Dataset Information

Modulation of proinflammatory cytokines and enzymes by polyherbal formulation Guggulutiktaka ghritam.

ABSTRACT:

Background

Guggulutiktaka ghritam is an ayurvedic medicine which has been traditionally used to treat various chronic inflammatory conditions. However, the mechanism of action of the Ayurvedic medication in control of inflammatory conditions has not been clearly evaluated.

Objective

In the current study, the effect of the Guggulutiktaka ghritam extract (GTG) on the lipoxygenase pathway and in the production of proinflammatory cytokines involved in the pathogenesis of chronic inflammation was studied.

Materials and methods

The effect of GTG in the production of leukotriene was determined by enzyme inhibition studies on 12- lipoxygenase. The assay was carried out by ferrous oxidation of xylenol orange (FOX assay) and was compared to a positive control nordihydroguaiaretic acid. The effect of GTG on the production of proinflammatory cytokines TNF-α and IL-1β in monocytes were studied. For this, the monocytes were pretreated with various concentrations of GTG and subsequently stimulated with lipopolysaccharide. The cytokines TNF-α and IL-1β produced were quantified by ELISA and the results were compared to positive controls Rolipram and Dexamethasone respectively. The gene expression studies were carried out using qRT-PCR. The IC₅₀ values were calculated and evaluated statistically.

Results

The result indicates that GTG in comparison to the positive control Nordihydroguaiaretic acid significantly reduced the activity of 12- lipoxygenase. Also, there was significant inhibition in the production of proinflammatory cytokines in LPS stimulated monocytes pretreated with GTG as compared to positive control Rolipram and Dexamethasone. There was significant downregulation of IL-1β gene in LPS stimulated monocytes pretreated with GTG as compared to control. These changes are further supported by Raman spectra obtained for GTG treated and untreated cells.

Conclusion

The study revealed that GTG is a leukotriene and cytokine inhibitor. The inhibition in the production of cytokines may be due to the down-regulation of genes for TNF-α and IL-1β. The study provides a scientific validation on the possible anti-inflammatory mechanism of action of this traditionally used medicine. Identification of bioactive molecules would aid in developing newer therapeutics for control of chronic inflammation.

SUBMITTER: Mohan MC

PROVIDER: S-EPMC8039337 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Similar Datasets

Project description:ObjectiveThe normal structure and function of articular cartilage are the result of a precisely balanced interaction between anabolic and catabolic processes. The transforming growth factor-beta (TGF-beta) family of growth factors generally exerts an anabolic or repair response; in contrast, proinflammatory cytokines such as interleukin 1 beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) exert a strong catabolic effect. Recent evidence has shown that IL-1beta, and TNF-alpha, and the TGF-beta signaling pathways share an antagonistic relationship. The aim of this study was to determine whether the modulation of the response of articular chondrocytes to TGF-beta by IL-1beta or TNF-alpha signaling pathways occurs through regulation of activity and availability of mothers against DPP (Drosophila) human homologue (Smad) proteins.MethodsHuman articular chondrocytes isolated from knee joints from patients with osteoarthritis (OA) or normal bovine chondrocytes were cultured in suspension in poly-(2-hydroxyethyl methacrylate)-coated dishes with either 10% fetal bovine serum media or serum-deprived media 6h before treatment with IL-1beta alone, TNF-alpha alone or IL-1beta followed by TGF-beta. Nuclear extracts were examined by electrophoretic mobility-shift assays (EMSA) for nuclear factor-kappa B (NF-kappaB) and Smad3/4 deoxyribonucleic acid (DNA) binding. Nuclear extracts were also subjected to the TranSignal Protein/DNA array (Panomics, Redwood City, CA) enabling the simultaneous semiquantitative assessment of DNA-binding activity of 54 different transcription factors. Nuclear phospho-Smad2/3 and total Smad7 protein expression in whole cell lysates were studied by Western blot. Cytoplasmic Smad7, type II collagen alpha 1 (COL2A1), aggrecan and SRY-related high mobility group-Box gene 9 (SOX-9) mRNA expression were measured by real-time polymerase chain reaction (PCR).ResultsThe DNA-binding activity of Smad3/4 in the TranSignal Protein/DNA array was downregulated by TNF-alpha (46%) or IL-1beta treatment (42%). EMSA analysis showed a consistent reduction in Smad3/4 DNA-binding activity in human articular chondrocytes treated with IL-1beta or TNF-alpha. TGF-beta-induced Smad3/4 DNA-binding activity and Smad2/3 phosphorylation were also reduced following pretreatment with IL-1beta in human OA and bovine chondrocytes. Real-time PCR and Western blot analysis showed that IL-1beta partially reversed the TGF-beta stimulation of Smad7 mRNA and protein levels in TGF-beta-treated human OA cells. In contrast, TGF-beta-stimulated COL2A1, aggrecan, and SOX-9 mRNA levels were abrogated by IL-1beta.ConclusionsIL-1beta or TNF-alpha exerted a suppressive effect on Smad3/4 DNA-binding activity in human articular chondrocytes, as well as on TGF-beta-induced stimulation of Smad3/4 DNA-binding activity and Smad2/3 phosphorylation in human OA and bovine articular chondrocytes. IL-1beta partially reversed the increase in TGF-beta-stimulated Smad7 mRNA or protein levels suggesting that Smad7 may not be involved in the suppression of TGF-beta signaling induced by IL-1beta or TNF-alpha in articular chondrocytes. The balance between the IL-1beta or TNF-alpha and the TGF-beta signaling pathways is crucial for maintenance of articular cartilage homeostasis and its disruption likely plays a substantial role in the pathogenesis of OA.

Project description:BackgroundNayopayam kwatha (NK) is a well-known polyherbal formulation widely used to cure respiratory ailments, heart problems, and postnatal difficulties. Literature suggests that so far no standardization protocol was developed for NK to validate its quality and purity.Objective(s)To develop a standardization protocol for NK based on the marker phytoconstituents present in the individual herbs of the formulation.Materials and methodsThe roots of bala [Sida cordifolia (B1) and Sida retusa (B2)], seeds of jeeraka (Cuminum cyminum), and rhizomes of nagara (Zingiber officinale) were the ingredients of NK. Since there were two source plants for bala, two sets of NK (NKB1 and NKB2) were prepared in the ratio 3:2:1 as per Vaidya Manorama and 10:1:1 as per Arogyaraksha Kalpadruma along with 1:1:1 as per the general way of Ayurvedic polyherbal decoctions. Both the individual herbs and the kwatha (decoction) prepared were analyzed in terms of pharmacognostical, organoleptic, and physcicochemical parameters as per the standard methods. Phytochemical analysis of the individual herbs resulted in the isolation of major phytoconstituents and the kwatha was quantified in terms of marker compounds with the aid of HPLC.ResultsHPLC quantification suggests that appreciable amount of marker phytoconstituents of individual herbs are present in the kwatha. Thus, the isolated compounds luteolin (C. cyminum), 6-gingerol (Z. officinale), β-sitosterol (S. retusa), and ecdysterone (S.cordifolia) can be used as markers to standardize NK.ConclusionCharacteristics of NK, as well as its individual drugs, were well-established. The present study of NK with respect to its phytochemistry revealed that the classical drug ratios of the polyherbal formulation are of utmost importance rather than the ingredients in equal proportion. The characterization parameters of individual herbs and kwatha described in this study may serve as a standard reference for quality control analysis of NK and the method developed in this study can be used as a reliable technique for standardization of NK to ensure the purity and quality of raw drugs used.

Project description:Background: Diosgenin, charantin, and hydroxychalcone are utilized for standardization of popular antidiabetic herbal drugs Trigonella foenum-graecum L. belonging to family Fabaceae, Momordica charantia L. belonging to family Cucurbitaceae, and Cinnamomum verum J. Presl belonging to family Lauraceae. However, no reports on the bioavailability of these markers were available. The present study was undertaken to determine the bioavailability and pharmacokinetic profile of the markers and formulations containing the herbs. Methods: The pharmacokinetic profile and absolute bioavailability of the pure active markers were determined in male Wistar rats by administrating individually the doses of 1.5 mg/kg i.v. and 15 mg/kg p.o., followed by estimation of serum levels of the markers at 0, 10, 30, 60, 120, and 240 mins till 24 h time points by a validated bioanalytical HPTLC method. Two standardized antidiabetic capsule formulations containing spray dried hydroalcoholic extracts of seeds of Trigonella foenum-graecum L. (42.8 mg equivalent to 0.95%w/w of diosgenin), fresh fruits of Momordica charantia L. (21.4 mg equivalent to 0.4% w/w of charantin), and bark of Cinnamomum verum J. Presl (10.71 mg equivalent to 0.079 %w/w hydroxychalcone) were prepared. In one formulation, piperine 1.5 mg was added along with the other herbal extracts mentioned. Bioavailability and pharmacokinetic profile of these two formulations were determined in male Wistar rats through estimating serum levels of active markers diosgenin, charantin, and hydroxychalcone at 0, 10, 30, 60, 120, and 240 mins till 24 h later oral administration of the formulations (Formulation without piperine F1 and formulation with Piperine F2). Results: Plasma concentrations were found to decline mono-exponentially following intravenous administration, and the mean elimination half-life (t1/2) was observed to be 7.93, 8.21, and 4.66 h, respectively. The absolute oral bioavailability of pure markers was observed to be 9.0 ± 0.2%, 8.18 ± 0.36%, and 10.54 ± 0.52% by the dose normalization method. The oral bioavailabilities of the formulations with respect to diosgenin, charantin, and hydroxychalcone were found to be 9.78, 10.743, and 8.07%, respectively. The formulation containing piperine indicated a significant (p < 0.01) increase in the bioavailabilities of all the marker compounds. Conclusion: In conclusion, diosgenin and charantin have low bioavailabilities as compared to hydroxychalcone. The bioavailabilities of all the three marker compounds can be increased exponentially with the addition of piperine.