Coherent phase microscopy: a new method of identification of intracellular structures based on their optical and morphometric parameters

A new method for the identification of intracellular structures of a living cell and obtaining the quantitative parameters characterizing these structures by means of coherent phase microscopy is proposed. The method is based on the analysis of the histogram of a cell phase image and its decomposition by phase height levels. In the spherical and cylindrical approximation of the cell, the method makes it possible to separate the contributions of phase-contrast intracellular structures to the integral refractive index of the whole cell. The calculation of refractive indices of intracellular structures is illustrated on a two-component model of a spherical cell. The possibility of determining the refractive indices of cellular organelles was shown by an example of cyanobacterium Anabaena variabilis ATCC 29413 cells and Cancer mammae breast cancer cells. Three most contrast cellular structures in the phase images of the cells were identified, and their refractive indices were determined. For Anabaena variabilis cells, these structures were the cell wall ( = 1.39), tylakoids ( = 1.47), and the nucleoid ( = 1.58); for breast cancer cells, these were the cytoplasm ( = 1.34), the nucleus ( = 1.36), and the nucleolus ( = 1.44). Using this method is possible to identify phase-contrast intracellular structures and calculate their refractive indices. This enables one to follow morphofunctional changes of not only the whole cell but also its individual organelles.     

Building and degradation of secondary cell walls: are there common patterns of lamellar assembly of cellulose microfibrils and cell wall delamination?

During cell wall formation and degradation, it is possible to detect cellulose microfibrils assembled into thicker and thinner lamellar structures, respectively, following inverse parallel patterns. The aim of this study was to analyse such patterns of microfibril aggregation and cell wall delamination. The thickness of microfibrils and lamellae was measured on digital images of both growing and degrading cell walls viewed by means of transmission electron microscopy. To objectively detect, measure and classify microfibrils and lamellae into thickness classes, a method based on the application of computerized image analysis combined with graphical and statistical methods was developed. The method allowed common classes of microfibrils and lamellae in cell walls to be identified from different origins. During both the formation and degradation of cell walls, a preferential formation of structures with specific thickness was evidenced. The results obtained with the developed method allowed objective analysis of patterns of microfibril aggregation and evidenced a trend of doubling/halving lamellar structures, during cell wall formation/degradation in materials from different origin and which have undergone different treatments.     

Ultrastructural observations on changes in cell shape in chromatophores of the sea urchin Centrostephanus longispinus

Summary Alterations in cell shape of the light-sensitive chromatophores of the sea urchin Centrostephanus longispinus were studied by scanning- and transmission electron microscopy. Transition of the aggregated to the dispersed state is accompanied by incorporation of vesicles into the membrane of the pigment cell. During dispersion a system of microtubules originating from centriole-like structures is established throughout the stellate cell. Within restricted areas of the cell, cytoplasmic differentiation and condensation is found. The possible functional significance of the findings is briefly discussed.     

Hemocytes of the centipede Scutigera coleoptrata (Chilopoda,Notostigmophora) with notes on their interactions with the tracheae

The hemocytes of Scutigera coleoptrata were investigated by light and electron microscopy. Four types of hemocytes were identified: prohemocytes, plasmatocytes, granulocytes, and spherulocytes. Only granulocytes could be distinguished from the three other types by May-Grünwald staining, as this is the only hemocyte type demonstrating an eosinophilic reaction. Shape and size give further indications for distinguishing the cell types. In addition, differentiation is possible on the basis of their ultrastructure. However, only a combination of all three methods (staining and light and electron microscopy) allows clear separation of the cell types. As transitional stages between the cell types occur in S. coleoptrata, it is likely that prohemocytes, plasmatocytes, and granulocytes are ontogenetic stages of a single cell lineage. Special cell components and their possible functions are described. Plasmatocytes exocytose tubular structures that probably play a role in coagulation processes. These tubular structures develop in the grana of plasmatocytes. Also, a special arrangement of microtubules and microfilaments was demonstrated. For the first time interactions between hemocytes and tracheae are documented within the Chilopoda. It is assumed that the hemocytes meet their oxygen requirements directly from the tracheae. Phylogenetic implications of the results are discussed.     

The cell biology of cell-in-cell structures

For decades, authors have described unusual cell structures, referred to as cell-in-cell structures, in which whole cells are found in the cytoplasm of other cells. One well-characterized process that results in the transient appearance of such structures is the engulfment of apoptotic cells by phagocytosis. However, many other types of cell-in-cell structure have been described that involve viable non-apoptotic cells. Some of these structures seem to form by the invasion of one cell into another, rather than by engulfment. The mechanisms of cell-in-cell formation and the possible physiological roles of these processes will be discussed.     

A mechanical explanation for cytoskeletal rings and helices in bacteria

Several bacterial proteins have been shown to polymerize into coils or rings on cell membranes. These include the cytoskeletal proteins MreB, FtsZ, and MinD, which together with other cell components make up what is being called the bacterial cytoskeleton. We believe that these shapes arise, at least in part, from the interaction of the inherent mechanical properties of the protein polymers and the constraints imposed by the curved cell membrane. This hypothesis, presented as a simple mechanical model, was tested with numerical energy-minimization methods from which we found that there are five low-energy polymer morphologies on a rod-shaped membrane: rings, lines, helices, loops, and polar-targeted circles. Analytic theory was used to understand the possible structures and to create phase diagrams that show which parameter combinations lead to which structures. Inverting the results, it is possible to infer the effective mechanical bending parameters of protein polymers from fluorescence images of their shapes. This theory also provides a plausible explanation for the morphological changes exhibited by the Z ring in a sporulating Bacillus subtilis; is used to calculate the mechanical force exerted on a cell membrane by a polymer; and allows predictions of polymer shapes in mutant cells.     

Dynamics of silica cell wall morphogenesis in the diatom Cyclotella cryptica: Substructure formation and the role of microfilaments

Diatoms are unicellular algae that make cell walls out of silica with highly ornate features on the nano- to microscale. The complexity and variety of diatom cell wall structures exceeds those possible with synthetic materials chemistry approaches. Understanding the design and assembly processes involved in diatom silicification should provide insight into patterning on the unicellular level, and information for biomimetic approaches for materials synthesis. In this report we examine the formation of distinct cell wall structures (valves and girdle bands) in the diatom Cyclotella cryptica by high resolution imaging using SEM, AFM, and fluorescence microscopy. Special attention was paid to imaging structural intermediates, which provided insight into the underlying design and assembly principles involved. Distinct stages in valve formation were identified, indicating a transition from a fractally organized structure to a dynamic pathway-dependent process. Substructures in the valves appeared to be pre-positioned prior to complete silicification, suggesting that organics responsible for these structures were pre-assembled and put in place. Microtubules and microfilamentous actin play significant roles in the positioning process, and actin is also important in the pathway-dependent expansion of the front of silicification. Our results indicate that even though all silica structures in C. cryptica are made of assemblies of nanoparticulate silica, control of meso- and microscale structure occurs on a higher order. It is apparent that diatoms integrate bottom up and top down control and synthesis mechanisms to form the diversity of structures possible.     

Presence of high concentrations of aluminum in neurons of a patient dying from a rapidly developing dementia

Ultrastructural observation of brain cells of a patient dead from senile dementia associated with myoclonies, has shown a number of intracytoplasmic dense structures. High concentration of Aluminium has been detected in these structures by electron probe X ray analysis. The clinical symptomatology, the evolution and the cell pathology observed in this patients are very similar to the classical features observed in dialysis encephalopathy the origin of which is an aluminium intoxication. In this patient, the possible origin of the intoxication is the absorption in the last 10 years of aluminium gels.     

High-salt solutions prevent reactivation of euglenoid movement in detergent-treated cell models ofEuglena gracilis

Summary Detergent-treated cell models ofEuglena gracilis showed rounded-up movement of the cell body upon addition of ATP and Ca²⁺. Reactivation of the cell models was inhibited when the cell models had been treated with solutions containing >150 mM NaCl or >300 mM KC1. When the supernatant of salt-extracted cell models was subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis, two distinctive bands of 120 and 160 kDa were found to be enriched. The cell membrane and associated cytoskeleton (pellicular complex) were isolated after treatment with salt solutions, and examined by electron microscopy to identify essential cortical structures required for reactivating rounding-up cell movement. Among three regularly arranged microtubules, only one and its associated structures were selectively dissolved from the pellicular strips, while the other pellicular elements remained intact. These structures were located in the groove region where sliding between strips is believed to occur during cell shape change. These results suggest a possible involvement of microtubule 2 and its associated bridges in active sliding between adjacent pellicular strips during euglenoid movement.     

SEM study of surface structures of the spathe in Cryptocoryne and Lagenandra (Araceae: Aroideae: Cryptocoryneae)

SEM studies of the spathe structures in the two closely related genera Cryptocoryne and Lagenandra show differences between the inner and outer surfaces, as well as in cell structures in the various parts of the spathe. The cell structure reveals patterns mat makes it possible to depict homologous structures of the spathe, even though the spathes of the two genera look different. The basal part of the kettle has a mucilage covering of the cells, interpreted as a hitherto unnoticed food source. The cells of the inner surface of the kettle and tube have downward pointing trichomes. On the second day of flowering these collapse and sink into the cell lumen, which is suggested to create a unique lattice-like structure that enables the insects to climb out of the kettle and tube. The cell structure of the flap shows that it is a prolongation and continuation of the spathe tube margin.     

Actin microfilaments in fungi

The cytoskeletal protein actin is among the most abundant proteins in nature. It is almost ubiquitous, occurring in all eukaryotes and in an ancestral form in prokaryotes. Actin monomers can polymerise to form microfilaments, structures that play a critical role in a number of fundamental cell processes in fungi such as morphogenesis, cytokinesis and the movement of organelles. Microfilaments are extremely dynamic structures and can be rapidly modified through their interactions with a number of actin binding proteins (ABPs). The purpose of the following review is to introduce actin and microfilaments in fungi to a general mycological audience and to provide a basic framework from which further study is possible.     

Fine structure of the plasmalemma surface of poplar parenchyma cells observed by the freeze etching technique

The plasmalemma surface of poplar parenchyma cells observed by the freeze etching technique is characterized by 11 nm particles, primary pit fields, lomasome-like structures and fibrillar structures. The most remarkable feature is the occurrence of fibrillar structures, which are considered to be the imprint of underlying microtubules on the plasmalemma surface. The previously reported observation on the possible appearance of microtubules at the cytoplasmic surface inside the plasmalemma is questionable. Although the fibrillar structures run almost perpendicular to the main cell axis, longitudinally oriented ones are also found, the occurrence of which is discussed in relation to the orientation of the cellulose microfibrils.     

COMPARISON OF THE GROWTH AND DIFFERENTIATION OF EPITHELIAL CELLS FROM NORMAL AND HYPERPLASTIC LENSES OF THE CHICK: STUDIES OF IN VITRO CELL CULTURES

Epithelial cells from hyperplastic lenses of a strain of chicks (Hy-1) selected for high growth rate were dissociated and cultured in vitro and compared with lens epithelial cells from a normal strain (N) in similar conditions. The hyperplastic lens cells showed remarkable motility and adhesiveness after dissociation and formed cell aggregates of various sizes before attaching to the substrate, giving a rather low plating efficiency. The lens structures (lentoid bodies) developed in partially confluent cultures of Hy-1 cells at least three days earlier than those in the cultures from normal control cells, in which the lens structures developed only after the cultures reached confluence. The results of culture at low cell density showed that the Hy-1 cell population consisted of at least two cell types different from each other in growth capacity. These striking differences in in vitro behaviour of dissociated cells from normal and hyperplastic lens epithelia and the results of clonal culture are discussed in relation to the possible mechanisms of abnormal morphogenesis and growth which are likely to be involved in the development of the hyperplastic lens in situ .     

THE FINE STRUCTURE OF TWO UNUSUAL STALKED BACTERIA

Two strains of bacteria that produce slender appendages (pseudostalks) from their lateral surfaces were studied using the electron microscope. The pseudostalks were shown to be extensions of the cytoplasm and peripheral membranes of the cell proper. Both strains of bacteria produce holdfasts at the poles of the cells by the means of which attachment can take place. The pseudostalks are not involved in the attachment of cells. No specialized intracytoplasmic structures are present at the point of juncture of pseudostalk and cell. A discussion of the possible functions of the pseudostalks, based on the electron microscope findings, is presented.     

Spontaneous senescence in the MDA-MB-231 cell line

Normal human somatic cells have a limited division potential when they grow in vitro. It is believed that shortening of telomeres, specialized structures at the ends of chromosomes, controls cell growth. When one telomere achieves a critical minimal length, the cell cycle control mechanism recognizes it as DNA damage and causes the cell's exit from the cycle in G1-phase. Because it is not possible to extend telomeres in normal cells, this non-dividing state is prolonged indefinitely, and is known as cellular senescence. The immortal cell line MDA-MB-231 has active telomerase, which prevents telomere shortening and allows cells' permanent divisions. However, there is a fraction of cells that do not divide over several days in culture as documented for some other tumour cell lines. Combination of methods has made it possible to isolate these non-growing cells and compare them with the fraction of fast-growing cells from the same culture. Although the non-growing fraction contains a significant percentage of typical senescent cells, both fractions have equal telomerase activity and telomere length. In this paper we discuss possible mechanisms that cause the appearance of this non-growing fraction of cells in cultures of MDA-MB-231, which indicate stress and genome instability rather than variation in telomerase activity or telomere shortening to affect individual cells.     

百合组织中细胞内生菌的分布与传播

在百合鳞茎、根、地上茎、叶和花蕾组织细胞中观察到细菌的分布。但各组织器官之间、细胞内所含细菌的数量差异很大。鳞茎组织细胞内含菌量最多。同一鳞茎,外围鳞片细胞内含菌量高于内侧。生长锥顶端分生组织细胞内未观察到细菌的分布。在生长锥中部有少量细菌出现,而基部则含有较多的细菌。百合鳞茎最外一层鳞片的外表皮中,细胞内有许多呈树丛状分布的类似侵染线的管状结构,它们与细胞壁发生联系,推测这些细菌可能是外源的。细菌随着植株的生长发育,由已成熟的含菌细胞向幼嫩的不含菌细胞中传播。细菌在细胞之间的传播可能是通过细胞壁上纹孔间的胞间连丝孔道。     

From the cell biology to the development of new chemotherapeutic approaches against trypanosomatids: dreams and reality

Members of the Trypanosomatidae family comprise a large number of species that are causative agents of important diseases such as sleeping sickness, Chagas' disease and Leishmaniasis. These organisms are also of biological interest since they are able to change the morphology according to the environment where they live, through a process of reversible cell transformation, and possess structures and organelles that are not found in mammalian cells. This review analyses the process of transformation, which takes place during the life cycle of Trypanosoma cruzi in the vertebrate and invertebrate hosts. Special attention is given to the interaction of the parasite with vertebrate cells. In addition, the present knowledge of structures and organelles such as the nucleus, the plasma membrane, the sub-pellicular microtubules, the flagellum, the kinetoplast-mitochondrion complex, the peroxisome (glycosome), the acidocalcisome and the structures and organelles involved in the endocytic pathway, is reviewed from a cell biology perspective. The possible use of available data for the development of new anti parasite drugs is also discussed.     

Centromere assembly and propagation

Centromere assembly provides a unique example of how elaborate protein structures can be assembled onto DNA, independent of sequence, and then stably propagated through numerous cell divisions. Here, we review the possible epigenetic strategies that organisms ranging from yeast to human use to assemble and propagate active centromeres.