Editor’s Choice Articles

Efavirenz (Sustiva^®) is a first-generation non-nucleoside reverse transcriptase inhibitor (NNRTI) used to treat human immunodeficiency virus (HIV) type 1 infection or to prevent the spread of HIV. In 1998, the FDA authorized efavirenz for the treatment of HIV-1 infection. Patients formerly required three 200 mg efavirenz capsules daily, which was rapidly updated to a 600 mg tablet that only required one tablet per day. However, when given 600 mg once daily, plasma efavirenz concentrations were linked not only to poor HIV suppression but also to toxicity. Clinical data suggested that the standard dose of efavirenz could be reduced without compromising its effectiveness, resulting in a reduction in side effects and making the drug more affordable. Therefore, ENCORE1 was performed to compare the efficiency and safeness of a reduced dose of efavirenz (400 mg) with the standard dose (600 mg) plus two NRTI in antiretroviral-naïve HIV-infected individuals. Nowadays, due to the emergence of integrase strand transfer inhibitors (INSTIs), some consider that it is time to stop using efavirenz as a first-line treatment on a global scale, in the parts of the world where that is possible. Efavirenz has been a primary first-line antiviral drug for more than 15 years. However, at this moment, the best use for efavirenz could be for pre-exposure prophylaxis (PrEP) and repurposing in medicine. Full article

The creatine precursor Guanidinoacetic Acid (GAA) accumulates in the genetic deficiency of the GuanidinoAcetate Methyl Transferase (GAMT) enzyme and it is believed to cause the seizures that often occur in this condition. However, evidence that it is indeed epileptogenic is scarce and we previously found that it does not cause neuronal hyperexcitation in in vitro brain slices. Here, we used Micro-Electrode Arrays (MEAs) to further investigate the electrophysiological effects of its acute and chronic administration in the networks of cultured neurons, either neocortical or hippocampal. We found that: (1) GAA at the 1 µM concentration, comparable to its concentration in normal cerebrospinal fluid, does not modify any of the parameters we investigated in either neuronal type; (2) at the 10 µM concentration, very similar to that found in the GAMT deficiency, it did not affect any of the parameters we tested except the bursting rate of neocortical networks and the burst duration of hippocampal networks, both of which were decreased, a change pointing in a direction opposite to epileptogenesis; (3) at the very high and unphysiological 100 µM concentration, it caused a decrease in all parameters, a change that again goes in the direction opposite to epileptogenesis. Our results confirm that GAA is not epileptogenic. Full article

The continuous remodeling of the tumor microenvironment (TME) during prostate tumorigenesis is emerging as a critical event that facilitates cancer growth, progression and drug-resistance. Recent advances have identified extensive communication networks that enable tumor–stroma cross-talk, and emphasized the functional importance of diverse, heterogeneous stromal fibroblast populations during malignant growth. Cancer-associated fibroblasts (CAFs) are a vital component of the TME, which mediate key oncogenic events including angiogenesis, immunosuppression, metastatic progression and therapeutic resistance, thus presenting an attractive therapeutic target. Nevertheless, how fibroblast heterogeneity, recruitment, cell-of-origin and differential functions contribute to prostate cancer remains to be fully delineated. Developing our molecular understanding of these processes is fundamental to developing new therapies and biomarkers that can ultimately improve clinical outcomes. In this review, we explore the current challenges surrounding fibroblast identification, discuss new mechanistic insights into fibroblast functions during normal prostate tissue homeostasis and tumorigenesis, and illustrate the diverse nature of fibroblast recruitment and CAF generation. We also highlight the promise of CAF-targeted therapies for the treatment of prostate cancer. Full article

Temoporfin (mTHPC) is approved in Europe for the photodynamic treatment of head and neck squamous cell carcinoma (HNSCC). Although it has a promising profile, its lipophilic character hampers the full exploitation of its potential due to high tendency of aggregation and a reduced ROS generation that compromise photodynamic therapy (PDT) efficacy. Moreover, for its clinical administration, mTHPC requires the presence of ethanol and propylene glycol as solvents, often causing adverse effects in the site of injection. In this paper we explored the efficiency of a new mTHPC formulation that uses human serum albumin (HSA) to disperse the photosensitizer in solution ([email protected]), investigating its anticancer potential in two HNSCC cell lines. Through a comprehensive characterization, we demonstrated that [email protected] is stable in physiological environment, does not aggregate, and is extremely efficient in PDT performance, due to its high singlet oxygen generation and the high dispersion as monomolecular form in HSA. This is supported by the computational identification of the specific binding pocket of mTHPC in HSA. Moreover, [email protected] induces cytotoxicity in both HNSCC cell lines, increasing intracellular ROS generation and the number of γ-H2AX foci, a cellular event involved in the global response to cellular stress. Taken together these results highlight the promising phototoxic profile of the complex, prompting further studies to assess its clinical potential. Full article

The progressively rising food-borne Campylobacter jejuni infections pose serious health problems and socioeconomic burdens. Given that antibiotic therapy is not recommended for most campylobacteriosis patients, novel treatment options include strategies targeting iron homeostasis that impacts both C. jejuni virulence and inflammatory cell damage caused by toxic oxygen species. In our preclinical intervention study, we tested potential disease-alleviating effects upon prophylactic oral application of the iron-chelating compound desferoxamine (DESF) in acute murine campylobacteriosis. Therefore, microbiota-depleted IL-10^−/− mice received synthetic DESF via the drinking water starting seven days before oral infection with C. jejuni strain 81-176. Results revealed that the DESF application did not reduce gastrointestinal pathogen loads but significantly improved the clinical outcome of infected mice at day 6 post-infection. This was accompanied by less pronounced colonic epithelial cell apoptosis, attenuated accumulation of neutrophils in the infected large intestines and abolished intestinal IFN-γ and even systemic MCP-1 secretion. In conclusion, our study highlights the applied murine campylobacteriosis model as suitable for investigating the role of iron in C. jejuni infection in vivo as demonstrated by the disease-alleviating effects of specific iron binding by oral DESF application in acute C. jejuni induced enterocolitis. Full article

The G2019S mutation in the leucine-rich repeat kinase 2 (LRRK2) gene is associated with late-onset Parkinson’s disease (PD). Although PD affects men and women differently, longitudinal studies examining sex- and age-dependent alterations in mice carrying the G2019S mutation are limited. We examined if behavioral and neurochemical dysfunctions, as well as neurodegeneration, occur in male and female BAC LRRK2-hG2019S (G2019S) mice, compared to their age-matched wild type littermates, at four age ranges. In the open field test, hyperlocomotion was observed in 10–12 month old male and 2–4.5 months old female G2019S mice. In the pole test, motor coordination was impaired in male G2019S mice from 15 months of age and in 20–21 months old female G2019S mice. In the striatum of G2019S male and female mice, the amounts of tyrosine hydroxylase (TH), measured with Western blotting, were unaltered. However, we found a decreased expression of the dopamine transporter in 20–21 month old male G2019S mice. The number of TH-positive neurons in the substantia nigra compacta was unaltered in 20–21 month old male and female G2019S mice. These results identify sex- and age-dependent differences in the occurrence of motor and neurochemical deficits in BAC LRRK2-hG2019S mice, and no degeneration of DA neurons. Full article

Three-dimensional (3D) bioprinted skin equivalents are highlighted as the new gold standard for alternative models to animal testing, as well as full-thickness wound healing. In this review, we focus on the advances and innovations of 3D bioprinting skin for skin regeneration, within the last five years. After a brief introduction to skin anatomy, 3D bioprinting methods and the remarkable features of recent studies are classified as advances in materials, structures, and functions. We will discuss several ways to improve the clinical potential of 3D bioprinted skin, with state-of-the-art printing technology and novel biomaterials. After the breakthrough in the bottleneck of the current studies, highly developed skin can be fabricated, comprising stratified epidermis, dermis, and hypodermis with blood vessels, nerves, muscles, and skin appendages. We hope that this review will be priming water for future research and clinical applications, that will guide us to break new ground for the next generation of skin regeneration. Full article

Ideal drug carriers should be stable in biological environments but eventually release their drug load once inside the targeted cells. These two aspects can be in contrast with each other, thus they need to be carefully tuned in order to achieve the desired properties for specific applications. Quantifying drug release profiles in biological environments or inside cells can be highly challenging, and standard methods to determine drug release kinetics in many cases cannot be applied to complex biological environments or cells. Within this context, the present work combined kinetic studies by flow cytometry with aging experiments in biological fluids and size-exclusion chromatography to determine drug release profiles in biological environments and inside cells. To this purpose, anionic and zwitterionic liposomes were used as model nanomedicines. By changing lipid composition, liposome stability in serum and intracellular release kinetics could be tuned and formulations with very different properties could be obtained. The methods presented can be used to characterize liposome release profiles in complex biological media, as well as inside cells. In this way, liposome composition can be tuned in order to achieve formulations with optimal balance between stability and release kinetics for specific applications. Full article

Cadmium (Cd) is a toxic metal for the human organism and for all ecosystems. Cd is naturally found at low levels; however, higher amounts of Cd in the environment result from human activities as it spreads into the air and water in the form of micropollutants as a consequence of industrial processes, pollution, waste incineration, and electronic waste recycling. The human body has a limited ability to respond to Cd exposure since the metal does not undergo metabolic degradation into less toxic species and is only poorly excreted. The extremely long biological half-life of Cd essentially makes it a cumulative toxin; chronic exposure causes harmful effects from the metal stored in the organs. The present paper considers exposure and potential health concerns due to environmental cadmium. Exposure to Cd compounds is primarily associated with an elevated risk of lung, kidney, prostate, and pancreatic cancer. Cd has also been linked to cancers of the breast, urinary system, and bladder. The multiple mechanisms of Cd-induced carcinogenesis include oxidative stress with the inhibition of antioxidant enzymes, the promotion of lipid peroxidation, and interference with DNA repair systems. Cd²⁺ can also replace essential metal ions, including redox-active ones. A total of 12 cancer types associated with specific genes coding for the Cd-metalloproteome were identified in this work. In addition, we summarize the proper treatments of Cd poisoning, based on the use of selected Cd detoxifying agents and chelators, and the potential for preventive approaches to counteract its chronic exposure. Full article

A decrease in the circulating levels of adiponectin in obesity increases the risk of metabolic complications, but the role of globular adiponectin, a truncated form produced by proteolytic cleavage, has not been defined. The objective of this investigation was to determine how globular adiponectin is generated and to determine whether this process impacts obesity. The cleavage of recombinant full-length adiponectin into globular adiponectin by plasma in vitro was used to identify Gly-93 as the N-terminal residue after proteolytic processing. The amino acid sequence of the cleavage site suggested thrombin was the protease responsible for cleavage, and inhibitors confirmed its likely involvement. The proteolytic site was modified, and this thrombin-resistant mutant protein was infused for 4 weeks into obese adiponectin-knockout mice that had been on a high-fat diet for 8 weeks. The mutation of the cleavage site ensured that globular adiponectin was not generated, and thus did not confound the actions of the full-length adiponectin. Mice infused with the mutant adiponectin accumulated less fat and had smaller adipocytes compared to mice treated with globular adiponectin, and concurrently had elevated fasting glucose. The data demonstrate that generation of globular adiponectin through the action of thrombin increases both adipose tissue mass and adipocyte size, but it has no effect on fasting glucose levels in the context of obesity. Full article

Brain organoids are invaluable tools for pathophysiological studies or drug screening, but there are still challenges to overcome in making them more reproducible and relevant. Recent advances in three-dimensional (3D) bioprinting of human neural organoids is an emerging approach that may overcome the limitations of self-organized organoids. It requires the development of optimal hydrogels, and a wealth of research has improved our knowledge about biomaterials both in terms of their intrinsic properties and their relevance on 3D culture of brain cells and tissue. Although biomaterials are rarely biologically neutral, few articles have reviewed their roles on neural cells. We here review the current knowledge on unmodified biomaterials amenable to support 3D bioprinting of neural organoids with a particular interest in their impact on cell homeostasis. Alginate is a particularly suitable bioink base for cell encapsulation. Gelatine is a valuable helper agent for 3D bioprinting due to its viscosity. Collagen, fibrin, hyaluronic acid and laminin provide biological support to adhesion, motility, differentiation or synaptogenesis and optimize the 3D culture of neural cells. Optimization of specialized hydrogels to direct differentiation of stem cells together with an increased resolution in phenotype analysis will further extend the spectrum of possible bioprinted brain disease models. Full article

Epidemiological studies and clinical observations show evidence of sexual dimorphism in brain responses to several neurological conditions. It is suggested that sex-related differences between men and women may have profound effects on disease susceptibility, pathophysiology, and progression. Sexual differences of the brain are achieved through the complex interplay of several factors contributing to this phenomenon, such as sex hormones, as well as genetic and epigenetic differences. Despite recent advances, the precise link between these factors and brain disorders is incompletely understood. This review aims to briefly outline the most relevant aspects that differ between men and women in ischemia and neurodegenerative disorders (AD, PD, HD, ALS, and SM). Recognition of disparities between both sexes could aid the development of individual approaches to ameliorate or slow the progression of intractable disorders. Full article

Melanogenesis is a major part of the environmental responses and tissue development of the integumentary system. The balance between reduction and oxidation (redox) governs pigmentary responses, for which coordination among epidermal resident cells is indispensable. Here, we review the current understanding of melanocyte biology with a particular focus on the “master regulator” of oxidative stress responses (i.e., the Kelch-like erythroid cell-derived protein with cap‘n’collar homology-associated protein 1-nuclear factor erythroid-2-related factor 2 system) and the autoimmune pigment disorder vitiligo. Our investigation revealed that the former is essential in pigmentogenesis, whereas the latter results from unbalanced redox homeostasis and/or defective intercellular communication in the interfollicular epidermis (IFE). Finally, we propose a model in which keratinocytes provide a “niche” for differentiated melanocytes and may “imprint” IFE pigmentation. Full article

Mosquito transgenesis and gene-drive technologies provide the basis for developing promising new tools for vector-borne disease prevention by either suppressing wild mosquito populations or reducing their capacity from transmitting pathogens. Many studies of the regulatory DNA and promoters of genes with robust sex-, tissue- and stage-specific expression profiles have supported the development of new tools and strategies that could bring mosquito-borne diseases under control. Although the list of regulatory elements available is significant, only a limited set of those can reliably drive spatial–temporal expression. Here, we review the advances in our ability to express beneficial and other genes in mosquitoes, and highlight the information needed for the development of new mosquito-control and anti-disease strategies. Full article

Background: Multi-omics delivers more biological insight than targeted investigations. We applied multi-omics to patients with heart failure with reduced ejection fraction (HFrEF). Methods: 46 patients with HFrEF and 20 controls underwent metabolomic profiling, including liquid/gas chromatography mass spectrometry (LC-MS/GC-MS) and solid-phase microextraction (SPME) volatilomics in plasma and urine. HFrEF was defined using left ventricular global longitudinal strain, ejection fraction and NTproBNP. A consumer breath acetone (BrACE) sensor validated results in n = 73. Results: 28 metabolites were identified by GCMS, 35 by LCMS and 4 volatiles by SPME in plasma and urine. Alanine, aspartate and glutamate, citric acid cycle, arginine biosynthesis, glyoxylate and dicarboxylate metabolism were altered in HFrEF. Plasma acetone correlated with NT-proBNP (r = 0.59, 95% CI 0.4 to 0.7), 2-oxovaleric and cis-aconitic acid, involved with ketone metabolism and mitochondrial energetics. BrACE > 1.5 ppm discriminated HF from other cardiac pathology (AUC 0.8, 95% CI 0.61 to 0.92, p < 0.0001). Conclusion: Breath acetone discriminated HFrEF from other cardiac pathology using a consumer sensor, but was not cardiac specific. Full article

In addition to the canonical Gs adenylyl cyclase pathway, the serotonin type 6 receptor (5-HT₆R) recruits additional signaling pathways that control cognitive function, brain development, and synaptic plasticity in an agonist-dependent and independent manner. Considering that aberrant constitutive and agonist-induced active states are involved in various pathological mechanisms, the development of biased ligands with different functional profiles at specific 5-HT₆R-elicited signaling pathways may provide a novel therapeutic perspective in the field of neurodegenerative and psychiatric diseases. Based on the structure of SB-258585, an inverse agonist at 5-HT₆R-operated Gs and Cdk5 signaling, we designed a series of 1-(arylsulfonyl-isoindol-2-yl)piperazine derivatives and synthesized them using a sustainable mechanochemical method. We identified the safe and metabolically stable biased ligand 3g, which behaves as a neutral antagonist at the 5-HT₆R-operated Gs signaling and displays inverse agonist activity at the Cdk5 pathway. Inversion of the sulfonamide bond combined with its incorporation into the isoindoline scaffold switched the functional profile of 3g at Gs signaling with no impact at the Cdk5 pathway. Compound 3g reduced the cytotoxicity of 6-OHDA and produced a glioprotective effect against rotenone-induced toxicity in C8-D1A astrocyte cell cultures. In view of these findings, compound 3g can be considered a promising biased ligand to investigate the role of the 5-HT₆R-elicited Gs and Cdk5 signaling pathways in neurodegenerative diseases. Full article

Polysaccharides isolated from Lentinula edodes are bioactive compounds with immunomodulatory properties. In our previous studies from L. edodes mycelium, we have isolated a selenium(Se)-enriched fraction (named Se-Le-30), a mixture of linear 1,4-α-glucan and linear 1,3-β- and 1,6-β-glucans. In this study, we analyzed the effects of Se-Le-30 on the activation and proliferation of human T lymphocytes stimulated by anti-CD3 and anti-CD3/CD28 antibodies (Abs) and on the production of cytokines by peripheral blood mononuclear cells (PBMCs). Se-Le-30 had effects on T cell proliferation induced by Abs against CD3 and CD28. It significantly inhibited the proliferation of CD3-stimulated CD4⁺ and CD8⁺ T cells and enhanced the proliferation of CD4⁺ T cells stimulated with anti-CD3/CD28 Ab. Moreover, Se-Le-30 downregulated the number of CD3-stimulated CD4⁺CD69⁺ cells, CD4⁺CD25⁺ cells, as well as CD8⁺CD25⁺ cells, and upregulated the expression of CD25 marker on CD4+ and CD8+ T cells activated with anti-CD3/CD28 Abs. Furthermore, Se-Le-30 enhanced the synthesis of IFN-γ by the unstimulated and anti-CD3/CD28-stimulated PBMCs, inhibited synthesis of IL-2 and IL-4 by CD3-stimulated cells, and augmented the synthesis of IL-6 and IL-10 by unstimulated, CD3-stimulated, and CD3/CD28-stimulated PBMCs. Together, we demonstrated that Se-Le-30 exerts immunomodulatory effects on human T lymphocytes. These observations are of importance for the prospective use of Se-Le-30 in research or as a therapeutic compound. Full article

Atopic dermatitis is a chronic inflammatory skin disease in which the overproduction of reactive oxygen species plays a pivotal role in the pathogenesis and persistence of inflammatory lesions. Phototherapy represents one of the most used therapeutic options, with benefits in the clinical picture. Studies have demonstrated the immunomodulatory effect of phototherapy and its role in reducing molecule hallmarks of oxidative stress. In this review, we report the data present in literature dealing with the main signaling molecular pathways involved in oxidative stress after phototherapy to target atopic dermatitis-affected cells. Since oxidative stress plays a pivotal role in the pathogenesis of atopic dermatitis and its flare-up, new research lines could be opened to study new drugs that act on this mechanism, perhaps in concert with phototherapy. Full article

Cancer metabolic reprogramming is essential for maintaining cancer cell survival and rapid replication. A common target of this metabolic reprogramming is one-carbon metabolism which is notable for its function in DNA synthesis, protein and DNA methylation, and antioxidant production. Polyamines are a key output of one-carbon metabolism with widespread effects on gene expression and signaling. As a result of these functions, one-carbon and polyamine metabolism have recently drawn a lot of interest for their part in cancer malignancy. Therapeutic inhibitors that target one-carbon and polyamine metabolism have thus been trialed as anticancer medications. The significance and future possibilities of one-carbon and polyamine metabolism as a target in cancer therapy are discussed in this review. Full article

Irisin is a hormone able to reproduce some of the positive effects of physical activity and diet. Recently, we demonstrated the presence of Irisin at the ovarian level as a potential physiological regulator of follicular function. Adipose tissue is crucial for reproductive function through its metabolic activity and the production of adipokines. At present, the exact nature of adipocyte precursors is still under debate, but an important role has been assigned to the population of adipose tissue mesenchymal stromal cells (ASCs) of perivascular origin. It should be noted that, when appropriately stimulated, ASCs can differentiate into preadipocytes and, subsequently, adipocytes. Therefore, this present study was undertaken to explore the potential effect of Irisin on ASCs, known for their high differentiative potential. Since Irisin expression in ASCs was confirmed by PCR, we tested its potential effects on the main functional activities of these cells, including proliferation (BrdU uptake); metabolic activity (ATP production); redox status, evaluated as the generation of free molecules such as superoxide anion and nitric oxide; and scavenger activities, assessed as both enzymatic (superoxide dismutase) and non-enzymatic antioxidant power. Moreover, we tested the effect of Irisin on ASCs adipogenic differentiation. BrdU uptake was significantly (p < 0.001) inhibited by Irisin, while ATP production was significantly (p < 0.05) increased. Both superoxide anion and nitric oxide generation were significantly increased (p < 0.001) by Irisin, while scavenger activity was significantly reduced (p < 0.05). Irisin was found to significantly (p < 0.05) inhibit ASCs adipogenic differentiation. Taken together, the present results suggest a potential local role of Irisin in the regulation of adipose tissue function. Full article

Mitochondria calcium is a double-edged sword. While low levels of calcium are essential to maintain optimal rates of ATP production, extreme levels of calcium overcoming the mitochondrial calcium retention capacity leads to loss of mitochondrial function. In moderate amounts, however, ATP synthesis rates are inhibited in a calcium-titratable manner. While the consequences of extreme calcium overload are well-known, the effects on mitochondrial function in the moderately loaded range remain enigmatic. These observations are associated with changes in the mitochondria ultrastructure and cristae network. The present mini review/perspective follows up on previous studies using well-established cryo–electron microscopy and poses an explanation for the observable depressed ATP synthesis rates in mitochondria during calcium-overloaded states. The results presented herein suggest that the inhibition of oxidative phosphorylation is not caused by a direct decoupling of energy metabolism via the opening of a calcium-sensitive, proteinaceous pore but rather a separate but related calcium-dependent phenomenon. Such inhibition during calcium-overloaded states points towards mitochondrial ultrastructural modifications, enzyme activity changes, or an interplay between both events. Full article

Chemotherapy-induced peripheral neuropathy (CIPN) is a common side effect of several chemotherapeutic agents, such as Paclitaxel. The main symptoms of CIPN are pain and numbness in the hands and feet. Paclitaxel is believed to accumulate in the dorsal root ganglia and free nerve endings. Novel therapeutic agents might help to mitigate or prevent Paclitaxel toxicity on dorsal root ganglion (DRG) neurons. Thus, we used primary DRG neurons as a model to investigate the potential neuroprotective effects of the endocannabinoid-like substance, palmitoylethanolamide (PEA). DRG neurons were isolated from cervical to sacral segments of spinal nerves of Wister rats (6–8 weeks old). After isolation and purification of neuronal cell populations, different concentrations of Paclitaxel (0.01–10 µM) or PEA (0.1–10 µM) or their combination were tested on cell viability by MTT assay at 24 h, 48, and 72 h post-treatment. Furthermore, morphometric analyses of neurite length and soma size for DRG neurons were performed. Adverse Paclitaxel effects on cell viability were apparent at 72 h post-treatment whereas Paclitaxel significantly reduced the neurite length in a concentration-dependent manner nearly at all investigated time points. However, Paclitaxel significantly increased the size of neuronal cell bodies at all time windows. These phenotypic effects were significantly reduced in neurons additionally treated with PEA, indicating the neuroprotective effect of PEA. PEA alone led to a significant increase in neuron viability regardless of PEA concentrations, apparent improvements in neurite outgrowth as well as a significant decrease in soma size of neurons at different investigated time points. Taken together, PEA showed promising protective effects against Paclitaxel-related toxicity on DRG neurons. Full article

Peripheral nerve injuries (PNI) are common and often result in lifelong disability. The peripheral nervous system has an inherent ability to regenerate following injury, yet complete functional recovery is rare. Despite advances in the diagnosis and repair of PNIs, many patients suffer from chronic pain, and sensory and motor dysfunction. One promising surgical adjunct is the application of intraoperative electrical stimulation (ES) to peripheral nerves. ES acts through second messenger cyclic AMP to augment the intrinsic molecular pathways of regeneration. Decades of animal studies have demonstrated that 20 Hz ES delivered post-surgically accelerates axonal outgrowth and end organ reinnervation. This work has been translated clinically in a series of randomized clinical trials, which suggest that ES can be used as an efficacious therapy to improve patient outcomes following PNIs. The aim of this review is to discuss the cellular physiology and the limitations of regeneration after peripheral nerve injuries. The proposed mechanisms of ES protocols and how they facilitate nerve regeneration depending on timing of administration are outlined. Finally, future directions of research that may provide new perspectives on the optimal delivery of ES following PNI are discussed. Full article

Severe gut motility disorders are characterized by the ineffective propulsion of intestinal contents. As a result, the patients develop disabling/distressful symptoms, such as nausea and vomiting along with altered bowel habits up to radiologically demonstrable intestinal sub-obstructive episodes. Chronic intestinal pseudo-obstruction (CIPO) is a typical clinical phenotype of severe gut dysmotility. This syndrome occurs due to changes altering the morpho-functional integrity of the intrinsic (enteric) innervation and extrinsic nerve supply (hence neuropathy), the interstitial cells of Cajal (ICC) (mesenchymopathy), and smooth muscle cells (myopathy). In the last years, several genes have been identified in different subsets of CIPO patients. The focus of this review is to cover the most recent update on enteric dysmotility related to CIPO, highlighting (a) forms with predominant underlying neuropathy, (b) forms with predominant myopathy, and (c) mitochondrial disorders with a clear gut dysfunction as part of their clinical phenotype. We will provide a thorough description of the genes that have been proven through recent evidence to cause neuro-(ICC)-myopathies leading to abnormal gut contractility patterns in CIPO. The discovery of susceptibility genes for this severe condition may pave the way for developing target therapies for enteric neuro-(ICC)-myopathies underlying CIPO and other forms of gut dysmotility. Full article

High-mobility group box 1 (HMGB1) is a damage-associated molecular pattern released by dying cells to stimulate the immune response. During cell death, HMGB1 is translocated from the nucleus to the cytoplasm and passively released. High levels of secreted HMGB1 are observed in the faeces of inflammatory bowel disease (IBD) patients, indicating its role in IBD pathophysiology and potential as a non-invasive IBD biomarker. HMGB1 is important in regulating neuronal damage in the central nervous system; its pathological activity is intertwined with oxidative stress and inflammation. In this study, HMGB1 expression in the enteric nervous system and its relevance to intestinal neuroinflammation is explored in organotypic cultures of the myenteric plexus exposed to oxidative stimuli and in Winnie mice with spontaneous chronic colitis. Oxidative stimuli induced cytoplasmic translocation of HMGB1 in myenteric neurons in organotypic preparations. HMGB1 translocation correlated with enteric neuronal loss and oxidative stress in the myenteric ganglia of Winnie mice. Inhibition of HMGB1 by glycyrrhizic acid ameliorated HMGB1 translocation and myenteric neuronal loss in Winnie mice. These data highlight modulation of HMGB1 signalling as a therapeutic strategy to reduce the consequences of enteric neuroinflammation in colitis, warranting the exploration of therapeutics acting on the HMGB1 pathway as an adjunct treatment with current anti-inflammatory agents. Full article

The geomagnetic field (GMF) is a natural component of Earth’s biosphere. GMF reduction to near-null values (NNMF) induces gene expression modulation that generates biomolecular, morphological, and developmental changes. Here, we evaluate the effect of NNMF on gene expression and reactive oxygen species (ROS) production in time-course experiments on Arabidopsis thaliana. Plants exposed to NNMF in a triaxial Helmholtz coils system were sampled from 10 min to 96 h to evaluate differentially expressed genes (DEGs) of oxidative stress responses by gene microarray. In 24–96 h developing stages, H₂O₂ and polyphenols were also analyzed from roots and shoots. A total of 194 DEGs involved in oxidative reactions were selected, many of which showed a fold change ≥±2 in at least one timing point. Heatmap clustering showed DEGs both between roots/shoots and among the different time points. NNMF induced a lower H₂O₂ than GMF, in agreement with the expression of ROS-related genes. Forty-four polyphenols were identified, the content of which progressively decreased during NNMF exposition time. The comparison between polyphenols content and DEGs showed overlapping patterns. These results indicate that GMF reduction induces metabolomic and transcriptomic modulation of ROS-scavenging enzymes and H₂O₂ production in A. thaliana, which is paralleled by the regulation of antioxidant polyphenols. Full article

Regulated cell death (RCD) is a signal-controlled process that not only eliminates infected, damaged, or aged cells but is also implicated in a variety of pathological conditions. The process of RCD is regulated by intracellular proteins that undergo varying levels of post-translational modifications, including mono- or polyubiquitination. Functionally, ubiquitination can affect protein abundance, localization, and activity. Like other post-translational modifications, ubiquitination is a dynamic and reversible process mediated by deubiquitinases, a large class of proteases that cleave ubiquitin from proteins and other substrates. The balance between ubiquitination and deubiquitination machinery determines cell fate under stressful conditions. Here, we review the latest advances in our understanding of the role of deubiquitinases in regulating the main types of RCD, including apoptosis, necroptosis, pyroptosis, and ferroptosis. This knowledge may contribute to identifying new protein degradation-related prognostic markers and therapeutic targets for human disease. Full article

Leucine-rich repeat kinase 2 (LRRK2) gene mutation is an autosomal dominant mutation associated with Parkinson’s disease (PD). Among LRRK2 gene mutations, the LRRK2 G2019S mutation is frequently involved in PD onset. Currently, diverse gene correction tools such as zinc finger nucleases (ZFNs), helper-dependent adenoviral vector (HDAdV), the bacterial artificial chromosome-based homologous recombination (BAC-based HR) system, and CRISPR/Cas9-homology-directed repair (HDR) or adenine base editor (ABE) are used in genome editing. Gene correction of the LRRK2 G2019S mutation has been applied whenever new gene therapy tools emerge, being mainly applied to induced pluripotent stem cells (LRRK2 G2019S-mutant iPSCs). Here, we comprehensively introduce the principles and methods of each programmable nuclease such as ZFN, CRISPR/Cas9-HDR or ABE applied to LRRK2 G2019S, as well as those of HDAdV or BAC-based HR systems used as nonprogrammable nuclease systems. Full article

Lysophosphatidic acid (LPA), a multifunctional endogenous phospholipid, plays a vital role in cellular homeostasis and the malignant behavior of cancer cells through G-protein-coupled receptors. However, the role of LPA in β-catenin-mediated gastric cancer is unknown. Here, we have noted the high expression of LPAR2 in human gastric cancer tissues, and that LPA treatment significantly increased the proliferation, migration, and invasion of human gastric cancer cells. Results from our biochemical experiments showed that an LPA exposure increased the expression of β-catenin and its nuclear localization, increased the phosphorylation of glycogen synthase kinase 3β (GSK-3β), decreased the expression of Axin2, and increased the expression of the target genes of the β-catenin signaling pathway. The LPA2 receptor (LPAR2) antagonist significantly reduced the LPA-induced nuclear localization of β-catenin, the primary signaling event. The knockdown of LPAR2 in the gastric cancer cell lines robustly reduced the LPA-induced β-catenin activity. An LPA exposure increased the ATP production by both oxidative phosphorylation and glycolysis, and this effect was abrogated with the addition of an LPAR2 antagonist and XAV393, which stabilizes the Axin and inhibits the β-catenin signaling pathway. Based on our findings, the possibility that LPA contributes to gastric cancer initiation and progression through the β-catenin signaling pathway as well as by the dysregulation of the energy metabolism via the LPAR2 receptor and Axin2, respectively, provides a novel insight into the mechanism of and possible therapeutic targets of gastric cancer. Full article

In vitro ACE-1 inhibitory peptides were characterised previously from a number of microalgal species including Spirulina platensis (peptide IAPG), Chlorella vulgaris (peptides FDL, AFL, VVPPA), Isochrysis galbana (peptide YMGLDLK), Chlorella sorokiniana (peptides IW and LW) and indeed Nannochloropsis oculata (peptides GMNNLTP and LEQ). The isolation of protein from Nannochloropsis oculata using a combination of ammonium salt precipitation and xylanase treatment of resulting biomass combined with molecular weight cut off filtration to produce a permeate and characterisation of bioactive peptides is described. The Angiotensin-1-converting enzyme (ACE-1) IC₅₀ value for the generated permeate fraction was 370 µg/mL. Ninety-five peptide sequences within the permeate fraction were determined using mass spectrometry and eight peptides were selected for chemical synthesis based on in silico analysis. Synthesized peptides were novel based on a search of the literature and relevant databases. In silico, simulated gastrointestinal digestion identified further peptides with bioactivities including ACE-1 inhibitory peptides and peptides with antithrombotic and calcium/calmodulin-dependent kinase II (CAMKII) inhibition. This work highlights the potential of Nannochloropsis oculata biomass as both a protein and bioactive peptide resource, which could be harnessed for use in the development of functional foods and feeds. Full article

RNase Y is a crucial component of genetic translation, acting as the key enzyme initiating mRNA decay in many Gram-positive bacteria. The N-terminal domain of Bacillus subtilis RNase Y (Nter-BsRNaseY) is thought to interact with various protein partners within a degradosome complex. Bioinformatics and biophysical analysis have previously shown that Nter-BsRNaseY, which is in equilibrium between a monomeric and a dimeric form, displays an elongated fold with a high content of α-helices. Using multidimensional heteronuclear NMR and AlphaFold models, here, we show that the Nter-BsRNaseY dimer is constituted of a long N-terminal parallel coiled-coil structure, linked by a turn to a C-terminal region composed of helices that display either a straight or bent conformation. The structural organization of the N-terminal domain is maintained within the AlphaFold model of the full-length RNase Y, with the turn allowing flexibility between the N- and C-terminal domains. The catalytic domain is globular, with two helices linking the KH and HD modules, followed by the C-terminal region. This latter region, with no function assigned up to now, is most likely involved in the dimerization of B. subtilis RNase Y together with the N-terminal coiled-coil structure. Full article

Benzodiazepines (BZDs) are a class of widely prescribed psychotropic drugs that target GABA_A receptors (GABA_ARs) to tune inhibitory synaptic signaling throughout the central nervous system. Despite knowing their molecular target for over 40 years, we still do not fully understand the mechanism of modulation at the level of the channel protein. Nonetheless, functional studies, together with recent cryo-EM structures of GABA_A(α1)₂(βX)₂(γ2)₁ receptors in complex with BZDs, provide a wealth of information to aid in addressing this gap in knowledge. Here, mechanistic interpretations of functional and structural evidence for the action of BZDs at GABA_A(α1)₂(βX)₂(γ2)₁ receptors are reviewed. The goal is not to describe each of the many studies that are relevant to this discussion nor to dissect in detail all the effects of individual mutations or perturbations but rather to highlight general mechanistic principles in the context of recent structural information. Full article

The sarco/endoplasmic reticulum Ca-ATPase (SERCA) is a P-type ion pump that transports Ca²⁺ from the cytosol into the endoplasmic/sarcoplasmic reticulum (ER/SR) in most mammalian cells. It is critically important in muscle, facilitating relaxation and enabling subsequent contraction. Increasing SERCA expression or specific activity can alleviate muscle dysfunction, most notably in the heart, and we seek to develop small-molecule drug candidates that activate SERCA. Therefore, we adapted an NADH-coupled assay, measuring Ca-dependent ATPase activity of SERCA, to high-throughput screening (HTS) format, and screened a 46,000-compound library of diverse chemical scaffolds. This HTS platform yielded numerous hits that reproducibly alter SERCA Ca-ATPase activity, with few false positives. The top 19 activating hits were further tested for effects on both Ca-ATPase and Ca²⁺ transport, in both cardiac and skeletal SR. Nearly all hits increased Ca²⁺ uptake in both cardiac and skeletal SR, with some showing isoform specificity. Furthermore, dual analysis of both activities identified compounds with a range of effects on Ca²⁺-uptake and ATPase, which fit into distinct classifications. Further study will be needed to identify which classifications are best suited for therapeutic use. These results reinforce the need for robust secondary assays and criteria for selection of lead compounds, before undergoing HTS on a larger scale. Full article

Mitochondria have beneficial effects on cells by producing ATP and contributing to various biosynthetic procedures. On the other hand, dysfunctional mitochondria have detrimental effects on cells by inducing cellular damage, inflammation, and causing apoptosis in response to various stimuli. Therefore, a series of mitochondrial quality control pathways are required for the physiological state of cells to be maintained. Recent research has provided solid evidence to support that mitochondria are ejected from cells for transcellular degradation or transferred to other cells as metabolic support or regulatory messengers. In this review, we summarize the current understanding of the regulation of mitochondrial transmigration across the plasma membranes and discuss the functional significance of this unexpected phenomenon, with an additional focus on the impact on the pathogenesis of cardiovascular diseases. We also provide some perspective concerning the unrevealed mechanisms underlying mitochondrial ejection as well as existing problems and challenges concerning the therapeutic application of mitochondrial ejection. Full article

Over the past few decades, the number of available structural bioinformatics pipelines, libraries, plugins, web resources and software has increased exponentially and become accessible to the broad realm of life scientists. This expansion has shaped the field as a tangled network of methods, algorithms and user interfaces. In recent years PyMOL, widely used software for biomolecules visualization and analysis, has started to play a key role in providing an open platform for the successful implementation of expert knowledge into an easy-to-use molecular graphics tool. This review outlines the plugins and features that make PyMOL an eligible environment for supporting structural bioinformatics analyses. Full article

Prion diseases are a class of neurodegenerative diseases that are uniquely infectious. Whilst their general replication mechanism is well understood, the components required for the formation and propagation of highly infectious prions are poorly characterized. The protein-only hypothesis posits that the prion protein (PrP) is the only component of the prion; however, additional co-factors are required for its assembly into infectious prions. These can be provided by brain homogenate, but synthetic lipids and non-coding RNA have also been used in vitro. Here, we review a range of experimental approaches, which generate PrP amyloid assemblies de novo. These synthetic PrP assemblies share some, but not necessarily all, properties of genuine infectious prions. We will discuss the different experimental approaches, how a prion is defined, the non-protein requirements of a prion, and provide an overview of the current state of prion amplification and generation in vitro. Full article

As one of the most transplanted tissues of the human body, bone has varying architectures, depending on its anatomical location. Therefore, bone defects ideally require bone substitutes with a similar structure and adequate strength comparable to native bones. Light-based three-dimensional (3D) printing methods allow the fabrication of biomimetic scaffolds with high resolution and mechanical properties that exceed the result of commonly used extrusion-based printing. Digital light processing (DLP) is known for its faster and more accurate printing than other 3D printing approaches. However, the development of biocompatible resins for light-based 3D printing is not as rapid as that of bio-inks for extrusion-based printing. In this study, we developed CSMA-2, a photopolymer based on Isosorbide, a renewable sugar derivative monomer. The CSMA-2 showed suitable rheological properties for DLP printing. Gyroid scaffolds with high resolution were successfully printed. The 3D-printed scaffolds also had a compressive modulus within the range of a human cancellous bone modulus. Human adipose-derived stem cells remained viable for up to 21 days of incubation on the scaffolds. A calcium deposition from the cells was also found on the scaffolds. The stem cells expressed osteogenic markers such as RUNX2, OCN, and OPN. These results indicated that the scaffolds supported the osteogenic differentiation of the progenitor cells. In summary, CSMA-2 is a promising material for 3D printing techniques with high resolution that allow the fabrication of complex biomimetic scaffolds for bone regeneration. Full article

Glaucoma is a progressive age-related disease of the visual system and the leading cause of irreversible blindness worldwide. Currently, intraocular pressure (IOP) is the only modifiable risk factor for the disease, but even as IOP is lowered, the pathology of the disease often progresses. Hence, effective clinical targets for the treatment of glaucoma remain elusive. Glaucoma shares comorbidities with a multitude of vascular diseases, and evidence in humans and animal models demonstrates an association between vascular dysfunction of the retina and glaucoma pathology. Integral to the survival of retinal ganglion cells (RGCs) is functional neurovascular coupling (NVC), providing RGCs with metabolic support in response to neuronal activity. NVC is mediated by cells of the neurovascular unit (NVU), which include vascular cells, glial cells, and neurons. Nitric oxide-cyclic guanosine monophosphate (NO-cGMP) signaling is a prime mediator of NVC between endothelial cells and neurons, but emerging evidence suggests that cGMP signaling is also important in the physiology of other cells of the NVU. NO-cGMP signaling has been implicated in glaucomatous neurodegeneration in humans and mice. In this review, we explore the role of cGMP signaling in the different cell types of the NVU and investigate the potential links between cGMP signaling, breakdown of neurovascular function, and glaucoma pathology. Full article

Secondary lymphedema is a chronic, debilitating disease and one of the most common side effects of oncologic surgery, substantially decreasing quality of life. Despite the progress conducted in lymphedema research, the underlying pathomechanisms remain elusive. Lymphedema is considered to be a disease affecting an isolated extremity, yet imaging studies suggest systemic changes of the lymphatic system in the affected patients. To evaluate potential systemic manifestations in lymphedema, we collected matched fat and skin tissue from the edematous and non-edematous side of the same 10 lymphedema patients as well as anatomically matched probes from control patients to evaluate whether known lymphedema manifestations are present systemically and in comparison to health controls. The lymphedematous tissue displayed various known hallmarks of lymphedema compared to the healthy controls, such as increased epidermis thickness, collagen deposition in the periadipocyte space and the distinct infiltration of CD4+ cells. Furthermore, morphological changes in the lymphatic vasculature between the affected and unaffected limb in the same lymphedema patient were visible. Surprisingly, an increased collagen deposition as well as CD4 expression were also detectable in the non-lymphedematous tissue of lymphedema patients, suggesting that lymphedema may trigger systemic changes beyond the affected extremity. Full article

The human organic cation transporter 1 (OCT1) is expressed in the liver and mediates hepatocellular uptake of organic cations. However, some studies have indicated that OCT1 could transport neutral or even anionic substrates. This capability is interesting concerning protein-substrate interactions and the clinical relevance of OCT1. To better understand the transport of neutral, anionic, or zwitterionic substrates, we used HEK293 cells overexpressing wild-type OCT1 and a variant in which we changed the putative substrate binding site (aspartate474) to a neutral amino acid. The uncharged drugs trimethoprim, lamivudine, and emtricitabine were good substrates of hOCT1. However, the uncharged drugs zalcitabine and lamotrigine, and the anionic levofloxacin, and prostaglandins E2 and F2α, were transported with lower activity. Finally, we could detect only extremely weak transport rates of acyclovir, ganciclovir, and stachydrine. Deleting aspartate474 had a similar transport-lowering effect on anionic substrates as on cationic substrates, indicating that aspartate474 might be relevant for intra-protein, rather than substrate-protein, interactions. Cellular uptake of the atypical substrates by the naturally occurring frequent variants OCT1*2 (methionine420del) and OCT1*3 (arginine61cysteine) was similarly reduced, as it is known for typical organic cations. Thus, to comprehensively understand the substrate spectrum and transport mechanisms of OCT1, one should also look at organic anions. Full article

The opportunistic pathogen Pseudomonas aeruginosa uses quorum sensing to control its virulence. One of its major signal molecules, the Pseudomonas quinolone signal PQS, has high affinity to membranes and is known to be trafficked mainly via outer membrane vesicles (OMVs). We previously reported that several 3-hydroxy-4(1H)-quinolone 2,4-dioxygenases (HQDs) catalyze the cleavage of PQS and thus act as quorum quenching enzymes. Further analysis showed that, in contrast to other HQDs, the activity of HQD from Streptomyces bingchenggensis (HQD_S._b.) was unexpectedly stabilized by culture supernatants of P. aeruginosa. Interestingly, the stabilizing effect was higher with supernatants from the strain PA14 than with supernatants from the strain PAO1. Heat treatment and lyophilization hardly affected the stabilizing effect; however, fractionation of the supernatant excluded small molecules as stabilizing agents. In a pull-down assay, HQD_S._b. appeared to interact with several P. aeruginosa proteins previously found in the OMV proteome. This prompted us to probe the physical interaction of HQD_S._b. with prepared extracellular membrane vesicles. Homo-FRET of fluorescently labeled HQD_S._b. indeed indicated a spatial clustering of the protein on the vesicles. Binding of a PQS-cleaving enzyme to the OMVs of P. aeruginosa may enhance PQS degradation and is highly reconcilable with its function as a quorum quenching enzyme. Full article

Macrophages regulate cutaneous wound healing by immune surveillance, tissue repair and remodelling. The depletion of dermal macrophages during the early and middle stages of wound healing has a detrimental impact on wound closure, characterised by reduced vessel density, fibroblast and myofibroblast proliferation, delayed re-epithelization and abated post-healing fibrosis and scar formation. However, in some animal species, oral mucosa and foetal life, cutaneous wounds can heal normally and remain scarless without any involvement of macrophages. These paradoxical observations have created much controversy on macrophages’ indispensable role in skin wound healing. Advanced knowledge gained by characterising macrophage subsets, their plasticity in switching phenotypes and molecular drivers provides new insights into their functional importance during cutaneous wound healing. In this review, we highlight the recent findings on skin macrophage subsets, their functional role in adult cutaneous wound healing and the potential benefits of targeting them for therapeutic use. Full article

Allostery is a property of biological macromolecules featuring cooperative ligand binding and regulation of ligand affinity by effectors. The definition was introduced by Monod and Jacob in 1963, and formally developed as the “concerted model” by Monod, Wyman, and Changeux in 1965. Since its inception, this model of cooperativity was seen as distinct from and not reducible to the “sequential model” originally formulated by Pauling in 1935, which was developed further by Koshland, Nemethy, and Filmer in 1966. However, it is difficult to decide which model is more appropriate from equilibrium or kinetics measurements alone. In this paper, we examine several cooperative proteins whose functional behavior, whether sequential or concerted, is established, and offer a combined approach based on functional and structural analysis. We find that isologous, mostly helical interfaces are common in cooperative proteins regardless of their mechanism. On the other hand, the relative contribution of tertiary and quaternary structural changes, as well as the asymmetry in the liganded state, may help distinguish between the two mechanisms. Full article

Corticosteroids, oral or transtympanic, remain the mainstay for inner ear diseases characterized by hearing fluctuation or sudden changes in hearing, including sudden sensorineural hearing loss (SSNHL), Meniere’s disease (MD), and autoimmune inner ear disease (AIED). Despite their use across these diseases, the rate of complete recovery remains low, and results across the literature demonstrates significant heterogeneity with respect to the effect of corticosteroids, suggesting a need to identify more efficacious treatment options. Previously, our group has cross-referenced steroid-responsive genes in the cochlea with published single-cell and single-nucleus transcriptome datasets to demonstrate that steroid-responsive differentially regulated genes are expressed in spiral ganglion neurons (SGN) and stria vascularis (SV) cell types. These differentially regulated genes represent potential druggable gene targets. We utilized multiple gene target databases (DrugBank, Pharos, and LINCS) to identify orally administered, FDA approved medications that potentially target these genes. We identified 42 candidate drugs that have been shown to interact with these genes, with an emphasis on safety profile, and tolerability. This study utilizes multiple databases to identify drugs that can target a number of druggable genes in otologic disorders that are commonly treated with steroids, providing a basis for establishing novel repurposing treatment trials. Full article

Transcriptional regulation is pivotal for all living organisms and is required for adequate response to environmental fluctuations and intercellular signaling molecules. For precise regulation of transcription, cells have evolved regulatory systems on the genome architecture, including the chromosome higher-order structure (e.g., chromatin loops), location of transcription factor (TF)-binding sequences, non-coding RNA (ncRNA) transcription, chromatin configuration (e.g., nucleosome positioning and histone modifications), and the topological state of the DNA double helix. To understand how these genome-chromatin architectures and their regulators establish tight and specific responses at the transcription stage, the fission yeast fbp1 gene has been analyzed as a model system for decades. The fission yeast fbp1 gene is tightly repressed in the presence of glucose, and this gene is induced by over three orders of magnitude upon glucose starvation with a cascade of multi-layered regulations on various levels of genome and chromatin architecture. In this review article, we summarize the multi-layered transcriptional regulatory systems revealed by the analysis of the fission yeast fbp1 gene as a model system. Full article

Postbiotics are a new category of biotics that have the potential to confer health benefits but, unlike probiotics, do not require living cells to induce health effects and thus are not subject to the food safety requirements that apply to live microorganisms. Postbiotics are defined as a “preparation of inanimate microorganisms and/or their components that confers a health benefit on the host”. Postbiotic components include short-chain fatty acids, exopolysaccharides, vitamins, teichoic acids, bacteriocins, enzymes and peptides in a non-purified inactivated cell preparation. While research into postbiotics is in its infancy, there is increasing evidence that postbiotics have the potential to modulate human health. Specifically, a number of postbiotics have been shown to improve gut health by strengthening the gut barrier, reducing inflammation and promoting antimicrobial activity against gut pathogens. Additionally, research is being conducted into the potential application of postbiotics to other areas of the body, including the skin, vagina and oral cavity. The purpose of this review is to set out the current research on postbiotics, demonstrate how postbiotics are currently used in commercial products and identify a number of knowledge gaps where further research is needed to identify the potential for future applications of postbiotics. Full article

Residual dipolar couplings (RDCs) are increasingly used for high-throughput NMR-based structural studies and to provide long-range angular constraints to validate and refine structures of various molecules determined by X-ray crystallography and NMR spectroscopy. RDCs of a given molecule can be measured in an anisotropic environment that aligns in an external magnetic field. Here, we demonstrate the first application of polymer-based nanodiscs for the measurement of RDCs from nucleic acids. Polymer-based nanodiscs prepared using negatively charged SMA-EA polymer and zwitterionic DMPC lipids were characterized by size-exclusion chromatography, ¹H NMR, dynamic light-scattering, and ²H NMR. The magnetically aligned polymer-nanodiscs were used as an alignment medium to measure RDCs from a ¹³C/¹⁵N-labeled fluoride riboswitch aptamer using 2D ARTSY-HSQC NMR experiments. The results showed that the alignment of nanodiscs is stable for nucleic acids and nanodisc-induced RDCs fit well with the previously determined solution structure of the riboswitch. These results demonstrate that SMA-EA-based lipid-nanodiscs can be used as a stable alignment medium for high-resolution structural and dynamical studies of nucleic acids, and they can also be applicable to study various other biomolecules and small molecules in general. Full article

Genetic alterations in the LRRK2 gene, encoding leucine-rich repeat kinase 2, are a common risk factor for Parkinson’s disease. How LRRK2 alterations lead to cell pathology is an area of ongoing investigation, however, multiple lines of evidence suggest a role for LRRK2 in lipid pathways. It is increasingly recognized that in addition to being energy reservoirs and structural entities, some lipids, including neural lipids, participate in signaling cascades. Early investigations revealed that LRRK2 localized to membranous and vesicular structures, suggesting an interaction of LRRK2 and lipids or lipid-associated proteins. LRRK2 substrates from the Rab GTPase family play a critical role in vesicle trafficking, lipid metabolism and lipid storage, all processes which rely on lipid dynamics. In addition, LRRK2 is associated with the phosphorylation and activity of enzymes that catabolize plasma membrane and lysosomal lipids. Furthermore, LRRK2 knockout studies have revealed that blood, brain and urine exhibit lipid level changes, including alterations to sterols, sphingolipids and phospholipids, respectively. In human LRRK2 mutation carriers, changes to sterols, sphingolipids, phospholipids, fatty acyls and glycerolipids are reported in multiple tissues. This review summarizes the evidence regarding associations between LRRK2 and lipids, and the functional consequences of LRRK2-associated lipid changes are discussed. Full article

Small-angle neutron scattering (SANS) is a powerful tool for studying biological membranes and model lipid bilayer membranes. The length scales probed by SANS, being from 1 nm to over 100 nm, are well-matched to the relevant length scales of the bilayer, particularly when it is in the form of a vesicle. However, it is the ability of SANS to differentiate between isotopes of hydrogen as well as the availability of deuterium labeled lipids that truly enable SANS to reveal details of membranes that are not accessible with the use of other techniques, such as small-angle X-ray scattering. In this work, an overview of the use of SANS for studying unilamellar lipid bilayer vesicles is presented. The technique is briefly presented, and the power of selective deuteration and contrast variation methods is discussed. Approaches to modeling SANS data from unilamellar lipid bilayer vesicles are presented. Finally, recent examples are discussed. While the emphasis is on studies of unilamellar vesicles, examples of the use of SANS to study intact cells are also presented. Full article

Calcium sparks are the elementary Ca²⁺ release events in excitation-contraction coupling that underlie the Ca²⁺ transient. The frequency-dependent contractile force generated by cardiac myocytes depends upon the characteristics of the Ca²⁺ transients. A stochastic computational local control model of a guinea pig ventricular cardiomyocyte was developed, to gain insight into mechanisms of force-frequency relationship (FFR). This required the creation of a new three-state RyR2 model that reproduced the adaptive behavior of RyR2, in which the RyR2 channels transition into a different state when exposed to prolonged elevated subspace [Ca²⁺]. The model simulations agree with previous experimental and modeling studies on interval-force relations. Unlike previous common pool models, this local control model displayed stable action potential trains at 7 Hz. The duration and the amplitude of the [Ca²⁺]_myo transients increase in pacing rates consistent with the experiments. The [Ca²⁺]_myo transient reaches its peak value at 4 Hz and decreases afterward, consistent with experimental force-frequency curves. The model predicts, in agreement with previous modeling studies of Jafri and co-workers, diastolic sarcoplasmic reticulum, [Ca²⁺]_sr, and RyR2 adaptation increase with the increased stimulation frequency, producing rising, rather than falling, amplitude of the myoplasmic [Ca²⁺] transients. However, the local control model also suggests that the reduction of the L-type Ca²⁺ current, with an increase in pacing frequency due to Ca²⁺-dependent inactivation, also plays a role in the negative slope of the FFR. In the simulations, the peak Ca²⁺ transient in the FFR correlated with the highest numbers of SR Ca²⁺ sparks: the larger average amplitudes of those sparks, and the longer duration of the Ca²⁺ sparks. Full article