Abnormal Reproductive Measures and Seminal Plasma Findings in Men With Thalassemia Major (TM) and Iron Overload

Blood ◽  
2013 ◽  
Vol 122 (21) ◽  
pp. 4707-4707
Author(s):  
Sylvia Titi Singer ◽  
Jerry Wang ◽  
Jung H Suh ◽  
David Killelea ◽  
Kristen Ivani ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Dna Damage ◽  
Trace Elements ◽  
Iron Overload ◽  
Seminal Plasma ◽  
Seminal Fluid ◽  
Sperm Quality ◽  
Sperm Count ◽  
Iron Concentration ◽  
Fertility Potential

Continuous improvement in the life span of TM patients allows patients to address critical issues of specific organ function. Among these hypogonadotrophic-hypogonadism (HH) and infertility are common complications, having a significant adverse impact on quality of life (QOL). While pregnancies are reported in TM women, paternity is less common in TM men, with significant gaps in our understanding of the underlying pathogenesis of male reduced fertility. Growing evidence indicates that oxidative stress negatively affects sperm quality and function, playing a central role in certain types of male infertility. In addition, abnormal low trace elements, in particular zinc, are important causes for infertility. We hypothesized that deleterious effects of iron overload impair sperm quality beyond that of iron deposition in the pituitary and the resultant HH state, and thus aimed to evaluate potential mechanisms for infertility in TM men: Systemic and pituitary iron burden (liver iron concentration; LIC, pituitary iron by MRI, ferritin), reproductive hormones (LH, FSH) and semen fluid were measured in 7 TM men (median age 26, range 21-30 years). Semen fluid parameters included volume, sperm count, motility, DNA damage determined by DNA Fragmentation Index (DFI), concentration of glutathione (GSH), a major cellular antioxidant which plays a central role in the defense against oxidative stress, and that of pertinent metal/trace elements (iron, zinc, copper). Results were compared to normal reference and Pearson correlation analysis was performed to identify markers that best predict fertility potential and highlight pathogenesis.Sperm and semen plasma analysisLab resultsImagingPtVol. mlCount Mill/mlMotility %DFI %Zn mg/LFe mg/LGSH µmol/LLH/FSH mIU/mLFerritin ng/mLLICMRI R213.51787513141.41.189.54.9/7.96685.413.5± 0.222.500-54.42.732.20.3/0.9369625.216.4± 0.731.400-6.72.530.880.0/0.0428822.816.8± 1.442.314659.7163.02.218.661.6/2.710926.614.5± 0.951.51137---7.52.8/3.419248.4-64.594758.6120.407.54-37752016.7± 0.671.800-80.22.341.060.07/0.330002019.5± 1.2nl>2>20>50<151400.3±0.12030-60/ 0.9-15<300<39.5-13 Semen analysis showed that 5/7 (70%) had either low sperm count or complete azoospermia, and 4/7 exhibited reduced sperm motility (57%). The three patients with a measurable sperm count did not exhibit increased DNA fragmentation (normal DFI). Overall, patients had low seminal plasma GSH concentration, indicating increased oxidative stress. Seminal plasma had elevated iron concentration in 5/6 patients; zinc levels were low in 4/6, while copper was undetectable in all patients (nl 0.064 mg/L). Patients with highest pituitary iron (2,3,7) had the lowest LH/FSH levels and azoospermia, but pituitary iron (R2) did not correlate with LH/FSH, with sperm or systemic iron measures. Seminal fluid GSH (p=0.001), iron (p=0.06), and zinc (p=0.02) inversely correlated with sperm motility. Systemic iron status, as measured by LIC and ferritin, was inversely correlated with GSH (p=0.02; 0.04, respectively) and with zinc (p=0.04; 0.04, respectively). Our data suggest that increased free iron, iron-induced oxidative damage and low trace elements are associated with impaired semen integrity and may explain low fertility potential in TM men, beyond the expected effect of pituitary iron toxicity and HH. These abnormalities, however, did not result in sperm DNA damage, in a small analyzed sample size. Oxidative and metal profiles in seminal fluid may be a valuable tool to improve the reproductive evaluation of TM men. Anti-oxidant treatment will likely have an important role in increasing their reproductive potential. Disclosures: No relevant conflicts of interest to declare.

Amelioration of sperm count and sperm quality by lycopene supplementation in irradiated mice

2020 ◽  
Vol 32 (12) ◽  
pp. 1040
Author(s):  
Małgorzata M. Dobrzyńska ◽  
Aneta Gajowik
Keyword(s):  
Dna Damage ◽  
Sperm Quality ◽  
Sperm Count ◽  
Combined Treatment ◽  
Treatment Groups ◽  
Patients With Cancer ◽  
Equivalent Time ◽  
Radiation Induced ◽  
Almost All ◽  
Harmful Effects

Male mice were exposed to lycopene (LYC; 0.15 and 0.30mg kg−1) and irradiation (0.5, 1 Gy) alone or in combination (0.5 Gy+0.15mg kg−1 LYC; 0.5 Gy+0.30mg kg−1 LYC; 1 Gy+0.15mg kg−1 LYC; 1 Gy+0.30mg kg−1 LYC) for 2 weeks. LYC administration in the drinking water was started 24h or on Day 8 after the first irradiation dose or equivalent time point for groups treated with LYC alone. Sperm count, motility, morphology and DNA damage were determined at the end of the 2-week treatment period. Irradiation deteriorated sperm count and quality. Supplementation with LYC from 24h significantly increased the sperm count compared with irradiation alone. In almost all combined treatment groups, the percentage of abnormal spermatozoa was significantly decreased compared with that after irradiation alone. In some cases, combined treatment reduced levels of DNA damage in gametes. Both doses of LYC administered from Day 8 significantly reduced the percentage of morphologically abnormal spermatozoa compared with that seen after 1 Gy irradiation and reduced DNA damage in all combined treatment groups. In conclusion, LYC supplementation after irradiation can ameliorate the harmful effects of irradiation on gametes. Mitigation of radiation-induced damage in germ cells following LYC administration may be useful for radiological accidents and to protect non-treated tissues in patients with cancer undergoing radiotherapy.

scholarly journals Effects of coadministration of DHA and vitamin E on spermatogram, seminal oxidative stress, and sperm phospholipids in asthenozoospermic men: a randomized controlled trial

2020 ◽  
Vol 112 (3) ◽  
pp. 707-719 ◽  
Author(s):  
Ghazaleh Eslamian ◽  
Naser Amirjannati ◽  
Nazanin Noori ◽  
Mohammad-Reza Sadeghi ◽  
Azita Hekmatdoost
Keyword(s):  
Oxidative Stress ◽  
Vitamin E ◽  
Sperm Motility ◽  
Seminal Plasma ◽  
Sperm Count ◽  
Controlled Trial ◽  
Sperm Concentration ◽  
Endocrine Function ◽  
Semen Parameters ◽  
Double Blind

ABSTRACT Background It is unknown which compounds in spermatozoa or seminal plasma may be involved in the regulation of sperm motility. Objectives The aim of this study was to investigate the effects of DHA (22:6n–3), vitamin E, and their probable interactions in men with asthenozoospermia. Methods A factorial, randomized, double-blind, placebo-controlled trial was conducted in infertility clinics in Tehran, Iran. The participants were idiopathic asthenozoospermic men aged 20–45 y, with normal endocrine function. Their concentration of spermatozoa and percentage of morphologically normal spermatozoa were equal to or above the lower reference limits, according to the fifth edition of the WHO guideline. Out of 717 men referred to the infertility clinics, 180 asthenozoospermic men were randomly assigned to 1 of 4 groups according to stratified blocked randomization by age and sperm concentration. Participants took daily 465 mg DHA plus 600 IU vitamin E (DE), 465 mg DHA plus placebo (DP), 600 IU vitamin E plus placebo (EP), or both placebo capsules (PP) for 12 wk. Sperm characteristics, oxidative stress of seminal plasma, serum and sperm membrane fatty acids, dietary intakes, anthropometric measurements, and physical activity were measured at baseline and after 12 wk. Results After the intervention, mean ± SD sperm progressive motility was greater in the DE group (27.9 ± 2.8) than in the DP (25.7 ± 3.4), EP (26.1 ± 2.8), and PP (25.8 ± 2.6) groups (P &lt; 0.05). Sperm count (P = 0.001) and concentration (P = 0.044) increased significantly in the DE group compared with the other 3 groups, whereas other semen parameters were not significantly different between the groups after the intervention. Serum concentrations of n–3 PUFAs were significantly higher in the DE and DP groups than in the EP and PP groups. Conclusions Combined DHA and vitamin E supplements led to increased sperm motility; however, no significant changes occurred in sperm morphology and vitality in asthenozoospermic men. This trial was registered at clinicaltrials.gov as NCT01846325.

Steroids in semen, their role in spermatogenesis, and the possible impact of endocrine disruptors

2013 ◽  
Vol 13 (1) ◽  
pp. 1-5 ◽  
Author(s):  
Richard Hampl ◽  
Jana Kubátová ◽  
Vladimír Sobotka ◽  
Jiří Heráček
Keyword(s):  
Blood Plasma ◽  
Endocrine Disruptors ◽  
Seminal Plasma ◽  
Hormone Receptors ◽  
Seminal Fluid ◽  
Semen Quality ◽  
Sperm Count ◽  
Environmental Chemicals ◽  
Male Reproduction ◽  
Steroid Hormone Receptors

AbstractThe data on hormonal steroids in the human seminal plasma and their role in spermatogenesis are summarized. The seminal steroid levels need not correlate with the blood plasma levels. The recent reports showed that androgen, especially dihydrotestosterone, and the estrogen levels in the seminal fluid may be used as the markers of spermatogenesis impairment. The estradiol concentration in the seminal plasma was higher than in the blood plasma, and its levels were significantly increased in men with impaired spermatogenesis. A good indicator for predicting the normal spermatogenesis, therefore, seems to be the testosterone/estradiol ratio. The seminal plasma also contains significant amounts of cortisol, which influences the androgen biosynthesis through its receptors in the Leydig cells. The local balance between cortisol and inactive cortisone is regulated by 11β-hydroxysteroid dehydrogenase, the activity of which may be affected by the environmental chemicals acting as the endocrine disruptors (EDCs). These compounds are believed to participate in worsening the semen quality – the sperm count, motility, and morphology, as witnessed in the recent last decades. As to the steroids’ role in the testis, the EDCs may act as antiandrogens by inhibiting the enzymes of testosterone biosynthesis, as the agonists or antagonists through their interaction with the steroid hormone receptors, or at the hypothalamic-pituitary-gonadal axis. Surprisingly, though the EDCs affect the steroid action in the testis, there is no report of a direct association between the concentrations of steroids and the EDCs in the seminal fluid. Therefore, measuring the steroids in the semen, along with the various EDCs, could help us better understand the role of the EDCs in the male reproduction.

scholarly journals Oxytocin in pig seminal plasma is positively related with in vivo fertility of inseminated sows

2021 ◽  
Vol 12 (1) ◽  
Author(s):  
Lorena Padilla ◽  
Marina López-Arjona ◽  
Silvia Martinez-Subiela ◽  
Heriberto Rodriguez-Martinez ◽  
Jordi Roca ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Sperm Quality ◽  
Sperm Count ◽  
Sperm Concentration ◽  
Reproductive Function ◽  
Peptide Hormone ◽  
Large White ◽  
Male Reproductive Function ◽  
Farrowing Rate

Abstract Background Identification of relevant in vivo biomarkers for fertility remains a challenge for the livestock industry. Concentrations of the small peptide hormone oxytocin (OXT), involved in male reproductive function and present in the seminal plasma (SP) of several species could be a robust one. This study characterized concentrations of SP-OXT in ejaculates from boars used in artificial insemination (AI) programs aiming to evaluate its relationship with sperm quality variables and in vivo fertility of their liquid-stored AI-semen. Seminal OXT concentrations (ng/mL) were measured in 169 ejaculates from 61 boars of the Duroc, Pietrain, Landrace and Large White breeds using a direct competitive immunoassay test based on AlphaLISA® technology. Ejaculate (ejaculate volume, sperm concentration, total sperm count) and sperm parameters (motility, viability, intracellular generation of reactive oxygen species, plasma membrane fluidity) were assessed at 0 h and 72 h in AI-semen samples stored at 17 °C. In vivo fertility included only 18 Large White and Landrace boars whose AI-semen was used to inseminated > 100 sows and evaluated both farrowing rate and litter size of 3,167 sows. Results The results showed that SP-OXT differed between boars and between ejaculates within boar (P < 0.05) but not between breeds (Duroc, Pietrain, Landrace and Large White). Ejaculates with higher SP-OXT concentration/mL (hierarchically grouped; P < 0.001) had larger volume and came from younger boars (P < 0.05). Ejaculates of boars showing positive farrowing rate deviation exhibited higher (P < 0.05) SP-OXT concentration/mL than those with negative farrowing rate deviation. Conclusion The SP concentrations of OXT are boar, ejaculate and age dependent, and positively related with ejaculate volume and farrowing rates of liquid-stored semen AI-doses.

scholarly journals Pyrethroids Toxicity to Male Reproductive System and Offspring as a Function of Oxidative Stress Induction: Rodent Studies

2021 ◽  
Vol 12 ◽  
Author(s):  
Xu Zhang ◽  
Tongtong Zhang ◽  
Xiaohan Ren ◽  
Xinglin Chen ◽  
ShangQian Wang ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Sperm Motility ◽  
Reproductive System ◽  
Sperm Quality ◽  
Sperm Count ◽  
Meta Analysis ◽  
Serum Testosterone ◽  
Male Reproductive System ◽  
Serum Testosterone Level ◽  
Testis Weight

Pyrethroids may be related to male reproductive system damage. However, the results of many previous studies are contradictory and uncertain. Therefore, a systematic review and a meta-analysis were performed to assess the relationship between pyrethroid exposure and male reproductive system damage. A total of 72 articles were identified, among which 57 were selected for meta-analysis, and 15 were selected for qualitative analysis. Pyrethroid exposure affected sperm count (SMD= -2.0424; 95% CI, -2.4699 to -1.6149), sperm motility (SMD=-3.606; 95% CI, -4.5172 to -2.6948), sperm morphology (SMD=2.686; 95% CI, 1.9744 to 3.3976), testis weight (SMD=-1.1591; 95% CI, -1.6145 to -0.7038), epididymal weight (SMD=-1.1576; 95% CI, -1.7455 to -0.5697), and serum testosterone level (SMD=-1.9194; 95% CI, -2.4589 to -1.3798) in the studies of rats. We found that gestational and lactational exposure to pyrethroids can reduce sperm count (SMD=1.8469; 95% CI, -2.9010 to -0.7927), sperm motility (SMD=-2.7151; 95% CI, -3.9574 to -1.4728), testis weight (SMD=-1.4361; 95% CI, -1.8873 to -0.9848), and epididymal weight (SMD=-0.6639; 95% CI, -0.9544 to -0.3733) of F1 offspring. Exposure to pyrethroids can increase malondialdehyde (SMD=3.3451; 95% CI 1.9914 to 4.6988) oxide in testes and can reduce the activities of glutathione (SMD=-2.075; 95% CI -3.0651 to -1.0848), superoxide dismutase (SMD=-2.4856; 95% CI -3.9612 to -1.0100), and catalase (SMD=-2.7564; 95% CI -3.9788 to -1.5340). Pyrethroid exposure and oxidative stress could damage male sperm quality. Gestational and lactational pyrethroid exposure affects the reproductive system of F1 offspring.

scholarly journals Seminal Plasma Proteomic Biomarkers of Oxidative Stress

2020 ◽  
Vol 21 (23) ◽  
pp. 9113
Author(s):  
Rossella Cannarella ◽  
Andrea Crafa ◽  
Federica Barbagallo ◽  
Laura M. Mongioì ◽  
Rosita A. Condorelli ◽  
...  
Keyword(s):  
Oxidative Stress ◽  
Male Infertility ◽  
Seminal Plasma ◽  
Molecular Mechanisms ◽  
Seminal Fluid ◽  
Accessory Gland ◽  
Male Accessory Gland ◽  
Abnormal Sperm ◽  
Clinical Conditions ◽  
Biomarkers Of Oxidative Stress

The prevalence of idiopathic male infertility is high, up to 75% of patients with abnormal sperm parameters. Hence, the research of its causes is mandatory. Oxidative stress (OS) can be responsible for male infertility in 30–80% of cases. In recent years, seminal plasma (SP) proteomics has developed as a useful tool to provide biomarkers of specific diseases. This systematic review aims to collect the available evidence on the changes of SP proteome in patients exposed to OS to provide possible SP biomarkers of sperm OS. To accomplish this, the following keyterms “seminal fluid proteome”, “seminal plasma proteome”, “oxidative stress”, and “sperm oxidative stress” were used and 137 records were found. Among these, 17 were finally included. Nine proteins involved with OS were found overexpressed in patients with OS. Twenty-three proteins were found differentially expressed in patients with clinical conditions associated with OS, such as varicocele, male accessory gland infection/inflammation, cigarette smoke, and obesity. These proteins do not seem to overlap among the clinical conditions taken into account. We speculate that specific SP proteins may mediate OS in different clinical conditions. Altogether, these results suggest that proteomics could help to better understand some of the molecular mechanisms involved in the pathogenesis of infertility. However, further studies are needed to identify potential biomarkers of male infertility with valuable clinical significance.

P–053 Concentration of bisphenol S in human seminal plasma negatively correlates with decreasing sperm concentration in ejaculate

2021 ◽  
Vol 36 (Supplement_1) ◽  
Author(s):  
K Franzová ◽  
M Ješeta ◽  
J Navrátilová ◽  
S Fialková ◽  
J Kalina ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Statistical Evaluation ◽  
Seminal Fluid ◽  
Sperm Count ◽  
Sperm Concentration ◽  
Dna Integrity ◽  
Negative Effects ◽  
Bisphenol S ◽  
Progressive Motility ◽  
Significant Difference

Abstract Study question Is there a relationship between concentration of bisphenol S in seminal fluid and spermiogram parameters? Summary answer Bisphenol S was detected in 81% of seminal plasma samples. Negative correlation was found between BPS concentration and total sperm count in normozoospermic men. What is known already Human spermatogenesis can be influenced by a range of chemicals present in our environment. Bisphenol S (BPS) is a very frequent compound commonly used as a softener in production of plastics, where it has replaced bisphenol A. It is an endocrine disruptor frequently associated with negative effects on reproduction. It has been observed that BPS can affect testicular development in rodent males. In addition, it has cytotoxic, reprotoxic and neurotoxic effects and induces the oxidative stress bringing negative effects on spermatogenesis. BPS has been detected in food, drinks or cosmetics. Its direct effect on spermatozoa or spermatogenesis is still unclear. Study design, size, duration A total of 38 patients (25 normozoospermic) aged 24 to 42 years, non-smokers, with BMI between 19,9–32,9 were included in this prospective study from 2018 to 2020. None of them had varicocele, urogenital infections or other urological problems. Their seminal plasma was separated by centrifugation. BPS was extracted using solvent extraction followed by preconcentration step. The samples were analysed on Agilent 6495 Triple Quadrupole. Two MS/MS transitions were used for quantitative LC-MS/MS analyses. Participants/materials, setting, methods All the men included in this study signed an informed consent and agreed with analyses of their samples. These analyses were approved by Ethical committee of University Hospital Brno. We evaluated the relationship between concentration of BPS in seminal plasma, sperm concentration, total sperm count, total motility, progressive motility, morphology and fragmentation of DNA in spermatozoa. Statistical evaluation was performed by one individual one-dimensional regression model (p-value lower than 0.05 were considered as statistically significant). Main results and the role of chance The examination revealed the presence of BPS in 31 samples of seminal plasma (81% of all the samples). In 6 samples, the concentration was under level of detection and in one sample under level of quantification. In 7 samples, a very high concentration was detected (&gt;0.1 ng/ml). These values were then compared to spermiogram parameters and sperm DNA integrity. There were no significant differences between the concentration of BPS and morphology of spermatozoa, progressive motility and total motility. In case of the DNA integrity, the opposite trend was observed, lower proportions of spermatozoa with fragmented DNA were found in samples with higher concentrations of BPS. Evaluation of sperm concentration and BPS concentration showed relationship of increasing BPS concentration with significantly lower sperm concentration, the differences were most obvious when only the normozoospermic men were compared. Evaluation of BPS concentration and total sperm count revealed the same trend with statistically significant difference in the category of normozoospermic men. Due to the small number of samples, a negative effect of extreme values on the statistical evaluation cannot be excluded. Currently, more analyses focused on detection of BPS in seminal plasma are carried out in order to obtain sufficiently larger data set. Limitations, reasons for caution A limitation is the number of samples included and analysed in this study, which slightly reduced the power of statistical analysis. Wider implications of the findings: These results document that BPS was present in 81% of analysed samples. Knowing the concentration of BPS in seminal fluid is important for understandig of impact of BPS on male fertility. Our future work will be focused on detection of other bisphenols in seminal plasma. Trial registration number MH CZ – DRO (FNBr, 65269705), AZV NV18–01–00544, Czech Ministry of Education, Youth and Sports (CZ.02.2.69/0.0/0.0/19_074/0012727)

Hydroxy-selenomethionine as an organic source of selenium in the diet improves boar reproductive performance in artificial insemination programs

2021 ◽  
Author(s):  
Ana Paula P Pavaneli ◽  
Cristian H G Martinez ◽  
Denis H Nakasone ◽  
Ana Carolina Pedrosa ◽  
Maitê V Mendonça ◽  
...  
Keyword(s):  
Seminal Plasma ◽  
Reproductive Performance ◽  
Artificial Insemination ◽  
Semen Quality ◽  
Sperm Quality ◽  
Sperm Count ◽  
Dietary Treatments ◽  
Gpx Activity ◽  
Organic Source

Abstract This study aimed to compare different selenium (Se) sources in the diet on boar's semen quality and fertility. For this, 28 boars aged 8 to 28 months were fed with the following dietary treatments for 95 days: 0.3 mg Se/kg as sodium selenite (SS, n = 14) and 0.3 mg Se/kg as hydroxy-selenomethionine (OH-SeMet, n = 14). During this period, two experiments were carried out. In experiment 1, the semen of all boars was evaluated every 2 weeks. Raw semen was initially evaluated for the processing of seminal doses, which were stored at 17 °C for 72 h, followed by sperm quality assessments. Furthermore, Se concentration and glutathione peroxidase (GPx) activity were measured in the seminal plasma. In experiment 2, 728 females were inseminated weekly with seminal doses from boars of the different experimental groups to further assess in vivo fertility and litter characteristics. Results demonstrated that boars fed OH-SeMet had more Se in their seminal plasma (p &lt; 0.05), showing the greater bioavailability of the organic source in the male reproductive system. Moreover, boars fed OH-SeMet tended (p &lt; 0.10) towards a higher total sperm count in the ejaculate (66.60 vs. 56.57 × 10 9 sperm), and the number of seminal doses (22.11 vs. 18.86; 3 × 10 9 sperm/dose) when compared to those fed SS. No effect of the dietary treatments was observed on GPx activity in seminal plasma (p &gt; 0.05), as well as on raw and stored semen quality (p &gt; 0.05). Under in vivo conditions, seminal doses from boars fed OH-SeMet tended (p &lt; 0.10) towards a higher pregnancy rate at weeks 3, 5, and 8, and also resulted in a higher (p &lt; 0.05) percentage of pregnant females in the overall period (99.30 vs. 97.00). In conclusion, the replacement of SS with OH-SeMet in boars' diet can improve sperm production and results in better reproductive performance for them, bringing greater productivity and profitability to artificial insemination centers and commercial pig farms.

Heightened Sulfur Amino Acid Oxidation in Plasma and Erythrocytes in β-Thalassemia Major.

Blood ◽  
2009 ◽  
Vol 114 (22) ◽  
pp. 4065-4065
Author(s):  
Anurag K. Agrawal ◽  
Jung H. Suh ◽  
Bruce N. Ames ◽  
Elliott P. Vichinsky ◽  
Ashutosh Lal
Keyword(s):  
Oxidative Stress ◽  
Amino Acid ◽  
Iron Overload ◽  
Metabolic Pathways ◽  
Iron Concentration ◽  
Thalassemia Major ◽  
Healthy Controls ◽  
Liver Iron ◽  
Redox States ◽  
Plasma Compartment

Abstract Abstract 4065 Poster Board III-1000 Background Oxidative stress in β-thalassemia major is a well documented problem thought in part to be due to transfusional iron overload. Sulfur amino acid (SAA)-derived metabolites, chiefly glutathione (GSH), are critical components of cellular antioxidant defense. Inability to adequately regulate endogenous antioxidant status may lead to further exacerbation of oxidative stress. To characterize whether alteration in SAA metabolism contributes to heightened oxidative stress in thalassemia, we utilized a novel liquid chromatography linked electrospray positive tandem mass spectrometric (LC-MS/MS) technique to simultaneously quantify the redox states of cysteine (Cys) and GSH as well as the concentrations of major amino acid-derived metabolites in plasma and erythrocytes. Methods After institutional review board approval, adult and pediatric subjects with β-thalassemia major were recruited for fasting blood samples drawn immediately prior to the next scheduled blood transfusion. Plasma and red blood cell samples were analyzed using the LC-MS/MS technique (J Chromatogr B 2009;in press) and compared with healthy controls. Results Twelve subjects with β-thalassemia major were recruited. The median age of the group was 26.5 years (range 11-41). The median liver iron concentration (LIC) determined by ferritometer was 11.9 mg/g dry-wt (range 1.0-34.8). The median ferritin level for the group was 1980 ng/mL (379-4730). Evidence of myocardial iron overload (T2* <20 msec) was present in 5 of 11 subjects. Plasma SAA redox analysis in these subjects showed significant oxidation of Cys and GSH. Total Cys redox status (Cysteine/2*Cystine ratios) decreased from 0.1 ± 0.01 in healthy controls to 0.08 ± 0.02 in thalassemia (p=0.02). Similarly, the plasma GSH redox state (GSH/2*GSSG ratio) decreased from 38.9 ± 13.7 in healthy controls to 6.7 ± 5.3 in thalassemia subjects (p=0.005). The total GSH and total Cys pools in the plasma did not differ between the two groups. Mirroring the patterns observed in the plasma compartment, the proportion of erythrocyte GSSG was also significantly elevated in thalassemia (0.65% vs. 0.04%, p=0.009), without a concomitant decline in total erythrocyte GSH pool. A significant 3-fold elevation in plasma cystathionine and S-adenosylmethionine levels suggests that key SAA metabolic pathways to augment synthesis of GSH may be up-regulated in thalassemia. In addition to alterations in SAA metabolite profiles, the thalassemia group was >3-fold deficient in plasma valine, spermine and citrulline compared with controls. We found that within this small group, the level of iron overload measured by serum ferritin, liver iron concentration and cardiac MRI T2* showed no correlation with the amount of oxidative stress measured by glutathione redox. Discussion Assessment of SAA redox states and metabolic pathways has the potential to be a novel important marker in β-thalassemia major. The alterations in both Cys and GSH redox states without concomitant decline in their total concentration suggests either a significant increased rate of SAA oxidation and/or a decline in the capacity for NADPH-dependent reduction of oxidized disulfides. The latter may be supported by the evidence of profound hypocitrullinemia in thalassemia, which has been noted in subjects with inherited mitochondrial disorders (Proc Nat Acad Sci U S A 2009;106:3941-5). Finally, we show that the oxidative environment in the plasma affects the transfused erythrocytes which acquire characteristics of abnormal thiol metabolites observed in the plasma compartment. The demonstration of significant oxidative stress even in subjects with low body iron burden suggests that currently acceptable LIC levels may not be optimal, or that other contributors to the pro-oxidant state should be sought in thalassemia. Disclosures: No relevant conflicts of interest to declare.

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