病毒载量和细胞因子在发热伴血小板减少综合征致病机制中的作用

Viral load and cytokines in the pathogenesis of severe fever with thrombocytopenia syndrome

目的：观察新型布尼亚病毒感染后病毒载量、血清细胞因子与组织损伤的关系，探讨该病毒感染水平对组织损伤及预后的影响。方法南京医科大学第一附属医院感染病科2011年5月至2012年7月门诊及住院的新型布尼亚病毒感染的严重发热伴血小板减少综合征（SFTS）患者24例，根据患者临床预后分为痊愈组21例和死亡组3例。另选取南京市中心血站健康献血者32名作为对照。采用荧光定量 PCR 检测血清病毒载量，用流式细胞仪动态定量检测患者血清中 Th1、Th2、Th17细胞因子，分析其与血清酶学、白细胞和血小板之间的关系。组间资料采用秩和检验，血清病毒载量与血清中细胞因子含量、血细胞计数、组织损伤的相关性采用 Spearman 相关分析。结果24例患者新型布尼亚病毒 RNA 检测结果均为阳性。痊愈组21例患者中有2例病毒载量＞7．0 lg 拷贝/mL。死亡组3例患者的病毒载量分别为6．7 lg 拷贝/mL、8．8 lg 拷贝/mL 和9．8 lg 拷贝/mL。SFTS 痊愈组患者在病程第5天和第7天，血清 IL-6分别为21．76和7．12 pg/mL，显著高于健康对照组的2．82 pg/mL，差异均有统计学意义（均 P ＜0．05）。SFTS 痊愈组患者病程第5天、第7天和第9天，血清 IL-10分别为14．33、14．13和3．01 pg/mL，显著高于健康对照组的1．56 pg/mL。死亡组患者在病程第7天、第9天，血清IL-6为137．61和1450．83 pg/mL，IL-10为50．26和49．43 pg/mL，较痊愈组患者增高，差异均有统计学意义（均 P ＜0．05）；病程中 SFTS 患者血清 IL-2、IL-4则显著低于健康对照组（均 P ＜0．05）。SFTS痊愈组患者白细胞和血小板计数的最低值出现在病程初期，血清 ALT、AST、乳酸脱氢酶（LDH）、CK 等均明显高于正常范围。相关性分析显示，SFTS 患者血清 IL-6、IL-10与血小板计数呈负相关（r 值分别为-0．390和-0．608，均 P ＜0．01），与病毒载量（r 值分别为0．560和0．758）、ALT（r 值分别为0．412和0．390）、AST（r 值分别为0．686和0．764）、LDH（r 值分别为0．633和0．677）、CK（r 值分别为0．527和0．636）等酶学指标呈正相关（均 P ＜0．01）。结论SFTS 患者血清病毒载量、IL-6、IL-10以及血清酶学水平与疾病严重程度密切相关，新型布尼亚病毒感染后血清中炎性因子和抑炎因子风暴，可能参与了SFTS 患者的免疫病理损伤。

Objective To observe the relationship of viral load,serum cytokines and tissue damage after severe fever with thrombocytopenia syndrome virus （SFTSV）infection,and to explore the impact of SFTSV levels on tissue injury and prognosis.Methods Twenty-four ambulatory and hospitalized patients who were infected with SFTSV were enrolled between May 2011 and July 2012 at Department of Infectious Disease, First Affiliated Hospital with Nanjiang Medical University. According to their prognosis,they were divided into cure and death group,while 32 healthy blood donators were also enrolled from center blood station in Nanjing as control.The serum SFTSV load was detected using fluorescence quantitative polymerase chain reaction （PCR）.The serum T helper （Th）1/Th2/Th17 cytokines in patients with severe fever with thrombocytopenia syndrome （SFTS）were determined dynamically and quantitatively by flow cytometry.The relationships between viral load,cytokines and serum enzymes, white blood cell （WBC）,platelet （PLT）counts were analyzed.Comparisons among groups were achieved by rank sum test and correlation analysis among serum cytokines,blood cell counts and tissue damage was done by Spearman correlation test.Results All of the 24 patients showed a positive reaction to SFTSV RNA.The SFTSV loads of 21 cured cases,those of 2 were 〉 7.0 lg copy/mL,and those of 3 death patients were 6.7 lg copy/mL,8.8 lg copy/mL and 9.8 lg copy/mL,respectively.Serum level of interleukin （IL）-6 （21 .76 pg/mL in day 5 and 7.12 pg/mL in day 7）and IL-10 （14.33 pg/mL in day 5 , 14.13 pg/mL in day 7 and 3.01 pg/mL in day 9）of cured patients were significantly higher than those of healthy controls （IL-6：2.82 pg/mL and IL-10：1 .56 pg/mL）（P 〈0.05 ）.At day 7 and day 9,serum levels of IL-6 of death cases were 137.61 pg/mL and 1 450.83 pg/mL,respectively and serum levels of IL-10 were 50.26 pg/mL and 49.43 pg/mL,respectively.Both of the indicators in the death group were significantly higher than those of cure group （P 〈0.05 ）.However,serum levels of IL-2 and IL-4 were significantly lower than those in healthy control group （P 〈0.05 ）.In the cure group,WBC and PLT counts were lowest during the early course of the disease,and serum alamine aminotransferase （ALT）, aspartate aminotransferase （AST ）, lactic dehydrogenase （LDH ） and creatine kinase （CK ） were significantly higher than their upper limits of normal.The correlation analysis showed that serum IL-6, IL-10 levels were negatively correlated with PLT count （r=-0.390 and -0.608,respectively;both P 〈0.01）,and positively correlated with SFTSV load （r=0.560 and 0.758,respectively）,ALT （r=0.412 and 0.390,respectively）,AST （r = 0.686 and 0.764,respectively）,LDH （r = 0.633 and 0.677, respectively）and CK （r =0.527 and 0.636,respectively）（all P 〈0.01 ）.Conclusions SFTSV load, IL-6,IL-10 and serum enzyme levels are closely related to the severity of the disease.The inflammatory and anti-inflammatory cytokine storm after SFTSV infection may be involved in the immune pathological injury in patients with SFTS.