Significance of Zn2+ in RyR1 for Structural Integrity and Ligand Binding: Insight from Molecular Dynamics.

Ryanodine receptor type 1 (RyR1) is a Ca2+-release channel central to skeletal muscle excitation-contraction (EC) coupling. RyR1's cryo-EM structures reveal a zinc-finger motif positioned within the cytoplasmic C-terminal domain (CTD). Yet, owing to limitations in cryo-EM resolution, RyR1 structures lack precision in detailing the metal coordination structure, prompting the need for an accurate model. In this study, we employed molecular dynamics (MD) simulations and the density functional theory (DFT) method to refine the binding characteristics of Zn2+ in the zinc-finger site of the RyR1 channel. Our findings also highlight substantial conformational changes in simulations conducted in the absence of Zn2+. Notably, we observed a loss of contact at the interface between protein domains proximal to the zinc-finger site, indicating a crucial role of Zn2+ in maintaining structural integrity and interdomain interactions within RyR1. Furthermore, this study provides valuable insights into the modulation of ATP, Ca2+, and caffeine binding, shedding light on the intricate relationship between Zn2+ coordination and the dynamic behavior of RyR1. Our integrative approach combining MD simulations and DFT calculations enhances our understanding of the molecular mechanisms governing ligand binding in RyR1.

Association of Coffee Consumption and Prediagnostic Caffeine Metabolites With Incident Parkinson Disease in a Population-Based Cohort.

BACKGROUND AND OBJECTIVES: Inverse associations between caffeine intake and Parkinson disease (PD) have been frequently implicated in human studies. However, no studies have quantified biomarkers of caffeine intake years before PD onset and investigated whether and which caffeine metabolites are related to PD. METHODS: Associations between self-reported total coffee consumption and future PD risk were examined in the EPIC4PD study, a prospective population-based cohort including 6 European countries. Cases with PD were identified through medical records and reviewed by expert neurologists. Hazard ratios (HRs) and 95% CIs for coffee consumption and PD incidence were estimated using Cox proportional hazards models. A case-control study nested within the EPIC4PD was conducted, recruiting cases with incident PD and matching each case with a control by age, sex, study center, and fasting status at blood collection. Caffeine metabolites were quantified by high-resolution mass spectrometry in baseline collected plasma samples. Using conditional logistic regression models, odds ratios (ORs) and 95% CIs were estimated for caffeine metabolites and PD risk. RESULTS: In the EPIC4PD cohort (comprising 184,024 individuals), the multivariable-adjusted HR comparing the highest coffee intake with nonconsumers was 0.63 (95% CI 0.46-0.88, p = 0.006). In the nested case-control study, which included 351 cases with incident PD and 351 matched controls, prediagnostic caffeine and its primary metabolites, paraxanthine and theophylline, were inversely associated with PD risk. The ORs were 0.80 (95% CI 0.67-0.95, p = 0.009), 0.82 (95% CI 0.69-0.96, p = 0.015), and 0.78 (95% CI 0.65-0.93, p = 0.005), respectively. Adjusting for smoking and alcohol consumption did not substantially change these results. DISCUSSION: This study demonstrates that the neuroprotection of coffee on PD is attributed to caffeine and its metabolites by detailed quantification of plasma caffeine and its metabolites years before diagnosis.

Caffeine and naloxone treated mesenchymal stem cells improve symptoms and reduce inflammation in a mouse model of ulcerative colitis.

BACKGROUND AND OBJECTIVE: Ulcerative colitis (UC) causes ulcers in the colon and rectum, leading to abdominal pain, diarrhea, and rectal bleeding, and if left untreated, can lead to serious complications. The therapeutic effects of mesenchymal stem cells (MSCs) on experimental models of UC have been proven. Since the microenvironment around these cells is crucial in maintaining cell proliferation, differentiation, metabolism, and overall function, this study aims to evaluation the role of caffeine and naloxone as a new microenvironment for MSCs in reducing inflammation and improving symptoms in an experimental model of UC. MATERIAL AND METHOD: A group of 40 outbred NMRI mice were studied and divided randomly into four equal groups (N = 10 each group). UC was induced in all groups using acetic acid. The first group (control) was treated with phosphate buffer saline (PBS), the second group with MSCs-Caffeine, the third with MSCs-Naloxone, and the fourth with Mesalazine. The disease activity index (DAI), tissue damage, myeloperoxidase (MPO) activity, nitric oxide (NO) levels, and the production of IL-1, IL-6, and TNF-α cytokines were evaluated. RESULT: Our research demonstrated that all treatments were effective in improving the symptoms and reducing inflammatory markers in mice with colitis. Among the two MSCs treatments, the MSCs-Caffeine was found to be the most potent in reducing the levels of NO, IL-1, IL-6, tissue damage (P < 0.001) and as well as TNF-α (P < 0.0001) in compared to the control group. CONCLUSION: MSCs treated with caffeine and naloxone can enhance the immunoregulatory potential of these. As a result, treated MSCs can lead to improved clinical signs and reduced inflammatory parameters in mice with UC, making this approach a useful way for controlling and treating the disease. However, additional research is needed to access the mechanism behind the stronger immune system regulatory effects of treated MSCs in UC treatment.

Long-term abuse of caffeine sodium benzoate induces endothelial cells injury and leads to coagulation dysfunction.

Our hospital admitted a patient who had difficulty in coagulation even after blood replacement, and the patient had abused caffeine sodium benzoate (CSB) for more than 20 years. Hence, we aimed to explore whether CSB may cause dysfunction in vascular endothelial cells and its possible mechanism. Low, medium, and high concentrations of serum of long-term CSB intake patients were used to treat HUVECs, with LPS as the positive control. MTT and CCK8 were performed to verify CSB's damaging effect on HUVECs. The expression of ET-1, ICAM-1, VCAM-1, and E-selectin were measured by ELISA. TUNEL assay and Matrigel tube formation assay were carried out to detect apoptosis and angiogenesis of HUVECs. Flow cytometry was applied to analyze cell cycles and expression of CD11b, PDGF, and ICAM-1. Expression of PDGF-BB and PCNA were examined by western blot. The activation of MAPK signaling pathway was detected by qRT-PCR and western blot. Intracellular Ca2+ density was detected by fluorescent probes. CCK8 assay showed high concentration of CSB inhibited cell viability. Cell proliferation and angiogenesis were inhibited by CSB. ET-1, ICAM-1, VCAM-1, and E-selectin upregulated in CSB groups. CSB enhanced apoptosis of HUVECs. CD11b, ICAM-1 increased and PDGF reduced in CSB groups. The expression level and phosphorylation level of MEK, ERK, JUN, and p38 in MAPK pathway elevated in CSB groups. The expression of PCNA and PDGF-BB was suppressed by CSB. Intracellular Ca2+ intensity was increased by CSB. Abuse of CSB injured HUVECs and caused coagulation disorders.

Extrapolating the Coffee and Caffeine (1,3,7-Trimethylxanthine) Effects on Exercise and Metabolism-A Concise Review.

The consumption of coffee and caffeine (1,3,7-trimethylxanthine) is part of many cultures worldwide. Their properties include serving as a neurostimulant aid, enhancing energy substrate levels, and improving general exercise performance. Both present therapeutic effects that can also be used to control chronic and metabolic diseases due to four mechanisms: adenosine receptor antagonism, increased catecholamine concentrations, a phosphodiesterase inhibitor, and a stimulator of calcium-release channels. Despite the individual genetic variabilities, distinct mechanisms have been demonstrated to improve physical performance, thermogenesis, lipolysis, insulin sensitivity, and hormonal modulation. Thus, coffee consumption and caffeine supplementation may enhance physical and mental performance and may improve metabolic variables, reducing oxidative stress, inflammation, and insulin resistance. Current data reveal vital aspects of coffee and caffeine consumption in specific populations, although further studies are needed to define clinical interventions with caffeine in obesity and chronic conditions.

Molecular insight into binding behavior of caffeine with lactoferrin: Spectroscopic, molecular docking, and simulation study.

The majority of bioactive substances in the human diet come from polyphenols. Here, we use spectroscopy, molecular docking, molecular dynamics simulations, and in vitro digestion to look at the relationship between caffeine (CAF) and bovine lactoferrin (BLF). The correlation analysis of the CAF-BLF fluorescence quenching process revealed that the reaction was spontaneous and that the CAF-BLF fluorescence quenching process may have been static. The predominant intrinsic binding forces were hydrogen bonds and van der Waals forces, which were also supported by molecular docking and molecular dynamics simulations. Through Fourier infrared and circular dichroism spectroscopy experiments, it was found that CAF changed the secondary structure of BLF and might bind to the hydrophobic amino acids of BLF. Compared with BLF, CAF-BLF showed inhibitory effects on digestion in simulated in vitro digestion. It will be helpful to better understand the interaction between CAF and BLF and provide the basis for the development of innovative dairy products.

Phase I study to assess the effect of adavosertib (AZD1775) on the pharmacokinetics of substrates of CYP1A2, CYP2C19, and CYP3A in patients with advanced solid tumors.

PURPOSE: Adavosertib may alter exposure to substrates of the cytochrome P450 (CYP) family of enzymes. This study assessed its effect on the pharmacokinetics of a cocktail of probe substrates for CYP3A (midazolam), CYP2C19 (omeprazole), and CYP1A2 (caffeine). METHODS: Period 1: patients with locally advanced or metastatic solid tumors received 'cocktail': caffeine 200 mg, omeprazole 20 mg, and midazolam 2 mg (single dose); period 2: after 7- to 14-day washout, patients received adavosertib 225 mg twice daily on days 1-3 (five doses), with cocktail on day 3. After cocktail alone or in combination with adavosertib administration, 24-h pharmacokinetic sampling occurred for probe substrates and their respective metabolites paraxanthine, 5-hydroxyomeprazole (5-HO), and 1'-hydroxymidazolam (1'-HM). Safety was assessed throughout. RESULTS: Of 33 patients (median age 60.0 years, range 41-83) receiving cocktail, 30 received adavosertib. Adavosertib co-administration increased caffeine, omeprazole, and midazolam exposure by 49%, 80%, and 55% (AUC0-12), respectively; AUC0-t increased by 61%, 98%, and 55%. Maximum plasma drug concentration (Cmax) increased by 4%, 46%, and 39%. Adavosertib co-administration increased 5-HO and 1'-HM exposure by 43% and 54% (AUC0-12) and 49% and 58% (AUC0-t), respectively; paraxanthine exposure was unchanged. Adavosertib co-administration decreased Cmax for paraxanthine and 5-HO by 19% and 7%; Cmax increased by 33% for 1'-HM. After receiving adavosertib, 19 (63%) patients had treatment-related adverse events (six [20%] grade ≥ 3). CONCLUSION: Adavosertib (225 mg bid) is a weak inhibitor of CYP1A2, CYP2C19, and CYP3A. CLINICALTRIALS: GOV: NCT03333824.

Caffeine can alleviate non-alcoholic fatty liver disease by augmenting LDLR expression via targeting EGFR.

Increasing low-density lipoprotein receptor (LDLR) protein levels represents a key strategy for the prevention and treatment. Berberine can reportedly alleviate non-alcoholic fatty liver disease (NAFLD) by increasing the LDLR expression in an ERK1/2 signaling-dependent manner of NAFLD. Studies have shown that caffeine can inhibit fat deposition in the livers of mice; however, caffeine has not been reported to alleviate NAFLD by augmenting the LDLR expression via targeting EGFR. Here, an MTT assay, western blotting, RT-qPCR, immunohistochemistry, and surface plasmon resonance (SPR) analysis were used to investigate the role of caffeine in low-density lipoprotein cholesterol (LDL-C) clearance both in vitro and in vivo. In vitro, we found that caffeine could activate the EGFR-ERK1/2 signaling pathway in HepG2 cells, leading to increased LDLR mRNA and protein expression, and this effect could be inhibited by cetuximab. The SPR assay results have indicated that caffeine may increase the LDLR expression by directly binding to the EGFR extracellular domain and activating the EGFR-ERK1/2 signaling pathway. In vivo, caffeine markedly improved fatty liver and related blood indices in ApoE KO mice with high-fat-diet-induced NAFLD. Consistent with our in vitro results, we found that caffeine could also activate EGFR-ERK1/2 signaling and promote the LDLR expression in ApoE KO mice. In summary, caffeine can enhance the LDLR expression by directly binding to EGFR and activating the EGFR-ERK1/2 signaling pathway. EGFR signaling may represent a novel target for the prevention and treatment of NAFLD.

Caffeine protects against hippocampal alterations in type 2 diabetic rats via modulation of gliosis, inflammation and apoptosis.

Type 2 diabetes (T2D) is implicated in the injury of several organs, including the brain resulting in neuronal damage, which may lead to cognitive impairment and dementia. Additionally, it is linked to inflammation, cytokine release, apoptosis and various degenerative conditions. Astrocytes and microglia might have a role in mediating these processes. Caffeine, a psychoactive beverage, has been shown to reduce the risk of cognitive and memory impairment. This study proposes anti-inflammatory and anti-apoptotic role of caffeine, which can be mediated via microglia/astrocyte activation and overexpression of pro-inflammatory molecules. T2D was induced in rats by feeding with high fat high sugar diet and injecting a single low dose streptozotocin (STZ) intraperitoneally. Other diabetic rats were given caffeine orally (in two doses) for 5 weeks, starting 1 week before STZ injection. Measurement of plasma cytokines, TNFα and IL6, was performed using enzyme-linked immunosorbent assay (ELISA) technique. After sacrificing animals, brains were obtained and processed for histological evaluation. Immunohistochemistry was also performed using the following primary antibodies, anti-astrocyte marker GFAP, anti-microglia marker CD11b and apoptotic marker (anti-cleaved caspase-3). There was upregulation of IL6 and TNF-α in diabetic rats. Additionally, histological evaluation of the hippocampus of diabetic rats revealed cellular degeneration. There was increased immunostaining of GFAP, CD11b and cleaved caspase-3 in diabetic rats. Pretreatment with caffeine to diabetic rats, resulted in improvement of structural changes and decrease in cytokine levels and immuno-markers, expression, and this was in a dose-dependent manner. In conclusion, caffeine had an ameliorative role in enhancing hippocampal degenerative changes in T2D.

Coffee Intake, Caffeine Metabolism Genotype, and Survival Among Men with Prostate Cancer.

BACKGROUND: Coffee intake may lower prostate cancer risk and progression, but postdiagnosis outcomes by caffeine metabolism genotype are not well characterized. OBJECTIVE: To evaluate associations between coffee intake, caffeine metabolism genotype, and survival in a large, multicenter study of men with prostate cancer. DESIGN, SETTING, AND PARTICIPANTS: Data from The PRACTICAL Consortium database for 5727 men with prostate cancer from seven US, Australian, and European studies were included. The cases included had data available for the CYP1A2 -163C>A rs762551 single-nucleotide variant associated with caffeine metabolism, coffee intake, and >6 mo of follow-up. OUTCOME MEASUREMENTS AND STATISTICAL ANALYSIS: Multivariable-adjusted Cox proportional hazards models across pooled patient-level data were used to compare the effect of coffee intake (categorized as low [reference], high, or none/very low) in relation to overall survival (OS) and prostate cancer-specific survival (PCSS), with stratified analyses conducted by clinical disease risk and genotype. RESULTS AND LIMITATIONS: High coffee intake appeared to be associated with longer PCSS (hazard ratio [HR] 0.85, 95% confidence interval [CI] 0.68-1.08; p = 0.18) and OS (HR 0.90, 95% CI 0.77-1.07; p = 0.24), although results were not statistically significant. In the group with clinically localized disease, high coffee intake was associated with longer PCSS (HR 0.66, 95% CI 0.44-0.98; p = 0.040), with comparable results for the group with advanced disease (HR 0.92, 95% CI 0.69-1.23; p = 0.6). High coffee intake was associated with longer PCSS among men with the CYP1A2 AA (HR 0.67, 95% CI 0.49-0.93; p = 0.017) but not the AC/CC genotype (p = 0.8); an interaction was detected (p = 0.042). No associations with OS were observed in subgroup analyses (p > 0.05). Limitations include the nominal statistical significance and residual confounding. CONCLUSIONS: Coffee intake was associated with longer PCSS among men with a CYP1A2 -163AA (*1F/*1F) genotype, a finding that will require further replication. PATIENT SUMMARY: It is likely that coffee intake is associated with longer prostate cancer-specific survival in certain groups, but more research is needed to fully understand which men may benefit and why.

Changes in adenosine receptors and neurotrophic factors in the SOD1G93A mouse model of amyotrophic lateral sclerosis: Modulation by chronic caffeine.

Amyotrophic lateral sclerosis (ALS) is characterized by the progressive degeneration of corticospinal tract motor neurons. Previous studies showed that adenosine-mediated neuromodulation is disturbed in ALS and that vascular endothelial growth factor (VEGF) has a neuroprotective function in ALS mouse models. We evaluated how adenosine (A1R and A2AR) and VEGF (VEGFA, VEGFB, VEGFR-1 and VEGFR-2) system markers are altered in the cortex and spinal cord of pre-symptomatic and symptomatic SOD1G93A mice. We then assessed if/how chronic treatment of SOD1G93A mice with a widely consumed adenosine receptor antagonist, caffeine, modulates VEGF system and/or the levels of Brain-derived Neurotrophic Factor (BDNF), known to be under control of A2AR. We found out decreases in A1R and increases in A2AR levels even before disease onset. Concerning the VEGF system, we detected increases of VEGFB and VEGFR-2 levels in the spinal cord at pre-symptomatic stage, which reverses at the symptomatic stage, and decreases of VEGFA levels in the cortex, in very late disease states. Chronic treatment with caffeine rescued cortical A1R levels in SOD1G93A mice, bringing them to control levels, while rendering VEGF signaling nearly unaffected. In contrast, BDNF levels were significantly affected in SOD1G93A mice treated with caffeine, being decreased in the cortex and increased in spinal the cord. Altogether, these findings suggest an early dysfunction of the adenosinergic system in ALS and highlights the possibility that the negative influence of caffeine previously reported in ALS animal models results from interference with BDNF rather than with the VEGF signaling molecules.

Influence of Aerobic and Anaerobic Moist Incubation on Selected Nonvolatile Constituents─Comparison to Traditionally Fermented Cocoa Beans.

Recently, moist incubation has been proposed as an alternative postharvest processing method for cocoa beans. During this treatment, unfermented and dried cocoa nibs are rehydrated with a lactic acid solution containing ethanol and subsequently incubated for 72 h at 45 °C before drying. Previous studies focused on the aroma formation during this treatment and the further processing of chocolate. The current study focused on the influence of aerobic and anaerobic moist incubation on selected nonvolatile components in comparison with the unfermented raw material and traditionally fermented cocoa. Total phenolic content and total flavan-3-ol content, contents of (+)-catechin, (-)-epicatechin, procyanidins B2 and C1, cinnamtannin A2, methylxanthines (theobromine and caffeine), contents of sugars (sucrose, d-glucose, and d-fructose) and free amino acids (17 proteinogenic amino acids) were determined. The fermentation index was also evaluated. The aerobically incubated and fermented cocoa showed low levels of phenolic compounds in comparison to the unfermented cocoa and the anaerobically incubated cocoa. The level of methylxanthines was unaffected by all treatments. The contents of reducing sugars were more than 2-fold higher after both incubation treatments compared to fermentation. The level of free amino acids liberated was highest after anaerobic incubation followed by fermentation and aerobic incubation. The aerobically incubated cocoa showed the highest FI, while the anaerobically incubated cocoa may be considered under-fermented (FI <1.0). Statistical analysis (ANOVA) showed significant differences between all treatments, which was verified by principal component analysis.

Assessment of Cosmetic and Dermatological Properties and Safety of Use of Model Skin Tonics with Kombucha-Fermented Red Berry Extracts.

Kombucha is a health-promoting beverage that is produced by fermenting sweetened tea using symbiotic cultures of bacteria belonging to the genus Acetobacter, Gluconobacter, and yeast of the genus Saccharomyces. This study compared the cosmetic and dermatological properties of the extracts of the following redberries: R. rubrum, F. vesca, and R. idaeus, and their ferments, which were obtained by fermentation for 10 and 20 days using tea fungus. For this purpose, the fermented and non-fermented extracts were compared in terms of their chemical composition using the HPLC/ESI-MS chromatographic method, demonstrating the high content of biologically active compounds that were present in the ferments. The antioxidant activity of the tested samples was evaluated using DPPH and ABTS tests, as well as by evaluating the scavenging of the external and intracellular free radicals. The cytotoxicity of the extracts and the ferments, as well as the cosmetic formulations, were also determined by conducting Alamar Blue and Neutral Red tests assessing the cell viability and metabolism using skin cell lines: fibroblasts and keratinocytes. In addition, application tests were conducted showing the positive effects of the model cosmetic tonics on the TEWL, the skin hydration, and the skin pH. The results indicate that both the extracts and the ferments that were obtained from kombucha can be valuable ingredients in cosmetic products.

BMSCs overexpressed ISL1 reduces the apoptosis of islet cells through ANLN carrying exosome, INHBA, and caffeine.

Early apoptosis of grafted islets is one of the main factors affecting the efficacy of islet transplantation. The combined transplantation of islet cells and bone marrow mesenchymal stem cells (BMSCs) can significantly improve the survival rate of grafted islets. Transcription factor insulin gene enhancer binding protein 1 (ISL1) is shown to promote the angiogenesis of grafted islets and the paracrine function of mesenchymal stem cells during the co-transplantation, yet the regulatory mechanism remains unclear. By using ISL1-overexpressing BMSCs and the subtherapeutic doses of islets for co-transplantation, we managed to reduce the apoptosis and improve the survival rate of the grafts. Our metabolomics and proteomics data suggested that ISL1 upregulates aniline (ANLN) and Inhibin beta A chain (INHBA), and stimulated the release of caffeine in the BMSCs. We then demonstrated that the upregulation of ANLN and INHBA was achieved by the binding of ISL1 to the promoter regions of the two genes. In addition, ISL1 could also promote BMSCs to release exosomes with high expression of ANLN, secrete INHBA and caffeine, and reduce streptozocin (STZ)-induced islets apoptosis. Thus, our study provides mechanical insight into the islet/BMSCs co-transplantation and paves the foundation for using conditioned medium to mimic the ISL1-overexpressing BMSCs co-transplantation.

Thebromine Targets Adenosine Receptors to Control Hippocampal Neuronal Function and Damage.

Theobromine is a caffeine metabolite most abundant in dark chocolate, of which consumption is linked with a lower risk of cognitive decline. However, the mechanisms through which theobromine affects neuronal function remain ill-defined. Using electrophysiological recordings in mouse hippocampal synapses, we now characterized the impact of a realistic concentration of theobromine on synaptic transmission and plasticity. Theobromine (30 µM) facilitated synaptic transmission while decreasing the magnitude of long-term potentiation (LTP), with both effects being blunted by adenosine deaminase (2 U/mL). The pharmacological blockade of A1R with DPCPX (100 nM) eliminated the theobromine-dependent facilitation of synaptic transmission, whereas the A2AR antagonist SCH58261 (50 nM), as well as the genetic deletion of A2AR, abrogated the theobromine-induced impairment of LTP. Furthermore, theobromine prevented LTP deficits and neuronal loss, respectively, in mouse hippocampal slices and neuronal cultures exposed to Aß1-42 peptides, considered a culprit of Alzheimer's disease. Overall, these results indicate that theobromine affects information flow via the antagonism of adenosine receptors, normalizing synaptic plasticity and affording neuroprotection in dementia-related conditions in a manner similar to caffeine.

Bitter Peptides YFYPEL, VAPFPEVF, and YQEPVLGPVRGPFPIIV, Released during Gastric Digestion of Casein, Stimulate Mechanisms of Gastric Acid Secretion via Bitter Taste Receptors TAS2R16 and TAS2R38.

Eating satiating, protein-rich foods is one of the key aspects of modern diet, although a bitter off-taste often limits the application of some proteins and protein hydrolysates, especially in processed foods. Previous studies of our group demonstrated that bitter-tasting food constituents, such as caffeine, stimulate mechanisms of gastric acid secretion as a signal of gastric satiation and a key process of gastric protein digestion via activation of bitter taste receptors (TAS2Rs). Here, we tried to elucidate whether dietary non-bitter-tasting casein is intra-gastrically degraded into bitter peptides that stimulate mechanisms of gastric acid secretion in physiologically achievable concentrations. An in vitro model of gastric digestion was verified by casein-fed pigs, and the peptides resulting from gastric digestion were identified by liquid chromatography-time-of-flight-mass spectrometry. The bitterness of five selected casein-derived peptides was validated by sensory analyses and by an in vitro screening approach based on human gastric parietal cells (HGT-1). For three of these peptides (YFYPEL, VAPFPEVF, and YQEPVLGPVRGPFPIIV), an upregulation of gene expression of TAS2R16 and TAS2R38 was observed. The functional involvement of these TAS2Rs was verified by siRNA knock-down (kd) experiments in HGT-1 cells. This resulted in a reduction of the mean proton secretion promoted by the peptides by up to 86.3 ± 9.9% for TAS2R16kd (p < 0.0001) cells and by up to 62.8 ± 7.0% for TAS2R38kd (p < 0.0001) cells compared with mock-transfected cells.

Anesthetics and Cell-Cell Communication: Potential Ca2+-Calmodulin Role in Gap Junction Channel Gating by Heptanol, Halothane and Isoflurane.

Cell-cell communication via gap junction channels is known to be inhibited by the anesthetics heptanol, halothane and isoflurane; however, despite numerous studies, the mechanism of gap junction channel gating by anesthetics is still poorly understood. In the early nineties, we reported that gating by anesthetics is strongly potentiated by caffeine and theophylline and inhibited by 4-Aminopyridine. Neither Ca2+ channel blockers nor 3-isobutyl-1-methylxanthine (IBMX), forskolin, CPT-cAMP, 8Br-cGMP, adenosine, phorbol ester or H7 had significant effects on gating by anesthetics. In our publication, we concluded that neither cytosolic Ca2+i nor pHi were involved, and suggested a direct effect of anesthetics on gap junction channel proteins. However, while a direct effect cannot be excluded, based on the potentiating effect of caffeine and theophylline added to anesthetics and data published over the past three decades, we are now reconsidering our earlier interpretation and propose an alternative hypothesis that uncoupling by heptanol, halothane and isoflurane may actually result from a rise in cytosolic Ca2+ concentration ([Ca2+]i) and consequential activation of calmodulin linked to gap junction proteins.

Coffee peel extracts ameliorate non-alcoholic fatty liver disease via a fibroblast growth factor 21-adiponectin signaling pathway.

Coffee peel (CP) contains abundant phytochemicals which might prevent non-alcoholic fatty liver disease (NAFLD). The present study aimed to identify the main phytochemicals in CP extracts, and to investigate whether CP extracts could ameliorate NAFLD through a hepatic fibroblast growth factor (FGF) 21-adiponectin signaling pathway. Caffeine and seven monomers of flavonoids were identified from CP extracts by using high-performance liquid chromatography-tandem mass spectrometry (HPLC-MS/MS). After 8 weeks of intervention, the mice fed a high-fat and high-sugar diet showed the pathophysiological characteristics of NAFLD. Treatment with CP extracts significantly alleviated hepatic steatosis and insulin resistance and reduced the concentrations of serum alanine transaminase, FGF21, and triglyceride, and hepatic interleukin-6, interleukin-1ß, and tumor necrosis factor-α, while increasing serum adiponectin concentrations. Meanwhile, CP extract supplementation significantly decreased the gene and protein expression levels of FGF21, while enhancing adiponectin expression levels. The present study demonstrated that CP extracts contained caffeine and seven monomers of flavonoids, and protected against NAFLD through regulating the FGF21-adiponectin signaling pathway.

Caffeine Ameliorates AKT-Driven Nonalcoholic Steatohepatitis by Suppressing De Novo Lipogenesis and MyD88 Palmitoylation.

Dysregulated hepatic lipogenesis represents a promising druggable target for treating nonalcoholic steatohepatitis (NASH). This work aims to evaluate the therapeutic efficacy of caffeine in a NASH mouse model displaying increased hepatic lipogenesis driven by constitutive hepatic overexpression of the active v-akt murine thymoma viral oncogene homolog (AKT). Caffeine was administered in the AKT mice to study the efficacy in vivo. AKT-transfected and insulin-stimulated human hepatoma cells were used for in vitro experiments. The results demonstrated that caffeine ameliorated hepatic steatosis and inflammatory injury in vivo. Mechanistically, caffeine repressed the AKT/mTORC1 and SREBP-1/ACC/FASN signaling in mice and in vitro. Furthermore, caffeine impaired NF-κB activation by stabilizing IκBα, resulting in a reduction of proinflammatory mediators interleukin-6 (IL-6) and tumor necrosis factor α (TNF-α). Notably, caffeine abolished mTORC1/FASN-dependent MyD88 palmitoylation, which could be essential for its anti-inflammatory potential. Collectively, these results suggest that caffeine consumption could be advantageous in the prevention and therapy of NASH, especially in the subset accompanied by increased de novo lipogenesis.

Role of Caffeine in the Age-related Neurodegenerative Diseases: A Review.

Caffeine, a simple purine alkaloid with the proper chemical name 1,3,7-trimethylpurine- 2,6-dione, is an abundant compound present in coffee, food and drugs. It interacts with various pathways of which antagonism of adenosine receptors is the most significant but the other physiological pathways can be influenced by caffeine as well. Interaction with glutamate and dopamine neurotransmission pathways, competition with other substrates on cytochrome P450, non-competitive inhibition of acetylcholinesterase, blocking of nicotinic acetylcholine receptor and competitive inhibition of cyclic nucleotide phosphodiesterase can be mentioned. Because of caffeine availability in foods, beverages and drugs, it has practical relevance even if the effect is weak. Intake of coffee containing edibles for a long period or even for a substantial part of life makes caffeine´s impact significant. Low acute and chronic toxicity of caffeine is another important specification. The discoveries from the last few years point to the fact that caffeine would interfere with the progression of some age-related neurodegenerative disorders like Alzheimer's and Parkinson's diseases and dementia with Lewy bodies. In this review article, the recent findings about caffeine´s impact on neurodegenerative diseases are presented and important facts about the caffeine effect, including the substantial discoveries, are described.