Change and significance of NKG2C and granzyme B labeled NK cell subsets in peripheral blood of children with purulent meningitis

doi:10.3969/j.issn.1673-8640.2023.05.011

Abstract

Abstract:

Objective To investigate the change characteristics and clinical significance of natural killer cell receptor 2C（NKG2C） and granzyme B（GZMB） labeled natural killer（NK） cell subsets in peripheral blood of children with purulent meningitis（PM）. Methods Totally，52 children with PM and 52 control children with non-central infectious diseases were enrolled. The children with PM were classified into good prognosis group（44 cases） and poor prognosis group（8 cases） according to the Glasgow Outcome Scale on discharge. NK cell subsets [NKG2C^- gamma-interferon gamma（INF-γ）⁺，NKG2C⁺INF-γ⁺，NKG2C⁺INF-γ^-，NKG2C^-GZMB⁺，NKG2C⁺GZMB⁺，NKG2C⁺GZMB^-] were determined by flow cytometry. The results of cerebrospinal fluid white blood cell（WBC） count，total protein（TP），glucose（Glu） and chlorine（Cl^-），peripheral blood WBC count，the percentage of neutrophil（NEUT%），serum procalcitonin（PCT） and high-sensitivity C-reactive protein（hs-CRP） were collected as well. Spearman rank correlation test was used to analyze the correlations. Receiver operating characteristic（ROC）curves were established to evaluate the values of the indicators in the diagnosis and prognosis of PM. Results The levels of NKG2C⁺，NKG2C^-GZMB⁺，NKG2C⁺GZMB⁺ and NKG2C⁺GZMB^- in PM group were higher than those in control group（P<0.05），and total NK cell percentage was lower than that in control group（P<0.05）. There was no NKG2C^-INF-γ⁺ and NKG2C⁺INF-γ⁺ NK cell in the 2 groups. The percentage of NK2G^-GZMB⁺ NK cells was positively correlated with cerebrospinal fluid WBC count，TP and serum PCT（r values were 0.286，0.558 and 0.740，P<0.05），and it was negatively correlated with cerebrospinal fluid Glu（r=-0.389，P<0.05）. The percentage of NKG2C⁺GZMB⁺ NK cells was positively correlated with cerebrospinal fluid WBC count，TP，peripheral blood WBC count and serum PCT（r=0.509，0.539，0.549 and 0.577，P<0.05），and it was negatively correlated with cerebrospinal fluid Glu（r=-0.391，P<0.05）. NKG2C⁺ GZMB^- NK cell percentage was positively correlated to cerebrospinal Glu level（r=0.624，P<0.05），and it was negatively correlated to cerebrospinal TP（r=-0.328，P<0.05）. The percentage and absolute value of total NK cells in PM group after treatment were higher than those before treatment（P<0.05）. Compared with before treatment，the percentage of NKG2C^-GZMB⁺ and NKG2C⁺GZMB⁺ NK cells in good prognosis group were decreased after treatment（P<0.05），while the percentage of NKG2C⁺GZMB^- NK cells was increased（P<0.05）. The percentage of NKG2C⁺GZMB⁺NK cells in poor prognosis group after treatment was increased（P<0.05）. ROC curve analysis results showed that NKG2C^-GZMB⁺ NK cell percentage had the highest diagnostic efficiency for PM，with the area under curve（AUC） of 0.980，the optimal cut-off value of 8.93%，the sensitivity of 92.31%，and the specificity of 98.08%. After treatment，NKG2C⁺GZMB^- NK cells had the highest efficiency in predicting poor prognosis in children with PM，with the AUC of 0.989，the optimal cut-off value of 5.51%，the sensitivity of 100.00%，and the specificity of 97.73%. Conclusions NKG2C^-GZMB⁺ NK cell subsets have a good value for the early diagnosis in children with PM. NKG2C⁺GZMB^- NK cells have a certain clinical value for prognostic prediction of PM.

Key words: Natural killer cell receptor 2C, Granzyme B, Natural killer cell, Purulent meningitis

CLC Number:

R446.1

YANG Ze, XIE Lili, TIAN Jing, XU Tongtong, ZHOU Guoqing, LIU Baojun, WU Huihui. Change and significance of NKG2C and granzyme B labeled NK cell subsets in peripheral blood of children with purulent meningitis[J]. Laboratory Medicine, 2023, 38(5): 466-474.

Figures/Tables 11

References 18

[1]	VOYTENKOV V B, VILNITZ A A, SKRIPCHENKO N V, et al. Quantitative electroencephalography indicators in children with acute purulent meningitis[J]. Neurosci Behav Physiol, 2022, 52(2):315-318. DOI
[2]	HE H, GENG T, CHEN P, et al. NK cells promote neutrophil recruitment in the brain during sepsis-induced neuroinflammation[J]. Sci Rep, 2016, 6:27711. DOI PMID
[3]	HAYASHI T, NAGAI S, FUJII H, et al. Critical roles of NK and CD8+ T cells in central nervous system listeriosis[J]. J Immunol, 2009, 182(10):6360-6368. DOI PMID
[4]	BEAULIEU A M. Memory responses by natural killer cells[J]. J Leukoc Biol, 2018, 104(6):1087-1096. DOI URL
[5]	LUETKE-EVERSLOH M, HAMMER Q, DUREK P, et al. Human cytomegalovirus drives epigenetic imprinting of the IFNG locus in NKG2Chi natural killer cells[J]. PLoS Pathog, 2014, 10(10):e1004441. DOI URL
[6]	CHOREÑO-PARRA J A, JIMÉNEZ-ÁLVAREZ L A, MALDONADO-DÍAZ E D, et al. Phenotype of peripheral nk cells in latent,active,and meningeal tuberculosis[J]. J Immunol Res, 2021, 2021:5517856.
[7]	RIBES S, NESSLER S, HEIDE E C, et al. The early adaptive immune response in the pathophysiological process of pneumococcal meningitis[J]. J Infect Dis, 2017, 215(1):150-158. DOI PMID
[8]	江载芳, 申昆玲, 沈颖. 诸福棠实用儿科学[M]. 8版. 北京: 人民卫生出版社, 2015.
[9]	PELTOLA H, PELKONEN T, ROINE I, et al. Predicting outcome of childhood bacterial meningitis with a single measurement of C-reactive protein[J]. Pediatr Infect Dis J, 2016, 35(6):617-621. DOI PMID
[10]	VELISSARIS D, PINTEA M, PANTZARIS N, et al. The role of procalcitonin in the diagnosis of meningitis:a literature review[J]. J Clin Med, 2018, 7(6):148. DOI URL
[11]	SONG T, LI L, SU B, et al. NKG2C+ natural killer cell function improves the control of HBV replication in individuals with acute HIV infection coinfected with HBV[J]. Medicine(Baltimore), 2020, 99(18):e20073.
[12]	GARAND M, GOODIER M, OWOLABI O, et al. Functional and phenotypic changes of natural killer cells in whole blood during Mycobacterium tuberculosis infection and disease[J]. Front Immunol, 2018, 9:257. DOI URL
[13]	KOBYZEVA P A, STRELTSOVA M A, EROKHINA S A, et al. CD56^dim CD57^- NKG2C⁺ NK cells retaining proliferative potential are possible precursors of CD57⁺ NKG2C⁺ memory-like NK cells[J]. J Leukoc Biol, 2020, 108(4):1379-1395. DOI URL
[14]	FOREL J M, CHICHE L, THOMAS G, et al. Phenotype and functions of natural killer cells in critically-ill septic patients[J]. PLoS One, 2012, 7(12):e50446. DOI URL
[15]	GIANNIKOPOULOS G, ANTONOPOULOU A, KALPAKOU G, et al. The functional role of natural killer cells early in clinical sepsis[J]. APMIS, 2013, 121(4):329-336. DOI PMID
[16]	BLOM K, CUAPIO A, SANDBERG J T, et al. Cell-mediated immune responses and immunopathogenesis of human tick-borne encephalitis virus-infection[J]. Front Immunol, 2018, 9:2174. DOI PMID
[17]	MA L, LI Q, CAI S, et al. The role of NK cells in fighting the virus infection and sepsis[J]. Int J Med Sci, 2021, 18(14):3236-3248. DOI PMID
[18]	MAUCOURANT C, FILIPOVIC I, PONZETTA A, et al. Natural killer cell immunotypes related to COVID-19 disease severity[J]. Sci Immunol, 2020, 5(50):eabd6832. DOI URL

组别	例数	脑脊液
组别	例数	WBC计数/（×10⁶/L）	Glu/（mmol/L）	TP/（g/L）	Cl^-/（mmol/L）
PM组	52	268.00（38.75~1 254.00）	2.05（0.96~2.74）	0.91（0.52~1.90）	118.30±6.96
对照组	52	8.50（3.25~52.50）	3.25（3.02~3.51）	0.18（0.15~0.23）	124.00±2.56
统计值		-8.12	-6.76	-7.66	5.84
P值		<0.001	<0.001	<0.001	<0.001
组别		外周血
组别		hs-CRP/（mg/L）	PCT/（ng/mL）	WBC计数/（×10⁹/L）	NEUT%
PM组		77.65（23.15~156.80）	1.23（0.24~11.50）	12.59（8.82~17.30）	70.40（44.48~83.53）
对照组		13.30（5.02~24.23）	0.12（0.06~0.34）	6.32（4.67~8.43）	47.95（32.75~60.63）
统计值		-6.23	-3.74	-5.98	-3.53
P值		<0.001	<0.001	<0.001	<0.001

组别	例数	脑脊液
组别	例数	WBC计数/（×10⁶/L）	Glu/（mmol/L）	TP/（g/L）	Cl^-/（mmol/L）
PM组	52	268.00（38.75~1 254.00）	2.05（0.96~2.74）	0.91（0.52~1.90）	118.30±6.96
对照组	52	8.50（3.25~52.50）	3.25（3.02~3.51）	0.18（0.15~0.23）	124.00±2.56
统计值		-8.12	-6.76	-7.66	5.84
P值		<0.001	<0.001	<0.001	<0.001
组别		外周血
组别		hs-CRP/（mg/L）	PCT/（ng/mL）	WBC计数/（×10⁹/L）	NEUT%
PM组		77.65（23.15~156.80）	1.23（0.24~11.50）	12.59（8.82~17.30）	70.40（44.48~83.53）
对照组		13.30（5.02~24.23）	0.12（0.06~0.34）	6.32（4.67~8.43）	47.95（32.75~60.63）
统计值		-6.23	-3.74	-5.98	-3.53
P值		<0.001	<0.001	<0.001	<0.001

项目	AUC（95%CI ^①）	P值	最佳临界值	敏感性/%	特异性/%
脑脊液Cl^-	0.815（0.730~0.900）	<0.000 1	120.00 mmol/L	61.54	94.23
脑脊液Glu	0.885（0.816~0.953）	<0.000 1	2.80 mmol/L	78.85	92.31
脑脊液TP	0.936（0.887~0.984）	<0.000 1	0.34 g/L	86.54	92.31
脑脊液WBC计数	0.899（0.956~1.000）	<0.000 1	95.00×10⁶/L	73.08	100.00
血清PCT	0.823（0.743~0.903）	<0.000 1	0.16 ng/L	84.62	65.38
血清hs-CRP	0.790（0.702~0.878）	<0.000 1	21.30 mg/L	75.00	73.08
外周血WBC计数	0.840（0.765~0.915）	<0.000 1	9.47×10⁹/L	73.08	84.62
外周血NEUT%	0.701（0.599~0.802）	0.000 4	65.15%	59.62	80.77
NKG2C^-GZMB⁺ NK细胞百分比	0.980（0.956~1.000）	<0.000 1	8.93%	92.31	98.08
NKG2C⁺GZMB⁺ NK细胞百分比	0.857（0.778~0.935）	<0.000 1	4.46%	73.08	92.31
NKG2C⁺GZMB^- NK细胞百分比	0.909（0.855~0.964）	<0.000 1	3.86%	84.62	86.54

项目	AUC（95%CI ^①）	P值	最佳临界值	敏感性/%	特异性/%
脑脊液Cl^-	0.815（0.730~0.900）	<0.000 1	120.00 mmol/L	61.54	94.23
脑脊液Glu	0.885（0.816~0.953）	<0.000 1	2.80 mmol/L	78.85	92.31
脑脊液TP	0.936（0.887~0.984）	<0.000 1	0.34 g/L	86.54	92.31
脑脊液WBC计数	0.899（0.956~1.000）	<0.000 1	95.00×10⁶/L	73.08	100.00
血清PCT	0.823（0.743~0.903）	<0.000 1	0.16 ng/L	84.62	65.38
血清hs-CRP	0.790（0.702~0.878）	<0.000 1	21.30 mg/L	75.00	73.08
外周血WBC计数	0.840（0.765~0.915）	<0.000 1	9.47×10⁹/L	73.08	84.62
外周血NEUT%	0.701（0.599~0.802）	0.000 4	65.15%	59.62	80.77
NKG2C^-GZMB⁺ NK细胞百分比	0.980（0.956~1.000）	<0.000 1	8.93%	92.31	98.08
NKG2C⁺GZMB⁺ NK细胞百分比	0.857（0.778~0.935）	<0.000 1	4.46%	73.08	92.31
NKG2C⁺GZMB^- NK细胞百分比	0.909（0.855~0.964）	<0.000 1	3.86%	84.62	86.54

组别	例数	PCT/（ng/mL）	hs-CRP/（mg/L）	外周血WBC计数/（×10⁹/L）	NEUT%
预后良好组	44
治疗前		0.70（0.18~10.23）	68.90（17.45~165.75）	11.81（7.76~17.3）	66.50（43.23~85.48）
治疗后		0.06（0.04~0.09）	2.07（0.76~7.43）	6.92（6.09~8.30）	47.10（34.88~63.85）
Z值		-6.655	-6.890	-5.262	-3.046
P值		<0.001	<0.001	<0.001	<0.001
预后不良组	8
治疗前		8.84（4.22~11.62）	35.15（29.20~52.83）	13.71（10.03~17.28）	77.05（71.13~83.20）
治疗后		5.03（3.07~5.81）^#	27.15（16.68~32.71）^#	11.03（9.59~11.90）^#	78.35（76.70~82.88）^#
Z值		-1.680	-1.890	-1.470	-0.525
P值		0.092	0.059	0.141	0.600
组别		总NK细胞百分比/%	NKG2C^-GZMB⁺ NK细胞百分比/%	NKG2C⁺ GZMB⁺ NK细胞百分比/%	NKG2C⁺ GZMB^- NK细胞百分比/%
预后良好组
治疗前		8.64（6.70~11.55）	21.85（17.83~29.35）	6.16（4.87~8.10）	2.95（1.88~3.79）
治疗后		10.89（4.74~16.09）	3.83（3.24~4.36）	1.59（1.02~2.10）	15.26（12.59~20.15）
Z值		-1.494	-8.053	-7.978	-8.836
P值		0.135	<0.001	<0.001	<0.001
预后不良组
治疗前		1.56（1.11~3.91）^*	9.53（7.61~11.46）^*	1.38（0.81~2.09）^*	2.06（1.64~3.33）^*
治疗后		4.13（3.18~5.60）^#	10.87（8.04~14.63）^#	3.42（2.34~3.93）^#	2.47（2.02~2.84）^#
Z值		-2.310	-0.840	-2.415	-0.735
P值		0.021	0.401	0.016	0.462