Physiology of red and white blood cells

Blood is made up of plasma and formed elements, which are red blood cells, white blood cells and platelets. The red blood cells (erythrocytes) make up the vast majority of the cells present in the blood. Their principal function is the transport of oxygen from the lungs to the tissues and the transport of carbon dioxide from those tissues back to the lungs. This is due to the presence of haemoglobin, a protein that binds easily and reversibly with oxygen. The affinity of haemoglobin for oxygen changes under certain conditions allowing increased offloading of oxygen at the respiring tissues as required. White blood cells (leucocytes) form the body's defence against invading pathogens. They can be subdivided into granulocytes and agranulocytes, which have different mechanisms of attack against those pathogens.     

An automated image analysis system can be beneficial in preclassification of leucocytes in children with hematological disease

This study was aimed to evaluate the analytical performance of an automated image analysis system (a pilot model of Diff Master^? Octavia) for the preclassification of leucocytes in children with hematological disease. Manual microscopy performed by pediatric hematologists was used as the reference method. Five mature cell class and blasts were evaluated. Diff Master Octavia correctly preclassified 87.4% of all leucocytes with a high reproducibility. The overall accuracy was found to be 93.0%. Clinical sensitivity was 97.7% and specificity was 76.0%. The average time per slide for Diff Master^? Octavia was 2.3 min lower than that of manual method. Our results indicated that the Diff Master^? Octavia can detect and preclassify leucocytes accurately; therefore, it can be used as an efficient and fast method in pediatric hematology routine. J. Clin. Lab. Anal. 25:71–75, 2011. © 2011 Wiley‐Liss, Inc.     

Comparative study of the expression of metalloproteases and their inhibitors in different localizations within primary tumours and in metastatic lymph nodes of breast cancer

Studies on metastasic lesions from human carcinomas are scarce. Therefore there is a need for such studies to identify the expression of the biological factors that will help in the assessment of the natural history of breast cancer. Here an immunohistochemical study was performed using tissue arrays and specific antibodies against matrix metalloproteinases (MMPs)‐1, 2, 7, 9, 11, 13, 14 and tissue inhibitors of metalloproteases (TIMPs)‐1, 2 and 3 in 39 patients with breast cancer. Specimens from 39 patients with node‐positive carcinomas were examined and the analysis was performed at the central core of the tumour, at the invasive front, and in the metastasic axillary lymph nodes (MALNs). Global expression of MMP‐1, 7 and 14, TIMP‐1, and 3, were significantly higher at the centre of the tumour compared with the invasive front or the MALNs. Significantly higher expression of MMP‐7 and 14, and TIMP‐3, by fibroblast‐like cells and mononuclear inflammatory cells (MICs) was seen in MALNs. In addition, in the tumour centre, the expression of MMP‐11 and TIMP‐1 and 2 by MICs, as well as TIMP‐2 expression by fibroblast‐like cells, were associated significantly with the occurrence of distant metastasis. In contrast, TIMP‐3 expression by tumour cells or by fibroblast‐like cells in this same tumour locations, as well as TIMP‐1 expression by fibroblast‐like cells at the invasive front, were associated significantly with poor prognosis. However, the expression of all of these biological factors in MALNs was not associated with the development of distant metastasis. Our data suggest that there is prognostic relevance to the expression of MMPs and TIMPs in the stromal cells of primary tumours, rather than to the expression of these enzymes in MALNs.     

Salmonid T cells assemble in the thymus, spleen and in novel interbranchial lymphoid tissue

In modern bony fishes, or teleost fish, the general lack of leucocyte markers has greatly hampered investigations of the anatomy of the immune system and its reactions involved in inflammatory responses. We have previously reported the cloning and sequencing of the salmon CD3 complex, molecules that are specifically expressed in T cells. Here, we generate and validate sera recognizing a peptide sequence of the CD3ε chain. Flow cytometry analysis revealed high numbers of CD3ε(+) or T cells in the thymus, gill and intestine, whereas lower numbers were detected in the head kidney, spleen and peripheral blood leucocytes. Subsequent morphological analysis showed accumulations of T cells in the thymus and spleen and in the newly discovered gill-located interbranchial lymphoid tissue. In the latter, the T cells are embedded in a meshwork of epithelial cells and in the spleen, they cluster in the white pulp surrounding ellipsoids. The anatomical organization of the salmonid thymic cortex and medulla seems to be composed of three layers consisting of a sub-epithelial medulla-like zone, an intermediate cortex-like zone and finally another cortex-like basal zone. Our study in the salmonid thymus reports a previously non-described tissue organization. In the intestinal tract, abundant T cells were found embedded in the epithelium. In non-lymphoid organs, the presence of T cells was limited. The results show that the interbranchial lymphoid tissue is quantitatively a very important site of T cell aggregation, strategically located to facilitate antigen encounter. The interbranchial lymphoid tissue has no resemblance to previously described lymphoid tissues.     

Interaction of amphotericin B lipid formulations and triazoles with human polymorphonuclear leucocytes for antifungal activity against Zygomycetes

The frequency of zygomycosis has increased considerably over recent years mainly in immunocompromised and diabetic patients. Little is known about the effects of host innate immunity against different Zygomycetes especially under the influence of antifungal agents. The antifungal activity of human polymorphonuclear leucocytes (PMN) in combination with liposomal amphotericin B (LAMB), amphotericin B lipid complex (ABLC), voriconazole (VRC) and posaconazole (PSC) against Rhizopus oryzae and Rhizopus microsporus, frequently isolated Zygomycetes, were studied and compared with Absidia corymbifera, a less pathogenic Zygomycete. Antifungal activity was evaluated as per cent of hyphal damage using the XTT metabolic assay. While A. corymbifera was more susceptible to PMN than the other two Zygomycetes, R. microsporus appeared to be the most susceptible to combined effects of amphotericin B formulations and VRC with PMN. LAMB exhibited synergistic activity with PMN in inducing hyphal damage to R. microsporus but not to the other fungi. In contrast, ABLC exhibited synergistic or additive activity with PMN against all three fungi. Among triazoles, only VRC exhibited additive effect with PMN against R. microsporus. Lipid formulations of amphotericin B and particularly ABLC interact with PMN predominantly in inducing augmented hyphal damage to three different species of Zygomycetes.     

姜黄素对脓毒症大鼠肾组织ICAM-1表达和MPO活性的影响

目的 观察姜黄素对脓毒症大鼠肾组织ICAM-1 mRNA表达、MPO活性的影响,探讨其保护肾功能的可能机制.方法 90只SPF级SD大鼠随机分为假手术组(Sham组)、盲肠结扎穿孔组(CLP组)、姜黄素治疗组(Cur组),CLP组和Cur组进行盲肠结扎穿孔术,Cur组予姜黄素液2mL·kg-1(即100mg·kg-1)腹腔注射,Sham组、CLP组给予2mL·kg-1生理盐水腹腔注射.各组大鼠于术后0、3、6、12、24、48 h 活杀动物取材,观察姜黄素处理后大鼠肾组织ICAM-1 mRNA表达、髓过氧化物酶(MPO)活性及血清尿素氮(BUN)、肌酐(Scr)的变化.结果 CLP术6h后Cur组肾组织ICAM-1 mRNA表达、MPO活性明显低于CLP组(P＜0.05),术后48h血清BUN 、Scr水平亦明显低于CLP组(P＜0.05).结论 姜黄素对脓毒症大鼠肾损伤的保护作用可能与其抑制ICAM-1表达,减少过量中性粒细胞在肾组织中聚集有关.     

The dorsal skinfold chamber: A valuable model for the in vivo evaluation of topical formulations

In this study, we introduce the mouse dorsal skinfold chamber model as a valuable approach for the in vivo evaluation of topical formulations. For this purpose, dorsal skinfold chambers were implanted into BALB/c mice. Tumor necrosis factor (TNF)‐α was administered to the chamber tissue for the local induction of inflammation followed by the application of diclofenac‐containing or diclofenac‐free (control) gel onto the skin of the chamber backside. Intravital fluorescence microscopy was repetitively performed throughout an observation period of 24 hours to study macromolecular leakage, leucocyte‐endothelial cell interactions and microhaemodynamic parameters. In addition, infiltration of the inflamed tissue with different immune cell subtypes was assessed by immunohistochemistry. In a second set of experiments, the effect of dermal application of a diclofenac‐containing gel on photochemically induced thrombus formation was analysed. It was observed that macromolecular leakage, numbers of adherent leucocytes and tissue infiltrating myeloperoxidase (MPO)‐positive neutrophilic granulocytes and CD68‐positive macrophages were significantly reduced in dorsal skinfold chambers treated with diclofenac‐containing gel when compared to controls. Moreover, the diclofenac‐containing gel exerted an anti‐thrombotic activity, as indicated by a significantly prolonged complete vessel occlusion time. These findings demonstrate that the mouse dorsal skinfold chamber represents a valid and versatile tool to evaluate the effects of topical formulations in vivo.     

Glutathione metabolism in activated human neutrophils: stimulation of glutathione synthesis and consumption of glutathione by reactive oxygen species

Since glutathione (GSH) is involved in the modulation of the function of polymorphonuclear leucocytes (PMN) such as phagocytosis and production of reactive oxygen species, the metabolism of GSH was studied in human PMN. The concentration of GSH in resting PMN amounted to 13.3 nmol 10(-7) PMN and remained stable over 100 min of incubation. Upon activation of PMN with phorbol myristate acetate intracellular GSH decreased to 50% of the resting concentration within 80 min. In the presence of buthionine sulfoximine, which inhibits the synthesis of GSH, the depletion of intracellular GSH was dramatically accelerated, indicating that activation of PMN is associated with a marked stimulation of GSH synthesis. Since a similar depletion of GSH was seen in the presence of propargylglycine, an inhibitor of the cystathionine pathway, most of the cysteine required for the resynthesis of GSH must originate from methionine and not from cysteine generated by the catabolism of GSH. Further studies showed that GSH is sequentially oxidized by O2-. and HOCl, first to GSSG and then to an unidentified compound, most likely a chloramine. In the presence of an adequate supply of GSH and NADPH which is required for the reduction of GSSG by glutathione reductase this further oxidation of GSSG was prevented. Thus, the highly toxic HOCl generated by PMN can be detoxified by the glutathione reducatase system. The capacity of PMN to re-synthesize GSH may be an important determinant of PMN function.     

激活粒细胞对大鼠离体心脏再灌注损伤的实验研究

本实验采用大鼠离体心脏模型，探讨激活粒细胞对大鼠离体心脏再灌注损伤的作用机制。实验过程观察冠状动脉流量，冠状动脉阻力，冠状动脉流出液中乳酸脱氢酶的活性及再二醛的含量。实验完毕后对心肌进行组织学检查及超微结构观察。