Deletion of the Caenorhabditis elegans homologues of the CLN3 gene, involved in human juvenile neuronal ceroid lipofuscinosis, causes a mild progeric phenotype

Summary The CLN3 gene is involved in juvenile neuronal ceroid lipofuscinosis (JNCL), or Batten–Spielmeyer–Vogt disease, a severe hereditary neurodegenerative lysosomal storage disorder characterized by progressive disease pathology, with loss of vision as the first symptom. Another characteristic of JNCL is the lysosomal accumulation of autofluorescent lipopigments, forming fingerprint storage patterns visible by electron microscopy. The function of the CLN3 protein is still unknown, although the evolutionarily conserved CLN3 protein is being functionally analysed using different experimental models. We have explored the potential of the nematode Caenorhabditis elegans as a model for Batten disease in order to bridge the gap between the unicellular yeast and very complex mouse JNCL models. C. elegans has three genes homologous to CLN3, for each of which deletion mutants were isolated. Cln-3.1 deletion mutants have a decreased lifespan, and cln-3.2 deletion mutants a decreased brood size. However, the neuronal or movement defects and aberrant lipopigment distribution or accumulation observed in JNCL were not found in the worms. To detect possible redundancy, single deletion mutants were crossed to obtain double and triple mutants, which were viable but showed no JNCL-specific defects. The cln-3 triple mutants show a more prominent decrease in lifespan and brood size, the latter most conspicuously at the end of the egg-laying period, suggesting premature ageing. To focus our functional analysis we examined the C. elegans cln-3 expression patterns, using promoter–GFP (green fluorescent protein) gene fusions. Fluorescence patterns suggest cln-3.1 expression in the intestine, cln-3.2 expression in the hypoderm, and cln-3.3 expression in intestinal muscle, male-specific posterior muscle and hypoderm. Further life stage- and tissue-specific analysis of the processes causing the phenotype of the cln-3 triple mutants may provide more information about the function of the cln-3 protein and contribute to a better understanding of the basic processes affected in Batten disease patients. Electronic supplementary material to this article is available at and is accessible to authorized users.     

Reversal of neuromotor and cognitive dysfunction in an enriched environment combined with multimodal early onset stimulation after traumatic brain injury in rats

This study was designed to investigate the additional benefits of a multimodal early onset stimulation (MEOS) paradigm when combined with enriched environment (EE) versus EE only and standard housing (SH) on the recovery after experimental traumatic brain injury (TBI). Male Sprague- Dawley rats were subjected to moderate lateral fluid percussion (LFP) brain injury (n = 40) or sham operation (n = 6). Thereafter, the injured and sham/EE + MEOS and EE only groups were placed into a complex EE consisting of tunnel-connected wide-bodied cages with various beddings, inclining platforms, and toys. Along with group living and environmental complexity, injured and sham/EE + MEOS animals were additionally exposed to a standardized paradigm of multimodal stimulation including auditory, visual, olfactory, and motor stimuli. In contrast, injured and sham/SH groups were housed individually without stimulation. A standardized composite neuroscore (NS) test was used to assess acute post-traumatic neuromotor deficits (24 h after injury) and recovery on days 7 and 15; recovery of cognitive function was assessed on days 11-15 using the Barnes maze. Neuromotor impairment was comparable in all injured animals at 24 h post-injury, but braininjured EE + MEOS rats performed significantly better than both brain-injured SH and EE groups when tested on post-injury days 7 and 15 (p = 0.004). Similarly, latencies to locate the hidden box under the Barnes maze platform were significantly shortened in EE + MEOS animals at day 15 (p = 0.003). These results indicate that the reversal of neuromotor and cognitive dysfunction after TBI can be substantially enhanced when MEOS is added to EE.     

Multimodal early onset stimulation combined with enriched environment is associated with reduced CNS lesion volume and enhanced reversal of neuromotor dysfunction after traumatic brain injury in rats

This study was designed to determine whether exposure to multimodal early onset stimulation (MEOS) combined with environmental enrichment (EE) after traumatic brain injury (TBI) would improve neurological recovery and to elucidate its morphological correlates. Male Sprague-Dawley rats were subjected to lateral fluid percussion (LFP) brain injury or to sham operation. After LFP, one-third of the animals (injured and sham) were placed under conditions of standard housing (SH), one-third were kept in EE only, and one-third received EE + MEOS. Assessment of neuromotor function 24 h post-injury using a standardized composite neuroscore test revealed an identical pattern of neurological impairment in all animals subjected to LFP. Neuromotor dysfunction in SH animals remained on a similar level throughout the experiment, while improvements were noted in both other groups 7 days post-injury (dpi). On 15 dpi, reversal of neuromotor dysfunction was significantly better in EE + MEOS animals vs. SH- and EE-only groups. In parallel, the comparison of lesion volume in EE + MEOS- vs. EE-only vs. SH rats revealed that animals exposed to EE + MEOS had consistently the lowest values (mm3, mean +/- SD; n = 6 rats in each group) as measured in serial brain sections immunostained for neuron-specific enolase (5.2 +/- 3.4 < or = 5.5 +/- 4.1 < 9.5 +/- 1.9), caspase 3-active/C3A (5.9 +/- 4.0 < or = 6.4 +/- 3.9 < 10.3 +/- 1.8) and glial fibrillary acidic protein (6.0 +/- 3.4 < or = 6.5 +/- 4.3 < 10.7 +/- 1.2). This first report on the effect of EE + MEOS treatment strongly indicates that the combined exposure reduces CNS scar formation and reverses neuromotor deficits after TBI in rats.     

A patient with sinus thrombosis associated with paroxysmal nocturnal hemoglobinuria

A 27-year-old woman with a history of aplastic anaemia complained of poor control of her right arm and hand, unsteady gait, and headache that increased while in a recumbent position. She was diagnosed with cerebral sinus thrombosis. Additional investigation revealed paroxysmal nocturnal haemoglobinuria (PNH). Treatment with heparin was initiated but stopped after the patient developed a brain haemorrhage. The patient recovered with no signs of residual symptoms and began taking oral anticoagulants as maintenance therapy. PNH is a rare acquired clonal disorder due to a defective expression of the glycosylphosphatidyl-inositol anchor membrane protein. It is characterised by haemolytic anaemia, diminished haematopoiesis, increased susceptibility for infections and a hypercoagulable state. Patients with aplastic anaemia have an increased risk of developing PNH. In patients with cerebral sinus thrombosis PNH should be considered as a possible underlying disorder. These patients should be questioned for possible clinical symptoms of PNH, such as acute abdominal pain or dark urine in the morning. For patients with these symptoms and in those with a history of aplastic anaemia or recurrent thrombosis, additional testing for PNH should be conducted.     

Infant mortality reviews in the Aberdeen Area of the Indian Health Service: strategies and outcomes

OBJECTIVE: To determine cause and manner of death for consecutive infant deaths in the Aberdeen Area of the Indian Health Service (AAIHS) from 1998 to 2002 and to identify risk markers for infant mortality. METHODS: Infant deaths in the AAIHS were identified from four data sources: death certificates from the four states in the AAIHS, deaths reported by local IHS Service Units, from obituaries in local and regional newspapers, and deaths reported by area hospitals. Each infant death is then sent to the local IHS service unit for review, where data from the infant and mother's chart is extracted and recorded. Local community factors, birth and death certificates, and autopsy reports are collected. The case is then reviewed at the Perinatal Infant Mortality Review (PIMR) meeting and a cause and manner of death is assigned. Summary data for the cohort was examined and then compared by mortality category and three age-at-death groups. RESULTS: Sudden infant death syndrome accounted for 33% of all infant deaths in the AAIHS. Prematurity was the second most prevalent cause-specific mortality category, accounting for 22% of all infant deaths. The authors found that infant mortality was surprisingly recurrent, with 32% of mothers of this infant having had a previous infant death. CONCLUSIONS: The PIMR committee requires substantial resources to support a review committee with appropriate expertise and their travel. Participation of local IHS staff and tribal members provides an important cultural and community perspective for the review process. Quality improvement changes are currently being implemented. These include increasing data on substance use, mental health needs, and reviews of fetal deaths. The process of mortality review has been very helpful in public education in the AAIHS.     

Effects of copper on enchytraeids in the field under differing soil moisture regimes

The aims of this study were to investigate the combined effects of drought stress and copper pollution on enchytraeids under natural conditions in the field and to compare the results of laboratory toxicity tests with results of the field study. Such studies were conducted to increase the understanding of interactions between chemicals and natural stressors and assess the predictive value of standardized laboratory tests with enchytraeids. The combined effect of copper and summer drought on enchytraeids was investigated in an old copper-contaminated field site at Hygum, Denmark, in three areas with different copper burdens. Each area consisted of five plots, which were divided into two subplots: one control and one drought subplot in which precipitation was excluded for a 45-d period during summer. Enchytraeids were sampled in spring (before the enforced drought began) and in autumn (after recovery from drought). Clear effects of copper were evident in both the field and the laboratory experiment. The field population density and species composition was highly affected by copper at concentrations in the range 300 to 500 mg Cu/kg dry soil and higher. In particular, a greatly impoverished species diversity was found in the copper-polluted areas. The effects of copper in the field compared reasonably well with the results of the laboratory tests. Surprisingly, possible effects of summer drought in the field were not detected in the autumn sampling, perhaps because of rapid recovery of the enchytraeid populations in both unpolluted and copper-polluted areas.     

The molecular genetics and morphometry-based endometrial intraepithelial neoplasia classification system predicts disease progression in endometrial hyperplasia more accurately than the 1994 World Health Organization classification system

BACKGROUND: The objective of this study was to compare the accuracy of disease progression prediction of the molecular genetics and morphometry-based Endometrial Intraepithelial Neoplasia (EIN) and World Health Organization 1994 (WHO94) classification systems in patients with endometrial hyperplasias. METHODS: A multicenter, multivariate analysis was conducted on 477 patients with endometrial hyperplasia who were required to have a 1-year minimum disease-free interval from the time of the index biopsy (1-18 years of follow-up). The results from that analysis were compared with the results from 197 patients who had < 1 year of follow-up. RESULTS: Twenty-four of 477 hyperplasias (5.0%) progressed to malignant disease over an average of 4 years (maximum, 10 years). According to the WHO94 classification, 16 of 123 atypical hyperplasias (13%) and 8 of 354 nonatypical hyperplasias (2.3%) progressed (hazard ratio [HR] = 7). Twenty-two of 118 EINs (19%) and 2 of 359 non-EINs (0.6%) progressed (HR = 45). EIN was prognostic within each WHO94 subcategory. Progression rates were 3% in simple hyperplasias, 22% in complex hyperplasias, 17% in simple atypical hyperplasias, and 38% in complex atypical hyperplasias with EIN, compared with progression rates of 0.0-2.0% in all hyperplasias if EIN was absent. EIN detected precancerous lesions (sensitivity, 92%) better than WHO94 atypical hyperplasias collectively (67%) or complex atypical hyperplasias alone (46%). In a Cox regression analysis, EIN was the strongest prognostic index of future endometrial carcinoma. The same was true for patients with < 1 year of follow-up (HR for EIN, atypical hyperplasia, and complex atypical hyperplasia: 58, 7, and 8, respectively). CONCLUSIONS: The EIN classification system predicted disease progression more accurately than the WHO94 classification and identified many women with benign changes that would have been regarded as high risk according to the WHO94 classification system.     

Successful treatment of low-grade oligodendroglial tumors with a chemotherapy regimen of procarbazine, lomustine, and vincristine

BACKGROUND: Anaplastic oligodendroglioma (OD) tumors, especially those with the combined loss of the short arm of chromosome 1 (1p) and the long arm of chromosome 19 (19q), are sensitive to chemotherapy. Only limited data are available on the role of chemotherapy in low-grade OD. The authors retrospectively studied the outcome of the procarbazine, lomustine, and vincristine (PCV) chemotherapy regimen in a group of 16 patients with newly diagnosed OD and 5 patients with recurrent low-grade OD. METHODS: Two groups of patients were studied: newly diagnosed patients with large OD and mixed oligoastrocytomas (OA) and patients with recurrent OD and OA after radiotherapy who still showed nonenhancing tumors. Treatment consisted of standard PCV chemotherapy. In the newly diagnosed and responding patients, radiotherapy was withheld until the time of disease recurrence. Responses were assessed by T2-weighted magnetic resonance image (MRI) scans. Loss of chromosome 1p and 19q was assessed using fluorescent in situ hybridization with locus-specific probes. RESULTS: Three of five patients with recurrent tumors responded. Thirteen of the 16 newly diagnosed patients showed evidence of response. The median time to disease progression in this group was >24 months. Only one of these patients experienced disease progression while receiving chemotherapy. Several patients showed a signficant clinical improvement despite only a modest improvement of the tumor on the MRI scans. Even patients without loss of 1p or 19q showed satisfactory responses. No TP53 mutations were found. CONCLUSIONS: Newly diagnosed patients with OD tumors, with or without loss of 1p/19q, responded to PCV chemotherapy. Up-front chemotherapy may be indicated especially for patients with large tumors. MRI scans were of limited value for the assessment of response. A Phase III trial should be initiated to compare radiotherapy with chemotherapy.