In-vitro Effects of Bacterial Melanin in Macrophage “RAW 264.7” Cell Culture

Many manufacturers use melanins from different sources to produce cosmetics. Melanin inhibits the expression of inflammation-associated enzymes, matrix metalloproteinases, and restores levels of cutaneous antioxidant enzymes.1 Study of hydrophilic bacterial melanin (BM) effects was initiated with experiments on plants. BM was obtained from the mutant strain of Bacillus Thuringiensis at the Institute of Biotechnology in Armenia. It was shown to be a potent stimulator of plant growth.2 BM has been tested in animal models to assess its effects on neuronal viability and regeneration. Topical injection of BM exhibited stimulatory action on axonal growth in experiments with injury of motor tracts and peripheral nerves,3 comanifested with better neuronal viability, neovascularization with extensive dilation of capillaries, suppressed inflammatory response and other effects (4). In our study the skin of animals was exposed to the pigment as well. Monitoring of the operated animals revealed stimulated hair growth in operated rats. To clarify this effect, we applied melanin solution (9 mg/mL for 10 consecutive days) to the larger cleaned lower back skin area (2 cm) in the experimental group rats (n = 12) and compared them to control animals with the same shaved skin area (n = 12). The skin of the study animals was observed every 3 days in order to register the hair regrowth pattern. One of the most commonly used hair growth assessment scale was offered by Matsuda et al, where the 0–4 scores are scaled the following way: no hair growth, less than 20%, 20%–39%, 40%–59%, 60%–79% and 80%–100% of hair growth.5 Another method used by many research groups is the self-designed scale of hair regrowth (high hair density and full, thick fur (Level 4)); moderate hair density with no visible skin area (Level 3), low hair density, with the visualization of the skin (Level 2), uneven hair growth on the tested area, skin easily seen (Level 1).6 Significant hair growth was observed in all 12 experimental rats treated topically with melanin. The average growth in the study and control groups was measured on the 10th, 20th, and 30th days after the topical use of melanin solution (Table 1). The rat head has the shortest hair regrowth period of 4 weeks. The growth rate decreases to the lower back where the molt period is around 8 weeks. Microscopic analysis showed that BM affects the hair anagen phase, prolonging it and increasing anagen/telogen ratio. The skin examination in the experimental group showed expressed vasodilation in skin areas exposed to melanin. This effect was not observed in the control rats. Many hair growth stimulants possess vasodilatory action (e.g., Minoxidil). However, the future for the alopecia management and hair stimulant development is not limited to vasodilators or antiandrogenic agents. The regenerative medicine has suggested new approaches for the hair growth technology including the use of stem cells and growth factors. The above mentioned stimulation of axonal (neuritic) growth by the hydrophilic melanin is mediated by endogenous neurotrophic growth factors, which include NGF, BDNF, GDNF, and IGF-1. The last factor is a confirmed regulator of hair growth cycle supporting the anagen stage and was shown to affect hair morphogenesis. These preliminary data constitute the background to assess possible use of the water-soluble and low-cost BM as a proper topical therapeutic agent for inducing hair growth in a case control study. As a hair-growth stimulant the BM acts with “double-hit effect” including vasodilation and enhanced regeneration. All authors have read and approved the final manuscript. P.T.R. and H.A.S designed the research study. P.T.R., H.A.S, A.S.V., K.H.O performed the research. A.S.V. and P.M.H analyzed data. P.T.R. and A.S.V. wrote the paper. The study was conducted in the frame of the international project supported by the U.S. Department U.S. Department of State, Bureau of International Security and Nonproliferation, Office of Cooperative Threat Reduction. ISTC Project PR-132.1.3. None declared. All authors have read and apprsoved the final version of the manuscript. The protocol was approved by the Institutional Animal Care and Ethics Committee of SNPO NAS RA in accordance with the European Communities Council Directive (86/609/EEC) on care and use of animals for experimental procedures (N8-6/19, 21 March 2019). All applicable institutional and/or national guidelines for the care and use of animals were followed. Not Applicable.

Depression is a globally recognized leading cause of mental health disability and mortality worldwide, severely affecting individuals' daily lives and relationships. At present, the limitation of first-line antidepressants has brought great difficulties to the treatment of depression. Therefore, the development of new antidepressants is imminent. Quercetin is considered as a potential antidepressant drug due to its potent anti-inflammatory, antioxidant, and neuroprotective effects. We explored the antidepressant effect of quercetin by establishing a mouse model (Adult male C57BL/6J mice) of depression induced by chronic unpredictable stress (CUS). Analysis of the antidepressant effect of quercetin was conducted using multi-omics. The sociability and the protein expression levels of Hsp90, XBP1 and iNOS in the hippocampus of mice were evaluated through behavioral and molecular biology experiments. We found that quercetin treatment alleviated social impairment in mice and could target Hsp90 to inhibit the upregulation of XBP1 and iNOS. In addition, based on microbiome as well as untargeted metabolomics results of hippocampal brain regions, quercetin was found to increase the abundance of Lactobacillus and modulate the levels of inosine, Creatinine, and Lithocholic acid metabolites in the hippocampus. To sum up, in the present study, we revealed the molecular mechanism of quercetin alleviating depression-like behavior in CUS mice by inhibiting CUS-induced endoplasmic reticulum stress, neuroinflammation, gut microbiota dysbiosis, and metabolic disorders through multi-omics joint analysis. This provides a complementary scientific explanation for the antidepressant effects of quercetin.

Parkinson’s disease is a chronic neurodegenerative disease characterized by a significant loss of dopaminergic neurons within the substantia nigra pars compacta region and a subsequent loss of dopamine within the striatum. A promising avenue of research has been the administration of growth factors to promote the survival of remaining midbrain neurons, although the mechanism by which they provide neuroprotection is not understood. Activin A, a member of the transforming growth factor β superfamily, has been shown to be a potent anti-inflammatory following acute brain injury and has been demonstrated to play a role in the neuroprotection of midbrain neurons against MPP+-induced degeneration in vitro. We hypothesized that activin A may offer similar anti-inflammatory and neuroprotective effects in in vivo mouse models of Parkinson’s disease. We found that activin A significantly attenuated the inflammatory response induced by both MPTP and intranigral administration of lipopolysaccharide in C57BL/6 mice. We found that administration of activin A promoted survival of dopaminergic and total neuron populations in the pars compacta region both 8 days and 8 weeks after MPTP-induced degeneration. Surprisingly, no corresponding protection of striatal dopamine levels was found. Furthermore, activin A failed to protect against loss of striatal dopamine transporter expression in the striatum, suggesting the neuroprotective action of activin A may be localized to the substantia nigra. Together, these results provide the first evidence that activin A exerts potent neuroprotection and anti-inflammatory effects in the MPTP and lipopolysaccharide mouse models of Parkinson’s disease.

Propofol reduces microglia activation and neurotoxicity through inhibition of extracellular vesicle release

Anti-Inflammatory and Neuroprotective Effects of DIPOPA (N,N-Diisopropyl-2-Oxopropanamide), an Ethyl Pyruvate Bioisoster, in the Postischemic Brain.

BackgroundCurcumin has been demonstrated to possess promising neuroprotective potential in Parkinson’s disease; however, its overall effects remain inconclusive, and its multiple mechanisms of action have not been systematically summarized.ObjectiveThis systematic review and meta-analysis aimed to evaluate the pharmacological effects of curcumin in animal models of Parkinson’s disease and to investigate its potential mechanisms involving antioxidant, anti-inflammatory, and neuroprotective effects, thereby providing a theoretical basis for its potential clinical application in Parkinson’s disease.MethodsA comprehensive search of four databases (EMBASE, PubMed, Web of Science, and the Cochrane Library) up to August 2025 identified 31 eligible studies involving a total of 552 animals. Methodological quality was assessed using the SYRCLE risk of bias tool. Standardized mean differences (SMDs) were calculated to evaluate the effects of curcumin on motor function, neurochemistry, inflammation, and oxidative stress in animal models of Parkinson’s disease.ResultsThe results demonstrated that curcumin intervention improved motor function in animal models of Parkinson’s disease, as evidenced by increased locomotor distance in the open field test (SMD = 1.25) and elevated mean velocity (SMD = 1.42), prolonged latency to fall in the rotarod test (SMD = 2.49), shortened descent time in the pole test (SMD = −1.16), and reduced traversal time on the balance beam (SMD = −2.27). Curcumin exhibited neuroprotective effects through increasing the number of tyrosine hydroxylase-positive cells (SMD = 2.12), maintaining dopamine levels (SMD = 4.11), and elevating 3,4-dihydroxyphenylacetic acid concentrations (SMD = 3.15). Regarding anti-inflammatory effects, curcumin significantly reduced multiple inflammatory markers, including interleukin-6 (SMD = −4.73), interleukin-1β (SMD = −3.30), tumor necrosis factor-α (SMD = −3.19), and nitric oxide (SMD = −4.91). With respect to antioxidant activity, curcumin significantly reduced malondialdehyde levels (SMD = −4.69) while increasing the activities of superoxide dismutase (SMD = 3.90), glutathione (SMD = 2.08), and catalase (SMD = 2.00).ConclusionCurcumin demonstrates significant neuroprotective effects in Parkinson’s disease animal models, improving motor deficits and neuronal integrity likely through multi-target mechanisms involving anti-inflammatory and antioxidant pathways.Systematic Review Registerhttps://www.crd.york.ac.uk/PROSPERO/view/CRD420251131257, identifier CRD420251131257

Anxiolytic and antidepressive potentials of rosmarinic acid: A review with a focus on antioxidant and anti-inflammatory effects

BackgroundLaquinimod is an immunomodulatory drug under clinical investigation for the treatment of the progressive form of multiple sclerosis (MS) with both anti-inflammatory and neuroprotective effects. Excitotoxicity, a prominent pathophysiological feature of MS and of its animal model, experimental autoimmune encephalomyelitis (EAE), involves glutamate transporter (GluT) dysfunction in glial cells.The aim of this study was to assess whether laquinimod might exert direct neuroprotective effects by interfering with the mechanisms of excitotoxicity linked to GluT function impairments in EAE.MethodsOsmotic minipumps allowing continuous intracerebroventricular (icv) infusion of laquinimod for 4 weeks were implanted into C57BL/6 mice before EAE induction. EAE cerebella were taken to perform western blot and qPCR experiments. For ex vivo experiments, EAE cerebellar slices were incubated with laquinimod before performing electrophysiology, western blot, and qPCR.ResultsIn vivo treatment with laquinimod attenuated EAE clinical score at the peak of the disease, without remarkable effects on inflammatory markers. In vitro application of laquinimod to EAE cerebellar slices prevented EAE-linked glutamatergic alterations without mitigating astrogliosis and inflammation. Moreover, such treatment induced an increase of Slcla3 mRNA coding for the glial glutamate–aspartate transporter (GLAST) without affecting the protein content. Concomitantly, laquinimod significantly increased the levels of the glial glutamate transporter 1 (GLT-1) protein and pharmacological blockade of GLT-1 function fully abolished laquinimod anti-excitotoxic effect.ConclusionsOverall, our results suggest that laquinimod protects against glutamate excitotoxicity of the cerebellum of EAE mice by bursting the expression of glial glutamate transporters, independently of its anti-inflammatory effects.

Inflammation and oxidative stress are key pathological drivers of many chronic and neurodegenerative disorders, prompting increasing interest in natural compounds that can safely modulate inflammatory responses and protect neural cells from oxidative damage. In this study, walnut shell pectin (WSP), extracted from Chilean walnut shells, was evaluated for its anti-inflammatory and neuroprotective potential using in vitro models. The anti-inflammatory effects were assessed in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages, while the neuroprotective effects were investigated in rotenone-treated SH-SY5Y neuroblastoma cells. WSP significantly reduced LPS-induced inflammation by downregulating the expression of pro-inflammatory cytokine TNF-α and enhancing the anti-inflammatory cytokine IL-10. In rotenone-exposed SH-SY5Y cells, WSP increased the activities of antioxidant enzymes, catalase and superoxide dismutase, thereby reducing oxidative stress and improving neuronal viability. The maximum protective concentration for both cell lines was determined to be 50 µg/ml. Overall, WSP exhibited strong anti-inflammatory and neuroprotective activities in vitro, highlighting its potential as a natural therapeutic compound for preventing and managing inflammation-associated and neurodegenerative disorders.

Inhibition of Glycogen Synthase Kinase 3β (GSK3β) Decreases Inflammatory Responses in Brain Endothelial Cells

Alpha-MSH is a tridecapeptide derived from proopiomelanocortin. Many studies over the last few years have provided evidence that alpha-MSH has potent protective and antiinflammatory effects. These effects can be elicited via centrally expressed melanocortin receptors that orchestrate descending neurogenic antiinflammatory pathways. alpha-MSH can also exert antiinflammatory and protective effects on cells of the immune system and on peripheral nonimmune cell types expressing melanocortin receptors. At the molecular level, alpha-MSH affects various pathways implicated in regulation of inflammation and protection, i.e., nuclear factor-kappaB activation, expression of adhesion molecules and chemokine receptors, production of proinflammatory cytokines and mediators, IL-10 synthesis, T cell proliferation and activity, inflammatory cell migration, expression of antioxidative enzymes, and apoptosis. The antiinflammatory effects of alpha-MSH have been validated in animal models of experimentally induced fever; irritant and allergic contact dermatitis, vasculitis, and fibrosis; ocular, gastrointestinal, brain, and allergic airway inflammation; and arthritis, but also in models of organ injury. One obstacle limiting the use of alpha-MSH in inflammatory disorders is its pigmentary effect. Due to its preserved antiinflammatory effect but lack of pigmentary action, the C-terminal tripeptide of alpha-MSH, KPV, has been delineated as an alternative for antiinflammatory therapy. KdPT, a derivative of KPV corresponding to amino acids 193-195 of IL-1beta, is also emerging as a tripeptide with antiinflammatory effects. The physiochemical properties and expected low costs of production render both agents suitable for the future treatment of immune-mediated inflammatory skin and bowel disease, fibrosis, allergic and inflammatory lung disease, ocular inflammation, and arthritis.

An updated and comprehensive review of the health benefits and pharmacological activities of hesperidin.

Stronger Neo-Minophagen C (SNMC) is a glycyrrhizin-containing preparation that is approved in Japan for the treatment of chronic hepatic diseases and is marketed in Japan, China, Korea, Taiwan, and India. Glycyrrhizin, a triterpene present in the roots and rhizomes of licorice (Glycyrrhiza glabra) has been shown to have anti-inflammatory, anti-oxidative, and anti-viral effects. In the present study, we demonstrated the marked neuroprotective effects of SNMC in the postischemic rat brain after middle cerebral artery occlusion (MCAO). We used 1 ml/kg of SNMC, which is within the dose range used for the treatment of patients with chronic hepatic disease. The administration of SNMC intravenously at 30 minutes before or 30 minutes and 3 hours after MCAO (60 minutes) reduces mean infarct volumes to 27.0±4.2%, 37.1±12.4%, and 67.8±5.8% of that of untreated controls, respectively. This neuroprotective effect is accompanied by improvements in motor impairment and neurological deficits. The administration of SNMC is shown to suppress microglia activation and neutrophil infiltration in the postischemic brain. In addition, SNMC suppresses lipopolysaccharide-induced nitrite production and proinflammatory cytokine induction in a microglia cell line, BV2. This indicates that the neuroprotective effect of SNMC might be due, at least in part, to an anti-inflammatiory effect. Interestingly, SNMC shows significantly higher neuroprotective potency compared to an equivalent dose of pure glycyrrhizin, in terms of reducing infarct volume and improving neurological deficits. Together these results indicate that SNMC, a glycyrrhizin-containing preparation developed for chronic liver disease, has a marked neuroprotective function in the postischemic brain via its anti-inflammatory effects.

Recently, neuroinflammation is thought to be one of the important causes of many neuropsychiatric diseases. Quercetin (QUER) is a natural flavonoid, and it is well known that QUER has antioxidative, anti-inflammatory, and neuroprotective effects. In our study, lipopolysaccharide (LPS) was injected into the lateral ventricle of rats to induce anxiety-like behaviors and neuroinflammation, and it was confirmed that chronic administration of QUER could improve anxiety-like symptoms. We also investigated the effects of QUER on inflammatory markers and its major mechanisms associated with inflammation in the hippocampus. Daily administration of QUER (10, 50, and 100 mg/kg) daily for 21 days significantly improved anxiety-like behaviors in the elevated plus-maze test and open field test. QUER administration significantly reduced inflammatory markers such as interleukin-6, interleukin-1β, cyclooxygenase-2, and nuclear factor-kappaB levels in the brain. In addition, QUER significantly increased the brain-derived neurotrophic factor (BDNF) mRNA level and decreased the nitric oxide synthase (iNOS) mRNA level. Therefore, our results have shown that QUER can improve anxiety-like behaviors caused by chronic neuroinflammation. This anxiolytic effect of QUER has been shown to be due to its anti-inflammatory effects and appropriate regulation of BDNF and iNOS expression. Thus, QUER provides the potential as a therapeutic agent to inhibit anxiety-like symptoms in neuropsychiatric diseases, such as anxiety.

PurposeMelittin is the main peptide component of melittin venom, with anti-allergic, anti-inflammatory, anti-arthritis, anti-cancer, and neuroprotective effects. Melittin has been confirmed to have anti-inflammatory effects in immune and liver cells and is widely used in treating arthritis and rheumatic diseases. At the same time, related studies have shown that melittin can enhance the expression of muscle differentiation related factors in mice. However, whether the melittin can improve the expression of stem cells into muscle differentiation and use it to treat Skeletal muscle atrophy, it has not yet been clarified.As "seed cells," stem cells have been widely concerned in the field of regenerative medicine research, such as embryonic stem cells, induced pluripotent stem cells, mesenchymal stem cells, etc., among which adipose mesenchymal stem cells (ADSCs) have excellent application potential in the treatment of Skeletal muscle atrophy because of their quantitative advantages, easy access, and inherent good differentiation potential and self-renewal ability. To investigate whether melittin can stimulate ADSCs MYOGenic differentiation and whether combined with ADSCs can treat Skeletal muscle atrophy.MethodsFour concentration ranges of melittin were set, namely Group A (20 mg/L, 40 mg/L, 80 mg/L, 160 mg/L, 320 mg/L), Group B (2 mg/L, 4 mg/L, 8 mg/L, 16 mg/L, 32 mg/L), Group C (0.2 mg/L, 0.4 mg/L, 0.8 mg/L, 1.6 mg/L, 3.2 mg/L), and Group D (0.02 mg/L, 0.04 mg/L, 0.08 mg/L, 0.16 mg/L, 0.32 mg/L), Intervention of adipose stem cells with different concentrations of melittin for 48 hours, followed by measurement of relative cell viability using the CCK-8 method.Melittin after 48 hours of intervention with ADSCs, Western blot was used to detect the expression levels of MYOGenic differentiation related factors.The control group (C), melittin group (M), dexamethasone group (D), and Dexamethasone+Melittin group (D+M) were set separately. After 12 hours of intervention with dexamethasone (10umol/L), the ADSCs were added with melittin for 48 hours. Western blot was used to detect MYOGenic differentiation related factors MYOGenic(MYOG), MYF5, MYOD, MYH4 p38 and p-p38 protein. In order to observe the process of melittin repairing cell atrophy, detected MYOD and MYOG using immunofluorescence.Randomly divided into control group (C), damage group (T), stem cell therapy group (SC), melittin combined with ADSCs therapy group (SC+M). Group C does not do any treatment. Group T, SC, and SC+M set up a mouse shoulder sleeve injury model, all from the left gang of the rats, and the suture is marked with surgical sutures. On the first day of surgery, The SC group of the muscle injecting ADSCs(1×106)in the SC group, and the ADSCs after the melittin intervention of the SC+M mouse is repeatedly treated every seven days. On the left, the tendon is extracted, and the muscles are extracted to perform related tissue-related testing such as Su Mu Jing-Yi Hong dyed (H&E dyed), immunohistochemistry, muscle fiber weight and diameter.ResultsCCK-8 Experiment shows that the suitable concentration of Melittin has no significant toxic effect on ADSCs.Western blot experiment shows that the appropriate concentration of Melittin can enhance the expression of muscle differentiation related factors in ADSCs.Western blot and immunofluorescence staining results show that the expression related factors of the D+M group are better than that of group D, and the expression related factors of forming muscle differentiation in group M is also better than group C.Melittin can promote ADSCs muscle differentiation through the p38-MAPK signaling pathway.Animal experiments have shown that the expression related factors of the SC+M treatment group’s muscle differentiation related factors is significantly higher than that of the SC therapy group, and the expression related factors of the two groups of rats of SC+M and SC groups are better than the T group Rat. Organic research has confirmed that rats who have been treated with ADSCs treated with melittin have significantly improved in muscle weight, muscle fiber diameter, and muscle fiber cross-sectional area. At the same time, the result of immunohistochemicals shows that the expression of MYOD and MYOG in the SC+M group is better than the SC group.ConclusionThe results of this study indicate that melittin can promote MYOGenic differentiation of ADSCs and increase the expression of MYOGenic differentiation related factors in atrophic ADSCs.Secondly, melittin can activate the p38-MAPK signaling pathway and promote the MYOGenic differentiation of ADSCs. Stem cells treated with melittin intervention can improve the recovery of muscle mass, muscle fiber diameter, and muscle fiber cross-sectional area in rats with skeletal muscle injury compared to rats treated with stem cells alone. Therefore, the above results provide insights into MYOGenic differentiation and may provide a potential alternative strategy for related diseases such as skeletal muscle atrophy.