CD_(14)^+单核细胞人类白细胞抗原-DR预测脓毒症预后及指导免疫调理治疗的初步临床研究

目的 :验证 CD+ 1 4 单核细胞人类白细胞抗原 DR( HL A DR)对于评估严重脓毒症患者免疫功能的作用 ;探讨胸腺 5肽 ( TP 5 )治疗免疫抑制的有效性。方法 :符合严重脓毒症标准 ,CD+ 1 4 单核细胞 HL A DR<30 %的患者被定义为脓毒症免疫抑制和代偿性抗炎症反应综合征 ( CARS)者纳入本研究 ,并接受 1m g TP 5肌肉注射 ,1次 / d,直至 CD+ 1 4 单核细胞 HL A DR>5 0 %或死亡。在 TP 5治疗前及结束治疗时分别测量 CD+ 1 4单核细胞 HL A DR和肿瘤坏死因子α( TNFα)、白介素 6 ( IL 6 )、IL 10和 IL 13。结果 :15例患者存活 ,5例死亡。经 TP 5治疗后所有患者的 CD+ 1 4 单核细胞 HL A DR均有不同程度提高 ,但仅存活者有显著差异。所有患者细胞因子均高于健康对照 ,治疗后存活患者的 TNFα、IL 6明显下降 ;死亡者各种细胞因子变化不明显。结论 :用 CD+ 1 4 单核细胞 HL A DR鉴别脓毒症免疫抑制并指导免疫刺激治疗是安全可靠的 ,且对评估预后有重要价值 ;TP 5可能对逆转免疫抑制有效 ,但需要严格的对照治疗研究确认 ;脓毒症的免疫抑制发生和逆转与促炎 /抗炎细胞因子平衡无关 ,确切机制有待深入探讨。

Expression of lipopolysaccharide binding protein and its receptor CD14 in experimental alcoholic liver disease

AIM: To evaluate the relationship between the expression of lipopolysaccharides (LPS) binding protein (LBP) and CD14 mRNA and the severity of liver injury in alcohol-fed rats. METHODS: Twenty Wistar rats were divided into two groups:ethanol-fed group (group E) and control group (group C). Group E was fed with ethanol(5-12 g x kg(-1) x d(-1)) and group C received dextrose instead of ethanol. Rats of the two groups were sacrificed at 4 weeks and 8 weeks. Levels of endotoxin and alanine transaminase (ALT) in blood were measured, and liver pathology was observed under light and electronic microscopy. Expressions of LBP and CD14 mRNA in liver tissues were determined by RT-PCR analysis. RESULTS: Plasma endotoxin levels were increased more significantly in group E(129+/-21) ng x L(-1) and (187+/-35) ng x L(-1) at 4 and 8 wk than in control rats(48+/-9) ng x L(-1) and (53+/-11) ng x L(-1), respectively (P【0.05). Mean values of plasma ALT levels were (1867+/-250) nkat x L(-1) and (2450+/-367) nkat x L(-1) in Group E. The values were increased more dramatically in ethanol-fed rats than in Group C after 4 and 8 weeks. In liver section from ethanol-fed rats, there were marked pathological changes (steatosis, cell infiltration and necrosis). In ethanol-fed rats, ethanol administration led to a significant increase in LBP and CD14 mRNA levels compared with the control group (P【0.05). CONCLUSION: Ethanol administration led to a significant increase in endotoxin levels in serum and LBP and CD14 mRNA expressions in liver tissues. The increase of LBP and CD14 mRNA expression might wake the liver more sensitive to endotoxin and liver injury.

Role of LPS/CD14/TLR4-mediated inflammation in necrotizing enterocolitis:Pathogenesis and therapeutic implications

AIM:To establish the roles of lipopolysaccharide (LPS)/CD14/toll-like receptor 4 (TLR4)-mediated inflammation in a rat model of human necrotizing enterocolitis (NEC).METHODS: Six pairs of intestinal samples from human NEC were collected before and after recovery for histological and molecular analysis of inflammatory cytokines and signaling components. In the rat NEC model, we isolated 10-cm jejunum segments and divided them into six groups (n=6) for sham operation, treatment with LPS, bowel distension, combined bowel distension and LPS stimulation, and two therapeutic groups. The potential eff icacy of a recombinant CD18 peptide and a monoclonal CD14 antibody was evaluated in the latter two groups. The serum and tissue levels of several inflammatory mediators were quantified by real-time polymerase chain reaction, ELISA and immunoblotting.RESULTS: Human acute phase NEC tissues displayed significant increases (P<0.05) in levels of TLR4, CD14, myeloid differentiation protein (MD)-2, tumor necrosis factor (TNF)-α and nuclear factor-κB when compared to those after recovery. The histological and inflammatory picture of human NEC was reproduced in rats that were treated with combined bowel distension and LPS, but not in the sham-operated and other control rats. Serum levels of interleukin-6 and TNF-α were also elevated. The NEC pathology was attenuated by treating the NEC rats with a monoclonal CD14 antibody or an LPS-neutralizing peptide.CONCLUSION:LPS and distension are required to produce the histological and inflammatory features of NEC. A potential treatment option is blocking LPS activation and leukocyte infi ltration.

Association between polymorphisms in the Toll-like receptor 4,CD14,and CARD15/NOD2and inflammatory bowel disease in the Greek population

AIM: Crohn's disease(CD)and ulcerative colitis(UC)are multifactorial diseases with a significant genetic background.Apart from CARD15/NOD2 gene, evidence is accumulating that molecules related to the innate immune response such as CD14 or Toll-like receptor 4 (TLR4), are involved in their pathogenesis. In further exploring the genetic background of these diseases, we investigated the variations in the CARD15/NOD2 gene (Arg702Trp,Gly908Arg and Leu1007fsinsC), and polymorphisms in the TLR4 gene (Asp299Gly and Thr399Ile) as well as in the promoter of the CD14 gene (T/C at position -159) in Greek patients with CD and UC.METHODS: DNA was obtained from 120 patients with CD,85 with UC and 100 healthy individuals. Genotyping was performed by allele specific PCR or by PCR-RFLP analysis.RESULTS: The 299Gly allele frequency of the TLR4 gene and the T allele and TT genotype frequendes of the CD14 promoter were significantly higher in CD patients only compared to healthy individuals (P = 0.026＜0.05; P = 0.0048＜0.01 and P= 0.047＜0.05 respectively). Concerning the NOD2/CARD15mutations the overall presence in CD patients was significantly higher than that in UC patients or in controls.Additionally, 51.67% of the CD patients were carriers of a TLR4 and/or CD14 polymorphic allele and at least one variant of the NOD2/CARD15, compared to 27% of the UC patients. It should be pointed out that both frequencies significantly increased as compared with the 10% frequency of multiple carriers found in healthy controls. A possible interaction of the NOD2/CARD15 with TLR4 and especially CD14, increased the risk of developing inflammatory bowel disease (IBD).CONCLUSION: Our results indicate that co-existence of a mutation in either the TLR4 or CD14 gene, and in NOD2/CARD15is associated with an increased susceptibility to developing CD compared to UC, and to developing either CD or UC compared to healthy individuals.

激素对烫伤大鼠肺脂多糖受体CD_(14)mRNA表达的影响

目的 探讨激素对烫伤后大鼠肺脂多糖受体 CD1 4 基因表达的影响及其对急性肺损伤和炎性反应综合征的治疗作用及机制。 方法 采用 SD大鼠 30 %～ 35 %三度烫伤模型 ,分别于 2 4、48h活杀 ,留取肺组织检测 CD1 4 m RNA的表达 ;留取血标本检测内毒素、IL-4的含量。 结果 烫伤后治疗组大鼠肺内皮细胞 CD1 4 m RNA表达与非治疗组比较有显著性差异 ,大剂量激素组烫伤后 2 4h IL-4水平与其他组比较有显著性差异。结论 大鼠在烫伤打击后 ,激素能抑制肺内皮细胞 CD1 4 m RNA表达上调 ,促进 IL -4的生成增加 。

CD_(44)V_6和CD_(15s)抗原在子宫内膜癌中的表达及其与临床相关性研究

目的 :探讨CD44V6、CD15s抗原表达与子宫内膜癌和侵袭转移的关系。方法 :应用免疫组化SABC方法对 5 0例子宫内膜癌标本进行CD44V6和CD15s抗原检测。结果 :CD44V6和CD15s在子宫内膜癌中阳性表达率分别为 74 %和 72 % ,两者呈正相关。CD44V6和CD15s表达与子宫内膜癌的临床分期和病理分级呈正相关 ,并随肿瘤细胞侵袭肌层深度的增加 ,阳性表达率增高。结论 :CD44V6、CD15s高表达提示子宫内膜癌生物学行为不良 ,联合检测CD44V6、CD15s可作为判断子宫内膜癌恶性程度、预测转移和评估预后的生物学指标。

组织芯片研究CD_(44)s和CD_(44)V_6在乳腺癌组织中的表达

目的 :研究CD44s和CD44V6的表达与乳腺导管癌分化、浸润及转移的关系。方法 :用组织芯片技术及免疫组织化学方法检测 1 2 0例乳腺导管癌标本和 2 0例正常乳腺标本中CD44s和CD44V6表达情况。结果 :CD44s和CD44V6在乳腺癌组织中表达阳性率分别为 4 6 % (5 5 1 2 0 )和 5 3% (6 4 1 2 0 ) ,在正常乳腺组织中均没有表达 ;CD44V6在低、中分化腺癌中表达阳性率分别为 70 % (2 4 34)和 4 6 % (4 0 86 ) ,差异有显著意义 ,χ2 =5 6 8,P <0 0 5。CD44s和CD44V6表达与乳腺导管癌淋巴结转移明显相关 ,P <0 0 1 ;而且与癌细胞浸润有关 ,P <0 0 1。结论 :CD44s和CD44V6过量表达与乳腺导管癌转移和浸润密切相关 ,联合检测CD44s和CD44V6能更准确地判断乳腺导管癌的转移状态。

结直肠癌中cyclin D1和CD_(44)V_6及E-cadherin蛋白的表达及其意义

目的 :探讨结直肠癌中cyclinD1、CD44V6、E cadherin(E cad)蛋白的表达。方法 :采用免疫组化SP法测定 48例结直肠癌中cyclinD1、CD44V6、E cad蛋白的表达。结果 :48例结直肠癌中cyclinD1、CD44V6、E cad表达的阳性率分别为5 8 3 % ( 2 8/4 8)、75 0 % ( 3 6/4 8)和 45 8%( 2 2 /4 8)。cyclinD1蛋白表达的阳性率 60岁以上年龄组高于 60岁以下年龄组 ,P<0 0 5 ,cyclinD1蛋白的表达与肿瘤组织分化程度呈负相关 ,P <0 0 1。CD44V6高表达及E cad低表达与结直肠癌Duke’s分期、浆膜浸润、淋巴结转移呈正相关 ,P<0 0 5。结论 :cyclinD1蛋白的表达与分化程度相关 ;CD44V6、E Cad蛋白的表达与结直肠癌浸润转移密切相关 ,检测cyclinD1、CD44V6、E cad蛋白是判断结直肠癌预后的客观指标。

血清胆碱酯酶、可溶性白细胞分化抗原-14、C反应蛋白与骨折术后感染相关性及临床预测价值分析

目的探究血清胆碱酯酶、可溶性白细胞分化抗原-14(sCD14)、C反应蛋白(CRP)与骨折术后感染相关性及临床预测价值。方法回顾性分析2017年3月至2019年7月淇县人民医院手术治疗的116例开放性骨折病人的临床资料,根据是否发生术后感染分为观察组(感染病人)53例,对照组(未感染病人)63例。研究两组病人的基线资料及术后1 d血清胆碱酯酶、sCD14、CRP水平,采用logistic回归分析骨折术后感染的危险因素,受试者工作特征(ROC)曲线分析血清各指标对术后感染的早期诊断价值。同时比较不同预后感染病人的基线资料及感染时、治疗2周、4周后各血清各指标水平,采用logistic回归分析骨折术后感染预后的影响因素,探讨治疗2周后的血清各指标水平对预后不良的预测价值。结果观察组手术时间(131.07±10.03)min长于对照组(108.45±10.57)min,术后1 d血清胆碱酯酶(4846.23±1355.07)U/L、sCD14(2.46±0.78)mg/L、CRP(96.54±23.49)mg/L高于对照组(3562.71±1290.69)U/L、(1.71±0.70)mg/L、(75.16±21.55)mg/L(均P<0.05);logistic回归显示,手术时间、术后1 d血清胆碱酯酶、sCD14、CRP水平升高均是开放性骨折术后感染的独立危险因素(P<0.05);ROC分析显示,术后1 d血清胆碱酯酶、sCD14、CRP对骨折术后感染诊断的曲线下面积(AUC)值均>0.7,具有较好的诊断价值,其中术后1 d血清胆碱酯酶、CRP诊断的灵敏度更高(69.81%),而sCD14诊断的特异度更高(90.48%);血清胆碱酯酶与sCD14、CRP水平呈正相关,血清sCD14与CRP呈正相关(P<0.05)。对术后感染病人进行3个月的随访,19例预后不良,34例预后良好;合并糖尿病、治疗2周后血清胆碱酯酶、sCD14、CRP水平升高是骨折术后感染病人预后不良的独立危险因素(P<0.05);ROC曲线分析显示,治疗2周的血清胆碱酯酶、sCD14、CRP水平对开放性骨折术后感染病人临床预后具有较好的预测价值(AUC均>0.7)。结论开放性骨折术后感染病人血清胆碱酯酶、sCD14、CRP水平显著升高,术后早期检测有助于感染的早期诊断。治疗2周后血清胆碱酯酶、sCD14、CRP水平对骨折术后感染病人预后具有较好的预测价值。

Cholecystokinin octapeptide inhibits the in vitro expression of CD14 in rat pulmonary interstitial macrophages induced by lipopolysaccharide

OBJECTIVE: To study the effect of cholecystokinin octapeptide (CCK-8) on lipopolysaccharide (LPS)-stimulated pulmonary interstitial macrophages (PIM) in vitro. METHODS: PIM were isolated and cultured in the presence or absence of LPS, CCK-8, proglumide (the antagonist of CCK receptors) and vehicle. The expression of membrane CD14 (mCD14) protein was assayed by flow cytometry and soluble CD14 (sCD14) in the supernatant was analyzed semi-quantitatively by Western blot. TNF-alpha in the supernatant was detected with ELISA. RESULTS: CCK-8, at concentrations of 10(-7) mol/L and 10(-6) mol/L, significantly inhibited the expression of mCD14. Release of sCD14 and TNF-alpha in the supernatant was up-regulated by LPS (1 microg/ml) but reduced by CCK-8. The effect of CCK-8 was inhibited by proglumide. CONCLUSION: CCK-8 negatively modulated several functions of LPS-stimulated PIM through CCK receptors. This may be one of the mechanisms for CCK-8 to alleviate inflammation in lung tissue during endotoxemia.

严重烧伤患者外周血单核细胞表面人白细胞DR抗原变化的初步研究

目的 :探讨烧伤后外周血单核细胞表面人白细胞 DR抗原受体 (HL A DR)的动态变化规律及意义。方法 :选取临床烧伤患者 30例 ,依据病程长短选取病程中 1～ 5个时间点静脉采血 ,以流式细胞仪测定外周血单核细胞 HL A DR的表达率 ,并根据烧伤程度分组进行分析。结果 :伤后患者外周血单核细胞 HL A DR的表达率明显降低 ,降低程度及持续时间与伤情有关 ,特重烧伤患者与中度烧伤患者〔(4.30± 1.5 0 ) %比(13.86 %± 2 .4 0 ) %〕、中度烧伤患者与轻度烧伤患者〔(13.86± 2 .4 0 ) %比 (5 8.80± 5 .6 0 ) %〕比较差异均显著(P均 <0 .0 1)。结论 :单核细胞 HL A DR的表达率是反映免疫功能的简单实用的指标。重症烧伤后免疫麻痹可持续较长时间 ,必要的免疫加强治疗有重要意义。

2型糖尿病患者血清可溶性CD_(14)水平的改变及其意义

在正常对照者 ,并发或未并发肾病的 2型糖尿病患者测定血清可溶性CD1 4(sCD1 4)、血糖、血脂、HbA1c、尿白蛋白等。结果显示血清sCD1

Mobilization of hematopoietic progenitor cells in patients with liver cirrhosis

AIM:To test the hypothesis that liver cirrhosis is associated with mobilization of hematopoietic progenitor cells. METHODS:Peripheral blood samples from 72 patients with liver cirrhosis of varying etiology were analyzed by flow cytometry.Identified progenitor cell subsets were immunoselected and used for functional assays in vitro. Plasma levels of stromal cell-derived factor-1(SDF-1) were measured using an enzyme linked immunosorbent assay.RESULTS:Progenitor cells with a CD133 + /CD45 + CD14 + phenotype were observed in 61%of th patients.Between 1%and 26%of the peripheral bloo mononuclear cells(MNCs)displayed this phenotype Furthermore,a distinct population of c-kit + progenito cells(between 1%and 38%of the MNCs)could b detected in 91%of the patients.Additionally,18% of the patients showed a population of progenito cells(between 1%and 68%of the MNCs)that wa characterized by expression of breast cancer resistanc protein-1.Further phenotypic analysis disclosed tha the circulating precursors expressed CXC chemokin receptor 4,the receptor for SDF-1.In line with thi finding,elevated plasma levels of SDF-1 were presen in all patients and were found to correlate with th number of mobilized CD133 + progenitor cells.

Effect of Early Intensive Insulin Therapy on Immune Function of Aged Patients with Severe Trauma

This study examined the effect of intensive insulin therapy on immune function and inflammatory factors at the early phase after severe trauma. At day 1, 3, 5, 7 after admission, subsets of CD4＋ helper T lymphocytes （Th1/Th2） and human leukocyte antigen （HLA）-DR expression on CD14＋ monocytes were flow cytometrically measured. Levels of cytokines, including tumor necrosis factor-alpha （TNF-α）, interleukin-6 （IL-6）, interleukin-10 （IL-10） and other immunity markers, such as IgA, IgG, IgM, C3, C4 and C reaction protein （CRP） were examined in two groups. The results showed that TNF-α, IL-6 and CRP levels in the intensive insulin therapy group were significantly lower than those in the conventional therapy group, whereas IL-10 levels were substantially increased after intensive insulin therapy. C3 level at day 3, 5, 7 and C4 levels at day 5, 7 were lower in the intensive therapy group than in the conventional therapy group. Th1/Th2 ratios decreased gradually over time in both groups, and were much lower at day 3, 5, 7 in intensive therapy group. There were significant differences among day 3 to day 7 after admission in HLA-DR expression in CD14＋ monocytes. It was concluded that the intensive insulin therapy could decrease pro-inflammatory cytokines and increase anti-inflammatory cytokines in the elderly suffering from severe trauma, at the same time, with complement recovery being delayed. Moreover, intensive insulin therapy promoted immune suppression and, therefore, measures need be taken to address the issue.

Immunophenotype of myeloid cells in myelodysplastic syndromes and its clinical implications

OBJECTIVE: To explore the immunophenotype of myeloid cells in myelodysplastic syndyomes (MDS) and its clinical implications. METHODS: A panel of monoclonal antibody was used to detect CD13+, CD33+, CD15+ and CD14+ antigens on the membrane surfaces of myeloid cells in the bone marrow from 51 MDS, 21 aplastic anemia (AA), 21 paroxysmal nocturnal hemoglobinuria (PNH) patients. 10 acute myeloblastic leukemia (AML) patients and 15 normal subjects by immunoenzymatic assay. The morphology and chromosome karyotype of bone marrow cells of MDS patients were also examined. RESULTS: CD14+, CD13+ and CD33+ cells in the bone marrow were more in MDS patients than in normal controls, AA patients and PNH patients. CD15+ cells in the bone marrow were less in MDS patients than in normal controls. CONCLUSIONS: The percentages of CD14+, CD13+ and CD33+ positive cells in the bone marrow of MDS patients were related to the percentage of myeloblasts, the chromosomal aberrations and the response to treatment. It indicated that there is immunophenotypic misexpression of myeloid cells in MDS patients. Immunophenotype analysis of myeloid cells might be useful for the diagnosis and treatment of MDS patients.

Lipopolysaccharide/Toll-like receptor 4 signaling pathway involved Qingdu decoction treating severe liver injury merging with endotoxemia

OBJECTIVE: To investigate the protective effects and underlying mechanism of Qingdu decoction(QDD) on experimental rats with severe liver injury induced by thioacetamide(TAA).METHODS: A total of 40 Wistar rats were randomly divided into normal group(n = 10) and experimental group(n = 30). Rats were administrated the same content of saline in normal group. The rats inthe experimental group were pretreated with TAA at dose of 12 mg/kg lasting 8 weeks, and from 9th week to 12 th week, with TAA at concentration of 36mg/kg. During the 9th week to 12 th week period,the rats were randomly divided into three subgroups(n = 10 each) simultaneously based on the treatment categories: model group, lactulose(LA,3.5 m L/kg) group and QDD(5.95 g/kg) group, orally once per day respectively. At the 12 th week, the content of serum alanine transaminase(ALT), aspartate transaminase(AST), total bilirubin(TBIL), endotoxin(ET) and tumor necrosis factor α(TNF-α) was detected by automatic biochemical analyzer. The plasma prothrombin time(PT), prothrombin time-international normalized ratio(PTR) and prothrombin time activity(PTA) were measured by automatic coagulation analyzer. The level of lipopolysaccharide(LPS)-binding protein(LBP), cluster differentiation 14(CD14) and Toll-like receptor 4(TLR4) expressions was measured by both western blot(WB) and real-time polymerase chain reaction(real-time PCR).RESULTS: Compared with the model group, hepatic morphology in the QDD group was improved under light microscope and transmission electron microscope; at the same time, the contents of serum ALT, AST, TBIL, ET and TNF-α, and level of LBP, CD14 and TLR4 expressions in liver tissues were significantly decreased compared with the model group(P < 0.05), while PTA in the QDD group was enhanced(P < 0.05).CONCLUSION: QDD has the functional effect on improving the injured liver through inhibiting the LPS/TLR4 signaling pathway thus decreasing the level of the inflammatory medicators.