Electron Crescent Distributions as a Manifestation of Diamagnetic Drift in an Electron-Scale Current Sheet: Magnetospheric Multiscale Observations Using New 7.5 ms Fast Plasma Investigation Moments.

We report Magnetospheric Multiscale observations of electron pressure gradient electric fields near a magnetic reconnection diffusion region using a new technique for extracting 7.5 ms electron moments from the Fast Plasma Investigation. We find that the deviation of the perpendicular electron bulk velocity from E × B drift in the interval where the out-of-plane current density is increasing can be explained by the diamagnetic drift. In the interval where the out-of-plane current is transitioning to in-plane current, the electron momentum equation is not satisfied at 7.5 ms resolution.

Genetic basis for expression of the idiotypic network. One unique Ig VH germline gene accounts for the major family of Ab1 and Ab3 (Ab1') antibodies of the GAT system.

Ig germline genes have been isolated from recombinant clones prepared in separate libraries constructed from adult BALB/c liver DNA either in pBR328 plasmid or in EMBL 3 phage. Three clones that gave a very strong positive hybridization signal with a VH anti-GAT-specific probe were completely characterized and sequenced. All three were greater than 95% homologous, with the exception of the 5' noncoding region, which was only 85% homologous but contained characteristic regulatory signals. One of these genes, H10, had a sequence that was completely identical to that of a cDNA derived from a GAT-specific BALB/c hybridoma. Southern blot analysis using Eco RI-digested DNA from rearranged GAT-specific hybridomas revealed that the same gene was used for other GAT-specific VH regions, including one differing from the H10 sequence by 12 nucleotides, which must have been generated by a somatic mechanism. The same H10 germline gene was also used, in most cases without any nucleotide substitution, in hybridomas of the Ab1' set of the GAT idiotypic cascade, suggesting that immunization with Ab2 (antiidiotypic) antibodies preferentially stimulates the direct expression of VH germline genes. Finally, the previous hypothesis that NPa and GAT VH genes were derived from the same germline gene was definitively confirmed, both from sequence data and Southern blot analysis.

Antigen receptor engagement turns off the V(D)J recombination machinery in human tonsil B cells.

The germinal center (GC) is an anatomic compartment found in peripheral lymphoid organs, wherein B cells undergo clonal expansion, somatic mutation, switch recombination, and reactivate immunoglobulin gene V(D)J recombination. As a result of somatic mutation, some GC B cells develop higher affinity antibodies, whereas others suffer mutations that decrease affinity, and still others may become self-reactive. It has been proposed that secondary V(D)J rearrangements in GCs might rescue B cells whose receptors are damaged by somatic mutations. Here we present evidence that mature human tonsil B cells coexpress conventional light chains and recombination associated genes, and that they extinguish recombination activating gene and terminal deoxynucleotidyl transferase expression when their receptors are cross-linked. Thus, the response of the recombinase to receptor engagement in peripheral B cells is the opposite of the response in developing B cells to the same stimulus. These observations suggest that receptor revision is a mechanism for receptor diversification that is turned off when antigen receptors are cross-linked by the cognate antigen.

Quantifying the Effect of Non-Larmor Motion of Electrons on the Pressure Tensor.

In space plasma, various effects of magnetic reconnection and turbulence cause the electron motion to significantly deviate from their Larmor orbits. Collectively these orbits affect the electron velocity distribution function and lead to the appearance of the "non-gyrotropic" elements in the pressure tensor. Quantification of this effect has important applications in space and laboratory plasma, one of which is tracing the electron diffusion region (EDR) of magnetic reconnection in space observations. Three different measures of agyrotropy of pressure tensor have previously been proposed, namely, A∅ e , Dng and Q. The multitude of contradictory measures has caused confusion within the community. We revisit the problem by considering the basic properties an agyrotropy measure should have. We show that A∅ e , Dng and Q are all defined based on the sum of the principle minors (i.e. the rotation invariant I 2) of the pressure tensor. We discuss in detail the problems of I 2-based measures and explain why they may produce ambiguous and biased results. We introduce a new measure AG constructed based on the determinant of the pressure tensor (i.e. the rotation invariant I 3) which does not suffer from the problems of I 2-based measures. We compare AG with other measures in 2 and 3-dimension particle-in-cell magnetic reconnection simulations, and show that AG can effectively trace the EDR of reconnection in both Harris and force-free current sheets. On the other hand, A∅ e does not show prominent peaks in the EDR and part of the separatrix in the force-free reconnection simulations, demonstrating that A∅ e does not measure all the non-gyrotropic effects in this case, and is not suitable for studying magnetic reconnection in more general situations other than Harris sheet reconnection.

Bone marrow cells in X-linked agammaglobulinemia express pre-B-specific genes (lambda-like and V pre-B) and present immunoglobulin V-D-J gene usage strongly biased to a fetal-like repertoire.

Expression of Ig and Ig-related genes has been studied in bone marrow cells from two patients with severe form of X-linked agammaglobulinemia (XLA). Phenotypic analysis revealed the presence of pre-B cells, in the absence of mature B cell markers. The pre-B-specific genes, lambda-like and V pre-B, were normally transcribed. Sequence analysis of 48 distinct V-D-J cDNA clones directly derived from XLA bone marrow cells indicated that they had characteristics of an early fetal pre-B repertoire. All VH families were identified, with a strong bias in the gene usage: a few VH genes were largely overexpressed, either germline or slightly mutated; most genes had been located 3' of the VH locus and were also used in fetal liver (8-13 wk of gestation). Short D regions, (resulting from D-D fusion, making usage of all D genes in both orientations with utilization of the three reading frames), restricted N diversity, and a fetal JH usage pattern were also observed. Taken together, our data suggest that the XLA defect does not alter V-D-J rearrangements nor the expression of mu, lambda-like, and V pre-B transcripts and most likely results in a poor efficiency of some critical steps of the B cell maturation.

A human non-XLA immunodeficiency disease characterized by blockage of B cell development at an early proB cell stage.

We report a detailed analysis of a B cell defect affecting a patient girl born from first cousin parents, characterized by a severe non-X-linked agammaglobulinemia with a total absence of CD19- cells in the periphery. In the bone marrow, CD19 expression was also highly impaired, resulting in the absence of both B and preB compartments. By contrast, CD34+CD10+, CD34psiL+, and some CD19+CD10+ mostly CD34+ early proB cells were present, although diminished. Semiquantitative RT-PCR analysis performed on mononuclear bone marrow cells indicated that lambda-like, VpreB, Rag-1, Rag-2, and TdT transcripts expressed during proB cell stages were found at normal levels whereas E2A, CD10, Syk, Pax-5, CD19, Igalpha, Igbeta, VH-Cmu, and Vkappa-Ckappa transcripts characteristic of later stages were severely depressed. This phenotype resembles that of Pax-5 knock-out mice, but since the coding sequence of the patient Pax-5 cDNA was shown to be normal, the defect might rather result from an altered regulation of this gene. All these data indicate that the patient suffers from a new genetic defect that results in an arrest of differentiation within the proB cell compartment, i.e., earlier than X-linked agammaglobulinemia, before the onset of Ig gene rearrangements.

Immunoglobulin heavy chain expression shapes the B cell receptor repertoire in human B cell development.

Developing B cells must pass a series of checkpoints that are regulated by membrane-bound Ig(mu) through the Igalpha-Igbeta signal transducers. To determine how Ig(mu) expression affects B cell development and Ab selection in humans we analyzed Ig gene rearrangements in pro-B cells from two patients who are unable to produce Ig(mu) proteins. We find that Ig(mu) expression does not affect V(H), D, or J(H) segment usage and is not required for human Igkappa and Iglambda recombination or expression. However, the heavy and light chains found in pro-B cells differed from those in peripheral B cells in that they showed unusually long CDR3s. In addition, the Igkappa repertoire in Ig(mu)-deficient pro-B cells was skewed to downstream Jkappas and upstream Vkappas, consistent with persistent secondary V(D)J rearrangements. Thus, Ig(mu) expression is not required for secondary V(D)J recombination in pro-B cells. However, B cell receptor expression shapes the Ab repertoire in humans and is essential for selection against Ab's with long CDR3s.

Quantitative trait loci analysis of powdery mildew disease resistance in the Arabidopsis thaliana accession kashmir-1.

Powdery mildew diseases are economically important diseases, caused by obligate biotrophic fungi of the Erysiphales. To understand the complex inheritance of resistance to the powdery mildew disease in the model plant Arabidopsis thaliana, quantitative trait loci analysis was performed using a set of recombinant inbred lines derived from a cross between the resistant accession Kashmir-1 and the susceptible accession Columbia glabrous1. We identified and mapped three independent powdery mildew quantitative disease resistance loci, which act additively to confer disease resistance. The locus with the strongest effect on resistance was mapped to a 500-kbp interval on chromosome III.

Fine characterization of childhood and adult acute lymphoblastic leukemia (ALL) by a proB and preB surrogate light chain-specific mAb and a proposal for a new B cell ALL classification.

The expression of the surrogate light chain (psiL) - made of the lambda-like (or lambda5) and the VpreB proteins - is a B cell-specific maturation marker. Using an anti-human VpreB mAb (4G7), we recently identified in human normal bone marrows, proB and preB cells that express the psiH-psiL proB (proBCR) and the mu-psiL preB (preBCR) receptors, respectively. In the present study, FACS and biochemical analysis confirm the broad proB and preB reactivity of the 4G7 mAb that contrasts with the narrow specificity of other available anti-psiL reagents for preB cells. This mAb was used to explore intracytoplasmic and cell surface expression of the VpreB protein on a series of 92 precursor B cell ALLs (from 40 child and 52 adult patients), in combination with 24 other mAbs. The major result concerns the identification within proB (or BI) and common (or BII) ALLs, of proBCR and proBCR+ ALLs that express the VpreB in the cytoplasm or at the cell surface, respectively. The percentage of ALLs within these two VpreB sub-groups differ considerably between the ALL origin. In the pediatric series, ALLs present in the majority a proBCR+ phenotype whereas we observed a proBCR+ phenotype for adult ALLs. Based on VpreB expression, and in combination with other published data, we propose a refined classification for precursor B cell ALLs.

Early occurrence of immunoglobulin isotype switching in human fetal liver.

A cDNA library prepared from a human fetal liver of the first trimester of gestation was screened with Ig C mu, C gamma, C kappa and C lambda probes. Ten heavy chain clones were isolated and characterized by restriction mapping and partial sequencing. The absence of Ig light chain clone and the presence of pre-B-specific lambda-like transcripts suggest that the immune compartment of this cDNA library was mostly derived from pre-B cells. Three transcripts of mu, gamma 2 and gamma 4 isotypes contained a V-D-J-C region with an open reading frame and used members of the VHIV, VHIII and VHI families, respectively. Seven clones were derived from sterile transcripts, one C mu and six C gamma. In addition to C mu exons, the sterile mu transcript contained the 5' flanking germline region. By contrast, the gamma sterile transcripts used a 5' sequence that was spliced from the I gamma 1 region onto the first C gamma 1 exon. In addition several of these transcripts were derived from alternative splicing. The simultaneous expression of both sterile and functional gamma transcripts suggests that the switch mechanism operates in normal fetal liver very early in ontogeny.

The human pre-B cell receptor: structural constraints for a tentative model of the pseudo-light (psi L) chain.

In human pre-B cells, the mu chain is associated with a surrogate light chain composed of the lambda-like and Vpre-B gene products. This pre-B cell receptor presumably triggers early steps of B cell differentiation, We have determined the NH2-terminal amino acid sequence of the lambda-like chain, showing that the mature chain results from the cleavage of a leader segment of 44 residues, leaving a polypeptide of 169 amino acids having partial features of the Ig light chain domains, with the exception of the first 50 amino acid NH2-terminal region. We have completed the nucleotide sequence of the Vpre-B gene, which appears to contain 126 residues in its mature form of which the 24 COOH-terminal portion was not Ig-related. Analysis of transfectants has provided direct evidence that lambda-like and Vpre-B chains assemble together even in the absence of heavy chain, prompting the search for a structural basis of this interaction. Comparison with the domain organization of the regular Ig lambda chain suggests that most of the psi L chain can be accommodated within a CL-VL-like structure, with an extra "subdomain" contributed by the non-Ig-like portions of both the lambda-like and Vpre-B polypeptides.

Monoclonal antibodies reveal three types of idiotypic determinants on MOPC 173: H-L-specific private and public idiotopes, and Vh-specific public idiotope(s).

Monoclonal anti-idiotypic antibodies against the IgG2a, K myeloma protein MOPC 173 were raised with A/J- and CBA-immune B-cells. The specificity of the antibodies was studied by hemagglutination and inhibition of a radioimmunoassay (RIA). A series of myeloma proteins, their H- and L-chains, free or reassociated in various combinations, were used as inhibitors. The RIA patterns allow recognition of three types of idiotypic determinants: private conformational idiotope(s), a public conformational idiotope, and public VH-determinant(s). Strain distribution of the public determinant was studied. A tentative correlation between amino acid positions and Id determinants is discussed.

A non-XLA primary deficiency causes the earliest known defect of B cell differentiation in humans: a comparison with an XLA case.

We report a detailed comparison of B cell defects in two patients, one XLA and one non-XLA. Both had severe agammaglobulinemia with a total absence of CD19+ cells in the periphery. In the non-XLA case, CD19 expression was also highly impaired in the bone marrow, resulting in the absence of both B and preB compartments. Early proB cells were present since CD34+CD10+ and some CD19+CD10+ mostly CD34+ were identified, although diminished. By contrast, in the XLA patient the CD34+CD19+ proB cells were increased whereas the CD34-CD19+ preB cell population was low. Semi-quantitative RT-PCR analysis performed on mononuclear bone marrow cells from the non-XLA patient indicated that lambda-like, VpreB, Rag-1, Rag-2 and TdT transcripts expressed during proB cell stages were found at normal levels whereas E2A, CD10, Syk, Pax-5, CD19, Ig alpha, Ig beta, VH-C mu and V kappa-C kappa transcripts characteristic of later stages were severely depressed. By contrast in the XLA patient most of these transcripts were observed in normal amounts. The phenotype of the non-XLA patient resembles that of Pax-5 or Ig beta knock-out mice, but since the coding sequence of both cDNAs were shown to be normal, the blockage might rather result from an altered regulation of one of these genes or from defect of other genes. All these data indicate that the non-XLA patient suffers from a new genetic defect that results in an arrest of differentiation within the proB cell compartment, before the onset of Ig gene rearrangements. From all agammaglobulinemias reported so far, including XLA cases and those resulting from C mu gene defects, the non-XLA patient exhibits the earliest blockage in the B cell differentiation pathway.

Organization and expression of the pseudo-light chain genes in human B-cell ontogeny.

In pre-B cells, the mu chain is expressed at the cell surface in association with a "light chain surrogate" encoded by the V pre-B and the lambda-like genes. This mu-psi-L complex presumably triggers early steps of the B cell differentiation, possibly by controlling the Ig gene rearrangements. In the humans, the lambda-like complex contains 3 genes, located in the 22q11.2-q12.3 band, telomeric to the IGCL locus, with which they share a similar organization, pointing to a common genetic origin. Only one lambda-like gene, 14.1, is functional and specifically expressed with V pre-B in pre-B cells. This expression starts in cells which still have the IGH locus in germline configuration (pro-B stage) and ceases as soon as the IGL loci rearrange. These pre-B specific transcripts provide useful markers of cells of the B lineage in both physiological and pathological situations.

Fetal versus adult PreB or B cells: the human VH repertoire.

At the preB stage, when only the IGH locus has rearranged, mu chains become expressed in association with the psi L chains, lambda-like and VpreB, thus forming the preB receptor. By the use of a monoclonal anti VpreB antibody, preB cells were isolated from two adult bone marrow samples, and the VH repertoire was analyzed and compared to fetal, XLA (X-linked agammaglobulinemia), and adult B repertoires. Most VH genes identified were also expressed in fetal liver, XLA bone marrow, and adult PBLs, with similar predominant usage of certain germline genes. Multiple D/D fusions, limited N diversity, and preferential use of JH4 with a low level of DQ52 usage were also identified. Few mutations could be observed, not specifically localized in CDR regions, that could be interpreted as not positively selected. Conversely, a shorter length of CDR3 appeared to be the hallmark of the preB step. Thus, the association of psi L chains with mu does not bring about a bias in the VH gene usage, but a first selection on the CDR3 region could be the result of recognition by given autoantigens or ligands different for preB cells and B cells.

Preoperative combination chemotherapy for advanced stage head and neck cancer. Promising early results.

We treated 19 consecutive patients with cisplatin, bleomycin, and methotrexate before definitive surgery or radiation therapy. Fourteen patients (74 percent) had partial or complete tumor regression after chemotherapy. With a minimum follow-up time of 27 months, none of the 4 patients who had a major histologic response relapsed, and only 2 of the remaining 15 patients continued disease-free. The achievement of a complete histologic response after preoperative chemotherapy may correlate with long-term disease-free survival after surgery and radiation therapy for head and neck cancer.

Use of supplemental steroids in patients having orthopaedic operations.

It is commonly thought that patients receiving exogenous glucocorticoids have suppression of the hypothalamic-pituitary-adrenal axis and need high supplemental doses of exogenous glucocorticoids (so-called stress steroids) to meet the demands of operative stress. Several reports have suggested that clinically important suppression of the hypothalamic-pituitary-adrenal axis is extremely uncommon and that the levels of glucocorticoids required for stress are much lower than previously believed. A prospective study of twenty-eight patients who had thirty-five major orthopaedic operations was conducted. No patient received stress steroids; they were given only the baseline immunosuppressive doses of glucocorticoids (mean dose, ten milligrams of prednisone). Clinical information (based on regular physical examinations for signs and symptoms of hypotension, myalgia, arthralgia, ileus, and fever) and laboratory data (serum sodium levels, eosinophil count, and twenty-four-hour urinary free-cortisol levels, determined at perioperative and non-stress postoperative time-periods) were obtained to document any evidence of adrenocortical insufficiency. There was no such evidence in any of the patients, who were monitored during their entire hospitalization. The levels of twenty-four-hour urinary free cortisol showed that all patients had endogenous adrenocortical function and, when this information was considered together with the clinical outcome, it was concluded that this level of function was sufficient to meet the demands of operative stress. Adrenocortical insufficiency in patients who have orthopaedic operations without receiving supplemental stress steroids appears to be much less common than previously thought. While biochemical testing of the function of the hypothalamic-pituitary-adrenal axis may sometimes reveal evidence of adrenal insufficiency, these tests do not predict the clinical outcome and may be too sensitive to guide decisions regarding treatment. Supplemental exogenous stress glucocorticoids may not be needed to meet the demands of operative stress in these patients.

In vitro activation of a nonproductive immunoglobulin allele by a single base pair insertion.

We have shown that one of the alleles in a hybridoma was nonproductive because the sequence in the N region of the heavy chain caused it to be out of frame and to terminate prematurely. This allele became productive, and the gamma 1 heavy chain that it encoded was secreted when an adenosine was inserted to produce an open reading frame. This event occurred at a very low frequency following mutagenesis of the cultured cells. This result suggests that similar sorts of events must occur in vivo when both productive and nonproductive alleles undergo frequent mutations and that new genes may be expressed in hybridomas as they are being subcloned.

[Structural analysis of immunoglobulins and the problem of antibody diversity. New approaches]. / Analyse de la structure des immunoglobulines et problème de la diversité des anticorps. Nouvelles approches.

The function of the various immunoglobulins is revealed by their structure. During humoral response to antigens, the host can modify the structure of its IgG's according to requirements. This is done by genetic processes and also by selection of the appropriate lymphocyte clones.