Hasil untuk "Biology (General)"

Yunhan Zhang, Ying Zhang, Jiaoyan Song et al.

Abstract Piezo1 plays a key role in the immune response during sepsis. To date, our understanding of the role of Piezo1 in inflammatory diseases has mostly been limited to influencing vasomotor function and regulating inflammatory infiltration. Whether and how Piezo1 in macrophages is involved in developing septic cardiac dysfunction has never been explored. Here, we have successfully established a mouse model with myeloid cell-specific knockdown of Piezo1. The intraperitoneal injection of lipopolysaccharide (LPS) resulted in a significant increase in cardiac macrophage infiltration, as well as an increase in the expression of inflammatory factors and the inflammatory response. However, myeloid cell-specific knockdown of Piezo1 impaired this response, leading to an increase in macrophage polarization towards the M2 type and the decreased inflammatory response. As a result, myocardial injury caused by sepsis was attenuated. We have also demonstrated that the PI3K/AKT pathway is significantly activated after Piezo1 knockdown, resulting in reduced myocardial dysfunction. Our data indicate that myeloid cell-specific knockdown of Piezo1 can influence macrophage polarization and thus exert cardioprotective effects in a murine model of sepsis, providing potential ideas and targets for the treatment of infectious cardiac dysfunction.

Janine Wesslowski, Sadia Safi, Michelle Rottmann et al.

Upon engagement of one of the nineteen secreted Wnt signaling proteins with one of the ten Frizzled transmembrane Wnt receptors (FZD_1–10), a wide variety of cellular Wnt signaling responses can be elicited, the selectivity of which depends on the following: (1) the specific Wnt-FZD pairing, (2) the participation of Wnt co-receptors and (3) the cellular context. Co-receptors play a pivotal role in guiding the specificity of Wnt signaling, most notably between β-catenin-dependent and -independent pathways, where co-receptors such as LRP5/6 and ROR1/2/PTK7 play major roles, respectively. It remains less understood how specific Wnt/FZD combinations contribute to the selectivity of downstream Wnt signaling, and we lack accurate comparative data on their binding properties under physiological conditions. Here, using fluorescently tagged Wnt3a, Wnt5a and Wnt16 proteins and cell lines expressing HiBiT-tagged Frizzled, we build on our ongoing efforts to provide a complete overview of the biophysical properties of all Wnt/FZD interactions using full-length proteins. Our real-time NanoBRET analysis using living cells expressing low receptor levels provides more accurate quantification of binding and will help us understand how these binary engagements control Wnt signaling outputs. We also provide evidence that LRP6 regulates the binding affinity of Wnt/FZD interactions in the trimeric Wnt-FZD-LRP6 complex.

Xuying Hu, Qian Zhang, Meiling Ge et al.

Deep-sea corals are critical to global exploration of deep-sea biodiversity, but research on them in the Indian Ocean is very limited. In this study, we report the first discovery of two new species of red corals at a depth of 1697 m in the western Indian Ocean. The species were identified through detailed morphological analysis, including microscopic examination of colony structure, branches and autozooids (assessing size, abundance and spacing), as well as scanning electron microscopy (SEM) of sclerite morphology and quantity. Two species for the genus Hemicorallium Gray, 1867 were found as new species, designated Hemicorallium indicodensum sp. nov. and Hemicorallium jiaolongensis sp. nov. H. indicodensum sp. nov. is characterised by densely branched structures and numerous globular double-club sclerites, distinguishing it from other species. Meanwhile, H. jiaolongensis sp. nov. features yellowish-white colonies with short rods with sharp, large radial projections in the tentacles. Four mitochondrial regions were used to reveal the phylogenetic relationship in Coralliidae, supporting the taxonomic placement of these two new species. This study provides significant new insights into the biodiversity of deep-sea corals in the Indian Ocean, enriching the known species pool. Additionally, a more comprehensive key to the genus Hemicorallium is provided, further enhancing our understanding of the group’s taxonomy.

Anne Longakit, Chloe Hess, Christina Zhang et al.

Immunohistochemistry (IHC) and immunofluorescence (IF) are fundamental molecular biology techniques to assess protein expression. However, the melanin present normally in the eye in the uveal tract (choroid, iris, and ciliary body) and the retinal pigment epithelium (RPE) poses a significant challenge for IHC and IF. This is because melanin interferes with both chromogenic and fluorescent detection methods. Additionally, formalin fixation, which is commonly used for IHC, can result in shrinkage and loss of cellular detail in the eye. This protocol provides an optimized approach using Davidson’s fixative with a hydrogen peroxide bleaching step to eliminate melanin interference in the mouse eye, improving the quality and interpretability of IHC analyses of the uveal tract and RPE. It is particularly useful for the analysis of uveal melanoma.

Kayla H Szymanik, Emily A Rex, Vamshikrishna R Pothireddy et al.

Proper recognition of viral pathogens is an essential part of the innate immune response. A common viral replicative intermediate and chemical signal that cells use to identify pathogens is the presence of a triphosphorylated 5' end (5'ppp) RNA, which activates the cytosolic RNA sensor RIG-I and initiates downstream antiviral signaling. While 5'pppRNA generated by viral RNA-dependent RNA polymerases (RdRps) can be a potent activator of the immune response, endogenous RNA polymerase III (RNAPIII) transcripts can retain the 5'ppp generated during transcription and induce a RIG-I-mediated immune response. We have previously shown that host RNA triphosphatase dual-specificity phosphatase 11 (DUSP11) can act on both host and viral RNAs, altering their levels and reducing their ability to induce RIG-I activation. Our previous work explored how experimentally altered DUSP11 activity can impact immune activation, prompting further exploration into natural contexts of altered DUSP11 activity. Here, we have identified viral DUSP11 homologs (vDUSP11s) present in some avipoxviruses. Consistent with the known functions of host DUSP11, we have shown that expression of vDUSP11s: 1) reduces levels of endogenous RNAPIII transcripts, 2) reduces a cell's sensitivity to 5'pppRNA-mediated immune activation, and 3) restores virus infection defects seen in the absence of DUSP11. Our results identify a context where DUSP11 activity has been co-opted by viruses to alter RNA metabolism and influence the outcome of infection.

Koki Toyoda, Shunsuke Kimura, Keiichiro Maki et al.

Effective gutta-percha/sealer removal is essential for predictable nonsurgical endodontic retreatment, but the effects of key operational parameters on nickel–titanium rotary removal instruments remain insufficiently understood. This study aimed to examine how varying downward loads and rotational speeds affect the removal efficiency and torque/force generation of the HyFlex Remover. Sixty transparent straight resin canals filled with gutta-percha and AH Plus sealer were prepared using the HyFlex Remover at two rotational speeds (400 and 800 rpm) and three downward loads (2, 3, and 4 N; <i>n</i> = 10/group). The removal rate, calculated using micro-computed tomography, as well as removal time, maximum force, maximum torque, and cumulative torque were recorded and evaluated using two-way analysis of variance with Bonferroni correction (α = 0.05). The removal rate was consistently higher at 400 rpm than at 800 rpm (<i>p</i> < 0.001), while removal time and cumulative torque were greater at 400 rpm (<i>p</i> < 0.001). Maximum torque differed only between the 800-2 N and 800-4 N groups (<i>p</i> = 0.006). Maximum force increased with higher loads (<i>p</i> < 0.001), and at 3 N and 4 N, it was lower at 400 rpm than at 800 rpm (3 N: <i>p</i> = 0.039, 4 N: <i>p</i> < 0.001). Overall, lower downward loads reduced torque but prolonged working time, whereas higher rotational speeds shortened both working time and torque but decreased the removal rate.

Callum M Ives, Linh Nguyen, Carl A Fogarty et al.

Glycosylation of the SARS-CoV-2 spike (S) protein represents a key target for viral evolution because it affects both viral evasion and fitness. Successful variations in the glycan shield are difficult to achieve though, as protein glycosylation is also critical to folding and structural stability. Within this framework, the identification of glycosylation sites that are structurally dispensable can provide insight into the evolutionary mechanisms of the shield and inform immune surveillance. In this work, we show through over 45 μs of cumulative sampling from conventional and enhanced molecular dynamics (MD) simulations, how the structure of the immunodominant S receptor binding domain (RBD) is regulated by N-glycosylation at N343 and how this glycan’s structural role changes from WHu-1, alpha (B.1.1.7), and beta (B.1.351), to the delta (B.1.617.2), and omicron (BA.1 and BA.2.86) variants. More specifically, we find that the amphipathic nature of the N-glycan is instrumental to preserve the structural integrity of the RBD hydrophobic core and that loss of glycosylation at N343 triggers a specific and consistent conformational change. We show how this change allosterically regulates the conformation of the receptor binding motif (RBM) in the WHu-1, alpha, and beta RBDs, but not in the delta and omicron variants, due to mutations that reinforce the RBD architecture. In support of these findings, we show that the binding of the RBD to monosialylated ganglioside co-receptors is highly dependent on N343 glycosylation in the WHu-1, but not in the delta RBD, and that affinity changes significantly across VoCs. Ultimately, the molecular and functional insight we provide in this work reinforces our understanding of the role of glycosylation in protein structure and function and it also allows us to identify the structural constraints within which the glycosylation site at N343 can become a hotspot for mutations in the SARS-CoV-2 S glycan shield.

Laia Cuesta-Casanovas, Jennifer Delgado-Martínez, Josep M. Cornet-Masana et al.

Abstract Background Development of precision medicine requires the identification of easily detectable and druggable biomarkers. Despite recent targeted drug approvals, prognosis of acute myeloid leukemia (AML) patients needs to be greatly improved, as relapse and refractory disease are still difficult to manage. Thus, new therapeutic approaches are needed. Based on in silico-generated preliminary data and the literature, the role of the prolactin (PRL)-mediated signaling was interrogated in AML. Methods Protein expression and cell viability were determined by flow cytometry. Repopulation capacity was studied in murine xenotransplantation assays. Gene expression was measured by qPCR and luciferase-reporters. SA-β-Gal staining was used as a senescence marker. Results The prolactin receptor (PRLR) was upregulated in AML cells, as compared to their healthy counterpart. The genetic and molecular inhibition of this receptor reduced the colony-forming potential. Disruption of the PRLR signaling, either using a mutant PRL or a dominant-negative isoform of PRLR, reduced the leukemia burden in vivo, in xenotransplantation assays. The expression levels of PRLR directly correlated with resistance to cytarabine. Indeed, acquired cytarabine resistance was accompanied with the induction of PRLR surface expression. The signaling associated to PRLR in AML was mainly mediated by Stat5, in contrast to the residual function of Stat3. In concordance, Stat5 mRNA was significantly overexpressed at mRNA levels in relapse AML samples. A senescence-like phenotype, measured by SA-β-gal staining, was induced upon enforced expression of PRLR in AML cells, partially dependent on ATR. Similar to the previously described chemoresistance-induced senescence in AML, no cell cycle arrest was observed. Additionally, the therapeutic potential of PRLR in AML was genetically validated. Conclusions These results support the role of PRLR as a therapeutic target for AML and the further development of drug discovery programs searching for specific PRLR inhibitors.

Ruibang Wu, Leizhen Huang, Qinghong Xia et al.

Intervertebral disc degeneration (IDD) is a major contributing factor to both lower back and neck pain. As IDD progresses, the intervertebral disc (IVD) loses its ability to maintain its disc height when subjected to axial loading. This failure in the weight-bearing capacity of the IVD is a characteristic feature of degeneration. Natural polymer-based hydrogel, derived from biological polymers, possesses biocompatibility and is able to mimic the structure of extracellular matrix, enabling them to support cellular behavior. However, their mechanical performance is relatively poor, thus limiting their application in IVD regeneration. In this study, we developed an injectable composite hydrogel, namely, Mel-MBG/SA, which is similar to natural weight-bearing IVD. Mesoporous bioactive glasses not only enhance hydrogels, but also act as carriers for melatonin (Mel) to suppress inflammation during IDD. The Mel-MBG/SA hydrogel further provides a mixed system with sustained Mel release to alleviate IL-1β-induced oxidative stress and relieve inflammation associated with IDD pathology. Furthermore, our study shows that this delivery system can effectively suppress inflammation in the rat tail model, which is expected to further promote IVD regeneration. This approach presents a novel strategy for promoting tissue regeneration by effectively modulating the inflammatory environment while harnessing the mechanical properties of the material.

Keisuke Hitachi, Masahiko Honda, Kunihiro Tsuchida

Skeletal muscle is a pivotal organ in humans that maintains locomotion and homeostasis. Muscle atrophy caused by sarcopenia and cachexia, which results in reduced muscle mass and impaired skeletal muscle function, is a serious health condition that decreases life longevity in humans. Recent studies have revealed the molecular mechanisms by which long non-coding RNAs (lncRNAs) regulate skeletal muscle mass and function through transcriptional regulation, fiber-type switching, and skeletal muscle cell proliferation. In addition, lncRNAs function as natural inhibitors of microRNAs and induce muscle hypertrophy or atrophy. Intriguingly, muscle atrophy modifies the expression of thousands of lncRNAs. Therefore, although their exact functions have not yet been fully elucidated, various novel lncRNAs associated with muscle atrophy have been identified. Here, we comprehensively review recent knowledge on the regulatory roles of lncRNAs in skeletal muscle atrophy. In addition, we discuss the issues and possibilities of targeting lncRNAs as a treatment for skeletal muscle atrophy and muscle wasting disorders in humans.

Justin Rashtian, Diana E. Chavkin, Zaher Merhi

Abstract A mounting body of the literature suggests that environmental chemicals found in food and water could affect female reproduction. Many worldwide daily-used products have been shown to contain chemicals that could incur adverse reproductive outcomes in the perinatal/neonatal periods, childhood, adolescence, and even adulthood. The potential impact of Bisphenol A (BPA), Phthalates and Perfluoroalkyl substances (PFAS) on female reproduction, in particular on puberty, PCOS pathogenesis, infertility, ovarian function, endometriosis, and recurrent pregnancy loss, in both humans and animals, will be discussed in this report in order to provide greater clinician and public awareness about the potential consequences of these chemicals. The effects of these substances could interfere with hormone biosynthesis/action and could potentially be transmitted to further generations. Thus proper education about these chemicals can help individuals decide to limit exposure, ultimately alleviating the risk on future generations.

Santiago Radío, Santiago Radío, Rafael Sebastián Fort et al.

Most signals involved in post-transcriptional regulatory networks are located in the untranslated regions (UTRs) of the mRNAs. Therefore, to deepen our understanding of gene expression regulation, delimitation of these regions with high accuracy is needed. The trypanosomatid lineage includes a variety of parasitic protozoans causing a significant worldwide burden on human health. Given their peculiar mechanisms of gene expression, these organisms depend on post-transcriptional regulation as the main level of gene expression control. In this context, the definition of the UTR regions becomes of key importance. We have developed UTR-mini-exon (UTRme), a graphical user interface (GUI) stand-alone application to identify and annotate 5′ and 3′ UTR regions in a highly accurate way. UTRme implements a multiple scoring system tailored to address the issue of false positive UTR assignment that frequently arise because of the characteristics of the intergenic regions. Even though it was developed for trypanosomatids, the tool can be used to predict 3′ sites in any eukaryote and 5′ UTRs in any organism where trans-splicing occurs (such as the model organism C. elegans). UTRme offers a way for non-bioinformaticians to precisely determine UTRs from transcriptomic data. The tool is freely available via the conda and github repositories.

Natalia Andreeva, Alexander Belyavsky

Coating tissue culture vessels with the components of the extracellular matrix such as fibronectin and collagens provides a more natural environment for primary cells in vitro and stimulates their proliferation. However, the effects of such protein layers are usually rather modest, which might be explained by the loss immobilized proteins due to their weak non-covalent association with the tissue culture plastic. Here we describe a simple protocol for a controlled fixation of fibronectin, vitronectin and collagen IV layers by formaldehyde, which substantially enhances the stimulation of primary cell proliferation by these extracellular proteins.

Lima A.C.J., Lara M.A.C.

Christopher E Laumer, Andreas Hejnol, Gonzalo Giribet

Flatworms number among the most diverse invertebrate phyla and represent the most biomedically significant branch of the major bilaterian clade Spiralia, but to date, deep evolutionary relationships within this group have been studied using only a single locus (the rRNA operon), leaving the origins of many key clades unclear. In this study, using a survey of genomes and transcriptomes representing all free-living flatworm orders, we provide resolution of platyhelminth interrelationships based on hundreds of nuclear protein-coding genes, exploring phylogenetic signal through concatenation as well as recently developed consensus approaches. These analyses robustly support a modern hypothesis of flatworm phylogeny, one which emphasizes the primacy of the often-overlooked ‘microturbellarian’ groups in understanding the major evolutionary transitions within Platyhelminthes: perhaps most notably, we propose a novel scenario for the interrelationships between free-living and vertebrate-parasitic flatworms, providing new opportunities to shed light on the origins and biological consequences of parasitism in these iconic invertebrates.

Ivan Lilyanov Dzhagalov, Katherine Grace Chen, Paul Herzmark et al.

The elimination of autoreactive T cells occurs via thymocyte apoptosis and removal by thymic phagocytes, but the sequence of events in vivo, and the relationship between thymocyte death and phagocytic clearance, are unknown. Here we address these questions by following a synchronized cohort of thymocytes undergoing negative selection within a three-dimensional thymic tissue environment, from the initial encounter with a negative selecting ligand to thymocyte death and clearance. Encounter with cognate peptide-MHC complexes results in rapid calcium flux and migratory arrest in auto-reactive thymocytes over a broad range of peptide concentrations, followed by a lag period in which gene expression changes occurred, but there was little sign of thymocyte death. Caspase 3 activation and thymocyte loss were first detectable at 2 and 3 hours, respectively, and entry of individual thymocytes into the death program occurred asynchronously over the next 10 hours. Two-photon time-lapse imaging revealed that thymocyte death and phagocytosis occurred simultaneously, often with thymocytes engulfed prior to changes in chromatin and membrane permeability. Our data provide a timeline for negative selection and reveal close coupling between cell death and clearance in the thymus.

Ximing Du, Abdulla S. Kazim, Andrew J. Brown et al.

The endosomal sorting complex required for transport (ESCRT) plays a crucial role in the degradation of ubiquitinated endosomal membrane proteins. Here, we report that Hrs, a key protein of the ESCRT-0 complex, is required for the transport of low-density lipoprotein-derived cholesterol from endosomes to the endoplasmic reticulum. This function of Hrs in cholesterol transport is distinct from its previously defined role in lysosomal sorting and downregulation of membrane receptors via the ESCRT pathway. In line with this, knocking down other ESCRT proteins does not cause prominent endosomal cholesterol accumulation. Importantly, the localization and biochemical properties of key cholesterol-sorting proteins, NPC1 and NPC2, appear to be unchanged upon Hrs knockdown. Our data identify Hrs as a regulator of endosomal cholesterol trafficking and provide additional insights into the budding of intralumenal vesicles.

David C. Diehl, Stephanie C. Toelle

Now available in Spanish, this is part of a series of publications by David C. Diehl and Stephanie C. Toelle that addresses media and its effects on young children, from birth to age six. The series is designed to reach both professional educators and parents and to provide research summaries, tips for parents, and internet resources for further information. Spanish versions published by the UF Department of Family Youth and Community Sciences, June 2011. UF/IFAS photo: Thomas Wright.  

Linda B. Bobroff, Leigh Ann Martin

Revised! FCS-8587-Eng, a 2-page fact sheet by Linda B. Bobroff and Leigh Ann Martin, lists trustworthy sites related to nutrition, health, food safety, and resources for older people. Published by the UF Department of Family Youth and Community Sciences, July 2009.