Metabolism and renal clearance of 111In-labeled DOTA-conjugated antibody fragments

Radiometal-labeled antibody fragments are promising reagents for radioimmunotherapy due to their high tumor uptake and rapid pharmacokinetics, but their therapeutic potentials are limited by high uptake and retention in the kidney. Identification of metabolic products is a first step in designing rationale approaches to lower kidney uptake. Previous studies in rats have shown that 111In-labeled DTPA-conjugated antibody fragments (via lysine residues) were degraded to an DTPA-epsilon-amino-lysine derivative and retained in the lysosomal compartments of the liver and kidney [Rogers et al. (1995) Cancer Res. 55, 5714s-5720s]. To determine the metabolic profile of another widely used metal-chelate, [111In]DOTA conjugated to lysines in antibody fragments via active ester chemistry, we analyzed kidney homogenates from nude mice injected with an [111In]DOTA-Fab generated enzymatically from the anti-lymphoma intact antibody Rituxan. The major kidney metabolite was identified as [111In]DOTA-epsilon-amino-lysine by comparison to an authentic synthetic standard. This end product was also identified in the urine, along with relatively small amounts of [111In]DOTA-Fab. Since injection of [111In]DOTA-epsilon-amino-lysine into nude mice resulted in rapid clearance into the urine without kidney retention, it is likely that the renal retention observed was due to kidney uptake of [111In]DOTA-Fab, followed by lysosomal degradation to [111In]DOTA-epsilon-amino-lysine, which is only slowly cleared from this compartment. This observation is supported by autoradiographs of the kidney showing rapid localization of radioactivity into the distal regions of the kidney cortex. To extend this analysis to clinical trials, we have also analyzed urine taken from a patient injected with the intact antibody [111In]DOTA-cT84.66. In that example, we found that the major radioactive species was also [111In]DOTA-epsilon-amino-lysine.     

粤北阴香不同器官中精油成分研究

采用水蒸汽同步蒸馏法分别对采自粤北的阴香的果、叶、枝条提取精油,并用气质联用仪对所提取的精油进行了成分分析和鉴定。共鉴定了41种成分,主要成分为龙脑（68．5％～73．8％）、芳香醇、丁子香烯、橙花叔醇、榄香烯、柠檬醛、莰烯、葑醇、愈创烯、香叶烯、枞萜、萜品醇、蒎烯等。阴香不同器官（果、叶、枝）中的精油成分及所占比例均有差异。     

Detection of RNAase inhibitor from different species and organs

Human placental alkaline RNAase inhibitor was purified to homogeneity. Activity was measured after each purification step. The final identification of the purified protein was done by two-dimensional polyacrylamide gel electrophoresis and by immunoblotting. Antibodies were prepared by immunization of rabbits with the highly purified inhibitor. The availability of the antiserum directed against the human inhibitor enabled the detection of RNAase inhibitor from various other organs and species. This procedure has the advantage over the usual activity test in that the inhibitor can be found even if its activity has been lost.     

Synergistic roles of antibody and interferon in noncytolytic clearance of Sindbis virus from different regions of the central nervous system

Sindbis virus (SINV) is an alphavirus that causes infection of neurons and encephalomyelitis in adult immunocompetent mice. Recovery can occur without apparent neurological damage. To better define the factors facilitating noncytolytic clearance of SINV in different regions of the central nervous system (CNS) and the roles of innate and adaptive immune responses at different times during infection, we have characterized SINV infection and clearance in the brain, brain stem, and spinal cords of severe combined immunodeficiency (SCID) and C57BL/6 (wild-type [WT]) mice and mice deficient in beta interferon (IFN-beta) (BKO), antibody (muMT), IFN-gamma (GKO), IFN-gamma receptor (GRKO), and both antibody and IFN-gamma (muMT/GKO). WT mice cleared infectious virus by day 8, while SCID mice had persistent virus replication at all sites. For 3 days after infection, BKO mice had higher titers at all sites than WT mice, despite similar IFN-alpha production, but cleared virus similarly. GKO and GRKO mice cleared infectious virus from all sites by days 8 to 10 and, like WT mice, displayed transient reactivation at 12 to 22 days. muMT mice did not clear virus from the brain, and clearance from the brain stem and lumbar spinal cord was delayed, followed by reactivation. Eighty-one days after infection, muMT/GKO mice had not cleared virus from any site, but titers were lower than for SCID mice. These studies show that IFN-beta is independently important for early control of CNS virus replication, that antiviral antibody is critical for clearance from the brain, and that both antibody and IFN-gamma contribute to prevention of reactivation after initial clearance.     

桑叶不同化学成分的降血糖作用

桑叶是一种重要的降血糖药用植物资源。以秋桑叶中提取的六种活性化学成分为材料,研究了六种提取物对四氧嘧啶诱导的糖尿病小鼠的降血糖效果。结果显示,各提取物均有一定的降血糖作用,其中以桑叶总多糖的功效最为显著,桑叶生物碱及桑叶黄酮苷类效果次之。     

Altered biodistribution of indium-111-labeled monoclonal antibody 96.5 to tumors and normal tissues of nude mice bearing human melanoma xenografts in visceral organs

Summary Human melanoma xenografts were produced in the subcutis, kidney, cecum and liver of different nude mice. An¹¹¹In-labeled anti-(human melanoma) monoclonal antibody (96.5) or an¹¹¹In-labeled nonspecific control monoclonal antibody (ZCE-025) was injected intravenously in separate groups of mice. Radioactive antibody accumulation was measured in tumor, blood, viscera, and carcasses. mAb 96.5 targeted specifically to tumor tissue regardless of site of growth. Tumors in the liver exhibited significantly (P <0.05) higher tumor-to-blood ratios (45±6, mean ±SEM) than xenografts at other visceral organs, the lowest value being found for subcutaneous melanoma (2.6±0.5). The differences in tumor-to-blood ratio were due to significant alterations of antibody biodistribution, since the actual antibody concentration in the different tumor sites was similar. The percentage of recovered anti-melanoma antibody per milliliter of blood in mice with visceral lesions (4.6±1.1%/ml) was significantly lower than that found in mice with subcutaneous tumors (9.5±1.4%/ml,P <0.05). Moreover, significantly higher levels (18.2±3.2%/g, 31.0±5.1%/g, respectively) of the melanoma mAb 96.5 were found in normal liver and spleen tissue recovered from mice with visceral tumors as compared to tissue from mice with subcutaneous tumors (9.2±0.9%/g, 13.5±1.9%/g, respectively;P <0.05). These results demonstrate that the presence of visceral tumor can significantly affect tumor-to-blood ratios, blood levels, and biodistribution of¹¹¹In-labeled mAb 96.5.This work was supported in part by funds from the National Institutes of Health, National Cancer Institute, Grant R35-CA42107 and Core Grant CA16672     

Trophic linkages between periphyton and grazing macroinvertebrates in rivers with different levels of catchment development

Stable isotope signatures (δ¹³C and δ¹⁵N) were used to compare trophic linkages between epilithic periphyton and three families of macroinvertebrates (Baetidae, Leptophlebiidae and Gripopterygidae) in riffles of two rivers with developed catchments (including agriculture, urbanization, impoundment and flow regulation) and two with undeveloped catchments (native forest with no major impoundments) in the Murrumbidgee River system, New South Wales, Australia. Periphyton had much higher average δ¹⁵N values and lower average C:N ratios in the developed rivers than in the undeveloped rivers, probably because of the combined effects of nutrient enrichment, upstream impoundment and alteration of riparian vegetation. The invertebrates were generally slightly depleted in ¹³C and ¹⁵N relative to expected values if they were assimilating whole periphyton alone, which suggests that they were assimilating periphyton components selectively or also consuming other foods. The match in isotope signatures between periphyton and invertebrates was only slightly weaker in the developed than in the undeveloped rivers, suggesting that development did not greatly disrupt trophic linkages between periphyton and these invertebrates. Handling editor: M. Power     

Physical and chemical characterization of lamb meat from different genotypes submitted to diet with different fibre contents

The objective of the study was to evaluate the effect of genotype and the dietary fibre concentration in the chemical composition and physical properties of lamb meat. Samples from 54 animals from Morada Nova and Santa Inês native breeds and Dorpper × Santa Inês half-breed were analyzed, which received two diets, one with 41.7% and another with 33.6% fibre content, until reaching the average slaughter weight of 30 kg. The design used was fully randomized 3 × 2 factorial, three genotypes and two diets with nine replicates. Colour and pH in Semimembranosus muscle were determined, together with the analyses of the water retention capacity, loss of weight due to cooking, shearing force, and chemical composition in Longissimus dorsi muscle. The factor genotype influenced the chemical composition of meat, with Santa Inês lamb and crossbreed showing the highest protein percentages. The diet with 33.6% fibre content provided meat with higher moisture percentage and lower protein value, and the loss of weight due to cooking and shearing force parameters did not interfere in other variables. In addition, lambs receiving higher fibre content diet produced meat with lower shear force values, having indicated to be more tenderness. Despite these variations, the meat of lambs from all genotypes and under the diets evaluated can be considered of good quality.     

The use of natural abundance stable isotopic ratios to indicate the presence of oxygen-containing chemical linkages between cellulose and lignin in plant cell walls

Qualitative and quantitative understanding of the chemical linkages between the three major biochemical components (cellulose, hemicellulose and lignin) of plant cell walls is crucial to the understanding of cell wall structure. Although there is convincing evidence for chemical bonds between hemicellulose and lignin and the absence of chemical bonds between hemicellulose and cellulose, there is no conclusive evidence for the presence of covalent bonds between cellulose and lignin. This is caused by the lack of selectivity of current GC/MS-, NMR- and IR-based methods for lignin characterisation as none of these techniques directly targets the possible ester and ether linkages between lignin and cellulose. We modified the widely-accepted “standard” three-step extraction method for isolating cellulose from plants by changing the order of the steps for hemicellulose and lignin removal (solubilisation with concentrated NaOH and oxidation with acetic acid-containing NaClO₂, respectively) so that cellulose and lignin could be isolated with the possible chemical bonds between them intact. These linkages were then cleaved with NaClO₂ reagent in aqueous media of contrasting ¹⁸O/¹⁶O ratios. We produced cellulose with higher purity (a lower level of residual hemicellulose and no detectable lignin) than that produced by the “standard” method. Oxidative artefacts may potentially be introduced at the lignin removal stage; but testing showed this to be minimal.Cellulose samples isolated from processing plant-derived cellulose–lignin mixtures in media of contrasting ¹⁸O/¹⁶O ratios were compared to provide the first quantitative evidence for the presence of oxygen-containing ester and ether bonds between cellulose and lignin in Zea mays leaves. However, no conclusive evidence for the presence or lack of similar bonds in Araucaria cunninghamii wood was obtained.     

Contribution of mineral nutrients from source to sink organs in rice under different nitrogen fertilization

The pot experiment with three treatments of nitrogen (N) topdressing was performed with the japonica rice cultivar viz. Huaidao 5. Remobilization of nine mineral nutrients including N, phosphorus (P), potassium (K), calcium (Ca), magnesium (Mg), iron (Fe), zinc (Zn), manganese (Mn), and copper (Cu) was measured from the source organs including bracts, leaf, and sheath to sink rice grain. Experimental results showed considerable contribution of bracts to grain for N, Mg, and Zn, with the averages contributions of 5.96, 12.56, and 12.34%, respectively, indicating a positive role of rice bracts in N, Mg, and Zn remobilization during grain filling. By contrast, minor contribution of bracts to grain P, K, and Cu was revealed, with the contribution rate being 0.99, 3.90, and 3.05%, respectively. Further, a net increase in Ca and Fe concentrations of bracts was detected, implying that bracts function as a sink of these mineral nutrients. In addition, grains produced at a moderate level of N topdressing had higher Fe and similar Zn concentration in comparison with those at high N level, suggesting the possibility of N management for maintaining Fe and Zn level under high yielding conditions.     

Different coexpressions of arthritis-relevant genes between different body organs and different brain regions in the normal mouse population

Structural changes in different parts of the brain in rheumatoid arthritis (RA) patients have been reported. RA is not regarded as a brain disease. Body organs such as spleen and lung produce RA-relevant genes. We hypothesized that the structural changes in the brain are caused by changes of gene expression in body organs. Changes in different parts of the brain may be affected by altered gene expressions in different body organs. This study explored whether an association between gene expressions of an organ or a body part varies in different brain structures. By examining the association of the 10 most altered genes from a mouse model of spontaneous arthritis in a normal mouse population, we found two groups of gene expression patterns between five brain structures and spleen. The correlation patterns between the prefrontal cortex, nucleus accumbens, and spleen were similar, while the associations between the other three parts of the brain and spleen showed a different pattern. Among overall patterns of the associations between body organs and brain structures, spleen and lung had a similar pattern, and patterns for kidney and liver were similar. Analysis of the five additional known arthritis-relevant genes produced similar results. Analysis of 10 nonrelevant-arthritis genes did not result in a strong association of gene expression or clearly segregated patterns. Our data suggest that abnormal gene expressions in different diseased body organs may influence structural changes in different brain parts.     

Genetic down-regulation of gibberellin results in semi-dwarf poplar but few non-target effects on chemical resistance and tolerance to defoliation

Aims Plant stature can be strongly modified via regulation of endogenous levels and signalling of the plant hormone gibberellin (GA). Down-regulation of GA can produce semi-dwarf tree varieties with improved qualities such as reduced susceptibility to wind damage, enhanced root growth and more compact cultivation. However, these modifications may have unintended, non-target consequences for defence against herbivores, via either of two mechanisms: (i) reduced biomass production may cause trade-offs with chemical resistance traits, as predicted by the growth-differentiation balance hypothesis, and (ii) altered biomass allocation to either roots or photosynthetic tissues may affect regrowth potential and thus tolerance to defoliation.     

Protective systems of immunocompetent organs in fishes from different ecological and systematic groups

This article presents the results of analysis of parameters of the immune, antioxidant, and monooxygenase systems in the immunocompetent organs (pro- and mesonephros, spleen, and liver) of species belonging to different systematic and ecological groups, namely, northern pike Esox lucius, zander Sander lucioperca, and bream Abramis brama. Although having the same features in the functioning of homeostatic mechanisms in the species under study, significant differences were revealed between bream and another two species with respect to the parameters of the nonspecific immunity of tissues and enzyme systems that perform protective functions. In all the three species, the basic role in processes of immunopoesis and nonspecific protection is played by the kidneys and spleen, while the greatest values of the parameters of antioxidant defenses and ethoxyresorufin-O-deethylase activity were recorded in the liver.     

DNA adducts of styrene-7,8-oxide in target and non-target organs for tumor induction in rat and mouse after repeated inhalation exposure to styrene

Styrene by inhalation had been shown to increase the lung tumor incidence in mice at 20 ppm and higher, but was not carcinogenic in rats at up to 1000 ppm. Styrene-7,8-oxide, the major metabolic intermediate, has weak electrophilic reactivity. Therefore, DNA adduct formation was expected at a low level and a 32P-postlabeling method for a determination of the two regioisomeric 2'-deoxyguanosyl-O6-adducts at the alpha(7)- and beta(8)-positions had been established. The first question was whether DNA adducts could be measured in the rat at the end of the 2 years exposure of a bioassay for carcinogenicity, even though tumor incidence was not increased. Liver samples of male and female CD rats were available for DNA adduct analysis. Adducts were above the limit of detection only in the highest dose group (1000 ppm), with median levels of 9 and 8 adducts per 10(7) nucleotides in males and females, respectively (sum of alpha- and beta-adducts). The result indicates that the rat liver tolerated a relatively high steady-state level of styrene-induced DNA adducts without detectable increase in tumor formation. The second question was whether different DNA adduct levels in the lung of rats and mice could account for the species difference in tumor incidence. Groups of female CD-1 mice were exposed for 2 weeks to 0, 40, and 160 ppm styrene (6h per day; 5 days per week), female CD rats were exposed to 0 and 500 ppm. In none of the lung DNA samples were adducts above a limit of detection of 1 adduct per 10(7) DNA nucleotides. The data indicate that species- and organ-specific tumor induction by styrene is not reflected by DNA adduct levels determined in tissue homogenate. The particular susceptibility of the mouse lung might have to be based on other reactive metabolites and DNA adducts, indirect DNA damage and/or cell-type specific toxicity and tumor promotion.     

Rate of clearance of interleukin-1 from the blood of normal and nephrectomized rats

Interleukin-1 (IL-1), a macrophage product, appears to be responsible for a wide variety of changes in animals during the early stages of infection and inflammation. However, the fate of injected IL-1 has not been established. The major pathway of removal from the circulation may be through the kidney, since IL-1 can be found in urine. The IL-1 for these studies was prepared from peritoneal macrophages of rats and rabbits. The clearance rate of IL-1 was determined by measuring the activity, for increasing plasma fibrinogen or releasing bone marrow neutrophils, which remained in the blood at various times after an iv injection. Both rat and rabbit IL-1 were removed rapidly from the blood of rats. The results indicate that less than 10% of the IL-1 was cleared by the kidney. So that rats with their kidneys removed showed only a slight decrease in the clearance rate of IL-1. This suggests that excretion through the kidney does not represent the major mechanism of IL-1 disposal.     

Characterization of blood pressure and morphological traits in cardiovascular-related organs in 13 different inbred mouse strains.

To better understand the contributions of various genetic backgrounds to complex quantitative phenotypes, we have measured several quantitative traits of cardiovascular interest [i.e., systolic blood pressure, weight (corrected by body weight) of several cardiac compartments and adrenals and kidneys, and histological correlates for kidneys and adrenals] in male and female mice from 13 different inbred strains. We selected strains so that each major genealogical group would be represented and to conform to priorities set by the Mouse Phenome Database project. Interstrain comparisons of phenotypes made it possible to identify strains that displayed values that belonged to either the low or the high end of the interstrain variance for quantitative traits, such as systolic blood pressure, body weight, left ventricular weight, and/or adrenocortical structure. For instance, both male and female C3H/HeJ and A/J mice displayed either low systolic blood pressure or low cardiac ventricular mass, respectively, and male C57BL6/J displayed low adrenal weight. Likewise, intersex comparisons made it possible to identify phenotypic values that were sexually dimorphic for some of the same traits. For instance, female AKR/J mice had relatively higher body weight and systolic blood pressure values than their male counterparts, perhaps constituting an animal model of the metabolic X syndrome. These strain- and sex-specific features will be of value both for future genetic and/or developmental studies and for the development of new animal models that will help in the generation of mechanistic hypotheses. All data have been deposited to the Mouse Phenome Database for future integration with the Mouse Genome Database and can be further analyzed and compared with tools available on the site.