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M R Shurin

Publications and source records attributed to M R Shurin.

At least 19 recordsLinked to original sources
PubMed2005

Comparative analysis of CD1a, S-100, CD83, and CD11c human dendritic cells in normal, premalignant, and malignant tissues.

A number of antibodies that recognize human dendritic cells (DC) have been identified. The main aim of this study was to compare and contrast different antigen retrieval techniques using both enzymatic and non-enzymatic treatments in order to determine the expression and distribution of several DC markers on formalin-fixed, paraffin-embedded tissues. Normal human lung, oral epithelial hyperplasia lesions, oral squamous cell carcinoma, and prostate adenocarcinoma tissues were evaluated using a panel of DC specific antibodies. The results of immunohistochemical staining for CD83, CD1a, CD11c, and S-100 DC markers were compared following the different antigen retrieval approaches. The overall best results for the analysis of tumor-associated DC were obtained with the enzymatic methods. Protease XXIV digestion was determined to be essential for detection of S-100 and CD11c positive DC, whereas trypsin and pepsin were required for the recognition of CD1a and CD83 expressing tumor-associated DC. These results could be easily adapted for routine practice and should be useful for characterization of the DC system in cancer patients for both diagnostic and prognostic purposes. In addition, standardized procedures for evaluating different subpopulations of tumor-associated DC should bring new insights in understanding of DC-tumor cell interaction.

Antigens, CD↗
PubMed2001

Mechanisms of dendritic cell-induced T cell proliferation in the primary MLR assay.

DC are unique antigen presenting cells, and their ability to induce proliferation of T cells in a mixed leukocyte reaction (MLR) assay is commonly used for the evaluation of their function. To determine the mechanisms involved in DC-induced T cell activation in a primary MLR assay, a variety of different agents were examined in this study that interfere with DNA synthesis, membrane organization, protein synthesis, and maturation induced by bacterial products. While only live DC were able to induce T cell proliferation in the MLR assay, irradiation of DC did not influence their stimulatory capacity. Fixation of DC membrane with paraformaldehyde resulted in a loss of DC capacity to induce T cell proliferation demonstrating that physical organization of the plasma membranes plays an important role in the induction of T cell activation. In addition, the pretreatment of DC with cycloheximide revealed that protein synthesis was not critical for the ability of DC to activate T cells. Finally, Staphylococcus aureus-mediated activation of DC significantly increased T cell proliferation and this effect was not dependent on IL-12 production of DC since DC generated from IL-12 knockout mice were not different from wild type DC. In summary, these data suggest that DC membrane structures are responsible for the antigen presentation and co-stimulation and play a key role in T cell recognition and activation by DC.

Animals↗
PubMed2001

Identification of delta- and mu-type opioid receptors on human and murine dendritic cells.

The purpose of this study was to evaluate mu- and delta-opioid receptors (OR) on human and murine dendritic cells (DC). Expression of mu- and delta-OR mRNA on DC was demonstrated by RT-PCR. The immunocytochemical and Western blot analyses revealed the expression of OR protein in DC. Radioreceptor assay demonstrated the specific saturated temperature-dependent binding of [3H]-labeled opioid ligand on DC and B(max)=2.8+/-0.3 fmol/10(6) cells and K(D)=4.8+/-1.0 nM were calculated by a Scatchard analysis. Finally, OR ligands DADLE and DAGO dose-dependently modulated the capacity of DC to induce T cell proliferation in an MLR assay. Importantly, expression of functional OR on DC was significantly increased upon TNF-alpha-induced DC maturation. Thus, these data suggest a new mechanism of opioid-dependent neuroendocrine immunomodulation.

Animals↗
PubMed2001

Human prostate cancer regulates generation and maturation of monocyte-derived dendritic cells.

BACKGROUND: The progression of prostate cancer is accompanied by a marked suppression of the immune system, including the apoptotic death of dendritic cells (DC) responsible for the induction of antitumor immunity. In this study, we evaluated whether prostate cancer might inhibit DC generation and maturation in vitro. METHODS: DC were generated from peripheral blood monocytes in the presence of the human prostate cell line LNCaP or nonmalignant cells, and characterized by light microscopy, FACScan analysis, and ability to stimulate T-cell proliferation. RESULTS: Prostate cancer significantly inhibited the conversion of monocytes into DC, which was assessed by the expression of DC markers CD1a and CD83. These cells were weak stimulators of T-cell proliferation, suggesting that DC generated in the prostate cancer microenvironment are functionally inhibited. CONCLUSIONS: Prostate cancer not only kills mature DC, but also inhibits their generation and maturation, resulting in decreased production of antigen-presenting cells and inhibition of their functional activity.

Antigens, CD↗
PubMed2001

Neuroblastoma-derived gangliosides inhibit dendritic cell generation and function.

Neuroblastoma (NB), a tumor of the sympathetic nervous system, is the most common extracranial solid tumor in children. NB-derived gangliosides inhibit the functional activity of T and natural killer cells, contribute to tumor-induced bone marrow suppression, and cause multiple alterations of hematopoiesis, resulting in pancytopenia. However, the role of gangliosides in the regulation of dendritic cell (DC) generation (dendropoiesis) has not been studied. Using murine and human NB cell lines, we demonstrated that coincubation of murine bone marrow progenitors or human CD34+ progenitor cells with NB cells resulted in a significant inhibition of dendropoiesis in vitro up to 90%. The number of DCs was assessed by FACScan determination of CD83+ or CD11c+ cells coexpressing MHC class II and CD86 molecules. In addition, inhibition of antigen-presenting properties of DCs cultured in the presence of NB cells was observed in allogeneic mixed leukocyte reaction (33,508 +/- 1,613 cpm for control DCs versus 17,428 +/- 152 cpm for NB-treated DCs; P < 0.05). Treatment of NB cells with 10 microM DL-threo-1-phenyl-2-decanolylamine-3-morpholino-1-propanol HCl, an inhibitor of glucosylceramide synthase, markedly abrogated ganglioside synthesis and was accompanied by blockade of NB ability to inhibit dendropoiesis. Furthermore, purified gangliosides added to DC cultures significantly inhibited DC generation. The percentage of CD83+ cells decreased from 51.8 +/- 6.1% in the control group to 12.9 +/- 2.7% in cultures treated with GD2 (P < 0.05). Thus, our results demonstrate that NB-derived gangliosides inhibit the generation of functionally active DCs and may play a role in tumor-induced immunosuppression and subsequent tumor escape from immune recognition and elimination.

Animals↗
PubMed2001

Human small cell lung carcinoma and carcinoid tumor regulate dendritic cell maturation and function.

The induction of apoptosis in dendritic cells (DC) is a key mechanism by which tumors escape immune recognition and elimination. In fact, a number of studies have showed the correlation between the number of DC within the tumor and the clinical prognosis, suggesting that increased infiltration of tumor tissue by DC was associated with better patient survival and low incidence of metastatic disease. We compared the number of DC and their distribution pattern in human small-cell lung carcinoma and bronchial carcinoid tumor (CT) tissues. Immunohistochemical analysis revealed the presence of cells expressing DC markers CD1a and CD83 in small-cell lung carcinoma tissues and the complete absence of these cells in CT samples. Next, we examined whether human lung tumor cells produce soluble factors that inhibit differentiation of hematopoietic precursors into mature DC. The addition of small-cell lung carcinoma-conditioned medium to CD34+ precursor cell cultures significantly inhibited colony-forming units of DC formation when compared with nontreated control DC cultures. Furthermore, DC generation and differentiation was completely abrogated in CD34+ cell cultures treated with CT-conditioned medium, suggesting that CT-derived factors blocked CD34+ cell differentiation into DC or induced their apoptosis. Finally, flow cytometry analysis of cultured DC confirmed these results. Thus, analysis of our data suggests that human lung tumors produce factors that inhibit DC generation or maturation and may also induce apoptotic death of DC precursors in vitro.

Antigens, CD↗
PubMed2001

TNF-alpha protects dendritic cells from prostate cancer-induced apoptosis.

We have recently shown that human prostate cancer (PCa) cells induced apoptotic death of the most potent antigen-presenting cells, dendritic cells (DC), which are responsible for the induction of specific antitumor immune responses. Here we have evaluated the effect of murine PCa cells RM-1 on the survival of immature and tumor necrosis factor-alpha (TNF-alpha)-stimulated mature DC. PCa cells and DC were co-incubated for 24-48 h and DC apoptosis was assessed by morphologic criteria, Annexin V assay, and TUNEL staining. We have shown that co-incubation of RM-1 cells with DC is accompanied by an increased level of DC apoptosis, which was mediated by decreased expression of anti-apoptotic protein Bcl-2. Stimulation of DC maturation by TNF-alpha resulted in increased resistance of DC to PCa-induced apoptosis. In TNF-alpha treated mature DC, but not in immature DC, the expression of Bcl-2 was not blocked after exposure to RM-1-derived factors. Thus, these data suggest that TNF-alpha-induced maturation of DC increases their resistance to PCa induced apoptosis. This is likely to be due to the stabilizing of the expression of anti-apoptotic protein Bcl-2. The difference in the sensitivity of mature and immature DC to PCa-induced cell death should be considered during the design of DC-based clinical trials for PCa patients.Prostate Cancer and Prostatic Diseases (2001) 4, 221-227.

Journal Article↗
PubMed2001

Effects of vitamin D (calcitriol) on transitional cell carcinoma of the bladder in vitro and in vivo.

PURPOSE: Vitamin D (calcitriol) has significant antiproliferative effects on various tumor cells in vitro and in vivo. In the clinical situation a major impediment to systemic administration of calcitriol is the side effect of hypercalcemia. To test the potential usefulness of calcitriol for bladder cancer treatment, we studied the antiproliferative effect of vitamin D on 2 human bladder cancer cell lines, 253j and T-24, in vitro. We also examined the in vivo effects of calcitriol in an animal model of bladder cancer using intravesical administration to avoid the toxicity of systemic calcitriol therapy. MATERIALS AND METHODS: The presence of vitamin D receptors in normal and neoplastic human bladder tissue, and tumor cells T-24 and 253j was determined by immunoblot analysis. Tumor cell proliferation in the presence or absence of calcitriol was determined using a crystal violet assay. Calcitriol induced apoptosis was determined by morphology, polyadenosine diphosphate ribose polymerase cleavage and annexin V binding. In vivo studies were performed by weekly intravesical instillation of calcitriol in female Fischer 344 rats after induction of tumors by N-methyl nitrosourea. Calcitriol administration was started 3 weeks after tumor induction for 7 doses at weekly intervals. RESULTS: Normal and neoplastic human bladder tissue, and the cell lines expressed vitamin D receptors. In the 253j and T-24 cell lines proliferation was significantly inhibited by calcitriol. Progressive cleavage of full length polyadenosine diphosphate ribose polymerase was observed in calcitriol treated cells starting as early as 4 hours after exposure. Similar changes were not observed in the control cells treated with vehicle (ethanol) alone. After 24 hours of treatment with calcitriol 45.8% of 253j cells bound annexin compared to 16.5% of control cells (chi-square p <0.001). Of the control animals 66% developed bladder tumors and 55% of the animals treated with calcitriol early (3 weeks) after tumor induction developed bladder tumors. Almost all of the tumors that developed in the calcitriol group were unifocal, and only 20% were invasive compared to 50% of those in the control animals. CONCLUSIONS: These results demonstrate that calcitriol inhibits proliferation and induces apoptosis in human bladder tumor cells in vitro, and may have therapeutic potential in bladder cancer. In vivo studies using an N-methylnitrosourea induced model of bladder cancer demonstrate that early institution of intravesical calcitriol therapy after carcinogen exposure results in fewer tumors, which are also less likely to be multifocal, high grade or invasive. With our protocol a short course of intravesical calcitriol administration did not result in any significant toxicity.

Administration, Intravesical↗
PubMed2001

Tumor necrosis factor-alpha-promoted expression of Bcl-2 and inhibition of mitochondrial cytochrome c release mediate resistance of mature dendritic cells to melanoma-induced apoptosis.

Melanoma escapes host defenses through a variety of means, including the elimination of immune effector cells within the tumor microenvironment. We have reported recently that murine and human tumors including melanoma induce premature apoptosis of dendritic cells both in vitro and in vivo. In this study, we have demonstrated that overexpression of the Bcl-2 protein family member Bcl-xL rescued murine dendritic cells (DCs) from melanoma-induced death in vitro. Another successful protection approach was tumor necrosis factor (TNF)-alpha-promoted sustained expression of the antiapoptotic protein Bcl-2 within dendritic cells. This effect of TNF-alpha was mediated by inhibition of mitochondrial cytochrome c release. Thus, both Bcl-xL and Bcl-2 enhance survival of dendritic cells within the tumor microenvironment. In addition, mature DCs were more resistant to melanoma-induced apoptosis than immature dendritic cells. This finding suggests a stage-dependent sensitivity of DCs to tumor-induced cell death. We conclude that: (a) mature DCs might be more suitable for the use of cancer vaccination; and (b) Bcl-2 protein family members such as Bcl-xL and Bcl-2 rescue DCs from tumor-induced premature apoptosis.

Animals↗
PubMed2000

Transduction of dendritic cells with Bcl-xL increases their resistance to prostate cancer-induced apoptosis and antitumor effect in mice.

We have shown that prostate cancer (PCa) causes apoptosis of dendritic cells (DC), which might block the development of specific antitumor immune responses. Analysis of murine prostatic carcinoma tissues revealed the significant decrease in intratumoral DC number during tumor progression. We demonstrated that the cytokine-mediated increase in DC survival was accompanied by an elevated expression of the anti-apoptotic protein Bcl-xL. Next, we evaluated the resistance to tumor-induced apoptosis and the antitumor efficiency of genetically engineered DC overexpressing Bcl-xL. DC were transduced with an adenoviral vector encoding the murine Bcl-xL gene and injected intratumorally. Data analysis revealed that treatment of PCa-bearing mice with Bcl-xL-transduced DC resulted in significant inhibition of tumor growth compared with the administration of nontransduced DC. Thus, our data suggest that the protection of DC from PCa-induced apoptosis might significantly increase the efficacy of DC-based therapies in cancer even in the absence of available tumor-specific Ags.

Animals↗
PubMed2000

Interleukin-12 and Flt3 ligand differentially promote dendropoiesis in vivo.

Interleukin-12 (IL-12) and Flt3 ligand (FL) regulate hematopoiesis by stimulating proliferation, differentiation and migration of progenitor and/or stem cells. In this study, we evaluated the in vivo effect of IL-12 alone or in combination with FL on dendritic cell (DC) generation and accumulation in murine spleen, lymph node, thymus, liver, and tumor tissues. Both cytokines induced accumulation of functional DC in lymphoid and non-lymphoid tissues. IL-12 promoted predominantly myeloid dendropoiesis, while FL induced both myeloid and lymphoid dendropoiesis. Combination treatment resulted in a dramatic increase in CD86+, and particularly, NLDC-145+ cells within the liver, which was largely due to cell proliferation. Combination therapy also revealed the ability of FL to protect bone marrow cell populations from IL-12-induced depletion in vivo. In vitro, we found a significant FL-induced up-regulation of IL-12 production by DC at both mRNA and protein levels. Thus, our study suggests that (i) the antitumor activity of IL-12 may, at least in part, be mediated by the stimulation of dendropoiesis and (ii) IL-12 might contribute to the antitumor activity of FL. Furthermore, induction of DC generation in vivo by a combination of IL-12 and FL might become a new approach for immunotherapy of cancer.

Animals↗
PubMed2000

Cytokine-mediated protection of human dendritic cells from prostate cancer-induced apoptosis is regulated by the Bcl-2 family of proteins.

Prostate cancer is the most common cancer in men in the United States, and second in cancer-induced mortality. It is likely that tumour-induced immunosuppression is one of the reasons for low treatment efficacy in patients with advanced prostate cancer. It has been recently demonstrated that prostate cancer tissue is almost devoid of dendritic cells (DC), the major antigen-presenting cells responsible for the induction of specific antitumour immune responses. In this study, we have tested the hypothesis that prostate cancer induces progressive suppression of the DC system. We found that co-incubation of human DC with three prostate cancer cell lines led to the high levels of premature apoptosis of DC, which were significantly higher than in DC cultures co-incubated with normal prostate cells or blood leucocytes. Stimulation of DC for 24 hours with CD40 ligand (CD154), IL-12 or IL-15 prior to their co-incubation with prostate cancer cells resulted in a significant increase in DC survival in the tumour microenvironment. Furthermore, activation of DC with these cytokines was also accompanied by increased expression of the anti-apoptotic protein Bcl-x(L) in DC, suggesting a possible mechanism involved in DC protection from apoptotic death. In summary, our data demonstrate that prostate cancer induces active elimination of DC in the tumour microenvironment. Stimulation of DC by CD154, IL-12 or IL-15 leads to an increased expression of the anti-apoptotic protein Bcl-x(L) and increased resistance of DC to prostate cancer-induced apoptosis. These results suggest a new mechanism of tumour escape from immune recognition and demonstrate the cytokine-based approaches which might significantly increase the efficacy of DC-based therapies for cancer.

Animals↗
PubMed1999

Effect of vascular endothelial growth factor and FLT3 ligand on dendritic cell generation in vivo.

The cytokine FLT3 ligand (FL) enhances dendritic cell (DC) generation and has therefore been proposed as a means to boost antitumor immunity. Vascular endothelial growth factor (VEGF) is produced by a large percentage of tumors and is required for development of tumor neovasculature. We previously showed that VEGF decreases DC production and function in vivo. In this study, we tested the hypothesis that VEGF regulates FL effects on DC generation. In seven experiments, four groups of mice were treated with PBS, VEGF alone (100 ng/h), FL alone (10 microgram/day), or with the combination of FL and VEGF. VEGF and PBS were administered continuously for 14 days via s.c. pumps. FL was given s.c. daily for 9 days, beginning on day 4. Tissues were collected and the number, phenotype, and function of lymph node, splenic, and thymic DCs were analyzed on day 14. As expected, treatment with FL resulted in a marked increase in the number of lymph node and spleen DCs and a smaller increase in thymic DC. Pretreatment of mice with VEGF inhibited these FL effects in lymph nodes and thymus by about 50%, whereas spleen DC numbers were undiminished by VEGF. VEGF treatment in vivo also inhibited the ability of FL to increase the number of hemopoietic precursor cells and the level of maturity exhibited by DC derived from these hemopoietic precursor cells in vitro. VEGF inhibited FL-inducible activation of transcription factor NF-kappaB. These data suggest that VEGF interferes with the ability of FL to promote dendritic cell differentiation from bone marrow progenitor cells in mice and therefore may decrease the therapeutic efficacy of FL in settings where increased numbers of DCs might provide clinical benefits.

Animals↗
PubMed1999

CD154 inhibits tumor-induced apoptosis in dendritic cells and tumor growth.

We have recently demonstrated that murine and human tumors induce apoptosis of dendritic cells (DC). Here, we evaluated the effect of CD40 ligation on the survival of tumor-associated DC and tumor growth. Retroviral transduction of MC38 colon carcinoma cells with the CD154 gene resulted in inhibition of tumor growth. This effect was abrogated in IL-12 knockout mice. Immunohistochemical analysis revealed an increase in CD11c+ (N418) and CD8+ but not NLDC-145+ cells in CD154-transfected tumors in wild-type mice. This increase was less pronounced in IL-12-deficient mice. In vitro, overexpression of CD154 on tumor cells significantly decreased the level of tumor-induced DC apoptosis. Surprisingly, the CD154-induced protection of DC from tumor-induced apoptosis was IL-12 independent in vitro, suggesting an IL-12-dependent and an IL-12-independent mechanism of CD154-induced anti-tumor immunity. Thus, our data suggest a new strategy to improve immunotherapy of cancer by protecting DC from tumor-induced apoptosis.

Animals↗
PubMed1999

Tumor's other immune targets: dendritic cells.

The induction of apoptosis in T cells is one of several mechanisms by which tumors escape immune recognition. We have investigated whether tumors induce apoptosis in dendritic cells (DC) by co-culture of murine or human DC with different tumor cell lines for 4-48 h. Analysis of DC morphological features, JAM assay, TUNEL, caspase-3-like and transglutaminase activity, Annexin V binding, and DNA fragmentation assays revealed a time- and dose-dependent induction of apoptosis in DC by tumor-derived factors. This finding is both effector and target specific. The mechanism of tumor-induced DC apoptosis involved regulation of Bcl-2 and Bax expression. Double staining of both murine and human tumor tissues confirmed that tumor-associated DC undergo apoptotic death in vivo. DC isolated from tumor tissue showed significantly higher levels of apoptosis as determined by TUNEL assay when compared with DC isolated from spleen. These findings demonstrate that tumors induce apoptosis in DC and suggest a new mechanism of tumor escape from immune recognition. DC protection from apoptosis will lead to improvement of DC-based immunotherapies for cancer and other immune diseases.

Animals↗