Practical context of enzymatic treatment for wound healing: A secreted protease approach (Review)

Skin wounds have been extensively studied as their healing represents a critical step towards achieving homeostasis following a traumatic event. Dependent on the severity of the damage, wounds are categorized as either acute or chronic. To date, chronic wounds have the highest economic impact as long term increases wound care costs. Chronic wounds affect 6.5 million patients in the United States with an annual estimated expense of $25 billion for the health care system. Among wound treatment categories, active wound care represents the fastest‑growing category due to its specific actions and lower costs. Within this category, proteases from various sources have been used as successful agents in debridement wound care. The wound healing process is predominantly mediated by matrix metalloproteinases (MMPs) that, when dysregulated, result in defective wound healing. Therapeutic activity has been described for animal secretions including fish epithelial mucus, maggot secretory products and snake venom, which contain secreted proteases (SPs). No further alternatives for use, sources or types of proteases used for wound healing have been found in the literature to date. Through the present review, the context of enzymatic wound care alternatives will be discussed. In addition, substrate homology of SPs and human MMPs will be compared and contrasted. The purpose of these discussions is to identify and propose the stages of wound healing in which SPs may be used as therapeutic agents to improve the wound healing process.Fil: Avila Rodríguez, María Isabela. Instituto Tecnologico de Monterrey. Escuela de Ingenieria y Ciencias.; MéxicoFil: Meléndez Martínez, David. Instituto Tecnologico de Monterrey. Escuela de Ingenieria y Ciencias.; MéxicoFil: Licona Cassani, Cuauhtemoc. Instituto Tecnologico de Monterrey. Escuela de Ingenieria y Ciencias.; MéxicoFil: Aguilar Yañez, José Manuel. Instituto Tecnologico de Monterrey. Escuela de Ingenieria y Ciencias.; México. Scicore Medical SAPI de CV; MéxicoFil: Benavides, Jorge. Instituto Tecnologico de Monterrey. Escuela de Ingenieria y Ciencias.; MéxicoFil: Sanchez, Mirna Lorena. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - La Plata. Instituto Multidisciplinario de Biología Celular. Provincia de Buenos Aires. Gobernación. Comisión de Investigaciones Científicas. Instituto Multidisciplinario de Biología Celular. Universidad Nacional de La Plata. Instituto Multidisciplinario de Biología Celular; Argentin

Different analysis strategies of 16S rRNA gene data from rodent studies generate contrasting views of gut bacterial communities associated with diet, health and obesity

Background One of the main functions of diet is to nurture the gut microbiota and this relationship affects the health of the host. However, different analysis strategies can generate different views on the relative abundance of each microbial taxon, which can affect our conclusions about the significance of diet to gut health in lean and obese subjects. Here we explored the impact of using different analysis strategies to study the gut microbiota in a context of diet, health and obesity. Methods Over 15 million 16S rRNA gene sequences from published studies involving dietary interventions in obese laboratory rodents were analyzed. Three strategies were used to assign the 16S sequences to Operational Taxonomic Units (OTUs) based on the GreenGenes reference OTU sequence files clustered at 97% and 99% similarity. Results Different strategies to select OTUs influenced the relative abundance of all bacterial taxa, but the magnitude of this phenomenon showed a strong study effect. Different taxa showed up to 20% difference in relative abundance within the same study, depending on the analysis strategy. Very few OTUs were shared among the samples. ANOSIM test on unweighted UniFrac distances showed that study, sequencing technique, animal model, and dietary treatment (in that order) were the most important factors explaining the differences in bacterial communities. Except for obesity status, the contribution of diet and other factors to explain the variability in bacterial communities was lower when using weighted UniFrac distances. Predicted functional profile and high-level phenotypes of the microbiota showed that each study was associated with unique features and patterns. Conclusions The results confirm previous findings showing a strong study effect on gut microbial composition and raise concerns about the impact of analysis strategies on the membership and composition of the gut microbiota. This study may be helpful to guide future research aiming to investigate the relationship between diet, health, and the gut microbiota

Comparative Economic Analysis Between Endogenous and Recombinant Production of Hyaluronic Acid

Hyaluronic acid (HA) is a biopolymer with a wide range of applications, mainly in the cosmetic and pharmaceutical sectors. Typical industrial-scale production utilizes organisms that generate HA during their developmental cycle, such as Streptococcus equi sub. zooepidemicus. However, a significant disadvantage of using Streptococcus equi sub. zooepidemicus is that it is a zoonotic pathogen, which use at industrial scale can create several risks. This creates opportunities for heterologous, or recombinant, production of HA. At an industrial scale, the recovery and purification of HA follow a series of precipitation and filtration steps. Current recombinant approaches are developing promising alternatives, although their industrial implementation has yet to be adequately assessed. The present study aims to create a theoretical framework to forecast the advantages and disadvantages of endogenous and recombinant strains in production with the same downstream strategy. The analyses included a selection of the best cost-related recombinant and endogenous production strategies, followed by a sensitivity analysis of different production variables in order to identify the three most critical parameters. Then, all variables were analyzed by varying them simultaneously and employing multiple linear regression. Results indicate that, regardless of HA source, production titer, recovery yield and bioreactor scale are the parameters that affect production costs the most. Current results indicate that recombinant production needs to improve current titer at least 2-fold in order to compete with costs of endogenous production. This study serves as a platform to inform decision-making for future developments and improvements in the recombinant production of HA

Primary recovery of hyaluronic acid produced in Streptococcus equi subsp. zooepidemicus using PEG-citrate aqueous two-phase systems

Given its biocompatibility, rheological, and physiological properties, hyaluronic acid (HA) has become a biomaterial of increasing interest with multiple applications in medicine and cosmetics. In recent decades, microbial fermentations have become an important source for the industrial production of HA. However, due to its final applications, microbial HA must undergo critical and long purification processes to ensure clinical and cosmetic grade purity. Aqueous two-phase systems (ATPS) have proven to be an efficient technique for the primary recovery of high-value biomolecules. Nevertheless, their implementation in HA downstream processing has been practically unexplored. In this work, polyethylene glycol (PEG)–citrate ATPS were used for the first time for the primary recovery of HA produced with an engineered strain of Streptococcus equi subsp. zooepidemicus. The effects of PEG molecular weight (MW), tie-line length (TLL), volume ratio (VR), and sample load on HA recovery and purity were studied with a clarified fermentation broth as feed material. HA was recovered in the salt-rich bottom phase, and its recovery increased when a PEG MW of 8000 g mol−1 was used. Lower VR values (0.38) favoured HA recovery, whereas purity was enhanced by a high VR (3.50). Meanwhile, sample load had a negative impact on both recovery and purity. The ATPS with the best performance was PEG 8000 g mol−1, TLL 43% (w/w), and VR 3.50, showing 79.4% HA recovery and 74.5% purity. This study demonstrated for the first time the potential of PEG–citrate ATPS as an effective primary recovery strategy for the downstream process of microbial HA

Spontaneous Bacterial Peritonitis: Physiopathological Mechanism and Clinical Manifestations

Changes in intestinal permeability have been determined to influence secondary inflammatory reactions and clinical manifestations such as spontaneous bacterial peritonitis (SBP) secondary to cirrhosis. As of yet, no in-depth exploration of the changes in the microbiota and how this influences cirrhosis to differ from clinically more severe cases than others has not begun. However, at the level of pathophysiological mechanism, it must be taken into account that due to the abuse of substances such as alcohol and chronic fatty liver disease, changes in the bacterial composition and intestinal permeability are induced. This set of changes in the bacterial composition (microbiome) and modification of the intestinal permeability could be related to the presence of ascites and spontaneous peritonitis secondary to cirrhosis, being of relevance the knowledge of the mechanisms underlying this phenomenon, as well as clinical manifestation. Prophylaxis and antibiotic treatment of SBP requires clinical knowledge for the treatment decisions based mainly on the presence of ascitic fluid, accompanied of risk factors, laboratory indexes such as PMN count and culture results, in order to determine the kind of molecule that will help to the SBP recovery or to amelioration symptoms, always taking care of not exceed the antibiotic consumption and restoring the microbiome imbalance

microbeMASST: A Taxonomically-informed Mass Spectrometry Search Tool for Microbial Metabolomics Data

microbeMASST, a taxonomically informed mass spectrometry (MS) search tool, tackles limited microbial metabolite annotation in untargeted metabolomics experiments. Leveraging a curated database of >60,000 microbial monocultures, users can search known and unknown MS/MS spectra and link them to their respective microbial producers via MS/MS fragmentation patterns. Identification of microbe-derived metabolites and relative producers without a priori knowledge will vastly enhance the understanding of microorganisms’ role in ecology and human health

A Taxonomically-informed Mass Spectrometry Search Tool for Microbial Metabolomics Data

MicrobeMASST, a taxonomically-informed mass spectrometry (MS) search tool, tackles limited microbial metabolite annotation in untargeted metabolomics experiments. Leveraging a curated database of >60,000 microbial monocultures, users can search known and unknown MS/MS spectra and link them to their respective microbial producers via MS/MS fragmentation patterns. Identification of microbial-derived metabolites and relative producers, without a priori knowledge, will vastly enhance the understanding of microorganisms’ role in ecology and human health

Temporal dynamics of the Saccharopolyspora erythraea phosphoproteome

Actinomycetes undergo a dramatic reorganization of metabolic and cellular machinery during a brief period of growth arrest (metabolic switch) preceding mycelia differentiation and the onset of secondary metabolite biosynthesis. This study explores the role of phosphorylation in coordinating the metabolic switch in the industrial actinomycete Saccharopolyspora erythraea. A total of 109 phosphopeptides from 88 proteins were detected across a 150-h fermentation using open-profile two-dimensional LC-MS proteomics and TiO2 enrichment. Quantitative analysis of the phosphopeptides and their unphosphorylated cognates was possible for 20 pairs that also displayed constant total protein expression. Enzymes from central carbon metabolism such as putative acetyl-coenzyme A carboxylase, isocitrate lyase, and 2-oxoglutarate dehydrogenase changed dramatically in the degree of phosphorylation during the stationary phase, suggesting metabolic rearrangement for the reutilization of substrates and the production of polyketide precursors. In addition, an enzyme involved in cellular response to environmental stress, trypsin-like serine protease (SACE_6340/NC_009142_6216), decreased in phosphorylation during the growth arrest stage. More important, enzymes related to the regulation of protein synthesis underwent rapid phosphorylation changes during this stage. Whereas the degree of phosphorylation of ribonuclease Rne/Rng (SACE_1406/NC_009142_1388) increased during the metabolic switch, that of two ribosomal proteins, S6 (SACE_7351/NC_009142_7233) and S32 (SACE_6101/NC_009142_5981), dramatically decreased during this stage of the fermentation, supporting the hypothesis that ribosome subpopulations differentially regulate translation before and after the metabolic switch. Overall, we show the great potential of phosphoproteomic studies to explain microbial physiology and specifically provide evidence of dynamic protein phosphorylation events across the developmental cycle of actinomycetes

Re-annotation of the Saccharopolyspora erythraea genome using a systems biology approach

Background: Accurate bacterial genome annotations provide a framework to understanding cellular functions, behavior and pathogenicity and are essential for metabolic engineering. Annotations based only on in silico predictions are inaccurate, particularly for large, high G + C content genomes due to the lack of similarities in gene length and gene organization to model organisms