Mitochondrial respiratory supercomplex association limits production of reactive oxygen species from Complex I

Evidence accumulated in the last ten years has demonstrated that a large proportion of the mitochondrial respiratory chain complexes in a variety of organisms is arranged in supramolecular assemblies called supercomplexes or respirasomes. Besides conferring a kinetic advantage (substrate channeling) and being required for the assembly and stability of Complex I, indirect considerations support the view that supercomplexes may also prevent excessive formation of reactive oxygen species (ROS) from the respiratory chain. Following this line of thought we have decided to directly investigate ROS production by Complex I under conditions in which the complex is arranged as a component of the supercomplex I1III2 or it is dissociated as an individual enzyme. The study has been addressed both in bovine heart mitochondrial membranes and in reconstituted proteoliposomes composed of complexes I and III in which the supramolecular organization of the respiratory assemblies is impaired by: (i) treatment either of bovine heart mitochondria or liposome-reconstituted supercomplex I-III with dodecyl maltoside; (ii) reconstitution of Complexes I and III at high phospholipids to protein ratio. The results of this investigation provide experimental evidence that the production of ROS is strongly increased in either model; supporting the view that disruption or prevention of the association between Complex I and Complex III by different means enhances the generation of superoxide from Complex I . This is the first demonstration that dissociation of the supercomplex I1III2 in the mitochondrial membrane is a cause of oxidative stress from Complex I. Previous work in our laboratory demonstrated that lipid peroxidation can dissociate the supramolecular assemblies; thus, here we confirm that preliminary conclusion that primary causes of oxidative stress may perpetuate reactive oxygen species (ROS) generation by a vicious circle involving supercomplex dissociation as a major determinant.

Role of Reactive Oxygen Species in signalling and oxidative stress

Results reported in this Thesis contribute to the comprehension of the complicated world of “redox biology”. ROS regulate signalling pathways both in physiological responses and in pathogenesis and progression of diseases. In cancer cells, the increase in ROS generation from metabolic abnormalities and oncogenic signalling may trigger a redox adaptation response, leading to an up-regulation of antioxidant capacity in order to maintain the ROS level below the toxic threshold. Thus, cancer cells would be more dependent on the antioxidant system and more vulnerable to further oxidative stress induced by exogenous ROS-generating agents or compounds that inhibit the antioxidant system. Results here reported indicate that the development of new drugs targeting specific Nox isoforms, responsible for intracellular ROS generation, or AQP isoforms, involved in the transport of extracellular H2O2 toward intracellular targets, might be an interesting novel anti-leukaemia strategy. Furthermore, also the use of CSD peptide, which simulate the VEGFR-2 segregation into caveolae in the inactive form, might be a strategy to stop the cellular response to VEGF signalling. As above stated, in the understanding of the redox biology, it is also important to identify and distinguish the molecular effectors that maintain normal biological and physiological responses, such as agents that stimulate our adaptation systems and elevate our endogenous antioxidant defences or other protective systems. Data here reported indicate that the nutraceutical compound sulforaphane and the Klotho protein are able to stimulate the HO-1 and Prx-1 expression, as well as the GSH levels, confirming their antioxidant and protective role. Finally, results here reported demonstrated that Stevia extracts are involved in insulin regulated glucose metabolism, suggesting that the use of these compounds goes beyond their sweetening power and may also offer therapeutic benefits hence improving the quality of life.

Reactive oxygen species mediate functional differences in human radial and internal thoracic arteries from smokers

Smoking not only increases the risk that coronary heart disease will develop but also morbidity and mortality in patients with known coronary atherosclerosis and after coronary artery bypass grafting. Excessive generation of reactive oxygen species (ROS) has been implicated as the final common pathway for the development of endothelial dysfunction in various cardiovascular risk factors. This study assessed the influence of smoking on two different human arteries routinely used as coronary artery bypass graft conduits.

Dystrophic cardiomyopathy: amplification of cellular damage by Ca2+ signalling and reactive oxygen species-generating pathways

AIMS: Cardiac myopathies are the second leading cause of death in patients with Duchenne and Becker muscular dystrophy, the two most common and severe forms of a disabling striated muscle disease. Although the genetic defect has been identified as mutations of the dystrophin gene, very little is known about the molecular and cellular events leading to progressive cardiac muscle damage. Dystrophin is a protein linking the cytoskeleton to a complex of transmembrane proteins that interact with the extracellular matrix. The fragility of the cell membrane resulting from the lack of dystrophin is thought to cause an excessive susceptibility to mechanical stress. Here, we examined cellular mechanisms linking the initial membrane damage to the dysfunction of dystrophic heart. METHODS AND RESULTS: Cardiac ventricular myocytes were enzymatically isolated from 5- to 9-month-old dystrophic mdx and wild-type (WT) mice. Cells were exposed to mechanical stress, applied as osmotic shock. Stress-induced cytosolic and mitochondrial Ca(2+) signals, production of reactive oxygen species (ROS), and mitochondrial membrane potential were monitored with confocal microscopy and fluorescent indicators. Pharmacological tools were used to scavenge ROS and to identify their possible sources. Osmotic shock triggered excessive cytosolic Ca(2+) signals, often lasting for several minutes, in 82% of mdx cells. In contrast, only 47% of the WT cardiomyocytes responded with transient and moderate intracellular Ca(2+) signals. On average, the reaction was 6-fold larger in mdx cells. Removal of extracellular Ca(2+) abolished these responses, implicating Ca(2+) influx as a trigger for abnormal Ca(2+) signalling. Our further experiments revealed that osmotic stress in mdx cells produced an increase in ROS production and mitochondrial Ca(2+) overload. The latter was followed by collapse of the mitochondrial membrane potential, an early sign of cell death. CONCLUSION: Overall, our findings reveal that excessive intracellular Ca(2+) signals and ROS generation link the initial sarcolemmal injury to mitochondrial dysfunctions. The latter possibly contribute to the loss of functional cardiac myocytes and heart failure in dystrophy. Understanding the sequence of events of dystrophic cell damage and the deleterious amplification systems involved, including several positive feed-back loops, may allow for a rational development of novel therapeutic strategies.

Nitric oxide induces cell death in canine cruciate ligament cells by activation of tyrosine kinase and reactive oxygen species

BACKGROUND: There is increasing evidence suggesting that development of progressive canine cranial cruciate ligament (CCL) rupture involves a gradual degeneration of the CCL itself, initiated by a combination of factors, ranging from mechanical to biochemical. To date, knowledge is lacking to what extent cruciate disease results from abnormal biomechanics on a normal ligament or contrary how far preliminary alterations of the ligament due to biochemical factors provoke abnormal biomechanics. This study is focused on nitric oxide (NO), one of the potential biochemical factors. The NO-donor sodium nitroprusside (SNP) has been used to study NO-dependent cell death in canine cranial and caudal cruciate ligament cells and to characterize signaling mechanisms during NO-stimulation. RESULTS: Sodium nitroprusside increased apoptotic cell death dose- and time-dependently in cruciate ligamentocytes. Cells from the CCL were more susceptible to apoptosis than CaCL cells. Caspase-3 processing in response to SNP was not detected. Testing major upstream and signal transducing pathways, NO-induced cruciate ligament cell death seemed to be mediated on different levels. Specific inhibition of tyrosine kinase significantly decreased SNP-induced cell death. Mitogen activated protein kinase ERK1 and 2 are activated upon NO and provide anti-apoptotic signals whereas p38 kinase and protein kinase C are not involved. Moreover, data showed that the inhibition reactive oxygen species (ROS) significantly reduced the level of cruciate ligament cell death. CONCLUSIONS: Our data support the hypothesis that canine cruciate ligamentocytes, independently from their origin (CCL or CaCL) follow crucial signaling pathways involved in NO-induced cell death. However, the difference on susceptibility upon NO-mediated apoptosis seems to be dependent on other pathways than on these tested in the present study. In both, CCL and CaCL, the activation of the tyrosine kinase and the generation of ROS reveal important signaling pathways. In perspective, new efforts to prevent the development and progression of cruciate disease may include strategies aimed at reducing ROS.

Reactive oxygen species-dependent mechanisms of N-(4-hydroxyphenyl)retinamide (4HPR)-induced apoptosis in human carcinoma cells

4HPR is a synthetic retinoid that has shown chemopreventive and therapeutic efficacy against premalignant and malignant lesions including oral leukoplakia, ovarian and breast cancer, and neuroblastoma. 4HPR induces apoptosis in various cancer cells and production of reactive oxygen species (ROS) has been suggested as a possible cause underlying these effects. However, the mechanisms governing these effects by 4HPR are not fully elucidated. In this study, we explored the mechanisms of 4HPR-induced ROS increase and apoptosis in human cancer cells. ^ First, we identified genes modulated by 4HPR using oligonucleotide gene expression arrays and found that they fall into specific functional canonical pathways and gene networks using Ingenuity Pathways Analysis®. Further analysis has shown that 4HPR induced up-regulation of Endoplasmic Reticulum (ER)-related genes such as Heat shock proteins 70 and 90 and the transcriptional factor, GADD153. These findings were validated using quantitative real-time PCR. ^ Second, we found that 4HPR induced extensive ER stress evidenced by dilation of the ER and endoribonuclease-mediated splicing and activation of the transcriptional factor, XBP-1. In addition, 4HPR induced the up-regulation of various ER stress-related genes and their protein products, as well as cleavage and activation of the ER specific Caspase-4. Concomitantly with XBP-1 splicing, all of these effects were dependent on ROS generation by 4HPR. Furthermore, chemical inhibition and RNA interference studies revealed a novel pro-apoptotic role for HSP70/A1A in 4HPR-mediated apoptosis. ^ Third, we observed rapid activation of the small GTPase Rac by 4HPR which was upstream of ROS generation. Inhibition of Rac activity or silencing of its expression by RNA interference inhibited ROS generation and apoptosis induction by 4HPR. siRNA targeting PAK1 and expression of a dominant negative Rac, decreased 4HPR-mediated ROS generation, while expression of a constitutive active Rac increased basal and 4HPR-induced ROS generation and PARP cleavage. Furthermore, metastatic cancer cells exhibited higher Rac activation, ROS generation, and cell growth inhibition due to 4HPR exposure compared to their primary cancer cell counterparts. ^ These findings provide novel insights into 4HPR-mediated ROS generation and apoptosis induction and support the use of ROS inducing agents such as 4HPR against metastatic cancer cells. ^

GMZ27 is a new organic arsenic derivative with anti-leukemic activity, via the induction of reactive oxygen species through NADPH oxidase and mitochondrial pathway that lead to leukemia cell apoptosis

Arsenic trioxide (ATO) is an inorganic arsenic derivative that is very effective against relapsed acute promyelocytic leukemia. It is being investigated as therapy for other cancers, but the risk/benefit ratio is questionable due to significant side effects. In contrast, organic arsenic derivatives (OAD) are known to be much less toxic than ATO. Based on high activity, we selected GMZ27 (dipropil-s-glycerol arsenic) for further study and have confirmed its potent activity against human acute leukemia cell lines. This anti-leukemic activity is significantly higher than that of ATO. Both in vivo and in vitro tests have shown that GMZ27 is significantly less toxic to normal bone marrow mononuclear cells and normal mice. Therefore, further study of the biological activity of GMZ27 was undertaken. ^ GMZ27, in contrast to ATO, can only marginally induce maturation of leukemic cells. GMZ27 has no effect on cell cycle. The anti-leukemic activity of GMZ27 against acute myeolocytic leukemia cells is not dependent upon degradation of PML-RARα fusion protein. GMZ27 causes dissipation of mitochondrial transmembrane potential, cleavage of caspase 9, caspase 3 activation. Further studies indicated that GMZ27 induces intracellular reactive oxygen species (ROS) production, and modification of intracellular ROS levels had profound effect on its potential to inhibit proliferation of leukemic cells. Therefore ROS production plays a major role in the anti-leukemic activity of GMZ27. ^ To identify how GMZ27 induces ROS, our studies focused on mitochondria and NADPH oxidase. The results indicated that the source of ROS generation induced by GMZ27 is dose dependent. At the low dose (0.3 uM) GMZ27 induces NADPH oxidase activity that leads to late ROS production, while at the high dose (2.0 uM) mitochondria function is disrupted and early ROS production is induced leading to dramatic cell apoptosis. Therefore, late, ROS production can be detected in mitochondria are depleted Rho-0 cells. Our work not only delineates a major biologic pathway for the anti-leukemic activity of GMZ27, but also discusses possible ways of enhancing the effect by the co-application of NADPH oxidase activator. Further study of this interaction may lead to achieving better therapeutic index.^

LOW REACTIVE OXYGEN SPECIES AND HIGH GLYCOLYSIS IN GLIOBLASTOMA STEM CELLS:MECHANISMS AND THERAPEUTIC IMPLICATIONS

Glioblastoma multiforme (GBM) is the most common and aggressive primary brain tumor with poor prognosis due in part to drug resistance and high incidence of tumor recurrence. The drug resistant and cancer recurrence phenotype may be ascribed to the presence of glioblastoma stem cells (GSCs), which seem to reside in special stem-cell niches in vivo and require special culture conditions including certain growth factors and serum-free medium to maintain their stemness in vitro. Exposure of GSCs to fetal bovine serum (FBS) can cause their differentiation, the underlying mechanism of which remains unknown. Reactive oxygen species (ROS) play an important role in normal stem cell differentiation, but their role in affecting cancer stem cell fate remains unclear. Whether the metabolic characteristics of GSCs are different from other glioblastoma cells and can be targeted are also unknown. In this study, we used several stem-like glioblastoma cell lines derived from clinical tissues by typical neurosphere culture system or orthotopic xenografts, and showed that addition of fetal bovine serum to the medium induced an increase of ROS, leading to aberrant differentiation and decreases of stem cell markers such as CD133. We found that exposure of GSCs to serum induced their differentiation through activation of mitochondrial respiration, leading to an increase in superoxide (O2-) generation and a profound ROS stress response manifested by upregulation of oxidative stress response pathway. This increase in mitochondrial ROS led to a down-regulation of molecules including SOX2, and Olig2, and Notch1 that are important for stem cell function and an upregulation of mitochondrial superoxide dismutase SOD2 that converts O2- to H2O2. Neutralization of ROS by antioxidant N-acetyl-cysteine in the serum-treated GSCs suppressed the increase of superoxide and partially rescued the expression of SOX2, Olig2, and Notch1, and prevented the serum-induced differentiation phenotype. Additionally, GSCs showed high dependence on glycolysis for energy production. The combination of a glycolytic inhibitor 3-BrOP and a chemotherapeutic agent BCNU depleted cellular ATP and inhibited the repair of BCNU-induced DNA damage, achieving strikingly synergistic killing effects in drug resistant GSCs. This study uncovers the metabolic properties of glioblastoma stem cells and suggests that mitochondrial function and cellular redox status may profoundly affect the fates of glioblastoma stem cells via a ROS-mediated mechanism, and that the active glycolytic metabolism in cancer stem cells may provide a biochemical basis for developing novel therapeutic strategies to effectively eliminate GSCs.

The anti-inflammatory actions of methotrexate are critically dependent upon the production of reactive oxygen species

1. The mechanism of action by which methotrexate (MTX) exerts its anti-inflammatory and immunosuppressive effects remains unclear. The aim of this study is to investigate the hypothesis that MTX exerts these effects via the production of reactive oxygen species (ROS). 2. Addition of MTX (100 nM-10 μM) to U937 monocytes induced a time and dose dependent increase in cytosolic peroxide [peroxide] cyt from 6-16 h. MTX also caused corresponding monocyte growth arrest, which was inhibited (P<0.05) by pre-treatment with N-acetylcysteine (NAC; 10 mM) or glutathione (GSH; 10 mM). In contrast, MTX induction of [peroxide] cyt in Jurkat T cells was more rapid (4 h; P<0.05), but was associated with significant apoptosis at 16 h at all doses tested (P<0.05) and was significantly inhibited by NAC or GSH (P<0.05). 3. MTX treatment of monocytes (10 nM-10 μM) for 16 h significantly reduced total GSH levels (P<0.05) independently of dose (P>0.05). However in T-cells, GSH levels were significantly elevated following 30 nM MTX treatment (P<0.05) but reduced by doses exceeding 1 μM compared to controls (P<0.05). 4. MTX treatment significantly reduced monocyte adhesion to 5 h and 24 h LPS (1 μg ml -1) activated human umbilical vein endothelial cells (HUVEC; P<0.05) but not to resting HUVEC. Pre-treatment with GSH prevented MTX-induced reduction in adhesion. 5. In conclusion, ROS generation by MTX is important for cytostasis in monocytes and cytotoxicity T-cells. Furthermore, MTX caused a reduction in monocyte adhesion to endothelial cells, where the mechanism of MTX action requires the production of ROS. Therefore its clinical efficacy can be attributed to multiple targets.

Ceramide and reactive oxygen species (ROS) as signal transduction molecules in inflammation

Reactive oxygen species (ROS) and the sphingolipid ceramide are each partly responsible for the intracellular signal transduction of a variety of physiological, pharmacological or environmental agents. Furthermore, the enhanced production of many of these agents, that utilise ROS and ceramide as signalling intermediates, is associated with the aetiologies of several vascular diseases (e.g. atherosclerosis) or disorders of inflammatory origin (e.g. rheumatoid arthritis; RA). Excessive monocyte recruitment and uncontrolled T cell activation are both strongly implicated in the chronic inflammatory responses that are associated with these pathologies. Therefore the aims of this thesis are (1) to further elucidate the cellular responses to modulations in intracellular ceramide/ROS levels in monocytes and T cells, in order to help resolve the mechanisms of progression of these diseases and (2) to examine both existing agents (methotrexate) and novel targets for possible therapeutic manipulation. Utilising synthetic, short chain ceramide to mimic the cellular responses to fluctuations in natural endogenous ceramide or, stimulation of CD95 to induce ceramide formation, it is described here that ceramide targets and manipulates two discrete sites responsible for ROS generation, preceding the cellular responses of growth arrest in U937 monocytes and apoptosis in Jurkat T-cells. In both cell types, transient elevations in mitochondrial ROS generation were observed. However, the prominent redox altering effects appear to be the ceramide-mediated reduction in cytosolic peroxide, the magnitude of which dictates in part the cellular response in U937 monocytes, Jurkat T-cells and primary human peripheral blood resting or PHA-activated T-cells in vitro. The application of synthetic ceramides to U937 monocytes for short (2 hours) or long (16 hours) treatment periods reduced the membrane expression of proteins associated with cell-cell interaction. Furthermore, ceramide treated U937 monocytes demonstrated reduced adhesion to 5 or 24 hour LPS activated human umbilical vein endothelial cells (HUVEC) but not resting HUVEC. Consequently it is hypothesised that the targeted treatment of monocytes from patients with cardiovascular diseases with short chain synthetic ceramide may reduce disease progression. Herein, the anti-inflammatory and immunosuppressant drug, methotrexate, is described to require ROS production for the induction of cytostasis or cytotoxicity in U937 monocytes and Jurkat T-cells respectively. Further, ROS are critical for methotrexate to abrogate monocyte interaction with activated HUVEC in vitro. The histological feature of RA of enhanced infiltration, survivability and hyporesponsiveness of T-cells within the diseased synovium has been suggested to arise from aberrant signalling. No difference in the concentrations of endogenous T-cell ceramide, the related lipid diacylglycerol (DAG) and cytosolic peroxide ex vivo was observed. TCR activation following PHA exposure in vitro for 72 hours did not induced maintained perturbations in DAG or ceramide in T-cells from RA patients or healthy individuals. However, T-cells from RA patients failed to upregulate cytosolic peroxide in response to PHA, unlike those from normals, despite expressing identical levels of the activation marker CD25. This inability to upregulate cytosolic peroxide may contribute to the T-cell pathology associated with RA by affecting the signalling capacity of redox sensitive biomolecules. These data highlight the importance of two distinctive cellular pools of ROS in mediating complex biological events associated with inflammatory disease and suggest that modulation of cellular ceramides represents a novel therapeutic strategy to minimise monocyte recruitment.

Reactive Oxygen Species via Redox Signaling to PI3K/AKT Pathway Contribute to the Malignant Growth of 4-Hydroxy Estradiol-Transformed Mammary Epithelial Cells

The purpose of this study was to investigate the effects of 17-β-estradiol (E2)-induced reactive oxygen species (ROS) on the induction of mammary tumorigenesis. We found that ROS-induced by repeated exposures to 4-hydroxy-estradiol (4-OH-E2), a predominant catechol metabolite of E2, caused transformation of normal human mammary epithelial MCF-10A cells with malignant growth in nude mice. This was evident from inhibition of estrogen-induced breast tumor formation in the xenograft model by both overexpression of catalase as well as by co-treatment with Ebselen. To understand how 4-OH-E2 induces this malignant phenotype through ROS, we investigated the effects of 4-OH-E2 on redox-sensitive signal transduction pathways. During the malignant transformation process we observed that 4-OH-E2 treatment increased AKT phosphorylation through PI3K activation. The PI3K-mediated phosphorylation of AKT in 4-OH-E2-treated cells was inhibited by ROS modifiers as well as by silencing of AKT expression. RNA interference of AKT markedly inhibited 4-OH-E2-induced in vitro tumor formation. The expression of cell cycle genes, cdc2, PRC1 and PCNA and one of transcription factors that control the expression of these genes – nuclear respiratory factor-1 (NRF-1) was significantly up-regulated during the 4-OH-E2-mediated malignant transformation process. The increased expression of these genes was inhibited by ROS modifiers as well as by silencing of AKT expression. These results indicate that 4-OH-E2-induced cell transformation may be mediated, in part, through redox-sensitive AKT signal transduction pathways by up-regulating the expression of cell cycle genes cdc2, PRC1 and PCNA, and the transcription factor – NRF-1. In summary, our study has demonstrated that: (i) 4-OH-E2 is one of the main estrogen metabolites that induce mammary tumorigenesis and (ii) ROS-mediated signaling leading to the activation of PI3K/AKT pathway plays an important role in the generation of 4-OH-E2-induced malignant phenotype of breast epithelial cells. In conclusion, ROS are important signaling molecules in the development of estrogen-induced malignant breast lesions.

The application of profluorescent nitroxides to detect reactive oxygen species derived from combustion-generated particulate matter: Cigarette smoke – a case study

Reactive oxygen species (ROS) and related free radicals are considered to be key factors underpinning the various adverse health effects associated with exposure to ambient particulate matter. Therefore, measurement of ROS is a crucial factor for assessing the potential toxicity of particles. In this work, a novel profluorescent nitroxide, BPEAnit, was investigated as a probe for detecting particle-derived ROS. BPEAnit has a very low fluorescence emission due to inherent quenching by the nitroxide group, but upon radical trapping or redox activity, a strong fluorescence is observed. BPEAnit was tested for detection of ROS present in mainstream and sidestream cigarette smoke. In the case of mainstream cigarette smoke, there was a linear increase in fluorescence intensity with an increasing number of cigarette puffs, equivalent to an average of 101 nmol ROS per cigarette based on the number of moles of the probe reacted. Sidestream cigarette smoke sampled from an environmental chamber exposed BPEAnit to much lower concentrations of particles, but still resulted in a clearly detectible increase in fluorescence intensity with sampling time. It was calculated that the amount of ROS was equivalent to 50 ± 2 nmol per mg of particulate matter; however, this value decreased with ageing of the particles in the chamber. Overall, BPEAnit was shown to provide a sensitive response related to the oxidative capacity of the particulate matter. These findings present a good basis for employing the new BPEAnit probe for the investigation of particle-related ROS generated from cigarette smoke as well as from other combustion sources.

Reactive Oxygen Species and Reactive Nitrogen Species in Epigenetic Modifications

For the normal homeostasis of a cell, there must be a balance between radical oxygen species/radical nitrogen species (ROS/RNS) production and the neutralization of these species by antioxidant scavenging. In times of stress, this balance is not maintained, and the result is oxidative stress. This stress can affect many pathways in the body and result in pathological consequences. Recent evidence suggests that ROS/RNS can affect the epigenetic regulation of genes by affecting the function of histone and DNA modifying enzymes, thus affecting phenotypic changes within the cellular environment. In the following chapter, we provide a broad overview of how oxidative stress induced by ROS/RNS can affect epigenetics, and using lung disease as our model we link the connection between these processes.

Effect of atmospheric aging on volatility and reactive oxygen species of biodiesel exhaust nano-particles

In the prospect of limited energy resources and climate change, effects of alternative biofuels on primary emissions are being extensively studied. Our two recent studies have shown that biodiesel fuel composition has a significant impact on primary particulate matter emissions. It was also shown that particulate matter caused by biodiesels was substantially different from the emissions due to petroleum diesel. Emissions appeared to have higher oxidative potential with the increase in oxygen content and decrease of carbon chain length and unsaturation levels of fuel molecules. Overall, both studies concluded that chemical composition of biodiesel is more important than its physical properties in controlling exhaust particle emissions. This suggests that the atmospheric aging processes, including secondary organic aerosol formation, of emissions from different fuels will be different as well. In this study, measurements were conducted on a modern common-rail diesel engine. To get more information on realistic properties of tested biodiesel particulate matter once they are released into the atmosphere, particulate matter was exposed to atmospheric oxidants, ozone and ultra-violet light; and the change in their properties was monitored for different biodiesel blends. Upon the exposure to oxidative agents, the chemical composition of the exhaust changes. It triggers the cascade of photochemical reactions resulting in the partitioning of semi-volatile compounds between the gas and particulate phase. In most of the cases, aging lead to the increase in volatility and oxidative potential, and the increment of change was mainly dependent on the chemical composition of fuels as the leading cause for the amount and the type of semi-volatile compounds present in the exhaust.