Purine metabolism in Toxoplasma gondii

We have studied the incorporation and interconversion of purines into nucleotides by freshly isolated Toxoplasma gondii. They did not synthesize nucleotides from formate, glycine, or serine. The purine bases hypoxanthine, xanthine, guanine, and adenine were incorporated at 9.2, 6.2, 5.1, and 4.3 pmol/10(7) cells/h, respectively. The purine nucleosides adenosine, inosine, guanosine, and xanthosine were incorporated at 110, 9.0, 2.7, and 0.3 pmol/10(7) cells/h, respectively. Guanine, xanthine, and their respective nucleosides labeled only guanine nucleotides. Inosine, hypoxanthine, and adenine labeled both adenine and guanine nucleotide pools at nearly equal ratios. Adenosine kinase was greater than 10-fold more active than the next most active enzyme in vitro. This is consistent with the metabolic data in vivo. No other nucleoside kinase or phosphotransferase activities were found. Phosphorylase activities were detected for guanosine and inosine; no other cleavage activities were detected. Deaminases were found for adenine and guanine. Phosphoribosyltransferase activities were detected for all four purine nucleobases. Interconversion occurs only in the direction of adenine to guanine nucleotides.     

Deletion mutagenesis of Tn10 Tet repressor — localization of regions important for dimerization and inducibility in vivo

The gene for the Tn 10 Tet repressor (TetR) was subjected to deletion mutagenesis. Screening for a transdominant operator-binding negative phenotype yielded 10 mutants with internal deletions. Three deletions extend from residue D5 to residues L41, W75, or Q76, respectively, and two contain deletions of the α-helix-turn-α-helix DNA-binding motif. Five deletions range from residue K84 to residues between R87 and K98. Since residues from the N-terminus up to position 98 are not necessary for dimerization, this must take place in the C-terminal half of the protein. Ability to dimerize was probed by introducing ochre non-sense codons (oc) at residues G138, H151, E159, l174, or K202. Koc202 shows wild-type in vivo operator-binding and inducibility by tetracycline indicating that the six C-terminal residues of TetR are not important for activity. Mutants with longer C-terminal truncations are inactive and not transdominant. They show reduced steady-state protein levels and are probably impaired in folding and degraded in vivo . Two mutants (Δ151–166, Δ164–166) with deletions in a region variable in primary structure and length among Tet repressers from different resistance determinants bind tet operator efficiently, but are not inducibie by tetracycline. This result indicates that these residues are not important for dimer formation in the operator-binding form.     

Phosphofructokinase of Leishmania donovani and Leishmania braziliensis and its role in glycolysis.

It was shown in an investigation of the phosphofructokinases of Leishmania donovani and Leishmania braziliensis that both enzymes are similar to that of Crithidia fasciculata. Although the enzymes are allosteric with respect to their substrates and require AMP for activation, there is no influence by other heterotropic modifiers. The Mg2+-ATP chelate activates these enzymes in a first order process and they can be inhibited by free ATP. The inhibition is reversed by the activator, AMP, in a competitive manner. The requirement for the nucleotide in L. donovani can be eliminated by decreasing the pH. The data indicate that phosphofructokinase, a pivotal enzyme in glycolysis for most organisms, probably does not play an important role in glycolysis in Leishmania.     

Breast MRI in nonpalpable breast lesions: a randomized trial with diagnostic and therapeutic outcome – MONET – study

Background

In orthodontic treatment, anchorage control is a fundamental aspect. Usually conventional mechanism for orthodontic anchorage control can be either extraoral or intraoral that is headgear or intermaxillary elastics. Their use are combined with various side effects such as tipping of occlusal plane or undesirable movements of teeth. Especially in cases, where key-teeth are missing, conventional anchorage defined as tooth-borne anchorage will meet limitations. Therefore, the use of endosseous implants for anchorage purposes are increasingly used to achieve positional stability and maximum anchorage.

Methods/Design

The intended study is designed as a prospective, multicenter randomized controlled trial (RCT), comparing and contrasting the effect of early loading of palatal implant therapy versus implant loading after 12 weeks post implantation using the new ortho-implant type II anchor system device (Orthosystem Straumann, Basel, Switzerland). 124 participants, mainly adult males or females, whose diagnoses require temporary stationary implant-based anchorage treatment will be randomized 1:1 to one of two treatment groups: group 1 will receive a loading of implant standard therapy after a healing period of 12 week (gold standard), whereas group 2 will receive an early loading of orthodontic implants within 1 week after implant insertion. Participants will be at least followed for 12 months after implant placement. The primary endpoint is to investigate the behavior of early loaded palatal implants in order to find out if shorter healing periods might be justified to accelerate active orthodontic treatment. Secondary outcomes will focus e.g. on achievement of orthodontic treatment goals and quantity of direct implant-bone interface of removed bone specimens. As tertiary objective, a histologic and microtomography evaluation of all retrieved implants will be performed to obtain data on the performance of the SLA surface in human bone evaluation of all retrieved implants. Additionally, resonance frequency analysis (RFA, Osstell? mentor) will be used at different times for clinically monitoring the implant stability and for histological comparison in order to measure the reliability of the resonance frequency measuring device.

Trial registration

Current Controlled Trials ISRCTN97142521.     

Single-chain Tet transregulators

We demonstrate here that the Tet repressor (TetR), a dimeric allosterical regulatory protein, can be converted to a fully functional monomer when connected by a 29 amino acid linker. TetR-based transregulators are widely used to regulate gene expression in eukaryotes. They can be fused to form single-chain (sc) Tet transregulators with two TetR moieties and one eukaryotic regulatory domain. Sc variants of transactivator and transsilencer exhibit the same regulatory properties as their respective dimeric counterparts in human cell lines. In particular, the reverse ‘tet-on’ phenotype of rtTA variants is also present in the sc variants. Coexpression of a reverse transactivator and sc transsilencer leads to reduced background expression and shows full activation upon induction. The data demonstrate that sc Tet transregulators exhibit the phenotype of their respective dimers and lack functional interference when coexpressed in the same cell.     

Successful immunotherapy with matrix metalloproteinase-derived peptides in adjuvant arthritis depends on the timing of peptide administration

We have recently found that matrix metalloproteinases (MMPs) are targets for T-cell and B-cell reactivity in experimental arthritis. In the present article, we investigate whether modulation of MMP-specific T-cell responses could influence the course of adjuvant arthritis (AA). Lewis rats were treated nasally with MMP peptides prior to or after AA induction. Administration of the MMP-10 or the MMP-16 peptide prior to AA induction reduced the arthritic symptoms. In contrast, administration of the MMP-10 peptide after AA induction aggravated the arthritic symptoms. The present study shows the possible usefulness of MMP peptides for immunotherapy. However, a clear understanding of proper timing of peptide administration is crucial for the development of such therapies.     

Differential role of proline-rich tyrosine kinase 2 and focal adhesion kinase in determining glioblastoma migration and proliferation

The propensity of malignant gliomas to invade surrounding brain tissue contributes to poor clinical outcome. Integrin-mediated adhesion to extracellular matrix regulates the migration and proliferation of many cell types, but its role in glioma progression is undefined. We investigated the role of the cytoplasmic tyrosine kinases FAK and Pyk2, potential integrin effectors, in the phenotypic determination of four different human glioblastoma cell lines. While FAK expression was similar between the four cell lines, increased FAK activity correlated with high proliferation and low migratory rates. In contrast, Pyk2 activity was significantly increased in migratory cell lines and depressed in proliferative cell lines. Overexpression of Pyk2 stimulated migration, whereas FAK overexpression inhibited cell migration and stimulated cellular proliferation. These data suggest that FAK and Pyk2 function as important signaling effectors in gliomas and indicate that their differential regulation may be determining factors in the temporal development of proliferative or migrational phenotypes.