Finding Order in Chaos: Quantitative Predictors of Chaos Terrain Morphology on Europa.

The mechanisms for chaos terrain formation on Europa have long been a source of debate in the scientific community. There exist numerous theoretical and numerical models for chaos formation, but to date there has been a lack of quantifiable observations that can be used to constrain models and permit comparison to the outputs of these chaos models. Here, we use mapping and statistical analysis to develop a quantitative description of chaos terrain and their observed morphologies. For nine chaos features, we map every block, or region of pre-existing terrain within disrupted matrix. We demonstrate that chaos terrains follow a continuous spectrum of morphologies between two endmembers, platy and knobby. We find that any given chaos terrain's morphology can be quantified by means of the linearized exponential slope of its cumulative block area distribution. This quantitative metric provides a new diagnostic parameter in future studies of chaos terrain formation and comparison.

Macrophage stimulating protein: purification, partial amino acid sequence, and cellular activity.

Macrophage stimulating protein (MSP) was purified to homogeneity from human blood plasma by selection of biologically active fractions obtained by sequential immunoaffinity and high pressure liquid ion exchange chromatography. By sodium dodecyl sulfate-polyacrylamide gel electrophoresis the molecular mass of MSP was 70 kilodaltons (kD); under reducing conditions two gel bands were seen, at 47 and 22 kD. The disulfide-linked two-chain structure of MSP was confirmed by separation of reduced and alkylated MSP chains. A computer search comparison of six partial sequences of MSP digests showed that MSP has not been recorded in data banks of protein sequences. Two MSP fragments had greater than 80% identity in overlaps of 12-16 residues to sequences in the protein family that includes human prothrombin, plasminogen, and hepatocyte growth factor. The concentration of purified MSP required for half-maximal biological activity was the order of 10(-10) M. In addition to making mouse resident peritoneal macrophages responses to chemoattractants, MSP caused the appearance of long cytoplasmic processes and pinocytic vesicles in freshly plated macrophages. MSP also caused phagocytosis via the C3b receptor, CR1. Whereas resident peritoneal macrophages bind but do not ingest sheep erythrocytes opsonized with IgM anti-Forssman antibody and mouse C3b, addition of MSP caused ingestion. Thus, MSP causes direct or indirect activation of two receptors of the mouse resident peritoneal macrophage, CR1 and the C5a receptor.

Purification and amino acid analysis of two human glioma-derived monocyte chemoattractants.

Two chemoattractants for human monocytes were purified to apparent homogeneity from the culture supernatant of a glioma cell line (U-105MG) by sequential chromatography on Orange A-Sepharose, an HPLC cation exchanger, and a reverse phase HPLC column. On SDS-PAGE gels under reducing or nonreducing conditions, the molecular masses of the two peptides glioma-derived chemotactic factor 1 and 2 were 15 and 13 kD, respectively. Amino acid composition of these molecules was almost identical, and differed from other cytokines that have been reported. The NH2 terminus of each peptide was apparently blocked. When tested for chemotactic efficacy, the peptides attracted approximately 30% of the monocytes added to chemotaxis chambers, at the optimal concentration of 10(-9) M. Potency and efficacy were comparable with that of FMLP, which is often used as a reference attractant. The activity was chemotactic rather than chemokinetic. In contrast to their interaction with human monocytes, the pure peptides did not attract neutrophils. These pure tumor-derived chemoattractants can now be compared with attractants produced by normal cells and evaluated for their biological significance in human neoplastic disease.

Molecular cloning of a human monocyte-derived neutrophil chemotactic factor (MDNCF) and the induction of MDNCF mRNA by interleukin 1 and tumor necrosis factor.

The cDNA coding for human monocyte-derived neutrophil-specific chemotactic factor (MDNCF) was cloned from LPS-stimulated human monocyte mRNA. The cDNA sequence codes for a polypeptide consisting of 99 amino acids, including a putative signal sequence. Comparison of the deduced amino acid sequence with the NH2-terminal amino acid sequence of natural MDNCF shows that the mature functional protein comprises 72 amino acids, beginning with serine at residue 28. The deduced amino acid sequence shows striking similarity to several platelet-derived factors, a v-src-induced protein, a growth-regulated gene product (gro), and an IFN-gamma inducible protein. The availability of the MDNCF cDNA enabled us to use it as a probe to identify inducers of MDNCF mRNA expression in human PBMC. MDNCF mRNA was increased greater than 10-fold within 1 h after stimulation with LPS, IL-1, or TNF, but not by IFN-gamma, IFN-alpha, or IL-2. Furthermore, we also determined that LPS, IL-1, and TNF stimulated the mononuclear cells to produce biologically active MDNCF. This observation may account for the in vivo capacity of IL-1 and TNF to induce netrophil infiltrates.

Identification of the ron gene product as the receptor for the human macrophage stimulating protein.

Macrophage-stimulating protein (MSP) is a member of the hepatocyte growth factor-scatter factor (HGF-SF) family. Labeled MSP bound to Madin-Darby canine kidney (MDCK) cells transfected with complementary DNA encoding Ron, a cell membrane protein tyrosine kinase. Cross-linking of 125I-labeled MSP to transfected cells (MDCK-RE7 cells) and immunoprecipitation by antibodies to Ron revealed a 220-kilodalton complex, a size consistent with that of MSP (80 kilodaltons) cross-linked to the beta chain of Ron (150 kilodaltons). The binding of 125I-labeled MSP to MDCK-RE7 cells was inhibited by unlabeled MSP, but not by HGF-SF. MSP caused phosphorylation of the beta chain of Ron and induced migration of MDCK-RE7 cells. These results establish the ron gene product as a specific cell-surface receptor for MSP.

Proteolytic cleavage and activation of pro-macrophage-stimulating protein by resident peritoneal macrophage membrane proteases.

Macrophage stimulating protein (MSP), which is secreted as biologically inactive pro-MSP, is activated to MSP by cleavage at a single peptide bond. Our objectives were to determine the form of MSP in circulating blood and to study proteolytic activation of pro-MSP by its target cell. Western blot of immunoaffinity-purified serum MSP showed that all the protein was pro-MSP, without detectable MSP. The circulating form of the protein is therefore pro-MSP, and conversion to MSP does not occur when blood is shed. Incubation of radiolabeled pro-MSP with murine peritoneal macrophages caused proteolytic cleavage to predominantly inactive fragments. Among several protease inhibitors, soybean trypsin inhibitor was one of two that inhibited nonspecific cleavage and revealed a macrophage proteolysis of pro-MSP, and certain concentrations enhanced cleavage to mature MSP. Macrophage membranes had nonspecific and specific pro-MSP proteolytic activity, which was not present in macrophage culture fluids. The results suggest that control of MSP activity can occur at the level of the target cell by proteolytic cleavage of pro-MSP to mature MSP or to inactive fragments.

Macrophages cultured in vitro release leukotriene B4 and neutrophil attractant/activation protein (interleukin 8) sequentially in response to stimulation with lipopolysaccharide and zymosan.

The capacity of lipopolysaccharide (LPS), zymosan, and calcium ionophore A23187 to induce neutrophil chemotactic activity (NCA), leukotriene B4 (LTB4), and neutrophil attractant/activation protein (NAP-1) release from human alveolar macrophages (AM) retrieved from normal nonsmokers was evaluated. LPS induced a dose-dependent release of LTB4 that began by 1 h, 4.0 +/- 3.2 ng/10(6) viable AM; peaked at 3 h, 24.7 +/- 13.5 ng/10(6) viable AM; and decreased by 24 h, 1.2 +/- 1.0 ng/10(6) viable AM (n = 8). Quantities of LTB4 in cell-free supernatants of AM stimulated with LPS were determined by reverse-phase high-performance liquid chromatography and corresponded well with results obtained by radioimmunoassay. By contrast, NAP-1 release began approximately 3-5 h after stimulation of AM with LPS, 197 +/- 192 ng/ml, and peaked at 24 h, 790 +/- 124 ng/ml. Release of NAP-1 was stimulus specific because A23187 evoked the release of LTB4 but not NAP-1, whereas LPS and zymosan induced the release of both LTB4 and NAP-1. The appearance of neutrophil chemotactic activity in supernatants of AM challenged with LPS for 3 h could be explained completely by the quantities of LTB4 present. After stimulation with LPS or zymosan for 24 h, AM had metabolized almost all generated LTB4. Preincubation of AM with nordihydroguiaretic acid (10(-4) M) completely abolished the appearance of NCA, LTB4, and NAP-1 in supernatants of AM challenged with LPS. Therefore, LPS and zymosan particles were potent stimuli of the sequential release of LTB4 and NAP-1 from AM.

Neutrophil attractant protein-1-immunoglobulin G immune complexes and free anti-NAP-1 antibody in normal human serum.

After obtaining data indicating the presence of a neutrophil attractant protein-1 (NAP-1)-IgG complex in normal human serum, we developed sandwich ELISAs that could quantify NAP-1 and NAP-1-IgG in mixtures of the two moieties. The ELISA for free NAP-1 used a monoclonal capture antibody that did not bind NAP-1-IgG. The ELISA for NAP-1-IgG was based on omission of the anti-NAP-1 detection antibody (required for the free NAP-1 ELISA) and on interaction of phosphatase-conjugated anti-human IgG with the human NAP-1-IgG complex. Gel filtration of immunoaffinity-purified NAP-1-IgG showed that the bulk of the complex comprised a single IgG. Binding between NAP-1 and antibody is strong, since 8 M urea at neutral or alkaline pH did not release NAP-1. However, at pH 2.0 in 9 M urea approximately 15% of the total NAP-1 could be dissociated from the complex. NAP-1-IgG was detected in 18 of 26 sera from normal humans. The mean serum concentration was 58 ng of IgG-bound NAP-1/ml, with an SEM of 16 and a range from undetectable to 247 ng/ml. NAP-1-IgG concentrations in paired sera drawn at a 1-mo interval were remarkably constant. Using an ELISA for free NAP-1 with a detection limit of 200 pg/ml, we found no free NAP-1 in the 26 sera. Free anti-NAP-1-IgG autoantibody was found in 9 of 26 sera by direct ELISA. IgG anti-NAP-1 of all nine sera was polyclonal, comprising both kappa and lambda isotypes; predominant subclasses were IgG2 and IgG3. NAP-1-IgG did not compete with 125I-NAP-1 for binding to neutrophils, which suggests that IgG anti-NAP-1 is a molecular trap that prevents binding of NAP-1 to neutrophils after it diffuses from production sites into the circulation.

Two independent signaling pathways mediate the antiapoptotic action of macrophage-stimulating protein on epithelial cells.

In addition to its effects on macrophage function, macrophage-stimulating protein (MSP) is a growth and motility factor for epithelial cells. The growth and survival of epithelial cells generally require two signals, one generated by interaction with extracellular matrix via integrins, the other initiated by a growth factor. Therefore we investigated the effect of MSP on epithelial cell survival. Survival of epithelial cells cultured overnight in serum-free medium was promoted by adhesion, which activated both the phosphatidylinositol 3'-kinase (PI3-K)/AKT and mitogen-activated protein kinase (MAPK) pathways, operating independently of one another. The number of apoptotic cells resulting from inhibition of either pathway alone was approximately doubled by simultaneous inhibition of both pathways. This shows that each pathway made a partial contribution to the prevention of apoptosis. In the presence of an inhibitor of either pathway, MSP increased the activity of the other pathway so that the single uninhibited pathway alone was sufficient to prevent apoptosis. In contrast to the results with adherent cells, although MSP also prevented apoptosis of cells in suspension (anoikis), its effect was mediated only by the PI3-K/AKT pathway. Despite activation of MAPK by MSP, anoikis was not prevented in suspended cells with a blocked PI3-K/AKT pathway. Thus, activation of MAPK alone is not sufficient to mediate MSP antiapoptotic effects. Cell adhesion generates an additional signal, which is essential for MSP to use MAPK in an antiapoptotic pathway. This may involve translocation of MSP-activated MAPK from the cytoplasm into the nucleus, which occurs only in adherent cells. Our results suggest that there is cross talk between cell matrix adhesion and growth factors in the regulation of cell survival via the MAPK pathway. Growth factors induce MAPK activation, and adhesion mediates MAPK translocation from the cytoplasm into the nucleus.

Oncogenic mutants of RON and MET receptor tyrosine kinases cause activation of the beta-catenin pathway.

beta-Catenin is an oncogenic protein involved in regulation of cell-cell adhesion and gene expression. Accumulation of cellular beta-catenin occurs in many types of human cancers. Four mechanisms are known to cause increases in beta-catenin: mutations of beta-catenin, adenomatous polyposis coli, or axin genes and activation of Wnt signaling. We report a new cause of beta-catenin accumulation involving oncogenic mutants of RON and MET receptor tyrosine kinases (RTKs). Cells transfected with oncogenic RON or MET were characterized by beta-catenin tyrosine phosphorylation and accumulation; constitutive activation of a Tcf transcriptional factor; and increased levels of beta-catenin/Tcf target oncogene proteins c-myc and cyclin D1. Interference with the beta-catenin pathway reduced the transforming potential of mutated RON and MET. Activation of beta-catenin by oncogenic RON and MET constitutes a new pathway, which might lead to cell transformation by these and other mutant growth factor RTKs.

Suppression of tumor growth in strain 2 guinea pigs by xenogeneic antitumor antibody.

Line 10 hepatoma cells were incubated with antiserum specific against line 10 cells (RaL10) and then tested for growth in syngeneic Wright strain 2 guinea pigs. Palpable tumors appeared in only 11 of 23 animals inoculated id with 10(5) RaL10-treated tumor cells, compared with an incidence of 21 of 23 for nonimmune rabbit serum (NRS)-treated cells and 23 of 23 for cells treated with syngeneic guinea pig serum. Animals inoculated with RaL10-treated tumor cells did not develop systemic tumor immunity. The long-term survival of guinea pigs receiving RaL10-treated tumor cells iv was 6 of 12 with a dose of 10(5) cells and 8 of 11 with 10(4) cells. None of the animals receiving 10(4) or 10(5) control tumor cells treated with NRS survived. RaL10 antiserum was not toxic to line 10 tumor cells in vitro, but mediated tumor-specific cytolytic reactions in the presence of fresh guinea pig serum or on the addition of peritoneal exudate cells from nonimmunized syngeneic guinea pigs.

Production and characterization of human glioma cell-derived monocyte chemotactic factor.

Since infiltration of monocytes into tumors may be mediated by tumor-derived chemoattractants, we characterized the monocyte-chemotactic activity (MCA) produced by glioma cell lines. The amount of MCA in the culture fluid of five lines tested differed by a factor of 25. U-105MG, the best producer, was selected for further study. After cells reached confluence and the medium was changed, MCA was detected by day 3 and remained at comparable levels on days 4 and 5. The molecular mass of MCA was approximately 17 kilodaltons, and the estimated isoelectric point ranged between pI 7 and pI 9. Because of the high constitutive production of MCA by U-105MG, sufficient material can be obtained for complete chemical characterization of this mediator of inflammation.

Extraction of tumor-specific antigen from cells and plasma membranes of line-10 hepatoma.

Tumor-specific antigen was extracted with 3 M KCl from line-10 guinea pig hepatoma cells. The yield of antigenic activity, estimated by production of delayed cutaneous hypersensitivity reactions in line-10 immune guinea pigs, was 10-30% of the antigen present in intact cells. By ultracentrifugation criteria, the extracted antigen was soluble. Gel filtration, ion exchange chromatography, and salting-out studies showed that the antigen was heterogeneous in size and net charge. The possibility that 3 M KCl extracted a homogeneous population of molecules associating into polymers of various sizes at low ionic strength was ruled out by heterogeneity on Sephadex G-200 chromatography at high ionic strength. After osmotic lysis of sucrose-loaded line-10 cells, whole plasma membranes or large membrane fragments were obtained in a yield of about 20%. The isolation procedure did not cause detectable loss of membrane antigenic activity. The membranes had 33 skin test U/mg membrane protein, compared to the intact cell value of 1.7 skin test U/mg cell protein. Extracts of plasma membranes had 10-20% of the antigenic activity of the starting membrane material. In contrast to the wide variety of proteins liberated from intact cells, much of the protein extracted from the membranes was in the molecular weight range above 250,000.

Protective tumor immunity induced by potassium chloride extracts of guinea pig hepatomas.

Tumor-specific antigens of cells of the diethylnitrosamine-induced hepatomas in strain-2 guinea pigs were extracted with 3 M KCl. Immunization of normal animals with the extracted tumor antigens in adjuvant protected them against a subsequent challenge with viable tumor cells. Extracted tumor-specific antigens were less effective immunogens than viable tumor cells for both of two antigenically distinct lines.

Interaction of BCG-activated macrophages with neoplastic and nonneoplastic cell lines in vitro : quantitation of the cytotoxic reaction by release of tritiated thymidine from prelabeled target cells.

Peritoneal cells from mice infected ip with Mycobacterium bovis, strain BCG, were cytotoxic to syngeneic tumor cells in vitro. Cytotoxicity was estimated by measurement of release of tritiated-thymidine (3-H-TDR) from prelabeled target cells. The cell responsible for tumor cytotoxicity was the macrophage. Macrophages from uninfected mice or from oil-, starch-, or thioglycollate-induced peritoneal exudates had little effect on labeled tumor monolayers. Tumoricidal macrophages were present at 3-7 days and persisted through 6 weeks after a single BCG injection. Two neoplastic/nonneoplastic cell-line pairs, all four of the cell lines derived from a cloned syngeneic embryo cell line, were used as target cells for BCG-activated macrophages. Both tumor cell lines released significantly more 3-H-TDR than did the two nonneoplastic lines. In a mixed neoplastic/nonneoplastic target cell population, BCG-activated macrophages selectively destroyed the neoplastic cells; nonneoplastic cells were not affected as "innocent bystanders".

Quantitative study of monocyte chemoattractant protein-1 (MCP-1) in cerebrospinal fluid and cyst fluid from patients with malignant glioma.

BACKGROUND: Monocyte chemoattractant protein-1 (MCP-1) is a 76-amino acid protein that attracts monocytes. In vitro studies have reported high levels of MCP-1 messenger RNA expression, as well as the presence of MCP-1, in malignant glioma cells. PURPOSE: Our purpose was to determine whether an MCP-1 assay could be used in a clinical setting 1) to differentiate malignant from benign gliomas and from nontumor disorders of the central nervous system and 2) to detect subarachnoid dissemination of glioma cells. METHODS: MCP-1 levels in cerebrospinal fluid (CSF) and cyst fluid were measured with a sandwich enzyme-linked immunosorbent assay (ELISA) that we had previously developed. We measured MCP-1 levels in CSF samples from 19 patients with malignant glioma (glioblastoma, 10; anaplastic astrocytoma, six; anaplastic oligodendroglioma, two; and ependymoblastoma, one), nine patients with benign glioma, and seven patients with nontumor disorders of the central nervous system. Cyst fluids from four patients with malignant glioma (anaplastic astrocytoma) were also tested. The correlation between MCP-1 concentration in the CSF and subarachnoid dissemination of malignant glioma cells was also studied. RESULTS: The MCP-1 concentration (mean +/- SE) in CSF samples from patients with malignant glioma (2.3 +/- 0.4 ng/mL) was significantly higher than that from patients with benign glioma (0.6 +/- 0.1 ng/mL) (P < .01) or from patients with no tumor (0.5 +/- 0.1 ng/mL) (P < .01). Furthermore, CSF samples from patients with subarachnoid dissemination of malignant glioma contained significantly higher amounts of MCP-1 than those from patients without dissemination (P < .05). Cyst fluids from four of the patients with malignant glioma contained high concentrations of MCP-1. CONCLUSIONS: These results indicate that MCP-1 is produced by malignant glioma in vivo as well as in vitro and suggest that testing for MCP-1 in CSF may be useful in the clinic to differentiate malignant glioma from benign glioma and to detect subarachnoid dissemination of the tumor cells. IMPLICATIONS: The MCP-1 ELISA in CSF may lead to more accurate diagnosis of malignant glioma and detection of subarachnoid dissemination of tumor cells, facilitating selection of patients with these conditions for appropriate therapy.

Characterization of macrophage chemotaxins in tumor cell cultures and comparison with lymphocyte-derived chemotactic factors.

Culture fluids from five murine sarcomas were chemotactic for syngeneic peritoneal macrophages in vitro. Peritoneal macrophages from mice infected with Mycobacterium bovis, strain Bacillus Calmette-Guérin, were more responsive to the chemotactic factor in tumor cultures than were normal macrophages. Peritoneal granulocytes, however, did not significantly respond to this factor. The level of chemotactic activity in tumor cultures paralleled cell growth for all five tumors; maximal levels occurred during log growth. Culture medium alone or fluids from proliferating spleen cell cultures stimulated with mitogens did not have detectable chemotactic activity. Chromatography of the tumor culture fluids resulted in a single peak of chemotactic activity in the 15,000-molecular weight range on Sephadex G-100 and at about 7.5 mmho/cm specific conductance on diethylaminoethyl cellulose. By both biological and physicochemical characteristics, the chemotactic activity in tumor culture fluids was different from mouse lymphocyte-derived chemotactic factor.

Requirement of phosphatidylinositol-3 kinase for epithelial cell migration activated by human macrophage stimulating protein.

Macrophage stimulating protein (MSP) is a ligand for the RON receptor protein tyrosine kinase. Activation of RON in murine resident macrophages results in cell shape change and migration. We studied cell movement induced by MSP in different types of human epithelial cells and the possible role of phosphatidylinositol-3 (PI-3) kinase in RON-mediated signal transduction. We observed specific and saturable binding of 125I-MSP to RON on several epithelial cell lines. In addition to activation and phosphorylation of RON, MSP also induced tyrosine phosphorylation of the PI-3 kinase p85 subunit in a time-dependent manner, with a peak at 15 min. Moreover, phosphorylated RON formed a complex with PI-3 kinase in both HK-NOC keratinocyte and RON cDNA-transfected MDCK cells. An in vitro protein interaction assay confirmed that PI-3 kinase from a lysate of MSP-activated cells bound to pure RON protein. MSP, at a concentration range of 1 to 5 nM, induced migration of three epithelial cell lines. This effect was inhibited by wortmannin, a specific inhibitor for PI-3 kinase, with an IC50 of 10 nM. MSP-induced shape change in murine resident peritoneal macrophages was also abolished by wortmannin. These data suggest that activation of PI-3 kinase is required for MSP-induced epithelial cell migration. The stimulation by MSP of epithelial cell movement may have implications for tissue repair, wound healing, and tumor metastasis.

Hepatic catabolism of intravenously administered pro-macrophage-stimulating protein in mice.

We injected 125I-pro-macrophage-stimulating protein (pro-MSP) intravenously into normal mice to determine its clearance from the circulation and to test for conversion of pro-MSP to the biologically active heterodimer in the absence of inflammation or tissue injury. Pro-MSP was cleared from the circulation with a half-life of approximately 100 min. This rapid clearance was not peculiar to 125I-pro-MSP, since clearance rates of unlabeled pro-MSP and of 125I-bovine serum albumin were comparable. The liver was the major locus of radioactivity 10-20 min after the intravenous injection of 125I-pro-MSP. By 90 min, over 60% of total recovered radioactivity was in the small intestine. Reflecting gastrointestinal transit, counts decreased in the small intestine and appeared in the colon by 180 min. Essentially all counts in urine and feces obtained at later times were soluble in trichloracetic acid. These findings reflected rapid hepatic proteolysis of pro-MSP to fragments undetectable by antibody to pro-MSP; within 20 min after intravenous administration, immunoprecipitable counts were only 22% of the total liver extract radioactivity. Comparison of sodium dodecyl sulfate-polyacrylamide gel electrophoresis and radioautography data for immunoprecipitated plasma and liver extract revealed no evidence for hepatic conversion of pro-MSP to MSP. Thus, the hepatic catabolic pathway of pro-MSP is degradative and does not yield mature MSP. The results support our view that MSP is not released into the circulation but is generated at specific extravascular loci by pro-MSP convertases.

Kinases involved in MSP/RON signaling.

Macrophage stimulating protein (MSP) belongs to the plasminogen-related kringle domain family. In addition to stimulation of macrophages, MSP acts on other cell types including epithelial and hematopoietic cells. The MSP receptor is a transmembrane tyrosine kinase called RON in humans and STK in mice. MSP/receptor interaction induces activation of signal transduction pathways that mediate MSP biological activities. Cytoplasmic kinases are intracellular messengers occupying an important role in signal transduction. We have identified kinases that participate in RON signaling. In addition to previously identified involvement of phosphatidylinositol 3-kinase (PI3-K), JNK, and MAPK, we found that FAK, c-Src, and AKT are rapidly and transiently activated by MSP. FAK, MAPK, and c-Src are involved in MSP-induced cell proliferation. MAPK and c-Src are components of one signal transduction cascade, and MAPK is downstream of c-Src. FAK also regulates MSP-induced cell growth, but via a path different from c-Src/MAPK. AKT kinase is a component of a separate branch of the RON/PI3-K pathway that mediates the MSP anti-apoptotic effect on epithelial cells. PI3-K regulates MSP-induced adhesion and motility but via downstream components different from AKT. Thus, occupancy of the RON receptor by MSP activates distinct signal transduction pathways that mediate several cellular responses.