Differential response of neuronal cells to a fusion protein of ciliary neurotrophic factor/soluble CNTF-receptor and leukemia inhibitory factor.

Ciliary neurotrophic factor (CNTF) displays neurotrophic activities on motor neurons and neural cell populations both in vivo and in vitro. On target cells lacking intrinsic expression of specific receptor alpha subunits cytokines of the IL-6 family only act in the presence of their specific agonistic soluble receptors. Here, we report the construction and expression of a CNTF/soluble CNTF-receptor (sCNTF-R) fusion protein (Hyper-CNTF) with enhanced biological activity on cells expressing gp130 and leukemia inhibitory factor receptor (LIF-R), but not membrane-bound CNTF-R. At the cDNA level, the C-terminus of the extracellular domain of human CNTF-R (amino acids 1-346) was linked via a single glycine residue to the N-terminus of human CNTF (amino acids 1-186). Recombinant Hyper-CNTF protein was expressed in COS-7 cells. Hyper-CNTF efficiently induced dose-dependent STAT3 phosphorylation and proliferation of BAF-3 cells stably transfected with gp130 and LIF-R cDNAs. While on BAF3/gp130/LIF-R cells, Hyper-CNTF and LIF exhibited similar biological responses, the activity of Hyper-CNTF on pheochromocytoma cells (PC12 cells) was quite distinct from that of LIF. In contrast to LIF, Hyper-CNTF stimulated neurite outgrowth of PC12 cells in a time- and dose-dependent manner correlating with the ability to phosphorylate MAP kinases. These data indicate that although LIF and Hyper-CNTF use the same heterodimeric receptor complex of gp130 and LIFR, only Hyper-CNTF induces neuronal differentiation. The therapeutic potential of Hyper-CNTF as a superagonistic neurotrophin is discussed.     

Intracellular photodynamic therapy with photosensitizer-nanoparticle conjugates: cancer therapy using a 'Trojan horse'.

Phthalocyanine-nanoparticle conjugates have been designed and synthesised for the delivery of hydrophobic photosensitizers for photodynamic therapy (PDT) of cancer. The phthalocyanine photosensitizer stabilized gold nanoparticles have an average diameter of 2-4 nm. The synthetic strategy interdigitates a phase transfer reagent between phthalocyanine molecules on the particle surface that solubilises the hydrophobic photosensitizer in polar solvents enabling delivery of the nanoparticle conjugates to cells. The phthalocyanine is present in the monomeric form on the nanoparticle surface, absorbs radiation maximally at 695 nm and catalytically produces the cytotoxic species singlet oxygen with high efficiency. These properties suggest that the phthalocyanine-nanoparticle conjugates are ideally suited for PDT. In a process that can be considered as cancer therapy using a 'Trojan horse', when the nanoparticle conjugates are incubated with HeLa cells (a cervical cancer cell line), they are taken up thus delivering the phthalocyanine photosensitizer directly into the cell interior. Irradiation of the nanoparticle conjugates within the HeLa cells induced substantial cell mortality through the photodynamic production of singlet oxygen. The PDT efficiency of the nanoparticle conjugates, determined using colorimetric assay, was twice that obtained using the free phthalocyanine derivative. Following PDT with the nanoparticle conjugates, morphological changes to the HeLa cellular structure were indicative of cell mortality via apoptosis. Further evidence of apoptosis was provided through the bioluminescent assay detection of caspase 3/7. Our results suggest that gold nanoparticle conjugates are an excellent vehicle for the delivery of surface bound hydrophobic photosensitizers for efficacious photodynamic therapy of cultured tumour cells.     

Incidence of chromosome 3, 7, 10, 11, 17 and X disomy in mature human sperm nuclei as determined by nonradioactive in situ hybridization

In situ hybridizations were performed on mature human sperm cells with biotin-labeled -satellite DNA probes specific for chromosomes 3, 7, 10, 11, 17, and X in order to reveal the disomy rate for each of these chromosomes. A total of 76 253 sperm nuclei from seven healthy probands aged 23–57 years were analyzed. An average of 12 000 sperm nuclei (at least 1500 per donor) showing hybridization were scored with each probe. The disomy rate as indicated by two distinct hybridization signals turned out to be similar for all chromosomes, ranging from 0.31% to 0.34%. There were no significant interindividual differences and no age correlation in the frequency of disomic sperm cells between the donors.     

Alexithymia,impulsiveness, emotion,and eating dyscontrol: similarities and differences between narcolepsy type 1 and type 2

Sleep and Biological Rhythms - Non-sleep symptoms, as depression, anxiety and overweight, are often encountered in narcoleptic patients. The purposes of this study are to evaluate mood,...     

Length–biomass equations to allow rapid assessment of semi-aquatic bug biomass in tropical streams

Length–biomass equations are relatively easy and cost-effective for deriving insect biomass. However, the exact relationship can vary between taxa and geographical regions. Semi-aquatic bugs are abundant and are indicators of freshwater quality, but there are no studies investigating the effect of habitat disturbance on their biomass, although it is useful in assessing ecological processes. We identified the best-fit length–biomass models to predict the biomass of semi-aquatic bugs (Hemiptera, Gerromorpha) collected from streams in Sabah, Malaysia. We used 259 juvenile and adult semi-aquatic bugs to compare a range of plausible length–biomass functions, and to assess whether relationships differed across the following families and body forms: (1) Cylindrostethinae, Gerrinae, and Ptilomerinae, which are subfamilies within Gerridae consisting of small-to-large bugs that have long and slender bodies, (2) Halobatinae, a subfamily within Gerridae, consisting of small-to-medium bugs with wide heads and thoraxes as well as short abdomens, and (3) Veliidae, which are small bugs with stout bodies. Estimation used five fitting functions – linear regression, polynomial regression order 2, 3, and 4, and power regression – on the following groupings: three body forms combined; each body form with life stages (juvenile and adult) combined; and each body form with life stages separated. Power regressions were the best fit in predicting the biomass of semi-aquatic bugs across life stages and body forms, and the predictive power of models was higher when the biomass of different body forms was calculated separately (specifically for Halobatinae and Veliidae). Splitting by life stages did not always result in additional improvement. The equations from this study expand the scope of possible future ecological research on semi-aquatic bugs, particularly in Southeast Asia, by allowing more studies to consider biomass-related questions.     

S-type lectins occur also in invertebrates: High conservation of the carbohydrate recognition domain in the lectin genes from the marine sponge Geodia cydonium

The marine sponge Geodia cydonium contains several lectins.The main component, called lectin-1, is composed of three tofour identical subunits. The subunits of the lectins were clonedfrom a cDNA library; two clones were obtained. From the deducedaa sequence of one clone, LECT-1, a mol. wt of 15 313 Da iscalculated; this value is in good agreement with mass spectrometricanalysis of 15 453 25 Da. The sequence of another clone, LECT-2,was analysed and the aa sequence was deduced (15 433 Da). Thetwo subunits have a framework sequence of 38 conserved aa whichare characteristic for the carbohydrate-binding site of vertebrateS-type lectins. Clustering of lectin sequences of various speciesfollowing their pairwise comparison establishes a dendrogram,which reveals that the sponge lectin could be considered asthe ancestor for vertebrate S-type lectins. Geodia cydonium lectin sponges S-type lectin     

Acetyl-CoA synthetase (ADP forming) in archaea,a novel enzyme involved in acetate formation and ATP synthesis

The anaerobic hyperthermophilic archaea Desulfurococcus amylolyticus, Hyperthermus butylicus, Thermococcus celer, Pyrococcus woesei, the hyperthermophilic bacteria Thermotoga maritima and Clostridium thermohydrosulfuricum and the aerobic mesophilic archaeon Halobacterium saccharovorum were grown either on complex media, on sugars or on pyruvate as carbon and energy sources. During growth acetate was formed as fermentation product by all organisms. The enzymes involved in acetyl-CoA formation from pyruvate and in acetate formation from acetyl-CoA were investigated:

1. Cell extracts of all species, both archaea and bacteria, catalyzed the coenzyme A-dependent oxidative decarboxylation of pyruvate with viologen dyes or with Clostridium pasteurianum ferredoxin as electron acceptors indicating a pyruvate: ferredoxin oxidoreductase to be operative in acetyl-CoA formation from pyruvate.
2. Cell extracts of all archaeal species, both hyperthermophiles (D. amylolyticus, H. butylicus, T. celer, P. woesei) and the mesophile H. saccharovorum, contained an acetyl-CoA synthetase (ADP forming), which catalyzes both acetate formation from acetyl-CoA and ATP synthesis from ADP and phosphate (P_i): Acetyl-CoA+ADP+P_i?Acetate + ATP+CoA. Phosphate acetyltransferase and acetate kinase could not be detected.
3. Cell extracts of the hyperthermophilic (eu)bacteria T. maritima and C. thermohydrosulfuricum contained phosphate acetyltransferase and acetate kinase rather than acetyl-CoA synthetase (ADP forming).

These data indicate that acetyl-CoA synthetase (ADP forming) represents a typical archaeal property rather than an enzyme specific for hyperthermophiles. It is proposed that in all acetate forming archaea the formation of acetate and of ATP from acetyl-CoA, ADP and P_i are catalyzed by acetyl-CoA synthetase (ADP forming), whereas in all acetate forming (eu)bacteria these reactions are catalyzed by two enzymes, phosphate acetyltransferase and acetate kinase.     

Age-Dependent Modifications and Further Localization of the CX 26-like Protein from Vicia faba L.

A strong age dependency together with alterations in the cellular distribution of CX 26 immunorelated protein(s) was found for differently developed leaves of Vicia faba L. With increasing age, an immunoreactive 40 kD band was observed in the soluble and microsomal fraction. In the cell wall protein preparation of young and fully differentiated leaves the 40 kD band was the minor constituent. A 33 kD polypeptide was dominantly localized in the microsomal fractions of all developmental stages and in SDS-extracts of total cell proteins of young leaves. A 21 kD protein together with a 16 kD polypeptide was associated with the cell wall fraction. The 21 kD protein, assumed to represent a plasmodesmatal constituent, was reduced with age. In SDS extracts, prepared from the different developmental stages of the leaves and of mesophyll protoplasts, the age-dependent appearance of the several immunostained bands was most obvious. A correlation of the 16, 33, and 40 kD bands to a turnover of the 21 kD protein is suggested. The reduced amount of the 21 kD protein with increasing age may be contemplated as an indication for a relative decrease of symplastic connections between cells of maturing leaves. This is in agreement with the results obtained by immunofluorescence studies using guard cell protoplasts. Here, observations pointed also to a reduction and final loss of CX 26-related protein at the protoplast surfaces.