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L H Margaritis

Publications and source records attributed to L H Margaritis.

At least 19 recordsLinked to original sources
PubMed2002

Implantation of MNNG crystals into a Triturus intact limb affects mitotic and labeling indices, regeneration rate, and morphogenesis in the contralateral, regenerating limb.

Experimental administration of chemical carcinogens to various mammals is highly effective in inducing malignant tumors. In contrast, treatment of regeneration-competent animals even with much higher doses of the same drugs only exceptionally leads to tumor-like growth. Usually, carcinogenic materials implanted or injected into a regenerating limb of urodele amphibia interfere with the regenerative process and frequently lead a). to growth retardation or arrest of regeneration, b). to development of a great variety of abnormal regenerates, and c). to generation of accessory, limb-like structures. Autonomous or experimental incidence of carcinogenesis is extremely low in animals endowed with strong regenerative capabilities. Of exceptional biological significance is the fact that such induced tumors usually regress spontaneously. This unique property of the regeneration-competent animals to resist carcinogenesis provides opportunities to compare non-cancerous alterations in the differentiated state of adult cells to those occurring in neoplasia. The mode of action of the chemical carcinogens on limb regeneration has not yet been clarified with certainty at the cellular and the molecular level. Several scientists claim that the above-mentioned effects might be attributed to local toxic influences of the drugs; therefore the present study was designed to investigate whether the administration of the carcinogen MNNG can affect cell proliferation, histogenesis, and morphogenesis at a region distant from the site of its implantation, even after a relatively long time period. To this end, 40 animals of the species Triturus cristatus had their right hindlimb surgically removed at the distal zeugopod. Then, a small microcrystal (approximately 5 micro g) of MNNG was inserted under the ventral aspect of the skin of the left tarsus in 20 of these animals (groups T and A; see below). Two months later, nine of the MNNG-treated animals were injected intraperitoneally with tritiated thymidine. After 2 h, six of these animals had their right hindlimb amputated at the distal zeugopod, whereas the rest were left to regenerate. The results were evaluated by camera lucida drawings, clearing in methyl benzoate, classical histology, and autoradiography. It was revealed that administration of MNNG at a somatic region (left hindlimb) reduces DNA synthesis and mitosis at a distant place (right hindlimb) even 2 months after MNNG implantation. Despite this, the rate of limb elongation is not substantially reduced. Classical histology revealed normal tissue structure throughout. All regenerated limbs displayed several teratogenic abnormalities.

Amputation, Surgical↗
PubMed2002

A novel case of haemoglobin H disease associated with clinical and morphological characteristics of congenital dyserythropoietic anaemia type I.

We report, for the first time, an unusual case of congenital anaemia with the clinical diagnosis of haemoglobin H disease complicated by morphological features at the light and electron microscopy level very similar to those of CDA-I. The red cell indices and the globin chain biosynthetic ratio were not characteristic of the defective haemoglobin genotype. The haematological, clinical and morphological data strongly suggest the novel coexistence of the two defects in a patient. The disease is characterised by a unique dyserythropoietic phenotype of diagnostic importance, which possibly brings new data regarding the reciprocal interaction between the two diseases, especially concerning a specific abnormality in globin chain synthesis in CDA-I, as previously suggested.

Adult↗
PubMed2001

Actin cytoskeleton reorganization of the apoptotic nurse cells during the late developmental stages of oogenesis in Dacus oleae.

In the present study, we demonstrate the actin cytoskeleton reorganization during nurse cells apoptosis of the olive fruit fly Dacus oleae. At the developmental stage 9A of oogenesis, the actin microfilaments are assembled in numerous ring canals and subcortically support all the nurse cells, as is shown by phalloidin-FITC staining. During the following stages, 9B and 10A, this structural pattern remains the same. The developmental stage 10B is characterized by actin microfilament rearrangement and formation of actin cables that are symmetrically organized around the nurse cell nuclei. At stage 11, when the dumping process begins, these actin cables seem to retain each nurse cell nucleus in the cell center, away from blocking the ring canals. The early stage 12 is characterized by an asynchronous nurse cell nuclear chromatin condensation, while at late stage 12 the actin cables become very thick, as adjacent ones overlap one another and traverse the disorganized apoptotic nurse cell nuclei that already have fragmented DNA, as is demonstrated by acridine orange staining and TUNEL assay. Finally, during stage 13, the apoptotic nuclear remnants are phagocytosed by the neighboring follicle cells. The data presented herein compared to previous reported results in Drosophila [Nezis et al., 2000: Eur J Cell Biol 79:610-620], demonstrate that actin cytoskeleton reorganization during nurse cell apoptosis is a developmentally regulated physiological mechanism, phylogenetically conserved in higher Dipteran.

Acridine Orange↗
PubMed2001

Differential sorting of constitutively co-secreted proteins in the ovarian follicle cells of Drosophila.

Conventional and freeze-fracture electron microscopy, immuno-electron microscopy of ovarian cryosections and confocal immunofluorescence were used to analyze the ovarian distribution of the major protein classes being secreted by the follicle cells during the vitellogenic and choriogenic stages of Drosophila oogenesis. Our results clearly demonstrated that at vitellogenic stages the follicle cells co-secrete constitutively vitelline membrane and yolk proteins that are either sorted into distinct secretory vesicles or they are segregated in different parts of bipartite vesicles by differential condensation. Following their exocytosis only the vitelline membrane proteins are incorporated into the forming vitelline membrane. The yolk proteins (along with their hemolymph circulating counterparts) diffuse through gaps amongst the incomplete vitelline membrane and are internalized through endocytosis by the oocyte where they are finally stored into modified lysosomes referred to as alpha-yolk granules. The unexpected immunolocalization of vitelline membrane antigens in the associated body of the alpha-yolk granules may indicate that this structure is a transient repository for the proteins being internalized into the oocyte along with the yolk proteins. In the early choriogenic follicle cells the vitelline membrane and early chorion proteins were found to be co-secreted and to be evenly intermixed into the same secretory vesicles. These findings illuminate new details concerning the follicle cells secretory and oocyte endocytic pathways and provide for the first time evidence for condensation-mediated sorting of constitutively secreted proteins in Drosophila.

Animals↗
PubMed2000

A mechanism for action of oscillating electric fields on cells.

The biological effects of electromagnetic fields have seriously concerned the scientific community and the public as well in the past decades as more and more evidence has accumulated about the hazardous consequences of so-called "electromagnetic pollution." This theoretical model is based on the simple hypothesis that an oscillating external electric field will exert an oscillating force to each of the free ions that exist on both sides of all plasma membranes and that can move across the membranes through transmembrane proteins. This external oscillating force will cause a forced vibration of each free ion. When the amplitude of the ions' forced vibration transcends some critical value, the oscillating ions can give a false signal for opening or closing channels that are voltage gated (or even mechanically gated), in this way disordering the electrochemical balance of the plasma membrane and consequently the whole cell function.

Cell Membrane↗
PubMed2000

Diversity of peripheral blood mononuclear cells as revealed by a novel multiple microgel "comet assay".

Multiple microgel comet assay (MMCA) is a metho-dological adaptation of the single-cell gel electrophoresis assay in which we have introduced the use of standard agarose plug molds in an attempt to improve and expand the applications of the assay. We focused on the study of the heterogeneity of peripheral blood mononuclear cells (PBMC) at the level of the basal single-strand breakage and the DNA damage induction caused by ionizing radiation. Differences among subpopulations were also investigated at the level of chromatin organization and methylation after NotI digestion of microgel-embedded cells. In parallel experiments, the NotI-digested nucleoids were also analyzed with the use of pulsed-field gel electrophoresis (PFGE) and the DNA migration patterns were compared with the corresponding patterns from the MMCA. Significant heterogeneity in the distribution of the oxidative DNA damage, as well as intracellular variations in the NotI digestion patterns were observed in the cell population of PBMC. The combined use of both the comet assay and PFGE provides a useful model for analysis of variation in DNA damage in individual cells as well as information on size of DNA fragments.

Chromatin↗
PubMed2000

Stage-specific apoptotic patterns during Drosophila oogenesis.

In the present study we demonstrate the existence of two apoptotic patterns in Drosophila nurse cells during oogenesis. One is developmentally regulated and normally occurs at stage 12 and the other is stage-specific and is sporadically observed at stages 7 and 8 of abnormally developed follicles. The apoptotic manifestation of the first pattern begins at stage 11 and is marked by a perinuclear rearrangement of the actin cytoskeleton and the development of extensive lobes and engulfments of the nurse cell nuclei located proximal to the oocyte. Consequently, at late stage 12 (12C), half of the nurse cell nuclei exhibit condensed chromatin, while at late stage 13 all the nuclei have fragmented DNA, as it is clearly shown by TUNEL assay. Finally, the apoptotic vesicles that are formed during stage 13, are phagocytosed by the neighboring follicle cells and at stage 14 the nurse cell nuclear remnants can be easily detected within the adjacent follicle cell phagosomes. In the second sporadic apoptotic pattern, all the nurse cell nuclei are highly condensed with fragmented DNA, accompanied by a completely disorganized actin cytoskeleton. When we induced apoptosis in Drosophila follicles through an etoposide and staurosporine in vitro treatment, we observed a similar pattern of stage-specific cell death at stages 7 and 8. These observations suggest a possible protective mechanism throughout Drosophila oogenesis that results in apoptosis of abnormal, damaged or spontaneously mutated follicles before they reach maturity.

Actins↗
PubMed1999

Biochemical and immunocytochemical analysis of vitellogenesis in the olive fruit fly Dacus (bactrocera) oleae (Diptera: Tephritidae).

Synthesis and selective accumulation of the major yolk proteins in the developing oocytes of the species Dacus oleae (Diptera: Tephritidae) was studied biochemically and by immunoelectron microscopy. In the hemolymph of adult females, two yolk proteins precursors (or vitellogenins) have been detected. They each exhibit a similar molecular weight and isoelectric point to their respective mature yolk proteins (or vitellins), while electrophoretic analysis of their synthetic profile shows that their levels in the hemolymph increase rapidly during development. Immunogold electron microscopy of ovarian sections, revealed that the hemolymph vitellogenins reach the oocyte through enlarged inter-follicular spaces and demonstrated vitellogenin synthesis by the follicle cells of the vitellogenic follicles. The newly synthesized vitellogenins follow a distinct secretory pathway into these cells as compared to other components being synthesized at the same time (e.g. the vitelline envelope proteins), since they were found in secretory vesicles that appeared to be differentiated from those destined to participate in the vitelline envelope. The vitellogenin-containing vesicles exocytose their contents directionally into the follicle cell/vitelline envelope boundary, and subsequently the vitellogenins diffuse among the gaps of the forming vitelline envelope and reach the oocyte plasma membrane. Their internalization by the oocyte includes the formation of an endocytic complex consisting of coated pits, coated vesicles, endosomes, transitional yolk bodies, and finally mature yolk bodies, in which the storage of the vitellins and other yolk proteins occur. These results are discussed in relation to data obtained from other Dipteran species.

Animals↗
PubMed1999

Mass determination of the unit cell of the innermost chorionic layer in Drosophilidae by scanning transmission electron microscopy.

The innermost chorionic layer (ICL) in eggshells of Drosophila melanogaster is a naturally occurring patchwork of thin three-dimensional crystalline plates located between the inner endochorion and the vitelline envelope. The mass-per-unit area of the ICL has been measured from scanning transmission electron microscope images of isolated unstained material and it was possible to distinguish up to four layers with the majority of the crystalline sheets being one to three layers thick. Taking into account the unit cell areas for the different crystals, we have estimated the mean ICL subunit sizes to be 36 kDa for Drosophila melanogaster, 35 kDa for Drosophila auraria, and 33 kDa for Drosophila teissieri. The results suggest that the three different Drosophilidae species have very similar average subunit masses.

Animals↗
PubMed1999

CD4 cytotoxic and dendritic cells in the immunopathologic lesion of Sjögren's syndrome.

The existence of CD4+ T lymphocytes with cytotoxic activity in minor salivary gland (MSG) biopsies from Sjögren's syndrome (SS) patients was investigated using in situ double immunohistochemistry technique. The presence of dendritic cells (DC) in SS lesions was examined by using single and double immunohistochemistry methods and a panel of different MoAbs to specific cell surface markers (i.e. CD3, CD11c, DRC). Furthermore, the ultrastructural morphology of DC was characterized by electron microscopy (EM). Immunogold labelling technique using the DRC surface marker was also applied. Finally, we investigated the existence of germinal centres (GC) in the salivary gland lesions of SS patients. Seven patients with primary SS and five patients with non-specific sialadenitis were the subjects of this study. Our results indicate the existence of a CD4+ cytotoxic cell population that utilizes perforin-mediated cell destructions as they expressed perforin mRNA. Quantitative analysis of these cells revealed that they comprised approximately 20% of the existing T lymphocytes. We also identified a population of CD4+ T cells that expressed the CD11c activation marker. Furthermore, we observed a distinct cell subtype which expressed the DRC cell surface marker. These cells had the characteristic ultrastructural morphology of DC and were DRC+ when examined by immunoelectron microscopy. Finally, the formation of GC structures in the histopathologic lesions of the salivary glands was observed. The above findings indicate that both CD4+ cytotoxic T lymphocytes (CTL) and DC may be involved in the initiation and perpetuation of SS pathogenesis. Moreover, the formation of GC in the lesions reveals a possible mechanism for in situ differentiation and proliferation of activated B lymphocytes.

Biopsy↗
PubMed1999

Origin of renewed spinal ganglia during tail regeneration in urodeles.

The source of the cells which form the spinal ganglia within the regenerating urodele tail is not yet indisputably known. Classical and modern experimental approaches trace the spinal cord as the most probable source. The aim of the present study was to further investigate this item by conventional histology, counting of mitotic figures, and estimating the labeling index. The main results can be summarized as follows: (a) The regenerated part of the tail contained only two bilaterally asymmetrical pairs of ganglia, with respect to the rostrocaudal (anterior-posterior) axis. (b) The anterior ganglia were slightly differentiated and appropriately localized; therefore, analysis was performed mainly in the posterior, still developing ganglia. (c) An anatomical continuation between the ventrolateral side of the regenerated spinal cord and the laterally forming spinal ganglion was noticed. There was some indication that many cells migrated out from the spinal cord towards the spinal ganglion, through the ventral root. (d) The mitotic and the labeling index along the regenerated spinal cord exhibited individual peaks near the level of each developing ganglion. The last two observations corroborate and reinforce the prevailing view that the progenitors of the spinal ganglion cells which are formed in the regenerating tail are of spinal cord origin.

Animals↗
PubMed1998

Immunolocalization of the temporally "early" secreted major structural chorion proteins, Dvs38 and Dvs36, in the eggshell layers and regions of Drosophila virilis.

We have shown by means of conventional electron microscopy that the eggshell of Drosophila virilis at the main body of the laid egg consists of the vitelline membrane and the multilayered chorion, which includes the wax layer, the innermost chorionic layer, the endochorion, and the exochorion, while several specialized regions of the eggshell are seen across the anterior-posterior axis of the egg. Biochemical analysis revealed the existence of six quantitatively enriched chorion proteins. Among them, Dvs38 and Dvs36 are synthesized when the innermost chorionic layer and the endochorion are assembled. Immunogold electron microscopy has shown that these two proteins are incorporated in the morphologically complete vitelline membrane apparently through an intercalation process and represent structural components of the endochorion in all the specialized regions of the eggshell. Additionally, by cytochemical means, the enzyme eggshell peroxidase, which is synthesized in parallel with Dvs38 and Dvs36, has been identified as a structural component of the innermost chorionic layer and the endochorion. These findings suggest a complex protein-protein recognition pattern during the formation of the eggshell since the cosecretion of its components (i.e., Dvs38, Dvs36 chorion proteins and eggshell peroxidase) does not recommend their colocalization in the eggshell sublayers and the timing of their synthesis is not related to their final position on the eggshell (i.e., the identification of Dvs38 and Dvs36 chorion proteins as vitelline membrane components).

Animals↗
PubMed1998

The formation of the functional chorion structure of Drosophila virilis involves inercalation of the "middle" and "late" major chorion proteins: a general model for chorion assembly in Drosophilidae.

We have shown by in vitro development of staged follicles in the presence of tritiated proline, as well as by immunoblot analysis, that the Dvs19 and DVS15 chorion proteins of Drosophila virilis are synthesized by the follicular epithelium and are incorporated in the chorion at the terminal stages of choriogenesis. Light microscopy immunolocalization has revealed that these two proteins represent structural components of the endochorion in all the specialized regions of the eggshell, while, contrary to their temporal pattern of secretion, electron microscopic immunolocalization has shown that these two proteins participate in endochorionic structures formed before the onset of their synthesis, thus suggesting an intercalation process during chorion formation. Furthermore, double immunolocalizations demonstrated that cosecreted proteins do not coexist in all secretory vesicles of the follicle cells, indicating that probably the final association of these chorionic components occurs extracellularly. On the basis of these data and those presented in the accompanying paper [Trougakos, I.P., and Margaritis, L.H. (1998) Immunolocalization of the temporally "early" secreted major structural chorion proteins, Dvs38 and Dvs36, in the eggshell layers and regions of Drosophila virilis, J. Struct. Biol. 123, 000-000.], a model for chorion assembly in Drosophilidae is proposed.

Animals↗
PubMed1998

Effects of gamma rays on the stability and size of DNA.

The effects of gamma radiation on the stability and size of mammalian DNA were studied by using thermal transition spectrophotometry and pulsed-field and standard agarose gel electrophoresis. The experiments were performed using deproteinized calf thymus DNA in buffered deaerated aqueous solutions. A dual dose response was observed: a tendency for increased helix stability at "low" doses (0-4 Gy) accompanied by a high tendency of the DNA molecules to interact, forming larger molecules, followed by a gradual increase of degradation and helix instability at higher doses. The results reported here for the low-dose region are consistent with the hypothesis of inter- and intramolecular interactions of covalent character (crosslinking) in irradiated DNA molecules.

Animals↗
PubMed1996

The eggshell of Drosophila melanogaster: IX. Synthesis and morphogenesis of the innermost chorionic layer.

Synthesis and morphogenesis of the innermost chorionic layer (ICL) was investigated by conventional EM methods, freeze-fracturing, tissue culture in Robb's medium, and EM autoradiography. Both autoradiography and fine structure results have shown that ICL-components are secreted prior to other chorion proteins. Their secretion starts on stage 12a but the first layer of ICL molecules is visible at stage 12b. Its thickness is gradually increased during the next stages, taking first, a bilaminar form along with the inner endochorion. Later, at the end of choriogenesis, ICL is detached from the endochorion and takes its final thickness and configuration, consisting of a 3-dimensional crystal, about 40 nm thick. The isolated ICL in conditions of air water interface is a monolayer crystal 10 nm thick. Studies on chorion mutants showed that the amount of protein secreted by the follicle cells is independent to the process of crystallisation. These data show how a proteinaceous extracellular substance is gradually assembled to form a 3-D crystal and how it can be organised to perform functions such as the physiological resistance of the insect eggs against water loss or water uptake, whenever they are laid on substrates with extreme environmental conditions. These functions are performed by ICL in conjunction with the underlying wax layer.

Animals↗
PubMed1993

The eggshell of Drosophila melanogaster. VIII. Morphogenesis of the wax layer during oogenesis.

Utilizing freeze-fracturing and conventional electron microscopy methods, we have studied the details of morphogenesis and construction of the wax layer envelope from Oregon R and mutants of Drosophila melanogaster eggs during oogenesis. The wax layer is synthesized and secreted by the follicular cells in the form of lipid vesicles during stage 10b. During secretion (stages 10b, 11 and 12) the lipid vesicles are accumulated on the vitelline membrane surface and become flat. At the late stages of choriogenesis (stages 13, 14) the lipid vesicles are compressed tightly between the vitelline membrane and the other already constructed eggshell layers, so the wax layer becomes very thin and is hardly seen in cross-fractured views.

Animals↗
PubMed1991

Structure and morphogenesis of the eggshell and micropylar apparatus in the olive fly, Dacus oleae (Diptera: Tephritidae).

The egg of the olive fly, Dacus oleae (Diptera, Tephritidae), is laid inside olives and the larva eventually destroys the fruit. The oocyte is surrounded by several distinct layers which are produced during choriogenesis. The chorion covering the main body of the egg outside of the vitelline membrane includes a "wax" layer, an innermost chorionic layer, an endochorion consisting of inner and outer layers separated by pillars and cavities similar to their counterparts in Drosophila melanogaster, as well as inner and outer exochorionic layers. The anterior pole is shaped like an inverted cup, which is chiefly hollow around its base and has very large openings communicating with the environment. Holes through the surface of the endochorion result from deposition of endochorionic substance around follicular cell microvilli. An opening at the apex of the cup provides an entrance for sperm entering the micropylar canal, which traverses the endochorion and continues into a "pocket" in a thickened vitelline protrusion. The micropylar canal is formed by deposition of endochorion and vitelline membrane around an elongated pair of follicular cell extensions. These extensions later degenerate and leave an empty canal about 5 microns in diameter and the narrower pocket about 1 micron in diameter. Respiration is thought to be facilitated by openings at the base of the anterior pole as well as by openings through the "plastron" around the main body of the shell.

Animals↗