Mature Cells Research Articles

Gamabufotalin loaded micro-nanocomposites for multimodal therapy of metastatic TNBC by efficiently inducing ICD

Gamabufotalin (CS-6), a main active compound derived from Chinese medicine Chansu, exhibits a robust inhibitory effect on programmed death-ligand 1 (PD-L1) in triple-negative breast cancer (TNBC) cells. Despite its potential for tumor therapy, the medical application of CS-6 is constrained by its hydrophobic nature, lack of targeting capability, and weak immunogenic cell death (ICD) effect. To address these limitations and improve the therapeutic efficiency of this drug against metastatic TNBC, we designed a new kind of CS-6@CPB-S.lux that integrates carboxy-Prussian blue nanoparticles (CPB NPs), CS-6, and attenuated Salmonella typhimurium (S.lux) for TNBC therapy. In vitro and in vivo results have confirmed that CS-6@CPB NPs were efficiently delivered to neoplastic tissue by the tumor hypoxic chemotaxis property of S.lux, wherein the nanomedicine induced significant tumor cell necroptosis and apoptosis via photothermal therapy (PTT) of CPB NPs and chemotherapy of CS-6, which elicited ICD and inhibited PD-L1 expression, resulting in dendritic cells (DCs) maturation and effector T cells activation to comprehensively eliminate tumors. Additionally, the CS-6@CPB-S.lux + Laser treatment significantly transformed the immunosuppressive tumor microenvironment (TME), enhancing antitumor immunity through promoting the polarization of tumor-associated macrophages into antitumorigenic M1 and reducing Tregs recruitment. Consequently, this comprehensive therapy not only inhibited primary and abscopal tumor progression but also prevented TNBC metastasis, which significantly prolonged survival time in animal models. In summary, these findings indicated an alternative approach for metastatic TNBC therapy.

Deferasirox, an iron chelator, impacts myeloid differentiation by modulating NF-kB activity via mitochondrial ROS.

The iron chelator deferasirox (DFX) is effective in the treatment of iron overload. In certain patients with myelodysplastic syndrome, DFX can also provide a dramatic therapeutic benefit, improving red blood cell production and decreasing transfusion requirements. Nuclear Factor-kappa B (NF-kB) signalling has been implicated as a potential mechanism behind this phenomenon, with studies focusing on the effect of DFX on haematopoietic progenitors. Here, we examine the phenotypic and transcriptional effects of DFX throughout myeloid cell maturation in both murine and human model systems. The effect of DFX depends on the stage of differentiation, with effects on mitochondrial reactive oxygen species (ROS) production and NF-kB pathway regulation that vary between progenitors and neutrophils. DFX triggers a greater increase in mitochondrial ROS production in neutrophils and this phenomenon is mitigated when cells are cultured in hypoxic conditions. Single-cell transcriptomic profiling revealed that DFX decreases the expression of NF-kB and MYC (c-Myc) targets in progenitors and decreases the expression of PU.1 (SPI1) gene targets in neutrophils. Together, these data suggest a role of DFX in impairing terminal maturation of band neutrophils.

Enhancing Tumor-Specific immunity with SLdacA: A attenuated Salmonella-mediated c-di-AMP delivery system targeting the STING pathway

The STING agonist stimulates an anti-tumor immune response by activating T cells, but its limited tumor-targeting specificity poses risks of cytokine storms or autoimmune reactions. Conversely, attenuated Salmonella typhimurium △ppGpp (S.t△ppGpp) exhibits superior tumor-targeting specificity and potent anti-tumor immunogenicity. However, the anti-tumor effects of Salmonella carrying STING agonists remain underexplored. In this study, we engineered a strain called SLdacA, utilizing S.t△ppGpp as a carrier, to produce c-di-AMP. This engineered strain effectively enhances dendritic cell maturation and M1-type macrophage polarization by inducing type I interferon production, thereby recruiting and activating effector T cells against tumor progression. This process is regulated by the STING/type I interferon pathway. Our findings indicate that utilizing S.t△ppGpp as a delivery vehicle for STING agonists holds promise as a strategy for synergistic bacterial-mediated immunotherapy.

Fmo5 plays a sex-specific role in goblet cell maturation and mucus barrier formation.

The intestine plays a key role in metabolism, nutrient and water absorption, and provides both physical and immunological defense against dietary and luminal antigens. The protective mucosal lining in the intestine is a critical component of intestinal barrier that when compromised, can lead to increased permeability, a defining characteristic of inflammatory bowel disease (IBD), among other intestinal diseases. Here, we define a new role for the flavin-containing monooxygenase (FMO) family of enzymes in maintaining a healthy intestinal epithelium. Using Caenorhabditis elegans we measure intestinal barrier function, actin expression, and intestinal damage response. In mice, we utilize an intestine-specific, tamoxifen- inducible knockout model of the mammalian homolog of Cefmo-2 , Fmo5, and assess histology, mucus barrier thickness, and goblet cell physiology. We also treat mice with the ER chaperone Tauroursodeoxycholic acid (TUDCA). In nematodes, we find Cefmo-2 is necessary and sufficient for intestinal barrier function, intestinal actin expression, and is induced by intestinal damage. In mice, we find striking changes to the intestine within two weeks following Fmo5 disruption. Alterations include sex-dependent changes in colon epithelial histology, goblet cell localization, and mucus barrier formation. These changes are significantly more severe in female mice, mirroring differences observed in IBD patients. Furthermore, we find increased protein folding stress in Fmo5 knockout animals and successfully rescue the severe female phenotype with addition of a chemical ER chaperone. Together, our results identify a highly conserved and novel role for Fmo5 in the mammalian intestine and support a key role for Fmo5 in maintenance of ER/protein homeostasis and proper mucus barrier formation.

Myokines May Be the Answer to the Beneficial Immunomodulation of Tailored Exercise-A Narrative Review.

Exercise can regulate the immune function, activate the activity of immune cells, and promote the health of the organism, but the mechanism is not clear. Skeletal muscle is a secretory organ that secretes bioactive substances known as myokines. Exercise promotes skeletal muscle contraction and the expression of myokines including irisin, IL-6, BDNF, etc. Here, we review nine myokines that are regulated by exercise. These myokines have been shown to be associated with immune responses and to regulate the proliferation, differentiation, and maturation of immune cells and enhance their function, thereby serving to improve the health of the organism. The aim of this article is to review the effects of myokines on intrinsic and adaptive immunity and the important role that exercise plays in them. It provides a theoretical basis for exercise to promote health and provides a potential mechanism for the correlation between muscle factor expression and immunity, as well as the involvement of exercise in body immunity. It also provides the possibility to find a suitable exercise training program for immune system diseases.

Selected stem cell populations in pediatric acute lymphoblastic leukemia.

Acute lymphoblastic leukemia is characterized by a disturbed maturation of hematopoietic stem cells (HSCs) resulting in development of a malignant clone. Despite relatively positive outcome, there are still instances of disease relapse occurring due to ineffective disease eradication or primary leukemic clone alterations. Unclear significance of stem cells in the course of ALL led us to investigate and establish crucial changes in two stem cell populations - very small embryonic-like stem cells (VSELs) and HSCs during the induction phase of treatment. In a retrospective study selected stem cells in peripheral blood and bone marrow of 60 pediatric ALL subjects and 48 healthy controls were subjected to flow cytometric analysis at 4 different time points. Both VSELs and HSCs were elevated at the moment of ALL diagnosis compared to healthy controls, but profoundly decline until day 15. Further observations revealed an increase in HSCs with a concomitant depletion of VSELs until week 12. ALL patients with high HSCs showed positive correlation with bone marrow blasts at diagnosis. Patients with lower VSELs or HSCs at diagnosis had slightly improved response to applied therapy. We observed higher initial bone marrow lymphoblast values in patients with lower VSELs or higher HSCs in the high-risk group. The significance of VSELs in predicting treatment outcome can be illustrated by lower day 15 MRD level of patients with lower VSELs at diagnosis. We found HSCs and VSELs to be valid participants in pediatric ALL with possible contribution in the neoplastic process and prediction of initial treatment outcome.

Ebola Virus Infection of Flt3-Dependent, Conventional Dendritic Cells and Antigen Cross-presentation Leads to High Levels of T-Cell Activation.

Previous studies have described that Ebola virus (EBOV) infection of human monocyte-derived dendritic cells (moDCs) inhibits dendritic cell (DC) maturation, resulting in poor T-cell activation. However, it is unknown how other DC subsets distinct from moDCs respond to EBOV infection. To better understand how DCs initiate T-cell activation during EBOV infection, we assessed the response of conventional mouse DCs (cDCs) to EBOV infection utilizing a recombinant EBOV expressing the model antigen ovalbumin. In contrast to moDCs, mouse cDC2s and cDC1s were poorly infected with EBOV but were highly activated. DCs were able to prime CD8 T cells via cross-presentation of antigens obtained from cell debris of EBOV-infected cells. EBOV infection further enhanced DC cross-presentation. Our findings indicate that EBOV infection of cDCs results in activation rather than inhibition, leading to high levels of T-cell activation. With that we propose a mechanistic explanation for the excess T-cell activation observed in human Ebola virus disease.

An Efficient Fabrication Approach for Multi-Cancer Responsive Chemoimmuno Co-Delivery Nanoparticles.

Background/Objectives: Cancer remains one of the leading causes of death, with breast, liver, and pancreatic cancers significantly contributing to this burden. Traditional treatments face issues including dose-limiting toxicity, drug resistance, and limited efficacy. Combining therapeutic agents can enhance effectiveness and reduce toxicity, but separate administration often leads to inefficiencies due to differing pharmacokinetics and biodistribution. Co-formulating hydrophobic chemotherapeutics such as paclitaxel (PTX) and hydrophilic immunologic agents such as polyinosinic-polycytidylic acid (Poly IC) is particularly challenging due to their distinct physicochemical properties. This study presents a novel and efficient approach for the co-delivery of PTX and Poly IC using chitosan-based nanoparticles. Method: Chitosan-PEG (CP) nanoparticles were developed to encapsulate both PTX and Poly IC, overcoming their differing physicochemical properties and enhancing therapeutic efficacy. Results: With an average size of ~100 nm, these nanoparticles facilitate efficient cellular uptake and stability. In vitro results showed that CP-PTX-Poly IC nanoparticles significantly reduced cancer cell viability in breast (4T1), liver (HepG2), and pancreatic (Pan02) cancer types, while also enhancing dendritic cell (DC) maturation. Conclusions: This dual-modal delivery system effectively combines chemotherapy and immunotherapy, offering a promising solution for more effective cancer treatment and improved outcomes.

The Enhancement of Regulatory T Cell Maturation and Th1/Th2 Balance through FOXP3 Expression by Lactobacillus paracasei in an Ovalbumin-Induced Allergic Skin Animal Model.

Chronic allergic skin conditions, including atopic dermatitis (AD), are characterized by pruritus, erythema, xerosis, desquamation, and inflammation, significantly impacting quality of life. Long-term steroid use, while common in treatment, carries the risk of adverse effects. Previous studies have demonstrated the potential of Lactobacillus paracasei subsp. paracasei NTU 101 (NTU 101) in alleviating AD symptoms from a preventive perspective. This study, however, focuses on exploring NTU 101's therapeutic potential by investigating its effects on regulatory T cell (Treg) maturation and Th1/Th2 balance. The results revealed that NTU 101 administration effectively reduced serum IgE levels and inflammatory cell infiltration in the skin, leading to a significant improvement in both epidermal and dermal thickness in the AD model. Additionally, NTU 101 modulated the immune response by increasing the proportion of CD4+/IL-4+ (Th2) cells in the spleen and concurrently enhancing FOXP3 expression in CD4+/CD25+ cells, which is critical for Treg cell development. This immune modulation was further associated with a rebalancing of the Th1/Th2 ratio, achieved by increasing the proportion of CD4+/IFN-γ+ (Th1) cells. Moreover, NTU 101 influenced the proportion of CD4+IL-17+ (Th17) cells, thereby supporting neutrophil maturation and promoting allergen clearance, ultimately mitigating AD symptoms. These findings underscore the potential of NTU 101 not only in managing AD symptoms but also in modulating key immune pathways involved in the pathogenesis of the disease, offering a promising alternative or adjunct to conventional steroid therapies.

BAFF promotes follicular helper T cell development and germinal center formation through BR3 signal.

T follicular helper (Tfh) cells represent an important subset of CD4+ T cells that is crucial to the maturation and differentiation of B cells and the production of high-affinity antibodies. Since BAFF, a vital B cell survival factor, is also crucial to B cell maturation and differentiation, we assessed the effects of BAFF on Tfh cell development and function. We demonstrate that deficiency of BAFF, but not of APRIL, markedly inhibits Tfh cell development, germinal center (GC) formation, and antigen-specific antibody production. The promoting effect of BAFF on Tfh cell development is dependent on expression of BR3 on T cells, and its promoting effect on GC formation is dependent on expression of BR3 on both T cells and B cells. BAFF directly promotes expression of the Tfh cell-characteristic genes via NF-κB signaling. This effect does need BR3 expression. Thus, BAFF not only has direct effects on B cells, but it also has direct effects on Tfh cell differentiation via engagement of BR3 which collectively promote GC formation and production of high-affinity antibodies. This dual effect of BAFF on B cells and Tfh cells may help explain the clinical utility of BAFF antagonists in the management of certain autoimmune diseases.

Multidimensional profiling of human T cells reveals high CD38 expression, marking recent thymic emigrants and age-related naive T cell remodeling

Thymic involution is a key factor in human immune aging, leading to reduced thymic output and a decline in recent thymic emigrant (RTE) naive Tcells in circulation. Currently, the precise definition of human RTEs and their corresponding cell surface markers lacks clarity. Analysis of single-cell RNA-seq/ATAC-seq data distinguished RTEs by the expression of SOX4, IKZF2, and TOX and CD38 protein, whereby surface CD38hi expression universally identified CD8+ and CD4+ RTEs. We further determined the dynamics of RTEs and mature cells in a cohort of 158 individuals, including age-associated transcriptional reprogramming and shifts in cytokine production. Spectral cytometry profiling revealed two axes of aging common to naive CD8+ and CD4+ Tcells: (1) a decrease in CD38++ cells (RTEs) and (2) an increase in CXCR3hi cells. Identification of RTEs enables direct assessment of thymic health. Furthermore, resolving the dynamics of naive Tcell remodeling yields insight into vaccination and infection responsiveness throughout aging.

Innate immune cell activation by adjuvant AS01 in human lymph node explants is age-independent.

Vaccine adjuvants are thought to work by stimulating innate immunity in the draining lymph node (LN), although this has not been proven in humans. To bridge data obtained in animals to humans, we have developed an in situ human LN explant model to investigate how adjuvants initiate immunity. Slices of explanted LNs were exposed to vaccine adjuvants and revealed responses that were not detectable in LN cell suspensions. We used this model to compare the liposome-based AS01 with its components MPL and QS-21, and TLR ligands. Liposomes were predominantly taken up by subcapsular sinus-lining macrophages, monocytes and dendritic cells. AS01 induced dendritic cell maturation and a strong pro-inflammatory cytokine response in intact LN slices but not in dissociated cell cultures, in contrast to R848. This suggests the onset of the immune response to AS01 requires a coordinated activation of LN cells in time and space. Consistent with the robust immune response observed in older adults with AS01-adjuvanted vaccines, the AS01 response in human LNs was independent of age, unlike R848. This human LN explant model is a valuable tool for studying the mechanism of action of adjuvants in humans and for screening new formulations to streamline vaccine development.

Rps3 Attenuates Gastric Precancerous Lesions by Promoting Dendritic Cells Maturation via AKT/β-Catenin Pathway.

This study aimed to investigate the dysregulated proteins and the underlying mechanisms of gastric precancerous lesions. Proteomic and phosphoproteomic methods were used to characterize the proteome and phosphoproteome profiles of N-methyl-N-nitro-N-nitrosoguanidine (MNNG)-induced gastric precancerous lesions. The hub differentially expressed proteins (DEPs) and phosphoproteins (DEPPs) were identified by using differential expression and protein-protein interaction network analyses. Western blot assay, quantitative reverse transcription (qRT)-PCR, and CCK-8 assays detected the expression of Rps3, N-cadherin, E-cadherin, AKT, p-AKT, and β-catenin and verified the roles of Rps3 on the MNNG-induced human gastric epithelial cell line (GES-1) cells. Hub DEPs and phosphoproteins Rps3, Akt1, and Ctnnb1 were significantly correlated with five dendritic cells (DCs) pathways, and Akt1 and Ctnnb1 were significantly negatively correlated with Rps3. MNNG administration markedly reduced the Rps3 mRNA and protein expression levels (all P < 0.05). Overexpression of Rps3 significantly inhibited tumorigenesis of MNNG-induced GES-1 cells (all P < 0.01) and changed the protein levels of N-cadherin, E-cadherin, AKT, p-AKT, and β-catenin. Similarly, SC79 treatment substantially increased the expression of interleukin (IL)-6, IL-10, and vascular endothelial growth factor (all P < 0.05). Rps3 was poorly expressed in precancerous gastric lesions. Correspondingly, overexpression of Rps3 promoted DC maturation via the AKT/β-catenin pathway, inhibiting the progression of gastric precancerous lesions.

In Vitro Proliferation of MG-63 Cells in Additively Manufactured Ti-6Al-4V Biomimetic Lattice Structures with Varying Strut Geometry and Porosity.

Lattice structures have demonstrated the ability to provide secondary stability in orthopedic implants by promoting internal bone growth. In response to the growing prevalence of lattices in orthopedic design, we investigated the effects of porosity and unit cell geometry in additively manufactured Ti-6Al-4V biomimetic lattice structures on the osteogenesis of human MG-63 osteoblastic cell lines in vitro. We analyzed glucose consumption, alkaline phosphatase (ALP) concentration, and end-of-culture cell count as markers for osteogenic growth. Two different strut geometries were utilized (cubic and body-centered cubic), along with four different pore sizes (400, 500, 600, and 900 µm, representing 40-90% porosity in a 10 mm cube), in addition to a solid specimen. Structural characterization was performed using scanning electron microscopy. The results indicated that lattices with a 900 µm pore size exhibited the highest glucose consumption, the greatest change in ALP activity, and the highest cell count when compared to other pore sizes. Cubic 900 µm lattice structures outperformed other specimens in facilitating the maturation of viable MG-63 cells from the formation to the mineralization phase of bone remodeling, offering the most promise for osseointegration in additively manufactured titanium implants in the future. However, irrespective of a particular pore size or unit cell geometry, it was found that all the lattices were capable of promoting osteogenic growth due to surface roughness in the printed parts.

Exploring the impact of tertiary lymphoid structures maturity in NSCLC: insights from TLS scoring.

Tertiary Lymphoid Structures (TLS) are lymphoid structures commonly associated with improved survival of cancer patients and response to immunotherapies. However, conflicting reports underscore the need to consider TLS heterogeneity and multiple features such as TLS size, composition, and maturation status, when assessing their functional impact. With the aim of gaining insights into TLS biology and evaluating the prognostic impact of TLS maturity in Non-Small Cell Lung Carcinoma (NSCLC), we developed a multiplex immunofluorescent (mIF) panel including T cell (CD3, CD8), B cell (CD20), Follicular Dendritic cell (FDC) (CD21, CD23) and mature dendritic cell (DC-LAMP) markers. We deployed this panel across a cohort of primary tumor resections from NSCLC patients (N=406) and established a mIF image analysis workstream to specifically detect TLS structures and evaluate the density of each cell phenotype. We assessed the prognostic significance of TLS size, number, and composition, to develop a TLS scoring system representative of TLS biology within a tumor. TLS relative area, (total TLS area divided by the total tumor area), was the most prognostic TLS feature (C-index: 0.54, p = 0.04). CD21 positivity was a marker driving the favorable prognostic impact, where CD21+ CD23- B cells (C-index: 0.57, p = 0.04) and CD21+ CD23- FDC (C-index: 0.58, p = 0.01) were the only prognostic cell phenotypes in TLS. Combining the three most robust prognostic TLS features: TLS relative area, the density of B cells, and FDC CD21+ CD23- we generated a TLS scoring system that demonstrated strong prognostic value in NSCLC when considering the effect of age, sex, histology, and smoking status. This TLS Score also demonstrated significant association with Immunoscore, EGFR mutational status and gene expression-based B-cell and TLS signature scores. It was not correlated with PD-L1 status in tumor cells or immune cells. In conclusion, we generated a prognostic TLS Score representative of the TLS heterogeneity and maturity undergoing within NSCLC tissues. This score could be used as a tool to explore how TLS presence and maturity impact the organization of the tumor microenvironment and support the discovery of spatial biomarker surrogates of TLS maturity, that could be used in the clinic.

Macrophage-Activated Products of Lacticaseibacillus paracasei N1115 Promote the Development of Primary Hippocampal Neurons

To make a preliminary investigation of the effect of the immune pathway mediated by live Lacticaseibacillus paracasei N1115 on the development of primary hippocampal neurons cultured in vitro. Live Lacticaseibacillus paracasei N1115 suspension of an appropriate concentration was used as the experimental group. Peptidoglycan (PGN) and lipopolysaccharide (LPS) were used as positive controls, and RPMI1640 medium served as the blank control. These were co-cultured with RAW264.7 cells to obtain the co-culture mediums and the total cellular RNA, and to measure the expression and secretion of cytokines. After centrifugation, the supernatants were co-cultured with primary hippocampal neurons at appropriate ratios. The co-culture mediums were collected, and the total cellular RNA was extracted to measure the expression of genes related to synaptic development in neurons. Following immunofluorescence staining of the primary hippocampal neurons, the presynaptic and presynaptic membrane-associated proteins, including synaptophysin (SYP) and the postsynaptic density protein 95 (PSD95), and neuronal cell maturation markers, including microtubule-associated protein 2 (MAP-2), and doublecortin (DCX) were quantitatively analyzed. Additionally, the morphological development of the neurons were measured. Compared with the blank control, the mRNA expression levels of interleukin (IL)-6 and tumor necrosis factor- α (TNF-α) increased by 7471% and 926%, respectively, after the RAW264.7 cells were treated with live Lacticaseibacillus paracasei N1115, while their secretion levels increased by 184.16 pg/mL and 12320.76 pg/mL, respectively, all showing statistically significant differences (P<0.05). The activation ability of N1115 live bacteria was stronger than that of PGN but weaker than that of LPS, showing statistically significant differences (P<0.05). Compared with the blank control, following the intervention with the supernatant from the co-culture of N1115 live bacteria and RAW264.7 cells, the viability of primary hippocampal neurons in the 10% supernatant intervention group increased by 19.25%, showing statistically significant differences (P<0.05), and the mRNA expression of SYP and PSD95 increased by 137% and 159%, respectively, showing statistically significant differences (P<0.05). The total neurite length (489.88 μm) of the neurons of the group intervened with the supernatant of N1115 live bacteria was increased compared to that of the blank control group (381.51 μm), and the cell body area (2092.22 μm2), maximum neurite length (184.78 μm), total neurite length, average neurite length (108.38 μm), and branching points (4.84 s) were higher than those in the two positive control groups, showing statistically significant differences (P<0.05). Lacticaseibacillus paracasei N1115 can significantly activate the immune regulatory function of macrophages, thereby promoting the morphological development and synaptic function of nerve cells.

An innovative approach to overcoming PD-1 resistance: Combined TIGIT blockade with nanophotothermal therapy

Photothermal immunotherapy, especially combined with immune checkpoint inhibitors (ICIs) targeting programmed cell death 1 (PD-1), marks a breakthrough in cancer treatment. However, therapy often triggers compensatory upregulation of other immune checkpoints. This study introduces an advanced strategy to enhance the efficacy of photothermal immunotherapy by utilizing a novel nanocarrier that targets both PD-1 and T-cell immunoreceptor with Ig and ITIM domains (TIGIT) immune checkpoints. The dual blockade could counteract the compensatory upregulation of immune checkpoints triggered by PD-1 targeted therapies alone. The nanocarrier of this work is composed of a ZrB2 photothermal core and a mesoporous silica layer, and finally coated with biocompatible polydopamine. This design could effectively deliver both TIGIT and PD-1 inhibitors directly to tumor sites, minimizing systemic side effects such as cytokine release syndrome. We demonstrate that this approach not only enhances cytotoxic responses of T and NK cells, but also promotes macrophage polarization towards a tumor-suppressing M1 phenotype dendritic cell maturation and immunogenic cell death. Our findings provide a promising avenue for cancer immunotherapy.

Role of IL-6 in Physiology and Pathology of the Ovary

Objective: We aimed to explore the roles of interleukin (IL)-6 in ovarian physiology and pathology. Mechanism: IL-6 is a common and important cytokine in numerous physiological and pathological processes. Findings in Brief: In the ovary, IL-6 is secreted by granulosa cells, and involved in many crucial physiological functions, including normal ovulation and menstrual cycle, hormone secretion, egg cell maturation, and even in the fertilization process of egg cells. The abnormal secretion of IL-6 is associated with premature ovarian failure, polycystic ovary syndrome and ovarian cancer. Conclusions: This review will summarize existing clues and explore the key roles of IL-6 in ovarian physiology and pathology, which will provide new targets for the treatment of common ovarian diseases.

PROTAC-mediated vimentin degradation promotes terminal erythroid differentiation of pluripotent stem cells

BackgroundHuman pluripotent stem cells (hPSCs), including human embryonic stem cells (hESCs) and induced pluripotent stem cells (hiPSCs), can undergo erythroid differentiation, offering a potentially invaluable resource for generating large quantities of erythroid cells. However, the majority of erythrocytes derived from hPSCs fail to enucleate compared with those derived from cord blood progenitors, with an unknown molecular basis for this difference. The expression of vimentin (VIM) is retained in erythroid cells differentiated from hPSCs but is absent in mature erythrocytes. Further exploration is required to ascertain whether VIM plays a critical role in enucleation and to elucidate the underlying mechanisms.MethodsIn this study, we established a hESC line with reversible vimentin degradation (dTAG-VIM-H9) using the proteolysis-targeting chimera (PROTAC) platform. Various time-course studies, including erythropoiesis from CD34+ human umbilical cord blood and three-dimensional (3D) organoid culture from hESCs, morphological analysis, quantitative real-time PCR (qRT-PCR), western blotting, flow cytometry, karyotyping, cytospin, Benzidine-Giemsa staining, immunofluorescence assay, and high-speed cell imaging analysis, were conducted to examine and compare the characteristics of hESCs and those with vimentin degradation, as well as their differentiated erythroid cells.ResultsVimentin expression diminished during normal erythropoiesis in CD34+ cord blood cells, whereas it persisted in erythroid cells differentiated from hESC. Depletion of vimentin using the degradation tag (dTAG) system promotes erythroid enucleation in dTAG-VIM-H9 cells. Nuclear polarization of erythroblasts is elevated by elimination of vimentin.ConclusionsVIM disappear during the normal maturation of erythroid cells, whereas they are retained in erythroid cells differentiated from hPSCs. We found that retention of vimentin during erythropoiesis impairs erythroid enucleation from hPSCs. Using the PROTAC platform, we validated that vimentin degradation by dTAG accelerates the enucleation rate in dTAG-VIM-H9 cells by enhancing nuclear polarization.Graphical

Assessment of Adipocyte Transduction Using Different AAV Capsid Variants.

Adeno-associated viruses (AAVs) are widely used as viral vectors for gene delivery in mammalian cells. We focused on the efficacy of the transduction of AAV2/5, 2/6, 2/8 and 2/9 expressing GFP in preadipocyte cells by live imaging microscopy using IncuCyte S3 and flow cytometry. Three transduction modes in 3T3-L1 preadipocyte cells assessed: AAV transduction in 3T3-L1 preadipocyte cells, transduction with further differentiation into mature adipocyte-like cells and the transduction of differentiated 3T3-L1 adipocytes. For the in vivo study, we injected AAV2/6, AAV2/8 and AAV2/9 in adipose tissue of C57BL6 mice, and the transduction capacity of AAV2/6, along with AAV2/8 and AAV2/9 was evaluated. AAV2/6 demonstrated the highest transduction efficiency in 3T3-L1 preadipocytes, as it was 1.5-2-fold more effective than AAV2/5, and AAV2/8 in the range of viral concentrations from 2 × 104 to 1.6 × 105 VG/cell. AAV2/5 and AAV2/8 showed transduction efficiencies similar to each other. The expression of GFP under the CMV promoter remained stable for up to 20 days. The induction of 3T3-L1 differentiation in three days after AAV transduction did not alter the GFP expression level, and AAV2/6 showed the best transduction efficiency. AAV2/6 demonstrated the ability to transduce mature adipocytes. These results were confirmed by in vivo studies on C57BL6 mice. AAV2/6 had the highest transducing activity on both inguinal and interscapular adipose tissue. Thus, AAV2/6 has demonstrated higher transduction efficacy compared to AAV2/5, AAV2/8 and AAV2/9 both in 3T3-L1 adipocytes and adipose tissue in vivo, which proves its usability along with AAV2/8 and AAV2/9 for gene delivery to adipocytes.