Miao Liu

BACKGROUND: TRP (transient receptor potential) cation channels play a crucial role in maintaining normal blood pressure and flow. We hypothesize that, during aging, TRPC6 (transient receptor potential cation channel subfamily C member 6)–mediated Ca ²⁺ influx promotes sarcoplasmic reticulum Ca ²⁺ refilling and RyR (ryanodine receptor) Ca ²⁺ sparks, thereby activating BK _Ca (large-conductance Ca ²⁺ -activated K ⁺ ) channels to limit myogenic arterial tone. METHODS: Using vascular smooth muscle cells (VSMCs) from resistance-sized small mesenteric arteries, we tested selective TRPC6 inhibitors (SH045 and SAR7334) and smooth muscle–specific TRPC6 knockout mice. Ca ²⁺ sparks, BK _Ca currents, and myogenic tone were measured in young (3–4 months) and old (12–14 months) mice. CaMKIIδ (Ca ²⁺ /calmodulin-dependent protein kinase IIδ) signaling was assessed pharmacologically and with a proteolysis-targeting chimera peptide degrader targeting CaMKIIδ. RESULTS: Inhibiting TRPC6 with SH045 or SAR7334 suppressed RyR-mediated Ca ²⁺ spark generation in VSMCs from old mice but not young mice. Similar results were found in smooth muscle–specific TRPC6 knockout mice. SH045 or SAR7334 also suppressed BK _Ca currents and elicited constriction in mesenteric arteries from old mice. These effects were due to increased translocation of TRPC6 channels to the VSMC plasma membrane, mediated by CaMKIIδ signaling. Using a proteolysis-targeting chimera peptide degrader targeting CaMKIIδ, we demonstrated the impact of CaMKIIδ on TRPC6 externalization and RyR activation in old VSMCs. CONCLUSIONS: Together, our findings identify age-dependent CaMKIIδ upregulation as a key regulator of TRPC6 membrane trafficking, thereby indirectly enhancing RyR-BKCa signaling through sarcoplasmic reticulum Ca ²⁺ handling and promoting VSMC hyperpolarization and vasodilation in aging.

The global incidence of intestinal diseases—including food allergy, inflammatory bowel disease (IBD), and colorectal cancer (CRC)—continues to escalate in parallel with the expansion of the “Western diet,” within which the high‐sugar diet (HSD) has emerged as a pivotal yet under‐recognized dietary driver. This review aims to define how HSD disrupts intestinal homeostasis and to establish a unified mechanistic framework that links three traditionally separate disorders—IBD, food allergy, and CRC—through a shared pathogenic axis. Preclinical and clinical studies over the past decades, along with foundational studies, where appropriate, were synthesized and stratified by evidence type (animal models, epidemiological cohorts, and clinical intervention trials), with particular attention to the distinct biological effects of fructose, glucose, sucrose, and high‐fructose corn syrup. The evidence indicates that HSD perturbs intestinal homeostasis through four interconnected mechanisms: remodeling of the gut microbiota toward dysbiosis with suppression of beneficial symbionts and short‐chain fatty acid (SCFA) production; dysregulation of host metabolism via SCFA depletion and a skewed bile acid profile; compromise of the intestinal barrier through mucus‐layer erosion and tight junction disassembly (“leaky gut”); and activation of innate and adaptive immune responses accompanied by immunometabolic reprogramming. These upstream events converge on a common “microbiota–barrier–immunity–metabolism” axis but diverge downstream into Th17/Th1‐driven mucosal inflammation in IBD, Th2/IgE‐mediated sensitization in food allergy, and a protumor microenvironment in CRC. In summary, HSD acts as a critical environmental stressor whose shared upstream mechanisms and divergent downstream outcomes help explain the concurrent rise of these intestinal disorders. Defining intestinal sugar‐sensing pathways and interindividual microbiome heterogeneity is expected to enable precision nutritional interventions—such as targeted dietary sugar management—for the prevention and treatment of intestinal diseases.

Abdominal aortic aneurysm (AAA) lacks effective pharmacotherapy. This study examines whether proprotein convertase subtilisin/kexin type 9 (PCSK9) drives ferroptosis in vascular smooth muscle cells (VSMCs) and whether its pharmacological degradation mitigates disease progression. PCSK9 is enriched in VSMCs of human AAA and in murine models induced by PPE or Ang II. SMC‐specific PCSK9 overexpression ( PCSK9 ^{SMC OE} ) increases aortic diameter, aggravates elastin fragmentation and collagen deposition and elevates MMP2/9 expression. Within aortic lesions, PCSK9 ^{SMC OE} enhances iron accumulation and lipid peroxidation while reducing glutathione GPX4, consistent with ferroptosis. In primary VSMCs, PCSK9 overexpression suppresses GPX4 and glutathione, increases malondialdehyde and Fe ²⁺ levels and impairs viability, whereas PCSK9 knockdown attenuates Ang II–induced ferroptosis. Mechanistically, PCSK9 triggers ferritinophagy, as shown by decreased ferritin heavy chain‐1 (FTH1) and nuclear receptor coactivator‐4 (NCOA4), an increased LC3‐II/I ratio and enhanced FTH1–LAMP1 colocalisation. Autophagy inhibition with bafilomycin A1 blocks Fe ²⁺ accumulation and rescues ferroptotic indices. The cell‐permeable peptide Cadd4 promotes PCSK9 degradation, restores FTH1 and NCOA4 and suppresses ferroptosis in VSMCs. In PPE and Ang II models, Cadd4 reduces aortic dilation, preserves medial structure and normalises ferroptosis and ferritinophagy markers. PCSK9 drives ferritinophagy‐dependent ferroptosis in VSMCs, and Cadd4 represents a promising therapeutic strategy for AAA.

Bladder cancer (BC), particularly muscle-invasive and metastatic disease, remains a major clinical challenge despite recent advances in immunotherapy. In this study, we aimed to identify a promising antitumor compound from five candidate small molecules and to explore its potential roles in BC progression. Through antiproliferative screening, cryptotanshinone (CTS) was identified as the promising candidate. Using both two-dimensional BC cell lines and three-dimensional bladder tumor organoid models, we comprehensively evaluated the effects of CTS on cell proliferation, migration, apoptosis, and organoid growth. To further explore the underlying mechanisms, transcriptomic sequencing based on bladder cancer organoid models, protein-protein interaction network analysis, and public databases (TCGA-BLCA, TIMER, and TISIDB) were integrated to examine immune-related pathways and potential molecular targets associated with CTS. GeneMANIA network prediction and molecular docking analyses were subsequently performed to investigate upstream regulatory networks and the potential interactions between CTS and key components of the cGAS-STING-IFN-I-JAK-STAT signaling pathway. Integrative analyses suggested that IFIT1, IFIT2, and IFIT3 may function as immune-associated genes potentially linked to BC progression, patient prognosis, immune cell infiltration, and PD-1/PD-L1 expression. Molecular docking results suggested that CTS may interact with core regulatory proteins within the cGAS-STING-IFN-I-JAK-STAT pathway, potentially influencing IFIT transcriptional regulation. Collectively, these findings indicate that CTS exhibits measurable antitumor and immunomodulatory effects, which may be associated with modulation of the cGAS-STING-IFN-I-JAK-STAT-IFIT signaling axis, supporting its potential as a small-molecule candidate for bladder cancer.

Systematic identification of functional non-coding regulatory variants remains a major challenge in human genetics. Conventional approaches such as large-scale CRISPR screening and genome-wide association studies (GWAS) are powerful but often prohibitively expensive, time-consuming, and experimentally intensive, limiting their scalability for locus-specific mechanistic studies. Recent advances in artificial intelligence offer the potential to partially replace or substantially augment these approaches by prioritizing regulatory variants with high functional likelihood. The RHD antigen, a major contributor to red blood cell alloimmunization, hemolytic transfusion reactions, and hemolytic disease of the fetus and newborn, serves as an excellent model for this paradigm, since coding variants alone do not fully account for differences in RHD expression. Here, we present an integrated artificial intelligence (AI)-guided and experimental framework to identify and validate functional non-coding regulatory variants governing RHD expression. We first applied AlphaGenome (AG), a deep-learning model released in 2025 for non-coding variant impact prediction, to systematically interrogate the RHD locus. By integrating multi-omics datasets, AG prioritized regulatory regions within the promoter, 5′ untranslated region (5′UTR), and intragenic regions. In silico deletion- and Single Nucleotide Polymorphism (SNP)-based perturbation analyses consistently predicted that variants within the promoter and its proximal regions, as well as within intragenic regions, exert strong suppressive effects on RHD expression. To experimentally validate these predictions, we performed CRISPR-mediated base editing in K562 cells at AG-prioritized non-coding SNP sites. Editing of a high-score predicted variant (chr1:25272434 G>A) achieved efficient base conversion and was accompanied by additional nearby edits, all predicted by AG to downregulate RHD expression. In contrast, editing of low-score predicted sites (chr1:25272422 C>T) produced much smaller functional effects. Quantitative polymerase chain reaction (qPCR) analysis of full-length RHD transcripts, together with flow cytometry–based analysis of RHD expression, confirmed strong concordance between AI-based predictions and transcriptional as well as phenotypic outcomes. Taken together, our results demonstrate that the combination of AI-guided regulatory variant prioritization and targeted base editing provides a potentially scalable and cost-effective alternative to traditional CRISPR screening for decoding functional non-coding variants in blood group genes, with direct implications for genomics-based RHD typing and transfusion medicine. To our knowledge, this study also represents the first validation of AlphaGenome predictions at the phenotypic level using wet-lab experiments.

Targeted protein degradation (TPD) represents a paradigm shift in drug discovery, moving beyond traditional binding‐based inhibition toward active removal of disease‐driving proteins. This approach has unlocked therapeutic possibilities for previously “undruggable” targets, including transcription factors like MYC and STAT3, mutant oncoproteins such as KRAS G12C, and scaffolding molecules lacking conventional binding pockets. Among TPD strategies, proteolysis‐targeting chimeras (PROTACs) have emerged as the leading clinical platform, with the first molecule entering trials in 2019 and progression to Phase III completion by 2024. This comprehensive review examines PROTAC development across diverse therapeutic areas, analyzing key targets including kinases, hormone receptors, antiapoptotic proteins, and epigenetic modulators. We evaluate clinical progression of breakthrough candidates such as ARV‐110 for prostate cancer, ARV‐471 for breast cancer, and BTK degraders, while discussing critical challenges including the “hook effect” and oral bioavailability limitations. The review explores future directions encompassing innovative delivery strategies, tissue‐specific degrader design, and approaches for expanding E3 ligase repertoires and overcoming resistance. This review provides essential foundations for rational target selection, molecular optimization, and clinical translation strategies. By integrating mechanistic insights with clinical realities, this analysis offers perspectives on PROTAC technology advancement and identifies opportunities for transforming treatment of complex diseases resistant to conventional therapies.

Proprotein convertase subtilisin/kexin type 9 (PCSK9) is a key regulator of low-density lipoprotein receptor (LDLR) degradation, which leads to increased levels of low-density lipoprotein cholesterol (LDL-C) and a heightened risk of hypercholesterolemia. Existing therapeutic strategies—such as monoclonal antibodies and gene-editing approaches—are limited by high production costs, potential safety concerns, and limited effectiveness in targeting intracellular PCSK9. Targeted protein degradation (TPD) represents a promising alternative capable of overcoming these limitations. In this study, we utilized computer-aided drug design (CADD) to develop a peptide-based degrader, Cadd4, aimed at selectively reducing PCSK9 levels. Through molecular docking and structural refinement, a peptide sequence with strong binding affinity to PCSK9 was identified. Functional evaluation was conducted both in vitro using human LX-2 cells and in vivo in high-fat diet (HFD)-induced hypercholesterolemic mouse models. Cellular uptake, PCSK9 degradation, and LDLR restoration were assessed via confocal microscopy, western blotting, and biochemical analysis. Safety was evaluated by monitoring liver enzyme levels and general physiological parameters. Cadd4 exhibited efficient intracellular delivery and selective PCSK9 degradation in LX-2 cell. In HFD-fed mice, hepatic PCSK9 expression was reduced by 38%, accompanied by a marked increase in LDLR levels. These changes translated into a 25% reduction in total plasma cholesterol and a 29% decrease in LDL-C concentrations. Biodistribution studies indicated liver-specific accumulation of Cadd4, consistent with its intended site of action. Experiments using human liver tissue further confirmed its ability to decrease PCSK9 and enhance LDLR levels. No signs of liver toxicity or adverse systemic effects were observed throughout the study. This work introduces Cadd4 as a CADD-designed peptide-based degrader capable of lowering plasma cholesterol by promoting LDLR recovery via PCSK9 degradation. Its intracellular mechanism of action addresses key gaps in current PCSK9-targeted therapies and supports the broader utility of TPD platforms in managing hypercholesterolemia. These findings offer a foundation for developing cost-effective, scalable therapies within the context of precision cardiovascular medicine.