Beta Changes Induced by Acute Hand Loss Model during NMES.

Neuromuscular electrical stimulation (NMES) has been demonstrated to effectively modulate cortical activities by evoking muscle contraction in upper limb and generating joint movements, which showed an excellent performance in motor rehabilitation. However, due to hand loss and cortical function reorganization induced by hand amputation, how neural activities in sensorimotor cortex response to NMES-evoked muscle contraction in the end of an amputation stump is not clear. In this paper, Ischemic nerve block (INB) technique was used to build an acute hand loss model, and 64-channel EEG signals were recorded from 11 healthy subjects to perform a 2×2 factorial design protocol, with the INB state and the current intensity as factors. The changes of NMES-evoked sensorimotor cortical activities were quantified by computing Beta-band event-related desynchronization (Beta ERD) patterns and the time-varying functional connectivity using adaptive directed transfer function (ADTF) before and during INB. The acute hand "loss" resulted in ipsilateral dominance of Beta ERD induced by NMES with two current intensities in the topographic maps, that is, ipsilateral Beta ERD was significantly higher than that the contralateral one (p<0.05). However, before INB, Beta ERD in the contralateral sensorimotor cortex induced by NMES above motor threshold was significantly higher than that in the ipsilateral area (p< 0.01). Meanwhile, whatever before or during INB, clustering coefficients of the ADTF network in sensorimotor cortex showed temporal dynamics during two NMES tasks. During INB, NMES above motor threshold-evoked lower clustering coefficients of the time-varying network in sensorimotor cortex than that before INB (p<0.05). The present results suggest that the loss of the hand proprioception will degrade cortical activities in the contralateral area, and increase cortical activities in the ipsilateral area compensatively responding to NMES. This finding may be particularly important to improve the reconstruction of the proprioception function of hand prosthesis.

The Kinematic-Muscle Synergies during Infant Crawling: A Pilot Study.

Many feature extraction algorithms have been separately used for kinematic or muscle synergy analysis during human movement. However, very few studies focus on the co-extraction of kinematic and muscle synergies. Therefore, the aim of this study was to propose a novel and efficient approach for extracting the kinematic-muscle synergies during infant crawling. Surface electromyography signals and three-dimensional joint trajectories were collected from 20 typically developing infants during self-paced hands-and-knees crawling. Angular accelerations of shoulder, elbow, hip and knee flexion/extension computing from those joint trajectories were divided into two independent directional positive degrees-of-freedom. The kinematic-muscle synergies and corresponding activation coefficients were extracted using the non-negative matrix factorization algorithm based on two selection criteria of synergy number (i.e., criterion 1: the total constraint, criterion 2: a combination of the total constraint and a local constraint for each joint/muscle). Then, the data of each joint/muscle were reconstructed by those synergies and corresponding activation coefficients. Our results indicated that the minimum number of kinematic-muscle synergies based on criterion 1 is less than that based on criterion 2. The data reconstruction of joint flexion/extension based on criterion 2 is better than that based on criterion 1, whereas the data reconstruction of muscles is similar between criterion 1 and 2. These promising results show the feasibility of applying the proposed approach to clinical assessments of motor function for infants.Clinical Relevance- Extracting kinematic-muscle synergies during infant crawling has the potential for professional therapists or rehabilitation physicians to conduct the early assessment and rehabilitation treatment of infants with the central nervous system disorders.

Synthetic gRNA/Cas9 ribonucleoprotein targeting HBV DNA inhibits viral replication.

The presence of hepatitis B virus (HBV) covalently closed circular (ccc) DNA (cccDNA), which serves as a template for viral replication and integration of HBV DNA into the host cell genome, sustains liver pathogenesis and constitutes an intractable barrier to the eradication of chronic HBV infection. The current antiviral therapy for HBV infection, using nucleos(t)ide analogues (NAs), can suppress HBV replication but cannot eliminate integrated HBV DNA and episomal cccDNA. Clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 is a powerful genetic tool that can edit integrated HBV DNA and minichromosomal cccDNA for gene therapy, but its expression and delivery require a viral vector, which poses safety concerns for therapeutic applications in humans. In the present study, we used synthetic guide RNA (gRNA)/Cas9-ribonucleoprotein (RNP) as a nonviral formulation to develop a novel CRISPR/Cas9-mediated gene therapy for eradicating HBV infection. We designed a series of gRNAs targeting multiple specific HBV genes and tested their antiviral efficacy and cytotoxicity in different HBV cellular models. Transfection of stably HBV-infected human hepatoma cell line HepG2.2.15 with HBV-specific gRNA/Cas9 RNPs resulted in a substantial reduction in HBV transcripts. Specifically, gRNA5 and/or gRNA9 RNPs significantly reduced HBV cccDNA, total HBV DNA, pregenomic RNA, and HBV antigen (HBsAg, HBeAg) levels. T7 endonuclease 1 (T7E1) cleavage assay and DNA sequencing confirmed specific HBV gene cleavage and mutations at or around the gRNA target sites. Notably, this gene-editing system did not alter cellular viability or proliferation in the treated cells. Because of their rapid DNA cleavage capability, low off-target effects, low risk of insertional mutagenesis, and readiness for use in clinical application, these results suggest that synthetic gRNA/Cas9 RNP-based gene-editing can be utilized as a promising therapeutic drug for eradicating chronic HBV infection.

ROS-Induced Mitochondrial Dysfunction in CD4 T Cells from ART-Controlled People Living with HIV.

We have previously demonstrated mitochondrial dysfunction in aging CD4 T cells from antiretroviral therapy (ART)-controlled people living with HIV (PLWH). However, the underlying mechanisms by which CD4 T cells develop mitochondrial dysfunction in PLWH remain unclear. In this study, we sought to elucidate the mechanism(s) of CD4 T cell mitochondrial compromise in ART-controlled PLWH. We first assessed the levels of reactive oxygen species (ROS), and we observed significantly increased cellular and mitochondrial ROS levels in CD4 T cells from PLWH compared to healthy subjects (HS). Furthermore, we observed a significant reduction in the levels of proteins responsible for antioxidant defense (superoxide dismutase 1, SOD1) and ROS-mediated DNA damage repair (apurinic/apyrimidinic endonuclease 1, APE1) in CD4 T cells from PLWH. Importantly, CRISPR/Cas9-mediated knockdown of SOD1 or APE1 in CD4 T cells from HS confirmed their roles in maintaining normal mitochondrial respiration via a p53-mediated pathway. Reconstitution of SOD1 or APE1 in CD4 T cells from PLWH successfully rescued mitochondrial function as evidenced by Seahorse analysis. These results indicate that ROS induces mitochondrial dysfunction, leading to premature T cell aging via dysregulation of SOD1 and APE1 during latent HIV infection.

Mitochondrial topoisomerase 1 inhibition induces topological DNA damage and T cell dysfunction in patients with chronic viral infection.

T cells are crucial for controlling viral infections; however, the mechanisms that dampen their responses during viral infections remain incompletely understood. Here, we studied the role and mechanisms of mitochondrial topoisomerase 1 (Top1mt) inhibition in mitochondrial dysfunction and T cell dysregulation using CD4 T cells from patients infected with HCV or HIV and compared it with CD4 T cells from healthy individuals following treatment with Top1 inhibitor - camptothecin (CPT). We found that Top1mt protein levels and enzymatic activity are significantly decreased, along with Top1 cleavage complex (Top1cc) formation, in mitochondria of CD4 T cells from HCV- and HIV-infected patients. Notably, treatment of healthy CD4 T cells with CPT caused similar changes, including inhibition of Top1mt, accumulation of Top1cc in mitochondria, increase in PARP1 cleavage, and decrease in mtDNA copy numbers. These molecular changes resulted in mitochondrial dysfunction, T cell dysregulation, and programmed cell death through multiple signaling pathways, recapitulating the phenotype we detected in CD4 T cells from HCV- and HIV-infected patients. Moreover, treatment of CD4 T cells from HCV or HIV patients with CPT further increased cellular and mitochondrial reactive oxygen species (ROS) production and cell apoptosis, demonstrating a critical role for Top1 in preventing mtDNA damage and cell death. These results provide new insights into the molecular mechanisms underlying immune dysregulation during viral infection and indicate that Top1 inhibition during chronic HCV or HIV infection can induce mtDNA damage and T cell dysfunction. Thus, reconstituting Top1mt protein may restore the mtDNA topology and T cell functions in humans with chronic viral infection.

Identification of virus-specific B-cell epitopes by convalescent plasma from COVID-19 patients.

Identification of immunologic epitopes against SARS-CoV-2 is crucial for the discovery of diagnostic, therapeutic, and preventive targets. In this study, we used a pan-coronavirus peptide microarray to screen for potential B-cell epitopes and validated the results with peptide-based ELISA. Specifically, we identified three linear B-cell epitopes on the SARS-CoV-2 proteome, which were recognized by convalescent plasma from COVID-19 patients. Interestingly, two epitopes (S 809-823 and R1ab 909-923) strongly reacted to convalescent plasma collected at the early phase (< 90 days) of COVID-19 symptom onset, whereas one epitope (M 5-19) reacted to convalescent plasma collected > 90 days after COVID-19 symptom onset. Neutralization assays using antibody depletion with the identified spike (S) peptides revealed that three S epitopes (S 557-571, S 789-803, and S 809-823) elicited neutralizing antibodies in COVID-19 patients. However, the levels of virus-specific antibody targeting S 789-803 only positively correlated with the neutralizing rates at the early phase (<60 days) after disease onset, and the antibody titers diminished quickly with no correlation to the neutralizing activity beyond two months after recovery from COVID-19. Importantly, stimulation of peripheral blood mononuclear cells from COVID-19-recovered patients with these SARS-CoV-2 S peptides resulted in poor virus-specific B cell activation, proliferation, differentiation into memory B cells, and production of immunoglobulin G (IgG) antibodies, despite the B-cells being functionally competent as demonstrated by their response to non-specific stimulation. Taken together, these findings indicate that these newly identified SARS-CoV-2-specific B-cell epitopes can elicit neutralizing antibodies, with titers and/or neutralizing activities declining significantly within 2-3 months in the convalescent plasma of COVID-19 patients.

Synthetic gRNA/Cas9 Ribonucleoprotein Inhibits HIV Reactivation and Replication.

The current antiretroviral therapy (ART) for human immunodeficiency virus (HIV) can halt viral replication but cannot eradicate HIV infection because proviral DNA integrated into the host genome remains genetically silent in reservoir cells and is replication-competent upon interruption or cessation of ART. CRISPR/Cas9-based technology is widely used to edit target genes via mutagenesis (i.e., nucleotide insertion/deletion and/or substitution) and thus can inactivate integrated proviral DNA. However, CRISPR/Cas9 delivery systems often require viral vectors, which pose safety concerns for therapeutic applications in humans. In this study, we used synthetic guide RNA (gRNA)/Cas9-ribonucleoprotein (RNP) as a non-viral formulation to develop a novel HIV gene therapy. We designed a series of gRNAs targeting different HIV genes crucial for HIV replication and tested their antiviral efficacy and cellular cytotoxicity in lymphoid and monocytic latent HIV cell lines. Compared with the scramble gRNA control, HIV-gRNA/Cas9 RNP-treated cells exhibited efficient viral suppression with no apparent cytotoxicity, as evidenced by the significant inhibition of latent HIV DNA reactivation and RNA replication. Moreover, HIV-gRNA/Cas9 RNP inhibited p24 antigen expression, suppressed infectious viral particle production, and generated specific DNA cleavages in the targeted HIV genes that are confirmed by DNA sequencing. Because of its rapid DNA cleavage, low off-target effects, low risk of insertional mutagenesis, easy production, and readiness for use in clinical application, this study provides a proof-of-concept that synthetic gRNA/Cas9 RNP drugs can be utilized as a novel therapeutic approach for HIV eradication.

Corrigendum: A matter of life or death: Productively infected and bystander CD4 T Cells in early HIV infection.

[This corrects the article DOI: 10.3389/fimmu.2020.626431.].

Plasma biomarkers for systemic inflammation in COVID-19 survivors.

BACKGROUND: While the majority of COVID-19 patients fully recover from the infection and become asymptomatic, a significant proportion of COVID-19 survivors experience a broad spectrum of symptoms lasting weeks to months post-infection, a phenomenon termed "post-acute sequelae of COVID-19 (PASC)." The aim of this study is to determine whether inflammatory proteins are dysregulated and can serve as potential biomarkers for systemic inflammation in COVID-19 survivors. METHODS: We determined the levels of inflammatory proteins in plasma from 22 coronavirus disease 2019 (COVID-19) long haulers (COV-LH), 22 COVID-19 asymptomatic survivors (COV-AS), and 22 healthy subjects (HS) using an Olink proteomics assay and assessed the results by a beads-based multiplex immunoassay. RESULTS: Compared to HS, we found that COVID-19 survivors still exhibited systemic inflammation, as evidenced by significant changes in the levels of multiple inflammatory proteins in plasma from both COV-LH and COV-AS. CXCL10 was the only protein that significantly upregulated in COV-LH compared with COV-AS and HS. CONCLUSIONS: Our results indicate that several inflammatory proteins remain aberrantly dysregulated in COVID-19 survivors and CXCL10 might serve as a potential biomarker to typify COV-LH. Further characterization of these signature inflammatory molecules might improve the understanding of the long-term impacts of COVID-19 and provide new targets for the diagnosis and treatment of COVID-19 survivors with PASC.

TRF2 inhibition rather than telomerase disruption drives CD4T cell dysfunction during chronic viral infection.

We investigated the role of telomerase and telomere repeat-binding factor 2 (TRF2 or TERF2) in T-cell dysfunction in chronic viral infection. We found that the expression and activity of telomerase in CD4+ T (CD4T) cells from patients with hepatitis C virus (HCV) infections or people living with HIV (PLWH) were intact, but TRF2 expression was significantly inhibited at the post-transcriptional level, suggesting that TRF2 inhibition is responsible for the CD4T cell dysfunction observed during chronic viral infection. Silencing TRF2 expression in CD4T cells derived from healthy subjects induced telomeric DNA damage and CD4T cell dysfunction without affecting telomerase activity or translocation - similar to what we observed in CD4T cells from HCV patients and PLWH. These findings indicate that premature T-cell aging and dysfunction during chronic HCV or HIV infection are primarily caused by chronic immune stimulation and T-cell overactivation and/or proliferation that induce telomeric DNA damage due to TRF2 inhibition, rather than telomerase disruption. This study suggests that restoring TRF2 presents a novel approach to prevent telomeric DNA damage and premature T-cell aging, thus rejuvenating T-cell functions during chronic viral infection.

Impacts of Motor Developmental Delay on the Inter-Joint Coordination Using Kinematic Synergies of Joint Angles During Infant Crawling.

Motor developmental delay (MDD) usually affects the inter-joint coordination for limb movement. However, the mechanism between the abnormal inter-joint coordination and MDD is still unclear, which poses a challenge for clinical diagnosis and motor rehabilitation of MDD in infant's early life. This study aimed to explore whether the joint activities of limbs during infant crawling are represented with kinematic synergies of joint angles, and evaluate the impacts of MDD on the inter-joint coordination using those synergies. 20 typically developing infants, 16 infants at risk of developmental delay, 11 infants at high risk of developmental delay and 13 infants with confirmed developmental delay were recruited for self-paced crawling on hands and knees. A motion capture system was employed to trace infants' limbs in space, and angles of shoulder, elbow, hip and knee over time were computed. Kinematic synergies were derived from joint angles using principal component analysis. Sample entropy and Spearman's rank correlation coefficients were calculated among those synergies to evaluate the crawling complexity and the symmetry of bilateral limbs, respectively. We found that the first two synergies with different contributions to the crawling movements sufficiently represented the joint angular profiles of limbs. MDD further delayed the development of motor function for lower limbs and mainly increased the crawling complexity of joint flexion/extension to some extent, but did not obviously change the symmetry of bilateral limbs. These results suggest that the time-varying kinematic synergy of joint angles is a potential index for objectively evaluating the abnormal inter-joint coordination affected by MDD.

Oxidative Stress Induces Mitochondrial Compromise in CD4 T Cells From Chronically HCV-Infected Individuals.

We have previously shown that chronic Hepatitis C virus (HCV) infection can induce DNA damage and immune dysfunctions with excessive oxidative stress in T cells. Furthermore, evidence suggests that HCV contributes to increased susceptibility to metabolic disorders. However, the underlying mechanisms by which HCV infection impairs cellular metabolism in CD4 T cells remain unclear. In this study, we evaluated mitochondrial mass and intracellular and mitochondrial reactive oxygen species (ROS) production by flow cytometry, mitochondrial DNA (mtDNA) content by real-time qPCR, cellular respiration by seahorse analyzer, and dysregulated mitochondrial-localized proteins by Liquid Chromatography-Mass Spectrometry (LC-MS) in CD4 T cells from chronic HCV-infected individuals and health subjects. Mitochondrial mass was decreased while intracellular and mitochondrial ROS were increased, expressions of master mitochondrial regulators peroxisome proliferator-activated receptor 1 alpha (PGC-1α) and mitochondrial transcription factor A (mtTFA) were down-regulated, and oxidative stress was increased while mitochondrial DNA copy numbers were reduced. Importantly, CRISPR/Cas9-mediated knockdown of mtTFA impaired cellular respiration and reduced mtDNA copy number. Furthermore, proteins responsible for mediating oxidative stress, apoptosis, and mtDNA maintenance were significantly altered in HCV-CD4 T cells. These results indicate that mitochondrial functions are compromised in HCV-CD4 T cells, likely via the deregulation of several mitochondrial regulatory proteins.

Multiple Sessions of Entorhinal Cortex Deep Brain Stimulation in C57BL/6J Mice Increases Exploratory Behavior and Hippocampal Neurogenesis.

Deep brain stimulation (DBS) has been a medical intervention for a variety of nervous system diseases and mental diseases. The input of DBS in the entorhinal cortex (EC) regulates the neurophysiological activities in its downstream regions, such as the dentate gyrus (DG) area. EC DBS may play a role in the treatment of diseases through hippocampal neurogenesis. This study we examined the effect of multiple sessions of EC DBS on the regulation of hippocampal neurogenesis. 4-month-old male C57BL/6J mice received bilateral multiple sessions of EC DBS (130 Hz, 90 µs, 100 µA, 1 h/d, 21 days), and the DBS parameters used are close to the high-frequency DBS parameters in clinical studies. The open field test (OFT) was used to test the exploratory behavior of mice, and hippocampal neurogenesis was detected by immunofluorescence staining with anti-doublecortin (DCX). We found that multiple sessions of EC DBS were tolerated in C57BL/6J mice, significantly increased exploratory behavior and the number of DCX-positive neurons in the DG area.Clinical Relevance- Hippocampal neurogenesis may be part of the reason for DBS to improve memory, and the results of this study show that multiple sessions of EC DBS increases exploratory behavior and hippocampal neurogenesis, which is conducive to the application of DBS in nervous system diseases and mental diseases related to memory impairment.

Angular Velocity Profiles of Upper Limb Joint Synergies in Reaching Movements: a pilot study.

The spatiotemporal kinematic synergy, a coupling of multiple degrees of freedom (DoF), runs through human activities of daily living (ADL). And it is an entry point for exploring the central nervous system's (CNS) control process of musculoskeletal system by analyzing the time-varying kinematic synergy. The aim of this study was to find more physiological properties from the angular velocity profiles of synergy. Ten healthy right-handed subjects were asked to reach target button at different locations. During reaching movement, the motion data of five right upper limb joints were recorded, and the synergistic patterns were extracted by PCA algorithm. Our results showed that the combinations of the first four synergies were sufficient to explain raw data. As far as possible to exclude the effects of individual and information differences, we found shoulder flexion/extension and elbow flexion/extension made distinct contribution in a period of time to the control procedure performed by CNS after targets were confirmed. Our preliminary results implied that reaching movements required comparatively constant scheduling of shoulder horizontal abduction/adduction, shoulder internal/external rotation and wrist ulnar/radial deviation by CNS, while scheduling of SFE and EFE depends on the objectives.Clinical relevance- The findings of this paper may provide a novel dynamic control evidence based on CNS for realizing near-natural control of assistive devices in motor rehabilitation area.

Study on the Establishment Process of Muscle Synergy Based on Cosine Similarity.

Muscle synergy is an important method for motor intention recognition in rehabilitation exoskeleton control. The use of the non-negative matrix factorization (NMF) to extract muscle synergy patterns often results in long calculation time due to the amount of data, which makes the effectiveness of synergy extraction low. In this paper, synergy matrices of the complete single-cycle signal while stretching and its segmented ones were extracted respectively. By studying the cosine similarity variation of synergy matrices between each continuous segment and the complete single-cycle EMG signals, it is found that there is a "building-stability-weakening" process on muscle synergy establishment. It is proposed to extract synergy mode with partial data from the "stable" segment, rather than using the complete single-cycle one, as similar result to single-cycle data synergy extraction could be obtained. The calculation time of NMF could be optimized by reducing the amount of data and the real-time characteristics of the synergy mode extraction could be improved at the same time. It is of great significance to use synergy matrix of NMF for motion intention recognition and exoskeleton control.Clinical Relevance- This paper studies the establishment process of the synergy mode, and proposes a method for quickly extracting the synergy mode, which can improve the effectiveness of the recognition of motion intention and is of great significance for the real-time control of the rehabilitation exoskeleton.

Selective oxidative stress induces dual damage to telomeres and mitochondria in human T cells.

Oxidative stress caused by excess reactive oxygen species (ROS) accelerates telomere erosion and mitochondrial injury, leading to impaired cellular functions and cell death. Whether oxidative stress-mediated telomere erosion induces mitochondrial injury, or vice versa, in human T cells-the major effectors of host adaptive immunity against infection and malignancy-is poorly understood due to the pleiotropic effects of ROS. Here we employed a novel chemoptogenetic tool that selectively produces a single oxygen (1 O2 ) only at telomeres or mitochondria in Jurkat T cells. We found that targeted 1 O2 production at telomeres triggered not only telomeric DNA damage but also mitochondrial dysfunction, resulting in T cell apoptotic death. Conversely, targeted 1 O2 formation at mitochondria induced not only mitochondrial injury but also telomeric DNA damage, leading to cellular crisis and apoptosis. Targeted oxidative stress at either telomeres or mitochondria increased ROS production, whereas blocking ROS formation during oxidative stress reversed the telomeric injury, mitochondrial dysfunction, and cellular apoptosis. Notably, the X-ray repair cross-complementing protein 1 (XRCC1) in the base excision repair (BER) pathway and multiple mitochondrial proteins in other cellular pathways were dysregulated by the targeted oxidative stress. By confining singlet 1 O2 formation to a single organelle, this study suggests that oxidative stress induces dual injury in T cells via crosstalk between telomeres and mitochondria. Further identification of these oxidation pathways may offer a novel approach to preserve mitochondrial functions, protect telomere integrity, and maintain T cell survival, which can be exploited to combat various immune aging-associated diseases.

Measurement and Analysis of Human Infant Crawling for Rehabilitation: A Narrative Review.

When a child shows signs of potential motor developmental disorders, early diagnosis of central nervous system (CNS) impairment is beneficial. Known as the first CNS-controlled mobility for most of infants, mobility during crawling usually has been used in clinical assessments to identify motor development disorders. The current clinical scales of motor development during crawling stage are relatively subjective. Objective and quantitative measures of infant crawling afford the possibilities to identify those infants who might benefit from early intervention, as well as the evaluation of intervention progress. Thus, increasing researchers have explored objective measurements of infant crawling in typical and atypical developing infants. However, there is a lack of comprehensive review on infant-crawling measurement and analysis toward bridging the gap between research crawling analysis and potential clinical applications. In this narrative review, we provide a practical overview of the most relevant measurements in human infant crawling, including acquisition techniques, data processing methods, features extraction, and the potential value in objective assessment of motor function in infancy; meanwhile, the possibilities to develop crawling training as early intervention to promote the locomotor function for infants with locomotor delays are also discussed.

SARS-CoV-2 specific memory T cell epitopes identified in COVID-19-recovered subjects.

The COVID-19 pandemic caused by SARS-CoV-2 infection poses a serious threat to public health. An explicit investigation of COVID-19 immune responses, particularly the host immunity in recovered subjects, will lay a foundation for the rational design of therapeutics and/or vaccines against future coronaviral outbreaks. Here, we examined virus-specific T cell responses and identified T cell epitopes using peptides spanning SARS-CoV-2 structural proteins. These peptides were used to stimulate peripheral blood mononuclear cells (PBMCs) derived from COVID-19-recovered subjects, followed by an analysis of IFN-γ-secreting T cells by enzyme-linked immunosorbent spot (ELISpot). We also evaluated virus-specific CD4 or CD8 T cell activation by flow cytometry assay. By screening 52 matrix pools (comprised of 315 peptides) of the spike (S) glycoprotein and 21 matrix pools (comprised of 102 peptides) spanning the nucleocapsid (N) protein, we identified 28 peptides from S protein and 5 peptides from N protein as immunodominant epitopes. The immunogenicity of these epitopes was confirmed by a second ELISpot using single peptide stimulation in memory T cells, and they were mapped by HLA restrictions. Notably, SARS-CoV-2 specific T cell responses positively correlated with B cell IgG and neutralizing antibody responses to the receptor-binding domain (RBD) of the S protein. Our results demonstrate that defined levels of SARS-CoV-2 specific T cell responses are generated in some, but not all, COVID-19-recovered subjects, fostering hope for the protection of a proportion of COVID-19-exposed individuals against reinfection. These results also suggest that these virus-specific T cell responses may induce protective immunity in unexposed individuals upon vaccination, using vaccines generated based on the immune epitopes identified in this study. However, SARS-CoV-2 S and N peptides are not potently immunogenic, and none of the single peptides could universally induce robust T cell responses, suggesting the necessity of using a multi-epitope strategy for COVID-19 vaccine design.

Immune Activation Induces Telomeric DNA Damage and Promotes Short-Lived Effector T Cell Differentiation in Chronic HCV Infection.

BACKGROUND AND AIMS: Hepatitis C virus (HCV) leads to a high rate of chronic infection and T cell dysfunction. Although it is well known that chronic antigenic stimulation is a driving force for impaired T cell functions, the precise mechanisms underlying immune activation-induced T cell dysfunctions during HCV infection remain elusive. APPROACH AND RESULTS: Here, we demonstrated that circulating CD4+ T cells from patients who are chronically HCV-infected exhibit an immune activation status, as evidenced by the overexpression of cell activation markers human leukocyte antigen-antigen D-related, glucose transporter 1, granzyme B, and the short-lived effector marker CD127- killer cell lectin-like receptor G1+ . In contrast, the expression of stem cell-like transcription factor T cell factor 1 and telomeric repeat-binding factor 2 (TRF2) are significantly reduced in CD4+ T cells from patients who are chronically HCV-infected compared with healthy participants (HP). Mechanistic studies revealed that CD4+ T cells from participants with HCV exhibit phosphoinositide 3-kinase/Akt/mammalian target of rapamycin signaling hyperactivation on T cell receptor stimulation, promoting proinflammatory effector cell differentiation, telomeric DNA damage, and cellular apoptosis. Inhibition of Akt signaling during T cell activation preserved the precursor memory cell population and prevented inflammatory effector cell expansion, DNA damage, and apoptotic death. Moreover, knockdown of TRF2 reduced HP T cell stemness and triggered telomeric DNA damage and cellular apoptosis, whereas overexpression of TRF2 in CD4 T cells prevented telomeric DNA damage. CONCLUSIONS: These results suggest that modulation of immune activation through inhibiting Akt signaling and protecting telomeres through enhancing TRF2 expression may open therapeutic strategies to fine tune the adaptive immune responses in the setting of persistent immune activation and inflammation during chronic HCV infection.

Mitochondrial Functions Are Compromised in CD4 T Cells From ART-Controlled PLHIV.

The hallmark of HIV/AIDS is a gradual depletion of CD4 T cells. Despite effective control by antiretroviral therapy (ART), a significant subgroup of people living with HIV (PLHIV) fails to achieve complete immune reconstitution, deemed as immune non-responders (INRs). The mechanisms underlying incomplete CD4 T cell recovery in PLHIV remain unclear. In this study, CD4 T cells from PLHIV were phenotyped and functionally characterized, focusing on their mitochondrial functions. The results show that while total CD4 T cells are diminished, cycling cells are expanded in PLHIV, especially in INRs. HIV-INR CD4 T cells are more activated, displaying exhausted and senescent phenotypes with compromised mitochondrial functions. Transcriptional profiling and flow cytometry analysis showed remarkable repression of mitochondrial transcription factor A (mtTFA) in CD4 T cells from PLHIV, leading to abnormal mitochondrial and T cell homeostasis. These results demonstrate a sequential cellular paradigm of T cell over-activation, proliferation, exhaustion, senescence, apoptosis, and depletion, which correlates with compromised mitochondrial functions. Therefore, reconstituting the mtTFA pathway may provide an adjunctive immunological approach to revitalizing CD4 T cells in ART-treated PLHIV, especially in INRs.