Cytochrome P450 enzymes and oxidative stress in tobacco/nicotine mediated HIV pathogenesis

Title from PDF of title page, viewed on June 8, 2016Dissertation advisor: Santosh KumarVitaIncludes bibliographical references (pages 147-161)Thesis (Ph.D.)--School of Pharmacy. University of Missouri--Kansas City, 2016In the US, the prevalence of cigarette smoking in the HIV-infected population is 50-70% compared to 15-20% in the general population, which further increases the risk of smoking-related health problems in this group. For example, HIV-infected smokers show decreased immune responses, poorer responses to antiretroviral therapy (ART), and greater risk of virological rebound, compared to HIV-infected non-smokers. Several in vivo and in vitro studies have shown that smoking/nicotine is associated with decreased immune responses, increased inflammation, increased oxidative stress, and increased numbers of opportunistic infections. Furthermore, in vitro studies have shown that smoking/nicotine enhances HIV replication in alveolar macrophages, microglia, and T cells. However, the mechanism by which smoking or nicotine increases HIV replication is largely unknown. As an exception, a report suggests that iron and oxidative stress are possible mechanisms of enhanced production of HIV by alveolar macrophages in cigarette smokers. The role of CYP enzymes has not been studied in context with smoking/nicotine and HIV pathogenesis. However, there are several elegant studies that show the involvement of CYP2A6, CYP2A13, CYP1A1, and CYP1B1 in smoking/nicotine-mediated toxicity leading to various types of cancers and hepatic toxicity. The majority of tobacco constituents, including nicotine, are metabolized and/or activated by CYP enzymes to generate reactive oxygen species (ROS) and/or reactive metabolites, NNK. Based on the fact that CYP pathways play a critical role in smoking mediated cell/organ toxicity, there is a critical need to explore the involvement of CYP pathways in smoking/nicotine-mediated HIV pathogenesis. The present study was based on the central hypothesis that tobacco/nicotine, or its metabolites, enhance HIV replication in monocytes/macrophages through CYP pathway. The key findings of our studies presented in this thesis indicate that (a) CYP2A6-mediated metabolism of nicotine increased the generation of ROS in HIV cell model, SVGA astrocytes, (b) Cigarette smoke condensate caused significant induction of CYP1A1, CYP2A6, ROS production, and cytotoxicity in U937 monocytes as well as enhanced HIV replication in HIV-infected primary macrophages, (c) Mild-to-moderate smoking increased viral load in HIV-infected individuals. Furthermore, our study suggested that smoking and HIV independently increase oxidative stress in the plasma as well as in monocytes. (d) There was a decrease in the level of nicotine and subsequent increase in the level of nicotine metabolites, suggesting an increase in nicotine metabolism in HIV-infected smokers compared with uninfected smokers. In conclusion, our in vitro and ex vivo results are consistent with the hypothesis that CYP and CYP-mediated oxidative stress by tobacco/nicotine are associated with increased viral load by smoking/smoking constituents. This study has clinical implications in terms of targeting CYP and oxidative stress pathways to find potentially novel therapeutic interventions, as well as drug dose adjustment to treat HIV-infected smokers effectively.Tobacco smoking mediated oxidative stress in organ toxicities: role of cytochrome P450 systems -- Tobacco smoking effect on HIV pathogenesis: role of cytochrome P450 isozymes -- General material and methods -- An LC-MS method for concurrent determination of nicotine metabolites and the role of CYP2A6 in nicotine metabolite mediated oxidative stress in SVGA astrocytes -- Cigarette smoke consensate (CSC) mediated oxidative stress and cytoxicity in U937 monocytes -- Effect of mild-tomoderate smoking on viral load, cytokines, oxidative stress, and cytochrome P450 enzymes in HIV-infected individuals -- Enhanced nicotine metabolism in HIV-positive smokers compared to HIV-negative smokers: simultaneous determination of nicotine and its four metabolites in their plasma using a simple and sensitive ESI-LC-MS/MS technique -- Summary and future direction

Enhanced oxidative stress by alcohol use in HIV+ patients: possible involvement of cytochrome P450 2E1 and antioxidant enzymes

BACKGROUND: Alcohol consumption is prevalent amongst HIV positive population. Importantly, chronic alcohol use is reported to exacerbate HIV pathogenesis. Although alcohol is known to increase oxidative stress, especially in the liver, there is no clinical evidence that alcohol increases oxidative stress in HIV positive patients. The mechanism by which alcohol increases oxidative stress in HIV positive patients is also unknown. METHODS: To examine the effects of alcohol use on oxidative stress we recruited HIV+ patients who reported mild-to-moderate alcohol use. Strict inclusion and exclusion criteria were applied to reduce the effect of other therapeutic drugs metabolized via the hepatic system as well as the effect of co-morbidities such as active tuberculosis on the interaction between alcohol and HIV infection, respectively. Blood samples were collected from HIV-negative alcohol-users and HIV positive alcohol-users followed by collection of plasma and isolation and fractionation of monocytes from peripheral blood. We then determined oxidative DNA damage, glutathione level, alcohol level, transcriptional level of cytochrome P450 2E1 (CYP2E1) and several antioxidant enzymes, and plasma level of cytokines. RESULTS: Compared to HIV-negative alcohol users, HIV-positive alcohol users demonstrated an increase in oxidative DNA damage in both plasma and CD14+ monocytes, as well as, a relative increase in oxidized/reduced glutathione (GSSG/GSH) in plasma samples. These results suggest an increase in oxidative stress in HIV-positive alcohol users compared with HIV-negative alcohol users. We also examined whether alcohol metabolism, perhaps by CYP2E1, and antioxidant enzymes are involved in alcohol-mediated increased oxidative stress in HIV-positive patients. The results showed a lower plasma alcohol level, which was associated with an increased level of CYP2E1 mRNA in monocytes, in HIV-positive alcohol users compared with HIV-negative alcohol users. Furthermore, the transcription of major antioxidants enzymes (catalase, SOD1, SOD2, GSTK1), and their transcription factor, Nrf2, were reduced in monocytes obtained from HIV positive alcohol users compared to the HIV-negative alcohol user group. However, no significant change in levels of five major cytokines/chemokines were observed between the two groups. CONCLUSIONS: The data suggests that alcohol increases oxidative stress in HIV+ patients, perhaps through CYP2E1- and antioxidant enzymes-mediated pathways. The enhanced oxidative stress is accompanied by a failure of cellular antioxidant mechanisms to maintain redox homeostasis. Overall, the enhanced oxidative stress in monocytes may exacerbate HIV pathogenesis in HIV positive alcohol users

Stelleninhaber geht – Wissen bleibt!

In Deutschland nimmt der Anteil älterer Arbeitnehmerinnen und Arbeitnehmer tendenziell zu. Deshalb muss sich die Bibliotheksleitung verstärkt auf das altersbedingte Ausscheiden älterer Arbeitnehmer einstellen. Eine langjährige Fachkraft verfügt über spezielles Erfahrungswissen im direkten Aufgabenfeld. Die Bibliotheksleitung muss den Transfer allen relevanten Wissens, dazu gehört das Erfahrungswissen, vom Stelleninhaber auf seinen Nachfolger ermöglichen und unterstützen. Am Beispiel der Universitätsbibliothek der Bergakademie Freiberg wird untersucht, wie das Wissensmanagement im Rahmen eines Stellenwechsels derzeit geregelt ist. Das geschieht mit Hilfe von Tiefeninterviews in verschiedenen Abteilungen. Die Auswertung der Interviews bildet die Basis für ein Konzept für das Wissensmanagement beim Stellenwechsel an der UB Freiberg. Das Konzept benennt u. a. Maßnahmen zur Identifikation des stellenbezogenen Wissens, Maßnahmen zur Dokumentation des relevanten Wissens und Instrumente zur Wissensweitergabe beim Stellenwechsel

Design and Development of IoT-Enabled Medicine Reminder Box

IoT (Internet of Things) refers to a system of internet-connected devices which are capable of sending and receiving the data without human intervention. This technology enabled the remote monitoring in health care sector which lead to keep the patients safe and heathy, and ensuring to deliver superlative care. Some people apparently should be taken care by the caretakers and other family members. This is not provided by everyone in today’s life. So, they may forget to consume medicines at the right time and may also forget what pill has to be taken. This paper aims to develop a device which alerts the patients to take medicine at the right time in an efficient manner. The proposed system has a facility which alerts the patients to take medicine on time and also acknowledges the medicine in-take through a Gmail notification. In the proposed system, the alarm is set up with the help of Blynk app indicating the patient to take the medicine. The IR sensor interfaced with the microcontroller unit determines the patient medicine in-take. The sensor detects the hand movement of the patient when he opens and takes the medicine from the slot provided. The corresponding message is sent to the caretaker via Gmail based on the IR sensor status.</jats:p

Effects of Cigarette Smoke Condensate on Oxidative Stress, Apoptotic Cell Death, and HIV Replication in Human Monocytic Cells.

While cigarette smoking is prevalent amongst HIV-infected patients, the effects of cigarette smoke constituents in cells of myeloid lineage are poorly known. Recently, we have shown that nicotine induces oxidative stress through cytochrome P450 (CYP) 2A6-mediated pathway in U937 monocytic cells. The present study was designed to examine the effect of cigarette smoke condensate (CSC), which contains majority of tobacco constituents, on oxidative stress, cytotoxicity, expression of CYP1A1, and/or HIV-1 replication in HIV-infected (U1) and uninfected U937 cells. The effects of CSC on induction of CYP1 enzymes in HIV-infected primary macrophages were also analyzed. The results showed that the CSC-mediated increase in production of reactive oxygen species (ROS) in U937 cells is dose- and time-dependent. Moreover, CSC treatment was found to induce cytotoxicity in U937 cells through the apoptotic pathway via activation of caspase-3. Importantly, pretreatment with vitamin C blocked the CSC-mediated production of ROS and induction of caspase-3 activity. In U1 cells, acute treatment of CSC increased ROS production at 6H (>2-fold) and both ROS (>2 fold) and HIV-1 replication (>3-fold) after chronic treatment. The CSC mediated effects were associated with robust induction in the expression of CYP1A1 mRNA upon acute CSC treatment of U937 and U1 cells (>20-fold), and upon chronic CSC treatment to U1 cells (>30-fold). In addition, the CYP1A1 induction in U937 cells was mediated through the aromatic hydrocarbon receptor pathway. Lastly, CSC, which is known to increase viral replication in primary macrophages, was also found to induce CYP1 enzymes in HIV-infected primary macrophages. While mRNA levels of both CYP1A1 and CYP1B1 were elevated following CSC treatment, only CYP1B1 protein levels were increased in HIV-infected primary macrophages. In conclusion, these results suggest a possible association between oxidative stress, CYP1 expression, and viral replication in CSC-treated cells of myeloid lineage. This study warrants a closer examination of the role of CYP1B1 in smoking-mediated enhanced HIV replication

Advancing Cancer Therapy with Present and Emerging Immuno-Oncology Approaches

Immuno-oncology (I-O) is a young and growing field on the frontier of cancer therapy. Contrary to cancer therapies that directly target malignant cells, I-O therapies stimulate the body\u27s immune system to target and attack the tumor, which is otherwise invisible to, or inhibiting the immune response. To this end, several methods have been developed: First, passive therapies that enable T-cells to fight the tumor without direct manipulation, typically through binding and modifying the intracellular signaling of surface receptors. Checkpoint inhibitors, perhaps the most well known of I-O therapies; are an example of such. These are monoclonal antibodies that block binding of the tumor cell at receptors that inactivate the T-cell. A variety of small molecules can achieve the same effect by affecting metabolic or signaling pathways to boost the immune response or prevent its attenuation. Drugs originally formulated for unrelated disease states are now being used to treat cancer under the I-O approach. Second, active therapies which often involve direct manipulations that occur in vitro and once introduced to the patient will directly attack the tumor. Adoptive cell transfer is the oldest of these methods. It involves the removal of T-cells from the body, which are then expanded and genetically modified for specificity toward tumor-associated antigens (TAAs), and then reintroduced to the patient. A similar approach is taken with cancer vaccines, where TAAs are identified and reintroduced with adjuvants to stimulate an immune response, sometimes in the context of antigen-presenting cells or viral vectors. Oncolytic viruses are genetically modified natural viruses for selectivity toward tumor cells. The resulting cytotoxicity has the potential to elicit an immune response that furthers tumor cell killing. A final active approach is bi-specific T-cell engagers. These modified antibodies act to link a T-cell and tumor cell through surface receptors and thereby forcibly generate immune recognition. The therapies in each of these subfields are all still very new and ongoing clinical trials could provide even further additions. The full therapeutic potential of the aforementioned therapies, alone or in combination, has yet to be realized, but holds great promise for the future of cancer treatment