It has been well known that a variety of pro-inflammatory cytokines and related signaling pathways are involved in the procession of DKD. Additionally, some anti-hyperglycemic agents and mineralocorticoid receptor antagonists (MRAs) that are effective in alleviating the progression of DKD have anti-inflammatory properties, which might have beneficial effects on delaying the progression of DKD. However, there is currently a lack of systematic overviews. In this review, we focus on the novel pro-inflammatory signaling pathways in the development of DKD, including the nuclear factor kappa B (NF-κB) signaling pathway, toll-like receptors (TLRs) and myeloid differentiation primary response 88 (TLRs/MyD88) signaling pathway, adenosine 5′-monophosphate-activated protein kinase (AMPK) signaling pathways, inflammasome activation, mitochondrial DNA (mtDNA) release as well as hypoxia-inducible factor-1(HIF-1) signaling pathway. We also discuss the related anti-inflammation mechanisms of metformin, finerenone, sodium-dependent glucose transporters 2 (SGLT2) inhibitors, Dipeptidyl peptidase-4 (DPP-4) inhibitors, Glucagon-like peptide-1 (GLP-1) receptor agonist and traditional Chinese medicines (TCM).Diabetic kidney disease (DKD) is a common complication of diabetes mellitus and a major cause of end-stage kidney disease (ESKD). The pathogenesis of DKD is very complex and not completely understood. Recently, accumulated evidence from  Diabetic kidney disease (DKD), which is also known as diabetic nephropathy before, is the most common microvascular complication of diabetes mellitus . The numPrevious studies demonstrate that many factors are implicated in the progression of DKD, such as metabolic abnormalities and hemodynamic changes caused by hyperglycemia and insulin resistance, etc . It has Inflammation is a biologically conserved immune defense mechanism . In humaIn this review, we discuss the role of some novel and key pro-inflammatory signaling pathways in the progression of DKD, such as the nuclear factor kappa B (NF-κB) signaling pathway, toll-like receptors (TLRs) and myeloid differentiation primary response 88 (TLRs/MyD88) signaling pathway, adenosine 5′-monophosphate-activated protein kinase (AMPK) signaling pathways, inflammasome activation, mitochondrial DNA (mtDNA) release and hypoxia-inducible factor-1(HIF) signaling pathway. We also introduce the role of anti-hyperglycemic drugs, non-steroidal selective MRA and traditional Chinese medicine in anti-inflammation during the treatment of DKD.Hyperglycemia and insulin resistance are the most common pathological characteristics in patients with DKD and can lead to systemic low-grade inflammation . HyperglThe transcription factor NF-κB is a key mediator of inflammation and NF-κB remains inactive while binding to the proteins of the IkappaB (IκB) family (mainly IκBα) . There aIt has been observed that activated NF-κB signaling pathway is associated with inflammation and fibrosis in DKD . ForestoToll-like receptors (TLRs) is an evolutionarily ancient family of pattern recognition receptors and an important component of innate immunity . ActivatIt has been demonstrated that the high glucose (HG) could activate TLRs-MyD88 signaling pathway in the kidneys of DKD, which is the initiator of renal interstitial fibrosis . Liu et Inflammasome was first proposed by Tschopp et al., in 2002, which could modulate inflammation by the activation of caspases . There aAccumulated data suggests that the inflammasome contributes to the progression of DKD . UpregulOther novel inflammasomes such as NLRP2, NLRP6, NLRP10, NLRP12, and AIM2 are also found to be involved in the regulation of inflammation and their expressions are upregulated in various kidney diseases . It has AMP-activated protein kinase (AMPK) is a serine/threonine kinase that is involved in regulating cellular metabolism by activating energy-producing pathways  Wang et, In addiInterestingly, AMPK signaling pathway was related to the renal inflammation . PreviouWhen mitochondria are dysfunctional, the mitochondrial DNA (mtDNA) translocates to cytoplasm and initiates the inflammation through various signaling pathways  Zhang e. First, As we know that proximal tubular cells of the kidney are rich in mitochondria and mitochondrial dysfunction and mtDNA release have been recognized in various kidney disease . A largeHypoxia-inducible factor (HIF) is conserved in biological evolution and is an important mediator of inflammation . The strRecent studies have shown that HIF signaling pathway is closely related to the development of renal interstitial fibrosis in DKD . The actIn addition to be involved in the pathogenesis of DKD, inflammation has recently been shown to play a role in diabetic cardiomyopathy . Under hThe treatments of DKD mainly include management of hyperglycemia, hypertension and hyperlipidemia. Actually, these drugs especially the nonsteroidal selective mineralocorticoid receptor antagonist (MRA) finerenone and some Chinese traditional medicine could alleviate the development of DKD by regulating the inflammation.Metformin has been used for treatment of diabetes mellitus since 1950s, and is now the most widely used anti-hyperglycemic drug . There’sNonsteroidal selective MRA is a novel drug with cardio-renal protection. Compared with traditional MRAs such as spironolactone and eplerenone, nonsteroidal selective MRA has significantly improved efficacy and safety. Finerenone  is currently approved by FDA for treatment of DKD. A series of clinical studies confirmed the reno-protective effect of finerenone in DKD. The phase III FIDELIO-DKD trial has shown that finerenone significantly delayed renal failure and eGFR decline. In addition to finerenone, several nonsteroidal MRAs are currently in development , such asIt has been well known that overactivation of MR is a key event for chronic inflammation by increasing the recruitment of neutrophil, macrophage and Th1 & Th17 cells and upregulating the expression of pro-inflammatory factors and fibrotic-related factors including TGF-α, endothelin 1, PAI-1, CTGF . There i2+ concentration and activating AMPK signaling pathway (Sodium-glucose cotransporter 2 inhibitor (SGLT2i) is a novel anti-hyperglycemic drug by inhibiting reabsorption of glucose in the kidney . Several pathway . Recent  pathway . Interes pathway . Birnbau pathway . In concGlucagon-like peptide 1 (GLP-1) is an incretin hormone that stimulates insulin secretion in response to food intake . GLP-1 bAstragalus, Cordyceps sinensis, Tripterygium wilfordii, Fructus arctii, Panax notoginseng, Berberine, etc. are also powerful weapons against DKD  such as inst DKD . A systeinst DKD . Furtherinst DKD . Some stinst DKD . The Triinst DKD . Guo et inst DKD . In addiinst DKD . All of In addition to TCM, nutraceuticals derived from Chinese herbal medicines could prevent DKD, for example, boswellic acid, curcumin and quercetin . A systeHowever, TCM has limitations such as low bioavailability, which greatly limits its broad application in the treatment of DKD . The stuMesenchymal Stem Cells (MSCs) and extracellular vesicles (EVs) are current research hotspots and pre-clinical agents for the treatment of DKD . PreviouThe EVs include exosomes, microvesicles and apoptotic bodies. It has been found that the EVs play an important role in the progression of DKD . Under tInflammation plays an important role in the progression of DKD . In thisIn addition, MSCs and EVs have shown the curative effect and anti-inflammatory effect in the treatment of DKD animal models . However


This article aims to provide an immunological explanation for the delayed side effects of a SARS-CoV-2 vaccine during an episode of natural infection. We reported a case of a 39-year-old male healthcare worker who complained about pruritus and discomfort around the injection site of an inactivated SARS-CoV-2 vaccine administrated 18, 17, and 13 months earlier. Those symptoms resembled the side effects previously experienced with one of the booster doses, and a sole erythematous papule was also documented. The patient was diagnosed with COVID-19 one or two days after noticing these local signs and symptoms, and high serum titers of immunoglobulin M (IgM) and immunoglobulin E (IgE) were found five weeks after the onset, along with SARS-CoV-2-specific immunoglobulin G (IgG) antibodies. Therefore, the OAS might be a plausible phenomenon to consider in individuals immunized with inactivated vaccines and exposed secondarily to a wild virus with antigenic variations.Although the  Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection emerged in the current coronavirus disease 2019 (COVID-19) pandemic, which has triggered a notable development of SARS-CoV-2 vaccines. Consequently, there have been considerable achievements in understanding the humoral and cellular immune responses following vaccination and natural infection. As most vaccines use a two-dose prime-boost approach to generate an immune response against SARS-CoV-2 antigens, individuals with a previous infection develop higher antibody titers than uninfected ones ["original antigenic sin" (OAS) effect at the immune system’s second exposure to a similar pathogen to which it was previously exposed, generating cross-reactive antibodies already present against the initial antigens, has been predicted against new variants of SARS-CoV-2 [However, the RS-CoV-2 .Since massive vaccination has been implemented, reports of adverse events following approved vaccines have increased worldwide. The most frequent cutaneous effects reported are localized at the injection site , with a We report significantly delayed side effects of an inactivated SARS-CoV-2 vaccine during an episode of natural infection in a case where the OAS is a plausible phenomenon. This paper aims to provide an immunology-based explanation of the clinical events reported and raises the emerging need to evaluate further the long-term safety and effectiveness profiles of current COVID-19 vaccines that received an emergency use or conditional marketing authorization.A 39-year-old male healthcare worker diagnosed with mild asthma received three doses of an inactivated SARS-CoV-2 vaccine  at 18, 17, and 13 months preceding his COVID-19 infection. Each vaccine was given intramuscularly (0.5 mL) in the right deltoid area; the second and third doses were associated with transient itchiness, redness (third dose only), and discomfort around the injection site with spontaneous resolution within 24-48 hours. His vaccination record showed the hepatitis B vaccine four years before, and no other vaccines followed SARS-CoV-2 immunization.After eight or nine days of being in close and continuous contact with a confirmed COVID-19 case, the patient started with pruritus and discomfort around the vaccine administration site, resembling the side effects experienced previously. The difference in this episode consists of the appearance of one sole erythematous papule, firm, slightly indurated, around 0.5-0.6 cm in diameter in the SARS-CoV-2 vaccine injection area, as shown in Figure Perilesional tenderness and pruritus lasted until Days 2-3 without warmth or lymphadenopathy, and the cutaneous events were solved spontaneously within one week, without complications. The complete blood counts before and after the COVID-19 infection were unremarkable. However, high serum titers of IgM  and IgE  were found five weeks after the onset of COVID-19 symptoms. At that time, anti-SARS-CoV-2 total antibodies were also reactive , along with SARS-CoV-2-specific IgG antibodies .It is known that the OAS phenomenon occurs in the second exposure to a similar pathogen (SARS-CoV-2 wild-type virus) to which it has previously been exposed (inactivated SARS-CoV-2 vaccine). Herein, overproduction of memory B cells could affect the naïve B cells activation capable of generating efficient antibodies, triggering immune evasion of the novel variants [As reported by Kostoff et al., numerous long-term potential issues concerning vaccines have been identified that can explain , at leasAntibodies originating from the inactivated vaccine may have specificity or cross-reactivity with immunodominant antigens circulating (or viremia) preceding virus detection on the nasopharyngeal swab. Irrespective of the viral variant causing the primary response, imprinting SARS-CoV-2 specific antibody reactions lead to a secondary response that favorably boosts against the primary antigen . MoreovePark et al. stated that the OAS might cause additional inflammatory responses in specific circumstances of antigenic variation . After iWhereas the Th17 role has been linked to an enhanced immune response producing eosinophilic pulmonary immunopathology in COVID-19 ,9. EosinAdditionally, at least one vaccine was given borderline to the recommended area in this case, which could have modified the antigenic depot exerted by alum adjuvants in the vaccine-site microenvironment, which in association with the effects described earlier, might have generated tertiary lymphoid structures (TLS). When analyzing the adverse events at the vaccination site, allergic relationship, Arthus phenomenon, and inflammation due to a TLS induction require a differential diagnosis. The first two hypersensitivity reactions commonly occur immediately, being ruled out in our case. Therefore, TLS reacting to successive cognate antigenic challenges in the OAS scenario is a plausible explanation. The stimulus caused by the infection may unleash the production of antibodies against the SARS-CoV-2 vaccine antigens, and they respond in the tertiary lymphoid follicles .Although an increase in the number of reports of skin manifestations has been documented in the context of SARS-CoV-2 infection and, more recently, after vaccination against COVID-19 , chance The OAS is a plausible phenomenon to consider in individuals immunized with inactivated vaccines and exposed secondarily to a wild virus with antigenic variations. Previous immunization status, vaccine type, site of injection, the timing between exposures, antigenic variations, and individual immune competency, among other factors, shall be considered in understanding the immune response against SARS-CoV-2. Consequently, there is an emerging need to evaluate further the long-term safety and effectiveness of COVID-19 vaccines against new variants and outcomes of natural infections.


Cardiovascular events such as myocarditis following mRNA COVID-19 vaccination are increasing. We present a 67-year-old postmenopausal woman with Takotsubo Syndrome and Graves’ disease after mRNA COVID-19 vaccination. She developed chest pain and shortness of breath one week after vaccination. An electrocardiogram revealed ST elevation in the precordial leads. Coronary angiography revealed the absence of obstructive coronary artery disease, and the left ventriculography showed a typical feature with apical ballooning. Laboratory workup showed the elevation of free T4 and thyrotropin receptor antibodies. It was presumed that Takotsubo Syndrome and Graves’ disease were probably related to the COVID-19 mRNA vaccination. The patient was treated with low-dose bisoprolol, diuretics, carbimazole, and steroid and discharged uneventfully. The mRNA COVID-19 vaccination is still safe and effective to defend against COVID-19 pandemic. However, clinicians should be aware of the possible cardiovascular adverse events other than myocarditis following vaccination. As of January 2022, the Coronavirus disease COVID-19) pandemic has affected about 352,700,000 patients worldwide. Vaccination against the disease has also been globally implemented with around nine to ten billion vaccine doses administrated. However, cardiovascular events after vaccine administration have been increasingly reported [9 pandemiTakotsubo syndrome is characterized by transient regional left ventricle systolic dysfunction. It mimics myocardial infarction but with the absence of angiographic evidence of obstructive coronary artery disease or acute plaque rupture. Usually, the regional wall motion abnormality extends beyond the territory perfused by a single epicardial coronary artery. Takotsubo syndrome is much more common in postmenopausal women, but its pathogenesis is not well elucidated. Vaccination can be a possible cause due to an immune inflammatory response. Herein, we present a case of Takotsubo Syndrome and Graves’ disease following mRNA COVID-19 vaccination.A 67-year-old postmenopausal female housekeeper presented to the emergency department with chest pain, palpitation, dyspnea on exertion, nausea, and diarrhea. The patient had no relevant past medical history and was not on any medication. One week earlier, she received her first dose of mRNA-1273 (Moderna) SARS-CoV-2 vaccination.On initial evaluation, the patient’s blood pressure was 106/69 mmHg. Her heart rate was 113 beats/min. She was afebrile. Her cardiac auscultation revealed regular and rapid heartbeats with grade 2 systolic murmurs over the left lower sternal border. Her respiratory and abdominal examinations were normal. An electrocardiogram (ECG) revealed sinus tachycardia and ST elevation in the precordial leads . A laborEchocardiography showed reduced left ventricular systolic function as an ejection fraction of 45% and akinesia of apex with preserved systolic wall motion in mid-ventricular and basal left ventricle of the heart . CoronarWe administered low-dose bisoprolol and diuretics. She also received carbimazole and steroids. On day 6, a follow-up ECG showed normal sinus rhythm. Repeat transthoracic echocardiography demonstrated the rapid recovery of apical ballooning of the left ventricle without any apical akinesia . ECG traOne week after discharge, the patient was followed up in the outpatient department. There was no chest pain, palpitation, or dyspnea on exertion. The laboratory results showed a thyroid-stimulating hormone level below 0.005 μIU/mL, and free T4 was above 77.7 ng/L. We prescribed bisoprolol and propylthiouracil (200 mg daily) for ongoing hyperthyroidism with Graves’ disease.The Phase III trial data for the mRNA-1273 SARS-CoV-2 vaccine reported no significant risk of cardiovascular reactions compared to a placebo [The present case describes a presentation of typical TTS and Graves’ disease after the first dose of mRNA-1273 SARS-CoV-2 vaccination in a postmenopausal woman. A systemic review has recruited 10 cases with TTS following COVID-19 vaccination between 1 January 2020 and 1 June 2022 . In contAlthough SARS-CoV-2 vaccination-associated myocarditis still could be a possible diagnosis in this case, the clinical features and typical apical ballooning in left ventriculography were strongly suggestive of TTS. The exact mechanisms of TTS after SARS-CoV-2 vaccination remains unclear. It may involve a systemic inflammatory reaction induced by vaccination with the overstimulation of sympathetic signaling. The mRNA-1273 SARS-CoV-2 vaccine encodes the stabilized prefusion SARS-CoV-2 spike protein. SARS-CoV-2 spike protein antibody might have cross-reacted with a variety of tissue antigens along with reacting to SARS-CoV-2 proteins . That coWe believe that SARS-CoV-2 vaccination-associated myocarditis and TTS are within the spectrum of immune response. The prognosis of TTS is distinct from that of myocarditis . LongituBoth TTS and Graves’ disease developed after SARS-CoV-2 vaccination in this case, though we could not exclude the causality between TTS and Graves’ disease. A multi-center observational study reported that 4 of 16 cases with TTS revealed subclinical or overt thyrotoxicosis . ThyrotoWe report a case in which the concomitant development of Takotsubo Syndrome and Graves’ Disease occurred after mRNA COVID-19 vaccination. The findings of the present report raise the possibility of an association between mRNA SARS-CoV-2 vaccination and TTS, particularly in postmenopausal women. Nevertheless, we still need further studies to establish the pathophysiologic mechanism of SARS-CoV-2 vaccination-associated complications.


Airborne pathogens, including SARS-CoV-2, are mainly contracted within the airway pathways, especially in the nasal epithelia, where inhaled air is mostly filtered in resting conditions. Mucosal immunity developing after SARS-CoV-2 infection or vaccination in this part of the body represents one of the most efficient deterrents for preventing viral infection. Nonetheless, the complete lack of such protection in SARS-CoV-2 naïve or seronegative subjects, the limited capacity of neutralizing new and highly mutated lineages, along with the progressive waning of mucosal immunity over time, lead the way to considering alternative strategies for constructing new walls that could stop or entrap the virus at the nasal mucosa surface, which is the area primarily colonized by the new SARS-CoV-2 Omicron sublineages. Among various infection preventive strategies, those based on generating physical barriers within the nose, aimed at impeding host cell penetration , or those preventing the association of SARS-CoV-2 with its cellular receptors  could be considered appealing perspectives. Provided that these agents are proven safe, comfortable, and compatible with daily life, we suggest prioritizing their usage in subjects at enhanced risk of contagion, during high-risk activities, as well as in patients more likely to develop severe forms of SARS-CoV-2 infection. Coronavirus Disease 2019 (COVID-19) is a life-threatening, highly infectious pathology that was first diagnosed at the end of 2019 in the Chinese city of Wuhan , and hasCOVID-19 is a typical respiratory infectious disease. Although it was earlier assumed that SARS-CoV-2 could only be transmitted with droplets, it has then become evident that contagion could also be driven by aerosols , and to The cell penetration of SARS-CoV-2 is mainly mediated by its spike protein, which binds effectively through its receptor-binding domain (RBD) to the human receptor angiotensin converting enzyme 2 (ACE2) upon proteolytic cleavage and activation by a number of human enzymes such as transmembrane serine protease 2 (TMPRSS2), furin, furin-like, trypsin-like and cathepsin proteases, among others . Cell enAltogether, these four mechanisms  as a result of a response to natural infection or vaccination . ConvincThe foremost issue is that such protection would not efficiently work in COVID-19 seronegative or SARS-CoV-2 naïve individuals, i.e., those who have not been previously infected by whatever type of SARS-CoV-2 variant. Even the protective role of preexisting immunity developed after exposure to other coronaviruses remains highly controversial, though mounting evidence suggests that it may not work efficiently to prevent SARS-CoV-2 infection .The second important caveat is that a former SARS-CoV-2 infection or a COVID-19 vaccination would not always provide sufficient protection when new and highly mutated lineages become prevalent. According to recent data published by Malato et al. , a previComparable considerations can be made for COVID-19 vaccination, whereby the administration of the former generation of “monovalent” vaccines , provides limited protection against infection by new Omicron sublineages. For example, the results of the recent meta-analysis published by Meggiolaro and colleagues  evidenceThe gradual waning of natural or vaccine-elicited immunity against SARS-CoV-2 is the third paradigmatic aspect that would contribute to limit the efficacy of mucosal immunity. Although we have previously seen that a previous SARS-CoV-2 infection or COVID-19 vaccination may be capable of triggering a robust mucosal immunity, the duration of such protection is limited over time. A study published by Isho and colleagues  showed tOwing to the important limitations characterizing anti-SARS-CoV-2 mucosal immunity highlighted in the previous paragraphs, development and implementation of “artificial” means of mucosal protection within the nose may be seen as an intriguing and appealing strategy for reducing the risk of SARS-CoV-2 (re)infection. Besides the administration of virucidal agents ,34, whicDe Vries et al. designed lipopeptide fusion inhibitors with the purpose of inhibiting membrane fusion between SARS-CoV-2 and the host cell . Daily iYip et al. tested the properties of a commercially available Chinese medicine nasal spray called Allergic Rhinitis Nose Drops (ARND) and containing 11 Chinese medicinal herbs . In a moFais et al. explored the efficacy of AM-301 nasal spray  against SARS-CoV-2 infection on a 3D-model of primary human nasal airway epithelium . AdminisThe clinical efficacy of a nasal spray containing 1% astodrimer sodium was tested by Paull et al. through intranasal administration in a K18-hACE2 mouse model . The adm2 was effective to prevent SARS-CoV-2 infection, but was also associated with almost total inhibition of virus release from infected cells.Bentley and Stanton investigated the effectiveness of three different hydroxypropyl methylcellulose (HPMC)-based nasal sprays  to prevent in vitro SARS-CoV-2 infection . Pre-tre2 IC, molecular testing reveled substantially lower intracellular SARS-CoV-2 RNA (between 3–5 orders of magnitude). In a separate investigation, Morokutti–Kurz et al. also demonstrated that 30 min pre-treatment with 10 μg/mL IC can efficiently inhibit by over five-fold the infection of Vero B4 cells with SARS-CoV-2 Spike Pseudotyped Lentivirus [Bovard and colleagues tested a nebulized iota-carrageenan (IC) nasal spray in a 3-D model of reconstituted nasal epithelium . At 48 antivirus . The effPyrć et al., investigated the effect of a positively charged polymer containing N-palmitoyl-N-monomethyl-N,N-dimethyl-N,N,N-trimethyl-6-O-glycolchitosan (GCPQ) in a model of human airway epithelial cells . The autZaderer et al., investigated the effect of ColdZyme mouth spray  administration to fully differentiated, polarized human epithelium cultured cells . Using SBalmforth and colleagues conducted a double blind, randomized, multi-centre clinical trial to explore the effectiveness of a prophylactic nasal spray for preventing SARS-CoV-2 infection . BrieflyComparable evidence emerged by the study of Figueroa and colleagues, using an IC-containing nasal spray, which was randomly administered with placebo to 394 Hospital workers . All stuPaolacci et al. administered a nasal spray containing α-cyclodextrin and hydroxytyrosol to 149 healthy volunteers at high risk of SARS-CoV-2 infection due to their occupation . During Preliminary but interesting evidence has also been provided that an anti-SARS-CoV-2 neutralizing monoclonal antibodies (mAbs)-based nasal spray may provide good protection against SARS-CoV-2 infection. In a small-scale clinical trial, Lin et al. explored the efficacy of a single nasal spray containing the anti-SARS-CoV-2 35B5 mAb, which was tested in 30 healthy volunteers . It was Recent evidence has been provided that the anticoagulant drug heparin displays pleiotropic antiviral properties, mostly by binding to the spike protein of SARS-CoV-2 and thus inhibiting host cell infection . It is hThe study protocol of a phase III, double-blind, randomized, single-centre clinical trial for establishing the efficacy of carrageenan-based nasal spray  for lowering the risk of SARS-CoV-2 infection was presented in September 2022 , but resAlthough widespread COVID-19 vaccination and gradual mitigation of SARS-CoV-2 pathogenicity have both synergically contributed to considerably attenuate the clinical impact of COVID-19 over time, this infectious disease is still causing a huge number of mild symptomatic infections , and continues to generate a significant burden of hospitalizations, especially those attributable to acute infection of unvaccinated and/or fragile individuals . The ideAirborne pathogens are mainly contracted within the respiratory pathways, especially through the nose, since airborne particles are mostly filtered within the nasal airway . The impAmong the various infection prevention strategies, those based on generation of physical barriers over nasal epithelial , or those hampering the association of the virus with its cellular receptors  are truly appealing perspectives. A vast array of compounds have been found effective for preventing SARS-CoV-2 infection in vitro or in animal models to date, whilst the number of sizeable randomized clinical trials is still very limited in humans. This evidence paves the way to planning comprehensive clinical trials, specifically aimed at testing, as well as comparing safety and effectiveness of these intranasally administered agents for preventing SARS-CoV-2 infection in humans before recommending their widespread administration.Nonetheless, provided that these agents will prove to be safe, comfortable, and compatible with daily life, we suggest that their usage should be prioritized in subjects at higher risk of contagion, during high-risk activities, as well as in patients at enhanced risk of developing severe COVID-19 illness .To this end, it is noteworthy that the German Society of Hospital Hygiene (DGKH) has recently endorsed the use of nasal sprays based on Carragelose, 3 times/daily in older people residing in elderly-care facilities or rehabilitation facilities and sharing communal activities, at family gatherings or professional meetings, schools, kindergartens and religious occasions . A clini


The coronavirus disease 2019 (COVID-19) pandemic is ongoing because of the repeated emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants, highlighting the importance of developing vaccines for variants that may continue to emerge. In the present review, we discuss humoral immune responses against SARS-CoV-2 with a focus on the antibody breadth to the variants. Recent studies have revealed that the temporal maturation of humoral immunity improves the antibody potency and breadth to the variants after infection or vaccination. Repeated vaccination or infection further accelerates the expansion of the antibody breadth. Memory B cells play a central role in this phenomenon, as the reactivity of the B-cell antigen receptor (BCR) on memory B cells is a key determinant of the antibody potency and breadth recalled upon vaccination or infection. The evolution of memory B cells remarkably improves the reactivity of BCR to antigenically distinct Omicron variants, to which the host has never been exposed. Thus, the evolution of memory B cells toward the variants constitutes an immunological basis for the durable and broad control of SARS-CoV-2 variants. Since the initial case was identified in December 2019, several vaccines that confer protection against SARS-CoV-2 have rapidly been developed and authorized for emergency use. COVID-19 vaccines efficiently confer protection against SARS-CoV-2 infection, hospitalization, and death [Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the causative virus of coronavirus disease 2019 (COVID-19), is currently threatening the lives of people worldwide. COVID-19 presents with mild to severe respiratory symptoms, and more than 500 million cases with more than 6 million deaths have been reported globally as of August 31, 2022  cells. Both long-lived B cells largely develop within germinal centers (GCs), where those with high-affinity B cell receptors (BCRs) to the antigens emerge with somatic hypermutations and are clonally selected , and late gradual (< 12 months) –62, rougThe antibody titers induced by COVID-19 vaccination have essentially the same kinetics. COVID-19 vaccines have been designed to activate B cells specific to the SARS-CoV-2 spike, as the spike is a target of protective antibodies, as described above. Several modalities of COVID-19 vaccines have been developed, such as mRNA, viral-vector, inactivated whole virus, and protein vaccines. Among these, immune responses induced by mRNA vaccines, which were approved for human use for the first time, have been intensely analyzed. Vaccination induces neutralizing antibodies to SARS-CoV-2 in the plasma, which reaches its peak at approximately 1 month after the 2nd vaccination and then declines. The neutralizing activity is low but still detectable 5 months after the 2nd dose –66. In lAlthough the antibody quantity decreases, plasma antibodies mature in terms of neutralizing potency and breadth over time. Neutralization potency, which has been reported to correlate with the prognosis of patients , increasAlthough plasma antibodies mature as mentioned above, the Omicron variants strongly escape vaccine-induced or convalescent serum polyclonal antibodies , 70–77. Meanwhile, current studies on vaccinated cohorts have revealed that humoral immunity improves reactivity to Omicron variants even after the 3rd vaccination by the ancestral Wuhan-based strain , 77–79. mem cells. As observed in influenza vaccine studies [mem cells specific to the spike are stably maintained or even increase in number from 1 to 6 months after the 2nd vaccination [mem cell numbers further increase after the 3rd vaccination [Exposure to the 3rd antigen with vaccination or breakthrough infection boosts the neutralizing titer to the same or higher level compared with the 2nd vaccination , 77–82.  studies , 7, Bmemcination –65. Thescination , 81.mem cells exhibit extended cross-reactivity to the virus variants compared to antibodies secreted by plasma cells. Even after the 2nd dose of the vaccine, 30–50% of Wuhan RBD-reactive Bmem cells are cross-reactive to the Omicron variants, and their numbers increase over time [mem cells retain BCRs which neutralize Omicron variants [mem cells, which differentiate into plasma cells secreting Omicron-neutralizing antibodies. Indeed, a fraction of Omicron-reactive Bmem cells exhibit an activated phenotype after the 3rd dose [Bver time , 81, 83.variants , 79, 83.mem cells to undergo maturation for a longer time to increase cross-reactivity before reactivation.The interval between antigen exposures is also a key factor in the cross-reactivity of humoral immune responses. A long interval between the 1st and 2nd doses of COVID-19 mRNA vaccines enhances the plasma neutralization titer against the Alpha, Beta, Gamma, and Delta variants compared with a short interval , 85. At mem cells have evolved their BCR repertoire to acquire broad reactivity to the variants over time after ancestral strain-based vaccination  skews antibody response towards the more specific HA head domain of the targeted strain . Concordmem cells, induced by COVID-19 vaccination, is indispensable for the development of universal vaccines that confer durable and broad protection against future SARS-CoV-2 variants and other pandemic viruses.Further investigation of the mechanism underlying the evolution of broadly reactive Bmem cells play a pivotal role in durable and broad humoral memory responses. Bmem cells evolve their reactivity to the variants over time, which are reactivated to secrete antibodies into circulation in response to subsequent antigen exposure. In summary, persistent maturation and repeated reactivation are the two key events involved in the evolution of humoral immune responses to SARS CoV-2. Although most COVID-19 vaccine studies have focused on responses to mRNA vaccines, some studies have reported that vaccines with other modalities induce Bmem cells of similar quality [mem cells induced by the Omicron-adapted vaccine booster is comparable to that of the ancestral booster in nonhuman primates [mem cells, which may be pivotal for the development of universal vaccines.The repeated emergence of SARS-CoV-2 variants escaping vaccine-induced immune responses has emphasized the importance of developing universal vaccines that induce protective immune memory toward a variety of variants, ideally including future escaping variants. Recent intense cohort studies on COVID-19-convalescent and vaccinated individuals have revealed that B quality , 110. Inprimates . Further


Since the mass vaccination for COVID-19, several case reports indicated the risk of autoimmune disease flare-ups after the vaccination. Among them, COVID-19 vaccine-induced glomerular diseases have drawn attention worldwide. The cases demonstrating the association between the mRNA vaccine and IgA nephropathy (IgAN) exacerbation had been noticed. Mostly mentioned, the flare-ups usually occurred after the second dose.We present a Taiwanese female with IgAN who developed gross hematuria within only six hours after the first dose of the Moderna vaccine.Six hours after the first dose of Moderna vaccine on 8 June 2021, the patient developed gross hematuria and significantly decreased urine output. All symptoms resolved spontaneously on the fifth day after the vaccination without any intervention. On the fourth day after the vaccination, the patient were able to back to her original condition.This was an intriguing case of IgAN flare-up following the first dose of mRNA-based COVID-19 vaccination. Numerous international reports demonstrated coronavirus disease 2019 (COVID-19) vaccines, especially for messenger ribonucleic acid (mRNA) vaccines, might trigger unexpected autoimmune responses and even the sudden onset of glomerular diseases.,4 However, among the previously reported cases collected by Bomback et al, the IgA nephropathy (IgAN) exacerbation usually occurred after the second dose and only 2 cases, which were reported by Perrin et al, occurred after the first dose. In this case report, we have a Taiwanese female with IgAN who developed gross hematuria within only 6 hours after the first dose of the Moderna vaccine.The COVID-19 vaccine-induced glomerular disease have drawn attention worldwide.A 40-year-old Taiwanese female with IgAN was proven by renal biopsy ten years ago. She has been under stable condition with medication of oral steroids and angiotensin II receptor antagonists ever since. She denied hypertension or hematuria within 3 months. She only stopped taking steroids for 7 days before the administration of the Moderna vaccine. 6 hours after the first dose of Moderna vaccine on June 8, 2021, she developed gross hematuria and significantly decreased urine output. She also reported nonspecific side effects including mild febrile (37.8°C), headache, stiff neck, and fatigue. All symptoms resolved spontaneously on the fifth day after the vaccination without any intervention.Thirty-eight days after the first dose, the patient received her second dose of the Moderna vaccine. This time, her symptoms of gross hematuria and fever episode over 38.6°C occurred 1 day later. The fever persisted only for 1 day and the gross hematuria accompanied by decreased urine output lasted for 4 days. She came to our outpatient department and followed up on her laboratory data of urine test on the same day of the second dose and 1 day after the development of gross hematuria, respectively. Hematuria deteriorated with an increase in urine red blood cells by a random urine test. On the fourth day after the vaccination, she had already been back to her original condition.,5 Exacerbation of immune-mediated glomerular diseases following the COVID-19 vaccine, especially for the mRNA-based vaccine (either Pfizer-BioNTech or Moderna) has been thoroughly reported by Bomback et al, with the collection of 6 reports including 10 cases of IgA nephropathy presenting as gross hematuria following the mRNA-based vaccine. Among these patients, 6 patients received Moderna vaccination while the other 4 received Pfizer. Most of these cases occurred gross hematuria within 2 days after the second dose. However, there were 2 cases reported by Perrin et al, in which one received Pfizer and the other take Moderna, to develop gross hematuria soon after the first dose.With mass vaccination for COVID-19, several case reports put the risk of autoimmune disease flare-ups after the vaccination in the spotlight. Previous studies have shown the potential link between influenza vaccination with flare-up of IgA nephropathy. Cases of reactivation of IgA vasculitis occurring after COVID-19 vaccination had also been reported. Furthermore, Rahim et al reported a recurrence of IgA nephropathy after COVID-19 vaccination in the same patient who had a similar episode 2 years earlier after receiving a recombinant zoster vaccine (Shingrix).IgAN is the most common glomerulonephritis caused by mesangial deposition of autoantibodies of IgA. Taken together, these results might suggest a link between the virus antigen-containing COVID-19 vaccine and an increase in anti-SARS-CoV-2 spike IgA. While coincidence cannot be excluded, the close temporal association within days between vaccination may suggest a potential pathogenetic linkage.All above studies indicated vaccine-induced IgA response is not uncommon and could take place at any vulnerable targets. To the best of our knowledge, severe COVID-19 illnesses can trigger an IgA response in the bronchial mucosa. but still, the mechanism remains unclear. Whether activation of autoreactive B cells after COVID-19 vaccination causes the preexisting or de novo mobilization of autoreactive B cells producing IgA (or both) remains to be established. According to the studies by Bomback et al, T cells are assumed to be the key mediators of COVID-19 vaccine-induced glomerular disease because these responses are similar to the infection of SARS-Cov-2 itself to cause activation of diverse autoimmune and alloimmune diseases affecting the kidney. Kudose et al also assumed it is a systemic cytokine-mediated flare, possibly via induction of heightened IgA1 antiglycan immune responses. Rahim et al suggested it is related to the delayed-type hypersensitivity owing to the response usually occurring after the second dose. But it became doubtful due to more and more reports of events occurring within 24 hours after the first dose, including 2 cases reported by Perrin et al and our case. Only one thing in common is all these cases highlight that in the absence of intervention, COVID-19 vaccine-associated IgAN and immunoglobulin A–associated vasculitis flairs may improve spontaneously.There are several debates concerning the link between autoimmune disease flares after COVID-19 vaccination,Without a doubt, these reports should not lead to vaccine hesitation as the benefits of vaccination strongly outweigh the potential risks. But they bring caution on patients with IgAN should be monitored closely following the COVID-19 vaccine. If the adverse events after the vaccination could not resolve, shifting to another vaccine should be taken into consideration. Further discussion of how to incorporate active glomerular disease after the COVID-19 vaccination into a current therapeutic plan for these patients is an issue.To sum up, we present an intriguing case of IgAN flare-up following the first dose of mRNA-based COVID-19 vaccination. The optimal therapy of COVID-19 vaccine-associated glomerular diseases including IgAN remains unclear. Our patient’s disease has been in remission with normal renal function and blood pressure after a complete 2-dose of vaccination. Nevertheless, further studies are required to investigate the exact pathogenesis of COVID-19 vaccine-induced IgAN. Is it necessary to shift to a different type of vaccine, such as traditional inactivated viral and adjuvanted protein vaccines, after the adverse events of the first dose? Is it possible for those adverse events processing speed up the deterioration of a patient’s renal function eventually? How to incorporate active glomerular disease after the COVID-19 vaccination into a current therapeutic plan for these patients has become an important issue.Conceptualization: Po-Hung Tseng, Shu-Han Chuang, Hung-Jen Shih, Sheng-Hsien Huang.Data curation: Po-Hung Tseng, Shu-Han Chuang.Formal analysis: Po-Hung Tseng.Funding acquisition: Hung-Jen Shih.Investigation: Po-Hung Tseng, Shu-Han Chuang.Methodology: Po-Hung Tseng, Shu-Han Chuang, Hung-Jen Shih, Hung-Jen Shih, Chin-Pao Chang.Project administration: Shu-Han Chuang.Resources: Yueh Pan, Hung-Jen Shih, Sheng-Hsien Huang.Software: Hung-Jen Shih.Supervision: Yueh Pan, Hung-Jen Shih, Chin-Pao Chang, Sheng-Hsien Huang.Validation: Po-Hung Tseng, Yueh Pan, Hung-Jen Shih.Visualization: Po-Hung Tseng, Shu-Han Chuang.Writing—original draft: Po-Hung Tseng, Shu-Han Chuang.Writing—review and editing: Po-Hung Tseng, Shu-Han Chuang, Yueh Pan, Hung-Jen Shih, Chin-Pao Chang, Sheng-Hsien Huang.


Viruses share many attributes in common with extracellular vesicles (EVs). The cellular machinery that is used for EV production, packaging of substrates and secretion is also commonly manipulated by viruses for replication, assembly and egress. Viruses can increase EV production or manipulate EVs to spread their own genetic material or proteins, while EVs can play a key role in regulating viral infections by transporting immunomodulatory molecules and viral antigens to initiate antiviral immune responses. Ultimately, the interactions between EVs and viruses are highly interconnected, which has led to interesting discoveries in their associated roles in the progression of different diseases, as well as the new promise of combinational therapeutics. In this review, we summarize the relationships between viruses and EVs and discuss major developments from the past five years in the engineering of virus-EV therapies. Multicellular organisms rely on intercellular communication to regulate many aspects of their physiology. It defines environmental niches that regulate cell growth and behavior, and it is essential for collective defense against host pathogens. The majority of intercellular communication is mediated via the transportation of bioactive molecules, such as proteins, nucleic acids, metabolites and lipids between cells ,2. PassaEVs are derived from multivesicular bodies (exosomes), the plasma membrane (microvesicles)  or otherEVs can stimulate immune responses against pathogens and tumors by transporting antigens and immune-stimulating factors, maintain cellular homeostasis by excreting harmful components, such as nuclear DNA in the cytoplasm ,10 and pEVs also have a deeply interwoven relationship with viruses ,17. ViruIn this review, we focus on the past decade of discoveries regarding the intertwined interactions of EVs and viruses. We begin by describing the biosynthetic pathway of EVs and discuss the ways in which viruses also utilize or manipulate these pathways for their own benefit. We examine the role of EVs in virus transportation and pathogenesis and discuss the relationships between EVs, viruses and the immune system. Lastly, we focus on the role of EVs in virus-associated disorders and discuss promising approaches for combining viruses and EVs as therapeutics, with a specific focus on cancer-related therapies.EVs are secreted from intraluminal vesicles (ILVs) contained in cellular multivesicular bodies (MVBs). The biosynthesis pathway begins with the maturation of endosomes into MVBs when membrane cargos recruit factors inducing the internal budding of the endosomal membrane and the formation of ILVs. These cargos can include proteins, nucleic acids, metabolites and lipids ,2. CargoPackaging of proteins into EVs depends on post-translational modifications—mostly ubiquitination and, in some cases, sumoylation —as well Non-classical sorting of proteins into EVs can also be achieved through ESCRT-independent pathways. ALIX can bind and transport non-ubiquitinated cargos inside MVBs, bypassing ESCRT-0, -I and -II . ALIX isIn contrast to protein cargos, the packaging of nucleic acids into EVs is less well understood but appears to depend on a variety of mechanisms. MicroRNAs (miRNAs) are packaged into EVs according to sequence motifs that are recognized by RNA-binding proteins, such as heterogeneous nuclear ribonucleoprotein A2B1 (hnRNPA2B1), synaptotagmin-binding cytoplasmic RNA-interaction protein (SYNCRIP), Argonaute2, Y-Box binding protein 1 (YBX-1), MEX3C, major vault protein (MVP) and La protein. These RNA-binding proteins then interact with MVBs membrane proteins, such as caveolin-1 (Cav-1), n-SMase-2 and VPS4, to facilitate the sorting of miRNAs into MVBs . For insTetraspanin small transmembrane proteins also have a significant role in the EV biosynthesis pathway. Tetraspanins are known for their role in mediating many physiological processes, such as cell migration, adhesion and signaling. Various tetraspanins are located inside MVBs, such as CD63, CD9 and CD81 . They suOnce cargos are packaged into ILVs, MVBs either fuse with lysosomes to be degraded or move to the plasma membrane, where they fuse and release ILVs to the surrounding environment through processes regulated by the soluble NSF attachment protein receptor (SNARE) complex . These sSecreted EVs have been found in many body fluids, such as blood, urine, saliva, tears and semen , and theFinally, EVs are internalized by specific recipient cells through EV-associated transmembrane molecules, such as extracellular matrix proteins , integrins, proteoglycans, lectins, glycolipids, phosphatidylserine (PS) and tetraspanins . EV bindViruses have adapted strategies to hijack EV biosynthesis machinery to aid in all stages of their life cycles. Both RNA and DNA viruses can acquire their viral envelope by using the ESCRT complex and Rab-GTPases. Enveloped viruses can acquire their viral envelope while entering MVBs, as observed for the human cytomegalovirus (HCMV) , Human hNon-enveloped viruses also hijack the ESCRT complex and secrete their virions or viral genome inside EVs, as observed with the Hepatitis A virus (HAV), Hepatitis E virus (HEV), Enterovirus 71 (EV71) and Bluetongue virus ,48,49,50Moreover, some viruses also hijack ESCRT and Rab-GTPase proteins to fulfill their replication cycle without entering MVBs. First, viruses recruit the ESCRT complex to acquire their viral envelope. For instance, the human immunodeficient virus (HIV) recruits ESCRT-I, -II,-III, ALIX and VPS4 to bud from the plasma membrane  Figure B2. SimilEVs have a prominent role in the viral spread and the infectivity of healthy cells. Many viruses pass infectious particles or genetic material to surrounding cells through EVs. This can provide both enveloped and non-enveloped viruses a cloak that hides them from immune surveillance while allowing virus dissemination, as mentioned above for HCMV, HHV-6, SARS-CoV-2, DV, HBV, HAV, HEV, EV71 and Bluetongue virus. For instance, DV-EVs containing virus-like particles transfer the virus to non-infected mosquito cells . MoreoveSome viruses alter the lipid and protein composition of EVs to increase binding and uptake into new cells. For instance, HAV, enteroviruses, EBV, Zika virus  and HIV The unique composition of EVs can also permit virus uptake in a receptor-independent fashion and alter viral tropism, as observed with EV71, HCV, HBV and severe fever with the thrombocytopenia syndrome (SFTS) virus. EV71-EVs containing the viral genome are integrated by recipient cells in a receptor-independent manner, as the knockdown of the viral receptor did not infer with virus uptake by recipient cells . SimilarEVs also increase cell susceptibility to virus infection by transporting virus-targeted host receptors to cells that do not normally express them at high levels. For instance, EVs transport SARS-CoV-2 viral receptor ACE2  and tetrVirus-EVs containing viral RNA can also facilitate the infection of recipient cells. HIV-infected monocyte-derived macrophages secrete microvesicles and EVs containing HIV-1 RNA, potentially HIV reverse transcriptase, host cytokines (interleukin (IL) -3, -4, -8, -17, leptin, tumor necrosis factor α (TNF-α)), antigen-presenting receptors and adhesion factors  and have specific lipid composition (PS). These HIV-1-EVs control infection of recipient T cells through CD4-independent clathrin-mediated endocytosis . NarayanVirus-EVs enhance viral particle transfer by packaging multiple viral particles. This packaging enables the transfer of multiple viral genomes and proteins to recipient cells, which improves survival against the cell antiviral immune response and increases overall infectivity ,75. ThisAedes albopictus and Aedes aegypti mosquitos) to mammalian hosts inside EVs. DV-infected mosquito cells secrete EVs containing both viral RNA  and proteins  that promote active infection in mosquito cells, mouse cells, human skin keratinocytes and blood endothelial cells [The encapsulation of virus particles in EVs increases the rate of virus transmission by providing safe transport between hosts. For instance, when noroviruses and rotaviruses are protected inside EVs, they are not degraded while transiting through stool and the gastrointestinal tracts. This improves the active fecal–oral transmission of these viruses . Anotheral cells . Interesal cells . Zika vial cells   elements such as Alu and LINE-1, which promote oncogenesis in recipient cells ,80. For In the previous section, we discussed the strategies adopted by viruses to hijack EV biosynthesis machinery to aid in their replication, egress and spread. In this section, we explore the relationship between EVs, viruses and the immune system and summarize the antiviral and host immunomodulatory signaling that is mediated by EVs in the context of viral infections .Virus-EVs containing either host or viral proteins or miRNAs can enhance all stages of the antiviral innate and adaptive immune response. Virus-EVs transport host miRNAs, molecules and proteins resulting in the activation of innate antiviral immune responses in uninfected recipient cells. For instance, in the late stages of IAV infection, when cells are undergoing apoptosis, the host Y5 non-coding RNA is degraded into miRNAs that are transported via EVs to uninfected recipient cells, where they induce IFN antiviral responses . During Secondly, virus-EVs stimulate the secretion of chemokines and pro-inflammatory cytokines by recipient cells, which induce activation and recruitment of immune cells to the infection site. As mentioned above, the IAV-EVs transporting IFIT and IFITM activate the secretion of IL-6, MCP-1 and TNF . RespiraLastly, virus-EVs can stimulate robust adaptive immune responses. As mentioned before, IAV-EVs and DV-EVs transporting IFITM could mediate this activation ,87,88. MThe antiviral immune response is also activated by EVs secreted by non-infected immune cells. EVs can protect the non-infected recipient cells from viral infection. For instance, during HBV infection, activated macrophages secrete IFN-α containing EVs, which are integrated by non-infected hepatocytes through TIM-1 (the host receptor for HAV), thus protecting cells against HBV infection . EVs alsSimilar to how viruses adapted the means to hijack host machinery and enhance their own replication, viruses have also found ways to suppress the antiviral immune responses by using EVs to transport host and viral immunomodulatory cargos from infected to uninfected cells. Some virus-EVs contain host miRNAs that inhibit the interferon (IFN) signaling in recipient cells, making them permissive to viral infection. EV71-EVs are enriched in host miR-146a, which represses the expression of signal transducer and activator of transcription 1 (STAT1), TNF receptor-associated factor 6 (TRAF6) and Interleukin 1 receptor-associated kinase 1 (IRAK1) and eventually suppresses the IFN-1 response in the recipient cells. Consequently, IFN-1-stimulated gene factors such as BST-2/tetherin are also inhibited. EV71-EVs integration by recipient cells not only enhances viral replication but also EV secretion by inhibiting the IFN antiviral immune response in the recipient cells since BST-2/tetherin is an inhibitor of Rab27a-dependent EV secretion . SimilarViruses have also developed strategies to evade the innate immune responses through the modulation of the EV biosynthesis pathways. Viruses, such as the Tomato bushy stunt tombusvirus, hijack the ESCRT machinery to form replication centers protected from immune surveillance ,54. PoliAs noted earlier, virus-EVs are often enriched in PS, which not only improves uptake into recipient cells, but also plays a role in the modulation of the immune response . PS is aVirus-EVs also transport viral proteins that lower immune recognition through antigen presentation. For instance, HSV-1-EVs contain the viral envelope glycoprotein B (gB), which downregulates major histocompatibility complex class II (MHC-II) expression and promotes MHC-II uptake and secretion in EVs, thereby reducing the ability of recipient cells to present viral antigens and initiate immune responses . SimilarThe transport of infectious material through EVs also provides an avenue to escape recognition by neutralizing antibodies. For example, HCV-EVs transporting viral RNA and proteins are only partially recognized by neutralizing IgG antibodies . SimilarFinally, some viruses-EVs transporting viral proteins permit active infection or apoptosis of immune cells. As mentioned previously, HBV-EVs lead to an active infection of the recipient NK cells despite the tropism of HBV for hepatic cells, which suppresses the antiviral immune response and promotes chronic infection . MoreoveThe interactions between viruses, EVs and the immune system can sometimes result in overstimulation and lead to chronic inflammatory disease. Zika virus-EVs secreted by mosquito cells that contain viral RNA, and the E-protein are integrated by naïve human endothelial vascular cells resulting in a pro-inflammatory and pro-coagulant cellular state . These ZSome virus-Evs transmit pathogen-associated molecular patterns (PAMPS), which activate PRRs in immune cells and promote the secretion of pro-inflammatory cytokines, eventually leading to chronic inflammation, as observed for HIV-1, Epstein–Barr virus (EBV), Ebola virus and HTLV-1. HIV-Evs containing TAR RNA and miRNA not only favor T cell infection, as mentioned above, but also support chronic inflammation ,102. BerVirus-EV-induced activation of PRRs in immune cells can also be achieved indirectly. HIV-EVs containing the Nef protein downregulates the ATP-binding cassette transporter type A1 (ABCA1) in recipient macrophages, leading to a cascade of cellular modifications. Downregulation of ABCA1 reduces cholesterol efflux, resulting in the inactivation of Cdc42, decreased actin polymerization and increased abundance of lipid rafts. This influences TLR-4 concentration in lipid rafts, potentiating ERK1/2 signaling and activating NLRP3 inflammasome and interleukin-1β (IL-1β) responses  Figure A.Finally, uninfected cells can also secrete EVs that promote chronic inflammation. During DV infection, IL-1β is secreted as part of the antiviral immune response, which promotes the secretion of EVs by non-infected platelets. These EVs containing various host proteins are detected by macrophages and neutrophils through CLEC5A and TLR-2, respectively, resulting in the secretion of inflammatory cytokines and activation of the neutrophile extracellular trap (NET), which lead to systemic inflammation, tissue damage, and vascular permeability  Figure A.HBV and HCV-EVs secreted by infected hepatocytes are integrated by hepatic stellate cells (HSCs), which cannot usually be infected by HBV and HCV, thus exasperating the disease to other cell types. HBV-infected cells express the oncogenic viral protein HBx, which induces EV biogenesis by interacting with the cellular CD9, CD81 and neutral sphingomyelinase 2 (N-SMase). The HBx protein and mRNA are secreted in HBV-EVs, then integrated by HSCs in which HBx stimulates cell proliferation, supporting HBV-associated liver disease . LikewisSuch viruses as the human papillomavirus (HPV), EBV, HIV-1 and HTLV-1 are widely known for their ability to cause cancer. Not surprisingly, EVs produced from cells infected with these viruses can also have oncogenic properties. Virus-EVs can aid in the development of tumors by transferring viral oncogenes or RNAs. For instance, HPV type 16 (HPV-16) infection increases the expression of more than 50 host microRNA (miRNA) in infected cells, ten of which are upregulated and selectively packaged into EVs as a result of the E6/E7 oncogene. These miRNAs inhibit apoptosis and senescence while inducing proliferation in both the infected and recipient cells ,111. In Virus-EVs can also cause cancer cells to adopt a more aggressive and invasive phenotype. For example, the EBV-infected cells express the viral latent membrane protein-1 (LMP-1), a constitutively active signaling protein mimicking CD40. LMP-1 can induce EV secretion through a CD63-dependent mechanism . In BurkCancer escape from immune surveillance can also be facilitated by the transportation of viral regulatory factors in virus-EVs. HPV-16-infected keratinocytes expressing E7 oncogene secrete plasma-membrane-derived EVs that inhibit the adaptive immune response. These HPV-16-EVs inhibit CD40 expression and IL-12 p40 subunit secretion by recipient Langerhans DCs in the epidermis, which reduces antigen presentation and activation of anti-HPV cytotoxic T cells . The LymUltimately, the relationships between EVs and viruses play an integral role in the spread and progression of multiple pathologies. A key contributor to these pathologies is the immune system. Virus-EVs act as potent immunomodulatory molecules that can serve to either activate or suppress the immune system. This can consequently lead to an onset of chronic inflammatory diseases or permit immune evasion and escape of other pathologies, such as cancer. However, the alternative can also be true. Virus-EV interactions can be a powerful immunotherapy strategy to target a variety of disorders.Both viruses and EVs have a wide range of therapeutic applications. EVs have reached clinical trials for a wide variety of different illnesses—from cancer ,122,123 In this respect, combinational virus-EV therapeutics have many promising characteristics that overcome the weaknesses of their individual counterparts. As we have discussed previously, EVs can promote the cell-to-cell spread of viral particles and genomes, permit infection of cells with low expression of the viral receptor and provide protection against immune responses. Hence, the viral therapies that spread via EVs, either naturally or by design, have the potential of reaching more target cells without clearance from the immune system. Viruses can also help overcome the low loading of therapeutic payloads in EVs by propagating in recipient cells and delivering prolonged expression of therapeutic transgenes. Virus infection is also known to promote EV production and secretion in many cells, so there is added potential for using engineered viruses with selectivity for different tissues to generate EVs with therapeutic payloads. In the following sections, we will discuss some of the recent developments using virus-EV strategies to treat different diseases .Adeno-associated virus (AAV) is a powerful tool for gene delivery that has been widely studied in clinical trials for therapies targeting the brain, spinal cord, liver and muscle . DespiteTo overcome these issues, AAVs can be targeted to EVs to provide protection against neutralizing antibodies while keeping the ability to target specific tissues and limiting off-target infection. Maguire et al. showed that the cells transfected with DNA for producing AAV particles also secreted EVs containing the fully functional AAV  . TransfeAAV-EVs have a high potential for the treatment of neurological disorders. AAV-EVs  avoid immune neutralization by host antibodies in vitro and in vivo  and are AAV-EVs also showed promising results against hemophilia B genetic disease. Meliani et al. transfected HEK293 cells with AAV8 plasmids encoding the human coagulation factor IX and later harvested AAV-EVs presenting TSG101 and CD9 specific markers. After intravenous injection, these AAV-EVs are protected from antibody recognition and successfully target and transduce liver cells, brain cells and skeletal muscle cells in mice. AAV-EVs could thus improve gene therapy safety and efficiency in liver genetic disease . FinallyThe intertwined relationship between EVs, viruses and activation of the immune response has a promising role in the development of oncolytic viral therapy. Oncolytic viruses (OVs) selectively infect tumor cells causing cell death, the release of tumor and viral antigens and activation of antiviral and anti-tumoral immune responses. Moreover, OVs can be genetically modified to express factors increasing the immune response, such as immune checkpoint inhibitors , cytokines  or T cell engagers ,151. ManThere are many OVs, such as VSV , HSV-1  and NDV Several studies have shown that EVs increase the spreading of oncolytic adenoviruses, enhance the immune response, and help target metastatic niches. Indeed, Ad5/3-D24-GMCSF leads to the secretion of EVs containing viral proteins and genomes that actively infect the recipient tumor cells, thus enhancing viral spreading  Figure B1. MoreoA major challenge with OV therapy is the presence of pre-existing anti-viral immunity. OVs are also often delivered via intratumoral injection, which limits their ability to target metastatic tumors. Whereas, intravenous injection of OVs faces issues with viral clearance in the vascular system and peripheral tissues by immune cells, neutralizing antibodies and other molecules . HoweverCancer cells can be used as OV-EVs producers, which will have a natural tropism for cancer cells  Figure C1. The OSimilarly, plasma membrane-derived EVs can also carry chemotherapeutic drugs (cisplatin)  and oncoOV-EVs can also be engineered using EV-mimetic nanovesicle drug loading technology, which allows the production of a much higher quantity of EVs containing the drug of interest. Producer cells are first transduced to express a drug and then passed step-by-step through smaller and smaller nanosized filters, eventually forcing the formation of EV-mimetic nanovesicles containing the drug  Figure C3. ZhangIn the past, our group has shown that it is possible to design and encode artificial miRNAs (amiRNAs) into oncolytic VSVΔ51 that get packaged into EVs and transmitted to surrounding uninfected cells . This stWe demonstrated that EVs produced by cancer cells upon infection with VSVΔ51 encoding an amiRNA targeting ARID1A, but not a non-targeting amiRNA, can sensitize uninfected cells to GSK126 . ThroughEV-mediated transfer of proteins, lipids or nucleic acids to recipient cells is essential in physiologic cell-to-cell communication, but also in viral infection and immune modulation. Both RNA and DNA viruses, whether they are enveloped or not, utilize the EVs pathway to secrete their viral particles, proteins and nucleic acids, but also to secrete host elements that benefit viral infection. Virus-EVs enhance viral infection by transferring viral elements to recipient cells and modulating their response toward the virus. Moreover, virus-EVs modulate many aspects of the immune system, leading to both antiviral and pro-viral responses that can drive a variety of autoimmune and chronic inflammatory diseases and cancers. EVs and viruses are both important vectors used in many therapies. In the field of oncology, EVs-based therapies and virotherapies are both being developed. Because of how virus-EVs can enhance viral spread and activate the immune response, the combination of OVs and EVs could potentiate these cancer therapies. Indeed, packaging of OV into EVs provides safe delivery and better uptake of the OV to the tumor, which enhances the overall efficacy of the virotherapy. Despite still being in development, this strategy has great potential in cancer therapy.


Cerebral ischemia refers to the symptom of insufficient blood supply to the brain. Cells of many different origins participate in the process of repairing damage after cerebral ischemia occurs, in which exosomes secreted by the cells play important roles. For their characteristics, such as small molecular weight, low immunogenicity, and the easy penetration of the blood–brain barrier (BBB), exosomes can mediate cell-to-cell communication under pathophysiological conditions. In cerebral ischemia, exosomes can reduce neuronal damage and improve the brain microenvironment by regulating inflammation, mediating pyroptosis, promoting axonal growth, and stimulating vascular remodeling. Therefore, exosomes have an excellent application prospect for the treatment of cerebral ischemia. This article reviews the roles and mechanisms of exosomes from different sources in cerebral ischemia and provides new ideas for the prevention and treatment of cerebral ischemia. Cerebral ischemia is a life-threatening cerebral vascular disease . As one Extracellular vesicles (EVs) are diverse nanoscale membrane vesicles with diameters ranging from 30 to 1000 nm that are released by cells . They arMost types of cells in the CNS and peripheral system can secrete exosomes, including microglia, neurons, astrocytes , and endNeural stem cells (NSCs) are a group of ectodermal progenitor cells that can differentiate into committed neural sub-types such as neurons, astrocytes, or oligodendrocytes . Recent As the unique immune cells of the CNS, microglia play essential roles in many cerebral physiological processes, including regulating immune responses and promoting neuronal survival . Brain dThe astrocyte is the largest type of glial cell. Astrocytes are widely distributed throughout the CNS. Their unique morphology can provide structural support to neurons. Furthermore, astrocytes have other functions, such as regulating neuroinflammatory responses, modulating synaptic activity, supplying energy to neurons, and maintaining the blood–brain barrier (BBB) . AstrocyExosomes derived from cerebral endothelial cells play essential roles in protecting neurons under a hypoxia state. Endothelial cell-derived exosomes (EC-exos) can protect neurons against I/R injury through the promotion of cell growth, migration, and invasion, and the inhibition of the apoptosis of SH-SY5Y nerve cells . MiRNA-1Based on above experiments and researches, it can be concluded that exosomes from CNS alleviate damage after cerebral ischemia and promote recovery through reducing inflammation, suppressing apoptosis, as well as enhancing neurogenesis and angiogenesis . Mesenchymal stem cells, considered as “sentinel and safe-guards of injury” , have beBone mesenchymal stem-cell-derived exosomes (BMSC) are a group of unique stem cells with great differential potential. The protective effect of BMSC-derived exosomes (BMSC-exos) is found to be linked to the inhibition of apoptosis in different studies. Various miRNAs can function in an ischemia-hypoxic brain environment taking BMSC-exos as carriers. BMSC-exo-miR-26a-5p mimics can dramatically reduce protein kinase CDK6 levels in BV-2 cells after oxygen–glucose deprivation/reoxygenation (OGD/R) treatment, as well as in brain tissues of the middle cerebral artery occlusion–reperfusion (MCAO/R) model. The downregulation of CDK6 mediated by exosomal miR-26a-5p might contribute to the attenuation of I/R injury in vivo, and the inhibition of microglia apoptosis in vitro . By down6 or 2 × 107 BMSC equivalents/kg) to young and aged mice. The results showed that sEVs at both doses promoted periinfarct angiogenesis in both types of rats. In addition, low-dose sEVs enhanced neurogenesis in the subventricular zone [Apart from the miRNA mentioned above, BMSC-exos are also found to function in protecting neurons through other mechanisms and pathways. It has been proven that the nucleotide-binding domain and leucine-rich repeat family protein 3 (NLRP3) inflammasome plays an essential role in neuronal damage induced by I/R . BMSC-exlar zone . Additiolar zone . Molecullar zone . This relar zone . BMSC-exlar zone . Buyang lar zone . Anotherlar zone .+ exosomes in poststroke might represent a novel treatment for stroke [Pigment epithelium-derived factor (PEDF) is a multifunctional protein that has neurotrophic, anti-inflammatory, and neuroprotective functions. Researchers have found that exosomes released by adipose-derived mesenchymal stem cells (ADSCs-exos) with increased PEDF content could further protect neurons against OGD-induced apoptosis in cerebral I/R through the activation of autophagy and the suppression of neuronal apoptosis. It is also observed that blocking autophagy could lead to a reduction in the effect of PEDF-containing exosomes . Exosomer stroke .Multiple studies have shown that human umbilical cord mesenchymal stem cell-derived exosomes (hUMSC-exos) can reduce microglia-mediated neuroinflammation, protecting against brain injury. Exosomes derived from umbilical cord mesenchymal stem cells can interfere with the Toll-like receptor 4 (TLR4) signaling of BV-2 microglia, which contributes to reducing neuroinflammation induced by microglia in perinatal brain injury . HUMSC-eResearch indicated that plasma-derived exosomes (PLA-exos) enriched with heat shock protein 70 (HSP70) showed better brain targeting and treatment effects. PLA-exos with inherited HSP70 can facilitate the migration and diapedesis of PLA-exos through an interaction with endothelial TLR4. Furthermore, PLA-exos-meditated HSP70 delivery can activate tight junction protein (TJP), which leads to the inhibition of mitochondria-meditated neuron cell apoptosis, the suppression of ROS accumulation, and the alleviation of BBB damage . EdaravoExosomes from remote ischemic preconditioning (RIPC) serum show neuroprotective effects via the upregulation of miRNA-126, thus reducing the expression of DNMTs3B in neurons and raising OGD tolerance in SH-SY5Y cells . HoweverAccording to the related studies we can summarize that the effects of exosomes from peripheral nervous system in brain ischemic stroke are mainly exerted through facilitating brain tissues functional recovery, inflammation reduction, as well as inhibition of cells pyroptosis and apoptosis .Apart from the above-mentioned sources, macrophages and some other cells can also react to brain injury via exosomes.Through the regulation of microglial polarity from phenotype M1 to anti-inflammatory phenotype M2, LPS-stimulated macrophage-derived exosomes have functional improvements and neuroprotection effects after ischemic stroke . MacrophHuman umbilical endothelial cells (HUECs) can reduce the apoptosis of neurons under OGD in a HUEC-neuron coculture assay. Researchers also found that Cav-1 upregulated by neurons during ischemia stroke can increase the neuron intake of extracellular vesicles derived from an endothelial cell, which attenuates apoptosis via exosomal miR-1290 and protects neurons . ExosomeTherefore, we can conclude that exosomes from other sources also have protective effects in cerebral ischemic stroke, and possess great potential as therapy of brain ischemia .It can be concluded from the above review that exosomes derived from diverse sources targeting at different brain cells  through


Anti-programmed death-1 (PD-1)/programmed death ligand 1 (PD-L1) immune checkpoint inhibitors (ICIs), combined with bevacizumab and platinum-based chemotherapy, have shown promising efficacy in treating metastatic non-squamous cell lung cancer in phase 3 clinical trials. However, drug-induced nephrotoxicity is an uncommon but threatening adverse effect when using this combination therapy, and should be evaluated and managed carefully. Here, we present two patients experiencing late-onset asymptomatic heavy proteinuria during the clinical trial. Kidney biopsies performed finally identified bevacizumab-induced thrombotic microangiopathy (TMA), and the proteinuria was decreased after discontinuing bevacizumab permanently. Our report suggests that a kidney biopsy is needed for those receiving ICIs in combination with bevacizumab and chemotherapy and experiencing nephrotoxicity such as heavy proteinuria. Drug-induced nephrotoxicity is an uncommon but threatening adverse effect when using ICIs combined with bevacizumab and chemotherapy.A kidney biopsy is indicated for metastatic non-squamous cell lung cancer patients experiencing nephrotoxicity during this combination therapy.The therapy of nivolumab in combination with carboplatin, paclitaxel, and bevacizumab was shown to be promising in the first-line treatment of advanced non-squamous, non-small-cell lung cancer (NSCLC) in a previous pivotal clinical trial ONO-4538-52/TASUKI-52  [The clinical treatment and follow-up courses of the two patients are summarized in A 74-year-old male patient without past major medical and medication history was diagnosed with stage IVa left upper lung adenocarcinoma with bilateral lung metastasis in May 2019. He began combination therapy regimens of nivolumab combined with bevacizumab, carboplatin, and paclitaxel in a phase 3 clinical trial in June 2019. He had a partial response (PR) to six cycles of nivolumab combined with bevacizumab, carboplatin, and paclitaxel, and this was followed by bevacizumab combined with nivolumab every three weeks as maintenance therapy. Regular dipstick urine analysis (UA), a hemogram, and biochemistry tests were performed during every cycle of therapy to follow the trial protocol. Proteinuria (dipstick UA 3+) with normal serum creatinine (1.08 mg/dL) was detected at the 20th cycle of therapy in August 2020. The patient had no recorded clinical symptoms such as fever, skin rashes, gross hematuria, decreased urine amount, or edema associated with the proteinuria. Twenty-four-hour total urine protein was 3.96 g/day, and the kidney ultrasonography did not show significant renal parenchymal abnormality . A kidneWe were the first to report on bevacizumab-induced TMA with pathology in immune checkpoint inhibitor (ICI), combined with bevacizumab and chemotherapy in advanced non-squamous NSCLC. In a previous trial IMPOWER150, which explored the efficacy of atezolizumab combined with carboplatin, paclitaxel, and bevacizumab therapy in NSCLC, the adverse event of proteinuria occurred in 12.5% of the study subjects. The evaluation and management of serious proteinuria in the same trial were rarely reported . A previThough ICIs had been reported to induce TMA in previous studies, the incidence was low. In addition, the patients who were diagnosed with ICI-induced TMA presented with the clinical feature of acute kidney injury (AKI) with obviously increased serum creatinine and proteinuria that was not heavy ,7. The tPrevious studies have reported that acute interstitial nephritis (AIN) is the most frequent pathological finding in ICI-induced nephrotoxicity ,7,8. In Our report suggests that regular UA and renal function tests are necessary in advanced NSCLC patients receiving ICIs combined with bevacizumab, and a kidney biopsy helps to determine the cause of drug-induced nephrotoxicity and proteinuria.


Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease in which motor neurons in spinal cord and motor cortex are progressively lost. About 15% cases of ALS also develop the frontotemporal dementia (FTD), in which the frontotemporal lobar degeneration (FTLD) occurs in the frontal and temporal lobes of the brain. Among the pathologic commonalities in ALS and FTD is ubiquitin-positive cytoplasmic aggregation of TDP-43 that may reflect both its loss-of-function and gain-of-toxicity from proteostasis impairment. Deep understanding of how protein quality control mechanisms regulate TDP-43 proteinopathies still remains elusive. Recently, a growing body of evidence indicates that ubiquitinating and deubiquitinating pathways are critically engaged in the fate decision of aberrant or pathological TDP-43 proteins. E3 ubiquitin ligases coupled with deubiquitinating enzymes may influence the TDP-43-associated proteotoxicity through diverse events, such as protein stability, translocation, and stress granule or inclusion formation. In this article, we recapitulate our current understanding of how ubiquitinating and deubiquitinating mechanisms can modulate TDP-43 protein quality and its pathogenic nature, thus shedding light on developing targeted therapies for ALS and FTD by harnessing protein degradation machinery. Amyotrophic lateral sclerosis (ALS) is among the most prevalent and fatal neurodegenerative diseases in which motor neurons in spinal cord and motor cortex are progressively lost. Patients diagnosed with ALS suffer from the gradual respiratory dysfunction, having the mean survival time of 3–5 years due to no effective therapeutic treatment . ApproxiA 414-amino acid TDP-43 protein contains an N-terminal domain with a nuclear localization signal (NLS), two RNA recognition motifs (RRM1 and RRM2) with a nuclear export signal (NES), mitochondria localization signals (M1-M5), and a glycine-rich C-terminal domain where multiple ALS-associated mutations were reported  Guerrer. This febona fide protein  interaction networks. Above certain threshold that can be held by protein quality control mechanisms–such as ubiquitin-proteasome system (UPS), heat shock response, or autophagy-lysosomal degradation pathway, TDP-43 proteopathies can become runaway or even aggravated by faulty proteostasis pathways. Moreover, aberrant accumulation of pathological TDP-43 species sequesters the critical UPS components, directly impairs the proteasome activity, induces the accumulation of insoluble polyubiquitinated proteins, and also depletes free ubiquitin pool, thereby leading to perturbed protein and ubiquitin homeostasis , and organelle homeostasis (as in mitochondria) . Under peostasis . In conseostasis ; 2) the eostasis . Accumuleostasis . In addieostasis .TARDBP gene (encoding TDP-43) have been associated with ALS and FTLD-TDP , E2 (ubiquitin conjugating enzyme), and E3 (ubiquitin ligase) enzymes . By geneUBQLN, VCP, p62, and OPTN  can exclusively reverse the ubiquitination process and thus are capable of stabilizing the target proteins. Besides, DUBs also play essential roles in ubiquitin recycling and ubiquitin chain editing, which may lead to altered subcellular localization of substrates and various responses to cellular signaling . Althougin vivo fly model. Thus, in this case, UBPY may serve as a protective factor against TDP-43-induced neurotoxicity (M719V), which was genetically identified from ALS and FTLD disease locus, exhibits significantly elevated K63-linkage specific deubiquitinase activity and caused the impairment in autophagosome-lysosome fusion pathway. This CYLDM719V mutant also increased cytoplasmic mislocalization of TDP-43 and reduced the axonal length (USP14, one of the major DUBs on the proteasome, can deubiquitinate and stabilize the protein substrates that are ubiquitinated at multiple sites . Given ttoxicity . DUBs mal length . Similarl length .Aberrant stress granule dynamics represent another pathological feature of TDP-43 proteinopathy . NotablyWhile ubiquitin-enriched cytoplasmic inclusion of TDP-43 is considered to be the proteopathic signature of ALS and FTD, the pathophysiological meaning of the ubiquitination has been a long-standing question. Dramatic alteration of TDP-43 localization, stability, and post-translational modifications under the pathological conditions may disturb its genuine functional interaction networks by losing nuclear functions or gaining cytoplasmic toxicity. On top of TDP-43’s cytoplasmic mislocalization and aggregation-prone property, the initial pathogenic cue may trigger ‘butterfly effect’ and eventually the TDP-43 proteinopathy would prevail. Reduced proteasomal or autophagic activity and free ubiquitin pool depletion will badly influence the quality control of TDP-43 protein, or rather aberrant or undesirable ubiquitination of TDP-43 from proteostasis failure may only further exacerbate the disease phenotype. In fact, ubiquitination-rich pathogenic inclusions or liquid condensates, if the modification is not cleared properly, may act as avid and irreversible absorbent chambers to sequester many important proteolysis components such as ubiquitin binding proteins and proteasome. In this sense, pharmacological modulation of the pathogenic TDP-43-specific ubiquitination or the protein turnover itself may provide promising therapeutic opportunities to cope with TDP-43 proteinopathies. Remodeling or editing the ubiquitin chains or fine control of TDP-43 half-life may favor its proteolysis flux or antagonize the disease progression. It should be also noted that the total depletion of TDP-43 is not a viable option because of its essentiality. Indeed, pharmacological activation of the UPS or lysosomal pathways have been shown to bear the promise for TDP-43 clearance. For example, as noted above, a small-molecule inhibitor targeting proteasome-bound USP14 was reported to accelerate TDP-43 turnover by enhancing the proteasome activity . Similar


By simultaneously transducing and amplifying, transistors offer advantages over simpler, electrode-based transducers in electrochemical biosensors. However, transistor-based biosensors typically use static  operation modes that are poorly suited for sensor architectures relying on the modulation of charge transfer kinetics to signal analyte binding. Thus motivated, here, we translate the AC “pulsed potential” approach typically used with electrochemical aptamer-based (EAB) sensors to an organic electrochemical transistor (OECT). Specifically, by applying a linearly sweeping square-wave potential to an aptamer-functionalized gate electrode, we produce current modulation across the transistor channel two orders of magnitude larger than seen for the equivalent, electrode-based biosensor. Unlike traditional EAB sensors, our aptamer-based OECT (AB-OECT) sensors critically maintain output current even with miniaturization. The pulsed transistor operation demonstrated here could be applied generally to sensors relying on kinetics-based signaling, expanding opportunities for noninvasive and high spatial resolution biosensing. This transistor platform amplifies kinetics-based signals, allowing for substantial biosensor miniaturization. ID) (kCT), a signal transduction mechanism that is difficult to monitor effectively using DC methods  have emerged as a particularly high-performance candidate due to the large signal gain associated with volumetric ion penetration throughout a conducting polymer channel, commonly composed of the commercially available blend poly doped with poly(styrene sulfonate) (PEDOT:PSS)  sensors are rapidly growing in importance because of their versatility and their exceptional selectivity  (12), wh2 electrodes, suggesting that our approach will enable sensing at high spatial resolution in the mapping of molecular analyte concentrations.Because of the small currents produced by surface-bound aptamers, an increasingly important limitation of traditional (electrode-based) EAB sensors is the difficulty of miniaturizing them below their current, few-millimeter length scales  is applied to the aptamer-functionalized working electrode, and the resulting Faradaic (redox-reaction-derived) current flowing to a separate counter electrode is measured. As noted, the transfer kinetics associated with this Faradaic current are typically measured using SWV. In this technique, each square voltage pulse drives a current transient decay. By sampling the current at the end of the pulse (the length of which is defined by the square-wave frequency), the measured current will be monotonically related to the charge transfer kinetics of the redox reporter. To optimize this strategy, the square-wave frequency is tuned to best distinguish between the transfer kinetics of the aptamer’s bound and unbound states (VG) sweep (IG) transients  sensors such that the target-recognizing aptamers are immobilized on the gate electrode . Rather G) sweep  that yieansients  caused bents ID; , which iW, d, and L are the channel width, thickness, and length, respectively . To maintain equilibrium across the circuit, the current flowing at the gate (in response to the redox reactions occurring on it) results in an equivalent charging of the channel (QG = QCH). Such behavior is not usually captured in standard DC OECTs, which are instead designed to maximize voltage drop at the channel side (CG >> CCH) so that the transconductance  is maximized. Instead, in our case, we use the current gain (β) of the OECT to amplify the redox current at the gateThe gate-to-channel modulation that drives AB-OECT sensors can be represented by a simple circuit consisting of two capacitors in series, with CG≈CCH, is optimal (fig. S1B). That is, while lower ratios increase the voltage drop at the gate, increasing the driving force for Faradaic reactions, they also yield lower overall gate currents owing to the reduced number of redox molecules on the smaller electrode surface. In contrast, higher ratios result in too low a voltage drop at the gate to drive the Faradaic reaction as well as increase the background signal from the charging of the electric double layer at the electrode surface, lowering the overall signal-to-noise ratio. Of note, both the device geometry and the drain voltage magnitude affect the redox peak position, which is effectively being referenced against the PEDOT:PSS channel . Upon binding the drug, in contrast, the aptamer folds into a configuration that brings the redox reporter closer to the electrode, increasing kCT. Of note, for the electrode-based system, it is possible to select square-wave frequencies at which the redox peak responds to rising target concentrations by either increasing  saturates around 70 Hz (kCT and OECT governing equations obtained from previous literature (fig. S3)  (2)6-S-S-(CH2)6-GGGACTTGGTTTAGGTAATGA-GTCCC-O-CH2-CHCH2OH-(CH2)4-NH-CO-(CH2)2-methylene blue-3′] was purchased from Sangon Biotech and used as supplied. A 2-μl aliquot of 100 μM aptamer in 1X phosphate buffered saline  was thawed and mixed with 4 μl of 10 mM tris(2-carboxyethyl)phosphine (Sigma-Aldrich) for 8 hours to reduce the disulfide bond. Next, the aptamer solution was diluted to 500 nM with PBS and placed in a polydimethylsiloxane (PDMS) well over plasma-activated gate (working) electrodes for an overnight immobilization. Next, electrodes were rinsed and placed in a 10 mM 6-mercapto-1-hexanol (Sigma-Aldrich) for 5 hours to passivate. Last, electrodes were rinsed with PBS and connected to channels via a PDMS well.The aminoglycoside aptamer [5′-HO-(CHVG sweeping from −600 to −100 mV. Electrode measurements were carried out on an Autolab potentiostat (Metrohm) using the gate electrode as working electrode with Ag/AgCl reference and platinum counter electrodes. For square-wave measurements on both the OECT and electrode, a 5-mV step and 20-mV amplitude were used, as adapted from previous work on the aminoglycoside aptamer (OECT measurements were collected using a Keysight B1500A semiconductor device analyzer. All OECT measurements used a