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Difficulties in testing the possible role of calcium as a Transmitter in the R.O.S. are discussed. A comparison is made with the Sarcoplasmic Reticulum system where calcium flux are easily measured. The latest results reviewed on intact cellular structures are highly indicative but not yet conclusive.
The transepithelial flux of chloride was increased by aldosterone treatment of amphibian skin and bladder and this was reflected by increased "shunt" conductance. The hormonal effect depended on the presence of chloride on the epithelial side of the preparation. These changes in tissue conductance and chloride permeability appear to be a direct effect of aldosterone as they did not occur when sodium transport was stimulated with vasopressin or hypotonicity. Chloride efflux was reduced in magnitude by indacrinone and DIDS, as well as after removal of chloride from the solution on the epithelial side of the preparations. These results suggest that, rather than merely diffusing along (a) paracellular pathway(s), chloride flows through (a) cellular structure(s), notably mitochondria-rich cells. These cells can therefore be considered as targets for aldosterone.
The site of hormone action is the cells of target tissues. Now that ultrastructural research has revealed the complexity of eukaryotic cells, we must ask which cellular structures are involved. Some hormones act at the cell membrane, mainly via adenylate cyclase and the "second messenger" cAMP. The cytosol and the mitochondria, the main sites of intermediate metabolism and enzyme activity, are apparently not involved, although in the past interaction of hormones with enzymes has been postulated. Most of the steroid hormones act at the level of the cell nucleus, regulating transcription of specific genes and inducing enzymes or other proteins. The mechanisms by means of which this regulation is achieved are not fully understood, but it is likely that derepression is involved. The active agent seems to be a complex of the hormone with a receptor protein.
Using a cystic lymphangioma of the greater omentum and a benign cystic mesothelioma as examples, the distinguishing characteristics of cystic peritoneal tumors are shown, using light microscopic and ultrastructural findings. A benign papillary mesothelioma is used for comparison. The cellular structures and growth rate of the mesotheliomas are contrasted with the tissue reactions which are typical for irritated serosa. The diffusely growing papillary mesothelioma is a very rare tumor, the cells of which are similar to normal serosa cells, but also show characteristics of other benign or malignant mesothelial tumors and of reactive proliferated mesothelial cells. The diffusely growing benign cystic mesothelioma has only been described in isolated cases and is characterized by cysts grouped in acini with mainly flat, localized cuboidal cell lining. The histochemical properties and cytological findings correspond closely to those of the papillary tumor or normal peritoneal lining cells. In contrast, the cystic lymphangioma probably represents a congenital defect with a slow growth rate. The structure is characterized by a sponge-like arrangement of smooth-walled cysts, in the walls of which smooth muscle cells and lymph follicles are embedded. The endothelium is also flat and ultrastructurally resembles that of lymph vessels.
During laparoscopy we performed 132 ovarian punch biopsies in seventy one amenorrhoeic patients. With increasing experience, laparoscopic ovarian biopsy is a valuable element in the examination of amenorrhoeic patients. Biopsy specimens with an edge length of 4-5 mm for surface and depth can be obtained during laparoscopy under general anaesthesia without any great risk or difficulty. When performed bilaterally, it provides a representative picture of the cellular structure and function of the ovaries and thus makes a contribution to the prognosis of a disturbance in the endocrine regulating-circuit of ovarian function. In addition, the ovarian biopsy provides information about the basic gonadotrophic situation of the patient as a longitudinal section. It should be included in the synopsis of all endocrine and genetic parameters of a patient, particularly in clinic in which the morphological substrate of disturbed functions and apparent diseases is still examined.
Serum proteins as constituents of vasogenic brain edema were visualized both macroscopically and microscopically applying a double layer immunofluorescence technique to paraffin embedded material derived from three experimental series: peritumorous edema following xeno-transplantation of glioma cells, edema after cerebral embolization with micropheres, and edema after unilateral MCA occlusion. Exclusively cats were used as experimental animals. The staining procedures resulted in selective green fluorescence of vessel contents as well as edema protein, which was demonstrable even macroscopically at times, where edema formation reaches a maximum in each experimental series. Microscopically, serum protein could be traced up to the end of observation time ranging from 1 to 4 weeks, where the specific fluorescence was related to cellular structures. As compared to other techniques employed in brain edema localization, immunostaining mainly offers the following advantages: avoidance of in vivo tracing, better structural resolution in paraffin than in freeze sections, high specificity and sensitivity in antigen localization.
Spontaneous neuronal activity was recorded in the cerebral cortex and the CA1 sector of the hippocampus in gerbils during and after 5-min ischemia, produced by bilateral clamping of the common carotid arteries. It was found that spontaneous activity in both cortical and CA1 neurons ceased within 60s after the onset of ischemia and that it began to reappear 10-20 min after the recirculation. During the next 24 h most CA1 neurons which were recorded showed hyperactivity. This was evident primarily by an increase in spike discharges, whereas recordings from the cerebral cortex were within the preocclusion ranges. On the 2nd day after ischemia, functioning CA1 neurons could not be found, as if they were in a state of functional death, although histological sections showed a general preservation of their cellular structure at that time.
Heterochronia, heterotopia and heterometria are important elements of the pathogenic process. Heterochronia has its historical roots in the philosophy of Aristotle. As the organs of living creatures and especially man have developed, the process has been one of fitting together a variety of evolutionary bricks. Each complex organ shows evidence of a gradual control of its cellular structure. Parts of organs observed at different stages of man's development all appear to retain certain heterologous (identical) organisational features. Thus, the different pathoclisis is explained. Classical Neuropathology has made headway in this field. Its methodical procedure was adopted - mutatis mutandis - by General Pathology. The inherent structural weaknesses within the organisation of the human brain are, in their organic type of differentiation, also present in the muscular heart. The right ventricle has developed mainly from the myoepicardium of the proampulla; the left ventricle has its origin in the metaampulla. The right ventricle consists mainly of priscomyocardium, the left one of neomyocardium. This is the basis on which are founded the main characteristics of the muscular structures, the arrangement of the coronary arteries and the organisation of the conduction system. A. Keith had already discovered in 1906 that the elements of specific muscle texture contain old structures. We believe that the pacemaker cell represents the historical paradigm of the working muscle cell of the heart (the transporter). Atrioventricular collateral connections (anastomoses) are only found within the priscomyocardium, the arteria coronaria dextra being the original. The coronaria sinistra was evolved later. From the summary of facts originating from the heterochronia of heart development we can draw conclusions about the development of the major heart diseases such as: Those which are dependent on interference with the ventricle walls following oxygen deprivation and the resulting toxic stress; Those which are characterised by infarction of the myocardium which occur in areas which have a predilection for such reaction and which can be set out in a predetermined topology; Those characterised by rhythmic interferences of the heart caused by atrioventricular collateral connections; as a result of an anthropological pathomorphology.
The clinico-pathologic findings in an additional case of gliomatosis cerebri are reported: a 60-year-old woman died 8 months after the onset of a progressive deterioration of both the neurologic and mental conditions. Neuropathologic examination disclosed wide demyelination of both hemispheres, communicating through the corpus callosum, extending downward along the internal capsule to the brainstem structures. Cellular stains showed the presence of elongated astrocytes, multinucleated cells, mitotic and anaplastic figures, involving the demyelinated areas and the neighboring regions, and allowed the diagnosis of gliomatosis cerebri diffusa. The nosologic and pathogenetic aspects of this rare entity are discussed.
The activities of several enzymes which metabolize xenobiotics were measured and compared in freshly isolated rabbit Clara cells (50-70% purity) and alveolar type II cells (80-95% purity) or microsomal preparations from the isolated cell fractions. The presence of 1 mM nicotinamide in protease and cell isolation buffers increased significantly 7-ethoxycoumarin (7-EC) deethylase and epoxide hydrolase activities in the isolated Clara and type II cells. Isolated Clara cell fractions metabolized 7-EC to umbelliferone at a rate of 241 +/- 27 pmoles/mg prot/min (mean +/- S.E., N =5), while the 7-EC deethylation rate in type II cells was 111 +/- 15 pmoles/mg prot/min. Coumarin hydroxylation activity, however, was more than ten times greater in the Clara cells than in the type II cells on a per mg cellular protein basis. N-oxidation of N,N-dimethylaniline, catalyzed by a flavin monooxygenase, was about 2 times as great in microsomes of Clara cells as in microsomes of type II cells. Epoxide hydrolase activity with benzo(a)pyrene 4,5-oxide as substrate was about 10 times higher in Clara cells than in type II cells. Because of the greater cellular, structural and functional heterogeneity in lung, differential distribution of enzymes responsible for xenobiotic metabolism in this tissue may contribute to cell selective chemical toxicity and carcinogenesis.
The morphology of the body wall and the gut in the midbody region of adult male and female Loa loa originating from patients in Gabon was studied by transmission and scanning electron microscopy. The cuticle of the dorsal and ventral regions consists of ten layers. In the lateral regions the cuticle is thicker and includes two additional layers. The thin hypodermis contains numerous transhypodermal fibres. A row of median cells is situated between the syncytia in each lateral chord. No intracellular bacteria were observed. The cross-sections of each of the four muscle sectors are comprised of approximately 12 muscle cells of the coelomyarian type. The plasm of the gut cells contains large vacuoles and several mitochondria. The intestinal wall surrounds a wide lumen filled with material which occasionally contains cellular structures. The morphology of L. loa is compared with that of adult Onchocerca volvulus and Brugia malayi. The gut of the adult L. loa has the typical nematode morphology, which might be an indication of its normal function in nutrition. The multilayered cuticle with the rather smooth surface, and the prominent muscles correspond to the migratory activity of this filaria.
Mono-ADP-ribosylation appears to be a reversible modification of proteins, which occurs in many eukaryotic and prokaryotic organisms. Multiple forms of arginine-specific ADP-ribosyltransferases have been purified and characterized from avian erythrocytes, chicken polymorphonuclear leukocytes and mammalian skeletal muscle. The avian transferases have similar molecular weights of approximately 28 kDa, but differ in physical, regulatory and kinetic properties and subcellular localization. Recently, a 38-kDa rabbit skeletal muscle ADP-ribosyltransferase was purified and cloned. The deduced amino acid sequence contained hydrophobic amino and carboxy termini, consistent with known signal sequences of glycosylphosphatidylinositol (GPI)-anchored proteins. This arginine-specific transferase was present on the surface of mouse myotubes and of NMU cells transfected with the cDNA and was released with phosphatidylinositol-specific phospholipase C. Arginine-specific ADP-ribosyltransferases thus appear to exhibit considerable diversity in their structure, cellular localization, regulation and physiological role.
Crithidia oncopelti, C. deanei, and C. desouzai are flagellates of the Trypanosomatidae family that present bacterium-like endosymbionts in their cytoplasm. Direct and indirect lectin-gold labeling techniques were used at the electron microscopic level in Lowicryl K4M-embedded cells to demonstrate the presence of intracellular lectin-binding sites. We used the lectins Ulex europaeus I, Griffonia simplicifolia II, Ricinus communis I, Arachis hypogaea, G. simplicifolia I, Wistaria floribunda, Limulus polyphemus, and Canavalia ensiformis, which recognize alpha-L-fucose, alpha- and beta-N-acetylglucosamine, beta-galactose and beta-N-acetylgalactosamine, beta-galactose, alpha-galactose, beta-N-acetylgalactosamine, sialic acid and alpha-D-mannose, and alpha-D-glucose residues, respectively. The nucleus was the cellular structure most frequently labeled by the lectins. The Golgi complex was seldom labeled, whereas the endoplasmic reticulum and the flagellar pocket presented a large number of binding sites. Symbionts had their two unit membranes weakly labeled by the different lectins but displayed no labeling of the space between the membranes.
This paper describes a new method for the rapid fixation of labile lipid pools in the brain. Perfusion of the brain with 0.9% saline containing esterase inhibitors (p-bromphenacyl-bromide and diisopropyl fluorophosphate), an antioxidant (nordihydroguaiaretic acid) and a Ca2+ chelator (EDTA) resulted in a substantial reduction in the levels of free fatty acids, a biochemical marker for the degradation of labile membrane lipids. Levels of unesterified polyunsaturated fatty acids in whole brain were decreased by 90-96% as compared to levels in brains perfused with saline alone. Levels of docosahexaenoic acid approximated levels obtained after microwave irradiation. Unlike microwave irradiation, this perfusion technique perserves the cellular structure of the brain, thereby allowing subcellular fractionation with minimal postmortem changes in lipid pools. The release of arachidonic acid during isolation of the P2 (synaptosomal) fraction was completely inhibited by the presence of the metabolic inactivators. The results of this study demonstrate a new and useful technique for the postmortem inactivation of enzymes responsible for the degradation of labile lipids in the brain. Further, the data underscore the key role of phospholipase A2 and Ca2+ in mediating the release and accumulation of free fatty acids in the ischemic brain.
The kinetic behaviour of succinate dehydrogenase [EC 1.3.99.1] in three fibre types of rat gastrocnemius was examined by a quantitative histochemical method without disruption of the cellular structure. 2-(2-Benzothiazolyl)-3-(4-phthalhydrazidyl)-5-styryl-t etrazolium chloride (BPST) and phenazine methosulphate were used as electron acceptors. On measurement of the absorbance value at 530 nm of BPST formazan, produced by the succinate dehydrogenase reaction in sections, it was found that the staining intensity of succinate dehydrogenase was linearly proportional to both the incubation time and the thickness of the slice therefore, the initial velocity of the staining could be calculated. By Michaelis-Menten (1913) treatment of the dependence of the initial velocity on the substrate concentration in the absence and the presence of a competitive inhibitor, malonate, the Km and Vmax values for succinate and the Ki value for malonate were obtained. The Km and Ki values of the three fibre types were similar. The ration of the Vmax values of type A, B and C fibres was 1.0:2.0:3.3. The temperature dependence of the kinetic parameters was very similar in the three fibre types. These findings confirm that the differences in the staining intensity of the three fibre types reflect differences in the amounts, but not the properties, of succinate dehydrogenase.
A theoretical concept is proposed, in order to explain some enigmatic aspects of cellular and molecular biology of eukaryotic organisms. Among these are the C-value paradox of DNA redundancy, the correlation of DNA content and cell size, the disruption of genes at DNA level, the "Chromosome field" data of Lima de Faria (Hereditas 93:1, 1980), the "quantal mitosis" proposition of Holtzer et al. (Curr. Top. Dev. Biol. 7:229 1972), the inheritance of morphological patterns, the relations of DNA and chromosome organisation to cellular structure and function, the molecular basis of speciation, etc. The basic proposition of the "Unified Matrix Hypothesis" is that the nuclear DNA has a direct morphogenic function, in addition to its coding function in protein synthesis. This additional genetic information is thought to be largely contained in the non-protein coding transcribed DNA, and in the untranscribed part of the genome.
125I incorporated in DNA is known to be exceptionally toxic. Values of D0 range from about 40 to about 90 decays for survival of mammalian cells. The effectiveness of 125I in DNA with respect to the induction of breaks of the DNA strands, however, appears to be comparatively low. The numbers of strand breaks per energy deposited in subnuclear cellular structures such as DNA is smaller for a disintegration of 125I than for gamma-rays. The difference in effectiveness diminishes with increasing mass of the considered sensitive volume. The apparent inefficiency of 125I-decay may, on one hand, result from a waste of local energy deposition. On the other hand, it may be caused by a multitude of local strand breaks (clusters) induced by 125I-decay which are measured as one break only by the conventionally applied techniques of strand break measurement. The apparent inefficiency of 125I may be evidence furthermore for the importance of considering not only the DNA as the sensitive target but with equal pertinence the gross sensitive volume, i.e. the whole cell nucleus. Further, for drawing meaningful comparisons, it may be necessary to take into consideration the microdosimetric event size distributions for the critical targets.
Subcellular distribution of a thiol-proteinase inhibitor protein was determined in the epidermis of the newborn rat by light and electron microscopy. This protein was highly soluble in basal cells and concentrated on ribosomes in the perinuclear region. Solubility in Tris buffer decreased in granular and cornified cells in which the protein appeared on polysomes which were attached on other cellular structures such as dense homogenous deposits and tonofilaments. The protein also appeared to be deposited on the plasma membrane and became insoluble in Tris buffer at 37 degrees C, but solubilized in 1 M phosphate buffer. Location of the protein around keratohyalin granules or by the plasma membrane suggested that the inhibitor protein bound to cysteine-rich protein of the epidermis with or without forming a thiol-proteinase inhibitor complex. The thiol-proteinase inhibitor protein seems to contribute to epidermal cell differentiation at multiple points through changes in its solubility and subcellular localization from basal cells to cornified cells.