Fig 1: Proposed model of FGFR activation pathways in response to live B. burgdorferi exposure (a) and intersectionality of the induced and downregulated FGFs with other neurological conditions (b). a Exposure of primary rhesus microglia to live B. burgdorferi upregulates the surface expression of FGFR1, FGFR2 and FGFR3 [1]. Host–pathogen interaction also induces expression of several FGFs such as FGF6, FGF10, FGF12, and FGF23, of which FGF6 (and likely FGF10 and FGF23) are secreted from the cells [2]. Whether FGF12 is secreted extracellularly is unclear. Ligand binding of FGF6 to FGFR1 induces phosphorylation of the receptor [3] and secretion of IL-6 and CXCL8. The intracellular signaling pathway is likely through MAPK pathways, particularly ERK, as have been demonstrated in our previous study in primary rhesus microglia [22]. While FGF6 was shown to activate FGFR1 in this study, it can also activate other FGFRs. Similarly, FGF10, shown to activate FGFR2 in the model, can also activate FGFR1, while FGF23 can activate FGFR3, FGFR2 and FGFR1 [86]. As FGF6, 10 and 12 only activated IL-6 and/or CXCL8, but the inhibition of FGFR1 individually by siRNA downregulated IL-6, CXCL8 as well as CCL2, it is likely that other than FGF23, non-FGF molecules present in the supernatant also likely activate this receptor. It should be noted that only autocrine effects of FGF binding FGFRs in microglia are shown. It is possible that some paracrine effects on other glial cells also occur and will be tested in future studies. Finally, our study also demonstrated that synthesis (or inhibition) of FGFs (except for FGF8, 23, and 16 & 21 to an extent) was also through FGFR1 (Additional file 1: SM4), as suppression of FGFR1 signaling with PD166866 modulated FGF levels [4]. b shows the known neurological roles of the FGFs from this study and others. [−] indicates (putative) negative roles, while [+] indicates (putative) positive effects of the indicated FGFs. The listed roles are not exhaustive. Please see the Discussion section for details. Upregulation of FGFs with deleterious effects, and downregulation of FGFs with ameliorative effects can contribute towards Lyme neuroborreliosis sequelae and other neuropathologies. Both figures created with BioRender.com
Fig 2: Effect of exogenous addition of FGFs on inflammatory mediator output and activation of FGFR1 pathway on primary rhesus microglia. a Various concentrations of recombinant human FGFs were added to enriched primary rhesus microglial (~ 80%) cells for 24 h. PBS/BSA (0.1%) was used as the solvent control. Supernatants were collected and analyzed for IL-6, CXCL8 and CCL2 expression by Multiplex ELISA. Lines within each cytokine/chemokine indicate that they were analyzed separately. 5 ng/ml data is representative of 2 experiments conducted on microglia derived from one frontal cortex tissue, while the higher concentration is representative of 2 experiments conducted on microglia derived from 2 different frontal cortex tissues. Data shown are from experiments that were performed with the same animal tissue. Bar represents standard deviation. Black asterisks represent statistically significant increase over PBS/BSA control. *p < 0.05, **p < 0.01, and ***p < 0.001. b Shows activation of FGFR1 pathway by addition of 5 ng/ml of FGF6 (+ DMSO) to primary rhesus microglial cells. Upregulation of pFGFR1 (green) is seen in cells that also stain for Iba1 (red). Bar represents 50 µm. Panel on the far-right shows the same data at a higher magnification (Bar represents 25 µm). c Shows the effect of PD166866 FGFR1 inhibitor on the inflammatory output in response to exogenous addition of FGF6. A representative experiment is shown of 2–3 experiments carried out on microglia derived from 2 different tissues. Black asterisks represent statistical differences in comparison to FGF6/DMSO control. **p < 0.01
Fig 3: Expression of specific upregulated FGFs in primary rhesus microglia in response to B. burgdorferi. The expressions of FGF6, FGF10, FGF12 and FGF23 was analyzed by immunofluorescence in primary rhesus microglia. a Shows immunofluorescent microscopy pictures of the specific FGFs upregulated (green) in response to the Lyme disease bacterium. The nuclei stained with DAPI is shown in blue. Representative pictures from 2 (FGF10, FGF23) to 3 (FGF6, FGF12) experiments are shown. Bar represents 50 µm. Panels on the far right in a are higher magnification images of FGF staining. Bar represents 25 µm. b Shows confocal microscopy pictures of the same FGFs (green) to be microglia specific by staining for Iba1 in red. Nuclear stain is in blue
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