Multiple types of data are required to identify the mechanisms influencing the spatial expansion of melanoma cell colonies

Background The expansion of cell colonies is driven by a delicate balance of several mechanisms including cell motility, cell-to-cell adhesion and cell proliferation. New approaches that can be used to independently identify and quantify the role of each mechanism will help us understand how each mechanism contributes to the expansion process. Standard mathematical modelling approaches to describe such cell colony expansion typically neglect cell-to-cell adhesion, despite the fact that cell-to-cell adhesion is thought to play an important role. Results We use a combined experimental and mathematical modelling approach to determine the cell diffusivity, D, cell-to-cell adhesion strength, q, and cell proliferation rate, ?, in an expanding colony of MM127 melanoma cells. Using a circular barrier assay, we extract several types of experimental data and use a mathematical model to independently estimate D, q and ?. In our first set of experiments, we suppress cell proliferation and analyse three different types of data to estimate D and q. We find that standard types of data, such as the area enclosed by the leading edge of the expanding colony and more detailed cell density profiles throughout the expanding colony, does not provide sufficient information to uniquely identify D and q. We find that additional data relating to the degree of cell-to-cell clustering is required to provide independent estimates of q, and in turn D. In our second set of experiments, where proliferation is not suppressed, we use data describing temporal changes in cell density to determine the cell proliferation rate. In summary, we find that our experiments are best described using the range D = 161 - 243 ?m2 hour-1, q = 0.3 - 0.5 (low to moderate strength) and ? = 0.0305 - 0.0398 hour-1, and with these parameters we can accurately predict the temporal variations in the spatial extent and cell density profile throughout the expanding melanoma cell colony. Conclusions Our systematic approach to identify the cell diffusivity, cell-to-cell adhesion strength and cell proliferation rate highlights the importance of integrating multiple types of data to accurately quantify the factors influencing the spatial expansion of melanoma cell colonies.

Packed bed bioreactor for the isolation and expansion of placental-derived Mesenchymal Stromal Cells

Large numbers of Mesenchymal stem/stromal cells (MSCs) are required for clinical relevant doses to treat a number of diseases. To economically manufacture these MSCs, an automated bioreactor system will be required. Herein we describe the development of a scalable closed-system, packed bed bioreactor suitable for large-scale MSCs expansion. The packed bed was formed from fused polystyrene pellets that were air plasma treated to endow them with a surface chemistry similar to traditional tissue culture plastic. The packed bed was encased within a gas permeable shell to decouple the medium nutrient supply and gas exchange. This enabled a significant reduction in medium flow rates, thus reducing shear and even facilitating single pass medium exchange. The system was optimised in a small-scale bioreactor format (160 cm2) with murine-derived green fluorescent protein-expressing MSCs, and then scaled-up to a 2800 cm2 format. We demonstrated that placental derived MSCs could be isolated directly within the bioreactor and subsequently expanded. Our results demonstrate that the closed system large-scale packed bed bioreactor is an effective and scalable tool for large-scale isolation and expansion of MSCs.

Implant removal by means of an expansion of the alveolar bony crest: Report of a clinical technique

Aim: To describe the adaptation of the Edentulous Ridge Expansion (E.R.E.) technique for implant removal. Material and Methods: The E.R.E. technique for the removal of failed implants is described in detail. A clinical case is also reported. In a patient carrying a full arch removable prosthesis in the upper jaw, sustained by two bars, two out of five implants were found to be fractured. Bucco-lingual partial-thickness flaps were used to access the fractured implants. The implants were subsequently removed applying the E.R.E. technique. Two recipient sites were prepared in the same position, using bone expanders, and two new implants were installed. Results: After 4 months of healing, the implants were integrated and a new bar was fabricated, and the old prosthesis readapted. Conclusion: The ERE technique may be successfully applied for the removal of failed implants, and the immediate or delayed reinstallation of new implants. © 2012 John Wiley & Sons A/S.

Characterization and optimization of antigen-specific T cell responses during ex vivo expansion of melanoma tumor-infiltrating lymphocytes

Treatment of metastatic melanoma with tumor reactive T cells (adoptive T cell therapy, ACT) is a promising approach associated with a high clinical response rate. However, further optimization of this treatment modality is required to increase the clinical response after this therapy. ACT in melanoma involves an initial phase (pre-REP) of tumor-infiltrating lymphocyte (TIL) expansion ex vivo from tumor isolates followed by a second phase, “rapid expansion protocol” (REP) generating the billions of cells used as the TIL infusion product. The main question addressed in this thesis was how the currently used REP affected the responsiveness of the CD8+ T cells to defined melanoma antigens. We hypothesized that the REP drives the TIL to further differentiate and become hyporesponsive to antigen restimulation, therefore, proper cytokine treatment or other ways to expand TIL is required to improve upon this outcome. We evaluated the response of CD8+ TIL to melanoma antigen restimulation using MART-1 peptide-pulsed mature DC in vitro. Post-REP TILs were mostly hypo-responsive with poor proliferation and higher apoptosis. Phenotypic analysis revealed that the expression of CD28 was significantly reduced in post-REP TILs. By sorting experiment and microarray analysis, we confirmed that the few CD28+ post-REP TILs had superior survival capacity and proliferated after restimulation. We then went on to investigate methods to maintain CD28 expression during the REP and improve TIL responsiveness. Firstly, IL-15 and IL-21 were found to synergize in maintaining TIL CD28 expression and antigenic responsiveness during REP. Secondly, we found IL-15 was superior as compared to IL-2 in supporting the long-term expansion of antigen-specific CD8+ TIL after restimulation. These results suggest that current expansion protocols used for adoptive T-cell therapy in melanoma yield largely hyporesponsive products containing CD8+ T cells unable to respond in vivo to re-stimulation with antigen. A modification of our current approaches by using IL-15+IL-21 as supporting cytokines in the REP, or/and administration of IL-15 instead of IL-2 after TIL infusion, may enhance the anti-tumor efficacy and long-term persistence of infused T cells in vivo.

Further expansion of the alien seaweed Caulerpa taxifolia var. distichophylla (Sonder) Verlaque, Huisman & Procacini (Ulvophyceae, Bryopsidales) in the Eastern Mediterranean Sea

Date of Acceptance: 27/04/2015 We are grateful to Andreas Antoniou (Dep. of Environment, Ministry of Agriculture, Rural Development & Environment, Cyprus) for his assistance in the preparation of the illustrations. We would also like to thank Dr. Sotiris Orfanidis (NAGREF – Fisheries Research Institute, Kavala, Greece) for his valuable advice and both the DFMR and HSR / HCMR Rhodes crew and George Hatiris for their help in samplings. Special thanks are due to Dinos Leonidou (SeaQuest Divers Cyprus) for accompanying the deep dive for sampling Caulerpa at Cavo Greco. We are grateful to the Total Foundation (Paris) for its funding support to this study within the framework of the project “Brown algal ecology and biodiversity in the eastern Mediterranean Sea” and to the MASTS pooling initiative (Marine Alliance for Science and Technology for Scotland, funded by the Scottish Funding Council and contributing institutions; grant reference HR09011).

The selective expansion of agricultural production in Latin America;

"United Nations publication. Sales no.: 1957.II.G.4."

Electricity : an expansion of Everett's Deschanel, part III, on the lines of modern electrical theory /

Mode of access: Internet.

Thermal expansion of some glasses in the system Li₂0-Ca0-Si0₂ /

This investigation was carried on through the cooperation of the United States Atomic Energy Commission (Contract Number AT-(40-1)--1080, and the Department of Ceramic Technology of the University of Alabama.

Thermal expansion of some glasses in the system Li₂0-B₂0₃-Si0₂ /

"Work performed under Contract No. AT(40-1)-1080."

The coefficient of thermal expansion of alpha uranium as fabricated by various methods /

"U.S. Atomic Energy Commission Contract AT(29-1)-1106."

Preliminary test on axial spindle expansion of nuber 739 Ex-Cell-O numerical tape control machine /

"U.S. Atomic Energy Commission Contract AT(29-1)-1106."

Thermal expansion of zirconium and zirconium-tin alloys up to 570°C /

"Contract AT-30-1-Gen-366."

Expansion of gas clouds and hypersonic jets bounded by a vacuum.

"SSD-TDR-62-96. Report no. TDR-169 (3230-12) TR-1. Contract no. AF 04(695)-169."