Eliminating elevated p53 signaling fails to rescue skeletal muscle defects or extend survival in lamin A/C-deficient mice.

Lamins A and C, encoded by the LMNA gene, are nuclear intermediate filaments that provide structural support to the nucleus and contribute to chromatin organization and transcriptional regulation. LMNA mutations cause muscular dystrophies, dilated cardiomyopathy, and other diseases. The mechanisms by which many LMNA mutations result in muscle-specific diseases have remained elusive, presenting a major hurdle in the development of effective treatments. Previous studies using striated muscle laminopathy mouse models found that cytoskeletal forces acting on mechanically fragile Lmna-mutant nuclei led to transient nuclear envelope rupture, extensive DNA damage, and activation of DNA damage response (DDR) pathways in skeletal muscle cells in vitro and in vivo. Furthermore, hearts of Lmna mutant mice have elevated activation of the tumor suppressor protein p53, a central regulator of DDR signaling. We hypothesized that elevated p53 activation could present a pathogenic mechanism in striated muscle laminopathies, and that eliminating p53 activation could improve muscle function and survival in laminopathy mouse models. Supporting a pathogenic function of p53 activation in muscle, stabilization of p53 was sufficient to reduce contractility and viability in wild-type muscle cells in vitro. Using three laminopathy models, we found that increased p53 activity in Lmna-mutant muscle cells primarily resulted from mechanically induced damage to the myonuclei, and not from altered transcriptional regulation due to loss of lamin A/C expression. However, global deletion of p53 in a severe muscle laminopathy model did not reduce the disease phenotype or increase survival, indicating that additional drivers of disease must contribute to the disease pathogenesis.

Exploring the Crosstalk Between LMNA and Splicing Machinery Gene Mutations in Dilated Cardiomyopathy.

Mutations in the LMNA gene, which encodes for the nuclear lamina proteins lamins A and C, are responsible for a diverse group of diseases known as laminopathies. One type of laminopathy is Dilated Cardiomyopathy (DCM), a heart muscle disease characterized by dilation of the left ventricle and impaired systolic function, often leading to heart failure and sudden cardiac death. LMNA is the second most commonly mutated gene in DCM. In addition to LMNA, mutations in more than 60 genes have been associated with DCM. The DCM-associated genes encode a variety of proteins including transcription factors, cytoskeletal, Ca2+-regulating, ion-channel, desmosomal, sarcomeric, and nuclear-membrane proteins. Another important category among DCM-causing genes emerged upon the identification of DCM-causing mutations in RNA binding motif protein 20 (RBM20), an alternative splicing factor that is chiefly expressed in the heart. In addition to RBM20, several essential splicing factors were validated, by employing mouse knock out models, to be embryonically lethal due to aberrant cardiogenesis. Furthermore, heart-specific deletion of some of these splicing factors was found to result in aberrant splicing of their targets and DCM development. In addition to splicing alterations, advances in next generation sequencing highlighted the association between splice-site mutations in several genes and DCM. This review summarizes LMNA mutations and splicing alterations in DCM and discusses how the interaction between LMNA and splicing regulators could possibly explain DCM disease mechanisms.

Growth media and assay plate material can impact on the effectiveness of cationic biocides and antibiotics against different bacterial species.

The effectiveness of several cationic disinfectants as well as colistin and polymyxin B were assessed under different growth conditions against Staphylococcus aureus, Escherichia coli and Pseudomonas aeruginosa strains. These conditions included different media (MH1, MH2, TSB and LB) and plate material (polypropylene and polystyrene). Results showed that Minimum inhibitory and bactericidal concentrations (MIC/MBC) values of colistin and polymyxin B were significantly lower on polypropylene plates when compared to polystyrene plates regardless of media used. There were also differences in MIC/MBC values to certain biocides e.g. chlorhexidine and octenidine particularly for S. aureus and E. coli strains, with polypropylene again showing lower values. Other biocides appear to be mostly unaffected by plate type. Whether biocide efficacy was altered by media composition was organism dependent with S. aureus and E. coli more affected than P. aeruginosa. Lower MIC values were more commonly associated with MH2 media and higher MIC values with TSB media for both polypropylene and polystyrene plates, although there were exceptions. Results obtained for standard strains were, in general, indicative for other S. aureus, E. coli and P. aeruginosa strains tested. This study demonstrates the importance of media composition and plate material on biocide effectiveness and highlights the need for optimized disinfectant testing methods. SIGNIFICANCE AND IMPACT OF THE STUDY: There are an increasing number of reports of bacterial strains that are multi-drug resistant. The use of biocides as part of infection control is crucial in helping to combat the spread of these particular strains. Unlike for antibiotics, there are few standardized measuring techniques to understand if an isolate has become more resistant to biocides. This study demonstrates the importance of media composition and plate material on variation and reporting of susceptibility of several bacterial species to specific cationic biocides. It is a useful comparison study to highlight the need to standardize biocide susceptibility testing.

Role of the pro-survival molecule Bfl-1 in melanoma.

Bfl-1 is a pro-survival Bcl-2 family member overexpressed in a subset of chemoresistant tumours, including melanoma. Here, we characterised the expression and regulation of Bfl-1 in normal and malignant melanocytes and determined its role in protecting these cells from chemotherapy-induced apoptosis. Bfl-1 was mitochondrially resident in both resting and apoptotic cells and experienced regulation by the proteasome and NFκB pathways. siRNA-mediated knockdown enhanced sensitivity towards various relevant drug treatments, with forced overexpression of Bfl-1 protective. These findings identify Bfl-1 as a contributor towards therapeutic resistance in melanoma cells and support the use of NFκB inhibitors alongside current treatment strategies.

The effect of the withdrawal of rofecoxib on prescribing patterns of COX-2 inhibitors in Scotland.

BACKGROUND: Concerns have been raised regarding the cardiovascular safety of the COX-2 inhibitors. In September 2004, rofecoxib was withdrawn from the market as a result of concerns regarding its cardiovascular safety. AIMS & METHODS: We set out to examine the effect of the withdrawal of rofecoxib on the prescription of other COX-2 inhibitors and nonselective nonsteroidal anti-inflammatory drugs (nsNSAIDs) in Scotland, using a national prescription database. RESULTS: The withdrawal of rofecoxib led to an initial increase in the prescription of celecoxib as prescribers presumably switched to this alternative agent. However, this rise was short-lived, presumably as a result of concerns that the safety concerning rofecoxib may be a class effect. A parallel increase in the prescription of diclofenac and ibuprofen was also noted, suggesting that prescribers were prescribing these medications as alternatives to COX-2 inhibitors. CONCLUSIONS: While prescribers and their patients may have initially interpreted safety concerns regarding rofecoxib to be drug specific, prescribers appear to have interpreted this effect to be class specific.

Bronchoscopic insertion of Gianturco stents for the palliation of malignant lung disease: 10 year experience.

Malignant large airway obstruction is life threatening and may not be amenable to urgent radiotherapy. Palliative airway stenting is difficult and traditionally carried out under general anaesthesia and fluoroscopy. We have shown that self expanding Gianturco metal stents can be placed under local anaesthesia using fibreoptic bronchoscopy and direct vision for the treatment of malignant airway tumours, and report our 10 year experience. All referrals for stenting referred to our unit between 1990 and 1999 were included, looking for histological type, number and site of stents, complications of the procedure, other interventions, and survival. One hundred and sixty two patients (average age 64 years, (range 21-89)) had 307 stents inserted during 167 procedures (144 primary lung tumours, 18 secondary malignancy). There were no operative deaths, but three patients developed a pneumothorax, one requiring intercostal drain insertion. Average survival following stent insertion was less for primary lung cancer than for secondary disease (103 vs. 431 days, P<0.001). There were no excess complications in a subgroup of 64 patients treated locally by oncologists, even when stenting was the primary procedure. This technique is useful in palliating life threatening airway obstruction, particularly for secondary cancer, and can be used in any centre undertaking fibreoptic bronchoscopy.

Functional characteristics of S-59 photochemically treated platelet concentrates derived from buffy coats.

BACKGROUND: A photochemical treatment (PCT) process for inactivation of infectious pathogens and leukocytes has been developed and evaluated using single-donor platelet concentrates. This study assessed the application of PCT to platelets prepared from pooled buffy coats. In this study, in vitro functional characteristics of PCT platelets were compared to control platelets prepared from pooled buffy coats using the approved platelet-additive solution T-Sol((R)). Platelets in platelet PAS III additive solution without PCT were evaluated as well. PCT also included the use of a psoralen (S-59) reduction device (SRD). MATERIALS AND METHODS: Four types of platelet concentrates were compared: (1) platelet concentrate in plasma/T-Sol; (2) platelet concentrate in plasma/PAS III; (3) platelet concentrate in plasma/PAS III, PCT, 9 h SRD and (4) platelet concentrate in plasma/PAS III, PCT, 16 h SRD. PCT occurred on the day after whole-blood collection. In vitro assay parameters included: pH, pO(2), pCO(2), HCO(-)(3), platelet count, mean platelet volume, plasma glucose, plasma lactate, total ATP, expression of p-selectin, hypotonic shock response and electron microscopy. RESULTS: The results indicate that PCT is compatible with platelet concentrates prepared from pooled buffy coats for up to 7 days of storage. CONCLUSION: The PCT process resulted in acceptable in vitro platelet functional characteristics and is currently in clinical trials to evaluate the haemostatic efficacy of PCT platelets in thrombocytopenic patients requiring multiple platelet transfusions.

Pneumococcal vaccine.

Streptococcus pneumoniae is a frequent cause of pneumonia and meningitis. This article looks at the pneumococcal vaccine, its uses, efficacy, and adverse effects and how vaccination may be improved. We also look at the role of the new conjugate vaccines.

Demonstration of insulin production and storage in insulinomas by in situ hybridization and immunocytochemistry.

Twelve cases of insulinoma were studied to assess the amount of hormone synthesis and hormone storage by the tumour and to see what effect a hormone-producing tumour has on the adjacent normal islets. This was investigated by performing in situ hybridization, which detects hormone messenger RNA, thus giving an indication of the degree of hormone synthesis by the tumour, and immunocytochemistry, which detects the hormone itself, thus giving an indication of the amount of hormone stored in the tumour cells. It was found that in most cases there was less hormone stored within the tumour cells than in adjacent islet cells. In a minority of cases, this decrease in stored hormone was associated with reduced hormone synthesis, but the majority of cases showed either equivalent or increased levels of hormone mRNA in the tumour cells compared with adjacent islets. In addition, it was noted that, unlike some other endocrine organs, the presence of a hormone-producing tumour within the pancreas did not appear to inhibit hormone synthesis in the adjacent normal tissue.

Surgery for lung cancer: age alone is not a contraindication.

A retrospective analysis of the clinical features, operative procedures, postoperative complications and subsequent survival of 70 (50 male) elderly patients undergoing surgery for lung cancer compared with 74 (53 male) younger patients treated at the same hospital during the same period was performed, to determine if elderly people with lung cancer are less likely to benefit from and/or tolerate surgery. The elderly group had to wait longer for operation (p = 0.001) and were more likely to have pre-existing disease (p = 0.019). In contrast, they had fewer recognised postoperative complications (p = 0.032) and there was no difference between the two groups in perioperative mortality and subsequent survival. Surgical treatment of localised lung cancer represents the best chance for cure and this study suggests that age should not be a consideration in the decision to operate or not. The patient's general state of health should be assessed and management decisions based on individual status rather than on age.

Serum indicators of free radical activity in idiopathic pulmonary fibrosis.

Serum levels of free radical activity were measured in 37 patients with idiopathic pulmonary fibrosis (IPF) and 16 control subjects. Three assays used were (1) simultaneously measured levels of the 9,11-diene conjugate of linoleic acid and 9,12-linoleic acid expressed as a percent molar ratio (%MR), a measure of free-radical-mediated lipid peroxidation; (2) thiobarbituric acid reactive substances (TBARS), one of which is malondialdehyde; (3) desferrioxamine-chelatable iron assay, a measure of the potential iron available to catalyze free radical generation. Mean %MR, TBARS and desferrioxamine-chelatable iron were all elevated initially in patients with IPF compared with control subjects (%MR, p < 0.0001; TBARS, p = 0.0013; desferrioxamine-chelatable iron, p = 0.0029). Furthermore, the serum %MR was higher in a subset of patients with clinically worsening IPF than in those patients with clinically stable disease (p = 0.002). Treatment did not appear to affect the three different serum indicators of free radical activity. Thus, lipid peroxidation appears to be increased in patients with IPF and is associated with an increase in desferrioxamine-chelatable iron levels. Serum % MR levels appeared to correlate with clinical disease activity, and they may have a role in monitoring disease activity.

Treatment of large airway obstruction in lung cancer using expandable metal stents inserted under direct vision via the fibreoptic bronchoscope.

BACKGROUND: Self-expanding metal stents have been used successfully to overcome large airway obstruction due to malignant pulmonary disease. The technique has been modified to place stents under direct vision using the fibreoptic bronchoscope. The effect of this procedure on lung function and patient well being was investigated in a large series of patients. METHODS: Fifty six patients with malignant tracheobronchial tumours were treated for symptoms of life threatening airways obstruction or collapse of a lung by the insertion of an expandable metal stent(s) under local anaesthetic using a fibreoptic bronchoscope. All had inoperable cancer and 33 had relapsed after or failed to respond to radiotherapy, chemotherapy, or surgery. Forty seven had primary bronchial carcinomas and nine had metastases from other tumour sites. All but two patients had the stents inserted at one sitting. Measurements were performed in most of the patients before and after stenting and included objective measures (pulmonary function tests, arterial blood gas tensions) and non-objective measures (patient well being, performance status). RESULTS: Overall, 77% of patients showed symptomatic improvement. In those patients in whom measurements were performed two thirds showed improvement in forced expiratory volume in one second (FEV1), forced vital capacity (FVC), peak expiratory flow rate (PEFR) and arterial oxygen tension (PaO2). Medical Research Council dyspnoea score (MRC), Karnofsky, and visual analogue scores (for both breathing and well being) improved in 81% of patients. There were no perioperative deaths. Fifty one patients have died since the procedure with a mean survival of 77 days (range 1-477), and five patients are still alive after a mean of 207 days (range 135-274). CONCLUSIONS: In suitable patients with either extraluminal or intraluminal tumour, or both, the insertion of expandable metal stents using a fibreoptic bronchoscope and local anaesthetic is a valuable addition to other palliative therapies in the treatment of lung cancer.

Counselling for an HIV test.

Doctors may feel uncomfortable with the prospect of discussing a human immunodeficiency virus (HIV) antibody test with their patient. This is in part because they do not enquire about high risk activity as part of a medical history. With increasing medical and public awareness of both the clinical manifestations and social implications of HIV infection, it is important that all doctors receive guidance on how to deal with these issues. Counselling is not the usual term used to describe obtaining informed consent. In the general medical setting, tests for hepatitis B and syphilis are routinely carried out without specific consent even though results of these tests may have profound effects on both the patient and their sexual partners. However society and ethical considerations have made HIV testing different. HIV testing will inevitably become more widespread, and thus become a more routine part of patient investigation and management.

Indicators of free radical activity in patients developing radiation pneumonitis.

PURPOSE: Radiation pneumonitis is thought to occur as the result of excess free radical generation following radiotherapy. Various in vitro studies have shown that large doses of irradiation can cause membrane lipid peroxidation and the oxidation of protein sulphuryl groups. We, therefore, studied two circulating markers of lipid peroxidation and an indicator of "catalytic iron" (potentially available iron to catalyze the generation of free radicals) in patients undergoing radiotherapy. METHODS AND MATERIALS: The 9,11 diene conjugate of 9,12 linoleic acid, expressed as their molar ratio (percentage molar ratio (MR)) and thiobarbituric acid reactive acid-substances (TBARS), as well as levels of circulating desferrioxamine-chelatable iron assay, were assayed. Serial blood samples were taken over a 3-month period in 25 patients with inoperable nonsmall cell lung cancer. RESULTS: Ten patients developed radiation pneumonitis. The patients who developed pneumonitis showed a tendency for the serum percentage molar ratio to increase after a week. The change in the percentage molar ratio between Time 0 and 1 week of radiotherapy was significantly higher in the group that subsequently developed pneumonitis compared to the group that did not (p = 0.002). The initial serum TBARS levels in patients were not significantly elevated compared to controls and there was no difference in the serum TBARS levels in the pneumonitis and nonpneumonitis groups throughout the study period. After 1 week of radiotherapy the group that subsequently developed pneumonitis had a significantly higher level of desferrioxamine-chelatable iron (DFx-iron) compared with the nonpneumonitis group (p = 0.05). CONCLUSION: These data suggest that both the percentage MR and DFx-iron appear to reflect an increased susceptibility to develop radiation pneumonitis and after 1 week of radiotherapy they indicate patients who are likely to subsequently develop pneumonitis. Hence, these indicators could indicate the group of patients that could benefit from intervention therapies with antioxidants.

The use of expandable metal stents for large airway obstruction in older patients.

OBJECTIVE: To review the indications, management, and outcome of 30 older patients who had expandable metal stents inserted for large airway obstruction. DESIGN: Information was collected retrospectively from case notes about presentation, radiographic appearances, pulmonary function, including arterial oxygen tension, and histology. Survival data were collected by reviewing hospital or General Practice records. MEASUREMENTS: Spirometry, peak expiratory flow rate, and blood gases were recorded before and after stent insertion. MAIN RESULTS: There was a significant improvement in the patient's mean forced expiratory volume in 1-second (FEV1) and mean peak expiratory flow rate (PEFR). The arterial oxygen tension (pO2) increased from 8.6 Kpa to 10.6 Kpa. The mean length of survival for the whole group was just under 5 months. CONCLUSIONS: Airway stenting for obstruction provides palliative and functional benefits in these severely disabled patients and a consequent improvement in quality of remaining life.