Fibromyalgia (FM) is a chronic syndrome characterized by widespread pain, fatigue, cognitive dysfunction and neuropsychiatric disturbances, associated with dysregulated oxidative stress and neuroinflammation. Emerging evidence suggests that microRNAs guiding these pathways, including miR-145-5p and miR-451a, may play a central role in FM pathogenesis. However, currently available pharmacological therapies provide limited efficacy and are frequently associated with adverse effects. Therefore, the present study investigated whether the methanolic extract of Hibiscus sabdariffa (MeOH-Exs), a polyphenol rich extract with known antioxidant and anti-inflammatory properties, alleviates reserpine (Res)-induced FM in rats through modulation of the neuroinflammation-oxidative stress-microRNA axis. FM was induced by Res (1 mg/kg, s.c., for three days), followed by 28-day treatment with MeOH-Exs (25 or 50 mg/kg, p.o.) compared with the standard drug Neurontin (Neu). Behavioral assessments, serum hydrogen peroxide, brain serotonin, peripheral expression of miR-145-5p and miR-451a, and spinal cord inflammatory and histopathological alterations were evaluated. MeOH-Exs significantly improved hyperalgesia, depressive- and anxiety-like behaviors, locomotion, and cognition relative to the Res group. The 50 mg/kg dose exerted the most pronounced effects, restoring expression of miR-145-5p and miR-451a, reducing oxidative stress, and elevating brain serotonin levels. Histological and immunohistochemical analyses confirmed attenuation of spinal cord inflammation and neuronal degeneration, caspase 3 expression and TNF-α immunoreactivity with effects comparable to or greater than Neu. Collectively, these findings indicate that Hibiscus sabdariffa may ameliorate FM - like menifistations by targeting an integrated biochemical pathway linking oxidative stress, neuroinflammation, apoptosis, microRNA dysregulation and spinal cord histopathological alterations. While the precise molecular mechanisms require further investigation, MeOH-Exs demonstrates a promising multi-target neuroprotective profile and may represent a potential complementary preclinical therapeutic candidate for FM.

Fibromyalgia (FM) is a debilitating nociplastic pain disorder increasingly recognized to involve skeletal muscle as an active contributor to peripheral nociception and central sensitization rather than a passive target of disease. Although angiotensin II promotes pain signaling through angiotensin type-1 (AT1) receptor activation, the role of skeletal muscle AT1 receptor signaling in orchestrating the inflammatory, trophic, and proteotoxic disturbances underlying FM remains unexplored. This study investigated, for the first time, whether AT1 receptor blockade by valsartan interrupts maladaptive muscle-derived signals that drive FM-associated pain and fatigue. Female rats were allocated into control groups (vehicle- and valsartan-treated) and reserpine-induced FM groups that either received oral valsartan for six consecutive days or remained untreated. Valsartan alleviated mechanical allodynia and hyperalgesia while enhancing motor performance and fatigue resistance. Mechanistically, FM muscle exhibited upregulation of lncRNA H19 accompanied by suppression of miR-29a-3p, fostering a pro-inflammatory milieu characterized by increased IL-6, CX3CL1, and IL-1β. These alterations coincided with activation of the AKT/CREB pathway, increased inhibitory phosphorylation of GSK3β, accumulation of Aβ1-42 and p-Tau, and diminished trophic support, evidenced by reduced IGF-1, BDNF, and p-TrkB. Notably, valsartan reversed these pathological cascades by suppressing lncRNA H19, restoring miR-29a-3p, attenuating IL-6 and CX3CL1, modulating AKT/CREB & GK3β signaling, reducing Aβ1-42 and p-Tau levels, and enhancing IGF-1, BDNF, and p-TrkB expression. Altogether, the present findings position skeletal muscle as an active orchestrator of FM pathology and highlight AT1 receptor blockade as a promising strategy to interrupt the muscle-to-brain signaling network that sustains chronic pain, fatigue, and impaired muscle performance.

Dendritic cell (DC) cross-presentation is essential for priming tumor-reactive CD8[+] T cells, but small-molecule approaches to enhance this process remain limited. A flow-cytometry-based natural-product screen identified reserpine (Res) as a candidate enhancer of antigen cross-presentation. Res increased surface H-2K[b]-SIINFEKL presentation in MutuDC cells and Flt3L-derived bone marrow DCs, and Res-conditioned DCs promoted OT-I CD8[+] T-cell IFN-γ production and antigen-specific B16-OVA tumor cell killing. In vivo, Res reduced B16 melanoma and MC38 colon tumor growth and improved anti-PD-1 responses in the B16 model. Anti-CD8 and anti-CD11c treatment attenuated Res-associated tumor control, consistent with contributions from CD8[+] T cells and CD11c[+] cells. Mechanistically, the effect persisted when Res was added after OVA pulsing and washing, and Res did not significantly alter direct peptide loading, fluorescent OVA uptake, or DQ-OVA-reported uptake-associated proteolysis under the tested conditions. Res increased 20S proteasome activity, and MG132, epoxomicin and bortezomib reduced Res-enhanced cross-presentation, supporting the involvement of a proteasome-dependent antigen-processing step. These findings identify Res as an immunopharmacological modulator of DC cross-presentation with potential relevance to antitumor immunotherapy.

Fibromyalgia is a chronic nociplastic pain disorder characterized by widespread pain and affective disturbances, in which oxidative stress and neuroinflammation are now recognized as central pathogenic mechanisms. However, current therapies provide only limited symptomatic relief. This study compared the therapeutic and molecular effects of dimethyl fumarate (DMF), an activator of nuclear factor erythroid 2-related factor 2 (NRF2), and cobalt protoporphyrin IX (CoPP), an inducer of heme oxygenase-1 (HO-1), in a reserpine-induced mouse model of fibromyalgia-like pain. Male and female C57BL/6J mice received repeated reserpine administration to induce mechanical allodynia, thermal hyperalgesia, cold allodynia, and depressive-like behaviors. Therapeutic efficacy was evaluated by behavioral testing and molecular analyses of the periaqueductal gray (PAG), anterior cingulate cortex (ACC), and spinal cord (SC). DMF produced a faster antinociceptive response than CoPP, although both compounds ultimately reversed nociceptive hypersensitivity and depressive-like behaviors in male and female mice. These behavioral alterations were associated with increased NLRP3 expression and activated AKT signaling, together with region-specific redox dysregulation involving differential regulation of NADPH oxidases and endogenous antioxidant defenses. Both treatments differentially modulated region-specific redox alterations, enhanced selected antioxidant defenses, and attenuated inflammatory and nociception-related signaling. These findings identify region-specific redox dysregulation as a prominent feature of fibromyalgia-like pathology, providing new mechanistic insight into the molecular basis of nociplastic pain. Collectively, these results demonstrate that DMF and CoPP treatments remodel region-specific redox networks while inhibiting both sensory and affective manifestations of fibromyalgia-like pain, supporting endogenous antioxidant pathways as promising mechanism-based therapeutic targets for fibromyalgia.

Multidrug resistance (MDR) mediated by ATP-binding cassette transporter G2 (ABCG2) remains a major clinical challenge in breast cancer chemotherapy, primarily due to its role in reducing intracellular adriamycin (ADR) concentrations. To address this challenge, we developed an integrated computational strategy for the rapid identification of potent ABCG2 inhibitors capable of reversing ADR resistance. We first established the Prediction of Protein-Ligand's pKd (PPLK), a specialized deep learning model that employs multimodal sequence-ligand representations and feedforward neural networks to accelerate the discovery of ABCG2 inhibitors. Next, computational virtual screening of a compound library (∼1.58 million) was conducted, coupled with molecular dynamics simulations, focusing on the key binding residues THR542, PHE439, ASN436, and SER440. Among the top-ranked candidates, reserpine was validated as a potent ABCG2 inhibitor, achieving 1.90-fold sensitization in ADR-resistant cells, thereby confirming the predictive accuracy of the PPLK model. Notably, we identified a small-molecule compound, F0161-0219, as a promising ABCG2 inhibitor, which led to a 1.94-fold sensitization enhancement in ADR-resistant cells. Direct target engagement of F0161-0219 with ABCG2 was confirmed by biolayer interferometry, DARTS, and CETSA assays. Collectively, our research underscores the potential of leveraging deep learning models and computational screening for the discovery of ABCG2 inhibitors.

Fibromyalgia (FM) is a chronic syndrome characterized by widespread pain, frequently accompanied by cognitive deficits and psychiatric symptoms. FM has been associated with large-scale functional alterations of brain networks involved in pain processing, particularly the default mode network (DMN) and the pain salience network. The infralimbic cortex (IL), a region of the medial prefrontal cortex (mPFC) considered part of the DMN, is functionally connected to regions involved in salience processing. However, the role of the IL in modulating FM-related manifestations remains poorly understood. Therefore, the objective of this study was to investigate the effect of IL neurostimulation on pain modulation and anxiety- and depression-like behaviors in an experimental rat model of FM. IL neurostimulation was performed using a deep-brain stimulation (DBS) device (20μA/100 Hz/15 s) 14 days after the experimental induction of FM using the reserpine pharmacological model (1 mg/kg, s.c., once daily for three consecutive days). IL neurostimulation reduced mechanical allodynia in the von Frey test and decreased both anxiety- and depression-like behaviors assessed using the open field, splash, and forced swim (FST) tests. These findings suggest that the IL plays an inhibitory role in modulating pain and psychiatric comorbidities in FM, suggesting potential circuits between the IL and the salience network that modulate the manifestations of FM.

Cannabidiol (CBD), a major non-psychoactive constituent of Cannabis sativa, exerts pleiotropic effects, including antipsychotic, anxiolytic, and neuroprotective properties, with low toxicity and minimal euphoric effects. Previous studies have demonstrated that CBD alleviates memory impairments associated with pathological conditions such as seizures, reserpine exposure, pneumococcal meningitis, and sepsis. However, its effects on memory processes, particularly inhibitory avoidance (IA) memory reconsolidation and consolidation, remain incompletely understood and inconsistently reported. Therefore, this study investigated the effects of CBD on IA memory reconsolidation and consolidation in mice and explored the underlying neurotrophic mechanisms.

5-Hydroxytryptophan (5-HTP) is a functional precursor of serotonin; however, achieving sustained 5-HTP production in vivo by engineered microbes remains challenging. To address this, we developed a genome-integrated engineered probiotic system enabling gut-local biosynthesis of 5-HTP. Escherichia coli Nissle 1917 (EcN), a clinically validated probiotic with an established safety profile for intestinal application, was selected as the chassis. Constitutive expression was employed to eliminate inducer dependence, while genomic integration was used to enhance genetic stability and avoid reliance on antibiotic selection for pathway maintenance. In addition, BH4 biosynthesis and regeneration modules were incorporated to address the limited endogenous BH4 availability in the gut environment. Different administration strategies were systematically evaluated to characterize in vivo performance. Notably, our results showed that the hTPH2 hydroxylation module, together with the BH4 biosynthesis and regeneration system, supported 5-HTP biosynthesis in EcN. The engineered EcN strain achieved a 5-HTP titer of 863.2 mg/L, the highest level reported in EcN-based systems. Under non-selective conditions, EcN[Pconst-5-HTP] carrying the chromosomally integrated pathway maintained stable 5-HTP production in vitro and retained 5-HTP-producing capacity after intestinal passage, whereas plasmid loss in non-integrated constructs resulted in a pronounced decrease in production. In vivo analysis showed that single-dose administration induced a transient elevation in plasma 5-HTP, whereas repeated dosing generated reproducible, time-dependent fluctuations in fecal bacterial abundance and gut-local biosynthetic activity. Treatment with the engineered probiotic further improved behavioral phenotypes and modulated brain 5-HT-related metabolites in a reserpine-induced mouse model. Together, these findings provide a translational framework linking in vitro 5-HTP production by engineered probiotics to their in vivo functional performance.

Tilia americana var. mexicana is a medicinal species whose reported pharmacological effects primarily target the central nervous system, including anxiolytic, antidepressant, and anticonvulsant activities, attributed in part to flavonoids such as tiliroside (8). In this study, 14 compounds were isolated and identified from the methanolic extract (Ta-MeOH) and evaluated by molecular docking against the monoamine oxidases MAO-A and MAO-B, enzymes implicated in Parkinson's disease (PD) due to their involvement in dopamine metabolism. Chemical analysis identified six terpenes: α- and β-amyrine (1-2), β-sitosterol (3), stigmasterol (4), ursolic acid (5), and β-sitosterol glucoside (6); the novel diglycosylated monoterpene 4α-terpineol sambubioside (7), characterized through its hexaacetate derivative (7a); the flavonoids tiliroside (8) and rutin (9); sucrose (10); and four phenolic compounds: scopoletin (11), caffeic acid (12), coumaric acid (13), and chlorogenic acid (14). Molecular docking against monoamine oxidases A and B (MAO-A and MAO-B) was used as a computational strategy to prioritize isolated metabolites for biological evaluation. β-Sitosterol glucoside (6), 4α-terpineol sambubioside (7), tiliroside (8) and rutin (9) exhibited the most favorable docking scores toward MAO-B, with predicted binding energies of -10.8733, -11.1434, -12.16986 and -12.9474 kcal/mol, respectively. Considering the docking results together with phytochemical and experimental criteria, (6) and (8) (1 mg/kg) were prioritized for biological evaluation, together with Ta-MeOH extract (100 mg/kg) and selected fractions (25 mg/kg), in a reserpine-induced mouse model of parkinsonism using L-DOPA (150 mg/kg) as a positive control. In the open field test, spontaneous locomotor activity was assessed by total crossings and expressed as AUC. Ta-MeOH significantly increased locomotor activity compared with the reserpine-treated group (AUC: 1281.8 vs. 551.7, respectively; p < 0.05). Tiliroside (8) also increased locomotor activity (AUC: 858). In the Rota-Rod test, fine motor coordination was assessed by latency to fall and expressed as AUC. Ta-MeOH showed the greatest recovery, with AUC values of 431, 89, and 34 at 4, 10, and 20 rpm, respectively. In addition, treatments derived from T. americana attenuated behavioral alterations induced by reserpine in the Irwin test. Overall, this study demonstrates that integrating phytochemical isolation, molecular docking, and in vivo pharmacological evaluation provides a useful strategy for prioritizing bioactive metabolites from T. americana. These findings support further pharmacological investigation of this medicinal species and its isolated metabolites, while additional studies are required to establish their molecular targets and mechanisms of action.

Supercritical fluid extraction coupled on-line with supercritical fluid chromatography creates a powerful multidimensional analysis platform, but its adoption is limited by the resource-heavy process of method development. This research presents a surrogate optimization approach as a better alternative to traditional design of experiments and response surface methodology. Unlike fixed experimental designs, the surrogate model iteratively updates the response surface after each run, facilitating effective global optimization through continuous exploration and exploitation. The study adjusts both SFE- and SFC-specific variables-such as back-pressure regulation and split flow-to reduce peak FWHM and enhance extraction efficiency. Analytes, specifically reserpine, vigabatrin, hydromorphone, and hydrocodone, covering diverse physicochemical profiles, were optimized sequentially, guided by a molecular similarity metric, to broaden the design space. The surrogate model identified critical factors, including modifier concentration, chromatography pressure, dynamic extraction time, and flow rate, with adjusted coefficient of determination (R[2]) values reaching 0.97 as more data was collected and the model was refined. Response surface and parallel coordinate analyses showed that hydrocodone, hydromorphone, and vigabatrin favored high modifier concentrations, while reserpine had a distinct optimum more sensitive to static extraction time. Sensitivity analysis over 62.5-1000 ng mL[-1] yielded detection limits/quantification limits of 6.9/23.0 and 2.4/8.1 ng mL[-1] for hydrocodone and reserpine, respectively. Sequential optimization enhanced sensitivity by 15%, demonstrating the scalability and transferability of the surrogate optimization framework for developing SFE-SFC methods.