自体骨髓间充质干细胞移植后心肌缝隙连接蛋白43表达与室性心动过速发生的关系

目的:缝隙连接蛋白43对维持心肌细胞的连接通讯功能、电信号传导和正常的节律性收缩起重要作用,其表达和分布的异常是多种室性心律失常的解剖学基础,建立小型猪急性心肌梗死模型.观察自体骨髓间充质干细胞移植后室性心动过速的发生及心肌缝隙连接蛋白43的表达方法:实验于2006-01/2007-01在河北省人民医院导管室完成。①材料:选取8～12月龄小型猪22头,由河北医科大学实验动物中心提供,体质量20～30 kg,随机数字表法分为细胞移植组12头、模型对照组10头.实验过程中对动物的处置符合动物伦理学际准②实验方法:无菌条件下抽取猪双侧股骨骨髓20 mL,percoll法 贴壁法分离培养骨髓间充质干细胞,待细胞生长达75%融合时用胰酶消化传代。将传至第2代细胞加入终浓度为10μmol/L的5-氮胞苷进行诱导,用胶体金标记12 h后继续培养20 d用于移植:两组小型猪均采用球囊堵闭法建立急性心肌梗死模型.心电图监测示相关至少2个导联ST段抬高大于0.2 mV、术后血肌钙蛋白和肌酸磷酸激酶同工酶升高超过正常的两倍为建模成功标准:细胞移植组于造模成功后经OTW球囊于第一对角支远端1 cm处再次阻断血流,注入经胶体金标记的10×10~7个骨髓间充质干细胞。③实验评估:于细胞移植后2 h及4周行电生理程序刺激.观察室性心动过速的发生情况。末次电生理检查后、采用免疫组化染色法检测心肌缝隙连接蛋白43的表达.计算其积分吸光度值。结果:①模型建立指标检测:与术前比较.造模后所有小型猪血肌钙蛋白含量和肌酸磷酸激酶同工酶活性均增高,峰值浓度分别为(21.3±3.6)μg/L和(178.3×41.4)IU/L,术中心电图ST段平均抬高(10.67±1.43)mm.证明急性心肌梗死模型成功建立。②骨髓间充质干细胞移植后室性心动过速的发生情况:与模型对照组诱发出室性心动过速的动物数量比较,术后2 h细胞移植组无明显变化(X~2=0.201,P=0.650),术后4周细胞移植组明显降低(X~2=4.455.P=0.035)。②骨髓间充质干细胞移植后梗死心肌缝隙连接蛋白43的表达:术后4周移植到梗死心肌的骨髓间充质干细胞与宿主心肌生长为一体,移植部位颜色变黑,苏木精-伊红染色示移植细胞的胞浆呈紫红色。细胞移植组心肌梗死区缝隙连接蛋白43积分吸光度值明显高于模型对照组(t=16,82.P=0.00),细胞移植组中未发生室性心动过速小型猪的梗死心肌缝隙连接蛋白43的表达明显高于发生室性心动过速小型猪(t=5.06,P=0.00)。结论:自体骨髓间充质干细胞移植可促进急性心肌梗死猪心肌缝隙连接蛋白43的表达,其表达程度可能与急性心肌梗死室性心动过速的发生有关。     

Insights into the initiation of type 2 immune responses

Type 2 immune responses, characterized by the differentiation of CD4+ T helper type 2 (Th2) cells and the production of the type 2 cytokines interleukin-4 (IL-4), IL-5, IL-9 and IL-13, are associated with parasitic helminth infections and inflammatory conditions such as asthma and allergies. Until recently the initiating factors associated with type 2 responses had been poorly understood. This review addresses the recent advances in identifying the diverse range of antigens/allergens associated with type 2 responses and the function, expression and sources of type-2-initiating cytokines (thymic stromal lymphopoietin, IL-25 and IL-33). We also discuss the latest findings regarding innate lymphoid cells, such as nuocytes, as early sources of type 2 cytokines and their importance in protective immunity to helminth infections. These developments represent major breakthroughs in our understanding of type 2 immunity, and highlight the increased complexity existing between the innate and adaptive arms of these responses. These additional steps in the type 2 immune pathway also offer potential targets for therapeutic intervention.     

Routine chest X-ray is not required after a low-risk central venous cannulation

Background: Knowledge of the radiographic catheter tip position after central venous cannulation is normally not required for short-term catheter use. Detection of a possible iatrogenic pneumothorax may nevertheless justify routine post-procedure chest X-ray. Our aim was to design a clinical decision rule to select patients for radiographic evaluation after central venous cannulation.
Methods: A total of 2230 catheterizations performed using external jugular, internal jugular or subclavian venous approaches during a 4-year period were included consecutively. Information on patient data and corresponding procedures was recorded prospectively. A post-procedure chest X-ray was obtained after each cannulation.
Results: Thirteen cases (0.58%) of cannulation-associated pneumothorax were identified. The risk of pneumothorax after a technically difficult (1.8%) or subclavian (1.6%) cannulation was significantly higher than after cannulation not considered as difficult (0.37%) or performed using other routes (0.33%). Clinical signs of pneumothorax within 8 h of cannulation were found in all seven patients with pneumothorax requiring specific treatment. A new clinical decision rule for radiographic evaluation after central venous cannulation based on the results of the present study shows that 48% of the post-procedure chest X-rays performed in our patients were clinically redundant.
Conclusion: Clinical symptoms were reported in all patients with pneumothorax requiring specific treatment. Approximately half of the post-procedure chest X-ray controls could be avoided using the proposed clinical decision rule to select patients for radiographic evaluation after central venous cannulation. A large prospective multi-centre study should be carried out to further evaluate this decision rule.     

The Development of a Novel Cancer Immunotherapeutic Platform Using Tumor-targeting Mesenchymal Stem Cells and a Protein Vaccine

An ideal anticancer strategy should target only the malignant cells but spare the normal ones. In this regard, we established a platform, consisting of an antigen-delivering vehicle and a protein vaccine, for developing an immunotherapeutic approach with the potential for eliminating various cancer types. Mesenchymal stem cells (MSCs) have been demonstrated capable of targeting tumors and integrating into the stroma. Moreover, we have developed a protein vaccine PE(ΔIII)-E7-KDEL3 which specifically recognized E7 antigen and elicited immunity against cervical cancer. Taking advantage of tumor-homing property of MSCs and PE(ΔIII)-E7-KDEL3, we used E6/E7-immortalized human MSCs (KP-hMSCs) as an E7 antigen-delivering vehicle to test if this protein vaccine could effectively eliminate non-E7-expressing tumor cells. Animals which received combined treatment of KP-hMSCs and PE(ΔIII)-E7-KDEL3 demonstrated a significant inhibition of tumor growth and lung-metastasis when compared to PE(ΔIII)-E7-KDEL3 only and KP-hMSCs only groups. The efficiency of tumor suppression correlated positively to the specific immune response induced by PE(ΔIII)-E7-KDEL3. In addition, this combined treatment inhibited tumor growth via inducing apoptosis. Our findings indicated that KP-hMSCs could be used as a tumor-targeting device and mediate antitumor effect of PE(ΔIII)-E7-KDEL3. We believe this strategy could serve as a platform for developing a universal vaccine for different cancer types.     

HUMAN PANCREATIC GROWTH HORMONE RELEASING HORMONE FAILS TO STIMULATE HUMAN GROWTH HORMONE BOTH IN CUSHING''S DISEASE AND IN CUSHING''S SYNDROME DUE TO ADRENOCORTICAL ADENOMA

An absent or severely blunted hGH response to an i.v. bolus injection (100 micrograms) of human pancreatic growth hormone releasing hormone (hpGRF 1-44) was found in seven female patients with Cushing's syndrome (five with pituitary dependent Cushing's disease and two due to an adrenal adenoma) and four men with pituitary dependent Cushing's disease. Three of the female and three of the male patients had an adequate hypoglycaemia after insulin administration. All these patients showed an absent or blunted hGH response after insulin induced hypoglycaemia. The GHRH data in these patients are in agreement with those in older literature on hGH responsiveness to stimuli such as L-dopa, arginine and insulin induced hypoglycaemia. It is concluded that hypercortisolism inhibits hGH release to various stimuli at the pituitary level.     

Secretion of Tumour Necrosis Factor α and Lymphotoxin α in Relation to Polymorphisms in the TNF Genes and HLA-DR Alleles. Relevance for Inflammatory Bowel Disease

The genes for tumour necrosis factor alpha (TNFα) and lymphotoxin alpha (LTα; TNFβ) are tandemly arranged in the central region of the MHC. They may, therefore, be of importance for the aetiology of MHC-associated diseases. The authors have prospectively studied the secretion of TNFα and LTα in relation to polymorphisms at positions -308 and -238 in the TNFα gene (TNFA), and two polymorphisms in the first intron of the LTα gene (LTA), as well as HLA-DR in 30 patients with chronic inflammatory bowel diseases (IBD) and 12 healthy controls. In the Dutch population, the alleles of these four polymorphisms are present in only five combinations, called TNF-haplotypes: TNF-C, -E, -H, -I, and -P. Significant associations between TNF haplotypes and TNFα and LTα secretion were found when PBMC were cultured with T-cell activators, irrespective of disease. Mean TNFα secretion of individuals carrying the HLA-DR3 associated TNF-E haplotype was significantly higher, as compared to individuals without this haplotype (26 441 pg/ml versus 19 629 pg/ml; P = 0.014). Individuals carrying the TNF-C haplotype produced the lowest amount of TNFα (17 408 pg/ml; P = 0.022). The TNF-C and TNF-E haplotypes differ only at position -308 in the promoter of TNFA. Individuals carrying the HLA-DR1 associated TNF-I haplotype produced significantly less LTα when compared to those who lack this haplotype (1979 pg/ml versus 3462 pg/ml; P = 0.006). As the TNF-I haplotype is also associated with low TNFα secretion, this haplotype thus defines a ‘low secretor phenotype’. In conclusion, this is the first study to show associations between TNF haplotypes and TNFα and LTα secretion when T-cell stimulators are used. These findings will contribute to define disease heterogeneity in IBD and may be of relevance for understanding the pathogenesis of autoimmune diseases.     

Capsaicin-sensitive extrinsic afferents are involved in acid-induced activation of distinct myenteric neurons in the rat stomach

Challenge of the rat gastric mucosa with 0.5 mol L(-1) HCl activates nitrergic neurons in the myenteric plexus as visualized by c-Fos immunohistochemistry. In the present study, we characterized the activated neurons more extensively by their chemical coding and investigated whether a neural pathway that involves capsaicin-sensitive extrinsic afferents and/or cholinergic neurons transmitting via nicotinic receptors contributes to the activation of myenteric neurons. In multiple labelling experiments, c-Fos was examined for co-localization with nitric oxide synthase (NOS), vasoactive intestinal peptide (VIP), neuropeptide Y (NPY), enkephalin (ENK), gastrin-releasing peptide (GRP), substance P (SP), calbindin D-28k (CALB) and neurofilament 145 (NF 145). All c-Fos-positive neurons were immunoreactive for NOS, VIP, NPY and NF 145, but not for SP, ENK, GRP and CALB. Nerve fibres co-expressing NOS, VIP and NPY were predominantly found in the external muscle layer and in the muscularis mucosae but rarely in the mucosa. Pre-treatment with capsaicin or hexamethonium or a combination of both pre-treatments reduced HCl-induced c-Fos expression by 54, 66 and 63%, respectively. Acid challenge of the stomach, therefore, leads to activation of presumably inhibitory motor neurons responsible for muscle relaxation. Activation of these neurons is partly mediated by capsaicin-sensitive afferents and involves ganglionic transmission via nicotinic receptors.     

Effect of nasal anaesthesia on lacrimal function after nasal allergen challenge

Ocular symptoms in allergic rhinoconjunctivitis arise partly from direct contact of the allergen with the conjunctiva and presumably partly from a naso-lacrimal reflex. The aim was to study the importance of this reflex in allergic rhinitis after topical anaesthesia of the nose. Ten patients with allergic rhinoconjunctivitis lo birch pollen were challenged with increasing intranasal doses of allergen until allergic symptoms occurred. The same dose of allergen was used in two other provocations, when the nasal cavity was anaesthetized with topical lidocaine in a double-blind rundomized cross-over manner. The effect on the eyes was evaluated by Shirmer's test, a routine method for measuring tear production. Lidocaine per se and/or allergen challenge had no significant effect on tear production. The naso-lacrimal reflex was not involved in the eye symptoms, in allergic conjunctivitis.