Luis Buelta

Inhibition of B cell death causes the development of an IgA nephropathy in (New Zealand white x C57BL/6)F(1)-bcl-2 transgenic mice.

Little is known about the pathogenic mechanisms of IgA nephropathy, despite being the most prevalent form of glomerulonephritis in humans. We report in this study that in (New Zealand White (NZW) × C57BL/6)F1 mice predisposed to autoimmune diseases, the expression of a human bcl-2 (hbcl-2) transgene in B cells promotes a CD4-dependent lupus-like syndrome characterized by IgG and IgA hypergammaglobulinemia, autoantibody production, and the development of a fatal glomerulonephritis. Histopathological analysis of glomerular lesions reveals that the glomerulonephritis observed in these animals resembles that of human IgA nephropathy. The overexpression of Bcl-2 in B cells selectively enhances systemic IgA immune responses to T-dependent Ags. Significantly, serum IgA purified from (NZW × C57BL/6)F1-hbcl-2 transgenic mice, but not from nontransgenic littermates, shows reduced levels of galactosylation and sialylation and an increased ability to deposit in the glomeruli, as observed in human patients with IgA nephropathy. Our results indicate that defects in the regulation of B lymphocyte survival associated with aberrant IgA glycosylation may be critically involved in the pathogenesis of IgA nephropathy, and that (NZW × C57BL/6)F1-hbcl-2 Tg mice provide a new experimental model for this form of glomerulonephritis.

Exacerbation of type II collagen–induced arthritis in apolipoprotein E–deficient mice in association with the expansion of Th1 and Th17 cells

OBJECTIVE To explore the bidirectional relationship between the development of rheumatoid arthritis (RA) and atherosclerosis using bovine type II collagen (CII)-immunized B10.RIII apoE(-/-) mice, a murine model of spontaneous atherosclerosis and collagen-induced arthritis (CIA). METHODS Male B10.RIII apoE(-/-) mice and wild-type controls were immunized with 150 μg of CII emulsified in Freunds complete adjuvant (CFA). The clinical, radiologic, and histopathologic severity of CIA, the levels of circulating IgG1 and IgG2a anti-CII antibodies, the expression of proinflammatory and antiinflammatory cytokines in the joints, and the percentages of Th1, Th17, and Treg lymphocytes in the draining lymph nodes were evaluated during CIA induction. In addition, the size of atherosclerotic lesions was assessed in these mice 8 weeks after CIA induction. RESULTS B10.RIII apoE(-/-) mice that were immunized with CII and CFA developed an exacerbated CIA that was accompanied by increased joint expression of multiple proinflammatory cytokines and by the expansion in the draining lymph nodes of Th1 and Th17 cells. In contrast, the size of vascular lesions in B10.RIII apoE(-/-) mice was not affected by the development of CIA. CONCLUSION Our findings indicate that a deficiency in apolipoprotein E and/or its consequences in cholesterol metabolism act as accelerating factors in autoimmunity by promoting Th1 and Th17 inflammatory responses.

Nuclear morphometry in prognosis of renal adenocarcinoma

Nuclear morphometry was carried out on 95 parenchymatous adenocarcinomas of the kidney treated by radical nephrectomy and hilar lymphadenectomy and followed up for at least five years. The study assessed nuclear area, nuclear perimeter, major diameter, nucleolar area, nuclear shape factor, and nuclear size. There was a significant statistical correlation between survival and the morphometric parameters and between the parameters themselves except for nuclear shape factor. The multiple regression proved that nuclear area is the factor which shows the greatest statistical significance for prognosis. Taking a mean nuclear area of 35 microns 2 allowed two prognostic groups to be established regardless of stage, with those below the threshold having a good prognosis and those above it having a poor prognosis: 96.7 percent of patients with a good prognosis survived after five years (60 months) compared with 17.2 percent of those with a poor prognosis.

Predictors of axillary lymph node metastases in patients with invasive breast carcinoma by a combination of classical and biological prognostic factors.

The presence of axillary lymph node metastases (ALNMs) is the most important prognostic factor in breast carcinoma. If ALNMs were predictable without performing axillary lymph node dissection (ALND), this procedure would not be necessary in selected patients. Using a combination of some of the new biological markers with the classical ones, our objective was I) to identify the best set of predictors of ALNMs, and II) to define predictive models with either high or low probability of ALNMs. We studied 102 patients with invasive breast carcinoma. All patients underwent ALND, and at least 10 axillary lymph nodes per case were obtained. In the primary tumour we evaluated size, histological subtype and grade, lymphatic/vascular invasion and margin. Hormone receptor status, MIB1 index, microvessel density, c-erbB-2 and cathepsin D expression were assessed by immunohistochemistry, and DNA ploidy and S-phase by flow cytometry. Risk factors for ALNMs were estimated by nonlinear logistic regression analysis. The best predictors of ALNMs were: tumour size > 2 cm [OR 6.45, 95% confidence interval (CI) 21.74 to 1.91], presence of lymphatic/vascular invasion [OR 4.95, CI (14.50 to 1.69)], infiltrative margin [OR 9.87 CI (37.44 to 2.60)] and high MIB-1 index [OR 8.39, CI (33.47 to 2.10)]. Two subsets had a very high risk of ALNMs: I) tumour size > 2 cm, with lymphatic/vascular invasion and infiltrative margin; 26 (89.66%) of 29 patients of this subgroup had ALNMs, and (II) tumour size > 2 cm, with lymphatic/vascular and high MIB1 index.; eight of the nine (89%) patients of this subgroup had ALNMs. We could also identify a two-variable model with a very low risk of ALNMs constituted by tumour with circumscribed margin and low MIB-1 index. Of the 19 patients showing these features, only 1 (5.26%) had ALNMs. Therefore, pathological features of the primary tumour can help to assess the risk for ALNM in invasive breast carcinoma. Such risk assessment might avoid regional surgical overtreatment.

CD4+CD25+ T Cell-Dependent Inhibition of Autoimmunity in Transgenic Mice Overexpressing Human Bcl-2 in T Lymphocytes

Regulation of lymphocyte survival is essential for the maintenance of lymphoid homeostasis preventing the development of autoimmune diseases. Recently, we described a systemic lupus erythematosus associated with an IgA nephropathy in autoimmune-prone (NZW × C57BL/6)F1 overexpressing human Bcl-2 (hBcl-2) in B cells (transgenic (Tg) 1). In the present study, we analyze in detail a second line of hBcl-2 Tg mice overexpressing the transgene in all B cells and in a fraction of CD4+ and CD8+ T cells (Tg2). We demonstrate here that the overexpression of hBcl-2 in T cells observed in Tg2 mice is associated with a resistance to the development of lupus disease and collagen type II-induced arthritis in both (NZW × C57BL/6)F1 and (DBA/1 × C57BL/6)F1 Tg2 mice, respectively. The disease-protective effect observed in autoimmune-prone Tg2 mice is accompanied by an increase of peripheral CD4+CD25+ hBcl-2+ regulatory T cells (Tregs), expressing glucocorticoid-induced TNFR, CTLA-4, and FoxP3. Furthermore, the in vivo depletion of CD4+CD25+ Tregs in (DBA/1 × C57BL/6)F1 Tg2 mice promotes the development of a severe collagen type II-induced arthritis. Taken together, our results indicate that the overexpression of hBcl-2 in CD4+ T cells alters the homeostatic mechanisms controlling the number of CD4+CD25+ Tregs resulting in the inhibition of autoimmune diseases.

Glandular Inclusions in Inguinal Hernia Sacs: A Clinicopathological Study of Six Cases

Glandular inclusions in inguinal hernia sacs are not frequent. We present six cases of inguinal hernia with this finding, which represents an incidence of 2.6% in males and shows a predominance in the prepubertal stage. Five patients showed cryptorchidism and two cases were related to congenital malformations of the single umbilical artery type and 47,XY chromosome disorder with chromosomal marker. The most important differential diagnosis must be made with normal histological structures such as the vas deferens or epididymis. The mean diameter of the inclusions was 0.1988 mm and there was a significant difference in size between the inclusions and the vas deferens, but not the epididymis. Differentiation from the latter is based on the absence of a well-developed muscular coat in the wall of the inclusions. It is important to recognize that these inclusions can occur in hernia sacs because of the clinical and medicolegal implications that arise if they are confused with true epididymis or vas deferens. They may arise from paratesticular embryonal remnants.

Prognostic factors in supraglottic laryngeal carcinoma

We carried out a retrospective study of patients with supraglottic carcinomas who were treated surgically at the Marques de Valdecilla Hospital (Santander, Spain) between 1978 and 1987 and who were followed up for at least 5 years. The Kaplan-Meier survival curves were calculated for 24 clinical, histologic, and morphometric parameters. Multivariate analysis was then performed by means of the Cox regression model. In the univariate analysis, survival was related to presence of capsule rupture of the involved lymph nodes (p = 0.00001), number of metastatic lymph nodes (p = 0.0002), postoperative TNM stage (p = 0.004), grade of cell differentiation (p = 0.001), presence of intratumoral necrosis (p = 0.01), and type of invasion (p = 0.04). The nuclear area did not have an influence on survival. Only the presence or absence of capsule rupture of the metastatic lymph nodes and the grade of cell differentiation were included in the final Cox model and proved to be parameters with independent prognostic significance.We carried out a retrospective study of patients with supraglottic carcinomas who were treated surgically at the Marques de Valdecilla Hospital (Santander, Spain) between 1978 and 1987 and who were followed up for at least 5 years. The Kaplan-Meier survival curves were calculated for 24 clinical, histologic, and morphometric parameters. Multivariate analysis was then performed by means of the Cox regression model. In the univariate analysis, survival was related to presence of capsule rupture of the involved lymph nodes (p = 0.00001), number of metastatic lymph nodes (p = 0.0002), postoperative TNM stage (p = 0.004), grade of cell differentiation (p = 0.001), presence of intratumoral necrosis (p = 0.01), and type of invasion (p = 0.04). The nuclear area did not have an influence on survival. Only the presence or absence of capsule rupture of the metastatic lymph nodes and the grade of cell differentiation were included in the final Cox model and proved to be parameters with independent prognostic significance. (Otolaryngol Head Neck Surg 1998;119:548–53.)

p27Kip1 inhibits systemic autoimmunity through the control of Treg cell activity and differentiation

OBJECTIVE Despite the importance of Treg cells in the maintenance of immunologic tolerance, the mechanisms that control their generation and activity are unknown. Since the cell cycle inhibitor p27(Kip1) (p27) was involved in T cell anergy, we undertook this study to explore its role in both Treg cell processes. METHODS The development of type II collagen-induced arthritis (CIA) and lupus-like abnormalities was compared between transgenic mice overexpressing human Bcl-2 in T cells (BCL2-TgT mice) and nontransgenic mice that were deficient or not deficient in p27. The contribution of Treg cells to disease evolution was also explored. Finally, the in vitro activity of Treg cells and their differentiation from naive CD4+ cells was compared between these strains of mice. RESULTS BCL2-TgT mice were protected against CIA by a Treg cell-dependent mechanism. In association with this protection, the overexpression of Bcl-2 in T cells enhanced the differentiation and activity of Treg cells. Both Bcl-2 effects were independent of its antiapoptotic activity but dependent on its capacity to induce the expression of p27 that augmented the strength of transforming growth factor β (TGFβ) signaling in T cells. Accordingly, down-modulation of p27 expression in BCL2-TgT mice promoted CIA. In addition, p27 deficiency in aged C57BL/6 mice reduced the number and activity of Treg cells and induced the development of mild lupus-like abnormalities. CONCLUSION Our results point to p27 as a critical regulator of Treg cell differentiation and function through the positive modulation of TGFβ signaling strength in T cells.

B-cell overexpression of Bcl-2 cooperates with p21 deficiency for the induction of autoimmunity and lymphomas

Genetic abnormalities predisposing to autoimmunity generally act in a cooperative manner affecting one or several mechanisms regulating immunological tolerance. In addition, many of these genetic abnormalities are also involved in the development of lymphoproliferative diseases. In the present study, we have determined the possible cooperation between deficiencies in members of the Cip/Kip family of cell cycle regulators (p21(WAF1/Cip1) or p27(kip1)) and the overexpression of human Bcl-2 in B lymphocytes in the induction of autoimmune and lymphoproliferative diseases in non-autoimmune C57BL/6 (B6) mice. Unlike single mutant mice, B6.p21(-/-) mice transgenic for human Bcl-2 in B cells developed a lethal autoimmune syndrome characterized by the production of autoantibodies, the prominent expansion of memory B and CD4(+) T cells and the development of severe glomerular lesions resembling IgA nephropathy. Furthermore, these mice presented a high incidence of B-cell lymphoproliferative disorders. Such genetic cooperation in the induction of autoimmunity was not observed in B6.p27(-/-) mice transgenic for human Bcl-2 in B cells. Altogether, what we have demonstrated here is the existence of preferential interactions among particular regulators of the G(1)/S transition of the cell cycle and B-cell survival in the induction of systemic autoimmune and lymphoproliferative diseases.

Effects of mycophenolate mofetil in the development of systemic lupus erythematosus in (NZBxNZW)F1 mice

SYSTEMIC LUPUS ERYTHEMATOSUS (SLE) is a chronic and progressive autoimmune syndrome characterized by the production of multiple autoantibodies (autoAb) resulting in the generation of circulating immune complexes (IC). The deposition of these IC leads to the development of tissue lesions such as glomerulonephritis. The study of several strains of mice that spontaneously develop an autoimmune syndrome resembling human SLE have been of considerable value to analyze cellular and molecular mechanisms responsible for the development of the disease as well as to explore the efficiency of different therapies using immunesuppressive drugs. One of these new immunosuppressive agents, Mycophenolate Mofetil (MMF), have been used successfully to prevent SLE in several lupus-prone mice such as MRL.1pr and (NZBxNZW)F1 mice. However, the precise mechanism by which MMF modulates the development of SLE in these animals remains controversial. Whereas in some studies the therapeutic effect of MMF is secondary to the inhibition of autoAb production, in others the administration of MMF blocks the development of IC-mediated glomerulonephritis but not the production of autoAb. To gain insight into the mechanisms responsible for the therapeutic effect of MMF, in the present study (NZBxNZW)F1 female mice have been treated with either high or low doses of MMF from 3 months of age, when increased serum levels of anti-nuclear autoAbs are already present. MATERIAL AND METHODS