The T cells specific for the carboxyl-terminal determinants of self (rat) heat-shock protein 65 escape tolerance induction and are involved in regulation of autoimmune arthritis

Immunization of Lewis rats with heat-killed Mycobacterium tuberculosis H37Ra leads to development of polyarthritis (adjuvant-induced arthritis; AA) that shares several features with human rheumatoid arthritis (RA). Immune response to the 65-kDa mycobacterial heat-shock protein (Bhsp65) is believed to be involved in induction of AA as well as in experimental modulation of this disease. However, the understanding of several critical aspects of the pathogenesis of AA in the Lewis rat has severely been hampered by the lack of information both regarding the level as well as epitope specificity of tolerance to the mammalian self (rat) homologue of Bhsp65, 65-kDa rat heat-shock protein (Rhsp65), and about the functional attributes of the T cell repertoire specific for this self protein. In this study, we established that tolerance to Rhsp65 in the Lewis rat is incomplete, and that the residual T cells primed upon challenge with this self hsp65 are disease regulating in nature. We also have defined the T cell epitopes in the C-terminal region within Rhsp65 that contribute predominantly to the immune reactivity as well as the AA-protective effect of this self protein. Furthermore, the T cells primed by peptides comprising these C-terminal determinants can be efficiently restimulated by the naturally generated epitopes from endogenous Rhsp65, suggesting that self hsp65 might also be involved in natural remission from acute AA. These novel first experimental insights into the self hsp65-directed regulatory T cell repertoire in AA would help develop better immunotherapeutic approaches for autoimmune arthritis.     

Mouse lysozyme-M knockout mice reveal how the self-determinant hierarchy shapes the T cell repertoire against this circulating self antigen in wild-type mice

We have studied T cell tolerance to defined determinants within ML-M using wild-type (WT; ML-M(+/+)) and LysMcre (ML-M(-/-)) C3H (H-2(k)) mice to determine the relative contribution of ML-M-derived epitopes vs those from other self Ags in selection of the ML-M-specific T cell repertoire. ML-M was totally nonimmunogenic in WT mice, but was rendered immunogenic in LysMcre mice. Most of the response to ML-M in LysMcre mice was directed to the immunodominant determinant region 105-119. This determinant is spontaneously displayed (without adding exogenous ML-M) by macrophages of WT, but not LysMcre, mice and is stimulatory for peptide 105-119 (p105-119)-primed T cells. Moreover, neonatal tolerization of LysMcre mice with p105-119 or ML-M abrogated the T cell response to subsequent challenge with ML-M or p105-119. Furthermore, p95-109 and p110-125 of ML-M were immunogenic in LysMcre mice, but not in WT mice, thereby representing subdominant, tolerance-inducing epitopes of ML-M. As expected, the T cell repertoire to cryptic ML determinants in WT mice was also intact in LysMcre mice. Furthermore, the pattern of response to the related homologue of ML-M, hen eggwhite lysozyme, was similar in these two groups of mice. Thus, several codominant T cell determinants within ML-M contribute significantly to tolerance induction, and the anti-cryptic T cell repertoire to ML-M was positively selected on non-ML-M self ligands. These results reveal that the induction of self tolerance to a multideterminant protein follows the quantitative hierarchy of self-determinant expression and are of relevance in understanding the pathogenesis of autoimmunity.     

Cellular immune responses in seronegative sexual contacts of acute hepatitis C patients

Acute hepatitis C virus (HCV) is typically defined as new viremia and antibody seroconversion. Rates and immunologic correlates of hepatitis C clearance have therefore been based on clearance of viremia only in individuals who initially had an antibody response. We sought to characterize the immunological correlates of clearance in patients with acute hepatitis C and their sexual contacts. We prospectively determined CD4(+) and CD8(+) cytotoxic T-lymphocyte responses in index patients with acute HCV and their sexual contacts who developed acute infection, either with or without spontaneous clearance, as well as those contacts who never developed viremia. Responses were measured using proliferation and ELISpot assays for CD4(+) and CD8(+) responses. We demonstrate in this prospective study that cellular immune responses can develop in exposed but persistently aviremic and antibody-negative individuals as well as those individuals with spontaneous clearance of acute HCV. These findings lend further credence to the importance of cellular immune responses in recovery from HCV and suggest that low exposure to HCV may lead to development of HCV-specific immune responses without ongoing HCV replication. This finding has important implications for HCV vaccine and therapeutic development.     

Bioreactor culture of oil palm (Elaeis guineensis) and effects of nitrogen source, inoculum size, and conditioned medium on biomass production

We report the successful culture of oil palm (Elaeis guineensis Jacq.) suspension cells in a bioreactor. In vitro propagation of this perennial monocotyledonous tree is an important part of the oil palm industry's approach to clonal propagation of high-yielding accessions. During culture of oil palm cells in a batch bioreactor, nutrients and extracellular metabolites were monitored, and kinetic parameters and nutrient-to-biomass conversion yields were calculated. The biomass increased approximately 3.5-fold per month, consistent with values reported for shake flask cultures. Although the carbon source was completely depleted by the end of the run, nitrogen sources remained in large excess and the sugar-to-biomass conversion yield remained low. Linear growth indicated that the cells were limited. The results obtained from the bioreactor runs indicated that we should be able to improve biomass production by carrying out optimization studies. Therefore, we initiated multi-factorial analyses using response surface experimental designs to investigate the effects of different nitrogen sources, as well as inoculum size and conditioned medium, on biomass production in flask cultures. Whereas glutamine does not have a significant effect on biomass production, ammonia has a positive effect up to an optimum concentration. Both inoculum density and conditioned medium have positive, synergistic effects on biomass production.     

Molecular mapping of<Emphasis Type="Italic"> Fusarium oxysporum</Emphasis> f. sp.<Emphasis Type="Italic"> ciceris</Emphasis> race 3 resistance gene in chickpea

Sequence-tagged microsatellite site (STMS) and sequence-tagged site (STS) markers linked closely to Fusarium oxysporum f. sp. ciceris race 3 resistance gene in chickpea were identified, and linkage between three wilt resistance genes was elucidated. The resistance to race 3 in chickpea germplasm accession WR-315 was inherited as a single gene, designated foc-3, in 100 F₇ recombinant inbred lines derived from the cross of WR-315 (resistant) × C-104 (susceptible). The foc-3 gene was mapped 0.6 cM from STMS markers TA96 and TA27 and STS marker CS27A. Another STMS marker, TA194, at 14.3 cM, flanked the gene on the other side. Linkage between foc-3 and two other chickpea wilt resistance genes, foc-1 (syn. h ₁ ) and foc-4, was established. foc-3 was mapped 9.8 cM from foc-1 and 8.7 cM from foc-4, whereas foc-1 and foc-4 are closely linked at 1.1 cM. The identification of closely linked markers to resistance genes will facilitate marker-assisted selection for introgression of the race 3 resistance gene to susceptible chickpea lines.Communicated by H.C. Becker     

Compatibility of alkaline xylanases from an alkaliphilic<Emphasis Type="Italic"> Bacillus</Emphasis> NCL (87-6-10) with commercial detergents and proteases

Alkaline xylanases from alkaliphilic Bacillus strains NCL (87-6-10) and Sam III were compared with the commercial xylanases Pulpzyme HC and Biopulp for their compatibility with detergents and proteases for laundry applications. Among the four xylanases evaluated, the enzyme from the alkaliphilic Bacillus strain NCL (87-6-10) was the most compatible. The enzyme retained its full activity (40 °C for 1 h) in the presence of detergents, whereas Pulpzyme HC and Sam III showed only 30% and 50% of their initial activity, respectively. Biopulp, though stable to detergents, had only marginal activity (5%)at pH 10. However, all four enzymes retained significant activity (80%) for 60 min in the presence of the proteases Alcalase and Conidiobolus protease. Supplementation of the enzyme enhanced the cleaning ability of the detergents.     

Synthesis and antitumour activity of pyrene-linked pyrrolo [2,1-c]benzodiazepine hybrids

Pyrene-linked pyrrolobenzodiazepine hybrids have been synthesized that exhibit potential anticancer activity in a number of human tumour cell lines. These hybrids also exhibit much enhanced DNA-binding ability in comparison to the parent pyrrolobenzodiazepine ring system (DC-81).     

Synthesis and DNA binding affinity of novel A-C8/C-C2-exo unsaturated alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers

The synthesis of novel A-C8/C-C2-exo unsaturated alkoxyamido-linked pyrrolo[2,1-c][1,4]benzodiazepine dimers is reported and these dimers show significant DNA binding affinity and they also exhibit moderate anticancer activity.     

Activity of some hepatic enzymes in schistosomiasis and concomitant alteration of arylsulfatase B

The levels of arylsulfatases A and B, alpha-amylase, aspartate transcarbamylase, and gamma-glutamyl transpeptidase were investigated during the infection of mice with schistosoma mansoni. This infection caused a significant (p < 0.001) increase in the activity of hepatic arylsulfatase B (ASB), aspartate transcarbamylases and gamma-glutamyl transpeptidase. A non-significant difference occurred for alpha-amylase (p < 0.3) and arylsulfatase A (p > 0.5) when compared to the control. The specific activity of hepatic ASB was progressively increased with the progression of the Schistosoma-infection. Moreover, the kinetic studies of hepatic ASB in Schistosoma-infection showed that a slight decrease in the value of K(m) and about a 40% increase in V(max) when compared to the control. In addition, the pH optimum of hepatic ASB was altered from 6 to 7 as a result of schistosomiasis. These observations suggest that there are schistosomiasis-associated changes of the catalytic and kinetic properties of hepatic ASB.     

Solid-phase synthesis of pyrrolo[2,1-c][1,4]benzodiazepine-5,11-diones

The solid-phase synthesis of biologically important pyrrolo[2,1-c][1,4]benzodiazepine-5,11-diones using Wang resin through amide formation and reductive cyclization procedures is described. Further, N10-substituents have been introduced in the final products and these have been cleaved from the solid support in good yields.