Dehydrozingerone ameliorates Lipopolysaccharide induced acute respiratory distress syndrome by inhibiting cytokine storm, oxidative stress via modulating the MAPK/NF-κB pathway.

BACKGROUND: Inflammation-mediated lung injury is a major cause of health problems in many countries and has been the leading cause of morbidity/mortality in intensive care units. In the current COVID-19 pandemic, the majority of the patients experienced serious pneumonia resulting from inflammation (Acute respiratory distress syndrome/ARDS). Pathogenic infections cause cytokine release syndrome (CRS) by hyperactivation of immune cells, which in turn release excessive cytokines causing ARDS. Currently, there are no standard therapies for viral, bacterial or pathogen-mediated CRS. PURPOSE: This study aimed to investigate and validate the protective effects of Dehydrozingerone (DHZ) against LPS induced lung cell injury by in-vitro and in-vivo models and to gain insights into the molecular mechanisms that mediate these therapeutic effects. METHODS: The therapeutic activity of DHZ was determined in in-vitro models by pre-treating the cells with DHZ and exposed to LPS to stimulate the inflammatory cascade of events. We analysed the effect of DHZ on LPS induced inflammatory cytokines, chemokines and cell damage markers expression/levels using various cell lines. We performed gene expression, ELISA, and western blot analysis to elucidate the effect of DHZ on inflammation and its modulation of MAPK and NF-κB pathways. Further, the prophylactic and therapeutic effect of DHZ was evaluated against the LPS induced ARDS model in rats. RESULTS: DHZ significantly (p < 0.01) attenuated the LPS induced ROS, inflammatory cytokine, chemokine gene expression and protein release in macrophages. Similarly, DHZ treatment protected the lung epithelial and endothelial cells by mitigating the LPS induced inflammatory events in a dose-dependent manner. In vivo analysis showed that DHZ treatment significantly (p < 0.001) mitigated the LPS induced ARDS pathophysiology of increase in the inflammatory cells in BALF, inflammatory cytokine and chemokines in lung tissues. LPS stimulated neutrophil-mediated events, apoptosis, alveolar wall thickening and alveolar inflammation were profoundly reduced by DHZ treatment in a rat model. CONCLUSION: This study demonstrates for the first time that DHZ has the potential to ameliorate LPS induced ARDS by inhibiting cytokine storm and oxidative through modulating the MAPK and NF-κB pathways. This data provides pre-clinical support to develop DHZ as a potential therapeutic agent against ARDS.

Supplementation of fenugreek with choline-docosahexaenoic acid attenuates menopause induced memory loss, BDNF and dendritic arborization in ovariectomized rats.

Cognitive impairment due to natural or surgical menopause is always associated with estrogen deficiency leading to reduced brain-derived neurotrophic factor (BDNF). Reduced BDNF levels in menopause affect neuronal maturation, survival, axonal and dendritic arborization and the maintenance of dendritic spine density. Conventional long-term estrogen replacement therapy reported causing the risk of venous thromboembolism and breast cancer. To overcome these undesirable effects, phytoestrogens have been used in menopause-induced condition without the risk of side effects. Therefore, the aim of the present study was to investigate the effect of dietary supplementation of fenugreek seed extract (FG) either alone or in combination with choline-DHA on BDNF and dendritic arborization of pyramidal neurons in CA1 and CA3 regions of the hippocampus in ovariectomized rats. Female Wistar rats of 9-10 months old were divided into six groups as normal control (NC); ovariectomy (OVX); OVX + FG; OVX + choline-DHA; OVX + FG + choline-DHA; and OVX + estradiol. All the groups, except NC, were ovariectomized. After 2 weeks of ovariectomy, dietary supplementation was initiated for a period of 30 days. After supplementation, behavioral studies, BDNF levels and dendritic arborization were estimated. Ovariectomized (OVX) rats showed reduced BDNF levels, dendritic branching points and dendritic intersections of pyramidal neurons in CA1 and CA3 regions of the hippocampus. OVX rats supplemented with FG with choline-DHA showed significantly improved BDNF levels, dendritic branching points and dendritic intersections. These results are demonstrating that FG with choline-DHA supplementation can be an alternative for estrogen replacement therapy to modulate menopause-induced learning and memory deficits.

Biochanin-A ameliorates pulmonary fibrosis by suppressing the TGF-ß mediated EMT, myofibroblasts differentiation and collagen deposition in in vitro and in vivo systems.

BACKGROUND: Idiopathic Pulmonary Fibrosis (IPF) is a progressive inflammatory disorder driven by a fibrotic cascade of events such as epithelial to mesenchymal transition, extracellular matrix production and collagen formation in the lungs in a sequential manner. IPF incidences were raising rapidly across the world. FDA approved pirfenidone and nintedanib (tyrosine kinase inhibitors) are being used as a first-line treatment drugs for IPF, however, neither the quality of life nor survival rates have been improved because of patient noncompliance due to multiple side effects. Thus, the development of novel therapeutic approaches targeting TGF-ß mediated cascade of fibrotic events is urgently needed to improve the survival of the patients suffering from devastating disease. PURPOSE: The aim of this study was to investigate and validate the anti-fibrotic properties of Biochanin-A (isoflavone) against TGF-ß mediated fibrosis in in vitro, ex vivo, in vivo models and to determine the molecular mechanisms that mediate these anti-fibrotic effects. METHODS: The therapeutic activity of BCA was determined in in vitro/ex vivo models. Cells were pre-treated with BCA and incubated in presence or absence of recombinant-TGF-ß to stimulate the fibrotic cascade of events. Pulmonary fibrosis was developed by intratracheal administration of bleomycin in rats. BCA treatment was given for 14 days from post bleomycin instillation and then various investigations (collagen content, fibrosis gene/protein expression and histopathological changes) were performed to assess the anti-fibrotic activity of BCA. RESULTS: In vitro/ex vivo (Primary normal, IPF cell line and primary IPF cells/ Precision cut mouse lung slices) experiments revealed that, BCA treatment significantly (p < 0.001) reduced the expression of TGF-ß modulated fibrotic genes/protein expressions (including their functions) which are involved in the cascade of fibrotic events. BCA treatment significantly (p < 0.01) reduced the bleomycin-induced inflammatory cell-infiltration, inflammatory markers expression, collagen deposition and expression of fibrotic markers in lung tissues equivalent or better than pirfenidone treatment. In addition, BCA treatment significantly (p < 0.001) attenuated the TGF-ß1/BLM-mediated increase of TGF-ß/Smad2/3 phosphorylation and resulted in the reduction of pathological abnormalities in lung tissues determined by histopathology observations. CONCLUSION: Collectively, BCA treatment demonstrated the remarkable therapeutic effects on TGF-ß/BLM mediated pulmonary fibrosis using IPF cells and rodent models. This current study may offer a novel treatment approach to halt and may be even rescue the devastating lung scarring of IPF.

Evaluation of the aphrodisiac potential of a chemically characterized aqueous extract of Tamarindus indica pulp.

ETHNOPHARMACOLOGICAL RELEVANCE: Tamarindus indica is an ingredient in the traditional aphrodisiac formulations in Africa and India. It is also a widely used food ingredient in other tropical countries. AIM OF THE STUDY: The present study was aimed to evaluate the aphrodisiac potential and reproductive safety profile of aqueous extract of Tamarindus indica in male Wistar rats. MATERIALS AND METHODS: The aqueous extract was prepared by maceration of pulp followed by reduction of volume in rotavapor under heat followed by freeze drying. The prepared extract was characterized for contents of total phenol, flavonoid, and saponin. It was also subjected to phytoconstituent analysis using GCMS. Further, the extract was evaluated for acute toxicity study. The aphrodisiac and reproductive toxicity potential were evaluated in animals after grouping them in four with six animals each namely, normal control, standard (Sildenafil citrate, 4mg/kg p.o.) and extract of Tamarindus indica treated groups at two dose levels, 125 and 250mg/kg p.o. The study was conducted for 54 days with daily once dosing of extract and standard. Equal number of females was grouped without treatment for evaluation of parameters of sexual desire (mount frequency and intromission frequency) and parameters of sexual arousal (mount latency and intromission latency). These parameters were evaluated on day 14, 28, 42 and 54. Animals were sacrificed on day 54, testes were removed and studied for histopathological changes. RESULTS: The extract showed 6.6mg gallic acid equivalent/g of total phenol, 2.3mg catechin equivalent/g of flavonoid and 11.6% saponin. Forty chemical constituents were identified by GCMS analysis. In acute toxicity study, the extract was found to be safe till 2000mg/kg p.o. Efficacy study showed significant (p<0.05) improvement in parameters of sexual desire (mount frequency and intromission frequency) and parameters of sexual arousal on all observed days except mount frequency for 125mg/kg on 42nd day and intromission frequency for both doses of tamarind compared to normal control. Improvements in these parameters were comparable to the standard drug. Histopathology study and sperm count suggested an increase in sperm production without any sign of toxicity in testis. Sperm motility significantly (p<0.05) increased in the treatment groups that received extract at 250mg/kg compared to normal control. CONCLUSION: Aqueous extract of Tamarindus indica possessed aphrodisiac activity together with spermatogenic potential.

Biological activity of a small molecule indole analog, 1-[(1H-indol-3-yl)methylene]-2-phenylhydrazine (HMPH), in chronic inflammation.

A synthetic small molecule, 1-[(1H-indol-3-yl)methylene]-2-phenylhydrazine (HMPH) was conveniently synthesised by a one-step reaction, purified and characterised by chromatographic and spectroscopic methods. HMPH scavenged free radicals and inhibited lipopolysaccharide (LPS)-induced ROS generation and NO release in RAW-264.7 cells without signs of any detectable cytotoxicity. HMPH inhibited lipid peroxidation (LPO) with IC50 of 135 ± 9 as against 58 ± 8 µM for α-tocopherol. Further, HMPH (>50 µM) significantly reduced the LPS-induced TNF-α release in mouse peritoneal macrophages and in human peripheral blood mononuclear cells (PBMCs). HMPH did not show any visible signs of toxicity in rats up to 400 mg/kg/intraperitoneal and 2000 mg/kg/oral. HMPH at 25 and 50 mg/kg attenuated neutrophil infiltration in air-pouch lavage and bronchoalveolar lavage (BAL) in rat models. HMPH also reduced myeloperoxidase (MPO), nitrite and TNF-α in air-pouch lavage in addition to MPO in plasma. HMPH reduced acute paw-inflammation in carrageenan-induced paw-edema. HMPH consistently decreased both ipsilateral and contralateral paw inflammation, minimised the clinical scores of arthritis, prevented body weight (B.wt.) loss, attenuated serum C-reactive protein (C-RP) and rheumatoid factors (RF) in rat model of adjuvant-induced arthritis. Histopathology and radio-graphical reports show that HMPH reduced bone erosion in both ipsilateral and contralateral paw joints. Failure to inhibit COX suggests that effectiveness of HMPH in both acute and chronic inflammation is mediated by a multimodal mechanism involving modulation of immunity, attenuating TNF-α, protecting bone attrition and reducing oxidative stress.

Assessment of the in vitro cytotoxicity and in vivo anti-tumor activity of the alcoholic stem bark extract/fractions of Mimusops elengi Linn.

Various parts of Mimusops elengi Linn. (Sapotaceae) have been used widely in traditional Indian medicine for the treatment of pain, inflammation and wounds. The study was conducted to explore the use of stem bark of M. elengi on pharmacological grounds and to evaluate the scientific basis of cytotoxic and anti-tumor activity. Extract/fractions were prepared and in vitro cytotoxicity was assessed using SRB assay. Most effective fractions were subjected to fluorescence microscopy based acridine orange/ethidium bromide (AO/EB) and Hoechst 33342 staining to determine apoptosis induction and DNA fragmentation assay. Comet and micronuclei assay were performed to assess genotoxicity. Cell cycle analysis was also performed. In vivo anti-tumor potential was evaluated by Ehrlich ascites carcinoma (EAC) model in mice. The alcoholic stem bark extract of M. elengi along with four fractions showed potential in vitro cytotoxicity in SRB assay. Of these, dichloromethane and ethyl acetate fractions were selected for further studies. The fractions revealed apoptosis inducing potential in AO/EB and Hoechst 33342 staining, which was further confirmed by DNA fragmentation assay. Genotoxic potential was revealed by comet and micronuclei assay. Fractions also exhibited specific cell cycle inhibition in G0/G1 phase. In EAC model, ethyl acetate fraction along with the standard (cisplatin) effectively reduced the increase in body weight compared to control and improved mean survival time. Both fractions were able to restore the altered hematological and biochemical parameters. Hence, M. elengi stem bark may be a possible therapeutic candidate having cytotoxic and anti-tumor potential.