PURPOSE: We purpose to determine the correlation of HER-2/neu and paxillin expression in ductal carcinoma in situ (DCIS), invasive ductal carcinoma with ductal carcinoma in situ (IDC with DCIS) and mucinous carcinoma. METHODS: To evaluate the expression of HER-2/neu and paxillin, the immunohistochemical staining was performed for 13 cases of DCIS, 13 cases of IDC with DCIS and 6 cases of mucinous carcinoma. RESULTS: The DCIS and IDC were associated with infiltration of the inflammatory cells, especially in the comedo type and solid type of tumor. In cases with infiltration of the inflammatory cells, HER-2/neu and paxillin were strongly expressed. When comparing the expression level of HER-2/neu from adjacent normal tissue between DCIS and IDC with DCIS, expression of HER-2/neu was similar to that of normal tissue adjacent to DCIS. However, in the adjoining normal ductal epithelial cells, paxillin was highly expressed in cells of all of the tumor types, and especially for IDC with DCIS. HER-2/neu and paxillin were not expressed in mucinous carcinoma cells in all cases. CONCLUSION: HER-2/neu in the DCIS and IDC with infiltration of inflammatory cells shows higher expression than non-inflammatory DCIS and IDC. If normal duct epithelial cells show a high level of HER-2/neu expression, the epithelial cells have a high probability of transformation into anaplastic cells. However, paxillin appears to have no value as a prognostic factor. The difference of expression of HER-2/neu between IDC with DCIS and DCIS suggests a different origin of tumor cells. The growth pattern of mucinous carcinoma cell is different from the that of DCIS or IDC cell, which grow slowly.
Adenocarcinoma, Mucinous ; Carcinoma, Ductal ; Carcinoma, Intraductal, Noninfiltrating ; Epithelial Cells ; Mucins ; Paxillin

PURPOSE: To evaluate adherence of human gingival fibroblasts (HGFs) to transmucosal abutment of dental implant with different surface conditions with time and to investigate the roles of focal adhesion linker proteins (FALPs) involved in HGFs adhesion to abutment surfaces. MATERIALS AND METHODS: Morphologies of cultured HGFs on titanium and ceramic discs with different surface were observed by scanning electron microscopy. Biocompatibility and focal adhesion were evaluated by ultrasonic wave application and cell viability assay. FALPs expression levels were assessed by RT-PCR and western blot. RESULTS: There seemed to be little difference in biocompatibility and adhesion strength of HGFs depending on the surface conditions and materials. In all experimental groups, the number of cells remaining on the disc surface after ultrasonic wave application increased more than 2 times at 3 days after seeding compared to 1-day cultured cells and this continued until 7 days of culture. FALPs expression levels, especially of vinculin and paxillin, also increased in 5-day cultured cells compared to 1-day cultured fibroblasts on the disc surface. CONCLUSION: These results might suggest that the strength of adhesion of fibroblasts to transmucosal abutment surfaces increases with time and it seemed to be related to expressions of FALPs.
Cell Survival ; Cells, Cultured ; Ceramics ; Dental Implants ; Fibroblasts ; Focal Adhesions ; Humans ; Microscopy, Electron, Scanning ; Paxillin ; Proteins ; Seeds ; Titanium ; Ultrasonics ; Vinculin

Paxillin is a regulatory component of the complex of cytoskeletal proteins that link the actin cytoskeleton to the plasma membrane. However, the role of paxillin during smooth muscle contraction is unclear. We investigated a possible role for the membrane-associated dense plaque protein paxillin in the regulation of contraction in rat aortic vascular smooth muscle. The tyrosine phosphorylation of paxillin, which was increased by norepinephrine, reached a peak level after 1 min stimulation and then decreased with time. However, norepinephrine induced a sustained contraction that reached a steady state 30 min after application. Pretreatment with tyrphostin, an inhibitor of tyrosine kinase, inhibited the tyrosine phosphorylation of paxillin and also the contraction stimulated by norepinephrine. Both inhibitions were concentration-dependent, and the degree of correlation between them was high. These results show that, in rat aortic smooth muscle, tyrosine kinase(s) activated by norepinephrine may phosphorylate the tyrosine residues of paxillin, thereby providing a source of regulation during vascular smooth muscle contraction.
Actin Cytoskeleton ; Animals ; Cell Membrane ; Cytoskeletal Proteins ; Muscle, Smooth ; Muscle, Smooth, Vascular* ; Norepinephrine ; Paxillin* ; Phosphorylation* ; Protein-Tyrosine Kinases ; Rats ; Tyrosine*

BACKGROUND: The role of small GTPase molecules is poorly understood under high glucose conditions. METHODS: We analyzed the expression pattern of Vav3 in skeletal muscle C2C12 cells under high glucose culture condition with reverse transcription-polymerase chain reaction and Western blot analysis. We also measured glucose uptake using isotope-labelled glucose. RESULTS: We showed that expression of Vav3 (a guanine nucleotide exchange factor for RhoA) increased. mRNA and protein levels in skeletal muscle C2C12 cells under high glucose conditions. The AMP-activated protein kinase (AMPK) activator AMPK agonist 5-aminoimidazole-4-carboxy-amide-1-d-ribofuranoside (AICAR) suppressed high glucose-induced Vav3 induction. In addition, exposure of cells to high glucose concentration increased the phosphorylation of PAK-1, a molecule downstream of RhoA. The phosphorylation of paxillin, a downstream molecule of PAK-1, was also increased by exposure to high glucose. Phosphorylation of these molecules was not observed in the presence of AICAR, indicating that AMPK is involved in the RhoA signal pathway under high glucose conditions. Knock down of Vav3 enhances metformin-mediated glucose uptake. Inhibition of AMPK blocked the increases of Vav3 knock down-induced glucose uptake. Metformin-mediated Glut4 translocation was also increased by Vav3 knock-down, suggesting that Vav3 is involved in metformin-mediated glucose uptake. CONCLUSION: These results demonstrate that Vav3 is involved in the process of metformin-mediated glucose regulation.
AMP-Activated Protein Kinases ; Blotting, Western ; Glucose* ; GTP Phosphohydrolases ; Guanine Nucleotide Exchange Factors ; Metformin ; Muscle, Skeletal ; Paxillin ; Phosphorylation ; RNA, Messenger ; Signal Transduction

PURPOSE: The fourth state of matter, plasma is known as an ionized gas with electrons, radicals and ions. The use of non-thermal plasma (NTP) in cancer research became possible because of the progresses in plasma medicine. Previous studies on the potential NTP-mediated cancer therapy have mainly concentrated on cancer cell apoptosis. In the present study, we compared the inhibitory effect of NTP on cell migration and invasion in the oral squamous cancer cell lines. MATERIALS AND METHODS: We used oral squamous cancer cell lines (SCC1483, MSKQLL1) and different gases (N₂, He, and Ar). To investigate the mechanism of plasma treatment, using different gases (N₂, He, and Ar) which induces anti-migration and anti-invasion properties, we performed wound healing assay, invasion assay and gelatin zymography. RESULTS: The results showed that NTP inhibits cancer cell migration and invasion of oral squamous cancer cell. In addition, focal adhesion kinase expression and matrix metalloproteinase-2/9 activity were also inhibited. CONCLUSION: The suppression of cancer cell invasion by NTP varied depending on the type of gas. Comparison of the three gases revealed that N₂ NTP inhibited cell migration and invasion most potently via decreased expression of focal adhesion kinase and matrix metalloproteinase activity.
Apoptosis ; Cell Line ; Cell Movement ; Epithelial Cells* ; Focal Adhesion Protein-Tyrosine Kinases ; Gases ; Gelatin ; Ions ; Neoplasms, Squamous Cell* ; Paxillin ; Plasma ; Plasma Gases* ; Wound Healing

The movement of leukocytes from the blood into peripheral tissues is a central feature of immune surveillance, but also contributes to the pathogenesis of inflammatory and autoimmune diseases. Integrins are a family of adhesion and signaling molecules made up of paired alpha and beta subunits, and the integrin alpha4beta1 plays a prominent role in the trafficking of mononuclear leukocytes. We have previously described the direct interaction of the signaling adaptor molecule paxillin with the cytoplasmic domain of the alpha4 integrin subunit. This interaction is critical for alpha4beta1 integrin dependent cell adhesion under shear flow conditions as it provides a needed connection to the actin cytoskeleton. Furthermore, the alpha4-paxillin interaction is required for effective alpha4beta1 dependent leukocyte migration and does so through the temporal and spatial regulation of the small GTPase Rac. These findings make the alpha4-paxillin interaction a potentially attractive therapeutic target in controlling leukocyte trafficking.
Protein Binding ; Paxillin/*metabolism/physiology ; Models, Biological ; Leukocytes/cytology/*metabolism ; Integrin alpha4beta1/metabolism/physiology ; Integrin alpha4/*metabolism/physiology ; Humans ; Cell Movement/*physiology ; Cell Adhesion/physiology

BACKGROUND: We aimed to determine the effect of fibronectin (FN)-immobilized microgrooved titanium (Ti) on human gingival fibroblast proliferation, gene expression and protein expression. METHODS: Photolithography was used to fabricate the microgrooved Ti, and amine funtionalization (silanization) was used for FN immobilization on titanium surfaces. Cell proliferation, gene expression and protein expression were analyzed, followed by multiple regression analysis for determining the influential factors on cell proliferation. RESULTS: FN-immobilized microgrooved Ti significantly enhanced the fibroblast proliferation in various timelines of culture, among which a burst of fivefold increase is induced at 96 h of culture compared to that on the control smooth Ti. We suggest a presence of the synergistic promotion effect of microgrooves and FN immobilization on fibroblast proliferation. Through a series of analyses on the expression of various genes and proteins involved in cell adhesion and proliferation, cyclin-dependent kinase 6, cyclin D1, integrin α5, oncogene c-Src, osteonectin, paxillin and talin-2 were determined as influential factors on promoting fibroblast proliferation induced by FN-immobilized microgrooved Ti. CONCLUSION: FN-immobilized microgrooved Ti can act as an effective surface for enhancing fibroblast proliferation, and can be used for promoting soft tissue response on the connective tissue attachment zone of biomaterial surfaces.
Cell Adhesion ; Cell Proliferation* ; Connective Tissue ; Cyclin D1 ; Cyclin-Dependent Kinase 6 ; Fibroblasts* ; Fibronectins ; Gene Expression ; Humans* ; Immobilization ; Oncogenes ; Osteonectin ; Paxillin ; Titanium*

BACKGROUND: Angiotensin II (Ang II) appears to play important roles in the pathogenesis of hypertension. However, the mechanism by which Ang II induces vascular smooth muscle contraction is not fully understood. The phosphorylation of myosin light chain (MLC) is an essential trigger of the cascade that initiates of smooth muscle contraction. In this study, we investigated the role of MLC phosphorylation on Ang II-induced vascular smooth muscle contraction. METHODS: Rat thoracic aortas were used as an experimental substrates. We measured isometric tension, myosin light chain phosphorylation, intracellular Ca2+ concentration, mitogen-activated protein kinase phosphorylation, and tyrosine phosphorylation. RESULTS: 100 nM Ang II increased smooth muscle contraction transiently in rat thoracic aorta. Ang II also increased intracellular Ca2+ and 20 kDa MLC phosphorylation. Pretreatment with 10microM verapamil and 30microM La3+ abolished the contraction developed at 30 seconds by Ang II, whereas pretreatment with 10microM verapamil and 30microM La3+ abolished the contraction and the intracellular Ca2+ increase induced at 2 minutes by Ang II. Moreover, pretreatment of 10microM verapamil, 30microM La3+ and 1microM thapsigargin abolished the contraction as well as intracellular Ca2+ increase developed at 30 seconds and 2 minutes by Ang II. However, MLC phosphorylation was not affected. GF109203X attenuated Ang II-induced contraction more so than ML-7. 100 nM Ang II increased tyrosine phosphorylation of mitogen-activated protein kinase, 68 and 125 kDa proteins. The 125 kDa protein was confirmed as paxillin in primary vascular smooth muscle cell culture. CONCLUSIONS: Ang II-induced contraction involves Ca2+-dependent and independent components, and Ca2+-dependent contraction by Ang II is mediated by voltage-dependent Ca2+ channel. Moreover, protein kinase C and the mitogen-activated protein kinase activation pathway are involved in Ang II-induced contraction.
Angiotensin II ; Animals ; Aorta, Thoracic ; Calcium ; Cell Culture Techniques ; Hypertension ; Muscle, Smooth* ; Muscle, Smooth, Vascular ; Myosin Light Chains ; Paxillin ; Phosphorylation ; Protein Kinase C ; Protein Kinases ; Rats* ; Thapsigargin ; Tyrosine ; Verapamil

Tumor angiogenesis induced by vascular endothelial cells (VECs) migration is a necessary condition for tumor growth and metastasis. The purpose of this study is to investigate the effect of focal adhesion kinase (FAK) inhibitor (50nmol/mL) on the adhesion and migration of endothelial cells(ECs) and the expression of focal adhesion proteins vinculin, talin and paxillin. Scratch wound migration assay was performed to examine the effect of FAK inhibitor with 50nmol/mL on ECs migration at 0, 5, 10, 30, 60 and 120min, respectively. And immunofluorescence analysis was performed to detect the expression of F-actin in ECs treated with FAK inhibitor within 2h. Western blot was carried out to determine the effect of FAK inhibitor on expression of vinculin, talin and paxillin proteins. The results showed that the migration distance and the expression of F-actin in ECs treated with FAK inhibitor decreased significantly compared with that of the controls, and the level of vinculin showed no significant difference with increasing of treated time of FAK inhibitor. However, the talin and paxillin showed an identical decreasing tendency in 5-10min, but slowly going up in 30min and then after subsequently decreasing. The results of this study proved that blocking phosphorylation of FAK could inhibit VECs adhesion and migration by downregulating focal adhesion proteins so that it may inhibit tumor angiogenesis. This may provide a new approach for tumor therapy.
Cell Adhesion ; Cell Movement ; physiology ; Cells, Cultured ; Endothelial Cells ; cytology ; metabolism ; Focal Adhesion Protein-Tyrosine Kinases ; antagonists & inhibitors ; metabolism ; Focal Adhesions ; metabolism ; physiology ; Humans ; Neoplasms ; blood supply ; Neovascularization, Pathologic ; Paxillin ; metabolism ; Talin ; metabolism ; Vinculin ; metabolism

OBJECTIVETo investigate changes in the adherent ability, the expression of adhesion related proteins Pyk2 and paxillin during HL-60 cells differentiation into granulocyte-monocyte induced by low-dose (LD) bufalin in combination with all-trans retinoic acid (ATRA), and to explore the effects of bortezomib on cellular adhesion and the expression of Pyk2 and paxillin.

METHODSThe expression of CD11b was detected by flow cytometry, cellular adherence ability by MTT assay, and the expressions of Pyk2, paxillin and tubulin by Western blot.

RESULTSThe combination of 5 nmol/L bufalin and 30 nmol/L ATRA induced HL-60 cells differentiation in a time-dependent manner, the percentages of CD11b positive cells treated for 2 d and 4 d being (20.0 +/- 2.8)% and (75.0 +/- 5.3)%, respectively, with the increasing of cellular adherence ability. Meanwhile the expressions of Pyk2 and Paxillin were also up-regulated in a time-dependent manner. Bortezomib suppressed HL-60 cell adhesion in a dose-dependent manner. At concentrations of 1 nmol/L and 10 nmol/L the adherence level were (7.8 +/- 0.1)% and (5.3 +/- 0.3)%, respectively, with down-regulation of Pyk2 but not Paxillin.

CONCLUSIONPyk2 is involved in the regulation of cellular adherence function. Bortezomib might inhibit HL-60 cells adhension function by down-regulation of Pyk2 expression.

Boronic Acids ; pharmacology ; Bortezomib ; Bufanolides ; pharmacology ; Cell Adhesion ; drug effects ; Cell Proliferation ; drug effects ; Focal Adhesion Kinase 2 ; metabolism ; HL-60 Cells ; Humans ; Paxillin ; metabolism ; Pyrazines ; pharmacology ; Tretinoin ; pharmacology