Cytotechnology最新文献

Inflammatory bowel disease (IBD) is a chronic, progressive, immune-mediated, gastrointestinal inflammatory disease with increasing occurrences in children. Collagen triple helix repeat containing 1 (CTHRC1), a migration-promoting protein, acts as a tumor-promoting factor in malignant tumors. However, functions and mechanisms of CTHRC1 in children with IBD remain unclear. This study aimed to determine the effects and mechanisms of CTHRC1 on dextran sodium sulfate (DSS)-treated HT-29 cells. HT-29 control cells were exposed to 2% DSS to develop an in vitro IBD model. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) and western blotting were used to assess CTHRC1 expression in serum of children with IBD and HT-29 cells. Cell viability and apoptosis were assessed using MTT and flow cytometry (FCM). Expressions of cleaved-Caspase3 and Caspase3 were determined by western blotting. The cytokine production (TNF-α, IL-1β and IL-6) in HT-29 cells was measured by ELISA assay. Activation or inactivation of NF-κB signaling pathway was confirmed by western blot assay. Results showed that CTHRC1 expression was upregulated in the IBD serum and HT-29 control cells. The level of CTHRC1 was lower in CTHRC1-siRNA transfected cells than in control siRNA-treated cells. Notably, silence of CTHRC1 markedly enhanced HT-29 cells viability, decreased apoptotic cells, suppressed cleaved-Caspase3 expression, inhibited cleaved-Caspase3/Caspase3 ratio, reduced the production of inflammatory cytokines, and blocked NF-κB signaling pathway induced by DSS. However, these effects were reversed following diprovocim treatment. Thus, that knockdown of CTHRC1 alleviated DSS-induced HT-29 cell injury by inhibiting the NF-κB signaling pathway in vitro, providing a new therapeutic target for IBD in children.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00705-x.

The long-term treatment of malignancies, particularly brain tumors, is challenged by abnormal protein expression and drug resistance. In terms of potency, selectivity, and overcoming drug resistance, Proteolysis Targeting Chimeras (PROTACs), a cutting-edge method used to selectively degrade target proteins, beats traditional inhibitors. This review summarizes recent research on using PROTACs as a therapeutic strategy for brain tumors, focusing on their mechanism, benefits, limitations, and the need for optimization. The review draws from a comprehensive search of peer-reviewed literature, scientific databases, and clinical trial databases. Articles published up to the knowledge cutoff date up to 14 April 2023 were included. Inclusion criteria covered PROTAC-based brain tumor therapies, including preclinical and early clinical studies, with no restrictions on design or publication type. We included studies using in vitro, in vivo brain tumor models, and human subjects. Eligible treatments involved PROTACs targeting proteins linked to brain tumor progression. We evaluated the selected studies for methodology, including design, sample size, and data analysis techniques. A narrative synthesis summarized key outcomes and trends in PROTAC-based brain tumor therapy. Recent research shows PROTACs selectively degrade brain tumor-related proteins with minimal off-target effects. They offer enhanced potency, selectivity, and the ability to combat resistance compared to traditional inhibitors. PROTACs hold promise for brain tumor treatment offering advantages over traditional inhibitors, but more research is needed to refine their mechanisms, efficacy, and safety. Larger-scale trials and translational studies are essential for assessing their clinical utility.

Intervertebral disc degeneration (IVDD) represents a major cause of lower back pain, whose prevalence rises with age. This study probed into the mechanism of microRNA (miR)-124-3p regulating function of nucleus pulposus cells (NPCs) by targeting calpain-1 (CAPN1). Rat IVD NPCs were cultured in vitro and transfected with miR-124-3p mimics, miR-124-3p inhibitor, oe-CAPN1 and their negative controls. The mRNA levels of miR-124-3p and CAPN1 were assessed by RT-qPCR. Cell proliferation, apoptosis and migration were evaluated by CCK-8, flow cytometry and Transwell assays. Levels of CAPN1 protein, apoptosis-related proteins (BAX, Cleaved-Caspase3, BCL-2) and extracellular matrix (ECM) proteins (Collagen II, Aggrecan, Fibronectin, Collagen I, matrix metalloproteinase [MMP]-13) were determined by Western blot. The target binding relationship between miR-124-3p and CAPN1 was verified by dual-luciferase assay. miR-124-3p overexpression facilitated NPC function and the maintenance of ECM homeostasis, as evidenced by increased NPC proliferation and migration, decreased apoptosis, elevated apoptosis-related protein BCL-2 level, diminished BAX and Cleaved-Caspase3 levels, reduced levels of ECM homeostasis-associated factors Collagen I and MMP-13 proteins, as well as raised levels of Collagen II, Aggrecan and Fibronectin proteins. Conversely, miR-124-3p knockdown brought about the opposite results. miR-124-3p targeted CAPN1. Furthermore, overexpression of CAPN1 partially reversed the regulatory effects of miR-124-3p on the ECM homeostasis, proliferation and migration in NPCs, and promoted apoptosis. miR-124-3p contributed to proliferation and migration of IVD NPCs, and reduced their apoptosis by inhibiting CAPN1 expression, thereby modulating ECM homeostasis and maintaining the function of IVD NPCs.

Cancer-associated fibroblasts (CAFs) have been shown to play a crucial role in the progression of non-small cell lung cancer (NSCLC). Exosomes derived from CAFs have emerged as important mediators of intercellular communication in the tumor microenvironment, contributing to cancer progression. Therefore, it is essential to further investigate the mechanisms by which CAF-derived exosomes regulate NSCLC. CAFs promoted NSCLC cell proliferation, invasion, and migration, while also suppressing radiosensitivity. We observed an upregulation of FAM83F expression in both NSCLC cells and NSCLC cells treated with conditioned medium from CAFs. Notably, CAF-derived exosomes were found to transfer FAM83F to NSCLC cells, thereby enhancing the malignant properties of the cancer cells. In contrast, FAM83F-deficient CAF-derived exosomes exerted inhibitory effects on NSCLC cell proliferation, invasion, and migration, while also sensitizing the cells to radiotherapy. FAM83F was found to interact with KIF23 in NSCLC cells, and the overexpression of KIF23 attenuated the effects induced by FAM83F-deficient exosomes in NSCLC cells. Moreover, FAM83F-deficient CAF-derived exosomes were effective in inhibiting tumor formation in vivo. Our findings highlight the crucial role of CAF-derived exosomal FAM83F in promoting NSCLC progression and conferring resistance to radiotherapy. Targeting this signaling pathway may offer promising therapeutic strategies for combating NSCLC progression and improving patient outcomes.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00713-x.

Targeting tumor angiogenesis with safe endogenous protein inhibitors is a promising therapeutic approach despite the plethora of the first line of emerging chemotherapeutic drugs. The extracellular matrix network in the blood vessel basement membrane and growth factors released from endothelial and tumor cells promote the neovascularization which supports the tumor growth. Contrastingly, small cleaved cryptic fragments of the C-terminal non collagenous domains of the same basement membrane display antiangiogenic effect. In the present study, full length α6(IV)NC1(Hexastatin) and its three subfragments α6S1(IV)NC1, α6S2(IV)NC1, and α6S3(IV)NC1 were validated for their pro-apoptotic and angio-inhibitory property. In order to construct the coding sequence of hexastatin and its three derivative partial peptide fragments were constructed with our proposed method, where the corresponding exons were amplified from the genomic DNA and then assembled together. Coding sequences were cloned and expressed using pLATE31 vector and recombinant proteins were purified with C-terminal His tag. The endogenous NC protein fragments of collagen IV were evaluated in vitro for their role in cytotoxicity on human umbilical vein endothelial cells (HUVECs). The results showed that the NC1 domain and its fragments inhibited the HUVECs cell proliferation, migration, invasion and induced apoptosis. The neovascularization inhibition was studied in in-vitro, via tube formation assay and in-vivo via the CAM Assay. The results showed that blood vessels and inter capillary network were inhibited in endothelial cells and also, in chick embryo treated with recombinant α6(IV)NC1 and its derivatives, except for α6S1(IV)NC1 and these endogenous protein inhibitors act as bio-therapeutics in inhibition of angiogenesis.

Because acute kidney injuries (AKI) are one of the critical health problems worldwide, studies on the risk factors, mechanisms, and treatment strategies seem necessary. Glycerol (GLY), known to induce cell necrosis via myoglobin accumulation in renal tubules, is widely used as an AKI model. This study aimed to evaluate the protective effects of gallic acid (GA) against GLY-induced AKI. The study utilized both in vivo and in vitro models. In vivo, healthy rats were divided into six groups: control (normal saline), GLY (10 mg/kg, intramuscularly), GLY + GA10 (10 mg/kg), GLY + GA50 (50 mg/kg), GLY + GA100 (100 mg/kg), and GA (100 mg/kg). GA was administered by gavage for seven consecutive days, followed by a single intramuscular injection of GLY. Kidney biomarkers, lactate dehydrogenase (LDH), oxidative stress markers, inflammatory indices, and histological parameters were assessed 72 h post-injection. In vitro, human embryonic kidney 2 (HK-2) cells were incubated with GLY and GA at different concentrations (30, 60, and 125 μg/ml) to evaluate cell viability, reactive oxygen species (ROS) production, oxidative stress, and inflammatory cytokines. GLY administration significantly elevated renal dysfunction markers, including blood urea nitrogen and creatinine, alongside oxidative stress and reduced cell viability. GA treatment improved kidney biomarkers, enhanced antioxidant enzyme activity, and reduced inflammatory cytokines. Histological analyses also showed improved kidney structural integrity in GA-treated rats compared to the GLY group. This study confirmed that GLY induces AKI through oxidative stress, inflammation, and structural damage. GA exhibited significant renal protective effects by enhancing antioxidant defenses and reducing inflammation. These findings support GA as a potential natural supplement for preventing or treating renal diseases.

Inflammatory bowel disease (IBD), a chronic gastrointestinal disorder, often emerges during childhood and poses significant challenges due to its adverse effects on growth, development, and psychosocial well-being. Circular RNAs (circRNAs) have been implicated in the pathogenesis of diverse diseases. However, the specific biological role and mechanisms of circRNA OMA1 in children with IBD remain largely unexplored. This study investigates the functions and mechanistic pathways of circRNA OMA1 in the progression of IBD. Quantitative real-time PCR (qRT-PCR) was employed to quantify circRNA OMA1 and miR-654-3p expression levels in the serum of children with IBD and in HT-29 cells. Downstream miRNA and mRNA targets of circRNA OMA1 were predicted using StarBase and validated via luciferase reporter assays. An in vitro IBD model was established by treating the human colonic epithelial cell line (HT-29) with 2% dextran sulfate sodium (DSS). Cell viability and apoptosis were assessed using the MTT assay and flow cytometry, respectively. Expression of the apoptosis-related protein cleaved caspase-3 was analyzed via western blotting, and proinflammatory cytokine levels (TNF-α, IL-1β, and IL-6) were measured using ELISA. The expression of circRNA OMA1 was notably lower in the serum of children with IBD and in DSS-treated HT-29 cells than in healthy controls, whereas miR-654-3p expression was upregulated. Bioinformatics analyses revealed a direct interaction between circRNA OMA1 and miR-654-3p. Overexpression of circRNA OMA1 through plasmid transfection increased circRNA OMA1 levels and suppressed miR-654-3p expression in HT-29 cells under both basal and DSS-stimulated conditions. Conversely, transfection with a miR-654-3p mimic reversed these effects. Upregulation of circRNA OMA1 ameliorated DSS-induced injury in HT-29 cells by enhancing cell viability, reducing apoptosis, and downregulating cleaved caspase-3 expression. Moreover, circRNA OMA1 overexpression inhibited the secretion of inflammatory cytokines TNF-α, IL-1β, and IL-6. However, these protective effects were partially reversed by treatment with the miR-654-3p mimic. Additionally, miR-654-3p was shown to directly target RAF1, negatively regulating its expression. The proliferation-promoting and apoptosis-suppressing effects of miR-654-3p inhibitor treatment were mitigated by RAF1-siRNA. Conclusion: Upregulation of circRNA OMA1 alleviates DSS-induced colonic cell apoptosis and inflammation by modulating the miR-654-3p/RAF1 axis. These findings suggest that circRNA OMA1 could be a promising biomarker for the diagnosis and treatment of IBD.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00703-z.

High expression of Fascin-1 involves high metastasis, high recurrence, and poor prognosis of cancers. However, the related regulatory mechanism in hepatocellular carcinoma (HCC) remains elusive. In this study, Fascin-1 was highly expressed in HCC tissues and cell lines. Fastin-1 protein levels and p-Akt1/Akt1 rate were increased by Akt activator SC79 and were decreased by Akt inhibitor LY294002. Silenced Fascin-1 suppressed cell proliferation, promoted cell apoptosis, suppressed cell invasion and epithelial-mesenchymal transition (EMT) in HCC cell lines. Also, silenced Fascin-1 induced cell cycle arrest in the G1 phase. Moreover, silenced Fascin-1 repressed invasion of HCC cells by inhibiting EMT. Besides, interference with Fascin-1 inhibited HCC cell growth, reduced Vimentin expressions and p-Akt1/Akt1 rate in vivo, while these impacts were abolished after injection of SC79. In conclusion, silencing Fascin-1 reduced the malignant growth of HCC, and this process was closely related to AKT inactivation.

Supplementary information: The online version contains supplementary material available at 10.1007/s10616-025-00707-9.

Burn injuries are complex, life-threatening events involving intricate cellular and molecular processes, including angiogenesis, which is vital for effective wound healing. Bletilla striata polysaccharide (BSP), a bioactive compound from Bletilla striata, exhibits anti-inflammatory and wound-healing properties. However, its impact on angiogenesis modulation, particularly through the synaptopodin-2-like (SCEL) gene, remains poorly understood. The effects of BSP on HMEC-1 cells exposed to lipopolysaccharide (LPS) were assessed using cell viability, migration, apoptosis, and angiogenesis assays. SCEL's role was explored through lentiviral transfection to manipulate SCEL expression. Animal models were employed to evaluate BSP's therapeutic potential in burn wound healing, with histological analysis, immunohistochemistry (IHC), and molecular assays to assess tissue repair and angiogenesis. BSP significantly alleviated LPS-induced damage in HMEC-1 cells by promoting cell survival, reducing apoptosis, and enhancing migration and angiogenesis. BSP treatment downregulated SCEL expression, reversing LPS-induced cellular damage. In SCEL-overexpressing cells and mice, BSP's beneficial effects on wound healing were attenuated, indicating SCEL's regulatory role in angiogenesis. In vivo, BSP accelerated burn wound closure, improved tissue organization, and enhanced angiogenesis, as evidenced by increased CD31 expression. SCEL overexpression impaired these effects, highlighting the essential role of SCEL downregulation in BSP-mediated healing. BSP promotes burn wound healing by modulating angiogenesis via SCEL downregulation, facilitating cell survival, migration, and vascularization. These findings position BSP as a promising therapeutic agent for burn wound treatment, with further investigation into SCEL's molecular mechanisms offering potential for novel wound care strategies.

This study aimed to investigate the role of circular RNAs (circRNAs) in sepsis-induced acute gastrointestinal injury (AGI), focusing on their potential as biomarkers and their involvement in disease progression. Peripheral blood samples from 14 patients with sepsis-induced AGI and healthy volunteers were collected. RNA sequencing was performed to profile circRNA and miRNA expression. Differential expression analysis identified key regulatory RNAs. Functional enrichment analysis was conducted to explore biological pathways, and circRNA-miRNA interaction networks were constructed. Significant differences in circRNA and miRNA expression profiles were observed between sepsis-induced AGI patients and healthy controls. Several circRNAs, including hsa_circ_0008381 and hsa_circ_0071375, exhibited stepwise expression increases correlating with AGI severity. Functional enrichment analysis indicated that the host genes of differentially expressed circRNAs are involved in key biological processes like protein ubiquitination, organelle maintenance, and cellular signaling pathways such as mitochondrial biogenesis and lipid metabolism. CircRNA-miRNA interaction networks suggested their role as miRNA sponges, regulating key downstream processes. This study demonstrated the potential of circRNAs as diagnostic biomarkers and therapeutic targets for sepsis-induced AGI. Further research is warranted to validate their clinical utility and unravel their mechanistic roles in AGI progression.