Leukocyte Membrane Enzymes Play the Cell Adhesion Game.

For a long time, proteins with enzymatic activity have not been usually considered to carry out other functions different from catalyzing chemical reactions within or outside the cell. Nevertheless, in the last few years several reports have uncovered the participation of numerous enzymes in other processes, placing them in the category of moonlighting proteins. Some moonlighting enzymes have been shown to participate in complex processes such as cell adhesion. Cell adhesion plays a physiological role in multiple processes: it enables cells to establish close contact with one another, allowing communication; it is a key step during cell migration; it is also involved in tightly binding neighboring cells in tissues, etc. Importantly, cell adhesion is also of great importance in pathophysiological scenarios like migration and metastasis establishment of cancer cells. Cell adhesion is strictly regulated through numerous switches: proteins, glycoproteins and other components of the cell membrane. Recently, several cell membrane enzymes have been reported to participate in distinct steps of the cell adhesion process. Here, we review a variety of examples of membrane bound enzymes participating in adhesion of immune cells.

Inhibition of dipeptidyl peptidase-4 averts free fatty acids deposition in the hearts of oral estrogen-progestin contraceptive-induced hyperinsulinemic female rats.

Free fatty acid (FFA) deposition in non-adipose tissues such as the heart is a characteristic of insulin resistant states which feature hyperinsulinemia and dipeptidyl peptidase-4 (DPP-4) activation. Estrogen-progestin oral contraceptives (OC) treatment reportedly increased DPP-4 activity in rat tissue, and DPP-4 inhibitors have anti-diabetic and anti-inflammatory properties. This study aims to investigate the effects of DPP-4 inhibition on cardiac FFA deposition in estrogen-progestin-treated female rats. From our data, estrogen-progestin OC exposure in female rats led to elevated plasma insulin, cardiac DPP-4 activity, FFA and triglyceride (TG) accumulation, TG/high-density lipoprotein cholesterol (TG/HDL-C) ratio, adenosine deaminase/xanthine oxidase/uric acid pathway (ADA/XO/UA), lipid peroxidation, glycogen synthase activity, and alanine phosphatase; whereas cardiac glucose-6-phosphate dehydrogenase, Na+/K+-ATPase and nitric oxide (NO) were decreased. However, DPP-4 inhibition resulted in decreased plasma insulin, cardiac DPP-4 activity, FFA, TG, TG/HDL-C ratio, and alkaline phosphatase. These were accompanied by reduced ADA/XO/UA pathway, lipid peroxidation, and augmented NO and Na+/K+-ATPase in estrogen-progestin OC-treated rats. DPP-4 inhibition attenuated cardiac lipid deposition accompanied by reduced activity in the ADA/XO/UA pathway in estrogen-progestin OC-treated female rats. DPP-4 is therefore a plausible therapeutic target in cardiometabolic disorders.

DPP4 Activity, Hyperinsulinemia, and Atherosclerosis.

CONTEXT: Obesity and type 2 diabetes are associated with chronic hyperinsulinemia, elevated plasma levels of dipeptidyl peptidase-4 (DPP4), and a pro-atherosclerotic milieu. EVIDENCE ACQUISITION: PubMed search of the term "insulin and atherosclerosis," "hyperinsulinemia," "atherosclerosis," or "cardiovascular outcomes" cross-referenced with "DPP4." Relevant research and review articles were reviewed. EVIDENCE SYNTHESIS: Hyperinsulinemia in the setting of insulin resistance promotes vascular inflammation, vascular smooth muscle cell growth, pathological cholesterol profile, hypertension, and recruitment of immune cells to the endothelium, all contributing to atherosclerosis. DPP4 has pleiotropic functions and its activity is elevated in obese humans. DPP4 mirrors hyperinsulinemia's atherogenic actions in the insulin resistant state, and genetic deletion of DPP4 protects rodents from developing insulin resistance and improves cardiovascular outcomes. DPP4 inhibition in pro-atherosclerotic preclinical models results in reduced inflammation and oxidative stress, improved endothelial function, and decreased atherosclerosis. Increased incretin levels may have contributed to but do not completely account for these benefits. Small clinical studies with DPP4 inhibitors demonstrate reduced carotid intimal thickening, improved endothelial function, and reduced arterial stiffness. To date, this has not been translated to cardiovascular risk reduction for individuals with type 2 diabetes with prior or exaggerated risk of cardiovascular disease. CONCLUSION: DPP4 may represent a key link between central obesity, insulin resistance, and atherosclerosis. The gaps in knowledge in DPP4 function and discrepancy in cardiovascular outcomes observed in preclinical and large-scale randomized controlled studies with DPP4 inhibitors warrant additional research.

The mechanism of sitagliptin inhibition of colorectal cancer cell lines' metastatic functionalities.

The cell membrane glycoprotein CD26 with peptidase activity (DPP4) and/or its soluble CD26/DPP4 counterpart expression and/or activity are altered in several cancers. Its role in metastasis development was recently highlighted by the discovery of CD26+ cancer stem cell subsets and the fact that clinical DPP4 inhibitors showed antimetastatic effects in animal models. Also, diabetic patients treated with the DPP4 inhibitor sitagliptin showed greater overall survival after colorectal or lung cancer surgery than patients under other diabetic therapies. However, the mechanism of action of these inhibitors in this context is unclear. We studied the role of CD26 and its DPP4 enzymatic activity in malignant cell features such as cell-to-cell homotypic aggregation, cancer cell motility, and invasion in a panel of human colorectal cancer (CRC) cell lines, avoiding models that include the physiological role of DPP4 in chemotaxis. Present results indicate that CD26 participates in the induction of cell invasion, motility, and aggregation of CD26-positive CRC cell lines. Moreover, only invasion and motility assays, which are collagen matrix-dependent, showed a decrease upon treatment with the DPP4 inhibitor sitagliptin. Sitagliptin showed opposite effects to those of transforming growth factor-ß1 on epithelial-to-mesenchymal transition and cell cycle, but this result does not explain its CD26/DPP4-dependent effect. These results contribute to the elucidation of the molecular mechanisms behind sitagliptin inhibition of metastatic traits. At the same time, this role of sitagliptin may help to define areas of medicine where DPP4 inhibitors might be introduced. However, they also suggest that additional tools against CD26 as a target might be used or developed for metastasis prevention in addition to gliptins.

Role of Milk-Derived Opioid Peptides and Proline Dipeptidyl Peptidase-4 in Autism Spectrum Disorders.

Opioid peptides released during digestion of dietary proteins such as casein, were suggested to contribute to autism development, leading to the announcement of opioid excess hypothesis of autism. This paper examines role of enzyme proline dipeptidyl peptidase-4 (DPPIV; EC 3.4.14.5) and it is exogenous substrate, ß-casomorphin-7 (BCM7) in autism etiology. Our study included measurements of DPPIV and BCM7 concentrations in serum and urine, which were analyzed with ELISA assays and activity of DPPIV was measured by colorimetric test. The effect of opioid peptides from hydrolysed bovine milk on DPPIV gene expression in peripheral blood mononuclear cells (PBMC) in autistic and healthy children was determined using the Real-Time PCR (Polymerase Chain Reaction) method. Our research included 51 healthy children and 86 children diagnosed with autism spectrum disorder (ASD, ICDF84). We determined that the concentration of BCM7 in serum was significantly, 1.6-fold, higher in the ASD group than in controls (p < 0.0001). Concentration of DPPIV was found to also be significantly higher in serum from ASD children compared to the control group (p < 0.01), while we did not notice significant difference in enzymatic activity of serum DPPIV between the two study groups. We confirmed correlation according to the gender between analyzed parameters. The inspiration for this study emanated from clinical experience of the daily diet role in relieving the symptoms of autism. Despite this, we have concluded that milk-derived opioid peptides and DPPIV are potentially factors in determining the pathogenesis of autism; conducted studies are still limited and require further research.

Biological function of dipeptidyl peptidase-4 on type 2 diabetes patients and diabetic mice.

BACKGROUND: Type 2 diabetes (TD2) is a sustained metabolic disorder, characterized by high blood glucose, insulin resistance (IR). Dipeptidyl peptidase-4 (DPP4) functions as an antigenic enzyme involved in hyperglycaemia, oxidative stress, and inflammation-associated IR. Therefore, association between DPP4 and TD2 warrants to be investigated. METHODS: In this study, blood samples of clinically diagnosed TD2 patients were harvested for biochemical tests. In addition, diabetic mice induced by high-fat diet (HFD) and single dose of streptozotocin (STZ) were used to assess the biological characteristics of DPP4 through biochemical and enzyme-linked immunosorbent assay (ELISA) tests, immunofluorescence staining, and western blot assay. RESULTS: Compared to controls, the clinical data of patients with TD2 resulted in increased contents of fasting blood glucose (FBG), glycated hemoglobin (HbA1c), homeostatic model assessment (HOMA)-IR, blood lipids of triglyceride (TG), total cholesterol (TC), low-density lipoprotein (LDL-C), and interleukin 6 (IL6) in plasma samples (p < 0.05). Notably, blood levels of DPP4 in TD2 patients were increased significantly in comparison to that in non-diabetic adults (p < 0.01). In animal study, diabetic mice showed increased levels of glucose, insulin, lipids, DPP4 activity in sera. Visibly, hepatocellular DPP4 expression was up-regulated in diabetic mice. Interestingly, DPP4 inhibitor-treated mice showed significantly reduced DPP4 expression in serum (p < 0.01), and lowered DPP4-positive cells and protein content in the liver were observed when compared to those in diabetic mice (p < 0.01). CONCLUSIONS: Collectively, these findings reveal that DPP4 biomolecule may be positively associated with TD2 development, and the underlying mechanism may be attributed to activation of DPP4 expression in liver cells.

Circulating Levels of Soluble Dipeptidyl Peptidase-4 Are Dissociated from Inflammation and Induced by Enzymatic DPP4 Inhibition.

Dipeptidyl peptidase-4 (DPP-4) controls glucose homeostasis through enzymatic termination of incretin action. We report that plasma DPP-4 activity correlates with body weight and fat mass, but not glucose control, in mice. Genetic disruption of adipocyte Dpp4 expression reduced plasma DPP-4 activity in older mice but did not perturb incretin levels or glucose homeostasis. Knockdown of hepatocyte Dpp4 completely abrogated the obesity-associated increase in plasma DPP-4 activity, reduced liver cytokine expression, and partially attenuated inflammation in adipose tissue without changes in incretin levels or glucose homeostasis. In contrast, circulating levels of soluble DPP4 (sDPP4) were dissociated from inflammation in mice with endothelial-selective or global genetic inactivation of Dpp4. Remarkably, inhibition of DPP-4 enzymatic activity upregulated circulating levels of sDPP4 originating from endothelial or hematopoietic cells without inducing systemic or localized inflammation. Collectively, these findings reveal unexpected complexity in regulation of soluble versus enzymatic DPP-4 and control of inflammation and glucose homeostasis.

Pharmacologic targeting of the diabetic stem cell mobilopathy.

Diabetes is a chronic metabolic disease characterized by hyperglycemia and several associated biochemical abnormalities. Diabetes leads to multiorgan complications that collectively reduce life expectancy. Hematopoietic stem cells (HSCs) are nested within bone marrow (BM) niches whence they can be mobilized to the peripheral circulation. Clinically, this is done for HSC collection and autologous or allogenic transplantation. A great amount of data from basic and clinical studies support that diabetic patients are poor HSC mobilizers owing to BM remodeling. Dysfunction of the BM shares pathophysiological features and pathways with typical chronic diabetic complications that affect other issues (e.g. the retina and the kidney). From a clinical perspective, impaired HSC mobilization translates into the failure to collect a minimum number of CD34+ cells to achieve a safe engraftment after transplantation. Furthermore, blunted mobilization is associated with reduced steady-state levels of circulating HSCs, which have been consistently described in diabetic patients and associated with increased risk of adverse outcomes, including cardiovascular events and death. In this review, we discuss the most clinically relevant pharmacological options to overcome impaired HSC mobilization in diabetes. These therapeutic strategies may result in an improved outcome of diabetic patients undergoing HSC transplantation and restore circulating HSC levels, thereby protecting from adverse cardiovascular outcomes.

The effects of dipeptidyl peptidase-4 on cardiac fibrosis in pressure overload-induced heart failure.

Dipeptidyl peptidase-4 (DPP-4) inhibitors are hypoglycemic agents. DPP-4 inhibitor has cardioprotective effects after transverse aortic constriction (TAC), but role of DPP-4 on cardiac fibrosis after TAC is not well known. Our aim was to determine the effects of DPP-4 on cardiac fibrosis in murine TAC model. Wild-type mice and DPP-4 knockout mice were subjected to TAC. Wild-type mice were then treated with vehicle or DPP-4 inhibitor. DPP-4 activities in serum and heart tissue were significantly increased at 2 weeks after TAC, but they were significantly decreased by DPP-4 inhibitor treatment. The inhibition of DPP-4 did not affect left ventricular hypertrophy, but improved cardiac function and decreased myocardial and perivascular fibrosis after TAC. The inhibition of DPP-4 decreased the collagen type III/I ratio in myocardium. These results suggest that DPP-4 inhibition ameliorates the progression of heart failure after TAC by changing the quality and quantity of cardiac fibrosis.

Association of plasma dipeptidyl peptidase-4 activity with non-alcoholic fatty liver disease in nondiabetic Chinese population.

OBJECTIVE: The pathogenesis of non-alcoholic fatty liver disease (NAFLD) is attributed to a "multi-hits hypothesis" involving insulin resistance, oxidative stress and inflammation. Dipeptidyl peptidase-4 (DPP4) was identified as a novel adipokine capable of enhancing the"multi-hits". Hence, we investigated the association between plasma DPP4 activity and NAFLD in nondiabetic Chinese population. DESIGN AND METHODS: We performed a cross-sectional study using data from 1105 subjects (36-79years) in Guilin between 2015 and 2016. Plasma DPP4 activity, homeostatic model assessment of insulin resistance (HOMA-IR), oxidative stress parameters, and inflammatory markers were measured in all participants. NAFLD and its severity were diagnosed by ultrasound after the exclusion of alcohol abuse and other liver diseases. RESULTS: Participants in the highest quartile of DPP4 activity had higher HOMA-IR, nitrotyrosine, 8-iso-PGF2a, interleukin-6, CRP, alanine aminotransferase, aspartate aminotransferase and γ-glutamyltransferase compared with those in the lowest quartile (all P<0.05). Plasma DPP4 activity gradually increased across the groups according to the ultrasonographic severity of steatosis (P<0.001 for the trend). In the highest DPP4 quartile, NAFLD risk was higher (odds ratio 1.88; 95% CI 1.04-3.37) than in the lowest quartile after adjustment for confounders. The risk for NAFLD increased more with higher levels of DPP4 activity, HOMA-IR, nitrotyrosine, 8-iso-PGF2a, interleukin-6 and CRP. CONCLUSIONS: Plasma DPP4 activity is significantly associated with NAFLD. The underlying mechanisms may be partly attributed to the interactions between insulin resistance, oxidative stress, inflammation, and DPP4.

Heterogeneity of molecular forms of dipeptidyl peptidase-IV and fibroblast activation protein in human glioblastomas.

BACKGROUND AND AIMS: Proteolytic enzymes contribute to the progression of various cancers. We previously reported increased expression of the proline specific peptidases dipeptidyl peptidase-IV (DPP-IV) and its closest paralogue fibroblast activation protein (FAP) in human glioblastomas. Here we analyze the molecular heterogeneity of DPP-IV and FAP in glioblastomas. METHODS: ELISA, isoelectric focusing, 1D and 2D electrophoresis followed by WB or enzyme overlay assay were utilized to analyze DPP-IV and FAP isoforms. Cell fractionation using a Percoll gradient and deglycosylation with PNGase F were performed to analyze the possible basis of DPP-IV and FAP microheterogeneity. RESULTS: Molecular forms of DPP-IV with an estimated molecular weight of 140-160 kDa and a pI predominantly 5.8 were detected in human glioblastoma; in some tumors additional isoforms with a more acidic (3.5-5.5) as well as alkaline (8.1) pI were revealed. Using 2D electrophoresis, two to three molecular forms of FAP with an alkaline (7.0-8.5) pI and an estimated MW of 120-140 kDa were identified in glioblastoma tissues. In glioma cell lines in vitro, several isoforms of both enzymes were expressed, however the alkalic forms present in glioblastoma tissues were not detected. Removal of N-linked oligosaccharides decreased the estimated molecular weight of both enzymes; the overall pattern of molecular forms nevertheless remained unchanged. CONCLUSION: Several isoforms of DPP-IV and FAP are present in glioblastoma tissue. The absence of alkaline isoforms of both enzymes in glioma cell lines however suggests that isoforms from other, most likely stromal, cell types contribute to the overall pattern seen in glioblastoma tissues.

Association of DPP-4 activity with BMD, body composition, and incident hip fracture: the Cardiovascular Health Study.

There was no association of plasma DPP-4 activity levels with bone mineral density (BMD), body composition, or incident hip fractures in a cohort of elderly community-dwelling adults. INTRODUCTION: Dipeptidyl peptidase IV (DPP-4) inactivates several key hormones including those that stimulate postprandial insulin secretion, and DPP-4 inhibitors (gliptins) are approved to treat diabetes. While DPP-4 is known to modulate osteogenesis, the relationship between DPP-4 activity and skeletal health is uncertain. The purpose of the present study was to examine possible associations between DPP-4 activity in elderly subjects enrolled in the Cardiovascular Health Study (CHS) and BMD, body composition measurements, and incident hip fractures. METHODS: All 1536 male and female CHS participants who had evaluable DXA scans and plasma for DPP-4 activity were included in the analyses. The association between (1) BMD of the total hip, femoral neck, lumbar spine, and total body; (2) body composition measurements (% lean, % fat, and total body mass); and (3) incident hip fractures and plasma levels of DPP-4 activity were determined. RESULTS: Mean plasma levels of DPP-4 activity were significantly higher in blacks (227 ± 78) compared with whites (216 ± 89) (p = 0.04). However, there was no significant association of DPP-4 activity with age or gender (p ≥ 0.14 for both). In multivariable adjusted models, there was no association of plasma DPP-4 activity with BMD overall (p ≥ 0.55 for all) or in gender stratified analyses (p ≥ 0.23). There was also no association of DPP-4 levels and incident hip fractures overall (p ≥ 0.24) or in gender stratified analyses (p ≥ 0.39). CONCLUSION: Plasma DPP-4 activity, within the endogenous physiological range, was significantly associated with race, but not with BMD, body composition, or incident hip fractures in elderly community-dwelling subjects.

[Role of dipeptidyl peptidase-4 and its inhibitor in the respiratory diseases].

Dipeptidyl peptidase-4 (DPP-4) is known as a highly conserved type II transmembraneous glycoprotein widely distributed in a variety of tissues and cells, and expressed in the peripheral blood as a soluble form. It has been reported that DPP4 play a distinct role in the physiological and pathological processes, such as immune regulation, inflammatory reaction, cell adhesion, and cell apoptosis. DPP4 inhibitor showes an incredible effect on the control of blood glucose and it is thought as a newly-developed drug for diabetes, especially in regulation of post-prandial glycemia. It has been reported that DPP4 plays a potential role in many respiratory diseases, especially in the pathogenesis of asthma.

Role of IL-26+CD26+CD4 T Cells in Pulmonary Chronic Graft-Versus-Host Disease and Treatment with Caveolin-1-Ig Fc Conjugate.

Obliterative bronchiolitis is the primary noninfectious pulmonary complication after allogeneic hematopoietic cell transplantation and the only pathognomonic manifestation of pulmonary chronic graft-versus-host disease (cGVHD). In our recent study, we identified a novel effect of IL-26, which is absent in rodents, on transplant related-obliterative bronchiolitis. Sublethally irradiated NOD/Shi-scidIL2rγnull mice transplanted with human umbilical cord blood gradually exhibited obliterative bronchiolitis with increased collagen deposition and predominant infiltration with human IL-26+CD26+CD4 T cells. Moreover, we showed that IL-26 increased collagen synthesis in fibroblasts in vitro and that collagen contents were increased in a murine GVHD model using IL26 transgenic mice. In vitro analysis demonstrated a significant increase in IL-26 production by CD4 T cells following CD26 costimulation, while immunoglobulin Fc domain fused with the N-terminal of caveolin-1, the ligand for CD26, (Cav-Ig) effectively inhibited production of IL-26. Administration of Cav-Ig before or after onset of GVHD impeded the development of clinical and histologic features of GVHD without interrupting engraftment of donor-derived human cells, with preservation of the graft-versus-leukemia effect. We concluded that cGVHD of the lungs is caused in part by IL-26+CD26+CD4 T cells, and that treatment with Cav-Ig could be beneficial for cGVHD prevention and therapy.

The role of bradykinin in lung ischemia-reperfusion injury in a rat lung transplantation model

ABSTRACT PURPOSE: To investigate the role of bradykinin in a rat lung transplantation (LTx) model and preliminarily discuss the relationship between bradykinin and CD26/DPP-4. METHODS: Rats were randomly divided into four groups: Control (CON), Sham, low potassium dextranglucose (LPD), and AB192 (n=15/group). Orthotopic single LTx was performed in the LPD and AB192 groups. The donor lungs were flush-perfused and preserved with low potassium dextranglucose (LPD) or LPD+CD26/DPP-4 catalytic inhibitor (AB192). LTx was performed after 18 h cold ischemia time and harvested two days post-LTx. Blood gas analysis (PO2), wet/dry weight ratio (W/D), myeloperoxidase activity (MPO), and lipid peroxidation (MDA) were analyzed at 48 hr after transplantation. Immunohistochemical (IHC) analysis was performed in the same sample and validated by Western-Blot. RESULTS: Compared to the LPD group, the AB192 group showed higher PO2, lower W/D ratio, and decreased MPO and MDA. IHC studies showed strong bradykinin β2 receptor (B2R) staining in the LPD group, especially in inflammatory cells, alveolar macrophages, and respiratory epithelial cells. Expression of B2R by Western-Blot was significantly different between the AB192 and LPD groups. CONCLUSION: Bradykinin may be a competitive substrate of DPP-4, and decreased bradykinin levels may enhance protective effects against ischemia/reperfusion injury during LTx.

Dipeptidyl Peptidase-4, Wound Healing, Scarring, and Fibrosis.

Scarring and fibrosis are an enormous public health concern, resulting in excessive morbidity and mortality in addition to countless lost health care dollars. Recent advances in cell and developmental biology promise a better understanding of scarring and fibrosis and may translate to new clinical therapies.

Inhibition of DPP-4 by alogliptin improves coronary flow reserve and left ventricular systolic function evaluated by phase contrast cine magnetic resonance imaging in patients with type 2 diabetes and coronary artery disease.

BACKGROUND: The present study determined whether dipeptidyl peptidase-4 (DPP-4) inhibition by alogliptin improves coronary flow reserve (CFR) and left ventricular election fraction (LVEF) in patients with type 2 DM and CAD. MATERIALS AND METHODS: Twenty patients with type 2 DM and known or suspected CAD were randomly allocated to receive diet therapy plus alogliptin (n=10; mean age, 73.3±6.6y) or a control group given diet therapy and glimepiride (n=10; mean age, 76.7±7.3y). Breath-hold PC cine MR images of the coronary sinus (CS) were acquired using a 1.5T MR scanner and 32 channel cardiac coils to assess blood flow of the CS at rest and during adenosine triphosphate (ATP) infusion. The CFR was calculated as CS blood flow during ATP infusion divided by that at rest. The CFR and LVEF were evaluated by MRI at baseline and at three months after starting therapy. RESULTS: Hemoglobin A1c (HbA1c) was significantly reduced in both groups (alogliptin, 7.2±0.6% to 6.6±0.5%, p=0.034; control, 6.9±0.4% to 6.4±0.3%, p=0.008). However, CFR and LVEF significantly improved only in the alogliptin group (alogliptin: CFR, 2.15±0.61 to 2.85±0.80, p=0.042; LVEF, 59.4±6.3% to 68.0±8.6%, p=0.03; control: CFR, 2.17±0.37 to 2.38±0.32, p=0.19; LVEF, 58.2±9.1 to 60.3±8.8%, p=0.61). The % increases in CFR and in LVEF positively correlated (R=0.47 by Spearman's correlation coefficient; p=0.036). CONCLUSION: The inhibition of DPP-4 by alogliptin improved CFR and LVEF evaluated by MRI in patients with type 2 DM and CAD and the improvement in CFR was associated with increased LV systolic function.

Modulation of Hematopoietic Chemokine Effects In Vitro and In Vivo by DPP-4/CD26.

Dipeptidyl peptidase 4 (DPP4)/CD26 truncates certain proteins, and this posttranslational modification can influence their activity. Truncated (T) colony-stimulating factors (CSFs) are decreased in potency for stimulating proliferation of hematopoietic progenitor cells (HPCs). T-CXCL12, a modified chemokine, is inactive as an HPC chemotactic, survival, and enhancing factor for replating or ex-vivo expansion of HPCs. Moreover, T-CSFs and T-CXCL12 specifically downmodulates the positively acting effects of their own full-length molecule. Other chemokines have DPP4 truncation sites. In the present study, we evaluated effects of DPP4 inhibition (by Diprotin A) or gene deletion of HPC on chemokine inhibition of multicytokine-stimulated HPC, and on chemokine-enhancing effects on single CSF-stimulated HPC proliferation, as well as effects of DPP4 treatment of a number of chemokines. Myelosuppressive effects of chemokines with, but not without, a DPP4 truncation site were greatly enhanced in inhibitory potency by pretreating target bone marrow (BM) cells with Diprotin A, or by assaying their activity on dpp4/cd26(-/-) BM cells. DPP4 treatment of myelosuppressive chemokines containing a DPP4 truncation site produced a nonmyelosuppressive molecule, but one which had the capacity to block suppression by that unmodified chemokine both in vitro and in vivo. Additionally, DPP4 treatment ablated the single cytokine-stimulated HPC-enhancing activity of CCL3/MIP-1α and CCL4/MIP-1ß, and blocked the enhancing activity of each unmodified molecule, in vitro and in vivo. These results highlight the functional posttranslational modulating effects of DPP4 on chemokine activities, and information offering additional biological insight into chemokine regulation of hematopoiesis.

Soluble DPP-4 up-regulates toll-like receptors and augments inflammatory reactions, which are ameliorated by vildagliptin or mannose-6-phosphate.

OBJECTIVE: Studies have shown that dipeptidyl peptidase-4 (DPP-4) inhibitors have anti-inflammatory effects. Soluble DPP-4 (sDPP-4) has been considered as an adipokine of which actions need to be further characterized. METHODS: We investigated the pro-inflammatory actions of sDPP-4 and the anti-inflammatory effects of DPP-4 inhibition, using vildagliptin, as an enzymatic inhibitor, and mannose-6-phosphate (M6P) as a competitive binding inhibitor. RESULTS: In lipopolysaccharide (LPS)-stimulated RAW264.7 cells, vildagliptin suppressed the increased expression of inducible nitric oxide synthase (iNOS) and phosphorylated JNK (pJNK), activation of the NF-κB pathway, and the resultant NO and proinflammatory cytokine production. Although sDPP-4 alone did not affect the protein level of iNOS or pJNK or the production of NO in RAW264.7 cells, it did amplify iNOS expression, NO responses, and proinflammatory cytokine production in LPS-stimulated RAW264 cells. As a probable mechanism, we found that sDPP-4 caused dose-dependent increases in the expression levels of toll-like receptor 4 (TLR4) and TLR2 in RAW264.7 cells, and that these alterations were inhibited by vildagliptin, M6P, or bisindolylmaleimide II, a protein kinase C inhibitor. Either vildagliptin or M6P suppressed iNOS expression and NO and cytokine production in LPS+DPP-4-co-stimulated macrophages, while combined treatment of the co-stimulated cells with both agents had increased anti-inflammatory effects compared with either treatment alone. Intravenous injection of sDPP-4 to C57BL/6J mice increased the expression of both TLRs in kidney and white adipose tissues. CONCLUSION: Our findings suggest that sDPP-4 enhances inflammatory actions via TLR pathway, while DPP-4 inhibition with either an enzymatic or binding inhibitor has anti-inflammatory effects.

The role of bradykinin in lung ischemia-reperfusion injury in a rat lung transplantation model.

PURPOSE:: To investigate the role of bradykinin in a rat lung transplantation (LTx) model and preliminarily discuss the relationship between bradykinin and CD26/DPP-4. METHODS:: Rats were randomly divided into four groups: Control (CON), Sham, low potassium dextranglucose (LPD), and AB192 (n=15/group). Orthotopic single LTx was performed in the LPD and AB192 groups. The donor lungs were flush-perfused and preserved with low potassium dextranglucose (LPD) or LPD+CD26/DPP-4 catalytic inhibitor (AB192). LTx was performed after 18 h cold ischemia time and harvested two days post-LTx. Blood gas analysis (PO2), wet/dry weight ratio (W/D), myeloperoxidase activity (MPO), and lipid peroxidation (MDA) were analyzed at 48 hr after transplantation. Immunohistochemical (IHC) analysis was performed in the same sample and validated by Western-Blot. RESULTS:: Compared to the LPD group, the AB192 group showed higher PO2, lower W/D ratio, and decreased MPO and MDA. IHC studies showed strong bradykinin ß2 receptor (B2R) staining in the LPD group, especially in inflammatory cells, alveolar macrophages, and respiratory epithelial cells. Expression of B2R by Western-Blot was significantly different between the AB192 and LPD groups. CONCLUSION:: Bradykinin may be a competitive substrate of DPP-4, and decreased bradykinin levels may enhance protective effects against ischemia/reperfusion injury during LTx.