Serological and genetic characterisation of a unique strain of adenovirus involved in an outbreak of epidemic keratoconjunctivitis

AIMS: To characterise a novel strain of adenovirus (Ad) type Ad8 (genome type Ad8I) involved in an epidemic keratoconjunctivitis (EKC) outbreak in Hiroshima city using serological testing and sequence analysis of the fibre and hexon gene. METHODS: A neutralisation test (NT) was performed in microtitre plates containing a confluent monolayer of A549 cells using 100 tissue culture infectious doses of virus and type specific antisera. The haemagglutination inhibition test was also carried out in microtitre plates with rat erythrocytes using four haemagglutination units of virus and twofold dilutions of serum. The fibre gene was sequenced by generating overlapping polymerase chain reaction products or by direct sequencing of genomic DNA. Primer selection was based on alignment of the fibre genes of human adenovirus serotypes Ad8, Ad19, Ad37, Ad9, and Ad15 available from Gene Bank. RESULTS: The virus strain was specifically neutralised by anti-Ad8 antibodies, although there was a major crossreaction with anti-Ad9 antibodies. Haemagglutination was equally inhibited by anti-Ad8 and anti-Ad9 antibodies. The predicted amino acid sequences of the hypervariable regions (HVRs) of the Ad8I hexon gene showed higher homology with Ad9 (83.3%) than with Ad8 (62.0%). However, the Ad8I fibre knob was more homologous to Ad8 (94.4%) than to Ad9 (91.6%). CONCLUSIONS: Ad8I is a unique strain of adenovirus because of its lower genomic homology with Ad8, major crossreactivity with Ad9 in NT, and mixed genetic organisation of HVRs of the hexon gene. These factors may have enabled the virus to circumvent acquired immunity, resulting in the outbreak.     

Synergistic effect of lymphokines and lipopolysaccharides on macrophage chemiluminescence. Appearance of spontaneous chemiluminescence and its correlation with cytotoxicity

A new type of chemiluminescence (CL) was observed in thioglycolate-elicited ICR mouse peritoneal macrophages which had been incubated for 24 h in medium containing lymphokines (LK) and lipopolysaccharides (LPS), and then exposed to a CL reagent solution containing luminol and phorbol myristate acetate (PMA). The new CL, which was clearly distinguishable from PMA-induced CL, appeared immediately after the addition of the CL reagent solution, reaching a maximal level at about 30 s, and disappeared rapidly. We have called the new CL 'spontaneous CL', since it appeared spontaneously without PMA triggering. When the macrophages were incubated with LK in the presence of 10 ng/ml of LPS, the spontaneous CL began to appear after about 4 h incubation, the maximal level being reached at about 12 h, after which it decreased gradually on further incubation. After 48 h, it could not be observed at all. The spontaneous CL could be observed only in macrophages simultaneously treated with LK and LPS, as in the case of cytotoxicity. The correlation between spontaneous CL and cytotoxicity was suggested by the following; they showed a similar dose dependency to LK, and neither of them could be induced in aged macrophages which had been incubated in vitro for 1 day before exposure to LK and LPS. These results suggest that spontaneous CL measurement could possibly replace the cytotoxicity test as a simple method for the determination of macrophage activating factor.     

Effects of drug adherence on patient outcomes to early treatment for Japanese cedar pollinosis]

Drug adherence is one of the important aspects in caring for patients with allergic rhinitis. To improve clinical efficacy of early treatment for Japanese cedar pollinosis (JCP), we evaluated the effect of drug adherence on patients' outcomes. Patients were randomly selected from 16 ENT clinical sites in Osaka and Wakayama between February 24 and March 8, 2003 (peak pollen season). Efficacy was assessed using patients' ratings of nasal and ocular symptoms and overall assessment in their condition compared with previous season ones. Costs include direct costs of the drugs used for treatment to JCP from January to February. Five hundred one patients taking early treatment were enrolled. Compared to low adherence patients, those who reported higher level of adherence significantly improved overall health condition, and achieved better symptom relief of rhinorrhea and nasal congestion. In multiple logistic regression analysis, the following factors were independent risk factors for low adherence: student (p=0.002), using OTC medications (p=0.006), and short-duration of medication (p=0.001). Low costs were also risk factor for low adherence. We conclude that taking medications for JCP for 22-28 days is the best way to enhance patients' outcomes.     

Development of T cells in SCID mice grafted with fetal thymus from AKR mice or F344 rats

To examine the development of T cells within an allogeneic or xenogeneic environment, we engrafted the fetal thymus from AKR mice or F344 rats under the kidney capsule of SCID mice (mTG and rTG mice). T lymphopoiesis developed in SCID mice 2 months after transplantation, although the ratio of CD4/CD8 in both experimental groups was different from that of normal control. T cells in mTG mice did not show in vitro proliferation or cytotoxicity against either host-type C.B-17 (H-2^d) or donor-type AKR (H-2^k) cells, while they exerted potent activities against third-party BIO (H-2^b) cells. In contrast, T cells in rTG mice exhibited proliferation against both host-type C.B-17 and donor-type F344 rat cells. Consistently, graft-vs.-host disease symptoms developed in these mice and histological examination showed impressive infiltration of lymphocytes into the skin or into the mucosal layers of the stomach. Activated state of T cells in rTG mice was also evidenced by the positive expression of interleukin-2 receptor. Taken together, fetal thymus appears to contain progenitor cells which are sufficient for in vivo reconstitution of T lymphopoiesis, but species-specific environment is important for the induction of tolerance. In mTG mice, Vβ6⁺ T cells reactive to donor Mls^a determinants and Vβ3⁺ T cells reactive to host Mls^c determinants were deleted, suggesting that tolerance was regulated mainly by clonal deletion. By contrast, Vβ11⁺ T cells reactive to Mls^f determinants were not deleted possibly due to the lack of their ligands.     

Serum valproate concentrations in epileptic children with favourable responses

The lower limits of the "therapeutic range" for serum levels of sodium valproate (VPA) were evaluated in epileptic children showing a benign clinical course. Twenty-five outpatients, aged 5 to 16 years, whose seizures were well controlled over three years with VPA alone, were studied. Venous blood was taken 1.1 to 5 hrs after the morning dose. Serum VPA concentrations at steady-state after receiving the maintenance doses to control seizures were determined by enzyme immunoassay. The patients were divided into three groups according to the seizure type and the age at onset; A and B: patients with tonic and/or clonic seizures, aged below 3 yrs (n = 11) and 3 to 11 yrs (n = 6), respectively, C: those with absence seizures, aged 4 to 11 yrs (n = 8). The serum concentrations in A (47.8 +/- 21.6 micrograms/ml, mean +/- SD) were significantly (p less than 0.02) lower than those in groups B and C (85.2 +/- 14.0 and 73.0 +/- 17.4 micrograms/ml, respectively). VPA concentrations below 50 micrograms/ml were seen in 6 patients (55%) in group A. It was concluded that many epileptic children, whose ages at onset were below 3 yrs, with tonic and/or clonic seizures could be controlled even with low initial serum concentrations below the "therapeutic range".     

Long-term follow-up of an individual with vitamin B6-dependent seizures

We report on a 31-year-old female with vitamin B6-dependent seizures whose seizure onset was in the neonatal period. Her elder brother had the same disorder and died in infancy. Administration of vitamin B6 was initiated in the postnatal period. At the age of 12 years 1 month, 2 months after withdrawal of vitamin B6, visual seizures began to occur frequently. Myoclonic seizures and occasional generalized convulsive seizures were also observed. At the same time, photoparoxysmal response and spontaneous diffuse spike-wave bursts were seen on her EEG. Myoclonic seizures were provoked by intermittent photic stimulation during the EEG. It is distinctive that visual seizures were one of the main seizure types in this patient, that her clinical course was relatively benign, and that she has normal intellectual outcome.     

Comparison of cyclosporin A and tacrolimus concentrations in whole blood between jejunal and ileal transplanted rats.

Most immunosuppresive drugs are absorbed from the intestine after oral administration, although there is some difference of bioavailability between ileum and jejunum. Using an orthotopic segmental small bowel transplantation (SBT) model in rats, we studied the pharmacokinetic profiles of cyclosporin A and tacrolimus concentrations after oral intake, comparing jejunal and ileal transplanted rats. Two types of segmental SBT (jejunal and ileal SBT) in a syngeneic combination were performed. After oral administration of cyclosporin A (10mgkg(-1) or tacrolimus (5 mg kg(-1)), pharmacokinetic data were obtained from the long-surviving rats transplanted with segmental SBT. To determine the effect of additional bile on cyclosporin absorption, an emulsion of cyclosporin A with fresh bile juice was re-challenged on segmental SBT rats before killing. A histological study was also performed by use of the intestinal grafts from the killed SBT rats. A higher concentration of cyclosporin A was observed in the ileum-grafted rats than in the rats which received the jejunal grafts. Oral bioavailability of cyclosporin A in ileal SBT rats tended to be increased by addition of fresh bile juice, but that in jejunal SBT rats did not change. On the other hand, there was no significant difference of tacrolimus concentration between jejunum- and ileum-transplanted rats. Histological studies showed that the superficial mucosal layer of both grafts, but especially the ileal graft, was markedly elongated compared with that of normal intestine. The present study showed that cyclosporin A was more actively absorbed from ileum than from jejunum in SBT, but tacrolimus was absorbed equally from both sites. These data suggest that cyclosporin A concentration is satisfactorily controlled in the segmental ileal graft, while there is no difference of tacrolimus absorption between ileal and jejunal graft.     

Pelvic insufficiency fractures in postmenopausal woman with advanced cervical cancer treated by radiotherapy.

PURPOSE: To assess the predisposing factors and clinical characteristics of pelvic insufficiency fractures (PIF) in postmenopausal women with pelvic irradiation. MATERIAL AND METHODS: A total 335 postmenopausal patients with cervical cancer of the intact uterus treated with radiation therapy between 1983 and 1998 were reviewed. Total external dose was delivered between 45 and 50.4 Gy with parallel opposed anteroposterior portals. Total brachytherapy dose at point A was delivered between 10 and 36 Gy. PIF were diagnosed by bone scintigraphy and confirmed by computed tomography. The cumulative incidence of symptomatic PIF was estimated by actuarial methods. Potential risk factors (age, weight, type II diabetes, delivery, menopause, total external dose, total brachytherapy dose) were assessed. RESULTS: Fifty-seven (17.0%) of 335 patients were diagnosed as having PIF. Forty-seven patients were symptomatic and ten were asymptomatic. Parameters carrying a significant association with PIF were body weight 49 kg or below (P=0.044) in stepwise logistic regression analysis. The cumulative incidence of symptomatic PIF at 5 years was 17.9% calculated by the Kaplan-Meier method. A body weight of 49 kg or below and more than three deliveries were identified as having a significant effect on symptomatic PIF in univariate analysis (P=0.021, P=0.003, log-rank test) and Cox life table regression analysis (P=0.038, P=0.013). Five patients required narcotic agents and eight patients required hospital admission. CONCLUSIONS: We should consider reducing the dose contribution to the sacrum and sacroilac joints, without underdosing the tumor, especially in postmenopausal women with many deliveries or low body weight.     

Subjects with <Emphasis Type="Italic">TNF-A-857TT</Emphasis> and <Emphasis Type="Italic">-1031TT</Emphasis> genotypes showed the highest <Emphasis Type="Italic">Helicobacter pylori</Emphasis> seropositive rate compared with those with other genotypes

Background A possible association between Helicobacter pylori seropositivity and tumor necrosis factor (TNF) A G-308A has been reported in Korea. The present study examined the associations of H. pylori with functional polymorphisms, TNF-A G-308A, C-857T, and T-1031C, and TNF-B A252G in Japanese subjects.Methods The total of 1374 study subjects included 241 outpatients who participated in an H. pylori eradication program (HPE), 679 first-visit outpatients (FVO) at a regional cancer hospital, and 454 local residents who received a health checkup examination (HCE).Results The frequency of the TNF-A -308A allele was only 1.3% of 480 chromosomes in the HPE group, so the FVO and HCE groups were not genotyped for that polymorphism. The genotype frequency of TNF-A C-857T was 69.2% CC, 27.7% CT, and 3.1% TT; that of TNF-A T-1031C was 69.4% TT, 28.1% TC, and 2.5% CC; and that of TNF-B A252G was 36.8% AA, 48.2% AG, and 15.0% GG. TNF-A -857T was tightly linked to TNF-A -1031T and TNF-B 252A. No significant associations between H. pylori seropositivity and polymorphisms of TNF-A C-857T and TNF-B A252G were observed. However, a reduced odds ratio adjusted for sex, age, and recruitment source was observed for TNF-A -1031CC (0.43; 95% confidence interval, 0.20–0.91) relative to TNF-A -1031TT. Subjects with TNF-A -857CC and -1031CC showed the lowest seropositivity (38.2% of 34 participants), while those with TNF-A -857TT and -1031TT showed the highest (66.7% of 42 participants).Conclusions This study suggests that the possibly high expression genotype of TNF-A may increase susceptibility to persistent H. pylori infection.