IL-2 in vivo activities and antitumor efficacy enhanced by an anti-IL-2 mAb

IL-2 is a potent immunostimulant and has been tested for clinical use, including in immunotherapy for cancers and HIV infection. Here we show that a widely used neutralizing anti-murine IL-2 mAb (S4B6) exhibits unexpected activities that enhance the treatment effects of IL-2 in vivo. Coinjection of the anti-IL-2 mAb with a plasmid carrying murine IL-2 cDNA significantly increased the serum IL-2 levels and induced a substantial increase in the division of CD8+ T and NK1.1(high) cells in vivo. Injection of the mAb premixed with recombinant murine IL-2 showed the same enhanced effect. A 5-day treatment with the anti-IL-2 mAb alone gradually increased the CD44(high)CD8+ population, and the increased population was maintained for >300 days, suggesting that the mAb can gradually maintain and potentially enhance the bioactivity of endogenous IL-2 for extended periods. Furthermore, combined treatment with the anti-IL-2 mAb plus the IL-2 plasmid markedly enhanced Ag-specific CTL activity in vivo and partially protected mice from tumor metastasis to the lungs, compared with the anti-IL-2 mAb or IL-2 plasmid alone. These results demonstrated IL-2-enhancing effects of the anti-IL-2 mAb in vivo and suggest that combining a neutralizing anti-IL-2 Ab with IL-2 gene delivery might be used effectively to enhance IL-2 functions in clinical applications.     

Analysis of Grassland Vegetation of the Southwest Heilongjiang Steppe (China) Using the Power Law

In 1997, we conducted a vegetation survey in three semi-arid natural grasslands (steppes) withdifferent livestock grazing intensities in Southwest Heilongjiang Province, China. The dominant grasslandspecies was the grass Stipa baicalensis Roshev. Grasslands with light, intermediate, and heavy grazingintensities were located 10, 5, and 2 km from a village, respectively. Villagers use the steppe to raise cattle,horses, sheep, and goats. Each of the three grasslands was surveyed by placing 100 quadrats (50 cm×50 cm)along a 50 m line transect. Each quadrat was divided into four equal areas (25 cm×25 cm; S-quadrats) and allplant species occurring in each of these smaller areas were identified and recorded. These data were summa-rized into frequency distributions and the percentage of S-quadrats containing a given species and thevariance of each species were estimated. The power law was applied to these estimates. The power law wasused to evaluate the spatial heterogeneity and frequency of occurrence for each species in the grasslandcommunity. The lightly grazed grassland exhibited high spatial heterogeneity (caused by large plant size),the highest species diversity, and a high occurrence of S. baicalensis. In contrast, the heavily grazedgrassland exhibited high spatial heterogeneity (caused by patchy populations of small plant size), lowspecies diversity, and a low occurrence of S. baicalensis. We judged that the heavily grazed grassland wasovergrazed and exclusion of livestock from the degraded areas is necessary for recovery.     

Enzymatic and Genetic Characterization of Carbon and Energy Metabolisms by Deep-Sea Hydrothermal Chemolithoautotrophic Isolates of Epsilonproteobacteria

The carbon and energy metabolisms of a variety of cultured chemolithoautotrophic Epsilonproteobacteria from deep-sea hydrothermal environments were characterized by both enzymatic and genetic analyses. All the Epsilonproteobacteria tested had all three key reductive tricarboxylic acid (rTCA) cycle enzymatic activities—ATP-dependent citrate lyase, pyruvate:ferredoxin oxidoreductase, and 2-oxoglutarate:ferredoxin oxidoreductase—while they had no ribulose 1,5-bisphosphate carboxylase (RubisCO) activity, the key enzyme in the Calvin-Benson cycle. These results paralleled the successful amplification of the key rTCA cycle genes aclB, porAB, and oorAB and the lack of success at amplifying the form I and II RubisCO genes, cbbL and cbbM. The combination of enzymatic and genetic analyses demonstrates that the Epsilonproteobacteria tested use the rTCA cycle for carbon assimilation. The energy metabolisms of deep-sea Epsilonproteobacteria were also well specified by the enzymatic and genetic characterization: hydrogen-oxidizing strains had evident soluble acceptor:methyl viologen hydrogenase activity and hydrogen uptake hydrogenase genes (hyn operon), while sulfur-oxidizing strains lacked both the enzyme activity and the genes. Although the energy metabolism of reduced sulfur compounds was not genetically analyzed and was not fully clarified, sulfur-oxidizing Epsilonproteobacteria showed enzyme activity of a potential sulfite:acceptor oxidoreductase for a direct oxidation pathway to sulfate but no activity of AMP-dependent adenosine 5′-phosphate sulfate reductase for a indirect oxidation pathway. No activity of thiosulfate-oxidizing enzymes was detected. The enzymatic and genetic characteristics described here were consistent with cellular carbon and energy metabolisms and suggest that molecular tools may have great potential for in situ elucidation of the ecophysiological roles of deep-sea Epsilonproteobacteria.     

Temporal and positional relationships between Mn uptake and low-pH-induced root hair formation in Lactuca sativa cv. Grand Rapids seedlings

We investigated whether low-pH-induced manganese (Mn) deficiency causes low-pH-induced root hair formation in lettuce seedlings. Both the number and length of root hairs increased in 0 μM Mn (Mn-free) at pH 6 and decreased in 3 mM Mn (excess Mn) at pH 4 compared with the values in 10 μM Mn (normal Mn). These results indicate an inhibitory effect of Mn on both root hair initiation and elongation. The time dependency of root hair induction caused by Mn deficiency corresponded to that caused by low pH. Within 1 h after the pH of the culture medium was reduced from pH 6 to pH 4, the Mn uptake by roots decreased to 43% of that at pH 6. These results suggest that low-pH-induced Mn deficiency promotes root hair formation. At low pH, the rate of Mn uptake was reduced in areas >2 mm from the root tip. Roots with low-pH-induced root hairs still showed low Mn uptake during 3 h of incubation at pH 6. Therefore, the additional root hairs induced by low pH did not compensate for the low-pH-induced decrease in Mn uptake     

UV-B radiation induces epithelial tumors in mice lacking DNA polymerase eta and mesenchymal tumors in mice deficient for DNA polymerase iota

DNA polymerase eta (Pol eta) is the product of the Polh gene, which is responsible for the group variant of xeroderma pigmentosum, a rare inherited recessive disease which is characterized by susceptibility to sunlight-induced skin cancer. We recently reported in a study of Polh mutant mice that Pol eta is involved in the somatic hypermutation of immunoglobulin genes, but the cancer predisposition of Polh-/- mice has not been examined until very recently. Another translesion synthesis polymerase, Pol iota, a Pol eta paralog encoded by the Poli gene, is naturally deficient in the 129 mouse strain, and the function of Pol iota is enigmatic. Here, we generated Polh Poli double-deficient mice and compared the tumor susceptibility of them with Polh- or Poli-deficient animals under the same genetic background. While Pol iota deficiency does not influence the UV sensitivity of mouse fibroblasts irrespective of Polh genotype, Polh Poli double-deficient mice show slightly earlier onset of skin tumor formation. Intriguingly, histological diagnosis after chronic treatment with UV light reveals that Pol iota deficiency leads to the formation of mesenchymal tumors, such as sarcomas, that are not observed in Polh(-/-) mice. These results suggest the involvement of the Pol eta and Pol iota proteins in UV-induced skin carcinogenesis.     

Suppression of protein l-isoaspartyl (d-aspartyl) methyltransferase results in hyperactivation of EGF-stimulated MEK-ERK signaling in cultured mammalian cells

l-Aspartyl (l-Asp) and l-asparaginyl residues in proteins isomerize or racemize to d,l-isoaspartyl (d,l-isoAsp) or d-aspartyl (d-Asp) residues during protein aging. These atypical aspartyl residues can interfere with the biological function of the protein and lead to cellular dysfunction. Protein l-isoaspartyl (d-aspartyl) methyltransferase (PIMT) is a repair enzyme that facilitates conversion of l-isoAsp and d-Asp to l-Asp. PIMT deficient mice exhibit accumulation of l-isoAsp in several tissues and die, on average, 12 days after birth from progressive epileptic seizures with grand mal and myoclonus features. However, little is known about the molecular mechanisms by which accumulation of the aberrant residues leads to cellular abnormalities. In this study, we established PIMT-knockdown cells using a short interfering RNA expression system and characterized the resultant molecular abnormalities in intracellular signaling pathways. PIMT-knockdown cells showed significant accumulation of proteins with isomerized residues, compared to control cells. In the PIMT-knockdown cells, Raf-1, MEK, and ERK, members of the MAPK cascade, were hyperphosphorylated after EGF stimulation compared to control cells. These results suggest that PIMT repair of abnormal proteins is necessary to maintain normal MAPK signaling.     

Small-scale species richness and its spatial variation in an alpine meadow on the Qinghai-Tibet Plateau

We investigated how the high small-scale species richness of an alpine meadow on the Qinghai-Tibet Plateau, China, is maintained. This area is characterized by strong wind and severe cold during long winters. In winter, most livestock is grazed on dead leaves in small pastures near farmers’ residences, whereas in the short summer, livestock is grazed in mountainous areas far from farmers’ residences. The number of plant species and the aboveground biomass were surveyed for three adjacent pastures differing in grazing management: a late-winter grazing pasture grazed moderately from 1 February to 30 April, an early-winter grazing pasture grazed lightly from 20 September to late October, and a whole-year grazing pasture grazed intensively throughout the entire year. In each pasture, we harvested the aboveground biomass from 80 or 100 quadrats of 0.01 m² along a transect and classified the contents by species. We observed 15.5–19.7 species per 0.01 m², which is high richness per 0.01 m² on a worldwide scale. The species richness in the two winter grazing pastures was higher than that in the whole-year grazing pasture. The spatial variation in species richness and species composition in the two winter grazing pastures in which species richness was high was greater than that in the whole-year grazing pasture in which species richness was lower. Most of the leaves that are preserved on the winter grazing pastures during summer are blown away by strong winds during winter, and the remaining leaves are completely exhausted in winter by livestock grazing. A pasture with a high richess is accompanied by a high spatial variation in species richness and species composition. There is a high possibility that the characteristic of spatial variation is also caused by traditional grazing practices in this area.     

Cardiovascular response to functional electrical stimulation and dynamic tilt table therapy to improve orthostatic tolerance

Orthostatic hypotension is a common condition for individuals with stroke or spinal cord injury. The inability to regulate the central nervous system will result in pooling of blood in the lower extremities leading to orthostatic intolerance. This study compared the use of functional electrical stimulation (FES) and passive leg movements to improve orthostatic tolerance during head-up tilt. Four trial conditions were assessed during head-up tilt: (1) rest, (2) isometric FES of the hamstring, gastrocnemius and quadriceps muscle group, (3) passive mobilization using the Erigo dynamic tilt table; and (4) dynamic FES (combined 2 and 3). Ten healthy male subjects experienced 70 degrees head-up tilt for 15 min under each trial condition. Heart rate, blood pressure and abdominal echograms of the inferior vena cava were recorded for each trial. Passive mobilization and dynamic FES resulted in an increase in intravascular blood volume, while isometric FES only resulted in elevating heart rate. No significant differences in blood pressure were observed under each condition. We conclude that FES combined with passive stepping movements may be an effective modality to increase circulating blood volume and thereby tolerance to postural hypotension in healthy subjects.