c-src deletion in mice leads to osteopetrosis as a result of reduced bone resorption due to an alteration of the osteoclast. We report that deletion/reduction of Src expression enhances osteoblast differentiation and bone formation, contributing to the increase in bone mass. Bone histomorphometry showed that bone formation was increased in Src null compared with wild-type mice. In vitro, alkaline phosphatase (ALP) activity and nodule mineralization were increased in primary calvarial cells and in SV40-immortalized osteoblasts from Src−/− relative to Src+/+ mice. Src-antisense oligodeoxynucleotides (AS-src) reduced Src levels by ∼60% and caused a similar increase in ALP activity and nodule mineralization in primary osteoblasts in vitro. Reduction in cell proliferation was observed in primary and immortalized Src−/− osteoblasts and in normal osteoblasts incubated with the AS-src. Semiquantitative reverse transcriptase-PCR revealed upregulation of ALP, Osf2/Cbfa1 transcription factor, PTH/PTHrP receptor, osteocalcin, and pro-alpha 2(I) collagen in Src-deficient osteoblasts. The expression of the bone matrix protein osteopontin remained unchanged. Based on these results, we conclude that the reduction of Src expression not only inhibits bone resorption, but also stimulates osteoblast differentiation and bone formation, suggesting that the osteogenic cells may contribute to the development of the osteopetrotic phenotype in Src-deficient mice.

Our study suggests considerable disagreement among pathologists on the diagnosis of melanoma versus other pigmented lesions. Tumor thickness and presence of ulceration are the most reproducible histologic features of cutaneous melanoma.

No abstract

We have previously reported that epidermal growth factor (EGF) stimulates branching morphogenesis of the fetal mouse submandibular gland (SMG) (M. Kashimata and E. W. Gresik, 1997, Dev. Dyn. 208, 149-161) and that the EGF receptor (EGFR) is localized principally, if not exclusively, on the epithelial components of the fetal SMG (E. W. Gresik, M. Kashimata, Y. Kadoya, R. Mathews, N. Minami, and S. Yamashina, 1997, J. Histochem. Cytochem. 45, 1651-1657). The EGFR is a receptor tyrosine kinase, and after binding of its ligand, it triggers several intracellular signaling cascades, among them the one activating the mitogen-activated protein kinases (MAPK) ERK-1/2. Here we investigated whether EGF utilizes the ERK-1/2 signaling cascade to stimulate branching morphogenesis in the fetal mouse SMG. SMG rudiments were collected as matched pairs at E14, E16, and E18 (E0 = day of vaginal plug); placed into wells of defined medium (BGJb); and exposed to EGF for 5 or 30 min or to medium alone (controls). By Western blotting we found that EGF induced the appearance of multiple bands of phosphotyrosine-containing proteins, including bands at 170 kDa and 44 kDa/42 kDa, presumably corresponding to the phosphorylated forms of EGFR and ERK-1/2, respectively. Other blots showed the specific appearance of the phosphorylated EGFR and of phospho-ERK-1/2 in response to EGF. Immunohistochemical staining for phosphotyrosine increased at the plasma membrane after EGF stimulation for 5 or 30 min. Diffuse cytoplasmic staining for MEK-1/2 (the MAPK kinase that activates ERK-1/2) increased near the cell membrane after EGF stimulation. Phospho-ERK-1/2 was localized in the nuclei of a few epithelial cells after EGF for 5 min, but in the nuclei of many cells after EGF for 30 min. PD98059, an inhibitor of phosphorylation and activation of MEK-1/2, by itself inhibited branching morphogenesis and, furthermore, decreased the stimulatory effect of EGF on branching. Western blots confirmed that this inhibitor blocked phosphorylation of ERK-1/2 in fetal SMGs exposed to EGF. These results show that components of the ERK-1/2 signaling cascade are present in epithelial cells of the fetal SMG, that they are activated by EGF, and that inhibition of this cascade perturbs branching morphogenesis. However, EGF did not cause phosphorylation of two other MAPKs, SAPK/JNK or p38MAPK, in fetal SMGs. These results imply that the ERK-1/2 signaling is responsible, at least in part, for the stimulatory effect of EGF on branching morphogenesis of the fetal mouse SMG.

No abstract

Three patients are described who presented with a glomerulopathy suggestive of lupus nephritis in the absence of other clinical and biological evidence of systemic lupus erythematosus (SLE). Renal biopsies showed a "full-house" immunofluorescence pattern and two patients also had cytoplasmic tubuloreticular inclusions by electron microscopy. All these patients developed antinuclear and anti-double-stranded DNA antibodies 3, 5, and 10 years after their original presentation. Subsequently, 1 patient also developed clinical symptoms of lupus. Reviewing all renal biopsies performed in our department, we found 14 additional patients who presented with a "full-house" immunofluorescence glomerulonephritis in the absence of other features of SLE. After a mean follow-up of 5.8 years, these patients have not developed serological or clinical evidence of SLE. We conclude that a "full-house" glomerulopathy in children may be the first symptom of SLE, especially when cytoplasmic tubuloreticular inclusions are detected. The appearance of other clinical and biological symptoms may be delayed by several years.

The carbohydrate histochemistry of normal human kidney has been investigated by the use of four peroxidase-labeled lectins at the light and electron microscopic level. The results show that the lectin of Lotus tetragonolobus, specific for L-fucose, binds exclusively to the proximal convoluted tubules of the nephron. While peanut and soybean lectins, specific for o-galactose and N-acetyl-D-galactosamine, respectively, are confined to the collecting ducts, wheat germ lectin, specific for sialic acid and N-acetyl-Dglucosamine, stains several parenchymal structures, in-Arachis hypogaea (peanut), GI3cine max (soybean), and Lotus let ragonolobus (winged pea) were obtained from Sigma Chemical Company.

Neutrophil adhesion to the vascular endothelium is enhanced during tissue ischemia and/or inflammation, conditions that are associated with tissue acidosis. This study examined the effects of hypercarbic acidosis (10 or 20% CO2) and of hypocarbic alkalosis (0% CO2) on human neutrophil CD18 and human aortic endothelial cell intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and E-selectin expression quantified by flow cytometry. Acidosis with 20% CO2 for 4 h decreased ICAM-1 to 60.6 +/- 9.7% of control. In contrast, alkalosis with 0% CO2 for 4 h enhanced ICAM-1 expression to 143.8 +/- 10.1% of control. There was no pH dependence of VCAM-1 or E-selectin expression. Tumor necrosis factor-alpha (TNF-alpha; 10 ng/ml) increased endothelial ICAM-1, E-selectin, and VCAM-1; under these conditions, acidosis with 20% CO2 blunted both ICAM-1 and E-selectin surface expression compared with 5% CO2-, TNF-alpha-treated cells. Hypercarbic acidosis with 20% CO2 increased neutrophil CD18 expression and enhanced neutrophil adhesion. This latter effect was inhibited by neutrophil pretreatment with an anti-CD18 monoclonal antibody. In contrast, when only endothelial cells were preincubated with the hypercarbic buffer, neutrophil adhesion diminished to 55.6 +/- 7.8% of control. The results suggest that acidosis generated during tissue ischemia/inflammation may induce CD18-mediated neutrophil adhesion despite a decrease in ICAM-1 expression.