Post-Bariatric Hypoglycemia: an Impaired Metabolic Response to a Meal.

AIMS/HYPOTHESIS: Post-bariatric hypoglycemia (PBH) is caused by postprandial hyperinsulinemia, due to anatomical alterations and changes in post-prandial metabolism after bariatric surgery. The mechanisms underlying the failing regulatory and compensatory systems are unclear. In this study, we investigated the differences in post-prandial hormones and metabolic profiles between patients with and without PBH. METHODS: We performed a mixed meal test (MMT) in 63 subjects before and 1 year after Roux-en-Y gastric bypass (RYGB) surgery. Blood was withdrawn at 0, 10, 20, 30, 60, and 120 min after ingestion of a standardized meal. Glucose, insulin, GLP-1, FGF-19, and FGF-21 were measured and untargeted metabolomics analysis was performed on blood plasma to analyze which hormonal and metabolic systems were altered between patients with and without PBH. RESULTS: Out of 63, a total of 21 subjects (33%) subjects developed PBH (glucose < 3.1 mmol/L) after surgery. Decreased glucose and increased insulin excursions during MMT were seen in PBH (p < 0.05). GLP-1, FGF-19, and FGF-21 were elevated after surgery (p < 0.001), but did not differ between PBH and non-PBH groups. We identified 20 metabolites possibly involved in carbohydrate metabolism which differed between the two groups, including increased carnitine and acylcholines in PBH. CONCLUSION: Overall, 33% of the subjects developed PBH 1 year after RYGB surgery. While GLP-1, FGF-19, and FGF-21 were similar in PBH and non-PBH patients, metabolomics analysis revealed changes in carnitine and acyclcholines that are possibly involved in energy metabolism, which may play a role in the occurrence of PBH.

GIP receptor antagonism eliminates paradoxical growth hormone secretion in some patients with acromegaly.

CONTEXT: About 30% of patients with active acromegaly experience paradoxically increased growth hormone (GH) secretion during the diagnostic oral glucose tolerance test (OGTT). Endogenous glucose-dependent insulinotropic polypeptide (GIP) is implicated in this paradoxical secretion. OBJECTIVE: We used the GIP receptor (GIPR) antagonist GIP(3-30)NH2 to test the hypothesis that GIP mediates this paradoxical response when GIPR is abundantly expressed in somatotropinomas. DESIGN, PATIENTS, SETTING, INTERVENTIONS: 25 treatment-naïve patients with acromegaly were enrolled. Each patient underwent one OGTT during simultaneous placebo infusion and one OGTT during a GIP(3-30)NH2 infusion. Blood samples were drawn at baseline and regularly after infusions to measure GH. We assessed pituitary adenoma size by magnetic resonance imaging and GIPR expression by immunohistochemistry on resected somatotropinomas. For mechanistic confirmation, we applied in vitro and ex vivo approaches. MAIN OUTCOME MEASURE: The effect of GIP(3-30)NH2 on paradoxical GH secretion during OGTT as a measure of GIP involvement. RESULTS: In four of seven patients with paradoxical GH secretion, GIP(3-30)NH2 infusion completely abolished the paradoxical response (P = 0.0003). Somatotrophs were available from three of four of these patients, all showing abundant GIPR expression. Adenoma size did not differ between patients with and without paradoxical GH secretion. CONCLUSIONS: Of 25 patients with acromegaly, seven had paradoxical GH secretion during OGTT, and pharmaceutical GIPR blockade abolished this secretion in four. Corresponding somatotroph adenomas abundantly expressed GIPR, suggesting a therapeutic target in this subpopulation of patients. In vitro and ex vivo analyses confirmed the role of GIP and the effects of the antagonist.

Expression of Secretin and its Receptor Along the Intestinal Tract in Type 2 Diabetes Patients and Healthy Controls.

CONTEXT: The hormone secretin (SCT) is released from intestinal S cells and acts via the SCT receptor (SCTR). Circulating SCT levels increase after Roux-en-Y gastric bypass surgery and have been associated with massive weight loss and high remission rates of type 2 diabetes (T2D) linked to these operations. Exogenous SCT was recently shown to reduce ad libitum food intake in healthy volunteers. OBJECTIVE: To understand SCT biology and its potential role in T2D pathophysiology, we examined the intestinal mucosal expression profile of SCT and SCTR and evaluated the density of S cells along the intestinal tract of individuals with T2D and healthy controls. METHODS: Using immunohistochemistry and messenger RNA (mRNA) sequencing, we analyzed intestinal mucosa biopsies sampled along the small intestine at 30-cm intervals and from 7 well-defined anatomical sites along the large intestine (during 2 sessions of double-balloon enteroscopy) in 12 individuals with T2D and 12 healthy controls. RESULTS: Both groups exhibited a progressive and similar decrease in SCT and SCTR mRNA expression and S-cell density along the small intestine, with reductions of 14, 100, and 50 times, respectively, in the ileum compared to the duodenum (used as reference). Negligible amounts of SCTR and SCT mRNA, as well as low S-cell density, were found in the large intestine. No significant differences were observed between the groups. CONCLUSION: SCT and SCTR mRNA expression and S-cell density were abundant in the duodenum and decreased along the small intestine. Very low SCT and SCTR mRNA levels and S-cell numbers were observed in the large intestine, without aberrations in individuals with T2D compared to healthy controls.

Histological Analyses of Capsular Contracture and Associated Risk Factors: A Systematic Review.

BACKGROUND: Capsular contracture is a severe complication to breast surgery with implants. Previous studies suggest multiple risk factors are associated with capsular contracture, but the etiology is still unknown. We performed a literature review to investigate existing studies on histological analyses of breast implant capsules and how clinical risk factors impact the capsule morphology. METHODS: The literature search was conducted in PubMed. Studies that performed histological analyses of breast implant capsules were included. Animal studies or studies with a study population of less than five patients were excluded. RESULTS: Fifty-two studies were included. The histological analyses showed that the breast implant capsules were organized in multiple layers with an inner layer of synovial-like metaplasia which was reported to diminish in capsules with capsular contracture. The remaining layers of the capsule mostly consisted of collagen. The alignment of the collagen fibers differed between contracted and non-contracted capsules, and capsules with higher Baker grade were generally thickest and contained more tissue inflammation. Studies investigating capsules affected by radiotherapy found a more pronounced inflammatory response and the capsules were generally thicker and fibrotic compared with nonirradiated capsules. CONCLUSIONS: The included studies offer valuable insights into the histological changes caused by capsular contracture and their relation to clinical risk factors. Further studies with larger sample sizes and more strict inclusion criteria are needed to further investigate implant capsules and the role of the synovial-like metaplasia for the development of capsular contracture. LEVEL OF EVIDENCE III: This journal requires that authors assign a level of evidence to each article. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors https://www.springer.com/00266 .

Decreased number of colonic tuft cells in quiescent ulcerative colitis patients.

BACKGROUND: Colonic tuft cells are epithelial chemosensory cells involved in barrier integrity, modulation of inflammatory responses and gut homeostasis. Recent evidence indicates an involvement of tuft cells in ulcerative colitis pathogenesis, though mechanisms remain largely unknown.Here, we quantified the colonic tuft cell population in patients with quiescent ulcerative colitis as compared to patients without identified colonic disease (controls). METHODS: In this retrospective study, we obtained endoscopic colonic sigmoid biopsies from 14 patients with quiescent ulcerative colitis and from 17 controls. In a blinded central-reading design, we identified tuft cells by immunohistochemistry using a cyclooxygenase-1 antibody as a marker and performed a simple counting by visual inspection. Poisson regression was employed for statistics and results were adjusted for gender, age and smoking status. RESULTS: Ulcerative colitis patients demonstrated a 55% reduced tuft cell count in colonic mucosa compared with the control group (95% confidence limit: range 31-71%, P = 0.0002). Ulcerative colitis patients had a mean tuft cells count of 46 tuft cells/mm2 (95% CI, 36-59), while controls demonstrated a mean of 104 tuft cells/mm2 (95% CI, 79-136). No interactions of other covariates, such as age, smoking status, total duration of ulcerative colitis disease and duration of clinical remission prior to study inclusion were detected between ulcerative colitis patients and controls. CONCLUSION: Quiescent ulcerative colitis patients have a relatively low number of colonic tuft cells. Further studies are warranted to explore the potential involvement of tuft cells in ulcerative colitis pathogenesis.

Immunomodulatory potential of Nisin A with application in wound healing.

Antimicrobial peptides can have a dual role with both antimicrobial activity against a broad range of bacteria and immunomodulatory effect, making them attractive as therapeutic treatment of difficult wounds. Nisin A is widely known for its antimicrobial activity, and a preliminary study demonstrated that it increased wound closure, but the mechanism behind its effect is unknown. The aim of this study is to elucidate the wound healing potential of Nisin A and the mechanism behind. First, an epithelial and endothelial cell line, human keratinocyte (HaCaT) and human umbilical vein endothelial cell, were used to demonstrate migration and proliferation effects in vitro. From HaCaT cells and peripheral blood mononuclear cell, changes in cytokine levels were shown by quantitative polymerase chain reaction and enzyme-linked immunosorbent assay. Second, the ex vivo porcine wound healing model was used to investigate the re-epithelization potential of Nisin A. Finally, the model Galleria mellonella was used to confirm antimicrobial activity and to investigate potential immunomodulatory effects in vivo. Here, we demonstrated that Nisin A affected migration significantly of both human umbilical vein endothelial cell and HaCaT cells (p < 0.05) but not proliferation, potentially by decreasing the levels of proinflammatory cytokines tumor necrosis factor-α, interleukin-6, and interleukin-8 (p < 0.001). Furthermore, Nisin A treatment diminished lipopolysaccharide-induced tumor necrosis factor-α levels from peripheral blood mononuclear cells and monocyte chemoattractant protein-1 from HaCaT cells (p < 0.001). Furthermore, Nisin A did not affect proliferation ex vivo either but increased re-epithelization of the porcine skin. Nisin A improved survival of G. mellonella significantly from Staphylococcus epidermidis (p < 0.001) but not from Escherichia coli, indicating that Nisin A did not help the larvae to survive the infection in a different than direct antimicrobial way. All together this makes Nisin A a potential treatment to use in wound healing, as it increases the mobility of skin cells, dampens the effect of lipopolysaccharide and proinflammatory cytokines, and decreases bacterial growth.

Glepaglutide, a novel long-acting glucagon-like peptide-2 analogue, for patients with short bowel syndrome: a randomised phase 2 trial.

BACKGROUND: Patients with short bowel syndrome might have impaired postprandial endogenous glucagon-like peptide-2 (GLP-2) secretion, which is required for optimal intestinal adaptation. We aimed to assess the therapeutic potential of glepaglutide, a novel long-acting GLP-2 analogue, for reducing faecal output and increasing intestinal absorption in patients with short bowel syndrome. METHODS: In this single-centre, double-blind, crossover, randomised phase 2 trial, adults (aged ≥18 to ≤90 years) with short bowel syndrome and with a faecal wet weight output of 1500 g/day or more were randomly assigned to receive one of six dose sequences of glepaglutide (10 mg, 1 mg; 10 mg, 0·1 mg; 1 mg, 10 mg; 1 mg, 0·1 mg; 0·1 mg, 10 mg; or 0·1 mg, 1 mg). Patients received daily subcutaneous injections of the first assigned dose of glepaglutide for 3 weeks, followed by a washout period of 4-8 weeks, and then the second dose of glepaglutide for 3 weeks. An unmasked statistician generated the randomisation list, and the trial investigator enrolled patients and assigned them their patient numbers. Trial investigators, patients, and other care providers were masked throughout the trial. The primary endpoint was the absolute change from baseline in faecal wet weight output, measured separately over the two treatment periods. Metabolic balance studies were done before and after each treatment period to assess the primary endpoint. Per-protocol analysis was used to assess the efficacy. Safety analysis was by intention to treat. This trial is registered at ClinicalTrials.gov, number NCT02690025, and has completed. FINDINGS: Of the 22 patients screened between Feb 5, 2016, and Jan 25, 2017, 18 patients were randomly assigned and treated with glepaglutide; 16 patients completed the trial. Treatment with 1 mg and 10 mg glepaglutide changed the adjusted mean faecal output by -592 g/day (95% CI -913 to -272; p=0·002) and -833 g/day (-1152 to -515; p=0·0002) from baseline, respectively. No changes were observed with 0·1 mg glepaglutide. Of the 18 patients who were randomly assigned to treatment, common treatment-related adverse events were stoma complications (13 [72%] patients), injection site reactions (11 [61%]), peripheral oedema (ten [56%]), nausea and abdominal pain (eight [44%] each), polyuria and fatigue (six [33%] each), abdominal distention, vomiting, and dizziness (five [28%] each); and cough and decreased appetite (four [22%] each). Related or possibly related serious adverse events were reported in two patients in the 0·1 mg dose group and two patients in the 10 mg dose group. These events included abdominal pain, stoma obstruction, catheter-related sepsis, and infection of unknown origin. No patients died during the trial. INTERPRETATION: Glepaglutide was well tolerated, and was associated with improved intestinal absorption in patients with short bowel syndrome with 1 mg and 10 mg glepaglutide, but not with 0·1 mg glepaglutide. Larger phase 3 clinical trials of longer durations have been initiated to fully assess the safety and efficacy of glepaglutide. FUNDING: Zealand Pharma.

Enteroendocrine K and L cells in healthy and type 2 diabetic individuals.

AIMS/HYPOTHESIS: Enteroendocrine K and L cells are pivotal in regulating appetite and glucose homeostasis. Knowledge of their distribution in humans is sparse and it is unknown whether alterations occur in type 2 diabetes. We aimed to evaluate the distribution of enteroendocrine K and L cells and relevant prohormone-processing enzymes (using immunohistochemical staining), and to evaluate the mRNA expression of the corresponding genes along the entire intestinal tract in individuals with type 2 diabetes and healthy participants. METHODS: In this cross-sectional study, 12 individuals with type 2 diabetes and 12 age- and BMI-matched healthy individuals underwent upper and lower double-balloon enteroscopy with mucosal biopsy retrieval from approximately every 30 cm of the small intestine and from seven specific anatomical locations in the large intestine. RESULTS: Significantly different densities for cells positive for chromogranin A (CgA), glucagon-like peptide-1, glucose-dependent insulinotropic polypeptide, peptide YY, prohormone convertase (PC) 1/3 and PC2 were observed along the intestinal tract. The expression of CHGA did not vary along the intestinal tract, but the mRNA expression of GCG, GIP, PYY, PCSK1 and PCSK2 differed along the intestinal tract. Lower counts of CgA-positive and PC1/3-positive cells, respectively, were observed in the small intestine of individuals with type 2 diabetes compared with healthy participants. In individuals with type 2 diabetes compared with healthy participants, the expression of GCG and PYY was greater in the colon, while the expression of GIP and PCSK1 was greater in the small intestine and colon, and the expression of PCSK2 was greater in the small intestine. CONCLUSIONS/INTERPRETATION: Our findings provide a detailed description of the distribution of enteroendocrine K and L cells and the expression of their products in the human intestinal tract and demonstrate significant differences between individuals with type 2 diabetes and healthy participants. TRIAL REGISTRATION: NCT03044860.

Macrophage Area Content and Phenotype in Hepatic and Adipose Tissue in Patients with Obesity Undergoing Roux-en-Y Gastric Bypass.

OBJECTIVE: To investigate hepatic and adipose tissue macrophage content in subjects with obesity and the role of adipose tissue macrophages in weight loss-induced improved insulin sensitivity (IS). METHODS: A cross-sectional and a longitudinal study were combined to investigate the role of macrophages in subcutaneous (SAT) and visceral (VAT) adipose tissue and the liver in obesity-induced impaired IS and improvements with weight loss. Macrophage markers (CD68, CD163, and CD206) in SAT, VAT, and the liver from patients with obesity were investigated. The same macrophage markers were investigated in SAT from 18 patients with obesity before and ∼18 months after a diet- and Roux-en-Y gastric bypass-induced weight loss. RESULTS: SAT macrophage markers did not decrease with weight loss, but macrophage concentration may have increased, concomitant with improved IS. Hepatic macrophage markers did not correlate to VAT mass or macrophage markers, but they were higher in patients with obesity compared with patients without obesity. Hepatic anti-inflammatory macrophage markers correlated positively with hepatic IS. VAT and SAT macrophage markers did not correlate. CONCLUSIONS: The results indicate that decreased SAT macrophage content is not a primary driver for weight loss-induced IS improvements, but a better hepatic CD163 and CD206 macrophage profile may contribute to improved glycemic control. SAT macrophage markers were not predictive for VAT macrophage markers.

Repeated Excessive Exercise Attenuates the Anti-Inflammatory Effects of Exercise in Older Men.

Introduction/Purpose: A number of studies have investigated the effect of training with a moderate exercise dose (3-6 h/weekly) on the inflammatory profile in blood, and the data are inconsistent. Cross-sectional studies indicate a positive effect of physical activity level on inflammation levels and risk of metabolic disease. However, it is not clear whether this may be dose dependent and if very prolonged repeated exercise therefore may be beneficial for low-grade inflammation. Based on this we studied how excessive repeated prolonged exercise influenced low-grade inflammation and adipose tissue anti-inflammatory macrophage content in six older male recreationally trained cyclists. Low-grade inflammation and adipose tissue macrophage content were investigated in six older trained men (age: 61 ± 4 years; VO2peak: 48 ± 2 mL kg-1 min-1) following repeated prolonged exercise. Methods: Cycling was performed daily for 14 days covering in total 2,706 km (1,681 miles). Maximal oxygen uptake (VO2peak) was measured before and after the cycling. Duration and intensity of the exercise were determined from heart rates sampled during cycling. An adipose tissue biopsy from subcutaneous abdominal fat and a blood sample were obtained at rest in the overnight fasted state before and after the cycling. Anti-inflammatory adipose tissue macrophages (ATM) were immunohistochemically stained in cross sectional sections using a CD163 binding antibody. The ATM and adipocyte sizes were analyzed blindly. Results: The cyclists exercised daily for 10 h and 31 ± 37 min and average intensity was 53 ± 1% of VO2peak. Body weight remained unchanged and VO2peak decreased by 6 ± 2% (P = 0.04). Plasma inflammatory cytokines, TNFα and IL-18 remained unchanged, as did hsCRP, but plasma IL-6 increased significantly. CD163 macrophage content remained unchanged, as did adipocyte cell size. The HbA1c was not significantly decreased, but there was a trend (P < 0.07) toward an increased insulin resistance as estimated by the Quicki Index. Conclusion: The regular prolonged exercise did not influence abdominal adipose tissue inflammation, but the higher plasma IL-6 concentration concurrent with a trend toward higher insulin resistance and decreased VO2peak implies that the excessive amount of exercise probably attenuated the possible potential anti-inflammatory effects of exercise.

Glucagon-like Peptide 1 Receptor Signaling in Acinar Cells Causes Growth-Dependent Release of Pancreatic Enzymes.

Incretin-based therapies are widely used for type 2 diabetes and now also for obesity, but they are associated with elevated plasma levels of pancreatic enzymes and perhaps a modestly increased risk of acute pancreatitis. However, little is known about the effects of the incretin hormone glucagon-like peptide 1 (GLP-1) on the exocrine pancreas. Here, we identify GLP-1 receptors on pancreatic acini and analyze the impact of receptor activation in humans, rodents, isolated acini, and cell lines from the exocrine pancreas. GLP-1 did not directly stimulate amylase or lipase release. However, we saw that GLP-1 induces phosphorylation of the epidermal growth factor receptor and activation of Foxo1, resulting in cell growth with concomitant enzyme release. Our work uncovers GLP-1-induced signaling pathways in the exocrine pancreas and suggests that increases in amylase and lipase levels in subjects treated with GLP-1 receptor agonists reflect adaptive growth rather than early-stage pancreatitis.

Mass spectrometry imaging of biomarker lipids for phagocytosis and signalling during focal cerebral ischaemia.

Focal cerebral ischaemia has an initial phase of inflammation and tissue injury followed by a later phase of resolution and repair. Mass spectrometry imaging (desorption electrospray ionization and matrix assisted laser desorption ionization) was applied on brain sections from mice 2 h, 24 h, 5d, 7d, and 20d after permanent focal cerebral ischaemia. Within 24 h, N-acyl-phosphatidylethanolamines, lysophosphatidylcholine, and ceramide accumulated, while sphingomyelin disappeared. At the later resolution stages, bis(monoacylglycero)phosphate (BMP(22:6/22:6)), 2-arachidonoyl-glycerol, ceramide-phosphate, sphingosine-1-phosphate, lysophosphatidylserine, and cholesteryl ester appeared. At day 5 to 7, dihydroxy derivates of docosahexaenoic and docosapentaenoic acid, some of which may be pro-resolving mediators, e.g. resolvins, were found in the injured area, and BMP(22:6/22:6) co-localized with the macrophage biomarker CD11b, and probably with cholesteryl ester. Mass spectrometry imaging can visualize spatiotemporal changes in the lipidome during the progression and resolution of focal cerebral inflammation and suggests that BMP(22:6/22:6) and N-acyl-phosphatidylethanolamines can be used as biomarkers for phagocytizing macrophages/microglia cells and dead neurones, respectively.

Effect of Roux-en-Y gastric bypass on the distribution and hormone expression of small-intestinal enteroendocrine cells in obese patients with type 2 diabetes.

AIMS/HYPOTHESIS: We studied the impact of Roux-en-Y gastric bypass (RYGB) on the density and hormonal gene expression of small-intestinal enteroendocrine cells in obese patients with type 2 diabetes. METHODS: Twelve patients with diabetes and 11 age- and BMI-matched controls underwent RYGB followed by enteroscopy ~10 months later. Mucosal biopsies taken during surgery and enteroscopy were immunohistochemically stained for glucagon-like peptide-1 (GLP-1), peptide YY (PYY), cholecystokinin (CCK), glucose-dependent insulinotropic polypeptide (GIP) and prohormone convertase 2 (PC2) and the expression of GCG (encoding preproglucagon), PYY, CCK, GIP, GHRL (encoding ghrelin), SCT (encoding secretin), NTS (encoding neurotensin) and NR1H4 (encoding farnesoid X receptor) was evaluated. RESULTS: The density of cells immunoreactive for GLP-1, CCK and GIP increased in patients after RYGB and the density of those immunoreactive for GLP-1, PYY, CCK and PC2 increased in controls. In both groups, GHRL, SCT and GIP mRNA was reduced after RYGB while PYY, CCK, NTS and NR1H4 gene expression was unaltered. GCG mRNA was upregulated in both groups. CONCLUSIONS/INTERPRETATION: Numerous alterations in the distribution of enteroendocrine cells and their expression of hormonal genes are seen after RYGB and include increased density of GLP-1-, PYY-, CCK-, GIP- and PC2-positive cells, reduced gene expression of GHRL, SCT and GIP and increased expression of GCG.

Activation of GLP-1 receptors on vascular smooth muscle cells reduces the autoregulatory response in afferent arterioles and increases renal blood flow.

Glucagon-like peptide (GLP)-1 has a range of extrapancreatic effects, including renal effects. The mechanisms are poorly understood, but GLP-1 receptors have been identified in the kidney. However, the exact cellular localization of the renal receptors is poorly described. The aim of the present study was to localize renal GLP-1 receptors and describe GLP-1-mediated effects on the renal vasculature. We hypothesized that renal GLP-1 receptors are located in the renal microcirculation and that activation of these affects renal autoregulation and increases renal blood flow. In vivo autoradiography using (125)I-labeled GLP-1, (125)I-labeled exendin-4 (GLP-1 analog), and (125)I-labeled exendin 9-39 (GLP-1 receptor antagonist) was performed in rodents to localize specific GLP-1 receptor binding. GLP-1-mediated effects on blood pressure, renal blood flow (RBF), heart rate, renin secretion, urinary flow rate, and Na(+) and K(+) excretion were investigated in anesthetized rats. Effects of GLP-1 on afferent arterioles were investigated in isolated mouse kidneys. Specific binding of (125)I-labeled GLP-1, (125)I-labeled exendin-4, and (125)I-labeled exendin 9-39 was observed in the renal vasculature, including afferent arterioles. Infusion of GLP-1 increased blood pressure, RBF, and urinary flow rate significantly in rats. Heart rate and plasma renin concentrations were unchanged. Exendin 9-39 inhibited the increase in RBF. In isolated murine kidneys, GLP-1 and exendin-4 significantly reduced the autoregulatory response of afferent arterioles in response to stepwise increases in pressure. We conclude that GLP-1 receptors are located in the renal vasculature, including afferent arterioles. Activation of these receptors reduces the autoregulatory response of afferent arterioles to acute pressure increases and increases RBF in normotensive rats.

NKG2D-dependent activation of dendritic epidermal T cells in contact hypersensitivity.

The interaction between keratinocytes (KCs) and skin-resident immune cells has an important role in induction of contact hypersensitivity. A specific subset of γδ T cells termed dendritic epidermal T cells (DETCs) are located in mouse epidermis, and we have recently shown that DETCs become activated and produce IL-17 in an IL-1ß-dependent manner during contact hypersensitivity. Various receptors on DETCs, including NKG2D, are involved in DETC responses against tumors and during wound healing. The ligands for NKG2D (NKG2DL) are stress-induced proteins such as mouse UL16-binding protein-like transcript 1 (Mult-1), histocompatibility 60 (H60), and retinoic acid early inducible-1 (Rae-1) in mice and major histocompatibility complex (MHC) class I-chain-related A (MICA), MHC class I-chain-related B, and UL16-binding protein in humans. Here, we show that allergens upregulate expression of the NKG2DL Mult-1, H60, and Rae-1 in cultured mouse KCs and of MICA in primary human KCs. We demonstrate that Mult-1 is expressed in mouse skin exposed to allergen. Furthermore, we find that the vast majority of DETCs in murine epidermis and skin-homing cutaneous lymphocyte-associated antigen positive γδ T cells in humans express NKG2D. Finally, we demonstrate that blocking of NKG2D partially inhibits allergen-induced DETC activation. These findings demonstrate that NKG2D and NKG2DL are involved in allergen-induced activation of DETCs and indicate that the NKG2D/NKG2DL pathway might be a potential target for treatment of contact hypersensitivity.

Epicutaneous exposure to nickel induces nickel allergy in mice via a MyD88-dependent and interleukin-1-dependent pathway.

BACKGROUND: Several attempts to establish a model in mice that reflects nickel allergy in humans have been made. Most models use intradermal injection of nickel in combination with adjuvant to induce nickel allergy. However, such models poorly reflect induction of nickel allergy following long-lasting epicutaneous exposure to nickel. OBJECTIVE: To develop a mouse model reflecting nickel allergy in humans induced by epicutaneous exposure to nickel, and to investigate the mechanisms involved in such allergic responses. METHODS: Mice were exposed to NiCl2 on the dorsal side of the ears. Inflammation was evaluated by the swelling and cell infiltration of the ears. T cell responses were determined as numbers of CD4+ and CD8+ T cells in the draining lymph nodes. Localization of nickel was examined by dimethylglyoxime staining. RESULTS: Epicutaneous exposure to nickel results in prolonged localization of nickel in the epidermis, and induces nickel allergy in mice. The allergic response to nickel following epicutaneous exposure is MyD88-dependent and interleukin (IL)-1 receptor-dependent, but independent of toll-like receptor (TLR)-4. CONCLUSION: This new model for nickel allergy that reflects epicutaneous exposure to nickel in humans shows that nickel allergy is dependent on MyD88 and IL-1 receptor signalling, but independent of TLR4.

HER4 and its cytoplasmic isoforms are associated with progression-free survival of malignant melanoma.

HER4 belongs to the epidermal growth factor (EGF) family. Mutations in HER4 are associated with malignant melanoma. This points to HER4 as an important receptor in malignant melanoma and also raises the question of whether the other receptors in the EGF system could be involved. RT-qPCR mRNA quantification was carried out of all four EGF receptors (EGFR, HER2, HER3, and HER4) and the HER4 cytoplasmic isoforms in lymph node metastases from patients with malignant melanoma. We related their expression to progression of the disease. HER4 expression was found to be an indicator of short progression-free survival (P=0.0340). Interestingly, of the two cytoplasmic splice variants of HER4, the association of CYT1 (P=0.0176) with progression-free survival was more pronounced than that for CYT2 (P=0.0458). Also, HER3 was associated with progression-free survival (P=0.0169), whereas no association was found for EGFR or HER2 with time to progression. Our results further emphasize the role of HER4 as an important oncogene in malignant melanoma and point to HER4 as a possible drug target in this disease.

Drug-induced hypothermia as beneficial treatment before and after cerebral ischemia.

OBJECTIVES: Hypothermia is still unproven as beneficial treatment in human stroke, although in animal models, conditioning the brain with hypothermia has induced tolerance to insults. Here, we delineate the feasibility of drug-induced mild hypothermia in reducing ischemic brain damage when conditioning before (preconditioning) and after (postconditioning) experimental stroke. METHODS: Hypothermia was induced in rats with a bolus of 6 mg/kg talipexole followed by 20 h continuous talipexole infusion of 6 mg/kg in total. Controls received similar treatment with saline. The core body temperature was continuously monitored. In preconditioning, hypothermia was terminated before either reversible occlusion of the middle cerebral artery (MCAO) for 60 min or global ischemia for 10 min with 2-vessel occlusion and hypotension. In postconditioning, rats experienced 60 min of MCAO before hypothermia was induced either immediately or with 3 h delay. Rats survived ischemia for 2, 7 or 90 days. Infarct volumes were quantified by stereology. Additional experiments of methodological relevance were included in the study. RESULTS: Talipexole induced mild hypothermia (35.1±1.1 to 36.0±0.5°C) for <20 h. Hypothermic pre- and postconditioning reduced infarct sizes by more than 60% as monitored during the first 90 days after experimental stroke (p<0.05). CONCLUSION: Talipexole is registered for use as a dopamine substitute in humans with Parkinson's disease. Although dosages cannot be directly translated to patients, our study exemplifies in an animal model that drug-induced hypothermia in a clinical setting might reduce cerebral ischemic damage before neuro- and cardiac surgical procedures and after stroke.

Glucagon-like peptide-1 (GLP-1) reduces mortality and improves lung function in a model of experimental obstructive lung disease in female mice.

The incretin hormone glucagon-like peptide-1 (GLP-1) is an important insulin secretagogue and GLP-1 analogs are used for the treatment of type 2 diabetes. GLP-1 displays antiinflammatory and surfactant-releasing effects. Thus, we hypothesize that treatment with GLP-1 analogs will improve pulmonary function in a mouse model of obstructive lung disease. Female mice were sensitized with injected ovalbumin and treated with GLP-1 receptor (GLP-1R) agonists. Exacerbation was induced with inhalations of ovalbumin and lipopolysaccharide. Lung function was evaluated with a measurement of enhanced pause in a whole-body plethysmograph. mRNA levels of GLP-1R, surfactants (SFTPs), and a number of inflammatory markers were measured. GLP-1R was highly expressed in lung tissue. Mice treated with GLP-1R agonists had a noticeably better clinical appearance than the control group. Enhanced pause increased dramatically at day 17 in all control mice, but the increase was significantly less in the groups of GLP-1R agonist-treated mice (P < .001). Survival proportions were significantly increased in GLP-1R agonist-treated mice (P < .01). SFTPB and SFTPA were down-regulated and the expression of inflammatory cytokines were increased in mice with obstructive lung disease, but levels were largely unaffected by GLP-1R agonist treatment. These results show that GLP-1R agonists have potential therapeutic potential in the treatment of obstructive pulmonary diseases, such as chronic obstructive pulmonary disease, by decreasing the severity of acute exacerbations. The mechanism of action does not seem to be the modulation of inflammation and SFTP expression.

The HER4 isoform JM-a/CYT2 relates to improved survival in bladder cancer patients but only if the estrogen receptor α is not expressed.

Bladder cancer tumors expressing human epidermal growth factor receptor 4 (HER4) demonstrate improved patient survival. HER4 isoforms and estrogen receptor alpha (ER-α) can form chaperone complexes causing cell-proliferation. We wanted to explore if HER4 isoforms and ER-α could correlate to poor prognosis in bladder cancers. We developed mRNA assays for HER4 isoforms (JM-a, JM-b, CYT1, and CYT2) and for ER-α. Expression was analyzed in tumors from 85 bladder cancer patients and compared to overall survival (median follow-up of 5.1 years). ER-α was expressed in 38% (n = 32) of tumors but did not correlate to survival (p = 0.4698). HER4 was expressed in 42% (n = 36) and in all cases as the ER-α binding isoform JM-a. The JM-a isoform can be alternatively spliced to either a CYT1 isoform (JM-a/CYT1) or a CYT2 isoform (JM-a/CYT2). All HER4 expressing tumors expressed the JM-a/CYT2 isoform and half of those (18/36) expressed both isoforms. JM-a/CYT2 expression correlated to improved survival (p = 0.004), but not when ER-α was co-expressed (p = 0.897). Immunohistochemistry revealed protein expression of HER4 and ER-α in tumor cells. Growth of RT4 bladder cancer cells, expressing both JM-a/CYT2 and ER-α was inhibited by the specific ER-α inhibitor raloxifene. Likewise, stable transfection with JM-a/CYT2 inhibited the growth of T24 bladder cancer cells, but only when ER-α was inhibited. Our results demonstrate that HER4 JM-a/CYT2 expressing bladder cancers relate to favorable prognosis when ER-α is not co-expressed. In vitro studies indicate that ER-α inhibition may be a useful treatment for patients with tumors expressing both ER-α and HER4 JM-a/CYT2.