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Evaluation of Mucuna pruriens extract as a potential treatment for Huntington’s disease: Antioxidant and anti-inflammatory mechanisms in rat models
*Corresponding author: Pallavi V. Bhosle, Department of Pharmacology, Swami Ramanand Teerth Marathwada University, Nanded, Maharashtra, India. pallavi1230@gmail.com
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Received: ,
Accepted: ,
How to cite this article: Bhosle PV, Wadher SJ. Evaluation of Mucuna pruriens extract as a potential treatment for Huntington’s disease: Antioxidant and anti-inflammatory mechanisms in rat models. J Neurosci Rural Pract. doi: 10.25259/JNRP_20_2025
Abstract
Objectives:
To evaluate the phytochemical profile and neuroprotective potential of Mucuna pruriens (MP) extract in Huntington’s disease (HD), focusing on its antioxidant and anti-inflammatory mechanisms using in vitro and in vivo models.
Materials and Methods:
Leaves and bark of Mucuna pruriens were sequentially extracted and tested for antioxidant activity. Huntington’s disease-like symptoms were induced in rats using 3-Nitropropionic acid, and the effects of ethanolic MP extract on behavioral, biochemical, and histopathological parameters were evaluated. Data were analyzed using Analysis of Variance (ANOVA) with P < 0.05 considered significant.
Results:
Phytochemical analysis revealed high levodopa content and targets associated with HD pathology. The extract significantly reduced reactive oxygen species, lipid peroxidation, and inflammatory mediators, demonstrating potent antioxidant and anti-inflammatory effects. In 3-NP-induced HD models, MP improved motor function, reduced oxidative stress, and preserved neuronal integrity, suggesting its potential to decelerate HD progression.
Conclusion:
MP extract exhibits promising neuroprotective effects against HD through antioxidant and anti-inflammatory pathways. Further exploration of its molecular targets and clinical applicability is warranted.
Keywords
Anti-inflammatory
Antioxidant
Huntington’s disease
Mucuna pruriens
Neuroprotection
INTRODUCTION
Background
Huntington’s disease (HD) is a progressive neurological ailment that affects roughly 5–10 individuals/100,000 worldwide, usually presenting in mid-adulthood. It is characterized by a relentless progression of cognitive decline, motor dysfunction, and psychiatric instability, significantly reducing patients’ quality of life and placing substantial emotional and physical burdens on caregivers.[1,2] The ailment arises from a genetic mutation characterized by the amplification of CAG trinucleotide repeats in the huntingtin (HTT) gene, resulting in the synthesis of a mutant HTT protein consisting of an aberrant polyglutamine sequence. This pathological protein triggers a cascade of neurotoxic events, resulting in selective neuronal degeneration, predominantly in the striatum and cortical regions.[3]
Animal models have been instrumental in advancing the understanding of HD and evaluating therapeutic approaches. The 3-nitropropionic acid (3-NP) model is frequently used, as it replicates HD-like neuropathology and behavioral deficits. By inhibiting succinate dehydrogenase, 3-NP induces mitochondrial dysfunction and energy failure, culminating in striatal neuron degeneration close to that seen in HD patients.[4,5] These models are essential for assessing the neuroprotective effects of potential treatments.
Despite extensive research, effective disease-modifying therapies for HD remain unavailable, necessitating innovative approaches. Natural products consisting of antioxidant and anti-inflammatory properties have garnered attention as potential therapeutic agents. Among them, Mucuna pruriens (MP), a medicinal legume, has shown promise due to its rich phytochemical content, including levodopa (L-DOPA), flavonoids, and alkaloids.[6] These compounds exhibit neuroprotective potential by mitigating oxidative stress and inflammation, key contributors to neurodegeneration.[7]
The present study aims to investigate the phytochemical profile and neuroprotective efficacy of MP extract in HD, focusing on its antioxidant and anti-inflammatory mechanisms. Utilizing in vitro and in vivo models, this study aims to shed light on MP’s medicinal potential for relieving HD symptoms and development.
MATERIALS AND METHODS
Authentication and collection of plant material
Fresh leaves and bark of MP were obtained from authenticated suppliers and verified by the Botanical Survey of India. The plant material was air-dried for 30 days, powdered, and stored for extraction.
Preparation of extracts
The powdered leaves and bark were first defatted using petroleum ether. Sequential extraction was then carried out using ethanol, chloroform, and finally distilled water, in increasing order of polarity. Each solvent extraction was performed using a Soxhlet apparatus for 60 h. For every solvent, a fresh batch of plant material was used to avoid overlapping phytoconstituents. All extracts were filtered, concentrated under reduced pressure, and stored at 4°C for further analysis.[8]
Organoleptic and physicochemical analysis
The organoleptic evaluation assessed the color, odor, taste, and texture of the plant parts. Physicochemical analysis determined moisture content, pH, extractive values, loss on drying, and ash content to assess purity and stability, following the World Health Organization guidelines.[9]
Preliminary phytochemical screening
Phytochemical screening was conducted on all extracts to detect bioactive compounds, including alkaloids, carbohydrates, glycosides, phenolics, proteins, fixed oils, saponins, sterols, flavonoids, steroids, and terpenoids.
Quantitative estimation
The total phenolic content was determined using the Folin–Ciocalteu procedure and reported as gallic acid equivalents per gram of extract. Using the aluminum chloride method, the total flavonoid concentration was measured and expressed as quercetin equivalents per gram of extract.
In vitro antioxidant assay
The antioxidant potential of MP extract was assessed through 2,2-diphenyl-1-picrylhydrazyl (DPPH) scavenging. Extracts (1–5 mg/mL) were incubated with 0.1 mM DPPH in ethanol, and absorbance at 517 nm was measured using ascorbic acid as a reference.[10] Hydrogen peroxide scavenging was tested with extracts (10–320 mg/mL), phosphate buffer (pH 7.4), and 40 mM H2O2, measuring absorbance at 230 nm using ascorbic acid as a control.[11] The reducing power assay used extracts (10–320 mg/mL) with ferricyanide, ferric chloride, and ascorbic acid, measuring absorbance at 700 nm to calculate EC50.[12] All assays were triplicated for reliability. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed ethanolic and chloroform extracts of MP leaves and bark maintained >75% cell viability after 24 h, confirming biocompatibility and therapeutic potential. Statistical analysis validated these findings.[13]
In vivo study
The experiment’s animals
Adult male Wistar rats (weighing 250–300 g) were housed under standard laboratory conditions (temperature: 23 ± 2°C, humidity: 55 ± 10%, light-dark cycle: 12:12 h) with access to a standard pellet diet and water. The study protocol was approved by the Institutional Animal Ethics Committee (IAEC) and adhered to the Committee for the Purpose of Control and Supervision of Experiments on Animals (CPCSEA) guidelines (animal house registration number 28762/POBC07CPCSEA). All animal experiments were conducted in accordance with the ARRIVE 2.0 guidelines.
Induction of HD-like symptoms
HD-like symptoms were induced by intraperitoneal injection of 3-NP at 10 mg/kg/day for 14 days. This neurotoxin inhibits succinate dehydrogenase, causing mitochondrial dysfunction and neuronal degeneration, mimicking HD.[14]
Acute toxicity study
Acute toxicity was assessed using Wistar rats (150–200 g) following OECD guidelines. Three groups (n = 3) received Group I – vehicle control (VC) (0.5% carboxymethyl cellulose [CMC]), Group II – 300 mg/kg of isolated phytoconstituent, and Group III – 2000 mg/kg of isolated phytoconstituent in 0.5% CMC. The rats were observed for 14 days to detect toxicity and determine the maximum tolerated dose.
Design of experiments and protocol for treatment
Animals were randomly assigned to the five experimental groups using a computer-generated random number sequence to reduce selection bias. The study included five groups of Wistar rats (n is 8/group) where Group I served as the VC with no treatment. For 2 weeks, Group II received intraperitoneal 3-NP at a dose of 10 mg/kg to induce neurological deficits. The ethanolic extract of MP (EEMP) was administered to Groups III, IV, and V at doses of 100 mg/kg, 200 mg/kg, and 400 mg/kg, respectively, alongside 3-NP, administered orally over the same period. A priori power analysis was not conducted for sample size estimation. The number of animals per group was based on previously published protocols, feasibility, and ethical considerations studies with similar experimental designs.
Body weight measurement
On the 1st and last day of the trial, body weight was measured. By comparing the end body weight to the starting body weight on the 1st day of the trial, the percent change in body weight was determined.
Behavioral assessment
The effect of behavioral parameters was assessed on the 8th, 11th, and 14th days. All assessments were conducted by investigators who were blinded to the treatment groups to minimize observer bias.
Assessment of motor activity
Motor activity and movement impairments were assessed using a combination of established methods. Locomotor activity was measured with an automated electronic activity meter (Opto-Varimex 4, Columbus Instruments, USA), which tracked horizontal and vertical movements through infrared beam interruptions along the x and y axes. Each animal’s activity was recorded individually for 2 min.[15] Behavioral impairments induced by 3-NP treatment were evaluated using a modified neurological scale, ranging from 0 (normal behavior) to 5 (recumbency due to severe limb dysfunction), based on Ludolph et al.[16] The string test, which measures forelimb strength indirectly by having the rat hang onto a steel wire strung 50 cm above a cushioned surface, was used to evaluate grip strength.[17] In addition, hind limb function and striatal degeneration were quantified using the limb withdrawal test, in which retraction times for each hind limb were measured after positioning the limbs in designated holes on a Perspex platform, following the methodology of Vis et al.[18] Together, these assessments provided a comprehensive evaluation of motor deficits and functional impairments.
Assessment of biochemical parameters
Tissue preparation
Rats were given ether anesthesia, and blood was drawn from them for biochemical analysis. After 24 h of the final treatment, animals were euthanized through cervical dislocation. The striatum, midbrain, and cortex were isolated, homogenized in 0.1 M phosphate buffer (pH 7.4) at 4°C, and centrifuged. The supernatant was used for biochemical analyses.
Biochemical analyses
All biochemical evaluations were performed by investigators blinded to the treatment groups to ensure objective data collection. Lipid peroxidation was quantified by measuring malondialdehyde levels using a thiobarbituric acid reaction, with absorbance recorded at 532 nm.[19] Reduced glutathione (GSH) levels were determined using Ellman’s method, with absorbance at 412 nm.[20] Catalase activity was analyzed based on hydrogen peroxide decomposition, expressed as mmol of H2O2 decomposed per minute per mg protein.[21] Superoxide dismutase (SOD) activity was assessed by its inhibition of epinephrine auto-oxidation at 480 nm.[22] Nitrite levels, reflecting nitric oxide production, were measured using a colorimetric assay with Griess reagent at 540 nm.[23] Bovine serum albumin was used as a standard; the biuret method was used to quantify the protein content. Pro-inflammatory cytokines (interleukin 1 beta (IL-1b), and tumor necrosis factor-alpha (TNF-a) were quantified by enzyme-linked immunosorbent assay using standard curves.[24]
Histopathological analysis
After being dried and preserved in 10% neutral-buffered formalin, paraffin was used to implant brain tissues, and hematoxylin and eosin were used to stain sections that were 5 mm thick for microscopic inspection at ×40 magnification.[25]
Statistical analysis
Data were presented as mean ± standard error of the mean. Two-group comparisons were performed using the unpaired t-test, while comparisons among multiple groups were conducted using one-way analysis of variance (ANOVA). Tukey’s post hoc test was applied only when ANOVA revealed statistically significant differences. Before applying parametric tests, data were evaluated for normality using the Shapiro–Wilk test and for homogeneity of variance using Levene’s test. Only datasets meeting these assumptions were subjected to ANOVA or t-test. All analyses were performed using the Statistical Package for the Social Sciences software version 21, with statistical significance set at P < 0.05. This statistical approach was applied to evaluate differences among groups and validate the study findings.
RESULTS
Organoleptic and physicochemical findings for Mucuna pruriens
The organoleptic evaluation of MP leaves and bark revealed a brown color, pleasant aroma, sweet-bitter taste, and coarse texture. Physicochemical analysis showed moisture content of 8% for leaves and 7.5% for bark, with pH values ranging from 4.0 to 7.0. Alcohol extraction gave the highest yields, 70% for leaves and 78.2% for bark. Loss on drying was 0.599% for leaves and 0.456% for bark. Ash content analysis indicated water-soluble ash of 2.134% (leaves) and 1.658% (bark), acid-insoluble ash of 0.567% (leaves) and 0.982% (bark), and sulfated ash of 4.03% (leaves) and 1.03% (bark).
Phytochemical analysis of Mucuna pruriens leaf and bark extracts
EEMP leaf and bark exhibited the highest levels of phenolics, flavonoids, and tannins compared to chloroform and water extracts. Leaf phenolic content ranged from 11.4 to 23.4 mg GAE/g, tannins from 5.9 to 7.9 mg GAE/g, and flavonoids from 14.7 to 30.18 mg RE/g, indicating strong antioxidant potential. Bark extracts contained phenolics (8.2–14.4 g GAE/100 g), tannins (0–7.2 g GAE/100 g), and flavonoids (11.6–18.11 g GAE/100 g).
In vitro antioxidant assay
DPPH radical-scavenging activity
The antioxidant potential of the extracts was evaluated through DPPH radical-scavenging activity. Among them, the ethanolic leaf extract of MP (ELEMP) showed the highest activity with the lowest IC50 value of 90.21 mg/mL, indicating strong radical-neutralizing ability. In comparison, other extracts such as chloroform leaf extract (CLEMP), ethanolic bark extract (EBEMP), and chloroform bark extract (CBEMP) had higher IC50 values, suggesting weaker activity. Ascorbic acid, the positive control, exhibited the strongest activity with an IC50 value of 42.50 mg/mL.
H2O2 scavenging activity
The extracts’ ability to scavenge H2O2 was evaluated at doses between 10 and 160 mg/mL, with ascorbic acid as the positive control. Ascorbic acid showed the highest inhibition, reaching 93.15% at 100 mg/mL. Among the extracts, ELEMP exhibited the strongest scavenging potential, with 77.15% inhibition at 160 mg/mL. CLEMP and EBEMP showed moderate activities, with inhibitions of 63.74% and 58.34%, respectively, at the highest concentrations, while CBEMP had the lowest activity, achieving a maximum inhibition of 57.51%.
Reducing power assay
The reducing power assay revealed significant antioxidant activity in all extracts. ELEMP exhibited the highest potential, with 66.78% inhibition at 160 mg/mL, followed by CLEMP at 62.33%. EBEMP and CBEMP showed moderate activity with inhibitions of 55.44% and 52.72%, respectively. Ascorbic acid, the control, had the highest inhibition at 87.28%.
Cytotoxicity study
The MTT assay indicated that ELEMP was non-toxic at concentrations up to 200 mg/mL, with cell viability above 75% at all doses (91.34% at 50 mg/mL, 90.54% at 100 mg/ML, and 85.32% at 200 mg/mL). Similarly, CLEMP, EBEMP, and CBEMP maintained viabilities above 80%, indicating their safety for biological use.
In vivo study
Acute toxicity evaluation of EEMP
The acute toxicity assessment of the ethanolic extract revealed no observable signs of toxicity or mortality in rats, even at doses up to 2000 mg/kg body weight, during the 14-day observation period.
Effect of Mucuna pruriens leaves extract on body weight in 3-NP-treated rats
The effect of ELEMP on body weight changes in 3-NP-treated rats is as follows:
VC: A slight increase in body weight (4.88 ± 0.68%).
3-NP: Induced a significant decrease in body weight (−26.76 ± 4.35%a)
ELEMP 100 + 3-NP: Marginal attenuation of weight loss (−25.17 ± 3.19%)
ELEMP 200 + 3-NP: Significant mitigation of weight loss (−15.96 ± 1.34%b,c)
ELEMP 400 + 3-NP: The highest dose exhibited maximum protection against weight loss (−10.47 ± 0.90%b,c).
Body weight was monitored over the treatment period, with percentage changes calculated from day 0 to day 14. Vehicle-treated animals maintained a stable weight, while 3-NP treatment caused significant weight loss. ELEMP at 200 and 400 mg/kg significantly reduced weight loss, with 400 mg/kg showing the strongest effect, while the 100 mg/kg dose had no significant impact.
Neurobehavioral study
The effects of ELEMP on locomotor activity, motor function scores, limb retraction time, and grip strength latency were evaluated in 3-NP-treated rats to assess its potential in improving motor function and behavior [Table 1].
| Treatment (mg/kg) | 0 Day | 7th Day | 14th Day |
|---|---|---|---|
| Effect of ELEMP on locomotor activity | |||
| VC | 388±5.45 | 400±7.21 | 395±6.68 |
| 3-NP | 367±8.32 | 238±12.66 (P=0.0004 vs. VC) | 178±9.45 (P=0.0001 vs. VC) |
| ELEMP 100+3-NP | 358±11.98 | 267±10.21 (P=0.041 vs. 3-N P) | 201±14.67 (P=0.038 vs. 3-NP) |
| ELEMP 200+3-NP | 400±19.29 | 310±15.87 (P=0.008 vs., 3-NP, P=0.034 vs. ELEMP100) | 309±14.82 (P=0.007 vs. 3-NP, P=0.030 vs. ELEMP100) |
| ELEMP 400+3-NP | 405±14.87 | 336±15.09b,d(P=0.003 vs. 3-NP, P=0.015 vs. ELEMP200) | 340±17.16b,d(P=0.002 vs. 3-NP, P=0.013 vs. ELEMP200) |
| Effect of ELEMP on motor function scores | |||
| VC | 5.8±0.3 | 6.0±0.7 | 6.2±0.6 |
| 3-NP | 5.9±0.8 | 3.6±0.5 (P=0.0002 vs. VC) | 1.8±0.4 (P=0.0001 vs. VC) |
| ELEMP 100+3-NP | 5.7±0.4 | 3.5±0.3 (P=0.060 vs. 3-NP) | 2.2±0.4 (P=0.050 vs. 3-NP) |
| ELEMP 200+3-NP | 5.8±0.6 | 4.4±0.5 (P=0.014 vs. 3-NP, P=0.045 vs. ELEMP100) | 3.7±0.4 (P=0.012 vs. 3-NP, P=0.043 vs. ELEMP100) |
| ELEMP 400+3-NP | 5.6±0.4 | 5.1±0.5b,d(P=0.008 vs. 3-NP, P=0.020 vs. ELEMP200) | 5.03±0.6b,d(P=0.006 vs. 3-NP, P=0.018 vs. ELEMP200) |
| Effect of ELEMP on limb retraction time | |||
| VC | 1.0±0.1 | 1.0±0.1 | 1.0±0.1 |
| 3-NP | 1.0±0.1 | 52.16±4.16 (P=0.0001 vs. VC) | 79.66±5.20 (P=0.0001 vs. VC) |
| ELEMP 100+3-NP | 0.9±0.1 | 48.29±5.39 (P=0.055 vs. 3-NP) | 73.17±4.28 (P=0.050 vs. 3-NP) |
| ELEMP 200+3-NP | 0.9±0.1 | 34.36±2.33 (P=0.010 vs. 3-NP, P=0.040 vs. ELEMP100) | 29.14±4.08 (P=0.008 vs. 3-NP, P=0.035 vs. ELEMP100) |
| ELEMP 400+3-NP | 1.0±0.1 | 27.32±2.90b,d(P=0.004 vs. 3-NP, P=0.022 vs. ELEMP200) | 18.22±4.98b,d(P=0.003 vs. 3-NP, P=0.018 vs. ELEMP200) |
| Effect of ELEMP on grip strength latency | |||
| VC | 50.7±4.15 | 53.59±3.19 | 55.21±5.7 |
| 3-NP | 54.7±3.89 | 12.12±0.07 (P=0.0001 vs. VC) | 5.67±0.70 (P=0.0001 vs. VC) |
| ELEMP 100+3-NP | 51.5±3.67 | 14.28±5.14 (P=0.052 vs. 3-NP) | 9.05±0.66 (P=0.049 vs. 3-NP) |
| ELEMP 200+3-NP | 55.4±5.87 | 26.16±3.18 (P=0.008 vs. 3-NP, P=0.037 vs. ELEMP100) | 28.78±2.08 (P=0.007 vs. 3-NP, P=0.032 vs. ELEMP100) |
| ELEMP 400+3-NP | 49.87±3.39 | 37.67±4.13b,d(P=0.002 vs. 3-NP, P=0.020 vs. ELEMP200) | 34.23±3.12b,d(P=0.002 vs. 3-NP, P=0.018 vs. ELEMP200) |
ELEMP significantly improved motor function and behavior in 3-NP-treated rats. While 3-NP caused declines in locomotor activity, motor scores, limb retraction, and grip strength, ELEMP (200 mg/kg and 400 mg/kg) markedly reversed these effects. The 400 mg/kg dose nearly restored locomotor activity to baseline by day 14 and showed the strongest improvements in motor scores, limb retraction time, and grip strength latency. The 100 mg/kg dose had minimal effects. These findings highlight ELEMP’s potential in managing 3-NP-induced motor impairments and neurodegenerative conditions.
Biochemical studies
The preventive benefits of ELEMP against 3-NP-induced neurotoxicity were validated by biochemical studies. 3-NP treatment reduced antioxidant defenses, as shown by lowered GSH levels and SOD and catalase activity, while dramatically raising nitrite levels, lipid peroxidation, thiobarbituric acid reactive substances, and pro-inflammatory cytokines (IL-1b and TNF-a). By decreasing oxidative damage, restoring antioxidant enzyme activity, and lowering cytokine concentrations, ELEMP treatment (200 or 400 mg/kg) successfully restored these alterations. These findings demonstrate ELEMP’s capacity to reduce oxidative stress and neuroinflammation brought on by 3-NP by acting as an antioxidant and anti-inflammatory [Table 2].
| Treatment group | Thiobarbituric acid reactive substances | Reduced glutathione | Superoxide dismutase | Catalase | Nitrite | Interleukin-1 Beta (pg/mg tissue) |
Tumor necrosis factor-alpha (pg/mg tissue) |
|---|---|---|---|---|---|---|---|
| VC | 2.82±0.07 | 5.92±0.09 | 4.38±0.06 | 3.51±0.04 | 1.18±0.08 | 170.08±11.68 | 120.67±14.27 |
| 3-NP | 8.21±0.80a(P=0.0002 vs. VC) | 1.67±0.04 (P=0.0001) | 1.32±0.02 (P=0.0001) | 1.09±0.02 (P=0.0001) | 5.19±0.60 (P=0.0003) | 732.23±14.36 (P=0.0001) | 621.57±42.49 (P=0.0001) |
| ELEMP 100+3-NP | 3.18±0.09b(P=0.042 vs. 3-NP) | 1.92±0.06 (P=0.065) | 1.48±0.03 (P=0.078) | 1.27±0.05 (P=0.082) | 4.99±0.55 (P=0.089) | 721.67±20.18 (P=0.073) | 604.34±26.29 (P=0.080) |
| ELEMP 200+3-NP | 5.14±0.47b,c(P=0.007 vs. 3-NP, P=0.018 vs. ELEM P100) | 3.08±0.04 (P=0.005, P=0.012) | 2.22±0.03 (P=0.006, P=0.013) | 1.79±0.03 (P=0.009, P=0.017) | 3.55±0.43 (P=0.011, P=0.019) | 440.43±18.98 (P=0.002, P=0.008) | 403.49±16.55 (P=0.001, P=0.007) |
| ELEMP 400+3-NP | 6.72±0.50b,d(P=0.001 vs. 3-NP, P=0.003 vs. ELEM P200) | 4.14±0.07b,d(P=0.0005, P=0.001) | 3.04±0.04b,d(P=0.0004, P=0.001) | 2.63±0.05b,d(P=0.0006, P=0.001) | 1.87±0.20b,d(P=0.0008, P=0.002) | 320.87±23.67b,d(P=0.0002, P=0.0009) | 280.67±31.89b,d(P=0.0001, P=0.0008) |
Histopathological impact of MP leaf extract on 3-NP-induced changes in rat brain regions
Effect of ELEMP on histopathological alterations in the striatum and cortex regions of control and 3-NP-treated rats, focusing on changes induced by 3-NP administration to model Huntington’s disease. Sections were visualized under a light microscope at a magnification of ×40 as shown in Figure 1.
- Histopathological changes in the striatum and cortex of control and 3-Nitropropionic acid (3-NP)-treated rats. (a) Control rats displayed normal histology in the striatum and cortex regions. (b) Rats treated with 3-NP alone showed extensive damage, with condensed pyknotic nuclei(red arrows) observed in both striatal and cortical areas. (c) In the ethanolic leaf extract of Mucuna pruriens (ELEMP) + 3-NP (100 mg/kg, p.o.) group, the striatum exhibited a reduced number of pyknotic nuclei(red arrows), indicating partial protection. (d) The ELEMP + 3-NP (200 mg/kg, p.o.) group demonstrated a more pronounced decrease in pyknotic nuclei(red arrows), reflecting improved neuroprotective effects. (e) The ELEMP + 3-NP (400 mg/kg, p.o.) group showed the greatest reduction in pyknotic nuclei(red arrows), with significantly better histological preservation in the striatum, underscoring enhanced efficacy at higher doses. (scale bar - 100 µm). All the sections were stained with hematoxylin and eosin (H&E).
DISCUSSION
Rats subjected to 3-NP-induced neurotoxicity were utilized to assess the neuroprotective benefits of MP. The 3-NP model is frequently used to mimic symptoms of HD[26] by inducing biochemical, behavioral, and histopathological alterations that closely resemble HD manifestations toxin,[27] and 3-NP induces oxidative stress and disrupts succinate dehydrogenase activity, impairing brain metabolism.[28]
This study highlighted the potential of MP, particularly its high L-DOPA content, in addressing HD pathology through antioxidant effects, neuroprotection, and modulation of inflammatory pathways. L-DOPA, a key component of MP, is known to enhance dopamine synthesis, which supports neural signaling and improves motor functions compromised in HD.[29] Phytochemical analysis confirms the presence of potent antioxidants such as flavonoids and phenolics, which exhibited strong radical scavenging activity in in vitro assays. These antioxidants neutralize reactive oxygen species, a primary contributor to cellular damage in HD.[30] The high antioxidant capacity of mucus extract, demonstrated through DPPH and hydrogen peroxide scavenging assays, indicates its potential to mitigate oxidative stress in neural cells and protect them from HD-induced degeneration.
The 3-NP-induced HD rat model demonstrated significant improvements in motor functioning, oxidative stress markers, and body weight in in vivo experiments. 3-NP administration led to weight loss, which may be attributed to metabolic disruptions and hypothalamic neuron degeneration, as well as striatal lesions and bradykinesia, which reduce food intake and appetite.[31] Treatment with MP significantly counteracted the changes in a dose-dependent manner. Behavioral assays, including locomotor activity and limb withdrawal tests, indicated that MP improved motor coordination and reduced neuromuscular impairments typically observed in HD models. Biochemical analyses showed reductions in lipid peroxidation and increased levels of antioxidant enzymes, such as catalase and glutathione. These findings suggest a reduction in oxidative stress and enhanced brain antioxidant defenses, which are often compromised in HD.
MP demonstrated significant effects on inflammatory cytokines, indicating its immunomodulatory role. In HD, elevated pro-inflammatory cytokines, including TNF-a and IL-1b, worsen neuronal damage. Treatment with MP significantly reduced these cytokines, suggesting it can suppress neuroinflammation by lowering microglial activation and pro-inflammatory signaling.
Histopathological analysis further supports its neuroprotective effects. The striatum and cortex of control rats showed normal histology. On the other hand, 3-NP therapy resulted in substantial harm. In the 3-NP group treated with lower doses of MP, a reduction in pyknotic nuclei was observed, indicating partial protection. Higher doses led to more pronounced reductions in pyknotic nuclei, with the highest dose showing the most significant neuroprotective effects.
The combined effects on oxidative stress, inflammation, and dopamine pathways highlight MP as a potential broad-spectrum neuroprotective agent in HD. It is a viable option for more exploration as a supplemental treatment for HD because of its capacity to stabilize dopamine circuits and lower oxidative stress and inflammation. Future studies should focus on isolating individual bioactive compounds within MP and exploring their specific roles in antioxidative, anti-inflammatory, and dopaminergic activities. Long-term studies on the safety and efficacy of these compounds in clinical settings would provide valuable insights for integrating MP into HD treatment regimens.
Limitations
This study has several limitations. The 3-NP model, while widely used, does not fully replicate the complex genetic basis of HD. Only male rats were used, limiting generalizability across sexes. Molecular mechanisms at the gene and protein levels were not explored. Furthermore, the lack of power analysis limits the robustness of sample size estimation. Finally, the translational applicability of the findings requires further validation through long-term and clinical studies.
Data availability statement
The datasets generated and/or analyzed during the current study are available from the corresponding author on reasonable request. Supplementary materials relevant to the experimental procedures are also available upon request.
CONCLUSION
The study demonstrates that MP possesses notable neuroprotective potential against HD-associated neurodegeneration. Its efficacy in mitigating oxidative stress, regulating inflammatory pathways, and enhancing dopaminergic function highlights its promise as a natural therapeutic candidate for managing HD symptoms.
Ethical approval:
The research/study was approved by the Institutional Review Board at Vivo Bio Tech Limited, number 28762/POBC07CPCSEA, dated 2024.
Declaration of patient consent:
Patient’s consent is not required as there are no patients in this study.
Conflicts of interest:
There are no conflicts of interest.
Use of Artificial Intelligence (AI)-Assisted Technology for Manuscript Preparation:
The authors confirm that there was no use of artificial intelligence (AI)-assisted technology for assisting in the writing or editing of the manuscript and no images were manipulated using AI.
Financial support and sponsorship: Nil.
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