Biomarkers in NeuropathogenicHIV-1 Subtypes

Neither it is possible to hit the HIV where it hides, nor to prevent the CNS infection and related CNS injury. In pre-HAART era HIV-associated dementia (HAD) was the most common (20-30%) worldwide among HIV infected AIDS people, especially subtype B predominant infections. However, in post-HAART era the less severe form of neurological disorders consists of a spectrum of neurological complications ranges from mild cognitive motor disorder (MCMD), HIV-1 associated neuro-cognitive disorder (HAND) and asymptomatic neurocognitive impairment (ANI) which are more or less equal among different subtypes. ^[1] Anti-retroviral therapy can detain the nature of HIV associated CNS disease severity, however milder forms of neurological dysfunction remain common. ^[1], ^[7]

Inflammatory molecules as Biomarkers

Previously, in patients without antiretroviral therapy, strong correlations were reported between CSF HIV RNA level and HAD.^[8] In contrast, by the use of CART, association between viral load, immune markers and neurological status are relatively weak. ^[9] In the post-HAART era, the point of difficulties in the neurological disorder underlying the nature of impairment rely chiefly on neuropsychological test performances. ^[10] The indistinguishable problem is to find the particular CNS damage or ongoing injury before and after CART, where laboratory based biomarkers stand with certain potential for HIV related CNS injury. ^[6] Although neurological damages occur by HIV were incompletely understood, many studies suggested that development of dementia correlated with the elevation level of MCP1 and RANTES,^[11] interferon-gamma inducible protein IP10 or CXCL10 in CSF.^[12] Large amount of immune activation marker such as neopterin, released by macrophages is negatively correlated with CD4+ T-cell concentration^[13] and level of CD38 increases in CSF. ^[14] In the absence oftherapy, inflammatory cytokines in serum and CSF such as TNF alpha, IFN gamma getmost of the HIV infection and indicates the systemic as well as CNS diseaseprogression. ^[15]However, in subjects with anti-retroviraltherapy, the CSF may not contain the associative quantity of inflammatory cytokines, whichis why to correlate the CNS disease condition, several CSF biomarkers molecules haveCathepsin B and cystatin B are lysosomal proteinssecreted by macrophages and microglia and the increased level in CSF indicatesusceptibility for neurotoxicity.Deficits in monocyteantioxidants may contribute to identify developed and progression of cognitiveimpairment.neurovirulence in post-HAARTera, mononuclear phagocyte secreted neurotoxic proteins that affect neurocognitiveimpairment have been used in several studies such as N-acetyl aspartate (NAA),arachidonic acid, PAF. The HIV RNA level in CSF toplasma seen in pre-HAART era correlates with neurocognitive status, moreover in post-HAART era the correlation is no longer seen. It is not clear as there are multiple factorsmicroglobulinquinolinicacid, matrix metalloproteinasegangiosides. ^[16] The progression of mildneurologic disorder is variable, sometimes patient suffers from mild dysfunction for a longerperiod of time and sometimes sever impairment occurs due to rapid progression ofneuronal dysfunction.Spatients ^[17]

Abnormally expressed protein as Biomarkers

The brain being the sanctuary of HIV, maintain a balanced immune privilege environment with neurotoxins as well as neuroprotective factors. Neopterin, neuro-metabolites such as glutamate, glutamine and glial metabolites like myo-inocytol and choline compounds were suggested to be CSF biomarker for neurological dysfunction.^[15] More importantly, neuroprotective factors such as BDNF, NT-3 used to get declined during neurocognitive impairment.^[18] Glycogen synthase kinase 3 beta level increases by any neurotoxins. GSK-3 beta inhibitors, phosphorylated form of GSK3 beta and 14-3-3 family of protein are used as neurological signature during CNS disease progression. Neuron specific enolase (NSE) and S100 beta, MBP and glial fibrillary acidic protein (GFAP) in CSF consider as putative marker of CNS disease progression in HIV patients.^[19]

Active HIV-related neurodegeneration can be correlated with neurofilament light chain (NFL) present in CSF. In HIV-1 subtype B infected patients, elevated level of neurofilament light chain in CSF have been reported by several investigators.^[20] Neopterin, the inflammatory biomarker of CNS immune activation have been reported in pre and post antiretroviral therapy by many groups.^[13], ^[21], ^[18] Unlike Alzheimer’s patients, the elevated level of t-tau protein in CSF of HAD patient is one of the defined pathways to neurological injury in brain.^[22] CSFMMP9 level increases in patients with opportunistic infections which indicate that the increase in MMP9 may degrade the extracellular matrix of blood brain barrier and trans-endothelial migration of virus may increase. In a collective study from four different broad groups (Milano, Sweden, California and Australia) before and after CART, ^[13] it has been reported that CSF neopterin is not simply a reflection of CNS inflammation; it may provide the degree of vulnerability of neural cells in brain.

Healthy individual contains 3- 5nmol/L neopterin in CSF, where as in HIV infected CART treated patients the level estimated to be 16-20nmol/L which indicates intrathecal immune activation.^[23] Often there is a good correlation between blood and CSF neopterin and study from a computed tomography demonstrated that significant correlation between blood neopterin concentration and loss of brain tissues. As many cytokines including IFN-γ have short half-lives, they might not be well reflected in CSF. In contrast, neopterin has long half-life and stability in biological fluid due to its non-polar chemical character; it appears to provide a stable indicator among HIV infected patients with progressive neurological diseases.

In a developing country like India, late diagnosis of HIV, due to irregular immuno-virological monitoring, the effect of opportunistic infections in patients with antiretroviral therapy has not been well defined. In addition, very few studies have been carried out in patients with opportunistic infections (OI). In the post HAART era, the neurological complications due to subtype B or C do not show much difference and report from clade B predominant countries follow the similar kind of observations. From the CNS HIV antiretroviral therapy effects research (CHARTER) studies, several reports suggested that although sever form of HAD drop down by antiretroviral therapy, milder form of cognitive impairment remain to be continued and prevalence of mild neurocognitive impairment increased in India ^[7], ^[15]. Most of the cognitive performances measured by international dementia scale is partly depends upon the educational status of HIV patients and it was found lower level of education has significant effect on the text performances. Neurocognitive impact of HIV-1 subtype C has been studied by various independent groups of south India and almost similar observation were found in the neuropsychological performances. Mild neurocognitive impairment is of 35.3 %, mild moderate impairment is 21.9% and moderate-sever impairment is 3.3% in the study sample ^[24]. In developed country, the milder forms are found as ANI 32.7%, MND 11.7% and HAD 2.4% and association of neuropsychological impairment correlate with sustained elevation of macrophage/microglia activation marker sCD163 in CSF and plasma.^[25]

Biomarkers of Subtype C intimately related to OI

In post-HAART era, apart from CD4 T-cell count, HIV viral load, quantization of immune activation marker in plasma, enormous amount of studies have been carried out in developed country to understand the neuropathological mechanism which may not be directly relevant to Indian scenario, because co-infections are intimately related to the immunological parameters of HIV patients in India and secondly opportunistic infections in CNS predominantly affect the subtype C associated neuropathogenesis. ^[5], ^[26] From a recent study of asymptomatic and symptomatic group of neurocognitive patients of subtype C HIV infected subjects from India, CSF biomarkers have been analyzed. As expected IL6, TNF alpha, beta-2 microglobulin and neopterin level are significantly higher in neurocognitive symptomatic groups in comparison to that of asymptomatic groups. ^[12] However, the specific parameter in asymptomatic groups which may contain the information about local pathogenic progression in CNS is lacking. Increased NFL in CNS has been predicted as vulnerability of CNS injury.^[22] Unfortunately, there is unavailability of this kind of study from Indian patients. Decline in certain neuro-protective factors like BDNF, NT-3 may explain the level of ongoing neurodegeneration in those asymptomatic groups which are worthwhile to predict the future neurological impairment. It is important to note that in addition to soluble CD14,CCl2, IL6 and general growth factors level may increase in CSF along with many other monocytic activation markers,^[12] but the decline in essential neurotrophic growth factors would make a special attention to identify a trend that give a signature molecule to predict the neuropathological status. Including viral and host response tissue regulatory CSF markers, many other CSF biomarkers have been reported in subtype B infected patients, but none of them systematically studied in drug naïve versus cART treated patient in India. Moreover, it is essential to consider the other associated neurological diseases in the patients as well as the immunological status if patient has more than one systemic opportunistic infection. Hence more CSF biomarkers need to assessed using new advanced techniques and these efforts are essential to understand the interaction of HIV-1 in brain for the longer period of stay. More importantly CSF NFL, neopterin, BDNF, NT-3 are the factors to examine as these CSF biomarkers might help to understand and monitor the immune reconstitution inflammatory syndrome (IRIS) and ongoing disease progression in brain.

Great potential of Biomarker for Neurodegenerative diseases

Biomarkers are believed to have great potential in early diagnosis of diseases, setting standard for developing new remedies to treat disease. To identify the biomarkers of several different neurodegenerative diseases, new advanced technology enabled the modern scientists. The biomarkers that have been recently recognised are phosphorylated tau protein and aggregated beta-amyloid peptide for Alzheimer’s Diseases, alpha-synuclein contained Lewy bodies and altered dopamine transporters. The light subunit of the neurofilament protein (NFL) is a major structural component of myelinated axons is essential to maintain axonal calibre and to facilitate effective nerve conduction, CSF concentrations of NFL provide a sensitive marker of CNS injury in a number of neurological diseases. In CSF, the light subunit of the neurofilament protein (NFL) concentrations across the spectrum of HIV-infection has been developed. ^[1], ^[7], ^[27]

Future prediction

CSF explores the possible global scenario of neuropsychological change. Novel biomarkers have been identified in CSF of the HIV infected patient with different opportunistic infection, prior to anti-retroviral and after anti-retroviral therapy. The significant differences in neurologically normal and declined patients are able to distinguish by biomarkers. The efforts of the critical evaluation of biomarkers lead to the identification of biomarkers of CART drug suppressive asymptomatic subjects, to delineate the neurological impairment mechanism from the various biochemical parameters. Also biomarkers are able to determine the increase in Blood brain barrier (BBB) permeability due to OIs and the impaired function of HBMECs, tight junction proteins and increase of matrix metalloproteinase altogether may allow the infiltration of activated monocytes to perform the oxidative damage to neuron as well as hamper the supportive activity of glial micro environment inside the CNS. Further, early CSF biomarkers are desperately needed to interpret CNS disease progression and for therapeutic intervention. The wide range CSF samples analysis may provide a trend to associate the disease progression at CNS level.