Long-term outcome of benign paroxysmal positional vertigo.

To clarify long-term outcome of benign paroxysmal positional vertigo (BPPV), 50 patients were followed up for a mean of 52 months. Overall recurrence rate by Kaplan-Meier estimation was 37% at 60 months. The patients with horizontal canal BPPV (n = 19) had a significantly higher recurrence rate (50%) at 60 months than those with posterior canal BPPV (n = 28; 26%). There was no significant association between recurrence rates and sex or age.

Construction of male and female PAC genomic libraries suitable for identification of Y-chromosome-specific clones from the liverwort, Marchantia polymorpha.

Unlike higher plants, the dioecious liverwort, Marchantia polymorpha, has uniquely small sex chromosomes, with X chromosomes present only in female gametophytes and Y chromosomes only in male gametophytes. We have constructed respective genomic libraries for male and female plantlets using a P1-derived artificial chromosome (pCYPAC2). With an average insert size of approximately 90 kb, each PAC library is estimated to cover the entire genome with a probability of more than 99.9%. Male-specific PAC clones were screened for by differential hybridization using male and female genomic DNAs as separate probes. Seventy male-specific PAC clones were identified. The male specificity of one of the clones, pMM4G7, was verified by Southern hybridization and PCR analysis. This clone was indeed located on the Y chromosome as verified by fluorescence in situ hybridization (FISH). This result shows that the Y chromosome contains unique sequences that are not present either on the X chromosome or any of the autosomes. Thus, the respective male and female libraries for M. polymorpha offer an opportunity to identify key genes involved in the process of sex differentiation and this unique system of sex determination.

Comparison of expressed sequence tags from male and female sexual organs of Marchantia polymorpha.

A total of 935 expressed sequence tags (ESTs) from male immature sexual organ were determined, of which 600 ESTs were assembled into 110 non-redundant groups, resulting in 445 unique EST sequences. Of these, 244 sequences shared significant similarities to known nucleotide or amino acid sequences in other organisms. The remaining 201 unique sequences showed no significant matches and thus are likely to be novel transcripts. ESTs from male and female immature sexual organs of a liverwort, Marchantia polymorpha, were compared to characterize gene expression patterns during sex differentiation. Ninety-nine male ESTs turned out to be common genes found also in the female library. Interestingly, one of the ESTs found only in male shows a significant similarity to the transformer-2 gene involved in sex determination in Drosophila. In female, several unique lectin ESTs were found that are not present in the male library.

Expressed sequence tags from immature female sexual organ of a liverwort, Marchantia polymorpha.

A total of 970 expressed sequence tag (EST) clones were generated from immature female sexual organ of a liverwort, Marchantia polymorpha. The 376 ESTs resulted in 123 redundant groups, thus the total number of unique sequences in the EST set was 717. Database search by BLAST algorithm showed that 302 of the unique sequences shared significant similarities to known nucleotide or amino acid sequences. Six unique sequences showed significant similarities to genes that are involved in flower development and sexual reproduction, such as cynarase, fimbriata-associated protein and S-receptor kinase genes. The remaining unique 415 sequences have no significant similarity with any database-registered genes or proteins. The redundant 123 ESTs implied the presence of gene families and abundant transcripts of unknown identity. Analyses of the coding sequences of 61 unique sequences, which contained no ambiguous bases in the predicted coding regions, highly homologous to known sequences at the amino acid level with a similarity score greater than 400, and with stop codons at similar positions as their possible orthologues, indicated the presence of biased codon usage and higher GC content within the coding sequences (50.4%) than that within 3' flanking sequences (41.9%).

Bryophyte 5S rDNA was inserted into 45S rDNA repeat units after the divergence from higher land plants.

The 5S ribosomal RNA genes (5S rDNA) are located independently from the 45S rDNA repeats containing 18S, 5.8S and 26S ribosomal RNA genes in higher eukaryotes. Southern blot and fluorescence in situ hybridization analyses demonstrated that the 5S rDNAs are encoded in the 45S rDNA repeat unit of a liverwort, Marchantia polymorpha, in contrast to higher plants. Sequencing analyses revealed that a single-repeat unit of the M. polymorpha nuclear rDNA, which is 16,103 bp in length, contained a 5S rDNA downstream of 18S, 5.8S and 26S rDNA. To our knowledge, this is the first report on co-localization of the 5S and 45S rDNAs in the rDNA repeat of land plants. Furthermore, we detected a 5S rDNA in the rDNA repeat of a moss, Funaria hygrometrica, by a homology search in a database. These findings suggest that there has been structural re-organization of the rDNAs after divergence of the bryophytes from the other plant species in the course of evolution.

[Human embryo freezing with dimethylsulfoxide (DMSO) and sucrose as cryoprotectants].

176 supernumerary human embryos following treatment by IVF were cryopreserved with dimethylsulfoxide (1.5M) and sucrose (0.1M) as cryoprotectants. The embryos were frozen at the early cleavage stage (48-96 hours after insemination) in the programmable freezer, by means of the protocol providing for slow cooling until -40 degrees C and slow thawing. The survival rate (50% or more blastomeres were intact) after thawing was higher for the embryos at the 8 cell stage to the morulla stage (60-96 hours after insemination) than for the embryos at the 4 to 6 cell stage (48-60 hours after insemination) (87% and 66%, p < 0.001), and the overall survival rate was 80% (140/176). Out of 62 transfer cycles, 11 pregnancies (17.7%) and 6 deliveries (9.7%) were achieved. No fetal malformation or chromosomal aberration was observed.

High glucose-6-phosphatase activity in osteoblasts in the metaphysis of femur of growing rats.

Glucose-6-phosphatase (G6Pase) activity was examined cytochemically in the metaphysis of femurs of 3- and 7-day-old rats. G6Pase and hexokinase activities were also examined biochemically in the femur and tibia of 3-day-old animals. The reaction product for G6Pase activity was seen in the endoplasmic reticulum and nuclear envelope of all cell types composing the metaphysis. The amount of the reaction product was abundant in osteoblasts, moderate in osteocytes, and moderate to scarce in osteoclasts and capillary endothelial cells. Biochemical G6Pase activity in the bones was higher than that in the brain, submandibular gland, or pancreas of the animals. Hexokinase activity in the bones was not different from that in the submandibular gland, pancreas, or kidney. The activity ratio of G6Pase and hexokinase in the bones (0.603) was greater than that in the submandibular gland, pancreas, or brain and smaller than that in the kidney. Possible physiological significances of the higher G6Pase activity in osteoblasts are discussed.

Glucose 6-phosphatase and glycogen phosphorylase activities in chondrocytes in epiphyseal cartilage of growing rats.

Glycogen, glycogen phosphorylase, and glucose 6-phosphatase (G6Pase) activities were examined cytochemically in chondrocytes of femoral epiphyseal cartilages and cartilaginous ribs of 3- and 7-day-old rats. G6Pase activity was also examined biochemically. Glycogen was abundant in chondrocytes of the reserve zone, while it became scarce in the cells of the proliferative zone. From the upper part (adjoining the proliferative zone) to the lower part of the hypertrophic zone, glycogen accumulated in chondrocytes and decreased in the cells of the degenerative zone. Inversely, glycogen phosphorylase a and G6Pase activities were relatively high in chondrocytes of the proliferative zone and upper hypertrophic zone and were low in the cells of the reserve zone, lower hypertrophic zone, and degenerative zone. The reaction product for G6Pase was present in the endoplasmic reticulum and nuclear envelope of all types of chondrocytes composing the cartilages, although the amounts of reaction product varied with the cell types in parallel with the histochemical results. Biochemical G6Pase activity was higher in epiphyseal cartilages than in cartilaginous ribs. The possible mechanism and significance of the accumulation and decrease of glycogen in chondrocytes of the epiphyseal cartilage were discussed.

Significance of increase in glucose 6-phosphatase activity in brown adipose cells of cold-exposed and starved mice.

Cytochemical and biochemical glucose 6-phosphatase (G6Pase) activity was examined in brown adipose tissues of normal, cold-exposed, or starved mice. In addition, G6Pase activity in white adipose tissue and hexokinase activity in brown and white adipose tissues were biochemically measured. In normal animals, the reaction product for G6Pase activity was localized in the endoplasmic reticulum and nuclear envelope of brown adipose cells. The amount of the reaction product increased in cold-exposed or starved animals. Biochemical G6Pase activity (259.7 +/- 48.5 ng Pi/min/mg protein) in brown adipose tissues of normal animals was higher when the value was compared with values of other organs. Biochemical G6Pase and hexokinase activities increased rapidly in brown adipose tissues of cold-exposed animals, and a close relation was found between activities of the two enzymes. In brown adipose tissues of animals starved for 3 days, biochemical G6Pase activity increased, but hexokinase activity did not change. In white adipose tissues of normal, cold-exposed, or starved animals, G6Pase activity was very low, although the enzyme activity increased slightly in animals starved for 3 days. The results show that the high G6Pase activity in brown adipose cells probably relates to thermogenesis in cold-exposed animals and may be concerned with glucose release into the blood in starved animals.

Physiological role of skeletal muscle glycogen in starved mice.

To study the physiological role of skeletal muscle glycogen in starved animals, effects of starvation on glycogen and glycogen phosphorylase (EC 2.4.1.1.) activity were studied in muscle fibers (morphologic study) and in whole muscles (biochemical study) of the rectus femoris muscle of mouse. Glycogen content in the liver of the starved animals was also measured. PAS reaction, strong in muscle fibers of fed animals, became weak predominantly in type IIB fibers after 2 days and almost disappeared after 4 days of starvation. Glycogen particles, numerous in the sarcoplasm between myofibrils of muscle fibers, decreased markedly predominantly in type IIB fibers after 2 days and almost disappeared after 4 days. Phosphorylase a activity, undetected in fibers of fed mice, appeared weak in type IIB fibers and very weak in type IIA fibers after 2 days and became moderate in type IIB fibers and weak in type IIA fibers after 4 days. Muscle glycogen content did not differ by 16 hours from the values of corresponding fed animals. However, liver glycogen content had already decreased after 8 hours and markedly so after 12 hours. The results support our hypothesis-"skeletal muscle glycogen is used for maintaining the blood glucose level in starved mice" (Hirose et al.: Anat. Rec., 216:133-138, 1986)-and show that type IIB fibers play a main role in maintaining the glucose level and that muscle glycogen is utilized after depletion of liver glycogen.