Zhejiang da xue xue bao. Yi xue ban = Journal of Zhejiang University. Medical sciences最新文献

Primary cilia are microtubule-based organelles that project from the cell surface. They are present in cells from single-celled eukaryotes to vertebrates, including humans. Recent studies have found that primary cilia are also widely distributed in multiple organs and tissues of the reproductive system, where they influence reproductive function by directly participating in or indirectly regulating related signaling pathways, thereby affecting fertility. Primary cilia participate in the regulation of oocyte meiosis and development. They also influence sperm maturation by regulating the homeostatic microenvironment required for spermiogenesis. By mediating Hedgehog (Hh) and Wnt signaling pathways, primary cilia regulate endometrial receptivity and decidual response, thereby influencing the embryo implantation rate. Furthermore, primary cilia control migration, invasion, differentiation, and vascular remodeling of human chorionic villi mesenchymal stromal cells and trophoblasts. Structural or functional impairment of primary cilia may disrupt placental vascular remodeling, leading to placental hypoplasia, potentially through the downregulation of downstream target genes of the Hh signaling pathway. Moreover, primary cilia may be involved in ovarian aging, ovulation, and endocrine function. This article reviews the research progress on the relationship between primary cilia and fertility, explores the potential mechanisms underlying the roles of primary cilia in gamete development, endometrial receptivity, decidualization, placental development, and ovarian reproductive endocrine function, aiming to provide new insights for fertility preservation and the prevention and treatment of human reproductive disorders.

Objectives: To investigate the role of PTEN-induced putative kinase 1 (PINK1) in regulating the viability, migration, and apoptosis of colorectal cancer (CRC) cells, and to explore its potential epigenetic mechanisms.

Methods: PINK1 was overex-pressed or knocked down in HCT116 and DLD1 CRC cell lines using lentiviral vectors, with efficiency verified by qRT-PCR and Western blotting. Cell proliferation was assessed using CCK-8 and colony formation. Cell migration was detected using wound healing and Transwell assays. Apoptosis was assessed using Hoechst 33258 staining. Protein levels of apoptosis-related and histone modification-related markers were analyzed by Western blotting. Genome-wide chromatin accessibility was profiled using assay for transposase-accessible chromatin with sequencing (ATAC-seq).

Results: PINK1 expression was significantly downregulated at both mRNA and protein levels in CRC tissues com-pared to normal tissues. PINK1 overexpression inhibited cell proliferation, colony formation, and migration in HCT116 and DLD1 cells (all P<0.05), whereas PINK1 knockdown promoted these malignant phenotypes (all P<0.05). PINK1 overexpression induced apoptosis, associated with decreased levels of anti-apoptotic proteins (MCL-1, BCL-2, BCL-XL) and increased pro-apoptotic BAX (all P<0.05), without altering p53 expression. Mechanistically, PINK1 overexpression reduced histone H3 lysine 9 trimethylation (H3K9me3) and histone H3 lysine 27 trimethylation (H3K27me3), and increased histone H3 lysine 9 acetylation (H3K9ac) and histone H3 lysine 27 acetylation (H3K27ac). It also downregulated key histone-modifying enzymes, including enhancer of Zeste homolog (EZH)2, EZH1, SUZ12, and histone deacetylase 3 (HDAC3) (all P<0.01). ATAC-seq revealed that PINK1 overexpression increased chromatin accessibility, particularly around transcription start sites.

Conclusions: PINK1 acts as a tumor suppressor in colorectal cancer by inhibiting proliferation and migration, promoting apoptosis, and remodeling the epigenetic landscape through altering histone modifications and enhancing chromatin accessibility.

Extracellular vesicles (EVs) are membrane-bound vesicles secreted by cells, including exosomes, microvesicles, and apoptotic bodies, which play critical roles in intercellular communication, material transport, and signal transduction. In recent years, increasing evidence has highlighted the essential function of EVs in early embryo development. By carrying bioactive molecules such as proteins, nucleic acids (e.g., mRNA and miRNA), and lipids, EVs regulate embryonic gene expression, cell proliferation, differ-entiation, and the microenvironment. Studies have shown that EVs derived from various segments of the female reproductive tract can enhance embryonic developmental potential, improve embryo quality, and facilitate implantation. Additionally, EVs secreted by embryos themselves participate in intercellular communication and play pivotal roles during embryogenesis. This review summarizes recent advances in understanding the functions of EVs in early embryo development, discusses their roles in mediating cell-to-cell com-munication and regulating gene expression, and explores the potential applications in reproductive medicine and clinical practice, offering new perspectives for optimizing assisted reproductive technologies.

With the increasing survival rates of cancer patients, the demand for fertility preservation in women has become increasingly prominent. Ovarian tissue cryopreservation and transplantation (OTCT) is an emerging fertility preservation technique that offers a unique advantage over embryo or oocyte cryopreservation, as it does not require ovarian stimulation. This makes it particularly suitable for prepubertal girls requiring urgent gonadotoxic therapy and reproductive-age women who cannot delay cancer treatment. Clinical evidence confirms that OTCT can effectively restore female fertility-especially the potential for natural conception-and restore ovarian endocrine function. The OTCT process involves key steps such as patient evaluation, tissue processing, cryopreservation, and transplantation. The patient's age at cryopreservation, ovarian reserve status, and prior exposure to gonadotoxic therapy significantly influence the outcomes of fertility preserva-tion. Optimal tissue preparation and the choice of cryopreservation method are critical for preserving ovarian tissue viability. During processing, the size of ovarian tissue fragments must be carefully controlled to balance freezing efficiency and post-transplantation viability, with adjustments based on individual patient factors. Slow freezing remains the mainstream clinical method, while vitrification is still considered experimental, with its efficacy and safety under ongoing investigation. The number, size, and transplantation site of ovarian tissue grafts impact their biological activity and functional outcomes. Both orthotopic and heterotopic transplantation can restore endocrine function, but orthotopic sites are superior for restoring fertility. A major safety concern in OTCT is the potential risk of reintroducing malignant or premalignant cells upon reimplantation. Innovative techniques such as in vitro maturation of oocytes and artificial ovaries are being explored to mitigate this risk. This review summarizes recent clinical advances in OTCT, with a focus on its indications, efficacy, implementation strategies, and safety profile, aiming to provide a reference for further research and clinical practice in this field.

Objectives: Ginkgolide B (GB), a natural bioactive compound, acts on multiple molecular targets; however, its potential multi-target-based therapeutic indications remain unexplored. This study used a network medicine framework to systematically predict novel therapeutic indications for GB, aiming to provide evidence to support its clinical repositioning.

Methods: GB targets were comprehensively identified by integrating data from TTD, CTD, and BindingDB databases, followed by literature-based validation. Disease-gene associations were compiled by integrating GWAS and OMIM datasets. The topological proximity between GB targets and disease modules within the human protein-protein interaction network was quantified using network proximity metrics, with statistical significance evaluated via Z-score and permutation testing. Candidate diseases were classified according to MeSH terms, visualized through network mapping, and further characterized by gene set similarity based on the Jaccard index. Experiments in RAW 264.7 macrophages were conducted to validate the inhibitory effect of GB on pro-inflammatory cytokines in cells.

Results: Twelve experimentally validated GB targets were identified, and a disease-gene set encompassing 680 diseases was established. Network proximity analysis revealed significant associations between GB and 22 diseases (Z<0, P<0.05), of which 11 have prior literature support and 11 represent novel predictions. Disease classification indicated that the predicted indications of GB were primarily enriched in immune and inflammatory disease categories, with notable examples including juvenile idiopathic arthritis (Z=－3.10) and Crohn's disease (Z=－3.05). In vitro experiments with macrophages demonstrated that GB suppresses the production of pro-inflammatory cytokines, thereby supporting its potential as a therapeutic agent for immune-mediated inflammatory diseases. Network analysis suggested that GB may exert pleiotropic regulatory effects across multiple diseases via key targets such as MAPK1. The strongest gene set similarity was observed between juvenile idiopathic arthritis and ankylosing spondylitis (Jaccard index=0.60).

Conclusions: This study demonstrates the utility of a network medicine approach in systematically predicting and partially validating novel indications for GB, particularly within immune-inflammatory disorders, including type 1 diabetes, inflammatory bowel disease, and rheumatoid arthritis. These findings provide a theoretical framework for the clinical repositioning of GB and highlight the potential utility of the network-based strategies in the systematic exploration of natural products.

Objectives: To characterize the biological properties of double-negative T (DNT) cells isolated from leukoreduction filter residues.

Methods: Leukoreduction filters containing residues from 400 mL whole blood units (n=6) were collected from a blood center. Filters were back-flushed with normal saline, and the eluate was concentrated to obtain leukoreduction filter residues. Leukocytes in the residues were counted by dual-fluorescence staining. DNT cells were then isolated from the residues using antibody-mediated adsorption and density gradient centrifugation. Both cryopreserved and fresh unstimulated DNT cells derived from the residues were subjected to in vitro culture. Following culture, cells were assessed for expansion fold, viability, immunophenotype, differentiation status, and cytotoxicity against target cells using dual-fluorescence staining and flow cytometry, with comparisons made to DNT cells derived from whole blood.

Results: The leukocyte recovery rate achieved through reverse flushing of the leukocyte reduction filter was (41.9±14.7)%. Compared to whole blood, the DNT cell starting material obtained from filter residues showed no significant difference in total T-cell content (P>0.05). However, the viability and purity of the resulting DNT cell starting materials were significantly lower (both P<0.05). After 17 days of culture, DNT cells from filter residues and whole blood showed no significant differences in expansion fold, immunophenotype, differentiation status, or cytotoxicity toward target cells (all P>0.05). However, the viability of DNT cells from residues was significantly lower than that of whole blood-derived DNT cells [(86.0±4.2)% vs. (92.2±1.2)%, P<0.05]. After thawing (post 3 or 15 days of cryopreservation) and 17 days of culture, DNT cell starting materials from residues showed comparable immunophenotype, expansion fold, and differentiation status to their non-cryopreserved counterparts from the same source (all P>0.05). However, the viability of DNT cells cryopreserved for 3 days [92.4% (91.8%, 92.8%)] and the cytotoxicity against target cells of those cryopreserved for 15 days [91.3% (89.4%, 95.1%)] were significantly higher than those of non-cryopreserved DNT cells [87.8% (82.0%, 89.0%) and 70.9% (67.3%, 80.2%), respectively] (P<0.05).

Conclusions: DNT cells derived from leukoreduction filter residues exhibited highly comparable characteristics to those from whole blood in terms of expansion, purity, differentiation, and biological potency. Furthermore, their biological activity post-cryopreservation and revival remained largely similar to non-cryopreserved cells. These findings suggest that leukoreduction filter residues represent a promising alternative source of starting material for manufacturing off-the-shelf, allogeneic DNT cell therapeutics.