Technical improvements in the clotted plasma droplet MIF assay in vitro.

The migration inhibitory activity of culture supernatants of rat and human lymphocytes stimulated with PHA and Con A-Sepharose was tested on cell migration from clotted plasma droplets. This technique was improved by using homologous as well as heterologous plasma and fibrinogen solutions for suspending the migratory cells, automatic micropipettes for performing the technique and purified cell populations as target. The effects of calcium chloride and of the cell concentration in the plasma droplets on the migration indices obtained in the MIF assay were tested.     

The apoptotic and autophagic properties of two natural occurring prodrugs,hyperoside and hypoxoside,against pancreatic cancer cell lines

Pancreatic cancer is only the 12th most common cancer, but the fourth leading cause of cancer-related deaths in the world. This is due to late prognosis, poor response to chemotherapy and early metastases. Natural prodrugs may play an important role in the treatment of pancreatic cancer. The main aim of this study was to determine the cytotoxicity of five natural prodrugs, namely harpagoside, hyperoside, hypoxoside, oleuropein and polydatin, by investigating apoptosis and autophagy as possible mechanism/s of action. Hypoxoside and hyperoside have shown selective cytotoxicity at IC50 values of ∼25 and 50 μM against INS-1 and MIA PaCa-2 pancreatic cancer cells, respectively. Hypoxoside and hyperoside induced G2/M phase arrest and caspase-3 activation in INS-1 and MIA PaCa-2 cells, respectively. Hoechst/phalloidin staining confirmed morphological changes, including condensed chromatin multinucleation, membrane blebbing and loss of cytoskeletal arrangement in INS-1 and MIA PaCa-2 cells. Acridine orange staining was absent in INS-1 (hypoxoside) and MIA PaCa-2 (hyperoside) treated cells, whereas LC3B expression was not significantly increased. INS-1 and MIA PaCa-2 treated cells favour the cell death pathway, apoptosis, over the cell survival pathway, autophagy.     

Proteinase-activated receptor-2-mediated signaling and inhibition of DNA synthesis in human pancreatic cancer cells

Summary Conclusion Proteinase-activated receptor-2 (PAR-2)-mediated effects contribute to the intracellular signaling network in pancreatic tumor cells. A role of PAR-2 as negative regulator in human pancreatic tumor growth might be implied. Background Using the human pancreatic tumor cell line MIA PaCa-2, we evaluated cellular effects of trypsin and the PAR-2-activating peptide SLIGRL on [Ca²⁺]_i mobilization, Ins(1,4,5)P₃ level, and protein kinase (PKC) activation. Furthermore, PAR-2 involvement in the regulation of cell proliferation has been estimated by measurement of [³H]thymidine incorporation in MIA PaCa-2 cells. Results Trypsin and the PAR-2 synthetic peptide agonist SLIGRL induced [Ca²⁺]_i mobilization, transient increase in inositol (1,4,5) triphosphate level, and PKC translocation in MIA PaCa-2 cells. In addition, SLIGRL induced a decrease in DNA synthesis in MIA PaCa-2 cells.     

Decreased calcitonin gene-related peptide expression in the dorsal root ganglia of TNF-deficient mice in a monoiodoacetate-induced knee osteoarthritis model

Background: The detailed mechanisms of knee osteoarthritis (OA) pain have not been clarified, but involvement of inflammatory cytokines such as tumor necrosis factor-alpha (TNF) has been suggested. The present study aimed to investigate the more detailed neurological involvement of TNF in joint pain using a TNF-knockout mouse OA model. Methods: The right knees of twelve-week-old C57BL/6J wild and TNF-deficient knockout (TNF-ko) mice (n=15, each group) were given a single intra-articular injection of 10 µg monoiodoacetate in 10 mL sterile saline. The left knees were only punctured as the control. Evaluations were performed immediately after the injection (baseline) and at 7, 14, and 28 days after the injection with a subsequent intra-articular injection of neurotracer into both knees. The animals were evaluated for immunofluorescence of the lumbar dorsal root ganglia (DRG) innervating the knee joints. The injected knees were observed macroscopically and mouse pain-related behaviors were scored. Results: Macroscopic observation showed similar knee OA development in both wild and TNF-ko mice. Calcitonin gene-related peptide (CGRP, a neuropeptide identified as a inflammatory pain-related biomarker) was significantly increased in DRG neurons innervating OA-induced knee joints with significantly less CGRP expression in TNF-ko animals. Pain-related behavior scoring showed a significant increase in pain in OA-induced joints, but there was no significant difference in pain observed between the wild and TNF-ko mice. Conclusions: The result of the present study indicates the possible association of TNF-alpha in OA pain but not OA development.     

Maternal immune activation leads to activated inflammatory macrophages in offspring

Several epidemiological studies have shown an association between infection or inflammation during pregnancy and increased risk of autism in the child. In addition, animal models have illustrated that maternal inflammation during gestation can cause autism-relevant behaviors in the offspring; so called maternal immune activation (MIA) models. More recently, permanent changes in T cell cytokine responses were reported in children with autism and in offspring of MIA mice; however, the cytokine responses of other immune cell populations have not been thoroughly investigated in these MIA models. Similar to changes in T cell function, we hypothesized that following MIA, offspring will have long-term changes in macrophage function. To test this theory, we utilized the poly (I:C) MIA mouse model in C57BL/6J mice and examined macrophage cytokine production in adult offspring. Pregnant dams were given either a single injection of 20 mg/kg polyinosinic–polycytidylic acid, poly (I:C), or saline delivered intraperitoneally on gestational day 12.5. When offspring of poly (I:C) treated dams reached 10 weeks of age, femurs were collected and bone marrow-derived macrophages were generated. Cytokine production was measured in bone marrow-derived macrophages incubated for 24 h in either growth media alone, LPS, IL-4/LPS, or IFN-γ/LPS. Following stimulation with LPS alone, or the combination of IFN-γ/LPS, macrophages from offspring of poly (I:C) treated dams produced higher levels of IL-12(p40) (p < 0.04) suggesting an increased M1 polarization. In addition, even without the presence of a polarizing cytokine or LPS stimulus, macrophages from offspring of poly (I:C) treated dams exhibited a higher production of CCL3 (p = 0.05). Moreover, CCL3 levels were further increased when stimulated with LPS, or polarized with either IL-4/LPS or IFN-γ/LPS (p < 0.05) suggesting a general increase in production of this chemokine. Collectively, these data suggest that MIA can produce lasting changes in macrophage function that are sustained into adulthood.     

愈合时间对微型种植体支抗稳定性的影响

目的：研究在相同力值牵引下,愈合时间对微型种植体支抗稳定性的影响。方法：在8只Beagle犬上颌根间牙槽骨中植入49颗微型种植体支抗,分别在即刻负载、愈合2周、愈合4周、愈合12周后施以150g水平牵引力,定期注射荧光标志物,负载2个月后处死动物。制作种植体-骨的不脱钙硬组织切片．进行荧光显微镜观察、组织形态学测量和推出试验。采用SAS6．12软件包中的单因素方差分析和SNK法分析各组间差异,相关性分析BIC、BSA值与生物力学性能参数的相关性。结果：4组标本的骨结合百分率（bone—to—implant contaet,BIC）和骨充填率（bone stlrface area,BSA）分别为（46．6±2．72）％、（44．08±3．62）％、（17．07±0．63）％、（32．85±3．01）％及（66．39±2．94）％、（64．96±5．11）％、（36．82±0．08）％、（70．33±4．07）％。愈合4周与其他3组之间均有显著性差异。4组标本种植体-骨界面的最大剪切力分别是（270．4±20．36）N、（270．4±6．97）N、（105.8±3．73）N、（335．07±17．22）N。愈合12周组与其他3组之间均有显著性差异。种植体-骨界面的总吸收能量情况与最大剪切力相似。微型种植体的生物力学性能与BSA的相关系数显著高于其与BIC的相关系数（心0．051。结论：愈合12周是一个较为理想的负载前愈合时间。即刻负载和愈合2周负载也可以接受,但必须注意种植体的即刻固位和即刻稳定性。而愈合4周负载则不利于种植体的稳定。     

193例特殊类型肺腺癌的临床特征及预后分析

  目的  分析“细支气管肺泡癌（bronchioloalveolar carcinoma BAC）及具有其特征的混合型腺癌”（2004年分型）的不利预后因素, 并经2011年肺腺癌新病理分型验证, 确定此类肿瘤的重要预后因素、阐明两种分型在评估临床预后的连贯性。  方法  回顾性分析天津医科大学附属肿瘤医院1974年3月至2007年5月收治并经病理学或细胞学确诊且随访资料完整的193例BAC及具其特征的混合型腺癌患者资料, 并将其中符合新分型的原位腺癌、微浸润癌与其他类型区别而单独分组, 记录性别、年龄、肿瘤大小、有无肿瘤家族史、吸烟史、TNM分期、临床症状持续时间及治疗方法等因素; 以Kaplan-Meier法计算生存率、Log-rank进行生存率显著性检验、Cox比例回归风险模型进行单因素、多因素分析, 评价各因素对预后的影响。  结果  全部患者的1年、3年、5年生存率分别为84.3%、60.6%、45.6%, 中位生存期为53个月。单因素分析显示: 确诊时临床分期（P < 0.001）、出现临床症状（P= 0.018）、原发病灶大小（P=0.039）、病理类型（P=0.028）及治疗方法选择（P=0.035）是影响预后的因素; Cox回归多因素分析显示: 仅确诊时临床分期是影响预后的独立因素; 其中符合新分型中的原位腺癌及微浸润癌患者预后最佳。  结论  BAC及具有其特征的混合型腺癌较其他非小细胞肺癌（NSCLC）预后好、临床表现无特异性, 确诊时的TNM分期是影响预后的独立因素; 在评估临床预后方面, 新肺腺癌分型与2004年分型具有连贯性, 且更细化、更利于指导临床。