Adjuvant chemotherapy in elderly patients with pancreatic cancer.

BACKGROUND: Adjuvant chemotherapy improves survival for patients with resected pancreatic cancer. Elderly patients are under-represented in Phase III clinical trials, and as a consequence the efficacy of adjuvant therapy in older patients with pancreatic cancer is not clear. We aimed to assess the use and efficacy of adjuvant chemotherapy in older patients with pancreatic cancer. METHODS: We assessed a community cohort of 439 patients with a diagnosis of pancreatic ductal adenocarcinoma who underwent operative resection in centres associated with the Australian Pancreatic Cancer Genome Initiative. RESULTS: The median age of the cohort was 67 years. Overall only 47% of all patients received adjuvant therapy. Patients who received adjuvant chemotherapy were predominantly younger, had later stage disease, more lymph node involvement and more evidence of perineural invasion than the group that did not receive adjuvant treatment. Overall, adjuvant chemotherapy was associated with prolonged survival (median 22.1 vs 15.8 months; P<0.0001). Older patients (aged ≥70) were less likely to receive adjuvant chemotherapy (51.5% vs 29.8%; P<0.0001). Older patients had a particularly poor outcome when adjuvant therapy was not delivered (median survival=13.1 months; HR 1.89, 95% CI: 1.27-2.78, P=0.002). CONCLUSION: Patients aged ≥70 are less likely to receive adjuvant therapy although it is associated with improved outcome. Increased use of adjuvant therapy in older individuals is encouraged as they constitute a large proportion of patients with pancreatic cancer.

Testing awareness of heart disease among West Australian women.

Cardiovascular disease [CVD] is the top cause of death in Australian women. Large studies in the US and Europe have shown that the majority of women do not consider CVD as their greatest health threat. Australian women's awareness has not previously been investigated. The aim of this cross-sectional survey [TAWDAH] was to assess Australian women's awareness of CVD their leading causes of death [LCD].

The prognostic and predictive value of serum CA19.9 in pancreatic cancer.

BACKGROUND: Current staging methods for pancreatic cancer (PC) are inadequate, and biomarkers to aid clinical decision making are lacking. Despite the availability of the serum marker carbohydrate antigen 19.9 (CA19.9) for over two decades, its precise role in the management of PC is yet to be defined, and as a consequence, it is not widely used. METHODS: We assessed the relationship between perioperative serum CA19.9 levels, survival and adjuvant chemotherapeutic responsiveness in a cohort of 260 patients who underwent operative resection for PC. RESULTS: By specifically assessing the subgroup of patients with detectable CA19.9, we identified potential utility at key clinical decision points. Low postoperative CA19.9 at 3 months (median survival 25.6 vs 14.8 months, P=0.0052) and before adjuvant chemotherapy were independent prognostic factors. Patients with postoperative CA 19.9 levels>90 U/ml did not benefit from adjuvant chemotherapy (P=0.7194) compared with those with a CA19.9 of ≤90 U/ml (median 26.0 vs 16.7 months, P=0.0108). Normalization of CA19.9 within 6 months of resection was also an independent favorable prognostic factor (median 29.9 vs 14.8 months, P=0.0004) and normal perioperative CA19.9 levels identified a good prognostic group, which was associated with a 5-year survival of 42%. CONCLUSIONS: Perioperative serum CA19.9 measurements are informative in patients with detectable CA19.9 (defined by serum levels of >5 U/ml) and have potential clinical utility in predicting outcome and response to adjuvant chemotherapy. Future clinical trials should prioritize incorporation of CA19.9 measurement at key decision points to prospectively validate these findings and facilitate implementation.

Delineating the Role of ßIV-Tubulins in Pancreatic Cancer: ßIVb-Tubulin Inhibition Sensitizes Pancreatic Cancer Cells to Vinca Alkaloids.

Pancreatic cancer (PC) is a lethal disease which is characterized by chemoresistance. Components of the cell cytoskeleton are therapeutic targets in cancer. ßIV-tubulin is one such component that has two isotypes-ßIVa and ßIVb. ßIVa and ßIVb isotypes only differ in two amino acids at their C-terminus. Studies have implicated ßIVa-tubulin or ßIVb-tubulin expression with chemoresistance in prostate, breast, ovarian and lung cancer. However, no studies have examined the role of ßIV-tubulin in PC or attempted to identify isotype specific roles in regulating cancer cell growth and chemosensitivity. We aimed to determine the role of ßIVa- or ßIVb-tubulin on PC growth and chemosensitivity. PC cells (MiaPaCa-2, HPAF-II, AsPC1) were treated with siRNA (control, ßIVa-tubulin or ßIVb-tubulin). The ability of PC cells to form colonies in the presence or absence of chemotherapy was measured by clonogenic assays. Inhibition of ßIVa-tubulin in PC cells had no effect chemosensitivity. In contrast, inhibition of ßIVb-tubulin in PC cells sensitized to vinca alkaloids (Vincristine, Vinorelbine and Vinblastine), which was accompanied by increased apoptosis and enhanced cell cycle arrest. We show for the first time that ßIVb-tubulin, but not ßIVa-tubulin, plays a role in regulating vinca alkaloid chemosensitivity in PC cells. The results from this study suggest ßIVb-tubulin may be a novel therapeutic target and predictor of vinca alkaloid sensitivity for PC and warrants further investigation.

Metabolic and ultrastructural characterization of guinea pig alveolar type II cells isolated by centrifugal elutriation.

Direct biochemical studies of the whole lung have been quite misleading because of the heterogeneity of the lung cell types. One of the advantages of studying the isolated cells is to be able to correlate specific metabolic functions with intracellular molecular events and to differentiate factors that affect the type II cell function directly. In the present study we have isolated type II cells from guinea pig lung with elastase and purified them by centrifugal elutriation. These cells fluoresce with phosphine 3R as the dye is specifically taken up by the lamellar bodies. In the electron micrographs, the type II cells display punctate villi, which underwent fragmentation in those cases where metrizamide density gradient was used. Mitochondria are scattered throughout the cytoplasm, and smooth endoplasmic reticulum is sparse. Type II cells possess large irregularly shaped nuclei with peripheral areas of dense hemochromatin and at least one prominent nucleolus. Ovoid lamellar bodies are the most prominent cellular inclusions. These bodies are present throughout the cytoplasm and contain a substructure of whorling and concentric laminations. Biochemical studies indicate that type II cells prepared by centrifugal elutriation are metabolically well preserved as seen from incorporation of [14C]leucine into cellular proteins, [methyl-14C]choline into cellular disaturated phosphatidylcholine and CDP[methyl-14C]choline into mitochondrial and microsomal phosphatidylcholine. Superiority of centrifugal elutriation over the commonly employed combination of discontinuous metrizamide gradient and cell elutriation is evident from the present study.

Satiety enhancement by selective orexin-1 receptor antagonist SB-334867: influence of test context and profile comparison with CCK-8S.

Acute systemic treatment with the selective orexin-1 (OX1R) antagonist SB-334867 reduces food intake in rats, an effect associated with an acceleration in behavioural satiety and unrelated to gross behavioural disruption, alterations in palatability, or toxicity. However, as enhanced satiety is behaviourally indexed by an earlier-than-normal transition from eating to resting, and since orexin-A has been implicated in mechanisms of arousal, it remains possible that sedation contributes to the anorectic effect of acute OX1R blockade. Previous work has shown that, when treated with SB-334867 (30 mg/kg, i.p.) 30 min before a 1h test with palatable food, rats begin to show appreciable levels of resting 10-15 min earlier than under control conditions (i.e. around 20 min versus 30-35 min into the session). The present results demonstrate that a 20 min increase in the injection-test interval (i.e. 50 min) had no significant impact on the anorectic, behavioural or weight gain effects of SB-334867 in non-deprived male rats. Most importantly, this altered treatment regimen led to a temporal profile of resting virtually identical to that previously observed with the more conventional 30 min injection-test interval. Although parallel studies indicated that the OX1R antagonist accelerated the onset of resting (and suppressed most active behaviours) even in the absence of food, an equianorectic dose of the natural satiety-related signal cholescystokinin octapeptide (CCK-8S; 5 microg/kg, i.p.) also produced very similar behavioural effects regardless of the presence of food. Together with evidence that SB-334867 preserves the structural integrity of natural feeding behaviour, does not induce nausea/illness or alter taste/palatability and fails to influence EEG measures of arousal/sleep, the present findings are consistent with the view that acute OX1R antagonism selectively enhances satiety. However, unlike the immediate short-circuiting of the satiety sequence induced by CCK-8S, the slower response to SB-334867 implies a more indirect mechanism of action.

Anorexia and weight loss in male rats 24 h following single dose treatment with orexin-1 receptor antagonist SB-334867.

Acute systemic treatment with the selective orexin-1 receptor antagonist SB-334867 (30 mg/kg, i.p.) has been reported not only to inhibit food intake and to accelerate behavioural satiety in rats, but also to produce a significant loss of bodyweight over the 24 h period post-dosing. The present studies were designed to test the hypothesis that the inhibition of weight gain following acute treatment with SB-334867 is due to a persistent anorectic action of the compound. In Experiment 1, the acute effects of SB-334867 (30 mg/kg, i.p.) on food intake and behaviour in a 1 h test with palatable mash were assessed as a function of injection-test interval. Results confirmed that, when administered 30 min prior to testing, SB-334867 significantly suppressed mash intake and accelerated behavioural satiety. More importantly, significant anorexia and behavioural change were also observed when animals were tested 24 h, but not 48 h, post-dosing. As previously reported, all animals treated with the orexin-1 receptor antagonist lost bodyweight over the 24 h period following acute treatment. The generality of these findings was confirmed in Experiment 2, where acute treatment with SB-334867 (30 mg/kg, i.p.) significantly suppressed home cage chow consumption over the 24 h period post-dosing, an effect also accompanied by a significant loss of bodyweight. The results of Experiment 3 showed that, following i.p. administration of 30 mg/kg, SB-334867 has good CNS penetration, reaches peak plasma and brain concentrations at 30 min, and maintains good exposure over 4 h post-dosing. Overall, current data support the hypothesis that a persistent anorectic action contributes to the significant loss of bodyweight observed 24 h following acute dosing with SB-334867. As the compound is virtually undetectable in plasma or brain beyond 8 h post-dosing, and since nothing is known about potentially active metabolites, we consider the possibility that single dose treatment with SB-334867 results in enduring alterations to the orexin-1 receptor and/or downstream signalling pathways.

Isolation and characterization of the [Ga2Al18O8(OH)36(H2O)12](8+) cluster: cationic variations on the Wells-Dawson topology.

The structural chemistry of Group 13 polyoxometalates lags far behind related negatively charged transition metal species and limits the development of advanced materials. A novel heterometallic cluster [Ga2Al18O8(OH)36(H2O)12](8+) (Ga2Al18) has been isolated using a supramolecular approach and structurally characterized using single-crystal X-ray diffraction. Ga2Al18 represents the Wells-Dawson structure polycations and variations in the structural topology may be related to the initial stabilization of the Keggin isomer. DFT calculations on the related ε-Keggins (GaAl12 and Al13), Ga2Al18, and theoretical Al2Al18 clusters reveal similar features of electronic structure, suggesting additional heteroatom substitution in other isostructural clusters should be possible.

Estrogen production in endometriosis and use of aromatase inhibitors to treat endometriosis.

Estrogen is the most important known factor that stimulates the growth of endometriosis. Estrogen delivery to endometriotic implants was classically viewed to be only via the circulating blood in an endocrine fashion. We recently uncovered an autocrine positive feedback mechanism, which favored the continuous production of estrogen and prostaglandin (PG)E2 in the endometriotic stromal cells. The enzyme, aromatase, is aberrantly expressed in endometriotic stromal cells and catalyzes the conversion of C19 steroids to estrogens, which then stimulate cyclooxygenase-2 to increase the levels of PGE2. PGE2, in turn, is a potent inducer of aromatase activity in endometriotic stromal cells. Aromatase is not expressed in the eutopic endometrium. Aromatase expression in endometriosis and its inhibition in eutopic endometrium are controlled by the competitive binding of a stimulatory transcription factor, steroidogenic factor-1, and an inhibitory factor, chicken ovalbumin upstream promoter-transcription factor to a regulatory element in the aromatase P450 gene promoter. In addition, we find that endometriotic tissue is deficient in 17beta-hydroxysteroid dehydrogenase type 2, which is normally expressed in eutopic endometrial glandular cells and inactivates estradiol-17beta to estrone. This deficiency is another aberration that favors higher levels of estradiol-17beta in endometriotic tissues in comparison with the eutopic endometrium. The clinical relevance of local aromatase expression in endometriosis was exemplified by the successful treatment of an unusually aggressive form of recurrent endometriosis in a postmenopausal woman using an aromatase inhibitor.

Disruption of estrogen receptor gene prevents 17 beta estradiol-induced angiogenesis in transgenic mice.

Estrogen is known to modulate angiogenesis, both under physiological and pathological conditions, and has been demonstrated to augment angiogenesis induced by bFGF in a mouse model. We have modified this mouse model and measured the apparent plasma volume in Matrigel plugs containing basic fibroblast growth factor (bFGF) in wild type and estrogen receptor knockout, ovariectomized mice in the presence and absence of exogenous 17 beta estradiol. The apparent plasma volume was determined by measuring the fluorescence of the excised plug 10 min. after injection of fluoroscein labeled dextran 150. In wild type mice exogenous 17 beta estradiol increased the apparent plasma volume of the Matrigel plug and the uterine weight significantly. In the estrogen receptor knockout mice exogenous 17 beta estradiol caused a small, but significant increase in uterine weight but was without effect on the apparent plasma volume of the Matrigel plug. It is concluded that functional estrogen receptors are essential for the augmentation of bFGF-induced angiogenesis by exogenous 17 beta estradiol in female mice.

Aromatase expression in endometriosis.

The conversion of C19 steroids to estrogens occurs in a number of tissues, such as the ovary and placenta, and is catalyzed by aromatase P450 (P450arom; the product of the CYP19 gene). P450arom expression has also been detected in a number of uterine tumors, such as leiomyomas and endometrial cancer. On the other hand, P450arom expression was undetectable in normal endometrial and myometrial tissues. The present study was conducted to determine the presence or absence of aromatase expression in peritoneal endometriotic implants and in the eutopic endometrium of women with endometriosis. Endometriotic implants in pelvic peritoneum (n = 17; e.g. posterior culdesac, bladder, and anterior culdesac) and eutopic endometrial curettings (n = 11) of 14 patients with histologically documented pelvic endometriosis were obtained at the time of laparoscopy or laparotomy. Pelvic peritoneal biopsies distal to endometriotic implants as well as normal endometrial tissues (n = 7) from disease-free women were used as negative controls. We used competitive RT-PCR technology employing an internal standard to amplify P450arom transcripts in total ribonucleic acid (RNA) isolated from these tissues. P450arom transcripts were detected in all endometriotic implants and in all eutopic endometrial tissues from patients with endometriosis. P450arom messenger RNA species were not detectable in endometrial tissues from disease-free women or in endometriosis-free peritoneal tissues. The highest levels of transcripts were detected in an endometriotic implant that involved the full thickness of the anterior abdominal wall. The P450arom transcript level within the core of this endometriotic mass was 4-fold higher than that in the surrounding adipose tissue. It has been shown recently that aromatase expression in various human tissues is regulated by the use of tissue-specific promoters via alternative splicing. To analyze promoter usage, we amplified by RT-PCR the most likely promoter-specific untranslated 5'-termini of P450arom transcripts in 2 endometriotic implants. It appears that these endometriotic implants use both the adipose-type promoter I.4 and gonadal-type promoter II for aromatase expression. The use of promoter I.4 for aromatase expression in adipose tissue has been recently observed to be regulated by members of the interleukin-6 (IL-6) cytokine family. Based on these findings, we examined by RT-PCR, IL-6 and IL-11 messenger RNA expression in 5 endometriotic tissues and 1 eutopic endometrial sample from a patient with endometriosis. We detected IL-6 and IL-11 transcripts in all endometriotic tissues and in the eutopic endometrial tissue sample studied. Our findings indicate that both eutopic endometrial tissues and endometriotic implants from patients with endometriosis are biochemically different from normal endometrial tissues of disease-free women. The presence of aromatase expression in eutopic endometrial tissues from patients with endometriosis may be related to the capability of implantation of these tissues on peritoneal surfaces. Furthermore, the possibility of estrogen production in these implants may serve to promote their growth. Increased IL-6 and IL-11 expression in these tissues suggests that P450arom expression in endometriosis may be regulated in part by these cytokines.

Progesterone receptor isoform A but not B is expressed in endometriosis.

We previously demonstrated that 17beta hydroxysteroid dehydrogenase type 2, the enzyme that inactivates estradiol to estrone, is expressed in luteal eutopic endometrium in response to progesterone but not in simultaneously biopsied peritoneal endometriotic tissue. This molecular evidence of progesterone resistance, together with the clinical observation of resistance of endometriosis to treatment with progestins, led us to determine the levels of progesterone receptor (PR) isoforms PR-A and PR-B in eutopic endometrial and extra-ovarian endometriotic tissues. It was proposed that progesterone action on target genes is mediated primarily by homodimers of PR-B, whereas the truncated variant PR-A acts as a repressor of PR-B function. Immunoprecipitation, followed by Western blot analysis, was performed to detect bands specific for PR-A and PR-B in paired samples of endometriotic and eutopic endometrial tissues simultaneously biopsed from 18 women undergoing laparoscopy during various phases of the menstrual cycle. PR-B was present in 17 of 18 eutopic endometrial samples, and its level increased in the preovulatory phase, as expected, whereas PR-A was detected in all samples (n = 18) with a similar, but less prominent, cyclic variation in its levels. In endometriotic samples, however, no detectable PR-B could be demonstrated, whereas PR-A was detected in all samples (n = 18), albeit in much lower levels and without any cyclic variation in contrast with the eutopic endometrium. Levels of PR-A and PR-B in endometriotic and eutopic endometrial tissues were determined and compared after normalization to total protein and estrogen receptor-alpha levels. Using RNase protection assay, we also demonstrated indirectly that only PR-A transcripts were present in endometriotic tissue samples (n = 8), whereas both PR-A and PR-B transcripts were readily detectable in all eutopic endometrial samples (n = 8). This was indicative that failure to detect PR-B protein in endometriotic tissues is due to the absence of PR-B transcripts. We conclude that progesterone resistance in endometriotic tissue from laboratory and clinical observations may be accounted for by the presence of the inhibitory PR isoform PR-A and the absence of the stimulatory isoform PR-B.

Deficient 17beta-hydroxysteroid dehydrogenase type 2 expression in endometriosis: failure to metabolize 17beta-estradiol.

Aberrant aromatase expression in stromal cells of endometriosis gives rise to conversion of circulating androstenedione to estrone in this tissue, whereas aromatase expression is absent in the eutopic endometrium. In this study, we initially demonstrated by Northern blotting transcripts of the reductive 17beta-hydroxysteroid dehydrogenase (17betaHSD) type 1, which catalyzes the conversion of estrone to 17beta-estradiol, in both eutopic endometrium and endometriosis. Thus, it follows that the product of the aromatase reaction, namely estrone, that is weakly estrogenic can be converted to the potent estrogen, 17beta-estradiol, in endometriotic tissues. It was previously demonstrated that progesterone stimulates the inactivation of 17beta-estradiol through conversion to estrone in eutopic endometrial epithelial cells. Subsequently, 17betaHSD type 2 was shown to catalyze this reaction, and its transcripts were detected in the epithelial cell component of the eutopic endometrium in secretory phase. Because 17beta-estradiol plays a critical role in the development and growth of endometriosis, we studied 17betaHSD-2 expression in endometriotic tissues and eutopic endometrium. We demonstrated, by Northern blotting, 17betaHSD-2 messenger ribonucleic acid (RNA) in all RNA samples of secretory eutopic endometrium (n=12) but not in secretory samples of endometriotic lesions (n=10), including paired samples of endometrium and endometriosis obtained simultaneously from eight patients. This messenger RNA was not detectable in any samples of proliferative eutopic endometrium or endometriosis (n=4) as expected. Next, we confirmed these findings by demonstration of immunoreactive 17betaHSD-2 in epithelial cells of secretory eutopic endometrium in 11 of 13 samples employing a monoclonal antibody against 17betaHSD-2, whereas 17betaHSD-2 was absent in paired secretory endometriotic tissues (n=4). Proliferative eutopic endometrial (n=8) and endometriotic (n=4) tissues were both negative for immunoreactive 17betaHSD-2, except for barely detectable levels in 1 eutopic endometrial sample. Finally, we sought to determine whether deficient 17betaHSD-2 expression in endometriotic tissues is due to impaired progesterone action in endometriosis. We determined by immunohistochemistry the expression of progesterone and estrogen receptors in these paired samples of secretory (n=4) and proliferative (n=4) eutopic endometrium and endometriosis, and no differences could be demonstrated. In conclusion, inactivation of 17beta-estradiol is impaired in endometriotic tissues due to deficient expression of 17betaHSD-2, which is normally expressed in eutopic endometrium in response to progesterone. The lack of 17betaHSD-2 expression in endometriosis is not due to alterations in the levels of immunoreactive progesterone or estrogen receptors in this tissue and may be related to an inhibitory aberration in the signaling pathway that regulates 17betaHSD-2 expression.

Differential effects of IFN-beta1b on the proliferation of human vascular smooth muscle and endothelial cells.

The effect of human interferon (IFN)-beta1b (Betaseron) on the proliferation of cultured human vascular smooth muscle and endothelial cells was tested in vitro. IFN-beta1b inhibited thymidine incorporation and growth of primary cultures of human aortic and coronary artery smooth muscle in a concentration-dependent manner. The same concentrations of IFN-beta1b did not inhibit thymidine incorporation or growth of primary cultures of human aortic or coronary artery endothelial cells. IFN-beta1b induced the expression of MxA (an antiviral protein induced by type I IFNs) in both smooth muscle and endothelial cells, suggesting that both cell types express receptors for type I IFNs. The growth-inhibitory effect of IFN-beta1b could be mimicked by commercially available human IFN-beta, but not by IFN-alpha2 or IFN-alpha8. The effect of IFN-beta1b was species specific, as it did not inhibit thymidine incorporation in aortic smooth muscle cells derived from pig, rabbit, rat, or mouse. The action of IFN-beta1b on smooth muscle cells persisted for at least 4 days following a 24 h preincubation with IFN-beta1b. Human vascular smooth muscle cells treated with IFN-beta1b did not release lactate dehydrogenase, nor did they show any morphologic change, suggesting that IFN-beta1b was not toxic to the human vascular smooth muscle cells. IFN-beta1b inhibited vascular smooth muscle growth while having no growth-inhibitory effect on endothelial cells obtained from the same blood vessel, making it a potential candidate for treating pathologic conditions where abnormal vascular smooth muscle proliferation is implicated, such as restenosis following balloon angioplasty or smooth muscle proliferation following vascular stenting.

Orexin-A, an hypothalamic peptide with analgesic properties.

The hypothalamic peptide orexin-A and the orexin-1 receptor are localized in areas of the brain and spinal cord associated with nociceptive processing. In the present study, localization was confirmed in the spinal cord and demonstrated in the dorsal root ganglion for both orexin-A and the orexin-1 receptor. The link with nociception was extended when orexin-A was shown to be analgesic when given i.v. but not s.c. in mouse and rat models of nociception and hyperalgesia. The efficacy of orexin-A was similar to that of morphine in the 50 degrees C hotplate test and the carrageenan-induced thermal hyperalgesia test. However, involvement of the opiate system in these effects was ruled out as they were blocked by the orexin-1 receptor antagonist SB-334867 but not naloxone. Orexin-1 receptor antagonists had no effect in acute nociceptive tests but under particular inflammatory conditions were pro-hyperalgesic, suggesting a tonic inhibitory orexin drive in these circumstances. These data demonstrate that the orexinergic system has a potential role in the modulation of nociceptive transmission.

A double-blind randomized placebo controlled trial of thromboprophylaxis in major elective general surgery using once daily injections of a low molecular weight heparin fragment (Fragmin).

The safety and efficacy of the low molecular weight heparin fragment (Fragmin) administered as a single daily injection of 2,500 anti Xa units has been evaluated in 183 patients undergoing major elective general surgery. The study was double-blinded and placebo controlled. The active agent, or placebo, was given subcutaneously with the preoperative medication and continued postoperatively for 5-9 days. Ninety five patients received Fragmin and 88 were randomized to receive the placebo. The clinical characteristics of the two treatment groups were similar. Fragmin significantly reduced the incidence of deep venous thrombosis, as detected by a positive 125I fibrinogen leg scan, relative to the placebo treated patients (4/95, 4.2% v. 14/88, 15.9%; p = 0.008). The thrombotic events occurred predominantly (73%) amongst patients with malignancy. Haemorrhagic endpoints necessitating discontinuation of the trial treatment were 4% in each group. No severe adverse reactions or drug related deaths occurred. These results indicate that 2,500 anti Xa units of Fragmin given only once daily is effective thromboprophylaxis for patients undergoing major elective abdominal surgery.

Calcium regulation in vascular smooth muscle contractility.

The contractile activity of vascular smooth muscle is regulated by control over the cytoplasmic calcium concentration. The intracellular calcium receptor is calmodulin, which, through stimulation of myosin light chain kinase, can activate 2 different contractile states. The calcium is supplied from the sarcoplasmic reticulum and the extracellular space; a minor component is supplied from the inner surface of the plasmalemma. The main intracellular messenger responsible for the transduction of receptor occupation and calcium release from the sarcoplasmic reticulum is IP3 and, to a lesser extent, calcium itself. The superficial location of the sarcoplasmic reticulum in vascular smooth muscle makes it the logical area for control of calcium entry due to calcium leak or through either or both types of calcium channel. The sarcoplasmic reticulum, therefore, acts as a "superficial calcium buffer barrier" and is probably the major system controlling free cytoplasmic calcium concentration in vascular smooth muscle.

EDRF release from canine coronary artery by lectins.

1. The effect of wheat germ agglutinin (WGA), concanavalin A (Con A) and lentil lectin agglutinin (LCA) were investigated on pre-contracted canine coronary artery rings in vitro. 2. In endothelium-intact canine coronary artery, contracted with the thromboxane A2-analogue, U46619, WGA relaxed the tissue in a concentration-dependent manner, with an inhibitory concentration (IC50) of 112 +/- 17 nM (n = 6). In the absence of an endothelium, WGA did not cause any relaxation of the tissue. 3. In endothelium-intact canine coronary artery, contracted with the thromboxane A2-analogue, U46619. LCA relaxed the tissue in a concentration-dependent manner, with an inhibitory concentration (IC50) of 423.1 +/- 41 nM (n = 6). In the absence of an endothelium, LCA produced a 20.1 +/- 1.1% (n = 6) relaxation at the highest concentration tested (3 microM). 4. Concanavalin A (Con A) relaxed canine coronary artery in a partial endothelium-dependent manner with an IC50 of 104 +/- 19 nM on endothelium-intact coronary artery and an IC50 of 1.3 +/- 0.3 microM (n = 6) on endothelium-denuded tissues. 5. The relaxation effects of WGA were attenuated by 1 mM NG-monomethyl L-arginine (L-NMMA) and completely inhibited by haemoglobin (3 microM), methylene blue (10 microM) and LY 83583 (30 microM). Ibuprofen had no effect on WGA-induced relaxation. 6. The relaxant effects of WGA were reversed by addition of 20 mM N-acetyl-D-glucosamine (GlcNAc) and N-acetyl-D-galactosamine (Ga1NAc) but not by alpha-mannose, D-(+)-galactose, and beta-lactose, whereas the endothelin-dependent relaxations to LCA and Con A were unaffected. 7.The endothelium-dependent relaxation induced by the lectins was unaffected by pretreatment of the tissue with 1 microM atropine.8. In the absence of extracellular calcium, WGA was also able to release EDRF suggesting that WGA acts through a second messenger system to release intracellular calcium.9. We suggest that WGA acts as an agonist to release EDRF from endothelial cells possibly by binding to a sugar moiety, specific receptor or adhesion molecules on the endothelial cell surface.

The effects of lectins on the release of EDRF from rabbit aorta.

The effects of the lectins, wheat germ agglutinin (WGA) and concanavalin A (Con A) on endothelium intact pre-contracted rabbit aorta were investigated. WGA (8.3 microM) produced endothelium-dependent relaxations, while Con A (4.3 microM) produced a partially endothelium-dependent relaxation. The endothelium-dependent relaxations to WGA were completely reversed by N-acetylglucosamine, haemoglobin and methylene blue and were partially reversed by NG-monomethyl-L-arginine (L-NMMA). It is suggested that WGA works as an agonist in releasing endothelium-derived relaxing factor (EDRF) from the endothelium of rabbit aorta by interacting with a glycosylated receptor on endothelial cells.

SB-334867-A: the first selective orexin-1 receptor antagonist.

The pharmacology of various peptide and non-peptide ligands was studied in Chinese hamster ovary (CHO) cells stably expressing human orexin-1 (OX(1)) or orexin-2 (OX(2)) receptors by measuring intracellular calcium ([Ca(2+)](i)) using Fluo-3AM. Orexin-A and orexin-B increased [Ca(2+)](i) in CHO-OX(1) (pEC(50)=8.38+/-0.04 and 7.26+/-0.05 respectively, n=12) and CHO-OX(2) (pEC(50)=8.20+/-0.03 and 8.26+/-0.04 respectively, n=8) cells. However, neuropeptide Y and secretin (10 pM - 10 microM) displayed neither agonist nor antagonist properties in either cell-line. SB-334867-A (1-(2-Methyylbenzoxanzol-6-yl)-3-[1,5]naphthyridin-4-yl-urea hydrochloride) inhibited the orexin-A (10 nM) and orexin-B (100 nM)-induced calcium responses (pK(B)=7.27+/-0.04 and 7.23+/-0.03 respectively, n=8), but had no effect on the UTP (3 microM)-induced calcium response in CHO-OX(1) cells. SB-334867-A (10 microM) also inhibited OX(2) mediated calcium responses (32.7+/-1.9% versus orexin-A). SB-334867-A was devoid of agonist properties in either cell-line. In conclusion, SB-334867-A is a non-peptide OX(1) selective receptor antagonist.