Purpose of Study Children infected with the human immunodeficiency virus, type 1 (HIV-1) have developmental delays due to viral infection of the central nervous system. While the exact mechanisms are still unclear, HIV-1 proviral DNA (HIV DNA) and monocyte chemoattractant protein 1 (MCP1) may play a role, neither of which have been studied extensively in children. Even with effective antiretroviral therapy, HIV DNA is still present, which may facilitate neurotoxicity. In conjunction, mutations in the MCP1 gene are associated with high MCP1 levels, which may further attract activated macrophages to the CNS having HIV DNA leading to further CNS toxicity. The objective of the study was to measure HIV DNA and characterize MCP1 genotypes in cerebrospinal fluid (CSF) from HIV-1-infected children. The significance of these parameters is that they may identify potential targets for intervention to improve growth and development in children, who are significantly affected by the virus.
Methods DNA was purified from CSF obtained from HIV-1-infected children as per Institutional Review Board guidelines. HIV DNA copy was measured by real-time PCR using standards set up with HIV GAG and β-globin primers; and quantified as HIV DNA copies per 106 cells. MCP1 genotype was assessed by amplifying a 361 bp region of the MCP1 gene and analyzing for a point mutation identified by the restriction site, PvuII, due to the single nucleotide polymorphism (SNP) at position 2578.
Results Good quality DNA was purified from 27 pediatric CSF specimens. By real-time PCR, all of the samples had detectable HIV DNA ranging from 4.0 × 10-3 to 9.4 × 10-6 copies/106 cells. MCP1 SNP analysis for the A2578G allele showed the mutation in 10 samples, 14 without the mutation, 3 were indeterminate.
Conclusion In this feasibility study, we showed for the first time a range of HIV DNA in CSF from HIV-1-infected children. We also demonstrated that the MCP1 genetic allele can be characterized from CSF cells. This study provides data to support measuring HIV DNA and assessing MCP1 genotype in CSF from HIV-1-infected children. The significance of this study is that the assays could potentially be incorporated in treatment and/or intervention strategies to compare CSF HIV DNA and MCP1 mutations to neurocognition in HIV-1-infected children. Both factors may be potentially important in growth and development of children infected with HIV-1.
Supported in part by grants P20RR16467; U54NS43049; G12RR03061; MH69173; RR03010; and U01A134853N.
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