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Sigma-1 Receptor Antagonist (BD1047) prior to Cocaine Reduces Cathepsin B Secretion in HIV-1 Models In Vivo and In Vitro.

紀錄類型:	書目-語言資料,手稿 : Monograph/item
正題名/作者:	Sigma-1 Receptor Antagonist (BD1047) prior to Cocaine Reduces Cathepsin B Secretion in HIV-1 Models In Vivo and In Vitro./
作者:	Velez-Lopez, Omar.
面頁冊數:	1 online resource (187 pages)
附註:	Source: Dissertations Abstracts International, Volume: 80-12, Section: B.
Contained By:	Dissertations Abstracts International80-12B.
標題:	Epidemiology. -
電子資源:	click for full text (PQDT)
ISBN:	9781392188958

Sigma-1 Receptor Antagonist (BD1047) prior to Cocaine Reduces Cathepsin B Secretion in HIV-1 Models In Vivo and In Vitro.
Velez-Lopez, Omar.

Sigma-1 Receptor Antagonist (BD1047) prior to Cocaine Reduces Cathepsin B Secretion in HIV-1 Models In Vivo and In Vitro. - 1 online resource (187 pages)

Source: Dissertations Abstracts International, Volume: 80-12, Section: B.

Thesis (Ph.D.)--University of Puerto Rico Medical Sciences (Puerto Rico), 2019.

Includes bibliographical references

Background: Human Immunodeficiency virus (HIV-1) is the etiological agent of acquired immunodeficiency syndrome (AIDS). Acquisition and progression of HIV-1 infection often leads to the development of neurocognitive disorders in at least 50% of the patients despite antiretroviral therapy. HIV-1 infected patients develop a spectrum of neurocognitive disorders, which are categorized in three stages: asymptomatic (ANI), mild neurocognitive, impairment (MND), HIV dementia (HAD) and are collectively termed as HIV-associated neurological disorders (HAND). Substance abuse is a problematic comorbidity among HIV-1 patients that increases the risk of infection and promotes viral progression. Among all substances of abuse, cocaine is the most commonly used drug by HIV-infected individuals that exacerbates the deleterious effects of HIV disease. For example, cocaine increases HIV-1 replication, disrupts the blood brain barrier, and enhances inflammatory states of microglia and other immune cells within the brain. One of the traits of the pathogenesis of HAND is the infiltration of perivascular macrophages into the brain with the concomitant secretion of viral proteins, neurotoxic, and inflammatory factors. One of these factors is cathepsin B (CATB), a lysosomal cysteine protease that induces neuronal apoptosis, and is increased in plasma and cerebrospinal fluid of HIV-1 infected patients (Cantres-Rosario et al., 2013). Cocaine potentiates CATB neurotoxicity both in vitro and in vivo (Zenon et al., 2014). One potential molecular target for assessing the role of cocaine in cathepsin B secretion in infected macrophages is through the modulation of the sigma-1 receptor (Sig1R). Sig1R is a small endoplasmic reticulum (ER) chaperone with versatile functions and a binding site for cocaine under physiological conditions (2-7 μM) (Sharkey et al., 1988). Originally classified as an opioid receptor N-allylnormetazocine (SKF 10,047), it was subsequently characterized as a transmembrane chaperone protein with different functions than those of classical opioid receptors. Its main cellular functions include controlling: endoplasmic reticulum stress (ER) responses, regulation of calcium efflux from the ER to the Golgi and mitochondria, control of sodium channels and protein folding among various other functions (Su et al., 2010).Modulation of sigma-1 (Sig1R) by cocaine increases oxidative species, cytokines and other factors that could promote lysosomal disruption (Boya & Kroemer 2008). Pharmacological modulation of this receptor with Sig1R antagonist (BD1047) prior to cocaine treatment has been shown to decrease: HIV-1 replication of microglia (Gekker et al., 2006), adhesion molecules in infected monocytes (Yao et al., 2010), reactive oxygen species and autophagy markers in astrocytes (Cao et al., 2016), among other functions. The focus of this study is to determine the role of Sig1R in CATB secretion and HIV-1 replication in macrophages exposed to cocaine. We hypothesized that pharmacological modulation of Sig1R by BD1047 prior to cocaine treatment could alter CATB secretion in HIV-1 infected macrophages both in vitro and in vivo. Methods: Monocyte derived-macrophages (MDM) from HIV-1 seronegative donors were isolated, infected with HIV-1ADA, and pretreated with Sig1R antagonist (BD1047) or Sig1R agonist (PRE-084) prior to cocaine exposure for 3,6,9 and 11 days post-infection (dpi) (Aim 1). MDM supernatants collected at these time intervals were assayed for HIV-1 replication, and supernatants from 11 dpi were used for pro-cathepsin B and total cathepsin B ELISA assays. Serum-free supernatants collected at 12 dpi was added to neuronal (HTB-11) cultures to measure apoptosis by TUNEL assays. In addition, experiments were conducted in vivo using the HIV encephalitis mouse model (HIVE) with treatments of mice with BD1047 (20 mg/g/weight) prior to cocaine exposure (15 mg/kg/weight) for fourteen (14) days (Aim 2). Protein from striatal tissue were isolated and Western blotted for cathepsin B, Sig1R, and microtubule associated protein (MAP-2). Paraffin- embedded tissues were also analyzed for the expression of CATB, synaptophysin (a presynaptic injury marker), cleaved-caspase-3 (total apoptosis), glial acidic fibrillary protein (GFAP) for astrogliosis, and p24 (HIV-1 capsid protein) for infection. Also, the levels of expression of CATB and Sig1R was measured in the post-mortem brains of HIV-1 infected cocaine users by immunohistochemistry. In aim 3, quantitative proteomics studies were performed on MDM lysates from 12 dpi using the Tandem Mass Tag (TMT) approach followed by mass spectrometry and Bioinformatics. We used a 10-plex TMT array, with samples from uninfected and infected MDM lysates treated with the Sig1R antagonist (BD1047) prior to cocaine versus cocaine exposed group without the antagonist. were digested, alkylated and reduced prior to combination with TMT tags. After tagging, samples were injected into the Q-Exactive LC/MS/MS Orbitrap and proteins were quantified using Proteome Discoverer program and R-programming analyses. (Abstract shortened by ProQuest).

Electronic reproduction.
Ann Arbor, Mich. :
ProQuest,
2024

Mode of access: World Wide Web

ISBN: 9781392188958Subjects--Topical Terms:

635923
Epidemiology.
Subjects--Index Terms:

Bd1047Index Terms--Genre/Form:

554714
Electronic books.

Sigma-1 Receptor Antagonist (BD1047) prior to Cocaine Reduces Cathepsin B Secretion in HIV-1 Models In Vivo and In Vitro.
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520 $a Background: Human Immunodeficiency virus (HIV-1) is the etiological agent of acquired immunodeficiency syndrome (AIDS). Acquisition and progression of HIV-1 infection often leads to the development of neurocognitive disorders in at least 50% of the patients despite antiretroviral therapy. HIV-1 infected patients develop a spectrum of neurocognitive disorders, which are categorized in three stages: asymptomatic (ANI), mild neurocognitive, impairment (MND), HIV dementia (HAD) and are collectively termed as HIV-associated neurological disorders (HAND). Substance abuse is a problematic comorbidity among HIV-1 patients that increases the risk of infection and promotes viral progression. Among all substances of abuse, cocaine is the most commonly used drug by HIV-infected individuals that exacerbates the deleterious effects of HIV disease. For example, cocaine increases HIV-1 replication, disrupts the blood brain barrier, and enhances inflammatory states of microglia and other immune cells within the brain. One of the traits of the pathogenesis of HAND is the infiltration of perivascular macrophages into the brain with the concomitant secretion of viral proteins, neurotoxic, and inflammatory factors. One of these factors is cathepsin B (CATB), a lysosomal cysteine protease that induces neuronal apoptosis, and is increased in plasma and cerebrospinal fluid of HIV-1 infected patients (Cantres-Rosario et al., 2013). Cocaine potentiates CATB neurotoxicity both in vitro and in vivo (Zenon et al., 2014). One potential molecular target for assessing the role of cocaine in cathepsin B secretion in infected macrophages is through the modulation of the sigma-1 receptor (Sig1R). Sig1R is a small endoplasmic reticulum (ER) chaperone with versatile functions and a binding site for cocaine under physiological conditions (2-7 μM) (Sharkey et al., 1988). Originally classified as an opioid receptor N-allylnormetazocine (SKF 10,047), it was subsequently characterized as a transmembrane chaperone protein with different functions than those of classical opioid receptors. Its main cellular functions include controlling: endoplasmic reticulum stress (ER) responses, regulation of calcium efflux from the ER to the Golgi and mitochondria, control of sodium channels and protein folding among various other functions (Su et al., 2010).Modulation of sigma-1 (Sig1R) by cocaine increases oxidative species, cytokines and other factors that could promote lysosomal disruption (Boya & Kroemer 2008). Pharmacological modulation of this receptor with Sig1R antagonist (BD1047) prior to cocaine treatment has been shown to decrease: HIV-1 replication of microglia (Gekker et al., 2006), adhesion molecules in infected monocytes (Yao et al., 2010), reactive oxygen species and autophagy markers in astrocytes (Cao et al., 2016), among other functions. The focus of this study is to determine the role of Sig1R in CATB secretion and HIV-1 replication in macrophages exposed to cocaine. We hypothesized that pharmacological modulation of Sig1R by BD1047 prior to cocaine treatment could alter CATB secretion in HIV-1 infected macrophages both in vitro and in vivo. Methods: Monocyte derived-macrophages (MDM) from HIV-1 seronegative donors were isolated, infected with HIV-1ADA, and pretreated with Sig1R antagonist (BD1047) or Sig1R agonist (PRE-084) prior to cocaine exposure for 3,6,9 and 11 days post-infection (dpi) (Aim 1). MDM supernatants collected at these time intervals were assayed for HIV-1 replication, and supernatants from 11 dpi were used for pro-cathepsin B and total cathepsin B ELISA assays. Serum-free supernatants collected at 12 dpi was added to neuronal (HTB-11) cultures to measure apoptosis by TUNEL assays. In addition, experiments were conducted in vivo using the HIV encephalitis mouse model (HIVE) with treatments of mice with BD1047 (20 mg/g/weight) prior to cocaine exposure (15 mg/kg/weight) for fourteen (14) days (Aim 2). Protein from striatal tissue were isolated and Western blotted for cathepsin B, Sig1R, and microtubule associated protein (MAP-2). Paraffin- embedded tissues were also analyzed for the expression of CATB, synaptophysin (a presynaptic injury marker), cleaved-caspase-3 (total apoptosis), glial acidic fibrillary protein (GFAP) for astrogliosis, and p24 (HIV-1 capsid protein) for infection. Also, the levels of expression of CATB and Sig1R was measured in the post-mortem brains of HIV-1 infected cocaine users by immunohistochemistry. In aim 3, quantitative proteomics studies were performed on MDM lysates from 12 dpi using the Tandem Mass Tag (TMT) approach followed by mass spectrometry and Bioinformatics. We used a 10-plex TMT array, with samples from uninfected and infected MDM lysates treated with the Sig1R antagonist (BD1047) prior to cocaine versus cocaine exposed group without the antagonist. were digested, alkylated and reduced prior to combination with TMT tags. After tagging, samples were injected into the Q-Exactive LC/MS/MS Orbitrap and proteins were quantified using Proteome Discoverer program and R-programming analyses. (Abstract shortened by ProQuest).
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