Transglutaminase 1-deficient recessive lamellar ichthyosis associated with a LINE-1 insertion in Jack Russell terrier dogs.

BACKGROUND: Congenital, nonepidermolytic cornification disorders phenotypically resembling human autosomal recessive ichthyosis have been described in purebred dog breeds, including Jack Russell terrier (JRT) dogs. One cause of gene mutation important to humans and dogs is transposon insertions. OBJECTIVES: To describe an autosomal recessive, severe nonepidermolytic ichthyosis resembling lamellar ichthyosis (LI) in JRT dogs due to insertion of a long interspersed nucleotide element (LINE-1) in the transglutaminase 1 (TGM1) gene. METHODS: Dogs were evaluated clinically, and skin samples were examined by light and electron microscopy. Phenotypic information and genotyping with a canine microsatellite marker suggested TGM1 to be a candidate gene. Genomic DNA samples and cDNA generated from epidermal RNA were examined. Consequences of the mutation were evaluated by Western blotting, quantitative reverse transcription-polymerase chain reaction (RT-PCR) and enzyme activity from cultured keratinocytes. RESULTS: Affected dogs had generalized severe hyperkeratosis. Histological examination defined laminated to compact hyperkeratosis without epidermolysis; ultrastructurally, cornified envelopes were thin. Affected dogs were homozygous for a 1980-bp insertion within intron 9 of TGM1. The sequence of the insertion was that of a canine LINE-1 element. Quantitative RT-PCR indicated a significant decrease in TGM1 mRNA in affected dogs compared with wild-type. TGM1 protein was markedly decreased on immunoblotting, and membrane-associated enzyme activity was diminished in affected dogs. CONCLUSIONS: Based on morphological and molecular features, this disease is homologous with TGM1-deficient LI in humans, clinically models LI better than the genetically modified mouse and represents its first spontaneous animal model. This is the first reported form of LI due to transposon insertion.

Activation of the redox-regulated molecular chaperone Hsp33--a two-step mechanism.

BACKGROUND: Hsp33 is a novel redox-regulated molecular chaperone. Hsp33 is present in the reducing environment of the cytosol and is, under normal conditions, inactive. The four highly conserved cysteines found in Hsp33 constitute a novel zinc binding motif. Upon exposure to oxidative stress, Hsp33's chaperone activity is turned on. This activation process is initiated by the formation of two intramolecular disulfide bonds. Recently, the 2.2 A crystal structure of Hsp33 has been solved, revealing that Hsp33 is present as a dimer in the structure (Vijayalakshmi et al., this issue, 367-375 [1]). RESULTS: We show here that oxidized, highly active Hsp33 is a dimer in solution. In contrast, reduced and inactive Hsp33 is monomeric. The incubation of reduced Hsp33 in H(2)O(2) leads to the simultaneous formation of two intramolecular disulfide bonds and the concomitant release of zinc. This concentration-independent step is followed by a concentration-dependent association reaction. The dimerization of Hsp33 requires highly temperature-sensitive structural rearrangements. This allows Hsp33's activation process to be greatly accelerated at heat shock temperatures. CONCLUSIONS: The regulation of Hsp33's chaperone function is highly sophisticated. On a transcriptional level, Hsp33 is under heat shock control. This increases the concentration of Hsp33 under heat and oxidative stress, a process that favors dimerization, a critical step in Hsp33's activation reaction. On a posttranslational level, Hsp33 is redox regulated. Dimerization of disulfide-bonded Hsp33 monomers leads to the formation of two extended, putative substrate binding sites. These sites might explain Hsp33's high and promiscuous affinity for unstructured protein folding intermediates.

Large-capacity separation of malignant cells and lymphocytes from the Furth mast cell tumor in a reorienting zonal rotor.

The Furth murine mastocytoma was adapted to the ascitic form and separated into fractions enriched with respect to lymphocytes and malignant cells by velocity sedimentation in the SZ-14 reorienting zonal rotor or in the isokinetic gradient. Lymphocytes were more highly purified (p less than 0.01) in the isokinetic gradient than in the zonal rotor, i.e., lymphocytes comprised 99.1% of the nucleated cells in the purest fraction from the isokinetic gradient and 80.1% of the nucleated cells in the purest fraction from the zonal rotor. Neoplastic mast cells were similarly purified by the two methods; they comprised 67.7 and 78.5% of the nucleated cells in the purest fractions from the isokinetic gradient and zonal rotor, respectively. Up to 160 million tumor cells can be purified in a single step with the reorienting zonal rotor, whereas 30 to 40 million cells per gradient approach the limit of the isokinetic gradient. After centrifugation in the zonal rotor, recovery was 85.6 +/- 12% (S.D.) of the cells layered over the gradient; and the separated tumor cells retained their ability to form tumors when transplanted into mice. The separation of large numbers of lymphocytes and malignant cells from the same tumor in the SZ-14 rotor should aid in the biochemical and immunological characterization of cancer.

P300 delay and attenuation in schizophrenia: reversal by neuroleptic medication.

BACKGROUND: P300 amplitude reduction in schizophrenia has been found by many investigators, but P300 latency generally has been reported to be normal; however, conflicting findings are present in the literature, and interpretation has been confounded by medication effects and methodological differences. METHODS: This study used a standard auditory oddball paradigm to compare the latency, amplitude, and topographic distribution of P300s in neuroleptic-free schizophrenic patients with those of healthy controls. The patients then were treated for 6 weeks with either remoxipride or haloperidol, and their P300s were reassessed. RESULTS: P300s were attenuated and delayed among neuroleptic-free patients. There was no evidence of peak lateralization or amplitude asymmetry over temporal areas. Subsequent neuroleptic medication normalized P300 latencies and increased P300 amplitudes, but the latter remained below normal limits over all except frontal areas. There were no correlations between P300 latency or amplitude and clinical symptomatology either before or after treatment. CONCLUSIONS: The finding of a P300 delay in neuroleptic-free schizophrenics that is normalized by neuroleptic medication has not been reported previously. Neuroleptic effects on P300 amplitude and latency appear to be independent of effects on clinical symptoms, and cannot be attributed to anticholinergic activity.

HIV and the brain: evidence of early involvement and progressive damage.

AIDS is often accompanied by progressive encephalopathy and 'subcortical' dementia, but there is uncertainty regarding how early the brain involvement may begin in the course of HIV infection. This study used a cognitive auditory 'oddball' paradigm to elicit sensory and cognitive event related potential (ERP) components from healthy controls and from patients at different stages of HIV infection. Sensory component latencies did not differ between groups, but cognitive components showed progressive delays corresponding to increasingly severe clinical stages of HIV infection. The earliest changes were found among asymptomatic HIV + patients, suggesting that this test is a sensitive indicator of early subclinical CNS damage. In contrast, neither frequency analysis nor nonlinear dynamical analysis of the EEG showed differences between healthy controls and patients.

Intracellular Staphylococcus aureus induces apoptosis in mouse osteoblasts.

Staphylococcus aureus invades osteoblasts and is the primary cause of osteomyelitis. This study examined the ability of S. aureus to induce apoptosis in a mouse osteoblast cell line. The presence of intracellular S. aureus was demonstrated by transmission electron microscopy. Light microscopy was utilized to examine morphological changes in the osteoblasts following killing of extracellular bacteria. Cell rounding was observed, and dark centers due to condensation of chromatin were noted in cells in infected osteoblast cultures. DNA was isolated from infected osteoblast cultures, and electrophoresis revealed the laddering effect characteristic of cells undergoing apoptosis. Additionally, an in situ cell death detection assay was utilized to label apoptosis-induced DNA strand breaks. Apoptotic nuclei were present, providing further evidence that S. aureus induces apoptosis in osteoblasts.

Flash P2 delay in primary degenerative dementia of the Alzheimer type.

1. Flash visual evoked responses of 31 patients meeting DSM-III-R criteria for primary degenerative dementia of the Alzheimer type were compared with 31 healthy controls. 2. P1 latency was normal (75 +/- 6 msec Alzheimer's; 75 +/- 6 msec controls). 3. P2 was significantly delayed (162 +/- 32 msec, 147 +/- 20 msec, p < 0.03) among patients. 4. P1-P2 interpeak latency was significantly increased (87 +/- 32 msec, 71 +/- 21 msec, p < 0.03) among patients.

The effects of thyroid hormones on the skin of beagle dogs.

The effects of hypothyroidism on canine skin were determined by comparing morphologic, morphometric, and hair cycle differences in skin biopsy samples from 3 groups of age- and gender-matched Beagle dogs: (1) euthyroid dogs; (2) dogs made hypothyroid by administration of 131I; and (3) dogs made hypothyroid and maintained in a euthyroid state by treatment with synthetic thyroxine. After 10 months of observation, there was slower regrowth of hair 2 months after clipping in the untreated-hypothyroid dogs. Untreated-hypothyroid dogs had a greater number of follicles in telogen and fewer hair shafts (ie, a greater number of hairless telogen follicles) than did the control group. The control dogs had a greater number of telogen follicles but the same number of hair shafts as the treated-hypothyroid group. Treated-hypothyroid dogs had the greatest number of follicles in the growing stage of the hair cycle (anagen). This study suggests that, at least in Beagles, induced hypothyroidism does not affect the pelage as dramatically as has been described in naturally occurring disease. This is because normal Beagles retain hair shafts in follicles for long periods, and the alopecia of hypothyroidism appears to evolve slowly because of the prolongation of this haired telogen stage. The evaluation of thyroxine-treated hypothyroid dogs demonstrates that thyroid hormone supplementation of Beagle dogs with induced hypothyroidism stimulates hair growth.

Characterization of a new human diploid myeloid leukemia cell line (PLB-985) with granulocytic and monocytic differentiating capacity.

A new human diploid cell line, designated PLB-985, has been established from the peripheral blood of a patient with acute nonlymphocytic leukemia (ANLL). Cells of this line are capable of granulocytic and monocytic maturation in the presence of inducing agents. By morphology, the analysis of surface antigens, and cytochemical staining PLB-985 cells are myelomonoblasts. Transmission electron microscopy reveals them to be devoid of neutrophilic primary or secondary granules and to have an open chromatin pattern with frequent nucleoli. The modal karyotype of the line is 46,XX, with no consistent marker chromosomes or recognizable translocations. Myelomonoblasts of this line form colonies in soft agar and induce tumors (chloromas) in nude mice. Growth of the cells in the presence of dimethyl sulfoxide, cis-retinoic acid, or dibutyryl cyclic adenosine monophosphate results in granulocytic maturation as determined by morphology, histochemical staining characteristics, and incorporation of 35S-methionine into the neutrophil primary granule proteinases elastase and cathepsin G. The tumor-promoting phorbol ester phorbol myristate acetate induces PLB-985 cells to differentiate as monocytes. Cells grown in the presence of this agent rapidly become adherent to plastic, display markedly increased phagocytosis of latex particles, stain positively for alpha-naphthyl acetate esterase, and lose the ability to synthesize the neutrophilic proteinases. Induction of differentiation along either pathway is accompanied by a marked decrease in myc oncogene transcription.

P2 flash visual evoked response delay may be a marker of cognitive dysfunction in healthy elderly volunteers.

In an earlier study, 31 healthy elderly volunteers had normal cognitive function as measured by the Mini-Mental State Examination. Twenty-seven returned for further memory testing using the Wechsler Memory Scale-Revised (WMS-R). The P2 latency of the flash visual evoked response was positively correlated with age (p = .0008), but was not significantly related to gender. Nine of these 27 putatively healthy subjects had a delayed P2, suggestive of dementia. Although unaware of any memory deficits, 5 of the 27 had WMS-R Visual Memory Span percentile scores 1 or more standard deviations less than age-matched controls. Four of the five also had a significantly delayed P2 component. This positive correlation, adjusted for age, between poor Visual Memory Span performance and a delayed P2 was statistically significant (p < .025). These findings suggest that a delayed P2 in putatively healthy subjects is indicative of a visuospatial deficit which might be a precursor of dementia later.

EEG-based, neural-net predictive classification of Alzheimer's disease versus control subjects is augmented by non-linear EEG measures.

Attempts to classify Alzheimer's disease (AD) subjects versus controls using spectral-band measures of electroencephalographic (EEG) data typically achieve around 80% success. This study assessed the ability of adding non-linear EEG measures and using a neural-net classification procedure to improve this performance level. The non-linear EEG measures were estimated correlation dimension ("dimensional complexity," or DCx) and saturation (degree of leveling-off of DCx with increasing embedding dimension). In a sample of 39 subjects (14 ADs, 25 controls), it was found that (a) the addition of non-linear EEG measures improved the classification accuracy of the AD/control status of subjects, and (b) a back-percolation neural net predictively classified the subjects much better than the standard linear techniques of multivariate discriminant analysis or nearest-neighbor discriminant analysis.

Use of 1,25(OH)2-vitamin D3 in prevention of renal osteodystrophy: preliminary observations.

Hypercalcaemia regularly occurs when "prophylactic" 1,25-dihydroxyvitamin D3 (1,25[OH)2D3) in a dose of 1.0 microgram daily is given to haemodialysis patients without clinical biochemical or radiological evidence of osteodystophy. This dose is too large for such patients. Hypercalcaemia may persist for several weeks after withdrawal of 1,25(OH)2D3, particularly when previous attempts at control by dosage reduction have failed. Hypercalcaemia is better managed by stopping 1,25(OH)2D3, albeit temporarily, than by reducing the dose.