Suboptimal growth temperatures enhance the biological activity of cultured cyanobacterium Gloeocapsa sp.

The cytotoxic, antibacterial, and antifungal activities of cyanobacterium Gloeocapsa sp. strain Gacheva 2007/R-06/1 were investigated and the possibility for an enhancement of these activities by changing the culture conditions evaluated. Fatty acids of this cyanobacterium were found to be active against Streptococcus pyogenes. Exopolysaccharides inhibited the growth of both Gram-positive and Gram-negative bacteria and the fungus Candida albicans. Both exopolysaccharides and fatty acid mixtures also significantly decreased the viability of human cervical carcinoma cells, HeLa. Greater biological activities were exhibited by Gloeocapsa sp., cultured at suboptimal temperatures (15–26°C) than at optimal and supraoptimal ones. In comparison with higher light intensity, the low-light cultivation stimulated the cytotoxicity of the fatty acids. In general, low temperatures decreased the growth of Gloeocapsa sp., but promoted its biological activity. Prolonged cultivation also had a beneficial impact on the bioactivity. Compared to 4 days, the 17-day cultivation resulted in fourfold higher antibacterial and antifungal activities of exopolysaccharides and more than twice increases in their cytotoxicity. The study revealed that this cyanobacterial isolate is a new source of natural products with potential for pharmacological and medical applications.     

Design and synthesis of new substrates of HtrA2 protease

HtrA2 belongs to the HtrA (high temperature requirement A) family of ATP-independent serine proteases. The primary function of HtrA2 includes maintaining the mitochondria homeostasis, cell death (by apoptosis, necrosis, or anoikis), and contribution to the cell signaling. Several recent reports have shown involvement of HtrA2 in development of cancer and neurodegenerative disorders. Here, we describe the profiling of HtrA2 protease substrate specificity via the combinatorial chemistry approach that led to the selection of novel intramolecularly quenched substrates. For all synthesized compounds, the highest HtrA2-mediated hydrolysis efficiency and selectivity among tested HtrA family members was observed for ABZ-Ile-Met-Thr-Abu-Tyr-Met-Phe-Tyr(3-NO₂)-NH₂, which displayed a specificity constant k_cat/K_M value of 14,535 M⁻¹ s⁻¹.     

The rising incidence of type 1 diabetes in south-eastern Poland. A study of the 0-29 year-old age group, 1980-1999

INTRODUCTION: Studies carried out over the last few years have provided information about the increase in the incidence of type 1 diabetes in different parts of the world including the European countries bordering Poland. THE AIM OF STUDY: The aim of this study was to determine the long-term trends in the incidence of type 1 diabetes over the 20 years between 1980 and 1999 and to compare the incidence during the decades preceding and following the 1989 economic and political transformation in Poland. MATERIAL AND METHODS: The registration of type 1 diabetes among people aged 0-29 was drawn up according to the DERI recommendations using three data sources. We calculated the age-standardised incidence rates for five-year age groups and determined the long-term trend in the incidence of type 1 diabetes in south-eastern Poland. RESULTS: A significant growth in the incidence of type 1 diabetes was observed among people aged 0-29 in the Rzeszów Province in the period between 1980 and 1999. The mean age-standardised incidence rate was 6.1/100,000, and a statistically significant difference was noted between the 1980s and the 1990s (5.3 [95%CI 4.5-6.0] and 6.8 [95%CI 5.9-7.6]). The male incidence of 6.7 significantly exceeded that for females--5.5/100,000. There was also a higher incidence in the group aged 0-14 in comparison with the group aged 15-29 (6.4 and 5.8/100,000 respectively). The highest incidence was found in boys aged 10-14 (11.5/100,000) and a significantly rising trend was observed in children of 0-4 years old. CONCLUSIONS: The mean incidence of type 1 diabetes among the study population was low. Nevertheless, we demonstrated a significantly increasing trend in the incidence during the 20-year observation period. The incidence in the 1990s, both in general and for males, was significantly higher when compared to the 1980s.     

Application of a novel highly sensitive activity-based probe for detection of cathepsin G

Cathepsins are crucial in antigen processing in the major histocompatibility complex class II (MHC II) pathway. Within the proteolytic machinery, three classes of proteases (i.e., cysteine, aspartic, and serine proteases) are present in the endocytic compartments. The combined action of these proteases generates antigenic peptides from antigens, which are loaded to MHC II molecules for CD4+ T cell presentation. Detection of active serine proteases in primary human antigen-presenting cells (APCs) is restricted because of the small numbers of cells isolated from the peripheral blood. For this purpose, we developed a novel highly sensitive α-aminoalkylphosphonate diphenyl ester (DAP) activity-based probe to detect the serine protease cathepsin G (CatG) in primary APCs and after Epstein-Barr virus (EBV) exposure. Although CatG activity was not altered after short-term exposure of EBV in primary myeloid dendritic cells 1 (mDC1s), the aspartic protease cathepsin D (CatD) was reduced, suggesting that EBV is responsible for mitigating the presentation of a model antigen tetanus toxoid C-fragment (TTCF) by reduction of CatD. In addition, CatG activity was reduced to background levels in B cells during cell culture; however, these findings were independent of EBV transformation. In conclusion, our activity-based probe can be used for both Western blot and 96-well-based high-throughput CatG detection when cell numbers are limited.     

First synthesis of alpha-aminoalkyl-(N-substituted)thiocarbamoyl-phosphinates: inhibitors of aminopeptidase N (APN/CD13) with the new zinc-binding group

OO-Di-trimethylsilyl esters of alpha-N-benzyloxycarbonylaminoalkylphosphinates (III) undergo triethylamine catalyzed addition to isothiocyanates to give after hydrolysis, a series of new alpha-aminoalkyl-(N-substituted)thiocarbamoyl-phosphinates. Thiocarbamoyl-phosphinate moiety can be included in the structures of the metalloproteinase inhibitors as the zinc-binding group and the new compounds reported here are good inhibitors of important aminopeptidase N(CD13) with IC(50) in range of 10.56-0.25 microM.     

High-resolution analysis of neuronal growth cone morphology by comparative atomic force and optical microscopy

Neuronal growth cones are motile sensory structures at the tip of axons, transducing guidance information into directional movements towards target cells. The morphology and dynamics of neuronal growth cones have been well characterized with optical techniques; however, very little quantitative information is available on the three-dimensional structure and mechanical properties of distinct subregions. In the present study, we imaged the large Aplysia growth cones after chemical fixation with the atomic force microscope (AFM) and directly compared our data with images acquired by light microscopy methods. Constant force imaging in contact mode in combination with force-distant measurements revealed an average height of 200 nm for the peripheral (P) domain, 800 nm for the transition (T) zone, and 1200 nm for the central (C) domain, respectively. The AFM images show that the filopodial F-actin bundles are stiffer than surrounding F-actin networks. Enlarged filopodia tips are 60 nm higher than the corresponding shafts. Measurements of the mechanical properties of the specific growth cone regions with the AFM revealed that the T zone is stiffer than the P and the C domain. Direct comparison of AFM and optical data acquired by differential interference contrast and fluorescence microscopy revealed a good correlation between these imaging methods. However, the AFM provides height and volume information at higher resolution than fluorescence methods frequently used to estimate the volume of cellular compartments. These findings suggest that AFM measurements on live growth cones will provide a quantitative understanding of how proteins can move between different growth cone regions.     

Apolipoprotein A-I gene polymorphism and susceptibility of non-insulin-dependent diabetes mellitus

A polymorphic DNA sequence of the apolipoprotein (apo) A-I gene region was studied in relation to non-insulin-dependent diabetes mellitus (NIDDM). We have examined 100 patients with NIDDM and 100 control subjects as well. The presence of a unique 2.5-kilobase (kb) EcoRI fragment was found in 13 diabetic patients and two control subjects with family history of diabetes. There were six homozygotes and nine heterozygotes among them. The analysis of clinical data did not confirm any linkage between the presence of a 2.5-kb fragment and atherosclerosis. These findings suggest that the observed defect in the apo A-I gene region may confer susceptibility to NIDDM.