Biochanin A mitigates ulcerative colitis and intestinal inflammation in mice by inhibiting MAPK/NF-kB (p65) axis.

Ulcerative colitis (UC) is a chronic problem of the intestine and relapsing in nature. Biochanin A is a nature-derived isoflavonoid and has numerous bioactivities. However, its role against UC and intestinal inflammation remains obscure. We aimed to comprehensively explore the pharmacological effect of biochanin A in alleviating colitis and to evaluate the potential mechanisms. Initially, we explored the anti-inflammatory action of biochanin A (15, 30, and 60 µM) by employing lipopolysaccharide (LPS)-activated RAW 264.7 cells. In RAW 264.7 cells under LPS stimulation, biochanin A inhibited the elevation of reactive oxygen species (ROS) (p < 0.0001), interleukin (IL)-1ß (p < 0.0001), IL-18 (p < 0.01), and tumor necrosis factor (TNF)-α (p < 0.01) release, nitrite production (p < 0.0001), and the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) proteins. Next, we studied the effectiveness of biochanin A (20 and 40 mg/kg) in mouse colitis induced with dextran sulfate sodium (DSS) by assessing colon length, disease activity index (DAI) scoring, and performing colonoscopy and histological analysis. The pro-inflammatory cytokines were estimated using ELISA. Western blot studies were performed to assess underlying mechanisms. In mice, biochanin A treatment alleviated DAI score (p < 0.0001), restored colon length (p < 0.05) and morphology, and re-established colon histopathology. Biochanin A affects the phosphorylation of proteins associated with NF-κB (p65) and mitogen-activated protein kinase (MAPK) axis and regulates colonic inflammation by reducing the expression of inflammatory cytokines and myeloperoxidase (MPO) activity. Altogether, our findings support the idea that the anticolitis potential of biochanin A is allied with anti-inflammatory activity by inhibiting the MAPK/NF-κB (p65) axis. Hence, biochanin A may be an alternative option to alleviate the risk of colitis.

Chronic kidney disease activates the HDAC6-inflammatory axis in the heart and contributes to myocardial remodeling in mice: inhibition of HDAC6 alleviates chronic kidney disease-induced myocardial remodeling.

Chronic kidney disease (CKD) adversely affects the heart. The underlying mechanism and the interplay between the kidney and the heart are still obscure. We examined the cardiac effect using the unilateral ureteral obstruction (UUO)-induced CKD pre-clinical model in mice. Echocardiography, histopathology of the heart, myocardial mRNA expression of ANP and BNP, the extent of fibrotic (TGF-ß, α-SMA, and collagen I) and epigenetic (histone deacetylases, namely HDAC3, HDAC4, and HDAC6) proteins, and myocardial inflammatory response were assessed. Six weeks of post-UUO surgery, we observed a compromised left-ventricular wall thickness and signs of cardiac hypertrophy, accumulation of fibrosis associated, and inflammatory proteins in the heart. In addition, we observed a perturbation of epigenetic proteins, especially HDAC3, HDAC4, and HDAC6, in the heart. Pharmacological inhibition of HDAC6 using ricolinostat (RIC) lessened cardiac damage and improved left-ventricular wall thickness. The RIC treatment substantially restored the serum cardiac injury markers, namely creatine kinase-MB and lactate dehydrogenase (LDH) activities, ANP and BNP mRNA expression, and heart histological changes. The extent of myocardial fibrotic proteins, phospho-NF-κB (p65), and pro-inflammatory cytokines (TNF-α, IL-18, and IL-1ß) were significantly decreased in the RIC treatment group. Further findings revealed the CKD-induced infiltration of CD3, CD8a, CD11c, and F4/80 positive inflammatory cells in the heart. Treatment with RIC substantially reduced the myocardial infiltration of these inflammatory cells. From these findings, we believe that CKD-induced myocardial HDAC6 perturbation has a deteriorative effect on the heart, and inhibition of HDAC6 can be a promising approach to alleviate CKD-induced myocardial remodeling.

Scopoletin alleviates high glucose-induced toxicity in human renal proximal tubular cells via inhibition of oxidative damage, epithelial-mesenchymal transition, and fibrogenesis.

BACKGROUND: Recent years of evidence suggest the crucial role of renal tubular cells in developing diabetic kidney disease. Scopoletin (SCOP) is a plant-based coumarin with numerous biological activities. This study aimed to determine the effect of SCOP on renal tubular cells in developing diabetic kidney disease and to elucidate mechanisms. METHODS AND RESULTS: In this study, SCOP was evaluated in vitro using renal proximal tubular (HK-2) cells under hyperglycemic conditions to understand its mechanism of action. In HK-2 cells, SCOP alleviated the high glucose-generated reactive oxygen species (ROS), restored the levels of reduced glutathione, and decreased lipid peroxidation. High glucose-induced alteration in the mitochondrial membrane potential was markedly restored in the SCOP-treated cells. Moreover, SCOP significantly reduced the high glucose-induced apoptotic cell population in the Annexin V-FITC flow cytometry study. Furthermore, high glucose markedly elevated the mRNA expression of fibrotic and extracellular matrix (ECM) components, namely, transforming growth factor (TGF)-ß, alfa-smooth muscle actin (α-SMA), collagen I, and collagen III, in HK-2 cells compared to the untreated cells. SCOP treatment reduced these mRNA expressions compared to the high glucose-treated cells. Collagen I and TGF-ß protein levels were also significantly reduced in the SCOP-treated cells. Further findings in HK-2 cells revealed that SCOP interfered with the epithelial-mesenchymal transition (EMT) in the high glucose-treated HK-2 cells by normalizing E-cadherin and downregulating the vimentin and α-SMA proteins. CONCLUSIONS: In conclusion, SCOP modulates the high glucose-generated renal tubular cell oxidative damage and accumulation of ECM components and may be a promising molecule against diabetic nephropathy.

Intranasal Delivery of Carvedilol- and Quercetin-Encapsulated Cationic Nanoliposomes for Cardiovascular Targeting: Formulation and In Vitro and Ex Vivo Studies.

Carvedilol (CVD), an adrenoreceptor blocker, is a hydrophobic Biopharmaceutics Classification System class II drug with poor oral bioavailability due to which frequent dosing is essential to attain pharmacological effects. Quercetin (QC), a polyphenolic compound, is a potent natural antioxidant, but its oral dosing is restricted due to poor aqueous solubility and low oral bioavailability. To overcome the common limitations of both drugs and to attain synergistic cardioprotective effects, we formulated CVD- and QC-encapsulated cationic nanoliposomes (NLPs) in situ gel (CVD/QC-L.O.F.) for intranasal administration. We designed CVD- and QC-loaded cationic nanoliposomal (NLPs) in situ gel (CVD/QC-L.O.F.) for intranasal administration. In vitro drug release studies of CVD/QC-L.O.F. (16.25%) exhibited 18.78 ± 0.57% of QC release and 91.38 ± 0.93% of CVD release for 120 h. Ex vivo nasal permeation studies of CVD/QC-L.O.F. demonstrated better permeation of QC (within 96 h), i.e., 75.09% compared to in vitro drug release, whereas CVD permeates within 48 h, indicating the better interaction between cationic NLPs and the negatively charged biological membrane. The developed nasal gel showed a sufficient mucoadhesive property, good spreadability, higher firmness, consistency, and cohesiveness, indicating suitability for membrane application and intranasal administration. CVD-NLPs, QC-NLPs, and CVD/QC-NLPs were evaluated for in vitro cytotoxicity, in vitro ROS-induced cell viability assessment, and a cellular uptake study using H9c2 rat cardiomyocytes. The highest in vitro cellular uptake of CVD/QC-cationic NLPs by H9c2 cells implies the benefit of QC loading within the CVD nanoliposomal carrier system and gives evidence for better interaction of NLPs carrying positive charges with the negatively charged biological cells. The in vitro H2O2-induced oxidative stress cell viability assessment of H9c2 cells established the intracellular antioxidant activity and cardioprotective effect of CVD/QC-cationic NLPs with low cytotoxicity. These findings suggest the potential of cationic NLPs as a suitable drug delivery carrier for CVD and QC combination for the intranasal route in the treatment of various cardiovascular diseases like hypertension, angina pectoris, etc. and for treating neurodegenerative disorders.

Imperatorin ameliorates kidney injury in diabetic mice by regulating the TGF-ß/Smad2/3 signaling axis, epithelial-to-mesenchymal transition, and renal inflammation.

Diabetic nephropathy (DN) is a serious concern in patients with diabetes mellitus. Prolonged hyperglycemia induces oxidative damage, chronic inflammation, and build-up of extracellular matrix (ECM) components in the renal cells, leading to kidney structural and functional changes. Imperatorin (IMP) is a naturally occurring furanocoumarin derivative with proven antioxidative and anti-inflammatory properties. We investigated whether IMP could improve DN and employed high glucose (HG)-induced HK-2 cells and high-fat diet-fed streptozotocin (HFD/STZ)-generated DN experimental model in C57BL/6 mice. In vitro, IMP effectively reduced the HG-activated reactive oxygen species generation, disturbance in the mitochondrial membrane potential (MMP) and epithelial-to-mesenchymal transition (EMT)-related markers, and the transforming growth factor (TGF)-ß and collagen 1 expression in HK-2 cells. In vivo, we found an elevation of serum creatinine, kidney histology alterations, and collagen build-up in the kidneys of the DN control group. Also, we found an altered expression of EMT-related markers, upregulation of the TGF-ß/Smad2/3 axis, and elevated pro-inflammatory molecules, TNF-α, IL-1ß, IL-18 and phospho-NF-kB (p65) in the DN control group. IMP treatment did not significantly reduce the blood glucose level compared to the DN control group. However, IMP treatment effectively improved renal damage by ameliorating kidney histological changes and serum renal injury markers. IMP treatment restored renal antioxidants and exhibited anti-inflammatory effects in the kidneys. Moreover, the abnormal manifestation of EMT-related attributes and elevated levels of TGF-ß, phospho-Smad2/3, and collagen 1 were also normalized in the IMP treatment group. Our findings highlight that IMP may be a potential candidate for treating DN.

Colon-targeted delivery of niclosamide from solid dispersion employing a pH-dependent polymer via hotmelt extrusion for the treatment of ulcerative colitis in mice.

Niclosamide (NCL) is repurposed to treat inflammatory bowel disease due to its anti-inflammatory properties and potential to reduce oxidative stress. This therapeutic activity remains challenging if administered directly due to its low solubility and high recrystallization tendency in gastric pH. Solid dispersions using pH-dependent polymer will be a better idea to improve the solubility, dissolution and targeted delivery at the colon. Hot melt extrusion was used to formulate a solid dispersion with 30% NCL utilising hydroxypropyl methylcellulose acetate succinate as a pH-dependent polymer. In vitro drug release studies revealed formulation (F1) containing 10%w/w Tween 80 showed minimal release (2.06%) at the end of 2 h, followed by 47.87% and 82.15% drug release at 6 h and 14 h, respectively, indicating the maximum amount of drug release in the colon. The drug release from the formulations containing no plasticiser and 5%w/w plasticiser was comparable to the pure crystalline drug (approximately 25%). Solid-state analysis confirmed particle conversion of crystalline NCL to amorphous form, and the optimised formulation was stable for 6 months without significant changes in dissolution profile. In contrast to pure NCL, the F1 formulation substantially reduced the disease activity index, colonic inflammation, histological alterations and oxidative damage in colitis mice. These findings reveal that the prepared formulation can potentially deliver the drug locally at the colon, making it an effective tool in treating ulcerative colitis.

Fisetin attenuates arsenic and fluoride subacute co-exposure induced neurotoxicity via regulating TNF-α mediated activation of NLRP3 inflammasome.

Groundwater is considered safe, however, the occurrence of contaminants like arsenic and fluoride has raised a major healthcare concern. Clinical studies suggested that arsenic and fluoride co-exposure induced neurotoxicity, however efforts to explore safe and effective management of such neurotoxicity are limited. Therefore, we investigated the ameliorative effect of Fisetin against arsenic and fluoride subacute co-exposure-induced neurotoxicity, and associated biochemical and molecular changes. Male BALB/c mice were exposed to Arsenic (NaAsO2: 50 mg/L) and fluoride (NaF: 50 mg/L) through drinking water and fisetin (5, 10, and 20 mg/kg/day) was administered orally for 28 days. The neurobehavioral changes were recorded in the open field, rotarod, grip strength, tail suspension, forced swim, and novel object recognition test. The co-exposure resulted in anxiety-like behaviour, loss of motor coordination, depression-like behaviour, and loss of novelty-based memory, along with enhanced prooxidant, inflammatory markers and loss of cortical and hippocampal neurons. The treatment with fisetin reversed the co-exposure-induced neurobehavioral deficit along with restoration of redox & inflammatory milieu, and cortical and hippocampal neuronal density. Apart from antioxidants, inhibition of TNF-α/ NLRP3 expression has been suggested as one of the plausible neuroprotective mechanisms of Fisetin in this study.

Role of lncRNAs in hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is among the deadliest cancer in human malignancies. It is the most common and severe type of primary liver cancer. However, the molecular mechanisms underlying HCC pathogenesis remain poorly understood. Long non-coding RNAs (lncRNAs), a new kind of RNA and epigenetic factors, play a crucial role in tumorigenesis and the progression of HCC. LncRNAs are capable of promoting the autophagy, proliferation, and migration of tumor cells by targeting and modulating the expression of downstream genes in signaling pathways related to cancer; these transcripts modify the activity and expression of various tumor suppressors and oncogenes. LncRNAs could act as biomarkers for treatment approaches such as immunotherapy, chemotherapy, and surgery to effectively treat HCC patients. Improved knowledge regarding the aetiology of HCC may result from an advanced understanding of lncRNAs. Enhanced oxidative stress in the mitochondrial and Endoplasmic reticulum leads to the activation of unfolded protein response pathway that plays a crucial role in the pathophysiology of hepatocellular carcinoma. The mutual regulation between LncRNAs and Endoplasmic reticulum (ER) stress in cancer and simultaneous activation of the unfolded protein response (UPR) pathway determines the fate of tumor cells in HCC. Mitochondria-associated lncRNAs work as essential components of several gene regulatory networks; abnormal regulation of mitochondria-associated lncRNAs may lead to oncogenesis, which provides further insight into the understanding of tumorigenesis and therapeutic strategies.

Biochanin A Ameliorates Nephropathy in High-Fat Diet/Streptozotocin-Induced Diabetic Rats: Effects on NF-kB/NLRP3 Axis, Pyroptosis, and Fibrosis.

Nephropathy is the most prevalent microvascular disorder in diabetes mellitus. Oxidative stress and inflammatory cascade provoked by the persistent hyperglycemic milieu play integral roles in the aggravation of renal injury and fibrosis. We explored the impact of biochanin A (BCA), an isoflavonoid, on the inflammatory response, nod-like receptor protein 3 (NLRP3) inflammasome activation, oxidative stress, and fibrosis in diabetic kidneys. A high-fat-diet/streptozotocin (HFD/STZ)-induced experimental model of diabetic nephropathy (DN) was established in Sprague Dawley rats, and in vitro studies were performed in high-glucose-induced renal tubular epithelial (NRK-52E) cells. Persistent hyperglycemia in diabetic rats was manifested by perturbation of renal function, marked histological alterations, and oxidative and inflammatory renal damage. Therapeutic intervention of BCA mitigated histological changes, improved renal function and antioxidant capacity, and suppressed phosphorylation of nuclear factor-kappa B (NF-κB) and nuclear factor-kappa B inhibitor alpha (IκBα) proteins. Our in vitro data reveal excessive superoxide generation, apoptosis, and altered mitochondrial membrane potential in NRK-52E cells that were cultured in a high-glucose (HG) environment were subsided by BCA intervention. Meanwhile, the upregulated expressions of NLRP3 and its associated proteins, the pyroptosis-indicative protein gasdermin-D (GSDMD) in the kidneys, and HG-stimulated NRK-52E cells were significantly ameliorated by BCA treatment. Additionally, BCA blunted transforming growth factor (TGF)-ß/Smad signaling and production of collagen I, collagen III, fibronectin, and alfa-smooth muscle actin (α-SMA) in diabetic kidneys. Our results indicate the plausible role of BCA in attenuating DN, presumably through modulation of the apoptotic cascade in renal tubular epithelial cells and the NF-κB/NLRP3 axis.

Anti-inflammatory activity of carvacrol protects the heart from lipopolysaccharide-induced cardiac dysfunction by inhibiting pyroptosis via NLRP3/Caspase1/Gasdermin D signaling axis.

AIMS: Lipopolysaccharide (LPS) is a well-known agent to induce septic conditions. Sepsis-induced cardiomyopathy has an overwhelming death rate. Carvacrol (CVL), a monoterpene phenol, has anti-inflammatory and antioxidant properties. This research aimed to investigate the effect of CVL on LPS-induced dysfunction in the heart. In this study, we evaluated the effect of CVL in LPS-stimulated H9c2 cardiomyoblast cells and Balb/C mice. MAIN METHODS: LPS was used to induce septic conditions in H9c2 cardiomyoblast cells in vitro and in Balb/C mice. A survival study was conducted to assess the survival rate of mice after LPS and/or CVL treatment. KEY FINDINGS: In vitro studies indicated that CVL inhibits reactive oxygen species (ROS) generation and abates pyroptosis mediated by NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome in H9c2 cells. In mice, CVL intervention improved the survival rate in septic conditions. The CVL administration markedly improved the echocardiographic parameters and alleviated the LPS-induced reduction in the ejection fraction (%) and fraction shortening (%). The CVL intervention restored the myocardial antioxidants and histopathological alterations and decreased the pro-inflammatory cytokine contents in the heart. Further findings disclosed that CVL reduced the protein levels of NLRP3, apoptosis-associated speck-like protein (ASC), caspase 1, interleukin (IL)-18, IL-1ß, and the pyroptosis-indicative protein, gasdermin-D (GSDMD) in the heart. The autophagy-indicative proteins, beclin 1 and p62 in the heart were also restored in the CVL-treated group. SIGNIFICANCE: Altogether, our findings demonstrated that CVL has a beneficial effect and can be a potential molecule against sepsis-induced myocardial dysfunction.

Sinigrin Attenuates the Dextran Sulfate Sodium-induced Colitis in Mice by Modulating the MAPK Pathway.

Ulcerative colitis (UC) is an intestinal inflammatory disease characterised by the loss of intestinal crypts, edema, mucosal ulceration, and infiltration of inflammatory cells in the mucosa. The current study aimed to investigate the protective and therapeutic effects of sinigrin and underlying mechanisms in a dextran sulfate sodium (DSS)-induced mouse model of ulcerative colitis. DSS-induced colitis models were used to demonstrate sinigrin's therapeutic/protective action. Mice were orally administered with sinigrin (15 mg/kg or 30 mg/kg) for a period of 12 days in both prophylactic and therapeutic models. Animal weights, stool consistency, and bleeding parameters were measured throughout the experimental period. After the experimental period, colon lengths were measured, and colon tissues were harvested to determine the levels of oxidative stress-inducing factors (nitrates and MDA levels) and anti-oxidant components (GSH, SOD, and catalase). Furthermore, gene expression analysis, IL-17 levels, and inflammatory marker expressions were measured using RT-qPCR, ELISA, and immunohistochemical methods respectively. Furthermore, histopathological observations and elucidation of the mechanism of action were determined using H&E analysis and Western blot analysis. Sinigrin treatment (in both prophylactic and therapeutic models) significantly mitigated the DSS-induced body weight loss, attenuated the colon length shrinkage, and improved the disease index score (p < 0.001). Further results revealed that sinigrin's protective/therapeutic effect is associated with a significant attenuation of pro­inflammatory cytokine production (p < 0.001), reversing the anti-oxidant enzyme levels (p < 0.001) and substantial improvement (2 folds) of the disruption of the colonic morphology in colon tissues compared to DSS control. Immunohistochemical analysis showed that sinigrin treatment remarkably reduced the DSS-induced myeloperoxidase, neutrophil elastase, and CD68 expression in colon tissues. Additionally, sinigrin successfully abrogated the DSS-induced IL-17 levels (p < 0.001) and improved the colonic barrier in colon tissues. Overall, these results demonstrated that sinigrin exerts protective and therapeutic effects on DSS­induced colitis, by enhancing the anti-oxidant enzymes and suppressing the intestinal inflammatory cascade of markers by regulating the MAPK pathway.

Alternanthera brasiliana L. extract alleviates carbon tetrachloride-induced liver injury and fibrotic changes in mice: Role of matrix metalloproteinases and TGF-ß/Smad axis.

ETHNOPHARMACOLOGICAL RELEVANCE: Alternanthera brasiliana L. is a flowering plant belonging to the family Amaranthaceae and is popularly known as "penicillin". It is used in folk medicine to treat infections, coughs, wound healing, and inflammatory diseases. AIM OF THE STUDY: We investigated the effect of Alternanthera brasiliana L. leaves hydroalcoholic extract (AB) against oxidative stress, inflammation, and fibrotic changes in an experimental model of carbon tetrachloride (CCl4)-induced liver injury and fibrosis in mice. MATERIALS AND METHODS: Thirty-six male Balb/C mice were randomized into five groups: normal control, AB control, CCl4 control, CCl4 + AB-200 mg/kg, and CCl4 + AB-400 mg/kg. In mice, liver injury was induced by intraperitoneal injection of CCl4 (20% in corn oil, 5 ml/kg body weight) thrice a week for six consecutive weeks. AB extract at two doses (200 mg/kg and 400 mg/kg body weight) was administered orally for six consecutive weeks. Liver injury-related serum markers (ALT, AST, ALP), antioxidants (GSH, GST, SOD, and vitamin C), pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α), interleukin (IL)-1ß, and IL-18, ultrasonographic and histological alterations, proteins of matrix metalloproteinases (MMPs) and tissue inhibitors of matrix metalloproteinase-1 (TIMP-1), nuclear factor-κB (p65) (NF-κB), nod-like receptor protein 3 (NLRP3), and TGF-ß/Smad signaling were accessed. LC-Q-TOF-MS/MS analysis of AB was performed. RESULTS: AB treatment significantly decreased the CCl4-induced rise in serum ALT, AST, and ALP activities and improved the histological alterations. Compared with the CCl4-treated group, treatment with AB significantly restored the hepatic antioxidants and reduced the pro-inflammatory cytokines in the liver. The antioxidant activity of AB may be attributed to its terpenoid constituents, which was confirmed by LC-Q-TOF-MS/MS analysis. The CCl4-induced rise in expression of MMP-2 and MMP-9 and decrease in TIMP-1 were markedly restored in the AB-treated groups. Further findings revealed a significant reduction in the protein levels of phospho-NF-κB (p65), NLRP3, TGF-ß, pSmad2/3, collagen I, and α-smooth muscle actin (α-SMA) in the AB treatment groups. CONCLUSIONS: The hepatoprotective effect of AB may be attributed to the high content of terpenoid compounds and alleviates liver injury and associated fibrotic changes through modulating MMPs, NF-κB (p65), and the TGF-ß/Smad axis.

Nuciferine alleviates intestinal inflammation by inhibiting MAPK/NF-κB and NLRP3/Caspase 1 pathways in vivo and in vitro.

Nuciferine (NCF) is an aporphine alkaloid and a principal bioactive constituent in the lotus plant. Herewith, we investigated the potential anti-inflammatory effect and underlying mechanisms of NCF employing dextran sulfate sodium (DSS)-induced ulcerative colitis in mice, a predominant intestinal inflammatory disease, and mouse RAW 264.7 cells in vitro. Lipopolysaccharide (LPS) was used to generate an inflammatory response in the RAW 264.7 cells. The disease activity index (DAI), colon morphology, colonoscopy, and colon histopathology were performed to assess experimental colitis. The biochemical assays, enzyme-linked immunosorbent assay (ELISA), and immunoblot analysis were performed to understand the underlying mechanisms. In RAW 264.7 cells, NCF pretreatment significantly decreased the expression of inducible nitric oxide synthase (iNOS), the expression and release of pro-inflammatory cytokines including interleukin (IL)-1ß, IL-18, and tumor necrosis factor-α (TNF-α) and interfered with the activation of mitogen-activated protein kinase (MAPK), nuclear factor-κB (NF-κB), and NOD-like family pyrin domain containing 3 (NLRP3) signaling pathways. The oral treatment of NCF substantially alleviated the DSS-induced DAI, increased colon length, and restored colon morphology and histology. Compared to the DSS-induced mice, the proteins involved in the activation of MAPK/NF-κB/NLRP3 pathways and the cytokines were markedly decreased in the NCF-treated mice. Moreover, the tight junction architecture of the colon was well-maintained in NCF treatment groups by regulating the expression of claudin-1 and zonula occludens-1 (ZO-1) proteins. All these findings suggest that NCF can be a promising molecule to modulate ulcerative colitis.

Carvacrol preserves antioxidant status and attenuates kidney fibrosis via modulation of TGF-ß1/Smad signaling and inflammation.

Chronic kidney disease (CKD) with diverse aetiologies is emerging as a challenging kidney disorder associated with inflammation and interstitial fibrosis. Carvacrol (CVL) is a bioactive monoterpenoid found abundantly in oregano, thyme, and bergamot, having diverse pharmacological benefits. However, the effect of CVL against fibrotic changes in the kidneys is poorly defined. In the current study, a robust mouse model of renal fibrosis induced through unilateral ureteral obstruction (UUO) is used to investigate the anti-fibrotic activity of CVL. The mice were treated with two different oral doses of CVL (25 mg kg-1 and 50 mg kg-1 body weight) for 14 consecutive days. The UUO induction resulted in impaired renal function, severe histological damage, and collagen deposition in the obstructed kidney. Our findings revealed profound activation of transforming growth factor-ß1 (TGF-ß1) and NF-κB (p65) signaling along with the downregulation of antioxidant proteins, nuclear factor-erythroid factor 2-related factor 2 (Nrf2), NAD(P)H: quinone oxidoreductase 1 (NQO1), and superoxide dismutase (SOD) in the obstructed kidney. CVL administration markedly recovered antioxidant proteins and kidney histological changes. In addition, CVL blunted the NF-κB (p65) phosphorylation and reduced the levels of tumor necrosis factor (TNF)-α, interleukin (IL)-1ß, IL-6, and cyclooxygenase 2 (COX-2) compared to the UUO control group. CVL also alleviated the increased fibrotic protein levels of TGF-ß1, pSmad2/3, collagen I, collagen III, fibronectin, and myofibroblast activation and epithelial-mesenchymal transition (EMT) markers, including alpha-smooth muscle actin (α-SMA), E-cadherin, and vimentin in the kidneys. Findings from in vitro study also confirmed that CVL inhibits the EMT process in TGF-ß1 stimulated renal tubular epithelial cells (NRK 52E cells). Collectively, our findings indicate that CVL administration attenuates kidney fibrosis by targeting oxidative stress and inflammation.

CRISPR-based point-of-care diagnostics incorporating Cas9, Cas12, and Cas13 enzymes advanced for SARS-CoV-2 detection.

An outbreak of the novel beta coronavirus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) first came to light in December 2019, which has unfolded rapidly and turned out to be a global pandemic. Early prognosis of viral contamination involves speedy intervention, disorder control, and good-sized management of the spread of disease. Reverse transcription-polymerase chain reaction, considered the gold standard test for detecting nucleic acids and pathogen diagnosis, provides high sensitivity and specificity. However, reliance on high-priced equipped kits, associated reagents, and skilled personnel slow down sickness detection. Lately, the improvement of clustered regularly interspaced short palindromic repeat (CRISPR)-Cas (CRISPR-associated protein)-based diagnostic systems has reshaped molecular diagnosis due to their low cost, simplicity, speed, efficiency, high sensitivity, specificity, and versatility, which is vital for accomplishing point-of-care diagnostics. We reviewed and summarized CRISPR-Cas-based point-of-care diagnostic strategies and research in these paintings while highlighting their characteristics and challenges for identifying SARS-CoV-2.

Up-regulation of Nrf2/HO-1 and inhibition of TGF-ß1/Smad2/3 signaling axis by daphnetin alleviates transverse aortic constriction-induced cardiac remodeling in mice.

Oxidative damage and accumulation of extracellular matrix (ECM) components play a crucial role in the adverse outcome of cardiac hypertrophy. Evidence suggests that nuclear factor erythroid-derived factor 2 related factor 2 (Nrf2) can modulate oxidative damage and adverse myocardial remodeling. Daphnetin (Daph) is a coumarin obtained from the plant genus Daphne species that exerts anti-oxidative and anti-inflammatory properties. Herein, we investigated the roles of Daph in transverse aortic constriction (TAC)-induced cardiac hypertrophy and fibrosis in mice. TAC-induced alterations in cardiac hypertrophy markers, histopathological changes, and cardiac function were markedly ameliorated by oral administration of Daph in mice. We found that Daph significantly reduced the reactive oxygen species (ROS) generation, increased the nuclear translocation of Nrf2, and consequently, reinstated the protein levels of NAD(P)H quinone dehydrogenase1 (NQO1), heme oxygenase-1 (HO-1), and other antioxidants in the heart. Besides, Daph significantly inhibited the TAC-induced accumulation of ECM components, including α-smooth muscle actin (α-SMA), collagen I, collagen III, and fibronectin, and interfered with the TGF-ß1/Smad2/3 signaling axis. Further studies revealed that TAC-induced terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) positive nuclei and the protein levels of Bax/Bcl2 ratio and cleaved caspase 3 were substantially decreased by Daph treatment. We further characterized the effect of Daph on angiotensin II (Ang-II)-stimulated H9c2 cardiomyoblast cells and observed that Daph markedly decreased the Ang-II induced increase in cell size, production of ROS, and proteins associated with apoptosis and fibrosis. Mechanistically, Daph alone treatment enhanced the protein levels of Nrf2, NQO1, and HO-1 in H9c2 cells. The inhibition of this axis by Si-Nrf2 transfection abolished the protective effect of Daph in H9c2 cells. Taken together, Daph effectively counteracted the TAC-induced cardiac hypertrophy and fibrosis by improving the Nrf2/HO-1 axis and inhibiting the TGF-ß1/Smad2/3 signaling axis.

Biochanin A alleviates unilateral ureteral obstruction-induced renal interstitial fibrosis and inflammation by inhibiting the TGF-ß1/Smad2/3 and NF-kB/NLRP3 signaling axis in mice.

AIMS: Tubulointerstitial fibrosis, a frequent complication of chronic kidney disease (CKD) is a major public health issue. Biochanin A (BCA), an isoflavone, has numerous pharmacological activities. However, its effect on renal fibrosis and underlying molecular mechanism has not yet been clarified. This study explored the effect of BCA on renal tubulointerstitial fibrosis and inflammation in mice. MAIN METHODS: The mouse model of unilateral ureteral obstruction (UUO) in vivo and transforming growth factor (TGF)-ß1 activated renal fibroblast (NRK 49F) cells in vitro model were used to assess the antifibrotic effect of BCA. Biochemical analysis, histopathology, western blotting, and immunofluorescent staining methods were performed to elucidate the mechanism of BCA. KEY FINDINGS: In vitro, BCA suppressed the expression of fibrogenic proteins in TGF-ß1-activated renal fibroblasts. The treatment with BCA displayed less tubular injury, prevented the aberrant accumulation of extracellular matrix (ECM) components, and inhibited the TGF-ß1/Smad2/3 signaling axis in the kidneys. Furthermore, BCA impeded the phosphorylation of NF-kB(p65) and blunted the expression of inflammatory genes in the obstructed kidneys. The UUO induced expressions of nod-like receptor protein 3 (NLRP3), active caspase 1, interleukin(IL)-18, and IL-1ß proteins were decreased in the BCA treated groups. We also found the increased expression of redox-sensitive nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase 1 (HO-1) proteins in BCA treated groups compared to the UUO control. SIGNIFICANCE: These findings indicate that BCA has a therapeutic benefit against renal fibrosis, and the ameliorative effect is mediated via inhibiting the TGF-ß1/Smad2/3 and NF-kB/NLRP3 signaling axis.

Biological Activities, Pharmacokinetics and Toxicity of Nootkatone: A Review.

Plant-based drugs have a significant impact on modern therapeutics due to their vast array of pharmacological activities. The integration of herbal plants in the current healthcare system has emerged as a new field of research. It can be used for the identification of novel lead compound candidates for future drug development. Nootkatone is a sesquiterpene derivative and an isolate of grapefruit. Shreds of evidence illustrate that nootkatone targets few molecular mechanisms to exhibit its pharmacological activity and yet needs more exploration. The current review is related to nootkatone, drafted through a literature search using research articles and books from different sources, including Science Direct, Google Scholar, Elsevier, PubMed, and Scopus. It has been reported to possess a wide range of pharmacological activities such as anti-inflammatory, anticancer, antibacterial, hepatoprotective, neuroprotective, and cardioprotective. Although preclinical studies in experimental animal models suggest that nootkatone has therapeutic potential, it is further warranted to evaluate its toxicity and pharmacokinetic parameters before being applied to humans. Hence, in the present review, we have summarized the scientific knowledge on nootkatone with a particular emphasis on its pharmacological properties to encourage researchers for further exploration in preclinical and clinical settings.

Perillyl Alcohol Attenuates NLRP3 Inflammasome Activation and Rescues Dopaminergic Neurons in Experimental In Vitro and In Vivo Models of Parkinson's Disease.

NLRP3 activation plays a key role in the initiation and progression of a variety of neurodegenerative diseases. However, understanding the molecular mechanisms involved in the bidirectional signaling required to activate the NLRP3 inflammasomes is the key to treating several diseases. Hence, the present study aimed to investigate the role of lipopolysaccharide (LPS) and hydrogen peroxide (H2O2) in activating NLRP3 inflammasome-driven neurodegeneration and elucidated the neuroprotective role of perillyl alcohol (PA) in in vitro and in vivo models of Parkinson's disease (PD). Initial priming of microglial cells with LPS following treatment with H2O2 induced NF-κB translocation to the nucleus with a robust generation of free radicals that act as signal 2 in augmenting NLRP3 inflammasome assembly and its downstream targets. PA treatment suppresses the nuclear translocation of NF-κB, enhances PARKIN translocation into the mitochondria, and maintains cellular redox homeostasis in both mouse and human microglial cells that limit NLRP3 inflammasome activation along with processing of active caspase-1, IL-1ß, and IL-18. To further correlate the in vitro study with the in vivo MPTP model, treatment with PA also inhibited the nuclear translocation of NF-κB and downregulated the NLRP3 inflammasome activation. PA administration upregulated various antioxidant enzymes' levels and restored the level of dopamine and other neurotransmitters in the striatum of the mouse brain, subsequently improving the behavioral activities. Therefore, we conclude that NLRP3 inflammasome activation required a signal from damaged mitochondria for its activation. Further pharmacological scavenging of free radicals restricts microglia activation and simultaneously supports neuronal survival via targeting the NLRP3 inflammasome pathway in PD.

Aloin alleviates pathological cardiac hypertrophy via modulation of the oxidative and fibrotic response.

AIMS: Pathological cardiac hypertrophy is a characteristic feature in many cardiovascular diseases (CVDs). Aloin is an anthraquinone glycoside from Aloe species, and the effect of aloin on cardiac hypertrophy and associated fibrotic changes have not been elucidated. This study investigated the effect of aloin against the isoproterenol (ISO)-induced cardiac hypertrophy in rats. MAIN METHODS: Cardiac hypertrophy experimental model was induced in rats by subcutaneous injection of ISO for 14 days. Meanwhile, the animals were administered orally with aloin at doses of 25 and 50 mg/kg/day. On the 15th day, cardiac echocardiography was performed, the heart was collected and subjected for histopathological, gene expression, and immunoblot studies. Additionally, the effect of aloin on ISO-induced hypertrophic changes in H9c2 cells was investigated. KEY FINDINGS: Aloin markedly alleviated ISO-induced heart injury, reduced cardiac hypertrophy, improved cardiac function, and histological alterations in the heart. Mechanistically, aloin attenuated ISO-induced fibrosis via inhibition of the levels of collagen I, α-smooth muscle actin (α-SMA), fibronectin, transforming growth factor-ß (TGF-ß) and pSmad2/3 proteins in the heart. Aloin alleviated ISO-induced myocardial oxidative damage and up-regulated the levels of antioxidant transcription factor nuclear factor erythroid 2-related factor 2 (Nrf2) and heme oxygenase-1 (HO-1) proteins. Moreover, aloin treatment attenuated ISO-induced hypertrophic changes and the generation of reactive oxygen species (ROS) in H9c2 cells in vitro. SIGNIFICANCE: Our findings demonstrated that aloin alleviated ISO-induced cardiac hypertrophy and fibrosis via inhibiting TGF-ß/pSmad2/3 signaling and restoring myocardial antioxidants, and therefore has promising therapeutic potential against cardiac hypertrophy and fibrosis.