Megakaryocyte interaction with subendothelial extracellular matrix is associated with adhesion, platelet-like shape change, and thromboxane A2 production

We have examined the morphological and secretory behavior of rat and guinea pig megakaryocytes exposed for up to 24 hours to extracellular matrix produced by cultured bovine endothelial cells. By phase-contrast microscopy of living cells and in more detail by scanning electron microscopy, the megakaryocytes showed a nonreversible adherence, an extensive formation of filopodia around the periphery like the rays of the sun, and a tendency toward flattening. These filopodia were generally linear with attenuated tips and were larger than, but resembled the filopodia of, rat or guinea pig platelets exposed to this extracellular matrix. In contrast, isolated megakaryocytes on glass or on uncoated plastic surfaces did not show these responses; adherence, in the face of gentle agitation before fixation, was minimal, with rare filopodia and no flattening. Megakaryocytes that interacted with the extracellular matrix produced significant amounts of thromboxane A2, but this did not occur on uncoated surfaces and could not be attributed to other contaminating cells in the megakaryocyte suspensions. The appearance in megakaryocytes of these typical platelet responses indicates that megakaryocytes acquire the functional capabilities of platelets by the synthesis and assembly of platelet substances and organelles. Thromboxane production by megakaryocytes stimulated by the extracellular matrix is a readily quantifiable measure of this capacity.     

Effects of P-MAPA Immunomodulator on Toll-Like Receptors and p53: Potential Therapeutic Strategies for Infectious Diseases and Cancer

Background

Compounds that can act as agonists for toll-like receptors (TLRs) may be promising candidates for the development of drugs against infectious diseases and cancer. The present study aimed to characterize the immunomodulatory effects of P-MAPA on TLRs in vitro and in vivo, as well as to investigate its potential as adjuvant therapy in infectious diseases and cancer.

Methods

For these purposes, the activity of P-MAPA on TLRs was assayed in vitro through NF-??B activation in HEK293 cells expressing a given TLR, and using an in vivo animal model for bladder cancer (BC). The antimicrobial activity of P-MAPA was tested against Mycobacterium tuberculosis (TB) in vitro in an MIC assay, and in vivo using an aerosol infection model of murine tuberculosis. Antitumor effects of P-MAPA were tested in an animal model with experimentally induced BC. Moxifloxacin (MXF) and Bacillus Calmette-Guerin (BCG) were used as positive controls in the animal models.

Results

The results showed that P-MAPA, administered alone or in combination with MXF, induced significant responses in vivo against TB. In contrast, the compound did not show antimicrobial activity in vitro. P-MAPA showed a significant stimulatory effect on human TLR2 and TLR4 in vitro. In BC, TLR2, TLR4 and p53 protein levels were significantly higher in the P-MAPA group than in the BCG group. The most common histopathological changes in each group were papillary carcinoma in BC group, low-grade intraepithelial neoplasia in BCG group and simple hyperplasia in P-MAPA group. Concerning the toxicological analysis performed during BC treatment, P-MAPA did not show evidence for hepatotoxicity and nephrotoxicity.

Conclusions

In conclusion, P-MAPA acted as TLR ligand in vitro and improved the immunological status in BC, increasing TLR2 and TLR4 protein levels. P-MAPA immunotherapy was more effective in restoring p53 and TLRs reactivities and showed significantly greater antitumor activity than BCG. The activation of TLRs and p53 may provide a hypothetical mechanism for the therapeutic effects in both cancer and infectious diseases. Taken together data obtained will encourage the further investigation of P-MAPA as a potential candidate for the treatment of cancer and infectious diseases.     

The Prevalence of Cryptosporidium spp. in Children,Taiz District,Yemen

Background

This is the first work done on cryptosporidiosis among the children in Taiz, Yemen.

Methods

A number of 712 samples were collected from children of different ages (ranging from 1 month to 12 years) from Dec 2006 to Aug 2007. The collected samples were examined by Sheather''s sugar floatation and Modified Ziehl- Neelsen stain as well as ELISA methods. The test results were statistically analyzed by SPSS software.

Results

The overall positive percentage was 43.7%. The higher incidence (36.2%) was occurred in males while the lowest incidence (32.7%) was observed in females (r=0.876; P=0.001). The correlation between infected cases and the type of drinking water was r =0.121. Among the cases examined by ELISA (92 cases), 26.1% were infected. The correlation between seropositivity and gender was r=0.652 (P=0.031).

Conclusion

Cryptosporidium spp. is a significant pathogen among children at Taiz. Fresh water supplies, education, eating habits and domestic animals are considered the main sources for transmission of cryptosporidiosis.     

Platelet activation status decreases after menopause

Objective.?The aim of the study was to investigate the impact of the climacterium (before and after menopause) on platelet activation.

Background.?Platelet activation has been associated to the risk of cardiovascular disease. There is much speculation about the relationship between platelet function and sex steroids, due to peculiarities of platelet action between the genders, including concerns about the influence of low estradiol status in menopausal women.

Methods.?By means of a cross-sectional study design, 37 female patients divided into two groups were compared. Group A consisted of ten women, mean age 43.9 years, in the premenopausal period, with normal estrogen levels; and Group B comprised 27 patients, mean age 53.0 years, who had all reached menopause. Platelet activation markers, namely P-selectin and glycoprotein IIb–IIIa complex (GPIIb–IIIa), were evaluated by flow cytometry with monoclonal antibodies. A binding index was calculated for both parameters (percentage of positive platelets?×?mean fluorescence of positive platelets). Also, thromboxane A₂ was quantified by means of its main plasma metabolite, thromboxane B₂, by enzyme immunoassay.

Results.?P-selectin and GPIIb–IIIa expression results revealed lower platelet activation status after menopause, as there was a decrease in both the percentage of P-selectin^?+? platelets and of GPIIb–IIIa mean fluorescence of positive platelets, lowering both binding indices. P-selectin binding index differed significantly between Group A (12.3?±?3, n?=?10) and Group B (6.2?±?2.9, n?=?27; mean?±?standard deviation (SD), p?<?0.001). GPIIb–IIIa binding index also differed significantly between both groups (Group A: 18.8?±?2.3, n?=?10 vs. Group B: 16.2?±?3.1, n?=?27; mean?±?SD, p?<?0.0018). Plasma concentration of thromboxane B₂ was 1.07?±?0.5?pg/well before menopause (Group A, n?=?10) and 1.9?±?4.1?pg/well after menopause (Group B, n?=?27), not significantly different (mean?±?SD, baseline?×?therapy, p?=?0.85).

Conclusions.?After the menopause, climacteric women – whose estradiol status is low – have a decreased activation platelet status compared with premenopausal women. Nevertheless, further studies on a larger sample are necessary for conclusive data regarding cardiovascular disease.     

超低出生体重儿红细胞输注的临床分析

探讨超低出生体重(ELBW)儿减少红细胞输注的可能性。方法对1989～1997年9年间256 例超低出生体重儿的红细胞输注进行临床分析。在此期间红细胞输注指征进行了3次制订,对检验样本的采 血量进行了严格的控制,部分病例应用了重组人促红细胞生成素治疗。结果1994年以后有1/4的ELBW儿 不需要输注红细胞,1989～1997年平均输血次数由7次降至2.7次(P＜0.01)。按出生体重累计红细胞输注 量由163.5mL/kg降至69.2mL/kg(P＜0.01),接受供血者人数由6.3人降至1.5人(P＜0.01)。红细胞输注 前的平均红细胞压积比,机械通气者由0.43降至0.34、自主呼吸者由0.41降至0.31。ELBW儿更加不成熟,平 均胎龄由27.4周减至26.0周,平均出生体重由833g降至741g。存活率仍达78％,住院时间没有延长,严重并 发症如视网膜病、动脉导管未闭、脑室出血的发生率没有增加。结论制订严格的红细胞输注指征有助于减少 输血次数和接受供血者人数,且患儿临床耐受良好。     

脑康泰胶囊对阿尔茨海默氏病大鼠学习记忆作用的影响

目的：观察脑康泰胶囊对阿尔茨海默氏病模型大鼠学习记忆作用的影响。方法：采用脑立体定向颅内注射啉酸所致阿尔第默氏病（Alzheimers disease.AD)大鼠模型的方法。结果：脑康泰胶囊可显著增加强AD大鼠被动学习和主动学习的能力。调节脑组织中单胺类递质含量及血清中相关激素水平,并显著改善AD模型大鼠的脑电图。结论：脑康泰胶囊显著改善AD大鼠学习记忆能力的作用,其机制与其调节中枢递质及激素水平等相关。     

一种新的层析等电点聚焦技术提纯葡萄球菌肠毒素D

我们成功地建立了一个新的层析等电点聚焦的三步程序.细菌培养液首先以CG-50吸附,洗脱出的粗毒素溶液在PBE_(94)柱上进行层析等电点聚焦.分别用5mMTris-Base pH9.4和Buffalyte_(8-4) pH5.4缓冲液洗脱,出现多个洗脱峰,即刻测定pH值.收集pH6.8～7.1的洗脱液,混合、浓缩,最后通过Sephacryl s-200过滤,以0.05M PBS pH6.8洗脱,出现3个峰,毒素在第1峰.毒素纯度为98%,回收率89%,p17.6,MW28500,SDS-PAGE上1条带.以~(125)I标记SED作探针的电泳转移和放射自显影均证明了提纯SED的免疫学特异性.免疫扩散试验,即使200μg/ml SED与不产毒素的FRI-184抗血清,也未有可见沉淀线.