Figure 5.
Dysregulated activation status and phagocytic activity in monocytes from participants with the LGR6 frameshift variant. Whole blood from participants from the variant (LGR6 variant) and control groups was incubated with antibodies against lineage markers to identify classical (A; CD14++CD16+), intermediate (B; CD14++CD16++) and nonclassical (C; CD14+CD16++) monocytes, and the expression of activation markers was determined using flow cytometry and fluorescently conjugated antibodies to these proteins. PLS-DA analysis (top) and VIP scores (bottom) based on circulating phagocyte activation markers in monocytes. (D) Monocytes from participants from the variant and control groups were isolated from whole blood and incubated with pHrodo Green S aureus BioParticles, and phagocytosis was assessed in real-time using high-content imaging. S aureus phagocytosis is expressed as the AUC of the increase in pHrodo Green signal over time divided by the number of cells. (E-F) Monocytes were incubated with 1 nM of MaR1 or vehicle (0.001%-0.01% ethanol) for 15 minutes followed by pHrodo Green S aureus BioParticles, and phagocytosis was assessed. (E) Time course of S aureus phagocytosis (expressed as the percentage of change from the vehicle) of monocytes when incubated with MaR1. (F) Spearman correlation analysis comparing LGR6 expression (percentage of cells expressing the receptor) against S aureus phagocytosis (AUC of the percentage of change from the vehicle over time). Yellow and green dots represent participants with or without the LGR6 frameshift variant, respectively; and the light blue background indicates a confidence interval of 95%. R and P values of the correlation are displayed. Statistical differences between variant and control were determined using the 1-tailed Mann-Whitney test, and P values are displayed. For panels A-C, results are representative of 3 for variants and 4 for controls. For panel D, results are representative of 8 for variants and 8 for controls. For panels E-F, results are representative of 7 for variants and 8 for controls.

Dysregulated activation status and phagocytic activity in monocytes from participants with the LGR6 frameshift variant. Whole blood from participants from the variant (LGR6 variant) and control groups was incubated with antibodies against lineage markers to identify classical (A; CD14++CD16+), intermediate (B; CD14++CD16++) and nonclassical (C; CD14+CD16++) monocytes, and the expression of activation markers was determined using flow cytometry and fluorescently conjugated antibodies to these proteins. PLS-DA analysis (top) and VIP scores (bottom) based on circulating phagocyte activation markers in monocytes. (D) Monocytes from participants from the variant and control groups were isolated from whole blood and incubated with pHrodo Green S aureus BioParticles, and phagocytosis was assessed in real-time using high-content imaging. S aureus phagocytosis is expressed as the AUC of the increase in pHrodo Green signal over time divided by the number of cells. (E-F) Monocytes were incubated with 1 nM of MaR1 or vehicle (0.001%-0.01% ethanol) for 15 minutes followed by pHrodo Green S aureus BioParticles, and phagocytosis was assessed. (E) Time course of S aureus phagocytosis (expressed as the percentage of change from the vehicle) of monocytes when incubated with MaR1. (F) Spearman correlation analysis comparing LGR6 expression (percentage of cells expressing the receptor) against S aureus phagocytosis (AUC of the percentage of change from the vehicle over time). Yellow and green dots represent participants with or without the LGR6 frameshift variant, respectively; and the light blue background indicates a confidence interval of 95%. R and P values of the correlation are displayed. Statistical differences between variant and control were determined using the 1-tailed Mann-Whitney test, and P values are displayed. For panels A-C, results are representative of 3 for variants and 4 for controls. For panel D, results are representative of 8 for variants and 8 for controls. For panels E-F, results are representative of 7 for variants and 8 for controls.

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