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ABSTRACT: Renal cell carcinoma (RCC) is highly resistant to chemotherapy and unresponsive to radio- and immunotherapy. Recently, we have documented that the histone deacetylase (HDAC)-inhibitor valproic acid (VPA) in combination with low-dosed interferon (IFN)-alpha significantly inhibits RCC proliferation and adhesion in vitro and in vivo. The current study investigated the effects of these compounds on gene transcription of metastatic RCC cell line Caki-1 after 3 and 5 days exposure.
To evaluate the gene expression profiles of the RCC cells, we performed microarray analysis using Affymetrix GeneChip. Selected significant genes were further validated by Real Time PCR.
Microarray revealed that VPA altered genes that are involved in cell growth, cell survival, immune response, cell motility and cell adhesion. Combination of VPA with IFN-alpha not only enhanced the effects on gene transcription but also resulted in the expression of novel genes, which were not induced by either VPA or IFN-alpha alone. Among the up-regulated genes were chemokines (CXCL10, CXCL11, CXCL16) and integrins (ITGA2, ITGA4, ITGA5, ITGA6, ITGA7). Genes encoding for adhesion molecules (NCAM1, ICAM1, VCAM1) were also modulated. Real Time PCR approved these findings.
This data provides insight into the molecular mechanism of action of the combined treatment of VPA and IFN-alpha in RCC. Implications are that the combined application of VPA and IFN-alpha may represent a more efficient alternative to existing therapy options for RCC.
World Journal of Urology 12/2011; 29(6):779-86. · 2.41 Impact Factor
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ABSTRACT: To evaluate the incidence of voiding dysfunction in older male renal transplant recipients.
Data for 103 patients aged 60 years or older (mean age, 65.7 years; group 1) who underwent transplantation at our center between January 1999 and August 2007 were compared with data for a group of 139 younger patients (mean age, 50.1 years; group 2) treated within the same time frame.
Postoperatively, 28 group 1 recipients (27%) and 26 group 2 recipients (19%) experienced voiding dysfunction after removal of the transurethral catheter (P = .12). The most common cause was bladder outlet obstruction due to benign prostatic hyperplasia in 26 patients in group 1 (25%) and 17 patients in group 2 (12%) (P = .009). Bladder neck contracture, urethral stricture, and detrusor underactivity were diagnosed in the other patients. Transurethral resection of the prostate gland was performed in 21 group 1 patients (20%) and 14 group 2 patients (10%) (P = .02) at a mean of 31.1 and 29.5 days, respectively (P = .23) after transplantation. Surgical procedures were performed without complication, and symptoms did not recur postoperatively.
Our data reveal a high incidence of voiding dysfunction in older male renal transplant recipients. High residual urine and urinary retention after renal transplantation may induce recurrent urinary tract infections, cause relevant complications, and seriously affect graft function. Recognizing the substantial effects of postoperative voiding dysfunction will enable optimum management of older kidney transplant recipients.
Transplantation Proceedings 07/2009; 41(5):1615-8. · 1.00 Impact Factor
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ABSTRACT: Conventional therapeutic approaches to treat advanced renal cell carcinoma (RCC) are of limited benefit. Receptor tyrosine kinase inhibitors (RTKI) may open up novel treatment options. In the present study, the effects of the RTKI AEE788 on the growth and adhesion capacity of RCC cell lines were evaluated in vitro.
RCC cells were treated with AEE788, and alterations of tumor growth and tumor cell interaction with vascular endothelium or extracellular matrix proteins were analyzed. Furthermore, the addition of interferon alpha (IFNalpha) was investigated to see whether it may enhance the anti-tumoral potential of AEE788.
AEE788 significantly blocked RCC cell growth and adhesion. Analysis of alpha- and beta-integrins revealed distinct alterations of the receptor expression profile and downregulation of integrin-dependent signaling. Growth-blocking effects were further enhanced when the AEE788-IFNalpha combination protocol was applied. In addition, downregulation of integrin-dependent signaling was more intense in the presence of a combination of AEE788 and IFNalpha than with AEE788 monotherapy.
AEE788 exerts significant anti-tumoral properties, particularly when combined with IFNalpha. AEE788 may therefore be an encouraging compound to treat advanced RCC.
Der Urologe 10/2008; 47(9):1175-81. · 0.50 Impact Factor
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ABSTRACT: Drug resistance to chemotherapy is often associated with increased malignancy in neuroblastoma (NB). In pursuit of alternative treatments for chemoresistant tumour cells, we tested the response of multidrug-resistant SKNSH and of vincristine (VCR)-, doxorubicin (DOX)-, or cisplatin (CDDP)-resistant UKF-NB-2, UKF-NB-3 or UKF-NB-6 NB tumour cell lines to valproic acid (VPA), a differentiation inducer currently in clinical trials. Drug resistance caused elevated NB adhesion (UKF-NB-2(VCR), UKF-NB-2(DOX), UKF-NB-2(CDDP), UKF-NB-3(VCR), UKF-NB-3(CDDP), UKF-NB-6(VCR), UKF-NB-6(CDDP)) to an endothelial cell monolayer, accompanied by downregulation of the adhesion receptor neural cell adhesion molecule (NCAM). Based on the UKF-NB-3 model, N-myc proteins were enhanced in UKF-NB-3(VCR) and UKF-NB-3(CDDP), compared to the drug naïve controls. p73 was diminished, whereas the p73 isoform deltaNp73 was upregulated in UKF-NB-3(VCR) and UKF-NB-3(CDDP). Valproic acid blocked adhesion of UKF-NB-3(VCR) and UKF-NB-3(CDDP), but not of UKF-NB-3(DOX), and induced the upregulation of NCAM surface expression, NCAM protein content and NCAM coding mRNA. Valproic acid diminished N-myc and enhanced p73 protein level, coupled with downregulation of deltaNp73 in UKF-NB-3(VCR) and UKF-NB-3(CDDP). Valproic acid also reverted enhanced adhesion properties of drug-resistant UKF-NB-2, UKF-NB-6 and SKNSH cells, and therefore may provide an alternative approach to the treatment of drug-resistant NB by blocking invasive processes.
British Journal of Cancer 07/2007; 96(11):1699-706. · 5.04 Impact Factor
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ABSTRACT: The branched-chain fatty acid valproate (valproic acid; VPA) displays antitumoral properties by blocking tumor growth, progression and invasion. Recent data have shown that VPA reduces the angiogenic activity of endothelial cells. The object of this study was to investigate whether endothelial modulation might also influence the level of chemotactic mediators. Endothelial cells were isolated from human umbilical cord veins (HUVEC) and treated with VPA-concentrations ranging from 0.125 mM to 1 mM. The mRNA level of CXC-chemokines was investigated by reverse transcriptase-polymerase chain reaction. The proliferative activity of HUVEC was measured as well. VPA evoked a striking increase in the neutrophil chemoattractants CXCL1, CXCL3, CXCL4, CXCL5 and a moderate increase in CXCL6 with maximal effects after a 3-day incubation period. Other CXC-chemokines and CXC-receptors remained unaffected. HUVEC growth was diminished time- and dose-dependently by VPA. We conclude that VPA treatment leads to alterations in the chemokine expression profile of endothelial cells. This might allow more neutrophils to reach the tumor area and trigger cytolysis.
International journal of clinical pharmacology and therapeutics 11/2004; 42(10):568-74. · 1.18 Impact Factor
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K Leckel,
W-D Beecken,
D Jonas,
E Oppermann,
M C Coman,
K-F Beck,
J Cinatl,
N P Hailer,
M K H Auth,
W O Bechstein,
M Shipkova, R A Blaheta
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ABSTRACT: Immunosuppression correlates with the development and recurrence of cancer. Mycophenolate mofetil (MMF) has been shown to reduce adhesion molecule expression and leucocyte recruitment into the donor organ. We have hypothesized that MMF might also prevent receptor-dependent tumour dissemination. Therefore, we have investigated the effects of MMF on tumour cell adhesion to human umbilical vein endothelial cells (HUVEC) and compared them with the effects on T cell-endothelial cell interactions. Influence of MMF on cellular adhesion to HUVEC was analysed using isolated CD4+ and CD8+ T cells, or WiDr colon adenocarcinoma cells as the model tumour. HUVEC receptors ICAM-1, VCAM-1, E-selectin and P-selectin were detected by flow cytometry, Western blot or Northern blot analysis. Binding activity of T cells or WiDr cells in the presence of MMF were measured using immobilized receptor globulin chimeras. MMF potently blocked both T cell and WiDr cell binding to endothelium by 80%. Surface expression of the endothelial cell receptors was reduced by MMF in a dose-dependent manner. E-selectin mRNA was concurrently reduced with a maximum effect at 1 microm. Interestingly, MMF acted differently on T cells and WiDr cells. Maximum efficacy of MMF was reached at 10 and 1 microm, respectively. Furthermore, MMF specifically suppressed T cell attachment to ICAM-1, VCAM-1 and P-selectin. In contrast, MMF prevented WiDr cell attachment to E-selectin. In conclusion, our data reveal distinct effects of MMF on both T cell adhesion and tumour cell adhesion to endothelial cells. This suggests that MMF not only interferes with the invasion of alloactivated T cells, but might also be of value in managing post-transplantation malignancy.
Clinical & Experimental Immunology 12/2003; 134(2):238-45. · 3.36 Impact Factor
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ABSTRACT: In vitro study on the effects of mycophenolate mofetil (MMF) on isolated human monocytes and endothelial cells.
Haematogenous macrophages play an essential role in the development of secondary damage following spinal cord injury (SCI), and there is evidence that the use of immunosuppressants such as MMF can reduce monocyte invasion and neuronal damage.
University Hospital for Orthopaedic Surgery, Frankfurt am Main, Germany.
The effects of MMF on the adhesion of human monocytes to human umbilical vein endothelial cells (HUVEC), monocyte binding to immobilised E-selectin, and monocyte expression of intercellular adhesion molecule (ICAM)-1, sialyl Lewis X (sLeX) and major histocompatibility complex (MHC)-II were studied. The binding of monocytes to E-selectin was examined by using purified and immobilised E-selectin fusion protein. Adhesion molecule expression was investigated by flow cytometry.
The binding of monocytes to HUVEC was significantly reduced by 30.1% after treatment of monocytes with MMF (10 microg/ml), whereas the pretreatment of HUVEC with MMF did not result in significant changes in monocyte adhesion. MMF forcefully inhibited monocyte binding to immobilised E-selectin by 55.7%. Furthermore, MMF significantly inhibited the upregulation of ICAM-1- and MHC-II-expression on monocytes stimulated with either lipopolysaccharide or interferon-gamma, whereas the expression of sLeX was not impaired. Toxic effects were excluded by propidium-iodide staining and measurement of fluorescein-diacetate metabolism.
MMF can downregulate important monocytic adhesion molecules and inhibits monocyte adhesion to endothelial cells, thus indicating that treatment with MMF could be beneficial after SCI.
This study was supported by the DFG (Ha 2721/1-3), the Paul und Ursula Klein-Stiftung and the Stiftung Friedrichsheim.
Spinal Cord 11/2003; 41(11):610-9. · 1.80 Impact Factor
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ABSTRACT: To evaluate the impact of passenger leukocytes in liver grafts on the rate of microchimerism induction after liver transplantation and to evaluate immunological changes thereafter based on serial donor-specific MLC's in these patients.
26 orthotopic liver transplant recipients were prospectively evaluated for immunological changes based on the co-transplantation of donor-derived leukocytes. Intraoperatively harvested liver biopsies and peripheral blood-lymphocytes of liver transplant recipients were sampled at various time points. Donor spleen cells were obtained during organ procurement.
HLA-PCR analysis demonstrated a stable pattern of microchimerism in 15 out of 26 patients. Microchimerism was detectable by PCR up to a mean of 7 weeks after transplantation, when chimerism in the peripheral blood became negative. Passenger donor leukocytes were present in all biopsies obtained during backtable preparation of the liver graft. For the 15 patients presenting microchimerism the rate of passenger leukocytes in the liver graft biopsies showed a mean of 155.8 leukocytes per mm 2 liver tissue (SD +/- 23.2 cells/mm2, range 121 to 217 cells per mm2 tissue). Otherwise patients without chimerism showed a mean of 90.4 passenger leukocytes per mm2 tissue (SD +/- 14.5 cells/mm2, range: 52 to 99 cells/mm2). Lymphocyte proliferation, determined by donor-specific "multiple" single-way- mixed-lymphocyte-cultures (dsmMLC) was reduced to a mean of 62.2 % of preoperative values (SD +/- 14.5 %, range 33 % to 88 %) in the 15 patients with stable microchimerism. Otherwise in the 11 patients without microchimerism dsmMLC results stayed at continuously higher levels with a mean of 106 % (SD +/- 13.4, range 92 % to 134 %).
The results from these studies of microchimerism and lymphocyte reactivity after liver transplantation suggest that the co-transplantation of donor leukocytes plays an important and active role in the modulation of the host-immune system.
Zentralblatt für Chirurgie 05/2003; 128(4):278-82. · 1.02 Impact Factor
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ABSTRACT: Over the past few years, hepatocyte transplantation has been considered as an alternative method for orthotopic liver transplantation for the treatment of various liver diseases. Beside curative approach for genetic metabolic deficiencies (familial hypercholesterolemia, hemophilia, etc.), it could be a useful tool for bridging the waiting period until an appropriate donor organ is obtained. In preclinical animal studies, hepatocytes injected intraperitoneally, intraportally or into the spleen settle down in the diseased liver. This enables genetic modification to correct inborn metabolic deficiencies and improves survival in acute liver failure. In 1992, the first clinical transplantation of isolated hepatocytes in 10 patients was performed. In 1998, Fox and coworkers described the successful transplantation of allogeneic liver cells in a child with Crigler-Najjar syndrome. Accomplished studies of Strom et al. resp. Bilir et al. of the same year proved the effectiveness of liver cell transplantation for transient treatment of acute liver failure. Prerequisite of this cell-based therapeutic strategy is a sufficient amount of highly differentiated hepatocytes, hence, a well established in-vitro cell-culture technique is necessary to yield a reproducible number of proliferating hepatocytes and to preserve the physiological cell function. This review discusses the different experimental approaches regarding the cultivation of human hepatocytes and also the use of alternative cell sources (like animal hepatocytes, immortalized cells of human origin, progenitor cells from fetal human liver/liver stem cells) for hepatocyte transplantation.
Zentralblatt für Chirurgie 05/2003; 128(4):283-90. · 1.02 Impact Factor
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J-U Vogel,
M Michaelis,
J Neyts, R A Blaheta,
R Snoeck,
G Andrei,
E De Clercq,
H F Rabenau,
J Kreuter,
J Cinatl,
H W Doerr
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ABSTRACT: Antiviral activity of the metal chelator ethylenediaminedisuccinic acid (EDDS) was examined in vitro against human cytomegalovirus (HCMV) wild type strains and strains that are resistant against ganciclovir (GCV) and cidofovir (HPMPC). EDDS inhibited the replication of wild-type as well as GCV- and HPMPC-resistant strains with a 50% effective concentration of 7.4-12 microg/ml. At concentrations of 100 microg/ml EDDS, unlike GCV or HPMPC, suppressed HCMV-induced up-regulation of intercellular adhesion molecule-1 (ICAM-1) and reduced T-cell adhesion to HCMV-infected cells in a monolayer adhesion model. In vitro EDDS inhibited murine cytomegalovirus (MCMV) replication (EC50 8.6 microg/ml) and caused in mice some protection against MCMV induced mortality at a non-toxic dose. Since immunopathological factors may play a significant role in HCMV disease it will be of interest to further study whether EDDS is effective in terms of modulation of inflammatory responses to HCMV infections.
Antiviral Research 08/2002; 55(1):179-88. · 4.30 Impact Factor
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ABSTRACT: Although the control of biliary ductular morphogenesis has received some attention particularly using isolated rat biliary epithelial cell models, the regulation of human bile duct formation is not well defined. In the present study, using a 3-dimensional culture model comprising primary human biliary epithelial cells (BECs) and coculture with primary human hepatocytes, we have sought to define the factors involved. We have shown that primary human BECs can be expanded on collagen gels in the absence of growth factors or serum. When plated in high density in double collagen gels, BECs established 3-dimensional structures that subsequently developed into well differentiated polarized luminal ducts. This morphogenic response occurred in the absence of hepatocyte growth factor (HGF) and epidermal growth factor. Strikingly, the addition of growth factors (in the presence of serum) resulted in loss of polarity although the cells retained growth responses to both factors. Coculture of BECs with autologous human hepatocytes enhanced the ability of low-density BECs to undergo ductulogenesis. This effect was mimicked by addition of conditioned medium from previous hepatocyte-BEC cocultures. These findings indicate that for human biliary ductular morphogenesis, epithelial cell-cell interactions are required but that mesenchymally derived factors such as HGF may not be important.
Hepatology 04/2001; 33(3):519-29. · 11.66 Impact Factor
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ABSTRACT: Several factors contribute to the maintenance of central nervous system immune privilege and astrocytes have been identified as a major source of immunomodulatory cytokines. To investigate whether hematogenous monocytes are immunologically deactivated by astrocyte-derived factors human monocytes were stimulated with lipopolysaccharide or interferon (IFN)-gamma and treated with the supernatant from pure astrocyte cultures, interleukin (IL)-4, IL-10, or with IL-1-receptor antagonist (1L-1-RA). Flow cytometry demonstrated that the supernatant from astrocyte cultures was the most potent agent in reducing the levels of major histocompatibility complex (MHC)-class-II- as well as intercellular adhesion molecule-1-expression, whereas IL-4, IL-10, and IL-1-RA had only marginal effects. The expression of leukocyte function antigen-1 and very late antigen-4 was not modulated by either factor. In conclusion, astrocytes seem to provide soluble factors that have the capacity to deactivate hematogenous monocytes.
Neuroscience Letters 02/2001; 298(1):33-6. · 2.11 Impact Factor
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ABSTRACT: Interaction of endothelial P-selectin with sialyl Lewis(x)-glycoprotein or P-selectin glycoprotein ligand (PSGL)-1 on leukocytes represents an early step in leukocyte recruitment. Redistribution of P-selectin to the endothelial cell surface occurs rapidly after challenge with several proinflammatory agents, for example, histamine, leucopterins, or lipopolysaccharide. We present evidence that prostaglandin E2 (PGE2) is an efficient inductor of surface P-selectin on cultured human umbilical vein endothelial cells (HUVEC). The increase in P-selectin-immunoreactivity coincided with redistribution of cytoplasmic P-selectin-reactive granulae to the endothelial cell surface, as visualized by confocal laser microscopic examination. CD4-T-cell adhesion to PGE2-stimulated HUVEC was also enhanced by a factor of 4, and blocking mAb directed against the binding site of P-selectin almost completely abrogated this increase in CD4-T-cell adhesion. In summary, our findings show that liberation of PGE2 is an important inductor of P-selectin surface expression on endothelial cells, resulting in enhanced recruitment of inflammatory cells.
Transplantation 08/2000; 70(1):236-40. · 4.00 Impact Factor
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ABSTRACT: Prostaglandin E2 (PGE2) is a powerful endogenous immune suppressant and interferes with various T-cell functions. However, it is not known in detail whether immunosuppressive drugs influence the PGE2-driven immune response in transplant patients. Therefore, we investigated the effect of several immunosuppressive compounds, in particular the novel drug mycophenolate mofetil (MMF), on endothelial PGE2 release.
Endothelial cells (HUVEC) were activated by either allogeneic CD4+ or CD8+ T cells, or by the cytokines interleukin-1 or gamma-interferon. Using an enzyme-linked immunosorbent assay, we analyzed PGE2 release of the activated HWEC in the presence of MMF, cyclosporine, or tacrolimus. As verapamil and mibefradil also possess immunosuppressive properties, they were included in the study as well.
Activation of HUVEC with interleukin-1 or T cells resulted in a drastic accumulation of PGE2 in the supernatant. Cyclosporine or tacrolimus had no effect on PGE2 release. However, Ca2+ channel blockers, when applied at higher dosages, caused a significant increase in PGE2. Interestingly, MMF strongly diminished the PGE2 level in the cell culture supernatant in a concentration-dependent manner.
The results demonstrate an inhibitory effect of MMF on PGE2 production, which may lower the benefits of the PGE2-triggered immune response after organ transplantation.
Transplantation 06/2000; 69(9):1977-81. · 4.00 Impact Factor
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ABSTRACT: Previously, experimental in vivo results showed that the productively and persistently human cytomegalovirus (HCMV)-infected neuroblastoma cell line UKF-NB-4AD169 exhibits a more malignant phenotype than the non-infected variant UKF-NB-4. To prove the assumption that enhanced malignancy may be due to enhanced invasive potential of the infected cells we studied interactions of both lines with monolayers of cultured endothelial cells. UKF-NB-4AD169 cells adhered to and transmigrated through endothelial monolayer to a significantly higher extent compared with UKF-NB4. Furthermore, the adhesion of UKF-NB-4AD169 but not of UKF-NB4 resulted in focal disruption of the monolayer integrity which facilitates tumor cell transmigration. Blocking antibodies directed against the beta1 integrin chain as well as beta1alpha5 on the tumor cells specifically inhibited adhesion in a concentration-dependent manner. When UKF-NB-4 were pretreated with a beta1 integrin activating antibody, focal disruption of the endothelial integrity also occurred. These findings lead us to suggest that HCMV infection activates beta1alpha5 in the host neuroblastoma cell which in turn enables these cells to tightly adhere to endothelial cells. In the presence of the protease inhibitor phenantroline, beta1alpha5-mediated adhesion was not impaired whereas UKF-NB4AD169-mediated endothelial monolayer permeabilization was dose dependently inhibited. We conclude that human cytomegalovirus infection contributes to augmented neuroblastoma invasiveness via adhesion of activated beta1alpha5 and subsequent matrix digestion by proteases.
Tissue Antigens 06/2000; 55(5):412-21. · 2.59 Impact Factor
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ABSTRACT: Human cytomegalovirus (HCMV) infection is associated with excessive proinflammatory immune responses such as cytokine/chemokine production or upregulation of adhesion molecules on the host cells. It is assumed that these features of HCMV-related immunopathology can not be treated effectively with currently available anti HCMV drugs. In the present study the efficacy of ganciclovir (GCV), foscarnet (PFA), cidofovir (HPMPC), and ISIS 2922, an antisense oligonucleotide complementary to HCMV immediate-early (IE) mRNA, was investigated on HCMV-induced secretion and functional activity of the C-X-C chemokine IL-8 and the expression of the intercellular adhesion molecule-1 (ICAM-1). As compared with mock-infected cells IL-8 production was increased up to 9-fold and ICAM-1 expression up to 4-fold in HCMV-infected fibroblasts. Treatment of infected cells with GCV (40 microM), PFA (200 microM) or HPMPC (2 microM) suppressed completely virus replication as demonstrated by quantification of late (L) antigen expression and infectious virus production. These drugs, however, failed to inhibit IE antigen expression and did not prevent HCMV-induced upregulation of IL-8 and ICAM-1. In contrast, ISIS 2922 (1 microM) suppressed both IE and L antigen expression by 99% and inhibited infectious virus production by 10(4)-fold. Moreover, ISIS 2922 significantly suppressed HCMV-induced upregulation of both IL-8 and ICAM-1 expression on the transcriptional and on the protein level. Our results indicate that ISIS 2922 but not inhibitors of HCMV DNA prevents HCMV-induced upregulation of IL-8 and ICAM-1, both hallmarks of inflammatory processes. Thus, inhibition of HCMV IE expression with ISIS 2922 may be an important strategy for the treatment of HCMV-related immunopathogenesis.
Journal of Medical Virology 04/2000; 60(3):313-23. · 2.82 Impact Factor
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R A Blaheta,
N P Hailer,
N Brude,
B Wittig,
K Leckel,
E Oppermann,
M Bachmann,
S Harder,
J Cinatl,
M Scholz,
J Bereiter-Hahn,
S Weber,
A Encke,
B H Markus
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ABSTRACT: Cyclosporine A (CsA) and tacrolimus prevent proliferation but not transendothelial migration of alloreactive lymphocytes into donor organs. As a result, serious adverse effects, such as nephrotoxicity and neurotoxicity, have been observed under CsA/tacrolimus therapy. The incorporation of new drugs with infiltration blocking properties might enhance the efficacy of the current immunosuppressive protocol, allowing lower CsA/tacrolimus dosage. Because Ca2+ plays a critical role in cell-cell interaction, the Ca2+-channel blocker verapamil might be a good cany. didate for supporting CsA/tacrolimus-based therapy.
A T-cell endothelial cell coculture model or immobilized immunoglobulin G globulin chimeras were employed to investigate how S- and R- verapamil interfere with the lymphocytic infiltration process. The expression and arrangement of membranous adhesion receptors and cytoskeletal F-actin filaments were analyzed by fluorometric method in the presence of. verapamil.
Both verapamil enantiomers strongly inhibited lymphocyte infiltration. CD4+ and CD8+ T-cells were influenced to a similar extent with regard to horizontal locomotion (CD4+=CD8+), but to a different extent with regard to adhesion and penetration (CD4+ > CD8+). Moreover, penetration was blocked to a higher extent than was adhesion. ID50-values were 31 microM (CD4+-adhesion) and 11 microM (CD4+-penetration). Verapamil reduced P-selectin expression on endothelial cells and effectively down-regulated binding of T-cells to immobilized P-selectin immunoglobulin G globulins (ID50=4.4 microM; CD4+). A verapamil-induced reduction of intracellular F-actin in T-lymphocytes was proven to be mainly responsible for diminished cell locomotion.
The prevention of CD4+ T-cell penetration by verapamil might argue for its use as an adjunct to CsA/tacrolimus-based immunosuppressive therapy.
Transplantation 02/2000; 69(4):588-97. · 4.00 Impact Factor
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ABSTRACT: A variety of cells such as leukocytes and tumor cells may adhere to endothelial cells and subsequently transmigrate into the solid tissue by involving specific intercellular molecular pathways. One important prerequisite for transendothelial migration is the loosening of endothelial cell-to-cell contact sites, which can be triggered by extravasating cells. Cytomegalovirus (CMV) has obviously evolved the ability not only to influence host cells floating in the blood stream to adhere to endothelial cells, but also to induce the formation of intercellular gaps within the endothelium, resulting in transendothelial migration. These features allow the virus to disseminate and evade the immune system. In coculture experiments with human endothelial monolayers and human CMV (HCMV)-infected neuroblastoma cells or leukocytes, changes in the integrity of the monolayer were observed and further analyzed on the molecular level. For example, HCMV may activate the integrin beta1alpha5 (VLA-5) that triggers adhesion to endothelial cells with subsequent focal disruption of endothelial cell-to-cell connections. It is hypothesized that a Ca(2+)-independent pathway following VLA-5 binding disconnects the cadherin-catenin-actin complex within the endothelial cells. The loss of cadherin function causes the loss of contact to the neighboring endothelial cells and thus could represent an important mechanism in HCMV-induced cellular transendothelial migration and disruption of the endothelial integrity.
Intervirology 02/1999; 42(5-6):350-6. · 2.34 Impact Factor
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R A Blaheta,
K Leckel,
B Wittig,
D Zenker,
E Oppermann,
S Harder,
M Scholz,
S Weber,
H Schuldes,
A Encke,
B H Markus
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ABSTRACT: The novel immunosuppressive drug mycophenolate mofetil (CellCept, MMF) blocks DNA-synthesis by the inhibition of the enzyme inosine monophosphate dehydrogenase (IMDH). IMDH is also involved in the synthesis of adhesion receptors which are known to play an important role in the regulation of cell-cell contacts. Therefore, application of MMF might lead to a reduction of cellular infiltrates in the course of transplant rejection. To evaluate the therapeutic value of MMF, we investigated to what extent MMF blocks T-lymphocyte infiltration in vitro with regard to (a) adhesion to endothelial cells, (b) horizontal migration along these cells and (c) penetration through the endothelial cells. The results demonstrated a strong inhibition of both CD4+ and CD8+ T-cell adhesion and penetration by MMF. The ID50 value for CD4+ T-cell adhesion was calculated to be 0.03 microM and the ID50 value for CD4+ T-cell penetration 1.21 microM. MMF did not significantly influence the horizontal migration of T-lymphocytes along the human vascular endothelial cell (HUVEC) borders. FACS-analysis revealed a diminished E-selectin and P-selectin expression on endothelial cell membranes in the presence of MMF. Although MMF did not interfere with the synthesis of T-cell adhesion ligands, the binding activity of lymphocytic leucocyte function associated antigen 1 (LFA-1), very late antigen 4 (VLA-4) and PSGL-1 (P-selectin glycoprotein ligand 1) to immobilized intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1 (VCAM-1) and P-selectin was impaired. Moreover, MMF prevented VLA-4 and PSGL-1 receptor accumulation on the membranes of T-cell pseudopodia. It can be concluded that MMF possesses potent infiltration blocking properties. MMF evoked down-regulation of specific endothelial membrane molecules and the loss of protein localization in the lymphocyte protrusions might be predominantly responsible for the observed blockade of cell adhesion and penetration.
Transplant Immunology 01/1999; 6(4):251-9. · 1.46 Impact Factor
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R A Blaheta,
N P Hailer,
N Brude,
B Wittig,
E Oppermann,
K Leckel,
S Harder,
M Scholz,
S Weber,
A Encke,
B H Markus
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ABSTRACT: Cyclosporin A reduces the mitotic activity of allosensitized lymphocytes, but fails to limit emigration of these cells into the donor organ. However, the modulation of both lymphocyte proliferation and infiltration are desirable characteristics of immunosuppressive therapy. The calcium-channel blocker, verapamil, has recently been shown to effectively prevent the transmigration of CD4+ and CD8+ T cells through allogeneic endothelium. Mibefradil (Ro 40-5967) represents a new generation of calcium antagonists with high potency and long-term activity. To evaluate the immunosuppressive potential of this drug, the influence of mibefradil on lymphocyte adhesion to, horizontal locomotion along, and penetration through allogeneic endothelium (HUVEC) was performed. When lymphocytes were prestimulated for 24 hr with mibefradil, adhesion and penetration were dose-dependently reduced. The adhesion ID50 values were 3.4 microM (CD4+ T cells) versus 9.2 microM (CD8+ T cells) and 2.1 microM (CD4+ T cells) versus 3.9 microM (CD8+ T cells) with regard to penetration. Mibefradil also effectively blocked horizontal locomotion. Specific down-regulation of T-cell binding to the P-selection receptor (ID50: CD4+ T cells, 0.8 microM: CD8+ T cells, 1.2 microM) and to the intracellular adhesion molecule-1 (ICAM-1) receptor (ID50: CD4+ T cells, 1.9 microM; CD8+ T cells, 1.5 microM) by mibefradil seems to be responsible for the decreased adhesion and penetration rates. Reduction of intracellular F-actin in T lymphocytes could diminish cell locomotion. In conclusion, the potent suppressive properties of mibefradil support its use as a co-medication in cyclosporin A-based immunosuppressive therapy.
Immunology 07/1998; 94(2):213-20. · 3.32 Impact Factor
