Biomedical subjects

H Asou

Publications and source records attributed to H Asou.

At least 37 records · Page 2Linked to original sources

PubMed1999

Activation of EVI1 transcripts with chromosomal translocation joining the TCRVbeta locus and the EVI1 gene in human acute undifferentiated leukemia cell line (Kasumi-3) with a complex translocation of der(3)t(3;7;8).

A cell line (Kasumi-3) established from acute myeloid leukemia (AML-M0) had unique phenotypes of undifferentiated leukemia cells with expression of both T cell and myeloid antigens. Kasumi-3 cells with t(3;7)(q26;q22) highly expressed a 6 kb transcript of EVI1, which is located on chromosome 3q26. Therefore, we further characterized the chromosomal breakpoint by pulsed-field gel electrophoresis near EVI1. We identified and isolated the chromosomal breakpoint at approximately 80 kb upstream from the 5' end of EVI1. Sequence analysis of the breakpoint revealed that the whole Vbeta region from T cell receptor beta (TCRbeta) at 7q35 was translocated to the upstream of EVI1. A 1.0 kb TCRbeta transcript was expressed in the Kasumi-3 cells, suggesting that TCRbeta rearrangement occurred as Dbeta-Jbeta joining events. Fluorescence in situ hybridization analysis revealed that the inverted chromosome 7q22-q35 segment between TCRbeta and the region proximal to the erythropoietin gene at 7q22 was translocated to the region distal to EVI1 in der(3). Since the telomeric region of chromosome 8 q was also translocated to the inverted chromosome 7q22-q35 segment in der(3), the chromosomal abnormalities of der(3) were defined as being der(3)t(3;7;8)(3pter-3q26::7q35-7q22::8q22 -8qter). It is suggested that a translocated enhancer element in the TCRbeta locus and/or loss of a negative regulatory element near EVI1 might function to enhance the EVI1 expression. Therefore, the enhanced EVI1 expression may contribute to the development of a subset of undifferentiated leukemia.

Acute Disease↗

PubMed1999

Dynamics of astrocyte adhesion as analyzed by a combination of atomic force microscopy and immuno-cytochemistry: the involvement of actin filaments and connexin 43 in the early stage of adhesion.

We observed the time-dependent morphological alteration of astrocytes during their adhesion by atomic force microscopy (AFM) and investigated the relationships between this morphological alteration and the localization of actin filaments and connexin 43 by immunocytochemistry. The fine processes observed as fine ridge-like structures by AFM were closely concerned with actin filaments by immunocytochemistry. During the adhesion of astrocytes, actin filaments appeared to be aligned regularly beyond the borders among different cells. Detectable connexin immunoreactivity was changed in the following regions: 1) the tips of fine cell processes and the cell margin when astrocytes started to adhere; 2) the border of cells when astrocytes tightly adhered; and 3) non-specific sites when astrocytes became a cluster. In the former two cases, the immunopositive spots for connexin were observed to colocalize with the tips of cell processes with actin filaments. These results strongly suggest that connexin associated with actin filaments at the tip of cell processes plays an important role in the early stage of the adhesion of astrocytes. These observations afford valuable clues for understanding the glial communication.

Actin Cytoskeleton↗

PubMed1999

Growth inhibition of myeloid leukemia cells by troglitazone, a ligand for peroxisome proliferator activated receptor gamma, and retinoids.

Peroxisome proliferator activated receptor gamma (PPARgamma) plays a central role in the process of adipocyte differentiation. This receptor and its heterodimeric partner, retinoid X receptor alpha (RXRalpha), form a DNA-binding complex that regulates transcription of adipocyte-specific genes. Troglitazone, an antidiabetic drug, has recently been identified as a synthetic ligand for PPARgamma. We studied the effects of troglitazone on proliferation and differentiation of normal and malignant hematopoietic cells. Expression of PPARgamma was easily detectable by Western blot analyses in all five myeloid leukemia cell lines. Troglitazone alone (10-5 M) did not induce differentiation in any of the cell lines; however, this compound suppressed the clonal growth (10-75% of inhibition) of all five myeloid leukemia cell lines. Myelomonocytic U937 cells, which were the most responsive to the growth suppressing effects of troglitazone, were arrested in the G1 phase of the cell cycle when cultured with this compound. Simultaneous treatment of myeloid leukemia cell lines with both troglitazone and a ligand that specifically binds either RXR (LG100268), or retinoic acid receptors (RAR, ATRA, ALART1550), or both (9-cis RA) resulted in additive suppression of clonal growth. In summary, our studies showed that troglitazone when combined with a retinoid was a moderately potent inhibitor of clonogenic growth of acute myeloid leukemia cells.

Alitretinoin↗

PubMed1999

Herpesvirus 8 inclusions in primary effusion lymphoma: report of a unique case with T-cell phenotype.

We describe a case of primary effusion lymphoma with T-cell phenotype, mixed genotype, and intranuclear herpesvirus inclusions visible with the light microscope. Cells were studied by immunohistochemical analysis, in situ hybridization, immunoglobulin and T-cell receptor gene rearrangement, and polymerase chain reaction. Primary effusion lymphoma cells with T-cell phenotype revealed herpesvirus 8 inclusions predominantly seen in apoptotic cells, suggesting that productive viral infection is associated with cell death. Clinical features were typical of primary effusion lymphoma. Cytologic, molecular genetic, and phenotypic features demonstrated a unique variant of primary effusion lymphoma.

Adult↗

PubMed1998

Monoclonal antibody 14F7, which recognizes a stage-specific immature oligodendrocyte surface molecule, inhibits oligodendrocyte differentiation mediated in co-culture with astrocytes.

Cells at an intermediate stage of oligodendrocyte lineage are not only well characterized by biochemical studies but also are likely to relate to the outcome of physiological events. To elucidate the molecular events leading to the development of oligodendrocyte lineage cells, we have raised monoclonal antibodies against stage-specific immature oligodendrocytes, which have previously been isolated by a novel oligodendrocyte-lineage cell culture technique (Sakurai et al.: J Neurosci Res 52:17-26, 1998). We have isolated a mouse monoclonal antibody termed 14F7 which predominantly labels stage-specific immature oligodendrocytes and have found that the expression of 14F7 immunoreactivity in the developing neonatal rat forebrain is closely associated with cells expressing the oligodendrocyte progenitor marker A2B5 and to immature oligodendrocyte expressing O4 antigen. 14F7+ cells were distributed in the ventricular and subventricular zone and the nearby forming corpus callosum as non-myelinating cells. In contrast to cell culture observations, 14F7+ cells were seen only in oligodendrocyte lineage cells. For instance, dissociated cell culture studies indicated that 14F7 labels a cell surface molecule, and its cellular distribution is coincident with all of O4+ cells and A2B5+ cells, and even A2B5- cells. By contrast, 14F7-positive cells did not label astrocytes and, furthermore, did not label myelin basic protein (MBP)-positive oligodendrocytes. 14F7 recognized a 48-kDa protein on sodium dodecyl sulfate polyacrylamide gel electrophoresis. 14F7 immunoreactivity was detectable in rat brain as early as embryonic day 18. Furthermore, in these cells, the total time for differentiation was extended, and on maturation, these cells subsequently expressed an array of myelin-specific proteins, which normally occurs by direct contact with type-1 astrocytes. However, in the presence of 14F7, stage-specific oligodendrocytes co-cultured with astrocytes completely failed to express MBP. These data suggest that the 14F7 antigen is a novel cell surface molecule that is expressed in the intermediate stage of oligodendrocyte-lineage cells, and it is expected that it regulates the differentiation of oligodendrocyte throughout development.

Animals↗

PubMed1998

19-nor vitamin-D analogs: a new class of potent inhibitors of proliferation and inducers of differentiation of human myeloid leukemia cell lines.

We have studied the in vitro biological activities and mechanisms of action of 1,25-dihydroxyvitamin D3 (1,25D3) and nine potent 1,25D3 analogs on proliferation and differentiation of myeloid leukemia cell lines (HL-60, retinoic acid-resistant HL-60 [RA-res HL-60], NB4 and Kasumi-1). The common novel structural motiff for almost all the analogs included removal of C-19 (19-nor); each also had unsaturation of the side chain. All the compounds were potent; for example, the concentration of analogs producing a 50% clonal inhibition (ED50) ranged between 1 x 10(-9) to 4 x 10(-11) mol/L when using the HL-60 cell line. The most active compound [1, 25(OH)2-16,23E-diene-26-trifluoro-19-nor-cholecalciferol (Ro 25-9716)] had an ED50 of 4 x 10(-11) mol/L; in contrast, the 1,25D3 produced an ED50 of 10(-9) mol/L with the HL-60 target cells. Ro 25-9716 (10(-9) mol/L, 3 days) was a strong inducer of myeloid differentiation because it caused 92% of the HL-60 cells to express CD11b and 75% of these cells to reduce nitroblue tetrazolium (NBT). This compound (10(-8) mol/L, 4 days) also caused HL-60 cells to arrest in the G1 phase of the cell cycle (88% cells in G1 v 48% of the untreated control cells). The p27(kip-1), a cyclin-dependent kinase inhibitor which is important in blocking the cell cycle, was induced more quickly and potently by Ro 25-9716 (10(-7) mol/L, 0 to 5 days) than by 1,25D3, suggesting a possible mechanism by which these analogs inhibit proliferation of leukemic growth. The NB4 promyelocytic leukemia cells cultured with the Ro 25-9716 were also inhibited in their clonal proliferation (ED50, 5 x 10(-11) mol/L) and their expression of CD11b was enhanced (80% positive [10(-9) mol/L, 4 days] v 27% untreated NB4 cells). Moreover, the combination of Ro 25-9716 (10(-9) mol/L) and all-trans retinoic acid (ATRA, 10(-7) mol/L) induced 92% of the NB4 cells to reduce NBT, whereas only 26% of the cells became NBT positive after a similar exposure to the combination of 1,25D3 and ATRA. Surprisingly, Ro 25-9716 also inhibited the clonal growth of poorly differentiated leukemia cell lines (RA-res HL-60 [ED50, 4 x 10(-9) mol/L] and Kasumi-1 [ED50, 5 x 10(-10) mol/L]). For HL-60 cells, Ro 25-9716 markedly decreased the percent of the cells in S phase of the cell cycle and increased the expression of the cyclin-dependent kinase inhibitor, p27(kip-1). In summary, 19-nor vitamin D3 compounds strongly induced differentiation and inhibited clonal proliferation of various myeloid leukemia cell lines, suggesting a therapeutic niche for their use in myeloid leukemia.

Cell Cycle↗

PubMed1998

Ligand for peroxisome proliferator-activated receptor gamma (troglitazone) has potent antitumor effect against human prostate cancer both in vitro and in vivo.

Troglitazone, a thiazolidinedione derivative, is a widely used antidiabetic drug that binds and activates peroxisome proliferator-activated receptor gamma (PPARgamma) and enhances insulin sensitivity. It induces differentiation of adipocytes, which highly express PPARgamma. We report that human prostate cancer cells expressed PPARgamma at prominent levels and normal prostate tissues had very low expression. Dose-response clonogenic assays of the PC-3 prostate cancer cell line with troglitazone showed an antiproliferative effect (ED50, 3 x 10(-7) M) and other PPARgamma ligands (BRL49653: ED50, 8 x 10(-8) M; 15-deoxy-delta12,14-prostaglandin J2: ED50, 2 x 10(-6) M; ciglitizone: ED50, not reached; indomethacin: ED50, not reached) showed similar effects. Combinations of troglitazone and a ligand specific for either retinoid X receptor or retinoic acid receptor did not show a synergistic effect. Pulse-exposure to troglitazone (10(-5) M) for different durations showed that 4 days of pulse-exposure to the agent irreversibly inhibited 50% clonal growth of PC-3 cells. Interestingly, PC-3 cells cultured with troglitazone (10(-5) M) showed dramatic morphological changes both by light and electron microscopy, suggesting that the cells became less malignant. Nevertheless, troglitazone did not affect either the cell cycle or several markers of differentiation. LNCaP cells constitutively produced prostate-specific antigen, and levels were markedly enhanced by all-trans-retinoic acid. Troglitazone (10(-5) M, 4 days) decreased by 50% the levels of prostate-specific antigen produced by these cells. In vivo treatment of PC-3 tumors growing in male BNX triple immunodeficient mice with oral troglitazone (500 mg/kg/day) produced significant inhibition of tumor growth (P = 0.01). The only objective side effect of troglitazone in mice was the elevation of serum transaminases. Short-term culture of four surgically obtained human prostate cancer tumors with troglitazone (10(-5) M, 4 days) produced marked and selective necrosis of the cancer cells (about 60%) but not the adjacent normal prostate cells. Taken together, these results suggest that troglitazone may be a useful therapeutic agent for the treatment of prostate cancer, especially in the setting of low disease burden.

Animals↗

PubMed1998

Ligands for peroxisome proliferator-activated receptorgamma and retinoic acid receptor inhibit growth and induce apoptosis of human breast cancer cells in vitro and in BNX mice.

Induction of differentiation and apoptosis in cancer cells through ligands of nuclear hormone receptors (NHRs) is a novel and promising approach to cancer therapy. All-trans-retinoic acid (ATRA), an RA receptor-specific NHR ligand, is now used for selective cancers. The NHR, peroxisome proliferator-activated receptor gamma (PPARgamma) is expressed in breast cancer cells. Activation of PPARgamma through a synthetic ligand, troglitazone (TGZ), and other PPARgamma-activators cause inhibition of proliferation and lipid accumulation in cultured breast cancer cells. TGZ (10(-5) M, 4 days) reversibly inhibits clonal growth of MCF7 breast cancer cells and the combination of TGZ (10(-5) M) and ATRA (10(-6) M, 4 days) synergistically and irreversibly inhibits growth and induces apoptosis of MCF7 cells, associated with a dramatic decrease of their bcl-2 protein levels. Similar effects are noted with in vitro cultured breast cancer tissues from patients, but not with normal breast epithelial cells. The observed apoptosis mediated by TGZ and ATRA may be related to the striking down-regulation of bcl-2, because forced over-expression of bcl-2 in MCF7 cells cultured with TGZ and ATRA blocks their cell death. TGZ significantly inhibits MCF7 tumor growth in triple immunodeficient mice. Combined administration of TGZ and ATRA causes prominent apoptosis and fibrosis of these tumors without toxic effects on the mice. Taken together, this combination may provide a novel, nontoxic and selective therapy for human breast cancers.

Adenocarcinoma↗

PubMed1998

Switching of platelet-activating factor acetylhydrolase catalytic subunits in developing rat brain.

In a previous study, we demonstrated that Platelet-activating Factor (PAF) acetylhydrolase purified from bovine brain cortical cytosol consists of two mutually homologous catalytic subunits (alpha1 and alpha2) and one putative regulatory beta subunit. The latter is a product of the LIS1 gene, which is defective in the Miller-Dieker syndrome, a form of lissencephaly. In this study, we examined the expression patterns of these three subunits in the developing rat brain. All three subunits were expressed in embryonic brain, whereas only alpha2 and beta subunit were detected in the adult brain by Western blotting. Biochemical analyses revealed that the alpha1/alpha2 heterodimer and alpha2/alpha2 homodimer are major catalytic units of embryonic and adult brain PAF acetylhydrolases, respectively. The alpha1 transcript and protein were detected predominantly in embryonic and postnatal neural tissues, such as the brain and spinal cord. Furthermore, we found using primary cultured cells isolated from neonatal rat brain that alpha1 protein were expressed only in neurons but not in glial cells and fibroblasts. In contrast, alpha2 and beta transcripts and proteins were detected both in neural and non-neural tissues, and their expression level was almost constant from fetal stages through adulthood. These results indicate that alpha1 expression is restricted to actively migrating neurons in rats and that switching of catalytic subunits from the alpha1/alpha2 heterodimer to the alpha2/alpha2 homodimer occurred in these cells during brain development, suggesting that PAF acetylhydrolase plays a role(s) in neuronal migration.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗

PubMed1998

Expression of the neural cell adhesion molecule in astrocytic tumors: an inverse correlation with malignancy.

BACKGROUND: Cell adhesion molecules are among the key factors in the development of the malignant potential of brain tumors. The aim of this study was to investigate the expression of the neural cell adhesion molecule (NCAM) in human astrocytic tumors and assess any relationship between NCAM expression and the degree of malignancy. METHODS: The expression of NCAM was examined in 52 astrocytic tumors by Western blot analysis. From them the authors selected 23 adult supratentorial ordinary astrocytic tumors and performed quantitative Western blot analysis for each isoform (NCAM 172-180, NCAM 145, NCAM 125-130) to investigate any correlation between the expression of each NCAM isoform and the histologic and biologic malignancy (histology, proliferating cell indices [PCIs] determined by MIB-1 immunohistochemistry, and manifestation on magnetic resonance images [MRIs]). Immunohistochemistry with antihuman NCAM monoclonal antibody was also performed on the tumors from which cryostat sections were available. RESULTS: Most of the astrocytomas and anaplastic astrocytomas revealed 3 bands at 180, 145, and 125-130kD, whereas in glioblastomas the bands tended to diminish. The expression of each NCAM isoform in astrocytic tumors decreased in proportion to the progression of the histologic malignancy, and the results were also corroborated by immunohistochemical evaluation. An inverse correlation was also observed between the amount of NCAM expression and MIB-1 PCIs. NCAM expression was hardly detectable in those tumors with highly invasive manifestation on MRIs. CONCLUSIONS: To the authors' knowledge, this study provides the first direct evidence that NCAM is down-regulated in the development of the malignancy of astrocytic tumors; and it is suggested that reduced NCAM expression might be involved in the development of biologic malignancy.

Adult↗

PubMed1998

Coordinate expression of L1 and 6B4 proteoglycan/phosphacan is correlated with the migration of mesencephalic dopaminergic neurons in mice.

Mesencephalic dopaminergic (DA) neurons of mice are generated from embryonic day 10 to 12 (E10-12) in the ventricular zone of the mesencephalon. They first migrate toward the ventral mesencephalon, and then turn laterally, or tangentially, in the basal part of the mesencephalon. With immunohistochemical analysis of E10-E15 ICR mice, we found that cell adhesion molecule L1 was transiently expressed on the median part of tangential fibers coincident with the lateral migration of DA neurons from E11 to E13, when neurons move along the tangential fibers toward their final destinations: the reticular formation, the substantia nigra pars compact, and the ventral tegmental area. While L1 expression was not observed in DA neurons, they expressed a chondroitin sulfate proteoglycan, 6B4 proteoglycan/phosphacan, which has been shown to bind to L1/Ng-CAM in vitro. These results suggest that the heterophilic interaction between 6B4 proteoglycan on the neurons and L1 on the fibers is involved in the lateral migration of mesencephalic DA neurons in mice.

Animals↗

PubMed1998

Differentiation of oligodendrocyte occurs in contact with astrocyte.

We have generated stage-specific oligodendrocytes, which are identified as immature oligodendrocytes by the expression of specific markers. These cells were generated by using 3 types of intrinsic and extrinsic factors for influencing the characteristics of oligodendrocyte-lineage cells. The factors included 10% serum in the medium to induce cell proliferation, bFGF in serum-free medium to increase survival and induce differentiation, and 0.25% trypsin to remove nonoligodendrocyte lineage cells. We have also demonstrated that it is possible to supply a huge amount of homogeneous populations of stage-specific immature oligodendrocytes after 35 days' culture. The maturation of these cells is controlled by interacting with astrocytes in vitro, resulting in the expression of MBP isoforms (14.0 and 18.5 kDa) that may participate in mature oligodendrocyte membrane compaction.

Animals↗

PubMed1998

Mechanisms of growth control of Kaposi's sarcoma-associated herpes virus-associated primary effusion lymphoma cells.

Primary effusion lymphoma (PEL) is a distinct clinicopathologic entity associated with Kaposi's sarcoma-associated herpes virus (KSHV). Several cytokines, including interleukin-6 (IL-6), basic fibroblast growth factor (bFGF), and platelet-derived growth factor (PDGF) may be important for survival of KS cells. However, little is known about the interaction of cytokines with KSHV-infected lymphocytes from PEL. Therefore, we investigated what cytokines were produced by KSHV-infected PEL cell lines (KS-1, BC-1, BC-2), what cytokine receptors were expressed by these cells, what response these cells had to selected cytokines, and what was the effect of IL-6 antisense phosphorothioated oligonucleotides. Reverse transcriptase-polymerase chain reaction (RT-PCR) and protein studies showed that these three cell lines produced IL-10, IL-6, and the receptors for IL-6. The granulocyte macrophage colony-stimulating factor (GM-CSF), IL-1beta, IL-8, IL-12, bFGF, PDGF, and c-kit transcripts were not detected in the cell lines. High levels (0.7 to 5 ng/mL/10(6) cells/48 hours) of IL-6 protein were consistently detected in supernatants of the cell lines by enzyme-linked immunosorbent assay (ELISA) tests. In clonogenic assays, interferon-alpha (IFN-alpha) and IFN-gamma suppressed the clonal growth of the PEL cells, but GM-CSF, IL-4, IL-6, IL-8, IL-10, and oncostatin M did not change it. We examined for several autocrine loops that have been suggested to occur in KS. Experiments using antisense oligonucleotides showed that the clonal growth of KS-1 and BC-1 was nearly 100% inhibited by IL-6 antisense oligonucleotides (10 micromol/L), but not at all by either oligonucleotides (</=10 micromol/L) to IL-6 sense, IL-6 scrambled, viral IL-6 (vIL-6) antisense, or IL-10 antisense. Furthermore, the IL-6 antisense oligonucleotides had no effect on two B-cell lymphoma cell lines, which were not infected with KSHV. Addition of IL-6 antibody did not inhibit clonal growth of any of the cell lines. Taken together, we have defined the cytokines and their receptors expressed on PEL cells and have found that these cells synthesized IL-6 and IL-6 receptors; interruption of this pathway by IL-6 antisense oligonucleotides specifically prevented the growth of these cells. These findings will offer potential new therapeutic strategies for PEL.

Cell Division↗

PubMed1998

Hyaluronectin is produced by oligodendrocytes and Schwann cells in vitro.

A hyaluronectin (HN)-like antigen was found in rat O-2A progenitors and oligodendrocytes, as well as in Schwann cells and in their culture medium. The HN-like antigen secreted in culture supernatants had a higher molecular mass than HN extracted from rat brain at acidic pH. In vitro the secreted HN-like antigen was spontaneously and slowly degraded into species whose Mr was close to that of HN found in acidic brain extract. In brain or nerve neutral pH extracts, both HN-like antigen and HN were present. The high Mr of the secreted antigen, the homology in amino acid sequences between HN and N-terminal domain of PG-M/versican, in addition to a positive hybridization between Schwann cell RNAs and a probe obtained with primers derived from HN sequences also found in versican suggested that HN is closely related to the large proteoglycan PG-M/versican. The presence in Schwann cell extract of a HN mRNA whose Mr was compatible with the size expected for HN showed that HN may be directly secreted by cells and not only the consequence of a proteolytic cleavage. The similarity of HN with PG-M (V3) suggested that HN found in vivo could be the result of an alternative splicing of a single gene. We conclude that HN as other members of the PG-M/versican family is a marker of oligodendrocytes and Schwann cells in culture.

Animals↗

PubMed1998

Methylation of the p16INK4A gene in multiple myeloma.

The p16INK4A (p16) binds to both cyclin D-CDK4 and cyclin D-CDK6 and inhibits the progression of the cell cycle from G1 to S phase. Loss of expression of this protein can occur by several mechanisms including structural alterations. Recent studies have suggested that the loss of expression of p16 can occur by hypermethylation of the gene. The methylation status of the p16 gene in multiple myeloma was examined in three myeloma cell lines (U266, RPMI8226 and IM9) and 16 primary myeloma samples using methylation-specific polymerase chain reaction (MSP). The U266 and RPMI8226 cell lines contained a completely methylated p16 gene and the IM9 line had a partially methylated p16 gene. Identical results were obtained by another polymerase chain reaction (PCR)-based methylation assay system as well as Southern blotting after using a methylation-sensitive restriction enzyme. The U266 cell line expressed no p16, and the IM9 had weak expression as determined by reverse transcript (RT-)PCR. The U266 cells began to express, and IM9 increased the accumulation of, the p16 RNA after treatment with the demethylating agent 5'-aza-2-deoxycytidine (10(-6)-10(-5) M). This suggested that the levels of methylation of the p16 gene detected by the MSP technique correlated with the regulation of transcription of this gene. Examination of the primary myeloma samples showed that eight of 16 (50%) contained a methylated p16 gene. We have previously found that alterations of the p16 gene, such as deletions and point mutations, are rare in primary multiple myeloma; none of the 16 samples included in this study had p16 gene alterations. Our results suggest that methylation of the p16 gene may contribute to the development and/or progression of multiple myeloma.

Aged↗

PubMed1997

Localization of Kaposi's sarcoma-associated herpesvirus in bone marrow biopsy samples from patients with multiple myeloma.

We have recently demonstrated the presence of Kaposi's sarcoma-associated herpesvirus (KSHV) in cultured bone marrow (BM) stromal dendritic cells from all patients with myeloma studied. To show that these findings were not an artifact of tissue culture, we performed in situ hybridization (ISH) and polymerase chain reaction (PCR) to detect KSHV in BM core biopsies. Using ISH to open reading frame-72 (ORF 72), we localized KSHV to BM dendritic cells in 17 of 20 patients with myeloma, 2 patients with plasmacytosis associated with the acquired immunodeficiency syndrome, and 1 case of aplastic anemia. In contrast, BM from normal subjects (n = 4) and patients with lymphoma and leukemia (n = 21) did not contain KSHV. PCR amplification with KSHV primers demonstrated product in fresh BM biopsy samples from 6 of 7 myeloma patients, whereas three normal marrows contained no amplified product. These findings suggest that KSHV, possibly through alterations in the BM microenvironment and production of viral interleukin-6 (vIL-6), may stimulate and maintain abnormal plasma cell proliferation in myeloma and related disorders.

AIDS-Related Opportunistic Infections↗

PubMed1997

PAF analogues capable of inhibiting PAF acetylhydrolase activity suppress migration of isolated rat cerebellar granule cells.

Intracellular platelet-activating factor (PAF) acetylhydrolase in the bovine brain is a heterotrimeric enzyme composed of alpha1, alpha2 and beta subunits. The trimeric enzyme may be involved in neural cell migration, since the human homolog of the non-catalytic beta subunit is a product of the LIS-1 gene which is a causative gene for Miller-Dieker syndrome. Miller-Dieker syndrome is a form of lissencephaly that is characterized by widespread agyria of the brain and defects of neuronal cell migration. In the present study, we attempted to determine whether the catalytic activity of either the alpha1 or alpha2 subunit is required for the regulation of granule cell migration. Granule cells prepared from rat cerebellum at postnatal day 0 express all three subunit proteins (alpha1, alpha2 and beta) as determined by western blotting. Granule cell migration, which was observed in vitro on a layer coated with laminin, was effectively blocked by PAF analogs which showed PAF receptor-antagonistic activity (CV-6209 and CV-3988) and PAF receptor-agonistic activity (carbamoyl PAF). These PAF analogs also inhibited the activity of bovine brain PAF acetylhydrolase. Cell migration was restored when the inhibitors were removed by washing the treated cells with buffer, indicating that the inhibitory effect of PAF analogs is reversible. Structurally-unrelated PAF antagonists (SM-12502, TCV-309 and YM-264), none of which showed any appreciable inhibitory activity against PAF acetylhydrolase, did not block granule cell migration under the same conditions. It is suggested that the catalytic activity of PAF acetylhydrolase may play a crucial role in neural cell migration.

1-Alkyl-2-acetylglycerophosphocholine Esterase↗

PubMed1997

19-nor-hexafluoride analogue of vitamin D3: a novel class of potent inhibitors of proliferation of human breast cell lines.

Breast cancer cells express vitamin D3 receptors and 1,25-dihydroxyvitamin D3 suppressed growth of these cells. We have synthesized six novel vitamin D3 analogues to identify those with expanded capacity to inhibit the proliferative ability of breast cancer cells. These analogues incorporated many of the structural motifs shown previously to have antiproliferative activity in several cell types. Six breast cancer cell lines were used as targets. Dose-response studies showed that each of the analogues had antiproliferative activities, and LH [1,25-(OH)2-16-ene-23-yne-26,27-F6-19-nor D3] was the most potent analogue, suppressing at 10(-11) M greater than 50% clonal proliferation (ED50) of the MCF-7 and SK-BR-3 breast cancer cells, increasing the proportion of MCF-7 cells in the G0-G1 phase, and decreasing those in the S phase of the cell cycle. Pulse-exposure studies showed that a 3-day exposure to LH (10(-7) M) in liquid culture was adequate to achieve a 50% inhibition of MCF-7 clonal growth in soft agar in the absence of the analogue, suggesting that the growth inhibition mediated by LH is irreversible. The cyclin-dependent kinase inhibitor known as p27Kip1 helps regulate the cell cycle and can mediate growth arrest in response to extracellular growth inhibitors. The analogue LH (10(-7) M) induced elevated expression of p27Kip1 in MCF-7 and SK-BR-3 cells. Taken together, these results indicate that LH is an extremely potent vitamin D3 analogue markedly inhibiting clonal growth of MCF-7 and SK-BR-3 cells with concomitant cell cycle arrest at G0-G1 and increased expression of p27Kip1. Compound LH is worthy of in vivo analysis for possible future clinical trials.

Antineoplastic Agents↗