Combinations of recently developed paradoxical concanavalin-A staining (PCS) and other routine histochemical procedures have made it possible to classify mucosubstances more precisely. By taking advantage of these sequences, the present study was undertaken both to characterize the mucinous contents of alimentary tracts of several animal species, and to compare the variability of epithelial mucosubstances with special reference to the mucous neck cells of the stomach. The alimentary tracts obtained from 7 species of fish, 5 amphibians, 3 reptiles, 4 birds, and 6 mammals were used. Acidity of epithelial mucosubstances was highly variable even among the corresponding mucous cells. On the contrary, concanavalin-A reactivity seemed to be consistent with each cell and species. The mucous neck cells, which were observed in amphibians, reptiles, and mammals, consistently exhibited a characteristic mucosubstance with stable class-III reactivity by PCS. Inversely, stable class-III reactivity was found only in species possessing mucous neck cells, and was widely distributed in the esophageal glands of the frog, snake, and man; in esophageal mucous cells of the frog and skink; in pyloric glands of amphibia, reptiles, and mammals; and in Brunner's glands of mammals. These mucous cells seem to form an unique group with morphological and histochemical similarities. It is likely that the stable class-III reactivity by PCS is a hallmark of the mucous neck cells and related glands and that, in addition, concanavalin-A reactivity of mucosubstances is evolutionarily more fundamental characteristic.

(SE) S U M M A R Y The morphology of the osteocyte changes during the cell's lifetime. Shortly after becoming buried in the matrix, an osteocyte is plump with a rich rough endoplasmic reticulum and a well-developed Golgi complex. This "immature" osteocyte reduces its number of organelles to become a "mature" osteocyte when it comes to reside deeper in the bone matrix. We hypothesized that mineralization of the surrounding matrix is the trigger for osteocyte maturation. To verify this, we prevented mineralization of newly formed matrix by administration of 1-hydroxyethylidene-1,1-bisphosphonate (HEBP) and then examined the morphological changes in the osteocytes in rats. In the HEBP group, matrix mineralization was disturbed, but matrix formation was not affected. The osteocytes found in the unmineralized matrix were immature. Mature osteocytes were seen in the corresponding mineralized matrix in the control group. The immature osteocytes in the unmineralized matrix failed to show immunoreactivity with anti-sclerostin antibody, whereas mature osteocytes in the mineralized matrix showed immunoreactivity in both control and HEBP groups. These findings suggest that mineralization of the matrix surrounding the osteocyte is the trigger for cytodifferentiation from a plump immature form to a mature osteocyte. The osteocyte appears to start secreting sclerostin only after it matures in the mineralized bone matrix. (J Histochem Cytochem 56:561-567, 2008)

The purpose of this study was to examine the morphological changes in alveolar bone osteocytes on the pressure side during experimental tooth movement, using quantitative evaluation on hematoxylin and eosin-stained sections, the TUNEL method, confocal laser scanning microscopy (CLSM), and transmission electron microscopy. In 8-week-old Wistar rats, the left first molar was forced to move mesially with an average load of 10 g by a nickel-titanium superelastic wire. After 6 hours, nuclear condensation and fragmentation appeared in osteocytes adjacent to the hyalinized periodontal ligament (PDL). These cells showed TUNEL-positive reaction. The number of osteocytes with apoptosis progressively increased up to 1 day. At 1 and 2 days, cytoplasmic and nuclear destruction and distribution within the lacunae occurred and increased up to 4 days. The proportion of necrotic osteocytes and near empty lacunae peaked at 2 and 4 days, respectively. At 7 days, necrotic osteocyte and empty lacunae numbers returned to the level of control bone, probably due to resorption of the alveolar bone containing apoptotic and necrotic osteocytes. Ultrastructually, the osteocytes showed apoptotic morphology at 6 and 12 hours and 1 day; at 2 and 4 days, several osteocytes exhibited characteristics of necrosis and destructive images of the surrounding bone matrix, which resulted in enlargement of the lacunae. The present results demonstrate that osteocytes in alveolar bone adjacent to the hyalinized PDL underwent cell death via apoptosis and "secondary necrosis" during orthodontic tooth movement, which may be associated with the subsequent bone resorption.

The middle portion of Meckel's cartilage (one of four portions that disappear with unique fate) degrades via hypertrophy and the cell death of chondrocytes and via the resorption of cartilage by chondroclasts. We have examined the immunolocalization of matrix metalloproteinase-2 (MMP-2), MMP-9, MMP-13, and MMP-14 (members of the MMP activation cascade) and galectin-3 (an endogenous substrate for MMP-9 and an anti-apoptotic factor) during resorption of Meckel's cartilage in embryonic mice and have compared the results with those of developing endochondral bones in hind limbs. MMP immunoreactivity, except for MMP-2, is present in nearly all chondrocytes in the middle portion of Meckel's cartilage. On embryonic day 15 (E15), faint MMP-2-immunoreactive and intense MMP-13-immunoreactive signals occur in the periosteal bone matrix deposited by periosteal osteoblasts on the lateral surface, whereas MMP-9 and MMP-14 are immunolocalized in the peripheral chondrocytes of Meckel's cartilage. The activation cascade of MMPs by face-to-face cross-talk between cells may thus contribute to the initiation of Meckel's cartilage degradation. On E16, immunopositive signaling for MMP-13 is detectable in the ruffled border of chondroclasts at the resorption front, whereas immunostaining for galectin-3 is present at all stages of chondrocyte differentiation, especially in hypertrophic chondrocytes adjacent to chondroclasts. Galectin-3-positive hypertrophic chondrocytes may therefore coordinate the resorption of calcified cartilage through cell-to-cell contact with chondroclasts. In metatarsal specimens from E16, MMPs are detected in osteoblasts, young osteocytes, and the bone matrix of the periosteal envelope, whereas galectin-3 immunoreactivity is intense in young periosteal osteocytes. In addition, intense MMP-9 and MMP-14 immunostaining has been preferentially found in pre-hypertrophic chondrocytes, although galectin-3 immunoreactivity markedly decreases in hypertrophic chondrocytes. These results indicate that the degradation of Meckel's cartilage involves an activation cascade of MMPs that differs from that in endochondral bone formation.

We examined epithelial regeneration in mechanically injured rat trachea using phenotypic markers that identify unique differentiated stages of epithelial cells. Following a focal denuding wound, the cells from the adjacent nonwounded epithelium flattened and migrated into the wounded site during the first 12 h. At 24 h, these cells dedifferentiated into poorly differentiated (PD) cells that did not precisely resemble any of the mature tracheal cells. Proliferation of PD cells produced a multilayered epithelium by 48 h. Mitotic activity, measured as mitotic rate (MR) following a 6-h colchicine metaphase blockade, was high at 24 h (MR 23.4%) and 48 h (MR 24.0%). These PD cells expressed keratin 14 and Griffonia simplicifolia I-isolectin B4 (GSI-B4) lectin binding sites, which are specific for basal cells in normal epithelium but did not react with secretory or ciliated cell markers. At 72 h, MR fell to 1.8% (control MR 0.38%). The wound was covered with a pseudostratified epithelium; secretory cell markers were present at the apex of differentiating columnar cells, and a few preciliated cells expressing ciliated cell markers appeared. Basal cells also became distinctly recognizable and expressed keratin 14 and GSI-B4 binding sites. Newly appearing secretory or ciliated cells also expressed these markers but lost them gradually as they acquired new sets of specific markers. During epithelial regeneration after mechanical injury, "dedifferentiation," "proliferation," and "redifferentiation" of epithelial cells occurred, and the PD cell was pivotal in this process.