Cytotechnology最新文献

{"title":"The oncogenic role of SPOCK1 in lung carcinoma by promoting immune evasion and its related mechanisms.","authors":"Geni Lv, Juan Chen, Dan Wei, Wenzhe Zhang, Wanhui Xie, Shuanying Yang","doi":"10.1007/s10616-025-00804-9","DOIUrl":"https://doi.org/10.1007/s10616-025-00804-9","url":null,"abstract":"<p><p>The progression of cancer is remarkable for its ability to evade the immune system. SPOCK1 plays crucial roles in lung carcinoma malignant phenotypes and CD8 + T cell infiltration. Here, we looked at how SPOCK1 drives immune evasion in lung cancer and unraveled the underlying mechanisms. Expression analyses were performed using quantitative PCR (qPCR), immunoblotting, or immunohistochemistry (IHC). Cell proliferation and viability were assessed by MTT assay. Cell apoptosis, invasion, and sphere formation were evaluated. The POU2F1-SPOCK1 relationship was analyzed by luciferase and ChIP assays. The ELAVL1-SPOCK1 relationship was verified by SPOCK1 mRNA stability analysis. In vivo validation of the POU2F1-SPOCK1 axis was performed using xenograft assays along with lentiviral rescue approach. Increased levels of SPOCK1 predicted poor clinical outcomes in lung carcinoma patients (n = 39) and were associated with PDL1 expression and the tumor mutational burden (TMB). SPOCK1 depletion suppressed the growth, invasion, and stemness of lung cancer cells. Moreover, SPOCK1 depletion increased TNF-α and IFN-γ secretion, enhanced CD8 + T cell viability, and suppressed CD8 + T cell apoptosis in vitro. Mechanistically, POU2F1 transcriptionally controlled SPOCK1 expression. SPOCK1 restoration reversed the impact of POU2F1 depletion on cancer cell malignant phenotypes and tumor immune evasion. Furthermore, ELAVL1 increased SPOCK1 mRNA stability to upregulate SPOCK1. Additionally, SPOCK1 increase rescued the growth of POU2F1-depleted A549 xenografts in vivo (n = 5 per group). Our findings demonstrate that SPOCK1 upregulation induced by POU2F1 or ELAVL1 contributes to lung carcinoma progression by sustaining cancer cell malignant phenotypes and promoting immune evasion, suggesting SPOCK1 as a potential target for lung cancer therapy.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00804-9.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"138"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12228940/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144583331","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"3D models to study therapy-induced senescence: where do we stand now?","authors":"Zahra Heydari, Alexander Malogolovkin, Olga Smirnova, Damir Lyukmanov, Alina Filimonova, Anastasia Shpichka, Massoud Vosough, Peter Timashev","doi":"10.1007/s10616-025-00774-y","DOIUrl":"https://doi.org/10.1007/s10616-025-00774-y","url":null,"abstract":"<p><p>Cellular senescence (CS) is a crucial tumor-suppressive phenomenon, inhibiting proliferation of cancerous cells. However, cancer therapies can also induce tumor cell senescence, generating senescent cells in tumoral and normal tissues. While initially beneficial, these senescent cells can paradoxically contribute to tumor recurrence, metastasis, and therapy resistance via the senescence-associated secretory phenotype (SASP). Due to the diverse and critical roles, cellular senescence could be a potential target in cancer biomedicine. To extend our understanding of therapy-induced senescence (TIS), developing experimental models is necessary. Currently TIS established models can be categorized into animal-based and laboratory models. These models are essential for advancing our knowledge of aging mechanisms and developing new treatment modalities. In vivo models of TIS have faced limitations, including poor immune system representation, oversimplified stromal complexity, and an inability to model functional vascular networks. Incorporating cutting-edge technologies such as 3D cultures, co-culturing, and tissue engineering can help researchers in creating in vitro models that closely mimic physiologically conditions. This review highlighted the current TIS challenges and advanced senotherapeutics. Finally, we discussed how to develop reliable in vitro models to better understanding TIS mechanisms.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"142"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12238474/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607754","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"S1P receptor 1 antagonist ponesimod alleviates postherpetic neuralgia in rats by normalizing spinal TRPV1 expression and inhibiting MAPK-mediated glial activation.","authors":"Meixin Liu, Yuanhong Wang","doi":"10.1007/s10616-025-00806-7","DOIUrl":"https://doi.org/10.1007/s10616-025-00806-7","url":null,"abstract":"<p><p>Postherpetic neuralgia (PHN) remains a clinically challenging neuropathic pain condition with limited treatment efficacy. This study investigated whether the S1PR1 modulator ponesimod alleviates PHN-like symptoms in a resiniferatoxin (RTX)-induced rat model and examined its mechanism of action. Rats with RTX-induced PHN received ponesimod (30 mg/kg) for 14 days, with subsets co-administered capsaicin (TRPV1 agonist) or AMG9810 (TRPV1 antagonist). Behavioral tests (mechanical allodynia, thermal hyperalgesia), spinal cord immunofluorescence (TRPV1, Iba-1, GFAP), cytokine ELISAs (TNF-α, IL-1β, IL-6), and Western blot (p38/JNK/ERK phosphorylation) were performed. Behavioral assessments revealed ponesimod significantly reversed both mechanical allodynia and thermal hyperalgesia compared to vehicle-treated controls, though not to baseline levels. Molecular analyses demonstrated ponesimod normalized spinal TRPV1 overexpression, suppressed glial activation, reduced proinflammatory cytokines (IL-1β, TNF-α, IL-6), and inhibited MAPK phosphorylation. Critically, TRPV1 agonist capsaicin co-administration abolished ponesimod's therapeutic effects by restoring pain behaviors, neuroinflammation, and p38 phosphorylation, while TRPV1 antagonist AMG9810 enhanced analgesic efficacy. These findings establish the efficacy of ponesimod against PHN-like pathology through S1PR1-mediated regulation of both neuroinflammation and TRPV1 expression, with TRPV1-p38 signaling serving as the key mechanistic axis for its therapeutic actions.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00806-7.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"141"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12238436/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144607756","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"UCHL3 regulates snail stability and promotes epithelial-mesenchymal transition in ovarian cancer.","authors":"Ziying Yang, Wen Wei, Daolin Nie, Menglei Zhang, Qiong Chen","doi":"10.1007/s10616-025-00811-w","DOIUrl":"https://doi.org/10.1007/s10616-025-00811-w","url":null,"abstract":"<p><p>Ovarian cancer (OC) is the most lethal gynecologic malignancy, characterized by high recurrence rates and resistance to platinum-based chemotherapy. Epithelial-mesenchymal transition (EMT) is central to OC progression, where the transcription factor Snail plays a pivotal role in downregulating E-cadherin and promoting invasive, mesenchymal phenotypes. While multiple deubiquitinating enzymes (DUBs) have been implicated in stabilizing oncogenic proteins, the specific function of ubiquitin C-terminal hydrolase L3 (UCHL3) in OC remains unclear. This study investigates whether UCHL3 regulates Snail stability and thereby drives EMT and OC progression. Publicly available datasets (TCGA + GTEx) were analyzed to compare UCHL3 mRNA levels across various tumors and corresponding normal tissues. In vitro, UCHL3 expression was measured by qPCR and Western blot in an immortalized ovarian epithelial cell line (IOSE80) and four OC cell lines (SKOV3, ES2, OVCAR3, and A2780). Stable knockdowns of UCHL3 were generated using shRNA in SKOV3 and A2780 cells. Proliferation was evaluated by CCK-8 and colony formation assays, while invasion and migration capabilities were assessed using Matrigel invasion and Transwell migration assays. EMT marker expression was examined by qPCR and Western blot. Co-immunoprecipitation (Co-IP) determined the interaction between UCHL3 and Snail, and the effect of UCHL3 on Snail ubiquitination was explored using immunoprecipitation in the presence of MG132. A cycloheximide chase assay confirmed Snail protein stability. UCHL3 was significantly overexpressed in OC tissues compared to normal controls. Silencing UCHL3 in OC cells markedly impaired cell proliferation, migration, and invasion. Concomitantly, knockdown of UCHL3 reversed EMT features, evidenced by increased E-cadherin and decreased N-cadherin, Vimentin, and Snail protein levels. Co-IP experiments demonstrated that UCHL3 directly interacts with Snail, and loss of UCHL3 elevated Snail ubiquitination, leading to accelerated Snail protein degradation. These findings indicate that UCHL3 deubiquitinates and stabilizes Snail, promoting OC cell invasiveness and EMT. Our study identifies UCHL3 as a critical regulator of Snail-mediated EMT in OC. By stabilizing Snail, UCHL3 fosters malignancy-associated phenotypes, including enhanced proliferation, migration, and invasion. These results underscore the potential of targeting UCHL3 as a therapeutic strategy to disrupt Snail-driven EMT and impede OC progression.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00811-w.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"152"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12274167/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144674099","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Exosomes carrying lncRNA FLG-AS1 overexpression vectors inhibit the tumorigenesis of oral squamous cell carcinoma via fat mass and obesity-associated protein-mediated m6A modification.","authors":"Xiaoyang Xia, Yan Liu","doi":"10.1007/s10616-025-00801-y","DOIUrl":"10.1007/s10616-025-00801-y","url":null,"abstract":"<p><p>Long noncoding RNAs (lncRNAs) selectively incorporated into exosomes contribute to the tumorigenesis of various cancers. However, the role of the lncRNA FLG-AS1 in oral squamous cell carcinoma (OSCC) has yet to be investigated. Hence, the current study aimed to evaluate the potential role of exosome-derived FLG-AS1 in OSCC. The expression of FLG-AS1 and fat mass and obesity-associated protein (FTO) in OSCC samples were determined via real-time quantitative polymerase chain reaction (RT-qPCR). The role of FLG-AS1 in OSCC was investigated by performing CCK8, flow cytometry, and transwell assays. Additionally, bioinformatics, methylated (m6A) RNA immunoprecipitation, and RT-qPCR were performed to evaluate the n6-methyladenosine (m6A) modification of FLG-AS1 and its underlying mechanism. Characterization of exosomal FLG-AS1 in OSCC cells was achieved through transmission electron microscopy, nanoparticle tracking analysis, Western blot analysis of CD63 and CD9, and RT-qPCR. Finally, the effects of exosomal FLG-AS1 on OSCC cell malignancy was analyzed using the aforementioned cell-based assays. Overall, our findings showed that low FLG-AS1 expression in OSCC restricted the malignant properties of OSCC cells. Furthermore, FTO reduced FLG-AS1 expression by mediating its m6A demethylation in OSCC cells. Exosomal FLG-AS1 expression was upregulated in OSCC cells, suggesting that it was packaged in the OSCC cell-derived exosomes, whereas exosome-derived FLG-AS1 overexpression vector reduced the malignant potential of the target OSCC cells. In conclusion, exosomal FLG-AS1 functions as a tumor suppressor in OSCC, while FTO can impair FLG-AS1 expression through m6A demethylation.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"133"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12214135/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144559479","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Metabolic changes in the one-carbon metabolism-related amino acids during etoposide-induced cellular senescence of neuronal cells.","authors":"Wang Jiahao, Jun Kameyama, Miyako Udono, Yoshinori Katakura","doi":"10.1007/s10616-025-00803-w","DOIUrl":"10.1007/s10616-025-00803-w","url":null,"abstract":"<p><p>A large-scale longitudinal epidemiological study by the Hisayama study revealed that the concentration of one-carbon metabolism-related amino acids in the serum changes with age and that there is a link between these fluctuations and the risk of developing dementia (Hata et al. in Am J Epidemiol 188:1637-1645, 2019; Mihara et al. in Sci Rep 12:12427, 2022). Therefore, the aim of this study was to focus on age-related changes in one-carbon metabolism-related amino acids and elucidate the regulatory basis of these changes. Treatment with etoposide, an anti-cancer drug, induced cellular senescence in SH-SY5Y cells, as indicated by increased senescence-associated β galactosidase activity and upregulated expression of senescence markers <i>p16</i> and <i>p21</i>. Liquid chromatography-mass spectrometry analysis revealed that the intracellular amino acid concentrations, particularly those involved in the one-carbon metabolism, were elevated in senescent cells, including those of methionine, S-adenosylmethionine, S-adenosylhomocysteine (SAH), homocysteine (Hcys), and related metabolites. The results of the expression analysis focused on the enzyme genes involved in Hcys metabolism and revealed that the induction of cellular senescence upregulated <i>adenosylhomocysteinase like 1/2</i> (<i>AHCYL1/L2</i>), which convert SAH to Hcys. Additionally, the genes involved in Hcys metabolism via the sulphuration pathway (<i>KYAT1/3</i> and <i>CTH</i>) were significantly upregulated. Because Hcys has been implicated in aging, further investigations focused on <i>AHCYL1/L2</i>. Gene knockdown of <i>AHCYL1/L2</i> in etoposide-treated cells reduced <i>p16</i> and <i>p21</i> expression, indicating that <i>AHCYL1/L2</i> is essential for cellular senescence induction. These findings suggest that Hcys accumulation and its metabolic enzymes play a crucial role in cellular senescence.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00803-w.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"131"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12209085/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144552555","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Transcription factor SP1 drives the malignant progression of oral squamous cell carcinoma and M2 macrophage polarization through transcription activation-mediated upregulation CLEC7A.","authors":"Zhen Xiao, Jiayi Li, Shuxia Fan, Yu Wang, Qingsong Jin","doi":"10.1007/s10616-025-00787-7","DOIUrl":"10.1007/s10616-025-00787-7","url":null,"abstract":"<p><p>Oral squamous cell carcinoma (OSCC) is the most common malignancy of the oral cavity. Previous studies have suggested that C-Type Lectin Domain Containing 7A (CLEC7A) could affect human cancer progression by regulating M2 macrophage polarization. However, the role of the molecular mechanism of CLEC7A involved in OSCC progression is poorly defined. GEPIA database was used to analyze CLEC7A and specificity protein 1 (SP1) expression, and the relationship between CLEC7A and M2-type macrophage markers (CD163 and MRC1). CLEC7A and SP1 levels were determined using real-time quantitative polymerase chain reaction (RT-qPCR). CLEC7A, SP1, RHOA, RAC1, E-cadherin, and Vimentin protein levels were detected using western blot. Cell proliferation, apoptosis, migration, and invasion were detected by Colony formation, flow cytometry, and Transwell assays. The proportion of CD11b⁺CD86⁺ positive cells was detected using flow cytometry. Binding between SP1 and CLEC7A promoter was predicted by JASPAR, and validated using Chromatin immunoprecipitation (ChIP) and dual-luciferase reporter assays. The biological role of SP1 on OSCC tumor growth was examined by the xenograft tumor model in vivo<i>.</i> CLEC7A and SP1 expression levels were increased in OSCC tissues and cell lines. Furthermore, CLEC7A deficiency could repress OSCC cell proliferation, migration, invasion, M2-type macrophage polarization, and induce cell apoptosis in vitro, as well as hinder tumor growth in vivo. At the molecular level, SP1 was a transcription factor of CLEC7A and promoted CLEC7A transcription via binding to its promoter regions. SP1-activated CLEC7A could facilitate OSCC cell malignant behaviors and M2 macrophage polarization, providing a possible therapeutic target for OSCC treatment.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00787-7.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"123"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12165943/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144301274","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Circular RNA-Sirt1 sponges miR-27b-3p to protect vascular smooth muscle cell injury during atherosclerosis through regulating the glutamine metabolism pathway.","authors":"Qian Song, Xiang Wang, Qinghua Zeng, Hui Xu, Lin Liu","doi":"10.1007/s10616-025-00759-x","DOIUrl":"10.1007/s10616-025-00759-x","url":null,"abstract":"<p><p>Atherosclerosis is a progressive pathological disorder resulting in various vital cardiovascular diseases such as myocardial infarction and stroke, leading to high mortality worldwide. Currently, the precise mechanisms of pathogenesis and progression of atherosclerosis remained unclear. Circular RNAs (circRNAs) have been implicated in vital processes of cardiovascular disease. In this study, we aimed to investigate the roles of circSirt1 in vascular smooth muscle cell (VSMC) injury during atherosclerosis. We found circSirt1 was significantly downregulated in VSMCs from atherosclerosis patients compared with those from healthy controls. Under oxidative stress, expression of circSirt1 was remarkedly suppressed in VSMCs. Notably, overexpression of circSirt1 effectively protected the oxidative stress-induced VSMC injury. On the other way, miRNA-27b-3p was high-expressed in VSMCs from atherosclerosis patients and was effectively induced under oxidative stress. Overexpression of miR-27b-3p exacerbated the oxidative stress-induced VSMC injury. From the non-coding RNA service, starBase, circSirt1 was predicted to interact with miR-27b-3p. This association was further validated by RNA pull-down and luciferase assays. We detected glutamine metabolism rate was depressed under oxidative stress and low glutamine supply rendered VSMCs more susceptible to oxidative stress. Furthermore, we identified the glutamine metabolism key enzyme, glutaminase (GLS) as a direct target of miR-27b-3p in VSMCs. miR-27b-3p blocked glutamine metabolism and promoted VSMC cell injury through direct targeting GLS. Finally, rescue experiments verified the circSirt1-protected VSMC injury was through regulating the miR-27b-3p-GLS axis that restoration of miR-27b-3p in circSirt1-overexpressed VSMCs successfully overrode the high-circSirt1-moduated miR-27b-3p and GLS expressions and the oxidative stress-induced VSMC injury. Summarily, these results unveiled vital roles and molecular mechanisms of circSirt1 in oxidative stress-induced VSMC injury during atherosclerosis by regulating the miR-27b-3p-GLS axis, indicating rescue of circSirt1 in VSMCs could be an effectively therapeutic approach to treat atherosclerosis.</p><p><strong>Supplementary information: </strong>The online version contains supplementary material available at 10.1007/s10616-025-00759-x.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"122"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12158877/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144301272","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Autoactivation of the HeLa 9903 test system during a full moon.","authors":"Daniel Krakowian, Martyna Lach, Przemysław Żemła, Dominika Gądarowska, Inga Mrzyk","doi":"10.1007/s10616-025-00817-4","DOIUrl":"10.1007/s10616-025-00817-4","url":null,"abstract":"<p><p>The HeLa 9903 cell line is used in the in vitro endocrine disruption assay (OECD 455). After treatment with an estrogen receptor agonist, the luminescence of these cells increases. However, some autoactivation of this test system may occur. The aim of this study was to try to explain the reason for the autoactivation of the HeLa 9903 cell line. Untreated cells were seeded on days with different moon phases and luminescence was measured the next day. Results from different moon phases were compared and statistically analyzed. An ELISA was also performed to check for the possible expression of 17β-estradiol. Statistical analysis revealed that the increase in luminescence in untreated cells was related to the phase of the moon. When the visibility of the moon was greater than approximately 87.7% (full moon), the likelihood of the cells autoactivating was greater. ELISA revealed that this change was not related to the production of 17β-estradiol by the cells (ligand-independent activation of the estrogen receptor). This is the first study to show a relationship between the phase of the moon and the activation of cells cultured in vitro. The results are important for planning the study using this test system.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"151"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12274154/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144674096","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Expression and localization of lysosomal-associated membrane protein 1- and perforin-based hybrid molecules in the murine cytotoxic T cell line CTLL-2.","authors":"Yuto Ueno, Rikako Yamaguchi, Yumiko Tanaka, Yasunobu Miyake, Keiko Takaki, Ayaka Nakao, Ryota Ozaki, Yuka Yokota, Itsuki Fukai, Sara Kuwada, Natsuki Fukuoka, Naoaki Okuda, Ai Nishida, Tomoo Shiba, Takao Kataoka","doi":"10.1007/s10616-025-00794-8","DOIUrl":"10.1007/s10616-025-00794-8","url":null,"abstract":"<p><p>Cytotoxic T lymphocytes (CTL) and natural killer cells harbor lytic granules as secretory lysosomes. Lytic granules contain perforin as a soluble protein that forms pores and is essential for target cell lysis. Lysosomal-associated membrane protein 1 (LAMP1) is a single transmembrane protein that is enriched in lysosomes and is also present in lytic granules. In this study, to investigate the feasibility of membrane-integral markers for lytic granules, LAMP1- and perforin-based hybrid molecules were constructed and their expression and localization were examined in the murine cytotoxic T-cell line CTLL-2. Stable CTLL-2 transfectants were established by nucleofection using linearized vectors encoding human LAMP1 (1-417) fused to enhanced green fluorescent protein (EGFP) and FLAG (LAMP1-EGFP-FLAG) and human perforin (1-555) fused to human LAMP1 (325-417), EGFP, and FLAG (Perforin-LAMP1-EGFP-FLAG). Confocal microscopy showed that LysoTracker Red colocalized with LAMP1-EGFP-FLAG and Perforin-LAMP1-EGFP-FLAG, and Perforin-LAMP1-EGFP-FLAG showed a broader cytoplasmic distribution than LAMP1-EGFP-FLAG. A Percoll density gradient centrifugation analysis revealed that LAMP1-EGFP-FLAG and Perforin-LAMP1-EGFP-FLAG were distributed in fractions associated with the endogenous expression of perforin and LAMP1. The effects of glycosylation inhibitors on the expression and intracellular transport of LAMP1-EGFP-FLAG were also investigated. Concanamycin A, an inhibitor of vacuolar-type H⁺-ATPase, was unable to induce the proteolytic degradation of Perforin-LAMP1-EGFP-FLAG, in contrast to perforin. The present results demonstrated that some LAMP1-EGFP-FLAG and Perforin-LAMP1-EGFP-FLAG localized to acidic granules or dense granules containing perforin and LAMP1 in CTLL-2 cells.</p>","PeriodicalId":10890,"journal":{"name":"Cytotechnology","volume":"77 4","pages":"129"},"PeriodicalIF":2.0,"publicationDate":"2025-08-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC12198104/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"144526772","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":4,"RegionCategory":"生物学","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}