Humans ; Laryngeal Neoplasms ; pathology ; Neoplastic Stem Cells ; cytology

At present, most patients with hematological malignancies are treated with conventional chemotherapy, but the relapse frequently occurs in these patients. It is increasingly recognized that the cause of relapse resides in a small portion of the leukemia stem/progenitor cell population which has self-renewal and multilineage differentiation potential. As traditional chemotherapeutic drugs only kill the majority of differentiated tumor cells and hardly affect the tumor stem/progenitor cell, that is why most of the chemotherapies did not achieve good results. In order to completely eradicate hematological malignancies, intrinsic properties of stem/progenitor cells must be studied and only the therapy targeting this population could make tumor curable. Targeting the surface markers which distinguish the tumor stem/progenitor cells from normal stem/progenitor cells, inducing the tumor stem/progenitor cell differentiation, disrupting the signal pathways and niche which regulate the tumor stem/progenitor cell self-renewal are all probable strategies. This review summarizes recent progress of research on leukemia stem/progenitor cell targeting therapy development.
Hematologic Neoplasms ; therapy ; Humans ; Neoplastic Stem Cells ; cytology

Stem/progenitor cells contribute to normal development of organs, but their mutation may lead to many human diseases including cancer. Stem/progenitor cells and some of the cancer cells have the same abilities of self-renewal and differentiation, suggesting that tumors were initiated from mutated stem/progenitor cells, the cancer stem/progenitor cells. The recent studies on the origin of the non-Hodgkin's lymphomas has proved that the second-hit aggravates gene mutation in lymphocytic progenitor cells, leading to the generation of lymphoma. This review summarizes the current advances in the studies of the second-hit at stem/progenitor cells in oncogenesis of B-cell non-Hodgkin's lymphoma.
Humans ; Leukemia ; etiology ; Lymphoma ; etiology ; Mutation ; Neoplastic Stem Cells ; cytology

MicroRNAs (miRNAs) are small non-coding RNAs that regulate messenger RNAs at the post-transcriptional level. They play an important role in the control of cell physiological functions, and their alterations have been related to cancer, where they can function as oncogenes or tumor suppressor genes. Recently, they have emerged as key regulators of "stemness", collaborating in the maintenance of pluripotency, control of self-renewal, and differen-tiation of stem cells. The miRNA pathway has been shown to be crucial in embryonic development and in embryonic stem (ES) cells, as shown by Dicer knockout analysis. Specific patterns of miRNAs have been reported to be expressed only in ES cells and in early phases of embryonic development. Moreover, many cancers present small populations of cells with stem cell characteristics, called cancer stem cells (CSCs). CSCs are responsible for relapse and treatment failure in many cancer patients, and the comparative analysis of expression patterns between ES cells and tumors can lead to the identification of a miRNA signature to define CSCs. Most of the key miRNAs identified to date in ES cells have been shown to play a role in tumor diagnosis or prognosis, and may well prove to be essential in cancer therapy in the foreseeable future.
Embryonic Stem Cells/cytology/*metabolism ; Humans ; MicroRNAs/genetics/*metabolism ; Models, Biological ; Neoplastic Stem Cells/cytology/*metabolism ; Signal Transduction/genetics/*physiology

As pioneer of tumor stem cell research, leukemia stem cell research has not only important theoretical significance, but also clinical application potential. The survival and development of stem cells are directly impacted by their microenvironment. The research on leukemia stem cells and their microenvironment are now becoming a hot topic. The author presumes that stem cells are a population with heterogenecity and hierarchy; any single cell from the population is difficult to form a clone; the interaction between the leukemia stem cell and its microenvironment can be described by the concept of leukemia stem cell niche. In this article, the leukemia cell population with heterogenecity and hierarchy as well as leukemia stem cell niche were summarized and discussed.
Cell Line, Tumor ; Humans ; Leukemia ; genetics ; pathology ; Neoplastic Stem Cells ; metabolism ; pathology ; Stem Cell Niche ; cytology ; Stromal Cells ; cytology ; immunology

Cancer stem cells are defined as rare cells in cancer tissues with indefinite potential for self-renewal that drives tumorigenesis. It was first extensively documented for leukaemia and multiple myeloma. It has also been found in solid cancers such as human breast cancer and nervous system tumors. Studies of cancer stem cell biology and mechanisms of tumorigenesis are lending insight into the origins of cancer and will ultimately yield new approaches to fight cancer.
Animals ; Cell Transformation, Neoplastic ; Humans ; Mice ; Neoplasms ; pathology ; therapy ; Neoplastic Stem Cells ; cytology ; Tumor Stem Cell Assay ; methods

OBJECTIVETo sort and identify side population (SP) cancer stem cells (CSC) in human prostate cancer (PCa) cell lines.

METHODSStem-like cells were isolated from five PCa cell lines Du145, IA8, LNCaP, TSU-Pr and PC-3 using FACS based on CD133+ CD44+ immunophenotype and SP in Hoechst staining. The in vitro growth pattern and tumorigenicity of SP stem cells were verified by soft agar colony-formation trial. LNCaP/SP cells were selected for further identification of stem cell properties using immunostaining, proliferation and invasion assay. Eventually, tumorigenicity and metastasis ability of LNCaP/SP were confirmed by xenograft experiments.

RESULTSThe percentages of CSCs of the CD133 CD44 + immunophenotype were extremely low in the five PCa cell lines. On the contrary, the percentages of the isolated SP cells were significantly higher in Du145 ([0.15 +/- 0.02]%), IA8 ([0.60 +/- 0.07 ]%), LNCaP ([0.8 +/- 0.1]%) and TSU-PrL ([2.0 +/- 0.4]%), but none was detected in PC-3. Besides, IA8/SP, LNCaP/SP and TSU-PrL/SP cells showed a significantly greater colony-forming efficiency than non-side population (NSP) cells (P < 0.05). Compared with LNCaP/NSP cells, LNCaP/SP cells exhibited high expressions of integrin alpha2, Nanog, CD44, OCT4 and ABCG2, remarkably enhanced invasive and proliferative potentials in vitro, and markedly increased tumorigenicity and metastasis (P < 0.01).

CONCLUSIONSP sorting is more suitable than CD133+ CD44+ selection for enriching CSCs from PCa cell lines, and LNCaP/ SP represents a typical CSC population.

Cell Line, Tumor ; cytology ; Cell Separation ; Humans ; Male ; Neoplastic Stem Cells ; cytology ; Prostatic Neoplasms ; Side-Population Cells ; cytology

This study was aimed to sort the side population (SP) cells from human multiple myeloma cell lines, then detect the biological characteristics of those SP cells. After Hoechst33342 staining, intracellular Hoechst33342 fluorescence staining differences of myeloma cell lines observed by the fluorescence microscopy. The fluorescence-activated cell sorting (FACS) technology was used to isolate SP cells and main population (MP) cells; proliferative capacity in vitro was determined by cell growth curve; the cell colony forming ability was compared by colony forming test. The CD138 expression was detected by flow cytometry. The expression of ABCG2 mRNA was detected by reverse transcription PCR; CCK-8 assay and colony forming test were used to evaluate the effect of bortezomib on the cell proliferation, vitality and colony forming ability of the two populations. The results showed that the myeloma cell lines had a small proportion of SP cells, especially, RPMI 8226 cells accounted for the highest proportion of SP cells (7.10 ± 2.69)%, which have also been confirmed under the fluorescence microscope; the proliferative activity and cell colony forming ability of SP cells were significantly higher than those of MP cells (P < 0.05). The expression levels of CD138 in SP and MP cells were not significantly different (P > 0.05). RT-PCR results showed that SP cells expressed the drug-resistance gene ABCG2, but MP cells hardly express these genes. The inhibition rate of bortezomib on SP cells was significantly lower than that on MP cells (P < 0.05), however, the difference was not significant (P > 0.05) at bortezomib 40 nmol/L. Bortezomib could reduce colony formation in the both two cell populations, but more severe reduction appeared in the MP cells. It is concluded that the myeloma cell line contain a small amount of SP cells with the cancer stem cell characteristics.
Cell Line, Tumor ; Cytological Techniques ; methods ; Humans ; Multiple Myeloma ; Neoplastic Stem Cells ; cytology ; Side-Population Cells ; cytology

OBJECTIVE: To investigate the optimizing conditions in isolation of the side population in laryngeal carcinoma cell line Hep-2. METHOD: Single-cell suspension cells were detached from the culture flask with trypsin EDTA, at a concentration of 1 x 10(6) cells/ml. (1) The trail Samples were incubated with Hoechst33342 at a concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml for 90 minutes. (2) They were incubated with Hoechst for 50, 70, 90, 110, 130 min in water bath individually. (3) The single-cell suspension were incubated Hoechst in water bath and in thermostat each. (4) The two different density of cells were harvested, which were 100% and 70%, and then di gest into single-cell suspension. Once incubation finished, suspended in phosphate buffered saline (PBS), then test SP% by flow cytometry. Among all groups,Verapamil hydrochloride was added to the control samples, incubated at 37 degrees C for 30 minutes, the other condition were keep the same with their trial groups. RESULT: (1) The percentage of Hoechst-negative cells in trial group was (39.96 +/- 0.24)%, (26.23 +/- 0.39)%. (18.79 +/- 0.02)%, (19.01 +/- 0.14)% at the concentration of 5 microg/ml, 9 microg/ml, 10 microg/ml, 11 microg/ml respectively, when the PI-positive cells were (30.45 +/- 0.63)%, (49.9 +/- 0.42)%, (50.12 +/- 0.68)%, (64.16 +/- 0.39)% separately. (2) Varying the duration of staining incubation showed that there was a typical FACS pattern and SP% was constant when the incubation was at least 90 min. (3) Compare to water bath, SP% was more than in thermostat, the SP% was (18.67 +/- 0.45)%, (22.6 +/- 0.50)% respectively; (4) Cell density is also responsible for SP%. The low density the cell is, the less in SP%. SPSS13.0 was used in statistical analysis, the groups were compared using t-Test. P < 0.05 was considered statistically significant. CONCLUSION The optimum concentration and duration of incubation of Hoechst33342 in isolation of the side population cells in laryngeal carcinoma cell line Hep-2 is 10 microg/ml and 90 min. Incubated in water bath is better than in thermostat. The best staining cell density is around 80%-90%.
Cell Count ; Cell Line, Tumor ; Flow Cytometry ; Humans ; Laryngeal Neoplasms ; pathology ; Neoplastic Stem Cells ; cytology ; Side-Population Cells ; cytology

OBJECTIVETo develop an effective method for isolating and culturing single cancer stem cells.

METHODSThe capillary glass tube was stretched on fire and connected to a sterile plastic tube to prepare the single cell separation apparatus. Single SW480 cell clone spheres in serum-free culture were marked with CD133 and CK7, and the single cancer stem cells were separated and cultivated in 96-well plates or microdrop covered by paraffin.

RESULTSSW480 cell clone formation rate was about 1.04%, and the cell clone spheres highly expressed CD133 with low CK7 expression. The isolation of the single cancer stem cells showed a success rate of 98.99% using the separation device. The cell division profile was comparable between the cell cultures in microdrop and 96-well plates in the initial 2 cell divisions (P>0.05), whereas prolonged cell division occurred afterwards in the microdrop culture as compared to 96-well plate culture. The cell population expansion of the single cancer stem cells was similar between microdrop culture (11.5%, 22/192) and 96-well plate culture (9.2%, 17/184) (P>0.05).

CONCLUSIONSSingle SW480 cells can develop into cancer stem cell spheres. Microdrop culture is convenient and stable, and can be the primary choice for single cancer stem cell culture.

Cell Culture Techniques ; methods ; Cell Line, Tumor ; Cell Separation ; methods ; Humans ; Neoplastic Stem Cells ; cytology