Determination of DNA ploidy in archival tissue from non-Hodgkin's lymphoma using flow and image cytometry.

Paraffin-embedded tissue from a series of 40 cases of diffuse, large cell lymphoma was analyzed by both flow and image cytometry to compare the ability of these techniques to detect DNA aneuploid populations. Image cytometry (ICM) was performed both on nuclear suspensions and tissue sections. Twenty cases (50%) were non-diploid by at least one method of analysis. Twenty-five percent of the cases were aneuploid by flow cytometry (FCM) alone. The majority of these cases were near-diploid tumors which could not be resolved by ICM. Peri-tetraploid peaks were identified by ICM of tissue sections alone in 15% of the cases. There was an apparent loss of these peri-tetraploid cells during the preparation of the nuclear suspensions. The remaining cases showed a good correlation between all three methods in the determination of DNA ploidy. Flow and image cytometry are complimentary techniques when applied to archival tissue, however aneuploid populations may be missed if ICM is not performed on tissue sections.     

In vitro effect of larval Schistosoma mansoni excretory-secretory products on phagocytosis-stimulated superoxide production in hemocytes from Biomphalaria glabrata

The in vitro production of the reactive oxygen metabolite superoxide (O2-) was confirmed in hemocytes from the schistosome intermediate host Biomphalaria glabrata. Active forms of the enzyme superoxide dismutase (SOD) inhibited reduction of nitroblue tetrazolium (NBT) to formazan in cells that had phagocytozed zymosan particles, whereas an inactivated form of SOD did not. Moreover, based on the prevalence of O2(-)-positive hemocytes and the relative intensity of NBT staining reactions, hemocytes from the Schistosoma mansoni-resistant 10-R2 strain of B. glabrata possessed an overall greater capacity for generating superoxide than did those from S. mansoni-susceptible M-line snails. Schistosoma mansoni excretory-secretory (E-S) products, released during in vitro transformation of miracidia to sporocysts, inhibited phagocytosis of zymosan particles and superoxide activity in hemocytes from both snail strains, but 10-R2 hemocytes maintained higher levels of phagocytosis and superoxide production than did M-line hemocytes. The dose-dependent decreases in phagocytosis observed in both snail strains in the presence of E-S products could not account fully for the concomitant decrease in superoxide levels detected, indicating that either a single E-S factor differentially affects phagocytosis and superoxide production, or that different E-S factors are involved in the specific interference of each of these hemocyte functions.     

A native nitrogen-fixing shrub facilitates weed invasion

Invasions by exotic weedy plants frequently occur in highly disturbed or otherwise anthropogenically altered habitats. Here we present evidence that, within California coastal prairie, invasion also can be facilitated by a native nitrogen-fixing shrub, bush lupine (Lupinus arboreus). Bush lupines fix nitrogen and grow rapidly, fertilizing the sandy soil with nitrogen-rich litter. The dense lupine canopy blocks light, restricting vegetative growth under bushes. Heavy insect herbivory kills lupines, opening exposed nitrogen-rich sites within the plant community. Eventual re-establishment of lupine occurs because of an abundant and long-lived seed bank. Lupine germination, rapid growth, shading and fertilization of sites, and then death after only a few years, results in a mosaic of nutrient-rich sites that are available to invading species. To determine the role of bush lupine death and nitrogen enrichment in community composition, we examined nutrient dynamics and plant community characteristics within a site only recently colonized by lupine, comparing patches where lupines had recently died or were experimentally killed with adjacent areas lacking lupine. In experimentally killed patches, instantaneous pool sizes of exchangeable ammonium and nitrate nitrogen were higher than in adjacent sites free of lupine. Seedlings of the introduced grass Bromus diandrus accumulated 48% greater root biomass and 93% more shoot biomass when grown in a greenhouse in soil collected under experimentally killed lupines compared to B. diandrus seedlings grown in soil collected at least 1 m away from lupines. At the end of the spring growing season, total above-ground live plant biomass was more than twice as great in dead lupine patches as in the adjacent lupine-free grassland, but dead lupine patches contained 47% fewer plant species and 57% fewer native species. Sites where lupines have repeatedly died and reestablished during recent decades support an interstitial grassland community high in productivity but low in diversity, composed of mostly weedy introduced annual plants. In contrast, at a site only recently colonized by bush lupines, the interstitial grassland consists of a less productive but more diverse set of native and introduced species. We suggest that repeated bouts of lupine germination, establishment, and death can convert a rich native plant community into a less diverse collection of introduced weeds.     

The cytolytic activity of pulmonary CD8+ lymphocytes, induced by infection with a vaccinia virus recombinant expressing the M2 protein of respiratory syncytial virus (RSV), correlates with resistance to RSV infection in mice.

Previous studies demonstrated that the pulmonary resistance to respiratory syncytial virus (RSV) challenge induced by immunization with a recombinant vaccinia virus expressing the M2 protein of RSV (vac-M2) was significantly greater 9 days after immunization than at 28 days and was mediated predominantly by CD8+ T cells. In this study, we have extended these findings and sought to determine whether the level of CD8+ cytotoxic T-lymphocyte (CTL) activity measured in vitro correlates with the resistance to RSV challenge in vivo. Three lines of evidence documented an association between the presence of pulmonary CTL activity and resistance to RSV challenge. First, vac-M2 immunization induced pulmonary CD8+ CTL activity and pulmonary resistance to RSV infection in BALB/c (H-2d) mice, whereas significant levels of pulmonary CTL activity and resistance to RSV infection were not seen in BALB.K (H-2k) or BALB.B (H-2b) mice. Second, pulmonary CD8+ CTL activity was not induced by infection with other vaccinia virus-RSV recombinants that did not induce resistance to RSV challenge. Third, the peak of pulmonary CTL activity correlated with the peak of resistance to RSV replication (day 6), with little resistance being observed 45 days after immunization. An accelerated clearance of virus was not observed when mice were challenged with RSV 45 days after immunization with vac-M2. The results indicate that resistance to RSV induced by immunization with vac-M2 is mainly mediated by primary pulmonary CTLs and that this resistance decreases to very low levels within 2 months following immunization. The implications for inclusion of CTL epitopes into RSV vaccines are discussed in the context of these observations.     

Influence of culture parameters on lignin metabolism byPhanerochaete chrysosporium

Culture parameters influencing metabolism of synthetic¹⁴C-lignins to¹⁴CO₂ in defined media have been studied in shallow batch cultures of the ligninolytic wood-destroying HymenomycetePhanerochaete chrysosporium Burds. Study of the effect of O₂ concentration in the gas phase above non-agitated cultures indicated essentially complete absence of attack on the lignin polymer at 5% O₂ in N₂, and a 2- to 3-fold enhancement by 100% O₂ as compared to air (21% O₂). Agitation of the cultures resulting in the formation of mycelial pellets greatly suppressed lignin decomposition. The optimum culture pH for lignin decomposition was 4 to 4.5, with marked suppression above 5.5 and below 3.5. The source of nutrient nitrogen (NO ₃ ^– , NH ₄ ⁺ , amino acids) had little influence on lignin decomposition, but the concentration of nitrogen was critical; decomposition at 24 mM was only 25–35% of that at 2.4 mM N. Thiamine was the only vitamin required for growth and lignin decomposition. Under the optimum conditions developed, decomposition of 5 mg of synthetic lignin was accompanied by utilization of approximately 100 mg of glucose. The influence of the various culture parameters was analogous for metabolism of synthetic lignin labeled in the ring-,side chain-, and methoxyl carbon atoms.     

Alteration of stimulated TSH and prolactin response in children treated with growth hormone.

The plasma TSH and prolactin responses to thyrotropin releasing hormone (TRH) were measured in 5 children with isolated growth hormone deficiency prior to, during and after the administration of human growth hormone (hGH). TSH and prolactin secretory patterns were not uniformly concordant. TSH responses to TRH infusion were suppressed in 4 subjects after 5 days or 1 month of hGH administration despite normal serum thyroxin concentrations. Prolactin responses were suppressed in all 5 subjects after 5 days of hGH administration. After 8 months of hGH therapy both TSH and prolactin responses returned toward pre-hGH values. Our finding that suppression of the TRH-induced TSH and prolactin secretory responses are reversible during hGH administration supports the concept of altered neuroregulation in this form of hypothalamic disorder.     

Identification of a Schistosoma mansoni sporocyst excretory-secretory antioxidant molecule and its effect on superoxide production by Biomphalaria glabrata hemocytes.

Excretory-secretory (E-S) products obtained during in vitro Schistosoma mansoni miracidium-to-sporocyst transformation were found to contain a 108-kDa polypeptide capable of scavenging both exogenously produced and M-line Biomphalaria glabrata hemocyte-derived superoxide (O2-) anions. Separation of crude transformation E-S products using HPLC and ion exchange chromatography resulted in the separation of two isoforms of the 108-kDa molecule. Using an in vitro phagocytosis assay, both isoforms were found to be capable of reducing O2- production by phagocytically stimulated M-line B. glabrata hemocytes without cell loss and without a concomitant reduction in phagocytosis. Although parasite antioxidant molecules appear to play a role in the evasion of host oxidative defense systems in several parasite-vertebrate systems, no previous reports of a parasite antioxidant capability against the potential of oxidative killing by invertebrate defense systems has been reported. In conjunction with the previously confirmed production of O2- by B. glabrata hemocytes and reports of reactive oxygen metabolite production by hemocytes from several molluscan species, these results indicate that reactive forms of oxygen and parasite antioxidant systems may play an important role in the determination of compatibility in the trematode-mollusc relationship.