Effects of low-level laser therapy on human osteoblastic cells grown on titanium

The aim of this study was to investigate the effects of low-level laser therapy (LLLT) by using gallium aluminum arsenide (GaAlAs) diode laser on human osteoblastic cells grown on titanium (Ti). Osteoblastic cells were obtained by enzymatic digestion of human alveolar bone and cultured on Ti discs for up to 17 days. Cells were exposed to LLLT at 3 J/cm2 (wavelength of 780 nm) at days 3 and 7 and non-irradiated cultures were used as control. LLLT treatment did not influence culture growth, ALP activity, and mineralized matrix formation. Analysis of cultures by epifluorescence microscopy revealed an area without cells in LLLT treated cultures, which was repopulated latter with proliferative and less differentiated cells. Gene expression of ALP, OC, BSP, and BMP-7 was higher in LLLT treated cultures, while Runx2, OPN, and OPG were lower. These results indicate that LLLT modulates cell responses in a complex way stimulating osteoblastic differentiation, which suggests possible benefits on implant osseointegration despite a transient deleterious effect immediately after laser irradiation.

Este estudo teve como objetivo investigar o efeito do laser diodo de gálio-alumínio-arsênio (GaAlAs) em células osteoblásticas humanas cultivadas sobre discos de Ti. Para tanto, células osteoblásticas foram obtidas por digestão enzimática de osso alveolar humano e cultivadas sobre discos de Ti por 17 dias. As células foram submetidas à irradiação no 3º e 7º dias na dose de 3 J/cm2 e comprimento de onda de 780 nm e células não irradiadas foram usadas como controle. A irradiação não alterou a proliferação celular, atividade de ALP e formação de matriz mineralizada. Microscopia por epifluorescência indicou que após 24 h da aplicação do laser, as culturas irradiadas apresentaram áreas sem células, que mais tarde foram repovoadas por células em fase de proliferação e menos diferenciadas. O laser aumentou a expressão gênica relativa da ALP, OC, BSP e BMP-7 e reduziu a de RUNX2, OPN e OPG. Os resultados indicam que a terapia com laser modula de forma complexa as respostas celulares, estimulando a diferenciação osteoblástica. Assim, é possível sugerir possíveis benefícios do laser na osseointegração de implantes de Ti apesar do efeito deletério às células imediatamente após a irradiação.

Effect of titanium surface roughness on human bone marrow cell proliferation and differentiation: an experimental study / Efeito da rugosidade de superfície de discos de titânio sobre a proliferação e diferenciação de células de medula óssea humana: estudo experimental

PURPOSE: To assess the proliferation and differentiation of human bone marrow-derived cells cultured on titanium surfaces with different roughness characteristics. METHODS: Cells obtained from the iliac crest of an adult human donor were routinely processed and cultured on titanium surfaces of varying roughness, according to their preparation method: polishing only (smooth surface) and polishing followed by etching with HF/HNO3 for 15 and 30 minutes (rough surfaces). Surfaces were assessed using scanning electronic microscopy and profilometry. RESULTS: Titanium disks etched with acid for 15 minutes allowed greater cell proliferation in all culture periods. The level of osteopontin and osteocalcin expression was increased in both acid-etched groups, which indicates an advanced stage of differentiation of cells into osteoblasts. CONCLUSIONS: Increased surface roughness accelerates the differentiation of undifferentiated mesenchymal cells into osteogenic lineage cells, but does not necessarily favor cell proliferation. An intermediate surface roughness of 0.5µm (acid etching for 15 minutes) favors both initial and final cell responses.

OBJETIVO: Avaliar a proliferação e diferenciação de células derivadas da medula óssea humana sobre superfícies de titânio com diferentes rugosidades de superfície. MÉTODOS: Células obtidas da crista ilíaca de um doador humano adulto foram rotineiramente processadas e cultivadas sobre superfícies de titânio preparadas através de polimento apenas, ou polimento seguido de condicionamento com HF/HNO3 por 15 e 30 minutos, para produzir superfícies com rugosidades variadas, conforme determinado por Microscopia Eletrônica de Varredura e perfilometria. RESULTADOS: Discos de titânio condicionados com ácido por 15 minutos permitiram maior proliferação celular em todos os períodos de cultura. O nível de expressão das proteínas osteopontina e osteocalcina estava aumentado em ambos os grupos condicionados com ácido, indicando que as células estavam comprometidas com o fenótipo osteogênico. CONCLUSÕES: O aumento na rugosidade de superfície acelera a diferenciação de células mesenquimais indiferenciadas em células de linhagem osteogênica, mas não necessariamente favorece a proliferação celular. Uma superfície com rugosidade intermediária de 0,5µm (condicionada com ácido por 15 minutos) favorece tanto as respostas celulares iniciais quanto as finais.

Influence of the microenvironment on gene and protein expression of odontogenic-like and osteogenic-like cells

Progenitor cells play an important biological role in tooth and bone formation, and previous analyses during bone and dentine induction have indicated that they may be a good alternative for tissue engineering. Thus, to clarify the influence of the microenvironment on protein and gene expression, MDPC-23 cells (mouse dental papilla cell line) and KUSA/A1 cells (bone marrow stromal cell line) were used, both in vitro cell culture and in intra-abdominal diffusion chambers implanted in 4-week-old male immunodefficient mice (SCID mice). Our results indicate that KUSA/A1 cells differentiated into osteoblast-like cells and induced bone tissue inside the chamber, whereas, MDPC-23 showed odontoblast-like characteristics but with a low ability to induce dentin formation. This study shows that MDPC-23 cells are especial cel ls, which possess morphological and functional characteristics of odontoblast-like cells expressing dentin sialophosphoprotein in vivo. In contrast, dentin sialophosphoprotein gene and protein expression was not detected in both cell lines in vitro. The intra-abdominal diffusion chamber appears as an interesting experimental model for studying phenotypic expression of dental pulp cells in vivo.

Immunohistochemical study of osteopontin in boar testis

The expression of osteopontin (OPN) in boar testis was studied. Western blot analysis detected 66- and 32-kDa OPN immunopositive bands in the testes of adult boars. In postnatal piglets, the 66-kDa OPN band was detected in the testes, but not the 32-kDa band. In the newborn testis, OPN immunostaining was seen in gonocytes and in some supporting cells in the seminiferous tubules, as well as in interstitial Leydig cells. In the adult boar testis, OPN immunoreactivity was detected in seminiferous tubules with varying intensities. Intense OPN immunostaining was seen in the residual bodies and acrosomes in the spermatids while, occasionally, OPN immunostaining was seen in spermatogonia and various stage of spermatocytes but in few Sertoli cells in the seminiferous tubules. In addition, Leydig cells in adult boars were weakly immunostained with OPN. These findings suggest that OPN is detected in the majority of germ cells and is involved in spermatogenesis in boar testis.