**2. Intracellular levels of GSH in viral-infected cells and related cellular alterations**

The JC virus is a double-stranded (ds) DNA virus from Polyomaviridae family. The mechanism of human-to-human transmission of the JC virus has not been established. It has been suggested that the ingestion of contaminated water and food represents the portal of entrance of this virus in human. The virus entry in the CNS goes through the blood-brain barrier (BBB), infecting the brain microvascular endothelium cells. The virus also infects B lymphocytes in the periphery that like a Trojan horse infiltrated the CNS in immunocompromised patients. There the virus infects oligodendrocytes and astrocytes [7]. JC virus is an etiologic agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease. Unfortunately, this disease is currently untreatable and fatal. The relationship between progressive multifocal leukoencephalopathy and GSH still remains unknown. Moreover, JC virus has also been related with CNS tumors, astrocytomas, glioblastomas, neuroblastomas, and medulloblastomas in immunosuppressed and nonimmunosuppressed individuals [8]. However, GSH and GSH-related enzymes constitute an important mechanism of drug and multidrug resistance to glioblastomas, as described below [9].

infiltration and inflammation, inducing a progressive damage [1], such as the progressive multifocal leukoencephalopathy with the John Cunningham virus (JC), AIDS-related dementia complex observed in HIV-1-infected patients, neurodevelopmental sequelae (mental retardation, cerebral palsy, and sensorineural hearing loss) caused by congenital cytomegalovirus (CMV) infection or cerebral mass lesions in immunocompromised adults CMV-infected, and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP) that affects the

In normal oxidative metabolism, the free radical formation is expected. During the 1950s, researchers observed the occurrence of reactive oxygen species (ROS) during molecule irradiation with X-rays and as an effect of normal enzyme metabolic activity. They started to propose that the formation of oxygen free radicals induced tissues and cell damage [2]. At the same time, it was suggested that the mice treatment with glutathione (GSH) inhibited the

GSH is a tripeptide synthesized in all mammalian cells from the amino acid precursors l-glutamate, l-cysteine, and glycine, through the reactions catalyzed by γ-glutamylcysteine and GSH synthetase. Physiologically, 98% of intracellular glutathione is found in reduced form, and only 2% is detected under oxidized form (GSSH) or joined with other molecules [4]. Glutathione (GSH) has an important role in cellular physiology and metabolism, including antioxidant activity and induction of cellular proliferation [5]. Furthermore, the GSHdependent antioxidant enzymes (glutathione peroxidase-1, glutathione reductase, glutathione S-transferase) cooperate and are interconnected reactions that eliminate ROS or controlled the redox state. Dysregulation of GSH synthesis was associated with many diseases, such as diabetes mellitus, cholestatic liver disease, endotoxemia, alcoholic liver disease, cancer, and neurodegenerative diseases. During aging the GSH content was decreased in the liver, lung, kidney, red blood cells, spleen lymphocytes, cerebral cortex, and cerebellum. This GSH concentration decline was related with the reduced expression of proteins involved in GSH synthesis. The GSH levels have been studied in Alzheimer's and Parkinson's diseases and others conditions [6]. However, the CNS is exposed to many situations that can induce a cell and tissue damage associated with ROS production. In this chapter, we will discuss some aspects of the balance of GSH levels and oxidative stress during viral infections in the CNS.

**2. Intracellular levels of GSH in viral-infected cells and related** 

The JC virus is a double-stranded (ds) DNA virus from Polyomaviridae family. The mechanism of human-to-human transmission of the JC virus has not been established. It has been suggested that the ingestion of contaminated water and food represents the portal of entrance of this virus in human. The virus entry in the CNS goes through the blood-brain barrier (BBB), infecting the brain microvascular endothelium cells. The virus also infects B lymphocytes in the periphery that like a Trojan horse infiltrated the CNS in immunocompromised patients. There the virus infects oligodendrocytes and astrocytes [7]. JC virus is an etiologic agent of progressive multifocal leukoencephalopathy (PML), a demyelinating disease. Unfortunately, this disease is currently

human T-cell lymphotropic virus type 1 (HTLV-1)-infected individuals.

animal deaths caused by X-ray irradiation [2, 3].

66 Glutathione in Health and Disease

**cellular alterations**

CMV is a member of *beta*-herpesvirus subfamily, in the family Herpesviridae. It is the largest human herpesvirus, with a 230-kb ds DNA genome infection. Virus is spread from infected individual to noninfected individual by body fluids, such as urine, saliva, blood, tears, semen, and breast milk. In addition, a CMV-infected woman can pass the virus to her developing baby during pregnancy [10]. Congenital CMV infection causes serious neurodevelopmental sequelae, including mental retardation, cerebral palsy, and sensorineural hearing loss. CMV also is an increasingly important opportunistic pathogen in immunocompromised patients, inducing cerebral mass lesions. Antiviral therapy of children with symptomatic CNS congenital CMV infection is effective at reducing the risk of long-term disabilities [11].

In muscle cells the CMV infection induces ROS production minutes after entry. This phenomenon is associated to the virus life cycle. The increase in ROS levels activates the transcriptional factor NF-κB, leading transactivation of the viral genes and inducing the transcription of viral proteins [12]. On the other hand, the infection induces increased levels of GSH to control the ROS generation in vitro. This GSH augment is essential to produce the viral progeny. These data suggested that CMV infection coordinates conditions where ROS levels should be controlled and oxidative stress minimized [13]. However, the CMV infection in peripheral blood erythrocytes of pregnant women induces reduced of GSH and GSH peroxidase levels, leading an increase of H2 O2 levels. These effects were associated with hemolytic anemia in pregnant women [14]. Although the CMV infection has been demonstrated in human brain cells in vitro*,* such as endothelial cells, astrocytes, neuronal cells, oligodendrocytes, and microglia [11], these studies did not investigate the role of GSH in CMV infection.

HTLV-1 and HIV were classified to the genus *Lentivirus* within the family of Retroviridae, subfamily Orthoretrovirinae. This virus infects leukocytes, which circulate in the blood and lymphatic vessels and may infiltrate in the spinal cord or brain, inducing a neurological diseases [7]. These viruses can be transmitted vertically from mother to child during transplacental transfer, delivery, or breastfeeding, by sexual contact and parenterally through the transfusion of the blood, organ transplant, and blood components or through contaminated needles.

HTLV-1 is the etiological agent of the adult T-cell leukemia/lymphoma and HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP), a chronic progressive disabling disease characterized by demyelination, axonal loss, neuronal degeneration, and gliosis. The main site of neurodegeneration is the thoracic spinal cord; this leads to a slowly progressive spastic paraparesis with low back pain and bowel, urinary, and sexual dysfunction. The treatment consists in diminishing the symptoms, using corticosteroid therapy [15, 16]. It was demonstrated that Tax, a HTLV-1-viral protein, induces an increase in ROS generation, causing DNA damage and cellular senescence [17]. Moreover, it was observed that the persistence of the virus in infected cells involves mitochondrial ROS production modulated by viral protein p13 [18]. The CD4+ T lymphocytes are the main targets of HTLV-1 infection, but it has been shown that other leukocytes and glial cells are also infected [19–21]. The infected cells can migrate to the spinal cord and induced the HAM/TSP development. HTLV-1-infected individuals present a spontaneous T-lymphocyte proliferation. This phenomenon is related to the HTLV-1-proviral load and the persistence of the infection. The spontaneous proliferation induced by HTLV-1 infection depends on intracellular GSH levels. Using a GSH synthesis inhibitor, DL-Buthionine-[S,R]-sulfoximine (BSO), the spontaneous proliferation induced by HTLV-1 was impaired in peripheral blood mononuclear cells (PBMC) from infected donors. On the other hand, the GSH precursor induces an increase in mitogen-stimulated cellular proliferation in HTLV-1-infected individuals [19]. Thus, modulation of GSH levels could be proposed as a therapeutic target in HTLV-1-associated diseases.

HIV infection is associated with acquired immune deficiency syndrome (AIDS) development. Combinations of antiretroviral drugs administered as highly active antiretroviral therapy (HAART) reduced the AIDS mortality. However, since 1997 it has been described that 10–20% of virus-infected individuals present HIV-associated dementia [22]. Moreover, the HIV infection also causes mild neurocognitive disorder and demyelinating neuropathy with motor and sensory impairments. The treatment of neurological diseases in HIV-1-infected individuals is established based at the symptoms in association or not with HAART. After virus entry in the CNS, it infects and replicates in microglia that acquire inflammatory phenotype, inducing a neurological damages [7]. Several groups showed a GSH deficiency in the HIV-infected tissues. Initially, the studies demonstrated that low levels of GSH were related to the impairment of T-cell functions. In this work, the authors observed an increase of survival of AIDS patients after the oral treatment with N-acetylcysteine (NAC) administration, a precursor of GSH [22, 23]. In addition, the thiols level analysis in the cerebrospinal fluid (CSF) from HIV-infected patients indicated a significantly reduction in GSH and cysteinyl-glycine levels. Furthermore, the treatment of HIV-infected patients with S-adenosylmethionine, a precursor of homocysteine that is used in GSH synthesis, induced an increase of GSH levels in CSF [24]. Together, these findings suggested the importance of GSH modulation during HIV infection, but the pathway involved to alter the GSH levels remained unknown. The mechanism of neurodegeneration involves the viral protein, gp41. Neurons' death was observed when these cells were incubated in the presence of lentivirus lytic peptide (LLP-1) that expresses the carboxy terminal cytoplasmic domain of gp41 from HIV-1. In addition, the incubation the neuron cell lines with LLP-1 induced a decrease in GSH levels, mitochondrial membrane depolarization, and H2 O2 production rapidly. The combination of GSH or NAC with LLP-1 prevented the mitochondrial membrane depolarization and cell death [25].

promoter regions of detoxifying and antioxidants genes [27]. Viral protein Tat enhanced cellular expression of Nrf2 and its translocation into the nucleus. Nrf2 overexpression inhibited the Tat effects, reducing the intracellular ROS and increasing intracellular levels of GSH [28]. The effects of gp120 and Tat can be observed in vivo using a mice model. The administration of gp120 and Tat together or alone decreased GSH and GSH peroxidase brain levels. Animals also presented a reduction of tight junction protein ZO-1, suggesting other effects into BBB, and exhibited augment in lipid peroxidation in the brain [29]. In **Figure 1**, the direct effects of HIV infection in GSH levels and the consequences of this modulation are summarized.

**Figure 1.** Levels of GSH during HIV-1 infection. In HIV-1-infected patients lower levels of GSH in the peripheral blood and CSF were observed. Alterations in intracellular levels of GSH cells were showed in vivo and in vitro. HIV proteins gp120, Tat, and gp41—reduced intracellular levels of GSH in endothelial cells, astrocytes, and neurons. The effects of

The Role of Glutathione in Viral Diseases of the Central Nervous System

http://dx.doi.org/10.5772/intechopen.76579

69

GSH is related with the transport of endogenous and exogenous molecules to extracellular medium. GSH is a physiological substrate of ABCC1. Multidrug resistance-related protein 1 (ABCC1) transports several compounds in a GSH-dependent manner; its activity could be stimulated by the GSH intracellular levels. The members of the ABCC family are ATPdependent efflux pumps, belonging to the ABC family of transport proteins, and they are also

**3. HTLV-1 and HIV-1 infection and GSH active transport**

HIV-1-viral proteins also involved BBB breakdown, lipid peroxidation, and cell death.

The development of neurological HIV disorders depends on virus entry in the CNS. To access the CNS, HIV virus particles and the infected cells induce the BBB disruption. HIV envelope protein gp120 and the regulatory virus protein, Tat, are involved in BBB breakdown. The incubation of endothelial brain cells with gp120 and Tat reduced the GSH intracellular levels and decreased GSH/GSSG ratio. These proteins also caused an increase in lipid peroxidation, suggesting that gp120 and Tat played an important role in BBB disruption by induction of oxidative cellular stress in endothelial cells [26].

The antioxidant response signals induce the activation of nuclear factor erythroid-derived 2-like-2 (Nrf2). It is a transcription factor that translocates into the nucleus and binds in the The Role of Glutathione in Viral Diseases of the Central Nervous System http://dx.doi.org/10.5772/intechopen.76579 69

shown that other leukocytes and glial cells are also infected [19–21]. The infected cells can migrate to the spinal cord and induced the HAM/TSP development. HTLV-1-infected individuals present a spontaneous T-lymphocyte proliferation. This phenomenon is related to the HTLV-1-proviral load and the persistence of the infection. The spontaneous proliferation induced by HTLV-1 infection depends on intracellular GSH levels. Using a GSH synthesis inhibitor, DL-Buthionine-[S,R]-sulfoximine (BSO), the spontaneous proliferation induced by HTLV-1 was impaired in peripheral blood mononuclear cells (PBMC) from infected donors. On the other hand, the GSH precursor induces an increase in mitogen-stimulated cellular proliferation in HTLV-1-infected individuals [19]. Thus, modulation of GSH levels could be

HIV infection is associated with acquired immune deficiency syndrome (AIDS) development. Combinations of antiretroviral drugs administered as highly active antiretroviral therapy (HAART) reduced the AIDS mortality. However, since 1997 it has been described that 10–20% of virus-infected individuals present HIV-associated dementia [22]. Moreover, the HIV infection also causes mild neurocognitive disorder and demyelinating neuropathy with motor and sensory impairments. The treatment of neurological diseases in HIV-1-infected individuals is established based at the symptoms in association or not with HAART. After virus entry in the CNS, it infects and replicates in microglia that acquire inflammatory phenotype, inducing a neurological damages [7]. Several groups showed a GSH deficiency in the HIV-infected tissues. Initially, the studies demonstrated that low levels of GSH were related to the impairment of T-cell functions. In this work, the authors observed an increase of survival of AIDS patients after the oral treatment with N-acetylcysteine (NAC) administration, a precursor of GSH [22, 23]. In addition, the thiols level analysis in the cerebrospinal fluid (CSF) from HIV-infected patients indicated a significantly reduction in GSH and cysteinyl-glycine levels. Furthermore, the treatment of HIV-infected patients with S-adenosylmethionine, a precursor of homocysteine that is used in GSH synthesis, induced an increase of GSH levels in CSF [24]. Together, these findings suggested the importance of GSH modulation during HIV infection, but the pathway involved to alter the GSH levels remained unknown. The mechanism of neurodegeneration involves the viral protein, gp41. Neurons' death was observed when these cells were incubated in the presence of lentivirus lytic peptide (LLP-1) that expresses the carboxy terminal cytoplasmic domain of gp41 from HIV-1. In addition, the incubation the neuron cell lines with LLP-1 induced a decrease in GSH levels,

O2

or NAC with LLP-1 prevented the mitochondrial membrane depolarization and cell death [25].

The development of neurological HIV disorders depends on virus entry in the CNS. To access the CNS, HIV virus particles and the infected cells induce the BBB disruption. HIV envelope protein gp120 and the regulatory virus protein, Tat, are involved in BBB breakdown. The incubation of endothelial brain cells with gp120 and Tat reduced the GSH intracellular levels and decreased GSH/GSSG ratio. These proteins also caused an increase in lipid peroxidation, suggesting that gp120 and Tat played an important role in BBB disruption by induction of

The antioxidant response signals induce the activation of nuclear factor erythroid-derived 2-like-2 (Nrf2). It is a transcription factor that translocates into the nucleus and binds in the

production rapidly. The combination of GSH

proposed as a therapeutic target in HTLV-1-associated diseases.

68 Glutathione in Health and Disease

mitochondrial membrane depolarization, and H2

oxidative cellular stress in endothelial cells [26].

**Figure 1.** Levels of GSH during HIV-1 infection. In HIV-1-infected patients lower levels of GSH in the peripheral blood and CSF were observed. Alterations in intracellular levels of GSH cells were showed in vivo and in vitro. HIV proteins gp120, Tat, and gp41—reduced intracellular levels of GSH in endothelial cells, astrocytes, and neurons. The effects of HIV-1-viral proteins also involved BBB breakdown, lipid peroxidation, and cell death.

promoter regions of detoxifying and antioxidants genes [27]. Viral protein Tat enhanced cellular expression of Nrf2 and its translocation into the nucleus. Nrf2 overexpression inhibited the Tat effects, reducing the intracellular ROS and increasing intracellular levels of GSH [28].

The effects of gp120 and Tat can be observed in vivo using a mice model. The administration of gp120 and Tat together or alone decreased GSH and GSH peroxidase brain levels. Animals also presented a reduction of tight junction protein ZO-1, suggesting other effects into BBB, and exhibited augment in lipid peroxidation in the brain [29]. In **Figure 1**, the direct effects of HIV infection in GSH levels and the consequences of this modulation are summarized.
