Cytotechnology最新文献

Rheumatoid arthritis (RA) is a long-lasting ailment mostly disturbing joints and managed with disease-modifying antirheumatic drug (DMARDs), corticosteroids, and NSAIDs, along with immunosuppressants, though these cause serious adverse effects. Citrus medica L., known for anti-inflammatory, antioxidant, and immunostimulatory properties, was examined for the anti-arthritic activity of its hydroalcoholic extract (HAECF) in Freund's Complete Adjuvant (FCA)-induced RA rats. In this work, HAECF was prepared by using a soxhlet apparatus. Furthermore, HAECF was taken into consideration to analyze phytochemical constitutions. Experimental groups received HAECF at doses of 200 mg/kg and 400 mg/kg orally for 28 days, and therapeutic efficiency was evaluated through body weight, paw volume, arthritis index (AI), hematological and antioxidant parameters, along with histopathological examination. HAECF confirmed significant dose-dependent anti-arthritic actions, as demonstrated by reduction in paw volume (2.35 ± 0.018 at 200 mg/kg and 2.26 ± 0.016 at 400 mg/kg) corresponding to a 43% and 45% reduction, respectively (p < 0.05), and an increase in body weight compared to FCA-induced RA rats. Hematological investigation showed refurbishment of RBC (4.40 ± 0.020 at 200 mg/kg and 4.84 ± 0.018 at 400 mg/kg) and hemoglobin (11.20 ± 0.201 g/dL at 200 mg/kg and 12.42 ± 0.009 g/dL at 400 mg/kg), while WBC, CRP, platelets, ESR, and rheumatoid factor were significantly (p < 0.05) decreased in HAECF-treated groups. Antioxidant markers like MDA, SOD, and catalase were also improved in a dose-dependent manner (p < 0.05). Histopathological examinations confirmed abridged synovial hyperplasia, joint damage, and pannus formation in HAECF-treated rats, supporting its anti-arthritic potential. Therefore, HAECF exhibits significant anti-arthritic potential through anti-inflammatory and antioxidant mechanisms, providing pharmacological justification for its use in RA management.

Glioblastoma (GBM) is a highly malignant central nervous system tumor known for its resistance to current treatment options. Efflux pumps play a crucial role in the development of multidrug resistance (MDR), leading to poor clinical outcomes. In this context, new treatment modalities targeting the cell cycle have gained attention in the recent years. Abemaciclib (ABE), a selective CDK4/6 inhibitor, inhibits the activity of different drug efflux pumps in several types of cancer. However, the role of ABE in regulating these pumps in GBM remains unclear. In this study, the therapeutic efficacy of ABE in the regulation of multidrug resistance pumps was evaluated in TMZ-resistant T98G and TMZ-sensitive U87MG GBM cells. The cells treated with different doses of ABE for 24 h and then CCK-8, Annexin V, cell cycle, AO/PI, RT-PCR and immunofluorescence analysis were performed. Our findings demonstrated that ABE significantly reduced the viability of GBM cell lines. However, the activity of the efflux pumps, especially P-gp and MRP1 pumps, was different due to TMZ resistance. Particularly, ABE treatment reduced the P-gp gene and protein levels in T98G cells. Additionally, ABE differentially affected the cell cycle in terms of TMZ-sensitivity. In conclusion, ABE treatment differentially regulated the inhibition of efflux pump activities in GBM cells in terms of TMZ resistance. Our findings may provide new insights into the potential use of ABE as a therapeutic agent to overcome MDR in TMZ-resistant GBM.

This study investigated the impact of culture surface rigidity on the proliferation and maintenance of undifferentiated human induced pluripotent stem cells (hiPSCs), which are crucial in regenerative medicine and stem cell-based organ therapy. Given the need for a substantial number of cells per patient, there is a pressing demand for methods that ensure homogeneous and efficient large-scale cultivation of hiPSCs. Mechanotransduction, the process by which cells, including hiPSCs, respond to mechanical stimuli, has gained significant attention. We evaluated the effects of varying culture surface rigidity, examining changes in morphology, gene expression, and differentiation tendencies. Soft gels were made by altering acrylamide gel polymerization on functionalized glass slides, which were then coated with laminin. Cell attachment rates were quantified 24 h after seeding which varied according to substrate rigidity. The apparent proliferation rate was highest at 5 kPa, suggesting that hiPSCs sense rigidity. We further confirmed that this mechanosensing occurred through activation of the Hippo signaling pathway. In conclusion, this study revealed that the adhesion and proliferation of hiPSCs are significantly influenced by culture surface rigidity, with 5 kPa identified as the optimal condition for proliferation. This understanding may help optimize cell culture conditions for future organ regeneration and therapeutic applications.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-026-00895-y.

It is believed that CSCs, which are resistant to chemotherapy and radiation, develop in certain situations, stay in the tissue niche after treatment, and cause disease recurrence. Targeting these cells, which are thought to be chemoresistant, may enable treatment to eliminate metastasis or recurrence. For this reason, we aimed to isolate CSCs from various breast cancer cell lines using a drug selection method in this study and then identify them via flow cytometry and fluorescent staining. MDA-123, MCF-7, and 4T1 were used as in vitro models, with HUVEC as controls. After isolation of stem cells from these cells by incubation with 5-fluorouracil (5-FU) and cisplatin (Cis), CSCs were identified by flow cytometry using CD24/CD44/CD133/EPCAM by assessing the rhodamine 123 (Rho 123). The results showed that 5-FU and Cis reduced the CD24 receptor in both cancer and normal cells, while the others showed varying expression levels. Administration of 5-FU and Cis significantly decreased Rho 123 fluorescence intensity in MDA-MB-231, MCF-7, 4T1, and HUVEC cells compared to the untreated group. A subpopulation of breast cancer cells with low CD24 and Rho123 expression was cancer stem cells. The 4T1 cells, which expressed low levels of CD24/CD133/EpCAM expression, had significantly lower Rho 123 levels than MDA-MB-231 and MCF-7 cells. This suggests that 4T1 cells may have greater chemoresistance and more aggressive properties. In conclusion, studying surface receptors and rhodamine uptake for the identification of CSCs is a powerful and valuable method for evaluating the efficacy of chemotherapy. The discovery of CSC markers that indicate therapeutic strategies could lead to clinical trials of medicines with potentially better therapeutic outcomes, particularly for recurring malignancies.

Adipose-derived stromal cells (ADSC) show promise for neuronal differentiation, but their utility is limited by late-stage cell death, which may be driven by endoplasmic reticulum stress (ERS). To investigate this mechanism, we employed an integrated approach combining immunocytochemistry, western blotting, single-cell RNA sequencing (scRNA-Seq), and transmission electron microscopy (TEM) to systematically profile ERS-related gene expression, dynamic changes of key proteins, and ultrastructural evolution of the ER during neuronal induction. Our results demonstrate that ERS pathways are activated throughout the differentiation process. In early stages, the endoplasmic reticulum (ER) chaperone GRP78 initially increased but markedly declined at 6 h and 8 h. Key UPR sensors IRE1α, XBP1s, PERK, and ATF6 peaked in undifferentiated ADSC and Pre-induction (Prei-1d) cells, then gradually decreased as differentiation progressed. In contrast, pro-apoptotic markers CHOP and Caspase-3 were continuously upregulated in later phases, accompanied by ultrastructural hallmarks of ER dilation, disrupted mitochondrial cristae, and cytoplasmic vacuolization. These findings indicate that ERS initially activates the unfolded protein response to maintain ER homeostasis and support differentiation, whereas sustained ERS at later stages shifts toward CHOP/Caspase-3-dependent apoptosis, leading to cellular injury. This study provides a theoretical basis for optimizing neuronal differentiation protocols through time-dependent modulation of ERS pathways.

NFIB, a pivotal transcription factor, intricately influences tumorigenesis by exerting dual roles as either an oncogenic promoter or a tumor-suppressive factor across a spectrum of tumor types. However, the specific impact of NFIB on bladder cancer remains poorly understood. This study aims to elucidate the biological function and molecular mechanism of NFIB in bladder cancer. We first found that the protein level of NFIB was downregulated in bladder cancer tissues compared to matched adjacent noncancerous tissues. Functional assays, including Wound healing and Transwell invasion assays, demonstrated that NFIB suppressed migration, invasion and epithelial-mesenchymal transition (EMT) of bladder cancer cells in vitro, wheras CCK8 assays showed NFIB had no significant effect on cell proliferation. In vivo experiments, including Xenograft and Nude mouse tail vein transfer assays, further supported these observations, indicating that NFIB inhibited lung metastasis of bladder cancer without affecting primary tumor growth. Finally, transcriptomic analysis confirmed that NFIB hindered the activation of the PI3K-AKT signaling pathway. Taken together, this study highlighted that NFIB serves as a tumor suppressor gene in bladder cancer, and suppresses cell migration, invasion and EMT through the modulation of the PI3K-AKT signaling pathway, revealing NFIB as a potential biomarker for monitoring early metastasis of bladder cancer and a target for therapy.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-026-00900-4.

Drug repurposing in oncology can reduce the time and cost of new drug development. Studies suggest that depression influences tumor progression, and some antidepressants exhibit anti-tumor effects. This study evaluated the effects of serotonergic antagonists-tropisetron, imipramine, ketanserin, and cyproheptadine-on AGS gastric cancer cells. The half-maximal inhibitory concentration (IC50) values of drugs were determined after 48 h in AGS cells using the MTT assay. Cell cycle distribution and apoptosis were evaluated by flow cytometry. Morphological changes were observed by Acridine Orange/Ethidium Bromide staining. The wound-healing assay was used to assess the effects of the drugs on cell migration. Real-time PCR was used to measure the expression of 5-hydroxytryptamine (5-HT) receptors (5-HT2A, 5-HT2B, 5-HT3A), serotonin transporter (SLC6A4/SERT), apoptosis-related genes (Bcl-2, Bax), and proliferating cell nuclear antigen (PCNA). All drugs significantly inhibited the growth of AGS cell in vitro. All four drugs induced apoptosis and inhibited cell migration with varying efficacies. Imipramine induced G1/S phase arrest, whereas tropisetron, ketanserin, and cyproheptadine increased the sub-G1 cell population. Gene expression analysis revealed decreased Bcl-2 and PCNA levels and increased Bax expression.These findings suggest the potential of tropisetron, imipramine, ketanserin, and cyproheptadine as repurposed therapeutic agents for gastric cancer.

Human adipose tissue-derived stem cells (hADSCs) are an attractive source for regenerative medicine. However, cryopreservation protocols-particularly with respect to optimal cell concentration-remain inadequately defined. hADSCs were isolated from adipose tissue of 12 donors (mean age: 31.8 ± 8.9 years; BMI: 22.9 ± 4.2). Second-passage cells were cryopreserved for two weeks at concentrations of 0.5 × 10⁶/mL, 1 × 10⁶/mL, 2 × 10⁶/mL, 5 × 10⁶/mL, and 10 × 10⁶/mL. Post-thaw viability, apoptosis, immunophenotype, proliferation, and tri-lineage differentiation were evaluated using standard assays. Cell viability increased significantly with higher cryopreservation concentrations, reaching 94.2 ± 2.0% at 10 × 10⁶/mL (p < 0.05 vs. 0.5 × 10⁶/mL). Early apoptosis decreased with increasing concentration, reaching its lowest level at 5 × 10⁶/mL (2.9 ± 0.5%; p < 0.05 vs. 10 × 10⁶/mL), but showed a slight increase at 10 × 10⁶/mL. Proliferation and tri-lineage differentiation into adipocytes, osteoblasts, and chondrocytes were maintained across all groups, as confirmed by histological staining and molecular analyses. Cryopreservation at 5 × 10⁶/mL offers the most favorable balance between high viability and minimal apoptosis while preserving proliferative and differentiation potential. This concentration likely represents an optimal condition for hADSC biobanking and clinical use.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00877-6.

Lung cancer is one of the most frequent cancers in the world and the main cause of cancer related deaths. Among them, non-small cell lung cancer (NSCLC) accounts for about 85% of lung cancer. Myristicin (1-allyl-5-methoxy-3,4-methylenedioxybenzene), an active aromatic compound, has been proved to have anti-cancer effects. However, the effects of myristicin on NSCLC are not fully illustrated. Our research aimed to elucidate the roles and explain the potential mechanism of myristicin in NSCLC. A549 and H1975 cells were exposed to 0.5, 1, 5mM myristicin for 48 h, EdU, flow cytometry, Transwell and wound healing migration assay were applied for analyzing cell proliferation, apoptosis, cycle, migration and invasion, respectively. Caspase 3 activity and cleaved-Caspase3 expression were determined by relevant kits and western blotting, respectively. Besides, the Epithelial-Mesenchymal Transition (EMT) related genes levels and Wnt/β-catenin pathway related genes expressions, including E-cadherin, N-cadherin, Wnt3a and β-catenin, were assessed by qRT-PCR, and western blot assays. The nuclear translocation of β-catenin in NSCLC cells was analyzed by immunofluorescence staining. Our data revealed that myristicin suppressed NSCLC proliferation, migration and invasion in a dose-dependent manner. Besides, myristicin led to cell apoptotic and G0/G1 arrest and enhancing Caspase3 activity in NSCLC. Moreover, myristicin inhibited NSCLC EMT and blocked Wnt/β-Catenin signaling pathway in a dose-dependent manner, as confirmed by enhanced E-cadherin expression, suppressed N-cadherin level, inhibited Wnt3a and β-catenin levels, and reduced nuclear translocation of β-catenin. Myristicin blocked the development of NSCLC via regulating cells proliferation, migration, invasion and EMT through deactivating the Wnt/β-catenin pathway, which provide a new therapeutic treatment for NSCLC in clinical.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-026-00893-0.

Glycation of low-density lipoprotein (LDL) occurs when blood glucose levels are high, as in diabetic patients. This can lead to abnormal cholesterol transport in the body. Glycated LDL (G-LDL) is harmful and triggers oxidative stress in human erythrocytes. The present study investigated the protective effect of crocin, found in saffron, against G-LDL-induced cytotoxicity, oxidative damage, and formation of reactive species in human erythrocytes. These parameters were assessed in isolated human erythrocytes exposed to 6 mg/ml G-LDL, with and without varying concentrations of crocin (0.5, 1.0, 1.5 mM). Increased hemolysis, methemoglobin, and oxoferrylHb were seen in G-LDL alone-incubated cells. A significant increase in reactive species in G-LDL-exposed erythrocytes led to enhanced oxidation of lipids, proteins, and thiols. The activities of certain key antioxidant and membrane-bound enzymes were reduced. The antioxidant capacity of cells was compromised as indicated by a diminished ability to neutralize free radicals and donate electrons. G-LDL significantly altered surface morphology, forming echinocytes and agglutinating the cells. All these characteristics were significantly restored when erythrocytes were pre-treated with crocin, before incubation with G-LDL, in a crocin concentration-dependent manner. Furthermore, erythrocytes incubated with 1.5 mM crocin alone did not show alterations in any of the above parameters, indicating that crocin was not toxic to these cells. These results clearly show that crocin is strongly cytoprotective against G-LDL-induced damage and toxicity in erythrocytes. Hence, it can be used as an efficient dietary antioxidant in various atherosclerotic cardiovascular disorders, as seen in diabetic patients.

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