Pharmacological enhancement of cholinergic neurotransmission alleviates neuroinflammation and improves functional outcomes in a triple transgenic mouse model of Alzheimer's disease.

Introduction: Alzheimer's disease (AD) is the most common neurodegenerative disorder affecting the elderly population worldwide. Due to the multifactorial nature of the disease, involving impairment of cholinergic neurotransmission and immune system, previous attempts to find effective treatments have faced challenges. Methods: In such scenario, we attempted to investigate the effects of alpha-glyceryl-phosphoryl-choline (α-GPC), a cholinomimetic molecule, on neuroinflammation and memory outcome in the triple transgenic mouse model of AD (3xTg-AD). Mice were enrolled at 4 months of age, treated orally with α-GPC dissolved in drinking water at a concentration resulting in an average daily dose of 100 mg/kg for 8 months and sacrificed at 12 months of age. Thereafter, inflammatory markers, as well as cognitive parameters, were measured. Results: Chronic α-GPC treatment reduced accumulation of amyloid deposits and led to a substantial re-balance of the inflammatory response of resident innate immune cells, astrocytes and microglia. Specifically, fluorescent immunohistochemistry and Western blot analysis showed that α-GPC contributed to reduction of cortical and hippocampal reactive astrocytes and pro-inflammatory microglia, concurrently increasing the expression of anti-inflammatory molecules. Whereas α-GPC beneficially affect the synaptic marker synaptophysin in the hippocampus. Furthermore, we observed that α-GPC was effective in restoring cognitive dysfunction, as measured by the Novel Object Recognition test, wherein 3xTg-AD mice treated with α-GPC significantly spent more time exploring the novel object compared to 3xTg-AD untreated mice. Discussion: In conclusion, chronic treatment with α-GPC exhibited a significant anti-inflammatory activity and sustained the key function of hippocampal synapses, crucial for the maintenance of a regular cognitive status. In light of our results, we suggest that α-GPC could be exploited as a promising therapeutic approach in early phases of AD.

Taming Microglia in Alzheimer's Disease: Exploring Potential Implications of Choline Alphoscerate via α7 nAChR Modulation.

Alzheimer's disease (AD), marked by cognitive impairment, predominantly affects the brain regions regulated by cholinergic innervation, such as the cerebral cortex and hippocampus. Cholinergic dysfunction, a key contributor to age-related cognitive decline, has spurred investigations into potential therapeutic interventions. We have previously shown that choline alphoscerate (α-GPC), a cholinergic neurotransmission-enhancing agent, protects from Aß-mediated neurotoxicity. Herein, we investigated the effects of α-GPC on the microglial phenotype in response to Aß via modulation of the nicotinic alpha-7 acetylcholine receptor (α7 nAChR). BV2 microglial cells were pre-treated for 1 h with α-GPC and were treated for 24, 48, and 72 h with Aß1-42 and/or α-BTX, a selective α7nAchR antagonist. Fluorescent immunocytochemistry and Western blot analysis showed that α-GPC was able to antagonize Aß-induced inflammatory effects. Of note, α-GPC exerted its anti-inflammatory effect by directly activating the α7nAChR receptor, as suggested by the induction of an increase in [Ca2+]i and Ach-like currents. Considering that cholinergic transmission appears crucial in regulating the inflammatory profiles of glial cells, its modulation emerges as a potential pharmaco-therapeutic target to improve outcomes in inflammatory neurodegenerative disorders, such as AD.

Systematic Review of Zinc's Benefits and Biological Effects on Oral Health.

BACKGROUND: This review was based on the following question: "What is the state-of-the-art regarding the effect of zinc exposure in the oral cavity on a population of adults and children, compared to dental products containing materials other than zinc, considering in vivo (clinical trials and observational studies) and in vitro studies?" according to a PICOS strategy format. This study aims to analyze zinc application in dental materials, with different compositions and chemical formulations, considering how mechanical and biological properties may influence its clinical applicability. METHODS: In vivo (clinical trials: controlled clinical trials (CCTs) and randomized controlled trials (RCTs); and observational studies: case control and cohort studies) trials or in vitro studies published in English or Italian during the last 10 years on children and adult patients with zinc exposure were included by three different reviewers using the MEDLINE (via PubMed), Scopus, and Web of Science electronic databases. RESULTS: Titles and abstracts were evaluated following the eligibility criteria. The full texts of eligible studies were then reviewed against the inclusion/exclusion criteria. Scientific and technical information of the 33 included studies were collected into evidence tables, reporting data on in vivo and in vitro studies. A narrative approach was adopted. CONCLUSIONS: Antibacterial activity was found to be the most studied property of zinc, but further investigations are needed to establish adjuvant zinc therapies in patients with oral disease.

Disease Modifying Strategies in Multiple Sclerosis: New Rays of Hope to Combat Disability?

Multiple sclerosis (MS) is the most prevalent chronic autoimmune inflammatory- demyelinating disorder of the central nervous system (CNS). It usually begins in young adulthood, mainly between the second and fourth decades of life. Usually, the clinical course is characterized by the involvement of multiple CNS functional systems and by different, often overlapping phenotypes. In the last decades, remarkable results have been achieved in the treatment of MS, particularly in the relapsing- remitting (RRMS) form, thus improving the long-term outcome for many patients. As deeper knowledge of MS pathogenesis and respective molecular targets keeps growing, nowadays, several lines of disease-modifying treatments (DMT) are available, an impressive change compared to the relative poverty of options available in the past. Current MS management by DMTs is aimed at reducing relapse frequency, ameliorating symptoms, and preventing clinical disability and progression. Notwithstanding the relevant increase in pharmacological options for the management of RRMS, research is now increasingly pointing to identify new molecules with high efficacy, particularly in progressive forms. Hence, future efforts should be concentrated on achieving a more extensive, if not exhaustive, understanding of the pathogenetic mechanisms underlying this phase of the disease in order to characterize novel molecules for therapeutic intervention. The purpose of this review is to provide a compact overview of the numerous currently approved treatments and future innovative approaches, including neuroprotective treatments as anti-LINGO-1 monoclonal antibody and cell therapies, for effective and safe management of MS, potentially leading to a cure for this disease.

Insight into Risk Factors, Pharmacogenetics/Genomics, and Management of Adverse Drug Reactions in Elderly: A Narrative Review.

The European Medicine Agency (EMA) has defined Adverse Drug Reactions (ADRs) as "a noxious and unintended response to a medicine", not including poisoning, accidental, or intentional overdoses. The ADR occurrence differs based on the approach adopted for defining and detecting them, the characteristics of the population under study, and the research setting. ADRs have a significant impact on morbidity and mortality, particularly among older adults, and represent a financial burden for health services. Between 30% and 60% of ADRs might be predictable and preventable, emerging as a result of inappropriate prescription, drug chemistry inherent toxicity, cell-specific drug toxicity, age- and sex-related anomalies in drug absorption, distribution, metabolism, and elimination (ADME), and drug-drug interactions (DDIs) in combination therapies or when a patient is treated with different drugs for concomitant disorders. This is particularly important in chronic diseases which require long-term treatments. Rapid developments in pharmacogenetics/genomics have improved the understanding of ADRs accompanied by more accurate prescriptions and reduction in unnecessary costs. To alleviate the burden of ADRs, especially in the elderly, interventions focused on pharmaceutical principles, such as medication review and reconciliation, should be integrated into a broader assessment of patients' characteristics, needs, and health priorities. Digital health interventions could offer valuable solutions to assist healthcare professionals in identifying inappropriate prescriptions and promoting patient adherence to pharmacotherapies.

TRAIL-R Deficient Mice Are Protected from Neurotoxic Effects of Amyloid-ß.

TRAIL, a member of TNF superfamily, is a potent inducer of neuronal death. Neurotoxic effects of TRAIL appear mediated by its death receptor TRAIL-R2/DR5. To assess the role of TRAIL/TRAIL-R2 pathway in AD-related neurodegeneration, we studied the impact of the treatment with amyloid-ß (Aß) upon cell viability and inflammation in TRAIL-R-deficient mice (TRAIL-R-/-). Here, we demonstrate that the lack of TRAIL-R2 protects from death cultured TRAIL-R-/- mouse embryonic hippocampal cells after treatment with either Aß1-42 or TRAIL. Consistently, stereotaxic injection of Aß1-42 resulted in blunted caspase activation, as well as in reduction of JNK phosphorylation and increased AKT phosphorylation in TRAIL-R-/- mice. Moreover, the lack of TRAIL-R2 was associated with blunted constitutive p53 expression in mice that have undergone Aß1-42 treatment, as well as in decrease of phosphorylated forms of tau and GSK3ß proteins. Likewise, TRAIL-R2 appears essential to both TRAIL and Aß-mediated neurotoxicity and inflammation. Indeed, hippocampi of TRAIL-R-/- mice challenged with Aß1-42, showed a slight expression of microglial (Iba-1) and astrocytic (GFAP) markers along with attenuated levels of IL-1ß, TNF-α, NOS2 and COX2. In conclusion, the bulk of these results demonstrate that the constitutive lack of TRAIL-R2 is associated with a substantial reduction of noxious effects of Aß1-42, providing further evidence on the prominent role played by TRAIL in course of Aß-related neurodegeneration and confirming that the TRAIL system represents a potential target for innovative AD therapy.

Role of Microglia and Astrocytes in Alzheimer's Disease: From Neuroinflammation to Ca2+ Homeostasis Dysregulation.

Alzheimer's disease (AD) is the most common form of dementia worldwide, with a complex, poorly understood pathogenesis. Cerebral atrophy, amyloid-ß (Aß) plaques, and neurofibrillary tangles represent the main pathological hallmarks of the AD brain. Recently, neuroinflammation has been recognized as a prominent feature of the AD brain and substantial evidence suggests that the inflammatory response modulates disease progression. Additionally, dysregulation of calcium (Ca2+) homeostasis represents another early factor involved in the AD pathogenesis, as intracellular Ca2+ concentration is essential to ensure proper cellular and neuronal functions. Although growing evidence supports the involvement of Ca2+ in the mechanisms of neurodegeneration-related inflammatory processes, scant data are available on its contribution in microglia and astrocytes functioning, both in health and throughout the AD continuum. Nevertheless, AD-related aberrant Ca2+ signalling in astrocytes and microglia is crucially involved in the mechanisms underpinning neuroinflammatory processes that, in turn, impact neuronal Ca2+ homeostasis and brain function. In this light, we attempted to provide an overview of the current understanding of the interactions between the glia cells-mediated inflammatory responses and the molecular mechanisms involved in Ca2+ homeostasis dysregulation in AD.

Experience and Prevalence of Dental Caries in Migrant and Nonmigrant Low-SES Families' Children Aged 3 to 5 Years in Italy.

Dental caries is a public health problem in children and is more prevalent in low-socioeconomic-status groups. The aim of this study is to assess the association between migrant families and the prevalence of caries among young children in Italy. This is a cross-sectional study. In the age range of 3 to 5 years, a total of 266 migrant children and 301 nonmigrant children were examined in three Italian charity dental centers. All children had families with low SES. The dmft was determined by intraoral examination performed by six pediatric dentist specialists to assess their dental health. In this study, the prevalence of caries (71%) and the mean dmft (3.68; SD: 1.52) of migrant children were statistically significantly higher than the percentage (52%) and the mean dmft of the nonmigrant control group (3.10; SD: 1.65) with no differences between genders. For the migrant children, the mean (0.49; SD: 0.32) of restorations (filled teeth) was statistically significantly lower than that of nonmigrant children (1.20; SD: 0.48). This study highlights that dmft values and the prevalence of caries are higher in migrant children than in nonmigrant children. In addition, the control group shows a higher level of dental caries than the national mean.

Intranasal Administration of a TRAIL Neutralizing Monoclonal Antibody Adsorbed in PLGA Nanoparticles and NLC Nanosystems: An In Vivo Study on a Mouse Model of Alzheimer's Disease.

Alzheimer's disease (AD) is a neurodegenerative disorder that progressively compromises cognitive functions. Tumor necrosis factor (TNF)-Related Apoptosis Inducing Ligand (TRAIL), a proinflammatory cytokine belonging to the TNF superfamily, appears to be a key player in the inflammatory/immune orchestra of the AD brain. Despite the ability of an anti-TRAIL monoclonal antibody to reach the brain producing beneficial effects in AD mice, we attempted to develop such a TRAIL-neutralizing monoclonal antibody adsorbed on lipid and polymeric nanocarriers, for intranasal administration, in a valid approach to overcome issues related to both high dose and drug transport across the blood-brain barrier. The two types of nanomedicines produced showed physico-chemical characteristics appropriate for intranasal administration. As confirmed by enzyme-linked immunosorbent assay (ELISA), both nanomedicines were able to form a complex with the antibody with an encapsulation efficiency of ≈99%. After testing in vitro the immunoneutralizing properties of the nanomedicines, the latter were intranasally administered in AD mice. The antibody-nanocarrier complexes were detectable in the brain in substantial amounts at concentrations significantly higher compared to the free form of the anti-TRAIL antibody. These data support the use of nanomedicine as an optimal method for the delivery of the TRAIL neutralizing antibody to the brain through the nose-to-brain route, aiming to improve the biological attributes of anti-TRAIL-based therapy for AD treatment.

Targeting the miRNA-155/TNFSF10 network restrains inflammatory response in the retina in a mouse model of Alzheimer's disease.

Age-related disorders, such as Alzheimer's disease (AD) and age-related macular degeneration (AMD) share common features such as amyloid-ß (Aß) protein accumulation. Retinal deposition of Aß aggregates in AMD patients has suggested a potential link between AMD and AD. In the present study, we analyzed the expression pattern of a focused set of miRNAs, previously found to be involved in both AD and AMD, in the retina of a triple transgenic mouse model of AD (3xTg-AD) at different time-points. Several miRNAs were differentially expressed in the retina of 3xTg-AD mice, compared to the retina of age-matched wild-type (WT) mice. In particular, bioinformatic analysis revealed that miR-155 had a central role in miRNA-gene network stability, regulating several pathways, including apoptotic and inflammatory signaling pathways modulated by TNF-related apoptosis-inducing ligand (TNFSF10). We showed that chronic treatment of 3xTg-AD mice with an anti-TNFSF10 monoclonal antibody was able to inhibit the retinal expression of miR-155, which inversely correlated with the expression of its molecular target SOCS-1. Moreover, the fine-tuned mechanism related to TNFSF10 immunoneutralization was tightly linked to modulation of TNFSF10 itself and its death receptor TNFRSF10B, along with cytokine production by microglia, reactive gliosis, and specific AD-related neuropathological hallmarks (i.e., Aß deposition and Tau phosphorylation) in the retina of 3xTg-AD mice. In conclusion, immunoneutralization of TNFSF10 significantly preserved the retinal tissue in 3xTg-AD mice, suggesting its potential therapeutic application in retinal degenerative disorders.

Stem Cells: Innovative Therapeutic Options for Neurodegenerative Diseases?

Neurodegenerative diseases are characterized by the progressive loss of structure and/or function of both neurons and glial cells, leading to different degrees of pathology and loss of cognition. The hypothesis of circuit reconstruction in the damaged brain via direct cell replacement has been pursued extensively so far. In this context, stem cells represent a useful option since they provide tissue restoration through the substitution of damaged neuronal cells with exogenous stem cells and create a neuro-protective environment through the release of bioactive molecules for healthy neurons, as well. These peculiar properties of stem cells are opening to potential therapeutic strategies for the treatment of severe neurodegenerative disorders, for which the absence of effective treatment options leads to an increasingly socio-economic burden. Currently, the introduction of new technologies in the field of stem cells and the implementation of alternative cell tissues sources are pointing to exciting frontiers in this area of research. Here, we provide an update of the current knowledge about source and administration routes of stem cells, and review light and shadows of cells replacement therapy for the treatment of the three main neurodegenerative disorders (Amyotrophic lateral sclerosis, Parkinson's, and Alzheimer's disease).

Brimonidine is Neuroprotective in Animal Paradigm of Retinal Ganglion Cell Damage.

To investigate the neuroprotective effect of brimonidine after retinal ischemia damage on mouse eye. Glaucoma is an optic neuropathy characterized by retinal ganglion cells (RGCs) death, irreversible peripheral and central visual field loss, and high intraocular pressure. Ischemia reperfusion (I/R) injury model was used in C57BL/6J mice to mimic conditions of glaucomatous neurodegeneration. Mouse eyes were treated topically with brimonidine and pattern electroretinogram were used to assess the retinal ganglion cells (RGCs) function. A wide range of inflammatory markers, as well as anti-inflammatory and neurotrophic molecules, were investigated to figure out the potential protective effects of brimonidine in mouse retina. In particular, brain-derived neurotrophic factor (BDNF), IL-6, tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and its death receptor DR-5, TNF-α, GFAP, Iba-1, NOS, IL-1ß and IL-10 were assessed in mouse retina that underwent to I/R insult with or without brimonidine treatment. Brimonidine provided remarkable RGCs protection in our paradigm. PERG amplitude values were significantly (p < 0.05) higher in brimonidine-treated eyes in comparison to I/R retinas. Retinal BDNF mRNA levels in the I/R group dropped significantly (p < 0.05) compared to the control group (normal mice); brimonidine treatment counteracted the downregulation of retinal BDNF mRNA in I/R eyes. Retinal inflammatory markers increased significantly (p < 0.05) in the I/R group and brimonidine treatment was able to revert that. The anti-inflammatory IL-10 decreased significantly (p < 0.05) after retinal I/R insult and increased significantly (p < 0.05) in the group treated with brimonidine. In conclusion, brimonidine was effective in preventing loss of function of RGCs and in regulating inflammatory biomarkers elicited by retinal I/R injury.

Beneficial Effects of Choline Alphoscerate on Amyloid-ß Neurotoxicity in an In vitro Model of Alzheimer's Disease.

BACKGROUND: Alzheimer's disease (AD) is the most common form of neurodegenerative disorder characterized by cognitive impairment, which represents an urgent public health concern. Given the worldwide impact of AD, there is a compelling need for effective therapies to slow down or halt this disorder. OBJECTIVE: Choline alphoscerate (α-GPC) represents a potentially effective cholinergic neurotransmission enhancing agent with an interesting clinical profile in cognitive dysfunctions improvement, although only scanty data are available about the mechanisms underlying such beneficial effects. METHODS: The SH-SY5Y neuronal cell line, differentiated for 1 week with 10 µm of all-trans-retinoic acid (RA), to achieve a switch towards a cholinergic phenotype, was used as an in vitro model of AD. SH-SY5Y cells were pre-treated for 1h with α-GPC (100nM) and treated for 72 h with Aß25-35 (10µM). RESULTS: α-GPC was able to antagonize Aß25-35 mediated neurotoxicity and attenuate the Aß-induced phosphorylation of the Tau protein. Moreover, α-GPC exerted its beneficial effects by employing the NGF/TrkA system, knocked down in AD and, consequently, by sustaining the expression level of synaptic vesicle proteins, such as synaptophysin. CONCLUSION: Taken together, our data suggest that α-GPC can have a role in neuroprotection in the course of toxic challenges with Aß. Thus, a deeper understanding of the mechanism underlying its beneficial effect, could provide new insights into potential future pharmacological applications of its functional cholinergic enhancement, with the aim to mitigate AD and could represent the basis for innovative therapy.

Repositioning of Immunomodulators: A Ray of Hope for Alzheimer's Disease?

Alzheimer's disease (AD) is the most common age-related neurodegenerative disorder characterized by cognitive decline and by the presence of amyloid ß plaques and neurofibrillary tangles in the brain. Despite recent advances in understanding its pathophysiological mechanisms, to date, there are no disease-modifying therapeutic options, to slow or halt the evolution of neurodegenerative processes in AD. Current pharmacological treatments only transiently mitigate the severity of symptoms, with modest or null overall improvement. Emerging evidence supports the concept that AD is affected by the impaired ability of the immune system to restrain the brain's pathology. Deep understanding of the relationship between the nervous and the immune system may provide a novel arena to develop effective and safe drugs for AD treatment. Considering the crucial role of inflammatory/immune pathways in AD, here we discuss the current status of the immuno-oncological, immunomodulatory and anti-TNF-α drugs which are being used in preclinical studies or in ongoing clinical trials by means of the drug-repositioning approach.

The immune system on the TRAIL of Alzheimer's disease.

Alzheimer's disease (AD) is the most common form of dementia, characterized by progressive degeneration and loss of neurons in specific regions of the central nervous system. Chronic activation of the immune cells resident in the brain, peripheral immune cell trafficking across the blood-brain barrier, and release of inflammatory and neurotoxic factors, appear critical contributors of the neuroinflammatory response that drives the progression of neurodegenerative processes in AD. As the neuro-immune network is impaired in course of AD, this review is aimed to point out the essential supportive role of innate and adaptive immune response either in normal brain as well as in brain recovery from injury. Since a fine-tuning of the immune response appears crucial to ensure proper nervous system functioning, we focused on the role of the TNF superfamily member, TNF-related apoptosis-inducing ligand (TRAIL), which modulates both the innate and adaptive immune response in the pathogenesis of several immunological disorders and, in particular, in AD-related neuroinflammation. We here summarized mounting evidence of potential involvement of TRAIL signaling in AD pathogenesis, with the aim to provide clearer insights about potential novel therapeutic approaches in AD.

Ocular Pharmacological Profile of Hydrocortisone in Dry Eye Disease.

To investigate the ocular pharmacological profile of hydrocortisone (HC) using in vitro and in vivo models of dry eye disease. Rabbit corneal epithelial cells (SIRCs) were used to assess the effect of HC in two paradigms of corneal damage: hyperosmotic stress and scratch-wound assay. Dry eye was induced in albino rabbits by topical administration of atropine sulfate or by injection of concanavalin A (ConA) into the lacrimal gland. TNFα, TNF-related apoptosis-inducing ligand (TRAIL), IL-1ß, and IL-8 were determined by ELISA or western blot in a corneal damage hyperosmotic in vitro model, with or without HC treatment. Inflammatory biomarkers, such as TNFα, IL-8, and MMP-9, were evaluated in tears of rabbit eye injected with ConA and treated with HC. Tear volume and tear film integrity, in both in vivo models, were evaluated by the Schirmer test and tear break-up time (TBUT). Ocular distribution of four formulations containing HC (0.001%, 0.003%, 0.005%, and 0.33%) was performed in the rabbit eye. Aqueous humor samples were collected after 15, 30, 60, and 90 min from instillation and then detected by LC-MS/MS. Hyperosmotic insult significantly activated protein expression of inflammatory biomarkers, which were significantly modulated by HC treatment. HC significantly enhanced the re-epithelialization of scratched SIRCs. Treatment with HC eye drops significantly reduced the tear concentrations of TNF-α, IL-8, and MMP-9 vs. vehicle in the ConA dry eye model. Moreover, HC significantly restored the tear volume and tear film integrity to levels of the control eyes, both in ConA- and atropine-induced dry eye paradigms. Finally, we demonstrated that HC crossed, in a dose-dependent manner, the corneal barrier when the eyes were topically treated with HC formulations (dose range 0.003-0.33%). No trace of HC was detected in the aqueous humor after ocular administration of eye drops containing the lowest dose of the drug (0.001%), indicating that, at this very low concentration, the drug did not pass the corneal barrier avoiding potential side effects such as intraocular pressure rise. Altogether, these data suggest that HC, at very low concentrations, has an important anti-inflammatory effect both in vitro and in vivo dry eye paradigms and a good safety profile.

Beneficial effects of curtailing immune susceptibility in an Alzheimer's disease model.

BACKGROUND: Currently, there are no effective therapeutic options for Alzheimer's disease, the most common, multifactorial form of dementia, characterized by anomalous amyloid accumulation in the brain. Growing evidence points to neuroinflammation as a major promoter of AD. We have previously shown that the proinflammatory cytokine TNFSF10 fuels AD neuroinflammation, and that its immunoneutralization results in improved cognition in the 3xTg-AD mouse. METHODS: Here, we hypothesize that inflammatory hallmarks of AD might parallel with central and peripheral immune response dysfunction. To verify such hypothesis, we used a triple transgenic mouse model of AD. 3xTg-AD mice were treated for 12 months with an anti-TNFSF10 antibody, and thereafter immune/inflammatory markers including COX2, iNOS, IL-1ß and TNF-α, CD3, GITR, and FoxP3 (markers of regulatory T cells) were measured in the spleen as well as in the hippocampus. RESULTS: Spleens displayed accumulation of amyloid-ß1-42 (Aß1-42), as well as high expression of Treg cell markers FoxP3 and GITR, in parallel with the increased levels of inflammatory markers COX2, iNOS, IL-1ß and TNF-α, and blunted IL-10 expression. Moreover, CD3 expression was increased in the hippocampus, consistently with FoxP3 and GITR. After chronic treatment of 3xTg-AD mice with an anti-TNFSF10 antibody, splenic FoxP3, GITR, and the above-mentioned inflammatory markers expression was restored to basal levels, while expression of IL-10 was increased. A similar picture was observed in the hippocampus. Such improvement of peripheral and CNS inflammatory/immune response was associated with decreased microglial activity in terms of TNFα production, as well as decreased expression of both amyloid and phosphorylated tau protein in the hippocampus of treated 3xTg-AD mice. Interestingly, we also reported an increased expression of both CD3 and FoxP3, in sections from human AD brain. CONCLUSIONS: We suggest that neuroinflammation in the brain of 3xTg-AD mice triggered by TNFSF10 might result in a more general overshooting of the immune response. Treatment with an anti-TNFSF10 antibody blunted inflammatory processes both in the spleen and hippocampus. These data confirm the detrimental role of TNFSF10 in neurodegeneration, and corroborate the hypothesis of the anti-TNFSF10 strategy as a potential treatment to improve outcomes in AD.

Tumor necrosis factor-related apoptosis-inducing ligand reduces the expression of the neuroprotective Na+ /Ca2+ exchanger isoform NCX3 in human neuroblastoma SH-SY5Y cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL), a cytokine belonging to the TNF superfamily, is regarded as a mediator of neurotoxicity. The constitutively expressed ion exchanger Na+ /Ca2+ exchanger isoform-3 (NCX3) has been shown to protect neurons from injury. Its expression is induced by nerve growth factor (NGF) through activation of its tyrosine kinase receptor trkA. The latter, in turn, activates downstream kinases, such as extracellular signal-regulated kinase (ERK) and the survival-related kinase protein kinase B (AKT). Here, we verified whether TRAIL could influence the expression of NCX3 via modulation of the NGF/trkA system. Differentiated human neuroblastoma SH-SY5Y cells were incubated with TRAIL and, subsequently, the expression of the NCX3 protein was studied at different times by means of western blot analysis. Then, the expression of the phosphorylated forms of either trkA, ERK or AKT was analyzed at identical intervals. Western blot analysis revealed that the expression of NCX3 protein decreased in a time-dependent fashion in SH-SY5Y cells treated with TRAIL, to reach its minimum at 48 h. On the other hand, p-trkA, p-ERK, and p-AKT expression was increased in cells treated with TRAIL after 6 and 16 h; then it declined to nearly undetectable levels after 48 h. Results indicate that the increase in TRAIL expression occurring during neuronal damage may be responsible of NCX3 down-regulation and weakens its neuroprotective effects. The TRAIL system could thus represent a potential target for treatment of neuronal damage characterized by NCX3 function impairment.

Redundant modulatory effects of proinflammatory cytokines in human osteoblastic cells in vitro.

OBJECTIVES: The aim of our study was to investigate possible interaction of IL-17, TRAIL, and TNF-α in the modulation of osteoblast homeostasis in vitro, using human differentiated osteoblastic Saos-2 cells as in vitro model. METHODS: The effects of these cytokines on osteoblastic cell viability were assessed, by MTT assay, alone or in combination, at different times and concentrations. The effects of IL-17 and TNF-α on the regulatory system of osteoclast activity RANK/RANKL/ OPG were evaluated by Western blot and ELISA techniques in cell culture media. Quantitative expression of RANKL, OPG and pro-inflammatory factors were analysed at the mRNA level by quantitative real time RT-PCR. RESULTS: Effects of IL-17, TNF-α and TRAIL on osteoblastic cell viability indicated that IL-17 alone, or in combination with TNF-α did not alter Saos-2 cell viability. On the other hand, TRAIL, as expected, exhibited time- and concentration-dependent cytotoxicity. The expression both RANKL and OPG were increased at the mRNA level and protein release by IL-17 and TNF-α, either alone or in combination. The analysis of IL-17 and TNF-α on pro-inflammatory molecules mRNA expression, such as CXC family chemokines CXCL-1 and CXCL-5, COX-2 and IL-6 demonstrated an increase in these pro-inflammatory cytokines with cooperative effects of the combination. CONCLUSIONS: Overall, these results suggest that IL-17, TRAIL and TNF-α sustain bone tissue inflammation associated with decrease of calcified component. To do so, they act redundantly each other, to amplify the inflammatory response in the bone. In conclusion, unravelling novel molecular targets within the bone-cytokine network represents a platform for innovative treatment of bone diseases due to immunological diseases such as psoriatic arthritis.