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Solute Carrier Transporters in Synovial Membrane and Hoffa’s Pad of Patients with Rheumatoid Arthritis

Damian Malinowski
Damian Malinowski
, 
Katarzyna Piotrowska
Katarzyna Piotrowska
, 
Marek Droździk
Marek Droździk
 y 
Andrzej Pawlik
Andrzej Pawlik
   | 27 jun 2024
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Article Category: Original
Publicado en línea: 27 jun 2024
Páginas: -
Recibido: 05 mar 2024
Aceptado: 13 may 2024
DOI: https://doi.org/10.2478/aite-2024-0014
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© 2024 Damian Malinowski et al., published by Sciendo
This work is licensed under the Creative Commons Attribution-NonCommercial-NoDerivatives 4.0 International License.

Fig 1.

Expression of SLC16A3, SLC19A1, SLC2A9, SLC22A18, SLC33A1, SLC7A5, SLC7A6, SLC02B1, and SLC04A1 membrane transporters in the SM. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier; SM, synovial membrane.
Expression of SLC16A3, SLC19A1, SLC2A9, SLC22A18, SLC33A1, SLC7A5, SLC7A6, SLC02B1, and SLC04A1 membrane transporters in the SM. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier; SM, synovial membrane.

Fig 2.

Expression of SLC15A2, SLC22A1, SLC22A3, SLC22A4, SLC22A5, SLC47A1, SLC51A, and SLC01C1 membrane transporters in the SM. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier; SM, synovial membrane.
Expression of SLC15A2, SLC22A1, SLC22A3, SLC22A4, SLC22A5, SLC47A1, SLC51A, and SLC01C1 membrane transporters in the SM. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier; SM, synovial membrane.

Fig 3.

Expression of SLC16A3, SLC19A1, SLC2A9, SLC22A18, SLC33A1, SLC7A5, SLC7A6, SLC02B1, and SLC04A1 membrane transporters in the Hoffa’s pad. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier.
Expression of SLC16A3, SLC19A1, SLC2A9, SLC22A18, SLC33A1, SLC7A5, SLC7A6, SLC02B1, and SLC04A1 membrane transporters in the Hoffa’s pad. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier.

Fig 4.

Expression of SLC15A2, SLC22A1, SLC22A3, SLC22A4, SLC22A5, SLC47A1, SLC51A, and SLC01C1 membrane transporters in the Hoffa’s pad. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, Rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier.
Expression of SLC15A2, SLC22A1, SLC22A3, SLC22A4, SLC22A5, SLC47A1, SLC51A, and SLC01C1 membrane transporters in the Hoffa’s pad. X: abbreviations of the examined SLC transporters superfamily. Y: RQ 2−ΔCT mean values (±SD); *p < 0.05. CT, cycle threshold; RA, Rheumatoid arthritis; RQ, relative quantification; SD, standard deviation; SLC, solute carrier.

Fig 5.

Transporter protein levels in SM and HFPs samples from RA and OA patients for PEPT2, SLC19A1, and MATE1 proteins. Pixel intensity acquired by ImageJ software. *Student’s t-test; #Mann–Whitney U-test. HFPs, Hoffa’s fat pads; OA, osteoarthritis; RA, rheumatoid arthritis; SLC, solute carrier; SM, synovial membrane.
Transporter protein levels in SM and HFPs samples from RA and OA patients for PEPT2, SLC19A1, and MATE1 proteins. Pixel intensity acquired by ImageJ software. *Student’s t-test; #Mann–Whitney U-test. HFPs, Hoffa’s fat pads; OA, osteoarthritis; RA, rheumatoid arthritis; SLC, solute carrier; SM, synovial membrane.

Fig 6.

Transporter protein expressions in SM samples from RA and OA patients. Transporters expression in RA patients panels (a, c, and e); transporters expressions in OA SM samples panels: (b, d, and f); green arrows—positive reaction of epithelial cells, red arrows—positive reaction of synoviocytes, red star—positive reaction in blood vessels; scale bar 50 μm; only representative images are presented. OA, osteoarthritis; RA, rheumatoid arthritis; SM, synovial membrane.
Transporter protein expressions in SM samples from RA and OA patients. Transporters expression in RA patients panels (a, c, and e); transporters expressions in OA SM samples panels: (b, d, and f); green arrows—positive reaction of epithelial cells, red arrows—positive reaction of synoviocytes, red star—positive reaction in blood vessels; scale bar 50 μm; only representative images are presented. OA, osteoarthritis; RA, rheumatoid arthritis; SM, synovial membrane.

Fig 7.

Transporter protein expressions in HFPs samples from RA and OA patients. Transporters expression in RA patients panels (a, c, and e); transporters expressions in OA HFP samples panels: (b, d, and f); scale bar 50 μm; only representative images are presented. HFPs, Hoffa’s fat pads; OA, osteoarthritis; RA, rheumatoid arthritis.
Transporter protein expressions in HFPs samples from RA and OA patients. Transporters expression in RA patients panels (a, c, and e); transporters expressions in OA HFP samples panels: (b, d, and f); scale bar 50 μm; only representative images are presented. HFPs, Hoffa’s fat pads; OA, osteoarthritis; RA, rheumatoid arthritis.

Fig 8.

SLC transporters: transport type of PEPT2 (cotransporter), RFC1 (exchanger) and MATE1 (exchanger) and their substrates. MPP, 1-methyl-4-phenylpyridinium; SLC, solute carrier; TEA, tetraethylammonium.
SLC transporters: transport type of PEPT2 (cotransporter), RFC1 (exchanger) and MATE1 (exchanger) and their substrates. MPP, 1-methyl-4-phenylpyridinium; SLC, solute carrier; TEA, tetraethylammonium.

IRS values for transportation proteins expression in SM samples in RA and OA patients

Protein RA OA
Median Average Min. Max. Median Average Min. Max
PEPT2 8 8.13 4 12 6 5.25 0 9
RFC1 8 7.93 4 12 6 5.63 1 12
MATE1 6 6.43 2 9 6 5.85 0 12

SLC transporters expression in SM and HFP

SM (mean ± SD) HFP (mean ± SD) p-value*
SLC0C1 ∆∆CT 0.437 ± 0.37 1.332 ± 0.90 0.088
SLC02B1 31.462 ± 63.66 41.233 ± 43.92 0.070
SLC04A1 14.388 ± 13.31 35.838 ± 21.45 0.036
SLC2A9 6.197 ± 1.22 10.295 ± 8.95 0.895
SLC7A5 13.45 ± 20.90 31.441 ± 28.92 0.239
SLC7A6 18.971 ± 15.31 121.553 ± 125.98 0.0005
SLC15A2 0.963 ± 1.05 2.6 ± 2.47 0.039
SLC16A3 154.528 ± 206.39 42.223 ± 30.48 0.041
SLC19A1 7.35 ± 9.30 22.44 ± 15.00 0.006
SLC22A1 1.289 ± 2.28 1.101 ± 0.89 0.307
SLC22A3 0.491 ± 0.69 2.386 ± 1.84 0.010
SLC22A4 2.081 ± 1.72 4.371 ± 2.93 0.064
SLC22A5 1.569 ± 1.49 6.656 ± 3.96 0.0008
SLC22A18 22.775 ± 16.89 8.694 ± 2.87 0.005
SLC33A1 9.382 ± 5.01 25.173 ± 9.62 0.0008
SLC47A1 2.632 ± 2.84 4.3612 ± 2.18 0.225
SLC51A 0.146 ± 0.17 0.397 ± 0.12 0.275

Correlation between the expression of SLC transporters in SM and clinical parameters

CRP WBC Age (years) Disease duration DAS28
SLC15A2 R −0.267 0.517 −0.385 0.017 −0.217
p-value 0.488 0.154 0.306 0.965 0.576
SLC22A18 R 0.188 −0.200 0.261 −0.118 0.176
p-value 0.603 0.580 0.466 0.746 0.627
SLC16A3 R 0.006 −0.091 −0.176 0.285 −0.018
p-value 0.987 0.803 0.626 0.425 0.960
SLC33A1 R 0.261 −0.382 0.122 0.415 0.188
p-value 0.467 0.276 0.738 0.233 0.603
SLC19A1 R −0.357 0.548 0.190 0.400 −0.333
p-value 0.385 0.160 0.651 0.326 0.420
SLC47A1 R −0.321 0.214 −0.500 0.436 −0.250
p-value 0.482 0.645 0.253 0.328 0.589
SLC2A9 R −0.783 0.100 −0.393 −0.339 −0.767
p-value 0.013 0.798 0.295 0.372 0.016
SLC51A R 1.000 0.500 −0.500 −0.866 1.000
p-value − 0.667 0.667 0.333 −
SLC22A1 R −0.238 0.595 −0.132 0.146 −0.238
p-value 0.570 0.120 0.756 0.729 0.570
SLC22A3 R −0.464 0.429 0.214 0.436 −0.429
p-value 0.294 0.337 0.645 0.328 0.337
SLC7A5 R −0.050 0.333 −0.519 0.373 −0.033
p-value 0.898 0.381 0.152 0.323 0.932
SLC22A4 R 0.036 0.036 −0.286 0.764 0.036
p-value 0.939 0.939 0.535 0.046 0.939
SLC7A6 R 0.200 0.455 0.195 0.415 0.212
p-value 0.580 0.187 0.590 0.233 0.556
SLC22A5 R −0.350 −0.033 0.259 0.602 −0.333
p-value 0.356 0.932 0.500 0.086 0.381
SLC01C1 R −0.700 −0.200 0.600 0.500 −0.700
p-value 0.188 0.747 0.285 0.391 0.188
SLC02B1 R −0.491 0.503 0.255 0.223 −0.503
p-value 0.150 0.138 0.476 0.536 0.138
SLC04A1 R 0.143 0.095 −0.048 0.424 0.024
p-value 0.736 0.823 0.911 0.295 0.955

Correlation between the expression of SLC transporters in Hoffa’s pad and clinical parameters

CRP WBC Age (years) Disease duration DAS28
SLC15A2 r 0.256 0.250 −0.357 0.382 0.255
p-value 0.579 0.589 0.432 0.398 0.582
SLC22A18 r −0.106 −0.261 0.292 0.344 0.085
p-value 0.770 0.467 0.413 0.331 0.815
SLC16A3 r 0.400 0.552 −0.195 0.368 0.530
p-value 0.252 0.098 0.590 0.295 0.115
SLC33A1 r −0.052 0.017 0.393 0.703 0.286
p-value 0.894 0.966 0.295 0.034 0.456
SLC19A1 r −0.355 −0.024 0.357 0.194 0.217
p-value 0.388 0.955 0.385 0.645 0.606
SLC47A1 r 0.473 0.821 −0.643 0.273 0.691
p-value 0.284 0.023 0.119 0.554 0.086
SLC2A9 r −0.279 −0.283 0.117 −0.177 0.185
p-value 0.468 0.460 0.765 0.648 0.634
SLC51A r 0.500 1.000 −0.500 0.866 1.000
p-value 0.667 − 0.667 0.333 −
SLC22A1 r −0.866 −0.500 0.500 −0.866 −0.866
p-value 0.333 0.667 0.667 0.333 0.333
SLC22A3 r −0.540 −0.667 0.733 0.203 −0.445
p-value 0.134 0.050 0.025 0.601 0.230
SLC7A5 r 0.516 0.486 0.200 0.088 0.530
p-value 0.295 0.329 0.704 0.868 0.280
SLC22A4 r −0.185 −0.357 0.821 0.327 −0.182
p-value 0.691 0.432 0.023 0.474 0.696
SLC7A6 r −0.256 −0.188 0.213 0.491 0.110
p-value 0.475 0.603 0.555 0.150 0.763
SLC22A5 r −0.669 −0.224 0.018 −0.153 −0.189
p-value 0.034 0.533 0.960 0.672 0.601
SLC01C1 r −0.906 −0.250 0.393 0.255 −0.436
p-value 0.005 0.589 0.383 0.582 0.328
SLC02B1 r −0.344 −0.030 −0.188 0.436 −0.268
p-value 0.331 0.934 0.602 0.208 0.454
SLC04A1 r 0.732 0.143 −0.119 −0.590 0.395
p-value 0.039 0.736 0.779 0.123 0.333

Antibodies used in the IHC procedure and the IRS evaluation

Antibody Host Manufacturer Rate of dilution Time of incubation/temperature
Anti-PEPT2 (SLC15A2) Rabbit Abcam, Cambridge, United Kingdom 1:75 1 h/RT
Anti-RFC1 (SLC19A1) Mouse Santa Cruz Biotechnology Inc., Dallas, TX, USA 1:100 1 h/RT
Anti-MATE1 (SLC47A1) Rabbit Sigma Prestige, Merck, Darmstadt, Germany 1:200 1 h/RT

TaqMan gene expression assays

SLC transporters Housekeeping genes
Gene symbol TaqMan assay ID Gene symbol TaqMan assay ID
SLC01A2 Hs00366488_m1 GUSB Hs99999908_m1
SLC01B1 Hs00272374_m1 RPS9 Hs02339424_g1
SLC01B3 Hs00351987_m1
SLC01C1 Hs00213714_m1
SLC02B1 Hs01030353_m1
SLC04A1 Hs00983988_m1
SLC04C1 Hs00698884_m1
SLC10A1 Hs00161820_m1
SLC10A2 Hs01001557_m1
SLC15A1 Hs00192639_m1
SLC15A2 Hs01113665_m1
SLC16A3 Hs00358829_m1
SLC19A1 Hs00953344_m1
SLC22A1 Hs00427552_m1
SLC22A11 Hs00945829_m1
SLC22A12 Hs00375985_m1
SLC22A18 Hs00180039_m1
SLC22A2 Hs01010726_m1
SLC22A3 Hs00222691_m1
SLC22A4 Hs01548718_m1
SLC22A5 Hs00929869_m1
SLC22A6 Hs00537914_m1
SLC22A7 Hs00198527_m1
SLC22A8 Hs00188599_m1
SLC22A9 Hs00375768_m1
SLC28A1 Hs00984403_m1
SLC28A2 Hs00188407_m1
SLC2A9 Hs00417125_m1
SLC33A1 Hs00270469_m1
SLC47A1 Hs00217320_m1
SLC47A2 Hs00945650_m1
SLC51A Hs00380895_m1
SLC51B Hs01057182_m1
SLC7A5 Hs01001183_m1
SLC7A6 Hs00187727_m1

IRS values for transportation proteins expression in HFP in RA and OA patients

Protein RA OA
Median Average Min. Max. Median Average Min. Max.
PEPT2 8 9.47 4 12 8 7.17 2 12
RFC1 12 9.06 1 12 4 4.93 0 12
MATE1 8 7.47 2 12 8 6.5 1 12
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