Prediction of Wound Failure in Patients with Head and Neck Cancer Treated with Free Flap Reconstruction: Utility of CT Perfusion and MR Perfusion in the Early Postoperative Period.

BACKGROUND AND PURPOSE: Free flap reconstruction in patients with head and neck cancer carries a risk of postoperative complications, and radiologic predictive factors have been limited. The aim of this study was to assess the factors that predict free flap reconstruction failure using CT and MR perfusion. MATERIALS AND METHODS: This single-center prospective study included 24 patients (mean age, 62.7 [SD, 9.0] years; 16 men) who had free flap reconstruction from January 2016 to May 2018. CT perfusion and dynamic contrast-enhanced MR imaging with conventional CT and MR imaging were performed between 2 and 4 days after the free flap surgery, and the wound assessments within 14 days after the surgery were conducted by the surgical team. The parameters of CT perfusion and dynamic contrast-enhanced MR imaging with conventional imaging findings and patient demographics were compared between the patients with successful free flap reconstruction and those with wound failure as appropriate. P < .05 was considered significant. RESULTS: There were 19 patients with successful free flap reconstruction and no wound complications (mean age, 63.9 [SD, 9.5] years; 14 men), while 5 patients had wound failure (mean age, 58.0 [SD, 5.7] years; 2 men). Blood flow, blood volume, MTT, and time maximum intensity projection (P = .007, .007, .015, and .004, respectively) in CT perfusion, and fractional plasma volume, volume transfer constant, peak enhancement, and time to maximum enhancement (P = .006, .039, .004, and .04, respectively) in dynamic contrast-enhanced MR imaging were significantly different between the 2 groups. CONCLUSIONS: CT perfusion and dynamic contrast-enhanced MR imaging are both promising imaging techniques to predict wound complications after head and neck free flap reconstruction.

Development and validation of a clinical prediction-score model for distant metastases in major salivary gland carcinoma.

BACKGROUND: The most common pattern of failure in major salivary gland carcinoma (SGC) is development of distant metastases (DMs). The objective of this study was to develop and validate a prediction score for DM in SGC. PATIENTS AND METHODS: Patients with SGC treated curatively at four tertiary cancer centers were divided into discovery (n = 619) and validation cohorts (n = 416). Multivariable analysis using competing risk regression was used to identify predictors of DM in the discovery cohort and create a prediction score of DM; the optimal score cut-off was determined using a minimal P value approach. The prediction score was subsequently evaluated in the validation cohort. The cumulative incidence and Kaplan-Meier methods were used to analyze DM and overall survival (OS), respectively. RESULTS: In the discovery cohort, DM predictors (risk coefficient) were: positive margin (0.6), pT3-4 (0.7), pN+ (0.7), lymphovascular invasion (0.8), and high-risk histology (1.2). High DM-risk SGC was defined by sum of coefficients greater than two. In the discovery cohort, the 5-year incidence of DM for high- versus low-risk SGC was 50% versus 8% (P < 0.01); this was similar in the validation cohort (44% versus 4%; P < 0.01). In the pooled cohorts, this model performed similarly in predicting distant-only failure (40% versus 6%, P < 0.01) and late (>2 years post surgery) DM (22% versus 4%; P < 0.01). Patients with high-risk SGC had an increased incidence of DM in the subgroup receiving postoperative radiation therapy (46% versus 8%; P < 0.01). The 5-year OS for high- versus low-risk SGC was 48% versus 92% (P < 0.01). CONCLUSION: This validated prediction-score model may be used to identify SGC patients at increased risk for DM and select those who may benefit from prospective evaluation of treatment intensification and/or surveillance strategies.

Predictors and Prevalence of Nodal Disease in Salvage Oropharyngectomy.

BACKGROUND: Patients with recurrent oropharyngeal cancer often require extensive salvage surgery. For patients with clinically N0 necks, the indication for concurrent neck dissection remains unclear. This study aimed to determine predictors, prevalence, and distribution of nodal disease in patients treated with salvage oropharyngectomy. METHODS: In a case series with data collection at a single tertiary academic National Cancer Institute (NCI)-designated comprehensive cancer center, this study analyzed patients treated with prior radiation or chemoradiation who had persistent, recurrent, or second primary squamous cell carcinoma of the oropharynx requiring oropharyngeal resection between 1998 and 2017 (n = 95). Clinical and oncologic characteristics and treatment outcomes were collected, and statistical analyses were performed. RESULTS: The overall rate of nodal positivity was 21% (24/95), and the rate of occult nodal disease was 6% (4/65). Ipsilateral and contralateral level 2 were the most common areas harboring positive nodes. Bivariate analysis showed female sex (p = 0.01), initial overall stage (p = 0.02), and N status (p = 0.03), as well as recurrent overall and T stage (p = 0.05) to be predictors of nodal disease. In the multivariate analysis, recurrent T stage continued to be significantly predictive of pathologic nodal disease. Both computed tomography (CT) and positron emission tomography-CT were moderately accurate in predicting nodal disease in the salvage setting (area under the curve, 0.79 and 0.80, respectively). CONCLUSION: Occult nodal disease is observed in few patients undergoing salvage oropharyngeal resection. This study identified factors predictive of nodal disease in patients undergoing salvage oropharyngectomy and appropriate diagnostic tests in this setting.

Tracking ultrafast hot-electron diffusion in space and time by ultrafast thermomodulation microscopy.

The ultrafast response of metals to light is governed by intriguing nonequilibrium dynamics involving the interplay of excited electrons and phonons. The coupling between them leads to nonlinear diffusion behavior on ultrashort time scales. Here, we use scanning ultrafast thermomodulation microscopy to image the spatiotemporal hot-electron diffusion in thin gold films. By tracking local transient reflectivity with 20-nm spatial precision and 0.25-ps temporal resolution, we reveal two distinct diffusion regimes: an initial rapid diffusion during the first few picoseconds, followed by about 100-fold slower diffusion at longer times. We find a slower initial diffusion than previously predicted for purely electronic diffusion. We develop a comprehensive three-dimensional model based on a two-temperature model and evaluation of the thermo-optical response, taking into account the delaying effect of electron-phonon coupling. Our simulations describe well the observed diffusion dynamics and let us identify the two diffusion regimes as hot-electron and phonon-limited thermal diffusion, respectively.

Novel Immunotherapeutic Approaches in Head and Neck Cancer.

Unresectable recurrent or metastatic head and neck cancer is an incurable disease with survival of approximately 12 months. Head and neck tumors exhibit numerous derangements in the tumor microenvironment that aid in immune evasion and may serve as targets for future therapies. Pembrolizumab is now approved as a first line therapy. Despite the promise of currently approved immunotherapies there continues to be low response rates and additional strategies are needed. Here, alterations in the immune microenvironment and current therapeutic strategies are reviewed with a focus on novel immunologic approaches.

Surveilling the Potential for Precision Medicine-driven PD-1/PD-L1-targeted Therapy in HNSCC.

Immunotherapy is becoming an accepted treatment modality for many patients with cancer and is now approved for use in platinum-refractory recurrent or metastatic head and neck squamous cell carcinoma (HNSCC). Despite these successes, a minority of patients with HNSCC receiving immunotherapy respond to treatment, and few undergo a complete response. Thus, there is a critical need to identify mechanisms regulating immune checkpoints in HNSCC such that one can predict who will benefit, and so novel combination strategies can be developed for non-responders. Here, we review the immunotherapy and molecular genetics literature to describe what is known about immune checkpoints in common genetic subsets of HNSCC. We highlight several highly recurrent genetic lesions that may serve as biomarkers or targets for combination immunotherapy in HNSCC.

Genomic sequencing and precision medicine in head and neck cancers.

Head and neck squamous cell carcinoma (HNSCC) remains a common and deadly disease. Historically, surgical and chemoradiation treatments have been met with modest success, and understanding of genetic drivers of HNSCC has been limited. With recent next generation sequencing studies focused on HNSCC, we are beginning to understand the genetic landscape of HNSCCs and are starting to identify and advance targeted options for patients. In this review, we describe current knowledge and recent advances in sequencing studies of HNSCC, discuss current limitations and future directions for further genomic analysis, and highlight the translational advances being undertaken to treat this important disease.

Supplementation of collagen scaffolds with SPARC to facilitate mineralization.

The extracellular matrix-associated protein, SPARC (Secreted Protein Acidic and Rich in Cysteine) is known to play a role in the mineralization of collagen in bone formation. The objectives of this study were to determine: 1) if SPARC supplementation of type 1 collagen scaffolds in vitro facilitated the binding of pre-formed HA nanoparticles added to the scaffolds; 2) if SPARC supplementation of the scaffolds enhanced the uptake of calcium and phosphorus from calcium phosphate solutions; and 3) if pretreatment in a calcium phosphate solution enhanced the subsequent binding of the nanoparticles. A related objective was to begin to determine the behavior of mesenchymal stem cells in the scaffolds when the constructs were grown in osteogenic medium. The calcium and phosphorus contents of the scaffolds were evaluated by inductively coupled plasma analysis, and the elastic modulus of the scaffolds determined by unconfined compression testing. Scaffolds were seeded with goat bone marrow-derived mesenchymal stem cells and the cell-seeded constructs grown in osteogenic medium. Supplementation of the scaffolds with as little as 0.008 % SPARC (by wt. of collagen) resulted in an increase in the binding of hydroxyapatite nanoparticles to the scaffold, but had no effect on incorporation of calcium or phosphorus from a calcium phosphate solution. The incorporation of hydroxyapatite nanoparticles into the scaffolds did not result in an increase in modulus. Supplementation of the scaffolds with SPARC and the increase in the binding of hydroxyapatite nanoparticles did not affect the proliferation of mesenchymal stem cells.

Injectable hydrogel materials for spinal cord regeneration: a review.

Spinal cord injury (SCI) presents a complex regenerative problem due to the multiple facets of growth inhibition that occur following trauma to the cord parenchyma and stroma. Clinically, SCI is further complicated by the heterogeneity in the size, shape and extent of human injuries. Many of these injuries do not breach the dura mater and have continuous viable axons through the injury site that can later lead to some degree of functional recovery. In these cases, surgical manipulation of the spinal cord by implanting a preformed scaffold or drug delivery device may lead to further damage. Given these circumstances, in situ-forming scaffolds are an attractive approach for SCI regeneration. These synthetic and natural polymers undergo a rapid transformation from liquid to gel upon injection into the cord tissue, conforming to the individual lesion site and directly integrating with the host tissue. Injectable materials can be formulated to have mechanical properties that closely match the native spinal cord extracellular matrix, and this may enhance axonal ingrowth. Such materials can also be loaded with cellular and molecular therapeutics to modulate the wound environment and enhance regeneration. This review will focus on the current status of in situ-forming materials for spinal cord repair. The advantages of, and requirements for, such polymers will be presented, and examples of the behavior of such systems in vitro and in vivo will be presented. There are helpful lessons to be learned from the investigations of injectable hydrogels for the treatment of SCI that apply to the use of these biomaterials for the treatment of lesions in other central nervous system tissues and in organs comprising other tissue types.

Correlation of radiographic and pathologic findings of dermal lymphatic invasion in head and neck squamous cell carcinoma.

HNSCC that involves the skin is able to invade the dermal lymphatic system. Currently there is no way to identify patients with dermal lymphatic invasion preoperatively. The purpose of this study is to determine whether CT can predict dermal lymphatic invasion. Medical records, CT scans, and corresponding histopathologic slides were reviewed of HNSCC patients with skin resected as part of their treatment. Dermal lymphatic invasion was defined radiographically as linear reticulations of the dermis and subcutaneous fat adjacent to the tumor. Twelve patients were identified with imaging suggestive of dermal lymphatic invasion. The corresponding pathology slides showed only 1 of the 12 patients had dermal lymphatic invasion, whereas the other 11 specimens showed peritumoral inflammation without evidence of tumor invasion. This study demonstrates that the linear areas of reticulation are most commonly caused by peritumoral inflammation and are not due to dermal lymphatic invasion.

Cell response to collagen-calcium phosphate cement scaffolds investigated for nonviral gene delivery.

Collagen-hydroxyapatite (HA) scaffolds for the non-viral delivery of a plasmid encoding the osteoinductive protein bone morphogenetic protein (BMP)-7 were developed. The collagen-HA was obtained by the combination of calcium phosphate cement in a collagen template. The effect on cell behavior of increasing amounts of HA in the scaffolds was evaluated. Collagen-HA scaffolds containing 13, 23 or 83 wt% HA were prepared. Cell proliferation was reduced in the 83% HA scaffold after 1 day compared to 13 and 23% HA, but by 14 days the number of cells in 83% HA considerably increased. Alkaline phosphatase (ALP) activity was 8 times higher for the 83% HA scaffolds. BMP-7 plasmid was incorporated into the 83% HA scaffold. The transfection was low, although significant levels of BMP7 were expressed, associated with an increase in cell proliferation.

The effects of glycosaminoglycan content on the compressive modulus of cartilage engineered in type II collagen scaffolds.

OBJECTIVE: The current study determined the unconfined compressive modulus of tissue-engineered constructs with varying sulfated glycosaminoglycan (GAG) density produced by goat articular chondrocytes in type II collagen scaffolds prepared with a range of cross-link densities and various times in culture. The purpose of this work is to establish a basis for future studies employing constructs of selected maturity (e.g., 25%, 50%, or 75% normal GAG content) for cartilage repair in vivo. METHODS: Porous scaffolds (8 mm diameter by 2 mm thick) were fabricated from porcine type II collagen by freeze-drying, followed by dehydrothermal treatment and carbodiimide cross-linking. In a pilot study, passage 3 adult caprine articular chondrocytes isolated from one goat were grown in scaffolds with six cross-link densities for 2, 3, 4, and 6 weeks (n=3). The goal was to select scaffold cross-link densities and times in culture that would produce constructs with approximately 25%, 50% and 75% the GAG density of native articular cartilage. Based on the results of the pilot study, chondrocytes from three goats were grown in scaffolds with two cross-link densities for three time periods: 3, 5, and 9 weeks (n=6; one of the cross-link groups was run in quadruplicate). The equilibrium modulus from unconfined compression testing of these samples was correlated with GAG content. RESULTS: There was a notable increase in GAG density with decreasing cross-link density. Histological analysis verified a chondrogenic phenotype and revealed various amounts of GAG and type II collagen-containing cartilage. The correlation between modulus and GAG density had a linear coefficient of determination of 0.60. One group with a mean GAG density of 22 microg/mm(3), which was 140% the GAG density of normal caprine articular cartilage, averaged a compressive modulus of 31.5 kPa, which was 10% of caprine articular cartilage tested in this study. CONCLUSIONS: The GAG density and modulus of tissue-engineered constructs can be controlled by the degree of cross-linking of type II collagen scaffolds and time in culture.

An integrated approach to distance learning with digital video in the French-speaking Virtual Medical University.

The aim of the French-speaking Virtual Medical University project (UMVF) is to share common resources and specific tools in order to improve medical training. Digital video on IP is an attractive tool for higher education but there are a number of obstacles to widespread implementation. This paper describes the UMVF approach to integrating digital video technologies and services in educational projects.

Collagen scaffolds for nonviral IGF-1 gene delivery in articular cartilage tissue engineering.

This study investigated the use of a type II collagen-glycosaminoglycan (CG) scaffold as a nonviral gene delivery vehicle for facilitating gene transfer to seeded adult articular chondrocytes to produce an elevated, prolonged and local expression of insulin-like growth factor (IGF)-1 for enhancing cartilage regeneration. Gene-supplemented CG (GSCG) scaffolds were synthesized by two methods: (1) soaking a pre-cross-linked CG scaffold in a plasmid solution followed by a freeze-drying process, and (2) chemically cross-linking the plasmid DNA to the scaffold. Two different plasmid solutions were also compared: (1) naked plasmid IGF-1 alone, and (2) plasmid IGF-1 with a lipid transfection reagent. Plasmid release studies revealed that cross-linking the plasmid to the CG scaffold prevented passive bolus release of plasmid and resulted in vector release controlled by scaffold degradation. In chondrocyte-seeded GSCG scaffolds, prolonged and elevated IGF-1 expression was enhanced by using the cross-linking method of plasmid incorporation along with the addition of the transfection reagent. The sustained level of IGF-1 overexpression resulted in significantly higher amounts of tissue formation, chondrocyte-like cells, GAG accumulation, and type II collagen production, compared to control scaffolds. These findings demonstrate that CG scaffolds can serve as nonviral gene delivery vehicles of microgram amounts of IGF-1 plasmid (<10 microg per scaffold) to provide a locally sustained therapeutic level of overexpressed IGF-1, resulting in enhanced cartilage formation.

Effect of expansion medium on ex vivo gene transfer and chondrogenesis in type II collagen-glycosaminoglycan scaffolds in vitro.

OBJECTIVE: Growth factors can profoundly affect the behaviour of chondrocytes during expansion and subsequent growth in three-dimensional (3-D) scaffolds. Prolonging such effects has stimulated investigation of the transfer of growth factor genes to chondrocytes. This study evaluated the effects of the monolayer expansion medium on the proliferation and cartilage matrix molecule synthesis of chondrocytes in 3-D pellet culture and in type II collagen-glycosaminoglycan (CG) scaffolds, and on ex vivo insulin-like growth factor-1 (IGF-1) gene transfer to articular chondrocytes in monolayer. The possibility of transfecting cells in 3-D culture using CG scaffolds was also investigated and the resulting effect of IGF-1 overexpression on glycosaminoglycan (GAG) biosynthesis in 3-D culture was assessed. METHODS: Two expansion media were compared-one that has been widely used for growing chondrocytes (Medium 1) and one that has been found to increase chondrocyte proliferation rates and preserve the redifferentiation potential of monolayer-expanded chondrocytes when subsequently placed in pellet cultures (Medium 2). Chondrocytes were expanded in monolayer culture and then 1) redifferentiated in 3-D culture, or 2) infected with the IGF-1 gene in monolayer or in type II CG scaffolds. RESULTS: The cell count for first passage chondrocytes was more than 3-fold higher when using Medium 2. In 3-D culture, cells expanded with Medium 2 and seeded in CG scaffolds produced more total GAG/DNA and displayed more intense immunohistochemical staining for collagen type II. Gene transfer and IGF-1 release kinetics from infected cells in monolayer were significantly affected by the composition of the expansion medium, the gene transfer method and time. IGF-1 gene transfer in CG scaffolds resulted in a 35-fold elevation in accumulated IGF-1 released from transfected Medium 2-expanded chondrocytes over controls, and resulted in a 40% increase in accumulated GAG/DNA. CONCLUSION: The composition of the expansion medium significantly affects monolayer proliferation of adult canine chondrocytes, GAG synthesis when the cells are subsequently grown in CG scaffolds, and ex vivo IGF-1 gene transfer.

Comparing ectopic bone growth induced by rhBMP-2 on an absorbable collagen sponge in rat and rabbit models.

Recombinant human Bone Morphogenetic Protein-2 (rhBMP-2) is currently employed as an autograft replacement for spinal fusion. The morphogen is incorporated onto its carrier, an absorbable collagen sponge (ACS), in the operating room. Although the effectiveness of the rhBMP-2/ACS implant in stimulating bone formation in human subjects has now been well established, further investigations of its use are necessary to deepen our understanding of its performance. The objective of the present study was to determine whether fluid released from the rhBMP-2/ACS implant could induce bone growth in tissue sites away from the implant site. We first measured the amount of protein in the fluid released from the rhBMP-2-soaked ACS during intraoperative handling. Variables included soak time and degree of compression. In the compression group that most closely approximated intraoperative conditions, more than 95% of the rhBMP-2 protein was retained by the ACS following a 15-min. soak time. This in vitro study was followed by an in vivo ectopic implant experiment using rat and rabbit models. The animal investigation compared the amount of bone induced by rhBMP-2 solution alone versus the de novo bone formation induced by rhBMP-2/ACS implants with varying concentrations of rhBMP-2. No ossicles were found at the sites where rhBMP-2 solution was injected in either animal species. Twenty-two of the 24 subcutaneous sites in the rats implanted with the rhBMP-2/ACS constructs displayed the presence of the typical 4- and 12-week ossicle. There were no noticeable differences in the size and shape of the ossicles after 4 and 12 weeks. There was a greater percentage of implant sites without ossicles in the rabbits, compared to the rats.

Optimizing a canine survival model of orthotopic lung transplantation.

INTRODUCTION: While acute models of orthotopic lung transplantation have been described in dogs, the technical considerations of developing a survival model in this species have not been elaborated. Herein, we describe optimization of a canine survival model of orthotopic lung transplantation. METHODS: Protocols of orthotopic left lung transplantation and single lung ventilation were established in acute experiments (n=9). Four dogs, serving as controls, received autologous, orthotopic lung transplants. Allogeneic transplants were performed in 16 DLA-identical and 16 DLA-mismatched unrelated recipient dogs. Selective right lung ventilation was utilized in all animals. A Malecot tube was left in the pleural space connected to a Heimlich valve for up to 24 hours. To date, animals have been followed up to 24 months by chest radiography, pulmonary function tests, bronchoscopy with lavage, and open biopsies. RESULTS: Long-term survival was achieved in 34/36 animals. Two recipients died intraoperatively secondary to cardiac arrest. All animals were extubated on the operating table, and in all cases the chest tube was removed within 24 hours. Major complications included thrombosis of the pulmonary artery and subcritical stenosis of bronchial anastamosis. One recipient underwent successful treatment of a small bowel intussusception. CONCLUSIONS: We report our experience in developing a survival canine model of orthotopic single lung transplantation. While short-term survival following canine lung transplantation is achievable, we report particular considerations that facilitate animal comfort, early extubation, and lung reexpansion in the immediate postoperative period, further optimizing use of this species for experimental modeling of long-term complications after lung transplantation.

Regeneration of articular cartilage.

Loss of articular cartilage from the ends of bones forming diarthrodial joints can be the source of profound pain and disability, and eventually lead to complete degeneration of the joint, necessitating total joint replacement. Until a few years ago, there seemed little hope of treating such defects. Novel surgical procedures and cell therapies have recently been found, however, to stimulate the formation of reparative tissue resulting in the relief of pain and restoration of function, at least for a limited time period. Moreover, studies of the healing of chondral defects in animal models have revealed that there is some potential for regeneration of this connective tissue. The introduction of certain biomaterial scaffolds along with selected surgical procedures and cell therapies has been demonstrated in animal studies to facilitate the cartilage reparative process and now offers the promise of extending the longevity of clinical treatments of cartilage defects. Collectively these findings provide the basis for the rational development of approaches for the more complete regeneration of articular cartilage, and demonstrate that meaningful clinical outcomes can be achieved even if complete regeneration is not achieved.

Hyaluronic acid-poly-D-lysine-based three-dimensional hydrogel for traumatic brain injury.

Brain tissue engineering in the postinjury brain represents a promising option for cellular replacement and rescue, providing a cell scaffold for either transplanted or resident cells. In this article, a hyaluronic acid (HA)-poly-D-lysine (PDL) copolymer hydrogel with an open porous structure and viscoelastic properties similar to neural tissue has been developed for brain tissue engineering. The chemicophysical properties of the hydrogel with HA:PDL ratios of 10:1, 5:1, and 4:1 were investigated by scanning electron microscopy (SEM) and X-ray photoelectron spectrometry. Neural cells cultured in the hydrogel were studied by phase-contrast microscope and SEM. The incorporation of PDL peptides into the HA-PDL hydrogel allowed for the modulation of neuronal cell adhesion and neural network formation. Macrophages and multinucleated foreign body giant cells found at the site of implantation of the hydrogel in the rat brain within the first weeks postimplantation decreased in numbers after 6 weeks, consistent with the host response to inert implants in numerous tissues. Of importance was the infiltration of the hydrogel by glial fibrillary acidic protein-positive cells-reactive astrocytes-by immunohistochemistry and the contiguity between the hydrogel and the surrounding tissue demonstrated by SEM. These findings indicated the compatibility of this hydrogel with brain tissue. Collectively, the results demonstrate the promise of an HA-PDL hydrogel as a scaffold material for the repair of defects in the brain.