The use of proteomics in identifying differentially expressed serum proteins in humans with type 2 diabetes

BACKGROUND: The aim of the study was to optimize protocols for finding and identifying serum proteins that are differentially expressed in persons with normal glucose tolerance (NGT) compared to individuals with type 2 diabetes mellitus (T2DM). Serum from persons with NGT and persons with T2DM was profiled using ProteinChip arrays and time-of-flight mass spectra were generated by surface enhanced laser desorption/ionization time-of-flight mass spectrometry (SELDI-TOF MS). RESULTS: Mass spectra from NGT- and T2DM-groups were compared. Fifteen proteins ranging from 5 to 79 kDa were differentially expressed (p < 0.05). Five of these proteins showed decreased and ten showed increased serum levels in individuals with T2DM. To be able to identify the proteins, the complexity of the sample was reduced by fractionation approaches. Subsequently, the purified fractions containing biomarkers were separated by one-dimensional SDS-polyacrylamide gel electrophoresis (SDS-PAGE) in two identical lanes. Protein bands of the first lane were excised and subjected to passive elution to recapture the biomarkers on ProteinChip arrays. The corresponding bands of the second lane were subjected to peptide-mass fingerprinting (PMF). Using this approach four of the differentially expressed proteins were identified as apolipoprotein C3 (9.4 kDa), transthyretin (13.9 kDa), albumin (66 kDa) and transferrin (79 kDa). Whereas apolipoprotein C3 and transthyretin were up-regulated, albumin and transferrin were down-regulated in T2DM. CONCLUSION: Protocols for protein profiling by SELDI-TOF MS and protein identification by fractionation, SDS-PAGE and PMF were optimized for serum from humans with T2DM. With these protocols differentially expressed proteins were discovered and identified when serum from NGT- and T2DM-individuals was analyzed

Therapeutic drug monitoring in inflammatory bowel disease : implementation, utilization, and barriers in clinical practice in Scandinavia

Background and aims Therapeutic drug monitoring (TDM) may optimize biologic and thiopurine therapies in inflammatory bowel disease (IBD). The study aimed to investigate implementation and utilization of TDM in Scandinavia. Methods A web-based questionnaire on the use of TDM was distributed to Scandinavian gastroenterologists via the national societies. Results In total, 297 IBD physicians prescribing biologic therapies, equally distributed between community and university hospitals, were included (response rate 42%) (Norway 118 (40%), Denmark 86 (29%), Sweden 50 (17%), Finland 33 (11%), Iceland 10 (3%)). Overall, TDM was applied during biologic therapies by 87%, and for TNF-inhibitors >90%. Among the users, reactive and proactive TDM were utilized by 90% and 63%, respectively. Danish physicians were significantly less inclined to use TDM compared to other Scandinavian countries; (58% vs 98%); OR 0.03 [0.01-0.09], p 10 IBD patients/week (p = 0.005). TDM scenarios were interpreted in accord with available evidence but with discrepancies for proactive TDM. The main barriers to TDM were lack of guidelines (51%) and time lag between sampling and results (49%). TDM of thiopurines was routinely used by 87%. Conclusion TDM of biologic and thiopurine therapies has been broadly implemented into clinical practice in Scandinavia. However, physicians call for TDM guidelines detailing indications and interpretations of test results along with improved test response times.Peer reviewe

Long-term follow-up with Granulocyte and Monocyte Apheresis re-treatment in patients with chronically active inflammatory bowel disease

<p>Abstract</p> <p>Background</p> <p>Patients with IBD and chronic inflammation refractory to conventional therapy often demonstrate higher risk of serious complications. Combinations of immunosuppression and biological treatment as well as surgical intervention are often used in this patient group. Hence, there is need for additional treatment options. In this observational study, focused on re-treatment and long-term results, Granulocyte/Monocyte Adsorption (GMA, Adacolumn^®) treatment has been investigated to study efficacy, safety and quality of life in IBD-patients with chronic activity.</p> <p>Methods</p> <p>Fifteen patients with ulcerative colitis and 25 patients with Crohn's disease, both groups with chronically active inflammation refractory to conventional medication were included in this observational study. The patients received 5-10 GMA sessions, and the clinical activity was assessed at baseline, after each completed course, and at week 10 and 20 by disease activity index, endoscopy and quality of life evaluation. Relapsed patients were re-treated by GMA in this follow-up study up to 58 months.</p> <p>Results</p> <p>Clinical response was seen in 85% and complete remission in 65% of the patients. Ten patients in the UC-group (66%) and 16 patients in the CD-group (64%) maintained clinical and endoscopic remission for an average of 14 months. Fourteen patients who relapsed after showing initial remission were re-treated with GMA and 13 (93%) went into a second remission. Following further relapses, all of seven patients were successfully re-treated for the third time, all of three patients for the fourth time and one for a fifth time.</p> <p>Conclusions</p> <p>IBD-patients with chronic inflammation despite conventional therapy seem to benefit from GMA. Re-treatment of relapsing remission patients seems to be effective.</p

The role of plasma membrane K+ and Ca2+ permeabilities for glucose induction of slow Ca2+ oscillations in pancreatic β-cells

AbstractIn individual pancreatic β-cells the rise of the cytoplasmic Ca2+ concentration ([Ca2+]i), induced by 11 mM glucose, is manifested either as oscillations (0.2–0.5 min−1) or as a sustained elevation. The significance of the plasma membrane permeability of Ca2+ and K+ for the establishment of these slow oscillations was investigated by dual wavelength microfluorometric measurements of [Ca2+]i in individual ob/ob mouse-β-cells loaded with fura-2. Increasing the extracellular Ca2+ to 10 mM or the addition of Ca2+ channel agonist BAY K 8644 (1 μM) or K+ channel blocker tetraethylammonium+ (TEA; 10–20 mM) caused steeper rises and higher peaks of the glucose-induced oscillations. However, when extracellular Ca2+ was lowered to 0.5 mM the oscillations were transformed into a sustained suprabasal level. When the β-cells exhibited glucose-stimulatedsustained elevation of [Ca2+]i in the presence of a physiological Ca2+ concentration (1.3 mM), it was possible to induce slow oscillations by promoting the entry of Ca2+ either by raising the extracellular Ca2+ concentration to 10 mM or adding TEA or BAY K 8644. The results indicate that glucose-induced slow oscillations of [Ca2+]i depend on the closure of ATP-regulated K+ channels and require that the rate of Ca2+ influx exceeds a critical level. Apart from an inherent periodicity in ATP production it is proposed that Ca2+-induced ATP consumption in the submembrane space contributes to the cyclic changes of the membrane potential determining periodic entry of Ca2+

Neural Control of Inflammation: Bioelectronic Medicine in Treatment of Chronic Inflammatory Disease

Inflammation is important for antimicrobial defense and for tissue repair after trauma. The inflammatory response and its resolution are both active processes that must be tightly regulated to maintain homeostasis. Excessive inflammation and nonresolving inflammation cause tissue damage and chronic disease, including autoinflammatory and cardiovascular diseases. An improved understanding of the cellular and molecular mechanisms that regulate inflammation has supported development of novel therapies for several inflammatory diseases, including rheumatoid arthritis and inflammatory bowel disease. Many of the specific anticytokine therapies carry a risk for excessive immunosuppression and serious side effects. The discovery of the inflammatory reflex and the increasingly detailed understanding of the molecular interactions between homeostatic neural reflexes and the immune system have laid the foundation for bioelectronic medicine in the field of inflammatory diseases. Neural interfaces and nerve stimulators are now being tested in human clinical trials and may, as the technology develops further, have advantages over conventional drugs in terms of better compliance, continuously adaptable control of dosing, better monitoring, and reduced risks for unwanted side effects. Here, we review the current mechanistic understanding of common autoinflammatory conditions, consider available therapies, and discuss the potential use of increasingly capable devices in the treatment of inflammatory disease