Proteolytic hydrolysis and purification of the LRP/alfa-2-macroglobulin receptor domain from alpha-macroglobulins

A new, easier and efficient purification method, using Sephacryl and DEAE-Sephacel, of the C-terminal fragment of two alpha-macroglobulins, alpha(2)-M and PZP, is presented. Two larger peptides were identified for each protein as the C-terminal fragment, with molecular weights of approximately 30 kDa and the N-terminal sequences were determined to be SSTQDTV for alpha(2)-M and VALHLS for PZP. The smaller peptides with molecular weights of 18 kDa correspond to a shorter C-terminal sequence of these proteins, and they were determined to be EEFPFA for alpha(2)-M and ALKVQTV for PZP, with no interfering sequences detected. The results confirmed the discriminatory capacity of the purification procedure and the purity of the fragments. This new methodology facilitates biological studies of alpha-macroglobulins, and will enable elucidation of the role the C-terminal region may exert to eliminate alpha-macroglobulin-proteinases complexes from the circulation by the LRP/receptor.     

The Genetic Analysis of Achiasmate Segregation in Drosophila Melanogaster. III. the Wild-Type Product of the Axs Gene Is Required for the Meiotic Segregation of Achiasmate Homologs

The regular segregation of achiasmate chromosomes in Drosophila melanogaster females is ensured by two distinct segregational systems. The segregation of achiasmate homologs is assured by the maintenance of heterochromatic pairing; while the segregation of heterologous chromosomes is ensured by a separate mechanism that may not require physical association. Axs(D) (Aberrant X segregation) is a dominant mutation that specifically impairs the segregation of achiasmate homologs; heterologous achiasmate segregations are not affected. As a result, achiasmate homologs frequently participate in heterologous segregations at meiosis I. We report the isolation of two intragenic revertants of the Axs(D) mutation (Axs(r2) and Axs(r3)) that exhibit a recessive meiotic phenotype identical to that observed in Axs(D)/Axs(D) females. A third revertant (Axs(r1)) exhibits no meiotic phenotype as a homozygote, but a meiotic defect is observed in Axs(r1)/Axs(r2) females. Therefore mutations at the Axs(D) locus define a gene necessary and specific for homologous achiasmate segregation during meiosis. We also characterize the interactions of mutations at the Axs locus with two other meiotic mutations (ald and ncd). Finally, we propose a model in which Axs(+) is required for the normal separation of paired achiasmate homologs. In the absence of Axs(+) function, the homologs are often unable to separate from each other and behave as a single segregational unit that is free to segregate from heterologous chromosomes.     

A Short Chromosomal Region with Major Roles in Yeast Chromosome III Meiotic Disjunction, Recombination and Double Strand Breaks

A multicopy plasmid was isolated from a yeast genomic library, whose presence resulted in a twofold increase in meiotic nondisjunction of chromosome III. The plasmid contains a 7.5-kb insert from the middle of the right arm of chromosome III, including the gene THR4. Using chromosomal fragments derived from chromosome III, we determined that the cloned region caused a significant, specific, cis-acting increase in chromosome III nondisjunction in the first meiotic division. The plasmid containing this segment exhibited high spontaneous meiotic integration into chromosome III (in 2.4% of the normal meiotic divisions) and a sixfold increase (15.5%) in integration in nondisjunctant meioses. Genetic analysis of the cloned region revealed that it contains a ``hot spot' for meiotic recombination. In DNA of rad50S mutant cells, a strong meiosis-induced double strand break (DSB) signal was detected in this region. We discuss the possible relationships between meiosis-induced DSBs, recombination and chromosome disjunction, and propose that recombinational hot spots may be ``pairing sites' for homologous chromosomes in meiosis.     

Multiple sites for double-strand breaks in whole meiotic chromosomes of Saccharomyces cerevisiae.

We present a scheme for locating double-strand breaks (DSBs) in meiotic chromosomes of Saccharomyces cerevisiae, based on the separation of large DNA molecules by pulsed field gel electrophoresis. Using a rad50S mutant, in which DSBs are not processed, we show that DSBs are widely induced in S. cerevisiae chromosomes during meiosis. Some of the DSBs accumulate at certain preferred sites. We present general profiles of DSBs in chromosomes III, V, VI and VII. A map of the 12 preferred sites on chromosome III is presented. At least some of these sites correlate with known 'hot spots' for meiotic recombination. The data are discussed in view of current models of meiotic recombination and chromosome segregation.     

Ethnic groups and high sensitivity C-reactive protein in Israel

High-sensitivity C-reactive protein (hs-CRP) is a biomarker that correlates with atherothrombotic risk and outcome. hs-CRP is influenced by various modifiable and non-modifiable factors. We studied the relationship between ethnic background and hs-CRP level, among the Jewish population in Israel. A total of 3659 men and 2180 women were divided into two ethnic groups (Ashkenazi and Sephardic Jews), based on the knowledge of Jewish immigration patterns throughout the centuries. Mean hs-CRP levels were calculated for each group and were adjusted for various factors known to influence hs-CRP. Sephardic Jews were found to have higher adjusted mean hs-CRP levels (2.0 mg l^-1 for men and 3.9 mg l^-1 for women) compared with Ashkenazi Jews (1.5 mg l^-1 for men and 2.9 mg l^-1 for women). Ethnic background emerged as an independent significant predictor of hs-CRP levels. We demonstrated that ethnicity is an important factor when considering hs-CRP as a marker of atherothrombotic risk.     

Skin Donors Positive for HBV Core Antibodies – to Bank or Not to Bank? An Overview from a Regional Tissue Bank

A prompt transplantation of skin allografts on patients with severe, large body area burns is a preferred treatment, but depends on a suitable supply of tissue donors. Limiting factors include donors' identification, families consent, and following the standards – exclusion due to assessed transmissible diseases. To increase the current rate of skin donations to our regional skin bank, we reviewed the data of all potential organ donors, identified at Soroka University Medical Center from October 1997 to December 2000 and evaluated the causes for exclusion, especially due to HBV serological profile. 114/168 (67.9%) patients did not meet the indicated standards for organ donation, among which 20/114 patients (17.5%) positive for anti-HBc (anti-HBc⁺). 54/168 persons were declared brain dead, with consents obtained from 21 families. To discuss the intriguing approval of skin from potential donors with anti-HBc⁺ serology, the literature was reviewed, specifically – the reported outcomes of organ transplants from anti-HBc⁺ donors, updates of HBV and skin, available tests, and finally a look for a safe commendable algorithm. The results suggested that HBV might be replicating in the skin, but proven communication of HBV has not been reported following grafting skin from anti-HBc⁺ donors. Unlike other procured organs and tissues, grafted banked skin is a temporary cover, storable up to six years, under appropriate conditions. Hence, banking of skin from anti-HBc⁺ donors might be considered for future grafting of patients with identical serological profiles, presumably immune to a subsequent HBV infection, until a further re-evaluation of the standards. This procedure is anticipated to increase the potential of organ and tissue donations, specifically skin.     

Decline in Serum Cholinesterase Activities Predicts 2-Year Major Adverse Cardiac Events

Parasympathetic activity influences long-term outcome in patients with cardiovascular disease, but the underlying mechanism(s) linking parasympathetic activity and the occurrence of major adverse cardiovascular events (MACEs) are incompletely understood. The aim of this pilot study was to evaluate the association between serum cholinesterase activities as parasympathetic biomarkers and the risk for the occurrence of MACEs. Cholinergic status was determined by measuring the cumulative capacity of serum acetylcholinesterase (AChE) and butyrylcholinesterase (BChE) to hydrolyze the AChE substrate acetylthiocholine. Cholinergic status was evaluated in randomly selected patients undergoing cardiac catheterization. The patients were divided into two groups of 100 patients in each group, with or without occurrence of MACEs during a follow-up period of 40 months. Cox regression models adjusted for potential clinical, metabolic and inflammatory confounders served to evaluate association with clinical outcome. We found that patients with MACE presented lower cholinergic status and AChE values at catheterization (1,127 ± 422 and 359 ± 153 nmol substrate hydrolyzed per minute per milliliter, respectively) than no-MACE patients (1,760 ± 546 and 508 ± 183 nmol substrate hydrolyzed per minute per milliliter, p < 0.001 and p < 0.001, respectively), whose levels were comparable to those of matched healthy controls (1,622 ± 303 and 504 ± 126 nmol substrate hydrolyzed per minute per milliliter, respectively). In a multivariate analysis, patients with AChE or total cholinergic status values below median showed conspicuously elevated risk for MACE (hazard ratio 1.85 [95% confidence interval [CI] 1.09–3.15, p = 0.02] and 2.21 [95% CI 1.22–4.00, p = 0.009]) compared with those above median, even after adjusting for potential confounders. We conclude that parasympathetic dysfunction expressed as reduced serum AChE and AChE activities in patients compared to healthy controls can together reflect impaired parasympathetic activity. This impairment predicts the risk of MACE up to 40 months in such patients. Monitoring these parasympathetic parameters might help in the risk stratification of patients with cardiovascular disease.     

Effects of a novel medial meniscus implant on the knee compartments: imaging and biomechanical aspects

Biomechanics and Modeling in Mechanobiology - The altered biomechanical function of the knee following partial meniscectomy results in ongoing articular cartilage overload, which may lead to...