Biomedical subjects

H Yamada

Publications and source records attributed to H Yamada.

At least 361 records · Page 20Linked to original sources

PubMed2000

Lignin-carbohydrate complexes: intestinal immune system modulating ingredients in kampo (Japanese herbal) medicine, juzen-taiho-to.

An extract preparation (TJ-48) of a Japanese herbal (Kampo) medicine, Juzen-Taiho-To has been found to show an enhancing activity on proliferation of bone marrow cells mediated by Peyer's patch cells (intestinal immune system modulating activity) (1). When TJ-48 was fractionated by MeOH- and water-extractions, EtOH precipitation and dialysis, water-soluble dialyzable (F-3) and polysaccharide fractions (F-5) both showed a significant intestinal immune system modulating activity in vitro; other fractions had no activity. Oral administration of F-3 (150 mg/kg) also showed the intestinal immune system modulating activity in C3H/HeJ mice. Two active substances were purified by gel filtration from F-3. Chemical analysis indicated that the active substances comprised lignin as well as a carbohydrate consisting mainly of arabinose, galactose, glucose and galacturonic acid. Activity was significantly reduced not only by NalO4 oxidation but also by NaClO2 treatment. Standard lignin showed no activity. Carbohydrate and lignin in the active substances were co-eluted by hydrophobic-interaction chromatography on phenyl-Sepharose, and the stimulating effect and lignin content of the substances did not change even after the hydrophobic-interaction chromatography. These results suggest that lignin-carbohydrate complexes are involved in intestinal immune system modulation by TJ-48.

Animals↗

PubMed2000

Use of angiotensin II stress pulsed tissue Doppler imaging to evaluate regional left ventricular contractility in patients with hypertrophic cardiomyopathy.

There is controversy concerning whether contract-ility in the nonhypertrophied region of the left ventricular (LV) wall is impaired or normal in patients with hypertrophic cardiomyopathy (HCM). Global LV systolic function decreases with increases in afterload in this disease. This study was performed to identify abnormalities in regional LV contractility along the long and short axes in the setting of HCM with the use of angiotensin II (AT-II) stress pulsed tissue Doppler imaging (PTDI). Angiotensin II was administered intravenously to patients with asymmetric septal hypertrophy (HCM group, n = 21) and age-matched normal volunteers (N group, n = 12). We then measured the percent LV fractional shortening (%FS) and end-systolic circumferential LV wall stress by M-mode echocardiography, LV ejection fraction (LVEF) by 2-dimensional echocardiography, and time-velocity integral (TVI) of LV outflow velocity by pulsed Doppler echocardiography. The peak first and second systolic LV wall motion velocities along the long (L-Sw(1) and L-Sw(2)) and short (S-Sw(1) and S-Sw(2)) axes were measured in the LV posterior wall and ventricular septum with the use of PTDI. The end-systolic circumferential LV wall stress at baseline was significantly lower in the HCM group. The L-Sw(1) and L-Sw(2) for the posterior wall were significantly lower in the HCM group, but the S-Sw(1) and S-Sw(2) for the posterior wall and ventricular septum were similar in the two groups. The %FS, LVEF, TVI, and systolic PTDI variables along both axes for the posterior wall decreased significantly, and end-systolic circumferential LV wall stress increased significantly at AT-II doses of 0.005 or 0.010 microg/kg per minute in the HCM group. No significant changes were found in either group in the systolic PTDI variables (except for L-Sw(1)) for the ventricular septum with AT-II infusion. Contractility along the long and short axes of the nonhypertrophied LV wall is easily impaired with increases in afterload in patients with HCM, resulting in a decrease in global LV systolic function. We found AT-II stress PTDI to be a safe and useful technique for evaluating the regional LV systolic function in this disease.

Analysis of Variance↗

PubMed2000

Evaluation of pulmonary gas exchange after lobectomy and simple thoracotomy.

Postoperative influence of simple thoracotomy and lobectomy on pulmonary gas exchange was investigated in 10 patients undergoing thoracotomy and lobectomy (L group) and 10 patients with simple thoracotomy (ST group). Gas exchange was assessed with hot-wire mass spectrometry and blood gas analyses preoperatively and postoperatively at 1, 3, 6, 12, 24 and 48 h and on days 4, 6 and 13. The preoperative values of alveolar-arterial oxygen tension gradient (AaDO2), ventilation-perfusion mismatch (V(A)/Q) and circulatory shunt (Qs/Qt) were maintained in the ST group. In the L group, however, AaDO2 and Qs/Qt showed good maintenance of gas exchange for only 6-12h after lobectomy and then deteriorated because of peripheral atelectasis, resulting in poorer values than those in the ST group on days 4 and 13. Careful postoperative management, including pulmonary toilet, is essential for maintenance of gas exchange and prevention of pulmonary complications.

Adult↗

PubMed2000

Tumor necrosis factor-alpha (TNF-alpha)-induced and interleukin-1 beta (IL-1 beta)-induced shedding of TNF receptors from gingival fibroblasts.

Tumor necrosis factor-alpha (TNF-alpha) exerts its functions by binding two different receptors (TNFR55 and TNFR75). Both TNFR55 and TNFR75 exist in cell-associated and soluble forms. Soluble TNF receptors (sTNFR), sTNFR55 and sTNFR75, are proteolytically shed upon inflammatory stimuli and then modulate various TNF-alpha bioactivities. As human gingival fibroblasts (HGF) can be potential targets for TNF-alpha in inflamed gingiva, we hypothesized that HGF partially modulate the cellular responses to TNF-alpha by regulating their own TNFR. In this study, the kinetics of expression of cell-associated and soluble forms of both receptors from cultured HGF in response to proinflammatory cytokines TNF-alpha and interleukin-1 beta (IL-1 beta) were investigated in vitro. Both TNF-alpha and IL-1 beta upregulated the gene expression of TNFR75 and did not affect that of TNFR55. TNF-alpha and IL-1 beta decreased binding of [(125)I]TNF-alpha to HGF. Moreover, TNF-alpha and IL-1 beta upregulated the release of sTNFR75 from HGF but not that of sTNFR55. These results suggest that HGF under inflammatory conditions may contribute to the inactivation of circulating TNF-alpha through the preferential induction and shedding of TNFR75.

Antigens, CD↗

PubMed2000

Unusual cytotoxic activities of thymus-independent, self-antigen-specific CD8(+) T cells.

We compared the cytotoxic activities of thymus-dependent and thymus-independent CD8(+) T cells. Thymus-dependent CD8(+) T cells, which are foreign antigen specific, acquired cytotoxic activity to tumor cells with a basal dose of the antigen peptides and to hybridoma cells expressing anti-TCR mAb only after differentiation into effector cytotoxic T lymphocytes (CTL). In contrast, thymus-independent CD8(+) T cells, which have been shown to be self-antigen specific, never showed cytotoxic activity to the target cells with a basal dose of the self-antigen peptide, while they could lyse hybridoma cells expressing anti-TCR mAb even without prior antigenic stimulation. Furthermore, the ex vivo cytotoxic activity of thymus-independent CD8(+) T cells was also observed against the target cells with high doses of the antigen peptides, which were not lysed by freshly isolated thymus-dependent CD8(+) T cells. Thus it is revealed that thymus-independent, self-antigen-specific CD8(+) T cells already acquire mature CTL functions in situ but have an increased threshold of TCR-mediated signaling for activation. These differences in cytotoxic activities between thymus-dependent and thymus-independent CD8(+) T cells suggest distinct roles of the two subsets of CD8(+) T cells in vivo.

Animals↗

PubMed2000

Clinical significance of major and minor bcr/abl chimeric transcripts in essential thrombocythemia.

BACKGROUND: Contradictory results have been reported in terms of detecting bcr/abl transcripts in patients with essential thrombocythemia. The aim of this study was to investigate whether the bcr/abl transcript could be found in patients with essential thrombocythemia (ET). METHODS: The bcr/abl transcript was amplified by the RT-nested PCR method using RNA extract from leukocytes taken from 14 essential thrombocythemia patients. The amplified DNAs were electrophoresed in 1% agarose and visualized with ethidium bromide. The DNA bands associated with the bcr/abl transcript were then extracted and followed by DNA sequence analysis. RESULTS: Major bcr/abl transcripts of the b3a2 type and minor ones of the e1a2 type were found in one and two ET patients, respectively. The incidence of bcr/abl transcripts was 21.4% (three of 14 patients). CONCLUSION: Our experiments confirmed that bcr/abl transcripts are present in some patients with essential thrombocythemia.

Adult↗

PubMed2000

Convenient and efficient in vitro folding of disulfide-containing globular protein from crude bacterial inclusion bodies.

We investigated how the folding yield of disulfide-containing globular proteins having positive net charges from crude bacterial inclusion bodies was affected by additives in the folding buffer. In screening folding conditions for human ribonucleases and its derivative, we found that addition of salt (about 0.4 M) to a folding buffer increased the folding yield. This suggested that electrostatic interaction between polyanionic impurities such as nucleic acids and cationic unfolded protein led to the formation of aggregates under the low-salt conditions. Since inclusion bodies were found to contain nucleic acids regardless of the electrostatic nature of the expressed protein, the electrostatic interaction between phosphate moieties of nucleic acids and basic amino acid residues of a denatured protein may be large enough to cause aggregation, and therefore the addition of salt in a folding buffer may generally be useful for promotion of protein folding from crude inclusion bodies. We further systematically investigated additives such as glycerol, guanidium chloride, and urea that are known to act as chemical chaperons, and found that these additives, together with salt, synergistically improved folding yield. This study, suggesting that the addition of salt into the folding buffer is one of the crucial points to be considered, may pave the way for a systematic investigation of the folding conditions of disulfide-containing foreign proteins from crude bacterial inclusion bodies.

Bacterial Proteins↗

PubMed2000

Stabilization of human RNase 1 by introduction of a disulfide bond between residues 4 and 118.

In order to stabilize human RNase 1 by introduction of an intramolecular cross-link, a mutant protein (4-118CL RNase 1), in which Arg4 and Val118 are replaced with cysteine residues and linked by a disulfide bond, was designed and expressed in Escherichia coli as inclusion bodies. The 4-118CL RNase 1 that refolded under redox conditions was a monomer without free SH groups and retained 11% of the activity of the wild-type recombinant RNase 1, indicating that the mutant enzyme was correctly folded with the formation of an additional disulfide bond between Cys4 and Cys118. From guanidium chloride denaturation experiments based on the assumption of a two-state transition for unfolding, it was demonstrated that the introduction of the present cross-link increased the thermodynamic stability of RNase 1 by 2.0 kcal/mol. This value was lower than that, 5.4 kcal/mol, theoretically calculated from the reduction of chain entropy of the unfolded state due to the introduction of the cross-link. These results suggest that the present cross-link also destabilized the folded state of RNase 1 by 3.4 kcal/mol. Along with the increase in the thermodynamic stability, the stability of RNase 1 against trypsin digestion was also significantly increased by the introduction of this cross-link. It is likely, although not proven, that stabilized human RNases are favorable for clinical use, because human RNase-based immunotoxins should have long half-lives as to proteolytic degradation after endocytosis.

Cysteine↗

PubMed2000

A comparative study of surgical approaches for cervical compressive myelopathy.

Since 1986, the authors have used anterior decompression and fusion to treat patients with one- or two-level lesions without spinal canal stenosis (Group A) and laminoplasty for patients with more than three-level lesions or spinal canal stenosis (Group P). The aim of this study was to compare surgical outcomes of anterior and posterior approaches for patients with cervical myelopathy because of spondylosis and disc herniation and to determine the cause of poor neurologic recovery after surgery. One hundred thirty-six patients were followed up for an average of 5.6 years. There were no significant differences in gender, preoperative neurologic deficits, axial symptoms, or duration of symptoms before surgery between the two groups. Mean recovery rates for disc herniations were 71.1% and 71.9% in Groups A and P, respectively. For spondylosis, mean recovery rates were 49.0% and 58.6% in Groups A and P, respectively. There were no differences in recovery rate for patients with either spinal disorder between Groups A and P. The neurologic recovery of patients with kyphotic spinal cord was inferior to that of patients with lordotic or straight spinal cord. It is possible that acquisition and maintenance of lordosis result in improvement of clinical outcomes after surgery for patients with myelopathy.

Aged↗

PubMed2000

Effects of the phosphodiesterase III inhibitors olprinone, milrinone, and amrinone on hepatosplanchnic oxygen metabolism.

OBJECTIVE: To measure the hepatic venous oxygen saturation in patients after cardiac surgery and to compare the effects of olprinone (OLP), a newly synthesized phosphodiesterase III inhibitor, with those of milrinone (MIL) and amrinone (AMR) on hepatosplanchnic oxygen dynamics. Phosphodiesterase III inhibitors are used to improve the hemodynamic state after cardiac surgery. However, the effect of these agents on the hepatosplanchnic circulation has not been investigated thoroughly. DESIGN: Prospective, randomized study. SETTING: University hospital intensive care unit (ICU). PATIENTS: Twenty-nine patients undergoing elective cardiac surgery. MEASUREMENTS AND MAIN RESULTS: In each patient, a 7.5-Fr oximeter catheter was placed in the hepatic vein via the right femoral vein. Catheterization was completed before admission to the ICU, and the study was performed 8 to 24 hrs after surgery, after obtaining stable systemic hemodynamics in the ICU. The patients were assigned randomly to three groups, and they received one of three drugs for 2 hrs (OLP group, 0.3 microg/kg/min of OLP; MIL group, 0.5 microg/kg/min of MIL; AMR group, 10 microg/kg/min of AMR). The authors did not change the patient's hemodynamic interventions, including catecholamines and vasodilators, throughout the study period. Arterial and hepatic venous blood gas data and hemodynamic data (via a pulmonary artery catheter) were obtained before and after drug infusion. Using these data, the authors calculated systemic oxygen delivery and consumption, the systemic oxygen extraction ratio and the hepatosplanchnic oxygen extraction ratio, and the change in hepatosplanchnic blood flow using Fick's equation. Although the increases in cardiac index were not significantly different among the three groups, hepatic venous oxygen saturation increased significantly only in the OLP group (from 47.1% +/-2.6% to 57.0% +/- 1.5% in the OLP group, from 48.4% +/- 2.3% to 50.9% +/- 2.6% in the MIL group, and from 49.8% +/- 3.6% to 50.8% + +/-.7% in the AMR group). The calculated hepatosplanchnic blood flow change was significantly larger in the OLP group than in the other groups (30.1% +/- 5.7% in the OLP group, 9.3% +/- 5.1% in the MIL group, and 2.6% +/- 6.5% in the AMR group). CONCLUSIONS: These results suggest that OLP enhances hepatosplanchnic blood flow and thus may be beneficial in protecting the hepatosplanchnic organs after cardiac surgery.

Amrinone↗

PubMed2000

Genistein potentiates the relaxation induced by beta1- and beta2-adrenoceptor activation in rat aortic rings.

In rat aortic rings, genistein, an inhibitor of tyrosine kinase, but not daidzein, an inactive analogue of genistein, potentiated the relaxation induced by isoproterenol. Atenolol, a beta1-adrenoceptor antagonist, or ICI-118,551, a beta2-adrenoceptor antagonist, inhibited the relaxation induced by isoproterenol. The potentiating effect of genistein on the relaxation induced by isoproterenol in the presence of ICI-118,551 was apparently greater than that in the presence of atenolol. In the presence of ICI-118,551, theophylline, an inhibitor of cyclic adenosine monophosphate (cAMP)-dependent phosphodiesterase (cAMP-PDE), markedly inhibited the potentiating effect of genistein on the isoproterenol-induced relaxation, whereas in the presence of atenolol, theophylline only partly inhibited the potentiating effect of genistein. The relaxation induced by forskolin, an activator of adenylyl cyclase, was potentiated by genistein or theophylline. In the presence of theophylline, the relaxation induced by forskolin was not further affected by genistein. Genistein also inhibited the activities of cAMP-PDE. In the presence of atenolol, but not ICI-118,551, iberiotoxin, an inhibitor of Ca-activated K channels, inhibited the relaxation induced by isoproterenol and the potentiating effect of genistein. In the presence of atenolol, quinacrine, an inhibitor of phospholipase A2, and metyrapone, an inhibitor of P-450 enzymes, but not alpha-naphthoflavone, an inhibitor of P-450 enzymes, indomethacin, a cyclooxygenase inhibitor, or AA861, a 5-lipoxygenase inhibitor, inhibited the potentiating effect of genistein. These results suggest that the potentiation of the beta1-adrenoceptor-induced relaxation by activation of genistein may mostly be due to inhibition of cAMP-PDE activities. In addition, the potentiation of the relaxation induced by activation of beta2-adrenoceptors by genistein may be related to the inhibition of arachidonic acid metabolism and cAMP-PDE activities.

Adrenergic beta-Agonists↗

PubMed2000

The inhibitory effect of KT3-671, a nonpeptide angiotensin-receptor antagonist, on rabbit and rat isolate vascular smooth muscles: a possible involvement of K(ATP) channels.

The vasoinhibitory effect of KT3-671, a recently synthesized nonpeptide angiotensin II (Ang II), AT1-receptor antagonist, and the factors affecting insurmountable antagonism of Ang II were examined in rabbit and rat isolated vascular smooth muscle preparations. In rabbit and rat aortic rings, KT3-671 caused insurmountable antagonism of Ang II. In addition, KT3-671 inhibited contractile responses to angiotensin III (Ang III). In rabbit isolated smooth muscles, KT3-671 was most effective in reducing the maximal contraction induced by Ang II in the renal artery followed by the basilar artery and the aorta. In rat renal arterial rings, KT3-671 (10(-5) M) inhibited the concentration-response curves of prostaglandin F2alpha and STA2. In rabbit and rat aortic rings without endothelium, the insurmountable antagonisms of Ang II by KT3-671 and EXP 3174 were changed to surmountable antagonism by pretreatment with DuP 753 and KT3-671, respectively. In addition, KT3-671 abolished the inhibitory effect of CV- 11974 in the rat aorta but not in the rabbit aorta. Indomethacin (10(-5) M) or the removal of endothelium did not affect the inhibitory effect of Ang II by CV-11974 or EXP 3174 but enhanced the insurmountable antagonism by KT3-671. ODQ (3 x 10(-6) M), N(G)-nitro-L-arginine (3 x 10(-4) M), 4-aminopyridine (3 x 10(-3) M), tetraethylammonium (TEA; 10(-3) M), or iberiotoxin (10(-7) M) did not affect the inhibitory action of KT3-671 or CV-11974. Methylene blue (3 x 10(-6) M), KCl (10(2) M), TEA (10(-2) M), or BaC12 (10(-4) M) changed the insurmountable antagonism by KT3-671 to surmountable antagonism and abolished the inhibitory effect of CV-11974. However, glibenclamide (3 x 10(-6) M) did not affect the inhibitory action of KT3-671 but reduced the insurmountable antagonism by CV- 11974. These results indicate that KT3-671 is an insurmountable antagonist of Ang II in the rabbit and rat aorta. The results in the rat aorta also suggest that K(ATP) channels may be involved in insurmountable antagonism of Ang II by KT3-671 and CV-11974. Key Words: KT3-671-Rabbit-Rat-Vascular smooth muscle-Angiotensin II-Insurmountable antagonist-K(TP)channels.

Angiotensin Receptor Antagonists↗

PubMed2000

Tumor necrosis factor-alpha represses androgen sensitivity in the LNCaP prostate cancer cell line.

PURPOSE: Prostate tumor progression is characterized by development of androgen independence and a heterogeneous distribution of the androgen receptor (AR). Tumor necrosis factor alpha (TNFalpha) has been demonstrated to contribute to the progression of several cancers and thus may play a role in prostate cancer progression. Accordingly, we examined if prostate cancers express TNFalpha and the effect of TNFalpha on androgen sensitivity and AR expression in LNCaP prostate cancer cells. MATERIALS AND METHODS: Immunohistochemical analysis of prostate tissues, ELISA, and northern blotting of LNCaP cell lines were carried out for detection of tumor necrosis factor-alpha (TNFalpha). To see the effect of TNFalpha on androgen receptor (AR), western blotting and northern blotting were performed after extraction of total protein and total RNA from LNCaP cells. Regulation of androgen-sensitivity by TNFalpha was investigated with cell proliferation assay and luciferase assay using PSA promoter after transfection of LNCaP cells. RESULTS: Immunohistochemical analysis demonstrated that TNFalpha protein was strongly expressed in epithelial cells of prostate cancer tissue but not in normal prostatic tissue. Basal level of TNFalpha in cell culture medium from LNCaP cells was very low. However, 12-O-tetradecanoylphorbol 13-acetate (TPA) induced TNFalpha secretion into medium up to 1600 pg/ml/day. Furthermore, 24 hr. post-TPA treatment TNFalpha mRNA levels were increased 15-fold compared to pre-treatment levels. TNFalpha (0 to 30 ng./ml. for 4 days) repressed AR protein and mRNA levels in a dose-dependent fashion in LNCaP cells. Pre-treatment of cells with actinomycin D treatment revealed that repression of mRNA levels was exerted at the post-transcriptional level. TNFalpha inhibited the ability of 10-9 M dihydrotestosterone (DHT) to induce LNCaP cell proliferation and activation of the prostate specific antigen (PSA) gene promoter. This inhibition was partially reversed by overexpression of transgenic androgen receptor. CONCLUSIONS: TNFalpha is present and inducible in prostate cancer cells and short-term TNFalpha diminishes androgen-sensitivity in LNCaP cells through down-regulation of AR protein and mRNA levels. These results suggest that TNFalpha may play a role in the initiation of an androgen-independent state in prostate cancer through its ability to inhibit AR sensitivity in prostate cancer.

Antibiotics, Antineoplastic↗

PubMed2000

Effect of long-term atrial-demand ventricular pacing on cardiac sympathetic activity.

It has been shown that either dual-chamber or atrial pacing may be better than ventricular single-chamber pacing, but the long-term effect of dual-chamber pacing on cardiac sympathetic activity is unclear. The aim of this study was to assess the effect of long-term dual-chamber pacing on cardiac sympathetic activity, compared with atrial pacing and unpaced individuals. We studied 11 patients with dual-chamber pacemakers (Group D), nine with atrial single-chamber pacemakers (Group A) over the long term (mean 44 +/- 36 months) and 10 normal individuals without cardiac pacing. All underwent myocardial 123I-metaiodobenzylguanidine (MIBG) imaging to assess cardiac sympathetic activity. The heart-to-mediastinum (H/M) MIBG uptake ratio and the MIBG washout rate from the myocardium were calculated. Echocardiography was performed in all patients with cardiac pacing to assess left ventricular function. In Group D, the H/M ratio on delayed images was significantly lower than that of Group A (1.82 +/- 0.51 vs 2.56 +/- 0.50, P < 0.001) and normal individuals (2.65 +/- 0.35, P < 0.05). The myocardial MIBG washout rate of Group D was significantly higher than that of either Group A (52 +/- 13% vs 36 +/- 8%, P < 0.01) or normal individuals (31 +/- 7%, P < 0.05). Neither the H/M ratio nor MIBG washout rate differed significantly between patients in Group A and normal individuals. Furthermore, the echocardiographic parameters did not differ significantly between the two pacing groups. We conclude that long-term ventricular pacing, even in the presence of atrioventricular synchrony, accelerates cardiac sympathetic activity without deteriorating left ventricular function.

3-Iodobenzylguanidine↗

PubMed2000

15N and 1H NMR study of histidine containing protein (HPr) from Staphylococcus carnosus at high pressure.

The pressure-induced changes in 15N enriched HPr from Staphylococcus carnosus were investigated by two-dimensional (2D) heteronuclear NMR spectroscopy at pressures ranging from atmospheric pressure up to 200 MPa. The NMR experiments allowed the simultaneous observation of the backbone and side-chain amide protons and nitrogens. Most of the resonances shift downfield with increasing pressure indicating generalized pressure-induced conformational changes. The average pressure-induced shifts for amide protons and nitrogens are 0.285 ppm GPa(-1) at 278 K and 2.20 ppm GPa(-1), respectively. At 298 K the corresponding values are 0.275 and 2.41 ppm GPa(-1). Proton and nitrogen pressure coefficients show a significant but rather small correlation (0.31) if determined for all amide resonances. When restricting the analysis to amide groups in the beta-pleated sheet, the correlation between these coefficients is with 0.59 significantly higher. As already described for other proteins, the amide proton pressure coefficients are strongly correlated to the corresponding hydrogen bond distances, and thus are indicators for the pressure-induced changes of the hydrogen bond lengths. The nitrogen shift changes appear to sense other physical phenomena such as changes of the local backbone conformation as well. Interpretation of the pressure-induced shifts in terms of structural changes in the HPr protein suggests the following picture: the four-stranded beta-pleated sheet of HPr protein is the least compressible part of the structure showing only small pressure effects. The two long helices a and c show intermediary effects that could be explained by a higher compressibility and a concomitant bending of the helices. The largest pressure coefficients are found in the active center region around His15 and in the regulatory helix b which includes the phosphorylation site Ser46 for the HPr kinase. This suggests that this part of the structure occurs in a number of different structural states whose equilibrium populations are shifted by pressure. In contrast to the surrounding residues of the active center loop that show large pressure effects, Ile14 has a very small proton and nitrogen pressure coefficient. It could represent some kind of anchoring point of the active center loop that holds it in the right place in space, whereas other parts of the loop adapt themselves to changing external conditions.

Amides↗

PubMed2000

IL-4 is required for defense against mycobacterial infection.

Although the involvement of T helper (Th1) cells is central to protection against intracellular bacteria, including Mycobacterium tuberculosis, the involvement of Th2 cells, characterized by potent interleukin (IL)-4 secretion in mycobacterial infection is still unclear. In order to clarify the role of IL-4 in murine tuberculosis, IL-4-deficient mutant mice, IL-4 knockout (IL-4 KO) mice, were utilized. The mice were infected with H37Rv, Kurono or BCG Pasteur via an airborne infection route by placing them in the exposure chamber of a Middlebrook airborne infection apparatus. Their capacity to control mycobacterial growth, granuloma formation, cytokine secretion, and nitric oxide (NO) production were examined. These mice developed large granulomas, but not necrotic lesions in the lungs, liver or spleen (P<0.05). This was consistent with a significant increase in lung colony-forming units (CFU). Compared with levels in wild-type mice, upon stimulation with mycobacteria, splenic IL-10 levels were low and IL-6 levels were intermediate, but interferon (IFN)-gamma and IL-12 levels were significantly higher. IL-18 levels were within the normal range. The level of NO production by alveolar macrophages of the IL-4 KO mice was similar to that of the wild-type mice. Granulomatous lesion development by IL-4 KO mice was inhibited significantly by treatment with exogenous recombinant IL-4. These findings were not specific to the IL-4 KO mice used. Our data show that IL-4 may play a protective role in defense against mycobacteria, although IFN-gamma and TNF-alpha play major roles in it. Our data do not rule out an IFN-gamma-independent function of IL-4 in controlling tuberculosis.

Animals↗

PubMed2000

Enhancing effect of an inhibitor of nitric oxide synthesis on bacillus Calmette-Guerin-induced macrophage cytotoxicity against murine bladder cancer cell line MBT-2 in vitro.

We studied the effect of an inhibitor of nitric oxide (NO) synthesis, N(G)-monomethyl-L-arginine (L-NMMA), on the Bacillus Calmette-Guérin (BCG)-induced antitumor activity of murine peritoneal exudate cells (PEC) against murine bladder cancer cell line MBT-2 in vitro. L-NMMA enhanced BCG-induced cytotoxic activity of PEC, as well as interferon (IFN)-gamma and tumor necrosis factor (TNF)-alpha production. The L-NMMA-induced enhancement was due to the prolonged survival of BCG in macrophages, because no enhancement of cytotoxicity was observed and neither IFN-gamma nor TNF-alpha production was significantly enhanced by killed BCG. Anti-TNF-alpha antibody (Ab) and anti-IFN-gammaAb reduced the L-NMMA-induced enhancement of the cytotoxicity. The depletion of T cells from PEC reduced the production of both IFN-gamma and TNF-alpha, as well as the enhancement of cytotoxicity induced by viable BCG plus L-NMMA. These results suggest that L-NMMA has an enhancing effect on BCG-induced macrophage cytotoxicity and the enhancement is partially mediated by T cells and their soluble products. Accordingly, NO inhibitor should be a valuable adjunct to BCG immunotherapy for bladder cancer.

Animals↗

PubMed2000

The association of transforming growth factor-beta 1 with myometrial invasion of endometrial carcinomas through effects on matrix metalloproteinase.

OBJECTIVE: The association of transforming growth factor-beta 1 (TGF-beta 1) with a matrix metalloproteinase (MMP) and a tissue inhibitor of metalloproteinase (TIMP), as well as myometrial invasion of endometrial cancer was studied. METHODS: The effects of TGF-beta 1 on cellular invasiveness, gelatinase activity, and expression of TIMP-1 were examined in 2 endometrial adenocarcinoma cell lines, KLE and Ishikawa. Plasma was obtained from 8 endometrial cancer patients with Stage-Ia disease, from 6 with Stage-Ib disease, and from 4 with Stage-Ic disease, and the levels of TGF-beta 1 were measured by enzyme immunoassays. The immunohistochemical expression of MMP-9, TIMP-1, TGF-beta 1, and TGF-beta receptor Type I in tumor tissue from the same patients also was detected. RESULTS: Invasiveness, gelatinase activity, and the expression of TIMP-1 were higher in KLE cells than in Ishikawa cells, and they were increased by treatment with rTGF-beta 1. The expression of TGF-beta receptor Type I was higher in KLE cells than in Ishikawa cells, which were unresponsive to exogenous TGF-beta 1. The plasma levels of TGF-beta 1 were greater in Stage-Ib and Stage-Ic patients than in Stage-Ia patients. MMP-9 and TGF-beta receptor Type I were expressed mainly in tumor cells, while TIMP-1 and TGF-beta 1 were localized in both tumor epithelial cells and stromal cells. MMP-9 and TIMP-1 were expressed only in Stage-Ib and Stage-Ic patients, although TGF-beta 1 and TGF-beta receptor Type I were ubiquitous. CONCLUSIONS: Myometrial invasion of endometrial cancers involves an increase in gelatinase activity, regulated to some extent by TGF-beta 1 in an autocrine or paracrine fashion.

Adenocarcinoma↗