Humans ; Proto-Oncogene Proteins p21(ras)/genetics* ; Prognosis ; Biomarkers, Tumor/genetics* ; Mutation/genetics* ; Colorectal Neoplasms/genetics* ; Proto-Oncogene Proteins B-raf/metabolism*

Objective To establish U251 cells with inhibited expression of interferon-γ inducible protein 30 (IFI30), and to investigate the effect of IFI30 on cell biological function as well as its underlying mechanism. Methods Three knockdown sequences which target IFI30 were designed online and 3 small interfering RNAs (siRNA) were synthesized. After transfection, the inhibition efficiency was detected by real-time quantitative PCR. The siRNA sequence with the highest inhibition efficiency was selected to create short hairpin RNA (shRNA) plasmids. The recombinant plasmids and packaging plasmids were co-transfected into HEK293T cells to prepare lentivirus. The glioma U251 cells were transfected with lentivirus, and the positive cells were screened by puromycin. CCK-8 assay, 5-ethyl-2'-deoxyuridine (EdU) and colony formation assays were used to analyze cell proliferation; the flow cytometry was used to analyze cell cycle and apoptosis; the Transwell^TM assay was used to detect cell invasion; the wound-healing assay was employed to detect cell migration, and western blot analysis to detect the protein expresison of cyclin D1, B-cell lymphoma factor 2 (Bcl2), epithelial cadherin (E-cadherin), neural cadherin (N-cadherin), signal transducer and activator of transcription 1 (STAT1). Results The sequence which effectively target IFI30 was screened and U251 cell line capable of inhibiting the IFI30 expression was successfully established. When IFI30 expression was knocked down, the proliferation of U251 cells was inhibited, along with increased ratio of cells in the phase G0/G1, the decreased phase S, the increased rate of cell apoptosis. The cell invasion and migration capabilities was also reduced. The decreased expression of cyclin D1, Bcl2 and N-cadherin were observed in U251 cells, and the expression of E-cadherin and the phosphorylation of STAT1 were found increased. Conclusion Knockdown of IFI30 inhibits the proliferation, invasion and migration of human glioma cell U251 and promotes its apoptosis by activating STAT1.
Humans ; Cyclin D1/genetics* ; HEK293 Cells ; Interferon-gamma ; RNA, Small Interfering ; Apoptosis/genetics* ; Cadherins ; Cell Proliferation/genetics* ; Glioma/genetics* ; Proto-Oncogene Proteins c-bcl-2 ; Oxidoreductases Acting on Sulfur Group Donors ; STAT1 Transcription Factor/genetics*

BACKGROUND: Lung cancer is a major threat to human health. The molecular mechanisms related to the occurrence and development of lung cancer are complex and poorly known. Exploring molecular markers related to the development of lung cancer is helpful to improve the effect of early diagnosis and treatment. Long non-coding RNA (lncRNA) THAP7-AS1 is known to be highly expressed in gastric cancer, but has been less studied in other cancers. The aim of the study is to explore the role and mechanism of methyltransferase-like 3 (METTL3) mediated up-regulation of N6-methyladenosine (m6A) modified lncRNA THAP7-AS1 expression in promoting the development of lung cancer. METHODS: Samples of 120 lung cancer and corresponding paracancerous tissues were collected. LncRNA microarrays were used to analyze differentially expressed lncRNAs. THAP7-AS1 levels were detected in lung cancer, adjacent normal tissues and lung cancer cell lines by quantitative reverse transcription-polymerase chain reaction (qRT-PCR). The diagnostic value of THAP7-AS1 in lung cancer and the relationship between THAP7-AS1 expression and survival rate and clinicopathological parameters were analyzed. Bioinformatics analysis, methylated RNA immunoprecipitation (meRIP), RNA pull-down and RNA-immunoprecipitation (RIP) assay were used to investigate the molecular regulation mechanism of THAP7-AS1. Cell proliferation, migration, invasion and tumorigenesis of SPC-A-1 and NCI-H1299 cells were determined by MTS, colony-formation, scratch, Transwell and xenotransplantation in vivo, respectively. Expression levels of phosphoinositide 3-kinase/protein kenase B (PI3K/AKT) signal pathway related protein were detected by Western blot. RESULTS: Expression levels of THAP7-AS1 were higher in lung cancer tissues and cell lines (P<0.05). THAP7-AS1 has certain diagnostic value in lung cancer [area under the curve (AUC)=0.737], and its expression associated with overall survival rate, tumor size, tumor-node-metastasis (TNM) stage and lymph node metastasis (P<0.05). METTL3-mediated m6A modification enhanced THAP7-AS1 expression. The cell proliferation, migration, invasion and the volume and mass of transplanted tumor were all higher in the THAP7-AS1 group compared with the NC group and sh-NC group of SPC-A-1 and NCI-H1299 cells, while the cell proliferation, migration and invasion were lower in the sh-THAP7-AS1 group (P<0.05). THAP7-AS1 binds specifically to Cullin 4B (CUL4B). The cell proliferation, migration, invasion, and expression levels of phosphatidylinositol-4,5-bisphosphate 3-kinase catalytic subunit alpha (PIK3CA), phosphoinositide-3 kinase, catalytic subunit delta (PIK3CD), phospho-phosphatidylinositol 3-kinase (p-PI3K), phospho-protein kinase B (p-AKT) and phospho-mammalian target of rapamycin (p-mTOR) were higher in the THAP7-AS1 group compared with the Vector group of SPC-A-1 and NCI-H1299 cells (P<0.05). CONCLUSIONS LncRNA THAP7-AS1 is stably expressed through m6A modification mediated by METTL3, and combines with CUL4B to activate PI3K/AKT signal pathway, which promotes the occurrence and development of lung cancer.
Humans ; Lung Neoplasms/pathology* ; RNA, Long Noncoding/metabolism* ; Phosphatidylinositol 3-Kinases/metabolism* ; Up-Regulation ; Proto-Oncogene Proteins c-akt/metabolism* ; Cell Line, Tumor ; Cell Proliferation/genetics* ; Gene Expression Regulation, Neoplastic ; Methyltransferases/metabolism* ; Cullin Proteins/genetics*

OBJECTIVE: To investigate the effect and possible mechanism of hydroxysafflor yellow A (HSYA) on human immortalized keratinocyte cell proliferation and migration. METHODS: HaCaT cells were treated with HSYA. Cell proliferation was detected by the cell counting kit-8 assay, and cell migration was measured using wound healing assay and Transwell migration assay. The mRNA and protein expression levels of heparin-binding epidermal growth factor (EGF)-like growth factor (HBEGF), EGF receptor (EGFR), phosphatidylinositol 3-kinase (PI3K), protein kinase B (AKT), mammalian target of rapamycin (mTOR), and hypoxia-inducible factor-1α (HIF-1α) were detected by quantitative real-time polymerase chain reaction (qRT-PCR) and Western blot, respectively. Circ_0084443-overexpressing HaCaT cells and empty plasmid HaCaT cells were constructed using the lentiviral stable transfection and treated with HSYA. The expression of circ_0084443 was detected by qRT-PCR. RESULTS: HSYA (800 µmol/L) significantly promoted HaCaT cell proliferation and migration (P<0.05 or P<0.01). It also increased the mRNA and protein expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and increased the phosphorylation levels of PI3K and AKT (P<0.05 or P<0.01). Furthermore, HSYA promoted HaCaT cell proliferation and migration via the HBEGF/EGFR and PI3K/AKT/mTOR signaling pathways (P<0.01). Circ_0084443 attenuated the mRNA expression levels of HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α (P<0.05). HSYA inhibited the circ_0084443 expression, further antagonized the inhibition of circ_0084443 on HBEGF, EGFR, PI3K, AKT, mTOR and HIF-1α, and promoted the proliferation of circ_0084443-overexpressing HaCaT cells (P<0.05 or P<0.01). However, HSYA could not influence the inhibitory effect of circ_0084443 on HaCaT cell migration (P>0.05). CONCLUSION HSYA played an accelerative role in HaCaT cell proliferation and migration, which may be attributable to activating HBEGF/EGFR and PI3K/AKT signaling pathways, and had a particular inhibitory effect on the keratinocyte negative regulator circ_0084443.
Humans ; Proto-Oncogene Proteins c-akt/metabolism* ; Phosphatidylinositol 3-Kinase ; Phosphatidylinositol 3-Kinases/metabolism* ; ErbB Receptors/genetics* ; TOR Serine-Threonine Kinases/metabolism* ; Cell Proliferation ; RNA, Messenger/genetics* ; Cell Movement ; Cell Line, Tumor ; Chalcone/analogs & derivatives* ; Quinones

Lung cancer is the malignant tumor with the highest incidence and mortality rate in China, among which non-small cell lung cancer (NSCLC) accounts for about 85%. BRAF mutation occurs about 1.5% to 5.5% in NSCLC patients, while BRAF V600 accounts for about 30% to 50% of all BRAF mutations. The overall prognosis of patients with BRAF-mutation is poor. At present, there are many clinical trials on BRAF-mutation NSCLC and new drugs constantly emerging. However, there is no standardized consensus on the diagnosis and treatment of BRAF-mutation NSCLC in China. The expert group of the Lung Cancer Professional Committee of the Chinese Anti-Cancer Association formulated this consensus by integrating foreign and domestic BRAF-mutation-related guidelines, consensus, and existing clinical trials, and combining with Chinese experts' clinical experience in the diagnosis and treatment of BRAF-mutation NSCLC. This consensus provides systematic recommendations for the clinical diagnosis and treatment process, rational drug choice, and adverse events management of BRAF-mutation NSCLC, aiming to provide reference for the standard of diagnosis and treatment of BRAF-mutation NSCLC.
Humans ; Carcinoma, Non-Small-Cell Lung/genetics* ; Lung Neoplasms/genetics* ; Proto-Oncogene Proteins B-raf/genetics* ; Consensus ; Mutation

OBJECTIVE: To observe the effect of electroacupuncture (EA) on liver protein kinase B (Akt)/forkhead box transcription factor 1 (FoxO1) signaling pathway in Zucker diabetic fatty (ZDF) rats, and to explore the possible mechanism of EA on improving liver insulin resistance of type 2 diabetes mellitus. METHODS: Twelve male 2-month-old ZDF rats were fed with high-fat diet for 4 weeks to establish diabetes model. After modeling, the rats were randomly divided into a model group and an EA group, with 6 rats in each group. In addition, six male Zucker lean (ZL) rats were used as the blank group. The rats in the EA group were treated with EA at bilateral "Zusanli" (ST 36), "Sanyinjiao" (SP 6), "Weiwanxiashu" (EX-B 3), and "Pishu" (BL 20). The ipsilateral "Zusanli" (ST 36) and "Weiwanxiashu" (EX-B 3) were connected to EA device, continuous wave, frequency of 15 Hz, 20 min each time, once a day, six times a week, for a total of 4 weeks. The fasting blood glucose (FBG) in each group was compared before modeling, before intervention and after intervention; the serum levels of insulin (INS) and C-peptide were measured by radioimmunoassay method, and the insulin resistance index (HOMA-IR) was calculated; HE staining method was used to observe the liver tissue morphology; Western blot method was used to detect the protein expression of Akt, FoxO1 and phosphoenolpyruvate carboxykinase (PEPCK) in the liver. RESULTS: Before intervention, compared with the blank group, FBG was increased in the model group and the EA group (P<0.01); after intervention, compared with the model group, FBG in the EA group was decreased (P<0.01). Compared with the blank group, the serum levels of INS and C-peptide, HOMA-IR, and the protein expression of hepatic FoxO1 and PEPCK were increased (P<0.01), while the protein expression of hepatic Akt was decreased (P<0.01) in the model group. Compared with the model group, the serum levels of INS and C-peptide, HOMA-IR, and the protein expression of hepatic FoxO1 and PEPCK were decreased (P<0.01), while the protein expression of hepatic Akt was increased (P<0.01) in the EA group. In the model group, the hepatocytes were structurally disordered and randomly arranged, with a large number of lipid vacuoles in the cytoplasm. In the EA group, the morphology of hepatocytes tended to be normal and lipid vacuoles were decreased. CONCLUSION EA could reduce FBG and HOMA-IR in ZDF rats, improve liver insulin resistance, which may be related to regulating Akt/FoxO1 signaling pathway.
Male ; Animals ; Rats ; Rats, Zucker ; Proto-Oncogene Proteins c-akt/genetics* ; Diabetes Mellitus, Type 2/therapy* ; Insulin Resistance ; C-Peptide ; Electroacupuncture ; Liver ; Signal Transduction ; Insulin ; Lipids

Humans ; Receptors, Chimeric Antigen/therapeutic use* ; DNA-Binding Proteins/genetics* ; Precursor B-Cell Lymphoblastic Leukemia-Lymphoma ; Proto-Oncogene Proteins c-bcl-2/genetics* ; Cell- and Tissue-Based Therapy ; Oncogene Proteins, Fusion/genetics* ; Translocation, Genetic

Objective: To summarize the pathological diagnosis, clinical features, treatment methods and outcomes of pediatric-type follicular lymphoma (PTFL). Methods: Clinical data including the pathology, clinical features, treatment methods, and follow-up results of 9 PTFL patients admitted to Henan Cancer Hospital from February 2017 to February 2023 were analyzed retrospectively. Results: The age of onset in 9 children was 6 to 18 years, all the patients were males. The clinical manifestation was local painless lymph node enlargement in the head and neck, with a stage of Ⅰ-Ⅱ. The histomorphological characteristics of PTFL were similar to those of classic follicular lymphoma (FL). The germinal center of most follicles were enlarged, the mantle zone disappeared, centroblasts were easily visible, and the histological grade were mostly grade Ⅲ, which may be accompanied by the "starry sky" phenomenon. Monoclonal peaks can be seen in B cell clonal rearrangements (BCR). Immunohistochemistry (IHC) showed CD20 positive, CD10 positive, Bcl-6 positive, Bcl-2 negative, C-myc negative, and Ki-67 was 70%-95%. Fluorescence in situ hybridization (FISH) test was negative for t (14, 18), Bcl-2 translocation, and C-myc translocation. Six cases underwent surgical resection, and 3 cases underwent surgical resection combined with chemotherapy. Up to February 2023, with a follow-up time of 45 to 72 months, all children survived without any recurrence and were in a complete remission state. Conclusions: PTFL is mainly characterized by adolescent male onset, with early clinical manifestations and pathological manifestations of high-level histological status, high proliferation index, and lack of t (14; 18)/Bcl-2 translocation and Bcl-2 expression. It is mainly treated by localized surgical excision and has a good prognosis.
Child ; Adolescent ; Humans ; Male ; Female ; Lymphoma, Follicular/pathology* ; Lymphoma, B-Cell/pathology* ; In Situ Hybridization, Fluorescence ; Retrospective Studies ; Proto-Oncogene Proteins c-bcl-2/genetics*

Objective: To investigate the effects of regenerating islet-derived protein 3A (REG3A) on the proliferation and invasion of glioma cells and its molecular mechanism. Methods: Five low-grade, five high-grade glioma tissues and ten adjacent tissues from glioma patients who underwent surgery at Linyi People's Hospital from October 17, 2017 to October 18, 2018 were collected. Human glioma cell lines (SF295, U251, TG905, A172, CRT) and a primary glioma cell line PT-1 were cultured in vitro. The protein and mRNA expressions of REG3A in these tissues and glioma cell lines were detected by Western blot and reverse transcription-quantitative real-time polymerase chain reaction (RT-qPCR). SF295 cells were infected with lentivirus and labeled as REG3A plasmid transfection group, and the TG905 cells were transfected with si-REG3A by liposome transfection reagent and labeled as si-REG3A transfection group. At the same time, the empty transfection control and blank control groups were set up. Glioma cells were treated with REG3A recombinant protein alone or in combination with Akt1/2 inhibitors. Cell counting kit-8 (CCK-8) and cell scratch assay were used to detect cell proliferation and invasion, respectively. Western blot was used to detect the protein expression of N-cadherin, vimentin and phosphorylation of Akt (p-Akt) in REG3A overexpressed and knockdown glioma cells. Results: RT-qPCR results showed that the mRNA expression levels of REG3A in glioma cells in each group were U251 (2.129±0.13), TG905 (2.22±0.59), CRT (5.02±0.31), A172 (6.62±1.34) and PT-1 (9.18±0.61), respectively, higher than its expression in SF295 cells (1.00±0.18, P<0.001). The mRNA expression level of REG3A in high-grade glioma tissue samples (3.18±2.92) was higher than that in the control group (1.00±1.14, P=0.031) and low-grade glioma group (0.90±0.67, P=0.014). The results of western blot and immunohistochemical staining were consistent with that of RT-qPCR. The migration rate of cells in si-REG3A transfection group [(60.57±5.30)%] was lower than that of the empty transfection group [(84.18±13.63)% (P=0.038)] and blank control group [(79.65±12.09)% (P=0.076)]. The results of the scratch experiment showed that the migration rate of cells in REG3A plasmid transfected cells in the SF295 group was (96.05±6.41)%, which was significantly higher than that of empty transfected cells [(74.47±8.23)%, P=0.021)]. REG3A recombinant protein could up-regulate the expression of N-cadherin, vimentin and p-Akt in SF295 cells. Compared with the control group [(100.00±2.53)%], the proliferation rate in the REG3A recombinant protein group [(117.70±10.24)%] was significantly up-regulated, and the proliferation rate in the REG3A recombinant protein+ Akt inhibitor group [(98.31±3.64)%] was significantly lower than that of the REG3A recombinant protein group (P=0.017). The migration rate of the REG3A recombinant protein+ Akt inhibitor group was (63.35±4.06)%, which was significantly lower than (89.26±11.07)% of the REG3A recombinant protein group (P=0.019). Conclusion: REG3A can promote the proliferation and invasion of human glioma cells by activating the PI3K/Akt signaling pathway.
Humans ; Cell Line, Tumor ; Cell Movement/genetics* ; Cell Proliferation/genetics* ; Glioma/genetics* ; Phosphatidylinositol 3-Kinases/metabolism* ; Protein Kinase Inhibitors ; Proto-Oncogene Proteins c-akt/metabolism* ; RNA, Messenger/metabolism* ; Signal Transduction ; Vimentin/metabolism*

Objective: PD-1/PD-L1 immune checkpoint treatment is effective for some triple-negative breast cancer populations with PD-L1 expression, but the response rate is still not satisfactory. This study aims to explore the mechanism of drug resistance to breast cancer anti-PD-1 therapies and the strategies for overcoming the resistance to PD-1therapies. Methods: By constructing a human triple-negative breast cancer drug-resistant cell line called BT-549R5 and a mouse breast cancer drug-resistant cell line called 4T1R3, and applying the whole-gene shRNA library screening, candidate drug resistance-associated molecules were obtained and verified by cytological experiments. The expression of Tyro3, Axl and MerTK of the TAM family in the 4T1R3 group was tested using the Western blot method. The down-regulation of CDK9 on the effect of T cells killing the BT-549R5 cells was observed through T cell killing tests, while the down-regulation of Tyro3 and CDK9 on the effect of anti-PD-1 therapies for transplanted breast tumors was observed in mouse tumor formation experiments. Results: The cell lines and animal models of breast cancer resistant to PD-1 treatment were successfully constructed. Tyro3, Axl and MerTK were highly expressed in 4T1R3 cells. Whole genome sequencing showed that Tyro3 and CDK9 were highly expressed in BT-549R5 cells. T cell killing experiment showed that the survival rate of BT-549R5 cells in the CDK9 down-regulated group and the control group decreased gradually with the increase of T cells, but the survival rate of BT-549R5 cells in the CDK9 down-regulated group decreased rapidly. Tumor formation experiment in mice showed that under anti-PD-1 treatment, the transplanted tumor in the 4T1R3 cell group grew rapidly compared with the 4T1 cell group (P<0.05), and the tumor volume of the 4T1R3 group was larger than that of the 4T1 group on Day 20. Nevertheless, the tumor growth rates in the CDK9-knockdown 4T1R3 cell group and the Tyro3-knockdown 4T1R3 cell group were similar to that of the 4T1 cell group, and the tumor volumes at day 20 were signiference lower than that of 4T1R3 cell group(P<0.05). Conclusions: Tyro3 and CDK9 are associated with the drug resistance to anti-PD-1 therapies for breast cancer. Inhibiting the expression of Tyro3 and CDK9 can reverse the drug resistance to breast cancer treatment.
Humans ; Animals ; Mice ; c-Mer Tyrosine Kinase/metabolism* ; Receptor Protein-Tyrosine Kinases/genetics* ; Axl Receptor Tyrosine Kinase ; Proto-Oncogene Proteins/metabolism* ; B7-H1 Antigen/genetics* ; Triple Negative Breast Neoplasms/genetics* ; Drug Resistance, Neoplasm ; Biomarkers ; Cell Line, Tumor ; Cyclin-Dependent Kinase 9