Peritoneal Cavity Of Mice Research Articles

Transplantation of human cells in the peritoneal cavity of immunodeficient mice for rapid assays of hepatitis B virus replication

Studies of natural hepatitis B virus infection must be restricted to humans or primates due to viral species-specificity. Alternative hepadnavirus animal models, e.g., woodchuck hepatitis virus in captive woodchucks, are not convenient, while in transgenic mice hepatitis B virus or viral proteins are expressed permanently through integrated genomes. Availability of small animal models that are easily produced and permit rapid assays will be quite helpful. We examined whether transplantation of human cells in the peritoneal cavity of mice will generate an appropriate mass of cells with hepatitis B virus replication. HepG2 2.2.15 cells were transplanted intraperitoneally into NOD/SCID mice. Replication of hepatitis B virus and viral gene expression was determined by analysis of blood and transplanted tissues with viral DNA and hepatitis B core antigen expression. Interruption of viral replication was examined. After intraperitoneal transplantation with microcarrier scaffolds, 2.2.15 cells engrafted and proliferated in the peritoneal cavity of NOD/SCID mice. Hepatitis B virus replicated in transplanted 2.2.15 cells as shown by hepatitis B core antigen expression. Moreover, viral particles were secreted into the blood. Hepatitis B virus replication was susceptible to conventional antiviral drug therapy, such as lamivudine, as well as experimental antiviral gene therapy with a synthetic mimic of an antiviral cellular microRNA. Intraperitoneal transplantation of human cells rapidly provided reservoirs of hepatitis B virus in mice. This simple xenotransplantation approach will be effective and convenient for studies of hepatitis B and other human viruses in vivo.

Molecular engineering of short half-life small peptides (VIP, αMSH and γ3MSH) fused to latency-associated peptide results in improved anti-inflammatory therapeutics

ObjectiveTo facilitate the targeting to inflammation sites of small anti-inflammatory peptides, with short half-lives, by fusion with the latency-associated peptide (LAP) of transforming growth factor β1 through a cleavable matrix...

Characterization of Toxoplasma gondii Proteins From Various Strains

Toxoplasma gondii the causative agent of toxoplasmosis, is a worldwide intracellular protozoan parasite that infects all warm blooded animals including humans. It can be devastating to immunocompromised humans and congenital transmission may result in severe clinical spectrum. It causes economic losses due to abortus in animals. Toxoplasmosis diagnosis depends on direct and indirect methods. Besides the Sabin-Feldman test, which is accepted to be the reference test, serologic tests such as ELISA and immunofluorescence antibody tests are means of indirect diagnosis. As detected antibodies in serologic tests are correlated with antigens that cause their synthesis, it is important to know different proteins of different strains. In this study RH, Ankara and TS-4 strains were used and differences between their proteins were examined. RH and Ankara strains were inoculated into the peritoneal cavity of mice. TS- 4 strain was produced in Vero cell culture. Tachyzoites collected by peritoneal wash were lysed and lyophilised. This was run on SDS-PAGE gel and protein bands were compared with a standard protein ladder after staining with polychromatic silver stain. It was observed that, while Ankara and RH strains had dense bands between 60-70 kDa and at 15 kDa, the most prominent bands of TS-4 strain were 60 ve 115 kDa bands. RH and Ankara strains have the same protein bands while TS-4 strain has different and fewer protein bands than the others.

Immunosuppressive effect of progesterone on dendritic cells in mice

Progesterone has been demonstrated to be involved in maintaining pregnancy by regulating immunocytes. Dendritic cells (DCs), the most potent triggers of the adaptive immune response, express receptors for steroid hormones and are regarded as one of the primary targets of progesterone. However, the functional modification of DCs by progesterone remains poorly understood. Here, we report that progesterone does not affect the morphology or apoptosis of murine bone marrow-derived DCs. Progesterone-treated DCs were characterized by decreased expression of Ia (MHC class II), CD80 and CD86, increased production of IL-10, and decreased secretion of IL-12. Compared with mature DCs (mDCs), activated progesterone-treated DCs had a reduced capacity to stimulate CD4(+) T cell proliferation. The observation that progesterone-treated DCs could attenuate delayed-type hypersensitivity (DTH) responses in vivo suggests that progesterone mediates suppressive DC activity. However, transfer of progesterone-treated DCs into the peritoneal cavity of mice did not elevate the percentage of CD4(+)CD25(+)Foxp3(+) regulatory T cells in the spleen. Overall, our study helps to increase understanding of the role of DCs exposed to progesterone in the maintenance of pregnancy.

The bradykinin B1 receptor antagonist R-954 inhibits Ehrlich tumor growth in rodents

The present study investigated the effects of a new bradykinin B1 receptor antagonist, R-954, on the development of Ehrlich ascitic tumor (EAT) induced by the intraperitoneal inoculation of EAT cells in mice and the formation of a solid tumor by the subcutaneous injection of the cells in rat paw. The development of the tumor was associated with an increase in mouse total cell counts in bone marrow (10.8-fold), ascitic fluid (14.6-fold), and blood (12.6-fold). R-954 (2mg/kg, s.c.) significantly reduced the ascitic fluid volume (63.7%) and the mouse weight gain (30.5%) after 10 consecutive days of treatment. The B1 antagonist as well as the anti-neoplasic drug vincristine also significantly inhibited the increase in total cell count in bone marrow, ascitic fluid, and blood. R-954 reduced significantly the total protein extravasation (57.3%), the production of nitric oxide (56%), PGE2 production (82%), and TNFα release (85.7%) in mice peritoneal cavity whereas vincristine reduced the release of these inflammatory mediators by 84–94%. The increase in paw edema after intraplantar injection of EAT cells was reduced by approximately 52% by either R-954 or vincristine treatment. In conclusion, this study presents for the first time the antitumoral activity of a new bradykinin B1 receptor antagonist on ascitic and solid tumors induced by Ehrlich cell inoculation in mice and rats.

RTX Toxin Enhances the Survival of Vibrio vulnificus During Infection by Protecting the Organism From Phagocytosis

Vibrio vulnificus is a marine bacterium causing serious septicemia and wound infection in humans. It produces an RTX toxin that can lyse a variety of cells and is important for virulence in mice. In this study, we explored the role of RTX in pathogenesis by characterizing an RTX-deficient mutant. This mutant showed an ∼2-log reduction in virulence for mice infected by various routes. Survival of the mutant at the infection site and subsequent spread into the bloodstream were impaired. In mice pretreated with cyclophosphamide to deplete the neutrophils, both the virulence and survival at the infection site of this mutant were enhanced. This mutant was further shown to be more readily cleared from the macrophage-rich mouse peritoneal cavity and phagocytosed by murine macrophages. These findings suggest that the RTX of V. vulnificus is required for bacterial survival during infection by protecting the organism from phagocytosis.

Leukotriene B4 Mediates Neutrophil Migration Induced by Heme

High concentrations of free heme found during hemolytic events or cell damage leads to inflammation, characterized by neutrophil recruitment and production of reactive oxygen species, through mechanisms not yet elucidated. In this study, we provide evidence that heme-induced neutrophilic inflammation depends on endogenous activity of the macrophage-derived lipid mediator leukotriene B(4) (LTB(4)). In vivo, heme-induced neutrophil recruitment into the peritoneal cavity of mice was attenuated by pretreatment with 5-lipoxygenase (5-LO) inhibitors and leukotriene B(4) receptor 1 (BLT1) receptor antagonists as well as in 5-LO knockout (5-LO(-/-)) mice. Heme administration in vivo increased peritoneal levels of LTB(4) prior to and during neutrophil recruitment. Evidence that LTB(4) was synthesized by resident macrophages, but not mast cells, included the following: 1) immuno-localization of heme-induced LTB(4) was compartmentalized exclusively within lipid bodies of resident macrophages; 2) an increase in the macrophage population enhanced heme-induced neutrophil migration; 3) depletion of resident mast cells did not affect heme-induced LTB(4) production or neutrophil influx; 4) increased levels of LTB(4) were found in heme-stimulated peritoneal cavities displaying increased macrophage numbers; and 5) in vitro, heme was able to activate directly macrophages to synthesize LTB(4). Our findings uncover a crucial role of LTB(4) in neutrophil migration induced by heme and suggest that beneficial therapeutic outcomes could be achieved by targeting the 5-LO pathway in the treatment of inflammation associated with hemolytic processes.

Nattectin a fish C-type lectin drives Th1 responses in vivo: Licenses macrophages to differentiate into cells exhibiting typical DC function

Considerable efforts are currently focused on the biology of DC in view of their possible clinical use as adjuvant for the generation of antigen-specific immunity and lifelong immunologic memory or for the treatment of tumors. We assessed the role of Nattectin a C-type lectin identified in the Thalassophryne nattereri fish venom in DC maturation. Nattectin induced a significant neutrophilic recruitment into peritoneal cavity of mice, followed by macrophages, with lipidic mediators and IL-12 p70 synthesis. Macrophages derived from 7day-Nattectin mice were CD11c+CD11blowLy6highF4/80Rhigh and express high levels of MHC class II and CD80 molecules. Culture of peritoneal exudates derived macrophages from 7day Nattectin-mice and immature BMDCs with Nattectin markedly increased the surface expression of CD40, CD80, CD86, and MHC class II in a dose-dependent manner, and the production of MMP-2 and MMP-9 distributed in nucleus and cytoplasm of cells, that was associated with strong activity in the culture supernatant. Nattectin treated DCs secreted IL-12 p70 and IL-10. The Nattectin-treated BMDC or macrophage-derived DCs were highly efficient at Ag capture. The specific immune response elicited by Nattectin was characterized by the production of specific antibodies IgG1 and mainly IgG2a with IL-10 and IFN-γ synthesis by splenic cells. These results enable us to address that Nattectin induces the recruitment of Ly6Chigh monocytes into the peritoneum, which exhibit a pro-inflammatory profile, where they differentiate into proliferating F4/80Rhigh macrophages. Macrophage-derived DCs mature in the presence of the cytokine milieu generated against Nattectin, exhibiting T cell co-stimulatory molecule expression and induced a Th1 polarized response.

Abstract 449: Tumor microenvironment induces profound immune suppression: Macrophages from peripheral and tumor locations differ in their degrees of inflammatory impairment

Abstract Macrophages are key players in the inflammatory response and exhibit significant roles in the different stages of tumor progression. We have previously reported the deficient functions of elicited macrophages recruited to the peritoneal cavity of mice bearing advanced D1-DMBA3 mammary tumors (T-PEMs). We showed that these T-PEMs are impaired in inflammatory activity due to diminished expression of transcription factors NFκB and C/EBP. We have also demonstrated that T-PEMs display neither M1 nor M2 phenotypes, exhibit deficiencies in the production of several inflammatory cytokines and proinflammatory signaling pathways intermediates and are less differentiated. Moreover, due in part to NFκB downregulation, increased apoptosis is observed in T-PEMs, a phenomenon which we reported for the first time that may contribute to increased myelopoiesis in tumor hosts. We have also recently examined the characteristics of blood monocytes from mice bearing the same mammary tumors and have demonstrated that these cells are neither inflammatory nor suppressive and are less differentiated, similar to the T-PEMs they later become. In the present study we investigate tumor-associated macrophages (TAMs) from the same animal tumor model and compare them with T-PEMs and with peritoneal macrophages from normal mice (N-PEMs). Our results show that TAMs downregulate IL-12p70, yet upregulate IL-12p40, IL-23 and IL-10. NFκB and C/EBP family members are decreased in TAMs as compared with T-PEMs, however NFκBp50 homodimers, STAT1/pSTAT1 and STAT3/pSTAT3 are all overexpressed. Interestingly, macrophages from tumor-bearing mice block T-cell proliferation, with TAMs exerting a more powerful inhibitory effect than T-PEMs. Also, TAMs are more prone to apoptosis than T-PEMs due to a higher expression of activated caspase 3 and downregulation of Bclx. TAMs also exhibit lower levels of differentiation than T-PEMs. In addition, our results show that T-PEMs and TAMs have equally impaired phagocytic capacity compared with fully-phagocytic N-PEMs. Moreover, TAMs constitutively express iNOS and produce nitric oxide as compared with T-PEMs, and they have constitutively lower arginase activity than T-PEMs. Furthermore, TAMs are Gr-1high and CD11blow, thus they are not classic myeloid-derived suppressor cells. Our results show that different locations in tumor hosts determine macrophage phenotypes and functions. The tumor microenvironment, as the location in most intimate contact with tumor cells, is the one that most alters macrophage activity. This is not only due to the occurrence of higher levels of tumor factors secreted by tumor cells, but also to the presence of other cell types that may interact with macrophages and modulate their activity. Targeting these tumor factors and cells in the tumor microenvironment will allow reversion of macrophage dysfunction. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 449. doi:10.1158/1538-7445.AM2011-449

Cutting Edge: Cyclic Polypeptide and Aminoglycoside Antibiotics Trigger IL-1β Secretion by Activating the NLRP3 Inflammasome

Clinical use of antibiotics is based on their capacity to inhibit bacterial growth via bacteriostatic or bacteriocidal effects. In this article, we show that the aminoglycoside antibiotic neomycin, the cyclic lipopeptide antibiotic polymyxin B, and the cyclic peptide antibiotics gramicidin and tyrothricin can induce IL-1β secretion in bone marrow dendritic cells and macrophages. LPS priming was required to trigger the transcription and translation of pro-IL-1β but was independent of TNFR or IL-1R signaling. All four antibiotics required the NLRP3 inflammasome, the adaptor ASC, and caspase-1 activation to secrete IL-1β, a process that depended on potassium efflux but was independent of P2X7 receptor. All four antibiotics induced neutrophil influx into the peritoneal cavity of mice, which required NLRP3 only in the case of polymyxin B. Together, certain antibiotics have the potential to directly activate innate immunity of the host.

Inhibition of nuclear factor‐kappaB suppresses peritoneal dissemination of gastric cancer by blocking cancer cell adhesion

Currently, patients with peritoneal dissemination of gastric cancer must accept a poor prognosis because there is no standard effective therapy. To inhibit peritoneal dissemination it is important to inhibit interactions between extracellular matrices (ECM) and cell surface integrins, which are important for cancer cell adhesion. Although nuclear factor-kappa B (NF-κB) is involved in various processes in cancer progression, its involvement in the expression of integrins has not been elucidated. We used a novel NF-κB inhibitor, dehydroxymethylepoxyquinomicin (DHMEQ), to study whether NF-κB blocks cancer cell adhesion via integrins in a gastric cancer dissemination model in mice and found that DHMEQ is a potent suppressor of cancer cell dissemination. Dehydroxymethylepoxyquinomicin suppressed the NF-κB activity of human gastric cancer cells NUGC-4 and 44As3Luc and blocked the adhesion of cancer cells to ECM when compared with the control. Dehydroxymethylepoxyquinomicin also inhibited expression of integrin (α2, α3, β1) in in vitro studies. In the in vivo model, we injected 44As3Luc cells pretreated with DHMEQ into the peritoneal cavity of mice and performed peritoneal lavage after the injection of cancer cells. Viable cancer cells in the peritoneal cavities were evaluated sequentially by in vivo imaging. In mice injected with DHMEQ-pretreated cells and lavaged, live cancer cells in the peritoneum were significantly reduced compared with the control, and these mice survived longer. These results indicate that DHMEQ could inhibit cancer cell adhesion to the peritoneum possibly by suppressing integrin expression. Nuclear factor-kappa B inhibition may be a new therapeutic option for suppressing postoperative cancer dissemination.

Characterization of immunomodulatory activity of eIF4A protein

Leishmania LeIF antigen, homologous to eukaryotic initiation factor eIF4A, was originally described as a Th1-type natural adjuvant and as an antigen that induces an IL-12-mediated Th1 response in the peripheral blood mononuclear cells (PBMC) of leishmaniasis patients. We showed previously that the induction of cytokines in monocytes of healthy subjects is not unique to the Leishmania protein. Indeed, 5 homologous proteins DEAD box in mammals and yeast were also able to induce the secretion of cytokines in monocytes of healthy subjects. In this study we aimed to validate eIF4A protein as a natural adjuvant. To achieve this objective, LeIF, eIF4A from mouse (MeIF4A) and Yeast (YeIF4A) were expressed and purified. The purified proteins were assessed for their ability to induce maturation of bone marrow-derived dendritic cells (BMDC). Their effect on DCs and their monocytic precursors in the peritoneal cavity of mice were analyzed. Our data showed that eIF4A proteins were able to activate BMDC to express co-stimulatory molecules and to produce IL-12p40/p70 and iNOS in vitro. Furthermore, eIF4A proteins induced inflammation in the peritoneal cavity of BALB/c mice similar to the well known adjuvant Alum. Indeed, injection of eIF4A proteins in combination with OVA protein induced a rapid recruitment into the peritoneal cavity of Ly6Chigh/CD11b+ inflammatory monocytes, Ly6Ghigh/CDb+ neutrophils and myeloid dendritic cells (MHC IIhigh-CD11c+). This study highlights the adjuvant activity of eIF4A protein and suggests that the immunomodulatory properties of eIF4A could be exploited in vaccination or immunotherapy protocols. Supported by grants from MESRST-Tunisia (LR00SP04), UNICEF/UNDP/WorldBank/WHO special programme TDR ({type:entrez-nucleotide,attrs:{text:A30134,term_id:21727281,term_text:A30134}}A30134) and a Fellowship from the International Network Institut Pasteur.

Role of Alumina Nanoporosity in Acute Cell Response

This work studied the effect of nanoporous alumina in acute cellular response in an in vivo model. Nanoporous alumina membranes, with pore size diameters of 20 and 200 nm, were fabricated by anodic oxidation of aluminium. The membranes were thereafter characterized in terms of pore size distribution and chemical composition. To evaluate acute inflammatory response, the membranes were implanted in the peritoneal cavity of mice. Cell recruitment to the implant site was determined by fluorescence activated cell sorting (FACS) analysis. Cell adhesion to material surfaces was studied in terms of cell number, type, and morphology using scanning electron microscopy (SEM) and immunocytochemical staining followed by fluorescence microscopy. The fabricated nanoporous alumina membranes were found to have narrow pore size distribution. The in vivo study showed that 200 nm alumina membranes induced stronger inflammatory response than 20 nm membranes. This was reflected by the number of implant-associated phagocytes and the number of cells recruited to the implantation site. Since both pore-size membranes possess similar chemical composition, we believe that the observed difference in cell recruitment and adhesion is an effect of the material nanotopography. Our results suggest that nanotopography can be used to subtly control the recruitment and adherence of phagocytic cells during the acute inflammatory response to alumina membranes.

Development of peritoneal macrophage along a dendritic cell lineage in response to uptake of oligomannose-coated liposomes

In this study, we investigate the potential of peritoneal macrophages to differentiate into dendritic cell (DCs) in response to preferential uptake of oligomannose-coated liposomes (OMLs). About 30% of peritoneal cells (PECs) preferentially took up OMLs that were administered into the peritoneal cavity. The OML-ingesting cells expressed CD11b and F4/80, but lacked CD11c expression, indicating that the OML-ingesting PECs with a CD11b highCD11c − phenotype are resident peritoneal macrophages. During in vitro cultivation, CD11c + cells arose among the PECs with ingested OMLs. CD11c + cells also developed among enriched peritoneal CD11b highCD11 − cells from OML-treated mice, and the resulting CD11c + cells expressed co-stimulatory molecules and MHC class II. In addition, OML-ingesting CD11b highCD11c + cells were found in spleen after the enriched peritoneal macrophages with ingested OMLs were transplanted in the peritoneal cavity of mice. These results show that a fraction of peritoneal macrophages can differentiate into mature DCs following uptake of OMLs.

CXCL4L1 inhibits angiogenesis and induces undirected endothelial cell migration without affecting endothelial cell proliferation and monocyte recruitment

The non-allelic variant of CXCL4/PF4, CXCL4L1/PF4alt, differs from CXCL4 in three amino acids of the C-terminal α-helix and has been characterized as a potent anti-angiogenic regulator. Although CXCL4 structurally belongs to the chemokine family, it does not behave like a 'classical' chemokine, lacking significant chemotactic properties. Specific hallmarks are its angiostatic, anti-proliferative activities, and proinflammatory functions, which can be conferred by heteromer-formation with CCL5/RANTES enhancing monocyte recruitment. Here we show that tube formation of endothelial cells was inhibited by CXCL4L1 and CXCL4, while only CXCL4L1 triggered chemokinesis of endothelial cells. The chemotactic response towards VEGF and bFGF was attenuated by both variants and CXCL4L1-induced chemokinesis was blocked by bFGF or VEGF. Endothelial cell proliferation was inhibited by CXCL4 (IC(50) 6.9 μg mL(-1)) but not by CXCL4L1, while both chemokines bound directly to VEGF and bFGF. Moreover, CXCL4 enhanced CCL5-induced monocyte arrest in flow adhesion experiments and monocyte recruitment into the mouse peritoneal cavity in vivo, whereas CXCL4L1 had no effect. CXCL4L1 revealed lower affinity to CCL5 than CXCL4, as quantified by isothermal fluorescence titration. As evidenced by the reduction of the activated partial thromboplastin time, CXCL4L1 showed a tendency towards less heparin-neutralizing activity than CXCL4 (IC(50) 2.45 vs 0.98 μg mL(-1)). CXCL4L1 may act angiostatically by causing random endothelial cell locomotion, disturbing directed migration towards angiogenic chemokines, serving as a homeostatic chemokine with a moderate structural distinction yet different functional profile from CXCL4.

Thalidomide Prevents the Progression of Peritoneal Fibrosis in Mice

Thalidomide is clinically recognized as a therapeutic agent for multiple myeloma and has been known to exert anti-angiogenic actions. Recent studies have suggested the involvement of angiogenesis in the progression of peritoneal fibrosis. The present study investigated the effects of thalidomide on the development of peritoneal fibrosis induced by injection of chlorhexidine gluconate (CG) into the mouse peritoneal cavity every other day for 3 weeks. Thalidomide was given orally every day. Peritoneal tissues were dissected out 21 days after CG injection. Expression of CD31 (as a marker of endothelial cells), proliferating cell nuclear antigen (PCNA), vascular endothelial growth factor (VEGF), α-smooth muscle actin (as a marker of myofibroblasts), type III collagen and transforming growth factor (TGF)-β was examined using immunohistochemistry. CG group showed thickening of the submesothelial zone and increased numbers of vessels and myofibroblasts. Large numbers of VEGF-, PCNA-, and TGF-β-positive cells were observed in the submesothelial area. Thalidomide treatment significantly ameliorated submesothelial thickening and angiogenesis, and decreased numbers of PCNA- and VEGF-expressing cells, myofibroblasts, and TGF-β-positive cells. Moreover, thalidomide attenuated peritoneal permeability for creatinine, compared to the CG group. Our results indicate the potential utility of thalidomide for preventing peritoneal fibrosis.

Impaired Cd14 and Cd36 Expression, Bacterial Clearance, and Toll-Like Receptor 4-Myd88 Signaling in Caveolin-1-Deleted Macrophages and Mice

An overwhelming immune response, particularly from macrophages, with gram-negative bacteria-induced sepsis plays a critical role in survival of and organ damage in infected patients. Caveolin-1 (Cav-1), a major structure protein of caveolae, regulates many cellular functions. We examined the vital role of Cav-1 in the response of macrophages and mice to bacteria or LPS exposure. Deletion of Cav-1 decreased the expression of CD14 and CD36 during macrophage differentiation and suppressed their phagocytotic ability. As well, the ability to kill bacteria was inhibited in Cav-1 macrophages and mice peritoneal cavity, tissue, and plasma, which was partly attributed to hindered expression of iNOS induced by bacteria or LPS. Furthermore, deletion of Cav-1 attenuated the expression of Toll-like receptor 4 and myeloid differentiation factor 88 and the activation of nuclear factor κB, all of which impeded the production of inflammatory cytokines in response to bacterial exposure in Cav-1 macrophages and mice. Thus, Cav-1 participates in the regulation of CD14, CD36, Toll-like receptor 4 and myeloid differentiation factor 88 protein expression and is crucial for the immune response of macrophages to bacterial infection. Cav-1 may be a therapeutic target in the treatment of sepsis.

A strategy for the engineering of insulin producing cells with a broad spectrum of defense properties

Insulin-producing pancreatic beta cells are known to be extremely susceptible to the oxidative stress and hypoxia generated following islet transplantation in diabetic patients. We hereby present a novel in vivo selection strategy based on the isolation of insulin-producing cells with enhanced protection after repeated rounds of encapsulation and xenotransplantation. Rat insulinoma INS-1 cells were encapsulated in alginate macrobeads and transplanted in the peritoneal cavity of mice. After 2 days the beads were retrieved and cells were recovered from alginate and propagated in vitro until submitted to a second round of encapsulation and transplantation. Three days later, the surviving cells, named INS-1m2, were isolated from the alginate beads and their protection and functional activity examined. Compared to parental INS-1 cells, the selected INS-1m2 cells were more resistant to hydrogen peroxide, nitric oxide, alloxan and hypoxia. This enhanced protection of the selected cells correlated with the increased level of catalase and poly (ADP-ribose) polymerase expression. Although selected cells expressed more insulin than parental cells, no change in their insulin response to glucose was observed. We conclude that the in vivo selection strategy is a powerful tool for the engineering of insulin producing cells with a broad spectrum of defense properties.

Development and prevention of postsurgical adhesions in a chimeric mouse model of experimental endometriosis

To examine the impact of a recent surgery on development of endometriosis-related adhesions in a chimeric model and to determine the therapeutic efficacy of pioglitazone (PIO). Human endometrial biopsies were maintained in E(2) with or without PIO for 24 h before intraperitoneal injection into immunocompromised mice also treated with or without PIO at multiple time points after peritoneal surgery. The presence and extent of adhesions were examined in animals relative to the initial establishment of experimental endometriosis. Medical school research center. Endometrial biopsies for experimental studies were provided by normally cycling women without a medical history indicative of endometriosis or adhesions. None. Examination of the development of endometriosis-related adhesions in an experimental model. Without therapeutic intervention, injection of E(2)-treated human endometrial tissue into mice near the time of peritoneal surgery resulted in multiple adhesions and extensive endometriotic-like disease. In contrast, PIO treatment reduced adhesive disease and experimental endometriosis related to surgical injury. The presence of human endometrial tissue fragments in the peritoneal cavity of mice with a recent surgical injury promoted development of both adhesive disease and experimental endometriosis. Targeting inflammation and angiogenesis with PIO therapy limited the development of postsurgical adhesions associated with ectopic endometrial growth.

LRRK2 is expressed in B-2 but not in B-1 B cells, and downregulated by cellular activation

LRRK2, the causal molecule of familial Parkinson's disease, is expressed strongly by one of the B cell subsets, B-2 cells, but not by the other subset, B-1 cells, in the mouse peritoneal cavity, spleen, and peripheral blood. Bone marrow pre-B cells or T cells exhibited little LRRK2 expression. LRRK2 expression was dramatically downregulated upon activation of B-2 cells with various types of stimulation. These results suggest that LRRK2, whose true function has not yet been clarified, may play some important role(s) in the development and function of B cells, particularly the maintenance of B-2 cells in a resting status.