Effect of miR-216a-5p on radiosensitivity of liver cancer cells via targeting KLF12 / 中华放射肿瘤学杂志

Objective:To evaluate the effect of miR-216a-5p on the radiosensitivity of liver cancer cells via targeting Krüppel-like transcription factors 12 (KLF12).Methods:Real-time reverse transcription PCR (RT-qPCR) was used to detect miR-216a-5p, KLF12 mRNA levels in normal human liver cells L-02 cells, and human hepatoma cells Huh7, HepG2, Hep3B and MHCC-97H cell lines. HepG2 cells were divided into 0, 2, 4, 6 and 8 Gy irradiation groups, and miR-216a-5p, KLF12 mRNA levels were compared among different groups. HepG2 cells overexpressing miR-216a-5p and / or KLF12 were constructed by plasmid transfection and divided into the miR-NC group (control group), miR-216a-5p group, miR-216a-5p+NC group, miR-216a-5p+KLF12 group, IR+miR-NC group, IR+miR-216a-5p group, IR+miR-216a-5p+NC group, IR+miR-216a-5p+KLF12 group, respectively. miR-216a-5p, KLF12 mRNA levels were compared among different groups. Clone formation assay was used to detect cell radiosensitivity. CCK-8 assay was employed to detect cell proliferation ability. Flow cytometry was adopted to detect cell apoptosis. Single factor ANOVA was used for inter group comparisons. LSD- t test was used for pairwise comparison. Results:Compared with L-02 cells, KLF12 mRNA levels were up-regulated, whereas miR-216a-5p levels were down-regulated in liver cancer cell lines (all P<0.05). Compared with 0 Gy group, KLF12 mRNA levels were down-regulated, whereas miR-216a-5p levels were up-regulated in 2, 4, 6, and 8 Gy groups (all P<0.05). Overexpression of miR-216a-5p or radiation therapy alone could enhance cell radiosensitivity and apoptosis levels, and reduce cell proliferation ability (all P<0.05). Simultaneous radiation treatment with overexpression of miR-216a-5p exerted more significant effects on cells (all P<0.05). Overexpression of KLF12 could partially reverse the aforementioned effects of overexpression of miR-216a-5p ( P<0.05). Conclusion:MiR-216a-5p reduces cell proliferation ability, enhances cell radiosensitivity and apoptosis via regulating KLF12.

M6A-mediated EGLN3 and FOSL2 enhance radioresistance in nasopharyngeal carcinoma / 中华放射肿瘤学杂志

Objective:To screen and verify the key radioresistance genes regulated by m6A methylation in nasopharyngeal carcinoma (NPC) based on the chip data and cell experiments.Methods:The microarray data of NPC radioresistance genes, m6A regulated genes and mRNA expression profiles of NPC genes were downloaded from Gene Expression Omnibus (GEO) database. The differential genes were screened and statistically analyzed by R software. The biological processes, signal pathways and interaction networks of these genes were analyzed by bioinformatics. The m6A regulatory factors were knocked down and the radioresistant strains were constructed. The above m6A differential radioresistant genes of NPC were screened and verified by real-time reverse transcription PCR (qRT-PCR) and Western blot. The m6A modification of screened genes and their direct binding ability with methyltransferase 3 (METTL3) were verified by methylated RNA immunoprecipitation qPCR (MeRIP-qPCR). The siRNA of selected genes was transfected into NPC cells, and after treatment with ionizing radiation, cell proliferation was detected by CCK-8 assay and EdU, apoptosis and cell cycle were detected by flow cytometry, and radiosensitivity was detected by clone formation assay. The trend of differences in the abundance of Fe 2+ and lipid peroxidation between the control and EGLN3 knockdown groups after ionizing radiation treatment was compared by paired t-test. Results:Chip data GSE48501 intersected with GSE200792 and GSE53819 to obtain 6 differential genes, including EGLN3, FOSL2, ADM, JUN, VEGFA and PRDM1. The target genes of EGLN3 and FOSL2 were further screened by TNMplot and KMplot, etc. The mRNA of the target genes directly bound to METTL3 and were subjected to its mediated modification of m6A. The target genes were up-regulated in the parental cells after irradiation in a dose and time gradient manner, which were also significantly up-regulated in radioresistant cells. After EGLN3 and FOSL2 were down regulated, the proliferation activity of NPC cells was more significantly decreased after irradiation, and the radiosensitization ratio was statistically significant compared with that of NPC cells without EGLN3 and FOSL2 down-regulation. After irradiation, EGLN3 down-regulated NPC cells significantly down-regulated glutathione peroxidase 4 (GPX4) expression, increased the abundance of Fe 2+ and lipid peroxidation, which played a role in radiosensitization by inducing ferroptosis. Conclusions:EGLN3 and FOSL2 play a role in radioresistance in NPC through METTL3 mediated m6A methylation. Down-regulation of EGLN3 combined with ionizing radiation can increase the intracellular Fe 2+ abundance and lipid peroxidation and down-reuglate the expression of GPX4 in NPC cells, which can enhance radiosensitization for NPC radiotherapy by the ferroptosis pathway.

Effect of lncRNA NEAT1 on radiosensitivity in breast cancer cells by regulating miR-149-5p/GPT2 axis / 中华放射肿瘤学杂志

Objective:To evaluate the effect of long non-coding RNA (lncRNA) nuclear enriched abundant transcript 1 (NEAT1) on the radiosensitivity in breast cancer cells by regulating the miR-149-5p/ glutamic pyruvic transaminase 2 (GPT2) axis.Methods:Real-time reverse transcription PCR (RT-qPCR) was used to detect NEAT1, miR-149-5p and glutamic pyruvic transaminase 2 (GPT2) mRNA levels in human breast cells MCF-10A, and human breast cancer cells MCF-7, MDA-MB-231 and MDA-MB-468, respectively. MCF-7 cells were divided into 0, 2, 4, 6 and 8 Gy irradiation groups. MCF-7 cells were divided into NEAT1 knockdown (si-NEAT1) group and control (si-NC) group, NEAT1 knockdown +miR-149-5p knockdown (si-NEAT1+anti-miR-149-5p) group and control (si-NEAT1+anti-miR-NC) group, NEAT1 knockdown + GPT2 overexpression (si-NEAT1+GPT2) group and control (si-NEAT1+NC) group. On the basis of the above grouping, irradiate each group of cells with 4 Gy radiation for 2 h, denoted as IR+si-NEAT1, IR+si-NC, IR+si-NEAT1+anti-miR-149-5p, IR+si-NEAT1+anti-miR-NC, IR+si-NEAT1+GPT2, IR+si-NEAT1+NC groups. Subsequently, MCF-7 cells were irradiated at a dose of 4 Gy and divided into the IR+si-NEAT1, IR+si-NC, IR+si-NEAT1+anti-miR-149-5p, IR+si-NEAT1+anti-miR-NC, IR+si-NEAT1+GPT2 and IR+si-NEAT1+NC groups. RT-qPCR was used to detect NEAT1, miR-149-5p, GPT2 mRNA levels in cells. Colony formation assay was used to detect cell radiosensitivity. CCK-8 assay was adopted to detect cell proliferation ability. The binding sites of NEAT1 and miR-149-5p were predicted by StarBase database. The binding sites of miR-149-5p and GPT2 were predicted by Targetscan database, and validated by dual luciferase assay. Single factor ANOVA was used for inter-group comparisons. LSD- t test was used for pairwise comparison. Results:Compared with MCF-10A cells, NEAT1 and GPT2 mRNA levels in cell lines were up-regulated, whereas miR-149-5p level was down-regulated (all P<0.05). Compared with the 0 Gy dose group, NEAT1 and GPT2 mRNA levels were down-regulated, while miR-149-5p levels were up-regulated in the 2, 4, 6, and 8 Gy dose groups (all P<0.05). Knockdown of NEAT1 expression or radiation alone could enhance cell radiosensitivity, and reduce cell proliferation ability (all P<0.05). Simultaneous radiation treatment with knockdown of NEAT1 expression could strengthen the above effects upon cells (all P<0.05). Knockdown of miR-149-5p expression or overexpression of GPT2 could partially reverse the aforementioned effects of knockdown of NEAT1 expression (all P<0.05). Conclusion:Knockdown of NEAT1 expression enhances breast cancer cell radiosensitivity, and reduces cell proliferation ability by regulating the miR-149-5p/GPT2 signal axis.

Effect of Survivin Down-Regulation by Egr1-Survivin shRNA Combined with Radiotherapy on Radiosensitivity of Esophageal Squamous Carcinoma / Efecto de la Regulación Negativa de Survivina por shRNA de Egr1-Survivina Combinado con Radioterapia sobre la Radiosensibilidad del Carcinoma Escamoso de Esófago

SUMMARY: This study is to investigate the effect of survivin down-regulation by Egr1-survivin shRNA combined with radiotherapy on the apoptosis and radiosensitivity of esophageal squamous cell carcinoma ECA109 and KYSE150 cells. ECA109 and KYSE150 cells were transfected with Egr1-survivin shRNA, and then treated with radiotherapy. After 24 h, the mRNA and protein levels of Egr1-survivin were detected by qPCR and Western-Blot. Cell cycle and apoptosis were detected by flow cytometry. Western blot also detected levels of cleavaged Caspase 3 and Caspase 9. YM155 was used as a positive control to inhibit survivin expression. The levels of survivin mRNA and protein in ECA109 and KYSE150 cells treated with Egr1-survivin shRNA combined with radiotherapy were significantly lower than those of the blank control group, the empty vector control group, and, the YM155 + radiotherapy group (P<0.05). Meanwhile, after survivin down-regulation, the ratio of G2 to S phase of ECA109 and KYSE150 cells increased significantly, leading to significant G2 and S phase arrest. Additionally, apoptosis of ECA109 and KYSE150 cells increased significantly (P <0.01). Further, protein levels of cleavaged Caspase 3 and Caspase 9 significantly increased in Egr1-survivin shRNA combined with radiotherapy group. Egr1-survivin shRNA combined with radiotherapy can down-regulate survivin expression, which further increases the apoptosis, and enhances the radiosensitivity of ECA109 and KYSE150 cells.

Este estudio tuvo como objetivo investigar el efecto de la regulación negativa de survivina por el shRNA de Egr1-survivina combinado con radioterapia sobre la apoptosis y la radiosensibilidad del carcinoma de células escamosas de esófago Células ECA109 y KYSE150. Las células ECA109 y KYSE150 se transfectaron con shRNA de survivina Egr1 y luego se trataron con radioterapia. Después de 24 h, los niveles de ARNm y proteína de Egr1-survivina se detectaron mediante qPCR y Western-Blot. El ciclo celular y la apoptosis se detectaron mediante citometría de flujo. La transferencia Western también detectó niveles de Caspasa 3 y Caspasa 9 escindidas. Se usó YM155 como control positivo para inhibir la expresión de survivina. Los niveles de ARNm y proteína de survivina en células ECA109 y KYSE150 tratadas con shRNA de survivina Egr1 combinado con radioterapia fueron significativamente más bajos que los del grupo control en blanco, el grupo control de vector vacío y el grupo de radioterapia YM155 + (P <0,05). Mientras tanto, después de la regulación negativa de survivina, la proporción entre las fases G2 y S de las células ECA109 y KYSE150 aumentó significativamente, lo que llevó a una detención significativa de las fases G2 y S. Además, la apoptosis de las células ECA109 y KYSE150 aumentó significativamente (P <0,01). Además, los niveles de proteína de Caspasa 3 y Caspasa 9 escindidas aumentaron significativamente en el shRNA de Egr1- survivina combinado con el grupo de radioterapia. El shRNA de survivina de Egr1 combinado con radioterapia puede regular negativamente la expresión de survivina, lo que aumenta aún más la apoptosis y mejora la radiosensibilidad de las células ECA109 y KYSE150.

Effect of miR-432-5p targeting DDX41 on proliferation,metastasis,and radiosensitivity of gastric cancer cells / 国际检验医学杂志

Objective To investigate the effects of microRNA(miR)-432-5p on the proliferation,metasta-sis and radiosensitivity of gastric cancer cells and the targeted regulatory relationship with DEAD-box helicase 41(DDX41)in this process.Methods The expressions of miR-432-5p and DDX41 in different gastric cancer cells were detected by quantitative real-time PCR(qRT-PCR).HGC-27 cells were transfected and divided into NC group(not transfected with plasmid),miR-NC group(transfected with miR-432-5p mimics negative con-trol),miR-432-5p group(transfected with miR-432-5p mimics),miR-432-5p+pcDNA-NC group(transfection miR-432-5p and pcDNA-DDX41 negative control)and miR-432-5p+pcDNA-DDX41 group(transfection miR-432-5p mimics and pcDNA-DDX41).After 48 h,qRT-PCR was used to detect the expression of miR-432-5p and DDX41 in each group of cells,MTT assay was used to detect the proliferation activity of each group of cells,Transwell assay was used to detect the migration ability of each group of cells,colony formation assay and flow cytometry were used to detect the radiosensitivity of each group of cells.The dual luciferase reporter gene assay was used to verify the targeting relationship between miR-432-5p and DDX41.The tumor growth of nude mice was detected,and the expression of DDX41 protein in tumor tissues was detected by Western blot.Results Compared with normal cells,cancer of the stomach cells miR-432-5p levels were decreased(P<0.05),DDX41 mRNA levels were increased(P<0.05).Compared with NC group and miR-NC group,the cell survival rate and DDX41 mRNA level in miR-432-5p group were decreased,the number of migrating cells was decreased(P<0.05),the miR-432-5p level was increased,and the radiosensitivity was enhanced(P<0.05).Overexpression of DDX41 reversed the effects of miR-432-5p on proliferation,migration and radiosensitivity of gastric cancer cells(P<0.05).Compared with NC group and miR-NC group,the volume and weight of tumor tissue and the expression of DDX41 protein in miR-432-5p group were significantly decreased(P<0.05).Con-clusion miR-432-5p through targeted cut DDX41 to inhibit the proliferation and metastasis of gastric cancer cells,enhance cell radiation sensitivity.

Effects of over-expression of E2F transcription factor 1 on radiosensitivity glioma U251 cells / 中华行为医学与脑科学杂志

Objective:To investigate the effects of over-expression of E2F transcription factor 1 (E2F1) on proliferation, invasion, apoptosis and radiosensitivity of glioma cell U251.Methods:Real-time quantitative PCR (qRT-PCR) were used to detect the differential expression of E2F1 mRNA in glioma cells LN18, SW1088, U251 and normal brain glial cells. The stable over-expression of E2F1 plasmid was constructed and transfected into U251 cells. qRT-PCR and Western blot test were used to detect the expression of E2F1, pituitary tumor transforming gene 1(PTTG1), C-Myc, B-cell lymphoma-2 (Bcl-2), Bcl2-associated X (Bax) mRNA and protein expression in the control group and E2F1 over-expression group.U251 cells were divided into control group(no X-ray irradiation), irradiation group(6 Gy dose of X-ray), and irradiation + E2F1 over-expression group(transfected with E2F1 first, then irradiated by 6 Gy of X-ray). Cell proliferation ability was detected by cell counting Kit-8(CCK-8) cell viability detection reagent, and cell invasion and migration ability were detected by Transwell chamber. Apoptosis and cell cycle were detected by flow cytometry.GraphPad Prism 8.0 was used for data analysis.The statistical methods were one-way ANOVA and independent sample t-test. Results:qRT-PCR showed that there was statistical difference in the mRNA levels of E2F1( F=201.92, P<0.05) in different cell lines.The expression levels of E2F1 mRNA in LN18(4.04±0.29), SW1088(3.19±0.16)and U251(4.66±0.20) cells were higher than those in HEB(1.02±0.07)cells ( q=27.00, 19.40, 32.52, all P<0.05). After successfully constructing U251 cells with stable over-expression of E2F1 plasmid, qRT-PCR and Western blot detection results showed that: the mRNA and protein levels of E2F1, PTTG1, C-Myc and Bcl-2 in E2F1 over-expression group were higher than those in control group ( t=77.16, 57.88, 4.63, 51.13, 7.50, 70.85, 8.38, 48.81, all P<0.05). Bax mRNA(0.20±0.01) and protein(0.66±0.01) levels were lower than those in control group((1.00±0.02), (0.94±0.01)), and the differences were statistically significant ( t=1.74, 54.65, both P<0.05). After X-ray irradiation (6 Gy), CCK8 detection results showed: the proliferation ability of the three groups at 24, 48, 72 and 96 h were significantly different ( F=95.41, 187.53, 1 158.49, 7 883.78, all P<0.05). The proliferation capacity of the irradiation group were lower than those of the control group at 24, 48, 72 and 96 h ( q=19.51, 27.20, 66.60, 174.9, all P<0.05). The proliferation capacity of irradiation + E2F1 over-expression group at 24, 48, 72 and 96 h were higher than those of irradiation group ( q=10.63, 10.81, 21.11, 60.90, all P<0.05). Transwell assay results showed that there were significant differences in cell invasion and migration ability among the three groups ( F=315.38, 681.10, both P<0.05). The invasion and migration ability of cells in the irradiation group were lower than those in the control group ( q=35.09, 12.76, both P<0.05), and the invasion and migration ability of cells in the irradiation + E2F1 over-expression group were higher than those in the irradiation group ( q=52.06, 22.81, both P<0.05). Flow cytometry showed that there were significant differences in apoptosis rate and percentage of cells in each cycle among the three groups ( F=667.63, 3 213.30, 3 011.26, 861.98, all P<0.05). The percentage of the apoptosis rate, S phase and G2 phase cells in the irradiation group were higher than those in the control group ( q=51.10, 89.39, 51.82, all P<0.05), while the percentage of G1 phase cells in the irradiation group was lower than that in the control group ( q=141.2, P<0.05). The apoptosis rate and percentage of S phase and G2 phase cells in the irradiation + E2F1 over-expression group were lower than those in the irradiation group ( q=18.87, 41.42, 29.31, all P<0.05), while the number of G1 phase cells in the irradiation + E2F1 over-expression group was lower than that in the irradiation group ( q=70.73, P<0.05). Conclusion:Over-expression of E2F1 can reduce the radiosensitivity of glioma U251 cells by regulating the expression of mRNA and protein of genes related to cell cycle and apoptosis, and E2F1 may be involved in the radioresistance of glioma cells.

The radiosensitization effect of STING agonist on cutaneous melanoma cells / 中华放射医学与防护杂志

Objective:To investigate the radiosensitizing effect and underlying mechanism of STING agonist (c-di-AMP) on cutaneous melanoma cells.Methods:Human cutaneous melanoma cells (A375) were divided into four groups: the control group, 10 μmol/L c-di-AMP group, X-ray irradiation group and X-ray irradiation combined with c-di-AMP group. The radiosensitizing effect of c-di-AMP on A375 cells was detected by CCK-8-based viability assay, lactate dehydrogenase (LDH) release assay, flow cytometry-based apoptosis assay, and colony formation assay. Western blot analysis was used to determine the expressions of cell death-related proteins.Results:In combination with 10 Gy X-ray irradiation, 10 μmol/L c-di-AMP showed significant radiosensitization effect in A375 cells, which was evidenced by decreased cell activity ( t=5.11, P<0.05), increased cytotoxicity ( t=10.15, P<0.05) and cell apoptosis ( t=4.41, P<0.05) and reduced clone viability( t=6.30, 3.55, 5.45, 3.55, P<0.05). The calculated radiosensitization ratio of c-di-AMP to A375 cells was 1.88. Moreover, 10 μmol/L c-di-AMP further increased the expressions of cell death-related proteins induced by radiation in A375 cells. Conclusions:The STING agonist c-di-AMP can be used as a radiosensitizer for cutaneous melanoma, which may provide a novel strategy for radiotherapy.

Effect of down-regulation of FABP5 on radiation damage of human keratinocytes / 中华放射医学与防护杂志

Objective:To investigate the effects of down-regulation of FABP5 (fatty acid binding protein 5) on radiation damage of skin cells, and explore underlying mechanism.Methods:A lentiviral vector with down-regulated FABP5 was constructed to infect human immortalized keratinocytes (HaCaT) cells, and the transfection efficiency was examined. The HaCaT cells were divided into blank control group, FABP5 down-regulation group (FABP5), radiation group (IR), and FABP5 down-regulation combined with radiation group (FABP5+ IR). After 6 MV X-ray radiation, cell proliferation viability was measured by CCK-8 assay, cell migration was detected by scratch assay, apoptosis was analyzed by flow cytometry, radiosensitivity was evaluated by cloning formation assay, and the cellular protein expressions of PARP1, γ-H2AX, AKT and p-AKT were detected by Western blot.Results:FABP5 was successfully knocked-down in both RNA level ( t=25.14, P<0.05) and protein level ( t=20.06, P<0.05). The down-regulation of FABP5 decreased the abilities of cells proliferation ( t=3.55, 5.88, 3.18, P<0.05) and migration ( t=15.44, P<0.05), but increased cell resistance to irradiation with a radiosensitization ratio of 0.782. The apoptosis rate of FABP5+ IR group was significantly lower than IR group (22.05±6.71)% vs. (9.82±1.45)%, t=3.08, P<0.05. The protein levels of PARP1 and γ-H2AX in FABP5+ IR group were also lower than those in the IR group 0.04±0.04, 0.11±0.06, 0.26±0.11, 0.22±0.07, 0.21±0.10, 0.52±0.22, 0.57±0.06, 0.43±0.02( t=2.83, 3.07, 4.50, 5.33, P<0.05), while the protein level of p-Akt in FABP5+ IR group was higher than that in IR group ( t=-16.24—3.02, P<0.05). Conclusions:Down-regulation of FABP5 inhibited cell proliferation and migration, increased radioresistance, and reduced radiation-induced apoptosis and DNA damage of skin cells probably through PI3K/AKT signaling pathway.

Effects of HMGB1 on clinical prognosis of esophagus squamous cell carcinoma patients after chemoradiotherapy and the radiosensitivity of xenograft in nude mice / 中华放射肿瘤学杂志

Objective:To evaluate the effects of high mobility group protein box 1 (HMGB1) on clinical prognosis of esophagus squamous cell carcinoma (ESCC) patients treated with chemoradiotherapy and the radiosensitivity of xenograft in nude mice.Methods:A total of 90 endoscopic biopsy specimens were obtained from ESCC patients treated with chemoradiotherapy. The expression level of HMGB1 was determined by immunohistochemical staining. High expression level was defined when staining was observed on ≥50% of the tumor cells. All patients were divided into the high expression group ( n=48) and low expression group ( n=42), and their survival information was retrospectively analyzed. Cell transfection was performed with the plasmid carrying human HMGB1-shRNA to knockdown HMGB1 expression in ECA109 cells and xenograft mouse models were established. The tumor volume and mass were calculated after irradiation with a dose of 15 Gy. The cell apoptosis in xenograft tissues were detected. Survival analysis was performed using Kaplan-Meier method. Univariate prognostic analysis was conducted by log-rank test. Intergroup comparison was performed by analysis of variance (ANOVA). Results:The expression level of HMGB1 was significantly associated with gross tumor volume, longest diameter of tumor, T staging and distant metastasis ( χ2=9.663, 5.625, 4.068, 7.146, all P<0.05). In the low expression group, the overall survival (OS) ( χ2=4.826, P=0.028), progression-free survival (PFS) ( χ2=4.390, P=0.036) were longer compared with that in the high expression group. Further analysis of HMGB1-high expression patients showed that the radiation dose and the combination of chemoradiotherapy did not significantly affect the OS or PFS of ESCC patients. We observed that knockdown of HMGB1 slowed the growth rate of xenograft, decreased the tumor volume and increased the apoptosis rate after irradiation. Conclusions:ESCC patients with high expression level of HMGB1 obtain poor prognosis after chemoradiotherapy, which can be enhanced by increasing the sensitivity to radiotherapy and chemotherapy. HMGB1 knockdown can effectively increase the radiosensitivity of xenograft in ESCC nude mice.

Research progress on radio-resistance mechanism of nasopharyngeal carcinoma / 中华放射肿瘤学杂志

Nasopharyngeal carcinoma is one of the most common malignant head and neck tumors, and radiotherapy is the main treatment. However, radio-resistance is a key cause of local recurrence of nasopharyngeal carcinoma. Therefore, overcoming the radio-resistance of nasopharyngeal carcinoma and enhancing the radiosensitivity have become urgent problems in the treatment of nasopharyngeal carcinoma, which also play a key role in improving the overall survival rate of patients. In this article, recent studies on DNA, non-coding RNA (ncRNA), protein and cell behaviors related to radio-resistance of nasopharyngeal carcinoma were reviewed, aiming to provide valuable ideas for clinical treatment of nasopharyngeal carcinoma.

Expression of NF45 in laryngeal squamous cell carcinoma and its effect on radiation sensitivity of tumor cells / 中华放射肿瘤学杂志

Objective:To investigate the expression of double-stranded RNA-binding protein nuclear factor 45 (NF45) in laryngeal squamous cell carcinoma (LSCC), and the effect of NF45 on the radiation sensitivity of LSCC cells and its mechanism.Methods:NF45 expression in LSCC and adjacent tissues was detected by real-time reverse transcription PCR (qRT-PCR) and immunohistochemical staining. The NF45-ShRNA lentivirus was transfected into Hep-2 cells, and cell transfection efficiency was determined by qRT-PCR and Western blot . Hep-2 cells were randomly divided into the control group, 2 Gy group, sh-NC+2 Gy group and sh-NF45+2 Gy group. Lentivirus infection and 2Gy X-ray irradiation treatment were carried out. Cell proliferation activity was assessed by CCK-8 assay. Apoptosis rate was determined by flow cytometry. Hep-2 cells in each group were treated with mCherry-EGFP-LC3B. The levels of autophagy were detected by immunofluorescence staining. The ratio of autophagy-related protein microtubule-associated protein 1 light chain 3 (LC3)-Ⅱ/LC3-Ⅰ and the expression levels of Beclin-1 and p62 proteins were determined by Western blot.Results:The expression level of NF45 in LSCC tissues was significantly higher than that in adjacent tissues ( P<0.01). The relative expression levels of NF45 mRNA and protein in Hep-2 cells infected with NF45-shRNA were significantly lower than those in the control and sh-NC groups (all P<0.05). Compared with the control group, the cell proliferation activity was decreased, the apoptosis rate was increased, the intracellular autophagy-lysosome were increased, the ratio of LC3-Ⅱ/LC3-Ⅰ was increased, the relative expression levels of Beclin-1 protein were up-regulated, and the relative expression levels of p62 protein were down-regulated in the 2 Gy, sh-NC+2 Gy and sh-NF45+2 Gy groups (all P<0.05). Compared with the 2 Gy group, the cell proliferation activity was decreased, the apoptosis rate was increased, the intracellular autophagy lysosomes were increased, the LC3-Ⅱ/LC3-Ⅰ ratio was increased, the relative expression of Beclin-1 protein was up-regulated, and the relative expression of p62 protein was down-regulated in the sh-NF45+2 Gy group (all P<0.05). Conclusions:The expression of NF45 is up-regulated in LSCC tissues. Targeted down-regulation of NF45 expression can inhibit the proliferation activity of LSCC cells, promote cell apoptosis, and improve the sensitivity of tumor cells to radiation. The mechanism may be related to the regulation of autophagy levels.

LncRNA TTN-AS1 affects the radiosensitivity of breast cancer cells by regulating miR-107/ HMGA1 signaling axis / 中华放射肿瘤学杂志

Objective:To investigate the effect and mechanism of long non-coding RNA (lncRNA) TTN-AS1 on the radiosensitivity of breast cancer cells. Methods:The expression of TTN-AS1 in breast cancer cells was detected by real-time reverse transcription PCR (qRT-PCR). MDA-MB-231 cells were divided into the 0 Gy group, 4 Gy group, negative control (NC) +4 Gy group, si- TTN-AS1+4 Gy group, si- TTN-AS1+ miR-107 inhibitor+4 Gy group, and si- TTN-AS1+ miR-107 inhibitor+si- HMGA1+4 Gy group. CCK-8 assay and flow cytometry were used to detect the proliferation and apoptosis rates in each group. Results:Compared with breast epithelial cells, TTN-AS1 was significantly highly expressed in breast cancer cell lines ( P<0.001). Compared with the NC+4 Gy group, the cell proliferation ability was significantly decreased ( P<0.05) and cell apoptosis was significantly increased ( P<0.001) in the si- TTN-AS1+4 Gy group. Compared with the 0 Gy group, the expression levels of TTN-AS1 and HMGA1 from 8 h to 24 h after radiotherapy were significantly up-regulated (both P<0.01), whereas the expression of miR-107 was significantly down-regulated from 8 h to 24 h after radiotherapy in the 4 Gy group ( P<0.001). The cell proliferation ability in the si- TTN-AS1+ miR-107 inhibitor+4 Gy group was significantly higher than that in the si- TTN-AS1+4 Gy group ( P<0.001), and cell apoptosis was significantly lower than that in the si- TTN-AS1+4 Gy group ( P<0.001). Compared with the si- TTN-AS1+ miR-107 inhibitor+4 Gy group, cell proliferation ability was significantly decreased ( P<0.001), whereas cell apoptosis was significantly increased in the si- TTN-AS1+ miR-107 inhibitor+si- HMGA1+4 Gy group ( P<0.001). Conclusion:TTN-AS1 can promote the radiosensitivity of breast cancer cells by regulating the miR-107/ HMGA1 signaling axis.

Study of the effect of PU-H71 on increasing radiosensitivity of cervical cancer cells / 中华放射肿瘤学杂志

Objective:To investigate the effect of heat shock protein 90 (Hsp90) inhibitor PU-H71 combined with X-ray on radioresistant human cervical cancer cells.Methods:The expression levels of Hsp90 gene between cervical cancer tissues and adjacent tissues were analyzed by bioinformatics. Radioresistant cervical cancer cell lines HeLa RR and SiHa RR were obtained by fractional irradiations (2 Gy per fraction, 30 fractions). The cell lines were divided into the control group (treated with dimethyl sulfoxide), irradiation alone group, PU-H71 group (treated with 0.5 μmol/L PU-H71), and PU-H71+irradiation group (irradiation at 24 h after treatment with 0.5 μmol/L PU-H71). Cell survival was detected by clonal formation assay. Immunofluorescence assay was used to detect γH2AX foci at 1, 6, and 24 h after cell treatment. The expression level of Rad51 protein at 1, 2, 6, 12, and 24 h after cell treatment was detected using Western blot. The expression level of phosphorylated DNA-dependent protein kinase catalytic subunit (p-DNA-PKcs) was measured at 2 h after cell treatment. Cell apoptosis at 48 h after cell treatment was assessed by flow cytometry. Results:PU-H71 enhanced the sensitivity of radioresistant cervical cancer cells to X-ray. Compared with the irradiation alone group, the radiation sensitization ratios (SER) of HeLa RR and SiHa RR cells at 10% survival were 1.36 and 1.27, and the apoptosis rates were increased by approximately 72.1% and 63.1% in the PU-H71+irradiation group, respectively. PU-H71 delayed the duration of γH2AX foci induced by X-ray, inhibited the phosphorylation of DNA-dependent protein kinase catalytic subunit (DNA-PKcs), thus preventing non-homologous end joining (NHEJ) repair and delaying homologous recombination repair.Conclusion:PU-H71 increases the radiosensitivity of radioresistant cervical cancer cells by inhibiting the repair pathway of DNA double-strand break, which is expected to be a radiosensitizer to enhance the efficacy of radiotherapy for cervical cancer.

Research progress on ferroptosis in radiosensitivity of malignant tumors / 中华放射肿瘤学杂志

As one of the most effective treatments for cancer, radiotherapy(RT)can eliminate the tumor cells and improve the survival rate of cancer patients. However, radiation resistance of tumor cells remarkably reduces the efficacy of RT. Therefore, it is significant to investigate the resistance mechanism and explore corresponding therapy for cancer. Ferroptosis is a novel mode of programmed cell death. Numerous studies have indicated an intimate connection between ferroptosis and tumor radiosensitivity. The mechanism involves lipid reactive oxygen species accumulation, glutathione metabolism and iron metabolism. It has been reported that ferroptosis inducers can act effectively and synergistically with RT, promote radiosensitivity and further improve tumor prognosis. In this article, relevant mechanisms and research progress were reviewed, and the future development direction of this field was discussed, aiming to provide reference for clinical trials and mechanism research of RT for malignant tumors.

Mechanism of the effect of arginine on radiosensitization of metastatic brain tumors / 中华放射肿瘤学杂志

Brain metastases are the most common intracranial malignancies, and radiotherapy is an effective treatment of controlling the localized lesions. However, conventional radiotherapy techniques have their own shortcomings that limit the effectiveness of radiotherapy. Metastatic brain tumors are highly likely to recur or progress after treatment. Clinical studies have shown that arginine can penetrate the blood-brain barrier and subsequently improve the radiosensitization of metastatic brain tumors. In this article, the mechanisms underlying the effect of arginine on the radiosensitization of metastatic brain tumors by inhibition of tumor glycolytic metabolism, reduction of DNA damage repair and alteration of tumor hemodynamic parameters were reviewed, aiming to provide new ideas for clinical research and treatment of brain metastases.

Effects of niraparib on the radiosensitivity of human esophageal cancer cells and its mechanism / 中华放射肿瘤学杂志

Objective:To evaluate the effect of niraparib, the poly (ADP-ribose) polymerase (PARP) inhibitor, on the radiosensitivity of esophageal squamous cell carcinoma (ESCC) and to preliminarily investigate its mechanism.Methods:Human esophageal squamous cell carcinoma cells ECA-109 and KYSE-150 were divided into the control, niraparib, single irradiation, combined (niraparib+irradiation) groups. Cell proliferation was measured by CCK-8 assay. The changes of cell survival rate were detected by colony formation assay. The changes of cell cycle and apoptosis were analyzed by flow cytometry. The number of γH2AX foci was detected by immunofluorescence, and the expression levels of PARP-1, cleaved-PARP, RAD51, mitogen-activated protein kinase (MAPK) [extracellular signal-regulated kinase 1 and 2 (ERK1/2) ] and p-MAPK (ERK1/2) proteins were determined by Western blot. All data were expressed as Mean±SD. Data between two groups conforming to normal distribution through the normality test were subject to independent sample t-test and multiple groups were analyzed using one-way ANOVA. Results:In human ESCC cells ECA-109 and KYSE-150, the proliferation of ESCC cells was significantly inhibited by niraparib combined with irradiation, and the values of average lethal dose (D 0), quasi-threshould dose(D q), survival fraction after 2 Gy irradiation (SF 2) in the combined group were decreased compared with those in the single irradiation group. The effect of irradiation alone on apoptosis of ECA-109 and KYSE-150 cells was limited. Compared to single irradiation group, irradiation combined with niraparib further increased the apoptosis rate in ESCC cells ( P=0.015, P=0.006). In ECA-109 cells, G 2/M phase arrest was significantly increased in combined group compared with irradiation alone group ( P<0.001). In ECA-109 cells, the number of γH2AX foci in combined group was higher than that in the single irradiation group after 2 h, and showed a significantly slower decay of γH2AX foci ( P<0.001). Moreover, niraparib combined with irradiation enhanced the radiation-induced cleavage of PARP-1 and down-regulated the expression of Rad51 and p-MAPK(ERK1/2). Conclusion:Niraparib can increase the radiosensitivity of esophageal cancer cells by inhibiting cell proliferation, promoting cell apoptosis, inhibiting the repair of DNA damage and regulating the MARK-ERK signaling pathway.

Radiosensitizing effect of PARP-1 inhibitor Olaparib on MCF-7 breast cancer and hypoxic imaging / 中国药理学通报

Aim To investigate the radiosensitizing effect of the PARP inhibitor Olaparib on MCF-7 breast cancer model and to monitor the radiosensitizing effect of Olaparib by F-Fluoroerythronitroimidazole (F-FETNIM) PET/CT. Methods MCF-7 breast cancer model was established and divided into control group, Olaparib group, irradiation group and Olaparib + irradiation group according to random number table method; tumor volume was measured to calculate tumor inhibition rate and survival time of tumor-bearing mice was counted.

Effect of Testin Gene on Radiosensitivity of Nasopharyngeal Cancer 5-8F Cells / 肿瘤防治研究

Objective To investigate the effect of TES gene on the radiosensitivity of nasopharyngeal carcinoma 5-8F cells. Methods Specimens of 5-8F cells (unprocessed group) and 5-8F cells with high TES expression (TES group) were irradiated at 0, 2, 4, 6, and 8 Gy radiation dose points.Cell clone formation experiment was conducted to draw the survival curve.Twenty-four BALB/c nude mice were randomly divided into four groups: nontransfection group, TES group, irradiation group, and TES irradiation group.A nude mouse model of nasopharyngeal carcinoma 5-8F cells was established.The length and diameter of the transplanted tumor were measured every three days, the tumor volume was calculated, and the growth curve of the transplanted tumor was drawn.After the mice were killed one month later, the tumor block was taken and weighed.The apoptosis of the transplanted tumor cells in each group was detected by flow cytometry. Results Compared with that in the unprocessed group, the survival rate of cells in the TES group was significantly lower (P < 0.01).The tumor growth rate and tumor mass of all four groups decreased in turn, while the apoptosis rate increased in turn.The TES irradiation group had the slowest tumor growth rate, greatest decrease in tumor weight, and highest apoptosis rate among the four groups.Multiple comparison revealed statistically significant differences between the groups (P < 0.05). Conclusion The testin gene can effectively improve the radiosensitivity of nasopharyngeal carcinoma 5-8F cells cultured in vitro.

Association between TOX gene expression level and radiosensitivity in lower-grade gliomas / 中华放射肿瘤学杂志

Objective:To investigate the relationship between the expression level of thymocyte selection-associated high mobility group box protein ( TOX) gene and the radiosensitivity of lower-grade glioma (LGG) patients. Methods:Using bioinformatics research methods, 474 LGG patients from The Cancer Genome Atlas (TCGA) database were selected as the test set (TCGA-474 set), and two different genetic data sets ( n=412 and n=171) from the Chinese Glioma Genome Atlas (CGGA) database were selected as the validation set (CGGA-412 set and CGGA-171 set). Patients were stratified based on whether received radiotherapy, and divided into the high and low TOX expression group according to the expression level of TOX gene in LGG. Survival curves of all patients were plotted. The overall survival (OS) and progression-free survival (PFS) of patients in the high and low TOX expression groups were compared and analyzed using log-rank test. Results:Multivariate analysis of OS in the TCGA-474 set showed that high expression of TOX was a protective factor for OS ( HR=0.061, 95% CI: 0.005-0.791, P=0.044). After stratification analysis based on radiotherapy and adjustment for confounding factors, the HR (95% CI) of patients with high TOX expression in the TCGA-474, CGGA-412, and CGGA-171 sets were 0.405 (0.261-0.629), 0.581 (0.418-0.806), and 0.464 (0.269-0.800), respectively, with P values of <0.001, 0.001, and 0.008, respectively. Among patients receiving radiotherapy in the TCGA-474 set, the OS and PFS of patients with high TOX expression were significantly longer than those in the low TOX expression group, and the differences were statistically significant (both P<0.001). The OS benefit of patients with high expression of TOX was significantly prolonged in both the CGGA-412 and CGGA-171 sets compared to those with low TOX expression, and the differences were statistically significant (both P<0.001). Conclusion:The high expression of TOX may be related to the radiosensitivity of LGG, which may be a gene marker of the radiosensitivity of LGG.

Heterogeneity of cancer-associated fibroblasts and regulating roles in cancer radiotherapy / 中华放射肿瘤学杂志

Cancer-associated fibroblasts (CAF) represent a crucial and abundant group of stromal cells in the tumor microenvironment (TME) and are effective targets for anti-tumor therapy. CAFs exhibit high heterogeneity and plasticity, which play a pivotal role in tumor initiation, progression, immune evasion, and therapy resistance. Radiotherapy (RT) is a fundamental treatment modality for malignant tumors that can reshape the TME through various mechanisms. Following RT, CAFs undergo a series of phenotypic changes and interact with multiple cells in the TME, promoting radioresistance and immune evasion through multiple pathways, such as enhancing tumor cell proliferation, modulating immune response, inducing angiogenesis, remodeling extracellular matrix, and reprogramming metabolism, etc., thereby affecting therapeutic effect. Targeting CAFs in combination with RT can improve anti-tumor efficacy and prognosis. In this article, research progress in CAFs in tumor RT was reviewed.