Any multicellular organism during its life is involved in relatively stable interactions with microorganisms. The organism and its microbiome make up a holobiont, possessing a unique set of ...characteristics and evolving as a whole system. This study aimed to evaluate the degree of the conservativeness of microbiomes associated with intertidal gastropods. We studied the composition and the geographic and phylogenetic variability of the gut and body surface microbiomes of five closely related sympatric Littorina (Neritrema) spp. and a more distant species, L. littorea, from the sister subgenus Littorina (Littorina). Although snail-associated microbiomes included many lineages (207-603), they were dominated by a small number of OTUs of the genera Psychromonas, Vibrio, and Psychrilyobacter. The geographic variability was greater than the interspecific differences at the same collection site. While the microbiomes of the six Littorina spp. did not differ at the high taxonomic level, the OTU composition differed between groups of cryptic species and subgenera. A few species-specific OTUs were detected within the collection sites; notably, such OTUs never dominated microbiomes. We conclude that the composition of the high-rank taxa of the associated microbiome ("scaffolding enterotype") is more evolutionarily conserved than the composition of the low-rank individual OTUs, which may be site- and / or species-specific.
Fibrobacter succinogenes is an important member of the rumen microbial community that converts plant biomass into nutrients usable by its host. This bacterium, which is also one of only two ...cultivated species in its phylum, is an efficient and prolific degrader of cellulose. Specifically, it has a particularly high activity against crystalline cellulose that requires close physical contact with this substrate. However, unlike other known cellulolytic microbes, it does not degrade cellulose using a cellulosome or by producing high extracellular titers of cellulase enzymes. To better understand the biology of F. succinogenes, we sequenced the genome of the type strain S85 to completion. A total of 3,085 open reading frames were predicted from its 3.84 Mbp genome. Analysis of sequences predicted to encode for carbohydrate-degrading enzymes revealed an unusually high number of genes that were classified into 49 different families of glycoside hydrolases, carbohydrate binding modules (CBMs), carbohydrate esterases, and polysaccharide lyases. Of the 31 identified cellulases, none contain CBMs in families 1, 2, and 3, typically associated with crystalline cellulose degradation. Polysaccharide hydrolysis and utilization assays showed that F. succinogenes was able to hydrolyze a number of polysaccharides, but could only utilize the hydrolytic products of cellulose. This suggests that F. succinogenes uses its array of hemicellulose-degrading enzymes to remove hemicelluloses to gain access to cellulose. This is reflected in its genome, as F. succinogenes lacks many of the genes necessary to transport and metabolize the hydrolytic products of non-cellulose polysaccharides. The F. succinogenes genome reveals a bacterium that specializes in cellulose as its sole energy source, and provides insight into a novel strategy for cellulose degradation.
Poly-ε-caprolactone (PCL) is a biodegradable polymer used in regenerative medicine. Mesenchymal stem cells (MSCs) play an important role in the regeneration of different tissues. The hydrophobicity ...and neutrality of a PCL surface reduce MSCs' adhesion and proliferation. In this study, PCL films were treated with arginine to improve surface hydrophilicity. The influences of arginine concentration, temperature, and solvent on PCL surface properties were investigated. PCL films treated with a solution of arginine in isopropyl alcohol were found to have the maximum number of amino groups. The greatest number of cells, 2 h after seeding, adhered to such films. It was shown that amino groups affect the interaction of cells with a modified surface and the hydrolysis reaction after treatment with isopropyl alcohol promotes the formation of adhesive focal contacts. Hence, our results illustrate that functional groups on the PCL surface after arginine solution treatment regulate MSC adhesion and focal contact formation.
Background. Retina is the highest oxygen-demanding and vascularized tissue in the body. Retinal development and function require proper vascularization and blood vessel function and integrity. Dll4 ...is most prominently expressed in the endothelium of angiogenic blood vessels and in quiescent arteries and capillaries in all tissues and organs of the mammalian species, and it is the key regulator of blood vessel sprouting. Results. Dll4 is a transmembrane protein that acts as a ligand for Notch receptors 1 and 4. Genetic deletion of Dll4 causes severe abnormalities in embryonic and postnatal vascular development. Deletion of even a single Dll4 allele results in almost complete embryonic lethality due to severe vascular abnormalities, the phenomenon called haploinsufficiency indicating the critical role of Dll4/Notch in vascular development. Dll4/Notch pathway interplays at multiple levels with other signaling pathways including VEGF, Wnt/Fzd, and genes controlling vascular toning. Multiple studies of the effects of Dll4 inhibition were performed in the developing retina to elucidate the key functions of Dll4 in normal and pathological angiogenesis. Several genetic approaches and therapeutic molecules were tested to evaluate the biological and therapeutic effects of acute and prolonged Dll4 inhibition in the eye and oncology. Conclusions. All current studies demonstrated that Dll4 controls blood vessel sprouting, growth, and remodeling in normal and pathological conditions as well as arterial-venous differentiation. Genetic and therapeutic Dll4 modulation studies show that Dll4 inhibition can promote blood vessel sprouting and might be useful to stimulate vessel growth in the ischemic retina and Dll4 is the key modulator of the postangiogenic vascular remodeling that ultimately defines vascular patterning.
The modeling of neuropathology on induced neurons obtained by cell reprogramming technologies can fill a gap between clinical trials and studies on model organisms for the development of treatment ...strategies for neurodegenerative diseases. Patient-specific models based on patients' cells play an important role in such studies. There are two ways to obtain induced neuronal cells. One is based on induced pluripotent stem cells. The other is based on direct reprogramming, which allows us to obtain mature neuronal cells from adult somatic cells, such as dermal fibroblasts. Moreover, the latter method makes it possible to better preserve the age-related aspects of neuropathology, which is valuable for diseases that occur with age. However, direct methods of reprogramming have a significant drawback associated with low cell viability during procedures. Furthermore, the number of reprogrammable neurons available for morphological and functional studies is limited by the initial number of somatic cells. In this article, we propose modifications of a previously developed direct reprogramming method, based on the combination of microRNA and transcription factors, which allowed us to obtain a population of functionally active induced striatal neurons (iSNs) with a high efficiency. We also overcame the problem of the presence of multinucleated neurons associated with the cellular division of starting fibroblasts. Synchronization cells in the G1 phase increased the homogeneity of the fibroblast population, increased the survival rate of induced neurons, and eliminated the presence of multinucleated cells at the end of the reprogramming procedure. We have demonstrated that iSNs are functionally active and able to form synaptic connections in co-cultures with mouse cortical neurons. The proposed modifications can also be used to obtain a population of other induced neuronal types, such as motor and dopaminergic ones, by selecting transcription factors that determine differentiation into a region-specific neuron.
Secretome of multipotent mesenchymal stromal cells (MSCs) is actively used in biomedical applications such as alveolar bone regeneration, treatment of cardiovascular disease, and neurodegenerative ...disorders. Nevertheless, hMSCs have low proliferative potential and production of the industrial quantity of their secretome might be challenging. Human fetal multipotent mesenchymal stromal cells (FetMSCs) isolated from early human embryo bone marrow are easy to expand and might be a potential source for pharmaceutical substances production based on their secretome. However, the secretome of FetMSCs was not previously analyzed. Here, we describe the secretome of FetMSCs using LC-MALDI shotgun proteomics. We identified 236 proteins. Functional annotation of the identified proteins revealed their involvement in angiogenesis, ossification, regulation of apoptosis, and immune response processes, which made it promising for biomedical applications. The proteins identified in the FetMSCs secretome are involved in the same biological processes as proteins from previously described adult hMSCs secretomes. Nevertheless, many of the common hMSCs secretome components (such as VEGF, FGF, Wnt and TGF-β) have not been identified in the FetMSCs secretome.
Ocular surface reconstruction is essential for treating corneal epithelial defects and vision recovery. Stem cell-based therapy demonstrates promising results but requires further research to ...elucidate stem cell survival, growth, and differentiation after transplantation in vivo. This study examined the corneal reconstruction promoted by EGFP-labeled limbal mesenchymal stem cells (L-MSCs-EGFP) and their fate after transplantation. EGFP labeling allowed us to evaluate the migration and survival rates of the transferred cells. L-MSCs-EGFP seeded onto decellularized human amniotic membrane (dHAM) were transplanted into rabbits with a modeled limbal stem cell deficiency. The localization and viability of the transplanted cells in animal tissue were analyzed using histology, immunohistochemistry, and confocal microscopy up to 3 months after transplantation. EGFP-labeled cells remained viable for the first 14 days after transplantation. By the 90th day, epithelialization of the rabbit corneas reached 90%, but the presence of viable labeled cells was not observed within the newly formed epithelium. Although labeled cells demonstrated low survivability in host tissue, the squamous corneal-like epithelium was partially restored by the 30th day after transplantation of the tissue-engineered graft. Overall, this study paves the way for further optimization of transplantation conditions and studying the mechanisms of corneal tissue restoration.
From the standpoints of both basic research and biotechnology, there is considerable interest in reaching a clearer understanding of the diversity of biological mechanisms employed during ...lignocellulose degradation. Globally, termites are an extremely successful group of wood-degrading organisms1 and are therefore important both for their roles in carbon turnover in the environment and as potential sources of biochemical catalysts for efforts aimed at converting wood into biofuels. Only recently have data supported any direct role for the symbiotic bacteria in the gut of the termite in cellulose and xylan hydrolysis. Here we use a metagenomic analysis of the bacterial community resident in the hindgut paunch of a wood-feeding 'higher' Nasutitermes species (which do not contain cellulose-fermenting protozoa) to show the presence of a large, diverse set of bacterial genes for cellulose and xylan hydrolysis. Many of these genes were expressed in vivo or had cellulase activity in vitro, and further analyses implicate spirochete and fibrobacter species in gut lignocellulose degradation. New insights into other important symbiotic functions including H2 metabolism, CO2-reductive acetogenesis and N2 fixation are also provided by this first system-wide gene analysis of a microbial community specialized towards plant lignocellulose degradation. Our results underscore how complex even a 1-l environment can be.
Microbes can play a prominent role in the evolution of their hosts, facilitating adaptation to various environments and promoting ecological divergence. The Wave and Crab ecotypes of the intertidal ...snail Littorina saxatilis is an evolutionary model of rapid and repeated adaptation to environmental gradients. While patterns of genomic divergence of the Littorina ecotypes along the shore gradients have been extensively studied, their microbiomes have been so far overlooked. The aim of the present study is to start filling this gap by comparing gut microbiome composition of the Wave and Crab ecotypes using metabarcoding approach. Since Littorina snails are micro‐grazers feeding on the intertidal biofilm, we also compare biofilm composition (i.e. typical snail diet) in the crab and wave habitats. In the results, we found that bacterial and eukaryotic biofilm composition varies between the typical habitats of the ecotypes. Further, the snail gut bacteriome was different from outer environments, being dominated by Gammaproteobacteria, Fusobacteria, Bacteroidia and Alphaproteobacteria. There were clear differences in the gut bacterial communities between the Crab and the Wave ecotypes as well as between the Wave ecotype snails from the low and high shores. These differences were both observed in the abundances and in the presence of different bacteria, as well as at different taxonomic level, from bacterial OTU's to families. Altogether, our first insights show that Littorina snails and their associated bacteria are a promising marine system to study co‐evolution of the microbes and their hosts, which can help us to predict the future for wild species in the face of rapidly changing marine environments.
Researchers are still working on the development of models that facilitate the accurate estimation of acoustic cavitation threshold. In this paper, we have analyzed the possibility of using the ...incubation time criterion to calculate the threshold of the onset of acoustic cavitation depending on the ultrasound frequency, hydrostatic pressure, and temperature of a liquid. This criterion has been successfully used by earlier studies to calculate the dynamic strength of solids and has recently been proposed in an adapted version for calculating the cavitation threshold. The analysis is carried out for various experimental data for water presented in the literature. Although the criterion assumes the use of macroparameters of a liquid, we also considered the possibility of taking into account the size of cavitation nuclei and its influence on the calculation result. We compared the results of cavitation threshold calculations done using the incubation time criterion of cavitation and the classical nucleation theory. Our results showed that the incubation time criterion more qualitatively models the results of experiments using only three parameters of the liquid. We then discussed a possible relationship between the parameters of the two approaches. The results of our study showed that the criterion under consideration has a good potential and can be conveniently used for applications where there are special requirements for ultrasound parameters, maximum negative pressure, and liquid temperature.
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