Extracellular Adenine Nucleotides and Adenosine Modulate the Growth and Survival of THP-1 Leukemia Cells

A new approach to improve the effectiveness of acute myeloid leukemia (AML) treatment is to use the properties of purinergic signaling molecules secreted into the bone marrow milieu in response to leukemic cell growth. Therefore, our study aimed to evaluate the effects of extracellular adenine nucleotides and adenosine on the growth and death parameters in the leukemic THP-1 cell line. Cells were exposed to ATP, ADP, AMP, adenosine and nonhydrolyzable analogues of ATP and ADP (ATPγS and ADPβS) in a 1–1000 μM broad concentration range. The basal mRNA expression of the P1 and P2 receptors was evaluated by real-time PCR. Changes in the processes of cell growth and death were assessed by flow cytometry analysis of proliferation, cell cycle and apoptosis. Chemotaxis toward stromal cell-derived factor-1 (SDF-1) was performed using the modified Boyden chamber assay, and chemokine receptor type 4 (CXCR4) surface expression was quantified by flow cytometry. We indicated several antileukemic actions. High micromolar concentrations (100–1000 μM) of extracellular adenine nucleotides and adenosine inhibit the growth of cells by arresting the cell cycle and/or inducing apoptosis. ATP is characterized by the highest potency and widest range of effects, and is responsible for the cell cycle arrest and the apoptosis induction. Compared to ATP, the effect of ADP is slightly weaker. Adenosine mostly has a cytotoxic effect, with the induction of apoptosis. The last studied nucleotide, AMP, demonstrated only a weak cytotoxic effect without affecting the cell cycle. In addition, cell migration towards SDF-1 was inhibited by low micromolar concentrations (10 μM). One of the reasons for this action of ATPγS and adenosine was a reduction in CXCR4 surface expression, but this only partially explains the mechanism of antimigratory action. In summary, extracellular adenine nucleotides and adenosine inhibit THP-1 cell growth, cause death of cells and modulate the functioning of the SDF-1/CXCR4 axis. Thus, they negatively affect the processes that are responsible for the progression of AML and the difficulties in AML treatment.

Download Full-text

Apoptosis Was Induced by Casticin in Acute Myelocytic Leukemia Cells

Blood ◽

10.1182/blood.v112.11.3991.3991 ◽

2008 ◽

Vol 112 (11) ◽

pp. 3991-3991

Author(s):

Jie Jin ◽

Jia-Kun Shen ◽

Hua-ping Du ◽

Min Yang ◽

Yun-Gui Wang

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Death ◽

Leukemic Cell ◽

K562 Cells ◽

Acute Myelocytic Leukemia ◽

Myelocytic Leukemia ◽

Cycle Arrest ◽

M Phase ◽

Ic50 Values

Abstract Casticin, a component from Vitex rotundifolia wich was widely used as an anti-inflammatory agent in Chinese traditional medicine, was reported to have anti-tumor activities in lung cancer and breast cancer. There are yet no reports on roles against acute myelocytic leukemia (AML). This study aims to elucidate the anti-leukemic activity of casticin on AML cells. We investigated the efficient efficacy and the mechanisms by which casticin triggers cell death in AML cells by analyzing cell cycle perturbations, apoptosis-related marker expression. Cell viability was measured by MTT method; apoptosis and cell cycle arrest were determined by flow cytometry and AV-PI assay. Western blot was performed to measure the apoptosis-related marker. Concentration-dependant cell deaths were observed in AML cell lines including K562, U937 and THP-1, with IC50 values of 24h (hours) being 47.4μM, 67.8μM and 61.7μM, respectively. Time-dependant cell deaths were also observed. At the concentration of 20μM casticin, 45.7%, 76.1% and 80.9% of K562 cells were inhibited at 24h, 48h and 72h, respectively; 24.7%, 30% and 61% of U937 cells were inhibited at 24h, 48h and 72h, respectively; while for THP-1, 29%, 41.8% and 53.9% were inhibited at 24h, 48h and 72h, respectively. Apoptosis was found using AV-PI staining by flow cytometry analysis. We observed an obvious G2/M phase increase prolongation in casticin treated K562 cells. BThe distribitions of G2/M phase were 2.9%, 33.6%, 75.3%, 54.9%, 29.7% and 27.0% in K562 cells after treated by 20μM casticin for 0h, 6h, 12h, 24h, 36h and 48h, respectively. Furthermore, apoptosis-related proteins, PARP and caspase 3, were cleaved in casticin treated K562 cells. Taken together, these results demonstrated that casticin can induce leukemic cell death through apoptosis, suggesting that casticin could be a promising therapeutic agent against acute myeloid leukemia.

Download Full-text

Essential Role for the MAML1 Co-Activator In T-ALL

Blood ◽

10.1182/blood.v116.21.2501.2501 ◽

2010 ◽

Vol 116 (21) ◽

pp. 2501-2501

Author(s):

Chunxia Cao ◽

Liang Tian ◽

Jian-Liang Li ◽

James D. Griffin ◽

Suming Huang ◽

...

Keyword(s):

Cell Growth ◽

Notch Signaling ◽

Leukemic Cell ◽

Genetic Alterations ◽

Leukemic Cells ◽

Cell Renewal ◽

Hematopoietic Stem ◽

Growth And Survival ◽

Oncogenic Pathways ◽

Cell Growth And Survival

Abstract Abstract 2501 T cell acute lymphoblastic leukemia (T-ALL) is the most common malignancy in children and accounts for nearly one third of all pediatric cancers. In this type of leukemia, lymphoid progenitor cells that are responsible for the generation of mature lymphocytes become genetically altered, leading to deregulated proliferation, survival, and clonal expansion. Two common genetic alterations frequently associated with this disease are mutations in the NOTCH1 cell-surface receptor and aberrant expression of the TAL1 transcription factor, with each abnormality detected in more than half of human T-ALL patients. The mutations in the NOTCH1 gene result in the aberrant activation of Notch signaling, a highly conserved signal transduction pathway that is critical for lymphocyte growth, maturation and survival. The constitutive activation of Notch signaling induces leukemia in mouse models and is required for human T-ALL leukemic cell growth and survival. On the other hand, TAL1 is required for the functions of hematopoietic stem cells and is essential for the generation of the erythroid and myeloid lineages. The ectopic activation of the TAL1 gene deregulates normal hematopoietic stem cell renewal and differentiation, leading to leukemia in cooperation with other oncogenes. Therefore, Notch and TAL1 oncogenic activities are critical for the initiation and maintenance of T-ALL. In this study, we investigated the role of a transcriptional co-activator, MAML1, in regulating NOTCH1 and TAL1 transforming activities in leukemic cells. In addition to its known function in co-activating Notch signaling, we found that MAML1 is a novel interacting partner for TAL1. MAML1 also enhanced TAL1 transcriptional activities, suggesting a role for MAML1 in TAL1-regulated transcription and leukemogenesis. A subset of T-ALL leukemic cells exhibit aberrant activation in both the NOTCH1 and TAL1 activities; thus, it suggests that these two genetic alterations cooperate in promoting leukemic cell growth and survival. Indeed, we found that the combined inhibition of both the pathways (via the pharmacological blockade of Notch signaling and shRNA-mediated TAL1 knockdown) results in synergistic responses in leukemic cells that carry genetic alterations in both the NOTCH1 and TAL1 genes, indicating that the two pathways synergize in promoting T-ALL. Since MAML1 appears to be a common key regulator for both TAL1 and Notch1 pathways, we next determined whether MAML1 expression level affects leukemic cell growth and survival. Gene knockdown studies suggest that MAML1 is essential for leukemic cell growth and survival by possibly regulating NOTCH1 and TAL1-mediated transcription. Overall, our data reveals a novel common regulatory mechanism for both NOTCH1 and TAL1 oncogenic pathways, and suggest that the manipulation of MAML1 expression or functional activities will affect leukemia initiation and progression. Therefore, our current studies focus on assessing the MAML1 co-activator as a target for these two oncogenic pathways. Disclosures: Griffin: Novartis: Consultancy, Research Funding.

Download Full-text

Diffuse Large B-Cell Lymphomas Utilize Endogenous and Exogenous Fatty Acids for Cell Growth and Survival

Blood ◽

10.1182/blood.v120.21.1581.1581 ◽

2012 ◽

Vol 120 (21) ◽

pp. 1581-1581

Author(s):

Frederick Lansigan ◽

Wilson L Davis ◽

Nancy Kuemmerle ◽

Leslie E Lupien ◽

Valeriya Posternak ◽

...

Keyword(s):

Flow Cytometry ◽

Fatty Acid ◽

Cell Growth ◽

B Cell ◽

Cancer Cells ◽

Cell Lines ◽

Functional Significance ◽

De Novo ◽

Growth And Survival ◽

Large B Cell

Abstract Abstract 1581 Background It is well-recognized that de novo long chain fatty acid (FA) synthesis, driven by the key enzyme fatty acid synthase (FASN), is crucial for the growth and survival of many types of cancer cells. We and others have observed FASN protein expression in diffuse large B-cell lymphoma (DLBCL) tumors. Furthermore, we have shown that higher levels of FASN in DLBCL tumors strongly predicted inferior survival, which was independent from the international prognostic index. We also recently demonstrated that, in addition to FA synthesis, various cancer cells can acquire FA from circulating lipoproteins, using the secreted enzyme lipoprotein lipase (LPL), and that this promotes cell growth. DLBCL, however, has never been examined in this regard. In this study, we investigated the functional significance of both de novo FA synthesis via FASN and exogenous FA uptake via LPL in DLBCL. Methods Levels of FASN and LPL mRNAs in DLBCL cell lines (SUDHL4, SUDHL10, OCI-LY3, OCI-LY19) were studied using reverse transcriptase polymerase chain reaction. We determined FASN and LPL protein expression by flow cytometry using a novel anti-LPL antibody that we developed. DLBCL cell lines were cultured +/− Cerulenin (an inhibitor of FASN), Orlistat (an inhibitor of FASN and LPL), or in lipoprotein-depleted serum +/− supplementation with very low density lipoprotein (VLDL) particles. The MTT assay was used to assess cell proliferation. Results DLBCL cell lines exhibited >10-fold variation in levels of FASN mRNA. Cerulenin and Orlistat each caused dose-dependent inhibition of proliferation of each cell line. The cells were partially rescued by the addition of palmitic acid, the FA product of FASN. Surprisingly, flow cytometry revealed that SUDHL4 and OCI-LY3 cells, which did not secrete LPL or show detectable LPL activity, displayed the enzyme on the cell surface. Moreover, in stark contrast to several other cancer cell lines, DLBCL cells were exquisitely sensitive to withdrawal of lipoproteins from the culture media. Indeed, 75–95% of the cells underwent apoptosis after only 24 hours in lipoprotein-depleted serum. In complete serum, the provision of VLDL particles did not rescue DLBCL cells from FA synthesis inhibition using Cerulenin, suggesting that the serum contains sufficient lipoproteins to saturate the FA uptake system. This prediction was validated in experiments utilizing lipoprotein-depleted serum, in which add-back of VLDL particles completely rescued the cells from Cerulenin-induced demise in a dose-related manner, with full restoration at approximately 100–200mcg/ml of VLDL. Conclusions Our data demonstrate that DLBCL cells employ both de novo FA synthesis via FASN and exogenous FA uptake using LPL to satisfy their strict requirement for FA. Interference with either pathway, using FASN inhibitors or lipoprotein-depleted serum, is cytotoxic indicating that neither alone is sufficient to support proliferation. Further, DLBCL cells show a striking dependency on exogenous FA of dietary origin compared with all other cancer cells we have examined. The observation that the cell lines can be rescued by provision of VLDL particles strongly supports the functional significance of the exogenous FA uptake pathway for DLBCL. Our data thus demonstrate that the extracellular lipase LPL is critical for the growth and survival of DLBCL cells. Surprisingly, the cells deploy LPL to their surface, and we speculate that this promotes efficient FA acquisition from circulating lipoproteins. Recognition that DLBCL relies on both synthesis and uptake of FA will provide guidance for drug development and dietary modifications to effectively target the metabolic requirements of this tumor. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

Aptamer TY04 Inhibits Multiple Myeloma Cell Growth Via Cell Cycle Arrest

Blood ◽

10.1182/blood.v122.21.5390.5390 ◽

2013 ◽

Vol 122 (21) ◽

pp. 5390-5390

Author(s):

Jing Liu ◽

Hong-Juan Dai ◽

Bian-Ying Ma ◽

Jian-Hui Song ◽

Hui-yong Chen ◽

...

Keyword(s):

Cell Cycle ◽

Multiple Myeloma ◽

Flow Cytometry ◽

Cell Growth ◽

Cell Lines ◽

Myeloma Cell ◽

Multiple Myeloma Cell ◽

Cycle Arrest ◽

M Phase ◽

Target Molecules

Abstract Multiple myeloma (MM), also known as plasma cell myeloma, is characterized by accumulation of clonal plasma cells in the bone marrow and overproduction of monoclonal immunoglobulin (Ig) in the blood or urine. MM accounts for approximately 10% of all hematologic malignancies. Despite recent advances in the understanding and treatment of this disease, MM remains an incurable disease in the vast majority. With conventional chemotherapy, the 5-year median survival rate for MM patients is approximately 25%. Aptamers are single-stranded RNA or DNA sequences that bind to target molecules with high affinity and specificity. Compared with antibodies, aptamers have unique advantages including easy chemical synthesis and modification, low toxicity, lack of immunogenicity, and rapid tissue penetration, Based on these advantages, aptamers show great potential for therapeutic application. The aptamer TY04 is a single-stranded DNA (ssDNA) generated by a method named cell-based systematic evolution of ligands by exponential enrichment (cell-SELEX), We found TY04 strongly inhibited the growth of multiple myeloma cell lines including MM1.S, NCI-H929, KM3 and OPM2,The concentration of TY04 to inhibit 50% cell growth (IC50) on MM1.S was 3.89 μM. In contrast, TY04 had no effect on the growth of non-tumor cell lines — immortal B lymphoblastoid cell lines. Next, we used MM1.S cell line as the model to study the mechanism of TY04 anti- multiple myeloma. Flow cytometry analysis showed that TY04 with the sequence specifically bind to MM1.S cells when compared with unselected ssDNA library control. To investigate whether the target molecules of TY04 are membrane proteins on cell surface, MM1.S cells were treated with trypsin and proteinase k for 2 or 10 minutes before incubation with TY04. The result revealed that TY04 lost partly recognition ability on treated cells, indicating that the target molecules were most likely membrane proteins. Furthermore, we evaluated the cell cycle distribution of MM1.S after TY04 treatment. We found that TY04 significantly caused cell-cycle arrest in G2/M phase. The percentage of G2/M phase cells increased from 30.1±1.56 to 53.2±6.36. To identify the underlying molecular mechanism, G2/M-related proteins were assayed by flow cytometry. Following TY04 treatment, a concomitant inhibition of ERK1/2, cyclin B, CDK1 and γ-tubulin expression occurred. Meanwhile, human cell cycle PCR array was used to analyze the expression of 84 genes key to cell cycle regulation in TY04-treated MM1.S cells. Our results indicated that aptamer TY04 decreased the genes expression of CCNB1, CCNB2, BIRC5, BRCA1 and CCNH, which were involved in the progress of G2/M phase. All these results are significant in that they provide a framework for further exploring the use of TY04 as a novel anti-multiple myeloma agent. Disclosures: No relevant conflicts of interest to declare.

Download Full-text

Efek Sinergis Kombinasi Ekstrak Air Akar Batu (Gerrardanthus Macrorhizus) dengan Doxorubicin pada Sel Kanker Payudara T47D

Buletin Penelitian Kesehatan ◽

10.22435/bpk.v46i3.878 ◽

2018 ◽

Vol 46 (3) ◽

pp. 183-190

Author(s):

Sari Haryanti

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cytotoxic Effect ◽

Clinical Results ◽

Morphological Alteration ◽

T47d Cells ◽

Natural Agents ◽

Chemotherapy Agent ◽

M Phase ◽

Apoptotic Induction

Doxorubicin is one of the most widely used chemotherapy agents in cancer therapy. The clinical results of doxorubicin are limited by cardiotoxic side effects which correlates with dose. Combination of doxorubicin with natural agents is a promising strategy for reducing its therapeutic doses as well as decreasing cardiotoxic effects. This study was conducted to investigate cytotoxic activity of caudex Gerrardanthus macrorhizus aqueous extract and its combination with doxorubicin on T47D cells. Dried caudex powder was extracted by infusion method, evaporated in oven 400C to get dried extract (GM). The MTT assay was performed to determine cytotoxic effect, either alone or in combination. Flow cytometry is used to observe cell cycle profile and apoptotic induction. GM alone did not exhibit cytotoxic effects but caused morphological alteration in T47D cells. Nevertheless, GM 40 μg/mL was able to improve cytotoxic effect of doxorubicin to 51%. Combination of doxorubicin 3 nM and 40 μg/mL GMA inhibited cell cycle in G2/M phase. This combination also resulted apoptotic induction, compared to untreated cell and each single treatment. Based on the results, caudex G. macrorhizus is potential to be further investigated as a co-chemotherapy agent with doxorubicin. Abstrak Doxorubicin adalah salah satu agen kemoterapi yang digunakan secara luas dalam terapi kanker. Hasil klinis doxorubicin dibatasi oleh kardiotoksisitas yang berkorelasi dengan besaran dosis. Kombinasi doxorubicin dengan bahan alam merupakan strategi yang menjanjikan untuk mengurangi dosis terapetik doxorubicin sehingga dapat menurunkan efek samping kardiotoksik. Penelitian ini dilakukan untuk mengkaji aktivitas sitotoksik ekstrak air caudex Gerrardanthus macrorhizus dan kombinasinya dengan doxorubicin pada sel T47D. Serbuk caudex G. macrorhizus (GM) kering diekstraksi dengan metode infusa, kemudian diuapkan dalam oven 400C. Uji MTT dilakukan untuk menguji efek sitotoksik ekstrak, baik tunggal maupun kombinasi dengan doxorubicin. Flow cytometry digunakan untuk mengetahui profil siklus sel dan induksi apoptosis. Hasil penelitian menunjukkan ekstrak tunggal tidak memberikan efek sitotoksik namun mengakibatkan perubahan morfologi pada sel T47D. Ekstrak GM 40 μg/mL mampu meningkatkan efek sitotoksik doxorubicin 3 nM hingga 51%. Kombinasi doxorubicin 3 nM dan ekstrak 40 μg/mL menghambat siklus sel pada fase G2/M dan meningkatkan induksi apoptosis, dibandingkan kontrol sel dan masing-masing perlakuan tunggalnya. Berdasarkan hasil penelitian ini, caudex G. macrorhizus berpotensi untuk diteliti lebih lanjut sebagai agen ko-kemoterapi dengan doxorubicin.

Download Full-text

Title: GPC3 in the Exosomes from Hepatocellular Carcinoma HepG2 Cells

10.21203/rs.3.rs-656873/v1 ◽

2021 ◽

Author(s):

Chunwen Pu ◽

Qi Wang ◽

Aijun Sun ◽

Ping Sun ◽

Hui Huang ◽

...

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Growth ◽

Liver Cancer ◽

Signaling Pathway ◽

Tumor Cells ◽

Hepg2 Cells ◽

Normal Liver ◽

Promoting Effect ◽

Normal Cells

Abstract Background Exosomes play an important role in regulating the growth in normal and abnormal cells. Exosomes secreted from tumor cells are also involved in regulating the growth behaviors of normal cells and tumor cells. Methods HepG2 cells, LO2 and HepG2 cells with GPC3 knocked down using shRNA (HepG2-shGPC3), were treated with different concentrations of GPC3. The effects of different concentrations of GPC3 on cell growth and apoptosis were determined using CCK8 and flow cytometry. HepG2 exosomes (Exo) and exosomes of HepG2 cells with GPC3 knocked down using shRNA (shGPC3-Exo) were used to treat LO2 and HepG2 cells separately. Cell growth was measured by CCK8 kit. The cell cycle and apoptosis were measured by flow cytometry. The expression of GPC3/WNT3A/β-catenin signal protein was determined by Western blotting. Results We found GPC3 has a two-way regulation between normal cells and HCC cells, which is the innovation of this research. After treating LO2 cells and HepG2 cells with GPC3, the LO2 cell cycle was blocked in the G0/G1 phase, while cell growth was inhibited and apoptosis was promoted; however, it appeared to promote the growth of HepG2 cells. Knocking down GPC3 can inhibit the growth and promote cell apoptosis of HepG2. In subsequent experiments, we found that GPC3 was expressed in both LO2 and HepG2 exosomes, and the expression of GPC3 in HepG2 exosomes is significantly higher than that of LO2 exosomes. These results suggested that GPC3 in exosomes has the potential to become a biomarker of HCC. In addition, HepG2 exosomes (Exo) can inhibit the growth of LO2 cells and promote apoptosis, which is consistent with the effect of GPC3 treatment. Further, we found that GPC3 in shGPC3-Exo had the same effect on LO2 cells as HepG2 exosomes (Exo), but the degree of influence was reduced. shGPC3-Exo showed a promoting effect on the growth of HepG2 cells. Therefore, GPC3 in exosomes plays a role in the growth of LO2 cells and HepG2 cells. Further studies have shown that GPC3 in liver cancer exosomes regulates the proliferation, apoptosis of LO2 and HepG2 cells through the Wnt /β-catenin signaling pathway. Conclusion GPC3 in the exosomes of liver cancer cells inhibits the growth of normal liver cells and promotes apoptosis by activating the Wnt/β-catenin signaling pathway, and assists the occurrence and development of HCC.

Download Full-text

LY294002 and Metformin Cooperatively Enhance the Inhibition of Growth and the Induction of Apoptosis of Ovarian Cancer Cells

International Journal of Gynecological Cancer ◽

10.1097/igc.0b013e3182322834 ◽

2012 ◽

Vol 22 (1) ◽

pp. 15-22 ◽

Cited By ~ 28

Author(s):

Cuilan Li ◽

Vincent Wing Sun Liu ◽

David Wai Chan ◽

Kwok Ming Yao ◽

Hextan Yuen Sheung Ngan

Keyword(s):

Cell Cycle ◽

Ovarian Cancer ◽

Flow Cytometry ◽

Cell Growth ◽

Cancer Cells ◽

Cancer Cell ◽

Ovarian Cancer Cell ◽

Mtor Pathway ◽

Ovarian Cancer Cells ◽

Cancer Cell Growth

BackgroundThe phosphoinositide 3 kinase (PI3K)/v-akt murine thymoma viral oncogene homolog (AKT)/mammalian target of rapamycin (mTOR) pathway is frequently aberrantly activated in ovarian cancer and confers the chemoresistant phenotype of ovarian cancer cells. LY294002 (PI3K inhibitor) and metformin (5′-adenosine monophosphate [AMP]-activated protein kinase [AMPK] activator) are 2 drugs that were known to inhibit mTOR expression through the AKT-dependent and AKT-independent pathways, respectively. In this study, we explored the effectiveness of LY294002 and metformin in combination on inhibition of ovarian cancer cell growth.MethodsWestern blotting was used to detect the changes of PI3K/AKT/mTOR and AMPK/acetyl-CoA carboxylase (ACC) signaling activities, cell cycle control, and apoptosis. Cell growth was evaluated by cell proliferation, colony formation, and soft agar assays. Flow cytometry was used to study cell cycle distribution and cell death upon drug treatment.ResultsOur study showed that LY294002 and metformin in combination could simultaneously enhance the repression of the PI3K/AKT/mTOR pathway and the activation of the AMPK/ACC pathway. The downstream target of AKT and AMPK, mTOR, was cooperatively repressed when the drugs were used together. The cell cycle regulatory factors, p53, p27, and p21, were up-regulated. On the other hand, caspase 3 and poly (ADP-ribose) polymerase activities involved in apoptosis were also activated. Cell growth assays indicated that LY294002 and metformin could effectively inhibit ovarian cancer cell growth. Flow cytometry analysis showed that the treatment of the 2 drugs mentioned above induced cell cycle arrest at G1 phase and increased sub-G1 apoptotic cells.ConclusionThe combinational use of LY294002 and metformin can enhance inhibition of the growth and induction of the apoptosis of ovarian cancer cells. Our results may provide significant insight into the future therapeutic regimens in ovarian cancer.

Download Full-text

Relationship among Several Key Cell Cycle Events in the Developmental Cyanobacterium Anabaena sp. Strain PCC 7120

Journal of Bacteriology ◽

10.1128/jb.00524-06 ◽

2006 ◽

Vol 188 (16) ◽

pp. 5958-5965 ◽

Cited By ~ 23

Author(s):

Samer Sakr ◽

Melilotus Thyssen ◽

Michel Denis ◽

Cheng-Cai Zhang

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Division ◽

Cell Growth ◽

Cell Size ◽

Dna Content ◽

Small Cells ◽

Filamentous Cyanobacterium ◽

Pcc 7120 ◽

Heterocyst Development

ABSTRACT When grown in the absence of a source of combined nitrogen, the filamentous cyanobacterium Anabaena sp. strain PCC 7120 develops, within 24 h, a differentiated cell type called a heterocyst that is specifically involved in the fixation of N2. Cell division is required for heterocyst development, suggesting that the cell cycle could control this developmental process. In this study, we investigated several key events of the cell cycle, such as cell growth, DNA synthesis, and cell division, and explored their relationships to heterocyst development. The results of analyses by flow cytometry indicated that the DNA content increased as the cell size expanded during cell growth. The DNA content of heterocysts corresponded to the subpopulation of vegetative cells that had a big cell size, presumably those at the late stages of cell growth. Consistent with these results, most proheterocysts exhibited two nucleoids, which were resolved into a single nucleoid in most mature heterocysts. The ring structure of FtsZ, a protein required for the initiation of bacterial cell division, was present predominantly in big cells and rarely in small cells. When cell division was inhibited and consequently cells became elongated, little change in DNA content was found by measurement using flow cytometry, suggesting that inhibition of cell division may block further synthesis of DNA. The overexpression of minC, which encodes an inhibitor of FtsZ polymerization, led to the inhibition of cell division, but cells expanded in spherical form to become giant cells; structures with several cells attached together in the form of a cloverleaf could be seen frequently. These results may indicate that the relative amounts of FtsZ and MinC affect not only cell division but also the placement of the cell division planes and the cell morphology. MinC overexpression blocked heterocyst differentiation, consistent with the requirement of cell division in the control of heterocyst development.

Download Full-text

Human monoclonal antibody BT32/A6 and a cell cycle—independent glioma-associated surface antigen

Journal of Neurosurgery ◽

10.3171/jns.1995.82.3.0475 ◽

1995 ◽

Vol 82 (3) ◽

pp. 475-480 ◽

Cited By ~ 4

Author(s):

Michael D. Dan ◽

Elizabeth M. Earley ◽

Mark C. Griffin ◽

Pradip K. Maiti ◽

Ashok K. Prashar ◽

...

Keyword(s):

Cell Cycle ◽

Monoclonal Antibody ◽

Flow Cytometry ◽

Growth Rate ◽

Cell Growth ◽

Surface Antigen ◽

Immunoglobulin M ◽

Cell Growth Rate ◽

Culture Viability ◽

A Cell

✓ The purpose of this study was to ascertain how various growth parameters may influence the labeling of SK-MG-1, a human glioma cell line, by BT32/A6, a human immunoglobulin M monoclonal antibody (MAb). By growing SKMG-1 cells at different culture split ratios, significant trends in cell growth rate, culture viability, and cell cycle state were produced. Labeling of SK-MG-1 cells by BT32/A6, however, was shown to be unaffected by culture split ratio (p > 0.05) and is therefore independent of cell growth rate, culture viability, and cell cycle state. Using flow cytometry and fluorescence-activated cell sorting, BT32/A6 was shown to label a cell surface antigen on viable, clonogenic cells of SK-MG-1. Approximately 100% of SK-MG-1 cells were shown by flow cytometry to express the BT32/A6 antigen. The recognition of a glioma-associated, cell cycle-independent surface antigen by MAb BT32/A6 makes it a promising candidate for further studies aimed at elucidating its usefulness as an adjunct in the treatment of human malignant gliomas.

Download Full-text

Inhibitory Effects of Trehalose on Malignant Melanoma Cell Growth: Implications for a Novel Topical Anticancer Agent on the Ocular Surface

ISRN Ophthalmology ◽

10.5402/2012/968493 ◽

2012 ◽

Vol 2012 ◽

pp. 1-9 ◽

Cited By ~ 6

Author(s):

Takashi Kudo ◽

Kimio Takeuchi ◽

Yu-ichi Ebina ◽

Mitsuru Nakazawa

Keyword(s):

Cell Cycle ◽

Flow Cytometry ◽

Cell Growth ◽

Malignant Melanoma ◽

Melanoma Cell ◽

Ocular Surface ◽

Melanoma Cells ◽

Apoptotic Cells ◽

Inhibitory Effects ◽

Malignant Melanoma Cell

Purpose. To investigate the inhibitory effects of trehalose on malignant melanoma cell growth. Methods. We cultured human malignant melanoma cells in a medium containing trehalose (control/2.5%/5.0%/7.5%/10.0%) and used the MTT assay to evaluate the growth activities. Subsequently, trehalose was topically instilled on subconjunctivally inoculated melanoma cells in F334/NJcl-rmu/rmu rats, followed by a histopathological evaluation of tumor growth. Using flow cytometry, we compared the distribution of the cell cycle, rate of apoptotic cells, and intracellular factors related to the cell cycle in cultured melanoma cells after trehalose treatment. Results. The MTT study showed that proliferation of melanoma cells was significantly inhibited by ≧ 5% of trehalose concentrations in the culture media. Subconjunctivally inoculated melanoma cell masses were significantly smaller in eyes administered trehalose as compared to controls. Flow cytometry analyses demonstrated that the trehalose groups had increased rates of G2/M phase cells and apoptotic cells in the cell culture. These cells also exhibited increased expressions of cell-cycle inhibitory factors. Conclusions. The current results show trehalose inhibits malignant melanoma cell growth by inducing G2/M cell cycle arrest and apoptosis, suggesting trehalose as a potential candidate for a topical agent to inhibit proliferation of malignant tumor cells of the ocular surface.

Download Full-text