Prevalence of FA-D2 Rare Complementation Group of Fanconi Anemia in Serbia

Objective

To investigate genetic subtypes of inherited bone marrow failure syndrome Fanconi anemia (FA) in Sebia. FA-D2 subtype was found to be the most frequent genetic subtype among investigated FA patients; specific observations of FA-D2 phenotype are pointed out.

Methods

Several biological endpoints of FA cells in vitro such as radiation-induced level of lymphocyte micronuclei (radiosensitivity), base line and radiation induced level of the DNA double strand breaks (DSBs), leukocyte apoptosis, and telomere capping function were assessed.

Results

The results indicate that all FA-D2 patients display radioresistant in vitro response, which is seen as significantly reduced yield of radiation-induced micronuclei. On the contrary, FA-A patients display radiosensitive in vitro response seen as increased number of radiation-induced micronuclei (MN). A massive elimination of irradiated cells via apoptosis is found in both FA-A and FA-D2 subtypes. In FA-A subtype apoptosis positively relates with the yield of radiation-induced MN, whereas in FA-D2 subtype apoptosis relates with a high percentage of cells carrying dysfunctional telomeres. The present results unequivocally demonstrate that cytokinesis-block micronucleus (CBMN) assay and analyses of telomere capping function can be used to distinguish FA-D2 and FA-A complementation groups.

Conclusions

Considering all biological endpoints were analyzed, it can be concluded that all FA patients are radiosensitive, regardless of their complementation group. Thus, using CBMN test and telomere capping function analysis can discriminate FA-A from FA-D2 complementation groups, which could be important for assessment the conditioning regimens prior to bone marrow transplantation.     

Postautologous stem cell transplantation engraftment syndrome: Improved treatment and outcomes

Postautologous stem cell transplantation (ASCT) engraftment syndrome (ES) is a well-known clinical complication; however, many aspects remain largely controversial. In this retrospective study, we reviewed records of 156 ASCTs done over 2 years in our institution. Our results show that 45 (34 multiple myeloma/amyloidosis and 11 lymphoma) of 156 adult patients (29%) were diagnosed with ES. Patients with ES were significantly more likely to have fever, rash, and diarrhea upon engraftment. Risk factors for ES included shorter time from diagnosis to ASCT (P = .029), and lower number of pretransplant treatment regimens (0.012). Post-ASCT, patients with ES had significantly lower absolute lymphocyte count on first engraftment day (P = .014). Most ES patients received treatment with steroids. Initial dose of prednisone/methylprednisolone was 2 mg/kg (n = 34), 1 mg/kg (n = 7), while four patients received 1000 mg IV with median length of therapy 7.5 days. One ES patient with inadequate steroid therapy died of ES complications, while another developed respiratory failure requiring intubation but had full recovery with steroids treatment. In conclusion, patients with shorter time from diagnosis to ASCT and with less prior therapy are more likely to develop ES. Overall survival of ES patients has improved with greater awareness of the diagnosis and earlier use of steroids.     

Galectin-3 deficiency enhances type 2 immune cell-mediated myocarditis in mice

Experimental autoimmune myocarditis (EAM) is a mouse model of immune-mediated myocarditis and cardiomyopathy. The role of Galectin-3 (Gal-3), a β-galactoside-binding lectin, in autoimmune myocarditis has not been studied. Therefore, the aim of this study was to delineate the role of Gal-3 in myosin peptide-induced autoimmune myocarditis in mice. EAM was induced in relatively resistant C57BL/6J mice (wild type, WT) and in mice with a targeted deletion of Gal-3 gene (Gal-3KO) by immunization with myosin peptide MyHCα_334–352. Gal-3KO mice developed more severe myocarditis and more pronounced heart hypertrophy than WT mice. Increased infiltration of CD45⁺ leucocytes, CD3⁺ T cells, F4/80⁺ macrophages, and eosinophils was observed in hearts of Gal-3KO mice compared to WT mice on day 21 after EAM induction. Moreover, hearts of Gal-3KO mice had more T helper type 2 (Th2) cells, alternatively activated M2 macrophages, higher amounts of IgG deposits, and higher serum levels of IL-4 and IL-33 than WT mice. Ablation of Gal-3 in Th1-dominant C57BL/6J mice that are relatively resistant to EAM resulted in more severe disease characterized by type 2 cardiac inflammation. The complex effects of Gal-3 on EAM progression might be important in the consideration of therapeutic options for the treatment of EAM.     

Alterations in anti-oxidative defence enzymes in erythrocytes from sporadic amyotrophic lateral sclerosis (SALS) and familial ALS patients.

BACKGROUND: Overproduction of nitric oxide (NO) and hydrogen peroxide (H(2)O(2)) may be an important factor in the pathogenesis of amyotrophic lateral sclerosis (ALS). Owing to their ability to permeate through biological membranes, excess NO and H(2)O(2) may be present in the media surrounding motor neurones. Anti-oxidative defence enzymes (ADEs) in erythrocytes are capable of detoxifying reactive oxygen species (produced endogenously or exogenously), but may also be structurally modified and inactivated by reactive oxygen and nitrogen species. Both balanced and coordinated ADE activities are of utmost importance for their correct physiological function. METHODS: We determined activity of the following ADEs: copper-zinc superoxide dismutase (CuZn SOD), catalase (CAT), glutathione peroxidase (GSH-Px) and glutathione reductase (GR) in erythrocytes from sporadic ALS patients [SALS (-/+)], familial ALS patients with the Leu144Phe mutation in the SOD1 gene [FALS (+/+)], asymptomatic carriers with the Leu144Phe mutation in the SOD1 gene (+/-), and control subjects (-/-). We also examined the in vitro effect of diethyldithiocarbamate (DDC) on CuZn SOD activity in erythrocytes from FALS patients, SALS patients and control subjects. RESULTS: The influence of the Leu144Phe mutation and/or disease was apparent for ADE activities measured in all three patient groups. The SOD1 gene mutation decreased CuZn SOD and GSH-Px activity (two-way ANOVA, significant mutation effect). We noted that the disease also contributed to decreased CuZn SOD activity in SALS patients in comparison with the control group (two-way ANOVA, mutation and disease effect). The disease also influenced CAT and GR activity. CAT activity was decreased in both SALS and FALS patients. In all three patient groups, GR activity was higher than in the control group. Finally, DDC inhibited CuZn SOD activity in erythrocytes from control subjects, FALS (Leu144Phe) patients and SALS patients; however, its effect was more pronounced and significant in FALS patients. CONCLUSIONS: Changes in erythrocyte ADE activities suggest that oxidative stress, involved in the motor neurone pathogenesis of SALS and FALS, also has systemic effects. Differences in ADE systems between the study groups revealed the presence of different types of oxidative pressure, indicating the potential additional benefit of individually designed anti-oxidant cocktail therapies.     

Clinical use of a novel audio pillow with recorded hypnotherapy instructions and music for anxiolysis during dental implant surgery: a prospective study

A prospective, comparative study of a novel audio pillow with hypnosis text and relaxation music was conducted in 82 dental-implant surgery patients to relieve anxiety over a 6-month period. Visual analogue scales combined with the Aachen Dental Treatment Fear Inventory (AZI) questionnaire were used to quantify patients' subjective feelings of fear. Blood pressure, heart rate, and capillary oxygen partial pressure were measured before, during, and after surgery. The AZI scores decreased in the hypnotherapy group (n = 44) and increased slightly in the control group; scores were significantly different between the groups (p = .000). During surgery, the average diastolic blood pressure and heart rate decreased in the hypnotherapy group and increased in controls. Thus, this audio pillow with relaxation music showed anxiolytic effects in patients during dental implantation procedures.     

The NutriChip project - translating technology into nutritional knowledge

Advances in food transformation have dramatically increased the diversity of products on the market and, consequently, exposed consumers to a complex spectrum of bioactive nutrients whose potential risks and benefits have mostly not been confidently demonstrated. Therefore, tools are needed to efficiently screen products for selected physiological properties before they enter the market. NutriChip is an interdisciplinary modular project funded by the Swiss programme Nano-Tera, which groups scientists from several areas of research with the aim of developing analytical strategies that will enable functional screening of foods. The project focuses on postprandial inflammatory stress, which potentially contributes to the development of chronic inflammatory diseases. The first module of the NutriChip project is composed of three in vitro biochemical steps that mimic the digestion process, intestinal absorption, and subsequent modulation of immune cells by the bioavailable nutrients. The second module is a miniaturised form of the first module (gut-on-a-chip) that integrates a microfluidic-based cell co-culture system and super-resolution imaging technologies to provide a physiologically relevant fluid flow environment and allows sensitive real-time analysis of the products screened in vitro. The third module aims at validating the in vitro screening model by assessing the nutritional properties of selected food products in humans. Because of the immunomodulatory properties of milk as well as its amenability to technological transformation, dairy products have been selected as model foods. The NutriChip project reflects the opening of food and nutrition sciences to state-of-the-art technologies, a key step in the translation of transdisciplinary knowledge into nutritional advice.     

Recessive ACTA1 variant causes congenital muscular dystrophy with rigid spine

Variants in ACTA1, which encodes α-skeletal actin, cause several congenital myopathies, most commonly nemaline myopathy. Autosomal recessive variants comprise approximately 10% of ACTA1 myopathy. All recessive variants reported to date have resulted in loss of skeletal α-actin expression from muscle and severe weakness from birth. Targeted next-generation sequencing in two brothers with congenital muscular dystrophy with rigid spine revealed homozygous missense variants in ACTA1. Skeletal α-actin expression was preserved in these patients. This report expands the clinical and histological phenotype of ACTA1 disease to include congenital muscular dystrophy with rigid spine and dystrophic features on muscle biopsy. This represents a new class of recessive ACTA1 variants, which do not abolish protein expression.     

The cost of systemic therapy for metastatic colorectal carcinoma in Slovenia: discrepancy analysis between cost and reimbursement

Background.

The aim of the study was to estimate the direct medical costs of metastatic colorectal cancer (mCRC) treated at the Institute of Oncology Ljubljana and to question the healthcare payment system in Slovenia.

Methods.

Using an internal patient database, the costs of mCRC patients were estimated in 2009 by examining (1) mCRC direct medical related costs, and (2) the cost difference between payment received by Slovenian health insurance and actual mCRC costs. Costs were analysed in the treatment phase of the disease by assessing the direct medical costs of hospital treatment with systemic therapy together with hospital treatment of side effects, without assessing radiotherapy or surgical treatment. Follow-up costs, indirect medical costs, and nonmedical costs were not included.

Results.

A total of 209 mCRC patients met all eligibility criteria. The direct medical costs of mCRC hospitalization with systemic therapy in Slovenia for 2009 were estimated as the cost of medications (cost of systemic therapy + cost of drugs for premedication) + labor cost (the cost of carrying out systemic treatment) + cost of lab tests + cost of imaging tests + KRAS testing cost + cost of hospital treatment due to side effects of mCRC treatment, and amounted to €3,914,697. The difference between the cost paid by health insurance and actual costs, estimated as direct medical costs of hospitalization of mCRC patients treated with systemic therapy at the Institute of Oncology Ljubljana in 2009, was €1,900,757.80.

Conclusions.

The costs paid to the Institute of Oncology Ljubljana by health insurance for treating mCRC with systemic therapy do not match the actual cost of treatment. In fact, the difference between the payment and the actual cost estimated as direct medical costs of hospitalization of mCRC patients treated with systemic therapy at the Institute of Oncology Ljubljana in 2009 was €1,900,757.80. The model Australian Refined Diagnosis Related Groups (AR-DRG) for cost assessment in oncology being currently used is probably one of the reasons for the discrepancy between pay-outs and actual costs. We propose new method for more precise cost assessment in oncology.     

Structure-Activity Relationship Study of Novel Peptoids That Mimic the Structure of Antimicrobial Peptides

The constant emergence of new bacterial strains that resist the effectiveness of marketed antimicrobials has led to an urgent demand for and intensive research on new classes of compounds to combat bacterial infections. Antimicrobial peptoids comprise one group of potential candidates for antimicrobial drug development. The present study highlights a library of 22 cationic amphipathic peptoids designed to target bacteria. All the peptoids share an overall net charge of +4 and are 8 to 9 residues long; however, the hydrophobicity and charge distribution along the abiotic backbone varied, thus allowing an examination of the structure-activity relationship within the library. In addition, the toxicity profiles of all peptoids were assessed in human red blood cells (hRBCs) and HeLa cells, revealing the low toxicity exerted by the majority of the peptoids. The structural optimization also identified two peptoid candidates, 3 and 4, with high selectivity ratios of 4 to 32 and 8 to 64, respectively, and a concentration-dependent bactericidal mode of action against Gram-negative Escherichia coli.