Demonstration of discrete secreted and membrane-bound ocular mucins in the dog

Our aims were to separate and characterize secreted canine ocular mucins, and to provide definitive evidence of membrane-bound mucins at the canine ocular surface. Mucus was collected by suction from the ocular surface of normal dogs and dispersed in guanidine hydrochloride and a cocktail of protease inhibitors. Caesium chloride density gradient centrifugation separated secreted mucins from membranes, which were collected from the top of the gradients. Membranes were extracted with octyl glucoside and screened using lectins and anti-mucin antibodies. Gradient fractions containing secreted mucins were constituted into pools on the basis of differential lectin and antibody staining. High molecular weight material from each pool was purified by gel filtration. This material, and the membrane extract, were reduced and alkylated. Vacuum blotting of separated materials after agarose gel electrophoresis was used to compare subunit structure. Density gradient profiles indicated three principal secreted glycoprotein peaks: one staining strongly with anti-mucin antibodies. Gel filtration demonstrated that each contained high molecular weight material. Vacuum blots demonstrated the presence of two secreted glycoproteins with differently sized subunits. On the basis of buoyant density, one of these may be lipid complexed. Membrane extracted material stained with anti-mucin antibodies, and vacuum blotting of this material provided evidence for two membrane-bound components. In conclusion, we have shown that normal canine ocular mucus contains two secreted mucins, each exhibiting different subunit structure; one of these mucins may undergo lipid complexation. Normal canine ocular mucus also contains two membrane-bound mucins: one of which is unique among membrane mucins in showing subunit structure.     

The interaction of peripheral blood leukocytes with alpha1-acid glycoprotein, its carbohydrate chains and neoglycoconjugates

The interaction of human peripheral blood leukocytes with alpha 1-acid glycoprotein (AGP), its glycoforms as well as neoglyco-conjugates representing carbohydrate chains of AGP or its fragments was studied by flow cytometry. It was shown that the main target cells for AGP as well as for conjugates of its carbohydrate chains with polyacrylamide (PAA) are monocytes and polymorphonuclear leukocytes but not lymphocytes. The interaction of AGP with monocytes and granulocytes are mediated by its carbohydrate chains: the binding of AGP with cells was inhibited by AGP, AGP oligosaccharides as well as conjugates of oligosaccharides and its fragments with PAA. The data obtained show the existence of monocyte (and granulocyte) receptors which interact with complex type sialooligosaccharides of AGP.     

Tissue specific expression of FMR-1 provides evidence for a functional role in fragile X syndrome

Prostaglandin E2 levels in isolated rat islets were increased from 64 +/- 11 pg/30 islets when incubated in medium containing 2 mM glucose to 115 +/- 9 pg/30 islets in medium containing 20 mM glucose. In contrast, glyceraldehyde (10 mM) reduced prostaglandin E2 levels to 29 +/- 6 pg/30 islets. Inhibition of glucose metabolism by mannoheptulose (10 mM) abolished the stimulatory effect of glucose on prostaglandin E2 levels and inhibited glucose-induced insulin release. The cyclooxygenase inhibitor, flurbiprofen (20 microM), did not affect insulin release caused by glucose or glyceraldehyde. In the presence of 1 mg/ml bovine serum albumin, insulin secretion induced by 20 mM glucose (6.9 +/- 1.1% of islet insulin content) was reduced by the lipoxygenase inhibitor BW755 C (20 microM) to 3.1 +/- 0.6%, and by the phospholipase A2 inhibitor, p-bromophenacyl bromide (10 microM), to 2.1 +/- 0.8%. In the absence of bovine serum albumin the inhibitory action of BW755 C and p-bromophenacyl bromide on glucose-induced insulin release was significantly more pronounced. These drugs whether in the presence or absence of bovine serum albumin, did not affect glyceraldehyde-stimulated insulin secretion. Glyceraldehyde (10 mM), potentiated glucose-induced insulin release in the presence of 2-8 mM glucose, but not for 10-20 mM glucose. Although the phospholipase A2 activator, melittin, initiated insulin release in the presence of 2 mM glucose and enhanced 10 mM glyceraldehyde-stimulated insulin secretion it had no effect on 20 mM glucose-induced insulin release. These two stimulatory effects of melittin on insulin release were totally abolished by p-bromophenacyl bromide.(ABSTRACT TRUNCATED AT 250 WORDS)     

c-KIT expression enhances the leukemogenic potential of 32D cells

The growth of human leukemic cells in culture and in vivo is dependent upon the presence of hematopoietic growth factors. Most populations of human leukemic acute myeloblastic leukemia (AML) cells express c-Kit on their surface and respond to Kit ligand (KL) in culture. To determine if this interaction was of potential significance in vivo we used a mouse model system. 32D cells, a murine IL-3-dependent myeloid cell line, were rendered KL responsive by transfection of the murine c-Kit. After injection of 32D or 32D-Kit cells into syngeneic hosts, animals bearing 32D-Kit cells, but not 32D cells, became moribund and were killed. These animals had circulating leukemic blast cells, infiltration of bone marrow, spleen, brain, liver, lung, and kidney. Cells recovered from some of the animals continued to be dependent upon IL-3 or KL for growth while in other cases the cells were factor independent. This model illustrates that the constitutive expression of c-Kit enhances the leukemic potential of 32D cells. The model will be useful for studying the progression of leukemia in vivo and testing whether interruption of the interaction of Kit and KL can affect the growth of leukemic cells.     

Pentacene-based organic thin-film transistors

Organic thin-film transistors using the fused-ring polycyclic aromatic hydrocarbon pentacene as the active electronic material have shown mobility as large as 0.7 cm²/V-s and on/off current ratio larger than 10⁸; both values are comparable to hydrogenated amorphous silicon devices. On the other hand, these and most other organic TFT's have an undesirably large subthreshold slope. We show here that the large subthreshold slope typically observed is not an intrinsic property of the organic semiconducting material and that devices with subthreshold slope similar to amorphous silicon devices are possible     

Endoneurial localisation of microvascular damage in human diabetic neuropathy

Twenty diabetic patients with neuropathy underwent clinical and neurophysiological evaluation together with a detailed morphometric assessment of capillary pathology in endoneurial and epineurial microvascular beds of the sural nerve. Morphological data were compared with ten non-diabetic control subjects. There were no significant differences in control subjects between basement membrane area, endothelial cell area, endothelial cell profile number or luminal area of endoneurial when compared with epineurial capillaries. In contrast, when compared with epineurial capillaries, endoneurial capillaries from diabetic patients demonstrated a significant increase in basement membrane (p < 0.001) and endothelial cell (p < 0.001) area and a significant reduction in luminal area (p < 0.001). There was no significant difference in endothelial cell profile number between endoneurial and epineurial capillaries amongst diabetic patients. Previous studies have demonstrated a good correlation between the degree of microangiopathy and measures of neuropathic severity. In the present study increased endoneurial capillary basement membrane area was significantly related to reduced peroneal nerve conduction velocity (p < 0.001), myelinated fibre density (p < 0.001) and elevated vibration (p < 0.05) and thermal (p < 0.001) perception. Increased endothelial cell area and reduced luminal size were related to a reduced peroneal nerve conduction (p < 0.05, p < 0.01, respectively), reduced myelinated fibre density (p < 0.05, p < 0.01) and elevated thermal perception (p < 0.05, p < 0.001). Epineurial capillary basement membrane, endothelial cell and luminal area failed to relate to measures of neuropathic severity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Pathogenesis of Mycobacterium bovis infection in cattle

This paper reviews the pathogenesis of Mycobacterium bovis infection in cattle, focusing on aspects relating to the host rather than the organism. A broad concept of pathogenesis has been considered and information is presented on sources and routes of infection, as well as the immune responses and pathology. In addition, data is presented on the excretion of M. bovis from tuberculous cattle.     

Genetic control of immunity to Heligmosomoides polygyrus: presentation of promiscuous antigens to parasite-specific T cell hybridomas

The role of MHC class II in the presentation of Heligmosomoides polygyrus antigens has been investigated, using a number of T cell hybridomas produced in A and E positive and negative mice. By using fixed and irradiated antigen presenting cells (APC), further evidence has emerged, to support earlier data, that there can be differential processing requirements during the presentation of H. polygyrus antigens by A and E molecules. In concordance with these earlier observations, this work provides further evidence than individual T cells can respond to antigen when presented by more than one MHC molecule. Previously, this evidence has been restricted to individual MHC molecules of the same haplotype, but these data show that H. polygyrus produces antigens which can be presented by both syngeneic and allogeneic MHC molecules. These antigens do not appear to be synonymous with the previously described H. polygyrus superantigen, as presentation is restricted to specific MHC haplotypes. It is proposed that H. polygyrus may produce these antigenic molecules as part of its strategy to manipulate the host immune system.     

Involution of the bovine mammary gland: histological and ultrastructural changes

Bovine mammary tissue was collected by surgical biopsy at intervals during involution for histological and ultrastructural observation. In lactating tissue (d 0 of involution, collected 8 h after the final milking), alveolar epithelial cells had marked ultrastructural evidence of lactation, including protein-containing secretory vesicles, lipid droplets, extensive rough endoplasmic reticulum, and numerous mitochondria. By d 2 of involution, alveolar epithelial cells contained large vacuoles apparently formed by coalescing of protein-containing secretory vesicles and lipid droplets. Large vacuoles were observed in epithelial cells until about the 3rd wk of involution. By d 2 of involution, the Golgi apparatus generally was not apparent. Rough endoplasmic reticulum and mitochondria were observed throughout the period studied, although in reduced amounts compared with their presence in lactating tissue. A marked increase in lysosomal or cytosegresomal structures in epithelial cells was not observed. There was no evidence of extensive sloughing of epithelial cells from the basement membrane. There was a progressive increase in the interalveolar area and a concurrent decrease in the alveolar luminal area as involution progressed. Ultrastructural examination showed that alveolar epithelial cells at d 21 and 30 of involution appear to be functionally active but not secreting milk components.