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Iron is an essential metal for all living organisms that is absorbed in the intestinal cells as a heme-chelated or free form. It is unclear how important plant-derived chelators, such as nicotianamine (NA), an organic small molecule that is ubiquitous in crops, vegetables, and various other foods, contribute to iron bioavailability in mammals. We performed electrophysiological assays with Xenopus laevis oocytes and radioactive tracer experiments with Caco-2 cells. The findings revealed that the proton-coupled amino acid transporter SLC36A1 (PAT1) transports iron in the form of NA-Fe (II) complex in vitro. Decreased expression of hPAT1 by RNA interference in Caco-2 cells reduced the uptake of NA-59Fe (II) complex. The uptake of inorganic 59Fe (II) was relatively unaffected. These results imply that PAT1 transports iron as a NA-Fe (II) complex. The rate of 59Fe absorption in the spleen, liver, and kidney was higher when mice were orally administered NA-59Fe (II) compared with free 59Fe (II). The profile of site-specific PAT1 expression in the mouse intestine coincided with those of NA and iron contents, which were the highest in the proximal jejunum. Orally administered NA-59Fe (II) complex in mice was detected in the proximal jejunum by thin layer chromatography. In contrast, much less 59Fe (or NA) was detected in the duodenum, where the divalent metal transporter SLC11A2 (DMT1) absorbs free Fe (II). The collective results revealed the role of PAT1 in NA-Fe (II) absorption in the intestine and potential implication of NA in iron uptake in mammals.

Tyrosine kinase inhibitors of epidermal growth factor receptor (EGFR) improve the survival of patients with lung adenocarcinoma, and determine the EGFR mutation status before treatment is necessary. In contrast to biopsy samples, cytological specimens are obtained less invasively and are useful for EGFR mutation analyses. Recently, novel antibodies against two major EGFR mutations were developed: SP111, which is specific for the E746-A750 deletion in exon 19; and SP125, which is specific for the L858R mutation. To the best of our knowledge, no study has evaluated cytological specimens using the two novel antibodies, thus their specificity and sensitivity were examined in surgical resection, and cytological lung adenocarcinoma samples in the present study. Previous screening for EGFR mutation status by molecular testing identified delE746-A750 in 3 cases and the L858R mutation in 7 cases; the other cases did not have the L858R or the delE746-A750 mutation. Using a four-grade scoring system (score 0 to 3+), the immunohistochemistry (IHC) and immunocytochemistry (ICC) results were compared with those of molecular testing. Using a score of ≥2 as positive, IHC and ICC using SP111 demonstrated sensitivities of 100 and 33.3%, and specificities of 100 and 100%, respectively. IHC and ICC using SP125 revealed sensitivities of 100 and 71.4%, and specificities of 100 and 100%, respectively. Therefore, screening for EGFR mutations by ICC may facilitate therapeutic decision-making, particularly in medical centers that are unable to perform molecular testing.

We report a case of systemic phaeohyphomycosis due to Exophiala dermatitidis in a 24-year-old man. At the age of 17, the patient had noticed cervical swellings. On palpation at his initial examination, more than a dozen firm lymph nodes between 1 and 5 cm in diameter were found on both sides of the neck and in both axillae. Examination of biopsy specimens of lymph nodes revealed pale brown hyphae in granulomatous lesions and Exophiala dermatitidis was isolated from mycological cultures of the local tissues. The clinical course was marked by an episode of jaundice six months later, and surgery disclosed a fist-sized inflammatory mass in the region of the head of the pancreas and the duodenum. The patient was treated by providing external biliary drainage and by the administration of antifungal agents, but two years later complained of diplopia and a slight heaviness in the head. Computed tomographic scans were made of the head, revealing multiple plum-sized masses in the brain. The patient died a year later. In Japan, 10 cases of systemic infection by this organism (including the present one) have been reported, all in patients of up to 30 years of age, with lesions appearing in the brain, lung, liver, digestive organs and lymph nodes. The prognosis is grave.

A number of tandemly reiterated sequences are present on the herpes simplex virus type 1 (HSV-1) DNA molecule of 152 kbp. While regions containing tandem reiterations were usually unstable, reiteration VII, which is present within the protein coding regions of gene US10 and US11, was stable; hence, reiteration VII could be used as a genetic marker. In the present study, the nucleotide sequences (159–213 bp) of a region encompassing reiteration VII of 62 HSV-1 isolates were compared with that of strain 17 as the standard strain, and the genetic variability of base substitutions, deletions, and multiplications was revealed. Base substitution was observed in nine residues on the region flanking reiteration VII and sixty-two HSV-1 isolates were classified into twelve groups based on these base substitutions. Deletions, which were present in all sixty-two isolates, were classified into six groups. Multiplications, which were present in 19 isolates having the same deletion (named del-2), were classified into four groups. The sixty-two isolates were classified into twenty patterns based on variations in the region encompassing reiteration VII, and the region encompassing reiteration VII was considered to be useful for studies on the molecular epidemiology and evolution of HSV-1. The lengths of these deletions and multiplications were multiples of 3; thus, a frame-shift mutation was not induced, and a mechanism to maintain the functions of US10 and US11 was suggested. A series of multiplications, which consisted of the duplication, triplication, and tetraplication of the same sequence, were found. Since all isolates with a multiplication had del-2, multiplications were assumed to be generated after the generation of del-2, and an isolate with del-2 was considered to have the ability to generate a multiplication. Recombination between a pair of direct repeats in and around reiteration VII was accountable for the generation of deletions and multiplications, indicating the recombinogenic property of the region encompassing reiteration VII. A correlation was revealed between a set of 20 DNA polymorphisms widely present on the HSV-1 genome and the base substitutions and deletions of the region encompassing reiteration VII, using discriminant analyses.

Summary Background Human parvovirus B19 (B19) infections exhibit various skin manifestations that are similar to and hence hard to distinguish from many other skin diseases. The virological diagnosis of B19 infection is usually based on time-consuming serological tests and polymerase chain reaction (PCR). Objectives In this study, a DNA amplification method, loop-mediated isothermal amplification (LAMP), was used for the diagnosis of B19 infection and was compared with PCR. Methods Ten patients with acute B19 infection and 16 patients with other skin diseases were enrolled. Sera and pharyngeal swabs were used directly as the templates in LAMP. The LAMP reaction was carried out at 63 °C for 1 h in a heat block. The reaction products were judged visually, by adding SYBR Green I® into the tubes, and by gel electrophoresis. Results B19 DNA was detected by LAMP in 10 sera and all of seven tested pharyngeal swabs of 10 patients with acute B19 infection but not in samples from 16 patients with other skin diseases. The results were in agreement with those obtained by PCR except for one case. The reason for the single discrepancy may be that the sensitivity of LAMP is 102 times higher than PCR. Conclusions Detection of B19 DNA by LAMP in serum and especially in the pharynx is a rapid and convenient method for the diagnosis of acute B19 infection.

To assess the role of nuclear factor kappaB (NFKB) in cellular radiosensitivity, three different IkappaB-alpha (also known as NFKBIA) expression plasmids, i.e., S-IkappaB (mutations at (32, 36)Ser), Y-IkappaB (a mutation at (42)Tyr), and SY-IkappaB, were constructed and introduced into human brain tumor M054 cells. The clones were named as M054-S8, M054-Y2 and M054-SY4, respectively. Compared to the parental cell line, M054-S8 and M054-Y2 cells were more sensitive to X rays while M054-SY4 cells exhibited the greatest sensitivity. After treatment with N-acetyl-Leu-Leu-norleucinal, a proteasome inhibitor, the X-ray sensitivity of M054-S8 and M054-SY4 cells did not change, while that of M054-Y2 cells and the parental cells was enhanced. An increase in X-ray sensitivity accompanied by a decrease in translocation of NFKB to the nucleus in parental cells was observed after treatment with pervanadate, an inhibitor of tyrosine phosphatase, as well as in M054-S8 and M054-SY4 cells. Repair of potentially lethal damage (PLD) was observed in the parental cells but not in the clones. Four hours after irradiation (8 Gy), the expression of TP53 and phospho-p53 ((15)Ser) was induced in the parental cells but not in M054-S8, M054-Y2 or M054-SY4 cells. Our data suggest that inhibition of IkappaB-alpha phosphorylation at serine or tyrosine acts independently in sensitizing cells to X rays. NFKB may play a role in determining radiosensitivity and PLD repair in malignant glioma cells; TP53 may also be involved.

Herpes simplex virus type 1 (HSV-1) strains belonging to the same genotype can possibly share biological properties and clinical manifestations common to the genotype. We classified previously 66 HSV-1 strains into 35 genotypes (F1-F35) using restriction fragment length polymorphism (RFLP) and F1 and F35 genotypes were revealed to be predominant [Arch. Virol. 13 (1993) 29]. It was found later that the F35 genotype seemed to be closely associated with eczema herpeticum [J. Med. Virol. 49 (1996) 329]. In the present study, a convenient method was developed for classification of two predominant genotypes by RFLP of polymerase chain reaction (RFLP-PCR). Using this method, genotypes of 21 strains isolated from eczema herpeticum were analyzed; seven of 21 strains (33.3%) were of F1 and five of 21 (23.8%) were of F35. Genotypes of 19 strains isolated from facial herpes other than eczema herpeticum were as follows; six of 19 (31.6%) strains were of F1 and one of 19 (5.3%) were of F35. Thus, strains belonging to F35 were appear to have been isolated more frequently from eczema herpeticum (5/21) than from facial herpes (1/19). These ratios showed a statistically significant difference. These results support the hypothesis that F35 strains is clearly associated with eczema herpeticum, in agreement with previous study. This is the first report of PCR-based approach for classification of HSV-1 strains into genotypes seeking an association of a genotype with clinical manifestation.

The antioxidant activities of 22 selected culinary herbs and spices (i.e. ginger, cinnamon, clove, bay, sage, rosemary, thyme, savory, oregano, sweet basil, parsley, coriander, tarragon, sansho, allspice, cumin, black and white peppercorns, nutmeg, caraway, dill and fennel) when they were added to pork homogenate were measured and expressed as a thiobarbituric acid (TBA) value. The addition of liquid extracts of all the herbs and spices significantly suppressed lipid oxidation of the pork, especially the extracts of sansho, sage and ginger, which showed the strongest inhibition of lipid oxidation.

It is established that extremely low frequency magnetic fields (ELFMF) at the flux densities, i.e., 5 mT and less, are not mutagenic. However, exposure to ELFMF enhances mutations induced by X-rays. In this study, we examined the effects of long-term exposure to 5 mT ELFMF on mutation induction and X-ray-induced mutations in human malignant glioma cells (MO54) with different mutant IkappaB-alpha (a critical inhibitor of NF-kappaB) genes. Cells were exposed or sham-exposed to 5 mT ELFMF for up to 8 days with or without initial X-rays (4 Gy), and the mutant frequency of hypoxanthine-guanine phosphoribosyl transferase (HPRT) gene was analyzed. An obvious increase in X-ray-induced mutations was observed after treatment with ELFMF in combination with X-irradiation in MO54 cells with tyrosine mutant IkappaB-alpha gene other than with serine mutant IkappaB-alpha gene or vector alone. Exposure to ELFMF alone increased mutations significantly in MO54 cells with tyrosine mutant IkappaB-alpha gene. In addition, X-ray-induced apoptoic cells were increased in MO54-V cells after exposure to ELFMF, while an anti-apoptotic effect of magnetic field was found in MO54-SY4 cells. Our data suggest that exposure to 5 mT ELFMF may induce mutations and enhance X-ray-induced mutations, resulting from the inactivation of NF-kappaB through the inhibition of tyrosine phosphorylation.

In the present study, we investigated the effects of acute and chronic systemic administration of MCI-225 (4-(2-fluorophenyl)-6-methyl-2-(1-piperazinyl)thieno[2,3-d]pyrimidine monohydrate hydrochloride), a newly-developed selective noradrenaline (NA) reuptake inhibitor with 5-HT3-receptor-blocking action, on extracellular NA levels in the hypothalamus of stressed and non-stressed rats by utilizing intracerebral microdialysis. Acute administration of MCI-225 (3 and 10 mg/kg, p.o.) significantly and dose-dependently increased extracellular NA levels in the hypothalamus in non-stressed rats. Footshock for 20 min also significantly increased NA levels in the hypothalamus of both groups of rats pretreated with vehicle and MCI-225. Although chronic administration of MCI-225 (3 or 10 mg/kg, p.o. for 14 days) did not alter the basal extracellular NA levels in the hypothalamus, the stress-induced increases in extracellular NA levels were significantly lower in rats chronically treated with MCI-225 (10 mg/kg) than those of rats pretreated with vehicle for the same period. The increase in extracellular NA levels induced by MCI-225 challenge (3 or 10 mg/kg, p.o.) were not different between rats chronically treated with MCI-225 or vehicle. These results suggest that MCI-225 enhances extracellular NA levels in the hypothalamus in both non-stressed and stressed rats by inhibiting NA uptake and that chronic systemic administration of MCI-225 did not alter basal extracellular NA levels, but reduced the increase in NA release caused by footshock stress. These data suggest the possibility that MCI-225 might possess anxiolytic and/or antidepressant properties.

To classify the clinical characteristics of ocular toxocariasis in Japan, the prevalence of antibodies to Toxocara antigens was examined in patients with uveitis of unknown aetiology. From 1982 to 1993, serum specimens of 383 cases and intraocular fluid samples of 22 cases were serologically screened for Toxocara infection with five immunodiagnostic tests. Fifty-five sera and 11 intravitreous fluid samples were estimated to have significantly high antibody levels against larval excretory–secretory (ES) antigens of T. canis. Eight cases were positive in both serum and vitreous fluid, and three were positive only in the vitreous fluid. Among the 58 antibody positive samples, 20 cases were omitted due to a lack of detailed description of ocular findings. The remaining 38 cases are described in this study. Of these 38 cases, 34 (89%) were older than 20 years of age. Ocular lesions were located in the posterior fundus in 11 cases, in the peripheral fundus in 18 cases, and in both areas in seven cases. Of the eight cases in which papillary oedema or redness was observed, chorioretinal lesions were also present in seven of them. Tractional retinal detachment was present in five cases. These observations suggest that ocular toxocariasis in Japan has a different clinical profile compared with those in the other countries, and indicate a need for revised classification of ocular toxocariasis.

We have previously demonstrated immunosuppression including reduced splenic natural killer cell activity (NKCA) in pregnant rats exposed to microwaves produced mainly by their thermal action. To examine the involvement of opioid systems in reduced NKCA in pregnant rats exposed to microwaves at a relatively low level (2 mW/cm2 incident power density at 2450 MHz for 90 min), we assayed beta-endorphin (betaEP) in blood, pituitary lobes, and placenta as well as splenic NKCA in virgin and/or pregnant rats. Although microwaves elevated colonic temperatures by 0.8 degreesC for virgin and 0.9 degreesC for pregnant rats, and betaEP in blood and anterior pituitary lobes (AP) significantly, it did not change blood corticosterone as an index of hypothalamic-pituitary adrenal axis. There were significant interactions between pregnancy and microwave exposure on splenic NKCA, betaEP in both blood and AP, and blood progesterone. Intra-peritoneal administration of opioid receptor antagonist naloxone prior to microwave exposure increased NKCA, blood, and placental betaEP in pregnant rats. Alterations in splenic NKCA, betaEP and progesterone in pregnant rats exposed to microwaves may be due to both thermal and nonthermal actions. These results suggest that NKCA reduced by microwaves during pregnancy is mediated by the pituitary opioid system.

In the post-genome era, several tools that have increased our global understanding of the molecular basis of several cell-based phenomena have been developed. However, proteomics has not been efficiently integrated with the other 'omics' (e.g. transcriptomics and metabolomics), because of the relatively low number of proteins identified by mass spectrometry (MS). Peptides from low-abundance proteins are often not detected by MS due to ionization suppression. To improve the number of peptide identifications in MS analyses, we propose three separation methodologies; namely, OFFGEL electrophoresis, 2D-liquid chromatography (LC) and the long monolithic silica-C18 capillary column method, with the common aim to decrease peptide complexity prior to MS analyses. Proteomics using the above three peptide separation methods were separately applied to protoplasts collected from the epidermal cell layer of Arabidopsis roots using fluorescence-activated cell sorting. In each method alone, 1,132, 836 and 795 proteins were specifically identified, respectively. This has allowed the identification of 1,493 proteins with no redundancy and with

Background: It has been reported that herpes simplex virus (HSV) DNA was detected in the oral cavity of patients with herpes labialis under various conditions such as during oral surgery. Objective: The frequency of detection of oral HSV DNA was compared between first or recurrent episodes of eczema herpeticum and recurrent type herpes labialis. Patients and Methods: Oral swabs were collected from 7 patients with eczema herpeticum and 9 with herpes labialis. The detection of oral HSV DNA was performed by the polymerase chain reaction method. Results: Oral HSV DNA was detected in 6 out of 7 patients (86%) with eczema herpeticum and 3 of 9 (33%) with herpes labialis. Conclusions: The high frequency of oral HSV DNA detection in eczema herpeticum suggests that subclinical herpetic lesions may develop in the oral cavity of patients with a first episode of eczema herpeticum or may occur during asymptomatic oral HSV shedding in people with recurrent eczema herpeticum.

Local application of muscimol through the striatal dialysis membrane caused a significant increase in both dopamine release and dopamine metabolism in the striatum of conscious rats, however, both elevations induced by muscimol were significantly lower in the kainic acid-lesioned striatum when assessed with in vivo brain microdialysis. These findings show that intra-striatal muscimol indirectly stimulates nigrostriatal dopaminergic function by possibly causing an inhibition of striatal γ-aminobutyric acid neurons.

The proto-oncogenes are thought to play important roles in the regulation of cellular growth and differentiation. In order to evaluate the role of proto-oncogenes in the regulation of growth of bronchial epithelial cells, we studied steady-state levels of fos, jun, and myc transcripts in response to fetal calf serum, bovine pituitary extract, and insulin. Extensively quiescent populations of bovine bronchial epithelial cells in growth factor-free medium were stimulated to divide by each of these three additives. We observed rapid but transient increases of fos, jun, and myc expression in association with such growth stimulation. There were no changes in tubulin mRNA levels over the same time periods. Other "growth factors" (epidermal growth factor, hydrocortisone, epinephrine, triiodothyronine, and transferrin) were also studied and did not affect either cell growth or expression of fos, jun, or myc. We further examined the effect of transforming growth factor-beta1 (TGF-beta1) on the above stimulatory effects. TGF-beta1 consistently inhibited the growth induced by fetal calf serum, bovine pituitary extract, or insulin and, interestingly, reduced proto-oncogene myc mRNA level without altering that of fos and jun. In conclusion, proto-oncogenes fos, jun, and myc appear to play a role in the regulation of growth response in bovine bronchial epithelial cells. It is also possible that TGF-beta1 exerts its growth inhibitory effect, at least in part, through the processes that involve the regulation of proto-oncogene myc transcription in these cells.

Herpes simplex virus type 1 (HSV-1) is a ubiquitous human pathogen. The a sequence of HSV-1 is the cis-acting site required for the cleavage and encapsidation of unit-length HSV-1 DNA from concatemeric forms. The consensus a sequence consists of (i) DR1 (direct repeat 1), (ii) Ub, (iii) a DR2 array [a repeat of various copy numbers of DR2 elements (11 or 12 bp)], (iv) a DR4 stretch and (v) Uc. In the present study, the nucleotide sequences of the a sequences of 26 HSV-1 isolates were determined and the DR4 stretches were classified into three groups. The state of a set of 20 DNA polymorphisms in the genomes of these HSV-1 isolates was determined previously. A correct classification rate of 100 % was achieved when discriminant analysis was performed between the DR4 stretch (criterion variable) and the set of 20 DNA polymorphisms (predictor variables), suggesting a close association of the DR4 stretch with HSV-1 diversification. DR2 elements of 9, 13 and 14 bp were detected in addition to those of 11 and 12 bp, and a correct classification rate of 93 % was achieved when discriminant analysis was performed between the DR2 array and the set of 20 DNA polymorphisms. Some DR2 elements of one HSV-1 isolate had the same nucleotide sequences as part of the adjacent DR4 stretch, and these variations were adequately explained by postulating recombination involving DR2 elements; hence, the DR2 array was deduced to be prone to recombination.

Background: A variable region, R2, on the varicella–zoster virus (VZV) genome contains a repeated 42-bp unit. Objective: The purpose of this study is the derivation of significance from tandem reiteration structure in the R2 region. Methods: Fifty-two specimens were collected from 52 patients with herpes zoster in Osaka and Tokyo, Japan. After treatment of the specimens to release viral DNA, the samples were amplified directly by polymerase chain reaction. In addition, 14 samples were collected from 7 of these zoster patients after valaciclovir or aciclovir therapy. Results: Analyses of the 52 specimens revealed that the number of repeats ranged from 4 to 13. Interestingly, the numbers of repeats among various VZV strains showed a normal distribution pattern, so that 6–9 repeats were found to be predominant in both Osaka (85%) and Tokyo (72%). The pre- and post-treatment strains taken from the same individuals showed the same numbers of repeats (7–9 in 6 cases and 11 in one). Conclusion: Our results suggest that the 6–9 repetitions of the 42-bp unit, with presumed stability, may offer these virus strains an advantage in virulence to human skin.

Extracellular dopamine (DA) levels in the steady state were not detected in the hippocampus of rats where sparse dopaminergic innervations were demonstrated. However, following L-DOPA (100 mg/kg i.p.) administration, extracellular DA was detected. Although tetrodotoxin (TTX, 1 μM) failed to prevent this phenomenon, the striatal DA peak disappeared with the addition of TTX (5 μM) into the perfusate and reappeared following L-DOPA (100 mg/kg i.p.) administration. These findings suggest that DA synthesis from L-DOPA, administered exogenously, is independent of the existence of dopaminergic neurons and that outflow of newly synthesized DA into the extracellular space is not coupled to neuronal activity.

A case of a 50-year-old man with recurrent herpes genitalis with two different isolated strains of herpes simplex virus (HSV) type 2 is reported. Morphologically, two types of cytopathic effects (CPE) induced by viruses were observed in the Vero cell cultures, one being a syncytial giant cell formation and the other a rounded cell formation without cell adhesion resembling the CPE induced by ordinary HSV type 2. In this case, it is clinically interesting that the patient complained of persistent pain on the glans and urethra without recurrence of herpetic lesions even after the involution of enanthema. The correlation between the persistent pain and neurovirulence of the two different HSV strains is discussed.

Estimation of LDL-chol and LDL-apo B is useful for the diagnosis of hyperapobetalipoproteinemia (normal LDL-chol with increased LDL-apo B), which is one of the most commonly occurring lipoprotein disorders associated with atherosclerotic cardiovascular diseases. The LDL-chol/LDL-apo B ratio reflects the level of small dense LDL, which is an important risk factor for IHD, CVD and ASO. In order to estimate LDL-apo B and LDL-chol/LDL-apo B ratio from blood chol, TG, HDL-chol and apo B values, we developed a formula for LDL-chol ¿0.94Chol- 0.94HDL-chol - 0.19TG¿, LDL-apo B ¿apo B - 0.09Chol + 0.09HDL-chol-0.08TG¿, and LDL-chol/LDL-apo B [¿0.94Chol-0.94HDL-chol - 0.19TG¿/¿apo B - 0.09Chol + 0.09HDL-chol-0.08TG¿] using ultracentrifugal data from 2179 subjects. These were calculated by the least squares method on the assumption that a certain compositional relationship exists between Chol, TG and apo B in VLDL, IDL and LDL. Friedewald's formula for LDL-chol (Chol - HDL-chol - 0.2TG) includes IDL-chol, but the present new formula theoretically excludes IDL-chol. It suggests a better estimation for the correct LDL-chol. Estimated LDL-apo B is useful for the diagnosis of hyperapobetalipoproteinemia and detection of small dense LDL. Without performing ultracentrifuge, additional information is obtained for the quantitative and qualitative alteration of LDL, such as small dense LDL. The above formulae and a new classification of lipoproteinemia including apo B were applied to the analyses of lipoprotein profiles of subjects with cardiovascular diseases, which were compared with those in the general population. Hyperapobetalipoproteinemia with high TG was observed 2-3 times more frequently in subjects with CAD, MI and ASO than in the Suita population. Lower ratios of LDL-chol/LDL-apo B, reflecting preponderance of small dense LDL, were observed in the above three groups. Type IIb and combined low HDL-chol were also frequent phenotypes in CAD, A-Th and ASO. The present formulae are useful for the detailed analyses of lipoprotein disorders in both qualitative as well as quantitative aspects.

OBJECTIVES: Microwaves produce various detrimental changes based on actions of heat or non-specific stress, although the effects of microwaves on pregnant organisms has not been uniform. This study was designed to clarify the effect of exposure to microwaves during pregnancy on endocrine and immune functions. METHODS: Natural killer cell activity and natural killer cell subsets in the spleen were measured, as well as some endocrine indicators in blood--corticosterone and adrenocorticotrophic hormone (ACTH) as indices of the hypothalamic-pituitary-adrenal axis--beta-endorphin, oestradiol, and progesterone in six female virgin rats and six pregnant rats (nine to 11 days gestation) exposed to microwaves at 10 mW/cm2 incident power density at 2450 MHz for 90 minutes. The same measurements were performed in control rats (six virgin and six pregnant rats). RESULTS: Skin temperature in virgin and pregnant rats increased immediately after exposure to microwaves. Although splenic activity of natural killer cells and any of the subset populations identified by the monoclonal antibodies CD16 and CD57 did not differ in virgin rats with or without exposure to microwaves, pregnant rats exposed to microwaves showed a significant reduction of splenic activity of natural killer cells and CD16+CD57-. Although corticosterone and ACTH increased, and oestradiol decreased in exposed virgin and pregnant rats, microwaves produced significant increases in beta-endorphin and progesterone only in pregnant rats. CONCLUSIONS: Microwaves at the power of 10 mW/cm2 produced activation of the hypothalamic-pituitary-adrenal axis and increased oestradiol in both virgin and pregnant rats, suggesting that microwaves greatly stress pregnant organisms. These findings in pregnant rats suggest that--with exposure to microwaves--pregnancy induces immunosuppression, which could result in successful maintainance of pregnancy. This enhancement of adaptability to heat stress with pregnancy may be mediated by activation of placental progesterone and placental or pituitary beta-endorphin.

To clarify the involvement of pregnancy in the response of the neuroendocrine-immune system to stress, we examined splenic natural killer-cell-activity-(NKCA) and its relevant central and blood parameters in female virgin and pregnant rats (10 to 11 days gestation) exposed to forced water-immersion stress with durations of 90 min and 180 min. Decreases in splenic NKCA, corticotropin releasing hormone (CRH) in the hypothalamus, and increases in progesterone (P), beta-endorphin (beta EP), and dopamine (DA) metabolic ratios in the frontal cortex and nucleus accumbens produced by stress were recognized in the virgin rats, but not in the pregnant rats. Pregnancy reduced splenic NKCA in rats without stress, but elevated it in the rats exposed to stress with a duration of 180 min. These findings suggest inhibitory effects of pregnancy on stress-induced immunosuppression and neuroendocrine changes, thereby promoting homeostasis in the neuroendocrine-immune system against stress. Such enhanced homeostasis associated with pregnancy seemed to be mediated by the activation of placental P and placental or pituitary beta EP in cooperation with mesocortical and mesolimbic DA systems and hypothalamic CRH.

Three children had systemic immediate-type reactions to Japanese encephalitis (JE) vaccine which contains gelatin as a stabilizer. We found that all the children had anti-gelatin IgE in their sera. Further, two of the three also had experienced systemic allergic reactions to gelatin-containing foods before the vaccination. We assume that the systemic allergic reactions to JE vaccines might be caused by gelatin in the JE vaccine.

Detection of varicella-zoster virus (VZV) DNA was carried out in the conjunctivas of 18 patients with herpes zoster by polymerase chain reaction. The rate of VZV DNA detection in conjunctivas was 3/4 in herpes zoster ophthalmicus associated with eruption on the dorsum nasi, 2/5 in herpes zoster ophthalmicus without eruption on the dorsum nasi, 1/2 in herpes zoster with eruption on the cheek and lower jaw, 1/2 in generalized herpes zoster and 0/5 in herpes zoster associated with eruption on the trunk or extremities, respectively. The reason for the detection of VZV DNA in conjunctivas is discussed.

A case of chromomycosis in which hyperthermia proved effective is reported. The patient was a 56-year-old male bean curd maker who, without any previous history of minor trauma, developed on the extensor side of the left upper arm an eczematous lesion that underwent gradual radial expansion. The lesion showed a well-defined, 7 x 10 cm infiltrated erythematous plaque with the central area healed and, at the upper and lower borders, adherent scales and crusts on the surface. Histological examination revealed granulomatous changes in the dermis, as well as sclerotic cells within giant cells and microabscesses. On culturing, Fonsecaea pedrosoi was isolated. The patient was treated with disposable chemical pocket warmers, which were secured over the lesion with a rather tight elastic bandage, so that they kept the affected area warm for 24 hours a day. After a month of such hyperthermic treatment, the erythema and infiltration had decreased considerably, and microscopic examination and culture of the crusts both yielded negative results. Examination of biopsy specimens of the lesion after the third month showed that it had cicatrized. The treatment was stopped after 4 months, and no relapse occurred. We also summarize the published results of local hyperthermic treatment of chromomycosis in Japan.

Transforming growth factor-beta 1 (TGF-beta 1) has been shown to induce squamous differentiation of cultured airway epithelial cells. It has also been shown to increase expression of matrix proteins and integrin receptors in cell culture of these and other cells. However, it is unknown if TGF-beta 1 affects expression of genes encoding intercellular junctional proteins. Therefore, we have investigated the effect of TGF-beta 1 on the expression of proteins and mRNAs for desmoplakins (DPs) I and II, desmosomal plaque proteins. Fibronectin, known to be induced by TGF-beta 1 was used as a positive control and tubulin as a negative control. Twenty-four hours after TGF-beta 1 stimulation, DP I and II mRNA levels assessed by Northern blotting analysis had increased significantly (DP I mRNA, 1.8-fold, P less than 0.05; DP II mRNA, 2.4-fold, P less than 0.04), thereby indicating pretranslational regulation of DP expression. By comparison, mRNA for fibronectin increased 8.1-fold whereas mRNA for tubulin was unchanged. Immunofluorescence using the monoclonal anti-DP I and II antibodies revealed dramatic increased expression of punctate DP structures after exposure to TGF-beta 1. Immunoblot analyses with polyclonal anti-DP I antibodies showed increased levels of both DP I (250 kD) and DP II (215 kD), with the DP I increase being more pronounced (DP I, 2.5-fold; DP II, 1.4-fold at 48 h relative to controls), suggesting translational regulation by TGF-beta 1. This study therefore demonstrates the ability of TGF-beta 1 to alter cellular phenotype by altering expression of proteins involved in intercellular junctions.(ABSTRACT TRUNCATED AT 250 WORDS)

We analyzed the differentiation of two strains isolated from the conjunctiva and rhinorrhea of a patient with herpetic keratitis by the restriction endonuclease digestion method of herpes simplex virus (HSV) DNA. As a result two strains were identified as the same one. This result suggests that HSV contained in tears flows into the nasal cavity via the lacrimal canaliculi.

A 26-year-old woman with erythema infectiosum is reported. She had small faint erythemas on her extremities and congestion of palpebral and bulbar conjunctiva of both eyes. Human parvovirus B19 (B19) DNA was detected in the specimen of the conjunctiva by the polymerase chain reaction. This result suggests that conjunctivitis in erythema infectiosum may cause the B19 infection.

In the goldfish brain, eleven groups of tyrosine hydroxylase (TH) -positive (catecholaminergic neurons and four groups of serotonin (5-HT) -positive (serotonergic) neurons are described as follows by immunofluorescence and light microscopic level of PAP technique: 1) The olfactory bulb (TH); 2) The area ventralis telencephali (TH); 3) The nucleus preopticus (TH); 4) The nucleus dorsolateralis thalami (TH); a) The pretectal area (5-HT); 5) The nucleus ventromedialis thalami (TH); 6) The nucleus posterioris periventricularis (PVO-accompanying cells) (TH); b) The nucleus recessus lateralis (NRL) (5-HT); 7) The rostro-medial part of the nucleus recessus lateralis (NRL) or the paraventricular organ (PVO) pars anterior (TH); b) PVO pars anterior (5-HT); 8) The nucleus posterior tuberis (NPT) (TH); c) The nucleus recessus posterioris (NRP) (5-HT); 9) The lateral isthmal region (TH); d) The raphe nuclei of the brainstem (5-HT); 10) The reticular formation of the medulla oblongata (TH); and 11) The post-obecular region (TH).By immunoelectron microscopy, two kinds of cerebrospinal fluid (CSF) -contacting neurons were confirmed in the PVO pars anterior. One contained TH-positive granules (80-120 nm) and the other 5-HT-immunoreactive granules (100-200 nm in diameter). In the telencephalon, preoptic nucleus (PON) and nucleus posterior tuberis (NPT), TH-positive non-CSF-contacting neurons showed similar ultrastructure such as well developed rough surfaced endoplasmic reticulum, and immunoreactive granules (106-110 nm).

Monoaminergic neuron systems have been studied in frontal, sagittal and horizontal sections of the bat brain by the indirect immunofluorescent method using antibodies to tyrosine hydroxylase (TH), dopamine-β-hydroxylase (DBH) and serotonin (5-HT).In general, the topographical distribution of monoaminergic neurons was found to be similar to that of the rat by Dahlstrom & Fuxe (3). However, some differences were found in the bat brain; namely the A3 group in the nucleus olivaris accessorius dorsalis and the A4 in the lateral part of the roof of the fourth ventricle were not observed.The 5-HT-immunofluorescent varicose fibers and terminals were widely spread throughout the brain, especially in the supra-ependymal parts of the ventricular walls, several cranial nerve nuclei such as V, VII, and Xth nerve nuclei, and the catecholaminergic cell groups (A1, A2, A6, A9, A10). In addition, both the dopaminergic neurons and 5-HT neurons were observed in the nucleus raphe dorsalis and the ventral tegmental area caudal to the interpeduncular nucleus.

By the immunofluorescence method, development of the cells containing catecholamine-synthesizing enzymes [tyrosine hydroxylase (TH), dopamine-β-hydroxylase (DBH) and phenylethanolamine-N-methyltransferase (PNMT)], serotonin (5-HT), calcitonin (Cal), somatostatin (Som), β-endorphin (End) and adrenocorticotropin (ACTH) was examined in the pituitary and tubero-infundibulum of Rana catesbeiana tadpoles, and the localization was compared with that of adult bullfrogs.A large number of End- and ACTH-like immunoreactive cells were observed in the rostral part of the pars distalis of the sadpole pituitary before metamorphosis (stage 31 by Gosner). At this time, 5-HT-positive cells were also observed throughout the pars distalis, though far less than those of the adult frog. TH-positive cells first appeared at Gosner's stage 40 together with the appearance of hindlimbs, but these cells were DBH-negative. The cells reacting with antiserum against DBH appeared after the resorption of the tail.TH-positive, DBH-negative dopaminergic fibers were observed around the capillary sinusoid of the pars distalis at Gosner's stage 31, but at stage 40 these fibers disappeared.TH-, 5-HT-, Cal-, Som- and End-immunoreactive neurons in the hypothalamus, and their fibers which may be derived from this brain area, in the neural lobe and median eminence, were already observed in the tadpole before metamorphosis (stage 31), as those in the adult frog.

Brains of the golden hamster were examined by immunofluorescent and peroxidase-antiperoxidase (PAP) methods using antibody to tyrosine hydroxylase (TH) or serotonin (5-HT).The localization of monoaminergic nerve cell bodies in the brain of the golden hamster was found to be similar to that of the rat by histofluorescent and immunofluorescent methods. Monoaminergic nerve fibers and terminals were widely distributed throughout the brain. Especially on the supraependymal part of the ventricle, 5-HT-positive varicose fibers were visualized, but a few TH-positive nerve processes in the ventromedial periventricular hypothalamus and in the caudal portion of the locus coeruleus were observed to penetrate into the ventricular lumen.By immunocytochemistry, TH- or 5-HT-positive reaction products were found in the cytoplasm except for cisternae of Golgi apparatus and mitochondria of neurons of Al cell group or raphe dorsalis. TH- or 5-HT-immunoreactive granules were occasionally found in each neuron, and their diameters were 100-140nm for TH or 110-120nm for 5-HT, respectively. Granules contained in nerve fibers were slightly smaller than those contained in nerve cell bodies.

Immunohisto-and immunocytochemical localizations of tyrosine hydroxylase (TH) were studied in the area postrema of hamster brain. By immunohistochemistry, TH-positive neurons with thick, smooth nerve processes were seen to be situated in close apposition to the blood vessels, and they were of oval-shaped, almost unipolar types.By immunocytochemistry, TH-positive reaction products were observed within the cytoplasm and granular vesicles but not in either the nucleus nor Golgi area of the TH-positive neurons. From the size of the granular vesicles, TH-positive cells in the area postrema were divided into two types of neurons, cells containing small granular vesicles (about 100 nm in diameter), and cells containing both small and large granular vesicles (larger than 150 nm in diameter). Small numbers of neurons with TH-negative reaction, were also found. These TH-negative neurons were agranular.

The present study revealed existence of a new dopaminergic nerve terminal plexus in the rat spinal cord, which was visualized by means of the peroxidase-antiperoxidase immunohistochemical technique utilizing antibody prepared against conjugated dopamine. Dopamine (DA)-immunoreactive nerve terminals occurred along the ventral motor column throughout all spinal levels, as well as previously noted presumed dopaminergic nerve terminals within the intermediolateral cell column, the dorsal horn and in the area surrounding the central canal. In these new terminals of the anterior column, fine DA-immunoreactive nerve terminals concentrated around motoneurons, suggesting that the dopaminergic nerve system may also be involved in somatic motor processes of the spinal cord.

Intracellular recordings were made from neurons in rabbit vesical pelvic ganglia (VPG), in vitro. Increasing the frequency of preganglionic-nerve stimulations from 0.1–1 Hz to 10–20 Hz facilitated fast excitatory postsynaptic potentials (EPSPs), resulting in a generation of action potentials. Acetylcholine-induced response was not altered during the facilitation of the fast EPSP. Serotonin blocked action potentials elicited by preganglionic-nerve stimulations at 0.1 Hz, while it caused no blockade at 10 Hz. Serotonin may potentiate the feature of high-pass filter [1] in transmission of VPG. Immunohistochemical study demonstrated serotonin-like varicose terminals in rabbit VPG.

臨床的に単純ヘルペスと診断した30症例を対象とし, ヘルペス病巣から採取した材料についてウイルス特異抗原の検出とウイルス分離を試み, その結果を発疹の形態別および病日別に比較検討し, つぎの結果を得た。 1) ウイルス特異抗原の検出法とウイルス分離法は皮膚単純ヘルペスのウイルス学的診断法として同程度に有用であつた。 2) ヘルペス発症後早期の水疱や膿疱を材料とした場合, ウイルス特異抗原の検出率が高く, この方式で迅速にウイルス学的診断を下すことが可能であつた。 3) 痂皮を材料とした場合では, 両方法を試みておくと, ひとつの方法が陰性の場合でも他の方法でウイルス学的診断が下せた。

Monoaminergic neurons of the brain of bullfrog were studied by means of immunohistocytochemistry using antibodies to catecholamine-synthesizing enzymes and serotonin (5-HT).Monoaminergic nerve cells were similarly localized as those reported by previous histofluorescence studies, but dopaminergic neurons were for the first time identified in the pretectal area. In the preoptic recess organ, a small number of 5-HT-containing liquor contacting neurons were found to coexist with a large number of dopaminergic ones. Neurons situated in and around the nucleus tractus solitarius were partly phenylethanolamine-N-methyltransferase-positive, supporting adrenaline to be of importance for the function of the frog brain. Moreover, the liquor contacting neurons of the paraventricular organ or nucleus infundibularis dorsalis and of the floor of the fourth ventricle were recognized as serotonergic and noradrenergic in nature, respectively.The ultrastructural study has shown that tyrosine hydroxylase (TH)- and 5-HT-positive reaction products were found in the ground substance and granules. The size of TH- or 5-HT-positive granules was variable from 90-200nm. Many differences between the intrinsic neurons in the brain parenchyma and the liquor contacting neurons were found in regard to the ultrastructure such as the number of immunoreactive granules and the nucleo-cytoplasmic ratio.

An antiserum was raised in a rabbit by immunization with taurine bound to bovine serum albumin. The antibody was purified by passage over an immunoabsorbent column (formyl-cellulofine-taurine) and it did not cross-react significantly with glutamate, aspartate, glycine, GABA (0.4%), glutamine, proline, cysteine, β-alanine, cysteic acid, carnosine or homocarnosine in enzyme-linked immunosorbent assays and nitrocellulose paper immunoblots. Immunocytochemical studies employing the peroxidase-antiperoxidase immunohistochemical technique revealed that many cerebellar Purkinje cells showed taurine-like immunoreactivity. Labelled axons could be followed within the white matter up to the deep cerebellar nuclei, where numerous puncta were observed. Immunoelectron microscopic examination revealed that labelled puncta were presynaptic terminals, and axo-dendritic or axo-somatic symmetrical synapses were observed on deep cerebellar nuclear neurons.

Antisera raised against glutamate or aspartate bound to bovine serum albumin were purified by affinity chromatography, and their specificities were verified by immunoblotting and by enzyme-linked immunosorbent assay. Immunohistochemical investigation using materials perfusion-fixed after long flushing demonstrated distinct laminar terminals with glutamate- or aspartate-like immunoreactivity throughout the limbic structures. This technique may offer a valuable tool for revealing the distribution of glutamatergic or aspartatergic nerve terminals.

Using a specific antibody against each of the 4 catecholamine-synthesizing enzymes, dopamine or serotonin, we compared the localization of these substances in rat and mouse spinal cord with two kinds of fixative, such as paraformaldehyde and glutaraldehyde, and two types of sectioning technique, i.e. cryostat and microslicer. In this study, we demonstrated aromatic L-amino acid decarboxylase (AADC)-like immunoreactivity (LI) in cerebrospinal fluid (CSF)-contacting neurons for the first time in the mouse. These cells and some AADC-immunoreactive cells localized outside the subependymal layer of mouse and rat spinal cord did not show tyrosine hydroxylase-, serotonin- and dopamine-LI with and without colchicine treatment.