Paw Edema Formation Research Articles

Guianensin, a Simulium guianense salivary protein, has broad anti-hemostatic and anti-inflammatory properties.

Salivary glands from blood-feeding arthropods secrete several molecules that inhibit mammalian hemostasis and facilitate blood feeding and pathogen transmission. The salivary functions from Simulium guianense, the main vector of Onchocerciasis in South America, remain largely understudied. Here, we have characterized a salivary protease inhibitor (Guianensin) from the blackfly Simulium guianense. A combination of bioinformatic and biophysical analyses, recombinant protein production, in vitro and in vivo experiments were utilized to characterize the molecula mechanism of action of Guianensin. Kinetics of Guianensin interaction with proteases involved in vertebrate inflammation and coagulation were carried out by surface plasmon resonance and isothermal titration calorimetry. Plasma recalcification and coagulometry and tail bleeding assays were performed to understand the role of Guianensin in coagulation. Guianensin was identified in the sialotranscriptome of adult S. guianense flies and belongs to the Kunitz domain of protease inhibitors. It targets various serine proteases involved in hemostasis and inflammation. Binding to these enzymes is highly specific to the catalytic site and is not detectable for their zymogens, the catalytic site-blocked human coagulation factor Xa (FXa), or thrombin. Accordingly, Guianensin significantly increased both PT (Prothrombin time) and aPTT (Activated partial thromboplastin time) in human plasma and consequently increased blood clotting time ex vivo. Guianensin also inhibited prothrombinase activity on endothelial cells. We show that Guianensin acts as a potent anti-inflammatory molecule on FXa-induced paw edema formation in mice. The information generated by this work highlights the biological functionality of Guianensin as an antithrombotic and anti-inflammatory protein that may play significant roles in blood feeding and pathogen transmission.

Anti-Inflammatory, Antinociceptive, Antipyretic, and Gastroprotective Effects of Eurycoma longifolia Jack Ethanolic Extract.

Tongkat ali (Eurycoma longifolia Jack) (ELJ) is a plant in the Simaroubaceae family. Its roots are used in traditional Thai medicine to treat inflammation, pain, and fever; however, the antiulcer abilities of its ethanolic extract have not been studied. This study examined the anti-inflammatory, antinociceptive, antipyretic, and gastroprotective effects of ethanolic ELJ extract in animal models and found that ELJ effectively reduced EPP-induced ear edema in a dose-dependent manner and that a high dose of ELJ inhibited carrageenan-induced hind paw edema formation. In cotton-pellet-induced granuloma formation, a high dose of ELJ suppressed the increases in wet granuloma weight but not dry or transudative weight. In the formalin-induced nociception study, ELJ had a significant dose-dependent inhibitory impact. Additionally, the study found that yeast-induced hyperthermia could be significantly reduced by antipyretic action at the highest dose of ELJ. In all the gastric ulcer models induced by chemical substances or physical activity, ELJ extracts at 150, 300, and 600 mg/kg also effectively prevented gastric ulcer formation. In the pyloric ligation model, however, the effects of ELJ extract on gastric volume, gastric pH, and total acidity were statistically insignificant. These findings support the current widespread use of Eurycoma longifolia Jack in traditional medicine, suggest the plant's medicinal potential for development of phytomedicines with anti-inflammatory, antinociceptive, and antipyretic properties, and support its use in the treatment of gastric ulcers due to its gastroprotective properties.

Design, optimization and evaluation of dexamethasone-loaded microneedles for inflammatory disorders

Dexamethasone (Dex) is a popular and highly potent anti-inflammatory drug, frequently used to treat a wide range of inflammatory disorders. However, the existing oral and parenteral delivery modes have several limitations, including systemic adverse effects and reduced patient compliance. This study aimed to develop a biodegradable microneedle (MN)-based transdermal delivery system capable of sustained, safe and effective delivery of Dex. A Quality by Design (QbD) approach was applied to design the Dex-loaded MN arrays. The formulation variables were optimized using a central composite design (CCD) model, generated with the statistical software package Design- Expert®. The optimized MNs were sharp, with heights ranging between 800 and 900 µm, appropriate for transdermal delivery. The MN arrays did not exhibit any cytotoxic effects on the fibroblast and keratinocyte cells. Moreover, the ex vivo studies confirmed the enhanced efficacy of MN-mediated skin permeation of Dex compared to passive permeation of drug solution. Finally, the in vivo anti-inflammatory efficacy was investigated using the carrageenan-induced rat paw edema model. The efficacy of the MN arrays to inhibit paw edema formation was found to be comparable to that of intravenous Dex injection and significantly greater than topical solution. Cytokine analysis also revealed that application of MN arrays downregulated the expressions of pro-inflammatory cytokines and upregulated the expressions of anti-inflammatory cytokines. Overall, the findings suggest that MN array could be a safe, easy, effective and minimally invasive alternative to the existing means of Dex delivery and could potentially be used for the treatment of inflammatory disorders.

Antinociceptive effect and identification of berberine alkaloid in Berberis ruscifolia extracts

Ethnopharmacological relevanceAerial parts (leaves and stems) of Berberis ruscifolia Lam. are a usual preparation as an analgesic, anti-inflammatory, antimalarial, antibacterial, and digestive in folk medicine. However, there were no previous studies of its chemical composition and biological activity related to analgesic effects. The objective of the studyThe evaluation of the anti-nociception of the infusion (I), the decoction (D), and the ethanolic extract (EE) obtained from aerial parts of B. ruscifolia and its main chemical constituent in them, in mouse models. Material and methodsThe chemical constituent of B. ruscifolia extracts was evaluated and quantified by LC-MS and HPLC methodology. The inhibition of nociception in mice was analyzed by formalin and acetic acid-induced contortions tests. Also, when the formalin test was performed to evaluate the antinociceptive activity, the inhibition of edema formation and the antipyretic effect of each extract were simultaneously evaluated in the same experiment. For the oral administration in the in vivo assays, doses ranging from 10 to 1000 mg/kg and 10–30 mg/kg were used for extract and the chemical compound, respectively. ResultsThe presence of berberine (Berb) was identified in the three evaluated extracts where the EE showed the highest content of this compound getting a yield of 2%, while in the I and D, Berb is present at 0.2%. The three extracts promoted a reduction of the contortions induced by acetic acid, being observed in EE the highest activity with 63 ± 6% of significant inhibition of the nociceptive behavior at a dose of 300 mg/kg, while D significantly inhibited 32 ± 12% at the same dose and for I at a dose of 1000 mg/kg an inhibition of 44 ± 8% was observed. Likewise, in the formalin trial, I and EE reduced nociception at a dose of 1000 (31 ± 5%) and 300 (35 ± 3%) mg/kg, respectively in the neurogenic phase, while in the second phase of the experiment, all the extracts evaluated showed an antinociceptive effect, with significant inhibition of I of 54 ± 6% and D of 44 ± 5% at a dose of 1000 mg/kg and for EE showed a 63 ± 2% inhibition at a dose of 300 mg/kg being the one with the highest antinociceptive activity. These extracts showed no inhibition in temperature and formalin-injected paw edema formation when compared to the control. As for Berb, at a 30 mg/kg dose, it showed significant inhibition of 70 ± 5% in the acetic acid-induced contortion test. ConclusionAltogether, the present results evidenced the analgesic properties of B. ruscifolia, scientific information presented for the first time, and also provided important knowledge not reported so far about the chemical composition of its extracts, by identifying the presence of Berb in them. Finally, we were able to conclude that the analgesic effect demonstrated by this medicinal plant is partly due to the presence of Berb.

Potent Analgesic Action of 2-acetoxy-5-(2-4 (trifluoromethyl)-phenethylamino)-benzoic Acid (Flusalazine) in Experimental Mice.

Nonsteroidal anti-inflammatory drugs (NSAIDs) and cyclooxygenase (COX)-2 selective inhibitors are the most widely used drugs to treat pain. Conventional NSAIDs and COX-2 selective inhibitors, however, cause several side effects such as gastric damage, kidney damage, and cardiovascular problems. Our previous study showed that 2-acetoxy-5-(2-4-(trifluoromethyl)-phenethylamino)-benzoic acid ie, flusalazine (also known as ND-07), which exerts dual actions by serving both as an anti-inflammatory agent and a free radical scavenger, is an effective and safe treatment for severe inflammatory diseases in mice. The goal of the present study was to examine the potential analgesic action and safety of flusalazine in mice models of pain. Flusalazine showed a significant analgesic effect in an acetic acid-induced abdominal constriction model. Likewise, total paw licking was reduced significantly in neurogenic (early stage) and inflammatory (late stage) pain induced by formalin in flusalazine-treated mice. In the tail immersion test, flusalazine significantly increased tail withdrawal time at 2 h after its administration. Also, the formation of paw edema in the flusalazine-treated group was significantly inhibited in a carrageenan-induced inflammatory pain model. Gastric damage was not induced by flusalazine even up to 1000 mg/kg, while aspirin and indomethacin caused critical gastric bleeding. These findings suggest that flusalazine's safety profile and analgesic effects have high translational potential for the clinical treatment of patients experiencing pain.

Preparation and in vitro–in vivo evaluation of QbD based acemetacin loaded transdermal patch formulations for rheumatic diseases

This research aimed to develop patches for transdermal delivery of acemetacin, which can be used to treat rheumatic diseasesand to determine their potential use. Patches were successfully created by solvent casting method using hydroxypropyl methylcellulose, propylene glycol, polyethylene glycol 400, tween 80, and dimethyl sulfoxide. Prepared patches were found using the Design of Experiments (DoE) method within the Quality by Design (QbD) approach. F1-ACM with a thickness of 0.1 ± 0.0 cm, a weight of 43.33 ± 6.29 mg, pH of 4.99 ± 0.24, moisture content of 18.33 ± 2.98%, a tensile strength of 9.196 ± 0.441 Mpa, elongation at break of 28.722 ± 0.803% and drug content of 100% was chosen as ideal formulation. 89.7% of ACM from F1-ACM was released in 5 min. F1-ACM significantly (p < 0.05) increased the response latency to the thermal stimulus at 90th (3.071 ± 0.517) and 120th (3.87 ± 0.332) min in the hot plate test. In the tail-flick experiment, F1-ACM significantly (p < 0.05) increased the reaction delay against heat stimuli at 90th (3.016 ± 0.695), 120th (2.884 ± 0.851), and 180th (2.893 ± 0.932) min. F1-ACM patch significantly (p < 0.001) inhibited paw edema formation at 1, 2, 3, 4, and 5 h after induction of inflammation as compared to the control group. Therefore, this formulation can be employed more efficiently for rheumatic disease.

In vivo and in silico anti-inflammatory properties of the sesquiterpene valencene

Valencene (VLN) is a sesquiterpene found in juices and essential oils of citrus species such as Cyperus rotundus. Considering the evidence that this species has anti-inflammatory effects, the present study aims to evaluate the anti-inflammatory activity of VLN in vivo and in silico. Swiss mice (n = 6) were orally treated according to their treatment groups as follows: VLN (10, 100 or 300 mg/kg), negative control (0.9% saline), and positive controls (indomethacin 25 mg/kg or promethazine 6 mg/kg). The anti-inflammatory activity was evaluated in murine models of acute and chronic inflammation. The inhibition of acute inflammation was evaluated in models of paw edema induced by different inflammatory agents (carrageenan, dextran, histamine, and arachidonic acid (AA)) and carrageenan-induced pleurisy and peritonitis. The modulation of chronic inflammation was evaluated in a granuloma model induced cotton pellets implantation. The interaction with inflammatory targets was evaluated in silico using molecular docking analysis. The administration of VLN to challenged mice significantly inhibited paw edema formation with no significant difference between the administered doses. The compound also reduced albumin extravasation, leukocyte recruitment, and the production of myeloperoxidase (MPO), IL-1β, and TNF-α in both pleural and peritoneal lavages. According to the mathematical-statistical model observed in silico analysis, this compound has favorable energy to interact with the cyclooxygenase enzyme (COX-2) and the histamine 1 (H1) receptor. Finally, animals treated with the sesquiterpene showed a reduction in both granuloma weight and concentration of total proteins in a chronic inflammation model. Given these findings, it is concluded that NLV presents promising pharmacological activity in murine models of acute and chronic inflammation.

Design, synthesis and in vivo evaluation of 1,4-dioxo-2-butenyl aryl amine derivatives as a promising anti-inflammatory drug prototype

Inflammation is a natural response of the organism to an infection, trauma, or cellular stress. Pain is the first symptom of acute and chronic inflammation. The standard class of medication to treat inflammatory pain is the nonsteroidal anti-inflammatory drug (NSAID). These drugs are associated with severe side effects such as gastric ulcers, gastritis, or internal bleeding. One of NSAIDs, Dipyrone® (metamizole) is largely used in many European and South American countries despite its dubious effectivity and its withdrawal from the market of several countries. Here, aiming to identify a new anti-inflammatory drug prototype based on Dipyrone® structure, a set of 27 molecules were virtually screened, and 4 compounds containing the active metabolite 4-aminoantipyrine and 1,4-dioxo-2-butenyl fragment were selected for docking, synthesis, and biological evaluation. The selection was based on the number of H-bonds and π- π stacking interactions between the inhibitor and the amino acids within the binding site of the enzyme. Carrageenan-induced paw edema, acetic acid-induced writhing, and formalin assays were used to evaluate inflammation and pain response. The selected compounds 1–4 inhibited the involvement of biogenic amines in the formation of paw edema. Compounds 1–4 also reduced pain in the inflammatory response phase. It has to be noted that 4-AA may cause agranulocytosis, which should be borne in mind when developing drug candidates of similar structure. Our new drug prototypes based on 4-aminoantipyrine and 1,4-dioxo-2-butenyl moieties open a gate for developing a prototype of nonsteroidal anti-inflammatory drugs.

In vitro and in vivo anti-inflammatory and anticoagulant activities of Myrciaria plinioides D. Legrand ethanol leaf extract

Myrciaria plinioides D. Legrand (Myrtaceae) is a native plant of Southern Brazil, which have potential in the food industry due to its edible fruits. Many plants belonging to this genus have been used for a variety of illnesses, including inflammatory disorders due to antioxidant properties. However, therapeutic uses of M. plinioides have been poorly studied. The aim of study was to assess the anti-inflammatory and anticoagulant activities of the ethanol leaf extract of M. plinioides. In M. plinioides extract-treated RAW 264.7 cells, assessments of cell viability, TNF-α release and p38 MAPK pathway-dependent protein expression were detected. In addition, rat paw edema models were used to analyze the anti-inflammatory effect of the extract. Macrophages cell line treated with M. plinioides extract showed a slight decrease in cell viability. In LPS-stimulated macrophages treated with different concentrations of the extract for 24 h, TNF-α release was inhibited, while modulation of p38 signaling pathway and inhibition of NF-κB p65 protein expression were dose-dependent. In rats, the extract inhibited the formation of paw edema, while an inhibitory effect on trypsin-like enzymes derived from mast cells was seen. Furthermore, the extract presented anticoagulant activity via extrinsic pathway, being able to block specifically factor Xa and thrombin. The study suggests that extract possess potent anti-inflammatory and anticoagulant effects. M. plinioides present great biological potential as a source for the development of anti-inflammatory and anticoagulant drugs. Additional studies can be proposed to better elucidate the mechanism by which M. plinioides exerts its effects.

Anti-Inflammatory Potential of the Oleoresin from the Amazonian Tree Copaifera reticulata with an Unusual Chemical Composition in Rats.

Copaifera reticulata Ducke is a popularly known species known as copaíba that is widely spread throughout the Amazon region. The tree yields an oleoresin which is extensively used in local traditional medicine mainly as an anti-inflammatory and antinociceptive agent. The aim of the present study was to assess the anti-inflammatory potential of this oleoresin obtained from a national forest in the central Amazon which presented an unusual chemical composition. The chemical composition of volatile compounds of oleoresin was analyzed by gas chromatography-mass spectrometry. The acute toxicity assay was performed with a single dose of 2000 mg/kg. The anti-inflammatory potential was evaluated by carrageenan-induced paw edema and air pouch assays using four different C. reticulata oleoresin concentrations (10, 100, and 400 mg/kg). The exudate was evaluated for nitrite concentration through the colorimetric method and for TNF-α, IL-1β, and PGE2 by ELISA. C. reticulata oleoresin collected in the Amazonian summer contained six major sesquiterpene compounds (β-bisabolene, cis-eudesma-6,11-diene, trans-α-bergamotene, β-selinene, α-selinene, and β-elemene) and was nontoxic at a dose of 2000 mg/kg, showing low acute toxicity. Different from oleoresin obtained from other sites of the Brazilian Amazon, the major volatile compound found was β-Bisabolene with 25.15%. This β-Bisabolene-rich oleoresin reduced the formation of paw edema induced by carrageenan and reduced the global number of cells in the air pouch assay, as well as exudate volume and nitrite, TNF-α, IL-1β, and prostaglandin E2 levels (p < 0.05). C. reticulata oleoresin with a high β-Bisabolene concentration showed anti-inflammatory activity, reducing vascular permeability and consequently edema formation, and thus reducing cell migration and the production of inflammatory cytokine, confirming its traditional use by local Amazonian communities.

Central and peripheral effects of environmental enrichment in a mouse model of arthritis

This study analyzed whether environmental enrichment (EE) modulates the nociceptive and inflammatory responses in the mouse model of arthritis induced by Complete Freund’s Adjuvant (CFA). Ninety male mice (C57BL/6-JUnib, 4-weeks-old; 20–25 g) were distributed into EE and standard (SE) groups. For EE, mice were kept in bigger cages using an alternation of materials to chew (wood and paper), for nesting (cotton), to use as hiding places (plastic tunnels), and for voluntary exercise (wheel running). Arthritis was induced by an injection of CFA (50 μL) into the right hind paw or saline solution in the control group. Separate groups received the anti-inflammatory drug dexamethasone (0.5 mg/kg; every 48 h). Inflammatory and pain measurements were performed from 1 to 35 days after CFA administration. EE per se reduced the acute paw edema formation and arthritis scores. The serum levels of tumor necrosis factor (TNF) were undetectable in any experimental groups. EE diminished the immunopositivity for the microglia marker IBA1 in the pre-frontal cortex, with slight changes for hippocampal GFAP-positive activated astrocytes. Finally, EE induced a marked increment of brain-derived nerve factor (BDNF) expression in the hippocampus, an effect that was fully prevented by dexamethasone. These data bring novel evidence on the peripheral and central effects of EE in a mouse arthritis model.

In-vivo studies to manifest the potent antiinflammatory and analgesic effect of the topical formulation, Pankajakasthuri orthoherb cream/ thermagel upon single application in wistar albino rats

Pain and inflammation are the most common and prominent symptoms of many diseases. Inflammation is the most pivotal response of a damaged tissue that induces various hostile stimuli such as infections, toxic chemical agents, and physical damage resulting in redness, edema, heat, and pain. Diclofenac sodium is the most widely prescribed medicine in the management of pain and inflammation. Lamentably, long-term use of this drug causes multiple harmful side effects. To overcome this, we have formulated Pankajakasthuri orthoherb cream/thermagel, a polyherbal topical preparation for inflammation and pain relief. The objective of the present study was to assess the anti-inflammatory and analgesic potential of Pankajakasthuri orthoherb cream/thermagel, when administered topically, using carrageenan-induced edema and formalin testing in experimental animals. Here, diclofenac sodium gel was used as a positive control for anti-inflammatory and analgesic activity. From the results of our study, it was clear that Pankajakasthuri orthoherb cream/ thermagel significantly inhibited the formation of the rat paw edema and this effect was significantly better than standard diclofenac sodium gel. Pankajakasthuri orthoherb cream/ thermagel recorded a positive result after 3 h of treatment and maximum reduction in edema was recorded after 18 h of treatment (96.72%). The analgesic effect of Pankajakasthuri orthoherb cream/thermagel was recorded mainly in the late phase (70.70%). This suggests that the activity may be due to peripheral action, when compared with the action of diclofenac sodium gel. Hence it can be concluded that Pankajakasthuri orthoherb cream/ thermagel can result in better anti-inflammatory and analgesic action for a longer duration upon single application compared with standard diclofenac sodium gel.

In vivo anti-inflammatory action evaluation of protease inhibitors from Cecropia hololeuca and their protein profile by electrophoretic and spectroscopic techniques

This work describes the electrophoretic and spectroscopic assessment of bioactive proteins from Cecropia hololeuca through SDS-PAGE, CE-UV, FT-IR and NMR, and their anti-inflammatory potential. In vitro assays were performed on murine macrophages J774A.1 and RAW 264.7 cells, and the in vivo anti-inflammatory potential on acute carrageenan edema in mice. The protein extract significantly inhibited the levels of NO, TNF-α, IL-1β and IL-6, and the expression of NF-κB in J774A.1 and RAW 264.7 cells, without causing toxicity to cells. In in vivo and ex vivo assay, the protein extract also reduced the formation of paw edema in animals, and the expression of IL-12 in macerate in the soft tissue of the paw. In the analysis of genotoxicity, the protein extract decreased the frequency of micronucleated erythrocytes when compared to the group treated with Dexamethasone. The analysis by SDS-PAGE, CE-UV, FT-IR and NMR corroborated the presence of proteins in the analyzed samples.

Lantana canescens (Kunth) inhibits inflammatory and hyperalgesic responses in murine models

Ethnopharmacological relevanceLantana canescens is popularly known in Brazil as “cidreirinha” or “chumbinho-branco”. It is found in Pantanal biome and its flowers and leaves are used in traditional medicine to treat pain and inflammation. Information about this species is limited to the activity of isolated essential oils. Studies with different extracts, composition, and biological properties are still scarce. Aim of this studyThe objective of this study was to evaluate the anti-inflammatory and anti-hyperalgesic activity of the hydroethanolic extract of L. canescens aerial parts. Materials and methodsThe hydroethanolic extract L. canescens aerial parts (HELc) was analyzed using HPLC-DAD-EM. Male and female Swiss mice weighing 18–25 g were used in the in vivo assays. Acute toxicity was assessed (2000 mg/kg); anti-inflammatory activity through paw edema, mast cell degranulation and peritonitis, and anti-hyperalgesic activity through abdominal writhing assays induced by acetic acid and formalin sensitization, were evaluated using the doses of 3, 30 and 300 mg/kg. ResultsThe phytochemical characterization of HELc confirmed the presence of glycosylated iridoids (theveside, theviridoside), verbascosides and flavonoids. The HELc did not present toxicity in the evaluated dose. HELc reduced formation of paw edema, degranulation of peritoneal mast cells and infiltration of polymorphonuclear cells into the animals peritoneal cavity. In addition, HELc decreased the number of abdominal writhing induced by acetic acid and the time of paw licking in the evaluation of formalin sensitization. ConclusionsThese results confirm the anti-inflammatory and anti-hyperalgesic effects of hydroethanolic extract of L. canescens, validating the use of this plant in folk medicine.

Anti-Inflammatory Potential of Aqueous Extract of Elsoltzia stachyodes on Experimental Models of Inflammation in Rats

Aim: Elsholtzia stachyodes is being used as a folk medicine to suppress the pain in various inflammation-related disorders in the North-eastern part of India. However, there is a lacking of evidence-based data on it. Hence, this study was conducted to evaluate the anti-inflammatory properties of an aqueous extract of Elsoltzia stachyodes (AEES) on experimental models of inflammation in rats. Materials and Methods: The qualitative phytochemical analysis tests were performed to check the presence of alkaloids, glycoside, flavonoids, saponin, tannin, and triterpenoids. Wistar (150–175 g) rats were used to evaluate anti-inflammatory property by carrageenan induced paw edema and cotton pellet-induced granulation tissue formation models. Indomethacin (10 mg/kg) and AEES were administered at doses of 50, 100, and 200 mg/kg in both the models. In case of carrageenan-induced paw edema model, the change in paw volume was estimated using plethysmometer. However, wounds weights of cotton pellet were measured in cotton pellet-induced granulation tissue formation model. Results were analyzed by one-way ANOVA. Results: The phytochemical analysis of AEES showed the presence of flavonoids, saponin, tannin, and triterpenoids. AEES showed a significant decrease in paw edema and weight of the cotton pellets in a dose-dependent manner in both the inflammatory models. Conclusion: The findings of the present study suggest that AEES exhibited significant anti-inflammatory activity and may have the potential of usefulness as an adjuvant therapy in inflammatory-related disorders.

Anti-inflammatory effects of FS48, the first potassium channel inhibitor from the salivary glands of the flea Xenopsylla cheopis

The voltage-gated potassium (Kv) 1.3 channel plays a crucial role in the immune responsiveness of T-lymphocytes and macrophages, presenting a potential target for treatment of immune- and inflammation related-diseases. FS48, a protein from the rodent flea Xenopsylla cheopis, shares the three disulfide bond feature of scorpion toxins. However, its three-dimensional structure and biological function are still unclear. In the present study, the structure of FS48 was evaluated by circular dichroism and homology modeling. We also described its in vitro ion channel activity using patch clamp recording and investigated its anti-inflammatory activity in LPS-induced Raw 264.7 macrophage cells and carrageenan-induced paw edema in mice. FS48 was found to adopt a common αββ structure and contain an atypical dyad motif. It dose-dependently exhibited the Kv1.3 channel in Raw 264.7 and HEK 293T cells, and its ability to block the channel pore was demonstrated by the kinetics of activation and competition binding with tetraethylammonium. FS48 also downregulated the secretion of proinflammatory molecules NO, IL-1β, TNF-α, and IL-6 by Raw 264.7 cells in a manner dependent on Kv1.3 channel blockage and the subsequent inactivation of the MAPK/NF-κB pathways. Finally, we observed that FS48 inhibited the paw edema formation, tissue myeloperoxidase activity, and inflammatory cell infiltrations in carrageenan-treated mice. We therefore conclude that FS48 identified from the flea saliva is a novel potassium channel inhibitor displaying anti-inflammatory activity. This discovery will promote understanding of the bloodsucking mechanism of the flea and provide a new template molecule for the design of Kv1.3 channel blockers.

Effects of photobiomodulation therapy on the local experimental envenoming by Bothrops leucurus snake

Bothrops leucurus is the major causative agent of snakebites in Brazil's Northeast. The systemic effects of its venom are effectively neutralized by antivenom therapy, preventing bitten patients' death. However, antivenom fails in neutralizing local effects that include intense pain, edema, bleeding, and myonecrosis. Such effects can lead to irreversible sequels, representing a clinically relevant issue for which there is no current effective treatment. Herein, the effects of photobiomodulation therapy (PBMT) were tested in the local actions induced by B. leucurus venom (BLV) in mice (n = 123 animals in 20 experimental groups). A continuous emission AlGaAs semiconductor diode laser was used in two wavelengths (660 or 780 nm). Mechanical nociceptive thresholds were assessed with the electronic von Frey apparatus. Local edema was determined by measuring the increase in paw thickness. Hemorrhage was quantified by digital measurement of the bleeding area. Myotoxicity was evaluated by serum creatine kinase (CK) activity and histopathological analysis. PBMT promoted anti-hypernociception in BLV-injected mice; irradiation with the 660 nm laser resulted in faster effect onset than the 780 nm laser. Both laser protocols reduced paw edema formation, whether irradiation was performed immediately or half an hour after venom injection. BLV-induced hemorrhage was not altered by PBMT. Laser irradiation delayed, but did not prevent myotoxicity caused by BLV, as shown by a late increase in CK activity and histopathological alterations. PBMT was effective in the control of some of the major local effects of BLV refractory to antivenom. It is a potential complementary therapy that could be used in bothropic envenoming, minimizing the morbidity of these snakebite accidents.

Anti-arthritic activity of D-carvone against complete Freund’s adjuvant-induced arthritis in rats through modulation of inflammatory cytokines

Chronic joint pain due to loss of cartilage function, degradation of subchondral bone, and related conditions are common plights of an arthritis patient. Antioxidant compounds could solve the problems in arthritic condition. The objective of this study was to evaluate the anti-arthritic activity of D-carvone against complete Freund’s adjuvant (CFA)-induced arthritis in rats. D-carvone was orally administered for 25 days at the doses of 30 and 60 mg/kg against CFA-induced arthritic rats. Changes in body weight, paw swelling, organ index, hematological parameters, oxidative stress markers, inflammatory cytokines, and histopathology were recorded. Oral treatment of D-carvone significantly improved the body weight, reduced the paw swelling, edema formation, and organ index in arthritic rats. The levels of white blood cells were reduced, red blood cells and hemoglobin levels were improved in D-carvone treated arthritic rats. Lipid peroxidation levels were lowered whereas enzymatic and non-enzymatic antioxidants were significantly elevated by D-carvone administration against arthritic rats. D-carvone significantly modulated inflammatory cytokine levels and improved the ankle joint pathology against CFA-induced arthritic inflammation. In conclusion, D-carvone proved significant anti-arthritic activity against CFA-induced arthritis in rats.

Machaerium acutifolium lectin inhibits inflammatory responses through cytokine modulation

Inflammatory response occurs when tissues are injured by pathogens, trauma, toxins, or heat. Lectins are proteins that recognize and bind reversibly to glycans and glycoconjugates and can modulate inflammatory responses in in vitro and in vivo models. As such, this study aimed to evaluate the potential of an anti-inflammatory lectin isolated from Machaerium acutifolium seeds (MaL) in mice and LPS-stimulated macrophage models. The protein was solubilized in sterile saline (0.9 % NaCl) immediately before treatment of mice by intraperitoneal routes at doses of 0.02 mg/kg, 1 mg/kg and 5 mg/kg. MaL significantly decreased inflammation in the formalin test, inhibited cell migration in experimental models of carrageenan-induced peritonitis, and blocked the formation of paw edema induced by carrageenan and dextran. In vitro studies showed that MaL downregulated the proinflammatory cytokine genes inducible nitric oxide synthase (iNOS) and tumor necrosis factor-α (TNF-α), but upregulated the anti-inflammatory IL-10 gene in LPS-stimulated macrophages. Therefore, this study suggests that MaL has an anti-inflammatory effect relative to modulated levels of pro- and anti-inflammatory cytokines, indicating that MaL can be used as a potential therapeutic agent in cellular inflammatory events.

Identification and quantification of oleanane triterpenoid saponins and potential analgesic and anti-inflammatory activities from the roots and rhizomes of Panax stipuleanatus

BackgroundPanax stipuleanatus represents a folk medicine for treatment of inflammation. However, lack of experimental data does not confirm its function. This article aims to investigate the analgesic and anti-inflammatory activities of triterpenoid saponins isolated from P. stipuleanatus. MethodsThe chemical characterization of P. stipuleanatus allowed the identification and quantitation of two major compounds. Analgesic effects of triterpenoid saponins were evaluated in two models of thermal- and chemical-stimulated acute pain. Anti-inflammatory effects of triterpenoid saponins were also evaluated using four models of acetic acid–induced vascular permeability, xylene-induced ear edema, carrageenan-induced paw edema, and cotton pellet–induced granuloma in mice. ResultsTwo triterpenoid saponins of stipuleanosides R1 (SP-R1) and R2 (SP-R2) were isolated and identified from P. stipuleanatus. The results showed that SP-R1 and SP-R2 significantly increased the latency time to thermal pain in the hot plate test and reduced the writhing response in the acetic acid–induced writhing test. SP-R1 and SP-R2 caused a significant decrease in vascular permeability, ear edema, paw edema, and granuloma formation in inflammatory models. Further studies showed that the levels of inflammatory mediators, nitric oxide, malondialdehyde, tumor necrosis factor-α, and interleukin 6 in paw tissues were downregulated by SP-R1 and SP-R2. In addition, the rational harvest of three- to five-year-old P. stipuleanatus was preferable to obtain a higher level of triterpenoid saponins. SP-R2 showed the highest content in P. stipuleanatus, which had potential as a chemical marker for quality control of P. stipuleanatus. ConclusionThis study provides important basic information about utilization of P. stipuleanatus resources for production of active triterpenoid saponins.