What are the barriers and facilitators to effective health promotion in urgent and emergency care? A systematic review.

BACKGROUND: There are potential health gains such as reducing early deaths, years spent in ill-health and costs to society and the health and care system by encouraging NHS staff to use encounters with patients to help individuals significantly reduce their risk of disease. Emergency department staff and paramedics are in a unique position to engage with a wide range of the population and to use these contacts as opportunities to help people improve their health. The aim of this research was to examine barriers and facilitators to effective health promotion by urgent and emergency care staff. METHODS: A systematic search of the literature was performed to review and synthesise published evidence relating to barriers and facilitators to effective health promotion by urgent and emergency care staff. Medical and social science databases were searched for articles published between January 2000 and December 2021 and the reference lists of included articles were hand searched. Two reviewers independently screened the studies and assessed risk of bias. Data was extracted using a bespoke form created for the study. RESULTS: A total of 19 papers were included in the study. Four themes capture the narratives of the included research papers: 1) should it be part of our job?; 2) staff comfort in broaching the topic; 3) format of health education; 4) competency and training needs. Whilst urgent and emergency care staff view health promotion as part of their job, time restraints and a lack of knowledge and experience are identified as barriers to undertaking health promotion interventions. Staff and patients have different priorities in terms of the health topics they feel should be addressed. Patients reported receiving books and leaflets as well as speaking with a knowledgeable person as their preferred health promotion approach. Staff often stated the need for more training. CONCLUSIONS: Few studies have investigated the barriers to health promotion interventions in urgent and emergency care settings and there is a lack of evidence about the acceptability of health promotion activity. Additional research is needed to determine whether extending the role of paramedics and emergency nurses to include health promotion interventions will be acceptable to staff and patients.

Maintaining pre-school children's health and wellbeing in the UK: a qualitative study of the views of migrant parents.

Background: There is evidence that key health behaviours of people who migrate deteriorate over time, which has a consequent impact upon the health of dependent children. As health in the early years sets the course for lifelong health, it is important to explore parents' views on maintaining children's health following migration. Methods: Five focus groups were held with parents of preschool children who had migrated to the UK within the last 10 years (n = 28). Parents originated from Romania, Poland, Somalia and Pakistan, with one group of Roma Gypsy parents. Data collection took place in January to March 2015. Results: All groups, apart from the Roma, perceived barriers to maintaining optimal health and well-being for their preschool children following migration to the UK. Eastern European parents experienced difficulties in ensuring family financial security, while parents from more established communities focused on barriers to children's exercise, play and nutrition. Conclusions: This study highlights aspects of public health where migrants and their children can experience adverse effects in the UK. These findings have implications for policymakers, commissioners and providers of health services who aim to promote good health among preschool children.

Polymeric prodrug combination to exploit the therapeutic potential of antimicrobial peptides against cancer cells.

Antimicrobial Peptides (AMPs) have unique anticancer properties, but their clinical application is currently limited by an inadequate margin of safety. A prodrug strategy associated with a combination therapy approach could address this limitation by increasing their therapeutic index and their efficacy. Accordingly, the first targeted anticancer polymeric prodrug candidates of AMPs, intended for combination therapy with another polymeric prodrug of an approved antineoplastic agent (doxorubicin), were synthesized as either a PEG-based dual-release prodrug or two individual pegylated prodrugs. The latter are based on a cathepsin B-labile peptide linker and an acid-sensitive acyl hydrazone bond for the AMP and doxorubicin prodrugs, respectively. Anticancer activities and toxicity differentials achieved with the free peptide and its polymer conjugates against ovarian, cancer and non-malignant, cells, indicate that protease-dependent reversible pegylation could be implemented to increase the therapeutic indices of AMPs in cancer therapy. The results obtained also show that this approach can be developed if the releasable PEG linker can be optimised to conciliate the attributes and restrictions of pegylation against proteases. In addition, combination of the polymeric prodrugs of the AMP and of doxorubicin provides additive antitumor effects which could be exploited to enhance the efficacy of the AMP candidate.

Investigation of the multifaceted iron acquisition strategies of Burkholderia cenocepacia.

Burkholderia cenocepacia is a bacterial pathogen which causes severe respiratory infections in cystic fibrosis (CF). These studies were aimed at gaining an insight into the iron acquisition strategies of B. cenocepacia. In iron restricted conditions, genes associated with the synthesis and utilisation of ornibactin (pvdA, orbA, orb F) were significantly upregulated compared to the expression of pyochelin associated genes (pchD, fptA). In the absence of alternative iron sources, B. cenocepacia J2315 and 715j utilised ferritin and haemin, but not transferrin or lactoferrin for growth. Significantly, mutants unable to produce ornibactin, (715j-orbI) or ornibactin and pyochelin, (715j-pobA), utilised haemin and ferritin more efficiently than the wild-type. Moreover, both mutants were also able to utilise lactoferrin for growth (P ≤ 0.01) and additionally 715j-pobA utilised transferrin (P ≤ 0.01), potentially facilitating adaptation to the host environment. Furthermore, B. cenocepacia increased ornibactin gene expression in response to pyoverdine from Pseudomonas aeruginosa (P ≤ 0.01), demonstrating the capacity to compete for iron in co-colonised niches. Pyoverdine also significantly diminished the growth of B. cenocepacia (P < 0.001) which was related to its iron chelating activity. In a study of three B. cenocepacia sequential clonal isolates obtained from a CF patient over a 3.5 year period, ornibactin upregulation in response to pyoverdine was less pronounced in the last isolate compared to the earlier isolates, as was growth in the presence of haemin and ferritin, indicating alternative iron acquisition mechanism(s) may dominate as chronic infection progresses. These data demonstrate the multifaceted iron acquisition strategies of B. cenocepacia and their capacity to be differentially activated in the presence of P. aeruginosa and during chronic infection.

Assessing the Efficacy of Active Learning to Support Student Performance Across Undergraduate Programmes in Biomedical Science.

Introduction: Active learning is a useful tool to enhance student engagement and support learning in diverse educational situations. We aimed to assess the efficacy of an active learning approach within a large interprofessional first year Medical Cell Biology module taken by six healthcare programmes across the School of Biomedical Sciences at Ulster University, United Kingdom. Materials and methods: An active learning approach was developed for weekly formative assessment using Smartwork to design a weekly interactive multiple-choice quiz to reinforce key concepts specifically for each lecture. We tracked and assessed student performance in the module overall and in each element of course work and exam for 2 years prior to and following the introduction of an active learning strategy to engage and support learning for students from all academic backgrounds and abilities. Results: Full engagement with active learning was significantly associated with an increased overall module performance as well as a significantly increased performance in each element of class test (No engagement vs. Full engagement, p < 0.001), exam (No Engagement vs. Full engagement, p < 0.05) and coursework (No engagement vs. Full engagement, p < 0.001) within this overall total (No Engagement vs. Full engagement, p < 0.01). Partial engagement with active learning was associated significantly improved class test (No engagement vs. partially engaged, p < 0.001) and coursework (No engagement vs. partially engaged, p < 0.05) performance. While a trend toward increased performance in exam and overall module mark was observed, these were not significant. Discussion: Active learning is a useful tool to support student learning across a range of healthcare programmes taken by students with differing backgrounds and academic abilities in an interprofessional and widening participation setting. Student engagement in active learning was highlighted as a key contributory factor to enhanced student performance in all aspects of assessment.

Peptides originally derived from Chilobrachys jingzhao tarantula venom possess beneficial effects on pancreatic beta cell health and function.

Clinical approval of the glucagon-like peptide-1 (GLP-1) mimetic exenatide for the treatment of type 2 diabetes highlights the therapeutic effectiveness of venom-derived peptides. In the present study, we examined and characterised the glucose-lowering potential of synthetic Jingzhaotoxin IX and Jingzhaotoxin XI peptides, which were originally isolated from the venom of the Chinese earth tarantula Chilobrachys jingzhao. Following confirmation of lack of beta-cell toxicity of synthetic peptides, assessment of enzymatic stability and effects on in vitro beta-cell function were studied, alongside putative mechanisms. Glucose homeostatic and appetite suppressive actions of Jingzhaotoxin IX and Jingzhaotoxin XI alone, or in combination with exenatide, were then assessed in normal overnight fasted C57BL/6 mice. Synthetic Jingzhaotoxin peptides were non-toxic and exhibited a decrease in mass of 6 Da in Krebs-Ringer bicarbonate buffer suggesting inhibitor cysteine knot (ICK)-like formation, but interestingly were liable to plasma enzyme degradation. The Jingzhaotoxin peptides evoked prominent insulin secretion from BRIN BD11 beta-cells, with activity somewhat characteristic of Kv2.1 channel binding. In addition, Jingzhaotoxin peptides enhanced beta-cell proliferation and provided significant protection against cytokine-induced apoptosis. When injected co-jointly with glucose, the Jingzhaotoxin peptides slightly decreased blood-glucose levels but had no effect on appetite in overnight fasted mice. Whilst the Jingzhaotoxin peptides did not enhance exenatide-induced benefits on glucose homeostasis, they augmented exenatide-mediated suppression of appetite. Taken together, these data highlight the therapeutic potential of tarantula venom-derived peptides, such as Jingzhaotoxin IX and Jingzhaotoxin XI either alone or in combination with exenatide, for diabetes and related obesity.

A novel peptide isolated from Aphonopelma chalcodes tarantula venom with benefits on pancreatic islet function and appetite control.

Proof-of-concept for therapeutic application of venom-derived compounds in diabetes is exemplified by the incretin mimetic, exenatide, originally extracted from the saliva of the venomous Heloderma suspectum lizard. In this regard, we have isolated and sequenced a novel 28 amino acid peptide named Δ-theraphotoxin-Ac1 (Δ-TRTX-AC1) from venom of the Mexican Blond tarantula spider Aphonopelma chalcodes, with potential therapeutic benefits for diabetes. Following confirmation of the structure and safety profile of the synthetic peptide, assessment of enzymatic stability and effects of Δ-TRTX-AC1 on in vitro beta-cell function were studied, alongside potential mechanisms. Glucose homeostatic and satiety actions of Δ-TRTX-AC1 alone, and in combination with exenatide, were then assessed in C57BL/6 mice. Synthetic Δ-TRTX-AC1 was shown to adopt a characteristic inhibitor cysteine knot (ICK)-like structure and was non-toxic to beta-cells. Δ-TRTX-AC1 evoked glucose-dependent insulin secretion from BRIN BD11 cells with bioactivity confirmed in murine islets. Insulin secretory potency was established to be dependent on KATP and Ca2+ channel beta-cell signalling. In addition, Δ-TRTX-AC1 enhanced beta-cell proliferation and provided significant protection against cytokine-induced apoptosis. When injected co-jointly with glucose in mice at a dose of 250 nmol/kg, Δ-TRTX-AC1 decreased blood-glucose levels and evoked a significant satiating effect. Moreover, whilst Δ-TRTX-AC1 did not enhance exenatide induced benefits on glucose homeostasis, the peptide significantly augmented exenatide mediated suppression of appetite. Together these data highlight the therapeutic potential of tarantula spider venom-derived peptides, such as Δ-TRTX-Ac1, for diabetes and related obesity.

Is sonographic intra-nodular vascularity a reliable predictor of thyroid malignancy? A UK tertiary centre study.

OBJECTIVE: British Thyroid Association 2014 guidelines emphasised ultrasound assessment of nodules. One ultrasonographic differentiator of debatable relevance is intra-nodular vascularity. This is the first UK study conducted to address this question. METHODS: Ultrasound reports for thyroid surgery patients over 10 years were retrospectively reviewed. Reports documenting 'intra-nodular vascularity or flow' were analysed. Reports identifying peripheral vascularity only or no intra-nodular flow formed the control group. Concordance with final histology was used to determine the odds ratio for malignancy. RESULTS: A total of 306 patients were included, and 119 (38.9 per cent) nodules demonstrated intra-nodular vascularity. Of these, 60 (50.4 per cent) were malignant compared with 42 per cent in the control group. Intra-nodular vascularity was not a statistically significant predictor of malignancy with an odds ratio of 1.39 (p = 0.18, 95 per cent confidence interval, 0.86-2.23). CONCLUSION: Intra-nodular vascularity in isolation was not a reliable predictor of malignancy. This supports other world literature studies. Although intra-nodular flow should not be relied upon in isolation, interpretation in conjunction with other suspicious findings enhances the predictive value.

The importance of postpartum glucose tolerance testing after pregnancies complicated by gestational diabetes.

AIMS: To review postpartum glucose tolerance in women with gestational diabetes and evaluate the role of formal 75 g oral glucose tolerance testing vs. fasting plasma glucose in screening for persistent abnormalities. METHODS: Retrospective study of 985 pregnancies over a 10 year period in a mixed ethnic cohort of women who underwent follow-up glucose tolerance testing at 6 weeks postpartum. Diagnosis obtained by oral glucose tolerance test was tested against that from the fasting plasma glucose value. RESULTS: There were 272 abnormal postpartum oral glucose tolerance test results (27.6%), with 109 women identified as having frank diabetes. Eleven of these (10%) had fasting plasma glucose < or =6.0 mmol/l, as did 62 of 114 cases of impaired glucose tolerance. A fasting plasma glucose concentration of > or =6.1 mmol/l correctly identified abnormal glucose tolerance in 199 of 272 cases (sensitivity 0.73). South Asian women were much more likely to have persistent abnormalities of glucose tolerance than were Europeans (32 vs. 15%, chi(2)P < 0.0001). CONCLUSIONS: A postpartum fasting plasma glucose measurement alone is not sensitive enough in our population to classify glucose tolerance status accurately. A formal postpartum oral glucose tolerance test is therefore needed to facilitate early detection and treatment.

Real-time PCR method for the quantification of Burkholderia cepacia complex attached to lung epithelial cells and inhibition of that attachment.

AIMS: To develop a rapid method to quantify the attachment of the cystic fibrosis pathogen, Burkholderia multivorans, to lung epithelial cells (16HBE14o(-)) using real-time PCR with a view to monitoring potential inhibition of lung cell attachment. METHODS AND RESULTS: Mammalian and bacterial DNA were purified from bacteria attached to lung epithelial cells. The relative amount of bacteria attached was determined by amplification of the recA gene relative to the human GAPDH gene, in the presence of SYBR Green. The method was thoroughly validated and shown to correlate well with traditional plating techniques. Inhibition of bacterial attachment with simple sugars was then evaluated by real-time PCR. Of the sugars examined, pre-incubation of B. multivorans with lactose, mannose and xylitol all decreased bacterial adherence to 16HBE14o(-) cells, while glucose and galactose had no significant effect. Pre-incubation with lactose had the greatest effect, resulting in reduced adhesion to 35% of untreated controls. CONCLUSIONS: This method can be used to quickly and effectively screen novel agents with higher affinities for bacterial adhesins. SIGNIFICANCE AND IMPACT OF THE STUDY: This method will enable the rapid development of novel agents to inhibit colonization by this pathogen from the environment.

Effects of an enzymatically stable C-terminal hexapseudopeptide fragment peptide of xenin-25, ψ-xenin-6, on pancreatic islet function and metabolism.

Xenin-25 undergoes rapid enzyme metabolism following secretion. Early studies demonstrated bioactivity of a C-terminal hexapeptide fragment of xenin-25, namely xenin-6, which were enhanced through introduction of a reduced N-terminal peptide bond, to yield Ψ-xenin-6. The present study was undertaken to define the biological actions and potential antidiabetic properties of Ψ-xenin-6. In vitro enzymatic stability, insulin and glucagon secretory activity, as well as effects on beta-cell survival were determined. Studies in mice were used to assess the impact of Ψ-xenin-6 on glucose homeostasis and satiety. Ψ-xenin-6 was resistant to murine plasma degradation. In BRIN-BD11 cells and isolated murine islets, Ψ-xenin-6 significantly stimulated insulin secretion, and prominently enhanced the insulinotropic actions of GIP. Xenin-6 and Ψ-xenin-6 had no impact on glucagon secretion, although xenin-6 partially reversed the glucagonotropic action of GIP. Further in vitro investigations revealed that, similar to GLP-1, Ψ-xenin-6 significantly augmented proliferation of human and rodent clonal beta-cells, whilst also fully protecting against cytokine-induced beta-cell cytotoxicity, with greater potency than xenin-25 and xenin-6. When administered to mice in combination with glucose, Ψ-xenin-6 significantly reduced glucose levels and enhanced glucose-induced insulin release, with a duration of biological action beyond 8 h. Ψ-xenin-6 also significantly enhanced the glucose-lowering action of GIP in vivo. In overnight fasted mice, Ψ-xenin-6 exhibited satiety actions at both 25 and 250 nmol/kg. These data demonstrates that Ψ-xenin-6 is a metabolically stable C-terminal fragment analogue of xenin-25, with a metabolic action profile that merits further study as a potential antidiabetic compound.

Novel dual incretin agonist peptide with antidiabetic and neuroprotective potential.

Glucose-dependent insulinotropic hormone (GIP) and glucagon-like peptide-1 (GLP-1) are incretin hormones that exert an array of beneficial actions on metabolism and cognitive function. GLP-1-based therapeutics have been highly successful in terms of obesity and diabetes management, however GIP therapies have found no clinical utility to date. In the present study we describe, for the first time, the therapeutic effectiveness of a novel GIP/GLP-1 hybrid peptide based on the amino acid sequences of GIP, GLP-1 and the clinically approved GLP-1 mimetic, exendin-4. The hybrid peptide, N-ac(d-Ala2)GIP/GLP-1-exe, was enzymatically stable for up to 12 h when incubated with DPP-4. N-ac(d-Ala2)GIP/GLP-1-exe significantly (P < 0.001) stimulated insulin secretion from BRIN-BD11 cells and isolated mouse islets, and evoked dose-dependent increases (P < 0.001) in cAMP production in both GIP-R and GLP-1-R transfected cells. In mice, injection of the hybrid in combination with glucose significantly (P < 0.001) reduced glucose and increased insulin concentrations, with metabolic actions evident (P < 0.05) 8 h post-injection. Twice-daily injection of N-ac(d-Ala2)GIP/GLP-1-exe to high fat fed (HFF) mice for 28 days significantly (P < 0.05-P < 0.001) reduced body weight, HbA1c, circulating glucose and insulin concentrations. Furthermore, both oral and i.p. glucose tolerance were improved (P < 0.001) and insulin sensitivity enhanced. The hybrid peptide also increased (P < 0.05-P < 0.001) beta cell number, islet area, pancreatic insulin content and islet insulin secretory responsiveness in HFF mice. Finally, N-ac(d-Ala2)GIP/GLP-1-exe treated mice exhibited improved (P < 0.01) recognition memory which was accompanied by enhanced (P < 0.05-P < 0.001) hippocampal neurogenesis, synapse formation and reduced neuronal oxidative stress. These data demonstrate for the first time the beneficial actions of the novel GIP/GLP-1 hybrid, N-ac(d-Ala2)GIP/GLP-1-exe, on glucose homeostasis and memory function in diabetes.

Dominant expression of multiple drug resistance after in vitro X-irradiation exposure in intraspecific Chinese hamster ovary hybrid cells.

BACKGROUND: Exposure of Chinese hamster ovary (CHO) cells to fractionated x irradiation in vitro has led to expression of a distinctive multiple-drug-resistant phenotype. This phenotype is characterized by overexpression of P-glycoprotein without an increase in P-glycoprotein messenger RNA or gene amplification; increased glutathione S-transferase (GST) activity; and resistance to vincristine, colchicine, and etoposide but not to doxorubicin. PURPOSE: To investigate whether this phenotype is dominant or recessive, we established intraspecific hybrids by fusion of x-ray-treated, drug-resistant CHO cells (DXR-10I or DXR-10II) with drug-sensitive CHO cells (E29). METHODS: Drug resistance levels were determined in the wild-type CHO cell line AuxB1, the drug-sensitive E29 line, the x-ray-pretreated lines, and the hybrid lines by colony-forming assay of cells grown in increasing concentrations of colchicine, vincristine, or doxorubicin. The hybrids were characterized by analysis of DNA content, P-glycoprotein expression by Western blotting, GST activity by use of 1-chloro-2,4-dinitrobenzene as substrate, and sensitivity to reversal of resistance to vincristine by exposure to verapamil. RESULTS: These hybrids proved resistant to colchicine (two-fold) and vincristine (five- to seven-fold) but not to doxorubicin. After the hybrids were exposed to verapamil, vincristine cytotoxicity was increased 10- to 12-fold. The hybrid lines exhibited levels of P-glycoprotein comparable to those of the unfused x-ray-treated parent cell line, suggesting that P-glycoprotein overexpression is a dominant trait in these hybrid lines. Interpretation of the role of increased GST activity in these hybrids was inconclusive because of the very high levels of GST in the drug-sensitive cell-fusion partner. CONCLUSIONS: The multiple-drug-resistant phenotype following x-ray treatment of CHO cells in vitro was dominantly expressed. Overall, these data are consistent with the hypothesis that this phenotype is a consequence of the dominant genetic alteration resulting from exposure to x irradiation. IMPLICATIONS: This work adds weight to our hypothesis that there is a biological basis for the expression of clinical drug resistance in certain patients whose tumors have been previously irradiated.

Characterization of the P-glycoprotein over-expressing drug resistance phenotype exhibited by Chinese hamster ovary cells following their in-vitro exposure to fractionated X-irradiation.

Exposure of the Chinese hamster ovarian AuxB1 cell line in vitro to fractionated X-irradiation generated sublines designated DXR-10, which proved resistant to multiple drugs and overexpressed P-glycoprotein (Pgp), as judged by Western blotting using the C219 monoclonal antibody. Further characterization of these irradiated DXR-10 sublines has provided evidence for: (i) the expression of cross-resistance to gramacidin D, taxol, puromycin and Navelbine, but not to daunomycin or mitoxantrone; (ii) overexpression of the class I Pgp, as judged by Western blotting using the C494 monoclonal antibody; (iii) decreased accumulation of 3H-vincristine, which could be enhanced by verapamil addition; (iv) unaltered accumulation and subcellular distribution of adriamycin; (v) significantly increased rhodamine 123 accumulation in the presence of verapamil; (vi) plasma-membrane ultrastructural modifications resulting in a significantly increased surface area; (vii) numerous clonal karyotypic alterations, with abnormalities involving the long arm of chromosome 1 being consistently identified; (viii) a lack of overexpression of sorcin; (ix) increased total glutathione levels and overexpression of glutathione S-transferase pi. The fact that only certain of these features are considered characteristic of the 'classic' multidrug-resistant CHRC5 cell line supports our earlier proposal that exposure to fractionated X-irradiation results in the expression of a unique drug-resistance phenotype.

Markov chain modelling for geriatric patient care.

OBJECTIVES: To show that Markov chain modelling can be applied to data on geriatric patients and use these models to assess the effects of covariates. METHODS: Phase-type distributions were fitted by maximum likelihood to data on times spent by the patients in hospital and in community-based care. Data on the different events that ended the patients' periods of care were used to estimate the dependence of the probabilities of these events on the phase from which the time in care ended. The age of the patients at admission to care and the year of admission were also included as covariates. RESULTS: Differential effects of these covariates were shown on the various parameters of the fitted model, and interpretations of these effects made. CONCLUSIONS: Models based on phase-type distributions were appropriate for describing times spent in care, as the ordered phases had an interpretable structure corresponding to increasing amounts of care being given.

Etretinate: effect of milk intake on absorption.

Since etretinate, an aromatic retinoid useful in the treatment of psoriasis and other skin disorders is lipid-soluble, it may be poorly absorbed in the absence of a fat load. We therefore studied serum concentrations of etretinate and its major metabolite (Ro 10-1670) after the controlled administration of etretinate. After an overnight fast, 6 Darier's disease and 4 psoriatic patients received a 1 mg/kg morning dose of etretinate with water or 1 pint of whole milk (fat load). There was a 260% increase (p less than 0.0005) in the mean of each patient's increase in the baseline-corrected peak serum concentration of etretinate after administration with milk (115 +/- 15 micrograms/dl) compared to after administration with water (32 +/- 4 micrograms/dl). Over a 24-h period there was an overall 296 +/- 26% (p less than 0.0005) increase in serum etretinate after administration with milk compared to water in 5 patients with Darier's disease. In contrast to the serum etretinate, there was a 17% mean decrease (p less than 0.025) in the corrected peak serum concentration of Ro 10-1670 in all 10 patients after administration of etretinate with milk compared to water. The net result of these alterations is that the mean corrected serum concentration of etretinate is higher than Ro 10-1670 at all time points measured after milk administration. In contrast, after administration of etretinate with water the major retinoid in the serum is Ro 10-1670. Establishing the clinical significance of these alterations may require controlled clinical trials.

Evidence of post-translational regulation of P-glycoprotein associated with the expression of a distinctive multiple drug-resistant phenotype in Chinese hamster ovary cells.

Chinese hamster ovary (CHO) cells following exposure to fractionated X-irradiation in vitro dominantly expressed a distinctive multiple drug-resistant phenotype, characterised by resistance to vinca alkaloids, epipodophyllotoxins and colchicine, but not to anthracyclines, together with overexpression of P-glycoprotein (Pgp), but without any concomitant elevation in Pgp mRNA (J Natl Cancer Inst 1990, 82, 607-612; 1992, 85, 48-53). To investigate the mechanism of this Pgp overexpression, Pgp stability ws examined in an X-irradiation pretreated subline and compared with that of two colchicine-selected drug-resistant CHO sublines. These studies revealed a slower turnover of Pgp in the X-irradiated cells (T1/2 > or = 40 h) relative to the drug-selected sublines (T1/2 = 17 h), indicating that Pgp overexpression appears to be differently regulated in these independently-derived resistant sublines. These data add support to our proposal that the development of drug resistance following X-irradiation may arise by a mechanism distinct from that operating after drug selection.

Brevinin-1 and multiple insulin-releasing peptides in the skin of the frog Rana palustris.

Few studies have comprehensively examined amphibian granular gland secretions for novel insulinotropic peptides. This study involved isolation and characterisation of biologically active peptides from the skin secretions of Rana palustris frogs. Crude secretions obtained by mild electrical stimulation from the dorsal skin surface were purified by reversed-phase HPLC on a semipreparative Vydac C18 column, yielding 80 fractions. These fractions were assayed for insulin-releasing activity using glucose-responsive BRIN-BD11 cells. Acute 20 min incubations were performed in Krebs Ringer bicarbonate buffer supplemented with 5.6 mmol/l glucose in the absence (control) and presence of various fractions. Fractions 29-54 and fractions 68-75 showed significant 2.0-6.5-fold increases in insulin-releasing activity (P<0.001). The fractions showing most prominent insulinotropic activity were further purified to single homogeneous peaks, which, on testing, evoked 1.5-2.8-fold increases in insulin release (P<0.001). The structures of the purified peptides were determined by mass spectrometry and N-terminal amino acid sequencing. Electrospray ionisation ion-trap mass spectrometry analysis revealed molecular masses of 2873.5-8560.4 Da. Sufficient material was isolated to determine the primary amino acid sequence of the 2873.5 Da peptide, revealing a 27 amino acid sequence, ALSILRGLEKLAKMGIALTNCKATKKC, repressing palustrin-1c. The database search for this peptide showed a 48% homology with brevinin-1, an antimicrobial peptide isolated from various Rana species, which itself stimulated insulin release from BRIN-BD11 cells in a concentration-dependent manner. In conclusion, the skin secretions of R. palustris frogs contain a novel class of peptides with insulin-releasing activity that merit further investigation.

Isolation and characterisation of an unexpected class of insulinotropic peptides in the skin of the frog Agalychnis litodryas.

Skin secretions of the frog Agalychnis litodryas were evaluated for the isolation and characterisation of novel insulinotropic peptides. Crude secretions obtained from young adult frogs by mild electrical stimulation of the dorsal skin surface were purified by reverse-phase high-performance liquid chromatography (HPLC) yielding 70 fractions. In acute 20-min incubations with glucose responsive BRIN-BD11 cells, fractions 39-42 (band 1) and fractions 44-46 (band 2) significantly stimulated insulin release by 2-3.5-fold compared with 5.6 mM glucose alone. Pooled fractions in band 1 and band 2 were rechromatographed to reveal 20 homogenous peptide peaks, which elicited significant 1.5-4-fold increases in insulin release. Mass spectrometry analyses indicated molecular masses of between 1649.2 and 4988.9 Da. The two peptides with the greatest insulin-releasing activity were directly subjected to N-terminal amino acid sequence analysis. The sequence of the 3020 Da peptide, called frog skin insulinotropic peptide or FSIP, was determined as AVWKDFLKNIGKAAGKAVLNSVTDMVNE, which has 79% homology with the C-terminal of the 75 amino acid dermaseptin BIV precursor. A partial N-terminal sequence was determined for the 2546.2 Da peptide as MLADVFEKIMGD... These data indicate that the skin secretions of A. litodryas frogs contain biologically active peptides which merit further evaluation as a new class of insulin secretagogues.