Sjøgren's syndrome-associated oxidative stress and mitochondrial dysfunction: Prospects for chemoprevention trials

Abstract

An involvement of oxidative stress (OS) was found in recent studies of Sjøgren's syndrome (SS) that reported significant changes in protein oxidation, myeloperoxidase activity, TNF-α, nitrotyrosine, and GSH levels in plasma from SS patients. Excess levels of OS markers, as oxidative DNA damage and propanoyl-lysine, were reported in saliva from SS patients. Previous reports concurred with a role of OS in SS pathogenesis, by showing a decreased expression of antioxidant activities in conjunctival epithelial cells of SS patients and in parotid gland tissue samples from SS patients. A link between OS and mitochondrial dysfunction (MDF) is recognized both on the grounds of the established role of mitochondria in reactive oxygen species (ROS) formation and by the occurrence of MDF in a set of OS-related disorders. Earlier studies detected mitochondrial alterations in cells from SS patients, related to the action of antimitochondrial autoantibodies, and affecting specific mitochondrial activities. Thus, a link between MDF and OS may be postulated in SS, prompting studies aimed at elucidating SS pathogenesis and in the prospect of chemoprevention trials in SS clinical management.     

Oxidative stress in Fanconi anaemia: from cells and molecules towards prospects in clinical management

Fanconi anaemia (FA) is a genetic disease featuring bone marrow failure, proneness to malignancies, and chromosomal instability. A line of studies has related FA to oxidative stress (OS). This review attempts to evaluate the evidence for FA-associated redox abnormalities in the literature from 1981 to 2010. Among 2170 journal articles on FA evaluated, 162 related FA with OS. Early studies reported excess oxygen toxicity in FA cells that accumulated oxidative DNA damage. Prooxidant states were found in white blood cells and body fluids from FA patients as excess luminol-dependent chemiluminescence, 8-hydroxy-deoxyguanosine, reduced glutathione/oxidized glutathione imbalance, and tumour necrosis factor-α. Some FA gene products involved in redox homeostasis can be summarized as follows: (a) FANCA, FANCC, and FANCG interact with cytochrome P450-related activities and/or respond to oxidative damage; (b) FANCD2 in OS response interacts with forkhead box O3 and ataxia telangiectasia mutated protein; (c) FANCG is found in mitochondria and interacts with PRDX3, and FA-G cells display distorted mitochondria and decreased peroxidase activity; (d) FANCJ (BACH1/BRIP1) is a repressor of haeme oxygenase-1 gene and senses oxidative base damage; (e) antioxidants, such as tempol and resveratrol decrease cancer incidence and haematopoietic defects in Fancd2(-/-) mice. The overall evidence for FA-associated OS may suggest designing chemoprevention studies aimed at delaying the onset of OS-related clinical complications.     

N-3 HUFAs affect fat deposition,susceptibility to oxidative stress,and apoptosis in Atlantic salmon visceral adipose tissue

We have investigated how n-3 highly unsaturated fatty acids (HUFAs) in the diet affect fatty acid (FA) utilization, fat storage and oxidative stress (OS) in Atlantic salmon (Salmo salar) white adipose tissue (WAT). Four groups of Atlantic salmon were fed for 21 weeks on one of the four diets supplemented with 23% (of dry matter) lipid. Docosahexaenoic acid (DHA; 22:6n-3) and eicosapentaenoic acid (EPA; 20:5n-3) levels increased from 10% of total FAs in the rapeseed oil (RO) diet, to 20% in the fish oil (FO) diet, and to 50% and 55% in the DHA-enriched and EPA-enriched diets, respectively. Increased dietary levels of n-3 HUFAs resulted in lower fat percentage in WAT. Furthermore, mitochondrial FA β-oxidation activity was higher in the FO group than it was in the RO group. The relative levels of DHA and EPA in phospholipids (PLs) from WAT and mitochondrial membranes increased with the increasing dietary levels of these HUFAs. In general, the mitochondrial membrane PLs were characterised by lower relative levels of n-3 HUFAs and higher relative levels of linoleic acid (LA; 18:2 n-6) than WAT membrane PLs. The predominance of LA relative to n-3 HUFAs in mitochondrial membrane PLs may help to protect these PLs from peroxidation. Cytochrome c oxidase measurements revealed higher incidence of disrupted mitochondrial membranes in the DHA and EPA dietary groups than in the FO and RO dietary groups. This disruption further affected the mitochondrial function, resulting in a marked reduction in FA β-oxidation capacities. The reduction in mitochondrial function and the increase in the activity of superoxide dismutase (SOD) in the DHA and EPA groups showed that high dietary dose of DHA and EPA resulted in oxidative stress (OS). The increased activity of caspase 3 in the high n-3 HUFA groups suggested the induction of apoptosis and increased incidence of cell death in WAT, which may be one of the factors explaining the lower fat percentage found in these groups.     

Association of low GLP-1 with oxidative stress is related to cardiac disease and outcome in patients with type 2 diabetes mellitus: A pilot study

Oxidative stress (OS) contributes to cardiovascular damage in type 2 diabetes mellitus (T2DM). The peptide glucagon-like peptide-1 (GLP-1) inhibits OS and exerts cardiovascular protective actions. Our aim was to investigate whether cardiac remodeling (CR) and cardiovascular events (CVE) are associated with circulating GLP-1 and biomarkers of OS in T2DM patients. We also studied GLP-1 antioxidant effects in a model of cardiomyocyte lipotoxicity. We examined 72 T2DM patients with no coronary or valve heart disease and 14 nondiabetic subjects. A median of 6 years follow-up information was obtained in 60 patients. Circulating GLP-1, dipeptidyl peptidase-4 activity, and biomarkers of OS were quantified. In T2DM patients, circulating GLP-1 decreased and OS biomarkers increased, compared with nondiabetics. Plasma GLP-1 was inversely correlated with serum 3-nitrotyrosine in T2DM patients. Patients showing high circulating 3-nitrotyrosine and low GLP-1 levels exhibited CR and higher risk for CVE, compared to the remaining patients. In palmitate-stimulated HL-1 cardiomyocytes, GLP-1 reduced cytosolic and mitochondrial oxidative stress, increased mitochondrial ATP synthase expression, partially restored mitochondrial membrane permeability and cytochrome c oxidase activity, blunted leakage of creatine to the extracellular medium, and inhibited oxidative damage in total and mitochondrial DNA. These results suggest that T2DM patients with reduced circulating GLP-1 and exacerbated OS may exhibit CR and be at higher risk for CVE. In addition, GLP-1 exerts antioxidant effects in HL-1 palmitate-overloaded cardiomyocytes. It is proposed that therapies aimed to increase GLP-1 may counteract OS, protect from CR, and prevent CVE in patients with T2DM.     

New insights into redox response modulation in Fanconi's anemia cells by hydrogen peroxide and glutathione depletors

Fanconi's anemia (FA) patients face severe pathological consequences. Bone marrow failure, the major cause of death in FA, accounting for as much as 80-90% of FA mortality, appears to be significantly linked to excessive apoptosis of hematopoietic cells induced by oxidative stress. However, 20-25% of FA patients develop malignancies of myeloid origin. A survival strategy for bone marrow and hematopoietic cells under selective pressure evidently exists. This study reports that lymphoblastoid cell lines derived from two FA patients displayed significant resistance to oxidative stress induced by treatments with H(2) O(2) and various glutathione (GSH) inhibitors that induce production of reactive oxygen species, GSH depletion and mitochondrial membrane depolarization. Among the various GSH inhibitors employed, FA cells appear particularly resistant to menadione (5 μm) and ethacrynic acid (ETA, 50 μm), two drugs that specifically target mitochondria. Even after pre-treatment with buthionine sulfoximine, a GSH synthesis inhibitor that induces enhanced induction of reactive oxygen species, FA cells maintain significant resistance to these drugs. These data suggest that the resistance to oxidative stress and the altered mitochondrial and metabolic functionality found in the FA mutant cells used in this study may indicate the survival strategy that is adopted in FA cells undergoing transformation. The study of redox and mitochondria regulation in FA may be of assistance in diagnosis of the disease and in the care of patients.     

Comparative studies of oxidative stress and mitochondrial function in aging

The oxidative stress theory and its correlate the mitochondrial theory of aging are among the most studied and widely accepted of all hypotheses of the mechanism of aging. To date, most of the supporting evidence for these theories has come from investigations using common model organisms such as Caenorhabditis elegans, Drosophila melanogaster, and laboratory rodents. However, comparative data from a wide range of endotherms provide equivocal support as to whether oxidative stress is merely a correlate, rather than a determinant, of species' maximum lifespan. The great majority of studies in this area have been devoted to the relationship between reactive oxygen species and maximal longevity in young adult organisms, with little emphasis on mitochondrial respiratory efficiency, age-related alterations in mitochondrial physiology or oxidative damage. The advantage of studying a broader spectrum of species is the broad range of virtually every biological phenotype/trait, such as lifespan, body weight and metabolic rate. Here we summarize the results from a number of comparative studies in an effort to correlate oxidant production and oxidative damage among many species with their maximal lifespan and briefly discuss the pitfalls and limitations. Based on current information, it is not possible to accept or dispute the oxidative stress theory of aging, nor can we exclude the possibility that private mechanisms might offer an explanation for the longevity of exceptionally long-lived animal models. Thus, there is need for more thorough and controlled investigations with more unconventional animal models for a deeper understanding of the role of oxidative stress in longevity.     

Defective mitochondrial peroxiredoxin-3 results in sensitivity to oxidative stress in Fanconi anemia

Cells from patients with Fanconi anemia (FA), an inherited disorder that includes bone marrow failure and cancer predisposition, have increased sensitivity to oxidative stress through an unknown mechanism. We demonstrate that the FA group G (FANCG) protein is found in mitochondria. Wild-type but not G546R mutant FANCG physically interacts with the mitochondrial peroxidase peroxiredoxin-3 (PRDX3). PRDX3 is deregulated in FA cells, including cleavage by a calpainlike cysteine protease and mislocalization. FA-G cells demonstrate distorted mitochondrial structures, and mitochondrial extracts have a sevenfold decrease in thioredoxin-dependent peroxidase activity. Transient overexpression of PRDX3 suppresses the sensitivity of FA-G cells to H2O2, and decreased PRDX3 expression increases sensitivity to mitomycin C. Cells from the FA-A and -C subtypes also have PRDX3 cleavage and decreased peroxidase activity. This study demonstrates a role for the FA proteins in mitochondria witsh sensitivity to oxidative stress resulting from diminished peroxidase activity. These defects may lead to apoptosis and the accumulation of oxidative DNA damage in bone marrow precursors.     

Oxidative stress induces mitochondrial fragmentation in frataxin-deficient cells

Friedreich ataxia (FA) is the most common recessive neurodegenerative disease. It is caused by deficiency in mitochondrial frataxin, which participates in iron-sulfur cluster assembly. Yeast cells lacking frataxin (Δyfh1 mutant) showed an increased proportion of fragmented mitochondria compared to wild-type. In addition, oxidative stress induced complete fragmentation of mitochondria in Δyfh1 cells. Genetically controlled inhibition of mitochondrial fission in these cells led to increased resistance to oxidative stress. Here we present evidence that in yeast frataxin-deficiency interferes with mitochondrial dynamics, which might therefore be relevant for the pathophysiology of FA.     

Increasing Fatty Acid Oxidation Remodels the Hypothalamic Neurometabolome to Mitigate Stress and Inflammation

Modification of hypothalamic fatty acid (FA) metabolism can improve energy homeostasis and prevent hyperphagia and excessive weight gain in diet-induced obesity (DIO) from a diet high in saturated fatty acids. We have shown previously that C75, a stimulator of carnitine palmitoyl transferase-1 (CPT-1) and fatty acid oxidation (FAOx), exerts at least some of its hypophagic effects via neuronal mechanisms in the hypothalamus. In the present work, we characterized the effects of C75 and another anorexigenic compound, the glycerol-3-phosphate acyltransferase (GPAT) inhibitor FSG67, on FA metabolism, metabolomics profiles, and metabolic stress responses in cultured hypothalamic neurons and hypothalamic neuronal cell lines during lipid excess with palmitate. Both compounds enhanced palmitate oxidation, increased ATP, and inactivated AMP-activated protein kinase (AMPK) in hypothalamic neurons in vitro. Lipidomics and untargeted metabolomics revealed that enhanced catabolism of FA decreased palmitate availability and prevented the production of fatty acylglycerols, ceramides, and cholesterol esters, lipids that are associated with lipotoxicity-provoked metabolic stress. This improved metabolic signature was accompanied by increased levels of reactive oxygen species (ROS), and yet favorable changes in oxidative stress, overt ER stress, and inflammation. We propose that enhancing FAOx in hypothalamic neurons exposed to excess lipids promotes metabolic remodeling that reduces local inflammatory and cell stress responses. This shift would restore mitochondrial function such that increased FAOx can produce hypothalamic neuronal ATP and lead to decreased food intake and body weight to improve systemic metabolism.     

Mitochondrial involvement in Parkinson's disease, Huntington's disease, hereditary spastic paraplegia and Friedreich's ataxia

Respiratory chain dysfunction has been identified in several neurodegenerative disorders. In Friedreich's ataxia (FA) and Huntington's disease (HD), where the respective mutations are in nuclear genes encoding non-respiratory chain mitochondrial proteins, the defects in oxidative phosphorylation are clearly secondary. In Parkinson's disease (PD) the situation is less clear, with some evidence for a primary role of mitochondrial DNA in at least a proportion of patients. The pattern of the respiratory chain defect may provide some clue to its cause; in PD there appears to be a selective complex I deficiency; in HD and FA the deficiencies are most severe in complex II/III with a less severe defect in complex IV. Aconitase activity in HD and FA is severely decreased in brain and muscle, respectively, but appears to be normal in PD brain. Free radical generation is thought to be of importance in both HD and FA, via excitotoxicity in HD and abnormal iron handling in FA. The oxidative damage observed in PD may be secondary to the mitochondrial defect. Whatever the cause(s) and sequence of events, respiratory chain deficiencies appear to play an important role in the pathogenesis of neurodegeneration. The mitochondrial abnormalities induced may converge on the function of the mitochondrion in apoptosis. This mode of cell death is thought to play an important role in neurodegenerative diseases and it is tempting to speculate that the observed mitochondrial defects in PD, HD and FA result directly in apoptotic cell death, or in the lowering of a cell's threshold to undergo apoptosis. Clarifying the role of mitochondria in pathogenesis may provide opportunities for the development of treatments designed to reverse or prevent neurodegeneration.     

Sensitivity to radiation of human normal, hyperthyroid, and neoplastic thyroid epithelial cells in primary culture

Samples of thyroid tissue removed surgically from 63 patients were cultured in vitro and exposed to X irradiation to investigate the radiosensitivities of various types of thyroid epithelial cells. A total of 76 samples were obtained, including neoplastic cells from patients with papillary carcinoma (PC) or follicular adenoma (FA), cells from hyperthyroidism (HY) patients, and normal cells from the surgical margins of PC and FA patients. Culturing of the cells was performed in a manner which has been shown to yield a predominance of epithelial cells. Results of colony formation assays indicated that cells from HY and FA patients were the least radiosensitive: when adjusted to the overall geometric mean plating efficiency of 5.5%, the average mean lethal dose Do was 97.6 cGy for HY cells and 96.7 and 94.3 cGy, respectively for neoplastic and normal cells from FA patients. Cells from PC patients were more radiosensitive, normal cells having an adjusted average Do of 85.0 cGy and PC cells a significantly (P = 0.05) lower average Do of 74.4 cGy. After allowing for this variation by cell type, in vitro radiosensitivity was not significantly related to age at surgery (P = 0.82) or sex (P = 0.10). These results suggest that malignant thyroid cells may be especially radiosensitive.     

Fluctuating asymmetry, size and mating success in males of Ischnura elegans (Vander Linden) (Odonata: Coenagrionidae)

Fluctuating asymmetry (FA) is thought to be an indicator of developmental stability and negatively related to male mating success in many animal taxa. We investigated the relationships between mating success of males, body size and FA for both wing length and number of setae on the legs in the damselfly Ischnura elegans. Males were classified as mated or unmated at the time of sampling. Fluctuating asymmetry, expressed as right-left differences, showed normal distributions without evidence of directional asymmetry or antisymmetry. Univariate analyses showed a significant negative correlation between size and mating success, and significant negative correlations between FA and mating success for both characters. On the other hand, with a multivariate analysis, new to studies on FA, the effect of body size was still significant but FA did not reach significance for either character. We conclude that the multivariate analysis should be used to assess the role of the different factors affecting mating success. Copyright 2000 The Association for the Study of Animal Behaviour.     

Chronic pancreatitis: role of oxidative stress and antioxidants

Abstract

Chronic pancreatitis (CP) is characterized by pain, and exocrine and endocrine insufficiency of pancreas. Several hypotheses have been put forward to explain the hitherto partially understood pathophysiology of CP. In the past decade, animal and clinical studies have suggested that an increased chronic oxidative stress (OS) plays a key role in pathophysiology of CP and perpetuates its clinical and histological symptoms (pain and fibrosis–necrosis, respectively). Mounting OS in pancreatic acinar cells is a result of overproduction of free radicals (FR) during xenobiotic metabolism. It has been shown that Phase I cytochrome P450 enzymes of xenobiotic pathway are induced when exposed to a xenobiotic overload including alcohol, tobacco, smoke and other dietary toxins, which exceeds the capacity of Phase II conjugation due to limited glutathione availability. Consequently, there is an overload of toxic metabolites as well as FR. Additionally, bioactivation of subsequently entering compounds may occur increasing their toxicity. Such an imbalance overwhelms the antioxidant capacity of the body resulting in undefended chronic OS that derails the normal physiology of pancreatic acinar cells since FR act as second messengers controlling the cellular signaling. OS hypothesis is further supported by the studies that demonstrated that antioxidant supplementation ameliorated pain. Moreover, animal studies have demonstrated a cessation of fibrotic cascade with antioxidant supplementation. In a recent large randomized controlled trial, it was demonstrated that antioxidant supplementation led to a significant reduction in pain, and also lowered the OS in patients with alcoholic or idiopathic CP.     

Identification and sizing of GAA trinucleotide repeat expansion, investigation for D-loop variations and mitochondrial deletions in Iranian patients with Friedreich's ataxia

Friedreich's Ataxia (FA) is the commonest genetic cause of ataxia and is associated with the expansion of a GAA repeat in intron 1 of the frataxin gene. Iron accumulation in the mitochondria of patients with FA would result in hypersensitivity to oxidative stress. Mitochondrial DNA (mtDNA) could be considered a candidate modifier factor for FA disease, since mitochondrial oxidative stress is thought to be involved in the pathogenesis of this disease. We studied 25 Iranian patients (16 females and 9 males) from 12 unrelated families. DNA from each patient was extracted and frequency and length of (GAA)(n) repeat was analyzed using a long-range polymerase chain reaction (PCR) test. Also we investigated impact of GAA size on neurological findings, age of onset and disease development. In order to identify polymorphic sites and genetic background, the sequence of two hypervariable regions (HVR-I and HVR-II) of mtDNA was obtained from FA patients harbouring GAA trinucletide expansions. Alignment was made with the revised cambridge reference sequence (rCRS) and any differences recorded as single base substitution (SBS), insertions and deletions. Homozygous GAA expansion was found in 21 (84%) of all cases. In four cases (16%), no expansion was observed, ruling out the diagnosis of Friedreich's ataxia. In cases with GAA expansions, ataxia, scoliosis and pes cavus, cardiac abnormalities and some neurological findings occurred more frequently than in our patients without GAA expansion. Molecular analysis was imperative for diagnosis of Friedreich's ataxia, not only for typical cases, but also for atypical ones. Diagnosis bases only on clinical findings is limited, however, it aids in better screening for suspected cases that should be tested. Our results showed that the rate of D-loop variations was higher in FA patients than control (P<0.05). mtDNA deletions were present in 76% of our patients representing mtDNA damage, which may be due to iron accumulation in mitochondria.     

FANCL supports Parkin-mediated mitophagy in a ubiquitin ligase-independent manner

Fanconi anemia (FA) is the most common inherited bone marrow failure syndrome. The FA proteins have functions in genome maintenance and in the cytoplasmic process of selective autophagy, beyond their canonical roles of repairing DNA interstrand cross-links. FA core complex proteins FANCC, FANCF, FANCL, FANCA, FANCD2, BRCA1 and BRCA2, which previously had no known direct functions outside the nucleus, have recently been implicated in mitophagy. Although mutations in FANCL account for only a very small number of cases in FA families, it plays a key role in the FA pathophysiology and might drive carcinogenesis. Here, we demonstrate that FANCL protein is present in mitochondria in the control and Oligomycin and Antimycin (OA)-treated cells and its ubiquitin ligase activity is not required for its localization to mitochondria. CRISPR/Cas9-mediated knockout of FANCL in HeLa cells overexpressing parkin results in increased sensitivity to mitochondrial stress and defective clearing of damaged mitochondria upon OA treatment. This defect was reversed by the reintroduction of either wild-type FANCL or FANCL(C307A), a mutant lacking ubiquitin ligase activity. To summarize, FANCL protects from mitochondrial stress and supports Parkin-mediated mitophagy in a ubiquitin ligase-independent manner.     

Caffeic acid phenethyl ester and its related compounds limit the functional alterations of the isolated mouse brain and liver mitochondria submitted to in vitro anoxia-reoxygenation: relationship to their antioxidant activities

It is an important therapeutic strategy to protect mitochondria from oxidative stress, especially during ischemia-reperfusion. In the present study, an attempt has been made to evaluate the protective effects of caffeic acid phenethyl ester (CAPE) and its related phenolic compounds on mouse brain and liver mitochondria injury induced by in vitro anoxia-reoxygenation. Added before anoxia or reoxygenation, CAPE markedly protected coupled respiration with the decrease in state 4 and the increases in state 3, respiratory control ratio (RCR) and ADP/O ratio in a concentration-dependent manner. CAPE effectively protected mitochondria by inhibiting the mitochondrial membranes fluidity decrease, the lipoperoxidation and the protein carbonylation increase, which indicated its protective action against the mitochondrial oxidative damage. Meanwhile, CAPE blocked the enhanced release of cardiolipin (CL) and cytochrome c (Cyt c). The related phenolic compounds like caffeic acid (CA), ferulic acid (FA) and ethyl ferulate (EF) also had different-degree protective effects. CAPE and CA were more potent than FA and EF. Their structural differences played the key role in their activity levels. These results suggest that CAPE and its related phenolic compounds protect mitochondria mainly correlated to their antioxidative activities and may be of interest for the prevention and therapy of ischemia-reperfusion injuries.     

Role of Fanconi DNA repair pathway in neural stem cell homeostasis

Defects in DNA repair pathways have been involved in collapse of early neurogenesis leading to brain development abnormalities and embryonic lethality. However, consequences of DNA repair defects in adult neural stem and progenitor cells and their potential contribution in ageing phenotype are poorly understood. The Fanconi anaemia (FA) pathway, which functions primarily as a DNA damage response system, has been examined in neural stem and progenitor cells during developmental and adult neurogenesis. We have shown that loss of fanca and fancg specifically provokes neural progenitor apoptosis during forebrain development, related to DNA repair defects, which persists in adulthood leading to depletion of the neural stem cell pool with ageing. In addition, neural stem cells from FA mice had a reduced capacity to self-renew in vitro. Here, we expand upon our recent work and give further data examining possible implication of oxidative stress. Therefore, FA phenotype might be interpreted as a premature ageing of stem cells, DNA damages being among the driving forces of ageing.     

Oxidative stress, endogenous antioxidants, alcohol, and hepatitis C: pathogenic interactions and therapeutic considerations

Hepatitis C virus (HCV) is a blood-borne pathogen that was identified as an etiologic agent of non-A, non-B hepatitis in 1989. HCV is estimated to have infected at least 170 million people worldwide. The majority of patients infected with HCV do not clear the virus and become chronically infected, and chronic HCV infection increases the risk for hepatic steatosis, cirrhosis, and hepatocellular carcinoma. HCV induces oxidative/nitrosative stress from multiple sources, including inducible nitric oxide synthase, the mitochondrial electron transport chain, hepatocyte NAD(P)H oxidases, and inflammation, while decreasing glutathione. The cumulative oxidative burden is likely to promote both hepatic and extrahepatic conditions precipitated by HCV through a combination of local and more distal effects of reactive species, and clinical, animal, and in vitro studies strongly point to a role of oxidative/nitrosative stress in HCV-induced pathogenesis. Oxidative stress and hepatopathogenesis induced by HCV are exacerbated by even low doses of alcohol. Alcohol and reactive species may have other effects on hepatitis C patients such as modulation of the host immune system, viral replication, and positive selection of HCV sequence variants that contribute to antiviral resistance. This review summarizes the current understanding of redox interactions of HCV, outlining key experimental findings, directions for future research, and potential applications to therapy.     

Divergent modulation of swine ileal microbiota by formic acid and methionine hydroxy analogue-free acid

Management of intestinal microbiota of monogastric animals has increased in importance since the ban of growth promoting antibiotics in many countries. Organic acids have been used as alternatives to antibiotics by many feed manufacturers. Regardless of the wide usage, the effect, dose response and mode of action of acids on intestinal microbes is poorly understood. In this study, we investigated the effects of dietary supplementation of three commonly used products, namely formic acid (FA) (90%), dl-methionine (DLM) (99%) and liquid methionine hydroxy analogue-free acid (88%), on ileal microbiota of pigs. Laboratory simulation system, mimicking swine ileum, was used to study the products at various concentrations and combinations. Furthermore, selected combinations were tested in a piglet trial to confirm the findings made in in vitro studies. FA turned out to have a dual effect on ileal microbiota. At concentrations below 0.5%, it significantly stimulated bacteria, but at higher inclusion rates it was highly inhibitory. This finding, which was consistent in in vitro and in vivo studies, implies that reducing the dose of FA does not lead to a diluted inhibitory effect, but in fact, an opposite, stimulatory effect on intestinal microbiota. It is highly important that feed compounders acknowledge this finding. Unlike FA, the inhibitory effect of methionine hydroxy analogue on ileal bacteria was linearly dose dependent and significant at inclusion levels above 0.2%, in vitro. Partial replacement of methionine hydroxy analogue by FA, or FA by methionine hydroxy analogue, led to an unpredictable outcome due to the dual effects of FA; e.g., a minor inclusion of added FA changed the inhibitory effect of methionine hydroxy analogue into microbial stimulation by FA. Inhibition of ileal microbiota by methionine hydroxy analogue was detected only in in vitro studies, suggesting that intact methionine hydroxy analogue may not have reached the ileum, in live animals. Therefore, if the target is to ensure the inhibitory effect of FA, the FA level in feed should be kept above 0.6%, and not reduced, if methionine hydroxy analogue is used as a methionine source instead of DLM. DLM was totally inert with regard to bacterial growth and metabolism, both in vitro and in vivo. The results of these studies reveal the importance of knowing how each acid product works. Inconsistent results in animal trials may have been partly due to quadratic dose-response effects of products, and unpredictable product combination effects.     

Mechanism, measurement, and prevention of oxidative stress in male reproductive physiology

Numerous factors influence male fertility. Among these factors is oxidative stress (OS), which has elicited an enormous interest in researchers in recent period. Reactive oxygen species (ROS) are continuously produced by various metabolic and physiologic processes. OS occurs when the delicate balance between the production of ROS and the inherent antioxidant capacity of the organism is distorted. Spermatozoa are particularly sensitive to ROS as their plasma membrane contains polyunsaturated fatty acids (PUFA), which oxidizes easily. They also lack cytoplasm to generate a robust preventive and repair mechanism against ROS. The transition metal ions that are found in the body have a catalytic effect in the generation of ROS. Lifestyle behaviours such as smoking and alcohol use and environmental pollution further enhance the generation of ROS and thus, cause destructive effects on various cellular organelles like mitochondria, sperm DNA etc. This article analyzes the detrimental effects of OS on male fertility, measurement of OS and effective ways to decrease or eliminate them completely. We have also provided information on oxidative stress in other systems of the body, which may be applied to future research in the field of reproductive biology.