Focal adhesion kinase contributes to proliferative potential of ErbB2 mammary tumour cells but is dispensable for ErbB2 mammary tumour induction in vivo

Introduction

Neu (HER2/ErbB2) is overexpressed in 25% to 30% of human breast cancer, correlating with a poor prognosis. Researchers in previous studies who used the mouse mammary tumor virus Neu-transgenic mouse model (MMTV-Neu) demonstrated that the Neu-YB line had increased production of CXCL12 and increased metastasis, whereas the Neu-YD line had decreased metastasis. In this study, we examined the role of increased production of CXCL12 in tumor cell invasion and malignancy.

Methods

We studied invasion in the tumor microenvironment using multiphoton intravital imaging, in vivo invasion and intravasation assays. CXCL12 signaling was altered by using the CXCR4 inhibitor AMD3100 or by increasing CXCL12 expression. The role of macrophage signaling in vivo was determined using a colony-stimulating factor 1 receptor (CSF-1R) blocking antibody.

Results

The Neu-YD strain was reduced in invasion, intravasation and metastasis compared to the Neu-YB and Neu deletion mutant (activated receptor) strains. Remarkably, in the Neu-YB strain, in vivo invasion to epidermal growth factor was dependent on both CXCL12-CXCR4 and CSF1-CSF-1R signaling. Neu-YB tumors had increased macrophage and microvessel density. Overexpression of CXCL12 in rat mammary adenocarcinoma cells increased in vivo invasion as well as microvessel and macrophage density.

Conclusions

Expression of CXCL12 by tumor cells results in increased macrophage and microvessel density and in vivo invasiveness.     

Blood pressure effects of intravenous apomorphine in conscious deoxycorticosterone-acetate salt-hypertensive rats

SUMMARY: The present study reports the effects of apomorphine (APO) on blood pressure and the principal site of action of this agonist in 4-week deoxycorticosterone-acetate (DOCA)-hypertensive conscious rats. In these preprations, intravenous (i.v.) administration of APO (0.50-1 mg/kg) induced short-lasting and dose-dependent decreases in mean arterial pressure. The hypotensive response to APO (0.3 mg/kg) was reversed into a significant pressor effect by i.v. hexamethonium (30 mg/kg), whereas it was enhanced by i.v. pretreatment with the vasopressor antagonist of arginine vasopressin (AVP) d(CH2)5Tyr(Me)AVP (10 microg/kg) and/or prazosin (1 mg/kg). This depressor effect was suppressed by the central and peripheral dopamine D2 receptor antagonist metoclopramide (5 mg/kg i.v.), unaffected by the selective dopamine D1 receptor antagonist SCH 23390 (0.2 mg/kg i.v.), partly reduced by intrathecal domperidone (40 microg per rat at T9-T10 level), a dopamine D2 receptor antagonist which does not cross the blood-brain barrier, and reversed into a significant pressor effect by i.v. domperidone (0.5 mg/kg). The latter pressor effect was fully abolished by combined i.v. pretreatment with the vasopressor antagonist of AVP and prazosin. These results show that, in conscious DOCA salt-hypertensive rats, APO induced a brief, initial depressor effect, which is opposed to a central pressor component. The depressor component is related to an inhibition of norepinephrine transmission through activation of dopamine D2 receptors, some of which are located in the spinal cord and some of which are located in the peripheral circulation. The central pressor component, which became manifest after peripheral dopamine D2 receptor blockade, appears to be related to an increase in vasopressin release and sympathetic tone through activation of brain dopamine D2 receptors.     

Blunted central bromocriptine-induced tachycardia in conscious,malnourished rats

Bromocriptine-induced tachycardia, persisting after adrenalectomy, is mediated by central dopamine D2 receptor stimulation through activation of the sympathetic outflow to the heart. The present study investigated the effects of malnutrition during pregnancy on bromocriptine-induced tachycardia in adult conscious rats. Malnourished rats were obtained by feeding dams a multideficient diet (providing 8% protein) during mating and pregnancy. Birth weight was significantly reduced in malnourished rats when compared to control rats born to dams fed standard commercially diet (23% protein) during mating and pregnancy. Baseline mean aortic pressure and heart rate in malnourished rats were comparable to those of well-nourished rats. Tachycardia (33+/-9 beats/min.), but not the hypotensive response to intravenous bromocriptine (150 microg/kg) was significantly reduced in malnourished rats, compared with control rats (70+/-10 beats/min.). In malnourished rats, pretreatment with intravenous domperidone (500 microg/kg) blocked the bromocriptine-induced hypotension, without affecting the tachycardia. Neither cardiac vagal (40+/-6 beats/min.) nor sympathetic tone (76+/-6 beats/min.) was significantly altered by multideficient diet-induced malnutrition (51+/-6 and 67+/-10 beats/min., respectively). In isolated perfused heart preparations from malnourished rats, positive inotropic response to isoproterenol (10-8 to 10-4 M) was not significantly different compared to that in control rats. In summary, malnutrition during foetal life blunted the bromocriptine-induced tachycardia, an effect that could be related to central dopamine D2 receptor desensitization rather than to impairment of autonomic regulation of the heart or cardiac beta-adrenoceptor desensitization.     

Protective effects of selenium on methimazole-induced anemia and oxidative stress in adult rats and their offspring

The present study investigates the potential ability of selenium, considered as an antioxidant with pharmacological property to alleviate oxidative stress and hematological parameter disorders induced by methimazole, an antithyroid drug. Pregnant Wistar rats were randomly divided into four groups of six each: group I served as negative control and received a standard diet; group II received 250 mg/L of methimazole in drinking water and a standard diet; group III received both methimazole (250 mg/L, orally) and selenium (0.5 mg/kg of diet) supplemented to the standard diet; group IV served as positive control and received a supplement of selenium in the diet (0.5 mg/kg of diet) as sodium selenite (Na(2)SeO(3)). Treatment was started from the 14th day of pregnancy until day 14 after delivery. Methimazole reduced the number of red blood cells, hemoglobin concentration and hematocrit in mothers and their pups. Besides, plasma iron, vitamins B(9), B(12), C and E levels were reduced. Lipid peroxidation increased, objectified by high malondialdehyde levels and lactate dehydrogenase activity in plasma, while glutathione, glutathione peroxidase, superoxide dismutase and catalase activities showed a significant decline. Co-administration of selenium through diet improved all the parameters cited above. It can be concluded that the administration of selenium alleviates methimazole-induced toxicity, thus demonstrating its antioxidant efficacy.     

Evaluation of the antioxidant, anti-inflammatory and hepatoprotective properties of vanillin in carbon tetrachloride-treated rats

The antioxidant and anti-inflammatory effects of vanillin are considered as important forces in the protection against liver injury and fibrosis. This study investigated the protective effects of vanillin against carbon tetrachoride (CCl(4))-induced hepatotoxicity in rat. Pretreatment with vanillin prior the administration of CCl(4) significantly prevented the decrease of protein synthesis and the increase in plasma alanine (ALT) and aspartate (AST) aminotransferases. Furthermore, it inhibited hepatic lipid peroxidation (MDA) and protein carbonyl (PCO) formation and attenuated the (CCl(4))-mediated depletion of antioxidant enzyme catalase and superoxide dismutase (SOD) activities and glutathione level (GSH) in the liver. In addition, vanillin markedly attenuated the expression levels of pro-inflammatory cytokines such as tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6) and prevented CCl(4)-induced hepatic cell alteration and necrosis, as indicated by liver histopathology. These findings suggest that the antioxidant and anti-inflammatory effects of vanillin against CCl(4)-induced acute liver injury may involve its ability to block CCl(4)-generated free radicals.     

Central bromocriptine-induced tachycardia is reversed to bradycardia in conscious, deoxycorticosterone acetate-salt hypertensive rats

A central dopaminergic origin has been demonstrated for the bromocriptine-induced tachycardia in conscious, normotensive rats. The present study investigated the effect of bromocriptine on heart rate and the principal site of action of this agonist in conscious, deoxycorticosterone acetate-salt hypertensive rats, in which altered central dopaminergic activity has been previously reported. Intravenous administration of bromocriptine (150 microg/kg) increased heart rate (49+/-5 beats/min.) in uninephrectomized control rats, while it induced a significant bradycardia (50+/-6 beats/min.) in deoxycorticosterone acetate-salt hypertensive rats. In the latter animals, intravenous (500 microg/kg) or intrathecal (40 microg/rat at T9-T10) pretreatment with domperidone, a selective dopamine D2 receptor antagonist that does not cross the blood-brain barrier, reduced partially, but significantly, the bradycardiac responses to bromocriptine (reduction of about 44% and 48% of the maximal effect, respectively). In contrast, the bromocriptine-induced bradycardia was fully abolished by intravenous pretreatment with metoclopramide (300 microg/kg), a dopamine D2 receptor antagonist that crosses the blood-brain barrier, or by combined pretreatment with intravenous and intrathecal domperidone. These results indicate that, in deoxycorticosterone acetate-salt hypertensive rats, bromocriptine decreases rather than increases heart rate, an effect that is mediated partly through a peripheral D2 dopaminergic mechanism and partly through stimulation of spinal dopamine D2 receptors. They further support the concept that, in normotensive, conscious rats, the central tachycardia of bromocriptine appears to predominate and to mask the bradycardia of this agonist at both peripheral and spinal dopamine D2 receptors.     

Rostrocaudal localization of cardiovascular responses induced by intrathecal administration of apomorphine in conscious, freely moving rats.

In conscious freely moving rats, administration of apomorphine (179 nmol), a dopamine receptor agonist, into the intrathecal (i.t.) space decreased mean aortic blood pressure (MBP) and heart rate (HR). Both the magnitude and the time of appearance of the response varied according to the spinal level of administration. The largest and immediately appearing effect was observed after the injection at the upper thoracic site, whereas the magnitude of the responses was smaller with an immediate or slightly delayed (0.5-1.5 min) onset at lower thoracic and midcervical levels. More caudal responses appeared to be due to spreading of the drug along the spinal axis (onset in 1-2 min after administration). Behavioral responses (stereotyped movements) were observed within 2-3 min after administration and were nearly the same whatever the site of administration. These results corroborate, as do those provided by i.t. injections of tritiated apomorphine, the spinal origin of cardiovascular effects of i.t. apomorphine. Furthermore, spinal transection at the T5-T7 level did not change the magnitude and duration of decreases in MBP and HR elicited by i.t. apomorphine injected at the T2-T4 level. Moreover, this procedure enhanced responses to i.t. administration at the T9-T10 level. In conclusion, these results favor the existence of a spinal site of action for the cardiovascular effects of apomorphine. Furthermore, they indicate that spinal transection is accompanied by development of a hypersensitive phenomenon (of a mechanism to be determined).     

Identification by array-Comparative Genomic Hybridization (array-CGH) of a large deletion of luteinizing hormone receptor gene combined with a missense mutation in a patient diagnosed with a 46,XY disorder of sex development and application to prenatal diagnosis

This paper reports the case of an infant presenting with sexual ambiguity at birth. The child presented with labia majora synechia, thready genital tubercle and perineal hypospadias. The karyotype was 46,XY. Low testosterone levels with no response to hCG administration, associated with high LH level for her age, high FSH level, high inhibin B levels and normal AMH indicated a lack of LH receptivity and prompted us to screen the LHCGR gene for mutations. A previously described missense mutation (p.Cys131Arg) was identified at homozygous state in the propositus and at heterozygous state in the mother. This variation, however, was not found in the father. Our attention was drawn by the presence of several single nucleotide polymorphisms (SNPs), identified at homozygous state without any paternal contribution from exon 1 to exon 10 of LHCGR, suggesting a paternal deletion. Array DNA analysis was performed revealing a large deletion extending from 61,493 to 135,344 bp and including the LHCGR gene. Adequate genetic counselling was provided. This paper describes the first application of prenatal diagnosis in LHCGR deficiency for 46,XY disorders of sex development with the subsequent delivery of a normal baby.