Influence of the primary disease on rehabilitation results in the early postoperative period in patients after orthotopic liver transplantation.

BACKGROUND: Rehabilitation after orthotropic liver transplantation (OLT) is a difficult clinical problem due to severe comorbidities and complications. We evaluated the rehabilitation progress in the early postoperative period of patients after OLT depending on the reason for liver transplantation. MATERIALS AND METHODS: We retrospectively evaluated 309 OLT recipients transplanted between 2003 and 2006, including 161 women and 148 men. We analyzed the results of early postoperative rehabilitation measured by the time to full, active, erection upright after surgery. We divided the patients into 5 groups due to etiology of liver failure: group A (n = 89; mean age, 46.7 years) consisted of patients with liver cirrhosis due to hepatitis B, C, or both; group B (n = 70; mean age, 43.8 years) included patients with primary biliary cirrhosis and primary sclerosing cholangitis; group C (n = 44; mean age, 46 years) conprised patients with postalcoholic liver cirrhosis; group D (n = 23; mean age, 29.7 years) had experienced acute or subacute liver failure; and group E (n = 83; mean age, 37.4 years) had other reasons of liver failure. All patients were included in an identical rehabilitation program. RESULTS: The mean time to full, active, erection upright was dependent on the reason for liver failure. The best result was observed in groups A and E (4.51 and 4.6 days, respectively), medium in groups B and C (5.3 and 5.02, respectively), and worst in group D (8.5 days). The differences between groups A, E, and D were significant. CONCLUSION: The best results were obtained in groups A and E, where full, active, erection upright was achieved at 4.51 and 4.60 days respectively, and worst in group D, where it was achieved on day 8.50. These results need to be taken into account in planning the rehabilitation process for OLT patients. When analyzing the correlation between full, active, erection upright and primary diseases, one of the factors contributing to the delay needs to be assumed to be the inability to develop compensating mechanisms in the cases of acute and subacute hepatic failures, resulting from the sudden development of the disease.

Factors determining the sward utilisation of winter pasture in cattle feeding.

In this review article, factors determining the sward utilisation of winter pasture in cattle feeding were defined and analyzed. The results from literature and own investigations have shown that yield and quality of autumn-saved herbage on winter pasture are determined by pratotechnical measures such as harvest date in winter and pre-utilisation date in summer, botanical composition of sward, and particularly the dominated grass species or community, nitrogen fertilisation and weather conditions in winter. The date of winter harvest as a dominating factor affects the development of dry matter yield as well as the digestibility of organic matter and nutrients concentration in herbage during winter. For the management of winter grazing systems it could be obtained that crude protein and energy concentration of the tested autumn-saved herbage met the requirements of suckler cows or beef cattle until the end of the year if they were pre-utilised in July. The weather conditions during autumn-winter period are important factor determining the accumulation of secondary metabolites formed by field fungi in herbage of winter pasture.

HbSC hemoglobinopathy suspected by chest x-ray and red blood cell morphology.

Thorax scan was performed for elucidation of a pulmonary problem in a Nigerian immigrant. The aspect of the vertebrae suggested sickle cell disease, of course without specification of the genotype. Routine hematological tests seemed compatible with an HbSC disease, showing typical laboratory features, namely a significant proportion of hyperchromic RBC, corresponding to secondary, non hereditary spherocytosis, presence of numerous target cells and occasional HbC crystals on Pappenheim stained blood films. The diagnosis of HbSC disease was confirmed by HPLC, iso-electric focusing and citrate agar electrophoresis of hemoglobin and by reverse phase HPLC of globin-chains. This case illustrates the importance of screening for hemoglobin anomalies as it is performed in a multiethnic country such as the Grand Duchy of Luxembourg

Short insertion in a hemoglobin chain: Hb Esch, an unstable alpha1 variant with duplication of the sequence Ala65-Leu-Thr-Asn68.

Hemoglobin (Hb) Esch, is an alpha1 variant, expressed at less than 5%, resulting from the duplication of the 12 nucleotides corresponding to CD65 through 68. The effect of this insertion is the repetition of the sequence Ala-Leu-Thr-Asn, which corresponds to the last turn of helix E. In this variant the presence of a one-turn elongated helix E causes instability and increased ligand affinity. Hb Esch was characterized by DNA sequencing and confirmed by electrospray mass spectrometry. Functional studies were performed by flash photolysis measurements on a fraction isolated by flatbed isoelectric focusing, which was enriched in the abnormal hemoglobin. Similar to other alpha chain variants due to short insertion (or deletion), Hb Esch probably results from a slipped mispairing mechanism. The stability of such modified proteins depends upon the region which is added or deleted and usually is more stable when involving a flexible loop or complete helix turn(s) near by.

Accumulation of mycotoxins in forage of winter pasture during prolonged utilisation of sward.

The concentration of mycotoxins in sward depending on time of pre-utilisation in summer and date of harvest in winter was analysed during the 2000-2002 vegetation seasons. Additionally, the yield of pasture sward was estimated. Higher concentrations of ochratoxin A--respectively: 0.55 and 050 ng/g DM--were found in the pasture sward harvested in December and January when compared to the sward collected in November (0.36 ng/g DM). The average concentration of zearalenone varied from 2.74 ng/g DM in December, to 6.00 ng/g DM in November and 9.77 ng/g DM in January. The highest concentration of ochratoxin A was 1.82 ng/g DM and zearalenone 47.89 ng/g DM. The highest number of samples in which ochratoxin A exceeded the level of 0.3 ng/g DM was noted in sward harvested in January (61.1%), while in the case of zearalenone, the number of samples containing this mycotoxin at a level exceeding 3.0 ng/g DM varied from 55.6% in December to 66.7% in November and January. It seems that the percentage of ochratoxin A positive samples and the toxin concentration level in the sward of winter pastures increased during the time of sward regrowth.

Hb Ernz [beta123(H1)Thr-->Asn] and Hb Renert [beta133(H11)Val-->Ala]: two new neutral variants revealed by reversed phase high performance liquid chromatography analysis.

Analysis of globin chains by reversed phase high performance liquid chromatography, used as an additional tool for characterizing hemoglobin variants, has led to the discovery of a new class of variants that display only differences in hydrophobicity. Two such variants are here described. Hb Ernz was found in a man of Italian origin who was polycythemic, and in two of his three daughters who were hematologically normal. Hb Renert, a slightly unstable variant, was found in a man from Cape Verde who also carried Hb S and presented with chronic hemolysis. The structural abnormalities were characterized by protein structure methods involving reversed phase high performance liquid chromatographic separations of globins and peptides, followed by mass spectrometry studies (electrospray, ion trap, tandem mass spectrometry).

Diversity among Field Populations of Bradyrhizobium japonicum in Poland.

Genetic structure in field populations of Bradyrhizobium japonicum isolated in Poland was determined by using several complementary techniques. Of the 10 field sites examined, only 4 contained populations of indigenous B. japonicum strains. The Polish bradyrhizobia were divided into at least two major groups on the basis of protein profiles on polyacrylamide gels, serological reaction with polyclonal antisera, repetitive extragenic palindromic PCR fingerprints of genomic DNA, and Southern hybridization analyses with nif and nod gene probes. Serological analyses indicated that 87.5% of the Polish B. japonicum isolates tested were in serogroups 123 and 129, while seven (12.5%) of the isolates tested belonged to their own unique serogroup. These seven strains also could be grouped together on the basis of repetitive extragenic palindromic PCR fingerprints, protein profiles, and Southern hybridization analyses. Cluster analyses indicated that the seven serologically undefined isolates were genetically dissimilar from the majority of the Polish B. japonicum strains. Moreover, immuno-cross-adsorption studies indicated that although the Polish B. japonicum strains reacted with polyclonal antisera prepared against strain USDA123, the majority failed to react with serogroup 123- and 129-specific antisera, suggesting that Polish bradyrhizobia comprise a unique group of root nodule bacteria which have only a few antigens in common with strains USDA123 and USDA129. Nodulation studies indicated that members of the serologically distinct group were very competitive for nodulation of Glycine max cv. Nawiko. None of the Polish serogroup 123 or 129 isolates were restricted for nodulation by USDA123- and USDA129-restricting soybean plant introduction genotypes. Taken together, our results indicate that while genetically diverse B. japonicum strains were isolated from some Polish soils, the majority of field sites contained no soybean-nodulating bacteria. In addition, despite the lack of long-term soybean production in Poland, field populations of unique B. japonicum strains are present in some Polish soils and these strains are very competitive for nodulation of currently used Polish soybean varieties.

Restriction of Nodulation by Bradyrhizobium japonicum Is Mediated by Factors Present in the Roots of Glycine max.

Reciprocal grafting experiments done using soybean plant introduction genotypes indicated that restriction of nodulation by Bradyrhizobium japonicum is determined by the genotype of the root and is dependent on plant growth temperature. Microscopic analyses indicated that the soybean plant introduction genotypes restrict nodulation of B. japonicum at symbiotic stages which occur both before and after the formation of nodule primordia.

Lectin staining of peritoneal mesothelial cells in vitro.

A survey of lectin-binding specificities present on rodent and human mesothelial cells propagated and maintained in tissue culture was made using fluorescein isothiocynate conjugated (FITC) lectins. Rodent and human cells exhibited cell-associated fluorescence following exposure to the FITC-lectins from C. ensiformis, T. vulgaris, A. hypogaea, E. cristagalli and B. simplicifolia, but not with lectins from G. max and D. biflorus. Rodent cells were also positive for FITC-M. pomifera lectin binding. Human, but not rodent, cells were positive for FITC-T. purpureas lectin binding. Exposure of rabbit mesothelial cells in vitro to FITC-lectins that bound to the cell surface resulted in the appearance of discrete loci of putatively intracellular fluorescence. Exposure of cells to ferritin-labelled T. vulgaris lectin at 37 degrees C for as little as 7.5 minutes resulted in the appearance of ferritin-size particles in intracellular vesicles. These results demonstrate 1. the presence of lectin-binding sites in and on peritoneal mesothelial cells from rodents and humans and 2. a possible role of such sites in mediating the entry of lectin-like endogenous molecules into the vacuolar apparatus of these cells.

In vivo and in vitro characterization of mesothelial lipid inclusions.

The nature of intracytoplasmic lipid inclusions found in cultured rabbit and rat peritoneal mesothelial cells was examined by ultrastructural and biochemical techniques. Transmission electron microscopy also demonstrated extracellular release of these lipid bodies. Differential fixation with tannic acid revealed 2 types of inclusions, lamellated (lamellar bodies) and nonlamellated (homogeneous). The lamellar bodies were found near or in the Golgi apparatus and on the cell surface where occasionally they were observed in exocytotic pouches. The homogeneous inclusions were the predominant species being found primarily intracellularly. Lipid bodies obtained from the culture media over the cells displayed on electron microscopy the same morphological characteristics as those seen intracellularly. Exposure of confluent cultures of mesothelial cells to the vital lipid stain Nile Red caused the appearance of intensely fluorescent droplets in or on the cells at wave lengths consistent with staining for phosphatidylcholine-rich vesicles. Incubation of the cells with (14C)-choline and subsequent analysis of phospholipid formation revealed high rates of (14C)-phosphatidylcholine addition to both intra- and extracellular lipid pools. Taken together, mesothelial cells exhibit lipid bodies similar in ultrastructure to the surfactant containing organelles of Type II pneumocytes.

Autosomal recessive polycystic kidney disease: characterization of human peritoneal and cystic kidney cells in vitro.

Renal cystic epithelia and peritoneal mesothelia from two humans with autosomal recessive polycystic kidney disease (ARPKD) were grown in culture. Cystic epithelial and mesothelial cells formed continuous monolayers in vitro. By electron microscopy, cystic renal cells exhibited a single apical cilium and numerous short, stubby microvilli, both in situ and in vitro. Mesothelial cells exhibited intra- and extracellular membrane-limited, lipid-filled vesicles and surface microvilli. Cystic kidney cells in vitro stained positive for lectins from Cancanavalia ensiformis (concanavalin A), Triticum vulgaris, Erythrina cristagalli, Ulex europeaus, and Arachis hypogaea. Immunocytochemical and lectin staining revealed the renal and peritoneal cells to be of collecting tubule and mesothelial origin, respectively. Both cell types showed large depositions of glycogen granules in the cytoplasm during propagation in certain culture media; in kidney cells, dibutyryl cyclic adenosine monophosphate (cAMP) abolished glycogen depositions. Glycogen deposition also was observed in liver tissue obtained by needle biopsy from one patient. No bacteria were cultured from nor endotoxin detected in the renal cyst fluid. Relative to serum, the cyst fluids contained low sodium, potassium, and chloride levels. Thus, cultured ARPKD cells demonstrate a number of characteristics that are different from cells derived from the autosomal dominant form of renal cystic disease (ADPKD).

Isolation and propagation in vitro of peritoneal mesothelial cells.

Mesothelial cells lining the peritoneal cavity are the primary site of molecular exchange during peritoneal dialysis, a life support system for over 50,000 patients worldwide. In this study, techniques are described for the isolation and propagation in culture of peritoneal mesothelial cells from rats and rabbits. For comparison, mesothelial cells were also obtained from the serosal surface of human colonic tissue. By electron microscopy the cultured cells were found to exhibit microvilli, a well-developed endoplasmic reticulum and golgi apparatus, micropinocytotic vesicles, and lipid-filled intracellular vesicles. Immunochemical probes revealed the expression by these cells in vitro of cytokeratin, fibronectin, vimentin, and keratin, but not von Willebrand factor. Mesothelial cells from rat, rabbit, and human exhibited contact inhibition, but differences in growth rates and dependence on supplements to the growth media. This work provides a multispecies comparison of the behavior of mesothelial cells in vitro for the purpose of developing an experimental system for the study of mesothelial cell biology and the role of these cells in peritoneal dialysis.

Coordinated synthesis of leghemoglobin and root protein R18 in yellow lupin.

Uninfected roots of yellow lupin contain an abundant 18 kDa protein (referred to as R18), absent in the mature nodules. Some properties of this polypeptide are apparently similar to those of lupin leghemoglobins. However, the lack of any immuno crossreaction between R18 and leghemoglobin and differences in N-terminal amino acid sequences indicate that these proteins are coded by different genes. The decrease in the content of R18 protein in developing nodule is associated with the increased synthesis of leghemoglobin. This implies coordination of both events.

Lupin leghemoglobins during root nodule development.

Two yellow lupin leghemoglobins, Lb I and Lb II, were purified to homogeneity using the HPLC technique for final separation. Lb I and Lb II were identified by the N-terminal sequences and their reaction with antibodies against electrophoretically pure leghemoglobin. The third Lb species was detected by the combined method of isoelectrofocusing and PAGE of Lb I. It seems that Lb III represents a posttranslational modification of Lb I. Developmental changes in Lb multiple forms were examined using the Western blotting method. The content of leghemoglobin, first detectable approximately 3 weeks after infection, increased up to 6-7 weeks, and then it remained at the same level until 8-9 weeks after the infection. At the early stages of nodule formation Lb I prevailed over Lb II, while later Lb II became the predominant form. This suggests physiological role of particular forms and precise regulation of the expression of Lb genes.

Identification of a 16-S RNA fragment crosslinked to protein S1 within Escherichia coli ribosomal 30-S subunits by the use of a crosslinking reagent: ethyl 4-azidobenzoylaminoacetimidate.

The bifunctional reagent ethyl 4-azidobenzoylaminoacetimidate was used to crosslink specifically ribosomal protein S1 to 16-S RNA within 30-S subunits. The reagent was attached to isolated protein S1. The modified protein was reassociated with protein-S1-depleted 30-S subunits and then crosslinked to the RNA molecule. The covalently bound 16-S RNA-protein S1 complex was isolated and the RNA fragment C-U-A-A-C-G-C-G-U-U-A-A-G-U-C-G-A-C-C-G-C-C-U-G-G-G-G-A-G (positions 861-889) was characterized to be crosslinked to protein S1.

Synthesis of a new reagent, ethyl 4-azidobenzoylaminoacetimidate, and its use for RNA-protein cross-linking within Escherichia coli ribosomal 30-S subunits.

A new reagent, ethyl 4-azidobenzoylaminoacetimidate, was prepared in a four-step synthesis starting from 4-aminobenzoic acid. This compound was used to cross-link RNA with proteins within the Escherichia coli 30-S ribosomal subunits. Following the reaction of the imidoester function with protein NH2 groups, photoactivation of the azide binds the other end of the reagent to RNA. The cross-linked proteins were labelled with 125I and identified by bidimensional gel electrophoresis. Proteins S3, S4, S5, S7, S9, S17, S18, and in a lower and more variable yield, S12, S13, S14 and S16 were bound to 16-S RNA. These results were confirmed by isolating cross-linked protein-oligonucleotide complexes from 30-S subunits containing 32P-labelled RNA.