Immunophenotypic characteristics of multipotent mesenchymal stromal cells that affect the efficacy of their use in the prevention of acute graft vs host disease

Table 1 Characteristics of donors, CFU-F and multipotent mesenchymal stromal cells

Donor MSCs	Gender male/female	Age (median), yr	Body mass index	CFU-F per 106 BM cells	Total MSC production for 4 passages, × 106	Time to P0, d
No GVHD	24/17	33.19 ± 1.70 (32)	24.88 ± 0.82 (23)	17.27 ± 3.39	13.93 ± 2.14	14.44 ± 0.42
Acute GVHD	5/6	41.27 ± 4.15 (45)	26.36 ± 1.58 (27)	11.46 ± 4.50	7.71 ± 2.40	14.73 ± 0.87

MSCs: Multipotent mesenchymal stromal cells; GVHD: Graft vs host disease.

Table 2 Changes in the mean fluorescence intensity of surface multipotent mesenchymal stromal cells untreated and treated with interleukin 1β, interferon gamma and tumour necrosis factor alpha antigens

MFI
	MSCs		MSCs treated with IL1β		MSCs treated with IFN		MSCs treated with TNF
Antigen	Effective	Ineffective	Effective	Ineffective	Effective	Ineffective	Effective	Ineffective
HLA-ABC	14561 ± 7741	8767 ± 1434	14740 ± 10211	8850 ± 994	15200 ± 9601	10208 ± 1189	21174 ± 1615	15706 ± 2938
HLA-DR	752 ± 1501	188 ± 42	610 ± 1541	149 ± 25	1077 ± 1531	423 ± 163	659 ± 1371	169 ± 38
CD54	4881 ± 371	4447 ± 1203	13773 ± 32181	4234 ± 1612	7889 ± 8161	4378 ± 1566	61708 ± 5998	65461 ± 11324
CD73	3047 ± 253	2354 ± 343	3396 ± 305	2226 ± 543	3414 ± 383	2678 ± 655	3395 ± 305	3370 ± 605
CD90	57679 ± 2626	49241 ± 7022	60117 ± 3990	45739 ± 9525	56500 ± 30721	40335 ± 4502	55799 ± 3471	45198 ± 7142
CD105	35682 ± 26231	21471 ± 3247	34402 ± 2490	29665 ± 6032	41171 ± 3118	30525 ± 5609	30431 ± 2471	24260 ± 3466
CD146	4345 ± 5341	1807 ± 482	3546 ± 497	3124 ± 688	3762 ± 479	2378 ± 707	3137 ± 387	3463 ± 539
CD274	293 ± 37		261 ± 38	227 ± 204	400 ± 104	304 ± 219	341 ± 54	322 ± 81
%CD54	56 ± 3	50 ± 9	57 ± 4	42 ± 8	66 ± 3	55 ± 8	89 ± 2	93 ± 4

¹Significant differences between effective and ineffective samples. MFI: Mean fluorescence intensity; MSC: Multipotent mesenchymal stromal cell; IL1β: Interleukin 1β; IFN: Interferon gamma; TNF: Tumour necrosis factor alpha.

Table 3 Changes in the mean fluorescence intensity of surface multipotent mesenchymal stromal cell antigens during co-cultivation with peripheral blood mononuclear cells expressed as the fold change relative to multipotent mesenchymal stromal cells cultured without peripheral blood mononuclear cells

Time of co-cultivation	1 d		4 d
Group of MSC samples	Effective	Ineffective	Effective	Ineffective
HLA-ABC	1.90 ± 0.11	1.53 ± 0.30	1.90 ± 0.15	1.67 ± 0.33
CD90	0.95 ± 0.04	0.79 ± 0.061	0.55 ± 0.04	0.49 ± 0.10
HLA-DR	134.37 ± 63.90	654.74 ± 363.49	682.42 ± 326.38	151.88 ± 96.44
CD105	1.08 ± 0.06	0.98 ± 0.14	0.64 ± 0.05	0.42 ± 0.07
CD146	0.82 ± 0.07	0.86 ± 0.30	22.16 ± 16.28	0.81 ± 0.21
CD54	60.59 ± 12.29	44.76 ± 8.84	33.08 ± 5.30	23.19 ± 7.64
CD274	434.64 ± 93.56	1.27 ± 0.281	573.44 ± 172.78	319.25 ± 233.40
CD73	1.05 ± 0.08	0.56 ± 0.131	150.36 ± 103.94	1.60 ± 0.54

¹Significant differences between effective and ineffective samples. MSC: Multipotent mesenchymal stromal cell.

Table 4 Discriminant function analysis summary

MSCs markers after 1 d co-cultivation with non-activated and activated lymphocytes (PHA)	Wilks'Lambda	Partial	F-remove	P-level	Coefficients of discriminant function
					No GVHD1	GVHD2
HLA-DR	0.572005	0.625895	11.95423	0.002488	-0.00358	0.0213
CD146	0.439602	0.814407	4.55774	0.045330	-0.01545	0.0095
CD73 PHA	0.517153	0.692280	8.89005	0.007375	4.30313	-0.6335
CD105	0.499794	0.716325	7.92028	0.010713	9.05802	-1.9000
CD105 PHA	0.419957	0.852503	3.46032	0.077629	2.65834	11.3187
CD54	0.404184	0.885773	2.57915	0.123954	-0.01228	0.0118

¹Constant for no graft vs host disease (GVHD) group – (-8.86214).

²Constant for GVHD group - (-10.0071). MSCs: Multipotent mesenchymal stromal cells; GVHD: Graft vs host disease.

Table 5 Classification of multipotent mesenchymal stromal cell samples

Sample number	GVHD development after prophylaxis	Posterior probabilities of sample classification		Squared mahalanobis distances from group centroids
		No GVHD	GVHD	No GVHD	GVHD
1	No GVHD	0.999996	0.000004	4.81908	26.37491
2	No GVHD	0.999831	0.000169	4.69845	18.56620
3	No GVHD	0.999937	0.000063	8.69194	24.53850
4	No GVHD	0.999973	0.000027	5.37220	22.94517
5	No GVHD	0.999999	0.000001	4.25232	27.68426
6	No GVHD	0.999513	0.000487	1.87460	13.62827
6	No GVHD	0.957820	0.042180	4.51571	7.26276
8	No GVHD	0.999812	0.000188	0.52715	14.18290
9	No GVHD	0.999849	0.000151	1.49937	15.60338
10	No GVHD	0.999975	0.000025	1.27143	18.96579
11	No GVHD	0.999962	0.000038	1.37772	18.21698
12	No GVHD	0.984365	0.015635	6.84328	11.62984
13	No GVHD	0.999843	0.000157	0.26067	14.27728
14	No GVHD	0.999493	0.000507	1.71004	13.38532
15	No GVHD	0.970256	0.029744	4.22516	7.69663
16	No GVHD	0.999991	0.000009	7.74281	27.47988
17	No o GVHD	0.999810	0.000190	6.84972	20.48673
18	No GVHD	0.999978	0.000022	2.57251	20.55185
191	GVHD	0.986823	0.013177	2.37048	7.50407
20	No GVHD	0.999810	0.000190	23.00573	36.64553
21	No GVHD	0.997370	0.002630	1.20348	9.58174
22	GVHD	0.000015	0.999985	44.40075	18.74916
23	GVHD	0.000033	0.999967	42.58649	18.44380
24	No GVHD	0.990716	0.009284	2.92989	8.77180
25	No GVHD	0.998626	0.001374	0.63394	10.31299
26	No GVHD	0.999170	0.000830	0.42912	11.11676
27	GVHD	0.096789	0.903211	15.96187	7.99662

¹Incorrect classifications. GVHD: Graft vs host disease.