Formalin Fixation at Low Temperature Better Preserves Nucleic Acid Integrity

Fixation with formalin, a widely adopted procedure to preserve tissue samples, leads to extensive degradation of nucleic acids and thereby compromises procedures like microarray-based gene expression profiling. We hypothesized that RNA fragmentation is caused by activation of RNAses during the interval between formalin penetration and tissue fixation. To prevent RNAse activation, a series of tissue samples were kept under-vacuum at 4°C until fixation and then fixed at 4°C, for 24 hours, in formalin followed by 4 hours in ethanol 95%. This cold-fixation (CF) procedure preserved DNA and RNA, so that RNA segments up to 660 bp were efficiently amplified. Histological and immunohistochemical features were fully comparable with those of standard fixation. Microarray-based gene expression profiles were comparable with those obtained on matched frozen samples for probes hybridizing within 700 bases from the reverse transcription start site. In conclusion, CF preserves tissues and nucleic acids, enabling reliable gene expression profiling of fixed tissues

Cross cultural adaptation of the Achilles tendon Total Rupture Score with reliability, validity and responsiveness evaluation.

PURPOSE: The Achilles tendon Total Rupture Score (ATRS) was developed because of the need for a reliable, valid and sensitive instrument to evaluate symptoms and their effects on physical activity in patients following either conservative or surgical management of an Achilles tendon rupture. Prior to using the score in larger randomized trial in an English-speaking population, we decided to perform reliability, validity and responsiveness evaluations of the English version of the ATRS. Even though the score was published in English, the actual English version has not be validated and compared to the results of the Swedish version. METHODS: From 2009 to 2010, all patients who received treatment for Achilles tendon rupture were followed up using the English version of the ATRS. Patients were asked to complete the score at 3, 6 and 12 months following treatment for Achilles tendon rupture. The ATRS was completed on arrival in the outpatient clinic and again following consultation. RESULTS: The outcomes of 49 (13 female and 36 male) patients were assessed. The mean (SD) age was 49 (12) years, and 27 patients had treatment for a left-sided rupture, 22 the right. All patients received treatment for ruptured Achilles tendons: 38 acute percutaneous repair, 1 open repair, 5 an Achilles tendon reconstruction using a Peroneus Brevis tendon transfer for delayed presentation, 1 gracilis augmented repair for re-rupture and 4 non-operative treatment for mid-portion rupture. The English version of ATRS was shown to have overall excellent reliability (ICC = 0.986). There was no significant difference between the results with the English version and the Swedish version when compared at the 6-month- or 12-month (n.s.) follow-up appointments. The effect size was 0.93. The minimal detectable change was 6.75 points. CONCLUSIONS: The ATRS was culturally adapted to English and shown to be a reliable, valid and responsive method of testing functional outcome following an Achilles tendon rupture

Alcohol consumption and synthesis of ethyl esters of fatty acids in adipose tissue

Ethyl esters of fatty acids (EEFA) have been found to be formed during ethanol metabolism. Human adipose tissue contains high concentrations of free fatty acids, the substrate for EEFA synthesis, and might therefore be a tissue with great potential for EEFA formation. In order to explore their potential usefulness as markers of alcohol abuse, the EEFA concentration and the activity of EEFA-synthesizing enzyme were therefore determined in adipose tissue from men belonging to the following categories: teetotalers, social drinkers, alcoholics under treatment, or established alcoholics found to have died as a result of alcohol intoxication. In order to estimate the half-life of EEFA and the synthase activity induction, the alcoholics were examined after different time periods of abstinence from alcohol. Comparisons were also made with several established markers of alcohol abuse. EEFA were not found in teetotalers, and were found in low concentrations in some of the social drinkers. EEFA were found in several alcoholics, and the forensic cases had high concentrations. EEFA-synthesizing enzyme activity was found in all subjects, increasing from teetotalers to social drinkers, and being 2-fold higher in alcoholics and 5-fold higher in dead alcoholics. The induction of the enzyme after abstinence appeared to have a half-life of the order of several weeks. Correlations were found between EEFA synthase activity and previously established markers of alcohol abuse known to remain for a long time period after abstinence, such as mean erythrocyte corpuscular volume. This preliminary study suggests the possibility that EEFA synthase induction in adipose tissue might have a longer half-life than previously used markers of alcohol abuse. It is therefore suggested that the induction of EEFA synthase might be a potentially useful new marker for alcohol abuse because of its apparent proportionality to alcohol intake over a prolonged time period, its presumed specificity, and long-term elevation after alcohol abstinence. This potential marker should be analysed furthe