Major depressive disorder (MDD) is a severe disease of unknown pathogenesis with a lifetime prevalence of ~10%. Therapy requires prolonged treatment that often fails. We have previously demonstrated that ceramide levels in the blood plasma of patients and in mice with experimental MDD are increased. Neutralization of blood plasma ceramide prevented experimental MDD in mice. Mechanistically, we demonstrated that blood plasma ceramide accumulated in endothelial cells of the hippocampus, inhibited phospholipase D (PLD) and thereby decreased phosphatidic acid in the hippocampus. Here, we demonstrate that phosphatidic acid binds to and controls the activity of phosphotyrosine phosphatase (PTP1B) in the hippocampus and thus determines tyrosine phosphorylation of a variety of cellular proteins including TrkB. Injection of PLD, phosphatidic acid, or inhibition of PTP1B abrogated MDD and normalized cellular tyrosine phosphorylation, including phosphorylation of TrkB and neurogenesis in the hippocampus. Most importantly, these treatments also rapidly normalized behavior of mice with experimental MDD. Since phosphatidic acid binds to and inhibits PTP1B, the lack of phosphatidic acid results in increased activity of PTP1B and thereby in reduced tyrosine phosphorylation of TrkB and other cellular proteins. Thus, our data indicate a novel pathogenetic mechanism of and a rapidly acting targeted treatment for MDD.
Interferon-gamma (IFN-γ) is a pleiotropic cytokine that plays a critical role in mediating an array of immune responses including promotes antiviral activity, facilitates macrophage activation, controls Th1/Th2 balance, and regulates cellular apoptosis and proliferation. A few articles have previously reviewed the effects of IFN-γ in the regulation of barrier permeability, but none of these articles focuses on barrier function of endothelial cells. This review aims to discuss the regulatory mechanisms of IFN-γ on endothelial barrier function and its underlying signaling pathways. Articles were retrieved from electronic databases such as PubMed and Google Scholar using keywords "Interferon-gamma", "endothelial cells", "barrier function", and "signaling pathway". The articles published between 2000 and 2022 that are related to the aforementioned topics were selected. A few journals published beyond this period were also included due to limited information available. The results showed that IFN-γ modulates endothelial barrier function, mainly involves small GTPases, STAT1-dependent pathway, p38 MAPK and nitric oxide. In conclusion, more in depth cellular and molecular studies are needed to elucidate the pathways of IFN-γ in the regulation of endothelial barrier function.
Endothelial cells line the inner surface of all blood and lymphatic vessels, creating a semi-permeable barrier regulating fluid and solute exchange between blood or lymph and their surrounding tissues. The ability of a virus to cross the endothelial barrier is an important mechanism that facilitates virus dissemination in the human body. Many viruses are reported to alter endothelial permeability and/or cause endothelial cell barrier disruption during infection, which is able to cause vascular leakage. The current study describes a real-time cell analysis (RTCA) protocol, using a commercial real-time cell analyzer to monitor endothelial integrity and permeability changes during Zika virus (ZIKV) infection of the human umbilical vein endothelial cells (HUVECs). The impedance signals recorded before and after ZIKV infection were translated to cell index (CI) values and analyzed. The RTCA protocol allows the detection of transient effects in the form of cell morphological changes during a viral infection. This assay could also be useful for studying changes in the vascular integrity of HUVECs in other experimental setups.
Matched MeSH terms: Human Umbilical Vein Endothelial Cells/metabolism
The tumor microenvironment (TME) is well-defined target for understanding tumor progression and various cell types. Major elements of the tumor microenvironment are the followings: endothelial cells, fibroblasts, signaling molecules, extracellular matrix, and infiltrating immune cells. MicroRNAs (miRNAs) are a group of small noncoding RNAs with major functions in the gene expression regulation at post-transcriptional level that have also appeared to exerts key functions in the cancer initiation/progression in diverse biological processes and the tumor microenvironment. This study summarized various roles of miRNAs in the complex interactions between the tumor and normal cells in their microenvironment.
Plasma leakage in severe dengue has been postulated to be associated with skewed cytokine immune responses. In this study, the association of cytokines with vascular permeability in dengue patients was investigated. Human serum samples collected from 48 persons (13 with dengue fever, 29 with dengue hemorrhagic fever, and 6 healthy) were subjected to cytokines analysis by using Luminex Multiplex Technology. Selected serum samples from patients with dengue hemorrhagic fever sera and recombinant human cytokines were then tested for roles on inducing vascular permeability by treatment of human umbilical vein endothelial cells. Confocal immunofluorescence staining indicated morphologic alteration of human umbilical vein endothelial cells treated with serum samples from patients with dengue hemorrhagic fever compared with serum samples from healthy persons. The findings suggest that cytokines produced during dengue hemorrhagic infections could induce alterations in the vascular endothelium, which may play a fundamental role in the pathophysiology of dengue.
The aim of this protocol presents an optimized procedure for the purification and cultivation of pBECs and to establish in vitro blood-brain barrier (BBB) models based on pBECs in mono-culture (MC), MC with astrocyte-conditioned medium (ACM), and non-contact co-culture (NCC) with astrocytes of porcine or rat origin. pBECs were isolated and cultured from fragments of capillaries from the brain cortices of domestic pigs 5-6 months old. These fragments were purified by careful removal of meninges, isolation and homogenization of grey matter, filtration, enzymatic digestion, and centrifugation. To further eliminate contaminating cells, the capillary fragments were cultured with puromycin-containing medium. When 60-95% confluent, pBECs growing from the capillary fragments were passaged to permeable membrane filter inserts and established in the models. To increase barrier tightness and BBB characteristic phenotype of pBECs, the cells were treated with the following differentiation factors: membrane permeant 8-CPT-cAMP (here abbreviated cAMP), hydrocortisone, and a phosphodiesterase inhibitor, RO-20-1724 (RO). The procedure was carried out over a period of 9-11 days, and when establishing the NCC model, the astrocytes were cultured 2-8 weeks in advance. Adherence to the described procedures in the protocol has allowed the establishment of endothelial layers with highly restricted paracellular permeability, with the NCC model showing an average transendothelial electrical resistance (TEER) of 1249 ± 80 Ω cm2, and paracellular permeability (Papp) for Lucifer Yellow of 0.90 10-6 ± 0.13 10-6 cm sec-1 (mean ± SEM, n=55). Further evaluation of this pBEC phenotype showed good expression of the tight junctional proteins claudin 5, ZO-1, occludin and adherens junction protein p120 catenin. The model presented can be used for a range of studies of the BBB in health and disease and, with the highly restrictive paracellular permeability, this model is suitable for studies of transport and intracellular trafficking.
Endothelial dysfunction in diabetes is generally attributed to oxidative stress, but this view is challenged by observations showing antioxidants do not eliminate diabetic vasculopathy. As an alternative to oxidative stress-induced dysfunction, we interrogated if impaired mitochondrial function in endothelial cells is central to endothelial dysfunction in the metabolic syndrome. We observed reduced coronary arteriolar vasodilation to the endothelium-dependent dilator, acetylcholine (Ach), in Zucker Obese Fatty rats (ZOF, 34 ± 15% [mean ± standard deviation] 10-3 M) compared to Zucker Lean rats (ZLN, 98 ± 11%). This reduction in dilation occurred concomitantly with mitochondrial DNA (mtDNA) strand lesions and reduced mitochondrial complex activities in the endothelium of ZOF versus ZLN. To demonstrate endothelial dysfunction is linked to impaired mitochondrial function, administration of a cell-permeable, mitochondria-directed endonuclease (mt-tat-EndoIII), to repair oxidatively modified DNA in ZOF, restored mitochondrial function and vasodilation to Ach (94 ± 13%). Conversely, administration of a cell-permeable, mitochondria-directed exonuclease (mt-tat-ExoIII) produced mtDNA strand breaks in ZLN, reduced mitochondrial complex activities and vasodilation to Ach in ZLN (42 ± 16%). To demonstrate that mitochondrial function is central to endothelium-dependent vasodilation, we introduced (via electroporation) liver mitochondria (from ZLN) into the endothelium of a mesenteric vessel from ZOF and restored endothelium-dependent dilation to vasoactive intestinal peptide (VIP at 10-5 M, 4 ± 3% vasodilation before mitochondrial transfer and 48 ± 36% after transfer). Finally, to demonstrate mitochondrial function is key to endothelium-dependent dilation, we administered oligomycin (mitochondrial ATP synthase inhibitor) and observed a reduction in endothelium-dependent dilation. We conclude that mitochondrial function is critical for endothelium-dependent vasodilation.
Inflammation drives coronary artery disease and atherosclerosis implications. Lipoprotein entry, retention, and oxidative modification cause endothelial damage, triggering innate and adaptive immune responses. Recruited immune cells orchestrate the early atherosclerotic lesions by releasing proinflammatory cytokines, expediting the foam cell formation, intraplaque haemorrhage, secretion of matrix-degrading enzymes, and lesion progression, eventually promoting coronary artery syndrome via various inflammatory cascades. In addition, soluble mediators disrupt the dynamic anti- and prothrombotic balance maintained by endothelial cells and pave the way for coronary artery disease such as angina pectoris. Recent studies have established a relationship between elevated levels of inflammatory markers, including C-reactive protein (CRP), interleukins (IL-6, IL-1β), and tumour necrosis factor-alpha (TNF-α) with the severity of CAD and the possibility of future cardiovascular events. High-sensitivity C-reactive protein (hs-CRP) is a marker for assessing systemic inflammation and predicting the risk of developing CAD based on its peak plasma levels. Hence, understanding cross-talk interactions of inflammation, atherogenesis, and CAD is highly warranted to recalculate the risk factors that activate and propagate arterial lesions and devise therapeutic strategies accordingly. Cholesterol-inflammation lowering agents (statins), monoclonal antibodies targeting IL-1 and IL-6 (canakinumab and tocilizumab), disease-modifying antirheumatic drugs (methotrexate), sodium-glucose transport protein-2 (SGLT2) inhibitors, colchicine and xanthene oxidase inhibitor (allopurinol) have shown promising results in reducing inflammation, regressing atherogenic plaque and modifying the course of CAD. Here, we review the complex interplay between inflammatory, endothelial, smooth muscle and foam cells. Moreover, the putative role of inflammation in atherosclerotic CAD, underlying mechanisms and potential therapeutic implications are also discussed herein.
In vitro blood-brain barrier (BBB) models from primary brain endothelial cells can closely resemble the in vivo BBB, offering valuable models to assay BBB functions and to screen potential central nervous system drugs. We have recently developed an in vitro BBB model using primary porcine brain endothelial cells. The model shows expression of tight junction proteins and high transendothelial electrical resistance, evidence for a restrictive paracellular pathway. Validation studies using small drug-like compounds demonstrated functional uptake and efflux transporters, showing the suitability of the model to assay drug permeability. However, one limitation of in vitro model permeability measurement is the presence of the aqueous boundary layer (ABL) resulting from inefficient stirring during the permeability assay. The ABL can be a rate-limiting step in permeation, particularly for lipophilic compounds, causing underestimation of the permeability. If the ABL effect is ignored, the permeability measured in vitro will not reflect the permeability in vivo. To address the issue, we explored the combination of in vitro permeability measurement using our porcine model with the pKa(FLUX) method in pCEL-X software to correct for the ABL effect and allow a detailed analysis of in vitro (transendothelial) permeability data, Papp. Published Papp using porcine models generated by our group and other groups are also analyzed. From the Papp, intrinsic transcellular permeability (P0) is derived by simultaneous refinement using a weighted nonlinear regression, taking into account permeability through the ABL, paracellular permeability and filter restrictions on permeation. The in vitro P0 derived for 22 compounds (35 measurements) showed good correlation with P0 derived from in situ brain perfusion data (r(2)=0.61). The analysis also gave evidence for carrier-mediated uptake of naloxone, propranolol and vinblastine. The combination of the in vitro porcine model and the software analysis provides a useful tool to better predict BBB permeability in vivo and gain better mechanistic information about BBB permeation.
Nitric oxide (NO), produced by endothelial nitric oxide synthase (eNOS), is a major antiatherogenic factor in the blood vessel. Oxidative stress plays an important role in the pathogenesis of various cardiovascular diseases, including atherosclerosis. Decreased availability of endothelial NO promotes the progression of endothelial dysfunction and atherosclerosis. Rutin is a flavonoid with multiple cardiovascular protective effects. This study aimed to investigate the effects of rutin on eNOS and NO production in cultured human umbilical vein endothelial cells (HUVEC). HUVEC were divided into four groups: control; oxidative stress induction with 180 μM H₂O₂; treatment with 300 μM rutin; and concomitant induction with rutin and H₂O₂ for 24 hours. HUVEC treated with rutin produced higher amount of NO compared to control (P < 0.01). In the oxidative stress-induced HUVEC, rutin successfully induced cells' NO production (P < 0.01). Rutin promoted NO production in HUVEC by inducing eNOS gene expression (P < 0.05), eNOS protein synthesis (P < 0.01), and eNOS activity (P < 0.05). Treatment with rutin also led to increased gene and protein expression of basic fibroblast growth factor (bFGF) in HUVEC. Therefore, upregulation of eNOS expression by rutin may be mediated by bFGF. The results showed that rutin may improve endothelial function by augmenting NO production in human endothelial cells.
Matched MeSH terms: Human Umbilical Vein Endothelial Cells/metabolism*
The effects of spaceflight on cardiovascular health are not necessarily seen immediately after astronauts have returned but can be delayed. It is important to investigate the long term effects of spaceflight on protein and gene expression of inflammation and endothelial activation as a predictor for the development of atherosclerosis and potential cardiovascular problems. The objectives of this study were to investigate the (a) protein and gene expression of inflammation and endothelial activation, (b) expression of nuclear factor kappa B (NFκB), signal transducer and activator of transcription-3 (STAT-3) and endothelial nitric oxide synthase (eNOS) in human umbilical vein endothelial cells (HUVEC) 3 months post-space flight travel compared to ground controls. HUVEC cultured on microcarriers in fluid processing apparatus were flown to the International Space Station (ISS) by the Soyuz TMA-11 rocket. After landing, the cells were detached from microcarriers and recultured in T-25 cm(2) culture flasks (Revived HUVEC). Soluble protein expression of IL-6, TNF-α, ICAM-1, VCAM-1 and e-selectin were measured by ELISA. Gene expression of these markers and in addition NFκB, STAT-3 and eNOS were measured. Spaceflight induced IL-6 and ICAM-1 remain elevated even after 3 months post spaceflight travel and this is mediated via STAT-3 pathway. The downregulation of eNOS expression in revived HUVEC cells suggests a reduced protection of the cells and the surrounding vessels against future insults that may lead to atherosclerosis. It would be crucial to explore preventive measures, in relation to atherosclerosis and its related complications.
INTRODUCTION: This study aims to assess microvascular endothelial function in obese compared to age matched lean controls. Serum lipid profile, fasting glucose, high sensitivity C-reactive protein (hs-CRP) and adiponectin levels were also determined.
METHODS: This cross-sectional study involved 36 healthy lean and 36 obese subjects. Microvascular endothelial function was assessed using Laser Doppler fluximetry and iontophoresis with acetylcholine and sodium nitroprusside.
RESULTS: Mean age of subjects was 26.54 ± 0.60 years. Obese subjects had higher systolic (118.8 ± 1.5 vs 105.7 ± 2.0 mmHg, p < 0.001) and diastolic blood pressure (71.61 ± 1.35 vs 64.53 ± 1.40 mmHg, p = 0.001), higher triglyceride (1.35 ± 0.13 vs 0.79 ± 0.05 mmol/l, p < 0.001), lower high density lipoprotein cholesterol (HDL-C) (1.43 ± 0.04 vs 1.62 ± 0.05 mmol/l, p = 0.003), higher hs-CRP (11.58 ± 1.88 vs 1.88 ± 0.35 mg/l, p < 0.001), and lower adiponectin levels (8.80 ± 0.43 vs 25.93 ± 0.40 μg/ml, p < 0.001) compared to lean subjects. Endothelial dependent vasodilatation was lower in obese compared to lean subjects (40.53 ± 6.59 vs 71.03 ± 7.13 AU, p = 0.001).
CONCLUSION: Microvascular endothelial function is reduced in obese compared to age matched controls. This is associated with higher BP, triglyceride and lower HDL-C and adiponectin levels in obese group.
Study site: not mentioned
Previously, several aurone derivatives were identified with promising neuroprotective activities. In developing these compounds to target the central nervous system (CNS), an assessment of their blood-brain barrier (BBB) permeability was performed using in vitro BBB models: parallel artificial membrane permeability assay-BBB which measures passive permeability and primary porcine brain endothelial cell model which enables determination of the involvement of active transport mechanism. Parallel artificial membrane permeability assay-BBB identified most compounds with high passive permeability, with 3 aurones having exceptional Pevalues highlighting the importance of basic amine moieties and optimal lipophilicity for good passive permeability. Bidirectional permeability assays with porcine brain endothelial cell showed a significant net influx permeation of the aurones indicating a facilitated uptake mechanism in contrast to donepezil, a CNS drug included in the evaluation which only displayed passive permeation. From pH-dependent permeability assay coupled with data analysis using pCEL-X software, intrinsic transcellular permeability (Po) of a representative aurone 4-3 was determined, considering factors such as the aqueous boundary layer that may hinder accurate in vitro to in vivo correlation. The Po value determined supported the in vivo feasibility of the aurone as a CNS-active compound.
Glioblastoma (GBM) is characterized by aberrant vascularization and a complex tumor microenvironment. The failure of anti-angiogenic therapies suggests pathways of GBM neovascularization, possibly attributable to glioblastoma stem cells (GSCs) and their interplay with the tumor microenvironment. It has been established that GSC-derived extracellular vesicles (GSC-EVs) and their cargoes are proangiogenic in vitro. To further elucidate EV-mediated mechanisms of neovascularization in vitro, we perform RNA-seq and DNA methylation profiling of human brain endothelial cells exposed to GSC-EVs. To correlate these results to tumors in vivo, we perform histoepigenetic analysis of GBM molecular profiles in the TCGA collection. Remarkably, GSC-EVs and normal vascular growth factors stimulate highly distinct gene regulatory responses that converge on angiogenesis. The response to GSC-EVs shows a footprint of post-transcriptional gene silencing by EV-derived miRNAs. Our results provide insights into targetable angiogenesis pathways in GBM and miRNA candidates for liquid biopsy biomarkers.
Antibacterial coating is important to prevent the colonization of medical devices by biofilm forming bacteria that would cause infection and sepsis in patients. Current coating techniques such as immobilization of antimicrobial compounds, time-releasing antibiotic agents and silver nanoparticles, require multiple processing steps, and they have low efficacy and low stability. We proposed a single-step plasma polymerization of an essential oil known as carvone to produce a moderately hydrophobic antibacterial coating (ppCar) with an average roughness of <1nm. ppCar had a static water contact angle of 78°, even after 10days of air aging and it maintained its stability throughout 24h of LB broth immersion. ppCar showed promising results in the live-dead fluorescence assay and crystal violet assay. The biofilm assay showed an effective reduction of E. coli and S. aureus bacteria by 86% and 84% respectively. ppCar is also shown to rupture the bacteria membrane for its bactericidal effects. The cytotoxicity test indicated that the coating is not cytotoxic to the human cell line. This study would be of interest to researcher keen on producing a bacteria-resistance and biocompatible coating on different substrates in a cost-effective manner.
Matched MeSH terms: Human Umbilical Vein Endothelial Cells/metabolism*
Experimental autoimmune encephalomyelitis (EAE) is a classical animal model of human multiple sclerosis (MS) that is most commonly used to study the neuropathology and therapeutic effects of the disease. Telocytes (TCs) are a specialized type of interstitial or mesenchymal cell first identified by Popescu in various tissues and organs. However, the existence, distribution and role of CD34+ stromal cells (SCs)/TCs in the EAE-induced mouse spleen remain to be elucidated. We conducted immunohistochemistry, immunofluorescence (double staining for CD34 and c-kit, vimentin, F4/80, CD163, Nanog, Sca-1, CD31 or tryptase) and transmission electron microscopy experiments to investigate the existence, distribution and role of CD34+ SCs/TCs in the EAE-induced mouse spleen. Interestingly, immunohistochemistry, double-immunofluorescence, and transmission electron microscopy results revealed that CD34+ SCs/TCs were significantly upregulated in the EAE mouse spleen. Immunohistochemical or double-immunofluorescence staining of CD34+ SCs/TCs showed positive expression for CD34, c-kit, vimentin, CD34/vimentin, c-kit/vimentin and CD34/c-kit, and negative expression for CD31 and tryptase. Transmission electron microscopy (TEM) results demonstrated that CD34+ SCs/TCs established close connections with lymphocytes, reticular cells, macrophages, endothelial cells and erythrocytes. Furthermore, we also found that M1 (F4/80) or M2 (CD163) macrophages, and haematopoietic, pluripotent stem cells were markedly increased in EAE mice. Our results suggest that CD34+ SCs/TCs are abundant and may play a contributing role in modulating the immune response, recruiting macrophages and proliferation of haematopoietic and pluripotent stem cells following injury to promote tissue repair and regeneration in EAE mouse spleens. This suggests that their transplantation combined with stem cells might represent a promising therapeutic target for the treatment and prevention of multiple autoimmune and chronic inflammatory disorders.
Systemic infections of Candida albicans, the most prevalent fungal pathogen in humans, are on the rise in recent years. However, the exact mode of pathogenesis of this fungus is still not well elucidated. Previous studies using C. albicans mutants locked into the yeast form via gene deletion found that this form was avirulent and did not induce significant differential expression of host genes in vitro. In this study, a high density of C. albicans was used to infect human umbilical vein endothelial cells (HUVEC), resulting in yeast-form infections, whilst a low density of C. albicans resulted in hyphae infections. Transcriptional profiling of HUVEC response to these infections showed that high densities of C. albicans induced a stronger, broader transcriptional response from HUVEC than low densities of C. albicans infection. Many of the genes that were significantly differentially expressed were involved in apoptosis and cell death. In addition, conditioned media from the high-density infections caused a significant reduction in HUVEC viability, suggesting that certain molecules released during C. albicans and HUVEC interactions were capable of causing cell death. This study has shown that C. albicans yeast-forms, at high densities, cannot be dismissed as avirulent, but instead could possibly contribute to C. albicans pathogenesis.
Matched MeSH terms: Human Umbilical Vein Endothelial Cells/metabolism*
Endothelial cell death due to increased reactive oxygen species (ROS) may contribute to the initial endothelial injury, which promotes atherosclerotic lesion formation. Piper sarmentosum (PS), a natural product, has been shown to have an antioxidant property, which is hypothesized to inhibit production of ROS and prevent cell injury. Thus, the present study was designed to determine the effects of PS on the hydrogen peroxide (H(2)O(2))-induced oxidative cell damage in cultured human umbilical vein endothelial cells (HUVECs). In this experiment, HUVECs were obtained by collagenase perfusion of the large vein in the umbilical cord and cultured in medium M200 supplemented with low serum growth supplementation (LSGS). HUVECs were treated with various concentrations of H(2)O(2) (0-1000 micromol/L) and it was observed that 180 micromol/L H(2)O(2) reduced cell viability by 50% as denoted by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. Using the above concentration as the positive control, the H(2)O(2)-induced HUVECs were concomitantly treated with various concentrations (100, 150, 250 and 300 microg/ml) of three different extracts (aqueous, methanol and hexane) of PS. Malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPX) levels showed a significant increase (P<0.05) in HUVECs compared to the negative control. However, PS extracts showed a protective effect on HUVECs from H(2)O(2)-induced cell apoptosis with a significant reduction in MDA, SOD, CAT and GPX levels (P<0.05). Furthermore, PS had exhibited ferric reducing antioxidant power with its high phenolic content. Hence, it was concluded that PS plays a beneficial role in reducing oxidative stress in H(2)O(2)-induced HUVECs.
While age-related insulin resistance and hyperinsulinemia are usually considered to be secondary to changes in muscle, the liver also plays a key role in whole-body insulin handling and its role in age-related changes in insulin homeostasis is largely unknown. Here, we show that patent pores called 'fenestrations' are essential for insulin transfer across the liver sinusoidal endothelium and that age-related loss of fenestrations causes an impaired insulin clearance and hyperinsulinemia, induces hepatic insulin resistance, impairs hepatic insulin signaling, and deranges glucose homeostasis. To further define the role of fenestrations in hepatic insulin signaling without any of the long-term adaptive responses that occur with aging, we induced acute defenestration using poloxamer 407 (P407), and this replicated many of the age-related changes in hepatic glucose and insulin handling. Loss of fenestrations in the liver sinusoidal endothelium is a hallmark of aging that has previously been shown to cause deficits in hepatic drug and lipoprotein metabolism and now insulin. Liver defenestration thus provides a new mechanism that potentially contributes to age-related insulin resistance.