Reversion of vegetation following the cessation of fertilizer application

Abstract. Tadham Moor in Somerset, England, is an exceptionally rich wetland site which has been mown for hay for many years, with stock grazing the aftermath, but with no history of any fertilizer use. A randomized blocks field experiment (1986–1989) was used to study the effects of five levels of nitrogen input treatments: 0 = control, 25, 50, 100 and 200 kg of N fertilizer per ha per yr. In Phase II of the experiment (1990–1993), each plot was split into two subplots. The allocated fertilizer treatment for the plot was continued in one, randomly selected, subplot but the treatment was discontinued in the other subplot. The experiment not only identified and quantified the changes occurring in the vegetation of hay meadows under different levels of N input, it also provided valuable insight into the dynamics of the sward upon the discontinuance of the treatments. The data for Phase II were used to estimate the time required by the changed vegetation (under different nitrogen treatments) to revert to a state comparable to that prevailing in the control plots. A method for estimating reversion times is described. The main difficulties in estimating the reversion times are identified, the choice of robust vegetation variables being critical. Reversion time estimation methods are presented and used to obtain working estimates for the four nitrogen treatments, applied for 5 yr. These estimates are 3, 5, 7 and 9 yr respectively. The validity of the estimates of 3 yr for the lowest nitrogen input treatment (25 kg /ha/yr) was checked using the available post cessation data.     

In Vitro Analysis of Breast Cancer Cell Line Tumourspheres and Primary Human Breast Epithelia Mammospheres Demonstrates Inter- and Intrasphere Heterogeneity

Mammosphere and breast tumoursphere culture have gained popularity as in vitro assays for propagating and analysing normal and cancer stem cells. Whether the spheres derived from different sources or parent cultures themselves are indeed single entities enriched in stem/progenitor cells compared to other culture formats has not been fully determined. We surveyed sphere-forming capacity across 26 breast cell lines, immunophenotyped spheres from six luminal- and basal-like lines by immunohistochemistry and flow cytometry and compared clonogenicity between sphere, adherent and matrigel culture formats using in vitro functional assays. Analyses revealed morphological and molecular intra- and inter-sphere heterogeneity, consistent with adherent parental cell line phenotypes. Flow cytometry showed sphere culture does not universally enrich for markers previously associated with stem cell phenotypes, although we found some cell-line specific changes between sphere and adherent formats. Sphere-forming efficiency was significantly lower than adherent or matrigel clonogenicity and constant over serial passage. Surprisingly, self-renewal capacity of sphere-derived cells was similar/lower than other culture formats. We observed significant correlation between long-term-proliferating-cell symmetric division rates in sphere and adherent cultures, suggesting functional overlap between the compartments sustaining them. Experiments with normal primary human mammary epithelia, including sorted luminal (MUC1⁺) and basal/myoepithelial (CD10⁺) cells revealed distinct luminal-like, basal-like and mesenchymal entities amongst primary mammospheres. Morphological and colony-forming-cell assay data suggested mammosphere culture may enrich for a luminal progenitor phenotype, or induce reversion/relaxation of the basal/mesenchymal in vitro selection occurring with adherent culture. Overall, cell line tumourspheres and primary mammospheres are not homogenous entities enriched for stem cells, suggesting a more cautious approach to interpreting data from these assays and careful consideration of its limitations. Sphere culture may represent an alternative 3-dimensional culture system which rather than universally ‘enriching’ for stem cells, has utility as one of a suite of functional assays that provide a read-out of progenitor activity.     

Mini review: Instrumentation for vibrational circular dichroism spectroscopy,still a role for dispersive instruments

Vibrational circular dichroism (VCD) has become a standard method for determination of absolute stereochemistry, particularly now that reliable commercial instrumentation has become available. These instruments use a now well‐documented Fourier transform infrared‐based approach to measure VCD that has virtually displaced initial dispersive infrared‐based designs. Nonetheless, many papers have appeared reporting dispersive VCD data, especially for biopolymers. Instrumentation designed with these original methods, particularly after more recent updates optimizing performance in selected spectral regions, has been shown still to have advantages for specific applications. This article presents a mini‐review of dispersive VCD instrument designs and includes sample spectra obtained for various biopolymer (particularly peptide) samples. Complementary reviews of Fourier transform‐VCD designs are broadly available.     

Gene expression profiles of mouse submandibular gland development: FGFR1 regulates branching morphogenesis in vitro through BMP- and FGF-dependent mechanisms

Analyses of gene expression profiles at five different stages of mouse submandibular salivary gland development provide insight into gland organogenesis and identify genes that may be critical at different stages. Genes with similar expression profiles were clustered, and RT-PCR was used to confirm the developmental changes. We focused on fibroblast growth factor receptor 1 (FGFR1), as its expression is highest early in gland development. We extended our array results and analyzed the developmental expression patterns of other FGFR and FGF isoforms. The functional significance of FGFR1 was confirmed by submandibular gland organ culture. Antisense oligonucleotides decreased expression of FGFR1 and reduced branching morphogenesis of the glands. Inhibiting FGFR1 signaling with SU5402, a FGFR1 tyrosine kinase inhibitor, reduced branching morphogenesis. SU5402 treatment decreased cell proliferation but did not increase apoptosis. Fgfr, Fgf and Bmp gene expression was localized to either the mesenchyme or the epithelium by PCR, and then measured over time by real time PCR after SU5402 treatment. FGFR1 signaling regulates Fgfr1, Fgf1, Fgf3 and Bmp7 expression and indirectly regulates Fgf7, Fgf10 and Bmp4. Exogenous FGFs and BMPs added to glands in culture reveal distinct effects on gland morphology. Glands cultured with SU5402 were then rescued with exogenous BMP7, FGF7 or FGF10. Taken together, our results suggest specific FGFs and BMPs play reciprocal roles in regulating branching morphogenesis and FGFR1 signaling plays a central role by regulating both FGF and BMP expression.