Paclitaxel enhances macrophage IL-12 production in tumor-bearing hosts through nitric oxide.

Tumor-induced macrophages (Mphis) mediate immunosuppression, in part, through increased production of factors that suppress T cell responsiveness and underproduction of positive regulatory cytokines. Pretreatment of tumor-bearing host (TBH) Mphis with the anticancer agent paclitaxel (Taxol) partially reverses tumor-induced Mphi suppressor activity, suggesting that paclitaxel may restore TBH Mphi production of proimmune factors. Because paclitaxel demonstrates LPS-mimetic capabilities and increased production of the LPS-induced immunostimulatory cytokine IL-12 could account for enhanced T cell responsiveness, we investigated whether paclitaxel induces Mphi IL-12 production. Tumor growth significantly down-regulated Mphi IL-12 p70 production through selective dysregulation of IL-12 p40 expression. LPS stimulation failed to overcome tumor-induced dysregulation of p40 expression. In contrast, paclitaxel significantly enhanced both normal host and TBH Mphi IL-12 p70 production in vitro, although TBH Mphi IL-12 production was lower than that of similarly treated normal host Mphis. Paclitaxel enhanced p40 expression in a dose-dependent manner. Through reconstituted Mphi IL-12 expression, paclitaxel pretreatment relieved tumor-induced Mphi suppression of T cell alloreactivity. Blocking Mphi NO suppressed paclitaxel's ability to induce IL-12 production. This suggests that paclitaxel-induced activities may involve a NO-mediated autocrine induction pathway. Collectively, these data demonstrate that paclitaxel restores IL-12 production in the TBH and ascribe a novel immunotherapeutic component to the pleiotropic activities of NO. Through its capacity to induce IL-12 production, paclitaxel may contribute to the correction of tumor-induced immune dysfunction.     

New class I and II HLA alleles strongly associated with opposite patterns of progression to AIDS.

The genetics of resistance to infection by HIV-1 cohort consists of 200 slow and 75 rapid progressors to AIDS corresponding to the extremes of HIV disease outcome of 20,000 Caucasians of European descent. A comprehensive analysis of HLA class I and class II genes in this highly informative cohort has identified HLA alleles associated with fast or slow progression, including several not described previously. A quantitative analysis shows an overall HLA influence independent of and equal in magnitude (for the protective effect) to the effect of the CCR5-Delta32 mutation. Among HLA class I genes, A29 (p = 0.001) and B22 (p < 0.0001) are significantly associated with rapid progression, whereas B14 (p = 0.001) and C8 (p = 0.004) are significantly associated with nonprogression. The class I alleles B27, B57, C14 (protective), and C16, as well as B35 (susceptible), are also influential, but their effects are less robust. Influence of class II alleles was only observed for DR11. These results confirm the influence of the immune system on disease progression and may have implications on peptide-based vaccine development.     

Chromatid damage induced by fluorescent light during G2 phase in normal and Gardner syndrome fibroblasts. Interpretation in terms of deficient DNA repair

Skin fibroblasts from Gardner syndrome (GS) compared with those from normal donors showed a significantly higher incidence of chromatid gaps and breaks following exposure to low-intensity, cool-white fluorescent light during G2 phase of the cell cycle. Considerable evidence supports the concept that chromatid gaps and breaks seen directly after exposure to DNA-damaging agents represent unrepaired DNA single- and double-strand breaks respectively. The changes in incidence of chromatid aberrations with time after light exposure are consistent with the sequence of events known to follow DNA damage and repair. Initially, the incidence of light-induced chromatid gaps was equivalent in GS and normal fibroblasts. In the normal cells, the chromatid gaps disappeared by 1 h post-exposure, presumably as a result of efficient repair of DNA single-strand breaks. In contrast, the incidence of gaps increased in GS cells by 0.5 h followed by a decrease at 1 h and concomitant increase in chromatid breaks. It appears from these findings that the increased incidence of chromatid damage in GS fibroblasts results from deficient repair of DNA single-strand breaks which arise from incomplete nucleotide excision of DNA damage during G2 phase.     

Improved microdialysis technique.

A simple, inexpensive method is described for dialysis of microliter amounts of aqueous samples against large volumes of solution with complete recovery of the fluid dialyzed. An example is given of application of the method to separation of [³H]inulin from a monosaccharide.     

Separation of precultured pollen from Nicotiana tabacum by aqueous polymer two-phase partition

Pollen isolated from cold treated and precultivated anthers of tobacco (Nicotiana tabacum L. var. Wisconsin 38) were separated into different fractions with counter-current distribution using an aqueous Dextran-polyethylene glycol two-phase system. It was possible to distinguish among eight pollen classes differing in developmental stage and in partitioning. A part of each fraction was cultivated for analysis of embryo formation. This was highest in a fraction with an intermediate to high partition in the phase system. Enriched in this fraction were also pollen that were fairly well stained with acetocarmine, contained several nuclei and had a relatively low mitochondrial activity. The enrichment of embryogenic pollen offers several advantages especially to physiological studies on embryogenesis.     

Characterization of transducin from bovine retinal rod outer segments. Participation of the amino-terminal region of T alpha in subunit interaction

The GTP-induced dissociation of T alpha from T beta gamma initiates the release of transducin from photolyzed rhodopsin and the subsequent activation of the cGMP phosphodiesterase. In this study, site-specific proteolysis and immunoprecipitation were used to map the domain of T alpha that interacts with T beta gamma. We found that Staphylococcus aureus V8 protease rapidly removes a small fragment from T alpha under native conditions, resulting in the formation of a single 38-kDa polypeptide (T alpha'). Under the same conditions, T beta gamma remains intact. A 4.5-fold decrease in the rate of T alpha cleavage by S. aureus protease was observed in the presence of T beta gamma, suggesting T beta gamma binding blocks the protease-sensitive site on T alpha. Amino acid sequence analysis indicated that T alpha' is derived from the cleavage of T alpha at Glu-21. The ability of T alpha' to interact with and activate the retinal phosphodiesterase is not diminished. However, T alpha' is unable to participate in T beta gamma-dependent activities such as the light-stimulated binding of guanine nucleotides, binding to photoexcited rhodopsin, and ADP-ribosylation catalyzed by pertussis toxin. Moreover, the anti-T alpha monoclonal antibody TF16 was able to precipitate T beta gamma in the presence of T alpha, but not with either T alpha' or T alpha-guanosine 5'-O-(3-thiotriphosphate). We conclude that the amino-terminal region of T alpha participates in T beta gamma interaction and discuss our results with respect to the known structure and function of transducin.     

Disruption of energy metabolism and smoltification during exposure of juvenile atlantic salmon (Salmo salar) to low pH

Juvenile Atlantic salmon were held for 76 days at pH 4.7 during the period when the final stages of smoltification normally occur. Control salmon (pH 6.5) had significant increases in weight, length and liver somatic index which were not observed in those held at low pH. Condition factor decreased in both groups but to a significantly greater extent in the low pH group. After 15 days levels of ADP and glucose were higher and AEC, CP and glycogen were lower in muscles of salmon exposed to low pH. These qualitative differences were maintained until the end of the experiment. ATP and total adenylate concentrations in muscle were lower after 62 days of exposure to low pH compared to controls. The levels of ATP, total adenylates, AEC and glucose were consistently higher in livers of salmon exposed to low pH than those of controls. There were no differences in liver concentrations of AMP, CP and glycogen between the two treatments or with time of exposure within each treatment. The results suggest that exposure of juvenile Atlantic salmon to low pH increased gluconeogenesis and decreased food intake to the detriment of growth.