Characterization of human CD4+EOMES+GzmK+ T-cell subsets unveils an uncoupling of suppressive functions from IL-10-producing capacities.

Human CD4+EOMES+ T cells are heterogeneous and contain Th1-cells, Tr1-cells, and CD4+CTL. Tr1- cells and non-classical EOMES+ Th1-cells displayed, respectively, anti- and pro-inflammatory cytokine profiles, but both expressed granzyme-K, produced IFN-γ, and suppressed T-cell proliferation. Diffusion map suggested a progressive CD4+T-cell differentiation from naïve to cytotoxic cells and identified EOMES+Th1-cells as putative Tr1-cell precursors (pre-Tr1).

Eomesodermin-expressing type 1 regulatory (EOMES+ Tr1)-like T cells: Basic biology and role in immune-mediated diseases.

Type 1 regulatory (Tr1) T cells are currently defined all T cells with regulatory functions that lack FOXP3 expression and produce IL-10. Tr1 cells are heterogeneous, and the different reported properties of Tr1-cell populations have caused some confusion in the field. Moreover, understanding the role of Tr1 cells in immune-mediated diseases has been hampered by the lack of a lineage-defining transcription factor. Several independent studies indicated recently that the transcription factor Eomesodermin (EOMES) could act as a lineage-defining transcription factor in a population of IL-10 and IFN-γ co-producing Tr1-like cells, since EOMES directly induces IFN-γ and cytotoxicity, enhances IL-10, and antagonizes alternative T-cell fates. Here, we review the known properties of EOMES+ Tr1-like cells. They share several key characteristics with other Tr1 cells (i.e., "Tr1-like"), namely high IL-10 production, cytotoxicity, and suppressive capabilities. Notably, they also share some features with FOXP3+ Tregs, like downregulation of IL-7R and CD40L. In addition, they possess several unique, EOMES-dependent features, that is, expression of GzmK and IFN-γ, and downregulation of type-17 cytokines. Published evidence indicates that EOMES+ Tr1-like cells play key roles in graft-versus-host disease, colitis, systemic autoimmunity and in tumors. Thus, EOMES+ Tr1-like cells are key players of the adaptive immune system that are involved in several different immune-mediated diseases.

SARS-COV-2 specific t-cells in patients with thyroid disorders related to COVID-19 are enriched in the thyroid and acquire a tissue-resident memory phenotype.

BACKGROUND: SARS-CoV-2 infections have been associated with the onset of thyroid disorders like classic subacute thyroiditis (SAT) or atypical SAT upon severe COVID disease (COV-A-SAT). Little is known about thyroid anti-viral immune responses. OBJECTIVES: To define the role of T-cells in COV-A-SAT. METHODS: T-cells from COV-A-SAT patients were analyzed by multi-dimensional flow cytometry, UMAP and DiffusionMap dimensionality reduction and FlowSOM clustering. T-cells from COVID-naïve healthy donors, patients with autoimmune thyroiditis (ATD) and with SAT following COVID vaccination were analyzed as controls. T-cells were analyzed four and eight months post-infection in peripheral blood and in thyroid specimen obtained by ultrasound-guided fine needle aspiration. SARS-COV2-specific T-cells were identified by cytokine production induced by SARS-COV2-derived peptides and with COVID peptide-loaded HLA multimers after HLA haplotyping. RESULTS: COV-A-SAT was associated with HLA-DRB1*13 and HLA-B*57. COV-A-SAT patients contained activated Th1- and cytotoxic CD4+ and CD8+ effector cells four months post-infection, which acquired a quiescent memory phenotype after eight months. Anti-SARS-CoV-2-specific T-cell responses were readily detectable in peripheral blood four months post-infection, but were reduced after eight months. CD4+ and CD8+ tissue-resident memory cells (TRM) were present in the thyroid, and circulating CXCR3+T-cells identified as their putative precursors. SARS-CoV-2-specific T-cells were enriched in the thyroid, and acquired a TRM phenotype eight months post-infection. CONCLUSIONS: The association of COV-A-SAT with specific HLA haplotypes suggests a genetic predisposition and a key role for T-cells. COV-A-SAT is characterized by a prolonged systemic anti-viral effector T-cell response and the late generation of COVID-specific TRM in the thyroid target tissue.

Ex vivo microRNA and gene expression profiling of human Tr1-like cells suggests a role for miR-92a and -125a in the regulation of EOMES and IL-10R.

Ex vivo gene expression and miRNA profiling of Eomes+ Tr1-like cells suggested that they represent a differentiation stage that is intermediate between Th1-cells and cytotoxic CD4+ T-cells. Several microRNAs were downregulated in Eomes+ Tr1-like cells that might inhibit Tr1-cell differentiation. In particular, miR-92a targeted Eomes, while miR-125a inhibited IFN-g and IL-10R expression.

Secretome of human adipose-derived mesenchymal stem cell relieves pain and neuroinflammation independently of the route of administration in experimental osteoarthritis.

OBJECTIVE: Treatment of pain associated with osteoarthritis (OA) is unsatisfactory and innovative approaches are needed. The secretome from human adipose-derived mesenchymal stem cells (hASC-Conditioned Medium, CM) has been successfully used to relieve painful symptoms in models of chronic pain. The aim of this study was to explore the efficacy of the hASC-CM to control pain and neuroinflammation in an animal model of OA. METHODS: OA was induced in mice by intra-articular monosodium-iodoacetate (MIA) injection. Thermal hyperalgesia and mechanical allodynia were assessed. Once hypersensitivity was established (7 days after MIA), hASC-CM was injected by IA, IPL and IV route and its effect monitored over time. Neuroinflammation in nerve, dorsal root ganglia and spinal cord was evaluated measuring proinflammatory markers and mediators by RT-qPCR. Protein content analysis of secretome by Mass Spectrometry was performed. RESULTS: A single injection with hASC-CM induced a fast and long lasting antihyperalgesic and antiallodynic effect. The IV route of administration appeared to be the most efficacious although all the treatments were effective. The effect on pain correlated with the ability of hASC-CM to reduce the neuroinflammatory condition in both the peripheral and central nervous system. Furthermore, the secretome analysis revealed 101 factors associated with immune regulation. CONCLUSION: We suggest that hASC-CM is a valid treatment option for controlling OA-related hypersensitivity, exerting a rapid and long lasting pain relief. The mechanisms underpinning its effects are likely linked to the positive modulation of neuroinflammation in peripheral and central nervous system that sustains peripheral and central sensitization.

Targeting prokineticin system counteracts hypersensitivity, neuroinflammation, and tissue damage in a mouse model of bortezomib-induced peripheral neuropathy.

BACKGROUND: Neuropathy is a dose-limiting side effect of many chemotherapeutics, including bortezomib. The mechanisms underlying this condition are not fully elucidated even if a contribution of neuroinflammation was suggested. Here, we investigated the role of a chemokine family, the prokineticins (PKs), in the development of bortezomib-induced peripheral neuropathy (BIPN), and we used a PK receptor antagonist to counteract the development and progression of the pathology. METHODS: Neuropathy was induced in male C57BL/6J mice by using a protocol capable to induce a detectable neuropathic phenotype limiting systemic side effects. The presence of allodynia (both mechanical and thermal) and thermal hyperalgesia was monitored over time. Mice were sacrificed at two different time points: 14 and 28 days after the first bortezomib (BTZ) injection. At these times, PK system activation (PK2 and PK-Rs), macrophage and glial activation markers, and cytokine production were evaluated in the main station involved in pain transmission (sciatic nerve, DRG, and spinal cord), and the effect of a PK receptors antagonist (PC1) on the same behavioral and biochemical parameters was assessed. Structural damage of DRG during BTZ treatment and an eventual protective effect of PC1 were also evaluated. RESULTS: BTZ induces in mice a dose-related allodynia and hyperalgesia and a progressive structural damage to the DRG. We observed a precocious increase of macrophage activation markers and unbalance of pro- and anti-inflammatory cytokines in sciatic nerve and DRG together with an upregulation of GFAP in the spinal cord. At higher BTZ cumulative dose PK2 and PK receptors are upregulated in the PNS and in the spinal cord. The therapeutic treatment with the PK-R antagonist PC1 counteracts the development of allodynia and hyperalgesia, ameliorates the structural damage in the PNS, decreases the levels of activated macrophage markers, and prevents full neuroimmune activation in the spinal cord. CONCLUSIONS: PK system may be a strategical pharmacological target to counteract BTZ-induced peripheral neuropathy. Blocking PK2 activity reduces progressive BTZ toxicity in the DRG, reducing neuroinflammation and structural damage to DRG, and it may prevent spinal cord sensitization.

Prokineticin 2 promotes and sustains neuroinflammation in vincristine treated mice: Focus on pain and emotional like behavior.

Vincristine (VCR) treatment is often associated to painful neuropathy. Its development is independent from antitumoral mechanism and involves neuroinflammation. We investigated the role of the chemokine prokineticin (PK)2 in a mouse model of VCR induced neuropathy using a PK-receptors (PK-R) antagonist to counteract its development. We also evaluated emotional like deficits in VCR mice. VCR (0,1 mg/kg) was i.p. injected in C57BL/6J male mice once a day for 14 consecutive days. Pain, anxiety and depressive like behaviors were assessed in animals. PK2, PK-Rs, cytokines, neuroinflammatory markers (CD68, CD11b, GFAP, TLR4) and ATF3 were evaluated in DRG, spinal cord, prefrontal cortex and hippocampus. The PK-Rs antagonist PC1, was s.c. injected (150 µg/kg) twice a day from day 7 (hypersensitivity state) until day 14. Its effect on pain and neuroinflammation was evaluated. VCR mice developed neuropathic pain but not mood alterations. After 7 days of VCR treatment we observed a neuroinflammatory condition in DRG with high levels of PK-Rs, TLR4, CD68, ATF3 and IL-1ß without relevant alterations in spinal cord. At day 14, an upregulation of PK system and a marked neuroinflammation was evident also in spinal cord. Moreover, at the same time, we observed initial alterations in supraspinal brain areas. PC1 treatment significantly counteracted neuropathic pain and blunted neuroinflammation.

Effect of Tapentadol on Splenic Cytokine Production in Mice.

BACKGROUND: Opioid drugs affect immunity, but not all opioid drugs share the same immunomodulatory properties. Tapentadol is an analgesic drug with a dual synergistic mechanism of action: µ-opioid receptor agonism and noradrenaline reuptake inhibition. Weaker µ-opioid receptor agonism combined with noradrenaline reuptake inhibition results in potent analgesia with reduced opioid side effects. We evaluated the impact of tapentadol on splenic cytokine in normal and in hyperalgesia/allodynia mice, comparing it with morphine and reboxetine, a noradrenaline reuptake inhibitor. METHODS: Tapentadol, reboxetine, and morphine were injected subcutaneously into naïve and mice that underwent sciatic nerve chronic constriction injury, and their effect on splenic cytokines (interferon-γ [IFN-γ], interleukin [IL]-2, IL-10, and IL-4) was measured by enzyme-linked immunosorbent assay after acute or chronic treatment. Nociceptive thresholds, thermal hyperalgesia, and allodynia also were assessed. Data were analyzed with 2-way analysis of variance (behavior) or 1-way analysis of variance (cytokines) followed by Bonferroni post hoc test. RESULTS: Primary outcomes of our study were the effects of drugs on splenic cytokines. Our data indicate that acute tapentadol did not modify cytokine production in comparison with animals that received saline, whereas morphine suppressed all the cytokines: saline versus morphine 10 mg/kg (mean difference [MD], 95% confidence interval [CI]: IFN-γ = 12,400 [7760, 17,040], P < .001; IL-2 = 216.2 [47.69, 384.7], P < .01; IL-10 = 868 [523.7, 1212], P < .001; and IL-4 = 17.26 [10.32, 24.20], P < .001). A significant difference also was present between morphine and tapentadol (morphine 10 mg/kg versus tapentadol 20 mg/kg: MD [95% CI]: IFN-γ = -11,600 [-16,240, -6960], P < .001; IL-2 = -334.2 [-502.7, -165.7], P < .001; IL-10 = -959 [-1303, -614.7], P < .001; IL-4 = -18.66 [-25.60, -11.72], P < .001). When chronically injected for 7 days, tapentadol and reboxetine did not significantly affect cytokines when compared with saline-treated animals. The immunoprofile of tapentadol was different from that of morphine also in mice that were in a condition of neuropathic pain. All cytokines appeared significantly decreased in mice that received a chronic constriction injury in comparison with sham animals but, after 7 days of treatment, with a similar antihyperalgesic profile, IL-10 and IL-4 were significantly increased in tapentadol and reboxetine animals in comparison with morphine mice (morphine versus tapentadol: MD [95% CI], IL-10 = -926.4 [-1664, -188.5], P < .01; IL-4 = -8.15 [-12.46, -3.84], P < .001). CONCLUSIONS: Acute and chronic tapentadol seem to be protective of splenic cytokines in contrast with morphine, which exerts a generalized suppression on all cytokines.

Effects of NSAIDs on the Release of Calcitonin Gene-Related Peptide and Prostaglandin E2 from Rat Trigeminal Ganglia.

Nonsteroidal anti-inflammatory drugs (NSAIDs) are frequently used to treat migraine, but the mechanisms of their effects in this pathology are not fully elucidated. The trigeminal ganglia and calcitonin gene-related peptide (CGRP) have been implicated in the pathophysiology of migraine. The release of CGRP and prostaglandin E2 (PGE2) from freshly isolated rat trigeminal ganglia was evaluated after oral administration of nimesulide, etoricoxib, and ketoprofen, NSAIDs with different pharmacological features. Thirty minutes after oral administration, nimesulide, 10 mg/Kg, decreased the GCRP release induced by an inflammatory soup, while the other NSAIDs were ineffective at this point in time. Two hours after oral nimesulide (5 and 10 mg/Kg) and ketoprofen (10 mg/Kg), but not of etoricoxib, a significant decrease in the CGRP release was observed. All drugs reduced PGE2, although with some differences in timing and doses, and the action on CGRP does not seem to be related to PGE2 inhibition. The reduction of CGRP release from rat trigeminal ganglia after nimesulide and ketoprofen may help to explain the mechanism of action of NSAIDs in migraine. Since at 30 minutes only nimesulide was effective in reducing CGRP release, these results suggest that this NSAID may exert a particularly rapid effect in patients with migraine.

Endothelial cellular senescence and tau accumulation: An interplay full of opportunities?

Recent research has shown that tau protein can be passed to neighboring cells, leading to cellular senescence in the endothelial cells present in the central nervous system (CNS). This discovery could potentially open new doors for testing novel therapeutic compounds that specifically target senescent cells (senolytics) or for identifying new biomarkers that can enable early detection of tauopathies and dementia.

Gold and silver nanoparticles in Alzheimer's and Parkinson's diagnostics and treatments.

Neurodegenerative diseases (NDs) impose substantial medical and public health burdens on people worldwide and represent one of the major threats to human health. The prevalence of these age-dependent disorders is dramatically increasing over time, a process intrinsically related to a constantly rising percentage of the elderly population in recent years. Among all the NDs, Alzheimer's and Parkinson's are considered the most debilitating as they cause memory and cognitive loss, as well as severely affecting basic physiological conditions such as the ability to move, speak, and breathe. There is an extreme need for new and more effective therapies to counteract these devastating diseases, as the available treatments are only able to slow down the pathogenic process without really stopping or resolving it. This review aims to elucidate the current nanotechnology-based tools representing a future hope for NDs treatment. Noble metal nano-systems, that is, gold and silver nanoparticles (NPs), have indeed unique physicochemical characteristics enabling them to deliver any pharmacological treatment in a more effective way within the central nervous system. This can potentially make NPs a new hope for reversing the actual therapeutic strategy based on slowing down an irreversible process into a more effective and permanent treatment.

An Intestinal Th17 Subset is Associated with Inflammation in Crohn's Disease and Activated by Adherent-invasive Escherichia coli.

IFNγ-producing ex-Th17 cells ['Th1/17'] were shown to play a key pathogenic role in experimental colitis and are abundant in the intestine. Here, we identified and characterised a novel, potentially colitogenic subset of Th17 cells in the intestine of patients with Crohn's disease [CD]. Human Th17 cells expressing CCR5 ['pTh17'] co-expressed T-bet and RORC/γt and produced very high levels of IL-17, together with IFN-γ. They had a gene signature of Th17 effector cells and were distinct from established Th1/17 cells. pTh17 cells, but not Th1/17 cells, were associated with intestinal inflammation in CD, and decreased upon successful anti-TNF therapy with infliximab. Conventional CCR5[-]Th17 cells differentiated to pTh17 cells with IL-23 in vitro. Moreover, anti-IL-23 therapy with risankizumab strongly reduced pTh17 cells in the intestine. Importantly, intestinal pTh17 cells were selectively activated by adherent-invasive Escherichia coli [AIEC], but not by a commensal/probiotic E. coli strain. AIEC induced high levels of IL-23 and RANTES from dendritic cells [DC]. Intestinal CCR5+Th1/17 cells responded instead to cytomegalovirus and were reduced in ulcerative colitis [UC], suggesting an unexpected protective role. In conclusion, we identified an IL-23-inducible subset of human intestinal Th17 cells. pTh17 cells produced high levels of pro-inflammatory cytokines, were selectively associated with intestinal inflammation in CD, and responded to CD-associated AIEC, suggesting a key colitogenic role.

Deep Phenotyping of T-Cells Derived From the Aneurysm Wall in a Pediatric Case of Subarachnoid Hemorrhage.

Intracranial aneurysms (IAs) are very rare in children, and the characteristics of the T-cells in the IA wall are largely unknown. A comatose 7-years-old child was admitted to our center because of a subarachnoid hemorrhage due to a ruptured giant aneurysm of the right middle cerebral artery. Two days after the aneurysm clipping the patient was fully awake with left hemiparesis. T-cells from the IA wall and from peripheral blood of this patient were analyzed by multi-dimensional flow cytometry. Unbiased analysis, based on the use of FlowSOM clustering and dimensionality reduction technique UMAP, indicated that there was virtually no overlap between circulating and tissue-infiltrating T-cells. Thus, naïve T-cells and canonical memory T-cells were largely restricted to peripheral blood, while CD4-CD8-T-cells were strongly enriched in the IA wall. The unique CD4+, CD8+ and CD4-CD8-T-cell clusters from the IA wall expressed high levels of CCR5, Granzyme B and CD69, displaying thus characteristics of cytotoxic and tissue-resident effector cells. Low Ki67 expression indicated that they were nevertheless in a resting state. Among regulatory T-cell subsets, Eomes+Tr1-like cells were strongly enriched in the IA wall. Finally, analysis of cytokine producing capacities unveiled that the IA wall contained poly-functional T-cells, which expressed predominantly IFN-γ, TNF and IL-2. CD4+T-cells co-expressed also CD40L, and produced some IL-17, GM-CSF and IL-10. This report provides to our knowledge the first detailed characterization of the human T-cell compartment in the IA wall.

Human airway organoids and microplastic fibers: A new exposure model for emerging contaminants.

Three-dimensional (3D) structured organoids are the most advanced in vitro models for studying human health effects, but their application to evaluate the biological effects associated with microplastic exposure was neglected until now. Fibers from synthetic clothes and fabrics are a major source of airborne microplastics, and their release from dryer machines is poorly understood. We quantified and characterized the microplastic fibers (MPFs) released in the exhaust filter of a household dryer and tested their effects on airway organoids (1, 10, and 50 µg mL-1) by optical microscopy, scanning electron microscopy (SEM), confocal microscopy and quantitative reverse transcription-polymerase chain reaction (qRT-PCR). While the presence of MPFs did not inhibit organoid growth, we observed a significant reduction of SCGB1A1 gene expression related to club cell functionality and a polarized cell growth along the fibers. The MPFs did not cause relevant inflammation or oxidative stress but were coated with a cellular layer, resulting in the inclusion of fibers in the organoid. This effect could have long-term implications regarding lung epithelial cells undergoing repair. This exposure study using human airway organoids proved suitability of the model for studying the effects of airborne microplastic contamination on humans and could form the basis for further research regarding the toxicological assessment of emerging contaminants such as micro- or nanoplastics.

Prokineticin Receptor Inhibition With PC1 Protects Mouse Primary Sensory Neurons From Neurotoxic Effects of Chemotherapeutic Drugs in vitro.

Neurotoxicity is a common side effect of chemotherapeutics that often leads to the development of chemotherapy-induced peripheral neuropathy (CIPN). The peptide Prokineticin 2 (PK2) has a key role in experimental models of CIPN and can be considered an insult-inducible endangering mediator. Since primary afferent sensory neurons are highly sensitive to anticancer drugs, giving rise to dysesthesias, the aim of our study was to evaluate the alterations induced by vincristine (VCR) and bortezomib (BTZ) exposure in sensory neuron cultures and the possible preventive effect of blocking PK2 signaling. Both VCR and BTZ induced a concentration-dependent reduction of total neurite length that was prevented by the PK receptor antagonist PC1. Antagonizing the PK system also reduced the upregulation of PK2, PK-R1, TLR4, IL-6, and IL-10 expression induced by chemotherapeutic drugs. In conclusion, inhibition of PK signaling with PC1 prevented the neurotoxic effects of chemotherapeutics, suggesting a promising strategy for neuroprotective therapies against the sensory neuron damage induced by exposure to these drugs.

Do All Opioid Drugs Share the Same Immunomodulatory Properties? A Review From Animal and Human Studies.

Suppression of the immune system has been constantly reported in the last years as a classical side effect of opioid drugs. Most of the studies on the immunological properties of opioids refer to morphine. Although morphine remains the "reference molecule," other semisynthetic and synthetic opioids are frequently used in the clinical practice. The primary objective of this review is to analyze the available literature on the immunomodulating properties of opioid drugs different from morphine in preclinical models and in the human. A search strategy was conducted in PubMed, Embase, and the Cochrane databases using the terms "immunosuppression," "immune system," "opioids," "Natural killer cells," "cytokines," and "lymphocytes." The results achieved concerning the effects of fentanyl, methadone, oxycodone, buprenorphine, remifentanil, tramadol, and tapentadol on immune responses in animal studies, in healthy volunteers and in patients are reported. With some limitations due to the different methods used to measure immune system parameters, the large range of opioid doses and the relatively scarce number of participants in the available studies, we conclude that it is not correct to generalize immunosuppression as a common side effect of all opioid molecules.

Layer- and subregion-specific electrophysiological and morphological changes of the medial prefrontal cortex in a mouse model of neuropathic pain.

Chronic neuropathic pain constitutes a serious public health problem, but the disease mechanisms are only partially understood. The involvement of different brain regions like the medial prefrontal cortex has already been established, but the comparison of the role of different subregions and layers is still inconclusive. In the current study, we performed patch-clamp recordings followed by anatomical reconstruction of pyramidal cells from different layers of the prelimbic and infralimbic subregions of the medial prefrontal cortex in neuropathic (spared nerve injury, SNI) and control mice. We found that in the prelimbic cortex, layer 2/3 pyramidal cells from SNI mice exhibited increased excitability compared to sham controls, whereas prelimbic layer 5 pyramidal neurons showed reduced excitability. Pyramidal cells in both layer 2/3 and layer 5 of the infralimbic subregion did not change their excitability, but layer 2/3 pyramidal cells displayed increased dendritic length and branching. Our findings support the view that chronic pain is associated with subregion- and layer-specific changes in the medial prefrontal cortex. They therefore provide new insights into the mechanisms underlying the chronification of pain.

Immune function after major surgical interventions: the effect of postoperative pain treatment.

INTRODUCTION: Impaired immune function during the perioperative period may be associated with worse short- and long-term outcomes. Morphine is considered a major contributor to immune modulation. PATIENTS AND METHODS: We performed a pilot study to investigate postoperative immune function by analyzing peripheral blood mononuclear cells' functionality and cytokine production in 16 patients undergoing major abdominal surgery. All patients were treated with intravenous (i.v.) patient-controlled analgesia with morphine and continuous wound infusion with ropivacaine+methylprednisolone for 24 hours. After 24 hours, patients were randomized into two groups, one continuing intrawound infusion and the other receiving only i.v. analgesia. We evaluated lymphoproliferation and cytokine production by peripheral blood mononuclear cells at the end of surgery and at 24 and 48 hours postoperatively. RESULTS: A significant reduction in TNF-α, IL-2, IFN-γ and lymphoproliferation was observed immediately after surgery, indicating impaired cell-mediated immunity. TNF-α and IFN-γ remained suppressed up to 48 hours after surgery, while a trend to normalization was observed for IL-2 and lymphoproliferation, irrespective of the treatment group. A significant inverse correlation was present between age and morphine and between age and lymphoproliferation. No negative correlation was present between morphine and cytokine production. We did not find any differences within the two groups between 24 and 48 hours in terms of morphine consumption and immune responses. CONCLUSION: A relevant depression of cell-mediated immunity is associated with major surgery and persists despite optimal analgesia. Even though morphine may participate in immunosuppression, we did not retrieve any dose-related effect.

Therapeutic effect of human adipose-derived stem cells and their secretome in experimental diabetic pain.

Painful neuropathy is one of the complications of diabetes mellitus that adversely affects patients'quality of life. Pharmacological treatments are not fully satisfactory, and novel approaches needed. In a preclinical mouse model of diabetes the effect of both human mesenchymal stromal cells from adipose tissue (hASC) and their conditioned medium (hASC-CM) was evaluated. Diabetes was induced by streptozotocin. After neuropathic hypersensitivity was established, mice were intravenously injected with either 1 × 106 hASC or with CM derived from 2 × 106 hASC. Both hASC and CM (secretome) reversed mechanical, thermal allodynia and thermal hyperalgesia, with a rapid and long lasting effect, maintained up to 12 weeks after treatments. In nerves, dorsal root ganglia and spinal cord of neuropathic mice we determined high IL-1ß, IL-6 and TNF-α and low IL-10 levels. Both treatments restored a correct pro/antinflammatory cytokine balance and prevented skin innervation loss. In spleens of streptozotocin-mice, both hASC and hASC-CM re-established Th1/Th2 balance that was shifted to Th1 during diabetes. Blood glucose levels were unaffected although diabetic animals regained weight, and kidney morphology was recovered by treatments. Our data show that hASC and hASC-CM treatments may be promising approaches for diabetic neuropathic pain, and suggest that cell effect is likely mediated by their secretome.