Systemic delivery of messenger RNA (mRNA) to non‐hepatocytes using lipid nanoparticles (LNPs) remains challenging. Inspired by anion coordination chemistry, here we report the rational chemical design of thiourea‐functionalized ionizable lipids (TUILs) for delivering mRNA potently and specifically to the secondary lymphoid organs (SLOs). The leading TUIL, namely 4A3‐LNSC8, features an impressive thiourea‐based linker capable of binding with various halide anions (F − , Cl − , I − ) through hydrogen‐bonding interaction, enabling precise regulation of LNP organotropism in vivo. When administered systemically, the representative Cl‐4A3‐LNSC8 LNPs exclusively redirected mRNA delivery from the liver to SLOs, yielding a 65‐fold and 29‐fold increase in splenic mRNA expression compared to DLin‐MC3‐DMA and SM102 LNPs with the addition of anionic lipid (18PA). Notably, upon intravenous injection of 0.2 mg kg −1 Cre mRNA, Cl‐4A3‐LNSC8 LNPs demonstrated strong tropism for splenic macrophages with high gene editing efficiency up to 65.7%, outperforming the current state‐of‐the‐art spleen‐targeted LNPs. Moreover, by leveraging iodine's CT contrast properties, I‐4A3‐LNSC8 LNPs mediated efficient theranostic mRNA delivery to lymph nodes, allowing early detection of lymphatic metastasis via dual‐modal CT and bioluminescence imaging. This work provides new insights into the development of cell‐specific ionizable lipids, advancing future applications of macrophage‐targeted therapies.

KRAB zinc finger proteins (KRAB-ZFPs) are the most abundant transcriptional regulators and epigenetic repressors in mammals and contribute to the silencing of transposable elements (TEs) during embryonic development. However, the early effectors of the KRAB-ZFP family during embryonic development and cell state transition remain largely unexplored. Here, we identify that zinc finger protein 560 (ZFP560) is involved in the regulation of early embryonic development and the transition from totipotency in mice. ZFP560 safeguards heterochromatin structure by recruiting KAP1 to facilitate the exit of totipotency. The deficiency of ZFP560 disrupted heterochromatin formation, significantly halting the transition from totipotency to pluripotency. Similarly, the overexpression of ZFP560 promotes exit from the MERVL-positive state and suppresses expression of 2-cell (2C) transcription factors by recruiting KAP1 via its KRAB domain, thereby reducing chromatin accessibility. Taken together, these findings reveal that ZFP560 is a highly expressed 2C-specific KRAB-ZFP transcription factor and a mediator that facilitates the exit from totipotency, providing insights into the epigenetic regulation of early embryonic development.

Mitochondrial oxidative stress plays a critical role in cancer development and progression. However, there is limited research on the relationship between mitochondrial oxidative stress and liver hepatocellular carcinoma (LIHC). Mitochondrial oxidative stress-related genes were collected from Genecards Portal. Prognosis-linked genes (PLGs) were identified by univariate Cox regression analysis. A risk model was constructed based on the PLGs using least absolute shrinkage and selection operator (LASSO) analysis. Receiver operating characteristic (ROC) curves were used to determine the predictive ability of the model. The expression levels of the prognostic genes were verified in the cell lines. Cell proliferation, apoptosis, and invasion assays were conducted to investigate the functional role of the target gene. We constructed a novel risk model based on 9 prognostic genes (CYP2C19, CASQ2, LPL, TXNRD1, CACNA1S, SLC6A3, OXTR, BIRC5, and MMP1). Survival analysis showed that patients with a low-risk score had a much better overall survival (OS). Prognostic risk score was found to be an independent predictor of prognosis. Patients in the high-risk group had a less favorable tumor microenvironment characterized by a lower degree of immune cell infiltration. Among the nine prognostic genes, MMP1, identified as the most promising candidate, demonstrated the capacity to enhance tumor cell proliferation and invasion. Our investigation reveals the oncogenic role of mitochondrial oxidative stress in LIHC. For the first time, we established a risk prediction model for mitochondrial oxidative stress in patients with LIHC. MMP1 has the potential to function as a promising biomarker in LIHC.

Chemotherapy and immunotherapy have shown no significant outcome for unresectable pancreatic ductal adenocarcinoma (PDAC). Multi-drug combination therapy has become a consensus in clinical trials to explore how to arouse anti-tumor immunity and meanwhile overcome the poorly tumoricidal effect and the stroma barrier that greatly hinders drug penetration. To address this challenge, a comprehensive strategy is proposed to fully utilize both the ferroptotic vulnerability of PDAC to potently irritate anti-tumor immunity and the desmoplasia-associated focal adhesion kinase (FAK) to wholly improve the immunosuppressive microenvironment via sustained release of drugs in an injectable hydrogel for increasing drug penetration in tumor location and averting systematic toxicity. The injectable hydrogel ED-M@CS/MC is hybridized with micelles loaded with erastin that exclusively induces ferroptosis and a FAK inhibitor defactinib for inhibiting stroma formation, and achieves sustained release of the drugs for up to 12 days. With only a single intratumoral injection, the combination treatment with erastin and defactinib produces further anti-tumor performance both in xenograft and KrasG12D-engineered primary PDAC mice and synergistically promotes the infiltration of CD8+ cytotoxic T cells and the reduction of type II macrophages. The findings may provide a novel promising strategy for the clinical treatment of PDAC.

Mammalian SWI/SNF (mSWI/SNF) ATP-dependent chromatin remodeling complexes play critical roles in regulating gene expression and DNA accessibility, and more than 20% of cancers have mutations in genes encoding chromatin remodeling complexes. The mSWI/SNF family comprises three distinct classes: canonical BAF (cBAF), PBAF, and non-canonical BAF (ncBAF). While the structures of cBAF and PBAF have been resolved by using cryo-electron microscopy (cryo-EM), the modular organization and assembly mechanism of ncBAF remain poorly understood. In this study, we first mapped the binding fragment of SMARCC1/SMARCD1 complex, then found that GLTSCR1(1041-1204) could form a stable complex with SMARCC1(447-966)/SMARCD1(129-515). Next, we purified the SMARCC1(447-966)/SMARCD1(117-515)/GLTSCR1(1041-1204)/BRD9(266-510) tetrameric complex. Finally, we assembled a stable and uniform SMARCC1(447-966)/SMARCD1(117-515)/GLTSCR1(1041-1204)/BRD9(266-510)/SMARCA4(289-464) quinary complex in vitro, which is ncBAF core module. These findings provide insight into the assembly mode of ncBAF complex, and lay the foundations for further solving its structure in the future.

Systemic mRNA delivery to specific cell types remains a great challenge. We herein report a new class of crown-like biodegradable ionizable lipids (CBILs) for predictable lung-selective mRNA delivery by leveraging the metal coordination chemistry. Each CBIL contains an impressive crown-like amino core that coordinates with various metal ions such as Zn2+ and further regulates the in vivo organ-targeting behavior of lipid nanoparticles (LNPs). The representative CBIL (Zn-9C-SCC-10)-formulated LNPs could exclusively deliver mRNA to the lung after systemic administration. Notably, following intravenous administration of 0.2 mg kg-1 Cre mRNA, Zn-9C-SCC-10 LNPs enabled the highly efficient gene editing of all lung epithelial and endothelial cells up to 43 and 61%, respectively, outperforming the current state-of-the-art LNPs in lung epithelial cell delivery. Moreover, compared to DLin-MC3-DMA LNPs with the addition of cationic lipid (DOTAP), our approach yielded a 44.6-fold enhancement in pulmonary mRNA expression and significantly improved biosafety in vivo. Taking advantage of paramagnetic gadolinium ion, Gd-12C-SCC-10 LNPs allowed the potent mRNA delivery to cancer cells and successfully illuminated lung tumors by magnetic and bioluminescent dual-mode imaging, facilitating the early discovery and diagnosis of lung cancer. This work will open a new avenue to rationally design predictable LNPs, as well as address the major challenges of mRNA delivery to specific cells in the lung tissues for treating a wide variety of diseases.

OBJECTIVES:This study aims to elucidate the microbial signatures associated with autoimmune diseases, particularly systemic lupus erythematosus (SLE) and inflammatory bowel disease (IBD), compared with colorectal cancer (CRC), to identify unique biomarkers and shared microbial mechanisms that could inform specific treatment protocols. METHODS:We analysed metagenomic datasets from patient cohorts with six autoimmune conditions-SLE, IBD, multiple sclerosis, myasthenia gravis, Graves' disease and ankylosing spondylitis-contrasting these with CRC metagenomes to delineate disease-specific microbial profiles. The study focused on identifying predictive biomarkers from species profiles and functional genes, integrating protein-protein interaction analyses to explore effector-like proteins and their targets in key signalling pathways. RESULTS:Distinct microbial signatures were identified across autoimmune disorders, with notable overlaps between SLE and IBD, suggesting shared microbial underpinnings. Significant predictive biomarkers highlighted the diverse microbial influences across these conditions. Protein-protein interaction analyses revealed interactions targeting glucocorticoid signalling, antigen presentation and interleukin-12 signalling pathways, offering insights into possible common disease mechanisms. Experimental validation confirmed interactions between the host protein glucocorticoid receptor (NR3C1) and specific gut bacteria-derived proteins, which may have therapeutic implications for inflammatory disorders like SLE and IBD. CONCLUSIONS:Our findings underscore the gut microbiome's critical role in autoimmune diseases, offering insights into shared and distinct microbial signatures. The study highlights the potential importance of microbial biomarkers in understanding disease mechanisms and guiding treatment strategies, paving the way for novel therapeutic approaches based on microbial profiles. TRIAL REGISTRATION NUMBER:NCT02394964.

A bioreducible Zn (II)-adenine multifunctional module (BS) and Tet1 peptide were used to modify lowmolecular-weight PEI3.5k (polyethyleneimine with molecular weight of 3.5 kDa)into a siRNA vector Zn-PB-T with high transfection efficiency in neurons. A GSH-responsive breakable disulfide spacer was introduced into BS to realize the controlled release of siRNA from the polyplexes in cytoplasm. Zn-PB showed >90% transfection rates in multiple cell lines (3 T3, HK-2, HepG2, 293 T, HeLa, PANC-1),and 1.8-folds higher EGFP knockdown rates than commercial Lipo2k in normal cell line 293 T and cancer cell line HepG2. And Zn-PB-T1 showed 4.7-4.9- and 8.0-8.1-folds higher transfection efficiency comparing to commercial Lipo(2k )and PEI25k (polyethyleneimine with molecular weight of 25 kDa) in PC12 cells respectively, 2.1-fold EGFP gene silencing efficiency (96.6% EGFP knockdown rates) superior to commercial Lipo2k in neurons. In Parkinson's model, Zn-PBT1/SNCA-siRNA can effectively protect neurons against MPP+-induced cell death and apoptosis, increasing the cell survival rate to 84.6% and reducing the cell apoptosis rate to 10.8%. This work demonstrated the promising application prospects of the resulting efficient siRNA carriers in siRNA-mediated gene therapy of Parkinson's disease.

Gene delivery to macrophages holds great promise for cancer immunotherapy. However, traditional gene delivery methods exhibit low transfection efficiency in macrophages. The star-shaped topological structure of polymers is known to encapsulate genes inside their cores, thereby facilitating sustained release of the genetic material. Herein, combining the structural advantages of star polymers and the transfection advantages of poly (β-amino ester)s (PAEs), we developed a novel linear oligomer grafting-onto strategy to synthesize a library of multi-terminal star structured PAEs (SPAEs), and evaluated their gene delivery efficiency in various tissue cells. The transfection with human hepatocellular carcinoma cells (HepG2, HCC-LM3 cells and MHCC-97H cells), rat normal liver cells (BRL-3 A cells), human ovarian cancer cells (A2780 cells), African green monkey kidney cells (Vero cells), human cervical cancer cells (HeLa cells), human chondrosarcoma cells (SW1353 cells), and difficult-to-transfect human epidermal keratinocytes (HaCaT cells) and normal human fibroblast cells (NHF cells) showed that SPAEs exhibited superior transfection profile. The GFP transfection efficiency of top-performing SPAEs in HeLa cells (96.1%) was 2.1-fold, and 3.2-fold higher compared to jetPEI and Lipo3000, respectively, indicating that the star-shaped topological structure can significantly enhance the transfection efficiency of PAEs. More importantly, the top-performing SPAEs could efficiently deliver Nod2 DNA to difficult-to-transfect RAW264.7 macrophages, with a high transfection efficiency of 33.9%, which could promote macrophage M1 polarization and enhanced CD8+ T cell response in co-incubation experiments. This work advances gene therapy by targeting difficult-to-transfect macrophages and remodeling the tumor immune microenvironment.

A highly efficient siRNA vector (Zn-PQD) capable of selectively silencing genes in cancer cells was obtained by using ROS-cleavable DED to crosslink low molecular weight (LMW) polyethylene imine (PEI) modified by selffluorescent metal coordinatied multifunctional module Zn-QS. Under the combined action of DED cross-linking and Zn-QS modification, Zn-PQD performs well in the siRNA delivery process in cancer cells, including siRNA condensation, cell uptake, endosome escape, and siRNA release. Zn-PQD exhibited higher transfection efficiency than commercial PEI25k and Lipo2k in multiple cancer cell lines including HepG2, HeLa, 4 T1, H520 and PANC-1, as well as cancer treatment-related stem cell rADSC. Ultimately, Zn-PQD can achieve extremely high and selective gene silencing effects in cancer cells (with a gene silencing rate of 98.3% in HepG2). This work is expected to provide an efficient and safe siRNA carrier for the future tumor siRNA therapy and its study of fluorescence mediated mechanism.

Immune checkpoint inhibitor-based cancer immunotherapy has shown promise as a potential treatment in the clinic. It has been reported that anti-PD-L1 combined with cisplatin treatment can improve the antitumor effect. However, the therapeutic outcome is limited due to the abundance of tumor stroma in pancreatic cancer (PC), which prevented the penetration of cisplatin and anti-PD-L1 into tumor regions, thus impeding the effectiveness in the treatment of PC. In this study, a nanocarrier-mediated codelivery system of hyaluronidase and cisplatin was constructed, which can degrade the stroma and promote cisplatin and anti-PD-L1 to penetrate the tumor stroma into the deep tumor, so as to suppress PC effectively. When combined the cisplatin nanocarrier system BPEI-SS-Pt/HAase@CaP (BSP/H@CaP) with an immune checkpoint inhibitor to overcome the poor therapeutic outcome of PC, the results indicated that the therapeutic effect of BSP/H@CaP combined with anti-PD-L1 was better than that of BSP/H@CaP and single anti-PD-L1 group. Because the stroma is degrading, a higher amount of BPEI-SS-Pt and anti-PD-L1 can enter the tumor stroma and reach the inner depths of the tumor for immune stimulation, leading to a synergistically augmented chemotherapy and immunotherapy for PC. The above combination therapy is useful for clinical translation to overcome the treatment resistance of matrix-rich PC.

大型仪器对于创新人才的培养具有不可替代的积极作用.如何使本科教学大型仪器在人才培养中发挥更大的作用,需要依据仪器自身特点,采取合适的教学方式,方能达到事半功倍的效果.以生物技术专业的"基于高效液相色谱仪的开放性实验项目"为例,探讨以开放探究性实验项目为载体的大型仪器实验教学模式.从项目实施的基础、实施过程、结题和答辩等方面介绍了该模式的全过程.实践表明,该模式不仅能够使学生熟练掌握大型仪器的使用,还能培养学生的科研能力,值得推广.

Highly branched poly(β-amino ester)s(HPAEs)have shown their great promise in gene delivery.How-ever,their broad molecular weight distribution(MWD)poses an additional challenge to the mechanistic understanding of the influence of molecular weight(MW)on their gene transfection activity.Using a stepwise precipitation strategy,HPAEs were fractionated.It is shown that MW has a significant effect on the transfection activity and cytotoxicity of HPAEs.The intermediate MW mediates higher transfection efficiency while maintaining high cell viability.Mechanistic studies show that the intermediate MW con-fers stronger DNA binding affinity to HPAEs,leading to the formulation of polyplexes with a relatively smaller size and more positive zeta potential.This study not only suggests a simple strategy to fraction-ate HPAEs with narrow MWD but also provides new insights into understanding the structure-property relationship,which would facilitate the clinical translation of HPAEs in gene therapy.

A novel quinoline‐based thioketal‐containing zinc (II) coordinative module (Zn‐QS) is designed to transform common low‐molecular‐weight (LMW) polyethyleneimine (PEI) into a high‐performance siRNA carrier (Zn‐PQ) with self‐fluorescence and reactive oxygen species (ROS)‐responsiveness. In the multifunctional module, a quinoline derivative, frequently serving as pharmacophore, is utilized as the highly biocompatible ligand, and the Zn (II) coordinative quinoline‐based ligand also provides the robust self‐fluorescence character. And the ROS‐responsive thioketal spacer is introduced to trigger the siRNA controlled release into cytoplasm. Consequently, by the synergistic effects between Zn‐QS and LMW PEI on improving several crucial transfection‐efficiency‐related steps, the representative Zn‐PQ4 exhibits very high transfection efficiency of 1.8 and 2.5‐folds commercial Lipo 2k and PEI 25k in stem cell rADSC, respectively, >90% of transfection rates in a variety of conventional cell lines (3T3, HK‐2, HepG2, 293T, HeLa, PANC‐1), and 98.4% and 99.9% EGFP knockdown rates in 293T and HepG2 cells, respectively. Furthermore, Zn‐PQ4/siRNA polyplexes can circumvent various obstacles existing in systemic application to accumulate in heart, liver, and kidney and greatly silence the DNA expression in vivo. This work demonstrates the promising application prospects of the resulting efficient siRNA carriers in siRNA‐mediated gene therapy and fluorescence‐guided research.

Extensive efforts have been dedicated to enhancing the expression of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) in cancer cells for the development of effective cancer treatments. However, highly safe and efficient delivery of TRAIL gene remains a significant challenge, especially using cationic polymers. Here, a series of highly branched-linear poly(β-amino ester)s (H-LPAEs) are developed through a unique oligomer branching strategy. H-LPAEs exhibit a more uniform distribution of linear segments and branching units, leading to excellent DNA condensation and favorable physicochemical properties of H-LPAE/DNA polyplexes. In SW1353 and BMSC cells, the optimized H-LPAEs, H-LPAE B4−S5−TMPTA , achieves superior gene transfection efficiency of 58.0% and 33.4%, which were 2.5-fold and 2.0-fold higher than that of the leading commercial gene transfection reagent, Lipofectamine 3000. Excitingly, H-LPAE B4−S5−TMPTA mediated 56.7% and 28.1% cell apoptosis in HepG2 cells and HeLa cells highlighting its potential application in cancer gene therapy. In addition, locally administered H-LPAE B4−S5−TMPTA delivered TRAIL DNA to HepG2 xenograft tumors and inhibited tumor growth in vivo. This study not only proposes a novel strategy for synthesizing poly(β-amino ester)s with a unique branched-linear topology but also identifies a promising candidate for highly efficient TRAIL gene transfection.

Oncoprotein SS18-SSX is a hallmark of synovial sarcomas. However, as a part of the SS18-SSX fusion protein, SS18's function remains unclear. Here, we depict the structures of both human SS18/BRG1 and yeast SNF11/SNF2 subcomplexes. Both subcomplexes assemble into heterodimers that share a similar conformation, suggesting that SNF11 might be a homologue of SS18 in chromatin remodeling complexes. Importantly, our study shows that the self-association of the intrinsically disordered region, QPGY domain, leads to liquid-liquid phase separation (LLPS) of SS18 or SS18-SSX and the subsequent recruitment of BRG1 into phase-separated condensates. Moreover, our results show that the tyrosine residues in the QPGY domain play a decisive role in the LLPS of SS18 or SS18-SSX. Perturbations of either SS18-SSX LLPS or SS18-SSX's binding to BRG1 impair NIH3T3 cell transformation by SS18-SSX. Our data demonstrate that both LLPS and assembling into chromatin remodelers contribute to the oncogenic activity of SS18-SSX in synovial sarcomas.

离心是生物学及相关学科教学和科研领域中最常用的实验技术之一.为提高实验操作效率和实验成功率,南开大学生物实验教学中心自行研制了一种"离心样品预处理装置",能够"一站式"完成离心样品预处理过程,主要包括样品混匀、定量取样与快速配平步骤,从而有效提高离心样品预处理操作速度,提高取样准确性与离心安全性.同时,本自制仪器具有性能稳定、操作简便、易于维护、节省空间的优点,并取得了良好的教学实践效果,对于实验教学改革和学生综合能力培养具有积极意义.

如何在实验教学条件持续发展的情况下,实现高等教育工作重心的转移,由规模扩张向内涵发展转变,是目前高校实验教学普遍面临的问题.实验室通过梳理、归纳、剖析制约实验教学质量的因素,提出通过优化实验教学体系和课程内容、以信息化改进教学方法和教学方式、以教改提升教学质量、提高实验室开放质量等举措多角度加强实验教学内涵建设.实践表明,这些举措有效提升了实验教学质量,有利于新时期创新人才培养.

Enzyme-regulated in situ self-assembly of peptides represents one versatile strategy in the creation of theranostic agents, which, however, is limited by the strong dependence on enzyme overexpression. Herein, we reported the self-amplifying assembly of peptides precisely in macrophages associated with enzyme expression for improving the anti-inflammatory efficacy of conventional drugs. The self-amplifying assembling system was created via coassembling an enzyme-responsive peptide with its derivative functionalized with a protein ligand. Reduction of the peptides by the enzyme NAD(P)H quinone dehydrogenase 1 (NQO1) led to the formation of nanofibers with high affinity to the protein, thereby facilitating NQO1 expression. The improved NQO1 level conversely promoted the assembly of the peptides into nanofibers, thus establishing an amplifying relationship between the peptide assembly and the NQO1 expression in macrophages. Utilization of the amplifying assembling system as vehicles for drug dexamethasone allowed for its passive targeting delivery to acute injured lungs. Both in vitro and in vivo studies confirmed the capability of the self-amplifying assembling system to enhance the anti-inflammatory efficacy of dexamethasone via simultaneous alleviation of the reactive oxygen species side effect and downregulation of proinflammatory cytokines. Our findings demonstrate the manipulation of the assembly of peptides in living cells with a regular enzyme level via a self-amplification process, thus providing a unique strategy for the creation of supramolecular theranostic agents in living cells.

Background Host-microbe interactions are crucial for normal physiological and immune system development and are implicated in a variety of diseases, including inflammatory bowel disease (IBD), colorectal cancer (CRC), obesity, and type 2 diabetes (T2D). Despite large-scale case-control studies aimed at identifying microbial taxa or genes involved in pathogeneses, the mechanisms linking them to disease have thus far remained elusive. Results To identify potential pathways through which human-associated bacteria impact host health, we leverage publicly-available interspecies protein-protein interaction (PPI) data to find clusters of microbiome-derived proteins with high sequence identity to known human-protein interactors. We observe differential targeting of putative human-interacting bacterial genes in nine independent metagenomic studies, finding evidence that the microbiome broadly targets human proteins involved in immune, oncogenic, apoptotic, and endocrine signaling pathways in relation to IBD, CRC, obesity, and T2D diagnoses. Conclusions This host-centric analysis provides a mechanistic hypothesis-generating platform and extensively adds human functional annotation to commensal bacterial proteins.