Potential of FX06 to prevent disease progression in hospitalized non-intubated COVID-19 patients — the randomized, EU-wide, placebo-controlled, phase II study design of IXION

Background: More than 2.7 million hospitalizations of COVID-19-infected patients have occurred in Europe alone since the outbreak of the coronavirus in 2020. Interventions against SARS-CoV-2 are still in high need to prevent admissions to ICUs worldwide. FX06, a naturally occurring peptide in humans and other mammals, has the potential to reduce capillary leak by improving endothelial dysfunction and thus preventing the deterioration of patients. With IXION, we want to investigate the potential of FX06 to prevent disease progression in hospitalized, non-intubated COVID-19 patients. Methods: IXION is an EU-wide, multicentre, placebo-controlled, double-blinded, parallel, randomized (2:1) phase II clinical study. Patient recruitment will start in September 2022 (to Q2/2023) in Germany, Italy, Lithuania, Spain, Romania, Portugal, and France. A total of 306 hospitalized patients (>= 18 years and < 75 years) with a positive SARS-CoV-2 PCR test and a COVID-19 severity of 4-6 according to the WHO scale will be enrolled. After randomization to FX06 or placebo, patients will be assessed until day 28 (and followed up until day 60). FX06 (2 x 200 mg per day) or placebo will be administered intravenously for 5 consecutive days. The primary endpoint is to demonstrate a difference in the proportion of patients with progressed/worsened disease state in patients receiving FX06 compared to patients receiving placebo. Secondary endpoints are lung function, oxygen saturation and breathing rate, systemic inflammation, survival, capillary refill time, duration of hospital stay, and drug accountability. Discussion: With IXION, the multidisciplinary consortium aims to deliver a new therapy in addition to standard care against SARS-CoV-2 for the clinical management of COVID-19 during mild and moderate stages. Potential limitations might refer to a lack of recruiting and drop-out due to various possible protocol violations. While we controlled for drop-outs in the same size estimation, recruitment problems may be subject to external problems difficult to control for

A study of precast concrete bridge units

"April 10, 1961.""Cooperative research project sponsored by Missouri State Highway Commission U.S. Bureau of Public Roads."On June 1, 1955 a research program to study design and economic criteria for precast bridge units was undertaken by the Engineering Experiment Station of the University of Missouri in cooperation with the Missouri State Highway Department and the Bureau of Public Roads. The primary purpose of this research project was to analyze typical sections under simulated loading conditions in order to determine both their behavior at nominal working loads and their ultimate capacities with respect to flexural or shear failures. It was further desired to study the behavior of a group of the precast structural units joined together to simulate a typical field installation. In order to accomplish these aims, a series of tests on full-scale and on half scale models of the standard precast bridge units presently utilized by the Missouri State Highway Department was undertaken. This report presents the results of these tests and the conclusions developed therefrom

PTCH-1 and MDM2 expression in ameloblastoma from a West African sub-population: implication for chemotherapeutics

Introduction: ameloblastoma is a slow growing, painless odontogenic swelling which can attain sizes that result in severe deformities of the craniofacial complex. It is the most commonly encountered odontogenic tumor in Nigeria. Surgical intervention is currently the method of treatment; however identification of altered molecular pathways may inform chemotherapeutic potential. The Protein Patched homolog 1 (PTCH-1) is overexpressed in ameloblastoma. Also, mutation in the MDM2 gene can reduce the tumor suppressor function of p53 and promote ameloblastoma growth. No study however has characterized the molecular profile of African cases of ameloblastoma with a view to developing chemotherapeutic alternatives. The objective was to characterize the PTCH-1 genetic profile of Ameloblastoma in Nigerian patients as a first step in investigating its potential for chemotherapeutic intervention. Methods: twenty-eight FFPE blocks of ameloblastoma cases from Nigerian patients were prepared for antibody processing to PTCH-1 (Polyclonal Anti-PTCH antibody ab39266) and MDM2 (Monoclonal Anti-MDM2 antibody (2A10) ab16895). Cytoplasmic brown staining was considered as positive for PTCH while nuclear staining was positive for MDM2. Results: moderate and strong expressions for PTCH in ameloblast and stellate reticulum were 78.6% and 60.7% respectively. Only 3 (10.7%) cases expressed MDM2. Conclusion: the importance of our study is that it supports, in theory, anti-PTCH/SHH chemotherapeutics for Nigerian ameloblastoma cases and also infers the possible additional use of anti-p53 agents

Platelet-rich fibrin secretome induces three dimensional angiogenic activation in vitro

Different tissue engineering techniques are used to support rapid vascularisation. A novel technique is the use of platelet-rich fibrin (PRF), an autologous source of growth factors. This study was the first to investigate the influence of PRF matrices, isolated following different centrifugation protocols, on human dermal vascular endothelial cells (ECs) in mono-culture and co-culture with human primary fibroblasts (HFs) as an in vitro model for tissue regeneration. Focus was placed on vascular structure formation and growth factor release. HFs and ECs were cultivated with PRF prepared using a high (710 ×g) or low (44 ×g) relative centrifugation force (RCF) over 14 d. Immunofluorescence staining and immunohistochemistry were used to evaluate the microvascular formation. Cell culture supernatants were collected for evaluation of growth factor release. The results showed a PRF-mediated effect on the induction of angiogenesis in ECs. Microvessel-like structure formation was promoted when ECs were combined with low-RCF PRF as compared to high-RCF PRF or control group. The percentage of vascular lumen area was significantly higher in low-RCF PRF, especially at day 7, which coincided with statistically significantly higher growth factor [vascular endothelial factor (VEGF), transforming growth factor β1 (TGF-β1) and platelet derived growth factor (PDGF)] concentration measured in low-RCF PRF as compared to high-RCF PRF or control group. In conclusion, reducing the RCF according to the low-speed centrifugation concept (LSCC) resulted in increased growth factor release and angiogenic structure formation with EC mono-culture, suggesting that PRF may be a highly beneficial therapeutic tool for tissue engineering applications

Mutations of FBN1 and genotype-phenotype correlations in Marfan syndrome and related fibrillinopathies

The Marfan syndrome (MFS) is a pleiotropic, autosomal dominant disorder of connective tissue with highly variable clinical manifestations including aortic dilatation and dissection, ectopia lentis, and a series of skeletal anomalies. Mutations in the gene for fibrillin-1 (FBN1) cause MFS, and at least 337 mainly unique mutations have been published to date. FBN1 mutations have been found not only in MFS but also in a range of connective tissue disorders collectively termed fibrillinopathies ranging from mild phenotypes, such as isolated ectopia lentis, to severe disorders including neonatal MFS, which generally leads to death within the first two years of life. The present article intends to provide an overview of mutations found in MFS and related disorders and to discuss potential genotype-phenotype correlations in MFS

Analysis of the in vitro degradation and the in vivo tissue response to bi-layered 3D-printed scaffolds combining PLA and biphasic PLA/bioglass components – Guidance of the inflammatory response as basis for osteochondral regeneration

The aim of the present study was the in vitro and in vivo analysis of a bi-layered 3D-printed scaffold combining a PLA layer and a biphasic PLA/bioglass G5 layer for regeneration of osteochondral defects in vivo Focus of the in vitro analysis was on the (molecular) weight loss and the morphological and mechanical variations after immersion in SBF. The in vivo study focused on analysis of the tissue reactions and differences in the implant bed vascularization using an established subcutaneous implantation model in CD-1 mice and established histological and histomorphometrical methods. Both scaffold parts kept their structural integrity, while changes in morphology were observed, especially for the PLA/G5 scaffold. Mechanical properties decreased with progressive degradation, while the PLA/G5 scaffolds presented higher compressive modulus than PLA scaffolds. The tissue reaction to PLA included low numbers of BMGCs and minimal vascularization of its implant beds, while the addition of G5 lead to higher numbers of BMGCs and a higher implant bed vascularization. Analysis revealed that the use of a bi-layered scaffold shows the ability to observe distinct in vivo response despite the physical proximity of PLA and PLA/G5 layers. Altogether, the results showed that the addition of G5 enables to reduce scaffold weight loss and to increase mechanical strength. Furthermore, the addition of G5 lead to a higher vascularization of the implant bed required as basis for bone tissue regeneration mediated by higher numbers of BMGCs, while within the PLA parts a significantly lower vascularization was found optimally for chondral regeneration. Thus, this data show that the analyzed bi-layered scaffold may serve as an ideal basis for the regeneration of osteochondral tissue defects. Additionally, the results show that it might be able to reduce the number of experimental animals required as it may be possible to analyze the tissue response to more than one implant in one experimental animal

Influence of concentration and preparation of platelet rich fibrin on human bone marrow mononuclear cells (in vitro)

Large bone defects have always been a big challenge. The use of bone marrow mononuclear cells (BMCs) combined with an osteoconductive scaffold has been proved a good alternative for the treatment of large bone defects. Another autologous source for tissue engineering is platelet rich fibrin (PRF). PRF is a blood concentrate system obtained through a one-step centrifugation. The generated 3D matrix of the PRF clot serves as a reservoir of growth factors. Those growth factors might support the regenerative response of BMC, and therefore the effect of PRF, centrifuged with either high medium (208 g) or low (60 g) relative centrifugation force (RCF) on BMCs was evaluated in vitro in the present study. The two PRF matrices obtained were initially characterized and compared to human serum. Significantly increased concentrations of insulin-like growth factor (IGF), soluble intercellular adhesion molecule-1 (sICAM1) and transforming growth factor (TGF)-β were found in PRF compared to human serum whereas VEGF concentration was not significantly altered. A dose-response study revealed no further activation of BMC’s metabolic activity, if concentration of both PRF matrices exceeded 10% (v/v). Effect of both PRF preparations [10%] on BMC was analyzed after 2, 7, and 14 days in comparison to human serum [10%]. Metabolic activity of BMC increased significantly in all groups on day 14. Furthermore, gene expression of matrix metalloproteinases (MMP)-2, −7, and −9 was significantly stimulated in BMC cultivated with the respective PRF matrices compared to human serum. Apoptotic activity of BMC incubated with PRF was not altered compared to BMC cultivated with serum. In conclusion, PRF could be used as a growth factor delivery system of autologous or allogeneic source with the capability of stimulating cells such as BMC

A low-speed centrifugation concept leads to cell accumulation and vascularization of solid platelet-rich fibrin: an experimental study in vivo

Platelet-rich fibrin (PRF) is generated from the patients’ own venous blood by a single centrifugation step without the additional use of anticoagulants. Based on the previously described LSCC (low-speed centrifugation concept), our group showed that modification of the centrifugation setting, that is, reducing the relative centrifugal force (RCF) and mildly increasing the centrifugation time, resulted in modified solid and liquid PRF-matrices with increased number of platelets, leukocytes, and growth factors’ concentrations. The aim of this study was to determine whether RCF reduction might also result in different tissue reactions toward the two PRF-based matrices, especially vascularization and cell distribution in vivo. Two centrifugation protocols (PRF-high [719 g] and PRF-medium [222 g]) were compared in a subcutaneous implantation model of SCID mice at 5 and 10 days. Histological and histomorphometrical analyses were performed to quantify lymphocyte, neutrophil, human macrophage, and monocyte populations. CD31 was used to detect newly formed vessels, while all human cells were detected by using human vimentin as a pan-cellular marker. The results demonstrated that PRF-high elicited a dense and stable fibrin structure and prevented cellular penetration of the host tissue. By contrast, PRF-medium was more porous, had a significantly higher in vivo vascularization rate, and included significantly more human cells, especially at day 10, compared to PRF-high. These findings highlight the possibility of modifying the structure and composition of PRF matrices and thus selectively altering their regenerative potential in vivo. Clinical studies now must evaluate the different PRF matrices for bone and soft-tissue regeneration to validate possible benefits using personalized preparation protocols

TGGE screening of the entire FBN1 coding sequence in 126 individuals with marfan syndrome and related fibrillinopathies

Mutations in the gene for fibrillin-1 (FBN1) cause Marfan syndrome (MFS), an autosomal dominant heritable disorder of connective tissue with prominent manifestations in the skeletal, ocular, and cardiovascular system. FBN1 mutations have also been identified in a series of related disorders of connective tissue collectively termed type-1 fibrillinopathies. We have developed temperature-gradient gel electrophoresis (TGGE) assays for all 65 FBN1 exons, screened 126 individuals with MFS, other type-1 fibrillinopathies, and other potentially related disorders of connective tissue for FBN1 mutations, and identified a total of 53 mutations, of which 33 are described here for the first time. Several mutations were identified in individuals with fibrillinopathies other than classic Marfan syndrome, including aneurysm of the ascending aorta with only minor skeletal anomalies, and several individuals with only skeletal and ocular involvement. The mutation detection rate in this study was 42% overall, but was only 12% in individuals not fulfilling the diagnostic criteria for MFS, suggesting that clinical overdiagnosis is one reason for the low detection rate observed for FBN1 mutation analysis. Hum Mutat 20:197-208, 2002. © 2002 Wiley-Liss, Inc