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Wednesday, September 26, 2018 7:35 AM–9:00 AM ePosters
The Spine Journal 18 (2018) S142 S225
NASS 33rd Annual Meeting (2018) – Proceedings
Wednesday, September 26, 2018 7:35 AM–9:00 AM ePosters P1. NF-κB inhibitor suppresses edema and promotes rhBMP-2mediated bone formation in spinal fusion D1XJuliane X GlaeserD,2X X PhD1, Phillip D3X X H. BehrensD4X,X MD2, Khosrowdad D5X X SalehiD,6X X BS2, 3 4 D9X X SheynD,10X X PhD, MSc , Zachary D1X X M. NaPierD,12X X MD5, D7XLea X KanimD,8X X MA , Dmitriy D13X X Jason M. CuellarD,14X X MD, PhD6, Hyun D15X X W. BaeD,16X X MD7; 1 Department of Orthopedics, Department of Surgery, Cedars-Sinai Medical Center, Board of Governors Regenerative Medicine Institute, Los Angeles, CA, USA; 2 Cedars-Sinai Medical Center, Los Angeles , CA, USA; 3 Spine Center, Cedars-Sinai Medical Center, Los Angeles, CA, USA; 4 Cedars-Sinai Medical Center, West Hollywood, CA, USA; 5 Cedars Sinai Orthopaedic Surgery, Los Angeles, CA, USA; 6 Los Angeles, CA, USA; 7 Spine Institute St. John’s Health Center, Los Angeles, CA, USA BACKGROUND CONTEXT: Loading of absorbable collagen sponges (ACS) with recombinant human bone morphogenic protein-2 (rhBMP-2) has been successfully used to enhance bone formation and to induce spinal fusion. However, side effects, such as soft-tissue edema and inflammation, have been reported. NEMO binding domain peptide (NBD) inhibits activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), a central regulator of immune response. PURPOSE: To investigate NBD's potential to reduce rhBMP-2-induced soft-tissue inflammation and to stimulate spinal fusion. STUDY DESIGN/SETTING: To evaluate inflammation, ACS containing either high dose rhBMP-2, rhBMP-2+NBD, NBD only or buffer were only implanted into intramuscular fusion beds of 32 rats. To analyze new bone formation in the presence of NBD, posterolateral intertransverse lumbar fusion procedures were performed on 16 rats. ACS implants were loaded with rhBMP-2 or rhBMP-2+NBD. PATIENT SAMPLE: No patients, animal study. OUTCOME MEASURES: T2-weighted relaxation time (T2-RT), histological analysis (H&E and Masson's trichrome staining), NF-κB binding assay, qPCR, manual palpation, and micro-computed tomography (µCT). METHODS: Edema formation at the implant sites was assessed using magnetic resonance imaging T2-weighted relaxation time (T2-RT). Cellular activity was measured by histological analysis of the implant-surrounding zones. NF-κB binding and gene expression of inflammatory markers, interleukin (IL)1β, IL6, IL18, chemokine ligand (CCL)2 and CCL3 were analyzed in the implants. Fusion efficacy was measured by manual palpation, µCT and bone histology. Statistics: separately for each dependent measure, analysis of variance was performed; appropriate post hoc tests for multiple comparisons using Tukey's honestly significant was applied. p≤.05 was considered significant. RESULTS: T2-RT values were increased in the BMP-2 group compared to BMP-2+NBD, NBD and ACS groups. No difference was detected between BMP-2+NBD versus NBD and ACS controls. Histological analysisof the implant-surrounding zones showed an increase in cellular activity in the BMP-2 group compared to BMP-2+NBD and controls. Presence of rhBMP-2 increased relative NF-κB binding and gene expression of inflammatory markers, interleukin (IL)1β, IL6, IL18, chemokine ligand (CCL)2 and CCL3 compared to controls. In the BMP-2+NBD group, cytokine
expression was blocked. No differences were found between BMP-2 +NBD and control groups. BMP-2+NBD resulted in a higher bone volume and reduced trabecular spacing compared to BMP-2, a higher number of levels fused, and similar structural properties of the bone tissue. CONCLUSIONS: In summary, NBD reduces soft-tissue edema formation, reduces recruitment of inflammatory cells, diminishes NF-κB binding and blocks transcription of NF-κB-regulated cytokines in response to highdose rhBMP-2 in rats. Furthermore, NBD stimulates bone formation in rhBMP-2-mediated spinal fusion, potentially through cross-talk of the NFκB pathway with other pathways. The results of this study might provide the basis to develop new therapeutic approaches for spinal fusion using graft material with a combinatory administration of rhBMP-2 and NBD. FDA DEVICE/DRUG STATUS: This abstract does not discuss or include any applicable devices or drugs. https://doi.org/10.1016/j.spinee.2018.06.539
P2. Methylene blue is an effective disclosing agent for identification of bacterial biofilms on spinal implants Jeremy D. Shaw, MD1, Nicholas N. Ashton, PhD2, Jeremy M. Gililland, MD3, Darrel S. Brodke, MD4, Brandon D. Lawrence, MD4, Erik N. Hansen, MD5, Dustin L. Williams, PhD3; 1 University of Utah Orthopaedics, Salt Lake City, UT, USA; 2 Salt Lake City, UT, USA; 3 University of Utah, Salt Lake City, UT, USA; 4 University Orthopaedic Center, Salt Lake City, UT, USA; 5 UCSF, San Francisco, CA, USA BACKGROUND CONTEXT: Tenacious bacterial biofilms pose a major challenge in treating deep spine infections. Biofilms provide bacteria substantial protection against antimicrobial agents, the host immune response, and are invisible to the naked eye. Biofilms are notoriously difficult to eradicate, and to that end, methylene blue has shown promise as a biofilm disclosing agent in the arthroplasty literature in both in vitro and in vivo settings. PURPOSE: The objective of the present study was to assess the intensity of methylene blue staining at varying concentrations as a biofilm disclosing agent in vitro for common biofilm forming bacterial infections and to determine performance characteristics across a range of spine implant materials. STUDY DESIGN/SETTING: Microbiology. METHODS: S. aureus biofilms were grown to maturity in bioreactors according to established lab protocol on titanium, cobalt chromium, and polyetherketone (PEEK) wafers. Biofilms were stained with 0.05% and 0.01% methylene blue solutions for 5-minutes and then washed with normal saline for 1-minute. Gross images were obtained to compare the visual sensitivity of the blue dye at different dilutions. Scanning electron microscopy was performed to confirm the presence or absence of biofilm on methylene blue stained areas. Images were compared to controls. RESULTS: S. aureus biofilms were grown for 7 days on double sided titanium, cobalt chromium, and PEEK wafers (n=4 each). There appeared to be a visible dose – dependent relationship based on the staining and dye concentration. At each dilution, biofilms demonstrated visible blue staining after immersion in methylene blue solution; however, blue dye was visible only where biofilms were present as confirmed by SEM. There was no evidence that methylene blue was able to stain titanium, cobalt chromium, or PEEK.
Refer to onsite annual meeting presentations and postmeeting proceedings for possible referenced figures and tables. Authors are responsible for accurately reporting disclosure and FDA device/drug status at time of abstract submission.
NASS 33rd Annual Meeting (2018) – Proceedings
Wednesday, September 26, 2018 7:35 AM–9:00 AM ePosters P1. NF-κB inhibitor suppresses edema and promotes rhBMP-2mediated bone formation in spinal fusion D1XJuliane X GlaeserD,2X X PhD1, Phillip D3X X H. BehrensD4X,X MD2, Khosrowdad D5X X SalehiD,6X X BS2, 3 4 D9X X SheynD,10X X PhD, MSc , Zachary D1X X M. NaPierD,12X X MD5, D7XLea X KanimD,8X X MA , Dmitriy D13X X Jason M. CuellarD,14X X MD, PhD6, Hyun D15X X W. BaeD,16X X MD7; 1 Department of Orthopedics, Department of Surgery, Cedars-Sinai Medical Center, Board of Governors Regenerative Medicine Institute, Los Angeles, CA, USA; 2 Cedars-Sinai Medical Center, Los Angeles , CA, USA; 3 Spine Center, Cedars-Sinai Medical Center, Los Angeles, CA, USA; 4 Cedars-Sinai Medical Center, West Hollywood, CA, USA; 5 Cedars Sinai Orthopaedic Surgery, Los Angeles, CA, USA; 6 Los Angeles, CA, USA; 7 Spine Institute St. John’s Health Center, Los Angeles, CA, USA BACKGROUND CONTEXT: Loading of absorbable collagen sponges (ACS) with recombinant human bone morphogenic protein-2 (rhBMP-2) has been successfully used to enhance bone formation and to induce spinal fusion. However, side effects, such as soft-tissue edema and inflammation, have been reported. NEMO binding domain peptide (NBD) inhibits activation of nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB), a central regulator of immune response. PURPOSE: To investigate NBD's potential to reduce rhBMP-2-induced soft-tissue inflammation and to stimulate spinal fusion. STUDY DESIGN/SETTING: To evaluate inflammation, ACS containing either high dose rhBMP-2, rhBMP-2+NBD, NBD only or buffer were only implanted into intramuscular fusion beds of 32 rats. To analyze new bone formation in the presence of NBD, posterolateral intertransverse lumbar fusion procedures were performed on 16 rats. ACS implants were loaded with rhBMP-2 or rhBMP-2+NBD. PATIENT SAMPLE: No patients, animal study. OUTCOME MEASURES: T2-weighted relaxation time (T2-RT), histological analysis (H&E and Masson's trichrome staining), NF-κB binding assay, qPCR, manual palpation, and micro-computed tomography (µCT). METHODS: Edema formation at the implant sites was assessed using magnetic resonance imaging T2-weighted relaxation time (T2-RT). Cellular activity was measured by histological analysis of the implant-surrounding zones. NF-κB binding and gene expression of inflammatory markers, interleukin (IL)1β, IL6, IL18, chemokine ligand (CCL)2 and CCL3 were analyzed in the implants. Fusion efficacy was measured by manual palpation, µCT and bone histology. Statistics: separately for each dependent measure, analysis of variance was performed; appropriate post hoc tests for multiple comparisons using Tukey's honestly significant was applied. p≤.05 was considered significant. RESULTS: T2-RT values were increased in the BMP-2 group compared to BMP-2+NBD, NBD and ACS groups. No difference was detected between BMP-2+NBD versus NBD and ACS controls. Histological analysisof the implant-surrounding zones showed an increase in cellular activity in the BMP-2 group compared to BMP-2+NBD and controls. Presence of rhBMP-2 increased relative NF-κB binding and gene expression of inflammatory markers, interleukin (IL)1β, IL6, IL18, chemokine ligand (CCL)2 and CCL3 compared to controls. In the BMP-2+NBD group, cytokine
expression was blocked. No differences were found between BMP-2 +NBD and control groups. BMP-2+NBD resulted in a higher bone volume and reduced trabecular spacing compared to BMP-2, a higher number of levels fused, and similar structural properties of the bone tissue. CONCLUSIONS: In summary, NBD reduces soft-tissue edema formation, reduces recruitment of inflammatory cells, diminishes NF-κB binding and blocks transcription of NF-κB-regulated cytokines in response to highdose rhBMP-2 in rats. Furthermore, NBD stimulates bone formation in rhBMP-2-mediated spinal fusion, potentially through cross-talk of the NFκB pathway with other pathways. The results of this study might provide the basis to develop new therapeutic approaches for spinal fusion using graft material with a combinatory administration of rhBMP-2 and NBD. FDA DEVICE/DRUG STATUS: This abstract does not discuss or include any applicable devices or drugs. https://doi.org/10.1016/j.spinee.2018.06.539
P2. Methylene blue is an effective disclosing agent for identification of bacterial biofilms on spinal implants Jeremy D. Shaw, MD1, Nicholas N. Ashton, PhD2, Jeremy M. Gililland, MD3, Darrel S. Brodke, MD4, Brandon D. Lawrence, MD4, Erik N. Hansen, MD5, Dustin L. Williams, PhD3; 1 University of Utah Orthopaedics, Salt Lake City, UT, USA; 2 Salt Lake City, UT, USA; 3 University of Utah, Salt Lake City, UT, USA; 4 University Orthopaedic Center, Salt Lake City, UT, USA; 5 UCSF, San Francisco, CA, USA BACKGROUND CONTEXT: Tenacious bacterial biofilms pose a major challenge in treating deep spine infections. Biofilms provide bacteria substantial protection against antimicrobial agents, the host immune response, and are invisible to the naked eye. Biofilms are notoriously difficult to eradicate, and to that end, methylene blue has shown promise as a biofilm disclosing agent in the arthroplasty literature in both in vitro and in vivo settings. PURPOSE: The objective of the present study was to assess the intensity of methylene blue staining at varying concentrations as a biofilm disclosing agent in vitro for common biofilm forming bacterial infections and to determine performance characteristics across a range of spine implant materials. STUDY DESIGN/SETTING: Microbiology. METHODS: S. aureus biofilms were grown to maturity in bioreactors according to established lab protocol on titanium, cobalt chromium, and polyetherketone (PEEK) wafers. Biofilms were stained with 0.05% and 0.01% methylene blue solutions for 5-minutes and then washed with normal saline for 1-minute. Gross images were obtained to compare the visual sensitivity of the blue dye at different dilutions. Scanning electron microscopy was performed to confirm the presence or absence of biofilm on methylene blue stained areas. Images were compared to controls. RESULTS: S. aureus biofilms were grown for 7 days on double sided titanium, cobalt chromium, and PEEK wafers (n=4 each). There appeared to be a visible dose – dependent relationship based on the staining and dye concentration. At each dilution, biofilms demonstrated visible blue staining after immersion in methylene blue solution; however, blue dye was visible only where biofilms were present as confirmed by SEM. There was no evidence that methylene blue was able to stain titanium, cobalt chromium, or PEEK.
Refer to onsite annual meeting presentations and postmeeting proceedings for possible referenced figures and tables. Authors are responsible for accurately reporting disclosure and FDA device/drug status at time of abstract submission.