Jochen G. Schneider

Loss of pain perception in diabetes is dependent on a receptor of the immunoglobulin superfamily

Molecular events that result in loss of pain perception are poorly understood in diabetic neuropathy. Our results show that the receptor for advanced glycation end products (RAGE), a receptor associated with sustained NF-kappaB activation in the diabetic microenvironment, has a central role in sensory neuronal dysfunction. In sural nerve biopsies, ligands of RAGE, the receptor itself, activated NF-kappaBp65, and IL-6 colocalized in the microvasculature of patients with diabetic neuropathy. Activation of NF-kappaB and NF-kappaB-dependent gene expression was upregulated in peripheral nerves of diabetic mice, induced by advanced glycation end products, and prevented by RAGE blockade. NF-kappaB activation was blunted in RAGE-null (RAGE(-/-)) mice compared with robust enhancement in strain-matched controls, even 6 months after diabetes induction. Loss of pain perception, indicative of long-standing diabetic neuropathy, was reversed in WT mice treated with soluble RAGE. Most importantly, loss of pain perception was largely prevented in RAGE(-/-) mice, although they were not protected from diabetes-induced loss of PGP9.5-positive plantar nerve fibers. These data demonstrate, for the first time to our knowledge, that the RAGE-NF-kappaB axis operates in diabetic neuropathy, by mediating functional sensory deficits, and that its inhibition may provide new therapeutic approaches.

Elevated plasma endothelin-1 levels in diabetes mellitus ☆

BACKGROUND This study compares plasma endothelin-1 (ET-1) levels in patients with diabetes mellitus or hypertension with healthy controls, and investigates whether ET-1 levels are correlated with glycemic control, metabolic parameters, and vascular complications. METHODS The study population consisted of 103 patients with type 1 diabetes, 124 patients with type 2 diabetes, 35 hypertensive patients without diabetes mellitus, and 99 controls. RESULTS Plasma ET-1 concentrations were significantly higher in patients with type 1 diabetes (0.28 +/- 0.34 fmol/mL, P =.001), type 2 diabetes (0.31 +/- 0.32 fmol/mL, P <.0001), and hypertension (0.35 +/- 0.26 fmol/mL, P <.0001) compared to controls (0.08 +/- 0.13 fmol/mL). Diabetic patients taking angiotensin converting enzyme (ACE) inhibitors had significantly lower plasma ET-1 levels than patients without (0.22 +/- 0.20 fmol/mL v 0.38 +/- 0.39 fmol/mL, P =.029). There were significant associations between ET-1 levels and age (r = 0.38, P <.05) and systolic blood pressure (BP) (r = 0.27, P <.05) in healthy controls. In diabetes we found only nonsignificant associations between ET-1 levels and age or vascular complications and a weak association between plasma ET-1 levels and glycemic control. CONCLUSIONS Patients with diabetes or hypertension have elevated ET-1 levels, but do not exhibit positive correlations between ET-1 levels and BP, which was observed in healthy controls. Increased ET-1 levels do not induce hypertension in diabetes, but were lower in diabetic patients taking ACE inhibitors compared to those without ACE inhibitors. There is no significant association between ET-1 levels and vascular complications. These findings suggest that the plasma ET-1 level is not a marker of endothelial dysfunction but changes in plasma ET-1 levels may precede vascular complications associated with hypertension and diabetes.

From meta-omics to causality: experimental models for human microbiome research.

Large-scale ‘meta-omic’ projects are greatly advancing our knowledge of the human microbiome and its specific role in governing health and disease states. A myriad of ongoing studies aim at identifying links between microbial community disequilibria (dysbiosis) and human diseases. However, due to the inherent complexity and heterogeneity of the human microbiome, cross-sectional, case–control and longitudinal studies may not have enough statistical power to allow causation to be deduced from patterns of association between variables in high-resolution omic datasets. Therefore, to move beyond reliance on the empirical method, experiments are critical. For these, robust experimental models are required that allow the systematic manipulation of variables to test the multitude of hypotheses, which arise from high-throughput molecular studies. Particularly promising in this respect are microfluidics-based in vitro co-culture systems, which allow high-throughput first-pass experiments aimed at proving cause-and-effect relationships prior to testing of hypotheses in animal models. This review focuses on widely used in vivo, in vitro, ex vivo and in silico approaches to study host-microbial community interactions. Such systems, either used in isolation or in a combinatory experimental approach, will allow systematic investigations of the impact of microbes on the health and disease of the human host. All the currently available models present pros and cons, which are described and discussed. Moreover, suggestions are made on how to develop future experimental models that not only allow the study of host-microbiota interactions but are also amenable to high-throughput experimentation.

De Novo Lipogenesis Maintains Vascular Homeostasis through Endothelial Nitric-oxide Synthase (eNOS) Palmitoylation

Endothelial dysfunction leads to lethal vascular complications in diabetes and related metabolic disorders. Here, we demonstrate that de novo lipogenesis, an insulin-dependent process driven by the multifunctional enzyme fatty-acid synthase (FAS), maintains endothelial function by targeting endothelial nitric-oxide synthase (eNOS) to the plasma membrane. In mice with endothelial inactivation of FAS (FASTie mice), eNOS membrane content and activity were decreased. eNOS and FAS were physically associated; eNOS palmitoylation was decreased in FAS-deficient cells, and incorporation of labeled carbon into eNOS-associated palmitate was FAS-dependent. FASTie mice manifested a proinflammatory state reflected as increases in vascular permeability, endothelial inflammatory markers, leukocyte migration, and susceptibility to LPS-induced death that was reversed with an NO donor. FAS-deficient endothelial cells showed deficient migratory capacity, and angiogenesis was decreased in FASTie mice subjected to hindlimb ischemia. Insulin induced FAS in endothelial cells freshly isolated from humans, and eNOS palmitoylation was decreased in mice with insulin-deficient or insulin-resistant diabetes. Thus, disrupting eNOS bioavailability through impaired lipogenesis identifies a novel mechanism coordinating nutritional status and tissue repair that may contribute to diabetic vascular disease.

Hedgehog signaling inhibition blocks growth of resistant tumors through effects on tumor microenvironment.

Hedgehog (Hh) signaling is implicated in bone development and cellular transformation. Here we show that inhibition of Hh pathway activity inhibits tumor growth through effects on the microenvironment. Pharmacologic inhibition of the Hh effector Smoothened (Smo) increased trabecular bone in vivo and inhibited osteoclastogenesis in vitro. In addition, enhanced Hh signaling due to heterozygosity of the Hh inhibitory receptor Patched (Ptch1(+/-)) increased bone resorption, suggesting direct regulation of osteoclast (OC) activity by the Hh pathway. Ptch1(+/-) mice had increased bone metastatic and subcutaneous tumor growth, suggesting that increased Hh activation in host cells promoted tumor growth. Subcutaneous growth of Hh-resistant tumor cells was inhibited by LDE225, a novel orally bioavailable SMO antagonist, consistent with effects on tumor microenvironment. Knockdown of the Hh ligand Sonic Hh (SHH) in these cells decreased subcutaneous tumor growth and decreased stromal cell production of interleukin-6, indicating that tumor-derived Hh ligands stimulated tumor growth in a paracrine fashion. Together our findings show that inhibition of the Hh pathway can reduce tumor burden, regardless of tumor Hh responsiveness, through effects on tumor cells, OCs, and stromal cells within the tumor microenvironment. Hh may be a promising therapeutic target for solid cancers and bone metastases.

Recent development and biomedical applications of probabilistic Boolean networks

Probabilistic Boolean network (PBN) modelling is a semi-quantitative approach widely used for the study of the topology and dynamic aspects of biological systems. The combined use of rule-based representation and probability makes PBN appealing for large-scale modelling of biological networks where degrees of uncertainty need to be considered.A considerable expansion of our knowledge in the field of theoretical research on PBN can be observed over the past few years, with a focus on network inference, network intervention and control. With respect to areas of applications, PBN is mainly used for the study of gene regulatory networks though with an increasing emergence in signal transduction, metabolic, and also physiological networks. At the same time, a number of computational tools, facilitating the modelling and analysis of PBNs, are continuously developed.A concise yet comprehensive review of the state-of-the-art on PBN modelling is offered in this article, including a comparative discussion on PBN versus similar models with respect to concepts and biomedical applications. Due to their many advantages, we consider PBN to stand as a suitable modelling framework for the description and analysis of complex biological systems, ranging from molecular to physiological levels.

Prevalence and determinants of osteoporosis in patients with type 1 and type 2 diabetes mellitus

BackgroundIncreased risk of osteoporosis and its clinical significance in patients with diabetes is controversial. We analyze osteoporosis prevalence and determinants of bone mineral density (BMD) in patients with type 1 and 2 diabetes.MethodsThree hundred and ninety-eight consecutive diabetic patients from a single outpatient clinic received a standardized questionnaire on osteoporosis risk factors, and were evaluated for diabetes-related complications, HbA1c levels, and lumbar spine (LS) and femoral neck (FN) BMD. Of these, 139 (71 men, 68 women) type 1 and 243 (115 men, 128 women) type 2 diabetes patients were included in the study. BMD (T-scores and values adjusted for age, BMI and duration of disease) was compared between patient groups and between patients with type 2 diabetes and population-based controls (255 men, 249 women).ResultsFor both genders, adjusted BMD was not different between the type 1 and type 2 diabetes groups but was higher in the type 2 group compared with controls (p < 0.0001). Osteoporosis prevalence (BMD T-score < −2.5 SD) at FN and LS was equivalent in the type 1 and type 2 diabetes groups, but lower in type 2 patients compared with controls (FN: 13.0% vs 21.2%, LS: 6.1% vs 14.9% men; FN: 21.9% vs 32.1%, LS: 9.4% vs 26.9% women). Osteoporosis prevalence was higher at FN-BMD than at LS-BMD. BMD was positively correlated with BMI and negatively correlated with age, but not correlated with diabetes-specific parameters (therapy, HbBA1c, micro- and macrovascular complications) in all subgroups. Fragility fracture prevalence was low (5.2%) and not different between diabetes groups. Fracture patients had lower BMDs compared with those without fractures; however, BMD T-score was above −2.5 SD in most patients.ConclusionsDiabetes-specific parameters did not predict BMD. Fracture occurrence was similar in both diabetes groups and related to lower BMD, but seems unrelated to the threshold T-score, <−2.5 SD. These results suggest that osteoporosis, and related fractures, is a clinically significant and commonly underestimated problem in diabetes patients.

The ADP receptor P2RY12 regulates osteoclast function and pathologic bone remodeling

The adenosine diphosphate (ADP) receptor P2RY12 (purinergic receptor P2Y, G protein coupled, 12) plays a critical role in platelet aggregation, and P2RY12 inhibitors are used clinically to prevent cardiac and cerebral thrombotic events. Extracellular ADP has also been shown to increase osteoclast (OC) activity, but the role of P2RY12 in OC biology is unknown. Here, we examined the role of mouse P2RY12 in OC function. Mice lacking P2ry12 had decreased OC activity and were partially protected from age-associated bone loss. P2ry12-/- OCs exhibited intact differentiation markers, but diminished resorptive function. Extracellular ADP enhanced OC adhesion and resorptive activity of WT, but not P2ry12-/-, OCs. In platelets, ADP stimulation of P2RY12 resulted in GTPase Ras-related protein (RAP1) activation and subsequent αIIbβ3 integrin activation. Likewise, we found that ADP stimulation induced RAP1 activation in WT and integrin β3 gene knockout (Itgb3-/-) OCs, but its effects were substantially blunted in P2ry12-/- OCs. In vivo, P2ry12-/- mice were partially protected from pathologic bone loss associated with serum transfer arthritis, tumor growth in bone, and ovariectomy-induced osteoporosis: all conditions associated with increased extracellular ADP. Finally, mice treated with the clinical inhibitor of P2RY12, clopidogrel, were protected from pathologic osteolysis. These results demonstrate that P2RY12 is the primary ADP receptor in OCs and suggest that P2RY12 inhibition is a potential therapeutic target for pathologic bone loss.

Integrated multi-omics of the human gut microbiome in a case study of familial type 1 diabetes.

1 Erratum: Integrated multi-omics of the human gut microbiome in a case study of familial type 1 diabetes Anna Heintz-Buschart, Patrick May, Cédric C. Laczny, Laura A. Lebrun, Camille Bellora, Abhimanyu Krishna, Linda Wampach, Jochen G. Schneider, Angela Hogan, Carine de Beaufort and Paul Wilmes Nature Microbiology 2, 16180 (2016); published 10 October 2016; corrected 24 October 2016 This Article should have been published under a Creative Commons licence according to the Nature policy on publishing the primary sequence of an organism’s genome for the first time. The editors apologize to the authors and to readers for this error. The manuscript is now open access and published under a CC-BY licence. All versions of the Article have been modified accordingly. ARTICLES NATURE MICROBIOLOGY DOI: 10.1038/NMICROBIOL.2016.227

Molecular and Clinical Evidence for an ARMC5 Tumor Syndrome: Concurrent Inactivating Germline and Somatic Mutations Are Associated With Both Primary Macronodular Adrenal Hyperplasia and Meningioma

CONTEXT Primary macronodular adrenal hyperplasia (PMAH) is a rare cause of Cushings syndrome, which may present in the context of different familial multitumor syndromes. Heterozygous inactivating germline mutations of armadillo repeat containing 5 (ARMC5) have very recently been described as cause for sporadic PMAH. Whether this genetic condition also causes familial PMAH in association with other neoplasias is unclear. OBJECTIVE The aim of the present study was to delineate the molecular cause in a large family with PMAH and other neoplasias. PATIENTS AND METHODS Whole-genome sequencing and comprehensive clinical and biochemical phenotyping was performed in members of a PMAH affected family. Nodules derived from adrenal surgery and pancreatic and meningeal tumor tissue were analyzed for accompanying somatic mutations in the identified target genes. RESULTS PMAH presenting either as overt or subclinical Cushings syndrome was accompanied by a heterozygous germline mutation in ARMC5 (p.A110fs*9) located on chromosome 16. Analysis of tumor tissue showed different somatic ARMC5 mutations in adrenal nodules supporting a second hit hypothesis with inactivation of a tumor suppressor gene. A damaging somatic ARMC5 mutation was also found in a concomitant meningioma (p.R502fs) but not in a pancreatic tumor, suggesting biallelic inactivation of ARMC5 as causal also for the intracranial meningioma. CONCLUSIONS Our analysis further confirms inherited inactivating ARMC5 mutations as a cause of familial PMAH and suggests an additional role for the development of concomitant intracranial meningiomas.