Cells In G1 Phase Research Articles

A Novel Cyclic Pentadepsipeptide, N-Methylsansalvamide, Suppresses Angiogenic Responses and Exhibits Antitumor Efficacy against Bladder Cancer.

Simple SummaryWe found a novel cyclic pentadepsipeptide, N-methylsansalvamide (MSSV), and evaluated its anti-tumor action against bladder cancer using in vitro and in vivo model systems. Additionally, we report its anti-angiogenic responses both in vitro and in vivo. Moreover, acute toxicity test and tissue staining for liver function revealed that orally administered MSSV (2000 mg/kg for 14 days) exerted no harmful effects as it did not cause animal death, undesirable weigh alteration, adverse clinical symptoms, and abnormal biochemical marker levels (AST, ALT).Here, we explored the anti-tumor efficacy of a cyclic pentadepsipeptide, N-methylsansalvamide (MSSV), in bladder cancer. MSSV inhibited the proliferation of both bladder cancer 5637 and T24 cells, which was attributed to the G1-phase cell cycle arrest, apoptosis induction, and alteration of mitogen-activated protein kinases (MAPKs) and protein kinase b (AKT) signaling pathways. Additionally, the treatment of bladder cancer cells with MSSV suppressed migratory and invasive potential via the transcription factor-mediated expression of matrix metalloproteinase 9 (MMP-9). MSSV abrogated vascular endothelial growth factor (VEGF)-induced angiogenic responses in vitro and in vivo. Furthermore, our result showed the potent anti-tumor efficacy of MSSV in a xenograft mouse model implanted with bladder cancer 5637 cells. Finally, acute toxicity test data obtained from blood biochemical test and liver staining indicated that the oral administration of MSSV at 2000 mg/kg caused no adverse cytotoxic effects. Our preclinical data described the potent anti-angiogenic and anti-tumor efficacy of MSSV and showed no signs of acute toxicity, thereby suggesting the putative potential of oral MSSV as a novel anti-tumor agent in bladder cancer treatment.

Melatonin Inhibits Oxidized Low Density Lipoprotein Induced Proliferation and Migration of Vascular Smooth Muscle Cells and Its Mechanism

The effect and molecular mechanism of melatonin on the vascular smooth muscle cells proliferation and vascular smooth muscle cells migration induced by oxidized low density lipoprotein was explored in this paper. The first scheme: induce vascular smooth muscle cells with series of oxidized low density lipoprotein concentrations for 48 h (0 μg/mL, 10 μg/mL, 20 μg/mL, 50 μg/mL and 100 μg/mL). The second scheme: 50 μg/mL oxidized low density lipoprotein was used to induce vascular smooth muscle cells s for 0 h, 6 h, 12 h, 24 h and 48 h. The third option: first induce vascular smooth muscle cells with 50 μg/mL oxidized low density lipoprotein for 24 h, then treat vascular smooth muscle cells with series of melatonin concentrations for 48 h (1 nmol/L, 1 μmol/L, 0.1 mmol/L, 1 mmol/L). Cell Counting Kit-8 method was used to detect cell proliferation. Cells were randomly divided into blank control group, melatonin group (0.1 mmol/L melatonin treatment vascular smooth muscle cells 48 h), oxidized low density lipoprotein group (vascular smooth muscle cells were induced by 50 μg/ml oxidized low density lipoprotein for 24 h), oxidized low density lipoprotein +melatonin group (50 μg/ml of oxidized low density lipoprotein induced vascular smooth muscle cells for 24 h and then treated vascular smooth muscle cells s with 0.1 mmol/L of melatonin for 48 h). After relative treatment, cell cycle was detected by flow cytometry and cell migration was detected by scratch test. Proliferating cell nuclear antigen, alpha smooth muscle actin, cyclin A and extracellular regulatory protein kinase were detected by immunofluorescence staining and Western blot 1/2 (extracellular regulatory protein kinase 1/2) signal pathway related protein expression. Compared with the blank control group, vascular smooth muscle cells s treated with 50 μg/ml oxidized low density lipoprotein could significantly induce cell proliferation. After 24 h of induction by oxidized low density lipoprotein (50 μg/ml), the number of cells in S phase and G1 phase was decreased significantly and the number of cells in G2 phase was increased significantly. Cell migration was promoted and proliferating cell nuclear antigen, cyclin A, Cyclina, p-extracellular regulatory protein kinase 1/2 protein expression was up-regulated, down regulating the p-p21 protein level. The difference was statistically significant (p<0.01). The melatonin with series concentrations (1 nmol/L, 1 μmol/L, 0.1 mmol/L and 1 mmol/L) could inhibit the proliferation of vascular smooth muscle cells induced by oxidized low density lipoprotein to varying degrees. After vascular smooth muscle cells treatment with oxidized low density lipoprotein (50 μg/ml) for 24 h and melatonin (0.1 mmol/L) for 48 h, the cells in G1 and S phase increased significantly, the cells in G2 phase decreased significantly, the cell migration was inhibited, which were compared with oxidized low density lipoprotein oxidized low density lipoprotein group. The expression of proliferating cell nuclear antigen, cyclin A, Cyclina, p-extracellular regulatory protein kinase 1/2 protein was decreased and the expression of p-p21 protein was up-regulated, the difference was statistically significant (p<0.01). Melatonin can significantly inhibit the proliferation and migration of vascular smooth muscle cells induced by oxidized low density lipoprotein and its mechanism may be related to activating p21 and blocking extracellular regulatory protein kinase 1/2 signaling pathway.

Chrysophanol inhibits the osteoglycin/mTOR and activats NF2 signaling pathways to reduce viability and proliferation of malignant meningioma cells

ABSTRACT Chrysophanol shows promising antitumor activity, but how it may work against malignant meningioma is poorly understood. In addition, osteoglycin (OGN) may help mediate the antitumor effects of chrysophanol; thus, this study investigated the potential antitumor mechanism of chrysophanol in malignant meningioma cultures. Meningioma cell line HBL-52 were incubated with varying doses of chrysophanol (0–90 μM) for different time points, and osteoglycin (OGN) was overexpressed or inhibited in some cell cultures to assess its roles. Cell viability was quantified using the CCK8 assay and colony formation assays, while effects on cell cycle distribution and apoptotic rates were examined by flow cytometry and enzyme-linked immunosorbent assays (ELISA) to detect histone DNA levels. Caspase-3 and −9 activities were detected by related commercial kits. Protein expression was assessed using Western blotting. Chrysophanol significantly reduced HBL-52 cell viability, based on reduced colony formation, and proliferation, based on low levels of bromodeoxyuridine incorporation. Annexin V/propidium iodide staining revealed a 30% increase in apoptotic cells at 90 μM chrysophanol (33.7% vs 3.3% in control cultures). Chrysophanol treatment greatly decreased the Bcl-2/Bax expression ratio and increased the expressions of cleaved caspase-3 and −9, and the activities of caspase-3 and −9. Chrysophanol blocked cells in G1 phase and inhibited the OGN/mTOR signaling cascade, but activated neurofibromatosis 2 (NF2) cascade. OGN overexpression activated mTOR, down-regulated NF2, and partially reversed growth inhibition by chrysophanol. Chrysophanol may be useful as a treatment against malignant meningioma by inhibiting OGN/mTOR signaling and activating NF2 signaling.

HBx Downregulates miR-422a Expression via Activation of FOXG1/Q1/E1 in HepG2 Cells

microRNA-422a (miR-422a) is downregulated in many hematopoietic tumors and solid tumors including hepatocellular carcinoma. We previously demonstrated that hepatitis B virus X protein (HBx) downregulated expression of miR-422a in HCC cell line HepG2 in vitro. However, we explore the mechanisms underlying this action. Forkhead box proteins (FOX) G1/Q1/E1 are known to negatively regulate miR-422a expression, and this prompted us to determine whether HBx suppresses miR-422a expression via activation of FOXG1/Q1/E1. The relationship between FOXG1/Q1/E1 and miR-422a in HepG2 cells stably expressing HBx was assessed with qRT-PCR. The correlation between HBx and FOXG1/Q1/E1 was determined with qRT-PCR and western blot in vitro. The cell cycle and CCK-8 assays were used to elucidate the consequence of miR-422a transfection in HepG2-hbx cells. FOXG1/Q1/E1 activated by HBx was found to be responsible, at least in part, for the downregulation of miR-422a in HepG2 cells. miR-422a transfection hampered the growth of HepG2-hbx cells by arresting cells in G1 phase. Both FOXG1/Q1/E1 and miR-422a may be suitable molecular targets for treatment of HBV-infected HCC.

Assaying Cell Cycle Status Using Flow Cytometry.

In this chapter, four methods are described that can be used to assess cell cycle status in flow cytometry. The first method is based on the simultaneous analysis of cellular DNA content using a fluorescent DNA dye (propidium iodide) and of a nuclear proliferation marker (Ki-67). The second is based on the differential staining of DNA and RNA using Hoechst 33342 and Pyronin Y: this method is particularly useful to distinguish quiescent cells in G0 phase from G1 cells. Finally, two methods are described based on DNA incorporation of the synthetic nucleosides BrdU and EdU.

Aspergillus fumigatus-derived demethoxyfumitremorgin C inhibits proliferation of PC3 human prostate cancer cells through p53/p21-dependent G1 arrest and apoptosis induction

Human prostate cancer is the second most frequently diagnosed cancer worldwide, and its incidence rate continues to increase. Advanced prostate cancer is more difficult to treat than early forms due to its chemotherapy resistance. There is need for more effective agents that can inhibit the progression of advanced prostate cancer. Demethoxyfumitremorgin C (DMFTC) was isolated from the fermentation extract of the marine fungus Aspergillus fumigatus. Antiproliferative activity of DMFTC against human prostate cancer PC3 cells was examined through cell cycle analysis by flow cytometry, the fluorescent nuclear imaging analysis with propidium iodide (PI), and proteins expression related to cell cycle arrest and apoptosis were investigated via Western blotting. DMFTC inhibited PC3 cells growth through G1 phase cell cycle arrest and apoptosis induction. It activated the tumor suppressor p53 and the Cdk inhibitor p21, which regulate the cell progression into the G1 phase. Additionally, PI-positive late apoptotic non-viable cells were increased and the expression levels of the G1-positive downstream regulators cyclin D, cyclin E, Cdk2, and Cdk4 were decreased by DMFTC treatment. These results suggest that DMFTC induces G1 arrest and apoptosis induction through regulation of p53/p21-dependent cyclin-Cdk complexes, and it may be a useful therapeutic agent for the treatment of human advanced prostate cancer.

Inhibition of PI3K/AKT signaling via ROS regulation is involved in Rhein-induced apoptosis and enhancement of oxaliplatin sensitivity in pancreatic cancer cells.

Several natural products have been demonstrated to both enhance the anti-tumor efficacy and alleviate the side effects of conventional chemotherapy drugs. Rhein, a main constituent of the Chinese herb rhubarb, has been shown to induce apoptosis in various cancer types. However, the exact pharmacological mechanisms controlling the influence of Rhein on chemotherapy drug effects in pancreatic cancer (PC) remain largely undefined. In this study, we found that Rhein inhibited the growth and proliferation of PC cells through G1 phase cell cycle arrest. Moreover, Rhein induced caspase-dependent mitochondrial apoptosis of PC cells through inactivation of the PI3K/AKT pathway. Combination treatment of Rhein and oxaliplatin synergistically enhanced apoptosis of PC cells through increased generation of intracellular reactive oxygen species (ROS) and inactivation of the PI3K/AKT pathway. Pre-treatment with the ROS scavenger N-acetyl-L-cysteine attenuated the combined treatment-induced apoptosis and restored the level of phosphorylated AKT, indicating that ROS is an upstream regulator of the PI3K/AKT pathway. The combination therapy also exhibited stronger anti-tumor effects compared with single drug treatments in vivo. Taken together, these data demonstrate that Rhein can induce apoptosis and enhance the oxaliplatin sensitivity of PC cells, suggesting that Rhein may be an effective strategy to overcome drug resistance in the chemotherapeutic treatment of PC.

Acetone Extract of &amp;lt;i&amp;gt;Dioscorea alata&amp;lt;/i&amp;gt; Inhibits Cell Proliferation in Cancer Cells

Dioscorea spp., White Yam has been shown to exhibit a wide range of nutritional and medicinal properties. However, the compounds associated with its medicinal functions have not been fully examined. The purpose of this study was to generate a chemoinformatic profile of the bioactive compounds present in Dioscorea. alata (D. alata) and to characterize their putative anti-cancer properties using prostate (DU145) and lung (A549) cancer cells. Chemoinformatic profiling of D. alata resulted in five bioactive extracts: hexane (DaJa-1), ether (DaJa-2), acetone (DaJa-3), ethanol (DaJa-4) and water (DaJa-5) were generated. The analytes present in the five bioactive extracts were dissolved in 0.1% DMSO and their anti-cancer activity were determined. We observed that the acetone extract (DaJa3) was the only extract capable of inducing greater than 90% cell death of DU 145 cell at 100 μg/mL. The order of growth inhibition of the extracts in DU-145 cell is DaJa-3 (IC50, 31.45 μg/mL) > DaJa-4 (IC50 60 μg/mL) > DaJa-1 (IC50 > 100 μg/mL) ≥ DaJa-2 (IC50 > 100 μg/mL) ≥ DaJa-5 (IC50 > 100 μg/mL). MTT cell viability, dye exclusion, caspase activity and microscopic assessment of apoptotic cells demonstrated that DaJa-3 displayed cytotoxicity to both lung and prostate cancer cells. The A549 cells were more sensitive toward DaJa-3 with an IC50 value of 22.28 μg/mL (CI 28.42 to 36.63 μg/mL), compared to that of DU145 cells with an IC50 value of 31.45 μg/mL (CI 27.58 to 35.86 μg/mL). It was also observed that DaJa-3 induces differential anti-proliferative activity in the cancer cells. The apoptotic response induced by DaJa-3 paralleled the level of cell cytotoxicity observed in both cell lines. DaJa-3 induces G2 phase cell cycle arrest in DU145 cells but G1 arrest in A549 cells. The level of key apoptotic regulator proteins was upregulated, suggested that DaJa-3 may be mediating its anti-cancer effect through activation of the intrinsic apoptotic pathway. Altogether, our data indicates that DaJa-3 derived from a staple food source (white Yam) contains unique active compounds that have specific biological properties that may prevent certain types of cancer or specific types of cancer.

Effects of nanopatterned-surface dishes on chondrocyte growth and cell cycle progression.

Discovering and developing ideal cell culture methods is important for cell biology, drug development, and cell therapy. Recent studies have explored and demonstrated the use of nanoscale structures and patterns that influence cell behavior, such as 3D scaffolds. In this study, we analyzed the effects of nanopatterned-surface dishes using chondrocytes as model cells. Chondrocytes grown on nanopatterned dishes exhibited rounded shapes. Interestingly, chondrocytes have a lower COL10 mRNA level when cultured using nanopatterned dishes. The nanopatterned dishes induced G0-/G1-phase cell cycle arrest and reduced the rate of proliferation. Our results suggest that nanoscale structures can directly control cellular behaviors and can be used for chondrocyte cell culture without causing chondrocytes to lose their functions. These results help to elucidate cellular responses and behaviors in native-like environments, and this information can be used to improve human health.

Curcumin analog B14 has high bioavailability and enhances the effect of anti-breast cancer cells in vitro and in vivo.

Curcumin has a variety of anticancer properties, but low bioavailability prevents its use in chemotherapeutic applications. To address this problem, we tested the efficacy of the synthetic curcumin analog B14 in breast cancer cells and explored the mechanism by which B14 inhibits proliferation and metastasis of breast cancer cells. We used the breast cancer cell line MCF‐7, MDA‐MB‐231 to study the anticancer effects of B14 and assessed cell viability, cell migration and invasion, cell cycle, and apoptosis, in addition, the antitumor effect of B14 in vivo was examined in mice bearing MDA‐MB‐231 cells. We found that, as the concentration of B14 increased, cell viability decreased in a dose‐dependent manner. Compound B14 exerted the best antitumor activity and selectivity for MCF‐7 and MDA‐M‐231 cells (IC50 = 8.84 μmol/L and 8.33 μmol/L, respectively), while its IC50 value for MCF‐10A breast epithelial cells was 34.96 μmol/L. B14 has been shown to be a multi‐targeted drug that alters the expression of cyclin D1, cyclin E1, and cyclin‐dependent kinase 2 (CDK2), and ultimately induces G1 phase cell cycle arrest. At the same time, B14 activates the mitochondrial apoptosis pathway in breast cancer cells. Furthermore, B14 was more effective than curcumin in inhibiting cell migration, invasion, and colony formation. In tumor‐bearing mice, analog B14 significantly reduced tumor growth and inhibited cell proliferation and angiogenesis. The pharmacokinetic test found that B14 was more stable than curcumin in vivo. Our data reveal the therapeutic potential of the curcumin analog B14 and the underlying mechanisms to fight breast cancer cells.

Alcohol dehydrogenase 1 and NAD(H)-linked methylglyoxal oxidoreductase reciprocally regulate glutathione-dependent enzyme activities in Candida albicans.

Glutathione reductase (Glr1) activity controls cellular glutathione and reactive oxygen species (ROS). We previously demonstrated two predominant methylglyoxal scavengers-NAD(H)-linked methylglyoxal oxidoreductase (Mgd1) and alcohol dehydrogenase 1 (Adh1)-in glutathione-depleted γ-glutamyl cysteinyl synthetase-disrupted Candida albicans. However, experimental evidence for Candida pathophysiology lacking the enzyme activities of Mgd1 and Adh1 on glutathione-dependent redox regulation remains unclear. Herein, we have aimed to demonstrate that glutathione-dependent enzyme activities coupled with cellular ROS changes is regulated by methylglyoxal accumulation in Δmgd1/Δadh1 double disruptants. Δmgd1/Δadh1 showed severe growth defects and G1-phase cell cycle arrest. The observed complementary and reciprocal methylglyoxal-oxidizing and methylglyoxalreducing activities between Δmgd1 and Δadh1 were not always exhibited in Δmgd1/Δadh1. Although intracellular accumulation of methylglyoxal and pyruvate was shown in all disruptants, to a greater or lesser degree, methylglyoxal was particularly accumulated in the Δmgd1/Δadh1 double disruptant. While cellular ROS significantly increased in Δmgd1 and Δadh1 as compared to the wild-type, Δmgd1/Δadh1 underwent a decrease in ROS in contrast to Δadh1. Despite the experimental findings underlining the importance of the undergoing unbalanced redox state of Δmgd1/Δadh1, glutathione-independent antioxidative enzyme activities did not change during proliferation and filamentation. Contrary to the significantly lowered glutathione content and Glr1 enzyme activity, the activity staining-based glutathione peroxidase activities concomitantly increased in this mutant. Additionally, the enhanced GLR1 transcript supported our results in Δmgd1/Δadh1, indicating that deficiencies of both Adh1 and Mgd1 activities stimulate specific glutathione-dependent enzyme activities. This suggests that glutathione-dependent redox regulation is evidently linked to C. albicans pathogenicity under the control of methylglyoxal-scavenging activities.

Inhibitory role of oleanolic acid and esculetin in HeLa cells involve multiple signaling pathways.

The global burden of cervical cancer from low and middle-income groups is increasing at alarming rates with more than half a million women being diagnosed every year. Although the disease is largely preventable when screened and diagnosed in earlier stages, the development of resistance and relapse had resulted in a poor prognosis. Therefore, a comprehensive approach needs to be put forward to understand and develop new preventive and therapeutic strategies to effectively combat cancer. Recently, much attention has been diverted to plant-derivatives for the treatment as they exhibit potent anti-cancer properties and side-effects caused by chemotherapeutic agents can also be prevented. Oleanolic acid and Esculetin are natural compounds known for their anti-cancer properties. Hence, the present study investigates the effect and mechanism of these compounds on cervical carcinoma, using HeLa cells. Posttreatment, it was observed that these compounds inhibited proliferation by both arresting the cells in the sub G1 phase and inducing senescence. Also, a marked reduction in the migration and cell survival was observed, as evidenced by results obtained from wound healing assay and Annexin V-FITC/PI staining. Furthermore, studies on the expression pattern of genes involved in major signaling pathways demonstrated a profound effect of these compounds. Taken together, the results of our study suggest that both Oleanolic acid and esculetin serve as a plausible therapeutic agent.

Phenolic Compounds from Polygonum chinense Induce Growth Inhibition and Apoptosis of Cervical Cancer SiHa Cells

Cervical cancer is considered to be one of the most serious malignant tumors in women. Natural compounds have been considered as important sources in the search for new anticancer agents. Polygonum chinense (PC) has been used as herbal medicine and Chinese cool tea. By activity-guided of the extracts from PC, PCwater shows good growth inhibition on SiHa cell, then by chromatographic analysis (HPLC and HPLC-MS/MS), we found twelve components, seven were phenolic compounds (PHE), two PHE named ellagic acid and corilagin were found to show strong growth inhibition effects in SiHa cell dose-dependently, while the seven phenolic compounds showed low inhibition on the common human HcerEpic cell. Further research found ellagic acid and corilagin induced G2 phase cell cycle arrest by upregulating levels of P53, Bcl-2, caspase 3, and caspase 9, while the Bax was reduced. These results suggested that PHE from PC might have potential anticancer effects against SiHa cells by acting through the apoptosis pathway, PHE from PC might have the potential to be used as a nutraceutical for the prevention and treatment of ovarian cancer.

Inhibition of the P53/P21 Pathway Attenuates the Effects of Senescent Nucleus Pulposus Cell-Derived Exosomes on the Senescence of Nucleus Pulposus Cells.

ObjectiveThe purpose of this paper is to investigate the effects of senescent nucleus pulposus cell (NPC)‐derived exosomes (SNPC‐Exo) and the roles of the P53/P21 pathway on the senescence of NPC.MethodsThe senescent phenotypes of NPC were induced by interleukin‐1β treatment. SNPC‐Exo was extracted from the culture medium of senescent NPC and purified by differential centrifugation. The structure of SNPC‐Exo was identified by transmission electron microscopy and western blot analysis was used to determine the exosomal marker proteins CD63 and Tsg101. Western blot analysis was performed to determine the relative expression levels of P16, P21, and P53 in NPC. Senescence‐associated β‐galactosidase (SA‐β‐gal) staining was used to stain the senescent NPC and a phase contrast microscope was used to observe and count the SA‐β‐gal staining of NPC. The proliferation of SNPC‐Exo‐treated NPC was assessed using growth curve analysis and the colony formation assay. The cell cycle of SNPC‐Exo‐treated NPC was determined by flow cytometry. NPC were transfected with siRNA to knock down P53 and P21 expression.ResultsInterleukin‐1β‐treated NPC had a higher percentage of SA‐β‐gal positive cells (45%) than the control group (20%) and showed an increase in the relative expression of P16, P21, and P53 (P < 0.05). SNPC‐Exo were positive for exosomal marker protein CD63 and Tsg 101 and negative for calnexin, and successfully internalized as previously described. SNPC‐Exo‐treated NPC showed an increase in the relative expression of P21 and P53 (P < 0.05). Compared with the control group, the SNPC‐Exo‐treated NPC showed a lower growth rate (3 times lower on the 5th day and 2 times lower on the 7th day), fewer colony‐forming units (12.0%), and a higher percentage of SA‐β‐gal‐positive NPC (50.0%). The SNPC‐Exo‐treated NPC contained more G1 phase cells (68.0%) and fewer S phase (15.5%) cells than the control group (53.0% in G1 phase, 33.5% in S phase). The expression of P21 and P53 significantly decreased in SNPC‐exo‐treated NPC after siRNA transfection (P < 0.05), followed by a higher growth rate (2 times higher on the 5th day and 1.5 times higher on the 7th day) and lower percentage of SA‐β‐gal‐positive NPC (22.5%). Moreover, the inhibition of the P53/P21 pathway promoted the SNPC‐Exo‐treated NPC to enter the S phase (from 15.5% to 25.3%).ConclusionThe inhibition of the P53/P21 pathway attenuated the senescence of NPC induced by SNPC‐Exo.

In silico APC/C substrate discovery reveals cell cycle-dependent degradation of UHRF1 and other chromatin regulators.

The anaphase-promoting complex/cyclosome (APC/C) is an E3 ubiquitin ligase and critical regulator of cell cycle progression. Despite its vital role, it has remained challenging to globally map APC/C substrates. By combining orthogonal features of known substrates, we predicted APC/C substrates in silico. This analysis identified many known substrates and suggested numerous candidates. Unexpectedly, chromatin regulatory proteins are enriched among putative substrates, and we show experimentally that several chromatin proteins bind APC/C, oscillate during the cell cycle, and are degraded following APC/C activation, consistent with being direct APC/C substrates. Additional analysis revealed detailed mechanisms of ubiquitylation for UHRF1, a key chromatin regulator involved in histone ubiquitylation and DNA methylation maintenance. Disrupting UHRF1 degradation at mitotic exit accelerates G1-phase cell cycle progression and perturbs global DNA methylation patterning in the genome. We conclude that APC/C coordinates crosstalk between cell cycle and chromatin regulatory proteins. This has potential consequences in normal cell physiology, where the chromatin environment changes depending on proliferative state, as well as in disease.

Expression and Importance of TMED2 in Multiple Myeloma Cells.

ObjectiveTMED2 is a member of the transmembrane emp24 domain (Tmed)/p24 protein family, which is significantly upregulated in breast cancer, ovarian cancer and other tumour tissues. The purpose of this study was to investigate the expression of TMED2 in MM cell lines and its effect on the biological behaviour of MM cell lines.MethodsReal-time quantitative PCR (RT-qPCR) was used to detect the expression of TMED2 in MM cell lines, including MM.1S and RPMI 8226 cells, and lentivirus vector-mediated TMED2 gene silencing was used to further study the effect of the downregulation of TMED2 expression on cell viability, the cell cycle, and apoptosis.ResultsBased on the RT-qPCR results, the expression of the TMED2 mRNA was increased in the MM cell lines MM.1S and RPMI 8226 compared with endogenous control GAPDH. The expression of the TMED2 mRNA was substantially reduced after transfection of the shRNA targeting TMED2 (shTMED2) in both MM cell lines. The CCK-8 assay showed significant decreases in the viability of MM.1S and RPMI 8226 cells, suggesting that the TMED2 gene plays an important role in the proliferation of these two cell lines. The cell cycle of MM.1S and RPMI 8226 cells was substantially altered by shTMED2, as evidenced by the increased number of cells in G1 phase and decreased number of cells in S and G2/M phases. The FACS analysis revealed a significant increase in the apoptosis of MM.1S and RPMI 8226 cells due to the increased activity of Caspase 3/7, suggesting that the TMED2 gene is significantly related to the apoptosis of these two cell lines.ConclusionBased on these results, TMED2 may play an important role in the pathogenesis of MM. This novel study may contribute to further investigations of useful biomarkers and potential therapeutic targets in patients with MM.

Studies on the Inhibitory Effects of Nano-Hydroxyapatite-Loaded As₂O₃ on Hepatoma Cells.

A stable monodisperse hydroxyapatite (HAP) nanoparticle suspension was prepared by chemical method-assisted ultrasound irradiation. HAP nanoparticles were characterized by atomic force microscopy (AFM) and particle size potentiometry. The effects of HAP nanoparticles on BEL-7402 human hepatocarcinoma cells were studied by MTT colorimetric assay and morphological observation. The mechanism of HAP nanoparticles was studied by analyzing single cell fluorescence element microregion, the change of ultrastructure and cell cycle. The experimental results show that HAP nanoparticles have an obvious inhibitory effect on BEL-7402 human hepatocarcinoma cells in vitro. By entering the cancer cells and blocking the progress of cell cycle, HAP nanoparticles induce the accumulation of cells in G1 phase, which leads to cancer cell swelling and apoptosis.

LncRNA MIR205HG Drives Esophageal Squamous Cell Carcinoma Progression by Regulating miR-214/SOX4 Axis.

BackgroundEsophageal squamous cell carcinoma (ESCC) is a common and fatal malignancy, which has posed a great challenge to public health, especially in China. Dysregulation of long non-coding RNAs is involved in the occurrence, development, invasion, and metastasis of multiple cancers including ESCC. However, little is known about the function of MIR205HG in ESCC.MethodsWe used qRT-PCR to detect the expression level of MIR205HG, miR-214, and SOX4 in human ESCC tissues and cell lines. Loss-of-functional assays were performed to test the impact of MIR205HG on cell proliferation, metastasis, and apoptosis process via CCK-8, transwell, and flow cell cytometry assays. Additionally, the downstream molecular mechanism of MIR205HG in ESCC was explored.ResultsHere, we found MIR205HG was substantially up-regulated in ESCC, and there was a positive correlation between MIR205HG expression and tumor size and lymphatic metastasis of ESCC patients. Inhibition of MIR205HG attenuated cell proliferation, migration, and invasion. Silencing MIR205HG increased G1 phase cell counts and decreased S phase cell counts, along with increased apoptotic cell populations. Notably, the rescue assays indicated that miR-214 could partly reverse the influence of MIR205HG on ESCC cell migration. We also found that SOX4 was a direct target mRNA of miR-214, and MIR205HG could act as a molecular sponge to regulate SOX4 expression in ESCC.ConclusionTaken together, our findings demonstrate that MIR205HG promotes ESCC progression by regulating the miR-214/SOX4 axis. MIR205HG may be a novel candidate target for ESCC diagnosis and therapy.

Effect of cell cycle synchronization on cadmium-induced apoptosis and necrosis in NRK-52E cells

ABSTRACT Heavy metal pollution is a problem that cannot be ignored. Due to the prevalence of cadmium in the environment and its harmful effects on humans, cadmium pollution has become a research hotspot recently. The mechanism of cadmium-induced toxicity has also drawn much attention and most studies have been conducted using whole cells, but the toxicological mechanism of cadmium remains unclear. In this study, we aimed to obtain NRK-52E cells at different growth stages by various methods and analyze the differences in cadmium toxicity. The results show that the cadmium sensitivity of cells in each phase was different and the late apoptotic rate was increased significantly after 5 µM Cd treatment. In addition, cadmium easily induces apoptosis of G0- and S-phase cells, as well as necrosis of S- and M-phase cells, but has no significant effect on G1-phase cells. Overall, we first explored the differences in the effects of cadmium on NRK-52E cells at various growth phases. Besides, the findings of this study might provide a theoretical basis for further exploration of the toxicological mechanism of cadmium.Abbreviations Cd: cadmium; CDK: cyclin-dependent kinases; DAPI 2-(4-amidinophenyl)-1H-indole-6-carboxamidine; TBST: Tris-buffered saline with Tween-20; PI: propidium iodide; DMEM: Dulbecco’s Modified Eagle Medium; BCA: bicinchoninic acid

Quinolinone inhibits proliferation of gastric cancer cells and induces their apoptosis via down-regulation of the expression of pro-oncogene c-Myc

Purpose: To determine the anti-proliferative potential of quinolinone against gastric cancer cells, and the underlying mechanism of action.Methods: Quinolinone-mediated proliferative changes were measured using 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay, while its effect on apoptosis was determined by flow cytometry. Transwell and wound healing assays were used for the determination of the effect of quinolinone on cell invasion and migration. The effect of quinolinone on protein expression levels were assayed with western blotting.Results: Quinolinone caused reduction in gastric cancer cell viability, but it had no effect on normal (GES-1) cells. Treatment with 8 μM quinolinone reduced the viability of SNU-5 and SGC-7901 cells to 32 and 27 %, respectively. Moreover, 8 μM quinolinone induced 67.90 and 71.54 % apoptosis in SNU-5 and SGC-7901 cells, respectively. Quinolinone significantly increased the population of cells in G1 phase, and suppressed migration potential (p &lt; 0.05). Furthermore, in quinolinone-treated cells, the expression levels of p-PI3K, c-Myc and p-AKT were much lower than those in untreated cells (p &lt; 0.05). Quinolinone also downregulated the expressions of MMP-2 and MMP-9, while it upregulated p21 expression in SNU-5 and SGC-7901 cells.Conclusion: Quinolinone suppresses the growth of SNU-5 and SGC-7901 gastric cancer cells via cell cycle arrest, induction of apoptosis and downregulation of the expressions of c-Myc and metalloproteinases. Thus, quinolinone may be developed as a potential drug candidate for the treatment of gastric cancer.&#x0D; Keywords: Gastric cancer, Apoptosis, Metalloproteinases, Phosphorylation