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Since the onset of the COVID-19 pandemic, a variety of diagnostic approaches, including RT-qPCR, RAPID, and LFA, have been adopted, with RT-qPCR emerging as the gold standard. However, a significant challenge in COVID-19 diagnostics is the wide range of symptoms presented by patients, necessitating early and accurate diagnosis for effective management. Although RT-qPCR is a precise molecular technique, it is not immune to false-negative results. In contrast, CRISPR-based detection methods for SARS-CoV-2 offer several advantages: they are cost-effective, time-efficient, highly sensitive, and specific, and they do not require sophisticated instruments. These methods also show promise for scalability, enabling diagnostic tests. CRISPR technology can be customized to target any genomic region of interest, making it a versatile tool with applications beyond diagnostics, including therapeutic development. The CRISPR/Cas systems provide precise gene targeting with immense potential for creating next-generation diagnostics and therapeutics. One of the key advantages of CRISPR/Cas-based therapeutics is the ability to perform multiplexing, where different sgRNAs or crRNAs can target multiple sites within the same gene, reducing the likelihood of viral escape mutants. Among the various CRISPR systems, CRISPR/Cas13 and CARVER (Cas13-assisted restriction of viral expression and readout) are particularly promising. These systems can target a broad range of single-stranded RNA viruses, making them suitable for the diagnosis and treatment of various viral diseases, including SARS-CoV-2. However, the efficacy and safety of CRISPR-based therapeutics must be thoroughly evaluated in pre-clinical and clinical settings. While CRISPR biotechnologies have not yet been fully harnessed to control the current COVID-19 pandemic, there is an optimism that the limitations of the CRISPR/Cas system can be overcome soon. This review discusses how CRISPR-based strategies can revolutionize disease diagnosis and therapeutic development, better preparing us for future viral threats.

Background: Ubiquitination is an important post-transcriptional modification crucial for maintaining cell homeostasis. As a deubiquitination enzyme, ubiquitin-specific protease 1 (USP1) is associated with tumor progression; however, its role in bladder cancer is unknown. This study aimed to analyze USP1 expression and study its roles in bladder cancer.

Methods: The web server GEPIA was used to analyze the USP1 expression. To explore USP1's function in bladder cancer, we constructed USP1-knockout cell lines in UMUC3 cells. A FLAG-USP1 (WT USP1) plasmid and a plasmid FLAG-USP1 C90S (catalytic-inactive mutant) were used to overexpress USP1 in T24 cells. CCK8, colony formation, and Transwell assays were used to assess cell viability, proliferation, and migration. RNA-sequencing (RNA-seq) and dual-luciferase reporter assays were performed to screen the pathway. Co-immunoprecipitation and immunofluorescence were used to explore the interaction between USP1 and c-MYC. A xenograft mouse model was used to study the role of USP1 in bladder cancer.

Results: USP1 expression was upregulated in human bladder cancer cells and correlated with poor patient prognosis. USP1 overexpression promoted cell proliferation, clone formation, and migration, and this was attenuated by genetic ablation of USP1. Furthermore, we observed that USP1 deficiency inhibited tumor formation in vivo. Mechanistically, the c-MYC pathway was remarkably activated compared with the other pathways. Furthermore, USP1 could interact with c-MYC and increase c-MYC's stability depending on the catalytic activity of USP1.

Conclusions: Our results suggested that high expression of USP1 promotes bladder cancer progression by stabilizing c-MYC; hence, USP1 may serve as a novel therapeutic target for treating bladder cancer.

Preeclampsia (preE) is a hypertensive disorder in pregnancies. It is the third leading cause of mortality among pregnant women and fetuses worldwide, and there is much we have yet to learn about its pathophysiology. One complication includes cerebral edema, which causes a breach of the blood-brain barrier (BBB). Urinary marinobufagenin (MBG) is elevated in a preE rat model prior to developing hypertension and proteinuria. We investigated what effect MBG has on the endothelial cell permeability of the BBB. Human brain microvascular endothelial cells (HBMECs) were utilized to examine the permeability caused by MBG. The phosphorylation of ERK1/2, Jnk, p38, and Src was evaluated after the treatment with MBG. Apoptosis was evaluated by examining caspase 3/7. MBG ≥ 1 nM inhibited the proliferation of HBMECs by 46-50%. MBG induced monolayer permeability, causing a decrease in the phosphorylation of ERK1/2 and the activated phosphorylation of Jnk, p38, and Src. MBG increased the caspase 3/7 expression, indicating the activation of apoptosis. Apoptotic signaling or the disruption of endothelia tight junction proteins was not observed when using the p38 inhibitor as a pretreatment in MBG-treated cells. The MBG-induced enhancement of the HBMEC monolayer permeability occurs by the downregulation of ERK1/2, the activation of Jnk, p38, Src, and apoptosis, resulting in the cleavage of tight junction proteins, and are attenuated by p38 inhibition.

Birds, especially the chick and hen, have been important biomedical research models for centuries due to the accessibility of the avian embryo and the early discovery of avian viruses. Comprehension of avian tumor virology was a milestone in basic cancer research, as was that of non-viral genesis, as it enabled the discovery of oncogenes. Furthermore, studies on avian viruses provided initial insights into Kaposi's sarcoma and EBV-induced diseases. However, the role of birds in human carcinogenesis extends beyond the realm of virology research. Utilization of CAM, the chorioallantoic membrane, an easily accessible extraembryonic tissue with rich vasculature, has enabled studies on tumor-induced angiogenesis and metastasis and the efficient screening of potential anti-cancer compounds. Also, the chick embryo alone is an effective preclinical in vivo patient-derived xenograft model, which is important for the development of personalized therapies. Furthermore, adult birds may also closely resemble human oncogenesis, as evidenced by the laying hen, which is the only animal model of a spontaneous form of ovarian cancer. Avian models may create an interesting alternative compared with mammalian models, enabling the creation of a relatively cost-effective and easy-to-maintain platform to address key questions in cancer biology.

The multiprotein Target of Rapamycin (TOR) Complex 1 (TORC1) is a serine/threonine kinase that stimulates anabolic metabolism and suppresses catabolism. Deregulation of TORC1 is implicated in various human pathologies, including cancer, epilepsy, and neurodegenerative disorders. The Gap Activity Towards Rags (GATOR) complex contains two subcomplexes: GATOR1, which inhibits TORC1 activity; and GATOR2, which counteracts GATOR1s function. Structural and biochemical studies have elucidated how GATOR1 regulates TORC1 activity by acting as a GTPase activating protein for Rag GTPase. However, while cryogenic electron microscopy has determined that the structure of the multi-protein GATOR2 complex is conserved from yeast to humans, how GATOR2 inhibits GATOR1 remains unclear. Here, we describe recent whole-animal studies in Drosophila that have yielded novel insights into GATOR2 function, including identifying a novel role for the GATOR2 subunit WDR59, redefining the core proteins sufficient for GATOR2 activity, and defining a TORC1-independent role for GATOR2 in the regulation of the lysosomal autophagic endomembrane system. Additionally, the recent characterization of a novel methionine receptor in Drosophila that acts through the GATOR2 complex suggests an attractive model for the evolution of species-specific nutrient sensors. Research on GATOR2 function in Drosophila highlights how whole-animal genetic models can be used to dissect intracellular signaling pathways to identify tissue-specific functions and functional redundancies that may be missed in studies confined to rapidly proliferating cell lines.

The role of stress granules (SGs) in pulmonary arterial hypertension (PAH) is unknown. We hypothesized that SG formation contributes to abnormal vascular phenotypes, and cardiac and skeletal muscle dysfunction in PAH. Using the rat Sugen/hypoxia (SU/Hx) model of PAH, we demonstrate the formation of SG puncta and increased expression of SG proteins compared to control animals in lungs, right ventricles, and soleus muscles. Acetazolamide (ACTZ) treatment ameliorated the disease and reduced SG formation in all of these tissues. Primary pulmonary artery smooth muscle cells (PASMCs) from diseased animals had increased SG protein expression and SG number after acute oxidative stress and this was ameliorated by ACTZ. Pharmacologic inhibition of SG formation or genetic ablation of the SG assembly protein (G3BP1) altered the SU/Hx-PASMC phenotype by decreasing proliferation, increasing apoptosis and modulating synthetic and contractile marker expression. In human PAH lungs, we found increased SG puncta in pulmonary arteries compared to control lungs and in human PAH-PASMCs we found increased SGs after acute oxidative stress compared to healthy PASMCs. Genetic ablation of G3BP1 in human PAH-PASMCs resulted in a phenotypic switch to a less synthetic and more contractile phenotype. We conclude that increased SG formation in PASMCs and other tissues may contribute to PAH pathogenesis.

Nicotinamide adenine dinucleotide (NAD⁺) is indispensable for the regulation of biological metabolism. Previous studies have revealed its role in aging and degenerative diseases, while crucially showing that supplementation with NAD⁺ or its precursors could ameliorate or reverse the progression of aging. Despite extensive evidence for the role and action of NAD⁺ in aging, its pharmacological activity on the skin, or even its mechanism, has not been elucidated. In this study, we established a novel approach to effectively utilize NAD⁺ for skin anti-aging by enhancing the pharmacological efficacy of exogenous NAD⁺ using a phytochemical complex consisting of quercetin, and enoxolone through inhibition of CD38. Through the comprehensive in vitro experiments based on human fibroblasts, we observed that exogenous NAD⁺ could exert protective effects against both extrinsic aging induced by ultraviolet light exposure and intrinsic aging. Additionally, we found that its effects were significantly boosted by quercetin and enoxolone. In this in-depth study, we demonstrated that these beneficial effects are mediated by improved sirtuin activation, autophagy, and mitochondrial functionality. Our approach is expected to verify the applicability of the topical application of NAD⁺ and offer more effective solutions for the unmet needs of patients and consumers who demand more effective anti-aging effects.

Thyroid hormones play a crucial role in the development of the central nervous system and are considered pivotal to cognitive functions in the adult brain. Recently, thyroid dysfunction has been associated with Alzheimer's disease. The aim of this study was to assess the neuroprotective effects of triiodothyronine (T3) on insulin signaling, neuroinflammation, apoptosis, and cognitive function in a streptozotocin (STZ)-induced sporadic Alzheimer's disease-like model. Male Wistar rats underwent stereotaxic surgery for intracerebroventricular injections of streptozotocin (STZ; 2 mg/kg) or vehicle in the lateral ventricles to induce an AD-like model. The animals received a daily dose of 1.5 μg of T3/100 g body weight or the same volume of vehicle for 30 days and were subdivided into four experimental groups: (1) animals receiving citrate treated with saline (Control = CTL); (2) animals receiving citrate treated with T3 (T3); (3) animals receiving STZ treated with saline (STZ); and (4) animals receiving STZ treated with T3 (STZ + T3). The novel object recognition test was used to measure cognitive function. Serum analysis, real-time RT-PCR, immunohistochemistry, and immunoblotting analyses were also carried out. Our results demonstrated that T3 treatment reversed cognitive impairment and increased Akt and GSK3 phosphorylation in the treated group, while also reducing microglial activation (Iba-1) and GFAP expression (reactive astrocytes), along with TNF-α, IL-6, and IL-1β levels in the hippocampus. Additionally, T3 treatment increased levels of the anti-apoptotic protein Bcl-2 and reduced the expression of the pro-apoptotic protein BAX in the hippocampus. Our study demonstrated that T3 could potentially protect neurons in an AD model induced by STZ.

Viruses are intracellular parasites that utilize organelles, signaling pathways, and the bioenergetics machinery of the cell to replicate the genome and synthesize proteins to build up new viral particles. Mitochondria are key to supporting the virus life cycle by sustaining energy production, metabolism, and synthesis of macromolecules. Mitochondria also contribute to the antiviral innate immune response. Here, we describe the different mechanisms involved in virus-mitochondria interactions. We analyze the effects of viral infections on the metabolism of glucose in the Warburg phenotype, glutamine, and fatty acids. We also describe how viruses directly regulate mitochondrial function through modulation of the activity of the electron transport chain, the generation of reactive oxygen species, the balance between fission and fusion, and the regulation of voltage-dependent anion channels. In addition, we discuss the evasion strategies used to avoid mitochondrial-associated mechanisms that inhibit viral replication. Overall, this review aims to provide a comprehensive view of how viruses modulate mitochondrial function to maintain their replicative capabilities.

Post-COVID-19 syndrome (PCS) is defined by the persistence or recurrence of symptoms after an initial acute SARS-CoV-2 infection [...].