Breast conservation therapy utilizing partial breast brachytherapy for early-stage cancer of the breast: a retrospective review from the Saint Luke's Cancer Institute.

BACKGROUND: Accelerated partial breast irradiation (APBI) is a convenient alternative to whole-breast irradiation, as less overall time is needed for completion. The use of APBI outside the framework of large prospective clinical trials has markedly increased. To our knowledge, no high-volume, community-based breast program has published their experience with APBI. METHODS: The records of 93 consecutive patients who underwent APBI utilizing Mammosite Radiation Therapy System from 2005 to 2010 at Saint Luke's Cancer Institute in Kansas City, MO, were retrospectively reviewed. The Kaplan-Meier method was used to estimate the ipsilateral breast recurrence rate and recurrence-free survival. RESULTS: Median age at diagnosis was 63 years (range, 45 to 86 y) and mean follow-up was 29 months. Patient stratification ASTRO consensus classifications for APBI was 37% suitable, 57% cautionary, and 6% unsuitable. The 3-year breast control rate was 98.7%. Three-year overall recurrence-free survival was 94.4%, and 3-year mastectomy-free survival was 97.4%. Using univariate analysis, no tumor or patient factors were associated with ipsilateral breast recurrence. However, tumor grade (P<0.05), stage (P=0.04), estrogen receptor status (P<0.001), progesterone receptor status (P<0.001), tumor size (P<0.001), and ASTRO suitability criteria (P=0.027) were associated with overall recurrence-free survival. No differences were observed when outcomes of patients with ductal carcinoma in situ were compared with those with invasive disease. CONCLUSIONS: In our high-volume community-based program, APBI outcomes are comparable with those reported from large academic institutions. We also found relationships between tumor stage, grade, negative estrogen receptor status, and ASTRO suitability criteria with overall recurrence rates. The continued careful application of APBI in appropriately selected patients appears warranted until phase III trials comparing this modality to whole-breast irradiation have matured.

Fatality involving complications of bupivacaine toxicity and hypersensitivity reaction.

This case represents unusual findings of elevated bupivacaine and tryptase concentrations following local anesthetic, bupivacaine, administered as a scalene nerve block for elective rotator cuff repair surgery. Following bupivacaine injection, the patient exhibited almost immediate seizure activity, bradycardia, and cardiac arrest. Resuscitative efforts including cardiopulmonary bypass restored a cardiac rhythm. However, the clinical medical status of the patient progressively declined and he died 7 h following administration of the local anesthetic. Autopsy revealed several abnormalities of the heart including cardiomegaly, myocardial bridging, and lipomatous hypertrophy of the intraatrial septum, which may have contributed to bradycardia and arrhythmia. Postmortem toxicology results revealed elevated bupivacaine and tryptase concentrations. Elevated postmortem bupivacaine concentrations 7 h following administration and abrupt onset of seizures indicate unintentional intravascular injection instead of nerve and tissue infiltration. An elevated postmortem tryptase concentration points to the possibility of a hypersensitivity reaction to bupivacaine.

Adenosine receptor regulation of coronary blood flow in Ossabaw miniature swine.

Adenosine clearly regulates coronary blood flow (CBF); however, contributions of specific adenosine receptor (AR) subtypes (A(1), A(2A), A(2B), A(3)) to CBF in swine have not been determined. ARs generally decrease (A(1), A(3)) or increase (A(2A), A(2B)) cyclic adenosine monophosphate, a major mediator of vasodilation. We hypothesized that A(1) antagonism potentiates coronary vasodilation and coronary stent deployment in dyslipidemic Ossabaw swine elicits impaired vasodilation to adenosine that is associated with increased A(1)/A(2A) expression. The left main coronary artery was accessed with a guiding catheter allowing intracoronary infusions. After placement of a flow wire into the left circumflex coronary artery the responses to bolus infusions of adenosine were obtained. Steady-state infusion of AR-specific agents was achieved by using a small catheter fed over the flow wire in control pigs. CBF was increased by the A(2)-nonselective agonist 2-phenylaminoadenosine (CV1808) in a dose-dependent manner. Baseline CBF was increased by the highly A(1)-selective antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), but not changed by other AR-specific agents. The nonselective A(2) antagonist 3,7-dimethyl-1-propargylxanthine and A(2A)-selective antagonist 4-(2-[7-amino-2-(2-furyl)[1,2,4]triazolo[2,3-a][1,3,5]triazin-5-ylamino]ethyl)phenol (ZM241385) abolished adenosine-induced CBF, whereas A(2B) and A(3) antagonism had no effect. Dyslipidemia and stenting decreased adenosine-induced CBF ∼70%, whereas A(1), A(2A), and A(2B) mRNA were up-regulated in dyslipidemic versus control >5-fold and there was no change in the ratio of A(1)/A(2A) protein in microvessels distal to the stent. In control Ossabaw swine A(1) antagonism by DPCPX positively regulated basal CBF. Impaired adenosine-induced CBF after stenting in dyslipidemia is most likely caused by the altered balance between A(1) and A(2A) signaling, not receptor expression.

Metabolic syndrome and coronary artery disease in Ossabaw compared with Yucatan swine.

Metabolic syndrome (MetS), a compilation of associated risk factors, increases the risk of type 2 diabetes and coronary artery disease (CAD, atherosclerosis), which can progress to the point of artery occlusion. Stents are the primary interventional treatment for occlusive CAD, and patients with MetS and hyperinsulinemia have increased restenosis. Because of its thrifty genotype, the Ossabaw pig is a model of MetS. We tested the hypothesis that, when fed high-fat diet, Ossabaw swine develop more features of MetS, greater native CAD, and greater stent-induced CAD than do Yucatan swine. Animals of each breed were divided randomly into 2 groups and fed 2 different calorie-matched diets for 40 wk: control diet (C) and high-fat, high-cholesterol atherogenic diet (H). A bare metal stent was placed in the circumflex artery, and pigs were allowed to recover for 3 wk. Characteristics of MetS, macrovascular and microvascular CAD, in-stent stenosis, and Ca(2+) signaling in coronary smooth muscle cells were evaluated. MetS characteristics including, obesity, glucose intolerance, hyperinsulinemia, and elevated arterial pressure were elevated in Ossabaw swine compared to Yucatan swine. Ossabaw swine with MetS had more extensive and diffuse native CAD and in-stent stenosis and impaired coronary blood flow regulation compared with Yucatan. In-stent atherosclerotic lesions in Ossabaw coronary arteries were less fibrous and more cellular. Coronary smooth muscle cells from Ossabaw had impaired Ca(2+) efflux and intracellular sequestration versus cells from Yucatan swine. Therefore, Ossabaw swine are a superior model of MetS, subsequent CAD, and cellular Ca(2+) signaling defects, whereas Yucatan swine are leaner and relatively resistant to MetS and CAD.

Short-term exercise training prevents micro- and macrovascular disease following coronary stenting.

The purpose of this study was to determine the effects of exercise on coronary blood flow and macrovascular atherosclerosis in response to stent deployment. Male Yucatan swine were placed on a control diet (C); on a high-fat/cholesterol diet (hypercholesterolemic; H); or on a high-fat/cholesterol diet and aerobically exercise trained (HX) starting after 36 wk on the diet. All pigs underwent coronary angiography and intravascular ultrasound (IVUS) guided placement of a bare metal stent in the circumflex coronary artery after 40 wk on diets and 3 wk later pigs underwent repeat angiography and IVUS and coronary blood flow (CBF) measurement. Average peak velocity (APV) was measured under basal conditions and in response to intracoronary application of the endothelium-independent vasodilator adenosine and the endothelium-dependent vasodilator bradykinin. There was a similar approximately 8-fold increase in total cholesterol in H and HX compared with control. Baseline CBF was increased above control and H in HX (P<0.05). At all doses adenosine-induced CBF was impaired in H, but preserved in HX. Similarly, bradykinin-induced CBF was impaired in H vs. control, yet was potentiated in HX. Microvessel density was decreased in H and preserved in HX vs. control. Native atheroma in HX was lower relative to H and control, while in-stent stenosis in HX was not different from H. Hyperlipidemia-induced microvascular dysfunction after stent deployment may be a result of reduction in microvessel density. This is the first report that short-term exercise training near the time of stenting prevents stent-induced microvascular dysfunction and attenuates native atheroma independent of changes in plasma cholesterol in this porcine model.

Exercise training decreases store-operated Ca2+entry associated with metabolic syndrome and coronary atherosclerosis.

AIMS: Stenting attenuates restenosis, but accelerated coronary artery disease (CAD) adjacent to the stent (peri-stent CAD) remains a concern in metabolic syndrome (MetS). Smooth muscle cell proliferation, a major mechanism of CAD, is mediated partly by myoplasmic Ca2+ dysregulation and store-operated Ca2+ entry (SOCE) via canonical transient receptor potential 1 (TRPC1) channels is proposed to play a key role. Exercise is known to prevent Ca2+ dysregulation in CAD. We tested the hypothesis that MetS increases SOCE and peri-stent CAD and exercise attenuates these events. METHODS AND RESULTS: Groups (n = 9 pigs each) were (i) healthy lean Ossabaw swine fed standard chow, (ii) excess calorie atherogenic diet fed (MetS), and (iii) aerobically exercise trained starting after 50 weeks of development of MetS (XMetS). Bare metal stents were placed after 54 weeks on diets, and CAD and SOCE were assessed 4 weeks later. Coronary cells were dispersed proximal to the stent (peri-stent) and from non-stent segments, and fura-2 fluorescence was used to assess SOCE, which was verified by Ni2+ blockade and insensitivity to nifedipine. XMetS pigs had increased physical work capacity and decreased LDL/HDL (P < 0.05), but no attenuation of robust insulin resistance, glucose intolerance, hypertriglyceridaemia, or hypertension. CAD was greater in peri-stented vs. non-stented artery segments. MetS had the greatest CAD, SOCE, and TRPC1 and STIM1 mRNA and protein expression, which were all attenuated in XMetS. CONCLUSION: This is the first report of the protective effect of exercise on native CAD, peri-stent CAD, SOCE, and molecular expression of TRPC1, STIM1, and Orai1 in MetS.

Leukemia inhibitory factor is upregulated in coronary arteries of Ossabaw miniature swine after stent placement.

Leukemia inhibitory factor (LIF), an IL-6 class cytokine, is reported to be antiatherosclerotic. Thus, we hypothesized that LIF expression might be altered during in-stent neointimal hyperplasia. Ossabaw miniature swine, a unique large-animal model of metabolic syndrome and cardiovascular disease, were used for these studies. Bare-metal stents were deployed in the left anterior descending and left circumflex coronary arteries. Stents were expanded to either 1.0 x luminal diameter (in accordance with current clinical practice) or 1.3 x (overexpansion). The development of in-stent neointimal hyperplasia was assessed 28-day postimplantation using intravascular ultrasound. The atherosclerotic coverage of the vessel wall was approximately five-fold higher in 1.0 x stents and approximately nine-fold higher in 1.3 x stents 4 weeks after deployment, compared with the same segments before stenting. LIF mRNA was elevated approximately 11-fold in stented segments, relative to unstented epicardial coronary arteries. LIF expression and the intima : media ratio were strongly correlated in 1.0 x stented vessels. Further studies to investigate the nature of the association between LIF and neointimal hyperplasia revealed that vascular smooth muscle cell proliferation was inhibited by LIF treatment in an in-vitro model of atherosclerosis (coronary artery organ culture). These novel and clinically relevant studies show that elevated LIF gene expression is predictive for in-stent neointimal hyperplasia, and suggest that LIF upregulation may be a compensatory mechanism in this setting.

Acute inhibition of the betaine transporter by ATP and adenosine in renal MDCK cells.

Extracellular ATP interacts with purinergic P2 receptors to regulate a range of physiological responses, including downregulation of transport activity in the nephron. ATP is released from cells by mechanical stimuli such as cell volume changes, and autocrine signaling by extracellular ATP could occur in renal medullary cells during diuresis. This was tested in Madin-Darby canine kidney (MDCK) cells, a model used frequently to study P1 and P2 receptor activity. ATP was released within 1 min after transfer from 500 to 300 mosmol/kgH2O medium. A 30-min incubation with ATP produced dose-dependent inhibition (0.01-0.10 mM) of the renal betaine/GABA transporter (BGT1) with little effect on other osmolyte transporters. Inhibition was reproduced by specific agonists for P2X (alpha,beta-methylene-ATP) and P2Y (UTP) receptors. Adenosine, the final product of ATP hydrolysis, also inhibited BGT1 but not taurine transport. Inhibition by ATP and adenosine was blocked by pertussis toxin and A73122, suggesting involvement of inhibitory G protein and PLC in postreceptor signaling. Both ATP and adenosine (0.1 mM) produced rapid increases in intracellular Ca2+, due to the mobilization of intracellular Ca2+ stores and Ca2+ influx. Blocking these Ca2+ increases with BAPTA-AM also blocked the action of ATP and adenosine on BGT1 transport. Finally, immunohistochemical studies indicated that inhibition of BGT1 transport may be due to endocytic accumulation of BGT1 proteins from the plasma membrane. We conclude that ATP and adenosine, through stimulation of PLC and intracellular Ca2+, may be rapidly acting regulators of BGT1 transport especially in response to a fall in extracellular osmolarity.

Impaired capsaicin-induced relaxation of coronary arteries in a porcine model of the metabolic syndrome.

Recent studies implicate channels of the transient receptor potential vanilloid family (e.g., TRPV1) in regulating vascular tone; however, little is known about these channels in the coronary circulation. Furthermore, it is unclear whether metabolic syndrome alters the function and/or expression of TRPV1. We tested the hypothesis that TRPV1 mediates coronary vasodilation through endothelium-dependent mechanisms that are impaired by the metabolic syndrome. Studies were conducted on coronary arteries from lean and obese male Ossabaw miniature swine. In lean pigs, capsaicin, a TRPV1 agonist, relaxed arteries in a dose-dependent manner (EC50 = 116 +/- 41 nM). Capsaicin-induced relaxation was blocked by the TRPV1 antagonist capsazepine, endothelial denudation, inhibition of nitric oxide synthase, and K+ channel antagonists. Capsaicin-induced relaxation was impaired in rings from pigs with metabolic syndrome (91 +/- 4% vs. 51 +/- 10% relaxation at 100 microM). TRPV1 immunoreactivity was prominent in coronary endothelial cells. TRPV1 protein expression was decreased 40 +/- 11% in obese pigs. Capsaicin (100 microM) elicited divalent cation influx that was abolished in endothelial cells from obese pigs. These data indicate that TRPV1 channels are functionally expressed in the coronary circulation and mediate endothelium-dependent vasodilation through a mechanism involving nitric oxide and K+ channels. Impaired capsaicin-induced vasodilation in the metabolic syndrome is associated with decreased expression of TRPV1 and cation influx.

Adenosine A1 receptors in neointimal hyperplasia and in-stent stenosis in Ossabaw miniature swine.

BACKGROUND: Stent-induced neointimal hyperplasia is a major cause of morbidity following stent deployment in patients with coronary artery disease. Importantly, however, mechanisms underlying stent-induced neointimal hyperplasia are unclear. This pathological response to stent placement is more aggressive when stents are over-expanded, suggesting that vascular injury may play a role. In this study we tested the hypothesis that adenosine A1 receptor upregulation is associated with neointimal hyperplasia within coronary artery stents. METHODS: Adult male Ossabaw swine were used as our experimental model. Neointima formation and gene expression were studied 4 weeks after coronary stents were placed at 1.0x or 1.3x luminal diameter. RESULTS: Neointima formation was observed in 1.0x stents and more than doubled in 1.3x stents, thus verifying the response to overexpansion injury. A1 receptor mRNA was increased four-fold and seven-fold in stents at 1.0x and 1.3x luminal diameter, suggesting that increased A1 receptor activity might contribute to stent-induced neointimal hyperplasia. Coronary artery organ culture model of arterial injury demonstrated A1 receptor activation increased DNA synthesis three-fold, an effect abolished by A1 receptor antagonism. CONCLUSION: Our data indicate that A1 receptor expression is increased within stents and that activation of A1 receptors increases smooth muscle cell proliferation. We suggest that inhibition of A1 receptor signaling may be a promising therapeutic target for management of in-stent stenosis.

Fetuin-A uptake in bovine vascular smooth muscle cells is calcium dependent and mediated by annexins.

Fetuin-A is a known inhibitor of vascular calcification in vitro. In arteries with calcification, there is increased immunostaining for fetuin-A. However, vascular smooth muscle cells (VSMC) do not synthesize fetuin-A, suggesting fetuin-A may be endocytosed to exert its inhibitory effects. To examine the mechanism by which fetuin-A is taken up in bovine VSMC (BVSMC), we examined living cells by confocal microscopy and determined the uptake of Cy5-labeled fetuin-A. The results demonstrated that fetuin-A was taken up in BVSMC only in the presence of extracellular calcium, whereas phosphorus had no effect. Additional studies demonstrated the calcium-dependent uptake was specific for fetuin-A and only observed in BVSMC and osteoblasts, but not epithelial, endothelial, or adipose cells. The uptake was dose dependent, but could not be inhibited by excess unlabeled fetuin-A, suggesting a fluid phase rather than a receptor-mediated process. Fetuin-A also induced a sustained increase in intracellular calcium in BVSMC in the presence of extracellular calcium, whereas there was no increase in the absence of extracellular calcium. To further characterize the uptake, we utilized an inhibitor of annexin calcium channel activity, demonstrating inhibition of both fetuin-A uptake and intracellular calcium increase. Finally, we demonstrate that fetuin-A binds to annexin II at the cell membrane of BVSMC. In summary, our study demonstrates calcium- and annexin-dependent uptake of fetuin-A that leads to a sustained rise in intracellular calcium. This regulated uptake may be a mechanism by which fetuin-A inhibits VSMC calcification in the presence of excess calcium.

Inhibition of the renal betaine transporter by calcium ions.

Chronic upregulation of the renal betaine/GABA transporter (BGT1) by hypertonic stress has been well documented, but it is not known whether BGT1 can be regulated acutely after insertion in the basolateral plasma membrane. Related transporters, such as the rat brain GABA transporter, can be rapidly removed from the plasma membrane through activation of G protein-coupled receptors. The goal of the present study was to determine whether acute changes in extracellular and/or intracellular Ca(2+) will regulate BGT1 transport activity at the plasma membrane level in Madin-Darby canine kidney cells subjected to 24-h hypertonic stress. After brief pretreatment with a Ca(2+)-free solution, the addition of extracellular Ca(2+) in the transport assay produced dose-dependent inhibition of Na(+)-GABA cotransport. Maximum inhibition was 49% at 2 mM Ca(2+) (P < 0.05). Fura 2 imaging confirmed that addition of 2 mM Ca(2+) produced a transient increase in intracellular Ca(2+) that preceded transport inhibition. Acute inhibition of Na(+)-GABA cotransport was reproduced by addition of thapsigargin (5 microM) and ionomycin (10 microM). Amino acid transport system A, assayed as a control, was not inhibited. Brief treatment with phorbol esters reproduced the specific inhibition of Na(+)-GABA cotransport, and the inhibition was blocked by staurosporine. Surface biotinylation confirmed that the response to phorbol esters was accompanied by loss of BGT1 protein from the plasma membrane, and immunohistochemistry showed a shift to an intracellular distribution. We conclude that BGT1 can be inhibited acutely by extracellular Ca(2+) through a mechanism involving BGT1 protein internalization, and protein kinase C may play a role.