Cissus Quadrangularis

Cissus quadrangularis (of the family vitaceae) is a joint and bone health herb (known to 'accelerate the rate of bone healing'^[1]) from Ayurveda under the name of Asthi Shrinkhala (Sanskrit)^[2] with is also used for treatment of digestive, asthmatic, and menstrual disorders as well as the health of the eyes and ears^[3][1] and less frequently for muscle pain (both smooth and contractile).^[4][5] The plant is known to grow indigenously in India, Malaysia, Sri Lanka, Thailand, and Africa where it is used medicinally and bears other names such as hadjod (Hindi) and 'Bone Setter' in reference to its bone healing properties.^[6]

Cissus quadrangularis is a traditional medicine for female health and for the health/function of both bones and joints, with some minor uses in digestive health

Cissus quadrangularis (aerial parts unless otherwise specified) tend to contain:

• 6′-O-trans-cinnamoyl-catalpol, 6-O-{2,3-dimethoxy}-t-cinnamoyl-catalpol, and 6-O-m-methoxy-benzoyl catalpol (acetylated glycosides of catalpol)^[7][8]
• Friedelin (pentacyclic triterpenoid), possibly up to 2.5% in 70% ethanolic extracts of the aerial parts (IND-HE)^[9][6]
• Two isomeric ketosteroids, onocer-7-ene-3α,21β-diol and onocer-7-ene-3β,21α-diol^[10] and some others including 7-oxoonocer-8-ene-3β,21α-diol^[10]
• δ-amyrin and δ-amyrone^[11][10]
• 3,3',4,4'-tetrahydroxybiphenyl^[10]
• Quercetin (0.007634-0.042649%^[12]) and Kaempferol^[11]
• Stilbene compounds such as Resveratrol (0.000264-0.000676%^[12]) or piceatannol,^[13] resveratrol glycosides,^[11] and other stilbenes such as pallidol, parthenocissine A, and quadrangularin A-C (both resveratrol dimers)^[7][13] genistein and daidzein^[7]
• Vitamin C at around 327mg per 100g^[14]
• Vitamin E at around 696mg per 100g^[14]
• β-sitosterol (1.15% methanolic extract, 0.5% ethyl acetate extracts, undetectable in water^[15]) and stimasterol (0.47%, 0.16%, and undetectable respectively^[15])

The main molecules of concern here seem to be the triterpenoids, where the very unique looking 'asymmetrical tetracyclic triterpenoids' (usually called ketosteroids) and freidelin seem active. Other bioactives may include the stilebenes such as resveratrol (and its unique dimers, such as Quadrangularin A) and the molecules based off of catalpol

Cissus Quadrangulus extract (stem) has an anti-oxidative phenolic capacity of 585.40+/-0.16mg gallic acid equivalents (GAE) per 100g^[14] which are thought to underlie some of the observed antioxidant effects^[14][16] and also contains a 16.2mg/kg alkaloid content (expressed by dry weight of the plant).^[17] Flavonoid content is 169.2+/-1.97mg QE (Quercetin equivalent) per 100g.^[17]

Cissus species of plants possess a gum (oleoresin) similar to some other nutritional herbs such as irvingia gabonensis or khaya grandifoliola,^[18] and these gums normally serve roles in either rapidly absorbing water in solution (demonstrated with khaya gum^[19]) or causing coingested drugs to have a time-release; this is thought to be the reason it has been used with the intent of an appetite suppressant.^[20][21]

The plant has a general antioxidant capacity (not overly impressive relative to some other herbs or pure antioxidants)

The whole plant of cissus quadrangularis (methanolic extract) appears to have relatively low binding affinity to the GABA_A benzodiazepine site in the concentration range of 0.1-10mg/mL.^[22]

While there are some GABAergic properties thought to occur due to the enhancement of sleep time induced by diazepam (see the sedation section), there is currently a lack of information on what bioactives are working and how they are working

When tested in a model of diazepam induced sleep, cissus quadrangularis (250-500mg/kg of the water extract) in mice appears to reduce sleep latency (by 47%) and prolong time asleep (a 10-fold increase from 21+/-8 minutes to 215+/-38 minutes) relative to the diazepam control at the higher dose only.^[4]

There appears to be an enhancement of benzodiazepine induced sleep time seen with cissus quadrangularis oral intake in high but reasonable doses, suggesting an enhancement of GABA_A signalling

250-500mg/kg of the water extract fed to mice an hour before seizure induction (Isonicotinic Hydrazide Acid (IHA) or electrical shock) was reported to reduce convulsions by 60-75% (electric shock) and delay seizures by 36-77% (IHA), both of which were significantly protective but less than the reference drug of 10mg/kg diazepam (83% reduction and 117% delay, respectively).^[4]

There appears to be respectable anticonvulsive properties associated with this plant based on preliminary animal evidence

The early phase of the formalin test (analgesic test) has shown benefit from cissus quadrangularis (10-40mg/kg of the methanolic extract; 11.82% yield) by inhibiting 22-47% of the licking response, outperforming the reference drug of aspirin (300mg/kg; 28%) despite aspirin being significantly more effective in the late phase (96% relative to 33-68%).^[23] There is also analgesia in the acetic-acid writhing test, with 10-40mg/kg exerting 34-72% inhibition while aspirin reached 66%^[23] and the water extract (250-500mg/kg; orally) has shown efficacy in a hot plate test with similar potency over 90 minutes.^[4]

In animal testing, cissus quadrangularis appears to be effective in reducing pain at oral doses low enough to be feasible with oral supplementation in humans

These analgesic effects have been noted in exercise trained men reporting (nonpathological) joint pain associated with exercise, where supplementaiton of 3,200mg of a cissus quadrangularis supplement reduced joint pain as assessed by the WOMAC rating scale (no placebo control nor reference drug for comparison).^[24]

Preliminary human evidence supports a pain killing effect in athletic persons associated with oral intake of the extract

In isolated endothelial cells (ECV304) cissus quadrangularis appears to be able to reduce H₂O₂ mediated oxidative damage with an IC₅₀ of 7.49+/-0.20mg/mL and secondary to this protective effect there was an increase in antioxidant enzymes and eNOS activity;^[12] this was thought to be due to the low resveratrol and quercetin content, as both molecules were significantly more protective.^[12]

Some possible, but not overly potent nor relevant, protective effects on the endothelium thought to be mostly due to two other dietary supplements which are minor components of cissus

Supplementation of cissus quadrangularis methanolic extract at 10% of an obesogenic diet over 60 days appears to be able to attenuate the increases in blood glucose and insulin, as well as improve insulin sensitivity, with a comparable potency to Metformin;^[14] there was no signifiant influence of cissus treatment to normal rats, and the dose may be too impractically high to be seen with standard oral supplementation of cissus in humans.

At this moment in time, there is no significant nor relevant antidiabetic properties demonstrated with standard doses of the supplement and higher oral doses seem to confer some protective effects in overfed rats (common to many supplements)

In a sample of obese persons (n=72), 300mg Cissus daily standardized to 2.5% ketosteroids was able to reduce body fat levels from 33.07+/-10.26% to 30.81+/-5.92% by 4 weeks and 28.23+/-6.12% by 10 weeks, in which only 10 weeks was significantly different than placebo.^[20] This dose was able to reduce body weight by 8lbs over 10 weeks, and was slightly more effective when paired with Irvingia Gabonensis.^[20]

Another study utilizing Cissus (standardized to 2.5% ketosteroids and 15% fiber) but with other confounds in the same capsules (green tea catechins and B-vitamins) found that obese persons had a weight reduction of 6.9% over 8 weeks and overweight persons 4.8% over the same time period; pairing a 2200kcal diet with the supplement in obese persons resulted in 8.5% body weight reduction.^[21] Obese persons lost 6% of their body fat and overweight persons 4.7% without diet.^[21] Another study using this same formulation found similar results, although did not have a group taking only Cissus without a dietary intervention.^[25]

At this moment in time, most studies on cissus quadrangularis and fat reduction are conducted in obese persons in which a reduction in food intake (and thus weight loss due to less food intake) cannot be ruled out. Due to this, the potential influence of study financing, and no plausible mechanism for cissus to reduce fat supplementation of cissus for the purpose of fat loss does not seem overly promising

250-500mg/kg of the water extract appears to have muscle relaxant properties within 30 minutes of oral ingestion in mice as assessed by a rotarod test, with the higher dose being nonsignificantly less impairing than 5mg/kg diazepam.^[4]

High doses of the water in mice (correlating to around 20-40mg/kg in humans, a feasible dose) appear to have muscle relaxing properties alongside the sedative propertis which can act 30 minutes after oral ingestion; while not shown to work in humans yet, it may be prudent to not take cissus as a pre-workout supplement due to this

In SaOS-2 osteoblasts, cissus quadrangularis (water extract of the aerial parts) at 1-10μg/mL was able to concentration-dependnetly increase the basal secretion of IGF-1 (mRNA increased by 42.6-69.26% and protein levels by 38.4-84.6%), IGF-II (mRNA by 35.51-77.95% and protein by 71.43-104%), and there was also an increase the expression of the IGF receptor with the mRNA being increased by 50.66-62.66% whereas the actual receptor content was increased by 27.39-67.8%;^[26] while there was no increase in the mRNA levels of the IGF binding protein which suppresses IGF activity (IGFBP-3), there was an increase in the protein content by 28.47-52.89% in the same concentration range.^[26]

These effects may be related to either the estrogenic properties of cissus quadrangularis (as estrogen itself can increase IGF secretion from osteoblasts^[27]) but this has not yet been confirmed.

There appears to be a concentration dependent increase in the secretion of insulin-like growth factors in osteoblasts at a concentration range which seems feasible to occur following oral ingestion

Cissus quadrangularis (ethanolic extract of aerial parts) at a range of 0.1-100µg/mL noted that there was no time-dependent increase in the proliferation of SaOS-2 osteoblasts at 0.1µg/mL (100nM) where proliferation was increased by 14-21%, but at 10µg/mL there was a time-dependent increase reaching 68-80% peaking after 48 hours.^[28]

Differentiation of osteoblasts also occurs in the active range due to an increase in alkaline phosphatase (ALP) secretion at 1-10µg/mL or the water extract (53-105% increase in mRNA^[28]) or 100-300µg/mL of the petroleum ether extract^[29] alongside an increase in the mRNA of RunX2 (66-118%) as well as its transcriptional activity with the water extract;^[28] the increase in ALP activity seems due to MAPK activation, mostly p38.^[30] And both mineral nodule formation and mineralization have been noted to be increased under the influence of cissus quadrangularis.^[28][30]

Elsewhere, the differentiation of mesenchymal stem cells into osteoblasts also appears to be enhanced with incubation of cissus (100-300µg/mL of the petroleum ether extract) in a manner that is additive with estrogens.^[29]

The proliferation of osteoblasts may be related to 6′-O-trans-cinnamoyl-catalpol which has been noted to have osteogenic activity by itself, which was active in the range of 1-1,000pM and similarly effective to rutin from allophylus serratus and a few phenolics in vitex negundo similar in structure to the one in cissus.^[8]

It seems that the extracts from cissus quadrangularis can promote osteoblastic proliferation and differentiation, and may influence the promotion of mesenchymal stem cells into osteoblasts

Collagen synthesis (mRNA levels) appears to be increased with 1-10µg/mL cissus quadrangularis in SaOS-2 (osteoblast) cells, to the degree of 85-106% over baseline.^[28]

Collagen synthesis has been noted to be increased in osteoblasts; significance to joints unknown

A study in exercise trained men who experienced chronic joint pain due to said exercise given 3,200mg of cissus quadrangularis daily for eight weeks noted that supplementation noted that supplementation was associated with a 31% reduction in total WOMAC score (self-reported joint pain and impairment) relative to baseline.^[24]

Preliminary evidence in athletic men with joint injuries arising from exercise appears to support a role for cissus quadrangularis in reducing joint pain and improving mobility

Cissus quadrangularis is famous in traditional medicine for healing bones, insofar that it is referred to as the 'bone setter'^[6] and is commonly used for this purpose according to surveys of traditional medicine usage (Ghana);^[31] while there has been a large amount of research in animal models that is seen as not relevant to humans (due to placing cissus quadrangularis directly into bone tissue during surgeries or injecting it^{[32][33][34][35][36]}) there does appear to be one (preliminary) study in humans where in persons with mandibular fractures noted benefit in regards to reducing pain after a week of supplementation, but beyond that week and in all other parameters (swelling, tenderness, mobility) it was not significantly better than placebo over the course of six weeks.^[37]

It is thought that the improvement in fractures is in part due to suppressing corticosteroid signalling (by acting as a receptor of the glucocorticoid receptor) and preserving anabolism of bone tissue, while also directly promoting osteoblastic proliferation and differentation;^[33][32] these claims have not yet been fully proven in oral studies.

The bone healing properties of cissus quadrangularis are one of its most popular traditional claims, but at this moment in time there does not appear to be good human evidence to support the usage of cissus for this goal

Oral intake of cissus quadrangularis (petroleum ether extract of the stem; 0.07% yield) in a rat model mimicking menopause (ovarectomized rats) at a dose of 500mg/kg for ninety days is able to fully prevent losses of bone strength and prevent up to 86% of the losses in bone thickness;^[38] cissus quadrangularis showed comparable efficacy to 5.4mg/kg of the SERM known as raloxifene, and while this study is duplciated in Medline^[39] the design has been replicated where a month of cissus quadrangularis supplementation (500mg/kg petroleum ether extract) was comparable in efficacy 25mg/kg raloxifene in ovarectomized rats.^[40]

In mice, 500mg/kg cissus quadrangularis has been noted to reverse the ovarectomy induced increase in inflammatory cytokines (TNF-α) and augment the increase in IL-1β from 39% to 322%;^[41] these changes were associated with near absolute preservation of corticol and cancellous bone mineral density and thickness.^[41]

The aerial parts (stem and leaves) of this plant have been noted elsewhere to be effective, where 75-100mg/kg of a phytoestrogen rich extract (70% ethanolic with 2.5% friedelin) in ovarectomized rats was able to increased circulating estrogen and Vitamin D in serum while the higher dose was comparable to estrogen in promoting bone strength and thickness but not density.^[9]

In rat studies, oral ingestion of various extracts of cissus quadrangularis appear to exert near maximal protection against the losses in bone strength seen with ovarectomy, although the one study measuring bone mineral density failed to find a significant protective effect of comparable potency

In ovarectomized rats (a model of estrogen deficiency), oral supplementation of a 70% ethanolic extract (2.5% friedelin content) has been noted to increase estrogen relative to the ovarectomized control by 194% (75mg/kg) and 232% (100mg/kg) which were statistically significant but less than the reference drug of 17β-estradiol injections (568%);^[9] despite the difference, both were comparable in preserving bone integrity.^[9] This study is duplicated in Pubmed (due to the exact same magnitude of serum estrogen increases), where estrogenic effects were confirmed by vaginal histology and uterine weight.^[6]

A 70% ethanolic extract of this plant appears to possess estrogenic properties related more to increasing serum estrogen rather than directly acting on estrogen receptors; practical significance in male rodents and humans not yet known

Compounds in Cissus Quadrangularis have been shown to act as glucocorticoid antagonists when placed in bone tissue (potency as assessed by IC₅₀ values not given), which would reduce their catabolic effects by occupying the receptor.^[33][32]

Potential role as a glucocorticoid antagonist (which would reduce the effects of cortisol) but this has not yet been linked to oral supplementation in any species

Due to the ability of cissus quadrangularis to act as a glucocorticoid antagonist, it has been proposed to possess anabolic/androgenic activity;^[32] no studies have yet addressed this topic.

Cissus quadrangularis ethyl acetate extract has been noted to suppress LPS induced activation of macrophages in the 50-400μg/mL range (IC₅₀ of 53.88μg/mL; full suppression of nitrite production at 400μg/mL)^[42] and the acetone is associated with inhibiting most inflammatory enzymes such as COX1 (IC₅₀ of 106.46μg/mL), COX2 (25.9μg/mL), and 5-LOX (550μg/mL).^[43] These antiinflammatory effects seem to be partly mediated by induction of heme-oxygenase 1 (HO-1) since inhibiting HO-1 attenuated the antiinflammatory effects seen with cissus ethyl acetate^[42] but since not all was prevented it suggests that the enzyme inhibitors in the acetone extract may be independently active.^[43]

The acetone extract has been further purified to an AFCQ extract, with more potent inhibitory properties on COX1 (IC₅₀ of 7μg/mL), COX2 (400ng/mL), and 5LOX (20μg/mL) and comprised of mostly tannin structures.^[43] This purified extract had an IC₅₀ value of 65μg/mL in reducing LPS induced nitrite formation,^[43] similar to the ethyl acetate extract elsewhere^[42] which is less potent on the aformentioned enzymes.

It appears that cissus quadrangularis contains antioxidant compounds that activate HO-1 which then suppresses inflammation, but also contains some currently unknown tannin-like structures that are potent COX inhibitors when tested in vitro; both of these mechanisms are thought to underlie antiinflammatory effects

The crude powder of cissus quadrangularis given to rats at the dose of 500-2,000mg/kg bodyweight one day and then again one hour before a carrageenin injections noted that the middle dose (1,000mg/kg) was able to significantly reduce the early phase swelling (indicative of histamine release) by 44%, outperforming 15mg/kg hydrocorticone.^[44]

Some very limited evidence suggests a respectable antihistamine effect, but this has not been replicated and the application of cissus as an antiallergic compound is still ambiguous (due to a lack of information on mast cells and T cells)

Cissus quadrangularis appears to have proton pump inhibitory (PPI) properties when the methanolic extract is tested in vitro of a potency (IC₅₀ in inhibiting the H⁺/K⁺ ATPase pump of 38μg/mL) similar to omeprazole (reference at 26μg/mL).^[17]

Cissus quadrangularis methanolic extract appears to have the ability to act as a proton pump inhibitor

In vitro, water extracts of cissus quadrangularis appear to have antibacterial properties against helicobacter pylori with MIC values in the range of 40μg/mL pending on when the plant said extract was derived from.^[45]

May have some protective effects against helicobacter pylori infections, practical significance of this information is not yet known

Pretreatment of cissus quadrangularis before ulceration by NSAID drugs appears to be nearly prevented with 250-500mg/kg of the methanolic extract associated with normalization of the damaged occurring to the gastric wall^[46] and reducing inflammatory and oxidative DNA damage seen in the stomach with NSAID ulceration.^[47] 1,000mg/kg of this extract for a week prior to ulceration from aspirin (NSAID) has been noted to normalize ulceration to control levels, a potency similar to 30mg/kg ranitidine (pharmaceutical H2 receptor inhibitor) by reducing ulceration 71.2-71.9%.^[48][49]

Cissus has shown accelerated healing against ulcer formation induced by acetic acid when cissus was given at 250-500mg/kg of a methanolic extract (3.2% yield) the day after ulceration and for one week afterwards, where cissus performed equally to the reference drug sucralfate;^[50] these rehabilitative effects were seen alongside an increased polyamine content, cell proliferation, and thymidine uptake in the gastric fluid of treated rats associated with a preservation of TGF-α.^[50] Polyamines themselves are known to be highly rehabilitative in regards to ulcers (reducing polyamines reduces the healing rate^[51]) and TGF-α is also a gastroprotective factor.^[52]

While the PPI properties of cissus quadrangularis are thought to play a role, the aforementioned 500mg/kg methanolic extract dosage (which is optimal, due to being more protective than 250mg/kg and 750mg/kg^[49]) appears to be associated with less neutrophil infiltration into gastric tissue.^[49][53]

While it hasn't been directly linked to the PPI properties of cissus, this herb appears to have relatively potent anti-ulcer properties when given in either a prevenative or rehabilitative manner based on preliminary animal evidence; the oral doses used in these studies seem to be higher than other studies (estimated human dose being 80-160mg/kg with the higher dose being optimal)

One study in rats has noted that cissus quadrangularus at 10% of the diet when they were fed an obesogenic diet (high fructose and high fat), supplementation over the course of 60 days was able to reduce the increase in hepatic peroxidation and liver enzymes in serum.^[14] There was no influence on liver enzymes, lipid peroxidation, or the lipid/cholesterol content of rats on a normal diet given cissus.^[14]

A lower dose has been used elsewhere, where 500mg/kg of the methanolic extract of cissus quadrangularus was able to reduce the liver damage done by rifampicin or isoniazid both with nonsignificantly less potency than 50mg/kg silymarins (from Milk thistle).^[54][55]

There appears to be a protective effect when high doses of the herb are ingested, but they may be too impractically high to be relevant

Cissus quadrangularis appears to be a (Thai) traditional medicine for hemhorroids^[23] and has been noted to be a venotropic agent when tested in isolated umbilical veins at a concentration of 100-400µg/mL which was similar in potency to 400µg/mL Daflon (a mixture of Diosmetin and Hesperidin at 9:1).^[23] Venotropic agents such as Daflon (other popular ones being Horse chestnut and pycnogenol) are known to also confer anti-hemhorroid properties, but when tested in humans supplementation of cissus quadrangularis has failed to outperform placebo despite Daflon being effective.^[56] There appears to be another study that cannot be located online reporting benefit with 500mg twice daily supplementation (reported vicariously through these two studies^[23][57]).

Should theoretically be a venotropic agent (increases blood flow in veins) and has been traditionally used as such, but the one study showing benefit cannot be located online to evaluated and the one that is available online failed to find a significant therapeutic effect

When 75-100mg/kg of the 70% ethanolic extract (2.5% friedelin) is given to a menopausal model of rats (ovarectomized) appeared to have libido promoting effects when the rats were exposed daily for over two months, with greater effects after two months relative to one; there were no immediate effects notable, and it appeared to be less significant than the reference drug of 17β-estradiol.^[6]

At least one study has noted pro-libido enhancing properties in ovarectomized rats. It is not certain if this will occur in rats not undergoing ovarectomy, since it may be due to an estrogen increase and it isn't certain if estrogen is increased in youthful female rats or humans

One study using 500mg/kg of the petroleum ether extract in pregnant rats noted that supplementation from the beginning of the second trimester until delivery was able to enhance bone mass (corticol and trabecular) in the newborn pups relative to control;^[58] this appears to have been reported previously with the ethanolic extract (750mg/kg),^[59] but while no complications were noted in either study there was no in depth toxicity testing on the pups.^[58][59]

Appears to promote bone growth in newborn pups when orally ingested by the mother starting from the second trimester, but insufficient toxicology testing has been conducted

One toxicology study found that, when using CQR-300 in rats, that a dose of 2500mg/kg bodyweight for 90 days was not associated with any observable side-effects (and established the NOAEL at this level), although some non-toxic changes occurred in blood clotting parameters at the higher doses.^[3] Previously, a rat study on dosages up to 3g/kg bodyweight over a period of 3 months did not find any abnormal effects of Cissus supplementation,^[60] the last dose (3g/kg) being 100-fold greater than the human therapeutic dose equivalent, and also noted non-toxic changes in blood parameters such as RBC count and clotting time (all within normal physiological range). From these results, it is postulated that a dose of 150g Cissus Quadrangularis daily is free from observable side-effects.^[3][60]

Elsewhere, investigating the petroleum ether extract which appears to concentration anti-osteoporotic effects has been noted to be safe up to 5,000mg/kg oral ingestion in acute toxicity studies (with a single dose not causing harm over 30 days of observation).^[38]

Two human trials using Cissus at dosages of 300mg daily (from CQR-300) found no observable side-effects^[25][21] and a pilot study of 3,200mg daily for eight weeks in exercise trained men has similarly failed to find adverse effects.^[24]

Appears to be safe at the doses commonly consumed, as toxicity has not been recorded in animal studies or traditionally via Ayurveda. The human studies using 300mg daily did not experience any differences from placebo in regards to side-effects

Cissus has been shown to produce genotoxic effects in vitro using a concentration of 500 and 1000ug/plate.^[61] Replicated these tests in revertant colonies failed to replicated the results, and in vivo tests do not show this method of damage.^[3]

Some data has noted a potential genotoxic effect in bacteria, but has failed to be replicated; practical significance of this information at this moment in time is not known but it is thought to not be relevant due to no observed genotoxicity in rodent models