Biomedical subjects
Publications and source records attributed to G Watanabe.
A 68 year-old woman underwent replacement of the descending thoracic aorta with a cryopreserved aortic homograft for the treatment of a Stanford B dissecting aneurysm. After surgery, the patient made an uneventful recovery without recurrent infection. In such cases, the use of aortic homografts to replace aortic segments is a promising and effective therapeutic option.
BACKGROUND: Platelets play pivotal roles in coronary thrombosis, and antiplatelet therapies are widely used for coronary artery disease (CAD). However, the effects of genetic variation in platelets on CAD are poorly understood. We have assessed the association between CAD and polymorphisms in a platelet receptor for von Willebrand factor, the glycoprotein (GP) Ib/IX complex, which mediates shear stress-dependent platelet activation. METHODS AND RESULTS: Genotypes of the alpha-chain of the receptor (GP Ib alpha, 145Thr/Met) were determined in 91 patients with myocardial infarction (MI) or angina pectoris whose lesions were confirmed by coronary angiography as well as in 105 individuals from the general population with no history of angina or other heart diseases and normal resting ECGs. There was no homozygote for Met/Met in either the control or patient groups. The prevalence of the Thr/Met genotype (T/M) in all patients was not significantly different from that in the control group. However, the frequency of T/M was significantly higher in patients aged < or = 60 years (31.8%) than in control subjects aged < or = 60 years (16.0%; P<.05, odds ratio=2.5). An association was also demonstrated between CAD and the other polymorphism of GP Ib alpha, a variable number of tandem repeats of a 13-amino acid sequence, which is known to be linked to the 145Thr/Met polymorphism. There was an association between the frequency of the T/M genotype and the angiographic severity of CAD: 11.1% for Gensini score < 40 versus 50.0% for Gensini score > or = 40 (P=.0015). There was no difference in the distribution of GP Ib alpha genotypes between patients with MI and those with angina pectoris. CONCLUSIONS: This study suggests that the presence of the Met allele in GP Ib alpha is a risk factor for the prevalence and severity of CAD in individuals aged < or = 60 years. The results need to be confirmed in a large-scale study of incident case subjects and matching control subjects.
We have analyzed a mutation responsible for a Hinf I polymorphism of the D1S80 system by Taq dye deoxy terminator cycle sequencing. A G-T transversion at the 58th nucleotide from the 5' end of the forward primer was identified. This mutation has been firmly established by the restriction enzymes. The type G was digested with Hinf I but not with Tsp 509 I. Conversely, the type T was digested with Tsp 509 I but not with Hinj I. We applied this technique to analyze D1S80 system in German and Japanese populations. In the Japanese population, the type T frequencies were much lower than those in the German population, and the type T was almost exclusively observed in the allele 24.
A broad array of stressors induce ACTH release from the anterior pituitary, with consequent stimulation of the adrenal cortex and release of glucocorticoids critical for survival of the animal. ACTH stimulates adrenocortical gene expression in vivo and inhibits adrenocortical cell proliferation. Binding of ACTH to its G-protein-coupled receptor stimulates the production of cAMP and activation of the protein kinase A pathway. The stress-activated protein kinases (SAPKs) (or c-Jun N-terminal kinases) and the extracellular signal-regulated kinases (ERKs) are members of the mitogen-activated protein kinase family of serine/threonine kinases, which have recently been implicated in G-protein-coupled receptor intracellular signaling. The SAPKs are preferentially induced by osmotic stress and UV light, whereas the ERKs are preferentially induced by growth factors and proliferative signals in cultured cells. In these studies, ACTH stimulated SAPK activity 3-4-fold both in the adrenal cortex in vivo and in the Y1 adrenocortical cell line. 12-O-Tetradecanoylphorbol-13-acetate but not cAMP induced SAPK activity in Y1 cells. The isoquinolinesulfonamide inhibitors H-8 and H-89 blocked ACTH induction of SAPK activity at protein kinase C inhibitory doses but not at protein kinase A inhibitory doses. The calcium chelating agent EGTA inhibited ACTH-induced SAPK activity and the calcium ionophore A23187 induced SAPK activity 3-fold. In contrast with the induction of SAPK by ACTH, ERK activity was inhibited in the adrenal cortex in vivo and in Y1 adrenal cells. Together these findings suggest that ACTH induces SAPK activity through a PKC and Ca+2-dependent pathway. The induction of SAPK and inhibition of ERK by ACTH in vivo may preferentially regulate target genes involved in the adrenocortical stress responses in the whole animal.
Rho small GTPase regulates cell morphology, adhesion and cytokinesis through the actin cytoskeleton. We have identified a protein, p140mDia, as a downstream effector of Rho. It is a mammalian homolog of Drosophila diaphanous, a protein required for cytokinesis, and belongs to a family of formin-related proteins containing repetitive polyproline stretches. p140mDia binds selectively to the GTP-bound form of Rho and also binds to profilin. p140mDia, profilin and RhoA are co-localized in the spreading lamellae of cultured fibroblasts. They are also co-localized in membrane ruffles of phorbol ester-stimulated sMDCK2 cells, which extend these structures in a Rho-dependent manner. The three proteins are recruited around phagocytic cups induced by fibronectin-coated beads. Their recruitment is not induced after Rho is inactivated by microinjection of botulinum C3 exoenzyme. Overexpression of p140mDia in COS-7 cells induced homogeneous actin filament formation. These results suggest that Rho regulates actin polymerization by targeting profilin via p140mDia beneath the specific plasma membranes.
While laparoscopic cholecystectomy is a standard therapeutic option for gallbladder stones, it is associated with a significant risk of injury to the gallbladder wall, which may result in the dispersion of free stones within the peritoneal cavity. However, the incidence and consequences of these dropped stones remains unclear. We report herein the cases of three patients in whom abdominal abscesses developed as a result of dropped stones during this procedure. Of particular interest was the relatively long interval from the procedure to the onset of symptoms and the unusual progression of the inflammation. These case reports strongly reinforce the risk of dropped stones during laparoscopic cholecystectomy.
We treated a case of paraesophageal hiatus hernia by laparoscopic repair. The procedure included a reduction of the gastric fundus and duodenal bulbus, closure of the diaphragmatic defect, mesh wrapping of the closure, gastropexy to the diaphragm, and a gastrostomy. Preoperative monitoring of the pH for 24 h showed no reflux. Intraoperative intraluminal manometry of the esophagus after hernia reduction showed the pressure of the lower esophageal sphincter to be normal, and thus an antireflux procedure was not deemed to be necessary. The patient was put on a soft diet from postoperative day 2. A postoperative upper gastrointestinal series showed no gastroesophageal reflux. No complications or recurrence of the hiatus hernia have been observed in the 12 months since the operation. Laparoscopic repair of a paraesophageal hiatus hernia with normal pressure of the lower esophageal sphincter, so that fundoplication is not needed, is thus considered to be possible.
The immunostimulant tumor necrosis factor-alpha (TNF alpha), produced by monocytes/macrophages in response to inflammatory disorders, regulates gene expression in part through induction of mitogen-activated protein kinases (MAPKs), including the stress-activated protein kinase (SAPK) (c-Jun N-terminal kinase [JNK]) and the extracellular signal-regulated kinases (ERKs). In testicular Leydig cells, the induction of steroidogenesis by cAMP is inhibited by TNF alpha. To examine the potential mechanisms governing the mutual inhibition between cAMP and TNF alpha in Leydig cells, the intracellular signaling pathways that contribute to AP-1-dependent gene expression were examined in the mouse MA-10 Leydig cell line. TNF alpha induced SAPK activity sixfold at 15 min, and the PKC inhibitor calphostin C reduced the induction of SAPK by 30%. cAMP induced SAPK activity twofold but reduced TNF alpha-induced SAPK activity. ERK activity was inhibited by both cAMP and TNFa. TNFa increased c-Jun protein, but only weakly induced FOS proteins (c-Fos, FosB, Fra-1, and Fra-2) whereas cAMP increased the abundance of several FOS proteins (c-Fos, FosB, Fra-1, and Fra-2), with little effect on c-Jun levels. AP-1 binding activity, assessed using electrophoretic mobility shift assays, was increased twofold by TNF alpha and fivefold by cAMP. Cyclic AMP alone induced AP-1-responsive reporter (p3TPLUX) activity threefold after 2 h with peak effect of 4-fold at 4 hr. AP-1 reporter was not induced by TNF alpha alone but in the presence of cAMP, TNF alpha induced AP-1 reporter activity 12-fold. In conclusion, TNF alpha and cAMP induce distinct components that separately contribute to the modulation of AP-1 activity in MA-10 cells.
We present a simple rapid reproducible polymerase chain reaction based technique, termed amplified product length polymorphism (APLP), as a new strategy for primer design for ABO genotyping. The method involves the use of primers differing in length and permits the identification of the major ABO genotypes (A1, A2, B, OA, OG, and O2) according to the molecular size of the allele-specific amplified products. Ten different primers designed to analyze the variations in nucleotide positions 261, 297, 796, 802, and 1059-1061 of cDNA are mixed in one reaction, and the amplified products are resolved on a polyacrylamide gel. Of eight PCR fragments (132 bp, 120 bp, 108 bp, 98 bp, 88 bp, 80 bp, 72 bp, and 64 bp), two to five are amplified in the reaction according to ABO genotypes. The new technique has been successfully applied to the genotyping of 221 peripheral blood samples from Japanese and Germans whose ABO phenotypes had previously been determined; a novel A allele (AG) was found in Japanese individuals.
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The effects of acute neutralization of endogenous inhibin on ovulation rate and circulating FSH levels were investigated. Nine or ten days after estrus, 5 heifers were given a single injection of 75 ml iv inhibin antiserum produced in a castrated male goat, while another 5 were given the same amount of a castrated male goat serum. All heifers were given injections of PGF2alpha im at 48 h and 60 h after the serum injection. Those exhibiting an estrus were artificially inseminated with frozen-thawed semen. Seven or eight days after the insemination, ova or embryos were collected using a non-surgical method. Administration of inhibin antiserum resulted in a significant increase in the number of medium-sized follicles compared with the number in the control animals. The number of large follicles in the inhibin-neutralized animals was 4.8 +/- 2.4 (mean +/- SEM; n = 5) on the day of estrus, while there was a single large follicles in the ovaries of control animals. Seven or eight days after estrus, 3 to 16 ova or embryos were recovered from 4 of 5 animals, and 64 % of the total ova/embryos were transferable. Administration of inhibin antiserum produced a significant increase in the concentrations of plasma FSH from 12 to 72 h after the serum injection compared with the levels in the control animals (P < 0.05). After the onset of estrus, preovulatory LH and FSH surges were noted in inhibin-neutralized animals and magnitude of the rise in each hormone was similar to the control animals. The present study demonstrates that a single injection of the inhibin antiserum induces multiple ovulations probably by enhancing FSH secretion, and that recovery of embryos is equal to that observation after an ordinary FSH treatment.
This study was carried out to examine the involvement of inhibin in the regulation of FSH secretion during the growth phase of the dominant follicle in the early luteal phase of cows. Six cows were given a single i.v. bolus injection of 100 ml inhibin antiserum raised against bovine 32-kDa inhibin in a castrated male goat, and five animals received the same amount of castrated male goat serum (control serum) on Day 5 of the estrous cycle (Day 0 = estrus). All animals in each group experienced a wave of follicular development after ovulation, and the dominant follicle was over 8.5 mm in diameter on Day 5. The corpus luteum was identified for each group on Day 5. Plasma concentrations of estradiol and progesterone gradually increased after ovulation and reached around 3.7 pg/ml and 3.0 ng/ml on Day 5, respectively, suggesting that the dominant follicle and corpus luteum were functional. Administration of inhibin antiserum produced a clear increase (P < 0.001) in plasma FSH within 8 hr compared with that in control animals. Plasma levels of luteizing hormone showed a moderate increase during 40 hr after the injection of antiserum (P = 0.08). A large number of antral follicles (4 mm in diameter) developed after the hypersecretion of FSH, coupled with the rise in plasma estradiol levels. These results clearly demonstrated that inhibin neutralization during the early luteal phase produces hypersecretion of FSH with a coincident stimulation of follicular development, indicating that inhibin is an important factor for the negative regulation of FSH secretion during the early luteal phase when secretion of estradiol and progesterone are normally high.
OBJECTIVE: Pulmonary hypertension and transient graft dysfunction may complicate the postoperative course of patients undergoing lung transplantation. Recent studies have suggested that ischemia followed by reperfusion impairs release of endothelium-derived nitric oxide (NO). We investigated the acute effect of inhaled NO on hemodynamics and the oxygen free radical scavenger system after reperfusion following lung ischemia. METHODS: Fourteen anesthetized mongrel dogs were ventilated mechanically. After median sternotomy the left lung was rendered ischemic by totally clamping the left pulmonary hilum. After 90 min, the left lung was reperfused for 150 min by unclamping of the left hilum and clamping the right pulmonary hilum so that all pulmonary blood flow was directed to the left lung. Seven dogs inhaled 30 parts per million (ppm) NO during reperfusion (NO group); the other seven dogs were ventilated without NO inhalation (control group). Hemodynamics, gas exchange, superoxide dismutase activity and lipid peroxide of pulmonary venous blood, and wet/dry ratio of reperfused lung were measured. RESULTS: Neither of the two groups showed any change in systemic blood pressure prior to or following reperfusion. Immediately after reperfusion, mean pulmonary arterial pressure was significantly less (23.2 +/- 4.2 mmHg) in the NO group than in the control group (32.7 +/- 5.8 mmHg), as had been throughout reperfusion. Pulmonary vascular resistance and right ventricular end diastolic pressure were lower after reperfusion in the NO group. Superoxide dismutase activity after reperfusion in the control group significantly decreased to 41% of preischemic value. In the NO group, however, no decrease was seen and a significantly higher value was observed. The wet/dry ratio of reperfused lung was 5.47 +/- 0.52 in the control group and 4.72 +/- 0.36 in the NO group, with decreased pulmonary moisture noted in the NO group. There was no difference in lipid peroxide between the two groups. CONCLUSION: NO inhalation suppressed pulmonary hypertension after reperfusion following lung ischemia without affecting systemic arterial pressure. As superoxide dismutase activity was not depressed in the NO group, NO inhalation might enhance suppression of oxygen free radicals. These findings suggest that NO inhalation may be therapeutically useful after lung transplantation.
To investigate the physiological importance of oestradiol-17beta and inhibin in the regulation of gonadotrophin secretion in the cyclic golden hamster, females were passively immunized against two hormones. When 200 microL antiserum against oestradiol-17beta (oestradiol-AS) was given on Day 3 (Day 1 = day of ovulation), the preovulatory gonadotrophin surge was completely blocked for 24 h and the length of the oestrous cycle was also prolonged for one day. In the group given 200 microL oestradiol-AS on Day 3, basal levels of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) increased slightly and superovulation (19.6+/-0.8, mean+/-s.e.m.) occurred. When 200 microL antiserum against inhibin (inhibin-AS) was given at 1100 hours on Day 3, a dramatic increase in plasma FSH and a slight increase in LH were noted, resulting in superovulation (38.2+/-2.6) on the expected Day 1. The present study indicates clearly that inhibin plays a major role in regulating the specific ovulation rate in the hamster through the control of FSH secretion. Present results also indicate that oestradiol-17beta suppresses basal LH secretion. Oestradiol-17beta may act as an indicator of the follicular maturation, and the high plasma concentration of oestradiol-17beta noted from Day 3 to Day 4 may play an important role in determining the timing of initiation of the preovulatory gonadotrophin surge.
The time course for loss of ability of Graafian follicles to secrete inhibin and estradiol was investigated during induced follicular atresia. Cyclic hamsters were hypophysectomized on Day 1 (estrus) and injected s.c. with 30 IU eCG. Thereafter, these animals were given a single i.p. injection of antiserum to eCG on the morning of Day 4 to induce follicular atresia in a rapid and predictable manner. A drastic fall in plasma levels of estradiol and testosterone was noted within 1 h, whereas relatively high levels of plasma inhibin were maintained until 12 h, followed by an abrupt decline by 24 h. The first histological signs of pyknosis in granulosa cells appeared by 4 h, and breakdown of the mural granulosa layer was observed in most follicles by 8-12 h after immunoneutralization of circulating eCG. According to immunohistochemical analysis, inhibin activity was unchanged in granulosa cells at 12 h followed by a slight decline by 24 h, whereas positive reaction for aromatase in these cells rapidly declined by 8 h. Immunoreactivity of 17alpha-hydroxylase/C17,20-lyase (CYP 17) was also reduced in theca cells by 8 h. These results indicate that granulosa cells continue to secrete inhibin during the process of follicular atresia, although these cells quickly shut off the secretion of estradiol, and theca cells shut off the secretion of testosterone. The present results indicate as well that a rapid decline of estradiol and testosterone in plasma is an early sign of atresia in antral follicles. These results, therefore, suggest that the loss of enzymatic activity of aromatase in granulosa cells and CYP 17 in theca cells is a part of the process of follicular atresia.
To examine the contributions of endogenous inhibin and estradiol to the regulation of FSH and LH secretion in the pregnant rat, some rats were passively immunized against inhibin and/or estradiol, and others were ovariectomized, on Days 5, 10, 15, and 20 of pregnancy. Ovarian and uterine venous blood was collected separately to confirm the sources of inhibin and steroid hormones during pregnancy. Immunoreactivity of inhibin in the placenta was also examined by RIA. Levels of inhibin in ovarian venous plasma were significantly higher than those in peripheral plasma during pregnancy. No difference was observed between the levels of inhibin in uterine venous plasma and peripheral plasma. No immunoreactivity of inhibin was detected in placental homogenate from rats at Days 10, 15, and 20. FSH secretion significantly increased after immunoneutralization of inhibin during pregnancy. A marked increase in FSH secretion was noted on Days 5 and 20, and the smallest increase was observed on Day 15. Administration of estradiol antiserum (AS) alone did not induce a significant increase in FSH secretion on any day of pregnancy. However, a synergistic effect of estradiol AS and inhibin AS was observed on Day 20. On Days 5, 10, and 20, administration of inhibin AS or estradiol AS induced a significant increase in LH secretion. A synergistic effect of inhibin AS and estradiol AS on LH secretion was observed on Day 5. On Days 5 and 10, significantly high LH secretion was noted in ovariectomized rats as compared with that in rats treated with both inhibin AS and estradiol AS, indicating that other ovarian hormones such as progesterone may be involved in the suppression of LH secretion in these stages of pregnancy. These data indicate that both inhibin and estradiol, predominantly secreted from the ovary, are involved in the regulation of gonadotropin secretion during pregnancy as during the estrous cycle in the rat.
The environmental pollutant 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD; dioxin) is a potent disrupter of vertebrate endocrine systems. It was shown previously that in utero and lactational (IUL) exposure to TCDD resulted in a reduction in serum estradiol concentrations; however, the mechanism for this remains unknown. In the current study, the effects of perinatal exposure to TCDD on the pituitary-ovarian axis were examined. Pregnant rats were given a single oral dose of 1 microg TCDD/kg or vehicle as control on gestation Day 15, and female pups were killed on postnatal Day 21. Pituitaries were assayed for gonadotropin beta-subunit mRNA; additional pituitaries were cultured for 4 h and the media were assayed for FSH. Gonadotropin receptor mRNAs from vehicle- and TCDD-exposed animals were compared, with some ovaries cultured and the media assayed for estrogen secretion. LH, FSH, progesterone, and androstenedione concentrations were determined in serum. IUL exposure to TCDD resulted in a significant reduction of pituitary FSHbeta mRNA. Although estrogen output was shown to be reduced, neither serum FSH nor LH concentration was increased significantly, and FSH secretion in vitro was not altered. Similarly, serum progesterone and androstenedione were not altered by TCDD exposure, while in vitro estrogen secretion was significantly reduced. These data suggest that TCDD did not act on serum gonadotropin concentrations. The reduction in the concentration of serum estrogen appears to result from direct or indirect actions on the ovary at some point following androstenedione production.