Standardized Centella asiatica extract ECa 233 alleviates pain hypersensitivity by modulating P2X3 in trigeminal neuropathic pain.

OBJECTIVE: During oral surgery and temporomandibular joint repositioning, pain hypersensitivity often occurs due to irritation or inflammation of the nerve endings in the orofacial region. This study aimed to investigate the effects of ECa 233, a Centella asiatica-standardized extract, on the development of mechanical hyperalgesia and allodynia induced by chronic constriction injury of the infraorbital nerve in mice. METHODOLOGY: The right infraorbital nerves of the mice were ligated. Oral carbamazepine (20 mg/kg) or ECa 233 (30, 100, or 300 mg/kg) was administered daily for 21 days. Von Frey and air-puff tests were performed on both sides of the whisker pad on days 0, 7, 14, and 21. Thereafter, the expression of purinergic receptor subtype 3 (P2X3) and voltage-gated sodium channel 1.7 (NaV1.7), a transmembrane protein, in the trigeminal ganglion and c-fos immunoreactivity-positive neurons in the trigeminal nucleus caudalis was assessed. RESULTS: After 21 days of infraorbital nerve ligation, the mice showed allodynia- and hyperalgesia-like behavior, P2X3 and NaV1.7 were upregulated in the trigeminal ganglion, and nociceptive activity increased in the trigeminal nucleus caudalis. However, the oral administration of carbamazepine (20 mg/kg), ECa 233 (100 mg/kg), or ECa 233 (300 mg/kg) mitigated these effects. Nevertheless, ECa 233 failed to affect NaV1.7 protein expression. CONCLUSION: Carbamazepine and ECa 233 can prevent pain hypersensitivity in mice. Considering the side effects of the long-term use of carbamazepine, ECa 233 monotherapy or combined ECa 233 and carbamazepine therapy can be used as an alternative for regulating the development of hypersensitivity in trigeminal pain. However, further detailed clinical studies should be conducted to provide comprehensive information on the use of ECa 233.

Standardized Centella asiatica extract ECa 233 alleviates pain hypersensitivity by modulating P2X3 in trigeminal neuropathic pain

Abstract During oral surgery and temporomandibular joint repositioning, pain hypersensitivity often occurs due to irritation or inflammation of the nerve endings in the orofacial region. Objective: This study aimed to investigate the effects of ECa 233, a Centella asiatica-standardized extract, on the development of mechanical hyperalgesia and allodynia induced by chronic constriction injury of the infraorbital nerve in mice. Methodology: The right infraorbital nerves of the mice were ligated. Oral carbamazepine (20 mg/kg) or ECa 233 (30, 100, or 300 mg/kg) was administered daily for 21 days. Von Frey and air-puff tests were performed on both sides of the whisker pad on days 0, 7, 14, and 21. Thereafter, the expression of purinergic receptor subtype 3 (P2X3) and voltage-gated sodium channel 1.7 (NaV1.7), a transmembrane protein, in the trigeminal ganglion and c-fos immunoreactivity-positive neurons in the trigeminal nucleus caudalis was assessed. Results: After 21 days of infraorbital nerve ligation, the mice showed allodynia- and hyperalgesia-like behavior, P2X3 and NaV1.7 were upregulated in the trigeminal ganglion, and nociceptive activity increased in the trigeminal nucleus caudalis. However, the oral administration of carbamazepine (20 mg/kg), ECa 233 (100 mg/kg), or ECa 233 (300 mg/kg) mitigated these effects. Nevertheless, ECa 233 failed to affect NaV1.7 protein expression. Conclusion: Carbamazepine and ECa 233 can prevent pain hypersensitivity in mice. Considering the side effects of the long-term use of carbamazepine, ECa 233 monotherapy or combined ECa 233 and carbamazepine therapy can be used as an alternative for regulating the development of hypersensitivity in trigeminal pain. However, further detailed clinical studies should be conducted to provide comprehensive information on the use of ECa 233.

Standardized Centella asiatica (ECa 233) extract decreased pain hypersensitivity development in a male mouse model of chronic inflammatory temporomandibular disorder.

Chronic inflammatory temporomandibular disorder (TMD) pain has a high prevalence, and available nonspecific treatments have adverse side effects. ECa 233, a standardized Centella asiatica extract, is highly anti-inflammatory and safe. We investigated its therapeutic effects by injecting complete Freund's adjuvant (CFA) into right temporomandibular joint of mice and administering either ibuprofen or ECa 233 (30, 100, and 300 mg/kg) for 28 days. Inflammatory and nociceptive markers, bone density, and pain hypersensitivity were examined. CFA decreased ipsilateral bone density, suggesting inflammation localization, which ipsilaterally caused immediate calcitonin gene-related peptide elevation in the trigeminal ganglia (TG) and trigeminal subnucleus caudalis (TNC), followed by late increase of NaV1.7 in TG and of p-CREB and activation of microglia in TNC. Contralaterally, only p-CREB and activated microglia in TNC showed delayed increase. Pain hypersensitivity, which developed early ipsilaterally, but late contralaterally, was reduced by ibuprofen and ECa 233 (30 or 100 mg/kg). However, ibuprofen and only 100-mg/kg ECa 233 effectively mitigated marker elevation. This suggests 30-mg/kg ECa 233 was antinociceptive, whereas 100-mg/kg ECa 233 was both anti-inflammatory and antinociceptive. ECa 233 may be alternatively and safely used for treating chronic inflammatory TMD pain, showing an inverted U-shaped dose-response relationship with maximal effect at 100 mg/kg.

Standardized Extract of Centella asiatica Prevents Fear Memory Deficit in 3xTg-AD Mice.

ECa 233 is a standardized extract of Centella asiatica (CA), an herb widely used in traditional Chinese and Ayurvedic medicine. Previous studies reported that ECa 233 enhanced memory retention and synaptic plasticity in the hippocampus of healthy rats. Because of this, we became curious whether ECa 233 has a therapeutic effect on the fear memory deficit in the triple transgenic Alzheimer's disease (3xTg-AD) model mice. Fear memory is a crucial emotional memory for survival that is found to be impaired in patients with early-onset Alzheimer's disease (AD). In this study, we orally administered ECa 233 (doses: 10, 30, and 100[Formula: see text]mg/kg) to 3xTg-AD mice, who were five months old, for 30 consecutive days. We found that ECa 233 prevented a cued fear memory deficit and enhanced hippocampal long-term potentiation (LTP) in 3xTg-AD mice. Subsequent proteomic and western blot analyses revealed increased expression levels of the molecules related to LTP induction and maintenance, including brain-derived neurotrophic factor (BDNF), tyrosine receptor kinase B (TrkB) and its network proteins, and extracellular signal-regulated kinase 1 and 2 (ERK1 and 2) in the hippocampi and amygdala of 3xTg-AD mice after ECa 233 pre-treatment. Our results indicate that ECa 233 is a promising potential herbal standardized extract that could be used in preventing the fear memory deficit and synaptic dysfunction before the early onset of AD.

Anti-nociceptive effects of ECa 233 a standardized extract of Centella asiatica (L.) Urban on chronic neuropathic orofacial pain in mice.

ETHNOPHARMACOLOGICAL RELEVANCE: ECa 233 is a standardized extract of Centella asiatica (L.) Urban, a herb traditionally used to treat a number of diseases including neurological disorders. Accordingly, ECa 233 showed benefits on animal models of cognitive deficits, chronic stress and Parkinson's disease. Analgesic activity of ECa 233 was shown in Tail's flick test in rodent and relieving aphthous ulcer pain in man. Moreover, acute and sub-chronic toxicity testing in rodents and pharmacokinetic study in healthy volunteers, clinical trial phase I demonstrated good safety profiles of ECa 233. AIM OF THE STUDY: This study aims to evaluate the anti-nociceptive effects of ECa 233 and its synergistic effect with gabapentin on chronic neuropathic orofacial pain after 3 weeks infraorbital nerve chronic constriction injury in mice. The peripheral and central nociceptive activities are also examined. MATERIALS AND METHODS: Chronic neuropathic orofacial pain was induced by 3 weeks infraorbital nerve chronic constriction injury. Mice were treated with ECa 233 (30, 100 and 300 mg/kg) and gabapentin (10 mg/kg) by oral gavage starting on day 21 and going on for 14 consecutive days. Mechanical hyperalgesia and allodynia were measured on day 7, 14, 21, 28 and 35 after infraorbital nerve chronic constriction injury. At the end of the experiment, mice were observed for the sedative effect using the locomotor activity, the calcitonin gen-related peptide in trigeminal ganglion and c-fos expression in trigeminal nucleus caudalis were investigated after euthanasia. RESULTS: Infraorbital nerve chronic constriction injury gradually induced marked ipsilateral mechanical hyperalgesia and allodynia. The maximum hyperalgesia and allodynia response presented on day 21 and the response was remained constant until day 35. Treatment with either 300 mg/kg ECa 233 or 10 mg/kg gabapentin were able to attenuate mechanical hyperalgesia and allodynia. The downregulation of calcitonin gen-related peptide on ipsilateral trigeminal ganglion were observed in ECa 233 at 100 and 300 mg/kg and 10 mg/kg gabapentin-treated groups. The c-fos expression on ipsilateral trigeminal nucleus caudalis was also decreased in 300 mg/kg ECa 233 and 10 mg/kg gabapentin-treated groups. CONCLUSION: ECa 233 reduced hyperalgesia and allodynia by modulating the peripheral calcitonin gen-related peptide expression consequently alleviate the nociceptive activity in trigeminal nucleus caudalis. Further clinical trial to proof ECa 233's efficacy in neuropathic pain in man as well as possible attributable mechanism of action should be further investigated.

A standardized extract of Centella asiatica (ECa 233) prevents temporomandibular joint osteoarthritis by modulating the expression of local inflammatory mediators in mice.

OBJECTIVES: To investigate the effect of a standardized extract of Centella asiatica (ECa 233), which has anti-inflammatory properties, on the local expression of the transient receptor potential vanilloid 1 (TRPV1), the acid-sensing ion channel subunit 3 (ASIC3), and the calcitonin gene-related peptide (CGRP) in the temporomandibular joint (TMJ) structure 21 days after injecting the TMJ with complete Freund's adjuvant (CFA). METHODOLOGY: A mouse model was induced by analyzing the CFA-injected TMJ on days 7, 14, and 21. We assessed TMJ histology by the osteoarthritis cartilage grade score. Then, we observed the effect of different ECa 233 concentrations (30, 100, and 300 mg/kg) and of 140 mg/kg ibuprofen doses on TRPV1, ASIC3, and CGRP local expression on day 21. RESULTS: Osteoarthritis cartilage scores were 1.17±0.37 and 3.83±0.68 on days 14 and 21, respectively, in the CFA group (n=5). On day 21, TRPV1, ASIC3, and CGRP expression significantly increased in the CFA group. In the ibuprofen-treated group, TRPV1 expression significantly decreased, but ASIC3 and CGRP showed no significant difference. All ECa 233 doses reduced TRPV1 expression, but the 100 mg/kg ECa 233 dose significantly decreased ASIC3 expression. CONCLUSIONS: TRPV1, ASIC3, and CGRP expression increased in mice with TMJ-OA on day 21. All ECa 233 and ibuprofen doses inhibited pathogenesis by modulating the local expression of TRPV1 and ASIC3. Therefore, ECa 233 was more effective than ibuprofen.

Acute enhancing effect of a standardized extract of Centella asiatica (ECa 233) on synaptic plasticity: an investigation via hippocampal long-term potentiation.

CONTEXT: ECa 233 is the standardized extract of Centella asiatica (L.) Urban. (Apiaceae). It contains at least 85% of triterpenoid glycosides and yields neuroprotective and memory-enhancing effects. However, the exact molecules exerting the effects might be triterpenic acid metabolites reproduced through gut metabolism after orally ingesting C. asiatica, not triterpenoid glycosides. OBJECTIVE: This study demonstrates the effect of unmetabolized ECa 233 on hippocampal synaptic plasticity after directly perfusing ECa 233 over acute brain slices. MATERIALS AND METHODS: The brain slices obtained from 7-week-old male Wistar rats were randomly divided into 4 groups. We perfused either artificial cerebrospinal fluid (ACSF), 0.01% DMSO, 10 µg/mL ECa 233, or 100 µg/mL on brain slices, and measured the long-term potentiation (LTP) magnitude to determine the synaptic plasticity of hippocampal circuits in each group. RESULTS: The LTP magnitude of ACSF, DMSO, 10 ug/mL ECa 233, and 100 ug/mL ECa 233 groups increased from 100% to 181.26 ± 38.19%, 148.74 ± 5.40%, 273.71 ± 56.66%, 182.17 ± 18.61%, respectively. ECa 233 at the concentration of 10 µg/mL robustly and significantly enhanced hippocampal LTP magnitude. The data indicates an improvement of the hippocampal synaptic plasticity. DISCUSSION AND CONCLUSIONS: This study emphasizes the effectiveness of triterpenoid glycosides in ECa 233 on synaptic plasticity enhancement. Therefore, this study supported and complimented the known effects of C. asiatica extract on the enhancement of synaptic plasticity, and subsequently, learning and memory, suggesting that ECa 233 could be a promising memory enhancing agent.

A standardized extract of Centella asiatica (ECa 233) prevents temporomandibular joint osteoarthritis by modulating the expression of local inflammatory mediators in mice

Abstract Objectives To investigate the effect of a standardized extract of Centella asiatica (ECa 233), which has anti-inflammatory properties, on the local expression of the transient receptor potential vanilloid 1 (TRPV1), the acid-sensing ion channel subunit 3 (ASIC3), and the calcitonin gene-related peptide (CGRP) in the temporomandibular joint (TMJ) structure 21 days after injecting the TMJ with complete Freund's adjuvant (CFA). Methodology A mouse model was induced by analyzing the CFA-injected TMJ on days 7, 14, and 21. We assessed TMJ histology by the osteoarthritis cartilage grade score. Then, we observed the effect of different ECa 233 concentrations (30, 100, and 300 mg/kg) and of 140 mg/kg ibuprofen doses on TRPV1, ASIC3, and CGRP local expression on day 21. Results Osteoarthritis cartilage scores were 1.17±0.37 and 3.83±0.68 on days 14 and 21, respectively, in the CFA group (n=5). On day 21, TRPV1, ASIC3, and CGRP expression significantly increased in the CFA group. In the ibuprofen-treated group, TRPV1 expression significantly decreased, but ASIC3 and CGRP showed no significant difference. All ECa 233 doses reduced TRPV1 expression, but the 100 mg/kg ECa 233 dose significantly decreased ASIC3 expression. Conclusions TRPV1, ASIC3, and CGRP expression increased in mice with TMJ-OA on day 21. All ECa 233 and ibuprofen doses inhibited pathogenesis by modulating the local expression of TRPV1 and ASIC3. Therefore, ECa 233 was more effective than ibuprofen.

The Effects of a Standardized Extract of Centella asiatica on Postlaser Resurfacing Wound Healing on the Face: A Split-Face, Double-Blind, Randomized, Placebo-Controlled Trial.

Background: Centella asiatica, a medicinal plant, has been used traditionally to promote wound healing. Its efficacy on promoting postlaser resurfacing wound healing is lacking. Methods: Thirty individuals with facial acne scars underwent a treatment with 2940 nm Er:YAG laser. Half side of the face was randomized to receive 0.05% w/w ECa 233 gel, a standardized extract of C. asiatica, and the other half a placebo gel. The gels were applied four times daily for 7 days then twice daily for 3 months. Erythema, melanin, and texture index (TI) from Antera3D,® and skin biophysics were obtained at baseline, days 2, 4, and 7, then every 2 weeks for the first month, and every month for 3 months. Three blinded dermatologists assessed the photographs and provided a grading scale of wound appearances. Results: The ECa 233 treated side exhibited significantly less erythema index over total follow-up by 0.03 U (coefficient = -0.03 [95% CI -0.06 to -0.0006]; p = 0.046). In keeping with the physicians' assessment that showed significantly higher improvements in skin erythema at days 2, 4, and 7 (p = 0.009, 0.0061, 0.012), crusting at days 2 (p = 0.02), and general wound appearance at days 2, 4, and 7 (p = 0.008, 0.001, 0.044), TI showed a trend toward better outcome in the ECa 233 group. Skin biophysics did not differ between the two groups. Conclusion: ECa 233 might be an option for postlaser treatment to improve wound appearance.

ECa 233 Suppresses LPS-Induced Proinflammatory Responses in Macrophages via Suppressing ERK1/2, p38 MAPK and Akt Pathways.

A current anti-inflammatory agent often targets the prevention of inflammatory disorder development. The standardized Centella asiatica ECa 233 extract has been previously reported for anti-inflammatory effect. This study aimed to investigate its anti-inflammatory effect and mechanisms of ECa 233 in lipopolysaccharide (LPS)-stimulated RAW264.7 macrophages, through 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, nitric oxide (NO) assay, reactive oxygen species (ROS) production assay, enzyme-linked immunosorbent assay (ELISA) and Western blot analysis. Our results found that ECa 233 significantly inhibited LPS-stimulated pro-inflammatory mediators production including ROS, NO and prostaglandin E2 (PGE2), and pro-inflammatory cytokines, including tumor necrosis factor (TNF)-α and interleukin (IL)-1ß without cytotoxicity. In addition, ECa 233 downregulated not only the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), but also the activation of nuclear factor-kappa B (NF-κB), activated protein kinase B (Akt), extracellular signal-regulated kinase (ERK1/2) and p38 mitogen-activated protein kinases (MAPK) induced by LPS. The inhibition of LPS-induced inflammation due to ECa 233 offered an opportunity as a tentatively potential candidate for the prevention and treatment of inflammatory diseases.

Inverted U-shaped response of a standardized extract of Centella asiatica (ECa 233) on memory enhancement.

The herb Centella asiatica has long been considered a memory tonic. A recent review found no strong evidence for improvement of cognitive function, suggesting negative results were due to limitations in dose, standardization and product variation. We used a standardized extract of C. asiatica (ECa 233) to study behavioral, cellular and molecular effects on learning and memory enhancement. ECa 233 (10, 30, and 100 mg/kg) was given orally to normal rats twice a day for 30 days. We used the Morris water maze to test spatial learning and performed acute brain slice recording to measure changes of synaptic plasticity in the hippocampus, a core brain region for memory formation. Plasticity-related protein expressions (NR2A, NR2B, PSD-95, BDNF and TrkB) in hippocampus was also measured. Rats receiving 10 and 30 mg/kg doses showed significantly enhanced memory retention, and hippocampal long-term potentiation; however, only the 30 mg/kg dose showed increased plasticity-related proteins. There was an inverted U-shaped response of ECa 233 on memory enhancement; 30 mg/kg maximally enhanced memory retention with an increase of synaptic plasticity and plasticity-related proteins in hippocampus. Our data clearly support the beneficial effect on memory retention of a standardized extract of Centella asiatica within a specific therapeutic range.

Safety and Pharmacokinetics of Standardized Extract of Centella asiatica (ECa 233) Capsules in Healthy Thai Volunteers: A Phase 1 Clinical Study.

The aim of this study was to investigate the safety and pharmacokinetic profiles of a newly developed, standardized extract of Centella asiatica (ECa 233) capsule in healthy Thai volunteers. This study was designed as an open-labeled, 2-sequence dosage, single- and repeated-dose study investigated under fasting conditions. Plasma concentrations of the parent compounds and their relative acid metabolites were measured and pharmacokinetic parameters were calculated using noncompartmental analysis. Tolerability was assessed based on physical examinations, monitoring of vital signs, clinical laboratory tests, and any observed adverse events. A key finding of this study was that the pharmacokinetics of ECa 233 in healthy volunteers did not correspond with its pharmacokinetics in animal studies. As indicated in human pharmacokinetic parameters, maximum plasma concentration and area under the curve of the parent compounds (madecassoside and asiaticoside) were very low, while their respective metabolites (madecassic acid and asiatic acid) demonstrated higher values. Based on the pharmacokinetic results observed in the dose comparison, accumulation of active metabolites after repeated dose is highly suggestive. In addition, the asiatic acid profile showed 2-fold increase in Cmax and AUC(0-t) after increasing dose from 250 to 500 mg of ECa 233. Lastly, the safety and tolerability evaluation illustrated that single and multiple doses in both 250 and 500 mg oral administration of ECa 233 were well tolerated, and none of the volunteers discontinued their participation due to adverse effects during the study.

Effects of Centella asiatica extract on antioxidant status and liver metabolome of rotenone-treated rats using GC-MS.

Centella asiatica has been used as a culinary vegetable or medicinal herb. In this study, the hepatoprotective effect of the standardized extract of C. asiatica (ECa233) in rotenone-treated rats was examined using a GC-MS-based metabolomic approach. ECa233 contains >80% triterpenoids with a ratio of madecassoside to asiaticoside of 1.5(±0.5):1. Rats were randomly divided into three groups (with six rats/group): sham negative control, rotenone positive control and the ECa233 test group. Rats in the ECa233 group received 10 mg/kg ECa233 orally for 20 days, followed by 2.5 mg/kg intraperitoneal rotenone injection to induce toxicity before being sacrificed. Metabolomic analysis showed that supplementation of ECa233 protected rat liver against rotenone toxicity. Pipecolinic acid was one of the most important metabolites; its level was decreased in the rotenone group as compared with the control. Supplementation with ECa233 before administration of rotenone raised pipecolinic acid to levels intermediate between controls and rotenone alone. The metabolomics approach also helped discover a possible new genuine epimetabolite in the present work. Antioxidant tests revealed that ECa233 inhibited lipid peroxidation and increased catalase activities in liver tissue.

Effect of Centella asiatica on pathophysiology of mild chronic cerebral hypoperfusion in rats.

OBJECTIVE: The present study investigated the effect of standardized Centella asiatica extract on cognition and hippocampal pathology of mild chronic cerebral hypoperfusion (CCH) that was induced by permanent right common carotid artery occlusion (RCO) in rats. MATERIALS AND METHODS: Sixty-four male Sprague-Dawley rats were randomly divided into four groups of Sham-veh, Sham-C. asiatica, RCO-veh and RCO-C. asiatica, which were further divided into short-term and long-term CCH induction. Oral treatments with 20 mg/kg C. asiatica initiated 24 hours and 12 months after CCH and continued for 14 consecutive days. According to the cognition and histopathological evaluation period, the experiment was divided into 2 sets of either 2 or 12 months of CCH. RESULTS: Results showed that 2-month CCH induced learning flexibility deficit associated with CA1 neuronal damage and internal capsule (IC) astroglia activation. Long-lasting (12 months) mild CCH induced spatial learning, memory and flexibility deficits associated with progressive dorsal hippocampal damage. Treatment with 20 mg/kg of C. asiatica improved learning flexibility deficit after 2 and 12 months of CCH. C. asiatica ameliorated neuronal damage in the dorsal hippocampus at 2 months of CCH when given 24 hours after CCH onset. Treatment with C. asiatica after 12 months of cerebral blood flow reduction improved memory and learning flexibility deficits and was associated with the dentate gyrus neuronal damage reduction. CONCLUSION: Our finding indicates the therapeutic potential of C. asiatica either when given immediately after ischemic insult or when administered one year after ischemic insult, in a CCH rat model.

Modulation of Neuronal Activity on Intercalated Neurons of Amygdala Might Underlie Anxiolytic Activity of a Standardized Extract of Centella asiatica ECa233.

GABAergic intercalated neurons of amygdala (ITCs) have recently been shown to be important in the suppression of fear-like behavior. Effects of ECa233 (a standardized extract of Centella asiatica), previously demonstrated anxiolytic activity, were then investigated on ITCs. Cluster of GABAergic neurons expressing fluorescence of GFP was identified in GAD67-GFP knock-in mice. We found that neurons of medial paracapsular ITC were GABAergic neurons exhibiting certain intrinsic electrophysiological properties similar to those demonstrated by ITC neurons at the same location in C57BL/6J mice. Therefore, we conducted experiments in both C57BL/6J mice and GAD67-GFP knock-in mice. Excitatory postsynaptic currents (EPSCs) were evoked by stimulation of the external capsule during the whole cell patch-clamp recordings from ITC neurons in brain slices. ECa233 was found to increase the EPSC peak amplitude in the ITC neurons by about 120%. The EPSCs in ITC neurons were completely abolished by the application of an AMPA receptor antagonist. Morphological assessment of the ITC neurons with biocytin demonstrated that most axons of the recorded neurons innervated the central nucleus of the amygdala (CeA). Therefore, it is highly likely that anxiolytic activity of ECa233 was mediated by increasing activation, via AMPA receptors, of excitatory synaptic input to the GABAergic ITC leading to depression of CeA neurons.

Neuroprotective effect of a standardized extract of Centella asiatica ECa233 in rotenone-induced parkinsonism rats.

BACKGROUND: Mitochondrial dysfunction and reactive oxygen species (ROS) generation cause dopaminergic neurodegeneration in Parkinson's disease. The neuroprotective approach is a promising strategy to slow disease progression in Parkinson's disease. A standardized extract of Centella asiatica ECa233 has been previously reported to have pharmacological effects in the central nervous system. PURPOSE: This study aimed to determine the neuroprotective effect and mechanisms of ECa233 in rotenone-induced parkinsonism rats. METHODS: Rats were orally given either vehicle or ECa233 (10, 30 and 100 mg/kg) for 20 consecutive days. Rotenone (2.5 mg/kg i.p.) was given to parkinsonism (PD) and ECa-treated rats from day 15 to 20. Locomotor activity was recorded on day 1, 14, 17 and 20. Tyrosine-hydroxylase (TH) immunohistological staining was used to determine dopaminergic neurons in the substantia nigra and striatum. Furthermore, mitochondrial complex I activity, malondialdehyde (MDA) levels, superoxide dismutase (SOD) and catalase protein expression were measured in brain tissue. RESULTS: Rats receiving ECa233 30 mg/kg showed a significant increase in distances (p < 0.01) together with a higher number and intensity of dopaminergic neurons in the substantia nigra and striatum (p < 0.001) compared to PD rats. ECa233 (30 mg/kg) protected against mitochondrial complex I inhibition, decreased MDA levels (p < 0.05) and increased SOD (p < 0.01) and catalase (p < 0.05) expression. CONCLUSION: ECa233 can protect against rotenone-induced motor deficits and dopaminergic neuronal death. These effects are mediated through the protection of mitochondrial complex I activity, the effects of antioxidants and the enhancement of antioxidant enzyme expression.

Induction of keratinocyte migration by ECa 233 is mediated through FAK/Akt, ERK, and p38 MAPK signaling.

Centella asiatica is widely considered the most important medicinal plant for treating and relieving skin diseases. Recently developed standardized extract of Centella asiatica ECa 233 has demonstrated positive effects on wound healing of incision and burn wound in rats. However, knowledge associated with wound healing mechanism of ECa 233 was scare. Therefore, this study aimed to investigate the effect and underlying molecular mechanisms of ECa 233 on the migration of a human keratinocyte cell line (HaCaT) using scratch wound healing assay. Formation of filopodia, a key protein in cell migration as well as signaling pathways possibly involved were subsequently assessed. It was found that HaCaT cell migration was significantly enhanced by ECa 233 in a concentration- and time-dependent manner. The filopodia formations were accordingly increased in exposure to ECa 233 at concentrations of 0.1-100 µg/ml. Furthermore, ECa 233 was found to significantly upregulate the expression of Rac1 and RhoA and to induce phosphorylation of FAK and Akt as well as ERK and p38 MAPK. Taken all together, it is suggestive that ECa 233 induces cell migration and subsequently promotes wound healing activity, through the activation of FAK, Akt, and MAPK signaling pathways thereby supporting the role of ECa 233 to be further developed for the clinical treatment of wound.

Comparative pharmacokinetics between madecassoside and asiaticoside presented in a standardised extract of Centella asiatica, ECa 233 and their respective pure compound given separately in rats.

1. ECa 233, the standardised extract of Centella asiatica, contains not less than 80% triterpenoid glycosides, in a madecassoside:asiaticoside ratio of 1.5 (±0.5):1. 2. The pharmacokinetic comparison of madecassoside and asiaticoside was performed in rats following intravenous and oral administration of ECa 233, or an equivalent dose of the individual compounds. Blood, tissues, urine and faeces were collected after dosing to determine drug and metabolite levels using liquid chromatography-tandem mass spectrometry. 3. Our study demonstrated that plasma levels of madecassoside, and to a lesser extent asiaticoside, were higher after administration of ECa 233 than the corresponding values for the pure compounds. There was a bidirectional interconversion between asiaticoside and madecassoside consistent with the increased exposure of madecassoside and asiaticoside in ECa 233. 4. Both madecassoside and asiaticoside appeared to be widely distributed in several organs and metabolized extensively; following intravenous administration of either compound, approximately 80-90% of the dose was recovered as madecassic acid and asiatic acid in the faeces.

Pharmacokinetics of a Standardized Extract of Centella asiatica ECa 233 in Rats.

ECa 233, a standardized extract of Centella asiatica, has been found to exhibit various positive neurological effects and to have a good safety profile. The present study aimed to explore the disposition kinetics of ECa 233, containing madecassoside (53.1 %) and asiaticoside (32.3 %), in rats. The extract was intravenously or orally administered at doses from 50 to 200 mg/kg. Plasma, tissues, urine, and feces were collected at time points from 0 to 48 h after dosing. The levels of madecassoside and asiaticoside, as well as their postulated triterpenic metabolites, madecassic acid and asiatic acid, in biological samples, were simultaneously measured by liquid chromatography-tandem mass spectrometry. The results showed that all animals had a good tolerability for ECa 233, whereas madecassic and asiatic acids were found in negligible amounts after pharmacokinetic assessment. Madecassoside and asiaticoside demonstrated rather similar absorption and tissue distribution profiles. They were rapidly absorbed, reaching maximum levels within 5-15 min after oral administration, but they had poor oral bioavailability, less than 1 %. Both triterpenoids were extensively distributed in the brain, stomach, and skin within 1 h and remained there for at least 4 h after dosing. Madecassoside and asiaticoside in ECa 233 were mainly excreted as an unchanged form after being injected, and exclusively as triterpenic acid metabolites in feces after oral administration. The pharmacokinetic results obtained could provide some guidance for an appropriate dosing regimen of ECa 233 in future studies. This study also provided the first evidence demonstrating the presence of madecassoside and asiaticoside in their target tissues.