Background
Methods
Design and data collection
Flow cytometric analysis of peripheral blood lymphocytes
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B cells subsets: anti- CD19 (ref: A07766), -CD27 (ref: 6607107), -CD21 (ref: PN IMU473U) and -sIgD (ref: 736000)
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CD8+ T-cell differentiation: anti-CD8 (ref: A07756), -CD45RA (ref: PN IM 271U), -CCR7 from R&D (Minneapolis, MN, USA ref: FAB197A)
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HLA-DR T-lymphocyte activation markers, we used the following panel of Mab: anti-CD45, -3, -4 and -8 (ref: 6607013) -HLA-DR (ref: PNA40579).
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CD25 T-lymphocyte activation marker: anti-CD3, -4 and -8 (ref: 6607013) -CD25 (ref: IM2646)
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CD28 T-lymphocyte activation marker: anti-CD3, -4 and -8 (ref: 6607013) -CD28 (ref: 6607108)
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CD38 T-lymphocyte activation marker: anti-CD3, -4 and -8 (ref: 6607013) -CD38 (ref: A07780)
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regulatory T-cell: anti-CD4 (ref: A07750), -CD25 (ref: IM2646) -CD127 (ref: PN IM1980U)
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NK and B cells: anti-CD45, -56, -19 and -3 (ref: 6607073) -CD16 (ref: A07766)
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T cells expressing the gamma/delta TCR: anti-Vdelta2 (ref: PN IM1464) and anti-pan-delta (ref: PN IM1418U).
Statistical analyses
Principal component analysis
Cluster analysis
PHID complication analyses
CVID substudy
Results
Patient characteristics
Sociodemographic and clinical characteristics
Characteristics | Distribution | |
---|---|---|
Sociodemographic data
| ||
Female, n (%) | 48 | (59) |
Median age at diagnosis in years (interquartile range) | 41 (35–54) | |
Median age at enrolment in years (interquartile range) | 46 (38–58) | |
Clinical data, n (%)
| ||
Respiratory tract infections
| ||
Otitis, sinusitis, nasopharyngitis, polyposis | 64 | (79) |
Bronchitis, pneumonia | 65 | (80) |
Bronchiectasis | 21 | (26) |
Gastrointestinal tract infection
| ||
Chronic diarrhea | 16 | (20) |
Acute diarrhea | 13 | (16) |
Lambliasis | 11 | (14) |
PHID complications: at least one of the following
| 31 | (38) |
Lymphoid hyperplasia | 16 | (20) |
Splenomegaly | 14 | (17) |
Autoimmune manifestations* | 14 | (17) |
Granulomatous disease† | 9 | (11) |
Villous atrophy | 5 | (6) |
Chronic inflammatory intestinal disease | 4 | (5) |
Therapeutic data, n (%)
| ||
Subcutaneous or intravenous immunoglobulin substitution | 66 | (81) |
Biological characteristics
Unit | Median | IQR | ||
---|---|---|---|---|
Total circulating lymphocytes (TCL)
| cells/mm3 | 1384 | (1017, 2000) | |
B cells
| CD19+ | cells/mm3 | 128 | (62, 193) |
% TCL | 9.37 | (6.21, 12.91) | ||
Naïve | CD27-IgD+ | % CD19+ | 74.01 | (56.42, 83.35) |
Switched | CD27-IgD- | % CD19+ | 2.62 | (1.86, 5.16) |
Marginal zone | CD27 + IgD+ | % CD19+ | 9.42 | (4.70, 15.37) |
Switched memory | CD27 + IgD- | % CD19+ | 8.07 | (3.63, 19.45) |
T cells
| CD3+ | cells/mm3 | 1078 | (746, 1585) |
CD4+ | cells/mm3 | 657 | (422, 941) | |
CD8+ | cells/mm3 | 329 | (228, 524) | |
CD4+/CD8+ ratio | 1.99 | (1.30, 2.70) | ||
CD8+ T cells
| ||||
Naïve | CD45RA + CCR7+ | % CD8+ | 18.01 | (6.79, 36.07) |
Central memory | CD45RA-CCR7+ | % CD8+ | 1.25 | (0.59, 2.48) |
Effector memory | CD45RA-CCR7- | % CD8+ | 34.39 | (24.47, 46.90) |
Terminal effector | CD45RA + CCR7- | % CD8+ | 33.48 | (23.31, 48.26) |
Immunosenescent | CD8 + CD57+ | % CD8+ | 15.97 | (8.04, 31.75) |
Activated T cells
| ||||
HLA-DR | CD3 + HLA-DR+ | % CD3+ | 12.87 | (7.99, 20.37) |
CD4 + HLA-DR+ | % CD4+ | 7.96 | (5.14, 13.79) | |
CD8 + HLA-DR+ | % CD8+ | 24.49 | (14.31, 41.11) | |
Regulatory T cells
| CD4 + CD25 + CD127- | cells/mm3% TCL | 32 1.10 | (16, 63) (0.51, 1.76) |
Natural Killer cells
| CD3-CD16 + CD56+ | cells/mm3 | 126 | (75, 194) |
Dendritic cells
| ||||
Myeloid | mDC | /ml | 9381 | (6597, 15611) |
Plasmacytoid | pDC | /ml | 4671 | (2810, 7427) |
Gammadelta cells
| cells/mm3 | 33 | (15, 72) | |
Gammadelta 2 cells | % GD | 56.32 | (22.19, 77.17) |
Principal component analysis of differentiation, activation and regulation markers of B and T-lymphocytes, natural killer cells, dendritic cells and T-cells expressing a gamma/delta TCR
Classification of PHID patients according to cluster analysis
Cluster 1 | Cluster 2 | Cluster 3 | Cluster 4 | Cluster 5 | ||
---|---|---|---|---|---|---|
n = 33 | n = 18 | n = 4 | n = 8 | n = 16 | ||
IgG | g/L | 4.37 (3.80, 5.32) | 5.73 (4.7, 6.3) | 4.68 (2.69, 5.29) | 1.44 (0.30, 2.71) | 3.69 (1.65, 4.22) |
IgA | g/L | 0.63 (0.14, 1.62) | 1.02 (0.07, 1.55) | 0.60 (0.04, 2.09) | 0.15 (0.01, 0.43) | 0.19 (0.05, 0.79) |
IgM | g/L | 0.55 (0.31, 1.09) | 0.48 (0.24, 0.78) | 0.54 (0.34, 0.62) | 0.16 (0.05, 0.42) | 0.27 (0.20, 0.47) |
B cells
| ||||||
CD19+ | cells/mm3 | 157 (128, 244) | 128 (102, 211) | 49 (20, 87) | 122 (94, 176) | 51 (29, 119) |
CD27-IgD+ | % CD19+ | 68.85 (52.03, 74.47) | 68.89 (57.94, 77.24) | 49.52 (10.73, 80.43) | 90.89 (82.94, 93.31) | 84.03 (70.93, 90.82) |
CD27-IgD- | % CD19+ | 2.80 (2.18, 4.38) | 3.90 (2.24, 6.42) | 6.13 (2.64, 23.40) | 2.40 (1.67, 2.58) | 2.65 (1.72, 4.22) |
CD27 + IgD+ | % CD19+ | 15.01 (10.11, 23.19) | 10.53 (4.42, 13.73) | 5.67 (2.43, 8.57) | 3.57 (2.21, 7.20) | 4.92 (1.32, 7.92) |
CD27 + IgD- | % CD19+ | 14.39 (7.08, 20.05) | 13.93 (7.26, 24.33) | 7.77 (2.43, 19.38) | 2.55 (0.94, 3.91) | 2.97 (0.36, 6.66) |
T cells
| ||||||
CD3+ | cells/mm3 | 1363 (1076, 1787) | 1045 (883, 1456) | 2756 (1472, 4183) | 831 (658, 1075) | 579 (407, 837) |
CD4+ | cells/mm3 | 835 (571, 1214) | 789 (573, 967) | 848 (609, 1227) | 382 (302, 565) | 308 (244, 565) |
CD8+ | cells/mm3 | 477 (358, 604) | 270 (159, 383) | 1621 (710, 2682) | 293 (196, 362) | 213 (141, 266) |
CD4+/CD8+ ratio | 1.89 (1.45, 2.42) | 2.77 (2.31, 4.08) | 0.60 (0.42, 1.06) | 1.26 (0.67, 2.65) | 1.89 (1.17, 2.61) | |
CD8+ T cells
| ||||||
CD45RA + CCR7+ | % CD8+ | 19.05 (11.11, 34.59) | 28.47 (13.32, 43.67) | 1.52 (0.56, 2.75) | 4.91 (1.53, 53.19) | 35.56 (10.29, 53.19) |
CD45RA-CCR7+ | % CD8+ | 0.83 (0.46, 1.90) | 4.60 (1.96, 7.97) | 0.53 (0.21, 0.82) | 0.82 (0.30, 1.20) | 1.37 (0.85, 1.96) |
CD45RA-CCR7- | % CD8+ | 31.28 (24.26, 36.96) | 43.13 (32.60, 57.14) | 36.82 (16.56, 57.25) | 39.24 (30.69, 59.20) | 29.14 (17.47, 42.55) |
CD45RA + CCR7- | % CD8+ | 41.48 (32.61, 52.32) | 22.30 (16.87, 27.10) | 40.75 (13.96, 59.90) | 48.28 (30.47, 63.41) | 29.25 (18.76, 39.37) |
CD8 + CD57+ | % CD8+ | 17.52 (9.42, 30.97) | 7.38 (4.41, 15.97) | 27.36 (19.36, 47.30) | 39.61 (33.78, 51.16) | 13.51 (8.35, 23.44) |
Activated T cells
| ||||||
CD3 + HLA-DR+ | % CD3+ | 12.04 (7.62, 16.95) | 9.87 (7.11, 12.83) | 52.00 (32.71, 59.56) | 46.56 (40.76, 57.25) | 13.65 (6.19, 20.33) |
CD4 + HLA-DR+ | % CD4+ | 5.88 (4.94, 9.17) | 6.75 (4.95, 9.13) | 48.30 (26.32, 56.25) | 43.87 (30.88, 49.95) | 11.54 (6.52, 18.33) |
CD8 + HLA-DR+ | % CD8+ | 25.48 (14.86, 31.93) | 19.73 (14.18, 26.44) | 53.48 (37.05, 65.52) | 62.14 (47.01, 76.39) | 15.12 (10.65, 28.64) |
Regulatory T cells
| cells/mm3 | 54 (33, 76) | 41 (24, 82) | 18 (13, 22) | 19 (12, 24) | 15 (8, 18) |
NK cells
| cells/mm3 | 143 (100, 214) | 174 (121, 230) | 268 (146, 388) | 75 (55, 103) | 41 (26, 70) |
Dendritic cells
| ||||||
Myeloid | /ml | 11940 (8751, 17805) | 11422 (6616, 13457) | 9122 (5662, 29989) | 7795 (4086, 10548) | 5420 (4107, 9683) |
Plasmacytoid | /ml | 7217 (4801, 11084) | 6152 (3599, 9836) | 2771 (1766, 8490) | 3073 (1351, 3618) | 2766 (2194, 4255) |
Gammadelta cells
| cells/mm3 | 44 (20, 74) | 27 (11, 54) | 396 (178, 579) | 19 (6, 92) | 16 (9, 26) |
Gammadelta 2
| % GD | 62.96 (40.85, 73.68) | 78.71 (75.00, 89.39) | 13.40 (8.77, 30.62) | 6.59 (4.09, 25.61) | 25.00 (12.41, 55.96) |
PHID complications according to clusters and T-cell activation
Cluster 1 | Cluster 2 | Cluster 3 | Cluster 4 | Cluster 5 | ||||||
---|---|---|---|---|---|---|---|---|---|---|
n = 33 | n = 18 | n = 4 | n = 8 | n = 16 | ||||||
n | % | n | % | n | % | n | % | n | % | |
Type of PHID
| ||||||||||
CVID | 23 | (70) | 8 | (44) | 3 | (75) | 8 | (100) | 13 | (82) |
IgG subclass deficiency | 10 | (30) | 10 | (56) | 0 | (0) | 0 | (0) | 1 | (6) |
Good’s syndrome | 0 | (0) | 0 | (0) | 1 | (25) | 0 | (0) | 2 | (12) |
PHID complications (at least one of the following)
|
5
|
(15)
|
6
|
(33)
|
3
|
(75)
|
7
|
(88)
|
10
|
(63)
|
Lymphoid hyperplasia | 2 | (6) | 5 | (28) | 1 | (25) | 3 | (38) | 5 | (31) |
Splenomegaly | 1 | (3) | 2 | (11) | 1 | (25) | 7 | (88) | 3 | (19) |
Autoimmune manifestations | 3 | (9) | 1 | (5) | 1 | (25) | 2 | (25) | 7 | (44) |
Granulomatous disease | 0 | (0) | 1 | (5) | 2 | (50) | 4 | (50) | 2 | (13) |
Villous atrophy | 0 | (0) | 2 | (11) | 0 | (0) | 0 | (0) | 3 | (19) |
Chronic inflammatory intestinal disease | 0 | (0) | 2 | (11) | 1 | (25) | 1 | (13) | 0 | (0) |
Unit | Odds ratio | 95% confidence interval | p-value | |
---|---|---|---|---|
B cells
| ||||
CD19+ | cells/mm3 | 0.65 | [0.36; 1.15] | 0.1373 |
CD27-IgD+ | % CD19+ | 1.49 | [0.89; 2.48] | 0.1310 |
CD27-IgD- | % CD19+ | 0.54 | [0.24; 1.23] | 0.1401 |
CD27 + IgD+ | % CD19+ | 0.62 | [0.34; 1.12] | 0.1124 |
CD27 + IgD- | % CD19+ | 0.93 | [0.29; 0.86] | 0.0119 |
T cells
| ||||
CD3+ | cells/mm3 | 0.68 | [0.40; 1.17] | 0,1646 |
CD4+ | cells/mm3 | 0.45 | [0.26; 0.78 ] | 0,0046 |
CD8+ | cells/mm3 | 1.04 | [0.66; 1.62 ] | 0.8818 |
CD4+/CD8+ ratio | 0.99 | [0.63; 1.56 ] | 0.9649 | |
CD8+ T cells
| ||||
CD45RA + CCR7+ | % CD8+ | 0.69 | [0.43; 1.13] | 0.1385 |
CD45RA-CCR7+ | % CD8+ | 0.75 | [0.46; 1.25] | 0.2710 |
CD45RA-CCR7- | % CD8+ | 1.38 | [0.86; 2.20] | 0.1800 |
CD45RA + CCR7- | % CD8+ | 0.77 | [0.48; 1.23] | 0.2666 |
CD8 + CD57+ | % CD8+ | 1.30 | [0.83; 2.06] | 0.2567 |
Activated T cells
| ||||
CD3 + HLA-DR+ | % CD3+ | 2.99 | [1.56; 5.73] | 0.0010 |
CD4 + HLA-DR+ | % CD4+ | 2.70 | [1.40; 5.23] | 0.0032 |
CD8 + HLA-DR+ | % CD8+ | 2.75 | [1.54; 4.90] | 0.0006 |
Regulatory T cells
| cells/mm3 | 0.41 | [0.22; 0.78] | 0.0065 |
Natural Killer cells
| cells/mm3 | 0.75 | [0.46; 1.21] | 0.2369 |
Dendritic cells
| ||||
Myeloid | /ml | 0.65 | [0.36; 1.17] | 0.1532 |
Plasmacytoid | /ml | 0.37 | [0.18; 0.73] | 0.0046 |
Gammadelta cells
| cells/mm3 | 1.30 | [0.79; 2.13] | 0.3028 |
Gammadelta 2 cells | % GD | 0.52 | [0.32; 0.85] | 0.0087 |
Classification of CVID patients according to EUROclass and CD8+ HLA-DR + marker
Model with EUROclass | Model with EUROclass and CD8+ HLA-DR+ | Total | |
---|---|---|---|
Predicted probabilities (%) | < 50% | ≥ 50% | |
Patients who experienced CVID complications
| |||
< 50% | 9 | 19 | |
≥ 50% | 1 | 5 | 6 |
Total | 10 | 15 | 25 |
Patients who didn’t experience CVID complications
| |||
< 50% | 22 | 5 | 27 |
≥ 50% | 2 | 1 | 3 |
Total | 24 | 6 | 30 |