Stella Maris de Freitas Lima

Shedding some light over the floral metabolism by arum lily (Zantedeschia aethiopica) spathe de novo transcriptome assembly.

Zantedeschia aethiopica is an evergreen perennial plant cultivated worldwide and commonly used for ornamental and medicinal purposes including the treatment of bacterial infections. However, the current understanding of molecular and physiological mechanisms in this plant is limited, in comparison to other non-model plants. In order to improve understanding of the biology of this botanical species, RNA-Seq technology was used for transcriptome assembly and characterization. Following Z. aethiopica spathe tissue RNA extraction, high-throughput RNA sequencing was performed with the aim of obtaining both abundant and rare transcript data. Functional profiling based on KEGG Orthology (KO) analysis highlighted contigs that were involved predominantly in genetic information (37%) and metabolism (34%) processes. Predicted proteins involved in the plant circadian system, hormone signal transduction, secondary metabolism and basal immunity are described here. In silico screening of the transcriptome data set for antimicrobial peptide (AMP) –encoding sequences was also carried out and three lipid transfer proteins (LTP) were identified as potential AMPs involved in plant defense. Spathe predicted protein maps were drawn, and suggested that major plant efforts are expended in guaranteeing the maintenance of cell homeostasis, characterized by high investment in carbohydrate, amino acid and energy metabolism as well as in genetic information.

Antimicrobial peptide-based treatment for endodontic infections — Biotechnological innovation in endodontics

The presence/persistence of microorganisms in the pulp and periapical area corresponds to the maintenance of an exacerbated immune response that leads to the start of periradicular bone resorption and its perpetuation. In endodontic treatment, the available intracanal medications do not have all the desirable properties in the context of endodontic infection and apical periodontitis; they need to include not only strong antimicrobial performance but also an immunomodulatory and reparative activity, without host damage. In addition, there are various levels of resistance to root canal medications. Thus, antimicrobial agents that effectively eliminate resistant species in root canals could potentially improve endodontic treatment. In the emergence of new therapies, an increasing number of studies on antimicrobial peptides (AMPs) have been seen over the past few years. AMPs are defense biomolecules produced in response to infection, and they have a wide spectrum of action against many oral microorganisms. There are some studies that correlate peptides and oral infections, including oral peptides, neuropeptides, and bacterial, fish, bovine and synthetic peptides. So far, there are around 120 published studies correlating endodontic microbiota with AMPs but, according to our knowledge, there are no registered patents in the American patent database. There are a considerable number of AMPs that exhibit excellent antimicrobial activity against endodontic microbiota at a small inhibitory concentration and modulate an exacerbated immune response, down-regulating bone resorption. All these reasons indicate the antimicrobial peptide-based endodontic treatment as an emerging and promising option.

Dentistry proteomics: From laboratory development to clinical practice

Despite all the dental information acquired over centuries and the importance of proteome research, the cross‐link between these two areas only emerged around mid‐nineties. Proteomic tools can help dentistry in the identification of risk factors, early diagnosis, prevention, and systematic control that will promote the evolution of treatment in all dentistry specialties. This review mainly focuses on the evolution of dentistry in different specialties based on proteomic research and how these tools can improve knowledge in dentistry. The subjects covered are an overview of proteomics in dentistry, specific information on different fields in dentistry (dental structure, restorative dentistry, endodontics, periodontics, oral pathology, oral surgery, and orthodontics) and future directions. There are many new proteomic technologies that have never been used in dentistry studies and some dentistry areas that have never been explored by proteomic tools. It is expected that a greater integration of these areas will help to understand what is still unknown in oral health and disease. J. Cell. Physiol. 228: 2271–2284, 2013.

NanoUPLC-MS(E) proteomic analysis of osteoclastogenesis downregulation by IL-4.

Bone resorption is an important factor in bone homeostasis and imbalance can cause several diseases. In osteoimmunology, IL-4 has been described as an important factor in promoting M2 macrophage profile. In order to shed some light on the effect of IL-4 on osteoclast precursors in the presence of RANKL, cytokines and nitric oxide (NO) production and the proteomic profile were analyzed. The presence of IL-4 in in vitro osteoclastogenesis provides production of TNF-α, IL-1α, IL-1β, IL-10 and IL-12 at basal cell levels. Regarding NO production, IL-4 was sufficient to increase the basal NO levels. Proteomic analyses identified 877 global proteins. IL-4 in in vitro RANKL-mediated osteoclastogenesis leads to the expression of 118 proteins. The presence of rIL-4 in in vitro rRANKL-mediated-osteoclastogenesis downregulated this process. However, the proteomics findings in the osteoclastogenesis demonstrated a much greater effect on osteoclast precursor cells than on RANKL-differentiated osteoclasts. These results suggest that the main effect of IL-4 in pre-osteoclast cells leads to a M2 macrophage activation, and this probably contributed to a reduction in osteoclastogenesis when both stimuli were used. This study noticed that IL-4 plays an important regulatory role in bone homeostasis due to its suppressive potential of precursor osteoclast cells.

Immune Response Profile against Persistent Endodontic Pathogens Candida albicans and Enterococcus faecalis In Vitro

INTRODUCTION Persistent microorganisms such as Candida albicans and Enterococcus faecalis might be directly related to endodontic treatment failure. The host response to these microorganisms impairs the reestablishment of intraradicular and periradicular health. METHODS The present investigation evaluated the expression of inflammatory mediators produced by RAW 264.7 cells in the presence of heat-killed antigens (HK) C. albicans and E. faecalis. Cultures of RAW cells were stimulated with both antigens in the presence or absence of recombinant interferon (rIFN)-γ. Parameters of cell viability, production of nitric oxide (NO), as well as the synthesis of interleukin (IL)-1α, IL-6, IL-10, IL-12, monocyte chemotactic protein-1, and tumor necrosis factor (TNF)-α were analyzed. RESULTS Results demonstrated that cell viability was especially reduced in antigens and rIFN-γ-stimulated groups. Groups stimulated with HK C. albicans upregulated IL-10 production. Otherwise, the addition of rIFN-γ to HK C. albicans upregulated TNF-α and NO production. Groups stimulated with HK E. faecalis upregulated TNF-α production. HK E. faecalis and rIFN-γ upregulated TNF-α and NO synthesis. The production of other cytokines remained unchanged by all stimuli. CONCLUSIONS Knowledge regarding the host mechanism of response to microorganisms that perpetuate endodontic infection and the periradicular lesions can contribute to optimization of endodontic therapy. The mentioned inflammatory mediators and virulence factors involved in endodontic failure might guide lesion progression and also be targets in the development of disinfectant and immunomodulatory agents.

The effects of glucose concentrations associated with lipopolysaccharide and interferon-gamma stimulus on mediators’ production of RAW 264.7 cells

HighlightsDiabetes mellitus include changes in immune response processes.High glucose in vitro does not change cell viability.High glucose may be sufficient to alter the in vitro mediators’ production.NO production is dose‐dependent in relation to the glucose concentrations. &NA; Diabetes mellitus (DM) is a metabolic disorder that results in the impairment of the metabolism of carbohydrates, proteins and lipids. It can give rise to various complications, mainly caused by chronic exposure of cells to high glucose concentrations, including changes in the immune response processes. The aim of this study was to verify the chemokine and cytokines production profile in the presence of different glucose concentrations and infection/inflammatory stimuli. To this end, cell viability and the production of chemokines, cytokines and nitric oxide (NO) were analyzed in RAW 264.7 cell culture. Results demonstrated that there was no change in cell viability after 6, 24 and 72 h. Different stimuli were unable to modify the monocyte chemoattractant protein (MCP)‐1 and tumor necrosis factor (TNF)‐&agr; production. Groups stimulated with lipopolysaccharides (LPS) and LPS and recombinant interferon (rIFN)‐&ggr; down‐regulated interleukin (IL)‐1&agr;, IL‐10 and IL‐12 and up‐regulated IL‐6 production. NO production maintained a pattern of increase, according to the increase in glucose concentrations, reaching its peak at 72 h. In summary, the results demonstrated that high glucose concentrations alone may be sufficient to alter the in vitro mediators’ production of RAW 264.7 cells.

Antimicrobial and immunomodulatory activity of host defense peptides, clavanins and LL-37, in vitro: An endodontic perspective

&NA; Endodontic treatment is mainly based on root canal disinfection and its failure may be motivated by microbial resistance. Endodontic therapy can be benefitted by host defense peptides (HDPs), which are multifunctional molecules that act against persistent infection and inflammation. This study aimed to evaluate the antimicrobial, cytotoxic and immunomodulatory activity of several HDPs, namely clavanin A, clavanin A modified (MO) and LL‐37, compared to intracanal medication Ca(OH)2. HDPs and Ca(OH)2 were evaluated by: (1) antimicrobial assays against Candida albicans and Enterococcus faecalis, (2) cytotoxicity assays and (3) cytokine tumor necrosis factor (TNF)‐&agr;, monocyte chemoattractant protein (MCP)‐1, interleukin (IL)‐1&agr;, IL‐6, IL‐10 and IL‐12 and nitric oxide (NO) production by RAW 264.7 cells incubated with or without heat‐killed (HK) C. albicans or E. faecalis combined or not with interferon‐&ggr;. The minimum inhibitory concentration (MIC) was established only for E. faecalis (LL‐37, 57 &mgr;M). Considering cytotoxicity, clavanin MO was able to reduce cell viability in many groups and demonstrated lowest LC50. The Ca(OH)2 up‐regulated the production of MCP‐1, TNF‐&agr;, IL‐12 and IL‐6 and down‐regulated IL‐1&agr;, IL‐10 and NO. Clavanins up‐regulated the TNF‐&agr; and NO and down‐regulated IL‐10 production. LL‐37 demonstrated up‐regulation of IL‐6 and TNF‐&agr; production and down‐regulation in IL‐10 and NO production. In conclusion, LL‐37 demonstrated better antibacterial potential. In addition, Ca(OH)2 demonstrated a proinflammatory response, while the HDPs modulated the inflammatory response from non‐interference with the active cytokines in the osteoclastogenesis process, probably promoting the health of periradicular tissues. HighlightsLL‐37 demonstrates better antimicrobial activity compared to Ca(OH)2 and clavanins against E. faecalis.Clavanin MO can be considered potentially cytotoxic.Ca(OH)2 can be considered a pro‐inflammatory agent due to up‐stimulation of MCP‐1, TNF‐&agr;, IL‐12 and IL‐6 production.

An Immunomodulatory Peptide Confers Protection in an Experimental Candidemia Murine Model

ABSTRACT Fungal Candida species are commensals present in the mammalian skin and mucous membranes. Candida spp. are capable of breaching the epithelial barrier of immunocompromised patients with neutrophil and cell-mediated immune dysfunctions and can also disseminate to multiple organs through the bloodstream. Here we examined the action of innate defense regulator 1018 (IDR-1018), a 12-amino-acid-residue peptide derived from bovine bactenecin (Bac2A): IDR-1018 showed weak antifungal and antibiofilm activity against a Candida albicans laboratory strain (ATCC 10231) and a clinical isolate (CI) (MICs of 32 and 64 μg · ml−1, respectively), while 8-fold lower concentrations led to dissolution of the fungal cells from preformed biofilms. IDR-1018 at 128 μg · ml−1 was not hemolytic when tested against murine red blood cells and also has not shown a cytotoxic effect on murine monocyte RAW 264.7 and primary murine macrophage cells at the tested concentrations. IDR-1018 modulated the cytokine profile during challenge of murine bone marrow-derived macrophages with heat-killed C. albicans (HKCA) antigens by increasing monocyte chemoattractant protein 1 (MCP-1) and interleukin-10 (IL-10) levels, while suppressing tumor necrosis factor alpha (TNF-α), IL-1β, IL-6, and IL-12 levels. Mice treated with IDR-1018 at 10 mg · kg−1 of body weight had an increased survival rate in the candidemia model compared with phosphate-buffered saline (PBS)-treated mice, together with a diminished kidney fungal burden. Thus, IDR-1018 was able to protect against murine experimental candidemia and has the potential as an adjunctive therapy.