Calcium-independent phospholipase A2beta-Akt signaling is involved in lipopolysaccharide-induced NADPH oxidase 1 expression and foam cell formation.

Foam cell formation is the most important process in atherosclerosis, and low density lipoprotein oxidation by reactive oxygen species (ROS) is the key step in the conversion of macrophages to foam cells. This study reveals the control mechanism of the gene for NADPH oxidase 1 (Nox1), which produces ROS in the formation of foam cells by stimulating TLR4. Treatment of macrophages by the TLR4 agonist LPS stimulated ROS production and ROS-mediated macrophage to foam cell conversion. This LPS-induced ROS production and foam cell formation could be abrogated by pretreatment of macrophages with N-acetyl cysteine or apocynin. LPS increased Nox1 promoter activity, and resultant expression of mRNA and protein. Small interfering RNA mediated inhibition of Nox1 expression decreased LPS-induced ROS production and foam cell formation. LPS-mediated Nox1 expression and the responses occurred in a calcium-independent phospholipase A(2) (iPLA(2))-dependent manner. The iPLA(2)beta-specific inhibitor S-BEL or iPLA(2)beta small interfering RNA attenuated LPS-induced Nox1 expression, ROS production, and foam cell formation. In addition, activation of iPLA(2)beta by LPS caused Akt phosphorylation and was followed by increased Nox1 expression. These results suggest that the binding of LPS and TLR4 increases Nox1 expression through the iPLA(2)beta-Akt signaling pathway, and control ROS production and foam cell formation.

Taxotere Induces Dephosphorylation of MET in Patient-derived Tumor Models.

BACKGROUND/AIM: Although molecular targeting therapy is an attractive treatment for cancer, resistance eventually develops in most cases. Here, we evaluated chemotherapeutic efficacy on non-small cell lung cancer (NSCLC) with acquired resistance to epidermal growth factor receptor inhibitors mechanistically. MATERIALS AND METHODS: Antitumor effects of taxotere were evaluated using multiple models, including xenograft, and patient-derived models developed from adenocarcinoma cancer patients. Protein expressions were analyzed after drug treatment. RESULTS: Taxotere inhibited tumor growth of NSCLC cells harboring drug resistance, and reduced the expression of phosphorylated MET proto-oncogene, receptor tyrosine kinase (MET). A tumor-inhibitory effect of taxotere was also demonstrated in vivo in xenografts in mice, patient-derived primary lung tumor cells and patient-derived xenograft with concomitant repression of phosphorylated MET expression. Chemotherapeutic and MET-targeting drug exhibited a synergistic cell growth-inhibitory effect. CONCLUSION: These results suggest that the anticancer drug taxane may be an adjuvant for lung tumors exhibiting enhanced signaling of MET networks.

Primary motor cortex activation by transcranial direct current stimulation in the human brain.

Transcranial direct current stimulation (tDCS) can modulate motor cortex excitability in the human brain. We attempted to demonstrate the cortical stimulation effect of tDCS on the primary motor cortex (M1) using functional MRI (fMRI). An fMRI study was performed for 11 right-handed healthy subjects at 1.5 T. Anodal tDCS was applied to the scalp over the central knob of the M1 in the left hemisphere. A constant current with an intensity of 1.0 mA was applied. The total fMRI paradigm consisted of three sessions with a 5-min resting period between each session. Each session consisted of five successive phases (resting-tDCS-tDCS-tDCS-tDCS), and each of the phases was performed for 21s. Our findings revealed that no cortical activation was detected in any of the stimulation phases except the fourth tDCS phase. In the result of group analysis for the fourth tDCS phase, the average map indicated that the central knob of the left primary motor cortex was activated. In addition, there were activations on the left supplementary motor cortex and the right posterior parietal cortex. We demonstrated that tDCS has a direct stimulation effect on the underlying cortex. It seems that tDCS is a useful modality for stimulating a target cortical region.

Cortical effect and functional recovery by the electromyography-triggered neuromuscular stimulation in chronic stroke patients.

We investigated the effect of electromyography (EMG)-triggered neuromuscular electrical stimulation (NMES; EMG-stim) on functional recovery of the hemiparetic hand and the related cortical activation pattern in chronic stroke patients. We enrolled 14 stroke patients, who were randomly assigned to the EMG-stim (n=7) or the control groups (n=7). The EMG-stim was applied to the wrist extensor of the EMG-stim group for two sessions (30 min/session) a day, five times per week for 10 weeks. Four functional tests (box and block, strength, the accuracy index, and the on/offset time of muscle contraction) and functional MRI (fMRI) were performed before and after treatment. fMRI was measured at 1.5 T in parallel with timed finger flexion-extension movements at a fixed rate. Following treatment, the EMG-stim group showed a significant improvement in all functional tests. The main cortical activation change with such functional improvement was shifted from the ipsilateral sensorimotor cortex (SMC) to the contralateral SMC. We demonstrated that 10-week EMG-stim can induce functional recovery and change of cortical activation pattern in the hemiparetic hand of chronic stroke patients.

Effect of constraint-induced movement therapy with modified opposition restriction orthosis in chronic hemiparetic patients with stroke.

OBJECTIVES: Constraint-induced movement therapy (CIMT) has been demonstrated to be effective in improving hemiparetic upper extremity function in stroke patients, but few studies have been performed to assess orthosis modification. We investigated the effect of the newly designed small orthosis named modified opposition restriction orthosis (MORO) in chronic hemiparetic patients with stroke. DESIGN: Twenty-one stroke patients were randomly assigned to the CIMT group or control group. Thirteen patients in the CIMT group wore MORO confining the thumb and index finger for at least 5 hours of each day, 7 days a week for 8 weeks. The affected upper extremity function was evaluated using the manual function test (MFT), Purdue Pegboard (PP) score, and motor activity log (MAL) at pre and post-CIMT. RESULTS: Four of the 13 patients in the CIMT group dropped out due to motivational problems, and 9 patients remained in the CIMT group at the end of the study. The patients in the CIMT group showed a mean improvement of 195.8% on MAL AOU (Amount of Use), 24.6% on PP score, and 5.5% on MFT. CONCLUSION: This new MORO would be effective for use in a CIMT program in chronic hemiparetic patients with stroke.

Brain activation pattern according to exercise complexity: a functional MRI study.

The aim of this study was to compare the areas of brain activation between complex and simple exercises in a unimanual hand and to assess the possibility of an exercise task for paretic hands following stroke. The subjects included 11 healthy right-handed volunteers. The complex exercise was a wooden ball rotation task with the unimanual hand and the simple exercise was a hand grasp task performed during a functional MRI scan. Stronger activation of the left primary sensorimotor cortex, the left premotor area, and the ipsilateral cerebellum emerged when the complex movement was performed. Ipsilateral activity was located in the primary sensory cortex and premotor area, and contralateral activity was shown in the left cerebellum. These results suggest that a unimanual ball rotation task may be appropriate for rehabilitation of a movable paretic hand in an early stage of stroke recovery, which should provide motor and sensory input using external stimuli, while the simple motor task may appropriate in a compensatory stage, and should inhibit the ipsilateral activity due to maladaptive plasticity.

Motor outcome according to the integrity of the corticospinal tract determined by diffusion tensor tractography in the early stage of corona radiata infarct.

Diffusion tensor tractography (DTT) is useful for exploring the state of the corticospinal tract (CST). An accurate estimation of the integrity of the CST in the early stage of a cerebral infarct would enable a determination of motor recovery. DTT was performed to classify CST integrity following a corona radiata infarct to evaluate if the procedure could characterize the motor outcome of the affected hand. Fifty-five patients with completely paralyzed hands due to a corona radiata infarct were recruited for the study, and DTT images were obtained within 7-30 days after a stroke. The DTI findings for the patients were classified into four groups. In type A, the CST was preserved around the infarct; in type B, the CST originated from a cortex other than the primary motor cortex; in type C, the CST was interrupted at the infarct; in type D, the CST failed to reach the infarct due to degeneration. Six months after a stroke, the motor function of the affected hand was evaluated with the motricity index (MI) for the hand, the Medical Research Council score (MRC) for finger extensors and the modified Brunnstrom classification (MBC). These indices were significantly influenced by the DTT type (p<0.05). The highest MI, MRC and MBC were seen in the DTT type A patients; the lowest MI, MRC and MBC were seen in the DTT type D patients (p<0.05). The integrity of the corticospinal tract determined by DTT obtained during the early stage of a corona radiata infarct seems to be helpful in predicting the motor outcome of the affected hand.

Motor outcome according to diffusion tensor tractography findings in the early stage of intracerebral hemorrhage.

We tried to investigate the motor outcome according to diffusion tensor tractography (DTT) findings for the corticospinal tract (CST) in the early stage for hemiparetic patients with intracerebral hemorrhage (ICH). Forty patients with severe paralysis of the affected side were enrolled. DTT was obtained in the early stage of the stroke (7-30 days) and was classified into four groups: type A, the CST originating from primary motor cortex was preserved around the hematoma; type B, the CST was similar to type A except the fiber originated from the adjacent areas to the primary motor cortex; type C, the CST was interrupted at or around the hematoma; and type D, the CST did not reach the hematoma due to degeneration (Fig. 1). Six months after onset, motor function was measured and the statistical influence of the DTT type was tested. Initially, none of the motor function scales of the affected side differed among the four DTT types. Six months after the onset of ICH, motor functions of the same side were significantly different according to DTT type (p<0.05). All motor scales were highest in the DTT type A group, and were lowest in the DTT type D group (p<0.0003). The early DTT findings for CST may be used to predict the motor outcome of the affected extremities in hemiparetic patients with ICH.

Cortical activation changes induced by visual biofeedback tracking training in chronic stroke patients.

OBJECTIVES: We tried to examine whether visual biofeedback tracking training (VBTT) can improve both the gait performance and cortical activation pattern in chronic stroke patients. DESIGN: We enrolled 10 chronic hemiparetic patients with stroke(mean age 46.3 +/- 5.19 years). The patients were randomly assigned to the training group (5 patients) or the control group (5 patients). VBTT was to follow the PC-generated sine waves with the knee joint electrogoniometer, and the two sine waves should appear as close to overlapping as possible on the PC monitor. The training was performed for 39 minutes/day, 5 days/week, for 4 weeks. Pre-training and post-training accuracy of tracking, functional status of gait, and functional MRI (fMRI) were measured. fMRI was performed at 1.5 T in parallel with timed knee flexion-extension movements at a fixed rate. RESULTS: The accuracy of the tracking performance, walking speed, and motor scale for gait improved in the training group. Primary sensorimotor cortex (SM1) cortical activation shifted significantly from the unaffected to the affected hemisphere in the training group. CONCLUSIONS: We demonstrated that cortical activation changes occurred with gait function improvement in chronic stroke patients throughout the 4-week VBTT program. It seems that the cortical reorganization was induced by VBTT.

Motor recovery via the peri-infarct area in patients with corona radiata infarct.

OBJECTIVES: Diffusion tensor image tracography (DTT) could be useful for exploration of the state of the corticospinal tract at the subcortical white matter level. We investigated the neural pathway associated with motor recovery in the patients with corona radiata infarct (CR) using DTT. DESIGN: Three hemiparetic patients who showed severe weakness of the affected upper extremity at stroke onset, were recruited. DTT was performed twice (subacute and chronic stage) using 1.5-T system. Three-dimensional reconstructions of the fiber tracts were obtained with FA <0.2, angle >45 degrees as termination criteria. RESULTS: In all patients, the motor function of the affected upper extremity has recovered to be functional state at second DTT scanning. The motor tracts of the affected hemisphere were observed to pass along the posterior portion of the CR infarct on both the first and second DTT. CONCLUSIONS: It seems that the motor function of the affected upper extremity of the patients recovered via the posterior portion of infarct. This finding may reflect functional reorganization of the motor pathway following damage to the corticospinal tract.

Development and validation of a subject-specific finite element model of the functional spinal unit to predict vertebral strength.

Finite element models of an isolated vertebral body cannot accurately predict compressive strength of the spinal column because, in life, compressive load is variably distributed across the vertebral body and neural arch. The purpose of this study was to develop and validate a patient-specific finite element model of a functional spinal unit, and then use the model to predict vertebral strength from medical images. A total of 16 cadaveric functional spinal units were scanned and then tested mechanically in bending and compression to generate a vertebral wedge fracture. Before testing, an image processing and finite element analysis framework (SpineVox-Pro), developed previously in MATLAB using ANSYS APDL, was used to generate a subject-specific finite element model with eight-node hexahedral elements. Transversely isotropic linear-elastic material properties were assigned to vertebrae, and simple homogeneous linear-elastic properties were assigned to the intervertebral disc. Forward bending loading conditions were applied to simulate manual handling. Results showed that vertebral strengths measured by experiment were positively correlated with strengths predicted by the functional spinal unit finite element model with von Mises or Drucker-Prager failure criteria ( R2 = 0.80-0.87), with areal bone mineral density measured by dual-energy X-ray absorptiometry ( R2 = 0.54) and with volumetric bone mineral density from quantitative computed tomography ( R2 = 0.79). Large-displacement non-linear analyses on all specimens did not improve predictions. We conclude that subject-specific finite element models of a functional spinal unit have potential to estimate the vertebral strength better than bone mineral density alone.

Activation of toll-like receptor-9 induces matrix metalloproteinase-9 expression through Akt and tumor necrosis factor-alpha signaling.

CpG oligodeoxunucleotide (ODN) plays an important role in immune cell function. The present study examined whether temporal control of toll-like receptor (TLR)-9 by CpG ODN can regulate the expression of matrix metalloproteinase-9 (MMP-9). CpG ODN induced the release of tumor necrosis factor (TNF)-alpha and the expression of TNF receptor (TNFR)-II, but not of TNFR-I, in a time-dependent manner and stimulated significant, though delayed, MMP-9 expression. The endosomal acidification inhibitors, chloroquine or bafilomycin A, inhibited CpG ODN-induced TNF-alpha, TNFR-II, and MMP-9 expression. CpG ODN induced the phosphorylation of Akt, and the inhibition of Akt by LY294002 suppressed CpG ODN-induced TNF-alpha, TNFR-II, and MMP-9 expressions. Moreover, neutralizing TNF-alpha antibody significantly suppressed CpG ODN-induced MMP-9 expression, suggesting the involvement of TNF-alpha. These observations suggest that CpG ODN may play important roles in macrophage activation by regulating the expression of MMP-9 via a TLR-9/Akt/TNF-alpha-dependent signaling pathway.

Substrate specificity of nonribosomal peptide synthetase modules responsible for the biosynthesis of the oligopeptide moiety of cephabacin in Lysobacter lactamgenus.

Lysobacter lactamgenus produces cephabacins, a class of beta-lactam antibiotics which have an oligopeptide moiety attached to the cephem ring at the C-3 position. The nonribosomal peptide synthetase (NRPS) system, which comprises four distinct modules, is required for the biosynthesis of this short oligopeptide, when one takes the chemical structure of these antibiotics into consideration. The cpbI gene, which has been identified in a region upstream of the pcbAB gene, encodes the NRPS - polyketide synthase hybrid complex, where NRPS is composed of three modules, while the cpbK gene -- which has been reported as being upstream of cpbI-- comprises a single NRPS module. An in silico protein analysis was able to partially reveal the specificity of each module. The four recombinant adenylation (A) domains from each NRPS module were heterologously expressed in Escherichia coli and purified. Biochemical data from ATP-PPi exchange assays indicated that L-arginine was an effective substrate for the A1 domain, while the A2, A3 and A4 domains activated L-alanine. These findings are in an agreement with the known chemical structure of cephabacins, as well as with the anticipated substrate specificity of the NRPS modules in CpbI and CpbK, which are involved in the assembly of the tetrapeptide at the C-3 position.

Effect of ethanol extract of dried Chinese yam (Dioscorea batatas) flour containing dioscin on gastrointestinal function in rat model.

In this study, a 40% ethanol extract of Chinese yam flour (Dioscoreae rhizoma), containing 177 +/- 58 microg/mL of dioscin, was tested in order to evaluate its pharmacological effects on the gastrointestinal tracts of Sprague-Dawley rats. Via the ingestion of the Chinese yam extract, the secretion of gastric acid was suppressed in the rats, and gastrointestinal motility increased by as much as 10%. The fecal quantity of rats fed on the Chinese yam extract also increased, by more than 40% as compared with that of the controls. The Chinese yam extract was found not to affect the growth of normal intestinal bacteria. However, a great deal of lactose-fermenting bacteria was observed in the fecal samples of rats fed for 6 weeks on 2% Chinese yam extract. This finding would appear to suggest that Chinese yam extract not only induces an improvement in digestive capability, but also affects the conversion of some intestinal flora to helpful bacteria. Our serochemical analyses indicated that serum glucose, neutral lipid, and total cholesterol levels were reduced to some degree by long-term feeding on Chinese yam extract. This finding bolsters the notion that Chinese yam extract may prove helpful as a digestion-aiding agent for patients suffering from hyperglycemia or hyperlipidemia.

MAP kinase activation is required for the MMP-9 induction by TNF-stimulation.

MMP-9 is a metalloproteinase capable of basement membrane degradation in vivo. Expression of MMP-9 can be found in normal conditions such as trophoblasts, osteoclasts, and leukocytes and their precursors. They also occur as well as in pathological conditions, such as the invasive growth of primary tumors, metastasis, angiogenesis, rheumatoid arthritis, and periodontal diseases. MMP-9 upregulation can be highly induced by a wide range of agents. These agents include growth factors, cytokines, cell-cell, and cell-ECM adhesion molecules, and agents altering cell shape. Here, we observed that TNF-alpha stimulated human monocytic cell line, HL-60 produced MMP-9 in a dose and time dependent manner. Real time PCR results indicated transcriptional upregulation of MMP-9 as early as 3 h post TNF-alpha stimulation. To investigate the signaling pathway underlined in TNF-alpha induced MMP-9 expression, three MAP kinase inhibitors were added to cells 1 h prior to TNF-alpha treatment. The ERK inhibitor completely abolished MMP-9 expression by TNF-alpha. But neither p38 MAP kinase nor JNK inhibitor had an effect on TNF-alpha induced MMP-9 expression, suggesting that ERK activation is required for the MMP-9 induction by TNF-alpha. Taken together, we found that TNF-alpha stimulation facilitates ERK activation, which results in the transcriptional upregulation of MMP-9 gene and subsequent MMP-9 production and secretion.

The 5-enolpyruvylshikimate-3-phosphate synthase of glyphosate-tolerant soybean expressed in Escherichia coli shows no severe allergenicity.

The recombinant gene was amplified from the chromosomal DNA of genetically-modified (GM) soybeans and identified as epsps encoding 5-enolpyruvylshikimate-3-phosphate synthase (EPSPS) which renders glyphosate resistance. The epsps structural gene was introduced in the pET28(a) plasmid for its expression in Escherichia coli BL21(DE3). It was confirmed that the maximal productivity of the EPSPS protein was achieved when cultivating the recombinant strain in a LB broth for 2 h after supplementing 1 mM isopropylbeta-D-thiogalactopyranoside (IPTG) in a 2 h-culture broth. Since the expressed EPSPS protein was found as an insoluble form in the inclusion body, it was extracted by 6 M urea after sonication, and then purified through immobilized nickel-affinity column chromatography to isolate EPSPS having a molecular mass of 57 kDa. When incubated in simulated gastric fluid containing pepsin at pH 1.5, the purified EPSPS protein was completely digested within 1 min. In addition, the passive cutaneous anaphylaxis reaction of the purified EPSPS protein was not observed in the Sprague Dawley rat system that was administered either orally or subcutaneously. Furthermore, treatment of the EPSPS protein to the culture of the sensitized peritoneal mast cells, or unsensitized but antisera-labeled mast cells, showed neither a remarkable change in the histamine release nor a cytokine production, including interleukin-4 (IL-4) and tumor necrosis factor-alpha (TNF-alpha). Thus, it can be concluded that the EPSPS protein in the GM soybean showed no significant allergenicity in the Sprague Dawley rats.

Glycogen synthase kinase 3beta and beta-catenin pathway is involved in toll-like receptor 4-mediated NADPH oxidase 1 expression in macrophages.

Macrophage activation contributes to the pathogenesis of atherosclerosis. In the vascular system, the major source of reactive oxygen species is the NADPH oxidase (Nox) family. Nox1 is induced by lipopolysaccharide (LPS) in macrophages, but the expression mechanism is not fully understood. We found that LPS causes beta-catenin accumulation by glycogen synthase kinase 3beta (GSK3beta) inactivation, and that beta-catenin accumulation increases Nox1 expression. LPS induced Nox1 mRNA expression and reactive oxygen species generation in Raw264.7 cells. Using bone marrow-derived macrophages from toll-like receptor 4 mutant mice, we also tested whether LPS-induced Nox1 expression is toll-like receptor 4 dependent. LPS caused GSK3beta phosphorylation, induced beta-catenin accumulation and increased nuclear translocation. The GSK3beta inhibitor LiCl potentiated LPS-induced Nox1 expression in accordance with beta-catenin accumulation and nuclear translocation. Conversely, ectopic expression of a constitutively active GSK3beta mutant severely attenuated Nox1 expression. These findings identify a novel regulatory pathway controlling Nox1 expression by LPS-stimulated macrophages.

Toll-like receptor 9-mediated inhibition of apoptosis occurs through suppression of FoxO3a activity and induction of FLIP expression.

Synthetic oligodeoxynucleotides (ODN) with a CpG-motif are recognized by Toll-like receptor 9 (TLR9) and pleiotropic immune responses are elicited. Stimulation of macrophages with TLR9 agonist prevented apoptosis induced by serum deprivation through increased expression of FLICE-like inhibitory protein (FLIP). CpG ODN-mediated anti-apoptosis depended on the TLR9-Akt-FoxO3a signaling pathway. Inhibition of TLR9 by small interfering (si) RNA or an inhibitor suppressed CpG ODN-mediated anti-apoptosis. Analysis of signaling pathways revealed that the anti-apoptotic effect of CpG ODN required phosphorylation of FoxO3a and its translocation from the nucleus to the cytosol. Overexpression of FoxO3a increased apoptosis induced by serum deprivation and CpG ODN blocked these effects through FLIP expression. In contrast, siRNA knock-down of FoxO3a decreased apoptosis by serum deprivation. In addition, Akt activation was involved in CpG ODN-induced phosphorylation of FoxO3a, expression of FLIP, and anti-apoptosis. Taken together, these results demonstrate the involvement of Akt-FoxO3a in TLR9-mediated anti-apoptosis and indicate that FoxO3a is a distinct regulator for FLIP expression.

Arsenic trioxide induces apoptosis in human colorectal adenocarcinoma HT-29 cells through ROS.

PURPOSE: Treatment with arsenic trioxide (As(2)O(3)) results in a wide range of cellular effects that includes induction of apoptosis, inhibition of cell growth, promotion or inhibition of cellular differentiation, and inhibition of angiogenesis through a variety of mechanisms. The mechanisms of As(2)O(3)-induced cell death have been mainly studied in hematological cancers, and those mechanisms in solid cancers have yet to be clearly defined. In this study, the mechanisms by which As(2)O(3) induces apoptosis in human colorectal adenocarcinoma HT-29 cells were investigated. MATERIALS AND METHODS: To examine the levels of apoptosis, HT-29 cells were treated with As(2)O(3) and then we measured the percentage of Annexin V binding cells, the amount of ROS production and the mitochondrial membrane potential. Western blot analysis was performed to identify the activated caspases after As(2)O(3) exposure, and we compared the possible target molecules of apoptosis. As(2)O(3) treatment induced the loss of the mitochondrial membrane potential and an increase of ROS, as well as activation of caspase-3, -7, -9 and -10. RESULTS: As(2)O(3) induced apoptosis via the production of reactive oxygen species and the loss of the mitochondrial membrane potential. As(2)O(3) induced the activation of caspase-3, -7, -9 and -10. Furthermore, As(2)O(3) treatment downregulates the Mcl-1 and Bcl-2 expressions, and the release of cytochrome c and an apoptosis-inducing factor (AIF). Pretreating the HT-29 cells with N-acetyl-L-cysteine, which is a thiol-containing antioxidant, inhibited the As(2)O(3)-induced apoptosis and caspase activation. CONCLUSION: Taken together, these results suggest that the generation of reactive oxygen species (ROS) by As(2)O(3) might play an important role in the regulation of As(2)O(3)-induced apoptosis. This cytotoxicity is mediated through a mitochondria-dependent apoptotic signal pathway in HT-29 cells.

Group IB secretory phospholipase A2 promotes matrix metalloproteinase-2-mediated cell migration via the phosphatidylinositol 3-kinase and Akt pathway.

Secretory phospholipase A(2) (sPLA(2)), abundantly expressed in various cells including fibroblasts, is able to promote proliferation and migration. Degradation of collagenous extracellular matrix by matrix metalloproteinase (MMP) plays a role in the pathogenesis of various destructive disorders, such as rheumatoid arthritis, tumor invasion, and metastasis. Here we show that group IB PLA(2) increased pro-MMP-2 activation in NIH3T3 fibroblasts. MMP-2 activity was stimulated by group IB PLA(2) in a dose- and time-dependent manner. Consistent with MMP-2 activation, sPLA(2) decreased expression of type IV collagen. These effects are due to the reduction of tissue inhibitor of metalloproteinase-2 (TIMP-2) and the activation of the membrane type1-MMP (MT1-MMP). The decrease of TIMP-2 levels in conditioned media and the increase of MT1-MMP levels in plasma membrane were observed. In addition, treatment of cells with decanoyl Arg-Val-Lys-Arg-chloromethyl ketone, an inhibitor of pro-MT1-MMP, suppressed sPLA(2)-mediated MMP-2 activation, whereas treatment with bafilomycin A1, an inhibitor of H(+)-ATPase, sustained MMP-2 activation by sPLA(2). The involvement of phosphatidylinositol 3-kinase (PI3K) and Akt in the regulation of MMP-2 activity was further suggested by the findings that PI3K and Akt were phosphorylated by sPLA(2). Expression of p85alpha and Akt mutants, or pretreatment of cells with LY294002, a PI3K inhibitor, attenuated sPLA(2)-induced MMP-2 activation and migration. Taken together, these results suggest that sPLA(2) increases the pro-MMP-2 activation and migration of fibroblasts via the PI3K and Akt-dependent pathway. Because MMP-2 is an important factor directly involved in the control of cell migration and the turnover of extracellular matrix, our study may provide a mechanism for sPLA(2)-promoted fibroblasts migration.