Research ProfileS
Proteome changes in HIVassociated dementia
therapy, which usually consists of a combination of antiretroviral agents, is working. “Currently, HAD therapy is monitored by neuropsychiatric evaluations,” Ciborowski says. “But it typically takes several months to see symptom improvement. We would like to have biomarkers to monitor the progress of therapy on a much shorter timescale.” To identify potential HAD biomarkers, Ciborowski and co-workers examined the CSF proteomes of nondemented (ND) HIV patients and those with HAD. Proteins and their fragments in CSF provide information on metabolic activity and pathological processes of
Cy3. Pairs of samples (HAD/ND) were run on 2DE gels, along with a pooled HIV-associated dementia (HAD), a sample containing equal amounts of neurological disorder that afflicts some both samples labeled with Cy2 for norpatients who have advanced HIV-1 malization. A total of 221 protein spots infections, is characterized by cognifrom the three gels were identified with tive, motor, and behavioral deficits. high confidence by LC/MS/MS. The HIV-1 virus is thought to enter To analyze CSF samples that had low the brain by infecting monocytes or concentrations of protein, Ciborowski macrophages, hijacking these immune and co-workers performed saturatedcells to penetrate the blood–brain barlabeling 2D DIGE to analyze six adrier. In the brain, the virus reprograms ditional protein samples (three HAD, infected macrophages, transforming three ND). Whereas minimal labeling the neuroprotective cells into chemifluorescently labels ~5% of proteins in cal-weapons factories that spew neurothe sample, saturated labeling adds toxic immunological and viral proteins. fluorophores to ~100% of proteins. This In addition, HIV-1 infects and procedure, which is used infrecauses the self-destruction of quently, allows smaller protein astroctyes, cells that supply nuamounts to be detected. PorpH 3.0 pH 10.0 trients to neurons. The resulttions of the six samples were 98 kDa ing neuronal damage induces labeled with Cy5 by saturated 62 kDa symptoms that range from mild labeling and run on separate 49 kDa cognitive or motor impairment 2DE gels, along with a pooled, to severe debilitation and death. Cy3-labeled internal control. The precise molecular etiology Because the same internal of HAD is unknown. control was present on all of 21 kDa To better understand HAD the gels, protein spots could progression, Pawel Ciborowski be matched and compared and co-workers at the Univer across multiple gels. For MS 14 kDa sity of Nebraska Medical Center, analysis, the remainders of the the University of Puerto Rico six samples were pooled and Medical Sciences, and the labeled with Cy3 for a single National Veterinary Research preparative gel. With LC/MS/ Identified proteins. Representative 2D DIGE gel (minimal labelInstitute (Poland) compared MS, the researchers identified ing) of CSF samples from HIV-1 patients with or without HAD. Circled spots indicate proteins identified by LC/MS/MS. the cerebrospinal fluid (CSF) 52 proteins that were differenproteomes of HIV-1-infected tially expressed in HAD and patients with or without HAD. ND. Differential expression of They report their results in this issue of the central nervous system. However, six of the identified proteins from both JPR (pp 4189−4199). proteomics analyses on CSF are chalexperiments was verified by western Because of a lack of conclusive lenging, because of the low protein blot analysis. diagnostic tests, HAD is currently a concentration (0.2−0.8 mg/mL; ~100× Although the differentially expressed diagnosis of exclusion made when less than that in serum) and the high proteins represent potential HAD bioopportunistic infections, psychiatric abundance of albumin, transferrin, markers, Ciborowski says that the disorders, and malignancies have been and immunoglobulins (which together results must be confirmed in a larger ruled out. Neuropsychiatric examinacompose >70% of CSF protein) in the cohort of CSF samples; these experitions combined with progressive imfluid. ments are in progress. Further studies mune suppression, high viral load, and The researchers used 2D difference also are needed to clarify whether cognitive dysfunction support a diaggel electrophoresis (2D DIGE) and LC/ differential expression of proteins in nosis of HAD. According to Ciborowski, MS/MS peptide sequencing to compare HAD reflects HIV-specific processes or “HAD biomarkers have been sought by the CSF proteomes of ND and HAD general neuroinflammation. “Some of different approaches, such as examinpatients. CSF samples were depleted the differentially expressed proteins in ing cytokine profiles, CD4 T-cell count, of the six most abundant proteins. The HAD are also differentially expressed levels of monocyte chemotactic protein depletion was performed with a mulin other neurodegenerative disorders, 1 (MCP1), viral load, and many others. tiple-affinity column. The researchers such as Alzheimer’s and Parkinson’s,” However, none of the potential HAD found that only 6 of the 38 CSF samples Ciborowski says. “I don’t believe that biomarkers has been able to predict the (3 HAD and 3 ND) contained sufficient there will be one silver-bullet biomarkonset or tempo of HAD, which is critical protein for analysis. Protein samples er for HAD. Future diagnoses will likely for the right treatment regime.” from patients with HAD were minimalinvolve a large panel of biomarkers in Reliable HAD biomarkers would ly labeled with Cy5 dye, and samples conjunction with psychiatric tests.” help physicians assess whether HAD from ND patients were labeled with Laura Cassiday
4106 Journal of Proteome Research • Vol. 6, No. 11, 2007
