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Biomedical subjects

J Wroblewski

Publications and source records attributed to J Wroblewski.

At least 19 recordsLinked to original sources
PubMed2002

Symptomatic abnormalities of dark adaptation in patients with EFEMP1 retinal dystrophy (Malattia Leventinese/Doyne honeycomb retinal dystrophy).

PURPOSE: To investigate the nature of symptomatic visual disturbance in patients with EFEMP1 retinal dystrophy in the absence of geographic atrophy or choroidal neovascularization. METHODS: Patients presenting to a tertiary referral centre underwent clinical evaluation, fluorescein angiography, colour contrast sensitivity, focal, pattern, and standard electroretinography, electrooculography, scotopic threshold perimetry and dark adaptometry. RESULTS: Clinical features included reduced central vision, difficulty passing from light to dark, and diffuse submacular and peripapillary deposits, which were hyperfluorescent by fluorescein angiography. Colour contrast thresholds were abnormal in all six patients studied and both pattern and focal electroretinograms were abnormal in five of six patients. The scotopic and mixed rod-cone single flash ERG was normal but two patients demonstrated reduced oscillatory potentials and one had borderline delayed 30 Hz responses. Scotopic thresholds were elevated and rod-mediated dark adaptation kinetics were markedly prolonged in all six patients when measured over the central visible confluent deposits. CONCLUSIONS: In patients with EFEMP1 retinal dystrophy with confluent macular deposits, scotopic sensitivity is reduced and dark adaptation kinetics are prolonged over the macular deposits but are normal elsewhere. These results emphasize the localised nature of functional deficits in some patients with EFEMP1 retinal dystrophy and correlate well with the patient's visual symptoms. Symptomatic visual dysfunction may precede the development of clinically evident geographic atrophy or choroidal neovascularization in this disorder.

Color Vision Defects↗
PubMed2002

Effects of irradiation on intestinal cells in vivo and in vitro.

The effects of irradiation on intestinal epithelial cells were analyzed in vivo and in vitro. The in vivo study was carried out on the rat small intestine and for the in vitro study the intestinal crypt cell-line IEC-6 was used. Rat intestine and IEC-6 cells were irradiated with X-ray doses ranging between 1-16 Gy. Energy-dispersive X-ray microanalysis was used for detection of the elemental changes in the cells. Cell morphology was investigated in the scanning electron microscope, DNA-synthesis by autoradiography of 3H-thymidine incorporating nuclei and proliferation by cell counting. Our results indicate that in vivo, in the crypt cells, the increasing doses of irradiation led to increased sodium and lowered potassium and phosphorus concentrations. Corresponding ion shifts were found in the irradiated IEC-6 cells. Cells continued to proliferate up to the dose of 8 Gy, although the proliferation rate became lower with increasing dose of irradiation. The increasing dose of irradiation significantly reduced DNA-synthesis (16 Gy decreased DNA-synthesis by 50%) which resulted in a complete inhibition of cell proliferation. Analysis of goblet cells also showed characteristic radiation-dependent elemental changes. Scanning electron microscopical investigation of cells in culture revealed that most of the control cells were flat and had rather smooth cell membranes. Irradiation led to the appearance of numerous different membrane manifestations (microvilli of varying length and distribution, and blebs). Frequency of differences in the topology of the cells was related to the dose of irradiation. Our study clearly demonstrates that even low doses of irradiation cause changes in the ionic composition of the cells and inhibit DNA-synthesis and cell proliferation. The effects observed in the crypt cells in vivo were the same as in the intestinal cell line in vitro, which indicates that IEC-6 cells can be used for investigation of side effects of radiation to the abdomen.

Animals↗
PubMed2001

Determining optimal surface roughness of TiO(2) blasted titanium implant material for attachment, proliferation and differentiation of cells derived from human mandibular alveolar bone.

In the complex process of bone formation at the implant-tissue interface, implant surface roughness is an important factor modulating osteoblastic function. In this study, primary cultures of osteoblast-like cells, derived from human mandibular bone, were used. The aim was to examine the effect of varying surface roughness of titanium implant material on cellular attachment, proliferation and differentiation. A recognized method of increasing surface roughness and enlarging the surface area of titanium implants is by blasting with titanium dioxide particles: the four specimen types in the study comprised surfaces which were machine-turned only, or blasted after turning, with 63-90 microm, 106-180 microm, or 180-300 microm TiO(2) particles, respectively. The specimens were analyzed by scanning electron microscopy and confocal laser scanning. The turned samples had the smoothest surfaces: average height deviation (S(a)) of 0.20 microm. The roughest were those blasted with 180-300 microm particles, S(a) value 1.38 microm. Blasting with intermediate particle sizes yielded S(a) values of 0.72 microm and 1.30 microm, respectively. Cell profile areas were measured using a semiautomatic interactive image analyzer. Figures were expressed as percentage of attachment. DNA synthesis was estimated by measuring the amount of [(3)H]-thymidine incorporation into trichloroacetic acid (TCA) insoluble cell precipitates. The specific activity of alkaline phosphatase was assayed using p-nitrophenylphosphate as a substrate. The ability of the cells to synthesize osteocalcin was investigated in serum-free culture medium using the ELSA-OST-NAT immunoradiometric kit. After 3 h of culture, the percentage of cellular attachment did not differ significantly between specimens blasted with 180-300 micromparticles and the turned specimens. All blasted surfaces showed significantly higher [(3)H]-thymidine incorporation than the turned surfaces (P<0.05), with the highest on the surfaces blasted with 180-300 microm particles. Osteocalcin synthesis by the cells in response to stimulation by 1,25(OH)2D3, was also significantly greater (P<0.05) on the surfaces blasted with TiO(2) particles. However, analysis of alkaline phosphatase activity disclosed no significant differences among the four surface modifications. It is concluded that in this cellular model, the proliferation and differentiation of cells derived from human mandibular bone is enhanced by surface roughness of the titanium implant. However, increasing the size of the blasting particles to 300 microm does not further increase the initial attachment of the cells compared to turned surfaces and those blasted with 63-90 microm particles.

Adolescent↗
PubMed2001

The characteristic cellular organization and CEACAM1 expression in the junctional epithelium of rats and mice are genetically programmed and not influenced by the bacterial microflora.

BACKGROUND: The epithelial cell adhesion molecule CEACAM1 exhibits an interesting dynamic expression during tooth development. It is first expressed in the reduced enamel epithelium, its expression then increases in the orally faced reduced epithelium and the overlying oral epithelium that then fuse to give rise to the junctional epithelium. The expression of CEACAM1 remains at high levels in the junctional epithelium, in contrast to the surrounding oral sulcular epithelium which shows much lower expression levels. We investigated if the high expression levels of CEACAM1 and the loosely organized cells characteristic of the junctional epithelium are genetically programmed or result from bacterial infiltration. METHODS: Oral tissues from germ-free rats and mice and animals with conventional bacterial flora were analyzed by transmission electron microscopy and immunohistochemical staining for CEACAM1. RESULTS: The junctional epithelium of both germ-free and conventional animals was identical with respect to both CEACAM1 expression and morphology. Also the presence of leukocytes was the same in both types of animals. CONCLUSIONS: The results indicate that the characteristic morphology and the high expression levels of CEACAM1 in the junctional epithelium are genetically programmed and not a result of bacterial infiltration. This suggests that CEACAM1 has an important role for the structural integrity of the junctional epithelium. This conclusion was supported by the observation that the junctional epithelium does not express any E-cadherin, which is another abundant epithelial cell adhesion molecule.

Animals↗
PubMed2000

Effects of titanium surfaces blasted with TiO2 particles on the initial attachment of cells derived from human mandibular bone. A scanning electron microscopic and histomorphometric analysis.

This study was performed to determine the effect of commercially pure titanium surfaces blasted with TiO2 particles on the biological responses of cells derived from human mandibular bone. The morphology and attachment of those cells were investigated on turned titanium surfaces (control) and surfaces blasted with 45 microns (standard), 45-63 microns, and 63-90 microns TiO2 particles. The surfaces were analyzed in a scanning electron microscope. Based on surface analyses reported elsewhere, the turned samples had the smoothest surfaces and the roughest were those blasted with the largest particles (63-90 microns). The cell profile areas were measured using a semi-automatic interactive image analyzer. The attachment was determined as a ratio of the area of cell profiles and the total micrograph area and was expressed as percentage of attachment. Morphologically, the cells were heterogeneous. In general, the cells had spread well on all titanium surfaces, indicating good attachment to both smooth and rough surfaces. After 1, 3 and 6 h, the percentage of cell attachment did not differ significantly between the surfaces blasted with 63-90 microns and the turned surfaces, but was significantly lower on the surfaces blasted with 45 microns or 45-63 microns particles. After 24 h the surfaces blasted with 63-90 microns particles had a higher rate of cell attachment than all the other surfaces including the controls. It is concluded that attachment and growth of cells originating from human mandibular bone in vitro, are influenced by the micro-texture of the implant surface.

Cell Adhesion↗
PubMed1999

Demonstration of estrogen receptor-beta immunoreactivity in human growth plate cartilage.

Estrogens affect longitudinal bone growth through their action on endochondral bone formation. Two estrogen receptors are known, the classical estrogen receptor-alpha (ER alpha), newly demonstrated in human growth plate cartilage, and a recently cloned estrogen receptor-beta (ER beta). The present study aimed to localize a possible expression of ER beta protein in human growth plates. Tissue samples were obtained from tibial and femoral growth plates in four female pubertal patients undergoing epiphyseal surgery. Immunohistochemistry, using two different ER beta-specific antibodies, demonstrated positive staining for ER beta in hypertrophic epiphyseal chondrocytes from all patients. No staining was noted in resting or proliferative chondrocytes. These data suggest that in addition to ER alpha, human epiphyseal chondrocytes also express ER beta. The physiological role of ER beta in the regulation of longitudinal bone growth in humans remains to be elucidated.

Adolescent↗
PubMed1998

Synthesis and metabotropic glutamate receptor activity of a 2-aminobicyclo[3.2.0]heptane-2,5-dicarboxylic acid, a molecule possessing an extended glutamate conformation.

A photochemical approach to the synthesis of the aminobicycloheptane 6a is reported. This compound assumes an extended glutamate conformation, and for this reason was created to further probe the structural features relevant to achieving selectivity for the subtypes of the metabotropic glutamate family of receptors.

Animals↗
PubMed1998

Dynamic expression of E-cadherin in ameloblasts and cementoblasts in mice.

During embryonic development, E-cadherin mediates intercellular adhesion in a variety of epithelia in a spatio-temporal pattern. We have analyzed the distribution of this cell adhesion molecule in the mouse during odontogenesis, at both mRNA and protein levels, in the mandibular first molars and incisors. E-cadherin was strongly expressed at the bell stage by the cells of the dental organ, and by the pre-secretory ameloblasts and the cells of stratum intermedium at the early mineralization stage. At the onset of enamel secretion, E-cadherin disappeared from the apical pool of the ameloblasts and was later absent from the post-secretory ameloblasts. E-cadherin was also found in Hertwig's root sheath and later in the cells producing acellular cementum. These findings indicate that E-cadherin may be involved in the polarization of the ameloblasts and in the early stages of cementogenesis.

Ameloblasts↗
PubMed1998

Rat alpha3/beta4 subtype of neuronal nicotinic acetylcholine receptor stably expressed in a transfected cell line: pharmacology of ligand binding and function.

We stably transfected human kidney embryonic 293 cells with the rat neuronal nicotinic acetylcholine receptor (nAChR) alpha3 and beta4 subunit genes. This new cell line, KXalpha3 beta4R2, expresses a high level of the alpha3/beta4 receptor subtype, which binds (+/-)- [3H]epibatidine with a Kd value of 304+/-16 pM and a Bmax value of 8942 +/- 115 fmol/mg protein. Comparison of nicotinic drugs in competing for alpha3/beta4 receptor binding sites in this cell line and the binding sites in rat forebrain (predominantly alpha4/beta2 receptors) revealed marked differences in their Ki values, but similar rank orders of potency for agonists were observed, with the exception of anatoxin-A. The affinity of the competitive antagonist dihydro-beta-erythroidine is >7000 times higher at alpha4/beta2 receptors in rat forebrain than at the alpha3/beta4 receptors in these cells. The alpha3/beta4 nAChRs expressed in this cell line are functional, and in response to nicotinic agonists, 86Rb+ efflux was increased to levels 8-10 times the basal levels. Acetylcholine, (-)-nicotine, cytisine, carbachol, and (+/-)-epibatidine all stimulated 86Rb+ efflux, which was blocked by mecamylamine. The EC50 values for acetylcholine and (-)-nicotine to stimulate 86Rb+ effluxes were 114 +/- 24 and 28 +/- 4 microM, respectively. The rank order of potency of nicotinic antagonists in blocking the function of this alpha3/beta4 receptor was mecamylamine > d-tubocurarine > dihydro-beta-erythroidine > hexamethonium. Mecamylamine, d-tubocurarine, and hexamethonium blocked the function by a noncompetitive mechanism, whereas dihydro-beta-erythroidine blocked the function competitively. The KXalpha3 beta4R2 cell line should prove to be a very useful model for studying this subtype of nAChRs.

Animals↗
PubMed1997

Expression of p53 and hsp70 in relation to apoptosis during Meckel's cartilage development in the mouse.

The process of Meckel's cartilage development was examined with regard to expression of p53, a tumor suppressor gene product and hsp70, a stress protein (heat-shock protein), in association with the occurrence of programmed cell death (apoptosis). Balb C mice embryos from embryonic days E13, E14, E15, E16, E17, E18 and 1- and 3-day-old pups were used. P53-positive cells were detected first at E15, and were found in the perichondrium of the distal part of Meckel's cartilage. During the degeneration process chondrocytes also became p53-positive. In contrast to p53, the expression of hsp70 was high and widespread in the early stages of development (E13-E15); however, it decreased with age, except for Meckel's cartilage, where hsp70 was found in the cytoplasm or nuclei of the hypertrophic cells. Apoptosis was first detected at E14-E15 in the perichondrium of the distal parts of Meckel's cartilage. The number of apoptotic bodies increased with age and the ongoing resorption of Meckel's cartilage. From the present study it can be concluded that expression of p53 and hsp70 varied during the development of Meckel's cartilage and that both proteins showed nuclear location in hypertrophic cells. No direct spatial or temporal correlation was observed between the expression of p53 and hsp70 and the occurrence of apoptotic bodies.

Animals↗
PubMed1997

Distribution of nestin in the developing mouse limb bud in vivo and in micro-mass cultures of cells isolated from limb buds.

Early skeletal muscle development is accompanied by changes in the composition of the cytoskeleton. In this report we analyze the distribution of the intermediate filament nestin in the developing mouse limb buds in vivo and in mesenchymal cells isolated from limb buds in vitro. The subcellular distribution of nestin mRNA and protein in muscle cells was also analyzed. We find a shift in nestin expression during early limb bud development. At embryonic day 11 (E11), low levels of nestin (protein) were expressed in the mesenchymal cells of the developing limb bud. Later, nestin mRNA and protein were down-regulated in the mesenchymal condensations undergoing chondrogenesis (E12 and E13), but remained expressed predominantly in the ectodermal cells and in the differentiating myoblasts. At E18, only muscle fibres, endothelial cells and nerves were nestin positive. This shift in expression was reproduced in vitro, in micro-mass cultures of mesenchymal cells. In E11 cultures, nestin protein was initially expressed in all cells. Upon formation of cartilage foci (after 2-3 days in culture), nestin immunoreactivity was not observed in cartilage, and low levels were detected in the cells located between the foci. A subpopulation of mono- and multinucleated cells, peripheral to the cartilage nodules, expressed the muscle-specific intermediate filament desmin protein together with high levels of nestin protein. The proportion of nestin-expressing cells could be changed by addition of specific signalling molecules. Insulin-like growth factors I and II (IGF I and II) increased the number of nestin-positive cells, while basic fibroblast growth factor (FGF) reduced the number of nestin-expressing cells. Finally, we present evidence for a different subcellular localization of nestin protein and mRNA: the mRNA is predominantly located in the ends of the muscle cell, whereas the protein is found in the central region. Intracellular localization of nestin mRNA may constitute an additional level of regulation of the cytoskeleton during muscle development.

Animals↗
PubMed1996

Characterization of chondrogenesis in cells isolated from limb buds in mouse.

Micromass cultures of mesenchymal cells isolated from limb buds of 11.5-day-old mouse fetuses were used to study chondrogenesis. After 3 days of culture, dense cell aggregates were observed. They then were converted into macroscopically visible cartilage foci during the following 2-4 days. Comparison of 2-, 4- and 7-day-old cultures has shown that the cells first expressed collagen type I, then switched to collagen type II expression as shown by immunohistochemistry and in situ hybridization. At day 7, proteoglycans were synthesized centrally in the foci. At the same time, most cells expressed collagen type II, with the highest expression in the periphery of the aggregates. The oncogene c-fos and homeodomain protein FS-1 were found in the cells expressing collagen type II, indicating that these transcription factors may be involved in the regulation of cell differentiation. The expression of alkaline phosphatase was detected first in mature cartilage foci (day 4) and increased during culture. Early in culture, DNA-replicating cells were uniformly distributed. With differentiation, the proliferating cells were present predominantly between the aggregates and their total number became significantly reduced. Our results indicate that the process of chondrogenesis in micromass cultures of mesenchymal cells mimics the differentiation process occurring during fetal development in vivo and can be directly studied by in situ hybridization, immunohistochemical and histochemical methods.

Animals↗
PubMed1995

Inhibitory effects of basic fibroblast growth factor on chondrocyte differentiation.

The role of basic fibroblast growth factor (bFGF) and insulin-like growth factor I (IGF-I) in cartilage growth was studied in primary cultures of rat rib growth plate chondrocytes. Growth factors effects on expression of the proto-oncogene c-fos, DNA synthesis, differentiation, and morphological changes were analyzed by in situ hybridization, 3H-thymidine incorporation, and light and fluorescence microscopy. In serum-deprived cells, bFGF induced a transient expression of c-fos with a maximal effect 15-30 minutes after stimulation. After 24 h of culture it had a slightly lower stimulatory effect on DNA synthesis than IGF-I, but became a significantly more potent mitogen than IGF-I after 48 and 72 h. The stimulatory effect of bFGF on DNA synthesis coincided with a decrease in collagen type II and IGF-II expression. In contrast, IGF-I alone stimulated expression of these genes. In bFGF-treated cultures, cell morphology and the appearance of actin filaments was changed. Polygonal chondrocytes became elongated, fibroblast-like, and the smooth actin filaments were brush-like and disrupted. Addition of IGF-I reduced these changes without affecting c-fos expression induced by bFGF. Our results suggest that bFGF stimulates cell proliferation by preventing terminal differentiation of chondrocytes. This effect is mediated by induction of c-fos expression and a decrease in the steady-state levels of transcripts for collagen II and IGF-II.

Actins↗
PubMed1995

C-CAM expression in odontogenesis and tooth eruption.

The distribution of the cell adhesion molecule C-CAM was analyzed during tooth development, eruption and formation of the junctional epithelium in rat molars by immunohistochemistry and in situ hybridization. C-CAM could not be detected during odontogenesis until the late bell stage; then only the mRNA was demonstrated in the odontoblasts and ameloblasts. Prior to eruption, a local increase in C-CAM (mRNA and protein) was observed in the reduced enamel epithelium. During eruption, high C-CAM levels were seen in the fusion zone between the oral epithelium and the reduced enamel epithelium. In the adult rat, C-CAM remained strongly expressed in the junctional epithelium. Our study indicates that C-CAM may play a role in odontogenesis and during formation of the epithelial structures involved in tooth eruption and formation of the junctional epithelium.

Ameloblasts↗