AT1 receptor autoantibodies mediate effects of metabolic syndrome on dopaminergic vulnerability

The metabolic syndrome has been associated to chronic peripheral inflammation and related with neuroinflammation and neurodegeneration, including Parkinson’s disease. However, the responsible mechanisms are unclear. Previous studies have involved the brain renin-angiotensin system in progression of Parkinson’s disease and the angiotensin receptor type 1 (AT1) has been recently revealed as a major marker of dopaminergic vulnerability in humans. Dysregulation of tissue renin-angiotensin system is a key common mechanism for all major components of metabolic syndrome. Circulating AT1 agonistic autoantibodies have been observed in several inflammation-related peripheral processes, and activation of AT1 receptors of endothelial cells, dopaminergic neurons and glial cells have been observed to disrupt endothelial blood -brain barrier and induce neurodegeneration, respectively. Using a rat model, we observed that metabolic syndrome induces overactivity of nigral pro-inflammatory renin-angiotensin system axis, leading to increase in oxidative stress and neuroinflammation and enhancing dopaminergic neurodegeneration, which was inhibited by treatment with AT1 receptor blockers (ARBs)S

Autoantibodies against ACE2 and angiotensin type-1 receptors increase severity of COVID-19

The renin-angiotensin system (RAS) plays a major role in COVID-19. Severity of several inflammation-related diseases has been associated with autoantibodies against RAS, particularly agonistic autoantibodies for angiotensin type-1 receptors (AA-AT1) and autoantibodies against ACE2 (AA-ACE2). Disease severity of COVID-19 patients was defined as mild, moderate or severe following the WHO Clinical Progression Scale and determined at medical discharge. Serum AA-AT1 and AA-ACE2 were measured in COVID-19 patients (n = 119) and non-infected controls (n = 23) using specific solid-phase, sandwich enzyme-linked immunosorbent assays. Serum LIGHT (TNFSF14; tumor necrosis factor ligand superfamily member 14) levels were measured with the corresponding assay kit. At diagnosis, AA-AT1 and AA-ACE2 levels were significantly higher in the COVID-19 group relative to controls, and we observed significant association between disease outcome and serum AA-AT1 and AA-ACE2 levels. Mild disease patients had significantly lower levels of AA-AT1 (p < 0.01) and AA-ACE2 (p < 0.001) than moderate and severe patients. No significant differences were detected between males and females. The increase in autoantibodies was not related to comorbidities potentially affecting COVID-19 severity. There was significant positive correlation between serum levels of AA-AT1 and LIGHT (TNFSF14; rPearson = 0.70, p < 0.001). Both AA-AT1 (by agonistic stimulation of AT1 receptors) and AA-ACE2 (by reducing conversion of Angiotensin II into Angiotensin 1-7) may lead to increase in AT1 receptor activity, enhance proinflammatory responses and severity of COVID-19 outcome. Patients with high levels of autoantibodies require more cautious control after diagnosis. Additionally, the results encourage further studies on the possible protective treatment with AT1 receptor blockers in COVID-19Axencia Galega de Innovación (IN845D 2020/20). Spanish Ministry of Economy and Competitiveness (RTI2018-098830-B-I00). Spanish Ministry of Health (PI17/00828, RD16/0011/0016 and CIBERNED). Galician Government (XUGA, ED431C 2018/10, ED431G/05). FEDER (Regional European Development Fund)S

The intracellular angiotensin system buffersdeleterious effects of the extracellular paracrine system

The‘classical’renin–angiotensin system (RAS) is a circulating system that controls blood pressure. Local/paracrine RAS,identified in a variety of tissues, including the brain, is involved in different functions and diseases, and RAS blockers arecommonly used in clinical practice. A third type of RAS (intracellular/intracrine RAS) has been observed in some types of cells,including neurons. However, its role is still unknown. The present results indicate that in brain cells the intracellular RAScounteracts the intracellular superoxide/H2O2and oxidative stress induced by the extracellular/paracrine angiotensin II acting onplasma membrane receptors. Activation of nuclear receptors by intracellular or internalized angiotensin triggers a number ofmechanisms that protect the cell, such as an increase in the levels of protective angiotensin type 2 receptors, intracellularangiotensin, PGC-1αand IGF-1/SIRT1. Interestingly, this protective mechanism is altered in isolated nuclei from brains of agedanimals. The present results indicate that at least in the brain, AT1 receptor blockers acting only on the extracellular or paracrineRAS may offer better protection of cellsWe thank Dr. Wei-Dong Le for providing the MES 23.5dopaminergic cell line, Dr. Manuel Serrano and Dr. Ruben Nogueiras for providing thetransgenic mice overexpressing SIRT1, and Dr. Daniel Henrion for providing the AT2KO mice. We thank Pilar Aldrey, Iria Novoa and Cristina Gianzo for their technicalassistance. This study was funded by the Spanish Ministry of Economy andCompetitiveness (BFU2015-70523), Spanish Ministry of Health (RD12/0019/0020,RD16/0011/0016 and CIBERNED), Galician Government (XUGA, GRC2014/002ED431G/05 and CIMUS accreditation 2016‐2019) and FEDER (Regional EuropeanDevelopment Fund)S

Microglial angiotensin type 2 receptors mediate sex-specific expression of inflammatory cytokines independently of circulating estrogen

Consellería de Cultura, Educación e Ordenación Universitaria, Xunta de Galicia, Grant/Award Numbers: XUGA, ED431C 2018/10, ED431G/05; Instituto de Salud Carlos III, Grant/Award Numbers: PI20/00385, RD16/0011/0016, CIBERNED; Secretaría de Estado de Investigación, Desarrollo e Innovación, Grant/Award Number: RTI2018-098830-B-I00; Regional European Development Fund (FEDER)There are sex differences in microglia, which can maintain sex-related gene expression and functional differences in the absence of circulating sex steroids. The angiotensin type 2 (AT2) receptors mediate anti-inflammatory actions in different tissues, including brain. In mice, we performed RT-PCR analysis of microglia isolated from adult brains and RNA scope in situ hybridization from males, females, ovariectomized females, orchiectomized males and brain masculinized females. We also compared wild type and AT2 knockout mice. The expression of AT2 receptors in microglial cells showed sex differences with much higher AT2 mRNA expression in females than in males, and this was not dependent on circulating gonadal hormones, as observed using ovariectomized females, brain masculinized females and orchiectomized males. These results suggest genomic reasons, possibly related to sex chromosome complement, for sex differences in AT2 expression in microglia, as the AT2 receptor gene is located in the X chromosome. Furthermore, sex differences in expression of AT2 receptors were associated to sex differences in microglial expression of key anti-inflammatory cytokines such as interleukin-10 and pro-inflammatory cytokines such as interleukin-1β and interleukin-6. In conclusion, sex differences in microglial AT2 receptor expression appear as a major factor contributing to sex differences in the neuroinflammatory responses beyond the effects of circulating steroidsThere are sex differences in microglia, which can maintain sex-related gene expression and functional differences in the absence of circulating sex steroids. The angiotensin type 2 (AT2) receptors mediate anti-inflammatory actions in different tissues, including brain. In mice, we performed RT-PCR analysis of microglia isolated from adult brains and RNA scope in situ hybridization from males, females, ovariectomized females, orchiectomized males and brain masculinized females. We also compared wild type and AT2 knockout mice. The expression of AT2 receptors in microglial cells showed sex differences with much higher AT2 mRNA expression in females than in males, and this was not dependent on circulating gonadal hormones, as observed using ovariectomized females, brain masculinized females and orchiectomized males. These results suggest genomic reasons, possibly related to sex chromosome complement, for sex differences in AT2 expression in microglia, as the AT2 receptor gene is located in the X chromosome. Furthermore, sex differences in expression of AT2 receptors were associated to sex differences in microglial expression of key anti-inflammatory cytokines such as interleukin-10 and pro-inflammatory cytokines such as interleukin-1β and interleukin-6. In conclusion, sex differences in microglial AT2 receptor expression appear as a major factor contributing to sex differences in the neuroinflammatory responses beyond the effects of circulating steroidsS

Renin angiotensin system and gender differences in dopaminergic degeneration

Background: There are sex differences in dopaminergic degeneration. Men are approximately two times as likely as premenopausal women of the same age to develop Parkinson’s disease (PD). It has been shown that the local renin angiotensin system (RAS) plays a prominent role in sex differences in the development of chronic renal and cardiovascular diseases, and there is a local RAS in the substantia nigra and dopaminergic cell loss is enhanced by angiotensin via type 1 (AT1) receptors. Results: In the present study, we observed that intrastriatal injection of 6-hydroxydopamine induced a marked loss of dopaminergic neurons in the substantia nigra of male rats, which was significantly higher than the loss induced in ovariectomized female rats given estrogen implants (i.e. rats with estrogen). However, the loss of dopaminergic neurons was significantly lower in male rats treated with the AT1 antagonist candesartan, and similar to that observed in female rats with estrogen. The involvement of the RAS in gender differences in dopaminergic degeneration was confirmed with AT1a-null mice lesioned with the dopaminergic neurotoxin MPTP. Significantly higher expression of AT1 receptors, angiotensin converting enzyme activity, and NADPH-oxidase complex activity, and much lower levels of AT2 receptors were observed in male rats than in female rats with estrogen. Conclusions: The results suggest that brain RAS plays a major role in the increased risk of developing PD in men, and that manipulation of brain RAS may be an efficient approach for neuroprotective treatment of PD in men, without the feminizing effects of estrogen.Funding: Spanish Ministry of Science and Innovation, Spanish Ministry of Health (RD06/0010/0013 and CIBERNED), Galician Government (XUGA) and FEDERS

Spanish Cell Therapy Network (TerCel): 15 years of successful collaborative translational research

In the current article we summarize the 15-year experience of the Spanish Cell Therapy Network (TerCel), a successful collaborative public initiative funded by the Spanish government for the support of nationwide translational research in this important area. Thirty-two research groups organized in three programs devoted to cardiovascular, neurodegenerative and immune-inflammatory diseases, respectively, currently form the network. Each program has three working packages focused on basic science, pre-clinical studies and clinical application. TerCel has contributed during this period to boost the translational research in cell therapy in Spain, setting up a network of Good Manufacturing Practice–certified cell manufacturing facilities– and increasing the number of translational research projects, publications, patents and clinical trials of the participating groups, especially those in collaboration. TerCel pays particular attention to the public-private collaboration, which, for instance, has led to the development of the first allogeneic cell therapy product approved by the European Medicines Agency, Darvadstrocel. The current collaborative work is focused on the development of multicenter phase 2 and 3 trials that could translate these therapies to clinical practice for the benefit of patients

Interaction between angiotensin type 1, type 2, and mas receptors to regulate adult neurogenesis in the brain ventricular–subventricular zone

The renin–angiotensin system (RAS), and particularly its angiotensin type-2 receptors (AT2), have been classically involved in processes of cell proliferation and maturation during development. However, the potential role of RAS in adult neurogenesis in the ventricular-subventricular zone (V-SVZ) and its aging-related alterations have not been investigated. In the present study, we analyzed the role of major RAS receptors on neurogenesis in the V-SVZ of adult mice and rats. In mice, we showed that the increase in proliferation of cells in this neurogenic niche was induced by activation of AT2 receptors but depended partially on the AT2-dependent antagonism of AT1 receptor expression, which restricted proliferation. Furthermore, we observed a functional dependence of AT2 receptor actions on Mas receptors. In rats, where the levels of the AT1 relative to those of AT2 receptor are much lower, pharmacological inhibition of the AT1 receptor alone was sufficient in increasing AT2 receptor levels and proliferation in the V-SVZ. Our data revealed that interactions between RAS receptors play a major role in the regulation of V-SVZ neurogenesis, particularly in proliferation, generation of neuroblasts, and migration to the olfactory bulb, both in young and aged brains, and suggest potential beneficial effects of RAS modulators on neurogenesis.This research was funded by Spanish grants from Ministerio de Economía y Competitividad (BFU2015-70523 and SAF2017-86690-R), Instituto de Salud Carlos III (Retic TERCEL RD16/0011/0016, RD16/0011/0017, and CIBERNED), Galician Government (XUGA, ED431C2018/10; ED431G/05), FEDER (Regional European Development Fund), Generalitat Valenciana (Prometeo 2017-030), and Fundación Emilio Botín-Banco SantanderS

Effects of Rho kinase inhibitors on grafts of dopaminergic cell precursors in a rat model of Parkinson's Disease

In models of Parkinson’s disease (PD), Rho kinase (ROCK) inhibitors have antiapoptotic and axonstabilizing effects on damaged neurons, decrease the neuroinflammatory response, and protect against dopaminergic neuron death and axonal retraction. ROCK inhibitors have also shown protective effects against apoptosis induced by handling and dissociation of several types of stem cells. However, the effect of ROCK inhibitors on dopaminergic cell grafts has not been investigated. In the present study, treatment of dopaminergic cell suspension with ROCK inhibitors yielded significant decreases in the number of surviving dopaminergic neurons, in the density of graft-derived dopaminergic fibers, and in graft vascularization. Dopaminergic neuron death also markedly increased in primary mesencephalic cultures when the cell suspension was treated with ROCK inhibitors before plating, which suggests that decreased angiogenesis is not the only factor leading to cell death in grafts. Interestingly, treatment of the host 6-hydroxydopamine-lesioned rats with ROCK inhibitors induced a slight, nonsignificant increase in the number of surviving neurons, as well as marked increases in the density of graft-derived dopaminergic fibers and the size of the striatal reinnervated area. The study findings discourage treatment of cell suspensions before grafting. However, treatment of the host induces a marked increase in graft-derived striatal reinnervation. Because ROCK inhibitors have also exerted neuroprotective effects in several models of PD, treatment of the host with ROCK inhibitors, currently used against vascular diseases in clinical practice, before and after grafting may be a useful adjuvant to cell therapy in PDThis work was supported by Spanish Ministry of Health (PI12/00798 and RD12/0019/0020) and Spanish Ministry of Economy and Competitiveness (BFU2012-3708)S

Neuroprotective effects of insulin-like growth factor II in a mouse model of Parkinson's disease

Progressive degeneration of the nigrostriatal dopaminergic pathway is a core, currently irreversible pathological hallmark of Parkinson’s disease (PD) that leads to a variety of motor and non-motor symptoms. Here, we aimed to study the potential neuroprotective effects of insulin-like growth factor II (IGF-II) in a PD mouse model based on the chronic administration of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine and probenecid (MPTP/p), which induces loss of dopaminergic neurons in the Substantia Nigra pars compacta (SNc). Male C57BL6/J mice (N=36) received a 5-week treatment with MPTP/p (or vehicle) and were co-treated with chronic IGF-II (or saline) from either the beginning of the procedure (plus an additional week, days 1-44) or once the MPTP/p insult was already triggered (days 21-44). Baseline and post-treatment measurements for motor performance in the Rotarod and self-grooming in an Open Field were taken. Likewise, dopaminergic (TH, DAT) and neuroinflammatory-related (GFAP) markers in the SNc and the dorsal striatum were studied by immunohistochemistry. Our results revealed that both early and delayed IGF-II co-treatment were successful in preventing motor and behavioral impairment in the MPTP/p model. Moreover, chronic IGF-II protected against MPTP/p-induced loss of dopaminergic neurons in the SNc and promoted a significant recovery of dopaminergic activity in the terminals located in the dorsal striatum, further reducing reactive astrocytosis in these brain regions. Thus, we demonstrated the neuroprotective role of IGF-II in a mouse model of PD, highlighting its potential as a promising therapeutical target for treating this disease. Funding: UMA18-FEDERJA-004, PID2020-113806RB-I00. Universidad de Málaga, Campus de Excelencia Internacional Andalucía Tech.Universidad de Málaga. Campus de Excelencia Internacional Andalucía Tech

Small extracellular vesicle targeting of hypothalamic AMPKα1 promotes weight loss in leptin receptor deficient mice

Background and aims: Leptin receptor (LEPR) deficiency promotes severe obesity and metabolic disorders. However, the current therapeutic options against this syndrome are scarce. Methods: db/db mice and their wildtypes were systemically treated with neuronal-targeted small extracellular vesicles (sEVs) harboring a plasmid encoding a dominant negative mutant of AMP-activated protein kinase alpha 1 (AMPKα1-DN) driven by steroidogenic factor 1 (SF1) promoter; this approach allowed to modulate AMPK activity, specifically in SF1 cells of the ventromedial nucleus of the hypothalamus (VMH). Animals were metabolically phenotyped. Results: db/db mice intravenously injected with SF1-AMPKα1-DN loaded sEVs showed a marked feeding-independent weight loss and decreased adiposity, associated with increased sympathetic tone, brown adipose tissue (BAT) thermogenesis and browning of white adipose tissue (WAT).Conclusion: Overall, this evidence indicates that specific modulation of hypothalamic AMPK using a sEV-based technology may be a suitable strategy against genetic forms of obesity, such as LEPR deficiencyMinisterio de Ciencia y Universidades co-funded by the FEDER Program of EU (CD: BFU2017-87721; RN: RTI2018-099413-B-I00 and RED2018-102379-T; ML: RTI2018-101840-B-I00, PID2021-128145NB-I00 and PDC2022-133958-I00). “la Caixa” Foundation (ID100010434), under the agreement LCF/PR/HR19/52160022 (ML); EuroNanoMed III (RA & ML: EURONANOMED2019-050-ENAMEP); European Research Council (RN: ERC Synergy Grant-2019-WATCH-810331)S