Phosphatidyl amino acids (p-AAs) are metabolites characterized by the phosphorylation of the hydroxyl, amino, carboxyl, and thiol groups of amino acids. Previous research has primarily focused on the phosphorylation sites within macromolecular proteins, with a particular emphasis on typical O-p-AAs. In this study, we established a prediction library of p-AAs based on existing knowledge. To improve detection rates of p-AAs in biological samples, we employed a chemical labeling-based LC-MS/MS method, utilizing p-[3,5-(dimethylamino)-2,4,6-triazine] benzene-1-sulfonyl piperazine (Tmt-PP) and its deuterated form (d12-Tmt-PP) as paired labeling reagents. A preliminary identification was performed by matching characteristic MS fragments with available standards. Additionally, strategies such as in vitro methods were implemented for further identification. The phosphatase treatment aids in identifying phosphate-modified metabolites by dephosphorylating them, while cell extract incubation helps determine if novel phosphorylated amino acids are generated in vivo. Ultimately, we identified 11 p-AAs, 6 of which are novel metabolites reported for the first time. A pseudotargeted metabolomics method covering 11 identified p-AAs was established and applied to investigate the differences between cisplatin-resistant non-small cell lung cancer (NSCLC) cells and their parental cells, as well as their derived exosomes. This approach enhances our understanding of the role of p-AAs in various health and disease conditions and contributes to the discovery of additional novel phosphatidyl metabolites.

Modulations of mitochondrial dysfunction, which involve a series of dynamic processes such as mitochondrial biogenesis, mitochondrial fusion and fission, mitochondrial transport, mitochondrial autophagy, mitochondrial apoptosis, and oxidative stress, play an important role in the onset and progression of stroke. With a better understanding of the critical role of mitochondrial dysfunction modulations in post-stroke neurological injury, these modulations have emerged as a potential target for stroke prevention and treatment. Additionally, since effective treatments for stroke are extremely limited and natural products currently offer some outstanding advantages, we focused on the findings and mechanisms of action related to the use of natural products for targeting mitochondrial dysfunction in the treatment of stroke. Natural products achieve neuroprotective through multi-target regulation of mitochondrial dysfunction encompassing the following processes: (1) Mitochondrial biogenesis: Cordyceps and hydroxysafflor yellow A activate the peroxisome proliferator-activated receptor gamma coactivator 1-alpha/nuclear respiratory factor pathway, promote mitochondrial DNA replication and respiratory chain protein synthesis, and thereby restore energy supply in the ischemic penumbra. (2) Mitochondrial dynamics balance: Ginsenoside Rb3 promotes Opa1-mediated neural stem cell migration and diffusion for recovery of damaged brain tissue. (3) Mitochondrial autophagy: Gypenoside XVII selectively eliminates damaged mitochondria via the phosphatase and tensin homolog-induced kinase 1/Parkin pathway and blocks reactive oxygen species and the NOD-like receptor protein 3 inflammasome cascade, thereby alleviating blood-brain barrier damage. (4) Anti-apoptotic mechanisms: Ginkgolide K inhibits Bax mitochondrial translocation and downregulates caspase-3/9 activity, reducing neuronal programmed death induced by ischemia-reperfusion. (5) Oxidative stress regulation: Scutellarin exerts antioxidant properties and improves neurological function by modulating the extracellular signal-regulated kinase 5-Kruppel-like factor 2-endothelial nitric oxide synthase signaling pathway. (6) Intercellular mitochondrial transport: Neuroprotective effects of Chrysophanol are associated with accelerated mitochondrial transfer from astrocytes to neurons. Existing studies have confirmed that natural products exhibit neuroprotective effects through multidimensional interventions targeting mitochondrial dysfunction in both ischemic and hemorrhagic stroke models. However, their clinical translation still faces challenges, such as the difficulty in standardization due to component complexity, insufficient cross-regional clinical data, and the lack of long-term safety evaluations. Future research should aim to integrate new technologies, such as single-cell sequencing and organoid models, to deeply explore the mitochondria-targeting mechanisms of natural products and validate their efficacy through multicenter clinical trials, providing theoretical support and translational pathways for the development of novel anti-stroke drugs.

The spatio-temporal detection of proteins in living cells has always been an ongoing challenge. This complexity stems from the dynamic nature of the cellular environment, in which proteins interact in a highly regulated manner. Some innovative methods collectively represent important advances in overcoming the challenges associated with spatiotemporal detection of proteins in living cells. They are essential for elucidating cell biology and advancing therapeutic diagnostic techniques. However, the spatio-temporal organization of living intracellular vessels still lacks sufficient characterization. At present, there is a lack of review articles that comprehensively review the progress in recent years. This review aims to fill this gap by providing a broad overview of protein analysis in single living cells. In this review, the recent technical advances in the spatio-temporal analysis of proteins in single living cells are investigated, and the prospects and challenges in this direction are briefly discussed.

BackgroundSalt stress poses a genuine threat to plants, impeding their growth, development, and yields. Platostoma palustre (Blume) A.J.Paton (P. palustre) is an important medicinal plant in tropical and subtropical regions; however, the molecular mechanisms underlying P. palustre response to salt stress remain poorly understood. To better understand the molecular response of P. palustre plants to salt stress, we employed an integrated approach based on physiological, biochemical, and transcriptomic analyses.ResultsIn this study, salt stress significantly restrained the growth of P. palustre and led to the accumulation of antioxidant enzymes (SOD, POD, and CAT). Besides, the chlorophyll content significantly decreased with the increase in NaCl concentration. Transcriptomic analysis revealed 8,679 differentially expressed genes (DEGs) (4,334 were up-regulated and 4,363 were down-regulated) between control (CK) and salt stress (150 mM NaCl). KEGG enrichment analysis showed that these DEGs were significantly enriched in “plant hormone signal transduction”, “MAPK signaling pathway-plant”, “photosynthesis - antenna proteins”, “starch and sucrose metabolism”, etc. Among the DEGs, 409 DEGs were identified as transcription factors (TFs), belonging to 25 families including MYB_superfamily, AP2/ERF, C2C2, bHLH, NAC, WRKY, and so on. In KEGG enrichment analysis of the identified TFs, 13 showed significant enrichment in “plant hormone signal transduction”. Notably, EIN3 (TRINITY_DN3357_c1_g2) and ERF1 (TRINITY_DN8842_c0_g1) involved in the ethylene (ETH) signaling were suggested potential candidates for salt stress response in P. palustre.ConclusionsThis study unravels key salt-responsive genes in P. palustre, facilitating the development of salinity-resistance varieties.

BackgroundThe emergence of innovative anticancer medicines has revitalized cancer treatment prospects, and improving the accessibility of innovative anticancer medicines is a key goal of national pharmaceutical policies. Despite significant efforts by the National Medical Security Administration to reform the National Reimbursement Drug List (NRDL), concerns regarding the utilization of these newly included anticancer drugs persist. This study aims to assess the accessibility of 23 Innovative Negotiated Anticancer Medicines (INAMs) in Jiangsu Province, a developed region in eastern China.MethodsA retrospective survey was conducted across 319 healthcare institutions in Jiangsu Province, with 285 included for analysis. Data were obtained from procurement records of 23 INAMs. We evaluated the use of these medicines based on three aspects: availability, price measured by the defined daily dose cost (DDDc), and affordability.Results43.5% of the 23 INAMs encountered difficulties in obtaining them, and 30.4% of 23 INAMs were very difficult to obtain. There was a notable disparity in the availability of 23 INAMs between secondary and tertiary healthcare institutions. The median DDDc for Group A and Group C showed a progressive annual reduction. The DDDc of 19 INAMs decreased during the study period, while three drugs remained unchanged, and one drug's DDDc actually increased. Family poverty caused by medicines was more pronounced among rural residents. The affordability of rural patients was significantly lower than that of urban patients (p < 0.05) in 2021 and 2022.ConclusionThe accessibility of INAMs improved from 2020 to 2022. However, the accessibility of INAMs is influenced by various factors, and disparities still exist in access across different healthcare institutions and between urban and rural areas. Effective and sustainable policies need to be developed to ensure equitable access to medicines for all patients.

Acute ischemic stroke (AIS) subtypes exhibit distinct pathophysiological mechanisms. While current classification methods predominantly depend on neuroimaging, there remains a critical need for sensitive biomarkers to complement imaging and enhance early subtype identification in clinical settings. This study employed metabolomics to identify such biomarkers. A total of 320 AIS patients within 48 h of symptom onset were enrolled, including 227 with large artery atherosclerosis (LAA) and 93 with small vessel occlusion (SVO). Participants were divided into a discovery cohort (n = 177) and a validation cohort (n = 143) based on enrollment order. Pseudotargeted serum metabolomic profiling was performed using liquid chromatography-tandem mass spectrometry (LC-MS/MS). Distinct metabolomic signatures were observed between LAA and SVO subtypes. Three differentiating metabolites—glycoursodeoxycholic acid, docosapentaenoic acid (22n-6), and FAHFA 38:4—were consistently identified and validated across both cohorts. Combined analysis of these metabolites significantly enhanced the discriminatory power for AIS subtype differentiation. This study presents these three circulating metabolites that have the potential to serve as novel biomarkers for the early differentiation between LAA and SVO subtypes of AIS.

This study aims to investigate the protective effects of curcumin (CUR) in high glucose (HG)-induced oxidative stress and apoptosis of primary cardiomyocytes by activating the Notch1 signaling pathway. CUR is a natural polyphenol isolated from turmeric rhizomes and is known for its antioxidant, anti-apoptotic, and anti-inflammatory effects, particularly relevant in diabetes.Therefore, we used neonatal rat cardiomyocytes exposed to HG conditions, followed by treatment with CUR and DAPT, respectively. We detected and assessed myocardial cells viability and antioxidant enzyme activity by CCK-8 reagent and antioxidant enzyme kit. Apoptosis was detected by flow cytometry. The production of reactive oxygen species was detected by fluorescence labeling, and the expression of related genes and proteins was detected by qRT-PCR and Western blot. HG-induced primary rat cardiomyocytes not only increased apoptosis and ROS production, but also decreased the activity of antioxidant enzymes and the expression of Notch1 and Hes1 proteins. After pre-treatment by CUR, surprisingly, we found that CUR markedly improved viability of HG-treated cardiomyocytes. The results showed that CUR could inhibit the apoptosis of rat cardiomyocytes, inhibit the production of intracellular ROS, and increase the activity of antioxidant enzymes. Further, we found that CUR can upregulate the expression of Notch1 and Hes1 proteins and related genes, suggesting that the protective effect of CUR on HG-induced damage involves the Notch1/Hes1 signaling. These results suggest that CUR protects cardiomyocytes from HG-induced oxidative stress by activating Notch1 and its downstream target genes.