Biochemical and biophysical research communications最新文献

Deep vein thrombosis (DVT) remains a significant health problem. Although animal models have provided significant insights into the DVT pathophysiology, time-course assessment in a same animal is technically limited. Recently, we reported a novel murine saphenous DVT model for in vivo visualization of spatiotemporal dynamics of inflammatory cells. This study further shed a light on the resolution and organization process of DVT using serial in vivo imaging technique. Similar with ferric chloride-induced thrombus model, our saphenous DVT model allowed serial in vivo imaging with fluorescence microscopy. However, unlike ferric chloride-induced thrombus model, we observed a significant decrease of DVT burden. Red blood cells area gradually decreased followed by fibrin and collagen deposition over time, although ferric chloride model induced platelet-rich arterial thrombus. Histological assessment revealed that neutrophils influx peaked 3 h after DVT induction, followed by macrophages' migration at 120 h' post-induction, indicating similar organization process with traditional stasis-induced DVT model. Ly6G/Ly6C positive cells at 3 h predicted the reduction of DVT burden (r > 0.8; P < 0.01), suggesting that inflammatory response at acute phase plays pivotal role in DVT resolution. MMP-9 expression was observed and colocalized with neutrophils at early timepoints in both traditional stasis-induced DVT model and our femoral imaging models. Taken together, our in vivo imaging model might allow better understanding of the resolution and organization processes in DVT.

Exercise benefits the brain, particularly the learning and memory center-the dorsal hippocampus (dHPC)-and holds promise for therapeutic applications addressing age-related cognitive deficits. While moderate-to-vigorous-intensity exercise is commonly recommended for health benefits, our translational research proposes the effectiveness of very-light-intensity exercise in enhancing cognitive functions. However, the intensity-dependent characteristics of HPC activation have yet to be fully delineated; therefore, there is no evidence of whether such easily accessible exercises for people of all ages and most fitness levels can activate HPC neurons. Here, we aimed to clarify this question using a physiologically sound rat exercise model. We used a previously established rat treadmill running model within a metabolic chamber and measured maximal oxygen uptake (V˙O_2max) during an incremental running test. Referring to the American College of Sports Medicine's V˙O_2max-based intensity classification, rats were assigned to one of five groups: resting control, very-light, light, moderate, and vigorous exercise intensity. We immunohistochemically assessed the effects of a single bout of exercise on dHPC neuronal activity and measured V˙O₂ and blood lactate as exercise intensity indicators. dHPC neuronal activity increased with exercise intensity, even at light-intensity without blood lactate accumulation, and correlated positively with increasing V˙O₂. The dorsal dentate gyrus and CA1 sub-regions were markedly activated even by very-light-intensity exercise. Our findings demonstrate the intensity-dependent activation of dHPC neurons, with very-light-intensity exercise as the minimal intensity threshold. These strongly support our hypothesis that very-light-intensity exercise serves as a viable memory-enhancing strategy, beneficial for various populations including low-fitness individuals and the elderly.

Molecular targeted cancer therapy is a rapidly developing field, driving progress toward greater treatment efficacy. However, targeted monotherapy often fails due to the development of multidrug resistance in tumors. The combination of multiple targeted agents emerges as a possible solution to enhance treatment outcomes by activating different signaling pathways. This study systematically investigates the combined effect of targeted agents for the oncomarkers HER2 and EpCAM on cancer cells. Specifically, the study examined the impact of anti-HER2 (DARP_9.29-LoPE) and anti-EpCAM (DARP_EC1-LoPE) immunotoxins on a panel of cancer cells expressing various levels of HER2 and EpCAM. Using the Chou-Talalay combination indices, the study revealed that cells with low HER2 expression and high EpCAM expression are not optimal targets for combined HER2/EpCAM therapy. In contrast, the most effective approach involves the usage of an equimolar ratio of immunotoxins for cells exhibiting high HER2 and moderate EpCAM expression, resulting in a synergistic therapeutic effect. These findings provide significant insights into optimizing combination anti-HER2/EpCAM therapies and hold promise for the development of more effective cancer treatment strategies.

Glioma is the most common primary intracranial malignant tumor in adults, with a poor prognosis. Exosomes released by tumor cells play a crucial role in tumor development, metastasis, angiogenesis, and other biological processes. Despite this significance, the precise molecular mechanisms governing exosome secretion and their impact on tumor progression remain incompletely understood. While Choline Kinase Alpha (CHKA) has been implicated in promoting various types of tumors, its specific role in glioma pathogenesis remains unclear. Our study initially demonstrates that CHKA enhances the proliferation, migration, and invasion abilities of glioma cells. Interestingly, CHKA also stimulates the release of exosomes from glioma cells. Mechanistically, reduced CHKA expression hampers exosome secretion by elevating autophagy levels in gliomas, whereas counteracting the autophagy elevation resulting from CHKA downregulation restores the release of exosomes. Notably, exosomes derived from glioma cells with normal CHKA expression exhibit a greater capacity to promote glioma progression compared to those derived from cells with low CHKA expression. Overall, our findings suggest that CHKA modulates exosome secretion via an autophagy-dependent pathway, thereby facilitating the proliferation, migration, and invasion of glioma cells.

Introduction: Paraoxonase-1 (PON1) is a crucial esterase in cardiovascular health, closely associated with HDL and known for its antioxidant and anti-inflammatory properties. Reduced PON1 activity has been linked to cardiovascular diseases. Lysophospholipids (LysoPLs), essential for cellular processes and immune responses, are implicated in the pathogenesis of cardiovascular diseases and are bound to lipoproteins, contributing to their diverse effects. Thus, we hypothesize that the relationship between PON1 and cardiovascular diseases may involve the modulation of LysoPLs by PON1. This study aims to investigate how PON1 potentially influences LysoPLs.

Methods: We quantified the levels of LysoPLs in HepG2 cells by using liquid chromatography-mass spectrometry, manipulating PON1 expression or knockdown.

Results: In cells overexpressing PON1, there was a significant increase in cellular levels of lysophosphatidylserine (LysoPS) and medium levels of LysoPS. Conversely, in cells with PON-1 knockdown, cellular levels of lysophosphatidylcholine (LysoPC) and medium levels of LysoPC showed a significant decrease.

Conclusions: PON1 is involved in modulating LysoPLs, which contribute to the antioxidant and anti-inflammatory properties of HDL, often attributed to PON1.

Bone morphogenetic protein (BMP)-3b, also known as growth differentiation factor (GDF)-10, belongs to the transforming growth factor (TGF)-β superfamily. Despite being named a BMP, BMP3b is considered as an intermediate between the TGFβ/activin/myostatin and BMP/GDF subgroups of the TGFβ superfamily. Myoblast differentiation is tightly regulated by various cytokines, including the TGFβ superfamily members. However, despite BMP3b supporting the maintenance of skeletal myofibers, myoblast differentiation induced by BMP3b remains unclear. In this study, BMP3 expression levels in isolated satellites were very low compared to those in the skeletal muscle tissues. We analyzed cardiotoxin-induced muscle regeneration. Intact muscle fiber size was larger in BMP3b null mice than in wild-type mice; however, regenerated muscle fiber size did not differ between the null and wild-type mice. Next, we analyzed the satellite cell-specific BMP3b-overexpressing (BMP3b Tg) mice. Intact fiber size was increased in BMP3b Tg mice. However, regenerating tibialis anterior muscle size was reduced in BMP3b Tg mice compared to that in control mice. BMP3b overexpression in C2C12 cells stimulated Smad2/3 signaling. Moreover, BMP3b overexpression and conditioned medium of BMP3b-expressing Chinese hamster ovary cells strongly suppressed myoblast differentiation by repressing transactivation. Overall, our data suggest that BMP3b is not necessary for muscle regeneration; however, excessive BMP3b interferes with muscle regeneration by suppressing myoblast differentiation.

During avian germ cell formation, primordial germ cells (PGCs) differentiate into prospermatogonia in testicular seminiferous tubules or into oogonia in the ovarian cortex in late-stage embryos. Although estrogenic endocrine-disrupting chemicals (EDCs) have been suggested to affect the differential fate of avian germ cells, there is currently no established method to examine the effects of EDCs on the differentiation potential of germline cells due to large amount of unidentified proteins present in avian germ cells. Regarding reliable molecular probes for the detection of germ cells that differentiated from the PGCs of Japanese quail, the prospermatogonium and oogonium, respectively, integrin beta1 (ITGB1), insulin-like growth factor 2-binding protein 1 (IGF2BP1), and stimulated by retinoic acid 8 (STRA8) were identified as marker proteins by RNA-seq and liquid chromatography tandem mass spectrometry analyses. This study also showed disordered germ cell formation in ovo following the addition of 100 nmol of diethylstilbestrol (DES), o, p'-dichloro-diphenyl-trichloroethane (o,p'-DDT), ethinylestradiol (EE), and bisphenol A. DES directly induced severe damage in germline cells by inhibiting their proliferation and subsequent differentiation into ITGB1-positive germ cells in males independently of disordered gonadal differentiation, while DES and o,p'-DDT decreased the number of female germ cells. In addition, EE toxicity was characterized by a reduction in IGF2BP1-germ cells due to the partial ovarian-like differentiation of male gonads. Furthermore, all EDCs exerted deleterious effects on female ovaries, which restricted differentiation into STRA8-positive oogonia. These results demonstrate that the bioaccumulation of estrogenic EDCs in birds during incubation may reduce male and female fertility.

Ammonium toxicity, resulting from prolonged use of ammonium as the sole nitrogen source, can lead to physiological and morphological disorders, ultimately stunting plant growth. Enhancing ammonium assimilation efficiency has been extensively explored as a strategy to mitigate ammonium toxicity. However, the role of AspAT, a key enzyme in nitrogen assimilation, remains underexplored. This study investigates the function of AspAT in alleviating ammonium toxicity and uncovers the underlying physiological mechanisms. The results show that two Populus AspAT genes, AspAT13 and AspAT15, exhibit the highest expression levels in roots and are induced by exogenous ammonium. Overexpression of AspAT13 and AspAT15 in transgenic plants results in increased root biomass. In these plants, the activities of key nitrogen assimilation enzymes (GS and GOGAT) are significantly enhanced, along with increases in soluble protein, soluble sugar, and free amino acid contents. Additionally, the activities of antioxidant enzymes, such as SOD and CAT, are elevated, and ammonium content in the roots is significantly reduced. Moreover, the levels of hormones, including IAA, ACC, IBA, and BR, are significantly increased in the roots of transgenic plants. Our findings suggest that AspAT13 and AspAT15 play essential roles in mitigating ammonium toxicity, a process closely linked to enhanced nitrogen assimilation, antioxidant systems, and the regulation of auxin and brassinosteroid (BR) signaling.

Objective and significance: Transforming growth factor-beta (TGF-β) plays a pivotal role in breast development by modulating tissue composition during the developmental phase. The TGFβ type II receptor (TGFβ RII) is implicated in breast cancer and represents a valuable therapeutic target. Due to the off-target side effects of many existing TGFβI/TGFβ RII inhibitors, a more targeted approach to drug discovery is necessary. This study used computational modeling and molecular dynamics simulations to screen the ChemBridge small molecule library against TGFβ RII.

Methods: This study employed high-throughput virtual screening, molecular dynamics simulations, and binding free energy calculations to identify potential inhibitors targeting TGF-β RII. MDA-MB 231 and MCF-7 breast cancer cells were used in anti-proliferative, tans-endothelial migration, and flow cytometric assays for in vitro validations.

Results: We identified 8-(2-methylphenyl)-9H-benzo[f]indeno[2,1-c]quinolin-9-one (C-5635020) as a potent and selective inhibitor. Protein-ligand modeling analysis revealed that C-5635020 targets the kinase domain of TGFβ RII with superior binding affinities compared to the standard drug, staurosporine. Computational results suggest that C-5635020 selectively binds and inhibits TGFβ RII activity, thereby controlling cell proliferation in breast cancer. In vitro, experiments corroborated these predictions, where C-5635020 inhibited TGFβ RII and p-Smad 2/3 positive population in MDAMB-231 and MCF-7 cells. The compound dose-dependently inhibited cell proliferation, trans-endothelial migration, and increased apoptosis in both breast cancer cell lines.

Conclusion: The strong binding affinity, stability, and favorable thermodynamics of C-5635020 with established in vitro efficacy highlight its potential as a lead compound for further preclinical and clinical developments for breast cancer treatment.

In budding yeast, endosomal sorting complex required for transport (ESCRT) mediates microautophagy by vacuolar membrane invagination into the vacuolar lumen, followed by Vps4-assisted membrane constriction and abscission. Here, we show that ESCRT elicits vacuolar fission in the absence of Vps4 after nutrient starvation, although vacuolar fusion is facilitated in wild-type cells in these conditions. ESCRT mediated vacuolar membrane invagination in vps4Δ cells, thereby causing vacuolar fission. It is known that vacuolar fission requires phosphatidylinositol 3,5-bisphosphate (PI(3,5)P2) and β-propellers that bind polyphosphoinositides (PROPPINs), PI(3,5)P2-binding proteins. However, PROPPIN, but not PI(3,5)P2, was dispensable for the ESCRT-mediated vacuolar fragmentation. Finally, we showed evidence that microlipophagy triggers vacuolar fission. Thus, disruption of the coordinated sequence of ESCRT-Vps4 operations in microautophagy leads to vacuolar fragmentation. This study provides insight into the ESCRT-Vps4 axis-dependent cellular disfunctions and related diseases.