The primary cilium is a ubiquitous cytoplasmic organelle of unknown function. Ultrastructural evidence of primary cilia in chondrocytes, and their colocalisation with the Golgi apparatus, has led to speculation that these structures are functionally linked. To investigate the relationship between these organelles, we examined the molecular anatomy of the microtubular cytoskeleton in the chondrocytes of chick embryo sterna. Thick cryosections were immunolabelled with antibodies directed against acetylated α-tubulin (C3B9), detyrosinated α-tubulin (ID5) and total α-tubulin (TAT), and imaged at high magnification using confocal laser scanning microscopy. Transmission electron microscopy confirmed the ultrastructure of the chondrocyte primary cilium and its structural relationship to the Golgi apparatus. Detyrosinated and acetylated α-tubulins were concentrated in the centrioles, centrosome and microtubule organising centre adjacent to the nucleus, with total α-tubulin distributed throughout the cytoplasm. ID5 stained the primary cilium at an incidence of 1 per cell, its colocalisation with C3B9 identifying the primary cilium as one of the most stable features of the microtubular cytoskeleton. Primary cilia varied from 1 to 4 µm in length, and 3 patterns of projection into the extracellular matrix were identified ; (1) full extension and matrix contact, with minor undulations along the length ; (2) partial extension and matrix contact, with a range of bending deflections ; (3) cilium reclined against the cell surface with minimal matrix contact. Ultrastructural studies identified direct connections between extracellular collagen fibres and the proteins which decorate ciliary microtubules, suggesting a matrix-cilium-Golgi continuum in hyaline chondrocytes. These results strengthen the hypothesis that the primary cilium acts as a ' cellular cybernetic probe ' capable of transducing environmental information from the extracellular matrix, communicating this information to the centrosome, and regulating the exocytosis of Golgi-derived secretory vesicles.

The composition and structural integrity of the pericellular microenvironment do influence the cellular response to experimental osmotic challenge. This suggests that the microenvironment functions in situ to mediate the chondrocyte response to physicochemical changes associated with joint loading.

No abstract

During insulin-dependent diabetes mellitus, immune cells infiltrate pancreatic islets progressively and mediate beta cell destruction over a prolonged asymptomatic prediabetic period. Apoptosis may be a major mechanism of beta cell loss during the disease. This process involves a proteolytic cascade in which upstream procaspases are activated which themselves activate downstream caspases, including caspase-3, a key enzyme involved in the terminal apoptotic cascade. Here dual-label immunohistochemistry was employed to examine the intra-islet expression, distribution and cellular sources of active caspase-3 in the non-obese diabetic (NOD) mouse given cyclophosphamide to accelerate diabetes. NOD mice were treated at day 95 and caspase-3 expression was studied at days 0, 4, 7, 11 and 14. Its expression was also correlated with advancing disease and compared with age-matched NOD mice treated with diluent alone. At day 0 (=day 95), caspase-3 immunolabelling was observed in several peri-islet and intra-islet macrophages, but not in CD4 and CD8 cells and only extremely rarely in beta cells. At day 4, only a few beta cells weakly expressed the enzyme, in the absence of significant insulitis. At day 7, caspase-3 expression was observed in a small proportion of intra-islet macrophages. At day 11, there was a marked increase in the number of intra-islet macrophages positive for caspase-3 while only a few CD4 cells expressed the enzyme. At day 14, caspase-3 labelling became prominent in a significant proportion of macrophages. Only a few CD4 and CD8 cells expressed the enzyme. Capase-3 labelling was also present in a proportion of macrophages in perivascular and exocrine regions. Surprisingly, beta cell labelling of caspase-3 at days 11 and 14 was rare. At this stage of heightened beta cell loss, a proportion of intra-islet interleukin-1beta-positive cells coexpressed the enzyme. Caspase-3 was also observed in numerous Fas-positive cells in heavily infiltrated islets. During this late stage, only a proportion of caspase-3-positive cells contained apoptotic nuclei, as judged by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL). We conclude that during cyclophosphamide-accelerated diabetes in the NOD mouse, the predominant immunolabelling of caspase-3 in intra-islet macrophages suggests that apoptosis of macrophages may be an important mechanism for its elimination. The virtual absence of caspase-3 immunolabelling in most beta cells even during heightened beta cell loss supports their rapid clearance following their death during insulin-dependent diabetes mellitus.

Today’s students have increased expectations for flexible learning options and evidence-based practice resources to be available to support curricular activities. We investigated: (i) the suitability of a static website for teaching ocular anatomy and physiology and an interactive version of the website with quiz and self-assessment activities and (ii) the usefulness of a blended online and in-lab environment to teach in Optometry. We administered a survey to compare responses of optometry students who had access to the interactive website, with those from students from a previous year who used the static version. We examined learning preferences of students in a focus group. Students were positive about the value of the website for their learning and the clarity of the website content. Nevertheless, objective comparison of pass rates for students using the static and interactive websites did not show significant changes. The majority of students commenting on the static website felt they did not get sufficient feedback via the website (67%) compared with only 22% from students who used self-assessments in the interactive website. Interestingly, users of the static website commented that it was perceived as just another resource while users of the interactive website commented on the usefulness of the material to review knowledge before laboratories. In the focus group, students reported they preferred a blended learning over the website alone even by students using the interactive website as they felt the need to revise content with the educator before the test. We conclude that there is acceptance of online learning methods due to the technologically ‘savvy’ environment of students in the first year of the Optometry programme but there is still dependence on the educator as the main administrator of their learning.