Enhancing collagen based nanoemulgel for effective topical delivery of Aceclofenac and Citronellol oil: Formulation, optimization, in-vitro evaluation, and in-vivo osteoarthritis study with a focus on HMGB-1/RAGE/NF-κB pathway, Klotho, and miR-499a.

The majority of conventional osteoarthritis (OA) treatments are based on molecular adjustment of certain signaling pathways associated with osteoarthritis (OA) pathogenesis, however there is a significant need to search for more effective and safe treatments. This study centers around formulating Aceclofenac (ACF) with high bioavailability in combination with Citronellol oil and collagen. The optimal concentrations of Citronellol oil/D-Limonene oil, Tween 80, and Transcutol HP were determined using a pseudoternary phase diagram. The formulated nanoemulsions were studied for thermophysical stability. Thermodynamically stable formula were analyzed for droplet size, zeta potential, and in-vitro permeation. Then, collagen based nanoemulsion were prepared to capitalize on its efficacy in reducing osteoarthritis side effects and characterized for nano size properties. Formulae F10 and F10C were chosen as optimum nanosize formula. Hense, they were prepared and characterized as nanoemulgel dosage form. The nanoemulgel formulae F10NEG1 and F10CNEG1 showed reasonable viscosity and spreadability, with complete drug release after 4 h. These formulae were chosen for further In vivo anti-OA study. Collagen based ACF/citronellol emugel were able to modulate HMGB-1/RAGE/NF-κB pathway, mitigating the production of inflammatory cytokine TNF-α. They were also able to modulate Klotho and miR-499, reducing serum CTXII and COMP, by reducing the cartilage destruction. Histological investigations validated the efficacy, safety, and superiority of Aceclofenac in combination with Citronellol oil and collagen (F10CNEG1) over solo the treated group (F10NEG1 and blank). Hence, the findings of the current work encourage the use of this promising combined formula in treatment of OA patients.

L-Carnitine augments probenecid anti-inflammatory effect in monoiodoacetate-induced knee osteoarthritis in rats: involvement of miRNA-373/P2X7/NLRP3/NF-κB milieu.

Osteoarthritis (OA) is a degenerative joint disease, whereas the underlying molecular trails involved in its pathogenesis are not fully elucidated. Hence, the current study aimed to investigate the role of miRNA-373/P2X7/NLRP3/NF-κB trajectory in its pathogenesis as well as the possible anti-inflammatory effects of probenecid and l-carnitine in ameliorating osteoarthritis via modulating this pathway. In the current study, male Sprague Dawley rats were used and monoiodoacetate (MIA)-induced knee osteoarthritis model was adopted. Probenecid and/or L-carnitine treatments for 14 days succeeded in reducing OA knee size and reestablishing motor coordination and joint mobility assessed by rotarod testing. Moreover, different treatments suppressed the elevated serum levels of IL-1ß, IL-18, IL-6, and TNF-α via tackling the miRNA-373/P2X7/NLRP3/NF-κB, witnessed as reductions in protein expressions of P2X7, NLRP3, cleaved caspase-1 and NF-κB. These were accompanied by increases in procaspase-1 and IκB protein expression and in miRNA-373 gene expression OA knee to various extents. In addition, different regimens reversed the abnormalities observed in the H and E as well as Safranin O-Fast green OA knees stained sections. Probenecid or l-carnitine solely showed comparable results on the aforementioned parameters, whereas the combination therapy had the most prominent effect on ameliorating the aforementioned parameters. In conclusion, l-carnitine augmented the probenecid's anti-inflammatory effect to attenuate MIA-induced osteoarthritis in rats by provoking the miRNA-373 level and inhibiting the P2X7/NLRP3/NF-κB milieu, leading to the suppression of serum inflammatory cytokines: IL-1ß, IL-18, IL-6, and TNF-α. These findings suggest the possibility of using probenecid and l-carnitine as a useful therapeutic option for treatment of osteoarthritis.

Antioxidant and anti-Alzheimer activities of Clivia miniata (Lindl) roots, bulbs, and aerial parts: In-vitro and in-silico studies.

Clivia miniata (Lindl) is a member of the family Amaryllidaceae known for its chemically diverse alkaloids with a wide range of biological activities. Many reports revealed a direct role of oxidative stress in the early stage of Alzheimer's disease (AD). Meanwhile, ß-site amyloid precursor protein cleavage enzyme 1 (BACE-1) is a molecular target for the treatment of AD. We aimed to investigate C. miniata root, bulb, and aerial part chemical profiling, antioxidant, BACE-1, and AChE enzyme inhibitory activities. Results showed that the total root had the most potent radical scavenging activity as compared to the total bulb and aerial part, respectively. Ethanol root extract had the most potent BACE-1 inhibitory activity (IC50 = 0.02 ± 0.001 µg/mL) as compared to the bulb and aerial part (IC50 = 0.93 ± 0.13, 1.80 ± 0.24 µg/mL), respectively. Moreover, the total root extract mitigated AChE enzyme activity more than total bulb and aerial fractions with IC50 values of (0.06 ± 0.02, 0.58 ± 0.3, and 1.89 ± 0.42 µg/mL, respectively. Bioassay-guided acid-base fractionation confirmed superior BACE-1 inhibitory activity of the root fractions particularly, methylene chloride and ethyl acetate fractions with (IC50 values of 0.21 ± 0.60 and 0.01 ± 0.001 µg/mL), respectively. UPLC-MS analysis of ethyl acetate and methylene chloride fractions of C. miniata root led to the identification of eight phenolics and thirteen alkaloids, respectively. Molecular docking studies against BACE-1 protein revealed that lycorine di-hexoside, miniatine, and cliviaaline were the most promising hits. Further investigation of anti-AD potential of the aforementioned small molecules is required.

Diosmin nanocrystal gel alleviates imiquimod-induced psoriasis in rats via modulating TLR7,8/NF-κB/micro RNA-31, AKT/mTOR/P70S6K milieu, and Tregs/Th17 balance.

Diosmin is a flavonoid with promising anti-inflammatory and antioxidant properties. However, it has difficult physicochemical characteristics since its solubility demands a pH level of 12, which has an impact on the drug's bioavailability. The aim of this work is the development and characterization of diosmin nanocrystals using anti-solvent precipitation technique to be used for topical treatment of psoriasis. Results revealed that diosmin nanocrystals stabilized with hydroxypropyl methylcellulose (HPMC E15) in ratio (diosmin:polymer; 1:1) reached the desired particle size (276.9 ± 16.49 nm); provided promising colloidal properties and possessed high drug release profile. Additionally, in-vivo assessment was carried out to evaluate and compare the activities of diosmin nanocrystal gel using three different doses and diosmin powder gel in alleviating imiquimod-induced psoriasis in rats and investigating their possible anti-inflammatory mechanisms. Herein, 125 mg of 5% imiquimod cream (IMQ) was applied topically for 5 consecutive days on the shaved backs of rats to induce psoriasis. Diosmin nanocrystal gel especially in the highest dose used offered the best anti-inflammatory effect. This was confirmed by causing the most statistically significant reduction in the psoriasis area severity index (PASI) score and the serum inflammatory cytokines levels. Furthermore, it was capable of maintaining the balance between T helper (Th17) and T regulatory (Treg) cells. Moreover, it tackled TLR7/8/NF-κB, miRNA-31, AKT/mTOR/P70S6K and elevated the TNFAIP3/A20 (a negative regulator of NF-κB) expression in psoriatic skin tissues. This highlights the role of diosmin nanocrystal gel in tackling imiquimod-induced psoriasis in rats, and thus it could be a novel promising therapy for psoriasis.

Gastroprotective and anti-Helicobacter pylori potentials of essential oils from the oleoresins of Araucaria bidwillii and Araucaria heterophylla.

Plant resins or oleoresins comprise a chemically complex mixture of different classes of compounds. Oleoresin of the genus Araucaria combines essential oil (EO) and resin. It possesses gastroprotective, cytotoxic, and timicrobial, antipyretic, and anti-inflammatory activities. The study aimed to investigate the EOs from the oleoresins of two Araucaria species, A. bidwillii and A. heterophylla, chemically and biologically for their gastroprotective, anti-inflammatory, antioxidant, and anti-Helicobacter pylori potentials. The chemical composition of both species cultivated in Egypt was analyzed with GC-MS and compared with those cultivated abroad using principal component analysis (PCA). There were 37 and 17 secondary metabolites identified in A. heterophylla and A. bidwillii, respectively. The EOs of both species showed a pronounced inhibitory effect on Helicobacter pylori activity in vitro. The gastroprotective effect was assessed in vivo using ethanol-induced gastric ulcer model in rats. Inflammatory cytokines, oxidative stress, and the nuclear factor-kappa B (NF-κB) biomarkers were assessed in the stomach tissues. The ulcer index and percentage of ulcer protection were determined. Stomach sections were examined histopathologically by staining with (H/E) and periodic acid Schiff (PAS). Moreover, the proliferative index was determined using the Ki-67 immunostaining. The treatment of rats with EOs (50, 100, and 200 mg/kg, orally) 1 hour prior to ethanol administration showed promising gastroprotective, anti-inflammatory, and antioxidant potentials. These findings declared the gastroprotective role played by both EOs with the superiority of A. bidwillii over A. heterophylla via modulation of oxidative stress/NF-κB/inflammatory cytokines. Their use can be recommended to protect against the recurrence of peptic ulcers.

Anti-neoplastic action of Cimetidine/Vitamin C on histamine and the PI3K/AKT/mTOR pathway in Ehrlich breast cancer.

The main focus of our study is to assess the anti-cancer activity of cimetidine and vitamin C via combating the tumor supportive role of mast cell mediators (histamine, VEGF, and TNF-α) within the tumor microenvironment and their effect on the protein kinase A(PKA)/insulin receptor substrate-1(IRS-1)/phosphatidylinositol-3-kinase (PI3K)/serine/threonine kinase-1 (AKT)/mammalian target of rapamycin (mTOR) cue in Ehrlich induced breast cancer in mice. In vitro study was carried out to evaluate the anti-proliferative activity and combination index (CI) of the combined drugs. Moreover, the Ehrlich model was induced in mice via subcutaneous injection of Ehrlich ascites carcinoma cells (EAC) in the mammary fat pad, and then they were left for 9 days to develop obvious solid breast tumor. The combination therapy possessed the best anti-proliferative effect, and a CI < 1 in the MCF7 cell line indicates a synergistic type of drug interaction. Regarding the in vivo study, the combination abated the elevation in the tumor volume, and serum tumor marker carcinoembryonic antigen (CEA) level. The serum vascular endothelial growth factor (VEGF) level and immunohistochemical staining for CD34 as markers of angiogenesis were mitigated. Additionally, it reverted the state of oxidative stress and inflammation. Meanwhile, it caused an increment in apoptosis, which prevents tumor survival. Furthermore, it tackled the elevated histamine and cyclic adenosine monophosphate (cAMP) levels, preventing the activation of the (PKA/IRS-1/PI3K/AKT/mTOR) cue. Finally, we concluded that the synergistic combination provided a promising anti-neoplastic effect via reducing the angiogenesis, oxidative stress, increasing apoptosis,as well as inhibiting the activation of PI3K/AKT/mTOR cue, and suggesting its use as a treatment option for breast cancer.

Galangin mitigates DOX-induced cognitive impairment in rats: Implication of NOX-1/Nrf-2/HMGB1/TLR4 and TNF-α/MAPKs/RIPK/MLKL/BDNF.

The cognitive and behavioral decline observed in cancer survivors who underwent doxorubicin (DOX)-based treatment raises the need for therapeutic interventions to counteract these complications. Galangin (GAL) is a flavonoid-based phytochemical with pronounced protective effects in various neurological disorders. However, its impact on DOX-provoked neurotoxicity has not been clarified. Hence, the current investigation aimed to explore the ability of GAL to ameliorate DOX-provoked chemo-brain in rats. DOX (2 mg/kg, once/week, i.p.) and GAL (50 mg/kg, 5 times/week., via gavage) were administered for four successive weeks. The MWM and EPM tests were used to evaluate memory disruption and anxiety-like behavior, respectively. Meanwhile, targeted biochemical markers and molecular signals were examined by the aid of ELISA, Western blotting, and immune-histochemistry. In contrast to DOX-impaired rats, GAL effectively preserved hippocampal neurons, improved cognitive/behavioral functions, and enhanced the expression of the cell repair/growth index, BDNF. The antioxidant feature of GAL was confirmed by the amelioration of MDA, NO and NOX-1, along with restoring the Nrf-2/HO-1/GSH cue. In addition, GAL displayed marked anti-inflammatory properties as verified by the suppression of the HMGB1/TLR4 nexus and p-NF-κB p65 to inhibit TNF-α, IL-6, IL-1ß, and iNOS. This inhibitory impact extended to entail astrocyte activation, as evidenced by the diminution of GFAP. These beneficial effects were associated with a notable reduction in p-p38MAPK, p-JNK1/2, and p-ERK1/2, as well as the necroptosis cascade p-RIPK1/p-RIPK3/p-MLKL. Together, these pleiotropic protective impacts advocate the concurrent use of GAL as an adjuvant agent for managing DOX-driven neurodegeneration and cognitive/behavioral deficits. DATA AVAILABILITY: The authors confirm that all relevant data are included in the supplementary materials.

Possible Implication of Nrf2, PPAR-γ and MAPKs Signaling in the Protective Role of Mangiferin against Renal Ischemia/Reperfusion in Rats.

Mangiferin (Mang) is a known glucosylxanthone that has proven its shielding effect against ischemia/reperfusion (Is/R). However, its full underlying mechanistic perspective against renal Is/R induced lesions is not fully revealed. Consequently, the purpose of this study is to track further non-investigated modulatory signals of Mang against the renal Is/R model involving nuclear factor erythroid 2-related factor (Nrf)2/heme oxygenase (HO)-1, peroxisome proliferator-activated receptor (PPAR)-γ/nuclear factor (NF)-κB, p38 mitogen-activated protein kinase (MAPK), and c-Jun N-terminal kinase (JNK) signaling. To ratify our aim, Mang was administrated (20 mg/kg, i.p for seven days) before the induction of bilateral Is/R. Mechanistic maneuver revealed that Mang balanced oxidative state via increasing the expression of the antioxidant Nrf2/HO-1 cue with subsequent enhancement of GSH besides MDA lessening. Additionally, Mang enhanced PPAR-γ mRNA expression and declined p-p38 MAPK and p-JNK expression with concomitant NF-κB downsizing leading to iNOS/NOx and TNF-α rebating. Furthermore, the Mang anti-apoptotic trait was affirmed by enriching Bcl-2 expression as well as decreasing Bax and caspase-3 expression. All these potentials were in the line with the molecular docking results and the improved histopathological findings and renal function biomarkers. Consequently, Mang provided plausible protective mechanisms against renal Is/R-related events, possibly by amending oxidative status, inflammatory mediators, and apoptotic cell death through the involvement of Nrf2, PPAR-γ, MAPK, JNK, and NF-κB signaling.

Micro RNAs 26b, 20a inversely correlate with GSK-3 ß/NF-κB/NLRP-3 pathway to highlight the additive promising effects of atorvastatin and quercetin in experimental induced arthritis.

Rheumatoid arthritis (RA) is an inflammatory disease with challenging therapeutic potential due to the implication of cross-talking intracellular pathways in the pathogenesis of the disease. This study aimed to evaluate the effects of the combination therapy of atorvastatin and quercetin on glycogen synthase kinase-3 beta/ nuclear factor kappa-B/ nucleotide-binding oligomerization domain-like receptor family pyrin domain containing-3 or inflammasome (GSK-3ß/NF-KB/NLRP-3) pathway as well as on microRNAs 26b and 20a (miR-26b, miR-20a) and to investigate the possible beneficial outcomes of the combination to offer a better treatment option than methotrexate (MTX) in adjuvant-induced arthritis (AIA). Assessment of arthritis progression, serum inflammatory, and oxidative parameters were done. The tibiotarsal tissue expression of the inflammatory parameters was evaluated. Western blot analysis was done to assess the expression level of the important members in the GSK-3ß/NF-κB/NLRP-3 pathway. Furthermore, the expression level of both microRNAs and serum level of transaminases were determined. All treatments, especially the combination regimen, abated arthritis progression, the elevated serum level of inflammatory and oxidative stress parameters in arthritic rats. Moreover, They down-regulated the gene expression of the important members of the aforementioned signaling pathway, amended the tissue levels of inflammatory parameters and elevated the expression level of miR-26b and miR-20a. Finally, we concluded that the combination therapy modulated miR-26b and miR-20a as well as GSK-3ß/NF-κB/NLRP-3 pathway, provided additive anti-inflammatory and anti-oxidant effects and offered an additional hepatoprotective effect as compared to untreated arthritic rats and MTX-treated groups, suggesting its promising role to be used as replacement therapy to MTX in RA.

Orchestrated modulation of rheumatoid arthritis via crosstalking intracellular signaling pathways.

Cell signaling is considered a part of a network for communication that regulates basic cellular activities. The ability of cells to communicate correctly to the surrounding environment has an important role in development, tissue repair, and immunity as well as normal tissue homeostasis. Dysregulated activation and crosstalk between many intracellular signaling pathways are implicated in the pathogenesis of rheumatoid arthritis (RA), such as the Janus Kinase/signal transducers and activators of transcription (JAK/STAT), Toll-like receptor/nuclear factor kappa B (TLR/NF-κB), phosphatidylinositide-3Kinase/protein kinase B/mammalian target of rapamycin (PI-3K/AKT/mTOR), the stress activated protein kinase/mitogen-activated protein kinase (SAPK/MAPK), and spleen tyrosine kinase (SYK) pathways. Other interrelated pathways that can be targeted to halt the inflammatory status in the disease are purinergic 2X7 receptor (P2X7R)/nucleotide binding oligomerization domain-like receptor family pyrin domain containing 3 or inflammasome (NLRP-3)/NF-κB and Notch pathways. In this review, we will show the orchestrated modulation in the pathogenesis of RA via the crossregulation between dysregulated signaling pathways which can mediate a sustained loop of activation for these signaling pathways as well as aggrevate the inflammatory condition. Also, this review will highlight many targets that can be useful in the development of more effective therapeutic options.

Sitagliptin and tofacitinib ameliorate adjuvant induced arthritis via modulating the cross talk between JAK/STAT and TLR-4/NF-κB signaling pathways.

AIMS: Rheumatoid arthritis is an autoimmune systemic disorder causing pain, swelling, stiffness, and disability in various joints. This work was designed to evaluate the effect of sitagliptin and tofacitinib on Janus kinase (JAK)/signaling transducer and activator of transcription (STAT) and toll like receptor (TLR-4)/nuclear factor kappa B (NF-κB) signaling pathways in adjuvant induced arthritis in rats. MATERIALS AND METHODS: Severity of arthritis was evaluated and serum was analyzed for inflammatory mediators. The mRNA and protein expression level of the most important members of the two signaling pathways were determined. Lipid profile, transaminases and renal function parameters were assessed. KEY FINDINGS: Sitagliptin and tofacitinib significantly decreased the level of inflammatory parameters, the mRNA and protein expression level of the members of JAK/STAT and TLR-4/NF-κB pathways with more prominent effect of sitagliptin on TLR-4/NF-κB pathway and more expected obvious effect of tofacitinib on JAK/STAT pathway. The combination offered additional anti-inflammatory effect by inhibiting the cross talk between these pathways as inhibition of NF-κB activation decreased the serum level of IL-6 preventing the activation of STAT-3 in tibiotarsal tissues. SIGNIFICANCE: The combination of tofacitinib and sitagliptin normalized serum lipids and blood glucose level which could offer protection against cardiovascular diseases and caused partial reversal of serum transaminases and creatinine levels which can protect against tofacitinb's related hepato and nephrotoxicity. We could conclude that the combination of Sitagliptin with tofacitinib can offer synergistic anti-inflammatory effect and more protective action against side effects of tofacitinib.