Effect of Commercially Available Wound Irrigation Solutions on Uninfected Host Tissue in a Murine Model.

Background: Commercially available irrigation solutions are used to reduce bacterial contamination and prevent surgical site infections. However, the effect of these solutions on the healing capacity of tissue has not been well-established. The purpose of this study was to investigate the effects of 5 commercially available irrigation solutions on host tissue in a murine model. Methods: There were 5 treatment groups: bacitracin, Clorpactin, Irrisept, Prontosan, Bactisure, and normal saline control. The irrigation solutions were applied to the wound for 30 seconds or 1 minute, as per the manufacturer's instructions, and then washed with normal saline. Mice were sacrificed at 3 days and 10 days. The tissue was examined histologically for inflammation, edema, granulation tissue formation, and re-epithelialization. Granulation tissue formation and re-epithelialization were surrogates for effective wound healing. Results: All of the irrigation solutions had negative effects on host tissue in the acute phase. The inflammation and edema were improved in the later phase (10 days). Recovery and healing of the open wounds were observed for all groups at 10 days. The antiseptic irrigation solutions had similar cytotoxic effects on host tissue at 3 days and did not have delayed or compromised wound healing at 10 days when compared to normal saline control. Conclusions: Single short-duration use of these commercially available antiseptic irrigation solutions appears to be safe in an uninfected wound. Data from this study will provide surgeons with useful information regarding the safety of using antiseptic wound irrigation solutions intraoperatively for prevention of surgical site infections.

Common Wound Irrigation Solutions Produce Different Responses in Infected vs Sterile Host Tissue: Murine Air Pouch Infection Model.

Background: Despite desirable microbicidal actions of irrigation solutions in surgical site infection treatment, several studies demonstrate potential cytotoxic effects. This study investigated tissue damage caused by irrigation solutions in the presence or absence of infection. Methods: Air pouches were created in 60 mice and evenly divided into 2 groups as infected with Staphylococcus aureus and control. Groups were then subdivided both by type of solution and by timing after irrigation. Solutions included control (0.9% saline), bacitracin (33 IU/ml), 0.2% sodium oxychlorosene, 0.05% chlorhexidine gluconate, and 0.013% benzalkonium chloride. Results: Inflammation decreased in infected pouches compared to the sterile ones for all solutions except bacitracin on day 0 and for all on day 7. On day 0, infected pouches had increased necrosis with bacitracin (P = .006), chlorhexidine gluconate (P = .18), and benzalkonium chloride (P = .07); on day 7, there was decreased necrosis in infected pouches for all solutions (P < .05) except for sodium oxychlorosene (P = .18). Edema decreased in infected pouches on day 0 for all solutions. On day 7, infected pouches had decreased edema with 0.9% saline, bacitracin, and benzalkonium chloride (P < .05) and increased edema with chlorhexidine gluconate (P < .05) and sodium oxychlorosene (P = .069). Bacitracin allowed for more bacteria growth than sodium oxychlorosene (P = .024), chlorhexidine gluconate (P = .025), and benzalkonium chloride (P = .025). Conclusions: The presence of bacteria led to less immediate tissue inflammation and edema, while tissue necrosis varied over time. The current study may guide surgeons on which solution to use and whether to irrigate a possibly sterile wound or joint.

Attachment and Growth of Fibroblasts and Tenocytes Within a Porous Titanium Scaffold: A Bioreactor Approach.

Background: Direct attachment of tendons to metallic implants is important in orthopedics. Tissue integration depends on scaffold microstructure and composition. This study evaluated the effect of pore size of titanium on the viability and function of fibroblasts and tenocytes in a dynamic bioreactor. Methods: Standardized Ti porous cylinders with 3 pore sizes (400, 700, and 1000 µm) were seeded with fibroblasts or tenocytes (4500 cells/µL) in silicon tubes. Cells were analyzed via alamarBlue (AB) assay in addition to scanning electron microscopy at day 7 (fibroblasts) or day 8 (tenocytes) and day 15. AB functions as a cell health indicator where functional living cells reduce the resazurin dye (blue) in the solution to resorufin (pink), and cell viability can be quantified via spectroscopy. Results: At day 7, fibroblasts cultured on all sizes reduced AB, with significant differences noted between 400 vs 1000 µm (P = .013) and 700 vs 1000 µm (P = .001). At day 15, fibroblasts reduced AB on all sizes with a significant difference noted between 700 vs 1000 µm (P = .004). Fibroblasts on all 3 pore sizes increased AB reduction from day 7 to day 15. Tenocytes reduced AB with significant differences between the 400 vs 700 µm (P = .049) and the 400 vs 1000 µm pore sizes at day 8. In contrast, tenocyte reduction of AB decreased from day 8 to day 15. Scanning electron microscopy performed on fibroblast cylinders showed fibroblasts reached the surface of the cylinders, confirming interconnectivity. Conclusions: While both fibroblasts and tenocytes penetrated the pores, fibroblasts preferred larger size, whereas tenocytes favored smaller size. Results are encouraging since soft-tissue attachment to a metallic scaffold is difficult but clinically desirable. Future studies could be performed in an in vivo animal model.

Irrigation Solutions Negatively Affect the Viability and Function of Human Fibroblasts: An in vitro Study.

Introduction: Multiple irrigation solutions are used in orthopedic surgeries although there are limited studies on their lasting effects on human tissues. The purpose of this work was to investigate the cytotoxic effects of the irrigation solutions Bacitracin, Clorpactin (sodium oxychlorosene), Irrisept (0.05% chlorhexidine gluconate), and Bactisure (ethanol 1%, acetic acid 0.6%, sodium acetate 0.2%, benzalkonium chloride 0.013%, and water) on 3D cultures of human fibroblasts. Methods: Two independent experiments with 6 replicates were performed for the following conditions: Control (saline), bacitracin, Clorpactin, Irrisept, and Bactisure. Human fibroblast cell sheets were exposed to these solutions (1 or 2 min), followed by three washes with warm saline. Cell sheets were then cultured for additional 5- and 7-day posttreatment. Cell viability was measured using the alamarBlue (AB) assay. The more cytotoxic the irrigant, the lower the AB reduction. Results: For 1-min exposure time, significant differences in AB reduction were noted in Clorpactin, Irrisept, and Bactisure groups compared to control at both 5 days (Clorpactin p = 0.0003, Irrisept p = 7.31 × 10-15, Bactisure p = 6.86 × 10-14) and 7 days posttreatment (all groups p < 0.0001). The results were similar in the 2-min exposure groups. Bacitracin-treated fibroblasts displayed no significant difference at all measurement times compared to control. Discussion: Impacts of irrigation solution exposure on cell viability were varied. Irrisept and Bactisure showed the highest cell toxicity even after a brief exposure (1 min), while bacitracin and Clor-pactin exposure showed smaller impacts on cell viability as compared to saline controls. This in vitro study provided insight into the effects of the irrigants on human cells and provides the groundwork essential to move to in vivo studies. Our findings raised the concern that some irrigation solutions may have negative impacts on wound healing and healthy cellular response.

FiberWire vs FiberTape: Comparison of Bacterial Adherence in a Murine Air Pouch Wound Model.

BACKGROUND: For high-tensile strength sutures, past research has largely focused on mechanical properties or bacterial adherence across various manufacturers. PURPOSE: This study investigated high-tensile strength sutures with different shapes but otherwise identical composition. The purpose was to evaluate the differences between high-tensile strength suture wire and suture tape relative to bacterial adherence and bacterial retention after washout. STUDY DESIGN: Controlled laboratory study. METHODS: Sutures were implanted in dorsal air pouches of 72 BALB/cJ mice. Experimental pouches were inoculated with Staphylococcus aureus; no bacteria were used in the control conditions. The mice were randomized into 3 groups: group 1 underwent suture extraction 7 days after implantation; group 2 underwent an irrigation procedure, followed by immediate suture extraction on day 7; and group 3 underwent an irrigation procedure on day 7, with delayed suture extraction on day 14 after implantation. The sutures were evaluated using confocal microscopy; electron microscopy; and spectrophotometry, through which optical density, as measured by the amount of scattered light, is directly correlated with the number of bacteria. Histological assessment was performed on the pouches. RESULTS: Optical density (mean ± SD) was significantly higher for FiberTape sutures than for FiberWire sutures, respectively, at the 2-hour time point for all groups (group 1, 0.0550 ± 0.0081 vs 0.0162 ± 0.006 [P = .0054]; group 2, 0.0225 ± 0.0049 vs 0.0056 ± 0.0006 [P = .0045]; group 3, 0.055 ± 0.0222 vs 0.0043 ± 0.0005 [P = .0103]). Additionally, groups 2 and 3 showed statistically significant results at the 4-hour time points (group 2, 0.0384 ± 0.0087 vs 0.0145 ± 0.0042 [P = .0280]; group 3, 0.0532 ± 0.0159 vs 0.0101 ± 0.0025 [P = .0058]). The wash fluid also demonstrated significantly greater optical density for the FiberTape than the FiberWire sutures, respectively, at the 2-hour time point for all groups (group 1, 0.1657 ± 0.0319 vs 0.0317 ± 0.008 [P = .0063]; group 2, 0.0522 ± 0.0156 vs 0.0127 ± 0.0022 [P = .0219]; group 3, 0.1707 ± 0.0205 vs 0.0191 ± 0.0053 [P < .0001]). No bacterial growth occurred in the control conditions. Histological assessment revealed only mild inflammation in the control groups as compared with more severe responses in the experimental groups at all time points. CONCLUSION: FiberTape was associated with increased bacterial adhesion as well as retention as compared with FiberWire in an in vivo murine wound model. CLINICAL RELEVANCE: This study demonstrates that suture design influences the occurrence of and ability to clear surgical infection and must be considered when selecting high-tensile strength sutures in a clinical setting.

Response profiles of circulating leukocytes and metal ions in patients with a modular dual-mobility hip implant.

INTRODUCTION: The purpose of this study was to retrospectively evaluate blood metal ion levels and leukocyte profiles in patients with modular dual-mobility hip implant (MDM) during a postoperative follow-up up to 2 years. METHODS: We recruited 49 patients in a retrospective cohort study and had postoperative follow-up up to 2 years. Blood concentrations of chromium (Cr), cobalt (Co) and serum cytokines were measured. Flow cytometry was used to quantify the subpopulations of leukocytes, including CD14+ and CD16+ monocytes, CD3+ T lymphocytes, CD19+ B lymphocytes, CD4+ Helper T-cells and CD45+RA memory vs. naïve T-cells. RESULTS: Clinical performances of implants were good during 2 years of follow-up. Cr levels were normal in all patients and only detectable in 1 patient (1.4µg/L, ref < 5.0µg/L). Co levels were mildly elevated in 4 patients at 1 year (mean 1.375µg/L, range 1.2-1.7µg/L, ref < 1.0µg/L) and in 2 patients at 2-year follow-up (both 1.2µg/L). Interestingly, Co level observed in 3 patients at 1 year converted to undetectable at their 2-year follow-up. Percentages of B cells, T cells and their subpopulations were within normal levels. There was no increase of CD16+ inflammatory monocytes. DISCUSSION: With the recent introduction of MDM systems there is potential for metal ion release from the interface between the acetabular shell and CoCr liner. Clinical results have been good and metal levels undetectable or within acceptable ranges at 1-2 years. There was no evidence of activated immune response, as manifested by constant circulating leukocyte profiles and no increase of CD16+ inflammatory monocytes.

Reinforcement of a new calcium phosphate cement with RGD-chitosan-fiber.

Calcium phosphate cement (CPC) has been widely used in orthopedic and dental applications. A critical limitation of CPC is low strength and high susceptibility to severe fracture. Surgeons can use it only to reconstruct non-stress bearing bone, raising the need for a tougher new generation of CPC. Fibers have been used as a reinforcement of CPC to improve the strength of a pure CPC scaffold. The RGD peptides (Arg-Gly-Asp) have been used to improve the biocompatibility of the scaffold, via physical adsorption. The purpose of this study was to develop a novel CPC scaffold reinforced by RGD peptide-bearing chitosan fibers (RGD-fiber-CPC). Our data showed that the RGD-fiber-CPC scaffold had an increased flexural strength, and stimulated new bone formation in an animal model. The RGD-fiber-CPC is a novel bone graft substitute in orthopedic surgery.

Prognostic significance of VEGF expression in osteosarcoma: a meta-analysis.

Vascular endothelial growth factor (VEGF) is considered as a prime mediator of angiogenesis and has been implicated in carcinogenesis and metastasis. Various studies examined the relationship between VEGF overexpression with the clinical outcome in patients with osteosarcoma but yielded conflicting results. Electronic databases updated to April 2013 were searched to find relevant studies. A meta-analysis was conducted with eligible studies which quantitatively evaluated the relationship between VEGF overexpression and survival of patients with osteosarcoma. Survival data were aggregated and quantitatively analyzed. We performed a meta-analysis of eight studies that evaluated the correlation between VEGF overexpression and survival in patients with osteosarcoma. Combined hazard ratios suggested that VEGF overexpression had an unfavorable impact on overall survival (hazard ratio (HR) = 1.75, 95% confidence interval (CI): 1.21-2.28) in patients with osteosarcoma for overall populations, 2.37 (1.35-3.39) in Asian studies but not in non-Asian studies (HR = 1.51, 95% CI: 0.89-2.14). No significant heterogeneity was observed among all studies. VEGF overexpression indicates a poor prognosis for patients with osteosarcoma. However, the prognostic value of VEGF on survival in osteosarcoma patients still needs further large-scale prospective trials to be clarified.

[Measurement on bone density of children's mandible by quantitative computed tomography].

OBJECTIVE: To explore the change of bone density of children's mandible measured by quantitative computed tomography (QCT). METHODS: 71 children aged between 10-16 years were measured by QCT in left mandible angle, middle of mandible and right mandible angle. The differences were analyzed according to the age and sex respectively. RESULTS: The density of mandible of left mandible angle, middle of mandible, right mandible angle were 44.29, 89.70, 54.31 mg/dL in the 10-12 ages group, and 63.85, 122.47, 70.23 mg/dL in the 13-16 ages group. CONCLUSION: Male youngster's mandible density was increased with the age between 10-12 and 13-16 ages. There were significant differences between left mandible angle, middle of mandible and right mandible angle. There were significant differences of mandible density between male and female.

Inhibiting wear particles-induced osteolysis with doxycycline.

AIM: To study the effect of doxycycline (DOX) on osteoclastogenesis, mature osteoclast fate and function, wear particles-induced osteoeolysis, and to provide some foundation for treating aseptic loosening and osteolysis after joint arthroplasty. METHODS: Osteoclasts were generated from mouse bone marrow monocytes with the receptor activator of NF-kappaB ligand and the macrophage colony stimulating factor. DOX at a concentration of 5, 10, 15, and 20 microg/mL was respectively added to the medium. Seven days later, the osteoclasts were determined through tartrate-resistant acid phosphatase (TRAP) staining. Mature osteoclasts were isolated from newborn rabbits and cultured for 3 d in 24-well plates or on bone slices. DOX at a concentration of 5, 10, 15, and 20 microg/mL was respectively added to the medium. After TRAP staining, the osteoclasts were counted, resorption on bone slices was quantified, and the area was calculated after toluidine blue and Mayer-hematoxylin staining. Polymethyl methacrylate (PMMA) or ultra-high molecular weight polyethylene (UHMWPE) particles were implanted on the calvariae of C57BL/J6 mice. DOX, at a dose of 2 and 10 mg x kg(-1) x d(-1), was respectively given intraperitoneally for 7 d. Seven days later, the calvariae were removed and processed for pathological analysis. RESULTS: DOX treatment effectively inhibited in vitro osteoclastogenesis, affected the fate of mature osteoclasts, and inhibited mature osteoclasts, causing bone resorption. In vivo data indicated that DOX strongly inhibited PMMA or UHMWPE-induced osteolysis and osteoclastogenesis. CONCLUSION: DOX can effectively inhibit osteoclastogenesis and affect mature osteoclast fate and suppress wear particles induced by osteolysis and osteoclastogenesis. DOX might be useful in the treatment or prevention of wear particles-induced osteolysis and aseptic loosening for its effect on osteoclast generation and mature osteoclast fate and function.

Functional expression of recombinant human stefin A in mammalian and bacterial cells.

Recombinant human cysteine protease inhibitor, stefin A, was expressed in both Escherichia coli and BS-C-1 monkey kidney cells utilizing pET and recombinant vaccinia virus systems, respectively. The expressed protein was purified and analyzed by SDS-PAGE and Western blot analysis utilizing a polyclonal antibody against rat cystatin alpha. In both cases the purified protein appeared as a single band corresponding to the molecular weight of stefin A ( approximately 10kDa). Viability of the expressed stefin A was determined by the inhibition of the plant cysteine protease, papain. Recombinant human stefin A expressed in both E. coli and BS-C-1 cells, was shown to almost completely inhibit papain. The expression of a fully functional recombinant human stefin A in the bacterial system provides a highly efficient tool for the production of large quantities of the protein. This can be an important tool in kinetic studies as well as in production of antibodies for other analytical studies (immunoblot, immunohistochemical studies, etc.). Expression in the mammalian cells, on the other hand, can provide a significant research tool to study the functional roles of stefin A in mammalian systems such as regulation of cysteine proteases.

Association between UHMWPE particle-induced inflammatory osteoclastogenesis and expression of RANKL, VEGF, and Flt-1 in vivo.

Wear debris-induced vascularized granulomatous periprosthetic tissue may augment the progress of prosthetic loosening, a major clinical problem after total joint replacement. The purpose of this study is to investigate the association of ultra-high-molecular-weight polyethylene (UHMWPE) particle-induced inflammatory osteoclastogenesis and expression of RANK/RANKL and VEGF/VEGF receptors (Flt-1 and Flk-1) using a mouse osteolysis model. UHMWPE particles were introduced into established air pouches on BALB/c mice, followed by implantation of calvaria bone from syngeneic littermates. Mice were injected with either recombinant VEGF or VEGF inhibitor (VEGF R2/F(c) Chimera). Mice without drug treatment, as well as mice injected with saline alone were included. Each group contains 10 mice. Pouch tissues were harvested 2 weeks after bone implantation for histological and molecular analysis. UHMWPE stimulation significantly increased VEGF gene expression, and exerted a lower enhancement effect on the gene expression of Flt-1 and Flk-1. UHMWPE-stimulated VEGF production was markedly reduced by VEGF inhibitor treatment. Immunofluorescent staining indicated that pouch tissue macrophages were the main source of both VEGF and Flt-1 production. A positive association was observed between tissue inflammation and the levels of VEGF and Flt-1 gene transcripts. Both RANK and RANKL gene transcripts were significantly increased by UHMWPE stimulation, which was subsequently reduced by VEGF inhibitor treatment (p<0.05). VEGF treatment increased TRAP(+) cells in pouches either with or without UHMWPE particle stimulation, and VEGF inhibitor treatment caused a significant reduction in the number of TRAP(+) cells in UHMWPE-containing pouches. This study suggests that VEGF has a role in the regulation of RANK/RANKL-mediated osteoclastogenesis, and warrant future investigations to elucidate the role of VEGF signaling in the pathogenesis of prosthetic loosening.