BBB disruption and dysfunction in a pathological state prompts the leakage of blood constituents into the CNS, infiltration of various cells, abnormal transfer and clearance of ions and molecules responsible for reduction and dysregulation of cerebral blood flow (CBF), further causing various neurological deficits [66].
Acute neurological diseases due to BBB dysfunction
Bacterial meningitis
Bacterial meningitis refers to inflammation of the subarachnoid space and meninges caused by various bacterial pathogens viz., Streptococcus pneumoniae, Neisseria meningitidis, Haemophilus influenzae, and Escherichia coli [67]. The inflammation also involves the brain cortex and spinal cord [68]. Bacterial invasion in the brain takes place through four processes, which include colonization, bacterial intrusion into the blood flow, survival in blood, and penetration into the subarachnoid space [28]. The consequent inflammation and CNS damage are brought about by a blend of pathogen and host factors. The primary manifestations associated with bacterial meningitis include pyrexia, cephalalgia, migraine, photophobia, and change in consciousness [69].
Blood-borne bacteria can subjugate the meninges through the choroid plexus (CPs) and CNS barrier capillaries which include the arachnoidal, pial, and brain parenchyma microvessels. H. influenzae interacts with the basolateral side of CPs ependymal monolayer and transcytoses to the cytoplasm and is released in ventricles by exocytosis. H. influenzae proficiently adheres to ECs [70]. Group B Streptococcus (GBS) and E. coli K1 are the two main causative agents of neonatal meningitis but both have different strategies of invasion in the subarachnoid space. Some E. coli K1 factors like AB-type toxin, cytotoxic necrotising factor-1 (CNF1), invasion of brain endothelial cells (IBE) aid bacterial invasion of ECs that actuate RHO family GTPases [71]. Interaction of bacterial factors with host cell receptors triggers various host cell signaling proteins [28]. Factors like Outer-Membrane Protein A (OmpA) mediate adhesion through N-acetyl glucosamine of the glycoprotein (GP96), FimH and the lipoprotein NLPI permit adhesion of bacteria to host cells [72]. N. meningitidis and S. pneumoniae have their natural niche in the human nasopharynx, from where they arrive in the respiratory tract and enter the bloodstream and cause septicemia [71]. Virulence factors of N. meningitidis allow its survival in blood and prevent its elimination by host effector cells. Type IV pili (T4P) are essential for adhesion to ECs. N. meningitidis adheres to CNS capillaries and actively crosses BBB by disrupting cell–cell junction in subarachnoid space [28].
Streptococcus pneumoniae can invade into the bloodstream through cavities of inner ear, intravascular space present within the tissues, or lungs. Once S. pneumoniae becomes bloodborne, it can invade the meninges through the olfactory neuron. The virulence factors that enable S. pneumoniae to survive in blood and brain parenchyma include capsule, pneumococcal surface proteins (PsP), and pneumolysin, also common for some other pathogens causing meningitis [73]. Encapsulated S. pneumoniae is more impervious to phagocytosis and facilitates successful colonization. PsP permit the adhesion of bacteria to ECs with the help of pneumococcal pilus-1. Further, Platelet Endothelial Cell Adhesion Molecule (PECAM-1) and poly Immunoglobulin receptor (plgR) promote translocation through BBB [74].
Numerous meningeal pathogens affect BBB integrity by interfering specifically with AJs or TJs. GBS and S. pneumoniae directly secrete a pore-forming toxin, thereby altering BBB integrity. E. coli K1, S. pneumoniae, and GBS enhance the production of nitric oxide from ECs via inducible nitric oxide synthase (iNOS), thus disrupting BBB integrity [75]. Moreover, pathogen-derived toxins enhance inflammatory chemokines or cytokines expression in response to infection by the host and can adversely affect BBB function [76].
The characteristic feature of epilepsy is the fallible and uncontrolled activity of either a part or of the entire CNS. A patient susceptible to epilepsy experiences attacks when excitability of the CNS transcends a certain critical threshold [73]. The main difference between seizure and epilepsy is that the former is a single occurrence while the latter is characterized by two or more provoked seizures [77]. Epilepsy has been categorized as Grand Mal, Petit Mal, and Focal Mal epilepsy. Grand Mal epilepsy is manifested as neuronal discharges in the entire brain. The person becomes unconscious and this state lasts for 3–4 min. Increased voltage and recurrence of electrical signals can occur over the entire cortex [78].
Petit Mal epilepsy is characterized by unconsciousness for 3–30 s and muscle contractions around the head. The brain wave pattern of Petit Mal epilepsy can be demonstrated by spike and dome pattern [79]. In vivo studies suggested it to be resulting from oscillations of inhibition and excitatory thalamic and corticothalamic neurons, which in turn, initiate Grand Mal epilepsy [80].
Focal Mal epilepsy arises from some localized organic lesion or functional abnormality which may be confined to a single area, such as brain scar tissue that pulls the adjacent neuronal tissue; a tumor that squeezes the region of the brain, or congenitally deranged local circuitry [78]. Epilepsy can be caused by strong emotional stimuli, traumatic lesions and alkalosis caused by over-breathing. Other symptoms observed in patients include increased microvascular density, disturbed GABAergic mechanisms [81], loss of TJs, IgG leakage in hippocampal resections [82], a short period of amnesia [83], sudden anxiety, discomfort, attack of abnormal rage, jerks, shock movements and difficulty in breathing [84].
BBB dysfunction emphatically relates with seizure frequency and is not related to neuronal loss. Epileptogenic injuries or seizures initiate the synthesis and secretion of proinflammatory molecules such as TNF-α, IL-1β, and High Mobility Group Box 1 (HMGB1) in glial cells which result in decreasing seizure threshold, which further contributes to seizure precipitation, and recurrence because of rapid changes in glutamate and γ-aminobutyric acid (GABA) receptor phosphorylation. This also leads to channelopathies which change intrinsic neural excitability [85].
Seizures may also lead to BBB disruption, and artificial opening of BBB causes synchronization of rat neuronal activity that leads to albumin and immunoglobulin G (IgG) neuropil eruption. Albumin alters the buffering capability of K+ in astrocytes which contributes to neuronal hyperexcitability as shown in Fig. 3 [88]. TGF-β released from other cell types also affects BBB integrity. Astrocytic TGFβ released enhances plasminogen activator inhibitor-1 (PAI-1), which in turn suppresses tissue plasminogen activator (tPA), resulting in BBB closure [89].
Activation of TNF-α and TGF-β Signaling by albumin during BBB breakdown induces rapid upregulation of genes associated to inflammation considering NF-ĸβ pathways and complimentary cascades, cytokines and chemokines (IL-6, CcL-2, CcD-7, Cd14) [86]. The pro-inflammatory molecules secreted by microglia and astrocytes contribute to voltage fluctuation in different parts of the brain. The figure is adapted from [87]
Traumatic brain injury (TBI)
Traumatic brain injury (TBI), also known as acquired injury on the head/brain due to swift trauma harms the brain and disturbs the normal functioning of the brain, further causing substantial disability and mortality. It usually results from a powerful blow of an object or jolt (external force) to the head. CSF provides buoyancy and protects the brain from external forces, allowing the brain to bounce away from the skull when hit in the head, thus behaving like a shock absorber, but when this shock exceeds the threshold, as in the case of TBI, the levels of inflammatory proteins, like IL-6, IL-8 and IL-10 increase in the CSF [90]. The symptoms include loss of consciousness, post-injury amnesia, headaches, migraine, dizziness, light and sound sensitivity, seizures, blurred vision, anxiety, alterations in aggression, memory lapses, lack of impulse control, impaired decision making, and post-traumatic stress, etc.
BBB restricts the therapeutic compounds from entering the brain. However, in case of brain injury, ruptured microvessels depolarize the barrier which allows blood components to enter immediately into the brain parenchyma [91]. Primary injury caused due to dysfunction like neuroinflammation and cell death further disrupts the walls of these microvessels, initiating coagulation cascade and resulting in secondary damage [92]. Intravascular coagulation limits the supply of blood to tissues, which causes oxygen shortage, leading to ischemia, and thus, blood-borne factors fibrinogen, thrombin, and albumin among others can gain access to the brain [66]. Ischemia damages the blood components and may start inflammatory cascades which hinder the proper functioning of ECs, ECM, pericytes, and astroglia cells. BBB dysfunction due to TBI can occur in two phases: in the initial phase, shear injury of microvessels may occur within hours of TBI. The second phase is the activation of inflammatory cells after 3 days of TBI which trigger alterations in the BBB permeability. In vivo analysis reported the role of BBB breakdown in initiating changes to transcriptional processes of the neurovascular network, further causing degenerative disorders like AD, psychological impairments, cognitive decline, and epilepsy [92].
Chronic neurological diseases
Alzheimer’s disease (AD)
AD is a brain disorder that leads to the progressive degeneration and death of brain cells. It results in a continuous decline in behavioral, thinking, and other social skills disrupting the person’s ability to function independently. Few symptoms of AD are dementia, anxiety, cognitive impairment, restlessness, fatigue, and dizziness, etc. [66]. Several genes like apolipoprotein (APOE4), presenilin-2 (PSEN2), presenilin-1(PSEN1), amyloid-beta precursor protein (APP), microtubule-associated protein tau (MAPT), etc., are also associated with a higher or lower risk of sporadic early or late-onset of AD (Fig. 4) [93]. The hyperphosphorylated tau neurofibrillary tangles (NFTs), Aβ plaques, neuronal loss, and cerebrovascular dysfunction are majorly proposed to contribute to AD pathophysiology and cognitive impairment [94]. NFTs caused by the accumulation of phosphorylated tau protein in the neuron can also lead to AD which is associated with chromosome no. 17q21. Moreover, tau is a microtubule-associated protein that facilitates axonal transport essential for neuronal signaling and trafficking [95]. In the normal brain, each tau molecule contains 2 to 3 phosphates, notwithstanding, the phosphoryl content increases by several folds in tauopathy patients. During hyperphosphorylation, tau protein dissociates from the microtubule, resulting in the spread of unbound microtubules and progressive accumulation of phosphorylated tau protein followed by the formation of NFTs [96]. On the other hand, PSEN1 and PSEN2 are the catalytic components of γ-secretase [97]. Mutations in human PSEN1 have been shown to promote the breakdown of BBB and cerebrovascular dysfunction [98]. PSEN2 mutations represent ~ 5% of all AD cases [99].
Schematic diagram showing mutations in several genes i.e. presenilin-1(PSEN1), presenilin-2 (PSEN2), amyloid-beta precursor protein (APP), apolipoprotein (APOE3), and apolipoprotein 4 (APOE4) related to increase risk of neurological disorders. Alzheimer’s disease (AD) is caused by mutations in the above genes and also by hyperphosphorylation of tau proteins in the distal part of axons in neuronal cells results in NFTs and cell death
Besides, APOE4 is one of the major genetic risk factors leading to sporadic and late-onset AD. APOE4 alleles increase AD risk by four times, compared with APOE3. The cerebrovascular system and neurons suffer toxic effects due to APOE [100]. Human APOE4 carriers may develop progressive BBB breakdown and pericyte degeneration, [101] early neurovascular dysfunction, [102] and reduced glucose uptake by the BBB.
The vascular capillary leakages of proteins like thrombin, IgG, fibrinogen, albumin, and hemosiderin in the hippocampus and prefrontal as well as entorhinal cortex of the AD patients have been observed. Approximately 40 APP mutations and co-localization of proteins with Aβ [103] responsible for causing AD have been identified. This causes cerebrovascular pathology, breakdown of BBB and cerebral amyloid angiopathy (CAA) [104]. CAA is caused by vascular degeneration of smooth muscle cells (SMCs) linked with the breakdown of the BBB at the arterial and arteriolar levels [105]. Besides, APOE4 carriers accelerate BBB breakdown via activation of pathway i.e., proinflammatory cyclophilin A (CypA)-MMP-9. Thus, causing degradation of endothelial TJs, proteins of basement membrane and increasing damage to BBB [66].
Amyotrophic lateral disease (ALS)
Amyotrophic lateral sclerosis (ALS) causes dysfunction of neurons controlling voluntary movements. It is also referred to as motor neuron disease or Lou Gehrig’s disease. The patients suffering from ALS experience loss of hands and arm functions, difficulty in walking, speaking and even breathing. Respiratory failure can often be the reason for death and the average survival time after ALS diagnosis is around three years [106]. Upper Motor Neuron (UMN) and Lower Motor Neuron (LMN) are specific neurons affected by ALS. UMN extends from the cerebral cortex or brain stem and carries motor information, while LMN extends from the brain stem to the skeletal muscles to cause movement [107].
ALS can either be familial or sporadic. The frequency of sporadic ALS is random and accounts for around 90% of cases, while familial ALS is inherited. Around 10% of cases are familial with dominant inheritance of mutation in around 15 genes such as TARDBP (transactive response DNA binding protein), ANG (angiogenin), FUS (RNA binding protein), and OPTN (optineurin), etc. [108]. The respiratory symptoms of the disease include dyspnoea (breathing difficulties), orthopnoea (breathing difficulties while lying flat), weak cough, excessive sleepiness, and cognitive impairment, etc.
The BBB and BSCB restrict the entry of erythrocytes and plasma components into the CNS. The disruption of these barriers has been observed in ALS patients with spinal cord or motor-cortex accumulation of proteins like fibrin and thrombin [109], reduced levels of TJ proteins, erythrocytes, haemoglobin and hemosiderin, which further cause generation of ROS, toxic to motor neurons (Fig. 5). Levels of adhesion molecules such as intercellular adhesion molecule (ICAM-1/2/ vascular cell adhesion molecules (VCAM)) are increased in ALS. The elevated ratio of CSF/serum albumin and reduced expression of BSCB TJs proteins has been reported in both forms of this disease [110]. The key role of pericytes in maintaining blood-CNS barriers has been well understood.
Reduced levels of TJs due to BBB dysfunction lead to pericyte degeneration causing infiltration of antibodies (IgG), intercellular adhesion molecule (ICAM-1/2/vascular cell adhesion molecules (VCAM), thrombin, plasminogen, haemoglobin (Hb) and iron (II) released from RBCs further produce ROS in the matrix, toxic for motor neurons in case of ALS. This figure is adapted from [66]
Huntington’s disease (HD)
Huntington’s disease (HD) is a progressive disorder characterized by flicking movements in the muscles followed by severe distortional movements in the whole body. In addition, severe cognitive problems also develop along with motor dysfunctions. The genetics of HD abnormality involves a repeat sequence of CAG (≥ 36) in exon 1 of chromosome 4 of the HTT gene, producing mutant Huntington protein (mHtt) that aggregates and further causes neurodegeneration [66]. CAG encodes for the amino acid glutamine in the molecular structure of an abnormal neuronal cell protein Huntington responsible for the symptoms; thus, it can be considered a polyglutamine disease. A normal individual has 10–35 repeats of CAG and a higher number of CAG repeats leads to the onset of symptoms in the individual [111].
HD causes gradual degeneration of the basal ganglia, also called the caudate nucleus and putamen [112]. The symptoms associated with HD are chorea (jerky involuntary movements that affect the hips, shoulders, and face), cognitive decline, dystonia (abnormal muscle tone which results in muscular spasm and posture abnormality), and behavioral difficulties [113]. A reduction in expression of proteins like claudin-5 and Occludin involved in the formation of BBB TJs and various markers such as IL-8 and metalloproteinase 1 tissue inhibitor are related to increased BBB permeability, which have been observed in HD patients [114, 115]. BBB breakdown in HD patients has been confirmed by detecting 2.5-fold increase in levels of extravascular fibrin deposition [103] as compared to controls. Recent studies of dynamic contrast-enhanced (DCE)-MRI in HD patients reflects increased BBB permeability in the caudate nucleus as well as an increase in the gray matter cerebral blood level.
Parkinson’s disease (PD)
PD is characterized by the accumulation of oligomeric α-synuclein (α-syn), and dopaminergic neuron degeneration observed in the part of substantia nigra- pars compacta (SNpc) which further prompts motor impairments [116]. Since pars compacta follows a nigrostriatal pathway that contributes in stimulating the cerebral cortex as well as initiating movement, the degeneration of pars compacta neurons ultimately results in a low movement state. The PD patients experience muscular stiffness in the limbs causing difficulty in normal walking, running, etc. Other symptoms are dizziness, cognitive impairments, dementia, loss of postural reflexes, and reduced facial expression.
Vascular dysfunction of the basal ganglia in PD patients leads to breakdown and dysfunction of BBB. In a study, Magnetic resonance imaging (MRI) demonstrated microbleeds, and diminished active efflux of xenobiotics as well as other potential toxins being reported by verapamil-PET [54]. Besides, increased 11C-verapamil uptake in frontal white matter regions was observed in comparison to controls [117]. BBB breakdown leads to the accumulation of neurotoxic fibrinogen, thrombin, plasminogen, and RBC extravasation. The release of Hb and Fe2+ generates ROS harmful to dopaminergic neurons. Proinflammatory cytokines like IL-1β, IFN-γ, and TNF as well as various MMPs, etc., released during neurodegeneration are shown in Fig. 6. Few recent studies highlighted the ability of α-syn to cross the BBB and its contribution in the accumulation of α-syn pool in the brain, further clearance from the brain across the BBB occurs through LRP1-mediated transcytosis.
Key pathological characteristics of Parkinson’s disease (PD) are depicted. These include cell death in the Substantia Nigra and Lewy bodies [aggregation of α-synuclein (α-syn),] affecting dopamine releasing neurons. BBB breakdown leads to accumulation of various blood components, leading to production of ROS and thus affecting dopaminergic astrocytes and neurons in case of PD. This figure is adapted from [118]
Missense mutations in DNA variants such as LRRK2 (Leucine-rich repeat kinase 2) have been associated with late-onset PD (> 50 years) [119]. Additionally, in vivo studies highlighted MDR1 mutations associated with familial as well as sporadic PD. MDR1 which is highly expressed at the BBB endothelium encodes for ABCB1 (P-glycoprotein) which shows decreased expression in the case of PD patients and leads to progression of PD [117].
Multiple sclerosis (MS)
Multiple sclerosis (MS) is a neurodegenerative disease that disrupts the BBB and eventually allows CD4+ T cells, peripheral macrophages, and B cells to enter into CNS. This triggers an inflammatory cascade that results in axonal loss and demyelination [120]. MS leads to demyelination of the nerves and causes inflammation, affecting the spinal cord, brain, and optic nerves of the CNS. Several genes of the Major Histocompatibility Complex (MHC) make the individual susceptible to MS. It has been considered that leukocytes, particularly the T-cells migrate to the BBB [121]. The symptoms include disturbances in motor, sensory, bladder and bowel, optic nerve functions, cognitive impairments, tingling, numbness, vision problems etc. An early BBB breakdown, fibrinogen accumulation, reduced expression of TJs, and endothelial degeneration are the characteristic features of MS. BBB dysregulation and trans-endothelial migration of activated leukocytes are among the earliest cerebrovascular abnormalities observed in MS, which induces inflammatory cytokines/chemokines release [122]. Activation of cerebral endothelial cells (CECs) by Th1 cytokines changes occludin and vascular endothelial cadherin (VE-Cadherin) phenotypes in TJs and AJs of CEC via several inflammatory genes [66]. Since most of the drugs are unable to penetrate the BBB, no effective drug against MS could be designed to date. However, a few years ago, the discovery of a peptide molecule, dNP2 which is permeable to the barrier in conjugation with cytotoxic T cells proved to be effective against the disease [123].
