Gastric cancer mesenchymal stem cells promote tumor glycolysis and chemoresistance by regulating B7H3 in gastric cancer cells.

Despite surgical treatment combined with multidrug therapy having made some progress, chemotherapy resistance is the main cause of recurrence and death of gastric cancer (GC). Gastric cancer mesenchymal stem cells (GCMSCs) have been reported to be correlated with the limited efficacy of chemotherapy in GC, but the mechanism of GCMSCs regulating GC resistance needs to be further studied. The gene set enrichment analysis (GSEA) was performed to explore the glycolysis-related pathways heterogeneity across different cell subpopulations. Glucose uptake and lactate production assays were used to evaluate the importance of B7H3 expression in GCMSCs-treated GC cells. The therapeutic efficacy of oxaliplatin (OXA) and paclitaxel (PTX) was determined using CCK-8 and colony formation assays. Signaling pathways altered by GCMSCs-CM were revealed by immunoblotting. The expression of TNF-α in GCMSCs and bone marrow mesenchymal stem cells (BMMSCs) was detected by western blot analysis and qPCR. Our results showed that the OXA and PTX resistance of GC cells were significantly enhanced in the GCMSCs-CM treated GC cells. Acquired OXA and PTX resistance was characterized by increased cell viability for OXA and PTX, the formation of cell colonies, and decreased levels of cell apoptosis, which were accompanied by reduced levels of cleaved caspase-3 and Bax expression, and increased levels of Bcl-2, HK2, MDR1, and B7H3 expression. Blocking TNF-α in GCMSCs-CM, B7H3 knockdown or the use of 2-DG, a key enzyme inhibitor of glycolysis in GC cells suppressed the OXA and PTX resistance of GC cells that had been treated with GCMSCs-CM. This study shows that GCMSCs-CM derived TNF-α could upregulate the expression of B7H3 of GC cells to promote tumor chemoresistance. Our results provide a new basis for the treatment of GC.

Cancer Stem Cells and the Tumor Microenvironment in Tumor Drug Resistance.

Although there has been some progress in the efficacy of anti-cancer drugs, drug resistance remains challenging. Cancer stem cells (CSCs) are self-renewing and differentiate into cancer tissues with tumor heterogeneity. CSCs are associated with the progression of breast, colon, and lung cancers. Hence, recent studies have focused on the role of CSCs in resistance to anti-cancer drugs. Increasing evidence suggests that CSCs interact with components of the tumor microenvironment (TME), such as vascular and immune cells, as well as various cytokines, and are regulated by multiple signaling pathways, thereby promoting drug resistance in various cancers. Therefore, it is important to clarify the mechanisms underlying the crosstalk between CSCs and the TME for the development of targeted anti-cancer therapies.

Gastric cancer mesenchymal stem cells via the CXCR2/HK2/PD-L1 pathway mediate immunosuppression.

BACKGROUND: Anti-PD-1 immunotherapy has emerged as an important therapeutic modality in advanced gastric cancer (GC). However, drug resistance frequently develops, limiting its effectiveness. METHODS: The role of gastric cancer mesenchymal stem cells (GCMSCs) in anti-PD-1 resistance was evaluated in vivo in NPGCD34+ or NCGPBMC xenograft mouse model. In addition, we investigated CD8+T cell infiltration and effector function by spectral cytometry and IHC. The effects of GCMSCs conditional medium (GCMSC-CM) on GC cell lines were characterized at the level of the proteome, secretome using western blot, and ELISA assays. RESULTS: We reported that GCMSCs mediated tolerance mechanisms contribute to tumor immunotherapy tolerance. GCMSC-CM attenuated the antitumor activity of PD-1 antibody and inhibited immune response in humanized mouse model. In GC cells under serum deprivation and hypoxia, GCMSC-CM promoted GC cells proliferation via upregulating PD-L1 expression. Mechanistically, GCMSC-derived IL-8 and AKT-mediated phosphorylation facilitated HK2 nuclear localization. Phosphorylated-HK2 promoted PD-L1 transcription by binding to HIF-1α. What is more, GCMSC-CM also induced lactate overproduction in GC cells in vitro and xenograft tumors in vivo, leading to impaired function of CD8+ T cells. Furthermore, CXCR1/2 receptor depletion, CXCR2 receptor antagonist AZD5069 and IL-8 neutralizing antibody application also significantly reversed GCMSCs mediated immunosuppression, restoring the antitumor capacity of PD-1 antibody. CONCLUSIONS: Our findings reveal that blocking GCMSCs-derived IL-8/CXCR2 pathway decreasing PD-L1 expression and lactate production, improving antitumor efficacy of anti-PD-1 immunotherapy, may be of value for the treatment of advanced gastric carcinoma.

Lactate induced mesenchymal stem cells activation promotes gastric cancer cells migration and proliferation.

Lactate extensively involves in gastric cancer (GC) progression, such as suppressing immune cells function and facilitating tumor angiogenesis. However, it remains unclear whether lactate promotes tumor progression by interacting with mesenchymal stem cells (MSCs), one of the major stroma components in GC. Here, we investigated the influence of lactate on the phenotype and function of MSCs. The migration of MSCs and the expression of several CAF markers in MSCs after lactate treatment were detected. We also evaluated the effect of lactate-primed MSCs on GC cells migration, proliferation, and programmed death ligand 1 (PD-L1) expression. It was found that lactate significantly activated MSCs, and increased fibroblast activation protein (FAP) expression via monocarboxylate transporter 1 (MCT1)/transforming growth factor-beta 1 (TGF-ß1) signaling. In addition, lactate-primed MSCs promoted GC cells migration and proliferation via PD-L1. Inhibiting MCT1 by AZD3965 abrogated lactate induced FAP expression and tumor-promoting potential of MSCs. Therefore, targeting MCT1/TGF-ß1/FAP axis in MSCs may serve as a potential strategy to restrain GC development.

Gastric cancer-derived mesenchymal stem cells promote gastric cancer cell lines migration by modulating CD276 expression.

CD276 has been studied in a variety of cancers and diseases, but its regulatory mechanisms in gastric cancer is still unclear. Mesenchymal stem cells (MSCs), one of the important members of tumor microenvironment, play an important role in the occurrence, development and metastasis of tumor, but the relationship between gastric cancer mesenchymal stem cells (GCMSCs) and CD276 in gastric cancer needs to be further explored. The differential expression of CD276 was identified via UCLAN and GEPIA databases. Then, the impacts of CD276 were calculated on clinical prognosis using the Kaplan-Meier plotter and Cox analysis. GO, KEGG and GSEA analysis were used to explore potential mechanism under CD276. Next, the expression of CD276 in gastric cell lines were detected by Western blot. Immunocoprecipitation was used to explore the association between CD276 and COL1A1. And the effect of condition medium (CM) from GCMSCs on gastric cell lines migration analyzed. GC-MSCs activated the AKT/c-Myc/mTOR pathway of gastric cell lines and upregulated CD276 expression. Moreover, the upregulation of CD276 promoted the migration of gastric cancer cells. Taken together, this study shown that GCMSCs could up-regulate the expression of CD276 of gastric cell lines to promote tumor migration. Our results provide a new basis for the treatment of gastric cancer.

Inhibition of CCCTC Binding Factor-Programmed Cell Death Ligand 1 Axis Suppresses Emergence of Chemoresistance Induced by Gastric Cancer-Derived Mesenchymal Stem Cells.

Background: Gastric cancer (GC) is the third leading cause of cancer-associated deaths worldwide. Stromal cells, especially mesenchymal stem cells (MSCs), play significant roles in the development of therapy resistance depending on their paracrine function. The PD-1/PD-L1 crosstalk between cancer and immune cells has been well studied. Emerging evidence suggests that PD-L1 also contributes to tumor resistance to therapy. Methods: Cell survival and apoptosis were assessed using CCK-8, colony formation, and flow cytometry assays. Protein alterations were analyzed via Western blot. Gene knockdown and overexpression were achieved with siRNA/shRNA and lentiviral infection, respectively. Drug effects on tumors in vivo were assessed with xenografts in nude mice. In addition, GC patient samples after chemotherapy treatment were collected to observe the relationship between chemotherapy effect and CTCF or PD-L1. Results: In response to 5-fluorouracil or paclitaxel treatment, GCMSC-CM enhanced the cell viability and decreased the apoptosis rate. Furthermore, blocking PD-L1 or CTCF in GC cells prevented GCMSC-induced drug resistance accompanied by a decline in cell stemness. Consistent with these in vitro observations, mice treated with GCMSC-CM showed a lower sensitivity to 5-fluorouracil. In addition, high expression of CTCF and PD-L1 was associated with poor chemotherapy progression in the clinic. Conclusion: Study results demonstrate a mechanism where GCMSC-CM promotes GC chemoresistance by upregulating CTCF-PD-L1 and provide strong evidence in support of targeting CTCF-PD-L1 signaling as a strategy to prevent resistance in the clinic.

Associations of the MTHFR rs1801133 polymorphism with gastric cancer risk in the Chinese Han population.

In recent years, increasing evidence has implicated the importance of mutations in the MTHFR gene in the risk of gastric cancer risk. A single nucleotide polymorphism (SNP) in the MTHFR gene (rs1801133) may serve a critical role in gastric cancer. A hospital-based case-controlled study was performed to assess the risk of the rs1801133 polymorphism on gastric cancer. A total of 307 patients with gastric cancer and 560 patients in the control group were recruited. Genomic DNA was extracted from peripheral blood and genotyped for rs1801133 using the ligase detection reaction. The relationship between rs1801133 and gastric cancer risk was evaluated by unconditional logistical regression analysis. The rs1801133-TT genotype was associated with a borderline significantly decreased risk of gastric cancer [(TT vs. CC, adjusted odds ratio (OR)=0.54, 95% confidence intervals (CI)=0.35-0.83; P=0.006; CT vs. CC, adjusted OR=0.59, 95% CI=0.44-0.79, P<0.001; and TT/CT vs. CC, adjusted OR=0.61, 95% CI=0.44-0.83, P=0.001), and further analysis showed the relationship was evident amongst older patients and patients who never drank alcohol. The C>T mutation at rs1801133 of the MTHFR gene was associated with a decreased risk of gastric cancer in older individuals and those who never drink.

Gastric cancer mesenchymal stem cells regulate PD-L1-CTCF enhancing cancer stem cell-like properties and tumorigenesis.

Rationale: Mesenchymal stem cells (MSCs) have been the focus of many studies because of their abilities to modulate immune responses, angiogenesis, and promote tumor growth and metastasis. Our previous work showed that gastric cancer MSCs (GCMSCs) promoted immune escape by secreting of IL-8, which induced programmed cell death ligand 1 (PD-L1) expression in GC cells. Mounting evidence has revealed that PD-L1 expression is related to intrinsic tumor cell properties. Here, we investigated whether GCMSCs maintained a pool of cancer stem cells (CSCs) through PD-L1 signaling and the specific underlying molecular mechanism. Methods: Stem cell surface markers, aldehyde dehydrogenase (ALDH) activity, migration and sphere formation abilities were tested to evaluate the stemness of GC cells. PD-L1-expressing lentivirus and PD-L1 specific siRNA were used to analyze the effects of PD-L1 on GC cells stemness. Annexin V/PI double staining was used to assess apoptosis of GC cells induced by chemotherapy. Co-Immunoprecipitation (Co-IP) and Mass spectrometry were employed to determine the PD-L1 binding partner in GC cells. PD-L1Negative and PD-L1Positive cells were sorted by flow cytometry and used for limiting dilution assays to verify the effect of PD-L1 on tumorigenic ability in GC cells. Results: The results showed that GCMSCs enhanced the CSC-like properties of GC cells through PD-L1, which led to the resistance of GC cells to chemotherapy. PD-L1 associated with CTCF to contribute to the stemness and self-renewal of GC cells. In vivo, PD-L1Positive GC cells had greater stemness potential and tumorigenicity than PD-L1Negative GC cells. The results also indicated that GC cells were heterogeneous, and that PD-L1 in GC cells had different reactivity to GCMSCs. Conclusions: Overall, our data indicated that GCMSCs enriched CSC-like cells in GC cells, which gives a new insight into the mechanism of GCMSCs prompting GC progression and provides a potential combined therapeutic target.

Microstructure and Mechanical Properties of 4Al Alumina-Forming Austenitic Steel after Cold-Rolling Deformation and Annealing.

Microstructural evolutions of the 4Al alumina-forming austenitic steel after cold rolling with different reductions from 5% to 30% and then annealing were investigated using electron backscattering diffraction (EBSD), X-ray diffraction (XRD) and transmission electron microscopy (TEM). Tensile properties and hardness were also measured. The results show that the average grain size gradually decreases with an increase in the cold-rolling reduction. The low angle grain boundaries (LAGBs) are dominant in the cold-rolled samples, but high angle grain boundaries (HAGBs) form in the annealed samples, indicating that the grains are refined under the action of dislocations. During cold rolling, high-density dislocations are initially introduced in the samples, which contributes to a large number of dislocations remaining after annealing. With the sustaining increase in cold-rolled deformation, the samples exhibit more excellent tensile strength and hardness due to the decrease in grain size and increase in dislocation density, especially for the samples subjected to 30% cold-rolling reduction. The contribution of dislocations on yield strength is more than 60%.

Exosomes Derived from Human Umbilical Cord Mesenchymal Stem Cells Promote Fibroblast-to-Myofibroblast Differentiation in Inflammatory Environments and Benefit Cardioprotective Effects.

Cardioprotective effects of exosomes derived from human umbilical cord mesenchymal stem cells (hucMSC-exosomes) postmyocardial infarction (post-MI) have been reported in our previous study. It is known that fibroblasts are pro-inflammatory phenotypes, while myofibroblasts are anti-inflammatory phenotypes. This study aimed to investigate whether hucMSC-exosomes promoted cardiac fibroblast-to-myofibroblast differentiation in inflammatory environments and protected cardiomyocytes. Rats were performed by permanent ligation of the left anterior descending coronary artery and underwent intramyocardial injection of hucMSC-exosomes or phosphate-buffered saline (PBS) in surgery. Fibroblasts were stimulated by lipopolysaccharide (LPS) to create inflammatory environments in vitro. Western blot and immunohistochemical and immunofluorescence staining for α-smooth muscle actin were used to demonstrate fibroblast-to-myofibroblast differentiation. Transwell migration assay and CCK-8 assay were used to evaluate migration and proliferation of fibroblasts. Reverse transcription-polymerase chain reaction, western blot, and immunohistochemical staining were used to detect expressions of inflammatory factors. To investigate cardioprotective effects, cardiomyocytes were treated with supernatant derived from fibroblasts pretreated with LPS or LPS plus hucMSC-exosomes in hypoxic environments. Cardiomyocyte apoptosis was determined using TUNEL assay and western blot. Results indicated that hucMSC-exosomes increased the density of myofibroblasts in infarct areas during inflammatory phases post-MI, promoted fibroblast-to-myofibroblast differentiation in inflammatory environments, and attenuated inflammatory responses in vitro and in vivo. Culture medium derived from fibroblasts pretreated with LPS plus hucMSC-exosomes reduced cardiomyocyte apoptosis. In vivo, apoptotic cells in acute myocardial infarction (AMI)+exosomes groups were also less than AMI+PBS groups. In conclusion, hucMSC-exosomes can promote fibroblast-to-myofibroblast differentiation in inflammatory environments, then protecting cardiomyocytes.

Precipitates and Particles Coarsening of 9Cr-1.7W-0.4Mo-Co Ferritic Heat-Resistant Steel after Isothermal Aging.

The precipitates obtained by EPE technology from the 9Cr-1.7W-0.4Mo-Co ferritic heat-resistant steel subject to isothermal aging were investigated using SEM, TEM and XRD. The particle size distribution and the coarsening kinetics of M23C6 with duration of isothermal aging were also analyzed with or without consideration of Laves phase. The results show that the isolated dislocations were detected in delta ferrite interior, and the precipitates on delta ferrite and martensite boundaries are obviously larger than other locations. Fe2W-Laves phase can only be found as duration of aging time to 2000 h, and is preferential to form adjacent to M23C6 particles. The small M23C6 particles firstly coarsen, but the large M23C6 are relatively stable during short aging. The total coarsening rate of M23C6 precipitates is 9.75 × 10-28m3s-1, and the coarsening of M23C6 depends on the formation of Laves phase.