Association Between Salivary Cytokines, Chemokines and Growth Factors and Salivary Gland Function in Children with Chronic Kidney Disease

Figures & data

Figure 1 Measurement of salivary inflammatory and anti-inflammatory profile by multiplex ELISA. Created with BioRender.com.

Table 1 Clinical Data of Children with Chronic Kidney Disease (CKD) Compared to Healthy Children (C)

	C (n = 30)	CKD (n = 30)	P-value
Sex (male/female)	16/14	16/14	ND
Age	13 (9–16)	13 (9–16)	>0.9999
Reasons of CKD
Glomerulopathies n (%)	0 (0)	10 (33.3)	ND
Urologic defects n (%)	0 (0)	9 (30)	ND
Nephropathies n (%)	0 (0)	5 (16.7)	ND
Renal dysplasia n (%)	0 (0)	5 (16.7)	ND
Undetermined etiology n (%)	0 (0)	1 (3.3)	ND
Kidney function
Serum Cr (mg/dL)	0.397 (0.3595–0.4642)	1.66 (0.9575–3.753)	<0.0001
Serum Urea (mg/dL)	17.38 (15.77–19.96)	58 (29.75–113)	<0.0001
eGFR (mL/min/1.73 m^2)	136.1 (131.2–141.5)	38.74 (16.2–89.6)	<0.0001
Albuminuria (mg/24 h)	ND	121.2 (10.38–335.5)	ND
Proteinuria (mg/24 h)	ND	150 (95.25–1029)	ND
Biochemical tests
Hemoglobin (g/dL)	ND	11.7 (10.6–14.1)	ND
Hematocrit (%)	ND	36.35 (31.38–39.33)	ND
Iron (µg/dL)	ND	82 (55.5–90)	ND
Drugs
ACEI n (%)	0 (0)	20 (66.7)	ND
β-blockers n (%)	0 (0)	7 (23.3)	ND
CCB n (%)	0 (0)	8 (26.7)	ND
Loop diuretics n (%)	0 (0)	21 (70)	ND
Iron n (%)	0 (0)	18 (60)	ND
Sialochemistry
SFR (mL/min)	0.475 (0.4–0.6175)	0.225 (0.1475–0.35)	<0.0001
TP (μg/mL)	1814 (1550–2054)	1280 (1041–1590)	<0.0001
SA (μg/mg protein)	0.225 (0.1775–0.3)	0.09 (0.07–0.1125)	<0.0001
Hyposalivation n (%)	0 (0)	14 (46.67)	ND
Dental examination
DMFT	10 (2–12)	10 (1–12)	>0.9999
dmft	3 (0–4)	3 (0–4)	>0.9999
PBI	0.2 (0–0.4)	0.2 (0–0.3)	>0.9999
GI	0.1 (0–0.2)	0.1 (0–0.2)	>0.9999
API (%)	20	23	>0.9999

Notes: Data are presented as the median (minimum – maximum). Bold indicates statistically significant results.

Abbreviations: ACEI, angiotensin converting enzyme inhibitors; API, Approximal Plaque Index; CCB, calcium channel blockers; dmft, decayed, missing, filled teeth index for deciduous teeth; DMFT, decayed, missing, filled teeth index for permanent teeth; eGFR, estimated glomerular filtration rate using Schwartz formula; Cr, creatinine; GI, gingival index; PBI, papilla bleeding index; SA, salivary amylase; SFR, salivary flow rate; TP, total protein.

Figure 2 Concentration of salivary pro-inflammatory and anti-inflammatory cytokines as well as Th1, Th2 and Th17 in the saliva of children with chronic kidney disease (CKD) compared to healthy children (C).

Figure 3 Concentration of salivary chemokines and growth factors in the saliva of children with chronic kidney disease (CKD) compared to healthy children (C).

Table 2 Multivariate Linear Regression Analysis Between the Secretory Function of the Salivary Glands and Salivary Inflammatory and Anti-Inflammatory Profile in Children with CKD and the Control

	β1 (SFR)			β2 (TP)			β3 (SA)
	Estimate	95% Confidence Interval	P value	Estimate	95% Confidence Interval	P value	Estimate	95% Confidence Interval	P value
IL-1β	−187.4	−395.5 to 20.78	0.0765	0.002668	−0.07026 to 0.07560	0.9415	−30.22	−445.6 to 385.2	0.884
TNF-α	−202.4	−363.4 to −41.46	0.0147	−0.04764	−0.1018 to 0.006485	0.0833	−34.86	−324.7 to 255.0	0.8104
IL-7	−0.4545	−11.61 to 10.70	0.9353	0.002207	−0.001570 to 0.005985	0.2467	12.25	−8.109 to 32.61	0.2332
IL-10	−40.53	−72.47 to −8.594	0.0139	0.0008253	−0.009683 to 0.01133	0.8753	−16.59	−81.73 to 48.55	0.6113
IL-1RA	−2139	−3771 to −507.0	0.0111	−0.375	−0.9275 to 0.1775	0.1794	101.6	−2876 to 3079	0.9457
IL-13	33.38	−1.523 to 68.28	0.0605	−0.001509	−0.01320 to 0.01019	0.7966	−62.22	−126.5 to 2.048	0.0575
INF-γ	−204.3	−340.5 to −68.01	0.004	−0.002708	−0.05011 to 0.04470	0.9092	−90.16	−333.1 to 152.8	0.4599
IL-2	−24.33	−52.92 to 4.255	0.0934	0.0005636	−0.009597 to 0.01072	0.9116	42.09	−9.277 to 93.46	0.1059
IL-12	−15.29	−30.90 to 0.3228	0.0548	−0.001702	−0.007039 to 0.003634	0.5252	52.12	22.10 to 82.14	0.001
IL-15	−40.46	−71.95 to −8.961	0.0128	−0.003247	−0.01391 to 0.007416	0.5443	−40.65	−98.12 to 16.81	0.1619
IL-4	−41.32	−72.37 to −10.26	0.0101	−0.002877	−72.37 to −10.27	0.5689	−22.69	−72.37 to −10.28	0.5689
IL-5	−48	−85.56 to −10.45	0.0133	−0.003634	−0.01594 to 0.008670	0.5558	−62.77	−136.0 to 10.45	0.0913
IL-6	−1835	−3077 to −593.9	0.0045	−0.1437	−0.5640 to 0.2766	0.4962	−597.8	−2863 to 1667	0.5991
IL-9	−192	−295.8 to −88.22	0.0005	−0.003858	−0.003858	0.8263	−189.5	−381.4 to 2.362	0.0528
IL-17	−43.39	−99.50 to 12.72	0.1269	0.002034	−0.01654 to 0.02061	0.827	−13.11	−117.6 to 91.43	0.8024
IP-10/CXCL10	−995.2	−2543 to 552.3	0.2013	−0.3874	−0.9356 to 0.1608	0.1611	1212	−1476 to 3900	0.3678
MCP-1/CCL-2	−2192	−4536 to 152.9	0.0663	0.05866	−0.7662 to 0.8835	0.887	−1893	−6111 to 2324	0.3714
MIP-1α/CCL3	−8914	−15,135 to −2692	0.0058	−0.4349	−2.611 to 1.741	0.6901	−2573	−13,717 to 8572	0.6901
MIP-1β/CCL4	−2672	−6862 to 1518	0.2063	−0.7304	−2.076 to 0.6155	0.2811	5787	−1816 to 13,390	0.1327
EOTAXIN/CCL11	−465.6	−826.2 to −104.9	0.0123	0.006195	−826.2 to −104.9	0.9194	−425.7	−1084 to 232.2	0.2002
RANTES/CCL5	−2953	−5982 to 76.61	0.0559	0.004459	−1.021 to 1.030	0.9931	−2837	−8364 to 2690	0.3082
IL-8/CXCL8	−2341	−5093 to 411.1	0.0939	−0.3955	−1.344 to 0.5530	0.407	4007	−918.1 to 8933	0.1087
G-CSF	−650.8	−1319 to 17.66	0.0561	−0.01748	−0.2259 to 0.1910	0.8669	1219	1.654 to 2436	0.0497
GM-CSF	−39.61	−74.71 to −4.510	0.0278	−0.004322	−0.01636 to 0.007717	0.4739	−6.46	−69.63 to 56.71	0.838
VEGF	63.72	−108.6 to 236.0	0.4609	−0.003759	−0.06296 to 0.05544	0.899	−0.06296 to 0.05544	−99.88 to 524.2	0.1781
FGF	−31.4	−57.07 to −5.732	0.0174	−0.002081	−0.01078 to 0.006619	0.6335	−6.97	−55.06 to 41.12	0.7725
PDGF	−74.97	−133.9 to −16.01	0.0138	−0.002557	−0.02245 to 0.01733	0.7973	31.58	−72.98 to 136.1	0.5468

Note: Bold indicates statistically significant results.

Abbreviations: IL-1β, interleukin 1β; TNF-α, tumor necrosis factor α; IL-7, interleukin 7; IL-10, interleukin 10; IL-1RA, interleukin 1RA; IL-13, interleukin 13; INF-γ, interferon γ; IL-12, interleukin 12; IL-2, interleukin 2; IL-15, interleukin 15; IL-4, interleukin 4; IL-5, interleukin 5; IL-6, interleukin 6; IL-9, interleukin 9; IL-17, interleukin 17; IP-10/CXCL10, chemokine (C-X-C motif) ligand 10/interferon gamma-induced protein 10; MCP-1/CCL2, monocyte chemoattractant protein-1; MIP-1α/CCL3, chemokine ligands 3/macrophage inflammatory protein 1α; MIP-1β/ CCL4, chemokine ligands 4/macrophage inflammatory protein 1β; CCL11/eotaxin, chemokine ligand 11/eotaxin; CCL5/RANTES, chemokine ligand 5/regulated on activation, normal T cell expressed and secreted; IL-8/CXCL8, interleukin 8; G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; VEGF, vascular endothelial growth factor; FGF basic, fibroblast growth factor; PDFG-BB, platelet-derived growth factor.

Table 3 Multivariate Linear Regression Analysis Between Renal Function and Salivary Inflammatory and Anti-Inflammatory Profile in Children with CKD and the Control

	β1 (Serum Cr)			β2 (Serum Urea)			β1 (eGFR)
	Estimate	95% Confidence Interval	P value	Estimate	95% Confidence Interval	P value	Estimate	95% Confidence Interval	P value
IL-1β	44,916	−14.05 to 56.29	0.2324	−0.1239	−1.640 to 1.392	0.8699	−0.1792	−1.081 to 0.7222	0.6905
TNF-α	52.04	24.16 to 79.93	0.0004	−1.935	−3.145 to −0.7244	0.0023	−0.8588	−1.562 to −0.1558	0.0176
IL-7	0.8119	−1.222 to 2.846	0.4273	0.0247	−0.06165 to 0.1110	0.5689	0.08248	0.03112 to 0.1338	0.0022
IL-10	2.909	−2.423 to 8.240	0.2785	0.1139	−0.1124 to 0.3403	0.317	0.03804	−0.09715 to 0.1732	0.5746
IL-1RA	96.21	−227.7 to 420.1	0.5542	1.584	−12.16 to 15.33	0.8183	−3.896	−12.07 to 4.282	0.344
IL-13	−4.296	−10.75 to 2.162	0.1876	−0.09585	−0.3711 to 0.1794	0.4875	−0.2638	−0.4483 to -0.07933	0.006
INF-γ	22.32	−2.810 to 47.44	0.0805	−0.4778	−1.544 to 0.5884	0.3727	−0.48	−1.118 to 0.1581	0.1372
IL-2	3.643	−1.253 to 8.538	0.141	0.01742	−0.1902 to 0.2251	0.141	0.103	−0.02489 to 0.2309	0.1118
IL-12	0.7658	−2.362 to 3.894	0.6255	0.07919	−0.05360 to 0.2120	0.2371	0.1001	0.02093 to 0.1792	0.0142
IL-15	1.884	−4.024 to 7.792	0.5256	0.1551	−0.09568 to 0.4059	0.2205	−0.01344	−0.1626 to 0.1357	0.8574
IL-4	7.024	1.428 to 12.62	0.0149	−0.1557	−0.3934 to 0.08203	0.1947	−0.06229	−0.2039 to 0.07933	0.3818
IL-5	0.6738	−5.746 to 7.094	0.834	0.2168	−0.05602 to 0.4896	0.1168	−0.07113	−0.2341 to 0.09181	0.3849
IL-6	354.1	139.3 to 568.8	0.0017	−10.69	−19.81 to −1.574	0.0224	−5.607	−11.03 to −0.1835	0.043
IL-9	20.62	−0.9466 to 42.18	0.0606	−0.8282	−1.743 to 0.08672	0.0751	−0.735	−1.281 to −0.1892	0.0092
IL-17	7.899	−2.035 to 17.83	0.1167	0.005244	−0.4165 to 0.4270	0.9802	0.08929	−0.1638 to 0.3423	0.4823
IP-10/CXCL10	107.5	−156.2 to 371.1	0.4151	−1.574	−12.75 to 9.605	0.7775	−1.396	−8.427 to 5.635	0.6905
MCP-1/CCL-2	1056	735.2 to 1377	<0.0001	−28.8	−43.07 to −14.52	0.0002	−0.1557	−9.174 to 8.863	0.9725
MIP-1α/CCL3	2042	1040 to 3044	0.0001	−59.6	−102.1 to −17.07	0.0069	−22.5	−47.93 to 2.940	0.0818
MIP-1β/CCL4	558.3	−213.2 to 1330	0.1524	−13.04	−45.78 to 19.71	0.4279	5.962	−13.62 to 25.55	0.5438
EOTAXIN/CCL11	47.22	−22.94 to 117.4	0.183	−0.5897	−3.568 to 2.389	0.6931	−0.9001	−2.672 to 0.8715	0.3132
RANTES/CCL5	708.3	123.3 to 1293	0.0185	−24.94	−49.77 to −0.1067	0.0491	−6.988	−21.76 to 7.783	0.3474
IL-8/CXCL8	390.6	−131.0 to 912.3	0.1391	−10.76	−32.91 to 11.38	0.3342	−2.281	−15.46 to 10.90	0.7301
G-CSF	186.2	80.76 to 291.7	0.0009	−5.799	−10.28 to −1.324	0.0122	0.8821	−1.821 to 3.585	0.5151
GM-CSF	4.503	−1.198 to 10.20	0.1188	0.06043	−0.1814 to 0.3023	0.6177	0.02139	−0.1261 to 0.1688	0.7718
VEGF	−7.344	−38.18 to 23.49	0.6343	0.862	−0.4460 to 2.170	0.1915	0.7562	−0.04027 to 1.553	0.0623
FGF	5.603	1.065 to 10.14	0.0165	−0.0919	−0.2845 to 0.1007	0.343	−0.0314	−0.1464 to 0.08365	0.5865
PDGF	34151	−2.252 to 18.11	0.1241	0.03751	−0.3949 to 0.4699	0.8624	0.07878	−0.1829 to 0.3404	0.5481

Abbreviations: IL-1β, interleukin 1β; TNF-α, tumor necrosis factor α; IL-7, interleukin 7; IL-10, interleukin 10; IL-1RA, interleukin 1RA; IL-13, interleukin 13; INF-γ, interferon γ; IL-12, interleukin 12; IL-2, interleukin 2; IL-15, interleukin 15; IL-4, interleukin 4; IL-5, interleukin 5; IL-6, interleukin 6; IL-9, interleukin 9; IL-17, interleukin 17; IP-10/CXCL10, chemokine (C-X-C motif) ligand 10/interferon gamma-induced protein 10; MCP-1/CCL2, monocyte chemoattractant protein-1; MIP-1α/CCL3, chemokine ligands 3/macrophage inflammatory protein 1α; MIP-1β/ CCL4, chemokine ligands 4/macrophage inflammatory protein 1β; CCL11/eotaxin, chemokine ligand 11/eotaxin; CCL5/RANTES, chemokine ligand 5/regulated on activation, normal T cell expressed and secreted; IL-8/CXCL8, interleukin 8; G-CSF, granulocyte colony-stimulating factor; GM-CSF, granulocyte-macrophage colony-stimulating factor; VEGF, vascular endothelial growth factor; FGF basic, fibroblast growth factor; PDFG-BB, platelet-derived growth factor. Bold indicates statistically significant results.

Figure 4 Analysis of the diagnostic utility of salivary MIP-1α/CCL3 in differentiating children with CKD and normal saliva secretion (NS) from patients with hyposalivation (HS).

Figure 5 Graphical conclusions from the study. Created with BioRender.com.