Antitumor effect of baicalin from the Scutellaria baicalensis radix extract in B-acute lymphoblastic leukemia with different chromosomal rearrangements.

Acute B-lymphoblastic leukemia (B-ALL) is the most common hematologic malignancy in children. Many cases of B-ALL harbor chromosomal translocations which are often critical determinants of prognosis. Most of them represent altered transcription factors that impact gene transcription or enhance signaling. B-ALLs harboring the mixed-lineage leukemia 1 (MLL1) gene rearrangements represent aggressive, high-risk type of early childhood leukemias that are usually associated with a very poor prognosis. Therefore, there is an urgent need for novel therapeutic agents as well as new treatment strategies. The objective was to examine the vitro inhibitory effects of Scutellaria baicalensis root extract (SBE) in B-ALL cell lines with different chromosomal rearrangements and in leukemic blasts derived from patients' bone marrow (BMCs). In this study we showed that baicalin which is the main component of the SBE possess antitumor activity against all leukemic cell lines especially those with MLL and PBX1 gene rearrangements. Baicalin inhibited cell proliferation, arrested the cell cycle at the G0/G1 phase, and induced cell death through caspase 3/7 activation. Moreover, baicalin treatment inhibited the glycogen synthase kinase-3ß (GSK-3ß) by suppressing its phosphorylation at Y216, and upregulated the downstream mediator of the cell cycle arrest - cyclin dependent kinase inhibitor p27Kip1. Bone marrow derived blasts from B-ALL patients also exhibited varied sensitivity towards baicalin with 72% patients sensitive to the SBE and baicalin treatment. Taken together, our findings provide new insights into the anti-cancer properties of baicalin by showing its diverse mode of action which might be related to the different genetic background.

Immune Consequences of in vitro Infection of Human Peripheral Blood Leukocytes with Vesicular Stomatitis Virus.

BACKGROUND: Oncolytic vesicular stomatitis virus (VSV) can be delivered intravenously to target primary and metastatic lesions, but the interaction between human peripheral blood leukocytes (PBLs) and VSV remains poorly understood. Our study aimed to assess the overall immunological consequences of ex vivo infection of PBLs with VSV. METHODS: Phenotypic analysis of lymphocyte subsets and apoptosis were evaluated with flow cytometry. Caspase 3/7 activity was detected by luminescence assay. Virus release was evaluated in a murine cell line (L929). Gene expression and cytokine/chemokine secretion were assessed by real-time PCR and multiplex assay, respectively. RESULTS: Ex vivo infection of PBLs with VSV elicited upregulated expression of RIG-I, MDA-5, tetherin, IFITM3, and MxA. VSV infection triggered rapid differentiation of blood monocytes into immature dendritic cells as well as their apoptosis, which depended on caspase 3/7 activation. Monocyte differentiation required infectious VSV, but loss of CD14+ cells was also associated with the presence of a cytokine/chemokine milieu produced in response to VSV infection. CONCLUSIONS: Systemic delivery is a major goal in the field of oncolytic viruses. Our results shed further light on immune mechanisms in response to VSV infection and the underlying VSV-PBL interactions bringing hope for improved cancer immunotherapies, particularly those based on intravenous delivery of oncolytic VSV.

The effects of killer cell immunoglobulin-like receptor (KIR) genes on susceptibility to HIV-1 infection in the Polish population.

Killer cell immunoglobulin-like receptors (KIR) are the most polymorphic receptors of natural killer (NK) cells. Their activity diversifies the functions of NK cells in the antiviral immune response, so the presence of certain KIR may affect transmission of HIV-1. The aim of the study was to evaluate the influence of KIR genes on the susceptibility to HIV-1 infection in the Polish population depending on the route of exposure. We determined the frequencies of activating (2DS1, 2DS2, 2DS3, 2DS4f, 2DS4del, 2DS5, 3DS1) and inhibitory (2DL1, 2DL2, 2DL3, 2DL5, 3DL1) KIRs in HIV-1-positive patients (n = 459), individuals exposed to HIV-1 but uninfected (EU, n = 118) and in uninfected, healthy blood donors (BD, n = 98). Analysis was performed using stepwise logistic regression. Apart from KIRs, CCR5-∆32, and CCR2-64I, alleles were also analyzed, as we knew or suspected that these features could affect susceptibility to HIV infection. The regression confirmed the protective effect of CCR5-∆32 (OR = 0.25, p = 0.006) and CCR2-64I (OR = 0.59, p = 0.032) against HIV infection. Among KIR genes, 2DL3 was found to be a protective factor (OR = 0.30, p = 0.015). A similar effect was seen for 3DS1 but only in intravenous drug users (IDUs) (OR = 0.30, p = 0.019), not in sexually exposed people. 2DL5 was found to be a factor facilitating HIV infection (OR = 2.13, p = 0.013). A similar effect was observed for 2DL2 but only in females (OR = 2.15, p = 0.040), and 2DS1 in IDUs (OR = 3.03, p = 0.022). Our results suggest a beneficial role of KIR3DS1 and 2DL3 supporting resistance to HIV infection and a harmful effect of 2DS1, 2DL5, and 2DL2 genes promoting HIV acquisition.

Nanoemulsion-based mucosal adjuvant induces apoptosis in human epithelial cells.

Nanoemulsions (NEs) are adjuvants that enhance antigen penetration of the nasal mucosa, increase cellular uptake of antigens by both epithelial and dendritic cells, and promote the migration of antigen-loaded dendritic cells to regional lymph nodes within 24-h of vaccine administration. The objective of this study was to elucidate cell death caused by W805EC NE and identify caspases and genes associated with death pathways. Consistent with this aim, we show that exposure of human epithelial cells (EC), both RPMI 2650 and FaDu, to NE results in the activation of caspases (1, 3/7, 6, 8, and 9) and the expression of genes involved in apoptotic as well as authophagy and necrosis pathways. Interestingly, the NE activates caspase 8 which promotes "immunogenic apoptosis". The rescue assay was employed to investigate the fate of RPMI 2650 cells treated with W805EC NE. After four-hour treatment with as little as 0.03% of NE no cells were rescued at 72h. Remarkably, immediately after four-hour treatment, the cells morphologically resembled untreated cells and most of the cells were alive. Altogether, these results suggest that NE induces death of human ECs through multiple pathways. Epithelial cell death caused by W805EC may have further implications on antigen uptake, processing, and presentation by DC's.

Deletion of the monkeypox virus inhibitor of complement enzymes locus impacts the adaptive immune response to monkeypox virus in a nonhuman primate model of infection.

Monkeypox virus (MPXV) is an orthopoxvirus closely related to variola virus, the causative agent of smallpox. Human MPXV infection results in a disease that is similar to smallpox and can also be fatal. Two clades of MPXV have been identified, with viruses of the central African clade displaying more pathogenic properties than those within the west African clade. The monkeypox inhibitor of complement enzymes (MOPICE), which is not expressed by viruses of the west African clade, has been hypothesized to be a main virulence factor responsible for increased pathogenic properties of central African strains of MPXV. To gain a better understanding of the role of MOPICE during MPXV-mediated disease, we compared the host adaptive immune response and disease severity following intrabronchial infection with MPXV-Zaire (n = 4), or a recombinant MPXV-Zaire (n = 4) lacking expression of MOPICE in rhesus macaques (RM). Data presented here demonstrate that infection of RM with MPXV leads to significant viral replication in the peripheral blood and lungs and results in the induction of a robust and sustained adaptive immune response against the virus. More importantly, we show that the loss of MOPICE expression results in enhanced viral replication in vivo, as well as a dampened adaptive immune response against MPXV. Taken together, these findings suggest that MOPICE modulates the anti-MPXV immune response and that this protein is not the sole virulence factor of the central African clade of MPXV.

Rhesus macaque rhadinovirus-associated non-Hodgkin lymphoma: animal model for KSHV-associated malignancies.

Rhesus macaque rhadinovirus (RRV) is closely related to Kaposi sarcoma-associated herpesvirus (KSHV) and is associated with the development of B-cell hyperplasia and persistent lymphadenopathy resembling multicentric Castleman disease in rhesus macaques (RMs) coinfected with simian immunodeficiency virus (SIV). Here we investigated whether RMs experimentally infected with SIV and RRV can develop other disease manifestations observed in HIV- and KSHV-infected patients. As reported earlier, inoculation of SIV-infected RMs with RRV results in persistent RRV infection, whereas immunocompetent animals infected with RRV exhibit viremia 2 weeks after infection, followed by a period of no virus detection until they are subsequently made immunodeficient by SIV infection. A subset of animals developed abnormal cellular proliferations characterized as extranodal lymphoma and a proliferative mesenchymal lesion. In situ hybridization and immunohistochemistry analysis indicate RRV is present in both malignancies, and DNA microarray analysis detected viral interleukin-6 (vIL-6) and viral FLICE-like inhibitory protein (vFLIP) transcripts. Reverse-transcriptase polymerase chain reaction analysis confirmed vIL-6 and vFLIP expression, and that of RRV open reading frames 72 and 73, homologs of KSHV open reading frames shown to be expressed in primary effusion lymphoma. These data support the utility of the RRV-/SIV-infected RM as an excellent animal model to investigate KSHV-like pathogenesis.