Use of Erythropoietin as adjuvant therapy in nerve reconstruction.

BACKGROUND AND AIMS: Adjuvant therapies may improve the outcome after nerve reconstruction. We analyzed the influence of recombinant human Erythropoietin (rHuEpo), which has proven angiogenic and neuroprotective effects, on the quality of peripheral nerve regeneration. METHODS: Thirty two female Lewis rats underwent nerve reconstruction by means of tubulization (groups I and II) or autologous sciatic nerve grafting (groups III and IV). Groups I and III received daily subcutaneous rHuEpo injections over 2 weeks (1,000 U/kg bw) with normal saline injections as controls (groups II and IV). Data on histology and muscle weight were collected after 7 weeks. Axon count and diameter were assessed by a new method based on digital segmentation. RESULTS: Atrophy of the tibial muscle was less severe in the rHuEpo-treated group compared to controls resulting in significant higher muscle weight quotients (p = 0.006). The same trend was found in the gastrocnemius muscle, but without being statistically significant. No significant differences in axon count or axon diameter were detected in the presence of rHuEpo treatments. CONCLUSION: Our findings give evidence for a positive effect of Erythropoietin on functional recovery after nerve grafting. Muscle recovery benefited from rHuEpo administration despite absence of improved neural morphology. Semi-automated axon detection facilitated accurate morphometrical assessment.

VEGF(165) and bFGF protein-based therapy in a slow release system to improve angiogenesis in a bioartificial dermal substitute in vitro and in vivo.

BACKGROUND: Angiogenesis can be enhanced by several growth factors, like vascular endothelial growth factor-165 (VEGF(165)) and basic fibroblast growth factor (bFGF). Delayed release of such growth factors could be provided by incorporation of growth factors in fibrin matrices. In this study, we present a slow release system for VEGF(165) and bFGF in fibrin sealant. MATERIALS AND METHODS: In vitro: Pieces of Integratrade mark matrix of 15 mm in diameter were prepared. Integratrade mark matrices were divided into four groups (A=control; B=fibrin sealant; C=fibrin sealant+growth factors; D=growth factors). In vivo: The bioartificial dermal templates were transplanted into a full-skin defect of the back of nu-nu mice. Four different groups included each six matrices at 2 and 4 weeks. RESULTS: In vitro: In groups C and D, continuous release of VEGF(165) and bFGF was eminent. The incorporation of growth factors into fibrin sealant evoked a prolonged growth factor release (p < 0.05). In vivo: A significantly higher amount of vessels was quantified in groups C and D compared to groups A and B (p < 0.001). CONCLUSIONS: A model of slow protein release by combining VEGF(165) and bFGF with fibrin sealant was produced. This model resulted in a prolonged bioavailability of growth factors in vivo for functional purposes. Fibrin and collagen can release growth factors in vivo and induce significant and faster neovascularisation in bioartificial dermal templates.

A technique to detect and to quantify fasciocutaneous blood vessels in small laboratory animals ex vivo.

PURPOSE: A microangiographical technique is described, which allows visualization of small and capillary blood vessels and quantification of fasciocutaneous blood vessels by means of digital computer analysis in very small laboratory animals. MATERIALS AND METHODS: The left carotid artery of 20 nu/nu mice was cannulated (26 gauge) and a mixture of gelatin, bariumsulfate, and green ink was injected according to standardized protocol. Fasciocutaneous blood vessels were visualized by digital mammography and analyzed for vessel length and vessel surface area as standardized units [SU] by computer program. RESULTS: With the described microangiography method, fasciocutaneous blood vessels down to capillary size level can be clearly visualized. Regions of interest (ROIs) can be defined and the containing vascular network quantified. Comparable results may be obtained by calculating the microvascular area index (MAI) and the microvascular length index (MLI), related to the ROIs size. Identical ROIs showed a high reproducibility for measured [SU] < 0.01 +/- 0.0012%. CONCLUSION: Combining microsurgical techniques, pharmacological knowledge, and modern digital image technology, we were able to visualize small and capillary blood vessels even in small laboratory animals. By using our own computer analytical program, quantification of vessels was reliable, highly reproducible, and fast.

[A microangiography technique to quantify fasciocutaneous blood vessels in small laboratory animals]. / Eine mikroangiographische Technik zur Quantifizierung fasziokutaner Blutgefässe am kleinen Versuchstier.

PURPOSE: A microangiographic technique is described, which allows visualization of small blood vessels with a diameter of approximately 20 microm and quantification of fasciocutaneous blood vessels by means of digital computer analysis in very small laboratory animals. METHOD: The left carotid artery of 45 nu/nu mice was cannulated (26 gauge) and a mixture of gelatine, barium sulfate and green ink was injected according to standardized protocol. Fasciocutaneous blood vessels were visualized by digital mammography and analyzed for vessel length and vessel surface area as standardized units (pixel) by computer program. RESULTS: With the described microangiography method fasciocutaneous blood vessels can be clearly visualized. Regions of interest (ROIs) can be defined and the containing vascular network quantified. Identical ROIs showed a high reproducibility for measured unit (pixel) < 6.5 +/- 2.4 %. By the use of digital image, processing the quantification of vessels was reliable, reproducible and fast. CONCLUSION: Combining microsurgical techniques, pharmacological knowledge and modern computer imaging analysis systems, we were able to visualize and quantify blood vessels with a diameter of approximately 20 microm even in small laboratory animals.