Gamma-irradiated RecD overproducers become permanent recB-/C- phenocopies for extrachromosomal DNA processing due to prolonged titration of RecBCD enzyme on damaged Escherichia coli chromosome

The RecBCD enzyme of Escherichia coli consists of three subunits RecB, RecC and RecD. RecBCD enzyme activities are regulated by its interaction with recombination hotspot Chi. Biochemical and genetic evidence suggest that interaction with Chi affects RecD subunit, and that RecD polypeptide overproduction antagonizes this interaction, suggesting that intact RecD replaces a Chi-modified one. We used bacteria with fragmented chromosomes due to double-strand breaks inflicted by UV and gamma-irradiation to explore in which way increased concentrations of RecBCD's individual subunits affect DNA metabolism. We confirmed that RecD overproduction alters RecBCD-dependent DNA repair and degradation in E. coli. Also, we found that RecB and RecC overproduction did not affect these processes. To determine the basis for the effects of RecD polypeptide overproduction, we monitored activities of RecBCD enzyme on gamma-damaged chromosomal DNA and, in parallel, on lambda and T4 2 phage DNA duplexes provided at intervals. We found that gamma-irradiated wild-type bacteria became transient, and RecD overproducers permanent recB(-)/C(-) phenocopies for processing phage DNA that is provided in parallel. Since this inability of irradiated bacteria to process extrachromosomal DNA substrates coincided in both cases with ongoing degradation of chromosomal DNA, which lasted much longer in RecD overproducers, we were led to conclude that the RecB(-)/C(-) phenotype is acquired as a consequence of RecBCD enzyme titration on damaged chromosomal DNA. This conclusion was corroborated by our observation that no inhibition of RecBCD activity occurs in gamma-irradiated RecBCD overproducers. Together, these results strongly indicate that RecD overproduction prevents dissociation of RecBCD enzyme from DNA substrate and thus increases its processivity.     

Transgene insertion in intronic sequences of Mdga2 gene shows methylation in an imprinted manner in an Acrodysplasia (Adp) mouse line

The Acrodysplasia (Adp) mutation arises from the insertion of a transgene containing a mouse metallothionein-promoted bovine papilloma virus and human growth hormone-releasing factor gene. Although the transgene is not expressed, mice that are hemizygous for the transgene show skull and paw deformities when the progeny receive the transgene paternally. To elucidate the molecular mechanisms underlying the mutant phenotype and the modified transmission pattern of the Adp phenotype, a junctional fragment around the transgene integration site was cloned. The transgene was inserted into the intronic sequences between exon 3 and exon 4 of the Mdga2 gene and the degree of methylation of the transgene and the severity of the phenotype were reciprocally related in that the transgene was highly or under methylated in normal and deformed mice, respectively. Thus, methylation of the transgene appears to regulate phenotypic expression and imprinting of Adp.     

Real-time expression of hTERT in primary melanoma biopsies

Skin melanoma is by far the most lethal skin cancer, it is unpredictable by nature and presents a severe diagnostic problem. One of the major issues in melanoma diagnostics is to differentiate it with confidence from a dysplastic nevus. Thus, the aim of this study was to evaluate hTERT expression on a spectrum of dermal lesions (from normal skin toprimary melanoma) in order to examine its possible role as a diagnostic marker in melanoma diagnostics. In this study we analyzed the expression of hTERT by real-time PCR on 58 freshly obtained biopsy samples (4 samples of normal skin, 12 dermal nevi, 23 dysplastic nevi, 19 primary melanomas). Our results showed slightly greater hTERT expression in dysplastic nevi than melanomas with major data overlap. Considering the given results, hTERT does not seem to be a reliable diagnostic marker for melanoma.     

Quality of life after cardiac surgery

The aim of this study was to determine a health-related quality of life two or more years after a cardiac surgery using the MOS-36 item short-form health survey questionnaire. 330 members of "Croatian society of patients who underwent cardiac surgery" were asked and 196 responders, 122 men and 74 women, were included in survey, all of them more than two years after surgery and older than fifty years of age. The answers were collected by phone. Women gained better results for Physical Functioning (p < 0.003) and men estimated better scores for psychical health (p < 0.001). Our patients gained some different results in respect to the Croatian sample of healthy people of the same age. They declared better General Health (p < 0.001), less Bodily Pain (p < 0.001), but inferior Role-Emotion (p < 0.001) and Social Functioning (p < 0.001). The estimations of QOL improving were comparable with the results of numerous shorter follow-ups.     

WRN mutations in Werner syndrome patients: genomic rearrangements, unusual intronic mutations and ethnic-specific alterations

Werner syndrome (WS) is an autosomal recessive segmental progeroid syndrome caused by null mutations at the WRN locus, which codes for a member of the RecQ family of DNA helicases. Since 1988, the International Registry of Werner syndrome had enrolled 130 molecularly confirmed WS cases from among 110 worldwide pedigrees. We now report 18 new mutations, including two genomic rearrangements, a deep intronic mutation resulting in a novel exon, a splice consensus mutation leading to utilization of the nearby splice site, and two rare missense mutations. We also review evidence for founder mutations among various ethnic/geographic groups. Founder WRN mutations had been previously reported in Japan and Northern Sardinia. Our Registry now suggests characteristic mutations originated in Morocco, Turkey, The Netherlands and elsewhere.     

A case report of an unrecognized nevoid melanoma in a young woman--clinicopathological diagnostic challenge

Nevoid melanoma is a rare form of melanoma histologically resembling benign melanocytic nevi and may be overlooked in routine histological sections. Authors are presenting a case of a 31-year-old woman who presented with bizarre pigmented skin lesions in the area of the postoperative scar on the back where, 6 years earlier, a "nevus pigmentosus epidermo-dermalis" was excised and hystologically confirmed in outer institution. The lesions were surgically removed and histopathological findings were characteristic for nevoid melanoma. Additionally, specimen of primary removed lesion was reexamined and primary nevoid melanoma was then recognized, therefore indicating that the lesions our patient presented with are nevoid melanoma recidivisms. Extensive diagnostic procedures showed no signs of melanoma dissemination. Three months later, the patient returned for consultation and presented with two new brownish-pigmented papules in the area of the new postoperative scar. The lesions were excised and new nevoid melanoma recidivism was confirmed. The patient remained under the regular follow up and, almost 9 years after the removal of primary nevoid melanoma, followed by two cutaneous recidivisms, remains disease-free. This case aims to highlight the problematic area in the analysis of pigmented skin lesions where nevoid melanoma represents one of the clinical and pathological diagnostic challenges.     

Early development of the endemic dace Telestes karsticus (Leuciscidae,Cypriniformes) in a Dinaric karst stream in Croatia

Early development of the endemic dace Telestes karsticus is described in a Dinaric karst stream in Croatia. Larvae were acquired from successful rearing in an aquarium. Basic morphometric and meristic measures of larvae were taken during 55 days of observation. The mean total length of larvae at the time of hatching was 6.45 mm. The yolk sac was completely absorbed in all larvae by day 9 post-hatching (PH), when the mouth also opened and independent feeding and intestinal function were observed. Flexion of the urostyle was completed between days 12–16 PH. The caudal fin became completely forked on day 23 PH. Formation of fin rays was complete on day 48 PH when fish fully resembled juvenile specimens. The number of soft ray fins corresponds to the adult. Understanding the early development is particularly valuable in assessing environmental impacts and recovering of this endangered species and it is essential for effective monitoring and its conservation. This is the first report of the early development for the endemic Telestes karsticus, pointing out the importance of the obtained results for the effective conservation of the species.     

Emerging Law of the Sea Issues in the Antarctic Maritime Area: A Heritage for the New Century?

This article focuses on three emerging law of the sea issues for states cooperating in management of the Antarctic and its maritime area. The first of these is no newcomer: How to regulate the dramatic increase in illegal, unregulated and unreported fishing of Patagonian toothfish (Dissostichus eleginoides) in the Southern Ocean? The second question, according to the letter of the UN Convention on the Law of the Sea, awaits the countries claiming sovereignty over portions of territory in the Antarctic 10 years from the entry into force of the Convention for each of them. The question here is what to do with the requirement contained in that Convention relating to the submission of information on the outer limit of the continental shelf beyond 200 nautical miles to the Commission on the Continental Shelf? Finally, there is a third tricky question: Who is competent to regulate, and accordingly to ban, mineral activities in the Southern Ocean seabed? Is it the International Seabed Authority as the global body, or the Antarctic Treaty Consultative Parties through their regional cooperation? This question may well never be put on the policy agenda for any global forum; but it may well be posed at any time and by any third party, whether in the UN General Assembly or, more likely, in the Assembly of the International Seabed Authority.     

Masticatory muscle and temporomandibular joint pain in Croatian war veterans with posttraumatic stress disorder

The aim of this study was to investigate the prevalence and intensity of masticatory muscle and temporomandibular joint (TMJ) pain in Croatian war veterans with posttraumatic stress disorder (PTSD). The examined group consisted of 100 Croatian war veterans, in whom PTSD had previously been diagnosed. Patients were compared with 92 subjects who had not taken part in the war and in whom PTSD was excluded by psychiatric examination. The clinical examination consisted of palpation of the masticatory muscles, the prominent neck musculature, and TMJ. The examination technique used and the definition of items were previously tested for reliability and validity. 93% of the subjects with PTSD had masticatory muscle tenderness compared to 45.65% of the subjects in the control group (chi2 = 51.46, p < 0.0001). The most frequent painful location in the subjects with PTSD was the left lateral pterygoid site in 88%, and in subjects of the control group the right lateral pterygoid site in 28.26% of cases. The most painful location in the PTSD group was the left lateral pterygoid site in 72%, and in the control group the left posterior digastric in 4.35% of cases. 58% of the subjects with PTSD had TMJ tenderness compared to 3.26% of subjects in the control group (chi2 = 66.23, p < 0.0001). The most frequent painful location of TMJ in both groups was the left posterior capsule; in the PTSD group 38% and in subjects in the control group 2.17% of cases. The most painful location was the left posterior capsule in 28% of subjects with PTSD, while not one subject in the control group reported severe painful sensitivity. The very high frequency and intensity of pain in subjects with PTSD confirms the effect of stress on muscle and joint sensitivity, i.e. perception of pain.     

Spontaneous perforation of the small intestine, a novel manifestation of classical homocystinuria in an adult with new cystathionine beta-synthetase gene mutations

The clinical picture of classical homocystinuria is diverse. This is the first report of an adult homocystinuric patient with non-traumatic spontaneous small bowel perforation. A 47-year old man presented with abdominal rebound tenderness, hypotension and tachycardia, anemia, and elevated markers of inflammation. Other routine laboratory tests were normal. Abdominal x-ray showed no free air. An emergency laparotomy revealed jejunal perforation in the left upper quadrant. Histologic specimen showed full-thickness nonspecific inflammation of the intestinal wall with granulocytic infiltration, hemorrhage and necrosis. Tuberculosis, actinomycosis and typhus were histologically and clinically excluded. After excluding all known possible causes of perforation, we presumed a causative relationship between homocystinuria and small bowel perforation. It could be hypothesized that connective tissue weakness in homocystinuria is a result of homocysteine interference with recombinant human fibrillin-1 fragments or cross-linking of collagen through permanent degradation of disulfide bridges and lysine amino acid residues in proteins. DNA analysis showed three detectable mutations in the cystathionine beta-synthetase gene, 1278T:c.833T>C, and two new mutations, V372G:c.1133T > G, and D520G:c.1558A > G in the aternatively spliced exon 15.