EFFECTS OF ENVIRONMENTAL ANTIOXIDANTS ON PRO-ANGIOGENIC ROS SIGNALLING IN ENDOTHELIAL COLONY FORMING CELLS IN VITRO

Introduction. Endothelial colony-forming cells (ECFCs) hold great cytotherapeutic potential for ischaemic disease. Whilst increasing evidence supports a key role for reactive oxygen species (ROS), specifically those derived from Nox NADPH oxidases, in the underlying angiogenic processes of these and other endothelial cells, such studies investigating the role of redox signalling may be hampered by the standard inclusion of antioxidant agents in endothelial cell media, such as phenol red. Aims. To study the effects of antioxidants present in culture media on pro-angiogenic function of ECFCs in vitro. Methods. Human ECFCs isolated from umbilical cord blood were maintained in media with and without antioxidant components (EGM2 and phenol red-free DMEM, respectively) prior to treatment with pro-oxidant PMA and assessment of their in vitro migratory capacity using a scratch-wound assay to measure pro-angiogenic activity. Results. Our previous work in our group indicated that PMA (500nM) increased ECFC migration in a both a superoxide and NADPH oxidase-dependent manner (control 18.6±2.8, PMA 32.7±6.6% wound closure; n=6, P<0.05), as indicated by attenuation with PEG-SOD and VAS2870. However, inconsistencies in the data generated under varying experimental conditions led us to hypothesise that antioxidant agents in the standard ECFC media may be influencing these effects. Indeed, a direct comparison of cell migration between ECFCs incubated in EGM2 DMEM demonstrated a clear trend towards higher migration in the latter (EGM2 9.0±4.5, DMEM 22.7±6.4%; n=3, P=NS). Similar to our previous EGM2 studies, cell migration was potentiated by PMA (control 11.6±1.6, PMA 25.1±2.8%; n=3, P<0.05), but at a lower dose (100nM), which is consistent with a reduction in media antioxidants. Notably, this response was attenuated by VAS2870 (PMA 37.6±7.3, PMA+VAS2870 10.3±2.9%; n=6, P<0.05), underlining a likely role for Nox NADPH oxidases. Conclusion. Taken together, these data indicate that ECFC migration is sensitive to different endothelial cell growth media, which appears to be dependent upon their antioxidant content. Although further experiments, such as quantification of cellular superoxide generation by dihydroethidium fluorescence may be required to confirm a specific role for antioxidants, such blunting of ROS signalling in vitro is clearly an important consideration which may significantly impact upon data interpretation.

Lake Kumphawapi revisited – The complex climatic and environmental record of a tropical wetland in NE Thailand

Kumphawapi, which is Thailand’s largest natural freshwater lake, contains a >10,000-year-long climatic and environmental archive. New data sets (stratigraphy, chronology, hydrogen isotopes, plant macrofossil and charcoal records) for two sedimentary sequences are here combined with earlier multi-proxy studies to provide a comprehensive reconstruction of past climatic and environmental changes for Northeast Thailand. Gradually higher moisture availability due to a strengthening of the summer monsoon led to the formation of a large shallow lake in the Kumphawapi basin between >10,700 and c. 7000 cal. BP. The marked increase in moisture availability and lower evaporation between c. 7000 and 6400 cal. BP favoured the growth and expansion of vegetation in and around the shallow lake. The increase in biomass led to gradual overgrowing and infilling, to an apparent lake level lowering and to the development of a wetland. Multiple hiatuses are apparent in all investigated sequences between c. 6500 and 1400 cal. BP and are explained by periodic desiccation events of the wetland and erosion due to the subsequent lake level rise. The rise in lake level, which started c. 2000 cal. BP and reached shallower parts c. 1400 cal. BP, is attributed to an increase in effective moisture availability. The timing of hydroclimatic conditions during the past 2000 years cannot be resolved because of chronological limitations.

Stone temperature and moisture variability under temperate environmental conditions: implications for sandstone weathering

Temperature and moisture conditions are key drivers of stone weathering processes in both natural and built environments. Given their importance in the breakdown of stone, a detailed understanding of their temporal and spatial variability is central to understanding present-day weathering behaviour and for predicting how climate change may influence the nature and rates of future stone decay.
Subsurface temperature and moisture data are reported from quarry fresh Peakmoor Sandstone samples exposed during summer (June–July) and late autumn / early winter (October–December) in a mid-latitude, temperate maritime environment. These data demonstrate that the subsurface thermal response of sandstone comprises numerous short-term (minutes), low magnitude fluctuations superimposed upon larger-scale diurnal heating and cooling cycles with distinct aspect-related differences. The short-term fluctuations create conditions in the outer 5–10 mm of stone that are much more ‘energetic’ in comparison to the more subdued thermal cycling that occurs deeper within the sandstone samples.
Data show that moisture dynamics are equally complex with a near-surface region (5–10 mm) in which frequent moisture cycling takes place and this, combined with the thermal dynamism exhibited by the same region may have significant implications for the nature and rate of weathering activity. Data indicate that moisture input from rainfall, particularly when it is wind-driven, can travel deep into the stone where it can prolong the time of wetness. This most often occurs during wetter winter months when moisture input is high and evaporative loss is low but can happen at any time during the year when the hydraulic connection between near-surface and deeper regions of the stone is disrupted with subsequent loss of moisture from depth slowing as it becomes reliant on vapour diffusion alone.
These data illustrate the complexity of temperature and moisture conditions in sandstone exposed to the ‘moderate’ conditions of a temperate maritime environment. They highlight differences in thermal and moisture cycling between near-surface (5–10 mm) and deeper regions within the stone and contribute towards a better understanding of the development of structural and mineralogical heterogeneity between the stone surface and substrate.