Proliferation Of B16 Cells Research Articles

A dual-pathway pyroptosis inducer based on Au–Cu2-xSe@ZIF-8 enhances tumor immunotherapy by disrupting the zinc ion homeostasis

The regulation of intracellular ionic homeostasis to trigger antigen-specific immune responses has attracted extensive interest in tumor therapy. In this study, we developed a dual-pathway nanoreactor, Au-Cu2-xSe@ZIF-8@P18 NPs (ACS-Z-P NPs), which targets danger-associated molecular patterns (DAMPs) and releases Zn2+ and reactive oxygen species (ROS) within the tumor microenvironment (TME). Zn2+ released from the metal–organic frameworks (MOFs) was deposited in the cytoplasm, leading to aberrant transcription levels of intracellular zinc-regulated proteins and DNA damage, thereby inducing pyroptosis and immunogenic cell death (ICD) dependent on caspase1/gasdermin D (GSDMD) pathway. Furthermore, upon laser irradiation, ACS-Z-P NPs could break through the limitations of inherent defects of immunosuppression in TME, enhance ROS generation through a Fenton-like reaction cascade, which subsequently triggered the activation of inflammatory vesicles and the release of damage-associated molecular patterns (DAMPs). This cascade effect led to the amplification of pyroptosis and immunogenic cell death (ICD), thereby remodeling the immunosuppressed TME. Consequently, this process improved dendritic cell (DC) antigen presentation and augmented anti-tumor T-cell responses, effectively initiating antigen-specific immune responses and further enhancing pyroptosis and ICD. This study explores the therapeutic properties of these mechanisms in detail. Statement of significanceThe synthesized Au-Cu2-xSe@ZIF-8@P18 nanoparticles (ACS-Z-Ps) can effectively enhance the bodyʼs immune response by regulating zinc ion levels within cells. This regulation leads to abnormal levels of zinc-regulated protein transcription and DNA damage, which induces cellular pyroptosis. As a result, antigen presentation to dendritic cells (DCs) is improved, and anti-tumor T-cell responses are enhanced.The ACS-Z-P NPs overcome the limitations of ROS deficiency and immunosuppression in the tumor microenvironment by using H2O2 in the tumor microenvironment through a Fenton-like reaction. This leads to an increased production of ROS and O2, remodeling of the immunosuppressed tumor microenvironment, and enhanced induction of cell pyroptosis and immunogenic cell death.ACS-Z-P NPs targeted B16 cells using the photosensitizer P18 in combination with PDT treatment. This approach significantly inhibited the proliferation of B16 cells and effectively inhibited tumor growth.

Tumor- and Fibroblast-Derived Cell-Free DNAs Differently Affect the Progression of B16 Melanoma In Vitro and In Vivo.

It is widely postulated that the majority of pathologically elevated extracellular or cell-free DNA (cfDNA) in cancer originates from tumor cells; however, evidence has emerged regarding the significant contributions of other cells from the tumor microenvironment. Here, the effect of cfDNA originating from murine B16 melanoma cells and L929 fibroblasts on B16 cells was investigated. It was found that cfDNAL929 increased the viability and migration properties of B16 cells in vitro and their invasiveness in vivo. In contrast, cfDNAB16 exhibited a negative effect on B16 cells, reducing their viability and migration in vitro, which in vivo led to decreased tumor size and metastasis number. It was shown that cell treatment with both cfDNAs resulted in an increase in the expression of genes encoding DNases and the oncogenes Braf, Kras, and Myc. cfDNAL929-treated cells were shown to experience oxidative stress. Gene expression changes in the case of cfDNAB16 treatment are well correlated with the observed decrease in proliferation and migration of B16 cells. The obtained data may indicate the possible involvement of fibroblast DNA in the tumor microenvironment in tumor progression and, potentially, in the formation of new tumor foci due to the transformation of normal cells.

Quercetin Limits Tumor Immune Escape through PDK1/CD47 Axis in Melanoma.

Quercetin (3,3[Formula: see text],4[Formula: see text],5,7-pentahydroxyflavone) is a bioactive plant-derived flavonoid, abundant in fruits and vegetables, that can effectively inhibit the growth of many types of tumors without toxicity. Nevertheless, the effect of quercetin on melanoma immunology has yet to be determined. This study aimed to investigate the role and mechanism of the antitumor immunity action of quercetin in melanoma through both in vivo and in vitro methods. Our research revealed that quercetin has the ability to boost antitumor immunity by modulating the tumor immune microenvironment through increasing the percentages of M1 macrophages, CD8[Formula: see text] T lymphocytes, and CD4[Formula: see text] T lymphocytes and promoting the secretion of IL-2 and IFN-[Formula: see text] from CD8[Formula: see text] T cells, consequently suppressing the growth of melanoma. Furthermore, we revealed that quercetin can inhibit cell proliferation and migration of B16 cells in a dose-dependent manner. In addition, down-regulating PDK1 can inhibit the mRNA and protein expression levels of CD47. In the rescue experiment, we overexpressed PDK1 and found that the protein and mRNA expression levels of CD47 increased correspondingly, while the addition of quercetin reversed this effect. Moreover, quercetin could stimulate the proliferation and enhance the function of CD8[Formula: see text] T cells. Therefore, our results identified a novel mechanism through which CD47 is regulated by quercetin to promote phagocytosis, and elucidated the regulation of quercetin on macrophages and CD8[Formula: see text] T cells in the tumor immune microenvironment. The use of quercetin as a therapeutic drug holds potential benefits for immunotherapy, enhancing the efficacy of existing treatments for melanoma.

Effect of up-regulation of micro-ribonucleic acid-22 and NLRP3 inhibition on viability of malignant melanoma cells

Purpose: To determine the effect of up-regulation of micro-ribonucleic acid-22 (miR-22) targeting nucleotide-binding oligomerization domain, leucine-rich repeat and pyrin domain-containing 3 (NLRP3) on the proliferation and invasiveness of malignant melanoma cells, as well as the underlying mechanisms.
 Methods: Mouse-derived melanoma B16 cells were subjected to thawing, sub-culture and transfection. The cells were assigned to groups (A1 - A5) in line with different transfections viz: A1 (miR-22 mimic overexpression-transfected), A2 (miR-22 inhibitor-transfected), A3 (miR-22 mimic+siNLRP3-transfected), A4 (miR-22 mimic NC sequence-transfected), and A5 (un-transfected).
 Results: The miR-22 expression was significantly up-regulated in A1, relative to A2, A3, A4, and A5 groups, but significantly lower, relative to A3, A4, and A5. The NLRP3 mRNA and protein levels were significantly lower in A1 than in other groups, but significantly up-regulated in A2, relative to A3, A4, and A5 (p < 0.05). Cell proliferation rate and colony formation rate were significantly lower in A1 group but were significantly up-regulated in A2, relative to A3, A4, and A5.
 Conclusion: Targeting NLRP3 inhibition by up-regulating miR-22 expression level significantly reduces the proliferation, invasiveness, and matrix metalloproteinase levels of melanoma B16 cells, thereby reducing the occurrence and development of malignant melanoma. The results provide some useful reference data for the treatment of malignant melanoma at the gene level.

Preparation of Lycium barbarum Active Glycopeptide and Investigation of Its Apoptotic Effects on Melanoma.

The increasing number of studies have shown that Lycium barbarum polysaccharides possess anti-tumor effects. However, the determination of the active ingredients and their mechanism against melanoma inhibition are still unknown. In this study, we aimed to investigate the mechanisms of action of Lycium barbarum active glycopeptide (LBAG) on melanoma. LBAG was extracted and isolated from the fruit of Lycium barbarum using aqueous alcoholic precipitation and identified using ultra-performance liquid chromatography-quadrupole-time of flightmass spectrometry. Various assays including cell apoptosis, cell cycle analysis, colony formation assay, cell scratch test, flow cytometry, and Western blot were performed to evaluate the effects of LBAG on melanoma. The results showed that LBAG has a molecular weight of 10-15 kDa and contains Man, Rha, GlcA, Glc, Gal, and Ara18 amino acids. Treatment with LBAG significantly decreased B16 cell proliferation and induced cell cycle arrest at the G0/G phase, accompanied by the accumulation of reactive oxygen species. Western blot analysis revealed that the phosphorylation of P38-MAPK and AKT, as well as the expression of N-acetyl-Lcysteine, were related to cell apoptosis and cell cycle regulation. In mouse xenografts, LBAG inhibited tumor growth through the P38-MAPK and AKT signaling pathways. In conclusion, the anti-melanoma activity of LBAG may induce apoptosis in cancer cells through ROSmediated activation of the P38-MAPK and AKT signaling pathways. These findings provide a foundation for further research on the anti-melanoma potential of LBAG.

Effects of Resveratrol on Mouse B16 Melanoma Cell Proliferation through the SHCBP1-ERK1/2 Signaling Pathway.

Melanoma originates from the malignant mutational transformation of melanocytes in the basal layer of the epidermal layer of the skin. It can easily spread and metastasize in the early stage, resulting in a poor prognosis. Therefore, it is particularly important to find effective antitumor adjuvant drugs to inhibit the occurrence and development of melanoma. In this study, we found that resveratrol, a polyphenolic compound from grape plants, can significantly inhibit the proliferation, colony formation and migration of mouse melanoma B16 cells. Notably, resveratrol was also found to inhibit the expression of SHCBP1 in B16 cells. Transcriptional analysis and cellular studies showed that SHCBP1 can activate the MAPK/ERK signaling pathway to regulate cyclin expression and promote the G1/S phase transition of the cell cycle by upregulating ERK1/2 phosphorylation levels. Resveratrol further downregulates the phosphorylation level of ERK1/2 by inhibiting SHCBP1 expression, thus inhibiting tumor cell proliferation. In conclusion, resveratrol inhibits the proliferation of B16 cells by regulating the ERK1/2 signaling pathway through SHCBP1. As an upstream protein of the ERK1/2 signaling pathway, SHCBP1 may be involved in the process of resveratrol-mediated inhibition of tumor cell proliferation.

Maprotiline Prompts an Antitumour Effect by Inhibiting PD-L1 Expression in Mice with Melanoma.

Research has revealed that the expression of PD-L1 is significantly upregulated in tumour cells and that the binding of programmed cell death protein 1 (PD-1) to programmed cell death 1 ligand 1 (PD-L1) inhibits the response of T cells, thereby suppressing tumour immunity. Therefore, blocking PD-L1/PD-1 signalling has become an important target in clinical immunotherapy. Some old drugs, namely, non-anticancer drugs, have also been found to have antitumour effects, and maprotiline is one of them. Maprotiline is a tetracyclic antidepressant that has been widely used to treat depression. However, it has not yet been reported whether maprotiline can exert an antitumour effect on melanoma. This study aimed to investigate the antitumour efficacy of maprotiline in mice with melanoma. In this study, female C57BL/6 mice were used to establish a tumour-bearing animal model. After treatment with maprotiline, the survival rate of mice was recorded daily. The expression of relevant proteins was detected by Western blotting, the proportion of immune cells was detected by flow cytometry, and the infiltration of immune cells in tumour tissue was detected by immunofluorescence staining. Maprotiline was found to inhibit the proliferation and migration of B16 cells while increasing cell apoptosis. Importantly, treatment with maprotiline decreased the expression of PD-L1 and increased the proportion of CD4+ T cells, CD8+ T cells, and NK cells in the spleen. It also increased the infiltration of CD4+ and CD8+ T cells in tumour tissue. Our research findings suggest that maprotiline enhances the antitumour immune response in mouse melanoma by inhibiting PD-L1 expression. This study may discover a new PD-L1 inhibitor, providing a novel therapeutic option for the clinical treatment of tumours.

Oxymatrine inhibits melanoma development by modulating the immune microenvironment and targeting the MYC/PD-L1 pathway

Oxymatrine, also known as ammothamnine or oxysophoridine, is a natural compound isolated from Sophora flavescens (in Chinese, Kushen), and many previous researchers have characterized its anti-inflammatory, anti-fibrotic and anti-tumor properties. However, the underlying anti-tumor immunological mechanism of oxymatrine remains elusive. In this study, we carried out experiments both in vitro and in vivo and investigated the anti-tumor effect of oxymatrine to inhibit the proliferation and migration of melanoma B16 cells, while promoting apoptosis. Oxymatrine upregulated CD4+ T, CD8+ T and NKT cells, downregulated Treg cells, promoted TNF-α secretion, and successfully modulated the immune microenvironment and ultimately suppressed melanoma development in subcutaneous tumor models established in mice. Evidence from network pharmacology and RNAseq suggested that possible targets of oxymatrine for melanoma treatment included PD-L1 and MYC. We observed oxymatrine inhibited PD-L1 and MYC expression in melanoma cells via qRT-PCR and western blotting analysis, and found MYC potentially regulated PD-L1 to mediate anti-tumor effects. These findings provide insight into the mechanism by which oxymatrine inhibits melanoma and enhances the anti-tumor immune effect. In summary, our study proposes a novel approach to suppress melanoma by targeting the MYC/PD-L1 pathway using oxymatrine, which may develop into a less toxic and more efficient anti-tumor agent for melanoma treatment.

Vitamin D3 regulates NSUN2 expression and inhibits melanoma cell proliferation and migration.

The activated form of vitamin D3 [1,25-dihydroxyvitamin D3; 1,25(OH)2D3] is important for various physiological processes, such as bone mineralization and calcium metabolism, and plays an anticancer role in numerous cancers as well. Its role in melanoma cells has yet to be proven. NOP2/Sun RNA methyltransferase 2 (NSUN2) is a typical RNA methyltransferase and is highly expressed in a variety of cancer cells. However, the molecular mechanisms underlying the role of 1,25(OH)2D3 and NSUN2 in melanoma cells remain largely unknown. The current study showed that 1,25(OH)2D3 could significantly and specifically inhibit the proliferation and migration of melanoma B16 cells. 1,25(OH)2D3 enhances vitamin D receptor expression while simultaneously reducing NSUN2 expression in melanoma cells. Subsequently, knockdown of NSUN2 suppressed B16 cell proliferation and migration. RNA-Seq results illuminated that DNA replication, cell proliferation and cell cycle pathways were enriched, and genes promoting these pathways were reduced after knocking down Nsun2. Dual-luciferase reporter assays showed that 1,25(OH)2D3 downregulated reporter gene expression was controlled by the Nsun2 promoter. The results suggest that 1,25(OH)2D3 binds to the vitamin D response element located upstream of the Nsun2 promoter to downregulate Nsun2 transcription activity and then affects the gene expression pattern related to cell proliferation and the cell cycle, thereby restraining B16 cell proliferation and migration.

Exosome-like Nanovesicles Derived from the Mucilage of Pinctada Martensii Exhibit Antitumor Activity against 143B Osteosarcoma Cells.

Osteosarcoma is prone to metastasis and has a low long-term survival rate. The drug treatment of osteosarcoma, side effects of treatment drugs, and prognosis of patients with lung metastasis continue to present significant challenges, and the efficacy of drugs used in the treatment of osteosarcoma remains low. The development of new therapeutic drugs is urgently needed. In this study, we successfully isolated Pinctada martensii mucilage exosome-like nanovesicles (PMMENs). Our findings demonstrated that PMMENs inhibited the viability and proliferation of 143B cells, induced apoptosis, and inhibited cell proliferation by suppressing the activation of the ERK1/2 and Wnt signaling pathways. Furthermore, PMMENs inhibited cell migration and invasion by downregulating N-cadherin, vimentin, and matrix metalloprotease-2 protein expression levels. Transcriptomic and metabolomic analyses revealed that differential genes were co-enriched with differential metabolites in cancer signaling pathways. These results suggest that PMMENs may exert anti-tumor activity by targeting the ERK1/2 and Wnt signaling pathways. Moreover, tumor xenograft model experiments showed that PMMENs can inhibit the growth of osteosarcoma in mice. Thus, PMMENs may be a potential anti-osteosarcoma drug.

A Systematic Study of Anti-Osteosarcoma Mechanism of pH-Sensitive Charge-Conversion Cinnamaldehyde Polymeric Prodrug Micelles In Vitro.

Osteosarcoma is an aggressive malignant neoplasm, and it is of great significance to the fabrication and investigation of the anti-tumor mechanism of nanomedicine in the treatment of osteosarcoma. Herein, a cinnamaldehyde polymeric prodrug micelle with pH-sensitive charge-conversion ability (mPEG-b-P(C7-co-CA)) was fabricated, and the anti-osteosarcoma mechanism of mPEG-b-P(C7-co-CA) micelle was investigated. mPEG-b-P(C7-co-CA) micelles were prepared by self-assembly method, and their diameter was 227 nm. mPEG-b-P(C7-co-CA) micelles could regulate the cell cycle and inhibit the proliferation of 143B cells, which was demonstrated by flow cytometry analysis, CCK-8 assay and 5-Ethynyl-2'-deoxyuridine (EdU) staining. The wound-healing assay and transwell assay showed that mPEG-b-P(C7-co-CA) micelles effectively inhibited the migration and invasion of 143B cells. It was proven that mPEG-b-P(C7-co-CA) micelles downregulated the levels of proliferation and apoptosis-related proteins and affected osteosarcoma migration and invasion by inhibiting the epithelial-mesenchymal transition (EMT). In addition, mPEG-b-P(C7-co-CA) micelles can also inhibit the transcriptional activity of the PI3K/Akt signaling pathway. Therefore, these findings provide new evidence for the pharmacological effects of mPEG-b-P(C7-co-CA) micelles.

PCSK9 facilitates melanoma pathogenesis via a network regulating tumor immunity

BackgroundPCSK9 regulates cholesterol homeostasis and promotes tumorigenesis. However, the relevance of these two actions and the mechanisms underlying PCSK9’s oncogenic roles in melanoma and other cancers remain unclear.MethodsPCSK9’s association with melanoma was analysed using the TCGA dataset. Empty vector (EV), PCSK9, gain-of-function (D374Y), and loss-of-function (Q152H) PCSK9 mutant were stably-expressed in murine melanoma B16 cells and studied for impact on B16 cell-derived oncogenesis in vitro and in vivo using syngeneic C57BL/6 and Pcsk9−/− mice. Intratumoral accumulation of cholesterol was determined. RNA-seq was performed on individual tumor types. Differentially-expressed genes (DEGs) were derived from the comparisons of B16 PCSK9, B16 D374Y, or B16 Q152H tumors to B16 EV allografts and analysed for pathway alterations.ResultsPCSK9 expression and its network negatively correlated with the survival probability of patients with melanoma. PCSK9 promoted B16 cell proliferation, migration, and growth in soft agar in vitro, formation of tumors in C57BL/6 mice in vivo, and accumulation of intratumoral cholesterol in a manner reflecting its regulation of the low-density lipoprotein receptor (LDLR): Q152H, EV, PCSK9, and D374Y. Tumor-associated T cells, CD8 + T cells, and NK cells were significantly increased in D374Y tumors along with upregulations of multiple immune checkpoints, IFNγ, and 143 genes associated with T cell dysfunction. Overlap of 36 genes between the D374Y DEGs and the PCSK9 DEGs predicted poor prognosis of melanoma and resistance to immune checkpoint blockade (ICB) therapy. CYTH4, DENND1C, AOAH, TBC1D10C, EPSTI1, GIMAP7, and FASL (FAS ligand) were novel predictors of ICB therapy and displayed high level of correlations with multiple immune checkpoints in melanoma and across 30 human cancers. We observed FAS ligand being among the most robust biomarkers of ICB treatment and constructed two novel and effective multigene panels predicting response to ICB therapy. The profiles of allografts produced by B16 EV, PCSK9, D374Y, and Q152H remained comparable in C57BL/6 and Pcsk9−/− mice.ConclusionsTumor-derived PCSK9 plays a critical role in melanoma pathogenesis. PCSK9’s oncogenic actions are associated with intratumoral cholesterol accumulation. PCSK9 systemically affects the immune system, contributing to melanoma immune evasion. Novel biomarkers derived from the PCSK9-network effectively predicted ICB therapy responses.

Analyzing the mechanism by which oyster peptides target IL-2 in melanoma cell apoptosis based on RNA-seq and m6A-seq.

Melanoma is a kind of skin cancer with high malignancy and strong proliferation and invasion abilities. Chemotherapy drugs in the clinic have the disadvantages of high price and high toxicity. Peptides are natural active ingredients that have many functions and are safe and effective. Previous studies have shown that oysters are rich in protein and have antitumor effects. In this study, a high-throughput strategy combined with MALDI TOF/TOF-MS and molecular docking was developed to screen peptides with antitumor functions from oyster hydrolysate. Three dominant peptides were predicted to have similar functions to IL-2 via molecular docking. Then, the activity of the peptides was confirmed in B16 cells, and we found that the three peptides increased the apoptosis of B16 cells. Furthermore, via RNA-seq and m6A-seq of B16 cells treated with the peptides, we found that ILADSAPR downregulates the expression of Pcna, Tlr4, and Ncbp2 and upregulates the expression of Bax, Bad, Pak4, Rasa2, Cct6, and Gbp2. ILADSAPR inhibited B16 cell proliferation and promoted cell apoptosis by regulating the expression of these genes. In addition, the result of metabolic pathway analysis also proved this point. This study provides a preliminary reference for antitumor research on oyster peptides.

EGCG regulated osteolytic microenvironment to enhance the antitumor effect of DOX on orthotopic osteosarcoma

Here, we demonstrated that EGCG could enhance the anti-osteosarcoma effect of DOX in vivo and in vitro. The combination of DOX and EGCG could inhibit the activity of osteoclasts at the tumor-bone interface, thereby preventing tumor-induced osteolysis and blocking the vicious circle. Taken together, EGCG may be used as an adjuvant chemotherapy drug to treat osteosarcoma. • EGCG enhanced DOX-induced proliferation inhibition of 143B Cells. • EGCG in synergy with DOX inhibited osteoclast differentiation and bone resorption in vitro. • EGCG in synergy with DOX inhibited AKT phosphorylation in 143B cells and BMMs. • EGCG could enhance the anti-osteosarcoma and anti- osteolysis effect of DOX in vivo. Osteosarcoma is the most common malignant bone tumor with the pathological essence of osteolysis. Doxorubicin (DOX) is common in adjuvant chemotherapy of osteosarcoma. However, it is required to address the conflict between enhancing its dose to improve efficacy and reducing its complications and chemotherapy resistance. Previous studies have demonstrated that EGCG has an antitumor and osteoclastogenesis inhibition effect. However, the combined effect of EGCG and DOX on osteosarcoma is still unclear. Here, EGCG was demonstrated to enhance the 143B cell proliferation inhibition and anti-osteoclastogenesis effect of DOX in vitro. The results suggest that EGCG in synergy with DOX inhibited AKT phosphorylation, which was involved in 143B cells proliferation and osteoclastogenesis. In vivo, the combination of DOX and EGCG could reduce tumor volume and osteolysis destruction compared with the DOX treatment group. This present study raised the possibility that EGCG may be used as an adjuvant chemotherapy drug to treat osteosarcoma.

A noncoding regulatory RNA Gm31932 induces cell cycle arrest and differentiation in melanoma via the miR-344d-3-5p/Prc1 (and Nuf2) axis

Emerging evidence has shown that long non-coding RNAs (lncRNAs) play an important role in inhibiting tumor cell proliferation and inducing differentiation. In this study, integrative analysis of whole transcriptome sequencing data demonstrated that lncRNA-Gm31932 is significantly decreased in all-trans retinoic acid (ATRA)-induced and sodium 4-phenylbutanoate (PB-4)-induced mouse melanoma B16 cells. Silencing lncRNA-Gm31932 could inhibit B16 cell proliferation, with cell cycle arrest at the G0/G1 phase and obvious differentiation characteristics, e.g., increased cell volume, melanin content and tyrosinase (Tyr) activity. Furthermore, a series of experiments (luciferase reporter assay, RNA pull-down assay, and western blotting) showed that lncRNA-Gm3932 down-regulated Prc1 and Nuf2 by competitively sponging miR-344d-3-5p, which subsequently reduced the expression of cell cycle-related proteins CDK2, CDC2, and Cyclin B1, and increased the expression of P21 and P27. Moreover, silencing lncRNA-Gm31932 could significantly inhibit tumor growth in B16 melanoma-bearing mice. Taken together, these results indicate that as a possible signaling pathway for ATRA and PB-4, lncRNA-Gm31932 can induce cell cycle arrest and differentiation via miR-344d-3-5p/Prc1 (and Nuf2) axis.

N6-Methyladenosine-Related lncRNAs Are Potential Prognostic Biomarkers and Correlated With Tumor Immune Microenvironment in Osteosarcoma.

N6-methyladenosine (m6A) and long non-coding RNAs (lncRNAs) play vital roles in the prognostic value and immune microenvironment of malignant tumors. Here, we constructed a m6A-related lncRNA signature in osteosarcoma samples from TCGA dataset and analyzed the association of the signature with tumor immune microenvironment. m6A-related lncRNAs were identified by performing Pearson’s correlation analysis and were used to construct a novel m6A-related lncRNA signature in osteosarcoma. Validation in testing and entire cohorts confirmed the satisfactory accuracy of the risk signature. Principal-component analysis verifies the grouping ability of the risk signature. Functional enrichment analyses connected immune with the risk signature based on the six m6A-related lncRNAs. When patients were separated into high- and low-risk group based on their risk scores, we found that patients in the high-risk group had lower stromal scores, immune scores, and ESTIMATE scores, while the tumor purity was higher in the high-risk group than that in the low-risk group. As for immune cell infiltration, the proportion of monocytes was significantly higher in the low-risk group than that in the high-risk group. Of the six lncRNAs, AC004812.2 was a protective factor in osteosarcoma and low expression of AC004812.2 predicted worse overall survival. Overexpression of AC004812.2 inhibited 143B cell proliferation and increased the expression levels of IGF2BP1 and YTHDF1. In all, our m6A-related lncRNA signature was a potential prognostic biomarker and correlated with tumor immune microenvironment and immune cell infiltration, and AC004812.2 might be an important regulator of m6A modification and a promising therapeutic target in osteosarcoma.

Physicochemical Properties and Biological Activities of Silver Carp Scale Peptide and Its Nanofiltration Fractions.

To explore the physicochemical properties and biological functions of silver carp scale peptide (SCSP), its molecular-weight fractions SCSP-I, II, and III obtained by nanofiltration were assessed for their solubility, emulsibility, free radical scavenging ability, effect on the proliferation of mouse B16 cells. The results showed that the solubility of each fraction of SCSP was higher than 90%, SCSP-II and III were higher than 95%. The antioxidant powers on ⦁OH, ⦁ and Fe3+ were ranked as SCSP-III > SCSP-II > SCSP-I > SCSP. All fractions of SCSP had no toxic or side effects in mouse B16 melanoma cells experiments in vitro. At a concentration of 0.01 mg/mL, the tyrosinase activity of B16 cells in the SCSP-II fraction was significantly lower than that of the α-arbutin (P < 0.05), at 65.37%. The molecular weight distribution of SCSP was 399–1404 Dalton and 13 peptide sequences were detected. Among them, SCSP-II contained many hydrophobic amino acids, and SCSP-III stood out for combining arginine with hydrophobic amino acids. This may be the reason why the low molecular-weight SCSPs show the strong antioxidant activity and strong tyrosinase inhibition. The work provides a data base for the development of SCSP and increases the possibility of its application.

Soy isoflavone metabolite equol inhibits cancer cell proliferation in a PAP associated domain containing 5-dependent and an estrogen receptor-independent manner

Isoflavone is a species of polyphenol found mainly in soy and soy products. Many studies have demonstrated its estrogen receptor (ER)-dependent action. Equol is an intestinal metabolite of a major soy isoflavone daidzein. We aimed to elucidate the mechanism for ER-independent actions of equol. Equol has been shown to inhibit proliferation of HeLa human cervical cancer cells and mouse melanoma B16 cells in an ER-independent manner. Using functional genetic screening, PAP associated domain containing 5 (PAPD5), which is a non-canonical poly(A) polymerase, was identified as an essential molecule in the ER-independent action. While peroral administration of equol inhibited tumor growth of control B16 cells subcutaneously inoculated in mice, it had little effect on the growth of PAPD5-ablated B16 cells. Intriguingly, equol progressed tumor growth of the PAPD5-ablated human breast cancer MCF-7 cells, which have high ERα expression. Equol has been found to induce polyadenylation of snoRNAs in a PAPD5-depdendent manner. Furthermore, peroral equol administration increased microRNA miR-320a expression in tumors. Together, these results suggest that equol may have a dual effect on ER-positive cancer cells, acting with, antiproliferative activity through PAPD5 and exhibiting proliferative activity via ERα and the former could be associated with miR-320a.

Endostatin inhibits the proliferation and migration of B16 cells by inducing macrophage polarity to M1‑type

Malignant melanoma is a common skin tumor that easily metastasizes and has a poor prognosis. Endostatin is an endogenous vascular endothelial inhibitor that mainly suppresses tumor growth by inhibiting the proliferation of vascular endothelial cells and by reducing the formation of tumor microvessels, however the immunological function of endostatin remains unclear. Previously, we have found that an over‑expression endostatin (pEndostatin) plasmid induced RAW264.7 cells' polarity to M1‑type macrophage. To elucidate the effect of M1‑type macrophages induced by endostatin on melanoma B16 cells, the present study transfected RAW264.7 cells with pEndostatin plasmid and co‑cultured them with B16 cells. Compared with the control group, the expression of matrix metalloproteinase (MMP)‑2, MMP‑9 and proliferating cell nuclear antigen in B16 cells was inhibited by M1‑type macrophages, but cleaved Caspase‑3 and cleaved Caspase‑8 were significantly upregulated and the ratio of Bax/Bcl‑2 was increased. These results indicated that M1 macrophages induced by pEndostatin plasmid inhibited the proliferation and migration of B16 cells and promoted their apoptosis. These findings suggest that the inhibitory effect of endostatin on melanoma is not limited to directly inhibiting tumor microvessel formation, but it may also be related to regulating changes in macrophage polarity.

Structures and anti-melanoma activities of two polysaccharides from Angelica sinensis (Oliv.) Diels

Polysaccharide is one of the necessary macromolecules in life activities, and it is also a very promising natural product for tumor prevention and treatment. In this study, two homogeneous polysaccharides (APS-4I and APS-4II) were isolated from Angelica sinensis (Oliv.) Diels. APS-4I was a linear glucan with molecular weight of 16.1 kDa, which was composed of 88.4% α-1,6-Glcp, 4.1% α-1,2-Glcp, 3.9% α-1,3-Glcp, and 2.8% α-T-Glcp. APS-4II was a novel polysaccharide with molecular weight of 11.1 kDa, which consisted of 55.4% α-1,6-Glcp, 10.4% α-1,3,5-Araf, 8.7% α-T-Araf, 9.2% α-1,5-Araf, 4.0% α-1,3-Araf, 3.6% α-1,4-Galp, and 9.1% β-1,3-Galp. NMR results demonstrated that APS-4II has a backbone composed of →6)-α-Glcp-(1 → 6)-α-Glcp-(1 → 5)-α-Araf-.(1 → 5)-α-Araf-(1 → 3,5)-α-Araf-(1 → 3)-β-Galp-(1 → 3)-β-Galp-(1 → 4)-α-Galp-(1 → 3)-α-Araf-(1 → 3,5)-α-Araf-(1→. Both APS-4I and APS-4II inhibited the tumor growth of B16-bearing mice, and the suppressive effect of APS-4II reached 64.7 ± 7.3%. Meanwhile, there were higher lymphocyte numbers and the levels of IL-2, IFN-γ, and TNF-α in peripheral blood of APS-4II-treated mice than those in APS-4I-treated mice. Furthermore, APS-4II showed a higher inhibitory effect on the proliferation of B16 cells and stronger promoting effects on the proliferation of splenocytes, the phagocytosis of peritoneal macrophages, and the cytotoxicity of NK cells. These results demonstrated that APS-4II could be a promising therapeutic agent for melanoma.