Analysis of cedar pollen time series: no evidence of low-dimensional chaotic behavior.

Much of the current interest in pollen time series analysis is motivated by the possibility that pollen series arise from low-dimensional chaotic systems. If this is the case, short-range prediction using nonlinear modeling is justified and would produce high-quality forecasts that could be useful in providing pollen alerts to allergy sufferers. To date, contradictory reports about the characterization of the dynamics of pollen series can be found in the literature. Pollen series have been alternatively described as featuring and not featuring deterministic chaotic behavior. We showed that the choice of test for detection of deterministic chaos in pollen series is difficult because pollen series exhibit [see text] power spectra. This is a characteristic that is also produced by colored noise series, which mimic deterministic chaos in most tests. We proposed to apply the Ikeguchi-Aihara test to properly detect the presence of deterministic chaos in pollen series. We examined the dynamics of cedar (Cryptomeria japonica) hourly pollen series by means of the Ikeguchi-Aihara test and concluded that these pollen series cannot be described as low-dimensional deterministic chaos. Therefore, the application of low-dimensional chaotic deterministic models to the prediction of short-range pollen concentration will not result in high-accuracy pollen forecasts even though these models may provide useful forecasts for certain applications. We believe that our conclusion can be generalized to pollen series from other wind-pollinated plant species, as wind speed, the forcing parameter of the pollen emission and transport, is best described as a nondeterministic series that originates in the high dimensionality of the atmosphere.

Overexpression of cyclin E and cyclin-dependent kinase 2 is correlated with development of hepatocellular carcinomas.

Mammalian cell cycle progression is regulated by the combined action of cyclins/cyclin-dependent kinases (CDKs) and CDK inhibitors. Abnormal expression as well as interaction of these proteins may result in malignant transformation of cells. To further address the role of these cell cycle proteins in hepatocellular carcinomas, we analyzed the expression of cyclin E and CDK2. A panel of livers with human hepatocellular carcinoma, liver cirrhosis, and chronic hepatitis were used as a human experimental system. The inbred LEC (Long-Evans with a cinnamon-like coat color) rats were used as an animal experimental HCC model. Immunohistochemical staining of serial paraffin sections was performed using antibodies to cyclin E and CDK2. The results showed that cyclin E and CDK2 were concurrently overexpressed in hepatocellular carcinomas both in human and rat livers. Western blot analysis and CDK2 kinase assay demonstrated expression levels of cyclin E and CDK2 and CDK2 kinase activity, respectively, and both were shown to increase along with the development of hepatocellular carcinomas. Analysis of the correlation between expression of cyclin E and CDK2 and clinicopathological parameters revealed a significant correlation between expression of cyclin E and tumor grade (P=0.013), and PCNA index (P=0.006) as well as CDK2 expression (P=0.015). Overexpression of CDK2 tended to be associated with poorly differentiated HCCs. The results suggest that overexpression of cyclin E and CDK2 plays an important role in the development of hepatocellular carcinoma.

Enhanced expression and activation of Ca(2+)/calmodulin-dependent protein kinase IV in hepatocellular carcinoma.

BACKGROUND: Ca(2+)/calmodulin-dependent protein kinase IV (CaM-kinase IV) is a multifunctional protein kinase that is expressed abundantly in the central nervous system and, to a lesser degree, in nonneuronal tissues such as the liver. In the current study, the authors demonstrated the expression of CaM-kinase IV in hepatocytes from hepatocellular carcinoma (HCC) in both humans and rats. METHODS: Immunoblotting and immunohistochemical analysis were performed to confirm the expression of CaM-kinase IV and CaM-kinase kinase in HCC occurring in both humans and rats. The kinase activity of CaM-kinase IV in the lysate of each of these liver supernatant fluids was measured using a specific substrate (peptide gamma) for this enzyme before and after phosphorylation by exogenously added CaM-kinase kinase. RESULTS: Marked positive staining of HCC hepatocytes was found and the subcellular staining pattern mainly was cytosolic. One immunoreactive band with a molecular weight of 64 kilodaltons, which was identical to an isoform of rat cerebellum CaM-kinase IV, was demonstrated by immunoblotting. Ca(2+)/calmodulin-dependent CaM-kinase IV activity was high in HCC and showed almost no difference in activity in specimens with and without CaM-kinase kinase phosphorylation. CONCLUSIONS: CaM-kinase IV was found to be expressed in HCC and might have been involved in the development of HCC. CaM-kinase IV that was expressed in cancerous hepatocytes was phosphorylated mainly by CaM-kinase kinase that also was expressed in tumor cells.

Polymorphisms of p21 cyclin-dependent kinase inhibitor and malignant skin tumors.

Polymorphisms of the cyclin-dependent kinase inhibitor gene p21 have recently been reported to be associated with several human cancers. To determine whether the polymorphisms are also associated with human skin cancers, we investigated the p21 polymorphisms in 165 healthy Japanese and 113 Japanese with malignant skin tumors: 30 squamous cell carcinoma, 20 malignant melanoma, 33 basal cell carcinoma, and 30 Bowen's disease. The p21 polymorphisms were characterized by single base substitutions in the following two sites: the last nucleotide of codon 31 of exon 2 and the site 20 nucleotides downstream from the 3' end of exon 3. The two polymorphic sites were reported to be firmly linked to each other. We have shown that the two sites were firmly linked to each other also in Japanese and that no associations of the polymorphisms with the skin cancers in Japanese were detected by statistical analysis. Although the p21 polymorphisms were found not to be involved with skin carcinogenesis, ethnic differences of the allele frequency distribution must be taken into account in studying the role of the p21 polymorphism in carcinogenesis.

Expression and subcellular localization of multifunctional calmodulin-dependent protein kinases-I, -II and -IV are altered in rat hippocampal CA1 neurons after induction of long-term potentiation.

Long-term potentiation (LTP) is considered to be associated with an increase in expression as well as activity of Ca(2+)/calmodulin-dependent protein kinases (CaMKs). LTP-induced and control hippocampal slices were studied by immunohistochemical and electronmicroscopic analyses using anti-CaMK-I, -II and -IV antibodies. All three kinases were demonstrated to increase their expression in CA1 neurons. CaMK-I was shown to mainly localize in the cytoplasm of the control and LTP-induced neurons, and a significant increase of immunoreactivity was observed in the latter neurons. A part of CaMK-I was found to translocate to the nuclei of LTP-induced hippocampal CA1 neurons. Direct evidence of the translocation of CaMK-II from cytoplasm to nuclei in LTP was demonstrated by immuno-electronmicroscopy. A significant increase in expression of CaMK-IV in the nuclei was also observed. Our data suggest that all the three CaMKs were actively involved in nuclear Ca(2+)-signaling in LTP.

Effects of luteinizing hormone, follicle-stimulating hormone, and epidermal growth factor on expression and kinase activity of cyclin-dependent kinase 5 in Leydig TM3 and Sertoli TM4 cell lines.

We examined the effects of luteinizing hormone (LH), follicle-stimulating hormone (FSH), and epidermal growth factor (EGF) on the expression and kinase activity of cyclin-dependent kinase 5 (Cdk5) in Leydig TM3 and Sertoli TM4 cell lines. Hormonal regulation of the expression and activity of Cdk5 by using normal and hypophysectomized rat testes was also investigated to elucidate its role. Cdk5 levels and kinase activity were significantly elevated in TM3 cells that were grown in the presence of 7.5% serum, EGF, or LH and were associated with an increase in testosterone production compared with controls. These increases were accompanied by an increase in proliferation of TM3 cells after treatment with serum or EGF but not with LH suggest that Cdk5 may be involved in cellular differentiation that is induced with LH treatment. In contrast, the presence of neither serum, EGF, nor FSH had a significant effect on Cdk5 activity levels in the Sertoli TM4 cell line, and there was no correlation with proliferative activity or transferrin levels. A significant decrease in Cdk5 expression and activity were noted in rat testis after hypophysectomy compared with normal rat testis and is associated with a simultaneous decrease in testosterone and transferrin levels. Immunohistochemical analysis revealed that Cdk5 was strongly expressed in the nuclei and cytoplasm of Leydig cells, Sertoli cells, spermatogonia, and peritubular cells of normal adult rat testis. After hypophysectomy, the pattern of Cdk5 staining differed markedly from that in normal rat testis and a profound reduction in staining of Cdk5 was observed in each tubule. Our results suggest that LH and EGF influence and modulate Cdk5 expression and activity in Leydig TM3 cells and may, conceivably, be involved in signal transduction cascades that are initiated by hormones or growth factors. Cdk5 in Sertoli TM4 cells is likely to possess some constitutive functions that are not affected by the cells' proliferation state. Moreover, Cdk5 is probably involved in the constitutive and hormonally stimulated activities of the rat testis, in addition to its involvement in cell proliferation.

Purification, identification and phosphorylation of annexin I from rat liver mitochondria.

Annexin was purified from rat liver mitochondria to an apparent homogeneity with a molecular weight of 35 kDa as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. The purified mitochondrial annexin (AXmito) was identified as annexin I by an immunoblot analysis using anti-annexin I antibody. The inhibitory effect of AXmito I on porcine pancreatic phospholipase A2 activity was as potent as that of bovine lung annexin I. The presence of annexin I in mitochondria was confirmed by an electron-microscopic study. AXmito I was shown to be phosphorylated by intrinsic protein tyrosine kinases on its tyrosine residues. This annexin was also phosphorylated by protein kinase C.

[Comparison of five patient-controlled analgesia drug delivery devices].

We report the performance and our impression of five patient-controlled analgesia (PCA) drug delivery devices commercially available; Atom PCA Pump 500, AP-II, Deltec CADD-PCA 5800, Sabratek 6060 and Verifuse. Each of these devices has unique features for PCA. However, these devices still leave some room for improvement. Especially, we hope that future devices will be lighter to carry and use dry batteries more economically. In order to use these devices effectively for the management of pain, it is important to understand their characteristics.

Calbrain, a novel two EF-hand calcium-binding protein that suppresses Ca2+/calmodulin-dependent protein kinase II activity in the brain.

A cDNA clone that encodes a novel Ca2+-binding protein was isolated from a human brain cDNA library. The gene for this clone, termed calbrain, encodes a 70-amino acid polypeptide with a predicted molecular mass of 8.06 kDa. The analysis of deduced amino acid sequence revealed that calbrain contains two putative EF-hand motifs that show significantly high homology to those of the calmodulin (CaM) family rather than two EF-hand protein families. By Northern hybridization analysis, an approximate 1.5-kilobase pair transcript of calbrain was detected exclusively in the brain, and in situ hybridization study revealed its abundant expression in the hippocampus, habenular area in the epithalamus, and in the cerebellum. A recombinant calbrain protein showed a Ca2+ binding capacity, suggesting the functional potency as a regulator of Ca2+-mediated cellular processes. Ca2+/calmodulin-dependent kinase II, the most abundant protein kinase in the hippocampus and strongly implicated in the basic neuronal functions, was used to evaluate the physiological roles of calbrain. Studies in vitro revealed that calbrain competitively inhibited CaM binding to Ca2+/calmodulin-dependent kinase II (Ki = 129 nM) and reduced its kinase activity and autophosphorylation.

Molecular cloning and characterization of a cysteine-rich 16.6-kDa prolamin in rice seeds.

An alcohol-soluble storage protein, a 16.6-kDa prolamin found in rice seeds, was purified from both the total protein body and purified type I protein body fractions. The partial amino acid sequences of three tryptic peptides generated from the purified polypeptide were analyzed. A part of the 16.6-kDa prolamin cDNA was amplified from developing seed mRNA by the reverse transcribed polymerase chain reaction using an oligo (dT) primer and a primer which was synthesized based on the partial amino acid sequence. The amplified product was used to isolate the full-length cDNA clone (lambda RP16) from a developing seed cDNA library. The cDNA has an open reading frame encoding a hydrophobic polypeptide of 149 amino acids. The polypeptide was rich in glutamine (20.0%), cysteine (10.0%), and methionine (6.9%). The cysteine content was higher than those of most other rice storage proteins. Messenger RNA of the 16.6-kDa prolamin was detected in seeds, but not in other aerial tissues.

[Tracheal intubation for emergent tracheostomy in a patient complicated with tracheal stenosis].

We report the management of anesthesia for emergent tracheostomy in a patient with severe tracheal stenosis. A 63-year-old male was scheduled for an emergency tracheostomy for severe tracheal stenosis due to the invasion of a thyroid cancer. A preoperative neck CT revealed the tracheal stenosis, extending from 1-2 cm below the vocal cord to the upper end of the sternum. The narrowest caliber was about 7 mm in transverse diameter. Moreover, the cancer was suspected to have a bleeding tendency. General anesthesia with endotracheal intubation was considered necessary to provide an open airway during tracheostomy. Anesthesia was induced with thiopental, and a 6.0 mm endotracheal tube with cuff was successfully introduced with a balloon introducer (AIRGUID E) using suxamethonium. We were able to perform tracheostomy uneventfully.

Osteoblastic differentiation is enhanced by rapamycin in rat osteoblast-like osteosarcoma (ROS 17/2.8) cells.

The effects of three immunosuppressants (rapamycin, FK506 and cyclosporin A) on the proliferation and differentiation of rat osteoblasts-like osteosarcoma cell line, ROS 17/2.8 (ROS) cells were examined in vitro. All immunosuppressants showed a direct inhibition on the proliferation of ROS cells with different potencies. Growth inhibition by rapamycin was stronger than that by FK506 or cyclosporin A. Rapamycin caused a significant increase in alkaline phosphatase (ALP) activity and in the expression of osteopontin and osteocalcin mRNAs. FK506 caused a moderate increase in ALP activity and a decreased expression of osteopontin mRNA. Cyclosporin A caused a decrease in ALP activity and in the expression of type 1 alpha 1 collagen mRNA. Our study indicates that rapamycin directly acts on ROS cells and induces osteoblastic differentiation, however, the effect of FK506 and cyclosporin A is weak. Rapamycin significantly enhances the differentiation induced by 1,25(OH)2-vitaminD3.

Expression of cyclin-dependent kinase 5 and associated cyclins in Leydig and Sertoli cells of the testis.

In this study, we examined the expression and subcellular localization of cyclin-dependent kinase 5 (Cdk5), cyclin D1, and cyclin E in Leydig and Sertoli cell lines that were cultured with 7.5, 1.0, 0.5, or 0% serum (mixture of a 2:1 ratio of horse serum and fetal bovine serum) and in the developing rat testis to verify the possible functions of Cdk5, cyclin D1, and cyclin E in the testis. The abundance of Cdk5 and cyclin E in the Leydig cell line, TM3, was significantly reduced at low serum concentrations. In contrast, serum concentration had no effect on Cdk5 and cyclin E levels in the Sertoli cell line, TM4. Cyclin D1 was detected by western blot analysis in TM4 cells only, and its abundance was serum dose dependent. The kinase activity of Cdk5 in TM3 and TM4 cells that were cultured at various serum concentrations coincided with the levels of Cdk5 expression. Immunohistochemical staining for Cdk5 and cyclin E revealed nuclear and cytoplasmic distribution, both in TM3 and TM4 cells. Moreover, cyclin D1 immunoreactivity was only detected in TM4 cells. In the developing rat testis, Cdk5 expression was most prominent at 2 and 3 weeks after birth. Cyclin D1 was strongly expressed at 1 and 2 weeks in premature rat testes. On the other hand, cyclin E was highly expressed in the adult testis. Immunohistochemical localization of Cdk5, cyclin D1, and cyclin E in 1-week-old and adult rat testes revealed expression in both Leydig and Sertoli cells. Our results suggest that Cdk5 in TM3 and Leydig cells of the testis might play a role in cell cycle regulation, whereas Cdk5 in TM4 and Sertoli cells of the adult testis might have some additional functions besides control of proliferation.

Synthesis and antinociceptive activity of [D-Ala2]Leu-enkephalin derivatives conjugated with the adamantane moiety.

Based on the physicochemical and pharmacological properties of drugs having an adamantane skeleton, an adamantane-based moiety was evaluated as a drug carrier for poorly absorbed compounds, including peptides, active towards the central nervous system (CNS). Seven [D-Ala2]Leu-enkephalin derivatives conjugated with an adamantane-based moiety at the C-terminus or N-terminus were prepared by the solution-phase method and their biological activities were examined. The compounds derivatized at the C-terminus through an ester or amide linkage were much more lipophilic than the parent peptide and exhibited moderate in vitro opioid activity (guinea-pig ileum assay). Among them, four derivatives (1, 2, 4, 5), exhibited significant antinociceptive effects in an in vivo assay (mouse tail-pressure test) after subcutaneous administration. This result suggests that the introduction of the lipophilic adamantane moiety into [D-Ala2]Leu-enkephalin would improve the permeation of the poorly absorbed parent peptide through the blood-brain-barrier (BBB) without loss of antinociceptive effect.

Reticalmin: a novel calcium/calmodulin-dependent protein kinase IV-like protein in rat retina.

Western blot analysis of 100,000 g supernatant of rat retina using a polyclonal anti-Ca2+/ calmodulin-dependent protein kinase IV (CaM-kinase IV) antibody revealed an immunoreactive mass of 35 kDa, termed reticalmin. Lower amount of a isoform of CaM-kinase IV was also expressed in rat retina. Reticalmin did not react with anti-CaM-kinase IV C-terminal peptide antibody which recognized alpha and beta isoforms of CaM-kinase IV and calspermin. Immunohistochemically reticalmin was shown to be localized mainly in the outer segment of photo-receptor cells, and in dendrites of inner plexiform layers and may be in nuclei of ganglion cells and some inner nuclear layer cells.

[Massive bleeding from the ruptured liver and the inferior vena cava controlled with autotransfusion and cerebral ischemia treated with mild hypothermia].

A patient suffered massive bleeding from the ruptured liver and laceration of the inferior vena cava due to traffic accident, and developed hypotension and decreased level of consciousness. The patient was transferred to our hospital for an emergency operation against intra-abdominal massive bleeding. This massive bleeding was controlled with autotransfusion using washing salvaging autotransfusion device (Cell Savor). Suspected brain ischemia was treated with intended mild hypothermia. When blood pressure decreased to 30 mmHg of systolic pressure over 7 minutes during the operation, suggesting the possible brain ischemia, mild hypothermia was maintained at 33.8 degrees C. Total bleeding volume was 16,700 ml, and total transfused volume was 10,700 ml. Of total transfused volume, 4,500 ml was washed salvaged blood using the intraoperative autotransfusion device. No neurological deficit was found during the postoperative course. The patient was discharged uneventfully on the 20th postoperative day. In conclusion, intraoperative autotransfusion with washed salvaged blood is a useful method for treatment of massive bleeding, and mild hypothermia is efficacious for protecting the brain from ischemia resulting from accidental hypotension.

Involvement of calmodulin-dependent protein kinases-I and -IV in long-term potentiation.

Multifunctional Ca2+/calmodulin-dependent protein kinases (CaMKs) are thought to be involved in the induction of long-term potentiation (LTP). In the present study, LTP was induced by theta burst stimulation in the Schaffer collateral area of the stratum radiatum in the hippocampal CA1 region of the rat hippocampus. LTP-induced and control hippocampal slices were studied by Western blot and immunohistochemical analyses using CaMK-I, -II and -IV antibodies. Increased amounts of all three CaMKs were found in LTP-induced hippocampal slices as indicated by Western blot as well as by the density of their immunoreactivity. Our data clearly shows that not only CaMK-II but also CaMK-I and -IV contribute to synaptic plasticity formed in LTP.

[Premixing lidocaine reduces the incidence and severity of pain on injection of propofol].

The purpose of this study was to confirm the effect of premixed lidocaine for the reduction of pain during injection of propofol in adult patients. We conducted a prospective, randomized, double-blind trial on 106 patients. In the study group (n = 54), lidocaine 40 mg (2 ml of lidocaine 2%) was added to 180 mg of propofol (18 ml). In the control group (n = 52), 2 ml of normal saline was added to 180 mg of propofol. The pain on injection was rated as none, mild, moderate, or severe. Eleven patients (20.4%) in the study group experienced pain compared with 25 (48.1%) in the control group. Thirteen in the control group complained moderate or severe pain compared with only one in the study group. In conclusion, lidocaine 40 mg premixed with 180 mg propofol significantly reduces the incidence and severity of pain associated with propofol injection.