Neuropathogenesis of Chikungunya infection: astrogliosis and innate immune activation

Abstract

Chikungunya, "that which bends up" in the Makonde dialect, is an emerging global health threat, with increasing incidence of neurological complications. Until 2013, Chikungunya infection had been largely restricted to East Africa and the Indian Ocean, with cases within the USA reported to be from foreign travel. However, in 2014, over 1 million suspected cases were reported in the Americas, and a recently infected human could serve as an unwitting reservoir for the virus resulting in an epidemic in the continental USA. Chikungunya infection is increasingly being associated with neurological sequelae. In this study, we sought to understand the role of astrocytes in the neuropathogenesis of Chikungunya infection. Even after virus has been cleared form the circulation, astrocytes were activated with regard to TLR2 expression. In addition, white matter astrocytes were hypertrophic, with increased arbor volume in gray matter astrocytes. Combined, these would alter the number and distribution of synapses that each astrocyte would be capable of forming. These results provide the first evidence that Chikungunya infection induces morphometric and innate immune activation of astrocytes in vivo. Perturbed glia-neuron signaling could be a major driving factor in the development of Chikungunya-associated neuropathology.

Keywords: Alphavirus; Chikungunya; Gliosis; Morphometry; Togavirus; Toll-like receptor.

Figure 1. Chikungunya infection induces qualitative changes in TLR2 expression

Control cynomolgus macaques (A) had diffuse expression of TLR2 (green), mainly on astrocytes (red). Following subcutaneous infection with Chikungunya, it TLR2 expression became more pronounced, oftentimes in a perinuclear location (B).

Figure 2. Chikungunya infection affects TLR2 expression on astrocytes

There were no significant differences in GFAP-expression in gray and white matter astrocytes of control animals and those infected with Chikugunya (A&B). Quantifying the subset of GFAP-positive astrocytes that were also positive for TLR2, we found no changes in double-labeled astrocytes after infection (C&D). There was an increase the total area of TLR2 expression in the gray matter of infected animals (E). Data is shown as mean ± SEM. Significance is set at *p < 0.05.

Figure 3. Cell body hypertrophy of white matter astrocytes in macaques infected with Chikugunya virus

There was cell body hypertrophy in white matter astrocytes of animals infected with Chikigunya virus (B). There were no differences in total arbor length (C, D) and surface area (not shown) observed between Chikugunya-infected animals and controls. However, total arbor volume was increased in gray matter astrocytes of infected animals indicating hypertrophy or swelling of the processes (E). No significant changes were observed in the total arbor volume in white matter astrocytes (F). This hypertrophy was mainly confined to the first three branches (G). Data is shown as mean ± SEM. Significance is set at *p < 0.05.

Figure 4. Decreased complexity of gray matter astrocytes in Chikugunya-infected macaques

There was no change in the number of processes leaving the cell bodies in either gray (A) or white matter (B) astrocytes. Gray matter astrocytes in CHIK-infected cynomolgus macaques had fewer bifurcations (C) and end points in their processes (E). These differences were no observed in white matter astrocytes (D&F). The decreased branching appeared to cluster between third and sixth order branches in gray matter (G). Data is shown as mean ± SEM. Significance is set at *p < 0.05.