Annona muricata L. peel extract inhibits carbohydrate metabolizing enzymes and reduces pancreatic ß-cells, inflammation, and apoptosis via upregulation of PI3K/AKT genes.

BACKGROUND AND OBJECTIVE: Annona muricata L. peel has been recognized for many ethnobotanical uses, including diabetes management. However, limited detailed scientific information about its mechanism of antidiabetic activity exists. The objective of this study was to evaluate the anti-diabetic properties of an aqueous extract of A. muricata peel (AEAMP) and its mechanism of action on alloxan-induced diabetic rats. METHODS: In vitro antidiabetic assays, such as α-amylase and α-glucosidase were analyzed on AEAMP. Alloxan monohydrate (150 mg/kg b.w) was used to induce diabetes in the rats. 150 mg/kg b.w positive control group doses of 6.67, 13.53, and 27.06 mg/kg were administered to 3 groups for twenty-one days. The positive control group was administered 30 mg/kg of metformin. The negative and normal control groups were administered distilled water. The fasting blood glucose, serum insulin, lipid profile, inflammatory cytokines, antioxidant markers, carbohydrate metabolizing enzymes, and liver glycogen were analyzed as well as PI3K/AKT and apoptotic markers PCNA and Bcl2 by RT-PCR. RESULTS: AEAMP inhibited α-amylase and α-glucosidase enzymes more effectively than acarbose. AEAMP reduced FBG levels, HOMA-IR, G6P, F-1,6-BP, MDA, TG, TC, AI, CRI, IL-6, TNF-α, and NF-κB in diabetic rats. Furthermore, in diabetic rats, AEAMP improved serum insulin levels, HOMA-ß, hexokinase, CAT, GST, and HDL-c. Liver PI3K, liver PCNA and pancreas PCNA were not significantly different in untreated diabetic rats when compared to normal rats suggesting alloxan induction of diabetes did not downregulate the mRNA expression of these genes. AEAMP significantly up-regulated expression of AKT and Bcl2 in the liver and pancreatic tissue. It is interesting that luteolin and resorcinol were among the constituents of AEAMP. CONCLUSIONS: AEAMP can improve ß-cell dysfunction by upregulating liver AKT and pancreatic PI3K and AKT genes, inhibiting carbohydrate metabolizing enzymes and preventing apoptosis by upregulating liver and pancreatic Bcl2. However, the potential limitation of this study is the unavailability of equipment and techniques for collecting more data for the study.

Antidiabetic Activity of Elephant Grass (Cenchrus Purpureus (Schumach.) Morrone) via Activation of PI3K/AkT Signaling Pathway, Oxidative Stress Inhibition, and Apoptosis in Wistar Rats.

Ethnopharmacological Relevance: The management of diabetes over the years has involved the use of herbal plants, which are now attracting interest. We assessed the antidiabetic properties of aqueous extract of C. purpureus shoots (AECPS) and the mechanism of action on pancreatic ß-cell dysfunction. Methods: This study was conducted using Thirty-six 36) male Wistar rats. The animals were divided into six equal groups (n = 6) and treatment was performed over 14 days. To induce diabetes in the rats, a single dose of 65 mg/kg body weight of alloxan was administered intraperitoneal along with 5% glucose. HPLC analysis was carried out to identified potential compounds in the extract. In vitro tests α-amylase, and α-glucosidase were analyzed. Body weight and fasting blood glucose (FBG) were measured. Biochemical parameters, such as serum insulin, liver glycogen, hexokinase, glucose-6-phosphate (G6P), fructose-1,6-bisphosphatase (F-1,6-BP), interleukin-6 (IL-6), tumor necrosis factor-alpha (TNF-α), and nuclear factor kappa B (NF-ĸB), were analyzed. Additionally, mRNA expressions of phosphatidylinositol 3-kinase/protein kinase B (PI3K/AKT), B-cell lymphoma 2 (Bcl-2), and proliferating cell nuclear antigen (PCNA) were each evaluated. Results: This in vitro study showed inhibitory potency of Cenchrus purpureus extract (AECPS) as compared with the positive controls. AECPS showed a gradual decrease in alloxan-induced increases in FBG, total cholesterol (TC), triglycerides (TG), low density lipoprotein (LDL-c), G6P, F-1,6-BP, malondialdehyde (MDA), IL-6, TNF-α, and NF-ĸB and increased alloxan-induced decreases in liver glycogen, hexokinase, and high density lipoprotein (HDL-c). The diabetic control group exhibited pancreatic dysfunction as evidenced by the reduction in serum insulin, homeostasis model assessment of ß-cell function (HOMA-ß), expressions of PI3K/AKT, Bcl-2, and PCNA combined with an elevation in homeostatic model assessment of insulin resistance (HOMA-IR). High performance liquid chromatography (HPLC) revealed 3-O-rutinoside, ellagic acid, catechin, rutin, and kaempferol in AECPS. Conclusion: AECPS showed efficient ameliorative actions against alloxan-induced pancreatic dysfunction, oxidative stress suppression as well as, inflammation, and apoptosis via the activation of PI3K/AKT signaling pathways.

Antidiabetic activity of avocado seeds (Persea americana Mill.) in diabetic rats via activation of PI3K/AKT signaling pathway.

The treatment of diabetes involves the use of herbal plants, attracting interest in their cost-effectiveness and efficacy. An aqueous extract of Persea americana seeds (AEPAS) was explored in this study as a possible therapeutic agent in rats with diabetes mellitus. The induction of diabetes in the rats was achieved by injecting 65 mg/kg body weight (BWt) of alloxan along with 5% glucose. This study was conducted using thirty-six (36) male Wistar rats. The animals were divided into 6 equal groups, (n = 6) and treated for 14 days. In vitro assays for total flavonoid, phenols, FRAP, DPPH, NO, α-amylase, and α-glucosidase, were performed. Biochemical indices fasting blood sugar (FBS), BWt, serum insulin, liver hexokinase, G6P, FBP, liver glycogen, IL-6, TNF-α, and NF-ĸB in the serum, were investigated as well as the mRNA expressions of PCNA, Bcl2, PI3K/Akt in the liver and pancreas. The in vitro analyses showed the potency of AEPAS against free radicals and its enzyme inhibitory potential as compared with the positive controls. AEPAS showed a marked decrease in alloxan-induced increases in FBG, TG, LDL-c, G6P, F-1, 6-BP, MDA, IL-6, TNF-α, and NF-ĸB and increased alloxan-induced decreases in liver glycogen, hexokinase, and HDL-c. The diabetic control group exhibited pancreatic dysfunction as evidenced by a reduction in serum insulin, HOMA-ß, expressions of PI3K/AKT, Bcl-2, and PCNA combined with an elevation in HOMA-IR. The HPLC revealed luteolin and myricetin to be the phytochemicals that were present in the highest concentration in AEPAS. The outcome of this research showed that the administration of AEPAS can promote the activation of the PI3K/AkT pathway and the inhibition of ß-cell death, which may be the primary mechanism by which AEPAS promotes insulin sensitivity and regulates glycolipid metabolism.

ABIN1 protein cooperates with TAX1BP1 and A20 proteins to inhibit antiviral signaling.

Upon virus infection, the innate immune response provides the first line of protection and rapidly induces type I interferons (IFNα/ß), which mediate potent antiviral effects. To maintain homeostasis and prevent autoimmunity, IFN production is tightly regulated; however, the mechanisms of negative regulation are poorly understood. Herein, we demonstrate that the A20 binding inhibitor of NF-κB 1 (ABIN1) is a novel negative regulator of antiviral signaling. Overexpression of ABIN1 inhibited IFN-ß promoter activation in response to virus infection or poly(I:C) transfection, whereas siRNA-mediated knockdown of ABIN1 enhanced IFN-ß production upon virus infection. ABIN1 interacted with the A20 regulatory molecule TAX1BP1 and was essential for the recruitment of TAX1BP1 and A20 to the noncanonical IκB kinases TBK1 and IKKi in response to poly(I:C) transfection. ABIN1 and TAX1BP1 together disrupted the interactions between the E3 ubiquitin ligase TRAF3 and TBK1/IKKi to attenuate lysine 63-linked polyubiquitination of TBK1/IKKi. Finally, an intact ubiquitin binding domain of ABIN1 was essential for ABIN1 to interact with TBK1/IKKi and inhibit IFN-ß production upon poly(I:C) transfection or virus infection. Together, these results suggest that ABIN1 requires its ubiquitin binding domain and cooperates with TAX1BP1 and A20 to restrict antiviral signaling.

Family physician attitudes about prescribing using a drug formulary.

BACKGROUND: Drug formularies have been created by third party payers to control prescription drug usage and manage costs. Physicians try to provide the best care for their patients. This research examines family physicians' attitudes regarding prescription reimbursement criteria, prescribing and advocacy for patients experiencing reimbursement barriers. METHODS: Focus groups were used to collect qualitative data on family physicians' prescribing decisions related to drug reimbursement guidelines. Forty-eight family physicians from four Ontario cities participated. Ethics approval for this study was received from the Hamilton Health Sciences/Faculty of Health Sciences Research Ethics Board at McMaster University. Four clinical scenarios were used to situate and initiate focus group discussions about prescribing decisions. Open-ended questions were used to probe physicians' experiences and attitudes and responses were audio recorded. NVivo software was used to assist in data analysis. RESULTS: Most physicians reported that drug reimbursement guidelines complicated their prescribing process and can require lengthy interpretation and advocacy for patients who require medication that is subject to reimbursement restrictions. CONCLUSION: Physicians do not generally see their role as being cost-containment monitors and observed that cumbersome reimbursement guidelines influence medication choice beyond the clinical needs of the patient, and produce unequal access to medication. They observed that frustration, discouragement, fatigue, and lack of appreciation can often contribute to family physicians' failure to advocate more for patients. Physicians argue cumbersome reimbursement regulations contribute to lower quality care and misuse of physicians' time increasing overall health care costs by adding unnecessary visits to family physicians, specialists, and emergency rooms.

TPL-2 negatively regulates interferon-beta production in macrophages and myeloid dendritic cells.

Stimulation of Toll-like receptors (TLRs) on macrophages and dendritic cells (DCs) by pathogen-derived products induces the production of cytokines, which play an important role in immune responses. Here, we investigated the role of the TPL-2 signaling pathway in TLR induction of interferon-beta (IFN-beta) and interleukin-10 (IL-10) in these cell types. It has previously been suggested that IFN-beta and IL-10 are coordinately regulated after TLR stimulation. However, in the absence of TPL-2 signaling, lipopolysaccharide (TLR4) and CpG (TLR9) stimulation resulted in increased production of IFN-beta while decreasing IL-10 production by both macrophages and myeloid DCs. In contrast, CpG induction of both IFN-alpha and IFN-beta by plasmacytoid DCs was decreased in the absence of TPL-2, although extracellular signal-regulated kinase (ERK) activation was blocked. Extracellular signal-related kinase-dependent negative regulation of IFN-beta in macrophages was IL-10-independent, required protein synthesis, and was recapitulated in TPL-2-deficient myeloid DCs by retroviral transduction of the ERK-dependent transcription factor c-fos.

Identification of eukaryotic open reading frames in metagenomic cDNA libraries made from environmental samples.

Here we describe the application of metagenomic technologies to construct cDNA libraries from RNA isolated from environmental samples. RNAlater (Ambion) was shown to stabilize RNA in environmental samples for periods of at least 3 months at -20 degrees C. Protocols for library construction were established on total RNA extracted from Acanthamoeba polyphaga trophozoites. The methodology was then used on algal mats from geothermal hot springs in Tengchong county, Yunnan Province, People's Republic of China, and activated sludge from a sewage treatment plant in Leicestershire, United Kingdom. The Tenchong libraries were dominated by RNA from prokaryotes, reflecting the mainly prokaryote microbial composition. The majority of these clones resulted from rRNA; only a few appeared to be derived from mRNA. In contrast, many clones from the activated sludge library had significant similarity to eukaryote mRNA-encoded protein sequences. A library was also made using polyadenylated RNA isolated from total RNA from activated sludge; many more clones in this library were related to eukaryotic mRNA sequences and proteins. Open reading frames (ORFs) up to 378 amino acids in size could be identified. Some resembled known proteins over their full length, e.g., 36% match to cystatin, 49% match to ribosomal protein L32, 63% match to ribosomal protein S16, 70% to CPC2 protein. The methodology described here permits the polyadenylated transcriptome to be isolated from environmental samples with no knowledge of the identity of the microorganisms in the sample or the necessity to culture them. It has many uses, including the identification of novel eukaryotic ORFs encoding proteins and enzymes.

Narrativizing errors of care: critical incident reporting in clinical practice.

This paper considers the rise across acute care settings in the industrialized world of techniques that encourage clinicians to record their experiences about adverse events they are personally involved in; that is, to share narratives about errors, mishaps or 'critical incidents'. The paper proposes that critical incident reporting and the 'root cause' investigations it affords, are both central to the effort to involve clinicians in managing and organizing their work, and a departure from established methods and approaches to achieve clinicians' involvement in these non-clinical domains of health care. We argue that critical incident narratives render visible details of the clinical work that have thus far only been discussed in closed, paperless meetings, and that, as narratives, they incite individuals to share personal experiences with parties previously excluded from knowledge about failure. Drawing on a study of 124 medical retrieval incident reports, the paper provides illustrations and interpretations of both the narrative and the meta-discursive dimensions of critical incident reporting. We suggest that, as a new and complex genre, critical incident reporting achieves three important objectives. First, it provides clinicians with a channel for dealing with incidents in a way that brings problems to light in a non-blaming way and that might therefore be morally satisfying and perhaps even therapeutic. Second, these narrations make available new spaces for the apprehension, identification and performance of self. Here, the incident report becomes a space where clinicians publicly perform concern about what happened. Third, incident reporting becomes the basis for radically altering the clinician-organization relationship. As a complex expression of clinical failure and its re-articulation into organizational meta-discourse, incident reporting puts doctors' selves and feelings at risk not just within the relative safety of personal or intra-professional relationships, but also in the normative context of organizational coordination, accountability, planning and management.

Bone mineral density after spinal cord injury: a reliable method for knee measurement.

OBJECTIVES: To test the interrater reliability of a standardized method to analyze knee bone mineral density (BMD) using dual-energy x-ray absorptiometry (DXA); to compare spine, hip, and knee BMD of people with spinal cord injury (SCI) with able-bodied controls; and to determine the relation between hip BMD and knee BMD in SCI and able-bodied subjects. DESIGN: Criterion standard and masked comparison. SETTING: Primary care university hospital. PARTICIPANTS: A convenience sample of 11 subjects with complete SCI was age and sex matched with 11 able-bodied control subjects. INTERVENTIONS: Not applicable. MAIN OUTCOME MEASURES: Four raters analyzed regions of interest according to operational definitions recently developed to standardize the analysis of BMD of the knee. Subjects with chronic SCI and matched controls underwent conventional DXA scans of the spine and hips and "less conventional" scans of the distal femurs and proximal tibias. The relation between hip and knee BMD was analyzed. RESULTS: The knee measurements were highly reliable (femur intraclass correlation coefficient model 2,1 [ICC(2,1)]=.98; tibia ICC(2,1)=.89). Subjects with SCI had lower BMD values than controls at all hip and knee sites (P<.05). Lumbar spine BMD did not differ between groups. Hip BMD was moderately predictive of distal femur BMD (R2=.67), but less correlated with the proximal tibia (R2=.38). CONCLUSIONS: Knee BMD can be reliably analyzed using DXA with this protocol. Subjects with SCI have diminished knee and hip BMD. Low hip BMD is associated with low distal femur BMD.

Glycoprotein 130 ligand oncostatin-M induces expression of vascular endothelial growth factor in human adult cardiac myocytes.

OBJECTIVE: In murine and rat cardiac myocytes the gp130 system transduces survival as well as hypertrophic signals and via induction of the expression of the potent angiogenic factor VEGF in these cells also indirectly contributes to cardiac repair processes through the development of new blood vessels. There are, however, species differences in receptor specificity and receptor crossreactivity in the gp130-gp130 ligand system. We asked whether gp130 signaling is also involved in the regulation of VEGF in human cardiac myocytes and if so which gp130 ligands are critical for such an effect. METHODS: Human adult cardiac myocytes (HACMs) were isolated from myocardial tissue and characterised by positive staining for myocardial actin, troponin-I and cardiotin. HACMs were treated with the gp130 ligands CT-1, IL-6, LIF or OSM and VEGF-1 was determined by a specific ELISA in the conditioned media of these cells. RT-PCR and Western blot analysis was used in order to detect gp130, IL-6-receptor, LIF-receptor or OSM-receptor specific protein and mRNA in human adult cardiac myocytes and for detection of VEGF-1 specific mRNA in cardiac myocytes after incubation with OSM. Pieces of myocardial tissue were incubated ex vivo in the presence and absence of OSM and VEGF was determined in supernatants of these cultures and immunohistochemistry was performed on the tissue using specific antibodies for VEGF-1. Immunohistochemistry was also employed to detect VEGF in sections from a healthy human heart and in a heart from a patient suffering from acute myocarditis. RESULTS: OSM, but not CT-1, IL-6 or LIF increased VEGF-1 production in human adult cardiac myocytes dose-dependently derived from five different donors. This selective stimulation of VEGF by gp130 ligands was also reflected by a specific receptor expression on these cells. We detected high levels of mRNA for gp130 and the OSM receptor in freshly isolated human cardiac myocytes but only low amounts of mRNA for the IL-6 receptor whereas mRNA for the LIF receptor was hardly detectable by RT-PCR. OSM receptor and IL-6 receptor were also detectable by Western blotting whereas LIF receptor was only present as a faint band. OSM also increased the expression of VEGF-1 mRNA in cardiac myocytes. When pieces of human myocardial tissue were incubated with the gp130 ligands in an ex vivo model only OSM resulted in an increase in VEGF-1 in the supernatants of these cultures. Furthermore, VEGF increased in tissue samples treated with OSM in cardiac myocytes as evidenced by immunohistochemistry. In addition, we found increased VEGF-1 expression in myocardial tissue from a patient suffering from acute myocarditis. CONCLUSION: The gp130-gp130 ligand system is also involved in VEGF regulation in human cardiac myocytes and OSM is the gp130 ligand responsible for this effect in the human system whereas LIF and CT-1 which had been shown to regulate VEGF expression in mouse and rat cardiac myocytes had no effect. Thus we have added OSM, which is produced by activated T lymphocytes and monocytes, to the list of regulatory molecules of VEGF production in the human heart. Our results lend further support to the notion that besides hypoxia, inflammation via induction of VEGF through autocrine or paracrine pathways plays a key role in (re)vascularisation of the myocardium.

Making patient-centered care come alive: achieving full integration of the patient's perspective.

A patient-centered model of care has profound implications for the way that care is planned, delivered, and evaluated. Although most leaders in healthcare organizations today embrace the basic tenets of a patient-centered philosophy, they often find that moving toward a patient-centered model requires an unanticipated level of commitment and significant adjustments in organizational structures. In this article, the authors describe how patients and families have been integrated into the care delivery model by involving them in planning, decision-making, and improvement processes at all levels of the organization.

An unexpected biochemical and functional interaction between gp130 and the EGF receptor family in breast cancer cells.

Oncostatin M (OSM), an interleukin-6 type cytokine, acts via the gp130 signaling receptor to inhibit proliferation and induce differentiation of breast cancer cells. EGF, a mitogen for breast cells, signals via EGFR/ErbB tyrosine kinase receptors which are implicated in breast cancer pathogenesis. Here we show paradoxically that EGF enhanced the OSM-induced inhibition of proliferation and induction of cellular differentiation in both estrogen receptor positive and negative breast cancer cells. This functional synergism was also seen with heregulin but not SCF, PDGF or IGF-1, indicating that it was specific to EGF-related growth factors. Immunoprecipitation experiments revealed that gp130 was constitutively associated with ErbB-2 and ErbB-3. There was a similar association between the OSMRbeta and ErbB-2. Furthermore, EGF unexpectedly induced tyrosine phosphorylation of gp130. We show that OSM induced phosphorylation of STAT3. Both OSM and EGF activated the p42/44 MAP kinases, but while the MEK inhibitor, PD98059, ablated the OSM-induced inhibition, it only partially ablated the inhibitory effects of OSM plus EGF. Thus, we have demonstrated that the receptors and signalling pathways of two apparently unrelated growth factors were intimately linked, resulting in an unexpected biological effect. This provides a new mechanism for generating signalling diversity and has potential clinical implications in breast cancer.

Recombinant Granulocyte-Macrophage Colony-Stimulating Factor (rGM-CSF) : A Review of its Pharmacological Properties and Prospective Role in the Management of Myelosuppression.

Recombinant granulocyte-macrophage colony-stimulating factor (rGM-CSF) is a polypeptide hormone produced through recombinant DNA technologies in glycosylated (yeast or mammalian expression systems) or nonglycosylated (Escherichia coli expression system) form. It is a multilineage haematopoietin which stimulates proliferation and differentiation of bone marrow myeloid progenitors and increases peripheral white blood cell counts when administered systemically. Treatment is generally well tolerated, although mild to moderate flu-like symptoms are common and rGM-CSF-induced fever and fluid retention may be problematic in occasional patients. rGM-CSF accelerates recovery of peripheral neutrophil counts after bone marrow transplantation, and results of a placebo-controlled randomised trial correlate this with reduced infectious episodes and shortened length of hospitalisation in patients with lymphoid malignancies. A substantial number of patients with graft failure after bone marrow transplantation also respond to rGM-CSF. The duration of myelosuppression secondary to cancer chemotherapy can be significantly reduced by rGM-CSF which has permitted investigation of antineoplastic dose-intensity escalation. In some haematopoietic disorders (e.g. aplastic anaemia, myelodysplasia and neutropenia secondary to HIV infection and antiviral therapy), rGM-CSF produces clinically useful increases in peripheral blood granulocyte counts, although the effect is generally not sustained after drug withdrawal. The potential for rGM-CSF to stimulate proliferation of the abnormal clone in myelodysplasia and in acute myelogenous leukaemia following induction therapy is of concern. Available data suggest, however, that with appropriate monitoring and exclusion of high-risk patients this serious potential risk can be avoided, and that myelopoiesis is enhanced in such patients by rGM-CSF treatment. Recombinant colony-stimulating factors are a new therapeutic modality; hence many aspects of their use remain to be clarified. Nonetheless, as one of a small group of novel agents rGM-CSF has major potential in the management of myelosuppression secondary to cytoreductive therapy with or without bone marrow transplantation, and in amelioration of disturbed myelopoiesis. It represents an important application of biotechnology to a difficult area of therapeutics.