Multielectron-Ion Coincidence Spectroscopy of Xe in Extreme Ultraviolet Laser Fields: Nonlinear Multiple Ionization via Double Core-Hole States.

Ultrafast multiphoton ionization of Xe in strong extreme ultraviolet free-electron laser (FEL) fields (91 eV, 30 fs, 1.6×10^{12} W/cm^{2}) has been investigated by multielectron-ion coincidence spectroscopy. The electron spectra recorded in coincidence with Xe^{4+} show characteristic features associated with two-photon absorption to the 4d^{-2} double core-hole (DCH) states and subsequent Auger decay. It is found that the pathway via the DCH states, which has eluded clear identification in previous studies, makes a large contribution to the multiple ionization, despite the long FEL pulse duration compared with the lifetime of the 4d core-hole states.

Routine measurements of factor VIII activity and inhibitor titer in the presence of emicizumab utilizing anti-idiotype monoclonal antibodies.

Essentials Emicizumab (Emi) affects the APTT-based assays of factor (F)VIII activity and inhibitor titer. A mixture of two anti-Emi monoclonal antibodies (mAb) effectively neutralized the Emi activity. Anti-Emi mAbs completely eliminated the influence of Emi on FVIII activity and inhibitor titer. The inclusion of anti-Emi mAbs in routine FVIII assays would be useful for Emi-treated patients. SUMMARY: Background Emicizumab is an anti-factor (F)IXa/X bispecific monoclonal antibody (mAb), mimicking the factor (F)VIIIa cofactor activity. Emicizumab does not require activation by thrombin and its shortening effect on the activated partial prothrombin time (APTT) is more pronounced than that of factor (F)VIII. APTT-based FVIII activity (FVIII:C) and FVIII inhibiter titer measurements are influenced by the presence of emicizumab. Aim To establish a reliable APTT-based assay to measure FVIII in the presence of emicizumab. Methods Plasmas from hemophilia A (HA) patients without or with inhibitors were studied using one-stage FVIII:C and Bethesda inhibitor assays. Two recombinant anti-idiotype mAbs to emicizumab (anti-emicizumab mAbs) were prepared, rcAQ8 to anti-FIXa-Fab and rcAJ540 to anti-FX-Fab. Results The combined anti-idiotype mAbs (2000 nm each) eliminated the effects of emicizumab on APTTs of HA plasmas without or with inhibitor by competitive inhibition of antibody binding to FIX(a)/FX(a). Measurements of FVIII coagulation activity in HA plasmas without inhibitor were overestimated in the presence of emicizumab (1 µm = ~150 µg mL-1 ) at all reference levels of FVIII. The addition of anti-emicizumab mAbs to the assay mixtures completely neutralized the emicizumab and facilitated accurate determination of FVIII:C. Anti-FVIII inhibitor titers were undetectable in the presence of emicizumab in HA plasmas with inhibitor or normal plasmas mixed with anti-FVIII neutralizing antibodies. These effects of emicizumab were completely counteracted by the addition of the anti-idiotype mAbs, allowing accurate assessment of inhibitor titers. Conclusion The in vitro inclusion of anti-emicizumab mAbs in the standard one-stage coagulation assays prevented interference by emicizumab and enabled accurate measurements of FVIII:C and inhibitor titers.

Tofogliflozin, a sodium/glucose cotransporter 2 inhibitor, attenuates body weight gain and fat accumulation in diabetic and obese animal models.

OBJECTIVE: Tofogliflozin, a highly selective inhibitor of sodium/glucose cotransporter 2 (SGLT2), induces urinary glucose excretion (UGE), improves hyperglycemia and reduces body weight in patients with Type 2 diabetes (T2D). The mechanisms of tofogliflozin on body weight reduction were investigated in detail with obese and diabetic animal models. METHODS: Diet-induced obese (DIO) rats and KKAy mice (a mouse model of diabetes with obesity) were fed diets containing tofogliflozin. Body weight, body composition, biochemical parameters and metabolic parameters were evaluated. RESULTS: In DIO rats tofogliflozin was administered for 9 weeks, UGE was induced and body weight gain was attenuated. Body fat mass decreased without significant change in bone mass or lean body mass. Food consumption (FC) increased without change in energy expenditure, and deduced total calorie balance (deduced total calorie balance=FC-UGE-energy expenditure) decreased. Respiratory quotient (RQ) and plasma triglyceride (TG) level decreased, and plasma total ketone body (TKB) level increased. Moreover, plasma leptin level, adipocyte cell size and proportion of CD68-positive cells in mesenteric adipose tissue decreased. In KKAy mice, tofogliflozin was administered for 3 or 5 weeks, plasma glucose level and body weight gain decreased together with a reduction in liver weight and TG content without a reduction in body water content. Combination therapy with tofogliflozin and pioglitazone suppressed pioglitazone-induced body weight gain and reduced glycated hemoglobin level more effectively than monotherapy with either pioglitazone or tofogliflozin alone. CONCLUSION: Body weight reduction with tofogliflozin is mainly due to calorie loss with increased UGE. In addition, tofogliflozin also induces a metabolic shift from carbohydrate oxidation to fatty acid oxidation, which may lead to prevention of fat accumulation and inflammation in adipose tissue and liver. Tofogliflozin may have the potential to prevent obesity, hepatic steatosis and improve insulin resistance as well as hyperglycemia.

Anti-factor IXa/X bispecific antibody (ACE910): hemostatic potency against ongoing bleeds in a hemophilia A model and the possibility of routine supplementation.

BACKGROUND: We previously reported that a humanized anti-factor IXa/X bispecific antibody, hBS23, mimics the function of FVIII even in the presence of FVIII inhibitors, and has preventive hemostatic activity against bleeding in an animal model of acquired hemophilia A. After further molecular engineering of hBS23, we recently identified an improved humanized bispecific antibody, ACE910, for clinical investigation. OBJECTIVES: To elucidate the in vivo hemostatic potency of ACE910 by examining its effect against ongoing bleeds, and to determine its pharmacokinetic parameters for discussion of its potency for prophylactic use. METHODS: A non-human primate model of acquired hemophilia A was established by injecting anti-primate FVIII neutralizing antibody. When bleeds emerged following an artificial bleed-inducing procedure, either ACE910 or recombinant porcine FVIII (rpoFVIII) was intravenously administered. rpoFVIII was additionally administered twice daily on the following 2 days. Bleeding symptoms were monitored for 3 days. A pharmacokinetic study and multiple-dosing simulations of ACE910 were also performed. RESULTS: A single bolus of 1 or 3 mg kg-1 ACE910 showed hemostatic activity comparable to that of 10 U kg-1 (twice daily) rpoFVIII against ongoing bleeds. The determined ACE910 pharmacokinetic parameters included a long half-life (3 weeks) and high subcutaneous bioavailability (nearly 100%). The simulation results based on pharmacokinetic parameters indicated that the above hemostatic level could be maintained with once-weekly subcutaneous administration of ACE910, suggesting the possibility of more effective prophylaxis. CONCLUSIONS: ACE910 may offer an alternative on-demand treatment option for patients with hemophilia A, as well as user-friendly and aggressive routine supplementation.

Anti-factor IXa/X bispecific antibody (ACE910): hemostatic potency against ongoing bleeds in a hemophilia A model and the possibility of routine supplementation.

BACKGROUND: We previously reported that a humanized anti-factor IXa/X bispecific antibody, hBS23, mimics the function of FVIII even in the presence of FVIII inhibitors, and has preventive hemostatic activity against bleeding in an animal model of acquired hemophilia A. After further molecular engineering of hBS23, we recently identified an improved humanized bispecific antibody, ACE910, for clinical investigation. OBJECTIVES: To elucidate the in vivo hemostatic potency of ACE910 by examining its effect against ongoing bleeds, and to determine its pharmacokinetic parameters for discussion of its potency for prophylactic use. METHODS: A nonhuman primate model of acquired hemophilia A was established by injecting anti-primate FVIII neutralizing antibody. When bleeds emerged following an artificial bleed-inducing procedure, either ACE910 or recombinant porcine FVIII (rpoFVIII) was intravenously administered. rpoFVIII was additionally administered twice daily on the following 2 days. Bleeding symptoms were monitored for 3 days. A pharmacokinetic study and multiple-dosing simulations of ACE910 were also performed. RESULTS: A single bolus of 1 or 3 mg kg⁻¹ ACE910 showed hemostatic activity comparable to that of 10 U kg⁻¹ (twice daily) rpoFVIII against ongoing bleeds. The determined ACE910 pharmacokinetic parameters included a long half-life (3 weeks) and high subcutaneous bioavailability (nearly 100%). The simulation results based on pharmacokinetic parameters indicated that the above hemostatic level could be maintained with once-weekly subcutaneous administration of ACE910, suggesting the possibility of more effective prophylaxis. CONCLUSIONS: ACE910 may offer an alternative on-demand treatment option for patients with hemophilia A, as well as user-friendly and aggressive routine supplementation.

Effect of SGLT2 inhibitors in a murine model of urinary tract infection with Candida albicans.

AIMS: Urinary tract infection (UTI) is a common clinical problem in diabetic patients; however, the relationship between UTI and glucosuria remains uncertain. To investigate the relationship, we examined the effect of glucosuria induced by sodium glucose cotransporter 2 (SGLT2) inhibitors on the progression of UTI in mice. METHODS: From 1 day before transurethral inoculation with Candida albicans, female mice were treated orally once a day with an SGLT2 inhibitor in different treatment regimens: (i) dapagliflozin at 10 mg/kg for 2, 3 or 7 days, (ii) dapagliflozin at 0.1, 1 or 10 mg/kg for 3 days and (iii) dapagliflozin, canagliflozin or tofogliflozin at 10 mg/kg for 3 days. To evaluate the ascending UTI, the kidneys were removed 6 days after the inoculation, and the number of viable C. albicans cells in kidney was measured as colony-forming units (CFU). RESULTS: In mice treated with dapagliflozin, the number of C. albicans CFU in kidney increased in accordance with both treatment duration and dose. The number of CFU significantly increased when mice were treated with 10 mg/kg dapagliflozin or canagliflozin but not tofogliflozin. With dapagliflozin and canagliflozin, urine glucose concentration (UGC) significantly increased up to 24 h after drug administration; with tofogliflozin, UGC significantly increased only up to 12 h after drug administration. CONCLUSIONS: Our data indicate that increased susceptibility to UTI is associated with a persistent increase in UGC.

Tofogliflozin, a novel sodium-glucose co-transporter 2 inhibitor, improves renal and pancreatic function in db/db mice.

BACKGROUND AND PURPOSE: Although inhibition of renal sodium-glucose co-transporter 2 (SGLT2) has a stable glucose-lowering effect in patients with type 2 diabetes, the effect of SGLT2 inhibition on renal dysfunction in type 2 diabetes remains to be determined. To evaluate the renoprotective effect of SGLT2 inhibition more precisely, we compared the effects of tofogliflozin (a specific SGLT2 inhibitor) with those of losartan (an angiotensin II receptor antagonist) on renal function and beta-cell function in db/db mice. EXPERIMENTAL APPROACH: The effects of 8-week tofogliflozin or losartan treatment on renal and beta-cell function were investigated in db/db mice by quantitative image analysis of glomerular size, mesangial matrix expansion and islet beta-cell mass. Blood glucose, glycated Hb and insulin levels, along with urinary albumin and creatinine were measured KEY RESULTS: Tofogliflozin suppressed plasma glucose and glycated Hb and preserved pancreatic beta-cell mass and plasma insulin levels. No improvement of glycaemic conditions or insulin level was observed with losartan treatment. Although the urinary albumin/creatinine ratio of untreated db/db mice gradually increased from baseline, tofogliflozin or losartan treatment prevented this increase (by 50-70%). Tofogliflozin, but not losartan, attenuated glomerular hypertrophy. Neither tofogliflozin nor losartan altered matrix expansion. CONCLUSIONS AND IMPLICATIONS: Long-term inhibition of renal SGLT2 by tofogliflozin not only preserved pancreatic beta-cell function, but also prevented kidney dysfunction in a mouse model of type 2 diabetes. These findings suggest that long-term use of tofogliflozin in patients with type 2 diabetes may prevent progression of diabetic nephropathy.

A multicenter study of leukocytapheresis in rheumatoid arthritis.

OBJECTIVE: To evaluate the efficacy and safety of leukocytapheresis (LCAP) in patients with rheumatoid arthritis (RA) that is refractory to disease modifying antirheumatic drugs (DMARDs), we conducted a prospective, multicenter, open-label clinical trial. METHODS: We enrolled 38 active RA patients, including 32 patients who showed an inadequate response to > or = 2 DMARDs and 6 patients with rapidly progressive RA. All patients continued drug therapy and were treated with 5 LCAP sessions conducted at 1-week intervals. The clinical response was evaluated at baseline before starting LCAP and at 4 weeks after the completion of all the LCAP sessions using the American College of Rheumatology (ACR) criteria and the 28-joint disease activity score (DAS28) of the European League Against Rheumatism (EULAR). RESULTS: Of the 35 patients who fulfilled the study's eligibility criteria, 24 (69%), 10 (29%), and 23 (66%) patients achieved 20% (ACR20), 50% (ACR50), and DAS28-C-reactive protein (CRP) EULAR improvement, respectively. The mean DAS28-CRP score of the 35 patients decreased significantly from 5.99 +/- 0.92 at baseline to 4.54 +/- 1.39 after treatment. Comparison analysis of the ACR20 responders and non-responders to LCAP revealed that 22 of 24 responders (92%) concomitantly received methotrexate, whereas significantly fewer, that is, 6 of 11 non-responders (55%) received methotrexate. Less frequent and transient mild-to-moderate adverse events, including nausea and headache, were seen in 12 of 189 LCAP sessions (6.3%). CONCLUSION: These results demonstrate the usefulness of LCAP in combination with DMARDs, particularly methotrexate, as an effective and safe treatment for refractory RA.

Suppressive effect of leflunomide metabolite (A77 1726) on metalloproteinase production in IL-1beta stimulated rheumatoid synovial fibroblasts.

Leflunomide, an isoxazol derivative structurally unrelated to other immunomodulatory drugs, has proven to be efficacious in the treatment of rheumatoid arthritis (RA). This study was conducted to elucidate the mechanism by which leflunomide mediated antirheumatic effects. We investigated the effects of A77 1726, leflunomide's active metabolite, on mitogen-activated protein kinase (MAPK) activation in IL-1beta-stimulated rheumatoid synovial fibroblasts. The effects of A77 1726 on the secretion of matrix metalloproteinases (MMPs) from rheumatoid synovial fibroblasts were also examined. A77 1726 partially suppressed IL-1beta-induced ERK1/2 and p38 kinase activation. In contrast, A77 1726 efficiently suppressed IL-1beta-stimulated JNK1/2 kinase activation. Although no suppressive effect was demonstrated on MMP-2, A77 1726 markedly inhibited MMP-1, 3, and 13 secretions from IL-1beta-stimulated rheumatoid synovial fibroblasts. Tissue inhibitor of metalloproteinases-1 (TIMP-1) was constitutively produced from rheumatoid synovial fibroblasts and the suppressive effects of A77 1726 on TIMP-1 production were minimal. Our results suggest that the suppression of the MAPK signalling pathway and MMP synthesis in rheumatoid synovial fibroblasts is a possible mechanism for the inhibitory activity of leflunomide against rheumatoid arthritis.

Anti-apoptogenic function of TGFbeta1 for human synovial cells: TGFbeta1 protects cultured synovial cells from mitochondrial perturbation induced by several apoptogenic stimuli.

OBJECTIVE: To investigate anti-apoptogenic mechanism of transforming growth factor beta1 (TGFbeta1) towards synovial cells. METHODS: Isolated synovial cells, treated or not with TGFbeta1, were cultured in the presence or absence of anti-Fas IgM, proteasome inhibitor Z-Leu-Leu-Leu-aldehyde (LLL-CHO), etoposide, or C2-ceramide. After cultivation, apoptosis of synovial cells was examined by the presence of hypodiploid DNA(+) cells, the presence of terminal deoxy (d)-UTP nick end labelling(+) cells (TUNEL(+) cells), activation of caspases, and disruption of mitochondrial transmembrane potential (DeltaPsim). RESULTS: Activation of caspase-9 and DeltaPsim was found in anti-Fas IgM treated synovial cells. The increment of both hypodiploid DNA(+) cells and TUNEL(+) cells accompanied by the activation of caspase-8 and caspase-3 was also determined in anti-Fas IgM treated synovial cells. These hallmarks for apoptosis induced by anti-Fas IgM were significantly suppressed in TGFbeta1 treated synovial cells. LLL-CHO, etoposide, and C2-ceramide also caused DeltaPsim, the increment of both hypodiploid DNA(+) cells and TUNEL(+) cells, and the activation of both Leu-Glu-His-Asp ase (LEHDase; caspase-9 like activity) and Asp-Glu-Val-Asp ase (DEVDase; caspase-3 like activity) in synovial cells. As determined in anti-Fas IgM treatment, TGFbeta1 significantly reduced apoptotic cell death of synovial cells induced by the above chemicals. CONCLUSIONS: The protective effect of TGFbeta1 for mitochondrial homoeostasis may be important in the anti-apoptogenic function of TGFbeta1 for synovial cells.

Atypical mycobacteriosis in two patients with rheumatoid arthritis.

Abstract We report two cases of rheumatoid arthritis (RA) with atypical mycobacteriosis. Opportunistic infections are critical complications for rheumatic diseases. The use of steroids or immunosuppressants may increase the risk of opportunistic infections. However, these reports are rare in that they demonstrate atypical mycobacterial infections as complications of RA, even though no immunosuppressive agents were used. We discuss the characteristics of atypical mycobacterial infections of the lung in RA.

A case of eosinophilic myositis associated with orbital myositis.

Abstract A 29-year-old woman was admitted to our hospital because of fever, myalgia, and ocular pain. She had been given norgesterone and ethinylestradiol orally. Laboratory data indicated the presence of systemic inflammation together with elevated levels of muscle enzymes. Magnetic resonance imaging revealed inflammatory lesions in skeletal and extraocular muscles. The diagnosis of eosinophilic myositis was based on histopathological examination of the deltoid muscle, which showed infiltration of eosinophils into the tissue with necrotizing muscle fibers. Prednisolone treatment resulted in marked clinical improvement. This is the first case report of eosinophilic myositis in which the extraocular muscles were affected.

Regulation of rheumatoid synoviocyte proliferation by endogenous p53 induction.

The p53 tumour suppressor protein protects cells from tumorigenic alterations by inducing either cell growth arrest or apoptosis. In the present study, we investigated the role of endogenous p53 expressed in rheumatoid arthritis synovial fibroblasts which show transformed-appearing phenotypes. Type B synovial cells (fibroblast-like synovial cells) were exposed to a proteasome inhibitor, carbobenzoxyl-leucinyl-leucinyl-leucinal (MG-132). During this process, the expressions of p53 and p21 were examined by Western blot. Cell cycle analysis of the synovial cells was determined by DNA staining using propidium iodide (PI). Inhibition of proteasome resulted in the accumulation of p53 which was followed by an increase in the amount of a cyclin-dependent kinase (CDK)-inhibitor, p21. As a consequence, the retinoblastoma gene product, Rb, remained in the hypophosphorylated state, thus preventing PDGF-stimulated synovial cells from progressing into S-phase. This study shows that endogenous p53, which is inducible in rheumatoid synovial cells, is functionally active based on the findings that its expression blocks the G1/S transition by inhibiting the CDK-mediated phosphorylation of Rb via p21 induction. Thus the induction of p53 using proteasome inhibitor may provide a new approach in the treatment of RA.

Validation of an analytical method for a potent antitumor agent, TZT-1027, in plasma using liquid chromatography-mass spectrometry.

A sensitive and specific analytical method for a potent antitumor agent, TZT-1027, in plasma has been developed using liquid chromatography-mass spectrometry (LC-MS) with [2H4]TZT-1027 as an internal standard (I.S.). A plasma sample was purified by solid-phase extraction on a C18 cartridge, followed by solvent extraction with diethyl ether. The extract was then injected into the LC-MS system. Chromatography was carried out on a C18 reversed-phase column using acetonitrile-0.05% trifluoroacetic acid (TFA) (55:45) as a mobile phase. Mass spectrometric analysis was performed in atmospheric pressure chemical ionization (APCI) mode with positive ion detection, and the protonated molecular ions ([M+H]+) of TZT-1027 and I.S. were monitored to allow quantitation. The method was applied to the determination of TZT-1027 in human, monkey, dog, rat and mouse plasma. As far as the sample preparation was concerned, good recoveries (73.5-99.1%) were obtained. The calibration curves were linear over the range of 0.25-100 ng per 1 ml of human, dog and rat plasma, per 0.5 ml of monkey plasma, and per 0.1 ml of mouse plasma. From the intra- and inter-day accuracy and precision, the present method satisfies the accepted criteria for bioanalytical method validation. TZT-1027 was stable when stored below -15 degrees C for 6 months in human plasma and for 3 weeks in plasma from other species. TZT-1027 was also stable in plasma through at least three freeze-thaw cycles.

Expression of membrane-type 1 matrix metalloproteinase in rheumatoid synovial cells.

Membrane-type 1 matrix metalloproteinase (MT1-MMP) is thought to be a putative regulator of pro-gelatinase A (MMP-2) in the rheumatoid synovium. In this study, we examined the effects of IL-1beta, one of the inflammatory cytokines, on the expression of MT1-MMP and the activation of pro-MMP-2 using rheumatoid synovial cells. We also studied the effects of KE-298 (2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid), a new disease-modifying anti-rheumatic drug (DMARD), on MT1-MMP expression of rheumatoid synovial cells. Type B synovial cells (fibroblast-like synovial cells) were cultured with KE-298 (25-100 microg/ml) in the presence of IL-1beta for 48 h. Activation of pro-MMP-2 secreted from synovial cells was analysed by gelatin zymography. Reverse transcription-polymerase chain reaction (RT-PCR) methods were used to detect MT1-MMP mRNA. MT1-MMP protein expression on synovial cells was examined by anti-MT1-MMP immunoblot. An active form of MMP-2 was demonstrated in the culture media conditioned by IL-1beta-stimulated synovial cells. In addition, MT1-MMP mRNA and protein expression of rheumatoid synovial cells were increased by IL-1beta treatment. KE-298 blocked this IL-1beta-induced pro-MMP-2 activation and MT1-MMP expression, but did not affect IL-1beta-induced tissue inhibitor of metalloproteinase-2 (TIMP-2) secretion from rheumatoid synovial cells. These findings indicate that activation of rheumatoid synovial cells by IL-1beta results in the induction of MT1-MMP expression. Given that MT1-MMP promotes matrix degradation by activating pro-MMP-2, these results suggest a novel mechanism whereby cytokine may contribute to articular destruction in rheumatoid arthritis (RA). KE-298 may prevent this process by down-regulating MT1-MMP expression.

[A study on the clinical features of pulmonary tuberculosis in elderly patients].

The clinical findings of pulmonary tuberculosis in elderly patients aged 75 years of age or older (elderly group) were compared with the middle-aged patients aged between 45 and 54 years of age (middle-aged group) and the young patients aged 34 years of age or younger (young group). The elderly patients who died in hospital were also compared with the elderly patients who survived. Study subjects were culture-positive pulmonary tuberculosis patients who were discharged from our hospital from December 1996 to November 1998. There were 79 patients in the elderly group, 95 in the middle-aged group, and 88 in the young group. The results were as follows. 1) The male/female ratio was significantly lower in the elderly group (1.9:1) than the middle-aged group (6.9:1). 2) Complication was noted more frequently in the elderly group. However, diabetes mellitus was noted less frequently in the elderly group (12.7%) than the middle-aged group (28.4%). 3) The frequency of cavitation was lower in the elderly group (59.5%) than the middle-aged group (87.4%). 4) The fever over 38 degrees C was noted less frequently in the elderly group (17.7%) than the other groups, while the frequency of the fever over 37 degrees C showed no significant difference between the elderly group and the other groups. 5) The frequencies of hypoalbuminemia and appetite loss were higher in the elderly group than the other groups. 6) The elderly group showed high mortality rate of 31.6%. The complication with cerebrovascular disease was noted significantly higher in the patients who died in hospital than those who survived. The frequency of widespread infiltrates, fever over 38 degrees C, neutrophilia, hypoalbuminemia and appetite loss were all significantly higher in the patients who died in hospital while their sputa were still positive on culture than those who survived. Our study clearly showed the features of elderly pulmonary tuberculosis patients in comparison with middle-aged patients and young patients. These features are very important to suspect the diagnosis of pulmonary tuberculosis in elderly patients with some atypical manifestation. Our study also suggests that the delay in diagnosing tuberculosis causes more frequently the patients' deterioration and death in elderly patients than in middle-aged patients and young patients.

[Tuberculosis of the elderly (above the age of 75) in national hospitals].

In Japan tuberculosis is becoming rapidly the disease of the elderly. We studied the background, the type and level of hospital cares needed, and the outcome of patients with pulmonary tuberculosis (sputum smear and/or culture positive) above the age of 75 who were admitted to 8 national hospitals during the period from January 1 to December 31, 1997. The study included 150 patients (male: 109, female: 41, mean age: 81.6), of whom 25% needed care in a single-bed room, 84.3% had underlying diseases (cardiovascular diseases: 43.3%, malignant diseases: 20.9%, neuro-psychiatric diseases: 13.4%), and 47.6% needed cares mainly in feeding and excretions. 92 patients (62.6%) improved and 45 patients (30.6%) died, of whom the cause of death was directly related to tuberculosis in 42. The mean hospital stay was 4.7 months. However, in 42 patients whose cause of death was related to tuberculosis, 66% died within 3 months, while in 102 patients who were discharged 71% stayed more than 3 months. The same comparison was done in 508 patients with bacteriologically proven tuberculosis above the age of 75 admitted to National Tokyo Hospital during the period from 1990 to 1999. The result was almost the same, among 133 patients died in the hospital 60% died within 3 months, while in 375 patients who were discharged hospital stay was more than 3 months in 70%. In the near future, the elderly will occupy more than 25% of the beds of the tuberculosis ward in Japan and most of them have underlying diseases other than tuberculosis. Because tuberculosis, once the disease of the young, is becoming rapidly the disease of the elderly, it is imperative for us to make necessary adjustments to meet this inevitable trend.

New disease-modifying antirheumatic drug 2 acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298) selectively augments activation-induced T cell death.

We examined in this study whether the newly developed disease-modifying antirheumatic drug (DMARD) 2-acetylthiomethyl-4-(4-methylphenyl)-4-oxobutanoic acid (KE-298) augments activation-induced T cell death. Peripheral blood (PB) T cells, isolated from healthy donors, were activated by incubation with interleukin-2 (IL-2) followed by further culture with 12-0-tetradecanoyl phorbol 13-acetate (PMA) and ionomycin in the presence or absence of KE-298. The apoptosis of activated T cells was examined by flow cytometric determination of hypodiploid DNA. Fas expression and caspase-3 activity in activated T cells were also examined by flow cytometry, and expression of Fas ligand (FasL), Bcl-2-related proteins, and X chromosome-linked inhibitor of apoptosis protein (XIAP) was determined by Western blot analysis. Apoptosis was not obvious in resting T cells and was not augmented by KE-298. In contrast, apoptosis was clearly detected in activated T cells (activation-induced T cell death) with the increment of caspase-3 activity, and incubation of these cells with KE-298 further enhanced apoptosis. Treatment of activated T cells with KE-298 increased Bax expression but decreased XIAP expression without affecting the expression of Fas/FasL. Thus caspase-3 activity in activated T cells appeared to be increased by KE-298. Our results suggest that the newly developed DMARD, KE-298, selectively augmented activation-induced T cell death. This finding may contribute to the therapeutic efficacy of KE-298 in rheumatoid arthritis (RA) patients and provide new insight into the pharmacologic action of DMARDs.