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Biomedical subjects

S Dias

Publications and source records attributed to S Dias.

At least 19 recordsLinked to original sources
PubMed2001

Inhibition of both paracrine and autocrine VEGF/ VEGFR-2 signaling pathways is essential to induce long-term remission of xenotransplanted human leukemias.

Antiangiogenic agents block the effects of tumor-derived angiogenic factors (paracrine factors), such as vascular endothelial growth factor (VEGF), on endothelial cells (EC), inhibiting the growth of solid tumors. However, whether inhibition of angiogenesis also may play a role in liquid tumors is not well established. We recently have shown that certain leukemias not only produce VEGF but also selectively express functional VEGF receptors (VEGFRs), such as VEGFR-2 (Flk-1, KDR) and VEGFR1 (Flt1), resulting in the generation of an autocrine loop. Here, we examined the relative contribution of paracrine (EC-dependent) and autocrine (EC-independent) VEGF/VEGFR signaling pathways, by using a human leukemia model, where autocrine and paracrine VEGF/VEGFR loops could be selectively inhibited by neutralizing mAbs specific for murine EC (paracrine pathway) or human tumor (autocrine) VEGFRs. Blocking either the paracrine or the autocrine VEGF/VEGFR-2 pathway delayed leukemic growth and engraftment in vivo, but failed to cure inoculated mice. Long-term remission with no evidence of disease was achieved only if mice were treated with mAbs against both murine and human VEGFR-2, whereas mAbs against human or murine VEGFR-1 had no effect on mice survival. Therefore, effective antiangiogenic therapies to treat VEGF-producing, VEGFR-expressing leukemias may require blocking both paracrine and autocrine VEGF/VEGFR-2 angiogenic loops to achieve remission and long-term cure.

Animals↗
PubMed2001

Clonal variation in phenotype and life span of human embryonic fibroblasts (MRC-5) transduced with the catalytic component of telomerase (hTERT).

Expression of telomerase (hTERT) in certain cell types has been shown to extend cellular life span without malignant transformation. We studied the phenotype of 26 telomerase-transduced fibroblast clones (TTFC) generated from a mass culture of hTERT retrovirally transduced MRC-5 cells. About two-thirds of the transduced clones senesced at the expected time or shortly thereafter, despite high levels of expression of telomerase and telomere length maintenance. The remaining one-third of the clones were "immortalized" (followed for over 200 cumulative population doublings). All clones maintained a nontransformed phenotype: contact inhibition, anchorage dependency, lack of tumor formation in nude mice, dose dependency to serum and growth factors, low expression of a matrix metalloproteinase associated with metastatic invasion (MMP-9) and high expression of its inhibitor TIMP-1, and no cytogenetic abnormalities by G-banding. In addition, fibroblast-specific biological parameters, such as colony size, production of collagenase, and response to MMC and gamma radiation were tightly regulated at the clonal and subclonal levels.

Animals↗
PubMed2001

Infection of endothelium with E1(-)E4(+), but not E1(-)E4(-), adenovirus gene transfer vectors enhances leukocyte adhesion and migration by modulation of ICAM-1, VCAM-1, CD34, and chemokine expression.

Intravascular introduction of replication-deficient adenoviral vectors (Advectors) provides an ideal model of delivery of transgenes for the treatment of various vascular abnormalities. On the basis of the knowledge that Advectors can induce inflammatory responses after intravascular administration, we speculated that cellular activation by Advector infection could directly modulate the endothelial cell (EC) adhesion molecule/chemokine expression repertoire. Infection of human umbilical vein ECs or bone marrow microvascular ECs with an E1(-)E4(+) Advector resulted in the upregulation of intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and CD34, but not E-selectin, P-selectin, CD36, CD13, CD44, HLA-DR or PECAM. Upregulation of ICAM-1, VCAM-1, and CD34 was apparent 12 hours after infection and persisted for weeks after infection. Selective induction of adhesion molecules was mediated by the presence of the E4 gene in the Advector, because infection of ECs with an E1(-)E4(-) Advector had no effect on adhesion molecule expression. ECs infected with E1(-)E4(+) Advector, but not those infected with E1(-)E4(-) Advector, supported the adhesion of leukocytes. Monoclonal antibodies to ICAM-1 and VCAM-1 inhibited adhesion of leukocytes to E1(-)E4(+)-infected ECS: Infection of the ECs with E1(-)E4(+) Advector, but not E1(-)E4(-) Advector, resulted in downregulation of expression of chemocytokines, including interleukin-8, MCP-1, RANTES, and GM-CSF. Nonetheless, a large number of leukocytes migrated through ECs infected with E1(-)E4(+), but not those infected with E1(-)E4(l-), in response to exogenous chemokines. These results demonstrate that infection of ECs with E1(-)E4(+) Advectors, but not E1(-)E4(-) Advectors, may directly augment inflammatory responses by upregulating expression of adhesion molecules and enhancing migration through Advector-infected ECs and suggest that E1(-)E4(-) Advectors may be a better choice for gene-transfer strategies directed to the ECS:

Adenoviridae↗
PubMed2001

Vascular endothelial growth factor and angiopoietin-1 stimulate postnatal hematopoiesis by recruitment of vasculogenic and hematopoietic stem cells.

Tyrosine kinase receptors for angiogenic factors vascular endothelial growth factor (VEGF) and angiopoietin-1 (Ang-1) are expressed not only by endothelial cells but also by subsets of hematopoietic stem cells (HSCs). To further define their role in the regulation of postnatal hematopoiesis and vasculogenesis, VEGF and Ang-1 plasma levels were elevated by injecting recombinant protein or adenoviral vectors expressing soluble VEGF(165), matrix-bound VEGF(189), or Ang-1 into mice. VEGF(165), but not VEGF(189), induced a rapid mobilization of HSCs and VEGF receptor (VEGFR)2(+) circulating endothelial precursor cells (CEPs). In contrast, Ang-1 induced delayed mobilization of CEPs and HSCs. Combined sustained elevation of Ang-1 and VEGF(165) was associated with an induction of hematopoiesis and increased marrow cellularity followed by proliferation of capillaries and expansion of sinusoidal space. Concomitant to this vascular remodeling, there was a transient depletion of hematopoietic activity in the marrow, which was compensated by an increase in mobilization and recruitment of HSCs and CEPs to the spleen resulting in splenomegaly. Neutralizing monoclonal antibody to VEGFR2 completely inhibited VEGF(165), but not Ang-1-induced mobilization and splenomegaly. These data suggest that temporal and regional activation of VEGF/VEGFR2 and Ang-1/Tie-2 signaling pathways are critical for mobilization and recruitment of HSCs and CEPs and may play a role in the physiology of postnatal angiogenesis and hematopoiesis.

Adenoviridae↗
PubMed2001

Vascular trauma induces rapid but transient mobilization of VEGFR2(+)AC133(+) endothelial precursor cells.

Bone marrow (BM)-derived circulating endothelial precursor cells (CEPs) are thought to play a role in postnatal angiogenesis. Emerging evidence suggests that angiogenic stress of vascular trauma may induce mobilization of CEPs to the peripheral circulation. In this regard, we studied the kinetics of CEP mobilization in two groups of patients who experienced acute vascular insult secondary to burns or coronary artery bypass grafting (CABG). In both burn and CABG patients, there was a consistent, rapid increase in the number of CEPs, determined by their surface expression pattern of vascular endothelial growth factor receptor 2 (VEGFR2), vascular endothelial cadherin (VE-cadherin), and AC133. Within the first 6 to 12 hours after injury, the percentage of CEPs in the peripheral blood of burn or CABG patients increased almost 50-fold, returning to basal levels within 48 to 72 hours. Mobilized cells also formed late-outgrowth endothelial colonies (CFU-ECs) in culture, indicating that a small, but significant, number of circulating endothelial cells were BM-derived CEPs. In parallel to the mobilization of CEPs, there was also a rapid elevation of VEGF plasma levels. Maximum VEGF levels were detected within 6 to 12 hours of vascular trauma and decreased to baseline levels after 48 to 72 hours. Acute elevation of VEGF in the mice plasma resulted in a similar kinetics of mobilization of VEGFR2(+) cells. On the basis of these results, we propose that vascular trauma may induce release of chemokines, such as VEGF, that promotes rapid mobilization of CEPs to the peripheral circulation. Strategies to improve the mobilization and incorporation of CEPs may contribute to the acceleration of vascularization of the injured vascular tissue.

AC133 Antigen↗
PubMed2001

Anagrelide metabolite induces thrombocytopenia in mice by inhibiting megakaryocyte maturation without inducing platelet aggregation.

OBJECTIVE: The mechanism for anagrelide's potent platelet lowering activity in human subjects is not well defined. Studies related to anagrelide function have been hampered by its lack of activity in nonhuman primates and water insolubility. In an effort to define the mechanism whereby anagrelide exerts its therapeutic effect, we identified a water-soluble metabolite (anagrelide.met). The availability of anagrelide.met allowed, for the first time, parallel in vitro and in vivo animal studies centered on the mechanisms by which anagrelide lowers platelet levels. MATERIALS AND METHODS: The effects of anagrelide.met on proliferation and maturation of mega-karyocytes (MKs) as well as platelet production were studied both in vitro and in vivo. RESULTS: Anagrelide.met is capable of blocking in vitro MK migration by 20% to 40%. At 100 ng/mL, anagrelide.met selectively blocked in vitro MK maturation, resulting in a 50% decrease in the total number of CD41a(+) MKs, corresponding with a 30% decrease in MK ploidy by day 10 and a 60% decrease by day 20. Daily intraperitoneal injections of anagrelide.met 100 microg into BALB/c mice was sufficient to significantly decrease platelet counts within 24 to 48 hours, stabilizing to 40 to 50% of normal levels by day 5. This was associated with a 45% decrease in the number of developing MKs and an increase in thrombopoietin levels. Anagrelide.met did not alter WBC counts, hematocrit, or bleeding time, or lead to any apparent signs of toxicity. Furthermore, unlike the parent anagrelide compound, anagrelide.met did not inhibit ADP-induced platelet aggregation even at high concentrations (10 microg/mL). CONCLUSIONS: We describe a cross-species reactive anagrelide metabolite that selectively inhibits MK maturation and migration, lowering platelet levels without influencing platelet aggregation.

Acetylcholinesterase↗
PubMed2001

Impaired recruitment of bone-marrow-derived endothelial and hematopoietic precursor cells blocks tumor angiogenesis and growth.

The role of bone marrow (BM)-derived precursor cells in tumor angiogenesis is not known. We demonstrate here that tumor angiogenesis is associated with recruitment of hematopoietic and circulating endothelial precursor cells (CEPs). We used the angiogenic defective, tumor resistant Id-mutant mice to show that transplantation of wild-type BM or vascular endothelial growth factor (VEGF)-mobilized stem cells restore tumor angiogenesis and growth. We detected donor-derived CEPs throughout the neovessels of tumors and Matrigel-plugs in an Id1+/-Id3-/- host, which were associated with VEGF-receptor-1-positive (VEGFR1+) myeloid cells. The angiogenic defect in Id-mutant mice was due to impaired VEGF-driven mobilization of VEGFR2+ CEPs and impaired proliferation and incorporation of VEGFR1+ cells. Although targeting of either VEGFR1 or VEGFR2 alone partially blocks the growth of tumors, inhibition of both VEGFR1 and VEGFR2 was necessary to completely ablate tumor growth. These data demonstrate that recruitment of VEGF-responsive BM-derived precursors is necessary and sufficient for tumor angiogenesis and suggest new clinical strategies to block tumor growth.

Animals↗
PubMed2001

Mobilization of endothelial and hematopoietic stem and progenitor cells by adenovector-mediated elevation of serum levels of SDF-1, VEGF, and angiopoietin-1.

The chemokine stroma-derived factor-1 (SDF-1) is produced within the bone marrow and mediates chemokinesis and chemotaxis on a variety of cell types that express the CXCR4 receptor. SDF-1-responsive cell types include monocytes and macrophages, B and T lymphocytes, platelets and megakaryocytes, and CD34+ cells, including both hematopoietic progenitors and stem cells. We have used intravenous injection of a replication-incompetent adenovector expressing the SDF-1 gene to elevate serum levels of SDF-1 in Balb/c and SCID mice. Within 3 to 5 days there was a marked leukocytosis, predominantly involving monocytes, and a three-fold increase in platelets. In addition, AdSDF-1 mobilized CFU-GM, CFU-s, and cells with long-term repopulating potential. We have identified a bone marrow-derived, circulating endothelial stem cell characterized by expression of the VEGFR2 (Flk-1/KDR). This cell exhibits a chemotactic and chemokinetic response to SDF-1 and VEGF. We have elevated serum levels of VEGF165 using intravenous adenovector gene delivery and compared this to an adenovector expressing angiopoietin-1 alone or in combination with VEGF. VEGF elevation was associated with rapid mobilization of hematopoietic stem and progenitor cells and a population of Flk-1-positive endothelial progenitors. In contrast angiopoietin induced a delayed mobilization of endothelial and hematopoietic progenitors. The combination of VEGF and angiopoietin produced a more prolonged elevation of these progenitors in the circulation with increased proliferation of capillaries and expansion of sinusoidal spaces in the marrow.

Adenoviridae↗
PubMed2000

Stromal-derived factor 1-induced megakaryocyte migration and platelet production is dependent on matrix metalloproteinases.

Despite the discovery of thrombopoietin (TPO) and its contribution to megakaryocytopoiesis, the exact mechanisms and sites of platelet production are unknown. It has been shown that mature megakaryocytes (MKs) functionally express the stromal-derived factor 1 (SDF-1) receptor, CXCR4. SDF-1-induced migration of mature MKs through endothelial cell layers results in increased platelet production. Because the migration of polyploid MKs from the bone marrow microenvironment requires remodeling of the perivascular extracellular matrix, it was hypothesized that mature polyploid MKs may express matrix metalloproteinases (MMPs), facilitating their exit into the bone marrow extravascular space. In this report, it is demonstrated that SDF-1 induces the expression and release of gelatinase B (MMP-9) by purified mature polyploid human MKs and an adeno-CXCR4-infected megakaryocytic cell line. Neutralizing antibody to MMP-9, but not MMP-2, blocked SDF-1-induced migration of MKs through reconstituted basement membrane, suggesting that expression of MMP-9 is critical for MK migration. Incubation of mature MKs with a synthetic MMP inhibitor, 5-phenyl-1,10-phenanthrolene, resulted in the inhibition of platelet formation, suggesting that the expression of MMPs is not only critical for megakaryocyte migration but also for subsequent platelet release. Confirming these results, adeno-SDF-1 injection into normal mice resulted in increased platelet counts, a process that could be blocked by a synthetic MMP inhibitor. These results suggest mobilization of MKs involves sequential expression and activation of chemokine receptors such as CXCR4, MMP-9, followed by transendothelial migration. MMP inhibitors may have potential use in the treatment of thrombotic and myeloproliferative disorders. (Blood. 2000;96:4152-4159)

Adult↗
PubMed2000

Arsenic trioxide induces dose- and time-dependent apoptosis of endothelium and may exert an antileukemic effect via inhibition of angiogenesis.

Arsenic trioxide (As(2)O(3)) has recently been used successfully in the treatment of acute promyelocytic leukemia and has been shown to induce partial differentiation and apoptosis of leukemic cells in vitro. However, the mechanism by which As(2)O(3) exerts its antileukemic effect remains uncertain. Emerging data suggest that the endothelium and angiogenesis play a seminal role in the proliferation of liquid tumors, such as leukemia. We have shown that activated endothelial cells release cytokines that may stimulate leukemic cell growth. Leukemic cells, in turn, can release endothelial growth factors, such as vascular endothelial growth factor (VEGF). On the basis of these observations, we hypothesized that As(2)O(3) may interrupt a reciprocal loop between leukemic cells and the endothelium by direct action on both cell types. We have shown that treatment of proliferating layers of human umbilical vein endothelial cells (HUVECs) with a variety of concentrations of As(2)O(3) results in a reproducible dose- and time-dependent sequence of events marked by change to an activated morphology, up-regulation of endothelial cell adhesion markers, and apoptosis. Also, treatment with As(2)O(3) caused inhibition of VEGF production in the leukemic cell line HEL. Finally, incubation of HUVECs with As(2)O(3) prevented capillary tubule and branch formation in an in vitro endothelial cell-differentiation assay. In conclusion, we believe that As(2)O(3 )interrupts a reciprocal stimulatory loop between leukemic cells and endothelial cells by causing apoptosis of both cell types and by inhibiting leukemic cell VEGF production. (Blood. 2000;96:1525-1530)

Antineoplastic Agents↗
PubMed2000

Exploiting changes in the tumour microenvironment with sequential cytokine and matrix metalloprotease inhibitor treatment in a murine breast cancer model.

The study of treatment-induced changes in the tumour microenvironment might lead to effective combinations of biological therapy. IL-12 induced tumour regression and cure of an experimental murine breast cancer, HTH-K, but only after long-term treatment that was associated with chronic toxicity. During IL-12 therapy, tumour levels of the matrix metalloprotease MMP-9 declined and its inhibitor TIMP-1 was strongly induced. We therefore administered alternate cycles of IL-12 and the MMP inhibitor Batimastat (BB94) to mice. Therapeutic efficacy was increased compared with short-term IL-12 therapy but without the chronic toxicity associated with long-term IL-12 treatment. Image analysis of treated tumours revealed that BB94 prevented regeneration of tumour and stromal compartments that normally occurred after short-term IL-12 therapy.

Angiogenesis Inhibitors↗
PubMed2000

Autocrine stimulation of VEGFR-2 activates human leukemic cell growth and migration.

Emerging data suggest that VEGF receptors are expressed by endothelial cells as well as hematopoietic stem cells. Therefore, we hypothesized that functional VEGF receptors may also be expressed in malignant counterparts of hematopoietic stem cells such as leukemias. We demonstrate that certain leukemias not only produce VEGF but also express functional VEGFR-2 in vivo and in vitro, resulting in the generation of an autocrine loop that may support leukemic cell survival and proliferation. Approximately 50% of freshly isolated leukemias expressed mRNA and protein for VEGFR-2. VEGF(165) induced phosphorylation of VEGFR-2 and increased proliferation of leukemic cells, demonstrating these receptors were functional. VEGF(165) also induced the expression of MMP-9 by leukemic cells and promoted their migration through reconstituted basement membrane. The neutralizing mAb IMC-1C11, specific to human VEGFR-2, inhibited leukemic cell survival in vitro and blocked VEGF(165)-mediated proliferation of leukemic cells and VEGF-induced leukemic cell migration. Xenotransplantation of primary leukemias and leukemic cell lines into immunocompromised nonobese diabetic mice resulted in significant elevation of human, but not murine, VEGF in plasma and death of inoculated mice within 3 weeks. Injection of IMC-1C11 inhibited proliferation of xenotransplanted human leukemias and significantly increased the survival of inoculated mice. Interruption of signaling by VEGFRs, particularly VEGFR-2, may provide a novel strategy for inhibiting leukemic cell proliferation.

Animals↗
PubMed2000

Mental health in medical students. A case control study using the 60 item General Health Questionnaire.

This paper describes a cross-sectional case control study to measure the prevalence of psychological morbidity in first year medical students and compare it to the prevalence in in a randomly selected control group of other first year students at Edinburgh University. The study was conducted anonymously using the 60 item General Health Questionnaire. Participation rates were over 90% in both subjects and controls. A total of 17% of medical students had symptoms of psychological morbidity which may benefit from treatment and a further 29% of medical students had symptoms of psychological distress which would be expected to remit spontaneously. A similar rate was found in the control group of students. This suggests that if medical students or doctors, later in their careers, fare badly in terms of mental health then this may well be related to aspects of their lives and is not an intrinsic characteristic.

Case-Control Studies↗
PubMed1999

Ethanol and erythrocyte membrane interaction: a hemorheologic perspective.

Previous studies have documented structural and functional changes induced by ethanol-erythrocyte membrane interaction. In order to perform an in vitro study on the effect of different ethanol concentrations on erythrocyte hemorheologic properties, blood samples were collected from 21 male donors at the Hospital of Santa Maria. Whole blood aliquots were incubated with ethanol solutions of rising concentrations. The following parameters were measured: erythrocyte aggregation, haemoglobin, carboxyhaemoglobin and methaemoglobin concentrations, hematocrit, plasma osmolality and erythrocyte membrane fluidity (fluorescence polarisation probes TMA-DPH and DPH). With ethanol blood concentrations of 45 mM a rise in plasma osmolality (0.352 Osm/kg H2O vs 0.310 Osm/kg H2O; p < 0.001) was verified. With 67 mM concentration a decrease of erythrocyte aggregation (11.03 vs 12.81; p < 0.05) and an increase in plasma osmolality (0.380 Osm/kg H2O vs 0.310 Osm/kg H2O; p < 0.001) were obtained. In conclusion, ethanol only changes erythrocyte aggregation for a concentration of 67 mM. These data could lead to future changes in therapeutic approaches to situations such as alcoholic coma.

Adult↗
PubMed1998

IL-12 regulates VEGF and MMPs in a murine breast cancer model.

In a murine model of breast cancer, IL-12 therapy exerts potent anti-angiogenic effects which contribute to tumor regression. After 7 days of treatment, levels of tumor VEGF protein decline markedly and are undetectable at 14 days. This decline is accompanied by a fall in MMP-9 and, as the tumors regress, an increase in its natural inhibitor, TIMP-1. A cell line established from the primary tumor produced VEGF in vitro. IFN-gamma reduced tumor cell production of VEGF over a 24-hr period in vitro, suggesting that IL-12-induced IFN-gamma may be responsible for the decline in VEGF levels in vivo. There is also in vitro evidence that IL-12 regulates stromal cell interactions, leading to decreased MMP-9 and increased TIMP-1 production. Thus, we suggest that at least 2 mechanisms are involved in IL-12 regulation of angiogenesis, removing the pro-angiogenic stimulus and blocking the release and activity of MMPs.

Animals↗
PubMed1998

Multiple molecular and cellular changes associated with tumour stasis and regression during IL-12 therapy of a murine breast cancer model.

IL-12 treatment of a murine transplantable breast carcinoma (HTH-K) led to tumour regression and cure which was related to the duration of treatment. We studied the sequential molecular and phenotypic changes in IL-12-treated tumours. IFN-gamma mRNA was detected 8 hr after the first treatment. mRNA expression for the IFN-gamma-inducible genes beta 2-microglobulin and indoleamine dioxygenase (IDO) was induced subsequently, together with the chemokine IP-10. IL-12-treated tumours had an abundant cellular infiltrate, consisting mainly of CD8+ T cells. mRNA for granzyme B and perforin also could be detected, suggesting that those cells were activated. After 7 days of daily therapy, tumours in IL-12-treated mice had a significant reduction in vasculature. Finally, the number of apoptotic tumour cells increased throughout IL-12 treatment. We compared the anti-tumour effects of IL-12 to those induced by IFN-gamma therapy, which caused initial tumour stasis but subsequent tumour progression. IFN-gamma induced beta 2-microglobulin and IDO over a 7-day period, but IP-10 was induced only transiently. IFN-gamma caused a lesser cellular infiltrate, a minor anti-angiogenic effect and a transient apoptotic effect. The success of IL-12 may be due to its ability to produce a distinct sequence of molecular and phenotypic changes in tumours, leading to an anti-tumour immune response, toxicity against tumour cells and an anti-angiogenic effect. Other cytokines, such as IFN-gamma, induce some, but not all, of these actions. Comparison of IL-12 and IFN-gamma suggests that sustained induction of IP-10 and activation of a resulting cellular infiltrate may be key changes in regressing tumours.

Animals↗