Clinical significance of expression of membrane type 1 matrix metalloproteinase and matrix metalloproteinase-2 in human head and neck squamous cell carcinoma

Three different membrane-type matrix metalloproteinases (MT-MMPs) activate in vitro the latent form of matrix metalloproteinase-2 (MMP-2), which is one of the key proteinases in invasion and metastasis of various cancers. We examined the mRNA expression of MT1, 2, and 3-MMPs and MMP-2 in cell lines of head and neck squamous cell carcinoma (HNSCC) and quantitated the relative expression levels in human HNSCC tissues by Northern blotting. The tissue localization of MT1-MMP and MMP-2 was determined by immunohistochemistry and in situ hybridization. Their implications in clinicopathologic factors were statistically evaluated. All cell lines examined consistently expressed MT1-MMP and MMP-2, but not MT2, 3-MMP. In the clinical specimens, there was a significant correlation in coexpression of messenger of RNA (P = .0005) and colocalization by immunohistochemistry (P < .0001) for MT1-MMP and MMP-2. Relative mRNA expression levels of MT1-MMP and MMP-2 in the carcinoma tissues were significantly higher than those of the control tissues (P = .0045 and P = .0122, respectively). Both mRNA expression level and immunopositivity of MT1-MMP significantly correlated with lymph node metastasis (P = .0081 and P = .0193, respectively), which was confirmed by multivariate logistic regression analysis. Immunoreaction of MT1-MMP and its mRNA expression were observed in both carcinoma cells and stromal cells. The localization of MMP-2 closely corresponded to that of MT1-MMP. These observations suggest that MT1-MMP possesses a role as a determinant of lymph node metastasis in HNSCC, and that concurrent expression of MT1-MMP and MMP-2 are involved in progression of HNSCC.     

Effect of high NaCl intake on Na+ and K+ transport in the rabbit distal convoluted tubule

Morphological studies have demonstrated that a chronic increase in distal Na⁺ delivery causes hypertrophy of the distal convoluted tubule (DCT). To examine whether high NaCl-intake also causes functional changes in the well defined DCT, we measured transmural voltage (V _T), lumen-to-bath Na⁺ flux (J _Na(LB)), and net K⁺ secretion (J _K(net)) in DCTs obtained from control rabbits and those on high NaCl-intake diets. The lumen negativeV _T was significantly greater in the high NaCl group than in the control group. The net K⁺ secretion (pmol mm^–1 min^–1) was greater in the high NaCl-intake group (54.1±13.0 vs 14.7±5.6). The K⁺ permeabïlities in both luminal and basolateral DCT membranes, as assessed by the K⁺-induced transepithelial voltage deflection inhibitable with Ba²⁺, were increased in the experimental group. The lumen-to-bath²²Na flux (pmol mm^–1 min^–1) was also greater in the experimental group (726±119 vs 396±65). TheV _T component inhibitable with amiloride was also elevated in the high NaCl-intake group. Furthermore, Na⁺–K⁺-ATPase activity of the DCT was higher in the experimental than in the control group. We conclude that high NaCl intake increases both Na⁺ reabsorption and K⁺ secretion by the DCT. This phenomenon is associated with an increased Na⁺–K⁺-ATPase activity along with increased Na⁺ and K⁺ permeabilities of the luminal membrane, and an increase in the K⁺ permeability of the basolateral membrane. Cellular mechanisms underlying these functional changes remain to be established.     

Neovascularization of transplanted central nervous tissue suspensions: an immunohistochemical study with laminin

Neovascularization of the dissociated central nervous tissue transplanted into the lateral ventricle of the rat was studied using laminin immunohistochemistry. A very high immunoreactive response to laminin was demonstrated in the presumably newly formed vessels within the transplants and the graft-host borders. The growing tips and fine spike-like sprouts called 'streamers' were also highly stained with laminin immunoreactivity. In contrast, laminin immunoreactivity was negligible in the vessels in the host brain. Therefore, these results indicate that laminin may be utilized as a marker for the newly formed vessels in neural transplantations.     

Possible involvement of keratinocyte growth factor and its receptor in enhanced epithelial-cell proliferation and acquired recurrence of middle-ear cholesteatoma

Middle-ear cholesteatoma is characterized by enhanced proliferation of epithelial cells and granular tissue formation. However, the molecular mechanism underlying these pathological changes is largely unknown. Keratinocyte growth factor (KGF) is a mesenchymal cell-derived paracrine growth factor that specifically stimulates epithelial cell proliferation. In the present study, we investigated the possible involvement of KGF and its receptor, KGFR, in the pathogenesis of cholesteatoma using in situ hybridization and immunohistochemistry, respectively. We examined 56 cholesteatoma specimens, and 8 normal skin areas as control. KGF and KGFR expression was examined by immunohistochemistry using rabbit anti-human KGF and anti-human KGFR polyclonal antisera raised in our laboratories against synthetic peptides corresponding to parts of human KGF and KGFR, respectively. KGF protein and mRNA were detected exclusively in stromal fibroblasts and infiltrating T lymphocytes in 80% of cholesteatoma cases, whereas KGFR protein and mRNA were localized in the epithelium in 72% of cases. Assessment of the proliferative activity of cholesteatoma using the labeling index for Ki-67 showed a significantly higher Ki-67 labeling index (66%) in KGF+/KGFR+ cases than other cases. There was a significant correlation between KGF+/KGFR+ expression and recurrence. Our results indicate the possible involvement of both KGF and KGFR in enhanced epithelial cell proliferative activity and recurrence of cholesteatoma.     

Human vascular smooth muscle cells and endothelial cells lack catalase activity and are susceptible to hydrogen peroxide

⁵Cr release as lytic and cell detachment as nonlytic injury were employed to estimate neutrophil-mediated injury of cultured human vascular smooth muscle cells and endothelial cells. The reagents hydrogen peroxide or hypoxanthine-xanthine oxidase produced dose-dependent killing and nonlytic cell detachment, which were specifically inhibited by catalase but not by superoxide dismutase. The concentration of hydrogen peroxide or xanthine oxidase to induce cell detachment was less than lytic dose, suggesting that cell detachment was a much more sensitive assay of injury. Neutrophil-mediated cell lysis averaged 15% at most and was mostly dependent on hydrogen peroxide, while neutrophil-mediated cell detachment was nearly 100% and its dependency on hydrogen peroxide varied from 46% to 60%. These results suggest that vascular smooth muscle cells and endothelial cells in neutrophil-mediated events are destroyed by a hydrogen peroxide-dependent process, mainly via a nonlytic cell detachment mechanism. There was no striking difference of sensitivity to hydrogen peroxide between vascular smooth muscle cells and endothelial cells. Vascular smooth muscle ceils and endothelial cells contained fairly high concentrations of superoxide dismutase, but not catalase, activity. The sensitivity of these cells to hydrogen peroxide but not to superoxide may arise from the fact that these cells lack intracellular catalase activity. The injury of vascular cells, which constitute important components of blood vessels, may lead to vascular injury and subsequent tissue damage.     

Molecular analysis of Cryptococcus neoformans mitochondrial cytochrome b gene sequences

Mitochondrial cytochrome b genes (cyt b) of 40 strains of Cryptococcus neoformans were partially sequenced to determine the genetic relations. With the exception of the type strain of C. neoformans var. neoformans, all strains contained introns in their sequences. Analysis of 386 bp of coding sequence from each strain under investigation revealed a total of 27 (6.99%) variable nucleotide sites and categorized isolates of C. neoformans into nine cyt b types. C. neoformans var. gattii included cyt b types I to V, and C. neoformans var. neoformans comprised types VI to IX. cyt b types were correlated with serotypes. All strains with cyt b types I, IV, and V were serotype B. All other strains except IFM 5878 (serotype B) with cyt b types II and III were serotype C. Serotype D strains had cyt b types VI and IX, and serotype A strains were cyt b type VIII. Of four serotype AD strains, one was cyt b type VII and the remaining three were type VIII. The phylogenetic tree based on deduced amino acid sequences divided the strains only into C. neoformans var. neoformans and C. neoformans var. gattii. These results indicate that cyt b sequences are effective for DNA typing as well as phylogenetic analysis of C. neoformans.     

Blood endotyping distinguishes the profile of vitiligo from that of other inflammatory and autoimmune skin diseases

1. Download : Download high-res image (180KB)
2. Download : Download full-size image

     

Cartilage regeneration using slow release of bone morphogenetic protein-2 from a gelatin sponge to treat experimental canine tracheomalacia: a preliminary report

We investigated whether saber sheath-type tracheomalacia could be treated by the slow release of bone morphogenetic protein (BMP)-2 from a gelatin sponge. A 1 cm gap was made in the middle portion of each of 10 consecutive tracheal cartilage rings in the canine cervix (control group, n = 3), then a gelatin sponge containing 12 microg of BMP-2 solution was implanted in the gap (12 microg group, n = 3). In another group (120 microg + P group, n = 3), the implanted gelatin sponge contained 120 microg of BMP-2 solution, and the gap was covered with periosteum. All of the control dogs developed saber sheath-type tracheomalacia, whereas tracheomalacia was not observed in the 12 microg and 120 microg + P groups. In the 12 microg group, fibrous cartilage was observed at the ends of the cartilage stumps. In the 120 microg + P group, newly formed bone and cartilage were observed to form a bridge between the cartilage stumps. The regeneration of cartilage or bone induced by the slow release of BMP-2 from a gelatin sponge might be useful for treatment of tracheomalacia.