Serum laminin assayed by Slot-Blot-ELISA in patients with combined viral hepatitis C and schistosomiasis

OBJECTIVES: Hepatic schistosomiasis and chronic hepatitis C virus (HCV) are the most prevalent agents causing hepatic fibrosis in humans. Laminin (LA) has been related to liver fibrosis and subsequent development of portal hypertension in chronic liver disease. There are no available data describing the pattern of laminin in combined HCV and schistosoma-infected patients, thus the rationale of this study was to assess the serum LA as an index of liver fibrosis in patients with schistosomiasis and/or chronic viral hepatitis C and to evaluate a developed Slot-Blot Enzyme-Linked Immunosorbant Assay (Slot-Blot-ELISA) as a method of estimation. DESIGNS AND METHODS: This study included four groups: group I included 34 patients with schistosomiasis, group II included 58 patients infected with HCV, group III included 68 patients with combined chronic viral hepatitis C and schistosomiasis and group IV included 50 healthy individuals who served as a control group. Serum LA was measured in the different groups quantitatively by ELISA and semi-quantitatively by Slot-Blot-ELISA. RESULTS: Significantly higher serum LA concentrations measured by ELISA were found in patients with combined chronic viral hepatitis C and schistosomiasis than in patients with either chronic HCV (P = 0.005) or schistosomiasis (P < 0.001) alone. Serum LA was significantly higher in the patient groups than the control group (P < 0.001). Serum LA concentration was positively correlated with fibrosis grading scores. Semi-quantitative results of serum LA using the developed SB-ELISA were found to have approximately the same power of ELISA results in different groups. The overall sensitivity, specificity, positive predictive value, negative predictive value and efficiency of ELISA for estimation of serum LA were 85.6%, 84.0%, 94.5%, 64.6% and 90%, respectively and for SB-ELISA were 87.5%, 82.0%, 94%, 67.2% and 88%, respectively. CONCLUSIONS: Serum LA was significantly increased in patients coinfected with HCV and Schistosoma mansoni. The newly developed Slot-Blot-ELISA is a simple, rapid and highly sensitive assay for detection of LA in hepatic fibrosis. Moreover, all steps were performed at room temperature without the need to use expensive equipment, and this may enhance the application of this assay in screening programs. Further study is warranted for confirmation of SB-ELISA reproducibility in a large population.     

Human papillomaviruses bind a basal extracellular matrix component secreted by keratinocytes which is distinct from a membrane-associated receptor

Human papillomaviruses (HPVs) have previously been shown to adsorb to cultured cells via membrane-associated heparan sulfate (HS) and alpha6 integrin. We demonstrate that cultured keratinocytes uniquely secrete a component into the basal extracellular matrix (ECM) which can function to adsorb HPV particles which can then be internalized by adherent cells. This uncharacterized basal ECM adsorption receptor was secreted by normal human epidermal keratinocytes (NHEK) and by each of the four keratinocyte-derived cell lines we examined, but not by non-keratinocyte cell lines. Multiple HPV types bound preferentially to this keratinocyte-specific receptor over the membrane-associated receptor, and binding to the basal ECM adsorption receptor was refractory to inhibition by heparin. Like the membrane-associated receptor, this basal ECM component was functional as an adsorption receptor in our in vitro infection model using HPV-11. Unlike particle adsorption, however, successful infection with HPV-11 virions remained sensitive to the pretreatment of virions with heparin. The secreted basal ECM receptor did not colocalize with antibodies against HS, perlecan, or alpha6 integrin, but colocalized with antibody against laminin-5, a marker of keratinocyte ECM and an abundant component of the basement membrane in mucosa and skin. These findings suggest a model for natural infections in which HPV virions, nonspecifically adsorbed to HS on suprabasal keratinocytes throughout an epithelial wound, might be transferred to mitotically active migrating keratinocytes via an intermediate association with the ECM secreted by these cells as they reestablish the basement membrane.     

VEGF和LN在大肠癌中的表达及与大肠癌预后的关系

目的：探讨大肠癌血管内皮生长因子(VEGF)、层黏连蛋白(LN)表达及与大肠癌预后的关系。方法：应用SP免疫组化法检测正常大肠黏膜、大肠腺瘤和大肠癌组织VEGF和LN的表达,并分析二者与大肠癌病理特征和预后的关系。结果：大肠癌VEGF和LN的表达均明显高于大肠腺瘤和正常大肠黏膜(P＜0.01,P＜0.05)。大肠癌VEGF和LN的表达均与淋巴结转移、分化程度、Duke’s分期和术后生存期有关(P＜0.01,P＜0.05)。结论：联合检测VEGF和LN可作为判断大肠癌浸润转移和评估预后的参考指标。     

Amino‐terminal fragments of laminin γ2 chain retract vascular endothelial cells and increase vascular permeability

Laminin γ2 (Lmγ2) chain, a subunit of laminin‐332, is a typical molecular marker of invading cancer cells, and its expression correlates with poor prognosis of cancer patients. It was previously found that forced expression of Lmγ2 in cancer cells promotes their invasive growth in nude mice. However, the mechanism of the tumor‐promoting activity of Lmγ2 remains unknown. Here we investigated the interaction between Lmγ2 and vascular endothelial cells. When treated with an N‐terminal proteolytic fragment of γ2 (γ2pf), HUVECs became markedly retracted or shrunken. The overexpression of Lmγ2 or treatment with γ2pf stimulated T‐24 bladder carcinoma cells to invade into the HUVEC monolayer and enhanced their transendothelial migration in vitro. Moreover, γ2pf increased endothelial permeability in vitro and in vivo. As the possible mechanisms, γ2pf activated ERK and p38 MAPK but inactivated Akt in HUVECs. Such effects of γ2pf led to prominent actin stress fiber formation in HUVECs, which was blocked by a ROCK inhibitor. In addition, γ2pf induced delocalization of VE‐cadherin and β‐catenin from the intercellular junction. As possible receptors, γ2pf interacted with heparan sulfate proteoglycans on the surface of HUVECs. Moreover, we localized the active site of γ2pf to the N‐terminal epidermal growth factor‐like repeat. These data suggest that the interaction between γ2pf and heparan sulfate proteoglycans induces cytoskeletal changes of endothelial cells, leading to the loss of endothelial barrier function and the enhanced transendothelial migration of cancer cells. These activities of Lmγ2 seem to support the aberrant growth of cancer cells.     

肾衰宁对糖尿病大鼠肾脏损伤的保护作用

目的 探讨肾衰宁对糖尿病大鼠肾脏的保护作用及可能机制.方法 雄性Vista大鼠24只,随机选取8只作为正常对照组(N组),其余16只注射链脲佐茵素(STZ)制成糖尿病模型,并随机分为糖尿病肾病组(DN组,n=8)和肾衰宁治疗组(S组,n=8),给药8周.检测血糖、肾功能、24 h尿蛋白定量等指标,并进行肾脏病理学检查及...     

Fractone-associated N-sulfated heparan sulfate shows reduced quantity in BTBR T+tf/J mice: a strong model of autism

BTBR T+tf/J (BTBR) mice show abnormal social, communicatory, and repetitive/stereotyped behaviors paralleling many of the symptoms of autism spectrum disorders. BTBR also show agenesis of the corpus callosum (CC) suggesting major perturbations of growth or guidance factors in the dorsal forebrain [1]. Heparan sulfate (HS) is a polysaccaride found in the brain and other animal tissues. It binds to a wide variety of ligands and through these ligands modulates a number of biological processes, including cell proliferation and differentiation, migration and guidance. It is aggregated on fractal-like structures (fractones) in the subventricular zone (SVZ), that may be visualized by laminin immunoreactivity (LAM-ir), as well as by HS immunoreactivity (HS-ir). We report that the lateral ventricles of BTBR mice were drastically reduced in area compared to C57BL/6J (B6) mice while the BTBR SVZ was significantly shorter than that of B6. In addition to much smaller fractones for BTBR, both HS and LAM-ir associated with fractones were significantly reduced in BTBR, and their anterior-posterior distributions were also altered. Finally, the ratio of HS to LAM in individual fractones was significantly higher in BTBR than in B6 mice. These data, in agreement with other findings linking HS to callosal development, suggest that variations in the quantity and distribution of HS in the SVZ of the lateral ventricles may be important modulators of the brain structural abnormalities of BTBR mice, and, potentially, contribute to the behavioral pathologies of these animals.     

抑制层黏素受体过表达在胆管癌细胞免疫逃逸中的作用

目的 探讨抑制层黏素受体(LNR)过表达下调Fas配体基因表达在胆管癌免疫逃逸中的作用.方法 采用流式细胞仪分选出过表达层黏素受体的胆管癌细胞QBC939,用脂质体转染法转染层黏素受体RNA干扰质粒和阴性对照质粒各6 μg至过表达层黏素受体的胆管癌细胞QBC939中,用实时荧光定量PCR法检测未转染组、阴性组和阳性组QBC939细胞Fas配体基因的表达情况.结果 分选后的QBC939细胞层黏素受体的表达率高达95%,继续转染层黏素受体RNA干扰质粒后,其FasL基因表达水平较未转染组和转染阴性对照质粒组明显降低.结论 LNR过表达在胆管癌免疫逃逸中的作用可能是通过Fas-FasL信号通路实现的.     

Effect of laminin environments and tumor factors on the biology of myeloid dendritic cells

Dendritic cells (DCs) are immune cells that surveil the organism for infections or malignancies and activate specific T lymphocytes initiating specific immune responses. Contrariwise, DCs have been show to participate in the development of diseases, among them some types of cancer by inducing angiogenesis or immunosuppression. The ultimate fate of DC functions regarding their role in disease or health is prompted by signals from the microenvironment. We have previously shown that the interaction of DCs with various extracellular matrix components modifies the immune properties and angiogenic potential of these cells.The objective of the current studies was to investigate the angiogenic and immune profile of murine myeloid DCs upon interaction with laminin environments, with a particular emphasis on ovarian cancer.Our results show that murine ovarian tumors produce several types of laminins, as determined by PCR analysis, and also that tumor-associated DCs, both from ascites or solid tumors express adhesion molecules capable of interacting with these molecules as determined by flow cytometry and PCR analysis. Further, we established that DCs cultured on laminin upregulate both AKT and MEK signaling pathways, and that long-term culture on laminin surfaces decreases the immunological capacities of these cells when compared to the same cells cultured on synthetic substrates. In addition, we observed that tumor conditioned media was able to modify the metabolic status of these cells, and also reprogram the development of DCs from bone marrow precursors towards the generation of myeloid-derived suppressor cells.Overall, these studies demonstrate that the interaction between soluble factors and extracellular matrix components of the ovarian cancer microenvironment shape the biology of DCs and thus help them become co-conspirators of tumor growth.     

Biofunctionalized peptide-based hydrogel as an injectable scaffold for BDNF delivery can improve regeneration after spinal cord injury

Background

The complex pathophysiological events occurring after traumatic spinal cord injuries (TSCI) make this devastating trauma still incurable. Peptide amphiphile (PA) hydrogels are nanobiomaterials displaying desirable properties for application in regenerative medicine because they are absorbable, injectable, allowing biofunctionalization, controlling release of trophic factors and mimic extracellular matrix (ECM). In this study, we explored the potentiality of the IKVAV-functionalized PA hydrogel to provide a permissive environment for cell migration and growth as well as sustained release of BDNF at the lesion after severe compression injury model.

肺腺癌中LN 5蛋白表达及其与EGFR突变关系的临床研究

目的 探讨肺腺癌组织中层粘连蛋白 5（LN-5）表达及其与人表皮生长因子受体（EGFR）基因突变关系。方法 收集2015年1月至2015年12月57例肺腺癌及其配对癌旁组织标本,采用免疫组化SP法检测LN 5蛋白表达,同时通过液相芯片技术检测EGFR基因突变状态,分析LN 5表达与肺腺癌临床病理特征的关系以及与EGFR基因突变的相关性。 结果肺腺癌组织中LN 5蛋白阳性表达率为21.1％（12／57）,癌旁组织中为1.7％（1／57）,差异有统计学意义（P＜0.05）。EGFR敏感突变（E19+E21）的发生率为43.9％（25／57）,EGFR非敏感突变（野生型+E20）的发生率为56.1％（32／57）。LN-5蛋白表达与淋巴结转移和EGFR突变有关（P＜0.05）,与TNM分期无关（P＞0.05）。LN 5表达与EGFR突变呈负相关（r=-0.283,P＜0.05）。结论 LN 5在肺腺癌组织中的表达升高,且其表达与EGFR突变状态具有相关性。