Highly Porous and Superabsorbent Biomaterial Made of Marine-Derived Polysaccharides and Ascorbic Acid as an Optimal Dressing for Exuding Wound Management

There are many modern wound dressings that have promising properties for repairing skin damage. However, due to various types of wounds and the problems they cause, there is still a great demand for new, effective healing strategies. The aim of this study was to create superabsorbent wound dressing made of marine-derived polysaccharides (agarose and chitosan) using the freeze-drying method. The secondary goal was its comprehensive evaluation for potential use as an external superabsorbent bandage for wounds with high exudation. Due to the well-known positive effect of ascorbic acid (vitamin C) on the healing process, biomaterial enriched with vitamin C was prepared and compared to the variant without the addition of ascorbic acid. It was shown that the produced foam-like wound dressing had a very porous structure, which was characterized by hydrophilicity, allowing a large amount of human fluids to be absorbed. According to in vitro tests on human fibroblasts, biomaterial was nontoxic and supportive to cell proliferation. Vitamin C-enriched dressing also had the ability to significantly reduce matrix metalloproteinase-2 production and to promote platelet-derived growth factor-BB synthesis by fibroblasts, which is desired during chronic wound treatment. The material has features of the eco-friendly wound care product since it was made of naturally-derived polysaccharides and was proved to be biodegradable. Importantly, despite degradable character, it was stable in the chronic and infected wound microenvironment, maintaining high integrity after 8-week incubation in the enzymatic solutions containing lysozyme and collagenases. The obtained results clearly showed that developed biomaterial possesses all necessary features of the external dressing for the management of exudate from both acute and chronic non-healing wounds.     

Evaluating spine micro-architectural texture (via TBS) discriminates major osteoporotic fractures from controls both as well as and independent of site matched BMD: the Eastern European TBS study

The aim of the study was to assess the clinical performance of the model combining areal bone mineral density (aBMD) at spine and microarchitecural texture (TBS) for the detection of the osteoporotic fracture. The Eastern European Study is a multicenter study (Serbia, Bulgaria, Romania and Ukraine) evaluating the role of TBS in routine clinical practice as a complement to aBMD. All scans were acquired on Hologic Discovery and GE Prodigy densitometers in a routine clinical manner. The additional clinical values of aBMD and TBS were analyzed using a two steps classification tree approach (aBMD followed by TBS tertiles) for all type of osteoporotic fracture (All-OP Fx). Sensitivity, specificity and accuracy of fracture detection as well as the Net Reclassification Index (NRI) were calculated. This study involves 1031 women subjects aged 45 and older recruited in east European countries. Clinical centers were cross-calibrated in terms of BMD and TBS. As expected, areal BMD (aBMD) at spine and TBS were only moderately correlated (r ² = 0.19). Prevalence rate for All-OP Fx was 26 %. Subjects with fracture have significant lower TBS and aBMD than subjects without fracture (p < 0.01). TBS remains associated with the fracture even after adjustment for age and aBMD with an OR of 1.27 [1.07–1.51]. When using aBMD T-score of ?2.5 and the lowest TBS tertile thresholds, both BMD and TBS were similar in terms of sensitivity (35 vs. 39 %), specificity (78 vs. 80 %) and accuracy (64 vs. 66 %). aBMD and TBS combination, induced a significant improvement in sensitivity (+28 %) and accuracy (+17 %) compared to aBMD alone whereas a moderate improvement was observed in terms of specificity (+9 %). The overall combination gain was 36 % as expressed using the NRI. aBMD and TBS combination decrease significantly the number of subjects needed to diagnose from 7 for aBMD alone to 2. In a multi-centre Eastern European cohort, we have shown that the use of TBS in addition to the aBMD permit to reclassified correctly more than one-third of the overall subjects. Furthermore, the number of subjects needed to diagnose fell to 2 subjects. Economical studies have to be performed to evaluate the gain induced by the use of TBS for the healthcare system.     

Sleep homeostasis and cortical synchronization: III. A high-density EEG study of sleep slow waves in humans

STUDY OBJECTIVES: The mechanisms responsible for the homeostatic decrease of slow-wave activity (SWA, defined in this study as electroencephalogram [EEG] power between 0.5 and 4.0 Hz) during sleep are unknown. In agreement with a recent hypothesis, in the first of 3 companion papers, large-scale computer simulations of the sleeping thalamocortical system showed that a decrease in cortical synaptic strength is sufficient to account for the decline in SWA. In the model, the reduction in SWA was accompanied by decreased incidence of high-amplitude slow waves, decreased wave slopes, and increased number of waves with multiple peaks. In a second companion paper in the rat, local field potential recordings during early and late sleep confirmed the predictions of the model. Here, we investigated the model's predictions in humans by using all-night high-density (hd)-EEG recordings to explore slow-wave parameters over the entire cortical mantle. DESIGN: 256-channel EEG recordings in humans over the course of an entire night's sleep. SETTING: Sound-attenuated sleep research room PATIENTS OR PARTICIPANTS: Seven healthy male subjects INTERVENTIONS: N/A. MEASUREMENTS AND RESULTS: During late sleep (non-rapid eye movement [NREM] episodes 3 and 4, toward morning), when compared with early sleep (NREM sleep episodes 1 and 2, at the beginning of the night), the analysis revealed (1) reduced SWA, (2) fewer large-amplitude slow waves, (3) decreased wave slopes, (4) more frequent multipeak waves. The decrease in slope between early and late sleep was present even when waves were directly matched by wave amplitude and slow-wave power in the background EEG. Finally, hd-EEG showed that multipeak waves have multiple cortical origins. CONCLUSIONS: In the human EEG, the decline of SWA during sleep is accompanied by changes in slow-wave parameters that were predicted by a computer model simulating a homeostatic reduction of cortical synaptic strength.     

Experimental platform utilising melting curve technology for detection of mutations in <Emphasis Type="Italic">Mycobacterium tuberculosis</Emphasis> isolates

Tuberculosis (TB) remains one of the most deadly infections with approximately a quarter of cases not being identified and/or treated mainly due to a lack of resources. Rapid detection of TB or drug-resistant TB enables timely adequate treatment and is a cornerstone of effective TB management. We evaluated the analytical performance of a single-tube assay for multidrug-resistant TB (MDR-TB) on an experimental platform utilising RT-PCR and melting curve analysis that could potentially be operated as a point-of-care (PoC) test in resource-constrained settings with a high burden of TB. Firstly, we developed and evaluated the prototype MDR-TB assay using specimens extracted from well-characterised TB isolates with a variety of distinct rifampicin and isoniazid resistance conferring mutations and nontuberculous Mycobacteria (NTM) strains. Secondly, we validated the experimental platform using 98 clinical sputum samples from pulmonary TB patients collected in high MDR-TB settings. The sensitivity of the platform for TB detection in clinical specimens was 75% for smear-negative and 92.6% for smear-positive sputum samples. The sensitivity of detection for rifampicin and isoniazid resistance was 88.9 and 96.0% and specificity was 87.5 and 100%, respectively. Observed limitations in sensitivity and specificity could be resolved by adjusting the sample preparation methodology and melting curve recognition algorithm. Overall technology could be considered a promising PoC methodology especially in resource-constrained settings based on its combined accuracy, convenience, simplicity, speed, and cost characteristics.     

Tumor-targeted bioconjugate based delivery of camptothecin: design, synthesis and in vitro evaluation.

Camptothecin (CPT) presents numerous challenges associated with optimal transport and delivery including variability in clinically observed effects, low target tissue concentrations and severe and unpredictable toxicity. The objective of the present study was to optimize the delivery of CPT by targeting it to cancer cells using an endogenous receptor system. A novel CPT bioconjugate was synthesized using carbodiimide chemistry with a linear poly(ethylene glycol) (PEG) and amino acid glycine as the spacer and linker respectively. Folic acid was used as the targeting ligand to take advantage of folate receptor mediated endocytosis. The bioconjugate was extensively characterized using MALDI, proton NMR, FT-IR and amino acid analysis. Furthermore, the bioconjugate was evaluated in vitro for specific targeting to folate receptor-expressing KB cells, a human nasopharyngeal carcinoma. Finally, the delivery system was evaluated for cytotoxicity using a MTT based assay. The results indicate significantly higher efficacy of the bioconjugate in comparison to CPT. A control conjugate without PEG demonstrated no improvement in efficacy over untargeted CPT emphasizing the importance of spacer between the anticancer compounds and targeting moiety. This bioconjugate represents the 'first-in-series' of targeted bioconjugates and serves as prototype for improving tumor cell concentration and efficacy.     

Lab-on-Chip-Based Platform for Fast Molecular Diagnosis of Multidrug-Resistant Tuberculosis

We evaluated the performance of the molecular lab-on-chip-based VerePLEX Biosystem for detection of multidrug-resistant tuberculosis (MDR-TB), obtaining a diagnostic accuracy of more than 97.8% compared to sequencing and MTBDRplus assay for Mycobacterium tuberculosis complex and rifampin and isoniazid resistance detection on clinical isolates and smear-positive specimens. The speed, user-friendly interface, and versatility make it suitable for routine laboratory use.Multidrug-resistant tuberculosis (MDR-TB) requires long and expensive treatment and often results in poor clinical outcome in both low- and high-income countries (1, 2). The World Health Organization (WHO) has endorsed specific molecular diagnostics to improve fast diagnosis of MDR-TB (3,–5). However, the genotypic diversity and geographical distribution of Mycobacterium tuberculosis complex (MTBC), together with the inability to provide appropriate interpretation of silent mutations and the limited versatility are some of the restraints undermining the effectiveness of the current tools on a global scale (6,–13).In the present study, we evaluated a lab-on-chip (LoC) device, developed by STMicroelectronics (Geneva, Switzerland) and marketed by Veredus Laboratories (Republic of Singapore) as the VerePLEX Biosystem, for the diagnosis of MDR-TB and detection of common nontuberculous mycobacteria (NTM). The molecular assay was evaluated on both clinical isolates and direct specimens in low- and high-burden settings.We tested 91 MTBC isolates (see Table S1 in the supplemental material) harboring different patterns of mutations in rpoB, katG, and inhA genes to evaluate the probes on the array listed in 14). An additional 116 MTBC culture-negative specimens were included in the analysis. DNA from isolates and specimens was extracted by thermal lysis and sonication as described elsewhere (15). Phenotypic drug susceptibility testing (DST) for rifampin (RIF) and isoniazid (INH) was performed according to international recommendations (16). Some of the specimens were tested in a representative high-burden setting in Uganda (Nsambya Hospital, Kampala, Uganda), by trained staff.

TABLE 1

Probes spotted onto the array and targeted mycobacterial species and MDR-TB targets included in the assay

Multiple Plasmodium falciparum Erythrocyte Membrane Protein 1 Variants per Genome Can Bind IgM via Its Fc Fragment Fcμ

The Plasmodium falciparum erythrocyte membrane protein 1 (PfEMP1) adhesive proteins expressed on the surfaces of infected erythrocytes (IEs) are of key importance in the pathogenesis of P. falciparum malaria. Several structurally and functionally defined PfEMP1 types have been associated with severe clinical manifestations, such as cerebral malaria in children and placental malaria in pregnant women. PfEMP1 that can bind the Fc part of IgM (Fcμ) characterizes one such type, although the functional significance of this IgM binding to PfEMP1 remains unclear. In this study, we report the identification and functional analysis of five IgM-binding PfEMP1 proteins encoded by P. falciparum NF54. In addition to the VAR2CSA-type PFL0030c protein, already known to bind Fcμ and to mediate chondroitin sulfate A (CSA)-specific adhesion of IEs in the placenta, we found four PfEMP1 proteins not previously known to bind IgM this way. Although they all contained Duffy binding-like ε (DBLε) domains similar to those in VAR2CSA-type PfEMP1, they did not mediate IE adhesion to CSA, and IgM binding did not shield IEs from phagocytosis of IgG-opsonized IEs. In this way, these new IgM-binding PfEMP1 proteins resemble the rosette-mediating and IgM-binding PfEMP1 HB3VAR06, but none of them mediated formation of rosettes. We could map the capacity for Fc-specific IgM binding to DBLε domains near the C terminus for three of the four PfEMP1 proteins tested. Our study provides new evidence regarding Fc-dependent binding of IgM to PfEMP1, which appears to be a common and multifunctional phenotype.     

Fleece bound sealing prevents pleural adhesions

This study assessed the value of haemostatic fleece (HF) in prevention of pleural adhesions in an experimental animal model. Forty rats were randomly assigned to four equal groups and underwent bilateral thoracotomy. In Group 1 standardized defects of 5 mm were generated in the visceral and the opposite parietal pleura without further coverage. In Group 2 a 5-mm piece of HF (TachoSil) was applied onto the intact pleura. In Group 3 a standardized pleural defect was completely covered by HF. The same kind of defect was only partially covered by HF in group 4 animals. Autopsy at 6 weeks (n=5, each group) revealed the fleece widely unchanged and covered by a smooth serous membrane. After 12 weeks (n=5, each group) the fleece had been completely resorbed. Histological studies revealed the area of the defect covered by regular mesothelium. In all animals pleural adhesions were detected only in the area without fleece coverage. In this experimental model HF prevented the development of pleural adhesions. This property may have clinical impact in patients with some probability of re-thoracotomy enabling to reduce the risk of pleural adhesions significantly.     

Limited Capability for Testing Mycobacterium tuberculosis for Susceptibility to New Drugs

We surveyed availability of phenotypic drug susceptibility testing for drug-resistant Mycobacterium tuberculosis in Europe. Of 27 laboratories, 17 tested for linezolid, 11 for clofazimine, 9 for bedaquiline, and 6 for delamanid during 2019. Our findings indicate that testing capacity for newer and repurposed tuberculosis drugs exists, but its availability is limited.     

Biotropic liquid crystal phase transformations in cellulose-producing bacterial communities

Biological functionality is often enabled by a fascinating variety of physical phenomena that emerge from orientational order of building blocks, a defining property of nematic liquid crystals that is also pervasive in nature. Out-of-equilibrium, “living” analogs of these technological materials are found in biological embodiments ranging from myelin sheath of neurons to extracellular matrices of bacterial biofilms and cuticles of beetles. However, physical underpinnings behind manifestations of orientational order in biological systems often remain unexplored. For example, while nematiclike birefringent domains of biofilms are found in many bacterial systems, the physics behind their formation is rarely known. Here, using cellulose-synthesizing Acetobacter xylinum bacteria, we reveal how biological activity leads to orientational ordering in fluid and gel analogs of these soft matter systems, both in water and on solid agar, with a topological defect found between the domains. Furthermore, the nutrient feeding direction plays a role like that of rubbing of confining surfaces in conventional liquid crystals, turning polydomain organization within the biofilms into a birefringent monocrystal-like order of both the extracellular matrix and the rod-like bacteria within it. We probe evolution of scalar orientational order parameters of cellulose nanofibers and bacteria associated with fluid-gel and isotropic-nematic transformations, showing how highly ordered active nematic fluids and gels evolve with time during biological-activity-driven, disorder-order transformation. With fluid and soft-gel nematics observed in a certain range of biological activity, this mesophase-exhibiting system is dubbed “biotropic,” analogously to thermotropic nematics that exhibit solely orientational order within a temperature range, promising technological and fundamental-science applications.

Many biological substances, like cholesterol, lipids, proteins, nucleic acids, microtubules, and viruses are known for their liquid crystalline (LC) phase behavior in either pure form or when in aqueous solutions and suspensions (1). The very first documented observations of LCs from well over a century ago utilized substances like derivatives of cholesterol extracted from living organisms (2, 3). Moreover, created within natural processes, LC-like structural organizations in tissues and key components of biological systems, like membranes, tend to endow complex functionality that remains superior to that of man-made materials (4, 5). Recognizing these important roles of LC-like organization, a biologist, D. Dervichian, once wrote “Liquid crystals stand between the isotropic liquid phase and the strongly organized state. Life stands between complete disorder, which is death and complete rigidity which is death again” (6). Understanding and recreating these LC-like nature’s elegant designs can revolutionize our ability of cost-effective mass-production of “living” materials and devices with attractive properties like low-cost, green manufacturing, self-healing and regeneration. However, how nature creates LC-like structures within out-of-equilibrium processes remains poorly understood despite of their diverse and ubiquitous observations (1–6).A particularly interesting form of a natural LC-like organization relates to bacterial biofilms, where birefringent domains of extracellular matrix and orientationally ordered bacteria have been reported for many microbiological systems (7–21). For example, Acetobacter xylinum (A. xylinum) bacteria biosynthesize cellulose filaments which can be organized in an orderly manner at the micrometer scales while exhibiting randomly aligned domains on larger scales within the three-dimensional (3D) reticulated biofilm (22–26). Limitations in controlling morphology and order within such biofilms hinder potential applications of bacterial cellulose-based materials (15–19). Here, we elucidate how a natural biological process leads to a spontaneous LC order of both cellulose nanofibers, which form an anisotropic extracellular matrix with strongly pronounced birefringence, and rod-like A. xylinum bacteria within a mesostructured active nematic system that can take both polydomain and monodomain structural forms. We probe orientationally ordered active nematic states in both aqueous environment and atop of solid agar surfaces. Using video-microscopy within different optical imaging modalities, we document the spontaneous emergence of nematic ordering in the course of biological activity. We find that directional feedings (and separately synthesized nematic hosts of colloidal nanocellulose used in our test experiments) break symmetry and lead to monodomain-like active nematic organizations, which we then use to quantitatively probe how orientational order of cellulose nanofibers and bacteria evolves with time. Holographic laser tweezers reveal onsets and details of formation of hydrogels with anisotropic networks of nanofibers. Characterization of distinct types of motions associated with A. xylinum’s nanocellulose production, as well as the estimation of order parameters (27, 28) in nematic and isotropic fluids and gels, reveals an active nematic system with disorder-to-order transformations driven by biological activity. Further, we show that such bacteria-made hydrogels can be readily converted to anisotropic aerogels and discuss how they may find diverse technological uses ranging from thermal insulation to scaffolding of biological tissue within LC-like matrices.