Biolistic nuclear transformation of Saccharomyces cerevisiae and other fungi

Summary The yeast Saccharomyces cerevisiae has been engineered to synthesize and secrete desulfato-hirudin (hirudin), a thrombin inhibitor from the leech Hirudo medicinalis. The synthetic gene coding for hirudin was expressed constitutively under the control of four size-variants of the yeast glyceraldehyde-3-phosphate dehydrogenase promoter (GAP) and cloned into a 2 based multicopy yeast vector. The constitutive action of the four promoter variants was confirmed by demonstrating that the expression and secretion of hirudin is growth-related. The different efficiencies of the promoter variants not only affected hirudin expression but also led to changes in several cellular parameters, such as cell growth, average plasmid copy number and plasmid stability. The observed changes show that yeast cells establish a specific equilibrium for each promoter variant. We conclude, that the adjustment of cellular parameters in response to the expression levels of a heterologous protein is regulated by two counteracting selective forces: (1) the need for complementation of the auxotrophic host marker by the plasmid-encoded selection gene which, in the case of dLEU2, requires several plasmid copies; and (2) a selective advantage of cells with a lower copy number enabling them to escape the burden of heterologous protein production.     

A genome-wide search for susceptibility genes in human systemic lupus erythematosus sib-pair families

Systemic lupus erythematosus (SLE) is an autoimmune multisystem inflammatory disease characterized by the production of pathogenic autoantibodies. Previous genetic studies have suggested associations with HLA Class II alleles, complement gene deficiencies, and Fc receptor polymorphisms; however, it is likely that other genes contribute to SLE susceptibility and pathogenesis. Here, we report the results of a genome-wide microsatellite marker screen in 105 SLE sib-pair families. By using multipoint nonparametric methods, the strongest evidence for linkage was found near the HLA locus (6p11-p21) [D6S257, logarithm of odds (lod) = 3.90, P = 0.000011] and at three additional regions: 16q13 (D16S415, lod = 3.64, P = 0.000022), 14q21–23 (D14S276, lod = 2.81, P = 0.00016), and 20p12 (D20S186, lod = 2.62, P = 0.00025). Another nine regions (1p36, 1p13, 1q42, 2p15, 2q21–33, 3cent-q11, 4q28, 11p15, and 15q26) were identified with lod scores ≥1.00. These data support the hypothesis that multiple genes, including one in the HLA region, influence susceptibility to human SLE.     

A computationally efficient method for automatic registration of orthogonal x-ray images with volumetric CT data

The paper presents a computationally efficient 3D-2D image registration algorithm for automatic pre-treatment validation in radiotherapy. The novel aspects of the algorithm include (a) a hybrid cost function based on partial digitally reconstructed radiographs (DRRs) generated along projected anatomical contours and a level set term for similarity measurement; and (b) a fast search method based on parabola fitting and sensitivity-based search order. Using CT and orthogonal x-ray images from a skull and a pelvis phantom, the proposed algorithm is compared with the conventional ray-casting full DRR based registration method. Not only is the algorithm shown to be computationally more efficient with registration time being reduced by a factor of 8, but also the algorithm is shown to offer 50% higher capture range allowing the initial patient displacement up to 15 mm (measured by mean target registration error). For the simulated data, high registration accuracy with average errors of 0.53 mm +/- 0.12 mm for translation and 0.61 +/- 0.29 degrees for rotation within the capture range has been achieved. For the tested phantom data, the algorithm has also shown to be robust without being affected by artificial markers in the image.     

Knee acoustic emission: a potential biomarker for quantitative assessment of joint ageing and degeneration

Based on a single time-point study of 34 healthy and 19 osteoarthritic knees in three different age groups (early, middle and late adulthood), this paper reports the potential of knee acoustic emission as a biomarker to monitor joint ageing and degeneration. Measurements were made of short transient high frequency acoustic emission signals generated by knee joints under stress during repeated sit-stand-sit movements along with joint angle. A statistically significant feature profile was established using a four-phase model of sit-stand-sit movements and two waveform features. The four-phase movement model is derived from joint angle measurement during repeated sit-stand-sit movements, and it consists of the ascending-acceleration and ascending-deceleration phases in the sit-to-stand movement, followed by the descending-acceleration and descending-deceleration phases in the stand-to-sit movement. The two statistically significant waveform features are extracted from AE measurement during repeated sit-stand-sit movements, and they consist of the peak magnitude value and average signal level of each AE burst. In addition to the use of bilateral plots, statistical distributions and 2D colour histograms to visualise the differences and similarities among participants, use of principal component analysis showed not only distinct data clusters corresponding to participating groups, but also an age- and disease-related trajectory progressing from the early adulthood healthy group to the late adulthood healthy group followed by the middle adulthood osteoarthritic group to the late adulthood osteoarthritic group. Furthermore, this trajectory shows increasing areas for each data cluster, with a highly compact cluster for the early adulthood healthy group at one end and a widely spread cluster for the late adulthood osteoarthritic group at the other end. From these results, a strong basis is formed for further development of knee acoustic emission as a convenient and non-invasive biomarker for quantitative assessment of joint ageing and degeneration.     

Synthesis and Anticonvulsant Activity of New 2-(4-Oxo-2-Thioxo-1,4-Dihydro-3(2h)-Quinazolinyl)Acetamides

Pharmaceutical Chemistry Journal - A search for new anticonvulsants led to a series of 2-(4-oxo-2-thioxo-1,4-dihydro-3(2H)-quinazolinyl)- acetamides that were synthesized by reacting...     

Exploratory study of a non-invasive method based on acoustic emission for assessing the dynamic integrity of knee joints

The paper presents the development of a new measurement system based on acoustic emission (AE) for assessing the dynamic integrity of knee joints, and the evaluation of its efficacy, through an exploratory study using healthy and osteoarthritic knees. For the former, the paper describes the system implemented to acquire joint angle-based AE and the protocol developed to obtain repeatable results. For the latter, the paper reports significant differences between healthy and osteoarthritic knees using statistical analysis of AE occurrence and distribution of AE features in different movement phases. Osteoarthritic knees are found to produce 6–10 times more acoustic emissions than healthy knees, with amplitudes which can be 20 dB higher, and durations which can be 10 times longer. These findings lead to a visual representation method of AE feature profiles based on multidimensional density of AE hits per repeated movement and the use of principal component analysis for objective assessment of the clinical status of knee joints.