Dataset Information

Acid bone lysates: what we eliminate by preparing demineralized bone matrix

ABSTRACT: Demineralized bone matrix is a widely used allograft where not only the inorganic mineral but also embedded growth factors are removed by hydrochloric acid. The cellular response to the growth factors released during the preparation of demineralized bone matrix, however, has not been studied. Here we investigated the in vitro impact of acid bone lysates (ABL) prepared from porcine cortical bone chips on oral fibroblasts. Proteomic analysis of ABL revealed a large spectrum of bone-derived proteins including TGF-β1. Whole genome microarrays and RT-PCR together with the pharmacologic blocking of TGF-β receptor type I kinase with SB431542 showed that ABL activates the TGF-β target genes interleukin 11, proteoglycan 4, and NADPH oxidase 4. Interleukin 11 expression was confirmed at the protein level by ELISA. Immunofluorescence and Western blot showed the nuclear localization of Smad2/3 and increased phosphorylation of Smad3 with ABL, respectively. This effect was independent of whether ABL was prepared from mandible, calvaria or tibia. ABL prepared with 0.1 N HCL was most effective compared to lysates obtained with 0.01 N and also 1.0 N HCl. These results demonstrate that TGF-β is a major growth factor that is removed upon the preparation of demineralized bone matrix.

INSTRUMENT(S): Q Exactive

ORGANISM(S): Homo Sapiens (human) Sus Scrofa Domesticus (domestic Pig)

TISSUE(S): Mandible, Gingival Fibroblast, Tibia, Calvarial Osteoblast, Fibroblast

SUBMITTER: Goran Mitulović

LAB HEAD: Goran Mitulović

PROVIDER: PXD010145 | Pride | 2018-11-04

REPOSITORIES: Pride

ACCESS DATA

Publications

Acid bone lysate activates TGFβ signalling in human oral fibroblasts.

Strauss Franz Josef FJ Stähli Alexandra A Beer Lucian L Mitulović Goran G Gilmozzi Valentina V Haspel Nina N Schwab Gerhild G Gruber Reinhard R

Scientific reports 20181030 1

Demineralized bone matrix is a widely used allograft from which not only the inorganic mineral but also embedded growth factors are removed by hydrochloric acid (HCl). The cellular response to the growth factors released during the preparation of demineralized bone matrix, however, has not been studied. Here we investigated the in vitro impact of acid bone lysate (ABL) prepared from porcine cortical bone chips on oral fibroblasts. Proteomic analysis of ABL revealed a large spectrum of bone-deriv ...[more]

PMID: 30375456

Similar Datasets

Project description:To evaluate the long-term growth potential of BCR-ABL-transduced primitive human hematopoietic cells, lin- cord blood cells containing an MSCV-BCR-ABL-IRES-GFP (BCR-ABL) or control-GFP transgene (MIG) were injected IP into fetal goats at 45-55 days of gestation. Six transplant goats were born alive. One was examined three weeks after birth and showed GFP+ cells in the blood, bone marrow (BM), liver, kidney, lung, heart, and both skeletal and smooth muscle. FISH analysis also showed the liver of this goat contained BCR-ABL-GFP transgenic cells. The remaining five goats appear normal although, in some, the WBC count is elevated 3- to 5-fold. GFP+ cells, including cells identifiable by FACS as human CD34+ cells, have been detected in the blood of all these goats. The presence of BCR-ABL-GFP transgenic cells in the BM and liver was confirmed by FISH analysis, and quantitative real-time PCR analysis of genomic DNA isolated from unpurified BM cells obtained from three of the transplant goats demonstrated 3-5Ã104 copies of the transgene per microgram of DNA. Microarray transcript profiling was performed on blood and liver tissues of normal goats, BCR-ABL chimeric goats, MIG chimeric goats, and normal human samples. RNA for human genes was detected in goats transplanted with cord blood cells but not in normal goats, and the RNA abundance of some genes in BCR-ABL chimeric goat blood was similar to or greater than levels observed in MIG goat blood or normal human samples. Quantitative RT-PCR confirmed the differential expression of several genes in goats carrying the BCR-ABL vs. control transgene. These results demonstrate long-term engraftment but slow expansion in a large animal model of primitive human hematopoietic cells transduced with a BCR-ABL fusion gene and transplanted in utero. This novel xenotransplant goat model should be useful for analyzing the initial phases of development of human CML and for assessing new therapies with potential long-term benefits. Experiment Overall Design: Total RNA was extracted from liver (L) and blood (B) samples of normal goats (ng), humans (hu), chimeric goats engrafted with human cord blood stem cells containing control (mig) vector, and chimeric goats engrafted with CML (bcrabl) vector. RNA samples were profiled on Affymetrix human U133A GeneChips and examined for differentially expressed genes in CML vs control goats, filtering for signals significantly above background levels observed in normal goat to select for specific human gene expression.

Dataset Information

Acid bone lysates: what we eliminate by preparing demineralized bone matrix

Publications

Acid bone lysate activates TGFβ signalling in human oral fibroblasts.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure