Towards application of water extract from heat-killed Lactococcus lactis H61 as a cosmetic ingredient.

We previously reported that oral administration of heat-killed Lactococcus lactis H61 improves certain human skin properties. For topical application of this strain, we reasoned that a bacterial cell extract obtained with an aqueous solvent could be readily formulated as a cosmetic ingredient. In the present study, we characterized the water extract from heat-killed H61. The extract had inhibitory activity for angiotensin-converting enzyme, which is known as suppression of inflammation of skin, and absorbed electromagnetic radiation in the UVB range. UVB-irradiated normal human epidermal keratinocytes (NHEKs) had lower viability than nonirradiated NHEKs. The NHEK survival rate was significantly higher in cells treated with the extract at 10 mg dried cells per ml prior to UVB exposure than in untreated cells or cells treated with lower extract concentrations. At this concentration, the extract also inhibited the production of interleukin-8 induced by UVB. The extract did not protect against hydrogen peroxide-induced cell damage. These data indicate that topical application of the H61 extract alleviates UVB damage and reduces inflammation in skin cells. The present study expands the potential application of strain H61 to its use as a cosmetic ingredient in addition to its use in the food industry. SIGNIFICANCE AND IMPACT OF THE STUDY: In our previous report, oral administration of heat-killed Lactococcus lactis H61 improved certain human skin properties. This study aimed exploring the potential topical use of this strain. The water extract derived from heat-killed cells with angiotensin-converting enzyme inhibitory activity, which is known as suppression of inflammation of skin, could protect normal human epidermal keratinocytes (NHEKs) from damage caused by UVB. Higher interleukin-8 production by UVB-exposed NHEKs than nontreated cells was suppressed by addition of the extract. The extract absorbed electromagnetic radiation in the UVB range. This extract could help in the maintenance of skin health by suppressing inflammation.

Effects of rare sugar D-allulose on acid production and probiotic activities of dairy lactic acid bacteria.

It has recently been reported that the rare sugar d-allulose has beneficial effects, including the suppression of postprandial blood glucose elevation in humans, and can be substituted for sucrose as a low-calorie food ingredient. To examine the applications of d-allulose in the dairy industry, we investigated the effects of d-allulose on the acid production of 8 strains of yogurt starter (Lactobacillus delbrueckii ssp. bulgaricus and Streptococcus thermophilus) and 4 strains of lactococci, including potential probiotic candidates derived from dairy products. Acid production by 2 L. delbrueckii ssp. bulgaricus yogurt starter strains in milk was suppressed by d-allulose, but this phenomenon was also observed in some strains with another sugar (xylose), a sugar alcohol (sorbitol), or both. In contrast, among the dairy probiotic candidates, Lactococcus lactis H61, which has beneficial effects for human skin when drunk as part of fermented milk, was the only strain that showed suppression of acid production in the presence of d-allulose. Strain H61 did not metabolize d-allulose. We did not observe suppression of acid production by strain H61 with the addition of xylose or sorbitol, and xylose and sorbitol were not metabolized by strain H61. The acid production of strain H61 after culture in a constituted medium (tryptone-yeast extract-glucose broth) was also suppressed with the addition of d-allulose, but growth efficiency and sugar fermentation style were not altered. Probiotic activities-such as the angiotensin-converting enzyme inhibitory activity of H61-fermented milk and the superoxide dismutase activity of H61 cells grown in tryptone-yeast extract-glucose broth-were not affected by d-allulose. d-Allulose may suppress acid production in certain lactic acid bacteria without altering their probiotic activity. It may be useful for developing new probiotic dairy products from probiotic strains such as Lactococcus lactis H61.

Different expression patterns of hepatic cytochrome P450 s during anaphylactic or lipopolysaccharide-induced inflammation.

Certain physiological states and diseases can alter the expression and activity of cytochrome P450 s (CYPs), which have the potential to cause unexpected adverse effects. We previously demonstrated that lipopolysaccharide (LPS)-induced inflammation attenuates the induction of CYPs by xenobiotics in mouse liver. In this study, to investigate whether anaphylaxis-induced inflammation affects the hepatic CYPs' expression, we examined the effects of ovalbumin (OVA)-induced anaphylaxis on constitutive CYP mRNA and protein expressions. We also compared these effects with those obtained with LPS treatment. In addition, we examined the tumor necrosis factor (TNF) alpha and interleukin (IL)-113 mRNA levels, because these cytokines are known to be induced by LPS treatment and anaphylactic reactions. LPS treatment decreased the constitutively expressed Cyp1a2, Cyp2c29, and Cyp3al 1 mRNAs, and increased the TNFalpha and IL-1beta mRNAs. LPS treatment also decreased the CYP1A2 and CYP3A protein levels. Anaphylaxis, on the other hand, did not change the levels of the constitutively expressed Cyp1a2, Cyp2c29, or Cyp3a1 1 mRNAs, although it increased the TNFalpha and IL-1beta mRNAs, as observed in the LPS-treated mice. These results suggest that anaphylaxis-induced inflammation had less effect than LPS-induced inflammation on these CYPs in the liver. In contrast, we observed that the expressions of Cyp2b10 mRNA and its protein were quite different from those of the other CYPs in both the anaphylactic and LPS-treated mice. Our findings strongly suggest that the alteration of the constitutive CYPs' expression levels during inflammation varies according to the immunostimulation pathway.

Resistance to cutaneous graft-vs.-host disease is not induced in T cell receptor delta gene-mutant mice.

The function of murine dendritic epidermal cells (dEC) remains largely speculative, probably because of the lack of a suitable in vivo model, although previous studies suggest that gamma/delta+ dEC may have originally evolved to serve as a self-protection mechanism(s). Our previous study demonstrated that the epidermis of mice that had spontaneously recovered from cutaneous graft-vs-host disease (GVHD) induced by local injection of CD4+ autoreactive T cells contained unexpectedly large numbers of dEC and became resistant to subsequent attempts to induce GVHD in a site-restricted manner, suggesting that the resistance is mediated by dEC. However, because alpha/beta+ dEC as well as gamma/delta+ dEC were greatly increased in number in the epidermis, it was unclear whether gamma/delta+ dEC are indeed responsible for this protection. The availability of this murine model and mice selectively lacking gamma/delta T cells as a result of disruption of the T cell receptor C delta gene segment allowed us to investigate the role of gamma/delta+ dEC. In the epidermis of gamma/delta T cell-deficient mice (delta-/-), a congenital lack of gamma/delta+ dEC was substituted for by alpha/beta+ dEC of either a CD4-8+ or a CD4-8- phenotype. After intradermal injection of the autoreactive T cells, delta-/- mice developed significantly enhanced delayed-type hypersensitivity responses and cutaneous GVHD, which persisted longer than in heterozygous littermate controls (delta+/-). Surprisingly, resistance to the cutaneous GVHD was not induced in the epidermis of delta-/- mice after spontaneous recovery from the GVHD, whereas the "susceptible" epidermis of delta-/+ mice contained large numbers of alpha/beta dEC comparable to those in "resistant" epidermis of delta+/- mice. Injection of day 16 fetal thymocytes from wild-type mice into delta-/- mice resulted in the appearance of donor-type gamma/delta+ dEC in the epidermis, and reconstitution with gamma/delta+ dEC restored the protective immune response of the epidermis against the GVHD to nearly normal levels. These results indicate that gamma/delta+ dEC are responsible for the site-restricted protection against cutaneous GVHD.

Loss of epidermal integrity by T cell-mediated attack induces long-term local resistance to subsequent attack. I. Induction of resistance correlates with increases in Thy-1+ epidermal cell numbers.

The cutaneous graft-versus-host disease (GVHD) lesions induced by intradermal injection of cloned autoreactive T cells have been shown to subside rapidly and the epidermis returns to normal 2 wk after injection. Those mice that had spontaneously recovered from the cutaneous GVHD became resistant to subsequent attempts to induce the cutaneous GVHD by the T cells while maintaining their activity to mount delayed-type hypersensitivity (DTH) responses and to induce the enlargement of the popliteal lymph nodes (PLN). The resistance appeared to be restricted to the epidermal structures of the injection sites, suggesting the involvement of locally acting suppression mechanisms. This local resistance was not specific for the clonotype used for the induction of the resistance. A loss of the epidermal integrity by an attack of T cells capable of producing cutaneous GVHD was a prerequisite for the induction of the resistance. By up to at least 8 mo after injection of the T cells, no mice became susceptible to the cutaneous GVHD again, provided that the T cells were injected into the same footpad sites that had initially received the T cells. This resistance correlated well with the great increase (20-30-fold) in Thy-1+ EC number. The great increase in the number of Thy-1+ EC following destruction of epidermal structures may be important in protecting the epidermal integrity from an additional attack by T cells.

Effects of fat-enriched diet and methionine on insulin sensitivity in lactating cows.

The hyperinsulinemic-euglycemic clamp (EGC) technique was used to investigate the effects of calcium salts of long-chain fatty acids (LCFA-Ca) and rumen-protected Met (RPM) on insulin sensitivity in the peripheral tissues of lactating cows. Six multiparous Holstein cows were used in a 3 × 3 Latin square experiment in each 14-d period. Dietary treatments were 0 (RPM0), 20 (RPM20), and 60 (RPM60) g/d of RPM, supplemented with a diet containing 1.5% of LCFA-Ca equal to 110% of the cows' ME requirement. And as a control for the 3 LCFA-Ca-containing diets, a dietary treatment without LCFA-Ca (Con) was also included. After a 10-d adaptation period, milk samples were collected for 4 d, and EGC experiments were performed on d 14 of each treatment period. Insulin solution was infused through a jugular vein catheter at a rate of 0.1, 0.2, 0.3, and 0.4 milliunits·kg BW-1·min-1 for 30 min and then at a rate of 0.5 milliunits·kg BW-1·min-1 for 60 min. Glucose solution was variably infused to maintain plasma glucose at steady state through the same catheter. Blood samples for measurements were taken using the contralateral catheter. Plasma total cholesterol, cholesterol ester, free cholesterol, and phospholipid concentrations in RPM0 and RPM20 were higher than those in Con, whereas the concentrations in RPM60 were low at the same degree of those in RPM0 (P < 0.05). Plasma Met concentration was greatest in RPM60 (P < 0.05). In the EGC experiment, the glucose infusion rate was greater in RPM60 than in RPM0 and RPM20 and an effective concentration of insulin resulting in 50% maximal glucose infusion rate was lower in RPM60 compared with RPM0 (P < 0.05), indicating that insulin sensitivity was intensified in RPM60. Although the insulin sensitivity evaluated from the EGC data in RPM0, RPM20, and RPM60 was not different from Con, a slight decline was observed in RPM0 and insulin sensitivity in RPM60 was higher than Con. Our results from the EGC experiment demonstrated that the feeding RPM lead to increased insulin sensitivity, which suggests that dietary Met affects lipid metabolism via insulin action in lactating dairy cows fed a LCFA-Ca-containing diet.

Epidermal T cells: their functional role and disease relevance for dermatologists.

T cells found within the epidermis in inflammatory dermatoses are generally accepted as making a major contribution to epidermal damage. On the other hand, those T cells residing in the murine epidermis are supposed to play an important role in protecting the epidermis from potentially dangerous immune reactions. Overwhelming evidence has accumulated that dendritic epidermal T cells (DETC) expressing monomorphic TCR gammadelta are responsible for the protection of epidermal structures against skin tumor, bacterial infection, and autoimmune attack. In animals congenitally lacking these gammadelta+ DETC, the epidermis is populated with bone marrow-derived TCR alphabeta+, CD8+ DETC. Although it remains unclear whether this subset of DETC could home to the epidermis to substitute for the physiologic function of gammadelta+ DETC or whether they would be pathologically relevant to epidermal injury, it should be noted that this subset represents the major fraction of T cells present in normal human epidermis and the most abundant in the lesional epidermis of fixed drug eruption (FDE). Because they are shown to kill target cells including keratinocytes upon stimulation and utilize a very limited range of TCR V alpha and Vbeta gene families, localized epidermal injury in FDE lesions would be mediated by activation of these epidermal T cells with autoaggressive potential. Epidermal T cells are thus likely to form several T-cell populations with different immunologic functions that are triggered by different modes of stimulation. Immune homeostasis in the epidermis would rely on a delicate balance between at least two types of epidermal T cells: autoaggressive T cells and protective T cells.

Lichenoid tissue reaction (LTR) induced by local transfer of Ia-reactive T-cell clones. II. LTR by epidermal invasion of cytotoxic lymphokine-producing autoreactive T cells.

Recent studies have suggested that T lymphocytes with helper phenotype may play an important role in the pathogenesis of lichenoid tissue reactions (LTR). In order to elucidate whether murine self-Ia-specific autoreactive T cells with helper phenotype can induce LTR in naive syngeneic hosts, two clones of autoreactive cloned T cells with both helper and cytotoxic activities, capable of producing the cytotoxic lymphokines gamma interferon and lymphotoxin in response to self-Ia antigens, were examined for their ability to induce LTR. One clone, BB5, when injected into footpads of syngeneic hosts, induced prominent epidermotropic cellular infiltrates that led to basal vacuolar degeneration and Civatte body formation; the other clone, C10, did not. No difference between BB5 and C10 was detected with respect to their ability to induce delayed-type hypersensitivity reactions. Immunohistochemical studies using anti-Lyt monoclonal antibodies showed that BB5-induced LTR was induced by the epidermal invasion of the injected BB5 cells themselves. We further observed that the infusion of BB5 cells not only resulted in Ia antigen expression on keratinocytes, but was also accompanied by a morphologic change in Langerhans cells at 24 h, when the epidermal invasion had not yet developed. However, similar observations were also observed with C10 cells, indicating that Ia expression on keratinocytes may not be a prerequisite for the induction of LTR.

In vitro migration of L3T4+ cloned T cells to epidermis: possible role for keratinocyte-derived factors.

Three types of L3T4+ cloned T cells with different antigen specificities, auto-, allo-, and antigen-reactive, were characterized with respect to their migratory potential using an in vitro migration assay under agar gel. Autoreactive T cells, BB5, and alloreactive T cells, SK 1, both of which have been proved to be epidermotropic in vivo, showed specific directional migration to the epidermis, whereas no directional migration was seen with non-epidermotropic cloned T cells and freshly isolated lymph node T cells. Both BB5 and SK 1 cells were equally attracted to all the epidermal fragments tested regardless of their I-A antigens. The directional migration of BB5 cells to the epidermis was significantly inhibited by the co-cultivation with the epidermis, but not the dermis. Studies with cell lines, the conditioned media (CM), and recombinant interleukin (IL) 1, 2, and 3 revealed that BB5 cells were chemotactically attracted to a transformed keratinocyte cell line PAM212 and, to a lesser extent, to the CM from PAM212 and IL-2, but not to IL-1 and IL 3. These results suggest that epidermotropic T cells may be preferentially trapped in an area with a high concentration of keratinocyte-derived growth factors as well as IL-2.

Lichenoid tissue reaction induced by local transfer of Ia-reactive T-cell clones. III. Role of Ia+ keratinocytes in the epidermotropic migration of the T cells.

The role of Ia+ keratinocytes in epidermotropism of T cells was analyzed by using self-IA specific autoreactive cloned T cells with epidermotropic nature (termed BB5) and those without it (termed C10). These T-cell clones were injected into the footpads of syngeneic nude mice whose keratinocytes had been induced to express Ia by the iv injection of normal mouse serum. Ia expression by keratinocytes was associated with the increased epidermal invasion of BB5 cells, but did not render C10 cells capable of migrating into the epidermis. The migration of the T cells to epidermis was also studied in vitro using a migration assay under agar. Ia expression by keratinocytes significantly enhanced the in vitro migration of BB5 cells to the epidermis, but had no effect on the migration of C10 cells and freshly isolated unstimulated lymphocytes. We surmise from these results that Ia+ keratinocytes may facilitate the epidermal invasion of the T cells with epidermotropic nature, but not those without it. However, the possibility certainly exists that the observed preferential migration of BB5 cells to Ia+ keratinocytes may be secondary to the alteration of other factors associated with the Ia expression. The injection of normal mouse serum was accompanied by an increase in the production of epidermal cell-derived thymocyte activating factor (ETAF) indicating that the increased ETAF production may have contributed to some of the observed preferential migration of BB5 cells.

Lichenoid tissue reaction induced by local transfer of Ia-reactive T-cell clones.

Epidermal basal cell damage in lichenoid tissue reactions (LTR) is considered to be the result of immunologic injury. In this study, we propose that LTR may be caused by local activation of Ia-reactive T cells. We have established allo-Iak-reactive helper T-cell clones and examined their behavior after adoptive transfer. We show that local transfer of 3 allo-Iak-reactive helper T-cell clones with different cross-reactivities and functions in vitro can cause delayed type hypersensitivity (DTH) reactions in vivo with antigen specificities identical to those demonstrated in vitro. Clone SK.1, when injected into appropriate recipients, caused massive dermal infiltrates of neutrophils and mononuclear cells. The latter were attracted to the epidermis and induced LTR-like basal cell degeneration which peaked at 72 h. Appropriate recipients were those strains of mice whose spleen cells were able to stimulate SK.1 cells to proliferate in vitro. Two other clones, SK.2.18 and SK.2.16, evoked significant DTH responses in their appropriate recipients, but the massive cellular infiltrates induced by either clone never invaded the epidermis or produced an LTR. The degeneration of epidermal cells caused by SK.1 cells did not correlate with the tested functions of this clone in vitro. The finding that only 1 of the 3 allo-Ia-reactive helper T-cell clones induced epidermotropic cellular infiltrates indicates that the infiltrative pattern of leukocytes in skin may depend on the particular T-cell clone that is activated.

Differential expression of lymphocyte function-associated antigen 1 (LFA-1) on epidermotropic and non-epidermotropic T-cell clones.

Various T-cell surface molecules are involved in T-cell adhesion, which is an essential requirement for epidermotropic migration of T cells. Our previous studies demonstrated that certain T-cell clones can migrate into the epidermis upon their intradermal inoculation into the footpads of recipient mice with relevant antigens, while other T-cell clones, despite their identical antigen specificities and functions, are non-epidermotropic. We therefore tested whether the differences in epidermotropism of these T cells could reside in the different levels of expression of T-cell surface molecules such as CD3, CD4, CD2, and lymphocyte function associated antigen 1 (LFA 1). The results of flow cytometric analysis showed that LFA-1 was preferentially expressed on the surface of epidermotropic T-cell clones, while non-epidermotropic T-cell clones were negative or very weakly positive for LFA-1 with one exception. After exposure to phorbol ester, epidermotropic clones with high levels of LFA-1 did not show any further up-regulation of LFA-1. In contrast, under identical conditions, significant up-regulation of LFA-1 was observed on non-epidermotropic T cells with low levels of LFA-1. However, even after exposure to phorbol ester, these T cells remained non-epidermotropic. These results suggest that the presence of high levels of LFA-1 on T cells is absolutely necessary for their epidermotropic migration, but its up-regulation is neither necessary nor sufficient to trigger the epidermotropic migration. High levels of LFA-1, regardless of cell activation, may be required to mediate stable cell adhesions leading to epidermotropic migration.

A new constitutively activating point mutation in the luteinizing hormone/choriogonadotropin receptor gene in cases of male-limited precocious puberty.

A single point mutation that encodes an aspartic acid (Asp578) to glycine substitution in the LH/CG receptor (LH/CGR) gene, D578G, was recently found in American patients with familial male-limited precocious puberty and in a Japanese patient with a sporadic form of the disorder. Transfection of the mutant, compared to the wild-type, LH/CGR complementary DNA into COS-7 cells results in higher basal cAMP production, but a normal agonist-induced response; the mutation is, therefore, proposed to constitutively activate Leydig cells and elevate serum testosterone, despite low levels of gonadotropin. In the current study we examined two additional Japanese patients with male-limited precocious puberty without a family history of the disease. We describe a heterozygous cytosine (C) to thymine (T) transition at nucleotide 1715 in both; the mutation encodes an alanine to valine substitution in codon 572 of transmembrane helix 6, A572V. Transfected into COS-7 cells, the A572V mutant exhibited the same constitutively high basal cAMP levels and normal agonist-induced cAMP response as the D578G mutant. We conclude that the constitutively higher cAMP levels caused by the A572V mutation led to Leydig cell activation and male-limited precocious puberty, as in the previously described D578G mutation. As the mother of one of the two patients had the same heterozygous mutation, this patient represents the first recognized case of inherited male-limited precocious puberty in the Japanese population. The previously described D578G mutant did not increase basal or agonist-induced inositol phosphate production in transfected COS-7 cells, or the number of LH/CGRs or their affinity for LH/CG. In contrast, transfection of the A572V mutation in COS-7 cells exhibited significantly higher inositol phosphate levels basally and at 10(-11) mol/L hCG, but significantly lower inositol phosphate levels at 10(-7) mol/L hCG. These data suggest that the A572V mutation of the LH/CGR may have effects on the guanine nucleotide binding protein which activates phospholipase C (Gq) coupling and phospholipase-C activation in addition to its effects on Gs coupling and activation of adenylyl cyclase. A572V-transfected cells also exhibited a higher affinity, despite an apparent decrease in the number of binding sites, for [125I]hCG, compared to transfectants with the wild-type LH/CGR. We hypothesize that these differences between the A572V and D578G mutations reflect a greater impact of the A572V mutation on receptor conformation.

Repression of fibrinolysis in scalded rats by administration of Serratia protease.

The oral administration Serratia protease (SP), an antiinflammatory oral drug, to scalded rats markedly repressed the activation of fibrinolysis induced by the scalding. The repression was most effective when SP was given 3 hr prior to scalding, and a statistically significant repression was observed at a dose of 5 mg/kg. The repressive effect of SP was dependent on its proteolytic activity and was far stronger than those of other proteases tested. The repression was observed also by the intravenous injection of SP at a dose as low as 0.2 microgram/kg, which corresponded to a blood concentration of about 4 ng/ml. In this case, too, the proteolytic activity was essential. In rat blood, SP existed as a complex with a plasma protease inhibitor, alpha 1-macroglobulin (alpha 1 M), with a molar binding ratio of 1:1, still retaining about 20% of its original caseinolytic activity. This ratio, together with the alpha 1 M concentration in rat plasma and the molecular weights of SP and alpha 1 M, enabled the estimation that at the effective SP concentration (4 ng/ml) only 1 out of 20,000 parts of alpha 1 M molecules in plasma took part in the complex formation.

GAPO syndrome: report on the first case in Japan.

We studied a 3.5-year-old Japanese boy with growth retardation, alopecia, pseudoanodontia, and bilateral papilledema. He was born of nonconsanguineous parents, but his paternal grandparents were related. From his characteristic physical manifestations, we diagnosed him as the first known case of GAPO syndrome in Japan and perhaps in the Mongoloid race. Our case had prominent dilatation of scalp veins and an audible intracranial bruit. Cranial angiography documented a narrowing of the sigmoid sinuses, with no flow to either jugular vein. We discuss here the relationships between optic atrophy and intracranial vascular changes in this syndrome.

Isolation and characterization of Streptoverticillium anticoagulant (SAC), a novel protein inhibitor of blood coagulation produced by Streptoverticillium cinnamoneum subsp. cinnamoneum.

Three Streptoverticillium anticoagulants, SAC I, II, and III, which strongly inhibit human intrinsic blood coagulation, were each isolated in a homogeneous form from a culture fluid of Streptoverticillium cinnamoneum subsp. cinnamoneum IFO 12852. SAC I, II, and III are simple proteins with molecular weights of around 12,000, and with isoelectric points of 9.7, 9.7, and 9.9, respectively. Their amino acid compositions are similar and each SAC possesses two disulfide bonds. The COOH-terminal residue of each of these proteins is phenylalanine. Together with the similarity of their protein chemical properties, the results of NH2-terminal amino acid sequence analysis of these SAC proteins strongly suggested that the deletion of Ser-Leu and Ser-Leu-Tyr from the NH2-terminus of SAC I (Ser-Leu-Tyr-Ala-Pro-...) results in the generation of SAC II and III, respectively. The amount of each SAC necessary to double the partial thromboplastin time was around 5 micrograms/ml. SAC I inhibited activated human factor XII and human plasma kallikrein. It also inhibited, but to a lesser extent, activated factor X. The inhibition constants (Ki) of SAC I toward activated factor XII and plasma kallikrein were 5.3 x 10(-8) and 7.2 x 10(-9) M, respectively. The SACs also inhibited some microbial serine proteases such as subtilisin Carlsberg and, to a lesser extent, mammalian serine proteases including bovine trypsin and alpha-chymotrypsin. Of these three inhibitors, only SAC I inhibited metalloproteases such as thermolysin in addition to these serine proteases.

Clinical study of biological response modifiers as maintenance therapy for hepatocellular carcinoma.

We conducted a randomized, controlled trial comparing 5-fluorouracil (5-FU) with or without biological response modifiers (BRMs) as a maintenance therapy for hepatocellular carcinoma (HCC) after treatment with percutaneous ethanol injection (PEI), transcatheter arterial embolization (TAE) or arterial infusion of antitumor agents (AI). A total of 58 cases of HCC were classified into 4 groups as follows: group I, PSK with 5-FU (n = 15); group II, lentinan with 5-FU (n = 15); group III, OK-432 with 5-FU (n = 12); and group IV, 5-FU alone as the control (n = 16). The mean survival time, mortality rate, time to progression, and T4/T8 ratio of lymphocytes in the peripheral blood were compared among the four groups. There was no significant difference in the background factors among the groups. In group I, the T4/T8 ratio of lymphocytes was reduced after the therapy. No significant difference was found among the groups in terms of the mean survival time, mortality rate, or time to progression. PEI for initial therapy was superior to the other therapies in terms of the mean survival time and mortality rate. These results suggest that the addition of BRM to maintenance therapy with 5-FU exerts no prognostic benefit on HCC patients treated with PEI, TAE, or AI.

Effects of TAK-029, a novel GPIIb/IIIa antagonist, on arterial thrombosis in guinea pigs, dogs and monkeys.

The antithrombotic and bleeding time (BT) prolonging effects of TAK-029, a novel GPIIb/IIIa antagonist, were examined in three arterial thrombosis models. In guinea pigs, TAK-029 at 30 micrograms/kg (i.v.) inhibited ADP-induced ex vivo platelet aggregation completely and prolonged BT to 4.5 times the control value 5 min after administration, and it prevented thrombotic occlusion in 2 out of 5 animals in a photochemically-induced basilar thrombosis model. TAK-029 at 100 micrograms/kg (i.v.) prolonged BT more than 9 times 5 min after administration, and it prevented thrombus formation for over 60 min. In dogs, TAK-029 at 30 micrograms/kg (i.v.) inhibited ADP-induced ex vivo platelet aggregation by 87% 5 min after administration, and it prevented thrombotic occlusion in injured and stenosed coronary arteries for 22 min without prolonging the BT. TAK-029 at 100 micrograms/ kg (i.v.) inhibited platelet aggregation completely and prolonged BT 3.6 times 5 min after administration, and it prevented thrombus formation for over 45 min. In monkeys, TAK-029 at 10 micrograms/kg (i.v.) inhibited ADP-induced ex vivo platelet aggregation by 84% and prolonged BT 4.6 times 5 min after the administration, and it prevented thrombotic occlusion in injured and stenosed carotid arteries for 24 min. TAK-029 at 30 micrograms/kg (i.v.) completely inhibited platelet aggregation and thrombus formation for over 60 min, and it prolonged BT more than 7.3 times 60 min after administration. In conclusion, TAK-029 exerted potent antithrombotic effects with BT prolongation in three different arterial thrombosis models. TAK-029 may be effective for the treatment of various arterial thrombotic diseases.

Molecular size difference of urinary heparan sulfates from normal individuals and genetic mucopolysaccharidoses.

Heparan sulfates were isolated from the urine of normal individuals and patients with genetic mucopolysaccharidoses after exhaustive digestion with chondroitinase ABC. Electrophoresis of these preparations on cellulose acetate membrane revealed one spot corresponding in mobility to reference heparan sulphate in barium acetate buffer, while electrophoresis in 0.1 M HCl resulted in two distinct spots for each case; one corresponded in migration rate to reference heparan sulfate, and the other was faster in mobility than reference heparan sulfate but slightly retarded when compared with reference heparin. On thin-layer gel filtration on Sephadex G-200 (superfine) heparan sulfate from normal urine was polydispersed in character and its molecular size was larger than those of other preparations. Heparan sulfates from Hunter's and Sanfilippo's urine were monodispersed and small in molecular size. The molecular size of heparan sulfate from Sanfilippo's urine was the smallest of all. Heparin sulfate from Hurler's urine appeared to be composed of two populations; one corresponded in molecular size to heparan sulfate from normal urine, and the other corresponded to that of Hunter's urine.

Psychotic symptoms in depression and borderline personality disorder.

The authors investigated the prevalence of psychotic symptoms in depression and borderline personality disorder employing the Diagnostic Interview for Borderlines. Clear-cut delusions and hallucinations were rare among the borderlines. However, derealization and depersonalization symptoms were common and were found to be prevalent as among depressives. The prevalence of these symptoms among patients with both borderline personality disorder and depression was similar to that among patients with only borderline personality disorder or depression. The relationship between depression and borderline personality disorder and the significance of psychotic symptoms in these disorders is discussed.