Macromolecular trafficking indicated by localization and turnover of sucrose transporters in enucleate sieve elements.

The leaf sucrose transporter SUT1 is essential for phloem loading and long-distance transport of assimilates. Both SUT1 messenger RNA (mRNA) and protein were shown to be diurnally regulated and to have high turnover rates. SUT1 protein was detected by immunolocalization in plasma membranes of enucleate sieve elements (SEs) in tobacco, potato, and tomato. Analysis by in situ hybridization showed that SUT1 mRNA localizes mainly to the SE and is preferentially associated with plasmodesmata. Antisense inhibition of SUT1 expression under control of a companion cell (CC)-specific promoter indicated synthesis of SUT1 mRNA in the CC. These results provide evidence for targeting of plant endogenous mRNA and potentially SUT1 protein through phloem plasmodesmata and for sucrose loading at the plasma membrane of SE.

Sentinel lymph node involvement and a high Breslow index are independent factors of risk for early relapse of melanoma.

AIM: The clinical relevance of sentinel lymph node (SLN) analysis was evaluated prospectively and compared with other known risk factors of relapse in early stage melanoma. METHODS: Surgery was guided by lymphoscintigraphy, blue dye and gamma probe detection. SLN were analysed by haematoxylin eosin (HE) histochemistry and multimarker immunohistochemistry (IHC). Disease free survival (DFS) was evaluated with Kaplan-Meier plots according to different parameters and Cox analyses of variance. RESULTS: From 210 patients a total of 381 SLN were excised. Lymphoscintigraphy identified all excised SLN with only 2 false positive lymphatic lakes. Fifty patients (24%) had tumour positive SLN. With a mean follow-up of 31.3 months, 29 tumour recurrences were observed, 19 (38%) in 50 SLN positive and 10 (6%) in 160 SLN negative patients. Strong predictive factors for early relapse (p < 0.0005) were SLN positivity and a high Breslow index. CONCLUSION: SLN tumour positivity is an independent factor of high risk for early relapse with a higher power of discrimination than the Breslow index.

Sugar transporters in higher plants--a diversity of roles and complex regulation.

Sugar-transport proteins play a crucial role in the cell-to-cell and long-distance distribution of sugars throughout the plant. In the past decade, genes encoding sugar transporters (or carriers) have been identified, functionally expressed in heterologous systems, and studied with respect to their spatial and temporal expression. Higher plants possess two distinct families of sugar carriers: the disaccharide transporters that primarily catalyse sucrose transport and the monosaccharide transporters that mediate the transport of a variable range of monosaccharides. The tissue and cellular expression pattern of the respective genes indicates their specific and sometimes unique physiological tasks. Some play a purely nutritional role and supply sugars to cells for growth and development, whereas others are involved in generating osmotic gradients required to drive mass flow or movement. Intriguingly, some carriers might be involved in signalling. Various levels of control regulate these sugar transporters during plant development and when the normal environment is perturbed. This article focuses on members of the monosaccharide transporter and disaccharide transporter families, providing details about their structure, function and regulation. The tissue and cellular distribution of these sugar transporters suggests that they have interesting physiological roles.

Sulla carnosa modulates root invertase activity in response to the inhibition of long-distance sucrose transport under magnesium deficiency.

Being the principal product of photosynthesis, sucrose is involved in many metabolic processes in plants. As magnesium (Mg) is phloem mobile, an inverse relationship between Mg shortage and sugar accumulation in leaves is often observed. Mg deficiency effects on carbohydrate contents and invertase activities were determined in Sulla carnosa Desf. Plants were grown hydroponically at different Mg concentrations (0.00, 0.01, 0.05 and 1.50 mM Mg) for one month. Mineral analysis showed that Mg contents were drastically diminished in shoots and roots mainly at 0.01 and 0.00 mM Mg. This decline was adversely associated with a significant increase of sucrose, fructose and mainly glucose in shoots of plants exposed to severe deficiency. By contrast, sugar contents were severely reduced in roots of these plants indicating an alteration of carbohydrate partitioning between shoots and roots of Mg-deficient plants. Cell wall invertase activity was highly enhanced in roots of Mg-deficient plants, while the vacuolar invertase activity was reduced at 0.00 mM Mg. This decrease of vacuolar invertase activity may indicate the sensibility of roots to Mg starvation resulting from sucrose transport inhibition. 14 CO2 labeling experiments were in accordance with these findings showing an inhibition of sucrose transport from source leaves to sink tissues (roots) under Mg depletion. The obtained results confirm previous findings about Mg involvement in photosynthate loading into phloem and add new insights into mechanisms evolved by S. carnosa to cope with Mg shortage in particular the increase of the activity of cell wall invertase.

Sucrose transporters in plants: update on function and structure.

In plants, sucrose is the major transport form for photoassimilated carbon and is both a source of carbon skeletons and energy for plant organs unable to perform photosynthesis (sink organs). As a molecule translocated over distance, sucrose has to pass through a number of membranes. Membrane transport of sucrose has therefore been considered for a long time as a major determinant of plant productivity. After several decades of physiological and biochemical experiments measuring the activity of sucrose carriers, unequivocal evidence came from the first identification of a cDNA coding a sucrose carrier (SoSUT1, Riesmeier et al. (1992) EMBO J. 11, 4705-4713). At present 20 different cDNAs encoding sucrose carriers have been identified in different plant species, in both dicots and monocots (one case). The total number is increasing rapidly and most importantly, it can be guessed from the results obtained for Arabidopsis, that in each species, sucrose transporters represent a gene family. The sequences are highly conserved and those carriers display the typical 12 transmembrane alpha-helices of members of the Major Facilitator superfamily. Yeast expression of those carriers indicate that they are all influx carriers, all cotransport sucrose and proton and that their affinity for sucrose is surprisingly similar (0.2-2 mM). All their characteristics are in agreement with those demonstrated at the physiological level in plants. These characteristics are discussed in relation to the function in plants and the few data available on the structure of those transporters in relation to their function are presented.

The sucrose carrier of the plant plasmalemma. III. Partial purification and reconstitution of active sucrose transport in liposomes.

The proteins from plasma membranes from sugar beet leaves were solubilized by 1% CHAPS and separated by size exclusion chromatography and by ion-exchange chromatography. The fractions enriched in sucrose transporter were monitored in three ways: differential labeling, ELISA, and reconstitution in proteoliposomes. When the plasma membranes were differentially labeled by N-ethylamaleimide in the presence of sucrose, a major peak of differential labeling was found at 120 kDa upon gel filtration. When this peak was recovered, denaturated by sodium dodecyl sulfate and reinjected on the gel filtration column, it yielded a peak of differential labeling at 42 kDa. When unlabeled membranes were used, the fractions eluted from the column were monitored by ELISA for their ability to recognize a serum directed against a 42 kDa previously identified as a putative sucrose carrier. The results paralleled those obtained by differential labeling, i.e. a major ELISA-reactive peak was found at 120 kDa upon gel filtration, and this peak yielded a peak most reactive at 40 kDa after denaturation. The 120 kDa peak prepared from unlabeled membranes was further separated on a Mono-Q column. The fractions were monitored by ELISA as described above, and reconstituted into proteoliposomes using asolectin. Active transport of sucrose, but not of valine could be observed with the reconstituted 120 kDa fraction. When the eluates from the Mono-Q column were reconstituted, the fractions exhibiting highest transport activity were enriched with a 42 kDa band. The data provide the first report concerning reconstitution of sucrose transport activity and confirm the involvement of a 42 kDa polypeptide in sucrose transport.

Sulfur dioxide inhibits the sucrose carrier of the plant plasma membrane.

Plasma membrane vesicles were prepared by phase partition from a microsomal fraction of broad bean (Vicia faba L.) leaf. In order to study the effects of sodium sulfite on active uptake of sucrose, the vesicles were artificially energized by a transmembrane pH gradient (delta pH) and/or a transmembrane electrical gradient (delta psi). At 1 mM, sulfite strongly inhibited sucrose uptake but did not affect the two components of the proton motive force, delta pH (measured by dimethyloxazolidine dione) and delta psi (measured by tetraphenylphosphonium). Moreover, sulfite did not inhibit the proton-pumping ATPase of the plasma membrane vesicles. These data demonstrate that sulfite may inhibit transport of photoassimilates in plant by a direct inhibition of the sucrose carrier of the plasma membrane.

Reconstitution of active sucrose transport in plant proteoliposomes.

The proteins of purified plasma membranes from sugar beet (Beta vulgaris L.) leaf were solubilized and separated on a size exclusion column. The fractions eluted from the column were monitored by ELISA with antibodies directed to a putative sucrose carrier protein. The peak most reactive in ELISA was approximately 120 kDa, and yielded a 40 kDa peak after denaturation by SDS. The 120-kDa peak was recovered and used for reconstitution experiments with asolectin. Upon imposition of an artificial pH gradient and electrical gradient, the obtained proteoliposomes exhibited active transport of sucrose, but not of valine. The active transport of sucrose was inhibited by N-ethylmaleimide and HgCl2.

Identification of a pollen-specific sucrose transporter-like protein NtSUT3 from tobacco.

Pollen cells are symplasmically isolated during maturation and germination. Pollen therefore needs to take up nutrients via membrane carriers. Physiological measurements on pollen indicate sucrose transport in the pollen tube. A cDNA encoding a pollen-specific sucrose transporter-like protein NtSUT3 was isolated from a tobacco pollen cDNA library. NtSUT3 expression is detected only in pollen and is restricted to late pollen development, pollen germination and pollen tube growth. Altogether these data indicate that pollen is supplied not only with glucose, but also with sucrose through a specific sucrose transporter. The respective contribution of each transport pathway may change during pollen tube growth.

Colocalization of collagen overexpression and inflammatory cell infiltration in the two-kidney one-clip rat model from the early days of hypertension onward.

In the first 6 days of hypertension, infiltrated mononuclear cells were colocalized with collagen (I) mRNA-overexpressing fibroblasts in the adventitial area of unclipped kidney. The number of adventitial infiltrated mononuclear cells was correlated with adventitial collagen (I) surface expansion. After 22 days of hypertension no collagen (I) mRNA-overexpressing fibroblasts or any increase in collagen area or mononuclear cell infiltration was observed. In the interstitium of unclipped kidney, collagen (I) mRNA overexpression, collagen (I) expansion and mononuclear cell infiltration began later, from the 7th day of hypertension, and kept increasing. In the clipped kidney, after expansion in the first 6 days of hypertension, the adventitial collagen remained stable. These results suggest that in the unclipped kidney fibroblastic activation begins within the first 6 days of hypertension in the adventitial area, but is transient, and fibrosis then spreads in the interstitium. Mononuclear cell infiltration is colocalized and correlated with adventitial and interstitial fibrosis. In the first 6 days, hypertension is not the only cause of fibrosis; the same level of adventitial fibrosis is detected in the nonhypertensive clipped kidney. All observed pathological phenomena could be detected within the first 3 days of hypertension.

Prevention of ischemia-reperfusion lung injury by sulfated Lewis(a) pentasaccharide. The Paris-Sud University Lung Transplantation Group.

Inhibition of polymorphonuclear neutrophil (PMN) adhesion to the pulmonary endothelium attenuates ischemia-reperfusion (I/R) lung injury. We hypothesized that 3'-sulfated Lewis(a) (SuLa), a potent ligand for the selectin adhesion molecules, may have a beneficial effect on I/R lung injury, as measured by the filtration coefficient (K(fc)), and reduce pulmonary sequestration of PMN as assessed by the lung myeloperoxidase (MPO) activity. Blood-perfused rat lungs were subjected to 30 min of perfusion, 60 min of warm ischemia, and 90 min of reperfusion after treatment with either SuLa (200 microg) or saline. Effects of SuLa on PMN adhesion to cultured human umbilical vein endothelial cells (HUVEC) stimulated with tumor necrosis factor-alpha and calcium ionophore were also investigated. Compared with preischemia conditions, I/R induced a significant increase in K(fc), which was attenuated with SuLa (80 +/- 8 vs. 30 +/- 5%; P < 0.001). SuLa reduced lung MPO and PMN adhesion to stimulated HUVEC. These results indicate that SuLa reduces I/R-induced lung injury and PMN accumulation in lung. This protective effect might be related to inhibition of PMN adhesion to endothelial cells.

Preeclampsia associated focal and segmental glomerulosclerosis and glomerular hypertrophy: a morphometric analysis.

Renal biopsies from hypertensive pregnant women performed 8 to 10 days postpartum were processed by morphometric analysis. We allocated the 74 patients into four groups according to the respective forms of pregnancy hypertension, i.e. preeclampsia and gestational hypertension. Groups I and II included preeclamptic women, with (group I) or without (group II) de novo FSGS. Groups III and IV included biopsies of women with isolated gestational hypertension, appeared during the third trimester (group III) or earlier (group IV). The control group included 17 biopsies from age-matched nonpregnant women presenting with isolated hematuria. Glomerular lesions of typical preeclampsia were seen in all the biopsies of groups I and II, and in some of women with gestational hypertension of groups III and IV. Our morphometric analysis of these renal biopsies showed a progressive increase in glomerular size from early gestational hypertension, gestational hypertension of the 3rd trimester, isolated preeclampsia, and finally preeclamptic nephropathy associated with FSGS. The largest glomeruli were seen in preeclamptic women with severe hypertension and histologic lesions of preeclampsia with FSGS. Thus, both systemic hypertension and glomerular hypertrophy seem necessary to induce FSGS in this type of pathology.

Satellite PET and lung cancer: a prospective study in surgical patients.

Positron emission tomography (PET) appears to be an innovative method for imaging the proliferative activity of malignant tissue, in particular by means of 18F-labelled fluorodeoxyglucose (FDG). The potential role of PET scanning was investigated in a satellite centre as an adjunct to conventional methods for estimating the likelihood of pulmonary malignancy. Therefore the sensitivity of detection of lung cancer in candidates was determined prior to exploratory or therapeutic thoracotomy by FDG PET imaging. The study involved 36 patients with abnormal chest roentgenogram and suspected lung cancer who were due for thoracotomy. The PET scans were evaluated qualitatively and semiquantitatively. Pulmonary malignancy was found in 31/36 patients and 29 had a focal increase in FDG pulmonary uptake. Benign pulmonary lesions were found in 5/36 patients, three of whom had a negative PET scan. The sensitivity of detection of lung cancer by FDG PET was therefore 93.5%. Bayesian study shows that FDG PET could be the most useful method in a population with a low prevalence of lung cancer. As illustrated by our study, a simple FDG PET scanning protocol in a satellite PET centre could provide adequate clinical information and help in deciding subsequent patient management.

Quantitative metabolic PET imaging of a plasma cell granuloma.

We report a patient who underwent surgical resection of two lung nodules that proved to be recurrent plasma cell granuloma, also known as inflammatory pseudotumor. Prior to surgery, positron emission tomography (PET) was performed with 18F-labeled fluoro-2-deoxy-D-glucose (18FDG) and rubidium-82 (82Rb). The 18FDG PET scan revealed that the nodules corresponded to two areas of intense uptake. PET imaging with 82Rb, the marker of flow, also showed intense uptake. Thus, PET demonstrated both a high degree of metabolic activity and increased perfusion. These features suggest a lesion with high cellular activity rather than a simple reparative process. The true nature of this lesion remains unknown.

The role of FDG-PET in the preoperative assessment of N-staging in head and neck cancer.

This prospective study based on 48 patients showed that FDG-PET has a significantly higher sensitivity for the detection of lymph node metastases compared with palpation and it appears that FDG-PET has a similar sensitivity to CT-scanning. According to our data, FDG-PET is a highly specific method in the evaluation of neck nodes. This new imaging technique allows a tridimensional study and is easy to interpret. Therefore, FDG-PET seems to be a valuable imaging technique for the detection of cervical lymph node metastasis.

[Oligosaccharide sulfate inhibitors of selectin-sugar interactions in inflammatory processes]. / Oligosaccharides sulfatés inhibiteurs des interactions sélectines-sucres intervenant dans les processus inflammatoires.

Leucocyte migration into lymphatic tissues or inflammatory sites depends upon the expression of adhesion molecules. Among these molecules, the selectins expressed on endothelial cells (E- and P-selectins) and leucocytes (L-selectin) recognize carbohydrate ligands such as sialyl Lewis A or sialyl Lewis X oligosaccharides due to the same positioning of NeuAc, Gal and Fuc residues in both isomeric structures. We have shown that the sialic acid residue could be replaced by a sulfate group such as in the sulfated Lewis A pentasaccharide, one of the most potent monovalent ligand for human E-selectin, which was shown to be very active in the prevention of ischemia reperfusion lung injury. In the same way, we have prepared through chemoenzymatic syntheses, two disulfated Lewis X pentasaccharides, the sulfated analogs of carbohydrate ligands found on GLYCAM 1, the natural receptor of L-selectin. Finally, based on the double recognition of L-selectin with Lewis type and glycosaminoglycan structures, we tentatively introduced a possible link between the selectin- and the integrin-mediated lymphocyte adhesion systems.

Stability and preliminary pharmacokinetic studies of 1-(2-chloroethyl)-3-[1-(5'-paranitrobenzoyl-2',3'-isopropylidene)-alpha, beta-D-ribofuranosyl]-1-nitrosourea (RFCNU), a nonimmunosuppressive nitrosourea.

Using high-performance liquid chromatography, the stability of RFCNU was monitored as a function of pH in aqueous buffers at 37 degrees C and as a function of temperature in plasma. The kinetics of degradation of RFCNU are apparently first-order. The log kappa-pH profile demonstrated the hydroxyl ion-catalyzed solvolysis and a maximum stability around pH 3.0. This analytic assay was reliable for quantitating intact RFCNU in biologic fluids. After administration of 400 mg of RFCNU orally to a female patient, no intact drug was excreted in the urine and plasma levels were very low.