Macrophage interleukin 1 response to injected silicone in a rat model.

Observations of silicone granuloma formation and migration of silicone to regional lymph nodes have indicated a need for more research into the possible immunological responses to silicone. The present study was undertaken to assess the effect of injected silicone particles on the ability of splenic macrophages to produce interleukin 1 (IL-1) and to determine the relative quantities produced. Lewis rats were divided into 4 groups: Group 1 animals (n = 3) were injected subcutaneously with sterile saline (2.5 ml) and served as control animals; Group 2 animals (n = 3) also served as control subjects, but macrophages isolated from these animals were exposed to lipopolysaccharide (LPS); Group 3 animals (n = 3) were injected subcutaneously with Freund's complete adjuvant (FCA) (2.5 ml) to serve as FCA control animals; and Group 4 animals (n = 3) received a subcutaneous injection of a sonicated slurry of equal parts FCA and silicone (2.5 ml each). IL-1 production was not significantly increased in splenic macrophages from animals exposed to the silicone slurry (p greater than 0.20) 8 months after injection as compared with control animals or animals given FCA alone. Macrophages exposed to LPS, a known mitogen, had significantly elevated IL-1 production. Subcutaneously injected silicone particles did not elicit an increase in IL-1 production in rat macrophages.     

Local tissue reaction to the subureteral injection of glutaraldehyde cross-linked bovine collagen in humans

Although the technique of subureteral injection has been widely accepted as an alternative to reimplantation in the treatment of vesicoureteral reflux, the choice of the material to be used is controversial. We have used glutaraldehyde cross-linked bovine collagen to correct vesicoureteral reflux within the context of a Food and Drug Administration approved investigational study. We report the local tissue reaction to the implanted collagen in 7 patients who underwent reimplantation 3 to 19 months after failed endoscopic therapy. Glutaraldehyde cross-linked bovine collagen engendered a minimal localized inflammatory reaction without causing granuloma formation. Subsequent reimplantation was not hindered by the presence of the implant materials.     

Histological evaluation of Permacol as a subcutaneous implant over a 20-week period in the rat model.

This study assessed the suitability of Permacol (a porcine derived, isocyanate cross linked collagen based biomaterial) as an alternative to autologous tissue in soft tissue reconstruction. The Sprague-Dawley rat was used as a model for subcutaneous implantation over a 20 week period and comparison made with two other porcine biomaterials (small intestinal submucosa and glycerol treated-ethylene oxide sterilised porcine dermis). Implants were scored histometrically on the degree of acute inflammation, chronic inflammation, fibrosis and stromal response. The vascularity and percentage composition of collagen within Permacol were assessed by stereology and seescan image analysis, respectively. In general terms, Permacol was well tolerated as a subcutaneous implant, with only a minor chronic inflammatory response remaining after a 20 week period of implantation. There was evidence of collagen degradation during this period and vascular ingrowth into Permacol was limited. Permacol has the potential for a broad range of applications in plastic surgery, but may benefit from modification to promote a more rapid degree of vascularisation.     

The osseous response to corundum blasted implant surfaces in a canine hip model.

The purpose of this study was to examine the radiographic and histologic response to corundum blasted implant surfaces of varying roughness in a canine total hip arthroplasty model. Three types of tapered femoral implants were made from titanium alloy and were identical in every respect except surface finish. The entire surface of the femoral implant had a 2.9-, 4.2-, or 6.7-micron average surface roughness (Ra) from blasting with 60-, 24-, or 16-grit corundum particles, respectively. Twenty-two stems in 11 dogs were evaluated at 6 months. Twenty-one of the stems showed osseointegration, whereas in one stem a fibrous interface developed. Abundant new periimplant bone formation occurred, particularly within the intramedullary canal where trabeculae spanned implant to endosteal cortex gaps as large as 5 mm. Bone apposition with the 60-, 24-, and 16-grit stems averaged 31.7%, 32%, and 27.9%, respectively; the differences were not statistically significant. However, the pattern of new bone formation was different in that the average length of each region of bone apposition for the 60- and 24-grit surfaces was 50% greater than that for the coarser 16-grit surface. The observations of this study indicate that because of their highly osteoconductive nature, corundum blasted surfaces represent an important and valuable technology for the design of noncemented implants.     

Skeletal response to dietary calcium in a rat model simulating weightlessness

Unweighting the hindlimbs of a rat by tail suspension leads to a decrease in bone in the unweighted hindlimbs, but not in the normally weighted forelimbs. We evaluated whether increments in dietary calcium could prevent this. Growing rats were fed diets ranging in calcium content from 0.1% to 2.4%. After the rats were suspended for two weeks, we found no differences between suspended and control animals fed the same diet with respect to calcium transport or serum levels of calcium, phosphorus, 1,25-dihydroxyvitamin D, and parathyroid hormone. In both groups, increasing dietary calcium reduced active intestinal calcium transport and serum 1,25-dihydroxyvitamin D levels. The calcium content of the tibia and lumbar vertebra (but not the humerus) was reduced in suspended rats compared to control rats fed the same diet. However, increasing dietary calcium increased the calcium content of all bones in both suspended and control animals. The bone formation rate at the tibiofibular junction (measured by double-label tetracycline) was reduced in the suspended animals compared to controls and was not altered by dietary calcium. However, the marrow area of the tibia, an indication of bone resorption, did not differ between suspended and control animals and was equally reduced in both groups when dietary calcium was increased. Our data suggest that the deleterious effects of skeletal unweighting on bone formation cannot be explained by changes in the calciotropic hormones and are not reversed by increments in dietary calcium. However, increasing dietary calcium can increase bone calcium, even in unweighted limbs, by decreasing bone resorption.     

Early inflammatory response of knee ligaments to prolotherapy in a rat model

Prolotherapy is an alternative injection‐based therapy for chronic musculoskeletal pain. Three different proliferants, D ‐glucose (dextrose), phenol‐glucose‐glycerine (P2G), and sodium morrhuate, used in prolotherapy are hypothesized to strengthen and reorganize chronically injured soft tissue and decrease pain through modulation of the inflammatory process. Our hypothesis is that commonly used prolotherapy solutions will induce inflammation (leukocyte and macrophage infiltration) in medial collateral ligaments (MCLs) compared to needlestick, saline injection, and no‐injection controls. MCLs of 84 Sprague‐ Dawley rats were injected one time at both the tibial and femoral insertions. Immunohistochemistry (IHC) was used to determine the inflammatory response at three locations (tibial and femoral insertions and midsubstance) 6, 24, and 72 h after dextrose injection compared to saline‐ and no‐injection controls and collagenase (positive control) (n = 4). qPCR was used to analyze gene expression 24 h postinjection (n = 4). Sodium morrhuate, P2G, and needlestick control were also investigated after 24 h (n = 4). In general, inflammation (CD43+, ED1+, and ED2+ cells) increased after prolotherapy injection compared to no‐injection control but did not increase consistently compared to saline and needlestick control injections. This response varied by both location and proliferant. Inflammation was observed at 6 and 24 h postinjection but was resolved by 72 h compared to no‐injection controls (p < 0.05). CD43+ leukocytes and ED2+ macrophages increased compared to needlestick and saline‐injection control, respectively, 24 h postinjection (p < 0.05). Prolotherapy injections created an inflammatory response, but this response was variable and overall, not uniformly different from that caused by saline injections or needlestick procedures. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:816–823, 2008     

Tensile strength and host response towards different polypropylene implant materials used for augmentation of fascial repair in a rat model

We compared inflammatory response, fibrosis and biomechanical properties of different polypropylene materials from one manufacturer (Tyco Healthcare) in a rat model for primary fascial repair. Full-thickness abdominal wall defects were primarily repaired using ‘overlay’ technique. Multifilament implants were Surgipro SPM and SPMW, the latter a wider-weave type of the former. Monofilament SPMM implants and polypropylene suture repair (Surgipro II) served as controls. Explants were evaluated macroscopically and changes in thickness, shrinkage and tensile strength were measured. Inflammatory and connective tissue response was assessed on haematoxylin-eosin and Movat stains. Immunohistochemistry was done to localise rat macrophages/monocytes. Multifilament materials induced a shorter acute inflammatory response and more pronounced chronic inflammatory reaction compared to monofilament implants. Macrophages could be found deep in interstices 7.5 by 12.5 μm. No difference in collagen deposition and neovascularisation was observed. At 90 days time point, explants reconstructed with tighter woven multifilament SPM were weaker than sutured or SPMM controls. Overall shrinkage of 10% was comparable for all groups.     

Histological analysis of achilles tendons in an overuse rat model

The purpose of this study was to design an animal model that induces histological changes in Achilles tendons consistent with those cited in the literature for human Achilles tendon disease. Sprague‐Dawley rats were subjected to 10° uphill treadmill running on a custom‐designed rodent treadmill and at a speed of 17 meters per minute for 1 h, five times per week, over a 12‐week treatment period. Subsequent histological analysis revealed alterations in the rat Achilles tendon that were generally consistent with those described in the literature for diseased human tendon tissues. These features include: decreased collagen fiber organization, more intense collagen staining, and increased cell nuclei numbers. Interestingly, though, immunohistochemical cell typing suggests that the observed increased cellularity does not include a significant inflammatory component but is secondary to increased numbers of endothelial cells (i.e., vascularization) and fibroblasts. These histological features likely represent a biological repair/remodeling response resulting from overuse running. © 2008 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 26:840–846, 2008     

Influence of mesh materials on collagen deposition in a rat model.

Alterations of the extracellular matrix (ECM) with its major component collagen are increasingly discussed as possible risk factors implicated in the development of abdominal-wall herniation. Because of the widespread use of alloplastic meshes for the surgical repair of hernias, an animal study was performed to analyze the influence of various mesh materials on the quantity and quality of collagen deposition. In 60 male Sprague-Dawley rats an abdominal replacement was performed using three different kinds of mesh materials: polyester (PE), a pure polypropylene (PP), and a composite mesh made of polypropylene and polyglactin (PG). A simple fascia suture repair served as control. The count of fibroblasts, the collagen/protein ratio, the type I/III collagen ratio, and the expression of basic fibroblast growth factor (b-FGF) at the interface were analyzed after 7, 21, and 90 days. The ratio of collagen to overall protein (microg/mg) showed significant differences comparing different mesh materials (sham controls 38.44 +/- 16.33 microg/mg, PE 68.5 +/- 23.8 microg/mg, PP 101.6 +/- 32.3 microg/mg, PG 49.6 +/- 11.6 microg/mg at day 90). The ratio of collagen type I/III increased over time in all groups. However, 90 days after mesh implantation the ratio was always significantly lowered compared to the controls. No significant difference was found comparing different mesh materials. The alteration of the scar composition is closely connected to an increased b-FGF expression. b-FGF and count of fibroblasts highly correlated (r =.95) and showed significant elevated levels compared to simple suture repair. The results of our study strongly support the notion that wound healing is affected by mesh implantation. The quality of the ECM deposition as determined by collagen type I/III ratio is impaired in general, whereas the quantity of ECM deposition is markedly influenced by the kind of mesh material.     

Evaluation of a cross-linked versus non-cross-linked collagen matrix in full-thickness skin defects

Autologous skin transplantation is the gold standard for treatment of full-thickness skin defects such as deep burn injuries, but has the disadvantages of limited donor sites and donor site morbidities. Alternative skin replacement products, such as xenografts and allografts, are not a permanent solution. Numerous manufactured skin substitutes already show promising approaches, but have limited efficacy. Therefore, wound dressings adaptable to the physiology of wound healing are still needed.In a randomized controlled in vivo study, a newly designed biocompatible collagen nonwoven matrix was compared to the Integra® bilayer dermal substitute and untreated controls in 48 full-thickness skin defects in a swine model.The take of all templates was complete, and all the tissue-engineered products accelerated dermal wound healing compared to the untreated controls, as identified by planimetric measurements. The higher collagen dose treatments and Integra®-covered wounds developed the thickest, cell-rich neoepidermal tissue in histological examination.The innovative biocompatible collagen matrix is flexibly applicable and modifiable, and offers potential as a carrier membrane for therapeutic supplemental products such as growth factors to further develop effective wound dressings.     

An in vivo evaluation of bone response to three implant surfaces using a rabbit intramedullary rod model

Our study was designed to evaluate osseointegration among implants with three surface treatments: plasma-sprayed titanium (P), plasma-sprayed titanium with hydroxyapatite (PHA), and chemical-textured titanium with hydroxyapatite (CHA). Average surface roughness (Ra) was 27 microns for the P group, 17 microns for the PHA group, and 26 microns for the CHA group. Bilateral distal intramedullary implants were placed in the femora of thirty rabbits. Histomorphometry of scanning electron microscopy images was used to analyze the amount of bone around the implants at 6 and 12 weeks after implantation. Greater amounts of osseointegration were observed in the hydroxyapatite-coated groups than in the noncoated group. For all implant surfaces, osseointegration was greater at the diaphyseal level compared to the metaphyseal level. No significant differences were seen in osseointegration between the 6 and 12 week time points. Although the average surface roughness of the P and the CHA groups was similar, osseointegration of the CHA implants was significantly greater. The results of this in vivo lapine study suggest that the presence of an hydroxyapatite coating enhances osseointegration despite similarities in average surface roughness.     

Elucidating the vascular response to burns with a new rat model.

Burn injury causes acute thrombosis and occlusion of vessels in the dermis directly killed by thermal energy. A vascular response also occurs in the uninjured dermis bordering the site of injury. Diminished blood flow leads to progressive ischemia and necrosis in the dermis beneath and surrounding the burn. If blood flow is maintained or restored in this area, the tissue survives. A noninvasive technique for studying dynamic changes in blood flow in this transitional dermis in rats is presented. A rectangular brass bar 19 mm wide with 5-mm transverse notches was heated in boiling water and applied to the skin surface for 20 seconds, making a "comb" burn composed of a row of four rectangular 10 x 19-mm full-thickness burns. Between the burns were 5 x 19-mm bands of uninjured skin, called "interspaces." After burning, blood flow near the surface of both the burn sites and the interspaces was monitored with a laser Doppler perfusion monitor for 24 hours. The vascular patency of blood vessels was directly visualized by latex vascular casts made 24 hours after burn. The possible prevention of progressive ischemia by injecting systemic ibuprofen was examined in this new model. Normal skin has a surface blood flow reading of 80 +/- 16 mV, burn sites have a reading of 11 +/- 4 mV, and interspaces have a reading of 21 +/- 4 mV at 24 hours postburn in untreated rats. Systemic ibuprofen given IM immediately postburn at 12.5 mg/kg increased blood flow to 80 +/- 28 mV within the interspaces, to 17 +/- 12 mV in the burn site, and to 80 +/- 9 mV in normal skin. The vascular casts showed an absence of patent vessels within both the burn sites and interspaces in untreated rats.(ABSTRACT TRUNCATED AT 250 WORDS)     

Early changes in serum osteocalcin and body weight are predictive of implant fixation in a rat model of implant loosening

Biomarkers are of interest to identify patients at risk for peri-implant osteolysis and aseptic loosening. We used a rat model of particle-induced peri-implant osteolysis to investigate if early changes in biomarkers were associated with subsequent implant fixation strength. Implants were placed in rat femora, which were then challenged with intra-articular knee injections of either clean polyethylene, lipopolysaccharide-doped polyethylene, or cobalt-chromium alloy particles, with particle-free vehicle serving as control (n ≥ 8 per group). Rats were weighed weekly, blood was collected at weeks 0, 3, 5, and 6, and locomotor behavior was assessed 4 days before study conclusion. Rats were euthanized 6 weeks post surgery. Week 6 serum was analyzed for five bone remodeling markers, while longitudinal serum was assessed for osteocalcin. Bone-implant contact, peri-implant trabecular architecture, and implant fixation strength were measured. Rats challenged with cobalt-chromium particles had a significant reduction in implant fixation strength compared with the vehicle-control group (P = .034). This group also had elevated serum osteocalcin (P = .005), depressed weight gain (P = .001) and less frequent rearing behavior (P = .029). Regardless of group, change in serum osteocalcin at week 3 (r = −.368; P = .046), change in weight at week 2 (r = .586; P < .001), as well as weight change at all other time intervals were associated with fixation strength. The finding that early alterations in serum osteocalcin and body weight were predictive of subsequent implant fixation strength supports continued investigation of biomarkers for early detection of peri-implant osteolysis and implant loosening. Further, change in biomarker levels was found to be more indicative of implant fixation status than any single measurement.     

Reconstituted bovine skin collagen enhances healing of bone wounds in the rat calvaria

Summary The value of reconstituted fibrillar collagen (Zyderm Collagen Implant I, a concentrated solution of pepsin-solubilized, bovin skin collagen) as a bone graft material was tested in 4 mm diameter surgically created defects of rat calvaria. All wounds were allowed to heal for 4 weeks, and were assessed both qualitatively and by computer-assisted morphometry. The fibrillar collagen was found to produce significantly more new bone than no graft or than heat-denatured fibrillar collagen. The fibrillar collagen was generally well tolerated, appeared to act as a hospitable osteoconductor, and became incorporated into the newly formed bone. The effect of collagen concentration was also tested by comparing the fibrillar collagen at 3.5% (Zyderm Collagen Implant I) with 6.5% suspension of collagen (Zyderm Collagen Implant II). There were no significant differences observed, but a definite trend was evident for Zyderm II to encourage more bone formation than Zyderm I. It is concluded that reconstituted fibrillar collagen is a hospitable, osteoconductive substance that enhances bone healing of calvarial defects in the rat.     

The immune response to carcinoma of the colon in a rat model: a temporal study

Carcinoma of the colon was induced in rats by treatment with 1, 2-dimethylhydrazine (DMH). Macroscopic tumours were observed from eight weeks after commencing DMH injections and rapidly increased in mass to cause death of the animals between 20 and 27 weeks later. Tumour immunity was assessed by the gross parameters of body, spleen and mesenteric lymph node weight and by in vitro tests of lymphocyte antitumour cytotoxicity and leucocyte migration inhibition by tumour and normal colon homogenates. Immune responses were recorded in control (saline treated) and in tumour (DMH treated) rats. Body weight of the tumour rats paralleled that of control animals until tumours were well advanced. At this time tumour animal body weight was significantly less than that of control animals. Spleen weight and in vitro lymphocyte antitumour cytotoxicity were significantly greater in tumour than in control rats and were so found soon after macroscopic tumours were observed. Mesenteric lymph node weight and leucocyte migration inhibition did not change until late in tumour development, but at this time there was an immune response in tumour animals. The study showed that carcinoma of the colon induced in rats was associated with an antitumour immune response and that this change occurred soon after macroscopic tumours were present. In spite of this the animals died of their neoplasms.     

Comparison of bovine collagen xenografts to autografts in the rabbit

The use of bovine tendon as a xenograft material in humans is attractive because of its ready availability and favorable mechanical characteristics. Previous research has shown that the fibroblasts and some extracellular proteoglycans and glycoproteins, not the collagen matrix itself, in bovine tendon are primarily responsible for its antigenicity. Various attempts have been made to decrease the antigenicity of these grafts. A chloroform/methanol (CM) extraction procedure has been developed that selectively removes the fibroblasts from bovine tendon without destroying the collagen matrix. The mechanical, immunologic, and local host tissue responses to these grafts were compared to autografts and to untreated and glutaraldehyde-treated bovine tendon xenografts. The humoral immune response to a purified bovine Type I collagen product was also studied. The central two-thirds of a rabbit Achilles tendon were replaced with a reversed autograft or an experimental graft. Histologic examination of one- and two- week specimens showed an acute inflammatory response to all grafts. Untreated grafts stimulated a severe inflammatory response and were almost completely resorbed by two weeks. Glutaraldehyde-treated grafts were encapsulated. Cellular repopulation was minimal and inflammatory response was more persistent than in the autograft and CM groups. Inflammatory response to CM-treated grafts was similar to that of autografts. The CM grafts repopulated rapidly with host cells. The mechanical strength of CM grafts was equal to autograft controls at 12 weeks. The mechanical strength of untreated and glutaraldehyde-treated grafts was significantly lower. Measurement of the humoral immune response to these grafts was conducted in an independent group of animals using an enzyme-linked immunosorbent assay. A significant antibody response to untreated, glutaraldehyde-fixed, and CM-treated grafts was detected at 30 days. Antibody titers to glutaraldehyde-fixed and untreated grafts remained elevated at 60 and 90 days. In the CM group, antibody titers decreased to the level of autograft controls by 90 days. No significant antibody response was detected toward purified bovine Type I collagen.