The expression of human glycerol-3-phosphate dehydrogenase in human/rodent somatic-cell hybrids

Our previous studies using rodent/human somatic-cell hybrids suggested that the expression of human mitochondrial glycerol-3-phosphate dehydrogenase (GPDM) is dependent on the presence of human mitochondria. This has now been tested directly by analysis of GPDM activity in a series of nine hybrid-cell lines, four segregating human chromosomes and five losing rodent chromosomes (reverse segregants). The chromosome composition of the hybrids was deduced from analysis of biochemical markers and examination of G- and G11-banded metaphase spreads and the mitochondrial content was determined by Southern blot analysis, using cloned mouse and human mtDNA sequences as probes. We found that the mtDNA species present in these hybrids correlated exactly with the pattern of chromosome segregation such that the conventional hybrids contained rodent mtDNA and the reverse segregants human mtDNA. However, the pattern of GPDM expression was not directly correlated with the species of chromosomes or mitochondria present: all the hybrids showed strong rodem GPDM activity and two from each class of hybrid also showed human GPDM activity but the other hybrids were negative for human GPDM. We conclude that rodent GPDM readily integrates into human mitochondria, that the expression of rodent GPDM is not dependent on the presence of rodent mitochondria, and that GPDM is not coded by mtDNA. Human GPDM either is not capable of being inserted into the rodent mitochondrial membrane or is regulated in some way in the hybrid cells by an unidentified rodent factor.     

Behavioral responses of three species of ground beetle (Coleoptera: Carabidae) to olfactory cues associated with prey and habitat

The responses ofPterostichus melanarius Illiger,Harpalus rufipes DeGeer, andNebria brevicollis Fabricius (Coleoptera: Carabidae) to olfactory cues of prey and habitat were studied in a four-arm continuous-airflow olfactometer. The process was semiautomated using time-lapse image analysis by microcomputer. The primary constituent of the aphid alarm pheromone, (E)--farnesene (EBF), was synthesized and tested for a kairomonal role in prey detection by carabids. In addition, individual beetles were exposed to odors from live aphids, live collembolans, and a crude extract of wheat. All three beetle species showed evidence of olfaction.P. melanarius responded to all the odors except collembolans,H. rufipes responded to EBF and wheat, andN. brevicollis to collembolans. The use of a defence allomone as a prey-finding kairomone by certain carabid species has implications for pest management. Manipulation of predator chemical ecology by the inclusion of behavior-modifying compounds in a crop spray mix with reduced amounts of insecticide may allow for efficient aphid control with less environmental contamination.     

Type VI collagen microfibrils: evidence for a structural association with hyaluronan

Type VI collagen, a widespread structural component of connective tissues, has been isolated in abundance from fetal bovine skin by a procedure involving bacterial collagenase digestion under nonreducing, nondenaturing conditions and gel filtration chromatography. Rotary shadowing electron microscopic analysis revealed that the collagen VI was predominantly in the form of extensive intact microfibrillar arrays. These microfibrils were seen in association with hyaluronan, which was identified by its ability to bind the G1 fragment of cartilage proteoglycan. Treatment with highly purified hyaluronidase largely disrupted the collagen VI microfibrils into component tetramers, double tetramers, and short microfibrillar sections. Subsequent incubation of disrupted collagen VI in the presence of hyaluronan facilitated a partial repolymerization of the microfibrils. In vitro binding studies have also demonstrated that type VI collagen binds hyaluronan with a relatively high affinity. These studies demonstrate that a specific structural relationship exists between type VI collagen and hyaluronan. This association is likely to be of primary importance in the growth and remodeling processes of connective tissues.     

Symptoms and Clinical Course of EHEC O104 Infection in Hospitalized Patients: A Prospective Single Center Study

Objectives

Shiga-toxin producing O157:H7 Entero Haemorrhagic E. coli (STEC/EHEC) is one of the most common causes of Haemolytic Uraemic Syndrome (HUS) related to infectious haemorrhagic colitis. Nearly all recommendations on clinical management of EHEC infections refer to this strain. The 2011 outbreak in Northern Europe was the first to be caused by the serotype O104:H4. This EHEC strain was found to carry genetic features of Entero Aggregative E. coli (EAEC) and extended spectrum β lactamase (ESBL). We report symptoms and complications in patients at one of the most affected centres of the 2011 EHEC O104 outbreak in Northern Germany.

Methods

The courses of patients admitted to our hospital due to bloody diarrhoea with suspected EHEC O104 infection were recorded prospectively. These data include the patients’ histories, clinical findings, and complications.

Results

EHEC O104 infection was confirmed in 61 patients (female = 37; mean age: 44±2 years). The frequency of HUS was 59% (36/61) in our cohort. An enteric colonisation with co-pathogens was found in 57%. Thirty-one (51%) patients were treated with plasma-separation/plasmapheresis, 16 (26%) with haemodialysis, and 7 (11%) with Eculizumab. Patients receiving antibiotic treatment (n = 37; 61%) experienced no apparent change in their clinical course. Twenty-six (43%) patients suffered from neurological symptoms. One 83-year-old patient died due to comorbidities after HUS was successfully treated.

Conclusions

EHEC O104:H4 infections differ markedly from earlier reports on O157:H7 induced enterocolitis in regard to epidemiology, symptomatology, and frequency of complications. We recommend a standard of practice for clinical monitoring and support the renaming of EHEC O104:H4 syndrome as “EAHEC disease”.     

Erythropoietin stimulates the coronary collateral development in patients with coronary chronic total occlusion

Objective

Erythropoietin (EPO) improves cardiac function and induces neovascularisation in post-myocardial infarction heart failure. The aim of this study was to analyse the association between the serum erythropoietin level and coronary collateral development in patients with coronary artery disease and chronic total occlusion.

Methods

A total of 168 patients consisting of 117 with coronary artery disease (CAD, (62 with chronic total occlusion (CTO), 55 without CTO)) and 51 with healthy coronary arteries were included in the study. The patients were assigned as coronary artery disease without CTO (group 0), CAD with CTO (group 1: poor collateral development, group 2: good collateral development) and normal coronary arteries (group 3).

Results

There was a significant positive correlation between serum EPO levels and the Rentrop scores in angiography (r = 0.243, p = 0.001). Similarly, a positive correlation was found between serum EPO levels and the Syntax scores (r = 0.253, p = 0.001). Echocardiography revealed a negative correlation between serum EPO levels and the cardiac ejection fraction (r = ?0.210, p = 0.006).

Conclusions

Serum EPO is a useful biomarker for coronary collateral development in patients with CTO.

     

Structural basis of type VI collagen dimer formation

We have determined the interactive sites required for dimer formation in type VI collagen. Despite the fact that type VI collagen is a heterotrimer composed of alpha1(VI), alpha2(VI), and alpha3(VI) chains, the formation of dimers is determined principally by interactions of the alpha2(VI) chain. Key components of this interaction are the metal ion-dependent adhesion site (MIDAS) motif of the alpha2C2 A-domain and the GER sequence in the helical domain of another alpha2(VI) chain. Replacement of the alpha2(VI) C2 domain with the alpha3(VI) domain abolished dimer formation, whereas alterations in the alpha2(VI) C1 domain did not disrupt dimer formation. When the helical sequences were investigated, replacement of the alpha2(VI) sequence GSPGERGDQ with the alpha3(VI) sequence GEKGERGDV abolished dimer formation. Mutating the Pro-108 to a Lys-108 in this alpha2(VI) sequence did not influence dimer formation and suggests that, unlike the integrin I-domain/triple-helix interaction, hydroxyproline is not required in collagen VI A-domain/helix interaction. These results demonstrate that the alpha2(VI) chain position in the assembled triple-helical molecule is critical for antiparallel dimer formation and identify the interacting collagenous and MIDAS sequences involved. These interactions underpin the subsequent assembly of type VI collagen.     

Fibrillin: from microfibril assembly to biomechanical function

Fibrillins form the structural framework of a unique and essential class of extracellular microfibrils that endow dynamic connective tissues with long-range elasticity. Their biological importance is emphasized by the linkage of fibrillin mutations to Marfan syndrome and related connective tissue disorders, which are associated with severe cardiovascular, ocular and skeletal defects. These microfibrils have a complex ultrastructure and it has proved a major challenge both to define their structural organization and to relate it to their biological function. However, new approaches have at last begun to reveal important insights into their molecular assembly, structural organization and biomechanical properties. This paper describes the current understanding of the molecular assembly of fibrillin molecules, the alignment of fibrillin molecules within microfibrils and the unique elastomeric properties of microfibrils.     

Two active-site tyrosyl residues of protein TrwC act sequentially at the origin of transfer during plasmid R388 conjugation

Protein TrwC is the relaxase-helicase responsible for the initiation and termination reactions of DNA processing during plasmid R388 conjugation. Site-directed mutagenesis was used to change to phenylalanine each of a set of four conserved tyrosyl residues in the sequence of the N-terminal relaxation domain of the protein. Simultaneous mutation of both Y18 and Y26 was required to abolish in vitro cleavage and strand-transfer reactions catalyzed by protein TrwC on oligonucleotides containing the nic site. Thus, both Y18 and Y26 could be involved independently in the formation of oligonucleotide-protein covalent complexes that constitute presumed intermediates of these reactions. This hypothesis was confirmed by the observation of Y18 and Y26-specific peptide-oligonucleotide adducts after protease digestion of TrwC and mutant derivatives. Finally mutation Y18F, but not mutation Y26F, abolished nic-cleavage of a supercoiled DNA containing the R388 origin of transfer (oriT). These data allowed the construction of a model for conjugative DNA processing in which Y18 specifically catalyzes the initial cleavage reaction, while Y26 is used for the second strand-transfer reaction, which terminates conjugation. The model suggests a control mechanism that can be effective at each conjugative replication cycle.     

Effect of EDTA and Tannic Acid on the Removal of Cd,Ni, Pb and Cu from Artificially Contaminated Soil by Althaea rosea Cavan

In this study an ornamental plant of Althaea rosea Cavan was investigated for its potential use in the removal of Cd, Ni, Pb and Cu from an artificially contaminated soil. Effect of two different chelating agents on the removal has also been studied by using EDTA (ethylenediaminetetracetic acid) and TA (tannic acid). Both EDTA and TA have led to higher heavy metal concentration in shoots and leaves compared to control plants. However EDTA is generally known as an effective agent in metal solubilisation of soil, in this study, TA was found more effective to induce metal accumulation in Althaea rosea Cavan under the studied conditions. In addition to this, EDTA is toxic to some species and restraining the growth of the plants. The higher BCF (Bio Concentration Factor) and TF (Translocation Factor) values obtained from stems and leaves by the effects of the chemical enhancers (EDTA and TA) show that Althaea rosea Cavan is a hyper accumulator for the studied metals and may be cultivated to clean the contaminated soils.