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M Weigert

Publications and source records attributed to M Weigert.

At least 19 recordsLinked to original sources
PubMed2001

Structural basis for autoantibody recognition of phosphatidylserine-beta 2 glycoprotein I and apoptotic cells.

Apoptotic cells contain nuclear autoantigens that may initiate a systemic autoimmune response. To explore the mechanism of antibody binding to apoptotic cells, 3H9, a murine autoantibody with dual specificity for phospholipids and DNA, was used. H chain mutants of 3H9 were constructed, expressed as single-chain Fv (scFv) in Escherichia coli, and assessed for binding to phosphatidylserine, an antigen expressed on apoptotic cells. Both 3H9 and its germline revertant bound to dioleoyl phosphatidylserine in ELISA, and binding was enhanced by beta 2 glycoprotein I (beta 2GPI), a plasma protein that selectively binds to apoptotic cells. Higher relative affinity for DOPS-beta 2GPI was achieved by the introduction of Arg residues into the 3H9 H chain variable region at positions previously shown to mediate DNA binding. Specificity of the two structurally most diverse scFv for apoptotic cells was shown by flow cytometry, and two populations of scFv-bound cells were identified by differences in propidium iodide staining. The results suggest that, in autoimmunity, B cells with Ig receptors for apoptotic cells and DNA are positively selected, and that the antibodies they produce have the potential to affect the clearance and processing of apoptotic cells.

Amino Acid Sequence↗
PubMed2001

Editors and editing of anti-DNA receptors.

Receptor editing is a means by which immature bone marrow B cells can become self-tolerant. Rearrangements of heavy (H) and/or light (L) chain genes are induced by encounter with autoantigens to change the specificity from self to nonself. We have developed site-directed transgenic mice (sd-tg) whose transgenes code for the H chain of antibodies that bind DNA. B cells that express the transgenic H chain associate mainly with four of the 93 functional Vkappa genes of the mouse. Numerous aspartate residues that might inhibit DNA binding by the V(H) domain distinguish these L chain Vkappa sequences, but engaging these Vkappa editors often requires multiple rearrangements. Among the edited B cells is a subset of multispecific cells that express multiple receptors. One consequence of multispecificity is partial autoreactivity; these multispecific B cells may contribute to autoimmunity.

Amino Acid Sequence↗
PubMed2001

Seasonal influences on in vitro fertilization and embryo transfer.

PURPOSE: The study was aimed to investigate the influence of time factors (day, week, month) on pregnancy rates and in vitro fertilization (IVF) and embryo transfer (ET) parameters. METHODS: 8,184 IVF-ET cycles, taking place in two IVF centers from 1992 to 1999, were analyzed. Multiple logistic and linear regression methods were performed as statistical analyses. RESULTS: Oocyte pickup on Tuesdays achieved a significantly higher mean number of oocytes (median: 7) and the highest pregnancy rate (PR) (33.4%) per ET There was a significant variation over the year, with the lowest PR/ET in July (25.71%) and the highest in December (35.5%). Concerning outcomes, the age factor and the number of embryos transferred had the highest significant (p < 0.0001) influence. CONCLUSIONS: The observed weekly rhythm for oocyte pick-ups is certainly due to preprogrammed ovarian stimulation used in our IVF programs. Age as well as the number of embryos transferred are the main influencing factors on a positive outcome and more predictive than seasonal aspects.

Adult↗
PubMed2000

Number and size of antral follicles as predictive factors in vitro fertilization and embryo transfer.

PURPOSE: The purpose of the study was to evaluate whether number and size of antral follicles can predict the outcome of in vitro fertilization-embryo transfer. METHODS: A total of 113 patients were prospectively included into this study. After 19 days of down-regulation, number and size of follicles were determined by using recent three-dimensional transvaginal ultrasound technology. Before application of gonadotropin, all follicles had been defined as antral follicles. According to size, antral follicles were categorized into four different groups: group I included antral follicles < 5 mm, group II follicles 5-10 mm; group III 11-20 mm; and group IV > 20 mm. Pregnant and non-pregnant patients were compared in terms of their number of antral follicles of group I-IV. These four groups were then compared regarding implantation rate, number of retrieved oocytes, endometrium thickness, and age. RESULTS: Pregnant patients showed an significant higher number of follicles with the size between 5 and 10 mm (P = 0.04). A significant correlation was found between number of retrieved oocytes and antral follicle size of 5-10 mm (P = 0.0001). Antral follicles with a diameter between 5 and 10 mm decreased significantly with age (P = 0.008). In group III and IV, a significant correlation was found between antral follicle size (P = 0.016) and serum estradiol level after gonadotropin-releasing hormone-agonist down-regulation (P = 0.011). CONCLUSIONS: We demonstrated that patients with a higher number of follicles between 5 and 10 mm showed a significantly higher pregnancy rate, whereas patients with a dominant number of antral follicles > 11 mm have a higher cancellation rate due to ovarian low response.

Embryo Transfer↗
PubMed2000

[Painfulness of transvaginal follicle aspiration for oocyte retrieval in in-vitro-fertilization].

OBJECTIVE: The aim of this study was to analyze the painfulness of a transvaginal ultrasound-guided follicle punction. MATERIAL AND METHODS: Patients could choose to have either a neuroleptanalgesia or to be supported by the partner or a psychotherapist during the oocyte pick up. Based on a questionnaire, 277 patients tried to describe their level of pain in a horizontal alphanumeric scale divided in 15 categories, while other painful physical interacts could be quantified, too. Furthermore, patients were asked for the cause of sterility, former IVF-ET treatments and whether the partner or the psychotherapist looked after them during the oocyte retrieval. Statistical analysis was performed using a standardized computer program (Stat View, Abacus Concepts, USA). RESULTS: The neuroleptanalgesia reduced the intensity of pain (average value of pain: 1.21 vs 9.26 without analgesia) in a significant way (p = 0.0001). Some patients compared the pain at oocyte retrieval with a bone fracture (8.08) or surgeries (10.12). More painful was a colic (13.67), infections (13.5), some diagnostic surgeries (12.09) or a delivery (11.91). Younger woman and patients with dysmenorrhea felt significantly more pain than others. Neither the presence of the partner or psychotherapist during punction nor the indication, number of previous IVF-ET treatments or a following pregnancy had any influence on the statistics. CONCLUSION: Age as well as the rate of problems with menstruation should help the patient to decide on whether to have the follicle punction with or without anesthesia.

Adult↗
PubMed1999

Somatic mutation and light chain rearrangement generate autoimmunity in anti-single-stranded DNA transgenic MRL/lpr mice.

Antibodies to single-stranded (ss)DNA are expressed in patients with systemic lupus erythematosus and in lupus-prone mouse models such as the MRL/Mp-lpr/lpr (MRL/lpr) strain. In nonautoimmune mice, B cells bearing immunoglobulin site-directed transgenes (sd-tgs) that code for anti-ssDNA are functionally silenced. In MRL/lpr autoimmune mice, the same sd-tgs are expressed in peripheral B cells and these autoantibodies gain the ability to bind other autoantigens such as double-stranded DNA and cell nuclei. These new specificities arise by somatic mutation of the anti-ssDNA sd-tgs and by secondary light chain rearrangement. Thus, B cells that in normal mice are anergic can be activated in MRL/lpr mice, which can lead to the generation of pathologic autoantibodies. In this paper, we provide the first direct evidence for peripheral rearrangement in vivo.

Amino Acid Sequence↗
PubMed1999

The effect of smoking on oocyte quality and hormonal parameters of patients undergoing in vitro fertilization-embryo transfer.

PURPOSE: The aim of the present study was to investigate the influence of smoking on different parameters such as oocyte count, embryo score, and basal hormone values within the scope of in vitro fertilization-embryo transfer (IVF-ET). METHODS: Eight hundred thirty-four women undergoing IVF-ET treatment were classified as smokers or nonsmokers on the basis of questionnaires. Additionally, we divided them into three groups according to their stimulation protocol--"combined stimulation" [I; clomiphene citrate plus human menopausal gonadotropin (hMG)], "ultrashort" [II; gonadotropin releasing hormone agonist (GnRHa) plus hMG or follicle-stimulating hormone (FSH)], and "long downregulation protocol" (III)--and further classified again as smokers or nonsmokers within the groups. RESULTS: In general, smoking patients were significantly (P = 0.0195) younger than nonsmokers and showed a significantly (P = 0.0379) lower embryo score and a tendency (P = 0.0931) to produce fewer oocytes. There was no significant difference concerning the number of normally or pathologically fertilized and transferred oocytes and embryos suitable for cryopreservation. Women who smoked had significantly (P = 0.0112) higher basal 17-beta-estradiol (E2), luteinizing hormone (LH) (P = 0.0001), and dehydroepian-drosteronesulfate (DHEAS) (P = 0.0039) levels, but their basal human prolactin (HPRL) levels were significantly (P = 0.0033) lower than those of nonsmokers. According to the stimulation protocol used, we found the following results. Smoking patients in group I showed a significantly (P = 0.023) lower embryo score and produced fewer oocytes (P = 0.0113), with fewer of them being fertilized (P = 0.0072) and transferred (P = 0.0067). Women who smoked had significantly (P = 0.0002) higher basal LH levels, but their HPRL levels were significantly (P = 0.031) lower than those of nonsmokers. Furthermore, they had a thinner endometrium on the day of embryo transfer (P = 0.0366). In group II we measured significantly elevated basal E2 levels (P = 0.0089) and higher LH values (P = 0.0092) in smokers. Group III showed a trend (P = 0.0565) toward lower HPRL values in smokers. CONCLUSIONS: Although the fertilization rate of oocytes and the pregnancy rate were not significantly different between smokers and nonsmokers, we found significantly alterated hormonal parameters and negatively influenced oocyte parameters, particularly after clomiphene stimulation. So we might consider using only GnRHa protocols for smoking patients. Additionally, we advise our patients to stop smoking before an IVF-ET treatment because of the complex effects of smoking on the reproductive and hormonal system.

Adult↗
PubMed1998

Regulation of anti-DNA B cells in recombination-activating gene-deficient mice.

Anti-DNA antibodies are regulated in normal individuals but are found in high concentration in the serum of systemic lupus erythematosus (SLE) patients and the MRL lpr/lpr mouse model of SLE. We previously studied the regulation of anti-double-stranded (ds)DNA and anti-single-stranded (ss)DNA B cells in a nonautoimmune background by generating mice carrying immunoglobulin transgenes coding for anti-DNAs derived from MRL lpr/lpr. Anti-dsDNA B cells undergo receptor editing, but anti-ssDNA B cells seem to be functionally silenced. Here we have investigated how anti-DNA B cells are regulated in recombination- activating gene (RAG)-2-/- mice. In this setting, anti-dsDNA B cells are eliminated by apoptosis in the bone marrow and anti-ssDNA B cells are partially activated.

Animals↗
PubMed1998

Fixing mismatches.

Explore the source record for details and available documents.

Adenosine Triphosphatases↗
PubMed1997

A solution to the rheumatoid factor paradox: pathologic rheumatoid factors can be tolerized by competition with natural rheumatoid factors.

Rheumatoid factors (RF) associated with arthritic joint erosion are only seen transiently, if at all, in nondiseased individuals. Therefore, a tolerance mechanism must exist that prevents pathologic RF B cells from expressing Abs. Surprisingly, it has been shown that pathologic RF B cells are not tolerized by any previously established tolerance mechanism such as deletion, receptor editing, anergy, or prevention of memory establishment. How are pathologic RF cells tolerized? By simulating the RF response with a cellular automaton model immune system, we demonstrate that pathologic RFs can be tolerized by the novel mechanism of "competitive tolerance" with natural, nonpathologic RFs. We then demonstrate that competitive tolerance can be broken when a sequestered pool of expanding B cells are inappropriately subjected to chronic stimulation (as appears to occur in MRL/lpr mice and in patients with rheumatoid arthritis).

Animals↗
PubMed1997

A Shannon entropy analysis of immunoglobulin and T cell receptor.

In 1970, before any antigen-bound immunoglobulin structure had been solved, Elvin Kabat proposed that regions of high amino acid diversity would be the antigen binding sites of immunoglobulin (Kabat, 1970). Conversely, sites of low variability were proposed to be structural, framework regions. This variability was defined by Wu and Kabat as the number of different amino acids found at a site divided by the relative frequency of the most common amino acid at that site (Wu and Kabat, 1970). Several groups have subsequently devised improvements of Kabat-Wu variability analysis (Litwin and Jores, 1992). While these methods are somewhat better than Kabat-Wu, they still suffer from Kabat-Wu's basic limitation: they account for only the most common one or two amino acids in estimating diversity. This leads to underestimates of low diversities and exaggerations of high diversities. Shannon information analysis eliminates serious bias and is more stable than Kabat-Wu and second generation measures of diversity (Jores et al. 1990; Wu and Kabat, 1970). Statistical reliability can be measured using Shannon analysis, and Shannon measurements can be provided with error estimates. Here we use Shannon's method to analyze the amino acid diversity at each site of T cell receptor Valpha and Vbeta to identify complementarity determining regions and framework sites. Our results reveal that the T cell receptor is significantly more diverse than immunoglobulin-suggesting T cell receptor has more than the previously-discovered four complementarity determining regions. These new complementarity determining regions may represent a larger antigen combining site, additional combining sites, or an evolutionary strategy to avoid inappropriate interaction with other molecules.

Animals↗
PubMed1997

Editing disease-associated autoantibodies.

We have generated site-directed transgenic mice whose transgenes code for anti-DNA antibodies. These antibodies are representative of the lupus-associated anti-DNAs seen in mouse models of autoimmunity and human SLE, and have the usual characteristics of pathogenic autoantibodies. As conventional transgenics in nonautoimmune mice, anti-DNA B cells have been shown to be deleted or inactivated. Autoreactive B cells can also escape negative regulation by a process called receptor editing. Here we describe two combined immunoglobulin H and L chain site-directed transgenic mouse models and characterize their editing phenotypes. One model, 3H9R/Vkappa4R, has a deletion-prone phenotype and undergoes editing, primarily by inactivation of the light chain by leap-frogging events. In the other model, 3H9R/Vkappa8R, B cells are susceptible to anergy and maintain most of their HR and LR chains. These studies clarify the relationship between editing and other mechanisms of tolerance.

Alleles↗
PubMed1995

Constitutive secretion of transgene-encoded IgG2b autoantibodies leads to symptoms of autoimmune disease.

Anti-DNA Ab are strictly regulated, except in autoimmunity, where they are expressed and may contribute to pathogenicity. To study constitutive anti-DNA Ab secretion in nonautoimmune mice, two anti-dsDNA H/L chain transgene combinations were constructed using an IgG2b C region with secretory but no transmembrane domain exons. One H/L combination, consisting of the VH3H9 H and V kappa 4 L chain transgenes, was chosen to recreate 3H9, an autoantibody that originally arose in an autoimmune MRL/lpr mouse; the other paired a higher affinity variant of VH3H9, 56R, with the same V kappa 4 L chain. Elevated titers of IgG2b along with normal levels of other isotypes were observed in transgene-positive mice, indicating that constitutive transgene-directed Ab secretion was achieved. Sera and hybridoma supernatants from VH3H9 gamma transgene-positive animals exhibited binding to dsDNA, ssDNA, and cardiolipin. Mice expressing the 56R gamma H chain and the V kappa 4 L chain showed enhanced binding. Expression of the transgenes correlated with signs of autoimmune disease, including prolonged plasma clotting in vitro, and reduced litter size. The results suggest that even a single autoreactive H chain that escapes tolerance may suffice to induce features of autoimmune disease.

Animals↗
PubMed1995

Light chain contribution to specificity in anti-DNA antibodies.

We studied mice expressing one of two H chain transgenes. Both transgenes expressed the same 3H9 anti-DNA VDJ, but differed in their constant domains. The IgM transgene efficiently induced tolerance and selected for a subset of endogenous L chains that prevented dsDNA binding. In contrast, the IgG2b secreted-only H chain allowed expression of a broad range of L chains, most of which yielded anti-dsDNA Ab. To deduce the features of L chains that affect DNA binding, we derived hybridomas from LPS-stimulated splenic B cells from the two transgene lines and compared the V kappa sequences of Ab they secreted. Identification of L chains with related sequences but different binding to ssDNA, dsDNA, and cardiolipin allowed us to pinpoint L chain residues that correlate with enhanced or reduced binding. Arginines at the junction of V kappa 1 or V kappa 8 regions and J kappa 1, and arginines or asparagines in CDR1 or CDR2 enhanced DNA binding. Negatively charged residues at the same positions were found to interfere with binding. Thus, we predict that appropriate amino acids at these positions may form contacts with DNA. The likely locations of contact residues in the combining site were evaluated by inspection of previously determined Ab structures. Our results indicate that L chains in anti-DNA Ab are able to modulate DNA binding and contribute contact sites for additional determinants on a complex autoantigen.

Amino Acid Sequence↗
PubMed1995

Light chain replacement: a new model for antibody gene rearrangement.

A functional B cell antigen receptor is thought to regulate antibody gene rearrangement either by stopping further rearrangement (exclusion) or by promoting additional rearrangement (editing). We have developed a new model to study the regulation of antibody gene rearrangement. In this model, we used gene targeting to replace the J kappa region with a functional V kappa-J kappa light chain gene. Two different strains of mice were created; one, V kappa 4R, has a V kappa 4-J kappa 4 rearrangement followed by a downstream J kappa 5 segment, while the other, V kappa 8R, has a V kappa 8-J kappa 5 light chain. Here, we analyze the influence of these functional light chains on light chain rearrangement. We show that some V kappa 4R and V kappa 8R B cells only have the V kappa R light chain rearrangement, whereas others undergo additional rearrangements. Additional rearrangement can occur not only at the other kappa allele or isotype (lambda), but also at the targeted locus in both V kappa 4R and V kappa 8R. Rearrangement to the downstream J kappa 5 segment is observed in V kappa 4R, as is deletion of the targeted locus in both V kappa 4R and V kappa 8R. The V kappa R models illustrate that a productively rearranged light chain can either terminate further rearrangement or allow further rearrangement. We attribute the latter to editing of autoantibodies and to corrections of dysfunctional receptors.

Animals↗
PubMed1995

The site and stage of anti-DNA B-cell deletion.

Antibodies to DNA and nucleoproteins are found in sera of individuals with systemic autoimmune disease. In the population (and in the autoimmune mouse strain MRL/lpr) there is a great variety of such antinuclear antibodies, but individuals with systemic lupus erythematosus or single MRL mice express a subset only of the antinuclear specificities found in the population. These observations have been interpreted to mean that these antibodies arise by immunization. The oligoclonal nature of the autoantibody response and the evidence of selection acting on somatically mutated autoantibodies favour this interpretation. Specific activation of autoantibodies in disease implies either that autoantibodies are regulated in non-diseased individuals or that autoantigen availability is variable. The former has been demonstrated in anti-DNA transgenic mice. In normal mice, transgene-encoded antibodies against double-stranded (ds) DNA are not expressed in serum or on B cells. Here we describe modified anti-dsDNA transgenic mice which allow us to study the site and developmental stage at which such B-cell regulation occurs. This model shows that in normal mice B cells expressing anti-DNA specificity are deleted in the bone marrow at a pre-B to immature B transitional stage.

Animals↗