Connective tissue growth factor is a positive regulator of epithelial–mesenchymal transition and promotes the adhesion with gastric cancer cells in human peritoneal mesothelial cells

Connective tissue growth factor (CTGF) is involved in human cancer development and progression. Epithelial to mesenchymal transition (EMT) plays an important role in many biological processes. In this study, we wished to investigate the role of CTGF in EMT of peritoneal mesothelial cells and the effects of CTGF on adhesion of gastric cancer cells to mesothelial cells. Human peritoneal mesothelial cells (HPMCs) were cultured with TGF-β₁ or various concentrations of CTGF for different time. The EMT process was monitored by morphology. Real-time RT-PCR and Western blot were used to evaluate the expression of vimentin, α-SMA , E-cadherin and β-catenin. RNA interference was used to achieve selective and specific knockdown of CTGF. We demonstrated that CTGF induced EMT of mesothelial cells in a dose- and time-dependent manner. HPMCs were exposed to TGF-β₁ also underwent EMT which was associated with the induction of CTGF expression. Transfection with CTGF siRNA was able to reverse the EMT partially after treatment of TGF-β₁. Moreover, the induced EMT of HPMCs was associated with an increased adhesion of gastric cancer cells to mesothelial cells. These findings suggest that CTGF is not only an important mediator but a potent activator of EMT in peritoneal mesothelial cells, which in turn promotes gastric cancer cell adhesion to peritoneum.     

ICAM-1 mediated tumor-mesothelial cell adhesion is modulated by IL-6 and TNF-alpha: a potential mechanism by which surgical trauma increases peritoneal metastases

Peritoneal metastases frequently occur in different gastrointestinal cancers and have a poor prognosis. It is known that surgical injury promotes tumor growth and local recurrence rates and also the degree of surgical trauma correlated with the amount of tumor implantation into the peritoneum. The mechanism that mediates tumor cell adhesion to the mesothelium is not fully understood. This study investigates the role of ICAM-1, an important mediator of trans-mesothelial leucocyte migration, in tumor-mesothelial interactions as the initial step in the development of peritoneal recurrence using an in vitro model incorporating mesothelial cell monolayer derived from omental samples. We also investigate how the cytokines interleukins 6 (IL-6) and tumor necrosis factor alpha (TNF-alpha) modulate this process. We demonstrate that ICAM-1 blockade reduces the ability of both pancreatic and colonic cancer cell lines to adhere to the mesothelium. Preincubation of the mesothelial cell monolayer with either IL-6 or TNF-alpha enhances tumor cell adhesion, and this is associated with an increased expression of ICAM-1. Mesothelial CD44 expression, which has previously been implicated in this process, was unaffected by these cytokines. The use of an inhibitory monoclonal antibody against ICAM-1 attenuated the enhanced adhesion mediated by IL-6 or TNF-alpha. This study suggests that mesothelial ICAM-1 plays a role in the adhesion of tumor cells to the peritoneum in the development of peritoneal metastases.     

In vitro Mesothelial Clearance Assay that Models the Early Steps of Ovarian Cancer Metastasis

Ovarian cancer is the fifth leading cause of cancer related deaths in the United States¹. Despite a positive initial response to therapies, 70 to 90 percent of women with ovarian cancer develop new metastases, and the recurrence is often fatal². It is, therefore, necessary to understand how secondary metastases arise in order to develop better treatments for intermediate and late stage ovarian cancer. Ovarian cancer metastasis occurs when malignant cells detach from the primary tumor site and disseminate throughout the peritoneal cavity. The disseminated cells can form multicellular clusters, or spheroids, that will either remain unattached, or implant onto organs within the peritoneal cavity³ (Figure 1, Movie 1). All of the organs within the peritoneal cavity are lined with a single, continuous, layer of mesothelial cells^4-6 (Figure 2). However, mesothelial cells are absent from underneath peritoneal tumor masses, as revealed by electron micrograph studies of excised human tumor tissue sections^3,5-7 (Figure 2). This suggests that mesothelial cells are excluded from underneath the tumor mass by an unknown process. Previous in vitro experiments demonstrated that primary ovarian cancer cells attach more efficiently to extracellular matrix than to mesothelial cells⁸, and more recent studies showed that primary peritoneal mesothelial cells actually provide a barrier to ovarian cancer cell adhesion and invasion (as compared to adhesion and invasion on substrates that were not covered with mesothelial cells)^9,10. This would suggest that mesothelial cells act as a barrier against ovarian cancer metastasis. The cellular and molecular mechanisms by which ovarian cancer cells breach this barrier, and exclude the mesothelium have, until recently, remained unknown. Here we describe the methodology for an in vitro assay that models the interaction between ovarian cancer cell spheroids and mesothelial cells in vivo (Figure 3, Movie 2). Our protocol was adapted from previously described methods for analyzing ovarian tumor cell interactions with mesothelial monolayers^8-16, and was first described in a report showing that ovarian tumor cells utilize an integrin –dependent activation of myosin and traction force to promote the exclusion of the mesothelial cells from under a tumor spheroid¹⁷. This model takes advantage of time-lapse fluorescence microscopy to monitor the two cell populations in real time, providing spatial and temporal information on the interaction. The ovarian cancer cells express red fluorescent protein (RFP) while the mesothelial cells express green fluorescent protein (GFP). RFP-expressing ovarian cancer cell spheroids attach to the GFP-expressing mesothelial monolayer. The spheroids spread, invade, and force the mesothelial cells aside creating a hole in the monolayer. This hole is visualized as the negative space (black) in the GFP image. The area of the hole can then be measured to quantitatively analyze differences in clearance activity between control and experimental populations of ovarian cancer and/ or mesothelial cells. This assay requires only a small number of ovarian cancer cells (100 cells per spheroid X 20-30 spheroids per condition), so it is feasible to perform this assay using precious primary tumor cell samples. Furthermore, this assay can be easily adapted for high throughput screening.     

The Influence of Reactive Oxygen Species on the Adhesion of Pancreatic Carcinoma Cells to the Peritoneum

Postoperative peritoneal carcinomatosis is a significant clinical problem after “curative” resection of pancreatic carcinoma. Peroperative surgical trauma activates a cascade of peritoneal defense mechanisms responsible for postoperative intra-abdominal tumor recurrence. Reactive oxygen species (ROS) play a pivotal role in this postoperative inflammatory reaction. This study explores the influence of ROS on adhesion of human pancreatic carcinoma cells to human mesothelial cells. Furthermore this study explores the influence of ROS on the presentation of adhesion molecules on Panc-1 and mesothelial cells. ROS were produced using the enzymatic reaction of xanthine with xanthine oxidase (X/XO). A reproducible in-vitro assay to study adhesion of human Panc-1 carcinoma tumor cells to a mesothelial cell monolayer of primary human mesothelial cells was used. Mesothelial monolayers were incubated with ROS produced prior to adhesion of the tumor cells. Incubation of the mesothelial cells with X/XO resulted in a significant increase (69.5%) in adhesion of Panc-1 in all patients. SOD/catalase, anti-oxidants, could reduce this increase by 56.7%. ROS significantly influenced the expression of the adhesion molecules ICAM-1, VCAM-1 and CD44h on mesothelial cells, but did not influence adhesion molecule expression on Panc-1. The ROS released during the post-operative inflammatory reaction may play an important role in the adhesion of pancreatic tumor cells to the mesothelium. Possibly by influencing adhesion molecule expression on mesothelial cells. Therefore ROS can partly be responsible for the enhanced post-operative intra-abdominal tumor recurrence.     

Mesothelial cells differentiate into fibroblast-like cells under the scirrhous gastric cancer microenvironment and promote peritoneal carcinomatosis in vitro and in vivo

Peritoneal metastases are one reason for the poor prognosis of scirrhous gastric cancer (SGC), and myofibroblast provides a favorable environment for the peritoneal dissemination of gastric cancer. The aim of this study was to determine whether myofibroblast originates from peritoneal mesothelial cells under the influence of the tumor microenvironment. Immunohistochemical studies of peritoneal biopsy specimens from patients with peritoneal lavage cytological (+) status demonstrate the expression of the epithelial markers cytokeratin in fibroblast-like cells entrapped in the stroma, suggesting that these cells stemmed from local conversion of mesothelial cells. To confirm this hypothesis in vitro, we co-incubated mesothelial cells with SGC or non-SGC to investigate morphology and function changes. As we expected, mesothelial cells undergo a transition from an epithelial phenotype to a mesenchymal phenotype with loss of epithelial morphology and decrease in the expression of cytokeratin and E-cadherin when exposed to conditioned medium from HSC-39, and the induction of mesothelial cells can be abolished using a neutralizing antibody to transforming growth factor-beta1 (TGF-β1) as well as by pre-treatment with SB431542. Moreover, we found that these mesothelial cells-derived cells exhibit functional properties of myofibroblasts, including the ability to increase adhesion and invasion of SGC. In summary, our current data demonstrated that mesothelial cells are a source of myofibroblasts under the SGC microenvironment which provide a favorable environment for the dissemination of gastric cancer; TGF-β1 produced by autocrine/paracrine in peritoneal cavity may play a central role in this pathogenesis.     

The Influence of Reactive Oxygen Species on the Adhesion of Pancreatic Carcinoma Cells to the Peritoneum

Postoperative peritoneal carcinomatosis is a significant clinical problem after “curative” resection of pancreatic carcinoma. Preoperative surgical trauma activates a cascade of peritoneal defense mechanisms responsible for postoperative intra-abdominal tumor recurrence. Reactive oxygen species (ROS) play a pivotal role in this postoperative inflammatory reaction. This study explores the influence of ROS on adhesion of human pancreatic carcinoma cells to human mesothelial cells. Furthermore this study explores the influence of ROS on the presentation of adhesion molecules on Panc-1 and mesothelial cells. ROS were produced using the enzymatic reaction of xanthine with xanthine oxidase (X/XO). A reproducible in vitro assay to study adhesion of human Panc-1 carcinoma tumor cells to a mesothelial cell monolayer of primary human mesothelial cells was used. Mesothelial monolayers were incubated with ROS produced prior to adhesion of the tumor cells. Incubation of the mesothelial cells with X/XO resulted in a significant increase (69.5%) in adhesion of Panc-1 in all patients. SOD/catalase, anti-oxidants, could reduce this increase by 56.7%. ROS significantly influenced the expression of the adhesion molecules ICAM-1, VCAM-1 and CD44h on mesothelial cells, but did not influence adhesion molecule expression on Panc-1. The ROS released during the post-operative inflammatory reaction may play an important role in the adhesion of pancreatic tumor cells to the mesothelium-possibly by influencing adhesion molecule expression on mesothelial cells. Therefore ROS can partly be responsible for the enhanced post-operative intra-abdominal tumor recurrence.Key words: reactive oxygen species, mesothelium, Panc-1     

Interactions of human ovarian tumor cells with human mesothelial cells grown on extracellular matrix : An in vitro model system for studying tumor cell adhesion and invasion

Human ovarian tumors metastasize by direct extension into the peritoneal cavity leading to tumor cell implantation onto peritoneal surfaces. Successful formation of peritoneal implants is dependent on the ability of ascitic tumor cells to infiltrate the mesothelium, and become firmly adherent to the underlying extracellular matrix (ECM). In order to investigate this process in more detail, an in vitro model system was developed employing human mesothelial cells grown on ECM-coated culture dishes. The ability of human ovarian carcinoma cells derived from ascitic fluid to attach to the mesothelial cell monolayer grown on ECM, ECM alone or plastic was quantitated with the use of 51Cr radio-labelled tumor cells. Tumor cells exhibited a more rapid and firmer attachment to ECM than to the mesothelial cells or to plastic. Using agitation to stimulate peritoneal fluid dynamics and shear forces in vivo, tumor cell arrest was found to be limited to the ECM, but it occurred at a slower rate than it did without agitation. Tumor cell attachment was also restricted to areas of exposed ECM in wounded mesothelium as assessed by phase-contrast microscopy. Morphologic alterations of the mesothelium induced by tumor cells were observed with the use of scanning electron microscopy (SEM) and immunohistochemical staining which included disruption of intercellular junctions leading to retraction of mesothelial cells, exposure of underlying ECM, subsequent attachment and proliferation on ECM. This model system would appear to be useful for elucidating mechanisms of ovarian tumor cell adhesion and proliferation, and for assessing various therapeutic modalities for their ability to block tumor cell implantation, invasion and growth on peritoneal surfaces.     

Role of Integrins and Evidence for two Distinct Mechanisms Mediating Human Colorectal Carcinoma Cell Interaction with Peritoneal Mesothelial Cells and Extracellular Matrix

Peritoneal carcinomatosis involves a series of events including tumor cell interactions with mesothelial cells and the extracellular matrix (ECM). We have studied the adhesive and invasive properties of four human colorectal carcinoma cell lines (Co115, HT29, SW480, SW620) confronted in vitro with a human mesothelial cell monolayer or with the ECM proteins collagen IV, laminin-1, fibronectin, tenascin-C and vitronectin. Quantitation was achieved following staining of tumor cells with the calcein-AM fluorescent dye. We found that all four cell lines rapidly adhered to a mesothelial cell monolayer. This adhesion event was not inhibitable by anti-integrin and anti-CD44 antibodies. Following initial attachment, the SW480 and SW620 cells invaded the mesothelial cell monolayer more aggressively than HT29 and Col 15 cells. All cell lines adhered to ECM proteins with each one exhibiting an individual adhesion pattern. Adhesion to matrix was completely integrin-dependent. When tested in an invasion assay, HT29 and Co115 cells crossed Matrigel-coated filters while SW480 and SW620 cells did not. This invasion was inhibited by anti-β1 integrin antibodies. Taken together, our results demonstrate that the initial colorectal tumor cell—mesothelial cell interaction occurs through an integrin-independent mechanism while adhesion to matrix proteins and invasion through Matrigel are integrin-dependent events. Furthermore, the different invasive capacity of SW480 and SW620 versus HT29 and Co115 cells upon interaction with a mesothelial cell monolayer or Matrigel suggests that these two invasion events may be mediated by distinct mechanisms.     

Role of Integrins and Evidence for two Distinct Mechanisms Mediating Human Colorectal Carcinoma Cell Interaction with Peritoneal Mesothelial Cells and Extracellular Matrix

Peritoneal carcinomatosis involves a series of events including tumor cell interactions with mesothelial cells and the extracellular matrix (ECM). We have studied the adhesive and invasive properties of four human colorectal carcinoma cell lines (Co115, HT29, SW480, SW620) confronted in vitro with a human mesothelial cell monolayer or with the ECM proteins collagen IV, laminin-1, fibronectin, tenascin-C and vitronectin. Quantitation was achieved following staining of tumor cells with the calcein-AM fluorescent dye. We found that all four cell lines rapidly adhered to a mesothelial cell monolayer. This adhesion event was not inhibitable by anti-integrin and anti-CD44 antibodies. Following initial attachment, the SW480 and SW620 cells invaded the mesothelial cell monolayer more aggressively than HT29 and Col 15 cells. All cell lines adhered to ECM proteins with each one exhibiting an individual adhesion pattern. Adhesion to matrix was completely integrin-dependent. When tested in an invasion assay, HT29 and Co115 cells crossed Matrigel-coated filters while SW480 and SW620 cells did not. This invasion was inhibited by anti-β1 integrin antibodies. Taken together, our results demonstrate that the initial colorectal tumor cell—mesothelial cell interaction occurs through an integrin-independent mechanism while adhesion to matrix proteins and invasion through Matrigel are integrin-dependent events. Furthermore, the different invasive capacity of SW480 and SW620 versus HT29 and Co115 cells upon interaction with a mesothelial cell monolayer or Matrigel suggests that these two invasion events may be mediated by distinct mechanisms.     

Modeling the Early Steps of Ovarian Cancer Dissemination in an Organotypic Culture of the Human Peritoneal Cavity

The pattern of ovarian cancer metastasis is markedly different from that of most other epithelial tumors, because it rarely spreads hematogenously. Instead, ovarian cancer cells exfoliated from the primary tumor are carried by peritoneal fluid to metastatic sites within the peritoneal cavity. These sites, most notably the abdominal peritoneum and omentum, are organs covered by a mesothelium-lined surface. To investigate the processes of ovarian cancer dissemination, we assembled a complex three-dimensional culture system that reconstructs the lining of the peritoneal cavity in vitro. Primary human fibroblasts and mesothelial cells were isolated from human omentum. The fibroblasts were then mixed with extracellular matrix and covered with a layer of the primary human mesothelial cells to mimic the peritoneal and omental surfaces encountered by metastasizing ovarian cancer cells. The resulting organotypic model is, as shown, used to examine the early steps of ovarian cancer dissemination, including cancer cell adhesion, invasion, and proliferation. This model has been used in a number of studies to investigate the role of the microenvironment (cellular and acellular) in early ovarian cancer dissemination. It has also been successfully adapted to high throughput screening and used to identify and test inhibitors of ovarian cancer metastasis.     

Assessment of Ovarian Cancer Spheroid Attachment and Invasion of Mesothelial Cells in Real Time

Ovarian cancers metastasize by shedding into the peritoneal fluid and dispersing to distal sites within the peritoneum. Monolayer cultures do not accurately model the behaviors of cancer cells within a nonadherent environment, as cancer cells inherently aggregate into multicellular structures which contribute to the metastatic process by attaching to and invading the peritoneal lining to form secondary tumors. To model this important stage of ovarian cancer metastasis, multicellular aggregates, or spheroids, can be generated from established ovarian cancer cell lines maintained under nonadherent conditions. To mimic the peritoneal microenvironment encountered by tumor cells in vivo, a spheroid-mesothelial co-culture model was established in which preformed spheroids are plated on top of a human mesothelial cell monolayer, formed over an extracellular matrix barrier. Methods were then developed using a real-time cell analyzer to conduct quantitative real time measurements of the invasive capacity of different ovarian cancer cell lines grown as spheroids. This approach allows for the continuous measurement of invasion over long periods of time, which has several advantages over traditional endpoint assays and more laborious real time microscopy image analyses. In short, this method enables a rapid, determination of factors which regulate the interactions between ovarian cancer spheroid cells invading through mesothelial and matrix barriers over time.     

Mesothelial vitronectin stimulates migration of ovarian cancer cells

Ovarian carcinomas, the most fatal gynaecological malignancies, are associated with poor prognosis predominantly because of a high recurrence rate. Ovarian cancer cells spread widely throughout the abdominal cavity leading to peritoneal metastasis. The influence of the mesothelial microenvironment on the biological mechanisms leading to cancer cell colonization of the mesothelium is poorly understood. This study aims to investigate whether mesothelial secretions affect the migration of ovarian cancer cells and focuses on the role of the adhesive molecule Vn (vitronectin) and its integrin receptors. An in vitro co‐culture model indicated that clusters of IGROV1 and SKOV3 cells adhere to MeT‐5A mesothelial cells preferentially at intercellular sites, invade the mesothelial monolayer and alter the integrity of the mesothelium. In addition, mesothelial CM (cell‐conditioned medium) induces migration of IGROV1 and SKOV3 cells in Boyden chambers and wound healing assays. Furthermore, blocking molecules directed against vitronectin or its αv integrin receptor decrease mesothelial‐CM‐induced migration by approximately 40% and 60–70% for IGROV1 and SKOV3 ovarian cancer cells, respectively, in Boyden chamber assays. Wound healing assays that allow cell migration to be measured over 24 h periods demonstrated that blocking molecules prevent the migration of IGROV1 and SKOV3 cells. Vitronectin is present in CM MeT‐5A (mesothelial conditioned medium) and in metastatic peritoneal tissue sections. The expression of vitronectin at the periphery of mesothelial cells and within ovarian cancer cell clusters suggests a potential role for this molecule during intraperitoneal implantation of ovarian cancer cells. Vitronectin could represent a target for the development of anti‐adhesive strategies to impede ovarian cancer dissemination.     

Synthesis and biological evaluation of sulfonyl acrylonitriles as novel inhibitors to peritoneal carcinomatosis

The vast majority of cancer patients die from metastasis, the process by which cancer cells spread to secondary tissues through body fluids. Peritoneal carcinomatosis is a type of metastasis in which cancer cells gain access to the intra-abdominal cavity and then implant in the peritoneum, the thin tissue that lines the abdominal wall and internal organs. Unfortunately, peritoneal carcinomatosis can occur following surgical resection of intra-abdominal malignancies. We previously reported proapoptotic activity of (2E)-3-[[4-(1,1-dimethylethyl)phenyl]sulfonyl]-2-propenenitrile (BAY 11-7085, 1) on colon and pancreatic cancer cells during adhesion and demonstrated that this compound could significantly inhibit peritoneal carcinomatosis in mice.(1,2) In order to determine the chemical basis of the anti-metastatic properties of BAY 11-7085, a series of analogs were synthesized and evaluated for their ability to induce apoptosis in pancreatic and ovarian cancer cells during adhesion to mesothelial cells, which line the surface of the peritoneum. The co-culture assay results were validated using a murine peritoneal carcinomatosis model. These analogs may greatly benefit patients undergoing surgical resections of colorectal, pancreatic, and ovarian cancers depending on their tolerability.     

Possible involvement of SDF-1α/CXCR4-DPPIV axis in TGF-β1-induced enhancement of migratory potential in human peritoneal mesothelial cells

We have previously reported that human peritoneal mesothelial cells (HPMCs) express a large amount of dipeptidyl peptidase IV (DPPIV) and that its expression is regulated by a variety of bioactive substances in malignant ascites from ovarian cancer patients. The aim of this study has been to examine the expression and role of the SDF-1α/CXCR4-DPPIV axis in HPMCs. We have demonstrated that the expression levels of DPPIV and E-cadherin in HPMCs decrease, following TGF-β1-induced morphological change, in a time- and concentration-dependent manner. Additionally, we show that both SDF-1α (a chemokine and substrate for DPPIV) and its receptor, CXCR4, are expressed on HPMCs, and that their expression levels are upregulated by TGF-β1 treatment, resulting in an increased migratory potential of HPMCs. Furthermore, the migratory potential of HPMCs is significantly enhanced in the presence of SDF-1α or DPPIV-specific inhibitor in the wound-healing assay. These results suggest that DPPIV and SDF-1α/CXCR4 play crucial roles in regulating the migratory potential of HPMCs, which may be involved in the re-epithelialization of denuded basement membrane at the site of peritoneal injury.     

Expression of matrix metalloproteinases-2 and -9 by cells isolated from the peritoneal fluid of women with ovarian carcinoma

OBJECTIVE: To investigate the level of expression of matrix metalloproteinases (MMP)-2 and -9 by cells isolated from the peritoneal fluid of women with ovarian carcinoma. STUDY DESIGN: Tumor tissue specimens and cells isolated from peritoneal fluid from 20 patients with epithelial ovarian carcinoma were examined for MMP-2 and -9 expression using immunostaining. Six benign peritoneal effusions containing mesothelial cells were also included in the study. RESULTS: Expression of both MMP-2 and -9 was noted in cancer cells in peritoneal fluid of all cases studied. Peritoneal fluid cancer cells showed increased expression of both MMP-2 and -9 relative to mesothelial cell expression of these MMPs. Positive immunoreactivity of these MMPs in primary tumor tissues was confirmed by immunohistochemistry. CONCLUSION: Our findings suggest that both MMP-2 and -9 are frequently overexpressed in ovarian cancer cells disseminated in the peritoneal cavity and that determination of cellular MMP-2 and -9 expression could be useful in distinguishing cancer cells from mesothelial cells in peritoneal fluid cytologic specimens from women with ovarian epithelial carcinoma.     

Nanotube action between human mesothelial cells reveals novel aspects of inflammatory responses

A well-known role of human peritoneal mesothelial cells (HPMCs), the resident cells of the peritoneal cavity, is the generation of an immune response during peritonitis by activation of T-cells via antigen presentation. Recent findings have shown that intercellular nanotubes (NTs) mediate functional connectivity between various cell types including immune cells - such as T-cells, natural killer (NK) cells or macrophages - by facilitating a spectrum of long range cell-cell interactions. Although of medical interest, the relevance of NT-related findings for human medical conditions and treatment, e.g. in relation to inflammatory processes, remains elusive, particularly due to a lack of appropriate in vivo data. Here, we show for the first time that primary cultures of patient derived HPMCs are functionally connected via membranous nanotubes. NT formation appears to be actin cytoskeleton dependent, mediated by the action of filopodia. Importantly, significant variances in NT numbers between different donors as a consequence of pathophysiological alterations were observable. Furthermore, we show that TNF-α induces nanotube formation and demonstrate a strong correlation of NT connectivity in accordance with the cellular cholesterol level and distribution, pointing to a complex involvement of NTs in inflammatory processes with potential impact for clinical treatment.     

A review of physiological and cellular mechanisms underlying fibrotic postoperative adhesion

Postoperative adhesions (PA) are fibrotic tissues that are the most common driver of long-term morbidity after abdominal and pelvic surgery. The optimal drug or material to prevent adhesion formation has not yet been discovered. Comprehensive understanding of cellular and molecular mechanisms of adhesion process stimulates the design of future anti-adhesive strategies. Recently, disruption of peritoneal mesothelial cells were suggested as the ''motor'' of PA formation, followed by a cascade of events (coagulation, inflammation, fibrinolysis) and influx of various immune cells, ultimately leading to a fibrous exudate. We showed that a variety of immune cells were recruited into adhesive peritoneal tissues in patients with small bowel obstruction caused by PA. The interactions among various types of immune cells contribute to PA development following peritoneal trauma. Our review focuses on the specific role of different immune cells in cellular and humoral mechanisms underpinning adhesion development.     

Impacts of icodextrin on integrin-mediated wound healing of peritoneal mesothelial cells

AimsExposure to glucose and its metabolites in peritoneal dialysis fluid (PDF) results in structural alterations of the peritoneal membrane. Icodextrin-containing PDF eliminates glucose and reduces deterioration of peritoneal membrane function, but direct effects of icodextrin molecules on peritoneal mesothelial cells have yet to be elucidated. We compared the impacts of icodextrin itself with those of glucose under PDF-free conditions on wound healing processes of injured mesothelial cell monolayers, focusing on integrin-mediated cell adhesion mechanisms.Main methodsRegeneration processes of the peritoneal mesothelial cell monolayer were investigated employing an in vitro wound healing assay of cultured rat peritoneal mesothelial cells treated with icodextrin powder- or glucose-dissolved culture medium without PDF, as well as icodextrin- or glucose-containing PDF. The effects of icodextrin on integrin-mediated cell adhesions were examined by immunocytochemistry and Western blotting against focal adhesion kinase (FAK).Key findingsCell migration over fibronectin was inhibited in conventional glucose-containing PDF, while icodextrin-containing PDF exerted no significant inhibitory effects. Culture medium containing 1.5% glucose without PDF also inhibited wound healing of mesothelial cells, while 7.5% icodextrin-dissolved culture medium without PDF had no inhibitory effects. Glucose suppressed cell motility by inhibiting tyrosine phosphorylation of FAK, formation of focal adhesions, and cell spreading, while icodextrin had no effects on any of these mesothelial cell functions.SignificanceOur results demonstrate icodextrin to have no adverse effects on wound healing processes of peritoneal mesothelial cells. Preservation of integrin-mediated cell adhesion might be one of the molecular mechanisms accounting for the superior biocompatibility of icodextrin-containing PDF.     

Malignant ascites of serous papillary ovarian adenocarcinoma. An immunocytochemical study of the tumor cells

In 17 malignant peritoneal effusions due to papillary serous adenocarcinoma of the ovary, the reaction patterns of the tumor cells to monoclonal antibodies (MAbs) against surface antigens were studied and compared with the reaction patterns of mesothelial cells in the same effusions. The following surface markers were used with the adhesive slide method: epithelial membrane antigen (EMA), human epithelium-specific cell surface antigen (HEA-125), human endothelial antigen (BMA-120), carcinoembryonic antigen (CEA 3-13), an antibody against natural killer cells and cytotoxic cells (BMA-070), granulocyte antigen (Leu M1) and leukocyte antigen of class I (HLA-1). In all cases, from 30% to 95% of the tumor cells reacted with EMA and HEA-125. Tumor cells showed a positive staining with CEA 3-13 in only five cases. In all cases, from 75% to 95% of the tumor cells reacted positively with BMA-120. The reactivity of a few mesothelial cells with EMA and of all mesothelial cells with BMA-120 did not interfere with the identification of positive tumor cells since the reaction patterns were different. Interestingly, our study demonstrated that BMA-070, an MAb identifying natural killer cells and cytotoxic cells, is also a most useful tumor marker. The same was found to be true for Leu M1, an MAb originally thought to react only with granulocytes. The tumor cells showed a partial or total loss of the expression of HLA-1 reactivity. Since all cases were immunocytochemically positive for tumor cells while conventional cytology was positive in only 13 of the cases, the immunocytochemical analysis of malignant peritoneal effusions due to papillary serous adenocarcinoma of the ovary seems able to improve the cytologic diagnosis of the fluids.