Effects of iron-, manganese-, or magnesium-deficiency on the growth and morphology of Euglena gracilis

Iron-, manganese-, or magnesium-deficiency has been induced in Euglena gracilis. Each arrests cell proliferation, decreases the intracellular content of the deficient metal, and increases that of several other metals. Light and electron microscopy of stationary phase cells reveal that Fe-deficient (-Fe) cells are similar in size and shape to control organisms. Magnesium-deficient (-Mg) cells, however, are larger, and approximately 14% are multilobed, containing 2 to 12 lobes of equal size emanating from a central region. Individual (-Mg) cells and each lobe of multilobed cells contain a single nucleus. Manganese-deficient (-Mn) organisms are morphologically more heterogeneous than (-Fe) or (-Mg) cells. Most are spherical and larger than controls. Approximately 15% are multilobed but, unlike (-Mg) cells, contain lobes of unequal size with either zero, one, or several nuclei present in each. Nuclei of (-Mn) cells differ in size and shape from those of control, (-Fe), or (-Mg) cells. All three deficient cell types accumulate large quantities of paramylon. Other cytoplasmic structures, however, appear normal. Addition of Fe, Mn, or Mg to the respective deficient stationary phase cultures reverses growth arrest and restores normal morphology. The results suggest that Fe-, Mn-, and Mg-deficiencies affect different stages of the E. gracilis cell cycle.     

Fibroblast Growth Factor May Regulate the Initiation of Oocyte Growth in the Developing Ovary of the Medaka, Oryzias latipes

The distribution of fibroblast growth factor (FGF) was investigated in developing and matured ovaries of the medaka, Oryzias latipes. In the fry, FGF localized in the cytoplasmic region of all oocytes in the ovary at the pre-vitellogenic stage. Before the initiation of vitellogenesis, it disappeared in the cytoplasmic region and newly appeared around each oocyte, and then it localized around all oocytes in the ovary at the vitellogenic stage. Interestingly, the change in FGF distribution was orderly occurring from the posterior to anterior region of the ovary. In the adult, FGF was detected by immunofluorescence staining around the oocytes. These results suggest that FGF plays a significant role in the initiation of oocyte development through follicle cells, and the expression of FGF is rigidly regulated in the developing ovary of O. latipes.     

Expression and localization of intestinal 15 kDa protein in the rat

Rat intestinal 15 kDa protein (I-15P) is highly homologous to porcine gastrotropin. We studied the occurrence, distribution and subcellular localization of I-15P in the entire rat body, using the immunocytochemistry to localize protein andin situ hybridization to localize mRNA. Both techniques demonstrated the expression of I-15P in the enterocytes of ileum, luteal cells of ovary and a subpopulation of steroid-endocrine cells of adrenal gland. Immuno-electron microscopy further demonstrated that I-15P is localized in both the cytoplasmic and nuclear matrix regions of these cells. The present results suggest roles of I-15P not only in the transport of bile salts but also in the metabolisms of certain steroid hormones.     

Assorbimento dopo colorazione al Fast-green di cellule della caliptra di Allium cepa localizzate in differenti regioni istologiche

Abstract

Fast-green absorption of cells localized in different regions of Allium cepa root cap.—The nuclear and cytoplasmic protein content of two cell population (C2 = columella cells; P2 = peripheral cells) has been determined by Barr and Stroud cytophotometer after Fast-green staining (after DNA extraction and without DNA extraction). The collected data seem indicate that: 1) histone content is higher in nuclei of C2 cells than in nuclei of P2 ones; 2) acid proteins content is higher in nuclei of P2 cells than in those of C2 ones; 3) the cytoplasmic proteins of the two cell populations are quite different in their Fast-green reactivity.     

Ovary cords organization in Hirudo troctina Johnson, 1816 and Limnatis nilotica (Savigny, 1822) (Clitellata,Hirudinea)

In both examined species of Hirudinea there are paired spheroid ovisacs, and within each ovisac two convoluted ovary cords occur. The morphology of the cords is characteristic: their apical end is club-shaped, the central part is narrow and may contain developing oocytes, whereas the basal end of the cord is irregularly shaped and composed of degenerating cells. The ovary cords are built of somatic and germ-line cells; the latter are united into syncytial cysts. Each germ cell in such a cyst has only one stable cytoplasmic bridge connecting it to the central anuclear cytoplasmic mass, the cytophore. Initially all germ-line cells in a given cyst are morphologically identical, then the fates of cells diversify. Most of them become nurse cells and eventually degenerate; the rest continue meiosis, gather macromolecules, cell organelles and nutritive material and become oocytes. The oogenesis found in the species studied should be regarded as meroistic. Previtellogenic oocytes protrude from the cord into the ovisac lumen, whereas the vitellogenic ones float freely in the ovisac lumen. The somatic cells found in the ovary cords are: follicular cells which form the envelope of the cord and are also found among germ cells inside the cord, and one, huge apical cell that always is located at the top of the club-shaped end of the ovary cord. The apical cell has several characteristic features, e.g., it forms long cytoplasmic projections filled with intermediate filaments and it is connected to the neighbouring cells (both somatic and germ-line) via hemidesmosomes. We suggest that the apical cell forms the niche for maintaining germ and somatic stem cells. Generally, the organization of the ovary cords found in both studied species is broadly similar to those described in other hirudiniform leeches studied to date.     

Immunohistochemical localization of androgen receptor and aromatase in the ovary of the pregnant pig

The distribution of androgen receptor (AR) and cytochrome P450 aromatase was investigated in paraffin sections of pregnant pig ovary. Ovarian follicles and corpora lutea were isolated from ovaries obtained on Days 10, 18, 32, 71 and 90 post coitum (p.c.). Androgen receptor was localized in the nuclei of granulosa cells of follicles of various sizes. In addition, some follicles demonstrated staining in the nuclei of theca interna cells. Stroma cells also exhibited a positive immunostaining. At early and mid pregnancy (up to Day 71) AR was expressed in the nuclei of luteal cells. Corpora lutea of Day 71 showed mainly cytoplasmic staining while on Day 90 almost all luteal cells showed staining exclusively in the cytoplasm. Immuno-staining for the presence of cytochrome P450 aromatase was very faint in all investigated ovarian structures. The results could suggest that the process of androgen aromatization plays a negligible role in the ovary of the pregnant pig.     

Ultrastructure and X-Ray Microanalysis of Blood Cells of Ascidia malaca

Ultrastructural studies of Ascidia malaca blood reveal particular cell types, which are characterized by a polymorphism in the organization of the electron dense cytoplasmic material. A new pathway of morula cell differentiation is suggested. X-ray microanalysis shows that vanadium is localized in vacuolated, granular and morular cells. Iron, which is accumulated by this species to a lesser degree than vanadium, is found in vacuolated amebocytes and, together with vanadium, in granular cells. Our results are discussed in the light of the relations between selective metal absorption in blood cells and their specialization and differentiation.     

Subcellular localization of copper in the root cells of Allium sativum by electron energy loss spectroscopy (EELS)

Ultrastructural investigation of the root cells of Allium sativum L. exposed to three different concentrations of Cu (1, 10 and 100 microM) for 9 days was carried out using transmission electron microscopy (TEM) and electron energy loss spectroscopy (EELS). The results presented here indicate that excess Cu induces ultrastructural changes such as strong vacuolization, condensed nuclear chromatin, decreased endoplasmic reticulum (ER) and ribosome and serious plasmolysis. EELS analysis indicated that electron dense granules containing Cu appeared in the cells after Cu treatment. The vacuoles of the root tip cells were the main Cu-accumulation site. Small amounts of copper were also localized to cytoplasmic vesicles or cell walls of cortical cells. The results of the present investigation have significant importance in further understanding the mechanisms of absorption, transportation and accumulation of heavy metals in plants grown in polluted soil.     

Structure and ultrastructure of the ovary in the South American Veturius sinuatus (Eschscholtz) (Coleoptera,Passalidae)

The morphoanatomy of the ovary in Veturius sinuatus (Eschscholtz) was studied by light and transmission electron microscopy. Data from the female gonad of this species provide more extended and precise knowledge regarding the organization of the ovary in Passalidae. Ovaries are composed of a pair of long telotrophic meroistic ovarioles, with some differences compared to the bauplan of this ovary type in Polyphaga (Coleoptera). The terminal filament has an enlarged proximal region with irregularly shaped cells in apparent degeneration process embedded in a membranous system. Globular structures with amorphous content associated with interstitial cells are distributed throughout the tropharium. Trophocytes develop with the reduction of the plasma membrane between sibling nurse cells of each cluster. Previtellogenic oocytes have an irregular shape and various cytoplasmic prolongations. As oogenesis advances, a single prolongation in the anterior part of the oocyte extends to the tropharium. The ovary structure is comparable to that found in other American species of passalids, and further, the conformation of the terminal filament could be a plesiomorphic character of the family.     

Factors affecting growth of the spiny lobsters Panulirus gracilis and Panulirus inflatus (Decapoda: Palinuridae) in Guerrero, México

The effects of sex, injuries, season and site on the growth of the spiny lobsters Panulirus gracilis, and P. inflatus, were studied through mark-recapture techniques in two sites with different ecological characteristics on the coast of Guerrero, México. Panulirus gracilis occurred in both sites, whereas P. inflatus occurred only in one site. All recaptured individuals were adults. Both species had similar intermolt periods, but P. gracilis had significantly higher growth rates (mm carapace length week-1) than P. inflatus as a result of a larger molt increment. Growth rates of males were higher than those of females in both species owing to larger molt increments and shorter intermolt periods in males. Injuries had no effect on growth rates in either species. Individuals of P. gracilis grew faster in site 1 than in site 2. Therefore, the effect of season on growth of P. gracilis was analyzed separately in each site. In site 2, growth rates of P. gracilis were similar in summer and in winter, whereas in site 1 both species had higher growth rates in winter than in summer. This could be due to spatial differences in processes related to changes in population density and food resources, which were documented in previous works. The overall results show that P. gracilis grows faster than P. inflatus, and that growth rates of both species are highly variable and are affected by environmental factors such as site and season, which should be taken into account when attempting to produce population growth curves for each species.     

Manganese-dependent ribonucleotide reductase from Propionibacterium freudenreichii subsp. Shermanii: partial purification, characteristics and role in DNA biosynthesis

Like Lactobacillus leichmanii, Rhizobium meliloti, and Euglena gracilis, P. freudenreichii implicates cobalamin in DNA anabolism via adenosylcobalamin-dependent ribonucleotide reductase. However, in the absence of corrinoids, P. freudenreichii is able to synthesize DNA with the involvement of an alternative ribonucleotide reductase, which is independent of adenosylcobalamin. This enzyme is localized in both the cytoplasm (80% of activity) and the cytoplasmic membrane (20% of activity), being loosely bound to the latter. Experiments with crude ribonucleotide reductase isolated from extracts of corrinoid-deficient cells showed that manganese specifically stimulates this enzyme and that it is composed of two protein subunits, a feature that is typical of all metal-containing reductases activated by molecular oxygen. Low concentrations of manganese ions enhanced DNA synthesis in corrinoid-deficient manganese-limited cells. This effect was prevented by the addition of 80 mM hydroxyurea, a specific inhibitor of metal-containing aerobic ribonucleotide reductases. It was concluded that, in adenosylcobalamin-deficient P. freudenreichii cells, DNA synthesis is provided with deoxyribosyl precursors through the functioning of manganese-dependent aerobic ribonucleotide reductase composed of two subunits.     

五种丛生状棕榈植物叶热值的月变化研究

对5种丛生状棕榈植物一欧洲棕、细叶棕竹、小箬棕、玲珑椰子、缨络椰子的叶片热值和灰分含量的月变化进行了研究,结果表明,5种植物具有较高的灰分含量,但灰分含量的月变化趋势各不相同;欧洲棕、细叶棕竹、小箬棕3种植物干重热值的月变化趋势有些差异,同属的缨络椰子、玲珑椰子干重热值的月变化趋势基本相似;缨络椰子、小箬棕的干重热值与灰分含量具有显著的线性相关(P<0．05),而其余3种植物的干重热值与灰分含量相关性不显著(P>0．05);从去灰分热值的月变化来看,细叶棕竹与小箬棕,缨络椰子与玲珑椰子的月变化趋势相似。     

The Protozoan Parasite Toxoplasma gondii Targets Proteins to Dense Granules and the Vacuolar Space Using Both Conserved and Unusual Mechanisms

All known proteins that accumulate in the vacuolar space surrounding the obligate intracellular protozoan parasite Toxoplasma gondii are derived from parasite dense granules. To determine if constitutive secretory vesicles could also mediate delivery to the vacuolar space, T. gondii was stably transfected with soluble Escherichia coli alkaline phosphatase and E. coli β-lactamase. Surprisingly, both foreign secretory reporters were delivered quantitatively into parasite dense granules and efficiently secreted into the vacuolar space. Addition of a glycosylphosphatidylinositol membrane anchor rerouted alkaline phosphatase to the parasite surface. Alkaline phosphatase fused to the transmembrane domain and cytoplasmic tail from the endogenous dense granule protein GRA4 localized to dense granules. The protein was secreted into a tuboreticular network in the vacuolar space, in a fashion dependent upon the cytoplasmic tail, but not upon a tyrosine-based motif within the tail. Alkaline phosphatase fused to the vesicular stomatitis virus G protein transmembrane domain and cytoplasmic tail localized primarily to the Golgi, although staining of dense granules and the intravacuolar network was also detected; truncating the cytoplasmic tail decreased Golgi staining and increased delivery to dense granules but blocked delivery to the intravacuolar network. Targeting of secreted proteins to T. gondii dense granules and the plasma membrane uses general mechanisms identified in higher eukaryotic cells but is simplified and exaggerated in scope, while targeting of secreted proteins beyond the boundaries of the parasite involves unusual sorting events.     

Dirofilaria immitis: ultrastructural aspects of oocyte development and zygote formation.

Ultrastructural observations of the ovary and uterus of Dirofilaria immitis reveal some characteristics of oogonia, oocytes, and uterine sperm. Oogonia are confined to the distal portion of the ovary including a blind tip, where a morphologically distinct terminal cap cell was not observed. These cells contain a nucleus with a nucleolus, numerous dense bodies, scanty ribosomes, lipid droplets, and an occasional mitochondrion. Endoplasmic reticulum is lacking and Golgi complexes were observed only in fully grown oogonia. Primary oocytes located in the middle portion of the ovary are large, elongate, and have a complete set of organelles including many small mitochondria, fragmentary endoplasmic reticulum, ribosomes, Golgi complexes, and very few dense bodies. These cells are arranged into many rosettes about central cytoplasmic masses, the rachises, to which they maintain cytoplasmic continuity by pseudopodlike processes. The rachises contain no organelle except a few dense granules and are bound by winding membranes. Oocytes from the proximal portion differ from those of the middle portion of the ovary in their larger size, round shape, absence of many organelles, presence of small dense granules, and lacking a rachis. Dense bodies are specific to the oogonia and exhibit DNase susceptibility and a positive reaction for a mitochondrial enzyme. These findings together with their decreased number and a concomitant increase of mitochondria in the oocytes suggest a relationship between these bodies and mitochondria.Uterine sperm of D. immitis are of the amoeboid type and contain several chromatin masses without a nuclear envelope, many mitochondria, and specialized membranous organelles referred to as mesosomelike vesicles. The vesicles are probably originated from the sperm plasma membrane. Upon fertilization, the entire spermatozoon penetrates the oocyte and its contents are gradually dissolved in the ooplasm with a simultaneous appearance of large numbers of ribosomes at the site of dissolution. Ribosomes were later found in the nucleus. A pronucleus was not observed. These findings are basically in agreement with those described for Ascaris but differ in the morphologic features and number of rachises, presence of dense bodies, absence of refringent granules in the oocytes and the absence of a refringent body and presence of several chromatin masses in the sperm.     

Ovulation and egg segregation in the tunic of a colonial ascidian,Diplosoma listerianum (Tunicata,Ascidiacea)

Summary The process of egg segregation in the tunic of the ovoviviparous ascidian Diplosoma listerianum was studied by light and electron microscopy. One egg at a time was seen to mature in each zooid. The eggs had large yolk and grew on the ovary wall enveloped in four layers: (1) outer follicle cells (OFC), long and rich in RER (rough endoplasmic reticulum) and with dense granules in the Golgi region; (2) flat inner follicle cells (IFC); (3) a loosely fibrillar vitelline coat (VC); (4) test cells encased on the egg surface. The growing egg protrudes from the ovary wall and presses on the contiguous epidermis. Granulocytes enter the space between the epidermis and the egg and insinuate cytoplasmic protrusions, disrupting the continuity of the OFC layer. At ovulation, OFC and IFC are discharged and form a post-ovulatory follicle (corpus luteum). The epidermis shrinks and closes, possibly by activation of microfilaments, causing the egg to be completely surrounded by the tunic. In the zooid, the wound caused by the passage of the egg is repaired both by contraction of the epidermis and by phagocytic activity. Altered spermatozoans are found in phagocytosing cells in the lumen of the ovary. These are presumably remnants of those which entered to fertilize the egg before segregation.     

FINE STRUCTURAL AND RADIOAUTOGRAPHIC OBSERVATIONS ON DENSE PERINUCLEAR CYTOPLASMIC MATERIAL IN TADPOLE OOCYTES

Dense fibrous material is first seen in association with mitochondria in tadpole oogonia but is most prominent in oocytes during the extended first meiotic prophase when it aggregates into dense bodies in the perinuclear cytoplasm. The origin of this material has been attributed to 350-A nuclear granules which form cytoplasmic streamers of fibrous material upon passage through nuclear pores. This has commonly been interpreted as the transfer of ribonucleoprotein to the cytoplasm for storage. However, cytochemical reactions for nucleic acids have indicated an absence of detectable RNA in this dense material, and the results of radioautographic studies with labeled uridine, thymidine, or actinomycin D argue against the presence of nucleic acids. When sites of incorporation of tritiated amino acids were radioautographically localized, an appreciable number of silver grains were present over the dense bodies. Uptake of certain amino acids occurs fairly promptly but the degree of labeling levels off after about 6 hr, suggesting a rapid turnover of the material in the dense bodies. Attention is drawn to the similarity of the dense bodies to structures present in germ cells of a number of other species, and possible functions of the dense bodies in germ cell differentiation are considered.     

Immunohistochemical evidence identifying the site of androgen production in the ovary of the protogynous grouper <Emphasis Type="Italic">Epinephelus merra</Emphasis>

Androgen plays an important role in the developing ovaries of female fish. However, little is known regarding either the sites of production of androgen or its functional roles. In the present study, we investigated immunohistochemically the localization of cholesterol-side-chain-cleavage (P450scc) and cytochrome P45011-hydroxylase (P45011) with antibodies P450scc and P45011 in the ovary of the female honeycomb grouper Epinephelus merra during its reproductive cycle. Clusters of strongly immunopositive cells, with 100–1000 cells in each cluster, against both P450scc and P45011, were observed throughout the annual reproductive cycle in tissue near blood vessels in the tunica ovary surrounding the outer periphery of the ovary. The ultrastructural characteristics of these cells showed that they were steroid-producing cells. In contrast, immunopositive cells against P450scc but not against P45011 were localized in the theca layer surrounding the outer periphery of oocytes. These results suggest that two distinct steroid biosynthesis sites exist in the ovary and that cells at the two sites differ functionally. The only cells that biosynthesize 11-ketotestosterone are found in clusters in the vicinity of blood vessels; they possibly play a physiological role in oocyte growth and gonadal restructuring during the sex change of individuals of this species. CREST, JST (Japan Science and Technology Corporation), and the Takeda Science Foundation provided grants supporting this work. This study was also partially supported by a grant for the 21st century COE project entitled, The Comprehensive Analyses of Biodiversity in Coral Reef and Island Ecosystems in Asian and Pacific Regions. We also gratefully acknowledge a Japanese Government (Monbukagakusho) Scholarship.