Evaluation of hyaluronic matrix efficacy in sinus augmentation: a randomized-controlled histomorphometric and micro-computed tomography analysis.

The objective of the present study was to test the hypothesis that the addition of hyaluronic acid-based matrix to collagenated heterologous bone graft for sinus augmentation would enhance bone formation compared to collagenated heterologous bone graft alone in the early healing period, by micro-computed tomography and histomorphometry. Thirteen systemically healthy patients requiring bilateral two-stage maxillary sinus augmentation (residual crest height≤4mm) were enrolled in this split-mouth prospective randomized controlled study. One sinus side as a control group was grafted with only collagenated heterologous bone graft; the other region as a test group was grafted with hyaluronic matrix and collagenated heterologous bone graft. Bone biopsy samples were taken after 4 months during the dental implant surgery and analyzed using micro-computed tomography and histomorphometric parameters. According to the micro-computed tomography and histomorphometric results, a significantly higher percentage of new bone was observed in the test group when compared to the control group after 4 months of healing. This study confirmed the hypothesis that the addition of hyaluronic matrix to collagenated heterologous bone graft for sinus augmentation enhances bone formation compared to collagenated heterologous bone graft alone in the early healing period.

Auricular cartilage repair using cryogel scaffolds loaded with BMP-7-expressing primary chondrocytes.

The loss of cartilage tissue due to trauma, tumour surgery or congenital defects, such as microtia and anotia, is one of the major concerns in head and neck surgery. Recently tissue-engineering approaches, including gene delivery, have been proposed for the regeneration of cartilage tissue. In this study, primary chondrocytes were genetically modified with plasmid-encoding bone morphogenetic protein-7 (BMP-7) via the commercially available non-viral Turbofect vector, with the aim of bringing ex vivo transfected chondrocytes to resynthesize BMP-7 in vitro as they would in vivo. Genetically modified cells were implanted into gelatin-oxidized dextran scaffolds and cartilage tissue formation was investigated in 15 × 15 mm auricular cartilage defects in vivo in 48 New Zealand (NZ) white rabbits for 4 months. The results were evaluated via histology and early gene expression. Early gene expression results indicated a strong effect of exogenous BMP-7 on matrix synthesis and chondrocyte growth. In addition, histological analysis results exhibited significantly better cartilage healing with BMP-7-modified (transfected) cells than in the non-modified (non-transfected) group and as well as the control.

3D ingrowth of bovine articular chondrocytes in biodegradable cryogel scaffolds for cartilage tissue engineering.

A feasibility study was undertaken to examine the potential of biodegradable HEMA-lactate-dextran (HEMA-LLA-D)-based cryogels as scaffolds for cartilage tissue engineering. This was a preliminary in vitro study giving essential information on the biocompatibility of cryogels with cartilage cells. HEMA-lactate (HEMA-LLA) and HEMA-LLA-D were synthesized and characterized by different techniques. Cryogel scaffolds with supermacroporous structures were produced by cryogenic treatment of these macromers. Chondrocytes obtained from bovine articular cartilage were seeded onto cylindrical cryogels and cultured. The samples were examined by several microcopical techniques for cell viability and morphological analyses were performed at two culture points. Histological study of the constructs revealed the cells' growth on the surface and within the scaffolds. Confocal microscopical images demonstrated that the majority of live vs. dead cells had been attached to and integrated with the pores of the scaffold. SEM analysis showed round to oval-shaped chondrocytic cells interconnected with each other by communicating junctions. The chondrocytes rapidly proliferated in the cryogels, manifesting that they fully covered the scaffold surface after 9 days and almost filled the spaces in the pores of the scaffold after 15 days of culture. Chondrocytes secreted significant amount of extracellular matrix in the scaffolds and exhibited highly interconnective morphology. Light and transmission electron microscopy revealed groups of active cartilage cells closely apposed to the cryogel. We concluded that cryogel scaffolds could be excellent candidates for cartilage tissue regeneration with their extraordinary properties, including soft, elastic nature, highly open interconnected pore structure and very rapid, controllable swellability.

A new concept in the treatment of extravasation injury: controlled drug delivery systems.

PURPOSE: To investigate the effectiveness of the intralesionally injected controlled granulocyte-monocyte colony stimulating factor (GM-CSF) releasing system in widening refractory extravasation wounds. METHODS: The determination of in vitro GM-CSF release from chitosan gel was the first, and in vivo effect of the molecule was the second step of the study. Thirty-five Wistar-Albino rats were randomly divided into 5 groups: 1) control group (adriamycin group) (n=7); 2) adriamycin+normal saline group (n=7); 3) adriamycin+chitosan group (n=7); 4) adriamycin+1 µg/mL GM-CSF-loaded chitosan group (n=7); and 5) adriamycin+10 µg/mL GM-CSF loaded chitosan group (n=7). The wound area was measured macroscopically and histological examination was carried out for wound healing and tissue response to the polymer. RESULTS: The best healing process was observed with the controlled released GM-CSF groups (groups 4 and 5). The 1 µg/mL GM-CSF loaded group showed superior wound healing than that of 10 µg/mL GM-CSF loaded gels. This result was correlated with the in vitro study which also showed increased drug release in the 1 µg/mL GM-CSF loaded group than the 10 µg/mL GMC-SF loaded gels. CONCLUSION: This study suggests that GM-CSF, applied with controlled drug delivery system, can supply dynamic treatment options with long-lasting activity in single-dose administration.

Chitosan and alginate scaffolds for bone tissue regeneration.

Polymeric scaffold for tissue regeneration was developed for veterinary applications. Oxytetracycline hydrochloride (OTC), which is a widely used antibiotic in veterinary medicine was chosen as the model compound. Gel formulations using chitosan and alginate were prepared in distilled water or in 1% (v/v) acetic acid solution. Sponges were also prepared by a freeze-drying process. Tripolyphosphate was used for cross-linking. Viscosity was decreased in the presence of OTC in chitosan gels whereas no difference was found with alginate gels. All gels showed pseudoplastic behaviour. Water absorption capacity was highest with chitosan/alginate sponges. The solvent used for preparation of the chitosan gels was found to affect the release of OTC. The release of OTC from the sponges was increased by cross-linking. Chitosan/alginate sponges showed the slowest and lowest drug release among the developed sponge formulations in this study. The formulations were found to be biocompatible, inducing no adverse reaction in vivo on surgically formed bone defects of radius of rabbits. The level of organization of the remodelled new bone in the treatment groups was better than that of control. Incorporation of OTC into formulations did not show any considerable enhancing effect.

In vivo performance of antibiotic embedded electrospun PCL membranes for prevention of abdominal adhesions.

The aim of this study was to prepare nonwoven materials from poly(epsilon-caprolactone) (PCL) and their antibiotic containing forms by electrospinning, so as to prevent postsurgery induced abdominal adhesions in rats. epsilon-Caprolactone was first polymerized by ring-opening polymerization, and then it was processed into matrices composed of nanofibers by electrospinning. A model antibiotic (Biteral) was embedded within a group of PCL membranes. In the rat model, defects on the abdominal walls in the peritoneum were made to induce adhesion. The plain or antibiotic embedded PCL membranes were implanted on the right side of the abdominal wall. No membrane implantation was made on the left side of the abdominal wall that served as control. Macroscopical and histological evaluations showed that using these barriers reduces the extent, type, and tenacity of adhesion. The antibiotic embedded membranes significantly eliminated postsurgery abdominal adhesions, and also improved healing.

Bone generation on PHBV matrices: an in vitro study.

Bone formation was investigated in vitro by culturing rat marrow stromal osteoblasts in biodegradable, macroporous poly(3-hydroxybutyric acid-co-3-hydroxyvaleric acid) (PHBV) matrices over a period of 60 days. Foams were prepared after solvent evaporation and solute leaching. PHBV solutions with different concentrations were prepared in chloroform: dichloromethane (1:2, v/v). In order to create a matrix with high porosity and uniform pore sizes, sieved sucrose crystals (300-500 microm) were used. PHBV foams were treated with rf-oxygen plasma (100 W 10 min) to modify their surface chemistry and hydrophilicity with the aim of increasing the reattachment of osteoblasts. Osteoblasts were isolated from rat bone marrow and seeded onto PHBV foams. The cell density on and in the foams was determined with MTS assay. MTS results showed that osteoblasts proliferated on PHBV. Twenty-one days after seeding of incubation, growth of osteoblasts on matrices and initiation of mineralization were observed by confocal laser scanning microscopy. Increasing ALP and osteocalcin secretion during 60 days confirmed the osteoblastic phenotype of the derived stromal cells. SEM, histological evaluations and confocal laser scanning microscopy showed that osteoblasts could grow inside the matrices and lead to mineralization. Cells exhibited spindle-like morphology and had a diameter of 10-30 microm. Based on these, it could confidently be stated that PHBV seems to be a promising polymeric matrix material for bone tissue engineering.

EGF containing gelatin-based wound dressings.

In case of bulk loss of tissue or non-healing wounds such as burns, trauma, diabetic, decubitus and venous stasis ulcers, a proper wound dressing is needed to cover the wound area, protect the damaged tissue, and if possible to activate the cell proliferation and stimulate the healing process. In this study, synthesis of a novel polymeric bilayer wound dressing containing epidermal growth-factor (EGF) -loaded microspheres was aimed. For this purpose, a natural, nontoxic and biocompatible material, gelatin, was chosen as the underlying layer and various porous matrices in sponge form were prepared from gelatin by freeze-drying technique. As the external layer, elastomeric polyurethane membranes were used. Two different doses of EGF was added into the prepared gelatin sponges (1 and 15 microg/cm2) to activate cell proliferation. EGF addition was carried out either in free form or within microspheres to achieve prolonged release of EGF for higher efficiency. The prepared systems were tested in in vivo experiments on full-thickness skin defects created on rabbits. At certain intervals, wound areas were measured and tissues from wound areas were biopsied and processed for histological examinations. The wound areas decreased upon low-dose EGF application but the difference between the affects of free EGF and microsphere loaded EGF was not so distinct. Upon increasing the dose of EGF by a factor of 15, it was observed that controlled release of EGF from microspheres provided a higher degree of reduction in the wound areas. Histological investigations showed that the prepared dressings were biocompatible and did not cause any mononuclear cell infiltration or foreign body reaction. The structure of the newly formed dermis was almost the same as that of the normal skin.

In vivo response to biodegradable controlled antibiotic release systems.

In this study, the major goal was to evaluate in vitro and in vivo findings by macroscopy, radiology, and histology to determine the effectiveness of therapy of experimental implant-related osteomyelitis with antibiotic carrier rods constructed of microbial polyesters. The polymers used were poly(3-hydroxybutyrate-co-4-hydroxyvalerate) [P(3-HB-co-4-HB)] and poly(3-hydroxybutyrate-co-3-hydroxy- valerate) [P(3-HB-co-3-HV)]. Both the Sulperazone and the Duocid-P(3-HB-co-4-HB) rods with a drug to polymer ratio of 1:1 (w/w) were effective in treating the bone infection that was experimentally initiated by inoculation of a hemolytic strain of Staphylococcus aureus (coagulase positive; phage type 52/52b) together with metal implants into the medullary area of rabbit tibia. Macroscopical data revealed that the effectiveness of therapy was apparent at week 6 for all categories tested. Radiological findings with Duocid- and Sulperazone-loaded P(3-HB-co-4-HB) rods improved significantly when judged by changes in periosteal elevation, widening of bone shaft, new bone formation, and soft-tissue deformation after 6 weeks of implantation. Histologically the signs of infection were found to subside by weeks 3 and 6. Inflammatory cells were replaced with bone-forming cells upon treatment with Sulperazone-P(3-HB-co-4-HB) and Duocid-P(3-HB-co-4-HB). Osteoblastic activity was prominent. Intramedullary inflammation, although still present, started to be replaced by fibrous or bony tissue. Histological findings presented the subsidence of infection. In summary, the antibiotic-loaded biopolymeric rods appeared to have potential as a new controlled-release system for the treatment of implant related osteomyelitis and chronic osteomyelitis.

Assessment of mineral density and atomic content of fracture callus by quantitative computerized tomography.

The mineral density and atomic numbers of elements in the periosteal callus and the cortex area of a healing fracture were measured by quantitative computerized tomography (QCT) to obtain accurate information on the mineralization process in rabbit tibia. The mineral density of the periosteal callus was highest on day 15 and decreased gradually throughout the experiment. This was initially detected by QCT, but not with conventional radiography. An apparent decrease in cortical bone density on days 28 and 42 after fracture was observed. Atomic numbers of elements in the cortex remained stable, indicating a possible homeostatic mechanism of mineral preservation at the fracture callus and involved cortical area. QCT may predict density alterations in the fracture callus more accurately than conventional X-ray. Further studies are essential to predict a relationship between mineral density, atomic numbers, and mechanical stability.

Sulbactam-cefoperazone polyhydroxybutyrate-co-hydroxyvalerate (PHBV) local antibiotic delivery system: in vivo effectiveness and biocompatibility in the treatment of implant-related experimental osteomyelitis.

In this study, a novel antibiotic carrier system for use in the treatment of implant-related and chronic osteomyelitis was developed. Sulbactam-cefoperazone was introduced to rods of polyhydroxybutyrate-co-hydroxyvalerate (22 mol % HV, w/w), a member of a family of microbial-origin polymer that is biodegradable, biocompatible, and osteoconductive due to its piezoelectric property. The antibiotic-loaded carrier was implanted into the infection site that was induced by Staphylococcus aureus inoculation into the rabbit tibia. The effectiveness of this was assessed macroscopically, radiographically, bacteriologically, and histopathologically. Findings of infection subsided on day 15 and almost complete remission was observed on day 30. The control side that contained antibiotic-free rods, however, worsened. These findings prompted us to conclude that the novel biodegradable antibiotic carrier developed in the present study seems to be a promising candidate for use in the treatment of severe bone infection.

Development of a calcium phosphate-gelatin composite as a bone substitute and its use in drug release.

This study was carried out to develop a calcium phosphate-gelatin composite implant that would mimic the structure and function of bone for use in filling voids or gaps and to release bioactive compounds like drugs, growth hormones into the implant site to assist healing. XDS analysis of the synthesized calcium phosphate revealed a calcium to phosphorus molar ratio of ca. 2.30, implying a less erodible material than hydroxyapatite (1.67). Release of the antibiotic gentamicin from the implant was with a burst, whether in situ or in vivo, followed by an almost constant release for about three months. It was found that the release rate could be decreased by increasing the density of the gelatin membrane. Upon implantation into rabbit tibia the release duration was substantially shortened (to about 4 weeks) with respect to the in situ tests basically due to the degradation of gelatin. In vivo studies with rabbits confirmed this degradation. The composite was perfectly biocompatible as shown by the histological studies. It, thus, has a great potential as a bone substitute material.

A study of adenosine treatment in experimental acute spinal cord injury. Effect on arachidonic acid metabolites.

STUDY DESIGN: A prospective, randomized, blinded experimental trauma study. STUDY OBJECTIVE: The effect of adenosine on arachidonic acid metabolites and lipid peroxidation was investigated in induced spinal cord injury. SUMMARY OF BACKGROUND DATA: Effects of adenosine in ischemia-reperfusion models have been studied, but no studies of adenosine's effect on direct trauma to the spinal cord have been reported. METHODS: Thirty-seven adult Wistar albino rats were randomly divided into four groups and underwent laminectomy. Group 1 underwent a sham operation. Group 2 received an intravenous adenosine infusion of 100 micrograms/kg per minute for 30 minutes. In Group 3, a standard spinal cord trauma of 50 g.cm strength was established at the lower thoracic level with a "weight-drop" technique, and Group 4 received an infusion of adenosine (100 micrograms/kg per minute) for 30 minutes after the trauma. RESULTS: Tissue prostaglandin E2 activity was significantly higher in adenosine-treated trauma groups when compared with that in other groups (P < 0.0001). The difference in tissue leukotriene C4 activity between control and trauma groups was significant (P < 0.05). Adenosine infusion after trauma limited the increases in lipid peroxidation, with the difference approaching significance at P = 0.06. The structure of myelin was well preserved in the adenosine-treated trauma group. However, the changes were irreversible in severely damaged areas. CONCLUSION: After acute spinal cord trauma, intravenous adenosine infusion of 100 micrograms/kg per minute could attenuate progression to secondary injury, but adenosine alone was not effective yet.

Ultrastructural findings of bone marrow in a case with malignant osteopetrosis following successful allogeneic bone marrow transplantation.

A nine-month-old female patient suffering from malignant osteopetrosis was evaluated by light and transmission electron microscopic study before and following allogeneic bone marrow transplantation (BMT). Bone marrow specimens were obtained from iliac crest biopsies. Before BMT, the bone marrow had an irregular appearance and was filled with bridging bony trabeculae devoid of cells. Following BMT, the marrow had an almost normal appearance with no myelofibrosis and a relatively regular distribution of hematopoletic cells. The osteocytes were visible in their lacunae in the bone matrix. Presence of bone resorbing and bone forming cell together demonstrated that the bone was beginning to gain its normal dynamic structure. These findings were in accordance with the clinical, laboratory and radiological data which showed the beneficial effect of the therapy.

Effects of octreotide and a-tocopherol on bacterial translocation in experimental intestinal obstruction: a microbiological, light and electronmicroscopical study.

BACKGROUND/AIMS: Bacterial translocation induced by intestinal obstruction is suggested to be due to increased intestinal luminal volume, leading to intestinal overgrowth with certain enteric microorganisms and intestinal mucosal damage. If this suggestion is true, maintenance of intestinal mucosal integrity by a cytoprotective agent, a-tocopherol, and inhibition of gastrointestinal secretions by octreotide should decrease the incidence of bacterial translocation and extent of mucosal injury due to intestinal obstruction. METHODS: Complete intestinal obstruction was created in the distal ileum of male Wistar Albino rats by a single 3-0 silk suture. The animals received subcutaneous injections of 1 ml of physiologic saline (group 1) (PS 24) and 1 ml of saline containing octreotide acetate (100 micrograms/kg) (group 2) (OC 24), at 0, 12 and 24 hours of obstruction. In group 3 (PS 48) and group 4 (OC 48), the rats were treated with subcutaneous physiologic saline (1 ml) and octreotide acetate (100 micrograms/kg), respectively, beginning at the time of obstruction and every 12 hours for 48 hours. The rats in group 5 (Toc 24), were pretreated with intramuscular a-tocopherol 500 mg/kg on day 1 and 8, and underwent laparotomy on day 9. A third dose of a-tocopherol was injected at the time of obstruction on day 9 and no treatment was given thereafter. We tested the incidence of bacterial translocation in systemic organs and circulation and evaluated the histopathological changes in all groups. RESULTS: Treatment with octreotide acetate was found to be ineffective in reducing the incidence of translocation, with no histopathological improvement. Mucosal damage scores, on the other hand, in the a-tocopherol group were statistically less than those in the octreotide and control groups (p < 0.05). Additionally, a-tocopherol treatment decreased the incidence of organ invasion with translocating bacteria, although this difference did not reach statistical significance. CONCLUSION: Octreotide acetate treatment in complete intestinal obstruction has no effect on the incidence of bacterial translocation. a-Tocopherol, on the other hand, has a cytoprotective effect on intestinal mucosa in intestinal obstruction which, in turn, is thought to decrease bacterial translocation when used in physiological doses and prophylactically.

Tween 80 and endothelium: functional reduction due to tissue damage.

The effects of Tween 80, a surface active agent which is frequently used for dispersion of water-insoluble substances, on endothelial function in rabbit aorta, were investigated in a cascade superfusion bioassay system by using a deendothelized precontracted rat aorta ring as a bioassay tissue. Prior incubation of rabbit thoracic aorta segments with higher concentrations of Tween 80, caused total inhibition of acetylcholine-induced endothelium derived relaxing factor (EDRF) release, while moderate concentrations had no significant effect. Perfusion of the donor aortae segments with 10 (-1) - 10 (-3) ml/1 Tween 80 also inhibited the EDRF release from donor aorta dose-dependently. Histopathological examination of the vasculary wall and endothelial structures revealed a significant desquamation of vasculary endothelium with the highest concentration of Tween 80 used in this study. These data suggest that the mechanism underlying the reduction in the release of EDRF from donor aorta segments may be the destruction by Tween 80 of endothelial lining.

Endothelin-1-induced oedema in rat and guinea-pig isolated perfused lungs.

Endothelin-1 caused an increase in perfusion pressure, bronchial resistance, lung weight and tracheal effusion when infused through the pulmonary artery of rat and guinea-pig isolated lungs. In contrast to vasoconstriction, the effects of endothelin-1 on bronchial resistance, lung weight and tracheal effusion were not concentration-dependent. Recovery from vasoconstriction occurred within 15-30 min when the lung was further perfused with Krebs buffer. Increases in lung weight, bronchial resistance and tracheal effusion induced by endothelin-1 were irreversible when infused at concentrations above 10(-10) M. UK 38,485, a thromboxane A2 synthesis inhibitor, partly prevented the increase in lung weight and tracheal effusion without altering the vasoconstriction induced by endothelin-1. Such an antagonism was not seen in guinea-pig lung at the concentration used. Iloprost, a stable analogue of prostacyclin, antagonized the effects of endothelin-1 on perfusion pressure and lung weight without reducing tracheal effusion in both rat and guinea-pig lungs. Pretreatment with allopurinol did not alter the effects of endothelin-1. These results were taken as evidence for the potent lung oedema-producing effect of the peptide which seems to be partially mediated by the secondary release of thromboxane A2.