BENNY WEISS - Academia.edu (original) (raw)

Papers by BENNY WEISS

Research paper thumbnail of El caseinato de sodio induce la diferenciación de las células hematopoyéticas multipotenciales 32D

INTRODUCCIÓN La hematopoyesis es controlada por un grupo de cito-cinas conocidas como factores de... more INTRODUCCIÓN La hematopoyesis es controlada por un grupo de cito-cinas conocidas como factores de crecimiento hematopo-yético e interleucinas, 1,2 sin embargo, existen biomolé-culas de origen y naturaleza distinta a las citocinas que afectan la proliferación y diferenciación de las células hematopoyéticas, como por ejemplo el ácido retinoico, miembro de la familia de las hormonas esteroides. 3,4 Sodium caseinate induces differentiation of the multipotential hemopoietic cell line 32D ABSTRACT Aim. To determine the role of sodium caseinate (CasNa) in the modulation of hemopoiesis. Materials and methods. 32D cells, a murine hemopoietic multipotential cell line dependent on interleukin-3 (IL-3) for proliferation and survival, were used. These cells were cultured with 0.5 ng/mL of IL-3, together with different concentrations of CasNa. We evaluated: proliferation (direct counting under the microscope and use of thymidine 3 H), morphological differentiation (giemsa staining), cytochemistry...

Research paper thumbnail of Differences in surface membrane ecto-ATPase and ecto-AMPase in normal and malignant cells I. Decrease in ecto-ATPase in myeloid leukemic cells and the independent regulation of ecto-ATPase and ecto-AMPase

Journal of Cellular Physiology, 1977

The hydrolysis of ATP and AMP by enzymes located on the external side of the plasma membrane (ect... more The hydrolysis of ATP and AMP by enzymes located on the external side of the plasma membrane (ecto-ATPase and ecto-AMPase) was studied in mouse myeloid leukemic cells, normal early myeloid cells, and normal mature granulocytes and macrophages. Nine clones of myeloid leukemic cells were used belonging to three groups that differ in their ability to be induced to differentiate by the differentiation-inducing protein MGI. These three groups consisted of MGI+D+ that can be induced to undergo complete differentiation, MGI+D- that can be induced to partially differentiate and MGI-D- with no induction of differentiation. The ecto-ATPase activity of normal early myeloid cells was similar to that of normal mature granulocytes and macrophages and higher than that of any of the leukemic cells. Among the leukemic cells, the MGI-D- cells had the highest level of ecto-ATPase activity. The behaviour of ecto-AMPase differed from that of ecto-ATPase. Some MGI-D- clones had a higher ecto-AMPase activity than normal cells and MGI+D- and MGI+D+ cells showed no detectable activity. Neither the ecto-ATP-ase nor ecto-AMPase activities changed after induction of differentiation in normal early myeloid or MGI+D+ leukemic cells. The results indicate that the myeloid leukemic cells had a decreased ability to hydrolyse external ATP, that there can be an independent regulation of ecto-ATPase and ecto-AMPase and that neither of these enzyme activities changed during differentiation.

Research paper thumbnail of Interleukin-1 beta (IL-1β) induces tumor necrosis factor alpha (TNF-α) expression on mouse myeloid multipotent cell line 32D cl3 and inhibits their proliferation

Research paper thumbnail of Sodium caseinate induces expression and secretion of murine multipotent myeloid cell line 32D macrophage colony-stimulating factor

Archives of Medical Research, 2004

Background. Evidence that sodium caseinate (CasNa) is capable of inhibiting proliferation of hema... more Background. Evidence that sodium caseinate (CasNa) is capable of inhibiting proliferation of hematopoietic precursor cell line 32D and inducing its differentiation into macrophage cells has recently been published. Taking into consideration that hematopoiesis is regulated by growth factors and that macrophage colony-stimulating factor (M-CSF) is a wellknown growth factor that induces differentiation of macrophages, in this work we evaluated whether CasNa is capable of inducing expression and secretion of M-CSF in 32D cells. Methods. We cultured 32D cells in presence and absence of CasNa and compared their proliferation and viability. RNA was extracted from cell lysates to evaluate expression of the gene for M-CSF and its receptor. Cultured conditioned media was used to evaluate presence of M-CSF. Results. Our results showed that CasNa inhibited proliferation of 32D cells and that conditioned media (CM) of these cultures contained M-CSF-like activity. Presence of M-CSF in CM was detected by inhibiting M-CSF activity with anti-M-CSF and presence of this growth factor was confirmed by ELISA assay. We also provided evidence that CasNa induced expression of mRNA for M-CSF in 32D cells as well as increased expression of mRNA for its receptor. Conclusions. CasNa inhibits proliferation of 32D cells and induces expression of the gene for M-CSF and that of its receptor. It also induces secretion of the bioactive form of M-CSF. Ć 2004 IMSS. Published by Elsevier Inc.

Research paper thumbnail of Sodium Caseinate (CasNa) Induces Mobilization of Hematopoietic Stem Cells in a BALB/c Mouse Model

Medical Science Monitor Basic Research, 2015

BACKGROUND Hematopoietic stem cells transplantation has high clinical potential against a wide va... more BACKGROUND Hematopoietic stem cells transplantation has high clinical potential against a wide variety of hematologic, metabolic, and autoimmune diseases and solid tumors. Clinically, hematopoietic stem cells derived from peripheral blood are currently used more than those obtained from sources such as bone marrow. However, mobilizing agents used in the clinic tend to fail in high rates, making the number of mobilized cells insufficient for transplantation. We investigated whether sodium caseinate induces functional mobilization of hematopoietic stem cells into peripheral blood of Balb/c mice. MATERIAL AND METHODS Using a mouse model, we administrated sodium caseinate or Plerixafor, a commercial mobilizing agent, and analyzed counts of hematopoietic stem cells in peripheral blood, and then cells were transplanted into lethally irradiated mice to restore hematopoiesis. All assays were performed at least twice. RESULTS We found that sodium caseinate increases the number of mononuclear cells in peripheral blood with the immunophenotype of hematopoietic stem cells (0.2 to 0.5% LSK cells), allowing them to form colonies of various cell lineages in semisolid medium (p<0.05). This effect is similar to that of Plerixafor, and cells transplanted into lethally irradiated mice can restore hematopoiesis at higher percentages than mononuclear cells mobilized by Plerixafor (40% vs. 20%, respectively). Further, a secondary transplant rescued a separate group of irradiated mice from death, proving definitive evidence of hematopoietic reconstitution after hematopoietic stem cells transplantation. Data are presented as mean ± standard deviation. To determine significant differences between the data, one-way ANOVA and the Tukey test were used. CONCLUSIONS Collectively these results show the utility of sodium caseinate as a mobilizer of hematopoietic stem cells and its potential clinical application in transplantation settings.

Research paper thumbnail of Evaluation of the antitumor activity of Rinvanil and Phenylacetylrinvanil on the cervical cancer tumor cell lines HeLa, CaSKi and ViBo

European Journal of Pharmacology, 2015

Capsaicin is a potent inducer of apoptosis in tumor cells as well as a selective agonist for the ... more Capsaicin is a potent inducer of apoptosis in tumor cells as well as a selective agonist for the transient receptor potential vanilloid 1 (TRPV1). The present study determined the IC50 and cytotoxic and apoptotic activities of the Capsaicin analogues Rinvanil and Phenylacetylrinvanil (PhAR) on three cervical cancer cell lines: HeLa, CaSKi and ViBo. These analogues possess an increased affinity for TRPV1 receptors. The IC50 obtained proved to be cytotoxic for all three cell lines; however, in the cells treated with Capsaicin both active caspase-3 and nuclear fragmentation were present. Capsaicin and its analogues also inhibited the normal proliferation of lymphocytes, suggesting that they are non-selective antitumor compounds. Finally, we discuss the possible loss of the relation between apoptosis and affinity to TRPV1, and the need for other strategies to synthesize Capsaicin analogues that can be useful in cancer treatments.

Research paper thumbnail of Sodium caseinate induces increased survival in leukaemic mouse J774 model

In vivo (Athens, Greece)

Acute myeloid leukaemia is a neoplastic disease of haematopoietic stem cells. Although there have... more Acute myeloid leukaemia is a neoplastic disease of haematopoietic stem cells. Although there have been recent advances regarding its treatment, mortality remains high. Consequently, therapeutic alternatives continue to be explored. In the present report, we present evidence that sodium caseinate (CasNa), a salt of the principal protein in milk, may possess important anti-leukaemic properties. J774 leukaemia macrophage-like cells were cultured with CasNa and proliferation, viability and differentiation were evaluated. These cells were also inoculated into BALB/c mice as a model of leukemia. We demonstrated that CasNa inhibits the in vitro proliferation and reduces viability of J774 cells, and leads to increased survival in vivo in a leukaemic mouse model. These data indicate that CasNa may be useful in leukaemia therapy.

Research paper thumbnail of Implication of tyrosine kinase receptor and steel factor in cell density-dependent growth in cervical cancers and leukemias

Cancer research, Jan 15, 2001

Cell-cell interaction is important in the expansion of leukemic cells and of solid tumors. Steel ... more Cell-cell interaction is important in the expansion of leukemic cells and of solid tumors. Steel factor (SF) or Kit ligand is produced as a membrane-bound form (mSF) and a soluble form. Because both primary gynecological tumors and primary leukemic cells from patients with acute myeloblastic leukemia (AML) have been shown to coexpress c-Kit and SF, we addressed the question of whether mSF could contribute to cell interaction in these cancers. Investigations on primary cervical carcinomas have been hindered by the fact that the cells do not grow in culture. We report herein the establishment of two cervical carcinoma cell lines, CALO and INBL, that reproduce the pattern of SF/c-Kit expression observed in primary tumor samples. In addition, these cells exhibit marked density-dependent growth much in the same way as AML blasts. Using an antisense strategy with phosphorothioate-modified oligonucleotides that specifically target SF without affecting other surface markers, we provide dire...

Research paper thumbnail of An HPV 16 L1-based chimeric human papilloma virus-like particles containing a string of epitopes produced in plants is able to elicit humoral and cytotoxic T-cell activity in mice

Virology Journal, 2009

Background: Even though two prophylactic vaccines against HPV are currently licensed, infections ... more Background: Even though two prophylactic vaccines against HPV are currently licensed, infections by the virus continue to be a major health problem mainly in developing countries. The cost of the vaccines limits wide-scale application in poor countries. A promising strategy for producing affordable and efficient vaccines involves the expression of recombinant immunogens in plants. Several HPV genes have been expressed in plants, including L1, which can self-assemble into virus-like particles. A plant-based, dual prophylactic/therapeutic vaccine remains an attractive possibility.

Research paper thumbnail of A novel HPV 16 L1-based chimeric virus-like particle containing E6 and E7 seroreactive epitopes permits highly specific detection of antibodies in patients with CIN 1 and HPV-16 infection

Virology Journal, 2011

The presence of IgG antibodies to HPV-16 L1-virus like particles (VLPs) in serum has been reporte... more The presence of IgG antibodies to HPV-16 L1-virus like particles (VLPs) in serum has been reported as a result of persistent exposure to the virus and as a marker of disease progression. However, detection of VLPspecific antibodies in sera does not always indicate a malignant lesion as positive results may also be due to a nonmalignant viral infection. Furthermore, malignant lesions are associated with an increased antibody titer for E6 and E7 proteins. The aim of this study was to develop an ELISA using a novel chimeric virus-like particle (cVLP) encoding an L1 protein fused with a string of HPV-16 E6 and E7 seroreactive epitopes to its C-terminus to be used for detection of HPV-16 specific antibodies in patients with cervical intraepithelial lesion grade 1 (CIN 1).

Research paper thumbnail of Isolation of a Proliferation Inhibitor Factor From Uterine Myomatosis Fibroblasts

Preparative Biochemistry and Biotechnology, 1999

In this work, we report the isolation of a factor from the culture supernatant of confluent fibro... more In this work, we report the isolation of a factor from the culture supernatant of confluent fibroblasts from human cervix with the diagnosis of uterine myomatosis. This factor possesses the capacity to inhibit the proliferation of normal fibroblasts. The proliferation inhibitor factor (PIF) was purified from the culture supernatant by precipitation with 80% ammonium sulfate, and by molecular sieve chromatography. Our results indicate that PIF is a protein of 23 kDa, which is highly sensitive to trypsin treatment, and is thermolabile, since temperatures equal to, or above, 60 degrees C eliminate the protein activity in 15 to 20 min. Western blot analyses identified no cross reactions of the purified PIF with TGF-alpha, TNFalpha, IFNgamma, or IL-1beta, suggesting that PIF is a new protein belonging to the group of factors secreted by fibroblasts able to inhibit cellular proliferation.

Research paper thumbnail of Detection of antibodies against a human papillomavirus (HPV) type 16 peptide that differentiate high-risk from low-risk HPV-associated low-grade squamous intraepithelial lesions

Journal of General Virology, 2004

A nonapeptide (16L1) was derived from the human papillomavirus type 16 (HPV-16) major capsid prot... more A nonapeptide (16L1) was derived from the human papillomavirus type 16 (HPV-16) major capsid protein and tested for detection of potential cross-reactive serum IgG and cervical IgA antibodies in low-and high-risk HPV-associated low-grade squamous intraepithelial lesions (LSIL) and cervical cancer patients by ELISA. The IgG response was similar in women with low-risk HPV-associated LSIL and controls (P=0?1). In contrast, more than 90 % of patients with high-risk HPV-associated LSIL were seropositive. Although tumours from cancer patients were all positive for the presence of high-risk HPV DNA, the level of seropositivity decreased significantly in this group (P<0?0001). Cervical IgA antibodies were also detected in a significantly high proportion of women with high-risk HPV-associated LSIL compared with controls. However, the proportion of IgA-positive patients was lower than the proportion of IgG seropositives. In conclusion, the 16L1 peptide appears to be a high-risk type-common epitope that induces cross-reactive antibodies in high-risk, but not low-risk, HPV-associated LSIL patients, allowing differentiation of high-and low-risk infected women at this stage of infection.

Research paper thumbnail of Expression of MICA, MICB and NKG2D in human leukemic myelomonocytic and cervical cancer cells

Journal of Experimental & Clinical Cancer Research, 2011

Background: Cancer cells are known to secrete the stress molecules MICA and MICB that activate cy... more Background: Cancer cells are known to secrete the stress molecules MICA and MICB that activate cytotoxicity by lymphocytes and NK cells through their NKG2D receptor as a mechanism of immunological defense. This work was undertaken to evaluate if cancer cells can also express this receptor as a possible mechanisms of depletion of MIC molecules and thus interfere with their immune recognition.

Research paper thumbnail of Cationic liposomes bearing IL-2 on their external surface induced mice leukocytes to kill human cervical cancer cells in vitro , and significantly reduced tumor burden in immunodepressed mice

Journal of Drug Targeting, 2011

Research paper thumbnail of Peripheral blood lymphocytes from low-grade squamous intraepithelial lesions patients recognize vaccine antigens in the presence of activated dendritic cells, and produced high levels of CD8 + IFNγ + T cells and low levels of IL-2 when induced to proliferate

Infectious Agents and Cancer, 2012

Background: Most infections with human papillomavirus (HPV) are resolved without clinical interve... more Background: Most infections with human papillomavirus (HPV) are resolved without clinical intervention, but a minority evolves into chronic lesions of distinct grades, including cervical-uterine cancer. It is known that in most cases the immune system mediates elimination of HPV infection. However, the mechanism of immune evasion leading to HPV persistence and development of early cervical lesions is not fully understood. The aim of the present work was to evaluate the potential of peripheral blood leukocytes (PBL) from low-grade squamous intraepithelial lesions (LSIL) patients to be activated ex-vivo by vaccine antigens, the participation of cytotoxic lymphocytes and regulatory T cells, and to determine the secretion of Th1 and Th2 cytokines mediated by stimulation of T cell receptors. Results: We found that PBL from LSIL patients showed a significantly lower proliferation rate to vaccine antigens as compared to that of healthy donors, even though there was not a difference in the presence of antibodies to those antigens in sera from both groups. We did not find differences in either the frequency of CD4 + CD25 + FoxP3+ in PBL, or the levels of IL-4, IL-5 and IL-10 in plasma or conditioned media from PBL incubated with TcR agonists in vitro, between the two groups. However, we detected a lower production of IL-2 and a higher proportion of CD8 + IFNγ + cells in PBL from LSIL patients as compared with PBL from normal donors. We also observed that PBL from patients infected by HPV-16 and −18 were not able to proliferate in the presence of soluble HPV antigens added to the culture; however, a high level of proliferation was attained when these antigens were presented by activated dendritic cells.

Research paper thumbnail of Identification of peptides presented by HLA class I molecules on cervical cancer cells with HPV-18 infection

Immunology Letters, 1999

In this work we eluted peptides from purified class I MHC molecules, isolated from a novel human ... more In this work we eluted peptides from purified class I MHC molecules, isolated from a novel human cervical carcinoma cell line (INBL), generated in our laboratory and positive for HPV-18 infection. A fraction of these peptides was capable of stimulating T lymphocytes obtained from a donor matched for HLA-Cw4 and who was also HPV-18 + . Direct N-terminal Edman degradation of these peptides, revealed the sequence (XQFPIFLQF) that matched 85% with the sequence NVFPIFLQM localized in between the 54 and 62 residues of the HPV-18 L1 protein. After stimulation with the synthetic peptide NVFPIFLQM, T lymphocytes from the donor were capable to lyse INBL cells. Our results provide evidence of the existence of naturally occurring viral epitopes presented on cervical cancer cells by the HLA-Cw4 allele, that could be useful for immunotherapy on this type of patient.

Research paper thumbnail of Alpha-, beta- and kappa-caseins inhibit the proliferation of the myeloid cell lines 32D cl3 and WEHI-3 and exhibit different differentiation properties

Immunobiology, 2008

We have recently shown that sodium caseinate (CasNa) was able to inhibit the proliferation of the... more We have recently shown that sodium caseinate (CasNa) was able to inhibit the proliferation of the myeloid cell line 32D cl3 in a non-toxic way, and that it also induced the expression of macrophage colony-stimulating factor (M-CSF). Casein is the main protein present in milk and is composed of alpha (a), beta (b) and kappa (k) subunits. This work was undertaken to evaluate if any one casein is responsible for the proliferation and differentiation properties found for CasNa on myeloid cells. Taking into consideration that 32D cl3 cells are considered to be non-malignant and dependent on IL-3 for proliferation, we also included for this study a leukemic cell line, WEHI-3, that does not depend on any external growth factor for its proliferation in order to evaluate if the growth inhibitory effect of caseins is also present for malignant cells. Our results showed that all caseins were inhibitory for the proliferation of either 32D cl3 and WEHI-3 and that only the 32D cl3 cells were induced to differentiate into the monocyte-macrophage lineage. In order to evaluate if CasNa was able to inhibit the proliferation of other myeloid cells we used J774 and P388 and found that they were also inhibited. We also determined that the different caseins exhibit different differentiation properties, with a-casein being the only one able to induce the secretion of M-CSF. We consider this work to open a new field of research, where casein, or its components, can be studied for their possible role in hematopoiesis and on the inhibition of malignant cell proliferation for therapeutic use.

Research paper thumbnail of Sodium caseinate induces secretion of macrophage colony-stimulating factor from neutrophils

Immunobiology, 2010

In this work we provide evidence that granulocytes produce macrophage colony-stimulating factor (... more In this work we provide evidence that granulocytes produce macrophage colony-stimulating factor (M-CSF) in the band cell stage and secrete it upon sodium caseinate-mediated differentiation to polymorphonuclear cells. We identified M-CSF in an enriched population of myeloid band cells from murine bone marrow using a chromophorelabeled monoclonal anti-M-CSF antibody. An ELISA assay was then used to detect secreted M-CSF in culture supernatants of enriched band cells differentiated to mature neutrophils using sodium caseinate. Colony formation in vitro by the supernatants from differentiating band cells was blocked by anti-M-CSF, thus suggesting that this factor is the only one responsible for this activity. Our data imply that casein can modulate hematopoiesis possibly via M-CSF production. Finally we discuss the possibility whether this M-CSF in concert with G-CSF could establish a cellular communication network between macrophages and granulocytes allowing them to simultaneously arrive at the inflammatory site.

Research paper thumbnail of Evidence that cervical cancer cells secrete IL-2, which becomes an autocrine growth factor

Cytokine, 2010

We present evidence that cervical cancer cells express a functional IL-2 receptor (IL-2R). In fac... more We present evidence that cervical cancer cells express a functional IL-2 receptor (IL-2R). In fact, by RT-PCR we obtained that the IL-2R is present in CALO, and INBL cells, and that it consisted of the alphaIL-2R, betaIL-2R, and gammaIL-2R chains. We also found that IL-2 is a growth factor for these cell lines, and unexpectedly that CALO and INBL themselves being cancer cells produce, and secrete IL-2. Antibodies against the alpha and beta subunits of the IL-2R inhibited cell proliferation thus hinting to a cell growth dependency on this factor. Our results thus provide evidence that the IL-2R on cervical cancer cells is part of an autocrine mechanism for its growth to the extent that, like lymphocytes, they produce and become partially dependent on this growth factor. We think that in view of our results caution should be taken when IL-2 is being considered for cancer therapy; in particular when the patient&#39;s cancer cells present the IL-2R, because as indicated by our results, the use of this factor could promote tumor growth. Finally, the possible implications of the expression of both IL-2, and IL-2R on cervical cancer cells on the immune escape mechanism of tumor cells are discussed.

Research paper thumbnail of Interleukin-2 (IL-2) receptor-βγ signalling is activated by c-Kit in the absence of IL-2, or by exogenous IL-2 via JAK3/STAT5 in human papillomavirus-associated cervical cancer

Cellular Signalling, 2004

Activation of the interleukin-2 receptor (IL-2R) induces signalling cascades promoting T cell pro... more Activation of the interleukin-2 receptor (IL-2R) induces signalling cascades promoting T cell proliferation. However, signal transduction pathways triggered in IL-2R-expressing solid tumours are unknown. This report shows that human papillomavirus (HPV)-associated cervical cancer cells express an IL-2R composed of beta and gamma chains (IL-2Rbetagamma), and that IL-2-mediated activation increases the phosphorylation of JAK3 and STAT5, stimulating cell proliferation. Interestingly, endogenous IL-2 is not produced by these cells, suggesting the activation of IL-2Rbetagamma by an alternative mechanism. Accordingly, we found that Stem Cell Factor (SCF)-activated c-Kit induces phosphorylation of the IL-2Rbeta chain in the absence of IL-2. Moreover, inhibition of IL-2Rbeta phosphorylation by blocking c-Kit tyrosine kinase activity abolishes both, IL-2 and SCF-mediated proliferation. Thus, these results demonstrate that IL-2 triggers a JAK3/STAT5 cascade in HPV-associated cervical cancer cells expressing IL-2Rbetagamma, and that this receptor can be alternatively activated by SCF-activated c-Kit in the absence of IL-2.

Research paper thumbnail of El caseinato de sodio induce la diferenciación de las células hematopoyéticas multipotenciales 32D

INTRODUCCIÓN La hematopoyesis es controlada por un grupo de cito-cinas conocidas como factores de... more INTRODUCCIÓN La hematopoyesis es controlada por un grupo de cito-cinas conocidas como factores de crecimiento hematopo-yético e interleucinas, 1,2 sin embargo, existen biomolé-culas de origen y naturaleza distinta a las citocinas que afectan la proliferación y diferenciación de las células hematopoyéticas, como por ejemplo el ácido retinoico, miembro de la familia de las hormonas esteroides. 3,4 Sodium caseinate induces differentiation of the multipotential hemopoietic cell line 32D ABSTRACT Aim. To determine the role of sodium caseinate (CasNa) in the modulation of hemopoiesis. Materials and methods. 32D cells, a murine hemopoietic multipotential cell line dependent on interleukin-3 (IL-3) for proliferation and survival, were used. These cells were cultured with 0.5 ng/mL of IL-3, together with different concentrations of CasNa. We evaluated: proliferation (direct counting under the microscope and use of thymidine 3 H), morphological differentiation (giemsa staining), cytochemistry...

Research paper thumbnail of Differences in surface membrane ecto-ATPase and ecto-AMPase in normal and malignant cells I. Decrease in ecto-ATPase in myeloid leukemic cells and the independent regulation of ecto-ATPase and ecto-AMPase

Journal of Cellular Physiology, 1977

The hydrolysis of ATP and AMP by enzymes located on the external side of the plasma membrane (ect... more The hydrolysis of ATP and AMP by enzymes located on the external side of the plasma membrane (ecto-ATPase and ecto-AMPase) was studied in mouse myeloid leukemic cells, normal early myeloid cells, and normal mature granulocytes and macrophages. Nine clones of myeloid leukemic cells were used belonging to three groups that differ in their ability to be induced to differentiate by the differentiation-inducing protein MGI. These three groups consisted of MGI+D+ that can be induced to undergo complete differentiation, MGI+D- that can be induced to partially differentiate and MGI-D- with no induction of differentiation. The ecto-ATPase activity of normal early myeloid cells was similar to that of normal mature granulocytes and macrophages and higher than that of any of the leukemic cells. Among the leukemic cells, the MGI-D- cells had the highest level of ecto-ATPase activity. The behaviour of ecto-AMPase differed from that of ecto-ATPase. Some MGI-D- clones had a higher ecto-AMPase activity than normal cells and MGI+D- and MGI+D+ cells showed no detectable activity. Neither the ecto-ATP-ase nor ecto-AMPase activities changed after induction of differentiation in normal early myeloid or MGI+D+ leukemic cells. The results indicate that the myeloid leukemic cells had a decreased ability to hydrolyse external ATP, that there can be an independent regulation of ecto-ATPase and ecto-AMPase and that neither of these enzyme activities changed during differentiation.

Research paper thumbnail of Interleukin-1 beta (IL-1β) induces tumor necrosis factor alpha (TNF-α) expression on mouse myeloid multipotent cell line 32D cl3 and inhibits their proliferation

Research paper thumbnail of Sodium caseinate induces expression and secretion of murine multipotent myeloid cell line 32D macrophage colony-stimulating factor

Archives of Medical Research, 2004

Background. Evidence that sodium caseinate (CasNa) is capable of inhibiting proliferation of hema... more Background. Evidence that sodium caseinate (CasNa) is capable of inhibiting proliferation of hematopoietic precursor cell line 32D and inducing its differentiation into macrophage cells has recently been published. Taking into consideration that hematopoiesis is regulated by growth factors and that macrophage colony-stimulating factor (M-CSF) is a wellknown growth factor that induces differentiation of macrophages, in this work we evaluated whether CasNa is capable of inducing expression and secretion of M-CSF in 32D cells. Methods. We cultured 32D cells in presence and absence of CasNa and compared their proliferation and viability. RNA was extracted from cell lysates to evaluate expression of the gene for M-CSF and its receptor. Cultured conditioned media was used to evaluate presence of M-CSF. Results. Our results showed that CasNa inhibited proliferation of 32D cells and that conditioned media (CM) of these cultures contained M-CSF-like activity. Presence of M-CSF in CM was detected by inhibiting M-CSF activity with anti-M-CSF and presence of this growth factor was confirmed by ELISA assay. We also provided evidence that CasNa induced expression of mRNA for M-CSF in 32D cells as well as increased expression of mRNA for its receptor. Conclusions. CasNa inhibits proliferation of 32D cells and induces expression of the gene for M-CSF and that of its receptor. It also induces secretion of the bioactive form of M-CSF. Ć 2004 IMSS. Published by Elsevier Inc.

Research paper thumbnail of Sodium Caseinate (CasNa) Induces Mobilization of Hematopoietic Stem Cells in a BALB/c Mouse Model

Medical Science Monitor Basic Research, 2015

BACKGROUND Hematopoietic stem cells transplantation has high clinical potential against a wide va... more BACKGROUND Hematopoietic stem cells transplantation has high clinical potential against a wide variety of hematologic, metabolic, and autoimmune diseases and solid tumors. Clinically, hematopoietic stem cells derived from peripheral blood are currently used more than those obtained from sources such as bone marrow. However, mobilizing agents used in the clinic tend to fail in high rates, making the number of mobilized cells insufficient for transplantation. We investigated whether sodium caseinate induces functional mobilization of hematopoietic stem cells into peripheral blood of Balb/c mice. MATERIAL AND METHODS Using a mouse model, we administrated sodium caseinate or Plerixafor, a commercial mobilizing agent, and analyzed counts of hematopoietic stem cells in peripheral blood, and then cells were transplanted into lethally irradiated mice to restore hematopoiesis. All assays were performed at least twice. RESULTS We found that sodium caseinate increases the number of mononuclear cells in peripheral blood with the immunophenotype of hematopoietic stem cells (0.2 to 0.5% LSK cells), allowing them to form colonies of various cell lineages in semisolid medium (p&amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;amp;lt;0.05). This effect is similar to that of Plerixafor, and cells transplanted into lethally irradiated mice can restore hematopoiesis at higher percentages than mononuclear cells mobilized by Plerixafor (40% vs. 20%, respectively). Further, a secondary transplant rescued a separate group of irradiated mice from death, proving definitive evidence of hematopoietic reconstitution after hematopoietic stem cells transplantation. Data are presented as mean ± standard deviation. To determine significant differences between the data, one-way ANOVA and the Tukey test were used. CONCLUSIONS Collectively these results show the utility of sodium caseinate as a mobilizer of hematopoietic stem cells and its potential clinical application in transplantation settings.

Research paper thumbnail of Evaluation of the antitumor activity of Rinvanil and Phenylacetylrinvanil on the cervical cancer tumor cell lines HeLa, CaSKi and ViBo

European Journal of Pharmacology, 2015

Capsaicin is a potent inducer of apoptosis in tumor cells as well as a selective agonist for the ... more Capsaicin is a potent inducer of apoptosis in tumor cells as well as a selective agonist for the transient receptor potential vanilloid 1 (TRPV1). The present study determined the IC50 and cytotoxic and apoptotic activities of the Capsaicin analogues Rinvanil and Phenylacetylrinvanil (PhAR) on three cervical cancer cell lines: HeLa, CaSKi and ViBo. These analogues possess an increased affinity for TRPV1 receptors. The IC50 obtained proved to be cytotoxic for all three cell lines; however, in the cells treated with Capsaicin both active caspase-3 and nuclear fragmentation were present. Capsaicin and its analogues also inhibited the normal proliferation of lymphocytes, suggesting that they are non-selective antitumor compounds. Finally, we discuss the possible loss of the relation between apoptosis and affinity to TRPV1, and the need for other strategies to synthesize Capsaicin analogues that can be useful in cancer treatments.

Research paper thumbnail of Sodium caseinate induces increased survival in leukaemic mouse J774 model

In vivo (Athens, Greece)

Acute myeloid leukaemia is a neoplastic disease of haematopoietic stem cells. Although there have... more Acute myeloid leukaemia is a neoplastic disease of haematopoietic stem cells. Although there have been recent advances regarding its treatment, mortality remains high. Consequently, therapeutic alternatives continue to be explored. In the present report, we present evidence that sodium caseinate (CasNa), a salt of the principal protein in milk, may possess important anti-leukaemic properties. J774 leukaemia macrophage-like cells were cultured with CasNa and proliferation, viability and differentiation were evaluated. These cells were also inoculated into BALB/c mice as a model of leukemia. We demonstrated that CasNa inhibits the in vitro proliferation and reduces viability of J774 cells, and leads to increased survival in vivo in a leukaemic mouse model. These data indicate that CasNa may be useful in leukaemia therapy.

Research paper thumbnail of Implication of tyrosine kinase receptor and steel factor in cell density-dependent growth in cervical cancers and leukemias

Cancer research, Jan 15, 2001

Cell-cell interaction is important in the expansion of leukemic cells and of solid tumors. Steel ... more Cell-cell interaction is important in the expansion of leukemic cells and of solid tumors. Steel factor (SF) or Kit ligand is produced as a membrane-bound form (mSF) and a soluble form. Because both primary gynecological tumors and primary leukemic cells from patients with acute myeloblastic leukemia (AML) have been shown to coexpress c-Kit and SF, we addressed the question of whether mSF could contribute to cell interaction in these cancers. Investigations on primary cervical carcinomas have been hindered by the fact that the cells do not grow in culture. We report herein the establishment of two cervical carcinoma cell lines, CALO and INBL, that reproduce the pattern of SF/c-Kit expression observed in primary tumor samples. In addition, these cells exhibit marked density-dependent growth much in the same way as AML blasts. Using an antisense strategy with phosphorothioate-modified oligonucleotides that specifically target SF without affecting other surface markers, we provide dire...

Research paper thumbnail of An HPV 16 L1-based chimeric human papilloma virus-like particles containing a string of epitopes produced in plants is able to elicit humoral and cytotoxic T-cell activity in mice

Virology Journal, 2009

Background: Even though two prophylactic vaccines against HPV are currently licensed, infections ... more Background: Even though two prophylactic vaccines against HPV are currently licensed, infections by the virus continue to be a major health problem mainly in developing countries. The cost of the vaccines limits wide-scale application in poor countries. A promising strategy for producing affordable and efficient vaccines involves the expression of recombinant immunogens in plants. Several HPV genes have been expressed in plants, including L1, which can self-assemble into virus-like particles. A plant-based, dual prophylactic/therapeutic vaccine remains an attractive possibility.

Research paper thumbnail of A novel HPV 16 L1-based chimeric virus-like particle containing E6 and E7 seroreactive epitopes permits highly specific detection of antibodies in patients with CIN 1 and HPV-16 infection

Virology Journal, 2011

The presence of IgG antibodies to HPV-16 L1-virus like particles (VLPs) in serum has been reporte... more The presence of IgG antibodies to HPV-16 L1-virus like particles (VLPs) in serum has been reported as a result of persistent exposure to the virus and as a marker of disease progression. However, detection of VLPspecific antibodies in sera does not always indicate a malignant lesion as positive results may also be due to a nonmalignant viral infection. Furthermore, malignant lesions are associated with an increased antibody titer for E6 and E7 proteins. The aim of this study was to develop an ELISA using a novel chimeric virus-like particle (cVLP) encoding an L1 protein fused with a string of HPV-16 E6 and E7 seroreactive epitopes to its C-terminus to be used for detection of HPV-16 specific antibodies in patients with cervical intraepithelial lesion grade 1 (CIN 1).

Research paper thumbnail of Isolation of a Proliferation Inhibitor Factor From Uterine Myomatosis Fibroblasts

Preparative Biochemistry and Biotechnology, 1999

In this work, we report the isolation of a factor from the culture supernatant of confluent fibro... more In this work, we report the isolation of a factor from the culture supernatant of confluent fibroblasts from human cervix with the diagnosis of uterine myomatosis. This factor possesses the capacity to inhibit the proliferation of normal fibroblasts. The proliferation inhibitor factor (PIF) was purified from the culture supernatant by precipitation with 80% ammonium sulfate, and by molecular sieve chromatography. Our results indicate that PIF is a protein of 23 kDa, which is highly sensitive to trypsin treatment, and is thermolabile, since temperatures equal to, or above, 60 degrees C eliminate the protein activity in 15 to 20 min. Western blot analyses identified no cross reactions of the purified PIF with TGF-alpha, TNFalpha, IFNgamma, or IL-1beta, suggesting that PIF is a new protein belonging to the group of factors secreted by fibroblasts able to inhibit cellular proliferation.

Research paper thumbnail of Detection of antibodies against a human papillomavirus (HPV) type 16 peptide that differentiate high-risk from low-risk HPV-associated low-grade squamous intraepithelial lesions

Journal of General Virology, 2004

A nonapeptide (16L1) was derived from the human papillomavirus type 16 (HPV-16) major capsid prot... more A nonapeptide (16L1) was derived from the human papillomavirus type 16 (HPV-16) major capsid protein and tested for detection of potential cross-reactive serum IgG and cervical IgA antibodies in low-and high-risk HPV-associated low-grade squamous intraepithelial lesions (LSIL) and cervical cancer patients by ELISA. The IgG response was similar in women with low-risk HPV-associated LSIL and controls (P=0?1). In contrast, more than 90 % of patients with high-risk HPV-associated LSIL were seropositive. Although tumours from cancer patients were all positive for the presence of high-risk HPV DNA, the level of seropositivity decreased significantly in this group (P<0?0001). Cervical IgA antibodies were also detected in a significantly high proportion of women with high-risk HPV-associated LSIL compared with controls. However, the proportion of IgA-positive patients was lower than the proportion of IgG seropositives. In conclusion, the 16L1 peptide appears to be a high-risk type-common epitope that induces cross-reactive antibodies in high-risk, but not low-risk, HPV-associated LSIL patients, allowing differentiation of high-and low-risk infected women at this stage of infection.

Research paper thumbnail of Expression of MICA, MICB and NKG2D in human leukemic myelomonocytic and cervical cancer cells

Journal of Experimental & Clinical Cancer Research, 2011

Background: Cancer cells are known to secrete the stress molecules MICA and MICB that activate cy... more Background: Cancer cells are known to secrete the stress molecules MICA and MICB that activate cytotoxicity by lymphocytes and NK cells through their NKG2D receptor as a mechanism of immunological defense. This work was undertaken to evaluate if cancer cells can also express this receptor as a possible mechanisms of depletion of MIC molecules and thus interfere with their immune recognition.

Research paper thumbnail of Cationic liposomes bearing IL-2 on their external surface induced mice leukocytes to kill human cervical cancer cells in vitro , and significantly reduced tumor burden in immunodepressed mice

Journal of Drug Targeting, 2011

Research paper thumbnail of Peripheral blood lymphocytes from low-grade squamous intraepithelial lesions patients recognize vaccine antigens in the presence of activated dendritic cells, and produced high levels of CD8 + IFNγ + T cells and low levels of IL-2 when induced to proliferate

Infectious Agents and Cancer, 2012

Background: Most infections with human papillomavirus (HPV) are resolved without clinical interve... more Background: Most infections with human papillomavirus (HPV) are resolved without clinical intervention, but a minority evolves into chronic lesions of distinct grades, including cervical-uterine cancer. It is known that in most cases the immune system mediates elimination of HPV infection. However, the mechanism of immune evasion leading to HPV persistence and development of early cervical lesions is not fully understood. The aim of the present work was to evaluate the potential of peripheral blood leukocytes (PBL) from low-grade squamous intraepithelial lesions (LSIL) patients to be activated ex-vivo by vaccine antigens, the participation of cytotoxic lymphocytes and regulatory T cells, and to determine the secretion of Th1 and Th2 cytokines mediated by stimulation of T cell receptors. Results: We found that PBL from LSIL patients showed a significantly lower proliferation rate to vaccine antigens as compared to that of healthy donors, even though there was not a difference in the presence of antibodies to those antigens in sera from both groups. We did not find differences in either the frequency of CD4 + CD25 + FoxP3+ in PBL, or the levels of IL-4, IL-5 and IL-10 in plasma or conditioned media from PBL incubated with TcR agonists in vitro, between the two groups. However, we detected a lower production of IL-2 and a higher proportion of CD8 + IFNγ + cells in PBL from LSIL patients as compared with PBL from normal donors. We also observed that PBL from patients infected by HPV-16 and −18 were not able to proliferate in the presence of soluble HPV antigens added to the culture; however, a high level of proliferation was attained when these antigens were presented by activated dendritic cells.

Research paper thumbnail of Identification of peptides presented by HLA class I molecules on cervical cancer cells with HPV-18 infection

Immunology Letters, 1999

In this work we eluted peptides from purified class I MHC molecules, isolated from a novel human ... more In this work we eluted peptides from purified class I MHC molecules, isolated from a novel human cervical carcinoma cell line (INBL), generated in our laboratory and positive for HPV-18 infection. A fraction of these peptides was capable of stimulating T lymphocytes obtained from a donor matched for HLA-Cw4 and who was also HPV-18 + . Direct N-terminal Edman degradation of these peptides, revealed the sequence (XQFPIFLQF) that matched 85% with the sequence NVFPIFLQM localized in between the 54 and 62 residues of the HPV-18 L1 protein. After stimulation with the synthetic peptide NVFPIFLQM, T lymphocytes from the donor were capable to lyse INBL cells. Our results provide evidence of the existence of naturally occurring viral epitopes presented on cervical cancer cells by the HLA-Cw4 allele, that could be useful for immunotherapy on this type of patient.

Research paper thumbnail of Alpha-, beta- and kappa-caseins inhibit the proliferation of the myeloid cell lines 32D cl3 and WEHI-3 and exhibit different differentiation properties

Immunobiology, 2008

We have recently shown that sodium caseinate (CasNa) was able to inhibit the proliferation of the... more We have recently shown that sodium caseinate (CasNa) was able to inhibit the proliferation of the myeloid cell line 32D cl3 in a non-toxic way, and that it also induced the expression of macrophage colony-stimulating factor (M-CSF). Casein is the main protein present in milk and is composed of alpha (a), beta (b) and kappa (k) subunits. This work was undertaken to evaluate if any one casein is responsible for the proliferation and differentiation properties found for CasNa on myeloid cells. Taking into consideration that 32D cl3 cells are considered to be non-malignant and dependent on IL-3 for proliferation, we also included for this study a leukemic cell line, WEHI-3, that does not depend on any external growth factor for its proliferation in order to evaluate if the growth inhibitory effect of caseins is also present for malignant cells. Our results showed that all caseins were inhibitory for the proliferation of either 32D cl3 and WEHI-3 and that only the 32D cl3 cells were induced to differentiate into the monocyte-macrophage lineage. In order to evaluate if CasNa was able to inhibit the proliferation of other myeloid cells we used J774 and P388 and found that they were also inhibited. We also determined that the different caseins exhibit different differentiation properties, with a-casein being the only one able to induce the secretion of M-CSF. We consider this work to open a new field of research, where casein, or its components, can be studied for their possible role in hematopoiesis and on the inhibition of malignant cell proliferation for therapeutic use.

Research paper thumbnail of Sodium caseinate induces secretion of macrophage colony-stimulating factor from neutrophils

Immunobiology, 2010

In this work we provide evidence that granulocytes produce macrophage colony-stimulating factor (... more In this work we provide evidence that granulocytes produce macrophage colony-stimulating factor (M-CSF) in the band cell stage and secrete it upon sodium caseinate-mediated differentiation to polymorphonuclear cells. We identified M-CSF in an enriched population of myeloid band cells from murine bone marrow using a chromophorelabeled monoclonal anti-M-CSF antibody. An ELISA assay was then used to detect secreted M-CSF in culture supernatants of enriched band cells differentiated to mature neutrophils using sodium caseinate. Colony formation in vitro by the supernatants from differentiating band cells was blocked by anti-M-CSF, thus suggesting that this factor is the only one responsible for this activity. Our data imply that casein can modulate hematopoiesis possibly via M-CSF production. Finally we discuss the possibility whether this M-CSF in concert with G-CSF could establish a cellular communication network between macrophages and granulocytes allowing them to simultaneously arrive at the inflammatory site.

Research paper thumbnail of Evidence that cervical cancer cells secrete IL-2, which becomes an autocrine growth factor

Cytokine, 2010

We present evidence that cervical cancer cells express a functional IL-2 receptor (IL-2R). In fac... more We present evidence that cervical cancer cells express a functional IL-2 receptor (IL-2R). In fact, by RT-PCR we obtained that the IL-2R is present in CALO, and INBL cells, and that it consisted of the alphaIL-2R, betaIL-2R, and gammaIL-2R chains. We also found that IL-2 is a growth factor for these cell lines, and unexpectedly that CALO and INBL themselves being cancer cells produce, and secrete IL-2. Antibodies against the alpha and beta subunits of the IL-2R inhibited cell proliferation thus hinting to a cell growth dependency on this factor. Our results thus provide evidence that the IL-2R on cervical cancer cells is part of an autocrine mechanism for its growth to the extent that, like lymphocytes, they produce and become partially dependent on this growth factor. We think that in view of our results caution should be taken when IL-2 is being considered for cancer therapy; in particular when the patient&#39;s cancer cells present the IL-2R, because as indicated by our results, the use of this factor could promote tumor growth. Finally, the possible implications of the expression of both IL-2, and IL-2R on cervical cancer cells on the immune escape mechanism of tumor cells are discussed.

Research paper thumbnail of Interleukin-2 (IL-2) receptor-βγ signalling is activated by c-Kit in the absence of IL-2, or by exogenous IL-2 via JAK3/STAT5 in human papillomavirus-associated cervical cancer

Cellular Signalling, 2004

Activation of the interleukin-2 receptor (IL-2R) induces signalling cascades promoting T cell pro... more Activation of the interleukin-2 receptor (IL-2R) induces signalling cascades promoting T cell proliferation. However, signal transduction pathways triggered in IL-2R-expressing solid tumours are unknown. This report shows that human papillomavirus (HPV)-associated cervical cancer cells express an IL-2R composed of beta and gamma chains (IL-2Rbetagamma), and that IL-2-mediated activation increases the phosphorylation of JAK3 and STAT5, stimulating cell proliferation. Interestingly, endogenous IL-2 is not produced by these cells, suggesting the activation of IL-2Rbetagamma by an alternative mechanism. Accordingly, we found that Stem Cell Factor (SCF)-activated c-Kit induces phosphorylation of the IL-2Rbeta chain in the absence of IL-2. Moreover, inhibition of IL-2Rbeta phosphorylation by blocking c-Kit tyrosine kinase activity abolishes both, IL-2 and SCF-mediated proliferation. Thus, these results demonstrate that IL-2 triggers a JAK3/STAT5 cascade in HPV-associated cervical cancer cells expressing IL-2Rbetagamma, and that this receptor can be alternatively activated by SCF-activated c-Kit in the absence of IL-2.