A Dexamethasone Prodrug Reduces the Renal Macrophage Response and Provides Enhanced Resolution of Established Murine Lupus Nephritis

We evaluated the ability of a macromolecular prodrug of dexamethasone (P-Dex) to treat lupus nephritis in (NZB × NZW)F1 mice. We also explored the mechanism underlying the anti-inflammatory effects of this prodrug. P-Dex eliminated albuminuria in most (NZB × NZW)F1 mice. Furthermore, P-Dex reduced the incidence of severe nephritis and extended lifespan in these mice. P-Dex treatment also prevented the development of lupus-associated hypertension and vasculitis. Although P-Dex did not reduce serum levels of anti-dsDNA antibodies or glomerular immune complexes, P-Dex reduced macrophage recruitment to the kidney and attenuated tubulointerstitial injury. In contrast to what was observed with free dexamethasone, P-Dex did not induce any deterioration of bone quality. However, P-Dex did lead to reduced peripheral white blood cell counts and adrenal gland atrophy. These results suggest that P-Dex is more effective and less toxic than free dexamethasone for the treatment of lupus nephritis in (NZB × NZW)F1 mice. Furthermore, the data suggest that P-Dex may treat nephritis by attenuating the renal inflammatory response to immune complexes, leading to decreased immune cell infiltration and diminished renal inflammation and injury.     

Testosterone Affects Neural Gene Expression Differently in Male and Female Juncos: A Role for Hormones in Mediating Sexual Dimorphism and Conflict

Despite sharing much of their genomes, males and females are often highly dimorphic, reflecting at least in part the resolution of sexual conflict in response to sexually antagonistic selection. Sexual dimorphism arises owing to sex differences in gene expression, and steroid hormones are often invoked as a proximate cause of sexual dimorphism. Experimental elevation of androgens can modify behavior, physiology, and gene expression, but knowledge of the role of hormones remains incomplete, including how the sexes differ in gene expression in response to hormones. We addressed these questions in a bird species with a long history of behavioral endocrinological and ecological study, the dark-eyed junco (Junco hyemalis), using a custom microarray. Focusing on two brain regions involved in sexually dimorphic behavior and regulation of hormone secretion, we identified 651 genes that differed in expression by sex in medial amygdala and 611 in hypothalamus. Additionally, we treated individuals of each sex with testosterone implants and identified many genes that may be related to previously identified phenotypic effects of testosterone treatment. Some of these genes relate to previously identified effects of testosterone-treatment and suggest that the multiple effects of testosterone may be mediated by modifying the expression of a small number of genes. Notably, testosterone-treatment tended to alter expression of different genes in each sex: only 4 of the 527 genes identified as significant in one sex or the other were significantly differentially expressed in both sexes. Hormonally regulated gene expression is a key mechanism underlying sexual dimorphism, and our study identifies specific genes that may mediate some of these processes.     

Homology of ciliary bands in Spiralian Trochophores

A number of hypotheses have been presented regarding the originsof the metazoans and, more specifically, the Bilateria. Usingvarious phylogenetic analyses, characteristics have been mappedon phylogenetic trees to infer ancestral body plans and lifehistory strategies of those ancestors. Many arguments on theevolution of the Bilateria are based on the presumed homologyof certain characteristics of extant larva and adults, includingvarious ciliated bands involved in feeding and locomotion. Thisarticle considers a recent study indicating that the second,downstream-collecting, ciliated band in the veliger larva ofthe gastropod mollusc, Crepidula fornicata, is actually derivedfrom secondary trochoblasts (derived from second quartet micromeres),that normally form part of the prototrochal band found in otherspiralian phyla (Hejnol et al. 2007). Despite previous arguments,these new findings suggest that the second ciliated band inthe veliger larva is not homologous to the metatroch found inthe trochophore larva of some other spiralians, such as theannelid, Polygordius lacteus. In the latter case, the metatrochwas reported to be formed by a different set of lineage precursors(derived from third quartet micromeres) (Woltereck 1904). Thesefindings have important implications for the interpretationof various hypotheses related to the evolution of metazoan phyla.     

Multiple triclosan targets in Trypanosoma brucei

Trypanosoma brucei genes encoding putative fatty acid synthesis enzymes are homologous to those encoding type II enzymes found in bacteria and organelles such as chloroplasts and mitochondria. It was therefore not surprising that triclosan, an inhibitor of type II enoyl-acyl carrier protein (enoyl-ACP) reductase, killed both procyclic forms and bloodstream forms of T. brucei in culture with 50% effective concentrations (EC(50)s) of 10 and 13 microM, respectively. Triclosan also inhibited cell-free fatty acid synthesis, though much higher concentrations were required (EC(50)s of 100 to 200 microM). Unexpectedly, 100 microM triclosan did not affect the elongation of [(3)H]laurate (C(12:0)) to myristate (C(14:0)) in cultured bloodstream form parasites, suggesting that triclosan killing of trypanosomes may not be through specific inhibition of enoyl-ACP reductase but through some other mechanism. Interestingly, 100 microM triclosan did reduce the level of incorporation of [(3)H]myristate into glycosyl phosphatidylinositol species (GPIs). Furthermore, we found that triclosan inhibited fatty acid remodeling in a cell-free assay in the same concentration range required for killing T. brucei in culture. In addition, we found that a similar concentration of triclosan also inhibited the myristate exchange pathway, which resides in a distinct subcellular compartment. However, GPI myristoylation and myristate exchange are specific to the bloodstream form parasite, yet triclosan kills both the bloodstream and procyclic forms. Therefore, triclosan killing may be due to a nonspecific perturbation of subcellular membrane structure leading to dysfunction in sensitive membrane-resident biochemical pathways.     

Role of programmed cell death in patterning the Drosophila antennal arista

Programmed cell death is a critical process for the patterning and sculpting of organs during development. The Drosophila arista, a feather-like structure at the tip of the antenna, is composed of a central core and several lateral branches. A homozygous viable mutation in the thread gene, which encodes an inhibitor of apoptosis protein, produces a branchless arista. We have found that mutations in the proapoptotic gene hid lead to numerous extra branches, suggesting that the level of cell death determines the number of branches in the arista. Consistent with this idea, we have found that thread mutants show excessive cell death restricted to the antennal imaginal disc during the middle third instar larval stage. These findings point to a narrow window of development in which regulation of programmed cell death is essential to the proper formation of the arista.     

TRAIL-R2 (DR5) mediates apoptosis of synovial fibroblasts in rheumatoid arthritis

TRAIL has been proposed as an anti-inflammatory cytokine in animal models of rheumatoid arthritis (RA). Using two agonistic mAbs specific for TRAIL-R1 (DR4) and TRAIL-R2 (DR5), we examined the expression and function of these death receptors in RA synovial fibroblast cells. The synovial tissues and primary synovial fibroblast cells isolated from patients with RA, but not those isolated from patients with osteoarthritis, selectively expressed high levels of cell surface DR5 and were highly susceptible to anti-DR5 Ab (TRA-8)-mediated apoptosis. In contrast, RA synoviocytes did not show increased expression of TRAIL-R1 (DR4), nor was there any difference in expression of Fas between RA and osteoarthritis synovial cells. In vitro TRA-8 induced apoptosis of RA synovial cells and inhibited production of matrix metalloproteinases induced by pro-inflammatory cytokines. In vivo TRA-8 effectively inhibited hypercellularity of a SV40-transformed RA synovial cell line and completely prevented bone erosion and cartilage destruction induced by these cells. These results indicate that increased DR5 expression and susceptibility to DR5-mediated apoptosis are characteristic of the proliferating synovial cells in RA. As highly proliferative transformed-appearing RA synovial cells play a crucial role in bone erosion and cartilage destruction in RA, the specific targeting of DR5 on RA synovial cells with an agonistic anti-DR5 Ab may be a potential therapy for RA.     

Novel Translation Products from Simian Immunodeficiency Virus SIVmac239 Env-Encoding mRNA Contain both Rev and Cryptic T-Cell Epitopes

Understanding the correlates of immune protection against human immunodeficiency virus and simian immunodeficiency virus (SIV) will require defining the entire cellular immune response against the viruses. Here, we define two novel translation products from the SIV env mRNA that are targeted by the T-cell response in SIV-infected rhesus macaques. The shorter product is a subset of the larger product, which contains both the first exon of the Rev protein and a translated portion of the rev intron. Our data suggest that the translation of viral alternate reading frames may be an important source of T-cell epitopes, including epitopes normally derived from functional proteins.The pathway from viral infection to the cellular immune response is not well understood. Despite the importance of T-cell responses in control of AIDS virus replication (1, 3, 8, 22), the sources of the peptides recognized by virus-specific T cells are still being discovered. AIDS virus-specific CD8⁺ T lymphocytes (CD8-TL) recognize complexes of major histocompatibility complex (MHC) class I and virus-derived epitopes presented on the surface of infected cells. These epitopes can be derived from exogenous viral proteins in the infecting virion (19, 20) or from de novo synthesis of viral proteins (9, 21). Additional sources of epitopes are also being explored (4, 6).CD8-TL can also recognize epitopes derived from translation of viral alternate reading frames (ARFs). Though CD8-TL specific for ARF-derived epitopes have been detected in human immunodeficiency virus (HIV) (2), they remain a largely unexplored source of epitopes that might elicit potent antiviral cellular immune responses. We recently showed that SIVmac239-infected rhesus macaques that spontaneously controlled viral replication, termed elite controllers, made immunodominant CD8-TL responses against an epitope (RHLAFKCLW, or cRW9) derived from an ARF of the env gene (15). This response selected for viral escape in vivo and suppressed viral replication in an in vitro assay. These findings imply that CD8-TL specific for ARF-derived epitopes might be an important component of the total AIDS virus-specific cellular immune response.Here, we show that the cRW9 epitope is translated as part of two distinct products that differ in size due to start codon usage. The larger and more frequent product contains both the first 23 amino acids of the Rev protein (exon 1) and 50 amino acids translated from the rev intron. The smaller is produced by translation initiation at a start codon within the rev intron and is a subset of the larger product. Finally, we show that these products are degraded after translation from the mature Env-encoding mRNA.     

Differential modulation of 3T3-L1 adipogenesis mediated by 11beta-hydroxysteroid dehydrogenase-1 levels

The localized activation of circulating glucocorticoids in vivo by the enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) plays a critical role in the development of the metabolic syndrome. However, the precise contribution of 11beta-HSD1 in the initiation of adipogenesis by inactive glucocorticoids is not fully understood. 3T3-L1 fibroblasts can be terminally differentiated to mature adipocytes in a glucocorticoid-dependent manner. Both inactive rodent dehydrocorticosterone and human cortisone were able to substitute for the synthetic glucocorticoid dexamethasone in 3T3-L1 adipogenesis, suggesting a potential role for 11beta-HSD1 in these effects. Differentiation of 3T3-L1 cells caused a strong increase in 11beta-HSD1 protein levels, which occurred late in the differentiation protocol. Reduction of 11beta-HSD1 activity in 3T3-L1 fibroblasts, achieved by pharmacological inhibition or adenovirally mediated delivery of short hairpin RNA constructs, specifically blocked the ability of inactive glucocorticoids to drive 3T3-L1 differentiation. However, even modest increases in exogenous 11beta-HSD1 expression in 3T3-L1 fibroblasts, to levels comparable with endogenous 11beta-HSD1 in differentiated 3T3-L1 adipocytes, were sufficient to block adipogenesis. Luciferase reporter assays indicated that overexpressed 11beta-HSD1 was catalyzing the inactivating dehydrogenase reaction, because the ability of both active and inactive glucocorticoids to activate the glucocorticoid receptor were largely suppressed. These results suggest that the temporal regulation of 11beta-HSD1 expression is tightly controlled in 3T3-L1 cells, so as to mediate the initiation of differentiation by inactive glucocorticoids and also to prevent the inhibitory activity of prematurely expressed 11beta-HSD1 during adipogenesis.