Cellular vaccine therapy for cancer

Observations that cells of the immune system are able to kill tumor cells both in vitro and in animal models have provided a compelling rationale for pursuit of a strategy whereby immune cells are administered as a therapeutic vaccine to patients with cancer. The successful outcome of this approach depends upon the ability to deliver this therapy in a manner in which a potent immune response is elicited. By harnessing the capacity of dendritic cells that are pivotal in priming the immune response and using gene therapy approaches to optimise the immune response, this may ultimately prove efficacious in the management of human cancer. Promising reports from recent clinical trials suggest that this may well be a realistic goal.     

The pathogenicity of long versus short fibre samples of amosite asbestos administered to rats by inhalation and intraperitoneal injection.

For many years it has been accepted that fibre dimensions are the most important factor in the development of asbestos related disease with long fibres being more dangerous than short for all types of asbestos. This information has been derived from in vitro experiments and injection or implantation experiments since the kilogramme quantities of specially prepared dusts that are necessary for long term inhalation have not been available. The present study has taken advantage of the availability of a sample of amosite produced so that almost all fibres were less than 5 micron in length. The effects of this dust were compared to dust prepared from raw amosite that contained a very high proportion of long fibres. Previous data from studies with UICC amosite, which was intermediate in length, were also available for comparison. At the end of 12 months of dust inhalation, significantly more short fibre amosite was present in the lung tissue compared to the long but while the long fibre dust caused the development of widespread pulmonary fibrosis, no fibrosis at all was found in animals treated with short fibre. One third of animals treated with long fibre dust developed pulmonary tumours or mesotheliomas but no pulmonary neoplasms were found in animals treated with short fibre dust. Following intraperitoneal injection, the long fibre amosite produced mesotheliomas in 95% of animals with a mean induction period of approximately 500 days. With short fibre dust, only a single mesothelioma developed after 837 days. In previous inhalation studies with UICC amosite, relatively little pulmonary fibrosis had developed and only two benign pulmonary tumours. This would suggest that to produce a significant carcinogenic response in rat lung tissue amosite fibres must be longer than those in the UICC preparation. Following the injection of UICC amosite, however, mesotheliomas developed in the same proportion of animals and with the same mean induction period as with long fibre dust. From this it would appear that while very short fibres exhibit little carcinogenicity to either lung or mesothelial tissues, mesotheliomas can be produced by dust preparations consisting of shorter fibres than are needed to produce tumours.     

Learning disability and epilepsy in an epidemiological sample of individuals with tuberous sclerosis complex

BACKGROUND: Intellectual impairments are a recognized feature of tuberous sclerosis complex (TSC), but the frequency and degree of intellectual impairments has not been systematically studied in large epidemiological samples using standardized measures. As such, the form of the IQ distribution (uni- or bi-modal) has not been established and the relationship between IQ and other features (e.g. epilepsy history) is poorly delineated. To address these shortcomings, we assessed the intellectual abilities of a large epidemiological sample of individuals with TSC, drawn from the 'Wessex' area of SW England and compared them with the abilities of their unaffected siblings. METHOD: Standardized tests were used to estimate the abilities of 108 (56 males, 52 females, median age = 25, range = 4-75) individuals with TSC and 29 unaffected siblings (14 males, 15 females, median age = 18, range = 6-55). Seizure history was obtained from informants and medical records. RESULTS: Estimated IQ was bi-modally distributed: 55.5% had an IQ in the normal range; 14% had mild to severe impairments: and 30.5% had profound disability (IQ < 21). Forty-four per cent of the individuals with TSC had an IQ < 70. In the subset of normally intelligent individuals with TSC, IQ was normally distributed with a mean of 93.6. This mean was significantly lower than the mean IQ of unaffected siblings (IQ = 105.6). All individuals with learning disability had a history of seizures that usually commenced before 12 months of age and that often presented as infantile spasms. Multivariate analyses indicated that a history of seizures as well as a history of infantile spasms was predictive of the degree of intellectual impairment. CONCLUSIONS: Intellectual abilities were bi-modally distributed in a representative sample of individuals with TSC. The likelihood of impairment was associated with a history of seizures, particularly infantile spasms. The genetic and brain basis of these findings requires further investigation.     

In Vivo Complementation of ureB Restores the Ability of Helicobacter pylori To Colonize

The objective of this study was to determine (i) if complementation of ureB-negative Helicobacter pylori restores colonization and (ii) if urease is a useful reporter for promoter activity in vivo. Strains used were M6, M6DeltaureB, and 10 recombinant derivatives of M6 or M6DeltaureB in which urease expression was under the control of different H. pylori promoters. Mice were orally inoculated with either the wild type or one of the mutant strains, and colonization, in vivo urease activity, and extent of gastritis were determined. Of eight M6DeltaureB recombinants tested, four colonized mice. Of those, three had the highest in vitro urease activity of any of the recombinants, significantly different from that of the noncolonizing mutants. The fourth colonizing recombinant, with ureB under control of the cag-15 promoter, had in vitro urease activity which did not differ significantly from the noncolonizing strains. In vivo, urease activities of the four colonizing transformants and the wild-type control were indistinguishable. There were no differences in gastritis or epithelial lesions between mice infected with M6 and those infected with the transformants. These results demonstrate that recovery of urease activity can restore colonizing ability to urease-negative H. pylori. They also suggest that cag-15 is upregulated in vivo, as was previously suggested by demonstrating that it is upregulated upon contact with epithelial cells. Finally, our results suggest that total urease activity and colonization density do not contribute to gastritis due to H. pylori.     

CD4+ T cell-mediated rejection of major histocompatibility complex class I-disparate grafts: A role for alloantibody

Experimental studies of the T cell requirement for rejection of class I major histocompatibility complex (MHC)-disparate grafts have generated controversy over both the autonomy of CD8⁺ T cells and the mechanism whereby CD4⁺ T cells are able to independently mediate rejection. In this study of rejection of RT1A^a class I MHC-disparate rat cardiac and skin allografts by high-responder PVG RT1^u recipients, we show that elimination of CD8⁺ T cells [by anti-CD8 monoclonal antibody (mAb) administration in vivo] fails to prolong graft survival, whereas partial depletion of CD4⁺ T cells (by anti-CD4 mAb treatment) markedly delays rejection of class I-disparate heart grafts, and marginally prolongs survival of skin grafts. Anti-CD4-treated PVG-RT1^u athymic nude rats reconstituted with CD8⁺ T cells failed to reject class I-disparate skin grafts for several weeks and eventual rejection correlated with re-emergence of a small number of donor derived CD4⁺ T cells. Conversely, anti-CD8-treated nude rats reconstituted with CD4⁺ T cells alone rapidly rejected class I-disparate skin grafts. Passive transfer of anti-class I immune serum to anti-CD4-treated euthymic recipients promptly restored their ability to specifically reject a class I-disparate heart graft. Similarly, passive transfer of immune serum to PVG-RT1^u nude rats bearing skin allografts caused destruction of class I-disparate but not third-party grafts. These results demonstrate that CD4⁺ T cells are both necessary and sufficient to cause rejection of class I-disparate heart and skin grafts in this model and that CD4⁺ T cell-dependent alloantibody plays a decisive role in effecting rejection.     

Lung dendritic cells are primed by inhaled particulate antigens, and retain MHC class II/antigenic peptide complexes in hilar lymph nodes for a prolonged period of time

Intratracheal (IT) administration of heat-killed Listeria monocytogenes (HKL) results in an influx of macrophage and dendritic cell (DC) precursors into the lung interstitium. Low-density, FcR+, interstitial lung cells isolated from rats instilled 24 hr before with HKL or vehicle alone, were > 90% Mar1+. After culturing with granulocyte-macrophage colony-stimulating factor (GM-CSF) for 3 days, up to 24% of the loosely adherent cells were DC that stimulated allogeneic T-cell proliferation in an mixed lymphocyte reaction (MLR) assay. After only an overnight incubation with GM-CSF, however, the capacity of interstitial Mar1+ cells to stimulate HKL immune T-cell proliferation without exogenous antigen was low. By contrast, when DC were isolated as major histocompatibility complex (MHC) class II+ cells from rat lungs at 1, 3, 7 and 14 days after HKL instillation and cultured overnight with GM-CSF, their antigen presentation capacity without added exogenous antigen was robust, but declined over the 2-week period. Interestingly, hilar lymph node DC maintained their HKL antigen-presenting capacity for up to 2 weeks after instillation of HKL. Following IT administration of PKH-26 labelled HKL, fluorescent or immunolabelled organisms were detected in OX62+ DC in airway epithelium, lung interstitium and hilar lymph nodes in situ and in MHC class II+ DC isolated from these sites. We conclude that newly immigrated Mar1+ lung DC precursors, while efficient in endocytosing particulate antigens, are incapable of eliciting a significant proliferative response from HKL-sensitized T cells. By contrast, MHC class II+ DC isolated from lungs and incubated overnight with GM-CSF induce vigorous antigen-specific T-cell proliferation. Antigen-loaded lung DC in hilar lymph nodes maintain their antigen presentation capacity for up to 2 weeks.     

Results of unrelated cord blood transplant in fanconi anemia patients: risk factor analysis for engraftment and survival.

We retrospectively analyzed results of unrelated cord blood transplantation (UCBT) in 93 Fanconi anemia (FA) patients. Median age at transplantation was 8.6 years (1-45). The units transplanted were HLA-A, -B, or -DRB1 identical in 12 cases, 1 HLA mismatch in 35 cases, and 2 or 3 HLA differences in 45 cases. The median number of nucleated cells (NC) and CD34+ cells infused of recipient weight was 4.9x10(7)/kg and 1.9x10(5)/kg, respectively. Participating centers selected the preparative regimen of their choice, in 57 patients (61%), it included Fludarabine. Graft-versus-host disease (GVHD) prophylaxis consisted mostly of cyclosporine with prednisone. Cumulative incidence (CI) of neutrophil recovery was 60+/-5% at day +60. In multivariate analysis, Fludarabine containing regimen and NC infused>or=4.9x10(7)/kg were associated with higher probability of recovery. CI of grade II-IV acute and of chronic GVHD (aGVHD, cGVHD) was 32%+/-5% and 16%+/-4%, respectively. Overall survival (OS) was 40%+/-5%. In multivariate analysis, factors associated with favorable outcome were use of Fludarabine in the conditioning regimen, number of NC infused>or=4.9x10(7)/kg, and negative cytomegalovirus (CMV) serology in the recipient. In conclusion, factors easily modifiable such as donor selection and a Fludarabine-containing regimen can considerably improve survival in FA patients given a UCBT. These data are the basis for designing prospective protocols.     

Persistent rat cytomegalovirus (RCMV) infection of the salivary glands contributes to the anti-RCMV humoral immune response

The salivary glands are the major sites of persistent replication of rat cytomegalovirus (RCMV). At several months post infection (pi), infectious RCMV is usually still produced in the salivary glands but not in any other organ or tissue of the rat. To investigate whether the persistence of RCMV in the salivary glands is crucial to the pathogenesis of viral infection, we monitored the progression of RCMV-induced disease in rats from which the salivary glands had been surgically removed (desalivated) as well as in sham-operated rats, both after a lethal and sublethal challenge with RCMV. Desalivation did not have a significant effect on either RCMV-induced morbidity or mortality. As expected, at 1 year pi, relatively high levels of infectious virus were detected in the salivary glands of sham-operated rats, whereas neither infectious virus nor RCMV DNA could be detected in liver, spleen and lungs of these animals. Infectious virus and viral DNA were also undetectable in organs from desalivated animals at 1 year pi. Surprisingly, a difference was found between desalivated and sham-operated rats in the titers of anti-RCMV IgG antibodies, which were significantly higher in sham-operated rats than in desalivated animals at 183, 295 and 365 days pi. This finding indicates that the persistence of RCMV in the salivary glands may contribute significantly to the anti-RCMV humoral immunity of infected rats.     

Detection of heat-stable antigens of Campylobacter jejuni and C. coli by direct agglutination and passive hemagglutination

The two serotyping schemes for the detection of heat-stable antigens of Campylobacter jejuni and Campylobacter coli use the same strains for antiserum production but differ in the detection systems used for identifying agglutination. The Penner method uses passive hemagglutination (PHA) while the Laboratory of Enteric Pathogens method uses the same antisera but in a whole-bacterial-cell direct agglutination (DA) protocol. C. jejuni produces a polysaccharide capsule, which is antigenic, and is the main component detected by the PHA method. The DA method will detect both capsule antigens and lipopolysaccharide (LPS) or lipooligosaccharide (LOS) surface antigens. Comparison of both methods by using a selection of isolates from human infection has shown a range of variation in agglutination specificity, reflecting the differences in antigens detected by the two methods. While 27.4% of the 416 C. jejuni isolates reacted with the antisera raised against the same type strains by either method, the majority showed a range of more complex relationships. None of the 37 C. coli isolates reacted with the same antiserum by both methods. Together the two schemes gave a total of 102 distinct combined serogroups for C. jejuni and 16 for C. coli. Thus, while some clonally related isolates share the same capsule and LOS or LPS antigens, other strains appear to have a common capsule antigen but differ in their LPS or LOS structures or vice versa.     

Intracellular mechanisms governing the acute phase of β-endorphin secretion from the corticotrope in vitro

It is not certain which protein kinase (A, C or both) is involved in the acute phase of β-endorphin (β-EP) release stimulated in the corticotrope by vasopressin (VP) and corticotropin-releasing factor (CRF). We have employed an isolated ovine anterior pituitary cell superfusion system to determine the dynamic effects of forskolin, a protein kinase A (PKA) stimulator, and phorbol 12-myristate 13-acetate (PMA), a protein kinase C (PKC) activator. Both secretagogues stimulated β-EP release within 5 min and therefore both PKA and PKC are potential mediators of the acute phase of hormonal stimulation of the corticotrope. Pretreatment with PMA specifically desensitized the pituitary cell columns to subsequent PMA exposure while not significantly altering sensitivity to forskolin or 50 mM KCl.