持续性细胞皱缩在人上皮细胞凋亡过程中的必要性

持续性细胞皱缩是凋亡发生的一个主要标志。近期研究发现细胞皱缩在细胞凋亡过程中并不是一个被动的次要事件。在各种细胞中,包括人上皮细胞,凋亡因子（apoptogen）刺激后马上发生全细胞皱缩,又称为凋亡性容积减小（apoptotic　volumede　crease,AVD）,继而发生caspase激活、DNA片段化、细胞破裂死亡。K^＋和Cl^-通道的激活导致KCl外流,诱导AVD发生。抑制AVD发生可以抑制细胞凋亡。AVD与调节性容积增加（regulatory　volume　increase,RVI）异常相伴发生时,人上皮性HeLa细胞发生持续性细胞皱缩。RVI功能受损时,高渗本身就能诱导HeLa细胞持续性细胞皱缩,继而凋亡。即使在正常渗透压、无凋亡因子刺激的情况下,将HeLa细胞置于缺乏Na^＋或Cl。的溶液也会导致细胞持续性皱缩,继而凋亡。因此,AVD诱导和RVI异常所导致的持续性细胞皱缩是人上皮细胞发生凋亡的首要条件。     

A promoter haplotype of the interleukin-18 gene is associated with juvenile idiopathic arthritis in the Japanese population

Recently, we reported that genetic polymorphisms within the human IL18 gene were associated with disease susceptibility to adult-onset Still's disease (AOSD), which is characterized by extraordinarily high serum levels of IL-18. Because high serum IL-18 induction has also been observed in the systemic type of juvenile idiopathic arthritis (JIA), we investigated whether similar genetic skewing is present in this disease. Three haplotypes, S01, S02, and S03, composed of 13 genetic polymorphisms covering two distinct promoter regions, were determined for 33 JIA patients, including 17 with systemic JIA, 10 with polyarthritis, and 6 with oligoarthritis. Haplotypes were also analyzed for 28 AOSD patients, 164 rheumatoid arthritis (RA) patients, 102 patients with collagen diseases, and 173 healthy control subjects. The frequency of individuals carrying a diplotype configuration (a combination of two haplotypes) of S01/S01 was significantly higher in the JIA patients, including all subgroups, than in the healthy controls (P = 0.0045, Fischer exact probability test; odds ratio (OR) = 3.55, 95% confidence interval (CI) = 1.55–8.14). In patients with systemic JIA, its frequency did not differ statistically from that of normal controls. Nevertheless, it is possible that haplotype S01 is associated with the phenotype of high IL-18 production in systemic JIA because the patients carrying S01/S01 showed significantly higher serum IL-18 levels compared with patients with other diplotype configurations (P = 0.017, Mann-Whitney U test). We confirmed that the frequency of the diplotype configuration of S01/S01 was significantly higher in AOSD patients than in healthy control subjects (P = 0.011, OR = 3.45, 95% CI = 1.42–8.36). Furthermore, the RA patients were also more predisposed to have S01/S01 (P = 0.018, OR = 2.00, 95% CI = 1.14–3.50) than the healthy control subjects, whereas the patients with collagen diseases did not. In summary, the diplotype configuration of S01/S01 was associated with susceptibility to JIA as well as AOSD and RA, and linked to significantly higher IL-18 production in systemic JIA. Possession of the diplotype configuration of S01/S01 would be one of the genetic risk factors for susceptibility to arthritis in the Japanese population.     

Virus-like particles associated with mass mortalities of the pen shell Atrina pectinata in Japan

Mass mortalities of the pen shell Atrina pectinata occurred in the fishing grounds of Ariake Bay, in southwestern Japan, during late spring and summer in 2003 and 2004. Histological examination revealed extensive necrosis in the epithelial cells of the kidney and gill, and impairment of the endothelial cells of the mantle arteria. Although cestode larvae belonging to the genus Tylocephalum were found in the mantle, adductor muscle, kidney, and digestive gland, their prevalence and the intensity of infection were low. Examinations of moribund pen shells for Haplosporidium spp. infection using PCR analysis and for Perkinsus spp. infection using Ray's fluid thioglycollate medium were negative. Unenveloped virus-like particles were detected by transmission electron microscopy in the cytoplasm of affected kidney and gill cells of moribund pen shells. They were icosahedral spherical and 50 to 55 nm in diameter. These virus-like particles found in moribund pen shells are different from those described in other marine mollusks, and may be the causative agent of the mass mortalities of pen shells.     

Islet-like cell clusters: viability, cell types, and subretinal transplantation in pancreatectomized cats

We investigated a method for isolating sufficient feline islets of Langerhans to restore normoglycaemia following transplantation into the subretinal space of pancreatectomized cats. Collagenase digestate of feline pancreas was maintained in serum-free tissue culture medium for 1-9 days. Viability of islet-like cell clusters (ICC) was assessed with ethidium bromide and fluorescein diacetate staining; cell types were identified immunohistochemically. After nine days, the ICC were transplanted. We estimated viable ICC in tissue culture at nine days as 3800 +/- 2000 (mean +/- SD) per pancreas. While numbers of beta cells decreased over time in culture, ductal cells increased. Bromodeoxyuridine labelling showed no proliferation of beta cells but extensive proliferation of ductal cells. Subretinal transplants of cultured ICC in the diabetic cats maintained normoglycaemia for up to 12 days, while they provoked massive lymphocyte infiltration indicating rejection. Islet transplantation into the feline subretinal space temporarily restored normoglycaemia. Our current method of culture achieved sufficient reduction of acinar cells but an insufficient yield of insulin-producing cells.     

The effect of IL-1alpha on the expression of matrix metalloproteinases, plasminogen activators, and their inhibitors in osteoblastic ROS 17/2.8 cells

Interleukin-1 (IL-1) plays key roles in altering bone matrix turnover. This turnover is regulated by matrix metalloproteinases (MMPs), tissue inhibitor of matrix metalloproteinases (TIMPs), and the plasminogen activation system, including tissue-type plasminogen activator (tPA), urokinase-type plasminogen activator (uPA) , and plasminogen activator inhibitor type-1 (PAI-1). In this study, we examined the effect of IL-1alpha on the expression of the MMPs, TIMPs, tPA, uPA, and PAI-1 genes in osteoblasts derived from the rat osteosarcoma cell line ROS 17/2.8. The cells were cultured in alpha-minimum essential medium containing 10% fetal bovine serum with 0 or 100 U/ml of IL-1alpha for up to 14 days. The levels of MMPs, TIMPs, uPA, tPA, and PAI-1 expression were estimated by determining the mRNA levels using real-time RT-PCR and by determining protein levels using ELISA. In IL-1alpha cultures, the expression levels of MMP-1, -2, -3, -13, and -14 exceeded that of the control through day 14 of culture, and the expression of MMPs increased markedly from the proliferative to the later stages of culture. The TIMP-1, -2, and -3 expression levels increased from the initial to the proliferative stages of culture. The expression of tPA increased greatly during the proliferative stage of culture, and uPA expression increased throughout the culture period, increasing markedly from the proliferative to the later stages of culture. In contrast, PAI-1 expression decreased in the presence of IL-1alpha through day 14. These results suggest that IL-1alpha stimulate bone matrix turnover by increasing MMPs, tPA, and uPA production and decreasing PAI-1 production by osteoblasts, and incline the turnover to the resolution.     

Identification and expression of human epiglycanin/MUC21: a novel transmembrane mucin

The gene for the human orthologue of mouse epiglycanin, a mucin expressed on mammary carcinoma TA3-Ha cells but not TA3-St cells, was identified by homology search to a mouse epiglycanin cDNA fragment identified by representational difference analysis between TA3-Ha and TA3-St cells. The open reading frame of this gene was cloned from human cervical carcinoma ME-180 cells. It consists of a mucin domain with 28 nonidentical tandem repeats of 45 nucleotides each corresponding to a threonine/serine-rich peptide, a stem domain, a transmembrane domain, and a cytoplasmic tail. The cloned cDNA with a FLAG sequence was expressed in K562 cells. A combination of immunoprecipitation with a polyclonal antibody specific for the cytoplasmic tail and Western blotting analysis with an anti-FLAG antibody and lectins revealed a mucin-like component as the gene product. Analysis by the use of tissue cDNA libraries indicated that the gene is expressed in lung, large intestine, thymus, and testis among 16 normal tissues tested. The polyclonal antibody specific for a synthetic peptide from the cytoplasmic tail, when tested for its reactivity with normal lung tissues, reacted with epithelia of bronchi and bronchioli but not with alveoli. All of 24 lung adenocarcinomas specimens tested were reactive with the antibody, whereas reactivity was observed with only 2 out of 24 squamous and none out of 24 small cell lung carcinomas. This is a novel transmembrane mucin and designated as MUC21.     

Synthesis and structure-activity relationships of a series of substituted 2-(1H-furo[2,3-g]indazol-1-yl)ethylamine derivatives as 5-HT2C receptor agonists

A series of novel indazole derivatives were synthesized, and their structure-activity relationships examined in order to identify potent and selective 5-HT2C receptor agonists. Among these compounds, (S)-2-(7-ethyl-1H-furo[2,3-g]indazol-1-yl)-1-methylethylamine (YM348) had a good in vitro profile, that is, high agonistic activity to the human 5-HT2C receptor subtype (EC50 = 1.0 nM) and high selectivity over 5-HT2A receptors. This compound was also effective in a rat penile erection model when administered p.o.     

Phase-specific developmental and reproductive strategies in the desert locust

Locusts modify developmental and reproductive traits over successive generations depending on the population density. A trade-off between developmental rate and body size and between progeny size and number is often observed in organisms. In this study, we present evidence that this rule is evaded by desert locusts, Schistocerca gregaria Forsk?l, which often undergo outbreaks. Under isolated conditions, large hatchlings, typical of the gregarious forms, grow faster but emerge as larger adults than do small hatchlings typical of the solitarious forms, except for some individuals of the latter group that undergo extra molting. Under crowded conditions, large and small hatchlings grow at a similar rate, but the former become larger adults than the latter. Small hatchlings show a trade-off between development time and body size at maturation, but this constraint is avoided by large hatchlings. Phase-specific, as well as body size-dependent, differences are also detected in reproductive performance. As adult body size increases, females of a solitarious line produce more but slightly smaller eggs, whereas those of a gregarious line produce more and larger eggs. Total egg mass per pod is larger in gregarious forms than in solitarious forms. A trade-off between egg size and number is shown by a solitarious line but not by a gregarious line that produces relatively large eggs with similar numbers of eggs per pod. These results suggest that phase transformation involves not just a shift of resource allocation but also an enhanced capability expressed in response to crowding.     

Enhancement of Newcastle Disease Virus-Induced Fusion of Mouse L Cells by Sodium Vanadate

Sodium vanadate enhanced Newcastle disease virus (NDV)-induced cell fusion in L cells, and there was a direct correlation between the degree of cell fusion and the dose of vanadate added. When anti-F protein of NDV monospecific antiserum was added to the culture fluid of L cells infected with NDV, the enhancement of cell fusion was suppressed. In contrast, neither anti-HN nor anti-M protein monospecific antiserum inhibited the enhancement. Incubation at low temperature (4 C) and addition of sodium azide to the culture fluid suppressed the enhancement. The suppression by azide was seen only when the drug was added within 5 min after the beginning of incubation of NDV-infected L cells with vanadate. On the other hand, incubation at low temperature inhibited the enhancement at any time during incubation with vanadate. Cytochalasin D also inhibited the enhancement if it was added at any time during incubation with vanadate.