Regulation of interleukin-2 (IL-2) gene expression by the p50 and p65 subunits of the DNA binding protein NF-kappa B was studied in nontransformed CD4+ T lymphocyte clones. A homodimeric complex of the NF-kappa B p50 subunit was found in resting T cells. The amount of p50-p50 complex decreased after full antigenic stimulation, whereas the amount of the NF-kappa B p50-p65 heterodimer was increased. Increased expression of the IL-2 gene and activity of the IL-2 kappa B DNA binding site correlated with a decrease in the p50-p50 complex. Overexpression of p50 repressed IL-2 promoter expression. The switch from p50-p50 to p50-p65 complexes depended on a protein that caused sequestration of the p50-p50 complex in the nucleus.

Anergy is a mechanism of T lymphocyte tolerance induced by antigen receptor stimulation in the absence of co-stimulation. Anergic T cells were shown to have a defect in antigen-induced transcription of the interleukin-2 gene. Analysis of the promoter indicated that the transcription factor AP-1 and its corresponding cis element were specifically down-regulated. Exposure of anergic T cells to interleukin-2 restored both antigen responsiveness and activity of the AP-1 element.

Increasing evidence suggests that HIV-1-specific cytotoxic T lymphocytes (CTLs) are a key host immune response to HIV-1 infection. Generation of CTL responses for prevention or therapy of HIV-1 infection has several intrinsic technical barriers such as antigen expression and presentation, the varying HLA restrictions between different individuals, and the potential for viral escape by sequence variation or surface molecule alteration on infected cells. A strategy to circumvent these limitations is the construction of a chimeric T cell receptor containing human CD4 or HIV-1-specific Ig sequences linked to the signaling domain of the T cell receptor chain (universal T cell receptor). CD8؉ CTLs transduced with this universal receptor can then bind and lyse infected cells that express surface HIV-1 gp120. We evaluated the ability of universal-receptorbearing CD8؉ cells from a seronegative donor to lyse acutely infected cells and inhibit HIV-1 replication in vitro. The kinetics of lysis and efficiency of inhibition were comparable to that of naturally occurring HIV-1-specific CTL clones isolated from infected individuals. Further study will be required to determine the utility of these cells as a therapeutic strategy in vivo.

Tuberculosis (TB) is the most deadly infectious disease in existence, and the only available vaccine, Bacillus Calmette-Guérin (BCG), is almost a century old and poorly protective. The immunological complexity of TB, coupled with rising resistance to antimicrobial therapies, necessitates a pipeline of diverse novel vaccines. Here, we show that Bacillus subtilis spores can be coated with a fusion protein 1 (“FP1”) consisting of Mycobacterium tuberculosis (Mtb) antigens Ag85B, ACR, and HBHA. The resultant vaccine, Spore-FP1, was tested in a murine low-dose Mtb aerosol challenge model. Mice were primed with subcutaneous BCG, followed by mucosal booster immunizations with Spore-FP1. We show that Spore-FP1 enhanced pulmonary control of Mtb, as evidenced by reduced bacterial burdens in the lungs. This was associated with elevated antigen-specific IgG and IgA titers in the serum and lung mucosal surface, respectively. Spore-FP1 immunization generated superior antigen-specific memory T-cell proliferation in both CD4+ and CD8+ compartments, alongside bolstered Th1-, Th17-, and Treg-type cytokine production, compared to BCG immunization alone. CD69+CD103+ tissue resident memory T-cells (Trm) were found within the lung parenchyma after mucosal immunization with Spore-FP1, confirming the advantages of mucosal delivery. Our data show that Spore-FP1 is a promising new TB vaccine that can successfully augment protection and immunogenicity in BCG-primed animals.

We have developed an immunotherapeutic approach with potential application in the treatment of viral and malignant disease. We show that primary CD8+ T cells isolated from peripheral blood can be genetically modified by retroviral transduction to express high levels of universal (major histocompatibility complex-unrestricted) chimeric T- cell receptors specific for human immunodeficiency virus (HIV) antigens. Two classes of HIV-specific URs in which the antigen-binding domain is comprised of either CD4 or a single-chain antibody are capable of activating a number of T-cell effector functions in response to target cells, including cytolysis, in a highly sensitive and specific manner. Importantly, we have addressed a number of issues which, although particularly relevant to the clinical application of this approach in the treatment of HIV infection, may also impact on the potential of UR immunotherapy for other disease targets. The UR immunotherapeutic system is particularly suited for evaluation in the clinical setting.

Obesity is a prominent metabolic disease that predisposes individuals to multiple comorbidities, including type 2 diabetes mellitus, cardiovascular diseases, and cancer. Elevated circulating levels of fatty acids contribute to the development of obesity, in part, by targeting the hypothalamus. Palmitate, the most abundant circulating saturated fatty acid, has been demonstrated to dysregulate NAMPT and circadian clock proteins, as well as induce neuroinflammation. These effects ultimately result in hypothalamic dysregulation of feeding behavior and energy homeostasis. NAMPT is the rate-limiting enzyme of the NAD+ salvage pathway and its expression is under the control of the circadian clock. NAD+ produced from NAMPT can modulate the circadian clock, demonstrating bidirectional interactions between circadian and metabolic pathways. Using NPY/AgRP-expressing mHypoE-46 neurons as well as the novel mHypoA-BMAL1-WT/F and mHypoA-BMAL1-KO/F cell lines, we studied whether there were any interactions between NAMPT and the core circadian clock protein BMAL1 in the palmitate-mediated induction of neuroinflammation. We report that palmitate altered Nampt, Bmal1, Per2 and the inflammatory genes Nf-κb, IκBα, Il-6, and Tlr4. Contrary to studies performed with peripheral tissues, the palmitate-mediated induction in Nampt was independent of BMAL1, and basal Nampt levels did not appear to exhibit rhythmic expression. Palmitate-induced downregulation of Bmal1 and Per2 was independent of NAMPT. However, NAMPT and BMAL1 were both involved in the regulation of Nf-κb, IκBα, Il-6, and Tlr4, as NAMPT inhibition resulted in the repression of basal Nf-κb and IκBα and normalized palmitate-mediated increases in Il-6, and Tlr4. On the other hand, BMAL1 deletion repressed basal Nf-κb, but increased basal Il-6. We conclude that NAMPT and BMAL1 do not interact at the transcriptional level in hypothalamic neurons, but are independently involved in the expression of inflammatory genes.

Vaccination is considered the most effective strategy for controlling tuberculosis (TB). The existing vaccine, the Bacille Calmette-Guérin (BCG), although partially protective, has a number of limitations. Therefore, there is a need for developing new TB vaccines and several strategies are currently exploited including the use of viral and bacterial delivery vectors. We have previously shown that Lactobacillus plantarum ( Lp ) producing Ag85B and ESAT-6 antigens fused to a dendritic cell-targeting peptide (referred to as Lp _DC) induced specific immune responses in mice. Here, we analyzed the ability of two Lp -based vaccines, Lp _DC and Lp _HBD (in which the DC-binding peptide was replaced by an HBD-domain directing the antigen to non-phagocytic cells) to activate antigen-presenting cells, induce specific immunity and protect mice from Mycobacterium tuberculosis infection. We tested two strategies: (i) Lp as BCG boosting vaccine (a heterologous regimen comprising parenteral BCG immunization followed by intranasal Lp boost), and (ii) Lp as primary vaccine (a homologous regimen including subcutaneous priming followed by intranasal boost). The results showed that both Lp constructs applied as a BCG boost induced specific cellular immunity, manifested in T cell proliferation, antigen-specific IFN-γ responses and multifunctional T cells phenotypes. More importantly, intranasal boost with Lp _DC or Lp _HBD enhanced protection offered by BCG, as shown by reduced M. tuberculosis counts in lungs. These findings suggest that Lp constructs could be developed as a potential mucosal vaccine platform against mycobacterial infections.

The best way to debunk a scientific dogma is to throw irrefutable evidence at it. This is especially true if the dogma in question has been nurtured over many decades, as is the case with the apparent redundancy of antibodies (Abs) against intracellular pathogens. Although not fully compelling yet, that ‘hard core’ evidence is nevertheless now slowly beginning to emerge. This is true for several clinically relevant infections but none more so than Mycobacterium tuberculosis , the archetype intracellular pathogen that poses a great health challenge to the mankind. Here, prompted by a spate of recent high-profile reports on the effects of Abs in various experimental models of tuberculosis, we step back and take a critical look at the progress that has been made in the last 5 years and highlight some of the strengths and shortcomings of the presented evidence. We conclude that the tide of the opinion has begun to turn in favour of Abs but we also caution against overinterpreting the currently available limited evidence. For, until definitive evidence that can withstand even the most rigorous of experimental tests is produced, the dogma may yet survive. Or indeed, we may find that the truth is hidden somewhere in between the dogma and the unfulfilled scientific prophecy.