Acute Effect of Caffeine on the Synthesis of Pro-Inflammatory Cytokines in the Hypothalamus and Choroid Plexus during Endotoxin-Induced Inflammation in a Female Sheep Model

This study was designed to determine the effect of acute caffeine (CAF) administration, which exerts a broad spectrum of anti-inflammatory activity, on the synthesis of pro-inflammatory cytokines and their receptors in the hypothalamus and choroid plexus (ChP) during acute inflammation caused by the injection of bacterial endotoxin—lipopolysaccharide (LPS). The experiment was performed on 24 female sheep randomly divided into four groups: control; LPS treated (iv.; 400 ng/kg of body mass (bm.)); CAF treated (iv.; 30 mg/kg of bm.); and LPS and CAF treated. The animals were euthanized 3 h after the treatment. It was found that acute administration of CAF suppressed the synthesis of interleukin (IL-1β) and tumor necrosis factor (TNF)α, but did not influence IL-6, in the hypothalamus during LPS-induced inflammation. The injection of CAF reduced the LPS-induced expression of TNF mRNA in the ChP. CAF lowered the gene expression of IL-6 cytokine family signal transducer (IL6ST) and TNF receptor superfamily member 1A (TNFRSF1) in the hypothalamus and IL-1 type II receptor (IL1R2) in the ChP. Our study on the sheep model suggests that CAF may attenuate the inflammatory response at the hypothalamic level and partly influence the inflammatory signal generated by the ChP cells. This suggests the potential of CAF to suppress neuroinflammatory processes induced by peripheral immune/inflammatory challenges.     

Preterm Intraventricular Hemorrhage-Induced Inflammatory Response in Human Choroid Plexus Epithelial Cells

Following an intraventricular hemorrhage (IVH), red blood cell lysis and hemoglobin (Hb) oxidation with the release of heme can cause sterile neuroinflammation. In this study, we measured Hb derivates and cellular adhesion molecules ICAM-1 and VCAM-1 with cell-free miRNAs in cerebrospinal fluid (CSF) samples obtained from Grade-III and Grade-IV preterm IVH infants (IVH-III and IVH-IV, respectively) at multiple time points between days 0–60 after the onset of IVH. Furthermore, human choroid plexus epithelial cells (HCPEpiCs) were incubated with IVH and non-IVH CSF (10 v/v %) for 24 h in vitro to investigate the IVH-induced inflammatory response that was investigated via: (i) HMOX1, IL8, VCAM1, and ICAM1 mRNAs as well as miR-155, miR-223, and miR-181b levels by RT-qPCR; (ii) nuclear translocation of the NF-κB p65 subunit by fluorescence microscopy; and (iii) reactive oxygen species (ROS) measurement. We found a time-dependent alteration of heme, IL-8, and adhesion molecules which revealed a prolonged elevation in IVH-IV vs. IVH-III with higher miR-155 and miR-181b expression at days 41–60. Exposure of HCPEpiCs to IVH CSF samples induced HMOX1, IL8, and ICAM1 mRNA levels along with increased ROS production via the NF-κB pathway activation but without cell death, as confirmed by the cell viability assay. Additionally, the enhanced intracellular miR-155 level was accompanied by lower miR-223 and miR-181b expression in HCPEpiCs after CSF treatment. Overall, choroid plexus epithelial cells exhibit an abnormal cell phenotype after interaction with pro-inflammatory CSF of IVH origin which may contribute to the development of later clinical complications in preterm IVH.     

Multiple Na,K-ATPase Subunits Colocalize in the Brush Border of Mouse Choroid Plexus Epithelial Cells

(1) Background: The unusual accumulation of Na,K-ATPase complexes in the brush border membrane of choroid plexus epithelial cells have intrigued researchers for decades. However, the full range of the expressed Na,K-ATPase subunits and their relation to the microvillus cytoskeleton remains unknown. (2) Methods: RT-PCR analysis, co-immunoprecipitation, native PAGE, mass spectrometry, and differential centrifugation were combined with high-resolution immunofluorescence histochemistry, proximity ligase assays, and stimulated emission depletion (STED) microscopy on mouse choroid plexus cells or tissues in order to resolve these issues. (3) Results: The choroid plexus epithelium expresses Na,K-ATPase subunits α1, α2, β1, β2, β3, and phospholemman. The α1, α2, β1, and β2, subunits are all localized to the brush border membrane, where they appear to form a complex. The ATPase complexes may stabilize in the brush border membrane via anchoring to microvillar actin indirectly through ankyrin-3 or directly via other co-precipitated proteins. Aquaporin 1 (AQP1) may form part of the proposed multi-protein complexes in contrast to another membrane protein, the Na-K-2Cl cotransporter 1 (NKCC1). NKCC1 expression seems necessary for full brush border membrane accumulation of the Na,K-ATPase in the choroid plexus. (4) Conclusion: A multitude of Na,K-ATPase subunits form molecular complexes in the choroid plexus brush border, which may bind to the cytoskeleton by various alternative actin binding proteins.     

Biomarkers Related to Synaptic Dysfunction to Discriminate Alzheimer’s Disease from Other Neurological Disorders

Recently, the synaptic proteins neurogranin (Ng) and α-synuclein (α-Syn) have attracted scientific interest as potential biomarkers for synaptic dysfunction in neurodegenerative diseases. In this study, we measured the CSF Ng and α-Syn concentrations in patients affected by AD (n = 69), non-AD neurodegenerative disorders (n-AD = 50) and non-degenerative disorders (n-ND, n = 98). The concentrations of CSF Ng and α-Syn were significantly higher in AD than in n-AD and n-ND. Moreover, the Aβ42/Ng and Aβ42/α-Syn ratios showed statistically significant differences between groups and discriminated AD patients from n-AD patients, better than Ng or α-Syn alone. Regression analyses showed an association of higher Ng concentrations with MMSE < 24, pathological Aβ 42/40 ratios, pTau, tTau and the ApoEε4 genotype. Aβ 42/Ng was associated with MMSE < 24, an AD-related FDG-PET pattern, the ApoEε4 genotype, pathological Aβ 42 levels and Aβ 42/40 ratios, pTau, and tTau. Moreover, APO-Eε4 carriers showed higher Ng concentrations than non-carriers. Our results support the idea that the Aβ 42/Ng ratio is a reliable index of synaptic dysfunction/degeneration able to discriminate AD from other neurological conditions.     

ADAM10 Plasma and CSF Levels Are Increased in Mild Alzheimer’s Disease

ADAM10 is the main α-secretase that participates in the non-amyloidogenic cleavage of amyloid precursor protein (APP) in neurons, inhibiting the production of β-amyloid peptide (Aβ) in Alzheimer’s disease (AD). Strong recent evidence indicates the importance of the localization of ADAM10 for its activity as a protease. In this study, we investigated ADAM10 activity in plasma and CSF samples of patients with amnestic mild cognitive impairment (aMCI) and mild AD compared with cognitively healthy controls. Our results indicated that plasma levels of soluble ADAM10 were significantly increased in the mild AD group, and that in these samples the protease was inactive, as determined by activity assays. The same results were observed in CSF samples, indicating that the increased plasma ADAM10 levels reflect the levels found in the central nervous system. In SH-SY5Y neuroblastoma cells, ADAM10 achieves its major protease activity in the fraction obtained from plasma membrane lysis, where the mature form of the enzyme is detected, confirming the importance of ADAM10 localization for its activity. Taken together, our results demonstrate the potential of plasma ADAM10 to act as a biomarker for AD, highlighting its advantages as a less invasive, easier, faster, and lower-cost processing procedure, compared to existing biomarkers.     

CSF Biomarkers Reflecting Protein Pathology and Axonal Degeneration Are Associated with Memory,Attentional, and Executive Functioning in Early-Stage Parkinson′s Disease

In early-stage Parkinson′s disease (PD), cognitive impairment is common, and a variety of cognitive domains including memory, attention, and executive functioning may be affected. Cerebrospinal fluid (CSF) biomarkers are potential markers of cognitive functioning. We aimed to explore whether CSF α-synuclein species, neurofilament light chain, amyloid-β₄₂, and tau are associated with cognitive performance in early-stage PD patients. CSF levels of total-α-synuclein and phosphorylated-α-synuclein, neurofilament light chain, amyloid-β₄₂, and total-tau and phosphorylated-tau were measured in 26 PD patients (disease duration ≤5 years and Hoehn and Yahr stage 1–2.5). Multivariable linear regression models, adjusted for age, gender, and educational level, were used to assess the relationship between CSF biomarker levels and memory, attention, executive and visuospatial function, and language performance scores. In 26 early-stage PD patients, attention and memory were the most commonly affected domains. A higher CSF phosphorylated-α-synuclein/total-α-synuclein ratio was associated with better executive functioning (sβ = 0.40). Higher CSF neurofilament light was associated with worse memory (sβ = −0.59), attentional (sβ = −0.32), and executive functioning (sβ = −0.35). Reduced CSF amyloid-β₄₂ levels were associated with poorer attentional functioning (sβ = 0.35). Higher CSF phosphorylated-tau was associated with worse language functioning (sβ = −0.33). Thus, CSF biomarker levels, in particular neurofilament light, were related to the most commonly affected cognitive domains in early-stage PD. This indicates that CSF biomarker levels may identify early-stage PD patients who are at an increased risk of developing cognitive impairment.     

Establishing In-House Cutoffs of CSF Alzheimer’s Disease Biomarkers for the AT(N) Stratification of the Alzheimer Center Barcelona Cohort

Background: Clinical diagnosis of Alzheimer’s disease (AD) increasingly incorporates CSF biomarkers. However, due to the intrinsic variability of the immunodetection techniques used to measure these biomarkers, establishing in-house cutoffs defining the positivity/negativity of CSF biomarkers is recommended. However, the cutoffs currently published are usually reported by using cross-sectional datasets, not providing evidence about its intrinsic prognostic value when applied to real-world memory clinic cases. Methods: We quantified CSF Aβ1-42, Aβ1-40, t-Tau, and p181Tau with standard INNOTEST^® ELISA and Lumipulse G^® chemiluminescence enzyme immunoassay (CLEIA) performed on the automated Lumipulse G600II. Determination of cutoffs included patients clinically diagnosed with probable Alzheimer’s disease (AD, n = 37) and subjective cognitive decline subjects (SCD, n = 45), cognitively stable for 3 years and with no evidence of brain amyloidosis in 18F-Florbetaben-labeled positron emission tomography (FBB-PET). To compare both methods, a subset of samples for Aβ1-42 (n = 519), t-Tau (n = 399), p181Tau (n = 77), and Aβ1-40 (n = 44) was analyzed. Kappa agreement of single biomarkers and Aβ1-42/Aβ1-40 was evaluated in an independent group of mild cognitive impairment (MCI) and dementia patients (n = 68). Next, established cutoffs were applied to a large real-world cohort of MCI subjects with follow-up data available (n = 647). Results: Cutoff values of Aβ1-42 and t-Tau were higher for CLEIA than for ELISA and similar for p181Tau. Spearman coefficients ranged between 0.81 for Aβ1-40 and 0.96 for p181TAU. Passing–Bablok analysis showed a systematic and proportional difference for all biomarkers but only systematic for Aβ1-40. Bland–Altman analysis showed an average difference between methods in favor of CLEIA. Kappa agreement for single biomarkers was good but lower for the Aβ1-42/Aβ1-40 ratio. Using the calculated cutoffs, we were able to stratify MCI subjects into four AT(N) categories. Kaplan–Meier analyses of AT(N) categories demonstrated gradual and differential dementia conversion rates (p = 9.815⁻²⁷). Multivariate Cox proportional hazard models corroborated these findings, demonstrating that the proposed AT(N) classifier has prognostic value. AT(N) categories are only modestly influenced by other known factors associated with disease progression. Conclusions: We established CLEIA and ELISA internal cutoffs to discriminate AD patients from amyloid-negative SCD individuals. The results obtained by both methods are not interchangeable but show good agreement. CLEIA is a good and faster alternative to manual ELISA for providing AT(N) classification of our patients. AT(N) categories have an impact on disease progression. AT(N) classifiers increase the certainty of the MCI prognosis, which can be instrumental in managing real-world MCI subjects.     

Specific Cerebrospinal Fluid SerpinA1 Isoform Pattern in Alzheimer’s Disease

SerpinA1 (α1-antitrypsin) is a soluble glycoprotein, the cerebrospinal fluid (CSF) isoforms of which showed disease-specific changes in neurodegenerative disorders that are still unexplored in Alz-heimer’s disease (AD). By means of capillary isoelectric focusing immunoassay, we investigated six serpinA1 isoforms in CSF samples of controls (n = 29), AD-MCI (n = 29), AD-dem (n = 26) and Lewy body disease (LBD, n = 59) patients and correlated the findings with CSF AD core biomarkers (Aβ42/40 ratio, p-tau, t-tau). Four CSF serpinA1 isoforms were differently expressed in AD patients compared to controls and LBD patients, especially isoforms 2 and 4. AD-specific changes were found since the MCI stage and significantly correlated with decreased Aβ42/40 (p < 0.05) and in-creased p-tau and t-tau levels in CSF (p < 0.001). Analysis of serpinA1 isoform provided good di-agnostic accuracy in discriminating AD patients versus controls (AUC = 0.80) and versus LBD patients (AUC = 0.92), with best results in patients in the dementia stage (AUC = 0.97). SerpinA1 isoform expression is altered in AD patients, suggesting a common, albeit disease-specific, in-volvement of serpinA1 in most neurodegenerative disorders.     

ROS-Scavengers,Osmoprotectants and Violaxanthin De-Epoxidation in Salt-Stressed Arabidopsis thaliana with Different Tocopherol Composition

To determine the role of α- and γ-tocopherol (TC), this study compared the response to salt stress (200 mM NaCl) in wild type (WT) Arabidopsis thaliana (L.) Heynh. And its two mutants: (1) totally TC-deficient vte1; (2) vte4 accumulating γ-TC instead of α-TC; and (3) tmt transgenic line overaccumulating α-TC. Raman spectra revealed that salt-exposed α-TC accumulating plants were more flexible in regulating chlorophyll, carotenoid and polysaccharide levels than TC deficient mutants, while the plants overaccumulating γ-TC had the lowest levels of these biocompounds. Tocopherol composition and NaCl concentration affected xanthophyll cycle by changing the rate of violaxanthin de-epoxidation and zeaxanthin formation. NaCl treated plants with altered TC composition accumulated less oligosaccharides than WT plants. α-TC deficient plants increased their oligosaccharide levels and reduced maltose amount, while excessive accumulation of α-TC corresponded with enhanced amounts of maltose. Salt-stressed TC-deficient mutants and tmt transgenic line exhibited greater proline levels than WT plants, lower chlorogenic acid levels, and lower activity of catalase and peroxidases. α-TC accumulating plants produced more methylated proline- and glycine- betaines, and showed greater activity of superoxide dismutase than γ-TC deficient plants. Under salt stress, α-TC demonstrated a stronger regulatory effect on carbon- and nitrogen-related metabolites reorganization and modulation of antioxidant patterns than γ-TC. This suggested different links of α- and γ-TCs with various metabolic pathways via various functions and metabolic loops.     

Quantitative Plasma Proteomics to Identify Candidate Biomarkers of Relapse in Pediatric/Adolescent Hodgkin Lymphoma

Classical pediatric Hodgkin Lymphoma (HL) is a rare malignancy. Therapeutic regimens for its management may be optimized by establishing treatment response early on. The aim of this study was to identify plasma protein biomarkers enabling the prediction of relapse in pediatric/adolescent HL patients treated under the pediatric EuroNet-PHL-C2 trial. We used untargeted liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based proteomics at the time of diagnosis—before any therapy—as semiquantitative method to profile plasma proteins specifically associated with relapse in 42 children with nodular sclerosing HL. In both the exploratory and the validation cohorts, six proteins (apolipoprotein E, C4b-binding protein α chain, clusterin, fibrinogen γ chain, prothrombin, and vitronectin) were more abundant in the plasma of patients whose HL relapsed (|fold change| ≥ 1.2, p < 0.05, Student’s t-test). Predicting protein function with the Gene Ontology classification model, the proteins were included in four biological processes (p < 0.01). Using immunoblotting and Luminex assays, we validated two of these candidate biomarkers—C4b-binding protein α chain and clusterin—linked to innate immune response function (GO:0045087). This study identified C4b-binding protein α chain and clusterin as candidate early plasma biomarkers of HL relapse, and important for the purpose of shedding light on the molecular scenario associated with immune response in patients treated under the EuroNet-PHL-C2 trial.     

Triterpenoids from the Roots of Rhaphiolepis indica var. tashiroi and Their Anti-Inflammatory Activity

Two new triterpenoids, 2α,3β-dihydroxyolean-11,13(18)-dien-19β,28-olide (1) and 3β,5β-dihydroxyglutinol (2), together with eight known compounds (3–10) were isolated from the roots of Rhaphiolepis indica var. tashiroi (Rosaceae). The structures of 1–10 were determined by spectroscopic techniques. Among these isolates, 2α,3β-dihydroxyolean-13(18)-en-28-oic acid (9) exhibited inhibitory effect on N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced superoxide production, with an IC₅₀ value of 16.50 μM.     

Altered Balance of Reelin Proteolytic Fragments in the Cerebrospinal Fluid of Alzheimer’s Disease Patients

Reelin binds to the apolipoprotein E receptor apoER2 to activate an intracellular signaling cascade. The proteolytic cleavage of reelin follows receptor binding but can also occur independently of its binding to receptors. This study assesses whether reelin proteolytic fragments are differentially affected in the cerebrospinal fluid (CSF) of Alzheimer’s disease (AD) subjects. CSF reelin species were analyzed by Western blotting, employing antibodies against the N- and C-terminal domains. In AD patients, we found a decrease in the 420 kDa full-length reelin compared with controls. In these patients, we also found an increase in the N-terminal 310 kDa fragment resulting from the cleavage at the so-called C-t site, whereas the 180 kDa fragment originated from the N-t site remained unchanged. Regarding the C-terminal proteolytic fragments, the 100 kDa fragment resulting from the cleavage at the C-t site also displayed increased levels, whilst the one resulting from the N-t site, the 250 kDa fragment, decreased. We also detected the presence of an aberrant reelin species with a molecular mass of around 500 kDa present in AD samples (34 of 43 cases), while it was absent in the 14 control cases analyzed. These 500 kDa species were only immunoreactive to N-terminal antibodies. We validated the occurrence of these aberrant reelin species in an Aβ42-treated reelin-overexpressing cell model. When we compared the AD samples from APOE genotype subgroups, we only found minor differences in the levels of reelin fragments associated to the APOE genotype, but interestingly, the levels of fragments of apoER2 were lower in APOE ε4 carriers with regards to APOE ε3/ε3. The altered proportion of reelin/apoER2 fragments and the occurrence of reelin aberrant species suggest a complex regulation of the reelin signaling pathway, which results impaired in AD subjects.     

Effect of γ-Cyclodextrin Inclusion Complex on the Absorption of R-α-Lipoic Acid in Rats

R-α-lipoic acid (RLA) is an endogenous organic acid, and works as a cofactor for mitochondrial enzymes and as a kind of antioxidant. Inclusion complexes of RLA with α-, β- or γ-cyclodextrins (CD) were prepared and orally administered as a suspension to rats. Among them, RLA/γ-CD showed the highest plasma exposure, and its area under the plasma concentration-time curve (AUC) of RLA was 2.2 times higher than that after oral administration of non-inclusion RLA. On the other hand, the AUC after oral administration of non-inclusion RLA and RLA/γ-CD to pylorus-ligated rats did not differ. However, the AUC after intraduodenal administration of RLA/γ-CD was 5.1 times higher than that of non-inclusion RLA, and was almost comparable to the AUC after intraduodenal administration of RLA-Na solution. Furthermore, the AUC after intraduodenal administration of RLA/γ-CD was not affected by biliary ligation or co-administration of an amylase inhibitor. These findings demonstrated that RLA was absorbed from the small intestine effectively when orally administered as a γ-CD inclusion complex, which could be easily dissolved in the lumen of the intestine. In conclusion, γ-CD inclusion complex is an appropriate formulation for supplying RLA as a drug or nutritional supplement with respect to absorption.     

Ingested Ketone Ester Leads to a Rapid Rise of Acetyl-CoA and Competes with Glucose Metabolism in the Brain of Non-Fasted Mice

The role of ketone bodies in the cerebral energy homeostasis of neurological diseases has begun to attract recent attention particularly in acute neurological diseases. In ketogenic therapies, ketosis is achieved by either a ketogenic diet or by the administration of exogenous ketone bodies. The oral ingestion of the ketone ester (KE), (R)-3-hydroxybutyl (R)-3-hydroxybutyrate, is a new method to generate rapid and significant ketosis (i.e., above 6 mmol/L) in humans. KE is hydrolyzed into β-hydroxybutyrate (βHB) and its precursor 1,3-butanediol. Here, we investigate the effect of oral KE administration (3 mg KE/g of body weight) on brain metabolism of non-fasted mice using liquid chromatography in tandem with mass spectrometry. Ketosis (Cmax = 6.83 ± 0.19 mmol/L) was obtained at Tmax = 30 min after oral KE-gavage. We found that βHB uptake into the brain strongly correlated with the plasma βHB concentration and was preferentially distributed in the neocortex. We showed for the first time that oral KE led to an increase of acetyl-CoA and citric cycle intermediates in the brain of non-fasted mice. Furthermore, we found that the increased level of acetyl-CoA inhibited glycolysis by a feedback mechanism and thus competed with glucose under physiological conditions. The brain pharmacodynamics of this oral KE strongly suggest that this agent should be considered for acute neurological diseases.     

Functional Characterization of Largemouth Bass (Micropterus salmoides) Soluble FcγR Homolog in Response to Bacterial Infection

Fc receptors (FcRs) are key players in antibody-dependent cellular phagocytosis (ADCP) with their specific recognition of the Fc portion of an immunoglobulin. Despite reports of FcγR-mediated phagocytosis in mammals, little is known about the effects of soluble FcγRs on the immune response. In this study, FcγRIα was cloned from the largemouth bass (Micropterus salmoides) (MsFcγRIα). Without a transmembrane segment or a cytoplasmic tail, MsFcγRIα was identified as a soluble form protein and widely distributed in the spleen, head kidney, and intestine. The native MsFcγRIα was detected in the serum of Nocardia seriolae-infected largemouth bass and the supernatants of transfected HEK293 cells. Additionally, it was verified that the transfected cells’ surface secreted MsFcRIα could bind to largemouth bass IgM. Moreover, the expression changes of MsFcγRIα, Syk, and Lyn indicated that MsFcγRIα was engaged in the acute phase response to bacteria, and the FcγR-mediated phagocytosis pathway was activated by Nocardia seriolae stimulation. Furthermore, recombinant MsFcγRIα could enhance both reactive oxygen species (ROS) and phagocytosis to Nocardia seriolae of leukocytes, presumably through the interaction of MsFcγRIα with a complement receptor. In conclusion, these findings provided a better understanding of the function of soluble FcγRs in the immune response and further shed light on the mechanism of phagocytosis in teleosts.     

The α2δ Calcium Channel Subunit Accessorily and Independently Affects the Biological Function of Ditylenchus destructor

The α₂δ subunit is a high-voltage activated (HVA) calcium channel (Ca_v1 and Ca_v2) auxiliary subunit that increases the density and function of HVA calcium channels in the plasma membrane of mammals. However, its function in plant parasitic nematodes remains unknown. In this study, we cloned the full-length cDNA sequence of the voltage-gated calcium channel (VGCC) α₂δ subunit (named DdCa_vα₂δ) in Ditylenchus destructor. We found that DdCa_vα₂δ tends to be expressed in the egg stage, followed by the J3 stage. RNA-DIG in situ hybridization experiments showed that the DdCa_vα₂δ subunit was expressed in the body wall, esophageal gland, uterus, post uterine, and spicules of D. destructor. The in vitro application of RNA interference (RNAi) affected the motility, reproduction, chemotaxis, stylet thrusting, and protein secretion of D. destructor to different degrees by targeting DdCα1D, DdCα1A, and DdCa_vα₂δ in J3 stages, respectively. Based on the results of RNAi experiments, it was hypothesized that L-type VGCC may affect the motility, chemotaxis, and stylet thrusting of D. destructor. Non-L-type VGCC may affect the protein secretion and reproduction of D. destructor. The DdCa_vα₂δ subunit gene also affected the motility, chemotaxis, and reproduction of D. destructor. These findings reveal the independent function of the VGCC α₂δ subunit in D. destructor as well as give a theoretical foundation for future research on plant parasitic nematode VGCC.     

Differential Effects of Furin Deficiency on Insulin Receptor Processing and Glucose Control in Liver and Pancreatic β Cells of Mice

The insulin receptor (IR) is critically involved in maintaining glucose homeostasis. It undergoes proteolytic cleavage by proprotein convertases, which is an essential step for its activation. The importance of the insulin receptor in liver is well established, but its role in pancreatic β cells is still controversial. In this study, we investigated the cleavage of the IR by the proprotein convertase FURIN in β cells and hepatocytes, and the contribution of the IR in pancreatic β cells and liver to glucose homeostasis. β-cell-specific Furin knockout (βFurKO) mice were glucose intolerant, but liver-specific Furin knockout (LFurKO) mice were normoglycemic. Processing of the IR was blocked in βFurKO cells, but unaffected in LFurKO mice. Most strikingly, glucose homeostasis in β-cell-specific IR knockout (βIRKO) mice was normal in younger mice (up to 20 weeks), and only mildly affected in older mice (24 weeks). In conclusion, FURIN cleaves the IR non-redundantly in β cells, but redundantly in liver. Furthermore, we demonstrated that the IR in β cells plays a limited role in glucose homeostasis.