Assessment of pain threshold in haemophilic patients.

Many patients with haemophilia (PWH) live with persistent end-stage arthritis, as a result of multiple joint haemarthrosis, and experience daily pain. For these people, pain becomes a central aspect of life. The aim of this study was to use mechanical pain thresholds (MPT) to characterize pain perception in different PWH groups. The groups tested were characterized by age, previous bleeding into joints, Hemophilia Joint Health Score (HJHS) and PAIN perception score in the HJHS scoring. A total of 23 PWH (haemophilia A) were included in this study (10 children, 13 adults). A total of 12 PWH suffered from repeated bleeding into some of the tested joints. Data were compared to those collected from 15 age-matched control subjects. The most significant differences in MPTs were found when the PWH were compared to the controls, based on the differences in PAIN score (PAIN score 1 and 2) in all the tested joints, except for the right knee. Similarly, the difference in MPT in ankle joints was confirmed when PWH with and without bleeding were compared to controls. Summarizing the outcomes, we can emphasize the potential usefulness of MPT as an objective tool in evaluating the pain of PWH.

[Recurrent intra-articular bleeding episodes in haemophiliacs. Treatment outcomes in the patients at the university hospital motol in 1985-2005]. / Efekt lécby recidivujícího nitrokloubního krvácení v souboru hemofiliku lécených ve FN v Motole v letech 1985-2005.

PURPOSE OF THE STUDY: Chronic synovitis is a common finding in people with haemophilia. It regularly appears after recurrent episodes of intra-articular bleeding. The bleeding originates from the subsynovial venous plexus underlying the capsule where a lack of thromboplastic activity has been demonstrated. Therefore, the changed synovium appears to be a treatment target. There are several methods which can be used to remove the synovial layer from the joint. The aim of our study was to asses the efficacy of different treatment approaches used in a group of haemophiliacs between 1985 and 2005 in our hospital. MATERIAL AND METHODS: A group of 30 patients with bleeding disorders was evaluated in the study. There were 29 men with haemophilia and one woman with von Wilebrandt factor deficiency. Their age ranged from 6 to 18 (median 13) years. They underwent a total of 68 interventions including surgical synovectomy (n=28), radionuclide synovectomy (n=33) and corticosteroid instillation (n=7). The necessity of a repeat intervention was used as a criterion of successful treatment. RESULTS: In the group of surgical synovectomies, 22% of the patients required repeat operations, in the group of radiation synovectomy, this was 9% and, in the group treated with corticosteroids, this was 43%. The average hospitalisation time was 50 days for surgical procedures (19-133 days) and 7 days for radiation synovectomy procedures (4-13 days). DISCUSSION: In 1994 Merchan presented seven excellent or good results in a group of 10 knees evaluated 1 year after treatment with methylprednisolone. Six years later he reported that "five years after completion of treatment, all results of the observed patients were poor". Generally, corticosteroids will reduce synovitis in the majority of patients but the effect is temporary. A complete remission is a very rare situation under corticosteroid treatment. The experience with surgical synovectomies is not recent and this method is described as carrying a high risk of complications and requiring a high amount of coagulating factor consumption. There are several recent reports on the application of Yttrium-90: in Madrid they evaluated treated joints (knees, ankles and elbows, n = 66) in 44 patients aged from 9 to 39 years. The results were good in less than half of the knees and ankles. The treatment of elbows was more successful. It was recommended to perform synoviorthesis at the early stages of synovitis. In Israel, they reported that a decrease in the number of bleeding episodes was achieved in 80% of 115 patients treated with Yttrium-90; in 15% of them, bleeding in the treated joints stopped completely. In Izmir, Yttrium was used in the treatment of knees, elbows, ankles and also shoulders in children and young adults (3-25 years). The method was found to be safe and effective. Brazilian authors have experience with the treatment of knees, ankles, elbows and shoulders too; they have concluded that this method represents an important resource for the treatment of chronic haemophilic synovitis and markedly reduces joint bleeding frequency and pain, irrespective of the radiographic stage and inhibitor status. While the European Association of Nuclear Medicine (EANM) recommend using 186Re-sulfide for treatment in medium-sized joints, Chinese authors have published a study comparing the effect of using three different doses of 186Re-sulfide in the treatment of chronic synovitis in knees. Their patients have received an amount of radionuclide according to the thickness of their synovial layer measured on MRI, with the result that 22 patients exhibited significant reduction in synovial thickness. A reduction in the number of bleeding episodes was reached in 71% of the patients within an 18-month period. No significant differences were found among the groups receiving different radioactivity doses. In Turkey, 35 elbows, 26 ankles and two shoulders in 49 patients aged between 3 and 30 years were treated with 186Re. The patients were followed up from 6 months to 3 years. At 6 months after the procedure, 81% of the elbows and 86% of the ankles with grade II synovitis were free from bleeding, as well as 53% and 44% of the elbows and ankles with grade III synovitis, respectively. CONCLUSIONS: Radiation synovectomy appears to be the method of choice in the treatment of recurrent bleeding in the joint cavity in people with haemophilia. The efficacy of surgical synovectomy is lower in comparison with radiation synovectomy. Risks associated with surgery and anaesthesia, the need of hospitalisation and a prolonged period of rehabilitation are bothering. On the contrary, the application of corticosteroids cannot be recommended as a good method to treat recurrent haemarthroses.

Methods for detection of circulating tumour cells and their clinical value in cancer patients.

Currently available analytical methods enable identification, detection and characterization of circulating tumour cells in the peripheral blood and disseminated tumour cells in the bone marrow of breast cancer patients. About 0.01 % of the circulating tumour cells observed in the blood are able to form metastases. Therefore, they could be used for estimation of the risk for metastatic relapse, as a diagnostic tool for patient stratification, early determination of the therapy failure, or potential risk of resistance to the given therapeutic intervention. New therapeutic molecular targets could be identified for management of cancer patients using circulating tumour cell detection. The following review summarizes introduced methods of circulating tumour cell detection and their possible application in clinics.

Adiponectin inhibits spontaneous and catecholamine-induced lipolysis in human adipocytes of non-obese subjects through AMPK-dependent mechanisms.

Adiponectin is an adipokine increasing glucose and fatty acid metabolism and improving insulin sensitivity. The aim of this study was to investigate the role of adiponectin in the regulation of adipocyte lipolysis. Human adipocytes isolated from biopsies obtained during surgical operations from 16 non-obese and 17 obese subjects were incubated with 1) human adiponectin (20 microg/ml) or 2) 0.5 mM AICAR - activator of AMPK (adenosine monophosphate activated protein kinase). Following these incubations, isoprenaline was added (10(-6) M) to investigate the influence of adiponectin and AICAR on catecholamine-induced lipolysis. Glycerol concentration was measured as lipolysis marker. We observed that adiponectin suppressed spontaneous lipolysis by 21 % and isoprenaline-induced lipolysis by 14 % in non-obese subjects. These effects were not detectable in obese individuals, but statistically significant differences in the effect of adiponectin between obese and non-obese were not revealed by two way ANOVA test. The inhibitory effect of AICAR and adiponectin on lipolysis was reversed by Compound C. Our results suggest, that adiponectin in physiological concentrations inhibits spontaneous as well as catecholamine-induced lipolysis. This effect might be lower in obese individuals and this regulation seems to involve AMPK.

HLA DRB1, DQB1 and insulin promoter VNTR polymorphisms: interactions and the association with adult-onset diabetes mellitus in Czech patients.

Both the human leucocyte antigen (HLA) DRB1 and the HLA DQB1 gene loci play a role in the development and progression of autoimmune diabetes mellitus (T1DM). Similarly, the insulin promoter variable number tandem repeats (INS-VNTR) polymorphism is also involved in the pathogenesis of diabetes mellitus (DM). We studied the association between each of these polymorphisms and DM diagnosed in patients older than age 35 years. Furthermore, we analysed possible interactions between HLA DRB1/DQB1 and INS-VNTR polymorphisms. Based on C-peptide and GADA levels we were able to distinguish three types of diabetes: T1DM, latent autoimmune diabetes in adults (LADA) and T2DM. INS-VNTR was genotyped indirectly by typing INS-23HphI A/T polymorphism. The genotype and allele frequencies of INS-23HphI did not differ between each of the diabetic groups and group of healthy subjects. We did, however, observe an association between the INS-23HphI alleles, genotypes and C-peptide secretion in all diabetic patients: A allele frequency was 86.2% in the C-peptide-negative group vs. 65.4% in the C-peptide-positive group (P(corr.) < 0.005); AA genotype was found to be 72.4% in the C-peptide-negative group vs. 42.6% in the C-peptide-positive groups (P(corr.) < 0.01). The HLA genotyping revealed a significantly higher frequency of HLA DRB1*03 allele in both T1DM and LADA groups when compared to healthy subjects: T1DM (25.7%) vs. control group (10.15%), odds ratio (OR) = 3.06, P < 0.05; LADA (27.6%) vs. control (10.15%), OR = 3.37, P < 0.01. The simultaneous presence of both HLA DRB1*04 and INS-23HphI AA genotype was detected in 37.5% of the T1DM group compared to only 9.2% of the healthy individuals group (OR = 5.9, P(corr.) < 0.007). We summarize that in the Central Bohemian population of the Czech Republic, the INS-23HphI A allele appears to be associated with a decrease in pancreatic beta cell secretory activity. HLA genotyping points to at least a partial difference in mechanism, which leads to T1DM and LADA development as well as a more diverse genetic predisposition in juvenile- and adult-onset diabetes. The simultaneous effect of HLA and INS-VNTR alleles/genotypes predispose individuals to an increased risk of diabetes development.

KCNJ11 E23K polymorphism and diabetes mellitus with adult onset in Czech patients.

In this work, we studied the association of the E23K polymorphism of the Kir6.2 ATP-sensitive potassium channels in 212 Czech patients with diabetes mellitus who were diagnosed after the age of 35. Patients were classified into T1DM, LADA and T2DM groups based on C-peptide and GADA levels. Carriers of the predisposing Kir6.2 E23K K allele showed no increased risk of either type of diabetes mellitus development. On the other hand, we found a correlation between E23K SNP of the KCNJ11 gene and C-peptide levels, which may be considered a measure of pancreatic beta-cell activity, although this correlation was not statistically significant. In conclusion, we failed to confirm the Kir6.2 E23K as a genetic marker for T1DM, LADA and T2DM in the Central Bohemian population of the Czech Republic.

Aetiological heterogeneity of asymptomatic hyperglycaemia in children and adolescents.

INTRODUCTION: Randomly estimated fasting hyperglycaemia in an asymptomatic individual may represent the first sign of pancreatic beta-cell dysfunction. OBJECTIVE: We aimed at specifying the genetic aetiology of asymptomatic hyperglycaemia in a cohort of children and adolescents. SUBJECTS AND METHODS: We analysed the aetiological diagnosis in 82 non-obese paediatric subjects (38 males) aged 0.2-18.5 years (median: 13.1) who were referred for elucidation of a randomly found blood glucose level above 5.5 mmol/l. In addition to fasting glycaemia and circulating levels of insulin and C-peptide, the subjects were tested by an oral glucose tolerance test and an intravenous glucose tolerance test and screened for mutations in the genes encoding glucokinase (GCK), HNF-1alpha (TCF1), Kir6.2 (KCNJ11) (if aged <2 years) and HNF-4alpha (HNF4A) (those with a positive family history of diabetes). RESULTS AND DISCUSSION: We identified 35 carriers of GCK mutations causing MODY2, two carriers of TCF1 mutations causing MODY3, one carrier of a HNF4A mutation causing MODY1 and one carrier of a KCNJ11 mutation causing permanent neonatal diabetes mellitus. Of the remaining patients, 11 progressed to type 1 diabetes mellitus (T1DM) and 9 had impaired glucose tolerance or diabetes mellitus of unknown origin. In 23 subjects, an impairment of blood glucose levels was not confirmed. We conclude that 39 of 82 paediatric patients (48%) with randomly found fasting hyperglycaemia suffered from single gene defect conditions, MODY2 being the most prevalent. An additional 11 patients (13%) progressed to overt T1DM. The aetiological diagnosis in asymptomatic hyperglycaemic children and adolescents is a clue to introducing an early and effective therapy or, in MODY2, to preventing any future extensive re-investigations.

Association of MHC class I chain related gene-A microsatellite polymorphism with the susceptibility to T1DM and LADA in Czech adult patients.

The results in this study suggest that microsatellite polymorphism within the transmembrane region of MIC-A gene is associated with genetic susceptibility to adult-onset of type 1 diabetes mellitus (T1DM), MIC-A5.1 allele, corrected P = 0.001, whereas it is not associated with latent autoimmune diabetes in adults (LADA) in Czech population. According to our findings, we can hypothesize that adult-onset T1DM and LADA may have partly different immunogenetic aetiopathogenesis.

The frequency of alleles of the Pro12Ala polymorphism in PPARgamma2 is different between healthy controls and patients with type 2 diabetes.

The aim of this initial case-control study was to determine the association between common Pro12Ala polymorphism in the PPARgamma2 gene and type 2 diabetes in the Czech Republic. Furthermore, the effect of this polymorphism on phenotypic characteristics and on levels of lipids (total cholesterol, HDL-cholesterol, LDL-cholesterol and triglycerides) was studied. One hundred thirty-three patients with type 2 diabetes and 97 control subjects were investigated. PCR and RFLP analysis were used for identification of individual genotypes. In the group of patients, three samples (2.26%) were identified as homozygous for the Ala/Ala genotype and 99 samples (74.44%) were homozygotes for the Pro/Pro genotype. Thirty-one samples (23.31%) were identified as Pro12Ala heterozygous. In the control group, six samples (6.19%) were homozygous for the Ala/Ala genotype and 61 samples (62.89%) were homozygotes for the Pro/Pro genotype. Thirty samples (30.93%) were identified as Pro12Ala heterozygous. The allele frequency for the Ala allele was lower in the type 2 diabetic group than in the control group (13.91% vs. 21.43%, P = 0.022). There was no difference (at P < 0.05) between the phenotypic characteristics (BMI, sex) studied in the group of patients according to the Pro12Ala genotype. There was no significant effect of the Pro12Ala polymorphism on lipid levels.

HLA in Czech adult patients with autoimmune diabetes mellitus: comparison with Czech children with type 1 diabetes and patients with type 2 diabetes.

Type 1 diabetes results from an autoimmune insulitis, associated with HLA class II alleles. The evidence about HLA allele association is not clear in patients diagnosed after 35 years of age. In this study we have analyzed HLA alleles of DQB1 and DRB1 genes by sequence specific primer (SSP)-PCR technique in adult patients with disease onset after 35 years of age. Two hundred and eighty-one patients were divided into three groups according to the insulin therapy, the level of C peptide (CP), and GAD antibodies (anti-GAD). Group 1 (type 1 diabetes in adults) was characterized by CP less than 200 pmol/L and anti-GAD more or less than 50 ng/mL (n = 80). All of them had insulin therapy within 6 months after diagnosis. Group 2 latent autoimmune diabetes mellitus in adults (LADA) was defined by a minimum 6-month-long phase after diagnosis without insulin therapy, and was characterized by CP more than 200 pmol/L and anti-GAD more than 50 ng/mL (n = 70). Group 3 (type 2 diabetes) was characterized by CP more than 200 pmol/L and anti-GAD less than 50 ng/mL (n = 131). None ever had insulin therapy. In group 1, there was increased frequency of DRB1*04 (45.0% vs. controls 14.1%, OR = 5.0, P < 0.0005) and DQB1*0302 alleles (43.3% vs. controls 11.1%, OR = 6.1, P < 0.00005). There was increased frequency of DRB1*03 and DQB1*0201, and decreased frequency of DQB1*0602 (3.3% vs. controls 20.2%), but it was not significant. In group 2, there was a significantly increased frequency of DRB1*03 only (50.0% vs. controls 21.2%, OR = 3.7, P < 0.05). Compared with children with type 1 diabetes and adults with type 2 diabetes (group 3), we conclude that the presence of predisposing DQB1 alleles in adults with type 1 diabetes decreases with the age, probably due to environmental factors. Only the DRB1*03, but not the DQB1 gene, becomes the main predisposing allele in LADA patients. These findings suggest that the presence of HLA-DQB1*0302 identifies patients at high risk of requiring insulin treatment. Type 1 diabetes mellitus (DM) in children or adults may have partly different immunogenetic etiopathogenesis than LADA.

[Association of HLA molecules with autoimmune diseases]. / Hypotézy asociácie HLA molekúl s ochoreniami autoimunitného charakteru.

The etiology of autoimmunity diseases includes an immunological reaction against various autoantigens. The HLA molecules play an important role in the antigen presentation process to the immune system. The HLA genes were found as the most significant genetic predisposition factor. The strongest association described in the literature is the association of HLA-B27 with ankylosing spondylitis. Associations were since usually observed with HLA-DR and DQ genes. We are also focused on Diabetes mellitus 1. type. Diabetes is an organ specific autoimmune disease characterised by destruction of beta-cells of Langerhansen pancreatic islets. The cause of the destruction process is not known yet, but it seems to be triggered by repeated presentation of self-antigen hGAD65 to the immune system. This review presents the most interesting solutions and hypotheses of the unanswered question about HLA association mechanism.