Prior viral infection primes cross-reactive CD8+ T cells that respond to mouse heart allografts.

Introduction: Significant evidence suggests a connection between transplant rejection and the presence of high levels of pre-existing memory T cells. Viral infection can elicit viral-specific memory T cells that cross-react with allo-MHC capable of driving allograft rejection in mice. Despite these advances, and despite their critical role in transplant rejection, a systematic study of allo-reactive memory T cells, their specificities, and the role of cross-reactivity with viral antigens has not been performed. Methods: Here, we established a model to identify, isolate, and characterize cross-reactive T cells using Nur77 reporter mice (C57BL/6 background), which transiently express GFP exclusively upon TCR engagement. We infected Nur77 mice with lymphocytic choriomeningitis virus (LCMV-Armstrong) to generate a robust memory compartment, where quiescent LCMV-specific memory CD8+ T cells could be readily tracked with MHC tetramer staining. Then, we transplanted LCMV immune mice with allogeneic hearts and monitored expression of GFP within MHC-tetramer defined viral-specific T cells as an indicator of their ability to cross-react with alloantigens. Results: Strikingly, prior LCMV infection significantly increased the kinetics and magnitude of rejection as well as CD8+ T cell recruitment into allogeneic, but not syngeneic, transplanted hearts, relative to non-infected controls. Interestingly, as early as day 1 after allogeneic heart transplant an average of ~8% of MHC-tetramer+ CD8+ T cells expressed GFP, in contrast to syngeneic heart transplants, where the frequency of viral-specific CD8+ T cells that were GFP+ was <1%. These data show that a significant percentage of viral-specific memory CD8+ T cells expressed T cell receptors that also recognized alloantigens in vivo. Notably, the frequency of cross-reactive CD8+ T cells differed depending upon the viral epitope. Further, TCR sequences derived from cross-reactive T cells harbored distinctive motifs that may provide insight into cross-reactivity and allo-specificity. Discussion: In sum, we have established a mouse model to track viral-specific, allo-specific, and cross-reactive T cells; revealing that prior infection elicits substantial numbers of viral-specific T cells that cross-react to alloantigen, respond very early after transplant, and may promote rapid rejection.

Subset-specific and temporal control of effector and memory CD4+ T cell survival.

Following their proliferative expansion and differentiation into effector cells like Th1, Tfh, and T central memory precursors (Tcmp), most effector CD4+ T cells die, while some survive and become memory cells. Here, we explored how Bcl-2 family members controlled the survival of CD4+ T cells during distinct phases of mouse acute LCMV infection. During expansion, we found that Th1 cells dominated the response, downregulated expression of Bcl-2, and did not require Bcl-2 for survival. Instead, they relied on the anti-apoptotic protein, A1 for survival. Similarly, Th17 cells in an EAE model also depended on A1 for survival. However, after the peak of the response, CD4+ effector T cells required Bcl-2 to counteract Bim to aid their transition into memory. This Bcl-2 dependence persisted in established memory CD4+ T cells. Combined, these data show a temporal switch in Bcl-2 family-mediated survival of CD4+ T cells over the course of an immune response. This knowledge can help improve T cell survival to boost immunity and conversely, target pathogenic T cells.

Pro-Inflammatory Alterations of Circulating Monocytes in Latent Tuberculosis Infection.

Background: Latent tuberculosis infection (LTBI) has been associated with increased cardiovascular risk. We investigated the activation and pro-inflammatory profile of monocytes in individuals with LTBI and their association with coronary artery disease (CAD). Methods: Individuals 40-70 years old in Lima, Peru, underwent QuantiFERON-TB testing to define LTBI, completed a coronary computed tomography angiography to evaluate CAD, and provided blood for monocyte profiling using flow cytometry. Cells were stimulated with lipopolysaccharide to assess interleukin-6 (IL-6) and tumor necrosis factor (TNF)-α responses. Results: The clinical characteristics of the LTBI (n = 28) and non-LTBI (n = 41) groups were similar. All monocyte subsets from LTBI individuals exhibited higher mean fluorescence intensity (MFI) of CX3CR1 and CD36 compared with non-LTBI individuals. LTBI individuals had an increased proportion of nonclassical monocytes expressing IL-6 (44.9 vs 26.9; P = .014), TNF-α (62.3 vs 35.1; P = .014), and TNF-α+IL-6+ (43.2 vs 36.6; P = .042). Among LTBI individuals, CAD was associated with lower CX3CR1 MFI on classical monocytes and lower CD36 MFI across all monocyte subsets. In multivariable analyses, lower CD36 MFI on total monocytes (b = -0.17; P = .002) and all subsets remained independently associated with CAD in LTBI. Conclusions: Individuals with LTBI have distinct monocyte alterations suggestive of an exacerbated inflammatory response and tissue migration. Whether these alterations contribute to cardiovascular disease pathogenesis warrants further investigation.

Restoring tolerance to ß-cells in Type 1 diabetes: Current and emerging strategies.

Type 1 diabetes (T1D) results from insulin insufficiency due to islet death and dysfunction following T cell-mediated autoimmune attack. The technical feasibility of durable, functional autologous islet restoration is progressing such that it presents the most likely long-term cure for T1D but cannot succeed without the necessary counterpart of clinically effective therapeutic strategies that prevent grafted islets' destruction by pre-existing anti-islet T cells. While advances have been made in broad immunosuppression to lower off-target effects, the risk of opportunistic infections and cancers remains a concern, especially for well-managed T1D patients. Current immunomodulatory strategies in development focus on autologous Treg expansion, treatments to decrease antigen presentation and T effector (Teff) activation, and broad depletion of T cells with or without hematopoietic stem cell transplants. Emerging strategies harnessing the intensified DNA damage response present in expanding T cells, exacerbating their already high sensitivity to apoptosis to abate autoreactive Teff cells.

T cells instruct myeloid cells to produce inflammasome-independent IL-1ß and cause autoimmunity.

The cytokine interleukin (IL)-1ß is a key mediator of antimicrobial immunity as well as autoimmune inflammation. Production of IL-1ß requires transcription by innate immune receptor signaling and maturational cleavage by inflammasomes. Whether this mechanism applies to IL-1ß production seen in T cell-driven autoimmune diseases remains unclear. Here, we describe an inflammasome-independent pathway of IL-1ß production that was triggered upon cognate interactions between effector CD4+ T cells and mononuclear phagocytes (MPs). The cytokine TNF produced by activated CD4+ T cells engaged its receptor TNFR on MPs, leading to pro-IL-1ß synthesis. Membrane-bound FasL, expressed by CD4+ T cells, activated death receptor Fas signaling in MPs, resulting in caspase-8-dependent pro-IL-1ß cleavage. The T cell-instructed IL-1ß resulted in systemic inflammation, whereas absence of TNFR or Fas signaling protected mice from CD4+ T cell-driven autoimmunity. The TNFR-Fas-caspase-8-dependent pathway provides a mechanistic explanation for IL-1ß production and its consequences in CD4+ T cell-driven autoimmune pathology.

Extending Remission and Reversing New-Onset Type 1 Diabetes by Targeted Ablation of Autoreactive T Cells.

Preserving endogenous insulin production is clinically advantageous and remains a vital unmet challenge in the treatment and reversal of type 1 diabetes. Although broad immunosuppression has had limited success in prolonging the so-called remission period, it comes at the cost of compromising beneficial immunity. Here, we used a novel strategy to specifically deplete the activated diabetogenic T cells that drive pathogenesis while preserving not only endogenous insulin production but also protective immunity. Effector T (Teff) cells, such as diabetogenic T cells, are naturally poised on the edge of apoptosis because of activation-induced DNA damage that stresses the p53 regulation of the cell cycle. We have found that using small molecular inhibitors that further potentiate p53 while inhibiting the G2/M cell cycle checkpoint control drives apoptosis of activated T cells in vivo. When delivered at the onset of disease, these inhibitors significantly reduce diabetogenic Teff cells, prolong remission, preserve functional islets, and protect islet allografts while leaving naive, memory, and regulatory T-cell populations functionally untouched. Thus, the targeted manipulation of p53 and cell cycle checkpoints represents a new therapeutic modality for the preservation of islet ß-cells in new-onset type 1 diabetes or after islet transplant.

Dying to protect: cell death and the control of T-cell homeostasis.

T cells play a critical role in immune responses as they specifically recognize peptide/MHC complexes with their T-cell receptors and initiate adaptive immune responses. While T cells are critical for performing appropriate effector functions and maintaining immune memory, they also can cause autoimmunity or neoplasia if misdirected or dysregulated. Thus, T cells must be tightly regulated from their development onward. Maintenance of appropriate T-cell homeostasis is essential to promote protective immunity and limit autoimmunity and neoplasia. This review will focus on the role of cell death in maintenance of T-cell homeostasis and outline novel therapeutic strategies tailored to manipulate cell death to limit T-cell survival (eg, autoimmunity and transplantation) or enhance T-cell survival (eg, vaccination and immune deficiency).

Manipulating DNA damage-response signaling for the treatment of immune-mediated diseases.

Antigen-activated lymphocytes undergo extraordinarily rapid cell division in the course of immune responses. We hypothesized that this unique aspect of lymphocyte biology leads to unusual genomic stress in recently antigen-activated lymphocytes and that targeted manipulation of DNA damage-response (DDR) signaling pathways would allow for selective therapeutic targeting of pathological T cells in disease contexts. Consistent with these hypotheses, we found that activated mouse and human T cells display a pronounced DDR in vitro and in vivo. Upon screening a variety of small-molecule compounds, we found that potentiation of p53 (via inhibition of MDM2) or impairment of cell cycle checkpoints (via inhibition of CHK1/2 or WEE1) led to the selective elimination of activated, pathological T cells in vivo. The combination of these strategies [which we termed "p53 potentiation with checkpoint abrogation" (PPCA)] displayed therapeutic benefits in preclinical disease models of hemophagocytic lymphohistiocytosis and multiple sclerosis, which are driven by foreign antigens or self-antigens, respectively. PPCA therapy targeted pathological T cells but did not compromise naive, regulatory, or quiescent memory T-cell pools, and had a modest nonimmune toxicity profile. Thus, PPCA is a therapeutic modality for selective, antigen-specific immune modulation with significant translational potential for diverse immune-mediated diseases.

The impaired and/or disabled anesthesiologist.

PURPOSE OF REVIEW: Impairment and/or disability resulting from any of a number of etiologies will afflict a significant number of anesthesiologists at some point during their career. The impaired anesthesiologist can be difficult to identify and challenging to manage. Questions will arise as to if, how, and when colleagues, family members, or friends should intercede if significant impairment is suspected.This review will examine the common sources of impairment among anesthesiologists and the professional implications of these conditions. We will discuss the obligations of an anesthesiologist and his/her colleagues when there is sufficient suspicion that he/she might be impaired. RECENT FINDINGS: Substance use disorder remains one of the commonest sources of impairment among both resident and attending anesthesiologists. Other common etiologies of impairment include various physical ailments, major psychiatric disorders, especially depression and burnout, and age related dementia. Many regulatory organizations, healthcare systems, and state licensing agencies have developed programmes and protocols with which to identify and direct into treatment those suspected of significant impairment. SUMMARY: Some degree of impairment will occur to one-third of anesthesiologists during the course of their career. It is important to understand how such impairments might impact the safe practice of anesthesiology.

Control of the Environment in the Operating Room.

There is a direct relationship between the quality of the environment of a workplace and the productivity and efficiency of the work accomplished. Components such as temperature, humidity, ventilation, drafts, lighting, and noise each contribute to the quality of the overall environment and the sense of well-being of those who work there.The modern operating room is a unique workplace with specific, and frequently conflicting, environmental requirements for each of the inhabitants. Even minor disturbances in the internal environment of the operating room can have serious ramifications on the comfort, effectiveness, and safety of each of the inhabitants. A cool, well-ventilated, and dry climate is optimal for many members of the surgical team. Any significant deviation from these objectives raises the risk of decreased efficiency and productivity and adverse surgical outcomes. A warmer, more humid, and quieter environment is necessary for the patient. If these requirements are not met, the risk of surgical morbidity and mortality is increased. An important task for the surgical team is to find the correct balance between these 2 opposed requirements. Several of the components of the operating room environment, especially room temperature and airflow patterns, are easily manipulated by the members of the surgical team. In the following discussion, we will examine these elements to better understand the clinical ramifications of adjustments and accommodations that are frequently made to meet the requirements of both the surgical staff and the patient.

The aging anesthesiologist.

PURPOSE OF REVIEW: The average age of anesthesiologists in the USA is increasing. Advancing age is accompanied by challenges and opportunities to the individual anesthesiologist and his/her colleagues. This article will discuss the science behind policies to assure continued competence among these aging physicians and safety for their patients. RECENT FINDINGS: There is growing evidence that aging anesthesiologists may be advantaged by a lifetime of experience but possibly disadvantaged under certain circumstances by lapses in current medical knowledge contributing to medical errors. Policies and procedures are emerging to assist in evaluating the continued competence of aging physicians. SUMMARY: The average age of practicing anesthesiologists in the USA is increasing. As physicians continue to practice into later years, it is critical that innovative continuing medical education programs and objective evaluations of clinical skills and competence focused upon this group continue to be developed to assure public safety.

Reversal of New-Onset Type 1 Diabetes With an Agonistic TLR4/MD-2 Monoclonal Antibody.

Type 1 diabetes (T1D) is currently an incurable disease, characterized by a silent prodromal phase followed by an acute clinical phase, reflecting progressive autoimmune destruction of insulin-producing pancreatic ß-cells. Autoreactive T cells play a major role in ß-cell destruction, but innate immune cell cytokines and costimulatory molecules critically affect T-cell functional status. We show that an agonistic monoclonal antibody to TLR4/MD-2 (TLR4-Ab) reverses new-onset diabetes in a high percentage of NOD mice. TLR4-Ab induces antigen-presenting cell (APC) tolerance in vitro and in vivo, resulting in an altered cytokine profile, decreased costimulatory molecule expression, and decreased T-cell proliferation in APC:T-cell assays. TLR4-Ab treatment increases T-regulatory cell (Treg) numbers in both the periphery and the pancreatic islet, predominantly expanding the Helios(+)Nrp-1(+)Foxp3(+) Treg subset. TLR4-Ab treatment in the absence of B cells in NOD.scid mice prevents subsequent T cell-mediated disease, further suggesting a major role for APC tolerization in disease protection. Specific stimulation of the innate immune system through TLR4/MD-2, therefore, can restore tolerance in the aberrant adaptive immune system and reverse new-onset T1D, suggesting a novel immunological approach to treatment of T1D in humans.

Eliminating encephalitogenic T cells without undermining protective immunity.

The current clinical approach for treating autoimmune diseases is to broadly blunt immune responses as a means of preventing autoimmune pathology. Among the major side effects of this strategy are depressed beneficial immunity and increased rates of infections and tumors. Using the experimental autoimmune encephalomyelitis model for human multiple sclerosis, we report a novel alternative approach for purging autoreactive T cells that spares beneficial immunity. The moderate and temporally limited use of etoposide, a topoisomerase inhibitor, to eliminate encephalitogenic T cells significantly reduces the onset and severity of experimental autoimmune encephalomyelitis, dampens cytokine production and overall pathology, while dramatically limiting the off-target effects on naive and memory adaptive immunity. Etoposide-treated mice show no or significantly ameliorated pathology with reduced antigenic spread, yet have normal T cell and T-dependent B cell responses to de novo antigenic challenges as well as unimpaired memory T cell responses to viral rechallenge. Thus, etoposide therapy can selectively ablate effector T cells and limit pathology in an animal model of autoimmunity while sparing protective immune responses. This strategy could lead to novel approaches for the treatment of autoimmune diseases with both enhanced efficacy and decreased treatment-associated morbidities.

Etoposide selectively ablates activated T cells to control the immunoregulatory disorder hemophagocytic lymphohistiocytosis.

Hemophagocytic lymphohistiocytosis (HLH) is an inborn disorder of immune regulation caused by mutations affecting perforin-dependent cytotoxicity. Defects in this pathway impair negative feedback between cytotoxic lymphocytes and APCs, leading to prolonged and pathologic activation of T cells. Etoposide, a widely used chemotherapeutic drug that inhibits topoisomerase II, is the mainstay of treatment for HLH, although its therapeutic mechanism remains unknown. We used a murine model of HLH, involving lymphocytic choriomeningitis virus infection of perforin-deficient mice, to study the activity and mechanism of etoposide for treating HLH and found that it substantially alleviated all symptoms of murine HLH and allowed prolonged survival. This therapeutic effect was relatively unique among chemotherapeutic agents tested, suggesting distinctive effects on the immune response. We found that the therapeutic mechanism of etoposide in this model system involved potent deletion of activated T cells and efficient suppression of inflammatory cytokine production. This effect was remarkably selective; etoposide did not exert a direct anti-inflammatory effect on macrophages or dendritic cells, and it did not cause deletion of quiescent naive or memory T cells. Finally, etoposide's immunomodulatory effects were similar in wild-type and perforin-deficient animals. Thus, etoposide treats HLH by selectively eliminating pathologic, activated T cells and may have usefulness as a novel immune modulator in a broad array of immunopathologic disorders.

United States anesthesiologists over 50: retirement decision making and workforce implications.

BACKGROUND: Anesthesiology is among the medical specialties expected to have physician shortage. With little known about older anesthesiologists' work effort and retirement decision making, the American Society of Anesthesiologists participated in a 2006 national survey of physicians aged 50-79 yr. METHODS: Samples of anesthesiologists and other specialists completed a survey of work activities, professional satisfaction, self-defined health and financial status, retirement plans and perspectives, and demographics. A complex survey design enabled adjustments for sampling and response-rate biases so that respondents' characteristics resembled those in the American Medical Association Physician Masterfile. Retirement decision making was modeled with multivariable ordinal logistic regression. Life-table analysis provided a forecast of likely clinical workforce trends over an ensuing 30 yr. RESULTS: Anesthesiologists (N = 3,222; response rate = 37%) reported a mean work week of 49.4 h and a mean retirement age of 62.7 yr, both values similar to those of other older physicians. Work week decreased with age, and part-time work increased. Women worked a shorter work week (mean, 47.9 vs. 49.7 h, P = 0.024), partly due to greater part-time work (20.2 vs. 13.1%, P value less than 0.001). Relative importance of factors reported among those leaving patient care differed by age cohort, subspecialty, and work status. Poor health was cited by 64% of anesthesiologists retiring in their 50s as compared with 43% of those retiring later (P = 0.039). CONCLUSIONS: This survey lends support for greater attention to potentially modifiable factors, such as workplace wellness and professional satisfaction, to prevent premature retirement. The growing trend in part-time work deserves further study.