Cutaneous polyarteritis nodosa (cPAN) is a rare, necrotizing vasculitis involving the small- and medium-sized vessels of the dermis and subcutaneous tissues. We report a severe case of cPAN in a patient with an atypical presentation of extensive bilateral lower extremity ulcerations with full-thickness necrosis managed at a burn center. A 70-year-old female with a three-month history of necrotizing cPAN to the bilateral lower extremities underwent surgical excision and autografting at an outside hospital. Postoperatively, she had total graft loss and was begun on prednisone. In the outpatient setting, she was tapered off prednisone and subsequently began to experience an acceleration of the disease process. She was then transferred to our regional burn center for bilateral escharotomy and management of her non-healing, tender, necrotic wounds with distinctive black-brown eschar. One year later, the patient’s wounds continue to decrease in size and heal with her daily regimen of 15 mg of prednisone, 50 mg of cyclophosphamide, and topical silver sulfadiazine application. With the increasing volume of non-burn wound admissions to burn centers primarily of dermatologic etiology, it becomes crucial for burn specialists to familiarize themselves with severe presentations of vasculitides, including cPAN. Further research is necessary to understand the atypical manifestations of this disease for more timely diagnosis and treatment. Cutaneous polyarteritis nodosa (cPAN) is a rare, necrotizing vasculitis that involves the small- and medium-sized vessels of the dermis and subcutaneous tissues. The classic clinical presentation of this disease is small 0.5-2 cm) subcutaneous nodules (80% of cases in a study of 79 patients), livedo reticularis (56%), and skin ulcers 49%) . Other r cm subcu9% . OtheAlthough dermatologists typically manage patients with cPAN, our experience with this extreme, gangrenous case of cPAN suggests that a multidisciplinary team approach that includes both dermatologists and burn specialists, such as one readily available in burn centers, may be beneficial for patient outcomes and recovery in such severe cases. With the increasing volume of non-burn wound admissions to burn centers primarily due to dermatologic and rheumatologic cases, it becomes crucial for burn specialists to familiarize themselves with severe presentations of vasculitides . In factA 70-year-old female presented to an outside hospital with a chief concern of ulcerations of unclear etiology to the bilateral lower extremities for three months. These ulcerations initially began as a small area of erythema on the right ankle that progressed to a blister and subsequent skin erosions. She underwent surgical excision and autografting at this hospital to address these concerns. Postoperatively, she had total graft loss and was begun on prednisone. In the outpatient setting, she was tapered off of her prednisone regimen and subsequently began to experience an acceleration of the disease process. She was then transferred to our regional burn center Intensive Care Unit for management of her non-healing wounds. Upon arrival at our burn center, she required bilateral escharotomy for her necrotic wounds. Her physical examination revealed tender, gangrenous wounds with a distinctive black-brown eschar Figures , 1b.The lower wounds were biopsied, revealing necrotizing vasculitis involving all layers of the vascular walls of the small and medium-sized arteries, exuberant fibrinoid degeneration of the vessel walls, dense perivascular and interstitial neutrophilic inflammation with neutrophilic debris and extravagated erythrocytes, and edema with an overlying necrotic epidermis Figures , 2b.Her past medical history was significant for hypertension, high cholesterol, papillary thyroid cancer status post total thyroidectomy and thymoma status post resection two months before the development of the wounds to her bilateral lower extremities. Serum protein electrophoresis was consistent with hypogammaglobulinemia and acute inflammation, and her IgG levels were markedly low . Her antinuclear antibody (ANA) titers were elevated , but perinuclear anti-neutrophil cytoplasmic antibodies (pANCA), cytoplasmic antineutrophil antibodies (cANCA), rheumatoid factor, anti-Ro, myeloperoxidase (MPO), C3 and C4, anti-cardiolipins, and serologies for human immunodeficiency virus, herpes simplex virus, and hepatitis B and C were all within normal limits or negative. The patient did not have any recent viral infections, and testing for Mycobacterium tuberculosis, staphylococcal and streptococcal infections were all negative.This patient was hospitalized for a total of 19 days at our burn center. Today, she continues to follow-up with our burn center to evaluate her response to her daily regimen of 15 mg of prednisone, 50 mg of cyclophosphamide, and topical silver sulfadiazine application. The patient cannot tolerate a dose lower than 10 mg of prednisone daily due to the exacerbation of her painful wounds. One year later, however, her wounds have significantly decreased in size and continue to heal but have moderate drainage with no signs of infection or cellulitis.cPAN was first described in 1931 as a distinct version of polyarteritis nodosa (PAN) that presented with similar clinical and histological cutaneous findings but lacked systemic manifestations . It is cThe clinical signs and symptoms of cPAN predominantly include subcutaneous nodules and livedo affecting the legs, with females being affected more commonly than males . While uHistopathology of cPAN demonstrates a fibrinoid, necrotizing vasculitis of the small- and medium-sized vessels . LaboratFrom an immunologic perspective, the patient presented following a thymectomy due to thymoma. There is a well-established relationship between thymoma and autoimmune disease, though the exact mechanism of autoimmunity is still debated . Some stDespite generally favorable prognosis in patients with cPAN, close follow-up is necessary to monitor recurrence as well as potential progression to systemic disease. The majority of the current literature suggests that cPAN does not progress to PAN, and there is only one case report to date that identifies two patients who solely had cutaneous manifestations for over a decade before later progressing to systemic PAN ,7,14. PaThis patient’s care ultimately depended on a multidisciplinary approach from her management team. Throughout her stay at our hospital, she received repetitive care from dermatologists, rheumatologists, physical therapists, social workers, and dieticians in addition to burn specialists. Of note, the dermatology team was crucial in diagnosing this patient with cPAN through dermatopathological evaluation and in providing pertinent management recommendations, such as transitioning to chronic immunosuppression and assessing whether repeat biopsies were necessary when lesions worsened. Rheumatology further assisted in the diagnostic process given the complexity of this patient’s presentation and helped determine which immunosuppressants would best benefit this patient.This report describes an extreme case of a patient with cPAN who ultimately required treatment at a burn center to manage her non-healing, ulcerative wounds that have substantially improved to date with a combination of prednisone, cyclophosphamide, and topical silver sulfadiazine. The patient’s case illustrates one of the many ways in which this rare and poorly understood vasculitis may manifest and provides an example of disease management. Isolated gangrenous necrosis with no systemic symptoms, while an uncommon manifestation of cPAN, should lead practitioners to include this disease on the differential and to even consider treatment in a burn center where patient care can be specialized and multifaceted.Conditions treated at burn centers are often stigmatizing given their visually noticeable manifestations that may negatively impact self-esteem, leaving patients to carry a major psychosocial burden both during their hospital stay as well as upon discharge. The multidisciplinary approach to patient care in modern-day burn centers is thus well-equipped to address these specific patient needs given the variety of training backgrounds from team members. Between intensive wound care, infection control, physical and occupational therapy, social work, and immediate availability for surgical care, burn centers have evolved to handle a variety of wounds of non-burn etiology, including severe vasculitides. While cPAN has historically been managed by dermatologic teams, we recommend a multidisciplinary approach to this disease that includes burn specialists, particularly in such extreme cases as this patient’s presentation that ultimately resulted in surgical interventions and aggressive pharmacologic therapy.


End stage heart failure is a major cause of death in the US. At present, organ transplant and left-ventricular assist devices remain the only viable treatments for these patients. Cardiac tissue engineering presents the possibility of a new option. Nanomaterials such as gold nanorods (AuNRs) and carbon nanotubes (CNTs) present unique properties that are beneficial for cardiac tissue engineering approaches. In particular, these nanomaterials can modulate electrical conductivity, hardness, and roughness of bulk materials to improve tissue functionality. Moreover, they can deliver bioactive cargo to affect cell phenotypes. This review covers recent advances in the use of nanomaterials for cardiac tissue engineering. Cardiovascular disease is a leading cause of morbidity and mortality across the United States and the world resulting in 800,000 and 17,000,000 annual deaths, respectively . The diaThere is a complex pathophysiology between myocardial infarction and heart failure. Although myocardial ischemia due to AHD or overt MI is the most common etiology of HF, it is important to note that there are other causes of HF. Ischemic insults result in gradual loss of functional myocardium and reductions in cardiac output. MI occurs when an atheromatous plaque ruptures within a coronary vessel, leading to occlusion of the vessel and loss of perfusion of the distal myocardium. If not intervened upon, the area of ischemic myocardium will infarct and progress through a series of well-defined healing stages . This prAdvanced heart failure refers to patients with persistent HF symptoms that significantly inhibit their daily lives, despite maximal medical therapy. Of the 6 million HF patients in the US, approximately 200,000 meet the criteria for having advanced heart failure . PatientTechniques in cardiac transplant have evolved such that transplant recipients now have an expected mean post-transplant life expectancy of 12–13 years . MortaliThe field of cardiac tissue engineering seeks to give patients with evidence of myocardial injury an additional therapy to potentially regenerate innate myocardial function. Nanomaterials, such as gold nanoparticles and carbon nanotubes, have unique electrical, mechanical, and thermal properties, which have positive therapeutic effects on electroactive cells, such as cardiomyocytes, which motivates the development of cardiac tissue engineering approaches. Moreover, they can serve as delivery systems for active biomolecules and/or drugs to further improve cardiac regeneration. This review covers recent advances in the use of nanomaterials for cardiac tissue engineering.Defined as materials with a characteristic dimension of 1–100 nm, nanoparticles have been increasingly investigated for medical applications, both as therapeutic and diagnostic tools ,24. TheiCurrently, most existing applications of nanoparticles in medicine rely upon intravenous injection of nanoparticles primarily for use in drug delivery or imaging. However, despite decades of work focused on delivery of nanoparticles to their target sites in vivo, the majority of intravenously-delivered nanoparticle doses do not actually reach target cellular niche. Chan and coworkers conducted a rigorous review of recent nanoparticle literature, finding that, on average, only 0.7% of nanoparticles targeted toward solid tumors reached their destination . NanoparThe incorporation of nanoparticles into tissue engineering has been developing more recently ,39. WithIn the upcoming sections of this review paper, we discuss the specific studies evaluating the incorporation of nanomaterials into cardiac tissue engineering approaches. We place special emphasis on carbon nanostructures and noble metal nanoparticles, which have been studied most extensively. In particular, carbon nanotubes and gold nanorods have spurred the most research efforts, and are discussed in the most detail.Carbon nanomaterials are biomaterials composed predominantly of carbon atoms with one dimension less than 100 nm. These nanomaterials fall into three categories: graphene, carbon nanotubes (CNTs), and fullerene, all of which have been discovered within the last four decades . Graphen2 hybridized carbon atoms, in contrast with bulk carbon materials such as diamond and graphite, which contain sp3 hybridized carbon atoms [2 geometry leaves open one p orbital per carbon atom, perpendicular with the plane of carbon atoms in these structures. These p orbitals can combine to form planar π molecular orbitals, which enable the sharing of electrons between many carbon atoms, resulting in the electrical conductivity (or semi-conductivity) of CNTs and graphene nanostructures. Additionally, these π orbitals permit the modification of carbon nanomaterials via π-π stacking with hydrophobic cargo. These materials also have high mechanical strength and stiffness. Furthermore, CNTs and graphene are anisotropic nanostructures that promote alignment of electrical cells, such as cardiomyocytes, preferentially in one direction. This increase in contractility of cells along the long axis mimics cardiac tissue. The conductivity, anisotropy, and mechanical strength of carbon nanostructures have been explored for therapeutic approaches in cardiomyocytes.As discussed in the excellent review paper by Hong et al., these interesting properties of carbon nanomaterials are due to their unique chemical structures . Specifion atoms . The hyb2+ ATPase (SERCA) and connexin 43 (Cx43), associated with gap junctions, were both also increased in the NRVMs grown on CNTs. Sun et al. identified additional components of signaling involved in cell-CNT interactions, finding that CNTs embedded in a gelatin methacrylate anhydride gel induce β1-integrin-mediated FAK  and RhoA (a cytosolic GTPase protein) upregulation in electrical and mechanical junction proteins [CNTs induce cardiomyocytes to function in a way that improves cardiomyocyte activity for cardiac tissue applications. Multiple studies have been conducted via electron microscopy and biochemical techniques in order to evaluate cardiomyocyte phenotype changes, due to the use of CNTs. Our laboratory’s work by Martinelli et al. in 2012 was the first to demonstrate the therapeutic effects of CNTs on electrophysiological behavior of cardiomyocytes . Our labproteins . These sAnisotropy has been harnessed in the field of cardiac tissue engineering as a whole, in order to improve biological outcomes, and CNTs provide an opportunity to impart anisotropy to materials. For example, 3D printed materials have been built with anisotropic properties that have guided the formation of endothelialized myocardium in a “heart on a chip,” which is capable of contracting . In thisThe use of electrical and mechanical properties of CNTs to influence stem cell differentiation and cardiomyocyte phenotypes in hydrogels has also been investigated. Ahadian et al. used a gelatin methacryloyl gel with aligned CNTs to differentiate embryoid bodies into cardiomyocytes. They achieved this by controlling the frequency, amplitude, and duration of an electrical stimulation through the gel . This apMechanical properties of CNTs have been investigated rigorously as cues for cardiomyocyte phenotype in other work. Yu et al. were able to show improvement in NRVM cell function by incorporating CNTs into type I collagen hydrogels . In the The influence of conductivity on atrial versus ventricular cardiomyocyte phenotype has also been investigated. Lee et al. performed a mechanistic study using GelMA scaffolds containing various CNTs, graphene oxide (GO), and reduced graphene oxide to evaluate the effects of conductivity on NRVM cell phenotypes on these scaffolds . This grThe method of preparing CNT-containing materials also plays a critical role in outcome. For example, Liu et al. created poly(ethylene glycol)-poly copolymer (PELA) fiber scaffolds using electrospinning, finding that the method used to incorporate the CNTs into the polymer had an effect on NRVM cell viability and function. If the CNTs were incorporated into the scaffolds coaxially within inner portions of the PELA fibers, the cells showed greater elongation and increased α-actinin and troponin I expression, versus materials with CNTs blended throughout the PELA, even though the PELA fibers with CNTs blended into them had greater conductivity and Young’s moduli. This study also found that cardiomyocytes on a coaxial scaffold with 5% CNT showed the best performance in terms of cell maturation, and this scaffold had lower toxicity .Electrospinning has been explored further by other investigators, who have taken advantage of the anisotropy of electrospun materials. Kharaziha et al. were able to incorporate CNTs into an aligned poly(glycerol sebacate):gelatin gel. They found that this scaffold had greater electrical conductivity and toughness compared to scaffolds without CNT. NRVM cells grown on this scaffold had more synchronous beating behavior and maintained cell viability . Wu et aSeveral other investigators have constructed CNT-containing hydrogels that have achieved favorable outcomes in vitro and in vivo. Pok et al. showed that a chitosan hydrogel with CNT can support beating NRVM cells at almost natural beating rates. The cells on the hydrogel also showed synchronous beating due to the improved conductivity of the gel . In anotGrowth factors can be delivered by modification of hydrogels containing carbon nanomaterials in order to optimize their properties. Paul et al. constructed GelMA hydrogels containing graphene oxide as a means of delivering plasmid DNA of VEGF to transfect adjacent endothelium and stimulate angiogenesis in peri-infarct regions of rat models . This stFullerene-derived carbon nanomaterials have also been studied for cardiac tissue engineering. The fullerene derivative fullerenol has been incorporated into injectable alginate hydrogels to improve the survival and proliferation of brown adipose derived stem cells and promotes their differentiation into cardiomyocytes . Hao et Stem cell encapsulation within CNT-containing gels has been explored to improve integration with heart tissue. Li et al. modified a poly (N-isopropylacrylamide) with CNT . The gelThe route of administration of CNT-containing materials has been considered in some studies, as non-invasive methods may be preferable for future clinical applications. Our research group has also developed a unique injectable formulation of CNTs chemically incorporated into a reversible thermal gel (RTG) that is liquid at room temperature and gels at body temperature . This woUnique from the aforementioned studies, CNTs have also been evaluated in vivo using models other than myocardial infarction models. In one recent study, McCauley et al. ablated the atrioventricular nodes of sheep models, and CNT fibers were sutured across the scars created by this process . AlthougIt should be noted that some studies involving CNTs showed that there is significant toxicity associated with their use in vivo, as reviewed by Liu et al. . InitialOverall, carbon nanomaterials have been investigated in great depth for cardiac tissue engineering, although the mechanism by which tissue engineering is achieved remains unclear and is perhaps multifactorial. Mechanotransduction, conductivity, anisotropy, and mechanical properties, such as Young’s modulus, are among the possible factors at play. Regardless, outcomes for cardiac tissue repair for carbon nanomaterial-containing scaffolds have been favorable in both in vitro and in vivo studies.Gold nanoparticles (AuNPs), gold nanorods (AuNRs), and silver nanoparticles have been studied extensively for biomedical applications ,27,28. VMixing AuNRs into hydrogels enhances cardiomyocyte differentiation into functional tissue. One method of incorporating AuNRs into hydrogels for tissue engineering is via mixing of AuNRs with UV-crosslinkable of gelatin-methacrylate (GelMA) prepolymers . CompareNavaei et al. have since performed a fascinating study to evaluate what property of the AuNRs in GelMA hydrogels is most beneficial for biological outcomes . In thisAuNRs have also been incorporated into collagen scaffolds . These cSpherical AuNPs have also been studied for cardiac tissue engineering. Baei et al. evaluated the effect of AuNPs added to chitosan thermosensitive hydrogels, which were seeded with mesenchymal stem cells (MSCs) . AuNPs iNair et al. integrated AuNPs into a porcine gall bladder-derived extracellular matrix to assess effect on H9C2 cardiomyoblast growth in vitro . These rModification of Au and AgNPs has also been performed, in order to influence tissue scaffold architecture. For example, Alarcon and coworkers experimented with the incorporation of peptide-coated AuNPs and AgNPs into injectable collagen-containing hydrogels to evaluate their effect on NRVM growth and macrophage polarization . The autOur group has recently investigated reverse thermal gels (RTGs) containing AuNPs for in vitro growth of NRVM cells . RTGs arPolymer nanoparticles have been investigated as well, though in significantly less depth than the above carbon nanomaterials or noble metal nanomaterials. One advantage of polymer nanoparticles is that they can be readily modified with drugs and biomolecules by a broad range of chemical techniques. These systems also tend to be more degradable, theoretically resulting in reduced toxicity. However, they do not have the same level of conductivity or hardness as the previously discussed nanomaterials.Mytulis edulis. This sea creature must withstand high forces to adhere to rocks, relying on adhesive foot proteins that are composed of high quantities of the catechol-containing amino acid 3,4-dihydroxy-l-phenylalanine (DOPA) [2 and amine-terminated silica surfaces [There has been a particular emphasis on the use of nanomaterials that use mussel-inspired dopamine or polydopamine chemistry. Developed by Messersmith et al., these chemical strategies draw inspiration from the use of the marine mussel e (DOPA) ,83. The surfaces . Based usurfaces . The versurfaces . Amines surfaces . Metals surfaces , PEG [85surfaces , and RGDsurfaces . PDA hassurfaces ,93,94,95Wang et al. designed a self-assembled nanoparticle composed of a bioactive drug (tanshinone IIA), coated this nanoparticle with PDA, and attached it to an ROS-polymer hydrogel via PDA-thiol bonds . The resConductive polypyrrole nanoparticles were linked using mussel-inspired chemistry to generate a cryogel with GelMA . When imIn another approach, carboxymethylcellulose nanoparticles were used by Kim et al. to induce differentiation of mature normal human dermal fibroblasts into induced cardiomyocyte cells by delivery of the drug 5-AZA to stop proliferation and plasmid DNA  to induce cardiogenesis . This meMagnetic nanoparticles composed of iron oxide have briefly been investigated as well. These systems have been particularly helpful in manipulation of cells or targeting applications. In one such study, magnetic ferumoxytol nanoparticles were used to label cardiosphere-derived stem cells and to target them into coronary arteries of rat post-myocardial infarction ischemia/reperfusion models, thereby addressing the ordinarily low level of stem cell retention and engraftment . In anotIron oxide nanoparticles have been found to have beneficial properties on cardiomyocyte growth as well. Supraparamagnetic iron oxide nanoparticles (SPIONs) have been incorporated into silk-fibroin nanofibers to promote proliferation of mouse embryonic cardiac cells . Like otVarious other nanoparticles have also been used in cardiac tissue engineering. Silicon NPs modified with atrial natriuretic peptide (ANP) have been used to target cardiomyocytes both in vitro and in vivo . In cellMotivated by the presence of lower selenium concentrations in patients with cardiomyopathy, Kalishwaralal et al. designed chitosan-selenium nanoparticles for cardiac patches with enhanced electrical conductivity and mechanical strength . This fiLiu et al. developed another chitosan-based cardiac patch formulation composed of TiO2 NPs in PEG-chitosan hydrogels . Young’sMicroRNAs (miRNAs) are single-stranded noncoding RNA strands, usually 20–25 nucleotides in length, which are capable of downregulating target messenger RNA (mRNA) through RNA interference ,110. StuMultiple miRNAs have been shown to be important in cardiac disease and cardiac repair. A study utilizing high-throughput screening to identify miRNAs involved in cardiomyocyte proliferation in rats showed that miR-590 and miR-199a promote ex vivo cardiomyocyte proliferation. Following myocardial infarction in mice, administration of these miRNAs stimulated cardiac repair with near complete recovery of cardiac function . A pig mThe main challenges of delivering miRNA therapy are that free RNA shows poor cellular uptake, short half-life, and significant off-target effects. These challenges make the delivery of free RNA for therapeutic purposes impractical. There have been many attempts to modify miRNAs to increase their stability and cellular uptake; however, off-target effects and the formation of toxic byproducts by degradation remain problematic. Alternative strategies such as the use of lipid formulations for the delivery of miRNAs has shown promise, but traditional transfection agents can have significant toxicity . NanoparThere has been a tremendous recent effort in the development of nanoparticle-containing materials for cardiac tissue engineering, much of which now focuses on the translation of these materials toward clinical solutions. The nanoparticles that have been most studied to date are carbon nanotubes, gold nanoparticles, and gold nanorods. These materials have successfully directed greater proliferation of cardiomyocytes and enhanced cardiomyocyte function. More recently, there have been efforts to deliver bioactive compounds via nanoparticles within these scaffolds. Other innovative approaches including magnetic in vivo manipulation of cardiomyocytes using iron oxide nanoparticles have also been attempted. Even though clinical data regarding the use of these nanomaterials is still lacking, there has been a recent emphasis on translational research that has involved the use in vivo models. These promising translational studies motivate clinical evaluation of these materials in the near future.As the nanomaterials community moves toward consideration of these materials in clinical research, there are several additional directions for future research in vitro and in vivo. First, greater mechanistic insights will be helpful in optimizing material design for cardiac regeneration. The addition of nanomaterials into bulk materials often change more than one property alone, including mechanical properties, electrical conductivity, characteristics of contact with cells, and roughness. However, relatively few efforts have been made to determine which properties are most relevant. Moreover, the biological mechanism by which cell phenotype is altered by nanoparticles alter cell phenotype must be further studied. Second, additional work will need to be done to assess the long-term safety and stability of these materials in vivo. Third, the delivery of biologically active molecules from nanoparticles for cardiac tissue engineering, such as miRNA, presents great promise to promote healing and merits deeper exploration. Building upon current nanoparticle research efforts in cardiac tissue engineering into these directions would be exciting for the treatment of ischemic heart disease and advanced heart failure, potentially representing an alternative to left ventricular assist devices and heart transplant.


Identifying the functional materials inside vaping devices can help inform efforts to understand risk. While nicotine E-cigarette components and metals have been characterized in several previous studies, the internal component compositions of tetrahydrocannabinol (THC) cartridge designs are not as well known. The 2019–20 e-cigarette or vaping product use associated lung injury (EVALI) outbreak has been associated with THC devices containing vitamin E acetate (VEA), possibly mediated by chemical reactions with internal cartridge components and high filament temperatures.We investigate the composition and internal components of 2019 EVALI patient-associated THC vaping devices compared to other THC and nicotine devices from 2016–19, specifically the metal, ceramic, and polymer components likely to be exposed to heat. To do this, we have disassembled forty-eight components from eight used and unused vaping devices under a microscope and analyzed them using X-ray fluorescence, scanning electron microscopy, and Fourier-transform infrared micro-spectroscopy.The two THC cartridges used by EVALI patients exhibited evidence of localized high temperatures, including charring of the ceramic heating elements and damaged wire surfaces. The newer THC cartridges possessed more ceramic and polymer insulation than older THC or nicotine devices. The combination of ceramics, metals, and high temperatures in newer THC cartridges is consistent with conditions hypothesized to produce VEA reactions during vaping. Nickel and chromium components were detected in all devices, and others contained copper, lead, tin, gold, silicon-rich rubbers, or fluorinated microplastics. These components have the potential to thermally degrade and volatilize if heated sufficiently. These findings do not imply that harmful exposures would occur under all usage conditions, and are most relevant to harm reduction efforts based on avoiding higher internal temperatures. This study was limited to a small sample of cartridges obtained from investigations. Future work should test more device types and internal temperatures under controlled usage conditions. Identifying the functional materials inside vaping devices can help inform efforts to understand risk. An outbreak of e-cigarette or vaping product use associated lung injury (EVALI) in 2019–20 resulted in hospital admissions and deaths. Vitamin E acetate (VEA) inside tetrahydrocannabinol (THC) vaping devices was identified as the most likely indicator of injury , 4, and A variety of nicotine E-cigarette cartridges and pods have been characterized by determining their design components , testingScanning electron microscopy with energy dispersive X-ray spectroscopy (SEM-EDS) and Fourier-transform infrared (FTIR) micro-spectroscopy are two methods previously used to infer potential environmental pathways from microscale elemental chemistry and morphology of consumer products –21. WillIn this paper, we determined the composition and internal components of EVALI patient-associated THC vaping devices compared to other THC and nicotine devices from 2016–19, specifically the metal, ceramic, and polymer components likely to be exposed to heat. To do this, we have disassembled forty-eight components from eight used and unused THC and nicotine vaping devices under a microscope and analyzed them using portable XRF, SEM-EDS, and limited FTIR micro-spectroscopy.Six THC vape cartridges and two nicotine e-cigarette devices were characterized . Two of The origins and operating voltages of the four THC cartridges from investigations are unknown, and their operating temperatures could not be tested due to the previous extraction of their liquids. The four new THC and nicotine products that were tested for comparison purposes had manufacturer-recommended power sources with batteries and operating voltages ranging from 200–1100 mAh and 2.8–6.0 V. It should be noted that various brands and types of power sources are available on the market, and a user may choose a different power source other than those recommended by the manufacturer. None of the manufacturers provided publicly available, validated measurement data for their internal heating temperatures.All pods and cartridges were disassembled with the aid of a vise, pliers, hand saw, and tweezers to determine their common functional components and were documented with reflected-light stereozoom microscopes at 6.3x – 80x magnification . The inferred plastic or glass construction of the tank walls for these devices is also listed in Components from two of the THC cartridges and both nicotine devices were mounted on Mylar X-ray film and analyzed using a portable XRF  with bench testing stand. Energy calibration was verified with a 316 steel standard. The detection limit varied by element between 10 ppm—1%. The minimum detectable atomic number was 15 . The spatial resolution of the XRF beam was approximately 1cm, and was convenient for integrating over entire pieces of the cartridge, such as the end caps containing the battery contacts.Internal components from all eight pods and cartridges were analyzed by SEM/EDS using an FEI XL30 Environmental SEM with Noran System 7 EDS  and SEMView 8000 electronics . Cartridge and pod components were further disassembled after portable XRF analysis to enable SEM-EDS imaging with 0.1 um spatial resolution. Samples were prepared on standard, aluminum SEM stubs with double-sided, carbon adhesive tabs and no conductive coatings applied. Samples were imaged at 20 kV and 20–2,500x using a tungsten source and an atomic-number-sensitive, back-scattered electron detector for rapid screening of heavy metals. X-ray energy-based element identifications were verified with a copper standard and had a spatial resolution of approximately 5 um. EDS quantitation is generally accurate only for flat specimens. Because some of these parts possessed substantial, variable topography on the order of 1 cm tall, elemental compositions were reported only in terms of qualitative presence/absence of major constituents, with an approximate 1% weight percent reporting limit.-1 spectral resolution from 700–4,000 cm-1. Identification of component mixtures was assigned manually with the aid of quantitative FTIR spectral searches and in-house and commercial libraries of over 100,000 spectra. The estimated limit of detection is on the order of 1%.Limited organic compound speciation was performed using a Nicolet iN-10MX FTIR Microscope with Omnic Picta software and a slide-on ATR detector with germanium crystal and MCT-A detector . Spectra were acquired with 4–8 cmTypical pod and cartridge components revealed during disassembly are summarized in Some design differences between device types were evident. All of the newer THC cartridges possessed ceramic heating elements. The filaments in the TKO and both CCell cartridges were embedded in 6 x 4 mm, tube-shaped ceramics Figs –4. The fThe heating element from an unused Hermes THC cartridge  was a 6 In contrast, the heating elements in the two unused THC cartridges of the older, unknown brands Figs  and 7 poThe used JUUL nicotine pod  differedThe EGO-CE6 coil head for a nicotine device  also appPortable XRF analyses performed on 11 components of the four devices are shown in Portable XRF analyses of both nicotine device filaments  yielded SEM-EDS results from 44 components, some of them obtained from further disassembly of pieces analyzed by XRF, are shown in Figs The fibrous end plug in the unused CCell was fiberglass, primarily Si with minor Al , while tThe ceramics in the EVALI-associated CCell and TKO and the unused CCell and Hermes, were all dominated by Si, with varying amounts of aluminum, sodium (Na), potassium (K), and P. All had pore dimensions on the order of 10 um, presumably to allow oil to soak into the heating elements. The fibrous insulation wraps around these ceramics exhibited predominantly C and a striated morphology consistent with semi-synthetic rayon fibers Figs  and 13E.All filaments were composed primarily of Fe, Cr, and Ni and were brazed to wire leads composed primarily of Ni, except for the unused CCell and two unknown brands, whose filaments were composed of Cr and Ni only. A wire lead from the EVALI-associated CCell had one region at the battery contact end that exhibited Ni, Fe, Cr, inconclusive Co, and sulfur. The JUUL did not possess wire leads. The eGo-CE6 was the only filament that possessed a slightly different metal signature at the joint, Cr and Ni, perhaps to aid in the bond . BecauseThe polymer sheaths around one wire lead each in the Hermes, Unknown #2, and eGo-CE6 all exhibited C and a strong fluorine (F) peak, consistent with a fluorinated polymer such as FEP (fluorinated ethylene propylene) or PTFE (polytetrafluoroethylene), or possibly a polymer coated with PFAS  or PFOS (perfluorooctanesulphonate) Figs  and 18.Higher magnification SEM images of the filaments and wire leads revealed the most surface damage in the used EVALI-associated cartridges. New filament and wire lead surfaces generally exhibited smooth surfaces with prominent grooves, similar to the used JUUL filament shown in FTIR of two polymer components in the Hermes cartridge are shown in . The besThe potential of the identified metals and ceramics to catalyze harmful VEA reactions or aerosolize is dependent on the internal device temperatures achieved in actual usage , 15, 23.The origins and usage conditions of the four THC cartridges from investigations are self-reported or unknown, and their operating temperatures could not be tested due to the previous extraction of their liquids. Nevertheless, the fact that both tested EVALI patient cartridges exhibited severe burn marks confirms that these cartridges were likely operated at high temperatures. This information could be used to help inform harm reduction efforts. In addition, knowledge of the materials in these four investigation-derived THC cartridges is potentially useful in the event that similar cartridges are used at high temperatures.The combination of metals and ceramics identified in the newer, 2019 THC devices is consistent with conditions hypothesized for the formation of ketenes from VEA when sufficiently high temperatures are present –8. The cThe Ni, Cr, Zn, Cu, Pb, Au, and Sn identified in these devices are potential sources of vaping exposures if heated enough to volatilize. Except for Au, these metals are consistent with those reported in previous nicotine device liquids and heated aerosol measurements –16. ThesSeveral of these product designs expose polymer insulation, plugs, or sheaths directly to heat, especially in the THC cartridges. The wire sheaths and insulation fibers were < 5 mm and thus can be considered microplastics . StudiesPortable XRF and SEM-EDS of the same nominal items were generally consistent but detected different elements in some cases, likely due to two main analytical differences: 1) The portable XRF did not detect lower-energy elements observed by SEM-EDS such as C and Si, and 2) The two techniques acquired data from different spatial scales. XRF signals were integrated across larger, centimeter-sized pieces of these products, while SEM-EDS focused on micrometer-sized regions from the disassembled components. For example, no Zr was detected by SEM-EDS in the EGO-CE6 filament or wick fibrils, so it is possible that the minor Zr detected by XRF was associated with heterogeneous regions of the wick not analyzed by SEM-EDS. Similarly, no Pb was detected in any of the individual battery contacts analyzed by SEM-EDS, so the minor Pb detected by XRF may have been in the metal end caps containing the battery contacts, which were too large to be included feasibly in the SEM-EDS analyses. In general, portable XRF was most useful for rapid scanning of heavy metals in the larger cartridge pieces, while SEM-EDS provided more detailed mineral, polymer, and metal compositions of their sub-components.The heavy metals identified in these devices are potential sources of chronic vaping exposures, though their ubiquitous presence in all tested products suggests that the metal components do not explain the relatively recent appearance of acute respiratory cases. It is possible that newer cartridges containing VEA may interact differently with these metals, especially if heated to higher temperatures. Notably, the newer cartridges associated with EVALI cases possessed the most insulating ceramic and polymer materials, and also exhibited more evidence of heat and thermal degradation than a used nicotine pod. These component compositions and high temperatures are consistent with conditions hypothesized to be sufficient for the formation of harmful chemicals (ethenone) from VEA. The fluorinated polymer components observed in some devices have the potential to thermally degrade and volatilize when heated. The apparent substitution of materials to achieve the same design goals, both between and within devices from the same vendors, suggests that the exact compositions of these components will continue to change over time.This work is limited by the fact that many of the tested devices were samples from real investigations with unknown origins and usage parameters. The observed burn marks inside THC cartridges, potential for catalysis, and metal and polymer compositions do not imply that exposures would occur under all usage conditions, and are most relevant to harm reduction efforts based on avoiding higher internal temperatures. Because this work represents a limited sample size, these data should be confirmed by future testing of other THC cartridge compositions, both historical and contemporary. More internal temperature testing of various THC product types is needed as a function of usage conditions, configurations, and liquid levels.


Although many patients do not receive reperfusion therapy because of delayed presentation and/or severity and location of infarct, new reperfusion approaches are expanding the window of intervention. Novel application of neuroprotective agents in combination with the latest methods of reperfusion provide a path to improved stroke intervention outcomes. We examine why neuroprotective agents have failed to translate to the clinic and provide suggestions for new approaches. New developments in recanalization therapy in combination with therapeutics evaluated in parallel animal models of disease will allow for novel, intra-arterial deployment of therapeutic agents over a vastly expanded therapeutic time window and with greater likelihood success. Although the field of neuronal, endothelial, and glial protective therapies has seen numerous large trials, the application of therapies in the context of newly developed reperfusion strategies is still in its infancy. Given modern imaging developments, evaluation of the penumbra will likely play a larger role in the evolving management of stroke. Increasingly more patients will be screened with neuroimaging to identify patients with adequate collateral blood supply allowing for delayed rescue of the penumbra. These patients will be ideal candidates for therapies such as reperfusion dependent therapeutic agents that pair optimally with cutting-edge reperfusion techniques. Stroke is the rapid development of disturbance of cerebral function attributed to the interruption of blood supply. Stroke is diagnosed by imaging evidence of infarct and associated clinical findings. Magnetic resonance imaging (MRI) is most sensitive to cerebral infarct using diffusion weighted techniques to compare the extent of the ischemic core with the extent of perfusion abnormality. Changes in diffusion weighted imaging are apparent in the hyperacute setting. Other modalities can diagnose cerebral infarct in the acute setting including computed tomographic (CT), CT angiogram/perfusion, and diagnostic cerebral angiogram. The two main types of stroke are ischemic and hemorrhagic, with incidences of 85 and 15%, respectively . Ischemiin situ formation of a thrombus on the plaque, causing arterial obstruction  . In the In recent pharmacological developments for stroke, attenuation of microcirculatory disturbances has relied on ablation of single factors in stroke pathogenesis with interventions including recombinant tissue plasminogen activator (rtPA), antioxidants, anti-intercellular adhesion molecule-1 (ICAM-1) antibodies, calcium-stabilizing agents, and anti-excitotoxic agents . HithertLarge amounts of oxygen and glucose are needed to sustain normal brain metabolism. For example, to sustain the propagation of action potentials, 70% of the ATP supplied to the brain is used by Na+/K+ ATPase ion pumps located on the plasma membrane . After gin vivo and in vitro experiments using functional or histological endpoints and showing improvement in focal models, reported that 912 drugs  have failed to achieve clinical success  and inefficient triage of patients needed to facilitate treatment within the recommended window of intervention; and third, adjunct therapies, e.g., collateral flow stimulants, NA-1, Uric acid, Hypothermia, and Activated protein C, in combination with endovascular therapy may enhance the effect of reperfusion .The consensus of STAIR X is that neuroprotective therapies for ischemic stroke have and continue to fail in clinical trials due to the complexity of stroke. First, stroke affects all the cells downstream to the occlusion, so a therapy targeting the neurovascular unit (Brain Cytoprotection) either by pleiotropic effects or improved reperfusion to the infarct core via collateral flow may be a better option compared to a single target approach that has been the primary focus of basic science research. Second, many of the failed clinical trials were designed as monotherapies instead of adjunctive therapies, which may be advantageously paired (pre or post) with current reperfusion approaches such as alteplase and thrombectomy devices. Third, failed monotherapeutic drugs should be reconsidered in adjunct to thrombectomy if they have shown significant improvement of outcomes in animal stroke models and demonstrated safety in phase II or III clinical trials .Another possible factor in limited translational success might be failure of animal models to adequately represent the diversity of patient populations. Due to the small window for reperfusion, a majority of patients do not receive reperfusion. Although estimates vary and rates of thrombectomy/embolectomy are increasing over time, recanalization rates in patients receiving either medical or surgical intervention for large vessel occlusions range from 11 to 40% –38. Rougpost-hoc subgroup analysis of the ESCAPE-NA1 trial demonstrated significantly improved outcomes in patients who received neuroprotectant nerinetide in combination with mechanical thrombectomy but without usual care alteplase  . A majorIntrinsic pathways are activated when mitochondria are damaged, causing membrane depolarization and permeabilization, and subsequently releasing several proapoptotic factors from the mitochondrial space . CyotochIn spontaneously hypertensive rats and aged female mice, 3K3A-APC, an altered version of activated protein c (APC) protease  that has <10% of APC's anticoagulant activity, reduced infarct volume and improved behavior . LiteratTargeting cell-survival pathways, metformin treatment in 40 newly diagnosed acute stroke patients with type 2 diabetes mellitus resulted in improved neurological function and reduced oxidative stress (increasing the expression of glutathione peroxidase and superoxide dismutase) . To elucSecondary to necrosis/apoptosis, neuroinflammation occurs after cytokines are released by immune and CNS-resident cells  . ClinicaTo date, inflammatory treatments are targeting immunosuppression via fingolimod  and inhibition of early platelet tethering and/or activation via glycoproteins. In a pilot trial, treatment with fingolimod FTY720), an immunosuppressive drug used in multiple sclerosis that inhibits T cell activation via downregulation of the sphingosine 1 phosphate receptor, reduced infarct volumes and increased neurological outcomes compared to non-treated patients 0, an imm. Once acBlood vessels carry blood from the heart to the tissue and are composed of arteries and arterioles (carry blood to tissue), the capillary bed (facilitate gas and nutrient exchange), and venules and veins (drain blood from tissue to the heart) . The bloBBB degeneration leads to pathological processes such as hemorrhagic transformation and/or edema, both of which exacerbate brain injury . MRI and+ and glutamate, and that astrocytic but not neuronal Na-K-Cl co-transporter is upregulated after ischemia, astrocytes are more susceptible to water inflow (swelling) than neurons , vascular smooth muscle cells, glial cells , and neurons . The proThe proximal aim of stroke therapy is to restore blood flow to the area of the infarction to reverse and minimize damage from oxygen and nutrient deprivation . In the A secondary and complementary aim of stroke therapy is to use pharmacological agents to attenuate mechanistically deleterious stroke pathways that are activated by ischemia and reperfusion . AlthougCurrent evidence supports endovascular thrombectomy in patients appropriately selected with advanced clinical image, including patients who have already received rtPA for thrombolysis , 101. PaN = 24) within 8 h after stroke improved functional outcomes compared to the placebo group (N = 21)  . Taken tin vivo studies have reported improved outcomes following ischemic stroke via inhibition of the following proteins involved in apoptosis, coagulation cascade, hypoxia induced cascade, and neuroinflammation: Caspase-3 (Recent developments in gene therapy such as RNA interference (RNAi), which are used as a natural defense against exogenous genes, can bind to specific mRNA to interfere with expression, thereby reducing the expression of upstream genes . Even thaspase-3 , Beclin1aspase-3 , Ask1   within 24–72 h of stroke onset resulted in a reduction of 1 point in median day-90 mRS compared to the non-treated control group, wherein no severe adverse events were observed . In a phN = 48) .N = 36). Patients enrolled in the trial demonstrated improved behavioral outcomes over the 12-months of follow-up, e.g., Barthel Index scores improved from a baseline of 11–27% at 6-months and to 35% at 12-months (P < 0.002)  investigating neurological function following intra-arterial injection of CTX0E03 cells  2–13 months after stroke, one of twenty-three and three of twenty-three subjects demonstrated improved motor function at 3 and 6–12 months, respectively, while no deleterious outcomes occurred related to the stem-cell therapy . In a Ph< 0.002) . Taken tIn 2015, several landmark clinical trials comparing the efficacy pharmacological and mechanical thrombectomy for treatment of large vessel ischemic stroke were published . These sStandardized and widespread acceptance of newly developed stent retriever and/or direct aspiration catheters from manufacturers will increase the fraction of stroke patients achieving timely reperfusion –131. AltBuilding on the foundation of recently established interventional techniques, we postulate several unique therapeutic applications that previously would not have been feasible. For instance, balloon occlusion techniques, when applied in combination with mechanical thrombectomy or direct aspiration, may transiently improve local bioavailability of intra-arterial therapies, delaying washout and systemic dilution, which may be especially efficacious in the context of blood brain barrier and endothelial-directed therapeutics. Theoretically, this could allow for much higher efficacy of therapeutics that would otherwise be limited in their systemic dosing or decreased in efficacy once normal, high-volume arterial perfusion is re-established. Similarly, intra-arterial administration of therapeutics at the moment of stent retriever deployment via a guide-catheter would ensure delivery of protective therapeutics to the at-risk vascular territory just before and during the moment of reperfusion.Given that the DEFUSE  and DAWNClinical research has established the principle of “time is brain,” meaning that time delay before intervention is related to the loss of brain tissue . This prThe concept of “time is brain” has been debated, but expert opinion holds that “time is brain” applies most directly to parenchyma lacking sufficient collaterals such that ischemia is primarily a watershed event without opportunity for rescue or subsistence level of perfusion supplied by collateral vessels. In real-world application, collateral circulation varies among individuals and directly affects the course of stroke injury .Taken together, recent developments provide a wide frontier for advancement of the field. Improved imaging technology and interpretation provide the opportunity to test adjunctive therapies selected to pair optimally with reperfusion or collateral status. The logistics of ensuring maximum implementation of new state-of-the-art techniques and widespread, high-quality training of interventionalists with the latest equipment will be an ongoing opportunity for improvement in overall outcomes. Specific devices and techniques will be matched with rationally selected adjunctive therapies. The relationship of time and stroke intervention will continue to evolve from first responders to delayed treatment.Because ~50% of withheld recanalization therapy is attributed to prehospital delays , extendiP = 0.055; N = 96)  and mortality. Molina et al. have reported acute recanalization (<6 h) to result in HT in ~20% of patients compared to ~50% of patients in delayed recanalized groups (<24 h) . Because N = 96) . Small sLack of research on the sequelae of delayed reperfusion is a major barrier preventing its application into clinical trials. Although there is a paucity of information on delayed reperfusion, some basic science researchers have started to investigate novel delayed reperfusion targets. For instance, Mcbride et al. have reported that following permanent MCAO, reperfusion at 3-days, and even 7-days, resulted in improved functional outcome compared to non-perfused animals . In summAlthough not exclusively a form of reperfusion injury, HT after reperfusion may be a primary representation and manifestation of the molecular pathways activated by reoxygenation, exposure to renewed systemic pressures, and delivery of pharmaceuticals such as tPA. Studies have tried to develop imaging criteria to identify high-risk patients. For example, Shinoyama et al. have suggested that time to peak (TTP) mapping, a perfusion-based imaging technique, on admission may identify stroke patients and quantify stroke severity . The sevIn the absences of reperfusion, penumbra is dependent on perfusion to the penumbra from collateral vessels . AlthougApproximately 325,000 ischemic strokes are caused by large vessel occlusion annually in the USA, and only ~20% are treated with recanalization therapy . If the We believe that neuroprotective agents in combination with reperfusion will reduce deleterious side-effects associated with ischemia and reperfusion, especially when tailored to select patient subgroups. An increasing fraction of stroke patients will be treated both within the 6 h window and beyond the traditional time window with improved outcomes. Objective evaluation of collateral vessel status will extend the window for intervention in some patients and will likely improve efficacy of delayed recanalization. In patients with poor quality collateral circulation, dedicated therapies will be deployed to sustain the penumbra until recanalization.The magnitude of stroke incidence justifies further investigation of interventions with potential for notable effect sizes . Even smThe idea and outline of the present review was made by NM, JC, and JZ. This manuscript was drafted by NM. Critical revisions of the manuscript were made by all authors.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.


Stroke-induced cognitive impairments affect the long-term quality of life. High-intensity interval training (HIIT) is now considered a promising strategy to enhance cognitive functions. This review is designed to examine the role of HIIT in promoting neuroplasticity processes and/or cognitive functions after stroke. The various methodological limitations related to the clinical relevance of studies on the exercise recommendations in individuals with stroke are first discussed. Then, the relevance of HIIT in improving neurotrophic factors expression, neurogenesis and synaptic plasticity is debated in both stroke and healthy individuals (humans and rodents). Moreover, HIIT may have a preventive role on stroke severity, as found in rodents. The potential role of HIIT in stroke rehabilitation is reinforced by findings showing its powerful neurogenic effect that might potentiate cognitive benefits induced by cognitive tasks. In addition, the clinical role of neuroplasticity observed in each hemisphere needs to be clarified by coupling more frequently to cellular/molecular measurements and behavioral testing. Stroke is one of the non-communicable diseases with the greatest number of disability-adjusted life years reflecting health loss . CognitiNumerous cognitive training strategies are employed in the clinic such as the use of diary, prompting devices, computers as well as remedial strategies through virtual reality, gaming and several memory tasks ,7. Some Beyond its roles in cardiorespiratory and muscular functions, endurance training can also be considered a part of cognitive rehabilitation in individuals with stroke . Moderat14Exercise regimens with higher intensities, such as high-intensity interval training (HIIT), emerge as encouraging alternatives to improve cardiovascular and brain health following stroke ,25,26,27The purpose of this review is to examine whether HIIT could be suitable for promoting neuroplasticity processes and/or cognitive functions after stroke by discussing findings from molecular to behavioral levels in both human and animal studies. The various methodological limitations in exercise studies, which could explain controversial findings, need to be first considered in this review to clarify both the clinical relevance of rodent studies and the impact of clinical studies on the exercise recommendations. Then, this review highlights the relevance of HIIT in promoting neuroplasticity and/or cognitive functions from single bout of HIIT protocols to training HIIT protocols in both healthy populations (young and older) and individuals with stroke. In addition, the preventive role of HIIT on the stroke severity in rodents (by starting HIIT program before stroke onset) is also discussed. To reinforce the potential role of HIIT in stroke rehabilitation, it is important to discuss the relevance of combining HIIT with cognitive tasks to potentiate their effects on neuroplasticity and cognitive performances.In both rodent and human studies, several methodological limitations keep us from finding optimal endurance programs to recover both cognitive and sensorimotor functions in individuals with stroke. These limitations are related to the heterogeneity of studied populations, the small number of patients, the variability of exercise types  , the timmax) separated by active  or passive recovery periods [2max, % of maximal heart rate or HRmax) during training sessions is not mentioned, despite the fact that it indicates the intensity level reached by patients. In other studies, “vigorous exercise” is defined as an exercise intensity sufficient to produce sweat [The intensity of repeated bouts of HIIT is above the lactate threshold , has recently been used because most individuals with stroke, and all rodents with cerebral ischemia can reach it during an incremental exercise test [LT is known to accurately distinguish between high and moderate running speeds, i.e., below SLT for low intensity and above for high intensity [However, when exercise physiological parameters are used to separate the low- and high-intensity running, moderate running speeds are observed around 17 m/min and the high intensities around 25 m/min in rats with cerebral ischemia . The useise test ,40. Furtntensity ,49,53. CWhen comparing various endurance regimens, the session workload is rarely individualized and standardized (work-matched exercise regimens) in both clinical and rodent studies, while it is strongly preconized in stroke exercise guidelines ,55. All Numerous studies advocated that endurance exercise should be initiated during the acute and subacute phases (first weeks or months) to promote a more effective long-term functional recovery ,56,57. RWhen HIIT starts from 6 to >24 months after stroke, no cognitive changes are found, although aerobic capacity and neuroplasticity are improved . In rodeIn contrast, a high dose of mobilization (mainly out-of-bed activity) very soon after stroke onset (<24 h) negatively impacts recovery as reported by a large controlled randomized trial , which iIn addition, physiological measurements related to neuroplasticity and cognition are also limited in exercise studies. Indeed, for ethical and technical reasons, the most common source for sampling BDNF in humans is peripheral blood . HoweverInterestingly, a preclinical study has proposed a new method to quantify in vivo the brain BDNF in freely moving mice by collecting microdialysate of cerebrospinal fluid during behavioral tasks or stress condition . This stSeveral potential molecular factors might mediate the effects of HIIT on neuroplasticity processes and/or cognitive improvements. First, skeletal muscles are able to communicate with other organs such as the brain through many released substances during exercise . Among oFndc5 gene expression in skeletal muscle and an increase in serum of its secreted form, irisin, after prolonged endurance exercise in mice and humans. It is postulated that irisin itself might be able to cross the blood–brain barrier (BBB) to induce these gene expression changes, or irisin might induce a factor x that can. When hippocampal Fndc5 is upregulated during training, Bdnf and other neuroprotective genes are also activated in the mice hippocampus [Fndc5 and Bdnf genes, thus improving cognition in a mouse model of Alzheimer’s disease [Moreover, recent evidence suggests a potential role in neuroplasticity after HIIT in both humans and rodents of the endurance exercise-induced myokine, the fibronectin type III domain-containing 5 (FNDC5) ,87,88,89 disease .Then, stroke is associated with a strong neuroinflammation that affects neuroplasticity processes within the core of lesion, the penumbra and the remote areas such as the spinal cord ,92,93,94Finally, it is found that genes associated with the inhibitory neurotransmitter gamma-aminobutyric acid (GABA), which regulate the subgranular zone (SGZ) niche of the stem cells by maintaining their quiescent state, were downregulated in rodents exposed to a 28-day running wheel . The decOverall, the impact of endurance training on cognitive benefits and cerebral plasticity was often investigated using MICT in healthy individuals . HoweverIn healthy children, a 4-week HIIT program enhances working memory, as observed with the digit span forward and the Tower of Hanoi test performance without modifying other cognitive tasks . OtherwiAn acute bout of high-intensity exercise is able to modulate complex motor behavior by improving motor skill acquisition and memorization in parallel with an increase in some biomarker concentrations  . For insOther authors found opposite conclusions by showing higher benefits of an acute MICT session on memory performance than HIIT . A meta-triceps brachii FNDC5 (fast-twitch muscle fibers) and hippocampal TrkB after HIIT, but not after MICT, in accordance with other authors who have demonstrated a link between myokines and neurotrophins expression [NTR). Similarly, higher levels of hippocampal BDNF and glial cell-line-derived neurotrophic factor (GDNF) expression are promoted by HIIT compared to MICT [On the one hand, an upregulation of TrkB, VEGF and peroxisome proliferator activator receptor γ coactivator-1α (PGC-1α) levels has been recently found following an 8-week HIIT in the rat hippocampus contrary to work-matched MICT. A positive correlation is also observed between the upregulation of pression . Howeverpression . In unpu to MICT . In line to MICT . Additio to MICT . Additio to MICT . Interes to MICT .A single session of HIIT improves antioxidant mechanisms reducing lipid peroxidation in the hippocampus . SimilarOn the other hand, some authors demonstrated higher effectiveness of MICT to stimulate hippocampal BDNF, IGF-1, VEGF as well as the mitochondrial marker, prohibitin, than a more fatiguing endurance training . Interes2 consumption, but no cognitive recovery is found [is found . Interesis found . Other ais found ,27. Indeis found   HIIT-induced functional and cognitive changes in rodents with cerebral ischemia. Rezaei et al. are the first to show that an 8-week HIIT can protect BBB integrity, decreasing inflammatory cells infiltration, thereby reducing cortical and total cerebral infarction volumes compared to MICT . MoreovePreconditioning HIIT also improves neurological score in rodents by preventing motor deficits and enhancing their recovery ,153. ThiRecent studies have reported promising effects of HIIT on cognitive functions in neurodegenerative diseases. In patients with middle cognitive impairments, HIIT combined to a ketogenic diet and memory training may reverse early stage memory loss . MoreoveIt seems that HIIT should be included in stroke rehabilitation for its beneficial effects on neuroplasticity processes. The clinical role of neuroplasticity observed in each hemisphere needs to be clarified by coupling more frequently cellular/molecular measurements and behavioral testing. Despite these results, HIIT induces very modest cognitive effects when performed alone in both healthy people and individuals with stroke. However, its powerful neurogenic effect might help to accentuate benefits induced by cognitive tasks. Based on these considerations, it is recommended to continue investigating the different modalities of HIIT on brain plasticity in terms of duration and/or intensity of both high-intensity intervals and recovery phases along with the type of recovery between series (active or passive) and the mode of HIIT exercise . It is noteworthy that this review is not designed to demonstrate a useless/ineffective role of MICT after stroke. In contrast, we believe that both low- and high-intensity training regimens might be complementary for brain health.


Post-traumatic stress disorder (PTSD) is a prevalent, debilitating, and costly psychiatric disorder. Evidenced-based psychotherapies, including Cognitive Processing Therapy (CPT), are effective in treating PTSD, although a fair proportion of individuals show limited benefit from such treatments. CPT requires cognitive demands such as encoding, recalling, and implementing new information, resulting in behavioral change that may improve PTSD symptoms. Individuals with PTSD show worse cognitive functioning than those without PTSD, particularly in acquisition of verbal memory. Therefore, memory dysfunction may limit treatment gains in CPT in some individuals with PTSD.Here, we present a protocol describing the Cognition and PsychoTherapy in PTSD (CPTPTSD) study, a prospective, observational study examining how cognitive functioning affects treatment response in CPT for PTSD (NCT# 03641924). The study aims to recruit 105 outpatient veterans with PTSD between the ages of 18 and 70 years. Prior to beginning 12 sessions of CPT, Veteran participants will have standardized assessments of mood and functioning and complete a comprehensive neurocognitive battery assessing episodic learning, attention and speed of processing, language ability, executive control, and emotional functioning. This study aims to fill gaps in the current literature by: (1) examining the specificity of memory effects on treatment response; (2) exploring how baseline cognitive functioning impacts functional outcomes; and (3) examining potential mechanisms, such as memory for treatment content, that might explain the effects of baseline memory functioning on PTSD symptom trajectory.If successful, this research could identify clinically relevant neurocognitive mechanisms that may impact PTSD psychotherapy and guide the development of individualized treatments for PTSD. Individuals who experience combat, sexual trauma, and other traumatic events are at higher risk for developing post-traumatic stress disorder (PTSD). PTSD is the most common mental health diagnosis received by returning Iraq and Afghanistan (OEF/OIF/OND) veterans at Veteran’s Administration (VA) facilities , with liCognitive Processing Therapy (CPT) is a trauma-focused psychotherapy strongly recommended for the treatment of PTSD . CPT is One factor that may impact psychotherapy in PTSD is cognitive functioning. Psychotherapy requires patients to encode and consolidate new information, organize this information into behavioral plans, and remember and execute these plans. Thus, the capability to retrieve specific details from one’s past and encode/retrieve treatment-relevant skills are highly relevant for therapeutic change across most types of psychotherapy . CriticaAccumulating research reports remarkably consistent effects of verbal memory deficits on psychotherapy outcomes in PTSD. In a small study of civilians in trauma-focused CBT, Furthermore, despite accumulating research in this area, no studies to date have examined the pathways by which cognitive functioning may affect treatment outcomes. For example, one pathway by which cognitive dysfunction may interfere with psychotherapy is by directly limiting treatment understanding, learning, or implementation, or interfering with a patient’s ability to re-contextualize memories. Evidence from research in substance use disorders and depression indicates that worse executive functioning, memory, and overall cognitive abilities are associated with poorer recall of treatment information , reducedIn sum, accumulating evidence indicates that memory dysfunction is prominent in PTSD and adversely affects psychotherapy outcomes. Identifying mechanisms underlying the relationship between memory dysfunction and poor treatment outcomes is crucial to provide empirical guidance regarding appropriate rehabilitation targets to reduce the impact of cognitive dysfunction on therapy outcomes. The specific rehabilitation approaches that are most appropriate for improving therapy outcomes in PTSD will depend on the specific deficits observed, their relationship with therapeutic and functional outcomes, and critical moderators or mediators of those relationships. Given convergent findings of associations between cognitive functioning and PTSD and its influence on psychotherapy outcomes, the current protocol was developed to provide a rigorous examination of such relationships using hypothesis-driven models. Notably, no prior studies have examined effects of cognitive functioning on psychotherapy outcomes as a primary aim or explored potential mechanisms underlying these relationships. The current protocol therefore aims to address several knowledge gaps in the literature.how episodic memory functioning might lead to poor treatment outcomes. Our primary hypothesis for this aim is that treatment recall at the treatment midpoint will mediate the association between verbal immediate memory at baseline and treatment outcomes. Finally, in an exploratory fashion, we also aim to examine whether alternative models of dysfunctional memory in PTSD, including impairment in autobiographical memory , but not overall cognitive performance, will interact with time to predict PTSD symptom changes at post-treatment, such that those with lower verbal learning performance at baseline will have less reduction in PTSD symptoms than those with higher verbal learning performance. Second, we aim to determine whether specific treatment-related variables  mediate the relationship between memory functioning and CPT response, helping to explain l memory , emotionl memory , or prosl memory , might bn = 105 after attrition; see Missing Data below) outpatient veteran participants aged between 18 and 70 who meet criteria for PTSD. Veteran outpatients are being recruited from the Corporal Michael J. Crescenz Veterans Affairs Medical Center (CMCVAMC) in Philadelphia, Pennsylvania.The Cognition and PsychoTherapy in PTSD: Mechanisms and Functional Outcomes  study is an observational, prospective study aiming to recruit 140  within the outpatient mental health clinic at CMCVAMC. Participants are referred to the study following evaluation from a behavioral health technician, in lieu of transfer to standard outpatient psychotherapy appointment for an EBP for PTSD, or by a pre-existing outpatient mental health provider. Inclusion criteria includes meeting DSM-5 criteria for PTSD via the Clinician Administered PTSD Scale (CAPS-5) and being between the ages of 18 and 70. Exclusion criteria includes moderate or severe substance use disorder (SUD) not in remission for >1 month; daily benzodiazepine use; severe psychiatric illness ; significant current suicidal or homicidal intent, including a specific plan; dementia, neurological disorder, or severe TBI ; inability to speak or read English; prior completion of CPT; and psychiatric hospitalization within 30 days of study entry. To enhance generalizability among our sample of veterans with PTSD, participants are allowed to concurrently take medications for PTSD and other mental  or physical health conditions or attend concurrent talk or supportive therapy.The first recruitment of veteran participants started in July 2019 with recruitment expected to be completed in March 2023. Participants undergo study screening to determine initial eligibility. Major assessments are conducted with participants at study entry (baseline), session 4, session 6, session 8, and at an endpoint assessment at post-treatment. For a full breakdown of all assessments administered at these time points see . BaselinFollowing each CPT visit, participants complete a brief questionnaire about adherence and application of treatment (see below), as well as the Patient Health Questionnaire PHQ-9;  to assesNote that with the outbreak of the novel Coronavirus-19 (COVID-19) in March of 2020, all study procedures have moved to a virtual platform, and verbal consent is being conducted, as approved by the CMC VAMC IRB.During the baseline visit, participants are interviewed by a trained and experienced masters-level clinician using the: (1) Clinician Administered PTSD Scale-5 , the golTwo assessments are administered to capture primary outcomes: (1) PTSD Checklist for DSM-5 PCL-5;  to assesTwo instruments are administered to capture covariate and predictor outcome variables. First, the PHQ-9, as mentioned previously, assesses depressive symptoms. Second, the Combat Experiences Scales CES;  is used The neurocognitive battery assesses participants’ functioning across a broad range of neurocognitive domains using both traditional paper and pencil testing and computerized measures from the Penn Computerized Neurocognitive Battery PennCNB; , 2010, aPresently, there are no ideal measures with extensive evidence of reliability and validity to assess treatment adherence, memory for treatment content, and treatment expectancy. Thus, measures with strong theoretical foundations and initial evidence of validity were chosen for administration. Treatment adherence is measured weekly by the Therapist- and Patient-Report versions of the Treatment Adherence Rating Scale TARS; . The TARTreatment expectancy is captured through three measures, including: (1) Expectancy of Therapeutic Outcome ETO; , a 5-iteParticipants also receive three additional measures of memory processes shown to be dysfunctional in PTSD, including the Autobiographical Memory Test AMT; , the EmoStatistical models used are linear or generalized linear mixed effects regression models, and we will use standard residual-based diagnostic approaches to check that model assumptions are satisfied . We will examine demographics as covariates in analyses to account for possible confounding of memory effects by demographics .z-score of CVLT-2 Trials 1–5 and WMS Logical Memory I. The comparison will be based on repeated responses obtained at baseline, mid-point, and end of treatment. The model for each outcome will include a random intercept to accommodate between-subject heterogeneity  and memory for treatment (PRT) mediate the effects of baseline verbal learning on end of treatment PTSD symptoms and overall functioning, we will use a multiple mediator model following Third, to determine whether aspects of memory are better predictors of treatment and functional outcomes than verbal memory in exploratory analyses, similar mixed effects models as the primary analyses will be conducted to examine the effects of baseline autobiographical memory, emotional memory, and prospective memory on overall PTSD symptom response.Finally, a small set of additional covariates potentially associated with PTSD psychotherapy outcomes, including TBI status and depression severity, will be included in reruns of our primary analyses. Model selection approaches based on BIC model fit comparisons to examine the separate and joint variance explained by primary and additional covariates will be used. We will also examine models with interactions between verbal memory and these additional covariates to assess whether they act as moderators of memory effects. Although exploratory in nature, these covariates will be included as baseline variables in the latent growth mediation models.n = 140, although the sample size of those who initiate CPT is expected to be n = 105.Based on our team’s prior studies, we anticipate a loss of up to 25% of veterans who enroll but do not start treatment (note that this study does not involve randomization). Thus, our planned enrollment is Sporadic missing items from a scale may yield a small amount of missing data. We will accommodate this in analyses by mean imputation of a total scale score, based on available items of all participants at that time point. Mixed-effects models and longitudinal mediation analyses make use of all data provided by participants. The analysis on the combined set of complete and incomplete data will be valid if the missing-at-random (MAR) assumption is met; in practical terms, this means that a future missed visit can be well-predicted on the basis of available data. We will perform sensitivity analyses using selection models  to assesz-scored versions of baseline verbal learning and the response, a one unit increase in verbal learning corresponded to an additional 0.15 unit decrease in the estimated change in PTSD outcomes. Based on 500 simulated datasets, we have 80% power for that observed effect for the PTSD outcome, and 81% power for slightly larger effects  for functioning measures. For mediation hypotheses of Aim 2, we use the methods of We regard our hypothesis on PTSD symptoms as separate from hypotheses on functioning, so we use a 5% alpha-level for both outcomes. Prior data  suggest This protocol paper describes an observational prospective study to examine the relationship between neurocognitive mechanisms and CPT outcomes in a sample of veterans with PTSD. Prior research has identified cognitive deficits in individuals with PTSD, with the largest effects in immediate verbal memory . MoreoveThis study has limitations. First, the protocol is not a randomized controlled trial (RCT), and therefore we will not be able to definitively know if symptom changes during CPT that are associated with neurocognitive functioning would have happened absent any intervention. However, conducting an RCT was not necessary to examine most relationships posed in our hypotheses. Second, we have faced numerous challenges with the outbreak of COVID-19, yet we have innovated and effectively transitioned much of our protocol to a virtual platform. Lastly, our study population is comprised solely of veterans, and results may not generalize to the civilian population. However, there is evidence that veteran and civilian PTSD rates are similar after experiencing similar traumas , and theThis project was approved and carried out with feedback from the Corporal Michael J. Crescenz Veterans Affairs Institutional Review Board. All subjects will provide written or verbal consent prior to enrollment into the study.Findings from this research will be disseminated in peer-reviewed journals and scientific meetings. We will also communicate results and implications to critical stakeholders in the veteran and PTSD research communities. Results will also inform the long-term goal of this program of research, which is to understand the role of cognitive dysfunction in psychotherapy treatment outcomes and identify novel targets for rehabilitation  that can be translated into clinical practice. Results from this study may also have wider clinical applicability, as they could lead to changes in other psychotherapeutic approaches for PTSD, as well as other disorders impacting cognition . For example, “fine-tuned” psychotherapies could be developed to enhance memory for treatment content by providing focused memory support interventions, which could be broadly applied to help facilitate treatment effectiveness. One novel and promising pathway is to train therapists to infuse psychotherapy treatment-as-usual with a memory support intervention to enhance patient memory for the content of treatment . This inThe studies involving human participants were reviewed and approved by the Corporal Michael J. Crescenz Veterans Affairs Institutional Review Board. The patients/participants provided their written or verbal informed consent to participate in this study.JCS obtained funding and is responsible for the overall study concept and design. DC and JCS drafted the manuscript. AA, CD, KL, JG, and GH provided critical revision of the manuscript for important intellectual content. KL also provided technical and statistical support. 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 terminology “gut-brain axis “points out a bidirectional relationship between the GI system and the central nervous system (CNS). To date, several researches have shown that migraine is associated with some gastrointestinal (GI) disorders such as Helicobacter pylori (HP) infection, irritable bowel syndrome (IBS), and celiac disease (CD). The present review article aims to discuss the direct and indirect evidence suggesting relationships between migraine and the gut-brain axis. However, the mechanisms explaining how the gut and the brain may interact in patients with migraine are not entirely clear. Studies suggest that this interaction seems to be influenced by multiple factors such as inflammatory mediators , gut microbiota profile, neuropeptides and serotonin pathway, stress hormones and nutritional substances. Neuropeptides including CGRP, SP, VIP, NPY are thought to have antimicrobial impact on a variety of the gut bacterial strains and thus speculated to be involved in the bidirectional relationship between the gut and the brain. According to the current knowledge, migraine headache in patients harboring HP might be improved following the bacteria eradication. Migraineurs with long headache history and high headache frequency have a higher chance of being diagnosed with IBS. IBS and migraine share some similarities and can alter gut microflora composition and thereby may affect the gut-brain axis and inflammatory status. Migraine has been also associated with CD and the condition should be searched particularly in patients with migraine with occipital and parieto-occipital calcification at brain neuroimaging. In those patients, gluten-free diet can also be effective in reducing migraine frequency. It has also been proposed that migraine may be improved by dietary approaches with beneficial effects on gut microbiota and gut-brain axis including appropriate consumption of fiber per day, adhering to a low glycemic index diet, supplementation with vitamin D, omega-3 and probiotics as well as weight loss dietary plans for overweight and obese patients. Based on global burden of headache reports in 2016, it was estimated that approximately 14% of the adult population worldwide suffer from migraine . The exaThe terminology “gut-brain axis “points out a bidirectional relationship between the GI system and the central nervous system (CNS). Brain normally regulates movements and functions of the GI tract (sensory and secretion). Hormonal factors through the hypothalamic pituitary adrenal (HPA) axis by mediating stress responses impact on the gut functions. On the other hand, GI system is believed to be able to affect the CNS. A number of the brain functions such as cognition, behavior and even nociception are under the influence of the gut system , 6. The The present review article aims to discuss the direct and indirect evidence suggesting relationships between migraine and the gut-brain axis. As it will be clarified later, this interrelationship seems to be influenced by multiple factors such as inflammatory mediators, gut microbiota profile, neuropeptides, stress hormones and nutritional substances. In this regard, at first we will take a look at the involvement of inflammation in migraine headache and role of gut microbiome. Afterwards, the role of the neuropeptides specifically serotonin pathway in relation to migraine and gut-brain axis will be explored. Later, the current evidence on the association between migraine and gastrointestinal (GI) disorders including Helicobacter pylori (HP) infection, irritable bowel syndrome (IBS), celiac disease (CD) and inflammatory bowel disease (IBD) will be described. Finally, the effects of probiotic supplementation on migraine and possibly effective dietary approaches for migraine patients including modifying carbohydrate and fat intake, vitamin supplementation and weight loss diets will be discussed.In the gut, immune cells and their inflammatory mediators such as interleukin (IL)-1β, IL-6 and IL-18, tumor necrosis factor alpha (TNF-훼), and interferon gamma (IFN)-훾 have been implicated as sensitizers of afferent endings and are known as inducers of visceral pain , 10. AddMost of the studies aiming to investigate the roles of gut microflora in different disorders, usually use microbiota deprivation or “germ-free” murine models. These animals are grown in a sterile condition and therefore are microbiota-deficient . In thisIt is noteworthy that the composition of the gut microbiota plays a major role in gut-brain axis. This happens via two mechanisms: indirect signaling, including microbiota-derived neurotransmitters, inflammatory molecules, and hormones; and direct connection with stimulating end terminals of the vagus nerve Fig. . Also inEscherichia coli, Enterococcus faecalis, and Lactobacillus acidophilus) and thus speculated to be involved in the bidirectional relationship between the gut and the brain [Lactobacillus paracasei administration attenuated this response [Substances such as CGRP, substance P (SP), vasoactive intestinal peptide (VIP), and neuropeptide Y (NPY), are thought to have antimicrobial impact on a variety of the gut bacterial strains , central sensitization, and by stimulating trigeminovascular system. Increased glutamate levels in the plasma and CSF of migraineurs in comparison with control subjects have been reported previously , 45. On NPY, a probable indicator of noradrenergic system function, affects cerebral blood flow through cerebral circulation regulation. It is worth mentioning that higher levels of NPY has been detected in ictal phases of migraineurs , 44. IntOne of the main biomarkers of migraine, CGRP, inhibits gastric acid secretion and may suppress food intake . As mentCholecystokinin (CCK) is synthesized by enteroendocrine cells in the mucosal lining of small intestine (I cells), mammalian brain  and midbrain), and spinal cord. This peptide inhibits gastric emptying and acid secretion, stimulates gall bladder contraction and pancreatic secretion, and provokes satiety feeling in the brain via CCK1 receptor. The presence of CCK1R on vagal afferent terminals lying in the wall of GI tract further confirms the hypothesis of endocrine and paracrine collaboration. The location of CCK1R in mesolimbic structures, hypothalamus and brain stem nucleus and its interaction with dopamine, serotonin, glutamate, hypothalamic hormones, and neuropeptides explains specific roles of CCK in behavior, mood, and extrapyramidal function , 50. LikIt has been proposed that tryptophan-kynurenine pathway might be one of the main ways by which GI microbiota may affect the function of the CNS due to the presence of synthetic enzymes in bacterial strains of the intestine that may produce tryptophan metabolites (such as quinolinic and kynurenic acids) , 56. As The role of gut microbiota in anxiety, depression and the HPA axis has been studied through assessing the effects of prebiotics and probiotics administration in these conditions –59. It hMoreover, the presence of serotonin receptors on immune cells including monocytes, macrophages, lymphocytes, and dendritic cells, may shed light on the impact of this neurotransmitter on immune modulation. This issue might also explain effects of serotonin on inflammatory state of the intestine and its role in increasing the susceptibility for GI diseases  .To date, several researches have shown that migraine is associated with some GI disturbances such as diarrhea, constipation , 63, dysBased on the findings of a meta-analysis of 5 case-control studies, about 45% of migraineurs harbor HP, while the prevalence rate among healthy controls was estimated at about 33% . It is nAccording to a review article findings, evidence supported that HP eradication may be associated with relief of migraine symptoms . Also, FHP infection is supposed to be related to a persistent chronic inflammatory state, which in turn may lead to increased production of inflammatory mediators and vasoactive compounds , 75. TheCollectively available data indicate that HP may impact on migraine symptoms. However, at the moment this is still a matter of research and there is no evidence to support systematic search of the infection in migraineurs nor the treatment of the infection specifically aimed to improve migraine symptoms.IBS and migraine share some similarities, i.e. both disorders are chronic, recurrent , and morThere is an established association between migraine and IBS, such that IBS was revealed to be common among migraineurs and migraine was reported to be prevalent among IBS suffers . A prevaMechanisms underlying the association are not entirely clear. Similar to migraine, IBS can alter gut microflora composition and thereby may affect the gut-brain axis and inflammatory status , 89–93. Also 5H-T has been postulated to play a role in the association of migraine and IBS. Serotonin release from entrochromaffin cells in the gastric pits of the stomach luminal epithelium stimulates gastric acid secretion, sensory and motor GI reflexes which in turn activates the ENS , 95, 96.CD is an autoimmune multisystem condition caused by gluten peptide in genetically susceptible individuals . AccordiIBD is a chronic relapsing-remitting inflammatory condition that consists of Crohn disease and ulcerative colitis . Data abDifferent dietary approaches have been suggested for subjects with migraine , but at Probiotic supplementation may modulate migraine attacks. Possible mechanisms of action are unclear and include promoting SCFA production in the gut and improving epithelial integrity of the intestine and enhancing inflammation by nuclear factor kappa-B (NF-κB) pathway suppression and therefore lowering proinflammatory cytokines levels , 124–127Some studies have explored the beneficial effects of probiotic administration on migraine headaches , 130. InIn an open-label trial in 40 migraineurs, 12-week supplementation with probiotic + minerals + vitamins + herbs, resulted in significant improvement in patients quality of life in approximately 80% of the subjects and pain relief in more than half of the migraineurs . In a fuGut microbiota fermented resistance carbohydrate to different metabolites, such as SCFAs . The dieChronic exposure to omega-3 in utero and early life, increases the proliferation of Lactobacillus and Bifidobacterium and sp., results in more SCFAs production . Four stVitamin D3 supplementation affected the intestinal microbiota composition. Supplementing healthy individuals with vitamin D3 for 8 weeks also significantly reduced Helicobacter sp. count . SeveralAnimal studies showed that obesity reduced gut permeability, reduced expression of tight junctions and could influence intestinal microbiota composition . Pieces The current evidence shows that the gut-brain axis may impact on migraine despite the mechanism explaining this interaction is not entirely clear. Generally, this interaction seems to be influenced by multiple factors such as inflammatory mediators , gut microbiota profile, neuropeptides and serotonin pathway, stress hormones and nutritional substances. Neuropeptides including CGRP, SP, VIP, NPY are thought to have antimicrobial impact on a variety of the gut bacterial strains and thus speculated to be involved in the bidirectional relationship between the gut and the brain. Additionally, there is comorbidity between migraine and a number of conditions including HP infection, IBS, IBD, and CD. According to the current knowledge, migraine headache in patients harboring HP might be improved following the bacteria eradication. Migraineurs with long headache history and high headache frequency have a higher chance of being diagnosed with IBS. IBS and migraine share some similarities and can alter gut microflora composition and thereby may affect the gut-brain axis and inflammatory status. Migraine has been also associated with CD and the condition should be searched particularly in patients with migraine with occipital and parieto-occipital calcification at brain neuroimaging. In those patients, gluten-free diet can also be effective in reducing migraine frequency. Diet strategies may impact on migraine course and could represent a valuable instrument to improve migraine management. However, no definite conclusion can be drawn because of the limited evidence on migraine management with diet. It can be hypothesized that prescribing dietary approaches with beneficial effects on gut microbiota and gut-brain axis including appropriate consumption of fiber per day, adhering to a low glycemic index diet, supplementation with vitamin D, omega-3 and probiotics as well as weight loss dietary plans (in case of obese patients) could lead to improvements in migraine associated features.