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Ana Cepero,1– 3,* Mónica Jiménez-Carretero,4,* Ylenia Jabalera,4 Lidia Gago,1– 3 Cristina Luque,1– 3 Laura Cabeza,1– 3 Consolación Melguizo,1– 3 Concepcion Jimenez-Lopez,4 José Prados1– 3 1Institute of Biopathology and Regenerative Medicine (IBIMER), Center of Biomedical Research (CIBM), University of Granada, Granada, 18100, Spain; 2Department of Anatomy and Embryology, Faculty of Medicine, University of Granada, Granada, 18071, Spain; 3Biosanitary Research Institute ibs.GRANADA, Granada, 18012, Spain; 4Department of Microbiology, Sciences School, University of Granada, Granada, 18002, Spain*These authors contributed equally to this workCorrespondence: Laura Cabeza; Concepción Jimenez-Lopez, Email lautea@ugr.es; cjl@ugr.esPurpose: The lack of specificity of conventional chemotherapy is one of the main difficulties to be solved in cancer therapy. Biomimetic magnetoliposomes are successful chemotherapy controlled-release systems, hyperthermia, and active targeting agents by functionalization of their surface with monoclonal antibodies. The membrane receptor Leucine-rich repeat-containing G-protein coupled receptor 5 (LGR5) stands out as colorectal cancer (CRC) biomarker and appears to be related to treatment resistance and the development of metastasis. The aim of this study was to assess the effectiveness and safety of LGR5-targeted biomimetic magnetoliposomes loaded with oxaliplatin (OXA) or 5-fluorouracil (5-FU) in the selective treatment of CRC and their possible application in hyperthermia.Methods: Synthesis, characterization and determination of heating capacity of magnetoliposomes transporting OXA or 5-FU (with and without LGR5 functionalization) were conducted. In vitro antitumoral activity was assayed in multiple colorectal cell lines at different times of exposition. In addition to this, cell internalization was studied by Prussian Blue staining, flow cytometry and fluorescence microscopy. In vivo acute toxicity of magnetoliposomes was performed to evaluate iron-related toxicity.Results: OXA and 5-FU loaded magnetoliposomes functionalized with LGR5 antibody showed higher cellular uptake than non-targeted nanoformulation with a reduction of the percentage of proliferation in colon cancer cell lines up to 3.2-fold of the IC50 value compared to that of free drug. The differences between non-targeted and targeted nanoformulations were more evident after short exposure times (4 and 8 hours). Interestingly, assays in the MC38 transduced cells with reduced LGR5 expression (MC38-L(-)), showed lower cell internalization of LGR5-targeted magnetoliposomes compared to non-transduced MC38 cell line. In addition, magnetoliposomes showed an in vitro favorable heating response under magnetic excitation and great iron-related biocompatibility data in vivo.Conclusion: Drug-loaded magnetoliposomes functionalized with anti-LGR5 antibodies could be a promising CRC treatment strategy for LGR5+ targeted chemotherapy, magnetic hyperthermia, and both in combination. Keywords: colorectal neoplasms, magnetoliposome, LGR5, oxaliplatin, 5-fluorouracil, magnetic hyperthermia

Cross-linking mass spectrometry (XL-MS) has seen significant improvements which have enhanced its utility for studying protein-protein interactions (PPIs), primarily due to the emergence of novel crosslinkers and the development of streamlined analysis workflows. Nevertheless, poor membrane permeability and side reactions with water limit the extent of productive intracellular crosslinking events that can be achieved with current crosslinkers. To address these problems, we have synthesized a novel crosslinker with o-nitrobenzyl-based photoresponsive groups. These o-nitrobenzyl ester (o-NBE) groups enhance the stability and hydrophobic properties of the crosslinker and add the potential for temporal resolution, i. e. the ability to control the initiation of the crosslinking reaction. Upon exposure to UV light the resulting aldehyde product reacts with adjacent amino groups and subsequent reductive amination of the formed Schiff-bases yields stable secondary amine linkages. This controlled activation mechanism enables precise UV-triggered protein crosslinking. We demonstrate proof-of principle of our o-NBE cross-linker to reliably detect PPIs by XL-MS using a recombinant model protein. We also demonstrate its ability to enter intact Hela cells, thereby indicating its future potential as a useful tool to study PPIs within the cellular environment., (© 2024 The Author(s). ChemBioChem published by Wiley-VCH GmbH.)

Targeting phospholipid biosynthesis, specifically phosphatidylcholine (PC), which is enhanced in tumor cells, has been proven a suitable antitumor strategy. In fact, the overexpression of the choline kinase α1 (ChoKα1) isoform has been found in malignant cells and tumors, thus becoming an excellent antitumor target. ChoKα1 inhibitors are being synthesized at the present that show a large inhibitory activity. Two of them have been chosen in this study as representatives of different structural families: a biscationic biphenyl derivative of thieno[3,2-d]pyrimidinium substituted with a cyclic amine (here referred to as Fa22) and a biscationic biphenyl thioethano derivative of 7-chloro-quinolinium substituted with a pyrrolidinic moiety (here referred to as PL48). However, the potential use of these types of compounds in systemic treatments is hampered because of their low specificity. In fact, to enter the cell and reach their target, these inhibitors use choline transporters and inhibit choline uptake, being that one of the causes of their toxicity. One way to solve this problem could be allowing their entrance into the cells by alternative ways. With this goal, MamC-mediated magnetic nanoparticles (BMNPs), already proven effective drug nanocarriers, have been used to immobilize Fa22 and PL48. The idea is to let BMNPs enter the cell (they enter the cell by endocytosis) carrying these molecules, and, therefore, offering another way in for these compounds. In the present study, we demonstrate that the coupling of Fa22 and PL48 to BMNPs allows these molecules to enter the tumoral cell without completely inhibiting choline uptake, so, therefore, the use of Fa22 and PL48 in these nanoformulations reduces the toxicity compared to that of the soluble drugs. Moreover, the nanoassemblies Fa22-BMNPs and PL48-BMNPs allow the combination of chemotherapy and local hyperthermia therapies for a enhanced cytotoxic effect on the tumoral HepG2 cell line. The consistency of the results, independently of the drug structure, may indicate that this behavior could be extended to other ChoKα1 inhibitors, opening up a possibility for their potential use in clinics., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Author(s). Published by Elsevier Inc. All rights reserved.)

The pupillary light reflex (PLR) adapts the amount of light reaching the retina, protecting it and improving image formation. Two PLR mechanisms have been described in vertebrates. First, the pretectum receives retinal inputs and projects to the Edinger-Westphal nucleus (EWN), which targets the ciliary ganglion through the oculomotor nerve (nIII). Postganglionic fibers enter the eye-globe, traveling to the iris sphincter muscle. Additionally, some vertebrates exhibit an iris-intrinsic PLR mechanism mediated by sphincter muscle cells that express melanopsin inducing muscle contraction. Given the high degree of conservation of the lamprey visual system, we investigated the mechanisms underlying the PLR to shed light onto their evolutionary origins. Recently, a PLR mediated by melanopsin was demonstrated in lampreys, suggested to be brain mediated. Remarkably, we found that PLR is instead mediated by direct retino-iridal cholinergic projections. This retina-mediated PLR acts synergistically with an iris-intrinsic mechanism that, as in other vertebrates, is mediated by melanopsin and has contribution of gap junctions between muscle fibers. In contrast, we show that lampreys lack the brain-mediated PLR. Our results suggest that two eye-intrinsic PLR mechanisms were present in early vertebrate evolution, whereas the brain-mediated PLR has a more recent origin., (© 2024. The Author(s).)

Animals constantly redirect their gaze away or towards relevant targets and, besides these goal-oriented responses, stabilizing movements clamp the visual scene avoiding image blurring. The vestibulo-ocular (VOR) and the optokinetic reflexes are the main contributors to gaze stabilization, whereas the optic tectum integrates multisensory information and generates orienting/evasive gaze movements in all vertebrates. Lampreys show a unique stepwise development of the visual system whose understanding provides important insights into the evolution and development of vertebrate vision. Although the developmental emergence of the visual components, and the retinofugal pathways have been described, the functional development of the visual system and the development of the downstream pathways controlling gaze are still unknown. Here, we show that VOR followed by light-evoked eye movements are the first to appear already in larvae, despite their burrowed lifestyle. However, the circuits controlling goal-oriented responses emerge later, in larvae in non-parasitic lampreys but during late metamorphosis in parasitic lampreys. The appearance of stabilizing responses earlier than goal-oriented in the lamprey development shows a stepwise transition from simpler to more complex visual systems, offering a unique opportunity to isolate the functioning of their underlying circuits., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Barandela, Núñez-González, Suzuki, Jiménez-López, Pombal and Pérez-Fernández.)

Macroalgae are an extensive resource for the obtention of bioactive compounds, mainly phenolic compounds, phlorotannins, and pigments. Fucoxanthin (Fx) is the most abundant pigment present in brown algae and has shown several useful bioactivities that can be used to fortify products in the food and cosmetic industries. Nevertheless, to date, there is still insufficient literature reporting on the extraction yield of Fx from U. pinnatifida species from green technologies. In this regard, the present study aims to optimize the extraction conditions to obtain the highest Fx yield from U. pinnatifida through emerging techniques, namely microwave-assisted extraction (MAE) and ultrasound-assisted extraction (UAE). These methods will be compared with the conventional methodologies of heat-assisted extraction (HAE) and Soxhlet-assisted extraction (SAE). According to our results, even though the extraction yield could be slightly higher when using MAE than UAE, the Fx concentration obtained from the alga was double when using UAE. Thus, the Fx ratio in the final extract reached values of 124.39 mg Fx/g E. However, the optimal conditions should also be considered since UAE needed 30 min to perform the extraction, whereas MAE was able to obtain 58.83 mg Fx/g E in only 3 min and 2 bar, meaning less energy expenditure and minimum cost function. To our knowledge, this study obtains the highest concentrations of Fx ever reported (58.83 mg Fx/g E for MAE and 124.39 mg Fx/g E for UAE), with low energy consumption and short times (3.00 min for MAE and 35.16 min for UAE). Any of these results could be selected for further experiments and proposed for industrial scaling-up.

Lipids exert their cellular functions in individual organelles, in some cases on the scale of even smaller, specialized membrane domains. Thus, the experimental capacity to precisely manipulate lipid levels at the subcellular level is crucial for studying lipid-related processes in cell biology. Photo-caged lipid probes which partition into specific cellular membranes prior to photoactivation have emerged as key tools for localized and selective perturbation of lipid concentration in living cells. In this review, we provide an overview of the recent advances in the area and outline which developments are still required for the methodology to be more widely implemented in the wider membrane biology community., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

India plays an important role in the production of oilseeds, which are mainly cultivated for future extraction of their oil. In addition to the energic and nutritional contribution of these seeds, oilseeds are rich sources of bioactive compounds (e.g., phenolic compounds, proteins, minerals). A regular and moderate dietary supplementation of oilseeds promotes health, prevents the appearance of certain diseases (e.g., cardiovascular diseases (CVDs), cancers) and delays the aging process. Due to their relevant content in nutraceutical molecules, oilseeds and some of their associated processing wastes have raised interest in food and pharmaceutical industries searching for innovative products whose application provides health benefits to consumers. Furthermore, a circular economy approach could be considered regarding the re-use of oilseeds' processing waste. The present article highlights the different oilseed types, the oilseeds-derived bioactive compounds as well as the health benefits associated with their consumption. In addition, the different types of extractive techniques that can be used to obtain vegetable oils rich from oilseeds, such as microwave-assisted extraction (MAE), ultrasonic-assisted extraction (UAE) and supercritical fluid extraction (SFE), are reported. We conclude that the development and improvement of oilseed markets and their byproducts could offer even more health benefits in the future, when added to other foods.

Dopamine is likely the most studied modulatory neurotransmitter, in great part due to characteristic motor deficits in Parkinson's disease that arise after the degeneration of the dopaminergic neurons in the substantia nigra pars compacta (SNc). The SNc, together with the ventral tegmental area (VTA), play a key role modulating motor responses through the basal ganglia. In contrast to the large amount of existing literature addressing the mammalian dopaminergic system, comparatively little is known in other vertebrate groups. However, in the last several years, numerous studies have been carried out in basal vertebrates, allowing a better understanding of the evolution of the dopaminergic system, especially the SNc/VTA. We provide an overview of existing research in basal vertebrates, mainly focusing on lampreys, belonging to the oldest group of extant vertebrates. The lamprey dopaminergic system and its role in modulating motor responses have been characterized in significant detail, both anatomically and functionally, providing the basis for understanding the evolution of the SNc/VTA in vertebrates. When considered alongside results from other early vertebrates, data in lampreys show that the key role of the SNc/VTA dopaminergic neurons modulating motor responses through the basal ganglia was already well developed early in vertebrate evolution.

Much interest has gained the biomineralization process carried out by magnetotactic bacteria. These bacteria are ubiquitous in natural environments and share the ability to passively align along the magnetic field lines and actively swim along them. This ability is due to their magnetosome chain, each magnetosome consisting on a magnetic crystal enveloped by a lipid bilayer membrane to which very unique proteins are associated. Magnetotactic bacteria exquisitely control magnetosome formation, making the magnetosomes the ideal magnetic nanoparticle of potential use in many technological applications. The difficulty to scale up magnetosome production has triggered the research on the in vitro production of biomimetic (magnetosome-like) magnetite nanoparticles. In this context, magnetosome proteins are being used to mediate such in vitro magnetite precipitation experiments. The present work reviews the knowledgement on the magnetosome proteins thought to have a role on the in vivo formation of magnetite crystals in the magnetosome, and the recombinant magnetosome proteins used in vitro to form biomimetic magnetite. It also summarizes the data provided in the literature on the biomimetic magnetite nanoparticles obtained from those in vitro experiments., (Copyright © 2016 Elsevier Inc. All rights reserved.)

The interaction of Candida albicans with phagocytes of the host's innate immune system is highly dynamic, and its outcome directly impacts the progression of infection. While the switch to hyphal growth within the macrophage is the most obvious physiological response, much of the genetic response reflects nutrient starvation: translational repression and induction of alternative carbon metabolism. Changes in amino acid metabolism are not seen, with the striking exception of arginine biosynthesis, which is upregulated in its entirety during coculture with macrophages. Using single-cell reporters, we showed here that arginine biosynthetic genes are induced specifically in phagocytosed cells. This induction is lower in magnitude than during arginine starvation in vitro and is driven not by an arginine deficiency within the phagocyte but instead by exposure to reactive oxygen species (ROS). Curiously, these genes are induced in a narrow window of sublethal ROS concentrations. C. albicans cells phagocytosed by primary macrophages deficient in the gp91(phox) subunit of the phagocyte oxidase do not express the ARG pathway, indicating that the induction is dependent on the phagocyte oxidative burst. C. albicans arg pathway mutants are retarded in germ tube and hypha formation within macrophages but are not notably more sensitive to ROS. We also find that the ARG pathway is regulated not by the general amino acid control response but by transcriptional regulators similar to the Saccharomyces cerevisiae ArgR complex. In summary, phagocytosis induces this single amino acid biosynthetic pathway in an ROS-dependent manner.

Alternative splicing is commonly used by the Metazoa to generate more than one protein from a gene. However, such diversification of the proteome by alternative splicing is much rarer in fungi. We describe here an ancient fungal alternative splicing event in which these two proteins are generated from a single alternatively spliced ancestral SKI7/HBS1 gene retained in many species in both the Ascomycota and Basidiomycota. While the ability to express two proteins from a single SKI7/HBS1 gene is conserved in many fungi, the exact mechanism by which they achieve this varies. The alternative splicing was lost in Saccharomyces cerevisiae following the whole-genome duplication event as these two genes subfunctionalized into the present functionally distinct HBS1 and SKI7 genes. When expressed in yeast, the single gene from Lachancea kluyveri generates two functionally distinct proteins. Expression of one of these proteins complements hbs1, but not ski7 mutations, while the other protein complements ski7, but not hbs1. This is the first known case of subfunctionalization by loss of alternative splicing in yeast. By coincidence, the ancestral alternatively spliced gene was also duplicated in Schizosaccharomyces pombe with subsequent subfunctionalization and loss of splicing. Similar subfunctionalization by loss of alternative splicing in fungi also explains the presence of two PTC7 genes in the budding yeast Tetrapisispora blattae, suggesting that this is a common mechanism to preserve duplicate alternatively spliced genes., Competing Interests: The authors have declared that no competing interests exist.

Introduction: endoscopic mucosal resection is an accepted technique for the treatment of proximal gastrointestinal tract superficial lesions., Objectives: to evaluate the efficacy and safety of this procedure in the proximal gastrointestinal tract., Material and Methods: forty one consecutive patients (23 males and 18 females, mean age of 61 ± 11.5 years) were included in our study. Fifty nine resections were performed in these patients in 69 sessions. Lesions treated consisted of elevated lesions with high grade dysplasia in the context of Barrett's esophagus (group A), high grade dysplasia appearing in random biopsies taken during the follow-up of Barrett's esophagus (group B) and superficial gastroduodenal lesions (group C). Snare resection after submucosal injection, band ligator-assisted or cap-assisted mucosal resection were the chosen techniques., Results: we resected 7 elevated lesions with high grade dysplasia in the context of Barrett's esophagus, 6 complete Barrett's esophagus with high grade dysplasia in 16 sequential sessions and 46 gastroduodenal superficial lesions (10 adenomas, 9 gastric superficial carcinomas, 18 carcinoid tumours and 9 lesions of different histological nature). Resections in the two first groups were complete in 100% of the cases, and in 97.9% of the cases in group C. Complications included 2 cases of limited deferred bleeding (groups A and B) and another two cases of stenosis with little clinical relevance in Group B., Conclusions: a) endoscopic mucosal resection is an efficient technique for the treatment of proximal gastrointestinal tract superficial lesions; b) it is a safe procedure with a low percentage of complications, which can generally be managed endoscopically; and c) in contrast with other ablative techniques, endoscopic mucosal resection offers the possibility of a pathologic analysis of the samples.

Introduction: exposure to ionizing radiation is associated with an increased risk of developing tumors. Patients with Crohn's disease (CD) usually require multiple imaging tests using this type of radiation., Objectives: the objectives of this study were to estimate the total dose of ionizing radiation received by patients with Crohn's disease during their course and to identify the factors associated with higher radiation doses., Methods: two hundred thirty-five CD patients diagnosed between 1972 and 2010 were included. The effective dose (ED) received by each patient was calculated retrospectively based on the number of gastrointestinal transit studies and computed tomography scans performed. Considering recent epidemiological studies, an ED greater than or equal to 50 mSv was used as the cut-off point for increased risk of developing cancer., Results: the mean ED received per patient was 33.4 mSv (95% CI 29.3-37.5). A total of 49 (20.8%) patients received an ED ≥ 50 mSv. The following factors were identified as independent predictors associated with an ED ≥ 50 mSv: Age older than 40 years, need for surgery, age under 16 years at diagnosis and disease duration over 8 years., Conclusions: a substantial proportion of patients with Crohn's disease receive high doses of potentially carcinogenic ionizing radiation. Identification of the most susceptible patients to receive high doses of radiation, monitoring of effective doses received and the use of imaging techniques that do not require ionizing radiation (MR enterography, abdominal ultrasound) could contribute in reducing patients' exposure to potentially carcinogenic ionizing radiation.

Like many organisms the fungal pathogen Candida albicans senses changes in the environmental CO(2) concentration. This response involves two major proteins: adenylyl cyclase and carbonic anhydrase (CA). Here, we demonstrate that CA expression is tightly controlled by the availability of CO(2) and identify the bZIP transcription factor Rca1p as the first CO(2) regulator of CA expression in yeast. We show that Rca1p upregulates CA expression during contact with mammalian phagocytes and demonstrate that serine 124 is critical for Rca1p signaling, which occurs independently of adenylyl cyclase. ChIP-chip analysis and the identification of Rca1p orthologs in the model yeast Saccharomyces cerevisiae (Cst6p) point to the broad significance of this novel pathway in fungi. By using advanced microscopy we visualize for the first time the impact of CO(2) build-up on gene expression in entire fungal populations with an exceptional level of detail. Our results present the bZIP protein Rca1p as the first fungal regulator of carbonic anhydrase, and reveal the existence of an adenylyl cyclase independent CO(2) sensing pathway in yeast. Rca1p appears to regulate cellular metabolism in response to CO(2) availability in environments as diverse as the phagosome, yeast communities or liquid culture.

A case of gastric metastasis of renal carcinomas is described. The patient had been nephrectomized 22 months ago, and she had yet presented another in appearance solitary metastasis in lung and brain, which had been resected. This is a very unusual localization in the spread of a renal tumor, and two cases alone exist in the literature up to date which had been diagnosed during life. In despite of surgical treatment, a short free interval after nephrectomy and the tumor aggressivity, with three foci of spread at less than a year, have been the pivotal factors determining a relatively short survival of 26 months after resection of primary tumor.