Morphological and biochemical alterations in growing rats induced by etretinate

Etretinate is an aromatic retinoid extensively used on Dermatology. Its toxic effects, however, reduce its application from a clinical point of view. In the present paper, we study etretinate intoxication of 48 growing Wistar rats. The intoxication was for 12 weeks using etretinate doses of 0.5 and 6 (mg/kg)/day. The concentrations of etretinate in plasma and liver were determined. Total seric cholesterol and triglyceride concentrations were analyzed. Structural and ultrastructural histological studies of liver samples were carried out. Continuous etretinate ingestions seem to produce an alteration in the detoxication of enzymatic complexes in the growing rats with both the concentrations, due to the increase in etretinate blood plasma observed during the study. There is a relationship between the etretinate dose and its blood plasma concentration and toxic effect, but there is not with etretinate concentration in the liver. The blood plasma concentration of cholesterol and triglycerides is not related to histological liver lesions. The histological study confirms hepatotoxicity with both doses. Nevertheless, the anatomopathological lesions observed do not seem to be related to the blood plasma and liver etretinate concentrations.     

Functional and structural alterations induced by copper in xanthine oxidase

Xanthine oxidase （XO）, a key enzyme in purine metabolism, produces reactive oxygen species causing vascular injuries and chronic heart failure. Here, copper＇s ability to alter XO activity and structure was investigated in vitro after pre-incubation of the enzyme with increasing Cue＋ concentrations for various periods of time. The enzymatic activity was measured by following XO-catalyzed xanthine oxidation to uric acid under steady-state kinetics conditions. Structural alterations were assessed by electronic absorption, fluorescence, and circular dichroism spectroscopy. Results showed that Cu^2＋ either stimulated or inhibited XO activity, depending on metal concentration and preincubation length, the latter also determining the inhibition type. Cu^2＋-xo complex formation was characterized by modifications in XO electronic absorption bands, intrinsic fluorescence, and α-helical and β-sheet content. Apparent dissociation constant values implied high- and low-affinity Cu^2＋ binding sites in the vicinity of the enzyme＇s reactive centers. Data indicated that Cu^2＋ binding to high-affinity sites caused alterations around XO molybdenum and flavin adenine dinucleotide centers, changes in secondary structure, and moderate activity inhibition; binding to low affinity sites caused alterations around all XO reactive centers including FeS, changes in tertiary structure as reflected by alterations in spectral properties, and drastic activity inhibition. Stimulation was attributed to transient stabilization of XO optimal conformation. Results also emphasized the potential role of copper in the regulation of XO activity stemming from its binding properties.     

Early biochemical alterations induced by 2-acetylaminofluorene in rat liver

Livers from rats fed the carcinogen 2-acetylaminofluorene (AAF) were analyzed at weekly or semiweekly intervals to correlate appearance of enzymatic markers in total liver homogenates with histochemical events accompanying formation of hyperplastic liver nodules. gamma-Glutamyltranspeptidase (gamma-GT)-positive foci appeared by day 11 and visible nodules were present by days 28-35. Specific activity of homogenate gamma-GT increased in parallel to formation of hyperplastic foci and nodules, declined and then rose again to 20-fold that of controls by day 77. Specific activity of ornithine decarboxylase increased in advance of that of gamma-GT, to a level of 8-fold above control during the period of formation of hyperplastic foci. An early response was a 2-fold rise in the specific activity of nucleoside diphosphate phosphatase during the first week of carcinogen administration. The specific activity of 5'-nucleotidase, known to increase during liver regeneration, declined as the animals aged and was not increased by the dietary AAF. The enzymatic alterations induced by AAF could not be mimicked by cell proliferation, diet stress or the hepatotoxicity induced by feeding 1.87% 4-acetamidophenol.     

Copper induced alterations of biochemical parameters in the gill and plasma of Oreochromis niloticus

The main objective of this study was to determine the effects of copper exposure on copper accumulated in branchial tissue, gill Na+/K+-ATPase activity and plasma Na+, Cl-, osmolality, protein, glucose and cortisol, in Oreochromis niloticus. Fish were experimentally exposed to 40 and 400 microg L(-1) of waterborne copper and sacrified after 0, 3, 7, 14 and 21 days. Copper accumulation and Na+/K+-ATPase activity were determined in branchial tissue, whereas osmolality, Na+, Cl-, protein, glucose and cortisol concentrations were measured in plasma samples. Gill copper accumulation increased linearly with exposure time and concentration, whereas gill Na+/K+-ATPase activity was maximally inhibited after 3 days of exposure and showed a significant negative correlation with copper tissue levels. Plasma Cl- values decreased with time of exposure but only at 400 microg L(-1) of copper. Plasma Na+, protein and osmolality decreased with exposure time at the highest copper concentration tested, whereas at 40 microg L(-1) of copper this effect was only observed after 21 days of exposure. Plasma glucose and cortisol levels increased in a dose and time dependent manner, while showing complex fluctuations during the intermediate exposure times. In conclusion, copper induces an early maximum inhibition of gill Na+/K+-ATPase activity in O. niloticus. The subsequent slow decrease in ion plasma levels was related to compensatory mechanisms involving a non-specific stress response that appeared overcome at long-term exposures.     

Effect of isoflavone from Flemingia vestita (Fabaceae) on the Ca2+ homeostasis in Raillietina echinobothrida, the cestode of domestic fowl

The alcoholic crude root-peel extract of Flemingia vestita and its major isoflavone, genistein, have been shown to have a vermifugal/vermicidal effect by causing a flaccid paralysis accompanied by alterations in the structural architecture of the tegumental interface and metabolic activity in Raillietina echinobothrida, the cestode of domestic fowl. In the present study, the crude root-peel extract and pure genistein were tested in vitro with respect to Ca2+ homeostasis and the occurrence of some metal ions was detected in the parasite. Live cestodes were incubated in pre-defined concentrations of the crude root-peel extract, genistein and praziquantel (as reference drug), till the paralysis time with simultaneous maintenance of respective controls. In the parasite tissue, a significant amount of Ca2+ (approximately 400 microg/g dry tissue wt) was found to be present besides magnesium, iron, zinc, lead and chromium, whilst manganese, cadmium and nickel were below the level of detection. The Ca2+ concentration was decreased significantly by 39%-49%, in the parasite tissue exposed to the test materials in comparison to the respective controls. There was also an increase in Ca2+ efflux by 91%-160% into the culture medium under similar treatments. The changes in Ca2+ homeostasis may be related to the rapid muscular contraction and consequent paralysis in the parasite due to the anthelmintic stress caused by the phytochemicals of F. vestita.     

Ultrastructural and biochemical alterations induced by propranolol treatment in experimental myocardial ischaemia

The effect of propranolol within the first ten days after a transient experimental myocardial ischaemia was investigated. As compared to the untreated control lot, the ultrastructural and biochemical lesions appeared to be much reduced and within reversible limits. Worthy of note was the slight affection of mitochondria revealed by the level of mitochondrial enzymes, the presence of lysosomal and phagolysosomal structures and capillaries of normal aspect, and the absence of cytolysis. The favourable effect of this beta-blocking drug might be attributed to its protective action on the cellular and capillary membrane systems.     

Reactive oxygen species induced structural alterations of substance P

Substance P (SP(1-11)) was exposed to a continuous flux of superoxide (O2-) or hydroxyl radicals ((.)OH) in a hypoxanthine (HX)/xanthine oxidase (86 mU) system in the presence of 1 mM deferoxamine and 40 mM D-mannitol or 50 muM FeCI(3). 6H(2)O and 50 muM EDTA, respectively. O2- caused fragmentation between the Phe(7) and Phe(8), whereas (.)OH induced cleavage also between the Phe(8) and Gly(9). Reactive oxygen species H(2)O(2) and HCIO did not cause fragmentation, but modification of the amino acid side chains and/or aggregation with altered hydrophobicity in reverse phase high performance liquid chromatography compared to native SP(1-11). Furthermore, exposure of SP(1-11) to phorbol myristate acetate preactivated neutrophils resuited in products similar to those observed upon exposure to superoxide or hydroxyl radicals in a cell-free HX/xanthine oxidase system. This study suggests that, in contrast to rigid proteins, fragmentation is relatively easily induced in a small peptide like SP(1-11), perhaps due to strain on the peptide and t-carbon bonds caused by the movable, random coil configuration acquired by SP(1-11) in an aqueous solution. Oxidative modification might modulate paracrine actions of SP(1-11) at site of inflammation.     

Spatial-resolved analysis of histological and biochemical alterations induced by water-soaking in melon fruit

Water-soaking, a physiological disorder characterised by a glassy texture of the flesh, depreciates greatly the commercial quality of early-season Charentais cantaloupe melons ( Cucumis melo L. cv. Talma). Although it is accepted that the genotype and a number of physiological and environmental factors play a role in the development of the syndrome, the intimate mechanisms responsible for water-soaking remain unknown. We report here on an integrated study of the development of water-soaking in fruit. Using nuclear magnetic resonance (NMR) imaging, we have shown that water mobility increased in the diseased tissues. Alteration of the cell wall and the presence of large intercellular spaces were correlated with a severe depletion of cell wall calcium. Water-soaking developed during the late stages of fruit ripening, but no correlation was found with ethylene biosynthesis. Thus, fruits in which ethylene action was blocked by 1-methylcyclopropene remained sensitive to water-soaking. Moreover, the expression of two genes encoding key enzymes in ethylene biosynthesis remained unchanged in response to water-soaking. The major changes observed concerned a protein implicated in calcium signalling processes. While the amount of total calmodulin, the ubiquitous calcium binding protein, was not modified, a particular calmodulin-binding protein (CaM-BP) was absent in water-soaked but not in sound mature tissues. This CaM-BP may be a marker or a determinant of this physiological disorder.     

Mercury and cadmium induced structural alterations in the taste buds of the fish Alburnus alburnus

The ultrastructural damages of the taste buds of the fish, Alburnus alburnus were studied after applying 0.05 microM and 0.5 microM mercury chloride as well as 0.1 microM and 1 microM cadmium chloride. The most conspicuous alterations were induced during the first week of heavy metal exposition. The main structural alterations are: 1) the swelling of sensory microvilli and cilia; 2) the extreme dilation of the rER tubules and nuclear membranes, which is most expressed after cadmium exposition; 3) the increase in the number of lysosomes and dens bodies, which is more expressed after mercury exposition; 4) the swelling of the innervating nerve fibres at the synaptic areas of the taste buds, especially after mercury exposition. The damaging processes induced by the applied dose of heavy metals did not increase after the first week of exposition. The taste buds showed regenerated structural appearance after two weeks of exposition to 1 microM CdCl2, while the evoked structural alterations could be detected even after two weeks of exposition to 0.5 microM HgCl2.     

X-rays induced alterations in mechanical and biochemical properties of isolated SH-SY5Y nuclei

BackgroundThe use of ionizing radiations in radiotherapy is an effective and very common cancer treatment after surgery. Although ionizing-radiation DNA damages are extensively investigated, little is known about their effects on the other nuclear components, since their variations when studied in whole cells can be difficult to decouple from those of the cytoplasmatic structures. The organization of nuclear components plays a functional role since they are directly involved in some of the nuclear response to chemical or physical stimuli. For this reason, studying the X-ray effects on nuclear components is a crucial step in radiobiology.Materials and methodsWe have used Atomic Force Microscopy (AFM) and micro-FTIR to examine the biomechanical and biochemical properties of hydrated fixed nuclei isolated from neuroblastoma (SH-SY5Y) cells irradiated by 2, 4, 6 and 8 Gy X-ray doses.ResultsThe experimental results have shown that, already at 2 Gy irradiation dose, the nuclei exhibit not only a DNA damage, but also relevant alterations of lipid saturation, protein secondary structure arrangement and a significant decrease in nuclear stiffness, which indicate a remarkable chromatin decondensation.Conclusions and general significanceThe present work demonstrates that a multi-technique approach, able to disclose multiple features, can be helpful to achieve a comprehensive picture of the X-ray irradiation effects of the nuclear components and distinguish them from those occurring at the level of cytoplasm.     

Differential binding of tetracyclines with serum albumin and induced structural alterations in drug-bound protein

Interaction of tetracycline (TC) derivatives viz. oxytetracycline, doxycycline, demeclocycline and chlorotetracycline with bovine serum albumin (BSA) and concomitant changes in protein conformation were studied using fluorescence quenching and circular dichroism measurements. Fluorescence data revealed the presence of one to three binding sites on BSA for different TC derivatives. Binding studies with the marker ligands, warfarin and bilirubin, elucidated site-I as a primary binding site for TCs on albumin. Scatchard analysis revealed the binding affinity (K_a) and capacity (n) for these derivatives vary in the range from 0.8 to 3.2×10⁶ l/mole and 1.3–3.4, respectively. Significant reduction (60–45%) in secondary structure (-helical content) of BSA was noticed upon interaction with different TC derivatives in presence of Cu (II) ions. High affinity binding of TCs with BSA signifies drug stability. However, excessive binding at higher TC concentrations in combination with Cu (II) induces conformational change in protein structure, which may exert detrimental effect on cellular protein.     

A relational theory of the structural changes induced in biological systems by alterations in environment

It is shown that a wide variety of structural alterations in both the “metabolic” and “genetic” apparatus of ( , ℜ)-systems can result from specific changes in the environment of such systems. A number of specific examples are investigated in order to demonstrate the scope of these alterations. Certain biological applications of this discussion are suggested, including a suggestion for a possible interpretation of the mitotic cycle. This research was supported by the United States Air Force through the Air Force Office of Scientific Research of the Air Research and Development Command, under Contract #AF 49 (638)-917.     

Sesamin attenuates behavioral,biochemical and histological alterations induced by reversible middle cerebral artery occlusion in the rats

Restoration of blood flow to an ischemic brain region is associated with generation of reactive oxygen species (ROS) with consequent reperfusion injury. ROS cause lipid peroxidation, protein oxidation, and DNA damage, all of which are deleterious to cells. So diminishing the production of free radicals and scavenging them may be a successful therapeutic strategy for the protection of brain tissue in cerebral stroke. The present study investigated the neuroprotective effect of sesamin (Sn) to reduce brain injury after middle cerebral artery occlusion (MCAO). The middle cerebral artery (MCA) of adult male Wistar rat was occluded for 2 h and reperfused for 22 h. Sesamin is the most abundant lignan in sesame seed oil is a potent antioxidant. Sesamin (30 mg/kg) was given orally twice, 30 min before the onset of ischemia and 12 h after reperfusion. The initial investigations revealed that sesamin reduced the neurological deficits in terms of behavior and reduced the level of thiobarbituric acid reactive species (TBARS), and protein carbonyl (PC) in the different areas of the brain when compared with the MCAO group. A significantly depleted level of glutathione and its dependent enzymes (glutathione peroxidase [GPx] and glutathione reductase [GR]) in MCAO group were protected significantly in MCAO group treated with sesamin. The present study suggests that sesamin may be able to attenuate the ischemic cell death and plays a crucial role as a neuroprotectant in regulating levels of reactive oxygen species in the rat brain. Thus, sesamin may be a potential compound in stroke therapy.     

Microdissection studies of the structural alterations induced in rat kidneys by experimental postischemic acute renal failure

A unique opportunity presented itself for a morphologic study of experimental unilateral acute renal failure (ARF) in male rats. The ARF had been induced in the rats by temporary occlusion (1h) of the left renal artery. Twenty-nine rats were divided into subsets as follows: 2-3 h, 24 h, 1 week, 2, 4, 8, and 12 weeks following release of occlusion. Microdissection showed a heterogeneous population of abnormally structured proximal tubules in which the regressive lesions of tubular necrosis were combined with the progressive reaction of repair. The lesions demonstrated are reminiscent of those which have been described in ARF in the human and in experimental animals. Many proximal tubules in the 2- to 3-hour subset presented 1-3 disruptive lesions (DLs) while greater numbers of proximal tubules from the 24-hour group presented 1-5 DLs. Many proximal tubules presented no DLs, but nearly all from the 24-hour subset (97-100%) displayed a squamate appearance which paralleled and was caused by acute tubular necrosis. At 1 week, a dilated pars recta was common, but by this time, the squamate pattern had disappeared. Many casts were present. At 2 weeks, many fewer casts were present in proximal tubules and none were seen at 4, 8 or 12 weeks. The nephrons, particularly the proximal tubules, presented a variety of structural alterations at 2, 4, 8 and 12 weeks. Changes of special interest include (1) the presence of swan-necks; (2) a distinctive squamate appearance of the proximal tubules in the animals killed at 24 h; (3) a spiral, curled appearance caused by differential hyperplasia in animals at 4, 8 and 12 weeks, and (4) a tendency for ischemic lesions to involve all layers of the renal cortex.     

Phospholipid supplementation reverses behavioral and biochemical alterations induced by n-3 polyunsaturated fatty acid deficiency in mice

This study investigated the effects of a diet deficient in alpha-linolenic acid followed or not by supplementation with phospholipids rich in n-3 polyunsaturated fatty acids (PUFA) on behavior and phospholipid fatty acid composition in selected brain regions. Three weeks before mating, two groups of mice were fed a semisynthetic diet containing both linoleic and alpha-linolenic acid or a diet deficient in alpha-linolenic acid. Pups were fed the same diet as their dams. At the age of 7 weeks, a part of the deficient group was supplemented with n-3 PUFA from either egg yolk or pig brain phospholipids for 2 months. In the open field, rearing activity was significantly reduced in the deficient group. In the elevated plus maze (anxiety protocol), the time spent on open arms was significantly smaller in deficient mice than in controls. Using the learning protocol with the same task, the alpha-linolenic acid deficiency induced a learning deficit. Rearing activity and learning deficits were completely restored by supplementation with egg yolk or cerebral phospholipids, though the level of anxiety remained significantly higher than that of controls. There were no differences among the 4 diet groups for either the Morris water maze or passive avoidance. In control mice, the level of 22:6 n-3 was significantly higher in the frontal cortex compared to all other regions analysed. The frontal cortex and the striatum were the most markedly affected by the deficiency. Supplementation with phospholipids restored normal fatty acid composition in brain regions except for frontal cortex. Egg yolk or cerebral phospholipids are an effective source of n-3 PUFA for reversing behavioral changes and altered fatty acid composition induced by a diet deficient in n-3 PUFA.