CD4+ T-epitope repertoire on the human acetylcholine receptor alpha subunit in severe myasthenia gravis: a study with synthetic peptides.

The alpha subunit of the nicotinic acetylcholine receptor (AChR) seems crucial in the pathogenesis of the autoimmune paralysis myasthenia gravis (MG) because it contains both the epitopes that dominate the antibody response against the AChR and those recognized by CD4+ AChR-specific T helper (Th) cells. To define the repertoire of anti-AChR Th cells, we investigated the response of unselected blood CD4+ cells or total lymphocytes, or both, from 22 MG patients to 20-residue overlapping synthetic peptides, screening the complete sequence of human-muscle AChR alpha subunit. Several epitopes were identified. Only the most severely affected patients recognized alpha subunit epitopes, and they were mainly young women. Detection of in vitro AChR-specific CD4+ response was facilitated by removal of the CD8+ cells because in two patients a clear response to several alpha subunit peptide sequences could be detected when CD(8+)-depleted cells were used, while their total peripheral blood mononuclear cell population did not respond to any alpha subunit peptide. Although each patient had a unique pattern of peptide recognition, four immunodominant regions recognized by long-term AChR-specific CD4+ T-cell lines, or flanking peptide sequences, were recognized most frequently (residues 48-67, 101-137, 293-337, and 308-437).     

Rat model of distraction osteogenesis

Prior studies of distraction osteogenesis in dog and rabbit models have shown predominantly intramembranous bone formation. Other models of fracture healing normally display mixtures of both endochondral and intramembranous bone formation. We have established a rat model of tibial lengthening that reliably reproduces the pattern of zonal osteogenesis previously observed in dog and rabbit models. A distraction rate of 0.25 mm twice a day with a 0-day latency period produced intramembranous bone with zones of progressive mineralization from collagen. With this protocol, rats bridged the distraction gap with a 25% increase in the tibial bone length. After 20 days of distraction and 50 days of consolidation, the three-point bending stiffness, as a percentage of the contralateral control, reached a level equivalent to that measured in the canine model for a 15% lengthening (28-day distraction and 84-day consolidation). Radiodensitometric analysis of the regenerate bones measured 97% of the unaffected contralateral tibial densities, and mineral analyses demonstrated that calcium and phosphorus levels in the regenerate bone reached 78% of contralateral tibial levels by day 70. We concluded that a rat model of distraction ostegenesis will be useful for a wide range of studies involving rapid intramembranous bone formation.     

Enhancement of fibronectin synthesis and fibrillogenesis by BMP-4 in cultured rat osteoblast.

The skeletal extracellular matrix produced by osteoblasts contains the glycoprotein fibronectin (Fn), which regulates the adhesion, differentiation, and function of osteoblasts. Fn fibrillogenesis is involved in the process of bone mineralization. Bone morphogenetic proteins (BMPs) can be isolated from organic bone matrix and are able to initiate de novo cartilage and bone formation. In this study, the effect of BMP-4 on Fn fibrillogenesis in cultured rat osteoblasts was examined. BMP-4 enhanced Fn synthesis and extracellular Fn assembly in primary cultured osteoblasts. In addition, the extracellular assembly of Fn from exogenously applied soluble human Fn was also increased by BMP-4. It has been reported that alpha5beta1 integrin is related to Fn fibrillogenesis. The synthesis of both alpha5 and beta1 integrins was upregulated by BMP-4. Immunocytochemistry showed that the clustering of alpha5 and beta1 integrins was also increased by BMP-4. BMP-4 increased fibril formation of Fn and the adhesion of osteoblasts onto Fn matrix, which was inhibited by disintegrin triflavin and Gly-Arg-Gly-Asp-Ser (GRGDS) peptide. Phosphorylation of extracellular signal-regulated kinase (ERK) and focal adhesion kinase (FAK) was increased by BMP-4. Enhancement of extracellular Fn fibrillogenesis and the mRNA expression of beta1 integrin by BMP-4 were inhibited by ERK kinase (MEK) inhibitor PD98059. These results suggest that the enhancement of extracellular Fn fibrillogenesis by BMP-4 in cultured osteoblasts is related to the increase of the synthesis of Fn and clustering of alpha5 and beta1 integrins. ERK is involved in the signaling pathway of BMP-4 in regulating Fn fibrillogenesis in osteoblasts.     

Detection of circulating antigen by McAb-AST for evaluating the efficacy of anti-Leishmania chemotherapy.

We have adapted the simple and sensitive McAb-antigen spot test (AST) for evaluating the efficacy of anti-Leishmania chemotherapy. Serum samples from 37 kala-azar patients were tested by McAb-AST, and all showed definite positive reactions before treatment. After a course of antimony treatment, 20 turned negative, coupled with the disappearance of clinical symptoms; another 12 cases responded with weak positivity accompanied by an improvement of clinical manifestations; and the remaining 5 antimony-resistant patients showed strong positive reactions, with their conditions gradually worsening. Furthermore, all 6 cases in which the diagnosis was missed by the bone marrow smear method turned McAb-AST negative after chemotherapy. These results suggest that McAb-AST can be used to evaluate the efficacy of chemotherapy as well as to avoid missed diagnosis by the bone marrow smear method.     

Three years' experience with levonorgestrel-releasing intrauterine device and Norplant-2 implants: a randomized comparative study

A randomized comparative study of levonorgestrel-releasing intrauterine device (LNG-IUD) and Norplant^R-2 implants was carried out in 200 women for 36 months. Three thousand one hundred woman-months of use were observed with each device. Only one pregnancy occurred in users of LNG-IUD at the 12th month of use. The change in bleeding pattern was the most frequent reason for discontinuation. The discontinuation rate for irregular bleeding with Norplant-2 was 17.3 and 26.8 at 24 and 36 months, respectively, as compared to 3.3 with LNG-IUD at both 24 and 36 months. The differences were statistically significant. Removal for amenorrhea and pain only occurred in acceptors of LNG-IUD. About 20–40% of women using Norplant-2 had prolonged bleeding through 36 months. The percentage of amenorrhea in LNG-IUD was the highest (29.3%) at the end of two years of use. More than 97% of subjects reported satisfaction with the methods used by themselves.

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Resumen Un estudio aleatorizado en el que se comparan los dispositivos intrauterinos que descargan levonorgestrel (LNG-IUD) con los implantes Norplant-2 se realizó en 200 mujeres durante 36 meses. Se observó un período de utilización correspondiente a tres mil cien meses-mujer con cada uno de los dispositivos. Hubo un solo embarazo, en el decimosegundo mes, entre las usuarias de LNG-IUD. La alteración del flujo menstrual fue la razón invocada con mayor frecuencia para el abandono del método. Los porcentajes de abandono por la irregularidad de la menstruación en el caso de los Norplant-2 fue de 17,3 y 26,8 a los 24 y 36 meses, respectivamente, en comparación con 3,3 con los LNG-IUD a los 24 y 36 meses. Estas diferencias son estadísticamente significativas. El retiro por amenorrea y dolores sólo ocurrió entre las mujeres que tenían los LNG-IUD. Los Norplant-2 ocasionaron sangrado prolongado durante los 36 meses estudiados en alrededor del 20 al 40% de las mujeres. El porcentaje de amenorrea con los LNG-IUD llegó al nivel máximo (29,3%) al cabo de dos años de uso. Más del 97% de las mujeres se mostraron satisfechas con los métodos que utilizaban.

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Resumé Une étude randomisée comparant les dispositifs intra-utûrins libérant du lévonorgestrel (LNG-IUD) aux implants Norplant-2 a été effectuée sur 200 femmes pendant 36 mois. On a ainsi observé une période d'utilisation correspondant à trois mille cent mois/femme avec chacun des dispositifs. Une seule grossesse s'est produite, au douzième mois, parmi les utilisatrices de LNG-IUD. L'altération du flux menstruel a été la raison le plus souvent invoquée pour l'abandon de la méthode. Les pourcentages d'abandon du fait de l'irrégularité de la menstruation dans le cas des implants se sont élevés à 17,3 et 26,8 après respectivement 24 et 36 mois, alors qu'ils n'atteignaient que 3,3 pour les LNG-IUD, tant à 24 qu'à 36 mois. Ces différences sont statistiquement significatives. Le retrait à la suite d'aménorrhée et de douleurs n'a été constaté que chez des femmes portant les LNG-IUD. Le Norplant-2 a occasionné des saignements prolongés pendant les 36 mois étudiés chez environ 20 à 40% des femmes. Dans le cas des LNG-IUD, le taux d'aménorrhée était le plus élevé (29,3%) au terme de deux années d'utilisation. Plus de 97% des patientes se sont dites satisfaites de la méthode qu'elles appliquaient.

     

Assessment of carbohydrate tolerance in pregnancy

A review is given of the various methods of assessing carbohydrate tolerance in pregnancy. Oral glucose tolerance screening and diagnostic tests have been in use for more than 25 years. They are easily administered, relatively inexpensive, and present reasonable sensitivity; therefore, they continue to be used quite extensively. However, lack of reproducibility of the results and side effects such as nausea, vomiting, and headache have led to the use of alternate methods including glucose polymer (Polycose) and standard breakfast meals. These methods have been reported to present satisfactory results in clinical practice. Glycosylated hemoglobin (HbA1c) and fructosamine assays are also alternate forms of testing carbohydrate metabolism HbA1c measurement have been proven insensitive as a screening test for gestational diabetes, while their use as an index of overall glucose control remains valuable. The role of fructosamine in the assessment of carbohydrate intolerance remains controversial with conflicting claims made by various investigators regarding its sensitivity in detecting gestational diabetes and its response to alterations in glycemic control. In this review, the relative advantages and disadvantages of each glucose tolerance test are discussed and recommendations are given regarding their utility in pregnancy.