Hind Paw Research Articles

Effect of the High-Intensity Interval Training on BDNF Level in Ischemic Stroke Rat Model on the Recovery of Motor Function

BACKGROUND: Stroke is one of the major causes of disability in the world. High-intensity interval training (HIIT) is known as a novel treatment to promote stroke recovery. However, the results differ in their effects on irisin, which is a regulator of brain-delivered neurotrophic factor (BDNF). Therefore, this study was conducted to investigate the effect of HIIT on BDNF and irisin levels in a rat model of ischemic stroke with middle cerebral artery occlusion (MCAO) induction on recovery motor function.METHODS: Rats were categorized into 4 groups: sham, MCAO, MCAO+moderate-intensity interval training (MIIT), and MCAO+HIIT. MCAO induction was performed to create the ischemic stroke rats model. The motor function was assessed through rotarod and footprint tests. Blood samples were obtained 6 days before MCAO and 14 days after MCAO to examine BDNF and irisin levels with enzyme-linked immunosorbent assay (ELISA). Brain tissue samples were collected 14 days after MCAO for histopathological examination of cortical tissue with hematoxylin-eosin (HE) staining.RESULTS: Rats in the MCAO+HIIT group exhibited an enhanced ability to walk on the rotarod (p=0.016). The stride-length hind paw right in the MCAO+HIIT group demonstrated a noteworthy increase in comparison to baseline value (p=0.036), and the stride-length fore paw right showed a significant increase in both the MCAO+HIIT (p=0.036) and MCAO groups (p=0.034). BDNF significantly improved in the MCAO+MIIT (p=0.043) and MCAO+HIIT groups (p=0.018). The irisin level only showed a significant enhancement in the MCAO+HIIT group (p=0.018).CONCLUSION: HIIT increased motor function, while BDNF level increased with HIIT and MIIT intervention. This preclinical research is useful for supporting the recovery of stroke patients by HIIT intervention.KEYWORDS: BDNF, HIIT, MIIT, irisin, ischemic stroke, MCAO

Stimulator of Interferon Genes Pathway Activation through the Controlled Release of STINGel Mediates Analgesia and Anti-Cancer Effects in Oral Squamous Cell Carcinoma.

Oral squamous cell carcinoma (OSCC) presents significant treatment challenges due to its poor survival and intense pain at the primary cancer site. Cancer pain is debilitating, contributes to diminished quality of life, and causes opioid tolerance. The stimulator of interferon genes (STING) agonism has been investigated as an anti-cancer strategy. We have developed STINGel, an extended-release formulation that prolongs the availability of STING agonists, which has demonstrated an enhanced anti-tumor effect in OSCC compared to STING agonist injection. This study investigates the impact of intra-tumoral STINGel on OSCC-induced pain using two separate OSCC models and nociceptive behavioral assays. Intra-tumoral STINGel significantly reduced mechanical allodynia in the orofacial cancer model and alleviated thermal and mechanical hyperalgesia in the hind paw model. To determine the cellular signaling cascade contributing to the antinociceptive effect, we performed an in-depth analysis of immune cell populations via single-cell RNA-seq. We demonstrated an increase in M1-like macrophages and N1-like neutrophils after STINGel treatment. The identified regulatory pathways controlled immune response activation, myeloid cell differentiation, and cytoplasmic translation. Functional pathway analysis demonstrated the suppression of translation at neuron synapses and the negative regulation of neuron projection development in M2-like macrophages after STINGel treatment. Importantly, STINGel treatment upregulated TGF-β pathway signaling between various cell populations and peripheral nervous system (PNS) macrophages and enhanced TGF-β signaling within the PNS itself. Overall, this study sheds light on the mechanisms underlying STINGel-mediated antinociception and anti-tumorigenic impact.

Interleukin-1α links peripheral CaV2.2 channel activation to rapid adaptive increases in heat sensitivity in skin

Neurons have the unique capacity to adapt output in response to changes in their environment. Within seconds, sensory nerve endings can become hypersensitive to stimuli in response to potentially damaging events. The underlying behavioral response is well studied, but several of the key signaling molecules that mediate sensory hypersensitivity remain unknown. We previously discovered that peripheral voltage-gated CaV2.2 channels in nerve endings in skin are essential for the rapid, transient increase in sensitivity to heat, but not to mechanical stimuli, that accompanies intradermal capsaicin. Here we report that the cytokine interleukin-1α (IL-1α), an alarmin, is necessary and sufficient to trigger rapid heat and mechanical hypersensitivity in skin. Of 20 cytokines screened, only IL-1α was consistently detected in hind paw interstitial fluid in response to intradermal capsaicin and, similar to behavioral sensitivity to heat, IL-1α levels were also dependent on peripheral CaV2.2 channel activity. Neutralizing IL-1α in skin significantly reduced capsaicin-induced changes in hind paw sensitivity to radiant heat and mechanical stimulation. Intradermal IL-1α enhances behavioral responses to stimuli and, in culture, IL-1α enhances the responsiveness of Trpv1-expressing sensory neurons. Together, our data suggest that IL-1α is the key cytokine that underlies rapid and reversible neuroinflammatory responses in skin.

Polymeric Nanocapsules Loaded with Lidocaine: A Promising Formulation for Topical Dental Anesthesia.

Lidocaine is the most commonly used local anesthetic worldwide, known for its rapid onset and moderate duration of anesthesia. However, it is short-lived and does not effectively promote effective topical anesthesia in the oral cavity when used alone. Our aim was to investigate whether an approximate 50% encapsulation of lidocaine in poly(ε-caprolactone) nanocapsules (LDC-Nano) would be able to increase its permeation and analgesic efficacy and reduce cytotoxicity. In this study, we characterized LDC-Nano and conducted MTT tests with HaCaT cells to assess their in vitro cytotoxicity. Additionally, in vitro permeation assays across the pig esophageal epithelium and the anesthetic efficacy of the hind paw incision model in rats were performed. Plain lidocaine (LDC) was compared with LDC-Nano and lidocaine hydrochloride plus epinephrine (LDC-Epi). The physicochemical characteristics of LDC-Nano were satisfactory (pH: 8.1 ± 0.21; polydispersity index: 0.08 ± 0.01; mean diameter (nm): 557.8 ± 22.7; and encapsulation efficiency (%): 51.8 ± 1.87) and remained stable for up to 4 months. LDC-Nano presented similar in vitro cytotoxicity to LDC but was higher than LDC-Epi (LD50: LDC = 0.48%; LDC-Nano = 0.47%; and LDC-Epi = 0.58%; p < 0.0001). Encapsulation increased the permeability coefficient about 6.6 times and about 7.5 the steady-state flux of lidocaine across the mucosal epithelium. Both encapsulation and epinephrine improved anesthesia duration, with epinephrine demonstrating superior efficacy (100% of animals were anesthetized up to 100, 30, and 20 min when LDC-Epi, LDC-nano, and LDC were used, respectively). Although LDC-Epi demonstrated superior in vivo anesthetic efficacy, the in vitro permeation and cytotoxicity of LDC-Nano indicate promising avenues for future research, particularly in exploring its potential application as a topical anesthetic in the oral cavity.

ETHNOPHARMACOLOGICAL INVESTIGATION OF PLEUROTUS OSTREATUS FOR ANTI-OXIDATIVE AND ANTI-INFLAMMATORY ACTIVITY IN EXPERIMENTAL ANIMALS

Objective: This particular study was aimed to evaluate the anti-oxidant effect of Pleurotus ostreatus using the in vitro method and also against inflammation. Methods: A methanolic extract of P. ostreatus (MEPO) was prepared by the cold maceration technique. Different tests. In vitro anti-oxidant activity was investigated using the 1,1-diphenyl-1-picryl hydrazyl (DPPH) method. Wistar rats were chosen for the study; animals weighing 150–200 g were divided into six groups of six each (n=6). Control animals were grouped as I, group II was administered with approximately 100 μL of 1% suspension of carrageenan in saline and injected into the plantar surface of the right hind paw, group III, IV, V, and VI were given with carrageenan, followed by diclofenac sodium (150 mg/kg body weight, p.o.) and MEPO at the dose of 200, 400, and 800 mg/kg orally for 21 days. The paw volume and percentage of inhibition of the paw were measured in all animals. Results: The IC50 values of the test extract in the DPPH free radical scavenging assay were found to be 44.02±0.09 as compared with the standard drug 35.01±0.12, which was considered significant (***p&lt;0.001). The test extract at the dose of 400 and 800 mg/kg significantly decreased (**p&lt;0.001) the paw volume thereby, the inflammation, the % percentage of inhibition in paw volume was compared to the positive control in the carrageenan-induced paw edema. Conclusion: The study explored a potential source for anti-oxidants in the MEPO, which also seems to be effective in chemical-induced inflammation.

Potensi Ekstrak Etanol Kulit Batang Jabon (Anthocephalus cadamba (Roxb) Miq.) Sebagai Antiinflamasi

This study investigates the anti-inflammatory properties of an ethanol extract from Jabon plant stem bark in male mice using the Rat Hind Paw Edema method. Twenty-five male mice were divided into five groups: negative control (Na-CMC 1%), positive control (mefenamic acid), and three groups receiving the extract at doses of 50 mg/kg, 100 mg/kg, and 200 mg/kg. Measurements were taken every 30 minutes for three hours using a plethysmometer. Analysis with one-way ANOVA and Duncan's test revealed significant anti-inflammatory effects for all doses, with the most effective being 100 mg/kg, showing 93.42% inhibition. The results suggest Jabon plant extract as a promising anti-inflammatory agent.

Neutrophil extracellular traps as a unique target in the treatment of inflammatory pain

Pain is a widespread motivation for seeking healthcare and stands as a substantial global public health concern. Despite comprehensive investigations into the mechanisms of pain sensitization induced by inflammation, efficacious treatments options remain scarce. Neutrophil extracellular traps (NETs) have been associated with the progression and tissue damage of diverse inflammatory diseases. This study aims to explore the impact of NETs on the progression of inflammatory pain and explore potential therapeutic approaches. Initially, we observed neutrophil infiltration and the formation of NETs in the left hind paw of mice with inflammatory pain induced by complete Freund's adjuvant (CFA). Furthermore, we employed the peptidyl arginine deiminase 4 (PAD4) inhibitor Cl-amidine (diluted at 50 mg/kg in saline, administered via tail vein injection once daily for three days) to impede NETs formation and administered DNase1 (diluted at 10 mg/kg in saline, once daily for three days) to break down NETs. We investigated the pathological importance of peripheral NETs formation in inflammatory pain and its influence on the activation of spinal dorsal horn microglia. The findings indicate that neutrophils infiltrating locally generate NETs, leading to an increased release of inflammatory mediators that worsen peripheral inflammatory reactions. Consequently, this results in the transmission of more harmful peripheral stimuli to the spinal cord, triggering microglial activation and NF-κB phosphorylation, thereby escalating neuroinflammation and fostering pain sensitization. Suppression of peripheral NETs can mitigate peripheral inflammation in mice with inflammatory pain, reverse mechanical and thermal hypersensitivity by suppressing microglial activation in the spinal cord, ultimately diminishing inflammatory pain. In conclusion, these discoveries propose that obstructing or intervening with NETs introduces a novel therapeutic avenue for addressing inflammatory pain.

Sodium Cromolyn Modulates Carrageenan-Induced Acute Inflammation Through the Interleukin-6 Levels

Background: Previous studies have indicated that sodium cromolyn does not negatively affect steroidal and nonsteroidal anti-inflammatory drugs (NSAIDs). Objective: This study aimed to investigate the anti-inflammatory effects of sodium cromolyn on carrageenan-induced paw edema in rats and assess its impact on IL-6, an anti-inflammatory cytokine. Methods: The carrageenan-induced paw edema model is widely used for studying inflammatory mechanisms and evaluating anti-inflammatory drugs. The test groups received intraperitoneal injections of sodium cromolyn at doses of 25, 50, and 100 mg/kg alongside a positive control group (indomethacin at 5 mg/kg) and a negative control group (which received the solvent, saline). Thirty minutes after drug administration, carrageenan (1% w/v) was injected into the right hind paws of the rats. Changes in paw volume were measured using a plethysmometer at 0.5, 1, 2, and 3 hours post-carrageenan injection. Blood and paw tissue samples were collected three hours after carrageenan administration, and IL-6 levels were determined using the standard rat Interleukin-6 ELISA kit. Results: The plasma and tissue IL-6 levels in the group treated with cromolyn (25 mg/kg) were significantly higher than those in the positive control group (indomethacin 5 mg/kg). Sodium cromolyn at a dose of 25 mg/kg had a negligible effect on reducing paw edema and IL-6 levels in serum and paw tissues compared to indomethacin (5 mg/kg). The IL-6 levels in plasma and tissues for the group receiving 50 mg/kg of cromolyn were not significantly different from those in the indomethacin group (5 mg/kg). However, IL-6 levels in the group treated with 100 mg/kg of cromolyn were significantly lower than those in the indomethacin group. Among the groups treated with sodium cromolyn, the highest and lowest IL-6 levels were observed in the groups receiving 25 mg/kg and 100 mg/kg of sodium cromolyn, respectively. The anti-inflammatory effects of sodium cromolyn at doses of 25 mg/kg, 50 mg/kg, and 100 mg/kg were analyzed, revealing that 25 mg/kg of cromolyn sodium was not significantly effective at any time point (P &lt; 0.05). Additionally, no significant differences (P &lt; 0.05) were observed between the 50 mg/kg and 100 mg/kg doses at any study hour. Sodium cromolyn at a 50 mg/kg dose was as effective as indomethacin (5 mg/kg) in significantly reducing paw edema. Conclusions: Sodium cromolyn, at doses of 50 and 100 mg/kg, effectively reduced paw edema and the concentration of serum and paw tissue IL-6 (P &lt; 0.05), demonstrating significant anti-inflammatory activity. Sodium cromolyn at a 50 mg/kg dose is recommended as the anti-inflammatory dose of this drug.

Bio fabrication of Fe2O3 NPs and their analgesia efficiency for post-operative epidural anaesthetic applications- study of chronic inflammatory pain model in mice

ABSTRACT The current study reported the synthesis of ferric oxide nanoparticles (Fe2O3 NPs) using the leaf extract of Amaranthus spinosus Linn and studied their analgesia efficiency in chronic inflammatory pain model in mice. X-RD, TEM results showed that the spherical Fe2O3 NPs having rhombohedral crystalline structure with an average particle size of 10 nm. Additionally, inflammatory pain models were designed through FCA injection on the CD1 mice hind paw with various Fe2O3 NPs doses and analgesic behaviour was monitored. The prepared Fe2O3 NPs showed a dose-dependent analgesic activity with considerable decrease in inflammatory cells, ROS production and pro-inflammatory markers. Additionally, the expression of enzymes responsible for ROS production were decreased. The outcomes of present work revealed the local administration of Fe2O3 NPs caused the substantial analgesia through reduction of pro-inflammatory signalling and inflammatory cellular infiltration along with micro-environmental free radical scavenging in mouse inflammatory pain model.

PEDF peptide plus hyaluronic acid stimulates cartilage regeneration in osteoarthritis via STAT3-mediated chondrogenesis.

Pigment epithelium-derived factor (PEDF) is known to induce several types of tissue regeneration by activating tissue-specific stem cells. Here, we investigated the therapeutic potential of PEDF 29-mer peptide in the damaged articular cartilage (AC) in rat osteoarthritis (OA). Mesenchymal stem/stromal cells (MSCs) were isolated from rat bone marrow (BM) and used to evaluate the impact of 29-mer on chondrogenic differentiation of BM-MSCs in culture. Knee OA was induced in rats by a single intra-articular injection of monosodium iodoacetate (MIA) in the right knees (set to day 0). The 29-mer dissolved in 5% hyaluronic acid (HA) was intra-articularly injected into right knees at day 8 and 12 after MIA injection. Subsequently, the therapeutic effect of the 29-mer/HA on OA was evaluated by the Osteoarthritis Research Society International (OARSI) histopathological scoring system and changes in hind paw weight distribution, respectively. The regeneration of chondrocytes in damaged AC was detected by dual-immunostaining of 5-bromo-2'-deoxyuridine (BrdU) and chondrogenic markers. The 29-mer promoted expansion and chondrogenic differentiation of BM-MSCs cultured in different defined media. MIA injection caused chondrocyte death throughout the AC, with cartilage degeneration thereafter. The 29-mer/HA treatment induced extensive chondrocyte regeneration in the damaged AC and suppressed MIA-induced synovitis, accompanied by the recovery of cartilage matrix. Pharmacological inhibitors of PEDF receptor (PEDFR) and signal transducer and activator of transcription 3 (STAT3) signalling substantially blocked the chondrogenic promoting activity of 29-mer on the cultured BM-MSCs and injured AC. The 29-mer/HA formulation effectively induces chondrocyte regeneration and formation of cartilage matrix in the damaged AC.

125 Contribution of Sensory Neuron-associated Macrophages to Neuropathic Pain via Up-regulation of Calcium Channel in C Fiber Sensory Neurons in Dorsal Root Ganglion: An Animal Model Study

INTRODUCTION: Animal models have demonstrated that sensory neuron-associated macrophages (sNAMs) play a key role in the initiation and persistence of neuropathic pain. The Cav3.2 calcium channel (Cav3.2) is also known to be involved in modulating neuropathic pain in peripheral sensory neurons. METHODS: Male Sprague-Dawley rats underwent unilateral tight ligation of the lumbar 5 spinal nerve and were divided into three groups: a control group (SHAM), a group that underwent ligation without, and with minocycline (SNL and SNL+Mino). The mechanical pain threshold of the hind paw was assessed with von-Frey filament application. Immunohistochemical staining techniques were used to evaluate the levels of sNAM, Cav3.2, and C-fiber sensory neurons with anti-Iba1, anti-Cav3.2, and IB4 antibodies, respectively. Cytokine levels were analyzed using cytokine arrays and western blotting. RESULTS: The SNL group had a significantly lower pain threshold compared to the control group (p &lt; 0.001). In the DRG, neuropathic pain led to an accumulation of macrophages and increased their ability to attach to Cav 3.2. The expression of Cav 3.2 then triggered the activation of IB4 cells in the DRG. Additionally, cytokine array and western blot analyses revealed a higher expression rate of TNF-α in the SNL group compared to the control group (p = 0.048). Treatment with minocycline resulted in increased pain threshold, decreased number of macrophages in the DRG, and interruption of the attachment between macrophages and Cav 3.2, leading to lower expression of TNF-α. CONCLUSIONS: In conclusion, the results of this study suggest that sNAMs may play a role in the development of neuropathic pain by increasing the expression of Cav3.2 through the upregulation of TNF-α release in peripheral sensory nociceptive neurons.

The nociceptive activity of peripheral sensory neurons is modulated by the neuronal membrane proteasome

Proteasomes are critical for peripheral nervous system (PNS) function. Here, we investigate mammalian PNS proteasomes and reveal the presence of the neuronal membrane proteasome (NMP). We show that specific inhibition of the NMP on distal nerve fibers innervating the mouse hind paw leads to reduction in mechanical and pain sensitivity. Through investigating PNS NMPs, we demonstrate their presence on the somata and proximal and distal axons of a subset of dorsal root ganglion (DRG) neurons. Single-cell RNA sequencing experiments reveal that the NMP-expressing DRGs are primarily MrgprA3+ and Cysltr2+. NMP inhibition in DRG cultures leads to cell-autonomous and non-cell-autonomous changes in Ca2+ signaling induced by KCl depolarization, αβ-meATP, or the pruritogen histamine. Taken together, these data support a model whereby NMPs are expressed on a subset of somatosensory DRGs to modulate signaling between neurons of distinct sensory modalities and indicate the NMP as a potential target for controlling pain.

Antinociception induced by rosuvastatin in mice: Modulation by opioid receptor antagonists, risperidone and l-name

Statins are widely used in cardiovascular disease as cholesterol lowering drugs. However, they also have other actions (pleiotropic effects) including antiproliferative, antithrombotic, neuroprotective, immunomodulatory, anti-inflammatory and antinociceptive activity. The aim of this study was to determine the antinociception properties of rosuvastatin in two murine models of pain and the involvement of opioid antagonists (naltrexone, naltrindole, norbinaltorphimine), risperidone, and L-NAME (L-NG-nitro arginine methyl ester) in this effect. Rosuvastatin was chosen among available statins because it is commonly prescribed and has high potency, efficacy, and an acceptable safety profile. Rosuvastatin antinociception was evaluated in the acetic acid writhing test and the formalin hind paw test by dose-response curves, before and after the i.p. administration of opioid antagonists, risperidone, and L-NAME. This work demonstrates that the assayed drugs modulate the antinociceptive effect of rosuvastatin in both experimental murine pain tests. The antinociception effect described for rosuvastatin may be due to a particular modulation induced by the opioid antagonists, risperidone and L-NAME. Given the broad effects of rosuvastatin, the results of this study may have novel clinical implications in the therapy of pain.

Effect of anethole and anethole+itraconazole assiciantion on experimental arthritis induced by Paracoccidioides brasiliensis

In this study, the effect of treatment with anethole (AN) and the combination anethole+itraconazole (AN+IT) compared to IT, on infectious arthritis induced by Paracoccidiodes brasiliensis, was evaluated. The animals were treated for 14 days at doses of anethole (AN - 62.5, 125 and 250 mg/kg), itraconazole (IT - 12.5, 25 and 50 mg/kg) and the combination of anethole+itraconazole (AN+ IT - 62.5 and 12.5 mg/kg). The parameters evaluated were: the development of knee edema, the number of leukocytes recruited into the joint cavity, the body weight of the animals, the walking capacity, the plasmatic concentration of nitric oxide, the concentration of TNF in the knee joint exudate, the production of anti-Pb antibodies and the activity of plasma transaminases (AST and ALT). Histological changes in the right knee joint of the hind paw were evaluated using Hematoxylin-eosin and Grocott staining. The results showed that treatment with monotherapies and combinations reduced knee joint edema, the number of leukocytes recruited into the synovial cavity and improved the animals' gait. The concentration of plasma NO, tissue TNF and anti-Pb were reduced by treatment with IT at all doses tested and with the AN+IT combination. Treatment with AN only reduced the plasma concentration of NO and tissue TNF at a high dose. Altogether, the data showed that treatments with IT monotherapy and the combination of AN + IT showed a similar inhibitory effect on the development of arthritis.

Mode of delivery alters sensitivity to thermal and chemical stimuli in adult rats: An experimental study.

The mode of delivery might prompt a long-lasting alteration in physiological and behavioral responsiveness in offspring. This study was intended to evaluate if the mode of delivery could alter sensitivity to thermal and chemical stimuli in female rats. 56 adult female Wistar rats (200-220 gr) that were born by vaginal or cesarean section (C-section) were used (n = 28/each). Inflammatory pain was induced by subcutaneous injection of formalin into the hind paw. The thermal nociceptive threshold was determined by tail-flick and hot plate tests. Besides, the Western blot test was used to evaluate the spinal cord levels of c-Fos and c-Jun proteins. Formalin-induced inflammation was significantly decreased in C-section group as compared to vaginally born rats (p 0.001). The baseline nociceptive threshed and morphine-induced analgesia were significantly increased in C-section groups in comparison to vaginally born rats. In addition, the levels of c-Fos and c-Jun proteins were significantly decreased in the spinal cord of C-section rats as compared to vaginally born animals (p 0.01). Morphine treatment could decrease the expression of c-Fos and c-Jun in the C-section group (p 0.05). Overall, C-section rats showed lower spinal nociceptive processing and neuronal activity later in life, compared to the vaginal born rats.

Extracorporeal photopheresis reduces inflammation and joint damage in a rheumatoid arthritis murine model

BackgroundRheumatoid arthritis (RA) is an autoimmune disease characterized by inflammatory reactions and tissue damage in the joints. Long-term drug use in clinical practice is often accompanied by adverse reactions. Extracorporeal photopheresis (ECP) is an immunomodulatory therapy with few side effects, offering a potential and safe therapeutic alternative for RA through the induction of immune tolerance. This study aimed to investigate the therapeutic effects of ECP on RA using a collagen-induced arthritis (CIA) murine model, as well as to explore its immunomodulatory effects in vivo. Additionally, particular attention was given to the significant role of monocytes during the ECP process.MethodsA murine model of rheumatoid arthritis was established by administering two injections of bovine type II collagen to DBA/1J mice. ECP, ECP-MD (mononuclear cells were depleted during the ECP), MTX, and PBS treatment were applied to the CIA mice. During the treatment process, clinical scores and body weight changes of CIA mice were closely monitored. After six treatment sessions, micro-CT images of the hind paws from live mice were captured. Ankle joints and paws of the mice were collected and processed for histological evaluation. Spleen samples were collected to measure the Th17/Treg cells ratio, and serum samples were collected to assess cytokine and anti-type II collagen IgG levels. Monocytes and dendritic cells populations before and after ECP in vitro were detected by flow cytometry.ResultECP therapy significantly attenuated the progression of CIA, alleviated the severity of clinical symptoms in CIA mice and effectively suppressed synovial hyperplasia, inflammation, and cartilage damage. There was an expansion in the percentage of CD3 + CD4 + CD25 + FoxP3 + Tregs and a decrease in CD3 + CD4 + IL17A + Th17 cells in vivo. Furthermore, ECP reduced the serum levels of pro-inflammatory cytokines IL-6 (53.47 ± 7.074 pg/mL vs 5.142 ± 1.779 pg/mL, P < 0.05) and IL-17A (3.077 ± 0.401 pg/mL vs 0.238 ± 0.082 pg/mlL, P < 0.0001) compared with PBS. Interestingly, the depletion of monocytes during the ECP process did not lead to any improvement in clinical symptoms or histological scores in CIA mice. Moreover, the imbalance in the Th17/Treg cells ratio became even more pronounced, accompanied by an augmented secretion of pro-inflammatory cytokines IL-6 and IL-17A. In vitro, compared with cells without ECP treatment, the proportion of CD11b + cells were significantly reduced (P < 0.01), the proportion of CD11c + cells were significantly elevated (P < 0.001) 24 h after ECP treatment. Additionally, the expression of MHC II (P < 0.0001), CD80 (P < 0.01), and CD86 (P < 0.001) was downregulated in CD11c + cells 24 h after ECP treatment.ConclusionOur study demonstrates that ECP exhibits a therapeutic effect comparable to conventional therapy in CIA mice, and the protective mechanisms of ECP against RA involve Th17/Treg cells ratio, which result in decreased IL-6 and IL-17A. Notably, monocytes derived from CIA mice are an indispensable part to the efficacy of ECP treatment, and the proportion of monocytes decreased and the proportion of tolerogenic dendritic cells increased after ECP treatment in vitro.

In vivo Anti-Inflammatory and Antinociceptive Activity Evaluation of Brassica Rapa Ssp. Chinensis Ethanolic Extract with In Vitro Thrombolytic and Anthelmintic Activity Test

Objective: Brassica rapa ssp. chinensis is a well-known vegetable plant (Family: Brassicaceae) often grown as a type of Chinese cabbage. The present study was designed to explore the thrombolytic, anthelmintic, anti-inflammatory, and antinociceptive activities of ethanolic extract of the Brassica rapa ssp. chinensis leaves. Methods: The in vitro thrombolytic activity of the leaf extract was evaluated by clot analysis using Streptokinase as a standard drug. The in vitro anthelmintic activity was assessed by applying four different concentrations of the plant extract (10 mg/ml, 20 mg/ml, 40 mg/ml, and 60mg/ml) and recording the time of paralysis and death of worms. Two different doses (100mg/kg and 250mg/kg b.w.) were administered to evaluate the anti-inflammatory activity of the carrageenan-induced paw edema model. The antinociceptive activity was evaluated by hot plate test in mice by two different doses (200mg/kg and 400 mg/kg b.w.). Results: The ethanolic leave extract had thrombolytic action, which broke up blood clots that had already formed. The ethanolic leaf extract showed both paralysis and faster worm death at those concentrations in the anthelmintic test. The extract showed significant anti-inflammatory action in pedal edema compared to the control group. The results show that the highest level of anti-inflammatory action against carrageenan-induced hind paw edema may be achieved with an ethanolic extract of Brassica rapa ssp. chinensis (250 mg/kg, p.o.). The proportion of maximum possible effect in the hot plate test was 82.49 percent in the antinociceptive evaluation. Conclusion: The pharmacological test of leaf extract endorsed the clinical uses of Brassica rapa ssp. chinensis in treating thrombosis, intestinal worms, and inflammation. Thus, this plant could be subjected to extensive bioactive compounds to discover novel therapeutic agents. Keywords: Brassica rapa ssp. chinensis, ethanolic extract, thrombolytic activity, anthelmintic activity, anti-inflammatory activity, antinociceptive activity

Formulation and Evaluation of Herbal Emulsion-Based Gel Containing Combined Essential Oils from Melaleuca alternifolia and Citrus hystrix

Aims: This study aimed to investigate the synergistic anti-inflammatory effects of combined essential oils from Melaleuca alternifolia and Citrus hystrix peel. Methodology: An emulsion-based gel containing combined essential oils from Melaleuca alternifolia and Citrus hystrix peel was topically applied to the injected area of the carrageenan-induced hind paw edema mouse model. Diclofenac sodium was used as a positive control. The inhibition percentage of protein denaturation in all mice was assessed and calculated based on paw volume (Pv). Results: The highest inhibition percentage, recorded at the third hour in the carrageenan-induced hind paw edema mouse model, was 44.76%, achieved by the mice treated with combined essential oils. This was followed by diclofenac-treated (37.14%), tea tree essential oil-treated (27.62%), and kaffir lime peel essential oil-treated mice (20.10%). A similar trend was observed after 24 hours of treatment, with the anti-inflammatory activity of combined essential oils-treated mice recorded at 46.74%, followed by diclofenac-treated (35.87%), tea tree essential oil-treated (24.97%) and kaffir lime peel essential oil-treated mice (11.97%). Conclusion: These findings confirm the synergistic anti-inflammatory effects of the combined essential oils-incorporated emulsion-based gel on the carrageenan-induced hind paw edema mouse model.

Real-time monitoring of cortical brain activity in response to acute pain using wide-area Ca2+ imaging

Previous human and rodent studies indicated that nociceptive stimuli activate many brain regions that is involved in the somatosensory and emotional sensation. Although these studies have identified several important brain regions involved in pain perception, it has been a challenge to observe neural activity directly and simultaneously in these multiple brain regions during pain perception. Using a transgenic mouse expressing G-CaMP7 in majority of astrocytes and a subpopulation of excitatory neurons, we recorded the brain activity in the mouse cerebral cortex during acute pain stimulation. Both of hind paw pinch and intraplantar administration of formalin caused strong transient increase of the fluorescence in several cortical regions, including primary somatosensory, motor and retrosplenial cortex. This increase of the fluorescence intensity was attenuated by the pretreatment with morphine. The present study provides important insight into the cortico-cortical network during pain perception.

Ketogenic Diet Alleviates Mechanical Allodynia in the Models of Inflammatory and Neuropathic Pain in Male Mice.

Inflammation is involved in the induction of chronic inflammatory and neuropathic pain. Moreover, the ketogenic diet, a high-fat, low-carbohydrate, and adequate protein diet, has an anti-inflammatory effect. Thus, we hypothesized that a ketogenic diet has a therapeutic effect on both types of chronic pain. In the present study, we investigated the effect of a ketogenic diet on mechanical allodynia, a chronic pain symptom, in formalin-induced chronic inflammatory pain and nerve injury-induced neuropathic pain models using adult male mice. Formalin injection into the hind paw induced mechanical allodynia in both the injected and intact hind paws, and the ketogenic diet alleviated mechanical allodynia in both hind paws. In addition, the ketogenic diet prevented formalin-induced edema. Furthermore, the diet alleviated mechanical allodynia induced by peripheral nerve injury. Thus, these findings indicate that a ketogenic diet has a therapeutic effect on chronic pain induced by inflammation and nerve injury.