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Introduction: The combination of patterns is a frequent and challenging situation in the daily laboratory routine of autoantibodies testing using HEp-2 cells indirect immunofluorescence assay (HEp-2-IFA). Recently, the Brazilian Consensus on Autoantibodies (BCA) named these combinations as complex patterns (CPs) and organized them into 3 subtypes: multiple, mixed, and composite. This study aimed to describe the most frequent combinations of HEp-2-IIF patterns according to this new nomenclature., Methods: Routine HEp-2-IFA results reported in January and June 2017 were reviewed using the new BCA classification. Visual pattern recognition was performed by experts on HEp-2-IFA readings, using the International Consensus on Antinuclear Antibodies (ANA) Patterns (ICAP) and BCA recommendations., Results: 54,990 serum samples from different patients were tested for ANA-HEp-2, and 11,478 (20.9%) were positive at a titer ≥ 1/80. Among these positive samples, 1,111 (9.7%) displayed CPs, divided into 95 different combinations. A higher proportion of CPs was observed in the pediatric age group. Multiple, mixed, and composite patterns were present in 85.3, 5.4, and 9.5% of the samples, respectively. In the multiple/mixed pattern group (n=1,005), double, triple, and quadruple combinations (ICAP/BCA codes) were observed in 97.7%, 2.2%, and 0.1%, respectively. The double nuclear pattern was the most prevalent combination observed (67.6%). The most common CPs registered were AC-4 (nuclear fine speckled) + AC-6,7 (nuclear discrete dots) (n=264); AC-2 (nuclear dense fine speckled) + AC-6,7 (n=201); AC-4+AC-8,9,10 (nucleolar) (n=129); and AC-3 (centromere)+AC-4 (n=124). All of these combinations were in the multiple subgroup., Conclusion: Almost 10% of positive results in the HEp-2 procedure displayed CPs. Among the 3 subtypes of CPs proposed, the multiple pattern was the most prevalent, especially in the pediatric population. The AC-4, AC-2, and AC-6,7 were the most prevalent single patterns observed in the combinations described in this study. There was a significant association between age and the prevalence of most combined patterns. The AC-4+AC-6,7 combination was the most prevalent complex pattern detected regardless of the age group. The AC-2+AC-6,7 was more prevalent in younger individuals. The concepts involved in the CPs definition should add value to the reading and interpretation of the HEp-2-IIF assay., Competing Interests: Authors WS, AC, DF and NG were employed by company Sabin Medicina and Health. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Santos, Cantuária, Félix, Guimarães and de Melo.)

The consumption of high-protein diets (HPD) is associated with resistance training (RT) due to effects on metabolism. However, little is known about these effects on cardiac tissue. This study aimed to investigate effects of HPD and RT on cardiac biomarkers. 18 rats were divided into normo-protein (NPD), and HPD groups: NPD-Control, NPD-RT, HPD-Control, and HPD-RT. Interleukin-6 (IL-6), tumour necrosis factor alpha (TNF-a), nitric oxide (NO), activity of metalloproteinase-2 (MMP-2), and vascular factor (VEGF) were analysed. RT was effective in regulating body weight, increasing strength, and reducing food consumption ( p  < .05). HPD induces higher levels of interleukin 6 ( p  = .0169), and lowers NO ( p  < .0001). When associated with RT, the HPD decreases levels of tumour necrosis factor alpha, while enhances NO, and MMP activity ( p  < .05). The association of RT with HDP decreases inflammatory parameters and indicates an enhancement in the molecular parameters of cardiac tissue.

Background: The prevalence of anti-cell autoantibodies detected by indirect immunofluorescence assay on HEp-2 cells (HEp-2-IIFA) increases with age and is higher in female sex. The number of medical specialties that use HEp-2-IIFA in the investigation of autoimmune diseases has increased lately. This study aimed to determine the prevalence and patterns of autoantibodies on HEp-2-IIFA according to demographics variables and referring medical specialties., Methods: A retrospective analysis of the HEp-2-IIFA carried out between January and June of 2017 was performed. The International Consensus on Antinuclear Antibodies Patterns (ICAP) and the Brazilian Consensus on Autoantibodies were used for patterns definition on visual reading of the slides. Anti-cell (AC) codes from ICAP and Brazilian AC codes (BAC) were used for patterns classification., Results: From 54,990 samples referred for HEp-2-IIF testing, 20.9% were positive at titer ≥ 1/80. HEp-2-IIFA positivity in females and males was 24% and 12%, respectively (p < 0.0001). The proportion of positive results in the 4 age groups analyzed: 0-19, 20-39, 40-59, and ≥ 60 years was 23.3, 20.2, 20.1, and 22.8%, respectively (p < 0.0001). Considering all positive sera (n = 11,478), AC-4 nuclear fine speckled (37.7%), AC-2 nuclear dense fine speckled (21.3%), BAC-3 nuclear quasi-homogeneous (10%) and mixed/composite patterns (8.8%) were the most prevalent patterns. The specialties that most requested HEp-2-IIFA were general practitioner (20.1%), dermatology (15%), gynecology (9.9%), rheumatology (8.5%), and cardiology (5.8%). HEp-2-IIFA positivity was higher in patients referred by rheumatologists (35.7% vs. 19.6%) (p < 0.0001). Moderate (46.4%) and high (10.8%) titers were more observed in patients referred by rheumatologists (p < 0.0001). We observed a high proportion of mixed and cytoplasmic patterns in samples referred by oncologists and a high proportion of BAC-3 (nuclear quasi-homogeneous) pattern in samples referred by pneumologists., Conclusions: One-fifth of the patients studied were HEp-2-IIFA-positive. The age groups with more positive results were 0-19 and ≥ 60 years. AC-4, AC-2, BAC-3 and mixed/composite patterns were the most frequent patterns observed. Rheumatologists requested only 8.5% of HEp-2-IIFA. Positive results and moderate to high titers of autoantibodies were more frequent in patients referred by rheumatologists., (© 2022. The Author(s).)

Aim: To evaluate the antimicrobial and immunomodulatory activity of double antibiotic paste (DAP) in an in vitro infection model., Methodology: The minimum inhibitory and bactericidal concentrations (MIC and MBC) and the antibiofilm activities (TTC assay) of DAP and its components (ciprofloxacin and metronidazole) were evaluated against Staphylococcus aureus and Enterococcus faecalis compared with triple antibiotic paste (TAP). The cellular viability of RAW 264.7 macrophages (24 and 72 h) and L929 fibroblasts (48 and 72 h) was evaluated by MTT. Furthermore, the production of TNF-α, IL-12, IL-6, IL-1α, IL-10 and NO (on RAW 264.7), besides IL-6, TGF-β and NO (on L929), stimulated with DAP in baseline and associated with heat-killed microbial-antigen conditions was measured by ELISA and Griess reaction. Data were analysed using the one-way ANOVA test with Bonferroni's corrections., Results: The MBC of pharmacopoeia DAP was similar to TAP for E. faecalis (0.25 μg.  mL -1 ) and lower for S. aureus (DAP 1 μg. mL -1 and TAP 2 μg. mL -1 ; p < .001). Ciprofloxacin was the most effective antibiofilm drug from the pastes (35% of reduction for E. faecalis and S. aureus; p < .0001), and both pastes had a similar antibiofilm eradication against both biofilm species (29% and 35% for S. aureus and 76% and 85% for E. faecalis; p < .0001). DAP was cytotoxic against the tested cells. DAP significantly upregulated IL-1α (p < .001), IL-6 (p < .0001), TNF-α (p < .01) and IL-12 (p < .05; in the absence of antigens) and significantly reduced IL-6 (p < .0001; in the presence of HK-S. aureus) and IL-10 (p < .05; in the presence of both antigens) on macrophages. Furthermore, DAP upregulated IL-6 (p < .001) and NO (p < .05; in the absence of antigens), IL-6 (p < .001; in the presence of HK-S. aureus) and reduced NO (p < .001; in the presence of HK-S. aureus)., Conclusions: Double antibiotic paste and TAP had similar antimicrobial activity against S. aureus and E. faecalis. DAP upregulated pro-inflammatory cytokines mainly in the absence of antigens and had pro- and anti-inflammatory activity in RAW 264.7 macrophages and L929 fibroblasts in the presence of antigens involved in pulp infections., (© 2021 International Endodontic Journal. Published by John Wiley & Sons Ltd.)

High-protein diets (HPDs) are widely accepted as a way to stimulate muscle protein synthesis when combined with resistance training (RT). However, the effects of HPDs on adipose tissue plasticity and local inflammation are yet to be determined. This study investigated the impact of HPDs on glucose control, adipocyte size, and epididymal adipose inflammatory biomarkers in resistance-trained rats. Eighteen Wistar rats were randomly assigned to four groups: normal-protein (NPD; 17% protein total dietary intake) and HPD (26.1% protein) without RT and NPD and HPD with RT. Trained groups received RT for 12 weeks with weights secured to their tails. Glucose and insulin tolerance tests, adipocyte size, and an array of cytokines were determined. While HPD without RT induced glucose intolerance, enlarged adipocytes, and increased TNF-α, MCP-1, and IL1-β levels in epididymal adipose tissue ( p < 0.05), RT diminished these deleterious effects, with the HPD + RT group displaying improved blood glucose control without inflammatory cytokine increases in epididymal adipose tissue ( p < 0.05). Furthermore, RT increased glutathione expression independent of diet ( p < 0.05). RT may offer protection against adipocyte hypertrophy, pro-inflammatory states, and glucose intolerance during HPDs. The results highlight the potential protective effects of RT to mitigate the maladaptive effects of HPDs.

Objectives: In order to evaluate host defense peptides (HDPs) HHC-10 and synoeca-MP activity in in vitro osteoclastogenesis process and in vivo induced apical periodontitis, testing the effect of molecules in the inflammatory response and in apical periodontitis size/volume after root canal treatment., Materials and Methods: In vitro osteoclastogenesis was assessed on bone marrow cell cultures extracted from mice, while in vivo endodontic treatment involved rats treated with Ca(OH) 2 or HDPs. In vitro osteoclasts were subjected to TRAP staining, and in vivo samples were evaluated by radiographic and tomographic exams, as well as histologic analysis., Results: None of the substances downregulated the in vitro osteoclastogenesis. Nevertheless, all treatments affected the average of apical periodontitis size in rats, although only teeth treated with HDPs demonstrated lower levels of the inflammatory process. These results demonstrated the in vivo potential of HDPs. Radiographic analysis suggested that HHC-10 and synoeca-MP-treated animals presented a similar lesion size than Ca(OH) 2 -treated animals after 7-day of endodontic treatment. However, tomography analysis demonstrated smaller lesion volume in synoeca-MP-treated animals than HHC-10 and Ca(OH) 2 -treated animals, after 7 days., Conclusions: These molecules demonstrated an auxiliary effect in endodontic treatment that might be related to its immunomodulatory ability, broad-spectrum antimicrobial activity, and possible induction of tissue repair at low concentrations. These results can encourage further investigations on the specific mechanisms of action in animal models to clarify the commercial applicability of these biomolecules for endodontic treatment., Clinical Significance: HDPs have the potential to be adjuvant substances in endodontic therapy due to its potential to reduce inflammation in apical periodontitis.

Regenerative therapies such as dental pulpal revascularization appear as an option for traumatized immature permanent teeth. However, the triple antibiotic paste - TAP (metronidazole, minocycline, and ciprofloxacin), used for these therapies, can generate cytotoxicity and dentin discoloration. In contrast, host defense peptides (HDPs) are promising antimicrobial and immunomodulatory biomolecules for dentistry. This study aimed to evaluate in vitro the antimicrobial activity (against Staphylococcus aureus and Enterococcus faecalis) and the immunomodulatory potential (by the evaluation of IL-1α, IL-6, IL-12, IL-10, TNF-α and NO, in RAW 264.7 macrophages and IL-6, TGF-β and NO, in L929 fibroblast) of synthetic peptides (DJK-6, IDR-1018, and IDR-1002), compared to TAP in an in vitro infection model containing heat-killed antigens from E. faecalis and S. aureus. Furthermore, the synergistic potential of ciprofloxacin and IDR-1002 was evaluated by checkerboard. Ciprofloxacin was the best antimicrobial of TAP, besides acting in synergism with IDR-1002. TAP was pro-inflammatory (p < 0.05), while the association of ciprofloxacin and IDR-1002 presented an anti-inflammatory profile mainly in the presence of both heat-killed antigens (p < 0.05). Based on these results, ciprofloxacin associated with IDR-1002 may demonstrate an efficient antimicrobial and immunomodulatory action in this in vitro model. Further in vivo studies may determine the real potential of this combination., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬‬This study aims to evaluate the in vitro antimicrobial and immunomodulatory activities and cytotoxicity of chlorhexidine (CHX) and synoeca-MP peptide alone or in combination against Pseudomonas aeruginosa. The antimicrobial property was evaluated by the determination of minimal inhibitory concentration, minimum bactericidal concentration, and planktonic bacteria and biofilm inhibition. Immunomodulatory activity was determined by enzyme-linked immunosorbent assay and nitric oxide production by the Griess reaction method. According to the results, synoeca-MP combined with CHX demonstrated antimicrobial effectiveness compared with its isolated use, in addition to immunomodulatory activity (upregulating MPC-1 and tumor necrosis factor-α and downregulating nitric oxide and interleukin-10). In this context, it is expected that the substances, together, could be capable of controlling bacterial infection and dissemination, besides potentiating macrophages' immune response against the studied microorganism. Moreover, reducing the CHX concentration by the addition of synoeca-MP peptide may, in a beneficial way, minimize the undesirable effects of both, CHX and synoeca-MP in a clinical setting., (© 2019 Wiley Periodicals, Inc.)

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)-α production. Groups stimulated with lipopolysaccharides (LPS) and LPS and recombinant interferon (rIFN)-γ down-regulated interleukin (IL)-1α, 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., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

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)-α, monocyte chemoattractant protein (MCP)-1, interleukin (IL)-1α, 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-γ. The minimum inhibitory concentration (MIC) was established only for E. faecalis (LL-37, 57μM). Considering cytotoxicity, clavanin MO was able to reduce cell viability in many groups and demonstrated lowest LC 50 . The Ca(OH) 2 up-regulated the production of MCP-1, TNF-α, IL-12 and IL-6 and down-regulated IL-1α, IL-10 and NO. Clavanins up-regulated the TNF-α and NO and down-regulated IL-10 production. LL-37 demonstrated up-regulation of IL-6 and TNF-α 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., (Copyright © 2017 Elsevier Inc. All rights reserved.)

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., (Copyright © 2017 American Society for Microbiology.)

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., (Copyright © 2015 Elsevier B.V. All rights reserved.)