Paraffin-embedded tissue as a source of RNA for gene expression analysis in oral malignancy

OBJECTIVE: To assess the feasibility of using archival oral mucosal tissue to examine gene expression at the ribonucleic acid (RNA) level.
MATERIALS AND METHODS: We describe the isolation of RNA from 8 nm sections of formalin-fixed paraffin-embedded oral mucosal tissue. RNA was reverse transcribed and three candidate genes amplified by polymerase chain reaction (PCR). The ribosomal protein S14 gene is a housekeeping gene which has been used as an internal standard in several quantitative PCR protocols. The thymidine kinase (TK) gene is expressed at low levels in most tissues and, with a well-documented genomic organisation, is a useful tool for discrimination between genomic DNA and cDNA. The RIa gene is reported to be overexpressed in many cancer cell lines, in malignant tissue and in vitro transformed cellS. RESULTS: The S14 gene, the TK gene and the RIα gene of the cAMP-dependent protein kinase (PKA) were amplified successfully from formalin-fixed paraffin-embedded tissue sections. The TK primer pair is a useful additional tool in the unambiguous identification of RNA-derived species.
CONCLUSION: RNA suitable for reverse transcribed (RT)-PCR was extracted from archival oral mucosal tissue. This should permit rapid sequence analysis of transcribed tumor suppressor genes and oncogenes in this material. Furthermore, the RT-PCR approach described may allow quantification of gene expression in oral mucosal archival material processed in a standard fashion.     

Mitochondrial function in brain tissue in primary degenerative dementia

Previous in vitro and in vivo studies of the brain in Alzheimer's disease indicated alterations in metabolism related to energy production although the relationships between these changes remains obscure. To help resolve this issue, in vitro oxygen uptake by homogenates of fresh samples of frontal neocortex from patients with dementia and neurosurgical controls has been examined as a measure of energy-related metabolism and mitochondrial function. Maximal respiratory rates (measured in the presence of an uncoupling agent) were similar for samples from 7 controls, 5 patients with Alzheimer's disease and two patients diagnosed clinically as Pick's disease, suggesting that there was little or no effect of these dementias on the maximal metabolic capacity of the tissue. However, under some conditions producing sub-maximal metabolic activity (which are of potentially greater physiological relevance) oxygen uptake rates were significantly elevated in the dementia group. The ratio of oxygen uptake rates in the presence and absence of ADP was significantly reduced (to 58% of control; P less than 0.02) for the dementia patients compared with controls, possibly indicative of partial mitochondrial uncoupling. These results indicate metabolic changes expressed in vitro which may be relevant to the pathogenesis of Alzheimer's disease and some related dementias.     

Diagnostic patterns of regional atrophy on MRI and regional cerebral blood flow change on SPECT in young onset patients with Alzheimer's disease, frontotemporal dementia and vascular dementia

OBJECTIVES: Alzheimer's disease (AD), frontotemporal dementia (FTD) and vascular dementia (VaD) are the three most common causes of young onset dementias. Most neuroimaging studies of these disorders have involved comparisons with normal controls. The aims of this study were to examine the clinical diagnostic value of magnetic resonance imaging (MRI) and single photon emission computed tomography (SPECT) (in combination and in isolation) in the differentiation of one form of dementia from another from amongst a group of AD, FTD and VaD. METHODS: T1 weighted MRI images and 99mTc-HMPAO SPECT images were obtained from consecutive patients with FTD (n=21), AD (n=23) and VaD (n=20) and rated visually by experienced neuroradiologists and nuclear medicine physicians. RESULTS: Asymmetrical atrophy was seen only in FTD. Frontotemporal dementia patients were the most atrophic whereas severe atrophy was rarely observed in VaD. Severe frontal atrophy (unilaterally or bilaterally) and/or asymmetrical atrophy on MRI is highly diagnostic (sensitivity 0.71, specificity 0.93, LR 10.24) of FTD from within a group of FTD and non-FTD (AD, VaD) patients. Mild or severe parietal atrophy with severe reduction in parietal regional cerebral blood flow on SPECT is diagnostic (sensitivity 0.71, specificity 0.76, LR 3.02) of AD from within a group of AD and non-AD (VaD, FTD) patients. CONCLUSION: Anatomical (MRI) and functional (SPECT) imaging provide different information and a combination of these modalities improves diagnostic specificity.     

CVA3 Adherence to Hypertension Therapy: The Effect of Initial Drug Choice

Health-care case management places pressure on decision makers to adopt treatment strategies that promote economic efficiency and hence profitability. Traditional costeffectiveness analysis (CEA), where the objective is to calculate cost-effectiveness ratios, can better inform decision making in markets where prices and efficacy vary widely. However, the threshold at which a given therapy becomes economically efficient relative to competing therapies is not evident from cost-effictiveness ratios alone.
OBJECTIVE: To illustrate the use of spatial techniques for identifying efficient treatment options, using statin therapy in secondary prevention of coronary heart disease (CHD) as a case study.
METHODS: We used a Markov model of CHD epidemiology and treatment to estimate cost-effectivness of 13 statin regimens versus no therapy in secondary prevention of CHD. Comparative efficacy was assessed using data from a recent trial (CURVES) that included these regimens. Patients were assumed to have a history of CHD with risk factors similar to those observed in the trial. CHD event risk was estimated using new subsequent-event risk equations from the Framingham Heart Study. Effectiveness was measured alternatively as gain in life expectancy and CHD events averted.
RESULTS: At usual starting doses, atorvastatin therapy provided the largest life expectancy gain and CHD event avoidance at the lowest cost per life-year gained ($12,900 and $23,400 for men and women, respectively), followed by simvastatin ($17,700 and $31,700), lovastatin ($18,800 and $33,700), pravastatin ($22,600 and $40,200), and fluvastatin ($23,800 and $42,000). Any desired level of effectiveness can be obtained at lowest cost with atorvastatin.
CONCLUSION: Economic efficiency is enhanced when atorvastatin is used to treat some or all patients requiring statin therapy in secondary prevention of CHD.     

Evaluation of the NINCDS-ADRDA criteria in the differentiation of Alzheimer's disease and frontotemporal dementia

OBJECTIVES: The diagnosis of Alzheimer's disease (AD) is now reliant on the use of NINCDS-ADRDA criteria. Other diseases causing dementia are being increasingly recognised--for example, frontotemporal dementia (FTD). Historically, these disorders have not been clearly demarcated from AD. This study assesses the capability of the NINCDS-ADRDA criteria to accurately distinguish AD from FTD in a series of pathologically proved cases. METHODS: The case records of 56 patients (30 with AD, 26 with FTD) who had undergone neuropsychological evaluation, brain imaging, and ultimately postmortem, were assessed in terms of whether at initial diagnosis the NINCDS-ADRDA criteria were successful in diagnosing those patients who had AD and excluding those who did not. RESULTS: (1) The overall sensitivity of the NINCDS-ADRDA criteria in diagnosing "probable" AD from 56 patients with cortical dementia (AD and FTD) was 0.93. However, the specificity was only 0.23; most patients with FTD also fulfilled NINCDS-ADRDA criteria for AD. (2) Cognitive deficits in the realms of orientation and praxis significantly increased the odds of a patient having AD compared with FTD, whereas deficits in problem solving significantly decreased the odds. Neuropsychological impairments in the domains of attention, language, perception, and memory as defined in the NINCDS-ADRDA statement did not contribute to the clinical differentiation of AD and FTD. CONCLUSION: NINCDS-ADRDA criteria fail accurately to differentiate AD from FTD. Suggestions to improve the diagnostic specificity of the current criteria are made.     

Further delineation of the KBG syndrome caused by ANKRD11 aberrations

Loss-of-function variants in ANKRD11 were identified as the cause of KBG syndrome, an autosomal dominant syndrome with specific dental, neurobehavioural, craniofacial and skeletal anomalies. We present the largest cohort of KBG syndrome cases confirmed by ANKRD11 variants reported so far, consisting of 20 patients from 13 families. Sixteen patients were molecularly diagnosed by Sanger sequencing of ANKRD11, one familial case and three sporadic patients were diagnosed through whole-exome sequencing and one patient was identified through genomewide array analysis. All patients were evaluated by a clinical geneticist. Detailed orofacial phenotyping, including orthodontic evaluation, intra-oral photographs and orthopantomograms, was performed in 10 patients and revealed besides the hallmark feature of macrodontia of central upper incisors, several additional dental anomalies as oligodontia, talon cusps and macrodontia of other teeth. Three-dimensional (3D) stereophotogrammetry was performed in 14 patients and 3D analysis of patients compared with controls showed consistent facial dysmorphisms comprising a bulbous nasal tip, upturned nose with a broad base and a round or triangular face. Many patients exhibited neurobehavioural problems, such as autism spectrum disorder or hyperactivity. One-third of patients presented with (conductive) hearing loss. Congenital heart defects, velopharyngeal insufficiency and hip anomalies were less frequent. On the basis of our observations, we recommend cardiac assessment in children and regular hearing tests in all individuals with a molecular diagnosis of KBG syndrome. As ANKRD11 is a relatively common gene in which sequence variants have been identified in individuals with neurodevelopmental disorders, it seems an important contributor to the aetiology of both sporadic and familial cases.     

The effects of a reduced exercise duration taper programme on performance and muscle enzymes of endurance cyclists

Summary The influence of tapering on the metabolic and performance parameters in endurance cyclists was investigated. Cyclists (n = 25) trained 5 days · week^–1, 60 min·day^–1, at 75–85% maximal oxygen consumption (VO_2max) for 8 weeks and were then randomly assigned to a taper group: 4D (4 days;n = 7), 8D (8 days;n = 6), CON (control, 4 days rest;n = 6), NOTAPER (non-taper, continued training;n = 6). Muscle biopsy specimens taken before and after training and tapering were analysed for carnitine palmityltransferase (CPT), citrate synthase, ß-hydroxyacyl CoA dehydrogenase (HOAD), cytochrome oxidase (CYTOX), lactate dehydrogenase, glycogen and protein. Significant increases inVO_2max (6%), a 60-min endurance cycle test (34.5%), oxidative enzymes (77–178%), glycogen (35%) and protein (34%) occurred following training. After the taper, HOAD and CPT decreased 25 % (P<0.05) and 26% respectively, in the CON. Post-taper CYTOX values were different (P<0.05) for 4D and 8D compared with CON. Muscle glycogen levels were increased (P<0.05) after tapering in the 4D, 8D and CON, but decreased in NOTAPER. Similarly, power output at ventilation threshold was significantly increased in the 4D (27.4 W) and 8D (27 W) groups, but decreased (22 W) in the NOTAPER. These findings suggest that tapering elicited a physiological adaptation by altering oxidative enzymes and muscle glycogen levels. Such an adaptation may influence endurance cycling during a laboratory performance test.     

Molecular genetic analysis of autosomal dominant cerebellar ataxia with retinal degeneration (ADCA type II) caused by CAG triplet repeat expansion

Autosomal dominant cerebellar ataxia with retinal degeneration (ADCAII) was previously mapped by linkage analysis studies to chromosome 3p12- p21.1 (SCA7). Positional cloning efforts have recently identified a novel gene, SCA7 , containing a translated CAG repeat, expanded in SCA7 patients. We cloned the SCA7 gene from a yeast artificial chromosome (YAC) clone contig spanning the SCA7 candidate region. Using a combination of genomic sequencing and cosmid-based exon trapping, two expressed sequence tags were identified. Sequencing of the corresponding cDNA clones and RT-PCR analysis identified the full- length SCA7 cDNA. Together, our sequence data defined the intron/exon boundaries of the first two coding exons of the SCA7 gene, with the first exon containing the expanded CAG repeat. Further, sequence comparison with the published SCA7 cDNA identified one additional putative exon in the 5'-UTR region of the SCA7 gene. The SCA7 gene was mapped on the YAC contig in the 2.5 cM interval between D3S1600 and D3S1287. In one extended Belgian SCA7 pedigree the expanded alleles ranged from 38 to at least 55 repeats with allele lengths being inversely correlated with onset age of ADCAII symptoms. The SCA7 repeats increased in length in successive generations. Normal alleles had from four to 18 repeats, with 10 repeats being the most common allele.      

Sensation Seeking, Trait and State Anxiety, and the Electrodermal Orienting Response

The relationship between sensation seeking and the orienting reflex (OR) using skin conductance change is investigated in two experiments. In Experiment I, high sensation seekers gave a greater initial OR In novel visual stimuli while not differing in habituation on subsequent trials. In Experiment II. the paradigm was extended to include auditory as well as visual stimuli. Again, high sensation seekers were found to be more arousable with respect to initial ORs while not differing in habituation rates. The results suggest that sensation seekers may be characterized as having strong excitatory CNS processes. In Experiment II, anxiety (trait and state) was also related to the OR. There were no effects due to trait anxiety but state anxiety did yield significant differences. The more highly anxious (state) subjects had weaker initial ORs than lows in both novel tones, but not to repeated tones. The findings with state anxiety are consistent with findings by others using anxiety neurotics as subjects.