Human neutrophil antigen‐1, -3, -4, and -5 allele and genotype frequencies in the Croatian blood donor population and their clinical significance

ObjectivesHuman neutrophil antigens (HNAs) and antibodies play an important role in allo- and autoimmunity associated with immune neutropenia and transfusion reactions. The aim of this study was to determine the HNA-1, -3, -4 and -5 allele and genotype frequencies in the Croatian blood donor population to assess the role of HNA-1, -3, -4, and -5 alleles in the development of neonatal alloimmune neutropenia and antibody-mediated transfusion-related acute lung injury.Material and methodsA total of 371 blood samples from unselected healthy blood donors were analyzed. Samples from all 371 donors were genotyped for HNA-1, samples from 160 donors were genotyped for HNA-3, and samples from 142 donors were genotyped for HNA-4 and HNA-5 using the polymerase chain reaction with sequence-specific primers (PCR-SSP) method.ResultsThe frequencies of the FCGR3B*01, FCGR3B*02 and FCGR3B*03 HNA-1 alleles were 0.393, 0.607 and 0.022, and of the SLC44A2*01 and SLC44A2*02 HNA-3 alleles 0.781 and 0.219, respectively. The frequencies of the ITGAM*01 and ITGAM*02 HNA-4 alleles were 0.796 and 0.204, and of the ITGAL*01 and ITGAL*02 HNA-5 alleles 0.718 and 0.282, respectively.ConclusionThese are the first results on the HNA allele and genotype frequencies in the Croatian blood donor population. We observed no deviations from previous reports on Caucasian populations. Determination of the HNA antigen frequencies in the population is important to estimate the risk of alloimmunization to HNA, especially the risk of fetal-maternal incompatibility and alloantibody production by transfusion of the HNA incompatible blood components.     

The regulation by phosphorylation of ‘priming' of phospholipase A2 activity in the neutrophil model system,differentiated HL60 cells

1. Differentiated HL60 cells have been utilized as a model system to examine the ‘priming'' of neutrophil phospholipase A₂ activity. In control cells activation of phospholipase A₂ by a 5 min stimulation with the chemotactic peptide formyl-methionyl-leucyl-phenylalanine (100 nM) was essentially undetectable. When cells were primed by preincubation with 5 μM cytochalasin B for 5 min arachidonate release, a measure of phospholipase A₂ activation, was observed within 20 s.
2. Priming by cytochalasin B did not involve or require a change in intracellular free calcium concentration.
3. Priming was associated with an increase in general protein tyrosine phosphorylation and could also be induced by the receptor tyrosine kinase agonist granulocyte macrophage colony-stimulating factor (GM-CSF, 20 ng ml⁻¹) and be mimicked by treatment with the phosphotyrosine phosphatase inhibitor perhydrovanadate (0.5 mM). However, an increase in MAP kinase activity was not involved in the priming process.
4. Western blot analysis demonstrated that phospholipase A₂ was phosphorylated in both control and primed cells, but that an increase in the amount of membrane associated enzyme was found in the primed cells.
5. Thus priming appears to be due to membrane association of the phospholipase and this may be regulated by tyrosine kinase activities.

     

Release of chemoattractants and neutrophil activation in acute myocardial infarction immediately after successful recanalization of the infarct-related vessel by angioplasty

The study investigated inflamatory responses in evolving myocardialinfarction. Fifteen patients with acute myocardial infarction,who had undergone balloon recanalization of the infarct-relatedcoronary artery within 4 h after onset of symptoms, were examined.Blood samples were obtained through the guiding catheter andfrom the pulmonary artery before and immediately after successfulrecanalization. After recanalization, plas from the pulmonaryartery was 47% (quartiles: l9%, 78; P =0·001) more chemotacticto neutrophils from normal donors than before recanalization.Furthermore, significant changes in neutrophil function werefound in the pulmonary artery. Compared to the values beforerecanalization, the nitroblue tetrazolium score rose by 31%(quartiles: 4%, 37% P=0·003), FMLP-stimulated superoxideanion production by 10% (quartiles: 0%, 39% P=0·020),and chemotaxis by 46% (quartiles: 0%, 81%, P=0·011),while neutrophil filterability decreased by 28% (quartiles:15%, 47%; P=0·010). No significant changes in neutrophilparameters were found in the arterial blood The study indicatesthat chemoattractants are released in the early reperfusionperiod of evolving myocardial infarction. These chemoattractantsmay act as inflammatory mediators causing neutrophil activation.     

Acute inflammatory responses to mechanical lesions in the CNS: differences between brain and spinal cord

Lesion-induced inflammatory responses in both brain and spinal cord have recently become a topic of active investigation. Using C57BL/6J mice, we compared the tissue reaction in these two central nervous system (CNS) compartments with mechanical lesions of similar size involving both grey and white matter. This evaluation included the quantitative assessment of neutrophils, lymphocytes and activated macrophages/microglia, as well as astrocyte activation, upregulation of vascular cell adhesion molecules (ICAM-1, VCAM-1, PECAM) and the extent of blood-brain barrier (BBB) breakdown. Time points analysed post-lesioning included 1, 2, 4 and 7 days (as well as 10 and 14 days for the BBB). We found clear evidence that the acute inflammatory response to traumatic injury is significantly greater in the spinal cord than in the cerebral cortex. The numbers of both neutrophils and macrophages recruited to the lesion site were significantly higher in the spinal cord than in the brain, and the recruitment of these cells into the surrounding parenchyma was also more widespread in the cord. The area of BBB breakdown was substantially larger in the spinal cord and vascular damage persisted for a longer period. In the brain, as in spinal cord, the area to which neutrophils were recruited correlated well with the area of BBB breakdown. It will be of interest to determine the extent to which the infiltration of inflammatory cells contributes, either directly or indirectly, to the vascular permeability and secondary tissue damage or, conversely, to local tissue repair in the brain and the spinal cord.     

Activation of neutrophil function by recombinant human granulocyte colony-stimulating factor improves the survival of rats with peritonitis

We examined the effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF) and cefmetazole sodium on survival, neutrophil count, and neutrophil function in rats with peritonitis produced by cecal ligation and puncture. Rats with peritonitis received either rhG-CSF (50 or 100 g/kg) with or without cefmetazole (50 mg/kg) for 3 days, cefmetazole alone, or no treatment and were evaluated as controls. The mortality rate of all treated rats was significantly lower than that of the untreated rats. The survival rate was 57.1% for the rats given both rhG-CSF and cefmetazole, but there was no significant improvement of survival as compared with cefmetazole therapy alone. Treatment with rhG-CSF at 100 |Gmg/kg caused the circulating neutrophil count to increase significantly. The phagocytic activity for latex beads and neutrophil H₂O₂ production showed a greater enhancement by phorbol myristate acetate (PMA) in the untreated rats, thus indicating that neutrophils from treated rats were more activated. These findings show that rhG-CSF can improve survival and neutrophil function in rats with peritonitis, while combined therapy with cefmetazole was also found to be beneficial.     

Role of neutrophils in the synergistic liver injury from monocrotaline and bacterial lipopolysaccharide exposure.

Synergistic liver injury develops in Sprague-Dawley rats from administration of a small, noninjurious dose (7.4 x 10(6) EU/kg) of bacterial lipopolysaccharide (LPS) given 4 h after a nontoxic dose (100 mg/kg) of the pyrrolizidine alkaloid, monocrotaline (MCT). Previous studies demonstrated that liver injury is mediated through inflammatory factors, such as Kupffer cells and tumor necrosis factor alpha (TNF-alpha), rather than through simple interaction between MCT and LPS. In the present study, the hypothesis that neutrophils (polymorphonuclear leukocytes or PMNs) are causally involved in this injury model is tested, and the interdependence between PMNs and other inflammatory components is explored. Hepatic PMN accumulation and the appearance of cytokine-induced neutrophil chemoattractant-1 in plasma preceded the onset of liver injury, suggesting that PMNs contribute to toxicity. Hepatic PMN accumulation was partially dependent on TNF-alpha. Prior depletion of PMNs in MCT/LPS-cotreated animals resulted in attenuation of both hepatic parenchymal cell (HPC) and sinusoidal endothelial cell (SEC) injury at 18 h. PMN depletion did not, however, protect against early SEC injury that occurred before the onset of HPC injury at 6 h. This observation suggests that SEC injury is not entirely dependent on PMNs in this model. In vitro, MCT caused PMNs to degranulate in a concentration-dependent manner. These results provide evidence that PMNs are critical to the HPC injury caused by MCT/LPS cotreatment and contribute to the progression of SEC injury.     

III期结直肠癌中NLRC3与预后和肿瘤免疫的关系

目的：探讨核苷酸寡聚化结构域样受体家族半胱天冬酶募集结构域蛋白3(nucleotide binding oligomerization domain-like receptor family caspase recruitment domain containing 3,NLRC3)与III期结直肠癌的预后及肿瘤免疫相关指标 的关系。方法：回顾性收集中南大学湘雅医院2012年至2013年122例经手术根治切除的III期结直肠癌患者的相关资 料。利用免疫组织化学法分析NLRC3与CD8+ T细胞的表达情况,利用患者的术前临床资料计算中性粒细胞/淋巴细 胞比值(neutrophil to lymphocyte ratio,NLR),并检测其微卫星稳定性。采用χ2检验分析NLRC3与临床病理因素之间的 关系,采用COX回归模型分析III期结直肠癌的独立预后因素。结果：在III期结直肠癌中,肿瘤组织CD8+ T细胞浸润 分期(χ2=27.79,P<0.01)、NLR值(χ2=6.35,P<0.05)、淋巴结转移分期(χ2=10.12,P<0.01)以及微卫星稳定性(χ2=6.05, P<0.05)与NLRC3的表达有关。NLRC3(OR=0.066,95% CI：0.020~0.218)、血管癌栓(OR=3.119,95% CI：1.547~6.286) 及NLR(OR=5.103,95% CI：2.465~10.563)对III期结直肠癌的5年总生存期(overall survival,OS)有影响(均P<0.05);另 外,NLRC3(OR=0.144,95% CI：0.055-0.377)、血管癌栓(OR=3.589,95% CI：1.859~6.932)及NLR(OR=2.939,95% CI： 1.509~5.723)对III期结直肠癌的无病生存期(disease free survival,DFS)同样有影响(均P<0.05)。结论：NLRC3,血管癌栓 和NLR是III期结直肠癌的独立预后因素。NLRC3通过抑制系统性炎症、促进局部抗肿瘤免疫而使III期结直肠癌患者 具有良好的预后。     

重组人Elafin转染气道上皮对炎性损伤因子攻击的保护作用

目的探讨重组人Elafin对炎症损伤因子中性粒细胞弹性蛋白酶(NE)攻击气道上皮保护作用的分子机制。方法通过构建人中性粒细胞弹性蛋白酶抑制剂Elafin真核表达载体pEGFP-C1-Elafin,并将其转染入NCI-H292细胞中,再用中性粒细胞弹性蛋白酶(NE)刺激24h后,用底物法测定培养上清中NE的活性,Westernbolt检测细胞中ZO-1表达。结果转染Ela-fin NE组培养上清液中NE活性与转染空载体的细胞相比较明显降低,而细胞内ZO-1蛋白含量明显增高。结论NE是引起气道慢性炎症的终效因子,通过转染重组人Elafin可对抗NE破坏气道上皮完整性的作用,增强气道抗感染能力。     

The overall benefits of empagliflozin treatment in adult siblings with glycogen storage disease type Ib: one year experience

IntroductionRecently published case reports suggest the benefit of empagliflozin use in subjects with glycogen storage disease Ib (GSD Ib).MethodsWe present the clinical and laboratory data of 2 adult brothers with GSD Ib treated with empagliflozin for 12 months.ResultsThere was no severe infection during administration of empagliflozin. The improvement of clinical symptoms of inflammatory bowel disease and arthritis along with reduction in serum CRP levels and urinary albumin excretion was noted. Neutrophil count increased, allowing for reduction or temporary withdrawal of G-CSF treatment.ConclusionsEmpagliflozin may be a new safe treatment in GSD Ib patients with an advanced stage of the disease.     

Assessment of neutrophil to lymphocyte ratio,C-reactive protein,mean platelet volume in obese,and nonobese patients with polycystic ovary syndrome

Polycystic ovary syndrome (PCOS) is associated with low-grade chronic inflammation.This was a retrospective case–control study.In the present study, the risk coefficients of neutrophil to lymphocyte ratio (NLR), high-sensitive C-reactive protein (hs-CRP), and mean platelet volume (MPV) in obese patients with PCOS were determined. This study was designed to investigate NLR, hs-CRP, and MPV levels in 68 obese patients with PCOS and 44 nonobese patients with PCOS, and our study group was matched with 47 obese and 43 nonobese controls, respectively.PCOS group had higher MPV, NLR, insulin, glucose, and HOMA-IR rates than those of the controls. Subgroup analyses revealed that the obese PCOS group had higher NLR, hs-CRP, and MPV levels compared to those of controls. The obese PCOS group had higher NLR, hs-CRP, and MPV levels compared to those of the nonobese PCOS group. The odds ratios and 95% confidence intervals of those variables (NLR, hs-CRP, MPV) were found significant (P < .05). NLR, hs-CRP, and MPV variables were found statistically significant in the analysis of receiver operating characteristics.Our study demonstrated that NLR, hs-CRP, and MPV levels are increased in patients with obese PCOS.