Cell Function Research Articles

Human umbilical cord mesenchymal stem cells small extracellular vesicles-derived miR-370-3p inhibits cervical precancerous lesions by targeting DHCR24

Abstract Background Cervical cancer is often caused by persistent high-risk human papillomavirus (HPV) infection, causing precancerous lesions. Human umbilical cord mesenchymal stem cells-derived small extracellular vesicles (hucMSC-sEV) exhibit diverse effects on tumors. This study investigates hucMSC-sEV, the impact and mechanisms on HPV-positive cervical precancerous lesion cells to provide new treatment insights. Materials and Methods We previously obtained hucMSC and hucMSC-sEV. In vitro experiments evaluated hucMSC-sEV effects on the proliferation and migration of S12 cells (derived from cervical precancerous lesions). Bioinformatics identified key microRNA components, and their impact on S12 cell proliferation and migration was investigated. The target gene of the microRNA component was predicted and confirmed via bioinformatics and dual-luciferase reporter assays. Lentiviral systems overexpressed target gene in S12 cells to examine the effects on microRNA impacts. SH-42 inhibitor was used to investigate target gene treatment potential. Immunohistochemistry assessed target gene expression in cervical precancerous lesions tissue. Results hucMSC-sEV significantly inhibited S12 cell proliferation and migration. Bioinformatics identified miR-370-3p as an effective cargo, which also suppressed S12 cell proliferation and migration. miR-370-3p was confirmed targeting DHCR24 (24-Dehydrocholesterol Reductase). DHCR24 overexpression reversed miR-370-3p’s inhibitory effects, while SH-42 counteracted DHCR24 overexpression’s promoting effects. Clinical specimen analysis supported these findings, demonstrating a positive correlation between DHCR24 protein expression and cervical precancerous lesions’ progression. Conclusions hucMSC-sEV inhibits S12 cell proliferation and migration, mediated by miR-370-3p targeting DHCR24 to regulate cellular cholesterol content. DHCR24 inhibition reduces the cholesterol level and cell functions, suggesting its potential as a therapeutic target in cervical precancerous lesions.

The role of micronuclei in chromatin elimination

Micronuclei are the extra-nuclear chromatin compartments separated from the primary nucleus and surrounded by their own nuclear envelope. For a long time it has been thought that micronuclei is the final stage of the pathological process in a cell. They have been used as biomarkers of the influence of genotoxic factors as well as of genome instability in various diseases. Nowadays, it is demonstrated that micronuclei could be involved in the cellular activities, affect the nuclear genome and lead to the changes in cell and tissue physiology. It is known that the formation of micronuclei is one of the steps in selective chromatin elimination in the ontogenesis of plant and animal species. The regions to be marked and eliminated from cell nucleus are recognized at the level of genome. This process is often accompanied by modifications with the heterochromatin formation, changes in the chromosome condensation and in the position of chromosomes in the nucleus. The processes observed in selective and non-selective chromatin elimination are similar to a great extent. The fact that the role of micronuclei in the cell functioning is not well-known yet, and the composition of the micronuclei and the ways of chromatin elimination could influence their role in the development of the pathogenesis, emphasizes the importance of additional studies for a more profound investigation of this phenomenon.

Etoposide targets 2A protease to inhibit enterovirus 71 replication

ABSTRACT Enterovirus 71 (EV71) is a major pathogen that causes hand, foot, and mouth disease (HFMD) in infants and children. Notably, no clinically approved drugs specifically target EV71. The EV71 2A protease (2A pro ), a cysteine protease produced by the virus, is essential for the virus’ replication and has a significant impact on the functioning of host cells. Thus, it presents a valuable target for the discovery of antiviral medications. In this study, based on the monomers and their derivatives in the Library of Traditional Chinese Medicine (TCM), we performed virtual screening and biological experiments. We identified a derivative of a traditional herbal monomer, Etoposide, commonly isolated from the roots and rhizomes of Podophyllum spp. Etoposide inhibited replication of EV71 A, B, C, and CVA16 viruses in a concentration-dependent manner in a variety of cell lines with minimal cytotoxicity. Furthermore, both molecular dynamics simulations and site-directed mutagenesis assays revealed that Etoposide inhibited the activity of the EV71 2A protease by mainly binding to two residues, Y89 and P107. The findings indicate that Etoposide serves as a promising inhibitor of the EV71 2A pro , demonstrating strong antiviral properties and positioning itself as a formidable candidate for clinical trials against EV71. IMPORTANCE We first used a drug screening approach focused on monomeric compounds and their derivatives from traditional Chinese medicine to identify an EV71 2A pro inhibitor—Etoposide. We then performed biological experiments to validate that Etoposide suppresses the replication of the EV71 virus in a concentration-dependent manner with minimal cytotoxicity to various cell lines. Remarkably, it shows inhibitory activity against EV71 A, B, C, and CVA16, suggesting that Etoposide may be a potential broad-spectrum inhibitor. We revealed a novel mechanism that Etoposide inhibits EV71 proliferation by targeting 2A pro , and the interactions with Y89 and P107 are of great importance. The findings suggest that Etoposide serves as a promising inhibitor of EV71 2A pro , demonstrating significant antiviral properties. It stands out as a strong candidate for broad-spectrum applications in clinical research.

Proposed Mechanisms of Cell Therapy for Alzheimer’s Disease

Alzheimer’s disease is a progressive neurodegenerative disorder characterized by mitochondria dysfunction, accumulation of beta-amyloid plaques, and hyperphosphorylated tau tangles in the brain leading to memory loss and cognitive deficits. There is currently no cure for this condition, but the potential of stem cells for the therapy of neurodegenerative pathologies is actively being researched. This review discusses preclinical and clinical studies that have used mouse models and human patients to investigate the use of novel types of stem cell treatment approaches. The findings provide valuable insights into the applications of stem cell-based therapies and include the use of neural, glial, mesenchymal, embryonic, and induced pluripotent stem cells. We cover current studies on stem cell replacement therapy where cells can functionally integrate into neural networks, replace damaged neurons, and strengthen impaired synaptic circuits in the brain. We address the paracrine action of stem cells acting via secreted factors to induce neuroregeneration and modify inflammatory responses. We focus on the neuroprotective functions of exosomes as well as their neurogenic and synaptogenic effects. We look into the shuttling of mitochondria through tunneling nanotubes that enables the transfer of healthy mitochondria by restoring the normal functioning of damaged cells, improving their metabolism, and reducing the level of apoptosis.

Abstract B012: Understanding the role of VISTA in regulating T cell function and the therapeutic potential of blocking VISTA in triple negative breast cancer

Abstract Triple-negative breast cancer (TNBC) is the most aggressive and highly metastatic breast cancer subtype with limited effective treatment options. Immunotherapy with immune checkpoint inhibitors (ICIs) targeting checkpoint molecules have been widely used and have shown therapeutic efficacy in treating multiple malignancies in the last decade. In TNBC, ICI targeting PD-1, alone or in combination with chemotherapy has been approved for the treatment of patients with both early and advanced stage of the disease. Although, there has been significant clinical benefit achieved from anti-PD-1 and chemotherapy, treatment efficacy is often limited to a subgroup of patients. The limited response to ICIs has been attributed to the increased infiltration of immunosuppressive cells (including MDSCs, neutrophils and tumor-associated macrophages), more importantly, the upregulation of alternative immune checkpoint receptors/ligands that may further suppress cytotoxic T cells (CD8+ T cells) in the tumor microenvironment (TME). Therefore, it has been suggested that targeting other immune checkpoint molecules may be more effective in bolstering anti-tumor immune response. V- domain Ig suppressor of T-cell activation (VISTA) is one of the immune checkpoint molecules attracting increasing attention in the context of anti-tumor immunity due to its broad expression across various cancer types and increased expression upon development of immune checkpoint resistance. However, the role of VISTA in regulating the anti-tumor immune response and its therapeutic potential in TNBC remains unclear. In our study, we observed that VISTA is expressed in TNBC patients, predominantly in neutrophils and macrophages. In the Py230 murine breast cancer model, VISTA expression increased upon paclitaxel (PTX), anti-PD-L1 and paclitaxel combined with anti-PD-L1 treatment compared to the control group, suggesting its potential role as a resistance mechanism in sustaining or further inhibiting T cells. In the Py230 model, VISTA blockade increased the infiltration and activation of CD8+ T cells, as evidenced by the higher number of CD8+ T cells, increased CD69+CD8+ T cells, and Granzyme B production, indicating that VISTA blockade improved CD8+ T cells mediated anti-tumor immunity. To functionally test the role of VISTA, we stimulated CD8+ T cells and co-cultured them with naïve macrophages or tumor educated macrophages in the presence or absence of VISTA blocking antibody. Tumor educated macrophages showed increased immunosuppressive activity which was abrogated with VISTA blockade. Moreover, we found that the immunosuppressive effect of macrophages on T cell activation needs direct cell-cell interaction between VISTA on macrophages and T cells. In conclusion, this study shows the mechanism of action of VISTA in the regulation of T cells mediated anti-tumor immune response and provides the rationale for targeting VISTA in TNBC. Citation Format: Maidinaimu Abudula, Yuliana Astuti, Meirion Raymant Raymant, Vijay Sharma, Michael Schmid, Ainhoa Mielgo. Understanding the role of VISTA in regulating T cell function and the therapeutic potential of blocking VISTA in triple negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr B012.

Abstract IA018: Determinants of B cell fate and function in cancer

Abstract Effector B cell responses in solid malignancies are associated with favorable response to immunotherapy. B cells can amplify anti-tumor immune responses via antibody production, antigen presentation, and pro-inflammatory cytokine release; yet B cells in cancer patients and tumor-bearing mice often fail to support these functions. We have evaluated the contribution of either systemic inflammatory cues or antigenic quality to B cell differentiation and function in cancer. First, we identify dysregulated transcriptional programs activated in pancreatic cancer-associated B cells that promote differentiation of naïve into immunosuppressive B cells and inhibit differentiation of anti-tumor plasma cells. Second, we find that systemic exposure to STING agonists potentiates expansion of immunosuppressive regulatory human and mouse IL35+ B lymphocytes. Finally, using a system where a B cell neoantigen is either soluble or tethered to the surface of cancer cells, we demonstrate that membrane-tethered antigen is sufficient to elicit activation of anti-tumor immunity and reduce tumor growth in a NK cell-dependent manner. Thus, inflammation and quality of antigen impact basic mechanisms governing B cell differentiation and the resulting immune response to solid malignancy with implications for vaccine engineering. Citation Format: Yuliya Pylayeva-Gupta. Determinants of B cell fate and function in cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr IA018.

Abstract B048: Neutrophil extracellular traps-targeting DNase I enhances the therapeutic efficacy of CAR-T cell adoptive immunotherapy in a syngeneic murine metastasis model

Abstract In recent years, chimeric antigen receptor (CAR) T cell therapy has emerged as a promising approach for treating various malignancies, most notably in hematologic cancers. However, its efficacy in solid tumors is often hindered by an immunosuppressive tumor microenvironment (TME), which, in particular, is frequently characterized by the presence of tumor-associated cell-free DNA (cfDNA), especially in the form of neutrophil extracellular traps (NETs). In adoptive immunotherapies NETs, composed primarily of neutrophil-derived DNA, histones and neutrophil granule enzymes (e.g., MPO, NE), act as physical barriers and induce the secretion of immunosuppressive factors that impair CAR T cell function. This study investigates the potential of co-administration of deoxyribonuclease I (DNase I) with CAR T cells in a syngeneic B16 melanoma murine model of lung metastasis. Bioluminescent imaging of melanoma metastatic processes has shown that a single injection of DNase I (10 mg/kg) suppresses B16-EGFR lung metastasis at early stages in comparison to the vehicle control group but does not improve survival. However, co-administration of DNase I with murine EGFR-CAR T cells significantly suppresses tumor burden, decreases the number of metastatic foci, and substantially prolongs survival compared to the CAR T cell monotherapy group. Degrading of NETs [RB1] by DNase I increases the amount of tumor-infiltrating T and CAR T cells and reduces the immunosuppressive effects of the TME. This is reflected in the CD8 population of tumor-infiltrating CAR T cells from the DNase I treated group, which have lower expression of PD-1 and TIM-3 exhaustion markers. This research highlights the critical role of the NETs in modulating CAR T cell efficacy and provides a compelling rationale for incorporating DNase I as an adjunctive treatment to improve therapeutic responses in patients undergoing CAR T cell therapy. However, further clinical studies are warranted to validate these findings and explore the translational potential of this combinatorial approach in enhancing cancer treatment. [RB1]Again - this takes the emphasis and the reader away from NETs and into the cfDNA universe. Citation Format: Alexey Stepanov, Wenjian Wang, Yingqin Hou, Ivan Chernikov Ivan Chernikov, Reid Bissonnette, George Tetz, Grigory Borisenko, Dmitry Genkin. Neutrophil extracellular traps-targeting DNase I enhances the therapeutic efficacy of CAR-T cell adoptive immunotherapy in a syngeneic murine metastasis model [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr B048.

Abstract C015: IL-27R signaling modulates protective versus pathogenic T cell responses in hepatocellular carcinoma

Abstract Liver cancer is the 3rd leading cause of cancer death. Hepatocellular carcinoma (HCC) is the major form of primary liver cancer, with a growing incidence rate estimated to surge by 50% in the coming twenty years. Although some success has been achieved with immune checkpoint blockade inhibitors, many HCC patients do not respond well to the treatments. Therefore, it is critical to better understand immune mechanisms regulating HCC to identify new biomarkers and develop novel immunotherapies for HCC patients. HCC is driven by chronic liver diseases and is frequently associated with chronic inflammation in the liver. Cytokines are small mediators of inflammation that play an essential role in inflammatory diseases and tumorigenesis. IL27 is a member of the IL6/IL12 superfamily with context-dependent roles in various inflammatory disorders and cancer. IL27 receptor (R) is expressed by most immune cells and several non-immune cells and plays pivotal roles in the regulation of immune responses. Recent work from our lab shows that IL27R signaling suppresses anti-cancer immune response in HCC, acting via the control of innate cytotoxic cells, and Il27ra -/- mice develop significantly less HCC. However, how IL27R signaling regulates T cell subsets in HCC in vivo remains elusive. Here, using diethylnitrosamine (DEN)-induced mouse model of HCC we found a reduction of regulatory T cells (Tregs) in tumor-bearing Il27ra-/-mice along with lowered HCC burden. ScRNA sequencing of CD45+ cells isolated from HCC tumors indicated a less proliferative phenotype of Tregs and more cytotoxic and less exhausted CD8 T cells in mice with Il27ra deficiency. Pathway analysis suggested that IL27R-deficient Tregs were more quiescent, as characterized by the downregulation of various metabolic pathways. Our newly generated Foxp3 cre Il27ra flox mice show a significant reduction of DEN-driven HCC accompanied by an increase of tumor-infiltrating activated CD8 T cells compared to IL27R sufficient controls. Treg-specific deletion of IL27R caused the upregulation of TCF1 in Tregs which has been recently demonstrated to control Treg immune-suppressive functions in colorectal cancer. Meanwhile, tumor-infiltrating effector CD8 T cells were increased in Foxp3 cre+ Il27ra flox mice relative to their Foxp3 cre- Il27ra flox littermate controls. Interestingly, CD8 T cell specific deletion of IL27R did not impact HCC growth in CD8 cre Il27ra flox mice. Overall, our data suggest that IL27R signaling suppresses anti-HCC immunity by enhancing the pro-tumorigenic Tregs and suppressing anti-tumor effector CD8 T cell functions. Citation Format: Zhengzheng Shi, Jiani Zhu, Aleksandra Mazitova, Anastasiia Marchenko, Sergei Grivennikov, Ekaterina Koltsova. IL-27R signaling modulates protective versus pathogenic T cell responses in hepatocellular carcinoma [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr C015.

Abstract B022: Investigating how monocytes impair NK cell cytotoxicity in lung metastasis of triple-negative breast cancer

Abstract Triple-negative breast cancer (TNBC) accounts for 20% of breast cancer (BC) cases worldwide, yet due to its aggressive phenotype and lack of targeted therapies it has the worst prognosis amongst BC subtypes. Up to 40% of TNBC patients will relapse after treatment and develop distant metastasis. Metastasis is the complex process by which disseminated tumor cells (DTCs) leave the primary tumor and grow at distal sites. Since most metastases occur within 3 years of diagnosis, the rapid onset and lack of effective treatments in metastatic TNBC (mTNBC) means less than 20% of patients survive after 4 years. Even though it is the most “immune-activated” BC subtype, and immune checkpoint blockade (ICB) has shown clinical benefits in early TNBC, there is a clear need for novel immunotherapeutic approaches for patients with mTNBC. Natural killer (NK) cells are innate lymphocytes that function as the body’s first line defense against metastatic cancer cells. NK cells eliminate DTCs in the blood or at distal sites via NK cell cytotoxicity (NKCC) and this process is known to be impaired in successful metastasis. We hypothesized that other cells within the tumor microenvironment (TME) can inhibit NKCC at the earliest stages of metastatic outgrowth. We have used Cherry-niche, an in vivo labelling tool, to characterize how TME cells change in early and established lung micro-metastases by single-cell RNA sequencing (scRNA-seq) and flow cytometry. Our initial analyses showed that monocytes are highly enriched, not only in the entire metastatic lung, but specifically in the established niche surrounding DTCs. Although we detected a slight increase in total NK cells, there was a shift in maturation status with fewer mature NK cells (CD27-CD11b+) and more immature NK cells (CD27+/-CD11b-) in the metastatic lung. Furthermore, NK cells were largely absent from the metastatic niche and those that were present displayed a dysfunctional phenotype. CellChat and NicheNet analyses of our scRNA-seq data predicted that several receptor-ligand interactions, including the cytokine macrophage migration inhibitory factor (MIF), can mediate the crosstalk between NK cells and monocytes. Next, we developed an in vitro 3D co-culture system to study the effect of blood-derived monocytes on human NKCC against mTNBC cells. We found that monocytes can suppress NKCC against MDA-MB-231 cells and decrease the viability of mature NK cells. Furthermore, recombinant MIF was able to directly inhibit NKCC, whilst a selective MIF antagonist was able to rescue monocyte-induced impairment of NKCC. These data implicate MIF as a critical monocyte-derived factor in the suppression of NK cell function against mTNBC cells. Future work will aim to identify the exact mechanism by which MIF inhibits NKCC, whilst also evaluate the effect of monocyte depletion and MIF inhibition on metastatic outgrowth and NK cell activation/maturation in vivo. This study will reveal novel inter-cellular dynamics occurring in the metastatic lung, as well as immunotherapeutic targets that may restore NKCC in mTNBC. Citation Format: Christos Ermogenous, Luigi Ombrato, Gordon Beattie. Investigating how monocytes impair NK cell cytotoxicity in lung metastasis of triple-negative breast cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Tumor-body Interactions: The Roles of Micro- and Macroenvironment in Cancer; 2024 Nov 17-20; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(22_Suppl):Abstract nr B022.

Resveratrol Enhances the Efficacy of Combined BM-MSCs Therapy for Rat Spinal Cord Injury via Modulation of the Sirt-1/NF-κB Signaling Pathway.

Spinal cord injury (SCI) represents a severe trauma to the central nervous system, resulting in significant disability and imposing heavy burdens on families and society. Pathophysiological changes following SCI often trigger secondary injuries that complicate treatment. Bone marrow mesenchymal stem cells (BM-MSCs) have become a focal point of research due to their multifunctionality and self-renewal capabilities; however, their survival and neuroprotective functions are compromised in inflammatory environments. Resveratrol, known for its anti-inflammatory, anti-aging, and anti-oxidative stress properties, has been extensively studied. This research focuses on the anti-inflammatory effects of resveratrol post-SCI and its combined application with BM-MSCs to treat rat spinal cord injuries, exploring both efficacy and mechanisms. In vivo experiments investigated changes in the Sirt-1 signaling pathway post-SCI, while in vitro studies examined the effects of resveratrol on BM-MSCs under inflammatory conditions. The assessment included recovery of motor function, neuronal survival, and apoptosis in SCI rats treated with resveratrol alone or in combination with BM-MSCs. Findings reveal a correlation between Sirt-1 and inflammation signaling pathways post-injury. Resveratrol significantly enhanced the survival and efficacy of BM-MSCs in inflammatory environments by upregulating Sirt-1 and downregulating NF-κB and other inflammatory markers, thereby reducing apoptosis. Combined treatment with resveratrol and BM-MSCs showed superior outcomes in motor function recovery and neuronal survival compared to treatment with BM-MSCs alone. This study offers a novel therapeutic strategy for SCI, enhancing stem cell survival and function through modulation of the Sirt-1/NF-κB pathway, providing a theoretical and experimental foundation for clinical applications.

LRRC8A drives NADPH oxidase-mediated mitochondrial dysfunction and inflammation in allergic rhinitis.

Allergic rhinitis (AR) is a complex disorder with variable pathogenesis. Increasing evidence suggests that the LRRC8A is involved in maintaining cellular stability, regulating immune cell activation and function, and playing significant roles in inflammation. However, the involvement of LRRC8A in AR inflammation and its underlying mechanisms remain unclear. LRRC8A expression in AR patients, confirmed by qRT-PCR and Western blotting, was analyzed to investigate its relationship with the clinical characteristics of AR patients. In vitro, IL-13 stimulated HNEpCs to establish a Th2 inflammation model, with subsequent LRRC8A knockout or overexpression. NOX1/NOX4 inhibitor (GKT137831) and chloride channel inhibitor (DCPIB) were utilized to investigate AR development mechanisms during LRRC8A overexpression. An OVA-induced AR model with nasal mucosa LRRC8A knockdown confirmed LRRC8A's regulatory role in AR inflammation. LRRC8A mRNA and protein levels were significantly elevated in AR patients, positively correlating with NADPH oxidase subunits and Th2 inflammatory markers. In vitro, IL-13 stimulation of HNEpCs resulted in upregulation of LRRC8A and increased expression of NOX1, NOX4, and p22phox, along with mitochondrial dysfunction and NF-κB pathway activation. The knockout of LRRC8A reversed these effects. In nasal mucosal epithelial cells, DCPIB and GKT137831 completely blocked mitochondrial dysfunction caused by the overexpression of LRRC8A, which led to up-regulation of NOX1, NOX4, and p22phox. In vivo, knocking down LRRC8A reduced eosinophil infiltration, downregulated the expression of NOX1, NOX4, p22phox IL-4, IL-5, and IL-13, and decreased NF-κB pathway activation. LRRC8A drives the upregulation of NOX1, NOX4, and p22phox, leading to ROS overproduction and mitochondrial dysfunction. It also activates NF-κB, ultimately leading to nasal mucosal epithelial inflammation. LRRC8A may be a potential target for the treatment of AR.

A Computational Pipeline for Identifying Gene Regulatory Networks: A Case Study of Response to Exercise.

Gene regulatory networks are foundational in the control of virtually all biological processes. These networks orchestrate a myriad of cell functions ranging from metabolic rate to the response to a drug or other intervention. The data required to accurately identify these control networks remains very cost and labor intensive typically leading to relatively sparse time course data that is largely incompatible with conventional data-driven model identification techniques. In this work, we combine empirical identification of gene-gene interactions with constraints describing the expected dynamic behavior of the network to infer regulatory dynamics from under-sampled data. We apply this to the identification of gene regulatory subnetworks recruited in groups of subjects participating in several different exercise interventions. Intervention-specific response networks are compared to one another and control actions driving differences are identified. We propose that this approach can extract statistically robust and biologically meaningful insights into gene regulatory dynamics from a dataset consisting of a small number of participants with very limited longitudinal sampling, for example pre- and post- intervention only.

Single-cell expression and immune infiltration analysis of polyamine metabolism in breast cancer.

Breast cancer is one of the most threatening women health diseases worldwide and its molecular heterogeneity offers a range of response to therapy. The role of polyamine metabolism is receiving increasing attention. Polyamine metabolism not only plays an important role in the occurrence and development of breast cancer, but also interacts with tumor immune microenvironment. In this work, we applied single-cell RNA-sequencing (scRNA-seq) and systems immunological approaches to interrogate immune cell infiltration gene-to-gene co-expressions in the bulk tumor transcriptomes of breast cancer. We acquired breast cancer sample data from The Cancer Genome Atlas (TCGA) and Gene Expression Omnibus (GEO), evaluated the infiltration status of 22 immune cell types using CIBERSORTx tool, respectively. By leveraging the Retrospective Breast sample of various technologies including gene expression and methylation, we identified 46 breast cancer proliferation-associated co-expression modules using weighted gene coexpression network analysis (WGCNA) approach along with machine learning models which in turn delineated single cell level expressions features that these selected module possessed. We observed substantial cellular heterogeneity in the breast cancer microenvironment, where lineage-specific gene expression patterns were highly associated with tumor progression. Moreover, we also identified the gene modules correlated with immune cell infiltration level that could function as regulators in response to tumors for immune therapy. Moreover, risk scores were correlated with immune cell function in different patient groups defined by high- and low-risk. The findings of this study shed a new light upon molecular classification prognostic assessment and personalized treatment in breast cancer.

H2S-Prdx4 axis mitigates Golgi stress to bolster tumor-reactive T cell immunotherapeutic response.

The role of tumor microenvironment (TME)-associated inadequate protein modification and trafficking due to insufficiency in Golgi function, leading to Golgi stress, in the regulation of T cell function is largely unknown. Here, we show that disruption of Golgi architecture under TME stress, identified by the decreased expression of GM130, was reverted upon treatment with hydrogen sulfide (H2S) donor GYY4137 or overexpressing cystathionine β-synthase (CBS), an enzyme involved in the biosynthesis of endogenous H2S, which also promoted stemness, antioxidant capacity, and increased protein translation, mediated in part by endoplasmic reticulum-Golgi shuttling of Peroxiredoxin-4. In in vivo models of melanoma and lymphoma, antitumor T cells conditioned ex vivo with exogenous H2S or overexpressing CBS demonstrated superior tumor control upon adoptive transfer. Further, T cells with high Golgi content exhibited unique metabolic and glycation signatures with enhanced antitumor capacity. These data suggest that strategies to mitigate Golgi network stress or using Golgihi tumor-reactive T cells can improve tumor control upon adoptive transfer.

Glucagon-like peptide-1 agonists: Role of the gut in hypoglycemia unawareness, and the rationale in type 1 diabetes

Type 1 diabetes is increasing and the majority of patients have poor glycemic control. Although advanced technology and nanoparticle use have greatly enhanced insulin delivery and glucose monitoring, weight gain and hypo-glycemia remain major challenges and a constant source of concern for patients with type 1 diabetes. Type 1 diabetes shares some pathophysiology with type 2 diabetes, and an overlap has been reported. The above observation created great interest in glucagon-like peptide-1 receptor agonists (GLP-1) as adjuvants for type 1 diabetes. Previous trials confirmed the positive influence of GLP-1 agonists on β cell function. However, hypoglycemia unawareness and dysregulated glucagon response have been previously reported in patients with recurrent hypoglycemia using GLP-1 agonists. Jin et al found that the source of glucagon dysregulation due to GLP-1 agonists resides in the gut. Plausible explanations could be gut nervous system dysregulation or gut microbiota disruption. This review evaluates the potential of GLP-1 agonists in managing type 1 diabetes, particularly focusing on their impact on glycemic control, weight management, and glucagon dysregulation. We provide a broader insight into the problem of type 1 diabetes mellitus management in the light of recent findings and provide future research directions.

Critical radicle length window governing loss of dehydration tolerance in germinated Perilla seeds: insights from physiological and transcriptomic analyses.

Perilla (Perilla frutescens L. Britt.) is an important oilseed and medicinal crop that frequently faces seasonal drought stress during seed germination, leading to a loss of dehydration tolerance (DT), which affects seed emergence and significantly reduces yield. DT has been successfully re-established for many species seeds. However, the physiological mechanisms and gene networks that regulate Perilla's response to DT loss remain unclear. Phenotypic analysis determined that the window for DT in Perilla seeds occurs at radicle lengths of 0-4mm. Integrating physiological and transcriptomic analyses revealed that the loss of DT promotes the production of reactive oxygen species (ROS) and regulates oxidase activity and gene expression. This implies that DT may influence seed germination by modulating ROS activity. Four radicle length (i.e., 0, 1, 3, and 4mm) stages were analyzed, and 262 differentially expressed genes (DEGs) were identified that responded to DT. The majority of these genes were associated with epigenetics, cell function, and transport mechanisms. Analysis of expression data shows that desiccation inhibits the signaling network of genes encoding small secreted peptides (SSPs) and receptor-like protein kinases (RLKs). Finally, a relevant network diagram of DT response was proposed. Based on this information, we have revealed the metabolism regulation maps of the four main pathways involving these DEGs (i.e., metabolic pathways, cell cycle, plant hormone signal transduction, and motor proteins). In conclusion, these findings deepen our understanding of gene network responses to DT during Perilla seed germination and provide potential target genes for the genetic improvement of drought resistance in this crop.

TET2 regulates early and late transitions in exhausted CD8+ T cell differentiation and limits CAR T cell function.

CD8+ T cell exhaustion hampers control of cancer and chronic infections and limits chimeric antigen receptor (CAR) T cell efficacy. Targeting TET2 in CAR T cells provides therapeutic benefit; however, TET2's role in exhausted T cell (TEX) development is unclear. In chronic lymphocytic choriomeningitis virus (LCMV) infection, TET2 drove conversion from stem cell-like TEX progenitors toward terminally differentiated and effector (TEFF)-like TEX. TET2 also enforced a terminally differentiated state in the early bifurcation between TEFF and TEX, indicating broad roles for TET2 in acquisition of effector biology. To exploit the therapeutic potential of TET2, we developed clinically actionable TET2-targeted CAR T cells by disrupting TET2 via knock-in of a safety switch alongside CAR knock-in at the TRAC locus. TET2-targeted CAR T cells exhibited restrained terminal exhaustion in vitro and enhanced antitumor responses in vivo. Thus, TET2 regulates fate transitions in TEX differentiation and can be targeted with a safety mechanism in CAR T cells for improved tumor control.

Enhancing Regeneration and Functionality of Excitable Tissues via Integrating Bioelectronics and Bioengineered Constructs

Abstract The inherent complexities of excitable cardiac, nervous, and skeletal muscle tissues pose great challenges in constructing artificial counterparts with bioelectrical, structural, and mechanical properties that closely resemble their natural states. Recent advances have increasingly revealed the beneficial impact of bioelectrical microenvironments on cellular behaviors, functional tissue regeneration, and improved therapeutic outcomes for these excitable tissues. This review aims to unveil the mechanisms by which electrical microenvironments enhance the regeneration and functionality of excitable cells and tissues, considering both endogenous electrical cues from electroactive biomaterials and exogenous electrical stimuli from external electronic systems. We explore the synergistic effects of these electrical microenvironments, combined with structural and mechanical guidance, on the regeneration of excitable tissues using tissue engineering scaffolds. Additionally, the emergence of micro/nanoscale bioelectronics has significantly expanded the scope of this field, enabling intimate interactions between implantable bioelectronics and excitable tissues at the cellular, tissue, and organ levels. These interactions allow for precise data extraction and localized modulation of cell and tissue functionalities through intricately designed electronic components according to physiological needs. The integration of tissue engineering and bioelectronics holds the potential to achieve optimal outcomes, highlighting a growing trend in engineering living tissue construct-bioelectronic hybrids for restoring and monitoring damaged excitable tissues. Furthermore, we envision critical challenges in engineering the next generation of these hybrids, focusing on integrated fabrication strategies, the development of ionic conductive biomaterials, and their convergence with biosensors.

A PSMA-targeted Tri-specific Killer Engager enhances NK cell cytotoxicity against prostate cancer.

Natural killer (NK) cell tumor infiltration is associated with good prognosis in patients with metastatic castration-resistant prostate cancer (mCRPC). NK cells recognize and kill targets by a process called natural cytotoxicity. We hypothesized that promoting an antigen-specific synapse with co-activation may enhance NK cell function in mCRPC. We describe a Tri-specific Killer Engager (TriKE) construct that engages with the activating receptor CD16 on NK cells, prostate-specific membrane antigen (PSMA) on mCRPC cells, and has an interleukin (IL)-15 moiety that is essential for NK cell survival, proliferation, and priming. We show that the PSMA TriKE specifically binds to PSMA-expressing cells and significantly enhances expansion, degranulation and cytokine production of NK cells derived from healthy donors or prostate cancer patients. Bystander killing of PSMA-negative was also achieved with PSMA TriKE treatment when co-cultured with PSMA-positive cells, suggesting potential PSMA TriKE benefit in controlling tumor antigen escape. When tested under physiologic conditions recapitulating the mCRPC tumor microenvironment (TME), NK cells treated with PSMA TriKE and prolonged exposure to hypoxia or MDSCs maintained their potent function while IL-15 treated NK cells showed greatly impaired cytotoxicity. Finally, in vivo testing of PSMA TriKE showed improved tumor control and survival of mice as compared to IL-15 and untreated control groups. In conclusion, PSMA TriKE demonstrates potential as a new therapy for advanced prostate cancer by providing additional signals to NK cells to maximize their anti-tumor potential in prostate cancer, especially in the setting of a hostile TME.

Targeting the mTOR-Autophagy Axis: Unveiling Therapeutic Potentials in Osteoporosis

Osteoporosis (OP) is a widespread age-related disorder marked by decreased bone density and increased fracture risk, presenting a significant public health challenge. Central to the development and progression of OP is the dysregulation of the mechanistic target of the rapamycin (mTOR)-signaling pathway, which plays a critical role in cellular processes including autophagy, growth, and proliferation. The mTOR-autophagy axis is emerging as a promising therapeutic target due to its regulatory capacity in bone metabolism and homeostasis. This review aims to (1) elucidate the role of mTOR signaling in bone metabolism and its dysregulation in OP, (2) explore the interplay between mTOR and autophagy in the context of bone cell activity, and (3) assess the therapeutic potential of targeting the mTOR pathway with modulators as innovative strategies for OP treatment. By examining the interactions among autophagy, mTOR, and OP, including insights from various types of OP and the impact on different bone cells, this review underscores the complexity of mTOR’s role in bone health. Despite advances, significant gaps remain in understanding the detailed mechanisms of mTOR’s effects on autophagy and bone cell function, highlighting the need for comprehensive clinical trials to establish the efficacy and safety of mTOR inhibitors in OP management. Future research directions include clarifying mTOR’s molecular interactions with bone metabolism and investigating the combined benefits of mTOR modulation with other therapeutic approaches. Addressing these challenges is crucial for developing more effective treatments and improving outcomes for individuals with OP, thereby unveiling the therapeutic potentials of targeting the mTOR-autophagy axis in this prevalent disease.