Nonsteroidal anti-inflammatory drug sensitizes Mycobacterium tuberculosis to endogenous and exogenous antimicrobials.

Existing drugs are slow to eradicate Mycobacterium tuberculosis (Mtb) in patients and have failed to control tuberculosis globally. One reason may be that host conditions impair Mtb's replication, reducing its sensitivity to most antiinfectives. We devised a high-throughput screen for compounds that kill Mtb when its replication has been halted by reactive nitrogen intermediates (RNIs), acid, hypoxia, and a fatty acid carbon source. At concentrations routinely achieved in human blood, oxyphenbutazone (OPB), an inexpensive anti-inflammatory drug, was selectively mycobactericidal to nonreplicating (NR) Mtb. Its cidal activity depended on mild acid and was augmented by RNIs and fatty acid. Acid and RNIs fostered OPB's 4-hydroxylation. The resultant 4-butyl-4-hydroxy-1-(4-hydroxyphenyl)-2-phenylpyrazolidine-3,5-dione (4-OH-OPB) killed both replicating and NR Mtb, including Mtb resistant to standard drugs. 4-OH-OPB depleted flavins and formed covalent adducts with N-acetyl-cysteine and mycothiol. 4-OH-OPB killed Mtb synergistically with oxidants and several antituberculosis drugs. Thus, conditions that block Mtb's replication modify OPB and enhance its cidal action. Modified OPB kills both replicating and NR Mtb and sensitizes both to host-derived and medicinal antimycobacterial agents.

Endocarditis caused by Rhodotorula infection.

Rhodotorula is an emerging opportunistic fungal pathogen that is rarely reported to cause endocarditis. We describe a case involving a patient who developed endocarditis due to Rhodotorula mucilaginosa and Staphylococcus epidermidis, proven by culture and histopathology. The case illustrates the unique diagnostic and therapeutic challenges relevant to Rhodotorula spp.

Novel Cephalosporins Selectively Active on Nonreplicating Mycobacterium tuberculosis.

We report two series of novel cephalosporins that are bactericidal to Mycobacterium tuberculosis alone of the pathogens tested, which only kill M. tuberculosis when its replication is halted by conditions resembling those believed to pertain in the host, and whose bactericidal activity is not dependent upon or enhanced by clavulanate, a ß-lactamase inhibitor. The two classes of cephalosporins bear an ester or alternatively an oxadiazole isostere at C-2 of the cephalosporin ring system, a position that is almost exclusively a carboxylic acid in clinically used agents in the class. Representatives of the series kill M. tuberculosis within macrophages without toxicity to the macrophages or other mammalian cells.

The apoptotic-cell receptor CR3, but not alphavbeta5, is a regulator of human dendritic-cell immunostimulatory function.

Dendritic cells (DCs) that capture apoptotic cells (ACs) in the steady state mediate peripheral tolerance to self-antigens. ACs are recognized by an array of receptors on DCs, the redundancy of which is not completely defined. We made use of an AC surrogate system to address the individual roles of the alphavbeta5 and complement receptors (CRs) in the phagocytosis and induction of immunity. CR3 and CR4, while substantially less efficient than alphavbeta5 in internalizing ACs, initiate signals that render DCs tolerogenic. Responding T cells show impaired proliferation and IFNgamma production and subsequently die by apoptosis. While tolerogenic DCs are not induced via alphavbeta5, coligation of CR3 and alphavbeta5 maintains the DC's tolerogenic profile. This immunomodulatory role, however, is countered by a significant inflammatory stimulus such as bacterial infection. Overall, our data suggest that under steady-state conditions, signaling via CRs predominates to render DCs tolerogenic.

A clinical grade cocktail of cytokines and PGE2 results in uniform maturation of human monocyte-derived dendritic cells: implications for immunotherapy.

Dendritic cells (DCs) can induce tumor- or pathogen-specific T cell responses in humans. We comprehensively compared the clinically available DC maturation stimuli for their ability to promote uniformly mature DCs that elicit higher levels of T cell responses. We compared the standard maturation stimulus, autologous monocyte-conditioned medium (MCM), with a synthetic double stranded RNA (poly I:C), soluble CD40 ligand trimer, and a defined cocktail of cytokines (TNF-alpha, IL-1 beta, IL-6) and PGE(2) to promote mature phenotype and function in human monocyte-derived DCs. The cocktail was the most efficient despite the lack of induction of IL-12p70. While these results support the use of the MCM-mimic cocktail in clinical DC immunotherapy trials, the roles of it's individual constituents remain to be completely defined.

Cutting edge: identification of the targets of clonal deletion in an unmanipulated thymus.

Autoreactive thymocytes can be eliminated by clonal deletion during their development in the thymus. The precise developmental stage(s) at which clonal deletion occurs in a normal thymus has been difficult to assess, in large part because of the absence of a specific marker for TCR-mediated apoptosis. In this report, we reveal that Nur77 expression can be used as a specific marker of clonal deletion in an unmanipulated thymus and directly identify TCRintCD4+CD8+ and semimature CD4+CD8- thymocytes as the principal targets of deletion. These data indicate that clonal deletion normally occurs at a relatively late stage of development, as cells mature from CD4+CD8+ thymocytes to single-positive T cells.