Mitochondrial Matrix Granules Research Articles

Sperm characters in the Hemiuridae (Digenea): first data on Aphanurus stossichii (Aphanurinae) and Ectenurus lepidus (Dinurinae)

The present work provides the first ultrastructural analysis of spermatozoa of two digeneans (Aphanurus stossichii (Monticelli, 1891) and Ectenurus lepidus Looss, 1907) belonging to the unexplored subfamilies of the Hemiuridae, namely, the Aphanurinae and the Dinurinae. In March 2019, these hemiurids were collected respectively from the digestive tract of the bogue Boops boops (Teleostei, Sparidae) and the Atlantic horse mackerel Trachurus trachurus (Teleostei, Carangidae) captured in the coastal zone of the Mediterranean Sea, off La Chebba (Tunisia). The ultrastructural study reveals that both spermatozoa exhibit the Bakhoum et al.'s type II of the digenean sperm cells characterized by the presence of two 9+'1' axonemes, an external ornamentation of the plasma membrane not associated with cortical microtubules and located in the anterior part of the spermatozoon, a single bundle of cortical microtubules, the maximum number of cortical microtubules located in a middle part of the sperm cell, and one mitochondrion. Moreover, they share several ultrastructural features with the studied spermatozoa of Hemiuridae such as the presence of two axonemes with the 9+'1' trepaxonematan pattern, a reduced number of parallel cortical microtubules organized into one field with their maximum number located in the median (A. stossichii) or posterior (E. lepidus) part of the spermatozoon, an external ornamentation of the plasma membrane in the anterior part of the spermatozoon, one mitochondrion, a nucleus, and a small amount of glycogen granules. However, the two studied hemiurids could be distinguished by the morphology of the anterior and posterior spermatozoon extremities and the presence of mitochondrial matrix granules in A. stossichii.

Sperm cell ultrastructure of the haplosplanchnid Haplosplanchnus caudatus (Platyhelminthes: Digenea) – An intestinal parasite of Mugil cephalus, and its potential phylogenetic application

The ultrastructural characters of mature spermatozoon of the digenean Haplosplanchnus caudatus were described by means of transmission electron microscopy (TEM). Live parasites were collected from the digestive tract of flathead grey mullet Mugil cephalus (Teleostei: Mugilidae) caught off the Gulf of Gabes in La Chebba (Tunisia). The present study provides the first ultrastructural data on a digenean belonging to the superfamily Haplosplanchnoidea. The spermatozoon of H. caudatus is a filiform cell which exhibits: (i) two axonemes with the 9+‘1’pattern of trepaxonematan Platyhelminthes having about the same length, (ii) an external ornamentation of the plasma membrane associated with cortical microtubules and located in the anterior part of the sperm cell, (iii) spine-like bodies, (iv) two mitochondria containing large mitochondrial matrix granules, (v) a nucleus, and (vi) granules of glycogen irregularly distributed. Moreover, the spermatozoon of H. caudatus displays the Quilichini et al.’s cryptogonimid type of posterior extremity. The mitochondrial matrix granules are described for the first time in a digenean sperm cell.

Effects of chronic exposure to environmentally relevant concentrations of waterborne depleted uranium on the digestive tract of zebrafish, Danio rerio

Uranium is a naturally occurring element, but activities linked to the nuclear fuel cycle can increase background levels in the surrounding waters. For this reason it is important to understand how this affects organisms residing in the water column. The objective of this study was to assess histopathological effects of uranium on the gut wall of a widely used model organism: zebrafish, Danio rerio. To this end we exposed zebrafish to 84 and 420 nM depleted uranium for over a month and then examined the histology of intestines of exposed individuals compared to controls.The gut wall of individuals exposed to 84 and 420 nM of uranium had large regions of degraded mucosa. Using transmission electron microscopy (TEM) coupled to energy-dispersive X-ray spectroscopy microanalysis (EDX) we found that uranium induced a decrease in the amount of calcium containing mitochondrial matrix granules per mitochondria. This is suggestive of perturbations to cellular metabolism and more specifically to cellular calcium homeostasis. TEM-EDX of the gut wall tissue further showed that some uranium was internalized in the nucleus of epithelial cells in the 420 nM treatment.Fluorescent in situ hybridization using specific probes to detect all eubacteria was performed on frozen sections of 6 individual fish in the 84 nM and 420 nM treatments. Bacterial colonization of the gut of individuals in the 420 nM seemed to differ from that of the controls and 84 nM individuals. We suggest that host–microbiota interactions are potentially disturbed in response to uranium induced stress.The damage induced by waterborne uranium to the gut wall did not seem to depend on the concentration of uranium in the media. We measure whole body residues of uranium at the end of the experiment and compute the mean dose rate absorbed for each condition. We discuss why effects might be uncoupled from external concentration and highlight that it is not so much the external concentration but the dynamics of internalization which are important players in the game.

Electron microscopic findings in non‐alcoholic fatty liver disease: Is there a difference between hepatosteatosis and steatohepatitis?

Non-alcoholic fatty liver disease has long been accepted as benign; however, recent evidence suggests that the disease may progress to cirrhosis and hepatocellular carcinoma, although the natural course of the disease is still unclear. This study was designed to comparatively evaluate electron microscopic features of non-alcoholic fatty liver (NAFL) and non-alcoholic steatohepatitis (NASH). Quantitative and semi-quantitative ultrastructural evaluations were performed on liver biopsies from 23 patients, 10 with NAFL and 13 with NASH. No statistically significant difference was noted between NAFL and NASH patients in ultrastructural features of hepatocytes including megamitochondria, intramitochondrial crystalline inclusions, mitochondrial matrix granules, foamy cytoplasmic appearance, electron-lucent and glycogen-containing nuclear regions, lipofuscin granules, or an increased frequency of vesicles containing electron-dense material in peribiliary Golgi zone; however, the mitochondrial diameter was significantly higher in the NASH patients. Intercellular distance and microvilli between hepatocytes, collagen and electron-dense material accumulation in the space of Disse, electron-dense material accumulation and microvillus density in bile canaliculi did not differ significantly between the groups. Our data show that, although NAFL and NASH can be distinguished by their distinct light microscopic features, ultrastructural characteristics are similar, which suggests that NAFL may also have the potential to progress to fibrosis and cirrhosis like NASH.

FINE STRUCTURAL MODIFICATIONS OF LIVER, PANCREAS AND BROWN ADIPOSE TISSUE MITOCHONDRIA FROM HIBERNATING, AROUSING AND EUTHERMIC DORMICE

An ultrastructural and morphometric study was performed on mitochondria of euthermic, hibernating and arousing hazel dormice (Muscardinus avellanarius), in order to investigate possible modifications during the seasonal cycle. Hepatocytes, pancreatic acinar cells and brown adipocytes were considered. Our results demonstrated that: (1) the general morphology of mitochondria of all cell types shows slight modifications during the seasonal cycle; (2) mitochondrial size and inner membrane length significantly increase from euthermia to hibernation and decrease upon arousal in all cell types; (3) mitochondrial matrix granules drastically increase in number during hibernation and decrease upon arousal in hepatocytes and pancreatic acinar cells, whereas they do not change in brown adipocytes. These structural modifications are probably related to the changes in cellular energy needs during the euthermia–hibernation–arousal cycle.

Mitochondrial matrix granules: their behavior during changing metabolic situations and their relationship to contact sites between inner and outer mitochondrial membranes.

Since their discovery in the early fifties mitochondrial granules have been the subject of many researches. Some twenty years ago two hypotheses on their function were introduced. Peachey thought that the granules were a sink of cations and that they would eventually regulate the concentrations of these ions. Alternatively, Barnard thought that the granules were precursors of the mitochondrial inner membrane. There are only a few data on organic constituents of the granules. Phospholipids (e.g., cardiolipin) glycoprotein or lipids, calcium precipitable lipoprotein, cytochrome c oxidase seem to be present in the granules. There has been much debate on whether calcium is present or not. Reports are mostly based on X-ray microanalysis, the result of which depends on preparation techniques. In heart muscle in stimulating situations the NMG (native matrix granules) move towards the inner membrane and are incorporated in it. They appear to create contact sites between inner and outer mitochondrial membranes in which enzymes can function efficiently. It is hypothetized that the system, NMG-contact sites, forms the structural basis of a regulatory mechanism, by which cells can cope with a high and sudden energy demand.

Ultrastructural characteristics of atrial, ventricular and subendocardial (Purkinje) cells of the fruit-eating bat Eidolon helvum.

The ultrastructure of the myocardium of the large (285 gm) fruit-eating bat (Eidolon helvum) provides further evidence that, among mammalian species, different types of cardiac myocytes are morphologically heterogeneous. Atrial cells of this species, like those of smaller bats previously studied, have few of the natriuretic hormone-containing granules. These granules are also small, with a mean diameter of 176 +/- 3.9 nm, in comparison with those of the rat (230 +/- 5.8 nm) and mouse (211 +/- 4.7 nm). In all types of myocytes of the bat investigated, mitochondrial matrix granules are more prominent than in those of the rat and mouse. Also distinctive are the Purkinje cells which, aside from their large size (30 microns) and lack of T-tubules, have atrial granule-like inclusions. These cells are well endowed with mitochondria (28% of cytoplasmic volume) and with lipid bodies but have few glycogen granules. Their myofibrillae (33%) are well aligned. The ventricular cells (average diameter 14 microns) have nearly equal volumes of mitochondria (41%) and myofibrils (47%). Mitochondrial cristae, however, are not compactly organized. Lipid material is found in the matrix of the mitochondria. Extensive evaginations of endothelial cells of the bat endocardium appear continuous with endothelial vacuoles. In these endothelial cells, there are also many dense granules of varying shapes and sizes.

The Occurrence of Intramitochondrial Ca2+ Granules in Valproate‐Induced Liver Injury

Two epileptic patients treated with anticonvulsants of valproic acid, phenobarbital, nitrazepam, and diphenylhydantoin developed anorexia, convulsions, and unconsciousness. The liver biopsy samples showed degenerated hepatocytes containing enlarged mitochondria with a distorted matrix. The matrix granules in the mitochondria were enlarged in size as compared with those of chronic hepatitis (P0 = 0.0009). X‐ray microanalysis has revealed the presence of calcium in large mitochondrial matrix granules. Both patients were thought to have valproic acid intoxication because they recovered quickly after the withdrawal of valproic acid.

The Occurrence of Intramitochondrial Ca2+ Granules in Valproate-Induced Liver Injury

Two epileptic patients treated with anticonvulsants of valproic acid, phenobarbital, nitrazepam, and diphenylhydantoin developed anorexia, convulsions, and unconsciousness. The liver biopsy samples showed degenerated hepatocytes containing enlarged mitochondria with a distorted matrix. The matrix granules in the mitochondria were enlarged in size as compared with those of chronic hepatitis (Po = 0.0009). X-ray microanalysis has revealed the presence of calcium in large mitochondrial matrix granules. Both patients were thought to have valproic acid intoxication because they recovered quickly after the withdrawal of valproic acid.

Coupled sodium-calcium transport in cultured chick heart cells

In cultured embryonic chick heart cells, alterations of extracellular Na (Nao) and Ca (Cao), intracellular Na (Nai) and Ca, extracellular pH, and membrane potential resulted in changes in Na and Ca contents that were consistent with sarcolemmal Na-Ca exchange. 24Na efflux measurements revealed a large ouabain-insensitive component, one-third of which was inhibited by removal of Cao. Incubating the cells in Na-free solution resulted in a rapid, 1.5- to 2-fold increase in total cell Ca that remained elevated for at least 15 min. Cells exposed for 15 min to Nao less than or equal to 20 mM became maximally loaded with Ca, whereas Ca loading fell off sharply at values of Nao greater than 20 mM. The movement of Na against its electrochemical gradient was shown to be associated with Ca accumulation. During Na-K pump inhibition (in 10(-4) M ouabain), Na initially rose 2- to 3-fold to a level below its equilibrium value; then, lowering Cao for 30 min from 1.25 to 0.75 mM caused a 26% elevation in Nai, whereas raising Cao from 1.25 to 2.7 mM resulted in a 25% fall in Nai against its electrochemical gradient. These data are consistent with Nai being maintained by a Na-Ca exchange during Na-K pump inhibition. In the presence of ouabain (10(-4) M), Ca uptake into intracellular organelles, e.g., mitochondria, was suggested by an increase in total cell Ca as well as the occurrence of mitochondrial matrix granules, which were shown qualitatively by X-ray analysis to contain Ca. Although matrix granules also occurred in mitochondria during Na-free incubation, they did not contain detectable amounts of Ca when examined under identical conditions of fixation and analysis.

The Individual and Combined Effects of Citrinin and Ochratoxin A on Renal Ultrastructure in Layer Chicks

Layer chicks fed 3.0 micrograms of ochratoxin A (OA) and 300 micrograms of citrinin (CTN) per gram of feed, alone and in combination, were evaluated for changes in renal ultrastructure. Feeding OA from 0 to 21 days of age was associated with proximal tubular intranuclear membrane-bound inclusions, elongated tortuous and ring-shaped mitochondria, enlarged mitochondrial matrix granules with hyaline centers, and an increase in number and size of peroxisomes and secondary lysosomes. Birds fed OA from 0 to 7 days and then given untreated ration had similar changes but to a lesser degree. Feeding CTN from 0 to 21 days of age was associated with similar proximal tubular nuclear inclusions, elongated tortuous and ring-shaped mitochondria, and an increase in size and number of peroxisomes and secondary lysosomes. Hyalinized mitochondrial matrix granules were not present, and some proximal tubular cells had cytoplasmic aggregates of smooth endoplasmic reticulum. Birds fed CTN from 0 to 7 days had similar but milder changes. Birds fed CTN + OA for 21 days had changes similar to those fed OA alone and also had cytoplasmic aggregates of smooth endoplasmic reticulum similar to those of CTN-fed birds. Again, changes in birds fed CTN + OA for 7 days were similar but milder.

Morphometric analysis of hepatocytes from rats subjected to compound 48/80-induced anaphylactic shock

Morphometric and biochemical techniques were used to analyze hepatic glycogen, endoplasmic reticulum, and mitochondrial matrix granules in rats treated with compound 48/80 to induce an anaphylactic-like state of shock. Thirty minutes after insult there was a significant decrease in glycogen and mitochondrial matrix granules, an increase in rough endoplasmic reticulum (RER), and no change in smooth endoplasmic reticulum (SER). Less glycogen in experimental rats substantiated a previously described glycogenolytic response to compound 48/80. The decrease in matrix granules implies a loss and/or shift in intramitochondrial calcium as occurs in epinephrine-induced glycogenolysis in the rat. Since other glycogenolytic agents, e.g. glucagon, and starvation stimulate an increase in SER presumably from RER, the present morphological data suggest the increase in RER may precede proliferation of SER from RER.

Human gingival fibroblast cell lines in vitro. II. Electron microscopic studies of fibrogenesis.

Eight human gingival fibroblast cell lines were cultivated in glass cover slip chambers and analyzed by electron microscopy for their capacity to synthesize an extracellular fibrillar matrix. Both microfibrils and amorphous components of elastic tissue and several forms of banded and unbanded collagen fibrils were elaborated by the fibroblasts. Banding periods of 64 nm and 100 nm were observed. Collagenases lysed collagen fibrils and in some cases altered them to display prominently banded FLS images. Numerous matrix vesicles were observed among the extracellular collagen fibrils and some were observed intracellularly either isolated in pinocytotic‐like vesicles or as clusters in larger vacuoles. Matrix granules also were observed among the extracellular fibrils. The Golgi sacs of synthesizing fibroblasts contained fine thread‐like parallel filaments with a banding period of 220 nm. The presence of annulate lamellae was related to culture aging and the appearance of large mitochondrial matrix granules was related to the effect of small increments of vitamin C. Two cells were shown to have phagocytosed collagen elaborated by other cells earlier, one contained an interiorized fibril with a 64 nm banding period and the other an interiorized fibril with a 100 nm banding period. A turnover of collagen in vitro was thus established.

Mitochondrial matrix granules in soft tissues: II. Isolation and initial characterization of a calcium-precipitable, soluble lipoprotein subfraction from brown fat and liver mitochondria

Isolated mitochondria from lamb brown fat and rat liver were subject to hypotonic shock to free the matrix granules. Total mitochondrial membranes were collected by a two-stage high-speed centrifugation. By addition of calcium ions to the supernatant, membrane-like structures were formed and collected by further centrifugation. This calcium-precipitable fraction was a phospholipoprotein with very low or absent marker enzyme activities for inner and outer membranes but with the same relative amount of cardiolipin as the inner membrane. It contains fewer proteins in somewhat different proportions than the total membrane fraction. A single incorporation experiment indicated that the calcium-precipitable fraction might be enriched in products of mitochondrial protein synthesis. The available evidence suggests that matrix granule material forms the major part of the calcium-precipitable fraction and that, as previously hypothesized, matrix granules are involved in the assembly of the mitochondrial inner membrane.

Mitochondrial matrix granules in soft tissues: I. Elemental composition by X-ray microanalysis

The elemental composition of individual matrix granules in mitochondria of rat brown fat, mouse gall bladder and guinea pig kidney has been examined by X-ray microanalysis. The matrix granules showed a similar elemental composition that was strongly dependent upon the method of sample preparation. Low-temperature oxygen plasma microincineration or wavelength-dispersive X-ray spectrometers were used to demonstrate the presence of phosphorus in matrix granules of osmium tetroxide-fixed specimens. Matrix granule osmiophilia was retained in glutaraldehyde-fixed brown fat only if exposure to polar organic solvents was avoided during subsequent steps, e.g. by cryosectioning. As normally prepared, matrix granules lack detectable calcium but had bound this at detectable levels after fixation in osmium tetroxide, but not glutaraldehyde, supplemented with 5 mM Ca 2+. The results demonstrate that mitochondrial matrix granules of normal soft tissues are not calcium phosphate deposits and contain phospholipids, apparently as a major constituent. Thus they provide evidence against the hypothesis that matrix granules are primarily involved in mitochondrial calcium sequestration and, indirectly, for the hypothesis that the granules may be related to inner membrane assembly.

Ultrastructure Of Cell Organelles By Scanning Electron Microscopy Of Thick Sections Surface‐Etched By An Oxygen Plasma

Kidney tissue double fixed in glutaraldehyde and osmium tetroxide and embedded in epoxy resin by standard techniques used for transmission electron microscopy was cut into section 1 micron or more thick and surface-etched by an oxygen plasma. Etching caused ash residues (possibly composed partly or organo-metallic complexes) of membranes and other etch resistant cell components to emerge as recognizable structures projecting upward from the surrounding embedment which was combusted and removed as volatile products. using the secondary electron mode for image formation, structural features of cells which could be imaged with clarity with the scanning electron microscopy included: profiles of peripheral and in-folded plasma membranes, the nuclear envelope and profiles of cut mitochondrial matrix granules, cristae and the outer limiting membranes. Resolution was better than that obtainable from most other methods of specimen preparation currently being used in scanning electron microscopy for viewing the internal structures of cells or organelles in bulk samples of tissue.

Phenformin-induced alterations of small intestinal function and mitochondrial structure in man.

Abstract The mechanism by which phenformin diminishes blood glucose concentrations in man is not clear. We studied the effect of the drug on intestinal absorption because of several recent reports which have suggested that at least part of the drug's action may be the impairment of glucose absorption. Jejunal absorption rates of glucose, water, and sodium were measured in human volunteers with a segmental perfusion technique before and 75 minutes after administration of 100 mg. of phenformin. The drug reduced mean glucose absorption from 4,532 μmoles per hour per segment to 2,341, water absorption from 228 ml. per hour per segment to −25, and sodium absorption from 1,700 μEq per hour per segment to 400. Although mucosal biopsies appeared normal by light microscopy, electron microscopy revealed disappearance of mitochondrial matrix granules in villus absorptive cells after the drug. We suggest that the diminished rates of active intestinal transport is the consequence of mitochondrial injury with phenformin and an important mechanism of action of the drug in man.