The factors controlling the microstructure and properties of collagen-based bioactive artificial dermis are reviewed. The second component, the pore diameter and porosity, the thickness of scaffold, the bioactive factors as well as the cross-linking density that are important parameters of artificial dermis should be carefully researched and designed. Experiment methods controlling these parameters are suggested.
Biocompatible Materials ; chemistry ; Collagen ; chemistry ; Dermis ; cytology ; Humans ; Porosity ; Skin, Artificial ; Tissue Engineering ; methods

OBJECTIVETo observe the effects of Angelica dahurica extracts on the biological characteristics of human dermal fibroblasts in vitro and to preliminary explore its possible therapeutic mechanism for wound healing.

METHODSThe optimal concentration of Angelica dahurica extracts was identified by analysing of proliferation activity of human normal fibroblasts (Fb) that treated with different concentration of Angelica dahurica extracts through thiazole blue (MTT) colorimetric assay. Cell cycle, collagen I and collagen III mRNA levels of the optimal Angelica dahurica extracts treated Fb were detected by flow cytometry (FCM) and real-time PCR techniques.

RESULTSAt concentrations of 5 × 10(-4) to 5 × 10(-2) g/L, the Angelica dahurica extracts significantly enhanced the proliferation of Fb. The most significant concentration was 5 × 10(-3) g/L (t = 5.79, P < 0.01), at which an increased percentage of G1 to S and S to G2 phase cells (t = 11.2, 5.69, 2.44, P < 0.05) as well as an increased level of collagen I (1.61 ± 0.26 vs. 1.00 ± 0.16) and collagen III mRNA (3.36 ± 0.40 vs. 1.00 ± 0.14) were obtained compared to the control group (t = 6.69, 7.64, P < 0.01).

CONCLUSIONSAngelica dahurica extracts can notably promote the proliferation of Fb and accelerating the cell cycle of Fb as well as up-regulating the expression of collagen I and collagen III, which may enhance the process of wound healing.

Angelica ; chemistry ; Cell Cycle ; drug effects ; Cell Proliferation ; drug effects ; Cells, Cultured ; Collagen ; metabolism ; Dermis ; cytology ; Fibroblasts ; cytology ; drug effects ; metabolism ; Humans ; Plant Extracts ; pharmacology

OBJECTIVETo investigate the effects of preemptive freezing with different temperature and cross-linking methods on the ultrastructure of collagen membrane and its influence on human fibroblast proliferation.

METHODSBovine collagen type I solution in concentration of 10 g/L was preliminarily frozen at -20 degrees C or - 80 degrees C for 12 hours, and lyophilized at -70 degrees C for 48 hours. The diameter of apertures in collagen membranes prepared with two different preliminary temperatures were observed by scanning electron microscope (SEM) and compared. The preliminary freezing temperature of - 80 degrees C was used for the following study. The apertures of collagen membrane performed with cross-linking glutaraldehyde and ultraviolet (UV) radiation cross-linking with glutaraldehyde (double cross - linking) after preliminary freezing were also compared. The proliferation of human fibroblasts inoculated in above cross-linking collagens were assessed by MTT assay, in terms of absorption value.

RESULTSThe mean diameter of apertures of collagen membrane pre-frozen at -20 degrees C was (172 +/- 374 microm, while that at -80 degrees C was (99 +/- 24) microm. The apertures of collagen membrane were reduced in size after glutaraldehyde cross-linking, while those of double cross-linking showed no change in size. There was obvious difference in absorption value of fibroblasts 8 days after seeding between above two cross-linking methods (1.534 +/- 0.013 for glutaraldehyde cross-linking, 3.778 +/- 0.010 for double cross-linking, P < 0.05).

CONCLUSIONThe collagen membrane after preliminary freezing at - 80 degrees C and double cross-linking with UV radiation and glutaraldehyde may be used as a dermal skeleton substitute.

Animals ; Cattle ; Cells, Cultured ; Coculture Techniques ; Collagen ; ultrastructure ; Cross-Linking Reagents ; chemistry ; Dermis ; cytology ; Fibroblasts ; cytology ; drug effects ; radiation effects ; Freezing ; Glutaral ; chemistry ; Humans ; Skin, Artificial ; Tissue Scaffolds ; Ultraviolet Rays

Apoptosis is regulated by interaction of antiapoptotic Bcl-2 family proteins with various proapoptotic proteins, several of which are also members of the Bcl-2 family. BNIP3 (formerly NIP3) is a proapoptotic mitochondrial protein classified in the Bcl-2 family based on limited sequence homology-3 (BH3) domain and COOH-terminal transmembrane domain. Sequence comparison of BNIP3 has indicated that there are several BNIP3 human homologs of this protein, like BNIP3L, Nix and BNIP3. We have cloned a new member of BNIP3 family from the cDNA library prepared from human dermal papilla cells and designated as BNIP3h. BNIP3h shows substantial homology with other BNIP3 family proteins. BNIP3h induced apoptosis from 24 hours after transfection in MCF7 cell lines and its apoptosis inducing activity is extended until 72 hours after transfection.
Amino Acid Sequence ; Apoptosis/*physiology ; Base Sequence ; Cells, Cultured ; Cloning, Molecular ; Dermis/chemistry/cytology ; Human ; Membrane Proteins/chemistry/*genetics/metabolism ; Mitochondria/chemistry ; Molecular Sequence Data ; Multigene Family ; Sequence Alignment ; Tissue Distribution ; Transfection ; Tumor Cells, Cultured

Nerve growth factor (NGF) can promote the regeneration of peripheral nerve as well as contraction and reepithelization of wound. We constructed a bioengineered dermis containing microencapsulated NGF-expressing NIH-3T3 cells and study the effect of the microencapsule to the bioengineered dermis and seed cells. NGF gene was transfected into NIH-3T3 cells and enclosed into alginate-poly-L-lysine-alginate (APA) microencapsulation and cultivated in vitro. Content of NGF in microencapsules culturing supernatant was measured by enzyme linked immunosorbent assay (ELISA) method. These microencapsules were co-cultured with epidermic cells and fibroblast cells. Bioengineered dermis was constructed with NGF-expressing micorencapsules as seed cells using tissue engineering method. NIH-3T3 microencapsules, empty microencapsules, normal culture media were control groups. After one week culture, the characteristics of the dermis were described by MTT test, the content of hydroxyproline (HP), HE staining and ultrastructure photograph. We found the NGF-expressing microencapsulates can secret NGF steadly after cultured 8w in vitro, promot the proliferation of epidermic cells and secret collagen of fibroblast cells. These functions can maintaine in bioengineered dermis. So NGF-expressing NIH-3T3 microencapsulates can promote the quality of bioengineered dermis.
Alginates ; chemistry ; Animals ; Biocompatible Materials ; chemistry ; Cell Proliferation ; Dermis ; cytology ; Gene Expression Regulation ; Mice ; NIH 3T3 Cells ; Nerve Growth Factor ; genetics ; Polylysine ; analogs & derivatives ; chemistry ; Skin Physiological Phenomena ; Tissue Engineering ; methods ; Transfection ; methods

Regeneration of periodontal tissue is the most promising method for restoring periodontal structures. To find a suitable bioactive three-dimensional scaffold promoting cell proliferation and differentiation is critical in periodontal tissue engineering. The objective of this study was to evaluate the biocompatibility of a novel porcine acellular dermal matrix as periodontal tissue scaffolds both in vitro and in vivo. The scaffolds in this study were purified porcine acellular dermal matrix (PADM) and hydroxyapatite-treated PADM (HA-PADM). The biodegradation patterns of the scaffolds were evaluated in vitro. The biocompatibility of the scaffolds in vivo was assessed by implanting them into the sacrospinal muscle of 20 New Zealand white rabbits. The hPDL cells were cultured with PADM or HA-PADM scaffolds for 3, 7, 14, 21 and 28 days. Cell viability assay, scanning electron microscopy (SEM), hematoxylin and eosin (H&E) staining, immunohistochemistry and confocal microscopy were used to evaluate the biocompatibility of the scaffolds. In vitro, both PADM and HA-PADM scaffolds displayed appropriate biodegradation pattern, and also, demonstrated favorable tissue compatibility without tissue necrosis, fibrosis and other abnormal response. The absorbance readings of the WST-1 assay were increased with the time course, suggesting the cell proliferation in the scaffolds. The hPDL cells attaching, spreading and morphology on the surface of the scaffold were visualized by SEM, H&E staining, immnuohistochemistry and confocal microscopy, demonstrated that hPDL cells were able to grow into the HA-PADM scaffolds and the amount of cells were growing up in the course of time. This study proved that HA-PADM scaffold had good biocompatibility in animals in vivo and appropriate biodegrading characteristics in vitro. The hPDL cells were able to proliferate and migrate into the scaffold. These observations may suggest that HA-PADM scaffold is a potential cell carrier for periodontal tissue regeneration.
Absorbable Implants ; Acellular Dermis ; Animals ; Biocompatible Materials ; chemistry ; Cell Adhesion ; physiology ; Cell Culture Techniques ; Cell Movement ; physiology ; Cell Proliferation ; Cell Shape ; physiology ; Cell Survival ; physiology ; Durapatite ; chemistry ; Fibrosis ; Humans ; Microscopy, Electron, Scanning ; Muscle, Skeletal ; surgery ; Necrosis ; Periodontal Ligament ; cytology ; surgery ; Rabbits ; Regeneration ; physiology ; Swine ; Tissue Engineering ; methods ; Tissue Scaffolds ; chemistry