BACKGROUNDUncontrolled cell division is one of the hallmarks of tumor growth. Researches have been focused on numerous molecules involved in this process. Cyclins are critical regulatory proteins of cell cycle progression and/or transcription. The present study aimed to investigate the anti-proliferative effect of cyclin L2, and to define its growth regulatory mechanisms using human lung adenocarcinoma cell line A549.

METHODSHuman cyclin L2 was transfected into human lung adenocarcinoma cells (A549 cell), and was expressed in a mammalian expression vector pcDNA3.1. The effects and mechanisms of the cyclin L2 in cell growth, cell cycle analysis and apoptosis were studied by 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT), flow cytometry or Western blot, respectively.

RESULTSOverexpression of cyclin L2 inhibited the growth of A549 cells. Cell cycle analysis in cells transfected with pCCNL2 revealed an increment in proportion in G0/G1 phase ((68.07 +/- 4.2)%) in contrast to (60.39 +/- 2.82)% of the cells transfected with mock vector. Apoptosis occurred in (7.25 +/- 0.98)% cells transfected with pCCNL2, as compared with (1.25 +/- 0.21)% of the mock vector control group. Cyclin L2-induced-G0/G1 arrest and apoptosis involved upregulation of caspase-3 and downregulation of Bcl-2 and survivin.

CONCLUSIONThe results indicate that overexpression of cyclin L2 protein may promote efficient growth inhibition of human lung adenocarcinoma cells by inducing G0/G1 cell cycle arrest and apoptosis.

Apoptosis ; Caspase 3 ; biosynthesis ; Cell Cycle ; Cell Line, Tumor ; Cyclins ; genetics ; physiology ; Humans ; Lung Neoplasms ; pathology ; Proto-Oncogene Proteins c-bcl-2 ; antagonists & inhibitors ; Transcription Factors ; genetics ; physiology ; Transfection

OBJECTIVETo investigate the influence of aerosols on the expression of cyclin B(1), cyclin C and proliferating cell nuclear antigen (PCNA) in wound tissue healing of burned rat models.

METHODSSprague Dawley (SD) rats were inflicted as the deep partial thickness burn models. Rats were randomly divided into experimental group and control group. The experimental group were treated with aerosols. Samples were collected in 1 approximately 10 postburn days. Immunohistochemistry and image analysis methods were conducted to examine the expression of cyclin B(1), cyclin C and PCNA in both experimental and control groups.

RESULTSThe expression of cyclin C in experimental group was detected in nucleus of skin basal cell on the second postburn day, increased evidently at the fifth days and sustained at high expression level up to the tenth days after treatment. The expression of cyclin C in experimental group was significantly higher than control group (P < 0.05). The expression of PCNA was first observed in skin basal cell nucleus and hair follicle cell nucleus in both experimental and control group on the third postburn day. The expression of PCNA increased evidently at the fifth days in experimental after treatment and that increased evidently at the seventh days in control group, which showed there were lots of active proliferation cell. And the difference of the expression of PCNA between experimental and control group was significant (P < 0.01). The expression of cyclin B(1) was detected in nucleus and cytoplasm of skin basal cell in both groups on the third postburn day, and no difference between the experimental and control group (P > 0.05).

CONCLUSIONSAerosols can up-regulate the expression of cyclin C and PCNA in skin basal cell nucleus. Therefore the aerosols can accelerate wound tissue healing.

Aerosols ; Animals ; Burns ; metabolism ; therapy ; Cyclin B ; biosynthesis ; Cyclin B1 ; Cyclin C ; Cyclins ; biosynthesis ; Disease Models, Animal ; Electric Stimulation Therapy ; methods ; Female ; Proliferating Cell Nuclear Antigen ; biosynthesis ; Rats ; Rats, Sprague-Dawley ; Wound Healing ; physiology

The pathogenesis of aplastic anemia (AA) was explored and the effects of AA serum on the expression of crucial cyclin D isoform (cyclin D3) in umbilical cord blood hematopoietic stem/progenitor cells were observed. The CD34+ cells were isolated from the cord blood with MIDI-MACS Semi-solid methylcellulose culture technique was used to measure the formation of CFU-GM; The expression level of cyclin D3 was assayed by semi-quantitative RT-PCR and Western-blot after the hematopoietic stem/progenitor cells were incubated in AA serum. The results showed that the AA serum could inhibit the formation of CFU-GM and down regulate the expression level of the cyclin D3 at the mRNA and protein level respectively. In conclusion, the AA serum could inhibit the proliferation of hematopoietic stem cells and down regulate level of cyclin D3, which might be one mechanism of hematopoiesis inhibition in AA.
Anemia, Aplastic ; blood ; Antigens, CD34 ; metabolism ; Cells, Cultured ; Colony-Forming Units Assay ; Cyclin D3 ; Cyclins ; biosynthesis ; genetics ; Female ; Fetal Blood ; cytology ; Hematopoietic Stem Cells ; cytology ; Humans ; Male ; Protein Isoforms ; biosynthesis ; genetics ; RNA, Messenger ; biosynthesis ; genetics ; Serum

Intimal hyperplasia is defined as the abnormal migration and proliferation of vascular smooth muscle cells (VSMCs) with deposition of extracellular matrix. However, the cell cycle regulatory mechanisms of injury-induced VSMC proliferation are largely unknown. To examine the expression kinetics of cell cycle regulatory factors which is known to be worked positively or negatively, we used rat balloon injury model. Marked induction of proliferating cell nuclear antigen (PCNA), G1/S cyclin-dependent kinase (cdk2), and its regulatory subunit (cyclin E) occurred between 1 and 3 days after balloon arterial injury, and this was sustained for up to 7 days and then declined. However, the induction of the negative regulators, p21 and p27, occurred between 3 and 5 days of injury, peaked after 7 and 14 days and was then sustained. VSMC proliferation after balloon catheter injury of the rat iliac artery is associated with coordinated expression of positive (cdk2, cyclin E and PCNA) and negative (p21, p27) regulators. Cell cycle regulators such as cdk2, cyclin E, p21, p27 may be suitable targets for the control of intimal hyperplasia.
Animals ; Arteries/*pathology ; Balloon Dilatation/*adverse effects ; Blotting, Western ; CDC2-CDC28 Kinases/biosynthesis ; Cell Cycle ; Cell Cycle Proteins/biosynthesis ; Cell Division ; Cyclin E/biosynthesis ; Cyclins/biosynthesis ; Endothelium, Vascular/pathology ; Extracellular Matrix/metabolism ; Hyperplasia/pathology ; Iliac Artery/pathology ; Immunohistochemistry ; Male ; Myocytes, Smooth Muscle/*cytology ; Proliferating Cell Nuclear Antigen/biosynthesis ; Rats ; Rats, Sprague-Dawley ; Support, Non-U.S. Gov't ; Time Factors ; Tumor Suppressor Proteins/biosynthesis

The pathogenesis of aplastic anemia (AA) was explored and the effects of AA serum on the expression of crucial cyclin D isoform (cyclin D3) in umbilical cord blood hematopoietic stem/progenitor cells were observed. The CD34+ cells were isolated from the cord blood with MIDI-MACS Semi-solid methylcellulose culture technique was used to measure the formation of CFU-GM; The expression level of cyclin D3 was assayed by semi-quantitative RT-PCR and Western-blot after the hematopoietic stem/progenitor cells were incubated in AA serum. The results showed that the AA serum could inhibit the formation of CFU-GM and down regulate the expression level of the cyclin D3 at the mRNA and protein level respectively. In conclusion, the AA serum could inhibit the proliferation of hematopoietic stem cells and down regulate level of cyclin D3, which might be one mechanism of hematopoiesis inhibition in AA.
Anemia, Aplastic/*blood ; Antigens, CD34/*metabolism ; Cells, Cultured ; Colony-Forming Units Assay ; Cyclins/*biosynthesis ; Cyclins/genetics ; Fetal Blood/cytology ; Hematopoietic Stem Cells/*cytology ; Protein Isoforms/biosynthesis ; Protein Isoforms/genetics ; RNA, Messenger/biosynthesis ; RNA, Messenger/genetics ; Serum

This paper was designed to investigate the expression of p53, p21 of A549 cell strains under hypoxic condition and the effect of trichostatin A (TSA), the inhibitor of histone deacetylasel (HDAC1) on their expression. The authors designed 1 normoxia group (control group) and 6 hypoxia groups (experimental group): hypoxia 6 h group (A), TSA+hypoxia 6 h (B), hypoxia 12 h group (C), hypoxia 24 h group (D), TSA+hypoxia 24 h (E), hypoxia 48 h group (F). The expression of HDAC1 in A549 cells was examined by using Western blot and the expression of p53, p21 in A549 cells and the effect of TSA on them were determined by using immunohistochemistry and reverse transcription-polymerase chain reaction (RT-PCR). The A value expressed by HDAC1 in A549 cell strains was 138+/-11 in the control group, 78+/-4, 86+/-5, 124+/-3, 120+/-9 in experimental groups A, C, D, F, respectively. The A value of the expression of the protein and mRNA of p53 in A549 cell strains were 0.12+/-0.02, 0.62+/-0.02 in the control group, 0.10+/-0.03, 0.32 +/-0.03; 0.11+/-0.01, 0.33+/-0.02; 0.13+/-0.03, 0.58+/-0.01; 0.12+/-0.02, 0.56+/-0.02 in experimental group A, B, D, E, respectively. The A value of the expression of the protein and mRNA of p21 in A549 cell strains were 0.17+/-0.03, 0.62+/-0.03 in the control group, 0.16+/-0.02, 0.50+/-0.02; 0.14+/-0.02, 0.36+/-0.02; 0.15+/-0.03, 0.49+/-0.03; 0.13+/-0.02, 0.33+/-0.02 in experimental groups A, B, D, E, respectively. These results indicate that the expression of HDAC1 is regulated by hypoxia and the effect of TSA is closely related to the expression of P21 under hypoxia condition.
Adenocarcinoma ; metabolism ; pathology ; Cell Hypoxia ; Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; biosynthesis ; genetics ; Histone Deacetylase Inhibitors ; Humans ; Hydroxamic Acids ; pharmacology ; Lung Neoplasms ; metabolism ; pathology ; RNA, Messenger ; biosynthesis ; genetics ; Tumor Cells, Cultured ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics

In order to explore the molecular mechanisms of sodium butyrate and trichostatin A on K562 cell proliferation/differentiation, K562 cells were grown in the absence or presence of sodium butyrate or trichostatin A. The percentage of viable cells was determined by trypan blue exclusion. Differentiation was determined by nitro-blue tetrazolium (NBT) reduction and cell surface adhesion molecules analyzed by FACS. Cell cycle distribution was studied after DNA staining by propidium iodide. Cell cycle regulatory proteins were detected by Western blot and reverse transcription-polymerase chain reaction. The results showed that sodium butyrate blocked cells mainly at the G0/G1 phase of the cell cycle, whereas trichostatin A arrested the cells at G2 phase. Sodium butyrate could down-regulate the mRNA expression of cyclin D1, but not affect its protein expression; down-regulate the protein expression of cyclin D3, but not affect its mRNA expression. Trichostatin A showed similar effects on cyclin D1 and D3 as sodium butyrate. Both sodium butyrate and trichostatin A could stimulate p21 expression of K562 cells at mRNA and protein levels. It may be concluded that sodium butyrate and trichostatin A could promote the proliferation/differentiation of the K562 cells, which might be contributed to the induced expression of cyclin D3 and p21 proteins.
Butyrates ; pharmacology ; Cell Cycle ; drug effects ; Cell Differentiation ; drug effects ; Cell Division ; drug effects ; Cyclin D ; Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; biosynthesis ; genetics ; Gene Expression ; drug effects ; Histone Deacetylase Inhibitors ; Humans ; Hydroxamic Acids ; pharmacology ; K562 Cells

Phagocytosis of serum- and IgG-opsonized zymosan (SOZ and IOZ, respectively) particles into J774A.1 macrophages induced apoptosis of the cells, accompanied by the expression of p21(WAF1), one of cyclin-dependent protein kinase (CDK) inhibitors. Furthermore, phagocytosis of SOZ and IOZ particles into macophages induced superoxide formation. Tat-superoxide dismutase (SOD), which is readily transduced into the cells using Tat-domain, protected the cells from the apoptosis induced by phagocytosis of SOZ and IOZ particles. lipopolysaccharide (LPS)/interferon-gamma (IFN-gamma) also caused the apoptosis of the cells. However, Tat-SOD could not protect the cells from LPS/IFN-gamma induced apoptosis, suggesting that apoptosis mechanisms involved are different from each other. In the present study, we determined the amounts of nitric oxide (NO) produced by SOZ, IOZ, and LPS/IFN-gamma, and found that SOZ and IOZ did not induce the generation of NO in macrophages, whereas LPS/ IFN-gamma did. The apoptosis due to phagocytosis was accompanied with the release of cytochrome c from mitochondrial membrane to cytosolic fraction. Furthermore, SOZ and IOZ induced the cleavage of procasapase-3 (35 kDa) to give rise to an active caspase-3 (20 kDa), which was blocked by Tat- SOD but not by 2-phenyl-4,4,5,5-tetramethylimidazoline-1-oxyl 3-oxide (PTIO), a scavenger of NO. On the other hand, LPS/IFN-gamma caused the activation of procaspase-3, which was blocked by PTIO but not by Tat-SOD. Taken together, phagocytosis of SOZ and IOZ particles induced apoptosis through superoxide but not NO in macrophages, accompanied with the release of cytochrome c and the activation of caspase-3.
Apoptosis/*immunology ; Caspases/metabolism ; Cell Line ; Cyclins/biosynthesis ; Cytochromes c/metabolism ; Immunoglobulin G/*immunology ; Interferon Type II/pharmacology ; Lipopolysaccharides/pharmacology ; Macrophages/*immunology/metabolism ; Nitric Oxide/*metabolism ; Opsonins/immunology ; Phagocytosis/*physiology ; Superoxide Dismutase/metabolism ; Superoxides/*metabolism ; Zymosan

OBJECTIVETo study effects of alternative transcripts of ING1 transfection on human cancer cell lines.

METHODSp47/ING1A and p33/ING1B expression vehicles were constructed and introduced into a human breast cancer cell line MCF-7 and a human lung cancer cell line PAa, both expressing wild-type p53 protein. Growth characteristics of the transfectants and potentially related genes were analyzed.

RESULTSThe levels of p47/ING1A and p33/ING1B protein elevated respectively in tumor cells of MCF-7 and PAa after transfected with p47/ING1A and p33/ING1B, and the latter was much higher than that of the former. Ectopic overexpression of p33/ING1B effectively blocked tumor cell growth and arrested cells in the G(0) approximately G(1) phase of the cell cycle (P < 0.01), while p47/ING1A gave no effect on cell growth or cell cycle. Tumor cells overexpressing p33/ING1B contained more p21(WAF1) protein than that of the control cells, with undisturbed p53 protein level.

CONCLUSIONSExpression of two different transcripts of ING1 may have different effects on tumor cell growth. p33/ING1B may cooperate with p53 in stimulating expression of p21(WAF1) gene, thus to arrest cell cycle and to inhibit tumor cell growth. p33/ING1B may be considered to be a candidate as a partner of p53 in gene therapy.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Alternative Splicing ; Breast Neoplasms ; genetics ; metabolism ; pathology ; Cell Cycle ; Cell Cycle Proteins ; Cell Division ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; biosynthesis ; DNA-Binding Proteins ; Genes, Tumor Suppressor ; Humans ; Inhibitor of Growth Protein 1 ; Intracellular Signaling Peptides and Proteins ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Nuclear Proteins ; Protein Biosynthesis ; Proteins ; genetics ; Transfection ; Tumor Suppressor Protein p53 ; biosynthesis ; Tumor Suppressor Proteins

OBJECTIVETo study the growth inhibitory effects of p21WAF1 and p53 overexpression in human lung adenocarcinoma cell line.

METHODSThe p21WAF1 and p53 gene were transfected respectively into a human lung adenocarcinoma cell line, GLC-82. Flow cytometry (FLC), transmission electron microscopy (EM) and TUNEL technique were used to evaluate cell growth and identify apoptosis.

RESULTSThe GLC-82 transfected by p21 plasmid showed increased cell number in G1 phase of cell cycle, decreased proliferation potential and decreased cloning efficiency. Apoptosis have not been detected neither on EM nor by TUNEL technique, whereas the GLC-82 infected by Ad-p53 showed significantly decreased proliferation potential and some of them even died, in addition apoptosis was confirmed by TUNEL technique.

CONCLUSIONThe results indicate that p21WAF1 and p53 can inhibit proliferation; p53 also can induce apoptosis of lung adenocarcinoma cell. Therefore, these two genes should have a wide application in gene therapy of tumors in future.

Adenocarcinoma ; genetics ; metabolism ; pathology ; Apoptosis ; Cell Line, Tumor ; Cyclin-Dependent Kinase Inhibitor p21 ; Cyclins ; biosynthesis ; genetics ; Gene Expression ; Humans ; Lung Neoplasms ; genetics ; metabolism ; pathology ; Tumor Suppressor Protein p53 ; biosynthesis ; genetics