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The use of horizontal starch gel electrophoresis to separate gray squirrel blood serum was investigated. Three sources of variation in the techniques were identified: (1) minor variation within-run; (2) variation across-run; (3) variation due to the sampling technique. The frequencies of the most anodal bands of 42 gray squirrel blood serum protein electropherograms were compared among several woodlots using chi-square contingency tables. On the basis of these frequencies, no differences were found among the woodlot populations of gray squirrels examined in this study. / Master of Science

Doctor of Philosophy / Department of Chemistry / Stefan H. Bossmann / Numerous proteases are known to be necessary for cancer development and progression including Matrix Metalloproteinases (MMPs), Tissue Serine Proteases, and Cathepsins. The goal of this research is to develop a Fe/Fe₃O₄ nanoparticle-based system for clinical diagnostics, which has the potential to measure the activity of cancer-associated proteases in biospecimens. Our nanoparticle-based “light switches” for measuring protease activity, consist of fluorescent cyanine dyes which are directly attached to Fe/Fe₃O₄ nanoparticles and porphyrins that are attached to Fe/Fe₃O₄ nanoparticles via consensus sequences. The consensus (cleavage) sequences can be cleaved in the presence of the correct protease, thus releasing a fluorescent dye from the Fe/Fe₃O₄ nanoparticle resulting in highly sensitive (down to 1 x 10⁻¹⁶ mol L⁻¹ for 12 proteases), selective, and fast nanoplatforms (required time: 60 min.). Upon escape, the emission intensity of the organic dye will significantly increase, which can be detected using fluorescence spectroscopy. In order to demonstrate the potential of this new technology of early recognition of various cancers several analysis types have been used. Blood and urine samples from human cancer patients and healthy volunteers, tissue and blood serum samples from human cancer patients, and canine urine and blood serum samples are some of those types. Blood samples from human cancer patients and healthy volunteers were used to demonstrate the potential of this new technology for the early recognition of breast and lung cancers. We were able to establish several proteases with diagnostic potential for breast cancer and non-small cell lung cancer. It is very likely that different cancers will feature different “protease signatures”, meaning that different proteases will be activated, depending on the origin of cancer. This permits the diagnosis of various solid tumors at different stages. Tissue samples were collected from normal tissues, from the boundary of the tumor and from the tumor of the same person. Performed fluorescence experiments clearly indicate that tissue samples from the tumor show the highest fluorescence indicating the highest concentration of the protease. Results can be used excellently in a diagnostic system for breast cancer. Based on our results measuring protease signatures offers an inexpensive and fast approach towards early cancer diagnostics.

Made available in DSpace on 2014-10-09T12:55:37Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:05:03Z (GMT). No. of bitstreams: 0 / Dissertacao (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

Made available in DSpace on 2014-10-09T12:55:37Z (GMT). No. of bitstreams: 0 / Made available in DSpace on 2014-10-09T14:05:03Z (GMT). No. of bitstreams: 0 / A prevalência do diabetes mellitus está aumentando de forma exponencial, adquirindo características epidêmicas em vários países. Nos países em desenvolvimento, particularmente, constitui-se de um grave problema de saúde pública. Dessa forma, o desenvolvimento de novas metodologias que auxiliem no diagnóstico e controle de doenças traz uma importante contribuição não somente ao tratamento de pacientes portadores desta doença como também no estabelecimento de políticas públicas de prevenção da mesma. A presença dos metais em amostras de soro humano tem sido utilizada por diversos autores para o diagnóstico da doença. Entretanto sua interpretação constitui-se ainda um desafio. A principal razão para esta dificuldade consiste no fato de que, na maioria dos estudos encontrados na literatura, o nível de metais tanto nos pacientes como no grupo de controle estão, muitas vezes, na mesma faixa de distribuição. Neste estudo, é discutida uma nova abordagem para a interpretação dos níveis de metais em amostras de soro de indivíduos portadores de diabetes mellitus tipo 2 e a possibilidade de sua utilização no diagnóstico da doença. Foram recrutados pelo Hospital da Universidade Federal de Juiz de Fora, 86 sujeitos com diabetes mellitus tipo 2 e 67 sujeitos controle. As amostras de sangue foram colhidas para separar o soro. Os elementos Cu, Mg, Mn, Se, V e Zn foram determinados pela da técnica de espectrometria de massa de alta resolução com fonte de plasma indutivamente acoplado. A interpretação dos dados foi realizada utilizando-se as técnicas de análise discriminante, componentes principais e de cluster. O conteúdo de metais nas amostras analisadas foi concordante com os dados da literatura. No entanto, a principal conclusão deste estudo é que as concentrações de cada elemento, por si mesmas, não podem ser utilizadas no diagnóstico de diabetes mellitus. Porém, a determinação de todos os metais associados à doença e a utilização de técnicas estatísticas multivariadas podem proporcionar uma excelente alternativa para esta avaliação. / Dissertacao (Mestrado) / IPEN/D / Instituto de Pesquisas Energeticas e Nucleares - IPEN-CNEN/SP

By a tracer study, using ⁸²Br, it is demonstrated that bromine is bound to serum proteins in vivo. ⁸²Br⁻ of high specific activity was injected into rabbits and serum removed one day later. Approximately ½% of the total ⁸²Br in the serum was found to be protein-bound at this stage. The application of various separation methods (electrophoresis, bromide exchange, denaturation followed by desalting) showed that one-third of the protein-bound bromine is loosely attached whereas two-thirds are firmly bound. After partial and complete enzymatic hydrolysis the bromine was found in the amino acid fraction. On the basis of the elution pattern of the amino acids on calibrated cation exchange resin columns it is concluded that the main portion of the radioactivity appeared to be associated with 3-bromo-L-tyrosine. Little, if any, bromine was observed in the serum lipids and in the thyroxine fraction isolated from serum proteins. / Thesis / Doctor of Philosophy (PhD)

Foi investigada a existência de proteção cruzada entre bacterinas bivalentes formuladas com um de três representantes do Sorogrupo Sejroe: hardjo (bacterina A), wolffi (bacterina B) e guaricura (bacterina C), e uma estirpe do sorovar pomona, empregada por ser patogênica para hamsters (Mesocricetus auratus) e possibilitar a realização do teste de potência com desafio. Os ensaios foram efetuados em hamsters machos, comparando-se os níveis de anticorpos aglutinantes e neutralizantes, respectivamente obtidos nos testes de soroaglutinação microscópica (SAM) e inibição de leptospiras in vitro (ICL). Os animais receberam duas doses de vacinas via subcutânea; aos dez dias da segunda dose, foram inoculados com culturas não inativadas dos respectivos sorovares do Sorogrupo Sejroe. Aos 21 dias pós-infecção (d.p.i.), os animais foram sangrados, os soros (n=8) foram agrupados em pools e submetidos aos testes de SAM e ICL. O teste de potência com desafio para o sorovar pomona foi adaptado do protocolo preconizado pelo United States Department of Agriculture, mas as vacinas não foram diluídas e o esquema de imunização empregou duas aplicações de 1,0mL pela via subcutânea em intervalo de dez dias; o desafio foi efetuado aos dez dias da segunda aplicação; os óbitos por leptospirose foram registrados e, aos 21 d.p.i., os sobreviventes foram sacrificados e a condição de portadores renais foi investigada através de cultivos de tecido renal para isolamento de leptospiras. No teste de potência com o sorovar pomona, o número de doses infectantes empregado para desafio (100) situou-se dentro da faixa preconizada (10 a 100); respectivamente para as bacterinas A, B e C, as proporções de mortes por leptospirose entre os animais vacinados foram de 1/10, 0/10 e 3/10, e as de portadores renais de leptospiras entre os sobreviventes foram 2/9, 1/10 e 2/7. Os resultados do teste SAM revelaram que a bacterina A induziu reações para os sorovares hardjo e wolffi; a bacterina B, para hardjo, wolffi e guaricura, e a bacterina C, apenas para a guaricura, e do teste de ICL, que animais vacinados com as bacterinas B ou C apresentaram proteção para hardjo, wolffi e guaricura; entretanto, a bacterina A conferiu proteção apenas para wolffi. Apesar das variações no poder imunogênico segundo a estirpe de leptospira empregada para a produção das bacterinas, houve proteção cruzada entre os sorovares hardjo, wolffi e guaricura. / The existence of cross-protection among bivalent bacterins, produced with one of three leptospires belonging to Serogroup Sejroe: hardjo (bacterin A), wolffi (bacterin B) and guaricura (bacterin C), and a strain of serovar pomona (included because of its pathogenicity to hamsters and the possibility of performing potency assay with challenge), was investigated in male hamsters (Mesocricetus auratus) by comparison of agglutinating and neutralizing antibodies titers, respectively measured by microscopic agglutination (MAT) and in vitro growth inhibition (GIT) tests. All animals received two doses of bacterins by subcutaneous route; after ten days from the second dose, they were inoculated with non-inactivated cultures of respective serovars of Serogroup Sejroe. At 21 post-challenge day (p.c.d.), all animals were bled and their sera (n=8) were joined in pools and tested by MAT and GIT. The potency assay with challenge performed only with serovar pomona was modified from protocol of USDA, but vaccines were not diluted and the immunization schedule employed two 1.0 mL vaccine doses by subcutaneous route with 10?day interval; the challenge was performed after ten days from the second dose; the number of deaths due to leptospirosis was registered; at 21 p.c.d., the survivors were sacrificed and their renal carrier state was investigated by culture of renal tissue for leptospires isolation. In the potency assay with serovar pomona, the number of infectious doses employed for challenge (100 infective units) was in accordance with the recommended range (10-1,000 infective units); respectively to bacterins A, B and C, proportions of deaths due to leptospirosis among vaccinated animals were 1/10, 0/10 and 3/10, and proportions of leptospires renal carrier among survivors were 2/9, 1/10 and 2/7. Results of MAT showed that bacterin A induced reactions against serovars hardjo and wolffi; bacterin B, against hardjo, wolffi and guaricura, and bacterin C, against guaricura; results of GIT revealed that vaccinated animals with bacterins B or C presented protection against serovar hardjo, wolffi and guaricura; however, bacterin A induced protection only against wolffi. A cross?protection was observed among serovars hardjo, wolffi and guaricura, although variations exist in the immunogenic capacity according to the strain of leptospires used for the bacterins production.

Dementia is the top global public health threat of the twenty first century. Within the dementia spectrum, Alzheimer’s disease (AD) is the most common type of dementia that occurs with aging and accounts for about 60% - 80% of diagnosed cases. But currently available discoveries failed to develop disease-modifying therapies for all patients living with AD. Recent discoveries can only partially slow down cognitive decline in a small subset of patients with limited effectiveness. The heterogeneity and complexity of the pathophysiology of AD indicate that a single drug approach may not be sufficient to prevent disease onset and progression. Human umbilical cord blood cells (HUCBC) and lithium treatment have shown promise against numerous neurological conditions, including AD. Yet, they also show significant unwanted, adverse effects. To address this barrier to yield successful treatments, we employed two key modifications to these two treatment strategy. We used human umbilical cord blood derived serum (HUCBS, also labeled as CBS) rather than HUCBC. We also utilized ionic cocrystal of lithium salicylate l-proline (LISPRO, also labeled as LP) instead of usual lithium salt. Both HUCBS and LISPRO have been shown to have strong neuroprotective, anti-inflammatory properties in separate studies conducted in transgenic AD mouse models. The studies detailed herein independently investigated the effectiveness of biological (HUCBS) and pharmacological (LISPRO) approaches in modulating the pathology and cognitive impairments in AD mouse models (e.g., 5XFAD, 3XTg-AD, APPswe/PS1dE9, and Tg2576). While administration of HUCBC stimulate anti-inflammatory pathways shown in previous studies, we found that HUCBS markedly promoted neurotrophic soluble amyloid precursor protein alpha (sAPPα) through non-amyloidogenic amyloid precursor protein (APP) processing pathway compared to adult (ABS) and aged blood serum (AgBS) in Chinese hamster ovary cells expressing wild type APP (CHO/APPwt). Using chromatographic fractionation, mass-spectrometry, and targeting complement proteins in cord blood serum fraction (αCBSF), we discovered the source of sAPPα in HUCBS as C1 complement protein. Further, intraperitoneal administration of αCBSF via osmotic minipump for 6 weeks showed prevention of cognitive impairment in 5XFAD mice assessed by novel object recognition, and Y-maze test. A series of recent studies have shown that lithium can prevent both AD- and age-associated cognitive decline. But, current United States Food and Drug Administration-approved lithium pharmaceutics (carbonate and citrate forms) have a narrow therapeutic window and unstable pharmaceutics that can cause toxicity without monitoring. Here we investigated the safety, pharmacokinetics, and therapeutic efficacy of LISPRO (LP), lithium salicylate (LS), and lithium carbonate (LC) in cell culture and mouse (B6129SF2/J, Tg2576, and 3xTg-AD) models. Cytokine profiles from the brain, plasma and splenocytes demonstrated that 8-week oral treatment with LISPRO downregulates pro-inflammatory cytokines, upregulates anti-inflammatory cytokines and suppress renal cyclooxygenase 2 (COX2) expression in Tg2576 mice. Pharmacokinetic studies indicated that LISPRO provides significantly higher brain and more steady plasma lithium levels in both B6129SF2/J and transgenic Tg2576 mice compared with lithium carbonate. Oral administration of LISPRO for 28 weeks significantly reduced β-amyloid plaques and tau phosphorylation. In addition, LISPRO significantly elevated pre-synaptic (synaptophysin) and post-synaptic protein (post synaptic density protein 95) expression in brains from transgenic 3XTg-AD mice. Moreover, female APPswe/PS1dE9 mice at 4 months of age were orally treated with LP, LS, or LC for 8- to 9- months at 2.25 mmol lithium/kg/day followed by measuring body weight, internal organs’ growth, and cognitive and non-cognitive function. LISPRO treatment prevented cognitive decline compared with transgenic APPswe/PS1dE9 cohort, as shown by shorter escape latency during training and probe trials in the Morris water maze and longer contextual freezing time during fear conditioning. As expected, LISPRO treatment also reduced depression assayed by tail suspension test and irritability assessed with the touch escape test. But, lithium treatment did not alter anxiety or locomotor activity as assessed by open field, elevated plus maze or accelerated rotarod tests. Taken together, these data indicate that both biological HUCBS and pharmacological LISPRO treatment may prove to be viable effective strategy for ameliorating Alzheimer’s like pathology and cognitive impairment in preclinical models.

Savo darbe tyrėme probiotinio preparato BACTOCELLR ir prebiotinio preparato AGRIMOSR įtaką lipidų kiekio pokyčiams japoninių putpelių (Coturnix coturnix japonica) organizme. Bandymo eigoje buvo tiriami šie rodikliai: bendrųjų lipidų kiekis kraujo serume, trigliceridų kiekis kraujo serume, cholesterolio kiekis kraujo serume, lipidų kiekis raumeniniame skrandyje, lipidų kiekis kepenyse, lipidų kiekis kiaušinio trynyje, vitamino A kiekis kiaušinio trynyje, trynio masė, kiaušinio masė. Atlikus eksperimentą ir išanalizavus tyrimų duomenis nustatyta, kad probiotinio preparato Bactocell ir prebiotinio preparato Agrimos įtakoje: bendrųjų lipidų kiekis kraujo serume bandomosiose grupėse vidutiniškai didėjo nuo 11,40 % iki 15,41 %, trigliceridų kiekis kraujo serume bandomosiose grupėse vidutiniškai didėjo nuo 1,25 % iki 8,05 %, cholesterolio kiekis kraujo serume bandomosiose grupėse vidutiniškai mažėjo nuo 2,35 % iki 3,08 %, lipidų kiekis raumeniniame skrandyje bandomosiose grupėse vidutiniškai didėjo nuo 16,67 % iki 33,33 %, lipidų kiekis kepenyse bandomosiose grupėse vidutiniškai didėjo nuo 14,29 % iki 33,33 %, lipidų kiekis kiaušinių tryniuose bandomosiose grupėse vidutiniškai didėjo nuo 7,11 % iki 14,29 %, vitamino A kiekis kiaušinio trynyje bandomosiose grupėse vidutiniškai didėjo nuo 8,51 % iki 14,89 %, trynio masė bandomosiose grupėse vidutiniškai didėjo nuo 12,93 % iki 20,11 %, kiaušinio masė bandomosiose grupėse vidutiniškai didėjo nuo 3,73 % iki 11,45 %, lyginant su... [toliau žr. visą tekstą] / We investigated and presented in this paper the influence of probiotic preparation BACTOCELLR and prebiotic preparation AGRIMOSR on the lipid quantity changes in the organisms of Japanese quails. During the experiment the following readings were examined: the total lipid quantity in the blood serum, the quantity of triglicerids in the blood serum, the quantity of cholesterol in blood serum, the quantity of lipids in the muscular stomach, the quantity of lipids in the liver, the quantity of lipids in egg yolk, the quantity of vitamin A in the egg yolk, egg yolk weight, and egg weight. After the experiment and data analysis we discovered that under the influence of probiotic preparation BACTOCELLR and prebiotic preparation AGRIMOSR, in comparison with the check-group, the total quantity of lipids in the blood serum of experimental groups raised from 11,40 % to 15,41 %; the quantity of triglicerids in the blood serum of experimental groups raised from 1,25 % to 8,05 %; the quantity of cholesterol in the blood serum of experimental groups fell from 2,35 % to 3,08 %; the quantity of lipids in the muscular stomach of experimental groups raised from 16,67 % to 33,33 %, the quantity of lipids in the liver of experimental groups fell from 14,29 % to 33,33 %, the quantity of lipids in the egg yolk of experimental groups raised from 7,11 % to 14,29 %; the quantity of vitamin A in the egg yolk of experimental groups raised from 8,51 % to 14,89 %, the yolk weight of experimental groups... [to full text]

The base for rearing healthy calves is a high level of passive immunity, which protects calves in a challenging period after birth. In examined herd of the Holstein cows was evaluated the influence of selected factors on the level of passive immunization in 57 calves, expressed as total protein content in calf blood serum. The selected factors were: the quality of colostrum, the time of first intake of colostrum after birth, the weight and sex of calves. The proper level of passive immunization was found only in 33 (57, 9%) calves in the studied breed. Insufficient immunization resulted in a high mortality which reached rate of 10% in 2017. The level of the passive immunization was most affected by the quality of colostrum (P < 0, 001) and the time of first drinking after birth (P < 0, 0230). The other observed factors (calf weight and gender) were not statistically significant. Adequate care for the youngest age category is crucial and influences the profitability of the breed.