The aim of this study was to evaluate the effects of recombinant human adenovirus p53 (rAd-p53; Gendicine) transfection and radiation at various time points following transfection. Cytotoxic effects and p53 protein expression levels were analyzed. rAd-p53 containing the human wild-type p53 gene was introduced into the human lung adenocarcinoma cell line A549, and cells were irradiated with a single dose of 6 MeV 4 Gy β rays. According to the time interval between rAd-p53 transfection and radiotherapy (RT), A549-transfected rAd-p53 cells were divided into 5 groups: radiation administered immediately after transfection (0 h-RT) group, after 3 h group (3 h-RT), after 6 h group (6 h-RT), after 24 h group (24 h-RT) and after 48 h group (48 h-RT). Cells with rAd-p53 transfection alone (Ad-p53) and with empty adenovirus (Ad) were included as the two control groups. Following 72 h of transfection, cell viability and growth were analyzed using MTT assays and flow cytometry, and p53 protein expression was analyzed using western blot analysis. From 0 h-RT to 48 h-RT, cell viability gradually decreased, while percentage of apoptotic cells and p53 protein expression gradually increased. The cell viability suppression rates in the 6 h-RT, 24 h-RT and 48 h-RT groups were 56.7±5.4, 60.8±6.0 and 68.9±6.6, respectively, which were significantly greater compared to that of the Ad-p53 (40.8±4.7), 0 h-RT (45.0±3.5) and 3 h-RT groups (47.0±4.3). No statistically significant differences were observed in the cell viability suppression rates among the 6 h-RT, 24 h-RT and 48 h-RT groups (P>0.05). Similar changes were observed in the percentage of apoptotic cells. The p53 protein expression level in the 6 h-RT group (0.856±0.092) was higher compared to that in the 3 h-RT group (0.643±0.089) (t=2.882; P=0.045), but not significantly different from that of the 24 h-RT group (1.193±0.202). The cell viability suppression rate and percentage of apoptotic cells was positively correlated with p53 protein expression in the A549 cells (P<0.05). Radiation may inhibit or damage p53 protein expression at the early stage of rAd-p53 transfection. To sensitize tumor cells to irradiation and achieve maximal cytotoxic effects, it is recommended to conduct RT at least 6 h following transfection with rAd-p53.
adenovirus p53; radiation therapy; gene therapy
Lung cancer is one of the leading causes of cancer-related mortality worldwide. Gastrointestinal metastasis from primary lung cancer is rare. Only a few reports have been published and the majority of described metastatic sites involved the small intestine. In the present study, we report the first case of primary lung adenocarcinoma with both gastric and colonic metastases. We also review the published literature of primary lung cancer with gastrointestinal metastasis.
lung cancer; gastrointestinal metastasis
CHOP (cyclophosphamide, doxorubicin, vincristine and prednisone)-like chemotherapy, in combination with rituximab (R-CHOP-like), improves outcome in patients with diffuse large B-cell lymphoma (DLBCL). We aimed to investigate the impact of rituximab on central nervous system (CNS) disease in adult patients. We studied 315 patients (aged 18–60 years old) from six hospitals between July 2003 and May 2008. All patients received CHOP-like (n=165) or R-CHOP-like (n=150) regimen every 3 weeks. With a median follow-up of 3.69 years, 10 patients (3.17%) developed CNS disease. The cumulative risk of CNS occurrence was not significantly different between the two treatment groups (P=0.871). We conclude that the addition of rituximab did not reduce the risk of CNS disease in adult patients with DLBCL.
central nervous system; diffuse large B-cell lymphoma; rituximab
Due to the favorable natural history in patients with low-grade gliomas (LGGs), there is no consensus on the treatment strategy following maximal safe surgical resection. A number of studies have been conducted to identify prognostic factors in patients treated for LGG. The present study evaluated the treatment outcomes as well as prognostic factors and their impact on overall survival (OS) and disease-free survival (DFS). We retrospectively reviewed 30 consecutive patients treated for LGG at the Department of Radiotherapy from February 2008 to July 2011. The patients underwent surgical intervention and postoperative radiotherapy. The response to radiotherapy was evaluated from six to eight weeks after the end of treatment using MRI analysis. Kaplan-Maier analysis was used for OS and DFS estimation. The endpoint was mortality as a result of any cause. Within a median follow-up of 21.8 months, 9 patients (30%) with disease progression were reported. The two- and five-year DFS and OS was 85.2 and 68.3% for DFS, and 84.3 and 63.4% for OS, respectively. The response to radiotherapy, evaluated in an MRI study, was found to be highly correlated with OS (p<0.0001). We also observed a significantly higher OS in patients with disease progression treated with salvage chemotherapy after the end of radiotherapy (p=0.08). Improved outcome among patients with LGG may be predicted by response to radiotherapy evaluated by MRI following termination of treatment.
low-grade gliomas; radiotherapy; prognostic factors
The transcription factor growth arrest and DNA damage-inducible gene 153 (GADD153), also known as CHOP, is considered to function as a proapoptotic molecule. Overexpression of GADD153 leads to cell cycle arrest and/or apoptosis. However, its clinical implications in non-small cell lung cancer (NSCLC) remain controversial. Therefore, we investigated the expression of GADD153 in stage I NSCLC using immunohistochemistry. Paraffin-embedded tissue sections from 76 patients, who were diagnosed with primary stage I NSCLC and had undergone a curative lung resection, were stained using an anti-GADD153 antibody. The intensity of GADD153 immunostaining was evaluated within the cell membrane and cytoplasm of invasive cancer components. The correlation between the intratumoral expression of GADD153 and various clinical parameters were explored. GADD153 was detected in 29 (38.2%) cases. No statistically significant difference in expression was demonstrated between stage IA and stage IB tumors (35.0 vs. 39.3%; P=0.735). The expression of GADD153 was not affected by histological subtypes or histological grades of differentiation. The intratumoral expression of GADD153 did not influence the overall survival rate (53.29 vs. 52.18 months; P=0.743) or disease-free survival rate (46.97 vs. 54.19 months; P=0.084) of stage I NSCLC patients. However, patients with GADD153 expression demonstrated an improved disease-specific survival rate (28.80 vs. 53.85 months; P=0.020). No patients with GADD153 expression demonstrated distant metastasis (P=0.029). These data suggest that GADD153 expression may be a valuable prognostic factor of early-stage NSCLC in patients who have undergone curative lung resection.
non-small cell lung cancer; growth arrest and DNA damage-inducible gene; immunohistochemistry; prognosis
In the present study, we reviewed 44 postgastrectomy adenocarcinoma patients who had hepatitis B and received treatment in the Abdominal Cancer Department of the West China Hospital between October 2006 and October 2010. Of these patients, 17 developed hepatic dysfunction. Radiotherapy is an independent risk factor to hepatic function on univariate and multivariate analysis. Grade III or IV hepatic dysfunction was developed by five patients, all of whom had received radiotherapy and had reactivated hepatic B virus (HBV). Radiotherapy is a significant risk factor to hepatic function in patients with postgastrectomy adenocarcinoma carrying HBV, thus we suggest excluding the liver from the radiation field. HBV reactivation plays a role in the development of grade III or IV hepatic dysfunction. Patients with reactivated HBV should immediately receive regular antiviral treatment.
hepatic dysfunction; radiotherapy; hepatitis B virus; gastric cancer; hepatitis B virus reactivation
Although the expression of keratin 7 (K7) and K20 is considered to be a useful factor in the differential diagnosis of intrahepatic cholangiocarcinoma (ICC) and metastatic colorectal carcinoma (CRC) of the liver, a proportion of typical ICC retains K20 expression. The frequency and biological significance of K20 expression in ICC remains unclear. We analyzed the expression of K7, K19 and K20 in 66 surgically resected liver tumors consisting of 46 ICCs and 20 metastatic CRCs of the liver and 20 corresponding primary CRCs. In the 46 ICCs, K7, K19 and K20 were expressed in 40 (87%), 45 (98%) and 16 (35%) cases, respectively. K7, K19 and K20 were expressed in 1 (5%), 20 (100%) and 16 (80%) of the 20 primary CRCs and 2 (10%), 20 (100%) and 16 (80%) of the 20 metastatic CRCs, respectively. A combined K7/K20 profile was identified as a good predictor for differentiating ICC and metastatic CRC. K20 expression in ICC was significantly associated with male gender (P=0.034), hilar location (P=0.026), intraductal papillary type (P=0.006), intestinal phenotype (P<0.001) and MUC2 expression (P=0.008). Univariate analysis identified that poor patient survival was significantly associated with histological grade (P=0.020), invasion depth (P=0.005), lymph node metastasis (P=0.012), tumor stage (P=0.004) and vessel invasion (P=0.023). The tumor stage (P=0.002) was a poor independent prognostic indicator, while MUC6 expression (P=0.036) was a good independent prognostic indicator. The survival rate in patients with K20-positive ICC was lower compared to that of patients with K20-negative ICC, but was not statistically significant. Furthermore, the combined K7/K20 immunophenotype was identified to be useful for differentiating ICC and metastatic CRC. K20-positive ICC displays specific characteristics with regards to tumor location and histological subtype. Additionally, MUC6 expression in ICC is a good independent prognostic factor, while K20 expression is more often associated with aggressive biological behavior.
keratin; cholangiocarcinoma; mucin; colorectal carcinoma
In our previous study, we reported that the cannabinoid receptors CB1 and CB2 are overexpressed in human hepatocellular carcinoma (HCC) tissues. Recently, the antitumor potential of the endogenous cannabinoid anandamide (AEA) has also been addressed. The present study was conducted to investigate the anti-proliferative effects of AEA in HCC cells. The human HCC cell line Huh7 was used. Cell proliferation was measured by MTT assay and flow cytometry. Apoptotic analysis was investigated by TUNEL assay. Real-time PCR and western blot analysis were used to analyze the expression of relevant molecules. The results of this study demonstrated that AEA inhibited the proliferation of Huh7 cells, resulted in G1 cell cycle arrest and induced apoptosis. Furthermore, downregulation of CDK4 and upregulation of p21 and Bak by AEA were observed. This study defines the anti-proliferative effects of anandamide in HCC cells and suggests that AEA has therapeutic potential in the management of HCC patients.
hepatocellular carcinoma; anandamide; proliferation
O6-methylguanine-DNA methyltransferase (MGMT) gene promoter hypermethylation is observed in a number of solid tumors and is correlated with the silencing of MGMT expression. In glioblastoma patients treated with the alkylating agent temozolomide, MGMT gene methylation status was shown to have predictive value in terms of prolonged overall survival. Recently, temozolomide has demonstrated promising activity in the treatment of soft tissue sarcomas, including those of the uterus. The tissue specimens involving tumor samples and normal uterine fragments were obtained from nine patients with smooth muscle uterine sarcoma, 11 with stromal uterine sarcoma and 17 with mixed uterine tumors. MGMT gene promoter methylation was analyzed by combined bisulfite restriction analysis (COBRA) while its expression levels were assessed using the real-time reverse transcription polymerase chain reaction (qRT-PCR). MGMT promoter methylation was observed in 27% of all tumor samples analyzed. When stratified by the disease type, 55.5% (5/9) of smooth muscle sarcomas, 23.5% (4/17) of mixed uterine tumor tissues and 9% (1/11) of stromal sarcomas showed MGMT methylation. The MGMT promoter methylation was associated with lower levels of gene expression in tumors when compared with those with an unmethylated promoter (P=0.0232) or normal tissues (P=0.0141). To conclude, MGMT promoter methylation and downregulation of gene expression is observed in a fraction of carcinosarcomas and non-epithelial malignant tumors of corpus uteri. The assessment of MGMT promoter methylation status may potentially identify patients who would benefit from temozolomide treatment.
uterine sarcoma; carcinosarcoma; O6-methylguanine-DNA methyltransferase; gene expression; gene methylation
In this study we report the pharmacokinetics and severe adverse effects of sunitinib in a woman with a gastrointestinal stromal tumor (GIST). A 60-year-old woman with small intestinal GIST developed severe thrombocytopenia (1.7×104/μl) following 1 week of treatment with sunitinib at 50 mg/day. Although the dose of sunitinib was reduced to 25 mg/day, platelet levels remained low. On day 7, the trough concentration of sunitinib plus SU12662 was 46.1 ng/ml and the area under the curve (AUC) was 1,393.0 ng·h/l. The dose was again reduced to 12.5 mg/day. However, the day after resumption of treatment, the patient developed symptoms of left heart failure due to myocardosis caused by sunitinib. Sunitinib has been reported to inhibit platelet-derived growth factor receptor (PDGFR) phosphorylation at concentrations over the range of 50–100 ng/ml (sunitinib plus SU12662) in vivo. In this case, the plasma concentration was sufficient to inhibit PDGFR at 25 or 50 mg/day. However, thrombocytopenia appeared at both dosages. Although the results in this case did not suggest a correlation between thrombocytopenia and plasma concentration, the degree of thrombocytopenia was decreased by reduction of the dose. In conclusion, the findings reported here indicate that the plasma concentration of sunitinib plus SU12662 is an important indicator to reduce adverse effects.
sunitinib; SU12662 (N-desethyl sunitinib); gastrointestinal stromal tumor; thrombocytopenia
Acute myeloid leukaemia results from the neoplastic transformation of haematopoietic stem cells. Although advances have been made in its treatment, the mortality rate remains high. As a result, therapeutic alternatives continue to be explored. In this study, we present evidence that suggests that casein, the principal protein in milk, possesses significant antileukaemic properties. We investigated whether casein inhibited the in vitro proliferation and induced the apoptosis of the mouse myelomonocytic leukaemia cell line WEHI-3. By contrast, under identical conditions, casein markedly promotes the proliferation of mouse normal mononuclear bone marrow cells. Since the selective elimination of leukaemia cells is an ideal therapeutic strategy, we also evaluated the antileukaemic potential of casein in vivo. The results showed that casein increases the survival of mice bearing WEHI-3-induced tumours, suggesting that this molecule is also capable of inhibiting the proliferation of these cells in vivo. The evidence that casein inhibited cell proliferation and induced apoptosis in leukaemia cells in vitro, but increased survival in vivo in a leukaemia mouse model, indicates that casein may be useful in leukaemia therapy.
sodium caseinate; BALB/c; acute myeloid leukaemia; cell line; leukaemia model
As a benign mesenchymal tumor, classic renal angiomyolipoma (AML) may obliterate the kidney parenchyma and cause renal hemorrhage. It has previously been reported that mesenchymal stem cells (MSCs) are involved in tumorigenesis; however, there have been no studies on stem cells with renal AML origin. In the present study, six females with classic renal AML received a partial or total nephrectomy. During surgery, tumor tissues were collected and culture expansion of adhesive fibroblastoid cells from these tissues was performed. We successfully isolated and cultured MSC-like cells from all six renal AML tumors. MSC characteristics, including morphology, immunophenotype and multidifferentiation potential were analyzed. Flow cytometry analysis revealed that these cells are highly similar to human bone marrow MSCs due to the expression of MSC-specific surface proteins, including CD29, CD44, CD73, CD90 and CD105. The stem cell-like nature of these cells is further supported by their adipogenic and osteogenic differentiation potentials when incubated in appropriate differentiation cocktails. Renal AML-derived adhesive cells possessing the characteristics of MSCs are described for the first time. They are a novel cell type which may be useful in future studies with regards to determining the role of stem cells in the formation and development of renal AML.
renal angiomyolipoma; renal tumor; mesenchymal stem cells
Extra-abdominal desmoid tumour is a rare type of tumour which is conventionally treated with wide local excision. However, wide local excision usually causes irreparable damage to the limbs. Nevertheless, few patients have been subjected to wide local excision and reconstructive surgery in a single procedure. A 52-year-old female reported a slow-growing lump in the right crus which had been growing for two years. The patient complained of persistent pain, especially at night, and was first subjected to lump resection in another hospital. The postoperative histopathological examination indicated muscle fibroma. Three months later, the lump recurred in the same position. The persistent pain induced by the tumour hampered her ability to walk. Wide local excision was conducted on the major parts of the gastrocnemius and soleus. After the pathological examination confirmed that the resection margin was negative, we performed reconstructive surgery on the Achilles tendon. The patient recovered plantarflexion function following the surgery and did not report any recurrence in the 6-year follow-up period. The desmoid tumour is a low-grade malignant tumour. Thus, the main focus of the treatment is to restore the function of the limbs to optimal capacity, such that the incidence of tumour recurrence is minimised.
desmoid tumour; surgery; aggressive fibromitosis; wide resection
The aim of this study was to investigate the mRNA and protein expression levels of the Syk gene as well as its promoter methylation in nasopharyngeal carcinoma (NPC) cell lines. The CNE-1 (highly differentiated), CNE-2 (poorly differentiated) and NP69 (non-cancerous human immortalized nasopharyngeal epithelial cells) cell lines were used in the present study. The MS-PCR, Q-RT-PCR and western blotting methods were used to examine the Syk gene promoter methylation levels and mRNA and protein expression in the three cell lines. The promoter methylation levels in CNE-1, CNE-2 and NP69 cells were 36%, 62% and 0, respectively. The mRNA levels in CNE-1 and CNE-2 cells were 42±3.5 and 28±2% of that in NP69, respectively; the protein levels in CNE-1 and CNE-2 cells were 36±4.5 and 16±2.5 of that in NP69, respectively; the statistical differences between groups were significant. The lower differentiation levels of the NPC cell lines correlate with lower levels of mRNA and protein expression of the Syk gene, as well as higher promoter methylation levels.
nasopharyngeal carcinoma; Syk gene; methylation; promoter
The aim of this study was to evaluate the volume doubling time (VDT) of lung cancer detected in our annual chest radiograph screening program and to compare it with those previously reported for computed tomography (CT) screening. In total, 209 patients who had a measurable tumor shadow and a history of participating in our chest radiograph mass screening program between 2006 and 2009 were included in this study. Indirect roentgenograms for patients with lung cancer were converted into digital images, and the section showing the tumor was enlarged on the monitor to a size of 0.01 mm. The mean VDT for all the patients was 158 days. Only 3.8% of the patients had a VDT of more than 400 days. In 140 patients with adenocarcinoma, the mean VDT was 177 days, and 5.0% of these patients had a VDT of more than 400 days. In the 44 patients with squamous cell carcinoma, the mean VDT was 133 days, and only 2.3% of these patients had a VDT of more than 400 days. These results were different from those previously reported for CT screening. In several reports on CT screening, more than 20% of the lung cancers had VDTs of more than 400 days. Since it is common knowledge that there are ‘indolent’ lung cancers with a VDT of more than 400 days, screening by annual chest radiography with rare overdiagnosis may need to be reconsidered.
volume doubling time; lung cancer; chest radiograph; mass screening program
The function of microsatellite instability (MSI) and the optimal panel of markers for epithelial ovarian cancer (EOC) are not well established. This study aimed to use the National Cancer Institute (NCI) markers BAT25, BAT26, D2S123, D5S346 and D17S250 to evaluate MSI in patients with ovarian serous cystadenocarcinoma, compared with ovarian serous cystadenoma and normal ovaries. A total of 37 patients were divided into three groups, as follows: cystadenocarcinoma (n=13), cystadenoma (n=10) and normal ovaries (n=14). DNA was extracted with TRIzol and quantified by spectrophotometry. MSI was evaluated by polymerase chain reaction (PCR), and classified as high (MSI-H), low (MSI-L) or stable (MSS). FIGO staging was I/II in 23.1% and III/IV in 76.9% of the cystadenocarcinoma group. Polymorphisms were found using at least one marker in 32 women, and were observed with D2S123 (83.7%), D17S250 (81.1%), D5S346 (72.9%), BAT25 (21.6%) and BAT26 (16.2%) markers. In the cystadenocarcinoma group, BAT25, BAT26, D2S123, D5S346 and D17S250 markers were positive in 30.8, 76.9, 53.8, 69.2 and 69.2% of patients, respectively. The same markers were positive in 30, 50, 40, 60 and 30% of the cystadenoma group, and 50, 71.4, 71.4, 64.3 and 63.3% in the normal ovary group, respectively. MSI-H was present in 84.6, 60 and 78.6% of the cystadenocarcinoma, cystadenoma and normal patients, respectively. MSI-L was detected in 0, 30 and 7.1%, and MSS was identified in 15.4, 10 and 14.3% of the cystadenocarcinoma, cystadenoma and normal patients, respectively. The frequency of MSI in both benign epithelial ovarian neoplasms and in normal ovaries was high, as well as in EOC, with no statistically significant difference between the groups. This suggests that MSI may arise as a consequence of the ovulatory process, and not solely as a feature of malignant ovarian tumors.
microsatellite instability; ovarian cancer; National Cancer Institute markers
In an attempt to clarify the clinical characteristics of synchronous primary endometrial and ovarian cancer (SPC), we reviewed the clinicopathological features of 13 cases treated in the Department of Gynecology and Obstetrics at Kyoto University Hospital over the last 6 years and compared them with 186 cases of primary uterine corpus cancer (PCC) and 136 cases of primary ovarian cancer (POC). Comparisons were performed based on clinicopathological factors, including age, BMI, parity, complication of thrombosis and FIGO stage. For SPC patients, the mean age was 51.5 years; 6 (46%) were nulliparous, and 7 (53%) had complicated thrombosis. All had well-differentiated endometrial cancer and 12 (92%) had endometrioid cancer in the ovary. The mean age of the SPC patients was significantly lower than that of the PCC patients (51.5 vs. 58.9 years). Thrombosis occurred in the SPC patients at a significantly higher rate than in both the PCC and POC patients. When the incidence of endometriosis and the regularity of menstruation were compared between patients who developed SPC with those who develop PCC at a young age (under 45 years), the SPC patients exhibited a significantly higher rate of endometriosis (100 vs. 35%), whereas the PCC patients exhibited a higher rate of irregular menstruation (53 vs. 15%, p=0.05). As for thrombosis, the age and FIGO stage of thrombosis-positive patients were significantly higher than those of thrombosis-negative patients in PCC and POC, while in SPC patients there was no such difference. In conclusion, this study demonstrated the differences in clinical features between SPC and PCC, and also novel features of SPC, namely endometriosis and thrombosis, which are essential in the management of this disease.
synchronous primary endometrial and ovarian cancer; endometriosis; thrombosis; thromboembolism; atypical endometriosis
Previous studies have demonstrated that interleukin-11 (IL-11) and the IL-11 receptor (IL-11R) are associated with the regulation of tumor progression and may play a significant role in bone metastases. The nonapeptide structure c(CGRRAGGSC) is a phage display-selected IL-11 mimic, which binds to IL-11R. The aim of this study is to investigate the binding characteristics of a cyclic nonapeptide c(CGRRAGGSC) in LNCaP human prostate cancer cells. To investigate its binding and uptake effects, c(CGRRAGGSC) was labeled with a fluorescent dye, LSS670. The binding location of LSS670 cyclic nonapeptide in LNCaP cells was investigated by fluorescence microscopy. Flow cytometry was used to detect the fluorescence of LSS670-c(CGRRAGGSC) in LNCaP cells. The binding of LSS670-c(CGRRAGGSC) in LNCaP cells was inhibited by unlabeled cyclic nonapeptide, depending on the varying density of c(CGRRAGGSC) and different time points. The molecular probe bound to the LNCaP cell membrane and cytoplasm through fluorescence tracing. In the saturation experiments performed in vitro, the Kd value was 3.2±0.02 nM and the Bmax value was 754±34 fmol/mg.pro. The 50% inhibiting concentration (IC50) was 6.31±0.12 nmol/l and the Ki value was 2.11±0.14 nmol/l in competitive inhibition experiments. Our results suggest that c(CGRRAGGSC) is able to specifically bind to LNCaP cells through a receptor-mediated pathway.
LNCaP cells; interleukin-11 analog; fluorescent dyes; prostate cancer
Severe hemorrhage following a prostatectomy is a rare and serious complication. A 63-year-old male with severe hemorrhage following radical prostatectomy which led to hypovolemic shock presented at our department and was treated with superselective internal iliac arterial embolization. At 6 months follow-up, the patient had recovered well, regained excellent urinary continence and the pelvic hematoma was absorbed using ultrasound examination. We concluded that rapid diagnosis by computed tomography angiography and early superselective embolization of internal iliac artery should be considered as the treatment of choice in severe hemorrhage cases following radical prostatectomy.
embolization; internal iliac arterial; hemorrhage; prostate; therapeutic
Inhibitor of growth 4 (ING4) is a member of the ING family and acts as a tumor suppressor protein. To investigate the impact of ING4 on breast cancer proliferation, the present study examined the antitumor effects caused by upregulation in the expression of ING4 and its possible mechanism of effect in MCF-7 cells. A plasmid-based expression system encoding the ING4 gene was used to construct a stable cell line and overexpress ING4 in MCF-7 cells. Real-time PCR and western blot analysis were used to detect the mRNA and protein expression levels of ING4, respectively. Cell growth was examined by methylthiazolyltetrazolium (MTT) assay. Cell cycle distribution and cell apoptosis were measured by flow cytometry. The expression of p21, p53 and bax genes were tested by real-time PCR and western blot analysis. The stably transfected cell lines pcDNA3.1(+)/ING4 (with the ING4 gene) and pcDNA3.1(+) (an empty vector) were established. The expression levels of ING4 mRNA and protein in the stable cell line expressing pcDNA3.1(+)/ING4 were significantly higher than those of the two control cell lines. The cell proliferation of stably transfected cells was inhibited, and the inhibitory rate was 62.58±2.93%. Based on the changes in cell cycle distribution in stably transfected cells compared with two control cell lines, a number of cells were blocked in the G0/G1 phase 67.82±3.78% (P<0.05). In addition, the apoptotic rate was significantly higher, at 31.51±3.02% (P<0.05). Real-time PCR revealed that p21 and bax mRNA expression were increased (P<0.01), but the expression of p53 did not change significantly (P>0.05) in the stably transfected cell lines. Western blot analysis results of p21, bax and p53 were in accordance with real-time PCR results. ING4 upregulation may inhibit breast cancer cell proliferation and accelerate the process of apoptosis. It is suggested that ING4 plays a significant role in the suppression of breast cancer progression.
inhibitor of growth 4; breast cancer; MCF-7
Cytotoxic nucleoside analogues are widely used in cancer chemotherapy. We used the cytosine arabinoside (Ara-C)-resistant erythroleukaemia cell line K562 and the Ara-C-sensitive myeloid leukaemia cell line HL60 to examine the differential expression of molecular markers. We found increased expression levels of deoxycytidine kinase (dCK) and human equilibrative nucleoside transporter 1 (hENT1) and decreased levels of multidrug resistance protein 5 (ABCC5) and ribonucleoside reductase subunit M1 (RRM1) expression in Ara-C-sensitive HL60 cells. We previously established the pemetrexed (MTA)-resistant small cell lung cancer cell lines PC6/MTA-0.4 and PC6/MTA-1.6 and found that MTA-resistant cells are more sensitive to gemcitabine (GEM) and Ara-C compared with parental PC-6 cells. We examined the molecular markers for GEM and Ara-C sensitivity in MTA-resistant cells and found increased gene expression of dCK and hENT1. Furthermore, treatment with MTA resulted in increased expression of dCK and hENT1 and decreased expression of ABCC5 and RRM1, concomitant with the alteration of the resistance to Ara-C in Ara-C-resistant K562 cells. These results provide evidence that the chemotherapeutic activity of the combination of MTA and cytotoxic nucleoside analogues is synergistic with regard to the alteration of metabolic molecules.
cytotoxic nucleoside analogue; pemetrexed; deoxycytidine kinase; human equilibrative nucleoside transporter 1; multidrug resistance protein 5; ribonucleoside reductase subunit M1
Epithelial ovarian cancer has the highest mortality of all gynecological cancers, and its progression is often without symptoms. Clinical outcome and survival may be improved if the disease is identified in the early stages. The objective of the study was to evaluate the utility of the serum biomarkers human epididymis protein 4 (HE4), soluble mesothelin-related protein (SMRP) and CA125 in the detection of ovarian cancer. In this retrospective study, the serum concentrations of CA125, HE4 protein and SMRP were measured in a cohort of 70 patients with epithelial ovarian cancer (EOC) compared with 78 healthy controls. Median serum levels of CA125 for ovarian cancer cases were 503.55±560.7 U/ml vs. 9.28±14.47 U/ml in the control group (p<0.001); for SMRP 5.13±7.64 nM vs. 1.02±0.89 nM (p<0.01); and for HE4 597.95±934.59 pM vs. 56.75±43.79 pM (p<0.001), respectively. Positive correlations between the clinical stage of EOC and CA125, HE4 and SMRP serum concentrations were found [(R=0.83; p<0.001); (R=0.64; p<0.001); (R=0.45; p<0.001), respectively]. Data analysis for the whole study group also revealed a significant correlation between plasma concentrations of CA125 and HE4 (R=0.45; p<0.001), between CA125 and SMRP (R=0.38; p<0.001) as well as HE4 and SMRP (R=0.51; p<0.001). Similar significant correlations between serum biomarker concentrations were also found in the ovarian cancer group [CA125 and HE4 (R=0.31; p<0.01); CA125 and SMRP (R=0.25; p<0.05); HE4 and SMRP (R=0.44, p<0.001), respectively]. A significant correlation was observed between the serous histological type of EOC and serum concentration of HE4 in the study group compared with other non-serous types of ovarian cancer (p<0.01). In conclusion, measuring CA125 in combination with new biomarkers such as SMRP and HE4 may improve the accuracy of ovarian cancer diagnosis, particularly in early detection of the disease.
ovarian cancer; biomarkers; human epididymis protein 4; soluble mesothelin related protein; CA125
Ganglioneuromas (GNs) arising from neural crest sympathogonia are rare benign neurogenic tumors. The most commonly affected sites are the posterior mediastinum, the retroperitoneum and the adrenal gland. GNs often present as a solitary, painless and slow-growing mass, and multiple occurrences in the cervical region are extremely rare. Here, we report a case of massive multiple cervical GN in a 4-year-old girl, and review cases of cervical GN that have been reported in the past 10 years. The results demonstrated that cervical GN, compared to other sites, is seldom secretory. The signs and symptoms of cervical GN are unspecific; the ultimate diagnosis of GN depends on pathological examination. Fine-needle aspiration biopsy has limited value in diagnosis. Surgical excision is the treatment of choice and the prognosis is excellent even in cases where complete excision cannot be achieved. Furthermore, GNs should be considered in patients with multiple masses in the neck.
neurogenic tumor; ganglioneuroma; children; cervical
The purpose of this study was to investigate the mechanism of glioma cell invasion in hypoxic conditions. We demonstrated that hypoxia increased cell invasion, matrix metalloproteinase-2 (MMP2) activity and time-dependent expression of hypoxia inducible factor-1α (HIF-1α) in human glioma cells. These data suggest that MMP2 may play a significant role in tumor invasion in hypoxic conditions. We investigated the mechanisms involved in the increased MMP2 activity and cell invasion in hypoxic conditions. Increased expression of phospho-Jun NH2-terminal kinase (p-JNK) and phospho-c-Jun (p-c-Jun) in glioma cells induced by hypoxia was detected. Furthermore, this effect may be reduced by inhibiting the JNK signaling pathway. We found that inhibition of RhoA geranylgeranylation by geranylgeranyltransferase inhibitor-2147 (GGTI-2147) or knockdown of RhoA by siRNA against RhoA reduced the expression of p-JNK and p-c-Jun, and decreased MMP2 activity and glioma cell invasion in hypoxic conditions. These data suggest a link among RhoA, JNK, c-Jun and MMP2 activity that is functionally involved in the increased glioma cell invasion induced by hypoxia.
glioma; cell invasion; hypoxia; U251 cell line; RhoA-JNK signaling pathway; matrix metalloproteinase-2
Transitional cell carcinoma (TCC) is one of the most common types of malignancies and a leading cause of genitourinary system cancer mortality worldwide. The tumor suppressor gene FOXO1, a member of the forkhead box O (FOXO) subfamily of transcription factors, is downregulated in a number of cancers, including TCC; however, the underlying mechanisms are poorly understood. In the present study, we used microRNA (miRNA) target prediction algorithms to identify a conserved potential miR-96 binding site in the 3′-untranslated region (3′-UTR) of FOXO1. Using quantitative real-time PCR (qRT-PCR) and northern blot analysis, we identified that miR-96 was downregulated in TCC tissues compared to normal bladder tissues (NB), suggesting that the loss of FOXO1 expression in TCC may be mediated by miR-96. To confirm this, we transfected pre-miR-96/anti-miR-96 into the T24 TCC cell line and revealed that miR-96 expression was sufficient to significantly reduce FOXO1 expression. Conversely, FOXO1 expression was not completely restored by the inhibition of miR-96 in T24 cells. Moreover, RNA silencing of FOXO1 significantly reduced miR-96 inhibitor-mediated T24 cell apoptosis. In conclusion, our study demonstrates that the miR-96 targeting of FOXO1 is upregulated in TCC; in addition, TCC tumorigenesis may be partly due to the ability of miR-96 to promote FOXO1 repression, thereby bypassing cell apoptosis controls.
bladder transitional cell carcinoma; miR-96; forkhead box protein O1; apoptosis