INTRODUCTION AND NEUROEPIDEMIOLOGY OF HIV/AIDS

AIDS was first recognized as a new and distinct clinical entity in 1981 1 and the HIV-1 as their casual agent in 1983 2. Since then, the HIV/AIDS epidemic has reached epidemic proportions with a total accumulative number of more than 60 million people, according to the Joint United Nations Programme on HIV/AIDS (UNAIDS) and WHO. The extensive spread of HIV-1 epidemics in Asia was not appreciated in the 1980s, however, several countries including China, India, Burma, Thailand, Indonesia and Vietnam have extremely growing epidemic 3. The first AIDS case in China was identified in Beijing in 1985 and since then the HIV-1 has spread rapidly through the whole country. By the end of 2002, more that 40, 000 HIV-1 cases were reported, and more than 2600 of cases were diagnosed with full-blown AIDS 4, 5. It is estimated that the present number of HIV-1 cases exceeds one million 6. By the year 2010 China is expected to reach 10 millions HIV-1 infected 7. The increase of prostitution, heterosexual transmission and injected drug users indicates the potential for HIV-1 spread in the future of China.

Neurological complications associated with HIV-1 are being recognized since the beginning of AIDS epidemic. The initial step in HIV-1 neuropathogenesis is viral entry into the central nervous system (CNS). This infiltration by HIV-1 can cause direct and/or indirect neurological complications 8. ADC is a common neurologic complication unique to HIV-1 infection 8. The prevalence of HIV-1-associated encephalitis is the sole manifestation in nearly 3–5% of patients with HIV-1 infection 9, with incidence ranging from 30 to 60% in the late stages of AIDS. In developing countries in Asia the three most opportunistic infections are TB, cryptococcosis, and Pneumocystis carinii pneumonia 10. Therefore, it is expected that secondary neurological complications due to TB and cryptococcosis will be the most common cause of morbility and mortality in HIV-1/AIDS cases in China. In addition, other frequent infections includes cytomegalovirus (CMV), toxoplasmosis and hepatitis C. Less common secondary neurological complications seen in patients with late stages of AIDS in developing countries includes JC virus associated progressive multifocal encephalopathy (PML) and HIV-1 associated malignancies primary CNS lymphoma. Overall in China, we expect HIV-1 associated neurological symptoms as a sole manifestation in nearly 50,000 of patients. In addition, we expect a high incidence of neurological complications ranging from 250,000 to more than half of a million of AIDS patients.

ART has clearly improved the morbidity and mortality in HIV-1 infected individuals by reducing plasma viral load and restoring immune function. Despite the overall improvement of outcome in those AIDS patients receiving ART, however, neurocognitive impairments continue to present in 10% of patients with HIV-1 associated dementia 11 and in up to 50% of patients with HIV-1 encephalopathy 12, 13. These findings could be attributed to AIDS patients developing resistance to or failure of ART 14, with the blood brain-barrier limiting access of anti-viral drugs to infected sites within the parenchyma 15. It is possible that cells from the CNS harbor latent HIV-1 and serve as a persistent reservoir of virus, responsible for viral reactivation and gradual neurocognitive decline.

NEUROTROPISM AND BRAIN COMPARTMENTALIZATION OF HIV-1

The HIV-1 is the agent that causes AIDS 2, and it is a RNA virus, which is a member of the retroviridae family. The genetic organization of HIV-1 is quite complex. In addition to the structural genes gag, pol and env possessed by all retroviruses, HIV-1 contains genes that encode for regulatory proteins: rev (regulator of virion structural protein), tat (trans-activator), and nef (negative regulatory factor); and genes that encode for proteins believed to be involved in virus maturation and release: vif (virion infectivity factor), vpu (virus protein U), and vpr (virus protein R). Based on relatedness of nucleotide sequences and the diversion seen among HIV-1 isolates from people of different countries, a classification of HIV-1 into subtypes or clades has been designed. Such subtypes, designed A through K, have envelope gene sequences that vary by 20% or more between subtypes.

Several studies have demonstrated that microglia/macrophages are important cellular reservoirs for productive HIV-1 infection in the brain 10, 16, 17, 18, 19, 20, 21, 22. In addition, it has been shown that astrocytes can be infected by HIV-1 in vitro and in vivo, by the detection of early regulatory genes such as nef and rev 16, 18, 23, 24, 25. In general, the selectivity of HIV-1 strains for different cell types is regulated by interactions between the viral envelope and cellular receptors. For example, HIV-1 infection of T helper lymphocytes and monocytes/macrophages is dependent upon the presence of cellular CD4 receptors 26, 27. In addition, some members of the chemokine receptor family are required in conjunction with CD4 as coreceptors for HIV-1 entry into target cells; T-tropic HIV-1 utilizes CXCR4 as a co-receptor 28, whereas M-tropic HIV-1 utilizes CCR5 and CCR3 29, 30, 31, 32, 33, 34. HIV-1 can also infect other cell types that do not express CD4, including brain-derived glial 35 and neuronal cells 36, 37, human skin fibroblasts 38, muscle cells 39, human trophoblast cells 40, follicular dendritic cells 41, colonic epithelial cells 42, fetal adrenal cells 43, and human liver carcinoma cells 44.

The V3 loop of gp120 has been identified as a primary determinant of HIV-1 cell tropism 45, 46, 47. Sequences within the V3 loop has been shown to be associated with either the M-tropic, or T tropic phenotypes 45, 48, 49. We have demonstrated the importance of the V3 loop of T-tropic HIV-1 as a primary determinant for infectivity of CD4-negative neuronal SK-N-MC cells 50, and astrocytes 51. Collectively, these data indicate that infection and/or affinity of HIV-1 for cells of the nervous system, a phenomenon called neurotropism, is regulated by the interaction between viral epitopes (V3 loop) and the receptors (see Fig. 1).

Figure 1
figure 1

Neuropathogenesis of HIV-1 Infection. Microglia infection by HIV-1 has been pathologically associated with “HIV-1 encephalitis”. The HIV-1 M tropic phenotype (V3 negative charges) is responsible for microglia infection in presence of CD4 receptor and CCR5 and CCR3 coreceptors. Neurons and astrocytes infection by HIV-1 has been associated with “HIV-1 encephalopathy”. The HIV-1 T tropic phenotype (V3 positive charges) is responsible for astrocyte/neuron infection in absence of CD4 receptor and some cases the presence of CXCR4.

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The genetic evolution of HIV-1 within the brain is distinct from that in lymphoid tissues and other organs 52, 53. The genetic compartmentalization of viral variants in the CNS suggest that adaptive changes occur in response to unique constraints within the brain microenvironment, including specific target cell populations and immune selection pressures 54.

In one study, 37 full-length HIV-1 envelope glycoproteins (env) genes were cloned directly from brain biopsy and blood samples from patients with AIDS 55. Phylogenetic analysis showed distinct clustering of brain relative to blood env sequences, indicating tissue-specific compartmentalization of the virus. However, no brain-specific signature sequence was identified. Furthermore, there were no significant differences in the numbers or positions of N-linked glycosylation sites between brain and blood env sequences. The patterns of coreceptor usage were heterogeneous, with no clear distinction between brain and blood env clones. These results suggest that HIV-1 envelopes in brain cannot be distinguished from those in blood on the basis of coreceptor usage or the number or positions of N-glycosylation sites, indicating that other properties underlie neurotropism.

Astrocytes and microglia has been identified as target cells for HIV-1 infection in the brain, whereas most studies show that viral DNA is rarely detected in neurons 10, 16, 19, 56, 57. HIV-1 has been molecularly characterized from pure populations of astrocytes, macrophages and multinucleated giants cells and it was isolated from brain tissue of ADC patients 58. The V3 region of the HIV-1 env gene was amplified from the pure-cell populations, and multiple clones were sequenced. The V3 env sequences were distinct in astrocytes compared with neighboring macrophages or multinucleated giants cells and were characteristic of CCR5-using HIV-1. These results demonstrate cell-specific compartmentalization of distinct R5-like viral strains in the CNS microenvironment.

NERVOUS SYSTEM: LATENCY AND RESERVOIR FOR HIV-1

The capacity of HIV-1 in establishing latent infection of CD4+ T cells may allow viral persistence despite immune responses and antiretroviral therapy. Measurements of infectious virus, viral RNA in plasma, viral DNA, and viral messenger RNA species from infected cells are suggesting that HIV-1 replication continues throughout the course of infection. During the asymptomatic phase of infection subsist an extremely low total body load of latently infected resting CD4+ T cells with replication-competent integrated provirus (<107 cells). The most prevalent form of HIV-1 DNA, in resting and activated CD4+ T cells, is a full-length, linear, unintegrated form that is not replication competent. The infection progresses even though at any given time, the lymphoid tissues which has integrated HIV-1 DNA, is present in a minute fraction of susceptible populations, including resting and activated CD4+ T cells, and macrophages 59, 60. Furthermore, replication-competent virus has been recovered from resting CD4+ T lymphocytes in patients on ART. This reservoir of latent virus should be considered when deciding to terminate treatment in ART responder patients 61. Most anti-viral therapies used against HIV-1 collect poorly in the CNS because of efflux systems located at the blood-brain barrier (BBB), which rapidly return these drugs back to the circulation 62, 63, 64. As such, HIV-1 within the CNS may act as a reservoir for the re-infection of peripheral tissues.

Human monocytes play an important role in mediating HIV-1 infection of the CNS, and monocytes-derived macrophages represent a major viral reservoir within the brain and other target organs. Microglia are endogenous brain macrophages that show distinct phenotypes such as expression of myeloid antigens, ramified morphology, and location within the neural parenchyma. Microglia play a significant role in the developing of HIV-1-associated encephalitis. Together with monocyte-derived (perivascular) macrophages, microglia represent a major target of HIV-1 infection and these cells are considered traditionally the “brain reservoir” of the virus 65.

Alternatively, due to the large number of astrocytes (1:10) and its crucial role in the brain homeostasis, this cell type is believed to play a significant role in the development of HIV-1-associated encephalopathy. Astrocytes is also serving as brain reservoir for HIV-1. Infection of astrocytes by HIV-1 impairs its function directly and/or indirectly 66, 67. Lastly, multipotential human brain-derived progenitor cells and progenitor-derived astrocytes, but not progenitor-derived neurons, are permissive for HIV-1 and support a latent infection that can be reactivated by differentiation or cytokine stimulation. Progenitor cells in the brain therefore may represent an additional reservoir for HIV-1. These cells may be a source of infected and/or impaired astrocytes in pediatric cases of HIV-1 encephalopathy. These cells may also represent long-term targets for infection in adult patients, who are often living with the infection for 2 or 3 decades before signs of neurological damage are apparent 68.

Several studies have shown a large HIV-1 DNA load in the brains of some AIDS patients 69, 70. In the host cell, retroviral DNAs exist in three main forms: unintegrated linear, unintegrated circular, and integrated (the provirus). Each of the three species of viral DNA are detectable in blood and brain of AIDS patients. Autopsy samples from patients with HIV encephalitis had a considerable higher proportion of unintegrated viral DNA 71, 72. High levels of unintegrated forms of retroviral DNA often correlate with superinfection and accompanying cytopathic effects.

NEUROLOGICAL MANIFESTATIONS OF HIV-1/AIDS

Neurological complications were recognized early in the AIDS epidemic as a consequence of HIV-1 infection. Neurological complications are detected in up to 60% of AIDS patients and are the result of direct (primary) HIV-1 infection and secondary (indirect) opportunistic infections and neoplasm 8 [See Fig. 2]. ADC or HIV-1-associated encephalitis is the most common neurological complication unique to HIV-1 infection 8. Neuropathological studies have suggested that more than 90% of all AIDS patients have nervous system abnormalities 8. These changes are primarily associated with opportunistic infections and neoplasms. Frequently, multiple, coexisting processes are found in the same patient 73. Neurological complications associated with HIV-1 infections vary geographically. Different profiles of neurological illness in AIDS patients are likely related to the geographic location of the reporting institution, the diagnostic studies available, and the percentage of patients from different groups, the time of stage of disease, time of analysis and the availability of ART.

Figure 2
figure 2

Neurological Complications of HIV-1/AIDS. Primary or direct complication of HIV-1 infection includes ADC, myelopathy, peripheral neuropathy and myopathy. Secondary opportunistic infections includes viral (CMV), bacterial (TB), fungal (criptococcus), and parasitic (toxoplasmosis) infections. The most common HIV-1 associated malignancy is the primary non-Hodgkin's CNS lymphoma.

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NeuroAIDS in Mexico and USA

To understand the pattern of HIV-1 associated neurological complications in Mexico in comparison with a population in USA, we conducted a cross-sectional and retrospective study in 120 AIDS patients from Mexico City, Mexico and 500 AIDS cases from Houston, Texas, USA 8. Neurological, laboratory, imaging and pathological examinations identified 40 Mexican and 130 USA patients with neurological complications. ADC was the most common complication in both groups, opportunistic infection such as intracranial tuberculoma was present only in the Mexican population. Tumors such as primary CNS lymphoma were more prevalent in the USA population. Similarly, PML occurred more commonly in the USA population. The different findings in the Mexican population likely reflect afflictions common to developing countries, a high prevalence of tuberculosis and a high mortality rate. These conditions preclude complication such as lymphoma and PML, which develop later in the natural course of HIV-1 infection. However, as HIV-1 patients from developing countries are living longer due to ART and prophylactic drugs for opportunistic infections, more common neurological complications are expected.

NeuroAIDS in Brazil

The first AIDS case in Brazil was reported in 1983. Since then, more than 360,000 cases of AIDS have been reported 74. One hospital which specializes in infectious disease, Institute Emilio Ribas, located in São Paulo, Brazil has admitted more than 30,000 AIDS patients in the last 15 years. Neurological complications have also critically impacted these patients. More than 2,000 patients with CNS cryptococcosis and almost 5,000 patients with CNS toxoplasmosis have been affected. At the Institute Emilio Ribas, a cohort study has been conducted investigating meningeal or meningoencephalitis syndrome. The most common causes for these syndromes were cryptococcosis, TB and syphilis. When brain expansive lesion syndrome predominates, the main causes included CNS toxoplasmosis, TB (tuberculomas or abscess), and primary CNS lymphoma 75. PML represents the most frequent cause of focal brain lesions without mass effect 76. The neurological complications of AIDS in this South American country highlight the importance of evaluating the neuroepidemiological setting in Brazil. For example, CNS TB represents the second most frequent cause of meningitis and expansive brain lesion 77, 78, 79. CNS toxoplasmosis, the most frequent CNS opportunistic disease, has been reduced to an incidence of 50 to 60% 80, which compares with a four-fold reduction in developed countries 81. In Brazil, CNS toxoplasmosis is considered as an AIDS-defining illness with severe immunodeficiency. CNS toxoplasmosis has a high mortality and disability rate, probably due to the epidemic characteristics of underdeveloped nations as heterosexual transmission, feminization and pauperization, and lack of ART 82. The prevalence of neurocognitives disorders and peripheral neuropathies in Brazilian AIDS patients has not being well defined. A molecular diagnosis of neurological opportunistic infections, referred to as “minimally invasive” will be part of a set of critical tools for prevalence analysis of international NeuroAIDS 83, 84. In summary, benefits of universal access programs to ART in Brazil are in some ways similar to developed countries, but there are urgent needs of a national network for surveillance, especially regarding incidence and the prevalence of neurological complications of HIV infection.

NeuroAIDS in India

India has the second largest burden of HIV related pathology next to sub-Saharan Africa. Neurological complications associated to HIV-1 infections, mainly clade C, are very common. The spectrum of HIV-1 associated complications reported within India (Bangalore in the south vs. Pune in the west) appears to be different. TB, cryptococcosis and toxoplasmosis are the major neuropathologies reflecting the endemicity and reactivation of latent infections 85. Viral infections and HIV-1 associated neoplasms such as primary CNS lymphoma are infrequent. ADC or HIV-1 associated encephalitis and myelopathies, are considered infrequent, though proper studies have just been initiated. Other HIV-1 associated complications including peripheral neuropathy have been reported. Future studies are important to understand the biology of neuroAIDS in India with its unique spectrum of opportunistic infections and HIV-1 clades.

FUTURE RESEARCH OF NEUROAIDS IN CHINA

Research of NeuroAIDS in China is essential to understand the impact and the biology of HIV-1 in the nervous system. Future studies would include, the molecular epidemiology and the description of opportunistic infections associated to HIV-1; the neuropathological description of primary and secondary HIV-1 complications in different high risk groups; the HIV-1 neurotropism and immune response studies for China's unique HIV-1 strains and recombinant forms derived from the nervous system, including experimental models such as the use of transgenic rats 86 to understand the HIV-1 neuropathogenesis of China; and the study of “potential” resistant virus, primarily when anti-retroviral therapy does not have full access in the brain. Latency, activation and reactivation of HIV-1 in the nervous system on HIV-1 in the brain are other major areas for future investigations, which will provide new insights into the development of therapeutic agents. If we can improve the neurological consequences of HIV-1/AIDS, we will improve the quality of life and the life expectancy of patients affected with this virus.