An Elastin-Derived Composite Matrix for Enhanced Vascularized and Innervated Bone Tissue Reconstruction: From Material Development to Preclinical Evaluation.

Despite progress in bone tissue engineering, reconstruction of large bone defects remains an important clinical challenge. Here, a biomaterial designed to recruit bone cells, endothelial cells, and neuronal fibers within the same matrix is developed, enabling bone tissue regeneration. The bioactive matrix is based on modified elastin-like polypeptides (ELPs) grafted with laminin-derived adhesion peptides IKVAV and YIGSR, and the SNA15 peptide for retention of hydroxyapatite (HA) particles. The composite matrix shows suitable porosity, interconnectivity, biocompatibility for endothelial cells, and the ability to support neurites outgrowth by sensory neurons. Subcutaneous implantation leads to the formation of osteoid tissue, characterized by the presence of bone cells, vascular networks, and neuronal structures, while minimizing inflammation. Using a rat femoral condyle defect model, longitudinal micro-CT analysis is performed, which demonstrates a significant increase in the volume of mineralized tissue when using the ELP-based matrix compared to empty defects and a commercially available control (Collapat). Furthermore, visible blood vessel networks and nerve fibers are observed within the lesions after a period of two weeks. By incorporating multiple key components that support cell growth, mineralization, and tissue integration, this ELP-based composite matrix provides a holistic and versatile solution to enhance bone tissue regeneration.

OCT to identify macular changes not clinically detectable in elderly patients with cataract.

PURPOSE: To demonstrate the importance of optical coherence tomography (OCT) in identifying macular changes not detected at clinical examination preceding cataract surgery in patients older than 60 years. SETTING: Hospital Oftalmológico Visão Laser, Santos, São Paulo, Brazil. DESING: Prospective case series. METHODS: Patients older than 60 years were selected for this cross-sectional prospective study conducted during preoperative examination for cataract surgery. Eyes with a previous diagnosis or clinical evidence of macular disease or with media opacity preventing OCT from being performed were excluded from the study. All the study participants underwent OCT and were then divided into 2 groups: patients with macular changes and patients without macular changes found on OCT. RESULTS: Of 364 eyes screened (212 patients), 300 eyes (180 patients) were included in the study. OCT identified macular changes in 40 eyes (13.3%), with age-related macular degeneration being found in 13 eyes (4.3%), epiretinal membrane in 12 eyes (4%), intraretinal fluid in 12 eyes (4%), and macular hole in 3 eyes (1%). In the group with macular changes, the mean age was 74.4 ± 6.3 years compared with 70.4 ± 6.7 years for those without changes ( P < .001). CONCLUSIONS: OCT proved an effective method for identifying macular diseases not detected at clinical evaluation before cataract surgery. Therefore, the relevance of performing OCT in these cases was confirmed and should be taken into consideration, particularly when evaluating patients older than 60 years.

Bone Regeneration in Small and Large Segmental Bone Defect Models after Radiotherapy Using Injectable Polymer-Based Biodegradable Materials Containing Strontium-Doped Hydroxyapatite Particles.

The reconstruction of bones following tumor excision and radiotherapy remains a challenge. Our previous study, performed using polysaccharide-based microbeads that contain hydroxyapatite, found that these have osteoconductivity and osteoinductive properties. New formulations of composite microbeads containing HA particles doped with strontium (Sr) at 8 or 50% were developed to improve their biological performance and were evaluated in ectopic sites. In the current research, we characterized the materials by phase-contrast microscopy, laser dynamic scattering particle size-measurements and phosphorus content, before their implantation into two different preclinical bone defect models in rats: the femoral condyle and the segmental bone. Eight weeks after the implantation in the femoral condyle, the histology and immunohistochemistry analyses showed that Sr-doped matrices at both 8% and 50% stimulate bone formation and vascularization. A more complex preclinical model of the irradiation procedure was then developed in rats within a critical-size bone segmental defect. In the non-irradiated sites, no significant differences between the non-doped and Sr-doped microbeads were observed in the bone regeneration. Interestingly, the Sr-doped microbeads at the 8% level of substitution outperformed the vascularization process by increasing new vessel formation in the irradiated sites. These results showed that the inclusion of strontium in the matrix-stimulated vascularization in a critical-size model of bone tissue regeneration after irradiation.

Chemical composition and biological activities of the essential oil from Eugenia stipitata McVaugh leaves.

In the present study, the volatile components and cytotoxic, antibacterial, antioxidant, and antiprotozoal activities of the essential oil obtained from the leaves of Eugenia stipitata McVaugh (Myrtaceae) grown in the Brazilian Northeast region (Araripe) were investigated. The essential oil was obtained by hydrodistillation. The leaves of E. stipitata provided an oil yield of 0.13 ± 0.01% (w/w). The volatile compounds in the essential oil of E. stipitata were analysed using gas chromatography, and the volatile chemical composition was mainly composed of ß-eudesmol (15.28%), γ-eudesmol (10.85%), elemol (10.21%) and caryophyllene oxide (6.65%). The essential oil of E. stipitata was highly selective against Leishmania braziliensis and L. infantum promastigotes. The essential oil exhibited good antibacterial activity. E. stipitata essential oil showed low free-radical scavenging activity. Our results suggest that the E. stipitata essential oil is a relevant source of the primary compounds required for the development of antibacterial and antiprotozoal drugs.

Effective recommendations towards healthy routines to preserve mental health during the COVID-19 pandemic

Objective: To assess the adherence to a set of evidence-based recommendations to support mental health during the coronavirus disease 2019 (COVID-19) pandemic and its association with depressive and anxiety symptoms. Methods: A team of health workers and researchers prepared the recommendations, formatted into three volumes (1: COVID-19 prevention; 2: Healthy habits; 3: Biological clock and sleep). Participants were randomized to receive only Volume 1 (control), Volumes 1 and 2, Volumes 1 and 3, or all volumes. We used a convenience sample of Portuguese-speaking participants over age 18 years. An online survey consisting of sociodemographic and behavioral questionnaires and mental health instruments (Patient Health Questionnaire-9 [PHQ-9] and Generalized Anxiety Disorder-7 [GAD-7]) was administered. At 14 and 28 days later, participants were invited to complete follow-up surveys, which also included questions regarding adherence to the recommendations. A total of 409 participants completed the study - mostly young adult women holding university degrees. Results: The set of recommendations contained in Volumes 2 and 3 was effective in protecting mental health, as suggested by significant associations of adherence with PHQ-9 and GAD-7 scores (reflecting anxiety and depression symptoms, respectively). Conclusion: The recommendations developed in this study could be useful to prevent negative mental health effects in the context of the pandemic and beyond.

First report on the chemical composition of leaf essential oil of Myrciaria pilosa Sobral & Couto and its antimicrobial and antivirulence activities against Staphylococcus aureus.

Myrciaria pilosa is a tree species of the Brazilian Caatinga biome. This paper is the first report on the chemical composition and the antimicrobial and antivirulence activities of essential oil extracted from its leaves. The oil was extracted by hydrodistillation. Chemical composition determined by GC-MS and CG-FID revealed 63 compounds; the sesquiterpenes guaiol (13.17%) and (E)-ß-caryophyllene (11.26%) dominated. Antimicrobial activity against strains of Staphylococcus aureus was evaluated by the broth microdilution method. It showed minimum inhibitory concentrations (MIC) of 5 µg/mL against evaluated strains and minimum bactericidal concentrations (MBC) ranging from 10 to 20 µg/mL. Evaluation of antivirulence activity showed reductions of 92.0% and 47.2%, respectively, in haemolytic action and production of staphyloxanthin. These findings show that the essential oil of M. pilosa has potential as an antimicrobial drug to control infection by multi-resistant strains of S. aureus.

Shockwave Therapy Plus Eccentric Exercises Versus Isolated Eccentric Exercises for Achilles Insertional Tendinopathy: A Double-Blinded Randomized Clinical Trial.

BACKGROUND: There remains a lack of consensus regarding the treatment of Achilles insertional tendinopathy. The condition is typically treated with eccentric exercises despite the absence of satisfactory and sustained results. Shockwave therapy was presented as an alternative, but there is a paucity of literature, with good outcomes, supporting its use. The purpose of the present single-center, double-blinded, placebo-controlled, randomized trial was to determine if the use of shockwave therapy in combination with eccentric exercises improves pain and function in patients with Achilles insertional tendinopathy. METHODS: A total of 119 patients with Achilles insertional tendinopathy were evaluated and enrolled in the study from February 2017 to February 2019. Patients were allocated to 1 of 2 treatment groups, eccentric exercises with extracorporeal shockwave therapy (SWT group) and eccentric exercises with sham shockwave therapy (control group). Three sessions of radial shockwaves (or sham treatment) were performed every 2 weeks and eccentric exercises were undertaken for 3 months. The primary outcome was the Victorian Institute of Sport Assessment-Achilles questionnaire (VISA-A) at 24 weeks. Secondary outcomes included the visual analogue scale, algometry, the Foot and Ankle Outcome Score, and the 12-Item Short Form Health Survey. RESULTS: Both groups showed significant improvement during the study period; however, there were no between-group differences in any of the outcomes (all p >0.05). At the 24-week evaluation, the SWT group exhibited a mean VISA-A of 63.2 (95% confidence interval, 8.0) compared with 62.3 (95% confidence interval, 6.9) in the control group (p = 0.876). There was a higher rate of failure (38.3%) but a lower rate of recurrence (17.0%) in the SWT group compared with the control group (11.5% and 34.6%, respectively; p = 0.002 and p = 0.047). There were no complications reported for either group. CONCLUSIONS: Extracorporeal shockwave therapy does not potentiate the effects of eccentric strengthening in the management of Achilles insertional tendinopathy. LEVEL OF EVIDENCE: Therapeutic Level I. See Instructions for Authors for a complete description of levels of evidence.

Tumor-intrinsic and -extrinsic (immune) gene signatures robustly predict overall survival and treatment response in high grade serous ovarian cancer patients.

In the present study, we developed a transcriptomic signature capable of predicting prognosis and response to primary therapy in high grade serous ovarian cancer (HGSOC). Proportional hazard analysis was performed on individual genes in the TCGA RNAseq data set containing 229 HGSOC patients. Ridge regression analysis was performed to select genes and develop multigenic models. Survival analysis identified 120 genes whose expression levels were associated with overall survival (OS) (HR = 1.49-2.46 or HR = 0.48-0.63). Ridge regression modeling selected 38 of the 120 genes for development of the final Ridge regression models. The consensus model based on plurality voting by 68 individual Ridge regression models classified 102 (45%) as low, 23 (10%) as moderate and 104 patients (45%) as high risk. The median OS was 31 months (HR = 7.63, 95% CI = 4.85-12.0, P < 1.0-10) and 77 months (HR = ref) in the high and low risk groups, respectively. The gene signature had two components: intrinsic (proliferation, metastasis, autophagy) and extrinsic (immune evasion). Moderate/high risk patients had more partial and non-responses to primary therapy than low risk patients (odds ratio = 4.54, P < 0.001). We concluded that the overall survival and response to primary therapy in ovarian cancer is best assessed using a combination of gene signatures. A combination of genes which combines both tumor intrinsic and extrinsic functions has the best prediction. Validation studies are warranted in the future.

Retrospective Validation of a 168-Gene Expression Signature for Glioma Classification on a Single Molecule Counting Platform.

Gene expression profiling has been shown to be comparable to other molecular methods for glioma classification. We sought to validate a gene-expression based glioma classification method. Formalin-fixed paraffin embedded tissue and flash frozen tissue collected at the Augusta University (AU) Pathology Department between 2000-2019 were identified and 2 mm cores were taken. The RNA was extracted from these cores after deparaffinization and bead homogenization. One hundred sixty-eight genes were evaluated in the RNA samples on the nCounter instrument. Forty-eight gliomas were classified using a supervised learning algorithm trained by using data from The Cancer Genome Atlas. An ensemble of 1000 linear support vector models classified 30 glioma samples into TP1 with classification confidence of 0.99. Glioma patients in TP1 group have a poorer survival (HR (95% CI) = 4.5 (1.3-15.4), p = 0.005) with median survival time of 12.1 months, compared to non-TP1 groups. Network analysis revealed that cell cycle genes play an important role in distinguishing TP1 from non-TP1 cases and that these genes may play an important role in glioma survival. This could be a good clinical pipeline for molecular classification of gliomas.

Chemical composition, antioxidant and antiprotozoal activity of Eugenia gracillima Kiaersk. leaves essential oil.

This work evaluated the volatile composition, antioxidant and antiprotozoal activities of the essential oil obtained from leaves of Eugenia gracillima Kiaersk. (EGEO) grown in Brazilian Northeast area (Araripe, Brazil). The volatile compounds of EGEO were analyzed by GC and GC-MS and its chemical composition is mainly composed of sesquiterpene hydrocarbons (91.22%), oxygenated sesquiterpenes (7.45%) and monoterpene (1.01%). The most abundant volatile constituents of the EGEO were germacrene D (16.10%), γ-muurolene (15.60%), bicyclogermacrene (8.53%), germacrene B (7.43%), and Δ-elemene (6.06%). The oil showed weak to moderate antioxidant activity. EGEO was highly selective to Leishmania braziliensis and Leishmania infantum promastigotes with selective indexes of 73.66 and 71.41, respectively. EGEO did not inhibit Trypanosoma cruzi. These data suggest that the E. gracillima essential oil is a relevant source of lead compounds for development of anti-Leishmania drugs.

Prevalence and clinical profile of refractory hypertension in a large cohort of patients with resistant hypertension.

Refractory hypertension (RfHT) is an extreme phenotype of resistant hypertension (RHT) and is considered uncontrolled blood pressure (BP) despite the use of five or more antihypertensives. The objective of this study was to characterize the prevalence and clinical profile of RfHT patients in a historical cohort of patients with RHT at two different times: before and after the introduction of spironolactone. First, this cross-sectional study evaluated 1048 RHT patients (72.3% females, mean [SD] age: 61.2 [11.3] years) referred to a hypertension clinic (prespironolactone period). All patients were submitted to a standard protocol including clinical and complementary exams. Second, the analysis evaluated patients after the introduction of spironolactone (postspironolactone period). Statistical analysis included bivariate comparisons between patients with RHT and patients with RfHT and logistic regressions to assess the independent correlations of RfHT. A total of 146 patients (13.9%) remained refractory despite the use of at least five antihypertensives (prespironolactone period). After the introduction of spironolactone, the prevalence increased to 17.6%. For any criterion, RfHT patients were younger and more obese. In the initial period, current smoking and left ventricular hypertrophy were independently correlated with RfHT. Furthermore, after spironolactone use, RfHT patients had lower aortic stiffness and peripheral artery disease (PAD), pointing to a lower cardiovascular risk despite the lack of BP control. Younger age and lower prevalence of PAD correlated independently with RfHT. In conclusion, there was a high prevalence of RfHT, especially in younger and obese patients, and spironolactone use seemed to reduce cardiovascular risk despite the lack of BP control.

Comparative analysis of transcriptomic profile, histology, and IDH mutation for classification of gliomas.

Gliomas are currently classified through integration of histology and mutation information, with new developments in DNA methylation classification. However, discrepancies exist amongst the major classification methods. This study sought to compare transcriptome-based classification to the established methods. RNAseq and microarray data were obtained for 1032 gliomas from the TCGA and 395 gliomas from REMBRANDT. Data were analyzed using unsupervised and supervised learning and other statistical methods. Global transcriptomic profiles defined four transcriptomic glioma subgroups with 91.4% concordance with the WHO-defined mutation subtypes. Using these subgroups, 168 genes were selected for the development of 1000 linear support vector classifiers (LSVC). Based on plurality voting of 1000 LSVC, the final ensemble classifier confidently classified all but 17 TCGA gliomas to one of the four transcriptomic profile (TP) groups. The classifier was validated using a gene expression microarray dataset. TP1 cases include IDHwt, glioblastoma high immune infiltration and cellular proliferation and poor survival prognosis. TP2a is characterized as IDHmut-codel, oligodendrogliomas with high tumor purity. TP2b tissue is mostly composed of neurons and few infiltrating malignant cells. TP3 exhibit increased NOTCH signaling, are astrocytoma and IDHmut-non-codel. TP groups are highly concordant with both WHO integrated histology and mutation classification as well as methylation-based classification of gliomas. Transcriptomic profiling provides a robust and objective method to classify gliomas with high agreement to the current WHO guidelines and may provide additional survival prediction to the current methods.

Characterization of ß-cyclodextrin/myrtenol complex and its protective effect against nociceptive behavior and cognitive impairment in a chronic musculoskeletal pain model.

Myrtenol has gained wide interest because of its pharmacological profiles, mainly for treatment of chronic diseases. To improve the solubility of myrtenol, the formation of inclusion complexes with ß-cyclodextrin was performed by physical mixture, kneading process or slurry complexation (SC) methods and characterized using thermal analysis, XRD, SEM and NMR. From these results, myrtenol complexed by SC was successfully complexed into ß-cyclodextrin cavity. The interaction between myrtenol and ß-cyclodextrin was confirmed by molecular docking. Hence, the SC ß-cyclodextrin-myrtenol complex was evaluate for its anti-hyperalgesic, anxiolytic and antioxidant activity in a fibromyalgia model. Results show that myrtenol and ß-cyclodextrin form a stable complex and have anti-hyperalgesic effect, improve the cognitive impairment caused and have an anxiolytic-like effect. Furthermore, the ß-cyclodextrin/myrtenol complex decrease lipoperoxidation, increased catalase activity and a reduce SOD/CAT ratio. Therefore, ß-cyclodextrin/myrtenol complex reduce painful behavior, improves motor skills and emotional behavior and decreases oxidative stress in a fibromyalgia model.

Improvement of human pancreatic islet quality after co-culture with human adipose-derived stem cells.

The aim of this study was to investigate whether co-culture of human islets with adipose-derived stem cells (ASCs) can improve islet quality and to evaluate which factors play a role in the protective effect of ASCs against islet dysfunction. Islets and ASCs were cultured in three experimental groups for 24 h, 48 h, and 72 h: 1) indirect co-culture of islets with ASC monolayer (Islets/ASCs); 2) islets alone; and 3) ASCs alone. Co-culture with ASCs improved islet viability and function in all culture time-points analyzed. VEGFA, HGF, IL6, IL8, IL10, CCL2, IL1B, and TNF protein levels were increased in supernatants of islet/ASC group compared to islets alone, mainly after 24 h. Moreover, VEGFA, IL6, CCL2, HIF1A, XIAP, CHOP, and NFKBIA genes were differentially expressed in islets from the co-culture condition compared to islets alone. In conclusion, co-culture of islets with ASCs promotes improvements in islet quality.

Osteogenic differentiation of bone marrow-derived mesenchymal stem cells on anodized niobium surface.

Currently, titanium and its alloys are the most used materials for biomedical applications. However, because of the high costs of these metals, new materials, such as niobium, have been researched. Niobium appears as a promising material due to its biocompatibility, and excellent corrosion resistance. In this work, anodized niobium samples were produced and characterized. Their capacity to support the osteogenic differentiation of bone marrow-derived mesenchymal stem cells (BM-MSCs) was also tested. The anodized niobium samples were characterized by SEM, profilometry, XPS, and wettability. BM-MSCs were cultured on the samples during 14 days, and tested for cell adhesion, metabolic activity, alkaline phosphatase activity, and mineralization. Results demonstrated that anodization promotes the formation of a hydrophilic nanoporous oxide layer on the Nb surface, which can contribute to the increase in the metabolic activity, and in osteogenic differentiation of BM-MSCs, as well as to the extracellular matrix mineralization.

Development of a cell-free and growth factor-free hydrogel capable of inducing angiogenesis and innervation after subcutaneous implantation.

Despite significant progress in the field of biomaterials for bone repair, the lack of attention to the vascular and nervous networks within bone implants could be one of the main reasons for the delayed or impaired recovery of bone defects. The design of innovative biomaterials should improve the host capacity of healing to restore a functional tissue, taking into account that the nerve systems closely interact with blood vessels in the bone tissue. The aim of this work is to develop a cell-free and growth factor-free hydrogel capable to promote angiogenesis and innervation. To this end, we have used elastin-like polypeptides (ELPs), poly(ethylene glycol) (PEG) and increasing concentrations of the adhesion peptide IKVAV (25% (w/w) representing 1.7 mM and 50% (w/w) representing 4.1 mM) to formulate and produce hydrogels. When characterized in vitro, hydrogels have fine-tunable rheological properties, microporous structure and are biocompatible. At the biological level, 50% IKVAV composition up-regulated Runx2, Osx, Spp1, Vegfa and Bmp2 in mesenchymal stromal cells and Tek in endothelial cells, and sustained the formation of long neurites in sensory neurons. When implanted subcutaneously in mice, hydrogels induced no signals of major inflammation and the 50% IKVAV composition induced higher vessel density and formation of nervous terminations in the peripheral tissue. This novel composite has important features for tissue engineering, showing higher osteogenic, angiogenic and innervation potential in vitro, being not inflammatory in vivo, and inducing angiogenesis and innervation subcutaneously. STATEMENT OF SIGNIFICANCE: One of the main limitations in the field of tissue engineering remains the sufficient vascularization and innervation during tissue repair. In this scope, the development of advanced biomaterials that can support these processes is of crucial importance. Here, we formulated different compositions of Elastin-like polypeptide-based hydrogels bearing the IKVAV adhesion sequence. These compositions showed controlled mechanical properties, and were degradable in vitro. Additionally, we could identify in vitro a composition capable to promote neurite formation and to modulate endothelial and mesenchymal stromal cells gene expression, in view of angiogenesis and osteogenesis, respectively. When tested in vivo, it showed no signs of major inflammation and induced the formation of a highly vascularized and innervated neotissue. In this sense, our approach represents a potential advance in the development of new strategies to promote tissue regeneration, taking into account both angiogenesis and innervation.

Anti-staphylococcal activity of Syagrus coronata essential oil: Biofilm eradication and in vivo action on Galleria mellonela infection model.

In this study, essential oil extracted from Syagrus coronata seeds (SCEO) was evaluated for antibacterial and antibiofilm activities against Staphylococcus aureus; in addition, Galleria mellonella model was used as an in vivo infection model. SCEO was mainly composed by fatty acids (89.79%) and sesquiterpenes (8.5%). The major components were octanoic acid, dodecanoic acid, decanoic acid and γ-eudesmol. SCEO showed bactericidal activity (minimal bactericidal concentration from 312 to 1250 µg/mL) against all tested S. aureus clinical isolates, which showed distinct biofilm-forming and multiple drug resistance phenotypes. SCEO weakly reduced biomass but remarkably decreased cell viability in pre-formed biofilms of S. aureus isolate UFPEDA-02 (ATCC-6538). Electron microscopy analysis showed that SCEO treatments decreased the number of bacterial cells (causing structural alterations) and lead to loss of the roughness in the multiple layers of the three-dimensional biofilm structure. In addition, overproduction of exopolymeric matrix was observed. SCEO at 31.2 mg/kg improved the survival of G. mellonela larvae inoculated with UFPEDA-02 isolate and reduced the bacterial load in hemolymph and melanization. In conclusion, SCEO is an antibacterial agent against S. aureus strains with different resistance phenotypes and able to disturb biofilm architecture. Our results show SCEO as a potential candidate to drug development.