Mass spectrometry-based proteomic analysis of proteins adsorbed by hexadecyl-immobilized cellulose bead column for the treatment of dialysis-related amyloidosis

Figures & data

Table 1. Baseline characteristics of ESKD patients undergoing HD with HICB column (HICB-HD) and regular HD.

	HD	HICB-HD	p Value
No. of patients	14	14
Case mix
Age, years	68.6 (62.4–77.1)	68.3 (62.6–71.0)	.476
Male sex	10	9	1
Body mass index, kg/m^2	21.5(18.9–22.9)	21.9 (19.1–23.7)	.701
Systolic blood pressure, mm Hg	135 (131–142)	140 (129–144)	.748
Cause of ESKD, DM/CGN/others	1/12/0/1	0/12/0/2	.531
Dialysis treatment
Duration of dialysis, years	28.3 (24.2–31.0)	32.9 (28.1–38.5)	.077
Single pool Kt/V	1.57 (1.43–1.70)	1.74 (1.54–1.85)	.129
Laboratory values
Blood hemoglobin, g/dL	10.5 (10.0–11.1)	10.7 (9.7–11.1)	.945
Albumin, g/dL	3.7 (3.5–3.8)	3.5 (3.4–3.7)	.594
Calcium, mg/dL	8.7 (8.3–9.1)	9.0 (8.8–9.6)	.051
Phosphate, mg/dL	5.4 (4.3–6.1)	4.6 (4.0–5.8)	.49
C-reactive protein, mg/dL	0.29 (0.13–0.49)	0.16 (0.09–0.26)	.113
Parathyroid hormone, ng/mL	127 (79–173)	101 (50–164)	.528
β2-Microglobulin, mg/L	27.4 (22.7–28.6)	23.4 (22.1–24.2)	.099
Histories of dialysis-related amyloidosis
Carpal tunnel syndrome	5 (36%)	14 (100%)	.128
Destructive spondyloarthropathy	2 (14%)	5 (36%)	.385
Joint arthropathy	2 (14%)	7 (50 %)	.103
Trigger finger	2 (14%)	3 (21%)	1

CGN: chronic glomerulonephritis; DM: diabetic mellitus; ESKD: end-stage kidney disease; HICB: hexadecyl-immobilized cellulose beads; HD: haemodialysis; NS: nephrosclerosis.

All values are expressed as the median (interquartile range) or number of patients (%). The Mann–Whitney and Chi-square tests were used to assess the differences between the HICB-HD and HD groups.

Table 2. Top 30 proteins with high SI_N extracted from HICBs after dialysis treatment of 14 DRA patients.

Protein symbol	Protein description	SIN
ALB	Albumin	296.6 ± 61.5
APOA4	Apolipoprotein A4	204.4 ± 46.7
B2M	Beta-2-microglobulin	193.9 ± 143.4
RBP4	Retinol binding protein 4	172.9 ± 55.4
LYZ	Lysozyme	156.5 ± 47.8
CFD	Complement factor D	109.4 ± 41.9
APOC3	Apolipoprotein C3	106.2 ± 58.9
IGLC2	Immunoglobulin lambda constant 2	89.9 ± 66
CST3	Cystatin C	88.8 ± 32.2
APOA1	Apolipoprotein A1	82.3 ± 26.9
TMSB4X	Thymosin beta 4 X-linked	75.5 ± 51.3
ANG	Angiogenin	70.2 ± 27
IGKC	Immunoglobulin kappa constant	66.3 ± 32.9
AMBP	Alpha-1-microglobulin/bikunin precursor	59.6 ± 20.4
RNASE1	Ribonuclease A family member 1	52.7 ± 31
TF	Transferrin	50.6 ± 16.7
APOA2	Apolipoprotein A2	48.6 ± 30.2
DEFA1	Defensin alpha 1	47 ± 27.7
IGFBP4	Insulin-like growth factor binding protein 4	43.1 ± 14.3
IGLC3	Immunoglobulin lambda constant 3	41.9 ± 62.5
APOH	Apolipoprotein H	38.9 ± 11
SERPINA1	Serpin family A member 1	34.2 ± 10.8
ORM1	Orosomucoid 1	33.9 ± 23.5
MGP	Matrix Gla protein	32.3 ± 25.9
TTR	Transthyretin	32.1 ± 17.5
RNASE4	Ribonuclease 4	31.7 ± 10.9
FGA	Fibrinogen alpha chain	31.1 ± 9.7
REG1A	Regenerating family member 1 alpha	30.8 ± 18.5
RARRES2	Retinoic acid receptor responder 2	30 ± 11.9
GC	Vitamin D-binding protein	27.9 ± 7.8

DRA: dialysis-related amyloidosis; HICB: hexadecyl-immobilized cellulose bead; SI_N: normalized spectral index.

All values are expressed as the mean ± standard deviation.

Table 3. Proteins adsorbed by HICBs and present in the amyloid lesions in carpal tunnel synovia of dialysis patients with DRA.

Protein symbol	Protein description	SIN in HICB	SIN in amyloid
APOA4	Apolipoprotein A4	204.4 ± 46.7	85.2 ± 81.5
B2M	Beta-2-microglobulin	193.9 ± 143.4	149.3 ± 125.2
LYZ	Lysozyme	156.5 ± 47.8	9.5 ± 8.9
CFD	Complement factor D	109.4 ± 41.9	4.5 ± 2.2
APOA1	Apolipoprotein A1	82.3 ± 27	16.6 ± 8.5
ANG	Angiogenin	70.3 ± 27	1.4 ± 1.8
AMBP	Alpha-1-microglobulin	59.6 ± 20.4	4.4 ± 3.6
MGP	Matrix Gla protein	32.3 ± 25.9	2.5 ± 4.3
FGA	Fibrinogen alpha chain	31.1 ± 9.7	55.5 ± 65.5
RPS27A	Ribosomal protein S27a	22.3 ± 6.5	9.4 ± 4
APOC1	Apolipoprotein C1	16.7 ± 7.7	1.8 ± 1.7
APOE	Apolipoprotein E	13.4 ± 5.6	22.6 ± 17.6
C9	Complement C9	9.6 ± 3.1	4.4 ± 8.1
CLU	Clusterin	5.3 ± 1.5	17.5 ± 14.8
FN1	Fibronectin 1	1.4 ± 0.4	13.3 ± 7.3
S100A6	S100 calcium binding protein A6	1.3 ± 1	47.7 ± 29.2
HIST1H1E	Histone cluster 1 H1 family member e	1.2 ± 0.5	7.4 ± 5.6
CLEC3B	C-type lectin domain family 3 member B	7 ± 3.2	3.9 ± 4.4
IGHG2	Immunoglobulin heavy constant gamma 2	5.9 ± 2.3	1.7 ± 1.3
HRG	Histidine rich glycoprotein	2.5 ± 1	1.5 ± 1.7
THBS4	Thrombospondin 4	1.7 ± 0.7	1.3 ± 1.3

DRA: dialysis-related amyloidosis; HICB: hexadecyl-immobilized cellulose bead; SI_N: normalized spectral index.

All values are expressed as the mean ± standard deviation.

Figure 3. In vitro Αβ_2M amyloid fibril formation with the amyloid-related proteins adsorbed by HICBs. Time courses of the aβ_2M amyloid fibril formation with lysozyme (A), angiogenin (B), complement factor D (C) and matrix Gla protein (D) were monitored using a thioflavin T (ThT) fluorescence assay. Maximum values of ThT fluorescence are shown in the reaction with lysozyme (E), angiogenin (F), complement factor D (G) and matrix Gla protein (H). Morphological analysis was performed using TEM in the reaction with lysozyme (I), angiogenin (J), complement factor D (K) and matrix Gla protein (L). The concentrations are shown at the top of the figures. Scale bars = 200 nm. HICB: hexadecyl-immobilized cellulose beads.

Data availability statement

Restrictions apply to the availability of the data analyzed in this study to preserve patient confidentiality. Data will be shared upon request to the corresponding author.