The goal of diabetes education programs (DEPs) is to improve patients' pattern-management habits for those with type 2 diabetes (T2D), though participation in DEPs is lower than expected due to low physician referral rates. This retrospective study examined secondary data of 162 T2D patients who had been referred to a DEP in a community center in RI to determine whether the source of referral affected patient attendance, participation persistence, and outcomes. Self-referred (n = 62) and physician-referred (n = 100) groups were analyzed for possible associations among the aforementioned variables. Chi-square (p = .04) and logistic regression (p = .04) indicated that the referral source for a DEP does have an effect on participation rates, while logistic regression showed that odds for self-referred patients to participate were 1.97 times higher. Multiple linear regression found no difference between the referral source and the number of sessions patients completed, though covariate analysis showed that age (p = .02) contributes to the model. Multiple linear regression showed no difference between the number of sessions attended and changes in hemoglobin A1c (HbA1c) levels. It is important to note that those who completed the program and reported pre- and post-program HbA1c levels (n = 7) all reported improved outcomes. This highlights the limitation of the small sample size (n = 7), which increased the possibility of a Type II error. This community center DEP model can serve as a blueprint, highlighting the importance of diabetes education and leading to positive social change by improving referral and participation rates and resulting in fewer complications, a decreased disease burden, and an improved quality of life.

Several studies have been conducted that link poor control of hemoglobin A1c (HbA1c) to an increased risk of heart disease. However, there are limited published studies that link HbA1c and heart disease based on ethnicity and gender. To address this gap in literature, the purpose of this study was to assess the association between HbA1c and heart disease in African American males (aged 30-64 years old) while controlling for education, income, and access to care. The research questions were focused on establishing an association between HbA1c values and heart disease in African American men through the lens of the health belief model. Secondary data (N=243) were used from the Nutritional Health and Examination Survey (2011-2016) and analyzed. Chi-square analysis and multiple logistic regression were conducted to test for an association between HbA1c and heart disease in men aged 30-64 years old. The variables of Hba1c values, various forms of heart disease, and stroke were tested while controlling for age, education, income, and access to care. Key results indicated no association between HbA1c and heart disease; yet, it is recommended that future research on the topic should include a larger sample of those with heart disease, from other sources, to better assess the outcome. The positive social change implications include the addition of research related to gender-specific outcomes and ethnicity-related risk between HbA1c and heart disease, which can be used to achieve better disease management and provide educational opportunities for diabetic African-American men.

THE TRYPANOLYTIC FACTOR OF HUMAN SERUM: A TROJAN HORSE African trypanosomes, the prototype of which is Trypanosoma brucei, are protozoan parasites of huge clinical, veterinary and economical importance. They develop in the body fluids of various mammals (including humans) where they face and manipulate many different aspects of the immune system. The extent of this interplay is pivotal to both host and parasite survival, and depending on parasite virulence and host susceptibility, infection duration ranges from some months to several years. At the end, host survival is invariably compromised. Humans and few other primates provide however a striking exception to this fatal outcome. They are indeed fully protected against most trypanosome infections through the presence in their blood of a so-called trypanosome lytic factor (TLF). The TLF is known to circulate mainly in the form of a high density lipoprotein particle characterized by the simultaneous presence of two primate-specific proteins: haptoglobin-related protein (Hpr) and apolipoprotein L-I (apoL-I). We have contributed to delineate the respective roles played by Hpr and apoL-I in the lysis process. ApoL-I was shown to be the exclusive toxin of the TLF. In its absence humans get fully susceptible to any trypanosome infection. The toxin was shown to kill the parasite after endocytosis through the generation of ionic pores in the lysosomal membrane. Those pores dissipate membrane potential and trigger the influx of chloride ions from the cytoplasm into the lysosomal compartment, leading to an eventually fatal uncontrolled osmotic phenomenon. ApoL-I efficient delivery to the parasite relies on Hpr. African trypanosomes indeed fulfil their heme nutritional requirements by receptor-mediated internalization of the complex formed by haptoglobin, an evolutionary conserved acute-phase protein, and hemoglobin, resulting from physiological intravascular hemolysis. This heme uptake by the auxotrophic parasites contributes to both growth rate and resistance against host oxidative burst. In human serum, the trypanosome receptor is unable to discriminate between Hp and the closely related TLF-bound Hpr, explaining TLF efficient endocytosis. As such, the TLF acts as a Trojan horse, killing the parasite from inside the cell after having deceived its vigilance through the high similarity between heme-delivering haptoglobin and toxin-associated Hpr.

Nature has always served as a model for mimicking and inspiration to humans in their efforts to improve their life. Researchers have been inspired by nature to produce biomimetic materials with molecular recognition properties by design rather than evolution. Molecular imprinting is one way to prepare such materials. Such smart materials with new functionalities are at the forefront of the development of a relevant number of ongoing and perspective applications ranging from consumer to space industry. Molecularly imprinted polymers were developed by mimicking the natural enzymes or antibodies that serve as host for binding target molecules. These imprints were used as a recognition element to substitute natural biomolecules in biosensors. The concept behind molecular imprinting is to mold a material (with the desired chemical properties) around individual molecules. Upon removal of the molecular templates, one is left with regions in the molded material that fit the shape of the template molecules. Thus, molecular imprinting results in materials that can selectively bind to molecules of interest. Imprinted materials resulted in applications ranging from chemical separation to bioanalytics. In this work attempts were made particularly in the development of molecularly imprinted polymer based thermometric sensors. The main effort was focused towards the development of an covalently imprinted polymer that would be able to selectively bind fructosyl valine (Fru-Val), the N-terminal constituent of hemoglobin A1c ß-chains. Taking into account the known advantages of imprinted polymers, e.g. robustness, thermal and chemical stability, imprinted materials were successfully used as a recognition element in the sensor. One of the serious problems associated with the development of MIP sensors and which lies in the absence of a generic procedure for the transformation of the polymer-template binding event into a detectable signal has been addressed by developing the "thermometric" approach. In general the developed approach gives a new insight on MIP/Analyte interactions. / In dem Bestreben, ihr eigenes Leben zu verbessern, haben die Menschen stets die Natur nachgeahmt und sich von ihr inspirieren lassen. Die Natur hat Forscher zur Erzeugung smarter biomimetischer Stoffe mit molekularen Erkennungseigenschaften nach dem Vorbild der Evolution inspiriert. Eine der Methoden zur Herstellung solcher Substanzen ist das molekulare Prägen. Smarte Materialien mit neuen Eigenschaften stehen an der Spitze der Entwicklung potentieller Anwendungen vom Verbraucher bis hin zur Raumfahrtindustrie. Durch Nachahmung von natürlichen Enzymen oder Antikörpern wurden molekular geprägte Polymere (MIPs) entwickelt, die der Bindung von Zielmolekülen dienen. Diese geprägten Polymere (imprints) wurden anstelle von Biomolekülen als Erkennungselemente in Biosensoren eingesetzt. Das Konzept, das dem molekularen Prägen zugrunde liegt, besteht in der Formung eines Polymers (mit den entsprechenden chemischen Eigenschaften) um einzelne Zielmoleküle herum. Nach Entfernen dieser molekularen Template bleiben Abdrücke im Polymer übrig, die der Form der Templatmoleküle entsprechen. Mit Hilfe des molekularen Prägens kann man also Stoffe herstellen, die sich selektiv an bestimmte Moleküle binden können. Geprägte Polymere finden breite Anwendung, etwa in chemischen Aufreinigungsprozessen und der Bioanalytik. Hauptanliegen der vorliegenden Arbeit war es, thermometrische Sensoren auf der Basis molekular geprägter Polymere zu entwickeln. Die Anstrengungen richteten sich vor allem auf die Entwicklung eines kovalent geprägten Polymers, das in der Lage ist, selektiv Fruktosyl-Valin (Fru-Val), den N-terminalen Bereich von Hämoglobin A1c, zu binden. Aufgrund der bekannten Vorzüge geprägter Polymere – z. B. Robustheit und thermische und chemische Stabilität – wurden geprägte Polymere erfolgreich als Erkennungselement im Sensor angewendet. Eine der größten Herausforderungen bei der Entwicklung von MIP-Sensoren, das Fehlen eines generischen Verfahrens zur Umwandlung der Bindungsreaktion in ein nachweisbares Signal, wurde mit der Entwicklung der thermometrischen Methode in Angriff genommen. Diese Methode führt allgemein zu neuen Einsichten in die Interaktionen zwischen MIP und Analyt.

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The need for an alternative to red cells in transfusions has led to the creation of hemoglobin-based oxygen carriers (HBOCs). However, evaluations of all products tested in clinical trials have noted cardiovascular complications, raising questions about their safety that led to the abandonment of all those products. It has been considered that the adverse side effects come from the scavenging of the vasodilator – nitric oxide (NO) by the deoxyheme sites of the hemoglobin derivatives. Another observation is that HBOCs with lower oxygen affinity than red cells release oxygen prematurely in arterioles, triggering an unwanted homeostatic response. Since the need for such a product remains critical, it is important to understand the reactivity patterns that contribute to the observed complications. Various alterations of the protein have been attempted in order to reduce HBOC-induced vasoconstriction. Recent reports suggest that a safe and effective product should be pure, homogenous and have a high molecular weight along with appropriate oxygenation properties. While these properties are clearly important, vasodilatory features of hemoglobin through its nitrite reductase activity may also act as an in situ source of NO. It follows that HBOCs with an enhanced ability to produce NO from endogenous nitrite may serve to counteract vasoactivity associated with NO-scavenging by hemoglobin. Here we characterize the effects of different protein modifications on the nitrite reductase activity of hemoglobin. We produced a variety of HBOCs that include cross-linked tetramers, polyethylene glycol (PEG) conjugates and bis-tetramers of hemoglobin. We report that the rate of NO production strongly depends on the conformational state of the protein, with R-state stabilized proteins (PEG-Hbs), exhibiting the fastest rates. In particular, we found that PEGylated bis-tetramers of hemoglobin (BT-PEG) exhibit increased nitrite reductase activity while retaining cooperativity and stability. Animal studies of BT-PEG demonstrated that this material is benign: it did not cause significant increases in systemic blood pressure in mice, the major side effect associated with existing HBOCs. BT-PEG exhibits an enhanced nitrite reductase activity together with sample purity and homogeneity, molecular size and shape, and appropriate oxygenation properties, characteristics of a clinically useful HBOC.

The need for an alternative to red cells in transfusions has led to the creation of hemoglobin-based oxygen carriers (HBOCs). However, evaluations of all products tested in clinical trials have noted cardiovascular complications, raising questions about their safety that led to the abandonment of all those products. It has been considered that the adverse side effects come from the scavenging of the vasodilator – nitric oxide (NO) by the deoxyheme sites of the hemoglobin derivatives. Another observation is that HBOCs with lower oxygen affinity than red cells release oxygen prematurely in arterioles, triggering an unwanted homeostatic response. Since the need for such a product remains critical, it is important to understand the reactivity patterns that contribute to the observed complications. Various alterations of the protein have been attempted in order to reduce HBOC-induced vasoconstriction. Recent reports suggest that a safe and effective product should be pure, homogenous and have a high molecular weight along with appropriate oxygenation properties. While these properties are clearly important, vasodilatory features of hemoglobin through its nitrite reductase activity may also act as an in situ source of NO. It follows that HBOCs with an enhanced ability to produce NO from endogenous nitrite may serve to counteract vasoactivity associated with NO-scavenging by hemoglobin. Here we characterize the effects of different protein modifications on the nitrite reductase activity of hemoglobin. We produced a variety of HBOCs that include cross-linked tetramers, polyethylene glycol (PEG) conjugates and bis-tetramers of hemoglobin. We report that the rate of NO production strongly depends on the conformational state of the protein, with R-state stabilized proteins (PEG-Hbs), exhibiting the fastest rates. In particular, we found that PEGylated bis-tetramers of hemoglobin (BT-PEG) exhibit increased nitrite reductase activity while retaining cooperativity and stability. Animal studies of BT-PEG demonstrated that this material is benign: it did not cause significant increases in systemic blood pressure in mice, the major side effect associated with existing HBOCs. BT-PEG exhibits an enhanced nitrite reductase activity together with sample purity and homogeneity, molecular size and shape, and appropriate oxygenation properties, characteristics of a clinically useful HBOC.

Due to the technology development, the diet habit has completely changed. It accompanied by the metabolite diseases relevant to blood glucose and lipids, which are dependent with the atherosclerosis and cardiovascular disease. In this study, we using matrix assisted laser desorption ionization time of flight mass spectrometry (MALDI-TOF/MS) to characterize triglycerides in human plasma. In the other, the glycosylated hemoglobin in human whole blood was detected by liquid electrospray laser desorption ionization (Liquid ELDI/MS). Triglycerides are energy source (9 kcal/g) in human body, derived from glycerol and three fatty acids. It is a main constituent of vegetable oil and animal fats. In clinical diagnosis, human plasma was mixed with triglyceride Kit to react to the final 520 nm UV-absorbing substance, then the concentration was quantified consistent with the calibration line by UV/Visible spectrometry. By the way, it needed Kit chemicals for one trial. MALDI-TOF/MS is a simple and easy method to operate to detect complex compounds in human plasma, only need to optimize the parameters (solvent collection, sample dilution, matrix selection, sample pretreatment ) to form a homogeneous crystals. The developed ¡§seed layer¡¨ method can reduce the sweet spot effect and cause a lower with-in spot variation (RSD < 20%) compared to ¡§premix¡¨ method (RSD >30%). Combined with statistic software 2D peak distribution, a semi-quantification can be observe of 24 different triglyceride concentration human plasmas. The level of glycosylated hemoglobin (HbA1c) in whole blood is currently the most important measurement of long-term control of the glycemic state of diabetes. As a result of the interferences of high concentrations of metabolites, proteins and salts in whole blood, tedious sample cleanup procedures must be performed prior to subjecting the sample solutions to conventional LC/MS and MALDI analyses for the detection of HbA1c. Electrospray laser desorption ionization mass spectrometry (ELDI/MS), a two-step ambient ionization technique, has been developed to characterize analytes directly from the liquid sample surface. One drop of the diluted hole blood (1/10, v/v in water) was placed on the stainless steel plate. The sample droplet was irradiated with a pulse laser, the desorbed analytes were post-ionized in an electrospray (ESI) plume (ESI solution: 70% methanol in water, 0.1% acetic acid), and the analyte ions were detected by a ion trap mass analyzer. Through this study, the protocol for efficiently characterizing HbA1c present in a drop of diluted whole blood with ELDI/MS was established. We successfully detected the ion signal of HbA1c with ELDI/MS. Quantification of the level of HbA1c in the whole blood of diabetic patients was achieved by calculating the ratio of the ion peak area of the glycosylated and non-glycosylated hemoglobin ions. A linear relationship exists for the quantitative results of HbA1c in whole blood of 20 diabetic patients obtained between ELDI/MS and that through conventional spectroscopic measurement.

Chapter 1. Deciphering the toxicity of Helicobacter pylori clinical isolates from gastric diseases patients using MALDI-TOF/MS and multivariate analysis. Helicobacter pylori (H. pyloyi) infection is associated with gastric diseases such as gastric polyp, chronic gastritis, gastric ulcer, gastric cancer, etc. In fact, most of the people infected not have the symptoms of gastric diseases due to the high degree of variability of gene with H. pyloyi and the specific immune responses of the hosts. In order to investigate the relationship between H.pylori and gastric diseases, the clinical strains of H. pylori isolated from patients from nine gastric diseases were extracted from the optimized extraction and analysis by MALDI-TOF/MS, then the high reproducible spectra were combined with multivariate statistical analysis including Principal Component Analysis (PCA), Hierarchical Cluster Analysis (HCA), Discriminant Analysis (DA) . In the result of PCA, there is no specific potential marker to discriminate the clinical strains to nine gastric diseases. In the result of HCA, the strains from different gastric diseases were clustered together means they have the similarity of the protein and metabolite. In the result of DA, the strains from gastric and non-gastric cancer were discriminanted by the discriminant function composed of thirty-eight discriminant variables in the spectra. This discriminant function would be confirmed by other clinical strains isolated from gastric diseases patients in the future and then would help to predict the the similarity of the protein and metabolite of the strains isolated from the gastric diseases patients whether gastric cancer or not. Chapter 2. Biomarker discovery in the stools of gastric cancer patients using MALDI-TOF/MS. According to the statistics of Republic 100 years from the Department of Health, cancer was the first of the ten lesding to death. With the modern change of eatiog habbits, gastrointestinal cancer has increased steadily. Gastrointestinal cancer accompanied occult gastrointestinal bleeding, and it is commonly detected by the fecal occult blood test (FOB). FOB including Guaiac-based fecal occult-blood test and immunochemical tests. Guaiac-based fecal occult-blood tests make use of the pseudoperoxidase activity of heme, and the reagent turns blue after oxidation by oxidants or peroxidases in the presence of an oxygen donor such as hydrogen peroxide, so it would have the potential of false-positive result. Immunochemical tests, which use antibodies detect against human hemoglobin with great sensitivity, but the tests are limited by loss of hemoglobin antigenicity at room temperature and require processing in a laboratory. In order to decrease the false-positive of detecting heme and decreasing the cost of the detection against hemoglobin in stools, in the study, we ues the distill water to extract the heme (m/z 616) and hemoglobin in stools and analysis with the reflectron and linear mode of MALDI-TOF/MS. In this study, at first, we used the stimulated stomach acid decomposing the hemoglobin to release the heme, to stimulate the gastrointestinal bleeding. Second, we used the distill water to extract the hemoglobin in stools, and detected by the linear mode of MALDI-TOF/MS, and the detection limit of MALDI-TOF/MS against hemoglobin in stool was better than the immunochemical tests. Third, the same strategy was applied to fifty-nine patients (including nineteen esophageal cancer patients, twenty gastric cancer patients and colorectal cancer patients) stools to detect heme and hemoglobin by MALDI-TOF/MS and the results were compared with the fecal occult blood test. In the detection of heme, MALDI-TOF/MS had not detect heme, but the Guaiac-based fecal occult-blood test had detected, it would be that the stools had the oxidants (not heme) to react the reagent. In addition, MALDI-TOF/MS had detected heme, but the Guaiac-based fecal occult-blood test had no results, those cases would be catched up in the future. In the detection of hemoglobin, using immunochemical tests to be the reference index, MALDI-TOF/MS had the false-negative result might come from the complicated matrix effect of stools, so that the hemoglobin could not form the good crystalline with matrix CHCA. The false-positive results of MALDI-TOF/MS might come from the criteria of hemoglobin signal.

published_or_final_version / Nursing Studies / Master / Master of Philosophy