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Biomedical subjects

K Khalili

Publications and source records attributed to K Khalili.

At least 37 records · Page 2Linked to original sources
PubMed2001

Activation of the IGF-IR system contributes to malignant growth of human and mouse medulloblastomas.

Insulin-like growth factor I receptor (IGF-IR) has been implicated in the normal and malignant growth of many cell types including cells from the central nervous system. In the cerebellar cortex IGF-IR mRNA is found in granular cells and IGF-I stimulation is mitogenic and protects cells from low-potassium-induced apoptosis. Since primitive neuroectodermal tumors/medulloblastomas (PNETs/medulloblastomas) are suspected to originate from the external cerebellar granular layer, it is reasonable to postulate that IGF-IR and/or its signaling molecules may contribute to the transformation of these poorly differentiated cells. To study activation of the IGF-IR system in medulloblastomas, we have utilized an antibody (anti-pY1316) that specifically recognizes the phosphorylated (active) form of the IGF-IR. Medulloblastoma biopsy specimens were positive when examined immunohistochemically with anti-Y1316 antibody. Further analysis of the IGF-IR system was performed in three human (Daoy, TE-671, D283 Med) and four mouse (BsB8, BsB13, Bs-1b, Bs-1c) medulloblastoma cell lines. All the murine cell lines examined express IGF-IR and PI3-kinase at relatively normal levels, and grossly overexpress IRS-1, when compared with normal mouse cerebellum. Within 15 min following IGF-I stimulation both mouse and human cell lines phosphorylate the beta subunit of the IGF-IR, IRS-1, Akt, and MAP kinases. They respond with cell proliferation when stimulated solely with IGF-I and are strongly inhibited when challenged with a dominant negative mutant of the IGF-IR (486/STOP), or with antisense oligonucleotides against the IGF-IR mRNA.

Animals↗
PubMed2001

Detection of JC virus DNA sequences and expression of the viral regulatory protein T-antigen in tumors of the central nervous system.

JC virus (JCV) is a neurotropic polyomavirus infecting greater than 70% of the human population worldwide during early childhood. Replication of JCV in brains of individuals with impaired immune systems results in the fatal demyelinating disease, progressive multifocal leukoencephalopathy (PML). Furthermore, JCV possesses an oncogenic potential and induces development of various neuroectodermal origin tumors including medulloblastomas and glioblastomas in experimental animals. The oncogenecity of JCV is attributed to the viral early gene product, T-antigen, which has the ability to associate with and functionally inactivate well-studied tumor suppressor proteins including p53 and pRB: The observations from laboratory animal experiments have provided a rationale for examining the presence of the JCV DNA sequence and expression of the viral oncogenic protein in human brain tumors. We have examined 85 clinical specimens from the United Kingdom, Greece, and the United States, representing various human brain tumors including oligodendroglioma, astrocytoma, pilocytic astrocytoma, oligoastrocytoma, anaplastic astrocytoma, anaplastic oligodendroglioma, glioblastoma multiforme, gliomatosis cerebri, gliosarcoma, ependymoma, and subependymoma, for their possible association with JCV. We performed gene amplification techniques using a pair of primers that recognize the JCV DNA sequence, and we demonstrated the presence of the viral early sequence in 49 (69%) of 71 samples. More importantly, our results from immunohistochemistry analysis revealed expression of JCV T-antigen in the nuclei of tumor cells in 28 (32.9%) of 85 tested samples. These observations, along with earlier in vitro and in vivo data on the transforming ability of this human neurotropic virus invite additional studies to re-evaluate the role of JCV in the pathogenesis of human brain tumors.

Animals↗
PubMed2001

Cell cycle regulation of NF-kappa b-binding activity in cells from human glioblastomas.

Glioblastoma multiforme is a highly malignant and anaplastic tumor of the central nervous system representing more than 50% of all malignant gliomas. The cell origin of this highly undifferentiated tumor remains obscure, although it is postulated that glioblastomas are developed from astrocytes. The rapid growth of the glioma and the state of its undifferentiation are attributed to the deregulation of several signal transduction pathways and cell cycle events. Recent studies showed diverse functions for the NF-kappa B/Rel family of inducible transcription factors including differentiation, apoptosis, oncogenesis, and cell cycle regulation. We sought to examine the level of NF-kappa B activity throughout the glioma's cell cycle. Results from band-shift studies indicated a biphasic NF-kappa B DNA-binding activity in the nuclei of cycling glioblastoma cells. We showed that NF-kappa B-binding activity maximizes in nuclear extracts at specific cell cycle stages including G0/G1, mid-late G1, and S phase. Results from Northern blotting studies revealed that the differential expression of the NF-kappa B subunits, p50 and p65, may not be responsible for cell cycle stage-specific association of NF-kappa B subunits with DNA. However, results from Western blotting analysis utilizing nuclear extracts from glioma cells throughout the cell cycle demonstrated that the nuclear accumulation of p50 and p65 perfectly correlates with their DNA-binding activity. These observations suggest that the nuclear translocation of the p50/p65 subunit of NF-kappa B in glioma cells is cell cycle stage-dependent and that is distinct from the differential mRNA expression of these genes during glioma cell cycling. The possible role of NF-kappa B in glioma cell formation and regulation of cellular genes by NF-kappa B in these tumor cells is discussed.

Active Transport, Cell Nucleus↗
PubMed2001

Regulation of Puralpha gene transcription: evidence for autoregulation of Puralpha promoter.

The single-stranded DNA and RNA binding protein, Puralpha, has recently received special attention as this protein, by associating with the specific nucleotide sequence (GGN repeats) and/or several important cellular and viral proteins regulates crucial biological events such as transcription, replication, and cell proliferation. In this study, we focused on the promoter activity of the Puralpha upstream DNA sequence and demonstrated that the sequence spanning 6,000 nucleotides upstream of the Puralpha transcription start site has promoter activity in various cell types. Results from promoter deletion studies revealed that this region encompasses various regulatory motifs which differentially participate in the promoter activity of Puralpha in various cells. The transcription start site of Puralpha is surrounded by the GA/GC-rich sequence which exhibits the ability to interact with Puralpha, suggesting a role for autoregulation of Puralpha transcription. Results from co-transfection studies revealed that ectopic expression of Puralpha reduced transcriptional activity of the Puralpha promoter and the region located between amino acid residues, 1-85 of Puralpha is important for the observed autoregulatory event. The regulatory protein of the human neurotropic virus, JCV, T-antigen, which interacts with Puralpha, decreased transcriptional activity of the Puralpha promoter. Co-expression of JCV T-antigen and Puralpha had no significant effect on the suppression of Puralpha gene transcription by either protein. The importance of this finding in light of earlier results showing down regulation of Puralpha during JCV infection of glial cells is discussed.

3T3 Cells↗
PubMed2001

Growth inhibition of glioblastoma cells by human Pur(alpha).

Pur(alpha) is a multifunctional DNA- and RNA-binding protein implicated in a variety of biological events including transcription and replication. Further, this protein has the ability to form a complex with several cellular proteins which are important for cell proliferation including the transcription factor, E2F-1. Pur(alpha) has a modular structure highlighted by alternating three basic aromatic class I and two acidic leucine-rich class II repeats in the central region of the protein. Here, we demonstrate that ectopic overexpression of Pur(alpha) suppresses proliferation of a variety of transformed and tumor cells including human glioblastoma. By utilizing various deletion mutants of Pur(alpha) in colony formation assay, we identified the region spanning the first class II repeat (residues 107-131) and the second class I repeat (residues 148-170) of Pur(alpha) which participate in growth inhibitory action of Pur(alpha). Results from protein transduction experiments using the synthetic peptides representing residues 109-131 and 123-154 of Pur(alpha) in fusion with the arginine rich domain of HIV-1 Tat revealed cellular internalization and nuclear appearance of the Tat-Pur(alpha) fusion peptide after 2 h and its detection in nuclei up to 24 h after treatment. Glioblastoma cells treated with Tat-Pur(alpha) (109-131) and Tat-Pur(alpha) (123-154) exhibited 41 and 47% decrease, respectively, in proliferation. Altogether these results illustrate the efficacy of Pur(alpha) in suppressing glioblastoma cell growth and provide evidence for the potential use of this protein and its derivative(s) in blocking proliferation of tumor cells.

Active Transport, Cell Nucleus↗
PubMed2001

Autoantibodies to TNFalpha in HIV-1 infection: prospects for anti-cytokine vaccine therapy.

Tumor necrosis factor alpha (TNFalpha) is a proinflammatory cytokine principally involved in the activation of lymphocytes in response to viral infection. TNFalpha also stimulates the production of other cytokines, activates NK cells and potentiates cell death and/or lysis in certain models of viral infection. Although TNFalpha might be expected to be a protective component of an antiviral immune response, several lines of evidence suggest that TNFalpha and other virally-induced cytokines actually may contribute to the pathogenesis of HIV infection. Based on the activation of HIV replication in response to TNFalpha, HIV appears to have evolved to take advantage of host cytokine activation pathways. Antibodies to TNFalpha are present in the serum of normal individuals as well as in certain autoimmune disorders, and may modulate disease progression in the setting of HIV infection. We examined TNFalpha-specific antibodies in HIV-infected non-progressors and healthy seronegatives; anti-TNFalpha antibody levels are significantly higher in GRIV seropositive slow/non-progressors (N = 120, mean = 0.24), compared to seronegative controls (N= 12, mean = 0.11). TNFalpha antibodies correlated positively with viral load, (P = 0.013, r = 0.282), and CD8+ cell count (P = 0.03, r = 0.258), and inversely with CD4+ cell count (P = 0.003, r = - 0.246), percent CD4+ cells (P = 0.008, r = -0.306), and CD4 :CD8 ratio (P = 0.033, r = - 0.251). TNFalpha antibodies also correlated positively with antibodies to peptides corresponding to the CD4 binding site of gp160 (P = 0.001, r = 0.384), the CD4 identity region (P = 0.016, r = 0.29), the V3 loop (P = 0.005, r = 0.34), and the amino terminus of Tat (P = 0.001, r = 0.395); TNFalpha antibodies also correlated positively with antibodies to Nef protein (P = 0.008, r = 0.302). The production of anti-TNFalpha antibodies appears to be an adaptive response to HIV infection and suggests the potential utility of modified cytokine vaccines in the treatment of HIV infections as well as AIDS-related and unrelated autoimmune and CNS disorders.

AIDS Vaccines↗
PubMed2001

Radiological features of epiploic appendagitis and segmental omental infarction.

Epiploic appendagitis and segmental omental infarction are more frequently encountered with the increased use of abdominal ultrasound and Computed tomography (CT) in the radiological assessment of the patient who presents clinically with acute abdominal pain. Recognition of specific imaging abnormalities enables the radiologist to make the correct diagnosis. This is important, as the appropriate management of both conditions is often conservative. Follow-up imaging features correlate with clinical improvement.

Abdomen, Acute↗
PubMed2001

Physical and functional interaction between viral and cellular proteins modulate JCV gene transcription.

The lytic phase of JC virus (JCV) appears to be highly complex and remains elusive. A growing body of experimental evidence suggests that the regulation of JCV gene expression and replication requires, in addition to the presence of specific transcription factors, cooperativity between viral and cellular regulatory proteins. This cooperativity may be accomplished by physical interaction of the participant proteins on and/or off the viral DNA sequence. Here, we present evidence of specific physical and functional interaction between a cellular factor, YB-1, and the JCV early protein, T-antigen, and showed that both proteins play important roles in JCV gene transcription. Additionally, our data indicate that YB-1 also functionally interact with another viral protein, designated agnoprotein, which is expressed late during the course of infection, adding further complexity to the currently known picture on JCV gene regulation.

Antigens, Viral, Tumor↗
PubMed2001

CNS invasion by CD14+/CD16+ peripheral blood-derived monocytes in HIV dementia: perivascular accumulation and reservoir of HIV infection.

Increases in circulating CD14+/CD16+ monocytes have been associated with HIV dementia; trafficking of these cells into the CNS has been proposed to play an important role in the pathogenesis of HIV-induced neurological disorders. This model suggests that events outside the CNS leading to monocyte activation initiate the process leading to HIV dementia. To investigate the role of this activated monocyte subset in the pathogenesis of HIV dementia, we examined brain specimens from patients with HIV encephalopathy (HIVE), HIV without encephalopathy, and seronegative controls. An accumulation of perivascular macrophages was observed in HIVE. The majority of these cells identified in microglial nodules and in the perivascular infiltrate were CD14+/CD16+. P24 antigen colocalized with both CD14 and CD16 suggesting that the CD14+/CD16+ macrophage is a major reservoir of HIV-1 infection in CNS. Using CD45/LCA staining, the perivascular macrophage was distinguished from resident microglia. In addition to perivascular and nodular localizations, CD16 also stained ramified cells throughout the white matter. These cells were more ramified and abundant than cells positive for CD14 in white matter. Double staining for p24 and CD16 suggests that these cells were often infected with HIV-1. The prominent distribution of CD14+ cells in HIVE prompted our analysis of soluble CD14 levels in cerebrospinal fluid. Higher levels of soluble CD14 (sCD14) were observed in patients with moderate-to-severe HIV dementia, suggesting the utility of sCD14 as a surrogate marker. CD14+/CD16+ monocytes may play a role in other neurological disorders and sCD14 may be useful for evaluating these conditions.

AIDS Dementia Complex↗
PubMed2001

Interaction of JC virus agno protein with T antigen modulates transcription and replication of the viral genome in glial cells.

In addition to encoding the structural and regulatory proteins, many viruses encode auxiliary proteins, some of which have been shown to play important roles in lytic and latent states of the viruses. The human neurotropic JC virus (JCV) genome encodes an auxiliary protein called Agno whose function remains unknown. Here, we investigated the functional role of JCV Agno protein on transcription and replication of the viral genome in glial cells. Results from transfection of human glial cells showed that Agno protein suppresses both T-antigen-mediated transcription of the viral late gene promoter and T-antigen-induced replication of viral DNA. Affinity chromatography and coimmunoprecipitation assays demonstrated that the Agno protein and T antigen physically interact with each other. Through the use of a series of deletion mutants, we demonstrated that the T-antigen-interacting region of Agno protein is localized to its amino-terminal half and the Agno-interacting domain of T antigen maps to its central portion. Furthermore, utilizing various Agno deletion mutants in functional studies, we confirmed the importance of the Agno-T antigen interaction in the observed down-modulation of T antigen function upon viral gene transcription and DNA replication by Agno protein. Taken together these data suggest that the Agno protein of JCV, which is produced late during the late phase of the lytic cycle, can physically and functionally interact with the viral early protein, T antigen, and downregulate viral gene expression and DNA replication. The importance of these observations in the lytic cycle of JCV is discussed.

Amino Acid Sequence↗
PubMed2001

Expression of a human polyomavirus oncoprotein and tumour suppressor proteins in medulloblastomas.

AIMS: Although the aetiology of medulloblastoma remains elusive, several lines of evidence suggest an association with the human neurotropic polyomavirus JC and its oncoprotein T antigen. The tumour forming properties of JC virus T antigen are the result, at least in part, of its ability to bind and inactivate tumour suppressor/cell cycle regulatory proteins, such as p53 and the retinoblastoma family of proteins. METHODS: To examine potential relations between these factors, immunohistochemistry was used to determine associations between the T antigen and the expression of p53 and the retinoblastoma proteins pRb, p107, and Rb2/p130 in eight medulloblastomas. RESULTS: Only the three medulloblastomas with T antigen expression also showed nuclear positivity with antibodies to p53. Although immunohistochemistry detected nuclear labelling for pRb in five of the cases, the three that were positive for T antigen showed the highest pRb labelling. The retinoblastoma related proteins p107 and Rb2/p130 were also immunopositive in most T antigen positive medulloblastomas. Double label immunohistochemistry also demonstrated p53 and pRb positivity in the same cells that were T antigen positive. CONCLUSIONS: These correlations suggest that associations between T antigen and p53 and/or T antigen and pRb occur in some of these tumours. These data provide indirect evidence that JC virus, acting through T antigen, might be involved in the formation and progression of medulloblastoma.

Adolescent↗
PubMed2001

Concomitant progressive multifocal leucoencephalopathy and primary central nervous system lymphoma expressing JC virus oncogenic protein, large T antigen.

This report describes the concomitant occurrence of the JC virus (JCV) induced demyelinating disease progressive multifocal leucoencephalopathy (PML) and a primary central nervous system lymphoma (PCNS-L) in a patient with AIDS. Postmortem neuropathological examination revealed characteristic features of PML including multiple lesions of demyelination, enlarged oligodendrocytes with hyperchromatic nuclei (many containing eosinophilic intranuclear inclusions), and enlarged astrocytes with bizarre hyperchromatic nuclei. Immunohistochemical analysis demonstrated the expression of the JCV capsid protein VP-1 in the nuclei of infected oligodendrocytes and astrocytes. The PCNS-L lesion located in the basal ganglia was highly cellular, distributed perivascularly, and consisted of large atypical plasmacytoid lymphocytes. Immunohistochemical examination of this neoplasm identified it to be of B cell origin. Moreover, expression of the JCV oncogenic protein, T antigen, was detected in the nuclei of the neoplastic lymphocytes. This study provides the first evidence for a possible association between JCV and PCNS-L.

AIDS-Related Opportunistic Infections↗
PubMed2001

Human neurotropic JC virus and its association with brain tumors.

JC virus (JCV) is a human polyomavirus known as the causative agent of the fatal demyelinating disease, Progressive Multifocal Leukoencephalopathy (PML). Further, in experimental animals this virus causes a broad range of tumors of central nervous system origin. Recent studies have suggested the association of JCV with several human tumors most notably malignant brain tumors of childhood, medulloblastoma. The development of tumors by JCV is most likely through mechanisms involving inactivation of tumor suppressors and de-regulation of signaling pathways such as Wnt by the viral early protein, T-antigen. The neurotropic nature of JCV along with the overwhelming evidence for its oncogenic potential in laboratory animals and its detection in significant numbers of human medulloblastomas invite the re-evaluation of the role for JCV in the development of human brain tumors.

Animals↗
PubMed2001

Aberrant localization of the neuronal class III beta-tubulin in astrocytomas.

BACKGROUND: The class III beta-tubulin isotype (betaIII) is widely regarded as a neuronal marker in development and neoplasia. In previous work, we have shown that the expression of betaIII in neuronal/neuroblastic tumors is differentiation dependent. In contrast, the aberrant localization of this isotype in certain nonneuronal neoplasms, such as epithelial neuroendocrine lung tumors, is associated with anaplastic potential. OBJECTIVE: To test the generality of this observation, we investigated the immunoreactivity profile of betaIII in astrocytomas. DESIGN: Sixty archival, surgically excised astrocytomas (8 pilocytic astrocytomas, WHO grade 1; 18 diffuse fibrillary astrocytomas, WHO grade 2; 4 anaplastic astrocytomas, WHO grade 3; and 30 glioblastomas, WHO grade 4), were studied by immunohistochemistry using anti-betaIII monoclonal (TuJ1) and polyclonal antibodies. A monoclonal antibody to Ki-67 nuclear antigen (NC-MM1) was used as a marker for cell proliferation. Antibodies to glial fibrillary acidic protein (GFAP) and BM89 synaptic vesicle antigen/synaptophysin were used as glial and neuronal markers, respectively. RESULTS: The betaIII immunoreactivity was significantly greater in high-grade astrocytomas (anaplastic astrocytomas and glioblastomas; median labeling index [MLI], 35%; interquartile range [IQR], 20%-47%) as compared with diffuse fibrillary astrocytomas (MLI, 4%; IQR, 0.2%-21%) (P <.0001) and was rarely detectable in pilocytic astrocytomas (MLI, 0%; IQR, 0%-0.5%) (P <.0001 vs high-grade astrocytomas; P <.01 vs diffuse fibrillary astrocytomas). A highly significant, grade-dependent relationship was observed between betaIII and Ki-67 labeling and malignancy, but this association was stronger for Ki-67 than for betaIII (betaIII, P <.006; Ki-67, P <.0001). There was co-localization of betaIII and GFAP in neoplastic astrocytes, but no BM89 synaptic vesicle antigen/synaptophysin staining was detected. CONCLUSIONS: In the context of astrocytic gliomas, betaIII immunoreactivity is associated with an ascending gradient of malignancy and thus may be a useful ancillary diagnostic marker. However, the significance of betaIII-positive phenotypes in diffuse fibrillary astrocytomas with respect to prognostic and predictive value requires further evaluation. Under certain neoplastic conditions, betaIII expression is not neuron specific, calling for a cautious interpretation of betaIII-positive phenotypes in brain tumors.

Adolescent↗
PubMed2001

Helix-destabilizing properties of the human single-stranded DNA- and RNA-binding protein Puralpha.

Puralpha is a ubiquitous nucleic acid-binding protein which has been implicated in the control of eukaryotic gene transcription. Further, Puralpha associates with DNA sequences positioned in close proximity to viral and cellular origins of replication suggesting a role for this protein in DNA replication. As initiation of transcription and replication require alteration in the structure of duplex DNA, we investigated the DNA unwinding activity of this single-stranded nucleic acid-binding protein. Here we demonstrate that Puralpha has the ability to displace an oligonucleotide annealed to single-stranded M13 DNA. The helix unwinding activity of Puralpha was dose-, time- and temperature-dependent and ATP-independent. Results from mapping studies revealed that the central region of Puralpha, spanning amino acids 72-274, was important for the helix-destabilizing activity of this protein. The region of Puralpha which was involved in the helix-destabilizing activity mapped to the DNA-binding domain of this protein. Results from heat inactivation experiments demonstrated that the helix-destabilizing activity of Puralpha correlates with its capacity to interact with DNA containing the PUR element. Taken together, these studies demonstrate that Puralpha possesses helix-destabilizing activity and that this activity maps to and correlates with its ability to interact with DNA.

Adenosine Triphosphate↗
PubMed2001

Oncogenic potential of human neurotropic virus: laboratory and clinical observations.

Cancer is a multi-step disease involving a series of genetic alterations that result in the loss of control of cell proliferation and differentiation. Such genetic alterations could emerge from the activation of oncogenes and the loss or malfunctioning of tumor suppressor gene activity. Our understanding of cancer has greatly increased through the use of DNA tumor viruses and their transforming proteins as a biological tool to decipher a cascade of events that lead to deregulation of cell proliferation and subsequent tumor formation. For the past ten years our laboratory has focused on the molecular biology of the human neurotropic papovavirus, JCV. This virus causes progressive multifocal leukoencephalopathy, a fatal neurodegenerative disease of the central nervous system in immunocompromised patients. JCV is a common human virus that infects more than 80% of humans but does not induce any obvious clinical symptoms. The increased incidence of acquired immune deficiency syndrome and the use of immunosuppressive chemotherapy have dramatically raised the incidence of PML. The coincidental occurrence of malignant astrocytes and oligodendrocytes in PML patients, coupled with the induction of glioblastoma in JCV-infected nonhuman primates, provides intriguing speculation on the association between JCV and CNS malignancies. In this report we discuss clinical data and laboratory observations pointing to the direct involvement of JCV in cancer.

Animals↗
PubMed2000

Cooperative interaction between HIV-1 regulatory proteins Tat and Vpr modulates transcription of the viral genome.

The virion-associated protein of human immunodeficiency virus, type 1 (HIV-1), Vpr, is a small protein with 96 amino acid residues that has the ability to modulate transcription of HIV-1 long terminal repeat (LTR) promoter activity and affects several cellular functions. In this study we have employed molecular approaches to further investigate the mechanism by which Vpr exerts its regulatory effect upon the LTR. We show that by structural and functional interaction with Tat, a potent viral regulatory protein, Vpr synergistically enhances the transcriptional activity of the HIV-1 LTR. Because Tat utilizes cyclin T1 and its partner, CDK9 to elevate the level of transcription from the LTR, we examined the cooperativity between Vpr, Tat, and cyclin T1/CDK9 on viral gene transcription. Results from co-transfection studies indicated superactivation of LTR by Tat and cyclin T1/CDK9 in the presence of wild type Vpr. This activation was not observed with the R73S mutant of Vpr, which contains arginine to serine transition at residue 73. Interestingly, expression of R73S mutant in cells exerts a negative effect on the observed superactivation of the LTR by Tat, cyclin T1/CDK9, and wild type Vpr. Results from protein-protein interaction studies indicated that Vpr is associated with both Tat and cyclin T1 in cells expressing these proteins. Use of deletion mutant proteins in binding studies revealed that the binding sites for Tat and Vpr within cyclin T1 are distinct and that association of these two viral proteins with cyclin T1 is independent from each other. These observations suggest a working model on the cooperative interaction of Vpr with viral and cellular proteins and its involvement in control of viral gene transcription and replication. Moreover identification of R73S mutant of Vpr provides a new therapeutic avenue for controlling HIV-1 gene transcription and replication in the infected cells.

Arginine↗