Qibin Fu

Dynamics between cancer cell subpopulations reveals a model coordinating with both hierarchical and stochastic concepts.

Tumors are often heterogeneous in which tumor cells of different phenotypes have distinct properties. For scientific and clinical interests, it is of fundamental importance to understand their properties and the dynamic variations among different phenotypes, specifically under radio- and/or chemo-therapy. Currently there are two controversial models describing tumor heterogeneity, the cancer stem cell (CSC) model and the stochastic model. To clarify the controversy, we measured probabilities of different division types and transitions of cells via in situ immunofluorescence. Based on the experiment data, we constructed a model that combines the CSC with the stochastic concepts, showing the existence of both distinctive CSC subpopulations and the stochastic transitions from NSCCs to CSCs. The results showed that the dynamic variations between CSCs and non-stem cancer cells (NSCCs) can be simulated with the model. Further studies also showed that the model can be used to describe the dynamics of the two subpopulations after radiation treatment. More importantly, analysis demonstrated that the experimental detectable equilibrium CSC proportion can be achieved only when the stochastic transitions from NSCCs to CSCs occur, indicating that tumor heterogeneity may exist in a model coordinating with both the CSC and the stochastic concepts. The mathematic model based on experimental parameters may contribute to a better understanding of the tumor heterogeneity, and provide references on the dynamics of CSC subpopulation during radiotherapy.

Atomic Layer Deposition Modified Track-Etched Conical Nanochannels for Protein Sensing

Nanopore-based devices have recently become popular tools to detect biomolecules at the single-molecule level. Unlike the long-chain nucleic acids, protein molecules are still quite challenging to detect, since the protein molecules are much smaller in size and usually travel too fast through the nanopore with poor signal-to-noise ratio of the induced transport signals. In this work, we demonstrate a new type of nanopore device based on atomic layer deposition (ALD) Al2O3 modified track-etched conical nanochannels for protein sensing. These devices show very promising properties of high protein (bovine serum albumin) capture rate with well time-resolved transport signals and excellent signal-to-noise ratio for the transport events. Also, a special mechanism involving transient process of ion redistribution inside the nanochannel is proposed to explain the unusual biphasic waveshapes of the current change induced by the protein transport.

Impact of cell dissociation on identification of breast cancer stem cells

BACKGROUND Fluorescence-activated cell sorting was commonly used for identification of cancer stem cells (CSCs), which relied on specific cell surface markers. And this approach makes it possible for us to study characteristics of CSCs in vitro. However, the pattern of membrane protein including surface makers might be vitally influenced during the dissociation of the adherent cells, thus it might heavily impact the quantity and quality of CSCs identified by flow cytometry. METHODS To address this question, in present study, three commonly used digestive reagents and two different temperatures were performed in MCF-7 cells to assay CD44(+)CD24(-) CSCs subpopulation. The potential of sorted CD44(+)CD24(-) cells from different digestion to form mammosphere in culture was also compared. RESULTS The results showed that trypsin, a commonly used reagent in CSCs studies, most aggressively reduced antigenicity for surface markers and make part of CD44(+)CD24(-) CSCs subpopulation cleaved into CD44(+)CD24(-) non-stem cancer cells (NSCCs). And it also increased the mammosphere formation efficiency of CD44(-)CD24(-) subpopulation. This cleavage effect is especially serious when cells are digested at 37°C. While accutase, a purified collagenase/neutral protease cocktail, provides the best balance of dissociation efficiency and antigen retention. CONCLUSION Taken together; these results indicate that enzymatic digestion process plays an important role in identification of CSCs with surface marker via flow cytometer, suggesting that researchers need to reconsider this process seriously.

Rescue of Targeted Nonstem-Like Cells from Bystander Stem-Like Cells in Human Fibrosarcoma HT1080

Cancer stem-like cells (CSCs) have been suggested to be the principal cause of tumor radioresistance, dormancy and recurrence after radiotherapy. However, little is known about CSC behavior in response to clinical radiotherapy, particularly with regard to CSC communication with bulk cancer cells. In this study, CSCs and nonstem-like cancer cells (NSCCs) were co-cultured, and defined cell types were chosen and irradiated, respectively, with proton microbeam. The bidirectional rescue effect in the combinations of the two cell types was then investigated. The results showed that out of all four combinations, only the targeted, proton irradiated NSCCs were protected by bystander CSCs and showed less accumulation of 53BP1, which is a widely used indicator for DNA double-strand breaks. In addition, supplementation with c-PTIO, a specific nitric oxide scavenger, can show a similar effect on targeted NSCCs. These results, showed that the rescue effect of CSCs on targeted NSCCs involves nitric oxide in the process, suggesting that the cellular communication between CSCs and NSCCs may be important in determining the survival of tumor cells after radiation therapy. To our knowledge, this is the first report demonstrating a rescue effect of CSCs to irradiated NSCCs that may help us better understand CSC behavior in response to cancer radiotherapy.

Target irradiation induced bystander effects between stem-like and non stem-like cancer cells

Tumors are heterogeneous in nature and consist of multiple cell types. Among them, cancer stem-like cells (CSCs) are suggested to be the principal cause of tumor metastasis, resistance and recurrence. Therefore, understanding the behavior of CSCs in direct and indirect irradiations is crucial for clinical radiotherapy. Here, the CSCs and their counterpart non stem-like cancer cells (NSCCs) in human HT1080 fibrosarcoma cell line were sorted and labeled, then the two cell subtypes were mixed together and chosen separately to be irradiated via a proton microbeam. The radiation-induced bystander effect (RIBE) between the CSCs and NSCCs was measured by imaging 53BP1 foci, a widely used indicator for DNA double strand break (DSB). CSCs were found to be less active than NSCCs in both the generation and the response of bystander signals. Moreover, the nitric oxide (NO) scavenger c-PTIO can effectively alleviate the bystander effect in bystander NSCCs but not in bystander CSCs, indicating a difference of the two cell subtypes in NO signal response. To our knowledge, this is the first report shedding light on the RIBE between CSCs and NSCCs, which might contribute to a further understanding of the out-of-field effect in cancer radiotherapy.

Association of elevated reactive oxygen species and hyperthermia induced radiosensitivity in cancer stem-like cells

Cancer stem-like cells (CSCs) are the principal causes of tumor radio-resistance, dormancy and recurrence after radiotherapy. Clinical trials show hyperthermia (HT) might be a potent radiation sensitizer. In this study, CSCs were found to be more susceptible to radiation when combined with HT treatment. Treated cells showed significantly reduced self-renewal, cell survival and proliferation in vitro, as well as significant reduced tumor formation in vivo. Further study demonstrated that the radiosensitization effect was associated with increased intracellular reactive oxygen species (ROS) level in CSCs, confirmed by modifying redox status in CSCs bidirectionally. Pharmacologic depletion of glutathione by buthionine sulphoximine mimicked HT induced radiosensitivity in CSCs. Antioxidant N-acetylcysteine could efficiently rescue HT induced radiosensitivity in CSCs. To our knowledge, this may be the first report suggesting the association between elevated intracellular ROS level and HT induced radiosensitization in human breast CSCs and pancreatic CSCs, which might provide new strategy for improving CSCs radiosensitivity.

Relative biological effectiveness for photons: implication of complex DNA double-strand breaks as critical lesions

Current knowledge in radiobiology ascribes the adverse biological effects of ionizing radiation primarily to the induction of DNA double-strand breaks (DSBs), which is supposed to be potentially lethal and may be converted to lethal damage due to misrepair. Soft and ultrasoft x-rays have been found to bear elevated biological effectiveness for cell killing compared with conventional x-rays or 60Co γ-rays. This phenomenon is qualitatively interpreted as the increased level of DSB induction for low energy photons, however, a thorough quantitative reasoning is lacking. Here, we systematically compared the relative biological effectiveness (RBE) with relative DSB induction for photons from several hundreds of eV up to MeV. Although there is an approximate two-fold increase in the yields of DSB for low energy photons found in our calculation and a large number of experimental measurements, it is far from enough to account for the three- to four-fold increase in RBE. Further theoretical investigations show that DSB complexity (additional single-strand breaks and base damage within 10 base pairs) increases notably for low energy photons, which largely reconciles the discrepancy between RBE and DSB induction. Our theoretical results are in line with accumulating experimental evidence that complex DSBs are refractory to repair machinery and may contribute predominantly to the formation of lethal damage.