Clearance dysfunction of trans-barrier transport and lymphatic drainage in cerebral small vessel disease: Review and prospect

Cerebral small vessel disease (CSVD) causes 20% – 25% of stroke and contributes to 45% of dementia cases worldwide. However, since its early symptoms are inconclusive in addition to the complexity of the pathological basis, there is a rather limited effective therapies and interventions. Recently, accumulating evidence suggested that various brain-waste-clearance dysfunctions are closely related to the pathogenesis and prognosis of CSVD, and after a comprehensive and systematic review we classified them into two broad categories: trans-barrier transport and lymphatic drainage. The former includes blood brain barrier and blood – cerebrospinal fluid barrier, and the latter, glymphatic-meningeal lymphatic system and intramural periarterial drainage pathway. We summarized the concepts and potential mechanisms of these clearance systems, proposing a relatively complete framework for elucidating their interactions with CSVD. In addition, we also discussed recent advances in therapeutic strategies targeting clearance dysfunction, which may be an important area for future CSVD research.


Introduction
Cerebral small vessel disease (CSVD) is a syndrome of clinical, neuroimaging, and neuropathological manifestations associated with aging (Wardlaw et al., 2013), and magnetic resonance imaging (MRI) images from patients with it show characteristic abnormalities, such as white matter hyperintensities (WMH), recent small subcortical infarcts, lacunes, cerebral microbleeds (Debette et al., 2019).It is a major factor leading to stroke and other cerebrovascular events, cognitive impairment or dementia (Cannistraro et al., 2019), but due to its insidious clinical symptoms and slow progression, the importance of early diagnosis and intervention for it has easily been overlooked or underestimated, and worse, the poor understanding of its underlying pathogenesis hinders the development of effective treatments.
The pathophysiology of CSVD is complex, and current research has identified several vascular manifestations of dysfunction, such as arteriolosclerosis, lipohyalinosis, andfibrinoid necrosis (Wardlaw et al., 2019).However, these findings focused on vascular wall damage cannot fully explain the full picture of its pathological process, such as the reasons for the variable lesion progression and symptoms and their frequent co-existence with neurodegenerative diseases remain unclear (Ter Telgte et al., 2018), which highlights the urgency and importance of further exploring its alternative mechanisms driving this complex overlapping pathology.
1 These authors contributed equally to this work.
exact relationships between them, and few studies have simultaneously focused on them in CSVD.Therefore, based on the main anatomical, physiological and pathological features, we classified these clearance systems into two broad categories: the trans-barrier transport and lymphatic drainage, aiming to provide a comprehensive description of the concepts and potential mechanisms of brain's clearance systems and how their dysfunction can be associated with, or might play a role in-the progress of CSVD.Furthermore, we also elaborate on the crosstalk between these systems, discuss the approaches for monitoring and intervening their dysfunction and suggest possible clinical applications that may improve risk prediction and therapeutic potential in individuals with CSVD.

Potential association of CSVD with brain clearance dysfunction
Due to the brain's high metabolic rate, inability to remove soluble metabolic wastes with potential neurotoxicity on time may lead to a variety of abnormalities, especially age-related nervous system diseases (Nedergaard, 2013), including CSVD.To date, several direct and indirect studies have confirmed that most subtypes of CSVD are associated with brain-waste-clearance dysfunction.
First, the incidence of type-I arteriolosclerosis, also known as agerelated and vascular risk-factor-related CSVD, just as its name implies, is associated with age, hypertension, diabetes, and other vascular risk factors, which considerably contributes considerably to the brainclearance rate (Kress et al., 2014).Epidemiological data suggest that in the pathogenesis of hypertension and the aging process, vasomotor activity and pulsation decreases and the BBB permeability changes correspond to slower clearance rates and may lead to small vessel lesions (van Veluw et al., 2020;Shah et al., 2012).Using the animal models of hypertension, researchers have demonstrated that chronic vascular injury by hypertension could activate the receptors for advanced glycation-end products, affect proteins clearance and promote their deposition (Yang et al., 2018).Analogously, the accumulation of misaggregated proteins may be a key neuropathological mechanism in the diabetic brain.Therefore, brain-waste-clearance dysfunction is likely to be the transit point between these vascular risk factors and CSVD.
Second, in cases of types II and III CSVD, including sporadic cerebral amyloid angiopathy (CAA) and cerebral autosomal dominant arteriopathy with subcortical infarcts and leukoencephalopathy (CADASIL), evidence of abnormal accumulation of Aβ, Notch3 and additional proteins, without increased production, strongly suggests impaired clearance mechanisms (Joutel et al., 2000;Weller et al., 1998;Hawkes et al., 2011).CAA is characterized histopathologically by the accumulation of Aβ deposits in the walls of small vessels, and ex vivo MRI confirmed that it may be the consequence of the dysfunctional perivascular clearance in the brain (Perosa et al., 2022).Mouse models have further demonstrated that smooth muscle cell (SMC) degeneration may be the cause of perivascular clearance defects in CAA (van Veluw et al., 2020).CADASIL is caused by a dominant mutation in the Notch3 gene, and the accumulation and deposition of granular osmiophilic material (GOM) is essential for the development of its pathology (Monet-Leprêtre et al., 2013), which occurs due to the failure to eliminate proteins from the artery walls.In fact, both CAA and CADASIL have been described as protein elimination failure angiopathies owing to their common characteristic of failure to eliminate proteins from the brain (Carare et al., 2013).
Reduced blood flow in CSVD may lead to hypoxia, neuronal death, and other neuropathological changes that exacerbate the deterioration of the brain environment, coupled with vascular dysfunction, ultimately leading to impaired waste clearance in the brain (Iliff et al., 2013a).Animal models have demonstrated that microinfarcts inhibit glymphatic clearance and lead to neurotoxic solute aggregation, consequently promoting neuroinflammation and accelerating synaptic loss, thereby exacerbating CSVD (Wang et al., 2017).In other words, the brain waste clearance disorder is likely to be one of the main causes of CSVD, and in turn, the pathological changes of CSVD could further intensify the clearance dysfunction, forming a vicious cycle.

Overview of the brain's waste clearance systems
Waste clearance from the brain involves both intracellular and extracellular mechanisms.Wa et al. (Boland et al., 2018) analyzed the three main modes of intracellular clearance in detail, and here we mainly discuss the extracellular clearance mechanisms: (1) flow out of the brain parenchyma into the blood through efflux transporters at the BBB and BCSFB, and (2) bulk CSF-ISF flow clearance, including the glymphatic-meningeal lymphatic system and IPAD pathway through the peri-and para-vascular spaces to the cervical lymph nodes (Fig. 1).However, before embarking on that, let us first provide a brief introduction to the concept of several important components and anatomical structures, which may contribute to a better understanding of various clearance systems of the brain and their relationships.

Fluid compartments within the cranium
Apart from blood, two major fluid compartments are found within the cranium consisted, namely cerebrospinal fluid (CSF) in the ventricles and subarachnoid spaces, and interstitial fluid (ISF) within the extracellular spaces of the brain, spinal cord, and lymph.Their compositions exhibit sharp boundaries and clear differences, but some results suggest that they also flow one into another in a continuous pathway (Bito and Davson, 1966;Cserr et al., 1977).The CSF is produced by the choroid plexuses (CP) within the ventricles of the brain and circulates through the subarachnoid space of the skull and spinal column to provide buoyancy and maintain fluid homeostasis of the brain and spinal cord.ISF bathes neurons and glia of the parenchyma and is the medium for the delivery in essential molecules and the removal of waste.Bulk CSF and ISF exchange, occurring macroscopically along perivascular pathways throughout brain tissue, plays a pivotal role in the waste clearance within the central nervous system (CNS) (Iliff et al., 2013b), and is proposed to be an efficient clearance route for cell debris and waste products of metabolism that are too large to exit across the endothelium of the blood vessels.Kounda et al. (Koundal et al., 2020) found that aging spontaneously hypertensive stroke prone rats (SHRSP) (Hannawi et al., 2021) showed a significant reduction in CSF and ISF transport, and several other models explicitly related to CSVD show similar anomalies (Wardlaw et al., 2013;Attier-Zmudka et al., 2019).Another interesting finding is that a recent study demonstrated the predictive potential of CSF flow changed in the development of cognitive impairment in patients with CSVD (Dobrynina et al., 2020).

Perivascular spaces
Perivascular spaces, also called Virchow-Robin spaces, are ISF-filled channels surrounding the brain's smaller arteries and veins and are externally limited by astrocytic end-feet (Wardlaw et al., 2020).In the glymphatic system and the IPAD, peri-arterial and peri-venous spaces might represent channels of CSF influx and ISF efflux, respectively (Rennels et al., 1985), contributing to draining brain waste metabolites (Rasmussen et al., 2022).It is also established that PVS are related to the BBB, the perineural lymphatic drainage, and the newly characterized meningeal lymphatic vessels (mLVs).We think it is fair to say that the PVS serve as critical scaffolds organizing solute exchange and clearance throughout brain tissue.In addition, PVS contain resident and migratory scavenger cells, which contribute to removing brain waste (Nimmerjahn et al., 2005).MRI-visible enlarged perivascular spaces (EPVS) indicates that excessive accumulation or failure to discharge the ISF increases the quantity of various wastes which clogs the PVS, resulting in remodeling and alteration of the fluid-clearance rate.For example, because the aggregation of C-reactive protein and inflammatory cells in PVS may lead to the remodeling and alteration of the fluid-clearance rate, some studies have proposed that inflammatory response may lead to the development of CSVD by triggering PVS dysfunction (Aribisala et al., 2014).What's worse, it can form a vicious cycle that further leads to toxin accumulation, hypoxia, and tissue damage.

Astrocytes AQP4
Aquaporin 4 (AQP4),a regulator of the transcellular transport of water, is one of 14 aquaporins only found on astrocytes and is primarily found in their end-feet facing the vasculature (Mader and Brimberg, 2019).Thespecific expression of AQP4 plays a key role not only in the perivascular CSF-ISF exchange in the glymphatic systems (Gomolka et al., 2023), but also in the preservation of glia limitans integrity at the BBB (Kaur et al., 2006).Loss of perivascular AQP4 localization is a common feature in the setting of aging and a number of neurological conditions, associated with impaired perivascular CSF-ISF exchange and slowed clearance of interstitial solutes (Kress et al., 2014;Wang et al., 2017), which can lead to neuroinflammation, demyelination, and neuron loss (Simon and Iliff, 2016), overlapping with the pathological characteristics of CSVD.Interestingly, recent studies have demonstrated that AQP4 supports circadian rhythms that can control brain clearance efficiency (Hablitz et al., 2020), so exploring how circadian timing affects brain's dynamics through AQP4 will help us understand the fundamental process of waste clearance regulation in the brain.

Transport across the blood brain barrier, blood-cerebrospinal fluid barrier
The CNS is protected against harmful substances contained in the blood by the BBB and the BCSFB (Hladky and Barrand, 2014).The clearance of some peptides, proteins and neurotoxins from the brain occurs via active transport at these two barriers.Transport and metabolic barrier mechanisms can be deregulated in different CNS diseases such as ischemia, inflammation, and infection, with potential consequences for the resolution of these pathologies.
BBB is comprised of vascular endothelial cells, tight junctions, and the underlying basement membrane.It is a structural and functional separation between circulating blood and CNS, which plays a key role in the precise regulation of neural processes by controlling the permeability of the nervous system.Recent findings suggest that BBB opening is accompanied by activation of metabolic waste clearance in the brain (Semyachkina-Glushkovskaya et al., 2017), specifically, small lipophilic molecules can pass through the BBB via free diffusion, and other molecules can be cleared into the blood directly at the BBB through specific transporter proteins, e.g., the low density lipoprotein receptor-related protein 1 and 2 (LRP-1 and LRP-2), P-glycoprotein (P-gp) and the receptor for advanced glycation end products (RAGE), which is an energydependent process (Pascale et al., 2011).Brain pathology often leads to loosening of the tight junctions, which compromises the integrity of the BBB, and there have been experiments which show that clearance of solutes from the CNS is reduced in the setting of BBB integrity disruption, initiating or exacerbating brain pathology (Bowman et al., 2018).
The BCSFB is localized in the CP of brain ventricles.Here, blood is separated from CSF by the walls of the surface arteries and veins in the leptomeninges, and by the walls of penetrating arterioles and venules surrounded by the PVS in the brain itself.The endothelial cells of these vessels, similar to those of BBB, are connected by tight junctions and constitute a relatively impermeable barrier (Reese and Karnovsky, 1967;van Deurs and Koehler, 1979).However, fenestrated endothelial cells of BCSFB are permeable to blood-borne macromolecules, allowing the entry of macromolecules from the blood into the brain.Specific transport mechanisms employing a variety of transmembrane proteins that enable selective passage of molecules form important components of the BCSFB.These transporters belong mainly to the ATP-binding cassette (ABC) transporter and solute carrier (SLC) super-families, which are crucial for moving metabolites and other harmful wastes out of the CNS (Ghersi-Egea et al., 2018;Dani et al., 2021;Usui et al., 2016).Indeed, studies have found that the changes in expression of the transporter genes at the BCSFB are opposite to those observed at the BBB during aging, and may be a compensatory mechanism to aid the failing BBB clearance pathways (Pascale et al., 2011).
In patients with CSVD, BBB dysfunction is evident even in normallooking white matter and increases with lesion burden (Topakian et al., 2010), which may explain pathological changes such as perivascular cell and protein infiltration, perivascular edema, and secondary axonal and neuronal damage in CSVD.To make matters worse, if BCSFB can't make full use of the compensatory function, neurotoxic substances may infiltrate the brain parenchyma and trigger neuroinflammation, thereby promoting the progression of CSVD.Evidence has shown that these barriers leakage is a possible early pathological event in the development of CSVD-associated brain injury, so measuring barriers function is beneficial in early screening and judging the condition of CSVD (Kerkhofs et al., 2021).However, there has been no comprehensive, brain-wide, tissue-specific in vivo assessment of BBB and BCSFB leakage, so little is known about the sequence of pathological events that lead up to their dysfunction.In addition, existing studies have yet to exhaustively define the influence of physiological regulation, regional variability, and pathology-associated changes in BBB-efflux-transporter expression and function on interstitial solute clearance, and the study of BCSFB is even more ambiguous,requiring deep exploration in experimental animal studies or humans.

Glymphatic-meningeal lymphatic system
The question of how the CNS can effectively clear intracranial waste and maintain fluid and tissue homeostasis in the absence of a functional lymphatic or drainage network has long puzzled scientists and clinicians.Fortunately, the glymphatic system and meningeal lymphatics have recently been characterized, finding a breach and setting up a brand-new area.

Glymphatic system
Glymphatic(glia + lymphatic) system was named for its dependence on glial water channels and its adoption of a clearance function similar to that of the peripheral lymphatic system (Plog and Nedergaard, 2018), which is considered to serve as the main pathway for the discharge of potentially neurotoxic soluble wastes (Boland et al., 2018).Concretely, it removes waste in three successive steps: (1)CSF flows into the PVS; (2) CSF enters the brain parenchyma through AQP4 water channels on the vascular astrocytic end-feet and exchanges with ISF; (3) ISF with waste products leaves the brain parenchyma through AQP4, flows into the cervical lymphatic system and peripheral body fluids along perivenous spaces.Glymphatic system's dysfunction characterized by a failure of solute clearance down-regulates the clearance of inflammatory cells, cytokines, and Aβ, subsequently exacerbating the inflammation burden and thus promoting the development of CSVD (Deane et al., 2003).Glymphatic system also serves multiple other purposes in CSVD-related neurophysiology, such as brain-wide delivery of nutrients (Lundgaard et al., 2015), the circulation and distribution of apolipoprotein E isoforms (Hablitz et al., 2020;Achariyar et al., 2017), and even astrocytic paracrine signaling and lipid molecules (Rangroo Thrane et al., 2013).In addition, glymphatic dysfunction in rats has already been associated with impaired cognitive functioning on behavioral tests (Hsu et al., 2023;Venkat et al., 2017).

Meningeal lymphatic drainage
Meningeal lymphatic vessels (mLVs), the latest addition into the ISF/ CSF outflow system, are found in the dura mater following the venous sinuses at the base of the skull, and are composed of a less ramified network of thin-walled initial lymphatic vessels (Aspelund et al., 2015;Louveau et al., 2015).They align with dural blood vessels and cranial nerves and traverse through the cranial foramen together with the venous sinuses, arteries, and cranial nerves, serving as a direct connection between the intracranial space and extracranial lymphatic vasculature (Louveau et al., 2015).Some lymphatic vessels traverse the cribriform plate toward the olfactory nerve.Fluorescent tracers injected into brain parenchyma and ventricles were observed along the mLVs, which drained primarily into the deep cervical lymph nodes (dcLNs) (Louveau et al., 2015).In contrast, in the absence of mLVs, cerebral macromolecule clearance was attenuated and drainage into dcLNs was almost completely abrogated, suggesting that macromolecules are cleared directly by mLVs to extracranial lymphoid tissue (Aspelund et al., 2015).Meanwhile, emerging studies have reported that they perform the function of CSF absorption and waste clearance, transporting immune cells and soluble substances to the peripheral lymph nodes (Da Mesquita et al., 2018a).

Glymphatic-meningeal connection
Glymphatic system enables bulk and net directional flow of CSF and ISF within the PVS and can drain into the peripheral lymphatic system via mLVs (Aspelund et al., 2015).That is to say, the mLVs may represent the second step in the drainage of the ISF from the brain parenchyma into the periphery after it has been exchanged with the CSF through glymphatic system, which is why they are collectively referred to as the glymphatic-meningeal lymphatic system.Da Mesquita et al. also showed that the glymphatic system function is regulated by meningeal lymphatic function (Rangroo Thrane et al., 2013).
The remarkable thing is that the specific pathways through which the CSF/ISF can reach the peripheral lymphatic system are currently an area of great controversy.In addition to mLVs, arachnoid villi or granulations (the outcroppings of arachnoid tissue that project into the dural venous sinuses) has been thought to serve as an outflow route through glymphatic system for decades (d'Avella et al., 1983), but no anatomical mechanism for how this process occurs has been settled upon.Some authors have suggested that they may only act as pressure valves that open to CSF/ISF flow under increased intracranial pressure (Coles et al., 2017).Another alternative mechanism is the perineural routes, such as the olfactory route through the cribriform plate, optic route, and some other cranial nerves are also in the process of exploring (Brierley and Field, 1948;Brierley, 1950;Pile-Spellman et al., 1984;Walter et al., 2006).In order to seek an improved understanding of the clearance pathways to the lymphatic system and the potential implications of these connections to the periphery for multiple neurological disorders, we need to establish a method of sensitively and specifically imaging the egress of CSF.
The impairment of glymphatic-meningeal lymphatic system drainage, which can induce waste aggregation in the brain, has been shown to be associated with CSVD-related risk factors, such as aging, hypertension, diabetes, and lipids metabolism (Tian et al., 2022).In rodent models, a mutually toxic effect of waste deposition and glymphatic and meningeal lymphatic functions has been shown (Da Mesquita et al., 2018b;Ding et al., 2021).Accordingly, we postulate that waste deposition in CSVD is not only a consequence of decreased glymphaticmeningeal lymphatic clearance but also a contributor to this process.Given that many studies have demonstrated that the glymphatic-mLVs failure is a prerequisite for the onset of dementia in neurodegenerative diseases such as Alzheimer's and Parkinson's disease (Da Mesquita et al., 2021;Harrison et al., 2020), we believe that turning attention to the function of the glymphatic-mLVs may lead to a breakthrough in understanding the interaction between CSVD and neurodegenerative diseases.

The intramural periarterial drainage pathway
Peri-and para-vascular clearance also occur in the IPAD pathway (Weller et al., 2008;Carare et al., 2014), which is similar and partially overlapped with the perivenous ISF clearance pathway of the glymphatic-meningeal lymphatic system.The difference is that in the IPAD pathway, after the exchange of CSF and ISF, ISF enter basement membrane (BM) of capillaries wall and drain out of the brain along BM in the tunica media of arterioles and arteries walls, driven by the vasomotion generated by rhythmic contraction and dilation of the arterial SMC (Albargothy et al., 2018), and then drains into the cervical lymph nodes (Morris et al., 2016).
The IPAD pathway and glymphatic-meningeal lymphatic system may cooperatively play a significant role in lymphatic drainage of brain waste, although the latter seems to have received more attention.Note that BM and cell components surrounding cortical arteries are compacted so that there is no PVS transporting CSF or ISF, which contradicts the statement in the glymphatic system (MacGregor Sharp et al., 2019), in which case the IPAD is all the more important.In addition, several experiments showed that the distribution of soluble tracers injected into the brain interstitium within the arterial wall along the IPAD pathway resembles the pathological deposition of Aβ in CAA (Arbel-Ornath et al., 2013;Preston et al., 2003), which indicates that the IPAD pathway has important implications in the pathogenesis of CAA.

Interrelation of various clearance pathways and their effect on CSVD progression
The characteristics of each clearance system have been studied extensively, and there is no doubt that they must work in symbiosis to ensure the proper removal of waste.Their functional abnormalities may lead to the superposition of brain waste accumulation, which can contribute to the progression of CSVD (Fig. 2).
Transport across barriers and lymphatic drainage have the same purpose in clearing interstitial solutes such as Aβ from the brain and in that sense likely serve complementary roles and seem to be partially overlapping mechanisms.If the amount of waste exceeds the efflux transporter capacity or the distance to the BBB or BCSFB is too large, it must be cleared through the ISF flow in the lymphatic drainage.The lymphatic convective flow can also stimulate barrier transport as the convective flow spreads wastes over a wider anatomical area and moves it more toward the BBB or BCSFB,thereby indirectly increasing transport efficiency.

Fig. 2.
Brain-waste-clearance dysfunction interacts with cerebral small vessel disease.Brain-waste-clearance system dysfunction has been gradually recognized as a key link in cerebral small vessel pathology, and the accumulation of metabolic waste plays an important role in the vicious circle of both.
It is precisely because of the close interaction between the various clearance mechanisms that the destruction of any mechanism may trigger a chain reaction of a series of events, which lead to the excessive accumulation of waste.BBB or BCSFB disintegration leads to the accumulation of blood-derived components in PVS, as well as the accumulation of plasma proteins along the vessel wall, serving as a physical impediment for clearance along these pathways, and the next likely step might leadto the stagnation of CSF influx and CSF plus ISF efflux (Weller et al., 2018).On the other hand, lymphatic drainage dysfunction will affect the cell function in the BBB and further lead to the impairment of the microstructure and pulsation of the surrounding cerebral arteries.
However, most of the current studies of brain-waste-clearance systems have thus far been restricted to animal models, and their specific anatomy and relative functional importance in have not been assessed yet.For example, the exact anatomical boundaries of PVS and the precise localization of mLVs have not been confirmed prior to highresolution in vivo imaging, nor is there sufficient evidence that chronic dysregulation of the brain-waste-clearance function is responsible for abnormal Aβ deposition in Alzheimer's disease and CAA.Therefore, future research should focus on addressing these issues, as well as improving the pathology-imaging correlations, and testing the main promising treatments.

Practical and clinical implications
Brain-waste-clearance dysfunction can occur years or even decades before the hallmark symptoms of neurological disorder manifest in presymptomatic individuals (Sperling et al., 2011), and continually promote and accelerate further deterioration of neurological function.It means that the monitoring and intervention aimed at waste clearance dysfunction may provide a new basis for evaluating the progression and prognosis of CSVD and formulating treatment strategies in the future.According to the framework established by us, specific biomarkers or imaging techniques can be explored based on these two mechanisms to better identify these dysfunctions, and take advantage of their overlapping or complementary components to develop efficient and individualized treatment regimens (Fig. 3).

Evaluating clearance function by imaging techniques
The visualization of waste clearance in the brain might be useful for evaluating disease progression in CSVD pathobiology, and the constantly evolving imaging techniques are important tools for evaluating waste clearance efficacy.
MRI quantification of subtle BBB dysfunction would be very useful for the early detection of CSVD.Dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) provides at present the most promising means for quantifying the regional BBB permeability (Thrippleton et al., 2019;Heye et al., 2016).But the technique is relatively immature, so basic research is still needed to advance or even replace current techniques, in order to improve the precision, accuracy, and feasibility of imaging BBB dysfunction.
Many researchers have also attempted to visualize and evaluate the function and structure of the lymphatic system in vivo using MRI, but precisely modelling to achieve quantitative measurements remains challenging (Jiang, 2019).As discussed above, the glymphatic system starts with CSF flowing through the PVS, so most recent advances are focused on imaging CSF flow or water diffusivity in the PVS.For instance, the emerging technique of diffusion tensor image analysis along the perivascular space (DTI-ALPS), a method to measure water diffusivity along the PVS in the human brain is promising for evaluating the activity of the glymphatic system (Zhang et al., 2021), but the correlation of ALPS-index with human glymphatic function has not yet been substantially and rigorously validated by pathophysiological studies so its utility in large queues warrants further testing.
In addition, the present data support the idea that the severity of EPVS, should be itself considered as a MRI marker of CSVD in the elderly (Zhu et al., 2010), which can be assessed using either visual rating scales or more sophisticated quantitative neuroimaging approaches on MRI (Paradise et al., 2020).More importantly, high degree of EPVS is associated with an increased risk of CSVD-induced cognitive impairment and dementia.Scientists have been trying to explore advanced techniques for more sensitive and accurate measurements of PVS and a lot of progress has been made thus far, such as 3D regression neural networks and 3D deep learning frameworks (Dubost et al., 2019;Williamson et al., 2022).Barisano et al. has comprehensively described the applications, advantages, and limitations of the latest technological developments (Barisano et al., 2022).However, it needs to be clarified whether the EPVS also reflects an increased ability to remove potentially neurotoxic metabolites from drainage.

Lifestyle changes and interventions to reduce the risk factors
In the early stages of impaired brain clearance, lifestyle changes and  interventions to reduce the risk factors, such as physical activity, nutritional intervention, and management of hypertension and diabetes mellitus, may be effective for the improvement or prevention of CSVD.In addition, sleep plays a prominent role in resistance to CSVD by improving brain-clearance efficiency.Thus, sleep may represent a key modifiable risk factor in the preclinical phases of CSVD, and has potential value in follow-up treatment.
First, research indicates that the clearance by PVS is enhanced during sleep (Berezuk et al., 2015), so sleep deprivation contributed to the accumulation of metabolic wastes and toxins in PVS and leads to dramatic glymphatic and meningeal lymphatic impairment.Second, during deep sleep the CSF production increases and the ISF volume expands by 60% via AQP4 compared with wakefulness, resulting in faster waste clearance (Xie et al., 2013), and lateral supine position of the head during sleep also seems to promote the lymphatic drainage efficiency (Lee et al., 2015).Finally, BBB permeability is dynamically controlled by circadian rhythms and sleep (Medina-Flores et al., 2020), so chronotherapy based on them may promote the clearance of metabolites along the BBB (Cuddapah et al., 2019).
To sum up, sleep provides an excellent landing pad for intervention and treatment of abnormalities in CSVD-related clearance mechanisms, as adhering to a healthy sleep pattern is a relatively simple way to maintain vascular health.Adequate sleep duration, consistent sleep patterns, avoidance of sleep fragmentation, and treatment of sleep disorders may constitute general recommendations for future interventions and treatments for CSVD by optimizing brain-waste clearance.

Development of new drugs targeting clearance mechanisms
Advance research on these clearance mechanisms contributes to pinpointing drug targets and providing an optimal strategy for CSVD.
Reducing BBB leakage may prevent the accumulation of permanent brain damage by improving the clearance of metabolic wastes and toxins in CSVD, and in this regard, the efficacy of drugs such as pinocembrin and baicalin have been demonstrated recently (Chen et al., 2018;Ma et al., 2018).Besides, therapy against APOE4 may be one of the ways to inhibit the damaging effect of glial cells on BBB (Jackson et al., 2022;Oue et al., 2023).In addition, several interesting potential therapies have also been explored to ameliorate the dysfunction of glymphatic system, such as cilostazol and some other phosphodiesterase inhibitors (Saito and Ihara, 2014), but further experimental studies are needed to confirm their efficiency.However, few studies targeting IPAD and the mLVs have been reported for the treatment of CSVD, and further exploration of the key regulatory sites and modulation mechanisms may offer improvements.
AQP4 is also a target of current interest for both improving drainage clearance function and transport across barriers.Potential treatment strategies include inhibition of AQP4 function, regulation of AQP4 expression and restoration of AQP4 polarization.For example, microRNA-29b overexpression can reduce BBB disruption after ischemic stroke via downregulating AQP4 expression (Wang et al., 2015;Ma et al., 2022).Some small molecule inhibitors have shown promise in the field of AQP4 regulation, such as acetazolamide and TGN-020 (Vandebroek and Yasui, 2020), but further studies are required to determine whether they can be applied to patients with CSVD.Interesting, statins, frequently used to treat CSVD, have also been shown to reduce reactive astrocyte proliferation and preserve the perivascular polarization of the AQP4 channel (Iliff et al., 2014); however, this pathway also requires more exploration of the potential molecular mechanisms involved.
Furthermore, the identification of genetic markers that predict increased susceptibility to removal efficiency by monitoring treatment response and the rapid development of genome-wide association studies and bioinformatics may provide us with new ideas and solutions.

Conclusions and perspectives
In this article, we provide a comprehensive review of trans-barrier transport and lymphatic drainage clearance, including the transport across the BBB and BCSFB, glymphatic-mLVs system and IPAD pathway.Based on the available evidence, we confirmed that all of them exert a vital role in the pathogenesis and progression of CSVD, and future studies should enhance our understanding of the dynamic interaction among them.Establishing emerging and more advanced neuroimaging approaches to assess directly their clearance efficiency may facilitate the earlier diagnosis of CSVD, while improving their clearance functions may represent an important goal for therapeutic strategies.For the latter, sleep improvement offers an easily implemented strategy, wherein AQP4 is a pivotal target.Our exploration provides inspiration for CSVD pathology studies and an exciting opportunity for early warning and intervention in related nervous system diseases, which will drive new research.

Fig. 1 .
Fig. 1.Overview of the brain-waste-clearance system.Through the (1) blood-brain barrier (BBB) and/or (2) blood-cerebrospinal fluid barrier (BCSFB), some small molecules can be transferred by free diffusion, and molecules that match transporters at the barrier can be transferred by active transport.(3) In the intramural periarterial drainage pathway (IPAD), interstitial fluid (ISF) enters basement membranes (BM) of capillaries wall and drain out of the brain along the BM in the tunica media of arterioles and arteries walls (4) The glymphatic system drains cerebrospinal fluid (CSF) with solutes into the brain via the peri-arterial and spaces, while ISF leave the brain via the peri-venous spaces.Subsequently, CSF /ISF is mainly transported along the meningeal lymphatic vessels (mLVs) into the lymph nodes and extracranial systemic circulation.BBB, blood-brain barrier; BCSFB, blood-cerebrospinal fluid barrier; IPAD, intramural periarterial drainage pathway; CSF, cerebrospinal fluid; ISF, interstitial fluid; BM, basement membrane.

Fig. 3 .
Fig. 3. Approaches and mechanisms for enhancing the brain's waste clearance, which play a key role in the treatment of CSVD.