Shear-Sensing by C-Reactive Protein: Linking Aortic Stenosis and Inflammation.

BACKGROUND: CRP (C-reactive protein) is a prototypical acute phase reactant. Upon dissociation of the pentameric isoform (pCRP [pentameric CRP]) into its monomeric subunits (mCRP [monomeric CRP]), it exhibits prothrombotic and proinflammatory activity. Pathophysiological shear rates as observed in aortic valve stenosis (AS) can influence protein conformation and function as observed with vWF (von Willebrand factor). Given the proinflammatory function of dissociated CRP and the important role of inflammation in the pathogenesis of AS, we investigated whether shear stress can modify CRP conformation and induce inflammatory effects relevant to AS. METHODS: To determine the effects of pathological shear rates on the function of human CRP, pCRP was subjected to pathophysiologically relevant shear rates and analyzed using biophysical and biochemical methods. To investigate the effect of shear on CRP conformation in vivo, we used a mouse model of arterial stenosis. Levels of mCRP and pCRP were measured in patients with severe AS pre- and post-transcatheter aortic valve implantation, and the presence of CRP was investigated on excised valves from patients undergoing aortic valve replacement surgery for severe AS. Microfluidic models of AS were then used to recapitulate the shear rates of patients with AS and to investigate this shear-dependent dissociation of pCRP and its inflammatory function. RESULTS: Exposed to high shear rates, pCRP dissociates into its proinflammatory monomers (mCRP) and aggregates into large particles. Our in vitro findings were further confirmed in a mouse carotid artery stenosis model, where the administration of human pCRP led to the deposition of mCRP poststenosis. Patients undergoing transcatheter aortic valve implantation demonstrated significantly higher mCRP bound to circulating microvesicles pre-transcatheter aortic valve implantation compared with post-transcatheter aortic valve implantation. Excised human stenotic aortic valves display mCRP deposition. pCRP dissociated in a microfluidic model of AS and induces endothelial cell activation as measured by increased ICAM-1 and P-selectin expression. mCRP also induces platelet activation and TGF-ß (transforming growth factor beta) expression on platelets. CONCLUSIONS: We identify a novel mechanism of shear-induced pCRP dissociation, which results in the activation of cells central to the development of AS. This novel mechanosensing mechanism of pCRP dissociation to mCRP is likely also relevant to other pathologies involving increased shear rates, such as in atherosclerotic and injured arteries.

Genetic Deficiency of TRAF5 Promotes Adipose Tissue Inflammation and Aggravates Diet-Induced Obesity in Mice.

Objective: The accumulation of inflammatory leukocytes is a prerequisite of adipose tissue inflammation during cardiometabolic disease. We previously reported that a genetic deficiency of the intracellular signaling adaptor TRAF5 (TNF [tumor necrosis factor] receptor-associated factor 5) accelerates atherosclerosis in mice by increasing inflammatory cell recruitment. Here, we tested the hypothesis that an impairment of TRAF5 signaling modulates adipose tissue inflammation and its metabolic complications in a model of diet-induced obesity in mice. Approach and Results: To induce diet-induced obesity and adipose tissue inflammation, wild-type or Traf5-/- mice consumed a high-fat diet for 18 weeks. Traf5-/- mice showed an increased weight gain, impaired insulin tolerance, and increased fasting blood glucose. Weight of livers and peripheral fat pads was increased in Traf5-/- mice, whereas lean tissue weight and growth were not affected. Flow cytometry of the stromal vascular fraction of visceral adipose tissue from Traf5-/- mice revealed an increase in cytotoxic T cells, CD11c+ macrophages, and increased gene expression of proinflammatory cytokines and chemokines. At the level of cell types, expression of TNF[alpha], MIP (macrophage inflammatory protein)-1[alpha], MCP (monocyte chemoattractant protein)-1, and RANTES (regulated on activation, normal T-cell expressed and secreted) was significantly upregulated in Traf5-deficient adipocytes but not in Traf5-deficient leukocytes from visceral adipose tissue. Finally, Traf5 expression was lower in adipocytes from obese patients and mice and recovered in adipose tissue of obese patients one year after bariatric surgery. Conclusions: We show that a genetic deficiency of TRAF5 in mice aggravates diet-induced obesity and its metabolic derangements by a proinflammatory response in adipocytes. Our data indicate that TRAF5 may promote anti-inflammatory and obesity-preventing signaling events in adipose tissue.

Age-related outcome of facial reanimation surgery using cross face nerve graft and gracilis free functional muscle transfer-A retrospective cohort study.

BACKGROUND: Free functional muscle transfer has become the criterion standard for the treatment of long-standing flaccid facial paralysis. Clinical experience suggests that a two-stage approach using a cross-face nerve graft (CFNG) as a donor nerve for free functional muscle transfers (FFMT) is less successful in older patients when compared to the pediatric population. However, clear data and scientific evidence are still rare. This study examines the age-related outcome of CFNG-driven FFMT. METHODS: Twenty-eight patients with a mean age of 20.73 years (ranging 5-51 years) who received two-stage facial reanimation with CFNG-driven gracilis FFMT at our institution from 1998 to 2019 were included. The ipsilateral sural nerve was used as CFNG. After 12 months, the ipsilateral gracilis muscle was used as FFMT. Patients were distributed equally into three cohorts according to their age. We assessed facial symmetry before and after facial reanimation measuring the angle between the interpupillary and the intermodiolar line (pupillo-modiolar angle). Additionally, the commissure height was measured using the Emotrics software. RESULTS: The mean follow-up of the pediatric, young adults and the middle-aged cohort was 29.5 ± 7.3, 24.9 ± 6.3, and 25.5 ± 12.4 months, respectively. One patient suffered flap loss due to flap ischemia. Four patients suffered insufficient innervation of the FFMT. Otherwise no major complication occurred. The likelihood of successful innervation of the FFMT was significantly higher in patients younger than 31 years (100% vs. 50%; p = .003). Smiling facial symmetry (pupillo-modiolar angle) significantly improved in the pediatric cohort (5-16 years; 8.68° ± 0.69° to 1.48° ± 0.67°; p < .001) and the young adults' cohort (17-30 years; 11.55° ± 1.95° to 4.62° ± 1.08°; p = .005), but improved only slightly in the middle-aged cohort (31-51 years; 11.77° ± 1.16° to 9.4° ± 1.8° p = .27). The postoperative smiling symmetry showed a significant correlation with increasing age (r = .62, p < .001). The smiling commissure height deviation significantly improved in the pediatric cohort (5-16 years; 6.5-2.3 mm; p = .006) and the postoperative result was significantly better than the middle-aged group (31-51 years; 2.3 vs. 7.5 mm; p = .02). CONCLUSIONS: The outcome of CFNG-driven gracilis FFMT is age-related. Static as well as dynamic facial symmetry after two-stage facial reanimation was best in the pediatric and young adult population. For older patients, other approaches like the nerve-to-masseter-driven FFMT should be considered.

Individualized Mini-Panel Sequencing of ctDNA Allows Tumor Monitoring in Complex Karyotype Sarcomas.

Soft tissue sarcomas (STS) are rare tumors of mesenchymal origin with high mortality. After curative resection, about one third of patients suffer from distant metastases. Tumor follow-up only covers a portion of recurrences and is associated with high cost and radiation burden. For metastasized STS, only limited inferences can be drawn from imaging data regarding therapy response. To date there are no established and evidence-based diagnostic biomarkers for STS due to their rarity and diversity. In a proof-of-concept study, circulating tumor DNA (ctDNA) was quantified in (n = 25) plasma samples obtained from (n = 3) patients with complex karyotype STS collected over three years. Genotyping of tumor tissue was performed by exome sequencing. Patient-individual mini-panels for targeted next-generation sequencing were designed encompassing up to 30 mutated regions of interest. Circulating free DNA (cfDNA) was purified from plasma and ctDNA quantified therein. ctDNA values were correlated with clinical parameters. ctDNA concentrations correlated with the tumor burden. In case of full remission, no ctDNA was detectable. Patients with a recurrence at a later stage showed low levels of ctDNA during clinical remission, indicating minimal residual disease. In active disease (primary tumor or metastatic disease), ctDNA was highly elevated. We observed direct response to treatment, with a ctDNA decline after tumor resections, radiotherapy, and chemotherapy. Quantification of ctDNA allows for the early detection of recurrence or metastases and can be used to monitor treatment response in STS. Therapeutic decisions can be made earlier, such as the continuation of a targeted adjuvant therapy or the implementation of extended imaging to detect recurrences. In metastatic disease, therapy can be adjusted promptly in case of no response. These advantages may lead to a survival benefit for patients in the future.

C-Reactive Protein and Its Structural Isoforms: An Evolutionary Conserved Marker and Central Player in Inflammatory Diseases and Beyond.

C-reactive protein (CRP) is an evolutionary highly conserved member of the pentraxin superfamily of proteins. CRP is widely used as a marker of inflammation, infection and for risk stratification of cardiovascular events. However, there is now a large body of evidence, that continues to evolve, detailing that CRP directly mediates inflammatory reactions and the innate immune response in the context of localised tissue injury. These data support the concept that the pentameric conformation of CRP dissociates into pro-inflammatory CRP isoforms termed pCRP* and monomeric CRP. These pro-inflammatory CRP isoforms undergo conformational changes that facilitate complement binding and immune cell activation and therefore demonstrate the ability to trigger complement activation, activate platelets, monocytes and endothelial cells. The dissociation of pCRP occurs on the surface of necrotic, apoptotic, and ischaemic cells, regular ß-sheet structures such as ß-amyloid, the membranes of activated cells (e.g., platelets, monocytes, and endothelial cells), and/or the surface of microparticles, the latter by binding to phosphocholine. Therefore, the deposition and localisation of these pro-inflammatory isoforms of CRP have been demonstrated to amplify inflammation and tissue damage in a broad range of clinical conditions including ischaemia/reperfusion injury, Alzheimer's disease, age-related macular degeneration and immune thrombocytopaenia. Given the potentially broad relevance of CRP to disease pathology, the development of inhibitors of CRP remains an area of active investigation, which may pave the way for novel therapeutics for a diverse range of inflammatory diseases.

Genotyping of circulating cell-free DNA enables noninvasive tumor detection in myxoid liposarcomas.

Soft tissue sarcomas (STS) are rare tumors of mesenchymal origin. About 50% of patients with STS experience relapse and more than 30% will die within 10 years after diagnosis. In this study we investigated circulating free DNA (cfDNA) and tumor-specific genetic alterations therein (circulating tumor DNA, ctDNA) as diagnostic biomarkers. Plasma concentrations and fragmentation of cfDNA was analyzed with quantitative PCR. Patients with STS (n = 64) had significantly higher plasma concentrations and increased fragmentation of cfDNA when compared to patients in complete remission (n = 19) and healthy controls (n = 41) (p < 0.01 and p < 0.001). Due to overlapping values between patients with STS and controls, the sensitivity and specificity of these assays is limited. Sensitive assays to detect genomic alterations in cfDNA of synovial sarcomas (t(X;18)), myxoid liposarcomas (t(12;16) and TERT C228T promoter mutation) and well-differentiated/de-differentiated liposarcomas (MDM2 amplifications) were established. ctDNA was quantified in nine liposarcoma patients during the course of their treatment. Levels of breakpoint t(12;16) and TERT C228T ctDNA correlated with the clinical course and tumor burden in patients with myxoid liposarcomas (n = 4). ctDNA could detect minimal residual disease and tumor recurrence. In contrast, detection of MDM2 amplifications was not sensitive enough to detect tumors in patients with well-differentiated/de-differentiated liposarcomas (n = 5). Genotyping of cfDNA for tumor specific genetic alterations is a feasible and promising approach for monitoring tumor activity in patients with myxoid liposarcomas. Detection of ctDNA during follow-up examinations despite negative standard imaging studies might warrant more sensitive imaging (e.g. PET-CT) or closer follow-up intervals to timely localize and treat recurrences.

Comparison of symmetry after smile reconstruction for flaccid facial paralysis with combined fascia lata grafts and functional gracilis transfer for static suspension or gracilis transfer alone.

PURPOSE: Facial paralysis has a profound impact on functionality and esthetics of the oral region. In patients with strong skin laxity and soft tissue ptosis, functional smile reconstruction is challenging due to the accentuated asymmetry at rest. Thus, the purpose of the study was to analyze facial symmetry in this patient clientele following a combination of dynamic reanimation with fascial strips for static suspension compared to functional gracilis transfer alone. METHODS: In 2014, we altered the single-stage approach for microsurgical smile reconstruction in patients with significant soft tissue ptosis by adding fascia lata grafts for static support. We evaluated 6 patients (mean age 57.8 ± 5.2, group A) who underwent the combined procedure, and compared their results to 6 patients with flaccid facial paralysis who were treated before 2014 and received a functional gracilis transfer alone (mean age 52.5 ± 7.5, group B). To test the efficacy of the technique, we retrospectively analyzed the correction of the oral asymmetry as well as nasal and philtral deviation by computer-assisted photograph analysis 6 months postoperatively. RESULTS: The comparative analysis revealed a significant postoperative improvement of the oral asymmetry (A: 90.0 ± 5.0% relative correction at rest vs. B: 62.6 ± 17.2%, P < .05), nasal (A: 0.4 ± 0.2 vs. B: 0.7 ± 0.4 mm, P < .05), and philtral deviation (A: 0.5 ± 0.6 vs. B: 2.8 ± 1.8 mm, P < .05) in group A. CONCLUSIONS: The combined procedure for dynamic facial reanimation allows for immediate correction of the oral asymmetry and improves overall outcome in patients with advanced soft tissue ptosis and oral asymmetry at rest.

Evaluation of the Implantable Doppler Probe for Free Flap Monitoring in Lower Limb Reconstruction.

BACKGROUND: Timely reexploration and reanastomoses can salvage failing free flaps. The use of the implantable Doppler probe provides direct evidence of vascular impairment of the microvascular anastomoses and allows for postoperative NPWT. The aim of this retrospective study was to compare the Doppler probe to conventional monitoring techniques for free flap monitoring in lower limb reconstruction and to identify risk factors for perfusion disturbance and reexploration. METHODS: All patients receiving free muscle flap reconstruction for lower limb soft tissue defects at our department from 2000 to 2013 were included, and all adverse events, timely detection of perfusion problems, and outcome of revision surgery were assessed by chart analysis. RESULTS: For lower limb reconstruction, 110 free muscle transfers were performed of which 41 muscle flaps were conventionally monitored and 69 flaps were monitored using the implantable Doppler probe. In 18 cases, the free muscle flaps needed revision because of perfusion disturbances. The salvage rate was 80% with monitoring by the implantable Doppler probe compared with 62.5% using conventional monitoring methods resulting in success rates of 95.7 and 92.7%, respectively. CONCLUSION: The use of the implantable Cook-Swartz Doppler probe represents a safe monitoring method for lower limb reconstruction, which allows for the additional use of NPWT. Higher salvage and revision success rates can be attributed to an earlier detection of perfusion impairment. However, a larger patient cohort is necessary to verify superiority over conventional postoperative monitoring.

Metabolic Response to XD14 Treatment in Human Breast Cancer Cell Line MCF-7.

XD14 is a 4-acyl pyrrole derivative, which was discovered by a high-throughput virtual screening experiment. XD14 inhibits bromodomain and extra-terminal domain (BET) proteins (BRD2, BRD3, BRD4 and BRDT) and consequently suppresses cell proliferation. In this study, metabolic profiling reveals the molecular effects in the human breast cancer cell line MCF-7 (Michigan Cancer Foundation-7) treated by XD14. A three-day time series experiment with two concentrations of XD14 was performed. Gas chromatography-mass spectrometry (GC-MS) was applied for untargeted profiling of treated and non-treated MCF-7 cells. The gained data sets were evaluated by several statistical methods: analysis of variance (ANOVA), clustering analysis, principle component analysis (PCA), and partial least squares discriminant analysis (PLS-DA). Cell proliferation was strongly inhibited by treatment with 50 µM XD14. Samples could be discriminated by time and XD14 concentration using PLS-DA. From the 117 identified metabolites, 67 were significantly altered after XD14 treatment. These metabolites include amino acids, fatty acids, Krebs cycle and glycolysis intermediates, as well as compounds of purine and pyrimidine metabolism. This massive intervention in energy metabolism and the lack of available nucleotides could explain the decreased proliferation rate of the cancer cells.

Dissociation of pentameric to monomeric C-reactive protein localizes and aggravates inflammation: in vivo proof of a powerful proinflammatory mechanism and a new anti-inflammatory strategy.

BACKGROUND: The relevance of the dissociation of circulating pentameric C-reactive protein (pCRP) to its monomeric subunits (mCRP) is poorly understood. We investigated the role of conformational C-reactive protein changes in vivo. METHODS AND RESULTS: We identified mCRP in inflamed human striated muscle, human atherosclerotic plaque, and infarcted myocardium (rat and human) and its colocalization with inflammatory cells, which suggests a general causal role of mCRP in inflammation. This was confirmed in rat intravital microscopy of lipopolysaccharide-induced cremasteric muscle inflammation. Intravenous pCRP administration significantly enhanced leukocyte rolling, adhesion, and transmigration via localized dissociation to mCRP in inflamed but not noninflamed cremaster muscle. This was confirmed in a rat model of myocardial infarction. Mechanistically, this process was dependent on exposure of lysophosphatidylcholine on activated cell membranes, which is generated after phospholipase A2 activation. These membrane changes could be visualized intravitally on endothelial cells, as could the colocalized mCRP generation. Blocking of phospholipase A2 abrogated C-reactive protein dissociation and thereby blunted the proinflammatory effects of C-reactive protein. Identifying the dissociation process as a therapeutic target, we stabilized pCRP using 1,6-bis(phosphocholine)-hexane, which prevented dissociation in vitro and in vivo and consequently inhibited the generation and proinflammatory activity of mCRP; notably, it also inhibited mCRP deposition and inflammation in rat myocardial infarction. CONCLUSIONS: These results provide in vivo evidence for a novel mechanism that localizes and aggravates inflammation via phospholipase A2-dependent dissociation of circulating pCRP to mCRP. mCRP is proposed as a pathogenic factor in atherosclerosis and myocardial infarction. Most importantly, the inhibition of pCRP dissociation represents a promising, novel anti-inflammatory therapeutic strategy.

Identification of a blood-borne miRNA signature of synovial sarcoma.

BACKGROUND: Synovial sarcoma account for approximately 10 % of all soft-tissue tumors and occur most frequently in young adults. A specific translocation in this sarcoma induces fusion of the SYT gene on chromosome 18 to the SSX genes on chromosome X, leading to proliferation of the tumor cells. The need for non-invasive biomarkers indicating recurrence and activity of this disease has sparked research into short non-coding RNA known as microRNA (miRNA). METHODS: Blood samples of patients with active synovial sarcoma and of synovial sarcoma patients in complete remission as well as of healthy donors and patients with active leiomyosarcoma, MPNST, Ewing sarcoma and liposarcoma were collected. Whole blood RNA was extracted and samples of patients with active synovial sarcoma and of healthy donors were analyzed using an Affymetrix GeneChip miRNA Array v. 4.0. qRT-PCR was carried out to confirm a panel of miRNAs which where differentially expressed in the miRNA array. This miRNA-panel was further evaluated in patients with synovial sarcoma in complete remission and patients with active leiomyosarcoma, MPNST, Ewing sarcoma and liposarcoma as well as in an independent cohort of synovial sarcoma patients. RESULTS: Unsupervised hierarchical clustering of the miRNA arrays separated patients with active synovial sarcoma from healthy controls. A panel of seven miRNAs (miR-99a-5p, miR-146b-5p, miR-148b-3p, miR-195-5p, miR-223-3p, miR-500b-3p and miR-505-3p) was further validated by qRT-PCR to be significantly upregulated in synovial sarcoma patients. Moreover, most of the analyzed miRNAs were shown to be significantly upregulated in synovial sarcoma patients compared to leiomyosarcoma, MPNST, Ewing sarcoma and liposarcoma patients. Validation of the miRNA panel in an independent cohort of synovial sarcoma patients confirmed higher expression levels compared to healthy controls and patients in complete remission. CONCLUSION: Our results have identified a specific whole blood miRNA signature that may serve as an independent biomarker for the diagnosis of local recurrence or distant metastasis of synovial sarcoma. It even distinguishes synovial sarcoma from other sarcoma subtypes, thus potentially serving as a specific biomarker for synovial sarcoma.

MicroRNA-155 aggravates ischemia-reperfusion injury by modulation of inflammatory cell recruitment and the respiratory oxidative burst.

The inflammatory sequelae of ischemia-reperfusion injury (IRI) are a major causal factor of tissue injury in various clinical settings. MicroRNAs (miRs) are short, non-coding RNAs, which regulate protein expression. Here, we investigated the role of miR-155 in IR-related tissue injury. Quantifying microRNA-expression levels in a human muscle tissue after IRI, we found miR-155 expression to be significantly increased and to correlate with the increased expression of TNF-α, IL-1ß, CD105, and Caspase3 as well as with leukocyte infiltration. The direct miR-155 target gene SOCS-1 was downregulated. In a mouse model of myocardial infarction, temporary LAD ligation and reperfusion injury resulted in a smaller area of necrosis in miR-155-/- animals compared to wildtype animals. To investigate the underlying mechanisms, we evaluated the effect of miR-155 on inflammatory cell recruitment by intravital microscopy and on the generation of reactive oxygen species (ROS) of macrophages. Our intravital imaging results demonstrated a decreased recruitment of inflammatory cells in miR-155-/- animals during IRI. The generation of ROS in leukocytic cells of miR-155-/- animals was also reduced. RNA silencing of the direct miR-155 target gene SOCS-1 abrogated this effect. In conclusion, miR-155 aggravates the inflammatory response, leukocyte infiltration and tissue damage in IRI via modulation of SOCS-1-dependent generation of ROS. MiR-155 is thus a potential target for the treatment or prevention of IRI.

A conformational change of C-reactive protein in burn wounds unmasks its proinflammatory properties.

Tissue damage in burn injury leads to a rapid increase of leukocytes and acute phase reactants. Plasma levels of C-reactive protein (CRP) rise within hours after the insult. No deficiency of this protein has been reported in humans, suggesting it plays a pivotal role in innate immunity. CRP in circulation is composed of five identical subunits [pentameric CRP (pCRP)]. Recently, deposits of structurally modified CRP (mCRP) have been found in inflammatory diseases. Little is known about this structural change and how it affects CRP functions. We analyzed CRP deposits in burn wounds and serum by immunohistochemistry, western blot and dot blot analysis. CRP was deposited in necrotic and inflamed tissue, but not in adjacent healthy tissue. Tissue deposited CRP was detected by mCRP-specific antibodies and structurally different from serum pCRP. mCRP but not pCRP induced reactive oxygen species production by monocytes and facilitated uptake of necrotic Jurkat cells by macrophages. In addition, it accelerated migration of keratinocytes in a scratch wound assay. The structural changes that occur in pCRP upon localization to damaged and inflamed tissue in burn wounds result in a functionally altered protein with distinct functions. mCRP exhibits opsonic, proinflammatory and promigratory properties which modulate wound healing.

C-reactive protein orchestrates acute allograft rejection in vascularized composite allotransplantation via selective activation of monocyte subsets.

INTRODUCTION: Despite advancements in transplant immunology and vascularized composite allotransplantation (VCA), the longevity of allografts remains hindered by the challenge of allograft rejection. The acute-phase response, an immune-inflammatory reaction to ischemia/reperfusion that occurs directly after allogeneic transplantation, serves as a catalyst for graft rejection. This immune response is orchestrated by acute-phase reactants through intricate crosstalk with the mononuclear phagocyte system. OBJECTIVE: C-reactive protein (CRP), a well-known marker of inflammation, possesses pro-inflammatory properties and exacerbates ischemia/reperfusion injury. Thus, we investigated how CRP impacts acute allograft rejection. METHODS: Prompted by clinical observations in facial VCAs, we employed a complex hindlimb transplantation model in rats to investigate the direct impact of CRP on transplant rejection. RESULTS: Our findings demonstrate that CRP expedites allograft rejection and diminishes allograft survival by selectively activating non-classical monocytes. Therapeutic stabilization of CRP abrogates this activating effect on monocytes, thereby attenuating acute allograft rejection. Intravital imagining of graft-infiltrating, recipient-derived monocytes during the early phase of acute rejection corroborated their differential regulation by CRP and their pivotal role in driving the initial stages of graft rejection. CONCLUSION: The differential activation of recipient-derived monocytes by CRP exacerbates the innate immune response and accelerates clinical allograft rejection. Thus, therapeutic targeting of CRP represents a novel and promising strategy for preventing acute allograft rejection and potentially mitigating chronic allograft rejection.

Special Issue: Evidence-Based Diagnosis and Management of Facial Nerve Disorders.

Although there has been a rapid increase in the number of new publications and studies in relation to the diagnostics, impacts and rehabilitation methods of facial nerve disorders, a general structure in evidence-based medicine is still difficult to establish [...].

Functional Outcome of Dual Reinnervation with Cross-Facial Nerve Graft and Masseteric Nerve Transfer for Facial Paralysis.

BACKGROUND: The combination of cross-facial nerve graft (CFNG) and masseteric nerve transfer (MNT) for reinnervation of facial paralysis may provide advantages of both neural sources. However, quantitative functional outcome reports with a larger number of patients are lacking in the literature. Here we describe our 8-year experience with this surgical technique. METHODS: Twenty patients that presented with a complete facial paralysis (duration <12 months) received dual reinnervation with CFNG and MNT. The functional outcome of the procedure was evaluated with the physician-graded outcome metric eFACE. The objective artificial intelligence-driven software Emotrics and FaceReader were used for oral commissure measurements and emotional expression assessment, respectively. RESULTS: The mean follow-up was 31.75±23.32 months. In the eFACE score, the nasolabial fold depth and oral commissure at rest improved significantly (p<0.05) towards a more balanced state after the surgery. Postoperatively, there was a significant decrease in oral commissure asymmetry while smiling (19.22±6.1mm to 12.19±7.52mm). For the emotionality expression, the median intensity score of happiness, as measured by the FaceReader software, increased significantly while smiling (0.28, IQR 0.13-0.64). In 5 (25%) patients, a secondary static midface suspension with fascia lata strip had to be performed due to an unsatisfactory resting symmetry. Older patients and patients with greater preoperative resting asymmetry were more likely to receive static midface suspension. CONCLUSIONS: Our results suggest that the combination of MNT and CFNG for reinnervation of facial paralysis provides good voluntary motion and may lessen the use of static midface suspension in the majority of patients.

Age-related outcomes of facial reanimation surgery using gracilis free functional muscle transfer innervated by the masseteric nerve: A retrospective cohort study.

BACKGROUND: The free functional muscle gracilis transfer is an established approach in facial reanimation surgery; however, the significance of its neurotization and the patient's age is still inconclusive. Several donor nerves are available for facial reanimation using the free functional gracilis muscle transfer. OBJECTIVE: This retrospective cohort study investigates whether the masseteric nerve is an equally reliable donor nerve in both older and younger patients. METHODS: We included 46 patients (13-71 years, male and female) who underwent nerve-to-masseter (NTM)-driven free functional muscle transfer (FFMT) between January 2008 and December 2019. Patients were distributed into three cohorts according to their age at surgery. We assessed the facial symmetry before and after surgery using the pupillo-modiolar angle. Commissure height and excursion deviation were measured with the Emotrics software. Patient-reported outcome measurements were taken using the Facial Clinimetric Examination (FaCE) scale. RESULTS: All patients had successful flap innervation, except for one patient in the middle-aged cohort (31-51 years). The postoperative facial symmetry at rest, smiling, and laughing was analyzed with the pupillo-modiolar angle and the Emotrics software and showed similar results between all cohorts. The FaCE scale showed similar scores for the middle-aged (31-51 years) cohort and the senior cohort (52-71 years). The social function score in the senior cohort was higher than in the middle-aged cohort, without statistical significance. One patient in the middle-aged (31-51 years) cohort and the senior cohort (52-71 years), respectively, underwent emergency revision due to impaired flap perfusion and could be salvaged. CONCLUSIONS: NTM-driven FFMT for facial reanimation is a safe and reliable procedure across all age groups of patients.

Secondary dynamic midface reanimation with gracilis free muscle transfer after failed reconstruction attempt: A 15-year experience.

BACKGROUND: The quantitative outcome of secondary reanimation after a failed primary reconstruction attempt for facial paralysis is rarely reported in the literature. This study aimed to investigate the feasibility of secondary reanimation with gracilis free muscle transfer (GFMT) and whether this outcome is influenced by the primary reconstruction. METHODS: Twelve patients with previously failed static procedures (static group, n = 6), temporal muscle transfer (temporal transfer group, n = 2), and GFMT (GFMT group, n = 4) were all secondarily reanimated with GFMT. The clinical outcome was graded with the eFACE metric. The objective oral commissure excursion was measured with Emotrics, and the artificial intelligence software FaceReader evaluated the intensity score (IS) of emotional expression. RESULTS: The mean follow-up was 40 ± 27 months. The eFACE metric showed a statistically significant (p < 0.05) postoperative improvement in the dynamic and smile scores across all groups. In the GFMT group, oral commissure with smile (75.75 ± 20.43 points), oral commissure excursion while smiling with teeth showing (32.7 ± 4.35 mm), and the intensity of happiness emotion while smiling without teeth showing (IS of 0.37 ± 0.23) were significantly lower as compared with the static group postoperatively (98.83 ± 2.86 points, p = 0.038; 41.7 ± 4.35 mm, p = 0.025; IS 0.83 ± 0.16, p = 0.01). CONCLUSIONS: Our data suggest that secondary dynamic reconstruction with GFMT is feasible should the primary reconstruction fail. The secondary GFMT appears to improve the outcome of primary GFMT; however, the oral commissure excursion while smiling might be lower than that in patients who had static procedures as primary reconstruction.

Dermatofibrosarcoma protuberans of the scalp: Surgical management in a multicentric series of 11 cases and systematic review of the literature.

BACKGROUND: Dermatofibrosarcoma protuberans is a rare, slow-growing soft-tissue malignancy originating in the dermis that is characterized by an infiltrating growth pattern with a marked tendency of local recurrence. Complete surgical resection with pathological margin clearance must be achieved to reduce the risk of tumor recurrence. Resulting defects often require extensive reconstructive procedures. Dermatofibrosarcoma protuberans of the scalp poses particular challenges owing to the proximity to the face and brain. This study aims to evaluate treatment options and proposes an algorithm for management of scalp dermatofibrosarcoma protuberans based on a multicentric case series and systematic review of the literature. METHODS: A retrospective multicentric chart analysis of 11 patients with scalp dermatofibrosarcoma protuberans who presented within the last 20 years was performed regarding demographic data, pathological tumor characteristics, and surgical management (resection and reconstruction). Additionally, a further 42 patients (44 cases) were identified through a systematic Preferred Reporting Systems for Systematic Reviews and Meta-Analysis-based review of the literature searching the Medline and Embase databases. RESULTS: In total, 30 cases were classified as primary and 20 cases as recurring scalp dermatofibrosarcoma protuberans (data from 5 cases were missing). The median tumor size was 24 cm2 (interquartile range 7.8-64), and the median defect size was 55.8 cm2 (interquartile range 48-112). Recurring scalp dermatofibrosarcoma protuberans was more often associated with invasion of deeper layers and required more extensive tumor resection to achieve negative margins. Within the subgroup that was managed with peripheral and deep en face margin assessment, no recurrence was observed. Most patients required local (41. 8%) or free flap (27.8%) reconstruction after dermatofibrosarcoma protuberans resection. CONCLUSION: Whenever possible, peripheral and deep en face margin assessment-based techniques should be preferred for resection of scalp dermatofibrosarcoma protuberans because they provide superior oncological safety while preserving uninvolved tissue. Patients with locally advanced and recurring scalp dermatofibrosarcoma protuberans often require multidisciplinary treatment including neurosurgery, radiotherapy, and microvascular reconstructive surgery and should be referred to a specialized center.

Multiparametric magnetic resonance imaging for radiation therapy response monitoring in soft tissue sarcomas: a histology and MRI co-registration algorithm.

Rationale: To establish a spatially exact co-registration procedure between in vivo multiparametric magnetic resonance imaging (mpMRI) and (immuno)histopathology of soft tissue sarcomas (STS) to identify imaging parameters that reflect radiation therapy response of STS. Methods: The mpMRI-Protocol included diffusion-weighted (DWI), intravoxel-incoherent motion (IVIM), and dynamic contrast-enhancing (DCE) imaging. The resection specimen was embedded in 6.5% agarose after initial fixation in formalin. To ensure identical alignment of histopathological sectioning and in vivo imaging, an ex vivo MRI scan of the specimen was rigidly co-registered with the in vivo mpMRI. The deviating angulation of the specimen to the in vivo location of the tumor was determined. The agarose block was trimmed accordingly. A second ex vivo MRI in a dedicated localizer with a 4 mm grid was performed, which was matched to a custom-built sectioning machine. Microtomy sections were stained with hematoxylin and eosin. Immunohistochemical staining was performed with anti-ALDH1A1 antibodies as a radioresistance and anti-MIB1 antibodies as a proliferation marker. Fusion of the digitized microtomy sections with the in vivo mpMRI was accomplished through nonrigid co-registration to the in vivo mpMRI. Co-registration accuracy was qualitatively assessed by visual assessment and quantitatively evaluated by computing target registration errors (TRE). Results: The study sample comprised nine tumor sections from three STS patients. Visual assessment after nonrigid co-registration showed a strong morphological correlation of the histopathological specimens with ex vivo MRI and in vivo mpMRI after neoadjuvant radiation therapy. Quantitative assessment of the co-registration procedure using TRE analysis of different pairs of pathology and MRI sections revealed highly accurate structural alignment, with a total median TRE of 2.25 mm (histology - ex vivo MRI), 2.22 mm (histology - in vivo mpMRI), and 2.02 mm (ex vivo MRI - in vivo mpMRI). There was no significant difference between TREs of the different pairs of sections or caudal, middle, and cranial tumor parts, respectively. Conclusion: Our initial results show a promising approach to obtaining accurate co-registration between histopathology and in vivo MRI for STS. In a larger cohort of patients, the method established here will enable the prospective identification and validation of in vivo imaging biomarkers for radiation therapy response prediction and monitoring in STS patients via precise molecular and cellular correlation.