Association between dexamethasone treatment and alterations in serum concentrations of trace metals.

Previously, a significant elevation in the serum levels of iron (Fe) was observed within a few days after the initiation of cisplatin (CDDP)-based chemotherapy. To clarify the underlying mechanisms, the serum concentration of hepcidin, a negative regulator of Fe release, was determined in the clinical samples obtained from six patients with cancer. The result showed that the serum concentration of hepcidin in patients receiving CDDP-based chemotherapy was significantly increased after 4-6 days of treatment, in comparison to the baseline level, suggesting that aforementioned excessive systemic Fe was not explained by the change of serum hepcidin level. All these patients received antiemetic premedication. We next evaluated of the effects of Pt-containing drugs and prophylactic antiemetic dexamethasone medication on the serum concentration of trace metals in mice, and on the hepatic and renal concentration of trace metals. The serum concentrations of Fe, Cu, and Zn in the CDDP-treated and oxaliplatin-treated mice were not significantly altered in comparison to those of the vehicle-treated control group. The serum concentrations of Fe, Cu, and Zn were increased after 24 h of dexamethasone treatment, compared to those of the control group (P < 0.05). The hepatic concentration of Mn was significantly reduced, whereas those of Fe and Cu inclined to diminish. The present findings suggest that dexamethasone can partly contribute to the changes in the serum concentrations of trace metals during anticancer chemotherapy.

Pathophysiological significance of the obese gene product, leptin, in ventromedial hypothalamus (VMH)-lesioned rats: evidence for loss of its satiety effect in VMH-lesioned rats.

To explore the pathophysiological significance of the obese (ob) gene product, leptin, in ventromedial hypothalamus (VMH)-lesioned rats, we examined the synthesis and secretion of leptin and its satiety effect in VMH-lesioned rats compared with those in sham-operated rats. Northern blot analysis revealed that ob gene expression is markedly augmented in the mesenteric and sc white adipose tissue, but remained unchanged in the epididymal white adipose tissue during the development of obesity in VMH-lesioned rats. Plasma leptin levels were relatively constant in sham-operated rats, but were elevated during the development of obesity in VMH-lesioned rats. In sham-operated rats, a single i.v. (1.0 mg/rat) or intracerebroventricular (2.0 micrograms/rat) injection of recombinant human leptin reduced food intake and body weight gain in sham-operated rats. By contrast, no significant effect on food intake or body weight gain was observed in VMH-lesioned rats. The present study provides evidence that VMH-lesioned rats overproduce leptin and increase its release but cannot respond to it and suggests that the loss of its satiety effect contributes to the development of obesity and the obesity-related phenotypes in VMH-lesioned rats.

Glucocorticoid regulation of leptin synthesis and secretion in humans: elevated plasma leptin levels in Cushing's syndrome.

Leptin, the obese (ob) gene product, is an adipocyte-derived satiety factor that is involved in the regulation of food ingestion and body weight. To investigate glucocorticoid regulation of leptin synthesis and secretion in humans, we measured plasma leptin levels in patients with Cushing's syndrome with adrenal or pituitary adenoma and in patients with iatrogenic Cushing's syndrome. Plasma leptin levels in patients with Cushing's syndrome were significantly elevated compared to those in nonobese healthy subjects and obese subjects without any metabolic or endocrine diseases at a given percentage of body fat by analysis of covariance. In patients with adrenal or pituitary adenoma, after the tumor resection, plasma leptin levels were reduced, with a concurrent decrease in plasma cortisol levels. With no significant changes in body weight, plasma leptin levels were also elevated significantly in lean healthy volunteers 24 h after the administration of 1 mg dexamethasone. Dexamethasone potently induced ob gene expression and leptin secretion in the organ culture of human adipose tissue. The data demonstrate that glucocorticoids act, at least in part, directly on the adipose tissue and increase leptin synthesis and secretion in humans.

Self-association of LIM-kinase 1 mediated by the interaction between an N-terminal LIM domain and a C-terminal kinase domain.

LIM-kinase 1 (LIMK1) and 2 (LIMK2) are members of a novel class of protein kinases containing two LIM motifs at the N-terminus. The LIM motif is thought to be involved in protein-protein interactions. We report here evidence that LIMK1 self-associates and also associates with LIMK2. In vivo and in vitro binding analyses using variously deleted mutants of LIMKI revealed that the self-association of LIMK1 was caused by interaction between the N-terminal LIM domain and the C-terminal kinase domain. The association of LIMK1 with itself and with LIMK2 is important for understanding how activities and functions of LIMK family kinases are regulated.

Factor VII and fibrinogen levels examined by age, sex, and other atherosclerotic risk factors in a Japanese population. The Jichi Medical School Cohort Study.

Factor VII coagulant activity (FVIIc) and fibrinogen (Fbg) levels have been investigated as cardiovascular risk factors. We studied these two factors with stratification for age, sex and blood pressure, and the relation with other atherosclerotic risk factors in a Japanese general population. The subjects were 3,139 Japanese (1,315 males and 1,824 females) aged 30 to 89 in 1992 and 1993. A linear increase with age was observed in the levels of Fbg in both men and women, but no differences were observed between men and women in each age group. A linear increase with age was also seen in the levels of FVIIc in women, but the levels of FVIIc in men were significantly higher for the age group 40-49 years than for any other age group. The levels of FVIIc in women were significantly higher than in men at age > or = 60 years. As concerning the effect of alcohol intake status, Fbg had a tendency to decrease with alcohol intake. Fbg and FVIIc levels were associated with an increase in smoking status in men, but no association was seen in women in either Fbg or FVIIc. FVIIc was positively correlated with age, body mass index, total cholesterol, triglycerides and fasting insulin level. Fbg was positively correlated with age, systolic blood pressure, diastolic blood pressure, total cholesterol, LDL-cholesterol and triglycerides in women, but Fbg had few positive correlations with risk factors in men. A comparison with previous Western studies showed that the Fbg levels of our Japanese population were lower than those of the Caucasians studied, but the present FVIIc levels were nearly the same level or slightly higher than theirs. The association of Fbg and FVIIc and with other cardiovascular risk factors in Japanese was similar to those observed in Caucasians.

Physical-strength tests and mortality among visitors to health-promotion centers in Japan.

A follow-up study was conducted to clarify the relationship between physical-strength level and risk of death from all causes and from cancer and cardiovascular disease. The 7286 persons who were examined at seven health-promotion centers throughout Japan between 1982 and 1987 were followed up. By January 1992, 6259 persons (85.9%) had been contacted by questionnaire. They included 3117 men (49.8% of all subjects studied) (average age 53.6 years at baseline, SD = 9.0 years, range 40-84 years), and 3142 women (50.2%) (average age 54.5 years at baseline, SD = 8.5 years, range 40-85 years). The follow-up period for each person averaged 6.1 years, for a total of 38,253 person-years. During this period, 155 deaths were reported. At baseline, five physical-strength tests (grip strength, side step, vertical jump, standing trunk flexion, and sit-ups) were performed. Five clinical laboratory tests (thickness of skinfold, blood sugar, total serum cholesterol, percent vital lung capacity, and blood pressure) were also conducted. The examinees were questioned about smoking status (current smoker, nonsmoker, and ex-smoker). Men with thicker skinfold [relative risk (RR) = 2.11] and higher levels of blood sugar (RR = 1.89) had an excess risk of death from all causes. Men with higher serum cholesterol (RR = 5.08), thicker skinfold (RR = 4.54), and elevated blood pressure (RR = 2.33) had an excess risk of death from cardiovascular disease. In women, no relationship was seen between clinical laboratory tests and an excess risk of death. Men exhibiting lower values for side step (RR = 2.43), vertical jump (RR = 2.37), sit-ups (RR = 1.93) and grip strength (RR = 1.92) also had an excess risk of death from all causes. Furthermore, men with lower heights for vertical jump (RR = 5.51) had an excess risk of death from cardiovascular disease. After adjustment for skinfold thickness, blood sugar, total serum cholesterol, blood pressure, percent vital lung capacity and smoking status, men with a lower level of side step, vertical jump, and grip strength had an excess risk of death from all causes. No such relationship was seen between physical-strength level and an excess risk of death in women. It is concluded that a low level of physical strength might be significantly correlated with subsequent health outcomes in men.

Binding sites for gonadotropins in human postmenopausal ovaries.

The binding of human LH and human FSH to postmenopausal ovarian tissue from 21 patients with cervical carcinoma was analyzed. The binding sites for FSH and LH were demonstrated in postmenopausal ovarian tissue. The surface-binding sites for gonadotropins were localized in the cells of cortical stroma of the postmenopausal ovary. In addition, diffuse cytoplasmic staining of endogenous estrogen and 3 beta-hydroxysteroid dehydrogenase activity were detected immunohistochemically and histochemically in the cells of the cortical stroma. Electron microscopic study also suggested steroidogenic function in the cells of the cortical stroma. The results of the present study suggest that postmenopausal ovaries contain specific binding sites for pituitary gonadotropins and play a role in ovarian steroidogenesis.

Transport of Na+ and HCO3- out of red blood cells is simultaneous with a chloride shift in canine and human whole blood exposed to CO2-rich gas.

To study the release of Na+ from erythrocytes, arterial whole blood obtained from anesthetized and heparinized dogs (n = 8), or human (n = 9) venous blood was aerated with a CO2-rich gas mixture (15% CO2, 25% O2, 60% N2) at a rate of 1 l/min. To examine possible mechanisms involved in the release of sodium ions into plasma, CO2-rich gas was also passed through whole blood in the presence of acetazolamide (n = 8), SITS (n = 8), or furosemide (n = 8). The changes in blood gas parameters, plasma protein concentration, hematocrit, and plasma sodium, chloride, and bicarbonate ion concentrations were examined. When CO2-rich gas was passed through treated and untreated canine or human whole blood, PCO2 and plasma sodium and bicarbonate contents per ml of blood increased, while pH and plasma chloride content per ml of whole blood decreased. After 0 to 5 min of CO2-rich gassing, the mean rates of change in plasma ion contents per ml of whole blood per min were related by the following equations: (1) delta MCI- = a (delta MNa+), (2) delta MHCO3- = b(delta MNa+), (3) delta MHCO3- approximately equal to delta MNa+ + delta MCl-, (4) b approximately equal to 1 + a. These results suggest that the release of Na+ and HCO3- from erythrocytes into plasma in canine and human whole blood occurred in a one-to-one ratio simultaneously with, and probably independently of, the well-known chloride shift during a period of 0 to 5 min after the start of CO2-rich gassing.

Determinants of hyperleptinaemia in an African population.

OBJECTIVE: To examine the determinants for elevated plasma leptin concentration in normal weight (NW), obese (OB), and morbidly obese (MO) individuals in Tanzania. DESIGN: Cross-sectional epidemiological study, the CARDIAC study. SETTING: Three areas in Tanzania; Dar es Salaam, urban (U), Handeni, rural (R) and Monduli, pastoralists (P), in August 1998. SUBJECTS: Five hundred and forty five participants from a random sample of 600 people aged 46-58 years. MAIN OUTCOME MEASURES: Plasma leptin concentrations, height, weight, body mass index (BMI), lipid profiles, haemoglobin A1c (HBA1c), and blood pressure (BP). RESULTS: Plasma leptin concentrations were higher in women than in men (women; 16.0 ng/mL, men; 3.1 ng/mL; p<0.0001). Women showed a higher mean body mass index (BMI), total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C) and high-density lipoprotein cholesterol (HDL-C) than men. In both genders, plasma leptin concentration, total cholesterol (TC), low density lipoprotein cholesterol (LDL-C), triglycerides (TG), systolic BP (SBP) and diastolic BP (DBP) were significantly higher in OB than in NW participants. MO women had significantly higher leptin concentration, SBP and DBP compared with the other two groups. In NW men, log leptin concentrations showed a direct correlation with weight, BMI, HBA1c, TC, LDL-C, TG, SBP and DBP (all p<0.0001 except TG; p<0.001), while among NW women and OB men, weight and BMI correlated positively with log leptin (all p<0.05). OB women observed a positive correlation between log leptin and weight, BMI and LDL-C. Regression analysis indicated that among NW subjects, gender, BMI and TC explained 53.9% of the variation in log leptin. In OB subjects, gender, BMI and LDL-C explained 51.7% of the variability in leptin levels. No relationship was found between log leptin and CVD risk factors among MO subjects. CONCLUSION: The most important determinants for hyperleptinaemia in NW participants were gender, BMI, TC, while in addition to these LDL-C, was an important determinant of leptin concentration in OB individuals. In MO women, the high leptin concentrations did not reflect the amount of adipose stores.

[Effect of L-ascorbic acid 2-phosphate on cultured rabbit keratocytes (the second report)].

We examined the effect of L-ascorbic acid 2-phosphate (P-Asc) on the proliferation of cultured rabbit keratocytes. P-Asc is a derivative of L-ascorbic acid and it yields more prolonged effects of vitamin C in solution than L-ascorbic acid. The proliferation of cultured rabbit keratocytes was promoted by the presence of P-Asc in culture medium for 10, 20, and 30 days. Transmission electron microscopic observations revealed that the cells were more multilayered in the presence of P-Asc (0.1mM) for thirty days than those in the absence of P-Asc. Moreover, this effect of P-Asc was attenuated by azetidine 2-carboxylic acid which is an inhibitor of collagen synthesis. Hence, it is suggested that the promotive effect of P-Asc on the growth of cultured keratocytes is related to the synthesis of collagen. Based on our observations, P-Asc may have a therapeutic effect on corneal stromal damages such as a corneal chemical burn and surgical trauma.

[Experimental study of phagocytosis of the cells observed on the surface of intraocular lens].

It has been reported that macrophages adhere to implanted intraocular lens (IOL) in a reaction to the foreign body, and transform themselves into epithelioid cells, fibroblast-like cells, and foreign body giant cells. We attempted to examine the activity of these cells by studying phagocytosis. Extracapsular lens extractions with implantation of posterior chamber IOLs were performed in rabbit eyes. After one week, the implants were removed and incubated in culture medium containing latex beads to investigate phagocytosis of these cells adhering to the implants by light microscopy, scanning and transmission electron microscopy. The cells observed were macrophages, epithelioid cells, fibroblast-like cells, and foreign body giant cells. The round macrophage was most active in phagocytizing latex beads, while the activity was weaker in the epithelioid cells, fibroblast-like cells, and foreign body giant cells. The macrophages on the implants had many ruffles on their surface, and many latex beads adhered to the cell surface. The epithelioid cells and foreign body giant cells had few ruffles with few latex beads on their surface. These results suggested that cellular activity of the macrophages on the implanted IOL might decrease as their shape transformed.

[Contribution of calmodulin to renin release in spontaneously hypertensive rats].

A comparative study was designed to evaluate the contribution of calmodulin to renin release from isolated glomeruli of spontaneously hypertensive rats (SHR, Okamoto and Aoki). Male 7-week-old SHR and age-matched control Wistar-Kyoto rat (WKY) were used in the present study. Glomeruli were isolated by the method of Beierwaltes et al. Isolated glomeruli were placed within the superfusion chamber and perfused with Krebs-Ringer solution at a constant flow of 0.3 ml/minute at 37 degrees C. Renin release was increased by calmodulin inhibitor, W-7 in both SHR and WKY. SHR showed higher maximal levels of renin release by W-7 compared to WKY. This result indicates that calmodulin plays an inhibitory role in renin release from juxtaglomerular cells. Calmodulin-mediated suppression mechanism in renin release is augmented in SHR.

[Participation of apoptosis in renal amyloidosis].

Renal amyloidosis shows symptoms of renal dysfunction due to the deposition of amyloid protein in the kidney. Recently, it was reported that apoptosis plays an important role in the pathogenesis of type-2 diabetes mellitus and Alzheimer's disease of which amyloid deposition is seen in the tissue. We investigated whether or not apoptosis and related factors are observed in renal amyloidosis. In situ nick end labeling (TUNEL) was performed in seven autopsied renal tissues with primary and secondary amyloidosis and 10 autopsied renal tissues without renal disease as the control. The number of TUNEL-positive cells was significantly increased in both the glomeruli and tubulus of the kidney with amyloidosis than in the control. Electron microscopic analysis was performed on one biopsied renal tissue with amyloidosis and six biopsied renal tissues with minor abnormalities as the control. Typical apoptotic cells were observed only in the former. Bax product, an inducer of apoptosis, and Bcl-2 protein, an inhibitor of apoptosis, were examined immunohistochemically in the seven autopsied renal tissues with amyloidosis and 10 autopsied control tissues. Bax was overexpressed in the tubulus and glomeruli of subjects with renal amyloidosis, compared to the normal controls. However, Bcl-2 protein was not detected in the glomeruli in any of the subjects examined. These results indicate that apoptotic cells are increased in number in renal amyloidosis and Bax overexpression may play an important role in this increase.

Fate of sperm tail components after incorporation into the hamster egg.

The changes in the cytoplasmic organelles of sperm tail in golden hamster eggs fertilized in vivo were observed by electron microscopy. Eggs were obtained from oviducts of hamsters that had been superovulated and inseminated by injection of cauda epididymal spermatozoa into the uteri. In the egg cytoplasm 10 hours after insemination, some of the mitochondria of the spermatozoon midpiece had begun to swell, and a number of multivesicular bodies were observed surrounding the midpiece. The fibrous sheath of the principal piece quickly disappeared prior to the first cleavage, whereas the axoneme and outer dense fibers were unaltered. During the two-cell stage, numerous multivesicular bodies gathered around the midpiece and fused with the mitochondria. The heterophagic vacuoles thus formed then gradually separated from the axial fibers. The outer dense fibers were disarranged and partially torn into small segments; then they seemed to dissociate into substructural granular components. The axonemal microtubules had begun to swell but were still present in the two blastomeres. It is indicated from these observations that at least the mitochondria of the tail constituents carried into the oocyte are digested into small molecular elements by the multivesicular bodies and are possibly distributed as nutrients for the blastomeres during the early stage of development.

A comparative epidemiological study on the effects of physical fitness on health level.

Clinical tests, a questionnaire on life style and mental health, and physical fitness tests were conducted on 3,132 individuals (1,796 men and 1,336 women) to find the relationships between physical fitness level and life style, mental health, or cardiovascular risk factors among Japanese. Individuals with an advanced physical fitness level also exhibited optimum mental health: the odds ratio (OR) and 95% confidence interval (CI) for their sensation of fatigue (fast recovery) was 1.16 (95% CI = 1.00 to 1.35) and that for mood recovery (fast) was 1.29 (95% CI = 1.12 to 1.49). Among those with an advanced physical fitness level, the following were rare (OR): daily smoking [0.78]; obesity (over 120%) [0.73 (95% CI = 0.62 to 0.87)]; thick subcutaneous fat (over 20 mm on the back) [0.77 (95% CI = 0.66 to 0.90)]; high blood sugar level [0.64 (95% CI = 0.50 to 0.82)]; and hypertension [0.67 (95% CI = 0.49 to 0.91)]. These are risks for circulatory diseases. These data show that risk factors associated with cardiovascular disease are more common in those with a low physical fitness level.

A comparative epidemiological study of the effects of regular exercise on health level.

STUDY OBJECT: To reveal the relationship between regular exercise and physical fitness, life style, mental health, and other cardiovascular risk factors among the Japanese. DESIGN: cross-sectional study. SUBJECTS: Three thousands, one hundred and thirty-two individuals (1,795 men and 1,336 women) over 40 years old who were examinees at one of seven centers for health promotion. METHODS: A questionnaire survey on the life style, a physical fitness tests and clinical laboratory tests were performed. RESULTS: Continuing regular exercise was associated with high levels of physical strength and their odds ratio (OR) was 2.09 (95% confidence interval(CI) 1.74-2.50). Many individuals in this group had good mental health: the OR for their sensation of fatigue (swift recovery) was 1.40(95% CI; 1.17-1.68) and that for mood recovery (fast) was 1.44(95% CI; 1.20-1.72). Thinness and obesity were scarce in this group. CONCLUSIONS: High physical fitness level, good mental health, standard body weight are more common in regular exercise group.

L-ascorbic acid 2-phosphate, a phosphate derivative of L-ascorbic acid, enhances the growth of cultured rabbit keratocytes.

We examined the effect of L-ascorbic acid 2-phosphate (P-Asc) on the proliferation of cultured rabbit keratocytes. P-Asc is a phosphate derivative of L-ascorbic acid and has more prolonged vitamin C activity in solution than does L-ascorbic acid. The proliferation of cultured keratocytes was promoted by the presence of P-Asc in culture medium. Transmission electron microscopic observations revealed that cells were more multi-layered after culture in the presence of P-Asc (0.1 mM) for 30 days than were those cultured in the absence of P-Asc. The effect of P-Asc was abrogated by L-azetidine 2-carboxylic acid, which is an analogue of proline that inhibits the production and secretion of collagen. Our observations support a therapeutic role for P-Asc in the repair of corneal stromal damage such as that caused by a corneal chemical burn.

Oral passive immunization against dental caries in rats by use of hen egg yolk antibodies specific for cell-associated glucosyltransferase of Streptococcus mutans.

The effect of polyclonal egg yolk immunoglobulin G antibodies (yIgG) raised against whole cells, cell-free (CF) glucosyltransferase (GTase), or cell-associated (CA) GTase of serotype c Streptococcus mutans was examined in terms of in vitro inhibition of virulence-related factors of S. mutans and protection of S. mutans-infected rats against the development of dental caries. Hens (18 weeks old) were immunized with formalin-treated whole cells, purified CF-GTase, or CA-GTase together with Freund's complete adjuvant. In addition, yIgG to surface protein antigen was used in some in vitro experiments for comparison with other antibodies. yIgG was purified by ammonium sulfate precipitation followed by DEAE-Sephacel column chromatography or fractional precipitation with ethanol. Purified yIgG was found to be a 220-kDa protein, which was dissociated into heavy and light chains upon addition of 2-mercaptoethanol. yIgG to whole cells and surface protein antigen gave a heavy aggregation of S. mutans organisms, while yIgG to CF- and CA-GTase specifically inhibited the enzymatic activity of the respective GTase. yIgG to CA-GTase and whole cells was found to clearly suppress the adherence of S. mutans cells to a glass surface. Specific-pathogen-free Sprague-Dawley rats that had been infected heavily and repeatedly with S. mutans and fed diet no. 2000 developed severe dental caries, while rats fed diet 2000 containing greater than or equal to 0.1% yIgG to CA-GTase showed a statistically significant reduction in dental plaque accumulation and caries development. Administration of yIgG to CF-GTase and whole cells failed to protect against caries. These results clearly suggest that yIgG to S. mutants CA-GTase specifically inhibited a virulence factor of this organism, i.e., insoluble glucan-synthesizing CA-GTase, resulting in a significant reduction in the development of dental caries.