BACKGROUNDThe palate is differently regulated and developed along the anterior-posterior axis. The Bmp signal pathway plays a crucial role in palatogenesis. Conditioned-inactivation of Bmp type I receptor Alk2 or Alk3 in the neural crest or craniofacial region leads to palatal cleft in mice. However, how different Bmp members are involved in palatogenesis remains to be elucidated. In the present study, mRNA expression patterns of Bmp2, Bmp3 and Bmp4 in the developing anterior and posterior palates were examined and compared, focusing on the fusion stage.

METHODSTo detect the expression of Bmp mRNA, antisense riboprobes were synthesized by in vitro transcription. Radioactive in situ hybridization was performed on sagital and coronal sections of mice head from E13 to E18.

RESULTSThe expression of these Bmps were developmentally regulated in the anterior and posterior palates prior to, during and after palatal fusion. During palatal fusion, Bmp4 expression shifted from the anterior to the posterior palate, Bmp2 was highly expressed in both the anterior and posterior palates in this process, whereas Bmp3 was only localized in the posterior palate. They showed generally non-overlapping pattern in their expression domains. Thereafter, their expression was detected in both the anterior and posterior palates regulating osteogenesis and myogenesis respectively.

CONCLUSIONSBmp signalling is involved in palatogenesis in multiple stages and has multiple roles in regulating anterior and posterior palatal development. Disturbances of Bmp signalling during palatogenesis might be a possible mechanism of cleft palate.

Animals ; Bone Morphogenetic Protein 2 ; Bone Morphogenetic Protein 3 ; Bone Morphogenetic Protein 4 ; Bone Morphogenetic Proteins ; genetics ; Female ; Gene Expression Regulation, Developmental ; Mice ; Palate ; embryology ; metabolism ; RNA, Messenger ; analysis ; Signal Transduction ; Transforming Growth Factor beta ; genetics

Animals ; Bone Morphogenetic Protein 2 ; metabolism ; Bone Morphogenetic Protein 4 ; metabolism ; Carrier Proteins ; pharmacology ; Liver ; metabolism ; Liver Regeneration ; Male ; Rats ; Rats, Wistar

OBJECTIVETo determine the expression of bone morphogenetic protein 2 and 4 (BMP-2 and BMP-4) in prostatic carcinoma (PCa) and investigate their relationship with clinical stage and Gleason score of tumor.

METHODSForty-eight PCa cases and 5 normal prostatic tissue were analysed for the expressions of BMP-2 and BMP-4 by Western bolt assay.

RESULTSThe optical densities of BMP-2 expressions in the tumor with Gleason score < or =5, 6-8, and > or = 9 were 7547.1 +/- 1964.12, 9657.4 +/- 2010.54, 12467.7 +/- 2496.75 and of BMP-4 expressions were 5174.4 +/- 1400.54, 5940.3 +/- 1587.42, 6332.1 +/- 1647.83, respectively. The optical densities of BMP-2 expressions in the tumor in T1 - T2 and T3 - T4 stages were 8003.37 +/- 1889.23, 12385.55 +/- 2506.72 and of BMP4 expressions were 5267.41 +/- 1 464.19, 6543.75 +/- 1668.46, respectively. There were significant differences between tissues with Gleason score < or =5 and > or =9 (P <0.01), and tissues in T1 - T2 and T3 - T4 stages, in expressions of BMP-2 protein. The expression of BMP-2 protein was significantly high in the PCa with bone metastasis compared with that without bone metastasis.

CONCLUSIONThe expressions of BMP-2 and BMP-4 increase with the progression of clinical stage and Gleason score compared with normal prostatic tissue. The expression of BMP-2 protein is significantly upregulated in bone metastasis of PCa, which indicates a poor prognosis.

Aged ; Aged, 80 and over ; Blotting, Western ; Bone Morphogenetic Protein 2 ; Bone Morphogenetic Protein 4 ; Bone Morphogenetic Proteins ; biosynthesis ; Humans ; Male ; Middle Aged ; Neoplasm Metastasis ; Neoplasm Staging ; Prostate ; metabolism ; Prostatic Neoplasms ; metabolism ; pathology ; Transforming Growth Factor beta ; biosynthesis

This study was aimed to compare the expression of BMP4 mRNA in the in-vivo tissue engineering bone constructed with Ca/P ceramics against the expression of BMP4 mRNA in the naturally healing bone. 20 porous Ca/P ceramics cylinders with Phi 5 mm x 8 mm were made and implanted into the dorsal muscles of 5 dogs. As control, one molar tooth was pulled out from each dog to create bone defect for the naturally healing bone at the same time. The specimens and the naturally healing bone were harvested at 1, 2, 4, 12 and 24 weeks post-implantation. After RNA extraction and reverse transcription, bone morphogenetic protein 4 (BMP4) and GAPDH mRNA were detected by real-time quantitative polymerase chain reaction (PCR) method. The results showed that the expression level of BMP4 mRNA of the in-vivo tissue engineering bone constructed with Ca/P ceramics was higher than that of the naturally healing bone in the period of experiment. However, the in-vivo tissue engineering bone had the same chronological order of BMP mRNA expression that the naturally healing bone did. As a bone substitute analogous to autologous bone, the in-vivo tissue engineering bone constructed with Ca/P ceramics has the potential for clinical application.
Animals ; Bone Morphogenetic Protein 4 ; genetics ; metabolism ; Bone Substitutes ; chemistry ; Calcium Phosphates ; chemistry ; Ceramics ; chemistry ; Dogs ; Humans ; Implants, Experimental ; RNA, Messenger ; genetics ; metabolism ; Tissue Engineering

BACKGROUNDTissue-engineering techniques combined with gene therapy have been recently reported to improve osteogenesis. In this study, tissue-engineered bone constructed by human Bone Morphogenetic Protein 4 (hBMP-4) gene-modified bone marrow stromal cells (bMSCs) was explored in an ectopic bone formation model in rabbits.

METHODSA pEGFP-hBMP-4 mammalian plasmid (EGFP: Enhanced Green Fluorescent Protein) was constructed by subcloning techniques. bMSCs obtained from rabbits were cultured and transfected with either pEGFP-hBMP-4, pEGFP or left uninfected in vitro. Transfer efficiency was detected through the expression of EGFP. Transcription of the target gene was detected by RT-PCR. Alkaline phosphatase (ALP) and Von Kossa tests were also conducted to explore the phenotypes of osteoblasts. The autologous bMSCs of the 3 groups were then combined with Natural Non-organic Bone (NNB), a porous hydroxyapatite implant with a dimension of 6 mm x 6 mm x 3 mm, at a concentration of 5 x 10(7) cells/ml. They were subsequently implanted into 6 rabbits subcutaneously using NNB alone as a blank control (6 implants per group). Four weeks after surgery, the implants were evaluated with histological staining and computerized analysis of new bone formation.

RESULTSpEGFP-hBMP-4 expression plasmid was constructed. Under optimal conditions, gene transfer efficiency reached more than 30%. Target gene transfer could strengthen the transcription of BMP-4, and increase the expression of ALP as well as the number of calcium nodules. In the ectopic animal model, NNB alone could not induce new bone formation. The new bone area formed in the bMSCs group was (17.2 +/- 7.1)%, and pEGFP group was (14.7 +/- 6.1)%, while pEGFP-hBMP-4 group was (29.5 +/- 8.2)%, which was the highest among the groups (F = 7.295, P < 0.01).

CONCLUSIONSThe mammalian hBMP-4 expression plasmid was successfully constructed and a comparatively high transfer efficiency was achieved. The gene transfer technique enhanced the expression of BMP-4 and promoted differentiation from bMSCs to osteoblasts. These in vivo results suggested that transfection of bMSCs with hBMP-4 might be a suitable method to enhance their inherent osteogenic capacity for bone tissue engineering applications.

Animals ; Bone Marrow Cells ; cytology ; metabolism ; Bone Morphogenetic Protein 4 ; Bone Morphogenetic Proteins ; genetics ; Cell Differentiation ; Gene Transfer Techniques ; Genetic Therapy ; Humans ; Osteogenesis ; Plasmids ; Rabbits ; Stromal Cells ; cytology ; metabolism ; Tissue Engineering

OBJECTIVESTo analyze retrospectively the formation and histological changes of sclerosis rim in patients with osteonecrosis of the femoral head (ONFH), and to study the relationship between bone morphogenetic proteins (BMP4) and sclerosis rim, so as to acquire experimental and theoretical basis on individualized treatment for ONFH patients.

METHODSFrom November 2005 to November 2007, 184 hips of steroid-induced ONFH inpatients were collected. The mean age was (47 ± 7) years, the patients were divided into high (more than 54 years old), middle (40 - 54 years old) and low (less than 40 years old) age groups. Their clinical data were analyzed retrospectively according to gender and age. Parts of the femoral heads were selected for the study, including 18 hips in high age group, 11 hips in low age group and 20 hips in middle age group. Each 10 hips were selected with or without sclerosis rim. The femoral heads were cut along middle coronal plane, their weight-bearing and non-weight-bearing areas were used for the study. The specimens were processed by routine HE staining and picric acid-Sirius red staining and electron microscopy preparation and immunohistochemistry stain. The average optical density of BMP4 protein was calculated by image analysis software.

RESULTSThe trabecular of sclerosis rim was thickening and disorder. But its osteocytes were normal and with high secretion. The ratio of sclerosis rim was 71.4% (105/147) in middle age ONFH patients, which was significantly higher than the low age group patients (45.5%, 5/11) and high age group patients (38.5%, 10/26) (P < 0.01). The optical density of BMP4 in middle age ONFH patients was 0.32 ± 0.14, which was significantly higher than the low age group 0.20 ± 0.17 and high age patients 0.19 ± 0.27 (P < 0.05). The optical density was 0.16 ± 0.11 in ONFH patients without sclerosis rim, which was significantly lower than with sclerosis rim (0.28 ± 0.13) (P < 0.01). The time from hip pain to joint replacement in patients with sclerosis rim was (49 ± 11) months, and (15 ± 2) months without sclerosis rim. There was significant difference between the two groups (P < 0.01).

CONCLUSIONSThe formation of sclerosis rim is positively related to the expression of BMP4, and high expression of BMP maybe promote the formation of sclerosis rim.

Adult ; Bone Morphogenetic Protein 4 ; metabolism ; Female ; Femur Head ; metabolism ; pathology ; Femur Head Necrosis ; metabolism ; pathology ; Humans ; Male ; Middle Aged ; Retrospective Studies

OBJECTIVEBone marrow stromal cells (MSCs) were transfected with human bone morphogenetic protein-4 (hBMP-4) gene in vitro to provide BMP gene modified cells for tissue-engineered bone.

METHODSMSCs were cultured and transfected with pEGFP-hBMP4, pEGFP plasmids respectively or left uninfected as control. Transcription of BMP-4 gene as well as gene transfection efficiency was tested. Morphological and growth feature of the transfected cells were valued. Alkaline phosphatase (ALP), von Kossa, and Osteocalcin (OC) were tested to determine the phenotypes of osteoblast.

RESULTSThe gene transfection efficiency was 20%-30%, based on GFP expression. RT-PCR showed that MSCs had low transcription of BMP-4 that was enhanced by the gene transfer. Morphological feature of MSCs transfected with pEGFP-hBMP-4 changed but growth curves did not show much difference among the groups. In pEGFP-hBMP-4 group, ALP positive stain area and the number of calcium nodules were increased, as well as the expression of OC.

CONCLUSIONSA high transfer efficiency of MSCs was achieved under optimized conditions. The gene transfer technique strengthened the transcription of BMP-4 and promoted differentiation from MSCs to osteoblasts. hBMP-4 transferred MSCs may serve as an ideal cell source for tissue-engineered bone.

Animals ; Bone Marrow Cells ; cytology ; Bone Morphogenetic Protein 4 ; Bone Morphogenetic Proteins ; biosynthesis ; genetics ; Cell Differentiation ; Cells, Cultured ; Gene Transfer Techniques ; Osteoblasts ; cytology ; metabolism ; Osteocalcin ; biosynthesis ; genetics ; Osteogenesis ; Rabbits ; Stromal Cells ; cytology ; Tissue Engineering ; Transfection

Distraction osteogenesis is currently a standard method of bone lengthening. It is a viable method for the treatment of short extremities as well as extensive bone defects, because large amounts of bone can be regenerated in the distraction gap. echanical stimulation by distraction induces biological responses of skeletal regeneration that is accomplished by a cascade of biologic processes that may include differentiation of pluripotential tissue, angiogenesis, mineralization, and remodeling. There are complex interactions between bone-forming osteoblasts and other cells present within the bone microenvironment, particularly vascular endothelial cells that may be pivotal members of a complex interactive communication network in bone. Regenerate bone forms by three modes of ossification, which include intramembranous, enchondral, and transchondroid ossifications, although intramembraneous bone formation is the predominant mechanism of ossification. In this review we discussed the coupling between angiogenesis and mineralization, the biological and mechanical factors affecting them, the cellular and molecular events occurring during distraction osteogenesis, and the emerging modalities to accelerate regenerate bone healing and remodeling.
Animals ; Biological Markers ; Bone Morphogenetic Protein 2 ; Bone Morphogenetic Protein 4 ; Bone Morphogenetic Proteins/genetics/metabolism ; Bone and Bones/radiography/ultrastructure ; Calcification, Physiologic/*physiology ; Collagen/metabolism ; Cytokines/metabolism ; Growth Substances/metabolism ; Humans ; Neovascularization, Physiologic/*physiology ; Osteoblasts/physiology ; *Osteogenesis, Distraction ; *Transforming Growth Factor beta

The objective of this study was to investigate the expression of BMP-4 in stromal cells in vitro derived from human aorta-gonad-mesonephros (AGM) region. Stromal cells derived from human AGM region (hAGM S1-S5) and fibroblasts derived from human fetal trunk (hFT) were cultured in vitro. RT-PCR was used to analyze the expression of BMP-4 in hAGM S1-S5 stromal cells at mRNA level. And BMP-4 level was detected in the supernatant of hAGM S1-S5 stromal cells by ELISA assay. hFT cells were used as control group. The results showed that the heterogenous hAGM S1-S5 stromal cells displyed shapes of fibroblast-like and endothelial-like cells. hAGM S1-S5 stromal cells expressed BMP-4 mRNA, but fetal trunk fibroblasts (hFT) did not express BMP-4 mRNA. In the supernatant of hAGM S1-S5 cells, BMP-4 could be detected by ELISA assay ana its levels were statistically higher than that in hFT group (p < 0.05), while there was no significant difference between groups of hAGM S1-S5 (p > 0.05). It is concluded that human AGM-derived stromal cells in vitro express BMP-4, and the establishment of a new culture system based on the feeder cells of AGM stroma would promote the differention of embryonic stem cells into hematopoietic stem cells at a high proportion.
Aorta ; cytology ; embryology ; metabolism ; Bone Morphogenetic Protein 4 ; metabolism ; Cells, Cultured ; Gonads ; cytology ; embryology ; metabolism ; Hematopoietic Stem Cells ; cytology ; Humans ; Mesonephros ; cytology ; embryology ; metabolism ; RNA, Messenger ; metabolism ; Stromal Cells ; metabolism

OBJECTIVETo investigate the role of bone morphogenetic protein 4 (BMP4) on the proliferation and apoptosis in glioma stem cells.

METHODSStem cells were isolated from a human glioma cell line U87 by using vincristine and characterized by immunofluorescence assay. Proliferation and apoptosis were determined by soft agar colony assay and flow cytometry; Cyclin D1, Bcl-2 and Bax were detected by Western blot analysis.

RESULTSBMP4 inhibited cell proliferation and promoted apoptosis in U87 glioma stem cells. Moreover, Bcl-2 and Cyclin D1 expression were decreased by BMP4, while Bax level was elevated.

CONCLUSIONBMP4 can inhibit U87 glioma stem cells proliferation through downregulating Cyclin D1 level, and promote apoptosis through induction of Bax expression and inhibition of Bcl-2 level. It suggests that BMP4 plays an important role in human glioma stem cell biology.

Apoptosis ; Bone Morphogenetic Protein 4 ; genetics ; metabolism ; Cell Line, Tumor ; Cell Proliferation ; Cyclin D1 ; genetics ; metabolism ; Gene Expression Regulation, Neoplastic ; Glioma ; genetics ; metabolism ; physiopathology ; Humans ; Neoplastic Stem Cells ; cytology ; metabolism ; Proto-Oncogene Proteins c-bcl-2 ; genetics ; metabolism