Publications by authors named "Filippo Ugolini"

14 Publications

  • Page 1 of 1

Digital Immunophenotyping Predicts Disease Free and Overall Survival in Early Stage Melanoma Patients.

Cells 2021 Feb 17;10(2). Epub 2021 Feb 17.

Division of Pathological Anatomy, Papa Giovanni XXIII Hospital, 24127 Bergamo, Italy.

Background: the prognostic significance of tumor infiltrating lymphocytes (TILs) in intermediate/thick primary cutaneous melanoma (PCM) remains controversial, partially because conventional evaluation is not reliable, due to inter-observer variability and diverse scoring methods. We aimed to assess the prognostic impact of the density and spatial distribution of immune cells in early stage intermediate/thick PCM.

Materials And Methods: digital image acquisition and quantitative analysis of tissue immune biomarkers (CD3, CD4, CD8, CD68, PD-L1, CD163, FOX-P3, and PD-1) was carried out in a training cohort, which included patients with primary PCM ≥ 2 mm diagnosed, treated, and followed-up prospectively in three Italian centers. Results were validated in an independent Italian cohort.

Results: in the training cohort, 100 Stage II-III melanoma patients were valuable. At multivariable analysis, a longer disease free survival (DFS) was statistically associated with higher levels of CD4 intratumoral T-cells (aHR [100 cell/mm increase] 0.98, 95%CI 0.95-1.00, = 0.041) and CD163 inner peritumoral (aHR [high vs. low] 0.56, 95%CI 0.32-0.99, = 0.047). A statistically significant longer DFS (aHR [high-high vs. low-low] 0.52, 95%CI 0.28-0.99, = 0.047) and overall survival (OS) (aHR [high-high vs. low-low] 0.39, 95%CI 0.18-0.85, = 0.018) was found in patients with a high density of both intratumoral CD8 T-cells and CD68 macrophages as compared to those with low density of both intratumoral CD8 T-cells and CD68 macrophages. Consistently, in the validation cohort, patients with high density of both intratumoral CD8 and CD3 T-cells were associated to a statistically better DFS (aHR[high-high vs. low-low] 0.24, 95%CI 0.10-0.56, < 0.001) and those with high density of both intratumoral CD8 and CD68 were associated to a statistically longer OS (aHR[high-high vs. low-low] 0.28, 95%CI 0.09-0.86, = 0.025).

Conclusion: our findings suggest that a specific preexisting profile of T cells and macrophages distribution in melanomas may predict the risk of recurrence and death with potential implications for the stratification of stage II-III melanoma patients.
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http://dx.doi.org/10.3390/cells10020422DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7922113PMC
February 2021

Protective Effect of Adenosine A Receptor Agonist, BAY60-6583, Against Transient Focal Brain Ischemia in Rat.

Front Pharmacol 2020 11;11:588757. Epub 2021 Feb 11.

Department of Neuroscience, Psychology, Drug Research and Child Health (NEUROFARBA), Division of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Cerebral ischemia is a multifactorial pathology characterized first by an acute injury, due to excitotoxicity, followed by a secondary brain injury that develops hours to days after ischemia. During ischemia, adenosine acts as an endogenous neuroprotectant. Few studies have investigated the role of A receptor in brain ischemia because of the low potency of adenosine for it and the few selective ligands developed so far. A receptors are scarcely but widely distributed in the brain on neurons, glial and endothelial cells and on hematopoietic cells, lymphocytes and neutrophils, where they exert mainly anti-inflammatory effects, inhibiting vascular adhesion and inflammatory cells migration. Aim of this work was to verify whether chronic administration of the A agonist, BAY60-6583 (0.1 mg/kg i.p., twice/day), starting 4 h after focal ischemia induced by transient (1 h) Middle Cerebral Artery occlusion (tMCAo) in the rat, was protective after the ischemic insult. BAY60-6583 improved the neurological deficit up to 7 days after tMCAo. Seven days after ischemia BAY60-6583 reduced significantly the ischemic brain damage in cortex and striatum, counteracted ischemia-induced neuronal death, reduced microglia activation and astrocytes alteration. Moreover, it decreased the expression of TNF-α and increased that of IL-10 in peripheral plasma. Two days after ischemia BAY60-6583 reduced blood cell infiltration in the ischemic cortex. The present study indicates that A receptors stimulation can attenuate the neuroinflammation that develops after ischemia, suggesting that A receptors may represent a new interesting pharmacological target to protect from degeneration after brain ischemia.
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http://dx.doi.org/10.3389/fphar.2020.588757DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7905306PMC
February 2021

Space-Dependent Glia-Neuron Interplay in the Hippocampus of Transgenic Models of β-Amyloid Deposition.

Int J Mol Sci 2020 Dec 11;21(24). Epub 2020 Dec 11.

Department of Health Sciences, Section of Clinical Pharmacology and Oncology, University of Florence, Viale Pieraccini 6, 50139 Firenze, Italy.

This review is focused on the description and discussion of the alterations of astrocytes and microglia interplay in models of Alzheimer's disease (AD). AD is an age-related neurodegenerative pathology with a slowly progressive and irreversible decline of cognitive functions. One of AD's histopathological hallmarks is the deposition of amyloid beta (Aβ) plaques in the brain. Long regarded as a non-specific, mere consequence of AD pathology, activation of microglia and astrocytes is now considered a key factor in both initiation and progression of the disease, and suppression of astrogliosis exacerbates neuropathology. Reactive astrocytes and microglia overexpress many cytokines, chemokines, and signaling molecules that activate or damage neighboring cells and their mutual interplay can result in virtuous/vicious cycles which differ in different brain regions. Heterogeneity of glia, either between or within a particular brain region, is likely to be relevant in healthy conditions and disease processes. Differential crosstalk between astrocytes and microglia in CA1 and CA3 areas of the hippocampus can be responsible for the differential sensitivity of the two areas to insults. Understanding the spatial differences and roles of glia will allow us to assess how these interactions can influence the state and progression of the disease, and will be critical for identifying therapeutic strategies.
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http://dx.doi.org/10.3390/ijms21249441DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7763751PMC
December 2020

An Overview on the Differential Interplay Among Neurons-Astrocytes-Microglia in CA1 and CA3 Hippocampus in Hypoxia/Ischemia.

Front Cell Neurosci 2020 11;14:585833. Epub 2020 Nov 11.

Department of Health Sciences, Section of Clinical Pharmacology and Oncology, University of Florence, Florence, Italy.

Neurons have been long regarded as the basic functional cells of the brain, whereas astrocytes and microglia have been regarded only as elements of support. However, proper intercommunication among neurons-astrocytes-microglia is of fundamental importance for the functional organization of the brain. Perturbation in the regulation of brain energy metabolism not only in neurons but also in astrocytes and microglia may be one of the pathophysiological mechanisms of neurodegeneration, especially in hypoxia/ischemia. Glial activation has long been considered detrimental for survival of neurons, but recently it appears that glial responses to an insult are not equal but vary in different brain areas. In this review, we first take into consideration the modifications of the vascular unit of the glymphatic system and glial metabolism in hypoxic conditions. Using the method of triple-labeling fluorescent immunohistochemistry coupled with confocal microscopy (TIC), we recently studied the interplay among neurons, astrocytes, and microglia in chronic brain hypoperfusion. We evaluated the quantitative and morpho-functional alterations of the neuron-astrocyte-microglia triads comparing the hippocampal CA1 area, more vulnerable to ischemia, to the CA3 area, less vulnerable. In these contiguous and interconnected areas, in the same experimental hypoxic conditions, astrocytes and microglia show differential, finely regulated, region-specific reactivities. In both areas, astrocytes and microglia form triad clusters with apoptotic, degenerating neurons. In the neuron-astrocyte-microglia triads, the cell body of a damaged neuron is infiltrated and bisected by branches of astrocyte that create a microscar around it while a microglial cell phagocytoses the damaged neuron. These coordinated actions are consistent with the scavenging and protective activities of microglia. In hypoxia, the neuron-astrocyte-microglia triads are more numerous in CA3 than in CA1, further indicating their protective effects. These data, taken from contiguous and interconnected hippocampal areas, demonstrate that glial response to the same hypoxic insult is not equal but varies significantly. Understanding the differences of glial reactivity is of great interest to explain the differential susceptibility of hippocampal areas to hypoxia/ischemia. Further studies may evidence the differential reactivity of glia in different brain areas, explaining the higher or lower sensitivity of these areas to different insults and whether glia may represent a target for future therapeutic interventions.
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http://dx.doi.org/10.3389/fncel.2020.585833DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7686560PMC
November 2020

Ethanol neurotoxicity is mediated by changes in expression, surface localization and functional properties of glutamate AMPA receptors.

J Neurochem 2020 Oct 26. Epub 2020 Oct 26.

Department of Neuroscience, Psychology, Drug Research and Child Health (NeuroFarBa), Section of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Modifications in the subunit composition of AMPA receptors (AMPARs) have been linked to the transition from physiological to pathological conditions in a number of contexts, including EtOH-induced neurotoxicity. Previous work from our laboratory showed that EtOH withdrawal causes CA1 pyramidal cell death in organotypic hippocampal slices and changes in the expression of AMPARs. Here, we investigated whether changes in expression and function of AMPARs may be causal for EtOH-induced neurotoxicity. To this aim, we examined the subunit composition, localization and function of AMPARs in hippocampal slices exposed to EtOH by using western blotting, surface expression assay, confocal microscopy and electrophysiology. We found that EtOH withdrawal specifically increases GluA1 protein signal in total homogenates, but not in the post-synaptic density-enriched fraction. This is suggestive of overall increase and redistribution of AMPARs to the extrasynaptic compartment. At functional level, AMPA-induced calcium influx was unexpectedly reduced, whereas AMPA-induced current was enhanced in CA1 pyramidal neurons following EtOH withdrawal, suggesting that increased AMPAR expression may lead to cell death because of elevated excitability, and not for a direct contribution on calcium influx. Finally, the neurotoxicity caused by EtOH withdrawal was attenuated by the non-selective AMPAR antagonist 2,3-dioxo-6-nitro-1,2,3,4-tetrahydrobenzo[f]quinoxaline-7-sulfonamide disodium salt as well as by the selective antagonist of GluA2-lacking AMPARs 1-naphthyl acetyl spermine. We conclude that EtOH neurotoxicity involves changes in expression, surface localization and functional properties of AMPARs, and propose GluA2-lacking AMPARs as amenable specific targets for the development of neuroprotective drugs in EtOH-withdrawal syndrome.
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http://dx.doi.org/10.1111/jnc.15223DOI Listing
October 2020

TRPA1 Expression in Synovial Sarcoma May Support Neural Origin.

Biomolecules 2020 10 15;10(10). Epub 2020 Oct 15.

Section of Clinical Pharmacology and Oncology, Department of Health Sciences, University of Florence, 50139 Florence, Italy.

Synovial sarcoma (SS) is a malignant mesenchymal soft tissue neoplasm. Despite its name, the cells of origin are not synovial cells, but rather neural, myogenic, or multipotent mesenchymal stem cells have been proposed as possible cells originators. Unlike other sarcomas, an unusual presentation of long-term pain at the tumor site has been documented, but the exact mechanisms have not been fully clarified yet. The transient receptor potential ankyrin 1 (TRPA1) is a nonselective cation channel mainly expressed in primary sensory neurons, where it functions as a pain sensor. TRPA1 have also been described in multiple non-excitable cells, including those derived from neural crest stem cells such as glial cells and, in particular, Schwann cell oligodendrocytes and astrocytes. We evaluated TRPA1 expression in SS. We selected a cohort of 41 SSs, and by immunohistochemistry, we studied TRPA1 expression TRPA1 was found in 92.6% of cases. Triple TRPA1/pS100/SOX10 and TRPA1/SLUG/SNAIL staining strongly supports a neural origin of SS. TRPA1 positivity was also observed in a subset of cases negative with pS100, SOX10 and/or SLUG/SNAIL, and these divergent phenotypes may reflect a process of tumor plasticity and dedifferentiation of neural-derived SSs. Given the functional diversity of TRPA1 and its expression in neuronal and non-neuronal multipotent neural crest stem cells, it remains to be determined whether TRPA1 expression in SSs neoplastic cells plays a role in the molecular mechanism associated with premonitory pain symptoms and tumor progression.
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http://dx.doi.org/10.3390/biom10101446DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7602570PMC
October 2020

Recognition of Cutaneous Melanoma on Digitized Histopathological Slides Artificial Intelligence Algorithm.

Front Oncol 2020 20;10:1559. Epub 2020 Aug 20.

Institute of Clinical Physiology, National Research Council, Pisa, Italy.

Increasing incidence of skin cancer combined with a shortage of dermatopathologists has increased the workload of pathology departments worldwide. In addition, the high intraobserver and interobserver variability in the assessment of melanocytic skin lesions can result in underestimated or overestimated diagnosis of melanoma. Thus, the development of new techniques for skin tumor diagnosis is essential to assist pathologists to standardize diagnoses and plan accurate patient treatment. Here, we describe the development of an artificial intelligence (AI) system that recognizes cutaneous melanoma from histopathological digitalized slides with clinically acceptable accuracy. Whole-slide digital images from 100 formalin-fixed paraffin-embedded primary cutaneous melanoma were used to train a convolutional neural network (CNN) based on a pretrained Inception-ResNet-v2 to accurately and automatically differentiate tumoral areas from healthy tissue. The CNN was trained by using 60 digital slides in which regions of interest (ROIs) of tumoral and healthy tissue were extracted by experienced dermatopathologists, while the other 40 slides were used as test datasets. A total of 1377 patches of healthy tissue and 2141 patches of melanoma were assessed in the training/validation set, while 791 patches of healthy tissue and 1122 patches of pathological tissue were evaluated in the test dataset. Considering the classification by expert dermatopathologists as reference, the trained deep net showed high accuracy (96.5%), sensitivity (95.7%), specificity (97.7%), F score (96.5%), and a Cohen's kappa of 0.929. Our data show that a deep learning system can be trained to recognize melanoma samples, achieving accuracies comparable to experienced dermatopathologists. Such an approach can offer a valuable aid in improving diagnostic efficiency when expert consultation is not available, as well as reducing interobserver variability. Further studies in larger data sets are necessary to verify whether the deep learning algorithm allows subclassification of different melanoma subtypes.
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http://dx.doi.org/10.3389/fonc.2020.01559DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7508308PMC
August 2020

Neuroprotective effects of mGluR5 activation through the PI3K/Akt pathway and the molecular switch of AMPA receptors.

Neuropharmacology 2020 01 7;162:107810. Epub 2019 Oct 7.

Department of Health Sciences, Unit of Clinical Pharmacology and Oncology, University of Florence, Viale G. Pieraccini 6, 50139, Florence, Italy.

Previous studies have demonstrated that antagonists of mGluR1, but not mGluR5, are neuroprotective in models of cerebral ischemia. To investigate the individual roles of mGlu1 and mGlu5 receptors in in vitro model of cerebral ischemia we used low doses of the non-selective group I agonist DHPG and mGlu1 and mGlu5 selective positive allosteric modulators (PAMs). In hippocampal slices subjected to 30 min oxygen-glucose deprivation (OGD), DHPG (1 μM) and the mGluR5 PAM (VU0092273) significantly reduced OGD-induced CA1 injury monitored by propidium iodide staining of the slices and quantitative analysis of CA1 neurons. In contrast, the mGluR1 PAM (VU0483605) showed no neuroprotection. These protective effects of DHPG and VU0092273 were prevented by inhibition of PI3K/Akt pathway by LY294002. The mGluR5 PAM (VU0092273) also prevented GluA2 down-regulation triggered by ischemic injury, via PI3K/Akt pathway, revealing a further contribution to its neuroprotective effects by reducing the excitotoxic effects of increased Ca influx through GluA2-lacking AMPA receptors. Furthermore, immunohistochemical assays confirmed the neuroprotective effect of VU0092273 and revealed activation of glia, indicating the involvement reactive astrogliosis in the mechanisms of neuroprotection. Our data suggest that selective activation/potentiation of mGluR5 signalling represents a promising strategy for the development of new interventions to reduce or prevent ischemia-induced neuronal death.
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http://dx.doi.org/10.1016/j.neuropharm.2019.107810DOI Listing
January 2020

Microglial distribution, branching, and clearance activity in aged rat hippocampus are affected by astrocyte meshwork integrity: evidence of a novel cell-cell interglial interaction.

FASEB J 2019 03 29;33(3):4007-4020. Epub 2018 Nov 29.

Department of Experimental and Clinical Medicine, University of Florence, Florence, Italy.

Aging and neurodegenerative diseases share a condition of neuroinflammation entailing the production of endogenous cell debris in the CNS that must be removed by microglia ( i.e., resident macrophages), to restore tissue homeostasis. In this context, extension of microglial cell branches toward cell debris underlies the mechanisms of microglial migration and phagocytosis. Amoeboid morphology and the consequent loss of microglial branch functionality characterizes dysregulated microglia. Microglial migration is assisted by another glial population, the astroglia, which forms a dense meshwork of cytoplasmic projections. Amoeboid microglia and disrupted astrocyte meshwork are consistent traits in aged CNS. In this study, we assessed a possible correlation between microglia and astroglia morphology in rat models of chronic neuroinflammation and aging, by 3-dimensional confocal analysis implemented with particle analysis. Our findings suggest that a microglia-astroglia interaction occurs in rat hippocampus via cell-cell contacts, mediating microglial cell branching in the presence of inflammation. In aged rats, the impairment of such an interaction correlates with altered distribution, morphology, and inefficient clearance by microglia. These data support the idea that generally accepted functional boundaries between microglia and astrocytes should be re-evaluated to better understand how their functions overlap and interact.-Lana, D., Ugolini, F., Wenk, G. L., Giovannini, M. G., Zecchi-Orlandini, S., Nosi, D. Microglial distribution, branching, and clearance activity in aged rat hippocampus are affected by astrocyte meshwork integrity: evidence of a novel cell-cell interglial interaction.
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http://dx.doi.org/10.1096/fj.201801539RDOI Listing
March 2019

Different Patterns of Neurodegeneration and Glia Activation in CA1 and CA3 Hippocampal Regions of TgCRND8 Mice.

Front Aging Neurosci 2018 13;10:372. Epub 2018 Nov 13.

Department of Health Sciences, Section of Clinical Pharmacology and Oncology, University of Florence, Florence, Italy.

We investigated the different patterns of neurodegeneration and glia activation in CA1 and CA3 hippocampal areas of TgCRND8 mice. The main feature of this transgenic model is the rapid development of the amyloid pathology, which starts already at 3 months of age. We performed immunohistochemical analyses to compare the different sensibility of the two hippocampal regions to neurodegeneration. We performed qualitative and quantitative evaluations by fluorescence immunohistochemistry with double or triple staining, followed by confocal microscopy and digital image analysis in stratum pyramidale (SP) and stratum radiatum (SR) of CA1 and CA3, separately. We evaluated time-dependent Aβ plaques deposition, expression of inflammatory markers, as well as quantitative and morphological alterations of neurons and glia in transgenic mice at 3 (Tg 3M) and 6 (Tg 6M) months of age, compared to WT mice. In CA1 SR of Tg 6M mice, we found significantly more Medium and Large plaques than in CA3. The pattern of neurodegeneration and astrocytes activation was different in the two areas, indicating higher sensitivity of CA1. In the CA1 SP of Tg 6M mice, we found signs of reactive astrogliosis, such as increase of astrocytes density in SP, increase of GFAP expression in SR, and elongation of astrocytes branches. We found also common patterns of glia activation and neurodegenerative processes in CA1 and CA3 of Tg 6M mice: significant increase of total and reactive microglia density in SP and SR, increased expression of TNFα, of iNOS, and IL1β in astrocytes and increased density of neurons-astrocytes-microglia triads. In CA1 SP, we found decrease of volume and number of pyramidal neurons, paralleled by increase of apoptosis, and, consequently, shrinkage of CA1 SP. These data demonstrate that in TgCRND8 mice, the responses of neurons and glia to neurodegenerative patterns induced by Aβ plaques deposition is not uniform in the two hippocampal areas, and in CA1 pyramidal neurons, the higher sensitivity may be related to the different plaque distribution in this area. All these modifications may be at the basis of memory loss, the peculiar symptom of AD, which was demonstrated in this transgenic mouse model of Aβ deposition, even at early stages.
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http://dx.doi.org/10.3389/fnagi.2018.00372DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6243135PMC
November 2018

The Selective Antagonism of Adenosine A Receptors Reduces the Synaptic Failure and Neuronal Death Induced by Oxygen and Glucose Deprivation in Rat CA1 Hippocampus .

Front Pharmacol 2018 24;9:399. Epub 2018 Apr 24.

Department of Neuroscience, Psychology, Drug Research and Child Health, NEUROFARBA, Section of Pharmacology and Toxicology, University of Florence, Florence, Italy.

Ischemia is a multifactorial pathology characterized by different events evolving in time. Immediately after the ischemic insult, primary brain damage is due to the massive increase of extracellular glutamate. Adenosine in the brain increases dramatically during ischemia in concentrations able to stimulate all its receptors, A, A, A, and A. Although adenosine exerts clear neuroprotective effects through A receptors during ischemia, the use of selective A receptor agonists is hampered by their undesirable peripheral side effects. So far, no evidence is available on the involvement of adenosine A receptors in cerebral ischemia. This study explored the role of adenosine A receptors on synaptic and cellular responses during oxygen and glucose deprivation (OGD) in the CA1 region of rat hippocampus . We conducted extracellular recordings of CA1 field excitatory post-synaptic potentials (fEPSPs); the extent of damage on neurons and glia was assessed by immunohistochemistry. Seven min OGD induced anoxic depolarization (AD) in all hippocampal slices tested and completely abolished fEPSPs that did not recover after return to normoxic condition. Seven minutes OGD was applied in the presence of the selective adenosine A receptor antagonists MRS1754 (500 nM) or PSB603 (50 nM), separately administered 15 min before, during and 5 min after OGD. Both antagonists were able to prevent or delay the appearance of AD and to modify synaptic responses after OGD, allowing significant recovery of neurotransmission. Adenosine A receptor antagonism also counteracted the reduction of neuronal density in CA1 stratum pyramidale, decreased apoptosis at least up to 3 h after the end of OGD, and maintained activated mTOR levels similar to those of controls, thus sparing neurons from the degenerative effects caused by the simil-ischemic conditions. Astrocytes significantly proliferated in CA1 stratum radiatum already 3 h after the end of OGD, possibly due to increased glutamate release. Areceptor antagonism significantly prevented astrocyte modifications. Both A receptor antagonists did not protect CA1 neurons from the neurodegeneration induced by glutamate application, indicating that the antagonistic effect is upstream of glutamate release. The selective antagonists of the adenosine A receptor subtype may thus represent a new class of neuroprotective drugs in ischemia.
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http://dx.doi.org/10.3389/fphar.2018.00399DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5928446PMC
April 2018

Alterations in the Interplay between Neurons, Astrocytes and Microglia in the Rat Dentate Gyrus in Experimental Models of Neurodegeneration.

Front Aging Neurosci 2017 11;9:296. Epub 2017 Sep 11.

Department of Health Sciences, Section of Pharmacology and Clinical Oncology, University of FlorenceFlorence, Italy.

The hippocampus is negatively affected by aging and neurodegenerative diseases leading to impaired learning and memory abilities. A diverse series of progressive modifications in the intercellular communication among neurons, astrocytes and microglia occur in the hippocampus during aging or inflammation. A detailed understanding of the neurobiological modifications that contribute to hippocampal dysfunction may reveal new targets for therapeutic intervention. The current study focussed on the interplay between neurons and astroglia in the Granule Layer (GL) and the Polymorphic Layer (PL) of the Dentate Gyrus (DG) of adult, aged and LPS-treated rats. In GL and PL of aged and LPS-treated rats, astrocytes were less numerous than in adult rats. In GL of LPS-treated rats, astrocytes acquired morphological features of reactive astrocytes, such as longer branches than was observed in adult rats. Total and activated microglia increased in the aged and LPS-treated rats, as compared to adult rats. In the GL of aged and LPS-treated rats many neurons were apoptotic. Neurons decreased significantly in GL and PL of aged but not in rats treated with LPS. In PL of aged and LPS-treated rats many damaged neurons were embraced by microglia cells and were infiltrated by branches of astrocyte, which appeared to be bisecting the cell body, forming triads. Reactive microglia had a scavenging activity of dying neurons, as shown by the presence of neuronal debris within their cytoplasm. The levels of the chemokine fractalkine (CX3CL1) increased in hippocampal homogenates of aged rats and rats treated with LPS, and CX3CL1 immunoreactivity colocalized with activated microglia cells. Here we demonstrated that in the DG of aged and LPS-treated rats, astrocytes and microglia cooperate and participate in phagocytosis/phagoptosis of apoptotic granular neurons. The differential expression/activation of astroglia and the alteration of their intercommunication may be responsible for the different susceptibility of the DG in comparison to the CA1 and CA3 hippocampal areas to neurodegeneration during aging and inflammation.
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http://dx.doi.org/10.3389/fnagi.2017.00296DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5601988PMC
September 2017

The anti-migraine component of butterbur extracts, isopetasin, desensitizes peptidergic nociceptors by acting on TRPA1 cation channel.

Br J Pharmacol 2017 Sep 20;174(17):2897-2911. Epub 2017 Jul 20.

Department of Health Sciences, Section of Clinical Pharmacology and Headache Center, University of Florence, Florence, Italy.

Background And Purpose: The mechanism of the anti-migraine action of extracts of butterbur [Petasites hybridus (L.) Gaertn.] is unknown. Here, we investigated the ability of isopetasin, a major constituent of these extracts, to specifically target TRPA1 channel and to affect functional responses relevant to migraine.

Experimental Approach: Single-cell calcium imaging and patch-clamp recordings in human and rodent TRPA1-expressing cells, neurogenic motor responses in rodent isolated urinary bladder, release of CGRP from mouse spinal cord in vitro and facial rubbing in mice and meningeal blood flow in rats were examined.

Key Results: Isopetasin induced (i) calcium responses and currents in rat/mouse trigeminal ganglion (TG) neurons and in cells expressing the human TRPA1, (ii) substance P-mediated contractions of rat isolated urinary bladders and (iii) CGRP release from mouse dorsal spinal cord, responses that were selectively abolished by genetic deletion or pharmacological antagonism of TRPA1 channels. Pre-exposure to isopetasin produced marked desensitization of allyl isothiocyanate (AITC, TRPA1 channel agonist)- or capsaicin (TRPV1 channel agonist)-evoked currents in rat TG neurons, contractions of rat or mouse bladder and CGRP release from mouse central terminals of primary sensory neurons. Repeated intragastric administration of isopetasin attenuated mouse facial rubbing, evoked by local AITC or capsaicin, and dilation of rat meningeal arteries by acrolein or ethanol (TRPA1 and TRPV1 channel agonists respectively).

Conclusion And Implications: Activation of TRPA1 channels by isopetasin results in excitation of neuropeptide-containing nociceptors, followed by marked heterologous neuronal desensitization. Such atten uation in pain and neurogenic inflammation may account for the anti-migraine action of butterbur.
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http://dx.doi.org/10.1111/bph.13917DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5554318PMC
September 2017

The neuron-astrocyte-microglia triad in CA3 after chronic cerebral hypoperfusion in the rat: Protective effect of dipyridamole.

Exp Gerontol 2017 10 9;96:46-62. Epub 2017 Jun 9.

Department of Health Sciences, Section of Pharmacology and Clinical Oncology, University of Florence, Viale Pieraccini 6, 50139 Firenze, Italy. Electronic address:

We investigated the quantitative and morphofunctional alterations of neuron-astrocyte-microglia triads in CA3 hippocampus, in comparison to CA1, after 2 Vessel Occlusion (2VO) and the protective effect of dipyridamole. We evaluated 3 experimental groups: sham-operated rats (sham, n=15), 2VO-operated rats treated with vehicle (2VO-vehicle, n=15), and 2VO-operated rats treated with dipyridamole from day 0 to day 7 (2VO-dipyridamole, n=15), 90days after 2VO. We analyzed Stratum Pyramidalis (SP), Stratum Lucidum (SL) and Stratum Radiatum (SR) of CA3. 1) ectopic neurons increased in SL and SR of 2VO-vehicle, and 2VO-dipyridamole rats; 2) apoptotic neurons increased in SP of 2VO-vehicle rats and dipyridamole reverted this effect; 3) astrocytes increased in SP, SL and SR of 2VO-vehicle and 2VO-dipyridamole rats; 4) TNF-α expression increased in astrocytes, blocked by dipyridamole, and in dendrites in SR of 2VO-vehicle rats; 5) total microglia increased in SL and SR of 2VO-vehicle and 2VO-dipyridamole rats; 6) triads increased in SR of 2VO-vehicle rats and dipyridamole reverted this effect. Microglia cooperated with astrocytes to phagocytosis of apoptotic neurons and debris, and engulfed ectopic non-fragmented neurons in SL of 2VO-vehicle and 2VO-dipyridamole rats, through a new mechanism called phagoptosis. CA3 showed a better adaptive capacity than CA1 to the ischemic insult, possibly due to the different behaviour of astrocytes and microglial cells. Dipyridamole had neuroprotective effects.
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http://dx.doi.org/10.1016/j.exger.2017.06.006DOI Listing
October 2017