CD4-independent protective cytotoxic T cells induced in early life by a non-replicative delivery system based on virus-like particles

The relative immaturity of the neonatal immune system limits CD4(+) Th1 and cytotoxic T lymphocyte (CTL) responses, and represents a significant challenge for the development of vaccines against intracellular pathogens. In this report, we demonstrate the ability of a non-replicative delivery system based on parvovirus-like particles (VLP) to induce CTL responses in the neonatal period. A single immunization of 1-week-old BALB/c mice with recombinant VLP carrying a CD8(+) T cell determinant from lymphocytic choriomeningitis virus (VLP-LCMV) induced antigen-specific CD8(+) cytotoxic T cells that were similar to those elicited by adult immunization, as assessed by cytotoxic activity, interferon (IFN)-gamma secretion, cytotoxic precursor cell frequencies, in vitro avidity for antigen and protective activity against viral challenge. These CTL responses are elicited within 2 weeks of a single immunization, in the absence of adjuvant and independently of the presence and help of CD4(+) T cells, highlighting the potential of VLP as candidate vaccine vectors in early life.     

Genetic analysis of cercarial emergence rhythms ofSchistosoma mansoni

Using two chronobiological variants ofSchistosoma mansoni (a blood fluke infecting man) from Guadeloupe (French West Indies), we carried out experimental crossbreeding between schistosomes with an early and those with a late cercarial shedding pattern. The results obtained on the F₁ (intermediate shedding patterns) and F₂ generations (early, intermediate, and late patterns) demonstrate that the cercarial emergence rhythms of schistosomes are genetically determined. This genetic variability is interpreted as a consequence of the selective pressure exerted by the two different hosts (man and rat) implicated in the life cycle ofS. mansoni from the Guadeloupean focus of schistosomiasis.This work received financial support from the UNDP/World Bank/WHO Special Programme for Research and Training in Tropical Diseases and the CNRS.     

Histological detection of minimal metastatic involvement in axillary sentinel nodes: a rational basis for a sensitive methodology usable in daily practice.

There is no consensus method for the histological analysis of axillary sentinel nodes (SN). This study aimed to (1) assess the rate of occult metastases in SN using large serial sectioning and immunohistochemistry (IHC), (2) evaluate whether occult metastases were predictive of metastases in the downstream axillary nodes, and (3) specify a methodology of analysis of SN that could be both sensitive and applicable in daily practice. One hundred three patients with breast carcinoma underwent SN biopsy and then axillary dissection. SN free of tumor at standard examination of one section were sectioned at six levels (150-microm intervals) and immunostained for cytokeratin. The number and localization of labeled metastatic cells (occult metastases) were recorded. In 29 of the 103 patients (28%), SN were found to be metastatic after standard examination. The SN of the remaining 74 patients were further analyzed using IHC. Occult metastases were detected in 35 of these patients (47.3%), leading to an overall SN involvement rate of 62% (29+35/103). In 33 of these 35 cases, the plurality and the dispersion of the immunostained cells implied that the screening of only 3 of the 6 levels would have led to the detection of diagnostic positive events. Only one of the 35 patients (2.8%) with occult metastases showed metastatic lymph node in the downstream axilla. In our series of axillary SN, the analysis of one standard histologic section and, when negative, of only three additional sections after IHC revealed >60% of metastasis or occult metastasis. Metastasis detected by standard analysis had a high predictive value of downstream node metastasis, whereas the predictive value of occult metastasis revealed by IHC was poor. The clinical significance of occult metastases in SN needs to be specified by long-term follow-up analysis.     

Activation of the CD3/T cell receptor (TcR) complex or of protein kinase C potentiate adenylyl cyclase stimulation in a tumoral T cell line: involvement of two distinct intracellular pathways.

A monoclonal antibody (OKT3) directed against the T cell receptor (TcR)/CD3 molecular complex, as well as a protein kinase C (PKC) activator (phorbol 12-myristate 13-acetate, PMA) were added to a culture of tumoral Jurkat T cells, in order to precise the sequence of intracellular signals leading to T cell activation. The experiments were performed in the presence or in absence of various stimulators of adenylate cyclase (AC) such as forskolin (FK), cholera toxin (CT) or prostaglandin E2 (PGE2). OKT3 increased inositol phosphate (IP) production; in parallel, it induced a slight accumulation of cAMP. The effect was markedly potentiated in presence of FK or CT, and to a lesser extent in the presence of PGE2. FK stimulated adenylate cyclase of Jurkat cell membranes, but the effect was not potentiated by OKT3, suggesting that potentiation of cAMP accumulation requires intact cells and is not mediated by direct receptor coupling. On the other hand, elevated cAMP accumulation induced a negative feedback on IP production. The effect of OKT3 on cAMP was mimicked by A23187, a Ca2+ ionophore, and abolished in the absence of extracellular Ca2+. PMA had the same effect as OKT3 on basal or FK- and CT-induced accumulation of cAMP. In contrast, it inhibited the PGE2 effect on the cyclic nucleotide. After desensitization of PKC by pretreatment with a high concentration of PMA, the phorbol ester was no longer effective. Under those conditions, facilitation by OKT3 of FK-induced accumulation of cAMP was preserved, whereas potentiation by the monoclonal antibody of the PGE2 stimulation of AC was even enhanced. The data indicate that cAMP accumulation indirectly elicited by phospholipase C activation is, at least partly, mediated by IP-dependent Ca2+ mobilization, while PKC is preferentially effective as an inhibitor of PGE2 stimulation.     

Thymic Medulla Epithelial Cells Acquire Specific Markers by Post-Mitotic Maturation

The development of thymocyte subsets and of the thymic epithelium in SCID and RAG-2^-/– mice was monitored after normal bone-marrow-cell transfer. The kinetics of thymic reconstitution and their relationships with cell proliferation were investigated by using bromodeoxyuridine to detect DNA-synthesizing cells among lymphoid cells by 3-color flow cytometry, and in epithelial compartments by staining frozen sections. Thymocytes started to express CD8 and CD4 10 days after transfer, simultaneously with extensive proliferation. The first mature CD4⁺ single-positive cells were generated, from resting CD4⁺CD8⁺ cells after day 15. During this day 10–15 period, many epithelial cells positive for cortexspecific or panepithelial markers were labeled with BrdUrd after pulse-injection. Organized medullary epithelium also developed after day,15, that is, synchronously with the appearance of mature thymocytes, but medullary cells were never found BrdUrd⁺. These results suggest that, in these models, the reconstitution of the thymic epithelial network proceeds through expansion of preexisting cortical or undifferentiated cells and by later maturation (acquisition of specific markers) of medullary cells. This last process is dependent of the presence of mature thymocytes.     

Analysis through mathematical models of the dynamics of the adrenal cortical response to ACTH in the rat

The distribution and metabolism of ACTH and the dynamics of the adrenal cortical response to this tropic hormone were characterized in the rat, through mathematical models involving data derived from experiments, where plasma corticosterone concentrations were measured following both single injections and infusions of ACTH. The models, which incorporate a previously established model of the dynamics of plasma corticosterone, were statistically validated. The simulated dynamics of the different processes linking ACTH secretion by the adenohypophysis to corticosterone secretion by the adrenal cortex include: (1) a variable MCR for plasma ACTH, modeled as the sum of a constant and a saturable degradation process; (2) the ability of the adrenal gland to secrete at a maximal rate aven after the plasma ACTH concentration has become negligible, modeled as the accumulation of an intermediary productZ directly controlled by the plasma ACTH concentration; (3) a saturable secretion with a small time constant and which, for single injections, always starts in the same fashion, modeled as a “synthesis process” whose input is a saturable function ofZ; (4) an immediate fall of the plasma corticosterone concentration at the end of the ACTH infusions, modeled as a “release process” also controlled byZ. Supported by grants from the Medical Research Council of Canada (MT-1205, MT-1555, and MA-4970), and from INSERM, France (CRL-76-5-020-4).     

The gene encoding CD23 leukocyte antigen (FCE2) is located on human chromosome 19

Using Southern blot analysis of DNAs from human×rodent cell hybrids, we have mapped the CD23 leukocyte antigen gene (FCE2) to human chromosome 19.     

5′-Cytosine DNA-methyltransferase mRNA levels in hereditary colon carcinoma

 DNA methylation plays an important part in the regulation of gene expression. Alterations in DNA methylation in tumours have been reported and have been used to generate hypotheses about mutagenesis and silencing of tumour suppressor genes. However, the underlying mechanism is still poorly understood, and conflicting data on the levels of overexpression of 5′-cytosine DNA methyltransferase in sporadic colon carcinoma have been published. We used a competitive RT-PCR assay for quantification of mRNA of 5′-cytosine DNA methyltransferase in colon biopsies obtained from patients with hereditary colon carcinoma syndromes and compared the results with those obtained in a control group. No significant difference was found between the flat mucosa of FAP patients and the mucosa of the control group. In FAP and HNPCC patients, the 5′-cytosine DNA methyltransferase mRNA levels of adenomas were significantly higher (P<0.05) than of flat mucosa in the same group, but both showed great variability from patient to patient. Our findings suggest that the mRNA levels of methyltransferase cannot be used as predictive marker for screening in families affected by hereditary colon carcinoma. Received: 20 July 1998 / Accepted: 21 September 1998     

PK/PD modeling of aminoglycoside nephrotoxicity

Aminoglycosides are bactericidial antibiotics with a serum concentration-dependent activity. They are mainly eliminated by the kidneys and the main difficulty arising in clinical use is their uptake by the renal cortex which leads to nephrotoxicity. An ototoxicity is also reported. We propose a PK/PD modelling of aminoglycoside nephrotoxicity which unifies more fourty years of physiological knowledge. This deterministic model successively describes the pharmacokinetics of aminoglycosides, their storage into renal cortex, their effect on renal cells, their consequences on the renal function through tubuloglomerular feedback and the changes in the serum concentrations of creatinine that is considered as a toxicity marker. The simulation of the model displays the leading effect of the shape and daily-time of administration schedule on the search for minimizing toxicity.