Differentially expressed proteins in the precancerous stage of rat hepatocarcinogenesis induced by diethylnitrosamine / 中华肝脏病杂志

<p><b>OBJECTIVE</b>To screen the differentially expressed proteins especially at the precancerous stage of diethylnitrosamine (DEN) induced hepatocarcinogenesis by comparative proteome research.</p><p><b>METHODS</b>Rats were divided into normal and DEN groups and sacrificed periodically. The liver samples were stained with gamma-glutamyl transpeptidase (GGT) and HE to distinguish the preneoplastic lesion (pre-HCC) from the normal and HCC tissues. The two-dimensional electrophoresis (2-DE) and mass spectrometry (MALDI-TOF-MS/MS) were then applied to analyze the differentially expressed protein between pre-HCC and normal tissues, pre-HCC and HCC, as well as HCC and normal tissues. A few of the candidate proteins such as laminin receptor 1 (67LR) and agmatinase were validated by Western blot and RT-PCR.</p><p><b>RESULTS</b>Totally, there were 82 proteins that differentially expressed two fold or more in one kind of tissues sample than the other, 47 of which occurred in the pre-HCC tissues. Eight proteins including 67LR were consistently up-regulated from normal tissue to pre-HCC and then to HCC tissues, while 22 proteins including agmatinase showed progressively down-regulated in these tissues samples.</p><p><b>CONCLUSION</b>The protein expression profiles are different during the process of hepatocarcinogenesis. Further study on the differentially expressed protein, especially these upregulated in the precancerous stage such as 67LR and agmatinase, might contribute to prevention and early diagnosis of human HCC.</p>

(99m)Tc-YIGSR as a receptor tracer in imaging the Ehrlich ascites tumor-bearing mice as compared with (99m)Tc-MIBI / 华中科技大学学报（医学）（英德文版）

The validity of (99m)Tc-YIGSR, a novel receptor radio-tracer, in imaging the Ehrlich ascites tumor was evaluated. YIGSR, a pentapeptide of laminin, was labeled with (99m)Tc by using a bifunctional chelator S-Acetly-NH(3)-MAG(3). The MIBI was labeled with (99m)Tc by following the kit instruction. The mice of tumor group were intravenously injected 1-2 mCi of (99m)Tc-YIGSR or (99m)Tc-MIBI via caudal vein, immobilized and imaged under a Gamma camera. The same procedure was performed in mice of blockade group, in which the unlabeled YIGSR was previously injected to block the receptor-recognition sites, and inflammation group serving as control. The reverse-phase Sep-Pak C(18) chromatogram was found to have an essentially complete conjugation between YIGSR and S-Acetly-NH(3)-MAG(3). The conjugated YIGSR could be radio-labeled successfully with (99m)Tc at room temperature and neutral pH, with a radio-labeling yield of 62%. Without the chelator S-Acetly-NH(3)-MAG(3), the YIGSR was labeled with (99m)Tc at an efficiency of 4%. The imagological study revealed obvious tumor accumulation of (99m)Tc-YIGSR 15 min after the injection, and the uptake peaked after 3 h with a tumor-to-muscle ratio (T/M) of 11.36. The radio-tracer was slowly cleared up and resulted in a T/M of 3.01 at the 8th h after the injection. As for blocked group, the tumor uptake of radiotracer was significantly lower, with the highest T/M being 4.61 after 3 h and 0.89 after 8 h. The T/M was 3.72 at the 3rd h and 1.29 at the 8th h after the (99m)Tc-YIGSR injection in the inflammatory group. The T/M was significantly higher in tumor group than in inflammatory group or control group (P<0.001). In the 99mTc-MIBI group, the T/M was 1.40 at the 3rd h and 0.55 at the 8th h after the injection, which showed a significant difference as compared with (99m)Tc-YIGSR (P<0.001). It is concluded that YIGSR can be successfully radiolabelled by using S-Acetly-NH(3)-MAG(3). (99m)Tc-YIGSR has many advantages in tumor imaging, such as quick and clear visualization, high sensitivity and specificity, and satisfactory target/non-target ratio (N/NT). It promises to be tumor radio-tracer.

Expression and activity analysis of human 67kD laminin receptor in Pichia pastoris / 生物工程学报

To carry out the secretive expression of human 67 kD laminin receptor (67LR), recombinant expression plasmid pPIC9K-67LR was constructed by inserting of 67LR cDNA into yeast expression vector pPIC9K. The 67LR protein was expressed in Pichia pastoris after induced by methanol, and about 12.56 mg electrophoresis purity 67LR could be obtained after the purification of 1L culture using affinity chromatograph column. In vitro competitive binding assay showed that target protein has an excellent biological activity. The successful expression of 67LR has placed a solid foundation for the research on structure and functions of 67LR.

(99m)Tc-YIGSR as a receptor tracer in imaging the Ehrlich ascites tumor-bearing mice as compared with (99m)Tc-MIBI / 华中科技大学学报（医学）（英德文版）

The validity of (99m)Tc-YIGSR, a novel receptor radio-tracer, in imaging the Ehrlich ascites tumor was evaluated. YIGSR, a pentapeptide of laminin, was labeled with (99m)Tc by using a bifunctional chelator S-Acetly-NH(3)-MAG(3). The MIBI was labeled with (99m)Tc by following the kit instruction. The mice of tumor group were intravenously injected 1-2 mCi of (99m)Tc-YIGSR or (99m)Tc-MIBI via caudal vein, immobilized and imaged under a Gamma camera. The same procedure was performed in mice of blockade group, in which the unlabeled YIGSR was previously injected to block the receptor-recognition sites, and inflammation group serving as control. The reverse-phase Sep-Pak C(18) chromatogram was found to have an essentially complete conjugation between YIGSR and S-Acetly-NH(3)-MAG(3). The conjugated YIGSR could be radio-labeled successfully with (99m)Tc at room temperature and neutral pH, with a radio-labeling yield of 62%. Without the chelator S-Acetly-NH(3)-MAG(3), the YIGSR was labeled with (99m)Tc at an efficiency of 4%. The imagological study revealed obvious tumor accumulation of (99m)Tc-YIGSR 15 min after the injection, and the uptake peaked after 3 h with a tumor-to-muscle ratio (T/M) of 11.36. The radio-tracer was slowly cleared up and resulted in a T/M of 3.01 at the 8th h after the injection. As for blocked group, the tumor uptake of radiotracer was significantly lower, with the highest T/M being 4.61 after 3 h and 0.89 after 8 h. The T/M was 3.72 at the 3rd h and 1.29 at the 8th h after the (99m)Tc-YIGSR injection in the inflammatory group. The T/M was significantly higher in tumor group than in inflammatory group or control group (P<0.001). In the 99mTc-MIBI group, the T/M was 1.40 at the 3rd h and 0.55 at the 8th h after the injection, which showed a significant difference as compared with (99m)Tc-YIGSR (P<0.001). It is concluded that YIGSR can be successfully radiolabelled by using S-Acetly-NH(3)-MAG(3). (99m)Tc-YIGSR has many advantages in tumor imaging, such as quick and clear visualization, high sensitivity and specificity, and satisfactory target/non-target ratio (N/NT). It promises to be tumor radio-tracer.

Expression and function of laminin receptor in laryngeal squamous cell carcinoma / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To investigate the role of 67 000 laminin receptor (LN-R) in the processes of invasion and metastasis of laryngeal squamous cell carcinoma.</p><p><b>METHODS</b>Expression of 67 000 LN-R mRNA in 20 cases with laryngeal squamous cell carcinomas and corresponding normal tissues was determined by RT-PCR, and the relationship between its expression level and patients' clinicopathological features was analyzed. Expression of 67 000 LN-R on the surface of AMC-HN-8 laryngeal carcinoma cells was examined by flow cytometry. The effect of 67 000 LN-R monoclonal antibody (MLuC5) on the adhesive and invasive abilities was observed by adhesion test and Boyden chamber invasiveness test in vitro.</p><p><b>RESULTS</b>The expression level of 67 000 LN-R mRNA in human laryngeal squamous cell carcinoma was significantly higher than that in normal tissues (P < 0.05). The expression level of 67 000 LN-R mRNA in laryngeal squamous cell carcinomas with cervical lymph node metastases was higher than in those without cervical lymph node metastases (P < 0.05). There was a significant negative correlation between the expression level of 67 000 LN-R mRNA and the degree of tumor differentiation, the level being higher in poorly differentiated tumors (P < 0.05). Flow cytometry showed that (80.9 +/- 0.9)% of AMC-HN-8 cells expressed 67 000 LN-R. MLuC5 inhibited the adhesion of AMC-HN-8 cells on LN, and after treated with MLuC5 for 60 and 120 minutes, the adhesion inhibition rate was 57.1% and 63.2%, respectively. The invasive ability to artificial basement membrane was reduced by MLuC5.</p><p><b>CONCLUSION</b>Laryngeal carcinoma overexpressing 67 000 LN-R has stronger invasiveness, and 67 000 LN-R monoclonal antibody may contribute to prevent metastasis of laryngeal carcinoma.</p>

Expression and regulation of integrin receptors in human trophoblast cells: role of estradiol and cytokines.

Embryo implantation and placentation are dynamic cellular events that require not only synchrony between the maternal environment and the embryo, but also complex cell-cell communication amongst the implanting blastocyst and the receptive endometrium through integrins, a large family of proteins involved in the attachment, migration, invasion and control of cellular functions. Integrins display dynamic temporal and spatial patterns of expression by the trophoblast cells during early pregnancy in humans. However, the precise mechanism of embryo implantation and the modulation of the integrin receptors during blastocyst attachment and further implantation remain elusive in the humans. The present study elucidates the expression and hormonal modulation of fibronectin, vitronectin and laminin integrin receptors by estradiol and IL-1alpha in human trophoblast cells. Human first trimester trophoblast cells showed the induction of the classical estrogen receptor (ER)-alpha by its own ligand, estradiol. Treatment with either estradiol or IL-1alpha induced the expressions of alpha4, alpha5, alpha6 and alpha(v) integrin receptor subunits at both the mRNA and protein levels, while expression of beta1 remained unaltered. Furthermore, estradiol upregulated the expression of IL-1alpha, thereby suggesting the possibility that estrogen may either directly or via the proinflammatory cytokine induces the expression of the cell surface integrin receptors. The findings delineate the role of hormones and the cytokines in modulating the adhesiveness and attachment of the trophoblast cells. This may reflect the in vivo scenario where the implanting embryo is surrounded by a hormone-cytokine rich uterine microenvironment that may precisely regulate the expression of integrins and thereby facilitate implantation.

Laminin and its alpha 6 integrin receptor in the regulation of human hepatocellular carcinoma cell phenotypes / 中华肿瘤杂志

<p><b>OBJECTIVE</b>To study the role of integrin alpha 6 in cell attachment, spread, survival/proliferation and differentiation of human hepatocellular carcinoma (HCC) BEL-7402 cells on various substrates by monoclonal antibody against the extracellular domain of alpha 6 subunit (IA6ED McAb).</p><p><b>METHODS</b>The effect of McAb on attachment and spread of BEL-7402 cells on LN or FN substrate was examined. MTT analysis was used to examine the cell survival/proliferation, gelatin zymography to the matrix metalloproteinases (MMPs) secreted by BEL-7402 cells, and microparticle immunoabsorbent kit to AFP secretion while the cells were cultured on LN-, FN- or Matrigel-coated substrates.</p><p><b>RESULTS</b>The cell attachment, spread and survival/proliferation were inhibited. Moreover importantly, the malignant cell dedifferentiation and abnormal differentiation on LN-coated substrate were also strongly inhibited by IA6ED McAb.</p><p><b>CONCLUSION</b>LN and integrin alpha 6 regulate human HCC cell phenotypes of survival/proliferation and differentiation. The phenotypes of dedifferentiation and abnormal differentiation can be reversed by blocking the interaction between integrin alpha 6 and LN using IA6ED McAb, which may lower metastatic potency of tumor cells.</p>

Aspectos biomoleculares del carcinoma basocelular y su relación con la metástasis / Biomolecular aspects of basal cell carcinoma and its relation with metastasis

El carcinoma basocelular es una neoplasia cutánea de crecimiento lento. Rara vez produce metástasis, siendo un modelo ideal para analizar los factores que condicionan la misma. Se han identificado numerosos factores de riesgo: radiación ultravioleta, la cual lesiona directamente al ADN induciendo una inmunodepresión selectiva, algunas cepas del virus papiloma humano y trastornos autosómicos recesivos, donde se identificaron mutaciones del gen supresor del crecimiento tumoral. Actualmente, numerosas líneas de investigación están abocadas al estudio de la baja incidencia de metástasis de éste tumor. En éste carcinoma no existe una disminución en las caderinas epiteliales, las cuales intervienen en la adherencia celular. Asimismo, existe disminución de los receptores de laminina y fibronectina. En la matriz extracelular no se ha observado un aumento de enzimas proteolíticas que la degraden. La inmunovigilancia tumoral está muy activada por medio de un aumento de linfocitos T citolíticos y la producción de interferón gamma. Concluimos que no sólo los queratinocitos neoplásicos intervienen en la metástasis, sino que la matriz extracelular juega un rol preponderante en la escasa incidencia de diseminación de éste tumor, por lo que un estudio exhaustivo de la misma permitiría ser aplicado a otras neoplasias y otorgar una mayor sobrevida

Interaçäo da proteína prion celular com laminina e STI-1 e suas possíveis implicações biológicas / Cellular prion protein interaction with laminin and STI-1 and its possible biological implications

A conversäo da proteína príon celular (PrPc) em sua isoforma anormal PrPsc está associada a uma série de doenças neurodegenerativas, genericamente designadas por doenças priônicas. Embora a literatura tenha enfatizado o estudo do PrPsc e o mecanismo de propagaçäo das doenças de príon, pouco tem sido feito para o entendimento do papel fisiológico do PrPc. Em 1997 nosso grupo descreveu um receptor/ligante para o PrPc utilizando o princípio da hidropaticidade complementar. Neste trabalho isolamos e identificamos este ligante de PrPc como sendo a STI-1 (Stress Inducible Protein-1). In vitro, a STI-1 interage com o PrPc de maneira específica, saturável e com alta afinidade (Kd=8x10-8M)...

Laminin/VLA-6 interactions and T cell function

A growing number of investigators consider extracellular matrix (ECM) proteins to be determinant factors in lymphocyte positioning and activation. One major ECM component is laminin, which is constitutively expressed in the thymus as well as in thymus-dependent areas of peripheral lymphoid organs. In the thymus, laminin is produced by epithelial and dendritic cells, and appears to mediate interactions with thymocytes through specific laminin receptors, in particular the integrin VLA-6. This receptor is also expressed by peripheral T cells, and is apparently involved in effector T cell migration and activation. We showed that CD4+ T lymphocytes from chronic chagasic mice exhibited an increase in the absolute and relative number of cells with high VLA-6 expression. Additionally, it is likely that VLA-6/laminin interactions are required for the development of the CD4+ T cell-dependent anti-myocardial autoreactive process that occurs in these animals. Lastly, laminin can bind to some cytokines, a fact that may represent an additional mechanism by which this extracellular matrix component modulates the behavior of T lymphocytes. Taken together, the present data strongly indicate that interactions involving laminin and VLA-6 are functionally linked to relevant events in T cell physiology, comprising entrance of pro-thymocytes into the thymus, intrathymic T cell migration and differentiation, as well as the functioning of mature T lymphocytes, including effector cells.

Adhesion to laminin is down-regulated upon retinoic acid-induced F9 cell differentiation: a role for alpha6/beta1 integrin

F9 mouse teratocarcinoma cells have a high capacity to adhere to laminin and we identified alpha6/beta1 integrin as the principal laminin-binding protein present in these cells. F9 cells differentiated into parietal endoderm when monolayer cultures were treated with retinoic acid and dibutyryl cyclic AMP. In this process a decreased adherence to laminin was observed due to a lower expression of alpha6/beta1 integrin on the cell surface