Alejandro Espinosa

Background/Objectives: Bursera bipinnata (DC.) Engl. resin (locally known as “copal blanco”) is traditionally used in Mexican ethnomedicine to treat infected wounds and skin inflammation, but the bioactive constituents underlying these effects remain largely uncharacterized. This study aimed to identify the compounds responsible for the wound-healing properties of the resin through bioactivity-guided fractionation and to evaluate their anti-inflammatory and antibacterial activities as complementary mechanisms supporting tissue repair. Methods: Crude resin (1.2–5.0 mg/mL) was assayed for anti-inflammatory activity in the TPA-induced ear-edema model in BALB/c mice, for antibacterial activity (MIC) against six clinically relevant strains, and for wound-healing activity in a murine excisional model with pirfenidone (PFD) as the reference drug (n = 5 per group). Bioactivity-guided fractionation followed by spectroscopic elucidation (¹H- and ¹³C-NMR, IR, EI-MS) led to the isolation of five constituents. Stigmasterol, the most active compound, was subsequently evaluated in an LPS-induced systemic inflammation model (oral administration, 20 mg/kg/day × 3 days) to characterize its immunomodulatory profile (TNF-α, IL-1β, IL-6, IFN-γ, IL-10) and in the wound-healing model to quantify local IL-6, IL-10 and TGF-β1 in skin homogenates. Results: The crude resin (5.0 mg/mL) achieved 99.63% wound closure at day 12 and a 49.08% reduction in TPA-induced ear edema, comparable to indomethacin (55.76%). The resin displayed selective antibacterial activity against Streptococcus pyogenes (MIC 125 µg/mL) and Salmonella typhimurium (MIC 250 µg/mL). Bioactivity-guided fractionation yielded the phytosterol stigmasterol (1), three lupane-type triterpenoids (lupeol acetate (2), lupenone (3), 3-epilupeol (5)), and the sesquiterpenoid caryophyllene oxide (4). At an equimolar 1 µM concentration, stigmasterol (1) shortened the mean wound-healing time to 10.3 ± 0.4 days, comparable to pirfenidone, and was associated with attenuation of systemic TNF-α, IL-1β and IL-6 peaks and with sustained local IL-10 and TGF-β1 expression. Histological assessment confirmed accelerated re-epithelialization and improved collagen organization. The resin was non-irritant in the OECD 404 acute dermal test (Primary Irritation Index = 0.00). Conclusions: These findings provide pharmacological evidence supporting the traditional use of B. bipinnata resin for wound healing. Stigmasterol (1), together with the lupane-type triterpenoids lupenone (3) and 3-epilupeol (5), were identified as key bioactive constituents. The data are consistent with a coordinated immunomodulation, in which stigmasterol is associated with reduced systemic pro-inflammatory signalling and increased local IL-10/TGF-β1 expression, an interpretation that should be confirmed in chronic and impaired wound-healing models.

Metabolomic profiling of the leaves, floral buds, flowers, and fruits of Ternstroemia lineata was conducted across the dry and rainy seasons in Mexico. The presence of katsumadin was determined and its antibacterial activity was evaluated. Multivariate data analysis was performed using proton nuclear magnetic resonance (¹H NMR) and Principal Component Analysis (PCA). Katsumadin was identified by ¹H NMR and, quantified by HPLC, and its antibacterial activity was assessed using a microdilution assay. The results revealed conserved ¹H NMR signals among leaf, flower buds, flowers and fruits, as well as signals that were either organ-specific or time-dependent. The spatial and temporal distribution of katsumadin was monitored over a nine-month period. During the dry season, katsumadin reached 1.5 mg/g DW in the leaves, whereas it was not detected during the rainy season. Higher katsumadin contents were observed in floral buds, fruits, and flowers (6.09 ± 0.10, 3.0 ± 0.79, and 3.4 ± 0.42 mg/g DW, respectively). In addition, katsumadin was evaluated against Salmonella Typhi and Pseudomonas aeruginosa, exhibiting a minimum inhibitory concentration and bactericidal effect at 0.5 mg/mL.

Neurocysticercosis is caused by the establishment of Taenia solium cysticerci in the central nervous system. The extraparenchymal form (ExP-NCC) is the most severe clinical presentation that may remain asymptomatic for years. Current treatment involves cysticidal drugs (albendazole and/or praziquantel) combined with glucocorticoids to manage the associated neuroinflammation; however, only ∼30% of patients respond effectively. This highlights the need to improve therapeutic strategies. Herein, the experimental murine model of human ExP-NCC was further characterized to improve its usefulness in testing new therapies. In humans, cysts grow slowly in the basal cisterns of the subarachnoid space, and patients become symptomatic years after the infection. Thus, a long-term follow-up was performed by using magnetic resonance imaging (MRI) with sequences allowing volumetric analysis. MRI confirmed NCC in 77% of infected rats, all exhibiting extraparenchymal localization and persistently elevated levels of HP10, a marker of viable cysticerci. Imaging also enabled precise cyst localization and estimation of the parasite-occupied volume.

Background: Multiple sclerosis (MS) is a demyelinating, neuroinflammatory, progressive disease that severely affects human health of young adults. Neuroinflammation (NI) and demyelination, as well as their interactions, are key therapeutic targets to halt or slow disease progression. Potent steroidal anti-inflammatory drugs such as methylprednisolone (MP) and remyelinating neurosteroids such as allopregnanolone (ALLO) could be co-administered intranasally to enhance their efficacy by providing direct access to the central nervous system (CNS). Methods: The individual and combined effects of MP and ALLO to control the clinical score of murine experimental autoimmune encephalitis (EAE), to preserve spinal cord tissue integrity, modulate cellular infiltration and gliosis, promote remyelination, and modify the expression of Aryl hydrocarbon receptor (AhR) were evaluated. In silico studies, to deep insight into the mechanisms involved for the treatments, were also conducted. Results: MP was the only treatment that significantly reduced the EAE severity, infiltration of inflammatory cells and ionized calcium-binding adapter molecule 1 (Iba-1) expression respect to those EAE non-treated mice but with no-significant differences between the three treatments. MP, ALLO and MP+ALLO significantly reduced tissue damage, AhR expression, and promoted remyelination. Overall, these results suggest that MP, with or without the co-administration with ALLO is an effective and safe strategy to reduce the inflammatory status and the progression of EAE. Despite the expectations of the use of ALLO to reduce the inflammation in EAE, its effect in the dose-scheme used herein is limited only to improve myelination, an effect that supports its usefulness in demyelinating diseases. These results indicate the interest in exploring different doses of ALLO to recommend its use. Conclusions: ALLO treatment mainly maintain the integrity of the spinal cord tissue and the presence of myelin without affecting NI and the clinical outcome. AhR could be involved in the effect observed in both, MP and ALLO treatments. These results will help in the development of a more efficient therapy for MS patients.

Background: Neurocysticercosis (NCC) is endemic in non-developed regions of the world. Two forms of NCC have been described, for which neurological morbidity depends on the location of the lesion, which can be either within the cerebral parenchyma or in extraparenchymal spaces. The extraparenchymal form (EXP-NCC) is considered the most severe form of NCC. EXP-NCC often requires several cycles of cysticidal treatment and the concomitant use of glucocorticoids to prevent increased inflammation, which could lead to intracranial hypertension and, in rare cases, to death. Thus, the improvement of EXP-NCC treatment is greatly needed. Methods: An experimental murine model of EXP-NCC, as an adequate model to evaluate new therapeutic approaches, and the parameters that support it are described. EXP-NCC was established by injecting 30 Taenia crassiceps cysticerci, which are less than 0.5 mm in diameter, into the cisterna magna of male and female Wistar rats. Results: Cyst implantation and infection progression were monitored by detecting the HP10 antigen and anti-cysticercal antibodies in the serum and cerebral spinal fluid (CSF) of infected rats and by magnetic resonance imaging. Higher HP10 levels were observed in CSF than in the sera, as in the case of human EXP-NCC. Low cell recruitment levels were observed surrounding established cysticerci in histological analysis, with a modest increase in GFAP and Iba1 expression in the parenchyma of female animals. Low cellularity in CSF and low levels of C-reactive protein are consistent with a weak inflammatory response to this infection. After 150 days of infection, EXP-NCC is accompanied by reduced levels of mononuclear cell proliferation, resembling the human disease. EXP-NCC does not affect the behavior or general status of the rats. Conclusions: This model will allow the evaluation of new approaches to control neuroinflammation and immunomodulatory treatments to restore and improve the specific anti-cysticercal immunity in EXP-NCC.

Dexamethasone (DXM) and methylprednisolone (MEP) are potent glucocorticoids used to control several inflammatory conditions. Evidence of delayed DXM reaching the central nervous system (CNS) as well as tachyphylaxis and systemic, undesirable side effects are the main limitations of peripheral delivery. Intranasal administration offers direct access to the brain as it bypasses the blood–brain barrier. The Mucosal Atomization Device is an optimal tool that can achieve rapid absorption into the CNS and the bloodstream across mucosal membranes. This study was designed to evaluate and compare the bioavailability of DXM and MEP after intranasal versus intravenous administration. Two open-label, balanced, randomized, two-treatment, two-period, two-sequence, single-dose, crossover studies were conducted, which involved healthy male and female adult volunteers. After intranasal administration, DXM and MEP were detected in plasma after the first sampling time. Mean peak concentrations of DXM and MEP were 86.61 ng/mL at 60 min and 843.2 ng/mL at 1.5 h post-administration, respectively. DXM and MEP showed high absolute bioavailability, with values of 80% and 95%, respectively. No adverse effects were observed. DXM and MEP systemic bioavailability by intranasal administration was comparable with the intravenous one, suggesting that the intranasal route can be used as a non-invasive and appropriate alternative for systemic drug delivery.

Inhalants are chemical substances that induce intoxication, and toluene is the main component of them. Increasing evidence indicates that a dependence on inhalants involves a state of chronic stress associated to the activation of immune cells in the central nervous system and release of proinflammatory mediators, especially in some brain areas such as the nucleus accumbens and frontal cortex, where the circuits of pleasure and reward are. In this study, anti-neuroinflammatory treatment based on a single dose of intranasal methylprednisolone was assessed in a murine model of chronic toluene exposure. The levels of proinflammatory mediators, expression levels of Iba-1 and GFAP, and histological changes in the frontal cortex and nucleus accumbens were evaluated after the treatment. The chronic exposure to toluene significantly increased the levels of TNF-α, IL-6, and NO, the expression of GFAP, and induced histological alterations in mouse brains. The treatment with intranasally administered MP significantly reduced the expression of TNF-α and NO and the expression of GFAP (p < 0.05); additionally, it reversed the central histological damage. These results indicate that intranasally administered methylprednisolone could be considered as a treatment to reverse neuroinflammation and histological damages associated with the use of inhalants.

Neuroinflammation (NI) is an important physiological process which promotes the tissue repair and homeostatic maintenance in the central nervous system (CNS) after different types of insults. However, when it is exacerbated and sustained in time, NI plays a critical role in the pathogenesis of different neurological diseases. The high systemic doses required for brain-specific targeting leads to severe undesirable effects. The intranasal (IN) route has been proposed as an alternative drug administration route for a better NI control. Herein, the brain biodistribution of intranasally administered dexamethasone (Dex) versus intravenous administered one is reported. A higher amount of dexamethasone was found in every brain analyzed regions of those brains of intranasally administered mice. HPLC analysis also revealed that IN administration allows Dx arrives faster and in a greater concentration to the brain in comparison with intravenous administration, data confirmed by immunofluorescence and HPLC analysis. These data support the proposal of the IN administration of Dx as an alternative for a more efficiently control of NI. Significance Statement This work highlights the biodistribution of dexamethasone after its intranasal administration. Intranasal administration allows for a faster arrival, better distribution and a higher concentration of the drug within the brain compared to its intravenous administration. These results explain some of the evidence shown by us in a previous work in which dexamethasone controls neuroinflammation in a murine stroke model and can be used to propose alternative treatments for neuroinflammatory diseases.