Co-localization of KLF6 and KLF4 with pregnancy-specific glycoproteins during human placenta development.

Pregnancy-specific glycoproteins (PSGs) are major placental proteins essential for the maintenance of normal gestation. However, little is known about their gene expression regulation during placentation. It was previously demonstrated that the human core promoter binding protein recently renamed Krüppel-like factor (KLF) 6 binds to a highly conserved sequence within the PSG promoters and is mainly expressed in human term placenta. Here, we determined the expression pattern of the 13 other KLFs during human placental development. We demonstrate that eight KLFs exhibit specific expression patterns in human placental tissues and membranes, in favor of a functional cooperation of specific KLFs during placentation. In addition, we demonstrate that KLF6, KLF4 and PSG proteins are co-expressed in same cell types of placental villi and membranes. This experimental evidence further strengthens the potential cross talk of both transcription factors for PSG gene regulation in vivo.

[Plasma homocysteine measurement: a study of pre-analytical variation factors for conditions for total plasma homocysteine concentration]. / Dosage de l'homocystéine plasmatique: étude des facteurs de variation préanalytiques sur la concentration en homocystéine plasmatique totale.

An increase in homocysteine, a sulphur amino acid, is nowdays considered as a risk factor for cardiovascular diseases, and is independent of other risk factors. Reference range for total plasma homocysteine level in adults is usually 5-15 mmol/l. Hyperhomocysteinemia is defined as a fasting total plasma homocysteine level > 15 mmol/l. There may be also graded increased risks for subjects with homocysteinemia from 10 to 15 mmol/l. However, no threshold has been defined, partly because of the lack of standardization in pre-analytical and analytical steps. The aim of the present work was to evaluate three pre-analytical parameters on plasma homocysteine levels: i) the influence of three anticoagulants (EDTA, sodium citrate and lithium heparin); ii) the delay period of blood sample on ice before centrifugation; and iii) the advantages of strong acidic citrate at room temperature. The mean concentrations of total plasma homocysteine were different in function of the anticoagulant. These differences (EDTA minus lithium heparin or EDTA minus sodium citrate) were less than 10% however the used methods and could explain the good correlation between the results. However we recommend to keep the anticoagulant constant in the same study. When EDTA blood samples were immediately put on crushed ice, the maximum delay period before centrifugation could reach 4 hours. If ice is unavailable, strong acidic citrate at room temperature is a good alternative until for 4 hours.

A novel human odorant-binding protein gene family resulting from genomic duplicons at 9q34: differential expression in the oral and genital spheres.

Lipocalins are carrier proteins for hydrophobic molecules in many biological fluids. In the oral sphere (nasal mucus, saliva, tears), they have an environmental biosensor function and are involved in the detection of odours and pheromones. Herein, we report the first identification of human lipocalins involved in odorant binding. They correspond to a gene family located on human chromosome 9q34 produced by genomic duplications: two new odorant-binding protein genes ( hOBP (IIa) and hOBP (IIb) ), the previously described tear lipocalin LCN1 gene and two new LCN1 pseudogenes. Although 95% similar in sequence, the two hOBP (II) genes were differentially expressed in secretory structures. hOBP (IIa) was strongly expressed in the nasal structures, salivary and lachrymal glands, and lung, therefore having an oral sphere profile. hOBP (IIb) was more strongly expressed in genital sphere organs such as the prostate and mammary glands. Both were expressed in the male deferent ducts and placenta. Surprisingly, alternatively spliced mRNAs resulting in proteins with different C-termini were generated from each of the two genes. The single LCN1 gene in humans generated a putative odorant-binding protein in nasal structures. Finally, based on the proposed successive genomic duplication history, we demonstrated the recruitment of exons within intronic DNA generating diversity. This is consistent with a positive selection pressure in vertebrate evolution in the intron-late hypothesis.

Lipolysis, fatness, gender and plasma leptin concentrations in healthy, normal-weight subjects.

BACKGROUND: Relationship between plasma leptin and adiposity and gender has been reported in adults. Effects of age on plasma leptin is unclear and regulation of leptin production by white adipose tissue is still poorly understood. OBJECTIVES: To study if age and parameters of lipolysis are related to plasma leptin concentrations. METHODS: Seventy-seven healthy, normal-weight subjects (age range 19-82 y.) had measurements of body composition (18oxygen dilution technique) and of fasting plasma levels of leptin, glycerol, and nonesterified fatty acids (NEFA). RESULTS: Plasma leptin was correlated to NEFA (r = 0.28) and glycerol (r = 0.48) concentrations. The relationship between %fat and plasma leptin was best fitted by an exponential (r2 = 0.82). In multiple regression %fat, body mass index, glycerol, and gender, but not fat mass, age or NEFA contributed independently to the variation in log plasma leptin. Log plasma leptin was higher in women than in men for a given glycerol concentration. CONCLUSION: Adiposity, lipolysis, and gender are related to plasma leptin in healthy humans.

Bioelectrical impedance analysis measurements of total body water and extracellular water in healthy elderly subjects.

OBJECTIVE: To address whether: (1) bioelectrical impedance analysis (BIA) can provide precise and accurate estimates of total body water (TBW) and extracellular water (ECW) in healthy elderly subjects, that display age-induced changes in body composition, (2) BIA models are improved by introducing variables related to geometrical body-shape and osmolarity. DESIGN: Cross-validation of available BIA models and models developed in the study. SUBJECTS: 58 healthy elderly subjects (31 women, 27 men, 66.8+/-4.7 y, mean +/- s.d.) MEASUREMENTS: BIA at 5, 50 and 100 kHz, 18O labelled water measurements of TBW, Br measurements of ECW, anthropometric variables, plasma osmolarity. RESULTS: Published BIA models for estimating TBW, entail various degrees of bias. Precise models (SEE of the models 0.8 L at 100 kHz, 1.0 L at 50 kHz) involving height2/resistance, weight, gender, circumferences and plasma osmolarity were established with data from 30 subjects chosen at random. Cross-validation of an independent group (n = 28) showed no bias (-1.5+/-3.2 L at 100 kHz, -1.4+/-3.2 L at 50 kHz, P = NS). CONCLUSION: We conclude that BIA models with increased accuracy and precision for predicting ECW and TBW can be derived in healthy elderly subjects. Repeated measures had a mean difference of 0.2+/-1.2 L.

[Defense proteins in the corneo-conjunctival epithelium]. / Protéines de défense de l'épithélium cornéo-conjonctival.

The eyes are protected by the mechanical action of blinking and the washing action of tears; besides maintaining comfort and serving as the optical surface, the tear film forms the first line of defence. This role is mainly due to specific action of secretory immunoglobulins and continuous presence of unspecific proteins such as lysozyme, lactoferrin and tear lipocalin.

Fine genetic mapping of LCN1/D9S1826 within 9q34.

LCN1 gene encodes the tear lipocalin; the lipocalins are a large and growing family of proteins characterized by their ability to bind small hydrophobic molecules. We report here the location of a dinucleotide repeat microsatellite marker (D9S1826) close to LCN1 gene. Using the CEPH reference families, the position of LCN1 is located within the 9q34 genetic map between D9S23 and D9S158.

Tear protein G originates from denatured tear specific prealbumin as revealed by two-dimensional electrophoresis.

One-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (1D-SDS PAGE) of the non-denatured low molecular weight (MW) tear proteins (dilution in phosphate buffered saline or in the non-ionic detergent Triton X100) revealed no protein G but a strongly marked 23-kD related to a tear specific prealbumin (TSP) subunit coming with the known 15, 17, 18 and 20-kD TSP subunits. Under mild denaturating conditions of sample preparation with SDS dilution just before electrophoresis, 23-kD protein decreased and a faint 32-kD protein G appeared. Under stronger denaturing conditions of sample preparation with SDS treatment (boiling or freeze-thaw cycles), 23-kD protein disappeared and two main protein G forms (32 and 34-kD) and additional bands (29, 36, 39, 42, 57 and 60-kD) appeared depending on the sample treatment. The isoelectric pH (pI) of these proteins ranged from pH 5.2 to pH 5.4. Different two-dimensional electrophoresis methods revealed that: - in presence of SDS, 23-kD protein was spontaneously changed into 17-kD TSP and such a phenomenon was partially reversible by using a non-ionic detergent (Triton X100), new proteins appeared under denaturating processes were related to various protein G forms and originated from TSP group, proteins G were produced by the aggregation of TSP subunit related to MW 17-kD/pI 5.0 corresponding to the major TSP subunit, disulfide bond formation was shown to play a major role in the aggregation process although protein G group was not totally reduced by dithiothreitol. Such results suggest that protein G is an in vitro experimental artifact due to denaturing conditions with SDS treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Analysis of proteins eluted from hemodialysis membranes.

To further investigate the types of interactions occurring between blood and hemodialysis membranes, proteins were sequentially eluted from used dialysers. Four different membranes (cuprophan, hemophan, cellulose acetate and polyacrylonitrile) were successively treated with a hydrogen bond cleaving agent (10 M urea), an ionic detergent destabilizing the hydrophobic interactions between apolar groups (SDS solution), and a hydroxylamine solution at alkaline pH to release postulated covalently bound C3 fragments. The eluted proteins were analyzed by SDS-PAGE and immunological techniques. Total protein determinations demonstrate different behaviour of the membranes as regards the 'protein cake'. Electrophoretic analysis suggests that qualitative and quantitative differences in the binding of the blood proteins are related to the membrane material. Complement fragment studies indicate that the complement activating potential of the dialysis membrane may not be determined by the availability of potential binding sites for activated C3b. An attempt is made to correlate these results with the biocompatibility concept.