Lysophosphatidic Acid Synthesis and its Receptors' Expression in the Bovine Oviduct During the Oestrous Cycle.

Lysophosphatidic acid (LPA) is a naturally occurring simple phospholipid which in the bovine reproductive system can be produced in the endometrium, corpus luteum, ovarian follicle and embryo. In this study, we examined the possibility that LPA receptors are expressed, and LPA synthesized, in the bovine oviduct. We found that the concentration of LPA was highest in infundibulum in the follicular phase of the oestrous cycle and was relatively high during the early-luteal phase in all examined parts of the oviduct. We also documented that LPA synthesis engages both available pathways for LPA production. The autotaxin (ATX) protein expression was significantly higher in the infundibulum compared to the isthmus during the follicular phase of the oestrous cycle. During the early-luteal phase of the oestrous cycle, ATX and phospholipase A2 (PLA2) protein expression was highest in ampulla, although the expression of LPARs was not as dynamic as LPA concentration in the oviduct tissue, and we presume that in the bovine oviduct, the most abundantly expressed receptor is LPAR2. In conclusion, our results indicate that the bovine oviduct is a site of LPA synthesis and a target for LPA action in the bovine reproductive tract. We documented that LPAR2 is the most abundantly expressed in the bovine oviduct. We hypothesize that in the bovine oviduct, LPA may be involved in the transport of gametes, fertilization and cellular signalling between the oviduct and cumulus-oocyte complex.

Human chorionic gonadotropin-like substance in nonendocrine tissues of normal subjects.

By means of two assay systems, a beta chain human chorionic gonadotropin radioimmunoassay and a radioreceptor gonadotropin assay, a chorionic gonadotropin-like substance was demonstrated in extracts of liver and colon obtained at autopsy from three patients who died of nonneoplastic disease. In contrast to placental chorionic gonadotropin, colon and liver chorionic gonadotropin was not bound to concanavalin A-Sepharose columns, indicating that this substance possessed little or no carbohydrate. Previous workers demonstrated that desialylated human chorionic gonadotropin possesses little or no bioactivity in vivo but retains full radioreceptor and radioimmunoassay activity in vitro. Our data suggest that the genome responsible for the human chorionic gonadotropin production is not completely suppressed in adult nonendocrine tissues, and that the chorionic gonadotropin produced by colon and liver has little or no bioactivity in vivo because of its low carbohydrate content. Since many normal tissues produce chorionic gonadotropin, bioactivity may be modulated by regulation of carbohydrate content.

RNA-seq analysis of diet-driven obesity and anti-obesity effects of quercetin glucoside or epigallocatechin gallate in Drosophila adults.

OBJECTIVE: High-fat diet (HFD) feeding stimulates fat accumulation in mammals and Drosophila. In the present study, we examined whether simultaneous feeding of familiar anti-obesity drugs, quercetin glycosides (QG) and epigallocatechin gallate (EGCG), to Drosophila has the same suppressive effect on fat accumulation as previously reported in rats and mice. To understand the underlying molecular mechanisms of HFD diet-induced obesity and the suppression effect of the drugs, we performed transcriptome analyses. MATERIALS AND METHODS: We induced extra fat accumulation by feeding Drosophila fly food containing 20% coconut oil and quantified the triglyceride accumulated in flies. The effects of anti-obesity drugs were also evaluated. We isolated total RNA from each sample and performed RNA-seq analyses and quantitive Real Time-Polymerase Chain Reaction (qRT-PCR) to investigate altered gene expression. RESULTS: The mRNA levels of several genes involved in lipid metabolism, glycolysis/gluconeogenesis, and anti-oxidative stress changed in HFD-fed adults. Moreover, the levels altered in those fed an HFD with QG or EGCG. The qRT-PCR further confirmed the RNA-seq data, suggesting that the expression of five essential genes for lipid metabolism changed in HFD-fed flies and altered in the flies treated with anti-obesity drugs. The most remarkable alteration was observed in the dHSL gene encoding a lipase involved in lipid-storage after HFD feeding and HFD with QG or EGCG. These alterations are consistent with HFD-induced fat accumulation as well as the anti-obesity effects of the drugs in mammals, suggesting that the genes play an important role in anti-obesity effects. CONCLUSIONS: These are the first reports to date of entire profiles of altered gene expression under the conditions of diet-induced obesity and its suppression by anti-obesity drugs in Drosophila.

Assessment of hemorrhage in pituitary macroadenoma by T2*-weighted gradient-echo MR imaging.

BACKGROUND AND PURPOSE: Intratumoral hemorrhage occurs frequently in pituitary macroadenoma and manifests as pituitary apoplexy and recent or old silent hemorrhage. T2*-weighted gradient-echo (GE) MR imaging is the most sensitive sequence for the detection of acute and old intracranial hemorrhage. T2*-weighted GE MR imaging was used to investigate intratumoral hemorrhage in pituitary macroadenomas. MATERIALS AND METHODS: Twenty-five consecutive patients who underwent total or subtotal resection of pituitary macroadenoma with heights from 17 to 53 mm, including 1 patient with classic pituitary apoplexy, underwent MR imaging before surgery, including T2*-weighted GE MR imaging. For histologic assessment of the hemorrhage in whole surgical specimens, we used hematoxylin-eosin staining. RESULTS: T2*-weighted GE MR imaging detected various types of dark lesions, such as "rim," "mass," "spot," and "diffuse" and combinations, indicating clinical and subclinical intratumoral hemorrhage in 12 of the 25 patients. The presence of intratumoral dark lesions on T2*-weighted GE MR imaging correlated significantly with the hemorrhagic findings on T1- and T2-weighted MR imaging (P < .02 and <.01, respectively), and the surgical and histologic hemorrhagic findings (P < .001 and <.001, respectively). CONCLUSION: T2*-weighted GE MR imaging could detect intratumoral hemorrhage in pituitary adenomas as various dark appearances. Therefore, this technique might be useful for the assessment of recent and old intratumoral hemorrhagic events in patients with pituitary macroadenomas.

Relationship of cigarette smoking and radiation exposure to cancer mortality in Hiroshima and Nagasaki.

Cancer mortality among 40,498 Hiroshima and Nagasaki residents was examined in relation to cigarette smoking habits and estimated atomic bomb radiation exposure level. Relative risk (RR) models that are either multiplicative or additive in the two exposures were emphasized. Most analyses were directed toward all nonhematologic (ANH) cancer, stomach cancer, lung cancer, or digestive tract cancer other than stomach cancer, for which there were, respectively, 1,725, 658, 281, and 338 deaths in the follow-up period for this study. Persons heavily exposed to both cigarette smoke and radiation were found to have significantly lower cancer mortality than multiplicative RR models would suggest for ANH cancer, stomach cancer, and digestive tract cancer other than stomach cancer. Surprisingly, the RR function appeared not only to be submultiplicative for some of these cancer site categories but also may be subadditive. The lung cancer RR function could not be distinguished from either a multiplicative or an additive form. The number of deaths was sufficient to permit some more detailed study of ANH cancer mortality: RR functions appeared to be consistent between males and females, though a paucity of heavy smoking females limits the precision of this comparison. The submultiplicative nature of the RR function mentioned above was particularly pronounced among persons who were relatively young (less than or equal to 30 yr of age) at the time of radiation exposure. The RR function for these younger subjects depends strongly on both radiation and cigarette smoke exposure levels. Even light smoking (approximately 5 cigarettes/day) for an extended period of time was associated with a large estimated RR. Implications of these findings are discussed in relation to human carcinogenesis models. As a byproduct, cancer mortality of several sites is significantly related to radiation exposure in this population, after accommodation for the possible confounding effects of cigarette smoking.

Correlates of cortical bone mass among premenopausal and postmenopausal Japanese women.

Few studies have examined the multifactorial risk factors of bone mass in Asian populations. This cross-sectional study was designed to explore relationships between bone mass and environmental variables, including dietary and life-style factors, in Japanese women living in Japan. A total of 178 Japanese women completed the study: 89 premenopausal, ages 35-40, and 89 postmenopausal, ages 55-60. Midradial bone mineral content (MBMC) and bone mineral content per unit area, referred to as bone density (MBMD), were measured using single-photon absorptiometry. The major results of this investigation were the following: (1) The postmenopausal women differed significantly from the premenopausal women in having lower radial bone parameters, lower mean height, later age of menarche, and higher dietary intakes of carbohydrates, vegetables, and milk with a lower intake of caffeine. (2) Current protein intake was a positive correlate of MBMC in both groups. (3) Intake of vegetables (leafy green, yellow, orange, and white) and current milk intake were positively associated with MBMC in the postmenopausal women. (4) For the premenopausal women, irregular menstrual cycles was a negative correlate of MBMC, and for the postmenopausal women, years of menopause was negatively associated with MBMC and MBMD. Longitudinal studies are needed to establish more conclusively associations among diet, life-style, and reproductive history and bone mass of Japanese women.

Calcium transport in perfused duodena from normal chicks: enhancement within fourteen minutes of exposure to 1,25-dihydroxyvitamin D3.

The effect of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3) on calcium transport was studied in vascularly perfused duodena of normal, vitamin D-replete chicks. Addition of 130 pM 1,25(OH)2D3 to the perfusate resulted in a significant increase in 45Ca transport from the lumen to the vascular effluent within 14 min; the transport rate rose to 140% of levels in comparable preparations exposed for 40 min to vehicle. No effects of 1,25(OH)2D3 were noted on the back flux or transfer of 45Ca from the vascular effluent to the lumen. Vascular perfusion with 100 microM colchicine, an antimicrotubular agent, abolished the rapid lumen-to-vascular effluent effect of 1,25(OH)2D3 on 45Ca transport, relative to preparations exposed to the secosteroid and 100 microM lumicolchicine, (a light inactivated analog of colchicine). Colchicine did not, however, alter basal 45Ca transport rates. Addition of 130 pM 1,25(OH)2D3 to the lumenal compartment of normal chicks or vascular perfusion of duodena from vitamin D-deficient birds failed to increase 45Ca transport above control levels. Perfusion of duodena from normal chicks with 650 pM 1,25(OH)2D3 further increased calcium transport to 170% of levels observed in preparations treated with 130 pM steroid, and 210% of levels in controls. Although 15 nM vitamin D3 had no effect, in one series of experiments 125 nM 25-hydroxyvitamin D3 elicited vascular calcium levels that were 185% of controls at 40 min. These results suggest that 1,25(OH)2D3 can act in vitamin D-replete animals to produce rapid unidirectional calcium transport responses (through unknown mechanisms), as well as by interaction with intestinal nuclear receptors in D-deficient animals to promote induction of protein(s) that support long acting calcium transport responses.

Hypercalcemia inhibits the rapid stimulatory effect on calcium transport in perfused duodena from normal chicks mediated in vitro by 1,25-dihydroxyvitamin D3.

We have previously reported that vascular perfusion of normal vitamin D3-replete chick duodena with physiological amounts of 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3] increases the movement of 45Ca2+ from the lumen to the venous effluent within 14 min under conditions of normal Ca2+ (0.9 mM) concentration in both the lumen and vascular perfusate. The present studies were designed to further explore details of this rapid 1,25-(OH)2D3 effect as a function of seco-steroid concentration and under conditions where the free Ca2+ concentrations in the perfusate were varied from 0.54-1.26 mM. Concentrations of 1,25-(OH)2D3 in the vascular perfusate ranging from 30-650 pM elicited an increasing stimulation of Ca transport, as judged by 45Ca levels in the venous effluent. At 0.98-6.5 nM 1,25-(OH)2D3, progressive inhibition of Ca transport was observed, yielding a biphasic dose-response curve. The optimal concentration of 650 pM 1,25-(OH)2D3 was used in subsequent experiments designed to study the effects of vascular Ca2+ levels on 45Ca transport mediated by the seco-steroid. The basal Ca2+ transport ratio, in the absence of 1,25-(OH)2D3, did not change when the divalent cation of the vascular perfusate was varied over the range 0.54-1.26 mM free calcium. However, the effect of 1,25-(OH)2D3 on 45Ca2+ transport was completely abolished in the group treated with 1.21 mM Ca2+ in the perfusate, but not in the groups treated with concentrations less than 1.17 mM Ca2+. These results suggest that the rapid intestinal calcium transport response to 1,25-(OH)2D3 may be modulated locally in part by the prevailing ionized Ca concentration of the vascular perfusate.

Additive effects of parathyroid hormone and calcitonin on adenosine 3',5'-monophosphate release in newly established perfusion system of rat femur.

A new perfusion system of isolated rat femora was established, and the effect of PTH or calcitonin (CT) on cAMP release from the adult bone was examined. Stimulation of cAMP release from the isolated perfused bone peaked rapidly between 5 and 10 min after human PTH (hPTH)-(1-34) or eel CT injection, respectively, and declined gradually toward the preinjection level. However CT exhibited a longer lived effect than PTH. Release of cAMP was still significantly greater than control at 60 min after a bolus injection of CT. The rate of cAMP release was directly related to the dose of PTH or CT. H2O2-oxidized PTH (biologically inactive) caused no increase in cAMP release. hPTH-(1-34) was markedly more potent than hPTH-(1-84) in stimulating cAMP release from the perfused bone on an equimolar basis. Simultaneous administration of 5 micrograms PTH and 5 micrograms CT at maximal stimulatory doses produced additive effects, indicating the presence of separate receptor sites for PTH and CT in the bone. In conclusion, this system provides us a means by which hormone actions on adult bone with integrity of the organ can be evaluated and therefore makes possible systemic investigation of the osteoblast-osteoclast function.

Impaired parathyroid hormone-stimulated adenosine 3',5'-monophosphate release by isolated perfused bones obtained from vitamin D-deficient rats.

The present studies were designed to explore the mechanism underlying skeletal refractoriness to PTH in a vitamin D-deficient animal by assessment of PTH-stimulated cAMP release from isolated perfused bone. In vitamin D-deficient (-D) rats both basal and PTH-stimulated cAMP release were markedly diminished, compared with that in vitamin D-replete (+D) rats. Isolated perfused bones from -D rats that had undergone parathyroidectomy 2 days before death still showed reduced cAMP release in response to PTH, compared with +D bones. To investigate which factors in terms of Ca, endogenous PTH, or vitamin D might primarily be responsible for the impaired PTH-stimulated cAMP release from -D bones, some -D rats were switched to a diet identical to the vitamin D-deficient diet but with high Ca content (4%) for 2 or 5 weeks before death. This schedule maintained normocalcemia despite vitamin D deficiency. PTH-stimulated cAMP release in these rats was increased to a level intermediate between that in -D rats and +D rats, indicating partial restoration of the impaired response to PTH in -D rats. These data indicate that skeletal refractoriness to PTH in vitamin D-deficient animals might, in part, be due to the impaired activation of adenylate cyclase, which cannot be explained entirely by hypocalcemia or associated secondary hyperparathyroidism. Vitamin D deficiency per se, therefore, may play a key role in the impaired cAMP response to PTH.

Monoamine oxidase in rat ovary during the estrous cycle. A histochemical study by a new coupled peroxidatic oxidation method.

A new coupled peroxidatic oxidation method for histochemical detection of monoamine oxidase (MAO) was applied to rat ovary. With this new method, fixed tissues could be used, and two forms of MAO could be identified by use of selective inhibitors. MAO activity was observed in the corpora lutea, interstitial gland cells, and blood vessels. In the corpora lutea, no activity was detected during the first estrous cycle, but strong activity was observed in the next two cycles. MAO in blood vessels showed characteristic changes of activity during the estrous cycle. The results suggest that MAO activity might possibly be involved in ovulation and progesterone metabolism in the ovary. Like other organs, rat ovary was found to contain two types of MAO; type A MAO was predominant in the corpora lutea. On the other hand, only one type of MAO, type B, was found in the blood vessels.

Acute systemic inflammatory response syndrome in subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: Systemic inflammatory response syndrome (SIRS) without infection is a well-known phenomenon that accompanies various acute cerebral insults. We sought to determine whether the initial SIRS score was associated with outcome in subarachnoid hemorrhage (SAH). METHODS: In 103 consecutive patients with SAH, the occurrence of SIRS was assessed according to the presence of >/=2 of the following: temperature of <36 degrees C or >38 degrees C, heart rate of >90 bpm, respiratory rate of >20 breaths/min, and white blood cell count of <4000/mm(3) or >12 000/mm(3). SIRS criteria and other prognostic parameters were evaluated as predictors of dichotomous Glasgow Outcome Scale score. RESULTS: SIRS was highly related to poor clinical grade (Hunt and Hess clinical grading scale), a large amount of SAH on CT (Fisher CT group), and high plasma glucose concentration on admission. By univariate analysis, the occurrence of SIRS was associated with higher mortality and morbidity rates than was the nonoccurrence (P<0.001). Among individual SIRS criteria, heart rate (P=0.003), respiration rate (P=0.003), and white blood cell count (P=0.03) were significant outcome predictors. By multivariate logistic regression analysis, the presence of SIRS independently predicted outcome. SIRS carried an increased risk of subsequent intracranial complications such as vasospasm and normal pressure hydrocephalus, as well as systemic complications. CONCLUSIONS: In SAH patients, SIRS on admission reflected the extent of tissue damage at onset and predicted further tissue disruption, producing clinical worsening and, ultimately, a poor outcome.

Growth hormone and prolactin release after injection of thyrotropin-releasing hormone in patients with depression.

The effects of thyrotropin-releasing hormone (TRH) on the release of growth hormone (GH), prolactin (PRL) and thyrotropin (TSH) were investigated in patients with depression. Intravenous injection of synthetic TRH (500 mug) caused a significant increase in plasma GH (peak value: 7.7 minus 35.0 ng/ml) in 8 of 13 patients with mental depression. After clinical recovery these patients had no response of plasma GH to TRH. TRH administration did not raise plasma GH in normal subjects examined. Plasma PRL responses to TRH were significantly enchanced (P smaller than 0.05) in depressed patients compared with control subjects. Plasma TSH responses to TRH were significantly blunted in patients with depression (P smaller than 0.05). These results suggest disorders in the hypothalamo-pituitary function in depression.

Blunted TSH and unaltered PRL responses to TRH following repeated administration of TRH in neurologic patients: a replication of neuroendocrine features of major depression.

A blunted thyrotropin (TSH) and an unaltered prolactin (PRL) responses to thyrotropin-releasing hormone (TRH) are widely recognized in neuroendocrinology of depression. We studied effects of repeated TRH administration of 1 mg/day for 10 days on the pituitary-thyroid axis function and PRL secretion in 16 euthyroid patients with neurological disorders. Although levels of serum thyroid hormones and of nonstimulated PRL were not affected by the treatment, baseline TSH levels were markedly inhibited. A blunted response of TSH to TRH was found without a significant effect on a PRL response to TRH after long-term treatment with TRH in four patients in whom a TRH test was performed. These changes are similar to those in depressed patients. TRH administration in this manner replicates a lowered sensitivity of thyrotrophs of the pituitary with a normal responsibility of lactotrophs in depression.

Increased sarcolemmal permeability in the cerebral artery during chronic spasm: an assessment using DNA-binding dyes and detection of apoptosis.

Alteration of sarcolemmal permeability was evaluated in the cerebral artery after subarachnoid hemorrhage. Significance of membrane dysfunction in the pathogenesis of chronic spasm and contribution of apoptosis were investigated in a canine model. Permeability of the smooth muscle cell (SMC) membrane was assessed by double staining with a hydrophilic (ethidium bromide [EB]) and a lipophilic (Hoechst 33342) DNA-binding dye. Quantitative observations were made with a ultraviolet-fluorescence microscope and a ultraviolet-laser confocal microscope. Occurrence of apoptosis was studied using electrophoresis and TUNEL method. In the normal arteries, nuclei of SMC were stained with Hoechst 33342 but not with EB. In the spastic arteries, SMC in the inner layer of the tunica media were stained with EB. The incidence of EB-positive cells reached maximum on day 7 (45 +/- 19%) and decreased in 2 to 4 weeks (13 +/- 5.2% and 5.0 +/- 2.1%, respectively), in parallel with amelioration of spasm. Electron and light microscopic observations revealed increased density of SMC cytoplasm with widening of the extracellular space. Necrosis was not evident. Apoptosis was not detected by the two methods. These results demonstrate that an augmentation in sarcolemmal permeability takes place during the course of chronic vasospasm and suggest its close correlation to pathogenesis.

Impaired calcium regulation of smooth muscle during chronic vasospasm following subarachnoid hemorrhage.

The intracellular calcium level was determined in the canine basilar artery to investigate whether Ca2+ regulation of its smooth muscle is altered during chronic vasospasm following subarachnoid hemorrhage. A double-hemorrhage model was used. The occurrence of vasospasm was confirmed angiographically 7 days after initial hemorrhage. The intracellular calcium concentration ([Ca2+]i) of smooth muscle was measured using Fura-2. Fluorescence to excitation at 340 and 356 nm was monitored and the ration R340/356 was used as the indicator of [Ca2+]i. When the extracellular calcium concentration ([Ca2+]e) was increased from pCa 8 to 2, [Ca2+]i also increased. In the spastic arteries, the [Ca2+]e - [Ca2+]i curve was elevated as compared with the normal arteries. Treatment with ionomycin elevated the curve in the normal group, but it had little effect in the spastic arteries. Values of [Ca2+]i, calculated in multiples of Kd, were greater in the spastic arteries. Diltiazem (10(-5) mol/L) partially suppressed the augmented [Ca2+]i signal in the spastic arteries, whereas it did not affect the curve in the control group. These results indicate that the calcium regulation of smooth muscle is impaired after subarachnoid hemorrhage, which may contribute to the pathogenesis of chronic vasospasm.

Somatostatin induces hyperpolarization in pancreatic islet alpha cells by activating a G protein-gated K+ channel.

Somatostatin inhibits glucagon-secretion from pancreatic alpha cells but its underlying mechanism is unknown. In mouse alpha cells, we found that somatostatin induced prominent hyperpolarization by activating a K+ channel, which was unaffected by tolbutamide but prevented by pre-treating the cells with pertussis toxin. The K+ channel was activated by intracellular GTP (with somatostatin), GTPgammaS or Gbetagamma subunits. It was thus identified as a G protein-gated K+ (K(G)) channel. RT-PCR and immunohistochemical analyses suggested the K(G) channel to be composed of Kir3.2c and Kir3.4. This study identified a novel ionic mechanism involved in somatostatin-inhibition of glucagon-secretion from pancreatic alpha cells.

Nonsteroidal antiinflammatory agents. 3. Synthesis of the positional isomers of 4'-chloro-5-methoxy-3-biphenylylacetic acid and their antiinflammatory and analgesic activities.

The positional isomers 3a-i of 4'-chloro-5-methoxy-3-biphenylylacetic acid [1 (DKA-9), R = 4-ClPh; R' = MeO] which is a newly developed nonsteroidal antiinflammatory agent, have been prepared and evaluated for antiinflammatory and analgesic activities using both the carrageenan-induced rat paw edema and AcOH writhing assays. The 3- and 4-biphenylylacetic acids 3a,d, which closely resemble 1 (R = 4-ClPh, R' = MeO) structurally, showed, by far, excellent activities compared with the other isomers in these assays. However, none of the compounds tested was more active than 1 (R = 4-ClPh; R' = MeO). In this series of compounds, structural requirements for good antiinflammatory activity seemed to be parallel to those for analgesic activity.

Nonsteroidal antiinflammatory agents. 2. Synthesis of 4',5-disubstituted 3-biphenylylacetic acids and their derivatives with antiinflammatory and analgesic activities.

A series of 5',5-disubstituted 3-biphenylylacetic acids (5a-n) and several alpha-methyl derivatives (5o-v) were prepared as analogues of a newly developed nonsteroidal antiinflammatory agent, 5'-chloro-5-methoxy-3-biphenylylacetic acid [1 (DKA-9), R = 4-C1Ph; R' = Me], and evaluated for antiinflammatory and analgesic activities using both carrageenan rat paw edema and AcOH writhing assays. Among them, 5-fluoro-3-biphenylylacetic acid (5m) showed the highest antiinflammatory activity, while 2-(3-biphenylyl)propionic acid (5o) showed the highest analgesic activity. However, they were less potent than 1 (R = 4-C1Ph; R' = Me) in these assays.