Daniela Barros

Ferroptosis is a newly identified form of regulated cell death characterized by iron accumulation and lipid peroxidation. Ferroptosis plays an essential role in the pathology of numerous diseases and has emerged as a key area of focus in studies of chronic kidney disease (CKD). CKD is a major public health problem with high incidence and mortality that is characterized by a gradual loss of kidney function over time. The severity and complexity of CKD combined with the limited knowledge of its underlying molecular mechanism(s) have led to increased interest in this disease area. Here, we summarize recent advances in our understanding of the regulatory mechanism(s) of ferroptosis and highlight recent studies describing its role in the pathogenesis and progression of CKD. We further discuss the potential therapeutic benefits of targeting ferroptosis for the treatment of CKD and the major hurdles to overcome for the translation of in vitro studies into the clinic.

Respiratory syncytial virus (RSV) infection is the most frequent cause of hospitalization in pediatric patients. Current systemic treatment and vaccines are not curative and re-infection is often associated with a more drastic incidence of the disease. Baicalin is a flavonoid isolated from Scutellaria baicalensis with potent anti-viral characteristics, namely against RSV. However, its precise mechanism of action remains unclear. Here, using in vitro methods and an in vivo murine model of RSV infection, we showed that baicalin inhibits RSV replication induces translational upregulation of type I interferons (IFNs), IFN-a and IFN-B, and reverses epithelial thickening in lung tissues. Moreover, baicalin inhibits transcription of the RSV nonstructural proteins NS1 and NS2. Molecular docking and surface plasmon resonance-based affinity analysis showed that baicalin also binds to the a-3 helix of the NS1 protein with an affinity constant of 1.119 Å~ 10−5 M. Polysome profiling showed that baicalin inhibits translation of the RSV matrix protein (M) RNA. Baicalin mediates increased release of the ribosomal protein L13a from the large ribosomal subunit, where the extra ribosomal subunit L13a inhibits M RNA translation. These results comprehensively establish the multiple mechanisms by which baicalin induces a potent innate immune response against RSV infection.

Sepsis-associated encephalopathy (SAE) is a diffuse central nervous system (CNS) dysfunction during sepsis, and is associated with increased mortality and poor outcomes in septic patients. Despite the high incidence and clinical relevance, the exact mechanisms driving SAE pathogenesis are not yet fully understood, and no specific therapeutic strategies are available. Regulatory T cells (Tregs ) have a role in SAE pathogenesis, thought to be related with alleviation of sepsis-induced hyper-inflammation and immune responses, promotion of T helper (Th) 2 cells functional shift, neuroinflammation resolution, improvement of the blood-brain barrier (BBB) function, among others. Moreover, in a clinical point of view, these cells have the potential value of improving neurological and psychiatric/mental symptoms in SAE patients. This review aims to provide a general overview of SAE from its initial clinical presentation to long-term cognitive impairment and summarizes the main features of its pathogenesis. Additionally, a detailed overview on the main mechanisms by which Tregs may impact SAE pathogenesis is given. Finally, and considering that Tregs may be a novel target for immunomodulatory intervention in SAE, different therapeutic options, aiming to boost peripheral and brain infiltration of Tregs , are discussed.

Laminin is a heterotrimeric glycoprotein with a key role in the formation and maintenance of the basement membrane architecture and properties, as well as on the modulation of several biological functions, including cell adhesion, migration, differentiation and matrix-mediated signaling. In the central nervous system (CNS), laminin is differentially expressed during development and homeostasis, with an impact on the modulation of cell function and fate. Within neurogenic niches, laminin is one of the most important and well described extracellular matrix (ECM) proteins. Specifically, efforts have been made to understand laminin assembly, domain architecture, and interaction of its different bioactive domains with cell surface receptors, soluble signaling molecules, and ECM proteins, to gain insight into the role of this ECM protein and its receptors on the modulation of neurogenesis, both in homeostasis and during repair. This is also expected to provide a rational basis for the design of biomaterial-based matrices mirroring the biological properties of the basement membrane of neural stem cell niches, for application in neural tissue repair and cell transplantation. This review provides a general overview of laminin structure and domain architecture, as well as the main biological functions mediated by this heterotrimeric glycoprotein. The expression and distribution of laminin in the CNS and, more specifically, its role within adult neural stem cell niches is summarized. Additionally, a detailed overview on the use of full-length laminin and laminin derived peptide/recombinant laminin fragments for the development of hydrogels for mimicking the neurogenic niche microenvironment is given. Finally, the main challenges associated with the development of laminin-inspired hydrogels and the hurdles to overcome for these to progress from bench to bedside are discussed.

Laminin incorporation into biological or synthetic hydrogels has been explored to recapitulate the dynamic nature and biological complexity of neural stem cell (NSC) niches. However, the strategies currently explored for laminin immobilization within three-dimensional (3D) matrices do not address a critical aspect influencing cell-matrix interactions, which is the control over laminin conformation and orientation upon immobilization. This is a key feature for the preservation of the protein bioactivity. In this work, we explored an affinity-based approach to mediate the site-selective immobilization of laminin into a degradable synthetic hydrogel. Specifically, a four-arm maleimide terminated poly(ethylene glycol) (PEG-4MAL) macromer was functionalized with a mono-PEGylated recombinant human N-terminal agrin (NtA) domain, to promote high affinity binding of laminin. Different NtA concentrations (10, 50 and 100 μM) were used to investigate the impact of NtA density on laminin incorporation, hydrogel biophysical properties, and biological outcome. Laminin was efficiently incorporated for all the conditions tested (laminin incorporation > 95%), and the developed hydrogels revealed mechanical properties (average storage modulus (G’) ranging from 187 to 256 Pa) within the values preferred for NSC proliferation and neurite branching and extension. Affinity-bound laminin PEG-4MAL hydrogels better preserve laminin bioactivity, compared to unmodified hydrogels and hydrogels containing physically entrapped laminin, this effect being dependent on NtA concentration. This was evidenced by the 10 μM NtA-functionalized PEG-4MAL gels incorporating laminin that support enhanced human NSC proliferation and neurite extension, compared to the latter. Overall, this work highlights the potential of the proposed engineered matrices to be used as defined 3D platforms for the establishment of artificial NSC niches and as extracellular matrix-mimetic microenvironments to support human NSC transplantation.

Laminin immobilization into diverse biological and synthetic matrices has been explored to replicate the microenvironment of stem cell niches and gain insight into the role of extracellular matrix (ECM) on stem cell behavior. However, the site-specific immobilization of this heterotrimeric glycoprotein and, consequently, control over its orientation and bioactivity has been a challenge that has limited many of the explored strategies to date. In this work, we established an affinity-based approach that takes advantage of the native high affinity interaction between laminin and the human N-terminal agrin (hNtA) domain. This interaction is expected to promote the site-selective immobilization of laminin to a specific substrate, while preserving the exposure of its key bioactive epitopes. Recombinant hNtA (rhNtA) domain was produced with high purity (> 90%) and successfully conjugated at its N-terminal with a thiol-terminated poly(ethylene glycol) (PEG) without affecting its affinity to laminin. Self-assembled monolayers (SAMs) of mono-PEGylated rhNtA on gold (mPEG rhNtA-SAMs) were then prepared to evaluate the effectiveness of this strategy. The site-specific immobilization of laminin onto mPEG rhNtA-SAMs was shown to better preserve protein bioactivity in comparison to laminin immobilized on SAMs of thiol-PEG-succinimidyl glutaramide (HS-PEG-SGA), used for the non-selective covalent immobilization of laminin, as evidenced by its enhanced ability to efficiently self-polymerize and mediate cell adhesion and spreading of human neural stem cells. These results highlight the potential of this novel strategy to be used as an alternative to the conventional immobilization approaches in a wide range of applications, including engineered coatings for neuroelectrodes and cell culture, as well as biofunctionalization of 3D matrices.