as non-enzymic activities, lipid peroxidation and immunohistochemical studies. Alloxan was administered as a single dose (75 mg/kg body weight) to induce diabetes in mice. A dose of ferulic acid (10 mg/kg body weight) and resveratrol (20 mg/kg body weight) were administrated orally, to the alloxan-induced diabetic mice. The levels of basic biochemical markers and lipid peroxidation were significantly (P<0.05) increased in alloxan-induced diabetic mice. The levels of antioxidants were significantly (P<0.05) decreased in liver, kidney and serum. Immunohistochemical studies in alloxan induced mice demonstrated a marked increase in the immunoreactivity of nuclear transcription factor (NF-κB). Treating the diabetic mice with doses of ferulic acid and resveratrol restored the changes in the above parameters analyzed. The present study, showed that ferulic acid and resveratrol exerted antioxidant as well as anti-diabetic effects, consequently alleviate liver, kidney and pancreas damage caused by alloxan-induced diabetes, probably through inhibition of the proinflammatory factor, NF-κB.

Detection and characterization of natural and inducible lectins in human serum.

This study was performed to detect and characterise the possible occurrence of natural and inducible lectins in human serum by hemagglutination method, wherein, the serum was treated using exogenous elicitors, namely, proteases and detergents. Natural and inducible lectins were detected and characterised in human serum. Untreated serum agglutinated buffalo and rabbit RBC, while serum treated with pronase, trypsin, α-chymotrypsin or SDS for the very first time, agglutinated hen/hen and sheep RBC within 15 min in a dosimetric manner. Cross adsorption test revealed that both trypsin and α-chymotrypsin-treated serum showed similar RBC adsorption pattern. The lectin activity in untreated, pronase-treated serum was cation independent and moderately sensitive/insensitive to calcium chelator EDTA, whereas, trypsin-treated serum was cation dependent as well as EDTA sensitive (sheep RBC), cation independent and EDTA insensitive (hen RBC). Hemagglutination of untreated serum was inhibited by certain glycosides and di-, oligo-saccharides, whereas, activity in pronase-treated serum was inhibited by hexosamines. By contrast, hemagglutination of trypsin-treated serum showed specificity for acetylated mannosamine as well as sialic acid for sheep RBC and certain glycoproteins for hen RBC. Thus, we have detected inducible lectins with distinct ligand binding specificity, upon treatment of human serum with proteases, namely, pronase and trypsin. Nevertheless, lectin activity was found in untreated human serum too with different ligand specificity.