FOXO1 regulates wound-healing responses in human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Forkhead box-O 1 (FOXO1) is a transcription factor actively involved in oral wound healing at the epithelial barrier. However, less is known regarding the role of FOXO1 during the tissue repair response in the connective tissue compartment. This study explored the involvement of FOXO1 in the modulation of fibroblast activity related to wound healing. METHODS: Primary cultures of human gingival fibroblasts were obtained from four healthy young donors. Myofibroblastic differentiation, collagen gel contraction, cell migration, cell spreading, and integrin activation were evaluated in the presence or absence of a FOXO1 inhibitor (AS1842856). Variations in mRNA and proteins of interest were evaluated through qRT-PCR and western blot, respectively. Distribution of actin, α-smooth muscle actin, and ß1 integrin was evaluated using immunofluorescence. FOXO1 and TGF-ß1 expression in gingival wound healing was assessed by immunohistochemistry in gingival wounds performed in C57BL/6 mice. Images were analyzed using ImageJ/Fiji. ANOVA or Kruskal-Wallis test followed by Tukey's or Dunn's post-hoc test was performed. All data are expressed as mean ± SD. p < .05 was considered statistically significant. RESULTS: FOXO1 inhibition caused a decrease in the expression of the myofibroblastic marker α-SMA along with a reduction in fibronectin, type I collagen, TGF-ß1, and ß1 integrin mRNA level. The FOXO1 inhibitor also caused decreases in cell migration, cell spreading, collagen gel contraction, and ß1 integrin activation. FOXO1 and TGF-ß1 were prominently expressed in gingival wounds in fibroblastic cells located at the wound bed. CONCLUSION: The present study indicates that FOXO1 plays an important role in the modulation of several wound-healing functions in gingival fibroblast. Moreover, our findings reveal an important regulatory role for FOXO1 on the differentiation of gingival myofibroblasts, the regulation of cell migration, and collagen contraction, all these functions being critical during tissue repair and fibrosis.

C1q and HBHA-specific IL-13 levels as surrogate plasma biomarkers for monitoring tuberculosis treatment efficacy: a cross-sectional cohort study in Paraguay.

Introduction: New diagnostic tools are needed to rapidly assess the efficacy of pulmonary tuberculosis (PTB) treatment. The aim of this study was to evaluate several immune biomarkers in an observational and cross-sectional cohort study conducted in Paraguay. Methods: Thirty-two patients with clinically and microbiologically confirmed PTB were evaluated before starting treatment (T0), after 2 months of treatment (T1) and at the end of treatment (T2). At each timepoint plasma levels of IFN-y, 17 pro- and anti-inflammatory cytokines/chemokines and complement factors C1q, C3 and C4 were assessed in unstimulated and Mtb-specific stimulated whole blood samples using QuantiFERON-TB gold plus and recombinant Mycobacterium smegmatis heparin binding hemagglutinin (rmsHBHA) as stimulation antigen. Complete blood counts and liver enzyme assays were also evaluated and correlated with biomarker levels in plasma. Results: In unstimulated plasma, C1q (P<0.001), C4 (P<0.001), hemoglobin (P<0.001), lymphocyte proportion (P<0.001) and absolute white blood cell count (P=0.01) were significantly higher in PTB patients at baseline than in cured patients. C1q and C4 levels were found to be related to Mycobacterium tuberculosis load in sputum. Finally, a combinatorial analysis identified a plasma host signature comprising the detection of C1q and IL-13 levels in response to rmsHBHA as a tool differentiating PTB patients from cured TB profiles, with an AUC of 0.92 (sensitivity 94% and specificity 79%). Conclusion: This observational study provides new insights on host immune responses throughout anti-TB treatment and emphasizes the role of host C1q and HBHA-specific IL-13 response as surrogate plasma biomarkers for monitoring TB treatment efficacy.

The Experience of Testing for Coronavirus Disease (COVID-19) at a Single Diagnostic Center in Paraguay before the Introduction of Vaccination.

Soon after the declaration of the COVID-19 pandemic, the Institute for Health Sciences Research (IICS) of the National University of Asunción, Paraguay became a testing laboratory (COVID-Lab) for SARS-CoV-2. The COVID-Lab testing performance was assessed from 1 April 2020 to 12 May 2021. The effect of the pandemic on the IICS and how the COVID-Lab contributed to the academic and research activities of the institute were also assessed. IICS researchers and staff adjusted their work schedules to support the COVID-Lab. Of the 13,082 nasopharyngeal/oropharyngeal swabs processed, 2704 (20.7%) tested positive for SARS-CoV-2 by RT-PCR. Of the individuals testing positive, 55.4% were female and 48.3% were aged 21-40 years. Challenges faced by the COVID-Lab were unstable reagent access and insufficient staff; shifting obligations regarding research, academic instruction, and grantsmanship; and the continuous demands from the public for information on COVID-19. The IICS provided essential testing and reported on the progress of the pandemic. IICS researchers gained better laboratory equipment and expertise in molecular SARS-CoV-2 testing but struggled to manage their conflicting educational and additional research obligations during the pandemic, which affected their productivity. Therefore, policies protecting the time and resources of the faculty and staff engaged in pandemic-related work or research are necessary components of healthcare emergency preparedness.

Protective Effect of Tilia americana var. mexicana Against Kainic Acid-induced Damage in Brain, Liver, and Kidney: Behavioral and Biochemical Changes.

Tiliaamericana var. mexicana (Tilia) possesses anticonvulsant, antioxidant, neuroprotective, and hepatoprotective activities. The spectrum of anticonvulsant activity in status epilepticus models has not been sufficiently explored. We evaluated the effects of ethyl acetate (EAc), and methanol (ME) extracts on kainic acid (KA)-induced seizures by measuring rats'behavior (severity and latency) and lipoperoxidation in different brain areas (cerebellum, brain hemispheres, cortex, and medulla), kidneys, and liver. Male Wistar rats were administered KA (10 mg/kg, i.p.) after three days of pretreatment with Tilia extract (100 mg/kg). The EAc and ME Tilia extracts significantly decreased the severity of phase 1 and phase 2 seizures, respectively. The ME Tilia extract increased the latency to seizure (27 ± 2 min) compared to the control (13 ± 2 min). The ME and EAc Tilia extracts significantly prevented the increased lipid peroxidation caused by KA-induced seizures in the cerebellum, brain hemispheres, cortex, medulla, liver, and kidneys. The vehicle olive oil (OO) also showed anticonvulsant effects, decreasing the severity of seizures to phase 3 and lipoperoxidation levels in the cerebellum, brain hemispheres, cortex, medulla, liver, and kidneys. The anticonvulsant activity of Tilia is mediated by antioxidant effects in central and systemic areas that involve synergistic interactions among the chemical constituents of these extracts (glucosides of quercetin and kaempferol), while vehicle OO showed the same effects, probably due to its constituent oleuropein.

Mechanical Properties and Antibacterial Effect on Mono-Strain of Streptococcus mutans of Orthodontic Cements Reinforced with Chlorhexidine-Modified Nanotubes.

Recently, several studies have introduced nanotechnology into the area of dental materials with the aim of improving their properties. The objective of this study is to determine the antibacterial and mechanical properties of type I glass ionomers reinforced with halloysite nanotubes modified with 2% chlorhexidine at concentrations of 5% and 10% relative to the total weight of the powder used to construct each sample. Regarding antibacterial effect, 200 samples were established and distributed into four experimental groups and six control groups (4 +ve and 2 -ve), with 20 samples each. The mechanical properties were evaluated in 270 samples, assessing microhardness (30 samples), compressive strength (120 samples), and setting time (120 samples). The groups were characterized by scanning electron microscopy and Fourier transform infrared spectroscopy, and the antibacterial activity of the ionomers was evaluated on Streptococcus mutans for 24 h. The control and positive control groups showed no antibacterial effect, while the experimental group with 5% concentration showed a zone of growth inhibition between 11.35 mm and 11.45 mm, and the group with 10% concentration showed a zone of growth inhibition between 12.50 mm and 13.20 mm. Statistical differences were observed between the experimental groups with 5% and 10% nanotubes. Regarding the mechanical properties, microhardness, and setting time, no statistical difference was found when compared with control groups, while compressive strength showed higher significant values, with ionomers modified with 10% concentration of nanotubes resulting in better compressive strength values. The incorporation of nanotubes at concentrations of 5% and 10% effectively inhibited the presence of S. mutans, particularly when the dose-response relationship was taken into account, with the advantage of maintaining and improving their mechanical properties.

SARS-CoV-2 Variants in Paraguay: Detection and Surveillance with an Economical and Scalable Molecular Protocol.

SARS-CoV-2 variant detection relies on resource-intensive whole-genome sequencing methods. We sought to develop a scalable protocol for variant detection and surveillance in Paraguay, pairing rRT-PCR for spike mutations with Nanopore sequencing. A total of 201 acute-phase nasopharyngeal samples were included. Samples were positive for the SARS-CoV-2 N2 target and tested with the Spike SNP assay to detect mutations associated with the following variants: alpha (501Y), beta/gamma (417variant/484K/501Y), delta (452R/478K), and lambda (452Q/490S). Spike SNP calls were confirmed using amplicon (Sanger) sequencing and whole-genome (Nanopore) sequencing on a subset of samples with confirmed variant lineages. Samples had a mean N2 Ct of 20.8 (SD 5.6); 198/201 samples (98.5%) tested positive in the Spike SNP assay. The most common genotype was 417variant/484K/501Y, detected in 102/198 samples (51.5%), which was consistent with the P.1 lineage (gamma variant) in Paraguay. No mutations (K417 only) were found in 64/198 (32.3%), and K417/484K was identified in 22/198 (11.1%), consistent with P.2 (zeta). Seven samples (3.5%) tested positive for 452R without 478K, and one sample with genotype K417/501Y was confirmed as B.1.1.7 (alpha). The results were confirmed using Sanger sequencing in 181/181 samples, and variant calls were consistent with Nanopore sequencing in 29/29 samples. The Spike SNP assay could improve population-level surveillance for mutations associated with SARS-CoV-2 variants and inform the judicious use of sequencing resources.

Exposure to Sub-Lethal Doses of Permethrin Is Associated with Neurotoxicity: Changes in Bioenergetics, Redox Markers, Neuroinflammation and Morphology.

Permethrin (PERM) is a member of the class I family of synthetic pyrethroids. Human use has shown that it affects different systems, with wide health dysfunctions. Our aim was to determine bioenergetics, neuroinflammation and morphology changes, as redox markers after subacute exposure to PERM in rats. We used MDA determination, protein carbonyl assay, mitochondrial O2 consumption, expression of pro-inflammatory cytokines and a deep histopathological analysis of the hippocampus. PERM (150 mg/kg and 300 mg/kg body weight/day, o.v.) increased lipoperoxidation and carbonylated proteins in a dose-dependent manner in the brain regions. The activities of antioxidant enzymes glutathione peroxidase, reductase, S-transferase, catalase, and superoxide dismutase showed an increase in all the different brain areas, with dose-dependent effects in the cerebellum. Cytokine profiles (IL-1ß, IL-6 and TNF-α) increased in a dose-dependent manner in different brain tissues. Exposure to 150 mg/kg of permethrin induced degenerated and/or dead neurons in the rat hippocampus and induced mitochondrial uncoupling and reduction of oxidative phosphorylation and significantly decreased the respiratory parameters state 3-associated respiration in complex I and II. PERM exposure at low doses induces reactive oxygen species production and imbalance in the enzymatic antioxidant system, increases gene expression of pro-inflammatory interleukins, and could lead to cell damage mediated by mitochondrial functional impairment.

Ketogenic Diet Provided During Three Months Increases KCC2 Expression but Not NKCC1 in the Rat Dentate Gyrus.

Ketogenic diet, a high fat and low carbohydrate diet, has been used as a non-pharmacological treatment in refractory epilepsy since 1920. In recent years, it has demonstrated to be effective in the treatment of numerous neurological and non-neurological diseases. Some neurological and neuropsychiatric disorders are known to be caused by gamma-aminobutyric acid (GABA)-mediated neurotransmission dysfunction. The strength and polarity of GABA-mediated neurotransmission are determined by the intracellular chloride concentration, which in turn is regulated by cation-chloride cotransporters NKCC1 and KCC2. Currently, it is unknown if the effect of ketogenic diet is due to the modulation of these cotransporters. Thus, we analyzed the effect of a ketogenic diet on the cation-chloride cotransporters expression in the dentate gyrus. We estimated the total number of NKCC1 immunoreactive (NKCC1-IR) neuronal and glial cells by stereology and determined KCC2 labeling intensity by densitometry in the molecular and granule layers as well as in the hilus of dentate gyrus of rats fed with normal or ketogenic diet for 3 months. The results indicated that ketogenic diet provided during 3 months increased KCC2 expression, but not NKCC1 in the dentate gyrus of the rat. The significant increase of KCC2 expression could explain, at least in part, the beneficial effect of ketogenic diet in the diseases where the GABAergic system is altered by increasing its inhibitory efficiency.

Use of dermal matrices to change gingival phenotypes

ABSTRACT: A variety of periodontal plastic surgery techniques have been proposed to correct aesthetic and functional problems of periodontal tissues. The most common procedure uses the connective tissue graft from the palate. However, patient discomfort and the limited quantity of palatal tissue results in many cases where the surgeon uses connective tissue substitutes. In this case report, we describe the use of a human dermal matrix for gingival volume augmentation in the lower incisors with a novel modified VISTA technique stabilizing the flap with sutures and cyanoacrylates, avoiding comorbidity of a second surgical site. The follow up at 6 months showed an increase in gingival thickness and a reduction in recessions length.

Uncoupled inflammatory, proliferative, and cytoskeletal responses in senescent human gingival fibroblasts.

BACKGROUND AND OBJECTIVE: Aging is characterized by a decline in tissue structure and function that may be explained by the development of cellular senescence. However, the acquisition of specific phenotypic responses in senescent gingival fibroblasts is still poorly understood. Here, we have analyzed whether proliferation of primary cultures of human gingival fibroblasts may affect different cell functions relevant to cellular senescence and tissue deterioration. METHODS: Human gingival fibroblasts from five young donors were expanded until cellular senescence was achieved. Cellular senescence was evaluated by determining modifications in cell size, cell proliferation, p16 and p21 mRNA levels, H2Ax phosphorylation, cell viability, and senescence-associated beta-galactosidase staining. Inflammation was evaluated by analyzing the secretion of cytokines and nuclear translocation of NF-κB. Collagen remodeling was evaluated using a collagen gel contraction assay. Immunofluorescence and confocal microscopy were used to determine changes in the localization of the cytoskeletal proteins. Data analysis was performed to identify changes between cultures of the same donor at early and late passages using the paired sample t test or the Wilcoxon matched-pairs signed-rank test. RESULTS: Late passage cells showed changes compatible with cellular senescence that included increased cell size, reduced cell proliferation, staining for SA-beta gal, phosphorylated H2Ax, and increased p16 and p21 mRNA levels. Late passage cells showed a decrease in collagen contraction and reduced co-localization between the cytoskeletal proteins actin and vinculin. Importantly, late passage cells neither demonstrated changes in the secretion of inflammatory cytokines nor NF-κB activation. CONCLUSION: Our results imply that cytoskeletal changes and inhibition of cell proliferation represent early modifications in the structure and function of senescent gingival fibroblasts that are not coupled with the acquisition of an inflammatory phenotype. Further studies are needed to clarify the impact of different senescence stages during aging of the periodontium.

Inhibitory effect of serotype a of Aggregatibacter actinomycetemcomitans on the increased destructive potential of serotype b.

OBJECTIVE: The serotype b of Aggregatibacter actinomycetemcomitans (A. actinomycetemcomitans) induces higher cytokine production in dendritic cells (DCs) compared with the other serotypes. However, this increased immunostimulatory potential was modified when DCs were co-infected with the other A. actinomycetemcomitans serotypes. This study aimed to analyze whether the production of interferon gamma (IFN-γ), C-reactive protein (CRP), matrix metalloproteinase (MMP)-2, and MMP-9, as well as the activity of osteoclasts, also varies when DCs are co-infected with the A. actinomycetemcomitans serotypes. MATERIALS AND METHODS: Human DCs were stimulated with the A. actinomycetemcomitans serotypes using the following stimulatory conditions: serotype a/b/c/a+b/a+c/b+c/a+b+c. The IFN-γ, CRP, and MMP-2 levels were quantified by ELISA. The active form of MMP-9 was quantified using fluorescent functional assays. The MMP-2 gelatinolytic activity was identified by zymogram. The osteoclast activity was determined by quantifying the TRAP expression and resorption-pit formation using cytochemistry and osteoassays. RESULTS: Higher levels of IFN-γ, CRP, MMP-2, MMP-9, and osteoclast activity were detected when DCs were stimulated with the serotype b of A. actinomycetemcomitans compared with the others. This increased immunostimulatory potential attributed to serotype b diminished when DCs were co-infected with the serotype a. CONCLUSIONS: This study provides new insights into the virulence of A. actinomycetemcomitans and reveals important differences in the immunostimulatory and pro-destructive potential among its serotypes.

Capsular-defective Porphyromonas gingivalis mutant strains induce less alveolar bone resorption than W50 wild-type strain due to a decreased Th1/Th17 immune response and less osteoclast activity.

BACKGROUND: Encapsulation of Porphyromonas gingivalis has been demonstrated as responsible of several host immunological changes, which have been associated with the pathogenesis of periodontitis. Using a murine model of periodontitis and two isogenic non-capsulated mutants of P. gingivalis, this study aimed to analyze whether P. gingivalis encapsulation induces more severe alveolar bone resorption, and whether this bone loss is associated with a T-helper (Th)1 and Th17-pattern of immune response. METHODS: Experimental periodontal infections were generated by oral inoculation with the encapsulated W50 wild-type strain or isogenic non-encapsulated ΔPG0116-PG0120 (GPA) and ΔPG0109-PG0118 (GPC) mutants of P. gingivalis. Periodontal infections induced with the encapsulated HG184 or non-encapsulated ATCC 33277 strains of P. gingivalis were used as controls. Alveolar bone resorption was analyzed using microcomputed tomography and scanning electron microscopy. The expression levels of Th1, Th2, Th17, or T regulatory-associated cytokines and RANKL, as well as the periodontal bacterial load, were quantified by quantitative polymerase chain reaction. The detection of Th1 and Th17 lymphocytes was analyzed by flow cytometry. RESULTS: In the periodontal lesions, both capsular-defective knockout mutant strains of P. gingivalis induced less alveolar bone resorption than the encapsulated W50 wild-type strain. This decreased bone loss was associated with a dismissed RANKL expression, decreased Th1- and Th17-type of cytokine expression, reduced Th1 and Th17 lymphocyte detection, and low osteoclast finding. CONCLUSION: These data demonstrate that encapsulation of P. gingivalis plays a key role in the alveolar bone resorption induced during periodontitis, and this bone loss is associated with a Th1- and Th17-pattern of immune response triggered in the periodontal lesions.

Dose-Dependent Behavioral and Antioxidant Effects of Quercetin and Methanolic and Acetonic Extracts from Heterotheca inuloides on Several Rat Tissues following Kainic Acid-Induced Status Epilepticus.

Kainic acid (KA) has been used to study the neurotoxicity induced after status epilepticus (SE) due to activation of excitatory amino acids with neuronal damage. Medicinal plants can protect against damage caused by KA-induced SE; in particular, organic extracts of Heterotheca inuloides and its metabolite quercetin display antioxidant activity and act as hepatoprotective agents. However, it is unknown whether these properties can protect against the hyperexcitability underlying the damage caused by KA-induced SE. Our aim was to study the protective effects (with regard to behavior and antioxidant activity) of administration of natural products methanolic (ME) and acetonic (AE) extracts and quercetin (Q) from H. inuloides at doses of 30 mg/kg (ME30, AE30, and Q30 groups), 100 mg/kg (ME100, AE100, and Q100 groups), and 300 mg/kg (ME300, AE300, and Q300 groups) against damage in brain regions of male Wistar rats treated with KA. We found dose-dependent effects on behavioral and biochemical studies in the all-natural product groups vs. the control group, with decreases in seizure severity (Racine's scale) and increases in seizure latency (p < 0.05 in the ME100, AE100, Q100, and Q300 groups and p < 0.01 in the AE300 and ME300 groups); on lipid peroxidation and carbonylated proteins in all brain tissues (p < 0.0001); and on GPx, GR, CAT, and SOD activities with all the treatments vs. KA (p ≤ 0.001). In addition, there were strong negative correlations between carbonyl levels and latency in the group treated with KA and in the group treated with methanolic extract in the presence of KA (r = -0.9919, p = 0.0084). This evidence suggests that organic extracts and quercetin from H. inuloides exert anticonvulsant effects via direct scavenging of reactive oxygen species (ROS) and modulation of antioxidant enzyme activity.

Congenital Zika Syndrome in Puerto Rico, Beyond Microcephaly, A Multiorgan Approach.

OBJECTIVE: Zika virus (ZIKV) infection was identified in Puerto Rico on December 2015, and the outbreak encouraged us to characterize clinical manifestations and laboratory findings of intrauterine exposed infants. METHODS: Retrospective medical record review of infants born to mothers with confirmed ZIKV infection during pregnancy was performed from January 2016-June 2017. We included patients admitted to UPH Neonatal Intensive Care Unit or referred for follow-up at UPH High Risk Clinics. The database was approved by the University of Puerto Rico, Medical Sciences Campus, IRB. RESULTS: 191 infants born to ZIKV positive mothers during pregnancy were identified. Normal head sonogram was found in 93% of the normo cephalic infants. Ocular findings were reported in 50% of the patients with microcephaly and 31% of the normo-cephalics. Fifteen newborns (7.8%) presented with microcephaly, of which 73% showed calcifications in head sonogram, and had severe anomalies on brain MRI. Auditory brainstem response test was performed on all newborns, 80% were within normal limits. CONCLUSION: Among the group of infants born to mothers with Zika positive test 4% had microcephaly. Of concern to us is the fact that 31% of normo cephalic infants had ocular manifestations and 7% of them had findings on head sonogram. While microcephaly is the severest form of Congenital Zika Syndrome, ocular manifestations might characterize the spectrum of disease. These findings reiterate the importance of detailed neonatal evaluations of exposed infants.

Association of Zika Virus Exposure in Utero with Ocular Phenotypes in a Group of Newborns in Puerto Rico Exposure with ocular phenotypes in newborns in Puerto Rico.

OBJECTIVE: After the outbreak in January 2016, researchers in Brazil reported the first cases of ophthalmic abnormalities in infants with microcephaly and presumed Zika virus (ZIKV) intrauterine infection. Screening for ocular lesions in all newborns exposed to ZIKV prenatally has been emphasized because of a chorioretinal macular scar found in a number normocephalic patient exposed to ZIKV. METHODS: A retrospective review of the medical records of infants born to mothers found to be ZIKV-positive during their pregnancies (January 2016-July 2017) was performed. We included all newborns and infants admitted to the NICU and/or receiving ambulatory care at the high-risk clinics at the University Pediatric Hospital, San Juan, Puerto Rico. The creation of this database was approved by the IRB of the University of Puerto Rico, Medical Sciences Campus. RESULTS: Records from 234 newborns born to Zika-positive mothers were identified. The ophthalmic evaluations of 95 patients were available. Sixty-three of them had normal findings (66%). Twelve of the 95 patients had microcephaly (12.6% of the cohort). Of the microcephalic group, half had normal ophthalmological findings. Of the normocephalic group (83/95), almost 31% had ocular findings: a small optic nerve in 3%, a double-ring sign in 10%, macular stippling in 8%, mottling in 4%, an oval optic nerve in 2%, a tilted nerve in 1%, and blunted foveal reflex in 1%. CONCLUSION: We report herein ZIKV associated ophthalmologic findings, similar to those published, in 34% of the infants with suspected/confirmed ZIKV fetal exposure. These data underline the importance of ocular examination in patients with suspected congenital Zika but without other findings on physical exam.

Docosahexaenoic acid protection in a rotenone induced Parkinson's model: Prevention of tubulin and synaptophysin loss, but no association with mitochondrial function.

Rotenone, a classic mitochondrial complex I inhibitor, leads to dopaminergic neuronal death resulting in a Parkinson's-like-disease. Docosahexaenoic acid (DHA) has shown neuroprotective effects in other experimental models of Parkinson's disease, but its effect on the rotenone-induced parkinsonism is still unknown. We tested whether DHA in vivo exerts a neuroprotective effect on rotenone-induced parkinsonism and explored the mechanisms involved, including mitochondrial function and ultrastructure as well as the expression of tubulin and synaptophysin. We pretreated eighty male Wistar rats with DHA (35 mg/kg/day) for seven days and then administered rotenone for eight days. We then measured rearing behavior, number of dopaminergic neurons, tyrosine hydroxylase content, tubulin and synaptophysin expression, mitochondrial complex I, respiratory control ratio, mitochondrial transmembrane potential, ATP production activity and mitochondrial ultrastructure. We found that in vivo DHA supply exerted a neuroprotective effect, evidenced by decreased dopaminergic neuron cell death. Although we detected rotenone induced mitochondrial ultrastructure alterations, these were not associated with mitochondrial dysfunction. Rotenone had no effect on mitochondrial complex I, respiratory control ratio, mitochondrial transmembrane potential or ATP production activity. DHA also prevented a rotenone-induced decrease in tubulin and synaptophysin expression. Our results support the neuroprotective effect of DHA on rotenone-induced parkinsonism, and a possible effect on early stage Parkinson's disease. This protective effect is not associated with mitochondrial function improvement, but rather with preventing loss of tubulin and synaptophysin, proteins relevant to synaptic transmission.

Alveolar bone resorption and Th1/Th17-associated immune response triggered during Aggregatibacter actinomycetemcomitans-induced experimental periodontitis are serotype-dependent.

BACKGROUND: Aggregatibacter actinomycetemcomitans expresses several virulence factors that may contribute to the pathogenesis of periodontitis. Based on the antigenicity of the O-polysaccharide component of the lipopolysaccharide (LPS), different A. actinomycetemcomitans serotypes have been described. Among them, serotype b has demonstrated a stronger capacity to trigger Th1 and Th17-associated cytokine, CC-chemokine, and CC-chemokine receptor production on immune cells in vitro. With a murine model of experimental periodontitis, this investigation aimed to analyze the alveolar bone resorption and the pattern of immune response triggered by the different A. actinomycetemcomitans serotypes within periodontal lesions. METHODS: For periodontal lesion induction, mice were orally infected with the different A. actinomycetemcomitans serotypes or their purified LPS. Alveolar bone resorption was analyzed using microcomputed tomography and scanning electron microscopy. Bacterial infection, receptor activator of nuclear factor-kappa B ligand (RANKL) and Th1 and Th17-associated cytokine, CC-chemokine, and CC-chemokine receptor levels were quantified by quantitative polymerase chain reaction (qPCR). T lymphocytes isolated from periodontal lesions were analyzed by flow cytometry. RESULTS: In periodontal lesions, serotype b of A. actinomycetemcomitans induced higher alveolar bone resorption and expression of RANKL compared with the other serotypes. In addition, serotype b induced greater levels of Th1- and Th17-related cytokines, CC-chemokines, and CC-chemokine receptors than the others. Similarly, higher numbers of infiltrating Th1 and Th17 lymphocytes were detected in serotype b-induced periodontal lesions. CONCLUSIONS: These results demonstrate that periodontal lesions induced with different A. actinomycetemcomitans serotypes elicited distinct alveolar bone resorption and immune response. In particular, serotype b was more pathogenic than the others and induced stronger Th1 and Th17 patterns of immune responses during experimental periodontitis.

Serotype a of Aggregatibacter actinomycetemcomitans down-regulates the increased serotype b-induced cytokine and chemokine production in dendritic cells.

OBJECTIVE: In Aggregatibacter actinomycetemcomitans, different serotypes have been described based on LPS antigenicity. Mixed infection with the different A. actinomycetemcomitans serotypes is frequent in periodontitis patients; accordingly, the role of this bacterial species in the pathogenesis of periodontitis may differ depending whether patients or periodontal lesions harbour one or more of the A. actinomycetemcomitans serotypes. We hypothesized that different combinations of these serotypes could be associated with distinct host responses and hence different inflammatory patterns. This investigation was aimed to assess whether the increased immuno-stimulatory potential attributed to the serotype b of A. actinomycetemcomitans on immune cells is able to be modified during co-infection with other A. actinomycetemcomitans serotypes. METHODS: Dendritic cells (DCs) were obtained from healthy subjects and stimulated with the different A. actinomycetemcomitans serotypes or their purified LPS using the following stimulatory conditions: serotype a, b, or c, and the combinations a+b, a+c, b+c, or a+b+c. The cytokine, CCR, and CCL levels were quantified by qPCR and ELISA. RESULTS: Higher levels of cytokines, CCRs, and CCLs were induced when DCs were stimulated with the serotype b of A. actinomycetemcomitans compared with the same cells stimulated with the other serotypes. When DCs were co-infected, these levels decreased in comparison with the serotype b-stimulation alone, in particular when the serotype a was present in the mixed infection. CONCLUSIONS: The increased immuno-stimulatory potential attributed to the serotype b was modified when DCs were co-infected with other A. actinomycetemcomitans serotypes, in particular, when the serotype a was present, the DC response diminished.

Regulatory T Lymphocytes in Periodontitis: A Translational View.

Periodontitis is a chronic immuno-inflammatory disease in which the disruption of the balance between host and microbiota interactions is key to the onset and progression of the disease. The immune homeostasis associated with periodontal health requires a regulated immuno-inflammatory response, during which the presence of regulatory T cells (Tregs) is essential to ensure a controlled response that minimizes collateral tissue damage. Since Tregs modulate both innate and adaptive immunity, pathological conditions that may resolve by the acquisition of immuno-tolerance, such as periodontitis, may benefit by the use of Treg immunotherapy. In recent years, many strategies have been proposed to take advantage of the immuno-suppressive capabilities of Tregs as immunotherapy, including the ex vivo and in vivo manipulation of the Treg compartment. Ongoing research in both basic and translational studies let us gain a better understanding of the diversity of Treg subsets, their phenotypic plasticity, and suppressive functions, which can be used as a substrate for new immunotherapies. Certainly, as our knowledge of Treg biology increases, we will be capable to develop new therapies designed to enhance the stability and function of Tregs during periodontitis.

Role of dendritic cells in peanut allergy.

INTRODUCTION: The prevalence of peanut allergy (PA) has increased, affecting approximately 1.1% of children in Western countries. PA causes life-threatening anaphylaxis and frequently persists for life. There are no standardized curative therapies for PA, and avoidance of peanuts remains the main therapeutic option. A better understanding of the pathogenesis of PA is essential to identify new treatment strategies. Intestinal dendritic cells (DCs) are essential in the induction and maintenance of food tolerance because they present dietary allergens to T cells, thereby directing subsequent immune responses. Areas covered: In this review, we discuss the factors related to the acquisition of oral tolerance to peanut proteins. We focus on intestinal DC-related aspects, including the latest advances in the biology of intestinal DC subtypes, effect of tolerance-inducing factors on DCs, effect of dietary components on oral tolerance, and role of DCs in peanut sensitization. Expert commentary: Given the increasing prevalence of PA, difficulty of avoiding peanut products, and the potentially serious accidental reactions, the development of novel therapies for PA is needed. The ability of DCs to trigger tolerance or immunity makes them an interesting target for new treatment strategies against PA.