Cytotechnology最新文献

Marine-derived functional ingredients are gaining attention for their therapeutic potential in skin health and inflammation. The wound healing and anti-inflammatory properties of Holothuria atra, an edible sea cucumber, remain underexplored. This study aimed to evaluate the in vitro wound healing activity of H. atra methanol extract and the in silico anti-tumor necrosis factor-α (TNF-α) of its metabolites. Cytotoxicity on human keratinocyte (HaCaT) cells was assessed, followed by scratch assays under normal and TNF-α-induced conditions. The components of the extract were identified based on their molecular masses, and key metabolites were docked against TNF-α binding sites. The extract showed 83.8 ± 2.5% cell viability at 100 ng/mL, which was used as the maximum concentration. In the scratch assay, wound closure increased with dose, reaching 53.2 ± 8.8% after 48 h at 100 ng/mL, compared to 18.4 ± 6.9% in the control (p < 0.0001). With TNF-α (10 ng/mL), cell migration dropped (1.0 ± 0.2% after 48 h); however, 100 ng/mL extract restored closure to 53.2 ± 15.1% (p < 0.001) after 48 h. GC-MS revealed major fatty acids, including arachidonic (17.5%) and palmitic acid (15.2%), while LC-MS identified ten saponins, six verified by standards. Docking showed metabolite binding to TNF-α (-9.29 to - 7.47 kcal/mol), particularly at Tyr119. In conclusion, H. atra methanol extract enhanced keratinocyte wound healing in vitro, and its metabolites showed TNF-α binding ability based on the molecular docking study. However, further in vivo validation is warranted.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00880-x.

Knee osteoarthritis (OA) is a prevalent degenerative joint disease and a leading cause of disability worldwide. While traditionally considered a mechanical disorder, accumulating evidence highlights the critical role of dysregulated immune responses in OA pathogenesis. Despite this, specific immune-related biomarkers for early diagnosis remain poorly defined. This study aims to systematically identify and validate immune-related molecular signatures involved in knee OA, providing potential diagnostic biomarkers and therapeutic targets. We applied integrated bioinformatics and machine-learning approaches to identify immune-related biomarkers specific to OA and validated their expression through in vitro experiments. Two immune-related hub genes, BCL6 and HMGB2, were found to be significantly downregulated in OA cartilage. Both demonstrated strong diagnostic performance and were consistently validated across external datasets and in vitro assays. This study identifies BCL6 and HMGB2 as robust immune-related biomarkers in knee OA, providing new insights into disease mechanisms and potential avenues for early diagnosis and therapeutic intervention.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00876-7.

This study was to investigate the therapeutic effect of Panax notoginseng-Bletilla striata (PN-BS) in reflux esophagitis (RE) and its molecular mechanism. Using the '4.2 mm pyloric clamp + 2/3 fundoplication' method, a rat model of RE was developed. RE cell model was established by exposing HET-1 A (esophageal epithelial cells) to bile salt. Esophageal mucosal injury was observed by HE staining, and epithelial barrier dysfunction was assessed using Toluidine blue staining. HET-1 A cell viability was measured by CCK-8. Inflammatory factors in tissues and cells were detected by enzyme-linked immunosorbent assay. Claudin-4, Claudin-5, NLRP3, cleaved-caspase-1, p-p38 MAPK, and p38 MAPK protein levels were detected by Western blot. PN-BS attenuated esophageal mucosal injury and inflammation and improved esophageal barrier dysfunction in RE rats. Panax notoginseng saponins (PNS, the main active ingredient of PN) and Bletilla striata polysaccharides (BSP, the main active ingredient of BS) attenuated acid and bile salt-induced esophageal barrier dysfunction. PNS and BSP inhibited NLRP3 inflammasomes and p38 MAPK pathway activation. An inhibitor of NLRP3 inflammasomes (MCC950) or an inhibitor of the p38 MAPK pathway (SB203580) further enhanced the ameliorative effects of PNS and BSP. PN-BS reduces esophageal barrier dysfunction by inhibiting the activation of NLRP3 inflammasomes and p38 MAPK pathway, thereby improving RE.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00858-9.

The imbalance of T helper 17 (Th17)/regulatory T (Treg) is pivotal in the development of chronic obstructive pulmonary disease (COPD). This study focused on evaluating the regulatory effects of short-chain fatty acids (SCFAs) on Th17/Treg balance in COPD. A COPD mouse model was induced by exposure to cigarette smoke (CS) and intranasal lipopolysaccharide (LPS) instillation. Mice were administered sodium acetate (NaA), sodium propionate (NaP), or sodium butyrate (NaB) via oral gavage. Lung function, histopathological changes, cytokine levels, and the distribution of Th17 and Treg cells were then evaluated. Epithelial cell injury induced by cigarette smoke extract (CSE) was assessed using the Cell Counting Kit-8 (CCK-8) assay, wound healing assay, and flow cytometry for apoptosis and T cell subset analysis. SCFAs treatment attenuated pathological damage in lung tissue, including reduced inflammatory cell infiltration and pulmonary fibrosis. SCFAs suppressed apoptosis and increased peak inspiratory flow (PIF) and peak expiratory flow (PEF). SCFAs effectively rebalanced the Th17/Treg axis by suppressing Th17 differentiation while promoting Treg cell expansion, which was accompanied by reduced levels of inflammatory signaling factors (IL-17, IL-6, IL-2, and TNF-α). Moreover, SCFAs enhanced CSE-induced MLE-12 cell viability and migration, and suppressed apoptosis. SCFAs blocked the differentiation of naïve CD4⁺ T cells into Th17 cells while facilitating their differentiation into Treg cells. SCFAs alleviate the pathological features of COPD by restoring the Th17/Treg balance and enhancing epithelial resilience, suggesting their promise as a therapeutic strategy for CS-related COPD.

The aggressive phenotype of colorectal cancer (CRC) is largely driven by interactions within the tumor microenvironment, specifically the co-occurrence of inflammation and hypoxic stress. While the pro-metastatic enzyme Kallikrein-related Peptidase 4 (KLK4) is known to contribute to dissemination, the precise molecular mechanism by which IL-6 and hypoxia converge to regulate KLK4 expression and subsequent metastatic potential remains to be elucidated. This study investigated the influence of the IL-6 cytokine on KLK4 gene expression and metastatic potential in the HT-29 colon cancer cell line under both normal and hypoxic conditions. Healthy non-cancerous endothelial cells (HUVEC) served as a comparative control. Expression was assessed via Real-Time PCR (mRNA) and Western blot (protein), while metastatic potential was determined by the scratch assay. Our findings demonstrate a substantial and marked upregulation in KLK4 gene and protein expression in HT-29 cells over a 48-hour period in response to IL-6, hypoxia, and the combined treatments. This increase in KLK4 was found to be associated with simultaneous upregulation of STAT-3 and p-STAT-3 proteins, strongly suggesting that the STAT-3 signaling pathway mediates this induction. The effects observed were tumor-specific: the non-cancerous HUVEC line showed only transient KLK4 changes and decreased proliferation in individual treatments. In sharp contrast, the combined IL-6 and hypoxia treatments significantly enhanced proliferative activity and metastatic potential in HT-29 cells. Western blot analysis collectively indicates that the augmented KLK4 expression in CRC cells is likely mediated through IL-6 and hypoxia-induced STAT-3 activation. These findings establish KLK4 as a potential downstream effector of the IL-6/STAT-3 pathway, offering a novel therapeutic target for mitigating metastatic potential in colon cancer.

Derived from Rehmanniae Radix, Rehmannioside A (ReA) appears to provide a defensive effect against diseases. This study was designed to investigate ReA's role and mechanism in cerebral ischemia-reperfusion injury (CI/RI). A transient middle cerebral artery occlusion (tMCAO) mouse model was developed. tMCAO mice were intraperitoneally injected with different doses of ReA for 3 days, and then underwent neurological function examination. Then, brain histopathology was observed by HE staining, neuronal apoptosis was observed by TUNEL staining, glial fibrillary acidic protein (GFAP) expression was detected by immunofluorescence staining to assess astrocyte activation, and blood-brain barrier (BBB) integrity was assessed by determining hemoglobin content and brain water content. p38 MAPK pathway-related proteins were detected by Western blot. Treatment with ReA in tMCAO mice showed a dose-dependent reduction in BBB damage, improvements in neurological function, decreased neuronal apoptosis, and inhibition of astrocyte activation. ReA inhibited p38 MAPK pathway activation, and the p38 MAPK pathway inhibitor SB203580 potentiated the ameliorative effects of ReA on CI/RI. ReA improves CI/RI by inhibiting astrocyte activation and reducing BBB damage through modulation of the p38 MAPK pathway.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00859-8.

This study investigates the characterization and biological effects of Dual Drug-Loaded Nanoparticles on HepG2/doxorubicin (DOX) cells, focusing on the anti-cancer ability of Doxorubicin/Curcumin-Polyethylene Glycol-Polycaprolactone Nanoparticles (DOX/Cur-PEG-PCL-NPs). DOX/Cur-PEG-PCL-NPs, along with single-drug and blank nanoparticles, were prepared and characterized by nanoparticle tracking analysis (NTA) for hydrodynamic diameter and Zeta potential. The morphology of DOX/Cur-PEG-PCL-NPs was characterized by transmission electron microscopy (TEM). Drug loading, encapsulation efficiency, and release profiles were evaluated. In HepG2/DOX cells, the DOX/Cur-NPs significantly reduced viability, migration, and invasion, while promoting apoptosis and reactive oxygen species (ROS) generation. Molecular analyses indicated downregulation of Nuclear Factor Erythroid 2-Related Factor 2 (Nrf2), NAD(P)H Quinone Dehydrogenase 1 (NQO1), Heme Oxygenase 1 (HO-1), and Multidrug Resistance Protein 1 (MDR1), and upregulation of Kelch-like ECH-associated protein 1 (Keap1), Bcl-2-associated X protein (Bax), and caspase-3. These results demonstrate that DOX/Cur-PEG-PCL-NPs enhance anticancer efficacy by modulating oxidative stress and apoptosis pathways. These results demonstrate that DOX/Cur-PEG-PCL-NPs effectively reverse chemoresistance and suppress tumor progression through modulation of the Nrf2 pathway and apoptosis induction, offering a promising strategy for targeted liver cancer therapy.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00855-y.

Apigenin (API) is a natural compound with an anti-cancer effect. This study aimed to investigate the mechanism of API against liver cancer. In vitro and in vivo, Huh7 cells and H22-induced liver orthotopic hepatoma model were constructed to assess the anti-tumor effect of API. Cell viability, apoptosis, mitochondrial membrane potential (MMP), reactive oxygen species (ROS), spleen lymphocyte differentiation, relative factors, and tumor pathological damage were determined using cell counting kit-8, flow cytometry, enzyme-linked immunosorbent assay, western blot, immunofluorescence, Hematoxylin-Eosin staining, and TUNEL. API inhibited liver cancer cell viability and promoted apoptosis both in vitro and in vivo. API increased ROS, interleukin (IL)-1β/6/8 and tumor necrosis factor (TNF)-α, expression of cleaved-Caspase-3/9, B-cell lymphoma-2 associated X protein, and phosphorylated nuclear transcription factor-kappa B proteins, while it reduced MMP of Huh7 cells. API increased interferon (IFN)-γ⁺ CD4⁺ cells (Th1) and decreased IL-4⁺ CD4⁺ cells (Th2) with increasing tumor TNF-α and IFN-γ and decreasing IL-1β and IL-4. The anti-tumor and T-cell regulating effects of API are similar to those of positive control cyclophosphamide. This study displayed that API has the potential to effectively prevent liver cancer by triggering mitochondrial apoptosis, thereby regulating the Th1/Th2 balance.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00868-7.

This study investigated whether the microrchidia family CW-type zinc finger 2(MORC2) decreases cellular response to paclitaxel by modulating glycolysis in ovarian cancer. In resistant ovarian cancer cell sublines for paclitaxel, including A2780/Taxol and SKOV3/Taxol, and parental cell lines, including A2780 and SKOV3, the differential expression of MORC2 was confirmed via qRT-PCR at the mRNA level and through western blot analysis at the protein level. shRNA interference was used to downregulate MORC2 in paclitaxel-resistant cells, and the actions of MORC2 reduction on paclitaxel resistance, cell cycle distribution, the uptake of glucose, and lactate production were investigated. As shown in A2780/Taxol and SKOV3/Taxol cells, MORC2 was highly expressed, and MORC2 protein expression was related to the time and concentration of paclitaxel. MORC2 downregulation made paclitaxel-resistant ovarian cancer cells more sensitive to paclitaxel, which indicated that more cells were arrested at the G0/G1 phase, suppressed glucose metabolism-related indicators (GLUT4, LDHA, and HK2). MORC2 knockdown reduced tumor growth and glycolysis, and enhanced paclitaxel sensitivity in vivo. These findings suggest MORC2 is upregulated in paclitaxel-resistant ovarian cancer cells, and that downregulation of MORC2 may reduce glycolysis and improve sensitivity to paclitaxel should be further examined.

This study aimed to investigate the regulatory role of chordin-like 2 (CHRDL2) in the Th17/Treg balance and its impact on the progression of osteoarthritis (OA). We evaluated the levels of CHRDL2 and Th17/Treg-related cytokines, and the proportions of Th17 and Treg cells in peripheral blood from both healthy subjects and OA patients. An OA mouse model was established by destabilization of the medial meniscus (DMM) surgery, and lentivirus-mediated overexpression and knockdown of CHRDL2 were conducted. The clinical and pathological manifestations of the mice were assessed, and knee joint cartilage damage was evaluated using histological staining. Additionally, we examined the levels of Treg/Th17-related inflammatory factors and transcription factors in peripheral blood, as well as the Treg/Th17 ratio. In both OA patients and mice, CHRDL2 expression was downregulated, with a significant increase in Th17 cell proportion and IL-17 levels, while Treg cell proportion and IL-10 levels were significantly decreased. Overexpression of CHRDL2 significantly improved the clinical and pathological manifestations in OA mice, corrected the Th17/Treg imbalance, reduced IL-17 and RORγt levels, and increased IL-10 and Foxp3 levels. However, knockdown of CHRDL2 results in the opposite effect. This study demonstrates that CHRDL2 can suppress OA progression by regulating the Th17/Treg balance and may serve as a key therapeutic target for alleviating immune dysregulation in OA.