Search results for: serum catalase

Authors: Gaweł Magdalena, Lipkowska Anna, Olbert Magdalena, Frąckiewicz Ewelina, Librowski Tadeusz, Nowak Gabriel, Pilc Andrzej

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Heme oxygenase-1 (HO-1), also known as heat shock protein 32 (HSP32), has been shown to be implicated in cytoprotection in various organs. Its activation plays a significant role in acute and chronic inflammation, protecting cells from oxidative injury and apoptosis. This inducible isoform of HO catalyzes the first and rate-limiting step in heme degradation to produce equimolar quantities of biologically active products: carbon monoxide (CO), free iron and biliverdin. CO has been reported to possess anti-apoptotic properties. Moreover, it inhibits the production of proinflammatory cytokines and stimulates the synthesis of the anti-inflammatory interleukin-10 (IL-10), as well as promotes vasodilatation at sites of inflammation. The second product of catalytic HO-1 activity, free cytotoxic iron, is promptly sequestered into the iron storage protein ferritin, which lowers the pro-oxidant state of the cell. The third product, biliverdin, is subsequently converted by biliverdin reductase into the bile pigment bilirubin, the most potent endogenous antioxidant among the constituents of human serum, which modulates immune effector functions and suppresses inflammatory response. Furthermore, being one of the so-called stress proteins, HO-1 adaptively responds to different stressors, such as reactive oxygen species (ROS), inflammatory cytokines and heavy metals and thus protects cells against such conditions as ischemia, hemorrhagic shock, heat shock or hypoxia. It is suggested that pharmacologic modulation of HO-1 may represent an effective strategy for prevention of stress and drug-induced gastrointestinal toxicity. HO-1 is constitutively expressed in normal gastric, intestinal and colonic mucosa and up-regulated during inflammation. It has been proven that HO-1 up-regulated by hemin, heme and cobalt-protoporphyrin ameliorates experimental colitis. In addition, the up-regulation of HO-1 partially explains the mechanism of action of 5-aminosalicylic acid (5-ASA), which is used clinically as an anti-colitis agent. In 2009 Ueda et al. has reported for the first time that mucosal protection by Polaprezinc, a chelate compound of zinc and L-carnosine used as an anti-ulcer drug in Japan, is also attributed to induction of HO-1 in the stomach. Since then, inducers of HO-1 are desired subject of research, as they may constitute therapeutically effective anti-ulcer drugs.

Keywords: heme oxygenase-1, gastric lesions, gastroprotection, Polaprezinc

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Authors: Farah Diab, Mohamad Khalil, Francesca Storace, Francesca Baldini, Piero Portincasaa, Giulio Lupidi, Laura Vergani

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Thymbra spicata is a thyme-like plant belonging to the Lamiaceae family that shows a global distribution, especially in the eastern Mediterranean region. Leaves of T. spicata contain large amounts of phenols such as phenolic acids (rosmarinic acid), phenolic monoterpenes (carvacrol), and flavonoids. In Lebanon, T. spicata is currently used as a culinary herb in salad and infusion, as well as for traditional medicinal purposes. Carvacrol (5-isopropyl-2-methyl phenol), the most abundant polyphenol in the organic extract and essential oils, has a great array of pharmacological properties. In fact, carvacrol is largely employed as a food additive and neutraceutical agent. Our aim is to investigate the beneficial effects of T. spicata ethanolic extract (TE) and its main component, carvacrol, using in vitro models of hepatic steatosis and endothelial dysfunction. As a further point, we focused on investigating if and how the binding of carvacrol to albumin, the physiological transporter for drugs in the blood, might be altered by the presence of high levels of fatty acids (FAs), thus impairing the carvacrol bio-distribution in vivo. For that reason, hepatic FaO cells treated with exogenous FAs such as oleate and palmitate mimic hepatosteatosis; endothelial HECV cells exposed to hydrogen peroxide are a model of endothelial dysfunction. In these models, we measured lipid accumulation, free radical production, lipoperoxidation, and nitric oxide release before and after treatment with carvacrol. The carvacrol binding to albumin with/without high levels of long-chain FAs was assessed by absorption and emission spectroscopies. Our findings show that both TE and carvacrol (i) counteracted lipid accumulation in hepatocytes by decreasing the intracellular and extracellular lipid contents in steatotic FaO cells; (ii) decreased oxidative stress in endothelial cells by significantly reducing lipoperoxidation and free radical production, as well as, attenuating the nitric oxide release; (ii) high levels of circulating FAs reduced the binding of carvacrol to albumin. The beneficial effects of TE and carvacrol on both hepatic and endothelial cells point to a nutraceutical potential. However, high levels of circulating FAs, such as those occurring in metabolic disorders, might hinder the carvacrol transport, bio-distribution, and pharmacodynamics.

Keywords: carvacrol, endothelial dysfunction, fatty acids, non-alcoholic fatty liver diseases, serum albumin

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Authors: Francisco J. Cedano, Laura M. Pinzón, Camila I. Castro, Felipe Salcedo, Juan P. Casas, Juan C. Briceño

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Complex fractures located in articular surfaces are challenging to treat and their reduction with conventional treatments could compromise the functionality of the affected limb. An adhesive material to treat those fractures is desirable for orthopedic surgeons. This adhesive must be biocompatible and have a high adhesion to bone surface in an aqueous environment. The proposed adhesive is based on chitosan, given its adhesive and biocompatibility properties. Chitosan is mixed with calcium carbonate and hydroxyapatite, which contribute to structural support and a gel like behavior, and glutaraldehyde is used as a cross-linking agent to keep the adhesive mechanical performance in aqueous environment. This work aims to evaluate the rheological, adhesion strength and biocompatibility properties of the proposed adhesive using in vitro tests. The gelification process of the adhesive was monitored by oscillatory rheometry in an ARG-2 TA Instruments rheometer, using a parallel plate geometry of 22 mm and a gap of 1 mm. Time sweep experiments were conducted at 1 Hz frequency, 1% strain and 37°C from 0 to 2400 s. Adhesion strength is measured using a butt joint test with bovine cancellous bone fragments as substrates. The test is conducted at 5 min, 20min and 24 hours after curing the adhesive under water at 37°C. Biocompatibility is evaluated by a cytotoxicity test in a fibroblast cell culture using MTT assay and SEM. Rheological results concluded that the average gelification time of the adhesive is 820±107 s, also it reaches storage modulus magnitudes up to 106 Pa; The adhesive show solid-like behavior. Butt joint test showed 28.6 ± 9.2 kPa of tensile bond strength for the adhesive cured for 24 hours. Also there was no significant difference in adhesion strength between 20 minutes and 24 hours. MTT showed 70 ± 23 % of active cells at sixth day of culture, this percentage is estimated respect to a positive control (only cells with culture medium and bovine serum). High vacuum SEM observation permitted to localize and study the morphology of fibroblasts presented in the adhesive. All captured fibroblasts presented in SEM typical flatted structure with filopodia growth attached to adhesive surface. This project reports an adhesive based on chitosan that is biocompatible due to high active cells presented in MTT test and these results were correlated using SEM. Also, it has adhesion properties in conditions that model the clinical application, and the adhesion strength do not decrease between 5 minutes and 24 hours.

Keywords: bioadhesive, bone adhesive, calcium carbonate, chitosan, hydroxyapatite, glutaraldehyde

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Authors: Oscar Rubiano, Oscar Ortiz, Natalia Morales, Lida Alfonso, Johana Alvarado, Adriana Gutierrez, Daniel Botero

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Athletes who usually use protein supplements, are those who practice strength and power sports, whose goal is to achieve a large muscle mass. However, it has also been explored in sports or endurance activities such as cycling, and where despite requiring high power, prominent muscle development can impede good competitive performance due to the determinant of body mass for good performance of the athlete body. This research shows, the effect with protein supplements establishes a protein - muscle mass ratio, although in a lesser proportion the relationship between protein types and muscle power. Thus, we intend to explore as a first approximation, the behavior of muscle power in lower limbs after the intake of two protein supplements from different sources. The aim of the study was to describe the behavior of muscle power in lower limbs after the consumption of animal protein (AP) and vegetable protein (VP) in four route cyclists from 18 to 20 years of the Bogota cycling league. The methodological design of this study is quantitative, with a non-probabilistic sampling, based on a pre-experimental model. The jumping power was evaluated before and after the intervention by means of the squat jump test (SJ), Counter movement jump (CMJ) and Abalacov (AB). Cyclists consumed a drink with whey protein and a soy isolate after training four times a week for three months. The amount of protein in each cyclist, was calculated according to body weight (0.5 g / kg of muscle mass). The results show that subjects who consumed PV improved muscle strength and landing strength. In contrast, the power and landing force decreased for subjects who consumed PA. For the group that consumed PV, the increase was positive at 164.26 watts, 135.70 watts and 33.96 watts for the AB, SJ and CMJ jumps respectively. While for PA, the differences of the medians were negative at -32.29 watts, -82.79 watts and -143.86 watts for the AB, SJ and CMJ jumps respectively. The differences of the medians in the AB jump were positive for both the PV (121.61 Newton) and PA (454.34 Newton) cases, however, the difference was greater for PA. For the SJ jump, the difference for the PA cases was 371.52 Newton, while for the PV cases the difference was negative -448.56 Newton, so the difference was greater in the SJ jump for PA. In jump CMJ, the differences of the medians were negative for the cases of PA and PV, being -7.05 for PA and - 958.2 for PV. So the difference was greater for PA. The conclusion of this study shows that serum protein supplementation showed no improvement in muscle power in the lower limbs of the cyclists studied, which could suggest that whey protein does not have a beneficial effect on performance in terms of power, either, showed an impact on body composition. In contrast, supplementation with soy isolate showed positive effects on muscle power, body.

Keywords: animal protein (AP), muscle power, supplements, vegetable protein (VP)

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Authors: Soniya Nityanand, Ekta Minocha, Manali Jain, Rohit Anthony Sinha, Chandra Prakash Chaturvedi

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Amniotic fluid stem cells (AFSC) have been shown to contribute towards the amelioration of Acute Renal Failure (ARF), but the mechanisms underlying the renoprotective effect are largely unknown. Therefore, the main goal of the current study was to evaluate the therapeutic efficacy of AFSC in a cisplatin-induced rat model of ARF and to investigate the underlying mechanisms responsible for its renoprotective effect. To study the therapeutic efficacy of AFSC, ARF was induced in Wistar rats by an intra-peritoneal injection of cisplatin, and five days after administration, the rats were randomized into two groups and injected with either AFSC or normal saline intravenously. On day 8 and 12 after cisplatin injection, i.e., day 3 and day7 post-therapy respectively, the blood biochemical parameters, histopathological changes, apoptosis and expression of pro-apoptotic, anti-apoptotic and autophagy-related proteins in renal tissues were studied in both groups of rats. Administration of AFSC in ARF rats resulted in improvement of renal function and attenuation of renal damage as reflected by significant decrease in blood urea nitrogen, serum creatinine levels, tubular cell apoptosis as assessed by Bax/Bcl2 ratio, and expression of the pro-apoptotic proteins viz. PUMA, Bax, cleaved caspase-3 and cleaved caspase-9 as compared to saline-treated group. Furthermore, in the AFSC-treated group as compared to saline-treated group, there was a significant increase in the activation of autophagy as evident by increased expression of LC3-II, ATG5, ATG7, Beclin1 and phospho-AMPK levels with a concomitant decrease in phospho-p70S6K and p62 expression levels. To further confirm whether the protective effects of AFSC on cisplatin-induced apoptosis were dependent on autophagy, chloroquine, an autophagy inhibitor was administered by the intra-peritoneal route. Chloroquine administration led to significant reduction in the anti-apoptotic effects of the AFSC therapy and further deterioration in the renal structure and function caused by cisplatin. Collectively, our results put forth that AFSC ameliorates cisplatin-induced ARF through induction of autophagy and inhibition of apoptosis. Furthermore, the protective effects of AFSC were blunted by chloroquine, highlighting that activation of autophagy is an important mechanism of action for the protective role of AFSC in cisplatin-induced renal injury.

Keywords: amniotic fluid stem cells, acute renal failure, autophagy, cisplatin

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Authors: Mohit Kumar, Beklashwar Salona, Shiv Murti, Mukesh Singh

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We report here a case of five-months old male baby, born as second child of non-consanguineous parents with no considerable history of genetic abnormality which was referred to our cytogenetic laboratory for chromosomal analysis. Physical dysmorphic facial features including mongoloid face, cleft palate, simian crease, and developmental delay were observed. We present this case with unique balanced autosomal translocation of t(3;10)(p21;p13). The risk of phenotypic abnormalities based on de novo balanced translocation was estimated to be 7%. The association of balanced chromosomal rearrangement with Down syndrome features such as multiple congenital anomalies, facial dysmorphism and congenital heart anomalies are very rare in a 5-months old male child. Trisomy-21 is not uncommon in chromosomal abnormality with the birth defect and balanced translocations are frequently observed in patients with secondary infertility or recurrent spontaneous abortion (RSA). Two ml heparinized peripheral blood cells cultured in RPMI-1640 for 72 hours supplemented with 20% fetal bovine serum, phytohemagglutinin (PHA), and antibiotics were used for chromosomal analysis. A total 30 metaphases images were captured using Olympus-BX51 microscope and analyzed using Bio-view karyotyping software through GTG-banding (G bands by trypsin and Giemsa) according to International System for Human Cytogenetic Nomenclature 2016. The results showed balanced translocation between short arm of chromosome # 3 and short arm of chromosome # 10. The karyotype of the child was found to be 46,XY,t(3;10)(p21; p13). Chromosomal abnormalities are one of the major causes of birth defect in new born babies. Also, balanced translocations are frequently observed in patients with secondary infertility or recurrent spontaneous abortion. The index case presented with dysmorphic facial features and had a balanced translocation 46,XY,t(3;10)(p21;p13). This translocation with break points at (p21; p13) has not been reported in the literature in a child with facial dysmorphism. To the best of our knowledge, this is the first report of novel balanced translocation t(3;10) with break points in a child with dysmorphic features. We found balanced chromosomal translocation instead of any trisomy or unbalanced aberrations along with some phenotypic abnormalities. Therefore, we suggest that such novel balanced translocation with abnormal phenotype should be reported in order to enable the pathologist, pediatrician, and gynecologist to have a better insight into the intricacies of chromosomal abnormalities and their associated phenotypic features. We hypothesized that dysmorphic features as seen in this case may be the result of change in the pattern of genes located at the breakpoint area in balanced translocations or may be due to deletion or mutation of genes located on the p-arm of chromosome # 3 and p-arm of chromosome # 10.

Keywords: balanced translocation, karyotyping, phenotypic abnormalities, facial dimorphisms

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Authors: Tamar Grdzelishvili, Zaza Sinauridze

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Introduction: Preeclampsia is a complication of the second trimester of pregnancy, which is characterized by high morbidity and multiorgan damage. Many complex pathogenic mechanisms are now implicated to be responsible for this disease (1). Preeclampsia is one of the leading causes of maternal mortality worldwide. Statistics are enough to convince you of the seriousness of this pathology: about 100,000 women die of preeclampsia every year. It occurs in 3-14% (varies significantly depending on racial origin or ethnicity and geographical region) of pregnant women, in 75% of cases - in a mild form, and in 25% - in a severe form. During severe pre-eclampsia-eclampsia, perinatal mortality increases by 5 times and stillbirth by 9.6 times. Considering that the only way to treat the disease is to end the pregnancy, the main thing is timely diagnosis and prevention of the disease. Identification of high-risk pregnant women for PE and giving prophylaxis would reduce the incidence of preterm PE. First-trimester screening model developed by the Fetal Medicine Foundation (FMF), which uses the Bayes-theorem to combine maternal characteristics and medical history together with measurements of mean arterial pressure, uterine artery pulsatility index, and serum placental growth factor, has been proven to be effective and have superior screening performance to that of traditional risk factor-based approach for the prediction of PE (2) Methods: Retrospective single center screening study. The study population consisted of women from the Tbilisi maternity hospital “Pineo medical ecosystem” who met the following criteria: they spoke Georgian, English, or Russian and agreed to participate in the study after discussing informed consent and answering questions. Prior to the study, the informed consent forms approved by the Institutional Review Board were obtained from the study subjects. Early assessment of preeclampsia was performed between 11-13 weeks of pregnancy. The following were evaluated: anamnesis, dopplerography of the uterine artery, mean arterial blood pressure, and biochemical parameter: Pregnancy-associated plasma protein A (PAPP-A). Individual risk assessment was performed with performed by Fast Screen 3.0 software ThermoFisher scientific. Results: A total of 513 women were recruited and through the study, 51 women were diagnosed with preeclampsia (34.5% in the pregnant women with high risk, 6.5% in the pregnant women with low risk; P<0.000 1). Conclusions: First-trimester screening combining maternal factors with uterine artery Doppler, blood pressure, and pregnancy-associated plasma protein-A is useful to predict PE in a routine care setting. More patient studies are needed for final conclusions. The research is still ongoing.

Keywords: first-trimester, preeclampsia, screening, pregnancy-associated plasma protein

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Authors: Safaa S. Hassan, Mohammed H. Elbakry, Safwat A. Mangoura, Zainab M. Omar

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Background: Liver fibrosis is the main reason for increased mortality in chronic liver disease. It has no standard treatment. Antioxidants from a variety of sources are capable of slowing or preventing oxidation of other molecules. Aim: to evaluate the hepatoprotective effect of vit. C, vit. E and gallic acid in comparison to silymarin in the rat model of carbon tetrachloride induced liver fibrosis and their possible mechanisms of action. Material& Methods: A total number of 60 adult male albino rats 160-200gm were divided into six equal groups; received subcutaneous (s.c) injection for 8 weeks. Group I: as control. Group II: received 1.5 mL/kg of CCL4 .Group III: CCL4 and co- treatment with silymarin 100mg/kg p.o. daily. Group IV: CCL4 and co-treatment with vit. C 50mg/kg p.o. daily. Group V: CCL4 and co-treatment with vit. E 200mg/kg. p.o. Group VI: CCL4 and co-treatment with Gallic acid 100mg/kg. p.o. daily. Liver was processed for histological and immunohistochemical examination. Levels of AST, ALT, ALP, reduced GSH, MDA, SOD and hydroxyproline concentration were measured and evaluated statistically. Results: Light and electron microscopic examination of liver of group II exhibited foci of altered cells with dense nuclei and vacuolated, granular cytoplasm, mononuclear cell infiltration in portal areas, profuse collagen fiber deposits were found around portal tract, more intense staining α-SMA-positive cells occupied most of the liver fibrosis tissue, electron lucent areas in the cytoplasm of the hepatocytes, margination of nuclear chromatin. Treatment by any of the antioxidants variably reduced the hepatic structural changes induced by CCL4. Biochemical analysis showed that carbon tetrachloride significantly increased the levels of serum AST, ALT, ALP, hepatic malondialdehyde and hydroxyproline content. Moreover, it decreased the activities of superoxide dismutase and glutathione. Treatment with silymarin, gallic acid, vit. C and vit. E decreased significantly the AST, ALT, and ALP levels in plasma, MDA and hydroxyproline and increased the activities of SOD and glutathione in liver tissue. The effect of administration of CCl4 was improved with the used antioxidants in variable degrees. The most efficient antioxidant was silymarin followed by gallic acid and vit. C then vit. E. It is possibly due to their antioxidant effect, free radical scavenging properties and the reduction of oxidant dependent activation and proliferation of HSCs. Conclusion: So these antioxidants can be a promising drugs candidate for ameliorating liver fibrosis better than the use of the drugs and their side effects.

Keywords: antioxidant, ccl4, gallic acid, liver fibrosis

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Authors: Madelyn N. Moawad, Hermine R. Z. Tadros, Mary G. Ghobrial, Ahmad R. Bassiouny, Kamal M. Kandeel, Athar Ata

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Seaweeds are potential renewable resources in marine environment. They provide an excellent source of bioactive substances such as dietary fibers and various functional polysaccharides that could potentially be used as ingredients for both human and animal health applications. The observations suggested that these bioactive compounds have strong antioxidant properties, which have beneficial effects on human health. The present research aimed at finding new chemical products from local marine macroalgae for natural medicinal uses and consumption for their nutritional values. Macroalgae samples were collected manually mainly from the Mediterranean Sea at shallow subtidal zone of Abu Qir Bay, Alexandria, Egypt. The chemical compositions of lyophilized materials of six selected macroalgal species; Colpomenia sinuosa, Sargassum linifolium, Padina pavonia, Pterocladiella capillacea, Laurencia pinnatifidia, and Caulerpa racemosa, were investigated for proteins using bovine serum albumin, and carbohydrates were assayed by phenol-sulfuric acid reaction. The macroalgae lipid was extracted with chloroform, methanol and phosphate buffer. Vitamins were extracted using trichloroacetic acid. Chlorophylls and total carotenoids were determined spectrophotometrically and total phenols were extracted with methanol. In addition, lipid-soluble, and water-soluble antioxidant, and anti α-glucosidase activities were measured spectrophotometrically. The antioxidant activity of hexane extracts was investigated using phosphomolybdenum reagent. The anti-α-glucosidase effect measurement was initiated by mixing α-glucosidase solution with p-nitrophenyl α-D-glucopyranoside. The results showed that the ash contents varied from 11.2 to 35.4 % on dry weight basis for P. capillacea and Laurencia pinnatifidia, respectively. The protein contents ranged from 5.63 % in brown macroalgae C. sinuosa to 8.73 % in P. pavonia. A relative wide range in carbohydrate contents was observed (20.06–46.75 %) for the test algal species. The highest lipid percentage was found in green alga C. racemosa (5.91%) followed by brown algae P. pavonia (3.57%) and C. sinuosa (2.64%). The phenolic contents varied from 1.32 mg GAE/g for C. sinuosa to 4.00 mg GAE/g in P. pavonia. The lipid-soluble compounds exhibited higher antioxidant capacity (73.18-145.95 µM/g) than that of the water-soluble ones ranging from 24.83 µM/g in C. racemosa to 74.07 µM/g in S. linifolium. The most potent anti-α-glucosidase activity was observed for P. pavonia with IC50 of 17.12 μg/ml followed by S. linifolium (IC50 = 71.75 μg/ml), C. racemosa (IC50 = 84.73 μg/ml), P. capillacea (IC50 = 92.16 μg/ml), C. sinuosa (IC50 = 112.44 μg/ml), and L. pinnatifida (IC50 = 115.11 μg/ml).

Keywords: α-glucosidase, lyophilized, macroalgae, spectrophotometrically

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Authors: Neha Batra Bali, Monika Tomar, Vinay Gupta

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A reagentless biosensor for detection of urea based on ZnO-CuO composite thin film is presented in following work. Biosensors have immense potential for varied applications ranging from environmental to clinical testing, health care, and cell analysis. Immense growth in the field of biosensors is due to the huge requirement in today’s world to develop techniques which are both cost effective and accurate for prevention of disease manifestation. The human body comprises of numerous biomolecules which in their optimum levels are essential for functioning. However mismanaged levels of these biomolecules result in major health issues. Urea is one of the key biomolecules of interest. Its estimation is of paramount significance not only for healthcare sector but also from environmental perspectives. If level of urea in human blood/serum is abnormal, i.e., above or below physiological range (15-40mg/dl)), it may lead to diseases like renal failure, hepatic failure, nephritic syndrome, cachexia, urinary tract obstruction, dehydration, shock, burns and gastrointestinal, etc. Various metal nanoparticles, conducting polymer, metal oxide thin films, etc. have been exploited to act as matrix to immobilize urease to fabricate urea biosensor. Amongst them, Zinc Oxide (ZnO), a semiconductor metal oxide with a wide band gap is of immense interest as an efficient matrix in biosensors by virtue of its natural abundance, biocompatibility, good electron communication feature and high isoelectric point (9.5). In spite of being such an attractive candidate, ZnO does not possess a redox couple of its own which necessitates the use of electroactive mediators for electron transfer between the enzyme and the electrode, thereby causing hindrance in realization of integrated and implantable biosensor. In the present work, an effort has been made to fabricate a matrix based on ZnO-CuO composite prepared by pulsed laser deposition (PLD) technique in order to incorporate redox properties in ZnO matrix and to utilize the same for reagentless biosensing applications. The prepared bioelectrode Urs/(ZnO-CuO)/ITO/glass exhibits high sensitivity (70µAmM⁻¹cm⁻²) for detection of urea (5-200 mg/dl) with high stability (shelf life ˃ 10 weeks) and good selectivity (interference ˂ 4%). The enhanced sensing response obtained for composite matrix is attributed to the efficient electron exchange between ZnO-CuO matrix and immobilized enzymes, and subsequently fast transfer of generated electrons to the electrode via matrix. The response is encouraging for fabricating reagentless urea biosensor based on ZnO-CuO matrix.

Keywords: biosensor, reagentless, urea, ZnO-CuO composite

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Authors: Honey Bryant, Elvina Chu

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Stroke is the leading cause of disability in adults in Canada and directly related to depression, anxiety, and sleep disorders; with an estimated annual cost of $50 billion in health care. Strokes not only impact the individual but society as a whole. Current stroke rehabilitation does not include Music Therapy, although it has success in clinical research in the use of stroke rehabilitation. This study examines the use of neurologic music therapy (NMT) in conjunction with stroke rehabilitation to improve sleep quality, reduce stress levels, and promote neurogenesis. Existing research on NMT in stroke is limited, which means any conclusive information gathered during this study will be significant. My novel hypotheses are a.) stroke patients will become less depressed and less anxious with improved sleep following NMT. b.) NMT will reduce stress levels and promote neurogenesis in stroke patients admitted for rehabilitation. c.) Beneficial effects of NMT will be sustained at least short-term following treatment. Participants were recruited from the in-patient stroke rehabilitation program at Providence Care Hospital in Kingston, Ontario, Canada. All participants-maintained stroke rehabilitation treatment as normal. The study was spilt into two groups, the first being Passive Music Listening (PML) and the second Neurologic Music Therapy (NMT). Each group underwent 10 sessions of intensive music therapy lasting 45 minutes for 10 consecutive days, excluding weekends. Psychiatric Assessments, Epworth Sleepiness Scale (ESS), Hospital Anxiety & Depression Rating Scale (HADS), and Music Engagement Questionnaire (MusEQ), were completed, followed by a general feedback interview. Physiological markers of stress were measured through blood pressure measurements and heart rate variability. Serum collections reviewed neurogenesis via Brain-derived neurotrophic factor (BDNF) and stress markers of cortisol levels. As this study is still on-going, a formal analysis of data has not been fully completed, although trends are following our hypotheses. A decrease in sleepiness and anxiety is seen upon the first cohort of PML. Feedback interviews have indicated most participants subjectively felt more relaxed and thought PML was useful in their recovery. If the hypothesis is supported, larger external funding which will allow for greater investigation of the use of NMT in stroke rehabilitation. As we know, NMT is not covered under Ontario Health Insurance Plan (OHIP), so there is limited scientific data surrounding its uses as a clinical tool. This research will provide detailed findings of the treatment of neuropsychiatric aspects of stroke. Concurrently, a passive music listening study is being designed to further review the use of PML in rehabilitation as well.

Keywords: music therapy, psychotherapy, neurologic music therapy, passive music listening, neuropsychiatry, counselling, behavioural, stroke, stroke rehabilitation, rehabilitation, neuroscience

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Authors: Clara Tramuta, Floris Irene, Daniela Manila Bianchi, Monica Pitti, Giulia Federica Cazzaniga, Lucia Decastelli

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This work presents the results obtained by the Regional Reference Centre for Salmonella Typing (CeRTiS) in a retrospective study aimed to investigate, through Pulsed-field Gel Electrophoresis (PFGE) analysis, the genetic relatedness of emerging Salmonella serotypes of human origin circulating in North-West of Italy. Furthermore, the goal of this work was to create a Regional database to facilitate foodborne outbreak investigation and to monitor them at an earlier stage. A total of 112 strains, isolated from 2016 to 2018 in hospital laboratories, were included in this study. The isolates were previously identified as Salmonella according to standard microbiological techniques and serotyping was performed according to ISO 6579-3 and the Kaufmann-White scheme using O and H antisera (Statens Serum Institut®). All strains were characterized by PFGE: analysis was conducted according to a standardized PulseNet protocol. The restriction enzyme XbaI was used to generate several distinguishable genomic fragments on the agarose gel. PFGE was performed on a CHEF Mapper system, separating large fragments and generating comparable genetic patterns. The agarose gel was then stained with GelRed® and photographed under ultraviolet transillumination. The PFGE patterns obtained from the 112 strains were compared using Bionumerics version 7.6 software with the Dice coefficient with 2% band tolerance and 2% optimization. For each serotype, the data obtained with the PFGE were compared according to the geographical origin and the year in which they were isolated. Salmonella strains were identified as follow: S. Derby n. 34; S. Infantis n. 38; S. Napoli n. 40. All the isolates had appreciable restricted digestion patterns ranging from approximately 40 to 1100 kb. In general, a fairly heterogeneous distribution of pulsotypes has emerged in the different provinces. Cluster analysis indicated high genetic similarity (≥ 83%) among strains of S. Derby (n. 30; 88%), S. Infantis (n. 36; 95%) and S. Napoli (n. 38; 95%) circulating in north-western Italy. The study underlines the genomic similarities shared by the emerging Salmonella strains in Northwest Italy and allowed to create a database to detect outbreaks in an early stage. Therefore, the results confirmed that PFGE is a powerful and discriminatory tool to investigate the genetic relationships among strains in order to monitoring and control Salmonellosis outbreak spread. Pulsed-field gel electrophoresis (PFGE) still represents one of the most suitable approaches to characterize strains, in particular for the laboratories for which NGS techniques are not available.

Keywords: emerging Salmonella serotypes, genetic characterization, human strains, PFGE

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Authors: Takaaki Nemoto

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Case presentation: A 69-year-old Japanese man presented with a low-grade fever and fatigue that had persisted for one month. The patient had an aortic dissection on the aortic arch 13 years prior, an abdominal aortic aneurysm seven years prior, and an aortic dissection on the distal aortic arch one year prior, which were all treated with artificial blood-vessel replacement surgery. Laboratory tests revealed an inflammatory response (CRP 7.61 mg/dl), high serum creatinine (Cr 1.4 mg/dL), and elevated transaminase (AST 47 IU/L, ALT 45 IU/L). The patient was admitted to our hospital on suspicion of prosthetic vascular graft infection. Following further workups on the inflammatory response, an enhanced chest computed tomography (CT) and a non-enhanced chest DWI (MRI) were performed. The patient was diagnosed with a pulmonary fistula and a prosthetic vascular graft infection on the distal aortic arch. After admission, the patient was administered Ceftriaxion and Vancomycine for 10 days, but his fever and inflammatory response did not improve. On day 13 of hospitalization, a lung fistula repair surgery and an omental filling operation were performed, and Meropenem and Vancomycine were administered. The fever and inflammatory response continued, and therefore we took repeated blood cultures. M. fortuitum was detected in a blood culture on day 16 of hospitalization. As a result, we changed the treatment regimen to Amikacin (400 mg/day), Meropenem (2 g/day), and Cefmetazole (4 g/day), and the fever and inflammatory response began to decrease gradually. We performed a test of sensitivity for Mycobacterium fortuitum, and found that the MIC was low for fluoroquinolone antibacterial agent. The clinical course was good, and the patient was discharged after a total of 8 weeks of intravenous drug administration. At discharge, we changed the treatment regimen to Levofloxacin (500 mg/day) and Clarithromycin (800 mg/day), and prescribed these two drugs as a long life suppressive therapy. Discussion: There are few cases of prosthetic vascular graft infection caused by mycobacteria, and a standard therapy remains to be established. For prosthetic vascular graft infections, it is ideal to provide surgical and medical treatment in parallel, but in this case, surgical treatment was difficult and, therefore, a conservative treatment was chosen. We attempted to increase the treatment success rate of this refractory disease by conducting a susceptibility test for mycobacteria and treating with different combinations of antimicrobial agents, which was ultimately effective. With our treatment approach, a good clinical course was obtained and continues at the present stage. Conclusion: Although prosthetic vascular graft infection resulting from mycobacteria is a refractory infectious disease, it may be curative to administer appropriate antibiotics based on the susceptibility test in addition to surgical treatment.

Keywords: prosthetic vascular graft infection, lung fistula, Mycobacterium fortuitum, conservative treatment

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Authors: Lívia Pinheiro Carvalho, Luciana Di Thommazo-Luporini, Rafael Luís Luporini, José Carlos Bonjorno Junior, Renata Pedrolongo Basso Vanelli, Manoel Carneiro de Oliveira Junior, Rodolfo de Paula Vieira, Renata Trimer, Renata G. Mendes, Mylène Aubertin-Leheudre, Audrey Borghi-Silva

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Rationale: Cardiorespiratory fitness (CRF) and functional capacity in young obese and normal-weight people are associated with metabolic and cardiovascular diseases and mortality. However, it remains unclear whether their metabolically healthy (MH) or at risk (AR) phenotype influences cardiorespiratory fitness in this vulnerable population such as obese adults but also in normal-weight people. HOMA insulin resistance index (HI) and leptin-adiponectin ratio (LA) are strong markers for characterizing those phenotypes that we hypothesized to be associated with physical fitness. We also hypothesized that an easy and feasible exercise test could identify a subpopulation at risk to develop metabolic and related disorders. Methods: Thirty-nine sedentary men and women (20-45y; 18.530 kg.m-2) underwent a clinical evaluation, including the six-minute step test (ST), a well-validated and reliable test for young people. Body composition assessment was done by a tetrapolar bioimpedance in a fasting state and in the folicular phase for women. A maximal cardiopulmonary exercise testing, as well as the ST, evaluated the oxygen uptake at the peak of the test (VO2peak) by an ergospirometer Oxycon Mobile. Lipids, glucose, insulin were analysed and the ELISA method quantified the serum leptin and adiponectin from blood samples. Volunteers were divided in two groups: AR or MH according to a HI cutoff of 1.95, which was previously determined in the literature. T-test for comparison between groups, Pearson´s test to correlate main variables and ROC analysis for discriminating AR from up-and-down cycles in ST (SC) were applied (p<0.05). Results: Higher LA, fat mass (FM) and lower HDL, SC, leg lean mass (LM) and VO2peak were found in AR than in MH. Significant correlations were found between VO2peak and SC (r= 0.80) as well as between LA and FM (r=0.87), VO2peak (r=-0.73), and SC (r=-0.65). Area under de curve showed moderate accuracy (0.75) of SC <173 to discriminate AR phenotype. Conclusion: Our study found that at risk obese and normal-weight subjects showed an unhealthy metabolism as well as a poor CRF and functional daily activity capacity. Additionally, a simple and less costly functional test associated with above-mentioned aspects is able to identify ‘at risk’ subjects for primary intervention with important clinical and health implications.

Keywords: aerobic capacity, exercise, fitness, metabolism, obesity, 6MST

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Authors: Mustafa M. Donma, Orkide Donma

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Obesity is known to be associated with many clinically important diseases including metabolic syndrome (MetS). Vitamin B₁₂ plays essential roles in fat and protein metabolisms and its cooperation with vitamin B₉ is well-known. The aim of this study is to investigate the possible contributions as well as associations of these micronutrients upon obesity and MetS during childhood. A total of 128 children admitted to Namik Kemal University, Medical Faculty, Department of Pediatrics Outpatient Clinics were included into the scope of this study. The mean age±SEM of 92 morbid obese (MO) children and 36 with MetS were 118.3±3.8 months and 129.5±6.4 months, respectively (p > 0.05). The study was approved by Namık Kemal University, Medical Faculty Ethics Committee. Written informed consent forms were obtained from the parents. Demographic features and anthropometric measurements were recorded. WHO BMI-for age percentiles were used. The values above 99 percentile were defined as MO. Components of MetS [waist circumference (WC), fasting blood glucose (FBG), triacylglycerol (TRG), high density lipoprotein cholesterol (HDL-Chol), systolic pressure (SP), diastolic pressure (DP)] were determined. Routine laboratory tests were performed. Serum vitamin B₁₂ concentrations were measured using electrochemiluminescence immunoassay. Vitamin B₉ was analyzed by an immunoassay analyzer. Values for vitamin B₁₂ < 148 pmol/L, 148-221 pmol/L, > 221 pmol/L were accepted as low, borderline and normal, respectively. Vitamin B₉ levels ≤ 4 mcg/L defined deficiency state. Statistical evaluations were performed by SPSSx Version 16.0. p≤0.05 was accepted as statistical significance level. Statistically higher body mass index (BMI), WC, hip circumference (C) and neck C were calculated in MetS group compared to children with MO. No difference was noted for head C. All MetS components differed between the groups (SP, DP p < 0.001; WC, FBG, TRG p < 0.01; HDL-Chol p < 0.05). Significantly decreased vitamin B₉ and vitamin B₁₂ levels were detected (p < 0.05) in children with MetS. In both groups percentage of folate deficiency was 5.5%. No cases were below < 148 pmol/L. However, in MO group 14.3% and in MetS group 22.2% of the cases were of borderline status. In MO group B₁₂ levels were negatively correlated with BMI, WC, hip C and head C, but not with neck C. WC, hip C, head C and neck C were all negatively correlated with HDL-Chol. None of these correlations were observed in the group of children with MetS. Strong positive correlation between FBG and insulin as well as strong negative correlation between TRG and HDL-Chol detected in MO children were lost in MetS group. Deficiency state end-products of both B₉ and B₁₂ may interfere with the expected profiles of MetS components. In this study, the alterations in MetS components affected vitamin B₁₂ metabolism and also its associations with anthropometric body measurements. Further increases in vitamin B₁₂ and vitamin B₉ deficiency in MetS associated with the increased vitamin B₁₂ as well as vitamin B₉ deficiency metabolites may add to MetS parameters.

Keywords: children, cobalamin, folate, metabolic syndrome, obesity

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Authors: Magdalena B. Kaziuk, Waldemar Kosiba, Marek J. Kuzniewski

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Introduction: Adipose tissue is a typical place for storage water-insoluble toxins in the body. It's connective tissue, where the intercellular substance consist of fat, which level in people with low physical activity should be 18-25% for women and 13-18% for men. Due to the fat distribution in the body we distinquish two types of obesity: android (visceral, abdominal) and gynoidal (gluteal-femoral, peripheral). Abdominal obesity increases the risk of complications of the cardiovascular system diseases, and impaired renal and liver function. Through the influence on disorders of metabolism, lipid metabolism, diabetes and hypertension, leading to emergence of the metabolic syndrome. So thus, obesity will especially overload kidney function in patients after transplantation. Aim: An attempt was made to estimate the impact of amount fat tissue on transplanted kidney function and excretory function of the liver in patients after Ktx. Material and Methods: The study included 108 patients (50 females, 58 male, age 46.5 +/- 12.9 years) with active kidney transplant after more than 3 months from the transplantation. An analysis of body composition was done by using electrical bioimpedance (BIA) and anthropometric measurements. Estimated basal metabolic rate (BMR), muscle mass, total body water content and the amount of body fat. Information about physical activity were obtained during clinical examination. Nutritional status, and type of obesity were determined by using indicators: Waist to Height Ratio (WHR) and Waist to Hip Ratio (WHR). Excretory functions of the transplanted kidney was rated by calculating the estimated renal glomerular filtration rate (eGFR) using the MDRD formula. Liver function was rated by total bilirubin and alanine aminotransferase levels ALT concentration in serum. In our patients haemolitic uremic syndrome (HUS) was excluded. Results: In 19.44% of patients had underweight, 22.37% of the respondents were with normal weight, 11.11% had overweight, and the rest were with obese (49.08%). People with android stature have a lower eGFR compared with those with the gynoidal stature (p = 0.004). All patients with obesity had higher amount of body fat from a few to several percent. The higher amount of body fat percentage, the lower eGFR had patients (p <0.001). Elevated ALT levels significantly correlated with a high fat content (p <0.02). Conclusion: Increased amount of body fat, particularly in the case of android obesity can be a predictor of kidney and liver damage. Due to that obese patients should have more frequent control of diagnostic functions of these organs and the intensive dietary proceedings, pharmacological and regular physical activity adapted to the current physical condition of patients after transplantation.

Keywords: obesity, body fat, kidney transplantation, glomerular filtration rate, liver function

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Authors: Vilma Sasyte, Vilma Viliene, Asta Raceviciute-Stupeliene, Agila Dauksiene, Romas Gruzauskas, Virginijus Slausgalvis, Jamal Al-Saifi

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The aim of the present study was to investigate the effect of butyrate, acids, medium-chain fatty acids and plant extract mixture on performance, blood and gastrointestinal tract characteristics of laying hens’. For the period of 8 weeks, 24 Hisex Brown laying hens were randomly assigned to 2 dietary treatments: 1) control wheat-corn-soybean meal based diet (Control group), 2) control diet supplemented with the mixture of butyrate, acids, medium chain fatty acids and plant extract (Lumance®) at the level of 1.5 g/kg of feed (Experimental group). Hens were fed with a crumbled diet at 125 g per day. Housing and feeding conditions were the same for all groups and met the requirements of growth for laying hens of Hisex Brown strain. In the blood serum total protein, bilirubin, cholesterol, DTL- and MTL- cholesterol, triglycerides, glucose, GGT, GOT, GPT, alkaline phosphatase, alpha amylase, contents of c-reactive protein, uric acid, and lipase were analyzed. Development of intestines and internal organs (intestinal length, intestinal weight, the weight of glandular and muscular stomach, pancreas, heart, and liver) were determined. The concentration of short chain fatty acids in caecal content was measured using the method of HPLC. The results of the present study showed that 1.5 g/kg supplementation of feed additive affected egg production and feed conversion ratio for the production of 1 kg of egg mass. Dietary supplementation of analyzed additive in the diets increased the concentration of triglycerides, GOT, alkaline phosphatase and decreased uric acid content compared with the control group (P<0.05). No significant difference for others blood indices in comparison to the control was observed. The addition of feed additives in laying hens’ diets increased intestinal weight by 11% and liver weight by 14% compared with the control group (P<0.05). The short chain fatty acids (propionic, acetic and butyric acids) in the caecum of laying hens in experimental groups decreased compared with the control group. The supplementation of the mixture of butyrate, acids, medium-chain fatty acids and plant extract at the level of 1.5 g/kg in the laying hens’ diets had the effect on the performance, some gastrointestinal tract function and blood parameters of laying hens.

Keywords: acids, butyrate, laying hens, MCFA, performance, plant extract, psysiological state

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Authors: Alireza Barari, Ahmad Abdi

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Regular body trainings cause adaption in various system in body. One of the important effect of body training is its effect on immune system. It seems that cytokines usually release after long period exercises or some exercises which cause skeletal muscular damages. If some of the cytokines which cause responses such as inflammation of cells in skeletal muscles, with manipulating of training program, it can be avoided or limited from those exercises which induct cytokines release. Ginger plant is a kind of medicinal plants which is known as a anti inflammation plant. This plant is as most precedence medicinal plants in medicine science especially in inflammation cure. The aim of the present study was the effect of selected resistance training and consumption of ginger extract on IL-1α and TNFα untrained young women. The population includes young women interested in participating in the study with the average of 30±2 years old from Abbas Abad city among which 32 participants were chosen randomly and divided into 4 four groups, resistance training (R), resistance training and ginger consumption(RG), Ginger consumption(G)and Control group(C). The training groups performed circuit resistance training at the intensity of 65-75% one repeat maximum, 3 days a week for 6 weeks. Besides resistance training, subjects were given either ginseng (5 mg/kg per day) or placebo. Prior to and 48 hours after interventions body composition was measured and blood samples were taken in order to assess serum levels of IL-1α and TNFα. Plasma levels of cytokines were measured with commercially available ELISA Kits.IL-1α kit and TNFα kit were used in this research. To demonstrate the effectiveness of the independent variable and the comparison between groups, t-test and ANOVA were used. To determine differences between the groups, the Scheffe test was used that showed significant changes in any of the variables. we observed that circuit resistance training in R and RG groups can significant decreased in weight and body mass index in untrained females (p<0.05). The results showed a significant decreased in the mean level of IL-1α levels before and after the training period in G group (p=0.046) and RG group (p=0.022). Comparison between groups also showed there was significant difference between groups R-RG and RG-C. Intergroup comparison results showed that the mean levels of TNFα before and after the training in group G (p=0.044) and RG (p=0.037), significantly decreased. Comparison between groups also showed there was significant difference between groups R–RG , R-G ,RG-C and G-C. The research shows that circuit resistance training with reducing overload method results in systemic inflammation had significant effect on IL-1α levels and TNFα. Of course, Ginger can counteract the negative effects of resistance training exercise on immune function and stability of the mast cell membrane. Considerable evidence supported the anti-inflammatory properties of ginger for several constituents, especially gingerols, shogaols, paradols, and zingerones, through decreased cytokine gene TNF α and IL-1Α expression and inhibition of cyclooxygenase 1 and 2. These established biological actions suggest that ingested ginger could block the increase in IL-1α.

Keywords: resistance training, ginger, IL-1α , TNFα

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Authors: Kathrina Aseanne Acapulco-Gomez, Shayne Julieane Morales, Tzar Francis Verame

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Mean platelet volume (MPV) is the most accurate measure of the size of platelets and is routinely measured by most automated hematological analyzers. Several studies have shown associations between MPV and cardiovascular risks and outcomes. Although its measurement may provide useful data, MPV remains to be a diagnostic tool that is yet to be included in routine clinical decision making. The aim of this systematic review and meta-analysis is to determine summary estimates of the diagnostic accuracy of mean platelet volume for the diagnosis of myocardial infarction among adult patients with angina and/or its equivalents in terms of sensitivity, specificity, diagnostic odds ratio, and likelihood ratios, and to determine the difference of the mean MPV values between those with MI and those in the non-MI controls. The primary search was done through search in electronic databases PubMed, Cochrane Review CENTRAL, HERDIN (Health Research and Development Information Network), Google Scholar, Philippine Journal of Pathology, and Philippine College of Physicians Philippine Journal of Internal Medicine. The reference list of original reports was also searched. Cross-sectional, cohort, and case-control articles studying the diagnostic performance of mean platelet volume in the diagnosis of acute myocardial infarction in adult patients were included in the study. Studies were included if: (1) CBC was taken upon presentation to the ER or upon admission (within 24 hours of symptom onset); (2) myocardial infarction was diagnosed with serum markers, ECG, or according to accepted guidelines by the Cardiology societies (American Heart Association (AHA), American College of Cardiology (ACC), European Society of Cardiology (ESC); and, (3) if outcomes were measured as significant difference AND/OR sensitivity and specificity. The authors independently screened for inclusion of all the identified potential studies as a result of the search. Eligible studies were appraised using well-defined criteria. Any disagreement between the reviewers was resolved through discussion and consensus. The overall mean MPV value of those with MI (9.702 fl; 95% CI 9.07 – 10.33) was higher than in those of the non-MI control group (8.85 fl; 95% CI 8.23 – 9.46). Interpretation of the calculated t-value of 2.0827 showed that there was a significant difference in the mean MPV values of those with MI and those of the non-MI controls. The summary sensitivity (Se) and specificity (Sp) for MPV were 0.66 (95% CI; 0.59 - 0.73) and 0.60 (95% CI; 0.43 – 0.75), respectively. The pooled diagnostic odds ratio (DOR) was 2.92 (95% CI; 1.90 – 4.50). The positive likelihood ratio of MPV in the diagnosis of myocardial infarction was 1.65 (95% CI; 1.20 – 22.27), and the negative likelihood ratio was 0.56 (95% CI; 0.50 – 0.64). The intended role for MPV in the diagnostic pathway of myocardial infarction would perhaps be best as a triage tool. With a DOR of 2.92, MPV values can discriminate between those who have MI and those without. For a patient with angina presenting with elevated MPV values, it is 1.65 times more likely that he has MI. Thus, it is implied that the decision to treat a patient with angina or its equivalents as a case of MI could be supported by an elevated MPV value.

Keywords: mean platelet volume, MPV, myocardial infarction, angina, chest pain

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Authors: Zain Ul Abadeen, Muhammad Zargham Khan, Muhammad Kashif Saleemi, Ahrar Khan, Ijaz Javed Hassan, Aisha Khatoon, Qasim Altaf

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Aflatoxins are secondary metabolites produced by toxigenic fungi, and there are four types of aflatoxins include AFB1, AFB2, AFG1 and AFG2. Aflatoxin B1 (AFB1) is considered as most toxic form. It is mainly responsible for the contamination of poultry feed and produces a condition called aflatoxicosis leads to immunosuppression in poultry birds. Saccharomyces cerevisiae is a single cell microorganism and acts as a source of growth factors, minerals and amino acids which improve the immunity and digestibility in poultry birds as probiotics. Saccharomyces cerevisiae is well recognized to cause the biological degradation of mycotoxins (toxin binder) because its cell wall contains β-glucans and mannans which specifically bind with aflatoxins and reduce their absorption or transfer them to some non-toxic compounds. The present study was designed to investigate the immunosuppressive effects of aflatoxins in broiler chicks and the reduction of severity of these effects by the use of Baker’s Yeast (Saccharomyces cerevisiae). One-day-old broiler chicks were procured from local hatchery and were divided into various groups (A-I). These groups were treated with different levels of AFB1 @ 400 µg/kg and 600 µg/kg along with different levels of Baker’s Yeast (Saccharomyces cerevisiae) 0.1% and 0.5 % in the feed. The total duration of the experiment was six weeks and different immunological parameters including the cellular immune response by injecting PHA-P (Phytohemagglutinin-P) in the skin of the birds, phagocytic function of mononuclear cells by Carbon clearance assay from blood samples and humoral immune response against intravenously injected sheep RBCs from the serum samples were determined. The birds from each group were slaughtered at the end of the experiment to determine the presence of gross lesions in the immune organs and these tissues were fixed in 10% neutral buffered formalin for histological investigations. The results showed that AFB1 intoxicated groups had reduced body weight gain, feed intake, organs weight and immunological responses compared to the control and Baker’s Yeast (Saccharomyces cerevisiae) treated groups. Different gross and histological degenerative changes were recorded in the immune organs of AFB1 intoxicated groups compared to control and Baker’s Yeast (Saccharomyces cerevisiae) treated groups. The present study concluded that Baker’s Yeast (Saccharomyces cerevisiae) addition in the feed helps to ameliorate the immunotoxigenic effects produced by AFB1 in broiler chicks.

Keywords: aflatoxins, body weight gain, feed intake, immunological response, toxigenic effect

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Authors: Anne-Margré C. Vink

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Background: Food-related allergies and intolerances are frequently occurring in dogs. Diagnosis and monitoring according to ‘Golden Standard’ of elimination efficiency are time-consuming, expensive, and requires expert clinical setting. In order to facilitate rapid and robust, quantitative testing of intolerance, and determining the individual offending foods, a serological test is implicated. Method: As we developed Medisynx IgG Human Screening Test ELISA before and the dog’s immune system is most similar to humans, we were able to develop Medisynx IgG Dog Screening Test ELISA as well. In this study, 47 dogs suffering from Canine Atopic Dermatitis (CAD) and several secondary induced reactions were included to participate in serological Medisynx IgG Dog Screening Test ELISA (within < 0,02 % SD). Results were expressed as titers relative to the standard OD readings to diagnose alimentary induced diseases and monitoring the efficacy of an individual eliminating diet in dogs. Split sample analysis was performed by independently sending 2 times 3 ml serum under two unique codes. Results: The veterinarian monitored these dogs to check dog’ results at least at 3, 7, 21, 49, 70 days and after period of 6 and 12 months on an individual negative diet and a positive challenge (retrospectively) at 6 months. Data of each dog were recorded in a screening form and reported that a complete recovery of all clinical manifestations was observed at or less than 70 days (between 50 and 70 days) in the majority of dogs(44 out of 47 dogs =93.6%). Conclusion: Challenge results showed a significant result of 100% in specificity as well as 100% positive predicted value. On the other hand, sensitivity was 95,7% and negative predictive value was 95,7%. In conclusion, an individual diet based on IgG ELISA in dogs provides a significant improvement of atopic dermatitis and pruritus including all other non-specific defined allergic skin reactions as erythema, itching, biting and gnawing at toes, as well as to several secondary manifestations like chronic diarrhoea, chronic constipation, otitis media, obesity, laziness or inactive behaviour, pain and muscular stiffness causing a movement disorders, excessive lacrimation, hyper behaviour, nervous behaviour and not possible to stay alone at home, anxiety, biting and aggressive behaviour and disobedience behaviour. Furthermore, we conclude that a relatively more severe systemic candidiasis, as shown by relatively higher titer (class 3 and 4 IgG reactions to Candida albicans), influence the duration of recovery from clinical manifestations in affected dogs. These findings are consistent with our preliminary human clinical studies.

Keywords: allergy, canine atopic dermatitis, CAD, food allergens, IgG-ELISA, food-incompatibility

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Authors: Ngwa Lodence Njwe

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This study investigated the response of hemodialysis patients to an established nutrition therapy guide accompanied by a 7-day menu plan administered for a month. End Stage Renal Disease (ESRD), also known as End Stage Kidney Disease (ESKD), is a non-communicable disease primarily caused by hypertension and diabetes, posing significant challenges in both developed and developing nations. Hemodialysis is a key treatment for these patients. In this experimental study, 100 hemodialysis patients from Douala General Hospital in Cameroon participated. A questionnaire was used to collect data on sociodemographic and anthropometric characteristics, health status, and dietary intake, while medical records provided biomedical data. The levels of the biochemical parameters (Phosphorus, calcium and hemoglobin) were determined before and one month after the distribution of the nutrition education guide and the use of a 7-day menu plan. The Phosphorus and Calcium levels were measured using an LTCC03 semi-automatic chemistry analyzer. Blood was collected from each patient into a test tube, allowed to clot and centrifuged. 50µl of the serum was aspirated by the analyzer for Ca and P level analysis, and results were read from the display. The hemoglobin level was measured using the URIT–12 hemoglobin Meter. The blood sample was collected by hand prick and placed in a strip, and the results were read from the screen. The means of the biochemical parameters were then computed. The most prevalent age group was 40-49 years, with males constituting 70% and females 30% of respondents. Among these patients, 80% were hypertensive, 3% had both hypertension and diabetes, 9% were hypertensive, diabetic, and obese, and 1% suffered from hypertension and heart failure. Analysis of anthropometric parameters revealed a high prevalence of underweight, overweight, and obesity, highlighting the urgent need for targeted nutrition interventions to modify cooking methods, enhance food choices, and increase dietary variety for improved quality of life. Before the nutrition therapy guide was implemented, average calcium levels were 73.05 mg/L for males and 89.44 mg/L for females; post-implementation, these values increased to 77.55 mg/L and 91.44 mg/L, respectively. Conversely, average phosphorus levels decreased from 42.05 mg/L for males and 43.55 mg/L for females to 41.05 mg/L and 39.3 mg/L, respectively, after the intervention. Additionally, average hemoglobin levels increased from 8.35 g/dL for males and 8.5 g/dL for females to 9.2 g/dL and 8.95 g/dL, respectively. The findings confirm that the nutrition therapy guide and the 7-day menu significantly impacted the biomedical parameters of hemodialysis patients, underscoring the need for ongoing nutrition education and counseling for this population.

Keywords: end stage kidney disease, nutrition therapy guide, nutritional status, anthropometric parameters, food frequency, biomedical data

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Authors: Mahmut Sozmen, Alparslan Kadir Devrim, Yonca Betil Kabak, Tuba Devrim

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Acute myocardial infarction is the leading cause of deaths in the worldwide. Mature cardiomyocytes do not have the ability to regenerate instead fibrous tissue proliferate and granulation tissue to fill out. Periostin is an extracellular matrix protein from fasciclin family and it plays an important role in the cell adhesion, migration, and growth of the organism. Periostin prevents apoptosis while stimulating cardiomyocytes. The main objective of this project is to investigate the effects of the recombinant murine periostin peptide administration for the cardiomyocyte regeneration in a rat model of acute myocardial infarction. The experiment was performed on 84 male rats (6 months old) in 4 group each contains 21 rats. Saline applied subcutaneously (1 ml/kg) two times with 24 hours intervals to the rats in control group (Group 1). Recombinant periostin peptide (1 μg/kg) dissolved in saline applied intraperitoneally in group 2 on 1, 3, 7, 14 and 21. days on same dates in group 4. Isoproterenol dissolved in saline applied intraperitoneally (85mg/kg/day) two times with 24 hours intervals to the groups 3 and 4. Rats in group 4 further received recombinant periostin peptide (1 μg/kg) dissolved in saline intraperitoneally starting one day after the final isoproterenol administration on days 1, 3, 7, 14 and 21. Following the final application of periostin rats continued to feed routinely with pelleted chow and water ad libitum for further seven days. At the end of 7th day rats sacrificed, blood and heart tissue samples collected for the immunohistochemical and biochemical analysis. Angiogenesis in response to tissue damage, is a highly dynamic process regulated by signals from the surrounding extracellular matrix and blood serum. In this project, VEGF, ANGPT, bFGF, TGFβ are the key factors that contribute to cardiomyocyte regeneration were investigated. Additionally, the relationship between mitosis and apoptosis (Bcl-2, Bax, PCNA, Ki-67, Phopho-Histone H3), cell cycle activators and inhibitors (Cyclin D1, D2, A2, Cdc2), the origin of regenerating cells (cKit and CD45) were examined. Present results revealed that periostin stimulated cardiomyocye cell-cycle re-entry in both normal and MCA damaged cardiomyocytes and increased angiogenesis. Thus, periostin contributes to cardiomyocyte regeneration during the healing period following myocardial infarction which provides a better understanding of its role of this mechanism, improving recovery rates and it is expected to contribute the lack of literature on this subject. Acknowledgement: This project was financially supported by Turkish Scientific Research Council- Agriculture, Forestry and Veterinary Research Support Group (TUBİTAK-TOVAG; Project No: 114O734), Ankara, TURKEY.

Keywords: cardiotoxicity, immunohistochemistry, isoproterenol, periostin

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Authors: Amal Balila, Maria Vahdati

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Earth is a green building material with very low embodied energy and almost zero greenhouse gas emissions. However, it lacks strength and durability in its natural state. By responsibly sourcing stabilisers, it's possible to enhance its strength. This research draws inspiration from the robustness of termite mounds, where termites incorporate glycoproteins from their saliva during construction. Biomimicry explores the potential of these termite stabilisers in producing bio-inspired adobe bricks. The meat industry generates significant waste during slaughter, including blood, skin, bones, tendons, gastrointestinal contents, and internal organs. While abundant, many meat by-products raise concerns regarding human consumption, religious orders, cultural and ethical beliefs, and also heavily contribute to environmental pollution. Extracting and utilising proteins from this waste is vital for reducing pollution and increasing profitability. Exploring the untapped potential of meat industry waste, this research investigates how glycoproteins could revolutionize adobe brick construction. Bovine serum albumin (BSA) from cows' blood and mucin from porcine stomachs were the chosen glycoproteins used as stabilisers for adobe brick production. Despite their wide usage across various fields, they have very limited utilisation in food processing. Thus, both were identified as potential stabilisers for adobe brick production in this study. Two soil types were utilised to prepare adobe bricks for testing, comparing controlled unstabilised bricks with glycoprotein-stabilised ones. All bricks underwent testing for unconfined compressive strength and erosion resistance. The primary finding of this study is the efficacy of BSA, a glycoprotein derived from cows' blood and a by-product of the beef industry, as an earth construction stabiliser. Adding 0.5% by weight of BSA resulted in a 17% and 41% increase in the unconfined compressive strength for British and Sudanese adobe bricks, respectively. Further, adding 5% by weight of BSA led to a 202% and 97% increase in the unconfined compressive strength for British and Sudanese adobe bricks, respectively. Moreover, using 0.1%, 0.2%, and 0.5% by weight of BSA resulted in erosion rate reductions of 30%, 48%, and 70% for British adobe bricks, respectively, with a 97% reduction observed for Sudanese adobe bricks at 0.5% by weight of BSA. However, mucin from the porcine stomach did not significantly improve the unconfined compressive strength of adobe bricks. Nevertheless, employing 0.1% and 0.2% by weight of mucin resulted in erosion rate reductions of 28% and 55% for British adobe bricks, respectively. These findings underscore BSA's efficiency as an earth construction stabiliser for wall construction and mucin's efficacy for wall render, showcasing their potential for sustainable and durable building practices.

Keywords: biomimicry, earth construction, industrial waste management, sustainable building materials, termite mounds.

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Authors: Wu Liching

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Aim This case aims to discuss the pathogenesis, clinical manifestations and medical treatment of strokes caused by mitochondrial gene mutations. Methods Diagnosis of ischemic stroke caused by mitochondrial gene defect by means of "next-generation sequencing mitochondrial DNA gene variation detection", imaging examination, neurological examination, and medical history; this study took samples from the neurology ward of a medical center in northern Taiwan cases diagnosed with acute cerebral infarction as the research objects. Result This case is a 49-year-old married woman with a rare disease, mitochondrial gene mutation inducing ischemic stroke. She has severe hearing impairment and needs to use hearing aids, and has a history of diabetes. During the patient’s hospitalization, the blood test showed that serum Lactate: 7.72 mmol/L, Lactate (CSF) 5.9 mmol/L. Through the collection of relevant medical history, neurological evaluation showed changes in consciousness and cognition, slow response in language expression, and brain magnetic resonance imaging examination showed subacute bilateral temporal lobe infarction, which was an atypical type of stroke. The lineage DNA gene has m.3243A>G known pathogenic mutation point, and its heteroplasmic level is 24.6%. This pathogenic point is located in MITOMAP and recorded as Mitochondrial Encephalopathy, Lactic Acidosis, and Stroke-like episodes (MELAS) , Leigh Syndrome and other disease-related pathogenic loci, this mutation is located in ClinVar and recorded as Pathogenic (dbSNP: rs199474657), so it is diagnosed as a case of stroke caused by a rare disease mitochondrial gene mutation. After medical treatment, there was no more seizure during hospitalization. After interventional rehabilitation, the patient's limb weakness, poor language function, and cognitive impairment have all improved significantly. Conclusion Mitochondrial disorders can also be associated with abnormalities in psychological, neurological, cerebral cortical function, and autonomic functions, as well as problems with internal medical diseases. Therefore, the differential diagnoses cover a wide range and are not easy to be diagnosed. After neurological evaluation, medical history collection, imaging and rare disease serological examination, atypical ischemic stroke caused by rare mitochondrial gene mutation was diagnosed. We hope that through this case, the diagnosis of rare disease mitochondrial gene variation leading to cerebral infarction will be more familiar to clinical medical staff, and this case report may help to improve the clinical diagnosis and treatment for patients with similar clinical symptoms in the future.

Keywords: acute stroke, MELAS, lactic acidosis, mitochondrial disorders

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Authors: Bipasa Dey, Varsha Panwar, Tanmay Dutta

Abstract:

Laccase, a multicopper oxidase (EC 1.10.3.2) have been at the forefront as a superior industrial biocatalyst. They are versatile in terms of bestowing sustainable and ecological catalytic reactions such as polymerisation, xenobiotic degradation and bioremediation of phenolic and non-phenolic compounds. Regardless of the wide biotechnological applications, the critical limiting factors viz. reusability, retrieval, and storage stability still prevail. This can cause an impediment in their applicability. Crosslinked enzyme aggregates (CLEAs) have emerged as a promising technique that rehabilitates these essential facets, albeit at the expense of their enzymatic activity. The carrier free crosslinking method prevails over the carrier-bound immobilisation in conferring high productivity, low production cost owing to the absence of additional carrier and circumvent any non-catalytic ballast which could dilute the volumetric activity. To the best of our knowledge, the ε-amino group of lysyl residue is speculated as the best choice for forming Schiff’s base with glutaraldehyde. Despite being most preferrable, excess glutaraldehyde can bring about disproportionate and undesirable crosslinking within the catalytic site and hence could deliver undesirable catalytic losses. Moreover, the surface distribution of lysine residues in Trametes versicolor laccase is significantly less. Thus, to mitigate the adverse effect of glutaraldehyde in conjunction with scaling down the degradation or catalytic loss of the enzyme, crosslinking with inert substances like gelatine, collagen, Bovine serum albumin (BSA) or excess lysine is practiced. Analogous to these molecules, sugars have been well known as a protein stabiliser. It helps to retain the structural integrity, specifically secondary structure of the protein during aggregation by changing the solvent properties. They are comprehended to avert protein denaturation or enzyme deactivation during precipitation. We prepared crosslinked enzyme aggregates (CLEAs) of laccase from T. versicolor with the aid of sugars. The sugar CLEAs were compared with the classic BSA and glutaraldehyde laccase CLEAs concerning physico-chemical properties. The activity recovery for the fructose CLEAs were found to be ~20% higher than the non-sugar CLEA. Moreover, the 𝐾𝑐𝑎𝑡𝐾𝑚⁄ values of the CLEAs were two and three-fold higher than BSA-CLEA and GACLEA, respectively. The half-life (t1/2) deciphered by sugar-CLEA was higher than the t1/2 of GA-CLEAs and free enzyme, portraying more thermal stability. Besides, it demonstrated extraordinarily high pH stability, which was analogous to BSA-CLEA. The promising attributes of increased storage stability and recyclability (>80%) gives more edge to the sugar-CLEAs over conventional CLEAs of their corresponding free enzyme. Thus, sugar-CLEA prevails in furnishing the rudimentary properties required for a biocatalyst and holds many prospects.

Keywords: cross-linked enzyme aggregates, laccase immobilization, enzyme reusability, enzyme stability

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Authors: Z. Karahaliloğlu, E. Yalçın, M. Demirbilek, E.B. Denkbaş

Abstract:

Critical-sized bone defects caused by trauma, bone diseases, prosthetic implant revision or tumor excision cannot be repaired by physiological regenerative processes. Current orthopedic applications for critical-sized bone defects are to use autologous bone grafts, bone allografts, or synthetic graft materials. However, these strategies are unable to solve completely the problem, and motivate the development of novel effective biological scaffolds for tissue engineering applications and regenerative medicine applications. In particular, scaffolds combined with a variety of bio-agents as fundamental tools emerge to provide the regeneration of damaged bone tissues due to their ability to promote cell growth and function. In this study, a magnetic silk fibroin (SF) hydrogel scaffold was prepared by electrogelation process of the concentrated Bombxy mori silk fibroin (8 %wt) aqueous solution. For enhancement of osteoblast-like cells (SaOS-2) growth and adhesion, basal fibroblast growth factor (bFGF) were conjugated physically to the HSA-coated magnetic nanoparticles (Fe3O4) and magnetic SF e-gel scaffolds were prepared by incorporation of Fe3O4, HSA (human serum albumin)=Fe3O4 and HSA=Fe3O4-bFGF nanoparticles. HSA=Fe3O4, HSA=Fe3O4-bFGF loaded and bare SF e-gels scaffolds were characterized using scanning electron microscopy (SEM.) For cell studies, human osteoblast-like cell line (SaOS-2) was used and an MTT assay was used to assess the cytotoxicity of magnetic silk fibroin e-gel scaffolds and cell density on these surfaces. For the evaluation osteogenic activation, ALP (alkaline phosphatase), the amount of mineralized calcium, total protein and collagen were studied. Fe3O4 nanoparticles were successfully synthesized and bFGF was conjugated to HSA=Fe3O4 nanoparticles with %97.5 of binding yield which has a particle size of 71.52±2.3 nm. Electron microscopy images of the prepared HSA and bFGF incorporated SF e-gel scaffolds showed a 3D porous morphology. In terms of water uptake results, bFGF conjugated HSA=Fe3O4 nanoparticles has the best water absorbability behavior among all groups. In the in-vitro cell culture studies realized using SaOS-2 cell line, the coating of Fe3O4 nanoparticles surface with a protein enhance the cell viability and HSA coating and bFGF conjugation, the both have an inductive effect in the cell proliferation. One of the markers of bone formation and osteoblast differentiation, according to the ALP activity and total protein results, HSA=Fe3O4-bFGF loaded SF e-gels had significantly enhanced ALP activity. Osteoblast cultured HSA=Fe3O4-bFGF loaded SF e-gels deposited more calcium compared with SF e-gel. The proposed magnetic scaffolds seem to be promising for bone tissue regeneration and used in future work for various applications.

Keywords: basic fibroblast growth factor (bFGF), e-gel, iron oxide nanoparticles, silk fibroin

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Authors: Antonio Munar-Frau, Sascha Klismoch, Manfred Schmolz, Federico Hernandez-Alfaro, Jordi Caballe-Serrano

Abstract:

Introduction: Biocompatibility of biomaterials has been proposed as one of the main criteria for treatment success. For guided bone regeneration (GBR), barrier membranes present a conflict given the number of origins and modifications of these materials. The biologic response to biomaterials is orchestrated by a series of events leading to the integration or rejection of the biomaterial, posing questions such as if a longer occlusive property may trigger an inflammatory reaction. Whole blood cultures are a solution to study the immune response to drugs or biomaterials during the first 24-48 hours. The aim of this study is to determine the early immune response of different origins and chemical modifications of barrier membranes. Materials & Methods: 5 different widely used barrier membranes were included in this study: Acellular dermal matrix (AlloDerm, LifeCell®), Porcine Peritoneum (BioGide, Geistlich Pharma®), Porcine Pericardium (Jason, Botiss Biomaterials GmbH®), Porcine Cross-linked collagen (Ossix Plus, Datum Dental®) and d-PTFE (Cytoplast TXT, Osteogenics Biomedical®). Blood samples were extracted from 3 different healthy donors and incubated with the different samples of barrier membranes for 24 hours. After the incubation time, serum samples were obtained and analyzed by means of biocompatibility assays taking into account 42 markers. Results: In an early stage of the inflammatory response, the Acellular dermal matrix, porcine peritoneum and porcine cross-linked collagen expressed similar patterns of cytokine expression with a great manifestation of ENA 78. Porcine pericardium and d-PTFE presented similar cytokine activation, especially for MMP-3 and MMP-9, although other cytokines were highlighted with lower expression. For the later immune response, Porcine peritoneum and acellular dermal matrix MCP-1 and IL-15 were evident. Porcine pericardium, porcine cross-linked collagen and d-PTFE presented a high expression of IL-16 and lower manifestation of other cytokines. Different behaviors depending on an earlier or later stage of the inflammation process were observed. Barrier membrane inflammatory expression does not only differ depending on the origin, variables such as treatment of the collagen and polymers may also have a great impact on the cytokine expression of the studied barrier membranes during inflammation. Conclusions: Surface treatment and modifications might affect the biocompatibility of the membranes, as different cytokine expressions were evidently depending on the origin of the biomaterial. This study is only a brushstroke regarding the biocompatibility of materials, as it is one of the pioneer studies for ex vivo barrier membranes assays. Studies regarding surface modification are needed in order to clarify mystifications of barrier membrane science.

Keywords: biomaterials, bone regeneration, biocompatibility, inflammation

Procedia PDF Downloads 158

Authors: S. A. Bhat, Ahmad Arif, Muneeb U. Rehman, Manzoor R Mir, S. Bilal, Ishraq Hussain, H. M Khan, S. Shanaz, M. I Mir, Sabhiya Majid

Abstract:

Background: Geologically Climatic conditions of the state range from sub-tropical (Jammu), temperate (Kashmir) to cold artic (Ladakh) zones, which exerts significant influence on its agro-climatic conditions. Gastrointestinal parasitism is a major problem in sheep production worldwide. Materials and Methods: The present study was to evaluate the resistance status of sheep breeds reared in Kashmir Valley for natural resistance against Haemonchus contortus by natural pasture challenge infection. Ten microsatellite markers were used in the study for evaluation of association of Ovar-MHC with parasitic resistance in association with biochemical and parasitological parameters. Following deworming, 500 animals were subjected to selected contaminated pastures in a vicinity of the livestock farms of SKUAST-K and Sheep Husbandry Kashmir. For each animal about 10-15 ml blood was collected aseptically for molecular and biochemical analysis. Weekly fecal samples (3g) were taken, directly from the rectum of all experimental animals and examined for Fecal egg count (FEC) with modified McMaster technique. Packed cell volume (PCV) was determined within 2-5 h of blood collection, all the biochemical parameters were determined in serum by semi automated analyzer. DNA was extracted from all the blood samples with phenol-chloroform method. Microsatellite analysis was done by denaturing sequencing gel electrophoresis Results: Overall sheep from Bakerwal breed followed by Corriediale breed performed relatively better in the trial; however difference between breeds remained low. Both significant (P<0.05) and non-significant differences with respect to resistance against haemonchosis were noted at different intervals in all the parameters.. All the animals were typed for the microsatellites INRA132, OarCP73, DRB1 (U0022), OLA-DQA2, BM1818, TFAP2A, HH56, BM1815, IL-3 and BM-1258. An association study including the effect of FEC, PCV, TSP, SA, LW, and the number of alleles within each marker was done. All microsatellite markers showed degree of heterozygosity of 0.72, 0.72, 0.75, 0.62, 0.84, 0.69, 0.66, 0.65, 0.73 and 0.68 respectively. Significant association between alleles and the parameters measured were only found for the OarCP73, OLA-DQA2 and BM1815 microsatellite marker. Standard alleles of the above markers showed significant effect on the TP, SA and body weight. The three sheep breeds included in the study responded differently to the nematode infection, which may be attributed to their differences in their natural resistance against nematodes. Conclusion: Our data confirms that some markers (OarCP73, OLA-DQA2 and BM1815) within Ovar-MHC are associated with phenotypic parameters of resistance and suggest superiority of Bakerwal sheep breed in natural resistance against Haemonchus contortus.

Keywords: Ovar-Mhc, ovine leukocyte antigen (OLA), sheep, parasitic resistance, Haemonchus contortus, phenotypic & genotypic markers

Procedia PDF Downloads 712

Authors: Eva Filová, Jana Daňková, Věra Sovková, Matej Daniel

Abstract:

Titanium alloys are biocompatible metals that are widely used in clinical practice as load bearing implants. The chemical modification may influence cell adhesion, proliferation, and differentiation as well as stiffness of the material. The aim of the study was to evaluate the adhesion, growth and differentiation of pig mesenchymal stem cells on the novel beta titanium alloy Ti36Nb6Ta compared to standard medical titanium alloy Ti6Al4V. Discs of Ti36Nb6Ta and Ti6Al4V alloy were sterilized by ethanol, put in 48-well plates, and seeded by pig mesenchymal stem cells at the density of 60×103/cm2 and cultured in Minimum essential medium (Sigma) supplemented with 10% fetal bovine serum and penicillin/streptomycin. Cell viability was evaluated using MTS assay (CellTiter 96® AQueous One Solution Cell Proliferation Assay;Promega), cell proliferation using Quant-iT™ ds DNA Assay Kit (Life Technologies). Cells were stained immunohistochemically using monoclonal antibody beta-actin, and secondary antibody conjugated with AlexaFluor®488 and subsequently the spread area of cells was measured. Cell differentiation was evaluated by alkaline phosphatase assay using p-nitrophenyl phosphate (pNPP) as a substrate; the reaction was stopped by NaOH, and the absorbance was measured at 405 nm. Osteocalcin, specific bone marker was stained immunohistochemically and subsequently visualized using confocal microscopy; the fluorescence intensity was analyzed and quantified. Moreover, gene expression of osteogenic markers osteocalcin and type I collagen was evaluated by real-time reverse transcription-PCR (qRT-PCR). For statistical evaluation, One-way ANOVA followed by Student-Newman-Keuls Method was used. For qRT-PCR, the nonparametric Kruskal-Wallis Test and Dunn's Multiple Comparison Test were used. The absorbance in MTS assay was significantly higher on titanium alloy Ti6Al4V compared to beta titanium alloy Ti36Nb6Ta on days 7 and 14. Mesenchymal stem cells were well spread on both alloys, but no difference in spread area was found. No differences in alkaline phosphatase assay, fluorescence intensity of osteocalcin as well as the expression of type I collagen, and osteocalcin genes were observed. Higher expression of type I collagen compared to osteocalcin was observed for cells on both alloys. Both beta titanium alloy Ti36Nb6Ta and titanium alloy Ti6Al4V Ti36Nb6Ta supported mesenchymal stem cellsˈ adhesion, proliferation and osteogenic differentiation. Novel beta titanium alloys Ti36Nb6Ta is a promising material for bone implantation. The project was supported by the Czech Science Foundation: grant No. 16-14758S, the Grant Agency of the Charles University, grant No. 1246314 and by the Ministry of Education, Youth and Sports NPU I: LO1309.

Keywords: beta titanium, cell growth, mesenchymal stem cells, titanium alloy, implant

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