Clinicopathological Significances and Prognostic Value of PPFIA4 in Colorectal Cancer.

Purpose: The PPFIA gene family (PPFIA1, PPFIA2, PPFIA3, and PPFIA4) is associated with multiple human diseases, particularly malignant tumors. However, the expression and prognostic value of the PPFIA family in human colorectal cancers (CRCs) have not been reported. Materials and methods: In this study, several databases, including Oncomine, UALCAN, and the cancer cell line encyclopedia, were used to compare differences in PPFIA1, PPFIA2, PPFIA3, and PPFIA4 expression between normal colon samples and CRCs. The expression levels of these four proteins were used to evaluate the survival of patients with CRC, as determined by the Cancer Genome Atlas Program (TCGA) portal and gene expression profiling interactive analysis (GEPIA) databases. Western blotting and reverse transcription-polymerase chain reaction were performed to detect protein and mRNA levels of PPFIA1, PPFIA3, and PPFIA4, respectively. Immunohistochemical (IHC) staining was used to detect the correlation between PPFIA4 expression and the degree of CRC malignancy. Furthermore, potential miRNAs targeting PPFIA4 in CRCs were studied and confirmed. Results: Bioinformatic analysis showed that the mRNA levels of PPFIA1, PPFIA3, and PPFIA4 were higher in CRC tissue samples than in normal colon tissue. Both mRNA and protein expression of PPFIA1, PPFIA3, and PPFIA4 were increased in the CRC cell lines LoVo and Hct116 compared with the normal colon epithelial cell line. Only PPFIA4 was associated with the prognosis of patients with CRC, which was confirmed by TCGA portal and GEPIA. IHC staining confirmed that the expression of PPFIA4 was higher in CRC tissues than in normal colon tissues and also increased in poorly differentiated CRC tissues and lymph node metastatic foci in comparison with well-differentiated CRC tissues and moderately differentiated CRC tissues. Functional annotation enrichment analysis indicated that the top 100 genes co-expressed with PPFIA4 were enriched in the G-protein coupled peptide receptor activity, leukotrience B4 receptor activity, and peroxisome proliferator-activated receptors and hypoxia-inducible factor-1 signaling pathways. In addition, miR-485-5p negatively regulates the expression of PPFIA4. Conclusion: PPFIA4 expression is associated with the development of CRCs and may be a novel potential prognostic marker for human CRCs.

PLK4 is a key molecule in the formation of PGCCs and promotes invasion and migration of progeny cells derived from PGCCs.

Purpose: Cancer stem cells (CSCs) are the evil source of tumor metastasis and recurrence. Polyploid giant cancer cells (PGCCs) that exhibit the characteristics of CSCs produced daughter cells via asymmetric division. The molecular mechanisms of daughter cells derived from PGCCs with high migration, invasion, and proliferation abilities in colorectal cancer (CRC) are explored in this paper based on the bioinformatics analysis. Materials and Methods: We characterized the expression of CSC-related genes in CRCs by analyzing the mRNAsi of The Cancer Genome Atlas and survival time. Weighted gene co-expression network analysis was performed to identify the modules of the hub and key genes. The migration, invasion, and proliferation abilities of cells, the expression of epithelial-mesenchymal transition (EMT)-related proteins and polo-like kinase 4 (PLK4) were compared in LoVo and Hct116 cells with and without bufalin treatment. In addition, the expression and subcellular location of cell division cycle 25C (CDC25C) in cells before and after PLK4 knockdown were assessed. Results: Eight hub genes were screened out and positively association with mRNAsi in CRCs based on bioinformatic analysis. Among them, checkpoint Kinase-1 (CHEK1), budding uninhibited by benzimidazoles 1 Homolog Beta (BUB1B) and PLK4 were closely associated with the prognosis of CRC patients. Bufalin could induce the formation of PGCCs in LoVo and Hct116 cell lines. PLK4 was overexpressed in PGCCs with progeny cells and progeny cells derived from PGCCs had strong migration and invasion abilities by expressing epithelial-mesenchymal transition (EMT)-related proteins. PLK4 could interact with CDC25C and promote CDC25C phosphorylation which was associated with the formation of PGCCs. Decreasing CDC25C expression in both LoVo and Hct116 PGCCs with progeny cells, while levels of pCDC25C-ser216 and pCDC25C-ser198 were increased in LoVo and decreased in Hct116 PGCCs with progeny cells. pCDC25C-ser216 located in the cytoplasm and pCDC25C-ser198 located in the nucleus in cells after bufalin treatment. Furthermore, expression of CDC25C, pCDC25C-ser216, and pCDC25C-ser198 was downregulated after PLK4 knockdown. Furthermore, the expression level of PLK4 was associated with differentiated degree, and lymph node metastasis in human CRC tissues. Conclusion: PLK4 contributes to the formation of PGCCs by regulating the expression of CDC25C and is associated with the expression and subcellular location of CDC25C, pCDC25C-ser216 and pCDC25C-ser198.

Glycine assists in efficient synthesis of herbal carbon dots with enhanced yield and performance.

As a special type of biomass, herbal medicine often contains a variety of biologically active substances, and taking it as a carbon source, it is expected to produce various types of biologically functional carbon dots (CDs). However, there are few reports in this field, especially in achieving enhanced performance of CDs by improving the utilization efficiency of active substances in medicinal materials. In this work, by adding glycine as an auxiliary agent in the preparation of CDs from herbal medicine (Exocarpium Citri Grandis), the carboxyl and amino groups of the adjuvant provided more reactive sites, which greatly improved the yield of CDs (about 6 times). More importantly, the antioxidant and biological activities of herbal CDs were also improved. By controlling the functional groups of adjuvants, the effects of carboxyl and amino groups in adjuvants on the synthesis of herbal CDs were compared. The results reveal that both carboxyl and amino groups can react with the substances in the carbon source, and the influence of amino groups was greater. After adding glycine, the size of the CDs became larger, resulting from the more abundant functional groups on the carbon skeleton, which was the main reason for the improved performance of the CDs. Finally, the biological activity experiment demonstrated that CDs derived from Exocarpium Citri Grandis and glycine could greatly enhance the vitality of cells and activate immune cells, which are expected to be applied in the field of cell reproduction and biological immunity. The method proposed in this work provides a potential strategy for high-yield preparation of CDs from biomass.

Immunoregulatory Activity of Herbal Tea-Derived Carbon Dots.

The activation of immune cells by immunoregulatory active substances can improve the body immunity. Carbon dots (CDs) with immunoregulatory activity are rarely reported. In this study, transmission electron microscopy results demonstrate the existence of CDs in herbal tea, while Fourier transform infrared and X-ray photoelectron spectroscopy results suggest the participation of polyphenol in herbal tea CD (H-CD) formation. The photoluminescence spectrum has shown that H-CDs have fluorescence emission at 565 nm and exhibit an excitation-dependent property. The toxicity and immunostimulatory activity of H-CDs on mouse macrophage RAW264.7 suggested that H-CDs had no toxicity to RAW264.7 cells. Meanwhile, compared with herbal tea, H-CDs have more obvious effect of promoting the expression of cyclooxygenase-2 (COX-2) and inducible nitric oxide synthase. In addition, the secretion of nitric oxide (NO) was promoted by H-CDs. This work suggests that H-CDs have stronger immunoregulatory function than that of original herbal tea, which provides a direction for the application of phenolic hydroxyl-modified CDs in the biomedical field.

The role of cell division control protein 42 in tumor and non-tumor diseases: A systematic review.

Rho-GTPases control a variety of cellular functions mainly by regulating microtubule and actin dynamics, affecting the cytoskeleton, and are important regulators of the structural plasticity of dendrites and spines. Members of the Rho-GTPase family include Ras-related C3 botulinum toxin substrate 1 (Rac1), RhoA (Ras homologous), and cell division control protein 42 (Cdc42). Cdc42 is involved in the regulation of a variety of tumor and non-tumor diseases through a cascade of multiple signaling pathways. Active Cdc42 can regulate intercellular adhesion, cytoskeleton formation, and cell cycle, thus affecting cell proliferation, transformation, and dynamic balance as well as migration and invasion of tumor cells by regulating the expression of effector proteins. Here we discuss the role of Cdc42 in promoting metastasis, invasion, epithelial-mesenchymal transformation and angiogenesis in malignant tumors. The significant role of Cdc42 in non-tumor diseases is also discussed. Since Cdc42 plays a central role in the development of various diseases, small molecule inhibitors targeting Cdc42 have important clinical significance in the prevention and treatment of these diseases.

Arsenic Trioxide Promotes Tumor Progression by Inducing the Formation of PGCCs and Embryonic Hemoglobin in Colon Cancer Cells.

Arsenic trioxide (ATO) has been used to treat acute promyelocytic leukemia. However, it is not effective in treating solid tumors such as colorectal cancer. We have previously reported that polyploid giant cancer cells (PGCCs) exhibiting the characteristics of cancer stem cells can be generated by various inducers. In this study, ATO was used to induce the formation of PGCCs in LoVo and Hct116 colon cancer cell lines. The migration, invasion, and proliferation abilities of colon cancer cells with and without ATO treatment were assessed by wound-healing, transwell, and plate colony formation assays. The expression of epithelial to mesenchymal transition-related proteins and erythroid differentiation-related proteins in colon cancer cells was further evaluated by western blot and immunocytochemical assays. LoVo and Hct116 cells were transfected with a eukaryotic expression vector for green fluorescent protein (GFP), red fluorescent protein (RFP), H2B-GFP, and H2B-mCherry to study PGCCs formation via cell fusion. WB and ICC assays were performed to assess the expression of cell fusion-related proteins. MG132, small interfering RNA-glial cell missing 1 (GCM1), and chromatin immunoprecipitation-polymerase chain reaction assays were performed to study the role of GCM1/syncytin-1-mediated cell fusion. Clinically, the significance of cell fusion-related proteins and erythroid differentiation-related proteins expression in human colorectal cancer tissues was evaluated. Results of our study showed that ATO induced the formation of PGCCs, and the daughter cells derived from PGCCs gained a mesenchymal phenotype and exhibited strong migration, invasion, and proliferation abilities. PGCCs also produced embryonic hemoglobin-delta and -zeta with strong oxygen-binding ability and erythroid differentiation-related proteins after ATO treatment. In addition, cell fusion was observed during the formation of PGCCs, indicated by the presence of yellow fluorescence via the GCM1/syncytin-1 signaling pathway. Clinically, the expression of cell fusion-related and erythroid differentiation-related proteins gradually increased with the progression of human colorectal cancer tissues. In conclusion, ATO can promote tumor progression by inducing the formation of PGCCs via GCM1/syncytin-1-mediated cell fusion. PGCCs can produce daughter cells with high invasion and migration abilities and embryonic hemoglobin with strong oxygen binding ability, promoting survival of tumor cells in a hypoxic microenvironment.

High Migration and Invasion Ability of PGCCs and Their Daughter Cells Associated With the Nuclear Localization of S100A10 Modified by SUMOylation.

Our previous studies have confirmed that cobalt chloride (CoCl2) or chemoradiotherapy could induce the formation of polyploid tumor giant cells (PGCCs). Polyploid giant cancer cells are a special subpopulation of cancer cells that contribute to solid tumor heterogeneity. The size of PGCC was at least three times larger than regular diploid cancer cells. PGCCs have the properties of cancer stem cells (CSCs) and can express CSC markers CD44 and CD133. Daughter cells derived from PGCCs have strong proliferation, infiltration and migration abilities. However, the detailed molecular mechanism of daughter cells expressing mesenchymal phenotype and displaying strong abilities of proliferation and migration is unclear. As a plasminogen receptor, S100A10 which is closely associated with the invasion and metastasis of malignant tumors, was highly expressed in PGCCs with their daughter cells. In this study, CoCl2 was used to induce the formation of PGCCs in LoVo and HCT116 CRC cells. Cell functional experiments, co-immunoprecipitation, MG132 and ginkgolic acid treatment, western blot, and ChIP-Seq were used to identify the mechanism of S100A10 nuclear location. The proliferation and migration abilities of PGCCs and their daughter cells decreased significantly after S100A10 knockdown. In the control cells, S100A10 was mainly ubiquitinated, while in PGCCs and daughter cells, S100A10 was mainly SUMOylated, which was associated with S100A10 nuclear location. After SUMO1 was inhibited, the nuclear S100A10 in PGCCs and daughter cells decreased, and their proliferation and migration abilities significantly decreased. ChIP-Seq combined with real-time fluorescent quantitative PCR showed that S100A10 regulated the expression of neutrophil defensin 3 (DEFA3), receptor-type tyrosine-protein phosphatase N2 (PTPRN2), and rho guanine nucleotide exchange factor 18 (ARHGEF18), which were associated with actin dynamics and cytoskeleton remodeling. The expression of S100A10 in the nuclei and cytoplasm of rectal cancer after neoadjuvant chemoradiation (nCRT) and liver metastases increased compared with that in rectal cancer without nCRT. Taken together, the expression and nuclear localization of S100A10 modified by SUMOylation were associated with the high proliferation and migration of PGCCs and their daughter cells, and the differentiation, metastases, and relapse of CRCs by regulating the expression of ARHGEF18, PTPRN2, and DEFA3.

Role of Transmembrane 4 L Six Family 1 in the Development and Progression of Cancer.

Transmembrane 4 L six family 1 (TM4SF1) is a protein with four transmembrane domains that belongs to the transmembrane 4 L six family members (TM4SFs). Structurally, TM4SF1 consists of four transmembrane domains (TM1-4), N- and C-terminal intracellular domains, two extracellular domains, a smaller domain between TM1 and TM2, and a larger domain between TM3 and TM4. Within the cell, TM4SF1 is located at the cell surface where it transmits extracellular signals into the cytoplasm. TM4SF1 interacts with tetraspanins, integrin, receptor tyrosine kinases, and other proteins to form tetraspanin-enriched microdomains. This interaction affects the pro-migratory activity of the cells, and thus it plays important roles in the development and progression of cancer. TM4SF1 has been shown to be overexpressed in many malignant tumors, including gliomas; malignant melanomas; and liver, prostate, breast, pancreatic, bladder, colon, lung, gastric, ovarian, and thyroid cancers. TM4SF1 promotes the migration and invasion of cancer cells by inducing epithelial-mesenchymal transition, self-renewal ability, tumor angiogenesis, invadopodia formation, and regulating the related signaling pathway. TM4SF1 is an independent prognostic indicator and biomarker in several cancers. It also promotes drug resistance, which is a major cause of therapeutic failure. These characteristics make TM4SF1 an attractive target for antibody-based immunotherapy. Here, we review the many functions of TM4SF1 in malignant tumors, with the aim to understand the interaction between its expression and the biological behaviors of cancer and to supply a basis for exploring new therapeutic targets.

Molecular Mechanism of Stem Cell Differentiation into Adipocytes and Adipocyte Differentiation of Malignant Tumor.

Adipogenesis is the process through which preadipocytes differentiate into adipocytes. During this process, the preadipocytes cease to proliferate, begin to accumulate lipid droplets, and develop morphologic and biochemical characteristics of mature adipocytes. Mesenchymal stem cells (MSCs) are a type of adult stem cells known for their high plasticity and capacity to generate mesodermal and nonmesodermal tissues. Many mature cell types can be generated from MSCs, including adipocyte, osteocyte, and chondrocyte. The differentiation of stem cells into multiple mature phenotypes is at the basis for tissue regeneration and repair. Cancer stem cells (CSCs) play a very important role in tumor development and have the potential to differentiate into multiple cell lineages. Accumulating evidence has shown that cancer cells can be induced to differentiate into various benign cells, such as adipocytes, fibrocytes, osteoblast, by a variety of small molecular compounds, which may provide new strategies for cancer treatment. Recent studies have reported that tumor cells undergoing epithelial-to-mesenchymal transition can be induced to differentiate into adipocytes. In this review, molecular mechanisms, signal pathways, and the roles of various biological processes in adipose differentiation are summarized. Understanding the molecular mechanism of adipogenesis and adipose differentiation of cancer cells may contribute to cancer treatments that involve inducing differentiation into benign cells.

FGFR2/STAT3 Signaling Pathway Involves in the Development of MMTV-Related Spontaneous Breast Cancer in TA2 Mice.

The Tientsin Albino 2 (TA2) mouse has a high incidence of spontaneous breast cancer (SBC) in the absence of external inducers or carcinogens. The initiation of SBC is related to mouse mammary tumor virus (MMTV) infection and pregnancy. Pathologic analysis showed that breast cancer cells in TA2 mice are triple negative. Our previous study confirmed that fibroblast growth factor receptor 2 (FGFR2) expression increased in SBC tissue compared to that in their corresponding normal breast tissues of TA2 mice. The present study focused on the function of the FGFR2/STAT3 signaling pathway in the initiation of SBC. In this study, the expression of FGF3, FGFR2, STAT3, p-STAT3Tyr705, and p-STAT3Ser727 was detected in serum and normal mammary gland tissues of TA2 mice with different number of pregnancies and SBC. The proliferation, invasiveness, and migration abilities of MA-891 cells from TA2 SBC were compared before and after cryptotanshinone and Stattic treatment. Transient siRNA transfection was used to detect the invasiveness, and migration abilities to avoid the off-targets effects. Downstream protein expression of STAT3 was also detected in MA-891 cells and TA2 xenografts from MA-891 inoculation. In addition, STAT3 expression was analyzed in 139 cases of human breast cancer including 117 cases of non-triple negative breast cancer (non-TNBC) (group I) and 22 cases of triple-negative breast cancer (TNBC) (group II). Results of our study confirmed that MMTV-LTR amplification, and FGFR2, p-STAT3Tyr705, p-STAT3Ser727 expression increased with the number of pregnancies in the breast tissue of TA2 mice and were the highest in SBC. Serum FGF3 expression of SBC was higher than it of TA2 mice with different number of pregnancies. After STAT3 was inhibited, the abilities of proliferation, invasiveness, and migration in MA-891 decreased and the expression levels of STAT3, p-STAT3Ser727, p-STAT3Tyr705, Bcl2, cyclin D1, and c-myc in MA-891 and animal xenografts were also down-regulated. In human breast cancer, STAT3 expression was significantly higher in TNBC than that in non-TNBC. Our results showed that the FGFR2/STAT3 signaling pathway may be related to SBC initiation in TA2 mice. Inhibition of STAT3 can decrease proliferation, invasiveness, and migration in MA-891 cells and the growth of TA2 xenografts.

Different p53 genotypes regulating different phosphorylation sites and subcellular location of CDC25C associated with the formation of polyploid giant cancer cells.

BACKGROUND: Our previous studies have confirmed that cobalt chloride (CoCl2) can induce the formation of polyploid giant cancer cells (PGCCs), which is the key to the heterogeneity of solid tumors. PGCC formation is closely related to the abnormal expression of cell cycle-related proteins and cell fusion. In this study, we investigated the molecular mechanism of PGCCs formation by detecting the expression of cell cycle-related proteins in mutant and wild-type p53 cancer cell lines. METHODS: HEY, BT-549, SKOv3 and MDA-MB-231 cells were treated with CoCl2 and the cell cycle was detected by flow cytometry. The expression and subcellular localization of cell cycle-related proteins, kinases, and P53 were compared before and after CoCl2 treatment. Immunoprecipitation was used to analyze the interacting proteins of pCDC25C-Ser216 and pCDC25C-Ser198. The clinicopathologic significances of these cell cycle-related proteins and protein kinases expression were studied. RESULTS: CoCl2 induced the formation of PGCCs and G2/M arrest. CDC25C, cyclin B1, and CDK1 expressions after CoCl2 treatment were lower than that in control cells. Cytoplasmic CDC25C was degraded by ubiquitin-dependent proteasome. The expression of P53 and phosphokinases including CHK1, CHK2, PLK1, and Aurora A increased after CoCl2 treatment. The expression of pCDC25C-Ser216 and pCDC25C-Ser198 depended upon the genotype of p53. The expressions of cell cycle-related proteins and kinases gradually increased with the development of ovarian cancer and breast cancer. CONCLUSION: CHK1, CHK2-pCDC25C-Ser216-cyclin B1-CDK1, and Aurora A-PLK1-pCDC25C-Ser198-cyclin B1-CDK1 signaling pathways may participate in the formation of PGCCs and different phosphorylation sites of CDC25C may be associated with the genotype of p53.

Clinicopathological Significances of Cancer Stem Cell-Associated HHEX Expression in Breast Cancer.

Aberrant expression of the transcription factor hematopoietic ally expressed homeobox/proline-rich homeodomain (HHEX/PRH) is implicated in numerous cancers. However, the association of HHEX with breast cancer (BC) remains unclear. In this study, HHEX mRNA and protein expression were analyzed using the Oncomine, UALCAN, GEPIA, TCGAportal, and HPA databases. We evaluated the effect of HHEX on clinicopathological parameters using Kaplan-Meier plotter, OncoLnc, TCGAportal, PROGgeneV2, and BC-GenExMiner. Western blotting was performed to compare the level of HHEX in breast samples of Tientsin Albino 2 mice, human breast precancerous lesions, benign breast tumors, and BC. The correlation between HHEX and cancer stem cells was investigated using the GEO (GSE52327 and GSE94865) and GEPIA datasets. Networks between HHEX and survival-related gene marker sets and microRNAs were analyzed using GEPIA, StarBase, and Cytoscape. Results of this study showed that HHEX expression in BC was significantly lower than those in breast precancerous lesions and benign breast tumors at both mRNA and protein levels. BC patients with lower HHEX expression had significantly worse overall survival and disease-free survival. Moreover, HHEX significantly affected the clinicopathology of BC. Specifically, low HHEX expression was correlated with the following groups of patients: age ≤51 years, ER-negative or PR-negative patients, HER-2 positive, triple-negative breast cancer, and basal-like BC. Immunohistochemical analysis of the breast samples showed significant differences of HHEX staining index (P < 0.001) among the three groups. To further investigate the mechanism, we determined the intersection of differentially expressed genes related to BC stem cells and those genes after HHEX expression was altered. This led to the identification of four potentially regulated genes-CXL12, BLNK, PAG1, and LPXN. Using StarBase and km-plotter, the negative regulation of HHEX expression and survival trends, including miR-130b, miR-30e, and miR-301b were joined into miRNA-HHEX-mRNA potential regulatory network. The abilities of proliferation, migration and invasion increased in MDA-MB-231 and BT-549 breast cancer cell lines after HHEX down expression and decreased after HHEX overexpression compared them in the control cells. In conclusion, these data suggest that HHEX expression is downregulated in BC and HHEX may regulate the development of BC through the stem cell-related genes.