Evaluating the immuno-expression patterns of HHV-6 and CD68 in Substantia Nigra in the background of excessive alcohol use: a histo-pathological analysis

Background. HHV-6 (Human Herpesvirus 6) is an omnipresent double stranded DNA virus that can reside in multiple organs including the brain. The virus has been previously shown to infect vascular cells, thereby evoking an immune response. Furthermore, excessive alcohol use has also been associated with endothelial cell dysfunction. The role of activated microglia as protector of homeostasis has been long established. Hence, it becomes essential to investigate the potential synergistic role of the HHV-6 infection and excessive alcohol intake in the dysregulation of homeostasis leading to cellular immune response mediated by activated microglia.

Aim. The present study aimed to evaluate HHV-6 antigens in micro-vessel walls and activated microglia/macrophages in the Substantia Nigra (SN) in the setting of the excessive alcohol use.

Methods. Grey and white matter in the SN region from 14 brain autopsy samples with history of alcohol addiction (alcoholics) were investigated in the present study, which were age-matched with 15 non-alcoholics (controls). Quantitative estimation of the HHV-6 positive micro-vessels was done using anti-HHV-6 antibody along with activated microglia/macrophages using anti-CD68 antibody. The immuno-expression was analysed using a light microscope (40×) and the results were analysed using Excel and R.

Results. The median age in the alcoholics group was 33.5±6.17 years compared with 26±5.75 years in the controls. HHV-6+ blood vessels were observed in 8 samples – 4 each in alcoholics and controls. Significantly more (p < 0.05) HHV-6+ micro-vessels were found in both grey and white matter in the alcoholics when compared with controls. More CD68+ cells were observed peri-vascularly in the alcoholics when compared with controls in the white matter while controls showed more CD68+ cells peri-vascularly in the grey matter. However, when adjusted for number of activated microglia/macrophages per 10 vessels per visual field, higher number of activated microglia were noted for alcoholics in both grey (10 activated microglia/10 vessels/visual field in alcoholics compared with 9 activated microglia/10 vessels/visual field) and white matter (11 activated microglia/10 vessels/visual field in alcoholics compared with 9 activated microglia/10 vessels/visual field).

Conclusion. An increased presence of CD68+ cells per 10 vessels per visual field peri-vascularly coupled with a higher number of HHV-6+ vessels in the alcoholics group indicates a role of both excessive alcohol use and HHV-6 infection in mediating the immune response via activation of perivascular microglia.

Acknowledgements. The authors declare the absence of conflict of interest. The present study was funded by FLPP project No. 2020/2-0069