Morphofunctional condition of kidney after disruption of its lymphatic drainage

Dimentions of the renal corpuscle, proximal and distal convoluted tubules have been studied in the dog kidney, normal and the morphofunctional state of the organ after cutting and ligation of its different lymphatic vessels (4 series of experiments in 158 dogs). The observations have been performed in 0.5, 1, 3, 10-160 days of the experiments. Histological, histochemical and morphometrical methods have been applied; the residual nitrogen in blood has been determined. During early stages, disturbances in the pathways of lymph outflow result in development of edema and albumin saturation of interstitium. granular distrophy of convoluted tubules epithelium in the kidneys. In 40-160 days stromal diffuse sclerosis of the medullary substance develops. The changes described and deterioration of the nitrogen-excretory function of the organ are especially pronounced after the disturbance in the pathways of lymph outflow are combined with a simultaneous extirpation of the contralateral kidney. Under these conditions the processes of the renal compensatory hypertrophy slow down. The morphofunctional changes after the disturbance in the pathways of lymph outflow from the compensatory hypertrophied kidney are the least pronounced.     

Amide I band of IR spectrum and structure of collagen and related polypeptides

The infrared amide I band of collagens (rat and cod skin) and related compounds (polyproline, polyglycine, and polytripeptides) was studied. Assignment of amide I-band components for polyproline II and polytripeptides (Gly-Pro-Pro)_n and (Gly-Pro-Gly)_n in the solid state and water solution was made. Three amide I components observed in the polypeptide spectra were attributed to three different peptide CO groups in each triplet. On the basis of this assignment, the interpretation of the amide I multicomponent structure in collagen and isomorphous oligo- and polypeptides was attempted. The ordering of intra- and intermolecular hydrogen bonds involving peptide CO groups in collagen and related compounds was discussed.     

The development of modified human Hsp70 (HSPA1A) and its production in the milk of transgenic mice

The production of major human heat shock protein Hsp70 (HSPA1A) in a eukaryotic expression system is needed for testing and possible medical applications. In this study, transgenic mice were produced containing wild-type human Hsp70 allele in the vector providing expression in the milk. The results indicated that human Hsp70 was readily expressed in the transgenic animals but did not apparently preserve its intact structure and, hence, it was not possible to purify the protein using conventional isolation techniques. It was suggested that the protein underwent glycosylation in the process of expression, and this quite common modification for proteins expressed in the milk complicated its isolation. To check this possibility, we mutated all presumptive sites of glycosylation and tested the properties of the resulting modified Hsp70 expressed in E. coli. The investigation demonstrated that the modified protein exhibited all beneficial properties of the wild-type Hsp70 and was even superior to the latter for a few parameters. Based on these results, a transgenic mouse strain was obtained which expressed the modified Hsp70 in milk and which was easy to isolate using ATP columns. Therefore, the developed construct can be explored in various bioreactors for reliable manufacture of high quality, uniform, and reproducible human Hsp70 for possible medical applications including neurodegenerative diseases and cancer.     

Inventing Engineered Organoids for end-stage liver failure patients

Journal of Molecular Histology - End-stage liver disease (ESLD) is a term used clinically in reference to a group of liver diseases with liver transplantation as the choice of treatment. Due to the...     

Purification and some catalytic properties of phosphoglucomutase from maize leaves

Phosphoglucomutase (EC 2.7.5.1, PGM) was purified to homogeneity from maize (Zea mays L.) leaves. The enzyme had specific activity 11. 7 U/mg protein and molecular mass (determined by gel-chromatography) of 133 +/- 4 kD. The molecular mass of PGM subunits determined by SDS-electrophoresis was 66 +/- 3 kD. The enzyme had Km for glucose-1-phosphate and glucose-1,6-diphosphate of 20.0 +/- 0.9 and 16.0 +/- 0.8 &mgr;M, respectively. Concentrations of glucose-1-phosphate and glucose-1,6-diphosphate above 3 and 0.4 mM, respectively, cause substrate inhibition. The enzyme activity was maximal at pH 8.0 and temperature 35 degreesC. Magnesium ions activate the enzyme and manganese ions inhibit it. 3-Phosphoglycerate is an uncompetitive inhibitor of the enzyme (Ki = 1.22 +/- 0.05 mM). Fructose-6-phosphate, 6-phosphogluconate, and ADP activate PGM, whereas ATP, UTP, and AMP inhibit the enzyme. Citrate was also a potent inhibitor, inhibitory effects of isocitrate and cis-aconitate being less pronounced.     

Comparison of Worm Control Strategies in Grazing Sheep in Denmark

Control of nematode parasites with reduced reliance on the use of anthelmintics was studied in 16 ewes with suckling twin lambs on contaminated pasture in Denmark. Ewes and lambs were treated with albendazole at turn-out 3 May. Ewes were removed from the groups on 26 July, and lambs were slaughtered on 11 October. The animals were allocated to 4 groups of 8 lambs and their 4 ewes. Group TS was treated with albendazole at weeks 3, 6 and 8 after turnout and set-stocked; group TM was similarly treated but moved to clean pasture in conjunction with the last drenching; group US was untreated and set-stocked, and group UM was left untreated but moved to clean pasture week 8 after turn-out. Supplementary feed was offered in June and August due to scarcity of pasture. Strategic treatments of ewes and lambs weeks 3, 6 and 8 after turn-out, with or without a move to clean pasture, were highly effective in controlling nematode infections for most of the season. This was reflected in better weight gains and carcass characteristics in the treated compared to untreated lambs, resulting in an average increase in the value of the product by 36%. The effect of moving without treatment (UM) on faecal egg counts was limited but peak pasture infectivity was reduced to less than 10% compared to the set-stocked group and weight gains of lambs were significantly better despite poor feed availability in late season. The study showed that under set-stocked conditions repeated anthelmintic treatments of both ewes and lambs in early season may ensure sufficient nematode control whereas moving animals to clean pasture without dosing was less efficient. The latter may, however, still be a viable option in organic and other production systems where routine use of anthelmintics is banned, particularly if weaning and moving are combined or a second move is performed.     

Purification and functional analysis of recombinant Acholeplasma laidlawii histone-like HU protein

HU is a most abundant DNA-binding protein in bacteria. This protein is conserved either in its heterodimeric form or in one of its homodimeric forms in all bacteria, in plant chloroplasts, and in some viruses. HU protein non-specifically binds and bends DNA as a hetero- or homodimer and can participate in DNA supercoiling and DNA condensation. It also takes part in some DNA functions such as replication, recombination, and repair. HU does not recognize any specific sequences but shows some specificity to cruciform DNA and to repair intermediates, e.g., nick, gap, bulge, 3′-overhang, etc. To understand the features of HU binding to DNA and repair intermediates, a fast and easy HU proteins purification procedure is required. Here we report overproduction and purification of the HU homodimers. The method of HU purification allows obtaining a pure recombinant non-tagged protein cloned in Escherichia coli. We applied this method for purification of Acholeplasma laidlawii HU and demonstrated that this protein possesses a DNA - binding activity and is free of contaminating nuclease activity. Besides that we have shown that expression of A. laidlawii ihf_hu gene in a slow-growing hupAB E. coli strain restores the wild-type growth indicating that aclHU can perform the basic functions of E. coli HU in vivo.     

Substrate specificity and reaction kinetics of an X-motif ribozyme

The X-motif is an in vitro-selected ribozyme that catalyzes RNA cleavage by an internal phosphoester transfer reaction. This ribozyme class is distinguished by the fact that it emerged as the dominant clone among at least 12 different classes of ribozymes when in vitro selection was conducted to favor the isolation of high-speed catalysts. We have examined the structural and kinetic properties of the X-motif in order to provide a framework for its application as an RNA-cleaving agent and to explore how this ribozyme catalyzes phosphoester transfer with a predicted rate constant that is similar to those exhibited by the four natural self-cleaving ribozymes. The secondary structure of the X-motif includes four stem elements that form a central unpaired junction. In a bimolecular format, two of these base-paired arms define the substrate specificity of the ribozyme and can be changed to target different RNAs for cleavage. The requirements for nucleotide identity at the cleavage site are GD, where D = G, A, or U and cleavage occurs between the two nucleotides. The ribozyme has an absolute requirement for a divalent cation cofactor and exhibits kinetic behavior that is consistent with the obligate binding of at least two metal ions.