The compositional abnormalities of lipoproteins in diabetic renal failure

BACKGROUND: Diabetic nephropathy (DN) is a common cause of chronic renal failure (CRF). Patients with DN have abnormal lipoprotein metabolism that can be influenced by both the impairment of renal function and the metabolic control of diabetes. The aim of the study was to explore the specific compositional lipoprotein abnormalities in patients with insulin-dependent DN in comparison with diabetic patients without nephropathy and non-diabetic CRF patients. METHODS: The lipid and apolipoprotein (apo) composition of major lipoprotein density classes was determined in 20 patients with insulin-dependent diabetes mellitus and nephropathy and compared with that in seven diabetic patients without nephropathy, 20 patients with non-diabetic CRF, and nine healthy control subjects. Lipoproteins isolated by preparative ultracentrifugation were very-low-density lipoproteins (VLDL), intermediate-density lipoproteins (IDL), low-density lipoproteins (LDL), and high-density lipoproteins (HDL). RESULTS: Patients with DN had a plasma lipid and apolipoprotein profile characteristic of renal dyslipoproteinaemia with increased concentrations of triglycerides and cholesterol, reduced levels of apoA-I and apoA-II and increased levels of apoB, apoC-II, apoC-III and apoE. These changes were more pronounced in diabetic than in non-diabetic patients with comparable degrees of renal failure. All density classes were characterized by abnormal concentration and composition of some lipid and apolipoprotein constituents. DN patients had a more than four-fold increase of VLDL mass, a three-fold increase of IDL mass, and a significant reduction of HDL mass compared to control subjects. They also had significantly higher concentrations of apoB, apoC-peptides and apoE particularly in VLDL and IDL, and to some extent in LDL. In HDL, DN patients had lower cholesterol, apoA-I, apoA-II and apoC-II levels than controls. The major compositional change in DN patients was a significant increase in the relative content of apoC-peptides in IDL and LDL. The lipoprotein abnormalities were more pronounced in patients with high HbA1c values. In addition, lower GFR and increased proteinuria were associated with higher concentrations of triglycerides and apoC peptides in VLDL, IDL, and LDL in DN patients. CONCLUSIONS: The results indicate that patients with DN share the characteristic features of dyslipidaemia of CRF with accumulation of intact or partially delipidized apoB-containing lipoproteins enriched in apoC-peptides and apoE, which are present not only in VLDL and IDL but also in LDL density range. The alterations are more marked in DN than in nondiabetic CRF patients reflecting the additional impact of metabolic control. Increased levels of these lipoproteins may represent risk factors for the accelerated development of atherosclerotic vascular disease in these patients.     

Semi-automatic segmentation of gated blood pool emission tomographic images by watersheds:application to the determination of right and left ejection fractions

Tomographic multi-gated blood pool scintigraphy (TMUGA) is a widely available method which permits simultaneous assessment of right and left ventricular ejection fractions. However, the widespread clinical use of this technique is impeded by the lack of segmentation methods dedicated to an automatic analysis of ventricular activities. In this study we evaluated how a watershed algorithm succeeds in providing semi-automatic segmentation of ventricular activities in order to measure right and left ejection fractions by TMUGA. The left ejection fractions of 30 patients were evaluated both with TMUGA and with planar multi-gated blood pool scintigraphy (PMUGA). Likewise, the right ejection fractions of 25 patients were evaluated with first-pass scintigraphy (FP) and with TMUGA. The watershed algorithm was applied to the reconstructed slices in order to group together the voxels whose activity came from one specific cardiac cavity. First, the results of the watershed algorithm were compared with manual drawing around left and right ventricles. Left ejection fractions evaluated by TMUGA with the watershed procedure were not significantly different (p=0. 30) from manual outlines whereas a small but significant difference was found for right ejection fractions (p=0.004). Then right and left ejection fractions evaluated by TMUGA (with the semi-automatic segmentation procedure) were compared with the results obtained by FP or PMUGA. Left ventricular ejection fractions evaluated by TMUGA showed an excellent correlation with those evaluated by PMUGA (r=0. 93; SEE=5.93%; slope=0.99; intercept = 4.17%). The measurements of these ejection fractions were significantly higher with TMUGA than with PMUGA (P<0.01). The interoperator variability for the measurement of left ejection fractions by TMUGA was 4.6%. Right ventricular ejection fractions evaluated by TMUGA showed a good correlation with those evaluated by FP (r = 0.81; SEE = 6.68%; slope = 1.00; intercept = 0.85%) and were not significantly different (P = 0.42). The interoperator variability with TMUGA was 6.7% for the right ventricle. Thus, the watershed algorithm proposed is an efficient segmentation tool for the semi-automatic analysis of right and left ventricular ejection fractions by TMUGA. Further studies are necessary to check whether this procedure can be used to evaluate ventricular volumes and cardiac outflow.     

Quality of life in patients with endocrine tumors of the gastrointestinal tract: patient and staff perceptions

Myo-inositol (mI) as a precursor in the phosphatidylinositol second messenger system has been reported to be reduced in depression. By means of proton-magnetic resonance spectroscopy (1H-MRS) the mI levels in the frontal brain were investigated in vivo in the present study. Twenty-two patients (mean age: 42.8 +/- 10.7 years) with depressive episodes according to ICD 10 (HAMD score > 17) were compared to 22 healthy subjects (28.0 +/- 5.3 years). Two voxels (30 x 20 x 20 mm3) in the frontal lobes were examined in a Siemens Magnetom SP 4000 at 1.5 T (STEAM sequence: TR = 3500 ms, TE = 55 ms). With the total creatine (Cr) as an internal standard, mI/Cr ratios were calculated to follow the mI levels. In the left frontal lobe, mI/Cr was 0.43 +/- 0.06 in depressive patients and 0.46 +/- 0.07 in healthy subjects; concerning the right frontal lobe, mI/Cr was 0.46 +/- 0.08 and 0.48 +/- 0.06, respectively. There were neither significant differences between the two groups nor between the hemispheres. Since there was a significant positive correlation (R = 0.6) between the age and the mI/Cr in the right frontal lobe of depressed patients, age matched pairs analysis was performed (n = 2 x 10, in each group: nine females, one male, < 40 years). In the right frontal lobe, the patients' mI/Cr of 0.40 +/- 0.05 was now significantly lower than the controls' mI/Cr of 0.45 +/- 0.06. However, most of the patients were on antidepressive medication. Interestingly, it was exactly this group of patients which showed significantly lower mI levels. We regard our investigation as a pilot study which suggests an influence of age and antidepressants on mI levels and should be taken into consideration in further investigations in depressive patients.     

Variability analysis of HIV-1 gp120 V3 region: IV. Distribution functions for intra- and inter-subtype amino acid hamming distances

Distribution functions for intra- and inter- HIV-1 V3-loop subtypes amino acid Hamming distances were calculated (850 V3-loop sequences from the Los Alamos HIV-1 Database (1996) were used). These functions have pronounced bell-like shape. Such shapes of the histograms for HIV-1 V3 intra- and inter-subtype distriutions are discussed to confirm the applicability of different hierarchical cluster procedures for HIV-1 V3 classification. Two-mode distribution for the subtype E could sertificate that this subtype includes two thinner taxons.     

New perspectives on the actions of sulphonylureas and hyperglycaemic sulphonamides on the pancreatic beta-cell

Sulphonylureas and the hyperglycaemic sulphonamide diazoxide are commonly employed in the therapy of non-insulin-dependent (Type 2) diabetes mellitus and insulinoma, respectively. Stimulatory effects of sulphonylureas on insulin secretion and the inhibitory action of diazoxide are thought to be primarily mediated through modulation of the activity of ATP-sensitive K+ channels (K(+)-ATP channels) in the beta-cell plasma membrane. Certain sulphonylureas are known to be internalised by the pancreatic B-cell. Recent studies suggest that these drugs and diazoxide can influence insulin secretion from electropermeabilized beta-cells in which K(+)-ATP channels and other plasma membrane ion channels are inoperative. This observation suggests that sulphonylureas and diazoxide interact with intracellular sites in the pancreatic B-cell which are directly involved in the regulation of the final stages of exocytosis.     

Location of type XV collagen in human tissues and its accumulation in the interstitial matrix of the fibrotic kidney

An antipeptide antibody was produced against the carboxyl-terminal noncollagenous domain of human type XV collagen and used to localize this recently described collagen in a number of human tissues. The most conspicuous findings were powerful staining of most of the capillaries and staining of the basement membrane (BM) zones of muscle cells. Not all of the BM zones were positive, however, as shown by the lack of staining in the developing fetal alveoli and some of the tubules in developing kidney. Nor was type XV collagen staining restricted to the BM zones, as some could be observed in the fibrillar collagen matrix of the papillary dermis and placental villi, for example. Interestingly, differences in the expression of type XV collagen could be observed during kidney development, and staining of fetal lung tissue suggested that changes in its expression may also occur during the formation of vascular structures. Another intriguing finding was pronounced renal interstitial type XV collagen staining in patients with kidney fibrosis due to different pathological processes. This suggests that the accumulation of type XV collagen may accompany fibrotic processes. Full-length human type XV collagen chains with an apparent molecular mass of approximately 200 kd were produced in insect cells using a baculovirus expression system. The fact that these had a markedly higher molecular mass than the 100- to 110-kd type XV collagen chains found in homogenates of heart and kidney tissue suggests either proteolytic processing during the synthesis of type XV collagen or an inability to solubilize complete molecules from tissues.     

Distribution of estramustine in the BT4C rat glioma model

When maintained under continuous selection with the folate inhibitor, methotrexate, cultured Aedes albopicfus mosquito cells amplify an 200 kb region of DNA containing the dihydrofolate reductase gene. To determine whether the amplicon contained additional coding regions, Southern blots of cosmid clones containing amplicon DNA were probed separately with reverse-transcribed mRNA from methotrexate-sensitive and methotrexate-resistant cells. Cosmid pWED118 contained five EcoRI fragments (A, B, C, F, G) ranging in size from 2 to 5 kb that hybridized with cDNA from resistant cells. Of these, fragments B and F also hybridized to probe representing mRNA from sensitive cells, and all but fragment G hybridized to repetitive DNA from wild-type cells. Fragment G, which appeared to encode a low copy number gene in wild-type cells that subsequently became part of the dihydrofolate reductase amplicon in methotrexate-resistant cells, hybridized strongly to a 7 kb band and more weakly to bands measuring 9 and 3 kb on Northern blots containing RNA from resistant cells. Fragment G contained a 1203 bp open reading frame, encoding 401 amino acids homologous to synaptic vesicle protein SV2, a member of a transmembrane transporter family expressed in neural and endocrine cells. The region of homology included the six N-terminal transmembrane domains, an internal cytoplasmic loop, a seventh transmembrane domain, and most of an intravesicular loop. This partial sequence, which appears to correspond to a truncated gene generated during formation of the dihydrofolate reductase amplicon, provides a useful basis for more extensive characterization of an important gene family that may be the target of novel insecticides.     

Screening of compounds interacting with HIV-1 proteinase using optical biosensor technology

A high-resolution optical biosensor assay for screening of low-molecular-weight compounds, using an immobilized protein target, has been developed. HIV-1 proteinase was immobilized on the sensor surface by direct amine coupling and a variety of inhibitors and noninteracting reference drugs were applied to the sensor surface in a continuous flow of buffer. The procedure did not require intrinsic reporter groups, substrates, inhibitors, or other ligands for detection. By using a reference protein, the signal could be corrected for the relatively large background signal caused by differences in dimethyl sulfoxide concentration between running and sample buffers. Substances binding with high affinity (Ki in nM range) required efficient regeneration of the sensor surface and washing of the injection system between sample cycles to get consistent results. Analysis was simplified by using report points, extracted during both association and dissociation phases, and a simple graphical display of data. The optimized assay could correctly distinguish HIV-1 inhibitors from other compounds in a randomized series, indicate differences in their interaction kinetics, and reveal artifacts due to nonspecific signals, incomplete regeneration, or carryover. The method is expected to be generally applicable to secondary screening of low-molecular-weight compound libraries with proteins as targets.