Alzheimer's disease neuropathology is exacerbated following traumatic brain injury. Neuroprotection by co-administration of nanowired mesenchymal stem cells and cerebrolysin with monoclonal antibodies to amyloid beta peptide

Abstract

Military personnel are prone to traumatic brain injury (TBI) that is one of the risk factors in developing Alzheimer's disease (AD) at a later stage. TBI induces breakdown of the blood-brain barrier (BBB) to serum proteins into the brain and leads to extravasation of plasma amyloid beta peptide (ΑβP) into the brain fluid compartments causing AD brain pathology. Thus, there is a need to expand our knowledge on the role of TBI in AD. In addition, exploration of the novel roles of nanomedicine in AD and TBI for neuroprotection is the need of the hour. Since stem cells and neurotrophic factors play important roles in TBI and in AD, it is likely that nanodelivery of these agents exert superior neuroprotection in TBI induced exacerbation of AD brain pathology. In this review, these aspects are examined in details based on our own investigations in the light of current scientific literature in the field. Our observations show that TBI exacerbates AD brain pathology and TiO₂ nanowired delivery of mesenchymal stem cells together with cerebrolysin—a balanced composition of several neurotrophic factors and active peptide fragments, and monoclonal antibodies to amyloid beta protein thwarted the development of neuropathology following TBI in AD, not reported earlier.

Introduction

Alzheimer's disease (AD) is a slowly progressing brain disease resulting in memory loss and other intellectual functions (Bondi et al., 2017; Kirova et al., 2015; Leli and Scott, 1982; Lyketsos et al., 2011; Shimada et al., 2001). The AD may develop over 20 years from early signs of inception resulting in gradually interfering with normal daily life (Besser et al., 2018; O'Bryant et al., 2015; Weiner et al., 2015). AD is not a disease of aging as it can appears in patients of 40 or 50 years of age known as younger onset (Juan and Adlard, 2019; Lane et al., 2018; Soria Lopez et al., 2019; Tromp et al., 2015). In 2016, AD represents the sixth leading causes of death in the United States of America (USA) (Alzheimer's Association, 2016; Calamia et al., 2016; Yeh et al., 2017). In the beginning memory loss in AD is mild that become severe with advancing age and disease (Jahn, 2013; Wang and Holtzman, 2020). People with AD normally live 8–10 years after initial diagnosis depending on the age and other co-morbidity factors (Alzheimer's Association, 2015; Mueller et al., 2019; Zhang et al., 2020a, Zhang et al., 2020b). Till date there are no cure for AD but available treatments could slow down the progression of disease and symptoms (Cortes-Canteli and Iadecola, 2020; Cummings et al., 2019; Shao et al., 2020). Thus, there is an urgent need to explore novel treatment strategies to alleviate brain pathology in AD or thwart the developmental processes of the disease.

The normal healthy human brain contains about 100 billions of neurons that are connected with each other with synapses for normal communication to maintain brain functions (Pfisterer and Khodosevich, 2017; Sporns et al., 2005; von Bartheld et al., 2016). About 100 trillions synapses allow flow of information through the neuronal circuits forming the cellular basis of memory, thoughts, emotion and other intellectual functions (Mesulam, 1998; Stampanoni Bassi et al., 2019; Zimmer, 2011). The number of neurons and their connections through synapses severely deteriorate during AD resulting in loss of memory and higher bran functions (Jackson et al., 2019; Meyer et al., 2019; Osborn et al., 2016). Deposits of the peptide fragment known as amyloid beta peptide (AβP) outside of the neurons and accumulation of the abnormal form of tau protein inside the neurons are the hallmark of AD brain (Blennow et al., 2006; Busche and Hyman, 2020; van der Kant et al., 2020). The AβP plaques contribute to cell death by interfering with neuronal communication at the synapses in AD (Leong et al., 2020; Malishev et al., 2019; Picone et al., 2020). The abnormal tau proteins known as tau tangles inhibit the transport of essential molecules and nutrients within the neurons resulting in cell death (Blurton-Jones and Laferla, 2006; Mata, 2018; Rodríguez-Martín et al., 2013). When the level of AβP deposits increases the tau tangles spread throughout the brain resulting in profound symptoms of AD and brain damage (Bassil et al., 2020; Bloom, 2014; Lowe et al., 2018).

The progress of AD with time results in shrinkage of the brain tissues together with the gradual enlargement of all the cerebral ventricles (Ferreira et al., 2015; Kuroda et al., 2020; Mentis et al., 2020; Ye et al., 2016). The hippocampus cells shrinking causes neurodegeneration resulting in memory decline (Jaroudi et al., 2017; Khanal et al., 2014; Mu and Gage, 2011; Tabatabaei-Jafari et al., 2017). With time AD spreads throughout the cerebral cortex and brain resulting in behavioral changes, higher mental and vital function deterioration affecting language, judgment, memory, daily life and other body systems until death within 9–10 years after diagnosis (Brookmeyer and Abdalla, 2019; Skaper et al., 2017; van Engelen et al., 2020).

The trigger of the molecular mechanisms leading to AβP deposits and/or abnormal or phosphorylated tau (p-tau) accumulation leading to AD pathology is still unclear (Gatti et al., 2020; Paroni et al., 2019; Wang and Zhang, 2018). In healthy conditions, very low levels of AβP and tau is present in the plasma that appears to be increased in patients with AD (Karikari et al., 2020; Mattsson et al., 2016, Mattsson et al., 2017; Schultz et al., 2019; Wei et al., 2017; Yun et al., 2020). It appears that in AD plasma AβP and tau are transported to the brain and in the cerebrospinal fluid (CSF) probably due to a compromise of the blood-brain barrier (BBB) and the blood-CSF-barrier (BCSFB) function (Sharma, 2004, Sharma, 2009; Sharma and Sharma, 2010; Sharma and Westman, 2004; Sharma et al., 2012). The healthy brain normally removes abnormal proteins from the brain via efflux transporters located at the cerebrovascular unit to maintain homeostasis (Bao et al., 2020; Iliff et al., 2012; Sharma, 2004). However, accumulation of the AβP and p-tau in AD brain suggests that these transporters or brain-blood barrier (bbb) is also affected (Hladky and Barrand, 2018; Jadiya et al., 2019).

One of the earliest precipitating factors in inducing AD appears to be traumatic brain injury (TBI) (Armstrong, 2019; Kempuraj et al., 2020; Wu et al., 2020). The linkage between brain trauma and neurodegenerative diseases is responsible for late onset of several chronic brain disorders (Bertogliat et al., 2020; Gardner and Yaffe, 2015; Ramos-Cejudo et al., 2018). Increasing evidences show that even a single mild TBI sustained in early life may lead to a cascade that could manifest in late development of AD-like disorders (Becker et al., 2018; Griesbach et al., 2018; Gupta and Sen, 2016). Several cases of boxers that get repetitive mild head trauma leads to neurodegenerative symptoms in some cases as early as in their 20s and 30s years of life (Alosco and Stern, 2019; McKee et al., 2018; Ossenkoppele et al., 2020). Repetitive head trauma causing chronic traumatic encephalopathy (CTE) results in several neurological disorders including AD (Iverson et al., 2019; McKee et al., 2009; Simom et al., 2017). Neurotrauma could induce cerebrovascular diseases (CBD) leading to the disruption of the BBB and BCSFB (Johanson et al., 2011; O'Keeffe et al., 2020; Sahyouni et al., 2017; Sharma and Johanson, 2007a, Sharma and Johanson, 2007b). These cerebrovascular disturbances may trigger AβP and p-tau accumulation leading to AD (Patnaik et al., 2018; Sharma et al., 2012, Sharma et al., 2016a, Sharma et al., 2016b). Military personnel are often prone to TBI resulting in post-traumatic stress disorders (PTSD) that are prevalent in soldiers associated with combat experiences (Adrian et al., 2018; Groer et al., 2015; Kritikos et al., 2019). Thus, it is quite likely that TBI could exacerbate AD brain pathology (Moore et al., 2020; Proessl et al., 2020; Silverberg et al., 2020). Accordingly, treatment modalities for improving or exploring patient care is needed to enhance clinical efficacy in situations of AD with TBI.

In this review we discuss novel therapeutic strategies to treat exacerbation of brain pathology in AD with TBI using nanomedicine based on our own observations. New evidences suggest that nanowired delivery of neurotrophic factors and stem cells could be the potential novel therapeutic agents in treating AD cases with TBI, not reported earlier.

Several lines of evidences suggest that impairment of cerebrovascular function is the prominent contributing factor in the pathophysiology of AD (Bell and Zlokovic, 2009; Nelson et al., 2016; Zlokovic, 2011). Pathological evaluation of albumin leakage within the AD brain suggests that breakdown of the BBB plays a key role (Montagne et al., 2017; Sweeney et al., 2018, Sweeney et al., 2019). The BBB resides in the cerebral capillaries that are connected with tight junctions (Abbott et al., 2010; Sharma, 2009; Sharma and Westman, 2004; Zlokovic, 2008). Thus, the cellular and subcellular structures comprising endothelial cell covering of basal lamina, glial end feet and neuronal contacts with the microvessels are actively separating the blood from brain parenchyma (Abbott et al., 2006; Liebner et al., 2018; Profaci et al., 2020). The BBB is thus maintaining strict microenvironment of the brain by controlling passage of essential nutrients through active and passive transport mechanisms (Banks, 2016; Erdő et al., 2017; Sharif et al., 2018). The BBB also prevents neurotoxins from entering into the brain (Alexander, 2018; Obermeier et al., 2013, Obermeier et al., 2016). The clearance function of the BBB using active transport and selective transporters maintains the healthy environment of the brain in normal conditions (Bell et al., 2007; Deane et al., 2009; Shibata et al., 2000). Deterioration of the BBB active maintenance of the brain function or reduction in the clearance capacity of toxins and unwanted substances results in microvascular abnormalities, microbleeds, protein deposits, neuroinflammation and neuronal cell death (Cai et al., 2018; Ransohoff, 2016; Sulhan et al., 2020). These pathological phenomena within the brain lead to decline of intellectual and cognitive functions promoting dementia and eventually AD (Eratne et al., 2018; Masters et al., 2015; Sweeney et al., 2019).

This idea is well substantiated by albumin ratio measurement and contrast material enhancement using magnetic resonance imaging (MRI) to detect BBB leakage in neurodegenerative diseases (van de Haar et al., 2016a, van de Haar et al., 2016b, van de Haar et al., 2017). Using these techniques neuroradiologists in the Netherlands examined BBB leakage in patients with early AD and compared with age-matched healthy controls. The authors used a dynamic contrast enhanced MRI with dual-time resolution that separates the leakage from the microvascular fillings (van de Haar et al., 2016a, van de Haar et al., 2016b). In addition, local blood plasma volume and its relationship with the BBB permeability and global cognition were also investigated.

The authors find increased BBB leakage in patients with early AD that was globally distributed within the cerebrum and correlates well with the declined cognitive performances (van de Haar et al., 2016a, van de Haar et al., 2016b). Using dynamic contrast-enhanced MRI for dual resolution the BBB leakage strength or rate is seen in the gray matter as well as in the white matter within the cerebrum (van de Haar et al., 2017). However, the leakage volume was greater than the leakage rate in early AD cases. Thus, BBB breakdown appears to be global in early AD cases (van de Haar et al., 2016a, van de Haar et al., 2016b, van de Haar et al., 2017).

Apart from BBB leakage the local plasma volume was also decreased in the areas showing BBB breakdown indicating ischemic hypoperfusion of the brain (Dong et al., 2018; Feng et al., 2019; Love and Miners, 2016). Ischemia induced cerebrovascular perfusion and endothelial cell dysfunction including leakage results in microbleed and small vessel diseases quite common in dementia and in AD (Arvanitakis et al., 2016; Blumenau et al., 2020; Hase et al., 2020). Another interesting point from this study showed a close correspondence between the strength of BBB leakage and decline in cognitive function (Kamintsky et al., 2020; Raja et al., 2018; Uemura et al., 2020). Although the mechanism of the BBB leakage in AD is still uncertain, it appears that endothelial cell permeability along with loss or damage of the tight junctions are both could be responsible for such leakage (Sharma, 2004, Sharma, 2009; Sharma et al., 2012; Sweeney et al., 2019). These observations indicate that both influx and efflux transport systems across the BBB is compromised in AD (Jeynes and Provias, 2011; Kim et al., 2020a, Kim et al., 2020b, Kim et al., 2020c; Zhou et al., 2017).

There are reasons to believe that BBB is a crucial determining factor for AβP and tau deposition in the AD brain (Kent et al., 2020; Laurent et al., 2018; Liu et al., 2019). Obviously, a breakdown of the influx and efflux transport system at the BBB will allow greater deposits and accumulation of these neurotoxins responsible for AD brain pathology (Sweeney et al., 2019; Xin et al., 2018; Zenaro et al., 2017). This is further evident from the findings that there is a close association between vascular inflammation and neuroinflammation in AD brain pathology (Klohs, 2019; Loera-Valencia et al., 2019; Sung et al., 2020). In AD, two key factors namely AβP deposition and neuroinflammation play predominant roles in brain pathology (Calsolaro and Edison, 2016). Although the precise mechanisms between the interaction of these two factors are not well known, it is amply clear that neuroinflammation predates the AβP deposition (González-Reyes et al., 2017; Hoozemans et al., 2006; Regen et al., 2017).

Several lines of evidences suggest that innate immune system plays pivotal role in inflammation and subsequent progression of AD (Gate et al., 2020; Park et al., 2020a, Park et al., 2020b; Webers et al., 2020). Activation of microglia and macrophages together with AβP and p-tau are responsible for AD brain pathology (Bartels et al., 2020; Heneka, 2020; Yuan et al., 2020). Activation of neuroimmune system allows clearance of AβP or other neurotoxic materials from the brain and leads to cell death of damaged neurons (Bedoui et al., 2018; Griffiths et al., 2007). However, in chronic activation of immune system leads to production of neurotoxic cytokines, chemokines and oxidative stress resulting in slow neuronal death in AD (Friker et al., 2020; Hoarau et al., 2011; Welikovitch et al., 2020).

Recent evidences show that one of the peripheral blood enzymes plasmin (PLS) is an important regulatory factor of neuroinflammation and brain pathology in AD (Baker et al., 2018, Baker et al., 2019; Paul et al., 2007). PLS enzyme is formed from cleavage of the primary blood protein plasminogen (PLG) that is synthesized in the liver (Huebner et al., 2018). The PLS is involved in the cell signaling, inflammatory responses, fibrinolysis and wound healing (Castellino and Ploplis, 2005). PLG plays a key role in activating inflammation and immune responses (Heissig et al., 2020). This is a chemoattractant for macrophages and monocytes have binding sites for PLG (Syrovets et al., 2012). Depletion of PLS pharmacologically results in increased survival rate due to less inflammation caused by excessive immunoactivation (Cuzner and Opdenakker, 1999; Gur-Wahnon et al., 2013). This suggests that PL is involved in production of proinflammatory cytokines and chemokines (Kang et al., 2020).

Since PLG and PLS are involved in neuroinflammatory behavior a possibility exists that pharmacological depletion of these enzymes may have some neuroprotective effects in AD.

The mechanisms behind the progression of AD brain pathology are still not well known. However, it appears that neuroinflammation is one of the key factors in contributing AD brain pathology (Newcombe et al., 2018; Regen et al., 2017; Wilkins and Swerdlow, 2016). The hallmark of AD includes AβP extracellular deposition and p-tau intracellular accumulation causing brain damage is preceded with activation of neuroimmune system and inflammatory responses of glial cells (LaFerla et al., 2007; Šimić et al., 2016; Villemagne et al., 2018). Activation of microglia could lead to clear AβP deposition through phagocytes up to some extent (Süß and Schlachetzki, 2020). However, chronic activation of immune system causes misfolding to AβP aggregation that is difficult to clear or dissolve in AD (Griciuc et al., 2013). In addition, AβP could activate proinflammatory cytokines through activating the glial cells by binding on the receptors present on them (Minter et al., 2016; Wang et al., 2015). The AβP can also activate proinflammatory cell signaling system in plasma and in cerebrospinal fluid (CSF) to induce neuroinflammation (Pillai et al., 2019; Sutton et al., 1999). This idea is further supported by the fact that depletion of AβP proinflammatory contact system results in decrease in neuroinflammation, brain pathology and cognitive dysfunction in AD (Kinney et al., 2018).

Several plasma factors are known activators of PLG and PLS affecting AD brain pathology (Baker et al., 2018; Simão et al., 2017). Depletion of blood coagulation factor XII (CFXII) responsible for intrinsic pathways for coagulation and immune contact activation is able to reduce brain pathology and neuroinflammation in AD (Chen et al., 2017). In addition CFXIIa, CFXI1, kallikrein and bradykinin that are key molecules for immune contact pathways are known activators of PLG and PLS (Göbel et al., 2019). When PLG and PLS are activated neuroinflammation is enhanced leading to brain pathology (Reuland and Church, 2020). CFXII is also activated by PLS that causes release of bradykinin—a proinflammatory agent and mediator of vascular permeability causing edema and neuroinflammation (Mugisho et al., 2019). In addition CFXII also induces thrombin generation another potent proinflammatory factor induces neuroinflammation through activation of protease receptors in the CNS (Iannucci et al., 2020).

One of the key factors in neuroinflammation is the deposition of fibrinogen within the CNS (Cortes-Canteli et al., 2012; Sulimai and Lominadze, 2020). When the BBB is compromised fibrin enters into the brain and deposited into several brain areas causing neuroinflammation (Cortes-Canteli et al., 2015). Deposition of fibrinogen is associated with depletion of PLG or PL in the CNS (Shaw et al., 2017). This idea is further supported by the fact that inhibition of PLG by tranexamic acid (TXA) enhances neuroinflammatory response in the CNS (Atsev and Tomov, 2020). Interestingly TXA is able to inhibit only free PLG or PL but could not affect cell bound PL activity (Baker et al., 2018).

Evidences show that AβP plaques are significantly decreased in PLG depletion in mice model of AD using antisense oligonucleotide (ASO) treatment (Baker et al., 2018). However, this decrease of AβP plaques with PLG depletion is unclear. There are reasons to believe that PL could enhance the cleavage of both a- and b-amyloid precursor protein (APP) (Baranello et al., 2015). There are reports that PLG activation (PA) is induced by AβP that could degrade oligomeric and fibrillar AβP (Tucker et al., 2000). Interestingly, tissue PLG activation (tPA) results in inhibition of AβP aggregation and neurotoxicity (Yang et al., 2020). Both PLG and tPA decreases with advancing age are associated with AD but their expression is increased around AβP plaques (Cacquevel et al., 2007). This indicates that PL is associated with tPA-induced proteolysis in AD (Nalivaeva et al., 2008). This aspect is further confirmed in experiments with genetic depletion of protein PAl-1, the major inhibitor of tPA that resulted in decrease in AβP deposition in mouse model of AD (Kutz et al., 2012). This observation clearly suggests an important role of PL in AβP clearance (Baker et al., 2018). PLG depletion thus reduces that ability to induce prominent neuroinflammation and therefore less AβP deposition in the AD brains (Baker et al., 2018, Baker et al., 2019).

Increased activation of PL is often seen in chronic inflammatory or autoimmune disorders (Cañas et al., 2015). In addition, PLG and tPA are found localized around AβP plaques that could further enhance local inflammatory responses (Riemenschneider et al., 2006). However, further studies are needed to examine the therapeutic effects of PLG in AD.

The signature characteristics of AD include extracellular deposition of AβP plaques, intracellular tangles of p-tau proteins, loss of synapses and neurons leading to brain pathology (Brun et al., 1995; Liu et al., 1999). However, apart from classical AD cases similar deposition of AβP is seen in TBI as well as PTSD cases (Emmerling et al., 2000; Mohamed et al., 2018; Raby et al., 1998). However, in each case the intensity and magnitude together with regional variation of AD deposition occurs. These specificities led to several functional imaging studies resulting in novel discoveries and fingerprinting of AβP deposits in several diseases (Veitch et al., 2019).

There exists a close relationship between AβP deposits and cognitive impairments in several imaging studies of TBI and AD brains of short or long-term survival (Abrahamson and Ikonomovic, 2020; Ayubcha et al., 2021). Increased AβP deposits found in TBI patients with PET studies that were largely confined into the posterior cingulate cortex, cerebellum and striatum very similar to that seen in AD cases (Hong et al., 2014; Stern et al., 2019; Ubukata et al., 2020; Yang et al., 2015). AβP plaques are seen in more than one third of all TBI patients irrespective of their age (Mohamed et al., 2018). This indicates that TBI is an important risk factor for AD development. Interestingly, AβP deposits in TBI is seen following short term injury and could not be found in many long-term survivor of brain trauma (Mohamed et al., 2020; Scott et al., 2016). This indicates that enhanced AβP clearance is active in eliminating AβP from the brain fluid microenvironment. However, when the clearance mechanisms are compromised AβP accumulation persists and spread through different parts of brain leading to full development of AD cases (Reddy and Oliver, 2019; Suzuki et al., 2015). Taken together these studies clearly suggest that TBI induce deposition of AβP is a risk factor for later development of AD brain pathology (Djordjevic et al., 2016; Jellinger, 2004; LoBue and Cullum, 2020).

Military personnel while engaged in combat operations are susceptible to TBI followed by PTSD (Chin and Zeber, 2020; Iljazi et al., 2020; Mac Donald et al., 2021; Moore et al., 2020). There are several reports suggesting that these military personnel are highly vulnerable to later development of AD brain pathology (Khachaturian and Khachaturian, 2014; Veitch et al., 2013; Weiner et al., 2013). Interestingly, like TBI, PTSD also shows deposition of AβP in specific brain areas (Mohamed et al., 2018, Mohamed et al., 2019). Thus, it is suggested that veterans with PTSD could have a twofold increase in the risk of developing AD (Chan et al., 2017; Desmarais et al., 2020; Günak et al., 2020; Rafferty et al., 2018). Recently, imaging studies of veterans with PTSD showed AβP deposition in the precuneus, frontal, temporal, parietal and anterior and posterior cingulate cortices associated with cognitive functional decline very similar to that of AD cases (Mohamed et al., 2018, Mohamed et al., 2020). These observations suggest that AβP could be a possible link between PTSD and AD.

It appears that stress related changes in hypothalamic-pituitary-adrenal (HPA) axis could play an important role between PTSD and AD. This is further evident from the fact that loss of the volume of hippocampus and associated cognitive decline is common in both the PTSD and AD (Bonne et al., 2008; Scheff et al., 2006; Schuff et al., 2009; Wang and Xiao, 2010). Several MRI studies in PTSD show decrease in volume of hippocampus, anterior cingulate cortex and prefrontal structures in patients and in AD (Boutet et al., 2014; Murphy et al., 2003; Parker et al., 2019; Schuff et al., 2009). In addition, PTSD and AD are associated with several neuroinflammatory factors including cytokines, chemokines and other neurodegenerative elements including AβP accumulation (Cai et al., 2014; Gill et al., 2018; Webers et al., 2020; Webster et al., 2015).

Studies showing good correlation in CSF markers of AβP and p-tau in TBI, PTSD and AD further suggest a close link between them (Blennow et al., 2010; Mohamed et al., 2018, Mohamed et al., 2019; Rabbito et al., 2020; Zou et al., 2020). This suggests an increased risk of TBI and or PTSD in development of AD brain pathology in later periods of life. However, slight and subtle differences in AβP accumulation occur in all three groups of patients with TBI, TBI and PTSD or AD (Mohamed et al., 2018). Thus, patients with TBI alone increased accumulation of AβP are seen in the cerebellum and precuneus (Hortobágyi et al., 2007; Mohamed et al., 2018). On the other hand, TBI and PTSD group showed a substantial increase in accumulation of AβP in the white matter besides the other brain regions (Mohamed et al., 2018, Mohamed et al., 2019). On the other hand PTSD alone exhibited AβP deposits in the frontal, occipital and temporal cortices (Mohamed et al., 2018). These differences in accumulation of AβP distribution show differential relationships between TBI, TBI and PTSD and PTSD alone in relation to development of AD brain pathology.

Mesenchymal stem cells (MSCs) are one of the prominent therapeutic tools in the treatment strategies of AD (Chakari-Khiavi et al., 2019; Kim et al., 2020a, Kim et al., 2020b, Kim et al., 2020c; Wang et al., 2019; Zhang et al., 2020a, Zhang et al., 2020b). Several studies have shown the efficacy of MSCs treatment in AD because of their immune modulatory and neurotrophic functions (Elia et al., 2019; Lo Furno et al., 2018; Mehrabadi et al., 2020; Zhang et al., 2020a, Zhang et al., 2020b). Some studies have also investigated the use of AD patients CSF samples as formulations of MSCs for the treatment of AD therapy (Benhamron et al., 2020; Lee et al., 2019).

The most important aspect of stem cell therapy in neurodegenerative disease like AD is to choose appropriate cell sources (Han et al., 2020). There are evidences that embryonic stem cells (ESCs), MSCs, brain-derived neural stem cells (NSCs) and induced pluripotent stem cells (iPSCs) are used in AD therapy (Farahzadi et al., 2020; Kim et al., 2020a, Kim et al., 2020b, Kim et al., 2020c; Liu et al., 2020; Lin et al., 2018; Penney et al., 2020). The MSCs induce development of mesenchymal tissue and can be harvested from umbilical cord blood (UCB-MSCs) or Wharton's jelly (Kim et al., 2015; Petukhova et al., 2019). In addition MSCs are present in adult stem cells niches, e.g., bone marrow and adipose tissues (Nakano et al., 2020; Nasiri et al., 2019; Qin et al., 2020; Reza-Zaldivar et al., 2018).

The MSCs are originally discovered from stroma of the bone marrow are the multipotent progenitor cells that are extensively used for neurodegenerative therapy due to their immune modulatory and neurotrophic properties (Esmaeilzade et al., 2019; Wang et al., 2019; Yang et al., 2016). AD is one of the most important neurodegenerative diseases for which no suitable therapeutic strategies are developed until today (Sharma et al., 2012, Sharma et al., 2016a, Sharma et al., 2016b). The MSCs are used in AD therapy largely because of their intrinsic ability to reduce AβP levels in the brain and attenuate neuroinflammation (Ding et al., 2018; Habisch et al., 2010; Yokokawa et al., 2019). In addition MSCs are also capable to enhance endogenous neurogenesis and improve behavioral performances in AD. Based on these evidences several clinical trials are underway to explore the safety and efficacy of MSCs in AD patients (Mohamed et al., 2018, Mohamed et al., 2019). Regarding the most efficient routes of MSCs administration in AD preclinical results suggests that intracerebroventricular (i.c.v.) or intraparenchymal injections yield superior effects as compared to intravenous (i.v.) or intra-arterial (i.a.) injections (Elia et al., 2019; Kim et al., 2015; Mohammadi et al., 2019).

Another important point is to use clinical grade of MSCs preparation as drug in AD therapy (Samsonraj et al., 2017). However, further studies are needed to further explore this aspect in greater details. Normally, the MSCs are used currently include various supplements to promote the survival of MSCs. This increases the possibilities of unavoidable interactions among cellular system of the recipients. Thus, this is important to enhance survival of MSCs in the recipient brain using other safe techniques such as nanodelivery of MSCs in AD (Andrzejewska et al., 2020; Sarnowska et al., 2013; Sharma et al., 2018).

Another way of optimizing MSCs formulation for AD therapy is the CSF samples of AD patients (Joerger-Messerli et al., 2016; Lee et al., 2019). This has several advantages. Since MSCs are normally administered into the CSF of patients using intracerebroventricular routes that have higher chances to penetrate into the brain parenchyma (Johanson et al., 2011; Sharma and Johanson, 2007a, Sharma and Johanson, 2007b). Since, the CSF flow is responsible for AβP clearance strategy in AD this route of administration helps MSCs to enhance positive effects (Sharma and Westman, 2004). CSF from AD patients represents brain fluid microenvironment and thus MSCs given through CSF of individual patients may better survive with the disease environment as compared to naive MSCs (Dehghanian et al., 2020).

Recent observations using administration of Wharton's jelly MSCs formulations with AD patients CSF resulted in gene expression that are known to block apoptosis, cell proliferation and neurogenesis (Bodart-Santos et al., 2019; Lee et al., 2019, Lee et al., 2020). Furthermore, this treatment enhances extracellular transport of AβP and exhibited neuroprotective and neurotrophic activity (Kim et al., 2020a, Kim et al., 2020b, Kim et al., 2020c). In addition, MSCs therapy in patients CSF increased expression of genes responsible for cell migration and cell adhesion indicating potential beneficial effects on cell survival (Benvenuti et al., 2006). These observations indicate that patients CSF could be used as optimal formulation for MSCs for effective AD therapy (Sharma et al., 2012, Sharma et al., 2016a, Sharma et al., 2016b, Sharma et al., 2018). However, further studies using other techniques such as nanodelivery of MSCs are needed to expand our knowledge in effective AD therapy.