Swimming training and herbal nanoformulations as natural remedies to improve sensory-motor impairment in rat midbrain tumor models: system biology, behavioral test, and experimental validation

Abstract

Motor impairment worsens health-related quality of life in patients with primary and metastatic midbrain tumors. Here, 56-male-Wistar rats were divided into eight groups: Normal group, Midbrain Tomur Model group, Model + Exe group, Model + Lipo, Model + Extract, Model + Lipo-Extract, Model + Extract-Exe, Model + Lipo-Extract + Exe. According to the aim, mid-brain tumor models were conducted by injections of the C6 glioma cell line (5 × 10⁵ cell suspension) and stereotaxic techniques in the substantia nigra area. Furthermore, consumption of nanoformulation of herbals extract (100 mg/kg/day), crude extract (100 mg/kg/day), and swimming training (30 min, 3 days/week) as interventional protocols were performed for 6 weeks. In addition, we evaluated the effect of polyherbal nanoliposomes containing four plant extracts and swimming training on the GABArα1/TRKB/DRD2/DRD1a/TH network in the substantia nigra of the midbrain tumor rat model. Data emphasized that DRD2 might be a druggable protein with the network's highest significance cut-point effect that could modulate sensory-motor impairment. Furthermore, we found Quercetin, Ginsenosides, Curcumin, and Rutin, as bioactive compounds present in Ginseng, Matthiola incana, Turmeric, and Green-Tea extracts, could bind over the DRD2 protein with approved binding affinity scores. Based on our data, swimming training, and nanoliposome-enriched combined supplements could consider effective complementary medicine for motor impairment recovery induced by the midbrain tumor in the substantia nigra area. Hence, regular swimming training and natural medicines rich in polyphenolic bioactive components and antioxidative effects could modify and improve the dopamine receptors' function.

Supplementary Information

The online version contains supplementary material available at 10.1007/s13205-023-03574-3.

Introduction

Parkinson's disease (PD) is a neurodegenerative condition which causes by the loss of dopaminergic neurons in the substantia nigra. Evidence has revealed that dopaminergic neuron impairment in the nigrostriatal pathway cause motor impairments and inability (Choi et al. 2017). Moreover, differential expression of genes involved in the cell cycle program, proliferation, survival, invasion/metastasis, cancer stemness, oxidative stress, inflammation, cell death, drug resistance, DNA repair, and DNA stability might play brain tumor development (Afshari et al. 2021).

While there is a significant advance in the prognosis and diagnosis of pathogenesis mechanism malignant brain tumors, Only 20–25% of patients have improved survival through approved ameliorate approaches (Van Tellingen et al. 2015). Several reports have shown that in half of the identified brain tumors with Parkinson's symptoms, lesions, and small lesions are seen in the substantia nigra of the basal ganglia nuclei (Ye et al. 2016; Margulies 1953; Yasuhara et al. 2009). In addition, brain tumors may cause motor impairment, such as limb weakness, plegia, ataxia, spasticity, and the inability to perform complicated movements (Amidei and Kushner 2015). Dopamine involves in various cognitive and behavioral processes, including motor activity, reward processing, and decision-making (De Corte et al. 2019).

Previous studies have shown that the expression level of Caspase-3 increased in dopamine-producing neurons (Pariyar et al. 2022; Hornykiewicz 1986; Nosheny et al. 2006; Schneider et al. 1987). This phenomenon is associated with decreased neurons expressing tyrosine hydroxylase (TH) (Pariyar et al. 2022). Tyrosine hydroxylase is an enzyme in the family of aromatic amino acid hydroxylase which is expressed in the central nervous system. Moreover, TH can catalyze the conversion of tyrosine to L-3 and 4-dihydroxyphenylalanine (L-DOPA) (Hortelano et al. 2010). After a series of downstream enzymatic reactions, L-DOPA is eventually converted to the messenger molecule and neurotransmitter dopamine (Reyes et al. 2013). Dopamine and dopamine receptors can play a critical role in controlling movement, emotions, and the reward system in the brain (Reyes et al. 2013). Furthermore, dopamine and dopamine receptors are vital in dopamine-producing neurons' death following substantia nigra damage (Reyes et al. 2013). In many neurological diseases involving the substantia nigra and striatum, dysfunction of dopamine receptors D1 and D2 at the cellular level has been observed. Based on the evidence, dysfunction of dopamine receptors leads to exacerbation of the disease process and loss of the population of dopamine-producing neurons (Mishra et al. 2018).

Another essential receptors in the brain are gamma-aminobutyric acid (GABA) (Van Tellingen et al. 2015). The GABA neurotransmitter regulates the inhibitory-excitatory balance that may be vital for brain function (Wu and Sun 2015). In many neurological disorders and brain injuries, dysfunction of the GABAergic system has been observed (Wu and Sun 2015). Tropomyosin receptor kinase B (TrkB) is another crucial receptor that plays a pivotal role in the proliferation and maintenance of dopamine-producing neurons. The binding of brain-derived neurotrophic factor (BDNF) growth factor to TrkB receptors switches on three critical messaging pathways of mitogen-activated protein kinase (MAPK), phosphoinositide 3-kinase (PI3K), and phospholipase C- gamma 1 (PLC-γ1), which have a crucial role in neurogenesis, survival, and differentiation of brain neurons. Studies have revealed that increasing TrKB receptor expression and triggering the BDNF/TrkB complex by gene transfer methods using viral vectors might increase the survival of dopamine-producing neurons remaining in the damaged area (Kang et al. 2017).

Today, there are a variety of therapies for PBT (primary brain tumors) and motor sensory impairment induced by PBT, including chemotherapy, radiation therapy, surgery, and targeted drugs, some of which have life-threatening side effects (Cote and Smith 2016; Erel-Akbaba et al. 2019). However, several resistance mechanisms such as insufficient penetration of chemotherapeutic drugs through the blood–brain barrier and an increased likelihood of temozolomide resistance in a subset of patients, pose significant challenges to existing treatment approaches (Eder and Kalman 2014). On the other hand, the pharmaceutical method is currently the primary therapeutic strategy for PD patients to reduce or manage motor symptoms (Wang et al. 2020). The therapy is often targeted to boost dopamine bioavailability, either by replenishing the dopamine precursors or preventing the breakdown of dopamine (Wang et al. 2020). During the first stages of therapy, levodopa (also known as L-dopa), the precursor of dopamine, is administered as the mainstay (Postuma et al. 2015).

In contrast to dopamine, levodopa may enter the brain and be converted into dopamine (Postuma et al. 2015). During the early stages of the illness, when therapy with levodopa is most effective, patients often experience a significant improvement in their symptoms, and this improvement is used as a diagnostic criterion for PD (Postuma et al. 2015). However, as the symptoms advance and more dopaminergic neurons in the substantia nigra are lost, the treatment efficacy declines. Motor fluctuations, "on/off" phenomena, and dyskinesias are typical long-term levodopa medication side effects (Wang et al. 2020). Advance-stage PD patients now face a treatment challenge due to levodopa-related comorbidities and impairment (Wang et al. 2020). Therefore, there is a need to find lower-risk approaches, complementary and conventional therapies to reduce the adverse effects (Johnson et al. 2019). Complementary and alternative medicine (CAMs) combined with commercial/conventional pharmaceuticals could be the most achieved as combinational medical therapy in pathogenic hallmark offset (Hajibabaie et al. 2022a, c). Conventional medical treatment is combined with CAMs techniques established to be safe and effective by scientific research (Barari et al. 2017; Abedpoor et al. 2022a).

Common brain tumor treatments include surgery, chemotherapy, radiation therapy, and targeted drug therapy (Cote and Smith 2016). However, common treatments have several side effects that can be life-threatening (Erel-Akbaba et al. 2019). Therefore, the need to find lower-risk or complementary and conventional therapies to reduce the side effects has been raised recently (Hajibabaie et al. 2022c). Today many countries consider physical exercise as a potential therapeutic approach (Abedpoor et al. 2022b). Exercise is recognized as a simple, inexpensive, and effective method to prevent and improve many inflammatory diseases (Morielli and Courneya 2020; Abedpoor et al. 2022a, b; Bahadorani et al. 2019; Hajibabaie et al. 2022b). Robert Newton et al. have displayed that despite the problems and complexities involved in treating brain tumors, aerobic training can positively and psychologically affect patients to reduce their anxiety and depression and increase the survival rate (Newton and Galvao 2008; Cormie et al. 2015). Exercise training has been identified as an affordable, simple, and effective therapy for preventing and treating a variety of inflammatory responses, oxidative stress, cell death, proliferation, survival, and invasion/metastasis (Zhang et al. 2016b). In most of the evidence, it has been observed that swimming exercises might decrease the formation of malignant cells (Barari et al. 2017; Zhang et al. 2016a; Almeida et al. 2009).

Moreover, swimming aerobic exercise could be improved neurogenesis and cognition, promote nerve anti-inflammatory, reduce neuropathic pain, and improve muscle mass and function (in mice models research) (Abedpoor et al. 2022b; Guo et al. 2020).

In addition, aerobic exercise and bioactive components derived from various plants have also been recognized as natural, complementary, and alternative treatments with low risk and side effects for treating many diseases (Mainardi et al. 2009; Adib-Hajbaghery et al. 2021; Hajibabaie et al. 2022c).

Based on phytochemical Investigations evidence, Turmeric, Green-Tea, Ginseng, and Matthiola incana are rich in Gallic acid, Caffeine, Epigallocatechin-3-gallate, Ferulic acid, Caffeic acid, Curcumin, Chlorogenic acid, Ginsenosides, Quercetin, Rutin, and Ellagic acid. Previous studies reported that Turmeric, Green-Tea, Ginseng, and Matthiola incana could have anti-inflammatory and antitumor effects, antioxidant substances, and high redox efficiency (Vishvakarma 2014; Kochman et al. 2020; Ratan et al. 2021; Liu et al. 2020; Miceli et al. 2021). Therefore, a compelling mixture of bioactive compounds can improve immune system activity, neurological function, cardiovascular protection, and anti-toxicity rate.

Overall, this study aimed to investigate the simultaneous effect of swimming training with a combined supplement containing four nanoliposome herbal extracts on activating the dopamine D1/D2 receptor molecular pathway in the substantia nigra of the brain tumor rat model. This study's primary goal is to appraise the effects of swimming training and a mix of herbal drugs (extract and nanoliposome forms) as a complementary strategy to improve sensory-motor impairment features in rats with brain tumors.

Materials and methods

Study design

This study evaluated the sensory-motor integration impairment induced by brain tumors based on the in-silico machine and in-vivo study. According to the aim, mid-brain tumor models were induced by injections of the C6 glioma cell line (5 × 10⁵ cell suspension) in the substantia nigra area of male Wistar rats. Moreover, we assessed the simultaneous effect of swimming training with a non-formulated combined supplement containing four plant extracts rich in polyphenolic compounds on activating the dopamine D1/D2 receptor molecular pathway in the substantia nigra of the brain tumor rat model. Consumption of nanoformulation of herbals extract (100 mg/kg/day), crude extract (100 mg/kg/day), and swimming training (30 min, 3-day/week) as interventional protocols were performed for 6 weeks. Notably, the small molecules derived from herbal medicines and 3D structures were detected using computational drug discovery and design tools. In addition, the beam test and functional sciatic index assessed nerve function and motor coordination. After 6 weeks of swimming training and consuming nanoliposomes enriched combined supplement, rats were sacrificed. Afterward, brain tissues were collected and transferred to the laboratory.

Bioinformatics tools in the identification of sensory-motor impairment induced by midbrain tumor

System biology was performed to recognize critical genes and molecular signaling pathways involved in the midbrain tumor pathogenesis and side symptoms such as sensory-motor integration impairment. For this purpose, we browsed Gene Expression Omnibus (GEO) database and selected a microarray dataset with ID number GSE7707 for Parkinson's and a microarray dataset with ID number GSE78895 for midbrain tumor (Akgül et al. 2018). Datasets were analyzed in R programing language software by applying MAS5 normalization and limma and s4vector packages. As a result, we displayed significant genes with a P value < 0.001 by the heatmap plot.

Moreover, to discover significant genes with differential expression patterns, we analyzed R outputs considering P value < 0.05 and logFC ± 0.5 as a cut-off threshold for Parkinson's dataset and P value < 0.05 and logFC ± 4 for the midbrain tumor dataset. The significant genes were used to construct the protein–protein interactions (PPIs) network in the STRING v.11.5 databases (Szklarczyk et al. 2021). On the other hand, we engaged the network's visual parameters, including degree, betweenness centrality, and closeness centrality in CytoScape v.3.6.0 software, to mark hub genes (Shannon et al. 2003). Moreover, we collected 244 nodes for Parkinson's and 145 nodes for the midbrain tumor. Data analysis indicated that 26 hab genes were common between Parkinson's and the midbrain tumor via the VENN diagram. Hence, considering a medium confidence score, we designed a PPIs network based on common hub genes. In addition, enrichment analysis performed by KOBAS-i (Bu et al. 2021), WikiPathway (Slenter et al. 2022), Reactome (Gillespie et al. 2022), and DAVID (Sherman et al. 2022).

Molecular docking studies

Computational drug discovery was conducted based on small molecules and 3D structures. This study investigated the binding affinity of bioactive compounds over the cut-point protein DRD2 in the pathogenesis network. Hence, we browsed the UniProt (UniProt: the universal protein knowledgebase in 2021 2021) and Protein Data Bank (PDB) (Burley et al. 2021) databases. We selected the best protein crystallography of DRD2 (PDB-ID: 6CM4) based on the X-ray method. Protein stricture preparation and optimization were conducted in Chimera 1.8.1 software (Pettersen et al. 2004). On the other hand, we scanned the previous studies to identify important bioactive compounds in Ginseng, Matthiola incana, Turmeric, and Green-Tea. We found 11 bioactive compounds in these botanical medicines, including Gallic acid, Caffeine, Epigallocatechin-3-gallate, Ferulic acid, Caffeic acid, Curcumin, Chlorogenic acid, Ginsenosides, Quercetin, Rutin, and Ellagic acid. In addition, 3D structures of bioactive compounds were collected from the PubChem database (Kim et al. 2021). We provided a small molecules library for selected compounds by OpenBable software. Molecular docking was analyzed by PyRx software to define the Vina search box dimension (X: 80.7253, Y: 61.1533, and Z: 87.7188). This chemoinformatics study considered binding affinity < -5 kcal/mol and RMSD < 2 as the acceptable threshold (Dallakyan and Olson 2015; Mosaddeghi et al. 2021).

Ethical code

This study followed the ARRIVE guidelines and was approved and performed according to protocols of laboratory animals by the Ethics Committee of Islamic Azad University, Khorasgan Branch (IR.IAU.KHUISF.REC.1400.335).

Grouping and treatment

The current study provided 56 male Wistar rats (230 ± 20 g) from Pasteur Institute (Karaj, Iran). The rats were housed under a standard condition (12 h dark/12 h light cycle, humidity: 60% ± 5%, and temperature: 23 ± 4 °C). The animals were access to food and water freely.

In this study, rats were distributed into eight groups:

(I) The rats considered as control group without treatment (we called this group as Control), (II) The rats induced the brain tumor model by injections of C6 glioma cell line and stereotaxic method (we called this group as model), (III) The rats induced the brain tumor model and treated swimming training (we called this groups as model + Exe), (IV) The rats induced the brain tumor model and treated Nanoliposomes (we called this groups as model + Lipo), (V) The rats induced the brain tumor model and treated combined supplement containing four plant extracts (we called this groups as model + Extract), (VI) The rats induced the brain tumor model and treated Nanoliposomes enriched combined supplement (we called this groups as model + Lipo-Extract), (VII) The rats induced the brain tumor model and treated combined supplement along with swimming training (we called this groups as model + Extract-Exe), (VIII) The rats induced the brain tumor model and treated Nanoliposomes enriched combined supplement along with swimming training (we called this groups as model + Lipo-Extract + Exe).

Inducing a brain tumor model

C6 glioma cell line (5 × 10⁵ in 10 μl cell suspension) injected in the substantia nigra (SN) region (deep ventral (DV): -7.6 mm, anterior–posterior (AP): -5.5 mm, medial–lateral (ML): − 2.4 mm; anterior–posterior (AP): − 5.2 mm; medial–lateral (ML): − 1.9 mm; deep ventral (DV): -8.2 mm) of male-Wistar rats based on the protocols and stereotaxic method (Bolcaen et al. 2021).

To inject the C6 glioma cell line, we used ketamine/xylazine (3:5) to anesthetize. After surgery, the animals were monitored (body temperature, wound healing, and behavior). Post-inoculation, animals were maintained in separate enclosures.

Administration of combined supplement

For groups treated with combined supplements, nanoliposomes and nanoliposomes enriched with combined supplements (final concentration 1 mM) (100 mg/kg/day for 6 weeks) administered by gavage.

Swimming training protocol

Rats performed swimming exercise training for 6 weeks. Rats swam to adapt to the environment and the swimming pool (10 min 3 days a week). Then the rats were subjected to swimming training 3 days a week (30 min. The intensity of the training protocol was moderated based on the literature review and World Health Organization (WHO) guidelines (Chen and Chang 2010; Boracı et al. 2020).

Preparation of nanoliposomes

Based on the literature review, we provided the safe dosage of herbal medicine extract (50 mg/kg Matthiola incana, 50 mg/kg Green-Tea, 50 mg/kg Turmeric, and 100 mg/kg Ginseng for oral administration/day). 100 gr of Green-Tea, Ginseng, Matthiola incana, and Turmeric freshly purchased from the herbal medicine center to prepare the extract. Firstly, herbs were washed with distilled water and placed in the oven at 40 °C for 7 days to dry. For the Mixing extraction, dried herbs were powdered by Electric mills, and a hydroalcoholic solution (70% ethanol and 30% water) was supplied at 40 °C for 48 h. Next, a solution was liquidized and filtered by a Buchner funnel. After that, it was placed in a distillation system with a rotary rotator at 50 rpm for 45–50 min and 50 °C. Herbal extract was dissolved in sterile distilled water to obtain a final concentration of 1 mM. After dissolving one gram of phosphatidyl colin in ethanol, a thin lipid film on the flask's wall was made by a rotavapor. The organic solvent was entirely evaporated under a vacuum, followed by adding distilled water and stirring the suspension for two hours under liquid nitrogen. The samples were then sonicated at 40 kHz and 40% of maximum power for 360 s to provide a homogenous solution (1 s On, 1 s Off). Samples of liposomes were preserved in glass bottles at 4 °C in the dark.

Measurements of liposome size and electrophoretic mobility

The liposome's mean size, electrophoretic mobility, and zeta potential were assessed based on DLS and empirical dilutions on Malvern Zetasizer Nano ZS (Malvern Instruments Ltd, UK, ZEN3600) and DTS Nano software v.6.12. Liposomes were prepared in triplicate (n = 3) after three runs, and the results were provided as vesicle size, polydispersity index, and zeta potential (Maherani et al. 2012). Morphological features of nanoliposome particles were achieved by scanning electron microscope (SEM, LEO 1430VP, Germany and UK).

Behavioral tests

Beam test

The Beam test was used to measure motor coordination. Rats were placed in the corner and allowed to walk to the other through the narrow at least three times. The narrow rod is 1–3 cm wide and was placed on a platform and rat cage. This test recorded the number of foot slips and the time required to cross the bar. Notably, the healthy rats could pass the bar without additional support, but rats with impaired forelimb or hind limb function were often placed on the restraint as support.

Sciatic functional index (SFI)

The Sciatic Functional Index (SFI) is a practical method for measuring the recovery of sciatic nerve function in rats after various experimental lesions and therapies. This SFI is made of wood (50 cm high, 7 cm wide, and 60 cm long). Using the methodology proposed by Bain et al., the following sciatic functional index was determined:

$$\text{SFI}=-38.3(\text{EPL}-\text{NPL})/(\text{NPL})+109.5(\text{ETS}-\text{NTS})/(\text{NTS})+13.3(\text{EIT}-\text{NIT})/(\text{NIT})-8.8$$

PL represented the print length, TS represented the toe spread, ITS represented the intermediate toe spread, E represented the experimental model, and N represented the normal model. A score of 100 demonstrated total disability, whereas a score of 0 indicated normal function.

Immunohistochemical assessment

Immunohistochemical (IHC) was performed to detect the D1 protein in the brain tissue. First, the hydrated tissue samples were placed in a TBS buffer and heated at 70 °C for 20 min for antigen retrieval. Then, the specimens were permeabilized for 20 min with a solution of 0.3 percent Triton x-100. The primary antibody DRD1 protein and (rabbit monoclonal, Recombinant Anti-Dopamine Receptor D1 antibody [EPR24102-105], ab279713, Abcam, MA, the USA at 1/1000 dilution) (Li et al. 2022) and TH protein (Rabbit polyclonal antibody to TH, ab137721, Abcam, MA, USA at 1/100 dilution) (Pyon et al. 2019) was incorporated after samples were cleaned in PBS and blocked with 10% goat serum for 40 min. In the following step, the samples were thoroughly washed with PBS and then incubated for 60 min at 37 °C in the dark with a Peroxidase-conjugated secondary antibody solution (goat Anti-Rabbit IgG, Abcam, ab97051, MA, USA at 1:50,000 diluted in PBS) (Zhang et al. 2022). Chromogen and substrate for the peroxidase enzyme coupled to goat Anti-Rabbit IgG were used following manufacturers' instructions. Counterstaining was completed using hematoxylin. A light microscope was used to examine the materials.

Quantitative gene expression assessment

Total RNA was isolated using a Trizol reagent in accordance with the manufacturer's instructions (Thermo Scientific, USA). Sample purity and concentration of RNA were determined by NanoDrop analysis using the OD 260/280 nm (Thermo Scientific, Waltham, MA, USA). Moreover, cDNA synthesis was conducted based on the instructions (TaKaRa, Kusatsu, Shiga Prefecture, Japan). Furthermore, SYBR Green dye and quantitative real-time PCR evaluated gene expression (Rotor-Gene 6000 instrument, Corbett Life Science, Mortlake, Australia). We utilized GAPDH as a reference gene for normalization. The 2^−ΔΔCt was used to determine mRNA expression. The primer list is indicated in Table 1.

Table 1.

The primers sequence

Gene	Sequence of primers	Size (bp)
TRKB	Forward: 5ʹ-TTATGCTTGCTGGTCTTGGGCTTC-3ʹ	146
	Reverse: 5ʹ-TCTGGGTCAATGCTGTTAGGTTCC-3ʹ
GABArα1	Forward: 5ʹ-CTCCTACAGCAACCAGCTATACCC-3ʹ	114
	Reverse: 5ʹ-GCGGTTTTGTCTCAGGCTTGAC-3ʹ
DRD2	Forward: 5ʹ-TGGAGGTGGTGGGTGAGTGG-3ʹ	228
	Reverse: 5ʹ-GTGGGGAGGAGATGGTGAAGGA-3ʹ
GAPDH	Forward: 5ʹ-AGGTCGGTGTGAACGGATTTG-3ʹ	123
	Reverse: 5ʹ-TGTAGACCATGTAGTTGAGGTCA-3ʹ

Crystal violet staining

The slides were deparaffinized, and the hydration steps were performed. The samples are placed in a 0.1% crystal violet (Sigma- C5042) solution for 10 min at a temperature of 50–37 °C. Then rinse with distilled water and soak in 95% alcohol for 2–3 min. The slides are washed and dried.

Statistical analyses

The sample size was calculated based on alpha level 0.05 and 90% power in this study. GraphPad Prism was used to analyze variance (Version 9 Graph Pad Software Inc., La Jolla, CA). The Kolmogorov–Smirnov test was used to estimate the normalizing distribution. In addition, data analysis was performed based on the One-way ANOVA with repeated measures followed by Tukey's post hoc. Moreover, we analyzed data based on the mean ± standard deviation (SD).

Result

Bioinformatics

Analysis of midbrain microarray samples in Parkinson's status showed 4094 genes with significant differential expression (P value < 0.05). Moreover, we indicated that 1504 genes were upregulated and 1228 genes were downregulated (logFC ± 0.5 thresholds). The heatmap visualization diagram presented considerable differential gene expression with a P value < 0.001 (Fig. 1A).

Fig. 1.

A The Heat Map plot tool in the R statistical language analysis demonstrated significant differences in gene expression (P value < 0.001 thresholds) in Parkinson's patients compared to healthy controls. B Based on the network's parameters, nodes with the most significant degree and betweenness centrality were ranked in the protein–protein interactions network of genes implicated in Parkinson's pathogenesis. C There are significant differences in gene expressions with P value < 0.001 scores between patients with midbrain cancer and healthy controls, as shown by the Heat Map plot tool in R statistical language analysis. D The protein–protein interactions network of genes implicated in the development of midbrain cancer was organized according to the network's properties

These data indicated that 244 hub genes (degree = 22, betweenness centrality = 0.001, and closeness centrality = 0.3) were associated with Parkinson's pathogenesis (Fig. 1B). Moreover, an analysis of the microarray dataset of the midbrain region's cancer marked 20,784 genes with significant differential expression compared to healthy midbrain samples (P value < 0.05). The heatmap visualization showed significant differential gene expression with P value < 0.001 (Fig. 1C). Among significant genes with differential expression in brain cancer, 583 were overexpressed, and 569 were downregulated (logFC threshold ± 4). Analysis of the protein–protein interactions (PPIs) marked 145 hub genes (degree = 20, betweenness centrality = 0.002, and closeness centrality = 0.2) associated with midbrain tumor pathogenesis (Fig. 1D).

VENN diagram showed 26 collaborative hab genes between Parkinson's and midbrain tumor, including Gabra1, Mybl2, Gabrg2, Calb1, Kif23, Exo1, Cd44, Brca1, Camk2a, Ntrk2, Th, Drd2, Slc32a1, Drd1a, Cacna1a, Vim, Cdk2, Aspm, Aurkb, Grin2b, Ptk2b, Gja1, Nes, Stat1, Prkcg, Skp2 (Fig. 2A and B). Enrichment analysis revealed that hub genes are involved in the gabaergic synapse, dopaminergic synapse, drug addiction, Parkinson's diseases, cell cycle, foxo signaling pathway, and p53 signaling pathways (P value < 0.05) (Fig. 2C). On the other hand, data analysis and data mining indicated that these genes had critical roles in the dopaminergic synapse pathway. Furthermore, the dopaminergic pathway is an effective signaling pathway in midbrain damage (Fig. 2D). Enrichment in WikiPathway human 2021 and Reactome 2016 database confirm these outcomes based on literature and evidence (Fig. 2E and F). The bioinformatics analysis and enrichment results selected Drd1a, Drd2, Gabra1, Gabrg2, and Ntrk2 as essential genes.

Fig. 2.

A The Venn diagram tool organized the list of the collaborative genes with differential expression significantly in the brain tumor and Parkinson's. B The protein–protein interaction network of common genes is designed based on modularity and eigenvector centrality characteristics. Nodes in this network with the same color suggest that the proteins are involved in the same pathways, indicating a strong association between proteins in the pathogenesis of midbrain cancer with Parkinson's hallmark. C Enrichment Outcomes from the analysis of common hub genes in the KOBAS-intelligent server suggest that hub genes are engaged in the dopaminergic synapse, the cell cycle, and the FoxO signaling pathway with a significance score of P value < 0.05, which could suggest that these molecular signaling pathways are influential pathways in glioblastoma and Parkinson's disease pathogenesis network as the significant molecular signaling pathways related to hub genes. D Following data mining of common hub genes in the DAVID database exhibited significant positions of the dopaminergic synapse pathway in the pathogenesis of brain tumors and Parkinson's. E and F Enrichment analysis in WikiPathway human 2021 and Reactome 2016 database highlighted the significant association between hub genes and molecular signaling pathways in midbrain cancer pathogenesis and Parkinson's status

Chemoinformatic

Based on PPIs network construction, we found that Drd2 could be a potent candidate for drug design and therapeutic approach by activating the D1/D2 dopamine receptor signaling pathway. Hence, we browsed the PDB server to choose the closest and best 3D structure of DRD2 protein. On the other hand, based on the literature review, we provided a 3D structure library including 11 bioactive compounds in Ginseng, Matthiola incana, Turmeric, and Green-Tea as effective botanical medicines. The library included Gallic acid, Caffeine, Epigallocatechin-3-gallate, Ferulic acid, Caffeic acid, Curcumin, Chlorogenic acid, Ginsenosides, Quercetin, Rutin, and Ellagic acid. We estimated the binding affinity between DRD2 protein and ligands library based on the molecular docking method and showed that all small molecules with a suitable binding affinity (binding affinity < -5 kcal/mol, and RMSD score < 2) bound to the active site of DRD2 protein (Table 2). The present chemoinformatic survey supplied the highest binding affinity for Quercetin, Ginsenosides, Curcumin, and Rutin, respectively (Fig. 3A–D).

Table 2.

Molecular Docking; score prediction of binding affinity between macromolecule DRD2 and bioactive molecules library

No	PubChem ID	Commercial name	Binding affinity (< − 5kdal/mol)	RMSD
1	Conformer3D_CID_370	Gallic acid	− 5.7	< 2
2	Conformer3D_CID_2519	Caffeine	− 5.8	< 2
3	Conformer3D_CID_65064	Epigallocatechin-3-gallate	− 8.2	< 2
4	Conformer3D_CID_445858	Ferulic acid	− 6.8	< 2
5	Conformer3D_CID_689043	Caffeic acid	− 7.3	< 2
6	Conformer3D_CID_969516	Curcumin	− 9.2	< 2
7	Conformer3D_CID_1794427	Chlorogenic acid	− 8.4	< 2
8	Conformer3D_CID_3086007	Ginsenosides	− 9.3	< 2
9	Conformer3D_CID_5280343	Quercetin	− 9.7	< 2
10	Conformer3D_CID_5280805	Rutin	− 9.0	< 2
11	Conformer3D_CID_5281855	Ellagic acid	− 8.6	< 2

Fig. 3.

A–D Molecular docking approach estimating predicted that all ligands in the library with suitable binding affinity scores bound to the active sites of the DRD2 protein. According to chemoinformatics prediction, Quercetin, Ginsenosides, Curcumin, and Rutin provided the most substantial binding affinity

Nanoliposomes characteristics

We measured the particle sizes of nanoliposome preparations after sonication. The nanoliposome particle size at 4 °C and 37 °C exhibited no significant instability between days 0 and 30. Moreover, the potential zeta test revealed that the nanoparticles have an electric charge of − 3.39 microvolts (Fig. 4A). DLS results indicated nanoparticle sizes from 60–600 nm; the most significant amount of nanoparticles generated was 100 nm (Fig. 4B). As shown by the scale bar in the DLS test, the size of the nanoparticles generated could be validated by SEM microscopic analysis results and their globular form and high density (Fig. 4C).

Fig. 4.

A According to the potential zeta test, the generated nanoparticles have an electric charge of − 3.39 microvolts. The nanocoatings were therefore found to be stable and negatively charged. B DLS results found nanoparticle sizes ranged from 60 to 600 nm, with most particles existing 100 nm in dimension. C The nanoparticles' size, spherical form, and density were visualized by SEM microscopic analysis results

Nanoliposomes enriched combined supplement along with swimming training improved the expression level of the D1 protein.

We found that the level of D1 protein significantly decreased in the model group compared with the control group (Fig. 5, P value < 0.0001). Moreover, based on our data, the protein level of the D1 was increased in model + Extract (Fig. 5, P < 0.0001) and model + Exe groups (Fig. 5, P = 0.002) compared with the model group. In the model + Lipo-Extract group, the D1 protein level was elevated compared with the model + Extract group (Fig. 5, P value < 0.0001), model + Exe group (Fig. 5, P = 0.0006), and model group (Fig. 5, P value < 0.0001). Notably, we found that the level of D1 protein in the model + Extract group was more than in the model + Exe group (Fig. 5, P = 0.0490). It should be noted that there was no difference between the model group and model + Lipo (Fig. 5, P = 0.9316). In addition, the expression level of the D1 protein was enhanced in model + Extract-Exe (Fig. 5, P value < 0.0001) and model + Lipo-Extract + Exe compared (Fig. 5, P value < 0.0001) with the model group. Notably, the model + Lipo-Extract + Exe group significantly increased the D1 protein compared with other groups (Fig. 5, P value < 0.0001). Moreover, we found that the level of D1 protein was enhanced in the model + Lipo-Extract + Exe group in comparison with model + Extract-Exe (Fig. 5, P value < 0.0001) and model + Lipo-Extract (Fig. 5, P value = 0.0009). Hence, nanoliposomes enriched combined supplement along with swimming training (model + Lipo-Extract + Exe) had more effect than the other interactions.

Fig. 5.

The effect of nanoliposomes enriched combined supplement along with swimming training on the expression level of the D1 protein. D1 protein level significantly reduced in Model group. TH protein level was improved in brain tissues of nanoliposomes enriched combined supplement along with swimming training. Data expressed as mean ± SD. + indicates statistically significant difference with Control group, * indicates statistically significant difference with Model group, @ indicates statistically significant difference with Model + Exe group, # indicates statistically significant difference with Model + Lipo group, & indicates statistically significant difference with Model + Extract group, ! indicates statistically significant difference with Model + Lipo-Extract group, ^ indicates statistically significant difference with Model + Extract + Exe group, $ indicates statistically significant difference with Model + Lipo-Extract + Exe. Magnification 100um

The tyrosine hydroxylase protein regulated by nanoliposomes enriched combined supplement and swimming training.

The Tyrosine hydroxylase protein (TH) was diminished in the model group compared with the Control group (Fig. 6, P value < 0.0001). Furthermore, we found that swimming training enhanced the protein level of the TH in rats induced in the brain tumor model (model + Exe group) compared with the model group (Fig. 6, P value = 0.0116). The data indicated that TH protein increased in the model + Extract group compared with the model group (Fig. 6, P value < 0.0001). Additionally, the protein level of TH was predominantly raised in a model + Lipo-Extract group compared with the model group (Fig. 6, P value < 0.0001). Besides, the protein level of the TH was elevated in the model + Extract-Exe group (Fig. 6, P value < 0.0001) and model + Lipo-Extract + Exe group (Fig. 6, P value < 0.0001) compared with the model group. The data indicated that the level of TH protein was increased in the model + Extract group compared with the model + Exe group (Fig. 6, P = 0.0323). Moreover, there was no difference between the model group and model + Lipo (Fig. 6, P = 0.9632).

Fig. 6.

Nanoliposomes enriched combined supplement along with swimming training improved the expression level of the TH protein. TH protein level significantly reduced in Model group. TH protein level was increased in brain tissues of nanoliposomes enriched combined supplement along with swimming training. Data expressed as mean ± SD. + indicates statistically significant difference with Control group, * indicates statistically significant difference with Model group, @ indicates statistically significant difference with Model + Exe group, # indicates statistically significant difference with Model + Lipo group, & indicates statistically significant difference with Model + Extract group, ! indicates statistically significant difference with Model + Lipo-Extract group, ^ indicates statistically significant difference with Model + Extract + Exe group, $ indicates statistically significant difference with Model + Lipo-Extract + Exe. Magnification 100um

Furthermore, the level of TH protein was improved in the model + Lipo-Extract + Exe group compared with the model + Extract-Exe (Fig. 6, P value < 0.0001) and model + Lipo-Extract group (Fig. 6, P value = 0.0079).

The relative expression of the GABArα1/TRKB/DRD2 network was enhanced by nanoliposomes enriched combined supplement along with swimming training

We found that the relative expressions of the GABArα1 (Fig. 7 A, P value < 0.0001), TRKB (Fig. 7B, P value < 0.0001), and DRD2 (Fig. 7C, P value < 0.0001) were decreased in the model group compared with the Control group. Moreover, the expression levels of the GABArα1, TRKB, and DRD2 were increased in the model + Exe group (Fig. 7A–C, P value = 0.0018, P value < 0.0001, P value = 0.0012) and model + Extract group (Fig. 7A–C, P value < 0.0001, P value < 0.0001, P value < 0.0001) compared with the model group. Also, the expression levels of the GABArα1 (Fig. 7A, P value = 0.0002), TRKB (Fig. 7B, P value < 0.0001), and DRD2 (Fig. 7C, P value < 0.0001) were enhanced in model + Extract group compared with model + Exe group. Moreover, there were no differences between the model group and model + Lipo (Fig. 7, 5P value > 0.9999, P value > 0.9999, P value = 0.9997). Furthermore, we demonstrated that the expression levels of the GABArα1 (Fig. 7A, P value < 0.0001), TRKB (Fig. 7B, P value < 0.0001), and DRD2 (Fig. 7C, P value < 0.0001) in the model + Lipo-Extract group were raised in comparison with the model group. On the other hand, the expression levels of the GABArα1, TRKB, and DRD2 were elevated in model + Extract-Exe (Fig. 7A–C, P value < 0.0001, P value = 0.0003, P value < 0.0001) and model + Lipo-Extract + Exe (Fig. 7A–C, P value < 0.0001, P value < 0.0001, P value < 0.0001) compared with the model group. Our data indicated that the relative expressions of the GABArα1/TRKB/DRD2 network were enhanced by nanoliposomes enriched combined supplement and swimming training (model + Lipo-Extract + Exe) and had more effect than the other interactions.

Fig. 7.

The effect of nanoliposomes enriched combined supplement along with swimming training on the relative expression of the GABArα1/TRKB/DRD2 network. The expression GABArα1/TRKB/DRD2 genes significantly decreased in Model group. GABArα1/TRKB/DRD2 transcript levels were amplified in brain tissues of nanoliposomes enriched combined supplement along with swimming training. Data expressed as mean ± SD. + indicates statistically significant difference with Control group, * indicates statistically significant difference with Model group, @ indicates statistically significant difference with Model + Exe group, # indicates statistically significant difference with Model + Lipo group, & indicates statistically significant difference with Model + Extract group, ! indicates statistically significant difference with Model + Lipo-Extract group, ^ indicates statistically significant difference with Model + Extract + Exe group, $ indicates statistically significant difference with Model + Lipo-Extract + Exe

Nanoliposomes enriched combined supplement and swimming training modified the population of neuro dopaminergic cells

The population of neuro dopaminergic cells in brain tissue was analyzed by Cresyl violet staining. The data indicated that the model group had the darkest cell number vs. the control group (Fig. 8, P value < 0.0001). Moreover, in model + Exe (Fig. 8, P value = 0.0002), model + Extract (Fig. 8, P value < 0.0001), model + Lipo-Extract (Fig. 8, P value < 0.0001), model + Extract-Exe groups (Fig. 8, P value < 0.0001) population of neuro dopaminergic cells in brain tissue were improved compared with the model group. Additionally, the population of neuro dopaminergic cells was improved in the model + Lipo-Extract + Exe group compared with the model + Extract-Exe (Fig. 6, P value < 0.0001) and model + Lipo-Extract (Fig. 6, P value = 0.0040). Furthermore, we found that nanoliposomes enriched with combined supplement and swimming training (model + Lipo-Extract + Exe) modified the population of neuro dopaminergic cells and had more effect than the other interactions (Fig. 8, P value < 0.0001).

Fig. 8.

Nanoliposomes enriched combined supplement, and swimming training modified the population of neuro dopaminergic cells. Samples were stained with Cresyl violet. The percentage of the dopaminergic cell calculated by image J software. The percentage of the dopaminergic cell significantly decreased in Model group. The population of neuro dopaminergic cells was enhanced in brain tissues of nanoliposomes enriched combined supplement along with swimming training. Data expressed as mean ± SD. + indicates statistically significant difference with Control group, * indicates statistically significant difference with Model group, @ indicates statistically significant difference with Model + Exe group, # indicates statistically significant difference with Model + Lipo group, & indicates statistically significant difference with Model + Extract group, ! indicates statistically significant difference with Model + Lipo-Extract group, ^ indicates statistically significant difference with Model + Extract + Exe group, $ indicates statistically significant difference with Model + Lipo-Extract + Exe. Magnification 20 um

Nerve function was improved by swimming training along with nanoliposomes enriched combined supplement

The SFI and beam test indicated that the nerve function and motor-sensory of the model group were reduced compared with the control group (Table 3, P value = 0.0144, P value < 0.0001). Moreover, the SFI and beam scores were improved in the Model + Exe and Model + Extract groups compared with the model group (Table 3, P. value < 0.0001, P. value = 0.0003). In addition, we found that the scores of the SFI and beam tests significantly ameliorated in the Model + Lipo-Extract group (Table 3, P value < 0.0001) and Model + Lipo-Extract + Exe group (Table 3, P value < 0.0001) compared with the model group. The behavior tests demonstrated that swimming training and nanoliposomes enriched combined supplement improved nerve function.

Table 3.

The behavior tests

	Sciatic functional index (SFI)	P value	Beam test	P value
Control	− 33.67 ± 10.6		7.667 ± 0.5774
Model	− 52 ± 8.544a	0.0144	4.833 ± 0.2887	< 0.0001
Model + Lipo	− 49 ± 7.937b	< 0.0001	6.25 ± 0.9899	0.0003
Model + Exe	− 39 ± 5.196b	0.7830	4.8 ± 0.3536	0.6101
Model + Extract	− 42.67 ± 8.145b	0.0004	5.333 ± 0.2887	0.0040
Model + Lipo-Extract	− 36.33 ± 3.055b	< 0.0001	6.333 ± 0.7638	< 0.0001
Model + Extract-Exe	− 35.33 ± 3.786b	< 0.0001	7.75 ± 0.3536	< 0.0001
Model + Lipo-Extract + Exe	− 39.67 ± 8.737b	0.0001	8.667 ± 0.7638	< 0.0001

^aIndicates a statistically significant difference with the Control group

^bIndicates a statistically significant difference with the Model group

Discussion

We pointed to DRD1a, DRD2, TRKB, and GABA as hub genes in activating the D1/D2 dopaminergic signaling pathway in the pathogenesis of Glioblastoma with Parkinson's status based on bioinformatic analysis. Enrichment demonstrated a significant link between common genes and GABAergic synapses, dopaminergic synapses, cell cycle, FoxO signaling pathways, and P53 signaling pathways (P value < 0.05). Text mining has indicated that molecular pathways could promise new strategies for preventing the pathogenesis, progression, and treatment of the hallmarks Glioblastoma and Parkinson's. Dopamine (DA) is the classic slow neurotransmitter and regulates several processes in the mammalian brain, including locomotion, motivation and reward, learning and memory, and endocrine regulation.

When dopamine is released from presynaptic axonal terminals, it binds to at least five different receptor subtypes in the central nervous system (CNS), which have been categorized into two groups: the D1-like receptors (D1Rs), which include the D1 and D5 receptors and are positively coupled to adenylyl cyclase and cAMP production, and the D2-like receptors (D2Rs), which include the D2, D3, and D4. DA modulates neuronal activity, synaptic plasticity, and behavior via various cAMP-dependent and -independent processes. As the primary autoreceptor of the dopaminergic system, D2Rs are localized presynaptically and control DA biosynthesis and release (Beaulieu and Gainetdinov 2011). The degradation of DA neurons in the substantia nigra is a significant neuropathological hallmark of Parkinson's disease, and dysfunctions in DA neuron circuits have been linked to several neuropsychiatric diseases (Wang et al. 2020). Ample evidence has revealed that α-synuclein is highly expressed in the presynaptic cleft of nerve cells and has a vital role in neurodegenerative diseases.

The pharmaceutical method is currently the primary therapeutic strategy for PD patients to reduce or manage motor symptoms. The therapy is often targeted to boost dopamine bioavailability, either by replenishing the dopamine precursors or by preventing the breakdown of Dopamine (Wang et al. 2020). During the initial stages of therapy, levodopa (also known as L-dopa), the precursor of dopamine, is administered as the mainstay. In contrast to Dopamine, Levodopa may enter the brain and be converted into dopamine there. However, levodopa conversion in the peripheral nervous system might cause side effects. Clinical investigations have demonstrated that mDA neurons produced from human fetal grafts could reinnervate the striatum, improve symptoms in patients with PD, and restore dopamine release (Parmar et al. 2020). Seamus P. Caragher et al. examined the influences of monoamine dopamine on Glioblastoma cells using human patient-derived xenograft lines and established that dopamine receptor 2 (DRD2) was overexpressed in the glioma-initiating cell population. Furthermore, the long-cycle activation of DRD2 amplified the sphere-forming potential of glioblastoma cells and tumor engraftment efficiency in both sex mice. DRD2 signaling could trigger the hypoxic response and functionally affects metabolism, according to mechanistic research. This research indicated that Glioblastoma cells produced and released dopamine. Thus, dopamine signaling might be a therapeutic target (Caragher et al. 2019). Neurotransmitters and brain-specific messengers as signaling molecules can influence molecular pathways and cellular activities in the central nervous system. Moreover, these cells' proliferation, quiescence, and differentiation status are also affected by neurotransmitters (Caragher et al. 2018).

More research is needed to fully understand how exercise might help people live longer and better lives (Damm et al. 2020). In addition, Shana M. Augustin et al. revealed that D2 dopamine receptors (D2Rs) could play a crucial role in motion in the striatum (Augustin et al. 2020).

On the other hand, Hines et al. have proven that dysregulation of GABAAR's clusters as interneuron transmitters, which include the two subunits might lead to an uncontrolled electrical disturbance in the brain and death, as well as anxiety-like phenotypic features in the Gabra2-1 animal model (Hines et al. 2018; Contreras et al. 2019). Johan Pallud et al. have highlighted that glutamate and GABA signaling are involved in oncogenesis, and unbalanced chloride homeostasis is associated with GABA depolarizing in the pyramidal cells. These indicators induce an imbalance between synaptic excitatory and underlying epileptic discharges in tumor patients (Pallud et al. 2014). Moreover, previous studies have indicated that the FOXO signaling pathway is critical in pathogenesis, tumorigenesis, cell cycle, cell proliferation, apoptosis, autophagy, protein expression, oxidative stress, and metabolism. Mohd Farhan et al. proposed that targeting the FOXO pathway's molecular signalings could be a potential strategy for preventing and treating cancers (Farhan et al. 2017; Richard and Jia-Hao 2020; Ciechomska et al. 2020).

In the current study, we provided the effect of nanoliposome enriched with a combined extract of Ginseng, Matthiola incana, Turmeric, and Green-Tea in ameliorating brain tumor rat models with Parkinson's symptoms. Ginseng has a comprehensive scope of therapeutic approaches due to its chemoprotective mechanism. Growing studies have indicated that Ginsenosides could be a medicinal effect on the immune system, neurological function, cardiovascular protection, anti-toxicity effect, and signal transduction process initiator (Palliyaguru et al. 2016; Lu et al. 2009; Ratan et al. 2021; Liu et al. 2020; Hajibabaie et al. 2020). Jian-Ming Lü et al. displayed the effects of the Ginseng compounds, especially ginsenosides, molecular mechanism, signal transduction pathways, vascular system, and ligand-receptor interactions (Lu et al. 2009). Nah et al. revealed the positive association of bioactive ginsenosides with central/peripheral nervous system function (Nah et al. 2007).

Moreover, evidence indicated that ginseng could provide neuroprotective properties as a therapeutic strategy in murine models of PD (Chen et al. 2005; Luo et al. 2011). Neurite attrition and shrinkage caused by MPP treatment of primary dopaminergic cultures are suppressed by Ginsenoside bioactive compounds (Rb1 and Rg1) (Luo et al. 2011). Furthermore, oral administration of the ginseng supplement effectively ameliorated TH-positive neurodegeneration in the substantia nigra and enhanced the sensory-motor function in MPTP-treated mouse models (Van Kampen et al. 2014; Rokot et al. 2016).

Growing evidence has revealed that consuming green tea, which has an antioxidative and antitumor effect, could prevent cancer development (Vishvakarma 2014; Basnet and Skalko-Basnet 2011). Furthermore, Joanna Kochman and colleagues have indicated that Green Tea could decrease inflammation and oxidative stress. Based on these data, Green Tea is considered the best beverage with unique chemical bioactive agents, anti-inflammatory substances, health-developer properties, and high redox efficiency in medicinal herbal classification (Kochman et al. 2020).

One of the crucial stages in avoiding PD pathophysiology is the inhibition of α-synuclein fibril formation and aggregation. Treating catechin-rich tea polyphenol extract restored motor deficits, recovered TH and dopamine concentration, and decreased the number of α-synuclein fibrils (Chen et al. 2015). Green Tea and epigallocatechin-3-gallate mitigated dopamine through the modulation of antioxidants superoxide dismutase (SOD) and catalase (CAT) in male C57/BL mice. In addition, Green Tea might avoid the death of dopaminergic neurons by blocking TH reduction (Levites et al. 2001; Sergi 2022; Malar et al. 2020). Also, Catechins could provide neuroprotection by elevating striatal protein kinase C-α (PKC-α) (Kujawska and Jodynis-Liebert 2018; Malar et al. 2020). Zhou et al. have shown that epigallocatechin-3-gallate modified peripheral immune reaction by lowering the CD4 + /CD8 + ratio and downregulating the TNF-α, IL-6 expression levels in C57BL/6 J mice model of PD (Zhou et al. 2018). These results were verified by Tseng and colleagues' research (Tseng et al. 2020).

Furthermore, Lee et al. have demonstrated that catechin intraperitoneal injection modulated the noradrenergic axis, leading to decreased immobility duration in a forcible swimming assessment and improved open-arm exploration in the plus-maze test (Lee et al. 2013).

Based on traditional Ayurvedic medicine, Turmeric, especially the bioactive compound curcumin, has been used as an herbal drug to treat inflammation and tumor therapy. Curcumin is a potent anti-inflammatory and antioxidant component that could effectively decrease tumor size and inhibit the metastasis process. Growing evidence has supported that curcumin might have anti-inflammatory and antioxidative effects. Moreover, curcumin is recognized as an effective natural compound with several therapeutic prospects in the treatment of neurodegenerative, cardiovascular, cancers, inflammation, infection, rheumatism, allergic, cell differentiation, and cell degeneration (Ryskalin et al. 2020; Shahcheraghi et al. 2019; Gersey et al. 2017; Su et al. 2021; Mohammad Gholinia Sarpoli et al. 2022).

One of the effective herbals identified as a tumorigenesis inhibitor is Matthiola incana. This plant appears to inhibit the growth of cancer cells with its antioxidant compounds and phenolic agents that decrease oxidative stress and cytotoxic properties. However, the molecular mechanism of Matthiola incana is still not well known (Miceli et al. 2019, 2021; Taviano et al. 2020). Maria Fernanda Taviano et al. mentioned that Matthiola incana with its cytotoxic, enzyme inhibitory, and antioxidant properties were noticed as potential pharmacological applications in some metabolic disorders such as obesity or type 2 diabetes and different tumors, such as colorectal adenocarcinoma and breast cancer. Also, Matthiola incana extracts acted as an enzyme inhibitor in neuroprotection (Taviano et al. 2020).

Taviano and coworkers indicated that Matthiola incana extracts could have a neuroprotective inhibitor. Based on this study, Matthiola incana extracts might regulate the monoamine oxidase A, tyrosinase, and acetylcholinesterase in the central nervous system (Taviano et al. 2020). In the present study, based on a cheminformatics survey and molecular docking analysis, we predicted the binding affinity of natural bioactive compounds derived from Ginseng, Matthiola incana, Turmeric, and Green Tea herbal plants on the DRD2 protein as a potent cut-off point in the dopaminergic signaling pathway. Accordingly, we proposed that natural bioactive compounds derived from Ginseng, Matthiola incana, Turmeric, and Green Tea could effectively influence the protein–protein interaction network from Glioblastoma and Parkinson's symptoms.

Nevertheless, these phenomenal chemicals have limited applicability due to their poor solubility, stability, bioavailability, and gastrointestinal decomposition process (which lowers the biological activity of bioactive compounds). Recent scientific advances have indicated that the bioavailability of chemical compounds might improve Parkinson's symptoms by using novel drug delivery methods. The bioavailability and efficiency of synthetic derivatives and molecular analogs of compounds have been found to be enhanced by nanoformulation, encapsulation in vehicles, and combining with adjuvants, hence expanding the spectrum of uses of bioactive substances (Sohn et al. 2021). As a result, liposomes have been created as a flexible assembly to overcome the limitations of herbal extract consumption in treating disorders. Moreover, liposomes are efficient carriers for administering herbal extracts with enhanced therapeutic benefits (Sogut et al. 2021).

Based on the in-vivo analysis, the consumed extract supplements increased the D1 protein and GABArα1/TRKB/DRD2 network. In addition, consumed extract supplements modified the population of neuro dopaminergic cells. Our data indicated that the administration of the nanoliposome-enriched supplement was more effective and practical than the extract supplements. The results demonstrated that the nanoliposomes enriched combined supplements significantly improved the D1 protein, GABArα1/TRKB/DRD2 network. Also, nanoliposome-enriched combined supplements modified the population of neuro dopaminergic cells.

Interestingly, we evaluated the effect of exercise training on rats that had been induced with a brain tumor model. In recent years, swimming exercise has been found to effectively treat neurodegenerative conditions and Glioblastoma. Several studies indicated that swimming could enhance oxidative and physical capacity (Boracı et al. 2020; Contarteze et al. 2008; Haobam et al. 2005). Moreover, natural abilities like swimming make it a popular choice for rat models, as it is easy for rats to do (Souza et al. 2009). Exercise has been shown to preserve dopaminergic neurons in animal models of Parkinson's disease while also increasing levels of neurotrophic (Boracı et al. 2020; Garza et al. 2004). Based on our data, swimming training increased the protein level of the D1, relative expression of the GABArα1/TRKB/DRD2 network, as well as ameliorating the population of neuro dopaminergic.

Furthermore, Boracı and colleagues have revealed that swimming training elevated the count of striatal calretinin-positive interneurons, providing novel avenues for neuroprotection mechanisms. Boracı indicated that striatal caretinin positive interneurons could play a significant role in the neuroprotective exercise processes in Parkinson's (PD). Regular swimming training and nanoliposomes enriched combined supplement could modulate motor sensory signs in midbrain tumor rat models. However, this study had some limitations; for example, we did not measure the tumor size or evaluate the tumor signs by natural therapy. Moreover, this study lacked data about chemical therapy and compared it with complementary medicine. On the other hand, we require to assess this natural remedies strategy in clinical trials.

Conclusion

Overall, we found that swimming training and nanoliposome-enriched combined supplements are suggested as effective complementary medicine for motor impairment recovery induced by the midbrain tumor in the substantia nigra area. Hence, regular swimming training and natural medicines rich in polyphenolic bioactive components and antioxidative effects could modify and improve the dopamine receptors' function.

Supplementary Information

Below is the link to the electronic supplementary material.

Abbreviations

PD

Parkinson's disease

WHO

World Health Organization

TT

Tumor-treating

CAMs

Complementary and alternative medicine

GEO

Gene Expression Omnibus

BT model

Brain tumor model

SN

Substantia nigra

DV

Deep ventral

AP

Anterior–posterior

ML

Medial–lateral

SFI

Sciatic functional index

SD

Standard deviation

L-DOPA

L-3 and 4-dihydroxyphenylalanine

GABA

Gamma-aminobutyric acid

TrkB

Tropomyosin receptor kinase B

BDNF

Brain-derived neurotrophic factor

MAPK

Mitogen-activated protein kinase

PI3K

Phosphoinositide 3-kinase

PLC-γ1

Phospholipase C- gamma 1

PBT

Primary brain tumors

TH

Tyrosine hydroxylase

PPIs

Protein–protein interactions

PDB

Protein Data Bank

DRD1

Dopamine receptor D1

DRD2

Dopamine receptor D2

IHC

Immunohistochemical

MPTP

1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine

MPP + 

1-Methyl-4-phenylpyridium

DA

Dopamine

CNS

Central nervous system

D1Rs

D1-like receptors

iMSNs

Medium spiny neurons

DOPAC

3,4-Dihydroxyphenylacetic acid

MTT

3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide

Funding

The authors declare that no funds, grants, or other support were received during the preparation of this manuscript.

Availability of data and materials

The raw data and the rest of the materials remain in Islamic Azad University Isfahan (Khorasgan) Branch and are available upon request. The datasets analyzed during the current study are available in the Gene Expression Omnibus database repository, https://www.ncbi.nlm.nih.gov/geo/, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE52139PERSISTENT WEB, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE21942, https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi, STRING server (https://string-db.org/), Venn diagram (https://bioinfogp.cnb.csic.es/tools/venny/), http://www.pantherdb.org/, https://reactome.org/, https://maayanlab.cloud/Harmonizome/dataset/Biocarta+Pathways, https://www.wikipathways.org/index.php/WikiPathways, https://www.kegg.jp/, http://mirwalk.umm.uni-heidelberg.de/, https://www.mirnet.ca/, http://mirdb.org/, http://snf-515788.vm.okeanos.grnet.gr/, https://www.rcsb.org/, https://www.rcsb.org/structure/3AGZ, https://www.rcsb.org/structure/4J6G, https://pubchem.ncbi.nlm.nih.gov/compound/Genistein, https://pubchem.ncbi.nlm.nih.gov/compound/Kaempferol, https://pubchem.ncbi.nlm.nih.gov/compound/Formononetin, https://pubchem.ncbi.nlm.nih.gov/compound/Riboflavin, https://pubchem.ncbi.nlm.nih.gov/compound/Retinol, https://pubchem.ncbi.nlm.nih.gov/compound/Naringenin, https://pubchem.ncbi.nlm.nih.gov/compound/Isosakuranetin, https://pubchem.ncbi.nlm.nih.gov/compound/24Methylenecholesterol, https://pubchem.ncbi.nlm.nih.gov/compound/Hesperetin.

Declarations

Conflict of interest

None of the authors has any conflicts of interest to disclose.

Ethical approval for animal usage

This study followed the ARRIVE guidelines (https://arriveguidelines.org) as well as approved and performed according to protocols of laboratory animals by the Ethics Committee of Islamic Azad University, Khorasgan Branch (IR.IAU.KHUISF.REC.1400.335).

Consent for publication

All authors support submission to this journal.