Potential Quality Markers Research Articles

Investigation of sperm telomere length as a potential marker of paternal genome integrity and semen quality

Recent studies have reported shorter sperm telomere length (STL) in men with idiopathic infertility. The aim of this study was to measure STL in semen samples from men to evaluate whether STL variation is associated with chromosomal abnormality, DNA fragmentation, traditional semen parameters, IVF outcome, or all four factors. A significant correlation between telomere length and diploidy was observed (P = 0.037). Additionally, STL was found to be positively associated with sperm count (P = 0.006); oligospermic samples had particularly short telomeres (0.9 ± 0.1 versus 1.4 ± 0.1; P = 0.0019). The results confirmed a link between sperm DNA fragmentation and aneuploidy, previously proposed (P = 0.009). A negative relationship was demonstrated between sperm concentration and aneuploidy and Sperm DNA framentation (P = 0.03, P < 0.0001, respectively). For a subset of 51 of the 73 sperm samples used for fertilization, IVF outcomes were known. A total of 17.6% of these samples had atypical STLs. None of these samples produced an ongoing pregnancy. In contrast, the pregnancy rate for samples that had STLs in the normal range was 35.7% (P = 0.044). In conclusion, STL has potential as a fast and inexpensive form of sperm quality assessment.

Platelet MicroRNAs: An Overview

MicroRNAs (miRNAs) are short ~22-nucleotide noncoding RNA that have been found to influence the expression of many genes and cellular processes by either repressing translation or degrading messenger RNA transcripts. Platelet miRNA expression has been shown to be perturbed during ex vivo storage of platelets and in platelet-associated disorders. Although bioinformatics-based miRNA target predictions have been established, direct experimental validation of the role of miRNAs in platelet biology has been rather slow. Target prediction studies are, nonetheless, valuable in directing the design of appropriate experiments to test specific miRNA:messenger RNA interactions relevant to the underlying mechanisms of platelet function in general and in disease as well as in ex vivo storage-associated "storage lesions," a collective term used to include physiologic, biochemical, and morphologic changes that occur in stored platelets. This brief review will focus on emerging human platelet miRNA studies to emphasize their potential role relevant to transfusion medicine field in terms of regulating platelet signaling pathways, markers of platelet associated disorders, and remote impactors of gene expression (intercellular biomodulators) as well as potential platelet quality markers of storage and pathogen reduction treatments.

Identification and Validation of a Potential Marker of Tissue Quality Using Gene Expression Analysis of Human Colorectal Tissue.

Correlative studies have identified numerous biomarkers that are individualizing therapy across many medical specialties, including oncology. Accurate interpretation of these studies requires the collection of tissue samples of sufficient quality. Tissue quality can be measured by changes in levels of gene expression and can be influenced by many factors including pre-analytical conditions, ischemic effects and the surgical collection procedure itself. However, as yet there are no reliable biomarkers of tissue quality at researchers’ disposal. The aim of the current study was to identify genes with expression patterns that fluctuated reproducibly in response to typical post-surgical stress (ischemia) in order to identify a specific marker of tissue quality. All tissue samples were obtained from patients with primary colorectal carcinoma (CRC) (N = 40) either via colonoscopy prior to surgery, or by surgical resection. Surgically resected tissue samples were divided into three groups and subjected to cold ischemia for 10, 20 or 45 minutes. Normal colorectal tissue and CRC tissue was analyzed using microarray and quantitative real-time PCR (qPCR). Comparing changes in gene expression between pre- and post-surgical tissue using microarray analysis identified a list of potential tissue quality biomarkers and this list was validated using qPCR. Results revealed that post-operative ischemia significantly alters gene expression in normal and CRC tissue samples. Both microarray analysis and qPCR revealed regulator of G-protein signaling 1 (RGS1) as a potential marker of CRC tissue quality and eukaryotic translation elongation factor 1 alpha 1 (EEF1A1) as a potential reference gene of post-operative tissue quality. Larger studies with additional time points and endpoints will be needed to confirm these results.

Identification of innovative potential quality markers in rocket and melon fresh-cut produce

Ready-to-eat fresh cut produce are exposed to pre- and postharvest abiotic stresses during the production chain. Our work aimed to identify stress responsive genes as new molecular markers of quality that can be widely applied to leaves and fruits and easily determined at any stage of the production chain. Stress responsive genes associated with quality losses were isolated in rocket and melon fresh-cut produce and their expression levels analyzed by quantitative real time PCR (qRT-PCR) at different time points after harvest at 20°C and 4°C. qRT-PCR results were supported by correlation analysis with physiological and biochemical determinations evaluated at the same conditions such as chlorophyll a fluorescence indices, total, reducing sugars, sucrose, ethylene, ascorbic acid, lipid peroxidation and reactive oxygen species. In both species the putative molecular markers increased their expression soon after harvest suggesting a possible use as novel and objective quality markers of fresh-cut produces.

Simplified sample preparation and rapid detection by RP‐HPLC/FLD of tocopherols and tocotrienols in margarines: Preliminary screening of plant fats—potential quality markers

A simplified, rapid, and accurate method for sample preparation and determination of tocopherols and tocotrienols (vitamin E) in margarine was developed. For this purpose, 28 margarines with different amounts of fat (20, 39, 60, 70, 75, and 80%) and of two types (spreads and cooking fats) were used. Sample preparation was reduced to a limited number of rapid steps: weighting, melting, diluting, vortexing, and centrifugation, following which the samples were directly injected to the HPLC system. The developed method prevented analyte losses and was characterized by good repeatability and reproducibility (1.4–5.5% and 2.9–9.9%, respectively) and accuracy range (97.9–101.2%) as well as recovery of tocopherols and tocotrienols when compared to the saponification method (89.4–104.4%), without any interference of other compounds from the margarine matrix and thus reduced the risk of potential measurement errors. Fat content in margarine samples did not have any negative impact on precision, accuracy or recovery of tocopherols and tocotrienols. Concentrations of tocopherol (T) and tocotrienol (T3) homologues in margarine samples depended on the fat content and ranged from 2.65–26.27 (α‐T), 0.02–0.70 (β‐T), 4.23–23.60 (γ‐T), 0.08–2.34 (δ‐T), 0.39–15.05 (α‐T3), 0.00–1.36 (β‐T3), 0.49–20.08 (γ‐T3), and 0.00–5.28 mg/100 g (δ‐T3).Practical applications: The necessity to ensure high quality and nutritional value of food products results in the need to develop new high quality analytical methods. This paper describes a simple, rapid and accurate method of sample preparation, preventing analyte losses, and facilitating fast determination of all tocopherol and tocotrienol homologues by RP‐HPLC/FLD in margarines containing different amounts of fat (20, 39, 60, 70, 75, and 80%). Margarines are a significant source of vitamin E in our daily diet; therefore, the developed method could be implemented in industry both in routine control and to improve nutrition information about the product provided on the label and also as the first screening test to assess the possible origin of the fat used for margarine production (with tocopherols and tocotrienols as quality markers).Tocopherol and tocotrienol homologues (α, β, γ, and δ) in margarine samples with different amounts of fat (20–80%) were successfully separated by using RP‐HPLC/FLD. The developed method is characterized by ease of implementation, speed, reliability, prevents analyte losses (quantification errors), saves solvents, and does not affect quality of analysis. As the developed method has been simplified and accelerated (reducing time and costs), it may be applied as the preliminary screening test to assess the possible origin of the fat used for margarine production (tocopherols and tocotrienols as quality markers).

Follicular fluid oocyte/cumulus-free DNA concentrations as a potential biomolecular marker of embryo quality and IVF outcome.

The present prospective study examined the follicular fluid oocyte/cumulus-free DNA concentrations (ff o/c-free DNA) during ovarian stimulation and the possible association between ff o/c-free DNA and embryological results such as embryo quality and pregnancy rate. Eighty-three women undergoing IV/ICSI-ET treatments were prospectively included in this study. ff o/c-free DNA was determined by conventional quantitative real time PCR-Sybr green detection approach. The 83 ff samples were categorized in two groups: group 1 (n = 62) with cumulus oocytes complexes (CoCs) ≥2 and group 2 (n = 21) with CoCs = 1. Group 1 revealed significant higher embryo quality in terms of mean score of embryo transfer (MSET), but lower ff o/c-free DNA concentrations compared to group 2. The two groups showed comparable pregnancy rates (positive hCG and clinical pregnancy). The higher the ff o/c-free DNA concentration, the lower the number of produced oocytes. ff o/c-free DNA did not seem to have any direct role in the IVF outcome. Further research is required to clarify whether ff o/c-free DNA is a biomolecular marker of embryo quality and IVF outcome.

70 PROTEIN EXPRESSION OF THE INSULIN-LIKE GROWTH FACTOR 1 RECEPTOR DURING BOVINE PRE-IMPLANTATION EMBRYONIC DEVELOPMENT

Insulin-like growth factor 1 (IGF1) is essential for regulating physiological processes such as growth and development of fetal and placental tissues (Bauer et al. 1998, Fowden 2003). During early embryonic development, IGF1 plays an important role, as it leads to a reduction of apoptosis and decreases early embryonic mortality (Block et al. 2007). The signal transduction of IGF1 is carried out by its specific binding to the membrane-embedded insulin-like growth factor 1 receptor (IGF1R). The expression of IGF1R is a potential quality marker of in vitro produced embryos (Liu et al. 1997, Yaseen et al. 2001). Thus far, analysis of the relative amount of specific transcripts is the method of choice to study bovine pre-implantation embryos, as information on protein expression is scarce. Therefore, it is of great interest to analyse protein expression and to determine if and to which extent these results differ from results obtained in previous mRNA expression analyses. In the present study, a total of 4800 cumulus-oocyte-complexes were deployed in 60 in vitro produced runs. The cleavage rates averaged 57.4 ± 7.3% and blastocyst rates were 27.6 ± 7.5% at Day 8 of culture. Embryos at the blastocyst stage were frozen and stored at –80°C for further experiments. The protein expression of the IGF1R during early embryonic development was investigated by Western blot analysis testing 8 different antibodies. Seven of these antibodies were commercially available and mainly not tested in the bovine species. Only 1 of these antibodies resulted in a weak signal for the IGF1R protein in bovine blastocysts. Therefore, a specific peptide antibody against 2 peptide sequences of the α unit of the bovine IGF1R was produced. The analysis of the IGF1R protein with this antibody resulted in the determination of a signal in a pool of 100 blastocysts, which was weaker than in the positive control (20 μg of bovine liver protein extract). The detection of the IGF1R protein localization was possible in all different stages of embryonic development from the zygote to the expanded blastocyst using immunfluorescence staining with the specific peptide antibody. The IGF1R protein was mainly expressed in the plasma membrane of single blastomeres and also weakly in the cytoplasm. As the early bovine embryo expresses IGF1R throughout all stages, the main function of IGF1 in embryonic development needs to be further elucidated. We gratefully acknowledge the financial support of the H. Wilhelm Schaumann Foundation (Hamburg, Germany).

Effects of variable protein and lipid proportion in gilthead sea bream (Sparus aurata) diets on fillet structure and quality

Juvenile gilthead sea bream (Sparus aurata) were fed with seven experimental diets containing different proportions of protein and lipid (P/L): 38/29, 41/26, 44/25, 47/23, 50/22, 54/21 and 57/20 per cent of dry matter, respectively. After 12 weeks, fish fed on diet 38/29 (P/L) showed the lowest food efficiency and final weight. A correlation between the hepatosomatic index and fat content in the diets was observed, together with a decrease in muscle glycogen in animals fed with a 38/29 (P/L) diet. Whilst no statistical differences were found in flesh texture, a clear relationship was observed between elasticity and fibre density. Fillet Chroma was altered in the animals fed with the 38/29 (P/L) diet. The same group showed a decrease in hepatic growth factor receptor (c-met) and proliferating cell nuclear antigen (PCNA) expression in white muscle. Protein content in the diets correlated with mammalian target of rapamycin (mTOR) phosphorylation in muscle. In conclusion, we have shown that the substitution of protein by lipid in sea bream diets can be performed up to a certain percentage without affecting fish growth, and we have demonstrated that several molecules such as MyoD, mTOR, PCNA and c-met are affected by the dietary ratio of protein to lipid and could be potential flesh quality markers.

Platelet lysate suppresses the expression of lipocalin-type prostaglandin D2 synthase that positively controls adipogenic differentiation of human mesenchymal stromal cells

Mesenchymal stromal cells (MSCs) have been shown to display a considerable therapeutic potential in cellular therapies. However, harmful adipogenic maldifferentiation of transplanted MSCs may seriously threaten the success of this therapeutic approach. We have previously demonstrated that using platelet lysate (PL) instead of widely used fetal calf serum (FCS) diminished lipid accumulation in adipogenically stimulated human MSCs and identified, among others, lipocalin-type prostaglandin D2 synthase (L-PGDS) as a gene suppressed in PL-supplemented MSCs. Here, we investigated the role of PL and putatively pro-adipogenic L-PGDS in human MSC adipogenesis.Next to strongly reduced levels of L-PGDS we show that PL-supplemented MSCs display markedly decreased expression of adipogenic master regulators and differentiation markers, both before and after induction of adipocyte differentiation. The low adipogenic differentiation capability of PL-supplemented MSCs could be partially restored by exogenous addition of L-PGDS protein. Conversely, siRNA-mediated downregulation of L-PGDS in FCS-supplemented MSCs profoundly reduced adipocyte differentiation. In contrast, inhibiting endogenous prostaglandin synthesis by aspirin did not reduce differentiation, suggesting that a mechanism such as lipid shuttling but not the prostaglandin D2 synthase activity of L-PGDS is critical for adipogenesis.Our data demonstrate that L-PGDS is a novel pro-adipogenic factor in human MSCs which might be of relevance in adipocyte metabolism and disease. L-PGDS gene expression is a potential quality marker for human MSCs, as it might predict unwanted adipogenic differentiation after MSC transplantation.

Management of long-term hypothyroidism: a potential marker of quality of medicines reconciliation in the intensive care unit†

Significant errors can be made during medication prescribing, dispensing and administration. One source of error and potential for harm is unintentional omission. Medicines reconciliation seeks to reduce the impact of this between transfer of care. In long-term hypothyroidism, patients are dependent upon levothyroxine and there are few contraindications to its prescription. We considered levothyroxine prescription in long-term hypothyroidism as a marker of medicines reconciliation on admission and during stay in the intensive care unit (ICU). A retrospective chart review was undertaken in a tertiary referral university ICU with all patients who were receiving long-term levothyroxine therapy identified. Notes were reviewed for the presence of thyroid-replacement prescription and for thyroid function tests, in addition to demographic, length of stay and mortality data. Thyroid-replacement therapy was not prescribed for more than 7 days in 23/133 (17.3%) patients and omitted entirely in three patients. A further 28/133 (21.1%) patients were intolerant of enteral feeding for more than 7 days and were thus unable to have oral levothyroxine administered. None of these patients received parenteral therapy. Thyroid function tests were performed in 104/133 (78.2%) patients. Prescription of chronic therapy, in this case thyroid-replacement therapy, was inadequate. This highlights the need for a progressive medicines-reconciliation process embedded within the daily ICU programme.

Pressure and oxygen debt on bypass – potential quality markers of perfusion?

No markers of quality of perfusion pressure and oxygen delivery during cardiopulmonary bypass (CPB), to complement rewarming rate, maximum temperature on rewarming, lowest haematocrit, and blood glucose, exist. Using the electronic acquisition of blood pressure on bypass (JOCAP system), the percentage of time perfusion pressure was below 30, 40, 50, 60 and 70 mmHg, average deviance, confidence interval, median, mode, standard deviation, variance, and average, maximum and cumulative oxygen debt were calculated. Numerous different readouts of achievement of maintenance of constant pressure on bypass and oxygen debt are now easily achievable with perfusion electronic data management systems. Mean, median, and mode offer poor discrimination of pressure control during CPB. Percentage of time perfusion pressure was below 30, 40, 50, 60 and 70 mmHg, average deviance, confidence interval, and standard deviation all have discriminatory power, but need clinical correlation for their significance. A composite score involving non-pressure readouts (e.g. oxygen delivery, arterial and venous saturations, and flow rates) may need to be integrated into any perfusion quality marker. Assessment of adequacy of constant perfusion pressure and oxygen delivery may allow the scientific evaluation of pressure and oxygen delivery on bypass for patients to be compared accurately. Currently, in studies involving CPB, blood pressure targets are stated with no quantitative assessment of adequacy of achievement of these targets. Electronic data monitoring during cardiopulmonary bypass, when correlated with clinical outcome, may help to provide a marker of quality of perfusion pressure during CPB and may, indeed, allow patient-specific perfusion pressure strategies to be developed.

Parameters affecting the accuracy of the MALDI-TOF MS determination of the phosphatidylcholine/lysophosphatidylcholine (PC/LPC) ratio as potential marker of spermatozoa quality

Matrix-assisted laser desorption and ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) is increasingly used to characterize (phospho)lipids. However, quantitative MALDI data are often questioned because ion suppression may occur if mixtures are analyzed. Therefore, relative (but no absolute) data are normally derived from the MALDI mass spectra of lipid mixtures. We are particularly interested in the phosphatidylcholine/lysophosphatidylcholine (PC/LPC) ratio because it seems to represent a suitable measure of the inflammatory activity. In this study, different parameters affecting the achievable accuracy of the MALDI-TOF MS determination of the PC/LPC ratio are compared. It will be shown that particularly the applied laser fluence as well as the used solvents influence the accuracies. Using artificial lipid mixtures it will be demonstrated that the PC/LPC ratio can be determined with an accuracy of about ±10% making the MALDI assay comparable to established methods. Finally, it will be shown that the optimized conditions are also useful to determine the PC/LPC ratios in human seminal plasma.

Proteomic profile of dry-cured ham relative to PRKAG3 or CAST genotype, level of salt and pastiness

Two-dimensional electrophoresis was used to compare dry-cured biceps femoris insoluble protein fraction according to genotype ( PRKAG3Ile199Val and CASTLys249Arg/Ser638Arg) as well as salt and pastiness level. The PRKAG3 affected mainly muscle metabolic enzymes, indicating its possible influence on muscle metabolism with heterozygotes Ile/Val appearing different from both homozygous genotypes. The effect of CAST was smaller, affecting the quantity of one actin fragment. Dry-cured ham salt and pastiness level affected a wide variety of protein spots including metabolic enzymes, plasma proteins, chaperones and myofibrillar proteins, including protein fragments, indicating the connection with proteolysis. Pastiness was associated with salt content, reflected also by the fact that many spots were affected by both factors. Despite the absence of extreme pastiness (or low salt samples), some protein spots (actin, MHC fragment, desmin fragment) exhibited important differences in intensity according to pastiness (and salt level) suggesting they could be used as potential quality markers.

Inter-chromosomal variation in aberration frequencies in human lymphocytes exposed to charged particles of LET between 0.5 and 55 keV/μm

Purpose: To investigate the distribution of chromosomal aberrations in chromosomes 2, 8 and 14 induced by charged particles, using the fluorescence in situ hybridisation (FISH) technique.Methods: Irradiation of peripheral blood from six healthy volunteers (four male and two female) was performed at the accelerators of the Joint Institute for Nuclear Research (JINR) in Dubna (Russia). Whole blood samples were irradiated with 2 and 3 Gy of protons (170 MeV/nucleon (n), linear energy transfer (LET) ≈ 0.5 keV/μm), 3.5 Gy of 12C ions (480 MeV/n, LET = 10.6 keV/μm), 3 Gy of 12C ions 500 MeV/n, LET = 12 keV/μm), 4 Gy of 7Li ions (30 MeV/n, LET ≈ 20 keV/μm) and 3 Gy of 11B ions (32 MeV/n, LET ≈ 55 keV/μm). Chromosomal aberrations were analysed in metaphase and prematurely condensed chromosomes (PCC) induced in G2-cells using calyculin A. Chromosomes 2, 8 and 14 were painted in different colours and aberrations scored with the help of an image-analysis system.Results: Chromosome 2 was generally less sensitive than expected on the basis of its DNA content. A higher than expected frequency of exchanges was found in chromosomes 8 and 14. On average, the dicentric frequency in chromosome 2 was higher than the translocation frequency, whereas variable dicentric to translocation ratios were observed in chromosomes 8 and 14. When aberrations in all painted chromosomes were summed up the ratio was close to 1. The frequency of complex aberrations correlated with LET.Conclusion: In lymphocytes of donors studied in this work chromosome 2 appears to be consistently less sensitive to protons and heavy ions than chromosomes 8 and 14. Complex aberrations appear to be a potential marker of radiation quality.

Dietary Total Antioxidant Capacity: A Novel Indicator of Diet Quality in Healthy Young Adults

Background: Overall diet quality measurements have been suggested as a useful tool to assess diet-disease relationships. Oxidative stress has been related to the development of obesity and other chronic diseases. Furthermore, antioxidant intake is being considered as protective against cell oxidative damage and related metabolic complications.Objective: To evaluate potential associations between the dietary total antioxidant capacity of foods (TAC), the energy density of the diet, and other relevant nutritional quality indexes in healthy young adults.Methods: Several anthropometric variables from 153 healthy participants (20.8 ± 2.7 years) included in this study were measured. Dietary intake was assessed by a validated food-frequency questionnaire, which was also used to calculate the dietary TAC and for daily energy intake adjustment.Results: Positive significant associations were found between dietary TAC and Mediterranean energy density hypothesis–oriented dietary scores (Mediterranean Diet Score, Alternate Mediterranean Diet Score, Modified Mediterranean Diet Score), non-Mediterranean hypothesis–oriented dietary scores (Healthy Eating Index, Alternate Healthy Eating Index, Diet Quality Index-International, Diet Quality Index-Revised), and diversity of food intake indicators (Recommended Food Score, Quantitative Index for Dietary Diversity in terms of total energy intake). The Mediterranean Diet Quality Index and Diet Quality Index scores (a Mediterranean and a non–Mediterranean hypothesis–oriented dietary score, respectively), whose lower values refer to a higher diet quality, decreased with higher values of dietary TAC. Energy density was also inversely associated with dietary TAC.Conclusion: These data suggest that dietary TAC, as a measure of antioxidant intake, may also be a potential marker of diet quality in healthy subjects, providing a novel approach to assess the role of antioxidant intake on health promotion and diet-based therapies.

Cut-off Value of Reactive Oxygen Species for Predicting Semen Quality and Fertilization Outcome

Though the role of reactive oxygen species (ROS) in male infertility is widely investigated worldwide, there is a lack of adequate information on the cut-off value of ROS beyond which ART outcome may be adversely affected. The objective of the present study is to establish an upper level of ROS in semen samples which can be considered as a potential marker of good semen quality. Semen ejaculates were randomly collected from 338 male partners of infertile couples. The upper critical limit (UCL) of ROS was calculated from the control chart of normozoospermic patients and found to be 0.75×106 counted photons per minute (cpm)/10 million cells. This was further validated by estimating ROS in 17 proven fertile men and 38 patients undergoing intra cytoplasmic sperm injection (ICSI). As expected, all abnormal semen samples exhibited higher ROS as compared to nomozoospermic and proven fertile samples. All semen samples from proven fertile volunteers were found to be <0.075×106 cpm/10 million cells. On the basis of the calculated UCL, ICSI patients were divided into two groups: Group I (<UCL) and Group II (>UCL). The semen parameters, fertilization rate and pregnancy outcome were found to be significantly affected in Group II. Significant difference in live birth-rates was observed between Group I (47.6%) and Group II (17.6%) while no live-birth was recorded for ROS level >0.1×106 cpm/10 million cells. It is concluded that the upper cut-off value of normal semen samples that correlates with good semen quality is, therefore, in the order of 0.075–0.1×106 cpm/10 million cells. In addition to the WHO [] semen analysis, this range is expected to assist andrologists and clinicians in predicting semen quality and fertilization outcome in patients with male factor infertility undergoing ICSI.

Cell Wall Fractions Isolated from Outer Layers of Rye Grain by Sequential Treatment with α-Amylase and Proteinase: Structural Investigation of Polymers in Two Ryes with Contrasting Breadmaking Quality

Recent studies have indicated that some structural features of arabinoxylans, the major cell wall polysaccharides, might be potential quality markers in the selection of rye breeding materials. To specify the most appropriate characteristics, the differences in the structure of cell wall components were studied in two ryes with high and low breadmaking qualities. Two cell wall fractions were isolated from the outer layers of the grain (pooled shorts and bran fractions) by a consecutive water extraction with alpha-amylase (WE-A) and proteinase K (WE-P). Polysaccharides predominated in the WE-A fraction (approximately 64%, mainly arabinoxylans). By contrast, the WE-P fraction contained mostly protein (approximately 63%), and its level of polysaccharides was relatively low (approximately 18%). The 1H NMR and sugar analysis of the ammonium sulfate precipitated subfractions revealed that the WE-A was built of four arabinoxylan populations with marked structural differences (arabinose-to-xylose ratios, Ara/Xyl, of approximately 0.3, 0.5, 0.8, and 1.2). Instead, the arabinoxylans present in the WE-P were generally enriched in disubstituted xylopyranosyl residues. The ratio of phenolic components to arabinose residues in the WE-P fraction (indicated by 1H NMR) and the proportion of polymers with the highest molecular weights in the WE-A fraction (revealed by HPSEC) distinguished well two ryes with diverse breadmaking qualities. Much less obvious differences between both ryes were observed in the ratio of amide I to amide II band intensities of FTIR spectra for the WE-P and in the level of phenolic acids and ferulic acid dehydrodimers for both cell wall preparations.

Expression of Retinoid X Receptor Transcripts and their Significance for Developmental Competence in In Vitro‐produced Pre‐implantation‐stage Bovine Embryos

Retinoid X receptors are frequently implicated in their role in embryonic development. However, there is no report regarding their expression in embryos with different qualities. In this study, expression pattern and levels of retinoid X receptor transcripts in different qualities and stages of in vitro-produced pre-implantation bovine embryos were examined using real-time polymerase chain reaction (PCR) techniques. Four independent cDNA pools per sample were prepared from mRNAs isolated from different developmental-stage embryos, after classifying them to be of good and bad quality. Primers were also designed to amplify products with PCR. Based on analysis, except retinoid X receptor gamma (RXRG) transcripts, the remaining two types (alpha and beta forms) were expressed in all samples with significantly higher (p < or = 0.05) levels until the eight-cell stage compared with the stages thereafter. However, the transcript levels for bad-quality embryos were consistently lower in all cases in comparison with good-quality embryos. RXRG was identified in 39% of the samples, but all in the pre-embryonic genome activation development stage. Therefore, strong expression patterns of these transcripts in earlier stages indicates their possible role during the maternal phase of embryo development, and the variations of copy number in embryos with different qualities substantiates their possible candidacy as potential quality markers. Moreover, identifying the transcript variations in different qualities and expression of RXRG at these stages is a novel input to the pre-implantation-stage knowledge.

Seminal carnitine and acetylcarnitine content and carnitine acetyltransferase activity in young Maremmano stallions

The reproductive characteristics and seminal carnitine and acetylcarnitine content as well as carnitine acetyltransferase activity of young Maremmano stallions ( n=25) are reported. The stallions were subjected to semen collection in November and January; in each trial two ejaculates were collected 1 h apart. The total motile morphologically normal spermatozoa (TMMNS) and the progressively motile spermatozoa at collection and during storage at +4°C were evaluated. Seminal l-carnitine (LC), acetylcarnitine (AC), pyruvate and lactate were measured using spectrophotometric methods, whereas carnitine acetyltransferase activity was measured by radioenzymatic methods. Since there were no major significant differences in seminal and biochemical characteristics between the November and January trials, data were also pooled for the first and second ejaculates. Significant differences ( P<0.001) were observed between the first and second ejaculates for sperm count (0.249±0.025 versus 0.133±0.014×10 9/ml), total number spermatozoa by ejaculate (12.81±1.23 versus 6.36±0.77×10 9), progressively motile spermatozoa (48.6±3.0 versus 52.6±3.0%) and TMMNS (3.35±0.50 versus 2.02±0.37×10 9). In the raw semen the LC and AC were significantly higher in the first ejaculate than in the second ( P<0.001), whereas, pyruvate and pyruvate/lactate ratio were higher in the second ejaculate ( P<0.05). Seminal plasma AC and LC concentrations resulted higher in the first ejaculate ( P<0.001). The pyruvate/lactate ratio was higher in the second ejaculate ( P<0.05). Both raw semen and seminal plasma LC and AC concentrations were positively correlated with spermatozoa concentration ( P<0.01); in raw semen AC was also correlated to TMMNS ( P<0.01). Lactate levels of raw semen was correlated to progressively motile spermatozoa after storage ( P<0.01). In the second ejaculate, significant correlations were also observed among AC/LC ratio in raw semen and progressively motile spermatozoa after 48 and 72 h of refrigeration. Furthermore, AC levels were correlated to lactate concentration. The positive correlation between LC, AC and spermatozoa concentration, and between AC and TMMNS indicated carnitine as potential semen quality marker. Moreover, the correlation between AC/LC ratio and progressive spermatozoa motility after refrigeration, suggests that carnitine may contribute towards improving the maintenance of spermatozoa viability during in vitro storage.

Analytical Tools for Rapid, Sensitive, Quantitative Identification of Potential Meat Quality Markers

Analytical Tools for Rapid, Sensitive, Quantitative Identification of Potential Meat Quality Markers