Association of the Testicular Lactate Dehydrogenase Isozyme with a Special Type of Mitochondria1 (original) (raw)
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Biology of Reproduction, 1995
The proposed dual intracellular distribution of the sperm-specific lactate dehydrogenase (EC 1.1.1.27) isozyme C (LDH C 4) has been based on indirect evidence. In order to obtain direct evidence on the localization of this LDH isozyme in mice, postembedding immunocytochemistry at ultrastructural level was performed on testes, epididymal spermatozoa, and isolated testicular mitochondria. The immunogold technique was applied to thin sections incubated first in partially purified specific anti-LDH C, rabbit IgG, and immunoreactive sites were detected with colloidal gold adsorbed to anti-rabbit IgG. In the testis, immunostaining was found in the cytoplasm of spermatocytes and spermatids and in the principal and middle pieces of differentiating spermatozoa. Spermatozoa from epididymis also exhibited heavy labeling of colloidal gold in their middle and principal pieces, but the immunostaining was weak in the special type of mitochondria present in spermatocytes, spermatids, and spermatozoa (sperm-type mitochondria, STM). The isolation of STM produced several morphological changes in comparison with those in situ, including an enhancement of the LDH C 4 labeling in the mitochondrial matrix. The other type of mitochondria (non-STM) from spermatocytes and nonspermatogenic cells were not immunostained and served as background control. The results presented here confirm previous findings, gathered by indirect methods, indicating a dual localization of LDH C in the cytosol of spermatocytes, spermatids, and spermatozoa, as well as in the matrix of sperm-type mitochondria.
Biology of Reproduction, 2006
Sperm mitochondria undergo remodeling during posttesticular maturation that includes extensive disulfide cross-linking of proteins of the outer membrane to form the insoluble mitochondrial capsule. The relationship of these changes to mitochondrial function in mature gametes is unclear. The phospholipid hydroperoxide glutathione peroxidase (GPX4; also termed PHGPx) represents a major disulfide bond-stabilized protein of the mitochondrial capsule, and it is readily released by disulfide-reducing agents. However, in addition to GPX4, we detected a second major protein of 26 kDa (MP26) that was eluted from purified hamster sperm tails by the disulfidereducing agent dithiothreitol. The objectives of the present study were to identify and characterize MP26 and to explore its potential role in mitochondrial function. Proteomic analysis of MP26 by matrix-assisted laser desorption/ionization time-offlight (MALDI-TOF) identified 14 peptides with sequence identity to a member of the short-chain dehydrogenase/ reductase superfamily termed P26h, which was implicated previously in hamster sperm-zona binding, and with high sequence similarity to mouse lung carbonyl reductase. Indirect immunofluorescence localized MP26 to the midpiece, and twodimensional PAGE and immunoblot analysis identified a single MP26 isoform of pI 9.0. Immunoblot analyses of cauda epididymal fluid and of purified sperm plasma membranes and mitochondria revealed the exclusive localization of MP26 to the mitochondrial fraction. These data indicate that MP26 does not function in zona binding; instead, like GPX4, it may be associated with the mitochondrial capsule and play an important role in sperm mitochondrial function.
Molecular Human Reproduction, 2001
The 'common' 4977 bp deletion in mitochondrial DNA (∆4977) is commonly used as an indicator of tissue deterioration in ageing and bioenergetic diseases. Deletion levels are normally measured by a serial dilution polymerase chain reaction (PCR) approach, where test reactions are compared with dilutions of control amplifications of DNA from a similar sized stable region of the mitochondrial genome. The end-point of this assay is the dilution that can just detect any PCR product; however, this is an inherently unstable measure. We constructed a chimaeric DNA construct that binds to both control and deletion primers with similar annealing properties. This was used in a competitive PCR assay to quantify ∆4977 in human testicular tissues that had been well-characterized using the serial dilution approach. We found the competitive assay to be highly replicable as it compares the PCR product of the construct with that of test DNA samples during the linear growth phase of the PCR reaction. Moreover, the serial dilution assay was shown to significantly overestimate the amounts of deleted mitochondrial DNA present. The assay promises to throw new light on the role of mitochondrial DNA deletions in tissue dysfunction and ageing, as such deletions can now be determined with high accuracy and repeatability and is much cheaper to apply than real-time fluorescent quantitative PCR. D.Mehmet et al. Cummins, J.M. (2001a) Cytoplasmic inheritance and its implications for mutation. With such an assay now available, accurate deteranimal biotechnology. Theriogenology, in press. mination of any increase in this mutation with ageing is now Cummins, J.M. (2001b) Mitochondria: potential roles in embryogenesis and possible. We concentrated here on a series of testicular tissues
Mitochondria, spermatogenesis and male infertility
Mitochondrion, 2010
Mitochondria and its role in male reproduction has remained an enigma since long. Similarly, etiology of male infertility in a large percentage of individuals, mainly primary infertility, has evaded concrete conclusions. Oxidative metabolism, energy production and free radical generation are the principal biological reactions occurring inside mitochondria. In addition to the above, mitochondria participates in an important process of apoptosis. Mitochondrial causes of infertility have triggered interest because of its presence in the tail of sperm and immense need of energy for sperm motility. Several studies on mitochondria have strongly suggested its role in fertility, some of which support mitochondrial role presenting numerous hypotheses, whereas others deny its very existence as a causative factor. We have, in the present review of existing literature, covered the role of mitochondria right from spermatogenesis to male infertility.
George Yu Article 2 Mitochondrial and Testosterone
Among the major physiological functions of steroid hormones is regulation of carbohydrate, fat, and protein metabolism. Mitochondria, through oxidative phosphorylation, play a critical role in modulating a host of complex cellular metabolic pathways to produce chemical energy to meet the metabolic demand for cellular function. Thus, androgens may regulate cellular metabolism and energy production by increased mitochondrial numbers, activation of respiratory chain components, and increased transcription of mitochondrial-encoded respiratory chain genes that code for enzymes responsible for oxidative phosphorylation. Androgen defi ciency is associated with increased insulin resistance, type 2 diabetes (T2DM), metabolic syndrome, obesity, and increased overall mortality. One common link among all these pathologies is mitochondrial dysfunction. Contemporary evidence exists suggesting that testosterone defi ciency (TD) contributes to mitochondrial dysfunction, including structural alterations and reduced expression and activities of metabolic enzymes. Here, we postulate that TD contributes to symptoms of fatigue, insulin resistance, T2DM, cardiovascular risk, and metabolic syndrome through a common mechanism involving impairment of mitochondrial function.
Genetic Association in the Maintenance of the Mitochondrial Microenvironment and Sperm Capacity
Oxidative Medicine and Cellular Longevity, 2021
Sperm motility is one of the major determinants of male fertility. Since sperm need a great deal of energy to support their fast movement by active metabolism, they are thus extremely vulnerable to oxidative damage by the reactive oxygen species (ROS) and other free radicals generated as byproducts in the electron transport chain. The present study is aimed at understanding the impact of a mitochondrial oxidizing/reducing microenvironment in the etiopathology of male infertility. We detected the mitochondrial DNA (mtDNA) 4,977 bp deletion in human sperm. We examined the gene mutation of ATP synthase 6 (ATPase6 m.T8993G) in ATP generation, the gene polymorphisms of uncoupling protein 2 (UCP2, G-866A) in the uncoupling of oxidative phosphorylation, the role of genes such as manganese superoxide dismutase (MnSOD, C47T) and catalase (CAT, C-262T) in the scavenging system in neutralizing reactive oxygen species, and the role of human 8-oxoguanine DNA glycosylase (hOGG1, C1245G) in 8-hydr...
Mitochondria functionality and sperm quality
Reproduction, 2013
Although mitochondria are best known for being the eukaryotic cell powerhouses, these organelles participate in various cellular functions besides ATP production, such as calcium homoeostasis, generation of reactive oxygen species (ROS), the intrinsic apoptotic pathway and steroid hormone biosynthesis. The aim of this review was to discuss the putative roles of mitochondria in mammalian sperm function and how they may relate to sperm quality and fertilisation ability, particularly in humans. Although paternal mitochondria are degraded inside the zygote, sperm mitochondrial functionality seems to be critical for fertilisation. Indeed, changes in mitochondrial integrity/functionality, namely defects in mitochondrial ultrastructure or in the mitochondrial genome, transcriptome or proteome, as well as low mitochondrial membrane potential or altered oxygen consumption, have been correlated with loss of sperm function (particularly with decreased motility). Results from genetically engine...
Correlation of sperm motility with mitochondrial enzymatic activities
Clinical Chemistry, 1998
Until now, little attention has been paid to the contribution of mitochondrial dysfunction to germinal tissue disorders. The target of this study was to investigate the relationship between sperm motility and mitochondrial respiratory chain enzyme activities. The results obtained showed that semen samples of control individuals (n = 33) have substantially higher activities of complexes I, II, and IV compared with those of asthenozoospermic subjects (n = 86). Moreover, a direct and positive correlation was found in the whole population studied between spermatozoa motility and all the mitochondrial respiratory complex activities assayed (I, II, I+III, II+III, and IV). The ratio of these enzymes to citrate synthase (a reliable enzymatic marker of mitochondrial volume) activities did not correlate with sperm motility. This suggests that motility depends largely on the mitochondrial volume within the sperm midpiece. These observations could be of physiopathological relevance because they...
PLOS One, 2011
Human sperm samples are very heterogeneous and include a low amount of truly functional gametes. Distinct strategies have been developed to characterize and isolate this specific subpopulation. In this study we have used fluorescence microscopy and fluorescence-activated cell sorting to determine if mitochondrial function, as assessed using mitochondrialsensitive probes, could be employed as a criterion to obtain more functional sperm from a given ejaculate. We first determined that mitochondrial activity correlated with the quality of distinct human samples, from healthy donors to patients with decreased semen quality. Furthermore, using fluorescence-activated cell sorting to separate sperm with active and inactive mitochondria we found that this was also true within samples. Indeed, sperm with active mitochondria defined a more functional subpopulation, which contained more capacitated and acrosome intact cells, sperm with lower chromatin damage, and, crucially, sperm more able to decondense and participate in early development using both chemical induction and injection into mature bovine oocytes. Furthermore, cell sorting using mitochondrial activity produced a more functional sperm subpopulation than classic swim-up, both in terms of improvement in a variety of functional sperm parameters and in statistical significance. In conclusion, whatever the true biological role of sperm mitochondria in fertilization, mitochondrial activity is a clear hallmark of human sperm functionality.