Subunit assembly of hemoglobin: an important determinant of hematologic phenotype

Hemoglobin's physiologic properties depend on the orderly assembly of its subunits in erythropoietic cells. The biosynthesis of alpha- and beta-globin polypeptide chains is normally balanced. Heme rapidly binds to the globin subunit, either during translation or shortly thereafter. The formation of the alpha beta-dimer is facilitated by electrostatic attraction of a positively charged alpha-subunit to a negatively charged beta-subunit. The alpha beta-dimer dissociates extremely slowly. The difference between the rate of dissociation of alpha beta- and alpha gamma-dimers with increasing pH explains the well-known alkaline resistance of Hb F. Two dimers combine to form the functioning alpha 2 beta 2-tetramer. This model of hemoglobin assembly explains the different levels of positively charged and negatively charged mutant hemoglobins that are encountered in heterozygotes and the effect of alpha-thalassemia and heme deficiency states in modifying the level of the variant hemoglobin as well as Hb A2. Electrostatic interactions also affect the binding of hemoglobin to the cytoplasmic surface of the red cell membrane and may underlie the formation of target cells. Enhanced binding of positively charged variants such as S and C trigger a normally dormant pathway for potassium and water loss. Thus, the positive charge on beta c is responsible for the two major contributors to the pathogenesis of Hb SC disease: increased proportion of Hb S and increased intracellular hemoglobin concentration. It is likely that electrostatic interactions play an important role in the assembly of a number of other multisubunit macromolecules, including membrane receptors, cytoskeletal proteins, and DNA binding proteins.     

Extracranial tumor vascularity: determination by dynamic CT scanning. Part II: The unit approach

Twenty-eight patients had combined conventional drip infusion CT scans. The information about the anatomic location of the lesion, its configuration, its cross-sectional appearance, its vascularity (as determined by dynamic signature curves), and its clinical presentation were considered as a single overall unit. This diagnostic approach allowed a diagnosis to be made on virtually all of these enhancing lesions without resorting to either a digital venous imaging study or angiographic procedure. In 17 of these cases, such an invasive second procedure was performed either to confirm the CT impression as part of this study or as part of a therapeutic embolization procedure.     

FC04.6Characterisation of dendritic cell responses to ascaridol, an autoxidation product of tea tree oil

A large number of monoterpenes and their degradation products are likely skin sensitizing agents (Hausen et al., 1999). Terpenes are very common e.g., as constituents of cosmetics, food and other daily used products.
We investigated responses to the endoperoxide 1,4‐epidioxy‐2‐p‐menthen (ascaridol), an autoxidation product of tea tree oil, using monocyte derived dendritic cells (MDDC). Therefore, we isolated peripheral blood mononuclear cells (PBMC) from 9 healthy donors by the standard Ficoll‐Paque gradient centrifugation.
Monocytes were isolated by adherence and incubated in media (RPMI 1640) containing GM‐CSF (800 units/ml), IL‐4 (1000 units/ml) and 10% autologous serum. The immature MDDC (day 6) were characterized by flow cytometry (CD1a+, CD14−, CD40, CD45, CD80, CD83, CD86, HLA‐DR and CCR‐7) and incubated with various concentrations of ascaridol (1–70 μg/ml). After one hour incubation time LPS was added (1 μg/ml) for 23 h. Cell culture supernatants were collected after 24 h for cytokine analysis.
IL‐12p40, IL‐12p70 and prostaglandin E2 were measured by ELISA, TNF‐alpha and IL‐2 were measured by flow cytometry (FACS). Methods of the quantification of steady state mRNA levels were established for IL‐12 and CCR7 (real‐time RT‐PCR). Ascaridol enhanced significantly IL‐12p70 production (120% up to 396%) by MDDC as well as mRNA levels for IL‐12 and CCR7. Moreover, we detected a distinct increase of TNF‐alpha (110% up to 146%) secretion, IL‐2 (135%) and PGE2 (102% up to 155%).
Totally, these results suggest that ascaridol may be a potent modulator of maturation and antigen presenting function of dendritic cells, and we performing further experiments to verify this hypothesis.
This study was supported by the Deutsche Forschungsgemeinschaft.     

Tissue and lineage-specific variation in inactive X chromosome expression of the murine Smcx gene

To understand how gene expression patterns are established on the inactive X chromosome during development, we have studied the murine gene Smcx, which is expressed from both the active and inactive mouse X chromosomes. In all tissues assayed, Smcx only partially escapes X inactivation, with expression levels from the inactive X allele approximately 30-65% that of the active X allele. Additionally, inactive X expression levels differed between extraembryonic and embryonic tissues and among different tissues from newborn and adult mice. Imprinted extraembryonic tissue had the lowest levels of inactive X Smcx expression, whereas the highest levels were in heart. These data suggest that the chromosomal basis of X inactivation differs among tissues, perhaps reflecting differences in the timing or regulation of inactivation in these cell lineages.      

Relationship among serotype G3P5A rotavirus strains isolated from different host species

The relationship among G3P5A rotavirus strains was analysed by restriction endonuclease assay of the VP4, VP6 and VP7 encoding genes, neutralization assay and phylogenetic analysis. The restriction patterns of the capsid encoding genes were species specific allowing the differentiation among the strains of different origin. The VP7 profiles differentiated human from animal strains more efficiently. The phylogenetic analysis of the VP4 gene demonstrated that HCR3A and K9 are closer related to each other than to other P5A strains. The same occurs to strains Ro1845 and Cat 97. The CU-1 virus appears to be an ancestor of the P5A strains by neutralization and phylogenetic analysis. The results placed the RRV strain definitely in a separate VP4 serotype and genotype from that of P5A strains. Restriction endonuclease assay of the capsid encoding genes seems to be a useful tool to identify the host species of rotavirus strains belonging to the same serotype and/or genotype.     

The mouse Mid1 gene: implications for the pathogenesis of Opitz syndrome and the evolution of the mammalian pseudoautosomal region

We have recently reported isolation of the gene responsible for X- linked Opitz G/BBB syndrome, a defect of midline development. MID1 is located on the distal short arm of the human X chromosome (Xp22. 3) and encodes a novel member of the B box family of zinc finger proteins. We have now cloned the murine homolog of MID1 and performed preliminary expression studies during development. Mid1 expression in undifferentiated cells in the central nervous, gastrointestinal and urogenital systems suggests that abnormal cell proliferation may underlie the defect in midline development characteristic of Opitz syndrome. We have also found that Mid1 is located within the mouse pseudoautosomal region (PAR) in Mus musculus , while it seems to be X- specific in Mus spretus. Therefore, Mid1 is likely to be a recent acquisition of the M. musculus PAR. Genetic and FISH analyses also demonstrated a high frequency of unequal crossovers in the murine PAR, creating spontaneous deletion/duplication events involving Mid1. These data provide evidence for the first time that genetic instability of the PAR may affect functionally important genes. In addition, we show that MID1 is the first example of a gene subject to X-inactivation in man while escaping it in mouse. These data contribute to a better understanding of the molecular content and evolution of the rodent PAR.      

异落新妇甙的结构研究

从百合科菝葵属植物土茯苓(Smilaxg labra Roxb.)根茎的乙醇提取物中分得一个新的天然化合物(I),命名为异落新妇甙(isoastilbin)。根据元素分析,UV,IR,¹H-NMR,¹³C-NMR,2DNMR及FAB-MS,确定化合物I的结构为5,7,3',5'-四羟基二氢黄酮醇-3-O-α-L-鼠李糖甙。