Tripartite containing motif 32 modulates proliferation of human neural precursor cells in HIV-1 neurodegeneration.

In addition to glial cells, HIV-1 infection occurs in multipotent human neural precursor cells (hNPCs) and induces quiescence in NPCs. HIV-1 infection of the brain alters hNPC stemness, leading to perturbed endogenous neurorestoration of the CNS following brain damage by HIV-1, compounding the severity of dementia in adult neuroAIDS cases. In pediatric neuroAIDS cases, HIV-1 infection of neural stem cell can lead to delayed developmental milestones and impaired cognition. Using primary cultures of human fetal brain-derived hNPCs, we gained novel insights into the role of a neural stem cell determinant, tripartite containing motif 32 (TRIM32), in HIV-1 Tat-induced quiescence of NPCs. Acute HIV-1 Tat treatment of hNPCs resulted in proliferation arrest but did not induce differentiation. Cellular localization and levels of TRIM32 are critical regulators of stemness of NPCs. HIV-1 Tat exposure increased nuclear localization and levels of TRIM32 in hNPCs. The in vitro findings were validated by studying TRIM32 localization and levels in frontal cortex of HIV-1-seropositive adult patients collected at post mortem as well as by infection of hNPCs by HIV-1. We observed increased percentage of cells with nuclear localization of TRIM32 in the subventricular zone (SVZ) as compared with age-matched controls. Our quest for probing into the mechanisms revealed that TRIM32 is targeted by miR-155 as downregulation of miR-155 by HIV-1 Tat resulted in upregulation of TRIM32 levels. Furthermore, miR-155 or siRNA against TRIM32 rescued HIV-1 Tat-induced quiescence in NPCs. Our findings suggest a novel molecular cascade involving miR-155 and TRIM32 leading to HIV-1 Tat-induced attenuated proliferation of hNPCs. The study also uncovered an unidentified role for miR-155 in modulating human neural stem cell proliferation, helping in better understanding of hNPCs and diseased brain.

The Notch co-repressor protein NKAP is highly expressed in adult mouse subventricular zone neural progenitor cells.

In the adult mammalian brain niches for neural stem cells are maintained, which enable a steady-state neurogenesis. This process is tightly regulated by multiple niche factors, including Notch and NF-κB signaling. The NF-κB-activating-protein (NKAP) has previously been shown to act as Notch co-repressor component by binding CIR and recruiting HDAC3 in T-cell development and furthermore to regulate NF-κB-dependent transcription. Here, we provide first evidence for the expression of NKAP in neurogenic cells of the adult mammalian brain. NKAP is highly expressed in Mash1(+) transit amplifying cells and PSA-NCAM(+) migrating neuroblasts throughout the subventricular zone (SVZ) and the rostral migratory stream (RMS), as well as in the hippocampus. We further show that NKAP expression levels are downregulated during the course of the RMS. Eventually, most differentiated cells in the olfactory bulb (OB) and the corpus callosum only display low levels of NKAP expression. Finally, large subsets of mature neurons in the OB, the hippocampus and the thalamus express NKAP at high levels, suggesting an additional role of NKAP outside of SVZ progenitor cells.

TRIM32-dependent transcription in adult neural progenitor cells regulates neuronal differentiation.

In the adult mammalian brain, neural stem cells in the subventricular zone continuously generate new neurons for the olfactory bulb. Cell fate commitment in these adult neural stem cells is regulated by cell fate-determining proteins. Here, we show that the cell fate-determinant TRIM32 is upregulated during differentiation of adult neural stem cells into olfactory bulb neurons. We further demonstrate that TRIM32 is necessary for the correct induction of neuronal differentiation in these cells. In the absence of TRIM32, neuroblasts differentiate slower and show gene expression profiles that are characteristic of immature cells. Interestingly, TRIM32 deficiency induces more neural progenitor cell proliferation and less cell death. Both effects accumulate in an overproduction of adult-generated olfactory bulb neurons of TRIM32 knockout mice. These results highlight the function of the cell fate-determinant TRIM32 for a balanced activity of the adult neurogenesis process.

Regulatory feedback loop between TP73 and TRIM32.

The p73 transcription factor is one of the members of the p53 family of tumor suppressors with unique biological functions in processes like neurogenesis, embryonic development and differentiation. For this reason, p73 activity is tightly regulated by multiple mechanisms, including transcription and post-translational modifications. Here, we identified a novel regulatory loop between TAp73 and the E3 ubiquitin ligase tripartite motif protein 32 (TRIM32). TRIM32, a new direct p73 transcriptional target in the context of neural progenitor cells, is differentially regulated by p73. Although TAp73 binds to the TRIM32 promoter and activates its expression, TAp73-induced TRIM32 expression is efficiently repressed by DNp73. TRIM32 in turn physically interacts with TAp73 and promotes its ubiquitination and degradation, impairing p73-dependent transcriptional activity. This mutual regulation between p73 and TRIM32 constitutes a novel feedback loop, which might have important implications in central nervous system development as well as relevance in oncogenesis, and thus emerges as a possible therapeutic target.

Cytosine arabinoside, idarubicin and divided dose etoposide for the treatment of acute myeloid leukemia in elderly patients.

Elderly patients with acute myeloid leukemia (AML) have an unfavourable prognoses due to low remission rates, short remission durations, and a high treatment related toxicity. Therefore, new chemotherapy regimens with curative potential, decreased toxicity, and applicability to the majority of these patients are still needed. For remission induction, AML patients > or = 61 years of age received one to three induction courses of the DIVA regimen (idarubicin 10 mg/m2/d days 1 and 3, etoposide 2 x 60 mg/m2/d every 12 hrs. days 2 to 5, and cytarabine 100 mg/m2/d as continuous i.v. infusion days 1 to 5). After achieving CR, patients received two additional courses of DIVA as consolidation therapy. Forty-two consecutive patients with de novo and secondary AML with a median age of 68 years were entered into this trial while six patients were judged ineligible for medical reasons. 62% of the patients achieved a CR, lasting for a median of 26 weeks. Toxicity was mainly hematologic with an early death rate of 12%. The median overall survival for all patients was 38 weeks, and 51 weeks for the 26 patients who achieved CR. Outcome was not significantly different for patients with de novo compared to secondary AML. In conclusion, DIVA showed a promising antileukemic activity and acceptable toxicity as remission induction therapy for de novo and secondary AML in this negligible selected group of elderly patients. However, relapse rate was high, indicating the need for novel approaches for consolidation and maintenance therapy.

Antineuronal antibody-associated paraneoplastic neuropathy in Hodgkin's disease.

Paraneoplastic neurological syndromes in patients with Hodgkin's disease are rare findings. Subacute, paraneoplastic cerebellar degeneration or autonomic dysfunctions were described before. In some of these cases, autoantibodies against central or peripheral nervous system structures were found in serum and CSF. We present a 30-year-old white male who developed a progredient, clinical and electrophysiological distal sensomotoric neuropathy. Six months after the beginning of the neurological disturbances, Hodgkin's disease (Stadium III BE) was diagnosed. Other reasons for neuropathy, such as direct impairment of the peripheral nervous system by tumor masses or drug-induced neuropathy, were excluded. Cerebrospinal fluid (CSF) analysis showed a mild pleocytosis, elevated total protein (9.8 g/l) and identical oligoclonal bands in serum and CSF. Blood-CSF barrier damage was detected by Reiber formula. Indirect immunofluorescence and western blot analysis demonstrated an autoantibody against peripheral and central nervous system structures in serum and CSF. Although the autoantibody responded to a 38-40 kDa-protein in western blot and showed nuclear staining of myenteric plexus and Purkinje cell nuclei in the immunofluorescence test, this antibody was shown to be not identical to anti-Hu. An intrathecal synthesis of the antineuronal antibody was detected by antibody specificity index. Tumor therapy, plasmapheresis and treatment with intravenous immunoglobulins did not improve the neuropathy. According to our knowledge this is the first case of antineuronal antibody-associated sensomotoric neuropathy in Hodgkin's disease.

Influence of neutrophil separation on the expression of adhesion molecules.

In many assays of polymorphonuclear neutrophil (PMN) function the first step is separation of PMN from whole blood. In the present investigation it was examined if PMN separation leads to an altered expression of neutrophil surface membrane adhesion molecules. Samples have been taken from 20 healthy volunteers (10 male, 10 female; 39.7 +/- 11.8 years of age). PMN activation was measured cytometrically using the following antibodies against PMN surface membrane receptors: L-selectin (CD 62 L), beta-2-integrin Mac-1 (CD 11b) and Intercellular Adhesion Molecule 1 (CD54). PMN activation was determined in whole blood and after separation of PMN using density gradients. After PMN separation all three adhesion molecules appeared increased but the effect was only statistically significant for CD 54 (Wilcoxon test). Data (mean fluorescence intensity in arbitrary units) were: CD 62 L: 62 +/- 37 in whole blood, 82 +/- 28 after separation; p = 0.0674, CD 11b: 94 +/- 55 in whole blood, 111 +/- 47 after separation; p = 0.1454, CD 54; 13 +/- 12 in whole blood, 81 +/- 35 after separation; p < 0.0001. With the present data available it can be assumed that separation of PMN from whole blood can influence the results of flow cytometric assays.

The BFM-protocol for HIV-negative Burkitt's lymphomas and L3 ALL in adult patients: a high chance for cure.

During a period of 9 years we used the pediatric BFM-NHL protocol for treatment of 14 adult patients with Burkitt's lymphoma or L3 acute lymphoblastic leukemia. Ten of 14 patients obtained a complete remission including 5/8 with stage-IV disease or B-ALL. After a median follow-up of 55 months none of these ten patients relapsed. The projected survival after 8 years is 71%. Toxicity was moderate, with one early death; a tumor lysis syndrome occurred in four patients. From our experience we conclude that the BFM-NHL protocol is very effective in adult patients, with a high cure rate and acceptable toxicity, even in advanced stages of disease.

[Sequential high-dose cytarabine therapy in combination with asparaginase in acute myeloid leukemia]. / Sequentielle hochdosierte Cytarabin-Therapie in Kombination mit Asparaginase bei der akuten myeloischen Leukämie.

Between 1984 and 1987 14 patients with acute non-lymphocytic leukemia were treated with sequential high-dose cytosine arabinoside in combination with asparaginase. Twelve patients were suffering from refractory leukemia; in these patients complete remissions were achieved in 58%. The efficacy of this schedule was much better in patients with substantial leukemia cell reduction due to antecedent conventional therapy and no more than 25% blast cells in the bone marrow. In this subgroup complete remissions were achieved in 75% and 86% respectively, taking into account only the completed treatment courses. Beside the well-known side-effects such as alopecia, nausea, vomiting and hepatotoxicity, we observed an increase in severe infections. Three patients died of pulmonary mycosis.

Effects of human alpha-interferon on granulocyte-macrophage progenitor cells (CFU-GM) in vitro.

Two preparations of human interferon (IFN)-alpha were assessed for their influence on granulocyte-macrophage progenitor cells (CFU-GM) in vitro. Both highly purified human IFN-alpha Ly and recombinant IFN-alpha 2a suppressed CFU-GM colony formation in a dose-dependent manner using low-density bone-marrow target cells. Suppression of CFU-GM colony formation was accompanied by an increase in clusters. However, depletion of monocytes, T lymphocytes and B lymphocytes from low-density bone-marrow cells resulted in insensitivity of progenitor cells to IFN-alpha. These results demonstrate that the effects of human IFN-alpha on myeloid progenitor cells (CFU-GM) are mediated by accessory cells within the bone marrow.

Inhibition of growth but not differentiation of normal and leukemic myeloid cells by methylthioadenosine.

Methylthioadenosine (MTA), a coproduct of polyamine biosynthesis, is known to inhibit proliferation in a variety of cell culture systems. In this paper, we show that while MTA inhibits the growth of the human promyelocytic cell line HL-60, it does not interfere with retinoic acid-induced granulocytic or phorbol ester-induced monocytic differentiation of these cells. MTA also inhibits proliferation induced by colony stimulating activity of normal human granulocytic precursor cells grown in suspension culture but does not suppress terminal differentiation of these cells. In contrast to the lack of effect of MTA on granulocytic differentiation which we report here, others have shown that MTA prevents terminal differentiation of murine erythroleukemia cells. That MTA is a normal cellular constituent which inhibits proliferation but not differentiation of normal granulopoietic cells and may have opposing effects on immature cells of erythroid lineage suggests a possible role for this compound in the regulation of hematopoiesis. In addition, MTA may be useful for studying the process of differentiation in the absence of cell proliferation in granulopoietic cells.

Effective treatment of lymphomas of Burkitt's type and B-ALL in adults.

Malignant lymphomas, Burkitt's type, and B-ALL are rarely encountered in adult patients. Rapid initial responses are usually followed by early relapse and death. In a pilot study four adult patients, two presenting with B-ALL, were successfully treated with an aggressive protocol developed by the BFM study group for childhood lymphomas of B-type. Rapid clearance of tumor masses was achieved in all patients; no relapse occurred during an observation period ranging from 19-33 months of complete remission.

[The influence on PHA-stimulation by inhibition of prostaglandin synthesis in vitro in patients with Hodgkin's disease (author's transl)]. / Zur Beeinflussung der PHA-Stimulation durch Hemmung der Prostaglandinsynthese in vitro bei Patienten mit Morbus Hodgkin.

Adherent mononuclear cells may have suppressor functions mediated by prostaglandins (PG). In the present study we tested a large number of normal donors and patients with Hodgkin's disease (HD) using PHA and the prostaglandin inhibitor indomethacin (IM). Stimulation of mononuclear cells from 24 healthy volunteers with PHA led to a mean response of 27 833 cpm; addition of IM caused a 32% increase of 3H-thymidine incorporation. The corresponding values for 30 patients with HD stages IIA-IVB were 14,064 cpm and 70% increase with IM. The effect of the drug was much more pronounced during relapse or progression than in untreated patients. There was an inverse relationship between PHA-response and per cent increase both in normal donors and Hodgkin patients. Depletion of adherent cells using Sephadex G-10 columns abolished the effect of IM completely, but PHA-stimulation was also slightly depressed. Our failure to observe an increase of the mitogen response after removal of monocytes may be related to the technique employed. However, an additional defect of Hodgkin lymphocytes must be considered.

C3 polymorphism in polyacrylamide-gel electrophoresis.

C3 polymorphism in polyacrylamide-gel electrophoresis was identified by crossed immunoelectrophoresis using anti-C3/C3c-serum. Through ageing, treatment of sera with cobra venom factor, endotoxin or with neuramindase, polymorphic bands were seen also in a conversion product and antigenically attributed to C3c. Rare phenotypes were observable in native C3 and in C3c.