Circ-NUP98 Promotes Lung Adenocarcinoma Development Through Regulating CBX1 by miR-188-3p.

Lung cancer has a high morbidity and mortality among malignant tumors, and lung adenocarcinoma (LUAD) is the main type of lung cancer. In recent years, circular RNAs (circRNAs) have been confirmed to play an important role in the generation and development of human cancer. However, the specific role and mechanism of circ-NUP98 in LUAD are still unclear and need to be further investigated. Circ-NUP98, microRNA-188-3p (miR-188-3p), and chromobox homolog 1 (CBX1) levels were detected by real-time quantitative polymerase chain reaction (RT-qPCR). Cell-counting Kit-8 (CCK-8), 5-Ethynyl-2'-deoxyuridine (EdU) assay, flow cytometry, wound healing, and transwell assay were used to observe LUAD cell proliferation, apoptosis, migration, invasion, and cell-cycle progression. Malondialdehyde (MDA) and superoxide dismutase (SOD) levels were examined using special assay kits. CyclinD1, Bcl-2-related X protein (Bax), matrix metalloproteinase 9 (MMP9) protein, and CBX1 protein levels were determined using Western blot. The interaction between miR-188-3p and circ-NUP98 or CBX1 was identified by dual-luciferase reporter and RNA immunoprecipitation (RIP) assay. In vivo efficacy of circ-NUP98 was evaluated in a xenograft tumor model. Besides, the expression of CBX1 and KI67 in the tumors was detected by immunohistochemical (IHC) assay. Circ-NUP98 and CBX1 expressions were upregulated in LUAD tissues and cells, and miR-188-3p was decreased. Downregulation of circ-NUP98 could inhibit the proliferation, migration, invasion, and oxidative stress, and promote apoptosis of LUAD cells. Mechanism experiments showed that circ-NUP98 acted as a sponge for miR-188-3p to increase CBX1 expression. Knockdown of circ-NUP98 could inhibit the growth of LUAD tumors in vivo. Circ-NUP98 might promote the malignant development of LUAD via the miR-188-3p/CBX1 axis, which might provide a potential new marker for early diagnosis of LUAD.

OIP5-AS1 modulates epigenetic regulator HDAC7 to enhance non-small cell lung cancer metastasis via miR-140-5p.

Long non-coding RNAs have been reported to be involved in non-small cell lung cancer (NSCLC) progression. However, whether Opa-interacting protein 5 antisense RNA 1 (OIP5-AS1) serves a role in NSCLC remains unclear. Bioinformatics analysis of The Cancer Genome Atlas datasets showed clinical significance and relevance of OIP5-AS1 in NSCLC. Western blotting and reverse transcription-quantitative PCR revealed protein and RNA expression levels of the genes [including OIP5-AS1, microRNA (miR)-140-5p, histone deacetylase 7 (HDAC7) and vascular endothelial growth factor A (VEGFA)]. Direct associations between the genes (miR-140-5p and OIP5-AS1, or miR-140-5p and HDAC7) were confirmed using a dual-luciferase reporter assay. Lymphatic vessel formation and invasion ability were detected using a lymphatic vessel formation assay and Transwell invasion assay. OIP5-AS1 knockdown attenuated lymphatic vessel length and invasion. The role of OIP5-AS1 was reverted by miR-140-5p. HDAC7 and VEGFA are downstream effectors of miR-140-5p-mediated NSCLC metastasis. OIP5-AS1, miR-140-5p, HDAC7 and VEGFA were all dysregulated in human clinical NSCLC tumor tissues. In conclusion, the present results demonstrated a novel mechanism for OIP5-AS1-induced metastatic phenotypes of NSCLC via the miR-140-5p/HDAC7/VEGFA axis.

Hsa_circ_0079662 induces the resistance mechanism of the chemotherapy drug oxaliplatin through the TNF-α pathway in human colon cancer.

The aim of the study was to research the biological functions of circRNA (hsa_circ_0079662) and its underlying mechanism in colorectal cancer. Drug-resistant cell lines (HT29-LOHP, HCT116-LOHP, HCT8-LOHP) were separately dealt with oxaliplatin concentration gradient (0.1-10 µmol/L). Real-time PCR, Western blotting, dual-luciferase assay, miRNA pull-down assay, coimmunoprecipitation and ELASA were performed to explore the mechanism of chemotherapy drug oxaliplatin resistance in CRC. The results showed that the expression of hsa_circ_0079662 was increased in drug-resistant cell lines by RT-PCR. The expression of HOXA9, TRIP6, Vcam-1, VEGFC, MMP3, MMP9 and MMP14 was higher by Western blotting. Interaction between HOXA9 and TRIP6 in CO-IP detection. Additionally, the cytokines TNF-α, IL-1 and IL-6 were also found. In conclusion, hsa_circ_0079662, as a ceRNA binding with hsa-mir-324-5p, can regulate target gene HOXA9 and induced the mechanism of chemotherapy drug oxaliplatin resistance in CRC through the TNF-α pathway in human colon cancer.

Effect of VEGFR 2 gene polymorphism V297I on clinical outcomes of advanced NSCLC patients treated with bevacizumab combining with chemotherapy as first line treatment / 中国肿瘤生物治疗杂志

@# Objective: To investigate the effect of VEGFR2 gene polymorphism V297I on the clinical outcomes in patients with advanced non-small cell lung cancer (NSCLC) treated with bevacizumab combining with chemotherapy. Methods:Atotal of 135 patients with advanced NSCLC, who were treated by bevacizumab plus platinum-based chemotherapy for first-line regimen, were included in this study. PCR-RFLP assay was used to detect the VEGFR2 genotypes in peripheral blood of patients and qPCR was used to detect the VEGFR2 mRNA in the cancer tissues of NSCLC patients. Logistic regression analysis was used to analyze the correlation between gene polymorphism and other variants, Kaplan-Meier assay to analyze the correlation between genotype and prognosis, and Cox regression model to analyze the risk factors for patients’PFS. Results: Of the polymorphisms analyzed, only polymorphism V297I was found to be of clinical significance. V297I locates in the coding region of VEGFR2, and it’s prevalence in the study population was as follows: CC genotype in 99 cases (73.33%), CT genotype in 33 cases (24.44%) and TT genotype in 3 cases (2.23%); the frequency of minor allele was 0.14, and the distribution of three genotypes was in accordance with Hardy-Weinberg equilibrium (P>0.05). The overall objective remission rate (ORR) of the 135 patients was 45.93%, the median progression free survival (mPFS) was 8.2 months and the median overall survival (mOS) was 20.8 months. The ORR, mPFS and mOS of patients with CT/TT genotype and CC genotype were 41.67%, 6.2 months, 18.9 months and 47.47%, 8.9 months and 21.5 months, respectively (all P<0.05).Additionally, the mRNAexpression of VEGFR2 in cancer tissues of the patients with CT/TT genotype was significantly higher than those with CC genotype (P< 0.01). The risk factors for patients’PFS included V297I, gender and ECOG score. Conclusion:Among advanced NSCLC patients treated by bevacizumab plus platinum-based chemotherapy, the polymorphism V297I of VEGFR2 may impact the clinical outcomes and prognosis of NSCLC patients treated with bevacizumab first line treatment by influencing the mRNAexpression of VEGFR2.

Association between genetic variation of kinase insert domain receptor and prognosis in colorectal cancer patients received 5-FU based adjuvant chemotherapy / 中国肿瘤生物治疗杂志

@# Objective: To investigate the association between genetic variation of kinase insert domain receptor (KDR) and the prognosis in colorectal cancer (CRC) patients received 5-FU based adjuvant chemotherapy. Methods: The clinical data of 176 CRC patients, who underwent surgical treatment at the Department of Anus and Intestine Surgery, People’s Hospital of Zhengzhou during January 2012 and December 2017, were retrospectively analyzed, and 93 cases of tumor tissues were collected for this study. The genotype of KDR polymorphism locus was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). qPCR was used to detect the expression of KDR mRNAin colorectal cancer tissues. The correlation between the polymorphism genotypes and other variables was analyzed by logistic regression model. The expression of different genotypes of KDR was analyzed by nonparametric test. The relationship between KDR genotype and prognosis of patients was analyzed by Kaplan-Meier survival analysis, and the other variables were adjusted by Cox risk scale model. Results: Of the polymorphisms analyzed, only rs2071559 was of clinical significance. The distribution frequency of KDR rs2071559 in 176 CRC patients was as follows: TT genotype in 95 cases (53.98%), TC genotype in 70 cases (39.77%) and CC genotype in 11 cases (6.25%); the minor allele frequency was 0.26; and the distribution of three genotypes was in accordance with Hardy-Weinberg's Equilibrium (P=0.690). The median disease free survival (mDFS) of patients carrying C allele and wild type TT genotype was 4.4 and 3.2 years, respectively (P<0.05); The median overall survival (mOS) of patients with TC/CC genotype and TT genotype was 5.2 and 4.0 years, respectively (P<0.05). After COX model modification, the effect of TC/CC genotype on mOS was still statistically significant (OR=0.55, P<0.05). The mRNA expression of KDR in cancer tissues of the patients with TC/CC genotypes were significantly lower than those of the wild type TT genotype (P<0.01). Conclusion: The polymorphism of KDR rs2071559 is associated with clinical outcomes in patients with colorectal cancer. KDR rs2071559 may affect the prognosis of colorectal cancer patients by affecting the mRNAexpression of KDR.

Long Noncoding RNA PlncRNA-1 Promotes Colorectal Cancer Cell Progression by Regulating the PI3K/Akt Signaling Pathway.

Accumulating evidence has indicated that long noncoding RNA (lncRNA) PlncRNA-1 plays an important regulatory role in cancers. However, the expression and biological functions of PlncRNA-1 in colorectal cancer (CRC) are still unclear. In the present study, we determined the expression of PlncRNA-1 in CRC and explored the function of PlncRNA-1 on CRC cell progression. The results showed that PlncRNA-1 was significantly increased in CRC tissues and cell lines; high PlncRNA-1 expression was associated with depth of invasion, lymph node metastasis, and TNM stage of CRC patients. Kaplan-Meier curve analysis showed that patients with high PlncRNA-1 expression had a poor overall survival. PlncRNA-1 knockdown remarkably reduced cell proliferation, migration, and invasion and promoted cell apoptosis in vitro. In vivo xenograft experiments showed that PlncRNA-1 inhibition significantly suppressed tumor growth. Finally, we used an agonist (740Y-P) of the PI3K/Akt signaling pathway; function assays showed that PlncRNA-1 exerted its effects by targeting the PI3K/Akt signaling pathway in CRC. Taken together, our data suggested that PlncRNA-1 might act as an oncogene in CRC progression and serve as a potential biomarker and therapeutic target for the treatment of CRC.

High expression of MICA/B prolongs disease-free survival in patients with HER2positive breast cancer / 中国肿瘤生物治疗杂志

@#[Abstract] Objective: To investigate the relationship between expression of MICA/B (MHC class I chain-related proteinA/B) and disease-free survival (DFS) of patients with HER2+(human epidermal growth factor receptor 2) breast cancer tissue. Methods: Twenty six cases of corresponding para-cancerous tissue and 100 cases of HER2+ breast cancer tissue that preserved in wax at Zhengzhou People’ s Hospital Affiliated to Southern Medical University from January 2009 to June 2010 were collected for this study. Expression of MICA/ B in these tissue samples was detected by immunohistochemistry; and the relationship between MICA/B expression with clinicopathologic features as well as DFS was analyzed with Kaplan-Meier survival curve. Results: The expression of MICA/B in adjacent paracancerous tissues was negative (0/26), however, it was highly positive in cancer tissues (92/100), and the percentage with high expression was 65%(65/100), the difference was significant (P<0.05). High MICA/B expression rate in stage I was significantly higher than that in stage Ⅱ-Ⅲ (77.55% vs 52.94%， P<0.05), and the high expression rate in stage T1 was also significantly higher than that in stage T2-T4 (75.00% vs 52.27%， P<0.05). High MICA/B expression rate in ER+, PR+ group (with positive number≥1%) was significantly lower than that in ER- , PR-group (ER： 52.38% vs 74.14%，PR： 51.35% vs 73.02%， all P<0.05). MICA/B expression was correlated with clinical stages, the expression of ER, PR and tumor size (all P<0.05), but not associated with menopausal status, histological grade and lymph node metastasis (all P>0.05). Over-expression of MICA/B was closely associated with much better 6-year DFS rate in patients no matter with or without targeted therapy (the targeted group: 90.6% vs 72.2%; the untargeted group: 78.4% vs 58.8%, all P<0.05). Conclusion: Over-expression of MICA/B in HER2+ breast cancer tissue is closely related to DFS, which may be served as a potential prognosis indicator for patients with HER2+ breast cancer.

Primary mucinous adenocarcinoma of the lung: A case report and review of the literature.

Mucinous adenocarcinoma is an unusual histological type of lung cancer, and its clinicopathological feature is distinctive from that of other histopathological types of lung adenocarcinoma. Mucinous adenocarcinoma has a mucus-producing function, which explains its name. The present study reports a rare case of a mucus-producing adenocarcinoma of the lung. A 60-year-old Chinese female patient was diagnosed with mucinous adenocarcinoma of the lung, which manifested as respiratory symptoms, including fever, cough and expectoration. The patient received thoracic exploratory operation and right pneumonectomy, since the above respiratory symptoms seriously affected her daily life, and other inspections could not establish the diagnosis. Histopathology revealed no mutations in epidermal growth factor receptor. The patient received adjuvant chemotherapy using taxol and cisplatin. However, metastases in the left lung were detected 7 months after the operation. Pemetrexed and cisplatin were used as the second-line treatment. The patient survived 3 years after the initial diagnosis. The present study reports a rare mucus-producing adenocarcinoma of the lung, which is of bad prognosis. Therefore, further studies on this type of cancer are urgently required.

Effect of Fructus Corni extract on B7-H6 expression in primary liver cancer of rats / 中国肿瘤临床

Objective:To investigate the effects of Fructus Corni extract on the B7-H6 expression in primary liver cancer cells of rats. Methods:Sixty SD rats were randomly divided into three groups, namely, model, matrine, and Cornus officinalis. The rat model bear-ing the primary liver cancer was induced by diethylnitrosamine, except for the rats in the control group. The rats in both the matrine and Cornus officinalis groups were fed with matrine and Cornus officinalis. The rats in model groups were fed with 0.9％sodium chlo-ride solution. The number of hepatocellular carcinoma nodules was calculated, and the tumor growth inhibition rate was also calculat-ed. The pathological changes of hepatic tissues in rats of each group were observed by hematoxylin and eosin staining. The expression levels of B7-H6 in these three groups were determined by immunohistochemistry and Western blot. Results:The number of liver nod-ules of the matrine and Fructus Corni group rats was lower than that of the model group (P<0.05). The tumor inhibition rate of the Cor-nus group was significantly higher than that of the matrine group (P<0.05). The tumor growth inhibition rate of the Cornus officinalis group was significantly higher than that of the matrine group (P<0.05). Immunohistochemistry showed that the positive expression of B7-H6 in the Cornus officinalis group and the matrine group was significantly higher than that in the model group (P<0.05), and the positive expression of B7-H6 in the Cornus officinalis group was significantly higher than that in the matrine group (P<0.05). Similarly, the protein expression of B7-H6 in the Cornus officinalis and matrine groups was significantly higher than that in the model group (P<0.05) by Western blot, while the protein expression of B7-H6 in the Cornus officinalis group was significantly higher than that in the matrine group (P<0.05). Conclusion:Fructus Corni extract may inhibit the growth of hepatocellular carcinoma through upregulating the B7-H6 expression.

Development of synthetic of peptide-functionalized liposome for enhanced targeted ovarian carcinoma therapy.

In this study, we report an active targeting liposomal formulation directed by a novel peptide (T7) that specifically binds to the transferrin receptor (TfR) overexpressed on ovarian carcinoma cells. The objectives of this study were to evaluate the in vitro and in vivo tumor drug targeting delivery of T7-anchored liposomes on A2780 cells. T7 conjugated to the distal end of DSPE-PEG2000-maleimide was incorporated into the liposomes via a post-insertion method, the liposome could keep stability in 50% FBS for more than 24 h. The uptake efficiency of T7-LP was 3.7 times higher than that of LP on A2780 cells. The anti-proliferative activity of T7-LP-PTX against A2780 cells was much stronger compared to that of LP-PTX and free PTX, respectively. The homing specificity and anticancer efficacy of T7-LP-PTX were also evaluated on the tumor spheroids, which revealed that T7-LP-PTX was more efficaciously internalized into tumor cells than LP. Compared to LP, T7-LP-PTX showed the highest accumulation capability into tumor spheroids, and the greatest tumor growth inhibitory effect in vitro. In the in vivo study, the T7-LP-PTX showed the best inhibition effect of the tumor growth for the A2780-bearing mice and tumor accumulation. In brief, the T7-LP may be an efficient targeting drug delivery system for ovarian carcinoma.

Down-regulation of long non-coding RNA GAS5 is associated with the prognosis of hepatocellular carcinoma.

INTRODUCTION: Long non-coding RNAs (lncRNAs) have been investigated as a new class of regulators of cellular processes, such as cell growth, apoptosis, and carcinogenesis. lncRNA GAS5 has recently been identified to be involved in tumorigenesis of several cancers such as breast cancer, lung cancer and renal cancer. However, the regulation of GAS5 in hepatocellular carcinoma (HCC) has not yet been reported before. METHODS: Expression of GAS5 in tumor and their normal matched tissues was determined by quantitative real-time PCR in n = 71 HCC patients, and its association with overall survival of patients was analyzed by statistical analysis. RESULTS: The expression level of GAS5 was reduced in HCC in comparison to normal matched tissues (P < 0.05). It is also proved that GAS5 expression was to be associated with HCC tumor size, lymphnode metastasis and clinical stage (P < 0.05). In addition, the Kaplan-Meier survival curves revealed that low GAS5 expression was associated with poor prognosis in HCC patients. GAS5 expression was an independent prognostic marker of overall HCC patient survival in a multivariate analysis. CONCLUSIONS: The study proved for the first time that GAS5 down regulated in a majority of HCC patients. Our results indicated that GAS5 expression was an independent prognostic factor for patients with liver cancer, which might be a potential valuable biomarker for HCC.

Development of synthetic of peptide-functionalized liposome for enhanced targeted ovarian carcinoma therapy.

In this study, we report an active targeting liposomal formulation directed by a novel peptide (T7) that specifically binds to the transferrin receptor (TfR) overexpressed on ovarian carcinoma cells. The objectives of this study were to evaluate the in vitro and in vivo tumor drug targeting delivery of T7-anchored liposomes on A2780 cells. T7 conjugated to the distal end of DSPE-PEG2000-maleimide was incorporated into the liposomes via a post-insertion method, the liposome could keep stability in 50% FBS for more than 24 h. The uptake efficiency of T7-LP was 3.7 times higher than that of LP on A2780 cells. The anti-proliferative activity of T7-LP-PTX against A2780 cells was much stronger compared to that of LP-PTX and free PTX, respectively. The homing specificity and anticancer efficacy of T7-LP-PTX were also evaluated on the tumor spheroids, which revealed that T7-LP-PTX was more efficaciously internalized into tumor cells than LP. Compared to LP, T7-LP-PTX showed the highest accumulation capability into tumor spheroids, and the greatest tumor growth inhibitory effect in vitro. In the in vivo study, the T7-LP-PTX showed the best inhibition effect of the tumor growth for the A2780-bearing mice and tumor accumulation. In brief, the T7-LP may be an efficient targeting drug delivery system for ovarian carcinoma.

Digestive tract hemorrhage due to complications with gastrointestinal stromal tumor treated with sunitinib: A case report.

Gastrointestinal stromal tumors (GISTs) are rare, and account for 1% of all gastrointestinal neoplasms. GISTs are the most frequent mesenchymal tumors of the gastrointestinal tract. However, the clinical and pathological characteristics of these neoplasms are not adequately understood. The best treatment approach for GISTs remains unclear. In the present study, we report a case of a GIST originating from the stomach. A digestive tract hemorrhage occurred as a complication of sunitinib treatment. This is the first report of a digestive tract hemorrhage due to sunitinib treatment.

[Expression of MICA/B protein in esophageal cancer and its clinical significance].

OBJECTIVE: To explore the expression of MICA/B in human esophageal cancer, and to analyze its correlation with clinicopathological features. METHODS: The expression of MICA/B in 40 cases of esophagus carcinoma and corresponding normal esophageal mucosa tissues were examined by immunohistochemistry. RESULTS: The positive rate of expression of MICA/B protein in the esophageal carcinoma was 75.0% (30/40), and that in the corresponding normal esophageal mucosa was 0 (0/40). Up-regulation of MICA/B expression was found in the esophageal carcinomas. The expression of MICA/B was related with histological grade of the esophageal carcinoma (P = 0.012). CONCLUSION: MICA/B protein plays an important role in the esophageal carcinogenesis, and my become a useful molecular marker for the diagnosis of esophageal carcinoma.

[Kaempferol-induced apoptosis of human esophageal squamous carcinoma Eca-109 cells and the mechanism].

OBJECTIVE: To evaluate the growth-inhibiting and pro-apoptotic effect of kaempferol in human esophageal squamous carcinoma Eca-109 cells and explore the mechanism. METHODS: The effect of kaempferol on Eca-109 cell proliferation in vitro was measured by MTT assay. TUNEL staining was used to detect the cell apoptosis following kaempferol treatment. The changes in Bax and Bcl-2 mRNA expressions in response to kaempferol treatment were determined by RT-PCR, and the caspase-3 and caspase-9 activities were evaluated using colorimetric assay. RESULTS: Kaempferol significantly inhibited Eca-109 cell proliferation (P<0.05) in a concentration-dependent manner and induced obvious cell apoptosis. RT-PCR showed that after kaempferol treatment caused up-regulated Bax and down-regulated Bcl-2 mRNA expression. The colorimetric assay revealed significantly increased caspase-3 and caspase-9 activities in Eca-109 cells following kaempferol treatment (P<0.01). CONCLUSION: Kaempferol can induce apoptosis of Eca-109 cells via a mitochondria-dependent pathway.

[Expression of NKG2D ligands on dendritic cells at different development stages and its effect on cytotoxicity of NK cells].

AIM: To investigate the expression of NKG2D ligands on dendritic cells(DC) at different development stages and its effect on cytotoxicity of natural killer(NK) cells. METHODS: The monocytes were cultured into immature dendritic cells(iDC) and mature dendritic cells(mDC) with cytokines. NK cells were obtained from normal peripheral blood by CD56 antibody magnetic isolation.The expression of NKG2D ligands (MICA/B, ULBP1-3) was detected by flow cytometry. Cytotoxicity of NK cells and the NK cells blocked with anti-NKG2D mAbs against iDC and mDC was tested using LDH-releasing method. RESULTS: IDC and mDC were of typical morphology and phenotypes. MICA, MICB, ULBP1, and ULBP3 were expressed on iDC and their expression rate was (32.39+/-8.30)%, (17.75+/-3.40)%, (26.71+/-6.48)%, (38.37+/-6.89)%, respectively. MICA and ULBP3 were expressed on mDC and their expression rate was (7.81+/-3.33)% and (8.36+/-2.42)%, respectively, which was lower than that on mDC (P<0.01). At the each E:T ratio cytotoxicity of NK cells against iDC was stronger than that against mDC (P<0.01). cytotoxicity of NK cells blocked with anti-NKG2D mAb against iDC was decreased compared with that of NK cells unblocked (P<0.05) while cytotoxicity of NK cells blocked with anti-NKG2D mAb against mDC showed no decrease compared with that of NK cells unblocked (P>0.05). CONCLUSION: The expression of NKG2D ligands on iDC is higher than that on mDC, which plays an important role in the cytotoxic effect of NK cells against iDC, but has no effect on that against mDC. NKG2D-NKG2D ligands shows one of the molecular mechanisms that NK cells kill iDC selectively.

[Effect of hyperthermia in combination with chemotherapy on K562/AO2 cells in vitro].

This study was purposed to investigate the inhibitory effect, apoptosis, Bcl-2 and P-gp expression of K562/AO2 cells by hyperthermia combined with adriamycin. The working concentration of adriamycin against K562/AO2 was determined by MTT assay. The hyperthermia and chemotherapy were used alone or in combination, then the cell survival rate was detected at 48 hours. The inhibitory effect was evaluated by MTT assay. The apoptosis rate, Bcl-2 and P-gp expression of K562/AO2 were determined by flow cytometry. The concentration of adriamycin in the experiment was defined as its IC(50) at 48 hours action. The results indicated that the hyperthermia at 40, 41 and 42 degrees C for 60 minutes showed obvious inhibitory effect on K562/AO2 cells (p < 0.01). Adriamycin chemotherapy combined with hyperthermia showed more obvious inhibitory effect on K562/AO2. According to flow cytometric analysis, the hyperthermia and adriamycin used alone or in combination could obviously increase the apoptosis rate and down-regulate Bcl-2 and P-gp expression of K562/AO2 cells (p < 0.01). It is concluded that the adriamycin chemotherapy combined with hyperthermia for 60 minutes shows obvious inhibitory effect on K562/AO2 cells, which increases the apoptosis rate and down-regulates expression of Bcl-2 and P-gp.

[Expression of NKG2D ligands in multidrug-resistant nasopharyngeal carcinoma cell line CNE2/DDP and their effects on cytotoxicity of natural killer cells].

OBJECTIVE: To analyze the expression of NKG2D ligands on human nasopharyngeal carcinoma cell line CNE2 and the multidrug-resistant lin CNE2/DDP and investigate its impact on cytotoxicity of natural killer (NK) cells. METHODS: Expression of NKG2D ligands on the surface of CNE2 and CNE2/DDP cells was analyzed by flow cytometry, and their HLA genotypes, along with inhibitory killer cell immunoglobulin-like receptors (KIRs) expressed on NK cells from 5 healthy donors, were determined by PCR with sequence specific primers. Cytotoxicity of NK cells against CNE2 and CNE2/DDP cells was evaluated by LDH-releasing assay at different effector-to-target ratios (E:T). In blocking experiments, different monoclonal antibodies (mAb) were added to the target cells at the E:T of 20:1 ratio. RESULTS: The HLA genotypes of CNE2 and CNE2/DDP cells were A2, 24, B18, 35, Cw4, 7, and the inhibitory KIR genotypes of the 5 healthy donors were KIR2DL1, KIR2DL3, KIR3DL1, and KIR3DL2, mismatched with the HLA -class I molecules expressed by the CNE2 and CNE2/DDP cells. The expression of MHC class I chain-related proteins A and B (MICA and MICB) on CNE2 cells was higher than that on CNE2/DDP cells (P<0.01), and ULBP1 and ULBP3 were not detectable. NK cells displayed highly in vitro cytotoxicity against CNE2 and CNE2/DDP cells with a mean cell lysis rate of (10.50-/+2.17)%, (4.98-/+0.95)%; (27.68-/+1.47) %, (15.48-/+2.10) %; (36.99-/+3.13) %, (28.46-/+4.30) %; (55.00-/+2.20) %, (40.95-/+2.21) %, respectively, corresponding to the E:T ratios of 5:1, 10:1, 20:1, and 30:1 (P<0.01). Blocking experiments confirmed that killing of CNE2 and CNE2/DDP cells by NK cells was efficiently inhibited by anti-MICA, anti-MICB, and anti-ULBP2 mAbs, whereas anti-ULBP1 and anti-ULBP3 mAbs had no effects on the cytotoxicity of the NK cells. CONCLUSION: Expression of NKG2D ligands on CNE2 and CNE2/DDP cells is correlated with NK cell-mediated lysis, and NK cells display higher cytotoxicity against parental CNE2 cells than the multidrug-resistant CNE2/DDP cells.

[Thermotherapy enhances the sensitivity of lymphoma cell line Raji to chemotherapy in vitro].

OBJECTIVE: To observe the effect of thermotherapy on the intracellular adriamycin concentration and apoptosis of Raji cells in vitro. METHODS: The working concentration of adriamycin against Raji cells was determined with MTT assay. Raji cells were subjected to thermotherapy (at 40 degrees Celsius;, 41 degrees Celsius; or 42 degrees Celsius;) and chemotherapy with adriamycin alone or in conjunction, and the cell survival rates were determined 48 h after the treatment. The cell growth inhibition effect of the treatment was evaluated with MTT assay, and the apoptotic rates of Raji cells and alteration of intracellular adriamycin concentration were determined by flow cytometry. RESULTS: The IC(50) of adriamycin was defined as the working concentration in the experiment. Thermotherapy at 40, 41 and 42 degrees Celsius; for 60 min in conjuction with chemotherapy significantly inhibited the growth of Raji cells (P<0.01). The results of flow cytometry showed that thermotherapy and adriamycin chemotherapy, used either alone or in combination, significantly increased the apoptotic rates of Raji cells (P<0.01), and thermotherapy remarkably increased the intracellular concentration of adriamycin. CONCLUSION: Adriamycin chemotherapy combined thermotherapy for 60 min can increase the intracellular concentration of adriamycin and the apoptosis rates of Raji cells.

[Expression of HLA class I molecules and MHC class I chain-related molecules A/B in K562 and K562/AO2 cell lines and their effects on cytotoxicity of NK cells].

The study was aimed to investigate the expression of HLA class I molecules and MHC class I chain-related molecules A/B (MICA/MICB) in K562 and adriamycin (ADM)-resistant K562 cell lines (K562/AO2) and their effect on cytotoxicity of NK cells. Expression of HLA class I molecules and MICA/MICB on the surface of K562 and K562/AO2 cell lines were analyzed by flow cytometry. Cytotoxicity of NK cells (isolated from 3 healthy persons) against K562 and K562/AO2 cells were detected by LDH releasing assay at different effect-to-target cell ratios (E:T). In blocking experiments, anti-MHC class I monoclonal antibody (McAb) (W6/32, a pan anti-HLA class I antibody) and anti-MHC class I chain-related molecules McAb (BAMO-1, specifically against MICA and MICB) were added to the target cells at E:T of 10:1. The results showed that the expression of MHC class I chain-related molecules on K562 was higher than that on K562/AO2 (P=0.000), and HLA class I molecules were not detectable on both cells. Cytotoxicities of NK cells against K562 and K562/AO2 cells were (29.32 +/- 0.12)%, (45.33 +/- 0.78)%, (58.37 +/- 0.87)%, (72.37 +/- 0.96)% and (12.47 +/- 0.91)%, (24.36 +/- 1.11)%, (33.29 +/- 1.03)%, (53.87 +/- 1.27)% at E:T ratios of 5:1, 10:1, 20:1 and 30:1 respectively (P=0.000), the cytotoxicity of NK cells on K562 cells was significantly higher than that on K562/A02 cells at different E:T ratios. Blocking experiments confirmed that at E:T of 10:1 killing of NK cells against K562 and K562/AO2 cells was efficiently inhibited by BAMO-1, whereas W6/32 had no effect on K562 and K562/AO2 cells. It is concluded that the expression of MHC class I chain-related molecules on K562 and K562/AO2 cells is correlated with NK cell-mediated lysis. NK cells display higher cytotoxicity against parental K562 cells than multi-drug resistant K562/AO2 cells. Down-regulation of MICA/B in multi-drug resistant tumor cell lines leads to reduction of susceptibility to NK lysis.