Design and feasibility of PASSIST, a passive instrument positioner.

Background And Purpose: During minimally invasive procedures, an assistant controls the camera and often a laparoscopic grasper. Ideally, the surgeon should be able to manipulate the instruments because the indirect way of control complicates the surgeon's observation and actions and disturbs eye-hand coordination. Reported replacements for the assistant are active positioners, "robots," such as the Aesop and the EndoAssist.

Haploinsufficiency of AML1 affects the temporal and spatial generation of hematopoietic stem cells in the mouse embryo.

The AML1:CBFbeta transcription factor complex is essential for definitive hematopoiesis. Null mutations in mouse AML1 result in midgestational lethality with a complete lack of fetal liver hematopoiesis. While the cell autonomous nature and expression pattern of AML1 suggest an intrinsic role for this transcription factor in the developing hematopoietic system, no direct link to a functional cell type has been made.

CFU-S(11) activity does not localize solely with the aorta in the aorta-gonad-mesonephros region.

The aorta-gonad-mesonephros (AGM) region is a potent hematopoietic site in the midgestation mouse conceptus and first contains colony-forming units-spleen day 11 (CFU-S(11)) at embryonic day 10 (E10). Because CFU-S(11) activity is present in the AGM region before the onset of hematopoietic stem cell (HSC) activity, CFU-S(11) activity in the complex developing vascular and urogenital regions of the AGM was localized. From E10 onward, CFU-S(11) activity is associated with the aortic vasculature, and is found also in the urogenital ridges (UGRs).

An intrinsic but cell-nonautonomous defect in GATA-1-overexpressing mouse erythroid cells.

GATA-1 is a tissue-specific transcription factor that is essential for the production of red blood cells. Here we show that overexpression of GATA-1 in erythroid cells inhibits their differentiation, leading to a lethal anaemia. Using chromosome-X-inactivation of a GATA-1 transgene and chimaeric animals, we show that this defect is intrinsic to erythroid cells, but nevertheless cell nonautonomous.

Definitive hematopoietic stem cells first develop within the major arterial regions of the mouse embryo.

The aorta-gonad-mesonephros (AGM) region is a potent hematopoietic site within the mammalian embryo body, and the first place from which hematopoietic stem cells (HSCs) emerge. Within the complex embryonic vascular, excretory and reproductive tissues of the AGM region, the precise location of HSC development is unknown. To determine where HSCs develop, we subdissected the AGM into aorta and urogenital ridge segments and transplanted the cells into irradiated adult recipients.

Cross-lineage T cell receptor gene rearrangements occur in more than ninety percent of childhood precursor-B acute lymphoblastic leukemias: alternative PCR targets for detection of minimal residual disease.

A large series of 202 childhood precursor-B cell acute lymphoblastic leukemia (ALL) patients was analyzed by Southern blotting (SB) for cross-lineage rearrangements and/or deletions in the T cell receptor TCRB, TCRG and TCRD loci. In 93% (187/201) of the precursor-B-ALL patients one or more genes were rearranged and/or deleted. TCRB gene rearrangements were found in 35% (69/196), TCRG gene rearrangements in 59% (113/192), TCRD gene rearrangements in 55% (112/202), and isolated monoallelic or biallelic deletions of TCRD loci in 34% (68/202) of the cases.

Prognostic value of minimal residual disease in acute lymphoblastic leukaemia in childhood.

Background: Sensitive techniques for detection of minimal residual disease (MRD) at degrees of one leukaemic cell per 10(3)-10(6) cells (10(-3)-10(-6)) during follow-up of children with acute lymphoblastic leukaemia (ALL) can provide insight into the effectiveness of cytotoxic treatment. However, it is not yet clear how information on MRD can be applied to treatment protocols.

Methods: We monitored 240 patients with childhood ALL who were treated according to national protocols of the International BFM Study Group.

L1-specific protection from tumor challenge elicited by HPV16 virus-like particles.

A single injection of HPV16 L1 virus-like particles induced potent CD8-mediated protection from tumor challenge by C3 cells, a line derived from embryonic mouse cells transfected with the HPV16 genome. L1 RNA, but not protein, was detected biochemically in C3 cells. These results indicate that low-level expression of HPV16 L1 can occur in proliferating cells and serve as a tumor vaccine target.

Bone marrow cellular composition in Listeria monocytogenes infected mice detected using ER-MP12 and ER-MP20 antibodies: a flow cytometric alternative to differential counting.

Detailed assessment of bone marrow cellular composition is essential in the evaluation of various experimental in vivo systems, such as expression of transgenes, null mutations and stimulation of host defence in infection. Traditional morphological analysis of mouse bone marrow is laborious, requires specific cytological expertise, and is somewhat subjective. As an alternative, we have examined whether double labelling of bone marrow with the anti-precursor monoclonal antibodies ER-MP12 and ER-MP20 could be used for differential analysis by flow cytometry, as these antibodies define six relatively homogeneous cell populations in mouse bone marrow.

Immunoglobulin and T cell receptor gene rearrangement patterns in acute lymphoblastic leukemia are less mature in adults than in children: implications for selection of PCR targets for detection of minimal residual disease.

In order to gain insight into immunoglobulin (Ig) and T cell receptor (TCR) gene rearrangements in adult acute lymphoblastic leukemia (ALL), we studied 48 adult patients: 26 with precursor-B-ALL and 22 with T-ALL. Southern blotting (SB) with multiple DNA probes for the IGH, IGK, TCRB, TCRG, TCRD and TAL1 loci revealed rearrangement patterns largely comparable to pediatric ALL, but several differences were found for precursor-B-ALL patients. Firstly, adult patients showed a lower level of oligoclonality in the IGH gene locus (five out of 26 patients; 19%) despite a comparable incidence of IGH gene rearrangements (24 out of 26 patients; 92%).

Chimeric papillomavirus virus-like particles elicit antitumor immunity against the E7 oncoprotein in an HPV16 tumor model.

Papillomavirus-like particles (VLPs) are a promising prophylactic vaccine candidate to prevent human papillomavirus (HPV) infections and associated epithelial neoplasia. However, they are unlikely to have therapeutic effects because the virion capsid proteins are not detected in the proliferating cells of the infected epithelia or in cervical carcinomas. To increase the number of viral antigen targets for cell-mediated immune responses in a VLP-based vaccine, we have generated stable chimeric VLPs consisting of the L1 major capsid protein plus the entire E7 (11 kDa) or E2 (43 kDa) nonstructural papillomavirus protein fused to the L2 minor capsid protein.

Immunization with human papillomavirus type 16 (HPV16) oncoprotein-loaded dendritic cells as well as protein in adjuvant induces MHC class I-restricted protection to HPV16-induced tumor cells.

Human papillomavirus (HPV) E6 and E7 oncoproteins are attractive targets for T-cell-based immunotherapy of cervical cancer. In this study, we demonstrate that dendritic cells (DCs) pulsed with HPV16 E7 protein are not only recognized in vitro by E7-specific CTLs but also elicit E7-specific CTL responses in vivo, associated with protection against a challenge with syngeneic HPV16-induced tumor cells. Vaccination with soluble E7 protein in incomplete Freund's adjuvant likewise induces E7-specific CTL responses associated with tumor protection.

Heterogeneity in junctional regions of immunoglobulin kappa deleting element rearrangements in B cell leukemias: a new molecular target for detection of minimal residual disease.

Virtually all immunoglobulin kappa (IGK) gene deletions are mediated via rearrangements of the so-called kappa deleting element (Kde). Kde rearrangements occur either to Vkappa gene segments (Vkappa-Kde rearrangements) or to the heptamer recombination signal sequence in the Jkappa-Ckappa intron. Kde rearrangements were analyzed by the polymerase chain reaction (PCR) and heteroduplex analysis in 130 B-lineage leukemias: 63 precursor-B-acute lymphoblastic leukemias (ALL) and 67 chronic B cell leukemias.

Interactions between organic anions, micelles and vesicles in model bile systems.

Biliary lipid secretion probably involves both 'micellization' and 'vesiculization' of bile-canalicular membrane lipids. Several hydrophilic organic anions inhibit the secretion of lipids into the bile without altering bile salt secretion [Verkade, Vonk and Kuipers (1995) Hepatology 21, 1174-1189]. Hydrophobic organic anions do not interfere with biliary lipid secretion.

Dietary n-3 fatty acids increase spleen size and postendotoxin circulating TNF in mice; role of macrophages, macrophage precursors, and colony-stimulating factor-1.

In experimental studies in mice, dietary supplementation with n-3 fatty acids (FA) alleviates inflammation and increases resistance to infection. Nevertheless, TNF production capacity was found to be increased in n-3 FA-fed mice. We previously found increased relative spleen weights in n-3 FA-fed mice.

High-level expression of the ER-MP58 antigen on mouse bone marrow hematopoietic progenitor cells marks commitment to the myeloid lineage.

Studies on the early events in the differentiation of the nonspecific immune system require the identification and isolation of myeloid-committed progenitor cells. Using the monoclonal antibodies (mAb) ER-MP12 and ER-MP20, generated against immortalized macrophage precursors, we have shown previously that the earliest macrophage colony-stimulating factor (M-CSF)-responsive cells in the bone marrow have the ER-MP12hi 20- phenotype. In addition, we found that the ER-MP12hi 20- subset (comprising about 2 % of total nucleated marrow) contains progenitor cells of all hematopoietic lineages.

Peptide specificity of alloreactive CD4 positive T lymphocytes directed against a major histocompatibility complex class I disparity.

The mouse strains C57BL/6 (B6, H2b) and Kbm1 mutant bm1 have a defined difference of three amino acids at position 152, 155, and 156 in the MHC class I K molecule. This causes a change in the side and the bottom of the antigen presenting groove of the K molecule resulting in strong allogeneic responses in vitro and in vivo. Here we report on the peptide specificity of CD4+ T cells of B6 origin directed against the Kbm1 mutant and speculate on the peptide specificity of CD8+ bm1-specific T lymphocytes of B6 origin.

Inactivation of Btk by insertion of lacZ reveals defects in B cell development only past the pre-B cell stage.

Bruton's tyrosine kinase (Btk) is a cytoplasmic protein kinase that is defective in X-linked agammaglobulinaemia in man and in X-linked immunodeficiency in the mouse. There is controversy regarding the stages of B cell development that are dependent on Btk function. To determine the point in B cell differentiation at which defects in Btk become apparent, we generated a mouse model by inactivating the Btk gene through an in-frame insertion of a lacZ reporter by homologous recombination in embryonic stem cells.

Detection of minimal residual disease in acute leukemia patients.

Diagnostic techniques, routinely used in clinical practice for monitoring acute leukemia patients, are able to detect only 1-5% of malignant cells. At present, two main techniques are being introduced for detection of minimal residual disease (MRD) in leukemia, namely immunological marker analysis and the polymerase chain reaction (PCR) technique with general sensitivity of 10(-4)-10(-5). Immunological marker analysis allows detection of unusual and aberrant immunophenotypes, and is usually performed by flow cytometry.

Characterization of the cholesterol crystallization-promoting low-density particle isolated from human bile.

Background & Aims: Biliary concanavalin A-binding glycoprotein (CABG) contains cholesterol crystallization-promoting activity that is not accounted for by the pronucleators that have been characterized in this fraction. The aim of this study was to isolate and characterize the missing activity.

Methods: Biliary glycoprotein was isolated using concanavalin A-Sepharose.

Induction of heat-stable antigen expression by phagocytosis is involved in in vitro activation of unprimed CTL by macrophages.

Costimulation is required for activation of unprimed CTL. While the costimulatory molecules B7 and heat stable Ag (HSA) play a role in CTL response induction generated by dendritic cells, HSA but not B7 contributes to primary CTL response induced by macrophages (M phi), but only if particulate material has been ingested. This is a finding that correlates well with the observation that soon after phagocytosis of latex beads by M phi, cell surface expression of HSA rapidly increases.

Phagocyte-induced antigen-specific activation of unprimed CD8+ T cells in vitro.

The strict segregation of the major histocompatibility complex (MHC) class I and class II loading pathways has been challenged by recent reports indicating that MHC class I molecules can acquire antigen in the phagocytic pathway. We now show that this alternative peptide loading pathway can be used efficiently to generate macrophages able to activate unprimed antigen-specific cytotoxic T cells in vitro. Short peptides (8-11 residues), administered in the phagocytic pathway at nanomolar concentrations, were found to be effective in specifically activating naïve cytotoxic T lymphocytes (CTL) in vitro, but longer peptides or whole protein antigen were not.

Leishmania (Viannia) lainsoni occupies a unique niche within the subgenus Viannia.

Leishmania lainsoni has recently been recognized as a new peripylarian species belonging to the subgenus Viannia and the L. braziliensis complex. It has been isolated from its sandfly vector, reservoir host and cutaneous lesions of human patients.