Ursolic acid inhibits NF-κB signaling and attenuates MMP-9/TIMP-1 in progressive osteoarthritis: a network pharmacology-based analysis.

Osteoarthritis (OA) is a degenerative joint disease, characterized by infiltration of monocytes into the synovial joint which promotes inflammation, stiffness, joint swelling, cartilage degradation and further bone destruction. The leaves of Ocimum forskolei have been used for inflammation-related disease management in traditional medicine. Additionally, the downregulation of NF-κB and the MMP/TIMP-1 ratio has been shown to protect against OA. The LC-HR-MS metabolic analysis of Ocimum yielded 19 putative compounds, among which ursolic acid (UA) was detected. Ursolic acid possesses significant anti-inflammatory effects and has been reported to downregulate oxidative stress and inflammatory biomarkers. It was tested on rats in a model of intra-articular carrageenan injection to investigate its efficacy on osteoarthritis progression. The UA emulgel exerted chondroprotective, analgesic and local anaesthetic efficacies confirmed via histopathological investigation and radiographical imaging. A network pharmacology followed by molecular docking highlighted TNF-α, TGF-ß and NF-κB as the top filtered genes. Quantitative real-time PCR analysis showed that UA significantly attenuated serum levels of TNF-α, IL-1ß, NF-κB, MMP-9/TIMP-1 and elevated levels of TGF-ß. Taken together, these results suggest that UA could serve as a functional food-derived phytochemical with a multi-targeted efficacy on progression of OA, regulating the immune and inflammatory responses, particularly, attenuating chondrocytes degeneration via suppression of NF-κB and MMP-9/TIMP-1. Accordingly, UA might be a promising alternative to conventional therapy for safe, easily applicable and effective management of OA.

Wound healing potential of Cystoseira/mesenchymal stem cells in immunosuppressed rats supported by overwhelming immuno-inflammatory crosstalk.

Wound healing, one of the most intricate and dynamic processes of the body, maintains skin integrity following trauma. One of the main issues that still exists is impaired wound healing, particularly for immunosuppressed patients. Recently, natural products from marine environments have been employed in wound-repairing activities. This work investigates the mesenchymal stem cells in the combined capacity of the bone marrow (BMMSC) for wound healing and Cystoseira sp. Algae extract in immunosuppressed rats. High-resolution liquid chromatography / MS investigation of Cystoseira extract revealed the prevalence of fatty acids that have wound-soothing potential. From constructed PPI network for wound healing and further analysis through molecular docking and molecular dynamics (MD) simulation experiments suggested that cystalgerone metabolite may be responsible for the wound healing-promoting effect of Cystoseira extract. According to the CD marker characterization of the BMMSC, 98.21% of them expressed CD90, and 97.1% expressed CD105. Sixteen d after immunity suppression (by 40 mg/kg hydrocortisone daily), an incision was made in the dorsal skin of the rat. The treatments were applied for 16 d and samples were taken from the tested groups on the 8th, 14th, and 16th days. The BMMSCs / Cystoseira group showed significantly improved wound closure, thickness, density of new layers, and skin elasticity than the control group (p < 0.001). The BMMSCs / Cystoseira combination significantly reduced the oxidative indicators, pro-inflammatory cytokines, and immune markers, according to the RT-PCR gene expression study. In order to delve deeper into the complex interconnections among wound healing-related biological targets and pinpoint key factors in this complex process, we engaged in network pharmacology and computational research. Subsequently, we conducted a comprehensive computational analysis, including reverse docking, free energy (ΔG) computation, and molecular dynamics simulations, on the molecular structures of the annotated compounds. The purpose of this investigation was to identify potential new targets for these chemicals as well as any potential interactions they may have with different signaling pathways related to the wound healing process. Our research indicates that the primary compounds of Cystoseira holds potential wound healing therapeutic activity. Although more safety testing and clinical studies are required, the combination has great potential for regenerative medicine and could be a revolutionary advance in the healing of the wounds of immunosuppressed patients.

Wound Restorative Power of Halimeda macroloba/ Mesenchymal Stem Cells in Immunocompromised Rats via Downregulating Inflammatory/Immune Cross Talk.

Impaired skin wound healing is still a major challenge, especially with immunocompromised patients who express delayed healing and are susceptible to infections. Injection of rat-derived bone marrow mesenchymal stem cells (BMMSCs) via the tail vein accelerates cutaneous wound healing via their paracrine activity. The present work aimed to investigate the combined wound-healing potential of BMMSCs and Halimeda macroloba algae extract in immunocompromised rats. High-resolution liquid chromatography-mass spectrometry (HR-LC-MS) investigation of the extract revealed the presence of variant phytochemicals, mostly phenolics, and terpenoids, known for their angiogenic, collagen-stimulating, anti-inflammatory, and antioxidant properties. The BMMSCs were isolated and characterized for CD markers, where they showed a positive expression of CD90 by 98.21% and CD105 by 97.1%. Twelve days after inducing immunocompromise (40 mg/kg hydrocortisone daily), a circular excision was created in the dorsal skin of rats and the treatments were continued for 16 days. The studied groups were sampled on days 4, 8, 12, and 16 after wounding. The gross/histopathological results revealed that the wound closure (99%), thickness, density of new epidermis and dermis, and skin elasticity in the healed wounds were considerably higher in the BMMSCs/Halimeda group than the control group (p < 0.05). RT-PCR gene expression analysis revealed that the BMMSCs/Halimeda extract combination had perfectly attenuated oxidative stress, proinflammatory cytokines, and NF-KB activation at day 16 of wounding. The combination holds promise for regenerative medicine, representing a revolutionary step in the wound healing of immunocompromised patients, with still a need for safety assessments and further clinical trials.

The Potential of Corchorus olitorius Seeds Buccal Films for Treatment of Recurrent Minor Aphthous Ulcerations in Human Volunteers.

Aphthous ulcers are very common disorders among different age groups and are very noxious and painful. The incidence of aphthous ulcer recurrence is very high and it may even last for a maximum of 6 days and usually, patients cannot stand its pain. This study aims to prepare a buccoadhesive fast dissolving film containing Corchorus olitorius seed extract to treat recurrent minor aphthous ulceration (RMAU) in addition to clinical experiments on human volunteers. An excision wound model was used to assess the in vivo wound healing potential of Corchorus olitorius L. seed extract, with a focus on wound healing molecular targets such as TGF-, TNF-, and IL-1. In addition, metabolomic profiling using HR-LCMS for the crude extract of Corchorus olitorius seeds was explored. Moreover, molecular docking experiments were performed to elucidate the binding confirmation of the isolated compounds with three molecular targets (TNF-α, IL-1ß, and GSK3). Additionally, the in vitro antioxidant potential of C. olitorius seed extract using both H2O2 and superoxide radical scavenging activity was examined. Clinical experiments on human volunteers revealed the efficiency of the prepared C. olitorius seeds buccal fast dissolving film (CoBFDF) in relieving pain and wound healing of RMAU. Moreover, the wound healing results revealed that C. olitorius seed extract enhanced wound closure rates (p ≤ 0.001), elevated TGF-ß levels and significantly downregulated TNF-α and IL-1ß in comparison to the Mebo-treated group. The phenotypical results were supported by biochemical and histopathological findings, while metabolomic profiling using HR-LCMS for the crude extract of Corchorus olitorius seeds yielded a total of 21 compounds belonging to diverse chemical classes. Finally, this study highlights the potential of C. olitorius seed extract in wound repair uncovering the most probable mechanisms of action using in silico analysis.

The Wound-Healing Potential of Olea europaea L. Cv. Arbequina Leaves Extract: An Integrated In Vitro, In Silico, and In Vivo Investigation.

Olea europaea L. Cv. Arbequina (OEA) (Oleaceae) is an olive variety species that has received little attention. Besides our previous work for the chemical profiling of OEA leaves using LC−HRESIMS, an additional 23 compounds are identified. An excision wound model is used to measure wound healing action. Wounds are provided with OEA (2% w/v) or MEBO® cream (marketed treatment). The wound closure rate related to vehicle-treated wounds is significantly increased by OEA. Comparing to vehicle wound tissues, significant levels of TGF-ß in OEA and MEBO® (p < 0.05) are displayed by gene expression patterns, with the most significant levels in OEA-treated wounds. Proinflammatory TNF-α and IL-1ß levels are substantially reduced in OEA-treated wounds. The capability of several lignan-related compounds to interact with MMP-1 is revealed by extensive in silico investigation of the major OEA compounds (i.e., inverse docking, molecular dynamics simulation, and ΔG calculation), and their role in the wound-healing process is also characterized. The potential of OEA as a potent MMP-1 inhibitor is shown in subsequent in vitro testing (IC50 = 88.0 ± 0.1 nM). In conclusion, OEA is introduced as an interesting therapeutic candidate that can effectively manage wound healing because of its anti-inflammatory and antioxidant properties.

Mechanistic Wound Healing and Antioxidant Potential of Moringa oleifera Seeds Extract Supported by Metabolic Profiling, In Silico Network Design, Molecular Docking, and In Vivo Studies.

Moringa oleifera Lam. (Moringaceae) is an adaptable plant with promising phytoconstituents, interesting medicinal uses, and nutritional importance. Chemical profiling of M. oleifera seeds assisted by LC-HRMS (HPLC system coupled to a high resolution mass detector) led to the dereplication of 19 metabolites. Additionally, the wound healing potential of M. oleifera seed extract was investigated in male New Zealand Dutch strain albino rabbits and supported by histopathological examinations. Moreover, the molecular mechanisms were investigated via different in vitro investigations and through analyzing the relative gene and protein expression patterns. When compared to the untreated and MEBO®-treated groups, topical administration of M. oleifera extract on excision wounds resulted in a substantial increase in wound healing rate (p < 0.001), elevating TGF-ß1, VEGF, Type I collagen relative expression, and reducing inflammatory markers such as IL-1ß and TNF-α. In vitro antioxidant assays showed that the extract displayed strong scavenging effects to peroxides and superoxide free radicals. In silico studies using a molecular docking approach against TNF-α, TGFBR1, and IL-1ß showed that some metabolites in M. oleifera seed extract can bind to the active sites of three wound-healing related proteins. Protein−protein interaction (PPI) and compound−protein interaction (CPI) networks were constructed as well. Quercetin, caffeic acid, and kaempferol showed the highest connectivity with the putative proteins. In silico drug likeness studies revealed that almost all compounds comply with both Lipinski's and Veber's rule. According to the previous findings, an in vitro study was carried out on the pure compounds, including quercetin, kaempferol, and caffeic acid (identified from M. oleifera) to validate the proposed approach and to verify their potential effectiveness. Their inhibitory potential was evaluated against the pro-inflammatory cytokine IL-6 and against the endopeptidase MMPs (matrix metalloproteinases) subtype I and II, with highest activity being observed for kaempferol. Hence, M. oleifera seeds could be a promising source of bioactive compounds with potential antioxidant and wound healing capabilities.

Antiulcer Potential of Psidium guajava Seed Extract Supported by Metabolic Profiling and Molecular Docking.

One of the most severe human health problems is gastric ulceration. The main aim of our study is to explore the gastroprotective effect of the Psidium guajava seeds extract (PGE). Metabolic profiling based on LC-HRMS for the extract led to the dereplication of 23 compounds (1-23). We carried out a gastric ulcer model induced by indomethacin in male albino rats in vivo and the extract of PGE was investigated at a dose of 300 mg/kg in comparison to cimetidine (100 mg/kg). Furthermore, the assessment of gastric mucosal lesions and histopathology investigation of gastric tissue was done. It has been proved that Psidium guajava seeds significantly decreased the ulcer index and protected the mucosa from lesions. The antiulcer effect of Psidium guajava seed extract, which has the power of reducing the ensuing inflammatory reactions, can counteract the inflammation induced by indomethacin by the downregulation of relative genes expression (IL-1ß, IL-6, and TNF-α). Moreover, PGE significantly downregulated the increased COX-2, TGF-ß, and IGF-1 relative genes expression, confirming its beneficial effect in ulcer healing. Moreover, the possible PGE antioxidant potential was determined by in vitro assays using hydrogen peroxide and superoxide radical scavenging and revealed high antioxidant potential. Additionally, on the putatively annotated metabolites, an in silico study was conducted, which emphasized the extract's antiulcer properties might be attributed to several sterols such as stigmasterol and campesterol. The present study provided evidence of Psidium guajava seeds considered as a potential natural gastroprotective agent.

Wound Healing and Antioxidant Capabilities of Zizyphus mauritiana Fruits: In-Vitro, In-Vivo, and Molecular Modeling Study.

LC-HRMS-assisted chemical profiling of Zizyphus mauritiana fruit extract (ZFE) led to the dereplication of 28 metabolites. Furthermore, wound healing activity of ZFE in 24 adult male New Zealand Dutch strain albino rabbits was investigated in-vivo supported by histopathological investigation. Additionally, the molecular mechanism was studied through different in-vitro investigations as well as, studying both relative gene expression and relative protein expression patterns. Moreover, the antioxidant activity of ZFE extract was examined using two in-vitro assays including hydrogen peroxide and superoxide radical scavenging activities that showed promising antioxidant potential. Topical application of the extract on excision wounds showed a significant increase in the wound healing rate (p < 0.001) in comparison to the untreated and MEBO®-treated groups, enhancing TGF-ß1, VEGF, Type I collagen expression, and suppressing inflammatory markers (TNF-α and IL-1ß). Moreover, an in silico molecular docking against TNFα, TGFBR1, and IL-1ß showed that some of the molecules identified in ZFE can bind to the three wound-healing related protein actives sites. Additionally, PASS computational calculation of antioxidant activity revealed potential activity of three phenolic compounds (Pa score > 0.5). Consequently, ZFE may be a potential alternative medication helping wound healing owing to its antioxidant and anti-inflammatory activities.

Antioxidant and Wound Healing Potential of Vitis vinifera Seeds Supported by Phytochemical Characterization and Docking Studies.

This study explored the in vivo wound healing potential of Vitis vinifera seed extract using an excision wound model with focus on wound healing molecular targets including TGFBR1, VEGF, TNF-α, and IL-1ß. The wound healing results revealed that V. vinifera seed extract enhanced wound closure rates (p < 0.001), elevated TGF-ß and VEGF levels, and significantly downregulated TNF-α and IL-1ß levels in comparison to the Mebo®-treated group. The phenotypical results were supported by biochemical and histopathological findings. Phytochemical investigation yielded a total of 36 compounds including twenty-seven compounds (1−27) identified from seed oil using GC-MS analysis, along with nine isolated compounds. Among the isolated compounds, one new benzofuran dimer (28) along with eight known ones (29−36) were identified. The structure of new compound was elucidated utilizing 1D/2D NMR, with HRESIMS analyses. Moreover, molecular docking experiments were performed to elucidate the molecular targets (TNF-α, TGFBR1, and IL-1ß) of the observed wound healing activity. Additionally, the in vitro antioxidant activity of V. vinifera seed extract along with two isolated compounds (ursolic acid 34, and ß-sitosterol-3-O-glucopyranoside 36) were explored. Our study highlights the potential of V. vinifera seed extract in wound repair uncovering the most probable mechanisms of action using in silico analysis.

Vincamine Modulates the Effect of Pantoprazole in Renal Ischemia/Reperfusion Injury by Attenuating MAPK and Apoptosis Signaling Pathways.

Pantoprazole has an antioxidant function against reactive oxygen species (ROS). Vincamine, a herbal candidate, is an indole alkaloid of clinical use against brain sclerosis. The aim of the present experiment is to evaluate, on a molecular level for the first time, the value of vincamine in addition to pantoprazole in treating experimentally induced renal ischemia/reperfusion injury (IRI). One-hundred-and-twenty-eight healthy male Wistar albino rats were included. Serum creatinine, blood urea nitrogen, and malondialdehyde levels were assessed. ELISA was used to estimate the pro-inflammatory cytokines. The expression of Bcl-2 and Bax genes was assessed by quantitative real-time PCR. ERK1/2, JNK1/2, p38, cleaved caspase-3, and NF-κB proteins expressions were estimated using western blot assay. The kidneys were also histopathologically studied. The IRI resulted in impaired cellular functions with increased creatinine, urea nitrogen, malondialdehyde, TNF-α, IL-6, and IL-1ß serum levels, and up-regulated NF-ĸB, JNK1/2, ERK1/2, p38, and cleaved caspase-3 proteins. Furthermore, it down-regulated the expression of the Bcl-2 gene and upregulated the Bax gene. The treatment with vincamine, in addition to pantoprazole multiple doses, significantly alleviated the biochemical and histopathological changes more than pantoprazole or vincamine alone, whether the dose is single or multiple, declaring their synergistic effect. In conclusion, vincamine with pantoprazole multiple doses mitigated the renal IRI through the inhibition of apoptosis, attenuation of the extracellular signaling pathways through proinflammatory cytokines' levels, and suppression of the MAPK (ERK1/2, JNK, p38)-NF-κB intracellular signaling pathway.

Pantoprazole Attenuates MAPK (ERK1/2, JNK, p38)-NF-κB and Apoptosis Signaling Pathways after Renal Ischemia/Reperfusion Injury in Rats.

Ischemia/reperfusion injury (IRI) in the kidney is the most common cause of acute renal dysfunction through different cell damage mechanisms. This study aimed to investigate, on molecular basics for the first time, the effect of pantoprazole on renal IRI in rats. Different biochemical parameters and oxidative stress markers were assessed. ELISA was used to estimate proinflammatory cytokines. qRT-PCR and western blot were used to investigate the gene and protein expression. Renal histopathological examination was also performed. IRI resulted in tissue damage, elevation of serum levels of creatinine, urea nitrogen, malondialdehyde, TNF-α, IL-6, IL-1ß, up-regulation of NF-κB, JNK1/2, ERK1/2, p38, and cleaved caspase-3 proteins. Furthermore, it up-regulated the expression of the Bax gene and down-regulated the expression of the Bcl-2 gene. Treatment of the injured rats with pantoprazole, either single dose or multiple doses, significantly alleviated IRI-induced biochemical and histopathological changes, attenuated the levels of proinflammatory cytokines, down-regulated the expression of NF-κB, JNK1/2, ERK1/2, p38, and cleaved caspase-3 proteins, and the Bax gene, and up-regulated Bcl-2 gene expression. Moreover, treatment with pantoprazole multiple doses has an ameliorative effect that is greater than pantoprazole single-dose. In conclusion, pantoprazole diminished renal IRI via suppression of apoptosis, attenuation of the pro-inflammatory cytokines' levels, and inhibition of the intracellular signaling pathway MAPK (ERK1/2, JNK, p38)-NF-κB.

Hyphaene thebaica (doum)-derived extract alleviates hyperglycemia in diabetic rats: a comprehensive in silico, in vitro and in vivo study.

In the present study, we investigated the hypoglycemic effect of different extracts (i.e. organic and aqueous) derived from the fruits of Hyphaene thebaica (doum) on male streptozotocin-induced diabetic rats. Blood glucose levels as well as the relative gene expression of insulin, TNF-α, and TGF-ß were determined in the pancreatic tissue of the experimental animals. Treatment of STZ-induced diabetic rats with aqueous extracts of the plant fruit over 7 weeks significantly reduced the elevated blood glucose and increased the relative expression of insulin, while the relative expression of inflammatory mediators (i.e. TNF-α and TGF-ß) was significantly reduced. Histopathological investigation also revealed that the aqueous extract treatment effectively reversed the ß-cell necrosis induced by STZ and restored its normal morphology. Furthermore, liquid chromatography high resolution mass spectrometry (LC-HRMS) and in silico chemical investigation of the aqueous extract elucidated its major bioactive phytochemicals (i.e. flavonoids) and putatively determined the pancreatic KATP channel as a target for these bioactive components. In vitro insulin secretion assay revealed that myricetin, luteolin, and apigenin were able to induce insulin secretion by human pancreatic cells (insulin production = 20.9 ± 1.3, 13.74 ± 1.8, and 11.33 ± 1.1 ng mL-1, respectively). Using molecular docking and dynamics simulations, we were able to shed the light on the insulin secretagogue's mode of action through these identified bioactive compounds and to determine the main structural elements required for its bioactivity. This comprehensive investigation of this native fruit will encourage future clinical studies to recommend edible and widely available fruits like doum to be a part of DM treatment plans.

A novel nasal co-loaded loratadine and sulpiride nanoemulsion with improved downregulation of TNF-α, TGF-ß and IL-1 in rabbit models of ovalbumin-induced allergic rhinitis.

PURPOSE: The work aimed to develop a co-loaded loratadine and sulpiride nasal nanoemulsion for allergic rhinitis management. METHODS: Compatibility studies were conducted adopting differential scanning calorimetry and Fourier transform infrared spectroscopy. Nanoemulsion formulations were prepared using soybean lecithin, olive oil and tween 80. Sodium cholate and glycerol were employed as co-surfactants. Nanoemulsions were assessed for viscosity, pH, droplet size, polydispersity index, zeta potential, electrical conductivity, entrapment, In vitro drug release and corresponding kinetics. Stability of the selected formulation was investigated. The biological effectiveness was evaluated in rabbit models of ovalbumin-induced allergic rhinitis by measuring TNF-α, TGF-ß and IL-1. RESULTS: Compatibility studies revealed absence of drug/drug interactions. Nanoemulsions exhibited > 90% entrapment efficiency. The selected nanoemulsion demonstrated small droplet size (85.2 ± 0.2 nm), low PDI (0.35 ± 0.0) and appropriate Zeta Potential (-23.3 ± 0.2) and stability. It also displayed enhanced in vitro drug release following the Higuashi Diffusion and Baker-Lonsdale models. The mean relative mRNA expression of TNF-α, IL-1 and TGF-ß significantly decreased from 9.59 ± 1.06, 4.15 ± 0.02 and 4.15 ± 0.02 to 1.28 ± 0.02, 1.93 ± 0.06 and 1.56 ± 0.02 respectively after treatment with the selected nanoemulsion formulation. CONCLUSION: The results reflected a promising potent effect of the combined loratadine and sulpiride nasal nanoemulsion in managing the symptoms of allergic rhinitis.

Antiulcer secondary metabolites from Elaeocarpus grandis, family Elaeocarpaceae, supported by in silico studies.

Elaeocarpus grandis has a very potent analgesic effect, especially to a δ-opioid receptor, but its antiulcer activity has not yet been validated. Therefore, the present study was carried out to evaluate the antiulcer potential of the total methanolic extract and its derived fractions of the aerial parts of the plant using an indomethacin-induced gastric ulcer method. One new compound, grandisine H (1), and five known compounds, P-methoxy benzaldehyde, methyl gallate, kaempferol, quercetin and heterophyllin A (2-6), were isolated from the ethyl acetate fraction, which was the most potent one with an ulcer index value of 5 ± 1.95 (mm) ** (*P < 0.05, **P < 0.01) and a preventive index of 92.9%, following a bioassay-guided fractionation. The isolated compounds were subjected to a molecular docking study in an attempt to explain their significant antiulcer potential, and the results revealed that kaempferol and quercetin bind to the active site of the M3 receptor with a strong binding affinity via strong hydrogen bonds of -6.081 kcal mol-1 and -6.013 kcal mol-1, respectively. Also, quercetin and heterophyllin A showed a binding affinity with the gastric proton pump receptor and a strong hydrogen bond interaction with the amino acid active sites in the case of an H2-modeled receptor. These results clarify the effectiveness and importance of the ethyl acetate fraction as a natural anti-ulcer remedy.