Effect of surface roughness on initial responses of osteoblast-like cells on two types of zirconia.

The aim of this study was to evaluate the effect of surface roughness on the initial attachment of mouse osteoblast-like cells on ceriastabilized zirconia/alumina nanocomposite (NANOZR) and yttria-stabilized zirconia (3Y-TZP) in comparison to those on pure titanium (Ti) and alumina oxide (AO). Specimens with smooth and rough surfaces were prepared by grinding with diamond paper or by sandblasting, respectively. For four substrates examined, the number of attached cells on the rough surface specimens was significantly higher than that on the smooth surface specimens (p < 0.05). Integrin alpha(5) and beta(1) expression had a greater increase in rough surface specimens than in smooth surface specimens. Actin cytoskeleton organization was, however, similar for both smooth and rough surface specimens. NANOZR and 3Y-TZP produced good cell attachment, similar to Ti and AO. The overall results demonstrated that NANOZR and 3Y-TZP with rough surface could provide good initial cell responses, adequate for future implant usage.

Initial osteoblast-like cell response to pure titanium and zirconia/alumina ceramics.

OBJECTIVES: Outstanding mechanical properties, resistance to scratching and high biocompatibility make zirconia/alumina ceramics interesting for dental applications. To solve the problem of the well-known low temperature degradation and to provide stable mechanical properties a novel zirconia alloy ((Y,Nb)-TZP/alumina) was developed. The aim of this study was to investigate the initial bone cell response to this new zirconia/alumina composite ceramic. METHODS: HOS cells were cultured on zirconia/alumina composite (Zc) and pure titanium (Ti) discs. Surface topography was examined by atomic force microscopy (AFM), cell morphology by scanning electron microscopy (SEM). Cell proliferation (MTS) and alkaline phosphatase activity was measured at 1, 4 and 8 days. The mRNA expression of Cycline D1, the cell cycle regulating gene, integrin beta 1, osteonectin (ON) and beta-actin were evaluated by RT-PCR analysis after 12, 24 and 48 h. RESULTS: Both substrates showed a very smooth character with R(a)-values in the range of 0.002-0.113 microm supporting a continuous cellular growth. After 8 days, cell proliferation on Zc was higher than on Ti. The mRNA expression of cyclin D1 showed similar activity after 48 h on both surfaces, ALP activity was higher on Zc after 8 days. ON expression however showed no difference between the two groups. SIGNIFICANCE: Our data demonstrate that this new zirconia composite ceramic showed at least equivalent or slightly better biological response of osteoblast-like HOS cells than pure titanium during a short-time cell culture period.

Cis-polyunsaturated fatty acids stimulate beta1 integrin-mediated adhesion of human breast carcinoma cells to type IV collagen by activating protein kinases C-epsilon and -mu.

We have investigated the effects of various fatty acids (FAs) on integrin-mediated MDA-MB-435 breast carcinoma cell adhesion to type IV collagen (collagen IV) in vitro. Arachidonic acid (AA) and linoleic acid both induced a dose-dependent increase in cell adhesion to collagen IV with no significant increase in nonspecific adhesion to polylysine and BSA. Oleic acid (a monounsaturated FA), AA methyl ester, and linoelaidic acid (a trans-isomer of linoleic acid) failed to stimulate adhesion to collagen IV, suggesting that these effects required cis-polyunsaturation and a free carboxylic moiety and that they were not due to membrane perturbations. Calphostin C, a protein kinase C (PKC) inhibitor, blocked cis-polyunsaturated FA (cis-PUFA)-induced cell adhesion in a dose-dependent manner, suggesting a role for a calcium-dependent PKC in this signal transduction pathway. Immunoblotting revealed that cis-PUFAs induced the translocation of PKCepsilon and PKCmu, two of the novel PKC isozymes, from the cytosol to the membrane. In contrast, a conventional PKC isozyme, PKCalpha, as well as the atypical isozymes, PKCzeta and PKCiota, did not translocate after cis-PUFA treatment. Function-blocking antibodies specific for alpha1, alpha2, and beta1, integrin subunits inhibited cell adhesion to collagen IV, whereas antibodies to alpha3 and alpha5 did not. No increase in the expression of these integrins on the cell surface was detected after the incubation of cells with cis-PUFAs, suggesting that there is an increase in the activity, but not in the amount, of these beta1, integrins. Altogether, these data suggest that cis-PUFAs enhance human breast cancer cell adhesion to collagen IV by selectively activating specific PKC isozymes, which leads to the activation of beta1 integrins.

Increased expression of integrin and receptor tyrosine kinase genes during autograft fusion in the sponge Geodia cydonium.

Recently cDNAs coding for cell surface molecules have been isolated from sponges. The molecules for alpha-integrin, galectin, and receptor tyrosine kinase (RTK), obtained from the marine sponge, Geodia cydonium, have been described earlier. In the present study also the cDNA for one putative beta-integrin has been identified from G. cydonium. The deduced aa sequence comprises the characteristic signatures, found in other metazoan beta-integrin molecules; the estimated size is 95,215 Da. To obtain first insights into the molecular events which proceed during autograft fusion, the expressions of these genes were determined on transcriptional and translational level. The cDNAs as well as antibodies raised against the recombinant sponge proteins alpha-integrin, RTK and galectin were used and Northern blot experiments and immunocytochemical analyses have been performed. The results show that transcription of the two subunits of an integrin receptor as well as of the RTK are strongly upregulated after grafting; levels of > 10-fold have been determined in the fusion zone of the grafts after a 10 days incubation. Immunofluorescence studies of sections through the fusion zone support these data. In contrast the transcription of the gene encoding galectin is drastically downregulated after grafting. In a parallel series of experiments the level of the heat-shock protein-70 was determined and it was found that it remained unchanged after grafting. We conclude that integrin subunits and the RTK molecule are involved in self-self recognition of sponge.

Effect of Astragalin on matrix secretion and beta 1 integrin mRNA expression in human mesangial cells.

OBJECTIVE: To investigate the effect of Astragalin on human renal mesangial cells. METHODS: Cultured human mesangial cells were treated with Astragalin and Astragalin serum in different concentrations in the presence or absence of PDGF-BB, the proliferation and type IV collagen secretion of mesangial cells were measured by MTT assay and ELISA, and expression of beta 1 integrin gene was estimated by reverse transcription-polymerase chain reaction (RT-PCR) method, suspectively. RESULTS: After 72 hours Astragalin or Astragalin serum treatment, the proliferation of mesangial cells induced by PDGF-BB was inhibited significantly in a dose-dependent manner compared with untreated controls (P < 0.05 and P < 0.01). After 24 hours of Astragalin or Astragalin serum treatment, the secretion of type IV collagen protein in presence of PDGF-BB was significantly decreased and beta 1 integrin mRNA level decreased significantly compared with untreated control (P < 0.05, P < 0.01). CONCLUSIONS: Astragalin inhibits cell proliferation and matrix over-synthesis which might be mediated, at least, partly by decrease of beta 1 integrin gene over-expression. The study suggested that Astragalin might play a role in preventing the progression of chronic renal diseases.

Immunogold localization of beta 1-integrin in bone: effect of glucocorticoids and insulin-like growth factor I on integrins and osteocyte formation.

Insulin-like growth factor I (IGF-I) and high-dose glucocorticoids exert opposite effects on bone formation. Because integrins are involved in cell and matrix organization, the effect of glucocorticoids and IGF-I on integrins was investigated in bone. An immunogold transmission electron microscopic (TEM) method was developed and applied to an organ culture system of 20-day fetal rat parietal bones, which mineralize in vitro. In parietal bone culture, 100 mM corticosterone treatment for 72 hr decreased calcification by 29%, disrupted osteoblast organization, and decreased the number of osteocytes. The quantity of osteoblast processes and the number of osteocytes per unit bone area were decreased by 48% and 56%, respectively. This effect was dose-dependent. The beta 1-integrin subunit was localized equally to apical and basal osteoblast surfaces by immunogold TEM. Compared to untreated control cultures, corticosterone (100 nM) decreased beta 1 by one third. In contrast, treatment with IGF-I for 72 hr increased calcification by 38%, cell processes by 71%, and osteocytes per unit area of bone by 107%. The number of gold particles localizing beta 1 on the osteoblast plasma membrane doubled, almost entirely on the apical surface of the osteoblast. Glucocorticoids and IGF-I had no significant effect on beta 1 levels in osteocytes. In conclusion, glucocorticoids and IGF-I modulate integrin levels on osteoblasts, and influence osteocyte formation and bone calcification. However, neither glucocorticoids nor IGF-I alter beta 1-integrin levels on osteocytes.