Two human liver cDNAs encode UDP-glucuronosyltransferases with 2 log differences in activity toward parallel substrates including hyodeoxycholic acid and certain estrogen derivatives.

Two human liver UDP-glucuronosyltransferase cDNA clones, HLUG25 [Jackson, M. R., et al. (1987) Biochem. J. 242, 581-588] and UDPGTh-2 [Ritter, J. K., et al. (1990) J. Biol. Chem. 266, 7900-7906] have previously been shown to encode isozymes active in the glucuronidation of hyodeoxycholic acid (HDCA) and certain estrogen derivatives (estriols and 3,4-catechol estrogens), respectively. Here we report that the UDPGTh-2-encoded isoform (udpgth-2) and the HLUG25-encoded isoform (udpgth-1) have parallel aglycon specificities. Following expression in COS-1 cells, each isoform metabolized three types of dihydroxy- or trihydroxy-substituted ring structures, including the 3,4-catechol estrogen (4-hydroxyestrone), estriol and 17-epiestriol, and HDCA, but the udpgth-2 isozyme is 100-fold more efficient than udpgth-1. udpgth-1 and udpgth-2 are 86% identical overall (76 differences out of 528 amino acids), including 55 differences in the first 300 amino acids of the amino terminus, a domain which confers isoform substrate specificity. The data indicate that a high level of conservation in the amino terminus is not required for the preservation of substrate selectivity. Analysis of glucuronidation activity encoded by UDPGTh-1/UDPGTh-2 chimeric cDNAs constructed at their common restriction sites, SacI (codon 297), NcoI (codon 385), and HhaI (codon 469), showed that nine amino acids between residues 385 and 469 are important for catalytic efficiency, suggesting that this region represents a domain which is critical for catalysis but distinct from that responsible for aglycon selection.(ABSTRACT TRUNCATED AT 250 WORDS)     

Reconstitution of EBV-directed T cell immunity by adoptive transfer of peptide-stimulated T cells in a patient after allogeneic stem cell transplantation for AITL

Reconstitution of the T cell repertoire after allogeneic stem cell transplantation is a long and often incomplete process. As a result, reactivation of Epstein-Barr virus (EBV) is a frequent complication that may be treated by adoptive transfer of donor-derived EBV-specific T cells. We generated donor-derived EBV-specific T cells by stimulation with peptides representing defined epitopes covering multiple HLA restrictions. T cells were adoptively transferred to a patient who had developed persisting high titers of EBV after allogeneic stem cell transplantation for angioimmunoblastic T-cell lymphoma (AITL). T cell receptor beta (TCRβ) deep sequencing showed that the T cell repertoire of the patient early after transplantation (day 60) was strongly reduced and only very low numbers of EBV-specific T cells were detectable. Manufacturing and in vitro expansion of donor-derived EBV-specific T cells resulted in enrichment of EBV epitope-specific, HLA-restricted T cells. Monitoring of T cell clonotypes at a molecular level after adoptive transfer revealed that the dominant TCR sequences from peptide-stimulated T cells persisted long-term and established an EBV-specific TCR clonotype repertoire in the host, with many of the EBV-specific TCRs present in the donor. This reconstituted repertoire was associated with immunological control of EBV and with lack of further AITL relapse.     

Functional analysis of phosphorylation of the mitotic centromere-associated kinesin by Aurora B kinase in human tumor cells

Mitotic centromere-associated kinesin (MCAK) is the best characterized member of the kinesin-13 family and plays important roles in microtubule dynamics during mitosis. Its activity and subcellular localization is tightly regulated by an orchestra of mitotic kinases, such as Aurora B. It is well known that serine 196 of MCAK is the major phosphorylation site of Aurora B in Xenopus leavis extracts and that this phosphorylation regulates its catalytic activity and subcellular localization. In the current study, we have addressed the conserved phosphorylation site serine 192 in human MCAK to characterize its function in more depth in human cancer cells. Our data confirm that S192 is the major phosphorylation site of Aurora B in human MCAK and that this phosphorylation has crucial roles in regulating its catalytic activity and localization at the kinetochore/centromere region in mitosis. Interfering with this phosphorylation leads to a delayed progression through prometa- and metaphase associated with mitotic defects in chromosome alignment and segregation. We show further that MCAK is involved in directional migration and invasion of tumor cells, and interestingly, interference with the S192 phosphorylation affects this capability of MCAK. These data provide the first molecular explanation for clinical observation, where an overexpression of MCAK was associated with lymphatic invasion and lymph node metastasis in gastric and colorectal cancer patients.     

Acute cholecystitis and thiazides.

Drugs purchased by a random sample (17 000) of the population of Jämtland county, Sweden, are continuously monitored. Patients who had been admitted to the county''s only hospital with acute cholecystitis and who were part of this sample were studied, and controls matched for age and sex were drawn from the sample. The purchase of thiazides and other drugs prescribed to the patients with acute cholecystitis was compared with that of the controls. The estimated relative risk of developing acute cholecystitis in patients who had purchased thiazides in the year before admission to hospital, as compared with those who had not, was 2.1 (95% confidence limit 1.1-3.9). As it has been reliably reported that the use of thiazides is not itself associated with cholelithiasis, the association found between thiazides and cholecystitis suggests that thiazides may increase the risk of acute cholecystitis developing in a patient with gall stones.     

Native tree regeneration in native tree plantations: understanding the contribution of Araucaria angustifolia to biodiversity conservation in the threatened Atlantic Forest in Argentina

Deforestation is a global process that has strongly affected the Atlantic Forest in South America, which has been recognised as a threatened biodiversity hotspot. An important proportion of deforested areas were converted to forest plantations. Araucaria angustifolia is a native tree to the Atlantic Forest, which has been largely exploited for wood production and is currently cultivated in commercial plantations. An important question is to what extent such native tree plantations can be managed to reduce biodiversity loss in a highly diverse and vulnerable forest region . We evaluated the effect of stand age, stand basal area, as a measure of stand density, and time since last logging on the density and richness of native tree regeneration in planted araucaria stands that were successively logged over 60 years, as well as the differences between successional groups in the response of plant density to stand variables. We also compared native tree species richness in planted araucaria stands to neighbouring native forest. Species richness was 71 in the planted stands (27 ha sampled) and 82 in native forest (18 ha sampled) which approximate the range of variation in species richness found in the native forests of the study area. The total abundance and species richness of native trees increased with stand age and time since last logging, but ecological groups differed in their response to such variables. Early secondary trees increased in abundance with stand age 3–8 times faster than climax or late secondary trees. Thus, the change in species composition is expected to continue for a long term. The difference in species richness between native forest and planted stands might be mainly explained by the difference in plant density. Therefore, species richness in plantations can contribute to local native tree diversity if practices that increase native tree density are implemented.     

Linked‐read sequencing enables haplotype‐resolved resequencing at population scale

The feasibility to sequence entire genomes of virtually any organism provides unprecedented insights into the evolutionary history of populations and species. Nevertheless, many population genomic inferences – including the quantification and dating of admixture, introgression and demographic events, and inference of selective sweeps – are still limited by the lack of high‐quality haplotype information. The newest generation of sequencing technology now promises significant progress. To establish the feasibility of haplotype‐resolved genome resequencing at population scale, we investigated properties of linked‐read sequencing data of songbirds of the genus Oenanthe across a range of sequencing depths. Our results based on the comparison of downsampled (25×, 20×, 15×, 10×, 7×, and 5×) with high‐coverage data (46–68×) of seven bird genomes mapped to a reference suggest that phasing contiguities and accuracies adequate for most population genomic analyses can be reached already with moderate sequencing effort. At 15× coverage, phased haplotypes span about 90% of the genome assembly, with 50% and 90% of phased sequences located in phase blocks longer than 1.25–4.6 Mb (N50) and 0.27–0.72 Mb (N90). Phasing accuracy reaches beyond 99% starting from 15× coverage. Higher coverages yielded higher contiguities (up to about 7 Mb/1 Mb [N50/N90] at 25× coverage), but only marginally improved phasing accuracy. Phase block contiguity improved with input DNA molecule length; thus, higher‐quality DNA may help keeping sequencing costs at bay. In conclusion, even for organisms with gigabase‐sized genomes like birds, linked‐read sequencing at moderate depth opens an affordable avenue towards haplotype‐resolved genome resequencing at population scale.     

Physical and morphological aspects of the eye of the manatee trichechus inunguis natterer 1883: (Sirenia: Mammalia)

The eyes and aspects of vision of the Amazonian manatee Trichechus inunguis NATTERER 1883: (Sirenia: mammalia) have been examined in five animals, in relation to previous findings for sirenians. This paper presents a review of previous findings confirming those upon gross ocular anatomy, retinal histology and ocular refraction. The confirmation of a primarily rod retina, high receptor: ganglion cell ratio and low refractive error under water were of note. A visual pigment based upon vitamin A₁ and a difference spectrum of λ max at 505 nm was in agreement with Dartnalls nomogram. The findings suggest a retina more suited to low light levels and at best, moderate visual acuity, though motion perception appears present and a low degree of binocular vision possible.     

Bacterial Viability and Physical Properties of Antibacterially Modified Experimental Dental Resin Composites

Purpose

To investigate the antibacterial effect and the effect on the material properties of a novel delivery system with Irgasan as active agent and methacrylated polymerizable Irgasan when added to experimental dental resin composites.

Materials and Methods

A delivery system based on novel polymeric hollow beads, loaded with Irgasan and methacrylated polymerizable Irgasan as active agents were used to manufacture three commonly formulated experimental resin composites. The non-modified resin was used as standard (ST). Material A contained the delivery system providing 4 % (m/m) Irgasan, material B contained 4 % (m/m) methacrylated Irgasan and material C 8 % (m/m) methacrylated Irgasan. Flexural strength (FS), flexural modulus (FM), water sorption (WS), solubility (SL), surface roughness R_a, polymerization shrinkage, contact angle Θ, total surface free energy γ_S and its apolar γ_S ^LW, polar γ_S ^AB, Lewis acid γ_S ⁺and base γ_S ^- term as well as bacterial viability were determined. Significance was p < 0.05.

Results

The materials A to C were not unacceptably influenced by the modifications and achieved the minimum values for FS, WS and SL as requested by EN ISO 4049 and did not differ from ST what was also found for R_a. Only A had lower FM than ST. Θ of A and C was higher and γ_S ^AB of A and B was lower than of ST. Materials A to C had higher γ_S ⁺ than ST. The antibacterial effect of materials A to C was significantly increased when compared with ST meaning that significantly less vital cells were found.

Conclusion

Dental resin composites with small quantities of a novel antibacterially doped delivery system or with an antibacterial monomer provided acceptable physical properties and good antibacterial effectiveness. The sorption material being part of the delivery system can be used as a vehicle for any other active agent.