Tumour necrosis factor-alpha-mediated human polymeric immunoglobulin receptor expression is regulated by both mitogen-activated protein kinase and phosphatidylinositol-3-kinase in HT-29 cell line.

Human polymeric immunoglobulin receptor (pIgR) is present on the surface of glandular epithelium, and it plays a crucial role in the mucosal immune defence. pIgR expression in HT-29 cells is up-regulated by one of the proinflammatory cytokines, tumour necrosis factor (TNF)-alpha. However, the mechanism used by the TNF-alpha-mediated signalling pathway has not been examined exclusively. To elucidate this mechanism in detail, HT-29 cells were cotreated with TNF-alpha and mitogen-activated protein kinase kinase (MAPKK, also called MEK1) inhibitor, PD98059, and the amount of free secretory component (SC) secreted into the culture medium was measured. The amount of free SC stimulated by TNF-alpha was increased by addition of PD98059. This up-regulation occurred at the transcriptional level. The amount of SC was also up-regulated by addition of TNF-alpha with U0126, an inhibitor of MEK1 and MEK2. Nuclear factor (NF)-kappaB activity and NF-kappaB binding to the kappaB2 site localized upstream of the pIgR gene did not change after coincubation of HT-29 cells with TNF-alpha and PD98059. The expression level of pIgR by TNF-alpha was decreased by LY294002, an inhibitor of phosphatidylinositol-3-kinase (PI3K), at the transcriptional level. Extracellular signal-regulated kinase (ERK)1/2 phosphorylation and NF-kappaB binding to the kappaB2 site were not affected by LY294002 treatment. These data suggest that TNF-alpha-mediated pIgR expression is negatively regulated by ERK pathway, which is independent of NF-kappaB. In addition, decrease of SC production by Ly294002 suggests that the presence of PI3K mediated regulation of SC production.

Bradykinin B(2) receptor gene polymorphism is associated with angiotensin-converting enzyme inhibitor-related cough.

The appearance of cough in association with angiotensin-converting enzyme (ACE) inhibitors is thought to be related to bradykinin, and it has been speculated that the elicitation of adverse effects is genetically predetermined. Several polymorphisms of the human bradykinin B(2) receptor gene may be involved in ACE inhibitor-related cough. To investigate this possibility, we identified the -58 thymine (T)/cytosine (C) polymorphism in subjects with ACE inhibitor-related cough. We classified the study population into 4 groups: subjects with and without cough that were treated with ACE inhibitors (n=30/30), nontreated essential hypertensive subjects (n=100), and normotensive subjects (n=100). The -58T/C was genotyped by the polymerase chain reaction single-strand conformation polymorphism method. The frequencies of the CC genotype and C allele of -58T/C were significantly higher in the nontreated hypertensive subjects than in the normotensive subjects. Conversely, the frequencies of the TT genotype and T allele were significantly higher in the subjects with cough than in the subjects without cough. These tendencies were more pronounced in females. Among the promoter assays of the human bradykinin B(2) receptor, -58T was found to have a higher transcription rate than that of -58C. This finding seems to suggest that the transcriptional activity of promoter might be involved in the appearance of ACE inhibitor-related cough. A genetic variant of the bradykinin receptor is involved in the elicitation of ACE inhibitor-related cough. It may be possible to predict the side effects of ACE inhibitors in advance.

Association of polymorphisms of the renin-angiotensin system and bradykinin B2 receptor with ACE-inhibitor-related cough.

The use of angiotensin-converting enzyme (ACE) inhibitors to treat hypertension has recently increased. However, their use is associated with a persistent dry cough in a significant percentage of such patients. The present study was designed to assess the contribution of polymorphisms as a genetic marker of ACE-inhibitor-related cough in a Japanese hypertensive population. Genotyping was carried out in 190 patients, 70 with cough and 120 without cough, who had been treated with ACE inhibitors. Polymorphisms of ACE insertion/deletion (I/D), angiotensin II type 1 receptor (1166A/C), type 2 receptor (3123C/A), and bradykinin B2 receptor (-58T/C, exon 1, I/D), were analyzed in these subjects. The TT genotype and T allele of bradykinin B2 receptor (-58T/C) were identified at a significantly higher frequency in the cough (+) patients than in the cough (-) patients. This difference was even more pronounced in women. However, there was no significant relationship between polymorphisms of ACE, angiotensin II receptors, or bradykinin B2 receptor exon 1, and occurrence of ACE-inhibitor-related cough. The transcriptional activity of the bradykinin B2 receptor promoter is involved in the occurrence of cough, and this new marker may provide a valuable tool to detect patients at risk of developing this side effect of ACE inhibitors. In conclusion, Susceptibility to develop cough is associated with a genetic variant of the bradykinin B2 receptor promoter; thus, it may be possible to identify those patients who will develop this adverse reaction to ACE inhibitors in advance.

[The case studies on oral administration of tegafur encapsulated with slow releasing granule (SF-SP) for three rectal patients].

Three rectal cancer patients with partial response (PR) were obtained by oral administration of 800-1200 mg/day of SF-SP. One of them showed I-A grade and other I-B in Karnofsky evaluation.

Impairment of mitochondrial respiratory activity of non-infarcted myocardium under severe pump failure in acute myocardial infarction.

We studied alterations in respiratory activity of mitochondria (Mt) in non-infarcted myocardium (NIZ) under severe pump failure complicated by acute myocardial infarction (AMI). Dogs in which AMI was induced were divided into two groups; one in which left ventricular systolic pressure (LVPs) was maintained higher than 70% of preligation level (ND group); and one in which LVPs was diminished to less than 70% (D group). Regional myocardial blood flow (MBF) in NIZ reduced significantly in proportion to decreases in LVPs and cardiac output (CO). State -III activity and RCR decreased in proportion to reductions in MBF, LVPs, and CO in Mt from NIZ of D group. Complex-I and DNP-stimulated ATPase activities were also reduced in NIZ of D group. Morphologic studies revealed slight swelling and fusion of mitochondria in NIZ cells of D group, but no changes such as the appearance of a dense deposit indicating ischemic damage were seen. Pump failure in AMI is likely to be caused partly by impaired function of Mt in NIZ induced by hypoperfusion. Improvement of metabolic impairment in NIZ is important in the treatment of pump failure.

[Experimental study on the effectiveness of BRM in radiation therapy--OK-432 and cyclophosphamide].

We have previously demonstrated that specific antitumor cell-mediated immunity in host mice was induced or enhanced by local irradiation to tumor. In this study, the immunological status of host mice with transplanted tumor were modified by using OK-432 as immunostimulator and cyclophosphamide as suppressor of suppressor T-cells before and/or after local irradiation to tumor. The results indicate that the combined use of immunomodifier and radiation is more effective in the tumor radiotherapy. Three factors as follows: the interval of drug and radiation, the times of treatments and dose of immunostimulator and immunosuppressor, are very important in these combined therapy.

Study on reperfusion injury on sarcoplasmic reticulum in acute myocardial ischemia.

Reperfusion injury in early myocardial ischemia was studied in the dog with special reference to sarcoplasmic reticulum (SR) and contraction bands. Acute myocardial ischemia (I) was induced by occlusion of the left anterior descending coronary artery (LAD) for 10, 20 and 30 min followed by reperfusion for 15 min (R). Ca(++)-ATPase activity of SR in 10-min-R-Group was significantly reduced to 60% of control activity, but activity of 10-min-I-Group remained near the control level in subendomyocardium (Endo). ATPase activity in 30-min-I-Group diminished to 60% of control activity in Endo and it was similar for 30-min-R-Group. In ischemic myocardium, composition of major ATPase protein decreased significantly in 30-min-I-Group and similar reduction was observed in 20-min-R-Group in Endo. In morphology proportion of appearance of contraction bands in Endo was significantly increased in 20-min or longer-R-Groups. These results suggest that reperfusion injury is likely to occur when coronary artery is reflowed after 10 min of ischemia. This may be caused by increased intracellular Ca++ at a very early stage of reperfusion period, and reperfusion injury may be induced due to acceleration in the necrotic process of the membrane system in the myocytes during ischemia.

Mitochondrial respiratory impairment in streptozotocin-induced diabetic rat heart.

The purpose of this study was to examine mitochondrial respiratory impairment in the diabetic heart. Diabetes mellitus was induced in male Wistar rats by intravenous injection of streptozotocin (STZ) for 2 to 16 weeks (Group D). In some of the diabetic rats, insulin was injected for 2 or 3 weeks prior to sacrifice (Group I). Fasting blood glucose was markedly elevated to greater than 300 mg/dl in Group D and was similar to normal glucose levels in Group I. At 2 weeks after STZ injection, state 3 was only 59.1% of that in the control. Complex I and complex V activities were also significantly reduced to 43.4% and 71.7% of those in the control, respectively. These reductions recovered with insulin treatment. This phenomenon persisted for 16 weeks. Morphological studies revealed swelling of the mitochondria and an increase in lipid droplets in diabetic cardiomyocytes, and these were also improved with insulin treatment. We conclude that in the diabetic heart, disturbance of energy production in cardiac mitochondria is generated by the impairment of oxidative phosphorylation due to depression of complex I and complex V. These changes may contribute the cardiac dysfunction that is a complication of diabetes mellitus.

The genetic factor in acute myocardial infarction with hypertension.

This study assessed the contribution of polymorphisms of angiotensin II (AngII) receptors and bradykinin B2 (BK-B2) receptor to hypertension and acute myocardial infarction (AMI) in a Japanese population: 150 subjects with essential hypertension, 150 subjects with AMI with/without hypertension, and 150 healthy, age- and sex-matched controls. Polymorphisms of the AngII type 1 receptor (1166 A/C) and type 2 receptor (3123 C/A), and the BK-B2 receptor (-58T/C, exon 1) were analyzed and significant differences of genotypes and allelic frequencies in the AngII type 2 receptor C/A and BK-B2 receptor -58T/C were found between the essential hypertension and control subjects. Further, a significantly higher incidence of the C allele of the BK-B2 receptor was seen in AMI subjects with hypertension compared with those without hypertension. Genetic variations in the AngII and BK-B2 receptors could prove to be significant pathophysiological mechanisms affecting essential hypertension and AMI, and genetic differences appear to be a new risk factor for these conditions.

Protective effect of captopril on ischemic myocardium.

The protective effect and mechanism of action of the angiotensin-converting enzyme inhibitor (ACE-I) captopril was investigated in organelles from ischemic myocardial cells in a canine coronary ligation model. Sarcoplasmic reticulum (SR) and mitochondrial fractions were extracted from ischemic and nonischemic myocardial cells from captopril- and saline-treated (control) hearts. Heart rate, cardiac output, and right ventricular systolic blood pressure were similar in the captopril-treated and control groups. Left ventricular systolic blood pressure (LVPs) decreased gradually to 89% of the baseline value after captopril administration, and to 78% of the baseline value after ligation. Ca-ATPase activity in the SR, the respiratory control ratio (RCR) in the mitochondria, and dinitrophenol (DNP)-stimulated ATPase activity were significantly higher in ischemic myocardium from the captopril-treated group than from the saline-treated (control) group. The SH group content of both organelles was higher in the captopril-treated group. Our results suggest that, in addition to their hemodynamic effects, ACE-I agents containing SH groups protect the myocardium from ischemic damage by preventing enzyme oxidation.

Congestive heart failure is associated with the rate of bone loss.

AIMS: To characterize relationships between mineral homeostasis, bone turnover, bone mass, and congestive heart failure (CHF), we evaluated 75 women with mild to moderate CHF. METHODS AND RESULTS: We examined the association in annual rate of change in spinal bone mineral density (BMD) with polymorphism of the vitamin D receptor (VDR) gene. Compared with the control group, the CHF group had reduced left ventricular ejection fraction (LVEF: 68.2 +/- 7.5% vs. 60.2 +/- 12.9%; P = 0.0249), human atrial natriuretic peptide (hANP) was elevated (hANP: 10.7 +/- 4.7 pmol L-1 vs. 25.8 +/- 24.2 pmol L-1; P = 0.001) and had lower peak VO2 (22.3 +/- 7.5 mL kg-1 min-1 vs. 15.8 +/- 7.4 mL kg-1 min-1; P = 0.0429). The CHF patients with the VDR FF genotype had a significantly high annual rate of decrease in BMD. In the CHF patients with the VDR FF genotype, urinary calcium excretion (FECa) was elevated (1.40 +/- 0.91% vs. 2.39 +/- 1.40%; P = 0.028), and serum bone-type alkaline phosphatase (B-ALP) was reduced (62.6 +/- 13.7 IU L-1 vs. 47.0 +/- 18.6 IU L-1; P = 0.0123). Also, FECa was correlated positively with furosemide dose (R = 0.881; P = 0.0087) and hANP concentrations (R = 0.635; P = 0.0147) and negatively with DeltaBMD (R = 0.72; P = 0.044) in the CHF patients with the VDR FF genotype. CONCLUSION: The CHF patients with the VDR FF genotype have higher rates of bone loss. These patients may need to increase their calcium intake and BMD may need to be followed more carefully over time.

The effects of dopamine, dobutamine and amrinone on mitochondrial function in cardiogenic shock.

The impairment of mitochondrial in non-infarcted myocardium under cardiogenic shock complicated by acute myocardial infarction was studied. We induced acute myocardial infarction in dogs by ligating the circumflex branch of the left coronary artery (LCX). On basis of left ventricular systolic pressure (LVPs) after 60 minutes, we divided the dogs into two groups: a group in which LVPs fell to below 70% of the pre-LCX ligation level, and a Control group in which LVPs remained more than 90%. The former group was further divided into four subgroups, depending on infusion of dopamine, dobutamine, amrinone or saline after 90 minutes. Mitochondria were prepared and mitochondrial respiratory activity determined. In the Saline group, hemodynamics became reduced to less than 70% of the preligation level after 120 minutes, however, in the Dopamine and Dobutamine groups, hemodynamics became restored to the preligation level. In the Amrinone group, LVPs decreased slightly, while cardiac output, LV Max. dp/dt and myocardial blood flow increased. In the Saline group, mitochondria in the non-infarcted myocardium functioned at a lower level of activity than that of the Control group. However, in the Dopamine, Dobutamine, and Amrinone groups, the mitochondria functioned at a higher level. Electron microscopy revealed mitochondrial damage in the Saline group only. The results indicate that an energy production disorder in the non-infarcted myocardium may have pathogenetic implications in cardiogenic shock associated with acute myocardial infarction, while dopamine, dobutamine, and amrinone improve mitochondrial function, and ultimately improve cardiac function.

Genetic background in patients with acute myocardial infarction.

The renin-angiotensin system is believed to play important roles in the development of acute myocardial infarction, and gene polymorphisms may also be involved. To investigate the genetic background in patients with acute myocardial infarction, we performed a case control study in a Japanese population. The study included 150 patients with acute myocardial infarction and 150 healthy, age- and sex-matched controls. We examined polymorphisms of angiotensin II type 1 receptor (1 166 A / C), type 2 receptor (3123 C / A), and bradykinin B2 receptor (-58 T / C) in these subjects. The allelic frequencies of angiotensin II type I receptor C and angiotensin II type 2 receptor A were significantly higher in the acute myocardial infarction subjects than in the control subjects, and this tendency was more significant in the younger patients. The combined ratios of angiotensin II type 1 receptor C and type 2 receptor A alleles in patients under 64 years old were significantly higher than in their older counterparts. However the total numbers of conventional coronary risk factors (hypertension, hypercholesterolemia, diabetes mellitus, and smoking) in individual subjects were not significantly different between younger and older patients. These polymorphisms were found to be involved in the development of acute myocardial infarction, particularly in the younger patients, and it was concluded that the incidence of acute myocardial infarction might be reduced by management from the genotypes.

Promoter polymorphism of the beta2 bradykinin receptor gene is associated with essential hypertension.

The present study examined the genetic contribution of the human beta2 bradykinin receptor gene in Japanese subjects with essential hypertension, and identified a -58T/C polymorphism of the core promoter that might be responsible for essential hypertension in Japanese. The study consisted of 100 hypertensive subjects and 100 age- and sex-matched controls. The allelic frequencies were 0.575 for the C allele and 0.425 for the T allele in hypertensive subjects, and 0.465 for the C allele and 0.535 for the T allele in normotensive subjects. The allelic frequencies were in Hardy-Weinberg equilibrium. Significant differences between hypertensive and normotensive subjects were seen in the genotypes distribution (p=0.049) and allelic frequencies (p=0.028), and the beta2 bradykinin receptor gene variant was associated with human essential hypertension in this Japanese population. This new marker may provide a valuable tool for assessing the risk for putative bradykinin-associated common diseases, such as hypertension, and cardiovascular diseases with genetic determinism. These results suggest that the -58 polymorphism of the human beta2 bradykinin receptor gene is an independent risk factor for essential hypertension in the Japanese population.

Generation of free radicals and the damage done to the sarcoplasmic reticulum during reperfusion injury following brief ischemia in the canine heart.

Free radical generation was studied by the electron spin resonance (ESR) technique using alpha-phenyl N tert butyl nitrone (PBN) in a brief ischemia-reperfusion model of the canine heart, and correlated with biochemical changes of the sarcoplasmic reticulum (SR). ESR spectra (aH=0.3-0.4mT, aN=1.43-1.58mT) were observed as PBN spin adducts, which peaked at levels 5-fold above the control levels at 5 min after reperfusion. The simulated coupling constants of PBN spin adducts suggested that the sample should contain at least 2 carbon-centered radicals at 5 min after reperfusion (radical A: aH=0.350mT, aN=1.485mT; radical B: aH=0.370mT, aN=1.615 mT). At this time point, a significant reduction in Ca-ATPase activity of the SR was found without degradation of the major ATPase protein. Superoxide dismutase (SOD) significantly reduced the intensity of the PBN spin adduct signals and preserved the Ca-ATPase activity of the SR to 80% of the control level. Reperfusion injury after brief ischemia may be the result of inactivation of intracellular Ca-ATPase by free radicals generated during reperfusion, and SOD contributes to the protective effect by scavenging the radicals.