Single nucleotide polymorphisms in the bovine SLC2A12 and SLC5A1 glucose transporter genes - the effect on gene expression and milk traits of Holstein Friesian cows.

In this study, novel single nucleotide polymorphisms (SNPs) were found in the 5'-regulatory regions (promoters) of the bovine glucose transporter (GT) genes SLC2A12 and SLC5A1. These polymorphisms were shown to associate with certain milk production traits in HF cows, including milk yield, milk composition, and somatic cell count. It was shown that the SNP g.-671C > G (NC_037336.1: g.72224078C > G) in the SLC2A12 gene could be an effective marker of cattle production traits and that genotypes CC and CG are associated with the best productivity. The polymorphisms found in the SLC5A1 gene promoter also influenced milk production traits in HF cows, albeit to a lesser extent, and we propose that these polymorphisms could be useful as genetic markers for milk production traits in marker-assisted selection (MAS); however, this must be confirmed on larger populations of cattle. In addition, the presence of polymorphisms within promoter regions appears to affect the expression of GT genes in the cow mammary gland and modify transcription factor (TF) binding capacity.

Comparative Evaluation of Sucrosomial Iron and Iron Oxide Nanoparticles as Oral Supplements in Iron Deficiency Anemia in Piglets.

Iron deficiency is the most common mammalian nutritional disorder. However, among mammalian species iron deficiency anemia (IDA), occurs regularly only in pigs. To cure IDA, piglets are routinely injected with high amounts of iron dextran (FeDex), which can lead to perturbations in iron homeostasis. Here, we evaluate the therapeutic efficacy of non-invasive supplementation with Sucrosomial iron (SI), a highly bioavailable iron supplement preventing IDA in humans and mice and various iron oxide nanoparticles (IONPs). Analysis of red blood cell indices and plasma iron parameters shows that not all iron preparations used in the study efficiently counteracted IDA comparable to FeDex-based supplementation. We found no signs of iron toxicity of any tested iron compounds, as evaluated based on the measurement of several toxicological markers that could indicate the occurrence of oxidative stress or inflammation. Neither SI nor IONPs increased hepcidin expression with alterations in ferroportin (FPN) protein level. Finally, the analysis of the piglet gut microbiota indicates the individual pattern of bacterial diversity across taxonomic levels, independent of the type of supplementation. In light of our results, SI but not IONPs used in the experiment emerges as a promising nutritional iron supplement, with a high potential to correct IDA in piglets.

The Role of Copper in the Regulation of Ferroportin Expression in Macrophages.

The critical function of ferroportin (Fpn) in maintaining iron homeostasis requires complex and multilevel control of its expression. Besides iron-dependent cellular and systemic control of Fpn expression, other metals also seem to be involved in regulating the Fpn gene. Here, we found that copper loading significantly enhanced Fpn transcription in an Nrf2-dependent manner in primary bone-marrow-derived macrophages (BMDMs). However, prolonged copper loading resulted in decreased Fpn protein abundance. Moreover, CuCl2 treatment induced Fpn expression in RAW 264.7 macrophages at both the mRNA and protein level. These data suggest that cell-type-specific regulations have an impact on Fpn protein stability after copper loading. Transcriptional suppression of Fpn after lipopolysaccharide (LPS) treatment contributes to increased iron storage inside macrophages and may result in anemia of inflammation. Here, we observed that in both primary BMDMs and RAW 264.7 macrophages, LPS treatment significantly decreased Fpn mRNA levels, but concomitant CuCl2 stimulation counteracted the transcriptional suppression of Fpn and restored its expression to the control level. Overall, we show that copper loading significantly enhances Fpn transcription in macrophages, while Fpn protein abundance in response to CuCl2 treatment, depending on macrophage type and factors specific to the macrophage population, can influence Fpn regulation in response to copper loading.

The effect of single-nucleotide polymorphism in the promoter region of bovine alpha-lactalbumin (LALBA) gene on LALBA expression in milk cells and milk traits of cows.

Polymorphisms of milk protein genes have been proposed as candidate markers for dairy production traits in cattle. In the present study, a polymorphism was detected in the 5'-flanking (promoter) region of the bovine alpha-lactalbumin (LALBA) gene, a T/C transition located at nucleotide -1,001 relative to the transcription start site g.-1001T > C (NC_037332.1:g.31183170T > C), which is recognizable with PstI restriction endonuclease. In silico analyses showed that this mutation created novel retinoid X receptor alpha and vitamin D receptor transcription factor binding sites. Real-time PCR found that cows with different genetic variants of the promoter demonstrated different levels of expression of LALBA mRNA in milk somatic cells (MSCs). The TT genotype cows demonstrated low expression, whereas those with CT demonstrated much higher expression (P < 0.05). ELISA analysis found milk LALBA protein levels also differed between the TT and CT cows (P < 0.05) and that these levels were not correlated with the mRNA abundance in MSC. Association analysis found that the g.-1001T > C polymorphism in the promoter region of the LALBA gene influenced milk production traits in Polish Holstein-Friesian cows. High daily milk yield and dry matter yield, and high lactose yield and concentration were associated with the TT genotype. The TT genotype cows also had a lower number of somatic cells in the milk, considered as an indicator of udder health status. Therefore, the TT genotype could be more desirable from the breeder's perspective.

Bone mineral density, thyroid function, and gonadal status in young adult survivors of childhood cancer.

INTRODUCTION: During the last years, changes in the diagnosis and treatment have caused a significant increase of the number of young adults who experienced cancer in childhood. This enlarging population is affected by many health problems, including multiple hormone deficiencies and bone mineral deficits. This is the first polish study assessing bone mineral density and endocrine status in young adult cancer survivors. MATERIAL AND METHODS: A total of 76 long-term survivors treated for pediatric cancer were identified. The mean age at the time of study was 24.1 ±3.5 years. Bone mineral density and TSH, fT3, fT4, FSH, LH, estradiol and testosterone level were assessed for each patient. RESULTS: Nine subjects were diagnosed with subclinical hypothyroidism. We found higher level of TSH in the study group, in comparison with control group (p = 0.015). Eighteen patients had increased level of FSH. In the study group higher number of patients with high FSH level was found in comparison with the control group (p = 0.049). A low BMD was observed in 7 patients whereas mild BMD deficits were found in 19 participants. CONCLUSIONS: In conclusion, our data show that young adult cancer survivors might experienced various hormonal problems including low bone mass, thyroid impairment and gonadal dysfunction. Some of the patients required treatment, but they were not diagnosed before this study. There is the lack of proper clinical assessment among adult childhood cancer survivors in Poland. Therefore, we demonstrated the need for a comprehensive plan for longitudinal follow-up for late effects in these population.

Serum 25-hydroxyvitamin D, osteocalcin, and parathormone status in children with meningomyelocele.

Sufficient vitamin D levels are required for normal skeletal development and mineralization. This is particularly important in children with meningomyelocele who are at an increased risk of osteoporosis. The purpose of this study was to assess serum 25-hydroxyvitamin D [25(OH)D] and the biochemical markers of bone metabolism (parathormone, osteocalcin, alkaline phosphatase, and electrolytes) in children with meningomyelocele. The patient group comprised 33 children with meningomyelocele. The mean 25(OH)D was 11.51 ± 7.87 ng/mL. A total of 97% of the subjects had a 25(OH)D level in the insufficient range (< 30 ng/mL) and 48.5% had a 25(OH)D level less than 10 ng/mL. Almost all patients had serum osteocalcin and phosphorus concentrations above the normal limits. The level of 25(OH)D negatively correlated with age and body weight. There were no correlations between the biochemical markers of bone metabolism and the ambulatory status. A significant correlation between serum 25(OH)D and osteoporosis was found.

Effect of the g.-723G-->T polymorphism in the bovine myogenic factor 5 (Myf5) gene promoter region on gene transcript level in the longissimus dorsi muscle and on meat traits of Polish Holstein-Friesian cattle.

Myogenic factor 5 (Myf5), a product of the Myf5 gene, belongs to the MRF family of basic helix-loop-helix transcription factors that regulate myogenesis. Their roles in muscle growth and development make their genes candidates for molecular markers of meat production in livestock, but nucleotide sequence polymorphism has not been thoroughly studied in MRF genes. We detected four single nucleotide polymorphisms (SNPs) within exon 1 of the Myf5 gene, encoding the NH-terminal transactivation domain of the Myf5 protein. Three of these mutations change the amino acid sequence. The distribution of these SNPs was highly skewed in cattle populations; most of the mutations were found in only a few or even single individuals. Of the nine SNPs found in the promoter region of Myf5, one (transversion g.-723G-->T) was represented by all three genotypes distributed in the cattle populations studied. This polymorphism showed an influence on Myf5 gene expression in the longissimus dorsi muscle and was associated with sirloin weight and fat weight in sirloin in carcasses of Holstein-Friesian cattle.

Association of polymorphisms in exons 2 and 10 of the insulin-like growth factor 2 ( IGF2) gene with milk production traits in Polish Holstein-Friesian cattle.

Insulin-like growth factor 2 (IGF2) is considered to be a regulator of post-natal growth and differentiation of the mammary gland. In the present work, associations of two single nucleotide polymorphisms in the bovine IGF2 gene with milk production traits were studied in dairy Holstein-Friesian cows: the already described g.8656C>T transition in exon 2 (RFLP-BsrI) and the newly found g.24507G>T transversion in exon 10 (RFLP-HaeIII), found by sequencing 273-bp exon 10 of the IGF2 gene in six individuals. Associations were analysed individually and in combination with the multi-trait repeatability test-day animal model. The CT/GT haplotype appeared to be associated with most of the milk traits studied (differences were significant at P < or = 0.001). The most frequent CT/GG haplotype seemed inferior to others in fat and protein content and daily yield of fat and protein but superior (together with the TT/GG genotype) when the daily milk yield is considered.

Expression in promoter variant of the ERalpha gene in bos taurus liver.

Due to the variant functions that estrogens play in the regulation of reproduction, development of the mammary gland, growth and differentiation of cells, estrogen receptors and their genes are considered as a candidates for the markers of production and functional traits in farm animals, including cattle. In the earliest study, a 2853-bp bovine ER gene 5'-region was PCR amplified and sequenced. Moreover, for the first time, a polymorphism was described within 5' region of the bovine ERalpha gene--A/G transition lying upstream at position 2591 from acceptor splice site +85, possibly within its promoter--which could be recognized with RFLP-BglI. In other study we are found second polymorphism-A/G transition at position 1213 from acceptor splice site +85, located in promoter for exon B. We have examined the specific mRNA expression of ERalpha in various genotypes using real-time RT-PCR. We used four animals from each genotype group--AG, GG for BglI and AA, AG for SnaBI--to analyse liver ERalpha expression at the level of Real-time PCR. Liver samples were taken from the 16 young Friesian bulls of the different ERalpha genotypes, slaughtered at the local abattoir. As shown by Real-Time PCR, on the livers of animals with different genotype ERalpha mRNA for BglI polymorphism we didn't found variability, but for SnaBI we have found variability between AG and AA genotypes.

[Fetal activin A in labor complicated by meconium-stained amniotic fluid]. / Plodowa aktywina A w porodzie powiklanym obecnoscia smólki w plynie owodniowym.

OBJECTIVE: During pregnancy the placenta and the fetal membranes are the main sources of activin A. An increased level of activin A has been found in the serum of women with preeclampsia, diabetes mellitus and intrauterine growth restriction. Meconium is the predictor for adverse perinatal outcome, such as meconium aspiration syndrome or brain damage. The aim of our study was to evaluate the levels of fetal activin A in labors complicated by meconium-stained amniotic fluid. MATERIAL AND METHODS: Cord blood samples were collected from 65 full-term neonates from single pregnancies. In each case, the hematological parameters of cord blood and activin A (ELISA--Oxford Bio-Innovation Activin A Assay Kit) were assessed. RESULTS: There were no significant differences in the concentration of activin A in cord blood between the group with and the group without meconium-stained amniotic fluid. The mean count of nucleated erythrocytes and white blood cells as well as the percentage of reticulocytes was significantly higher in the meconium group. There were no significant differences between concentration of fetal activin A in a vaginal delivery (0.58 +/- 0.38 ng/ml) and cesarean section after labor (0.44 +/- 0.32 ng/ml) or elective cesarean section (0.62 +/- 0.47 ng/ml) groups. There were also no correlations between the levels of activin A and the parameters of fetal acid base status or cord blood hematological values. CONCLUSIONS: Fetal activin A has a limited significance for diagnosing fetal hypoxia in labors complicated by meconium-stained amniotic fluid. There were no correlations between the parameters of fetal acid base status and fetal activin A. The levels of fetal activin A do not depend on the mode of the delivery. Meconium-stained amniotic fluid resulted in significant changes of the hematological variables in cord blood.

Elevated tear fluid levels of MIP-1alpha in patients with cystic fibrosis.

Cystic fibrosis (CF) is the commonest multisystem genetic disease of white races, caused by mutations in the cystic fibrosis transmembrane regulator (CFTR), encoded on the long arm of chromosome 7. Mutations in the CFTR gene result in defective sodium, chloride, and water transport in the epithelial cells of the respiratory, hepatobiliary, gastrointestinal, and reproductive tracts, the pancreas, and the eye. The pathogenesis of ocular changes in CF is still unknown, but CF belongs to the large pathologic group of ocular surface epithelial diseases, termed keratoconjunctivitis sicca (KCS), that develop in dry eye syndrome. The aim of this study was to evaluate the levels of macrophage inflammatory protein-1alpha (MIP-1alpha) in the tear fluid of CF patients. We also investigated the correlation between the tear levels of this chemokine and clinical severity of CF and ocular surface disease. We studied 25 patients with CF with a mean age of 14 years. Chemokine levels were determined by ELISA. Complete ophthalmic examination, including dry eye tests, were used to study the ocular surface. The tear levels of MIP-1alpha in the CF patients were significantly higher when compared with healthy controls. We found a negative correlation between the tear levels of MIP-1alpha and clinical severity in CF patients and a positive correlation between the tear levels of MIP-1alpha and the presence of dry eye findings in CF patients. This current study indicates that chemokines play an important role in the ongoing inflammatory response. Our findings may help to explain one of the key factors contributing to the pathogenesis of ocular surface changes in CF patients.

[Concentration of interleukine-8 in tears in patients with cystic fibrosis]. / Stezenie interleukiny-8 w plynie lzowym u pacjentów z mukowiscydoza.

PURPOSE: To evaluate IL-8 concentration in tears fluid in cystic fibrosis patients. MATERIAL AND METHODS: Tears samples were collected from 18 CF Patients and 18 normal controls. Cytokine level was determined by ELISA. RESULTS: A significant increase of IL-8 concentration in tears fluid was found in CF patients compared with controls. CONCLUSIONS: Increase IL-8 concentration in tears fluid in CF patients suggests a role of immunologic processes in the pathogenesis of the ocular changes, and may be a marker of the inflammatory status in ocular surface in cystic fibrosis.

IL-8 and IFN-gamma in tear fluid of patients with cystic fibrosis.

Cystic fibrosis (CF) is inherited as an autosomal recessive disorder. It is caused by mutations in the protein-coding gene of chromosome 7, resulting in chronic pulmonary disease and pancreatic insufficiency. The disease affects all secretory epithelia, including the eye. The pathogenesis of ocular changes in CF is still unknown, but the involvement of immunologic processes in patients with CF has been studied in recent years. We measured interleukin-8 (IL-8) and interferon-gamma (IFN-gamma) levels in tears in a group of patients and a group of normal controls to determine if the levels of these cytokines are elevated in CF. The levels of these cytokines in tears and the clinical severity of CF and eye disease were compared. Tear samples were collected from 24 patients with CF at the department of pediatric diseases, Medical University of Bialystok, Poland. Cytokine levels were determined by ELISA. Ophthalmic examinations, including tests for keratoconjunctivitis sicca (dry eye), were used to study the ocular surface. The tear levels of IL-8 and IFN-gamma in the CF patients were significantly higher than those in controls. The clinical severity of CF correlated significantly with the IL-8 and IFN-gamma levels. We found positive correlation between the tear levels of IFN-gamma and dry eye findings in CF patients. Our results suggest that the inflammatory cytokines IL-8 and IFN-gamma may play key roles in the regulation of ocular surface inflammation and the immunologic reaction in patients with CF. The tear levels of IL-8 and IFN-gamma may be candidate markers for evaluation of the clinical status of CF and eye disease. These findings help to provide a new insight into the pathogenesis of dry eye in patients with CF and provide potential targets for therapy.

[Inflammatory mediators and hematological parameters in cord blood in labor complicated by meconium-stained amniotic fluid]. / Mediatory zapalenia oraz parametry hematologiczne krwi plodu w porodzie powiklanym obecnoscia smolki w plynie owodniowym.

OBJECTIVE: Meconium-stained amniotic fluid at term gestation is a predictor for adverse perinatal outcome and is associated with increased peripartum infections, independent of other risk factors. The aim of our study was to evaluate concentrations of inflammatory mediators (such as cytokine IL-6 and intracellular adhesion molecule ICAM-1) and values of hematological parameters of cord blood in presence or absence of meconium in amniotic fluid in term labor. MATERIAL AND METHODS: Cord blood samples were obtained from 66 term normal neonates immediately after birth, Soluble ICAM-1 and IL-6 concentrations were measured with ELISA R&D Systems kits. The umbilical blood specimen was analyzed using an automated hematology cell analyzer. Blood films were stained using May-Grünwald-Giemsa method. RESULTS: There were no difference in concentrations of ICAM-1 and IL-6 in cord blood in groups with or without meconium-stained amniotic fluid. The mean count of umbilical nucleated red blood cells and white blood cells was significantly higher in meconium group. There was no correlation between the cord blood hematological values and ICAM-1 or IL-6. There was also no correlation between IL-6 and ICAM-I and duration of labor. The mode of delivery influenced cord blood IL-6 levels. CONCLUSIONS: There was no influence of meconium-stained amniotic fluid on cord blood IL-6 and ICAM-1 levels. Changes in hematological parameters in cord blood in meconium passage can suggest either fetus hypoxia or infection. Significant differences of concentrations of fetal IL-6 were associated with the mode of delivery.

[IFN-gamma in tear fluid in patients with cystic fibrosis]. / Interferon-gamma w plynie lzowym u pacjentów z mukowiscydoza.

PURPOSE: To evaluate IFN-gamma concentration in tear fluid in CF patients. MATERIAL AND METHODS: Tear samples were collected from 15 CF patients at the age 10-21, and from 15 patients in control group at the age 11-20. Cytokine levels were determined by ELISA. RESULTS: The concentration of IFN-gamma in tear fluid in CF patients was 10.75+/-2.23 pg/ml, and 4.06+/-0.57 pg/ml in control group. CONCLUSIONS: The results of this study indicate that increased concentration of IFN-gamma in the tear fluid may be an important factor in the pathogenesis of dry eye in CF patients. They also open new perspectives related to therapeutic management.

Relationship of serum interleukin-7 concentration and the coagulation state in children with nephrotic syndrome.

BACKGROUND: Enhanced platelet reactivity may play a significant role in the hypercoagulable state of nephrotic syndrome (NS). Thrombocytosis with platelet aggregation cause the release of some cytokines, among them interleukin-7 (IL-7). The aim of the study was to evaluate serum IL-7 levels in children with the symptoms of NS and to determine a correlation between its concentration and platelet count, other hemostatic factors, and NS intensity. METHODS: The study was performed in two groups. I--the examined group of 26 children with NS (12 boys, 14 girls) aged 6.8 +/- 2.1 years, subjected to two examinations: A--before treatment, B--during treatment with prednisone (60 mg/kg 24 h after albuminuria regression); and II--the control group (C) of 20 healthy children. Serum IL-7 level was assayed by enzyme-linked immunosorbent assay method using a R & D Quantikine set. RESULTS: In group I, IL-7 level in examination A (33.33 +/- 33.24 pg/mL) was higher than in the control subjects (P < 0.01). In examination B, IL-7 concentration was reduced to the level of 10.86 +/- 5.22 pg/mL and did not differ from the controls (P > 0.05). A positive correlation was observed between IL-7 and platelet count and serum fibrinogen level. A negative correlation was noted with antithrombin III concentration. No correlation was found with serum levels of albumin and cholesterol or urine protein. CONCLUSION: In children with NS, serum IL-7 level increases proportionally to the elevated platelet count and other hemostatic components, but shows no correlation with serum albumin or urine protein.

[Evaluation of neutrophils adhesive molecules CD11b/CD18 in the course of photochemotherapy (PUVA) for psoriasis vulgaris]. / Ocena czasteczek adhezyjnych CD11b/CD18 granulocytów obojetnochlonnych w przebiegu fotochemoterapii (PUVA) luszczycy pospolitej.

Psoriasis is a common, chronic dermatological disorder. Inflammatory cells assemble in epidermis in the early stage of acute psoriasis. Accumulation of neutrophils at the site of inflammation depends on the expression of adhesion molecules. A variety of stimuli--mediators or selectins can active CD11b/CD18 molecules on the neutrophils surface. The aim of our study was expression of CD11b/CD18 molecules without and after stimulation by fMLP (formyl-Met-Leu-Phe) in patients with psoriasis vulgaris treated by PUVA. Expression of beta2-integrins was estimated with MFI (mean fluorescence intensity). Neutrophils in patient with acute psoriasis showed an increase in CD11b/CD18 expression. It means about an activation of neutrophils and particular role of those molecules in granulocyte adhesion to epidermis in psoriasis.

Evaluation of selected peripheral blood leukocyte functions in patients with various forms of periodontal disease.

INTRODUCTION: Periodontitis (P) is an infectious disease that develops in the supporting tissues of the tooth. One of the risk factors leading to it may be dysfunction of some immune system cells. Therefore, the object of the study was to assess selected functions of peripheral blood leukocytes in patients with various forms of P. As leukocytes are able to secrete interleukin (IL)-4 and IL-6, concentrations of their soluble receptors and the expression of their membrane receptors were investigated. MATERIAL AND METHODS: Twenty generally healthy subjects with aggressive (AP)and chronic periodontitis (CP)were enrolled in the study. The control group consisted of 8 healthy subjects,with no changes in periodontium. Peripheral blood mononuclear cells (PBMCs) were isolated and cultured. Levels of IL-4,IL-6,and their soluble receptors sIL-4R and sIL-6R were determined in the supernatant by ELISA. The expressions of cell surface IL-4R and IL-6R were assayed on PBMC using flow cytometry. RESULTS: No statistically significant differences were found in the selected parameters between people with periodontal disease and healthy controls. However, in subjects with AP, there was an increasing tendency in IL-6 concentration and IL-4R expression on PBMCs. CONCLUSIONS: Our results show that leukocytes play a significant part in P and their activity is probably lesion-dependent. Estimation of the cytokines secreted by leukocytes may facilitate differentiation and prognosis of the disease progression.