Aloin inhibits gastric cancer cell proliferation and migration by suppressing the STAT3/HMGB1 signaling pathway / 南方医科大学学报

OBJECTIVE@#To investigate the molecular mechanism underlying the inhibitory effect of aloin on the proliferation and migration of gastric cancer cells.@*METHODS@#Human gastric cancer MGC-803 cells treated with 100, 200 and 300 μg/mL aloin were examined for changes in cell viability, proliferation and migration abilities using CCK-8, EdU and Transwell assays. HMGB1 mRNA level in the cells was detected with RT-qPCR, and the protein expressions of HMGB1, cyclin B1, cyclin E1, E-cadherin, MMP-2, MMP-9 and p-STAT3 were determined using Western blotting. JASPAR database was used to predict the binding of STAT3 to HMGB1 promoter. In a BALB/c-Nu mouse model bearing subcutaneous MGC-803 cell xenograft, the effect of intraperitoneal injection of aloin (50 mg/kg) on tumor growth was observed. The protein expressions of HMGB1, cyclin B1, cyclin E1, E-cadherin, MMP-2, MMP-9 and p-STAT3 in the tumor tissue was examined using Western blotting, and tumor metastasis in the liver and lung tissues was detected using HE staining.@*RESULTS@#Treatment with aloin concentration-dependently inhibited the viability of MGC-803 cells (P < 0.05), significantly reduced the number of EdU-positive cells (P < 0.01), and attenuated the migration ability of the cells (P < 0.01). Aloin treatment dose-dependently down-regulated HMGB1 mRNA expression (P < 0.01), lowered the protein expressions of HMGB1, cyclin B1, cyclin E1, MMP-2, MMP-9 and p-STAT3, and up-regulated E-cadherin expression in MGC-803 cells. Prediction based on JASPAR database suggested that STAT3 could bind to the promoter region of HMGB1. In the tumor-bearing mice, aloin treatment significantly reduced the tumor size and weight (P < 0.01), lowered the protein expressions of cyclin B1, cyclin E1, MMP-2, MMP-9, HMGB1 and p-STAT3 and increased the expression of E-cadherin in the tumor tissue (P < 0.01).@*CONCLUSION@#Aloin attenuates the proliferation and migration of gastric cancer cells by inhibiting the STAT3/HMGB1 signaling pathway.

Regorafenib Suppresses Migration of and Induces Cell Cycle Arrest and Apoptosis in MCF-7 Cells

This study investigated the mechanism underlying the suppression of estrogen receptor-positive MCF-7 cell growth by regorafenib. MCF-7 cells were treated with regorafenib, and the effect of regorafenib on multiple cancer-associated pathways was evaluated. Although regorafenib effectively inhibited the proliferation of MCF-7 cells, it had no effect on the proliferation of the normal breast epithelial cell line MCF10A. Regorafenib suppressed MCF-7 cell migration, probably by regulating the homeostatic expression of matrix metalloproteinases and the tissue inhibitor of MMPs. Furthermore, it upregulated p21 expression, downregulated cyclin B1 and cyclin D1 expresssions, and caused cell cycle arrest. In addition, regorafenib induced apoptosis in MCF-7 cells by reducing Mcl-1 expression and activating caspase signaling. These results demonstrate that regorafenib has the potential to be an effective drug for treating breast cancer

Protein Expression of Cyclin B1, Transferrin Receptor, and Fibronectin Is Correlated with the Prognosis of Adrenal Cortical Carcinoma

BACKGROUND: Adrenal cortical carcinoma (ACC) is a rare cancer with a variable prognosis. Several prognostic factors of ACC have been previously reported, but a proteomic analysis has not yet been performed. This study aimed to investigate prognostic biomarkers for ACC using a proteomic approach.METHODS: We used reverse-phase protein array data from The Cancer Proteome Atlas, and identified differentially expressed proteins in metastatic ACCs. Multivariate Cox regression analysis adjusted by age and staging was used for survival analysis, and the C-index and category-free net reclassification improvement (cfNRI) were utilized to evaluate additive prognostic value.RESULTS: In 46 patients with ACC, cyclin B1, transferrin receptor (TfR1), and fibronectin were significantly overexpressed in patients with distant metastasis. In multivariate models, high expression of cyclin B1 and TfR1 was significantly associated with mortality (hazard ratio [HR], 6.13; 95% confidence interval [CI], 1.02 to 36.7; and HR, 6.59; 95% CI, 1.14 to 38.2; respectively), whereas high fibronectin expression was not (HR, 3.92; 95% CI, 0.75 to 20.4). Combinations of high cyclin B1/high TfR1, high cyclin B1/high fibronectin, and high TfR1/high fibronectin were strongly associated with mortality ([HR, 13.72; 95% CI, 1.89 to 99.66], [HR, 9.22; 95% CI, 1.34 to 63.55], and [HR, 18.59; 95% CI, 2.54 to 135.88], respectively). In reclassification analyses, cyclin B1, TfR1, fibronectin, and combinations thereof improved the prognostic performance (C-index, 0.78 to 0.82–0.86; cfNRI, all P values <0.05).CONCLUSION: In ACC patients, the overexpression of cyclin B1, TfR1, and fibronectin and combinations thereof were associated with poor prognosis.

Discovery of an Indirubin Derivative as a Novel c-Met Kinase Inhibitor with In Vitro Anti-Tumor Effects

The c-Met protein is a receptor tyrosine kinase involved in cell growth, proliferation, survival, and angiogenesis of several human tumors. Overexpression of c-Met has been found in gastric cancers and correlated with a poor prognosis. Indirubin is the active component of Danggui Longhui Wan, which is a traditional Chinese antileukemic recipe. In the present study, we tested the anti-cancer effects of an indirubin derivative, LDD-1937, on human gastric cancer cells SNU-638. When we performed the in vitro kinase assay against the c-Met activity, LDD-1937 inhibited the activity of c-Met. This result was confirmed by immunoblot and immunofluorescence of phosphorylated c-Met. Immunoblot analysis showed that LDD-1937 decreased the expression of the Erk1/2, STAT3, STAT5, and Akt, downstream proteins of c-Met. In addition, LDD-1937 reduced the cell viability and suppressed colony formation and migration of SNU-638 cells. Furthermore, LDD-1937 induced G2/M phase arrest in the SNU-638 cells by decreasing the expression levels of cyclin B1 and CDC2. Cleaved-PARP, an apoptosis-related protein, was up-regulated in cells treated with LDD-1937. Overall, this study suggests that LDD-1937 may be a novel small-molecule with therapeutic potential for selectively inhibiting c-Met and c-Met downstream pathways in human gastric cancers overexpressing c-Met.

Helicobacter pylori inhibited cell proliferation in human periodontal ligament fibroblasts through the Cdc25C/CDK1/cyclinB1 signaling cascade

PURPOSE: Several studies have shown that the oral cavity is a secondary location for Helicobacter pylori colonization and that H. pylori is associated with the severity of periodontitis. This study investigated whether H. pylori had an effect on the periodontium. We established an invasion model of a standard strain of H. pylori in human periodontal ligament fibroblasts (hPDLFs), and evaluated the effects of H. pylori on cell proliferation and cell cycle progression. METHODS: Different concentrations of H. pylori were used to infect hPDLFs, with 6 hours of co-culture. The multiplicity of infection in the low- and high-concentration groups was 10:1 and 100:1, respectively. The Cell Counting Kit-8 method and Ki-67 immunofluorescence were used to detect cell proliferation. Flow cytometry, quantitative real-time polymerase chain reaction, and western blots were used to detect cell cycle progression. In the high-concentration group, the invasion of H. pylori was observed by transmission electron microscopy. RESULTS: It was found that H. pylori invaded the fibroblasts, with cytoplasmic localization. Analyses of cell proliferation and flow cytometry showed that H. pylori inhibited the proliferation of periodontal fibroblasts by causing G2 phase arrest. The inhibition of proliferation and G2 phase arrest were more obvious in the high-concentration group. In the low-concentration group, the G2 phase regulatory factors cyclin dependent kinase 1 (CDK1) and cell division cycle 25C (Cdc25C) were upregulated, while cyclin B1 was inhibited. However, in the high-concentration group, cyclin B1 was upregulated and CDK1 was inhibited. Furthermore, the deactivated states of tyrosine phosphorylation of CDK1 (CDK1-Y15) and serine phosphorylation of Cdc25C (Cdc25C-S216) were upregulated after H. pylori infection. CONCLUSIONS: In our model, H. pylori inhibited the proliferation of hPDLFs and exerted an invasive effect, causing G2 phase arrest via the Cdc25C/CDK1/cyclin B1 signaling cascade. Its inhibitory effect on proliferation was stronger in the high-concentration group.

Genistein Inhibits Proliferation of BRCA1 Mutated Breast Cancer Cells: The GPR30-Akt Axis as a Potential Target

BACKGROUND: BRCA1 mutated breast cancer cells exhibit the elevated cell proliferation and the higher metastatic potential. G protein-coupled receptor 30 (GPR30) has been shown to regulate growth of hormonally responsive cancers, such as ovarian and breast cancers, and high expression of GPR30 is found in estrogen receptor (ER)-negative breast cancer cells. ER-negative breast cancer patients often have a mutation in the tumor suppressor gene, BRCA1. This study explored antiproliferative effects of genistein, a chemopreventive isoflavone present in legumes, and underlying molecular mechanisms in triple negative breast cancer cells with or without functionally active BRCA1.METHODS: Expression of BRCA1, GPR30 and Nrf2 was measured by Western blot analysis. Reactive oxygen species (ROS) accumulation was monitored by using the fluorescence-generating probe, 2’,7’-dichlorofluorescein diacetate. The effects of genistein on breast cancer cell viability and proliferation were assessed by the MTT, migration and clonogenic assays.RESULTS: The expression of GPR30 was dramatically elevated at both transcriptional and translational levels in BRCA1 mutated breast cancer cells compared to cells with wild-type BRCA1. Notably, there was diminished Akt phosporylation in GPR30 silenced cells. Treatment of BRCA1 silenced breast cancer cells with genistein resulted in the down-regulation of GPR30 expression and the inhibition of Akt phosphorylation as well as the reduced cell viability, migration and colony formation. Genistein caused cell cycle arrest at the G₂/M phase in BRCA1-mutant cells through down-regulation of cyclin B1 expression. Furthermore, BRCA1-mutant breast cancer cells exhibited higher levels of intracellular ROS than those in the wild-type cells. Genistein treatment lowered the ROS levels through up-regulation of Nrf2 expression.CONCLUSIONS: Lack of functional BRCA1 activates GPR30 signaling, thereby stimulating Akt phosphorylation and cell proliferation. Genistein induces G2/M phase arrest by down-regulating cyclin B1 expression, which is attributable to its suppression of GPR30 activation and Akt phosphorylation in BRCA1 impaired breast cancer cells.

Artesunate inhibits proliferation of glioblastoma cells by arresting cell cycle / 中国中药杂志

Glioblastoma is a common brain tumor and the overall survival rate of the patients is very low, so it is an effective way to develop the potential chemotherapy or adjuvant chemotherapy drugs in glioblastoma treatment. As a well-known antimalarial drug, artesunate(ARTs) has clear side effects, and recently it has been reported to have antitumor effects, but rarely reported in glioblastoma. Different concentrations of ARTs were used to treat the glioblastoma cells, and then the inhibitory effect of ARTs on glioblastoma proliferation was detected by MTT assay; Ki67 immunofluorescence assay was used to detect the proliferation of cells; Soft agar experiment was used to explain the clonal formation abilities ; Flow Cytometry was used to detect the cell cycle; and Western blot assay was used to determine the expression of key cell cycle protein. MTT assay results indicated that ARTs-treated glioblastoma cell A172, U251, U87 were significantly inhibited in a time-and-dose dependent manner as compared to the control group(DMSO treatment group). Soft agar experiment showed that ARTs could significantly reduce the clonal formation ability of glioblastoma. Furthermore, Flow cytometry analysis showed that ARTs could obviously increase the cell proportion in G₀/G₁ phase and reduce the cell proportion in S phase. Western blot results showed that the expressions of cell cycle-related proteins CDK2, CDK4, cyclin D1 and cyclin B1 were all obviously down-regulated. Above all, ARTs may inhibit the proliferation of glioblastoma cells by arresting cell cycle in G₀/G₁ phase through down-regulating the expression of CDK2, CDK4, cyclin D1, cyclin B1. These results may not only provide a novel method for rediscovering and reusing ARTs but also provide a new potential drug for treating glioblastoma.

A Novel Germline Mutation in BRCA1 Causes Exon 20 Skipping in a Korean Family with a History of Breast Cancer / 한국유방암학회지

Germline mutations in the BRCA1 and BRCA2 genes are strong genetic factors for predispositions to breast, ovarian, and other related cancers. This report describes a family with a history of breast and ovarian cancers that harbored a novel BRCA1 germline mutation. A single nucleotide deletion in intron 20, namely c.5332+4delA, was detected in a 43-year-old patient with breast cancer. This mutation led to the skipping of exon 20, which in turn resulted in the production of a truncated BRCA1 protein that was 1773 amino acids in length. The mother of the proband had died due to ovarian cancer and had harbored the same germline mutation. Ectopically expressed mutant BRCA1 protein interacted with the BARD1 protein, but showed a reduced transcriptional function, as demonstrated by the expression of cyclin B1. This novel germline mutation in the BRCA1 gene caused familial breast and ovarian cancers.

Cellular Effects of Troglitazone on YD15 Tongue Carcinoma Cells

An FDA approved drug for the treatment of type II diabetes, Troglitazone (TRO), a peroxisome proliferator–activated receptor gamma agonist, is withdrawn due to severe idiosyncratic hepatotoxicity. In the search for new applications of TRO, we investigated the cellular effects of TRO on YD15 tongue carcinoma cells. TRO suppressed the growth of YD15 cells in the MTT assay. The inhibition of cell growth was accompanied by the induction of cell cycle arrest at G₀/G₁ and apoptosis, which are confirmed by flow cytometry and western blotting. TRO also suppressed the expression of cell cycle proteins such as cyclin D1, cdk2, cdk4, cyclin B1, cdk1(or cdc2), cyclin E1 and cyclin A. The inhibition of cell cycle proteins was coincident with the up-regulation of p21(CIP1/WAF1) and p27(KIP1). In addition, TRO induces the activation of caspase-3 and caspase-7, as well as the cleavage of PARP. Further, TRO suppressed the expressions of Bcl-2 without affecting the expressions of Bad and Bax. Overall, our data supports that TRO induces cell cycle arrest and apoptosis on YD15 cells.

Circadian rhythm variation of the clock genes Per1 and cell cycle related genes in different stages of carcinogenesis of buccal mucosa in animal model / 中华口腔医学杂志

<p><b>OBJECTIVE</b>To investigate the expression and circadian rhythm variation of biological clock gene Per1 and cell cycle genes p53, CyclinD1, cyclin-dependent kinases (CDK1), CyclinB1 in different stages of carcinogenesis in buccal mucosa and its relationship with the development of buccal mucosa carcinoma.</p><p><b>METHODS</b>Ninety golden hamsters were housed under 12 hours light-12 hours dark cycles, and the model of buccal squamous cell carcinoma was established by using the dimethylbenzanthracene (DMBA) to smear the golden hamster buccal mucosa. Before the DMBA was used and after DMBA was used 6 weeks and 14 weeks respectively, the golden hamsters were sacrificed at 6 different time points (5 rats per time point) within 24 hour, including 4, 8, 12, 16, 20 and 24 hour after lights onset (HALO), and the normal buccal mucosa, precancerous lesions and cancer tissue were obtained, respectively. HE stained sections were prepared to observe the canceration of each tissue. Real time RT-PCR was used to detect the mRNA expression of Per1, p53, CyclinD1, CDK1 and CyclinB1, and a cosine analysis method was applied to determine the circadian rhythm variation of Per1, p53, CyclinD1, CDK1 and CyclinB1 mRNA expression, which were characterized by median, amplitude and acrophase.</p><p><b>RESULTS</b>The expression of Per1, p53, CDK1 and CyclinD1 mRNA in 6 different time points within 24 hours in the tissues of three different stages of carcinogenesis had circadian rhythm, respectively. However, the CyclinB1 mRNA was expressed with circadian rhythm just in normal and cancer tissue (P < 0.05), while in precancerous lesions the circadian rhythm was in disorder (P > 0.05). As the development of carcinoma, the median of Per1 and p53 mRNA expression were significantly decreased (P < 0.05), yet the median of CDK1, CyclinB1 and CyclinD1 mRNA expression were significantly increased (P < 0.05). The amplitude of Per1, p53 and CyclinD1 mRNA expression was significantly decreased as the development of carcinoma (P < 0.05), however the amplitude of CDK1 mRNA expression was significantly increased (P < 0.05). In addition, there was no significant difference in the amplitude of CyclinB1 mRNA expression. The time that the peak expression value of Per1 and CDK1 mRNA appeared (Acrophase) in precancerous lesions was remarkably earlier than that in normal tissues, but the acrophase of p53 and CyclinD1 mRNA expression was remarkably delayed. Moreover, the acrophase of CDK1 and CyclinB1 mRNA expression in cancer tissues was obviously earlier than that in normal tissues, yet there was no significant variation in acrophase of Per1, p53, CyclinD1 mRNA expression between normal tissues and cancer tissues.</p><p><b>CONCLUSIONS</b>The circadian rhythm of clock gene Per1 and cell cycle genes p53, CyclinD1, CDK1, CyclinB1 expression remarkably varied with the occurrence and development of carcinoma. Further research into the interaction between circadian and cell cycle of two cycle activity and relationship with the carcinogenesis may providenew ideas and methods of individual treatment and the mechanism of carcinogenesis.</p>

Effects of β-catenin on differentially expressed genes in multiple myeloma / 华中科技大学学报（医学）（英德文版）

This study aimed to identify the differentially expressed genes after silencing of β-catenin in multiple myeloma transduced with β-catenin shRNA. The DNA microarray dataset GSE17385 was downloaded from Gene Expression Omnibus, including 3 samples of MM1.S (human multiple myeloma cell lines) cells transduced with control shRNA and 3 samples of MM1.S cells transduced with β-catenin shRNA. Then the differentially expressed genes (DEGs) were screened by using Limma. Their underlying functions were analyzed by employing Gene Ontology and Kyoto Encyclopedia of Genes and Genomes pathway enrichment analyses. Moreover, DEGs annotation was conducted based on the databases of tumor associated genes, tumor suppressed genes and the transcriptional regulation from patterns to profiles. Furthermore, the protein-protein interaction (PPI) relationship was obtained from STRING and the protein-protein interaction network and the functional modules were visualized by Cytoscape. Then, the pathway enrichment for the DEGs in the functional module was performed. A total of 301 DEGs, including 124 up-regulated and 117 down-regulated DEGs, were screened. Functional enrichment showed that CCNB1 and CDK1 were significantly related to the function of cell proliferation. FOS and JUN were related to innate immune response-activating signal transduction. Pathway enrichment analysis indicated that CCNB1 and CDK1 were most significantly enriched in the pathway of cell cycle. Besides, FOS and JUN were significantly enriched in the Toll-like receptor signaling pathway. FOXM1 was identified as a transcription factor. Moreover, there existed interactions among CCNB1, FOXM1 and CDK1 in PPI network. The expression of FOS, JUN, CCNB1, FOXM1 and CDK1 may be affected by β-catenin in multiple myeloma.

Fucoidan induces apoptosis of HepG2 cells by down-regulating p-Stat3 / 华中科技大学学报（医学）（英德文版）

Fucoidan is one of the main bioactive components of polysaccharides. The current study was focused on the anti-tumor effects of fucoidan on human heptoma cell line HepG2 and the possible mechanisms. Fucoidan treatment resulted in cell cycle arrest and apoptosis of HepG2 cells in a dose-dependent manner detected by MTT assay, flow cytometry and fluorescent microscopy. The results of flow cytometric analysis revealed that fucoidan induced G2/M arrest in the cell cycle progression. Hoechst 33258 and Annexin V/PI staining results showed that the apoptotic cell number was increased, which was associated with a dose-dependent up-regulation of Bax and down-regulation of Bcl-2 and p-Stat3. In parallel, the up-regulation of p53 and the increase in reactive oxygen species were also observed, which may play important roles in the inhibition of HepG2 growth by fucoidan. In the meantime, Cyclin B1 and CDK1 were down-regulated by fucoidan treatment. Down-regulation of p-Stat3 by fucoidan resulted in apoptosis and an increase in ROS in response to fucoidan exposure. We therefore concluded that fucoidan induces apoptosis through the down-regulation of p-Stat3. These results suggest that fucoidan may be used as a novel anti-cancer agent for hepatocarcinoma.

Different responses of cell cycle between rat vascular smooth muscle cells and vascular endothelial cells to paclitaxel / 华中科技大学学报（医学）（英德文版）

Although previous reports showed drug-eluting stent (DES) could effectively inhibit neointima formation, in-stent restenosis (ISR) remains an important obstacle. The purpose of this study was to investigate different effects of paclitaxel on proliferation and cell cycle regulators between vascular smooth muscle cells (VSMCs) and vascular endothelial cells (VECs) of rats in vitro. The cultured VSMCs and VECs of rats from the same tissues were examined by using immunohistochemistry, flow cytometry and Western blotting in control and paclitaxel-treated groups. The results showed paclitaxel could effectively inhibit proliferation of VSMCs and VECs. However, as compared with VECs, proliferation of VSMCs in paclitaxel-treated group decreased less rapidly. The percentage of cells in G0-G1 and G2-M phases was reduced, and that in S phase increased after treatment for 72 h. The expression of cyclin D1 and B1, p27 and PCNA in VSMCs of paclitaxel-treated group was up-regulated, but that of p21 down-regulated as compared with VECs. It is concluded that there are significant differences in the expression of cell cycle regulators and proliferation rate between paclitaxel-treated VSMCs and paclitaxel-treated VECs, suggesting that the G1-S checkpoint regulated by paclitaxel may play a critical role in the development of complications of DES, which provides new strategies for treatments of ISR.

Enhanced growth inhibition by combined two pathway inhibitors on K-ras mutated non-small cell lung cancer cells / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To evaluate the effect of combined targeting of MEK and PI3K signaling pathways on K-ras mutated non-small cell lung cancer cell line A549 cells and the relevant mechanisms.</p><p><b>METHODS</b>A549 cells were treated with different concentrations of two inhibitors. Growth inhibition was determined by MTT assay. According to the results of MTT test, the cells were divided into four groups: the control group, PI3K inhibitor group (GDC-0941,0.5 and 5.0 µmol/L), combination group I (0.5 µmol/L AZD6244+0.5 µmol/L GDC-0941) and combination group II (5.0 µmol/L AZD6244+5.0 µmol/L GDC-0941). The cell cycle and apoptosis were analyzed by flow cytometry. The expression of proteins related to apoptosis was tested with Western blot.</p><p><b>RESULTS</b>Both GDC-0941 and AZD6244 inhibited the cell proliferation. The combination group II led to a stronger growth inhibition. The combination group I showed an antagonistic effect and combination group II showed an additive or synergistic effect. Compared with the control group, the combination group I led to reduced apoptotic rate [(20.70 ± 0.99)% vs. (18.65 ± 0.92 )%, P > 0.05]; Combination group II exhibited enhanced apoptotic rate [(37.85 ± 3.18)% vs. (52.27 ± 4.36)%, P < 0.01]. In addition, in the combination group II, more A549 cells were arrested in G0/G1 phase and decreased S phase (P < 0.01), due to the reduced expressions of CyclinD1 and Cyclin B1, the increased cleaved PARP and the diminished ratio of Bcl-2/Bax.</p><p><b>CONCLUSIONS</b>For single K-ras mutated NSCLC cell line A549 cells, combination of RAS/MEK/ERK and PI3K/AKT/mTOR inhibition showed synergistic effects depending on the drug doses. Double pathways targeted therapy may be beneficial for these patients.</p>

Regulation mechanism of eIF3 P170 on developing myocardial cell cycle / 中南大学学报(医学版)

OBJECTIVE@#To investigate the expression of eIF3P170, cdc2, cyclinB1 and cyclinD1 in developing cardiac myocytes, and the correlation between eIF3P170 with cdc2, cyclin D1, and cyclin B1 in mice.@*METHODS@#Mouse cardiac myocytes were obtained at different time points. RT-PCR was employed to detect the expression of eIF3P170, cdc2, cyclin D1 and cyclin B1 mRNA.@*RESULTS@#Expressions of eIF3P170, cdc2, cyclinD1 and cyclinB1 mRNA were higher in the embryonic Day 13, 15, 18 and postnatal Day 1, 2, 3, 5. Expressions at postnatal Day 5 reached the highest (all P values<0.05 vs other time points), and then the expressions of these genes gradually decreased to the weakest at postnatal Day 30 (all P values<0.05 vs other time points). The mRNA expression of eIF3P170 was positively correlated with cdc2, cyclin D1 and cyclin B1 mRNA expression respectively.@*CONCLUSION@#The mRNA expressions of eIF3 P170, cdc2, cyclin D1 and cyclin B1 in the embryo and the early life after birth are high. They reach the maximum at postnatal Day 5, then gradually decreased.

Types of primary cyclins expressed in gastric carcinoma and their relationship with clinicopathological features / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To characterize the human primary cyclins (D1, E, A, B1) expressed in gastric carcinoma, and to clarify the relationship between the types of expressed primary cyclins and clinicopathological features of gastric carcinoma.</p><p><b>METHODS</b>Primary cyclins (D1, E, A, B1) expressed in single cells separated from 68 cases gastric carcinoma tissues were analyzed by flow cytometry. We classified the gastric carcinomas by different types of the expressed primary cyclins, and explore the roles of primary cyclins expressed in cell cycle and the expression patterns of the cyclins. The results were analyzed together with clinicopathological features.</p><p><b>RESULTS</b>The patterns of expressed primary cyclins could be classified into five types. The proportion was 10.3% (7/68), 22.1% (15/68), 25.0% (17/68), 29.4% (20/68), and 13.2% (9/68), respectively, from type I to type V. Each type could be, according to the degree of in-cycle cyclins expressed, divided into different sub-types. The types of primary cyclins expressed were strongly linked to invasive depth and lymph node metastasis of the gastric carcinoma (P < 0.01). The rates of lymph node metastasis were 26.6%, 43.8%, 82.3%, 95.0%, and 100.0%, respectively, from type I to type V. The type of primary cyclins expressed was also significantly associated with disease stage (TNM stage). The proportion of stage IV disease was 0, 6.7%, 17.6%, 25.0% and 55.6%, respectively, from type I to type V. It was shown that there were relationships between the sub-types of primary cyclins expressed and different growth-types, degree of cell differentiation, or, the tumor gross types (P < 0.01).</p><p><b>CONCLUSIONS</b>The types of primary cyclins expression are different in the process of the occurrence, development and metastasis of gastric carcinoma, and are correlated with clinicopathological features of gastric carcinoma.</p>

Growth inhibition of combined pathway inhibitors on KRAS mutated non-small cell lung cancer cell line / 中华病理学杂志

<p><b>OBJECTIVE</b>To investigate the effect of the selective PI3K inhibitor and MEK inhibitor on KRAS and PTEN co-mutated non-small cell lung cancer cell line NCI-H157 and the relevant mechanisms.</p><p><b>METHODS</b>NCI-H157 was cultured routinely and treated with different concentrations of the two inhibitors. Cell proliferation was detected by MTT cell cycle assay. Based on the MTT results the cells were divided into four groups: the control group, PI3K inhibitor group (GDC-0941, 0.5 and 5.0 µmol/L), combination group I (0.5 µmol/L AZD6244 + 0.5 µmol/L GDC-0941) and combination group II (5.0 µmol/L AZD6244 + 5.0 µmol/L GDC-0941). Colony formation assay was performed to detect colony formation efficiency. The cell cycle and apoptosis were analyzed by flow cytometry. The expression of protein related to apoptosis was tested with Western blot.</p><p><b>RESULTS</b>Cell growth was inhibited by the two inhibitors. Combination groups led to stronger cell proliferation inhibition: combination group Ishowed synergistic effect of their actions and combination group II showed an additive effect; in both groups, there were decreased colony number [(77.2 ± 1.54)/well vs (61.50 ± 2.12)/well, P < 0.01] and [(51.00 ± 4.00)/ well vs (22.50 ± 3.53)/well, P < 0.01]; and enhanced apoptotic ratios [(18.30 ± 0.82)% vs (21.32 ± 0.56)%, P < 0.01] and [(27.14 ± 1.58)% vs (42.45 ± 4.42)%, P < 0.01]. In addition, compared to the PI3K inhibitor alone group, the NCI-H157 cells in the combination groups showed increased G0/G1 phase and decreased S phase (P < 0.01). Western blotting showed that the combination groups demonstrated significantly decreased expression of cyclin D1 and cyclin B1, increased p21 and cleaved PARP and decreased bcl-2/bax ratio, compared to the PI3K inhibitor only group.</p><p><b>CONCLUSION</b>The combined inhibition of PI3K (AZD6244) and MEK (GDC-0941) has synergistic effects on the proliferation of NCI-H157 cells, but such effects appear to be in a dose-dependent manner.</p>

Ethanol Elicits Inhibitory Effect on the Growth and Proliferation of Tongue Carcinoma Cells by Inducing Cell Cycle Arrest

Cellular effects of ethanol in YD-15 tongue carcinoma cells were assessed by MTT assay, caspase activity assay, Western blotting and flow cytometry. Ethanol inhibited the growth and proliferation of YD-15 cells in a dose- and time-dependent manner in an MTT assay. The effects of ethanol on cell cycle control at low percent range of ethanol concentration (0 to 1.5%), the condition not inducing YD-15 cell death, was investigated after exposing cells to alcohol for a certain period of time. Western blotting on the expression of cell cycle inhibitors showed that p21 and p27 was up-regulated as ethanol concentration increases from 0 to 1.5% whilst the cell cycle regulators, cdk1, cdk2, and cdk4 as well as Cyclin A, Cyclin B1 and Cyclin E1, were gradually down-regulated. Flow cytometric analysis of cell cycle distribution revealed that YD-15 cells exposed to 1.5% ethanol for 24 h was mainly arrested at G2/M phase. However, ethanol induced apoptosis in YD-15 cells exposed to 2.5% or higher percent of ethanol. The cleaved PARP, a marker of caspase-3 mediated apoptosis, and the activation of caspase-3 and -7 were detected by caspase activity assay or Western blotting. Our results suggest that ethanol elicits inhibitory effect on the growth and proliferation of YD-15 tongue carcinoma cells by mediating cell cycle arrest at G2/M at low concentration range and ultimately induces apoptosis under the condition of high concentration.

Expression of Cyclin A, B1, D1, D3, and E in Non-Small Lung Cancers

PURPOSE: Cyclins, and their associated cyclin dependent kinases, regulate progression of the cell cycle through the G1 phase and into the S-phase during the DNA replication process. Cyclin E regulation is an important event in cell proliferation. Despite its importance, abnormalities of these genes and their protein products have yet to be found in lits asoociation with lung cancer. MATERIALS AND METHODS: The relationships between expression of cyclin A, cyclin B1, cyclin D1, cyclin D3, and cyclin E and clinicopathologic factors were investigated in 103 cases with non-small cell carcinomas, using immunohistochemical analysis. RESULTS: The positive immunoreactivity was observed in 51 cases (50%) for cyclin A, 33 cases (32%) for cyclin B1, 83 cases (81%) for cyclin D1, 19 cases (18%) for cyclin D3, and 11 cases (11%) for cyclin E. Expression of cyclin E was significant for lymph node metastasis (p=0.004, Chi-square test). There was no relationship between cyclin A, B1, D1, and E and histological typing, tumor size, lymph node metastasis, or pathological tumor, node and metastasis staging. CONCLUSION: These findings suggest that the expression of cyclin E played a role, to some degree, in the lymph node metastasis.

Mechanism of radiosensitization of 3-methyladenine in human hypopharynx cancer cells / 中华耳鼻咽喉头颈外科杂志

<p><b>OBJECTIVE</b>To investigate the radiosensitizing effect and its mechanism of 3-MA in human hypopharynx cancer cells.</p><p><b>METHODS</b>5 mmol/L of 3-MA combined with 2 Gy or 4 Gy of X-ray was utilized to deal with Fadu cells, and the cell livability (cloning efficiency) and DNA lesion severity (tail moment) of each groups was examined by clonogenic survival assay and comet assay, then differences were compared between groups by independent-sample T test. Fadu cells were then treated with different dose of 3-MA (1, 2, 5, 10 mmol/L), the alteration of cell cycle distribution was detected by flow cytometer, and differences among groups were analyzed through one-way analysis of variance. The expression of p62 and cyclinB1 in each group was examined by western blot.</p><p><b>RESULTS</b>The livability and DNA lesion severity of cells treated with 3-MA alone showed no notable variation. Compared with non-3-MA groups, the cloning efficiency of cells treated with 3-MA decreased much more after irradiated with 2 Gy or 4 Gy of X-ray (t = 13.41 or 13.98, P < 0.001), and the cells showed a more serious DNA lesion (t = 7.07 or 6.91, P < 0.001). The G2/M percentages of cells in the control group and groups treated with 1, 2, 5, 10 mmol/L of 3-MA were 17.10 ± 1.20, 23.30 ± 2.3, 39.90 ± 3.12, 58.47 ± 1.65, 76.13 ± 3.51 and differences among groups were statistically significant (F = 278.4, P < 0.05). The expression of p62 in cells treated with 3-MA showed a dose-dependent increase, while cyclinB1 showed a dose-dependent decrease.</p><p><b>CONCLUSIONS</b>The autophagy inhibitor 3-MA could enhance radiosensitivity of human hypopharynx cancer cells by inducing G2/M arrest and enhancing irradiation-induced DNA damage.</p>