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The effect on extracellular matrix content is believed to be an average of several serum derived compounds acting in opposition. The aim of the study is to determine whether whole serum of rats with myocardial infarction may modify the accumulation of extracellular matrix in cultures of myofibroblasts isolated from the myocardial infarction scar. A second aim is to determine whether the tested serum can also degranulate the mast cells. Serum was collected from rats with sham myocardial infarction, rats with myocardial infarction induced by coronary artery ligation and control animals. The experiments were carried out on myocardial infarction scar myofibroblasts or mast cells from the peritoneal cavity. The cultures were divided into three groups containing eight cultures each: one treated with serum from control rats, from animals after sham operation or from those after myocardial infarction. In all groups, the serum was used at concentrations of 10%, 20% or 30%. The total collagen content (Woesner method) glycosaminoglycan level (Farandale method), cell proliferation (BrdU), histamine secretion from mast cells (spectrofluorymetry), β1 integrin and α-smooth muscle actin expression (flow cytometry) were evaluated. Isolated cells were α-smooth muscle actin positive and identified as myofibroblasts. Serum derived from rats with myocardial infarction increased collagen and glycosaminoglycan content in the cultures and modified myofibroblast proliferation in a concentration-dependent manner. The serum also results in an imbalance between collagen and glycosaminoglycan levels. The content of β1 integrin was not influenced by myocardial infarction serum. The serum of rats with myocardial infarction is involved in regulation of collagen and glycosaminoglycan content in myofibroblast cultures, as well as the modification of their proliferation. These changes were not accompanied with integrin β1 density variations. The serum of the myocardial infarction rats did not influence the mast cell degranulation.

Introduction: Melatonin was found to inhibit forskolin-stimulated oxytocin (OT) and vasopressin (VP) release in vitro. The purpose of the present investigation was to evaluate the contribution of the cyclic 3’,5’-adenosine monophosphate/protein kinase A (cAMP/PKA) signalling pathway in melatonin-dependent inhibition of OT and VP secretion from the rat hypothalamo-neurohypophysial (H-NH) system in vitro. Material and methods: The H-NH explants were placed in 1 ml of normal Krebs-Ringer (nK-R) buffer and first preincubated for 30 min in control buffer or in the presence of PKA inhibitor, i.e. cAMPS-Rp or H-89. Next, they were incubated in nK-R buffer {fluid F1} and then in buffer as F1 enriched with melatonin (10–9 M or 10–7 M) and/or PKA activator, i.e. cAMP analogue (8-Br-cAMP), or their vehicles {fluid F2}. After 20 min of incubation in fluid F1 and then F2, the media were collected and frozen, to be assayed for OT and VP by the RIA. Results: 8-Br-cAMP increased OT and VP secretion when the H-NH explants were preincubated in control medium, while PKA inhibitors eliminated its stimulatory effect on OT and VP release. Melatonin (10–7 M) diminished basal OT and VP output from the H-NH system, and inhibited (at both concentrations studied) the cAMP analogue-stimulated release of both neurohormones under control conditions. The effect of melatonin on OT and VP release was completely blocked when cAMPS-Rp, but not H-89, was used to disrupt the cAMP/ /PKA pathway. Conclusions: Melatonin employs the cAMP/PKA signalling pathway to inhibit OT and VP secretion from the rat H-NH system; nonethe­less, other cAMP-mediated mechanisms are not excluded.

Kallmann syndrome type 1 (KS1) is a heterogeneous disorder where hypogonadotropic hypogonadism (HH) associated with an impaired sense of smell is observed. The aim of this study was to investigate the usefulness of the multiplex ligation-dependent probe amplification (MLPA) technique for differential diagnosis in comparison with molecular cytogenetics - fluorescence in situ hybridisation (FISH) or traditional PCR analysis and propose a diagnostic approach for patients with KS.Karyotype and PCR analysis in two related patients and other family members were performed, followed by MLPA dosage sensitive analysis.In the proband and his maternal uncle, the PCR allowed the detection of a large deletion within the KAL1 gene, from exon 4 to 14 (c.469-?_6314+?del). The deletion was also diagnosed in three female carriers in the presented family. These results were proved by the MLPA technique. Moreover, we traced the presence of the region located downstream and upstream to the KAL1 gene on Xq22.32. However, FISH analysis failed to reveal any deletion in the critical region for KS. Simultaneously, we report difficulties connected with the PCR technique based on the primers for KAL1 amplification presented in the literature. We designed primers that are specific to the X chromosome and bypass pseudogene KALY amplification.FISH analysis is a convenient screening technique, but in the presented family it failed to detect the deletion. Therefore, in the face of a distinctive manifestation of KS, a subsequent molecular assay should be introduced. The MLPA is a useful technique for differential diagnosis in patients with HH combined with smell impairment.

A simple method for the determination of 2-thiohydantoins based on the induction of the iodine-azide reaction was developed. The consumption of iodine is proportional to the amount of the inductor. The inducing properties of different compounds (S 2− ) were estimated from the induction coefficient ([I]/[S]) which indicates how many times the consumption of iodine in the induced reaction is higher than that in the iodimetric reaction. Under analytically optimum conditions the induction coefficients were in the range 270–307 for six 2-thiohydantoins and 138–192 for four 3-phenyl-2-thiohydantoins. The applicability of the iodine-azide reaction for the determination of microgram amounts of 2-thiohydantoins after their separation by thin-layer chromatography is demonstrated. The iodine-azide method is sensitive, selective, accurate, cheap and rapid.