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Year : 2021  |  Volume : 11  |  Issue : 2  |  Page : 137-147

Neuroprotective Effect of Tephrosia purpurea Against Mitochondrial Dysfunction by Regulation of the Caspase3/9 and Pink1/Parkin Complexes

Neuroscience Laboratory, Centre for Molecular and Nanomedical Sciences, School of Bio and Chemical Engineering, Sathyabama Institute of Science and Technology, Chennai, Tamil Nadu, India

Date of Submission07-Jan-2021
Date of Decision01-Feb-2021
Date of Acceptance16-Feb-2021
Date of Web Publication22-Apr-2021

Correspondence Address:
Rajaretinam Rajesh Kannan
Centre for Molecular and Nanomedical Sciences, Sathyabama Institute of Science and Technology, Chennai 600119, Tamil Nadu
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Source of Support: None, Conflict of Interest: None

DOI: 10.4103/ijnpnd.ijnpnd_1_21

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Background: Tephrosia pupurea is a perennial shrub that has been widely incorporated in Indian traditional medicine for its anti-inflammatory and hepatoprotective effects. Recent studies have identified T. purpurea as a source of acetylcholine esterase inhibitors. Aim: In this study, we have established the potential of T. purpurea as a potential source of drugs against Parkinsonism using an oxidopamine (6-OHDA) model. Methods: Metabolomics profiling of T. purpurea extract (TPE) was obtained using the HR-LCMS method. Enzymatic activities of Catalase, Glutathione, Superoxide Dismutase and Malondialdehyde were measured in vitro. Reactive Oxygen Species generation capacity and the mitochondrial membrane potential were also determined. The zebrafish embryos were treated with oxidopamine along with varying concentrations of T. purpurea extract and the swimming pattern and total distance travelled was evaluated. The mRNA expression of mitophagy related genes were measured using RT-PCR studies. Results: The metabolite profile of T. purpurea identified the presence of various polyphenols such as Genistein, Esculetin, and Chrysin that have neuroprotective effects. 6-OHDA-induced PD causes an increase in oxidative stress, reactive oxygen species generation, and affects mitochondrial stability. There was a significant increase in the catalase, glutathione, and superoxide dismutase levels and a decrease in Malondialdehyde and Reactive Oxygen Species levels in cells treated with TPE when compared to 6-OHDA treated cells. We then treated zebrafish embryos with 6-OHDA along with varying concentrations of T. purpurea extract, and the mRNA expression and swimming pattern were evaluated. The embryos cotreated with TPE showed improved swim pattern similar to untreated embryos, whereas those treated with the positive control failed to do so. T. purpurea extract also significantly decreased the expressions of casp3, casp9, lrrk2, and increased pink1 and parkin expression. Conclusion: Our study identifies Tephrosia purpurea extract as a viable candidate against 6-OHDA induced-neurotoxicity, and further studies of its effect in models of neurodegenerative diseases are required.

Keywords: Caspases, oxidopamine, Pink1/Parkin, Tephrosia purpurea, zebrafish

How to cite this article:
Kesh S, Kannan RR. Neuroprotective Effect of Tephrosia purpurea Against Mitochondrial Dysfunction by Regulation of the Caspase3/9 and Pink1/Parkin Complexes. Int J Nutr Pharmacol Neurol Dis 2021;11:137-47

How to cite this URL:
Kesh S, Kannan RR. Neuroprotective Effect of Tephrosia purpurea Against Mitochondrial Dysfunction by Regulation of the Caspase3/9 and Pink1/Parkin Complexes. Int J Nutr Pharmacol Neurol Dis [serial online] 2021 [cited 2022 Dec 5];11:137-47. Available from:

   Introduction Top

Plants have long been used as a source to identify and harvest molecules that are beneficial to human health. Because most plants are consumable and nontoxic to humans, they pose as viable candidates for the screening of therapeutic compounds and drugs. The phytoconstituents of several plants have been isolated and studied extensively for their properties.[1] These plant metabolites have high therapeutic efficacy against multiple conditions such as cancer, liver disorders, microbial infections, and neurodegenerative disorders, whereas lying low on the spectrum of side effects.[2],[3],[4] Ethanopharmacology studies have helped identify the sources of drugs that have been used commonly and traditionally. Screening of these traditionally used sources would help in discovering therapeutic natural molecules from them.[5]

Tephrosia pupurea (Linn.) Pers. is a perennial plant belonging to the family Fabaceae that is found throughout the Indian subcontinent and Sri Lanka. Locally, this plant is known as “Sarapunkha” and is used in Ayurveda and traditional medicine concoctions.[6],[7],[8] Ayurvedic literature refers to the plant as “sarva warnavishapak” due to its potential in healing all wound types. Traditionally this plant is most sought after to heal wounds and treat liver and gastric ailments.[9],[10],[11],[12],[13],[14] Whole-plant extracts have been shown to cure inflammatory conditions, ulcers, and tumors. The roots, stem, leaves, and pods are also used separately to treat ailments.[12],[13],[14]

Parkinson disease (PD) is a neurodegenerative disease characterized by loss of neurons in the brain causing dementia and involuntary movements such as tremors, bradykinesia, and loss of rigidity.[15] Mitochondrial dysfunction has been deemed as one of the major causes contributing toward the pathogenesis of PD. DJ-1, PINK1, and Parkin are involved in the quality control of mitochondria and cell survival. The PINK1/Parkin complex has been studied extensively and loss of function of these genes has been reported in cases of autosomal recessive PD.[16] Leucine-Rich Repeat Kinase 2 (LRRK2), a serine/threonine kinase, involved in mitochondrial functioning, is the cause of sporadic and autosomal dominant familial PD.[17] Mitochondrial dysfunction characterized by calcium imbalance, loss of membrane stability, impaired trafficking, increase in oxidative stress, and mitophagy are associated with the pathogenesis of PD and involve PINK1 (PARK6), Parkin (PARK2), and LRRK2 (PARK8).[18] Targeting of these mitochondria-related genes implicated in PD would help modulate the pathways involved in the disease and help identify therapeutic targets against PD.

Oxidopamine (6-OHDA) causes neurotoxicity by the inhibition of mitochondrial complexes. The primary mode of action of oxidopamine is an increase in oxidative stress leading to mitochondrial dysfunction.[19] This has been widely used to model neurodegenerative conditions in animals.[20] Zebrafish is a strong model to mimic and study mammalian disorders and to help screen for potential drugs.[21] The analogy of their genes to humans further their standing as a sought-after model.

In this study, we aimed to assess the neuroprotective effects of Tephrosia purpurea in a PD model in vivo and in vitro, to determine its anti-apoptotic activity, and its ability to modify mitochondria-related genes involved in PD.

   Materials and Methods Top

Preparation of methanol extract

The leaves of Tephrosia purpurea (500 g) were collected, washed under running tap water; shade dried, and coarsely powdered using a mechanical pulverizer. The plant powder was then soaked in methanol at room temperature in the ratio of 1:4 (w/v) and was left undisturbed for 72 hours. This extract was filtered using Whatman filter paper No.1, to which methanol was added and the process continued till the color of the filtrate was pale. All the filtrates were concentrated to 2% of their original volume under reduced pressure and the Tephrosia Purpurea extract (TPE) was collected in an amber bottle and kept at room temperature.

Phytochemical analysis of the extract

The High Resolution Liquid Chromatograph Mass Spectrometer (HR-LCMS) of the extract was carried out at the Sophisticated Analytical Instrument Facility (SAIF), IIT Bombay, India. The sample was analyzed using the Q-Exactive Plus– Orbitrap MS, Thermo Scientific and the data were acquired using the Thermo Scientific Xcalibur (Version software. The sample run was for a total of 30 minutes in a Hypersil Gold 3 micron 100 × 2.1 mm column (Thermo Scientific) with Solvent A: 0.1% formic acid in Milli-Q water and Solvent B: methanol. The result obtained was then analysed using the Compound Discoverer 2.1 SP1 software.

Cell culture model

The SH-SY5Y Human Neuroblastoma Cell line was obtained from National Centre for Cell Science (NCCS), Pune, India. The cell line was maintained in a Dulbecco’s Modified Eagle’s medium (DMEM) that was supplemented with fetal bovine serum (FBS), penicillin, and streptomycin in a CO2 incubator.

Cell viability assay

Firstly, the cell viability was measured against varying concentrations of TPE to determine the lethality of the extract using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) reagent. The cells were suspended in 96-well plates and incubated with the MTT reagent for 24 hours following which it was washed and a stopping reagent was added. The absorbance was measured at 570 nm. After establishing the cytotoxicity of the TPE, the protective effect of TPE against 6-OHDA induced toxicity was determined.

Superoxide dismutase and catalase assays

Superoxide dismutase (SOD) and catalase (CAT) levels were measured to determine the anti-oxidant potential of the cells against cell damage. The SOD levels were measured using SOD Kit (R&D systems INC) and measured at 450 nm using an ELISA reader. CAT activity was measured using Catalase Activity Assay Kit (Abcam) at an emission of 587 nm.

Glutathione and lipid peroxidation assays

Glutathione (GSH) activity was measured as per the manufacturer’s protocol using the GSH Detection Assay Kit (Abcam, USA). The absorbance was measured at 380 nm. The malondialdehyde (MDA) activity was assessed using the TBARS/MDA Assay Kit (Cayman Chemicals, USA) and the absorbance was measured at 530 nm. This assay is indicative of the lipid peroxidation activity.

Reactive oxygen species assay

The reactive oxygen species (ROS) generation assay was carried out using the Cellular ROS Assay Kit from Abcam, USA. The kit measures the increase in red fluorescence emission which occurs as a result of cell damage at 605 nm.

Mitochondrial membrane potential assay

The mitochondrial membrane potential (MMP) of the cells was analyzed using a fluorescent cationic probe, JC-1, and measured using a flow cytometer (BD FACS Calibur). This dye can be measured by its uptake into the mitochondria of healthy, live cells facilitated by the membrane potential (Δψ). Loss of mitochondrial membrane potential in apoptotic and damaged cells affects the uptake of the dye.

Animal husbandry and breeding

Wild-type AB strain Zebrafish were reared and maintained under 14:10 light and dark conditions with a temperature of 26 to 28°C. Sexually mature adult fishes were chosen for breeding. The male and female fishes were placed in breeding tanks in the ratio 2:1, respectively. They were left undisturbed overnight and the embryos were collected the next day, screened, and reared in E3 medium. The embryos were treated with the neurotoxin and varying drug concentrations in 24-well plates under optimum conditions.

Ethical Statement

The handling and experimentation on zebrafish were carried out in a humanly manner following established guidelines for housing and rearing of Danio rerio. The study was conducted under the supervision of the Institutional Bioethical Committee (IBC) (No.2017/201679609/RRK8-23). All experimental protocols were in accordance with the ARRIVE guidelines and OECD guidelines for toxicity analyzing experiments.

Fish embryo toxicity

The embryos were exposed to varying concentrations of TPE and their mortality, hatching rate, and morphology were observed till 96 hours postfertilization (hpf) according to the OECD guidelines.[22] Any developmental teratology such as spinal bending, tail bending, absence of eyes or mouth, and edemas were observed. Working standards of the solutions were prepared by dilution to the required concentrations with a final concentration of 0.1% DMSO in the wells.

Locomotion and behavior

Oxidopamine- and TPE-treated embryos were placed individually in a 96-well plate. Post acclimatization for 30 minutes, the movement of the embryos was recorded for 2 minutes and their swim pattern was analyzed using UMATracker software. The total distance travelled was also calculated.

Real-time PCR analysis

Two dpf (days post fertilization) zebrafish embryos were placed in the solutions and treated for 72 hours. Posttreatment, 30 embryos per treatment were taken and their RNA was isolated using RNAzol Reagent (Sigma). cDNA conversion of the collected RNA was carried out using the RevertAid First Strand cDNA synthesis kit (Thermo Scientific). The real-time PCR was carried out in the QuantStudio Real-Time PCR System (Applied Biosystems). Real-time PCR analysis was carried out for the genes lrrk2, prkn, pink1, casp3, casp9, and β-actin. The sequences of the primers are mentioned in [Table 1]. The expression was calculated using the ΔΔCt method where β-actin was the internal reference.
Table 1 Primer sequences

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Statistical analysis

The data obtained were analyzed using one-way ANOVA and Sidak’s multiple comparison tests using Graph Pad Prism software. Mean values were calculated for the data and the P-values < 0.05 (95% confidence interval) were considered as statistically significant.

   Results Top

Identification of metabolites in T. purpurea extract

HR-LCMS analysis [Figure 1] of the methanolic extract of T. purpurea helped identify 21 compounds in the positive ion mode [Table 2] and four compounds in the negative ion mode [Table 3]. The extract was rich in flavonoids with traces of coumarins, steroids, and alkaloids.
Figure 1 HR-LCMS of T. purpurea extract. Total ion chromatogram (TIC) of TPE in (a) positive ion mode and (b) negative ion mode. Abbreviation: TPE, T. purpurea extract

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Table 2 Compounds identified in HR-LCMS in positive ion mode

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Table 3 Compounds identified in HR-LCMS in negative ion mode

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TPE protects against 6-OHDA toxicity

Prior to establishing the protective effects of TPE against 6-OHDA, we tested the cytotoxicity of TPE individually. TPE treatment failed to cause lethality in the cells at higher concentrations up to 500 μg/ml [Figure 2]a]. We summarized that the extract was not toxic to the cells. Treatment with 25 µM 6-OHDA induced a significant decrease in the cell viability of the SH-SY5Y cells. Treatment with TPE on 6-OHDA induced cells significantly improved the cell viability that establishes the cytoprotective effect of TPE against 6-OHDA [Figure 2]b.
Figure 2 Effect of TPE on 6-OHDA induced toxicity in SH-SY5Y cells. (a) Cell viability in response to TPE. (b) Cell viability in response to 6-OHDA + TPE. (Means significantly differ by Sidak’s multiple comparison test at P < 0.05). Abbreviations: 6-OHDA, oxidopamine; TPE, T. purpurea extract

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TPE alleviates oxidative stress markers and ROS generation

6-OHDA treatment significantly decreased the levels of CAT, SOD, and GSH, whereas increasing the level of MDA. Treatment with TPE significantly reversed the activity of these biomarkers [Figure 3]. Although the levels of CAT, SOD, and GSH improved significantly on treatment with TPE, the level of MDA was reduced indicating the ability of TPE in alleviating oxidative stress damage. Treatment with the positive control also significantly reversed the levels of the oxidative stress markers. ROS generation was also increased on 6-OHDA treatment signifying oxidative damage. Treatment with TPE significantly reduced the levels of ROS generation [Figure 4].
Figure 3 Measurement of (a) catalase (CAT) (b) glutathione (GSH) (c) superoxide dismutase (SOD), and (d) lipid peroxidation (MDA). 6-OHDA treatment resulted in a dramatic decrease in CAT, GSH, and SOD levels when compared to the control, whereas MDA was increased. Co-treatment with 62.5 and 125 μg/mL TPE in 6-OHDA induced cells showed a significant increase in the enzyme level of CAT, GSH, and SOD, whereas the MDA levels were significantly decreased. Abbreviations: 6-OHDA, oxidopamine; CAT, catalase; GSH, glutathionine; SOD, superoxide dismutase; TPE, T. purpurea extract

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Figure 4 Measurement of ROS. 6-OHDA treatment significantly increased the production of ROS when compared to the control. Co-treatment with 62.5 and 125 μg/mL TPE in 6-OHDA induced cells showed a significant decrease in the ROS level. Abbreviations: 6-OHDA, oxidopamine; ROS, reactive oxygen species; TPE, T. purpurea extract

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TPE improves mitochondrial membrane potential

Treatment of SH-SY5Y cells with 6-OHDA caused a significant decrease in the membrane potential. The uptake of JC-1 is impaired in cells with a depolarized membrane due to which the staining fluoresces in FL-1 but not in FL-2. Cells that fluoresce in both FL-1 and FL-2 have a healthy, functioning, and polarized membrane that facilitated the uptake of JC-1 inside the membrane. Treatment with TPE improved the membrane potential of the mitochondria significantly [Figure 5].
Figure 5 Effect of TPE on mitochondrial membrane potential. 6-OHDA treatment significantly decreased the membrane potential of the cells. Treatment with TPE increased the MMP significantly. (Means significantly differ by Sidak multiple comparison test at P < 0.05)

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Developmental and phenotypic effects of TPE in Zebrafish embryos

The zebrafish embryos were treated with increasing concentrations of TPE to determine the toxicity and lethality. Pericardial edema and body axis bending was observed at concentrations above 60 μg/mL but the teratological effects were not significant. Clotting of blood in the pericardium was observed at concentrations above 80 μg/mL that were absent in the control fishes and lower concentrations of TPE. At concentrations 100 μg/mL and higher, no hatching was observed [Supplementary Figure 1].

TPE rescues 6-OHDA induced locomotion impairment

6-OHDA-treated embryos showed a disruptive and reduced swim pattern when compared to the control fishes. Treatment with TPE at 10 and 20 μg/mL showed a significant improvement in the swim pattern [Figure 6] and the total distance traveled [Figure 7] when compared to the fishes treated with 6-OHDA [250 µM]. Treatment with l-DOPA (50 µM) failed to improve the motor impairment of the embryos.
Figure 6 Locomotion of zebrafish larvae. Treatment with TPE reversed the effects of 6-OHDA and improved the swimming pattern of the zebrafish larvae

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Figure 7 Total distance traveled by zebrafish. Treatment with TPE significantly increased the total distance traveled by the fishes when compared to 6-OHDA-treated group. The data are represented as a mean of the total group. (Means significantly differ by Sidak multiple comparison test at P < 0.05). Abbreviations: 6-OHDA, oxidopamine; TPE, T. purpurea extract

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Anti-apoptotic activity of TPE

The mRNA expression of apoptotic genes casp3 and casp9 was upregulated following 6-OHDA treatment concurrent with cell damage. Treatment with TPE reduced the expressions of both casp3 and casp9, whereas treatment with l-DOPA only rescued casp9 expression [Figure 8].
Figure 8 Effect of TPE on mRNA expression of apoptotic genes (a) casp3 (b) casp9. The upregulation in mRNA expression of casp3 and casp9 as a result of 6-OHDA treatment was reversed on treatment with TPE. (Means significantly differ by Sidak multiple comparison test at P < 0.05). Abbreviations: 6-OHDA, oxidopamine; TPE, T. purpurea extract

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TPE modulates mitophagy-related genes

The expression of pink1, parkin, and gsk3b were significantly modified on the treatment of zebrafish embryos with 6-OHDA. Although the expressions of pink1 and prkn were downregulated on 6-OHDA treatment, the expression of lrrk2 was upregulated. Treatment with l-DOPA failed to rescue the expression of lrrk2. Treatment with TPE significantly increased pink1 and prkn expressions, whereas simultaneously reducing lrrk2 expression [Figure 9].
Figure 9 Effect of TPE on mRNA expression of serine/threonine kinases (a) pink1, (b) prkn, and (c) lrrk2. The downregulation of pink1 and parkin were rescued on treatment with TPE. Treatment with TPE also downregulated the expression of lrrk2 compared to 6-OHDA treated group. (Means significantly differ by Sidak multiple comparison test at P < 0.05). Abbreviations: 6-OHDA, oxidopamine; TPE, T. purpurea extract

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   Discussion Top

Our current study has shown the neuroprotective potential of T. purpurea extract against 6-OHDA-induced toxicity in in vivo and in vitro models. This study thus highlights Tephrosia purpurea as a potential source to identify and isolate neuroprotective compounds against mitochondrial dysfunction in neurodegenerative conditions.

Oxidative stress is a vital factor that influences mitochondrial functioning and the progression of neurodegenerative diseases such as PD. Tephrosia purpurea has long been used in traditional and Indian medicinal practices for its hepatoprotective,[13],[23] anti-inflammatory,[24],[25] antiulcer,[12],[26] antimicrobial,[27],[28] anti-allergic,[29],[30] antidiabetic,[31],[32] anticancer,[33],[34] and wound healing[9] properties. The plant has high anti-oxidant potential and free radical scavenging capacity establishing its therapeutic ability to reverse oxidative stress damage.[35],[36] Although traditionally T. purpurea is not used for its neuroprotective properties, research has highlighted it as a candidate against neurodegenerative conditions. Recent studies have shown acetylcholinesterase inhibitory activity of molecules isolated from T. purpurea extract.[37],[38],[39],[40] The phytochemical analysis of TPE determined its rich flavonoid content and identified the presence of flavonoids, flavones, glyosidic flavonoids, isoflavones, and prenylflavonoids similar to previous reports. Flavonoid-rich fractions of Tephrosia purpurea have been reported for their therapeutic benefits against various disorders.[29],[32],[41]

Oxidopamine aptly mimics Parkinsonian-like conditions by causing oxidative stress and impairing the mitochondrial complex. The hallmarks of 6-OHDA induced neurotoxicity include an elevation of oxidative stress markers, increased ROS generation, mitochondrial dysfunction, and cell death.[42] The activities of catalase, glutathione, and superoxide dismutase are measured to assess the extent of damage by oxidative stress.[43] A decrease in these biomarkers indicates an increase in oxidative stress in the cell. Lipid peroxidation levels are measured by measuring the malondialdehyde levels that is a by-product of lipid peroxidation and is increased during oxidative damage to the cells. In our study, treatment with 6-OHDA significantly decreased the levels of CAT, SOD, and GSH, whereas reducing MDA levels in the SH-SY5Y cells. Treatment with TPE significantly reversed the activity of these biomarkers.

As mentioned previously, 6-OHDA also causes an increase in ROS production and mitochondrial dysfunction. To assess this, we measured the generation of ROS in response to 6-OHDA and the MMP of the cells. Concurrent with previous studies, there was a significant increase in ROS generation and a significant loss in the membrane potential of the mitochondria.[44] Loss of membrane polarity of the mitochondria is indicative of mitochondrial damage. The stability of the mitochondrial membrane is important for robust cell functioning and the loss of membrane stability has is an indicator of the onset of PD in patients. The increase in ROS generation in the cells as a result of 6-OHDA was significantly reduced when treated with TPE. Furthermore, treatment with TPE also improved the MMP indicative of its ability to restore polarity and stability to the mitochondrial membrane.

Zebrafish is a well-defined and malleable model to study neurodegenerative conditions.[45] Studies show that treatment with neurotoxins can induce Parkinsonian conditions both on a behavioral and molecular level. Consistent with previous reports, we observed a decrease in the distance traveled by the fish when treated with 6-OHDA.[46] The fish also exhibited prolonged periods of stagnation compared to the control fish. These behavioral patterns are a paradigm of neurodegeneration. Treatment with TPE significantly improved the locomotion of the fish, whereas the positive control, levodopa, failed to elicit the same.

Oxidative stress damage in neurodegeneration involves dysfunction and modification of several apoptotic and signaling pathways. Mutations in genes involved in these pathways can lead to the onset and progression of the disease.[47] The intrinsic apoptotic pathway, also known as the mitochondria-mediated pathway, has been an indicator of cell death and mitochondrial dysfunction in PD.[48],[49] The activation of caspase-9 initiates the pathway and activates caspase-3. The increase in the expressions of caspase-3 and caspase-9 has been studied in PD models for their involvement in neuronal death.[50],[51] To assess the involvement of the intrinsic pathway, we observed the levels of casp3 and casp9 expression in a 6-OHDA induced zebrafish model. Both casp3 and casp9 levels were elevated on treatment with 6-OHDA. Treatment with TPE significantly reduced the expressions of both casp3 and casp9 while levodopa only reduced casp9 expression significantly.

The process of removing defective mitochondria from the cells by the process of autophagy, called mitophagy, is observed to be compromised in patients with PD. Impairment of mitophagy, in turn, leads to the accumulation of damaged mitochondria resulting in neuronal death and neurodegeneration. Various genes and their proteins such as PINK1, Parkin, LRRK2, SNCA, VPS35, and CHCHD2 that are implicated in familial PD are also involved in maintaining mitochondrial homeostasis.[18] LRRK2 and PTEN (phosphatase and tensin homolog)-induced putative kinase 1 (PINK1) have been extensively studied in PD patients. Both LRRK2 and PINK1 mutations have been shown to cause early onset and familial PD in humans.[17] Loss of function of PINK1 leads to defective mitochondrial pathologies and results in the loss of dopaminergic neurons.[52] The PINK1-Parkin complex is the most common and prominent mitophagy pathway studied in the progression of PD. LRRK2 along with the PINK1-Parkin complex is implicated in damaged mitochondrial characteristics such as increase in oxidative stress, defective mitophagy, incorrect mitochondrial trafficking, and impaired electron transport chain.[53] Studies have identified inhibition of LRRK2 or activation of PINK1 and Parkin as potential therapeutic solutions for PD.[54],[55] Hence, these pathways are studied for identifying possible therapeutic targets that would effectively mitigate the disease condition. Our work focused on studying the expression levels of mitophagy related genes- pink1, prkn, and lrrk2 in an oxidopamine model of zebrafish. Our study showed that 6-OHDA treatment resulted in an upregulated expression of lrrk2, whereas pink1 and prkn expressions were downregulated. Treatment with our extract, TPE, significantly reduced the expression of lrrk2 while upregulating pink1 and prkn expressions in the zebrafish model. This is conclusive of the protective and therapeutic effect of TPE against mitochondrial dysfunction involved in PD.

   Conclusion Top

Our findings illustrate the neuroprotective potential of T. purpurea against 6-OHDA-induced neurotoxicity by increasing CAT, SOD, and GSH levels while decreasing MDA and ROS generation levels and protecting the mitochondrial membrane potential. T. purpurea successfully improved the locomotor impairments in zebrafish embryos. Furthermore, TPE reduced the expression of the caspases while mitigating the oxidopamine induced changes in the expression of mitophagy-related genes.


We acknowledge and thank Dr.Anandan Balakrishnan, Department of Genetics, Dr ALM PGIBMS Campus, University of Madras, Taramani, Chennai, Tamil Nadu, India for supporting and providing resources to facilitate the RT-PCR studies.

Financial support and sponsorship

This research was funded and supported by the Department of Science and Technology, India under INSPIRE Fellowship [No. DST/INSPIRE Fellowship/2016/IF160639].

Conflicts of interest

There are no conflicts of interest.

   References Top

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  [Figure 1], [Figure 2], [Figure 3], [Figure 4], [Figure 5], [Figure 6], [Figure 7], [Figure 8], [Figure 9]

  [Table 1], [Table 2], [Table 3]


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