Ultrasound simulation training to meet the 2021 Royal College of Radiologists' curriculum for radiology trainees: South East London experience.

AIM: To enhance ultrasound teaching delivery to radiology trainees using a simulation course matched to the 2021 Royal College of Radiologists (RCR) curriculum. MATERIAL AND METHODS: An ultrasound simulation training course was designed for specialty trainees (ST) 1 in radiology, which was based on the 2021 RCR curriculum and covered the top ultrasound training priorities. The course was piloted initially on two occasions in a 1-day format to the August 2021 and the March 2022 ST1 intake trainees. Based on the feedback, a comprehensive 4-day course was developed and delivered between October and December 2022 for the August 2022 ST1 intake, funded by Health Education England. The outcomes measured were subjective trainee feedback using numerical scores and free text. RESULTS: All King's College Hospital NHS Foundation Trust radiology ST1 trainees from the August 2021 to the August 2022 intake participated in ultrasound simulation training. The training matched the RCR curriculum and increased the trainees' confidence and competency in medical ultrasound. CONCLUSIONS: Ultrasound simulation training can be successfully delivered to ST1 trainees to match the 2021 RCR curriculum and enhance training in medical ultrasound for radiologists.

Background levels of micro-organisms in the busy urban environment of transport hubs.

AIMS: We constantly interact with our surrounding microbiome, including the micro-organisms present in highly populated public places. However, data on everyday exposure to background levels of micro-organisms are limited. To address this, bacteria and fungi were collected and enumerated in settled dust from railway stations. METHODS AND RESULTS: Samples were collected weekly for 52 weeks, from up to three pre-determined surfaces in each of 17 railway stations in England and Scotland. Trained staff at each station took surface wipes, sending them to the laboratory for culture-based analysis for total bacteria and fungi. Maximum yields of bacteria at the stations were 107 -108 colony forming units (CFU) per cm2 , and 104 -105 CFU per cm2 for fungi. CONCLUSIONS: There was evidence of seasonal trends, with bacterial numbers rising from spring through to winter, while fungal numbers peaked in autumn. Microbial numbers were similar in samples taken at the same time at a given station. Influences on contamination levels were likely to be a combination of passenger numbers and station layout, with dust generated from construction work also contributing. SIGNIFICANCE AND IMPACT OF THE STUDY: A baseline of typical human exposure to micro-organisms in public transport hubs was established through the generation of a comprehensive database.

A longitudinal assessment of periodontal health status in 53 Labrador retrievers.

OBJECTIVES: To determine the incidence and rates of progression of gingivitis and periodontitis in Labrador retrievers. MATERIALS AND METHODS: Fifty-three dogs, aged 1·1 to 5·9 years, had their periodontal health assessed every 6 months for up to 2 years. The extent of gingivitis and periodontitis was measured around the whole gingival margin of every tooth under general anaesthesia. RESULTS: All dogs had gingivitis at the initial assessment. The majority (64·2%) of tooth aspects had very mild gingivitis. The palatal/lingual aspect of all tooth types was most likely to show bleeding when probed: 63·0% of these aspects had mild or moderate gingivitis. Over 2 years, 56·6% of dogs developed periodontitis and dogs as young as 1·9 years were affected. There was a significant positive correlation between the proportion of teeth with periodontitis and age. In total, 124 teeth (5·7%) developed periodontitis; 88 (71·0%) of these were incisors. The palatal/lingual aspect of the incisors developed the disease first (2·8% of incisor aspects). CLINICAL SIGNIFICANCE: Periodontitis developed in regions that are difficult to see in conscious dogs implying that detection and treatment of disease requires periodic sedation or anaesthesia.

Testicular Microlithiasis: Is Sonographic Surveillance Necessary? Single Centre 14 Year Experience in 442 Patients with Testicular Microlithiasis.

PURPOSE: Increased prevalence of germ cell tumour (GCT) is seen with testicular microlithiasis (TM) suggesting TM is a premalignant condition with US surveillance advocated. We present a cohort of patients with TM followed up in a single centre and deliberate on the value of US surveillance. MATERIALS AND METHODS: A retrospective analysis of subjects with underlying US diagnosis of TM between 1998 and 2012. One-yearly US follow-up was offered to all patients with TM and a database maintained. Any co-existing tumour at presentation with TM was recorded. TM was divided into limited (< 5 microliths/field), classical (≥ 5 microliths/field) and florid ('snowstorm' appearance). Patient demographics, follow-up details and the development of any scrotal abnormalities were recorded. The radiological and histological findings were documented when a testicular lesion occurred during the follow-up period. RESULTS: 20 224 patients were examined: 867/20 224 (4.3 %) had TM. 21/867 (2.4 %) patients had histology proven malignant tumours at presentation. All TM patients consented to follow-up with 442/867 (51.0 %) achieving this and entering into a follow-up program (mean duration 28 months, range 8 - 165 months). Two patients developed primary GCT during the follow up period. One patient (limited TM) had undergone a previous orchiectomy for contralateral GCT and developed a palpable mass at follow up month 21. The other (limited TM) had an atrophic testis; a tumour was found on US at follow up month 62. CONCLUSION: Two patients of 442 (0.5 %) followed up for all forms of TM in a single centre developed a GCT over a mean duration of 28 months, both had independent risk factors for the development of GCT. These findings suggest that US surveillance is not required when TM is the only abnormality in the absence of any clinical risk factors for the development of GCT.

The point mutation UCH-L1 C152A protects primary neurons against cyclopentenone prostaglandin-induced cytotoxicity: implications for post-ischemic neuronal injury.

Cyclopentenone prostaglandins (CyPGs), such as 15-deoxy-Δ(12,14)-prostaglandin J2 (15dPGJ2), are reactive prostaglandin metabolites exerting a variety of biological effects. CyPGs are produced in ischemic brain and disrupt the ubiquitin-proteasome system (UPS). Ubiquitin-C-terminal hydrolase L1 (UCH-L1) is a brain-specific deubiquitinating enzyme that has been linked to neurodegenerative diseases. Using tandem mass spectrometry (MS) analyses, we found that the C152 site of UCH-L1 is adducted by CyPGs. Mutation of C152 to alanine (C152A) inhibited CyPG modification and conserved recombinant UCH-L1 protein hydrolase activity after 15dPGJ2 treatment. A knock-in (KI) mouse expressing the UCH-L1 C152A mutation was constructed with the bacterial artificial chromosome (BAC) technique. Brain expression and distribution of UCH-L1 in the KI mouse was similar to that of wild type (WT) as determined by western blotting. Primary cortical neurons derived from KI mice were resistant to 15dPGJ2 cytotoxicity compared with neurons from WT mice as detected by the WST-1 cell viability assay and caspase-3 and poly ADP ribose polymerase (PARP) cleavage. This protective effect was accompanied with significantly less ubiquitinated protein accumulation and aggregation as well as less UCH-L1 aggregation in C152A KI primary neurons after 15dPGJ2 treatment. Additionally, 15dPGJ2-induced axonal injury was also significantly attenuated in KI neurons as compared with WT. Taken together, these studies indicate that UCH-L1 function is important in hypoxic neuronal death, and the C152 site of UCH-L1 has a significant role in neuronal survival after hypoxic/ischemic injury.

Cancer mortality in the meat and delicatessen departments of supermarkets (1950-2006).

Meat cutters and meat wrappers in the meat department of supermarkets are exposed to oncogenic viruses present in raw meat from cattle, pigs, sheep, and poultry, and their products (unpasteurized milk and raw eggs). Up to the mid 1970s, meat wrappers were also exposed to carcinogens present in fumes emitted from the machine used to wrap meat. Because of this we studied cancer mortality in a cohort of 10,701 workers in the meat and delicatessen departments of supermarkets, and we report here the findings after the third follow-up. Standardized mortality ratios (SMR) were estimated in the cohort as a whole and in race/sex subgroups, using the US population for comparison. Study subjects were followed up from January 1950 to December 2006. Significantly increased SMRs of 1.3 (95% CI, 1.2-1.5), and 2.7 (95% CI, 1.2-5.3) were recorded for cancers of the lung, and tonsils/oropharynx, respectively, in the entire cohort, affecting nearly all race/sex subgroups. SMRs of 4.6 (95% CI, 1.0-13.6) for cancer of the floor of the mouth, and 2.8 (95% CI, 1.3-5.3) for cancer of the gall bladder and biliary tract were recorded only in White male meatcutters. Significantly decreased SMRs were observed for a few cancers. It is not known if the observed excess of cancers is a result of occupational exposures. However, substantial evidence points to fumes from the wrapping machine as a possible candidate for explaining the excess in female meat wrappers. Nested case-control studies that can examine risks from occupational exposures in greater detail, and adequately control for confounding factors are now needed, to permit specifically investigate the role of the oncogenic viruses, fumes and non-occupational risk factors in the occurrence of these cancers. The findings are important, not only occupationally but also because the general population may also experience these exposures, albeit to a lesser degree.

Frailty and Pain: Two Related Conditions.

Frailty is a multidimensional syndrome, involving functional, nutritional, biological and psychological aspects. This condition, defined as a decreased resistance to internal and external stressors, is predictive of adverse health outcomes, including disability and mortality. Importantly, the frailty syndrome is usually considered a reversible condition, thus amenable of specific preventive interventions. Persistent pain in older adults is very common and has multiple determinants. This symptom represents a determinant of accelerated aging. In the present paper, we discuss available evidence examining the association between these two conditions. Despite the high prevalence of these two conditions and their shared underlying mechanisms, our search only retrieved few relevant studies. Most of them reported a relationship between pain (or analgesics consumption) and different operational definitions of frailty. Pain may represent a relevant risk factor as well as a potential target for interventions against the frailty syndrome, but further studies are needed.

Co-Presence of Multimorbidity and Disability with Frailty: An Examination of Heterogeneity in the Frail Older Population.

BACKGROUND: Frailty is often associated with multimorbidity and disability. OBJECTIVES: We investigated heterogeneity in the frail older population by characterizing five subpopulations according to quantitative biological markers, multimorbidity and disability, and examined their association with mortality and nursing home admission. DESIGN: Observational study. PARTICIPANTS: Participants (n=4,414) were from the population-based Age Gene/Environment Susceptibility Reykjavik Study. MEASUREMENTS: Frailty was defined by ≥ 3 of five characteristics: weight loss, weakness, reduced energy levels, slowness and physical inactivity. Multimorbidity was assessed using a simple disease count, based on 13 prevalent conditions. Disability was assessed by five activities of daily living; participants who had difficulty with one or more tasks were considered disabled. Differences among frail subpopulations were based on the co-presence of multimorbidity and disability. Differences among the following subpopulations were examined: 1) Non-frail (reference group); 2) Frail only; 3) Frail with disability; 4) Frailty with multimorbidity; 5) Frail with disability and multimorbidity. RESULTS: Frailty was present in 10.7% (n=473). Frailty was associated with increased risk for mortality (OR 1.40; 95% CI 1.15-1.69) and nursing home admission (OR 1.50; 95% CI 1.16-1.93); risks differed by subpopulations. Compared to the non-frail, the frail only group had poorer cognition and increased inflammation levels but did not have increased risk for mortality (OR 1.40; 95% CI 0.84-2.33) or nursing home admission (OR 1.01; 95% CI 0.46-2.21). Compared to the non-frail, the other frail subpopulations had significantly poorer cognition, increased inflammation levels, more white matter lesions, higher levels of calcium, glucose and red cell distribution width and increased risk for mortality and nursing home admission. CONCLUSIONS: The adverse health risks associated with frailty in the general older adult population may primarily be driven by increased disease burden and disability.

Genetically diverse Clostridium difficile strains harboring abundant prophages in an estuarine environment.

Clostridium difficile is the leading cause of antibiotic-associated diarrheal disease in health care settings across the world. Despite its pathogenic capacity, it can be carried asymptomatically and has been found in terrestrial and marine ecosystems outside hospital environments. Little is known about these environmental strains, and few studies have been conducted on estuarine systems. Although prophage abundance and diversity are known to occur within clinical strains, prophage carriage within environmental strains of C. difficile has not previously been explored. In this study, we isolated C. difficile from sites sampled in two consecutive years in an English estuarine system. Isolates were characterized by PCR ribotype, antibiotic resistance, and motility. The prevalence and diversity of prophages were detected by transmission electron microscopy (TEM) and a phage-specific PCR assay. We show that a dynamic and diverse population of C. difficile exists within these sediments and that it includes isolates of ribotypes which are associated with severe clinical infections and those which are more frequently isolated from outside the hospital environment. Prophage carriage was found to be high (75%), demonstrating that phages play a role in the biology of these strains.

On the difficulties of isolating Clostridium difficile from hospital environments.

Spores of Clostridium difficile were deposited on to a stainless steel surface and subsequently exposed to a chlorine-releasing disinfectant (dichloroisocyanurate). Recovery of the spores was carried out using RODAC plates containing a variety of selective and non-selective agars. The non-selective agar media yielded higher recoveries of both control and chlorine-stressed spores. Our results show that the antibiotics used in selective media imposed an additional stress on both disinfectant-treated and untreated spores resulting in considerably reduced recoveries. This could lead to a serious underestimate of the extent of environmental contamination by this organism.

Development and in vivo evaluation of silver sulfadiazine loaded hydrogel consisting polyvinyl alcohol and chitosan for severe burns.

A new Hydrogel containing silver Sulfadiazine (SSD) was developed for enhanced burns wound healing. The hydrogel was prepared by cross-linking of PVA and Chitosan by freeze thawing method. Their gel properties, moisture retaining capacity, fluid uptake capacity, in vitro release study, in vivo burn healing effect were evaluated. Chitosan and PVA cross linking decreased gel fraction upto 70% determined the good gel properties. This cross linked hydrogel increased the Swelling ratio and Water vapour transmission rate (WVTR) which provides the sustained release of drug and moist environment for healing respectively. The hydrogel containing 7.5% of PVA, 0.75% of chitosan found to have increased gel strength, higher water vapour transmission rate and fluid uptake capacity suitable for faster healing of burns. This hydrogel also sustained the release of 1% SSD required for longer antimicrobial activity and found better in vivo burn healing capacity as compared to marketed preparation. Thus hydrogel containing 7.5% of PVA, 0.75% of chitosan and 1% SSD is a potential burns dressing with better gel properties and excellent burns healing capacity.

Effect of vitamin C deficiency during postnatal development on adult behavior: functional phenotype of Gulo-/- knockout mice.

Organisms using oxygen for aerobic respiration require antioxidants to balance the production of reactive oxygen species during metabolic processes. Various species--including humans and other primates--suffer mutations in the GULO gene encoding L-gulono-γ-lactone oxidase; GULO is the rate-limiting enzyme in the biosynthesis of ascorbate, an important cellular antioxidant. Animals lacking the ability to synthesize vitamin C develop scurvy without dietary supplementation. The Gulo-/- knockout (KO) mouse requires oral supplemental vitamin C; without this supplementation the animal dies with a scorbutic condition within several weeks. Vitamin C is known to be most abundant in the brain, where it is believed to play important roles in neuroprotection, neurotransmission and neuromodulation. We therefore hypothesized that ascorbate deficiency in Gulo-/- KO mice might lead to an abnormal behavioral phenotype. We established the amount of ascorbate in the drinking water (220 ppm) necessary for generating a chronic low-ascorbate status in the brain, yet clinically the mice appeared healthy throughout 100 days postpartum at which time all behavioral-phenotyping tests were completed. Compared with Gulo+/+ wild-type littermates, ascorbate-deficient Gulo-/- mice were found to be less active in moving in their environment; when in water, these mice swam more slowly in some tests, consistent with a mild motor deficit. We found no evidence of cognitive, anxiety or sensorimotor-gating problems. Despite being less active, Gulo-/- mice exhibited exaggerated hyperactivity to the dopaminergic agonist methamphetamine. The subnormal movement, combined with hypersensitivity to a dopamine agonist, point to developmental ascorbate deficiency causing long-term striatal dysfunction.

European ancestry and resting metabolic rate in older African Americans.

BACKGROUND/OBJECTIVES: Resting metabolic rate (RMR) contributes 60-80% of total energy expenditure and is consistently lower in populations of African descent compared with populations of European populations. Determination of European ancestry (EA) through single nucleotide polymorphism (SNP) analysis would provide an initial step for identifying genetic associations that contribute to low RMR. We sought to evaluate the association between RMR and EA in African Americans. SUBJECTS/METHODS: RMR was measured by indirect calorimetry in 141 African American men and women (aged 74.7±3.0 years) enrolled in a substudy of the Health, Aging and Body Composition Study. Ancestry informative markers were used to estimate individual percent EA. Multivariate regression was used to assess the association between RMR and EA after adjustments for soft tissue fat-free mass (STFFM), fat mass, age, study site, physical activity level and sex. RESULTS: Mean EA was 23.8±16% (range: 0.1-70.7%) and there were no differences by sex. Following adjustments, each percent EA was associated with a 1.6 kcal/day (95% Confidence interval: 0.42, 2.7 kcal/day) higher RMR (P=0.008). This equates to a 160 kcal/day lower RMR in a population of completely African ancestry, with one of completely European ancestry. Additional adjustment for trunk STFFM that partially accounts for high-metabolic rate organs did not affect this association. CONCLUSIONS: EA in African Americans is strongly associated with higher RMR. The data suggest that population differences in RMR may be due to genetic variants.

Targeting CYP450 modulation to decrease the risk of induced cataract in the experimental model.

BACKGROUND: Diabetes is one of the major causes of cataract. Some drugs prescribed for the treatment of diabetes are the modulators of CYP450, which may alter the risk of cataract. OBJECTIVE: To study the effect of CYP450 modulation in galactosemic cataract. MATERIALS AND METHODS: Male Sprague-Dawley suckling rats were allotted to four groups (n = 6), as follows: Group 1: Normal control, Group 2: Galactose control, Group 3: CYP450 inhibitor pretreated and Group 4: CYP450 inducer pretreated. Cataract was induced in animals of all groups except group 1 by feeding them galactose (50%), 21 days after parturition. From the eighteenth day of life, CYP450 inhibitor (nifedipine; 8.1 mg/kg) and CYP450 inducer (pioglitazone; 3.8 mg/kg) were given orally to groups 3 and 4, respectively. The maturation pattern of the cataract was observed by an operating microscope, every third day. Biochemical changes in the lenses of all groups, for example, CYP450 activity expressed as ΅M NADPH oxidized / unit time, alterations in the levels of total proteins, soluble proteins, and reduced glutathione (GSH) following the induction of cataract, were estimated. RESULTS: The microscopic examination of the lenses indicated that CYP450 inhibitor pre-treatment delayed (fourteenth day) the occurrence of cataract, while CYP450 inducer pretreatment demonstrated an early (ninth day) cataract as compared to galactose control rats (twelfth day). A significant decrease and increase in CYP450 activity was observed with the CYP450 inhibitor and inducer pre-treatment, respectively. There was no alteration in the GSH level, but a significant increase in total and soluble protein was found in groups 3 and 4 as compared to group 2. CONCLUSION: CYP450 may have a role in the initiation of cataract without any effect on the maturation pattern, as revealed by the delayed occurrence of cataract with the CYP450 inhibitor and an early onset of cataract with the CYP450 inducer.

Evaluation of the effect of Myrica sapida on bronchoconstriction and bronchial hyperreactivity.

The present investigation was undertaken to evaluate the bronchodilator and bronchial hyperreactivity of the stem bark of Myrica sapida. Experimental models studied were histamine induced bronchospasm in guinea pigs, bronchoalveolar lavage fluid (BALF) in egg albumin sensitized guinea pigs, histamine release from the lung tissues of sensitized guinea pigs and histopathological studies. Ethanolic extract of M. sapida (75 mg/kg, p.o., for 7 days) showed significant protection against histamine aerosol induced bronchospasm. Significant decrease in the total and differential leukocyte counts in BALF and prevention of egg albumin induced histamine release from chopped lung tissues of sensitized guinea pigs was observed on chronic administration of ethanolic extract of M. sapida (75 mg/kg, p.o., for 15 days). Histological examination of the section of lung from sensitized guinea pigs treated with ethanolic extract of M. sapida (75 mg/kg, p.o., for 15 days) was comparable to that of the control group. These results suggest that M. sapida possesses not only bronchodilator activity but also decreases bronchial hyperresponsiveness by decreasing the infiltration of inflammatory mediators like eosinophils, neutrophils in BALF and inhibiting histamine release from lungs of sensitized guinea pigs.

Characterisation of smoking behaviour across the life course and its impact on decline in lung function and all-cause mortality: evidence from a British birth cohort.

OBJECTIVES: To describe smoking trajectories from early adolescence into mid-life and to examine the effects of these trajectories on health and all-cause mortality. METHODS: A nationally representative birth cohort study including 3387 men and women followed up since their birth in 1946 in England, Scotland and Wales. The main outcome measure is all-cause mortality by age 60 years and rate of decline in forced expiratory volume in 1 second (FEV(1)). RESULTS: Eighteen per cent of the sample were categorised as lifelong smokers (smokers at all six waves at ages 20, 25, 31, 36, 43, 53 years), of whom 90% had begun smoking by age 18 years. By age 60 years, 10% of all lifelong smokers had died. They had a threefold increase in mortality rate compared with never smokers (hazard ratio (HR) 3.2, 95% confidence interval (CI) 2.1 to 4.8). For predominantly smokers (smokers for at least four of the six data collections), mortality rate remained higher than never smokers (HR 1.6, 95% CI 1.0 to 2.5). Predominantly non-smokers did not differ from those who never smoked (HR 1.3, 95% CI 0.9 to 2.0). Using the most recent smoking status available, current smokers had more than double the risk of mortality compared with never smokers (HR 2.4, 95% CI 1.6 to 3.5). Lifelong smokers and predominantly smokers had a greater rate of decline in lung function than never smokers (regression coefficients -18 ml/year, 95% CI -22 to -13; -6, 95% CI -10.3 to -1.7 respectively). For current smokers, the decline was 8.4 ml/year (95% CI -12.0 to -5.0) faster than never smokers. CONCLUSIONS: The strength and differentiation of adverse effects identified by using simplified smoking behaviours has highlighted the advantages of obtaining further information on lifelong smoking behaviour from former smokers, rather than just current smoking status.

Studies in release behavior of diltiazem HCl from matrix tablets containing (hydroxypropyl)methyl cellulose and xanthan gum.

(Hydroxypropyl)methyl cellulose and xanthan gum were used as hydrophilic matrixing agents for preparing modified release tablets of diltiazem HCl. The amount of (Hydroxypropyl)methyl cellulose and xanthan gum exhibited significant effect on drug release from the tablets prepared by direct compression technique. Xanthan gum showed a higher ability to retard the drug release than (Hydroxypropyl)methyl cellulose. A 2(2) + 1 factorial design was adopted to study the effect of amount of (Hydroxypropyl)methyl cellulose and xanthan gum on percent drug released in first hour (Y60) and the time required for 90% drug dissolution (t90). A response surface plot is generated for investigating the effect of the independent variables on t90. The tablets containing 90 mg diltiazem HCl, 45 mg (Hydroxypropyl)methyl cellulose and 45 mg xanthan gum showed drug release upto 12 h. The value of similarity factor, f2, for the selected batch was found to be 85.1 when the dissolution study was carried out in water or simulated gastric fluid, indicating pH independent drug dissolution. The selected batch also showed a comparable release profile with a market product (f2 = 60.2). Linear relationship was observed between percent drug released and degree of swelling. The kinetics of the drug release fitted well to the Hixson-Crowell equation. It can be concluded that by using a suitable blend of (Hydroxypropyl)methyl cellulose and xanthan gum desired modified drug release can be achieved.

The relationship between human papillomavirus and p53 gene in conjunctival squamous cell carcinoma.

PURPOSE: The p53 tumor-suppressor gene has been documented to exist in mutated forms in many types of squamous cell carcinoma in the body. Also in conjunctival squamous cell carcinoma, human papillomavirus (HPV) is accepted as an oncogenic factor. The objective of our study was to establish a correlation between p53 overexpression and the presence of HPV infection within tumor tissues from patients with conjunctival squamous cell carcinoma. METHODS: Tissue sections obtained from paraffin-embedded conjunctival squamous cell carcinoma specimens from 23 patients were examined with light microscopy, polymerase chain reaction (PCR), and immunohistochemistry. RESULTS: Seventy-eight percent of tumors were positive for p53, whereas 22% were positive for HPV. The proportion of patients positive for both p53 and HPV was 17%, whereas another 17% of the patients were negative for both p53 and HPV. Therefore no significant disproportion was found in the distribution of patients' HPV status and p53 status (p = 1.00). No significant correlation or linear association was found between the HPV status and p53 status (r = 0.022; p = 0.920). CONCLUSION: We could not show any statistical association between abnormal p53 gene-product expression by immunohistochemistry in conjunctival squamous cell carcinomas and HPV infection by PCR detection techniques.

Stimulation or endothelin-1 secretion by human breast cancer cells through protein kinase A activation: a possible novel paracrine loop involving breast fibroblast-derived prostaglandin E2.

Breast cancer cells secrete endothelin-1 (ET-1), which may act as a paracrine mitogen in breast tumours. The paracrine factors and signal transduction pathways responsible for regulating ET-1 production in breast cancer are unknown. In this study we have examined the involvement of the protein kinase A (PKA) signalling pathway in the control of ET-1 secretion in the human breast cancer cell line MCF-7. Treatment of MCF-7 cells with various agents that activate protein kinase A (PKA) through increases in intracellular cAMP levels including forskolin, cholera toxin (ChT), the cAMP analogue 8-Br-cAMP, or the cAMP phosphodiesterase inhibitor, 3-isobutyl-1-methyl-xanthine (IBMX) all markedly increased ET-1 release. Prostaglandin E2 (PGE2) while stimulating cAMP production, but not inositol lipid hydrolysis also significantly stimulated ET-1 release. Activation of PKC by 2-O-tetradecanoyl phorbol 13-acetate (TPA) also stimulated ET-1 secretion in MCF-7 cells. The PKA inhibitor H-89 attenuated the ET-1 response to PGE2, forskolin and ChT, but not that due to the PKC agonist TPA. The possibility that human breast fibroblasts (HBFs) are a target for ET-1 action with regard to PGE2 production was also investigated, and revealed that while HBFs were unresponsive to ET-1 alone, pretreatment with the cytokine IL-beta greatly potentiated PGE2 release in response to ET-1. In conclusion our results show that activation of either the PKA or PKC signalling pathways in human breast cancer cells increases ET-1 secretion. We also found that HBFs release PGE2 after treatment with ET-1 and that PGE2 itself stimulates ET-1 production in MCF-7 cells. The implication of this potential novel paracrine loop may be significant in view of the high levels of PGE2 and ET-1 found in malignant breast tissues.