Non-classical monocytes predict progression of carotid artery bifurcation intima-media thickness in HIV-infected individuals on stable antiretroviral therapy

Figures & data

Figure 1 Multiparametric flow cytometry gating strategy to phenotype four distinct monocyte sub-populations from peripheral blood based on CD16 and CD14 expression: (1) classical monocytes lacking CD16 expression (CD14++CD16-) and those expressing CD16 comprised (2) intermediate (CD14++CD16+) and (3) non-classical (CD14low/+CD16++) monocytes. A fourth MO subset which we have termed (4) “transitional” monocyte subset is characterized by reduced but still detectable levels of CD14 (CD14 + CD16-).

Table 1 Baseline characteristics. Values reported as median (Q1, Q3), except for frequency count, n (%)

	All Subjects (N = 50)
Variables
Demographics
Age, median years (interquartile range)	49 (46, 56)
Male, n (%)	42 (84%)
Ethnicity, White, n (%)	27 (54%)
Clinical Data
BMI, kg/m^2	26 (25, 29)
hsCRP, mg/L	0.9 (0.6, 2.1)
Framingham risk score (ATP 3)	0.045 (0.02, 0.10)
Framingham risk class
Low, n (%)	38 (76)
Intermediate, n (%)	4 (8)
High, n (%)	8 (16)
History of CVD event, n (%)	6 (12)
Metabolic syndrome, n (%)	8 (16)
Systolic blood pressure, median mmHg (IQR)	122 (116, 133)
Total cholesterol, median mg/dL (IQR)	181 (155, 209)
HDL cholesterol, median mg/dL (IQR)	46 (34, 57)
LDL cholesterol, median mg/dL (IQR)	115 (97, 132)
Triglyceride, mg/dL (IQR)	106 (77, 167)
Diabetes, n (%)	1 (2)
Current smoker, n (%)	5 (10)
Protease Inhibitor use	26 (52)
Current Atazanavir use, n (%)	16 (32)
HIV data
Nadir CD4 cell count cells/mmᶟ	135 (29, 235)
Undetectable plasma HIV-1 RNA, n (%)	42 (84)
CD4+ T cell count, median cell/ul	461 (317, 578)
Carotid Intima-media Thickness (cIMT) measures
Right Common Carotid, mm	0.73 (0.66, 0.83)
Right Bifurcation, mm	0.80 (0.73, 0.87)
Maximal plaque thickness, mm	2.89 (2.48, 3.39)
Number of plaques (plaque > 2 mm), n (%)	9 (18)
Biomakers
hsCRP, ng/mL (IQR)	0.60 (0.90, 2.10)
Log IFN-γ, pg/mL (IQR)	−0.21 (–0.44, 0.02)
Log IL-6, pg/mL (IQR)	0.14 (–0.07, 0.38)
Log IL-8, pg/mL (IQR)	0.54 (0.44, 0.63)
Log IL-10, pg/mL (IQR)	0.21 (–0.04, 0.65)
Log MCP-1, pg/mL (IQR)	2.13 (2.04, 2.21)
Log MMP-9, ng/mL (IQR)	1.71 (1.54, 1.86)
Log E-selectin, ng/mL (IQR)	1.49 (1.30, 1.66)
Log sICAM-1, ng/mL (IQR)	2.12 (2.03, 2.19)
Log sVCAM-1, ng/mL (IQR)	3.07 (3.00, 3.11)
Log TNF-α, pg/mL (IQR)	0.50 (0.27, 0.63)
log_tPAI-1, ng/mL (IQR)	1.96 (1.86, 2.06)
CD8 Activation %	10.5 (8.1, 16.7)
CD8 Activation count, cells/uL (IQR)	88 (39, 113)
log CD8 Activation count	1.95 (1.59, 2.05)
Monocyte Sub-Types
Classical, % (IQR)	78.3 (72.6, 82.5)
Intermediate, % (IQR)	1.36 (0. 57, 4.41)
Transitional, % (IQR)	13.6 (10.5, 15.6)
Non-Classical, % (IQR)	5.44 (4.48, 8.61)
log Absolute total monocyte count (cells/L)	8.62 (8.51, 8.71)
log Classical (CD14++CD16^–)	8.51 (8.36, 8.62)
log Intermediate (CD14++CD16+)	6.82 (6.38, 7.19)
log Transitional (CD14+CD16^–)	7.76 (7.61, 7.89)
log Non-classical (CD14+/lowCD16++)	7.33 (7.16, 7.48)

Table 2 Correlation of monocyte and monocyte subtypes with carotid intima-media thickness, baseline and year 2

	All Monocytes (from CBC)	Classical Monocytes, % (CD14++CD16^–)	Intermediate Monocytes, % (CD14++CD16+)	Transitional Monocytes, % (CD14+CD16-)	Non-Classical Monocytes, % (CD14+/lowCD16++)
CIMT at bifurcation region (BIF)
CIMTBIF, baseline	0.224	−0.270	0.181	0.308	−0.105
CIMTBIF, year 2	0.221	−0.277	0.187	0.300	−0.078
Change in CIMTBIF	−0.015	−0.106	0.086	−0.086	0.355*
CIMT at the common carotid artery (CCA)
CIMTCCA, baseline	0.113	−0.300	0.067	0.366*	−0.042
CIMTCCA, year 2	0.137	−0.278	0.070	0.350*	−0.066
Change in CIMTCCA	0.075	0.155	0.038	−0.096	−0.230

* p < 0.05.

Table 3 Correlation between monocyte subtypes and biomarkers

	log hsCRP	log IFN-γ	log IL-10	log IL-6	log IL-8	log MCP-1	log MMP-9	log selectin	log sICAM-1	log sVCAM-1	log TNF-α	log tPa-1
CIMTCCA, Baseline	0.181	−0.179	−0.157	0.044	0.149	0.030	0.058	0.068	−0.116	0.327*	0.079	0.114
CIMTCCA, Year 2	0.123	−0.169	−0.132	0.039	0.145	0.025	0.045	0.044	−0.130	0.346*	0.070	0.080
CIMTCCA, Change	−0.159	0.134	0.202	−0.046	−0.089	−0.039	−0.089	−0.152	−0.025	−0.049	−0.079	−0.225
CIMTBIF, Baseline	0.214	−0.056	−0.017	0.006	0.002	−0.072	0.080	−0.142	−0.168	0.160	0.035	0.034
CIMTBIF,Year 2	0.173	−0.043	−0.013	0.006	−0.008	−0.041	0.080	−0.142	−0.161	0.176	0.056	0.016
CIMTBIF, Change	−0.257	0.174	0.053	0.008	−0.133	0.421*	0.006	−0.022	0.082	0.240	0.298*	−0.248

* p < 0.05.

Table 4 Multivariable linear regression analysis of the association of non-classical monocyte subset percentage and MCP-1 to the change of CIMT_BIF from baseline adjusting for Framingham Risk Score and baseline CIMT_BIF

	Regression coefficient	Standard Error	T-value	p-value	95% Confidence Interval
Regressions by individual biomarker
CIMTBIF, baseline	0.005	0.012	0.45	0.652	−0.018	0.029
Framingham Risk Score	0.026	0.027	0.98	0.336	−0.028	0.080
Non-classical monocyte, %	0.132	0.058	2.30	0.028*	0.016	0.249
CIMTBIF, baseline	0.010	0.010	0.98	0.331	−0.010	0.029
Framingham Risk Score	−0.002	0.024	−0.08	0.939	−0.050	0.047
MCP-1	0.031	0.009	3.24	0.002*	0.012	0.050
Regression by both biomarkers
CIMTBIF, baseline	0.010	0.011	0.90	0.374	−0.012	0.032
Framingham Risk Score	0.023	0.025	0.93	0.361	−0.027	0.073
Non-classical monocyte, %	0.087	0.056	1.56	0.128	−0.026	0.201
MCP-1	0.025	0.010	2.63	0.013*	0.006	0.045

* p < 0.05.