Double Thionated Pyrimidine Nucleobases: Molecular Tools with Tunable Photoproperties.

Sulfur-substituted nucleobases are DNA and RNA base derivatives that exhibit extremely efficient photoinduced intersystem crossing (ISC) dynamics into the lowest-energy triplet state. The long-lived and reactive triplet states of sulfur-substituted nucleobases are crucial due to their wide range of potential applications in medicine, structural biology, and the development of organic light-emitting diodes (OLEDs) and other emerging technologies. However, a comprehensive understanding of non-negligible wavelength-dependent changes in the internal conversion (IC) and ISC events is still lacking. Here, we study the underlying mechanism using joint experimental gas-phase time-resolved photoelectron spectroscopy (TRPES) and theoretical quantum chemistry methods. We combine 2,4-dithiouracil (2,4-DTU) TRPES experimental data with computational analysis of the different photodecay processes, which are induced by increasing excitation energies along the entire linear absorption (LA) ultraviolet (UV) spectrum. Our results show how the double-thionated uracil (U), i.e., 2,4-DTU, appears as a versatile photoactivatable instrument. Multiple decay processes can be initiated with different ISC rates or triplet-state lifetimes that resemble the distinctive behavior of the singly substituted 2- or 4-thiouracil (2-TU or 4-TU). We obtained a clear partition of the LA spectrum based on the dominant photoinduced process. Our work clarifies the reasons behind the wavelength-dependent changes in the IC, ISC, and triplet-state lifetimes in doubly thionated U, becoming a biological system of utmost importance for wavelength-controlled applications. These mechanistic details and photoproperties are transferable to closely related molecular systems such as thionated thymines.

Glucose-stimulated insulin secretion depends on FFA1 and Gq in neonatal mouse islets.

AIMS/HYPOTHESIS: After birth, the neonatal islets gradually acquire glucose-responsive insulin secretion, a process that is subjected to maternal imprinting. Although NEFA are major components of breastmilk and insulin secretagogues, their role for functional maturation of neonatal beta cells is still unclear. NEFA are the endogenous ligands of fatty acid receptor 1 (FFA1, encoded by Ffar1 in mice), a Gq-coupled receptor with stimulatory effect on insulin secretion. This study investigates the role of FFA1 in neonatal beta cell function and in the adaptation of offspring beta cells to parental high-fat feeding. METHODS: Wild-type (WT) and Ffar1-/- mice were fed high-fat (HFD) or chow diet (CD) for 8 weeks before mating, and during gestation and lactation. Blood variables, pancreas weight and insulin content were assessed in 1-, 6-, 11- and 26-day old (P1-P26) offspring. Beta cell mass and proliferation were determined in P1-P26 pancreatic tissue sections. FFA1/Gq dependence of insulin secretion was evaluated in isolated islets and INS-1E cells using pharmacological inhibitors and siRNA strategy. Transcriptome analysis was conducted in isolated islets. RESULTS: Blood glucose levels were higher in CD-fed Ffar1-/- P6-offspring compared with CD-fed WT P6-offspring. Accordingly, glucose-stimulated insulin secretion (GSIS) and its potentiation by palmitate were impaired in CD Ffar1-/- P6-islets. In CD WT P6-islets, insulin secretion was stimulated four- to fivefold by glucose and five- and sixfold over GSIS by palmitate and exendin-4, respectively. Although parental HFD increased blood glucose in WT P6-offspring, it did not change insulin secretion from WT P6-islets. In contrast, parental HFD abolished glucose responsiveness (i.e. GSIS) in Ffar1-/- P6-islets. Inhibition of Gq by FR900359 or YM-254890 in WT P6-islets mimicked the effect of Ffar1 deletion, i.e. suppression of GSIS and of palmitate-augmented GSIS. The blockage of Gi/o by pertussis toxin (PTX) enhanced (100-fold) GSIS in WT P6-islets and rendered Ffar1-/- P6-islets glucose responsive, suggesting constitutive activation of Gi/o. In WT P6-islets, FR900359 cancelled 90% of PTX-mediated stimulation, while in Ffar1-/- P6-islets it completely abolished PTX-elevated GSIS. The secretory defect of Ffar1-/- P6-islets did not originate from insufficient beta cells, since beta cell mass increased with the offspring's age irrespective of genotype and diet. In spite of that, in the breastfed offspring (i.e. P1-P11) beta cell proliferation and pancreatic insulin content had a genotype- and diet-driven dynamic. Under CD, the highest proliferation rate was reached by the Ffar1-/- P6 offspring (3.95% vs 1.88% in WT P6), whose islets also showed increased mRNA levels of genes (e.g. Fos, Egr1, Jun) typically high in immature beta cells. Although parental HFD increased beta cell proliferation in both WT (4.48%) and Ffar1-/- (5.19%) P11 offspring, only the WT offspring significantly increased their pancreatic insulin content upon parental HFD (5.18 µg under CD to 16.93 µg under HFD). CONCLUSIONS/INTERPRETATION: FFA1 promotes glucose-responsive insulin secretion and functional maturation of newborn islets and is required for adaptive offspring insulin secretion in the face of metabolic challenge, such as parental HFD.

Charge Dynamics of a CuO Thin Film on Picosecond to Microsecond Timescales Revealed by Transient Absorption Spectroscopy.

Understanding the mechanism of charge dynamics in photocatalysts is the key to design and optimize more efficient materials for renewable energy applications. In this study, the charge dynamics of a CuO thin film are unraveled via transient absorption spectroscopy (TAS) on the picosecond to microsecond timescale for three different excitation energies, i.e., above, near, and below the band gap, to explore the role of incoherent broadband light sources. The shape of the ps-TAS spectra changes with the delay time, while that of the ns-TAS spectra is invariant for all the excitation energies. Regardless of the excitations, three time constants, τ1 ∼ 0.34-0.59 ps, τ2 ∼ 162-175 ns, and τ3 ∼ 2.5-3.3 µs, are resolved, indicating the dominating charge dynamics at very different timescales. Based on these observations, the UV-vis absorption spectrum, and previous findings in the literature, a compelling transition energy diagram is proposed. Two conduction bands and two defect (deep and shallow) states dominate the initial photo-induced electron transitions, and a sub-valence band energy state is involved in the subsequent transient absorption. By solving the rate equations for the pump-induced population dynamics and implementing the assumed Lorentzian absorption spectral shape between two energy states, the TAS spectra are modeled which capture the main spectral and time-dependent features for t > 1 ps. By further considering the contributions from free-electron absorption during very early delay times, the modeled spectra reproduce the experimental spectra very well over the entire time range and under different excitation conditions.

The Effect of Methylation on the Triplet-State Dynamics of 2-Thiouracil: Time-Resolved Photoelectron Spectroscopy of 2-Thiothymine.

The ultrafast internal conversion and intersystem crossing dynamics of 2-thiouracil (2TU) and 2-thiothymine (2TT) are studied using time-resolved photoelectron spectroscopy to investigate the effect of methylation on the deactivation mechanism. Like other thiobases, the triplet manifold is populated with high quantum yields via the lowest singlet excited state, which is dark in absorption. This study focuses on the lowest triplet state and the role of two minima, with sulfur-out-of-plane and slightly boat-like geometries, in the intersystem crossing dynamics back to the ground state.

Effects of adrenergic-stimulated lipolysis and cytokine production on in vitro mouse adipose tissue-islet interactions.

Inflammatory cytokines and non-esterified fatty acids (NEFAs) are obesity-linked factors that disturb insulin secretion. The aim of this study was to investigate whether pancreatic adipose tissue (pWAT) is able to generate a NEFA/cytokine overload within the pancreatic environment and as consequence to impact on insulin secretion. Pancreatic fat is a minor fat depot, therefore we used high-fat diet (HFD) feeding to induce pancreatic steatosis in mice. Relative Adipoq and Lep mRNA levels were higher in pWAT of HFD compared to chow diet mice. Regardless of HFD, Adipoq and Lep mRNA levels of pWAT were at least 10-times lower than those of epididymal fat (eWAT). Lipolysis stimulating receptors Adrb3 and Npr1 were expressed in pWAT and eWAT, and HFD reduced their expression in eWAT only. In accordance, HFD impaired lipolysis in eWAT but not in pWAT. Despite expression of Npr mRNA, lipolysis was stimulated solely by the adrenergic agonists, isoproterenol and adrenaline. Short term co-incubation of islets with CD/HFD pWAT did not alter insulin secretion. In the presence of CD/HFD eWAT, glucose stimulated insulin secretion only upon isoproterenol-induced lipolysis, i.e. in the presence of elevated NEFA. Isoproterenol augmented Il1b and Il6 mRNA levels both in pWAT and eWAT. These results suggest that an increased sympathetic activity enhances NEFA and cytokine load of the adipose microenvironment, including that of pancreatic fat, and by doing so it may alter beta-cell function.

Spectroscopic investigation of the electronic and excited state properties of para-substituted tetraphenyl porphyrins and their electrochemically generated ions.

Porphyrins play pivotal roles in many crucial biological processes including photosynthesis. However, there is still a knowledge gap in understanding electronic and excited state implications associated with functionalization of the porphyrin ring system. These effects can have electrochemical and spectroscopic signatures that reveal the complex nature of these somewhat minor substitutions, beyond simple inductive or electronic effect correlations. To obtain a deeper insight into the influences of porphyrin functionalization, four free-base, meso-substituted porphyrins: tetraphenyl porphyrin (TPP), tetra(4-hydroxyphenyl) porphyrin (THPP), tetra(4-carboxyphenyl) porphyrin (TCPP), and tetra(4-nitrophenyl) porphyrin (TNPP), were synthesized, characterized, and investigated. The influence of various substituents, (-hydroxy,-carboxy, and -nitro) in the para position of the meso-substituted phenyl moieties were evaluated by spectroelectrochemical techniques (absorption and fluorescence), femtosecond transient absorption spectroscopy, cyclic and differential pulse voltammetry, ultraviolet photoelectron spectroscopy (UPS), and time-dependent density functional theory (TD-DFT). Spectral features were evaluated for the neutral porphyrins and differences observed among the various porphyrins were further explained using rendered frontier molecular orbitals pertaining to the relevant transitions. Electrochemically generated anionic and cationic porphyrin species indicate similar absorbance spectroscopic signatures attributed to a red-shift in the Soret band. Emissive behavior reveals the emergence of one new fluorescence decay pathway for the ionic porphyrin, distinct from the neutral macrocycle. Femtosecond transient absorption spectroscopy analysis provided further analysis of the implications on the excited-state as a function of the para substituent of the free-base meso-substituted tetraphenyl porphyrins. Herein, we provide an in-depth and comprehensive analysis of the electronic and excited state effects associated with systematically varying the induced dipole at the methine bridge of the free-base porphyrin macrocycle and the spectroscopic signatures related to the neutral, anionic, and cationic species of these porphyrins.

Metabolic implications of pancreatic fat accumulation.

Fat accumulation outside subcutaneous adipose tissue often has unfavourable effects on systemic metabolism. In addition to non-alcoholic fatty liver disease, which has received considerable attention, pancreatic fat has become an important area of research throughout the past 10 years. While a number of diagnostic approaches are available to quantify pancreatic fat, multi-echo Dixon MRI is currently the most developed method. Initial studies have shown associations between pancreatic fat and the metabolic syndrome, impaired glucose metabolism and type 2 diabetes mellitus. Pancreatic fat is linked to reduced insulin secretion, at least under specific circumstances such as prediabetes, low BMI and increased genetic risk of type 2 diabetes mellitus. This Review summarizes the possible causes and metabolic consequences of pancreatic fat accumulation. In addition, potential therapeutic approaches for addressing pancreatic fat accumulation are discussed.

Femtosecond photodecarbonylation of photo-ODIBO studied by stimulated Raman spectroscopy and density functional theory.

Photo-oxa-dibenzocyclooctyne (Photo-ODIBO) undergoes photodecarbonylation under UV excitation to its bright S2 state, forming a highly reactive cyclooctyne, ODIBO. Following 321 nm excitation with sub-50 fs actinic pulses, the excited state evolution and cyclopropenone bond cleavage with CO release were characterized using femtosecond stimulated Raman spectroscopy and time-dependent density functional theory Raman calculations. Analysis of the photo-ODIBO S2 CO Raman band revealed multi-exponential intensity, peak splitting and frequency-shift dynamics. This suggests a stepwise cleavage of the two C-C bonds in the cyclopropenone structure that is completed within <300 fs after excitation. Evidence of intramolecular vibrational relaxation on the S2 state, concurrent with photodecarbonylation, with dynamics matching previous electronic transient absorption spectroscopy, was also observed. This confirms an excited state, as opposed to ground state, photodecarbonylation mechanism resulting in a vibronically excited photoproduct, ODIBO.

The hepatokine fetuin-A disrupts functional maturation of pancreatic beta cells.

AIMS/HYPOTHESIS: Neonatal beta cells carry out a programme of postnatal functional maturation to achieve full glucose responsiveness. A partial loss of the mature phenotype of adult beta cells may contribute to a reduction of functional beta cell mass and accelerate the onset of type 2 diabetes. We previously found that fetuin-A, a hepatokine increasingly secreted by the fatty liver and a determinant of type 2 diabetes, inhibits glucose-stimulated insulin secretion (GSIS) of human islets. Since fetuin-A is a ubiquitous fetal glycoprotein that declines peripartum, we examined here whether fetuin-A interferes with the functional maturity of beta cells. METHODS: The effects of fetuin-A were assessed during in vitro maturation of porcine neonatal islet cell clusters (NICCs) and in adult human islets. Expression alterations were examined via microarray, RNA sequencing and reverse transcription quantitative real-time PCR (qRT-PCR), proteins were analysed by western blotting and immunostaining, and insulin secretion was quantified in static incubations. RESULTS: NICC maturation was accompanied by the gain of glucose-responsive insulin secretion (twofold stimulation), backed up by mRNA upregulation of genes governing beta cell identity and function, such as NEUROD1, UCN3, ABCC8 and CASR (Log2 fold change [Log2FC] > 1.6). An active TGFß receptor (TGFBR)-SMAD2/3 pathway facilitates NICC maturation, since the TGFBR inhibitor SB431542 counteracted the upregulation of aforementioned genes and de-repressed ALDOB, a gene disallowed in mature beta cells. In fetuin-A-treated NICCs, upregulation of beta cell markers and the onset of glucose responsiveness were suppressed. Concomitantly, SMAD2/3 phosphorylation was inhibited. Transcriptome analysis confirmed inhibitory effects of fetuin-A and SB431542 on TGFß-1- and SMAD2/3-regulated transcription. However, contrary to SB431542 and regardless of cMYC upregulation, fetuin-A inhibited beta cell proliferation (0.27 ± 0.08% vs 1.0 ± 0.1% Ki67-positive cells in control NICCs). This effect was sustained by reduced expression (Log2FC ≤ -2.4) of FOXM1, CENPA, CDK1 or TOP2A. In agreement, the number of insulin-positive cells was lower in fetuin-A-treated NICCs than in control NICCs (14.4 ± 1.2% and 22.3 ± 1.1%, respectively). In adult human islets fetuin-A abolished glucose responsiveness, i.e. 1.7- and 1.1-fold change over 2.8 mmol/l glucose in control- and fetuin-A-cultured islets, respectively. In addition, fetuin-A reduced SMAD2/3 phosphorylation and suppressed expression of proliferative genes. Of note, in non-diabetic humans, plasma fetuin-A was negatively correlated (p = 0.013) with islet beta cell area. CONCLUSIONS/INTERPRETATION: Our results suggest that the perinatal decline of fetuin-A relieves TGFBR signalling in islets, a process that facilitates functional maturation of neonatal beta cells. Functional maturity remains revocable in later life, and the occurrence of a metabolically unhealthy milieu, such as liver steatosis and elevated plasma fetuin-A, can impair both function and adaptive proliferation of beta cells. DATA AVAILABILITY: The RNAseq datasets and computer code produced in this study are available in the Gene Expression Omnibus (GEO): GSE144950; https://www.ncbi.nlm.nih.gov/geo/query/acc.cgi?acc=GSE144950.

Pancreatic fat cells of humans with type 2 diabetes display reduced adipogenic and lipolytic activity.

Obesity, especially visceral fat accumulation, increases the risk of type 2 diabetes (T2D). The purpose of this study was to investigate the impact of T2D on the pancreatic fat depot. Pancreatic fat pads from 17 partial pancreatectomized patients (PPP) were collected, pancreatic preadipocytes isolated, and in vitro differentiated. Patients were grouped using HbA1c into normal glucose tolerant (NGT), prediabetic (PD), and T2D. Transcriptome profiles of preadipocytes and adipocytes were assessed by RNAseq. Insulin sensitivity was estimated by quantifying AKT phosphorylation on Western blots. Lipogenic capacity was assessed with oil red O staining, lipolytic activity via fatty acid release. Secreted factors were measured using ELISA. Comparative transcriptome analysis of preadipocytes and adipocytes indicates defective upregulation of genes governing adipogenesis (NR1H3), lipogenesis (FASN, SCD, ELOVL6, and FADS1), and lipolysis (LIPE) during differentiation of cells from T2D-PPP. In addition, the ratio of leptin/adiponectin mRNA was higher in T2D than in NGT-PPP. Preadipocytes and adipocytes of NGT-PPP were more insulin sensitive than T2D-PPP cells in regard to AKT phosphorylation. Triglyceride accumulation was similar in NGT and T2D adipocytes. Despite a high expression of the receptors NPR1 and NPR2 in NGT and T2D adipocytes, lipolysis was stimulated by ANP 1.74-fold in NGT cells only. This stimulation was further increased by the PDE5 inhibitor dipyridamole (3.09-fold). Dipyridamole and forskolin increased lipolysis receptor independently 1.88-fold and 1.48-fold, respectively, solely in NGT cells. In conclusion, the metabolic status persistently affects differentiation and lipolysis of pancreatic adipocytes. These alterations could aggravate the development of T2D.

Ultrafast transient absorption spectroscopy of the photodecarbonylation of photo-oxadibenzocyclooctyne (photo-ODIBO).

The ultrafast dynamics of photo-OxaDiBenzocycloOctyne (photo-ODIBO) photo-dissociation was studied using femtosecond transient absorption spectroscopy. Steady-state UV-Vis, time-dependent density functional theory, and 350 nm and 321 nm transient absorption studies are reported. Photo-ODIBO excitation with 321 nm and 350 nm light-induced photodecarbonylation of the cyclopropenone functional group results in the formation of ODIBO. The presence of the photoproduct was confirmed by the results of steady-state photolysis experiments and the observation of absorption signatures of ODIBO in the photo-ODIBO transient absorption spectra. Analysis of the latter revealed the underlying photochemical mechanisms and associated time constants, following excitation of the samples. The dynamics show a multi-exponential decay process, following the dissociation of photo-ODIBO into an excited state of the photoproduct ODIBO within <294 fs after 321 nm excitation. 350 nm excitation, on the other hand, is shown to produce ground state ODIBO via an intermediate species. Additional transient absorption measurements were performed directly on the photoproduct ODIBO to help distinguish spectral signatures associated with these processes.

FFA2-, but not FFA3-agonists inhibit GSIS of human pseudoislets: a comparative study with mouse islets and rat INS-1E cells.

The expression of short chain fatty acid receptors FFA2 and FFA3 in pancreatic islets raised interest in using them as drug targets for treating hyperglycemia in humans. This study aims to examine the efficacy of synthetic FFA2- and FFA3-ligands to modulate glucose-stimulated insulin secretion (GSIS) in human pseudoislets which display intact glucose responsiveness. The FFA2-agonists 4-CMTB and TUG-1375 inhibited GSIS, an effect reversed by the FFA2-antagonist CATPB. GSIS itself was not augmented by CATPB. The FFA3-agonists FHQC and 1-MCPC did not affect GSIS in human pseudoislets. For further drug evaluation we used mouse islets. The CATPB-sensitive inhibitory effect of 100 µM 4-CMTB on GSIS was recapitulated. The inhibition was partially sensitive to the Gi/o-protein inhibitor pertussis toxin. A previously described FFA2-dependent increase of GSIS was observed with lower concentrations of 4-CMTB (10 and 30 µM). The stimulatory effect of 4-CMTB on secretion was prevented by the Gq-protein inhibitor FR900359. As in human pseudoislets, in mouse islets relative mRNA levels were FFAR2 > FFAR3 and FFA3-agonists did not affect GSIS. The FFA3-agonists, however, inhibited GSIS in a pertussis toxin-sensitive manner in INS-1E cells and this correlated with relative mRNA levels of Ffar3 > > Ffar2. Thus, in humans, when FFA2-activation impedes GSIS, FFA2-antagonism may reduce glycemia.

Pancreatic Steatosis Associates With Impaired Insulin Secretion in Genetically Predisposed Individuals.

CONTEXT: Pancreatic steatosis leading to beta-cell failure might be involved in type 2 diabetes (T2D) pathogenesis. OBJECTIVE: We hypothesized that the genetic background modulates the effect of pancreatic fat on beta-cell function and investigated genotype × pancreatic fat interactions on insulin secretion. DESIGN: Two observational studies. SETTING: University hospital. PATIENTS OR PARTICIPANTS: A total of 360 nondiabetic individuals with elevated risk for T2D (Tuebingen Family Study [TUEF]), and 64 patients undergoing pancreatectomy (Pancreas Biobank [PB], HbA1c <9%, no insulin therapy). MAIN OUTCOME MEASURES: Insulin secretion calculated from 5-point oral glucose tolerance test (TUEF) and fasting blood collection before surgery (PB). A genome-wide polygenic score for T2D was computed from 484,788 genotyped variants. The interaction of magnetic resonance imaging-measured and histologically quantified pancreatic fat with the polygenic score was investigated. Partitioned risk scores using genome-wide significant variants were also computed to gain insight into potential mechanisms. RESULTS: Pancreatic steatosis interacted with genome-wide polygenic score on insulin secretion (P = 0.003), which was similar in the replication cohort with histological measurements (P = 0.03). There was a negative association between pancreatic fat and insulin secretion in participants with high genetic risk, whereas individuals with low genetic risk showed a positive correlation between pancreatic fat and insulin secretion. Consistent interactions were found with insulin resistance-specific and a liver/lipid-specific polygenic scores. CONCLUSIONS: The associations suggest that pancreatic steatosis only impairs beta-cell function in subjects at high genetic risk for diabetes. Genetically determined insulin resistance specifically renders pancreatic fat deleterious for insulin secretion.

Internal conversion and intersystem crossing dynamics of uracil upon double thionation: a time-resolved photoelectron spectroscopy study in the gas phase.

The photophysical properties of 2,4-dithiouracil (2,4-DTU) in the gas phase are studied by time-resolved photoelectron spectroscopy (TRPES) with three different excitation wavelengths in direct extension of previous work on uracil (U), 2-thiouracil (2-TU) and 4-thiouracil (4-TU). Non-radiative deactivation in the canonical nucleobases like uracil mainly occurs via internal conversion (IC) along singlet excited states, although intersystem crossing (ISC) to a long-lived triplet state was confirmed to play a minor role. In thionated uracils, ISC to the triplet state becomes ultrafast and highly efficient with a quantum yield near unity; however, the lifetime of the triplet state is strongly dependent on the position of the sulfur atom. In 2-TU, ISC back to the ground state occurs within a few hundred picoseconds, whereas the population remains trapped in the lowest triplet state in the case of 4-TU. Upon doubling the degree of thionation, ISC remains highly efficient and dominates the photophysics of 2,4-DTU. However, several low-lying excited states contribute to competing IC and ISC pathways and a complex deactivation mechanism, which is evaluated here based on TRPES measurements and discussed in the context of the singly thionated uracils.

Glucose Responsiveness of ß-Cells Depends on Fatty Acids.

Glucose-stimulated insulin secretion (GSIS) is the gold standard for ß-cell function. Both experimental and clinical diabetology, i. e., preceding transplantation of isolated human islets, depend on functional testing. However, multiple factors influence GSIS rendering the comparison of different in vitro tests of glucose responsiveness difficult. This study examined the influence of bovine serum albumin (BSA)-coupled fatty acids on GSIS. Isolated islet preparations of human donors and of 12-months old mice displayed impaired GSIS in the presence of 0.5% FFA-free BSA compared to 0.5% BSA (fraction V, not deprived from fatty acids). In aged INS-1E cells, i. e. at a high passage number, GSIS became highly sensitive to FFA-free BSA. Readdition of 30 µM palmitate or 30 µM oleate to FFA-free BSA did not rescue GSIS, while the addition of 100 µM palmitate and the raise of extracellular Ca2+from 1.3 to 2.6 mM improved glucose responsiveness. A high concentration of palmitate (600 µM), which fully activates FFA1, largely restored insulin secretion. The FFA1-agonist TUG-469 also increased insulin secretion but to a lesser extent than palmitate. Glucose- and TUG-induced Ca2+oscillations were impaired in glucose-unresponsive, i. e., aged INS-1E cells. These results suggest that fatty acid deprivation (FFA-free BSA) impairs GSIS mainly through an effect on Ca2+sensitivity.

Glucose homeostasis is regulated by pancreatic ß-cell cilia via endosomal EphA-processing.

Diabetes mellitus affects one in eleven adults worldwide. Most suffer from Type 2 Diabetes which features elevated blood glucose levels and an inability to adequately secrete or respond to insulin. Insulin producing ß-cells have primary cilia which are implicated in the regulation of glucose metabolism, insulin signaling and secretion. To better understand how ß-cell cilia affect glucose handling, we ablate cilia from mature ß-cells by deleting key cilia component Ift88. Here we report that glucose homeostasis and insulin secretion deteriorate over 12 weeks post-induction. Cilia/basal body components are required to suppress spontaneous auto-activation of EphA3 and hyper-phosphorylation of EphA receptors inhibits insulin secretion. In ß-cells, loss of cilia/basal body function leads to polarity defects and epithelial-to-mesenchymal transition. Defective insulin secretion from IFT88-depleted human islets and elevated pEPHA3 in islets from diabetic donors both point to a role for cilia/basal body proteins in human glucose homeostasis.

Glucose, adrenaline and palmitate antagonistically regulate insulin and glucagon secretion in human pseudoislets.

Isolated human islets do not always meet the quality standards required for transplant survival and reliable functional in vitro studies. The formation of pseudoislets, i.e. the reaggregation of a defined number of islet cells after dissociation, improves insulin secretion. We present a simple method of pseudoislet formation from human islet cells and assess the transcriptome and function of isolated human islets and pseudoislets from the same organ donors. Following pseudoislet formation, insulin content/DNA and mRNA/RPS13 resembled that of islets. In pseudoislets, glucose-stimulated insulin secretion (GSIS) was significantly higher (8-13-fold) than in islets (2-4-fold). GSIS of pseudoislets was partly inhibited by the glucagon-like peptide-1 receptor (GLP-1R) antagonist exendin-9. The stimulatory effects of palmitate and forskolin at 12 mM glucose were also significantly higher in pseudoislets than in islets. Further analysis of pseudoislets revealed that regulation of secretion and insulin and glucagon content was maintained over a longer culture period (6-14 d). While adrenaline inhibited GSIS, adrenaline together with palmitate stimulated glucagon secretion 2-fold at low glucose, an effect suppressed by high glucose. Transcriptome analysis revealed that, unlike islets, pseudoislets were deprived of exocrine and endothelial cells. In conclusion, pseudoislet formation restores functional integrity of human islet cells and allows long-term in vitro testing.

What role do fat cells play in pancreatic tissue?

BACKGROUND: It is now generally accepted that obesity is a major risk factor for type 2 diabetes mellitus (T2DM). Hepatic steatosis in particular, as well as visceral and ectopic fat accumulation within tissues, is associated with the development of the disease. We recently presented the first study on isolated human pancreatic adipocytes and their interaction with islets [Gerst, F., Wagner, R., Kaiser, G., Panse, M., Heni, M., Machann, J., et al., 2017. Metabolic crosstalk between fatty pancreas and fatty liver: effects on local inflammation and insulin secretion. Diabetologia 60(11):2240-2251.]. The results indicate that the function of adipocytes depends on the overall metabolic status in humans which, in turn, differentially affects islet hormone release. SCOPE OF REVIEW: This review summarizes former and recent studies on factors derived from adipocytes and their effects on insulin-secreting ß-cells, with particular emphasis on the human pancreas. The adipocyte secretome is discussed with a special focus on its influence on insulin secretion, ß-cell survival and apoptotic ß-cell death. MAJOR CONCLUSIONS: Human pancreatic adipocytes store lipids and release adipokines, metabolites, and pro-inflammatory molecules in response to the overall metabolic, humoral, and neuronal status. The differentially regulated adipocyte secretome impacts on endocrine function, i.e., insulin secretion, ß-cell survival and death which interferes with glycemic control. This review attempts to explain why the extent of pancreatic steatosis is associated with reduced insulin secretion in some studies but not in others.