Accession-Specific Haplotypes of the Internal Transcribed Spacer Region in Arabidopsis thaliana--A Means for Barcoding Populations

Eukaryote genomes contain multiple copies of nuclear ribosomal DNA (nrDNA) harboring both highly conserved and variable regions. This has made nrDNA the most popular genetic marker for phylogenetic studies and the region of choice for barcoding projects. Furthermore, many scientists believe that all copies of nrDNA within one nucleus are practically identical due to concerted evolution. Here, we investigate the model plant species Arabidopsis thaliana for intragenomic variation of the internal transcribed spacer (ITS) region of nrDNA. Based on a modified deep sequencing approach, we provide a comprehensive list of ITS polymorphisms present in the two most widely used accessions of A. thaliana-Col-0 and Ler. Interestingly, we found that some polymorphisms are shared between these genetically very distinct accessions. On the other hand, the high number of accession-specific polymorphisms shows that each accession can be clearly and easily characterized by its specific ITS polymorphism patterns and haplotypes. Network analysis based on the detected haplotypes demonstrates that the study of ITS polymorphism patterns and haplotypes is an extremely powerful tool for population genetics. Using the methods proposed here, it will now be possible to extend the traditionally species-bound barcoding concept to populations.     

Identification of evolutionarily conserved genetic regulators of cellular aging

To identify new genetic regulators of cellular aging and senescence, we performed genome-wide comparative RNA profiling with selected human cellular model systems, reflecting replicative senescence, stress-induced premature senescence, and distinct other forms of cellular aging. Gene expression profiles were measured, analyzed, and entered into a newly generated database referred to as the GiSAO database. Bioinformatic analysis revealed a set of new candidate genes, conserved across the majority of the cellular aging models, which were so far not associated with cellular aging, and highlighted several new pathways that potentially play a role in cellular aging. Several candidate genes obtained through this analysis have been confirmed by functional experiments, thereby validating the experimental approach. The effect of genetic deletion on chronological lifespan in yeast was assessed for 93 genes where (i) functional homologues were found in the yeast genome and (ii) the deletion strain was viable. We identified several genes whose deletion led to significant changes of chronological lifespan in yeast, featuring both lifespan shortening and lifespan extension. In conclusion, an unbiased screen across species uncovered several so far unrecognized molecular pathways for cellular aging that are conserved in evolution.     

A study of replacement of timolol-pilocarpine with latanoprost in pseudoexfoliation glaucoma

The aim of the study was to evaluate the efficacy of replacing current dual local therapy (timolol and pilocarpine) with latanoprost 0.005% in 71 pseudoexfoliation glaucoma patients with controlled intraocular pressure (IOP). 39 patients switched to latanoprost 0.005%) and 32 patients continued timolol-pilocarpine therapy. Mean diurnal (IOP) was measured at baseline, after 0.5, 1, 3 and 6 months of treatment. After 6 months 38 patients with latanoprost and 30 patients with timolol-pilocarpine had completed the study. At baseline the mean diurnal IOP was 20.4 +/- 2.0 mmHg for patients in latanoprost treatment group and 21.4 +/- 2.1 mmHg for patients in timolol-pilocarpine group. At the end of the study, after 6 months of treatment, the mean diurnal IOP values were 16.6 +/- 2.4 and 17.9 +/- 2.0 mmHg respectively. IOP was statistically significantly reduced from baseline (p < 0.001). The mean diurnal IOP change from baseline was -3.3 +/- 0.5 mmHg (mean +/- SEM, ANCOVA) for the patients treated with latanoprost and -3.2 +/- 0.4 mmHg for the patients treated with timolol + pilocarpine. This difference in IOP reduction between groups was not statistically significant (z = 0.69; p = 0.49). This study showed that combination therapy (timolol plus pilocarpine) in pseudoexfoliation glaucoma can effectively be replaced by latanoprost monotherapy.     

Dental and minor physical anomalies in children with developmental disorders--a discriminant analysis

A discriminant analysis was performed in a sample of 303 children with developmental disorders (DD) and 303 healthy controls (C) in order to test whether some oro-dental and physical minor anomalies could discriminate these groups of children. DD sample comprised 176 mentally retarded (MR) children. 70 children with impaired hearing (IH) and 57 children with impaired vision (IV). The control group included 303 healthy subjects, matched for sex and age. The analysis comprised seven common oral and dental anomalies: median diastema, hypodontia, impacted teeth, microdontia, dens invaginatus, upper lip frenulum and frenulum of the tongue. Minor physical anomalies were assessed by the method proposed by Waldrop et al., as the average number of minor anomalies per individual (W1) and as the weighted score of minor anomalies (W2). Three discriminant functions were obtained by analysis of nine initial variables. Distinct discrimination and considerable distances were found between the centroids of the controls and all groups of DD children. The first two discriminant functions were significant for discrimination between the groups and they explained 98.6% of the total variance. The first function contained 90.2% of information and was defined by the number and weighted scores of minor anomalies. The second variable explained 8.4% of the total variability and was defined by three dental anomalies. The results obtained by the discriminant analysis show that application of dental and minor physical anomalies enables discrimination between the group of healthy children and the groups of children with different developmental disorders.     

Activating immunity in the liver. II. IFN-beta attenuates NK cell-dependent liver injury triggered by liver NKT cell activation

Dendritic cell (DC)-dependent activation of liver NKT cells triggered by a single i.v. injection of a low dose (10-100 ng/mouse) of alpha-galactosyl ceramide (alphaGalCer) into mice induces liver injury. This response is particularly evident in HBs-tg B6 mice that express a transgene-encoded hepatitis B surface Ag in the liver. Liver injury following alphaGalCer injection is suppressed in mice depleted of NK cells, indicating that NK cells play a role in NK T cell-initiated liver injury. In vitro, liver NKT cells provide a CD80/86-dependent signal to alphaGalCer-pulsed liver DC to release IL-12 p70 that stimulates the IFN-gamma response of NKT and NK cells. Adoptive transfer of NKT cell-activated liver DC into the liver of nontreated, normal (immunocompetent), or immunodeficient (RAG(-/-) or HBs-tg/RAG(-/-)) hosts via the portal vein elicited IFN-gamma responses of liver NK cells in situ. IFN-beta down-regulates the pathogenic IL-12/IFN-gamma cytokine cascade triggered by NKT cell/DC/NK cell interactions in the liver. Pretreating liver DC in vitro with IFN-beta suppressed their IL-12 (but not IL-10) release in response to CD40 ligation or specific (alphaGalCer-dependent) interaction with liver NKT cells and down-regulated the IFN-gamma response of the specifically activated liver NKT cells. In vivo, IFN-beta attenuated the NKT cell-triggered induction of liver immunopathology. This study identifies interacting subsets of the hepatic innate immune system (and cytokines that up- and down-regulate these interactions) activated early in immune-mediated liver pathology.     

The effect of temperature on midgut and brain protein profiles in Morimus funereus larvae (Coleoptera: Cerambycidae)

The 7-days shift of M. funereus larvae, from nature to a constant temperature of 23 degrees C led to changes in midgut and brain protein quality and quantity. The changes in midgut protein profiles are characterized by an intensified protein band Mr of 29 kD, the absence of protein Mr of 22 kD and less intense bands Mr of 8.5-2.5 kD. Electrophoretic patterns of brain proteins showed less intense Mr of 66-2.5 kD protein bands.     

Mortality of cerebrovascular diseases in Croatia--1958-1997

The aim of this investigation was to analyze secular trend of mortality from cerebrovascular diseases in Croatia and its regional characteristics. The research comprised all deaths from cerebrovascular diseases in Croatia in persons aged between 35 and 74 years over the period 1958-1997. The investigated period is divided in eight 5-year periods, and for that 5-year periods proportional mortality rates, standardized mortality rates and specific mortality rates, according to the age and gender were calculated. Number of all deaths in the population aged 35-74 in Croatia, by 5-year periods rose from 18,913 to 26,788 (increase of 42%), deaths from cerebrovascular diseases from 2831 to 3959 (increase of 40%). Proportional mortality rate for this disease increased from 9.0% in the first 5-year period to 14.8% in the last 5-year period (increase of 64%). Standardized mortality rates for cerebrovascular diseases increased from 118 to 206 per 100,000 inhabitants (increase of 75%). The specific mortality rates over a 5-year period have shown a trend of increase in all men age groups and stagnation or decrease in women age groups. At the same time the rates standardized by age and sex increased by 62%. Standardized mortality rates for cerebrovascular diseases in continental communities (Osijek, Varazdin) are much higher (twice or even threefold) than those in coastal communities (Split, Rijeka). A data analysis showed that, although mortality trends of cerebrovascular diseases stagnated or even declined in some communities during the recent years, the secular trend for the entire country had a tendency of constant rise over the whole period of research. Therefore, the short-term prognosis predicts further increase of both the number and rates of deaths from cerebrovascular diseases in our country.     

Muscle contracture emulating system for studying artificially induced pathological gait in intact individuals

When studying pathological gait it is important to correctly identify primary gait anomalies originating from damage to the central nervous and musculoskeletal system and separate them from compensatory changes of gait pattern, which is often challenging due to the lack of knowledge related to biomechanics of pathological gait. A mechanical system consisting of specially designed trousers, special shoe arrangement, and elastic ropes attached to selected locations on the trousers and shoes is proposed to allow emulation of muscle contractures of soleus (SOL) and gastrocnemius (GAS) muscles and both SOL-GAS. The main objective of this study was to evaluate and compare gait variability as recorded in normal gait and when being constrained with the proposed system. Six neurologically and orthopedically intact volunteers walked along a 7-m walkway while gait kinematics and kinetics were recorded using VICON motion analysis system and two AMTI forceplates. Statistical analysis of coefficient of variation of kinematics and kinetics as recorded in normal walking and during the most constrained SOL-GAS condition showed comparable gait variability. Inspection of resulting group averaged gait patterns revealed considerable resemblance to a selected clinical example of spastic diplegia, indicating that the proposed mechanical system potentially represents a novel method for studying emulated pathological gait arising from artificially induced muscle contractures in neurologically intact individuals.