An analytical electron microscope study of the kinetics

The detection and quantification of the segregation of Bi to grain boundaries in Cu using Analytical Electron Microscopy is demonstrated and the effects of time and temperature are observed. The experimental data are compared with the theoretical prediction of McLean’s segregation isotherm. Special grain boundaries are also considered. The analysis shows that the detection of the variation of segregation with time and temperature as well as grain boundary characteristics is possible using analytical electron microscopy and that the quantification techniques used here agree well with theoretical calculations of the segregant levels.     

Stereological image analysis of cultured human melanocytes observed by transmission electron microscopy

The aims of the study were to write an image analysis (IA) program allowing the stereological quantification of human epidermal melanocyte melanization at the ultrastructural level and to specify the suitable preparative methods, in keeping with IA limits and stereological principles. Micrographs of cultured human melanocytes obtained in transmission electron microscopy were digitized with a scanner. The key step of the designed IA program is a thresholding based on the gray levels. Hence, gray level histograms (pixel frequency as a function of gray level) of melanocyte images exhibit a peak specific to melanin. The gray level thresholding used consists in isolating the melanin pixels that form profiles on a binary image and in storing the numerical data produced for a given melanocyte profile. These primary data are used to calculate numerous parameters via stereology with melanocyte cytoplasm and melanized melanosome as main reference spaces. The most important stereological parameters obtained are v(mi,cy) (melanin volume per average cell), v(mi,m) (melanin volume per average melanized melanosome), and nm (number of melanized melanosomes per average cell), and their validity is discussed. Melanocytes embedded in situ were abandoned for stereological reasons but pelleted melanocytes were found suitable. Using this computerized tool and stereology, we are able to perform quantitative studies producing varied data even from small cell samples. To our knowledge, this is the first stereological approach for quantifying intracellular melanization. A quantitative comparison of spectrophotometrical results (melanin assay) with stereological results obtained in ultraviolet B-irradiated Caucasian epidermal melanocytes will be performed in order to appraise this method.     

An electron microscope study of the axonemal ultrastructure in human spermatozoa from male smokers and nonsmokers

OBJECTIVE: To investigate possible abnormalities or deterioration of the sperm axonemal ultrastructure in men who have smoked a large quantity of cigarettes (> 20 per day) for a prolonged period. DESIGN: Semen specimens were collected by patients via masturbation; qualitative characteristics of the sperm were assessed and ultrastructural analysis of the sperm axoneme was performed using standard operating procedures for electron transmission microscopy. SETTING: The Andrology Institute of Lexington, Lexington, Kentucky, and the Department of Histology and Embryology, University of Salonika, Greece (collaborative effort). PATIENT(S): Twenty-nine men (mean age +/- SD, 30.7 +/- 2.1 years) who smoked a mean (+/- SD) of 30.7 +/- 2.1 cigarettes per day for 10.7 +/- 0.7 years and 15 men who never smoked (mean age +/- SD, 30.4 +/- 2.2 years) participated in this study. MAIN OUTCOME MEASURE(S): Ultrastructural organization of the sperm axoneme in male smokers and nonsmokers. RESULT(S): Changes in the number and the arrangement of axonemal microtubules were noted in the smoker group when compared to the nonsmoker group. The incidence of axonemal abnormalities was higher in spermatozoa from smokers compared with that in spermatozoa from nonsmokers. CONCLUSION(S): Smoking a large quantity of cigarettes per day, under the conditions of the current study, severely affected the ultrastructure of the flagellum and, more specifically, it affected the axoneme of the human spermatozoon.     

The human uterotubal junction: a scanning electron microscope study during different phases of the menstrual cycle

Uterotubal junctions from surgically extirpated human uteri were examined. The specimens were obtained during different phases of the menstrual cycle. The interstitial portions of the tubes together with the cornual areas were dissected, excised, and their luminal surfaces exposed. The specimens were then processed for scanning electron microscopy. The surface epithelium of both the cornual endometrium and interstitial endosalpins. Ciliated cells were more numerous in the endosalpinx. Cyclic changes in ciliated cells were minimal, while cyclic secretory activity was demonstrated, especially in the endometrium. The transitional area between the endometrium and the endosalpinx was characterized by a marked increase in the number of ciliated cells, and a tendency of the secretory cells to assume a flattened, polygonal shape. These morphologic features suggest a possible role in the transport and/or maintenance of spermatozoa and/or ova.     

Cytoadherence of Plasmodium falciparum to intercellular adhesion molecule 1 and chondroitin-4-sulfate expressed by the syncytiotrophoblast in the human placenta

Late stages of Plasmodium falciparum-infected erythrocytes (IRBCs) frequently sequester in the placentas of pregnant women, a phenomenon associated with low birth weight of the offspring. To investigate the physiological mechanism of this sequestration, we developed an in vitro assay for studying the cytoadherence of IRBCs to cultured term human trophoblasts. The capacity for binding to the syncytiotrophoblast varied greatly among P. falciparum isolates and was mediated by intercellular adhesion molecule 1 (ICAM-1), as binding was totally inhibited by 84H10, a monoclonal antibody specific for ICAM-1. Binding of the P. falciparum line RP5 to the syncytiotrophoblast involves chondroitin-4-sulfate (CSA), as this binding was dramatically impaired by addition of free CSA to the binding medium or by preincubation of the syncytiotrophoblast with chondroitinase ABC. ICAM-1 and CSA were visualized on the syncytiotrophoblast by immunofluorescence, while CD36, E-selectin, and vascular cell adhesion molecule 1 were not expressed even on tumor necrosis factor alpha (TNF-alpha)-stimulated syncytiotrophoblast tissue, and monoclonal antibodies against these cell adhesion molecules did not inhibit cytoadherence. ICAM-1 expression and cytoadherence of wild isolates was upregulated by TNF-alpha, a cytokine that can be secreted by the numerous mononuclear phagocytes present in malaria-infected placentas. These results suggest that cytoadherence may be involved in the placental sequestration and broaden the understanding of the physiopathology of the malaria-infected placenta.     

Localization of vitamin A in the small intestine of mouse. An electron microscope radioautographic study

Localization of vitamin A in the small intestine of mice was studied with electron microscope radioautography after administration of tritiated vitamin A. The label was concentrated over lipid droplets in cells distributed in the lamina propria and the submucous layer. The cells were similar both to fibroblasts and to fat-storing cells in their morphological features. The name "Vitamin A-Storing Cell" is proposed for these labeled cells, including the fat-storing cell in the liver.     

Isolation from human placenta of the IgG transporter, FcRn, and localization to the syncytiotrophoblast: implications for maternal-fetal antibody transport

The IgG transporter responsible for ferrying maternal IgG across the human placenta to fetal circulation has not been identified, although the human homologue of the neonatal rat Fc receptor (FcRn), a heterodimer with pH-dependent IgG affinity, structurally similar to MHC Class I molecules, was recently proposed as a candidate. Affirming this hypothesis, we describe herein the specific copurification from human placenta of 46- and 14-kDa proteins by IgG affinity at acid pH. The larger protein, characterized by its amino acid sequence and by immunoblot, is the alpha-chain of human FcRn (hFcRn). The smaller is beta2-microglobulin. Their coisolation by ligand affinity suggests that they comprise the hFcRn heterodimer. Placenta sections stained immunohistochemically with anti-hFcRn alpha-chain peptide Abs show extensive expression of hFcRn in the syncytiotrophoblast and traces in the endothelium and other unidentified cells of the villus stroma. We find alpha-chain mRNA by Northern analysis in human placenta and in human trophoblast-like cell lines (JEG-3, ED27) but not in a human myelocytic cell line (HL60). We suggest that the placental hFcRn heterodimer may transport IgG to the fetus by a mechanism in which maternal IgG is pinocytosed nonspecifically and then carried to fetal tissues by a pH gradient from acidic endosomes to the pH-neutral basolateral surface of the syncytiotrophoblast. Furthermore, the known characteristics of FcRn suggest a wider function, that it is the receptor postulated by Brambell in the 1960s to regulate tissue and serum IgG concentrations by controlling IgG transport and catabolism.     

A scanning electron microscope study of the interstitial tissue of the canine testis

Scanning electron microscopy (SEM) is a potent tool that is especially valuable in interpreting the three-dimensional relationships of cells within tissues. This type of information is obtainable from thin sections in transmission electron microscopy (TEM) only by reconstructions of serial sections. The arrangement of the interstitial cells of the testis in relation to the capillaries and lymphatic channels, in particular, is easier to visualize in SEM than in TEM. Cytoplasmic constituents, as well as cell surface modifications, are demonstrable by this technique. The presence of droplets, presumably lipid droplets, both within and on the Leydig cells and the lymphatic endothelial cells, is quite evident. Other cytoplasmic structures are also apparent. For example, the possible functional significance of "openings" that are seen by SEM on the septa that surround lipid droplets is discussed relative to the appearance of the same area as seen in thin sections or in freeze-fracture replicas. SEM should become a very useful method for studying cytological and morphological alterations that occur in testicular tissue that is subjected to physical or chemical manipulation.     

Thy-1 immunolabeled thymocyte microdomains studied with the atomic force microscope and the electron microscope

The atomic force microscope (AFM) and the transmission electron microscope (TEM) have been used to study the morphology of isolated mouse thymocyte microdomains and Thy-1 antigen distribution at the surface of these structures. AFM images were recorded in air in the contact mode on membrane vesicles deposited on previously heated tissue culture plastic sheets and indirectly immunolabeled for Thy-1 expression with colloidal gold-conjugated secondary antibodies. AFM images of untreated plastic plates showed a very characteristic network of streaks 20-200 nm wide. Heating the plastic removed the streaks and provided flat surfaces (r.m.s. 1 nm). This substrate allowed strong adsorption and homogeneous spreading of the vesicles and easy manipulations during immunolabeling experiments. Vesicles flattened on the substrate without losing their morphology. The 10-nm membrane-bound gold beads were reproducibly imaged without degradation by repeated tip scanning. The observed microdomains had a mean diameter of 184 +/- 76 nm, and 65% of them were specifically labeled. Images obtained with the TEM on the same vesicles, deposited on carbon-coated grids and negatively stained, confirmed the AFM observations. The size distribution of the microdomains was quite similar, but the number of beads per vesicle was significantly higher, and 76% of the vesicles were labeled. The difference may be explained 1) by removal of beads from the vesicles in the additional washing step with water, which was necessary for the AFM; 2) by tip-sample convolution; and 3) by statistical fluctuations.     

Acetylcholinesterase distribution in chick spinal cord cultures. A light and electron microscope study

In vitro studies and animal experiments were started for the purpose of following the migration of chloramphenicol marked with 14C from polymerised polymethylmetacrylate cylinders. Test-cylinders were submerged in a physiological saline solution and the 14C concentration followed over a period of 34 days. Two series were started with the cylinders being submerged at intervals of 5 and 40 min after the start of polymerisation. These in vitro studies showed that initially the migration of active substances marked with 14C from the plastic cylinders was extremely high but remained then constant over the whole test period. Over a period of at least 20 days a higher 14C migration was evident in those cylinders submerged in the elution 5 min after the start of polymerisation. In our in vivo studies we implanted test-cylinders into the femur of rabbits. The migration of active substances marked with 14C resulted in 14C-concentrations being present in the surrounding tissues. During the phase of exudation, depending on the operative process, the concentration increased and reached values that remained constant during the phase of recanalisation of the surrounding tissues over a period of up to 6 weeks. Thereafter the concentration of active substances fell. The max. concentration rates of active substances within the boundary layer were between 15 and 20 mug/g humidity weight. After 8 weeks we found concentration rates of approx. 1 mug/g humidity weight. On examining the 14C distribution within the implanted plastic cylinders we observed that in the course of time the boundary layer increased in thickness and a concentration gradient towards the centre had developed. The spread of diffusion gradually seized the deeper layers so that a long-lasting presence of active substances may be expected.     

Regeneration of the node of Ranvier: a light and electron microscope study

Regeneration of the node of Ranvier was investigated in the rat peroneal nerve 10-60 days after nerve crush, by light and electron microscopy. At 10 and 20 days after crush nodes of Ranvier were clearly identifiable by electron microscopy but had a relatively simple structure. At 40 days after crush however nodes were highly differentiated showing specialised features such as paranodal bulbs, nodal constriction of the axon, paranodal Schwann cell mitochondria, nodal Schwann cell microvilli, and nodal gap substance. By light microscopy some nodes were identifiable as early as 20 days after crush. At both 30 and 60 days after crush regenerated internodes were uniformly short (means of 275 micronm and 339 micronm respectively).     

Relief of the endothelial surface of human arteries according to scanning electron microscope findings

The work presents data on the relief of the endothelial surface of initial portions of the common carotid artery and the common iliac artery of adult man obtained by the method of scanning electron microscopy. In middle-aged humans this relief has all the characteristic features which were noted in studying the aorta. Considerable changes of the endothelial surface relief were shown in elderly and old people in connection with the development of atherosclerosis. Postmortem alterations can result in sharp distortion of the endothelial surface relief.     

Subchronic toxicity of pentachlorobiphenyl congeners n. 126 or 118 in the rat liver. An electron microscope study

3,3',4,4',5-Pentachlorobiphenyl (PCB) or congener n. 126 and 2,3',4,4',5-pentachlorobiphenyl or congener n. 118 were given independently to male and female Sprague-Dawley weanling rats. Experimental diets were prepared by dissolving the congeners in 4% corn oil. The congeners were administered as follows: congener n. 126--groups of three animals, either male or female, in each group were placed on the respective diets containing 0.1, 1.0, 10.0 ppb congener, 5.0 micrograms/kg bw loading dose + 10.0, or 100 ppb; congener n. 118--the females were dosed with 2, 20, 200, and 2,000 ppb congener, and the males received 10, 100, 1,000, 10,000 ppb. Thirteen weeks after the start of dosing with the two congeners, liver samples were obtained from all the animals and prepared for electron microscopy. In the congener n. 126-exposed animals, the alterations noted in a dose-related fashion consisted of a marked increase in the profiles of smooth endoplasmic reticulum (SER), and in the heightened number of lipid droplets in many parenchymal cells. Mitochondria showed abnormalities such as dumb-bell shapes, and the cristae parallel to the long axis of the organelle. Lipofuscin granules were numerous in the liver of animals that received 100 ppb of the congener; notably the females of the treatment group expressed this trait more abundantly than the males of the group. We conclude that the compound is mildly toxic. In the animals administered congener n. 118, the alterations were revealed in the liver of both male and female animals in a dose-related manner, also the most evident hepatocyte architectural modifications included an augmentation of SER profiles, mitochondrial aberrations, and an elevated number of lysosomal elements and lipid droplets. Abnormal shapes, and cristae in atypical orientation comprised mitochondrial aberrations. Alterations in the liver morphology of the females were qualitatively similar to those in the males; however, the dose levels used in the latter were five-folds of that which were given to the females. We conclude that the females are more sensitive than the males of the species to congener n. 118. We further conclude that congener n. 118 is less toxic than n. 126 since the lesions were induced by several-folds high dose levels used for the former.     

A microscopical study of wound repair in the human placenta

In order to fulfill its many functions as the selective interface between maternal and fetal circulations it is imperative that the human placenta remains intact and in good operational order. That damage of some sort occurs during its short but extremely active life seems inevitable given the dynamic environment in which the placenta exists, and evidence has accumulated that disruption is indeed a regular event. The implications of such damage, one could speculate, may impact on functions such as transport and hormone secretion as well as mutual protection against attack by maternal and fetal immune systems. Consequently, it would seem a theoretical necessity for discontinuities in the placenta surface to be repaired as soon as possible. We have used a combination of ex vivo observation, in vitro modelling, immunohistochemistry and correlative microscopy to provide evidence for a wound response in the placenta and to begin dissecting the detail of how this may operate. Evidence for small lesions caused by fusion and subsequent tearing of the syncytiotrophoblast in vivo, as well as plugging of such wounds by underlying cells is shown. We also identify a putative role for migratory cytotrophoblasts in the healing of larger scale injuries and demonstrate that certain molecules, common to wound repair in other tissues, appear to be involved in placenta repair also. Taken together these results clearly show that the human placenta is capable of a degree of self-maintenance by activating what appears to be an endogenous wound healing mechanism.     

In vitro perfusion studies of the human placenta. IV. Some characteristics of the glucose transport system in the human placenta

Cross infection has become a serious risk to hospitalized patients. Potential sources of infection by anaesthetic apparatus and equipment and the danger arising from disregard of proper asepsis are discussed. Prophylactic and hygienic measures to minimize these hazards are reviewed. Since patients receiving intensive therapy are particularly are risk very high hygienic standards are a "must" in these units. The need for thoroughly and regularly checking all equipment for contamination is emphasized.