Search Results (13)

Background and Objectives: Alzheimer’s disease (AD) is a complex neurodegenerative disorder marked by cholinergic deficits, oxidative stress, amyloid-β (Aβ) aggregation, and tau hyperphosphorylation. Caffeic acid (CA), a naturally occurring hydroxycinnamic acid, has emerged as a promising neuroprotective candidate due to its antioxidant, anti-inflammatory, and enzyme-inhibitory properties. This review systematically evaluates recent in vitro and in vivo evidence regarding the therapeutic potential of CA in AD models and examines the impact of delivery systems and derivatives on its efficacy and bioavailability. Materials and Methods: A systematic literature search was conducted in the PubMed, Scopus, and Web of Science databases, adhering to the PRISMA 2020 guidelines. Studies published between January 2021 and April 2025 were included in this review. Eligible studies investigated the effects of CA or CA-enriched extracts on AD-relevant mechanisms using in vitro, in vivo, and in silico models. After screening 101 articles, 44 met the inclusion criteria and were included in the final qualitative synthesis of the study. Results: In vitro studies have confirmed that CA modulates cholinergic activity by inhibiting AChE and BChE and exerting antioxidant and anti-amyloidogenic effects. In vivo studies using pharmacological, genetic, and metabolic AD models have demonstrated improvements in cognitive function, reduction in oxidative stress, inflammation, and Aβ and tau pathologies following CA administration. Advanced delivery platforms, such as solid lipid nanoparticles, transferrin-functionalized liposomes, and carbon dot systems, have significantly enhanced CA’s brain bioavailability and therapeutic efficacy. CA derivatives, including caffeic acid phenethyl ester and nitro-substituted analogs, exhibit improved pharmacokinetic and neuroprotective profiles. Conclusions: This review provides evidence supporting the use of CA as a promising multitarget agent against AD pathology. Its therapeutic potential is further enhanced by nanotechnology-based delivery systems and chemical modifications that overcome the limitations of bioavailability. Continued preclinical evaluation and translational studies are warranted to support its clinical development as an AD intervention. Full article

Background/Objectives: The limited translatability of preclinical experimental findings to patients remains an obstacle for successful treatment of brain diseases. Relevant models to elucidate mechanisms behind brain pathogenesis, including cell-specific contributions and cell-cell interactions, and support successful targeting and prediction of drug responses in humans are urgently needed, given the species differences in brain and blood-brain barrier (BBB) functions. Human microphysiological systems (MPS), such as Organ-Chips, are emerging as a promising approach to address these challenges. Here, we examined and advanced a Brain-Chip that recapitulates aspects of the human cortical parenchyma and the BBB in one model. Methods: We utilized human primary astrocytes and pericytes, human induced pluripotent stem cell (hiPSC)-derived cortical neurons, and hiPSC-derived brain microvascular endothelial-like cells and included for the first time on-chip hiPSC-derived microglia. Results: Using Tumor necrosis factor alpha (TNFα) to emulate neuroinflammation, we demonstrate that our model recapitulates in vivo-relevant responses. Importantly, we show microglia-derived responses, highlighting the Brain-Chip’s sensitivity to capture cell-specific contributions in human disease-associated pathology. We then tested BBB crossing of human transferrin receptor antibodies and conjugated adeno-associated viruses. We demonstrate successful in vitro/in vivo correlation in identifying crossing differences, underscoring the model’s capacity as a screening platform for BBB crossing therapeutic strategies and ability to predict in vivo responses. Conclusions: These findings highlight the potential of the Brain-Chip as a reliable and time-efficient model to support therapeutic development and provide mechanistic insights into brain diseases, adding to the growing evidence supporting the value of MPS in translational research and drug discovery. Full article

Cerebrospinal fluid (CSF) plays an important role in the homeostasis of the brain. We previously reported that CSF major glycoproteins are biosynthesized in the brain, i.e., lipocalin-type prostaglandin D2 synthase (L-PGDS) and transferrin isoforms carrying unique glycans. Although these glycoproteins are secreted from distinct cell types, their CSF levels have been found to be highly correlated with each other in cases of neurodegenerative disorders. The aim of this study was to examine these marker levels and their correlations in other neurological diseases, such as depression and schizophrenia, and disorders featuring abnormal CSF metabolism, including spontaneous intracranial hypotension (SIH) and idiopathic normal pressure hydrocephalus (iNPH). Brain-derived marker levels were found to be highly correlated with each other in the CSF of depression and schizophrenia patients. SIH is caused by CSF leakage, which is suspected to induce hypovolemia and a compensatory increase in CSF production. In SIH, the brain-derived markers were 2–3-fold higher than in other diseases, and, regardless of their diverse levels, they were found to be correlated with each other. Another abnormality of the CSF metabolism, iNPH, is possibly caused by the reduced absorption of CSF, which secondarily induces CSF accumulation in the ventricle; the excess CSF compresses the brain’s parenchyma to induce dementia. One potential treatment is a “shunt operation” to bypass excess CSF from the ventricles to the peritoneal cavity, leading to the attenuation of dementia. After the shunt operation, marker levels began to increase within a week and then further increased by 2–2.5-fold at three, six, and twelve months post-operation, at which point symptoms had gradually attenuated. Notably, the marker levels were found to be correlated with each other in the post-operative period. In conclusion, the brain-derived major glycoprotein markers were highly correlated in the CSF of patients with different neurological diseases, and their correlations were maintained even after surgical intervention. These results suggest that brain-derived proteins could be biomarkers of CSF production. Full article

The glycosylation of proteins plays an important role in neurological development and disease. Glycoproteomic studies on cerebrospinal fluid (CSF) are a valuable tool to gain insight into brain glycosylation and its changes in disease. However, it is important to consider that most proteins in CSFs originate from the blood and enter the CSF across the blood–CSF barrier, thus not reflecting the glycosylation status of the brain. Here, we apply a glycoproteomics method to human CSF, focusing on differences between brain- and blood-derived proteins. To facilitate the analysis of the glycan site occupancy, we refrain from glycopeptide enrichment. In healthy individuals, we describe the presence of heterogeneous brain-type N-glycans on prostaglandin H2-D isomerase alongside the dominant plasma-type N-glycans for proteins such as transferrin or haptoglobin, showing the tissue specificity of protein glycosylation. We apply our methodology to patients diagnosed with various genetic glycosylation disorders who have neurological impairments. In patients with severe glycosylation alterations, we observe that heavily truncated glycans and a complete loss of glycans are more pronounced in brain-derived proteins. We speculate that a similar effect can be observed in other neurological diseases where a focus on brain-derived proteins in the CSF could be similarly beneficial to gain insight into disease-related changes. Full article

Iron is an essential nutrient in the body. However, iron generates oxidative stress and hence needs to be bound to carrier proteins such as the glycoprotein transferrin (Tf) in body fluids. We previously reported that cerebrospinal fluid contains Tf glycan-isoforms that are derived from the brain, but their origins at the cellular level in the brain have not yet been elucidated. In the present report, we described the localization of Tf protein and mRNA in mouse and human brain tissue. In situ hybridization of mouse brain tissue revealed that Tf mRNA is expressed by different cell types such as epithelial cells in the choroid plexus, oligodendrocyte-like cells in the medulla, and neurons in the cortex, hippocampus, and basal ganglia. In contrast, Tf protein was barely detected by immunohistochemistry in hippocampal and some cortical neurons, but it was detected in other types of cells such as oligodendrocyte-like cells and choroid plexus epithelial cells. The results showed that Tf mRNA is expressed by neural cells, while Tf protein is expressed in different brain regions, though at very low levels in hippocampal neurons. Low Tf level in the hippocampus may increases susceptibility to iron-induced oxidative stress, and account for neuron death in neurodegenerative diseases. Full article

The cerebrospinal fluid (CSF) plays an important role in homeostasis of the brain. We previously demonstrated that major CSF proteins such as lipocalin-type prostaglandin D2 synthase (L-PGDS) and transferrin (Tf) that are biosynthesized in the brain could be biomarkers of altered CSF production. Here we report that the levels of these brain-derived CSF proteins correlated well with each other across various neurodegenerative diseases, including Alzheimer’s disease (AD). In addition, protein levels tended to be increased in the CSF samples of AD patients compared with the other diseases. Patients at memory clinics were classified into three categories, consisting of AD (n = 61), mild cognitive impairment (MCI) (n = 42), and cognitively normal (CN) (n = 23), with MMSE scores of 20.4 ± 4.2, 26.9 ± 1.7, and 29.0 ± 1.6, respectively. In each category, CSF protein levels were highly correlated with each other. In CN subjects, increased CSF protein levels correlated well with those of AD markers, including amyloid-β and tau protein, whereas in MCI and AD subjects, correlations declined with AD markers except p-tau. Future follow-up on each clinical subject may provide a clue that the CSF proteins would be AD-related biomarkers. Full article

A series of studies indicated that iron distribution that partly derives from transferrin-bound iron in the peripheral nervous system in the brain may act in processes such as myelination and brain development. However, the relationship between schizophrenia, its psychotic symptoms, and the transferrin (TF) gene has not been systematically explored. Our study aimed to investigate how a particular polymorphism of the transferrin gene, rs3811655, affects the superoxide dismutase (SOD), malondialdehyde (MDA), psychotic symptoms, cognition, or the mediation model between antioxidant enzymes and cognition via symptoms. A total of 564 patients with chronic schizophrenia and 468 healthy control subjects were recruited. The psychotic symptoms and cognition were assessed by the Positive and Negative Syndrome Scale (PANSS) and the Repeatable Battery for the Assessment of Neuropsychological Status (RBANS), respectively. Furthermore, the serum SOD, MDA activity, and transferrin gene polymorphism were measured in patients. Our results demonstrated that patients with the G allele possessed more severe negative symptoms, worse cognitive performance with respect to attention, and higher serum Mn-SOD activity. Additionally, the rs3811655 polymorphism may act as a moderator in the association between Cu/Zn-SOD activity and cognition, as well as psychotic symptoms in patients suffering from schizophrenia. According to this study, the single nucleotide polymorphism (SNP) rs3811655 polymorphism may fail to contribute to the susceptibility of schizophrenia in an individual but is involved in the iron-induced oxidative stress disturbance and cognitive impairment in schizophrenia. This deepens our understanding of the critical role of iron-induced oxidative stress that might underlie the pathophysiology of schizophrenia. Full article

A major bottleneck in the successful development of central nervous system (CNS) drugs is the discovery and design of molecules that can cross the blood-brain barrier (BBB). Nano-delivery strategies are a promising approach that take advantage of natural portals of entry into the brain such as monoclonal antibodies (mAbs) targeting endogenous BBB receptors. However, the main selected mAbs rely on targeting broadly expressed receptors, such as the transferrin and insulin receptors, and in selection processes that do not fully mimic the native receptor conformation, leading to mistargeting and a low fraction of the administered dose effectively reaching the brain. Thus, there is an urgent need to identify new BBB receptors and explore novel antibody selection approaches that can allow a more selective delivery into the brain. Considering that in vitro models fail to completely mimic brain structure complexity, we explored an in vivo cell immunization approach to construct a rabbit derived single-domain antibody (sdAb) library towards BBB endothelial cell receptors. The sdAb antibody library was used in an in vivo phage display screening as a functional selection of novel BBB targeting antibodies. Following three rounds of selections, next generation sequencing analysis, in vitro brain endothelial barrier (BEB) model screenings and in vivo biodistribution studies, five potential sdAbs were identified, three of which reaching >0.6% ID/g in the brain. To validate the brain drug delivery proof-of-concept, the most promising sdAb, namely RG3, was conjugated at the surface of liposomes encapsulated with a model drug, the pan-histone deacetylase inhibitor panobinostat (PAN). The translocation efficiency and activity of the conjugate liposome was determined in a dual functional in vitro BEB-glioblastoma model. The RG3 conjugated PAN liposomes enabled an efficient BEB translocation and presented a potent antitumoral activity against LN229 glioblastoma cells without influencing BEB integrity. In conclusion, our in vivo screening approach allowed the selection of highly specific nano-antibody scaffolds with promising properties for brain targeting and drug delivery. Full article

Herein, GSH-sensitive hyaluronic acid-poly(lactic-co-glycolic acid) (HA-SS-PLGA) was synthesized. Surface modification of PLGA with hyaluronic acid produced a highly stable micelle at physiological pH while a micelle was destabilized at a higher GSH level. Fluorescence microscopy results showed that rhodamine-encapsulated micelle was taken up by brain cancer cells, while competitive inhibition was observed in the presence of free HA and free transferrin. In vitro cytotoxicity results revealed that transferrin-targeted nanoformulated AUY922 (TF-NP-AUY922) shows higher cytotoxicity than either free AUY922 or non-targeted AUY922-loaded micelles (NP-AUY922). In comparison to the control groups, free AUY922, TF-NP-AUY922 or NP-AUY922 treatment revealed the upregulation of HSP70, while the expression of HSP90 client proteins was simultaneously depleted. In addition, the treatment group induced caspase-dependent PARP cleavage and the upregulation of p53 expression, which plays a key role in apoptosis of brain cancer cells. In vivo and ex vivo biodistribution studies showed that cypate-loaded micelle was taken up and accumulated in the tumor regions. Furthermore, in vivo therapeutic efficacy studies revealed that the AUY922-loaded micelle significantly suppressed tumor growth in comparison to the free AUY922, or control groups using tumor-bearing NOD-SCID mice. Moreover, biochemical index and histological analysis revealed synthesized micelle does not show any significant cytotoxicity to the selected major organs. Overall, a synthesized micelle is the best carrier for AUY922 to enhance the therapeutic efficiency of brain cancer. Full article

Alzheimer’s disease (AD) is a fatal neurodegenerative disorder caused by protein misfolding and aggregation, affecting brain function and causing dementia. Amyloid beta (Aβ), a peptide deriving from amyloid precursor protein (APP) cleavage by-and γ-secretases, is considered a pathological hallmark of AD. Our previous study, together with several lines of evidence, identified a strict link between APP, Aβ and 37/67kDa laminin receptor (LR), finding the possibility to regulate intracellular APP localization and maturation through modulation of the receptor. Here, we report that in fibroblasts from familial AD (fAD), APP was prevalently expressed as an immature isoform and accumulated preferentially in the transferrin-positive recycling compartment rather than in the Golgi apparatus. Moreover, besides the altered mitochondrial network exhibited by fAD patient cells, the levels of pAkt and pGSK3 were reduced in respect to healthy control fibroblasts and were accompanied by an increased amount of secreted Aβ in conditioned medium from cell cultures. Interestingly, these features were reversed by inhibition of 37/67kDa LR by NSC47924 a small molecule that was able to rescue the “typical” APP localization in the Golgi apparatus, with consequences on the Aβ level and mitochondrial network. Altogether, these findings suggest that 37/67kDa LR modulation may represent a useful tool to control APP trafficking and Aβ levels with implications in Alzheimer’s disease. Full article

The non-human primate (NHP)-brain endothelium constitutes an essential alternative to human in the prediction of molecule trafficking across the blood–brain barrier (BBB). This study presents a comparison between the NHP transcriptome of freshly isolated brain microcapillaries and in vitro-selected brain endothelial cells (BECs), focusing on important BBB features, namely tight junctions, receptors mediating transcytosis (RMT), ABC and SLC transporters, given its relevance as an alternative model for the molecule trafficking prediction across the BBB and identification of new brain-specific transport mechanisms. In vitro BECs conserved most of the BBB key elements for barrier integrity and control of molecular trafficking. The function of RMT via the transferrin receptor (TFRC) was characterized in this NHP-BBB model, where both human transferrin and anti-hTFRC antibody showed increased apical-to-basolateral passage in comparison to control molecules. In parallel, eventual BBB-related regional differences were Investig.igated in seven-day in vitro-selected BECs from five brain structures: brainstem, cerebellum, cortex, hippocampus, and striatum. Our analysis retrieved few differences in the brain endothelium across brain regions, suggesting a rather homogeneous BBB function across the brain parenchyma. The presently established NHP-derived BBB model closely mimics the physiological BBB, thus representing a ready-to-use tool for assessment of the penetration of biotherapeutics into the human CNS. Full article

Tubulinopathies are rare neurological disorders caused by alterations in tubulin structure and function, giving rise to a wide range of brain abnormalities involving neuronal proliferation, migration, differentiation and axon guidance. TUBB is one of the ten β-tubulin encoding genes present in the human genome and is broadly expressed in the developing central nervous system and the skin. Mutations in TUBB are responsible for two distinct pathological conditions: the first is characterized by microcephaly and complex structural brain malformations and the second, also known as “circumferential skin creases Kunze type” (CSC-KT), is associated to neurological features, excess skin folding and growth retardation. We used a combination of immunocytochemical and cellular approaches to explore, on patients’ derived fibroblasts, the functional consequences of two TUBB variants: the novel mutation (p.N52S), associated with basal ganglia and cerebellar dysgenesis, and the previously reported variant (p.M73T), linked to microcephaly, corpus callosum agenesis and CSC-KT skin phenotype. Our results demonstrate that these variants impair microtubule (MT) function and dynamics. Most importantly, our studies show an altered epidermal growth factor (EGF) and transferrin (Tf) intracellular vesicle trafficking in both patients’ fibroblasts, suggesting a specific role of TUBB in MT-dependent vesicular transport. Full article

Magnetic nanoparticles have great prospects for drug delivery purposes, as they can be designed with various surface coatings and conjugated with drugs and targeting moieties. They also have a unique potential for precise delivery when guided by magnetic force. The blood-brain barrier (BBB) denotes the interface between the blood and brain parenchyma and hinders the majority of drugs from entering the brain. Red fluorescent magnetic nanoparticles were encapsulated in liposomes and conjugated to antibodies targeting the rat transferrin receptor (OX26) to form magnetic immunoliposomes. These magnetic immunoliposomes enhanced the uptake by rat brain capillary endothelial cells (BCECs) in vitro. In situ brain perfusion in young rats high in the endogenous expression of transferrin receptors by BCECs, revealed enhanced uptake of magnetic immunoliposomes when compared to naked magnetic nanoparticles or non-targeted magnetic liposomes. When applying the external magnetic force, the magnetic nanoparticles were detected in the brain parenchyma, suggesting transport across the BBB. Ultrastructural examination of the immunoliposomes, unfortunately, was unable to confirm a complete encapsulation of all naked nanoparticles within the liposomes, suggesting that the data on the brain could derive from particles being released from the liposomes under influence of external magnetic force; hence hypothesizes on external magnetic force as a qualifier for dragging targeted magnetic immunoliposomes through the BBB. In conclusion, our results suggest that transport of magnetic nanoparticles present in BCECs by targeted delivery to the transferrin receptor may undergo further transport into the brain when applying magnetic force. While magnetic immunoliposomes are targetable to BCECs, their design to enable further transport across the BBB when applying external magnetic force needs further improvement. Full article