Holoclone Forming Cells from Pancreatic Cancer Cells Enrich Tumor Initiating Cells and Represent a Novel Model for Study of Cancer Stem Cells

Pancreatic cancer is one of the direct causes of cancer-related death. High level of chemoresistance is one of the major obstacles of clinical treatment. In recent years, cancer stem cells have been widely identified and indicated as the origin of chemoresistance in multi-types of solid tumors. Increasing evidences suggest that cancer stem cells reside in the cells capable of forming holoclones continuously. However, in pancreatic cancer, holoclone-forming cells have not been characterized yet. Therefore, the goal of our present study was to indentify the holoclone-forming pancreatic cancer stem cells and develop an in vitro continuous colony formation system, which will greatly facilitate the study of pancreatic cancer stem cells.Pancreatic cancer cell line BxPC3 was submitted to monoclonal cultivation to generate colonies. Based on the morphologies, colonies were classified and analyzed for their capacities of secondary colony formation, long-term survival in vitro, tumor formation in vivo, and drug resistance. Flowcytometry and quantitative RT-PCR were performed to detect the expression level of cancer stem cells associated cell surface markers, regulatory genes and microRNAs in distinct types of colonies. Three types of colonies with distinct morphologies were identified and termed as holo-, mero-, and paraclones, in which only holoclones generated descendant colonies of all three types in further passages. Compared to mero- and paraclones, holoclones possessed higher capacities of long-term survival, tumor initiation, and chemoresistance. The preferential expression of cancer stem cells related marker (CXCR4), regulatory genes (BMI1, GLI1, and GLI2) and microRNAs (miR-214, miR-21, miR-221, miR-222 and miR-155) in holoclones were also highlighted.Our results indicate that the pancreatic tumor-initiating cells with high level of chemoresistance were enriched in holoclones derived from BxPC3 cell line. Generation of holoclones can serve as a novel model for studying cancer stem cells, and attribute to developing new anti-cancer drugs

Combined immune checkpoint protein blockade and low dose whole body irradiation as immunotherapy for myeloma

BACKGROUND: Multiple myeloma is characterized by the presence of transformed neoplastic plasma cells in the bone marrow and is generally considered to be an incurable disease. Successful treatments will likely require multi-faceted approaches incorporating conventional drug therapies, immunotherapy and other novel treatments. Our lab previously showed that a combination of transient lymphodepletion (sublethal whole body irradiation) and PD-1/PD-L1 blockade generated anti-myeloma T cell reactivity capable of eliminating established disease. We hypothesized that blocking a combination of checkpoint receptors in the context of low-dose, lymphodepleting whole body radiation would boost anti-tumor immunity. METHODS: To test our central hypothesis, we utilized a 5T33 murine multiple myeloma model. Myeloma-bearing mice were treated with a low dose of whole body irradiation and combinations of blocking antibodies to PD-L1, LAG-3, TIM-3, CD48 (the ligand for 2B4) and CTLA4. RESULTS: Temporal phenotypic analysis of bone marrow from myeloma-bearing mice demonstrated that elevated percentages of PD-1, 2B4, LAG-3 and TIM-3 proteins were expressed on T cells. When PD-L1 blockade was combined with blocking antibodies to LAG-3, TIM-3 or CTLA4, synergistic or additive increases in survival were observed (survival rates improved from ~30% to >80%). The increased survival rates correlated with increased frequencies of tumor-reactive CD8 and CD4 T cells. When stimulated in vitro with myeloma cells, CD8 T cells from treated mice produced elevated levels proinflammatory cytokines. Cytokines were spontaneously released from CD4 T cells isolated from mice treated with PD-L1 plus CTLA4 blocking antibodies. CONCLUSIONS: These data indicate that blocking PD-1/PD-L1 interactions in conjunction with other immune checkpoint proteins provides synergistic anti-tumor efficacy following lymphodepletive doses of whole body irradiation. This strategy is a promising combination strategy for myeloma and other hematologic malignancies. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s40425-014-0043-z) contains supplementary material, which is available to authorized users

Watershed Urbanization Linked to Differences in Stream Bacterial Community Composition

Urbanization strongly influences headwater stream chemistry and hydrology, but little is known about how these conditions impact bacterial community composition. We predicted that urbanization would impact bacterial community composition, but that stream water column bacterial communities would be most strongly linked to urbanization at a watershed-scale, as measured by impervious cover, while sediment bacterial communities would correlate with environmental conditions at the scale of stream reaches. To test this hypothesis, we determined bacterial community composition in the water column and sediment of headwater streams located across a gradient of watershed impervious cover using high-throughput 16S rRNA gene amplicon sequencing. Alpha diversity metrics did not show a strong response to catchment urbanization, but beta diversity was significantly related to watershed impervious cover with significant differences also found between water column and sediment samples. Samples grouped primarily according to habitat—water column vs. sediment—with a significant response to watershed impervious cover nested within each habitat type. Compositional shifts for communities in urbanized streams indicated an increase in taxa associated with human activity including bacteria from the genus Polynucleobacter, which is widespread, but has been associated with eutrophic conditions in larger water bodies. Another indicator of communities in urbanized streams was an OTU from the genus Gallionella, which is linked to corrosion of water distribution systems. To identify changes in bacterial community interactions, bacterial co-occurrence networks were generated from urban and forested samples. The urbanized co-occurrence network was much smaller and had fewer co-occurrence events per taxon than forested equivalents, indicating a loss of keystone taxa with urbanization. Our results suggest that urbanization has significant impacts on the community composition of headwater streams, and suggest that processes driving these changes in urbanized water column vs. sediment environments are distinct

A System level analysis of ecosystem responses to hurricane impacts on a coastal region.

Tropical cyclones are major disturbances for coastal systems. Hurricane Harvey made landfall in Texas, USA, on August 25, 2017 as a category 4 storm. There were two distinct disturbances associated with this storm that were spatially decoupled: (1) high winds causing direct damage and storm surge, and (2) high rains causing scouring floods and significant discharge of fresh water carrying carbon and nutrients to estuaries. Here, we provide a synthesis of the effects of Hurricane Harvey on biogeochemical, hydrographic, and biotic components of freshwater and estuarine systems and their comparative resistance and resilience to wind- and rain-driven disturbances. Wind-driven disturbances were most severe along the coastal barrier islands and lower estuaries, damaging mangroves and seagrass and increasing sediment coarseness. Rain-driven disturbances were most pronounced within freshwater streams and the upper estuaries. Large volumes of freshwater run-off reduced the abundance of riverine fauna and caused hypoxic and hyposaline conditions in the estuaries for over a week. In response to this freshwater input event, benthic fauna diversity and abundance decreased, but mobile fauna such as estuarine fishes did not markedly change. Although hydrographic and biogeochemical components were highly perturbed, they returned to baseline conditions within days. In contrast, biotic components demonstrated lower magnitude changes, but some of these organisms, particularly the sedentary flora and fauna, required weeks to months to return to pre-storm conditions, and some did not recover within the 6 months reported here. Our synthesis illustrates that resistance and resilience of system components may negatively co-vary and that structural components of coastal systems may be the most vulnerable to long-term changes following tropical cyclones