OBJECTIVETo explore transient expression of the eukaryotic expression plasmid carrying human platelet-derived growth factor-B (hPDGF-B) in gingival fibroblasts of Beagle dog.

METHODSPlasmid carrying hPDGF-B (EX-A0380-M03) was amplified and identified, and then transfected into gingival fibroblasts of Beagle dog. Reverse transcription polymerase chain reaction (RT-PCR), immunocytochemistry, enzyme-linked immunosorbent assay (ELISA) and Western bolt were choose to detect the expression of hPDGF-B.

RESULTSTarget gene carried by EX-A0380-M03 was hPDGF-B. Green fluorescence protein (GFP) expressed by transfected gingival fibroblasts was observed under inverted phase contrast fluorescence microscope (IPCFM) (after 24 hours) and the transfection efficiency was 18%-38% (after 48 hours). Serials other methods (RT-PCR, immunocytochemistry, and ELISA) mentioned above also convinced that cells expressed hPDGF-B, and the protein that was a kind of fusion protein composed of PDGF-BB and GFP was identified by Western blot.

CONCLUSIONEukaryotic expression plasmid carrying hPDGF-B was transfected into gingival fibroblasts successfully, and a kind of fusion protein was expressed.

Animals ; Dogs ; Fibroblasts ; Gingiva ; Plasmids ; Platelet-Derived Growth Factor ; Proto-Oncogene Proteins c-sis ; Transfection

No abstract available.
Osteoblasts* ; Platelet-Derived Growth Factor*

OBJECTIVETo investigate the synergistic effect of transforming growth factor-beta1 (TGF-beta1) and platelet-derived growth factor-BB (PDGF-BB) on the expression of integrin beta3, in periodontal membrane of rat orthodontic tooth.

METHODSAn orthodontic tooth movement model was established. Up to 32 experimental rats were randomly divided into four groups according to a random number table. The four groups were injected with 1% PBS, TGF-beta1 (5 ng), PDGF-BB (10 ng), and combined TGF-beta1 (5 ng) and PDGF-BB (10 ng) in the buccal submucosal, respectively. The volume injected in each group was 0.1 mL. The animals were then sacrificed on the 10th day. The left maxillary first molar and periodontal tissue were taken. Different expressions of integrin beta3 were detected in periodontal tissues through immunohistochemistry. Mean optical density (OD) values of the positive fields were examined. The data obtained were analyzed through ANOVA. The data followed normal distribution, and were compared via t-test.

RESULTSCompared with the control groups, the expression of integrin beta3 was higher in the experimental groupin tension sides (P < 0.01). Significant differences in tension sides between the single-injection groups and the combined group were observed (P < 0.01). Compared with the control groups, the expression of integrin beta3 was higher in the experimental group in compression sides (P < 0.05). In addition, there was no significant differences in compression sides between the single-injection groups and the combined group (P > 0.05).

CONCLUSIONIn terms of local regulatory factors, TGF-beta1 combined with PDGF-BB enhance the expression of integrin beta3 in the periodontal membrane and accelerate periodontal remodeling. The synergistic effect of the two growth factors is better than the single growth factor.

Animals ; Integrin beta3 ; Molar ; Periodontal Ligament ; Platelet-Derived Growth Factor ; Proto-Oncogene Proteins c-sis ; Rats ; Tooth Movement Techniques ; Transforming Growth Factor beta1

OBJECTIVETo study the expression and significance of platelet derived growth factor (PDGF) and its receptor (PDGFR) in liver tissues of patients with chronic hepatitis fibrosis and liver cirrhosis.

METHODSThe expression, distribution, quantitation, and correlation of PDGF-A, PDGF-B, PDGFR-alpha, PDGFR-beta, and alpha-SMA in the liver tissues were analyzed by immunohistochemical techniques in 21 patients with chronic hepatitis and 42 patients with liver cirrhosis.

RESULTSIn the liver tissues of chronic hepatitis and liver cirrhosis, PDGF and its receptor and alpha-SMA mainly distributed in the fibrotic septa and the infiltration area of inflammation, particularly in branch spindle-shaped cells (activated HSC). The expression of PDGF-B and PDGFR-beta was stronger than that of PDGF-A and PDGFR-alpha with a significant difference between them (P<0.05 approximately 0.01). The expression and distribution of alpha-SMA was basically identical with the expression and distribution of PDGF-A, PDGF-B and PDGFR-alpha, PDGFR-beta and quantitative analysis showed a positive correlation (r=0.606, P<0.001).

CONCLUSIONSPDGF and PDGFR play a key role in liver fibrogenesis and development. The biologic effects of PDGF are elicited through activising HSC. Inhibiting PDGF and its receptor is a new approach to the treatment of liver fibrosis.

Actins ; biosynthesis ; physiology ; Adolescent ; Adult ; Aged ; Female ; Hepatocytes ; metabolism ; Humans ; Liver Cirrhosis ; metabolism ; Male ; Middle Aged ; Muscle, Smooth ; chemistry ; Platelet-Derived Growth Factor ; biosynthesis ; physiology ; Proto-Oncogene Proteins c-sis ; biosynthesis ; physiology ; Receptor, Platelet-Derived Growth Factor alpha ; biosynthesis ; physiology ; Receptor, Platelet-Derived Growth Factor beta ; biosynthesis ; physiology

BACKGROUND: Platelet-rich plasma (PRP) has more concentrated platelets than normal plasma (approximately 150-400x10(3) cell/dL). Platelets excrete several growth factors and cytokines that are associated with the healing and regeneration process. However, even though PRP is widely used, the mechanism or actual effect is presently unclear. Therefore, this study was performed to investigate the levels of growth factors and platelet concentration rate. METHODS: Autologous blood for preparing PRP was obtained from healthy subjects aged 25 to 35 years. The samples were divided into 4 experimental groups (inactivated whole blood, inactivated PRP, activated whole blood with thrombin and calcium chloride, and activated PRP). The platelet counts in the blood were analyzed and the growth factors were quantitatively measured. A statistical analysis was performed by using Dunn's multiple comparison test. RESULTS: In the blood cell analysis, the platelet count of the PRP group was approximately 4.25 times higher than that of the whole blood group. In the quantitative analysis of growth factors, the platelet-derived growth factor (PDGF)-AB, PDGF-BB, and transforming growth factor-beta of the inactivated and activated PRP groups were higher than those of the inactivated and activated whole blood groups (P<0.05). CONCLUSIONS: In this study, the platelet count and the levels of PDGF-AB and PDGF-BB in the PRP were determined. Further, more research is required on the bioactivity level of the growth factors secreted during the process of PRP preparation and the potency of growth factors that can be exerted physiologically in vivo.
Aged ; Blood Cells ; Blood Group Antigens ; Blood Platelets ; Calcium Chloride ; Cytokines ; Humans ; Intercellular Signaling Peptides and Proteins ; Plasma ; Platelet Count ; Platelet-Derived Growth Factor ; Platelet-Rich Plasma ; Proto-Oncogene Proteins c-sis ; Regeneration ; Thrombin ; Transforming Growth Factors

Humans ; Platelet-Derived Growth Factor ; Pulmonary Fibrosis

OBJECTIVETo acquire sufficient PDGF-BB protein and provide the basis for the further studies of its role on the fracture healing and trauma reconstruction and its clinical applications.

METHODSConstructed the prokaryotic expression vector pQE-PDGF-B with the gene rearrangement technique, and the monomeric form of recombinant PDGF-B expressed in E. coli M15.

RESULTSPDGF-B mature peptide gene was inserted into prokaryotic expression vector pQE30, which was confirmed by PCR, enzyme digestion and sequencing identification; the expressed products of pQE-PDGF-B in E. coli showed a single protein on SDS-PAGE, and their expression level was about 15% of the total bacterial protein. The molecular weight of the purified PDGF-B protein was about 15 KDs on SDS-PAGE.

CONCLUSIONSThe construction of recombinant plasmid and preparation of the monomeric protein of PDGF-B provides a solid foundation for further studying the function of PDGF-BB and producing biologically PDGF-BB protein.

Cloning, Molecular ; Escherichia coli ; genetics ; metabolism ; Genetic Vectors ; Humans ; In Vitro Techniques ; Platelet-Derived Growth Factor ; biosynthesis ; genetics ; Proto-Oncogene Proteins c-sis ; genetics ; Recombinant Proteins ; biosynthesis ; genetics ; Transfection

Adolescent ; Adult ; Biomarkers ; Female ; Hepatitis B, Chronic ; metabolism ; Humans ; Liver ; chemistry ; Liver Cirrhosis ; blood ; therapy ; Male ; Middle Aged ; Platelet-Derived Growth Factor ; analysis ; Proto-Oncogene Proteins c-sis

Adult ; Female ; Hepatitis B e Antigens ; blood ; Hepatitis B, Chronic ; blood ; Humans ; Male ; Middle Aged ; Platelet-Derived Growth Factor ; metabolism ; Proto-Oncogene Proteins c-sis ; Young Adult

Cell Line ; Epidermal Growth Factor ; biosynthesis ; genetics ; Fibroblast Growth Factor 2 ; biosynthesis ; genetics ; Hepatocytes ; metabolism ; Humans ; Interleukin-10 ; pharmacology ; Platelet-Derived Growth Factor ; pharmacology ; Proto-Oncogene Proteins c-sis ; RNA, Messenger ; biosynthesis ; genetics