Tissue Autoradiography Research Articles

Schistosoma mansoni: Migration potential of normal and radiation attenuated parasites in naive guinea pigs

Compressed tissue autoradiography using [ 75Se]selenomethionine labelled parasites has been used to investigate the migration potential of normal and radiation attenuated cercariae of Schistosoma mansoni in naive guinea pigs. By Day 14 after infection, 44% of normal parasites were detected as reduced silver foci in the liver; this value corresponded well with the number of liver parasites recovered by retrograde perfusion of the hepatic portal system on Day 42 (42% of the challenge). In contrast, cercariae subjected to 50 krad of gamma irradiation failed to migrate out of the skin. The migration capacity of 20 krad irradiated parasites was less severely affected in that about half of the challenge parasites reached the lungs, but virtually none moved to the liver. These data are discussed in relation to the kinetics of immunity induced in guinea pigs by infection or vaccination with normal or radiation attenuated parasites.

In vitro labeling and tissue autoradiography of splenomegaly associated with portal hypertension.

Sinus hyperplasia occurring in the congestive splenomegaly associated with portal hypertension is a interesting model of steady-state hyperplasia in an adult tissue. In vitro labeling of human splenic tissue with 3H-thymidine and the demonstration of S-phase cells by autoradiography was performed to investigate the proliferative activity of sinus-lining and cordal reticular cells in congestive splenomegaly compared with that in the normal spleen. The labeling index (%) of sinus-lining cells after 2 h incubation was 0.07 +/- 0.02 in the normal spleen and 0.26 +/- 0.03 in congestive splenomegaly (t = 4.553, P less than 0.005, n = 11). This result indicated that sinus-lining cells have a long life span and that their proliferative activity is increased in congestive splenomegaly. On the other hand, the labeling of cordal reticular cells and arterial and venous endothelial cells was sparse or absent in both congestive splenomegaly controls, and these cells do not appear to be involved in sinus hyperplasia.

Translocation of fixed carbon from symbiotic bacteria to host tissues in the gutless bivalve Solemya reidi

Solemya reidii (Bernard, 1980) were collected by Van Veen grab near a sewage outfall in California, USA, in spring/early summer of 1984 and 1985. Fixation of carbon dioxide by symbiotic chemoautotrophic bacteria in the gills, followed by translocation of the fixed carbon to symbiontfree tissues of the clam has been demonstrated using NaH14CO3 in two types of experiments. The first approach used the technique of tissue autoradiography to determine the exact sites of fixation of carbon dioxide and subsequent utilization of the fixed carbon compounds. Initial deposition (>95% after 10 min) was in the well defined portion of the gill filaments where the symbiotic bacteria reside. The heaviest redeposition of labeled carbon after a 2 d chase period was in the various glandular tissues of the clams (8 to 38% of the gill level). Levels in two of these glandular tissues decreased after a 5 d chase period, probably due to secretion. Significant labeling of muscle tissue, gonads, and the tips of the gill filaments occurred only after the chase periods. In the second approach, rates of initial carbon fixation by symbiotic bacteria and translocation into clam tissues were determined using pulse-chase experiments followed by dissections and assay of the clam tissues for radioactivity. Freshly caught clams fixed carbon at an average rate of 0.8 μmol CO2 g-1 wet weight h-1. In these fresh clams, 88% of the fixed carbon was found in the gills after 2 min incubation periods, with only 44% remaining in the gills after 1 to 1.7 h incubations. Individuals maintained in the laboratory for 100 d had only 5.8% of the RuBP carboxylase (a diagnostic enzyme for autotrophs) activity in their gills compared to freshly caught clams. After 2 min incubations in NaH14CO3, only 21% of the carbon fixed by the “old” clams was found in their gills, and the rate of incoporation into gill tissue was 5.7% of the rate in fresh individuals. These experiments demonstrate that the symbiotic bacteria fix carbon at a high rate, that a significant portion of the carbon fixed by the symbiotic bacteria (>45%) is translocated to the host, and that the translocated carbon is used by the host.

The utilization of [3H] sugars by non-malignant and malignant human breast.

The utilization of [3H] sugars and leucine by non-malignant and malignant human breast has been assessed using an organ culture technique with subsequent tissue autoradiography. The uptake of sugars by normal and hyperplastic breast was generally constant, with some differences observed in the utilization of galactose by acini of normal and hyperplastic tissues. After 24 h incubation localization was predominantly at the luminal cell periphery. The utilization of sugars by carcinomas was much more variable. Differences were observed between adjacent cells and cell groups of the same tumour. The uptake of individual sugars within a carcinoma was also varied being either similar to, or greater or lesser than normal breast. Variation between carcinomas was also present. No correlation between type and differentiation was noted in this respect, but there was between localization of sugars and differentiation. Better differentiated areas in tumours showed patterns similar to non-malignant breast whilst localization in poorly differentiated cell groups was cytoplasmic. The uptake of leucine was more constant and proved to be a useful indicator of viability. While this approach cannot give information with regard to differences in glycoprotein structure between non-malignant and malignant breast, it has been of value in determining the heterogeneity of tumour cells with regard to the enzymes involved in glycosylation. As such it would be of use in assessing the uniformity of response to agents modifying glycosylation.

Techniques for locating isotopically labelled schistosomula of Schistosoma mansoni in host tissues for ultrastructural investigations.

The use of 75 Selenomethionine labelled cercariae of Schistosoma mansoni for ultrastructural localization of parasites in host tissues has been evaluated. Both gamma counting of tissue, and autoradiography of resin-embedded tissue were successful. The autoradiographic technique was more sensitive and schistosomula were readily located in pulmonary tissue up to 24 days post-infection.

Site requirements and kinetics of immune-dependent elimination of intravascularly administered lung stage schistosomula in mice immunized with highly irradiated cercariae of Schistosoma mansoni.

Experiments were performed to compare the migration and survival of 75Se-labeled schistosomes, introduced by percutaneous cercarial exposure or by intravascular administration of 7-day-old lung stage schistosomula, in control and irradiated cercaria-immunized mice. Schistosomula were intravascularly introduced into the lungs, systemic organs and liver by injection via the femoral vein (FV), left ventricle (LV), and superior mesenteric vein (SMV), respectively. The fate of challenge larvae was examined by autoradiography of host tissues and by recovery of adult worms. It was found that both normal and immune elimination were site-dependent. In control mice 45%-60% of cercarial penetrants and lung schistosomula injected into the FV and LV were recoverable as adult worms, while a significantly greater number (70%-85%) were recoverable when lung schistosomula were injected into the SMV. In immunized mice, parasites introduced as either cercariae or FV-injected schistosomula were both highly sensitive to immune elimination. LV-injected schistosomula were also sensitive but to a slightly lesser degree. In contrast, schistosomula placed directly in the liver by SMV injection were totally insensitive to immune elimination. It was concluded that elimination of schistosomula in irradiated cercaria-immunized mice occurs in the lungs and/or in the systemic organs, but not in the liver. Also, it was concluded that immune elimination is not a rapid process, since more than 7 days were required after intravascular challenge for the development of demonstrable differences between control and immunized mice.

Timing of sulphogalactolipid biosynthesis in the rat testis studied by tissue autoradiography.

The testicular synthesis of sulphatoxygalactosylacylalkylglycerol (SGG) has been studied in the rat by autoradiography of frozen tissue sections following in vivo metabolic labelling. The results are consistent with the synthesis of this major mammalian germ-cell glycolipid at the zygotene and early pachytene stages of spermatogenesis. Further synthesis of SGG is prevented by the appearance of an inhibitor of galactolipid sulphotransferase activity at the mid-pachytene stage.

De novo cellular synthesis of sulfated proteoglycans of the developing renal glomerulus in vivo.

The site of cellular synthesis of glomerular proteoglycans was investigated in developing glomeruli of 4- to 5-day-old rats. [35S]Sulfate was administered intravenously and animals were sacrificed 15 min to 12 hr later. The outermost layers of the kidney cortices were utilized for characterization of proteoglycans and electron microscopic autoradiography. Sepharose CL-6B chromatography and cellulose acetate electrophoresis revealed that most (approximately equal to 96%) of the radioactivity was associated with heparan sulfate-proteoglycan synthesized during maturation of glomerular capillaries. Tissue autoradiography revealed the following: (i) during the S-shaped body stage, there is rapid incorporation of [35S]sulfate by mesenchymal cells into the cleft region (site for development of future glomerular extracellular matrices); (ii) during the precapillary stage, mesenchyme-derived cells showed higher incorporation of radioisotope than did epithelial cells; and (iii) during the mature capillary stage, all glomerular cell types (mesangial, endothelial, and epithelial) incorporated [35S]sulfate, incorporation by mesangial cells being the greatest. Radiolabeling was also higher in the mesangial matrix than in the glomerular basement membrane of peripheral capillary loops. Synthesis of a single major species of sulfated glycosaminoglycan by cells of different embryologic origin may be unique to glomerular capillaries.

Autoradiographic and biochemical studies of drug distribution in the liver. III. [14C]Aminotriazole.

Whole body autoradiography revealed that the distribution pattern of [14C]aminotriazole in the mouse liver was homogeneous after intravenous administration of the labeled agent and then became heterogeneous (or reticular). Microautoradiography by dry-mounting method revealed that the macroscopic heterogeneous pattern was due to the central localization of the radioactive compound in the hepatic lobule. The present studies indicated that the heterogeneous distribution could be explained as follows. The amount of [14C]aminotriazole circulated to the liver was large since the compound was not so significantly distributed in non-hepatic tissues: distribution pattern was homogeneous in the liver. This was shown by whole body autoradiography and radiometry of tissues. A part of [14C]aminotriazole radioactivity present in the liver was gradually bound covalently to hepatic macromolecules. This was shown by whole body autoradiography after whole body sections of the mouse were extracted by acid, and by the biochemical fractionation of the liver. The covalently bound radioactivity alone became apparent in centrilobular hepatocytes: the distribution was heterogeneous. This was shown by microautoradiography and by the finding that the elimination rate of the bound radioactivity was slower than that of unbound radioactivity.

Autoradiographic and biochemical studies of drug distribution in the liver. I. [14C]Dehydrocorydaline.

Whole body autoradiography revealed that the distribution pattern of [14C]dehydrocorydaline in the mouse and rat liver was heterogeneous (or reticular) regardless of time after intravenous administration of the labeled agent. Microautoradiography by dry-mounting method revealed that the macroscopic heterogeneous pattern was due to the periportal localization of the radioactive compound in the hepatic lobule. By comparison with [14C]salicylid acid, [14C]diphenylhydantoin and [14C]p-chlorophenoxyacetic acid whose distribution pattern are homogeneous in the liver, the present studies indicated that the existence and persistence of heterogeneous distribution of [14C]dehydrocorydaline in the liver had the following causes: 1. Shortly after intravenous administration, the amount of [14C]dehydrocorydaline circulated to the liver was greatly restricted by its significant distribution in non-hepatic tissues. This was shown by the whole body autoradiography, radiometry of tissues and quantitative comparison of volumes of distribution in non-hepatic tissues. Therefore, 2. perilobular hepatocytes alone could take up [14C]dehydrocorydaline and consequently, centrilobular cells were unavailable to it: heterogeneous distribution pattern is formed. This was shown by microautoradiography as described above, and by the rapid and significant uptake of [14C]dehydrocorydaline by isolated hepatocytes in vitro and by the liver to which the labeled agents were continuously administered in situ. It was also substantiated by the more homogeneous distribution pattern in the liver of the rat to which greater amount of [14C]dehydrocorydaline was gradually given into the portal vein and of the mouse with allyl formate-induced perilobular damage. 3. Redistribution of [14C]dehydrocorydaline scarcely occurred in the whole body and therefore radioactive substance was not significantly supplied to the liver: the distribution pattern remained unchanged. This was shown by the whole body autoradiography and radiometry of tissues.

The migration and survival of gamma-irradiatedSchistosoma mansonilarvae and the duration of host—parasite contact in relation to the induction of resistance in mice

The migration in mice of 20, 50 and 90 krad. 60Co-irradiated Schistosoma mansoni larvae, biosynthetically radio-isotope labelled with [75Se]-selenomethionine, was evaluated by autoradiography of compressed tissues and compared to the migration of non-irradiated 75Se-labelled larvae. The migration of 20 krad.-irradiated schistosomula between skin and lungs was slightly delayed but otherwise paralleled the migration of normal, non-irradiated schistosomula during the first 8 days following exposure. By day 8 over 90% of both non-irradiated and 20 krad.-irradiated organisms were located in the lungs. In contrast to non-irradiated organisms, however, only a small proportion of 20 krad. organisms migrated to the liver. The delay in migration between skin and lungs was more pronounced with 50 krad.-irradiated schistosomula. Nevertheless, 45-93% of 50 krad.-irradiated organisms migrated to the lungs by 8 days post-exposure. Over 90% of the 50 krad. larvae detected in the mouse on day 21 were in the lungs; no more than an occasional 50 krad.-irradiated organism was ever detected in the liver. In three experiments, over 85% of the 90 krad.-irradiated organisms were retained in the skin; in a fourth experiment about half of the 90 krad.-irradiated organisms migrated as far as the lungs. As with 50 krad. organisms, only an occasional 90 krad. organism was ever detected in the liver. Removal of the skin exposure site within the first 4 days of immunization with either 50 or 90 krad.-irradiated cercariae completely blocked the induction of resistance. Removal between the 4th and 6th days gave variable results. Mice had to be in contact with the irradiated larvae for a minimum of 8-11 days to stimulate a level of resistance comparable to that of mice whose immunization site was not removed.

Stimulation of wound healing, using brain extract with fibroblast growth factor (FGF) activity: III. Electron microscopy, autoradiography, and ultrastructural autoradiography of granulation tissue

A bovine brain fraction with FGF activity (fibroblast growth factor) increased the formation of granulation tissue of rats in an experimental wound healing model. It was characterised by intensified formation of new capillaries, stimulation of the synthesis function of fibroblasts and myofibroblasts, and proliferation of freshly developed granulation tissue. The results obtained from light microscopy and autoradiographic electron microscopy with 3H-thymidine suggested significant increase in the labelling of activated, undifferentiated endothelial cells in vascular sprouts and of differentiated endothelial cells in developed capillaries. FGF S3 appeared to speed up the differentiation of freshly formed endothelial cells. Also labelled were pericytes and other fibroblastoid cells, undifferentiated mesenchymal cells, juvenile undifferentiated fibroblasts, mature fibroblasts, myofibroblasts, and macrophages. Pericytes are presumed to be preferential precursors of myofibroblasts, on account of their occurrence in time and similar ultrastructural parameters. Close neighbourhood of lymphocytes and labelled fibroblasts gave rise to the assumption of specific effects, such as stimulation by lymphokines.

Alpha-particle autoradiography in CR-39: a technique for quantitative assessment of alpha-emitters in biological tissue

The techniques for alpha -particle autoradiography based on the plastic nuclear track detector CR-39, previously reported, have been developed considerably. The techniques are applied to alpha -autoradiography of human lung tissue in particular but are applicable to any biological tissue. The most important developments are: (i) Improvements in the manufacture and pre-etching of the plastic. These allow activities as low as approximately 10-15 Ci g-1 to be determined. (ii) High resolution alpha -particle spectroscopy in CR-39 plastic based on the analysis of the structure of the etched track. This enables the energy of individual alpha -particles to be determined to approximately 35 keV. (iii) Calculation of the effective thickness of tissue sampled by the plastic. This relates the tissue activity to the track density on the plastic. (iv) A deconvolution analysis which takes the distribution of track length and dip angle in the plastic and determines the alpha -particle range spectrum and distribution of tissue activity with height above the plastic surface. This enables both the absolute abundance and the microdistribution of alpha -active nuclides present to be determined. (v) The analysis of radon diffusion in tissue to determine the mean random diffusion distance in tissue and plastic.

Functional neuroanatomical mapping in insects by [ 3H]2-deoxy-D-glucose at electron microscopical resolution

Electron microscope autoradiography of insect nervous tissue prepared for ‘activity staining’ by the deoxyglucose method provides information on metabolic activity of small identifiable nerve fibers. Grain counts over two sets of axon profiles selected for high and low radioactivity, respectively, indicate that the dynamic range of the present method comprises about one order of magnitude. It is demonstrated that, although a small fraction of the radioactive label is lost during flotation of the thin sections on a water surface, the distribution of the remaining label is not appreciably altered and reflects stimulus-induced nervous activity.

Can Corneal Graft Be Used as a Steroid Reservoir?

Because we found that steroid can retard autolysis of the stored cornea, we planned to add the agent to the tissue culture medium used for storing isolated corneas. The purpose of our experiment was to determine the rate at which steroid enters the stored cornea and the rate at which it will be released from the graft postoperatively. Rabbit corneas were stored for 2 days at 4°C in MK medium containing radiolabeled hydrocortisone. Discs from these corneas were grafted onto allogeneic animals. Identical discs were used for determining the amount of the drugs present per disc by scintillation counting. Autoradiography of the donor tissue and recipient cornea containing the graft was also done. Both scintillation counting and autoradiography showed that hydrocortisone entered the corneal tissue stored at 4°C and that by 24 hours postoperatively, most of the steroid disappeared from the graft and the graft-bed. In view of the rapid disappearance of steroid from the graft and graft-bed postoperatively, it appears that corneal donor tissue stored in steroid solution should not create a problem with respect to wound healing and microbial invasion.

Lyt phenotype and lectin binding properties of mouse lymphocytes which enter lymph nodes.

The objective of this study was to determine how Lyt phenotype and specific lectin binding properties influenced the ability of thymus-derived lymphocytes to enter lymph nodes. Both short term 51Cr quantitative migration experiments and tissue autoradiography experiments using 3H-adenosine labeled cells were done. Results demonstrate that Lyt phenotype per se is insufficient to determine migration pattern and that other factors, including surface carbohydrate composition, are involved.KeywordsLymph NodeInguinal Lymph NodeMouse LymphocyteEnter Lymph NodeNylon Wool ColumnThese keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

The microdistribution of α-active nuclei in the human lung I - techniques

ABSTRACT We present further developments of our technique for high sensitivity high resolution α-particle autoradiography of human lung tissue using CR-39 plastic. High resolution measurements of α-track length and dip angle on the auto radiograph are made. The distributions of these quantities allow the distribution of α-active elements present to be calculated. Any localisation of height emission up to 30 μm above the plastic surface can also be found. Using a distribution of distance to nearest neighbour of α-tracks on the autoradiograph the Radon diffusion coefficient is measured. This information is necessary in order to calculate the thickness of tissue sampled for Rn222 and daughter nuclei.

Distribution of pressure-induced fast axonal transport abnormalities in primate optic nerve. An autoradiographic study.

The distribution of transport abnormalities in primate optic nerve from eyes subjected to five hours of pressure elevation (perfusion pressure of 35 mm Hg) was studied. Tissue autoradiography and electron microscopy were used to localize regions of the lamina cribrosa with increased transport interruption. A preferential involvement by this transport abnormality involved the superior, temporal, and inferior portions, to the exclusion of the nasal portion, of the optic nerve head. This observation supports the hypothesis that transport interruption seen in this model may be pertinent to the study of clinical glaucomatous neuropathy.

The fate of isoprenaline in the isolated rabbit aorta. Radiochemical and morphologic observations.

The fate of isoprenaline (ISO) was studied in the intact rabbit aorta, as well as in the isolated adventitia and isolated media, by means of liquid scintillation counting of 3H-ISO and 3H-O-methylisoprenaline (OMI) as well as by autoradiography of tissues incubated with 2μM ISO. The 3 preparations accumulated and O-methylated ISO; O-methylation was proportionally higher in the intact vessel than in its constituents. The isolated adventitia differed from the other preparations in reaching a steady-state of cortexone-resistant accumulation after 8 min of incubation and in exhibiting an O-methylating capacity which was partly resistant to the COMT inhibitor, U-0521. The effect of cocaine gave evidence for a participation of neuronal uptake in the accumulation of the amine in the intact aorta. In the media, most of the accumulation occurred in smooth muscle cells and was reduced by cortexone and increased by U-0521; elastin and collagen, present both in the media and the adventitia, showed accumulation which was not affected by the inhibitory drugs studied. The results show that in the rabbit aorta there is a corticosteroid-resistant uptake mechanism (which predominates in the adventitia) which involves structures devoid of COMT activity. Furthermore, smooth muscle cells represent, in the media, the extraneuronal metabolizing site of loss. The adventitia is a complex layer, with different types of cells which may intervene in accumulation and metabolism of ISO. Therefore, it is concluded that the isolated media represents an acceptable model for the study of both corticosteroidsensitive and-resistant extraneuronal mechanisms, whereas the isolated adventitia is characterized by the presence of neuronal and extraneuronal mechanisms.

A technique for high-sensitivity alpha autoradiography of bronchial epithelium tissue

A technique for low-level alpha-particle autoradiography of bronchial epithelium utilising the plastic track detector, CR-39, is described. This plastic is new to the field of nuclear track detection and is very sensitive to alpha -particles. The recording properties of CR-39 for alpha -particles are described in detail and the technique for autoradiography discussed. This technique includes two methods of background reduction enabling activities as low as alpha -particles cm-2 to be detected. The location of the point of emission of an alpha -particle from the tissue surface can be determined to an accuracy of a few mu m. Determination of the lower limit to the energy of individual alpha -particles is possible from measurements of their range in the plastic. Examples are given of the determination of the microdistribution of alpha -particle active nuclei in bronchial tissue including the observation of two 'hot-spots' in an epithelium sample which are attributed to the presence of small particles of uranium with its daughter products.