In order to calibrate stem cell exploitation for cellular therapy inneurodegenerative diseases, fundamental and preclinical research in NHP(nonhuman primate)models is crucial. Indeed, it is consensually recognized that it is notpossible to directly extrapolate results obtained in rodent models to humanpatients. A large diversity of neurological pathologies should benefit fromcellular therapy based on neural differentiation of stem cells. In thecontext of this special issue of Primate Biology on NHP stem cells, wedescribe past and recent advances on cell replacement in the NHP model ofParkinson's disease (PD). From the different grafting procedures to the variouscell types transplanted, we review here diverse approaches forcell-replacement therapy and their related therapeutic potential on behaviorand function in the NHP model of PD. PD is defined by the presence ofA better comprehension of the physiopathology of PD and the establishment ofnew therapies requires an in-depth investigation of early stages of thedisease, including pathophysiological characterization of (i) evolving non-motorsymptoms, (ii) the restructuring of the central nervous system induced by DA lesioninitiation and (iii) non-motor behavioral manifestations of the disease. Earlyonset of cognitive troubles linked to PD is now recognized , and they are in part due to fronto-striatal loop dysfunction  that can degenerateinto psychological and behavioral troubles, e.g., dementia, with psychiatric symptoms beingidentified in 40–80 % of patients presenting cognitive deficits . Early sleep and circadian disorders have been increasinglyobserved in the majority of patients, concerning many physiologic circadian rhythms. Thus, it appears essential to precisely studythe evolution of neuronal reorganization at play during early stages of thedisease for the identification of new therapeutic targets and also tocharacterize the multiparametric impact of therapeutic treatment on motorand non-motor aspects of PD.The study of behavioral, physiological, anatomical and biochemicalconsequences of DA neuronal death in the basal ganglia was greatly facilitatedby the availability of neurotoxins capable of inducing a highly selectivedeath of DA neurons in animals, e.g.,1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) or 6-hydroxydopamine(6-OHDA) for primate and rodent models .These animal models of PD have been and are still essentials for thedevelopment and amelioration of new therapeutic strategies. It is now wellacknowledged that testing in nonhuman primate (NHP) models is a safe andrequisite preclinical step before any translation to the clinic of braintransplantation. NHPs are more appropriate than non-primates for in vivoscreening, because of their relative closeness to humans, notably regardingbrain organization. In particular, primates share precise cortical developmentphases and associated compartmental growth of inter-areal connections ; late frontal lobe development, both phylogenetically andontogenetically; compartmentalization of meso-frontal projections accordingto a mediolateral gradient ; and functional organization of cortical and subcortical striatalafferents . All these features are critically involved in theevolution and manifestation of PD at different stages.The low-dose MPTP monkey model  is the model of choicefor translational study because it presents a parkinsonian syndromecharacterized by all critical aspects of PD, including a slow progressiveevolution of symptoms, e.g., it replicates the typical motor symptoms used forprimary clinical diagnosis of parkinsonism with response to classical DAtherapy , characteristic pattern of nigrostriataldenervation observed in PD patients , early and long-lasting non-motor symptoms , and increasedMPTP rodent models have been useful to unravel mechanisms underlying DAneuron loss; however, their DA system and sensitivity to MPTP vary highlyacross species  anddiffer from primates . One explanation is thatrodents express extremely small amounts of neuromelanin , which is abundant in human brainstems and is proposed to have a role inlong-lasting toxicity of MPTP in primates .The NHP MPTP model hence reproduces a large repertoire of motor andnon-motor impairments found in PD patients and is thus perfectly suited fora multiparametric evaluation of the therapeutic efficacy of cell transplantas well as for developing and refining techniques for improving integration ofgrafted cells and minimizing potential side effects of the graft. Here wefocus on preclinical investigations done in the gold standard NHP MPTP model. We provide a non-exhaustive review ofdifferent procedures using transplantation of various cell sources as atherapeutic approach in the NHP MPTP model of PD and provide an update onlinked therapeutic behavioral and functional outcomes.2The first attempts to treat PD with cell replacement date back to the early1970s, with the idea of reestablishing striatal DA transmission and restoring aregulated release of DA in the striatum. Two different cell sources for auto-and allografts  paved the way for brain repair through celltransplantation. Adrenal medulla grafts which contained chromaffin cells thatsynthesized DA were grafted in 6-OHDA rats as an alternative source ofcatecholamine-producing cells with the benefit of avoiding ethical and immunological issues linked to thegraft of fetal tissue. Even if testing in the 6-OHDA NHP model returnedminimal survival of transplanted adrenal medullary tissue , it was rapidly followed by first clinical trials in the few patientsreporting clinical improvement .However, grafts in larger cohorts of PD patients returned ratherunsatisfactory results, calling for more careful investigations at thepreclinical level .2.1The fetal ventral mesencephalon (fVM), which generates DA neurons duringdevelopment, constituted another promising candidate for cell grafting. Inthe late 1970s, transplantation of rodent fVM tissue intothe lateral ventricle adjacent to the putamen of 6-OHDA-lesioned rats as analternative to L-DOPA treatment was first performed  and showed that fVMtissue could survive, innervate the host striatum, release dopamine and reverse manyof the behavioral deficits in this PD model. The functional and behavioraloutcome of brain transplantation was then investigated further in 6-OHDA ratsfor several years until the first trial of transplanting human fetal DAneurons was performed in the same rodent model . At thetime, testing of fVM striatal grafts on the recently developed NHP MPTP model returned satisfactory results, with behavioral recoveryin monkeys exhibiting mild to severe parkinsonism ,confirming trials in the rodent model of PD. First clinical trials  were again initially promising  butwere later held back due to efficiency concerns following studies with a largernumber of patients and randomized, double-blind, placebo-controlled protocols that showed no significantdifference between grafted patients and placebo, with several patientsdeveloping graft-induced dyskinesia.Nevertheless, retrospective analyses andreports on those clinical trials of fVM striatal graftsdemonstrated their efficiency either clinically ,histologically , behaviorally  orfunctionally . Based on all these evidences, a newclinical trial of fVM transplantation has been started on a cohort ofpatients with improved grafting procedures and selection of patients  with the aim of preparing for the move to stem-cell-basedtransplantation in humans while waiting for preclinical investigationreports in animal models.The development of the MPTP NHP model of PD contributed enormously to thestandardization of the methodology, due to the possibility of studying largecohorts of monkeys – in particular, in determining the optimal stage of thedonor fetuses , the conditioningof the grafted sample, the site of transplantation  andthe mechanism of graft-induced recovery . Morerecently, transplantation methods and sites of placement of the transplantedcells were revisited and refined in the NHP model .All the transplantation studies performed with fVM in NHPs were based on theassumption that fVM was enriched in immature DA neurons, although most ofthe studies were performed without any detailed analyses of the cellularcontent of the grafted tissue. However, fVM contains a heterogeneous populationof cells whose composition fluctuates during development and comprises arelatively low proportion of DA progenitors divided in two subtypes:(1) progenitors that will give rise to A9 neurons of the SNpc that express theG-protein-gated inwardly rectifying KProgress in the field of stem cells brings hope that this type ofcell therapy could be generalized to treat PD patients. A number ofpluripotent stem cells (PSCs) have been tested in NHPs, isolated either fromearly stage embryos  or from reprogrammedsomatic cells . PSCs have the capacityto become any cell types in the body, including dopaminergic progenitors andneurons. They thus constitute an infinite source of cells for transplantationinto PD patients.We will now focus on the transplantable DA cell types generated from primatePSCs, which represent the closest to clinical application. Human ESCs  and monkeyESCs  were first used, recently followed by human iPSCs and monkey iPSCs .2.2Various protocols available for the generation of DA neurons from human andNHP PSCs were adapted from those developed with mouse ESCs . Early protocols aimed atfirst inducing neural differentiation of PSCs generally by culturing the PSCswith stromal cells (PA6 cells or MS5 mouse lines) or in the presence ofmedium conditioned by these cells . Other protocols forneural differentiation involved suspension cultures to generate embryoidbodies and culture in serum-free medium . These protocols enable a significant enrichment of the population intoneural progenitors that expressed NESTIN, SOX1,PSA-N-CAM ,PAX6 and SOX2.Midbrain DA specification of these neural precursors can then be induced byaddition of FGF8, a mid- and hindbrain organizing morphogen, and SHH, aventralizing morphogen , and/or by treatment with FGF2 and FGF20 – a secreted protein thatenhances the survival of primary DA neurons . Characterization of the cells showed thatDA neurons express midbrain DA neuron markers such as NURR1 and LMX1A, LMX1B,FOXA2, OTX2, CORIN, PITX3, factors that control specification anddifferentiation of midbrain DA neurons during mouse development , and GIRK2, which is the A9-specific marker . They also express tyrosine hydroxylase (TH), and the dopaminetransporter (DAT), and they produce dopamine, confirming that they arefunctional DA neurons .Although these methods enabled efficient DA differentiation, the culturesusually comprise a high percentage of glial cells and multiple neuronsubtypes, such as GABAergic, cholinergic and serotonergic neurons . Complete and robust midbrain specificationwas recently obtained via a floor plateintermediate stage from human and NHP PSCs , using a modifieddual-SMADinhibition protocol . However, major differences in human and rodent ventralmidbrain development have recently been identified .In light of these results, studies have been engaged to develop and refinethe existing protocols and tools to generate DA neurons from primate PSCsin vitro that better mimic their in vivo counterparts. In this line,identification of a set of new markers of human mesencephalic DA progenitors enabled Kirkeby and collaborators todevelop a differentiation protocol that leads to increased yield of DAprogenitors from human ESCs . These markers enableproper prediction of the resulting DA neuron content in the graft and willenable further standardization of DA differentiation protocols from PSCs.These last generation of PSC-derived DA progenitors are produced undercareful Good Manufacturing Practice (GMP) laboratory conditions  and bare authentic fVMfeatures: they are able to reinnervate the lesioned striatum and functionwith equal potency to fVM tissue upon striatal transplantation in the PDrodent brain . Based on recent preclinical in vivo assessments mostlyperformed in the rodent model and more recently in the NHP model, severalclinical trials have been recently launched worldwide using human ESC DAcells, such as the European trial “STEM-PD” .DA neurons can also be generated by direct reprogramming of somatic cellssuch as mouse and human fibroblasts, using overexpression of differenttranscription factors and midbrain-specific factors such as LMX1A, ASCL1, NURR1,BRN2, FOXA2, NGN2, SOX2 and MYT1L . These induced DA (iDA) cells ledto reduction of motor symptoms when transplanted in the striatum of PD rodentmodels  and represent an interesting cell source thatstill needs to be pre-validated in the NHP model.The intended application of reprogrammed cells (either iPSCs or iDA cells)and their derivatives is autologous transplantation. However, their isolationand deep characterization usually needs several months, which constitutes animportant drawback when considering that transplantations have to be donewhen the system is still reparable. Furthermore, because they are derivedfrom PD patients, they may present genetic and epigenetic alterations andmay be more susceptible to the pathological processes induced in PD.2.3Multipotent neural stem cells (NSCs) present several advantages over DAprogenitors or neurons. They constitute homogeneous cell populations that canbe expanded on a very large scale and extensively characterized, and they canbe kept as frozen stocks that are ready to use for transplantation. Thisensures high levels of batch-to-batch consistency and enables optimaltraceability of the grafted cells. Another advantage of NSCs is that they arecapable of giving rise to neuronal and non-neuronal progenies, such asastrocytes and oligodendrocytes. They are thus potentially capable ofrespondingand adapting to the host environment, making them one of the best candidates torestore a functional equilibrium in the nigrostriatal system .Human NSCs have been isolated from fetal brains  or from PSCs  throughadaptation of the protocol used to derive mouse NSCs . They express the stem cell markers NESTIN, SOX2,VIMENTIN and MUSASHI; are karyotypically normal; and can be amplified for longperiods of time in culture without losing their properties . Following testing for various contaminants , they constitute working cell banksprepared under GMP-grade conditions . NSC lines can further be carefully preselectedfor transplantation on the basis of their capacity to differentiate into DAprogenitors in response to DA induction in vitro and to give riseto various neuronal and glial cell types upon transplantation into differentregions of the mouse brain in vivo .Human NSCs have shown very promising results upon injection in SN andstriatum of MPTP monkeys . Although transplantations ofhuman cells into the brain of NHPs bring crucial information, NSC safety andefficiency within an NHP species still need to be confirmed. This would implythe isolation and deep characterization of NHP NSC lines that are stillsparse .2.4Autologous neural cell ecosystems (or ANCE) represent an interestingalternative for autologous transplantation. These cells are derived from theadult monkey cortex; they do not require genetic modification and can beexpanded in only a few weeks in vitro . They do not express specific DA markers, but progenitor markerssuch as GFAP ,NEUROFILAMENT, VIMENTIN and NESTIN . It isstill unclear whether these cells are able to differentiate into DA neurons. They showed very good survival and integration uponauto transplantation in striatum and SN of the PD NHP brain, which is associated withbehavioral recovery . However, this system needs carefulcharacterization in the NHP model to evaluate its usefulness in futureclinical application.Human retinal pigment epithelial (hRPE) cells may constitute another suitablecell type for transplantation in PD. They are isolated from the inner layerof the retina of postmortem fetal eyes and produce L-DOPA as a precursor oftheir characteristic eumelanin pigment through the activity of TH. They canbe easily expanded in culture and stored as frozen stocks, allowingextensive characterization and testing before transplantation. hRPE cellsattached to gelatin microcarriers (GM) show enhanced survival and improve motordeficits after striatal grafts in rodents  and in NHP models of PD . RPE-GM are currentlyused in human PD patients under the name of Spheramine. The mechanism of action of RPE cells may notbe the production of dopamine, but constant in situ release of lowphysiological level of levodopa, which may stimulate DA synthesis fromsurviving DA neurons.In 2011, the results of the first randomized, double-blind,placebo-controlled trial of RPE cell transplantation in PD patients showedthat hRPE cells provided no antiparkinsonian benefits over sham surgery . Optimal therapeutic benefit could be reached afteradministration of low doses of levodopa. These results have been recentlyconfirmed in NHPs where unilateral striatal transplantation of hRPE-GM wasnot sufficient to completely reverse the motor symptoms of inducedparkinsonism . Studies performed in the NHP model will becrucial to refining the characteristics of these RPE cells used fortransplantation, in particular the optimal age of the donors. Fetal hRPEcells indeed showed higher survival than those obtained from neonatal donors,at least in the rat PD model . The NHPmodel may also help in evaluating the therapeutic effect of other sources ofhRPE cells and other formulations of microcarriers to increase cell survival.2.5As far as xenografts are concerned, human-grafted cells are easilydistinguished from the host primate brain using human-specific markers, suchas the human cytoplasmic markers hCy  and STEM121 . In contrast,localizing the NHP cells upon transplantation in NHP models necessitateslabeling cells before grafting.fVM tissue cannot be labeled prior to transplantation; its progenies arethus commonly detected by simple TH immunofluorescent staining. This strategyenables locating the transplanted tissue, but the migration of the graftedcells and connection with host THIn contrast to fVM tissue, NSCs, DA progenitors and DA neurons can easily belabeled in vitro, prior to grafting. This was originally performedthrough incorporation of BrdU . The disadvantage of this labeling is that not allcells become labeled with BrdU, leading to underestimation of cell survival. With the emergence of newtechnologies, cells are now frequently labeled with fluorescent proteins(GFP or RFP) after electroporation of a GFP-expressing plasmid and antibioticselection , or lentiviral infection . Although the efficiency and stability of the labeling were not alwaysdocumented, this type of labeling remains essential to assess the migrationand facilitates visualization of neurite extension of the grafted cells. The fluorescent dies PKH26 and PKH67 can also becited , but their propensity to fadeovertime makes them inappropriate for long-term labeling.So far, GFP and RFP labeling seems most appropriate for evaluating integrationand differentiation rates of grafted cells. However, some studies reportedGFP-induced cytotoxicity  and, morerecently, dose-dependent toxicity in DA neurons in rats after cytoplasmic GFPtransfection . While this toxicityseems to be more associated with the level of expression of this reportergene in transfected cells, this should be carefully taken into account whileevaluating the efficacy of GFP-expressing transplant in NHPs.It is noteworthy that senescent cells found in the aged and degenerativebrain produce autofluorescence that interferes with detection of specificfluorescent signals, implying rigorous manual tracing analyses oftransplanted cells in the case of fluorescent labeling .Noninvasive methods developed for tracking cells in real time after graftingmight also help to improve the design of future clinical celltransplantation. A nice example is cell labeling with superparamagnetic ironoxide (SPIO)nanocomposites, which allow cell tracing in living animals usingMRI .33.1In studies performed with fVM, transplanted cells in the striatum consist ofsmall pieces of fVM tissue .It is noteworthy that early DA neurons tend to survive better upon graftingin the brain of PD monkeys than do terminally differentiated DA neurons, aspreviously shown in rodents . For example, VM tissueisolated from early stage fetuses showed a higher survival rate than that ofolder stage fetuses , as was previously shown in rodents . Similarly, early DA progenitors derivedfrom primate PSCs (D21–D28 neurospheres) produced larger grafts  and showed a higher survival rate  than did late DA neurons . Nevertheless, the survival rate never exceeded 5 %.In contrast to DA neurons, uncommitted neural cells show a much highersurvival rate that is generally close to 10 % for NSCs and over 40 % in the case of autografts of immature neural cells derivedfrom the adult cortex . The higher survival rateof immature cells, as compared to that of terminally differentiated neurons,could be explained by their multipotent features, which allow them to bestadapt to environmental fluctuations. They also exhibit minimal neuriteoutgrowth and may thus be less sensitive than differentiated cells tomechanical stress during their isolation and transplantation procedure.Interestingly, increasing the number of grafted cells did not improve cellsurvival proportionally in MPTP NHPs, regardless of their origin . It has been hypothesized that ahigh cell number might induce host rejection in the case ofallotransplantation and that it may also exhaust the supply of neurotrophicfactors that are present in low amounts in the adult diseased brain.Grafted cell survival can be improved through implantation of micrografts orcells over several sites within striatum or SN of NHPs , confirming earlier rodent studies . Another strategy to increase cell survival of grafted cells involvesdelivery of neurotrophic factors combined with cell transplantation . In this context, glial cell-derivedneurotrophic factor (GDNF) is one of the most widely tested neurotrophicfactors, and it is well known to promote survival of DA neurons in vitro and in vivo in different animal models of PDincluding NHPs, where it prevents degeneration of DA neurons . The neurotrophic actionsof GDNF in PD have been extensively described elsewhere .Neurexophilin 3 (NXPH3), a synapse-related peptide, may be also be used incombination with cell grafts for promoting cell survival. It was recentlyshown to support survival of mouse iPSC-derived DA neurons in vitroand in vivo, when combined with grafted cells .\opttitle{Number of TH $+$ cells required for functionalrecovery}Despite the low-cell surviving rate of the grafted cells, several studies showedbehavioral improvement upon cell grafting in MPTP NHPs (Table 2). Thissuggests that a minimal restoration of normal DA innervation in the striatumis sufficient for functional recovery, as previously shown in the rodentmodel . However, a certain threshold of THThe required number of surviving TH3.2The principal focus when analyzing the graft composition is on DA neurons.The most widespread marker of DA neurons is tyrosine hydroxylase, theenzyme that catalyzes the conversion of tyrosine to dihydroxyphenylalanine,which is the first step in the biosynthesis of dopamine. However, TH is also involvedin the synthesis of other catecholamines such as epinephrine andnorepinephrine. The identity of DA neurons is thus usually refined bystudying the expression of DAT through immunostaining analyses or measurement of the bindingpotential of As previously mentioned, PSC dopaminergic differentiation in vitro leads tovariable amounts of THNSCs, isolated either from fetuses or PSCs, generally poorly differentiatedinto TH3.3One of the critical issues of cell therapy in PD is the capacity of thetransplanted cells to grow axons and reinnervate the DA-denervated hoststriatum over distances that are relevant for the size of the human brain.The adult brain has been suspected of no longer being capable of eliciting anddirecting axonal outgrowth from the SN to the striatum. Grafted cells havethus often been placed ectopically into the striatum, which is the site of lostdopaminergic input .When transplanted in the lesioned rodent striatum, DA neurons from human fVMand PSCs show extensive reinnervation of striatal and extra-striatal targetstructures , and, when grafted inthe SN, they project axons over long distances and reinnervate the relevantA9 and A10 host target structures , as was observedfor rodent DA neurons .In contrast, their capacity to reinnervate distant targets seems ratherlimited in the NHP brain, with THTo promote reinnervation of the nigrostriatal circuitry, multipleintrastriatal and intranigral fVM grafts have been used as “bridge grafts”that attract the growth of neurites from grafted DA neurons in the rat and NHP models. Using this strategy, fVMgrafts placed in the SN extended neurites over long distance preferentiallyto striatal co-grafts, suggesting that axon guidance cues are still presentin the lesioned brain to guide the growing axons from the grafted DA neuronsto their appropriate targets.The nature of these guidance cues is still poorly known in the lesioned NHPbrain. Several molecules such as the NETRIN, SLIT, EPHRIN and SEMAPHORIN families ofsecreted proteins, whose axon guidance activities have been extensivelystudied in the nervous system, have been shown to affect embryonic DA axonsand may be involved in the regulation of axonal outgrowth of transplantedcells in the lesioned NHP brain. NETRIN-1 attracts whereas SLIT-2 repelsrodent midbrain DA neurons in vitro and in vivo , and both proteins affect human PSC DA neuron outgrowthin vitro . Signaling through the EPHRIN familyreceptor EphB1 and ligand EPHRIN-B2 is involved in the regulation of axonalgrowth of developing DA neurons in rodents . EphA4 receptor and EPHRIN-B2 ligand are expressed in the adult NHPbrain, including the CdN, putamen and SN , and may play arole in directing axonal outgrowth of grafted cells. SEMAPHORINs areexpressed in the rodent striatum and may be involved in the establishment ofDA projections from the midbrain to the striatum during embryonic development. SHHsignalling is involved in DA axon pathfinding and determination of thestructural diversity of the DA projections during rodent development and mayalso promote axonal growth of grafted cells . Whetherthese guidance cues persist in the adult or lesioned NHP brain is stillunknown.In addition to its action on DA neuron survival, GDNF also stimulatesoutgrowth of DA neurons after lesion or grafting in the rodent  andNHP brain . Combination of GDNF and NETRIN-1 was found to support directedlong-distance growth of DA axons from rodent fVM grafts .However, recent studies showed that GDNF delivery combined with fVM graft didnot lead to increased functional improvement in the PD NHP model , highlighting the need to clarify the benefit of GDNF delivery inthis context.Besides axonal outgrowth, migration of the transplanted cells is alsoinvolved in the reinnervation of the lesioned striatum over long distances.In the rodent model, transplanted primate DA neurons extensively migrate, evenreaching the contralateral hemisphere . In theNHP model, the sparse data available suggest that migration of DA neurons islimited, as judged by the lack of expression of the migrating neuroblast marker,doublecortin . In contrast to DA neurons,primate NSCs show widespread migration throughout the MPTP-lesioned NHP brain as was previously shown in various rodent models . In particular, NSC progenies were observedmigrating along the nigrostriatal pathway, from the caudate to the putamen. The phenomenon of migrationof immature cells is exemplified by unilateral injections, where the graftedcells were found migrating to the opposite hemisphere . Interestingly, human NSCs spontaneously andpreferentially migrate to the region of cellular loss over long distances inthe lesioned PD NHP brain , suggesting that migrationis not a random event. The signals that direct NSC migratory pattern in theMPTP model are not known. Progenies of human NSCs express the chemokinereceptor CXCR4, suggesting that chemokine-dependent mechanisms are involvedin the regulation of their migration .Immature cells develop very slowly in the host environment, and this may takeseveral months before they generate terminally differentiated progenies,including fully functional mature DA neurons that extend projections in thenigrostriatal pathway. Studies of axonal outgrowth and cell migration arethus more appropriate in the NHP model, which allows long-term analyses aswell as studies to assess the capacity of grafted cells to innervate the hostbrain over sufficiently long distances to provide good innervation of theremote putamen in primates.3.4From the earlier transplantation studies of adrenal medullar cells  to the most recent studies of NSCs and DA neurontransplantation , the functional effects of graftedcells were hypothesized to be obtained not exclusively by a cell-replacementmechanisms but also through diffuse release of neurotrophic stimuli andneuroprotective support on host circuitry . NSCs can express and produce in situa wide array of transmembrane and trophic molecules capable of promotingtissue repair. GDNF and BDNF are expressed in glial cells derived from thegrafted NSCs, in both the rodent and NHP models, and may provide trophicsupport to the pathological host milieu .Close physical associations have commonly been observed between grafted cellsand host cells, indicating intercellular relationships. For example, DAneurons from transplanted fVM tissue or isolated from PSCs establish synapticcontacts with host striatal neurons by targeting dendrites and somata ofspiny neurons , as visualized by synaptophysin staining.Differentiated progenies of hNSCs were also found in close contact with hostTH3.5One major concern with regard to the use of PSCs-based therapy is the risk ofovergrowth or development of tumors. In the NHP model, tumor formation orovergrowth was generally not detected following transplantation ofPSC-derived DA neurons  or immature NSCs, as judged by the negative staining for Ki67, amarker of proliferative cells . Asexpected, pluripotent nuclear  and membrane markers  were also not found in these grafts .Proliferating cells have occasionally been detected after transplantation ofhiPSC-derived DA neurospheres .However, prolonged PSC DA differentiation in vitro enabled a drasticreduction of tumorigenicity  as previously shown in the rat.However, depending on the protocol used to obtain DA neurons, the population ofgrafted cells may still contain immature proliferating cells that may, in somecases, be incapable of differentiating in the host tissue and eventually formtumors in vivo. The possibility of uncontrolled growth of the grafted cellsin vivo should still be a matter of concern. This highlights the need fordeep characterization of the grafted cells and long-term preclinicalstudies in the NHP model to validate the safety of each cell type.4The brain has long been positioned as immune privileged, but it is now wellestablished that this privilege is not absolute and that immunologicalrejection processes can occur in the CNS . Transplanted tissue or cells might thus berecognized as foreign, leading to their rejection after transplantation inthe brain. Fetal VM tissue expresses MHC antigens that can elicit an immuneresponse in the case of host mismatch . Expression levelof MHC molecules is low in human PSCs and NSCs  but can rapidly be induced in inflammatory conditions orfollowing differentiation in vitro .Xenografts in the MPTP-treated NHPs have thus usually been performed underimmunosuppression (Table 2). In the immunosuppressed environment,transplanted cells elicit only weak immune reaction, with minimal glialscarring and host IBA1In the case of allografts in NHPs, immunosuppression has exceptionally beenused, and, when evaluated, immune response or inflammatory reaction wasgenerally found to be weak (Table 1b).However, direct comparison of allo- and autografts of iPSCs in healthy NHPsreported immune reaction in allografts, with the presence of host microglialcells that expressed MHC-II, IBA1These recent studies suggest that immunosuppression can only be withdrawn inthe autologous models . Direct comparison of autograft and allografts in the lesionedNHP brain might ultimately confirm the efficacy and safety of autologousiPSC-derived or iDA cells.5Callithrix jacchus), parkinsonism can be modeled with MPTP,6-OHDA or also through overexpression of In marmosets (Cercopithecus aethiops sabaeus)all rely on the model developed by Redmond and collaborators  and cynomolgus (Macaca fascicularis) monkeys, MPTP is administered either acutely, leading to rapidnigrostriatal lesion and expression of motor symptoms, or chronically overweeks to months, inducing a progressive DA lesion and slowly evolvingnon-motor and motor symptoms. The main difference between these models isthat acute MPTP models induce a topography of the DA lesion different thanthat produced by chronic MPTP intoxication, following which the ventralstriatum is the most preserved structure comparable to what is observed in PD patients .Sanchez-Pernaute and collaborators grafted DA cells derived from ESCs (cyno-1line) in the anterior and posterior part of the right Put of one parkinsoniancynomolgus monkey after repeated intravenous MPTP injections implanted iPSCs in the anterior and posterior parts ofCdN; in the anterior, mid- and posterior parts of Put; and in SN did not report any in vivofunctional,e.g., with PET-scan, nor behavioral outcome of the grafts (6 studies usedasymptomatic MPTP monkeys and one non-MPTP-treated monkeys), focusingexclusively on postmortem evaluation of host–graft integration . In theseconditions it is impossible to infer the degree of integration required forany behavioral or functional recovery. However, even if those studies arenecessary for testing different types of transplant before investigation ina larger number of animals, reporting clinical motor score has been very welldocumented for many parkinsonian models and should be an imperative standard,considering that those investigations require sacrifice of NHPs. From theremaining studies, about half of them reported in vivo functional outcomefollowing graft in addition to behavioral outcome assessing clinical motorsymptoms; however, some studies just reported qualitatively those functionalchanges . Surprisingly, and even so non-motor symptoms are recognized ashaving a great impact on quality of life in PD patients , no graft study using the NHP MPTP model to date has reported effectson non-motor symptoms. We found one study in the 6-OHDA marmoset modelreporting effects of fVM grafts on a version of the detour task used toassess integrity of DA innervation to the frontal cortex .Assessing non-motor symptoms has just lately being explored in a rodent modelof PD  and will hopefully pave the way for systematicfollow-up of non-motor symptoms in cell-repair research for PD.6Grafting in NHPs will enable the development of techniques for more detailedin vivo and postmortem follow-up of the fate of the cells that can betranslated to humans. Because NHP and human cells share the samecharacteristics and cell signalling regulations, NHPs are potentially the mostappropriate for in vivo screening.https://ec.europa.eu/health/scientific_committees/consultations/public_consultations/scheer_consultation_03_enSCHEER provides recommendations, after public consultation, on how to advancetraining, improvement of techniques and protocols, sharing of knowledge,removal of barriers and research needs for NHP use.. Progress was made, butit was not enough to justify a reduction in the use of NHPs in neurodegenerativedisease research. Rather, both advances in recent promising techniques and theneed for more preclinical design studies in NHPs call for a controlledincrease in the use of NHPs in neurodegenerative research. In return, rigorousindividual follow-up with pertinent functional imaging evaluations during thedifferent intoxication and transplantations phases (pre/post) should bestandard practice for preclinical observations.However, the burdens from working with NHPs are significant in terms of time andcost investments needed to pursue preclinical investigations and/or providesignificant contribution to scientific knowledge about the underlyingmechanisms of PD. Limitations are often required by ethic committees whichfollow a country's directives; for example, recently the European Commission asked theScientific Committee on Health, Environmental and Emerging Risks (SCHEER) toupdate the opinion on the use of nonhuman primates inresearchWe have seen that site of transplantation is critical for behavioralrecovery. Studies originally placed transplants into the striatum, the targetof DA neurons for practical reasons, and showed that placement into Put leadto better motor recovery compared to caudate transplants . Multiple small transplants  are also an efficient solution to implant more cells, but not in onesingle location, which has been showed to be detrimental . Combination of multiple target areas might promotereconstruction of the nigrostriatal pathway but still needs further refinement inNHPs. No clear evidence of nigrostriatal reconstruction has been shown sofar, neither with combined grafts in striatum and SN  nor in SN grafts with trophic factors in the striatum. However, interhemispheric cell migration is possible, and transplanted cells favorthe natural nigrostriatal connectivity patterns for selective neuriticoutgrowth . Indeed, basal ganglia circuitry andespecially nigrostriatal and striato-cortical connectivity patterns are welldescribed in NHPs . According to these schemes, it is more than likely that graftsplaced at given striatal sub-compartments would influence the specificsystem they support – e.g., graft placed in the caudal CdN might be moreefficient in affecting cognitive troubles, whereas putaminal grafts mightcorrelate better with motor recovery. With increased understanding of growthand guidance molecules affecting DA neurons, it may be feasible to placetransplants in the damaged SN and direct the growth of axons into targetregions for reconstruction of midbrain DA circuitry. Our established andongoing understandings of the molecular cues which support directed growthof DA neurons form an important basis for the refinement and optimization ofgrafting procedures. Adding supporting factors for survival and axonaloutgrowth of grafted cells  and combining different cell types will be the next step in therefinement of the technique.The latest developments in the field of neural tissue engineering could not beimplemented here, but shall certainly be taken into account in futuretransplantation strategies in PD NHPs. Various biocompatible and biodegradablebiomaterials are currently being developed and may enhance survival andintegration of the grafted cells .While the DA system is not the only neurotransmitter system altered, DAremains central in PD. DA plays an essential role in (i) multiple cerebralfunctions of the frontal lobe, e.g., in performance monitoring or inmotivational aspects of behavior; (ii) motorfunction via the extrapyramidalsystem; and (iii) circadian regulation of behavior through interactions withlateral habenula and locus coeruleus. This multidimensional aspect of DAshould trigger a multiparametric follow-up of the NHP model of PD. Longitudinaland simultaneous multiparametric follow-ups  should be carried out asmuch as possible given the NHP PD model chosen in future experiments. Indeed,non-motor symptoms have only been reproduced through repeated low-dose MPTPregimen . Overall, very few studies follow uptransplanted monkeys for more than a year . However, a longeroutcome period should be considered (2–4 years post-implantation) in orderto assess potential long-term side effects, graft rejection or changes infunctional in vivo measures, especially in cases that present good motorrecovery. All these criteria will enable full validation of the safety andefficiency of cell grafting before clinical translation.Finally, it should be noticed that recently emerging NHP models of PD, e.g.,overexpression of


Intra-aortic balloon counterpulsation is increasingly used in acute myocardial infarction complicated by cardiogenic shock. The aim of this study was to explore the preference, effect, and prognosis of intra-aortic balloon counterpulsation in acute myocardial infarction complicated by cardiogenic shock patients.  Data of acute myocardial infarction complicated by cardiogenic shock patients at the Fourth Medical Center of PLA General Hospital were collected retrospectively. A propensity score was calculated with a logistic regression which contained clinically meaningful variables and variables selected by Lasso and then used to match the control group. The cumulative incidence curve and Gray's test were employed to analyse the effect and prognosis of intra-aortic balloon counterpulsation on mortality. P = 0.454).  A total of 1962 acute myocardial infarction cases admitted between May 2015 and November 2018 were identified, and 223 cases with acute myocardial infarction complicated by cardiogenic shock were included as the study cohort, which contained 34 cases that received IABP and 189 cases that did not receive IABP. Patients with higher alanine aminotransferase , higher triglyceride , and higher blood glucose  had a higher probability of receiving intra-aortic balloon counterpulsation. In the propensity score matching analysis, 34 cases received intra-aortic balloon counterpulsation and 102 matched controls were included in the comparison. By comparing the cumulative incidence of in-hospital mortality, there was no statistically significant difference between the intra-aortic balloon counterpulsation group and matched control group ( The use of intra-aortic balloon counterpulsation may not improve the prognosis of the acute myocardial infarction complicated by cardiogenic shock patients. At present, despite marked advances in medical treatment and revascularization techniques, acute myocardial infarction complicated by cardiogenic shock (AMI-cardiogenic shock) has still higher mortality –4. In thTherefore, the use of IABP supports hemodynamics, but evidence on improving the prognosis of AMI-cardiogenic shock patients was still controversial. We established a retrospective cohort study from a single center, to explore the impact of IABP on the mortality of patients with AMI-cardiogenic shock. By matching patients who have received IABP and patients who have not received IABP, we explored the mortality of patients with AMI-cardiogenic shock in the hospital. In addition, we explored the prognostic factors for mortality in AMI-cardiogenic shock patients receiving IABP. Recommendations are given clinically in order to better assist the treatment of AMI-cardiogenic shock and improve the survival rate of patients.A retrospective cohort was established to investigate mortality associated with IABP for acute myocardial infarction complicated by cardiogenic shock. Patients admitted in the Fourth Medical Center of PLA General Hospital between May 2015 and November 2018 with a diagnosis of acute myocardial infarction were identified from the electronic medical records. The exclusion criterion was acute myocardial infarction not complicated by cardiogenic shock.Demographic, clinical, laboratory, and clinical outcome data were obtained from the hospital's electronic clinical medical records. At the first clinical consultation, demographic, clinical, and laboratory data were collected within 24 hours after admission. The outcome of interest was in-hospital death, with discharge as the competing risk event, and event time was defined as the time from admission to either event whichever came first.Continuous variables were presented as median and interquartile range (IQR), and categorical variables were presented as number and its corresponding percentage. Variables with a right-skewed distribution were log-transformed before being included in the analysis. Missing values were imputed with single imputation. Univariate logistic regression was used to explore the risk factors of receiving IABP in AMI-cardiogenic shock patients, and a cause-specific Cox model was used to explore the prognostic factors of mortality in AMI-cardiogenic shock patients receiving IABP.To ensure the assumption of positivity  is met in the propensity score analysis, we used a two-step approach in exploring the predictors of receiving IABP and matching IABP patients with controls.In the first step, variables that led to (quasi-) complete separation were automatically identified as predictors (with an OR being infinite), and a deterministic matching based on these variables was performed . In step two, we used Lasso for variable selection to determine other predictors of receiving IABP, after excluding those predictors already identified in step one. The selected variables were used as matching variables (in addition to age and sex) in the propensity score matching analysis (with a ratio of 3 : 1). The number of variables selected by Lasso was determined with the 1-in-10 rule of thumb: number of selected variables = max. The cumulative incidence curve and Gray's test were used to compare the in-hospital mortality of patients receiving IABP and matched controls. Statistical analyses were conducted using R software (version 3.6.1) and packages dplyr, mice, glmnet, Hmisc, AUC, survival, cmprsk, descry, and forestplot.A total of 1962 acute myocardial infarction cases admitted between May 2015 and November 2018 were identified. After excluding 1739 cases not complicated by cardiogenic shock, 223 cases with AMI-cardiogenic shock were included as the study cohort, which contained 34 (15.25%) cases that received IABP and 189 cases that did not receive IABP .Univariate logistic regression revealed that patients with higher drinking history, heart rate, hyperlipidemia, ischemic cardiomyopathy, diabetes, alanine aminotransferase, aspartate aminotransferase, cholesterol, triglyceride, glycosylated hemoglobin, and blood glucose and younger age had a higher probability of receiving IABP . All IABAfter excluding non-IABP patients in cardiac insufficiency and nonhepatic insufficiency groups, each IABP patient was matched with three controls by the propensity score calculated based on age, sex, alanine aminotransferase, triglyceride, and blood glucose. Finally, 102 patients not receiving IABP were selected as the control group . The disP = 0.454; When comparing the cumulative incidence of in-hospital mortality, there was no statistically significant difference between the IABP group and the matched control group (Univariate Cox regression revealed that patients receiving IABP with a higher heart rate and serum potassium had a higher probability of death .This retrospective cohort study revealed that the use of IABP cannot improve the mortality of AMI-cardiogenic shock patients. Despite the fact that there are still great controversies about the use of IABP that can improve the prognosis of AMI-cardiogenic shock patients and some observational studies revealed a trend toward lower mortality for patients if they received IABP, these findings may result from the clear inequality of baseline risk factors , 19. In From a pathophysiological point of view, the main feature of death in AMI-cardiogenic shock is unstable hemodynamics with reduced systolic and mean arterial pressures that lead to reduced oxygen supply to vital organs , while tOur results revealed that higher heart rate and serum potassium may lead to higher mortality in AMI-cardiogenic shock patients receiving IABP. A previous study revealed that serum potassium and heart rate can increase the risk of mortality of patients with AMI . TherefoOur study has several limitations. First, the sample size of our study is not large enough. Therefore, the research may not have good statistical power, which may be one of the reasons why statistically significant result was not observed. Therefore, a larger cohort is needed to further study the effect of IABP on the mortality of patients with acute myocardial infarction complicated by cardiogenic shock. Second, this is a retrospective single-center study, and some laboratory tests were not taken within the first day after admission. We used an imputation method to deal with these missing values; however, a single imputation may lead to uncertainty in results. A prospective cohort will provide stronger evidence.In conclusion, the use of intra-aortic balloon counterpulsation may not improve the prognosis of the acute myocardial infarction complicated by cardiogenic shock patients.


Although great breakthroughs have been made in cancer treatment following the development of targeted therapy and immune therapy, resistance against anti-cancer drugs remains one of the most challenging conundrums. Considerable effort has been made to discover the underlying mechanisms through which malignant tumor cells acquire or develop resistance to anti-cancer treatment. The Hippo signaling pathway appears to play an important role in this process. This review focuses on how components in the human Hippo signaling pathway contribute to drug resistance in a variety of cancer types. This article also summarizes current pharmacological interventions that are able to target the Hippo signaling pathway and serve as potential anti-cancer therapeutics.Chemotherapy represents one of the most efficacious strategies to treat cancer patients, bringing advantageous changes at least temporarily even to those patients with incurable malignancies. However, most patients respond poorly after a certain number of cycles of treatment due to the development of drug resistance. Resistance to drugs administrated to cancer patients greatly limits the benefits that patients can achieve and continues to be a severe clinical difficulty. Among the mechanisms which have been uncovered to mediate anti-cancer drug resistance, the Hippo signaling pathway is gaining increasing attention due to the remarkable oncogenic activities of its components  and their druggable properties. This review will highlight current understanding of how the Hippo signaling pathway regulates anti-cancer drug resistance in tumor cells, and currently available pharmacological interventions targeting the Hippo pathway to eradicate malignant cells and potentially treat cancer patients. Drosophila and humans, as well as how the activation or inactivation of these molecules  leads to drug resistance. We also summarize the pharmacological agents that enable the intensive anti-cancer efficiency by targeting the Hippo pathway.As a powerful weapon to kill aberrantly fast-growing tumor cells, chemotherapy represents one of the most efficacious treatment strategies for the majority of cancer patients, immensely prolonging progression-free survival and improving quality of life. However, most cancers remain largely incurable, due to their early asymptomatic nature and late diagnosis, local and distant metastasis, and the development of therapeutic resistance. Although combinations of multiple therapeutic agents have become the paradigm for cancer therapy as initial solutions to the resistance against single-agent treatment, drug resistance continues to be a huge obstacle . Till noDrosophila melanogaster, following the identification of four tumor suppressors, Warts  and diap1 (inhibitor of apoptosis), thus accelerating cell proliferation and diminishing cell apoptosis . D. D167]. Cyclin-dependent kinase 9 (CDK9) is the catalytic subunit of the positive transcription elongation factor b (P-TEFb) and phosphorylates the carboxyl-terminal domain (CTD) of RNA polymerase II (RNA-Pol II) . Cancer In addition, another study points out Bromodomain-containing protein 4 (BRD4), a general regulator of RNA-Pol II-dependent transcription, is recruited to the promoter of YAP/TAZ target genes by YAP/TAZ-bound enhancers, thus boosting the transcription of genes involved in tumorigenesis . Hence, Hyperactivation of YAP/TAZ, or loss-of-function of tumor suppressors in the Hippo pathway is able to mediate tumor cell resistance against anti-cancer therapeutic drugs. Depletion or inhibition of YAP/TAZ through pharmacological agents, or re-expression of tumor suppressors in the Hippo pathway reverts the drug sensitivity, suggesting intervention of YAP/TAZ activity is a prospective strategy to overcome drug resistance in cancer cells. In addition to controlling cell growth and apoptosis, the Hippo pathway is also implicated in drug resistance, as documented in a variety of cancer types. Studies indicate significant superiority of pharmacologically targeting the Hippo pathway, particularly YAP/TAZ, over current agents to eliminate cancer cells. However, several critical questions need to be answered for successful future translation of these preclinical studies into powerful clinical strategies. Given that targeting the Hippo pathway shows attractive perspectives in anti-cancer treatment, well-designed clinical trials will be necessary for the successful practice of evidence-based health care. Indeed, FDA-approved non-anti-cancer drugs, such as verteporfin, simvastatin and ibudilast, are under active clinical trials to evaluate their effectiveness in cancer therapy. Considering the reasonable safety of these medicines that have been applied in clinic for many years, their paths to be incorporated into standard cancer treatment would presumably be much shorter than newly invented compounds. This scenario also applies to FDA-approved anti-cancer agents, for example, specific kinase inhibitors that suppress YAP/TAZ activities. Decipher of these mechanisms broaden the scope of the drug applications in cancer treatment, supplying more medications for the cancer types which are relevantly rare and have limited therapeutic options. However, it should be underlined that simply relying on existing drugs or chemicals is not sufficient to solve the drug resistance problems. Inventions of novel compounds with elevated anti-cancer efficiency, substrate specificity and attenuated adverse effects are in demand. For example, although BET inhibitors significantly enhance the vulnerability of melanoma cell under RAF inhibitor treatment, selective BET inhibitors, especially inhibitors selectively targeting BRD4, are scarcely available. In-depth illustration of the protein quaternary structure, protein-to-protein interactions and structure dynamics might advance the design of improved inhibitors.In addition, clinical trials will provide feasible methods for oncologists to select the patients who are thought most likely to benefit from the treatment, which could be generally explained as the maximum of drug efficacy and the minimum of unfavorable clinical outcomes. Although YAP/TAZ are frequently activated in various cancers, the efficacy of YAP/TAZ-targeted treatment and the possible side effects may vary from patient to patient, which might require a personalized therapy for each individual. Specific biomarker tests  appear to represent the most common method to predict drug response, which also need to be investigated to anticipate the outcomes of YAP/TAZ-targeted therapy. And cohort studies are also valuable strategies to stratify patients depending on their shared characteristics. It should be pointed out, in contradiction to studies defining YAP as an oncogene, some studies also showed a tumor suppressive function for YAP. In breast cancer, loss of YAP protects cancer cells from anoikis, and YAP silencing promotes cancer cell migration and invasion . YAP knoOver the past years, although enormous efforts have been made to identify the complicated functions, regulations, and networks of the Hippo pathway, the underlying mechanisms how this signaling drives drug resistance remain largely obscure. YAP/TAZ hyperactivity or overexpression is apparently not the ultimate answer to this question. What are the downstream genes induced by YAP/TAZ and then more directly implicated in drug resistance? How does the Hippo pathway collaborate with other signaling to regulate YAP/TAZ activation in driving drug resistance? These mechanisms may clue more powerful multi- drug combinations in terms of further beneficial therapeutics for cancer patients. Moreover, stemness and microenvironment-directed drug resistance are two critical features of malignant cells, but relatively less investigated. What is the influence of YAP/TAZ-targeted treatment in cancer stem cell biology and tumor microenvironment? Deciphering these genetic, molecular, and cellular players related to YAP/TAZ biology will elicit more innovations in anti-cancer treatment and define more potentials of YAP/TAZ-target therapy.


Data relating to research design, sample population, type of electro-cosmetic devices used, were extracted.The search strategy identified 50 studies, 30 of which were potentially relevant.Our research is in favor of moderate periodical use of cosmetic medical devices in patients bearing tumors, in any stage, like in healthy people. Special consideration is dedicated to massage, manipulation, and pressure delivery upon the cytoskeleton of cancer cells that has proven to be sensitive to mechanical stress at least in some specific locally relapsing cancers such as osteosarcoma. Genetic and epigenetic events define the evolution from normal cells to the malignant state, up to death. Tumor progression is determined by molecular mechanisms, almost completely identified, acting progressively step by step until the end of the life. In 2000, Hanahan and Weinberg developed the first 6 hallmarks of cancer that became 10 in 2011 (The 10 cancer characteristics acquired by the cells during cancer development are:(1)Uncontrolled proliferation: healthy cells actively control the growth cycle and cell division ensuring homeostasis and maintenance of tissue architecture and function. Cancer cells counteract these signals resulting in uncontrolled cell growth (l growth .(2)Altered metabolism: the cancer cell metabolism is mainly based on aerobic glycolysis that breaks down glucose in aerobic conditions (nditions .(3)Apoptosis: the healthy cells avoid aging and DNA/RNA mutations/deletions, with physiological death and regeneration; the cancer cells ignore this pathway.(4)Replicative immortality: normal cellular aging is consistent with shortening of telomeres; cancer cells do not reduce their length.(5)Block of the onco-suppressor genes: onco-suppressors have been discovered by experimental activation–inactivation in mice or human cancer tissues. They code for proteins that can inhibit and/or downstage the proliferation (feration .(6)Genomic instability: gene mutations and chromosome fragmentation, cross over, and partial loss, increase into cancerized nuclei. Normal cells counteract partially these damages activating their endogenous DNA repair system.(7)Inflammation: cancer cells express very often inflammatory cytokines that help cancer growth.(8)Immune defenses: the check point of immune control of normal cell reproduction is lost in the majority of malignant tumors.(9)Metastasis: cancer cells can colonize far from their primary site through active cytoskeleton motility via adhesion receptors and coagulation–fibrinolytic interactions.(10)Angiogenesis: newly formed blood vessels arise into and around the cancer masses to supply oxygen and nutrients and output carbon dioxide and waste catabolites  with dominant autosomal forms,•Mismatch repair syndrome,•Family shapes.In natural conditions, the cells, after 50–60 reproductive cycles, undergo apoptosis, but in cancer cells, telomerase persists actively rendering them young and immortal. Exponential growth, accordingly, with the Gompertz’s law, is the result of continuous multiplication: later on, the cell turnover reaches a plateau, stabilizes, then slows down to balanced reproduction/versus death cycles. Viable cancer cells can rest quiescent for years either stopping the mitotic cycle or equally balancing proliferation and necrosis , 3.The growth, invasion, and formation of tumor metastases are induced by new vascularization, triggered by cancer cells through neo angiogenic cytokines . The newTumor angiogenesis is mediated by molecules or factors such as:•Fibroblastic growth factor (FGF),•Vascular endothelial growth factor (VEGF),•Vascular permeability factor (VPF),•Angiogenin,•Epidermal growth factor (EGF).The tumor necrosis factor alpha (TNFα) is primarily engaged in host’s defense against bacterial and viral infection. Involved both in physiological and pathological processes, it causes cancer cell necrosis and apoptosis. Finally, TNF has emerged as an important risk factor for cancer progression, invasion, and metastases and is a key intermediary of chronic inflammation associated with cancer . It bindTumor pathology is obviously multifactorial, with variable role of genes, environment, nutrition, lifestyle , 6. SeveIn the recent years, the use of complementary alternative medicine (CAM) has increased sharply, used as an adjuvant tool in oncological diseases and palliative care , 13. A vThe most common CAM massage techniques are:•Ayurvedic massage restores the balance between mind and body in such a way that it can give the individual a psychophysical well-being.•Chinese Tui Na massage relaxes muscles, improves bone system performance, and eliminates stress.•Do-in massage is one of the best methods to combat stress.•Massage with acupressure is effective especially in sleep disturbances.•Thai massage releases energy and is supposed to promote detoxification.•Shiatsu massage reduces stress and anxiety, promotes psychophysical relaxation, counteracts insomnia, and relieves ailments and joint pain.•Zonal massage aids detoxification and reduces joint pain.Aromatherapy (ATM), integrated with manual massage, is a very appealing approach in neuroaesthetics as assessed by Boehm et al. . His sysThe mechanisms leading to benefits and recovery in the daily life are summarized as follows:•Psychophysical relaxation; induced by the olfactory characteristics of aromatherapy.•Childhood regression; determined by manual contact of the massage.•Better perception of one’s own image.•Restoration of the mind-to-body ratio.in vivo model with MG-63 (human OS cells), turned to osteosarcomatous after GFP-lentivirus transfection, and then implanted, into the right and left joint . The massage was repeated on the tumor site twice a week, not in the control group (MT-). The mice were sacrificed when the tumor size reached roughly the 2% of the body weight, and at the autopsy, bilateral legs, liver, lungs, lymph nodes, and serum were kept for immunohistochemistry. The rabbit anti-GFP antibody  marked MG-63 cells in the organs of tumor-transferred mice. The mice treated with MT had reduced survival with the highest recurrence rate or metastasis, accordingly with the human osteosarcoma study. Babahosseini et al. , evokes deactivation of a limbic–paralimbic–neocortical network, up to the limbic system, as well as activation of somatosensory brain regions. The evidence-based studies show that the application of acupuncture reduces chemotherapy-induced nausea, leading to greater control of pain and anxiety. Last, but not least, the acupuncture is a highly safe and reliable integrative cancer therapy.A 2010 Hui ,•Electroporation,•Electromagnetic inductors,•High- and low-frequency magnetotherapy,•Heat-generating instruments such as radio frequency,•Tecar,•Electrotherapy,•Cryotherapy.These active principles, in our opinion, have no absolute contraindications in cancer because they release low energy across a very limited depth across (skin–dermis–subcutis) for the cosmetic restoration of epidermis collagen reticular and elastic fibers. Electromedical instruments that exploit vacuum therapy or activate venous, arterial, and lymphatic circulation (pressotherapy) are applied in places of the body where congestion and stasis are subsequently caused by wide lymph node excisions or dense collagen scars (like in the forearm post mastectomy lymphedema or “brass gross” or in heavily irradiated areas). Some experimental and clinical investigations focused on the cancer cells spreading risk through the endothelial gaps of the capillaries, resulting in metastatic progression. Although this hypothesis may be supposedly theoretically reasonable, the huge, complex network of receptors, adhesive systems, and coagulation fibrinolysis, and permeability factors exclude that low power energies have any direct independent chance to promote instant growth and dissemination of cancer. In the Craig’s  study, e•adhesion of cancer cells to matrix proteins;•stimulates the proliferation of cancer cells.In a murine model of Co26 and Co51 cancer cells implanted into a surgical wound, the burden of 15 mmHg pressure on tumor cells reduced the survival rate of the animals, in comparison with cells under normal pressure or pretreated with colchicine. Cancer lines from the colon and breast express the same behavior through adhesion to matrix proteins, endothelial cell monolayers, and experimental surgical wound margins with intrinsic signal variations in integrin-binding affinity.in situ to invasive carcinoma [for example, expression of E-cadherin calcium dependents is reduced in carcinomas  in different diseases. Herrero et al.  assessedRadial shock high-energy mechanical waves are generated by an electrode triggered in a chamber insulated by a rubber membrane in contact with the skin. They pass through the skin and spread radially into the body. Widely used to treat kidney stones, they are effective also in bone, joint, and tendon diseases, and even in cancer. The shock wave-spreading pathways are still unclear, but they produce mechanical cell rupture and increased cellular permeability with two basic mechanisms of action:(1)Direct energy delivery enhanced by the reflectivity amplification either on soft (muscles and tendons) or hard  tissues and clusters.(2)Indirect effect of “cavitation” on the cell membranes when the energy exceeds the elastic compliance of the tissue.Both these action mechanisms activate the microvascular network and reduce the local inflammation, stimulating also osteogenesis in porotic or fractured bone. Two cell lines were experimentally treated with SW, one cancerized (CAKI-2) and the other normal (HK-2 and HRE): shock waves were highly detrimental for tumor cells rather than normal ones .Ultrasound is a vibratory acoustic wave. Ultrasonic rays focus upon one target and spread harmlessly through the surrounding tissues. These are selectively absorbed accordingly with the emission frequency and tissue and organ density and also histomorphological structure. Temperature increases in the target areas, especially with high-intensity focused ultrasound (HIFU). HIFU have been experimentally used to treat pancreatic cancer , with encouraging results in terms of surrogate endpoint  administration in combination with systemic chemotherapy, reducing the abdominal pain offering survival benefits and a few low-grade complications . In his Even if further specific systematic investigation is lacking, considering that HIFU is successfully endorsed to locally destroy cancer cells in the clinical setting, we suppose that indication of the same energy for cosmetic purposes is safe. As to low power ultrasounds, their safety is testified by the diagnostic use in obstetrics, gynecology, and with higher acoustic intensity, for remodeling subcutaneous fat .Laser and pulsed light (IPL) systems, widely used in aesthetics and dermatology, are considered fairly safe. However, the literature reports some possible side effects .•Edema,•Flittena or blisters from burn,•Hypopigmentation,•Hyperpigmentation,•Scarring.All laser and IPL systems use non-ionizing radiation that do not interfere with DNA. Studies denied any cause–effect relationship between laser aesthetic use and cancers . AesthetRadiofrequency is used in Wi-Fi systems, radio, television, and microwave ovens. The electromedical and cosmetical devices work with energies in the range of 400 kHz, with the following different delivery systems:•Monopolar radiofrequency: it is made of two handles that generate opposite negative and positive energy, overheating the trespassed tissues down to the deep layers of the dermis and hypoderma. The body and the face are the usual targets of these instruments.•Bipolar resistive radiofrequency: it delivers electric charges through two electrodes combined in the same handle. A higher localized thermal concentration is, thus, achieved.•Phototherapy or photoporation: the absorbed electromagnetic energy induces different effects depending on the wavelength and can help low-molecular weight cosmeceuticals to pass across the skin barrier.•The capacitive radiofrequency: works deep into the tissues through the placement of insulated electrodes, exciting the interstitial water molecules with heating effect.•The resisting radiofrequency: heats the tissues by electrodes; when delivered through the fat, greater thermal effect induces lipolysis.Generally speaking, the cosmetic final goal of the radiofrequencies is to reach the deep dermis and subacute, strengthening its texture, thickness, and elasticity. We did not find any, even if anecdotal, literature contribution about aesthetic radiofrequency administration to cancer patients. Our personal experience is that they effectively tighten the skin in the face, arms, and legs inducing a very good, even if temporary, rejuvenation effect; no untoward effects have been observed, with the warning of achieving an adequate hydration level of the skin before radiofrequency heat delivery. In the experimental setting, a 2-year study  exposed Cryotherapy is widely used for cosmetic purposes in the following methods:•Localized cryotherapy for single or multiple benign lesions. The sudden contact of the skin with low temperatures is painless and concentrated on single skin spots: it removes benign and malignant skin tumors and smoothens the skin surface improving translucency.•Cryosauna is a vertical cabin in which liquid nitrogen is released in a gaseous state (-140°C for about 3 min). The patients stay for a few seconds into it, for a certain number of sessions, just the time to activate subcutaneous fat cell lipolysis and mesenchymal reticular and collagenous fiber turnover.•Cryotherapy associated with electroporation and chromotherapy is addressed to localized areas of the body. The synchronous effects of electroporation currents or chromotherapy, with the sudden cooling of the target, enhance the mesenchymal cells turnover and reduce fat and cellulitis.Cryotherapy is a historical weapon for the local treatment of cancers either of the skin or for the deep organs such as the liver, kidneys, etc. Freezing methods are safe: Zang  confirme•MRC5 fibroblasts are extremely sensitive to hyperthermia, but more resistant to hypothermia.•T98G and A549 are thermo-tolerant and thermosensitive to varying degrees.•In the U87MG thermal-sensitive cellular line, hyperthermia triggered caspase 9.•In the thermo-tolerant T98G and A549 cell lines, caspase 9 levels decreased.•Hyperthermia strongly induced HSP90 levels in T98G, while a sharp decrease was recorded in the PC3 and U87MG thermosensitive cell lines.•Hyperthermia sensitized the cell lines of thermo-sensitive cancer to cisplatin and temozolomide, while its sensitizing effect decreased in thermo-tolerant cell lines.On the basis of this investigation, we conclude that either moderate hyperthermia, or namely, hypothermia, reduces the vitality and/proliferation in all the above-described cell lines.in vitro/in vivo dedicated studies or anecdotal clinical reports, or any other reasonable statement. The cancer cells are very vulnerable to almost all the physical existing energies due to their simplified antioxidant, defense, or DNA repair intrinsic potential. Most of the energies delivered by aesthetic electromedical instruments have been, or are currently, used with higher power to locally destroy cancer cells. The genetic and molecular mechanisms modulating cancer turnover investigated, so far, either in the induction or spreading or terminal phase, interact so finely and sequentially that any putative epigenetic interaction generated by this class of instruments seems gross and unrealistic. Our conclusions are in favor of moderate periodical use of cosmetic medical devices in patients bearing tumors, in any stage, like in healthy people.Our report, based on the literature screening and our personal long-standing clinical experience, was intended to verify whether the warning against the use of electromedical instruments in the cosmetic professional or medical cancer patient settings is consistent with evident oncological risks supported by experimental Special consideration is dedicated to massage, manipulation, and pressure delivery upon the cytoskeleton of cancer cells that has proven to be sensitive to mechanical stress at least in some specific locally relapsing cancers such as osteosarcoma. Cancer mechanotransduction, a new investigational oncological profile, gives us some warnings about the due care when treating manually or with mechanical patients affected by malignant tumors, and let us hope in further new, instrumental, and molecular strategies to cope with cancer diffusion and metastasis. Obviously, during and after the aesthetic sessions, a medical judgment about the general individual trophic and local conditions of the skin and teguments, especially during chemo and oncological therapies, has to be carefully considered and adequately managed. Our strongly convinced opinion is that onco aesthetic care is, today, an extraordinary aid along the obscure tunnel of impending cancer diagnosis, treatments, and follow-up, giving a smart psychological and self-esteem support to the patients. It will be extremely helpful to avoid perceptions of discrimination and adequately cope with adverse destiny.BP and MV: results and conclusions. LP and AM: materials and methods. VP: introduction. All authors contributed to the article and approved the submitted version.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.


The complex physiological signal transduction networks that respond to the dual challenges of inflammatory and oxidative stress are major factors that promote the development of cardiovascular pathologies. These signaling networks contribute to the development of age-related diseases, suggesting crosstalk between the development of aging and cardiovascular disease. Inhibition and/or attenuation of these signaling networks also delays the onset of disease. Therefore, a concept of targeting the signaling networks that are involved in inflammation and oxidative stress may represent a novel treatment paradigm for many types of heart disease. In this review, we discuss the molecular mechanisms associated with the physiological responses to inflammation and oxidative stress especially in heart failure with preserved ejection fraction and emphasize the nature of the crosstalk of these signaling processes as well as possible therapeutic implications for cardiovascular medicine. Heart failure (HF) is increasing in prevalence and now affects over 10 % of those aged 70 years and over. HF is characterized by the activation of the sympathetic nervous and renin-angiotensin-aldosterone systems, in a process of neuroendocrine activation that is associated with oxidative stress in the myocardium and vasculature. Oxidative stress occurs in the myocardium  is characterized by high myocardial diastolic stiffness. Diastolic dysfunction is defined as the inability to fill the ventricle to an adequate preload volume  at acceptably low pressures. Patients with abnormal diastolic function in particular HF with preserved ejection fraction (HFpEF) showed a complex remodeling of cardiomyocyte structure and function, in addition to a remodeling of the non-myocyte compartment  induce cellular changes that are at least partially mediated via oxidative or nitrosative stress , IL-6, monocyte chemoattractant protein 1, and other chemokine ligands, all of which lead to monocyte recruitment and macrophage activation. Increased peripheral inflammation, monocytosis, and monocyte differentiation to anti-inflammatory/profibrotic M2 macrophages have been associated with HFpEF in a population with a very high prevalence of metabolic comorbidities , c-Jun N-terminal kinases, and p38 MAPKs in the myocardium  in various cell compartments, including the cytoplasm, cell membrane, endoplasmic reticulum (ER), mitochondria, and peroxisome, all as through basal metabolic activity. In cardiomyocytes, ROS typically originate from several intracellular sources, including mitochondria, NOS, and enzymes such as xanthine oxidase, NADPH oxidase (NOX), and cytochrome p450 . Changes in intracellular calcium cycling are either causal or adaptive  , sarco-/endoplasmic reticulum Ca2+-ATPase (SERCA), ryanodine receptors (RyR), and myofilament proteins such as cardiac troponin I (cTnI), myosin-binding protein C (cMyBPC), and the giant protein titin –mediated phosphorylation targets several proteins involved in calcium handling, including the –regulated kinases (ERK), and calcium calmodulin–dependent protein kinase II (CaMKII)  Fig. .The titin protein consists of two main isoforms, N2BA (long and compliant) and N2B (short and stiff), and acts as a molecular spring with a crucial role in LV diastolic stiffness –directed PKG signaling is initiated by soluble guanylate cyclase (sGC) or particulate GC (pGC), both lyase enzymes that convert guanosine triphosphate (GTP) to cyclic guanosine monophosphate. sGC is activated by NO, which then generates cGMP from GTP Fig.  Klaiber; b. pGC Modifications of cardiomyocyte function in HF are associated with alterations in cGMP-PKG signaling due to oxidative stress (indirect ROS effect on cardiomyocyte modifications) Fig. . NormallOxidative stress is a major stimulator of epithelial cell function and can induce DNA damage and repair. During DNA repair, cells are prevented from entering S phase of the cell cycle  phosphorylates several cardiac calcium handling and myofilament proteins to modulate excitation–contraction coupling, apoptosis, and gene transcription  is a conserved enzyme that controls glucose flux through the canonical Embden-Meyerhof glycolytic pathway . It also mediates cell death via nuclear translocation under conditions of oxidative stress. GAPDH is a glycolytic enzyme that is responsible for the sixth step of glycolysis   1 and 2 cascades are markedly activated in cardiomyocytes by virtually all hypertrophic stimuli, inhibition of the cascade suppresses at least some aspects of the hypertrophic response, and constitutive activation of the pathway produces compensated cardiac hypertrophy in transgenic mice . Many studies showed suppression of several signaling pathways and in particular the cGMP-PKG pathway in HFpEF patients and animal models of HFpEF. This reduction is possibly due to inflammation and oxidative stress making it a suggestive treatment target option for these patients. However, as many heart diseases are characterized by increased inflammation and oxidative stress, one may think that targeting both could be a treatment option for many sub-groups. Indeed, our recent work suggested that increased oxidative stress and inflammation may play a major role in the deterioration of LV diastolic function, as we showed that NO-sGC-cGMP-PKG signaling is reduced in HFpEF patients and in an HFpEF animal model (Kolijn et al. l-arginine attenuate the expression of many pro-inflammatory cytokines (De Caterina et al. The current treatment options for many forms of heart disease in particular for HFpEF patients are very limited and no drug has yet been shown to improve cardiac or specifically diastolic function in these patients. Many of the tested compounds were ineffective in reducing morbidity and mortality although in many cases proven to be effective for the treatment of HFrEF. Accordingly, several clinical trials have failed to show that any of these proposed drugs have positive effects in HFpEF patients. Patient management is presently limited to amelioration of symptoms and the treatment of common comorbidities such as hypertension, diabetes, obesity, and atrial fibrillation. Some trials such as the recent PARAGON-HF trial that investigated the effects of sacubitril/valsartan in patients with HFrEF showed evidence of a very heterogeneous response to treatment (Solomon et al. It remains very critical to understand the complex interactions of oxidative and nitrosative stress with pro-inflammatory mechanisms, metabolic dysfunction, signaling pathways, and the redox modification of proteins characteristic of heart failure to design novel approaches to therapeutic strategies for each heart failure phenotype.