Search results for: bovine macrophages

Authors: Raheela Akhtar, Zafar Iqbal Chaudhary, Yongqun Oliver He, Muhammad Younus, Aftab Ahmad Anjum

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Brucella abortus is an intracellular pathogen affecting macrophages. Macrophages release some components such as lysozymes (LZ), reactive oxygen species (ROS) and reactive nitrite intermediates (RNI) which are important tools against intracellular survival of Brucella. The antibrucella activity of bovine and murine macrophages was compared following stimulation with Brucella abortus lipopolysaccharides. Our results revealed that murine macrophages were ten times more potent to produce antibrucella components than bovine macrophages. The differential production of these components explained the differential Brucella killing ability of these species that was measured in terms of intramacrophagic survival of Brucella in murine and bovine macrophages.

Keywords: bovine macrophages, Brucella abortus, cell stimulation, cytokines, Murine macrophages

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Authors: Abdul Matin, Salik Nawaz, Suk-Yul Jung

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Balamuthia mandrillaris is well known to cause fatal Balamuthia amoebic encephalitis (BAE). Amoebic transmission into the central nervous system (CNS), haematogenous spread is thought to be the prime step, followed by blood-brain barrier (BBB) dissemination. Macrophages are considered to be the foremost line of defense and present in excessive numbers during amoebic infections. The aim of the present investigation was to evaluate the effects of macrophages alone or primed with cytokines on the biological characteristics of Balamuthia in vitro. Using human brain microvascular endothelial cells (HBMEC), which constitutes the BBB, we have shown that Balamuthia demonstrated > 90% binding and > 70% cytotoxicity to host cells. However, macrophages further increased amoebic binding and Balamuthia-mediated cell cytotoxicity. Furthermore, macrophages exhibited no amoebicidal effect against Balamuthia. Zymography assay demonstrated that macrophages exhibited no inhibitory effect on proteolytic activity of Balamuthia. Overall, to our best knowledge, we have shown for the first time macrophages has no inhibitory effects on the biological properties of Balamuthia in vitro. This also strengthened the concept that how and why Balamuthia can cause infections in both immuno-competent and immuno-compromised individuals.

Keywords: Balamuthia mandrillaris, macrophages, cytokines, human brain microvascular endothelial cells, Balamuthia amoebic encephalitis

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Authors: Mayada Mazher, Ahmed Moustafa, Ahmed Abdellatif

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Background: Autophagy is a eukaryotic, highly conserved catabolic process implicated in many pathophysiologies such as wound healing. Autophagy-associated genes serve as a scaffolding platform for signal transduction of macrophage polarization during the inflammatory phase of wound healing and tissue repair process. In the current study, we report a model for the interplay between autophagy-associated genes and macrophages polarization associated genes. Methods: In silico analysis was performed on 249 autophagy-related genes retrieved from the public autophagy database and gene expression data retrieved from Gene Expression Omnibus (GEO); GSE81922 and GSE69607 microarray data macrophages polarization 199 DEGS. An integrated protein-protein interaction network was constructed for autophagy and macrophage gene sets. The gene sets were then used for GO terms pathway enrichment analysis. Common transcription factors for autophagy and macrophages' polarization were identified. Finally, microRNAs enriched in both autophagy and macrophages were predicated. Results: In silico prediction of common transcription factors in DEGs macrophages and autophagy gene sets revealed a new role for the transcription factors, HOMEZ, GABPA, ELK1 and REL, that commonly regulate macrophages associated genes: IL6,IL1M, IL1B, NOS1, SOC3 and autophagy-related genes: Atg12, Rictor, Rb1cc1, Gaparab1, Atg16l1. Conclusions: Autophagy and macrophages' polarization are interdependent cellular processes, and both autophagy-related proteins and macrophages' polarization related proteins coordinate in tissue remodelling via transcription factors and microRNAs regulatory network. The current work highlights a potential new role for transcription factors HOMEZ, GABPA, ELK1 and REL in wound healing.

Keywords: autophagy related proteins, integrated network analysis, macrophages polarization M1 and M2, tissue remodelling

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Authors: Mohd Sukri Hassan, Ahlam Inayatullah Badrul Munir, M. Husaini A. Rahman

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Halal authentication and verification in supplement capsules are highly required as the gelatine available in the market can be from halal or non-halal sources. It is an obligation for Muslim to consume and use the halal consumer goods. At present, real-time polymerase chain reaction (RT-PCR) is the most common technique being used for the detection of porcine and bovine DNA in gelatine due to high sensitivity of the technique and higher stability of DNA compared to protein. In this study, twenty samples of supplements capsules from different products with different Halal logos were analyzed for porcine and bovine DNA using RT-PCR. Standard bovine and porcine gelatine from eurofins at a range of concentration from 10^-1 to 10^-5 ng/µl were used to determine the linearity range, limit of detection and specificity on RT-PCR (SYBR Green method). RT-PCR detected porcine (two samples), bovine (four samples) and mixture of porcine and bovine (six samples). The samples were also tested using FT-IR technique where normalized peak of IR spectra were pre-processed using Savitsky Golay method before Principal Components Analysis (PCA) was performed on the database. Scores plot of PCA shows three clusters of samples; bovine, porcine and mixture (bovine and porcine). The RT-PCR and FT-IR with chemometrics technique were found to give same results for porcine gelatine samples which can be used for Halal authentication.

Keywords: halal, real-time PCR, gelatine, chemometrics

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Authors: Ahlam Inayatullah Badrul Munir, M. Husaini A. Rahman, Mohd Sukri Hassan

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Real-time PCR is a molecular biology technique that is currently being widely used for halal services to differentiating between porcine and bovine DNA. The useful of technique become very important for student or workers (who works in the laboratory) to learn how the technique could be run smoothly without fail. Same concept with conventional PCR, real-time PCR also needed DNA template, primer, enzyme polymerase, dNTP, and buffer. The difference is in real-time PCR, have additional component namely fluorescent dye. The most common use of fluorescent dye in real-time PCR is SYBR green. The purpose of this study was to find out how sensitive, specific and efficient real-time PCR technique was combined with SYBR green method and specific primers of CYT b. The results showed that real-time PCR technique using SYBR Green, capable of detecting porcine and bovine DNA concentrations up to 0.0001 µl/ng. The level of efficiency for both types of DNA was 91% (90-110). Not only that in specific primer CYT b bovine primer could detect only bovine DNA, and porcine primer could detect only porcine primer. So, from the study could be concluded that real-time PCR technique that was combined with specific primer CYT b and SYBR green method, was sensitive, specific and efficient to detect porcine and bovine DNA.

Keywords: sensitivity, specificity, efficiency, real-time PCR, SYBR green, Cytochrome b, porcine DNA, bovine DNA

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Authors: Berrin Şentürk

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In this study, due to the recurrence of bovine tuberculosis, in the same areas, the risk factors for the disease were determined and evaluated at the local level. This study was carried out in 32 farms where the disease was detected in the district and center of Samsun province in 2014. Predetermined risk factors, such as farm, environmental and economic risks, were investigated with the survey method. It was predetermined that risks in the three groups are similar to the risk variables of the disease on the global scale. These risk factors that increase the susceptibility of the infection must be understood by the herd owners. The risk-based contagious disease management system approach should be applied for bovine tuberculosis by farmers, animal health professionals and public and private sector decision makers.

Keywords: bovine tuberculosis, disease management, control, outbreak, risk analysis

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Authors: Kidane Workelul, Yohans Tekle, Guesh Negash, Haftay Abraha, Nigus Abebe Shumuye, Yisehak Tsegaye Redda

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Bovine papillomatosis (wart) is one of the economically important bovine skin diseases worldwide, caused by a group of viruses named papillomaviruses (PVs). However, it has often been misdiagnosed as other skin diseases and remained untreated. In order to determine the status of the diseases, twenty-two farms were visited, and fourteen infected cattle with cutaneous papillomatosis were identified from a total of 235. Papilloma biopsies were taken for molecular and histopathological characterization, the therapeutic trial of an autogenous vaccine was evaluated on infected animals. The overall status of bovine papillomatosis in this study was calculated as 5.96% (14/235). The disease was found to be statistically significant in the age groups less than two years (X² = 26.69, P = 0.0001). The more prominent histologically characterized lesions in the sampled tissue were identified as squamous papilloma and fibro-papilloma. The Polymerase Chain Reaction (PCR) based identification revealed that all the clinically and histo-pathologically characterized papillomatosis cases were found to be infected with Bovine Papilloma Virus1(BPV1), indicating that BPV1 was the most common and sole causative agent of the diseases in the study area. In immunizing active bovine papillomatosis, an autogenous vaccine therapeutic trial demonstrated excellent results, with practically full recovery and no recurrence of the infection. Hence, it is concluded that bovine papillomatosis is an economically important disease of young age group cattle as well as a treatable disease. So, the production of marketable autogenous vaccines against bovine papillomatosis should be started and given at an early stage.

Keywords: autogenous vaccine, bovine papillomatosis, bovine papilloma virus1 clinical-pathology, polymerase chine reaction, wart

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Authors: Daniel M. Barry

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Since more than half a century ago, attempts have been made to culture cells and embryos outside the body (in vitro or ex vivo). This was done with different culture media and in various “incubators”. In the present study two different culture media were used: a standard TCM-199 culture medium and first trimester amniotic fluid (BAF) collected sterilely from pregnant cows after slaughter. Two different culture conditions were also investigated, the standard laboratory CO2 incubator versus culturing bovine embryos in the vagina of a goat doe. Two experiments were done: Firstly the permeability of different receptacles to CO2 gas was analyzed for possible culture in the vagina. Four-well plates and straws were used to incubate TCM-199 and BAF for a period of 120 h in the presence or absence of 5% CO2 gas. The pH values were measured and recorded every 24 h. In the second experiment pre-compacted morula stage bovine embryos were cultured in the above culture media in sealed 0.25 mL straws in a standard laboratory incubator and in the vagina of a goat doe. Evaluation was done on (1) stage of development and (2) number of blastomeres after 96 h of culture. In the first experiment it was shown that the CO2 gas diffused out of the 4-well plate as well as through the wall of the straws in the absence of CO2 gas, while in the presence of CO2 the pH of both media stabilized between 7.3 and 7.5. This meant that the semen straws were permeable to CO2 gas and could therefore be used as receptacles for culturing early stage bovine embryos. In the second experiment no statistical differences (p>0.05) were found in the number of pre-compacted bovine embryos that developed to the blastocyst stage, or the hatched blastocyst stage, neither for the culture medium used, or the method of culturing in the two incubators. Neither was there any difference (p>0.05) in the number of blastomeres that developed at the blastocyst stage between the two types of incubators. The bovine embryos tended to develop more blastomeres when cultured in BAF than when cultured in TCM-199 in both the standard laboratory incubator and when using the vagina of a goat doe as an incubator.

Keywords: alternative culture, bovine embryos, vagina, bovine amniotic fluid, incubator

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Authors: Hainan Chen, Jina Qing, Xiao Zhu, Ling Gao, Ampadu O. Jackson, Min Zhang, Kai Yin

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Objective: Editing macrophage activation to dampen inflammatory diseases by promoting the repolarization of inflammatory (M1) macrophages to anti-inflammatory (M2) macrophages is highly associated with mitochondrial respiration. Recent studies have suggested that mitochondrial apolipoprotein A-1 binding protein (APOA1BP) was essential for the cellular metabolite NADHX repair to NADH, which is necessary for the mitochondrial function. The exact role of APOA1BP in the repolarization of M1 to M2, however, is uncertain. Material and method: THP-1-derived macrophages were incubated with LPS (10 ng/ml) or/and IL-4 (100 U/ml) for 24 hours. Biochemical parameters of oxidative phosphorylation and M1/M2 markers were analyzed after overexpression of APOA1BP in cells. Results: Compared with control and IL-4-exposed M2 cells, APOA1BP was downregulated in M1 macrophages. APOA1BP restored the decline in mitochondrial function to improve metabolic and phenotypic reprogramming of M1 to M2 macrophages. Blocking oxidative phosphorylation by oligomycin blunts the effects of APOA1BP on M1 to M2 repolarization. Mechanistically, LPS triggered the hydration of NADH and increased its hydrate NADHX which inhibit cellular NADH dehydrogenases, a key component of electron transport chain for oxidative phosphorylation. APOA1BP decreased the level of NADHX via converting R-NADHX to biologically useful S-NADHX. The mutant of APOA1BP aspartate188, the binding site of NADHX, fail to repair oxidative phosphorylation, thereby preventing repolarization. Conclusions: Restoring mitochondrial function by increasing mitochondrial APOA1BP might be useful to improve the reprogramming of inflammatory macrophages into anti-inflammatory cells to control inflammatory diseases.

Keywords: inflammatory diseases, macrophage repolarization, mitochondrial respiration, apolipoprotein A-1 binding protein, NADHX, NADH

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Authors: Roman Marsalek

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Preparation of nano-particles of cerium oxide and adsorption of bovine serum albumine on them were studied. Particle size distribution and influence of pH on zeta potential of prepared CeO2 were determined. Average size of prepared cerium oxide nano-particles was 9 nm. The simultaneous measurements of the bovine serum albumine adsorption and zeta potential determination of the (adsorption) suspensions were carried out. The adsorption isotherms were found to be of typical Langmuir type; values of the bovine serum albumin adsorption capacities were calculated. Increasing of pH led to decrease of zeta potential and decrease of adsorption capacity of cerium oxide nano-particles. The maximum adsorption capacity was found for strongly acid suspension (am=118 mg/g). The samples of nanoceria with positive zeta potential adsorbed more bovine serum albumine on the other hand, the samples with negative zeta potential showed little or no protein adsorption. Surface charge or better say zeta potential of CeO2 nano-particles plays the key role in adsorption of proteins on such type of materials.

Keywords: adsorption, BSA, cerium oxide nanoparticles, zeta potential, albumin

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Authors: K. Nikovics, D. Riccobono, M. Oger, H. Morin, L. Barbier, T. Poyot, X. Holy, A. Bendahmane, M. Drouet, A. L. Favier

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Muscle regeneration after injury (as irradiation) is of great importance. However, the molecular and cellular mechanisms are still unclear. Cytokines are believed to play fundamental role in the different stages of muscle regeneration. They are secreted by many cell populations, but the predominant producers are macrophages and helper T cells. On the other hand, it has been shown that adipose tissue derived stromal/stem cell (ASC) injection could improve muscle regeneration. Stem cells probably induce the coordinated modulations of gene expression in different macrophage cells. Therefore, we investigated the patterns and timing of changes in gene expression of different cytokines occurring upon stem cells loading. Muscle regeneration was studied in an irradiated muscle of minipig animal model in presence or absence of ASC treatment (irradiated and treated with ASCs, IRR+ASC; irradiated not-treated with ASCs, IRR; and non-irradiated no-IRR). We characterized macrophage populations by immunolabeling in the different conditions. In our study, we found mostly M2 and a few M1 macrophages in the IRR+ASC samples. However, only few M2b macrophages were noticed in the IRR muscles. In addition, we found intensive fibrosis in the IRR samples. With in situ hybridization and immunolabeling, we analyzed the cytokine expression of the different macrophages and we showed that M2d macrophage are the most abundant in the IRR+ASC samples. By in situ hybridization, strong expression of the transforming growth factor β (TGF-β) was observed in the IRR+ASC but very week in the IRR samples. But when we analyzed TGF-β level with immunolabeling the expression was very different: many M2 macrophages showed week expression in IRR+ASC and few cells expressing stronger level in IRR muscles. Therefore, we investigated the MMP expressions in the different muscles. Our data showed that the M2 macrophages of the IRR+ASC muscle expressed MMP2 proteins. Our working hypothesis is that MMP2 expression of the M2 macrophages can decrease fibrosis in the IRR+ASC muscle by capturing TGF-β.

Keywords: adipose tissue derived stromal/stem cell, cytokine, macrophage, muscle regeneration

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Authors: Ekaterina Zubkova, Irina Beloglazova, Iurii Stafeev, Konsyantin Dergilev, Yelena Parfyonova, Mikhail Menshikov

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Mesenchymal stromal cells from adipose tissue (MSC) currently are widely used in regenerative medicine to restore the function of damaged tissues, but that is significantly hampered by their heterogeneity. One of the modern approaches to overcoming this obstacle is the polarization of cell subpopulations into a specific phenotype under the influence of cytokines and other factors that activate receptors and signal transmission to cells. We polarized MSC with factors affecting the inflammatory signaling and functional properties of cells, followed by verification of their expression profile and ability to affect the polarization of macrophages. RT-PCR evaluation showed that cells treated with LPS, interleukin-17, tumor necrosis factor α (TNF α), primarily express pro-inflammatory factors and cytokines, and after treatment with polyninosin polycytidic acid and interleukin-4 (IL4) anti-inflammatory factors and some proinflammatory factors. MSC polarized with pro-inflammatory cytokines showed a more robust pro-angiogenic effect in fibrin gel bead 3D angiogenesis assay. Further, we evaluated the possibility of paracrine effects of MSCs on the polarization of intact macrophages. Polarization efficiency was assesed by expression of M1/M2 phenotype markers CD80 and CD206. We showed that conditioned media from MSC preincubated in the presence of IL-4 cause an increase in CD206 expression similar to that observed in M2 macrophages. Conditioned media from MSC polarized in the presence of LPS or TNF-α increased the expression of CD80 antigen in macrophages, similar to that observed in M1 macrophages. In other cases, a pronounced paracrine effect of MSC on the polarization of macrophages was not detected. Thus, our study showed that the polarization of MSC along the pro-inflammatory or anti-inflammatory pathway allows us to obtain cell subpopulations that have a multidirectional modulating effect on the polarization of macrophages. (RFBR grants 20-015-00405 and 18-015-00398.)

Keywords: angiogenesis, cytokines, mesenchymal, polarization, inflammation

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Authors: Chaoran Liu

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Bioactive polysaccharides and polysaccharide-protein complex derived from mushrooms and fungi have a wide range of immunomodulatory activity with low side-effects and have therefore the potential to be developed as an adjuvant in cancer therapies. Mushrooms sclerotium is rich in polysaccharides and the polysaccharides isolated from the sclerotium of Polyporus rhinocerus have shown potent in vivo and in vitro immunomodulatory effects. Macrophages are considered to be an important component of the innate immune response against bacterial infection and cancer. To better understanding the immunomodulatory effects and its underlying mechanisms of sclerotial water-soluble polysaccharides extracted from P. rhinocerus on macrophages, the objectives of this study are to purify the water-soluble novel sclerotial polysaccharides and to characterize the structure and properties as well as to study the detailed molecular mechanisms of the in vitro immunomodulating effects in murine macrophages. The hot water-soluble fraction PRW from the sclerotium of P. rhinocerus was obtained using solvent extraction. PRW was further fractionated by membrane ultrafiltration to a give a fraction (PRW1) with molecular mass less than 50 kDa. PRW1 was characterized to be a polysaccharide-protein complex composed of 45.7% polysaccharide and 44.2% protein. The chemical structure of the carbohydrate moiety of PRW1 was elucidated by GC and FTIR to be mainly beta-D-glucan with trace amount of galactose and mannose. The immunomodulatory effects of PRW1 on murine RAW 264.7 macrophages were demonstrated in terms of the increase in nitric oxide production and cytokine production. Mechanistically, PRW1 initiates ERK phosphorylation to activate macrophages within 15 min and significantly improves the expression level of inducible NOS (iNOS) from 6 h after treatment. In summary, this study indicates that PRW1 is a potent immunomodulatory agent for macrophages and suggests that mushroom sclerotia from Polyporus rhinocerus requires for further investigation in cancer research.

Keywords: Polyporus rhinocerus, mushroom sclerotia, Polysaccharide-Protein Complex, macrophage activation

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Authors: Hamid Mustafa, Huson J. Heather, Kim Eiusoo, Adeela Ajmal, Tad S. Sonstegard

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The objective of this study was to evaluate the informativeness of Bovine high density SNPs genotyping in Sahiwal cattle population. This is a first attempt to assess the Bovine HD SNP genotyping array in any Pakistani indigenous cattle population. To evaluate these SNPs on genome wide scale, we considered 777,962 SNPs spanning the whole autosomal and X chromosomes in Sahiwal cattle population. Fifteen (15) non related gDNA samples were genotyped with the bovine HD infinium. Approximately 500,939 SNPs were found polymorphic (MAF > 0.05) in Sahiwal cattle population. The results of this study indicate potential application of Bovine High Density SNP genotyping in Pakistani indigenous cattle population. The information generated from this array can be applied in genetic prediction, characterization and genome wide association studies of Pakistani Sahiwal cattle population.

Keywords: Sahiwal cattle, polymorphic SNPs, genotyping, Pakistan

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Authors: H. Jeries, N. Volkova, C. G. Iglesias, M. Najjar, M. Rosenblat, M. Aviram, T. Hayek

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Background: Synthetic forms of glucocorticoids (e.g., prednisone, prednisolone) are anti-inflammatory drugs which are widely used in clinical practice. The role of glucocorticoids (GCs) in cardiovascular diseases including atherosclerosis is highly controversial, and their impact on macrophage foam cell formation is still unknown. Our aim was to investigate the effects of prednisone or its active metabolite, prednisolone, on macrophage oxidative stress and lipid metabolism using in-vivo, ex-vivo and in-vitro systems. Methods: The in-vivo study included C57BL/6 mice which were intraperitoneally injected with prednisone or prednisolone (5mg/kg) for 4 weeks, followed by lipid metabolism analyses in the mice aorta, and in peritoneal macrophages (MPM). In the ex-vivo study, we analyzed the effect of serum samples obtained from 9 healthy volunteers before or after treatment with oral prednisone (20mg for 5 days), on J774A.1 macrophage atherogenicity. In-vitro studies were conducted using J774A.1 macrophages, human monocyte derived macrophages (HMDM) and fibroblasts. Cells were incubated with increasing concentrations (0-200 ng/ml) of prednisone or prednisolone, followed by determination of cellular oxidative status, triglyceride and cholesterol metabolism. Results: Prednisone or prednisolone treatment resulted in a significant reduction in triglycerides and mainly in cholesterol cellular accumulation in MPM or in J774A.1 macrophages incubated with human serum. Similar resulted were noted in HMDM or in J774A.1 macrophages which were directly incubated with the GCs. These effects were associated with GCs inhibitory effect on triglycerides and cholesterol biosynthesis rates, throughout downregulation of diacylglycerol acyltransferase1 (DGAT1) expression, and of the sterol regulatory element binding protein (SREBP2) and HMGCR expression, respectively. In parallel to prednisone or prednisolone induced reduction in macrophage triglyceride content, paraoxonase 2 (PON2) expression was significantly upregulated. GCs-induced reduction of cellular triglyceride and cholesterol mass was mediated by the GCs receptors on macrophages since the GCs receptor antagonist (RU 486) abolished these effects. In fibroblasts, unlike macrophages, prednisone or prednisolone showed no anti-atherogenic effects. Conclusions: Prednisone or prednisolone are anti-atherogenic since they protected macrophages from lipid accumulation and foam cell formation.

Keywords: atherosclerosis, cholesterol, foam cell, macrophage, prednisone, prednisolone, triglycerides

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Authors: Wenxiao Liu, Kun Zhang, Yongqing Li

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Bovine herpesvirus 1, a member of the genus Variellovirus of the subfamily Alphaherpesvirinae, has caused severe economic cost to the bovine industry. In this study, BoHV-1 glycerol protein gD was expressed in insect cells, and the purified gD was immunized in the Balb/C mice to generate monoclonal antibodies. Based on hybridoma cell fusion techniques, 20 monoclonal antibodies against Bovine herpesvirus 1 have been obtained. Further, mAb 3F8 with neutralizing activity and gD were applied to develop a blocking enzyme-linked immunosorbent assay (Elisa) for detecting neutralizing antibodies against BoHV-1, which shows a significant correlation between the blocking Elisa and VNT. The sensitivity and specificity of the test were estimated to be 94.59% and 93.42%, respectively. Furthermore, antibody pairing tests revealed that mAb 1B6 conjugated to fluorescence microspheres was used as the capture antibody, and mAb 3F9 was used as the detectable antibody to establish the immunochromatographic assay (ICS). The ICS was conducted to detect BoHV-1 in bovine samples with high sensitivity, specificity, and good stability. Clinical sample testing revealed that the results of ICS and real-time PCR have a coincidence rate of 95.42%. Our research confirmed that the ICS is a rapid and reliable method for the diagnosis of BoHV-1. In conclusion, our results lay a solid foundation for the prevention and control of BoHV-1 infection.

Keywords: bovine disease, BoHV-1, ELISA, ICS assay

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Authors: Emily Simango, T. Chitura

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Background: Bovine brucellosis is among the most neglected zoonotic diseases in developing countries, where it is endemic and a growing challenge to public health. The development of cost-effective control measures for the disease can only be affirmed by the knowledge of the disease epidemiology and the ability to define its risk profiles. The aim of the study was to document the trend of bovine brucellosis and the control measures adopted following reported cases during the period 2014 to 2019 in South Africa. Methods: Data on confirmed cases of bovine brucellosis was retrieved from the website of the World Organisation of Animal Health (WOAH). Data was analysed using the Statistical Package for Social Sciences (IBM SPSS, 2022) version 29.0. Descriptive analysis (frequencies and percentages) and the Analysis of variance (ANOVA) were utilized for statistical significance (p<0.05). Results: The data retrieved in our study revealed an overall average bovine brucellosis prevalence of 8.48. There were statistically significant differences in bovine brucellosis prevalence across the provinces for the years 2016 and 2019 (p≥0.05), with the Eastern Cape Province having the highest prevalence in both instances. Documented control measures for the disease were limited to killing and disposal of disease cases as well as vaccination of susceptible animals. Conclusion: Bovine brucellosis is real in South Africa, with the risk profiles differing across the provinces. Information on brucellosis control measures in South Africa, as reported to the WOAH, is not comprehensive.

Keywords: zoonotic, endemic, Eastern Cape province, vaccination

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Authors: M. N. Boukhatem, M. A. Ferhat, K. Mansour

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Bovine raw milk represents a favorable environment for the growth of several food-spoilage strains and some pathogens. It must meet stringent standards to ensure the highest microbiological and toxicological qualities.In order to assess the microbiological risks associated with the consumption of this food, we conducted this study to determine the microbiological quality of bovine raw milk (54 samples) commercialized at the state of Blida (Algeria). The samples analyzed were unsatisfactory in terms of total flora where 61.11% of samples were considered as non acceptable in terms of quality standards, fecal coliforms (40.74%), fecal streptococci (55.55%) and staphylococci (74.07%). Salmonella and Clostridium strains were not detected in all the samples. Furthermore, antibiotic residues were found in 26% of analysed samples. These results reflect non-compliance with the rules of good hygiene practices at milking, storage, transportatio, and sale of milk. Bovine raw milk consumed presents a serious health risk to the population of the study areas.The livestock coaching actors and dissemination of good hygiene practices throughout the production chain are needed to improve the quality of local milk.

Keywords: bovine raw milk, microbiological quality, fecal coliforms, antibiotic residue, Blida state

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Authors: Hernandez Pardo Diego F., Guiza Arguello Viviana R., Coy Echeverria Ana, Viejo Abrante Fernando

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The biological hydroxyapatite obtained from bovine bone arouses great interest in its application as a material for bone regeneration due to its better bioactive behavior in comparison with synthetic hydroxyapatite. For this reason, the objective of the present investigation was to determine the effect of chemical and thermal treatments in obtaining biological bovine hydroxyapatite of high purity and crystallinity. Two different chemical reagents were evaluated (NaOH and HCl) with the aim to remove the organic matrix of the bovine cortical bone. On the other hand, for analyzing the effect of thermal treatment temperature was ranged between 500 and 1000°C for a holding time of 4 hours. To accomplish the above, the materials before and after the chemical and thermal treatments were characterized by elemental compositional analysis (CHN), infrared spectroscopy by Fourier transform (FTIR), RAMAN spectroscopy, scanning electron microscopy (SEM), thermogravimetric analysis (TGA) and X-ray diffraction (XRD) and energy dispersion X-ray spectroscopy (EDS). The results allowed to establish that NaOH is more effective in the removal of the organic matrix of the bone when compared to HCl, whereas a thermal treatment at 700ºC for 4 hours was enough to obtain biological hydroxyapatite of high purity and crystallinity.

Keywords: bovine bone, hydroxyapatite, biomaterials, thermal treatment

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Authors: Fernanda Gonçalves, Karla Alcântara, Vanessa Moura, Patrícia Nolasco, Jorge Kalil, Maristela Hernandez

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Introduction: Atherosclerosis is a chronic disease where two striking features are observed: retention of lipids and inflammation. Understanding the interaction between immune cells and lipoproteins involved in atherogenesis are urgent challenges, since cardiovascular diseases are the leading cause of death worldwide. Macrophages are critical to the development of atherosclerotic plaques and in the perpetuation of inflammation in these lesions. These cells are also directly involved in unstable plaque rupture. Recently different populations of macrophages are being identified in atherosclerotic lesions. Although the presence of M2 macrophages (macrophages activated by the alternative pathway, eg. The IL-4) has been identified, the function of these cells in atherosclerosis is not yet defined. M2 macrophages have a high endocytic capacity, they promote remodeling of tissues and to have anti-inflammatory activity. However, in atherosclerosis, especially unstable plaques, severe inflammatory reaction, accumulation of cellular debris and intense degradation of the tissue is observed. Thus, it is possible that the M2 macrophages have altered function (phenotype) in atherosclerosis. Objective: Our aim is to evaluate if the presence of oxidized LDL alters the phenotype and function of M2 macrophages in vitro. Methods: For this, we will evaluate whether the addition of lipoprotein in M2 macrophages differentiated in vitro with IL -4 induces 1) a reduction in the secretion of anti-inflammatory cytokines (CBA and ELISA), 2) secretion of inflammatory cytokines (CBA and ELISA), 3) expression of cell activation markers (Flow cytometry), 4) alteration in gene expression of molecules adhesion and extracellular matrix (Real-Time PCR) and 5) Matrix degradation (confocal microscopy). Results: In oxLDL stimulated M2 macrophages cultures we did not find any differences in the expression of the cell surface markers tested, including: HLA-DR, CD80, CD86, CD206, CD163 and CD36. Also, cultures stimulated with oxLDL had similar phagocytic capacity when compared to unstimulated cells. However, in the supernatant of these cultures an increase in the secretion of the pro-inflammatory cytokine IL-8 was detected. No significant changes where observed in IL-6, IL-10, IL-12 and IL-1b levels. The culture supernatant also induced massive extracellular matrix (produced by mouse embryo fibroblast) filaments degradation. When evaluating the expression of 84 extracellular matrix and adhesion molecules genes, we observed that the stimulation of oxLDL in M2 macrophages decreased 47% of the genes and increased the expression of only 3% of the genes. In particular we noted that oxLDL inhibit the expression of 60% of the genes constituents of extracellular matrix and collagen expressed by these cells, including fibronectin1 and collagen VI. We also observed a decrease in the expression of matrix protease inhibitors, such as TIMP 2. On the opposite, the matricellular protein thrombospondin had a 12 fold increase in gene expression. In the presence of native LDL 90% of the genes had no altered expression. Conclusion: M2 macrophages stimulated with oxLDL secrete the pro-inflammatory cytokine IL-8, have an altered extracellular matrix constituents gene expression, and promote the degradation of extracellular matrix. M2 macrophages may contribute to the perpetuation of inflammation in atherosclerosis and to plaque rupture.

Keywords: atherosclerosis, LDL, macrophages, m2

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Authors: Nermin Isik, Ozlem Derinbay Ekici, Oguzhan Avci

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The aim of this study was to determination of role infection in neonatal calves in Central Anatolian, Turkey. A total 300 fecal samples were collected from diarrheic neonatal calves, aged between 0–90 days from Konya, Karaman, and Aksaray from January to April 2014. Fecal specimens from calves with clinically diarrheic symptoms were examined for the presence of Bovine Coronavirus, Bovine Rotavirus, Cryptosporidium sp., and E. coli by commercially available capture direct enzyme linked immunosorbent assay (ELISA) kit and Modified Ziehl Neelsen method (MZN). Calves were grouped according to their age as follows: 1-14, 15-29, and 30-90 days. Cryptosporidium sp. infection was detected in 52.8%, 58.8%, and 39.2% by ELISA and 33.9%, 47%, 26.7% by MZN in the respective age groups. The seroprevalance of Rotavirus (12.5 %, 40 %, 12.5 %), Coronavirus (2.5%, 0%, 3.5%) and E. coli (5%, 4.7%, 8.9%) infections were determined according to the age groups respectively. Cryptosporidium sp. was the most detected enteropathogen (52 %) of calves and coronavirus was the least detected (2 %). The detection rate of the mixed enfection was 12.3%. In conclusion, it must be evaluated by mix infections in calves with diarrhea. These results will provide an important contribution against the factors that cause diarrhea

Keywords: cryptosporidium sp., bovine coronavirus, bovine rotavirus, E.coli, calve, ELISA

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Authors: Huseyin A. Yavasoglu, Yusuf Ziya Tengiz, Ali Göksenli

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In this study, an automatic feeding machine for different type and class of bovine animals is designed. Daily nutrition of a bovine consists of grass, corn, straw, silage, oat, wheat and different vitamins and minerals. The amount and mixture amount of each of the nutrition depends on different parameters of the bovine. These parameters are; age, sex, weight and maternity of the bovine, also outside temperature. The problem in a farm is to constitute the correct mixture and amount of nutrition for each animal. Faulty nutrition will cause an insufficient feeding of the animal concluding in an unhealthy bovine. To solve this problem, a new automatic feeding machine is designed. Travelling of the machine is performed by four tires, which is pulled by a tractor. The carrier consists of eight bins, which each of them carries a nutrition type. Capacity of each unit is 250 kg. At the bottom of each chamber is a sensor measuring the weight of the food inside. A funnel is at the bottom of each chamber by which open/close function is controlled by a valve. Each animal will carry a RFID tag including ID on its ear. A receiver on the feeding machine will read this ID and by given previous information by the operator (veterinarian), the system will detect the amount of each nutrition unit which will be given to the selected animal for feeding. In the system, each bin will open its exit gate by the help of the valve under the control of PLC (Programmable Logic Controller). The amount of each nutrition type will be controlled by measuring the open/close time. The exit canals of the bins are collected in a reservoir. To achieve a homogenous nitration, the collected feed will be mixed by a worm gear. Further the mixture will be transported by a help of a funnel to the feeding unit of the animal. The feeding process can be performed in 100 seconds. After feeding of the animal, the tractor pulls the travelling machine to the next animal. By the help of this system animals can be feeded by right amount and mixture of nutrition

Keywords: bovine, feeding, nutrition, transportation, automatic

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Authors: Vijaya Madhuri Devraj, Swarnalatha Guditi, Kiran Kumar Bokara, Gangadhar Taduri

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Cell-based strategies may open therapeutic approaches that promote tolerance through manipulation of macrophages to increase long-term transplant survival rates and minimize side effects of the current immune suppressive regimens. The aim of the present study was, therefore, to test and compare the therapeutic potential of MSC and Tregs on macrophage polarization to develop an alternate cell-based treatment option in kidney transplantation. In the current protocol, macrophages from kidney transplant recipients with graft dysfunction were co-cultured with MSCs and Treg cells with and without cell-cell contact on transwell plates, further to quantitatively assess macrophage polarization in response to MSC and Treg treatment over time, M1 and M2 cell surface markers were used. Additionally, multiple soluble analytes were analyzed in cell supernatant by using bead-based immunoassays. Furthermore, to confirm our findings, gene expression analysis was done. MSCs induced the formation of M2 macrophages more than Tregs when macrophages M0 were cultured in transwell without cell contact. From this, we deduced the mechanism that soluble factors present in the MSCs condition media are involved in skewing of macrophages towards type 2 macrophages; similarly, in co-culture with cell-cell contact, MSCs resulted in more M2 type macrophages than Tregs. And an important finding of this study is the combination of both MSC-Treg showed significantly effective and consistent results in both with and without cell contact setups. Hence, it is suggestive to prefer MSCs over Tregs for secretome-based therapy and a combination of both for either therapy for effective transplantation outcomes. Our findings underline a key role of Tregs and MSCs in promoting macrophage polarization towards anti-inflammatory type. The study has great importance in prolongation of allograft and patient survival without any rejection by cell-based therapy, which induce self-tolerance and controlling infection.

Keywords: graft rejection, graft tolerance, macrophage polarization, mesenchymal stem cells, regulatory T cells, transplant immunology

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Authors: Sulhee Lee, Geon Kim, Young-Seo Park

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Unlike the traditional milk sterilization methods (LTLT, HTST, or UHT), pulsed electric field (PEF) technology is a non-thermal pasteurization process. This technology minimizes energy required for heat treatment in food processing, changes in sensory properties, and physical losses. In this study, structural changes of bovine milk proteins, the amount of immunoproteins such as IgG, and their storability by PEF treatment were examined. When the changes of protein content in PEF-treated milk were examined using HPLC, the amounts of α-casein and β-lactoglobulin were reduced over 40% each, whereas those of κ-casein and β-casein did not change. The amount of α-casein in HTST milk was reduced to 50%. When PEF was applied to milk at the energy level of 250 kJ, the amounts of IgG, IgA, β-lactoglobulin (β-LG), lactoferrin, and α-lactalbumin (α-LA) decreased by 43, 41, 35, 63, and 45%, respectively. When milk was sterilized by LTLT process followed by PEF process at the level of 150 kJ, the concentrations of IgG, IgA, β-LG, lactoferrin, and α-LA were 56.6, 10.6, 554, 2.8 and 660.1 μg/mL, respectively. When the bovine milk was sterilized by LTLT process followed by PEF process at the energy level of 180 kJ, storability of immunoproteins of milk was the highest and the concentrations of IgG, IgA, and β-LG decreased by 79.5, 6.5, and 134.5 μg/mL, respectively, when compared with the initial concentrations of those proteins. When bovine milk was stored at 4℃ after sterilization through HTST sterilizer followed by PEF process at the energy level of 200 kJ, the amount of lactoferrin decreased 7.3% after 36 days of storage, whereas that of lactoferrin of raw milk decreased 16.4%. Our results showed that PEF treatment did not change the protein structure nor induce protein denaturation in milk significantly when compared with LTLT or HTST sterilization. Also, LTLT or HTST process in combination with PEF were more effective than LTLT only or HTST only process in the conservation of immunoproteins in bovine milk.

Keywords: pulsed electric field, bovine milk, immunoproteins, sterilization

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Authors: Sachin K. Samuchiwal, Barbara Balestrieri, Amanda Paskavitz, Hannah Raff, Joshua A. Boyce

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IL-33, a recently discovered member of the IL-1 cytokine family, binds to the TLR/IL1R super family receptor ST2 and induces type 2 immune responses. IL-33 is constitutively expressed in structural cells at barrier sites such as skin, lung, and intestine, and also inducibly expressed by hematopoietic cells including macrophages. Stimulation of macrophages by Lipopolysaccharide (LPS) can induce de novo IL-33 expression, and also causes the production of prostaglandin-E2 (PGE2) via cyclooxygenase (COX)-2 and microsomal PGE2 synthase-1 (mPGES-1). Because PGE2 can regulate macrophage functions through both autocrine and paracrine mechanisms, the potential interplay of endogenous PGE2 on IL-33 production was explored. Bone-marrow derived murine macrophages (bmMF) that lack either mPGES-1 or EP2 receptor expression were stimulated with LPS in the absence or presence of exogenous PGE2 along with pharmacological agonists and antagonists. The study results demonstrate that endogenous PGE2 markedly enhances LPS-induced IL-33 production by bmMFs via EP2 receptors. Moreover, exogenous PGE2 can amplify LPS-induced IL-33 expression dominantly by EP2 and partly by EP4 receptors by a pathway involving cAMP and exchange protein activated by cAMP (EPAC), but not protein kinase A (PKA). Though both IL-33 production and PGE2 generation in response to LPS require activation of both p38 MAPK and NF-κB, PGE2 did not influence this activation. In conclusion, it is demonstrated that endogenous PGE2 signaling through EP2 and EP4 receptors is a prerequisite for LPS-induced IL-33 production in bmMFs and the underlying cAMP mediated pathway involves EPAC. Since IL-33 is a critical pro-inflammatory cytokine in various pathological disorders, this PGE2-EP2/EP4-cAMP mediated pathway can be exploited to intervene in IL-33 driven pathologies.

Keywords: bone marrow macrophages, EPAC, IL-33, PGE2

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Authors: Natapol Pumipuntu

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Staphylococcus argenteus is the emerging species of S. aureus complex. It was generally misidentified as S. aureus by standard techniques and their features. S. argenteus is possibly emerging in both humans and animals, as well as increasing worldwide distribution. The objective of this study was to differentiate and identify S. argenteus from S. aureus, which has been collected and isolated from milk samples of subclinical bovine mastitis cases in Maha Sarakham province, Northeastern of Thailand. Twenty-one isolates of S. aureus, which confirmed by conventional methods and immune-agglutination method were analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) and multilocus sequence typing (MLST). The result from MALDI-TOF MS and MLST showed 6 from 42 isolates were confirmed as S. argenteus, and 36 isolates were S. aureus, respectively. This study indicated that the identification and classification method by using MALDI-TOF MS and MLST could accurately differentiate the emerging species, S. argenteus, from S. aureus complex which usually misdiagnosed. In addition, the identification of S. argenteus seems to be very limited despite the fact that it may be the important causative pathogen in bovine mastitis as well as pathogenic bacteria in food and milk. Therefore, it is very necessary for both bovine medicine and veterinary public health to emphasize and recognize this bacterial pathogen as the emerging disease of Staphylococcal bacteria and need further study about S. argenteus infection.

Keywords: Staphylococcus argenteus, subclinical bovine mastitis, Staphylococcus aureus complex, mass spectrometry, MLST

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Authors: Andrei-Marius Sandu

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This study aims to analyze the bovine meat and milk sector for the period 2007-2013. For the period analyzed, it is known that Romania has benefited from EU funding through the National Rural Development Programme 2007-2013. In this programme, there were measures that addressed exclusively the animal husbandry sector in Romania. This paper presents data on bovine production of meat, milk and livestock in Romania, but also data on the price and impact the European Funds implementation had on them.

Keywords: European funds, measures, national rural development programme, price

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Authors: Eduardo Kanagushiku Pereira, Frank Gregory Cavalcante da Silva, Barbara Soares Gonçalves, Ana Lúcia Bergamasco Galastri, Ronei Luciano Mamoni

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The inflammatory response initiates after the recognition of pathogens by receptors expressed by innate immune cells. Among these receptors, the NLRP3 was associated with the recognition of pathogenic fungi in experimental models. NLRP3 operates forming a multiproteic complex called inflammasome, which actives caspase-1, responsible for the production of the inflammatory cytokines IL-1beta and IL-18. In this study, we aimed to investigate the involvement of NLRP3 in the inflammatory response elicited in macrophages against Paracoccidioides brasiliensis (Pb), the etiologic agent of PCM. Macrophages were differentiated from THP-1 cells by treatment with phorbol-myristate-acetate. Following differentiation, macrophages were stimulated by Pb yeast cells for 24 hours, after previous treatment with specific NLRP3 (3,4-methylenedioxy-beta-nitrostyrene) and/or caspase-1 (VX-765) inhibitors, or specific inhibitors of pathways involved in NLRP3 activation such as: Reactive Oxigen Species (ROS) production (N-Acetyl-L-cysteine), K+ efflux (Glibenclamide) or phagossome acidification (Bafilomycin). Quantification of IL-1beta and IL-18 in supernatants was performed by ELISA. Our results showed that the production of IL-1beta and IL-18 by THP-1-derived-macrophages stimulated with Pb yeast cells was dependent on NLRP3 and caspase-1 activation, once the presence of their specific inhibitors diminished the production of these cytokines. Furthermore, we found that the major pathways involved in NLRP3 activation, after Pb recognition, were dependent on ROS production and K+ efflux. In conclusion, our results showed that NLRP3 participates in the recognition of Pb yeast cells by macrophages, leading to the activation of the NLRP3-inflammasome and production of IL-1beta and IL-18. Together, these cytokines can induce an inflammatory response against P. brasiliensis, essential for the establishment of the initial inflammatory response and for the development of the subsequent acquired immune response.

Keywords: inflammation, IL-1beta, IL-18, NLRP3, Paracoccidioidomycosis

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Authors: Anjali Roy, Mirza S. Baig

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Atherosclerosis is a chronic inflammatory disease characterized by the formation of lipid rich plaque enriched with necrotic core, modified lipid accumulation, smooth muscle cells, endothelial cells, leucocytes and macrophages. Macrophage foam cells play a critical role in the occurrence and development of inflammatory atherosclerotic plaque. Foam cells are the fat-laden macrophages in the initial stage atherosclerotic lesion formation. Foam cells are an indication of plaque build-up, or atherosclerosis, which is commonly associated with increased risk of heart attack and stroke as a result of arterial narrowing and hardening. The mechanisms that drive atherosclerotic plaque progression remain largely unknown. Dissecting the molecular mechanism involved in process of macrophage foam cell formation will help to develop therapeutic interventions for atherosclerosis. To investigate the mechanism, we studied the role of nitric oxide synthase 1(NOS1)-mediated nitric oxide (NO) on low-density lipoprotein (LDL) uptake by bone marrow derived macrophages (BMDM). Using confocal microscopy, we found that incubation of macrophages with NOS1 inhibitor, TRIM (1-(2-Trifluoromethylphenyl) imidazole) or L-NAME (N omega-nitro-L-arginine methyl ester) prior to LDL treatment significantly reduces the LDL uptake by BMDM. Further, addition of NO donor (DEA NONOate) in NOS1 inhibitor treated macrophages recovers the LDL uptake. Our data strongly suggest that NOS1 derived NO regulates LDL uptake by macrophages and foam cell formation. Moreover, we also checked proinflammatory cytokine mRNA expression through real time PCR in BMDM treated with LDL and copper oxidized LDL (OxLDL) in presences and absences of inhibitor. Normal LDL does not evoke cytokine expression whereas OxLDL induced proinflammatory cytokine expression which significantly reduced in presences of NOS1 inhibitor. Rapid NOS-1-derived NO and its stable derivative formation act as signaling agents for inducible NOS-2 expression in endothelial cells, leading to endothelial vascular wall lining disruption and dysfunctioning. This study highlights the role of NOS1 as critical players of foam cell formation and would reveal much about the key molecular proteins involved in atherosclerosis. Thus, targeting NOS1 would be a useful strategy in reducing LDL uptake by macrophages at early stage of disease and hence dampening the atherosclerosis progression.

Keywords: atherosclerosis, NOS1, inflammation, oxidized LDL

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Authors: Blanca Fatela Cantillo, Silvia Iglesias Cerrato, Guadalupe Garrido Ceca

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Purpose: Bleb dysfunction is a late complication following glaucoma filtration surgery. We describe our surgical technique for excision and reconstruction of a hypertrophic bleb complication using bovine pericardium patch graft (Tutopatch®) and amniotic membrane. Material and methods: The case report presents a hypertrophic bleb over the cornea with good intraocular pressure control. The hanging bleb without leak caused dysesthesia and high irregular astigmatism. Bleb reconstruction involved the excision of corneal fibrous material and avascular conjunctiva, preserving the original scleral and tennon. Bovine pericardium patch graft (Tutopatch®) was sited over these with fixed sutures, reinforcing the underlying scleral, and the conjunctiva advanced. The superior epithelium corneal defect was covered using an amniotic membrane. Conclusion: Repair of bleb dysfunction with varied techniques has been reported, including conjunctival advancement, use of scleral patch graft, dural patch graft, or pericardium. Additional use of amniotic membrane promotes epithelialization and exhibits anti-fibrotic and anti-inflammatory features. Reconstruction with bovine pericardium patch graft and amniotic membrane resulted in pain relief, visual rehabilitation, and good aesthetic results, with preservation of bleb function.

Keywords: reconstruction, hypertrophic bleb, bovine pericardium, amniotic membrane, dysesthesia of the bleb

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