Mutations in ATP1A3 cause Alternating Hemiplegia of Childhood (AHC) by disrupting function of the neuronal Na+/K+ ATPase. Published studies to date indicate 2 recurrent mutations, D801N and E815K, ...and a more severe phenotype in the E815K cohort. We performed mutation analysis and retrospective genotype-phenotype correlations in all eligible patients with AHC enrolled in the US AHC Foundation registry from 1997-2012. Clinical data were abstracted from standardized caregivers' questionnaires and medical records and confirmed by expert clinicians. We identified ATP1A3 mutations by Sanger and whole genome sequencing, and compared phenotypes within and between 4 groups of subjects, those with D801N, E815K, other ATP1A3 or no ATP1A3 mutations. We identified heterozygous ATP1A3 mutations in 154 of 187 (82%) AHC patients. Of 34 unique mutations, 31 (91%) are missense, and 16 (47%) had not been previously reported. Concordant with prior studies, more than 2/3 of all mutations are clusteredin exons 17 and 18. Of 143 simplex occurrences, 58 had D801N (40%), 38 had E815K(26%) and 11 had G947R (8%) mutations corrected.Patients with an E815K mutation demonstrate an earlier age of onset, more severe motor impairment and a higher prevalence of status epilepticus. This study further expands the number and spectrum of ATP1A3 mutations associated with AHC and confirms a more deleterious effect of the E815K mutation on selected neurologic outcomes. However, the complexity of the disorder and the extensive phenotypic variability among subgroups merits caution and emphasizes the need for further studies.
Circadian systems provide a fitness advantage to organisms by allowing them to adapt to daily changes of environmental cues, such as light/dark cycles. The molecular mechanism underlying the ...circadian clock has been well characterized. However, how internal circadian clocks are entrained with regular daily light/dark cycles remains unclear. By collecting and analyzing indirect calorimetry (IC) data from more than 2000 wild-type mice available from the International Mouse Phenotyping Consortium (IMPC), we show that the onset time and peak phase of activity and food intake rhythms are reliable parameters for screening defects of circadian misalignment. We developed a machine learning algorithm to quantify these two parameters in our misalignment screen (SyncScreener) with existing datasets and used it to screen 750 mutant mouse lines from five IMPC phenotyping centres. Mutants of five genes (Slc7a11, Rhbdl1, Spop, Ctc1 and Oxtr) were found to be associated with altered patterns of activity or food intake. By further studying the Slc7a11tm1a/tm1a mice, we confirmed its advanced activity phase phenotype in response to a simulated jetlag and skeleton photoperiod stimuli. Disruption of Slc7a11 affected the intercellular communication in the suprachiasmatic nucleus, suggesting a defect in synchronization of clock neurons. Our study has established a systematic phenotype analysis approach that can be used to uncover the mechanism of circadian entrainment in mice.
Formalin-fixed and paraffin-embedded lymph node biopsy specimens from 52 untreated patients with newly diagnosed diffuse large cell (n = 48) or mixed cell (n = 4) non-Hodgkin's lymphoma (NHL) were ...analyzed for DNA content and proliferative activity (PA) by flow cytometry. The results obtained by flow cytometry were compared with the results of cytogenetic studies performed on 28 of the specimens. The median age of the patients was 65 years (range, 15-84 years) and the male to female ratio was 3 to 2. All patients were uniformly staged and uniformly treated with cyclophosphamide, doxorubicin, procarbazine, bleomycin, vincristine, and prednisone. The flow cytometric results were compared statistically by univariate analysis with the rate and duration of complete remission and survival. Tumors with low PA (greater than or equal to 80% of cells in G0/G1 phase) were found in 65% of the patients; 74% of those with low PA versus only 44% of those with high PA achieved an initial complete remission (P less than 0.02). DNA aneuploidy was detected in tumors of 56% of the patients and was associated with a significantly longer duration of complete remission (P less than 0.01). Both low PA and aneuploidy independently predicted longer survival. The predicted 2-year actuarial survival for patients with tumors with low PA was 68% versus 10% for those with high PA (P less than 0.01). Similarly, the 2-year survival of patients with aneuploid tumors was 60% versus 36% for those with diploid tumors (P less than 0.01). The combination of PA and DNA content categorized the patients into four groups with decreasing 2-year survivals: low PA/aneuploid (n = 20), 77%; low PA/diploid (n = 14), 57%; high PA/aneuploid (n = 9), 32%; high PA/diploid (n = 9), 0%. The flow cytometric results correlated well with those of the cytogenetic studies. We conclude that low PA and DNA aneuploidy, both separately and in combination, predict a favorable clinical outcome for patients with diffuse mixed cell and large cell NHL.
Specific cytogenetic changes such as t(14;18) and t(8;14) are associated with specific histologic subtypes of non-Hodgkin's lymphoma (NHL) and may predict disease outcome. Nonspecific cytogenetic ...changes include other structural rearrangements or numerical changes such as monosomies and trisomies, which may cause changes in total cellular DNA content. In many solid tumors, the presence of abnormal DNA content may be predictive of clinical behavior. NHL biopsies, however, contain normal (diploid) as well as abnormal cells, and DNA changes in the peridiploid range are detectable by cytogenetic analysis, but not consistently by flow cytometry. In the present study, we performed flow cytometric and cytogenetic analysis of DNA on biopsies from 129 patients with non-Hodgkin's lymphoma (NHL). Cytogenetic studies were successful on 88 (68%) of the samples. There was 55% concordance between flow cytometric and cytogenetic techniques in detecting aneuploid DNA content, with the majority of discrepancies occurring in the peridiploid range. We also detected six samples which were aneuploid by flow cytometry, but diploid by cytogenetics. We suggest that a reasonable approach to determine DNA content, as it relates to prediction of outcome in NHL, would be to combine data from both of these techniques and analyze the results in terms of ranges of DNA rather than by categorizing as diploid versus aneuploid.
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