Non-Union Treatment in the Foot, Ankle, and Lower Leg: A Multicenter Retrospective Study Comparing Conventional Treatment with the Human Allogeneic Cortical Bone Screw (Shark Screw®).

Addressing non-unions involves stabilizing the affected area through osteosynthesis and improving bone biology using bone grafts. However, there is no consensus on the optimal treatment method. This study aims to compare outcomes of non-union surgery using conventional treatment methods (metal hardware ± graft) versus osteosynthesis with the human allogeneic cortical bone screw (Shark Screw®) alone or in combination with a metallic plate. Thirty-four patients underwent conventional treatment, while twenty-eight cases received one or more Shark Screws®. Patient demographics, bone healing, time to bone healing, and complications were assessed. Results revealed a healing rate of 96.4% for the Shark Screw® group, compared to 82.3% for the conventionally treated group. The Shark Screw® group exhibited a tendency for faster bone healing (9.4 ± 3.2 vs. 12.9 ± 8.5 weeks, p = 0.05061). Hardware irritations led to six metal removals in the conventional group versus two in the Shark Screw® group. The Shark Screw® emerges as a promising option for personalized non-union treatment in the foot, ankle, and select lower leg cases, facilitating effective osteosynthesis and grafting within a single construct and promoting high union rates, low complications, and a rapid healing process.

Structural and biochemical characterization of human Schlafen 5.

The Schlafen family belongs to the interferon-stimulated genes and its members are involved in cell cycle regulation, T cell quiescence, inhibition of viral replication, DNA-repair and tRNA processing. Here, we present the cryo-EM structure of full-length human Schlafen 5 (SLFN5) and the high-resolution crystal structure of the highly conserved N-terminal core domain. We show that the core domain does not resemble an ATPase-like fold and neither binds nor hydrolyzes ATP. SLFN5 binds tRNA as well as single- and double-stranded DNA, suggesting a potential role in transcriptional regulation. Unlike rat Slfn13 or human SLFN11, human SLFN5 did not cleave tRNA. Based on the structure, we identified two residues in proximity to the zinc finger motif that decreased DNA binding when mutated. These results indicate that Schlafen proteins have divergent enzymatic functions and provide a structural platform for future biochemical and genetic studies.

Efficient treatment of murine acute GvHD by in vitro expanded donor regulatory T cells.

Acute graft-versus-host disease (aGvHD) is a frequent complication after allogeneic bone marrow/stem cell transplantation (BMT/SCT) induced by co-transplanted alloreactive conventional donor T cells. We previously demonstrated that the adoptive transfer of donor CD4+CD25+Foxp3+ regulatory T cells (Treg) at the time of BMT prevents aGvHD in murine models. Yet, the therapeutic potential of donor Treg for the treatment of established aGvHD has not yet been studied in detail. We now used in vitro expanded phenotypically and functionally stable murine Treg to explore their therapeutic efficacy in haploidentical aGvHD models. Upon transfer donor Treg ameliorate clinical and histologic signs of aGvHD and significantly improve survival. They migrate to lymphoid as well as aGvHD target organs, predominantly the gastrointestinal tract, where they inhibit the proliferation of conventional T cells, reduce the influx of myeloid cells, and the accumulation of inflammatory cytokines. Successfully treated animals restore aGvHD-induced tissue damage in target organs and lymphoid tissues, thereby supporting lymphocyte reconstitution. The therapeutically applied Treg population survives long term without conversion into pathogenic effector T cells. These results demonstrate that donor Treg not only prevent aGvHD, but are also efficacious for the treatment of this life-threatening BMT complication.

Lost in Datafication? - A Typology of (Emotion) Data Contextualization.

This article elaborates on the meaning of "context" for data created in interdisciplinary research on emotions. Particularly with regard to the potential reuse of scientific data, the elicitation of contexts can contribute to a better assessment of emotion data. Beyond a discussion of social scientific conceptualizations of "context" focusing on the situational and cultural contexts and their respective interrelations, this article presents the findings of an empirical study on datafication processes in interdisciplinary emotion research. Based on 123 survey responses and 15 in-depth interviews, a multitude of contextual dimensions will be reviewed. The typology of contexts, ranging from method-specific aspects and researchers' subjectivities to the contextual embeddedness of the research objects, provides a schema suitable for various epistemological approaches. The proposed typology can serve as a framework for emotion researchers to reflect on their research practice and interactions with research participants. The empirical findings also show the limitations of contextualization pertaining to tacit knowledge, implicit knowledge, embodied emotions and ethical considerations. The article concludes with suggestions for further research, pointing to intercultural settings, the integration of contexts and particular scenarios for data reuse.

The association between acute graft-versus-host disease and antimicrobial peptide expression in the gastrointestinal tract after allogeneic stem cell transplantation.

Intestinal microbiota disruption is associated with acute gastrointestinal (GI) Graft-versus-Host Disease (GvHD) and poor outcome after allogeneic stem cell transplantation (ASCT). Here, in a retrospective analysis of 200 patients undergoing ASCT at the Regensburg University Medical Center, we assessed the relative expression of Paneth cell antimicrobial peptides (AMPs), Human Defensins (HD) 5 and 6 and regenerating islet-derived 3α (Reg3α), in 292 human intestinal biopsies as well as Reg3α serum levels in relation to acute GI GvHD. In the absence of GI GvHD, the relative expression of Paneth cell AMPs was significantly higher in the small intestine (duodenum to ileum) than in the stomach and large intestine (cecum to rectum) for Reg3α (p≤0.001), HD5 (p≤0.002) and HD6 (p≤0.02). Acute stage 2-4 GI GvHD was associated with reduced expression of AMPs in the small intestine (p≤0.01) in comparison to stage 0-1 disease, accompanied by a decrease in Paneth cell count in case of severe acute GI GvHD (p<0.001). The opposite held true for the large intestine as we found stage 2-4 GI GvHD correlated with significantly higher expression of HD5, HD6, and Reg3α compared to mild or no acute GI GvHD (p≤0.002). Severe GI GvHD in both the lower and the upper GI tract also correlated with higher serum concentrations of Reg3α (p = 0.002). As indirect markers of intestinal microbiome diversity low levels of urinary 3-indoxyl sulfate levels were associated with severe stages of acute GI GvHD compared to mild stage or no acute GI GvHD (p = 0.05). In conclusion, acute GI GvHD correlates with intestinal expression of HD5, HD6 and Reg3α as well as Reg3α serum levels and is associated with intestinal dysbiosis.

* Demineralized Bone Matrix as a Carrier for Bone Morphogenetic Protein-2: Burst Release Combined with Long-Term Binding and Osteoinductive Activity Evaluated In Vitro and In Vivo.

To allow bone defect regeneration, autologous bone grafting still represents the gold standard. However, autograft harvesting has limitations, including an additional surgery, donor site morbidity, and limited availability. Demineralized bone matrix (DBM) would represent an alternative, yet lacks sufficient osteoinductive properties. Combining DBM with a potent agent, such as bone morphogenetic protein-2 (BMP-2) might be a feasible alternative approach, optimizing an established grafting material with strong osteoinductive properties. A unique mixing device has been developed that enables perioperative handling to reach a homogeneous and standardized paste for bone defect filling. DBM proved in vitro to be a suitable carrier for BMP-2, with a documented release over 56 days at concentrations sufficient to stimulate osteogenic differentiation. At the end of the elution experiment, 56 days, bioactive BMP was still captured within the DBM. Using a sheep drill hole defect model, DBM perioperatively mixed with BMP-2 showed strong osteoinductive properties comparable to those of autologous bone and outnumbering the one of DBM alone or empty defects. Bone defect healing was enabled at diaphyseal and metaphyseal defects and thus BMP-2-doped DBM represented an easy perioperative enriching method and an efficient carrier for BMP-2. With the comparability to the clinical gold standard autologous bone, DBM mixed with BMP-2 might serve as possible alternative grafting material enabling a controlled osteogenic stimulation.

Splenic pooling and loss of VCAM-1 causes an engraftment defect in patients with myelofibrosis after allogeneic hematopoietic stem cell transplantation.

Myelofibrosis is a myeloproliferative neoplasm that results in cytopenia, bone marrow fibrosis and extramedullary hematopoiesis. Allogeneic hematopoietic stem cell transplantation is the only curative treatment but is associated with a risk of delayed engraftment and graft failure. In this study, patients with myelofibrosis (n=31) and acute myeloid leukemia (n=31) were analyzed for time to engraftment, graft failure and engraftment-related factors. Early and late neutrophil engraftment and late thrombocyte engraftment were significantly delayed in patients with myelofibrosis as compared to acute myeloid leukemia, and graft failure only occurred in myelofibrosis (6%). Only spleen size had a significant influence on engraftment efficiency in myelofibrosis patients. To analyze the cause for the engraftment defect, clearance of hematopoietic stem cells from peripheral blood was measured and immunohistological staining of bone marrow sections was performed. Numbers of circulating CD34+ were significantly reduced at early time points in myelofibrosis patients, whereas CD34+CD38- and colony-forming cells showed no significant difference in clearance. Staining of bone marrow sections for homing proteins revealed a loss of VCAM-1 in myelofibrosis with a corresponding significant increase in the level of soluble VCAM-1 within the peripheral blood. In conclusion, our data suggest that reduced engraftment and graft failure in myelofibrosis patients is caused by an early pooling of CD34+ hematopoietic stem cells in the spleen and a bone marrow homing defect caused by the loss of VCAM-1. Improved engraftment in myelofibrosis might be achieved by approaches that reduce spleen size and cleavage of VCAM-1 in these patients prior to hematopoietic stem cell transplantation.

Telomere shortening in enterocytes of patients with uncontrolled acute intestinal graft-versus-host disease.

Acute intestinal graft-versus-host disease (aGVHD) refractory to immunosuppressive treatment is a serious complication after allogenic hematopoietic stem cell transplantation (HSCT). The underlying mechanisms of refractory aGVHD of the gut are not fully understood. Although telomere length (TL) reflects the replicative history of a cell, critically short telomeres have been associated with replicative exhaustion and tissue failure. In this study, we demonstrate that enterocytes of patients with refractory intestinal aGVHD show significantly increased proliferation, which translates into significant and critical telomere attrition following HSCT as compared with unaffected patients undergoing HSCT. Calculated telomere loss in aGVHD patients is 190 bp/wk, thereby massively exceeding physiological steady-state TL shortening rates such as in lymphocytes (∼50 bp/y). Our data support the hypothesis that increased compensatory proliferation following continued tissue damage can result in massive telomere loss in enterocytes of aGVHD patients. The present study introduces aGVHD-triggered increased cellular turnover and telomere loss with subsequent replicative exhaustion as a mechanism for refractory gut GVHD that is compatible with the long-term clinical aspect of the disease and provides a basis for stem cell protective therapies in the treatment of aGVHD.

Retinal Involvement in a Patient with Cerebral Manifestation of Chronic Graft-Versus-Host-Disease.

BACKGROUND: We report a 35-year-old female patient with cerebral manifestations of chronic graft-versus-host disease (cGvHD) and putative retinal involvement after allogeneic peripheral blood stem cell transplantation (alloHSCT). PATIENT AND METHODS: The patient experienced recurrent episodes of fever and encephalitic signs 7 months after alloHSCT during taper of immunosuppression (IS). RESULTS: Cerebral magnetic resonance imaging (MRI) showed non-gadolinium-enhancing confluent periventricular lesions and cerebrospinal fluid inflammation. After exclusion of infectious causes, treatment with steroids and antiepileptics improved cognitive deficits. Steroid reduction provoked a relapse responding to IS. 2 years later, she complained of right-sided blurred vision and floaters; both eyes showed whitish circumscribed retinal infiltrations, cellular infiltration of the vitreous and mild bilateral optic disc edema. Oncological and neurological work-up ruled out infectious diseases and other GvHD manifestations. Symptoms and signs resolved under continued systemic IS, leaving pigmented retinal scars. After IS withdrawal, classical cutaneous cGvHD developed, resolving on systemic IS. 94 months after transplantation, she is doing well. CONCLUSION: To our knowledge, this is the first observation of retinal involvement of cerebral cGvHD manifestations with retinal infiltrations documented in the absence of other causes and in parallel to periventricular lesions in cerebral MRI. Based on bone marrow histology, we discuss a small vessel pathophysiology of cGvHD.

Consensus diagnostic histopathological criteria for acute gastrointestinal graft versus host disease improve interobserver reproducibility.

Graft versus host disease (GvHD) is a clinically important complication after allogeneic hematopoietic stem cell transplantation (HSCT). Its diagnosis relies on clinical and histopathological findings. In order to evaluate and improve inter-institutional diagnostic agreement on histological diagnosis and grading of acute gastrointestinal GvHD, we conducted a round robin test, which included 33 biopsies from 23 patients after HSCT. Five pathologists from different institutions independently evaluated the original sections from the biopsies submitted for diagnosis. Based on their results, consensus qualitative criteria for the assessment of typical histological features of GvHD (e.g., apoptosis, crypt destruction, mucosa denudation) were proposed, including detailed descriptions as well as histological images. In a second round robin test with involvement of the same pathologists, the reproducibility of both diagnosis and grading had improved. Remaining differences were mostly related to differential diagnostic considerations, including viral infection or toxic side effects of medication, which should be resolved by integrating histopathological findings with proper clinical information.

NIH Consensus development project on criteria for clinical trials in chronic graft-versus-host disease: II. The 2014 Pathology Working Group Report.

The 2005 National Institute of Health (NIH) Consensus Conference outlined histopathological diagnostic criteria for the major organ systems affected by both acute and chronic graft-versus-host disease (GVHD). The 2014 Consensus Conference led to this updated document with new information from histopathological studies of GVHD in the gut, liver, skin, and oral mucosa and an expanded discussion of GVHD in the lungs and kidneys. The recommendations for final histological diagnostic categories have been simplified from 4 categories to 3: no GVHD, possible GVHD, and likely GVHD, based on better reproducibility achieved by combining the previous categories of "consistent with GVHD" and "definite GVHD" into the single category of "likely GVHD." Issues remain in the histopathological characterization of GVHD, particularly with respect to the threshold of histological changes required for diagnostic certainty. Guidance is provided for the incorporation of biopsy information into prospective clinical studies of GVHD, particularly with respect to biomarker validation.

Preventive azithromycin treatment reduces noninfectious lung injury and acute graft-versus-host disease in a murine model of allogeneic hematopoietic cell transplantation.

Noninfectious lung injury and acute graft-versus-host disease (GVHD) after allogeneic hematopoietic cell transplantation (allo-HCT) are associated with significant morbidity and mortality. Azithromycin is widely used in allogeneic HCT recipients for pulmonary chronic GVHD, although current data appear controversial. We induced GVHD and noninfectious lung injury in lethally irradiated B6D2F1 mice by transplanting bone marrow and splenic T cells from allogeneic C57BL/6 mice. Experimental groups were treated with oral azithromycin starting on day 14 until the end of week 6 or week 14 after transplantation. Azithromycin treatment resulted in improved survival and decreased lung injury; the latter characterized by improved pulmonary function, reduced peribronchial and perivascular inflammatory cell infiltrates along with diminished collagen deposition, and a decrease in lung cytokine and chemokine expression. Azithromycin also improved intestinal GVHD but did not affect liver GVHD at week 6 early after transplantation. At week 14, azithromycin decreased liver GVHD but had no effect on intestinal GVHD. In vitro, allogeneic antigen-presenting cell (APC)- dependent T cell proliferation and cytokine production were suppressed by azithromycin and inversely correlated with relative regulatory T cell (Treg) expansion, whereas no effect was seen when T cell proliferation occurred APC independently through CD3/CD28-stimulation. Further, azithromycin reduced alloreactive T cell expansion but increased Treg expansion in vivo with corresponding downregulation of MHC II on CD11c(+) dendritic cells. These results demonstrate that preventive administration of azithromycin can reduce the severity of acute GVHD and noninfectious lung injury after allo-HCT, supporting further investigation in clinical trials.

Tumor-Induced Osteomalacia: Increased Level of FGF-23 in a Patient with a Phosphaturic Mesenchymal Tumor at the Tibia Expressing Periostin.

In our case, a 45-year-old male patient had multiple fractures accompanied by hypophosphatemia. FGF-23 levels were significantly increased, and total body magnetic resonance imaging (MRI) revealed a tumor mass located at the distal tibia leading to the diagnosis of tumor-induced osteomalacia (TIO). After resection of the tumor, hypophosphatemia and the increased levels of FGF-23 normalized within a few days. Subsequent microscopic examination and immunohistochemical analysis revealed a phosphaturic mesenchymal tumor mixed connective tissue variant (PMTMCT) showing a positive expression of somatostatin receptor 2A (SSTR2A), CD68, and Periostin. Electron microscopy demonstrated a poorly differentiated mesenchymal tumor with a multifocal giant cell component and evidence of neurosecretory-granules. However, the resected margins showed no tumor-free tissue, and therefore a subsequent postoperative radiotherapy was performed. The patient is still in complete remission after 34 months. Tumor resection of PMTMCTs is the therapy of choice. Subsequent radiotherapy in case of incompletely resected tumors can be an important option to avoid recurrence or metastasis even though this occurs rarely. The prognostic value of expression of Periostin has to be evaluated more precisely in a larger series of patients with TIO.

Cytostatic conditioning in experimental allogeneic bone marrow transplantation: Busulfan causes less early gastrointestinal toxicity but Treosulfan results in improved immune reconstitution.

BACKGROUND: Acute graft versus host disease (aGVHD) after allogeneic bone marrow transplantation (allo-BMT) is associated with significant morbidity and mortality. We evaluated the impact of the conditioning regimen on aGVHD comparing Treosulfan (Treo) and Busulfan (Bu) with total body irradiation (TBI). METHODS: Using a haploidentical murine model, B6D2F1 mice conditioned with Bu (100 mg/kg)/Fludarabine (Flu, 500 mg/kg) or Treo (6000 mg/kg)/Flu (500 mg/kg) or TBI with 14 Gy received bone marrow cells and splenocytes (20 × 10(6)) from either syngeneic (B6D2F1) or allogeneic (C57BL/6N) donors in order to analyze aGVHD outcome. RESULTS: Conditioning with Bu/Flu or Treo/Flu resulted in significantly reduced aGVHD severity and improved survival (p < 0.05) after allo-BMT compared to TBI. On day 5 after allo-BMT, the organ damages of Bu/Flu conditioned animals were significantly reduced in association with diminished expression of tumor necrosis factor in serum compared to Treo/Flu. Interestingly, the early toxicity of Treo/Flu did not result in significantly higher aGVHD severity; furthermore, a significantly improved immune reconstitution of B220-positive B cells was observed at day 42 after Treo/Flu conditioning compared to Bu/Flu. CONCLUSION: Conditioning with Treo/Flu or Bu/Flu results in decreased aGVHD severity compared to TBI. Moreover, Treo/Flu was associated with improved immune reconstitution despite the early toxicity.

Consensus on the histopathological evaluation of liver biopsies from patients following allogeneic hematopoietic cell transplantation.

After allogeneic hematopoietic cell transplantation (alloHCT) liver biopsy is performed for enigmatic liver disorders when noninvasive diagnostic steps have failed in establishing a definitive diagnosis. This document provides an updated consensus on the prerequisites for proper evaluation of liver biopsies in alloHCT patients and the histological diagnostic criteria for liver graft-versus-host disease (GvHD). The Working Group's recommendations for the histological diagnosis of liver GvHD were derived from the peer-reviewed literature and from the consensus diagnosis of a total of 30 coded liver biopsies. Acceptance of the recommendations was tested by a survey distributed to all HCT centers in Austria, Germany and Switzerland. Consensus was achieved for biopsy indications, methods of sample acquisition and processing, reporting and interpretation of biopsy findings. As GvHD is variably treated and the treatment modalities have changed over time, the panel endorses the use of more frequent biopsies in clinical studies in order to improve the present challenging clinical and diagnostic situation.

Entrapment syndrome of multiple nerves in graft-versus-host disease.

INTRODUCTION: Peripheral nerve entrapment syndromes are associated with hereditary neuropathy with liability to pressure palsies and a variety of rheumatic and endocrinological diseases. METHODS: We report a patient with entrapment syndromes of multiple nerves associated with chronic graft-versus-host-disease (GVHD) after allogeneic hematopoietic stem cell transplantation. Nerve ultrasound, histology, and ultrastructural changes were assessed. RESULTS: The 51-year-old man had developed severe deep dermal sclerosis due to chronic GVHD with a progressive polyneuropathy and entrapment syndromes of multiple nerves. Pre-stenotic enlargement was shown by nerve ultrasound. Histology demonstrated fibrosis of the epineurium with scarce infiltration of macrophages. Electron microscopy demonstrated alterations of the myelin sheaths and marked depletion of normal-sized myelinated nerve fibers. CONCLUSIONS: In addition to polyneuropathy, chronic GVHD can be associated with peripheral nerve entrapment syndromes and should be added to the differential diagnosis of compressive neuropathies.

Low Paneth cell numbers at onset of gastrointestinal graft-versus-host disease identify patients at high risk for nonrelapse mortality.

Acute graft-versus-host disease (GVHD) of the gastrointestinal (GI) tract is an often lethal complication of allogeneic hematopoietic cell transplant. Clinical severity correlates with outcomes, but histopathologic grading is primarily used to confirm the clinical diagnosis. One barrier to using histopathologic grading to predict clinical outcomes is inter-grader variability among transplant centers. Recent experimental models have shown that the loss of Paneth cells, which are located in the small intestine and help regulate the GI microbiome by secreting antimicrobial peptides, correlates with clinical GVHD severity. Because Paneth cells are easy to identify and quantify by light microscopy, we evaluated the mean number of Paneth cells per high-powered field (hpf) in 116 duodenal biopsies obtained at diagnosis of GI GVHD at 2 different centers with their clinical outcomes. Paneth cell counts were reproducible between centers (r(2) = 0.81; P < .0001). Lower numbers of Paneth cells at diagnosis correlated with clinically more severe GI GVHD (P < .0001) and less likelihood of response to GVHD treatment (P < .0001). A threshold of 4 Paneth cells per hpf stratified patients according to nonrelapse mortality (28% vs 56%; P = .004). We conclude that the enumeration of duodenal Paneth cells is a readily available index of disease severity that provides important information regarding GVHD prognosis.

Murine cytomegalovirus immediate-early 1 gene expression correlates with increased GVHD after allogeneic hematopoietic cell transplantation in recipients reactivating from latent infection.

The success of allogeneic (allo) hematopoietic cell transplantation (HCT) is limited by its treatment related complications, mostly graft versus host disease (GVHD) and fungal and viral infections. CMV reactivation after HCT has been associated with increased morbidity and mortality, and a causal relation between GVHD, immunosuppressive therapy and vice versa has been postulated. Using a low GVHD severity murine HCT model, we assessed the role of MCMV reactivation and GVHD development. BALB/c mice were infected with either murine CMV (MCMV) or mock and monitored for 25 weeks to establish latency, followed by sublethal irradiation conditioning and infusion of bone marrow plus splenocytes from either syngeneic (syn) BALB/c or allo B10.D2 donors. Engraftment of allo donor cells was confirmed by PCR for D2Mit265 gene product size. Day+100 mortality and overall GVHD severity in allo MCMV pre-infected recipients was higher than in allo mock controls. Pathologic changes of lung and liver GVHD in immediate-early gene 1 (IE1) positive recipients were significantly increased compared to mock controls, and were only slightly increased in IE1 negative. No significant gut injury was seen in any group. Aggravated lung injury in IE1 positive recipients correlated with higher BAL cell counts both for total cells and for CD4+ T cells when compared with mock controls, and also with protein expression of lung IFN-gamma and liver TNF. No evidence for CMV specific morphologic changes was seen on histopathology in any organ of IE1 positive recipients, suggesting that CMV reactivation is related to increased GVHD severity but does not require active CMV disease, strengthening the concept of a reciprocal relationship between CMV and GVHD.