Gotu Kola

Last Updated: September 28 2022

Centella asiatica (Gotu Kola) is a traditional medicine mainly renowned for its cognitive enhancing properties (usually alongside bacopa monnieri) and its ability to regenerate wound healing. It appears effective on both parameters in preclinical evidence, and may also be anti-rheumatic.

Gotu Kola is most often used for.



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    1.

    Sources and Structure

    1.1

    Sources

    Centella asiatica (of the family Apiaceae, also known as Umbellifere[1]) is a rasayana herb from Ayurveda that is also referred to as Gotu kola (from Traditional Chinese Medicine[1]), Indian Pennywort, Jal Brahmi and Mandookaparni[2] as well as the Japanese term 'Tsubokusa'.[3] It is botanically synonymous with Hydrocotyle asiatica[4] and a few of its names (Indian Pennywort and Brahmi) and ranks (Aindri, Medhya Rasayana) are used synonymously with Bacopa monnieri, as the two have an intertwined medicinal history.[1]

    This herb appears to be traditionally used for leprosy, varicose veins, a 'blood purifier', ulcers, lupus and certain eczemas, general longevity, and of mental retardation[4][1] as well as one of its more popular usages being both internal and topical use for skin health (wound, scar, and burn healing).[5] It is commonly used alongside bacopa monnieri for the purpose of cognitive enhancement, insofar that the term 'Brahmi' (used to refer to bacopa monnieri) sometimes is used to refer to the combination of the two herbs.[6]

    The herb itself has been reported to be tasteless and odorless with small fan-shaped green leaves with white or light purple-to-pink or white flowers; it bears small oval shaped fruits and grows in watery areas.[1]

    Centella asiatica is an ayurvedic herb that is seen as a cognitive enhancer, with its past being fairly intertwined with that of bacopa monnieri. Beyond neural regeneration, it appears to be used for purposes related to anti-inflammation (lupus and eczema), skin regeneration, and circulation (varicose veins)

    1.2

    Composition

    The aerial parts of centella asiatica tend to contain:

    • Asiatic acid (0.72-0.98% dry weight[7])[8] and its glycoside known as Asiaticoside (0.3% dry weight in chloroform, methanol, and ethyl acetate fractions yet 0.04% in water and ethanolic[4] but much higher (1.63-2%) in the plant extract[7]). Asiatic acid consists of 26.7% of the total triterpenoids[9]
    • Madecassic acid (0.72-0.95% dry weight[7]) and its glycoside Madecassoside (1.27-1.7%[7]); the isomer of Madecassoside being Asiaticoside B (or Terminoloside as a synonym)[7] and madecassic acid saponins consist of 25.53% total saponins[9]
    • Madasiatic acid,[10] Asiaticoside F-G,[11] quadranoside IV,[11] and Sceffoleoside A (0.0068%)[10]
    • Thankuniside and Isothankuniside (glycosides)[1]
    • Centellasaponins B (0.0063%), C (0.0084%), and D (0.0036%)[10] as well as Centellosides A and B[12]
    • Brahmoside and Brahminoside[1]
    • Ursolic acid (Triterpenoid)[8]
    • Other triterpenoids such as 2α,3β,20,23-tetrahydroxyurs-28-oic acid,[13] 11,12-dehydroursolic acid lactone, pomolic acid, corosolic acid, 2α,3α-dihydroxyurs-12-en-28-oic acid, and 3-epimaslinic acid[8]
    • Centellin and Centellicin[14]
    • Rosmarinic acid[8]
    • Ginsenosides Mc, Rk1, Rg5, Rd2, and Y[12]
    • (20R)-ginsenoside Rg3 and (20S)-ginsenoside Rg3[12]
    • Notoginsenoside ST-4 and Fe[12]
    • Gypenoside Iβ and η[12]
    • 8-acetoxy-1,9-pentadecadiene-4,6-diyn-3-ol[8]
    • The alkaloid hydrochotine[1]
    • Apigenin at a comparable concentration as bacopa monnieri[15]
    • Rutin (yet no Quercetin or Quercetrin content)[15]
    • Irbic acid (3,5-O-dicaffeoyl-4-O-malonilquinic acid)[16] and other phenolic acids including 3,5-Di-O-caffeoyl quinic acid, 1,5-di-O-caffeoyl quinic acid, 3,4-di-O-caffeoyl quinic acid, and 4,5-di-O-caffeoyl quinic acid[3]
    • Kaempferol,[3] kaempferol-3-O-β-D-glucoside,[3] and Castilliferol (ie. kaempferol-3-p-coumarate)[17]
    • Quercetin,[3] quercetin-3-O-β-D-glucoside,[3] and castillicetin (Quercetin-3-caffeate)[17]
    • Chlorogenic acid[3] and Isochlorogenic acid[17]
    • Cadiyenol[18]
    • Vitamin C at 255.7mg/100g (0.25%)[19]
    • β-carotene (42.5-73.7μg/g of the leaves[20]), trans-lutein (82.6-133.5μg/g[20]), trans-neoxanthin (5.9-14.2μg/g[20]), trans-violaxanthin (20.5-26.3μg/g[20]), and other unidentified carotenoid structures[20] with total carotenoid reaching 17.58mg/100g[19]
    • Anthocyanins at up to 37.6mg/100g[19]
    • Chlorophyll at 29.91mg/100g[19]
    • Iron (29-74.3mg/100g[20]), potassium (3,079-6,295mg/100g[20]), copper (2.6-6.4mg/100g[20]), zinc (11.3-45.3mg/100g[19][20]), calcium (1,150.9-2,206.1mg/100g[20][19]), sodium (1,114.2-2,597.1mg/100g[20]), manganese (54.5mg/100g[19]), magnesium (240.7-841.3mg/100g[20]), cobalt (1.34+/-0.03mg/100g[19])
    • β-sitosterol 3-Oβ-glucopyranoside[8] as well as both campesterol and stigmasterol[1]

    image

    The main bioactives appear to be the triterpenoid structures, with the two major triterpenoids being asiatic acid and madecassic acid (those two and their glycosides being at around 2% of plant weight each and being a quarter of total triterpenoids each). The ursolic acid and rosmarinic acid contents may also be bioactive, and oddly this plant seems to contain ginsensosides that are not found in any species of ginseng

    As a vegetable product, centella asiatica has an 83.43% moisture content.[19] The dry weight (overall weight minus moisture) is 13.26% ash content, 8.24% fiber, 6.13% protein, 0.66% fat and the remaining carbohydrate.[19]

    Overall, centella asiatica appears to have a relatively low flavonoid and phenolic content[21] although a high tannin concentration (20-25%).[1] The total centelloside content ('Centelloside' being a term used to collectively refer to asiatic acid, madecassic acid, and their glycosides) is concentrated in the leaf extracts[22]) at around 20-80mg/g (2-8%) in the leaves depending on age and nutrient supply.[23]

    There may be a metal chelating property with centella asiatica, with the hexane fraction showing a peak metal chelating activity (IC50 of 90µg/mL) in vitro with iron sulfate.[24]

    Centella asiatica appears to have antioxidant properties with both the aqueous and ethanolic extracts as assessed by FRAP, DPPH, iron reduction and nitric oxide scavenging; the water extract exceeding the ethanolic in potency[21] and being comparable in potency to Grape seed extract,[25] Vitamin C,[25] and bacopa monnieri.[21]

    1.3

    Structure and Properties

    The two main components (asiatic acid and medecassic acid) are of the saponin class, but in particular they are pentacyclic (five ringed) triterpenoid saponins of the ursane class; a structurally designation similar to corosolic acid and ursolic acid.

    Asiaticoside appears to be poorly soluble in both ethanol and water and more soluble in chloroform, ethyl acetate, and methanol.[4]

    1.4

    Formulations

    ECa-233 is a standardized extract of centella asiatica (no less than 80% triterpenoid saponins with a ratio of madecassoside to asiaticoside in the range of 1.5+/-0.5).[26][27]

    2.

    Pharmacology

    2.1

    Serum

    An oral intake of 100mg/kg madecassoside in rats appears to reach a maximum concentration (Cmax) of 303.75+/-28.53ng/mL at (Tmax) 0.9 hours after ingestion, exhibiting a half-life of 3.47+/-0.68 hours.[28] A double peak was noted, suggesting variations in intestinal absorption of enterohepatic circulation (cause not confirmed).[28]

    In humans, 30-60mg of the total triterpenoid fraction taken twice daily over the course of six days (with measurements after the first dose and after the week of dosing) noted that 30mg was able to raise plasma levels to around 700+/-110ng/mL after a single dose and chronic treatment rose this to 1.03+/-0.05µg/mL (Tmax of 4.1-4.5 hours) whereas 60mg acutely rose plasma levels to 1.36+/-0.13µg/mL and chronic treatment to 1.69+/-0.07µg/mL (Tmax of 4.2 hours).[29]

    The half life in humans was quite variable, with differing values with acute intake of 30mg (2.20+/-0.30 hours) and 60mg (3.40+/-0.68) as well as chronic intake of 30mg (6.33+/-1.82) and 60mg (10.28+/-1.8).[29]

    Oral intake of madecassoside at high doses reaches a nanomolar concentration in the blood, while it appears that the standard recommended doses of total triterpenoid saponins (30-60mg twice daily) reaches the 1-2µg/mL range

    2.2

    Distribution

    Asiatic acid is known to bind to human serum albumin, particularly the IIA subdomain, which was mostly a hydrophobic interaction with a ΔG of −6.36 kcal/M-1.[30]

    2.3

    Phase I Enzymatica Interactions

    In vitro, the CYP2C19 enzyme is inhibited by centella asiatica extracts at 615.1μg/mL (water extract), 30.2μg/mL (dichloromethane), 61.2μg/mL (ethanol) and 112.5μg/mL (hexane) with inhibition being noted with isolated asiatic acid (117.5μg/mL or 240.4μM), asiaticoside (408.3μg/mL or 425.7μM), madecassic acid (92.5μg/mL or 183.3μM) and rosmarinic acid (2265.5μM).[31]

    The inhibition on CYP2C19 appears to be significantly higher than serum levels, and so while there is technically inhibition the practical relevance of this information is not known

    3.

    Molecular Targets

    3.1

    MAPKs

    The MAPKs are three proteins involved in cellular growth and proliferation known as ERK (or MAPK), JNK (or SAPK), and p38. Centella asiatica has been noted to inhibit ERK and p38 activation in cardiomyocytes,[32] p38 in endothelial cells,[33] and all three in macrophages.[34]

    Asiaticoside has been noted to induce transcription of DUSPP and DUSP3 fairly rapidly after incubation at 30μg/mL (fibroblasts)[35] which are known to be negative regulators of the MAPK family.[36][37] It was also noted that, in macrophages, all MAPKs were successfully suppressed in the presence of LPS due to inhibiting the activation of Raf-1[34] which is known to mediate the induction of MAPKs in response to LPS.[38][39]

    In general, the MAPKs appear to be inhibited although which particular MAPK (p38, ERK, or JNK) that is inhibited depends on the tissue in question; this can be traced back to either induction of DUSPP and DUSP3 genes (which suppress MAPK activity) or dysregulating the ability of Ras/Raf-1 to increase MAPK activity

    In neuronal cells exclusively, centella asiatica appears to phosphorylate (activate) ERK[26][40][41] which underlies the protective and growth effects of this herb on brain cells.

    Activation of the BDNF receptor is known to increase ERK activity in neurons[42] and centella asiatica itself increases BDNF in vivo[43] suggesting a plausible mechanisms that is exclusive to neurons. An alternate and more plausible hypothesis is seen where asiaticoside stimulates CREB phosphorylation which then causes an increase in BDNF secretion[40] and this increase in CREB phosphorylation is blocked by inhibiting the upstream regulator of ERK (a protein known as MEK1[40] which couples Ras/Raf-1 signalling to ERK[44]) and the asiaticoside induced neuronal growth is blocked by MEK1 inhibition.[41]

    Another alternate explanation, enhancing the activities of NGF (another growth factor) at 1μM asiaticoside, is also tied into this pathway as it relies on MEK1 activation[41] so regardless of which growth factor is causative the protein of MEK1 and the overall Ras/Raf-1/MEK1/ERK pathway is implicated.

    ERK phosphorylation is increased due to MEK1 activation, and this is known to cause neuronal growth in vitro (and since this has been observed in animals following oral ingestion of the plant, it is thought to be biologically relevant). While the exact mechanisms are not fully elucidated, it seems that MEK1 activation (unknown how this is activated) causes an increase in CREB phorphorylation and a greater production of BDNF

    3.2

    Cytokines

    In isolated THP-1 macrophages, asiaticoside has been found to augment the MCP-1 induced release of the macrophage-derived cytokine known as IL-1β, which occurs at a remarkably low concentration of 10pM to 100nM.[45]

    The increase in MCP-1 induced IL-1β concentrations appears to occur at incredibly low concentrations (picomolar range) and thus this mechanism is most likely relevant to oral supplementation of centella asiatica; possibly related to the anti-rheumatic properties of this herb

    3.3

    Phospholipase A2

    Centella asiatica has also been shown to inhibit phospholipase A2 on cellular surfaces in vitro[46] in cerebellum slices in a concentration dependent manner between 31.5–500µg/mL (250-500µg/mL nearly abolishing activity of iPLA2 and 12.5µg/mL being the lowest dose effective against cPLA2 whereas 1.25µg/mL was ineffective).[47] While thought to be neuroprotective (since elevated phospholipase A2 activity in the brain causes cellular changes associated with neurodegeneration[48]), it is uncertain what role this plays since the more resistant variant (iPLA2) consists of up to 80% of phospholipase A2 in the brain[47] and serum levels of asiaticoside are known to reach up to 1-2µg/mL in humans[29] which is in the ineffective range for cPLA2.

    While there may be a direct inhibitory effect on phospholipase A2 variants in the brain, they both appear to occur at concentrations that are above what is seen with oral supplementation of centella asiatica and they may not occur following oral ingestion

    4.

    Neurology

    4.1

    Cholinergic Neurotransmission

    Young mice given 200mg/kg of centella asiatica for 15 days appear to have a modest increase in hippocampal acetylcholinesterase activity[49] although 150-300mg/kg over 6 weeks in a model of aluminmum toxicity (which would normally increased acetylcholinesterase) prevented such an increase.[50]

    Concentrations of up to 250μg/mL of the water extract (lacking the standard saponins) have failed to inhibit acetylcholinesterase activity.[51]

    Overall, there do not appear to be any significant or known interactions with cholinergic neurotransmission

    4.2

    Glutaminergic Neurotransmission

    Incubation of centella asiatica water extract (lacking the standard saponins) at concentrations up to 200μg/mL have failed to protect cells from glutamate induced cell death[51] whereas isolated asiatic acid has been shown to protect cells from glutamate induced cell death at 10nM (100mg/kg oral intake in mice).[52]

    The saponins (asiatic acid) appear to be protective against glutamate induced cell death, and the mechanisms underlying this are not yet known

    4.3

    Neurogenesis

    Neurotrophy (growth of neurons) appears to occur via activation of the MERK/ERK or the PI3K/Akt pathways,[53][54] which are downstream of several neurotrophic factors such as BDNF, NT-3, and NGF acting on their receptors.[55][42] Centella asiatica saponins have been found to phosphorylate ERK and Akt and abolishing either of these pathways prevents the neurotrophic actions of this herb.[26]

    In isolated IMR-32 neuroblastoma cells, a centella asiatica extract (52% madecassoside and 32% asiaticoside) at 1-100μg/mL (but not 100nM) failed to proliferate neurons yet promoted neurite outgrowth in a time dependent but not concentration dependent manner with a potency comparable to 100ng/mL BDNF.[26] Elsewhere, in the presence of NGF (another neuronal growth factor) ethanolic extracts of centella asiatica were shown to augment NGF-induced neurite outgrowth thought to be mostly due to isolated asiatic acid (active at 1μM) which acted via ERK.[41]

    The saponins in centella asiatica appear to promote neuronal growth

    In a rat model of Parkinson's Disease (MPTP toxicity), it was noted that while neither control nor the toxin control experienced any changes in BDNF levels the groups given oral madecassoside (15-60mg/kg) experienced a dose-dependent increase in BDNF concentrations reaching about 1.5-fold at the highest tested dose.[43]

    At least one study has traced back the increase in BDNF to CREB phosphorylation, and the CREB phosphorylation to the activation of MAPKs.[40]

    Centella asiatica may induce BDNF expression after oral, which is likely due to CREB being phosphorylation from MAPKs in neurons

    Oral ingestion of centella asiatica has been confirmed to increase dendritic arborization at 200mg/kg for 15 days in young mice (apical, basal, and branching points),[49] at 1.58-4.74g/kg (dry leaf equivalent given via juice) for six weeks in rat pups in the amygdala,[56] and in rat pups given the same dosage of juice but in the hippocampus.[57]

    The aforementioned studies were in youth, although increased dendritic arborization has been noted in adult rat hippocampuses given the leaf juice at the highest aforementioned dose (4.74mg/kg appeared to be significant whereas 1.58-3.16mg/kg were not)[58] as well as the amygdala.[59] The increases also took six weeks to manifest, but reached a 47–118% increase in apical dendritic intersections (111% increase in branching points) and 47–90% increase in basal intersections (105% increase in branching points).[58]

    When investigating the number of dendritic interactions within a particular distance from the neuron, all dendritic interactions between 20-100 microns in length appear to be enhanced.[57][49][56] Furthermore, there appears to be a time-related as well as dose-related response with up to four weeks being required for increased dendritic arborization.[49][56][57] Adult subjects also experience increases in most lengths of dendrites and have a time-related benefit, but the time may be more delayed (requiring six weeks for a dosage that is effective in four for youth).[58]

    This increased growth of neurons appears to occur in living creatures who ingest the herb at reasonable dosages. While this occurs in both youth and adults rats, adults seem a bit more resistant to the changes

    4.4

    Neuroprotection

    Centella asiatica has shown neuroprotective efficacy at an oral intake of 150-300mg/kg in rats when ingested over six weeks alongside aluminum (neurotoxic dose),[50] against a model of stroke when 100-300mg/kg of the water extract is preloaded for 21 days where centella asiatica exerts a dose dependnet effective with maximal efficacy in reducing infarct volume from 40% to 10%,[60] protects mice against β-amyloid toxicity,[51] monosodium glutamate,[61] and oral doses as low as 5mg/kg centella asiatica (8.0–8.6% saponins and 0.8% asiaticoside) for 10 days prior to 3-NPA toxicity have shown some protective effects.[62][63]

    When looking at various toxic components, centella asiatica appears to be generally neuroprotective and with a potency that isn't overly remarkable but still somewhat respectable. In particular, it has been noted to occur at low oral doses suggesting that this is relevant to oral supplementation of centella asiatica

    Seven days ingestion of madecassoside (15-60mg/kg) prior to an injection of MPTP (neurotoxin that mimicks early phase Parkinson's Disease[64]) was able to dose-dependently reduce motor deficits as assessed by a ladder walking test, with 60mg/kg effectively normalizing performance relative to control.[43] 30-60mg/kg were also able to significantly preserve dopamine and attenuate changes in oxidative (Glutathione and TBARS) and genetic (Bcl but not Bax) alterations, but neither dose exerted absolute protection.[43]

    An increase in BDNF was seen with madecassoside[43] which is known to protect neurons from MPTP[65][66] via inducing Bcl-2 and reducing cellular death[67][68] (more cell preservation causing a preservation of dopamine synthesis) suggesting that this is the mechanism of action.

    Centella asiatica is thought to be neuroprotective via the same mechanism that it induces neuronal growth, via inducing BDNF protein levels

    4.5

    Anxiety

    In non-stressed mice, 100-300mg/kg of the ECa-233 extract showed slightly anxiolytic properties[27] and a juice extract for 15 days in otherwise normal mice has shown anxiolytic properties in a hole-board test.[49] When looking at various extracts of centella asiatica, it seems that while the water extract (200mg/kg) is ineffective in reducing anxiety on an elevated plus maze test both the chloroform and methanolic extracts at the same dose are effective.[69]

    In stressed mice, the increase in corticosterone is hindered with 10mg/kg and abolished by 30-100mg/kg (similar to 2mg/kg and 10mg/kg diazepam) whereas anxiety is abolished by 100mg/kg;[27] the isolated madecassoside (16mg/kg) and asiaticoside (10mg/kg) in doses equivalent to the 30mg/kg dosage were significantly less effective.[27]

    Animal studies suggest anxiolytic properties somewhat comparable to diazepam albeit requiring a higher dosage, and these may occur following a single dose of the supplement. They may occur even if the subject is not inherently stressed, albeit to a lesser degree than anxious/stressed subjects

    A 70% ethanolic extract of the aerial parts of centella asiatica at 500mg twice daily (1,000mg total) in persons with generalized anxiety disorder over the course of 60 days noted time dependent reductions in anxiety that reached 13.1% at 30 days and 26% at the end of the trial.[2]

    Comparable reductions were noted in self-reported stress (12.5% at 30 days and 23.2% at trial end) and depression (10.2% and 21.8%, respectively).[2]

    Anxiety reduction has been reported in humans given oral supplementation of the plant, and this appears to be due to the saponin content

    4.6

    Analgesia

    1-10mg/kg asiatic acid appears to have analgesic effects against acetic-acid induced writhing, the late phase of the formalin test, and a carrageenan test with 10mg/kg asiatic acid being comparable to 10mg/kg indomethacin. Other parameters measured such as serum cytokines (TNF-α and IL-1β) and oxidative changes were identical between 10mg/kg asiatic acid or indomethacin yet asiatic acid seemed more effective at reducing COX-2 and NF-kB activation.[70]

    Appears to have pain killing properties

    4.7

    Depression and Mood

    Oral ingestion of 250-750mg centella asiatica extract (3% polyphenolics and 5% asiaticoside plus asiatic acid) given either acutely or over the course of 1-2 months in otherwise healthy older adults there was a trend to increase P300 latency (failed to reach statistical significance) and while P300 amplitide and N100 latency were wholly unaffected there appears to be a significant increase in N100 amplitude after two months of supplementation to near doubling.[71] These changes were associated with an increase in calmness, contentment, and alertness.[71]

    4.8

    Attention

    An increase in attention has been noted with 1,000mg centella asiatica in two divided doses, reaching 13.4% after 30 days and 27.8% after 60 days, which is thought to be secondary to reduced symptoms of anxiety and stress in persons with generalized anxiety disorder.[2] Elsewhere, otherwise healthy older adults given 250-750mg of an extract containing 5% asiaticoside plus asiatic acid for two months there was a significant reduction in choice, numerical working memory, and spatial memory reaction time but not simple reaction nor digit vigilance time.[71]

    The usage of centella asiatica for ADHD has only been tested in conjunction with a large amount of herbs (Paeoniae Alba, Ashwagandha, Spirulina, Bacopa Monnieri, and Lemon Balm) which, while effective, cannot be used to evaluate the efficacy of centella asiatica itself.[72]

    A single dose of 12g of centella asiatica (basic herb) was able to significantly attenuate the acoustic startle response (ASR) in otherwise healthy persons at 30-60 minutes after ingestion without significantly affecting mood state, suggesting anxiolytic and anti-fear properties.[73]

    4.9

    Memory and Learning

    Mechanistically, in vitro application of a centella asiatica extract has noted CREB phosphorylation in neuroblastoma cells at a concentration range of 1-200µg/mL with an EC50 of 26µg/mL; this was due to the saponin content, where asiaticoside and madecassoside possess EC50 values of 3.8µM and 5.7µM respectively.[40] This CREB phosphorylation was blocked with the MAPK inhibitor PD98059 (MEK1 inhibitor, MEK1 mediating the influence of Raf-1 on ERK[44]) and enhanced CREB phosphorylation was shown in vitro to enhance BDNF secretion[40] which is thought to underlie the brain's arborization[57] and regeneration;[41] two phenomena seen with centella asiatica.

    It was confirmed that PKA, nitric oxide, and the NMDA receptor (all known to stimulate CREB) were uninvolved with the actions of centella asiatica and its components.[40]

    Appears to enhance CREB phosphorylation, and this occurs at a low enough concentration that it is probably relevant to oral supplementation (perhaps higher doses, however). This is secondary to MAPK activation, and ultimately results in increased BDNF production

    When looking at studies assessing the anti-amnesiac effects of centella asiatica, it appears to have protective effects against β-amyloid (as assessed by the Tg2576 genetic strain of mice) at 200mg/kg of the water extract,[51] against streptozotocin-induced cognitive impairment (100-300mg/kg),[74] D-galactose induced amnesia,[75] and pentylenetetrazole-induced cognitive impairment with the 100-300mg/kg dose.[76] When looking at studies investigating solely saponins, asiatic acid at 100mg/kg is able to reduce cognitive impairment in a model of glutamate excitotoxicity.[52]

    In looking at anti-amnesiac mechanisms, a beneficial trend in oxidation is seen[51][74][75][76] despite the water extract (used in some studies and free of both asiatic acid and madecassic acid) not possessing direct antioxidative efficacy.[51]

    In regards to the anti-amnesiac effects of centella asiatica, there appears to be components in the water extract (excludes the saponins) as well as the saponins themselves which have protective effects on neurons. The anti-amnesiac effects are respectable but not absolute even at the highest effective tested dose (300mg/kg in rats)

    Mice (young and adult) given 200mg/kg centella asiatica daily for 15 days appeared to experience increased performance in a radial arm maze[49] and elsewhere a study over 14 days using 200mg/kg of one of three extracts (water, methanolic, or chloroform) noted that only the water extract was effective and that while 200mg/kg was more effective than 100mg/kg that it was equivalent to 300mg/kg (assessed via step through, step down, and shuttle box experiments).[69] Leaf extracts (water) have shown improvements in adult rat cognitive elsewhere.[77]

    Asiatic acid in isolation has shown cognitive enhancing properties, albeit at an oral dose of 30mg/kg in rats (1-10mg/kg inactive) as assessed by active and passive avoidance tasks.[78]

    The centella asiatica extract does appear to have inhernet cognitive enhancing properties, and while isolated asiaticoside is implicated in cognitive enhancement it seems that the water extract (lacking saponins) is also effective; suggesting that there are multiple bioactive components

    5.

    Cardiovascular Health

    5.1

    Cardiac Tissue

    Madecassoside is known to have antiinflammatory effects in immune cells[79] and this appears to extend to cardiomyocytes at a concentration of 100-300µM when preloaded before LPS by 2-4 hours.[32] This protective effect in cardiomyocytes was associated with activation of ERK and p38 (but not JNK) resulting in inhibition of NF-kB.[32]

    The antiinflammatory properties of madecassoside appear to extend to cardiomyocytes

    Oral ingestion of 20mg/kg madecassoside to rats for five days with the final dose two hours prior to LPS infusion is able to attenuate the changes in blood pressure and heart rate.[32] 2-50mg/kg madecassoside prior to ischemia/reperfusion dose-dependently reduced infarct size, with the higher two doses (10-50mg/kg) being comparable to the reference drug of 10mg/kg nifedipine.[80] This study also noted less potency in normalizing markers of cell damage (LDH and CK) yet more potency in normalizing antioxidant biomarkers (SOD and MDA).[80]

    Madecassoside appears to be active when orally ingested in rodents, and can protect from cardiac stressors when preloaded

    5.2

    Blood Pressure

    Acute administration of 30mg/kg asiatic acid to rats (but not 1-10mg/kg) is able to slightly and acutely reduce systolic and diastolic blood presure.[78]

    5.3

    Clotting

    A slight inhibitory effect on platelet reactivity is noted with oral ingestion of 3,5-di-O-caffeoyl quinic acid at 4mg/kg bodyweight twice daily over 14 days in rats;[3] flavonoids of centella asiatica and asiaticoside are ineffective.[3]

    5.4

    Endothelium

    30-300µM madecassoside (but not 10µM) is able to attenuate the damage done by H2O2 attenuated the changes in glutathione and MDA in a concentration dependent manner, but only up to around half restoration;[33] This was associated with inhibition of MAPK phosphorylation (although ERK was unaffected)[33] which is known to be an upstream regulator of caspase-3[81] and its reduction may explain the increased mitochondrial integrity with madecassoside.[33]

    5.5

    Circulation

    One trial has assessed the effects of centella asiatica on normal controls, persons with moderate superficial venous hypertension, and those with postphlebitic limbs and severe venous hypertension.[82] 60mg centella asiatica thrice daily was able to benefit microcirculation in all subjects albeit it was significantly more effective in those with more severe symptoms.[82]

    In diabetics with microangiopathy, 60mg of the saponins from centella asiatica twice daily over the course of six months, there was a significant improvement in microcirculation relative to both control and placebo.[83] This same dose for 12 months in diabetics with both microangiopathy and diabetic neuropathy also noted benefits to the degree of 38% (improved venous response) associated with a reduction in edema in extremities approximately 28% less than placebo control;[84] those without neuropathy also benefitted from 12 months of treatment, but to a lesser degree of 22.7% for venous responsiveness and 9.5% for edema.[84]

    In subjects with mild to moderate superficial venous disease given triterpenoids from centella asiatica before a three hour flight (60mg of saponins thrice daily for the two days preceding the flight), supplementation was associated with significantly less ankle and leg swelling (relative to placebo).[85]

    5.6

    Chronic Venous Insufficiency

    Chronic venous insufficieny is a circulatory disorder that is caused by the obstruction or reflux of blood flow in the veins due to abnormalities of the venous wall and valves,[86] and tends to result in varicose veins (aesthetic) and venous ulcers (medically important) as a side-effect of high venous blood pressure[87] resulting in reduced quality of life.[88] Centella asiatica has been traditionally used for circulatory disorders, and similar to some other supplements (Pycnogenol and Horse Chestnut) it has been investigated for its therapeutic usage in CVI.[89]

    Mechanistically, centella asiatica and its saponins are known to inhibit elastase and hyaluronidase[90] with a potency greater than horse chestnut.[91] Hyaluronidase is known to degrade hyaluronic acid (component of capillaries) and elastase degrades numerous targets in the extracellular matrix (elastin, collagen, proteoglycans, fibronectin) and it is thought their overactivity can augment plasma leakage through the endothelial wall (which results in edema).[91][92]

    Centella asiatica is known to be used to treat chronic venous insufficiency, a circulatory disorder primarily associated with the veins.

    One review[89] has noted eight trials that investgiated the role of centella asiatica in chronic venous insufficient, three of which involved venous insufficient[93][94][95] and five of which involved venous hypertension.[96][97][98][99][100] While one study used 60mg of centella asiatica total saponins once daily only[93] and another studies used 30mg twice[96] or thrice[99] daily, most studies have used a twice daily approach at 60mg[100][95][98][97][96] yet all studies note efficacy over placebo.

    Relatively low oral doses of the saponins from centella asiatica appear to be reliable in the treatment of chronic venous insufficiency and in improving microcirculation; other symptoms of chronic venous insufficiency, such as leg pain/edema as well as varicose veins, appear to be reduced alongisde other symptoms

    6.

    Interactions with Glucose Metabolism

    6.1

    Mechanisms

    5-20mg/kg oral intake of asiatic acid over 45 days in streptozotocin-induced diabetic rats was able to dose-dependently reverse the changes in blood glucose and insulin, although the highest dose was less potent than the refernece drug of 600μg/kg glibenclamide[101] aside from restoring HbA1c and enzymatic function (Hexokinase, G6P, and F-1,6-BP) where they were equal.[101]

    Non-diabetic rats given 20mg/kg asiatic acid do not experience any changes in glucose or insulin concentrations.[101]

    6.2

    Diabetes

    In diabetic patients with wounds, thrice daily supplementation of a centella asiatica supplement containing 100mg asiaticoside for 21 days seemed to be associated with a significantly faster rate of wound contraction yet suppression on the rate of granulation.[102]

    7.

    Inflammation and Immunology

    7.1

    Macrophages

    Asiatic acid and asisaticoside at 30-120µM were both able to suppress LPS-induced inflammation in RAW 264.7 macrophages as assessed by cytokine secretion as well as both nitrite and PGE2 production, these were due to inhibition of NF-kB;[34] the inhibition of NF-kB was secondary to inhibiting phosphorylation of p38, ERK1/2, and JNK (MAPKs) and this inhibition was due to preventing phosphorylation of Raf-1 (60-120µM abolishing phosphorylation)[34] which is known to stimulate the MAPKs.[38][39]

    Madecassoside has also been confirmed to inhibit NF-kB activation in macrophages[79] although elsewhere 10-100µM has failed to reduce nitric oxide production in LPS activated macrophages.[103]

    The triterpenoids appear to have antiinflammatory effects at moderate to higher concentrations, and this is associated with suppression of NF-kB activation

    Asiaticoside, in vitro with THP-1 macrophages, appears to augment the MCP-1 induced secretion of IL-1β at concentrations of 10pM to 100nM without inherently having an effect in the absence of MCP-1; the increase in secretion was around 30%.[45]

    Asiaticoside appears to augment a macrophage dependent release of IL-1β, and this occurs at remarkably low concentrations

    7.2

    Fever

    In fevers, macrophages activation and induction of the COX enzymes will produce PGE2. This prostaglandin binds to its receptors in the brain to suppress heat losses and augment heat retention[104][105] causing fever symptoms. Due to the traditional usage of centella asiatica as an antipyretic and its ability to suppress COX expression elsewhere, it has been investigated in fevers.

    Three days supplementation of isolated asiaticoside (5-45mg/kg) to rats the subject to lipopolysaccharide (LPS) infusion was able to attenuate the rise in body heat seen with LPS induced fever, and pretreatment of 45mg/kg asiaticoside was as potent as an infusion of 50mg/kg paracetamol given once after LPS.[106] These anti-fever effects are associated with increases in serum IL-10 (which is known to reduce fevers via inducing heme-oxygenase 1 or HO-1[107][108]) and HO-1 was confirmed to be increased with asiaticoside; blocking HO-1 prevents the antipyretic effects of asiaticoside.[109]

    May have anti-pyretic (fever reducing) properties secondary to causing an increase in circulating IL-10, which then causes an increase in HO-1 activity

    7.3

    Virology

    Centella asiatica (methanolic extract) appears to have anti-viral properties against herpes simplex, with inhibitory properties against HSV-1 (EC50 of 362.40µg/mL) and HSV-2 (EC50 of 298.84µg/mL), a potency significantly lesser than the reference drug (Acyclovir 0.14-0.15µg/mL) and the herbs Maclura cochinchinensis and Magnifera indica (both around 20µg/mL).[110] This moderate level of potency has been replicated elsewhere.[111]

    This may be related to the asiaticoside content.[110]

    Technically there are anti-herpes simplex properties, hindering viral replication, but the potency seems to be significantly lesser than other herbs and the reference drugs. May still be useful for topical application, but the concentrations may be too high for oral ingestion

    7.4

    Rheumatoid Arthritis

    In collagen-induced arthritis in mice, oral ingestion of 10-40mg/kg for 20 days was able to dose-dependently reduce inflammation, and this appears to be associated with reductions in anti-CII antibodies (21.1-62% reductions at 20-40mg/kg) and reduced immune cell reactivity to said antibodies.[103] This has been replicated elsewhere with 10-30mg/kg madecassoside, where it was noted that circulating inflammatory cytokines were reduced and the antiinflammatory IL-10 upregulated alongside improvements in arthritis symptoms[112] and the potency of madecassoside (10-30mg/kg) appears comparable to indomethacin (10mg/kg)[112] yet lesser than dexamethasone (1mg/kg).[103]

    8.

    Interactions with Hormones

    8.1

    Testosterone

    In rats who consumed centella asiatica at a dose that impairs fertility (10-100mg/kg aqueous extract), it appears that testosterone is reduced in a dose dependent manner.[113]

    9.

    Interactions with Organ Systems

    9.1

    Stomach

    Oral ingestion of a centella asiatica extract (hot water extract) at 50-500mg/kg bodyweight prior to alcohol ingestion if able to reduce ulcer formation by 58-82%[114] and a subsequent test using 200-600mg/kg of the leaf juice (twice daily for five days) was able to protect against all tested stressors (aspirin, ethanol, stress, and pyloric ligation) with a potency comparable to 250mg/kg sucralfate.[115]

    Appears to have somewhat respectable anti-ulcer properties in the stomach

    9.2

    Kidney

    Centella asiatica appears to be used in traditional chinese medicine for the purposes of being a kidney tonic and promoting diuresis.[116]

    In rats subject to adriamycin-induced renal toxicity (a model for nephrotic syndrome), oral 8-32mg/kg asiaticoside for four weeks was able to reduce protein losses in the urine with 16-32mg/kg being as potent as 25mg/kg prednisone;[116] these protective effects were thought to be related to the preservation in integrity of the podocytes (last barrier against protein losses in the urine[117]).[116]

    Asiaticoside appears to be protective of the kidneys against animal models of nephrotic syndrome

    9.3

    Testicles

    Water extracts of centella asiatica (200mg/kg) were able to nearly normalize adverse changes in lead-induced impairments in sperm cell morphology, motility, and count[118] without inherently affecting sperm function or enzymatic (3β-HSD and 17β-HSD) activity.[118]

    It has been noted that centella asiatica extracts (aqueous extract of dry leaves) at 10-100mg/kg to rats have increased testicular weight yet caused abnormal morphology of sperm cells[119] thought to be related to the glycosides isothankuniside and thankuniside.[1] A later study noted a decrease in testicular weight alongside dose-dependent decreases in sperm cell viabiltiy, motility, and count which were associated with increased apoptosis in sperm cells.[113] It is unclear why the results from this research group differ from the other ones.

    10.

    Interactions with Aesthetics

    10.1

    Skin

    30μg/mL asiaticoside has been noted to induce genes associated with skin cell proliferation and collagen synthesis[35] and to promote skin cell migration, attachment, and growth in vitro[120] while collagen type I and III mRNA has been noted to be increased at 1-10μM concentrations of both asiaticoside and madecassoside (the latter being more potent).[121]

    In keloid fibroblasts (keloids refer to excessive skin proliferation and scar formation, and tend to be exacerbated by TGF-β1 since it plays a role in tissue regeneration[122] but is dysregulated in prolonged and impaired wound healing resulting in scar formation[123]), asiaticoside at 100-500μg/mL inhibits keloid proliferation in a concentration and time dependent manner over five days of incubation (without affecting normal fibroblasts, reducing keloids to 35-63% of baseline) associated with increasing Smad7 expression and downregulating TGF-β receptors which suppressed collagen synthesis.[124]

    When looking at what happens to skin cells, they appear to experience an increase in collagen synthesis and enhanced growth and proliferation when they are subject to centella asiatica glycosides (asiaticoside and madecassoside); the opposite seems to occur in scar tissue

    When looking at the enzymes of elastase and hyaluronidase asiaticoside shows inhibitory potential against their activity with IC50 values of 19.45+/-0.25µg/mL (elastase) and 18.63+/-0.33µg/mL, a potency comparable to ursolic acid[90] and greater than other medicinal plants such as clitoria ternatea[125] and horse chestnut.[91]

    Madecassoside showed an increase in TIMP-1 mRNA (which normally sequesters MMP-1[126] which would preserve collagen, this also being seen with asiaticoside[35]) but the ratio of TIMP-1:MMP-1 was unaltered suggesting no change in collagen breakdown.[121] Elsewhere, asiaticoside has been shown to reduce MMP-1 activity in vitro[90] with a potency comparable to oleanolic acid.

    Both madecassoside (3-10μM) and asiaticoside (10μM) also stimulated TGF-β1 and TβRII mRNA and Smad3 protein levels while downregulating Smad7; the aglycones were inactive[121] and this stimulation of collagen synthesis and TGF-β1 mRNA is independent of the TGF-β1 receptor.[127]

    There appears to be a beneficial trend in enzymes and proteins involved in skin synthesis, with a downregulation of MMP-1, elastase, and hyaluronidase activiy with an increase in transforming growth factor mRNA levels

    There appears to be an increase in macrophage (but not leucocyte) concentration as well as some cytokines (MCP-1, VEGF, IL-1β) in the wound exudate of rats topically treated with asiaticoside, and macrophages in vitro were the cause of the increased IL-1β when in the presence of MCP-1 (but not inherently).[45]

    There appears to be increased macrophage recruitment to wounds associated with asiaticoside

    Elastase and hyaluronidase are both involved in loss of skin elasticity with sun exposure[128] and MMP-1 is involved in wrinkle formation;[129] due to the inhibitory potential of asiatocisde against all three aforementioned proteins, it is investigated as a topical agent against skin aging.[90]

    Elsewhere, centella asiatica appears to suppress pro-apoptotic changes in mRNA that occurs following UVB exposure.[130][131]

    Centella asiatica may possess anti-aging properties on the skin when topically applied, secondary to reducing the damaging effects of UV radiation. While not tested in a living model yet, it appears more potent than other supplements at this goal

    10.2

    Wound Healing

    Centella asiatica appears to be a commonly ingested or topically applied herb for the purpose of wound healing, similar to aloe vera.[5] Of the phases of wound healing (reducing bacterial contamination, reducing inflammation, and promoting resynthesis) centella asiatica appears to be recommended for the specific purpose of promoting resynthesis of tissue.[5]

    Appears to be traditionally used both topically (applied to the skin) and internally (consumed) for the purpose of wound regeneration

    When looking at burn healing rates, rats subject to burns and then fed either the glycosides (6-24mg/kg) or the aglycones (3-12mg/kg) noted accelerated burn healing rates with madecassoside relative to asiaticoside (although both were effective) while the aglycones were ineffective.[121] Elsewhere, isolated madecassoside (6-24mg/kg) in burned rats appears to cause dose and time dependent benefits to wound closure associated with increased angiogenesis[132]

    Topically applied centella asiatica herb extract (0.002-0.005%) or lower concentrations of isolated asiaticoside (10pg to 100ng per wound in rats) is able to reduce wound size from a burn over 20 days of observation.[45]

    Appears to have the ability to enhance burn healing rates when orally ingested as well as topically applied

    In a punch wound model, 0.2% topical asiaticoside is able to increase wound tensile strength (57%) and collagen content when 0.1% or lower was ineffective and in diabetic rats (wound healing rate is attenuated[133]) a higher concentration of 0.4% was required to reduce wound size by 42% and increase tensile strength by 66%.[134]

    Oral intake of 1mg/kg asiaticoside was found to accelerate wound healing by reducing the wound area by 28% and increased collagen content by 76%, but half this dose and 10-fold the dose were both ineffective.[134]

    Appears to possess wound healing properties when tested in non-burn models (physical trauma)

    10.3

    Stretch Marks

    Centella asiatica has been used alongside other nutraceuticals (Vitamin E,[135] rosehip oil,[136] and hydroxyprolisilane CN[136] which is hydroxyproline and silicium) for the treatment of striae gravidarum (stretch marks), which is a condition caused by either suppression of fibroblast proliferation (glucocorticoid usage) or mechanical stretching that exceeds the rate of firboblast proliferation.

    While the combination therapy appears highly effective,[135][136] there are currently no studies asssessing the efficacy of centella asiatica in isolation.

    May have a role in reducing stretch marks, although this is not yet ascertained

    11.

    Interactions with Cancer Metabolism

    11.1

    Breast Cancer

    In MCF-7 and MDA-MB-231 breast cancer cells, asiatic acid is able to induce S-G2/M phase arrest and induce apoptosis with IC50 values of 5.95μM and 8.12μM, respectively[137] and 10μM of asiatic acid can induce 95% apoptosis over 48 hours in vitro.[138] This was due to inhibition of two MAPKs (p38 and ERK) causing an increase in p21/WAF1 independent of p53 (as asiatic acid did not influence p53) and also independent of JNK and Fas/Fas Ligand.[137] The overall protein content of p38 and ERK are not influenced, just their phosphorylation status.[137]

    The aforementioned apoptosis is a mitochondrial-dependent pathway, and while p38 was required for the phase arrest both p38 and ERK played a role in mitochondrial-dependent apoptosis[137] ultimately causing DNA fragmentation in these cancer cells.[138]

    Asiatic acid signals via p38 to cause cell cycle arrest in breast cancer cells, and via both p38 and ERK to cause cellular apoptosis; these occur at a concentration probably relevant to oral supplementation

    11.2

    Lung Cancer

    Asiatic acid appears to have antiproliferative effects on A549 lung carcinoma cells with an IC50 of 44.95+/-5.14µM and on PC9/G cells with an IC50 of 58.04+/-2.90µM.[139]

    Possible antiproliferative effects at higher than normal concentrations of asiatic acid

    11.3

    Melanoma

    SK-MEL-2 cells appear to experience apoptosis with asiatic acid with an IC50 of around 40µM (despite a clear drop to less than 20% viability at 50µM) which was associated with an increase in oxidative damage from the mitochondrial pathway yet no alterations in p53.[140]

    Appears to induce apoptosis, possibly similar to the mechanisms seen in breast cancer cells due to no p53 involvement

    11.4

    Liver Cancer

    In isolated HepG2 cells (hepatoma), asiatic acid is able to cause intracellular calcium influx at around an IC50 of 30µM yet reaches 20% cell viability at 40µM.[141] This increase in calcium influx is known to mediate apoptosis[142] via activating p53[143] and increased p53 was noted to be secondary to calcium influx in these cells.[141] Elsewhere, asiatic acid has noted a similar IC50 for apoptosis at 45μM in HepG2 cells.[30]

    As evidenced in prostatic cancer cells, this increased calcium influx may be from disrupting the endoplasmic reticulum (an intracellular storage of calcium).[144]

    Appears to cause calcium influx at moderate to higher concentrations, causing an increase in p53

    11.5

    Prostate

    PPC-1 prostatic cancer cells incubated with asiatic acid is able to induce cell death with an LD50 of 42+/-3.5µM peaking at eight hours of incubation. This was associated with the mitochondrial dependent pathway (increasing levels of caspase 2, 3, and 8 but not 9)[144] but there appears to also be a mitochondrial independent pathway causing apoptosis which involved calcium release from physically disrupting the endoplasmic reticulum.[144]

    Appears to induce apoptosis in prostatic cells via two independent mechanisms

    12.

    Nutrient-Nutrient Interactions

    12.1

    Vitamin E

    While both the ethanolic extract of centella asiatica and Vitamin E show concentration-dependent antioxidant properties in vitro, the combination was highly synergistic as the theoretically additive IC50 values (gained by adding the individual IC50 of vitamin E and centella asiatica and assuming they are complementary) was reduced from 594-15,394ng/mL (depending on extraction process) down to 14.9-584.2ng/mL (2-3% of the theoretical value).[145] This was thought to be due to centella asiatica recycling vitamin E in the same manner that Vitamin C does.[145]

    On in vitro antioxidant tests, centella asiatica appears to be highly synergistic with Vitamin E

    13.

    Safety and Toxicology

    13.1

    General

    An acute oral dose of 10g/kg of the centella asiatica extract ECa-233 (80% saponins) failed to exert acute toxicity to mice when observed over 14 days, and in a subchronic study doses of up to 1g/kg daily over the course of 90 days was not associated with any clinically relevant toxicological symptoms.[146]

    13.2

    Case Studies

    There is a case study of a 15 year old girl on lymecycline (acne medication) for eight weeks who took an undisclosed herbal medication for her acne of which one ingredient was centella asiatica (dosage and other ingredients not disclosed); that being said, injury occurred upon rechallenge suggesting the product in question was causative.[147] While the acne mediciation belongs to a class of molecules (tetracyclines) that may induce liver damage, these tend to occur within 4-5 days of ingestion.[148]

    Elsewhere, three women have had reactions to centella asiatica itself (jaundice alongside liver inflammation and apoptosis/necrosis) and one women had a reaction to reintroduction of the same product; all subjects were successfully treated with ursodeoxycholic acid.[149]

    There are a few case studies that pinpoint causation on the products containing centella asiatica as symptoms reoccur with reintroduction, this isn't fully causative to say the herb causes liver damage but reasonable to suspect it and it isn't ascertained what made these particular women susceptable

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