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Introduction The aim of the present study was to find the relation of exercise-induced biomarkers (antioxidant, muscle damage, and inflammatory markers) with endurance capacity and anaerobic power. The study also aimed to develop predicting regression models for maximal oxygen uptake (V̇O2max) and relative anaerobic power (Wpeak) to specify the essential performance limiting elements. Material and Methods Eighty-six endurance male players (i.e., football (n = 39) and field hockey (n = 47)) were selected as test subjects for the present study. Muscle damage indices (creatine kinase (CK), lactate dehydrogenase (LDH), cortisol), inflammatory markers (interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α)), antioxidant variables (malondialde-hyde (MDA), superoxide dismutase (SOD), glutathione (GSH), glutathione peroxidase (GPx)) and performance variables (indicated as V̇O2max and Wpeak) were assessed using standard protocols. Results The most significant (sig p ---lt--- 0.001) prediction of V̇O2max = (0.763) MDA+ (5.644) SOD+ (0.039) GSH- (0.154) GPx+ (0.002) LDH- (0.011) CK+ (0.038) cortisol+ (1.232) IL+ (1.135) TNF+ 20.018. The strongest correlations were found between V̇O2max vs MDA (R2 = 0.852), V̇O2max vs IL-6 (R2 = 0.589), V̇O2max vs TNF-α (R2 = 0.385). Conclusions Artificial neural network perceptron model depicted stronger prediction of V̇O2max (R2 = 0.872) in comparison to Wpeak (R2 = 0.271), with MDA and CK as the major predictors for V̇O2max and Wpeak, respectively. Among all biomarkers, MDA, IL-6, and TNF-α were identified as the most valuable indicators to predict endurance capacity significantly. While MDA, SOD, GPx, IL-6, and TNF-α were strongly correlated with V̇O2max and LDH, cortisol was strongly correlated with Wpeak. Contrarily, exercise-induced biomarkers failed to predict anaerobic power.

Creatine kinase (CK), lactate dehydrogenase (LDH) and cortisol are widely accepted as biological markers. The purpose of the study was to frame the reference interval for muscle damage indices (CK, LDH) and cortisol in the young athletic population of various sports disciplines. 260 young male players [i.e., football (n=62), hockey (n=60), gymnastics (n=36), swimming (n=28), table tennis (n=25), sprint-jump-throw (n=36) and middle-long distance running (n=13)] were recruited for the study (mean age = 15.6±1.59 yrs). Assay of LDH, CK and cortisol was done using the standard enzymatic protocol. The reference interval was calculated by following the Clinical and Laboratory Standard Institute (CLSI) C28-A3 guideline and “MedCalc” software (version 19) with a 90% confidence interval. Serum LDH range was from 148.00-324.00 IU/L with a mean of 233.2±34.74 and a median around 236.25. Serum CK ranged from 17.00-43.50 IU/L with a mean of 28.93±5.23 IU/L and a median around 28.00. Cortisol ranged from 4.99-15.78 µg/dl with a mean of 9.31±2.09 µg/dl and a median around 8.90. The present study confers 165.63 - 303.43 IU/L, 19.00 – 40.09 IU/L and 6.07-14.15 µg/dl as the reference interval values for LDH, CK and cortisol, respectively. The present finding will guide the researchers to avoid misinterpretation of muscle damage indices values during any phase of competitive training of sports person.

Stem-like progenitor exhausted CD8 T cells are critical for maintaining long-term resistance during chronic infections and cancer, and represent an important checkpoint blockade immunotherapy target for functional reinvigoration and disease control. Hence, there is vigorous interest in understanding the ontogenesis of TCF-1Histem-like exhausted CD8 T cells, and in defining the signals that promote their development. Here, we show that virus-specific TCF1HiGzmBLoCD8 T cells develop during early stages of chronic viral infection, and are progressively deprogrammed from the stem-like lineage towards terminal differentiation by strong IL-2 signals. In vivo fate-tracking studies show that strong T cell-intrinsic IL-2 signals through the high affinity heterotrimeric IL-2Rα/β/γ receptor drive skewed development of terminally differentiated TCF-1LoGzmBHicells, which largely die in chronic antigenic environment. In contrast, tempered IL-2 signals through the intermediate affinity IL-2Rβ/γ heterodimer, or delayed priming in diminished IL-2 milieu support preferential development of TCF-1Histem-like exhausted CD8 T cells, capable of long-term persistence and potent responsiveness to PD-1 therapy in later stages of chronic viral infection. In human tumors as well, single cell RNA-seq analyses of tumor infiltrating lymphocytes from melanoma, human papillomavirus+ head and neck cancer and lung cancer patients revealed an inverse relationship between IL-2 signaling signature and T cell stemness. Moreover, melanoma patients with enriched IL-2 signaling signature showed poor responses to checkpoint blockade immunotherapy. Collectively, these findings bear relevance to clinical immunotherapies of chronic viral infections and cancers, and support exogenous IL-2 signal manipulation prior to therapy as a strategy for augmenting long-term clinical outcomes.ONE SENTENCE SUMMARYProgramming of exhausted T cell fates by differential IL-2

Subacute necrotizing encephalopathy, or Leigh syndrome (LS), is the most common paediatric presentation of genetic mitochondrial disease. LS is a multi-system disorder with severe neurologic, metabolic, and musculoskeletal symptoms. The presence of progressive, symmetric, necrotizing lesions in the brainstem are a defining feature of the disease, and the major cause of morbidity and mortality, but the mechanisms underlying their pathogenesis have been elusive. Recently, we demonstrated that high-dose pexidartinib, a CSF1R inhibitor, prevents LS CNS lesions and systemic disease in theNdufs4(-/-) mouse model of LS. While the dose-response in this study implicated peripheral immune cells, the immune populations involved have not yet been elucidated. Here, we used a targeted genetic tool, deletion of the colony stimulating factor 1 receptor (CSF1R) macrophage super-enhancer FIRE (Csf1rΔFIRE), to specifically deplete microglia and define the role of microglia in the pathogenesis of LS. Homozygosity for theCsf1rΔFIRE allele ablates microglia in both control andNdufs4(-/-) animals, but onset of CNS lesions and sequalae in theNdufs4(-/-), including mortality, are only marginally impacted by microglia depletion. The overall development of necrotizing CNS lesions is not altered, though microglia remain absent. Finally, histologic analysis of brainstem lesions provides direct evidence of a causal role for peripheral macrophages in the characteristic CNS lesions. These data demonstrate that peripheral macrophages play a key role in the pathogenesis disease in theNdufs4(-/-) model.

Inhibitory signaling in dysfunctional CD8 T cells through the programmed cell death 1 (PD-1) axis is well established in chronic viral infections and cancers. PD-1 is also transiently induced to high concentrations during priming of acute infections and immunizations, yet its impact on the development of long-lived antigen-independent T cell memory remains unclear. In addition to its expected role in restraining clonal effector expansion, here, we show that PD-1 expression on antigen-specific CD8 T cells is required for the development of a durable CD8 T cell memory pool after antigen clearance. Loss of T cell–specific PD-1 signaling led to increased contraction and a defect in antigen-independent renewal of memory CD8 T cells in response to homeostatic cytokine signals, thus resulting in attrition of the memory pool over time. Whereas exhausted CD8 T cells regain function after PD-1 checkpoint blockade during chronic viral infection, the preexisting pool of resting functional bystander memory CD8 T cells established in response to a previously administered immunogen decreased. Metabolically, PD-1 signals were necessary for regulating the critical balance of mTOR-dependent anabolic glycolysis and fatty acid oxidation programs to meet the bioenergetic needs of quiescent CD8 T cell memory. These results define PD-1 as a key metabolic regulator of protective T cell immunity. Furthermore, these results have potential clinical implications for preexisting CD8 T cell memory during PD-1 checkpoint blockade therapy.

Type-I interferon (IFN-I) signals exert a critical role in disease progression during viral infections. However, the immunomodulatory mechanisms by which IFN-I dictates disease outcomes remain to be fully defined. Here we report that IFN-I signals mediate thymic atrophy in viral infections, with more severe and prolonged loss of thymic output and unique kinetics and subtypes of IFN-α/β expression in chronic infection compared to acute infection. Loss of thymic output was linked to inhibition of early stages of thymopoiesis (DN1-DN2 transition, and DN3 proliferation) and pronounced apoptosis during the late DP stage. Notably, infection-associated thymic defects were largely abrogated upon ablation of IFNαβR and partially mitigated in the absence of CD8 T cells, thus implicating direct as well as indirect effects of IFN-I on thymocytes. These findings provide mechanistic underpinnings for immunotherapeutic strategies targeting IFN-1 signals to manipulate disease outcomes during chronic infections and cancers.

A major obstacle in current T cell-based immunotherapy against solid tumors is the widely observed T cell exhaustion and loss of functionality within the tumor microenvironment (TME). We, and others, have demonstrated that chronic antigenic signaling is the predominant factor that drives TCR tumor-reactive, and CAR T cell exhaustion in the TME. To combat this exhaustion, we have engineered CD19-targeted third generation CAR constructs to express pro-memory transcription factors associated with longevity, polyfunctionality and/or enhanced responsiveness to immune checkpoint blockade in cancer immunotherapy. Using an immunocompetent murine model of syngeneic solid tumors expressing truncated human-CD19 (hCD19t), we demonstrate that ectopic expression of pro-memory transcription factors in the 4th generation transcription factor-CARs (TF-CARs) augments CAR T cell therapeutic efficacy against solid tumors. Amongst CAR T cells recovered from established solid tumors, the 4th generation TF-CAR T cells retained a memory-precursor effector (MPEC) T cell like phenotype, while the 3rd generation CAR T cells largely displayed a terminally exhausted phenotype (TEx). Importantly, the inclusion of select pro-memory transcription factors did not compromise CAR T cell proliferation or expression of the effector cytokines IFN-γ and TNFα following restimulation. In addition to superior control of primary tumors, the TF-CAR T cells instilled greater long-term protection from tumor relapse in mice, which cleared primary tumors (6/7) compared to 3rd generation CAR T cells (3/7). Collectively, these data present incorporation of pro-memory transcription factors as an attractive engineering strategy to intrinsically combat CAR T cell exhaustion in the TME and promote long-term CAR T cell memory and protection from tumor relapse. Citation Format: Steven J. Reed, Samantha J. Tower, Ryma Toumi, Vandana Kalia, Surojit Sarkar. Incorporation of pro-memory transcription factors in fourth generation CARs enhances CAR T cell immunotherapy against solid tumors in an immunocompetent murine model [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 5580.

Vitamin D, a key nutrient/prohormone classically associated with skeletal health, is also an important immunomodulator, with pleotropic effects on innate and adaptive immune cells. Outcomes of several chronic, autoimmune, and infectious diseases are linked to vitamin D. Emergent correlations of vitamin D insufficiency with coronavirus‐induced disease 2019 (COVID‐19) severity, alongside empirical and clinical evidence of immunoregulation by vitamin D in other pulmonary diseases, have prompted proposals of vitamin D supplementation to curb the COVID‐19 public health toll. In this review paper, we engage an immunological lens to discuss potential mechanisms by which vitamin D signals might regulate respiratory disease severity in severe acute respiratory syndrome coronavirus 2 (SARS‐CoV2) infections, vis a vis other pulmonary infections. It is proposed that vitamin D signals temper lung inflammatory cascades during SARS‐CoV2 infection, and insufficiency of vitamin D causes increased inflammatory cytokine storm, thus leading to exacerbated respiratory disease. Additionally, analogous to studies of reduced cancer incidence, the dosage of vitamin D compounds administered to patients near the upper limit of safety may serve to maximize immune health benefits and mitigate inflammation and disease severity in SARS‐CoV2 infections. We further deliberate on the importance of statistically powered clinical correlative and interventional studies, and the need for in‐depth basic research into vitamin D‐dependent host determinants of respiratory disease severity., Graphical Abstract Review on Vitamin D‐dependent immunoregulatory mechanisms underlying respiratory disease severity in SARS‐CoV2 infections.

c-Myc is a major transcription factor expressed upon activation of CD8 T cells that drives their clonal expansion and metabolism. The role of c-Myc in regulating the outcome of exhausted CD8 T cells during chronic infection and cancer remains poorly defined. In this study, we used a unique model of c-Myc conditional ablation following antigen priming of CD8 T cells to investigate whether c-Myc exerts a role in CD8 T cell exhaustion in chronic antigenic settings. Our studies show that targeted deficiency of c-Myc in antigen-specific CD8 T cells resulted in decreased cell numbers but promoted the generation of stem-cell-like TCF-1+ less exhausted CD8 T cells. These cells are key therapeutic target for overcoming CD8+ T cells exhaustion by providing the proliferative burst after PD1 checkpoint blockade immunotherapy (CBI). We, therefore combined anti-PD-L1 therapy and c-Myc inhibition to determine whether c-Myc inhibition might synergize to improve the therapeutic outcome. Our data using c-Myc inhibitor alone revealed an increased frequency of TCF-1+ CD8 T cells. However, the number of antigen specific CD8 T cells and the production of effector cytokines (IFN-γ and TNF-α) remained identical to untreated group. Notably, the combination of c-Myc inhibitor and PD-1 CBI significantly promoted the proliferation of antigen-specific CD8 T cells and the production of IFN-γ and TNF-α. Overall, findings support a role for c-Myc in defining the fate of exhausted CD8 T cells, and illustrate a potential pathway for the optimization of check point blockade immunotherapy of chronic infections and cancer. Supported by NIH R21AI154363 NIH R01AI32819

IL-2 signaling at the time of CD8 T cell activation is a critical factor dictating effector and memory CD8 T cell differentiation. However, how IL-2 signaling impacts exhausted CD8 T cell fate outcomes remains poorly defined. Here, we address the role of IL-2 signaling in regulating the differentiation of the stem-like and terminally exhausted CD8 T cells. Prolonged expression of the high affinity IL-2 receptor (IL-2Ra, CD25), and resultant increase in IL-2 signaling drives terminal differentiation of effector cells during acute infection. We hypothesized that curtailing this signal would prevent terminal differentiation and promote the formation of less exhausted stem-cell like TCF-1+CD8T cells in chronic antigenic setting. Our studies show that blocking IL-2 signals at the early phase of chronic infection significantly increased the proportions of TCF-1+CD8T cells. Similarly, knocking down Il2ra augmented the development of TCF-1+CD8T cells. The therapeutic success of PD-1/PD-L1 checkpoint blockade immunotherapy (CBI) relies primarily on stem-cell like cells for vigorous expansion and effector responses. Thus, we next investigated how FACS-purified CD25Hi and CD25Lo cells adoptively transferred into infection-matched chronically infected recipients respond to PD-1 CBI. In contrast to CD25Hi cells, CD25Lo cells underwent vigorous proliferation after PD-1 CBI and produced high amount of effector molecules (GzmB, TNF-α and IFN-γ). Collectively, these data highlight the role of IL-2 signaling in the regulation of CD8 T cell exhaustion, which may be therapeutically exploited to optimize treatments of chronic infections and cancers with immune checkpoint inhibitors. Supported by NIH R21AI154363 NIH R01AI32819

Effector CAR T cell persistence and transition to memory have been widely associated with long-term protection from hematological tumor relapse. In the context of solid tumors, anti-tumor T cell exhaustion in the tumor microenvironment (TME) leads to T cell dysfunction and impaired memory formation. However, in solid tumor models a vast number of infused CAR T cells traffic to lymphoid organs (such as spleen and nondraining lymph nodes), which are bereft of CAR-antigen. The fate of these CAR T cells which have undergone ex vivo activation, viral transduction, and expansion regimens prior to transfer back into patients remains unclear. Using a third generation CAR targeting human(h)CD19t, and syngeneic murine tumor lines expressing hCD19t tumor-associated antigen, we show that CAR T cells residing in antigen-free sites of tumor-bearing immunocompetent mice transition to fully functional memory cells. Using viral infection-induced memory T cells as a benchmark, memory CAR T cells displayed hallmark memory characteristics including homeostatic proliferation, rapid polyfunctional cytokine production upon restimulation, and secondary expansion in response to cognate antigenic stimulation. Finally, memory CAR T cells in naive mice, or in mice with primary tumor remission, were protected against lethal rechallenge with heterologous hCD19t-expressing tumors. Results from this study are among the first to directly compare canonical virus-induced memory T cells with memory CAR T cells, and provide valuable insight into the characteristics and function of memory CAR T cells in an immunocompetent model for solid tumors. Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under Award Number F32CA265056, and the Pediatric Cancer Research Foundation

T cell exhaustion in the tumor microenvironment (TME) has hindered the success of T cell-based immunotherapy against multiple types of solid tumors. To combat this exhaustion, we have engineered a CD19-targeted fourth generation CAR construct to express the pro-memory transcription factor Inhibitor of DNA Binding 3 (Id3). Id3 critical for the formation of long-lived memory T cells in viral infection models, and retrovirally-induced expression of Id3 in T cells has been shown to enhance the survival of effector T cells in both acute infection and tumor models.Using an immunocompetent murine model of syngeneic solid tumors expressing truncated human-CD19 (hCD19t), we demonstrate that ectopic expression of Id3 via 4th generation CARs containing Id3 (Id3-CAR) augments CAR T cell therapeutic efficacy against solid tumors. Amongst cells recovered from established solid tumors, Id3-CAR T cells retained a memory-precursor effector cell (MPEC) like phenotype including elevated expression of TCF-1, while the 3rd generation CAR T cells largely displayed a terminally exhausted (TEx) phenotype. Importantly, the inclusion of Id3 did not compromise CAR T cell proliferation or expression of the effector cytokines IFN-γ and TNFα following restimulation. In addition to superior control of primary tumors, the Id3-CAR T cells instilled greater long-term protection from tumor relapse in mice, which cleared primary tumors (6/7) compared to 3rd generation CAR T cells (3/7). Highlighting Id3 as a candidate, these data present incorporation of pro-memory transcription factors as an attractive engineering strategy to intrinsically combat CAR T cell exhaustion in the TME and promote long-term CAR T cell memory and protection from tumor relapse. Research reported in this publication was supported by the National Cancer Institute of the National Institutes of Health under Award Number F32CA265056, and Pediatric Cancer Research Foundation.

Differential interleukin-2 (IL-2) signaling and production are associated with disparate effector and memory fates. Whether the IL-2 signals perceived by CD8 T cells come from autocrine or paracrine sources, the timing of IL-2 signaling and their differential impact on CD8 T cell responses remain unclear. Using distinct models of germline and conditional IL-2 ablation in post-thymic CD8 T cells, this study shows that paracrine IL-2 is sufficient to drive optimal primary expansion, effector and memory differentiation, and metabolic function. In contrast, autocrine IL-2 is uniquely required during primary expansion to program robust secondary expansion potential in memory-fated cells. This study further shows that IL-2 production by antigen-specific CD8 T cells is largely independent of CD4 licensing of dendritic cells (DCs) in inflammatory infections with robust DC activation. These findings bear implications for immunizations and adoptive T cell immunotherapies, where effector and memory functions may be commandeered through IL-2 programming.

Background: Anthropometric and physiologic properties have influence on exercise performance in sepak takraw and physiological profiles of athletes differ between sports and playing positions. Objectives: Present study was aimed to determine the effect of playing position on anthropometric, body composition and physical fitness parameters in young male sepak takraw players. Methods: The cross-sectional study was conducted on three positional groups of total 44 male Indian sepak takraw players (spiker/killer, n = 14; tekong, n = 14; and feeder, n = 16) of national level. Anthropometric parameters, body composition profile, physical fitness variables including 30m flying, 6X10m shuttle run, vertical jump, sit up, endurance and anaerobic power were assessed by following standard protocols. Results: Significant differences were observed in height, weight and leg length among the positional groups. Almost all body composition parameters were significantly (P < 0.01) higher among ‘tekong’ position except fat mass. Handgrip strength, back strength, vertical jump, sit up, VO2max, explosive leg power and maximum anaerobic power were found to be significantly differed among the three positional groups of sepak takraw. Regression prediction model identified back strength (P = 0.034), peak explosive power (P < 0.001) and leg length (P < 0.001) as the significant predictor of vertical jump (R2 = 0.930 and P < 0.001). Conclusions: Players of ‘tekong’ position were better in anthropometric and body composition profiles than ‘spiker and feeder’. Whereas, ‘spiker’ position was found to have higher back strength, explosive power, vertical jump and sit up abilities. Regression model identifies back strength, peak explosive power and leg length as the significant predictors of performance in sepak takraw.

Introduction High-intensity interval training (HIIT) has previously been reported having the effect of training period on altering oxidant status, muscle damage and performance. The present study was aimed to understand and evaluate the adaptive response of 8 weeks HIIT on muscle damage indices, inflammatory markers, oxidative stress variables and physical fitness parameters. Methods Forty young endurance male players [i.e., football (n = 20) and field hockey (n = 20)] were recruited under two groups i.e., control and HIIT. 8 weeks long 3 h/day of sprint-HIIT was intervened for thrice/week. HIIT workouts includes total 4 sets/session (divided into 2 phase × 2 sets × 2 min) of all-out sprint workout (at 90–95% of HRmax with work: rest = 1:1). Muscle damage indices (CK and LDH), inflammatory markers (IL-6 and TNF-α), oxidative stress variables (MDA, SOD, GSH and GPx) and physical fitness variables (VO2max, Wpeak and VJ) were assessed via following standard protocols. Result The HIIT resulted to significantly(p Conclusion Antioxidant redox imbalance confers inflammatory oxidative stress condition which further leads to muscle damage and that may cause due to HIIT induced temporary hypoxic condition which contrarily induced overtraining effect but with improving performance variables.

Differential IL-2 signaling is implicated in memory CD8 T cell fates. Autocrine IL-2 production is a hallmark of polyfunctional, long-lived lymphoid memory cells. Whether the IL-2 signals perceived by CD8 T cells come from autocrine or paracrine sources, the timing of these signals, and their differential impact on the qualitative outcomes of CD8 T cell responses are not clearly understood. We used two distinct models of germline and conditional autocrine IL-2 ablation in post-thymic CD8 T cells to specifically dissect the requirement and timing of autocrine IL-2. Our studies show that autocrine IL-2 is uniquely required during primary CTL responses to program the key memory property of robust recall responses. Ablation of autocrine IL-2 in fully differentiated wild type memory cells immediately prior to rechallenge did not impact their recall expansion potential. Furthermore, we found that paracrine IL-2 could effectively supplant autocrine IL-2 for primary expansion, effector differentiation and memory maintenance, but not recall expansion. Likewise, our data highlight the redundancy of autocrine and paracrine IL-2 in optimally driving equivalent metabolic programs associated with effector and memory states. While CD4 licensing of DCs is needed for optimal programming of autocrine IL-2 in CD8 T cells, we found that the requirement for CD4 help is less critical for autocrine IL-2 production in the context of inflammatory infections. These findings have implications in adoptive T cell immunotherapies where expansion of engineered cells is largely driven by paracrine IL-2, which can potentially impact their long-term sentinel function.

The general paradigm is that the tumor microenvironment is non-conducive to sustaining productive cytotoxic T cell responses due to suppressive and counter-immunity T cell extrinsic factors. It has been shown that the tumor microenvironment can harbor large numbers of bystander memory cells. However, it is not very clear if given the right stimulation these bystander memory cells can be commandeered to utilize the available resources of the tumor microenvironment, get optimally activated and elaborate productive immune responses. This has wide-ranging implications in hijacking the tumor-resident bystander memory cells for efficacious tumor control. In this study we show that the tumor microenvironment is capable of harboring ultra-functional bystander memory cells, which display complete panoply of memory markers. These tumor–resident bystander memory cells are not only capable of elaborating robust ex vivo and transplantable memory T cell functions, but they can also efficiently utilize the resources within the tumor microenvironment to elaborate in situ recall effector functions. We further show that these memory T cells utilize CXCR3 as a key chemokine receptor for trafficking to the tumor sites. Our data identify bystander tumor-resident memory cells as Spartan warriors to mediate enhanced tumor control during adoptive T cell immunotherapy.

BackgroundScientific researches available on anthropometric and physiological characteristics of female rugby players in India are very scanty.ObjectivesThe present study was aimed to evaluate and compared different anthropometric, body composition and physical fitness parameters and also to investigate whether any distinctive characteristics exist among above variables according to their specific playing position.MethodsTwenty five Indian national women rugby players (forward, n = 12, mean age = 20.9 ± 3.30 and back, n = 13, mean age = 19.7 ± 2.48) of pre-competitive phase were chosen as subject from the national coaching camp. They were evaluated for various anthropometric, body composition parameters, hemoglobin, blood glucose level and physical fitness profile by following standard procedure.ResultsForward players had significantly higher body weight, BMI, fat percentage, sum of skinfold, total body potassium, total body calcium and glycogen content as compared to the back. Whereas, players of back position had significantly higher relative muscle mass (MM/Wt.), relative body cell mass (BCM/Wt.), agility, standing broad jump, VO2max and relative peak anaerobic power than their forward counterparts. Linear regression model identified agility (β = 0.59) and sum of skinfold (β = -0.45) as significant predictor of peak anaerobic power (adjusted R2 = 0.61) of these players.ConclusionsPresent study indicated the players of back position were faster, more agile and aerobically fit with having lower body weight and fat percent as compared to their forward counterparts. Whereas, principal component analysis identified BMI, sum of skinfold and agility as the main confounding variables to categorize the present rugby players between forward and back positional groups.

Background: The dominant role of physical training is reflected through the exercise induced physiological and biochemical alterations in athletes. Objective: Present study was aimed to evaluate the variability of muscular cell damage indices among young Indian female players of different sports discipline with respect to their training. Methods: Body height, weight and body mass index were assessed in 34 female gymnasts, 30 athletes, 28 swimmers and 35 footballers following standard procedures. Fat mass, fat free mass and muscle mass (MM) were recorded . Maximum aerobic capacity (VO2max) was assessed through beep test. Venous blood samples were analyzed for muscle damage markers including serum creatine kinase MB (CK-MB), lactate dehydrogenase (LDH); and cortisol . Results: Swimmers were the tallest and heaviest whereas gymnasts were the shortest and lightest (p

It is now well known that in the mammalian body vitamin D is converted by successive hydroxylations to 1,25-dihydroxyvitamin D (1,25D), a steroid-like hormone with pleiotropic properties. These include important contributions to the control of cell proliferation, survival and differentiation, as well as the regulation of immune responses in disease. Here, we present recent advances in current understanding of the role of 1,25D in myelopoiesis and lymphopoiesis, and the potential of 1,25D and analogs (vitamin D derivatives; VDDs) for the control of hematopoietic malignancies. The reasons for the unimpressive results of most clinical studies of the therapeutic effects of VDDs in leukemia and related diseases may include the lack of a precise rationale for the conduct of these studies. Further, clinical trials to date have generally used extremely heterogeneous patient populations and, in many cases, small numbers of patients, generally without controls. Although low calcemic VDDs have been used and combined with agents that can increase the leukemia cell killing or differentiation effects in acute leukemias, the sequencing of agents used for combination therapy should to be more clearly delineated. Most importantly, it is recommended that in future clinical trials the rationale for the basis of the enhancing action of drug combinations should be clearly articulated and the effects on anticancer immunity should also be evaluated.

Immune memory cells are poised to rapidly expand and elaborate effector functions upon reinfection yet exist in a functionally quiescent state. The paradigm is that memory T cells remain inactive due to lack of T cell receptor (TCR) stimuli. Here, we report that regulatory T (Treg) cells orchestrate memory T cell quiescence by suppressing effector and proliferation programs through inhibitory receptor, cytotoxic-T-lymphocyte-associated protein-4 (CTLA-4). Loss of Treg cells resulted in activation of genome-wide transcriptional programs characteristic of effector T cells and drove transitioning as well as established memory CD8(+) T cells toward terminally differentiated KLRG-1(hi)IL-7Rα(lo)GzmB(hi) phenotype, with compromised metabolic fitness, longevity, polyfunctionality, and protective efficacy. CTLA-4 functionally replaced Treg cells in trans to rescue memory T cell defects and restore homeostasis. These studies present the CTLA-4-CD28-CD80/CD86 axis as a potential target to accelerate vaccine-induced immunity and improve T cell memory quality in current cancer immunotherapies proposing transient Treg cell ablation.

Highly active killer T cells mediate a stable standoff during controlled persistent infections. In this issue of Immunity , Robey and colleagues describe a unique antigen-addicted T cell population bearing characteristics of both effector and memory CD8 + T cells that provides a continuous supply of potent killer T cells to curb Toxoplasma gondii growth during latency.

Progress towards a safe and effective vaccine for the prevention of tularemia has been hindered by a lack of knowledge regarding the correlates of protective adaptive immunity and a lack of tools to generate this knowledge. CD8+T cells are essential for protective immunity against virulent strains ofFrancisella tularensis, but to-date, it has not been possible to study these cells in a pathogen-specific manner. Here, we report the development of a tool for expression of the model antigen ovalbumin (OVA) inF. tularensis, which allows for the study of CD8+T cell responses to the bacterium. We demonstrate that in response to intranasal infection with theF. tularensisLive Vaccine Strain, pathogen-specific CD8+T cells expand after the first week and produce IFN-γ but not IL-17. Effector and central memory subsets develop with disparate kinetics in the lungs, draining lymph node and spleen. Notably,F. tularensis-specific cells are poorly retained in the lungs after clearance of infection. We also show that intranasal vaccination leads to more pathogen-specific CD8+T cells in the lung-draining lymph node compared to scarification vaccination, but that an intranasal booster overcomes this difference. Together, our data show that this novel tool can be used to study multiple aspects of the CD8+T cell response toF. tularensis. Use of this tool will enhance our understanding of immunity to this deadly pathogen.

Early after priming, effector CD8 T cells are distinguished into memory precursor and short-lived effector cell subsets (MPECs and SLECs). Here, we delineated a distinct in vivo heterogeneity in killer cell lectin-like receptor G1 (KLRG-1) expression, which was strongly associated with diverse MPEC and SLEC fates. These in vivo MPECs and SLECs expressed equivalent levels of cytotoxic molecules and effector cytokines. Using a unique in vivo degranulation assay, we found that the MPECs and SLECs similarly encountered infected target cells and elaborated equivalent levels of cytotoxicity in vivo. These data provide direct in vivo evidence that memory-fated cells pass through a robust effector phase. Additionally, the preferential localization of the MPECs in the lymph nodes, where a lesser degree of cytotoxicity was elaborated, suggests that the MPECs may be protected from excessive stimulation and terminal differentiation by virtue of their differential tissue localization. These data provide novel mechanistic insights into the linear decreasing potential model of memory differentiation.

IL-2 is a key driver of T cell responses critical for the conversion of resting antigen specific T cells into glycolytic powerhouses that undergo massive expansion and elicit cytolytic function to clear infections. Recent studies have used ex vivo antigen challenge combined with somatic IL-2 deletion to show that both paracrine and autocrine IL-2 signals are indispensable for the formation of highly functional memory CD8 T cells. Despite this critical role of autocrine IL-2 for memory T cell fate, the in vivo history and fate of autocrine IL-2 producing CD8 T cells remains unknown. We generated a mouse model in which autocrine IL-2 production leads to transcription of TdTomato, leading to an indelibly marked T cell. Following acute infection, a proportion of antigen specific CD8 T cells made autocrine IL-2, however, more than half of the T cell response had no history of IL-2 production. Furthermore, contrary to their ex vivo functionality, memory precursor and terminal effector T cells had similar history of in vivo IL-2 production. Despite autocrine IL-2 production being a benchmark of highly functional memory T cells, we observed no increase in the ex vivo polyfunctionality of T cells that produced in vivo IL-2 during infection. Finally, while strong TCR engagement is critical for the recruitment of robust T cell responses, we saw that in vivo autocrine IL-2 production is more effectively elicited by lower affinity TCR signals. Thus, we see that while proportion of T cells rely on autocrine IL-2 during infection, the ability of a CD8 T cell to recruit autocrine IL-2 function heavily depends on the avidity of the TCR to the peptide-MHC complex. Together these data provide the first insights into the in vivo function of IL-2 in antigen specific CD8 T cells.

Chimeric antigen receptor (CAR) T cells therapies have made substantial progress in treating blood cancers in humans. However, CAR T therapies currently have several limitations, including the lack of a persistent anti-tumor memory population to limit relapse and poor efficacy against solid tumors. Many studies on CAR T therapies have used immunocompromised xenograft mouse models, limiting the translational impact of the results. Furthermore, mouse models have largely focused on cancers in the blood while neglecting solid tumors. To improve the relevance of preclinical studies of CAR T cell therapy and to understand the mechanisms underlying some of the current CAR T therapy limitations, we have created a novel immunocompetent mouse model. This model consists of three components: murine CD8 T cells expressing the human CD19 CAR, syngeneic murine tumor cell lines expressing a truncated human CD19 (CD19t) serving as the tumor antigen, and an immunocompetent mouse tolerized to human CD19t and the CD19 CAR. Our results demonstrate that the CD19 CAR T cells can efficiently target the CD19t-expressing tumor lines. Furthermore, tolerized recipient mice do not reject implanted syngeneic CD19t-expressing tumors or adoptively transferred CD19 CAR-expressing T cells. This model now provides a platform to study the in vivo biology of CAR T cells with respect to localization, exhaustion, and functional reinvigoration; to investigate best approaches to treating multiple solid tumors with distinct microenvironments; and to test the preclinical efficacy of novel CAR T strategies.

The high affinity IL-2 receptor (IL-2Ra, CD25) is a key activation marker, which is transiently and heterogeneously expressed during the priming . While IL-2 signals are deemed critical for expansion of antigen-specific CD8 T cells, the role of IL-2 in programing the fate of exhausted CD8 T cells in vivo is poorly understood. Exhausted CD8 T cells are broadly divided into 2 subsets: one with brightest levels of inhibitory receptors such as PD-1, and lower mitochondrial spare respiratory capacity (SRC) – these cells are terminally exhausted and are largely unresponsive to PD-1 checkpoint blockade immunotherapy. The other subset has intermediate levels of inhibitory receptors and retains higher mitochondrial SRC – these are the clinically relevant cells that are highly responsive to PD-1 blockade. Based on the ability of IL-2 signals to drive glycolytic burnout in T cells, we hypothesized that the stochastic heterogeneity in in vivo assimilation of IL-2 signals during priming drives distinct CD8 T cell fates late during exhaustion. We sorted in vivo primed CD8 T cells based on their heterogeneity in CD25 expression, and adoptively transferred them into infection-matched chronically infected recipients. We observed that the exhausted CD8 T cells generated from CD25Hi donor cells phenotypically and functionally resembled terminally exhausted cells. Furthermore, knocking down the expression of CD25 or Special AT-rich sequence binding protein 1 (SATB1), which is known to regulate CD25 expression, resulted in better survival and function of exhausted CD8 T cells. Our data underscores the critical role of IL-2 signals in the exhaustion of CD8 T cells and has clinical significance in current combination checkpoint blockade immunotherapies.

Long-lived plasma cells that reside in the bone marrow constitutively produce antibody in the absence of antigen and are the cellular basis of durable humoral immunity. The generation of these long-lived plasma cells depends upon a series of highly orchestrated interactions between antigen-specific CD4 T cells and B cells and the formation of germinal centers (GCs). In this study, we have examined the role of the cytokine interleukin-21 (IL-21) in regulating humoral immunity during acute viral infections. Using IL-21 receptor-deficient (IL-21R −/− ) mice, we found that virus-specific CD4 T cells were generated after infection with lymphocytic choriomeningitis virus (LCMV) and that these CD4 T cells differentiated into T follicular helper (T FH )-like cells in the absence of IL-21 signaling. There was also no defect in the formation of GCs, although after day 15 these GCs disappeared faster in IL-21R −/− mice than in wild-type mice. Isotype switching and the initial LCMV-specific IgG response were normal in IL-21R −/− mice. However, these mice exhibited a profound defect in generating long-lived plasma cells and in sustaining antibody levels over time. Similar results were seen after infection of IL-21R −/− mice with vesicular stomatitis virus and influenza virus. Using chimeric mice containing wild-type or IL-21R −/− CD4 T cells and B cells, we showed that both B and CD4 T cells need IL-21 signaling for generating long-term humoral immunity. Taken together, our results highlight the importance of IL-21 in humoral immunity to viruses.

The precise microRNAs and their target cellular processes involved in generation of durable T-cell immunity remain undefined. Here we show a dynamic regulation of microRNAs as CD8 T cells differentiate from naïve to effector and memory states, with short-lived effectors transiently expressing higher levels of oncogenic miR-17-92 compared with the relatively less proliferating memory-fated effectors. Conditional CD8 T-cell-intrinsic gain or loss of expression of miR-17-92 in mature cells after activation resulted in striking reciprocal effects compared with wild-type counterparts in the same infection milieu-miR-17-92 deletion resulted in lesser proliferation of antigen-specific cells during primary expansion while favoring enhanced IL-7Rα and Bcl-2 expression and multicytokine polyfunctionality; in contrast, constitutive expression of miR-17-92 promoted terminal effector differentiation, with decreased formation of polyfunctional lymphoid memory cells. Increased proliferation upon miR-17-92 overexpression correlated with decreased expression of tumor suppressor PTEN and increased PI3K-AKT-mTOR signaling. Thus, these studies identify miR17-92 as a critical regulator of CD8 T-cell expansion and effector and memory lineages in the physiological context of acute infection, and present miR-17-92 as a potential target for modulating immunologic outcome after vaccination or immunotherapeutic treatments of cancer, chronic infections, or autoimmune disorders.

PD-1 is an inhibitory receptor that has a major role in T cell dysfunction during chronic infections and cancer. While demethylation of the PD-1 promoter DNA is observed in exhausted T cells isolated from chronically infected individuals, little is known about when this stable demethylation of PD-1 promoter DNA is programmed during the course of a chronic infection. To assess if PD-1 promoter DNA demethylation is impacted by prolonged stimulation during effector phase of chronic infection, we adoptively transferred virus-specific day 8 effector CD8 T cells from mice infected with lymphocytic choriomeningitis virus (LCMV) clone 13 into recipient mice that had cleared an acute infection. We observed that LCMV-specific CD8 T cells from chronically infected mice maintained their surface expression of PD-1 even after transfer into acute immune mice until day 45 posttransfer. Interestingly, the PD-1 transcriptional regulatory region continued to remain unmethylated in these donor CD8 T cells generated from a chronic infection. The observed maintenance of PD-1 surface expression and the demethylated PD-1 promoter were not a result of residual antigen in the recipient mice, because similar results were seen when chronic infection-induced effector cells were transferred into mice infected with a variant strain of LCMV (LCMV V35A) bearing a mutation in the cognate major histocompatibility complex class I (MHC-I) epitope that is recognized by the donor CD8 T cells. Importantly, the maintenance of PD-1 promoter demethylation in memory CD8 T cells was coupled with impaired clonal expansion and higher PD-1 re-expression upon secondary challenge. These data show that the imprinting of the epigenetic program of the inhibitory receptor PD-1 occurs during the effector phase of chronic viral infection. IMPORTANCE Since PD-1 is a major inhibitory receptor regulating T cell dysfunction during chronic viral infection and cancers, a better understanding of the mechanisms that regulate PD-1 expression is important. In this work, we demonstrate that the PD-1 epigenetic program in antigen-specific CD8 T cells is fixed during the priming phase of chronic infection.

An important question in memory development is understanding the differences between effector CD8 T cells that die versus effector cells that survive and give rise to memory cells. In this study, we provide a comprehensive phenotypic, functional, and genomic profiling of terminal effectors and memory precursors. Using killer cell lectin-like receptor G1 as a marker to distinguish these effector subsets, we found that despite their diverse cell fates, both subsets possessed remarkably similar gene expression profiles and functioned as equally potent killer cells. However, only the memory precursors were capable of making interleukin (IL) 2, thus defining a novel effector cell that was cytotoxic, expressed granzyme B, and produced inflammatory cytokines in addition to IL-2. This effector population then differentiated into long-lived protective memory T cells capable of self-renewal and rapid recall responses. Experiments to understand the signals that regulate the generation of terminal effectors versus memory precursors showed that cells that continued to receive antigenic stimulation during the later stages of infection were more likely to become terminal effectors. Importantly, curtailing antigenic stimulation toward the tail end of the acute infection enhanced the generation of memory cells. These studies support the decreasing potential model of memory differentiation and show that the duration of antigenic stimulation is a critical regulator of memory formation.

The memory T cell pool serves as a relatively long-lived heterogeneous repository of antigen-experienced T cells that "remember" previous encounters with antigen. While heterogeneity in the memory T cell pool is now well established, signals regulating the generation of this memory T cell heterogeneity are not fully understood. Two articles in this issue of the European Journal of Immunology highlight the importance of the strength of antigenic stimulation in regulating the generation of phenotypically and functionally distinct CD4+ T cell memory subsets. New insights are also provided into key molecular players that likely mediate differences in homeostatic and secondary expansion between the memory subsets. See accompanying articles: http://dx.doi.org/10.1002/eji200737611 and http://dx.doi.org/10.1002/eji200737852

A key feature of the adaptive immune system is the ability of cytotoxic T lymphocytes (CTLs) to eliminate infected or diseased target cells in an antigen-specific manner, and then ‘remember’ the first encounter by way of long-lived memory CD8 T cells. Memory cells are capable of rapidly culling any future incursions owing to their heightened responsiveness and ready localization at portals of pathogen entry and sites of immune activation. Thus, understanding how memory CD8 T cells develop has been of long-standing interest to the field. This knowledge is not only significant for our vaccine development efforts aimed at inducing lifelong protective immunity, but also bears relevance to selectively potentiating effector CTLs during immunotherapy of cancer or chronic infections, or suppressing deleterious CTL functions during autoimmunity and transplantation. Here we review our current understanding of the mechanisms underlying the development and maintenance of robust CD8 T cell immunological memory that have emerged from the murine models of acute infections, and also discuss how some of these findings have been extended to humans. Recent discoveries in cellular and metabolic, transcriptional, and epigenetic control of memory with a diverse array of localization and functional properties are presented in the context of promising immunological interventions for treatment and vaccination.

The discovery of vitamin D receptor (VDR) expression in immune cells has opened up a new area of research into immunoregulation by vitamin D, a niche that is distinct from its classical role in skeletal health. Today, about three decades since this discovery, numerous cellular and molecular targets of vitamin D in the immune system have been delineated. Moreover, strong clinical associations between vitamin D status and the incidence/severity of many immune-regulated disorders (e.g. infectious diseases, cancers and autoimmunity) have prompted the idea of using vitamin D supplementation to manipulate disease outcome. While much is known about the effects of vitamin D on innate immune responses and helper T (T(H)) cell immunity, there has been relatively limited progress on the frontier of cytotoxic T lymphocyte (CTL) immunity--an arm of host cellular adaptive immunity that is crucial for the control of such intracellular pathogens as human immunodeficiency virus (HIV), tuberculosis (TB), malaria, and hepatitis C virus (HCV). In this review, we discuss the strong historical and clinical link between vitamin D and infectious diseases that involves cytotoxic T lymphocyte (CTL) immunity, present our current understanding as well as critical knowledge gaps in the realm of vitamin D regulation of host CTL responses, and highlight potential regulatory connections between vitamin D and effector and memory CD8 T cell differentiation events during infections.

Cryopreservation of peripheral blood mononuclear cells (PBMC) from animal model studies and clinical trials is utilized as a primary method for long-term storage of PBMC for future in vitro and in vivo applications. The objective of this study was to define the mechanistic pathways involved in cryopreservation-induced apoptosis of CD4+ T-cells in PBMC, and to evaluate a cytokine treatment of the cryopreserved samples to rescue apoptosis for the potential future use of the cryopreserved PBMC. Using cryopreserved PBMC samples isolated from naive and Simian immunodeficiency virus (SIV)-infected rhesus macaques as a model, frozen PBMC showed significantly increased levels of apoptosis-induced CD4+ T-cell death compared to fresh PBMC over a 5-day culture period as detected by Annexin V/PI and trypan blue staining. Mechanistic studies using a broad-spectrum caspase inhibitor z-VAD demonstrated a crucial involvement of caspases in cryopreservation-induced apoptosis of CD4+ T-cells. Furthermore, the ability of z-VAD to inhibit both mitochondrial membrane perturbation and apoptotic cell death implicated the involvement of caspase-mediated mitochondrial membrane damage in cryopreservation-induced apoptosis of CD4+ T-cells. Due to their known properties to promote T-cell survival and inhibit apoptosis, we evaluated the ability of IL-2, IL-4, and IL-7 combination cytokine treatment of the cryopreserved cells to rescue apoptosis of the CD4+ T-cells. The cytokine treatment resulted in a significant inhibition (p

Dysregulation of cytokines and chemokines during human immunodeficiency virus 1 (HIV-1) and simian immunodeficiency virus (SIV) infection is thought to be critical in the progression of acquired immunodeficiency syndrome (AIDS). To evaluate the potential role of Th1-agonist chemokines in disease progression during AIDS, we assessed CXCL9/MIG and CXCL10/IP-10 expression simultaneously in the periphery and lymphoid tissues of SIV-infected animals at a single-cell level by flow cytometry. We optimized intracellular staining and analysis of CXCL9/MIG and CXCL10/IP-10 production in human leukocyte antigen (HLA)-DR + macaque cells by flow cytometry using cross-reactive antibodies against human chemokines. We observed an upregulation of CXCL9/MIG and CXCL10/IP-10 production in both the periphery and lymph nodes of infected animals compared with naive controls. Animals with higher viral loads had higher levels of CXCL9/MIG and CXCL10/ IP-10 producing cells compared with animals with low viral loads. Analysis of cells bearing the receptor (CXCR3) for CXCL9/MIG and CXCL10/IP-10 revealed increased number of CXCR3 + cells in the lymph nodes of infected animals. Importantly, an inverse correlation (P < 0.05) between CXCL9/MIG and CXCL10/IP-10 production, both in the periphery and lymph nodes, and peripheral CD4 + T-cell numbers was observed. These findings provide further evidence that dysregulation of Th1 agonist chemokines might contribute to the ultimate immunopathology during AIDS.

It is now well known that in the mammalian body vitamin D is converted by successive hydroxylations to 1,25-dihydroxyvitamin D (1,25D), a steroid-like hormone with pleiotropic properties. These include important contributions to the control of cell proliferation, survival and differentiation, as well as the regulation of immune responses in disease. Here, we present recent advances in current understanding of the role of 1,25D in myelopoiesis and lymphopoiesis, and the potential of 1,25D and analogs (vitamin D derivatives; VDDs) for the control of hematopoietic malignancies. The reasons for the unimpressive results of most clinical studies of the therapeutic effects of VDDs in leukemia and related diseases may include the lack of a precise rationale for the conduct of these studies. Further, clinical trials to date have generally used extremely heterogeneous patient populations and, in many cases, small numbers of patients, generally without controls. Although low calcemic VDDs have been used and combined with agents that can increase the leukemia cell killing or differentiation effects in acute leukemias, the sequencing of agents used for combination therapy should to be more clearly delineated. Most importantly, it is recommended that in future clinical trials the rationale for the basis of the enhancing action of drug combinations should be clearly articulated and the effects on anticancer immunity should also be evaluated.

Virus-specific CD4+ T cells (Th) play a crucial role in the control of lentiviral replication. To better understand the epitope-specificity of CD4+ Th repertoire to the envelope glycoprotein (Env) of simian immunodeficiency virus (SIV), we analyzed Th responses to 20-mer overlapping Env peptides in eight genetically heterogeneous macaques chronically infected with live attenuated SIV. A set of 19 'broadly reactive' Th peptide-epitopes was defined from the distinct sets of responder peptides for individual macaques. The majority of broadly reactive peptide-epitopes (14 of 19) were uniformly distributed on the transmembrane (TM) domain of Env. Only five broadly reactive responder peptides localized to the surface domain (SU) of Env, and they were all confined to two non-glycosylated regions towards its carboxyl-terminus. This first comprehensive report of Env peptide-specific Th responses associated with attenuated SIV vaccine immunity indicates a profound influence of glycosylation on the development of Th responses and has important implications for acquired immunodeficiency syndrome (AIDS) vaccine development.

Ag-specific CD4+ Th cells play a key role in the development, maturation, and maintenance of pathogen-specific humoral and cellular immune responses. To define the fine specificity of broadly reactive Th responses associated with mature immunity in a lentiviral system, we analyzed peptide-specific Th responses in eight macaques chronically infected with a reference live attenuated SIV at 12–14 mo postinoculation. All macaques had stable immunocompetent Th cells at the time of analysis, and a unique array of Th responses to 20-mer overlapping peptides from envelope (Env) and Gag was identified for each macaque, which were then used to define a set of 31 broadly reactive peptide epitopes. Only 5 of the 31 broadly reactive Th epitope peptides mapped to the surface (SU) domain of Env. Interestingly, these were all confined to two conserved nonglycosylated regions toward the carboxyl terminus of SU, suggesting a structural influence of glycosylation on development of Th responses. Gag and the Env transmembrane proteins contained the majority of broadly reactive peptide epitopes (12 and 14 peptides, respectively), which were uniformly distributed throughout their sequence. This study defines for the first time broadly reactive Th epitope peptides of SIV Env and Gag proteins that are associated with enduring broadly protective vaccine immunity to attenuated SIV, which may be used for the design and evaluation of experimental vaccines. Moreover, the data suggest that extensive glycosylation of SU may provide yet another immune escape mechanism developed by lentiviruses to restrict the breadth of Th repertoire to SU, a major immunologically exposed protein of the virus.