Probiotic-educated Tregs are more potent than naïve Tregs for immune tolerance in stressed new-born mice.

When new-born mice are subjected to acute maternal separation stress, cow-milk based formula feeding, and brief recurrent hypoxia with cold stress, they develop gut inflammation similar to the phenotype of neonatal necrotizing enterocolitis, characterised by an increase in gut mucosal effector T (Teffs) and reduced Foxp3+ regulatory T (Tregs) cells. The imbalance can be prevented by probiotic Limosilactobacillus reuteri DSM 17938 (LR 17938). We hypothesised that LR 17938 could potentiate a tolerogenic function of Tregs. To analyse whether LR 17938 can educate Tregs to improve their tolerogenic potency during neonatal stress, we isolated T cells (Tregs and Teffs) from 'donor' mice fed with either LR 17938 (107 cfu) or control media. The cells were adoptively transferred (AT) by intraperitoneal injection (5 × 105 cells/mouse) to new-born (d5) recipient mice. Mice were then separated from their dams, fed formula by gavage, and exposed to hypoxia and cold stress (NeoStress) for 4 days. We analysed the percentage of Tregs in CD4+T helper cells in the intestine (INT) and mesenteric lymph nodes (MLN) of recipient mice. We found that: (1) the percentage of Tregs in the INT and MLN following NeoStress were significantly reduced compared to dam-fed unstressed mice; (2) AT of either naïve Tregs or LR-educated Tregs to mice with Neostress increased the percentage of Tregs in the INT and MLN compared to the percentage in NeoStress mice without Treg treatment; however, LR-educated Tregs increased the Tregs significantly more than naïve Tregs; and (3) AT of LR-educated Tregs reduced pro-inflammatory CD44+Foxp3-NonTregs and inflammatory CX3CR1+ dendritic cells in the intestinal mucosa of NeoStress mice. In conclusion, adoptive transfer of Tregs promotes the generation of and/or migration of endogenous Tregs in the intestinal mucosa of recipient mice. Importantly, probiotic-educated Tregs are more potent than naïve Tregs to enhance immune tolerance following neonatal stress.

A cross-sectional study of non-attendance among patients at a US hemophilia treatment center 2010-2014.

INTRODUCTION: Among patients with chronic disease, non-attendance at scheduled healthcare visits is associated with poor outcomes. The impact of non-attendance among patients with bleeding disorders is unknown. METHODS: Scheduling and medical record data over a 5-year period for all individuals with at least one scheduled appointment during 2010-2014 at a US Hemophilia Treatment Center (HTC) were analysed. Non-attendance rates were calculated as the number of non-attended visits divided by the number of years as a patient during the time period. Consistent non-attenders were patients who did not attend more than one scheduled appointment per person-year on average. Logistic regression determined characteristics associated with consistent non-attendance and Poisson regression estimated adjusted incidence rate ratios (aIRRs) describing associations between non-attendance and emergency department (ED) visits and hospitalizations. RESULTS: There were 8028 appointments scheduled for 950 individuals; 12% were not attended. Consistent non-attenders (n = 62; 7% of the HTC patient population) accounted for over one-third of non-attended appointments and over one-quarter of hospitalizations. Characteristics associated with consistent non-attendance included public health insurance and black race. Higher non-attendance rates were associated with more ED visits (aIRR 1.78; 95% CI: 1.37-2.30) and hospitalizations (aIRR 2.73; 95% CI: 2.18-3.42). Consistent non-attenders had more ED visits (aIRR 2.49; 95% CI: 1.56-3.96) and hospitalizations (aIRR 4.73; 95% CI: 2.96-7.57) compared with patients who never missed appointments. CONCLUSIONS: Frequent non-attendance identified a small but at-risk population. Interventions to improve disease management that target them may have an impact on health outcomes and healthcare utilization.

A randomized comparison between neurostimulation and ultrasound-guided lateral femoral cutaneous nerve block.

BACKGROUND: This prospective, randomized trial compared neurostimulation (NS) and ultrasound (US) guided lateral femoral cutaneous nerve (LFCN) block. We hypothesized that US would result in a shorter total anesthesia-related time (sum of performance and onset times). METHODS: Twenty-one volunteers were enrolled. The right lower limb was randomized to an NS- or US-guided LFCN block. The alternate technique was employed for the left lower limb. With NS, paresthesias were sought in the lateral thigh at a stimulatory threshold of 0.6 mA (pulse width=0.3 ms; frequency=2 Hz) or lower. With US, local anesthetic was deposited under the inguinal ligament, ventral to the iliopsoas muscle. In both groups, 5 mL of lidocaine 2% were used to anesthetize the nerve. During the procedure of the block, the performance time and number of needle passes were recorded. Subsequently, a blinded observer assessed sensory block in the lateral thigh every minute until 20 minutes. Success was defined as loss of pinprick sensation at a point midway between the anterior superior iliac spine and the lateral knee line. The blinded observer also assessed the areas of sensory block in the anterior, medial, lateral, and posterior aspects of the thigh and mapped this distribution onto a corresponding grid. RESULTS: Both modalities provided comparable success rates (76.2%-95.2%), performance times (162.1 to 231.3 seconds), onset times (300.0 to 307.5 seconds) and total anesthesia related-times (480.1 to 554.0 seconds). However US required fewer needle passes (3.2±2.9 vs 9.5±12.2; P=.009). There were no intergroup differences in terms of the distribution of the anesthetized cutaneous areas. However considerable variability was encountered between individuals and between the 2 sides of a same subject. The most common areas of sensory loss included the central lateral two-eighths anteriorly and the central antero-inferior three-eighths laterally. CONCLUSION: Ultrasound guidance and NS provide similar success rates and total anesthesia-related times for LFCN block. The territory of the LFCN displays wide inter- and intra-individual variability.

Reliability of pressure waveform analysis to determine correct epidural needle placement in labouring women.

Pressure waveform analysis provides a reliable confirmatory adjunct to the loss-of-resistance technique to identify the epidural space during thoracic epidural anaesthesia, but its role remains controversial in lumbar epidural analgesia during labour. We performed an observational study in 100 labouring women of the sensitivity and specificity of waveform analysis to determine the correct location of the epidural needle. After obtaining loss-of-resistance, the anaesthetist injected 5 ml saline through the epidural needle (accounting for the volume already used in the loss-of-resistance). Sterile extension tubing, connected to a pressure transducer, was attached to the needle. An investigator determined the presence or absence of a pulsatile waveform, synchronised with the heart rate, on a monitor screen that was not in the view of the anaesthetist or the parturient. A bolus of 4 ml lidocaine 2% with adrenaline 5 µg.ml-1 was administered, and the epidural block was assessed after 15 min. Three women displayed no sensory block at 15 min. The results showed: epidural block present, epidural waveform present 93; epidural block absent, epidural waveform absent 2; epidural block present, epidural waveform absent 4; epidural block absent, epidural waveform present 1. Compared with the use of a local anaesthetic bolus to ascertain the epidural space, the sensitivity, specificity, positive and negative predictive values of waveform analysis were 95.9%, 66.7%, 98.9% and 33.3%, respectively. Epidural waveform analysis provides a simple adjunct to loss-of-resistance for confirming needle placement during performance of obstetric epidurals, however, further studies are required before its routine implementation in clinical practice.

Physician trust and depression influence adherence to factor replacement: a single-centre cross-sectional study.

INTRODUCTION: Poor adherence to factor replacement therapy among patients with haemophilia can lead to joint bleeding and eventual disability. AIM: The aim of this study was to determine patient-related characteristics associated with adherence to factor replacement in adults with haemophilia. METHODS: Adults with haemophilia were recruited to participate in this cross-sectional study. Adherence was measured using either the Validated Hemophilia Regimen Treatment Adherence Scale (VERITAS)-Pro or the VERITAS-PRN questionnaire. Simple and multiple regression analyses that controlled for confounding were performed to determine the association between patient-related characteristics and adherence to factor replacement therapy. RESULTS: Of the 99 subjects enrolled, all were men; 91% had haemophilia A and 78% had severe disease. Age ranged from 18 to 62 years. Most (95%) had functional health literacy; but only 23% were numerate. Mean adherence scores were 45.6 (SD 18) and 51.0 (SD 15) for those on a prophylactic and those on an episodic regimen, respectively, with a lower score indicating better adherence. On multivariable analysis, being on any chronic medication, longer duration followed at our haemophilia treatment centre, higher physician trust and better quality of life were associated with higher adherence. A history of depression was associated with lower adherence. CONCLUSION: Two potentially modifiable characteristics, physician trust and depression, were identified as motivator and barrier to adherence to factor replacement therapy. Promoting a high level of trust between the patient and the healthcare team as well as identifying and treating depression may impact adherence to factor replacement therapy and accordingly reduce joint destruction.

High fat induces acute and chronic inflammation in the hypothalamus: effect of high-fat diet, palmitate and TNF-α on appetite-regulating NPY neurons.

BACKGROUND: Consumption of dietary fat is one of the key factors leading to obesity. High-fat diet (HFD)-induced obesity is characterized by induction of inflammation in the hypothalamus; however, the temporal regulation of proinflammatory markers and their impact on hypothalamic appetite-regulating neuropeptide Y/agouti-related peptide (NPY/AgRP) neurons remains undefined. METHODS: Mice were injected with an acute lipid infusion for 24 h or fed a HFD over 8-20 weeks. Characterized mouse NPY/AgRP hypothalamic cell lines were used for in vitro experimentation. Immunohistochemistry in brain slices or quantitative real-time PCR in cell lines, was performed to determine changes in the expression of key inflammatory markers and neuropeptides. RESULTS: Hypothalamic inflammation, indicated by tumor necrosis factor (TNF)-α expression and astrocytosis in the arcuate nucleus, was evident following acute lipid infusion. HFD for 8 weeks suppressed TNF-α, while significantly increasing heat-shock protein 70 and ciliary neurotrophic factor, both neuroprotective components. HFD for 20 weeks induced TNF-α expression in NPY/AgRP neurons, suggesting a detrimental temporal regulatory mechanism. Using NPY/AgRP hypothalamic cell lines, we found that palmitate provoked a mixed inflammatory response on a panel of inflammatory and endoplasmic reticulum (ER) stress genes, whereas TNF-α significantly upregulated IκBα, nuclear factor (NF)-κB and interleukin-6 mRNA levels. Palmitate and TNF-α exposure predominantly induced NPY mRNA levels. Utilizing an I kappa B kinase ß (IKKß) inhibitor, we demonstrated that these effects potentially occur via the inflammatory IKKß/NF-κB pathway. CONCLUSIONS: These findings indicate that acute lipid and chronic HFD feeding in vivo, as well as acute palmitate and TNF-α exposure in vitro, induce markers of inflammation or ER stress in the hypothalamic appetite-stimulating NPY/AgRP neurons over time, which may contribute to a dramatic alteration in NPY/AgRP content or expression. Acute and chronic HFD feeding in vivo temporally regulates arcuate TNF-α expression with reactive astrocytosis, which suggests a time-dependent neurotrophic or neurotoxic role of lipids.

Distress in patients with bleeding disorders: a single institutional cross-sectional study.

INTRODUCTION: Distress may affect a patient's ability to cope with and manage disease. AIM: To report distress prevalence in adult patients with bleeding disorders and determine whether specific clinical and health characteristics, including disease severity and employment status, are associated with distress. METHODS: Patients who visited a Haemophilia Treatment Centre (HTC) between January 1st, 2012 through February 28th, 2014 and who completed a distress screen, pain screen and questionnaire were evaluated cross sectionally. Distress was measured by the National Comprehensive Cancer Network Distress Management Tool, which allowed patients to rate recent distress on a 0-10 point scale. A rating of five or more was categorized as high distress. Pain was measured by the Brief Pain Inventory Short Form, which asked patients to rate pain types on 0-10 point scales. Patients reported employment and other demographic and behavioural information on the questionnaire. Primary diagnosis, age, HIV and HCV status were abstracted from medical records. Adjusted logistic regression was used to identify distress associations. RESULTS: High distress prevalence among 152 patients with bleeding disorders was 31.6%. Unemployment, disability, greater depressive symptoms and higher pain were associated with high distress in multivariable models. Bleeding disorder diagnosis, race/ethnicity, HIV/HCV status and on-demand treatment regimen were not associated with high distress. CONCLUSION: Distress among patients with congenital bleeding disorders followed at a comprehensive HTC was high and similar to that reported among patients with cancer. Future research should determine whether distress impacts clinical outcomes in patients with bleeding disorders as demonstrated in other chronic disorders.

Forkhead box protein 3(+) regulatory T cells and Helios(+) subset in perinatally acquired HIV.

Forkhead box protein 3 (FoxP3)(+) regulatory T cells (Tregs ) are important not only in regulating the development of autoimmune conditions, but also in chronic infectious diseases. Given their cardinal function in suppressing immune activation, research has focused upon whether they play a detrimental role in chronic infections, particularly HIV. While the role of Tregs in HIV has been investigated intensively, it remains an unresolved topic. However, it is generally accepted that Tregs are susceptible to HIV infection and are preferentially preserved over conventional CD4(+) T cells. It is unknown whether the peripheral-induced or the thymic-derived Tregs are more susceptible to HIV cytotoxicity. It has been recognized that Tregs can be segregated into two subsets based on Helios expression, with the vast majority being Helios(+) . This study examines the impact of HIV infection on total Tregs and their Helios subsets in a perinatal-acquired HIV-infected paediatric population. The finding indicates a selective expansion or survival of Tregs in association with CD4 depletion and increased viraemia. The Helios(+) and Helios(-) subsets within Tregs appear to be equally affected. However, the Helios(+) Tregs seem to be more preserved in patients with low CD4(+) ≤ 25% and detectable plasma HIV RNA >20 copies/ml. In this group, the frequencies of Tregs are increased, but their numbers appear insufficient to restrain immune activation. In conclusion, our findings suggest that both Helios subsets of Tregs are susceptible to HIV infection and are preferentially preserved compared to conventional CD4(+) T cells.

Actual colonoscopy: what are the risks of perforation?

Recent studies have suggested that virtual colonoscopy (VC) and actual colonoscopy (AC) have similar efficacy for detection of polyps >6 mm. However, procedural risks with emerging technology such as VC need to be assessed before widespread implementation. We propose to demonstrate complication rates after AC that can be used for a comparative benchmark in VC. From 1994 to 1999 all patients undergoing AC who sustained perforation that required operation were analyzed for the mortality and complications. There were 26,162 consecutive colonoscopies that required 21 operations for perforation. Of these 16,948 (65%) colonoscopies were diagnostic and 9,214 (35%) were therapeutic with 11 (0.06%) and 10 (0.11%) operations respectively. Overall risk for colonoscopic perforation that requires operation was one in 1,246 (one in 1,541 for diagnostic and one in 921 for therapeutic). Five perforations were oversewn, 15 were resected (five with stoma), and one was drained. One patient died. There were two reoperations. Mortality was 0.006 per cent (one in 16,948) for diagnostic and zero for therapeutic colonoscopy. Overall risk for perforation that requires operation or mortality after AC is low. Virtual colonoscopists who propose screening and subsequent therapeutic interventions need to report high volume without complications as the perforation rate requiring operation was one in 1,246.

Promoting cancer prevention activities among Vietnamese physicians in California.

BACKGROUND: To promote prevention and early detection of cancer, the authors conducted a three-year intervention targeting Vietnamese physicians in solo practice in California. METHODS: Twenty subjects who had received their medical training in Vietnam were recruited into a randomized controlled trial. The intervention included computerized or manual cancer screening reminders, continuing medical education seminars, Vietnamese-language health education materials, newsletters, and oncology data-query programs. Evaluation included chart audits for eight targeted activities pre- and post-intervention. RESULTS: Before the intervention, annual physician performance rates were low for all eight activities: routine checkups (65.6%), Pap testing (13.8%), pelvic examinations (19.8%), clinical breast examinations (13.3%), mammography (6.4%), hepatitis B serologies (21.9%), hepatitis B immunizations (12.8%), and smoking cessation counseling (1.6%). After the intervention, performance rates increased significantly for smoking cessation counseling (p = 0.02), Pap testing (p = 0.004), and pelvic examinations (p = 0.01). CONCLUSIONS: The results demonstrate the efficacy of an intervention targeting Vietnamese primary care physicians in promoting smoking cessation counseling, Pap testing, and pelvic examinations, but not other cancer prevention activities.

Needle-localized breast biopsy for mammographic abnormalities: a community hospital experience.

Increased awareness of benefits of early detection of breast cancer has resulted in increased numbers of screening mammographies and breast biopsies for nonpalpable lesions. Tertiary hospital studies have demonstrated positive biopsy rates from abnormal mammographic findings at 18 to 32 per cent. We examined the effectiveness of needle biopsy for nonpalpable radiographic abnormalities in our community hospital. We reviewed 167 records of patients biopsied over a 2-year period. Mammographic assessment, biopsy, and pathological assessment were performed using accepted methods. Malignancy was detected in 34 of 167 biopsies (20%). The biopsy yield rate was highest for mammographic findings of spiculated or stellate masses (75%, P < 0.01). Most biopsies (83%) were performed because of mammographic findings of microcalcifications or circumscribed enlarging masses/nodular developing densities for a positive biopsy yield rate of 16 per cent. Rates were higher in patients with personal (44%) or family history (30%) of breast cancer and in postmenopausal women (30%). These results demonstrate that 1) factors such as age, personal or family history of breast cancer, and certain mammographic features of breast lesions are associated with high biopsy yield rates, and 2) the biopsy yield rate in our community setting is comparable to tertiary hospital experience.

Synergistic responses of steroidal estrogens in vitro (yeast) and in vivo (turtles).

Many environmental agents exert estrogenic activity. Previous studies from our laboratories demonstrated that certain combinations of environmental estrogens (i) reverse the sex of male turtle embryos in a synergistic manner (Bergeron et al., (1994) Environ. Hlth Perspect. 102, 780-782), and (ii) synergistically transactivate the human estrogen receptor (hER) in yeast and mammalian cells (Arnold et al., (1996) Science 272, 1489-1492). Because our findings with synthetic estrogens suggested that combinations of naturally-occurring steroidal estrogens might also produce synergistic activity of the ER, we used the same model systems to measure the activity of combinations of steroidal estrogens. The activity of combinations of estrone, estradiol-17beta or estradiol-17alpha in yeast strains expressing hER was synergistic at submaximal concentrations of both estrogenic compounds. However, synergy was not observed with mixtures of estrogens when the concentration of one of the estrogens alone was maximally active in yeast. Ligand-binding assays in yeast performed with various radiolabeled estrogens suggested that multiple estrogens may interact with the receptor. The estrogen-dependent process of sex-reversal of turtle embryos incubated at a male-producing temperature was used to determine whether steroidal estrogens also had synergistic activity in vivo. In this instance, a combination of estriol and estradiol-17beta was effective in reversing the gonadal sex of turtle embryos from males to females in a synergistic manner. Our results suggest that the synergy of some combinations of estrogens, synthetic or steroidal, may play a role in the estrogen-dependent process of sexual development in certain species.

Evaluation of clinical and environmental anti-estrogens with human estrogen receptor expressed in Saccharomyces cerevisiae: a novel role for ABC-cassette transporters in mediating anti-estrogenic activity.

The effectiveness of anti-estrogens in treating estrogen-dependent diseases is limited by the acquired resistance of some diseases to anti-estrogens. This effect could occur by the export of anti-estrogens by cell membrane transport proteins. To study this phenomenon we have expressed human estrogen receptor (hER) and an estrogen-sensitive reporter in wild-type yeast and two transport-defective strains. In the wild-type strain, the most effective anti-estrogen was nafoxidine. 4-Hydroxy tamoxifen and clomiphene were inactive whereas tamoxifen had significant inhibitory activity in the wild-type strain. Using a strain missing the ABC-cassette transporter Snq2, clomiphene had anti-estrogenic activity. 4-Hydroxy tamoxifen had anti-estrogenic activity only in yeast lacking the transporter Pdr5. Whole cell binding assays indicated that 4-hydroxy tamoxifen is exported by Pdr5. Environmental chemicals such as polychlorinated biphenyls function as partial estrogens and anti-estrogens in yeast. In the absence of Pdr5 or Snq2, the estrogenic activity of 4-hydroxy, 2',4',6'-trichloro biphenyl (3-PCB) was substantially reduced in comparison to its activity in the wild-type strain. Interestingly, the antiestrogenic activity of 3-PCB was equivalent in the wild-type and transporter-defective strains. Our results suggest a novel role for ABC-cassette transporters in regulating the activity of clinical and environmental anti-estrogens.

Several synthetic chemicals inhibit progesterone receptor-mediated transactivation in yeast.

The human progesterone receptor (hPR) B-form and a progesterone-sensitive reporter were expressed in yeast and used to screen a library of synthetic chemicals for their ability to function as agonists or antagonists of hPR. The transcriptional activity of hPR was not increased in the presence of over 40 individual chemicals. Seven chemicals decreased progesterone-dependent activity in yeast. The most effective chemicals were 6-hydroxychrysene, 1-hydroxypyrene, 4-hydroxy, 2',4',6'-trichloro biphenyl, and 4-hydroxy, 2',3',4',5'-tetrachloro biphenyl. The decrease of progesterone-mediated transactivation strongly correlated with their displacement of [3H]progesterone from hPR. The absence of the hydroxyl group on the above chemicals completely abolished their inhibitory activity. The other chemicals which decreased progesterone activity were endosulfan II, endosulfan sulfate, and lindane. These chemicals did not inhibit [3H]progesterone binding, suggesting that they inhibit progesterone action by interacting with a region of hPR distinct from binding [3H]progesterone or by a mechanism independent of hPR. These results highlight the utility of yeast for screening hormonally-active chemicals. In addition, hydroxylation appears to be essential for the interaction of some chemicals with hPR.

The anti-estrogenic activity of selected polynuclear aromatic hydrocarbons in yeast expressing human estrogen receptor.

Polynuclear aromatic hydrocarbons (PAH) represent a large class of chemicals present in environment. We have used yeast strain ER(wt) expressing human estrogen receptor (hER) and an estrogen-sensitive reporter to characterize the estrogenic or anti-estrogenic activities of 21 PAHs. The PAHs did not exhibit estrogenic activity in yeast strain ER(wt). Four of the PAHs, dibenz[a,h]anthracene, 6-hydroxy-chrysene, 2,3-benzofluorene, and benzo(a)pyrene, inhibited estradiol-dependent reporter activity in strain ER(wt). A mutant hER lacking the amino terminus expressed in yeast was inhibited by the four PAHs to a lesser extent than the full-length hER. 6-hydroxy-chrysene, 2,3-benzofluorene, and benzo(a)pyrene, but not dibenz[a,h]anthracene, weakly displaced [3H]estradiol from the hER in a competition binding assay. A strong correlation between the inhibition of [3H]estradiol-binding from the hER and the reduction of hER-mediated transactivation in yeast was not observed. These observations suggest that the PAHs dibenz[a,h]-anthracene, 6-hydroxy-chrysene, 2,3-benzofluorene, and benzo(a)pyrene may antagonize activity in yeast by binding to an anti-estrogen binding site on the hER or by mechanisms independent of the hER.

Inhibition of progesterone receptor activity in yeast by synthetic chemicals.

Numerous synthetic chemicals have estrogenic activity by interacting with the estrogen receptor. In this report, we test the hypothesis that some estrogenic chemicals may also modulate the human progesterone receptor (hPR) signaling pathway. This was evaluated by examining synthetic chemicals for their ability to modulate the activity of hPR expressed in yeast. The transcriptional activity of hPR was not increased in the presence of several synthetic chemicals. However, the estrogenic chemicals p-nonylphenol and 4-tert-octyphenol, and pentachlorophenol, effectively inhibited the activity of the hPR in yeast. Competition binding studies indicated these chemicals effectively competed with radiolabeled R5020, a synthetic progestin, for binding to the hPR in yeast. These results indicate that some synthetic chemicals directly inhibit the activity of hPR in yeast. The observations that some estrogenic chemicals can also inhibit hPR activity suggest a potential mechanism for the reported potent estrogenic activities of these chemicals.

The inhibition of estrogen receptor-mediated responses by chloro-S-triazine-derived compounds is dependent on estradiol concentration in yeast.

The chloro-S-triazine derived compounds atrazine, atrazine desisopropyl, cyanazine, and simazine are commonly used herbicides. These compounds do not have estrogenic activity in yeast expressing human estrogen receptor (hER) and an estrogen-sensitive reporter. In the presence of a concentration of estradiol (20 nM) that induced maximal reporter activity in yeast, the triazines did not inhibit reporter activity. However, the triazines decreased reporter activity in a dose dependent manner in the presence of a submaximal concentration of estradiol (0.5 nM). The estradiol-dependent activity of a mutant hER lacking the amino terminus was not inhibited by the triazines in yeast. Competition binding assays demonstrated that the triazines displaced radiolabeled estradiol from recombinant hER. These results suggest that the ability of the triazines to inhibit estrogen receptor-mediated responses in yeast occur through their interaction with hER and is dependent on the concentration of estradiol.