The neuroprotective role of miR-221 in PD: Insights from in vitro and in vivo Studies
miR-221 plays a crucial role in promoting neuronal survival and protecting against oxidative stress in PD through several pathways (Figure 2 ). Autophagy is a process that helps maintain healthy cells by removing damaged or nonfunctional organelles and recycling their materials (Vargas, Hamasaki, Kawabata, Youle, & Yoshimori, 2023). It also destroys pathogens and removes long-lived proteins and aggregates to maintain cell homeostasis (Kocaturk & Gozuacik, 2018). There are three types of autophagy observed in mammalian cells: macroautophagy, chaperone-mediated autophagy, and microautophagy (Sato et al., 2016). This process is important for keeping cells healthy and has been linked to PD (Cerri & Blandini, 2019). When autophagy is not functioning properly, it can contribute to the development of PD (Cheung & Ip, 2009). Regulating autophagy is a proposed treatment strategy for PD (Rakowski, Porębski, & Grzelak, 2022). Another process called mitophagy, which helps remove damaged mitochondria from cells, may also play a role in protecting brain cells in PD (Goiran, Eldeeb, Zorca, & Fon, 2022).
A long non-coding RNA called LncRNA-SNHG1 is significantly increased in PD (H. Wang, Wang, Zhang, & Zhao, 2021). It has been shown to increase the formation of protein clumps called α-Syn aggregates and promote inflammation in the brain (L. Zhao & Wang, 2019b). mTOR is an enzyme that has a key role in controlling cellular growth, division, and viability (Ali et al., 2022; Barzegar Behrooz et al., 2022). It is a portion of two different protein complexes, mTORC1 and mTORC2, which have different targets and functions within the cell (Jhanwar-Uniyal, Dominguez, Mohan, Tobias, & Gandhi, 2022). Signals such as growth factors and nutrient levels activate mTORC1 which controls processes such as protein production, cell growth, and autophagy (Movahhed et al., 2022). mTORC2 is activated by growth factors and controls cell survival, metabolism, and the organization of the cell’s internal structure (Y. Sun et al., 2023). When mTOR signaling is not functioning properly, it can facilitate the development of various diseases, including cancer, diabetes, and neurological disorders (Russo, Citraro, Constanti, & De Sarro, 2012; Suhara, Baba, Shimada, Higa, & Matsui, 2017; Zou, Tao, Li, & Zhu, 2020). p27 is a protein that helps control cell division by inhibiting the activity of certain enzymes called cyclin-dependent kinases (Guiley et al., 2019). Functioning as a tumor suppressor, it prevents uncontrolled cell proliferation and has been linked to various types of cancer (Razavipour, Harikumar, & Slingerland, 2020). It has been demonstrated that p27 takes part in controlling autophagy and apoptosis in PD (Lv, Wang, Zhong, & Huang, 2020). LC3 II is a protein found on the membranes of autophagosomes, which are structures involved in autophagy (R. Wang et al., 2022). The level of LC3 II is related to the number of autophagosomes in a cell (Kabeya et al., 2000). A study by Qian and colleagues suggests that reducing levels of lncRNA-SNHG1 can increase autophagy and protect against cell death in PD by affecting the miR-221/222/p27/mTOR signaling pathway. They found that reducing lncRNA-SNHG1 levels increased the number of autophagosomes and reduced cell damage in a model of PD. This was due to decreased interaction between lncRNA-SNHG1 and miR-221/222, which resulted in lower levels of p27 and increased activity of the mTOR pathway. Blocking miR-221/222 reversed the effects of reducing lncRNA-SNHG1 levels on p27 and mTOR. These results suggest that targeting the interaction between lncRNA-SNHG1, miR-221/222, p27, and mTOR could be a potential treatment strategy for PD (Qian et al., 2019).
NPTX2 is a protein concentrated in certain brain tissues and cells affected by PD (Moran et al., 2008; Shao, Shan, Ru, & Ma, 2020). HOTAIR is a type of lncRNA that is associated with various diseases, including PD, thyroid cancer, and gallbladder cancer (H.-m. Li et al., 2017; M.-z. Ma et al., 2014; S. Wang, Zhang, Guo, Rong, & Liu, 2017). It can affect gene expression by interacting with proteins that modify DNA structure and with proteins that control gene activity (Spokoini-Stern et al., 2020). Lang and colleagues proposed that HOTAIR participated in promoting autophagy in certain brain cells affected by PD in mice. They found that HOTAIR levels were increased in the brains of mice with chemically induced PD and that reducing HOTAIR levels decreased autophagy and improved movement in these mice. They also discovered that HOTAIR affects the levels of the NPTX2 protein by interacting with miR-221-3p and that NPTX2 plays a role in regulating autophagy in PD. These findings indicate that targeting the interaction between HOTAIR, miR-221-3p, and NPTX2 could be a potential treatment strategy for PD (Lang et al., 2020).
Crocin is a natural pigment found in saffron that gives it its bright red color (Pan, Wen, Ma, Qin, & Feng, 2022). It possesses antioxidant, neuroprotective, and anti-inflammatory effects according to studies (Kermanshahi et al., 2020; Nam et al., 2010). Research has looked at the prospect of crocin as a treatment for various diseases, including PD, AD, and cancer (Ahmed et al., 2020; Veisi et al., 2020). PI3K enzyme has a crucial part in transmitting signals within cells that control cell growth, division, differentiation, and survival (Juntilla & Koretzky, 2008). It is activated by signals from outside the cell, such as growth factors and hormones, and converts one type of molecule (PIP2) into another (PIP3) (Luo, Manning, & Cantley, 2003). PIP3 then activates other signaling molecules within the cell, such as Akt and mTOR, which control various cellular processes (Park et al., 2010). Akt is an enzyme that has a crucial part in transmitting signals within cells that control cell growth, division, differentiation, and survival (Dickson & Rhodes, 2004; Macintyre et al., 2011). Akt is stimulated by various signaling components, such as growth factors, cytokines, and hormones, and it phosphorylates downstream signaling molecules, such as mTOR, GSK-3β, and FoxO3a, which regulate various cellular processes (B.-G. Li, Hasselgren, & Fang, 2005; Salama, Abdel-Latif, Abbas, Hekmat, & Schaalan, 2020). A study by Rania and colleagues showed that crocin, a natural pigment found in saffron, can protect against PD in rats exposed to the chemical rotenone. This protective effect is achieved through the interaction between the PI3K/Akt/mTOR signaling pathway and increased levels of two miRNAs, miRNA-7 and miRNA-221. The PI3K/Akt/mTOR pathway is stimulated by crocin, increasing the activity of some proteins while reducing brain cell death. Crocin also reduces cell death and levels of a protein called α-syn. These results provide new information about how crocin may protect against PD (Salama et al., 2020).
PTEN is a tumor suppressor gene that produces a protein that acts as a phosphatase, removing phosphate groups from specific proteins (Pérez-Ramírez, Cañadas-Garre, Molina, Faus-Dáder, & Calleja-Hernández, 2015). It helps maintain balance in the PI3K/AKT pathway and may suppress tumor growth independently of its role in the PI3K pathway (Dubrovska et al., 2009). As described by the research conducted by Li et al. miR-221 may have a protective effect in PD. Their research showed that levels of miR-221 were substantially reduced in cell models of PD induced by the chemical 6-OHDA. When miR-221 levels were increased, the cells showed improved viability and growth and a decrease in cell death. They found that AKT phosphorylation is significant in PD as it is involved in regulating cell survival and apoptosis. miR-221 promotes AKT phosphorylation by targeting PTEN, thereby activating the PI3K/AKT pathway and potentially contributing to the protective role of miR-221 in PD. In summary, miR-221 can be a potential treatment goal for PD (L. Li et al., 2018).
Apoptosis is a natural mechanism observed in multicellular organisms, facilitating the elimination of damaged or unnecessary cells from the body (D’arcy, 2019). This process is tightly regulated through intricate signaling pathways and involves the activation of caspases, a class of enzymes that act in targeted molecule degradation and subsequent cellular breakdown (Boice & Bouchier-Hayes, 2020). Apoptosis, or ”programmed cell death,” is a mechanism that involves the Bax/caspase-3 signaling pathway (Zamanian et al., 2017; Zhu et al., 2022). BAX is a member of the Bcl-2 family of proteins, which are part of the apoptosis regulators and either promote or inhibit cell death. Its primary location is in the cytoplasm of cells. During apoptosis, Bax undergoes a conformational change and translocates from the cytoplasm to the mitochondria (Ansari, Ball, Wase, Novak, & Haqqi, 2021). Caspase-3, in turn, acts as a protease that selectively cleaves specific substrates, leading to the dismantling of the cell (P. Yadav, Yadav, Jain, & Vaidya, 2021). Bim is a pro-apoptotic protein that participates in controlling how cells undergo apoptosis. It belongs to the Bcl-2 family of proteins, which are essential for regulating the apoptosis of neurons (Kim et al., 2017; Linseman et al., 2002). It has roles in the regulation of numerous types of neurons, such as sympathetic neurons, cerebellar granule neurons, and cortical neurons (Linseman et al., 2002). Bim is regulated by transcriptional factors such as FOXO, Egr-1, and the JNK/c-Jun pathway (Xie et al., 2011). About PD, the expression of Bim is controlled by the activation of the JNK/c-jun signaling pathway and the decrease in its translocation to the mitochondrial field (Hu et al., 2019). According to the research, there was a notable decrease in miR-221 levels in both the blood of PD patients and in the brain tissue of mice with a chemically induced form of the disease. The study showed that introducing additional miR-221 prevented the death of dopamine-producing neurons in the mice and influenced the Bax/caspase-3 pathway by interacting with Bim, a protein that promotes cell death. These are evidence that miR-221 could be a potential target for developing treatments to protect dopamine-producing neurons and alleviate symptoms of PD (Yao et al., 2023).
DJ-1 is a gene that produces a protein that helps protect cells from oxidative stress (Kahle, Waak, & Gasser, 2009). Changes in the DJ-1 gene have accompanied the primary development of PD (Kahle et al., 2009). DJ-1 has been shown to regulate the thioredoxin/ASK1 complex, which protects against oxidative damage, and to adjust miR-221 levels to promote neuronal survival in the face of oxidative stress (Oh et al., 2018). DJ-1 has been shown to help prevent cellular demise by protecting against oxidative stress and it also protects nerve cells due to its movement to the mitochondria driven by cysteine-sulfinic acid (Canet-Avilés et al., 2004). Oh et al. discovered that DJ-1, a gene product associated with PD, promotes neuronal preservation in the face of oxidative stress. Their study found that when miR-221 was overexpressed in SH-SY5Y cells, cell viability increased significantly following treatment with MPP+, a dopaminergic neurotoxin that induces oxidative stress. Specifically, they found that DJ-1 adjusts miR-221 levels to shield the neurons from oxidative stress. They also discovered that when DJ-1 was knocked down, neurite outgrowth decreased and susceptibility to cell death caused by oxidative stress increased, while overexpression of miR-221 reduced these effects. Additionally, they found that miR-221 targets transcripts of apoptotic proteins, including BIM, FOXO3a, BMF, and BNIP3L, which are predicted to be suppressed by miR-221 (Oh et al., 2018). In conclusion, DJ-1 and miR-221 could be anticipated endpoints for the development of therapies for PD.
The SNCA gene is associated with PD and produces a protein called α-syn. Changes in the SNCA gene have been associated with inherited types of PD. (Deng & Yuan, 2014). This protein is a key constituent of the protein aggregates known as Lewy bodies, which are present in the brains of individuals with PD (Fields, Bengoa-Vergniory, & Wade-Martins, 2019). The PARK2 gene is also associated with PD and produces a protein called parkin. Changes in the PARK2 gene have been associated with early-onset forms of PD (Nuytemans, Theuns, Cruts, & Van Broeckhoven, 2010). Parkin helps break down damaged or misfolded proteins in cells (Kilarski et al., 2012). When parkin doesn’t function properly, toxic proteins can build up in cells which can lead to the development of PD (Kilarski et al., 2012). Tatura et al. reported that miR-221 is upregulated in their study and its upregulation is consistent with the observed decrease in SNCA transcript levels. Additionally, the downregulation of miR-221 is consistent with its potential role in the onset and development of PD (Tatura et al., 2016).
TfR2, or transferrin receptor 2, is a protein that helps regulate the balance of iron in the body (Piperno et al., 2004). It acts as a carrier for transferrin, a molecule that binds to iron and transports it to different parts of the body (Robb & Wessling-Resnick, 2004). TfR2 shares some similarities with another transferrin receptor protein called TfR1, but its specific function in iron regulation is not yet fully understood (Kleven, Jue, & Enns, 2018). TfR2 is present in various tissues, like the liver as well as pancreas, and brain, and mutations in the TfR2 gene have been detected in a rare genetic disorder called hereditary hemochromatosis type 3, which is characterized by iron overload in the body (Bao et al., 2016; Yamamoto et al., 2002). Asci et al. used various techniques, including bioinformatics tools and miRNA inhibitors, to identify and manipulate miRNAs that may be involved in regulating TfR2 expression. They found that inhibiting endogenous levels of miR-221 in SH-SY5Y cells increases the expression of TfR2, and both the mRNA and protein expression of TfR2 can be reduced by miR-221. The research also showed that microRNA 221 interacts with a specific part of the TfR2 messenger RNA molecule and reduces its activity. This suggests that miR-221 can decrease the production of TfR2 protein. In conclusion, miR-221 may help adjust TfR2 levels in certain cells and could provide insight into the development of PD (Asci et al., 2013).
Paraoxonase 1 (PON1) is an enzyme that helps protect against oxidative stress (Kotur-Stevuljevic et al., 2015). PD is characterized by a disproportion between acetylcholine (ACh) and dopamine (DA) in the CNS (Rizzi & Tan, 2017). This is mainly due to the degeneration of dopaminergic neurons in the substantia nigra (Lehéricy, Sharman, Santos, Paquin, & Gallea, 2012). Acetylcholinesterase can be attacked and inhibited by free radicals such as peroxide and organophosphate substances, which causes an increase in acetylcholine and worsens the imbalance between ACh and DA (Al-Baggou, Naser, & Mohammad, 2011; Pearson & Patel, 2016). To block the imbalance between ACh and DA, the PON1 enzyme suppresses these free radicals (Menini & Gugliucci, 2014). Arylesterase, (ARE) is an enzyme that is involved in the metabolism of organophosphate compounds (Piras et al., 2021). According to a report by Ghita and Deeb, serum levels of miRNAs (miR-214, miR-141, and miR-221) and antioxidants (uric acid, PON1, and ARE) were significantly lower in cases with PD in comparison to healthy individuals (Ghit & Deeb, 2022). UPDRS-III is utilized to evaluate the movement-related symptoms of PD, including shaking, stiffness, slowed movement, and balance issues (M. Lu et al., 2020). The scale has a range of 0 to 108, where a higher score signifies more severe motor symptoms (Thobois et al., 2002). UPDRS-V is utilized to evaluate the daily living activities and independence of individuals with PD (Shu, Qian, & Wang, 2020). The scale has a range of 0 to 100, where a higher score signifies greater dependence on others for performing daily activities (Tibar et al., 2018).
The Area Under the Curve (AUC) value ranges from 0 to 1, with 0.5 indicating a test with no discriminatory ability (equivalent to random chance) and 1 indicating a perfect test that can accurately distinguish between the two groups (Walter, 2005; Wolever, 2004). Ma et al. discovered that serum miR-221 was considerably reduced in individuals with PD and had a positive correlation with UPDRS-III and UPDRS-V scores, showing its potential as an indicator of PD severity. The AUC value for predicting PD using serum miR-221 was 0.787, suggestive of its prospective use as a biomarker for diagnosing PD. These findings suggest that downregulated serum miR-221 might be considered a biomarker for the assessment of PD (W. Ma et al., 2016).