Cell-related Markers Research Articles

Exploring the significance and potential mechanisms of hippo pathway-associated genes in prognosis of glioma patients

PurposeDespite the efforts of countless researchers to develop glioma treatment strategies, the current therapeutic effect of glioma is still not ideal, and it is necessary to further explore the mechanism to guide treatment. Thus, this study aims to introduce a novel approach for predicting patient prognosis and guiding further treatment interventions.MethodsInitially, we conducted a differential gene expression analysis to identify Hippo pathway-associated genes overexpressed in tumors and determined genes correlated with prognosis. Subsequently, employing cluster analysis, we categorized samples into two groups and performed further analyses including prediction, immune cell infiltration abundance, and drug response rates. We utilized weighted gene co-expression analysis to reveal gene sets with high co-variation, delineate inter-sample gene correlation patterns, and conduct enrichment analysis. Prognostic models were built using ten machine learning algorithms combined in 101 different combinations, followed by evaluation and validation. Immune infiltration analysis, differential expression analysis of depleted T cell-related markers, drug sensitivity analysis, and exploration of pathway dysregulation were performed for different risk groups. Quality control and batch integration were performed, and single-cell data were analyzed using dimensionality reduction clustering algorithms and annotation tools to evaluate the activity of the prognostic model in malignant cells.ResultsWe conducted data filtering to identify genes overexpressed in tumors, intersecting these genes with Hippo pathway-related genes, identifying 62 genes correlated with prognosis, and performing cluster analysis to divide tumor tissues into two groups. Cluster 2 exhibited a poorer prognosis and demonstrated differences in immune cell infiltration. Utilizing weighted gene co-expression analysis on Cluster 2, we identified gene modules, conducted functional enrichment analysis, and delineated pathways. Employing a combined model based on ten machine learning algorithm combinations, we selected the optimal prognostic model system and validated the model’s predictive ability within the dataset. Through immune-related analysis and drug sensitivity analysis, we uncovered differences in immune infiltration and varying sensitivities to chemotherapy drugs. Additionally, the enrichment analysis of gene set revealed discrepancies in upregulation within relevant pathways between the high and low-risk groups. Finally, annotation and evaluation of malignant cells via single-cell analysis showed increased activity of the prognostic model and variations in distribution across different prognostic levels in malignant cells.ConclusionThis study introduces a novel approach utilizing the Hippo pathway and associated genes for glioma prognosis research, demonstrating the potential and significance of this method in evaluating the outcome for patients with glioma. These findings hold substantial clinical significance in guiding therapy and predicting outcomes for individuals diagnosed with glioma, offering significant clinical utility.

The relationship between the tertiary lymphoid structure and immune-infiltrating cells in gastrointestinal cancers: A systematic review and meta-analysis.

This study systematically evaluated the relationship between tertiary lymphoid structures (TLS) and clinical pathological features as well as immune infiltrating cells in gastrointestinal cancers. We searched Web of science, Pubmed, Embase, and Cochrane Library for studies that met the requirements as of July 1, 2023, and the odds ratio, the corresponding 95% confidence interval or mean and standard deviation, were included in the analysis. We eventually included 20 studies, involving a total of 4856 patients. TLS were found to be significantly associated with T stage, N stage, TNM stage, and tumor size. Moreover, patients with positive TLS showed significantly elevated expression of T-cell related markers, including CD3, CD4, CD8, CD45RO; B-cell related markers, such as CD11c and CD20; and dendritic cell-related marker CD103. On the other hand, positive TLS correlated significantly with low expression of FOXP3 and CD68. Additionally, there was a significant positive association between TLS and overall infiltration of tumor-infiltrating lymphocytes. The presence of TLS is significantly correlated with the infiltration of various immune cells in gastrointestinal cancers. To determine the ideal balance between the presence of mature TLS and appropriate immune cell infiltration, further high-quality and multicenter clinical studies need to be conducted.

Rapamycin Potentiates the Antitumor Effect of 5-Fluorouracil in Colon Cancer by Enhancing Autophagy

Objective: To investigate whether the mTOR inhibitor rapamycin can enhance the growth suppression effect of 5-fluorouracil (5-FU) on colon cancer cells. Methods: CCK8 assays were used to examine cell survival. Flow cytometry and Western blotting were employed to detect cell proliferation, apoptosis, and related markers. Results: The combination of rapamycin and 5-FU exhibited greater cytotoxicity in cells compared to rapamycin or 5-FU alone. Notably, cells in the G0/G1 phase increased while cells in the S phase decreased in the combination group, consistent with changes in the levels of Cyclin D1 and PCNA. Rapamycin enhanced 5-FU-induced apoptosis in vitro by inducing caspase-dependent apoptosis, which is Bax/Bcl-2 related. Conclusion: The combination of rapamycin and 5-FU showed a significant synergistic anticancer effect by enhancing autophagy. This study supports that the combination of rapamycin with 5-FU is an effective and feasible approach for colorectal cancer treatment.

Role of PI3K/AKT signaling pathway involved in self-renewing and maintaining biological properties of chicken primordial germ cells

Avian primordial germ cells (PGCs) are important culture cells for the production of transgenic chickens and preservation of the genetic resources of endangered species; however, culturing these cells in vitro proves challenging. Although the proliferation of chicken PGCs is dependent on insulin, the underlying molecular mechanisms remain unclear. In the present study, we explored the expression of the PI3K/AKT signaling pathway in PGCs, investigated its effects on PGC self-renewal and biological properties, and identified the underlying mechanisms. Our findings indicated that although supplementation with the PI3K/AKT activator IGF-1 failed to promote proliferation under the assessed culture conditions, the PI3K/AKT inhibitor LY294002 resulted in retarded cell proliferation and reduced expression of germ cell-related markers. We further demonstrated that inhibition of PI3K/AKT regulates the cell cycle and promotes apoptosis in PGCs by activating the expression of BAX and inhibiting that of Bcl-2. These findings indicated that the PI3K/AKT pathway is required for cell renewal, apoptosis, and maintenance of the reproductive potential in chicken PGCs. This study aimed to provide a theoretical basis for the optimization and improvement of a culture system for chicken PGCs and provide insights into the self-renewal of vertebrate PGCs as well as potential evolutionary changes in this unique cell population.

Dendritic Cell-Related Immune Marker CD1C for Predicting Prognosis and Immunotherapy Opportunities of Lung Adenocarcinoma Patients.

Lung adenocarcinoma (LUAD) is the most frequent type of lung cancer with a high mortality rate. Here, we aim to explore novel immune-related biomarkers for LUAD patients. Datasets, mRNA expression profiles, and clinical data concerned with LUAD were obtained from Gene Expression Omnibus (GEO) database and The Cancer Genome Atlas (TCGA), respectively. Differential expression analysis was performed to obtain differentially expressed genes (DEGs). Based on DEGs, we conducted functional enrichment analyses. Subsequently, Kaplan‑Meier (KM) was performed to analyze survival differences among different groups. Furthermore, immune cell infiltration proportion was calculated by CIBERSORT and TIMER. The relationship between gene and immune response was analyzed using Tumor Immune System Interactions (TISIDB) database. Finally, Pearson correlation analysis was performed between CD1C and six immune checkpoints. We identified dendritic cells (DCs)-related expression profiles from four LUAD samples. DCs' immune marker CD1C in LUAD was selected by univariate Cox regression analysis. Low CD1C expression patients had a poor prognosis. A total of 332 DEGs were identified in high and low CD1C expression groups, which primarily enriched in 348 GO terms and 30 KEGG pathways. There were significant differences in the infiltration proportion of 17 immune cells between high and low CD1C expression groups. Most immunomodulators, chemokines, and chemokine receptors were positively associated with CD1C expression. Six immune checkpoints were also positively correlated with CD1C expression. DCs related immunomarker CD1C probably plays a pivotal part in prognosis and immunotherapy of LUAD via a joint analysis of single-cell and bulk sequencing data.

Identification of Stem Cell-related Gene Markers by Comprehensive Transcriptome Analysis to Predict the Prognosis and Immunotherapy of Lung Adenocarcinoma.

Cancer stem cells (CSCs) contribute to metastasis and drug resistance to immunotherapy in lung adenocarcinoma (LUAD), so the stemness evaluation of cancer cells is of great significance. The single-cell RNA sequencing (scRNA-seq) data of the GSE149655 dataset were collected and analyzed. Malignant cells were distinguished by CopyKAT. CytoTRACE score of marker genes in malignant cells was counted by CytoTRACE to construct the stemness score formula. Sample stemness score in TCGA was determined by the formula and divided into high-, medium- and low-stemness score groups. LASSO and COX regression analyses were carried out to screen the key genes related to the prognosis of LUAD from the differentially expressed genes (DEGs) in high- and low-stemness score groups and a risk score model was constructed. Seven types of cells were identified from a total of 4 samples, and 193 marker genes of 3455 malignant cells were identified. There were 1098 DEGs between low- and high-stemness score groups of TCGA, of which CPS1, CENPK, GJB3, and TPSB2 constituted gene signatures. The 4-gene signature could independently evaluate LUAD survival in the training and validation sets and showed an acceptable area under the receiver operator characteristic (ROC) curves (AUCs). This study provides insights into the cellular heterogeneity of LUAD and develops a new cancer stemness evaluation indicator and a 4-gene signature as a potential tool for evaluating the response of LUAD to immune checkpoint blockade (ICB) therapy or antineoplastic therapy.

Basal differentiation and expression status of SOX2 and KLF4 in basal layers are prognostic factors for disease-specific survival in oral squamous cell carcinoma.

Basal differentiation in oral squamous cell carcinoma is usually detected at invasive sites. However, its significance as a prognostic value has been poorly investigated. COL17 was selected as a basal differentiation marker because of its stable expression in the basal-like cells of oral squamous cell carcinoma. Sixty-five cases of oral squamous cell carcinoma were subclassified into COL17-high (30 cases) and -low (35 cases) types, and the prognostic value was analyzed by Cox regression analysis. In addition, the stem cell markers such as SOX2, KLF4, MYC as well as the stem cell-related markers BMI1, EZH2, and YAP and its paralog TAZ, were immunohistochemically analyzed. Their prognostic values were investigated along with their COL17 status by Cox regression analysis. No significant difference was observed between the COL17-high and -low groups in the disease-specific survival and recurrence-free survival in oral squamous cell carcinoma. When the COL17-high and -low categories were combined with the SOX2, KLF4, EZH2, or YAP/TAZ status in the basal layers, together with gender and age as covariates, the hazard ratios reached 3.3, 3.7, 2.8, and 3.1, respectively. In addition, multivariate analysis, including COL17, SOX2, and KLF4, with gender and age as covariates, showed a significantly poor prognosis for disease-specific survival. Based on the relatively high hazard ratios, it is indicated that basal differentiation and the expression status of SOX2 and KLF4 in the basal layers are prognostic factors for oral squamous cell carcinoma.

Immune system-related plasma extracellular vesicles in healthy aging.

To identify age-related plasma extracellular vehicle (EVs) phenotypes in healthy adults. EV proteomics by high-resolution mass spectrometry to evaluate EV protein stability and discover age-associated EV proteins (n=4 with 4 serial freeze-thaws each); validation by high-resolution flow cytometry and EV cytokine quantification by multiplex ELISA (n=28 healthy donors, aged 18-83 years); quantification of WI-38 fibroblast cell proliferation response to co-culture with PKH67-labeled young and old plasma EVs. The EV samples from these plasma specimens were previously characterized for bilayer structure, intra-vesicle mitochondria and cytokines, and hematopoietic cell-related surface markers. Compared with matched exo-EVs (EV-depleted supernatants), endo-EVs (EV-associated) had higher mean TNF-α and IL-27, lower mean IL-6, IL-11, IFN-γ, and IL-17A/F, and similar mean IL-1β, IL-21, and IL-22 concentrations. Some endo-EV and exo-EV cytokine concentrations were correlated, including TNF-α, IL-27, IL-6, IL-1β, and IFN-γ, but not IL-11, IL-17A/F, IL-21 or IL-22. Endo-EV IFN-γ and exo-EV IL-17A/F and IL-21 declined with age. By proteomics and confirmed by flow cytometry, we identified age-associated decline of fibrinogen (FGA, FGB and FGG) in EVs. Age-related EV proteins indicated predominant origins in the liver and innate immune system. WI-38 cells (>95%) internalized similar amounts of young and old plasma EVs, but cells that internalized PKH67-EVs, particularly young EVs, underwent significantly greater cell proliferation. Endo-EV and exo-EV cytokines function as different biomarkers. The observed healthy aging EV phenotype reflected a downregulation of EV fibrinogen subpopulations consistent with the absence of a pro-coagulant and pro-inflammatory condition common with age-related disease.

Machine learning constructs a T cell-related signature for predicting prognosis and drug sensitivity in ovarian cancer.

The leading cause of death related to gynecologic cancer is ovarian cancer, which typically has a poor prognosis. T cells are referred to as key mediators of immunosurveillance and tumor eradication, and unbalanced regulation or lack of T cells in tumors result in immunotherapy resistance. The identification of T cell related markers depended on single-cell RNA-seq analysis. Using data from multiple datasets, including TCGA, GSE14764, GSE26193, GSE26712, and GSE140082, we constructed a prognostic signature called TRS (T cell-related signature) using 10 different machine learning algorithms. The correlation between TRS and drug sensitivity were analyzed using the data from GSE91061 and IMvigor210 dataset. PlsRcox method based TRS was as a risk factor for the clinical outcome of ovarian cancer patients. In comparison with stage, grade and many prognostic signatures, the performance of our TRS in evaluating the clinical outcome was better in ovarian cancer. TRS-based risk score showed distinct association with the level of ESTIMATE score, immune-related function score and immune cells. Moreover, TRS could be used to predict the immunotherapy response and chemotherapy response in ovarian cancer. In conclusion, we constructed a powerful TRS in ovarian cancer, which could accurately predict the clinical outcome of patients and be used to predict the immunotherapy response and chemotherapy response.

ChemR23 signaling ameliorates brain injury via inhibiting NLRP3 inflammasome-mediated neuronal pyroptosis in ischemic stroke

BackgroundInflammatory response has been recognized as a pivotal pathophysiological process during cerebral ischemia. ChemR23 signaling is involved in the pathophysiology of various inflammatory diseases. Nevertheless, the role of ChemR23 signaling in ischemic stroke remains largely unknown.MethodsPermanent ischemic stroke mouse model was accomplished by middle cerebral artery occlusion (MCAO). Resolvin E1 (RvE1) or chemerin-9 (C-9), the agonists of ChemR23, were administered by intracerebroventricular (i.c.v) injection before MCAO induction. Then, analysis of neurobehavioral deficits and brain sampling were done at Day 1 after MCAO. The brain samples were further analyzed by histological staining, immunofluorescence, RNA sequencing, ELISA, transmission electron microscope, and western blots. Furthermore, oxygen–glucose deprivation (OGD) was employed in SH-SY5Y to mimic MCAO in vitro, and ChemR23 signaling pathway was further studied by overexpression of ChemR23 or administration of related agonists or antagonists. Analysis of cell death and related pathway markers were performed.ResultsChemR23 expression was upregulated following MCAO. Under in vitro and in vivo ischemic conditions, ChemR23 deficiency or inhibition contributed to excessive NLRP3-mediated maturation and release of IL-1β and IL-18, as well as enhanced cleavage of GSDMD-N and neuronal pyroptosis. These influences ultimately aggravated brain injury and neuronal damage. On the other hand, ChemR23 activation by RvE1 or C-9 mitigated the above pathophysiological abnormalities in vivo and in vitro, and overexpression of ChemR23 in SH-SY5Y cells also rescued OGD-induced neuronal pyroptosis. Blockade of NLRP3 mimics the protective effects of ChemR23 activation in vitro.ConclusionOur data indicated that ChemR23 modulates NLRP3 inflammasome-mediated neuronal pyroptosis in ischemic stroke. Activation of ChemR23 may serve as a promising potential target for neuroprotection in cerebral ischemia.

A sequential algorithm for CSF-testing using new and conventional B cell-related markers for the diagnosis of multiple sclerosis

A sequential algorithm for CSF-testing using new and conventional B cell-related markers for the diagnosis of multiple sclerosis

Transcription Factor MITF Inhibits the Transcription of CPT1B to Regulate Fatty Acid β-Oxidation and Thus Affects Stemness in Lung Adenocarcinoma Cells

Introduction: Cancer stem cells (CSCs) play critical roles in lung adenocarcinoma (LUAD) progression, and fatty acid oxidation is key for CSC growth and survival. Therefore, investigating the molecular mechanisms regulating fatty acid β-oxidation in LUAD is important for its treatment. Methods: Bioinformatics analysis assessed CPT1B and MITF expression and their correlation in LUAD tissues, as well as the pathways enriched by CPT1B. qRT-PCR assessed expression of CPT1B and MITF, while CCK-8 and sphere-forming assays were used to measure cell viability and stemness, respectively. Dual staining detected lipid accumulation, while kits were used to measure fatty acid β-oxidation and glycerol content. qRT-PCR was used to assay expression of lipid oxidation genes. Western blot was used to examine expression of stem cell-related markers. Dual-luciferase assay and ChIP assay were used to verify the binding relationship between MITF and CPT1B. Results: CPT1B was found to be highly expressed in LUAD and enriched in linoleic acid metabolism pathway and α-linolenic acid metabolism pathway. Functional experiments showed that CPT1B could promote stemness in LUAD cells by regulating fatty acid β-oxidation. Additionally, CPT1B was found to be regulated by the upstream transcription factor MITF, which was lowly expressed in LUAD and could downregulate CPT1B expression. Rescue experiments revealed that CPT1B/MITF axis could affect stemness in LUAD cells by regulating fatty acid β-oxidation. Conclusion: Transcription factor MITF inhibited transcription of CPT1B to regulate fatty acid β-oxidation, thereby suppressing stemness in LUAD cells. MITF and CPT1B may become new targets for LUAD.

Jin-Fu-An decoction manipulation of macrophage polarization via β-catenin (CTNNB1) synergizes with cisplatin in lung cancer

Previous studies have demonstrated that tumor-associated macrophages (TAMs) exhibiting an M2 phenotype contribute significantly to the pathogenesis of various cancer types, including lung cancer. Therapeutic approaches targeting TAMs have the potential to complement and synergize with conventional chemotherapy and immunotherapy. Through database analysis, it has become evident that the expression of CTNNB1 (β-catenin) is predominantly localized in macrophages, and its presence is associated with unfavorable outcomes in the absence of CD8+ cells. Jin-Fu-An decoction (JFAD) has been utilized as an adjunct to augment current clinical interventions. By conducting a network pharmacological analysis, we discovered that CTNNB1 is a significant target of JFAD. Experiments were conducted to examine the impact of JFAD on macrophage polarization both in vitro and in vivo. Furthermore, the study investigated the combined effect of JFAD and cisplatin (CDDP) on mitigating adverse reactions and prolonging survival in subcutaneously transplanted tumor models and orthotopic lung cancer models. The percentage of M1 and M2 macrophages in the tumor and spleen were measured using flow cytometry. Additionally, the levels of β-catenin, M1, and M2 macrophage markers were measured by Western blotting and qPCR, while CD8 and iNOS protein expression was analyzed via immunohistochemistry. Our research findings indicate that JFAD has the ability to modulate the transformation of M2 macrophages into M1 macrophages, augment the anticancer efficacy of CDDP, and diminish the expression of cell-related markers in M2 cells. This regulatory effect may potentially be associated with the downregulation of β-catenin expression.

UBTF Tandem Duplications in Pediatric MDS and AML: Implications for Clinical Screening and Diagnosis

Recent genomic studies in adult and pediatric acute myeloid leukemia (AML) demonstrated recurrent in-frame tandem duplications (TD) in exon 13 of upstream binding transcription factor ( UBTF) (PMID: 35176137, PMID:37085611). UBTF-TDs are subtype-defining genomic alterations associated with poor outcomes that account for ~4.3% of AMLs in childhood and up to 3% in adult AMLs under the age of 60. UBTF-TD AMLs are characterized by HOXA/HOXB dysregulation, trisomy 8 or normal cytogenetics and frequent FLT3-ITD and/or WT1 mutations. A more comprehensive investigation into the clinicopathological features of UBTF-TD AMLs is needed to better understand disease features for appropriate diagnosis and to develop more effective therapies. Here, we present 94 unique pediatric cases harboring a tandem duplication in exon 13 of UBTF with a median size of 60bp (range 21-1173bp). Of these 94 cases, 6 (6.4%) were diagnosed with childhood MDS with increased blasts (WHO), suggesting that like in adults, UBTF-TDs are not only confined to AML. UBTF-TD AMLs are associated with an overall lower median white blood cell (WBC) when compared to other common HOXA/B dysregulated leukemias ( UBTF-TD median = 10.2x10 9/L, NPM1-mutation median = 21.8x10 9/L, NUP98-rearranged ( NUP98r) median = 97x10 9/L). Available immunophenotyping on 32 U BTF-TD tumors demonstrated positivity for HLA-DR (31/31; 100%), CD117 (28/29; 95%), and CD34 (26/30; 86%) with variable positivity for CD7 (17/30; 56%) and CD64 (13/25; 52%). A limited number of cases tested for CD123 (n=11) and CD11c (n=15) were all positive for the respective marker. Furthermore, morphologic assessment of these cases revealed pleomorphic blasts with occasional Auer rods and increased immature myeloid cells with salmon-colored granules, commonly with background multilineage dysplasia and increased erythroid precursors. At the transcriptional level, UBTF-TD leukemias are largely similar to NPM1-mutated and NUP98::NSD1 AMLs with the exception of UBTF-TD cases having marked increase in expression of histone genes (e.g., HIST1H4F and HIST1H2B1) and a subset of posterior HOXB cluster genes (e.g., HOXB9). Further inspection using UMAP (Uniform Manifold Approximation and Projection) of expression profiles across different pediatric AML cohorts revealed two unclassified cases of AML that clustered with the UBTF-TD subtype ( Figure 1). One case had a WT1p.S364* and the other a somatic GATA2 p.R338fs and FLT3-ITD, but did not contain an exon 13 duplication or other defining alterations. Upon closer inspection of the UBTF gene in these cases, we uncovered that each case had an in-frame tandem duplication in exon 9 of UBTF encoding a region between HMG box 2 and 3 with features similar to those in exon 13 ( Figure 2). We therefore postulated that exon 9 TDs ( UBTF-e9) could represent a functionally equivalent subgroup of UBTF-TDalterations. To test this hypothesis, cord-blood CD34+ (cbCD34+) cells were transduced to test the transforming potential of UBTF-e9 duplications. Like with UBTF-e13 duplications, cbCD34+ cells expressing UBTF-e9 duplications displayed increased proliferation compared to UBTF -WT and lentiviral empty vector control in liquid culture; and increased colony counts and replating capacity, increased CD117 expression, and a more blast-like morphology in CFU assays. Collectively, these data suggest that like TDs in exon 13, exon 9 TDs are also sufficient to promote leukemogenic phenotypes. In conclusion, we find that UBTF-TDs occur in both MDS and AML and are associated with lower WBC count, myelodysplasia, and stem cell-related surface marker expression. We have also expanded the spectrum of UBTFalterations in myeloid neoplasms through the identification of rare UBTF duplications in exon 9 that induce a similar transcriptional signature to exon 13 UBTF-TDs. The recent identification of UBTF-TDs in adult and pediatric high-risk myeloid neoplasms has highlighted the importance of understanding and appropriately diagnosing tumors with this genomic alteration, including a new class of alterations that will not be detected by PCR-based screening strategies that specifically target exon 13 of UBTF.

FKBP5 associated CD8 T cell infiltration is a novel prognostic biomarker in luminal B breast cancer.

To investigate the relationship between FKBP prolyl isomerase 5 (FKBP5) gene expression and CD8 T cells in tumour progression and immunology of the luminal B subtype of breast cancer (LBBC) using bioinformatics analyses. The Gene Expression Profiling Interactive Analysis 2, Human Protein Atlas and breast cancer gene-expression miner v4.5 databases were used for data mining and analysing FKBP5, its co-expressed genes and CD8 T cell-related markers. The Tumor IMmune Estimation Resource 2.0 database was used for analysing the correlation and prognosis of FKBP5 and CD8 T cell infiltration level in LBBC. Upregulated FKBP5 expression was correlated with improved survival in LBBC. Upregulated FKBP5-related CD8 T cell markers were also demonstrated to be significantly correlated with better survival in LBBC and might play a role in the biological activity of FKBP5. These findings suggest that FKBP5 and its associated CD8 T cell infiltration are potential benign prognostic indicators for LBBC.

Heterogeneity of cancer stem cell-related marker expression is associated with three-dimensional structures in malignant pleural effusion produced by lung adenocarcinoma.

Cancer stem cells have been described in lung adenocarcinoma-associated malignant pleural effusion. They show clinically important features, including the ability to initiate new tumours and resistance to treatments. However, their correlation with the three-dimensional tumour structures in the effusion is not well understood. Cell blocks produced from lung adenocarcinoma patients' pleural effusion were examined for cancer stem cell-related markers Nanog and CD133 using immunocytochemistry. The three-dimensional cancer cell structures and CD133 expression patterns were visualized with tissue-clearing technology. The expression patterns were correlated with tumour cell structures, genetic variants and clinical outcomes. Thirty-nine patients were analysed. Moderate-to-strong Nanog expression was detected in 27 cases (69%), while CD133 was expressed by more than 1% of cancer cells in 11 cases (28%). Nanog expression was more homogenous within individual specimens, while CD133 expression was detected in single tumour cells or cells within small clusters instead of larger structures in 8 of the 11 positive cases (73%). Although no statistically significant correlation between the markers and tumour genetic variants or patient survival was observed, we recorded seven cases with follow-up specimens after cancer treatment, and four (57%) showed a change in stem cell-related marker expression corresponding to treatment response. Lung adenocarcinoma cells in the pleural effusion show variable expression of cancer stem cell-related markers, some showing a correlation with the size of cell clusters. Their expression level is potentially correlated with cancer treatment effects.

Leptin/lipopolysaccharide-treated dendritic cell vaccine improved cellular immune responses in an animal model of breast cancer

Purpose In dendritic cells (DCs), leptin as an immune-regulating hormone, increases the IL-12 generation whereas it reduces the IL-10 production, thus contributing to TH1 cell differentiation. Using a murine model of breast cancer (BC), we evaluated the impacts of the Leptin and/or lipopolysaccharide (LPS)-treated DC vaccine on various T-cell-related immunological markers. Materials and methods Tumors were established in mice by subcutaneously injecting 7 × 105 4T1 cells into the right flank. Mice received the DC vaccines pretreated with Leptin, LPS, and both Leptin/LPS, on days 12 and 19 following tumor induction. The animals were sacrificed on day 26 and after that the frequency of the splenic cytotoxic T lymphocytes (CTLs) and TH1 cells; interferon gamma (IFN-γ), interleukin 12 (IL-12) and tumor growth factor beta (TGF-β) generation by tumor lysate-stimulated spleen cells, and the mRNA expression of T-bet, FOXP3 and Granzyme B in the tumors were measured with flow cytometry, ELISA and real-time PCR methods, respectively. Results Leptin/LPS-treated mDC group was more efficient in blunting tumor growth (p = .0002), increasing survival rate (p = .001), and preventing metastasis in comparison with the untreated tumor-bearing mice (UT-control). In comparison to the UT-control group, treatment with Leptin/LPS-treated mDC also significantly increased the splenic frequencies of CTLs (p < .001) and TH1 cells (p < .01); promoted the production of IFN-γ (p < .0001) and IL-12 (p < .001) by splenocytes; enhanced the T-bet (p < .05) and Granzyme B (p < .001) expression, whereas decreased the TGF-β and FOXP3 expression (p < .05). Conclusion Compared to the Leptin-treated mDC and LPS-treated mDC vaccines, the Leptin/LPS-treated mDC vaccine was more effective in inhibiting BC development and boosting immune responses against tumor.

MMP12 serves as an immune cell-related marker of disease status and prognosis in lung squamous cell carcinoma.

Worldwide, lung squamous cell carcinoma (LUSC) has wreaked havoc on humanity. Matrix metallopeptidase 12 (MMP12) plays an essential role in a variety of cancers. This study aimed to reveal the expression, clinical significance, and potential molecular mechanisms of MMP12 in LUSC. There were 2,738 messenger RNA (mRNA) samples from several multicenter databases used to detect MMP12 expression in LUSC, and 125 tissue samples were validated by immunohistochemistry (IHC) experiments. Receiver operator characteristic (ROC) curves, Kaplan-Meier curves, and univariate and multivariate Cox regression analyses were used to assess the clinical value of MMP12 in LUSC. The potential molecular mechanisms of MMP12 were explored by gene enrichment analysis and immune correlation analysis. Furthermore, single-cell sequencing was used to determine the distribution of MMP12 in multiple tumor microenvironment cells. MMP12 was significantly overexpressed at the mRNA level (p<0.05, SMD = 3.13, 95% CI [2.51-3.75]), which was verified at the protein level (p<0.001) by internal IHC experiments. MMP12 expression could be used to differentiate LUSC samples from normal samples, and overexpression of MMP12 itself implied a worse clinical prognosis and higher levels of immune cell infiltration in LUSC patients. MMP12 was involved in cancer development and progression through two immune-related signaling pathways. The high expression of MMP12 in LUSC might act as an antigen-presenting cell-associated tumor neoantigen and activate the body's immune response. MMP12 expression is upregulated in LUSC and high expression of MMP12 serves as a risk factor for LUSC patients. MMP12 may be involved in cancer development by participating in immune-related signaling pathways and elevating the level of immune cell infiltration.

NCAPG stimulates lung adenocarcinoma cell stemness through aerobic glycolysis.

Cancer stem cells are pivotal in cancer progression and therapy, including lung adenocarcinoma (LUAD). High NCAPG level is implicated in malignant tumorigenesis, but investigations on NCAPG and LUAD stem cells are warranted. Hence, projecting the impact of NCAPG on cell stemness and the targeted therapy for LUAD is of the essence. Bioinformatics analyzed NCAPG expression in LUAD tissues. qRT-PCR assayed NCAPG expression in LUAD cells. CCK-8 assessed cell viability and cell sphere-forming assay measured sphere-forming ability. Western blot assessed expression of stem cell-related markers (CD133, CD44, Oct-4) and specific genes (HK2, PKM2, LDHA) related to glycolysis metabolism pathway. Cellular glycolytic capacity was assayed by glycolytic metabolites pyruvic acid, lactate, citrate, and malate assay kits, and extracellular acidification rate and oxygen consumption rate analyzers. NCAPG was upregulated in LUAD and enriched in the aerobic glycolysis pathway, and its expression was positively correlated with that of glycolytic marker genes. Cell function assays revealed that NCAPG stimulated proliferation, stemness, and glycolytic activity of LUAD cells. Rescue experiments unveiled that 2-DG (glycolysis inhibitor) was able to reverse the stimulative impact of NCAPG overexpression on proliferation, stemness, and glycolytic activity of LUAD cells. NCAPG stimulated LUAD cell stemness through activation of glycolysis pathway. NCAPG may be possible biomarker for diagnosis and target for treatment of LUAD.

Psoralidin Induced Differentiation from Adipose-derived Stem Cells to Nucleus Pulposus-like Cells by TGF-β/Smad Signaling.

Psoralidin (PL) could affect the differentiation of bone marrow mesenchymal stem cells (BMSCs). The role of PL is still unclear in adipose-derived stem cells (ADSCs). This study aimed to investigate the effects of PL on ADSCs differentiation into nucleus pulposus-like cells and the TGF-β/Smad signaling pathway. The proliferation and apoptosis of ADSCs were detected. The nucleus pulposus cell-related markers (CD24, BASP1, KRT19, and Aggrecan) and TGF-β/Smad signaling pathway indexes were analyzed. The results showed that compared to the control group, the cell activity was increased in the PL group, and the apoptosis rate was decreased. The mRNA and protein levels of nucleus pulposus cells markers (CD24, BASP1, KRT19, Aggrecan, and Collagen Type II) and TGF-β/Smad signaling pathway-related indexes (TGF-β, SMAD2, and SMAD3) were increased in PL group. After treatment with PL and TGF-β silencing, the TGF-β/Smad signaling pathway-related indicators (TGF-β, SMAD2, and SMAD3) and nucleus pulposus cells markers (CD24, BASP1, KRT19, Aggrecan, and Collagen Type II) were found to be higher in the sh-TGF-β +PL group than in the sh-TGF-β group. In conclusion, our study showed that PL might induce the differentiation of ADSCs to nucleus pulposus cells through the TGF-β/Smad signaling pathway. It might have the potential application value in the treatment of intervertebral disc degeneration.