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PubMed2004

In vitro synthesis of a crystalline (1-->3,1-->4)-beta-D-glucan by a mutated (1-->3,1-->4)-beta-D-glucanase from Bacillus.

Oligo- and poly-saccharides have a large number of important biological functions, and they occur in natural composite materials, such as plant cell walls, where they self-assemble during biosynthesis in a poorly understood manner. They can also be used for the formation of artificial composite materials with industrial applications. Fundamental and applied research in biology and nanobiotechnology would benefit from the possibility of synthesizing tailor-made oligo-/poly-saccharides. In the present paper, we demonstrate that such syntheses are possible using genetically modified glycoside hydrolases, i.e. glycosynthases. The ability of the endoglycosynthase derived from Bacillus (1-->3,1-->4)-beta-D-glucanase to catalyse self-condensation of sugar donors was exploited for the in vitro synthesis of a regular polysaccharide. The specificity of the enzyme allowed the polymerization of alpha-laminaribiosyl fluoride via the formation of (1-->4)-beta-linkages to yield a new linear crystalline (1-->3,1-->4)-beta-D-glucan with a repeating 4betaG3betaG unit. MS and methylation analyses indicated that the in vitro product consisted of a mixture of oligosaccharides, the one having a degree of polymerization of 12 being the most abundant. Morphological characterization revealed that the (1-->3,1-->4)-beta-D-glucan forms spherulites which are composed of platelet crystals. X-ray and electron diffraction analyses allowed the proposition of a putative crystallographic structure which corresponds to a monoclinic unit cell with a =0.834 nm, b =0.825 nm, c =2.04 nm and gamma=90.5 degrees. The dimensions of the ab plane are similar to those of cellulose I(beta), but the length of the c -axis is nearly twice that of cellulose I. It is proposed that four glucose residues are present in an extended conformation along the c -axis of the unit cell. The data presented show that glycosynthases represent promising enzymic systems for the synthesis of novel polysaccharides with specific and controlled structures, and for the analysis in vitro of the mechanisms of polymerization and crystallization of polysaccharides.

Bacillus↗

PubMed2001

Choroid plexus and ageing in rats: a morphometric and ultrastructural study.

Choroid plexuses (CP) are intraventricular structures involved in the production of cerebrospinal fluid (CSF) and in the synthesis and transport of numerous CSF components. Age-related modifications of the CP structure are still ill defined. We performed an ultrastructural and morphometric study of ageing CP in nine Sprague-Dawley rats 6, 18 and 30 months of age. Epithelial cells of CP villi were cubic in shape at 6 months, more dome-like at 18 months, and significantly flattened at 30 months of age. Epithelial basement membranes were thin and regular at 6 months, significantly thicker at 18 months and thicker and irregular at 30 months. Intravillous stroma increased nonhomogeneously with age. The ageing of CP in rats is characterized morphologically by epithelial atrophy, irregular fibrosis of the stroma and a thickening of epithelial basement membranes. These modifications suggest an alteration of secretory and filtrating functions in ageing CP.

Aging↗

PubMed1997

Proliferation and differentiation of cells from explants of fetal rat stomach.

The current understanding of the mechanisms controlling the proliferation and differentiation of the stem cells of the gastric oxyntic glands is limited. The aim of the present study was to develop a method for investigating proliferation and differentiation of undifferentiated cells from fetal rat stomach. Outgrowth of cells was initiated from explants of 16-day-old fetal rat stomachs. At this stage of the fetal development the gastric epithelial cells are undifferentiated. The explants were cultured in DMEM/F-12 medium supplemented with fetal calf serum only, or fetal calf serum combined with either hydrocortisone or pentagastrin. Morphological characterization by means of light microscopy, dye staining and immunostaining was used to identify the growing cells. Both hydrocortisone and pentagastrin accelerated the differentiation towards H,K-ATPase-positive cells, mucus-producing cells and other epithelial cells. H,K-ATPase-positive cells, which were identified by immunostaining with a monoclonal antibody reacting with the alpha-subunit of the H,K-ATPase, grew on top of the confluent layer of epithelioid and fibroblastoid cells. With this method in vitro investigations of the mechanisms of proliferation and differentiation of gastric mucosal cells are possible. Although by different mechanisms, both hydrocortisone and pentagastrin appear to play a regulatory role in these processes.

Animals↗

PubMed1998

Near-haploid common acute lymphoblastic leukaemia of childhood with a second hyperdiploid line: a DNA ploidy and fluorescence in-situ hybridization study.

Near-haploidy is a rare cytogenetic finding in childhood acute lymphoblastic leukaemia (ALL) and is associated with a poor prognosis. A second hyperdiploid line, occurring presumably by endoreduplication of the near-haploid stemline, is often observed. We present a case of common ALL in relapse characterized morphologically by a dual population of small and large lymphoblasts. Cytogenetic analysis supplemented with fluorescence in-situ hybridization (FISH) studies localized near-haploidy and hyperdiploidy to the small and large blast population respectively. DNA ploidy determination confirmed two abnormal clones with near-haploidy as the predominant one. A novel t(9;12)(q11;q13) was present in the near-haploid clone and was duplicated in the hyperdiploid clone. This finding identified cells bearing near-haploidy to be the clonogenic population following malignant transformation and confirmed endoreduplication as the mechanism for the presence of associated hyperdiploidy.

Child↗

PubMed1997

The c-erbB-2/neu proto-oncogene in human pituitary tumours.

OBJECTIVE: The aetiology of most pituitary tumours remains unknown. We have examined the potential role of neu receptor proto-oncogene in human pituitary tumorigenesis. MATERIALS AND METHODS: Ten non-tumorous pituitary glands and 16 morphologically characterized functional and clinically non-functioning pituitary adenomas were studied. Protein expression was examined by immunohistochemistry, mRNA expression by RT-PCR and competitive PCR, gene amplification by differential PCR, and point mutations by DNA sequencing. RESULTS: Cytoplasmic positivity for neu was identified in a few scattered cells of the non-tumorous adenohypophysis using an antibody to the intracytoplasmic domain of neu, but no membrane staining was found with an antibody to the extracellular domain; the latter is said to reflect gene amplification. mRNA transcript signals of the expected size were identified in the normal adenohypophysis and in all 16 adenomas examined. No increase in the degree of mRNA expression, however, was noted in the different tumour types compared to normal human pituitary tissue as determined by competitive PCR. As neu can be activated to an oncogene by a point mutation in the transmembrane region, nucleotide substitutions in this domain were investigated. Direct sequencing of codon 659 revealed no point mutations in any of the tumours. Furthermore, since amplification of neu has been noted in various human malignancies, DNA from these tumours was examined by differential PCR. No detectable differences were noted between the neu gene and the single-copy reference gene IFN-gamma. CONCLUSION: The neu gene is expressed in adenohypophysial cells and their tumours. In pituitary adenomas, this expression is not associated with gene amplification or activating mutations to suggest a direct role in pituitary tumorigenesis.

Adenoma↗

PubMed1998

Cytoplasmic staining of erbB-2 but not mRNA levels correlates with differentiation in human thyroid neoplasia.

OBJECTIVE: Amplification and overexpression of the erbB-2 proto-oncogene in human carcinomas may have prognostic significance. Its role in thyroid carcinoma is controversial. We investigated human thyroid tumours for erbB-2 gene amplification, activating mutations in the transmembrane domain, quantitative mRNA expression and protein expression. MATERIALS AND METHODS: DNA and mRNA were extracted from 47 morphologically characterized, frozen thyroid tumours including 10 nodular hyperplasias, 3 follicular carcinomas and 34 papillary carcinomas (4 with tall-cell features, 2 with insular and 2 with anaplastic de-differentiation). DNA amplification was analysed by differential PCR. The transmembrane domain of erbB-2 was sequenced in all tumours for activating mutations in position 659. Levels of mRNA expression were determined by competitive mRNA RT-PCR. Immunohistochemistry (IHC) for erbB-2 protein expression in corresponding paraffin-embedded samples was evaluated. RESULTS: Our results showed no DNA amplification of erbB-2. Sequencing of the transmembrane domain of erbB-2 revealed no activating mutations. The level of mRNA expression was variable, 11 papillary carcinomas showing statistically significant elevated mRNA levels compared with corresponding normal thyroid tissue; however, this did not correlate with other indicators of poor prognosis. In contrast to elevated mRNA levels in tumours, the level of protein staining correlated with degree of differentiation, normal and hyperplastic tissue being strongly positive and poorly differentiated tumours negative. CONCLUSION: There are no mutations or amplifications of the erbB-2 gene in human thyroid tumours. Elevated erbB-2 mRNA expression in some thyroid tumours is not associated with clinical features of poor prognosis; however, the significance of the elevated mRNA levels is unclear, as it did not result in protein overexpression. Instead, cytoplasmic erbB-2 protein detection by IHC correlates with differentiation of human thyroid tumours and may be a feature of good prognosis.

Carcinoma, Papillary↗

PubMed1998

Assessment of fibre orientation in reinforced concrete using Fourier image transform.

In this study, ribbon-shaped amorphous cast-iron fibres were used to reinforce a concrete matrix. X-ray photographs have been taken to detect fibres in situ. Their orientation has been investigated by automatic image analysis methods. However, this measurement should not be influenced by the digitization on the square frame of the analyser. For that purpose, the Fourier transform was used rather than the rose of direction method. This analysis revealed the transverse isotropic nature of the spatial arrangement of these fibres, whose axis of revolution corresponds to the concrete casting axis. Such a morphological characterization of the fibre-reinforced concrete reveals its mechanical behaviour.

Journal Article↗

PubMed2000

Mitochondrial membrane potential differentiates cells resistant to apoptosis in hybridoma cultures.

Previous research has implicated mitochondrial physiology and, by extension, respiratory capacity in the initiation and progress of apoptosis of cells in culture and tissue environments. This hypothesis was tested by separating a hybridoma cell population into subpopulations of varying mitochondrial membrane potential (MMP) using Rhodamine 123 stain and fluorescence-activated cell sorter analysis and subjecting them to two apoptosis inducers, rotenone and staurosporin. Apoptotic death was characterized morphologically through the determination of apoptosis-related chromatin condensation and biochemically through the measurement of caspase-3 enzymatic activity. We found dramatic differences in the apoptotic death kinetics for the subpopulations, with the high MMP cells showing higher resistance to apoptotic death. After incubation with 30 microM rotenone, the low MMP cells exhibited one-third of the viability of the high MMP cells and a three-fold increase in the capsase-3 enzymatic activity. No changes were observed in the DNA content or the cell cycle distributions of the two cell subpopulations, which maintained their mean MMP difference after 20 generations. These results suggest that heterogeneity exists in mammalian cell populations with respect to mitochondrial physiology, which correlates with resistance to apoptotic death.

Amino Acid Chloromethyl Ketones↗

PubMed2000

Pathology of the arteries in the central nervous system with special reference to their dilatation: blood flow.

Arteries that belong to the central nervous system (CNS) have thick and monotonous internal elastic lamina (IEL), Willis ring, and no collateral branch to the systemic circulation. These characteristics suggest that the circulation of the CNS is constant. In order to know how flow controls the arterial configuration of the CNS, flow of the vertebral and basilar arteries was increased experimentally using eight adult male Japanese white rabbits. Flow increase was induced by the simple ligation of the bilateral common carotid arteries (ligation group, n = 4) or by the ligation of the bilateral common carotid arteries at 1 week after arteriovenous fistula (AVF) between the left common carotid artery and the left external jugular vein (ligation plus AVF group, n = 4). Cineangiography revealed distinct flow increase in the ligation group at 5 weeks after ligation and in the ligation plus AVF group at 5 weeks after AVF (4 weeks after ligation). Vertebral and basilar arteries were remarkably dilated and elongated. Histopathologically, severe disruption of the IEL and focal thinning of the media were distinct. Endothelial cells were preserved and there was no inflammatory cell infiltration. These morphological features are consistent with increased flow-induced adaptive remodeling. It is suggested that the constancy of the flow may give the arterial tree of the CNS these morphological characterizations.

Animals↗

PubMed1998

Trichobilharzia regenti n. sp. (Schistosomatidae, Bilharziellinae), a new nasal schistosome from Europe.

Members of the genus Trichobilharzia are parasitic in visceral or nasal body parts of their avian hosts. The examination of water snails in South Bohemia revealed a schistosome infection in Radix peregra snails. The experimental infection of ducklings (Anas platyrhynchos, Cairina moschata) confirmed that a new Trichobilharzia species--T. regenti n. sp.--was found. The adults, eggs and larvae (miracidia and cercariae) of the species were morphologically characterized. The adult occupy the nasal blood vessels where they lay the eggs; the miracidia hatch from the eggs directly in the tissue and leave the host during drinking/feeding of the infected birds.

Animals↗

PubMed2002

Electrocardiographic T-wave inversion: differential diagnosis in the chest pain patient.

Inverted T waves produced by myocardial ischemia are classically narrow and symmetric. T-wave inversion (TWI) associated with an acute coronary syndrome (ACS) is morphologically characterized by an isoelectric ST segment that is usually bowed upward (ie, concave) and followed by a sharp symmetric downstroke. The terms coronary T wave and coved T wave have been used to describe these ischemic TWIs. Prominent, deeply inverted, and widely splayed T waves are more characteristic of non-ACS conditions such as juvenile T-wave patterns, left ventricular hypertrophy, acute myocarditis, Wolff-Parkinson-White syndrome, acute pulmonary embolism, cerebrovascular accident, bundle branch block, and later stages of pericarditis.

Adult↗

PubMed1975

[Whipple's disease].

Whipple's disease is characterized morphologically by macrophages in the small intestine which store PAS-postive material, as well as coarsening and atrophy of the intestinal villi, with pseudocystic cavitations. Without treatment it ends fatally due to irreversible diarrhoea and cachexia. Until 1963, only 90 cases had been diagnosed, most of them at autopsy. Intestinal biopsy has facilitated the diagnosis in life. Treatment with antibiotics results in dramitic improvement and remission for many years. From this an at least partial bacterial cause of the disease has been deducted, but it has not been possible so far to identigy more precisely any causative bacteria. Two cases of Whple's disease with some special features are reported. Both occurred in women (previous reports were almost entirely of men). The severely ill patients were cured by tetracycline within a few weeks and have remained completely well at follow-up examinations. The morphological changes in the intestinal mucosa had partially regressed.

Adult↗

PubMed1975

[Diagnostic criteria of idiopathic hypertrophic subaortic stenosis in the echocardiogram(author's transl)].

Idiopathic hypertrophic subaortic stenosis (IHSS) is morphologically characterized by ventricular septal hypertrophy. It is asymmetrical because there is no corresponding hypertrophy of the posterior wall of the ventricle. The proportion between septal thickness and posterior wall thickness is more than 1.2. In addition, the anterior mitral leaflet moves towards the ventricular septum during the ventricular systole. Finally, the aortic cusps may close prematurely, even during ventricular systole, if there is a marked outflow-tract obstruction. The thickness of the septum and posterior wall, as well as the movement of the mitral and the aortic valves, can be easily registered by the echocardiograph. IHSS is, therefore, more easily diagnosed by this non-invasive method than by any other method. The echocardiogram demonstrates (1) asymmetrical septal hypertrophy, (2) anterior movement of the anterior and frequently also the posterior mitral leaflet in midsystole, (3) partial or complete closure of the aortic valve in mid-systole, (4) relatively small end-diastolic and systolic diameters of the left ventricle, (5) delayed early-systolic closure movement of the anterior mitral leaflet in the sense of a functional mitral stenosis, (6) decreased systolic septal movement.

Adult↗

PubMed2000

Thyrocyte-interleukin-1 interactions.

Autoimmune thyroid disease is the most common organ-specific autoimmune disease and is a very common cause of thyroid dysfunction such as autoimmune hypothyroidism, Graves' disease and postpartum thyroiditis. The thyroid gland from patients with autoimmune thyroid disease is morphologically characterized by massive infiltration of lymphoid cells. The interleukin-1 (IL-1) family of molecules is together with other cytokines an integral component of the complex intercellular communication required to mount and control an immune response. IL-1alpha/beta in moderate and high concentrations reversibly inhibit thyroid cell function, while IL-1beta in low concentrations stimulates thyroid cell function. The biphasic, non-cytotoxic and reversible influence of IL-1 supports a role of IL-1 in the physiological regulation of thyroid cell function. IL-1 stimulates the guanylate mediated pathways and inhibits the adenylate cyclase mediated pathways. All IL-1 effects are counteracted by IL-1 receptor antagonist indicating that the effects are exerted through activation of specific IL-1 receptors on thyrocytes. Furthermore, IL-1 induces or enhances expression of a number of immunologically active molecules such as adhesion molecules, cytokines, and complement regulatory proteins in thyroid epithelial cells. IL-1 may thus play a role during physiological as well as pathophysiological conditions contributing to for example the euthyroid sick syndrome and development of thyroid autoimmunity. This review summarizes current literature on the phenomenological in vitro influence of IL-1 on the thyroid cell.

Animals↗

PubMedDate not supplied

Compatible and incompetent Paxillus involutus isolates for ectomycorrhiza formation in vitro with poplar (Populus x canescens) differ in H2O2 production.

Isolates of Paxillus involutus (Batsch) Fr. collected from different hosts and environmental conditions were screened for their ability to form ectomycorrhizal symbiosis with hybrid poplar P. x canescens (= Populus tremula L. x P. alba) in vitro. The ability to form ectomycorrhiza varied between the fungal isolates and was not correlated with the growth rate of the fungi on agar-based medium. The isolate MAJ, which was capable of mycorrhiza synthesis under axenic conditions, and the incompetent isolate NAU were characterized morphologically and anatomically. MAJ formed a typical hyphal mantle and a Hartig net, whereas NAU was not able to penetrate the host cell walls and caused thickenings of the outer cell walls of the host. MAJ, but not NAU, displayed strong H2O2 accumulation in the outer hyphal mantle. Increases in H2O2 in the outer epidermal walls and adjacent hyphae of the incompetent isolate were moderate. No increases of H2O2 in response to the mycobionts were found inside roots. Suggested functions of H2O2 production in the outer hyphal mantle of the compatible interaction are: growth regulation of the host's roots, defence against other invading microbes, or increasing plant-innate immunity. The system established here for P. x canescens compatible and incompetent fungal associations will be useful to take advantage of genomic information now available for poplar to study tree-fungal interactions at the molecular and physiological level.

Basidiomycota↗

PubMed2005

[Idiopathic hypertrophic cranial pachymeningitis].

BACKGROUND: Idiopathic hypertrophic cranial pachymeningitis (IHCP) is a chronic inflammatory disease of unknown origin that is morphologically characterized by a local fibrotic thickening of the dura mater with a sterile lymphocytic infiltrate. Patients usually present with chronic headache, ataxia and cranial nerve palsies. Because of the great diversity of symptoms and the fact that IHCP can be associated with many other disorders, there is often a long time between the occurrence of clinical features and making the diagnosis. During this time symptoms can further increase. In addition, this is the third case report describing bone involvement in this disorder. CASE REPORT: In May 2000, a 51-year-old woman presented with reduced visual acuity on both eyes, complete palsies of the fourth and sixth cranial nerve, a partial palsy of the third cranial nerve and numbness and pain concerning the area of the fifth cranial nerve on the left side, painful eye movement on the left side, headache, and numbness and palsies on the left side of her body. Magnetic resonance imaging of the brain showed a focal gadolinium-enhanced small thickening of the dura mater close to the left hemisphere and a homogeneous bone thickening in the same area close to the thickened dura mater. Cerebrospinal fluid analysis revealed an elevated cell count (lymphocytic pleocytosis), routine blood counts and serum chemistry showed an elevated white blood count, an elevated C-reactive protein (CRP) and erythrocyte sedimentation rate (ESR). Therapy with corticosteroids resulted in a complete remission in November 2000 except for a persistent numbness of the left side of her face and body and reduced visual acuity on both sides. The following three years were characterized by repeated clinical deteriorations followed by a temporary increase of the daily steroid dose. Altogether, daily oral steroid therapy could slow down the progression of disease and improve the visual acuity when compared with the first examination. CONCLUSIONS: Chronic headache, ataxia and cranial nerve palsies in combination with inflammation and dural thickening should call to mind idiopathic hypertrophic cranial pachymeningitis. In principle, every ophthalmological or neurological symptom can be associated with IHCP.

Cranial Nerve Diseases↗

PubMed1987

[Morphologic criteria of prognosis and clinical course in ovarian tumors of the borderline type].

Borderline tumors of the ovary are morphologically characterized by histologic and cellular criteria and by lack of evidence of invasion. During an observation period of 12 years at Heidelberg University Gynecological Clinic, 49 patients with borderline tumors underwent surgery and the clinical course was followed up. In 15 cases the tumor was at an advanced stage (FIGO III/IV); chemotherapy was instituted after surgery in 12 of these women. None of the patients with early-stage tumors (FIGO I/II) were lost due to the tumor during the period of observation; the death rate among patients with stage III/IV tumors (FIGO) was 40%. However, the death of three of these women was also due to their advanced age and high internal risks. As in the case of ovarian carcinomas, the survival rates improve if the postoperative residual tumor mass is smaller than 2 cm. Subsequent histologic work-up of archived, formalin-fixed tumor tissue and metastases thereof showed that two forms of the metastasizing borderline tumor exist: one with a favorable prognosis, a "pure" form with borderline changes in the ovary and metastases, and one with a less favorable prognosis, with borderline changes in the ovary and destructive-invasive portions of tumor in the metastazation areas. Impulse cytophotometric studies (ICP) showed that all primary tumors had the same degree of ploidy i.e., diploid. However, detection of an aneuploid distribution pattern in a metastasis correlated with subsequent histological confirmation of tumor invasion and rapid tumor death. The S-phase fraction was low, at 2.02% +/- 1.3.(ABSTRACT TRUNCATED AT 250 WORDS)

Combined Modality Therapy↗

PubMed1986

Molecular basis of hereditary elliptocytosis due to protein 4.1 deficiency.

Hereditary elliptocytosis is a heterogeneous group of red-cell disorders characterized morphologically by oval-shaped erythrocytes. It is an infrequent but well-defined cause of hemolytic anemia. To investigate the molecular abnormality in this disorder, we examined the arrangement of the gene encoding protein 4.1, a membrane skeletal protein of human erythrocytes, in an Algerian family with hereditary elliptocytosis and severe hemolytic anemia due to complete deficiency of protein 4.1. Southern blot analysis of genomic DNA from hematologically normal and abnormal members of this family showed that the mutant gene had a DNA rearrangement upstream from the initiation codon for translation. The mRNA from the mutant locus was aberrantly spliced, and that abnormality was the most likely explanation for the deficiency of protein 4.1 in the family. We assigned the gene for erythrocyte protein 4.1 to the short arm (p) of chromosome 1, within a region from band 32 to the terminus (1p32----1pter). Other cases of hereditary elliptocytosis of unknown cause have been mapped to the same region of chromosome 1 by linkage to the Rh locus. The chromosomal location of the gene that codes for protein 4.1 suggests that hereditary elliptocytosis in one class of patients with the disorder may be caused by a mutation of this gene.

Blood Proteins↗