Culture supernatants of breast cancer cell line MDA-MB-231 treated with parthenolide inhibit the proliferation, migration, and lumen formation capacity of human umbilical vein endothelial cells

Background  Parthenolide has been tested for anti-tumor activities, such as anti-proliferation and pro-apoptosis in recent studies. However, little is known about its role in the process of tumor angiogenesis. This study aims to investigate the effects and potential mechanisms of parthenolide on the proliferation, migration and lumen formation capacity of human umbilical vein endothelial cells.

Methods  Different concentrations of parthenolide were applied to the human breast cancer cell line MDA-MB-231 cells. After 24-hour incubation, the culture supernatants were harvested and used to treat human umbilical vein endothelial cells for 24 hours. Then an inverted fluorescence phase contrast microscope was used to evaluate the human umbilical vein endothelial cells. The secretion of vascular endothelial growth factor (VEGF), interleukin (IL)-8 and matrix metalloproteinases (MMP)-9 in the culture supernatant of the MDA-MB-231 cells was then measured with enzyme-linked immunosorbent assay (ELISA) assays.

Results  Suppression of proliferation, migration, and the lumen formation capacity of human umbilical vein endothelial cells was observed in the presence of the culture supernatants from the breast cancer cell line treated with different concentrations of parthenolide. Parthenolide decreased the levels of the angiogenic factors MMP-9, VEGF, and IL-8 secreted by the MDA-MB-231 cells.

Conclusions  Parthenolide may suppress angiogenesis through decreasing angiogenic factors secreted by breast cancer cells to interfere with the proliferation, migration and lumen-like structure formation of endothelial cells, thereby inhibiting tumor growth. It is a promising potential anti-angiogenic drug.

     

Cytotoxic effects of mono-（2-ethylhexyl） phthalate on human embryonic stem cells

Background  Mono-(2-ethylhexyl) phthalate (MEHP), the metabolite of di-(2-ethylhexyl) phthalate (DEHP), was suspected to be toxic to human embryos. This study contributes to investigating its toxic effects by an embryonic stem cell test (EST) based on two human embryonic stem cell (hESCs) lines.

Methods  CH1 established in our own lab and H1, a federally registered cell line were two human embryonic stem cell lines used in this test. Four endpoint measurements were performed consisting of cell viability, proliferation ability, apoptosis as well as changes of gene expression patterns after spontaneous differentiation were determined. For measuring effects on the first three endpoints, the cells were treated with various concentrations of MEHP dissolved in dimethyl sulfoxide (DMSO) and only with DMSO which served as control and harvested after 5 days. For measuring effects during spontaneous differentiation, the RNA of embryoid bodies (EBs) formed after 8 days’ MEHP exposure was collected and changes in differentiation specific gene expression patterns were analyzed by quantitative real time RT-PCR.

Results  As a result the viability and proliferation ability of both cell lines decreased significantly at 1000 µmol/L MEHP, while there was no effect on apoptosis or cell morphology. In addition MEHP also changed the gene expression pattern in the EBs of both cell lines.

Conclusion  MEHP in a high dose was cytotoxic and affected the development of hESCs, which indicates its embryo toxicity in human embryos.

     

Role of microRNA-181a in the apoptosis of tubular epithelial cell induced by cisplatin 

Background  Cisplatin (DDP) is one of most effective and most commonly used therapeutic agent in treating tumors, it can accumulate in the kidney and lead to acute renal failure. MicroRNA-181a can induce cell apoptosis by suppressing the expression of Bcl-2 family. In the present study, we investigated the role of microRNA-181a in the apoptosis of tubular epithelial cell induced by DDP.

Methods  HK-2 cells were cultured, transfected with microRNA-181a inhibitor for 48 hours, and stimulated with 50 µmol/L cisplatin for 24 hours. MicroRNA-181a expression was analyzed by real time PCR, and cell apoptosis was detected by flow cytometry. Moreover, Bcl-2 and Bcl-2-associated X protein (Bax) expression were measured by Western blotting.

Results  MicroRNA-181a expression significantly down-regulated in cells transfected with microRNA-181a inhibitor, compared with that in untransfectd cells (21.19±2.01 vs. 38.87±1.97, P <0.05). Cell apoptosis induced by DDP significantly decreased in cells transfected with MicroRNA-181a inhibitor. Compared with DDP treated cells alone, Bcl-2 expression strikingly was up-regulated and Bax expression was down-regulated in cells transfected with microRNA-181a inhibitor.

Conclusion  One pathway of DDP induces apoptosis of tubular epithelial cell by suppressing Bcl-2 expression is achieved by regulating the target gene of MicroRNA-181a.

     

Sodium valproate induces mitochondria-dependent apoptosis in human hepatoblastoma cells

Background  Sodium valproate inhibits proliferation in neuroblastoma and glioma cells, and inhibits proliferation and induces apoptosis in hepatoblastoma cells. Information describing the molecular pathways of the antitumor effects of sodium valproate is limited; therefore, we explored the mechanisms of action of sodium valproate in the human hepatoblastoma cell line, HepG2.

Methods  The effects of sodium valproate on the proliferation of HepG2 cells were evaluated by the Walsh-schema transform and colony formation assays. Sodium valproate-induced apoptosis in HepG2 cells was investigated with fluorescence microscopy to detect morphological changes; by flow cytometry to calculate DNA ploidy and apoptotic cell percentages; with Western blotting analyses to determine c-Jun N-terminal kinases (JNK), p-JNK, Bcl-2, Bax, and caspase-3 and -9 protein expression levels; and using JC-1 fluorescence microscopy to detect the membrane potential of mitochondria. Statistical analyses were performed using one-way analysis of variance by SPSS 13.0 software.

Results  Our results indicated that sodium valproate treatment inhibited the proliferation of HepG2 cells in a dose-dependent manner. Sodium valproate induced apoptosis in HepG2 cells as it: caused morphologic changes associated with apoptosis, including condensed and fragmented chromatin; increased the percentage of hypodiploid cells in a dose-dependent manner; increased the percentage of annexin V-positive/propidium iodide-negative cells from 9.52% to 74.87%; decreased JNK and increased phosphate-JNK protein expression levels; reduced the membrane potential of mitochondria; decreased the ratio of Bcl-2/Bax; and activated caspases-3 and -9.

Conclusion  Sodium valproate inhibited the proliferation of HepG2 cells, triggered mitochondria-dependent HepG2 cell apoptosis and activated JNK.

     

Induction of actin disruption and downregulation of P-glycoprotein expression by solamargine in multidrug-resistant K562/A02 cells

Background  Solamargine (SM), a steroidal glycoalkaloid isolated from the Chinese herb Solanum incanum, has been shown to inhibit the growth of some cancer cell lines and induce significant apoptosis. However, the effects of SM on multidrug-resistant (MDR) cells and the molecular mechanisms involved are poorly understood. The purpose of this study was to evaluate the anti-MDR effects of SM and the associated mechanisms in MDR K562/A02 cells.

Methods  The cytotoxicity of SM was measured by 3-(4,5-dimethylthiazol)-2,5-diphenyltetrazolium bromide (MTT) assay. The 14′,6-diamidino-2-phenylindole (DAPI) nuclear staining and flow cytometry were used to detect SM-induced apoptosis. The mRNA expression of P-glycoprotein (P-gp) was investigated by real-time PCR (RT-PCR). Western blotting was used to determine the expression of Bcl-2, Bax, and actin. The changes in the morphology of actin were examined with immunofluorescence staining.

Results  MTT results showed that SM effectively killed the MDR sublines K562/A02, KB/VCR, and H460/paclitaxel (Taxol), and their parental cell lines K562, KB, and H460 to an equivalent or more sensitive degree. Based on the results by flow cytometry and immunostaining, the pro-apoptotic effects of SM were observed in MDR K562/A02 cells. Furthermore, the RT-PCR results showed that SM induced the downregulation of MDR1 mRNA. In addition, the expression of P-gp and actin was decreased in the SM-treated cells, as measured by western blotting and immunostaining.

Conclusions  These results demonstrate that SM effectively triggers apoptosis in MDR tumor cells, which is associated with actin disruption and downregulation of MDR1 expression. This compound may merit further investigation as a potential therapeutic agent that bypasses the MDR mechanism for the treatment of MDR tumors. 

     

Rapamycin and 3-methyladenine regulate apoptosis and autophagy in bone-derived endothelial progenitor cells

 

Background  Mammalian target of rapamycin (mTOR) is involved in a caspase independent form of programmed cell death called autophagy. The aim of this research was to investigate the effects of rapamycin and 3-methyladenine (3-MA) on autophagy, proliferation, apoptosis, and cell-cycle parameters of rat bone marrow-derived endothelial progenitor cells (EPCs).

Methods  Mononuclear cells isolated from rat bone marrow were treated with rapamycin (0.01, 0.1, 1, or 10 μg/L) or 3-MA (1.25, 2.5, 5, or 10 mmol/L) for 24 hours. Expression of the autophagy marker protein LC3-II was analyzed by Western blotting. Apoptosis and cell-cycle progression were analyzed by flow cytometry. Cell proliferation was measured using the MTT assay.

Results  Rapamycin treatment of EPCs induced apoptosis and autophagy and inhibited proliferation and cell-cycle progression in a dose-dependent manner. Treatment with 5 mmol/L 3-MA promoted cell proliferation; in contrast, treatment with 10 mmol/L 3-MA promoted apoptosis and induced S-phase arrest.

Conclusions  Rapamycin treatment of EPCs induced apoptosis and autophagy. Low concentrations of 3-MA had no significant effect on the proliferation and apoptosis of EPCs; The 5 mmol/L group promoted cell proliferation, but had no effect on the apoptosis; the 10 mmol/L group inhibited the proliferation and promoted apoptosis through the cell cycle.

     

Resveratrol Induces Apoptosis in Pancreatic cancer cells

Background  Pancreatic cancer is one of the most lethal human cancers with a very low survival rate of 5 years. Conventional cancer treatments including surgery, radiation, chemotherapy or combinations of these show little effect on this disease. Several proteins have been proved critical to the development and the progression of pancreatic cancer. The aim of this study was to investigate the effect of resveratrol on apoptosis in pancreatic cancer cells.

Methods  Several pancreatic cancer cell lines were screened by resveratrol, and its toxicity was tested by normal pancreatic cells. Western blotting was then performed to analyze the molecular mechanism of resveratrol induced apoptosis of pancreatic cancer cell lines.

Results  In the screened pancreatic cancer cell lines, capan-2 and colo357 showed high sensitivity to resveratrol induced apoptosis. Resveratrol exhibited insignificant toxicity to normal pancreatic cells. In resveratrol sensitive cells, capan-2 and colo357, the activation of caspase-3 was detected and showed significant caspase-3 activation upon resveratrol treatment; p53 and p21 were also detected up-regulated upon resveratrol treatment.

Conclusion  Resveratrol provides a promising anti-tumor stratagy to fight against pancreatic cancer.

     

Specific regulator of eosinophil apoptosis: Siglec-8—new hope for bronchial asthma treatment

Objective  It is known that Siglec-8 is selectively expressed on human eosinophils at a high level and mediates eosinophil apoptosis when crosslinked with its antibody. The aim of our review is to elucidate the molecular and biological characteristic of Siglec-8 and then discuss the function and possible mechanisms of Siglec-8 in eosinophils. Thereby, we will expand our understanding to the regulation of eosinophil apoptosis, and provide important clues to the treatment of asthma and other hyper-eosinophilic diseases.

Data sources  Most articles were identified by searching of PubMed online resources using the key term Siglecs.

Study selection  Mainly original milestone articles and critical reviews written by major pioneer investigators in the field were selected.

Results  Siglec-8 is selectively expressed on human eosinophil and can specifically induce eosinophil apoptosis.

Conclusion  The restricted expression of Siglec-8 on human eosinophil and the rapid progress in understanding its role as cell signaling and activation of death receptors have made it an attractive target for treatment of asthma and other hyper-eosinophilic diseases.

     

Effect of pcDNA3.1- vascular endothelial growth factor 165 recombined vector on vascular buds in rabbit vertebral cartilage endplate

Background  This study aimed to investigate the effect of pcDNA3.1-vascular endothelial growth factor (VEGF)165 vector on vertebral cartilage endplate vascular buds and intervertebral discs.

Methods  Rabbits were randomly assigned to the control and experimental groups with 10 in each. In the experimental group, we anesthetized the rabbits and exposed the front vertebral body. Using the mark of the longitudinal ossature of the front vertebral body of the lumbar vertebrae, we advanced a needle at the central point of the front fourth and fifth lumbar intervertebral discs and injected 20 µl pcDNA3.1-VEGF165. Similarly, in the control group, we injected 20 µl pcDNA3.1. At 4 and 8 weeks post-injection, we examined the changes of the vertebral cartilage endplate using X-ray radiograph, histology, and scanning electron microscopy.

Results  The vertebral cartilage endplate calcification and degeneration in the experimental group were less than those in the control group at 8 weeks post-operation. The average number and diameter of vascular buds obviously increased in the experimental group at 4 and 8 weeks post-operation. The number of vascular buds and the diameter in the region of the inner annulus increased when compared to those in the area near the nucleus pulposus.

Conclusions  The pcDNA3.1-VEGF165 plasmid can increase the average number and diameter of vascular buds and decelerate intervertebral disc degeneration.

     

Proteomic analysis for detecting serum biomarkers related to smoking in humans

Background  Smoking is the leading cause of death in the world. This study focused on the difference of the serum proteomic profiling between healthy smokers and nonsmokers in order to find smoking-specific serum biomarkers.

Methods  Pattern-based proteomic profiling of 100 serum samples (from 50 Chinese male smokers and 50 matched nonsmokers) was performed through magnetic bead fractionation coupled with matrix-assisted laser desorption/ionization time-of-flight mass spectrometry analysis (MALDI-TOF-MS) and resulting data were statistically analyzed by Ciphergen ProteinChip software 3.0.2.

Results  We found 72 serum peaks were significantly different between smokers and nonsmokers (P <0.05). Marker peaks of mass-to-charge ratio (m/z) 3159.13, 7561.03 and 9407.32 were smoking-specific.

Conclusion  The preliminary data suggested that smoking-specific serum biomarkers could be detected in humans.

     

Antiviral therapy of decompensated hepatitis B virus-related cirrhosis

Objective  To review the development, mechanism, necessity and limitation of antiviral therapy in decompensated hepatitis B virus-related cirrhosis.

Data sources  Most information was pulled from a literature search (Pubmed 2000 to 2011) using the keywords of antiviral and decompensated hepatitis B virus-related cirrhosis. Relevant book chapters were also reviewed.

Study selection  Well-controlled, prospective landmark studies and review articles on antiviral therapy in decompesated hepatitis B virus-related cirrhosis were selected.

Results  Specific antiviral agents not only control viral replication, which permits liver transplantation, but also improve liver function so significantly that patients could be removed from the transplant waiting list. However, the emergence of drug-resistant mutants can result in treatment failure. Combination therapy is a save-strategy in drug-resistant.

Conclusions  Although the treatment of end-stage liver disease is still a challenge worldwide, antiviral therapy has altered the natural history of hepatitis B patients with decompensated cirrhosis. The approval of the new generation of antivirals is opening new perspectives for finding the optimal antiviral treatment for patients with decompensated cirrhosis and preventing antiviral resistance. A combination of antivirals may be one of the future strategies for fulfilling these goals.

     

Effects of estrogen on collagen gel contraction by human retinal glial cells

Background  There are definite gender differences in patients with macular holes. Menopausal women over 50 years are most affected. We aimed to observe the effect of estrogen on collagen gel contraction by cultured human retinal glial cells. It is speculated that estrogen could strengthen the tensile stress of the macula by maintaining the correct morphology and contraction.

Methods  Estrogen was used to determine its effects on collagen gel contraction, and its function was measured using morphological changes in cells. Human retinal glial cells were cultured in collagen solution. The cells were then exposed to collagen gels and the degree of contraction of the gel was determined.

Results  Estrogen at differing concentrations had no effect on the growth of human retinal glial cells. However, after exposed to collagen gel block, less contraction was noted in the estrogen-treated group than in the control group.

Conclusions  Estrogen can inhibit collagen gel contraction by glial cells. These results suggest a mechanism for macular hole formation, which is observed in menopausal females.