Biochimica et biophysica acta. Molecular cell research最新文献

Exposure to the non-protein amino acid cyanotoxin β–N-methylamino-L-alanine (BMAA), released by cyanobacteria found in many water reservoirs has been associated with neurodegenerative diseases. We previously demonstrated that BMAA induced cell death in both retina photoreceptors (PHRs) and amacrine neurons by triggering different molecular pathways, as activation of NMDA receptors and formation of carbamate-adducts was only observed in amacrine cell death. We established that activation of Retinoid X Receptors (RXR) protects retinal cells, including retina pigment epithelial (RPE) cells from oxidative stress-induced apoptosis. We now investigated the mechanisms underlying BMAA toxicity in these cells and those involved in RXR protection.

BMAA addition to rat retinal neurons during early development in vitro increased reactive oxygen species (ROS) generation and polyADP ribose polymers (PAR) formation, while pre-treatment with serine (Ser) before BMAA addition decreased PHR death. Notably, RXR activation with the HX630 agonist prevented BMAA-induced death in both neuronal types, reducing ROS generation, preserving mitochondrial potential, and decreasing TUNEL-positive cells and PAR formation. This suggests that BMAA promoted PHR death by substituting Ser in polypeptide chains and by inducing polyADP ribose polymerase activation. BMAA induced cell death in ARPE-19 cells, a human epithelial cell line; RXR activation prevented this death, decreasing ROS generation and caspase 3/7 activity.

These findings suggest that RXR activation prevents BMAA harmful effects on retinal neurons and RPE cells, supporting this activation as a broad-spectrum strategy for treating retina degenerations.

Polycystic ovary syndrome (PCOS) is a common endocrine disorder characterized by chronic ovulation dysfunction and overproduction of androgens. Women with PCOS are commonly accompanied by insulin resistance (IR), which can impair insulin sensitivity and elevate blood glucose levels. IR promotes ovarian cysts, ovulatory dysfunction, and menstrual irregularities in women patients, leading to the pathogenesis of PCOS. Secreted frizzled-related protein 4 (SFRP4), a secreted glycoprotein, exhibits significantly elevated expression levels in obese individuals with IR and PCOS. Whereas, whether it plays a role in regulating IR-induced PCOS still has yet to be understood. In this study, we respectively established in vitro IR-induced hyperandrogenism in human ovarian granular cells and in vivo IR-induced PCOS models in mice to investigate the action mechanisms of SFRP4 in modulating IR-induced PCOS. Here, we revealed that SFRP4 expression levels in both mRNA and protein were remarkably upregulated in the IR-induced hyperandrogenism with elevated testosterone in the human ovarian granulosa cell line KGN. Under normal conditions without hyperandrogenism, overexpressing SFRP4 triggered the remarkable elevation of testosterone along with the increased nuclear translocation of β-catenin, cell apoptosis and proinflammatory cytokine IL-6. Furthermore, we found that phytopharmaceutical disruption of SFRP4 by genistein ameliorated IR-induced increase in testosterone in ovarian granular cells, and IR-induced PCOS in high-fat diet obese mice. Our study reveals that SFRP4 contributes to IR-induced PCOS by triggering ovarian granulosa cell hyperandrogenism and apoptosis through the nuclear β-catenin/IL-6 signaling axis. Elucidating the role of SFRP4 in PCOS may provide a novel therapeutic strategy for IR-related PCOS therapy.

Intermittent fasting exerts a profound beneficial influence on a spectrum of diseases through various mechanisms including regulation of immune responses, elimination of senescent- and pathogenic cells and improvement of stem cell-based tissue regeneration in a disease- and tissue-dependent manner. Our previous study demonstrated that alternate-day fasting (ADF) led to alleviation of xerostomia and sialadenitis in non-obese diabetic (NOD) mice, a well-defined model of Sjögren's syndrome (SS). This present study delved into the previously unexplored impacts of ADF in this disease setting and revealed that ADF increases the proportion of salivary gland stem cells (SGSCs), defined as the EpCAM^hi cell population among the lineage marker negative submandibular gland (SMG) cells. Furthermore, ADF downregulated the expression of p16^INK4a, a cellular senescence marker, which was concomitant with increased apoptosis and decreased expression and activity of NLRP3 inflammasomes in the SMGs, particularly in the SGSC-residing ductal compartments. RNA-sequencing analysis of purified SGSCs from NOD mice revealed that the significantly downregulated genes by ADF were mainly associated with sugar metabolism, amino acid biosynthetic process and MAPK signaling pathway, whereas the significantly upregulated genes related to fatty acid metabolic processes, among others. Collectively, these findings indicate that ADF increases the SGSC proportion, accompanied by a modulation of the SGSC property and a switch from sugar- to fatty acid-based metabolism. These findings lay the foundation for further investigation into the functionality of SGSCs influenced by ADF and shed light on the cellular and molecular mechanisms by which ADF exerts beneficial actions on salivary gland restoration in SS.

Resistance to endocrine therapy is a major clinical challenge in estrogen receptor (ER)-positive breast cancer. Obesity is associated with the clinical response to ER-positive breast cancers; however, the mechanism underlying obesity-induced resistance to endocrine therapy in ER-positive breast cancers remains unclear. In this study, we investigated the molecular mechanisms underlying obesity-induced resistance to tamoxifen (TAM), an anti-estrogen agent, in the ER-positive breast cancer cell line MCF-7 using differentiated adipocyte-conditioned medium (D-CM). Treatment of the cells with D-CM promoted TAM resistance by reducing TAM-induced apoptosis. The expression levels of the ERα target genes were higher in D-CM-treated cells than those in untreated ones. In contrast, when the cells were cultured in the presence of TAM, the expression levels were decreased, with or without D-CM. Moreover, the expression of the markers for cancer stem-like cells (CSCs) and mammosphere formation was enhanced by co-treating with D-CM and TAM, compared with TAM alone. The phosphatidylinositol-3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway was activated in MCF-7 cells by D-CM treatment, even in the presence of TAM. Inhibition of the PI3K/Akt/mTOR pathway decreased the expression levels of the CSC markers, suppressed mammosphere formation, and resensitized to TAM via inducing apoptosis in D-CM-treated cells. These results indicate that the conditioned medium of differentiated adipocytes promoted TAM resistance by inducing the CSC phenotype through activation of the PI3K/Akt/mTOR pathway in ER-positive breast cancer cells. Thus, the PI3K/Akt/mTOR pathway may be a therapeutic target in obese patients with ER-positive breast cancers.

Protein kinase D (PKD) is a family of serine/threonine kinases that play important roles in various signalling pathways in cells, including neuronal cells. In the nervous system, PKD has been shown to be involved in learning and memory formation by regulating neurotransmitter release, neurite outgrowth and dendrite development, synapse formation and synaptic plasticity. In addition, PKD has been implicated in pain perception or neuroprotection during oxidative stress. Dysregulation of PKD expression and activity has been linked to several neurological disorders, including autism and epilepsy. In this review, we summarize the current knowledge on the function of the PKD family members in neuronal cells, including the spatial regulation of their downstream signalling pathways. We will further discuss the potential role of PKD in the pathogenesis of neurological disorders.

The Ku heterodimer (Ku70/Ku80) is central to the non-homologous end-joining (NHEJ) pathway. Ku binds to the broken DNA ends and promotes the assembly of the DNA repair complex. The N-terminal Ku70 von Willebrand A (vWA) domain is known to mediate protein-protein interactions important for the repair process. In particular, the D192 and D195 residues within helix 5 of the Ku70 vWA domain were shown to be essential for NHEJ function, although the precise role of these residues was not identified. Here, we set up a miniTurbo screening system to identify Ku70 D192/D195 residue-specific interactors in a conditional, human Ku70-knockout cell line in response to DNA damage. Using fusion protein constructs of Ku70 wild-type and mutant (D192A/D195R) with miniTurbo, we identified a number of candidate proximal interactors in response to DNA damage treatment, including DNA Ligase IV (LigIV), a known and essential NHEJ complex member. Interestingly, LigIV was enriched in our wildtype screen but not the Ku70 D192A/D195R screen, suggesting its interaction is disrupted by the mutation. Validation experiments demonstrated that the DNA damage-induced interaction between Ku70 and LigIV was disrupted by the Ku70 D192A/D195R mutations. Our findings provide greater detail about the interaction surface between the Ku70 vWA domain and LigIV and offer strong evidence that the D192 and D195 residues are important for NHEJ completion through an interaction with LigIV. Altogether, this work reveals novel potential proximal interactors of Ku in response to DNA damage and identifies Ku70 D192/D195 residues as essential for LigIV interaction with Ku during NHEJ.

Ca²⁺ signalling is pivotal in T cell activation, an essential process in adaptive immune responses. Key to this activation are Ca²⁺ microdomains, which are transient increases in cytosolic Ca²⁺ concentration occurring within narrow regions between the endoplasmic reticulum (ER) and the plasma membrane (PM), lasting a few tens of milliseconds. Adhesion Dependent Ca²⁺ Microdomains (ADCM) rely on store-operated Ca²⁺ entry (SOCE) via the ORAI/STIM system. The nanometric scale at which these microdomains form poses challenges for direct experimental observation.

Following the previous work of Gil et al. [1], which introduced a three-dimensional model of the ER-PM junction, this study combines a detailed description of the Ca²⁺ fluxes at the junction with stochastic dynamics of a cluster of D-myo-inositol 1,4,5 trisphosphate receptors (IP₃R) located in the ER surrounding the junction. Because the consideration of Ca²⁺ release through the IP₃R calls for the simulation of a portion of the cytoplasm considerably larger than the junction, our study also investigates the spatial distribution of PMCAs, revealing their likely localization outside the ER-PM junction. Simulations indicate that Ca²⁺ puffs implying the opening of 2–6 IP₃Rs create ADCMs by provoking local depletions of ER Ca²⁺ stimulating Ca²⁺ entry through the ORAI1 channels. Such conditions allow the reproduction of the amplitude, duration and spatial extent of the observed ADCMs. By integrating advanced computational techniques with insights from experimental studies, our approach provides valuable information on the mechanisms governing early Ca²⁺ signalling in T cell activation, paving the way for a deeper understanding of immune responses.

Introduction

Angiogenesis is closely related to renal fibrosis; however, its basic mechanism remains unclear. In our study, we found that nuclear receptor 4A1 (NR4A1) inhibits vascular endothelial growth factor A (VEGFA)-induced angiogenesis, ameliorating renal fibrosis.

Methods

We prepared a renal fibrosis animal model with unilateral ureteral obstruction (UUO) and NR4A1 knockdown UUO mice model, Using Human umbilical vein endothelial cells (HUVECs) to conduct all in vitro experiments. We then detected and analyzed the expression levels of NR4A1 and other genes related to angiogenesis and fibrosis.

Results

The angiogenesis related genes, such as VEGFA, vascular endothelial growth factor receptor-2 (VEGFR-2), endoglin (CD105), as well as the expression of fibrosis related genes that included, α-smooth muscle actin (α-SMA), Vimentin, and Collagen I are all significantly increased in the UUO rat model. In addition, the expression of NR4A1 of the kidney tissue of UUO rats was significantly reduced. Therefore, according to the above results, we speculated that angiogenesis may exacerbate renal fibrosis and NR4A1 may repress renal fibrosis by inhibiting angiogenesis. To further verify the above results, we used VEGFA to stimulate HUVECs with (or without) overexpression or knockdown of NR4A1. The results showed that with prolonged stimulation using VEGFA, the expression of NR4A1 decreases. Overexpression of NR4A1 significantly inhibits the expression of related indicators of angiogenesis and renal fibrosis. Furthermore, knockdown of NR4A1 induces endothelial cell proliferation and migration; therefore, exacerbating angiogenesis and fibrosis. Finally, the results of NR4A1 knockdown UUO mice showed that knockdown of NR4A1 can aggravating kidney damage and induce the expression of angiogenesis and renal fibrosis related indicators, while UUO can significantly induce kidney damage, angiogenesis and renal fibrosis. When knockdown of NR4A1, renal kidney damage, angiogenesis and fibrosis becomes more severe than UUO. Thus, all of these results indicate that NR4A1 can ameliorate renal fibrosis by inhibiting angiogenesis.

Conclusions

NR4A1 can inhibit angiogenesis to ameliorate renal fibrosis.

Friedreich's ataxia (FA) is one of the most frequent inherited recessive ataxias characterized by a progressive sensory and spinocerebellar ataxia. The main causative mutation is a GAA repeat expansion in the first intron of the frataxin (FXN) gene which leads to a transcriptional silencing of the gene resulting in a deficit in FXN protein. The nature of the mutation (an unstable GAA expansion), as well as the multi-systemic nature of the disease (with neural and non-neural sites affected) make the generation of models for Friedreich's ataxia quite challenging. Over the years, several cellular and animal models for FA have been developed. These models are all complementary and possess their own strengths to investigate different aspects of the disease, such as the epigenetics of the locus or the pathophysiology of the disease, as well as being used to developed novel therapeutic approaches. This review will explore the recent advancements in the different mammalian models developed for FA.

Obesity causes a range of tissue dysfunctions that increases the risk for morbidity and mortality. Protein kinase D (PKD) represents a family of stress-activated intracellular signalling proteins that regulate essential processes such as cell proliferation and differentiation, cell survival, and exocytosis. Evidence suggests that PKD regulates the cellular adaptations to the obese environment in metabolically important tissues and drives the development of a variety of diseases. This review explores the role that PKD plays in tissue dysfunction in obesity, with special consideration of the development of obesity-mediated cardiomyopathy, a distinct cardiovascular disease that occurs in the absence of common comorbidities and leads to eventual heart failure and death. The downstream mechanisms mediated by PKD that could contribute to dysfunctions observed in the heart and other metabolically important tissues in obesity, and the predicted cell types involved are discussed to suggest potential targets for the development of therapeutics against obesity-related disease.