How does proline-repeat motif length in the proteins of teeth and bones relate to the evolution of vertebrates? Counterintuitively, longer repeat stretches are associated with smaller aggregated subunits within a supramolecular matrix, resulting in enhanced crystal length in mammalian versus amphibian tooth enamel.

During the recent decade, the periodontal attachment apparatus has turned into one of the premier areas of the body for the development of novel tissue engineering strategies. In the present review we are using a developmental biology approach to characterize current concepts in periodontal regeneration and to discuss strategies for future applications in periodontal therapies. In order to decipher the developmental make-up of the periodontal region, we have followed the path of the migratory neural crest as it gives rise to periodontal progenitor tissues, which in turn are subjected to the influence of diverse craniofacial extracellular matrices and peptide growth factors. Based on this developmental perspective, we have conducted a systematic analysis of the factors, progenitor cells, and matrices used in current periodontal tissue engineering approaches. We propose that the developmental history of a tissue is a highly instructive design template for the discovery of novel bioengineering tools and approaches.

Macrophage spreading requires the microtubule cytoskeleton and protein kinase C (PKC). The mechanism of involvement of the microtubules and PKC in this event is not fully understood. Dynamitin is a subunit of dynactin, which is important for linking the microtubuledependent motor protein dynein to vesicle membranes. We report that dynamitin is a Ca 2؉ /calmodulin-binding protein and that dynamitin binds directly to macrophage-enriched myristoylated alanine-rice C kinase substrate (MacMARCKS), a membrane-associated PKC substrate involved in macrophage spreading and integrin activation. Dynamitin was found to copurify with Mac-MARCKS both during MacMARCKS purification with conventional chromatography and during the immunoabsorption of MacMARCKS using anti-MacMARCKS antibody. Vice versa, MacMARCKS was also found to cosediment with the 20 S dynactin complex. We determined that the effector domain of MacMARCKS is required to interact with the N-terminal domain of dynamitin. MacMARCKS and dynamitin also partially colocalized at peripheral regions of macrophages and in the cell-cell border of 293 epithelial cells. Treatment with phorbol esters abolished this colocalization. Disrupting the interaction with a short peptide derived from the MacMARCKS-binding domain of dynamitin caused macrophages to spread and flatten. These data suggest that the dynamitin-MacMARCKS interaction is involved in cell spreading. Furthermore, the regulation of this interaction by PKC and Ca 2؉ /calmodulin provides a possible regulatory mechanism for cell adhesion and spreading.

The role angiogenesis plays in atopic dermatitis is not well understood. The authors previously demonstrated ultrastructurally dermal microvascular angiogenesis in the IL-4-transgenic mouse model of atopic dermatitis. Here, they determine the angiogenic factors involved in dermal microvascular angiogenesis, regulatory function of inflammatory cytokines on the VEGF-A production, and microvascular permeability in this model. Computer-assisted photometric analyses for immunofluorescence-labeled CD31 demonstrated a progressive increase in blood vessel number, diameter, and percent dermal areas occupied by CD31(+) vessels as the disease evolves in transgenic mice from before disease onset through early and late skin lesions. Similar findings were documented for VEGR2(+) vessels. Quantification of skin angiogenic factor mRNAs showed progressive increase of transcripts of VEGF-A, but not VEGF-B, VEGF-C, or VEGF-D. ELISA showed a similar increase of VEGF-A in the serum and skin of transgenic mice. IL-6 and IFN-gamma stimulated VEGF-A mRNA production in the skin and in primary keratinocytes of transgenic mice. Other skin angiogenic factors that increased included Ang-1, Ang-2, GBP-1, and VE-cadherin. Microvascular leakage began in the transgenic mouse skin before disease onset and peaked in the late stage. In conclusion, IL-6 and IFN-gamma may play important roles in upregulation of VEGF-A, along with other pro-angiogenic factors, to induce dermal microvascular angiogenesis.

The extracellular tooth enamel matrix is a unique, protein-rich environment that provides the structural basis for the growth of long and parallel oriented enamel crystals. Here we have conducted a series of in vivo and in vitro studies to characterize the changes in matrix shape and organization that take place during the transition from ameloblast intravesicular matrices to extracellular subunit compartments and pericrystalline sheath proteins, and correlated these changes with stages of amelogenin matrix protein posttranslational processing. Our transmission electron microscopic studies revealed a 2.5-fold difference in matrix subunit compartment dimensions between secretory vesicle and extracellular enamel protein matrix as well as conformational changes in matrix structure between vesicles, stippled materials, and pericrystalline matrix. Enamel crystal growth in organ culture demonstrated granular mineral deposits associated with the enamel matrix framework, dot-like mineral deposits along elongating initial enamel crystallites, and dramatic changes in enamel matrix configuration following the onset of enamel crystal formation. Atomic force micrographs provided evidence for the presence of both linear and hexagonal/ring-shaped full-length recombinant amelogenin protein assemblies on mica surfaces, while nickel-staining of the N-terminal amelogenin N92 His-tag revealed 20 nm diameter oval and globular amelogenin assemblies in N92 amelogenin matrices. Western blot analysis comparing loosely bound and mineral-associated protein fractions of developing porcine enamel organs, superficial and deep enamel layers demonstrated (i) a single, full-length amelogenin band in the enamel organ followed by 3 kDa cleavage upon entry into the enamel layer, (ii) a close association of 8–16 kDa C-terminal amelogenin cleavage products with the growing enamel apatite crystal surface, and (iii) a remaining pool of N-terminal amelogenin fragments loosely retained between the crystalline phases of the deep enamel layer. Together, our data establish a temporo-spatial correlation between amelogenin protein processing and the changes in enamel matrix configuration that take place during the transition from intracellular vesicle compartments to extracellular matrix assemblies and the formation of protein coats along elongating apatite crystal surfaces. In conclusion, our study suggests that enzymatic cleavage of the amelogenin enamel matrix protein plays a key role in the patterning of the organic matrix framework as it affects enamel apatite crystal growth and habit.