Real-time identification of life-threatening necrotizing soft-tissue infections using indocyanine green fluorescence imaging.

Significance: Necrotizing soft-tissue infections (NSTIs) are life-threatening infections with a cumulative case fatality rate of 21%. The initial presentation of an NSTI is non-specific, frequently leading to misdiagnosis and delays in care. No current strategies yield an accurate, real-time diagnosis of an NSTI. Aim: A first-in-kind, observational, clinical pilot study tested the hypothesis that measurable fluorescence signal voids occur in NSTI-affected tissues following intravenous administration and imaging of perfusion-based indocyanine green (ICG) fluorescence. This hypothesis is based on the established knowledge that NSTI is associated with local microvascular thrombosis. Approach: Adult patients presenting to the Emergency Department of a tertiary care medical center at high risk for NSTI were prospectively enrolled and imaged with a commercial fluorescence imager. Single-frame fluorescence snapshot and first-pass perfusion kinetic parameters-ingress slope (IS), time-to-peak (TTP) intensity, and maximum fluorescence intensity (IMAX)-were quantified using a dynamic contrast-enhanced fluorescence imaging technique. Clinical variables (comorbidities, blood laboratory values), fluorescence parameters, and fluorescence signal-to-background ratios (SBRs) were compared to final infection diagnosis. Results: Fourteen patients were enrolled and imaged (six NSTI, six cellulitis, one diabetes mellitus-associated gangrene, and one osteomyelitis). Clinical variables demonstrated no statistically significant differences between NSTI and non-NSTI patient groups (p-value≥0.22). All NSTI cases exhibited prominent fluorescence signal voids in affected tissues, including tissue features not visible to the naked eye. All cellulitis cases exhibited a hyperemic response with increased fluorescence and no distinct signal voids. Median lesion-to-background tissue SBRs based on snapshot, IS, TTP, and IMAX parameter maps ranged from 3.2 to 9.1, 2.2 to 33.8, 1.0 to 7.5, and 1.5 to 12.7, respectively, for the NSTI patient group. All fluorescence parameters except TTP demonstrated statistically significant differences between NSTI and cellulitis patient groups (p-value<0.05). Conclusions: Real-time, accurate discrimination of NSTIs compared with non-necrotizing infections may be possible with perfusion-based ICG fluorescence imaging.

Neoadjuvant Therapies Do Not Reduce Epidermal Growth Factor Receptor (EGFR) Expression or EGFR-Targeted Fluorescence in a Murine Model of Soft-Tissue Sarcomas.

PURPOSE: ABY-029, an epidermal growth factor receptor (EGFR)-targeted, synthetic Affibody peptide labeled with a near-infrared fluorophore, is under investigation for fluorescence-guided surgery of sarcomas. To date, studies using ABY-029 have occurred in tumors naïve to chemotherapy (CTx) and radiation therapy (RTx), although these neoadjuvant therapies are frequently used for sarcoma treatment in humans. The goal of this study was to evaluate the impact of CTx and RTx on tumor EGFR expression and ABY-029 fluorescence of human soft-tissue sarcoma xenografts in a murine model. PROCEDURES: Immunodeficient mice (n = 98) were divided into five sarcoma xenograft groups and three treatment groups - CTx only, RTx only, and CTx followed by RTx, plus controls. Four hours post-injection of ABY-029, animals were sacrificed followed by immediate fluorescence imaging of ex vivo adipose, muscle, nerve, and tumor tissues. Histological hematoxylin and eosin staining confirmed tumor type, and immunohistochemistry staining determined EGFR, cluster of differentiation 31 (CD31), and smooth muscle actin (SMA) expression levels. Correlation analysis (Pearson's correlation coefficients, r) and linear regression (unstandardized coefficient estimates, B) were used to determine statistical relationships in molecular expression and tissue fluorescence between xenografts and treatment groups. RESULTS: Neoadjuvant therapies had no broad impact on EGFR expression (|B|≤ 7.0, p ≥ 0.4) or on mean tissue fluorescence (any tissue type, (|B|≤ 2329.0, p ≥ 0.1). Mean tumor fluorescence was significantly related to EGFR expression (r = 0.26, p = 0.01), as expected. CONCLUSION: Results suggest that ABY-029 as an EGFR-targeted, fluorescent probe is not negatively impacted by neoadjuvant soft-tissue sarcoma therapies, although validation in humans is required.

Pathologic Femur Fracture in an Immunocompetent Healthy Young Adult due to Acute Osteomyelitis.

An immunocompetent 33-year-old woman presented with a pathologic femur fracture after one month of progressively worsening right thigh pain. Open biopsy demonstrated acute suppurative osteomyelitis despite the lack of clinical risk factors. The polymicrobial infection was successfully treated with three operative procedures and culture-specific antibiotic agents. Acute osteomyelitis, while an uncommon cause of pathologic fracture, must always be on the differential diagnosis, even when no obvious predisposing factors are present. When investigating for an infectious etiology in cases such as our own, considering immunodeficiency syndromes alongside the more typical causes of osteomyelitis is encouraged.

Bone Tumor Risk Stratification and Management System: A Consensus Guideline from the ACR Bone Reporting and Data System Committee.

The assessment and subsequent management of a potentially neoplastic bone lesion seen at diagnostic radiography is often complicated by diagnostic uncertainty and inconsistent management recommendations. Appropriate clinical management should be directed by risk of malignancy. Herein, the ACR-sponsored Bone Reporting and Data System (Bone-RADS) Committee, consisting of academic leaders in the fields of musculoskeletal oncology imaging and orthopedic oncology, presents the novel Bone-RADS scoring system to aid in risk assignment and provide risk-aligned management suggestions. When viewed in the proper clinical context, a newly identified bone lesion can be risk stratified as having very low, low, intermediate, or high risk of malignancy. Radiographic features predictive of risk are reviewed include margination, pattern of periosteal reaction, depth of endosteal erosion, pathological fracture, and extra-osseous soft tissue mass. Other radiographic features predictive of histopathology are also briefly discussed. To apply the Bone-RADS scoring system to a potentially neoplastic bone lesion, radiographic features predictive of risk are each given a point value. Point values are summed to yield a point total, which can be translated to a Bone-RADS score (1-4) with corresponding risk assignment (very low, low, intermediate, high). For each score, evidence-based and best practice consensus management suggestions are outlined. Examples of each Bone-RADS scores are presented, and a standardized diagnostic radiography report template is provided.

In vitro transcription of self-assembling DNA nanoparticles.

Nucleic acid nanoparticles are playing an increasingly important role in biomolecular diagnostics and therapeutics as well as a variety of other areas. The unique attributes of self-assembling DNA nanoparticles provide a potentially valuable addition or alternative to the lipid-based nanoparticles that are currently used to ferry nucleic acids in living systems. To explore this possibility, we have assessed the ability of self-assembling DNA nanoparticles to be constructed from complete gene cassettes that are capable of gene expression in vitro. In the current report, we describe the somewhat counter-intuitive result that despite extensive crossovers (the stereochemical analogs of Holliday junctions) and variations in architecture, these DNA nanoparticles are amenable to gene expression as evidenced by T7 RNA polymerase-driven transcription of a reporter gene in vitro. These findings, coupled with the vastly malleable architecture and chemistry of self-assembling DNA nanoparticles, warrant further investigation of their utility in biomedical genetics.

Use of Freshly Amputated Human Limbs for Pre-Clinical Evaluation of Molecular-Targeted Fluorescent Probes.

We have co-developed a first-in-kind model of fluorophore testing in freshly amputated human limbs. Ex vivo human tissue provides a unique opportunity for the testing of pre-clinical fluorescent agents, collection of imaging data, and histopathologic examination in human tissue prior to performing in vivo experiments. Existing pre-clinical fluorescent agent studies rely primarily on animal models, which do not directly predict fluorophore performance in humans and can result in wasted resources and time if an agent proves ineffective in early human trials. Because fluorophores have no desired therapeutic effect, their clinical utility is based solely on their safety and ability to highlight tissues of interest. Advancing to human trials even via the FDA's phase 0/microdose pathway still requires substantial resources, single-species pharmacokinetic testing, and toxicity testing. In a recently concluded study using amputated human lower limbs, we were able to test successfully a nerve-specific fluorophore in pre-clinical development. This study used systemic administration via vascular cannulization and a cardiac perfusion pump. We envision that this model may assist with early lead agent testing selection for fluorophores with various targets and mechanisms.

First demonstration of a novel nerve-targeting fluorophore in a cohort of ex vivo human tissues.

Iatrogenic nerve injury is a common complication across all surgical specialties. Better nerve visualization and identification during surgery will improve outcomes and reduce nerve injuries. The Gibbs Laboratory at Oregon Health and Science University has developed a library of near-infrared, nerve-specific fluorophores to highlight nerves intraoperatively and aid surgeons in nerve identification and visualization; the current lead agent is LGW16-03. Prior to this study, testing of LGW16-03 was restricted to animal models; therefore, it was unknown how LGW16-03 performs in human tissue. To advance LGW16-03 to clinic, we sought to test this current lead agent in ex vivo human tissues from a cohort of patients and determine if the route of administration affects LGW16-03 fluorescence contrast between nerves and adjacent background tissues (muscle and adipose). LGW16-03 was applied to ex vivo human tissue from lower limb amputations via two strategies: (1) systemic administration of the fluorophore using our first-in-kind model for fluorophore testing, and (2) topical application of the fluorophore. Results showed no statistical difference between topical and systemic administration. However, in vivo human validation of these findings is required.

Early identification of life-threatening soft-tissue infection using dynamic fluorescence imaging: first-in-kind clinical study of first-pass kinetics.

Necrotizing soft-tissue infections (NSTIs) are aggressive and deadly. Immediate surgical debridement is standard-of-care, but patients often present with non-specific symptoms, thereby delaying treatment. Because NSTIs cause microvascular thrombosis, we hypothesized that perfusion imaging using indocyanine green (ICG) would show diminished fluorescence signal in NSTI-affected tissues, particularly compared to non-necrotizing, superficial infections. Through a first-in-kind clinical study, we performed first-pass ICG fluorescence perfusion imaging of patients with suspected NSTIs. Early results support our hypothesis that ICG signal voids occur in NSTI-affected tissues and that dynamic contrast-enhanced fluorescence parameters reveal tissue kinetics that may be related to disease progression and extent.

Proceduralist criteria for evaluating interface utility of novel imaging modalities in early phase clinical trials: evaluating the need for standardized criteria.

Accelerating innovation in the space of fluorescence imaging for surgical applications has increased interest in safely and expediently advancing these technologies to clinic through Food and Drug Administration-(FDA-) compliant trials. Conventional metrics for early phase trials include drug safety, tolerability, dosing, and pharmacokinetics. Most procedural imaging technologies rely on administration of an exogenous fluorophore and concurrent use of an imaging system; both of which must receive FDA approval to proceed to clinic. Because fluorophores are classified as medical imaging agents, criteria for establishing dose are different, and arguably more complicated, than therapeutic drugs. Since no therapeutic effect is desired, medical imaging agents are ideally administered at the lowest dose that achieves adequate target differentiation. Because procedural imaging modalities are intended to enhance and/or ease proceduralists' identification or assessment of tissues, beneficial effects of these technologies may manifest in the form of qualitative endpoints such as: 1) confidence; 2) decision-making; and 3) satisfaction with the specified procedure. Due to the rapid expansion of medical imaging technologies, we believe that our field requires standardized criteria to evaluate existing and emerging technologies objectively so that both quantitative and qualitative aspects of their use may be measured and useful comparisons to assess their relative value may occur. Here, we present a 15-item consensus-based survey instrument to assess the utility of novel imaging technologies from the proceduralist's standpoint.

Automated motion artifact correction for dynamic contrast-enhanced fluorescence imaging during open orthopedic surgery.

Indocyanine green (ICG)-based dynamic contrast-enhanced fluorescence imaging (DCE-FI) can objectively assess bone perfusion intraoperatively. However, it is susceptible to motion artifacts due to patient's involuntary respiration during the 4.5-minute DCE-FI data acquisition. An automated motion correction approach based on mutual information (MI) frameby-frame was developed to overcome this problem. In this approach, MIs were calculated between the reference and the adjacent frame translated and the maximal MI corresponded to the optimal translation. The images obtained from eighteen amputation cases were utilized to validate the approach and the results show that this correction can significantly reduce the motion artifacts and can improve the accuracy of bone perfusion assessment.

Acceptability and Feasibility of Delivering Decision Aids to Veterans for Management of Knee Osteoarthritis - A Pilot Study.

Introduction: Decision aids are effective tools in facilitating patient-centered care and patient involvement in the decision-making process. Given unique barriers to providing patient-centered care for Veterans, implementation of decision aids may improve overall quality of care. We aimed to assess the acceptability and feasibility of video-based and pamphlet-based decision aid use in Veterans with knee osteoarthritis. Materials and Methods: Veterans considering treatment for knee osteoarthritis received either an online video-based aid, pamphlet-based aid, or both before their surgical consult. At their visit, patients completed written pre-visit and post-visit questionnaires. The pre-visit questionnaire included questions about the patient's demographics, decision-making preferences, experiences using the assigned decision aids, and the Hip-Knee Decision Quality Instrument. The post-visit questionnaire assessed the patient's overall experience with the decision-making process and how use of the decision aid influenced their discussion with the physician. Results: All 16 patients who received the pamphlet-based aid reviewed the decision aid before their visit, compared to only five of the 12 patients who received the video-based aid. Thirteen of 20 patients indicated that they preferred to share treatment decision-making with their physician. Seventeen of 20 patients believed they would feel comfortable questioning the treatment recommendation of their surgeon after decision aid use. Most patients reported a positive experience using their decision aid, regardless of modality, and found it easily comprehensible and useful in visit preparation. A preference for a pamphlet-based aid was expressed by the majority of patients. Conclusion: Veterans considering treatment for knee osteoarthritis are well prepared to engage in a patient-centered care experience. Most patients preferred sharing the decision-making process with their physician and felt comfortable questioning them about treatment recommendations. Decision aids helped Veterans feel more informed about their treatment options and improved engagement and discussion with their physician. Pamphlet-based aids were utilized more reliably than video-based aids.

Preclinical evaluation of molecularly targeted fluorescent probes in perfused amputated human limbs.

Significance: This first-in-kind, perfused, and amputated human limb model allows for the collection of human data in preclinical selection of lead fluorescent agents. The model facilitates more accurate selection and testing of fluorophores with human-specific physiology, such as differential uptake and signal in fat between animal and human models with zero risk to human patients. Preclinical testing using this approach may also allow for the determination of tissue toxicity, clearance time of fluorophores, and the production of harmful metabolites. Aim: This study was conducted to determine the fluorescence intensity values and tissue specificity of a preclinical, nerve tissue targeted fluorophore, as well as the capacity of this first-in-kind model to be used for lead fluorescent agent selection in the future. Approach: Freshly amputated human limbs were perfused for 30 min prior to in situ and ex vivo imaging of nerves with both open-field and closed-field commercial fluorescence imaging systems. Results: In situ, open-field imaging demonstrated a signal-to-background ratio (SBR) of 4.7 when comparing the nerve with adjacent muscle tissue. Closed-field imaging demonstrated an SBR of 3.8 when the nerve was compared with adipose tissue and 4.8 when the nerve was compared with muscle. Conclusions: This model demonstrates an opportunity for preclinical testing, evaluation, and selection of fluorophores for use in clinical trials as well as an opportunity to study peripheral pathologies in a controlled environment.

Current and Future Applications of Fluorescence Guidance in Orthopaedic Surgery.

Fluorescence-guided surgery (FGS) is an evolving field that seeks to identify important anatomic structures or physiologic phenomena with helpful relevance to the execution of surgical procedures. Fluorescence labeling occurs generally via the administration of fluorescent reporters that may be molecularly targeted, enzyme-activated, or untargeted, vascular probes. Fluorescence guidance has substantially changed care strategies in numerous surgical fields; however, investigation and adoption in orthopaedic surgery have lagged. FGS shows the potential for improving patient care in orthopaedics via several applications including disease diagnosis, perfusion-based tissue healing capacity assessment, infection/tumor eradication, and anatomic structure identification. This review highlights current and future applications of fluorescence guidance in orthopaedics and identifies key challenges to translation and potential solutions.

Fluorescence guidance improves the accuracy of radiological imaging-guided surgical navigation.

BACKGROUND: Imaging-based navigation technologies require static referencing between the target anatomy and the optical sensors. Imaging-based navigation is therefore well suited to operations involving bony anatomy; however, these technologies have not translated to soft-tissue surgery. We sought to determine if fluorescence imaging complement conventional, radiological imaging-based navigation to guide the dissection of soft-tissue phantom tumors. METHODS: Using a human tissue-simulating model, we created tumor phantoms with physiologically accurate optical density and contrast concentrations. Phantoms were dissected using all possible combinations of computed tomography (CT), magnetic resonance, and fluorescence imaging; controls were included. The data were margin accuracy, margin status, tumor spatial alignment, and dissection duration. RESULTS: Margin accuracy was higher for combined navigation modalities compared to individual navigation modalities, and accuracy was highest with combined CT and fluorescence navigation (p = 0.045). Margin status improved with combined CT and fluorescence imaging. CONCLUSIONS: At present, imaging-based navigation has limited application in guiding soft-tissue tumor operations due to its inability to compensate for positional changes during surgery. This study indicates that fluorescence guidance enhances the accuracy of imaging-based navigation and may be best viewed as a synergistic technology, rather than a competing one.

Extraneuraxial Hemangioblastoma: An Unusual Soft Tissue Neoplasm that Mimics More Common Entities but Carries Distinct Clinical Implications.

Hemangioblastoma, one of the characteristic tumors associated with Von Hippel-Lindau (VHL) disease, most often presents in the central nervous system (CNS) but can uncommonly arise in extraneuraxial, or previously referred to as peripheral, locations. Without the clinical context of known VHL disease, hemangioblastoma may not enter the differential for a soft tissue mass outside the CNS. Here, we present two patients with diagnostically challenging extraneuraxial hemangioblastoma to highlight the importance of considering this entity within the differential diagnosis of soft tissue neoplasms containing clear cells and delicate vasculature. We review the relevant diagnostic features, including a suggested immunohistochemical panel, along with the potential associated clinical implications of making this diagnosis. It is recommended that affected patients be offered genetic counseling to assess for underlying VHL disease.

Intraoperative assessment of bone viability through improved analysis and visualization of dynamic contrast-enhanced fluorescence imaging: technique report.

Bone devitalization is believed to be a critical determinant of complications such as infection or nonunion. However, intraoperative assessment of bone devitalization, particularly in open fractures and infections, remains highly subjective resulting in variation in treatment. Optical imaging tools, particularly dynamic contrast-enhanced fluorescence imaging, can provide real-time, intraoperative assessment of bone and soft tissue perfusion, which informs the tissues' ability to heal and fight infection. We describe a novel technique to apply indocyanine green-based fluorescence imaging, using a device that is frequently used in the operating room to assess skin or flap perfusion in plastic surgery, to assess bone and deep tissue perfusion in three pertinent cases: (1) a chronic infection/nonunion after a Gustilo type 3A tibia fracture (patient 1), (2) an acute Gustilo type 3C tibia open fracture with extensive degloving/soft tissue stripping (patient 2), and (3) an atrophic nonunion of the humerus (patient 3). In all three cases, fluorescence imaging (both time-specific fluorescence and maximum fluorescence) and derived kinetic maps of time-to-peak, ingress slope, and egress slope demonstrated clear spatial variation in perfusion that corresponded to the patient pathogenesis. The impact of this information on patient outcome will need to be evaluated in future clinical trials; however, these cases demonstrate in principle that optical imaging information has the potential to inform surgical practice, reduce the variation in treatment, and improve outcomes observed in these challenging patients.

Epidermal growth factor-targeted fluorescence is unaffected by standard neoadjuvant therapies in human sarcomas.

Curative surgery for other many cancers requires that the tumor be removed with a zone of normal tissue surrounding the tumor with 'negative' margins. Sarcomas, cancers of the bones, muscles, and fat, require WLE for cure. Unfortunately, 'positive' margins occur in 20-25% of sarcoma surgeries, associated with cancer recurrence and reduced survival. Our group successfully tested a small-molecule fluorophore (ABY-029) in sarcomas that targets the epidermal growth factor receptor. We sought to evaluate human sarcoma xenografts for epidermal growth factor receptor expression and binding of ABY-029 with and without exposure to standard presurgical chemotherapy and radiation. We inoculated groups of 24 NSG mice with five cell lines (120 mice total). Eight mice from each cell line received: 1) radiation alone; 2) chemotherapy alone; or 3) chemotherapy and radiation. We administered ABY-029 2-4 hours before surgery. Tumor and biopsy portions of background tissues were removed. All tissues were imaged on a LI-COR Odyssey and processed in pathology. There were no significant reductions in epidermal growth factor receptor expression or in ABY-029-mediated fluorescence in tumors exposed to chemotherapy, radiation, or both. fluorescence-guided surgery demonstrates strong promise to improve curative surgical cancer care, particularly for sarcomas where the positive margin rate is substantial. Fluorophore performance must be evaluated under circumstances that duplicate accurately the biological milieu relevant to a particular cancer. This work shows that human sarcoma xenografts subjected to standard therapies do not demonstrate a change in epidermal growth factor receptor expression or in epidermal growth factor receptor-targeted fluorescence, thereby indicating that epidermal growth factor receptor-targeted fluorescence-guided surgery should be feasible under normal therapeutic conditions in the clinic.

Dynamic contrast-enhanced fluorescence imaging compared with MR imaging in evaluating bone perfusion during open orthopedic surgery.

ICG-based dynamic contrast-enhanced fluorescence imaging (DCE-FI) and intraoperative DCE- magnetic resonance imaging (MRI) have been carried out nearly simultaneously in three lower extremity bone infection cases to investigate the relationship between these two imaging modalities for assessing bone blood perfusion during open orthopedic surgeries. Time-intensity curves in the corresponding regions of interest of two modalities were derived for comparison. The results demonstrated that ICG-based DCE-FI has higher sensitivity to perfusion changes while DCE-MRI provides superior and supplemental depth-related perfusion information. Research applying the depth-related perfusion information derived from MRI to improve the overall analytic modeling of intraoperative DCE-FI is ongoing.

Use of the PRECICE nail for distraction osteogenesis after tumor resection.

INTRODUCTION: Reconstructing long bone defects following intercalary tumor resection presents an exciting challenge with a greater range of surgical solutions than more typical situations requiring arthroplasty. Segmental bone transport (distraction osteogenesis) is the least utilized option for intercalary reconstruction; however, it arguably provides patients with the most desirable result. Distraction osteogenesis can be used in the management of multiple skeletal conditions including deformity (congenital or acquired), or in the presence of bone defects (by trauma or planned surgical excision). Lack of broader adoption of transport is likely due to the highly technical demands and common complications of utilizing fine-wire fixators via the Ilizarov method. More recently, internal lengthening nails such as the PRECICE nail have been employed to facilitate distraction osteogenesis without the added complexity of external fixation. AREAS COVERED: This review will examine the literature on the indications, design, and safety of the PRECICE nail (NuVasive) for intercalary reconstruction after tumor resection. EXPERT OPINION: Bone transport using the PRECICE nail represents a viable alternative to Ilizarov distraction and has the benefit of avoiding the complications of an external fixator. For large defects, the PRECICE nail can be supplemented with a locking plate for additional stability and maintenance of limb length.

Applications of artificial intelligence in orthopaedic surgery.

The practice of medicine is rapidly transforming as a result of technological breakthroughs. Artificial intelligence (AI) systems are becoming more and more relevant in medicine and orthopaedic surgery as a result of the nearly exponential growth in computer processing power, cloud based computing, and development, and refining of medical-task specific software algorithms. Because of the extensive role of technologies such as medical imaging that bring high sensitivity, specificity, and positive/negative prognostic value to management of orthopaedic disorders, the field is particularly ripe for the application of machine-based integration of imaging studies, among other applications. Through this review, we seek to promote awareness in the orthopaedics community of the current accomplishments and projected uses of AI and ML as described in the literature. We summarize the current state of the art in the use of ML and AI in five key orthopaedic disciplines: joint reconstruction, spine, orthopaedic oncology, trauma, and sports medicine.