CD26 Deficiency Controls Macrophage Polarization Markers and Signal Transducers during Colitis Development and Resolution.

Ulcerative colitis (UC) is a multifactorial condition characterized by a destructive immune response that failed to be attenuated by common regulatory mechanisms which reduce inflammation and promote mucosa healing. The inhibition of CD26, a multifunctional glycoprotein that controls the immune response via its dipeptidyl peptidase (DP) 4 enzyme activity, was proven to have beneficial effects in various autoimmune inflammatory diseases. The polarization of macrophages into either pro-inflammatory M1 or anti-inflammatory M2 subclass is a key intersection that mediates the immune-inflammatory process in UC. Hence, we hypothesized that the deficiency of CD26 affects that process in the dextran sulfate sodium (DSS)-induced model of UC. We found that mRNA expression of M2 markers arginase 1 and Fizz were increased, while the expression of M1 marker inducible NO synthase was downregulated in CD26-/- mice. Decreased STAT1 mRNA, as well as upregulated pSTAT6 and pSTAT3, additionally support the demonstrated activation of M2 macrophages under CD26 deficiency. Finally, we investigated DP8 and DP9, proteins with DP4-like activity, and found that CD26 deficiency is not a key factor for the noted upregulation of their expression in UC. In conclusion, we demonstrate that CD26 deficiency regulates macrophage polarization toward the anti-inflammatory M2 phenotype, which is driven by STAT6/STAT3 signaling pathways. This process is additionally enhanced by the reduction of M1 differentiation via the suppression of proinflammatory STAT1. Therefore, further studies should investigate the clinical potential of CD26 inhibitors in the treatment of UC.

The influence of CD26+ and CD26- fibroblasts on the regeneration of human dermo-epidermal skin substitutes.

CD26, also known as dipeptidyl peptidase IV (DPPIV), is a multifunctional transmembrane protein playing a significant role in the cutaneous wound healing processes in the mouse skin. However, only scarce data are available regarding the distribution and function of this protein in the human skin. Therefore, the aim of this study was to investigate the impact of CD26 deficiency in human primary fibroblasts on the regeneration of human tissue-engineered skin substitutes in vivo. Dermo-epidermal skin analogs, based on collagen type I hydrogels, were populated either with human CD26+ or CD26knockout fibroblasts and seeded with human epidermal keratinocytes. These skin substitutes were transplanted onto the back of immune-incompetent rodents. Three weeks post-transplantation, the grafts were excised and analyzed with respect to specific epidermal and dermal maturation markers. For the first time, we show here that the expression of CD26 protein in human dermis is age-dependent. Furthermore, we prove that CD26+ fibroblasts are more active in the production of extracellular matrix (ECM) both in vitro and in vivo and are necessary to achieve rapid epidermal and dermal homeostasis after transplantation.

Lung development and immune status under chronic LPS exposure in rat pups with and without CD26/DPP4 deficiency.

Dipeptidyl-peptidase IV (CD26), a multifactorial integral type II protein, is expressed in the lungs during development and is involved in inflammation processes. We tested whether daily LPS administration influences the CD26-dependent retardation in morphological lung development and induces alterations in the immune status. Newborn Fischer rats with and without CD26 deficiency were nebulized with 1 µg LPS/2 ml NaCl for 10 min from days postpartum (dpp) 3 to 9. We used stereological methods and fluorescence activated cell sorting (FACS) to determine morphological lung maturation and alterations in the pulmonary leukocyte content on dpp 7, 10, and 14. Daily LPS application did not change the lung volume but resulted in a significant retardation of alveolarization in both substrains proved by significantly lower values of septal surface and volume as well as higher mean free distances in airspaces. Looking at the immune status after LPS exposure compared to controls, a significantly higher percentage of B lymphocytes and decrease of CD4+CD25+ T cells were found in both subtypes, on dpp7 a significantly higher percentage of CD4 T+ cells in CD26+ pups, and a significantly higher percentage of monocytes in CD26- pups. The percentage of T cells was significantly higher in the CD26-deficient group on each dpp. Thus, daily postnatal exposition to low doses of LPS for 1 week resulted in a delay in formation of secondary septa, which remained up to dpp 14 in CD26- pups. The retardation was accompanied by moderate parenchymal inflammation and CD26-dependent changes in the pulmonary immune cell composition.

Dipeptidyl Peptidase-4 deficiency effectively protects the brain and neurological function in rodent after acute Hemorrhagic Stroke.

This study tested the hypothesis that abrogated dipeptidyl peptidase-4 (DPP4) activity played a crucial role on reducing stroke volume and preserving neurological function in rodent after acute hemorrhagic stroke (AHS). Animals (n=6/each group) were categorized into group 1 (sham-control of F344 rat), group 2 (sham-control of DPP4-deficiency rat), group 3 [AHS by right cerebral injection of autologous blood (100 µL) in F344 rat], group 4 (AHS + sitagliptin/600 mg/kg 3 h prior to and at 3 h then once per day after AHS) and group 5 (AHS in DPP4-deficiency rat). The results of corner test showed the neurological function was significantly improved from days 3, 7, and 14 in groups 4 and 5 than in group 3 (all p<0.001). By days 1 and 14 after AHS procedure, the circulating levels of SDF-1α and GLP-1 were significantly increased from groups 1/2 to group 5 (all p<0.001), whereas circulating DPP4 activity was significantly increased in group 3 than other groups (all p<0.001). The brain ischemic area (BIA) was highest in group 3, lowest in groups 1/2 and significantly lower in group 5 than in group 4 (all p<0.0001). The protein expressions of oxidative-stress/inflammatory/apoptotic/cell-proliferation signaling, and the cellular expressions of inflammatory/DNA-damaged biomarkers exhibited a similar pattern to BIA among the groups (all p<0.01). In conclusion, deprivation of DPP4 activity protected the brain from AHS damage and preserved neurological function.

Incorporating Pharmacological Target-Mediated Drug Disposition (TMDD) in a Whole-Body Physiologically Based Pharmacokinetic (PBPK) Model of Linagliptin in Rat and Scale-up to Human.

Linagliptin demonstrates substantial nonlinear pharmacokinetics due to its saturable binding to its pharmacological target dipeptidyl peptide 4 (DPP-4), a phenomenon known as target-mediated drug disposition (TMDD). In the current study, we established a novel whole-body physiologically-based pharmacokinetic (PBPK)-TMDD model for linagliptin. This comprehensive model contains plasma and 14 tissue compartments, among which TMDD binding process was incorporated in 9 of them, namely the plasma, kidney, liver, spleen, lung, skin, salivary gland, thymus, and reproductive organs. Our final model adequately captured the concentration-time profiles of linagliptin in both plasma and various tissues in both wildtype rats and DPP4-deficient rats following different doses. The association rate constant (kon) in plasma and tissues were estimated to be 0.943 and 0.00680 nM-1 h-1, respectively, and dissociation rate constant (koff), in plasma and tissues were estimated to be 0.0698 and 0.00880 h-1, respectively. The binding affinity of linagliptin to DPP-4 (Kd) was predicted to be higher in plasma (0.0740 nM) than that in tissue (1.29 nM). When scaled up to a human, this model captured the substantial and complex nonlinear pharmacokinetic behavior of linagliptin in human adults that is characterized by less-than dose-proportional increase in plasma exposure, dose-dependent clearance and volume of distribution, as well as long terminal half-life with minimal accumulation after repeated doses. Our modeling work is not only novel but also of high significance as the whole-body PBPK-TMDD model platform developed using linagliptin as the model compound could be applied to other small-molecule compounds exhibiting TMDD to facilitate their optimal dose selection. Graphical abstract.

Deficiency in Dipeptidyl Peptidase-4 Promotes Chemoresistance through the CXCL12/CXCR4/mTOR/TGFß Signaling Pathway in Breast Cancer Cells.

Dipeptidyl peptidase (DPP)-4, a molecular target of DPP-4 inhibitors, which are type 2 diabetes drugs, is expressed in a variety of cell types, tissues and organs. DPP-4 has been shown to be involved in cancer biology, and we have recently shown that a DPP-4 inhibitor promoted the epithelial mesenchymal transition (EMT) in breast cancer cells. The EMT is known to associate with chemotherapy resistance via the induction of ATP-binding cassette (ABC) transporters in cancer cells. Here, we demonstrated that deficiency in DPP-4 promoted chemotherapy resistance via the CXCL12/CXCR4/mTOR axis, activating the TGFß signaling pathway via the expression of ABC transporters. DPP-4 inhibition enhanced ABC transporters in vivo and in vitro. Doxorubicin (DOX) further induced ABC transporters in DPP-4-deficient 4T1 cells, and the induction of ABC transporters was suppressed by either the CXCR4 inhibitor AMD3100, the mTOR inhibitor rapamycin or a neutralizing TGFß (1, 2 and 3) antibody(N-TGFß). Knockdown of snail, an EMT-inducible transcription factor, suppressed ABC transporter levels in DOX-treated DPP-4-deficient 4T1 cells. In an allograft mouse model, however, the effects of DOX in either primary tumor or metastasis were not statistically different between control and DPP-4-kd 4T1. Taken together, our findings suggest that DPP-4 inhibitors potentiate chemotherapy resistance via the induction of ABC transporters by the CXCL12/CXCR4/mTOR/TGFß signaling pathway in breast cancer cells.

Attenuation of diabetic kidney injury in DPP4-deficient rats; role of GLP-1 on the suppression of AGE formation by inducing glyoxalase 1.

Dipeptidyl peptidase 4 (DPP4) inactivates incretin hormone glucagon-like peptide-1. DPP4 inhibitors may exert beneficial effects on diabetic nephropathy (DN) independently of glycemic control; however, the mechanisms underlying are not fully understood. Here, we investigated the mechanisms of the beneficial effects of DPP4 inhibition on DN using DPP4-deficient (DPP4-def) rats and rat mesangial cells.Blood glucose and HbA1c significantly increased by streptozotocin (STZ) and no differences were between WT-STZ and DPP4-def-STZ. The albumin level in urine decreased significantly and the albumin/creatinine ratio decreased slightly in DPP4-def-STZ. The glomerular volume in DPP4-def-STZ significantly decreased compared with that of WT-STZ. Advanced glycation end products formation, receptor for AGE (RAGE) protein expression, and its downstream inflammatory cytokines and fibrotic factors in kidney tissue, were significantly suppressed in the DPP4-def-STZ compared to the WT-STZ with increasing glyoxalase-1 (GLO-1) expression responsible for the detoxification of methylglyoxal (MGO). In vitro, exendin-4 suppressed MGO-induced AGEs production by enhancing the expression of GLO-1 and nuclear factor-erythroid 2 p45 subunit-related factor 2, resulting in decreasing pro-inflammatory cytokine levels. This effect was abolished by GLO-1 siRNA.Our data suggest that endogenously increased GLP-1 in DPP4-deficient rats contributes to the attenuation of DN partially by regulating AGEs formation via upregulation of GLO-1 expression.

Postnatal morphological lung development of wild type and CD26/DPP4 deficient rat pups in dependency of LPS exposure.

BACKGROUND: Rodents are born with morphological immature lungs and an intact surfactant system. CD26/DPP4 is a multifactorial transmembrane integral type II protein, which is involved in physiological and pathophysiological processes and is already expressed during development. CD26/DPP4, called CD26 in the following, is able to enhance or dampen differently triggered inflammation. LPS exposure often used to simulate perinatal infection delays lung development. OBJECTIVE: A perinatal LPS rat model was used to test the hypothesis that CD26 deficiency modulates LPS-induced retardation in morphological lung development. METHODS: New born Fischer CD26 positive (CD26+) and deficient (CD26-) rats were exposed to LPS on postnatal day (day post partum, dpp) 3 and 5. Morphological parameters of lung development were determined stereologically. Lung development was analysed in 7, 10 14 and 21day old rats. RESULTS: Compared to controls LPS application resulted (1) in a mild inflammation independent of the strain, (2) in significantly lower total surface and volume of alveolar septa combined with significantly higher total volume of airspaces and alveolar size on dpp 7 in both substrains. However, compared to controls in LPS treated CD26- rats significant lower values of total septal surface and volume combined with higher values of total parenchymal airspaces and alveolar size were found until the end of classical alveolarization (dpp14). In LPS treated CD26+ rat pups the retardation was abolished already on dpp 10. CONCLUSION: In absence of CD26, LPS enhances the delay of morphological lung development. Morphological recovery was slower after the end of LPS exposure in CD26 deficient lungs.

Differential severity of LPS-induced lung injury in CD26/DPP4 positive and deficient F344 rats.

BACKGROUND: Lipopolysaccharide (LPS) induced inflammation often leads to lung injury, in which pulmonary recruitment of neutrophils plays a pivotal role. Inflammatory processes are influenced by CD26/DPP4, highly expressed in lungs. Asthma induced CD26/DPP4 deficient (CD26/DPP4⁻) Fischer (F) 344 rats suffering from a transport block in the rER caused by a point mutation showed reduced pulmonary inflammation and reduced expression of immunomodulating surfactant proteins (SP). The degree of LPS induced lung injury in CD26/DPP4 deficient rats has not been investigated so far. OBJECTIVE: We hypothesize that LPS induced lung injury leads not only to an attenuated inflammation but also to a reduced SP expression and decreased structural damage in CD26/DPP4⁻ rats. METHODS: Both genotypes were intratracheally instilled with 250 µl LPS or with 250 µl 0.9% NaCl. Nine hours later animals were killed and either bronchoalveolar lavage was carried out to determine inflammatory cells and surface tension or lung blocks were removed and processed for histology, immunohistochemistry, electron microscopy or qRt-PCR analyses and Western Blot analyses. RESULTS: Signs of acute lung injury, such as structural damage of the blood gas barrier occurred only sporadically in both genotypes. LPS-induced CD26/DPP4⁻ rats showed decreased gene expression of SP-A and SP-D and reduced signs of lung inflammation associated with a reduced alveolar influx of macrophages and neutrophils. CONCLUSIONS: Less pulmonary inflammation combined with less structural alterations and minor expression of immunomodulating SP may be an indication of the critical role of CD26/DPP4 in regulating lung inflammation.

Efficient Reconstitution of Hepatic Microvasculature by Endothelin Receptor Antagonism in Liver Sinusoidal Endothelial Cells.

Reconstitution of healthy endothelial cells in vascular beds offers opportunities for mechanisms in tissue homeostasis, organ regeneration, and correction of deficient functions. Liver sinusoidal endothelial cells express unique functions, and their transplantation is relevant for disease models and for cell therapy. As molecular targets for improving transplanted cell engraftment and proliferation will be highly significant, this study determined whether ETA/B receptor antagonism by the drug bosentan could overcome cell losses due to cell transplantation-induced cytotoxicity. Cell engraftment and proliferation assays were performed with healthy wild-type liver sinusoidal endothelial cells transplanted into the liver of dipeptidylpeptidase IV knockout mice. Transplanted cells were identified in tissues by enzyme histochemistry. Cells with prospective ETA/B antagonism engrafted significantly better in hepatic sinusoids. Moreover, these cells underwent multiple rounds of division under liver repopulation conditions. The gains of ETA/B antagonism resulted from benefits in cell viability and membrane integrity. Also, in bosentan-treated cells, mitochondrial homeostasis was better maintained with less oxidative stress and DNA damage after injuries. Intracellular effects of ETA/B antagonism were transduced by conservation of ataxia telangiectasia mutated protein, which directs DNA damage response. Therefore, ETA/B antagonism in donor cells will advance vascular reconstitution. Extensive experience with ETA/B antagonists will facilitate translation in people.

Role of hypothalamus-pituitary-adrenal axis modulation in the stress-resilient phenotype of DPP4-deficient rats.

BACKGROUND: Dipeptidyl peptidase 4 (DPP4, CD26) is a moonlighting enzyme responsible for the proteolytic inactivation of neuropeptide Y (NPY), a peptide known for its anxiolytic effect in the central nervous system. Our previous work revealed a stress-resilient phenotype and a potentiation of short-term fear extinction in a congenic rat model deficient for DPP4 activity (DPP4mut). Here, we investigated neuroendocrine mechanisms underlying the phenotype of the DPP4mut animals. We studied the function of the hypothalamus-pituitary-adrenal (HPA) axis including the expression levels of its key genes and explored the possibility of structural NPY system changes. METHODS AND RESULTS: We find decreased expression of Nr3c1 (glucocorticoid receptor - GR) and Fkbp5 (FK506 binding protein 5) in the amygdala and the hypothalamus of the DPP4mut rats, as well as the lower stress-induced peripheral corticosterone (CORT) levels. We detect no significant alterations in basal and DEX-induced CORT levels in the DPP4mut animals. The abundance of NPY-ergic neurons in the basolateral amygdala, dentate gyrus and hippocampus did not differ between the DPP4mut and their wild type littermates. CONCLUSION: DPP4mut rats show blunted CORT response in line with their lower behavioral stress-response profile. These results are consistent with the hypothesis that increased central NPY levels elevate the threshold of stress response. We suggest that changes in the expression levels of key HPA axis genes (Nr3c1 and Fkbp5) are a consequence of the altered stress-perception of DPP4mut animals, thus further contributing to the stress-resilient phenotype.

Advanced glycation end products evoke inflammatory reactions in proximal tubular cells via autocrine production of dipeptidyl peptidase-4.

We have previously shown that albuminuria and renal levels of advanced glycation end products (AGEs), receptor for AGEs (RAGE), and oxidative stress are suppressed in dipeptidyl peptidase-4 (DPP-4)-deficient diabetic rats, thus suggesting the crosstalk between AGE-RAGE axis and DPP-4 in experimental diabetic nephropathy. Therefore, we examined here the role of DPP-4 in AGE-evoked inflammatory reactions in human proximal tubular cells. Proteins were extracted from proximal tubular cells, and conditioned medium was collected, both of which were subjected to western blot analysis using anti-DPP-4 antibody. RAGE-aptamer was prepared using a systemic evolution of ligands by exponential enrichment. NF-κB p65 and monocyte chemoattractant protein-1 (MCP-1) gene expression was analyzed by reverse transcription-polymerase chain reaction. AGEs significantly increased DPP-4 expression and soluble DPP-4 production by tubular cells, the latter of which was attenuated by RAGE-aptamer or an anti-oxidant, N-acetylcysteine. AGEs or DPP-4 up-regulated NF-κB p65 or MCP-1 mRNA levels in tubular cells, which were suppressed by linagliptin, an inhibitor of DPP-4. AGEs stimulated NF-κB p65 gene expression in tubular cells isolated from control rats, but not from DPP-4-deficient rats. Our present results suggest that the AGE-RAGE-mediated oxidative stress could evoke inflammatory reactions in proximal tubular cells via autocrine production of DPP-4.

The effect of CD26-deficiency on dipeptidyl peptidase 8 and 9 expression profiles in a mouse model of Crohn's disease.

The involvement of dipeptidyl peptidase (DP) IV/CD26 (DPP IV/CD26) family members in the pathogenesis of Crohn's disease (CD), an autoimmune inflammatory condition of the gut, is effected mainly through proteolytic cleavage of immunomodulatory substrates and DPP IV/CD26's costimulatory function. DP8 and DP9 are proteases with diverse functions including cell interactions, apoptosis, and immune response but their localization remains to be clarified. We assessed the impact of DPP IV/CD26 deficiency (CD26-/- ) on the expression profiles of DP8 and DP9 by qPCR and immunodetection as well as quantified DP8/9 enzyme activity in distinctive phases of a chemically-induced CD model in mice. CD26-/- did not affect colon DP8 mRNA expression, while the physiological concentration of DP8 protein is decreased in CD26-/- mice but rises in inflammation (P < 0.05). On the other hand, DP9 mRNA level is significantly increased in CD26-/- mice in inflammation as well as healing with the DP9 concentration being almost twofold increased (P < 0.05) in all experimental points in CD26-/- mice compared to wild-type indicating the expected up-regulation in CD26-/- conditions. Surprisingly, dominantly intracellular DP8 and DP9 were found in abundance in serum. DP8/9 activity is decreased in the inflamed colon, whereas its contribution to the overall serum DPP IV/CD26-like activity is negligible, suggesting the importance of their extra-enzymatic functions. To summarize, CD induction generated gene, protein and enzymatic changes of DP8 and DP9 so their involvement in inflammation development and/or healing process is implicated, especially in CD26-/- , and the question of their subcellular localization should be revised.

Hepatocyte-secreted DPP4 in obesity promotes adipose inflammation and insulin resistance.

Obesity-induced metabolic disease involves functional integration among several organs via circulating factors, but little is known about crosstalk between liver and visceral adipose tissue (VAT). In obesity, VAT becomes populated with inflammatory adipose tissue macrophages (ATMs). In obese humans, there is a close correlation between adipose tissue inflammation and insulin resistance, and in obese mice, blocking systemic or ATM inflammation improves insulin sensitivity. However, processes that promote pathological adipose tissue inflammation in obesity are incompletely understood. Here we show that obesity in mice stimulates hepatocytes to synthesize and secrete dipeptidyl peptidase 4 (DPP4), which acts with plasma factor Xa to inflame ATMs. Silencing expression of DPP4 in hepatocytes suppresses inflammation of VAT and insulin resistance; however, a similar effect is not seen with the orally administered DPP4 inhibitor sitagliptin. Inflammation and insulin resistance are also suppressed by silencing expression of caveolin-1 or PAR2 in ATMs; these proteins mediate the actions of DPP4 and factor Xa, respectively. Thus, hepatocyte DPP4 promotes VAT inflammation and insulin resistance in obesity, and targeting this pathway may have metabolic benefits that are distinct from those observed with oral DPP4 inhibitors.

Chronic Psychological Stress Accelerates Vascular Senescence and Impairs Ischemia-Induced Neovascularization: The Role of Dipeptidyl Peptidase-4/Glucagon-Like Peptide-1-Adiponectin Axis.

BACKGROUND: Exposure to psychosocial stress is a risk factor for cardiovascular disease, including vascular aging and regeneration. Given that dipeptidyl peptidase-4 (DPP4) regulates several intracellular signaling pathways associated with the glucagon-like peptide-1 (GLP-1) metabolism, we investigated the role of DPP4/GLP-1 axis in vascular senescence and ischemia-induced neovascularization in mice under chronic stress, with a special focus on adiponectin -mediated peroxisome proliferator activated receptor-γ/its co-activator 1α (PGC-1α) activation. METHODS AND RESULTS: Seven-week-old mice subjected to restraint stress for 4 weeks underwent ischemic surgery and were kept under immobilization stress conditions. Mice that underwent ischemic surgery alone served as controls. We demonstrated that stress impaired the recovery of the ischemic/normal blood-flow ratio throughout the follow-up period and capillary formation. On postoperative day 4, stressed mice showed the following: increased levels of plasma and ischemic muscle DPP4 and decreased levels of GLP-1 and adiponectin in plasma and phospho-AMP-activated protein kinase α (p-AMPKα), vascular endothelial growth factor, peroxisome proliferator activated receptor-γ, PGC-1α, and Sirt1 proteins and insulin receptor 1 and glucose transporter 4 genes in the ischemic tissues, vessels, and/or adipose tissues and numbers of circulating endothelial CD31+/c-Kit+ progenitor cells. Chronic stress accelerated aortic senescence and impaired aortic endothelial sprouting. DPP4 inhibition and GLP-1 receptor activation improved these changes; these benefits were abrogated by adiponectin blocking and genetic depletion. CONCLUSIONS: These results indicate that the DPP4/GLP-1-adiponectin axis is a novel therapeutic target for the treatment of vascular aging and cardiovascular disease under chronic stress conditions.

Dipeptidyl Peptidase-4 Regulates Hematopoietic Stem Cell Activation in Response to Chronic Stress.

BACKGROUND: DPP4 (Dipeptidyl peptidase-4)-GLP-1 (glucagon-like peptide-1) and its receptor (GLP-1R) axis has been involved in several intracellular signaling pathways. The Adrß3 (ß3-adrenergic receptor)/CXCL12 (C-X-C motif chemokine 12) signal was required for the hematopoiesis. We investigated the novel molecular requirements between DPP4-GLP-1/GLP-1 and Adrß3/CXCL12 signals in bone marrow (BM) hematopoietic stem cell (HSC) activation in response to chronic stress. METHODS AND RESULTS: Male 8-week-old mice were subjected to 4-week intermittent restrain stress and orally treated with vehicle or the DPP4 inhibitor anagliptin (30 mg/kg per day). Control mice were left undisturbed. The stress increased the blood and brain DPP4 levels, the plasma epinephrine and norepinephrine levels, and the BM niche cell Adrß3 expression, and it decreased the plasma GLP-1 levels and the brain GLP-1R and BM CXCL12 expressions. These changes were reversed by DPP4 inhibition. The stress activated BM sca-1highc-KithighCD48lowCD150high HSC proliferation, giving rise to high levels of blood leukocytes and monocytes. The stress-activated HSC proliferation was reversed by DPP4 depletion and by GLP-1R activation. Finally, the selective pharmacological blocking of Adrß3 mitigated HSC activation, accompanied by an improvement of CXCL12 gene expression in BM niche cells in response to chronic stress. CONCLUSIONS: These findings suggest that DPP4 can regulate chronic stress-induced BM HSC activation and inflammatory cell production via an Adrß3/CXCL12-dependent mechanism that is mediated by the GLP-1/GLP-1R axis, suggesting that the DPP4 inhibition or the GLP-1R stimulation may have applications for treating inflammatory diseases.

Combination of exendin-4 and DPP-4 silencing promoted angiogenesis of human coronary artery endothelial cells via activation of PI3K/Akt pathway.

This study was aimed to explore the combined effects of Exendin-4 with dipeptidyl peptidase-IV (DPP-4) silencing on vascular endothelial growth factor (VEGF)-induced cell proliferation and angiogenesis in Human Coronary Artery Endothelial Cells (HCAECs), as well as the underlying molecular mechanisms which were involved in this process. HCAECs were treated by exendin-4, small interfering RNA (siRNA) targeting DPP-4 (DPP-4-siRNA) or exendin-4 plus DPP-4-siRNA, respectively. Cell migration, proliferation and angiogenesis in vitro were assessed by scratch-wound assay, MTT, tran swell assay, and matrigel tube formation, respectively. Cell apoptosis was investigated by TUNEL assay. Expression of apoptosis and PI3K/Akt related proteins were assessed by Western blotting. Incubation of HCAECs with exendin-4 and silencing of DPP-4 both caused an increase in cell proliferation, migration and tube formation, while a significant decrease in apoptosis (all p<0.05). Furthermore, the combination of the exendin-4 and silencing of DPP-4 had additional effects on HCAECs. Protein levels of p-Akt and p-PI3K were markedly increased by exendin-4 incubation, silencing of DPP-4 in HCAECs. These results suggest that combination of exendin-4 and silencing of DPP-4 had additional promoted effects on angiogenesis of HCAECs via activation of PI3K/Akt pathway. Our study indicated an alternative therapeutic strategy for atherosclerotic neovascularization.

A possible role for CD26/DPPIV enzyme activity in the regulation of psoriatic pruritus.

BACKGROUND: Psoriasis (PSO) is one of the most common chronic inflammatory skin diseases, and pruritus affects approximately 60-90% of patients with PSO. However, the pathogenesis of pruritus in PSO remains unclear. Dipeptidyl peptidase IV (DPPIV) enzyme activity is involved in the regulation of peptide hormones, chemokines and neurotransmitters. OBJECTIVES: Our aim is to evaluate for a potential association between DPPIV and an increased risk of pruritus, and to identify possible underlying treatment targets in affected patients. METHODS: Utilizing clinical serum samples of PSO patients and in vivo experimental pruritus models, we evaluated for a potential association between DPPIV and an increased risk for pruritus, and attempted to identify possible underlying treatment targets in pruritus of PSO. RESULTS: We first showed that levels of DPPIV enzyme activity in sera of patients with PSO were significantly increased compared to those of healthy controls. We next evaluated levels of substance-P (SP), which is a neurotransmitter for pruritus and a substrate for DPPIV enzyme. Truncated form SP cleaved by DPPIV was significantly increased in sera of PSO. In an in vivo pruritus model induced by SP, scratching was decreased by treatment with a DPPIV inhibitor. Moreover, DPPIV-knockout mice showed attenuation of scratching induced by SP. Finally, scratching was decreased following the administration of a DPPIV inhibitor in an imiquimod-induced PSO model. On the other hand, scratching induced by imiquimod was increased in DPPIV overexpressing-mice. CONCLUSIONS: These results suggest that inhibition of DPPIV enzyme activity regulates pruritus in PSO.

Glucagon-like peptide-1 receptor signalling reduces microvascular thrombosis, nitro-oxidative stress and platelet activation in endotoxaemic mice.

BACKGROUND AND PURPOSE: Excessive inflammation in sepsis causes microvascular thrombosis and thrombocytopenia associated with organ dysfunction and high mortality. The present studies aimed to investigate whether inhibition of dipeptidyl peptidase-4 (DPP-4) and supplementation with glucagon-like peptide-1 (GLP-1) receptor agonists improved endotoxaemia-associated microvascular thrombosis via immunomodulatory effects. EXPERIMENTAL APPROACH: Endotoxaemia was induced in C57BL/6J mice by a single injection of LPS (17.5 mg kg-1 for survival and 10 mg kg-1 for all other studies). For survival studies, treatment was started 6 h after LPS injection. For all other studies, drugs were injected 48 h before LPS treatment. KEY RESULTS: Mice treated with LPS alone showed severe thrombocytopenia, microvascular thrombosis in the pulmonary circulation (fluorescence imaging), increased LDH activity, endothelial dysfunction and increased markers of inflammation in aorta and whole blood (leukocyte-dependent oxidative burst, nitrosyl-iron haemoglobin, a marker of nitrosative stress, and expression of inducible NOS). Treatment with the DPP-4 inhibitor linagliptin or the GLP-1 receptor agonist liraglutide, as well as genetic deletion of DPP-4 (DPP4-/- mice) improved all these parameters. In GLP-1 receptor-deficient mice, both linagliptin and liraglutide lost their beneficial effects and improvement of prognosis. Incubation of platelets and cultured monocytes (containing GLP-1 receptor protein) with GLP-1 receptor agonists inhibited the monocytic oxidative burst and platelet activation, with a GLP-1 receptor-dependent elevation of cAMP levels and PKA activation. CONCLUSIONS AND IMPLICATIONS: GLP-1 receptor activation in platelets by linagliptin and liraglutide strongly attenuated endotoxaemia-induced microvascular thrombosis and mortality by a cAMP/PKA-dependent mechanism, preventing systemic inflammation, vascular dysfunction and end organ damage. LINKED ARTICLES: This article is part of a themed section on Redox Biology and Oxidative Stress in Health and Disease. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v174.12/issuetoc.

Involvement of DPP IV/CD26 in cutaneous wound healing process in mice.

Dipeptidyl peptidase IV (DPP IV/CD26) is a widely distributed multifunctional protein that plays a significant role in different physiological as well as pathological processes having a broad spectrum of bioactive substrates and immunomodulative properties. It has potential influence on different processes crucial for wound healing, including cell adhesion, migration, apoptosis, and extracellular matrix degradation. However, despite its known enzymatic and immunomodulative functions, limited data characterize the role of DPP IV/CD26 in cutaneous wound healing mechanisms. The aim of this study was to investigate the process of wound healing in conditions of CD26 deficiency in order to obtain better insights on the role of DPP IV/CD26 in cutaneous regeneration. Experimental wounds were made on the dorsal part of CD26 deficient (CD26-/- ) and wild-type mice (C57BL/6). The process of cutaneous wound healing was monitored on defined time schedule postwounding by macroscopic, microscopic, and biochemical analyses. Obtained results revealed a better rate of wound closure, revascularization and cell proliferation in CD26-/- mice, with enhanced local expression of hypoxia-inducible factor 1α and vascular endothelial growth factor. CD26 deficiency induced prompt macrophage recruitment at the site of skin damage but did not influence mobilization of T-cells in comparison with wild-type mice. CD26-/- mice have significantly higher values of IP-10 in serum and control skins compared with wild-type mice but values in wounds did not differ significantly on days 2, 4, and 7 of wound healing. DPP IV/CD26 activity was found to be decreased 4 days postwounding in serum and 2, 4, and 7 days postwounding in wounds of wild-type animals compared with control skins. These findings contribute to better understanding of wound healing mechanisms and could give a support in finding new therapeutic approaches for wound healing and tissue regeneration.