An overview on recent patents and technologies on nanoparticles for nucleic acid delivery.

INTRODUCTION: Nucleic acid-based therapeutics offer groundbreaking potential for treating genetic diseases and advancing next-generation vaccines. Despite their promise, challenges in efficient delivery persist due to the properties of nucleic acids. Nanoparticles (NPs) serve as vital carriers, facilitating effective delivery to target cells, and addressing these challenges. Understanding the global landscape of patents in this field is essential for fostering innovation and guiding decision-making for researchers, the pharmaceutical industry, and regulatory agencies. AREAS COVERED: This review provides a comprehensive overview of patent compositions, applications, and manufacturing aspects concerning NPs as nucleic acid delivery systems. It delves into temporal trends, protection locations, market dynamics, and the most influential technological domains. In this work, we provide valuable insights into the advancements and potential of NP-based nucleic acid delivery systems, with a special focus on their pivotal role in advancing cutting-edge therapeutic solutions. EXPERT OPINION: Investment in NPs for nucleic acid delivery has significantly surged in recent years. However, translating these therapies into clinical practice faces obstacles, including the need for robust clinical evidence, regulatory compliance, and streamlined manufacturing processes. To address these challenges, our review article summarizes recent advances. We aim to engage researchers worldwide in the development of these promising technologies.

High-Content Imaging-Based Assay for SARS-CoV-2-Neutralizing Antibodies.

The COVID-19 pandemic and the consequent emergence of new SARS-CoV-2 variants of concern necessitates the determination of populational serum potency against the virus. Here, we standardized and validated an imaging-based method to quantify neutralizing antibodies against lentiviral particles expressing the spike glycoprotein (pseudovirus). This method was found to efficiently quantify viral titers based on ZsGreen-positive cells and detect changes in human serum neutralization capacity induced by vaccination with up to two doses of CoronaVac, Comirnaty, or Covishield vaccines. The imaging-based protocol was also used to quantify serum potency against pseudoviruses expressing spikes from Delta, Omicron BA.1.1.529, and BA.4/5. Our results revealed increases in serum potency after one and two doses of the vaccines evaluated and demonstrated that Delta and Omicron variants escape from antibody neutralization. The method presented herein represents a valuable tool for the screening of antibodies and small molecules capable of blocking viral entry and could be used to evaluate humoral immunity developed by different populations and for vaccine development.

Solidagenone from Solidago chilensis Meyen Protects against Acute Peritonitis and Lipopolysaccharide-Induced Shock by Regulating NF-κB Signaling Pathway.

Anti-inflammatory agents are widely used for the treatment of inflammatory diseases. Nevertheless, the associated side effects of the available drugs make it necessary to search for new anti-inflammatory drugs. Here, we investigated the anti-inflammatory activity of solidagenone. Initially, we observed that a single dose of 30, 60, or 90 mg/kg of solidagenone did not result in mortality or elicit any discernible signs of toxicity in mice. At the same doses, solidagenone promoted a significant reduction in the migration of neutrophils in an acute peritonitis model and decreased mortality in a lipopolysaccharide-induced endotoxic shock model. Interestingly, treatment with solidagenone conferred a protective effect against leukopenia and thrombocytopenia, hematological disorders commonly observed in sepsis conditions. In addition, treatment with all the doses of solidagenone promoted a significant reduction in nitric oxide, TNF-α, and IL-1ß levels relative to the LPS-stimulated vehicle-treated cultures. Furthermore, gene expression and in silico analyses also supported the modulation of the NF-κB pathway by solidagenone. Finally, in silico pharmacokinetics predictions indicated a favorable drugability profile for solidagenone. Taken together, the findings of the present investigation show that solidagenone exhibits significant anti-inflammatory properties in acute experimental models, potentially through the modulation of the NF-κB signaling pathway.

Ayahuasca and its major component harmine promote antinociceptive effects in mouse models of acute and chronic pain.

ETHNOPHARMACOLOGICAL RELEVANCE: Ayahuasca (AYA) is a psychedelic brew used in religious ceremonies. It is broadly used as a sacred medicine for treating several ailments, including pain of various origins. AIM OF THE STUDY: To investigate the antinociceptive effects of AYA and its mechanisms in preclinical models of acute and chronic pain in mice, in particular during experimental neuropathy. MATERIALS AND METHODS: The antinociceptive effects of AYA administered orally were assessed in the following models of pain: formalin test, Complete Freund's Adjuvant (CFA)-induced inflammation, tail flick test, and partial sciatic nerve ligation model of neuropathic pain. Antagonism assays and Fos immunohistochemistry in the brain were performed. AYA-induced toxicity was investigated. AYA was chemically characterized. The antinociceptive effect of harmine, the major component present in AYA, was investigated. RESULTS: AYA (24-3000 µL/kg) dose-dependently reduced formalin-induced pain-like behaviors and CFA-induced mechanical allodynia but did not affect CFA-induced paw edema or tail flick latency. During experimental neuropathy, single treatments with AYA (24-3000 µL/kg) reduced mechanical allodynia; daily treatments once or twice a day for 14 days promoted consistent and sustained antinociception. The antinociceptive effect of AYA (600 µL/kg) was reverted by bicuculline (1 mg/kg) and methysergide (5 mg/kg), but not by naloxone (5 mg/kg), phaclofen (2 mg/kg), and rimonabant (10 mg/kg), suggesting the roles of GABAA and serotonergic receptors. AYA increased Fos expression in the ventrolateral periaqueductal gray and nucleus raphe magnus after 1 h, but not after 6 h or 14 days of daily treatments. AYA (600 µL/kg) twice a day for 14 days did not alter mice's motor function, spontaneous locomotion, body weight, food and water intake, hematological, biochemical, and histopathological parameters. Harmine (3.5 mg/kg) promoted consistent antinociception during experimental neuropathy. CONCLUSIONS: AYA promotes consistent antinociceptive effects in different mouse models of pain without inducing detectable toxic effects. Harmine is at least partially accountable for the antinociceptive properties of AYA.

Exotic fruits patents trends: An overview based on technological prospection with a focus on Amazonian.

The Amazon rainforest encompasses one of the largest biodiversities of the world and is home to a wide variety of food and therapeutic plants. Due to the diversity of components, the fruits of the Amazon biome possess essential physicochemical, nutritional, and pharmacological properties, strengthening the idea that fruit consumption may provide benefits to human health. Thus, the objective of this study was to investigate the current scenario of the use of Amazonian fruits on the development of food, pharmaceutical, nutraceutical, or cosmetic products through the study of filed patents. A prospecting strategy conducted focusing on patents was used to investigate the application of the following fruits: Euterpe oleracea, Oenocarpus bacaba, Caryocar brasiliense, Garcinia gardneriana, Nephelium lappaceum, and Astrocaryum vulgare. A total of 264 patent documents were found. In 2016, a peak of 33 applications was reached, followed by a peak in 2019 with 32 applications. The study is distributed in three main application areas: cosmetics, pharmaceuticals, and food. The Asian continent was the region with the world leadership in this theme, followed by Brazil. Thus, technological prospection studies can foster investments in translational research to elucidate the effects and properties of Amazonian fruits, which can generate sustainable development of new products with industrial potential.

Cleomin Exerts Acute Antinociceptive Effects in Mice via GABAB and Muscarinic Receptors.

Cleomin, a 1,3-oxazolidine-2-thione, was recently isolated from Neocalyptrocalyx longifolium, a species traditionally used for treating painful conditions. Reports about the pharmacological activities of cleomin are lacking. Here, the antinociceptive effects of cleomin were investigated using mice models of pain, namely the formalin, the cold plate, and the tail flick tests. Motor integrity was assessed in the rota-rod test. Antagonism assays and in silico docking analyses were performed to investigate the putative mechanisms of action. Cleomin (12.5-25 mg/kg), at doses that did not induce motor impairment, induced dose-dependent antinociception in both early and late phases of the formalin test and reduced nociceptive behaviors in both the cold plate and tail flick tests. Pretreatments with phaclofen and atropine attenuated the antinociceptive effects of cleomin, implicating the involvement of GABAB and muscarinic receptors. In silico docking studies suggested satisfactory coupling between cleomin and GABAB and M2 receptors, hence corroborating their role in cleomin's activity. Pretreatments with naloxone, yohimbine, bicuculline, and methysergide did not affect the antinociception of cleomin. In silico pharmacokinetics prediction showed a good drug ability profile of cleomin. In conclusion, cleomin promoted antinociception mediated by GABAB and muscarinic receptors. These findings support further investigation of the analgesic potential of cleomin.

Antinociceptive and anti-inflammatory properties of α-D-mannan from the yeast Kluyveromyces marxianus: evidence for a role in interleukin-6 inhibition.

The management of inflammatory states typically involves non-steroidal anti-inflammatory drugs (NSAIDs) and opiates. Understanding the mechanisms underlying the processing of nociceptive information from potential alternatives such as some polysaccharides may enable new and meaningful therapeutic approaches. In this study, α-D-mannan isolated from the Kluyveromyces marxianus cell wall produced antinociceptive effects in models of inflammatory pain (formalin and complete Freund's adjuvant tests). Furthermore, α-D-mannan reduced paw edema and interleukin-6 (IL-6) production after carrageenan-induced inflammation. The polysaccharide α-D-mannan was characterized by gas chromatography-mass spectrometry, methylation analysis, and spectroscopic techniques. Moreover, the Doehlert experimental design was applied to find the optimal conditions for biomass production, with the best conditions being 10.8 g/L and 117 h for the glucose concentration and the fermentation time, respectively. These results indicate that α-D-mannan from K. marxianus exerts anti-inflammatory and antinociceptive effects in mice, possibly via a mechanism dependent on the inhibition of IL-6 production.

Therapeutic Potential of Natural Products in the Treatment of Schistosomiasis.

It is estimated that 250 million people worldwide are affected by schistosomiasis. Disease transmission is related to the poor sanitation and hygiene habits that affect residents of impoverished regions in tropical and subtropical countries. The main species responsible for causing disease in humans are Schistosoma Mansoni, S. japonicum, and S. haematobium, each with different geographic distributions. Praziquantel is the drug predominantly used to treat this disease, which offers low effectiveness against immature and juvenile parasite forms. In addition, reports of drug resistance prompt the development of novel therapeutic approaches. Natural products represent an important source of new compounds, especially those obtained from plant sources. This review compiles data from several in vitro and in vivo studies evaluating various compounds and essential oils derived from plants with cercaricidal and molluscicidal activities against both juvenile and adult forms of the parasite. Finally, this review provides an important discussion on recent advances in molecular and computational tools deemed fundamental for more rapid and effective screening of new compounds, allowing for the optimization of time and resources.

DNA methylation-based depiction of the immune microenvironment and immune-associated long non-coding RNAs in oral cavity squamous cell carcinomas.

Oral cavity squamous cell carcinoma (OSCC) is a complex and dynamic disease characterized by clinicopathological and molecular heterogeneity. Spatial and temporal heterogeneity of cell subpopulations has been associated with cancer progression and implicated in the prognosis and therapy response. Emerging evidence indicates that aberrant epigenetic profiles in OSCC may foster an immunosuppressive tumor microenvironment by modulating the expression of immune-related long non-coding RNAs (lncRNAs). DNA methylation analysis was performed in 46 matched OSCC and normal adjacent tissue samples using a genome-wide platform (Infinium HumanMethylation450 BeadChip). Reference-based computational deconvolution (MethylCIBERSORT) was applied to infer the immune cell composition of the bulk samples. The expression levels of genes encoding immune markers and differentially methylated lncRNAs were investigated using The Cancer Genome Atlas dataset. OSCC specimens presented distinct immune cell composition, including the enrichment of monocyte lineage cells, natural killer cells, cytotoxic T-lymphocytes, regulatory T-lymphocytes, and neutrophils. In contrast, B-lymphocytes, effector T-lymphocytes, and fibroblasts were diminished in tumor samples. The hypomethylation of three immune-associated lncRNAs (MEG3, MIR155HG, and WFDC21P) at individual CpG sites was confirmed by bisulfite-pyrosequencing. Also, the upregulation of a set of immune markers (FOXP3, GZMB, IL10, IL2RA, TGFB, IFNG, TDO2, IDO1, and HIF1A) was detected. The immune cell composition, immune markers alteration, and dysregulation of immune-associated lncRNAs reinforce the impact of the immune microenvironment in OSCC. These concurrent factors contribute to tumor heterogeneity, suggesting that epi-immunotherapy could be an efficient alternative to treat OSCC.

A randomized clinical trial to evaluate the efficacy of L-carnitine L-tartrate to modulate the effects of SARS-CoV-2 infection.

Introduction: L-carnitine (LC) has been associated with inflammatory mediator reduction and with downregulating the angiotensin-converting enzyme-2 (ACE2) receptor, which is the target of SARS-CoV-2 attachment. Methods: This pilot phase 2 randomized, double-blind placebo-controlled trial contained two cohorts. Cohort 1 comprised 101 individuals with negative RT-PCR SARS-CoV-2 test results who cohabitated with an individual diagnosed with SARS-CoV-2 infection. Cohort 2 comprised 122 individuals with positive SARS-CoV-2 RT-PCR test results who were asymptomatic or had mild COVID-19 pneumonia symptoms. Participants in each cohort were randomized 1:1 to receive either 2 g elemental oral LC supplementation or placebo daily for 21 days. Primary endpoints included adverse events, SARS-CoV-2 infection incidence in Cohort 1, and disease progressions in Cohort 2. Secondary endpoints included between-group laboratory profile comparisons and Cohort 2 ACE1/ACE2 plasma levels. Disease progression was compared between the Cohort 2 groups using chest computed tomography. Results: In Cohort 1, two SARS-CoV-2 infections occurred in each group. The common adverse events included headache, dyspnea, and tiredness. In Cohort 2, platelet counts were elevated, and fibrinogen levels reduced in the LC group compared with those of the placebo group. Conclusion: Our study showed that LC was well-tolerated and suggests it modulates coagulation pathways. Furthermore, chest computed tomography images of the Cohort 2 LC group showed significant lung lesion improvement, suggesting that LC may slow COVID-19 progression.

An Overview of the Conventional and Novel Methods Employed for SARS-CoV-2 Neutralizing Antibody Measurement.

SARS-CoV-2 is the etiological agent of the coronavirus disease-19 (COVID-19) and is responsible for the pandemic that started in 2020. The virus enters the host cell through the interaction of its spike glycoprotein with the angiotensin converting enzyme-2 (ACE2) on the host cell's surface. Antibodies present an important role during the infection and pathogenesis due to many reasons, including the neutralization of viruses by binding to different spike epitopes. Therefore, measuring the neutralizing antibody titers in the whole population is important for COVID-19's epidemiology. Different methods are described in the literature, and some have been used to validate the main vaccines used worldwide. In this review, we discuss the main methods used to quantify neutralizing antibody titers, their advantages and limitations, as well as new approaches to determineACE2/spike blockage by antibodies.

Clinical Trials with Mesenchymal Stem Cell Therapies for Osteoarthritis: Challenges in the Regeneration of Articular Cartilage.

Osteoarthritis (OA) is a whole-joint disease primarily characterized by the deterioration of hyaline cartilage. Current treatments include microfracture and chondrocyte implantation as early surgical strategies that can be combined with scaffolds to repair osteochondral lesions; however, intra-articular (IA) injections or implantations of mesenchymal stem cells (MSCs) are new approaches that have presented encouraging therapeutic results in animal models and humans. We critically reviewed clinical trials with MSC therapies for OA, focusing on their effectiveness, quality, and outcomes in the regeneration of articular cartilage. Several sources of autologous or allogeneic MSCs were used in the clinical trials. Minor adverse events were generally reported, indicating that IA applications of MSCs are potentially safe. The evaluation of articular cartilage regeneration in human clinical trials is challenging, particularly in the inflammatory environment of osteoarthritic joints. Our findings indicate that IA injections of MSCs are efficacious in the treatment of OA and the regeneration of cartilage, but that they may be insufficient for the full repair of articular cartilage defects. The possible interference of clinical and quality variables in the outcomes suggests that robust clinical trials are still necessary for generating reliable evidence with which to support these treatments. We suggest that the administration of just-sufficient doses of viable cells in appropriate regimens is critical to achieve effective and durable effects. In terms of future perspectives, genetic modification, complex products with extracellular vesicles derived from MSCs, cell encapsulation in hydrogels, and 3D bioprinted tissue engineering are promising approaches with which to improve MSC therapies for OA.

Toxic profile of marinobufagin from poisonous Amazon toads and antitumoral effects on human colorectal carcinomas.

ETHNOPHARMACOLOGICAL RELEVANCE: South Americans natives have extensively used the toad "kururu" to reduce/treat skin infections, cutaneous lesions and sores. They release secretions rich in bufadienolides, polyhydroxy steroids with well-documented cardiotonic and antiproliferative actions, but in vivo antitumoral evaluations in mammals are rare, and toxicological safety has been left in second place. AIMS OF THE STUDY: This investigation used in silico, in vitro and in vivo tools to evaluate acute and subacute toxic effects of marinobufagin and the anticancer action in tumor-bearing mice models. MATERIALS AND METHODS: Initially, in silico toxic predictions were performed, followed by in vitro assays using human and murine normal and tumor lines. Next, acute and subacute studies on mice investigated the behavior, hematological and intestinal transit profile and antitumoral activity of marinobufagin in sarcoma 180- and HCT-116 colorectal carcinoma-transplanted mice for 7 and 15 days, respectively. Ex vivo and in vivo cytogenetic assays in Sarcoma 180 and bone marrow cells and histopathological examinations were also executed. RESULTS: In silico studies revealed ecotoxicological effects on crustaceans (Daphnia sp.), fishes (Pimephales promelas and Oryzias latipes), and algae. A 24-h marinobufagin-induced acute toxicity included signals of central activity, mainly (vocal frenzy, absence of body tonus, increased ventilation, ataxia, and equilibrium loss), and convulsions and death at 10 mg/kg. The bufadienolide presented effective in vitro cytotoxic action on human lines of colorectal carcinomas in a similar way to ouabain and tumor reduction in marinobufagin-treated SCID-bearing HCT-116 heterotopic xenografts. Animals under subacute nonlethal doses exhibited a decrease in creatinine clearance with normal levels of blood urea, probably as a result of a marinobufagin-induced renal perfusion fall. Nevertheless, only minor morphological side effects were identified in kidneys, livers, hearts and lungs. CONCLUSIONS: Marinobufagin has in vitro and in vivo anticancer action on colorectal carcinoma and mild and reversible alterations in key metabolic organs without direct chemotherapy-induced gastrointestinal effects at subacute exposure, but it causes acute ataxia, equilibrium loss, convulsions and death at higher acute exposure.

Myocardial infarction or myocarditis? A case report and review of a myocardial adverse event associated with mRNA vaccine.

A 23-year-old man started with chest pain 8 h after his first Pfizer-BioNTech COVID-19 vaccination. ECG evaluation showed sinus tachycardia with ST-segment elevation in D1, AVL, V5, and V6, the findings compatible with acute subepicardial myocardial damage. However, cardiac MRI documented myocardial fibrosis, with cardiac late enhancement non-ischemic pattern with diffuse edema. He had no other symptoms to suggest another etiology than the vaccination. The patient was hospitalized and received corticosteroid (prednisolone) daily. Then, 2 weeks after hospitalization, all laboratory parameters and ECG were normal and the patient was discharged from the hospital. The patient had a history of Wolf-Parkinson White that was corrected with ablation when he was 11 years old. This report calls attention to myocardial adverse reaction risk for mRNA COVID-19 vaccines for people with a previous cardiac disease history.

Antileshimania, anti-Trypanosoma cruzi and antimicrobial activities of scandenin and 4'-O-methylderrone from Deguelia costata.

Scandenin and 4'-O-methylderrone were isolated from the ethanol extract of the roots and dichloromethane extract of the leaves of Deguelia costata (Benth.) A.M.G. Azevedo & R.A. Camargo, respectively. These compounds and their extracts had their antiprotozoal, antibacterial, antifungal, and cytotoxic activities tested. All samples were active for amastigotes of the T. cruzi, with EC50 values varying from 34.5 to 9.8 µg mL-1. The 4'-O-methylderrone and scandenin showed better leishmanicidal action against the promastigote of L. amazonensis, with EC50 of 43.3 and 45.9 µg mL-1, respectively, when compared to their extracts. All extracts and scandenin showed activities against Staphylococcus sp, Bacillus sp, and Candida sp. The compounds did not show cytotoxicity on rat macrophages. As confirmed by spectroscopic analyses, the extracts are rich in phenolics, mainly isoflavonoids. The study of D. costata is a promising strategy for discovering isoflavones and 4-hydroxy-3-phenylcoumarins with antiprotozoal, antibacterial, and antifungal activities.

Natural Dibenzo[b,f]oxepines, Pacharin and Bauhiniastatin-1, Isolated from Bauhinia acuruana Induce Apoptosis on Breast Cancer Cells via MCL-1 Protein Reduction.

Herein, we describe the antiproliferative effects of two natural dibenzo [b,f]oxepines, pacharin and bauhiniastatin-1, isolated from Bauhinia acuruana on a breast cancer cell line and the mode of action underlying the cytotoxicity. Both compounds were cytotoxic in a panel of six tumor lines analyzed by the MTT assay, and IC50 values ranged from 7.8 to 45.1 µM, including human breast adenocarcinoma (MCF-7) cells. In contrast, none of the compounds were cytotoxic on normal human peripheral blood mononuclear cells (IC50 > 100 µM). Human breast adenocarcinoma (MCF-7) cells treated with pacharin or bauhiniastatin-1 20 µM for 24 h presented a reduction in cell volume and intensification of chromatin condensation, DNA fragmentation, and apoptotic cells. These findings became more evident after 48 h of exposure. Antiapoptotic B-cell lymphoma-2 family members, such as myeloid cell leukemia-1 and B-cell lymphoma-extra large, are important targets in cancer cells since their overexpression confers resistance to cancer treatments. A significant reduction of the myeloid cell leukemia-1 protein levels in human breast adenocarcinoma (MCF-7) cells after 24 h of treatment with pacharin or bauhiniastatin-1 at 20 µM was observed, while the B-cell lymphoma-extra large protein content was reduced in bauhiniastatin-1-treated cells at 40 µM only. The cytotoxic effects of pacharin and bauhiniastatin-1 are likely linked to myeloid cell leukemia-1 inhibition, which leads to the apoptosis of breast adenocarcinoma cells.

Molecular Hybridization Strategy on the Design, Synthesis, and Structural Characterization of Ferrocene-N-acyl Hydrazones as Immunomodulatory Agents.

Immunomodulatory agents are widely used for the treatment of immune-mediated diseases, but the range of side effects of the available drugs makes necessary the search for new immunomodulatory drugs. Here, we investigated the immunomodulatory activity of new ferrocenyl-N-acyl hydrazones derivatives (SintMed(141−156). The evaluated N-acyl hydrazones did not show cytotoxicity at the tested concentrations, presenting CC50 values greater than 50 µM. In addition, all ferrocenyl-N-acyl hydrazones modulated nitrite production in immortalized macrophages, showing inhibition values between 14.4% and 74.2%. By presenting a better activity profile, the ferrocenyl-N-acyl hydrazones SintMed149 and SintMed150 also had their cytotoxicity and anti-inflammatory effect evaluated in cultures of peritoneal macrophages. The molecules were not cytotoxic at any of the concentrations tested in peritoneal macrophages and were able to significantly reduce (p < 0.05) the production of nitrite, TNF-α, and IL-1ß. Interestingly, both molecules significantly reduced the production of IL-2 and IFN-γ in cultured splenocytes activated with concanavalin A. Moreover, SintMed150 did not show signs of acute toxicity in animals treated with 50 or 100 mg/kg. Finally, we observed that ferrocenyl-N-acyl hydrazone SintMed150 at 100 mg/kg reduced the migration of neutrophils (44.6%) in an acute peritonitis model and increased animal survival by 20% in an LPS-induced endotoxic shock model. These findings suggest that such compounds have therapeutic potential to be used to treat diseases of inflammatory origin.

Unveiling Targets for Treating Postoperative Pain: The Role of the TNF-α/p38 MAPK/NF-κB/Nav1.8 and Nav1.9 Pathways in the Mouse Model of Incisional Pain.

Although the mouse model of incisional pain is broadly used, the mechanisms underlying plantar incision-induced nociception are not fully understood. This work investigates the role of Nav1.8 and Nav1.9 sodium channels in nociceptive sensitization following plantar incision in mice and the signaling pathway modulating these channels. A surgical incision was made in the plantar hind paw of male Swiss mice. Nociceptive thresholds were assessed by von Frey filaments. Gene expression of Nav1.8, Nav1.9, TNF-α, and COX-2 was evaluated by Real-Time PCR in dorsal root ganglia (DRG). Knockdown mice for Nav1.8 and Nav1.9 were produced by antisense oligodeoxynucleotides intrathecal treatments. Local levels of TNF-α and PGE2 were immunoenzymatically determined. Incised mice exhibited hypernociception and upregulated expression of Nav1.8 and Nav1.9 in DRG. Antisense oligodeoxynucleotides reduced hypernociception and downregulated Nav1.8 and Nav1.9. TNF-α and COX-2/PGE2 were upregulated in DRG and plantar skin. Inhibition of TNF-α and COX-2 reduced hypernociception, but only TNF-α inhibition downregulated Nav1.8 and Nav1.9. Antagonizing NF-κB and p38 mitogen-activated protein kinase (MAPK), but not ERK or JNK, reduced both hypernociception and hyperexpression of Nav1.8 and Nav1.9. This study proposes the contribution of the TNF-α/p38/NF-κB/Nav1.8 and Nav1.9 pathways to the pathophysiology of the mouse model of incisional pain.

Anti-Inflammatory Activities of Betulinic Acid: A Review.

Inflammatory diseases have a high prevalence and has become of great interest due to the increase in life expectancy and the costs to the health care system worldwide. Chronic diseases require long-term treatment frequently using corticosteroids and non-steroidal anti-inflammatory drugs, which are associated with diverse side effects and risk of toxicity. Betulinic acid, a lupane-type pentacyclic triterpene, is a potential lead compound for the development of new anti-inflammatory treatments, and a large number of derivatives have been produced and tested. The potential of betulinic acid and its derivatives has been shown in a number of pre-clinical studies using different experimental models. Moreover, several molecular mechanisms of action have also been described. Here we reviewed the potential use of betulinic acid as a promissory lead compound with anti-inflammatory activity and the perspectives for its use in the treatment of inflammatory conditions.

A Betulinic Acid Derivative, BA5, Induces G0/G1 Cell Arrest, Apoptosis Like-Death, and Morphological Alterations in Leishmania sp.

Leishmaniasis are endemic diseases caused by different species of intracellular parasites of the genus Leishmania. Due to the high toxicity and drug resistance of current antileishmanial drugs, it is necessary to identify new and more effective drugs. Previously, we investigated the immunomodulatory and anti-Trypanosoma cruzi action of BA5, a derivative of betulinic acid. In the present study, we investigated the in vitro activity of BA5 against different species of Leishmania and their action mechanism. BA5 exhibited low cytotoxicity against macrophages and inhibited the proliferation of promastigote forms of Leishmania amazonensis (IC50 = 4.5 ± 1.1 µM), Leishmania major (IC50 = 3.0 ± 0.8 µM), Leishmania braziliensis (IC50 = 0.9 ± 1.1 µM) and Leishmania infantum (IC50 = 0.15 ± 0.05 µM). Incubation with BA5 reduced the percentage of Leishmania amazonensis-infected macrophages and the number of intracellular parasites (IC50 = 4.1 ± 0.7 µM). To understand the mechanism of action underlying BA5 antileishmanial activity (incubation at IC50/2, IC50 or 2xIC50 values of the drug), we investigated ultrastructural changes by scanning electron microscopy and evaluated cell cycle, membrane mitochondrial potential, and cell death against promastigote forms of Leishmania amazonensis by flow cytometry. Promastigotes incubated with BA5 presented membrane blebbing, flagella damage, increased size, and body deformation. Flow cytometry analysis showed that parasite death is mainly caused by apoptosis-like death, arrested cell cycle in G0/G1 phase and did not alter the membrane mitochondrial potential of Leishmania amazonensis. Surprisingly, the combination of BA5 and amphotericin B, an assay used to determine the degree of drug interaction, revealed synergistic effects (CI = 0.15 ± 0.09) on promastigotes forms of Leishmania amazonensis. In conclusion, BA5 compound is an effective and selective antileishmanial agent.