Xianglin Hou

Regeneration of uterine horns in rats by collagen scaffolds loaded with collagen-binding human basic fibroblast growth factor.

Severe damages of uterine endometrium which prevent embryos from implantation and placentation finally often result in infertility or pregnant complications. There is lack of effective treatments due to the limitation of native materials available and complexity of the function and internal environment of uterus. In the present study, a collagen targeting basic fibroblast growth factor (bFGF) delivery system was constructed by a collagen membrane loaded with bFGF fused a collagen-binding domain (CBD) to the N-terminal which limits the diffusion of bFGF from collagen. We tested the bFGF delivery system in rats under the severe uterine damage model (partial rat uterine horn excision/reconstruction), and found this delivery system improved regeneration abilities of uterine endometrium and muscular cells, improved vascularization, as well as better pregnancy outcomes in rats. Therefore, this targeting delivery system may be an effective strategy for uterine tissue regeneration.

Regeneration of full-thickness abdominal wall defects in rats using collagen scaffolds loaded with collagen-binding basic fibroblast growth factor

Biomaterials are increasingly used in the repair of tissue defects. The aim of the present study was to evaluate a new composite biomaterial for reconstruction of a 2 × 2.5 cm full-thickness abdominal wall defect. In this study, the collagen membrane was activated with the engineered human basic fibroblast growth factor (bFGF). To enhance the binding of bFGF to collagen membranes, a specific peptide of collagen-binding domain (CBD) was fused to the N-terminal of bFGF. After implantation, little adhesion was caused in collagen/CBD-bFGF, collagen/NAT-bFGF and collagen/PBS groups. Moreover, collagen/CBD-bFGF group could effectively promote the vascularization at 30 d after surgery and significantly accelerate the integration of myofibers into the collagen material at 90 d after surgery compared to the other two groups. Due to the replacement of the myofibers in materials, the mechanical strength of implanted biomaterials in collagen/CBD-bFGF group was also greater than the other two groups at 90 d after surgery. Thus, the collagen/CBD-bFGF composite biomaterial was promising for the treatment of full-thickness abdominal wall defect.

Linear Ordered Collagen Scaffolds Loaded with Collagen-Binding Neurotrophin-3 Promote Axonal Regeneration and Partial Functional Recovery after Complete Spinal Cord Transection

Neurotrophin-3 (NT3) is an important neurotrophic factor for spinal cord injury (SCI) repair. However, constant exchange of cerebrospinal fluid often decreases the effective dosage of NT3 at the targeted injury site. In the present study, a recombinant collagen-binding NT3 (CBD-NT3), consisting of a collagen-binding domain (CBD) and native NT3, was constructed. Linear rat-tail collagen (LRTC) was used as a physical carrier for CBD-NT3 to construct a LRTC/C3 system. The collagen-binding ability of CBD-NT3 was verified, and the bioactivity of CBD-NT3 was assayed with neurite outgrowth of dorsal root ganglia (DRG) explants and DRG cells in vitro. After complete spinal cord transection in rats, LRTC/CBD-NT3 or the LRTC/NT3 system was transplanted into the injury site. Hindlimb locomotion recovery was closely observed using the Basso-Beattie-Bresnahan (BBB) locomotor rating scale and the grid walk test. Significant improvement was observed in the LRTC/CBD-NT3 group. The results of regenerating nerve fiber and anterograde tracing of biotinylated dextran amine (BDA)-labeled corticospinal tract (CST) fibers demonstrated axonal regeneration of LRTC/CBD-NT3 in the injured spinal cord. Serotonin fiber regrowth also illustrated the effectiveness of LRTC/CBD-NT3. Thus, collagen-binding NT3 with LRTC may provide an effective method for treating SCI.

Promotion of neuronal differentiation of neural progenitor cells by using EGFR antibody functionalized collagen scaffolds for spinal cord injury repair

The main challenge for neural progenitor cell (NPC)-mediated repair of spinal cord injury (SCI) is lack of favorable environment to direct its differentiation towards neurons rather than glial cells. The myelin associated inhibitors have been demonstrated to promote NPC differentiation into glial lineage. Herein, to inhibit the downstream signaling activated by myelin associated inhibitors, cetuximab, an epidermal growth factor receptor (EGFR) neutralizing antibody, functionalized collagen scaffold has been developed as a vehicle for NPC implantation. It was found that collagen-cetuximab 1 μg scaffolds enhanced neuronal differentiation and inhibited astrocytic differentiation of NPCs exposed to myelin proteins significantly in vitro. To test the therapeutic effect in vivo, NPCs expressing green fluorescent protein (GFP)-embedded scaffolds have been implanted into the 4 mm-long hemisection lesion of rats. We found that the collagen-cetuximab 5 μg scaffolds induced neuronal differentiation and decreased astrocytic differentiation of NPCs, enhanced axon regeneration, and promoted functional recovery markedly. A well-functionalized scaffold was constructed to improve the recovery of SCI, which could promote the neuronal differentiation of neural progenitor cells in vivo.

Bladder Regeneration by Collagen Scaffolds With Collagen Binding Human Basic Fibroblast Growth Factor

PURPOSE Studies show that basic fibroblast growth factor can promote bladder regeneration. However, the lack of targeting delivery approaches limits its clinical application. We investigated a collagen based targeting system for bladder regeneration. A collagen binding domain was added to the native basic fibroblast growth factor N-terminal to allow it to bind to collagen. MATERIALS AND METHODS Sprague-Dawley rats underwent partial cystectomy. Collagen scaffolds loaded with collagen binding domain basic fibroblast growth factor, native basic fibroblast growth factor or phosphate buffered saline were grafted to the remaining host bladders, respectively. At days 30 and 90 reconstructed bladders were evaluated by histological analysis and urodynamics. RESULTS This targeting basic fibroblast growth factor delivery system induced satisfying bladder histological structures. It promoted more vascularization and smooth muscle cell ingrowth. Urodynamics revealed well accommodated bladder tissue with volume capacity and compliance. CONCLUSIONS Results show that the targeting delivery system consisting of collagen binding domain basic fibroblast growth factor and collagen membranes induced better bladder regeneration at the injury site. Thus, this targeting delivery system may be an effective strategy for bladder regeneration with potential clinical applications.

The osteogenic effect of bone morphogenetic protein-2 on the collagen scaffold conjugated with antibodies

Considerable research has been focused on the exploration of bone morphogenetic protein-2 (BMP(2)) delivery vehicles for achieving prolonged availability and maintaining efficient local concentration at the bone injury sites. In this study, heterobifunctional cross-linkers Sulfo-SMCC and cyclic thioimidate compound Trauts Reagent were used to conjugate monoclonal polyhistidine antibody on collagen scaffold demineralized bone matrix (DBM) to create specific binding between BMP(2) containing six histidines tag (His-BMP(2)) and DBM. According to the optimized cross-linking reagent concentration, more polyhistidine antibodies conjugated on DBM with 5mg/ml Trauts Reagent and 25ug/ml Sulfo-SMCC than physical adsorption. Monoclonal antibodies conjugated DBM (MAbs-DBM) could bind more His-BMP(2) than DBM and achieved controlled release in vitro. The alkaline phophatase (AP) activity of C2C12 cells on MAbs-DBM indicated that His-BMP(2) retained on MAbs-DBM preserved the function to induce osteogenic differentiation. His-BMP(2)/MAbs-DBM induced more ectopic bone formation (AP activity assay and histochemistry stain) than control group after subcutaneous implantation. The results demonstrated that antibody-collagen system could be useful for maintaining higher local therapy concentration of growth factors at the injury sites.

Acceleration of diabetic wound healing by collagen-binding vascular endothelial growth factor in diabetic rat model

AIMS Vascular endothelial growth factor (VEGF) is an important active protein for the induction of angiogenesis and plays an important role in the tissue regeneration of diabetic wounds. In this study, we used collagen-binding VEGF in a diabetic rat model to investigate the effects of this new method. METHODS We produced a fusion protein (CBD-VEGF) consisting of VEGF and a collagen-binding domain (CBD), which allowed VEGF to bind to collagen. The diabetic rat models were made by injected streptozocin (STZ) peritoneally and removed full thickness skin on the back. All the rats were randomly divided into 3 groups: PBS group (n=24), NAT-VEGF group (n=24), and CBD-VEGF group (n=24). After model establishment, the dissolved drugs were evenly given on the wounds using syringe. The healing rates were calculated and compared among the groups and the tissues of the wound were taken and evaluated for histological analysis. RESULTS The CBD-VEGF group showed better result in wound healing rate, better vascularization and higher amount of VEGF in the wound granulation tissue compared with NAT-VEGF group and control. CONCLUSIONS Topical application of CBD-VEGF can promote diabetic wound healing in rat model, which could potentially provide a better therapeutic option for diabetic wounds.

The effect of collagen-binding vascular endothelial growth factor on the remodeling of scarred rat uterus following full-thickness injury.

Serious injuries of uterine which lead to scar formation will finally result in infertility or pregnancy complications. There are few effective methods to treat such damages because of the shortage of native tissues. Vascular endothelial growth factor (VEGF) is important for the formation of new vessels and re-epithelialization of endometrium. Here we produced a collagen-binding VEGF by fusing a collagen-binding domain to the N-terminal of native VEGF. After injection into a rat scarred uterus model (partial of rat uterine horn was excised and left for scar formation) the collagen targeting VEGF promoted remodeling of the scarred uterus including the regeneration of endometrium, muscular cells, and vascularization and improved pregnancy outcomes. Thus, collagen-binding VEGF may be a pragmatic solution for the treatment of severe uterine damages.

Extrahepatic bile duct regeneration in pigs using collagen scaffolds loaded with human collagen-binding bFGF

Extrahepatic bile duct defects and their complications are benign lesions but with malignant outcomes. Extrahepatic bile duct regeneration at the injury site could be important for the repair. In our previous work, a human basic fibroblast growth factor (bFGF) fused with a collagen-binding domain (CBD) was produced to activate the collagen membrane to obtain targeted tissue regeneration. This collagen/growth factor functional biomaterial could promote the regeneration of skin, bladder and full-thickness abdominal wall by accelerating vascularization and cellularization of autologous tissues. We speculate that the functional biomaterial could also provide the repairing effect on extrahepatic bile duct injuries. Using a pig extrahepatic bile duct injury model, we found that the collagen/CBD-bFGF composite biomaterial could significantly promote the extrahepatic bile duct regeneration at the injury site without causing structure deformation or hepatic dysfunction during both short- and long-time observations.

Promotion of diabetic wound healing by collagen scaffold with collagen-binding vascular endothelial growth factor in a diabetic rat model

Aims: Studies show that VEGF can promote tissue regeneration in diabetic wounds. The aim of this study was to evaluate the effects of a new composite biomaterial, a collagen scaffold with CBD‐VEGF, for wound healing in a diabetic rat model. Materials and methods: We produced a collagen scaffold loaded with CBD‐VEGF, which allowed VEGF to bind to the collagen scaffold. The diabetic rat model was constructed by injecting streptozocin (STZ) peritoneally and removing a 2 x 2.5 cm thick slice of skin from the back of the animal. Animals were randomly divided into 4 groups: blank control (BC Group, n = 24), collagen scaffold loaded with PBS (PBS Group, n = 24), collagen scaffold loaded with NAT‐VEGF (NAT‐VEGF Group, n = 24), and collagen scaffold loaded with CBD‐VEGF (CBD‐VEGF Group, n = 24). Wounds of the BC Group were covered with gauze and those of the PBS, NAT‐VEGF and CBD‐VEGF Groups were grafted by corresponding collagen scaffolds, respectively. Healing rates were calculated and compared among groups. Wound tissue was evaluated by histologic analysis. Results: The CBD‐VEGF group showed a higher wound healing rate, better vascularization and higher level of VEGF in the granulation tissue wound compared with NAT‐VEGF and PBS groups. Conclusions: The collagen scaffold with CBD‐VEGF promoted wound healing in a diabetic rat model, which could potentially provide better therapeutic options for the treatment of diabetic wounds. Copyright