Bone marrow cells contribute to seven different endothelial cell populations in the heart.

Understanding the mechanisms underlying vascular regeneration in the heart is crucial for developing novel therapeutic strategies for myocardial ischemia. This study investigates the contribution of bone marrow-derived cells to endothelial cell populations in the heart, and their role in cardiac function and coronary circulation following repetitive ischemia (RI). Chimeric rats were created by transplanting BM cells from GFP female rats into irradiated male recipients. After engraftment chimeras were subjected to RI for 17 days. Vascular growth was assessed from recovery of cardiac function and increases in myocardial blood flow during LAD occlusion. After sorting GFP+ BM cells from heart and bone of Control and RI rats, single-cell RNA sequencing was implemented to determine the fate of BM cells. Our in vivo RI model demonstrated an improvement in cardiac function and myocardial blood flow after 17 days of RI with increased capillary density in the rats subjected to RI compared to Controls. Single-cell RNA sequencing of bone marrow cells isolated from rats' hearts identified distinct endothelial cell (EC) subpopulations. These ECs exhibited heterogeneous gene expression profiles and were enriched for markers of capillary, artery, lymphatic, venous, and immune ECs. Furthermore, BM-derived ECs in the RI group showed an angiogenic profile, characterized by upregulated genes associated with blood vessel development and angiogenesis. This study elucidates the heterogeneity of bone marrow-derived endothelial cells in the heart and their response to repetitive ischemia, laying the groundwork for targeting specific subpopulations for therapeutic angiogenesis in myocardial ischemia.

The fatty acid omega hydroxylase genes (CYP4 family) in the progression of metabolic dysfunction-associated steatotic liver disease (MASLD): An RNA sequence database analysis and review.

Fatty acid omega hydroxylase P450s consist of enzymes that hydroxylate various chain-length saturated and unsaturated fatty acids (FAs) and bioactive eicosanoid lipids. The human cytochrome P450 gene 4 family (CYP4) consists of 12 members that are associated with several human diseases. However, their role in the progression of metabolic dysfunction-associated fatty liver disease (MASLD) remains largely unknown. It has long been thought that the induction of CYP4 family P450 during fasting and starvation prevents FA-related lipotoxicity through FA metabolism to dicarboxylic acids that are chain-shortened in peroxisomes and then transported to the mitochondria for complete oxidation. Several studies have revealed that peroxisome succinate transported to the mitochondria is used for gluconeogenesis during fasting and starvation, and recent evidence suggests that peroxisome acetate can be utilized for lipogenesis and lipid droplet formation as well as epigenetic modification of gene transcription. In addition, omega hydroxylation of the bioactive eicosanoid arachidonic acid to 20-Hydroxyeicosatetraenoic acid (20-HETE) is essential for activating the GPR75 receptor, leading to vasoconstriction and cell proliferation. Several mouse models of diet-induced MASLD have revealed the induction of selective CYP4A members and the suppression of CYP4F during steatosis and steatohepatitis, suggesting a critical metabolic role in the progression of fatty liver disease. Thus, to further investigate the functional roles of CYP4 genes, we analyzed the differential gene expression of 12 members of CYP4 gene family in datasets from the Gene Expression Omnibus (GEO) from patients with steatosis, steatohepatitis, fibrosis, cirrhosis, and hepatocellular carcinoma. We also observed the differential expression of various CYP4 genes in the progression of MASLD, indicating that different CYP4 members may have unique functional roles in the metabolism of specific FAs and eicosanoids at various stages of fatty liver disease. These results suggest that targeting selective members of the CYP4A family is a viable therapeutic approach for treating and managing MASLD.

Krüppel-like factor 10 protects against metabolic dysfunction-associated steatohepatitis by regulating HNF4α-mediated metabolic pathways.

BACKGROUND: Krüppel-like factor 10 (KLF10), a zinc finger transcription factor, plays a pivotal role in modulating TGF-ß-mediated cellular processes such as growth, apoptosis, and differentiation. Recent studies have implicated KLF10 in regulating lipid metabolism and glucose homeostasis. This study aimed to elucidate the precise role of hepatic KLF10 in developing metabolic dysfunction-associated steatohepatitis (MASH) in diet-induced obese mice. METHODS: We investigated hepatic KLF10 expression under metabolic stress and the effects of overexpression or ablation of hepatic KLF10 on MASH development and lipidemia. We also determined whether hepatocyte nuclear factor 4α (HNF4α) mediated the metabolic effects of KLF10. RESULTS: Hepatic KLF10 was downregulated in MASH patients and genetically or diet-induced obese mice. AAV8-mediated overexpression of KLF10 in hepatocytes prevented Western diet-induced hypercholesterolemia and steatohepatitis, whereas inactivation of hepatocyte KLF10 aggravated Western diet-induced steatohepatitis. Mechanistically, KLF10 reduced hepatic triglyceride and free fatty acid levels by inducing lipolysis and fatty acid oxidation and inhibiting lipogenesis, and reducing hepatic cholesterol levels by promoting bile acid synthesis. KLF10 highly induced HNF4α expression by directly binding to its promoter. The beneficial effect of KLF10 on MASH development was abolished in mice lacking hepatocyte HNF4α. In addition, the inactivation of KLF10 in hepatic stellate cells exacerbated Western diet-induced liver fibrosis by activating the TGF-ß/SMAD2/3 pathway. CONCLUSIONS: Our data collectively suggest that the transcription factor KLF10 plays a hepatoprotective role in MASH development by inducing HNF4α. Targeting hepatic KLF10 may offer a promising strategy for treating MASH.

Hepatic FOXA3 overexpression prevents Western diet-induced obesity and MASH through TGR5.

Forkhead transcription factor 3 (FOXA3) has been shown to regulate metabolism and development. Hepatic FOXA3 is reduced in obesity and fatty liver disease. However, the role of hepatic FOXA3 in regulating obesity or steatohepatitis remains to be investigated. In this work, C57BL/6 mice were i.v. injected with AAV8-ALB-FOXA3 or the control virus. The mice were then fed a chow or Western diet for 16 weeks. The role of hepatic FOXA3 in energy metabolism and steatohepatitis was investigated. Plasma bile acid composition and the role of Takeda G protein-coupled receptor 5 (TGR5) in mediating the metabolic effects of FOXA3 were determined. Overexpression of hepatic FOXA3 reduced hepatic steatosis in chow-fed mice and attenuated Western diet-induced obesity and steatohepatitis. FOXA3 induced lipolysis and inhibited hepatic genes involved in bile acid uptake, resulting in elevated plasma bile acids. The beneficial effects of hepatic FOXA3 overexpression on Western diet-induced obesity and steatohepatitis were abolished in Tgr5-/- mice. Our data demonstrate that overexpression of hepatic FOXA3 prevents Western diet-induced obesity and steatohepatitis via activation of TGR5.

Is miR-21 A Therapeutic Target in Cardiovascular Disease?

microRNA-21 (miR-21) serves a multitude of functions at the molecular level through its regulation of messenger RNA. Previous research has sparked interest in the role of miR-21 as a potential therapeutic target in cardiovascular diseases. miR-21 expression contributes to the differentiation, proliferation, and maturation of many cell types, such as fibroblasts, endothelial cells, cardiomyocytes, and endothelial progenitor cells. The function of miR-21 depends upon its expression level in the specific cell types and downstream targets, which determine cell fate. Under pathological conditions, the expression level of miR-21 is altered, leading to abnormal gene regulation of downstream signaling and cardiovascular diseases such as hypertension, cardiac hypertrophy and fibrosis, atherosclerosis, and heart failure. Agomirs or antagomirs can be introduced into the respective tissue type to reverse or stop the progression of the disease. Exosomes in the extracellular vesicles, which mediate many cellular events with high biocompatibility, have a high potential of efficiently delivering miR-21 to their targeted cells. The critical role of miR-21 in cardiovascular disease (CVD) is indisputable, but there are controversial reports on the function of miR-21 in the same disease. This discrepancy sparks interest in better understanding the role of miR-21 in different tissues under different stages of various diseases and the mechanism of how miR-21 inhibitors work.

Hepatocyte Sirtuin 6 Protects against Atherosclerosis and Steatohepatitis by Regulating Lipid Homeostasis.

Histone deacetylase Sirtuin 6 (SIRT6) regulates many biological processes. SIRT6 is known to regulate hepatic lipid metabolism and inhibit the development of nonalcoholic fatty liver disease (NAFLD). We aimed to investigate the role of hepatocyte SIRT6 in the development of atherosclerosis and further characterize the mechanism underlying SIRT6's effect on NAFLD. Ldlr-/- mice overexpressing or lacking hepatocyte SIRT6 were fed a Western diet for 16 weeks. The role of hepatic SIRT6 in the development of nonalcoholic steatohepatitis (NASH), atherosclerosis, and obesity was investigated. We also investigated whether p53 participates in the pathogenesis of NAFLD in mice overexpressing hepatic SIRT6. Our data show that loss of hepatocyte SIRT6 aggravated the development of NAFLD, atherosclerosis, and obesity in Ldlr-/- mice, whereas adeno-associated virus (AAV)-mediated overexpression of human SIRT6 in the liver had opposite effects. Mechanistically, hepatocyte SIRT6 likely inhibited the development of NAFLD by inhibiting lipogenesis, lipid droplet formation, and p53 signaling. Hepatocyte SIRT6 also likely inhibited the development of atherosclerosis by inhibiting intestinal lipid absorption and hepatic VLDL secretion. Hepatic SIRT6 also increased energy expenditure. In conclusion, our data indicate that hepatocyte SIRT6 protects against atherosclerosis, NAFLD, and obesity by regulating lipid metabolism in the liver and intestine.

Takotsubo syndrome is a coronary microvascular disease: experimental evidence.

BACKGROUND AND AIMS: Takotsubo syndrome (TTS) is a conundrum without consensus about the cause. In a murine model of coronary microvascular dysfunction (CMD), abnormalities in myocardial perfusion played a key role in the development of TTS. METHODS AND RESULTS: Vascular Kv1.5 channels connect coronary blood flow to myocardial metabolism and their deletion mimics the phenotype of CMD. To determine if TTS is related to CMD, wild-type (WT), Kv1.5-/-, and TgKv1.5-/- (Kv1.5-/- with smooth muscle-specific expression Kv1.5 channels) mice were studied following transaortic constriction (TAC). Measurements of left ventricular (LV) fractional shortening (FS) in base and apex, and myocardial blood flow (MBF) were completed with standard and contrast echocardiography. Ribonucleic Acid deep sequencing was performed on LV apex and base from WT and Kv1.5-/- (control and TAC). Changes in gene expression were confirmed by real-time-polymerase chain reaction. MBF was increased with chromonar or by smooth muscle expression of Kv1.5 channels in the TgKv1.5-/-. TAC-induced systolic apical ballooning in Kv1.5-/-, shown as negative FS (P < 0.05 vs. base), which was not observed in WT, Kv1.5-/- with chromonar, or TgKv1.5-/-. Following TAC in Kv1.5-/-, MBF was lower in LV apex than in base. Increasing MBF with either chromonar or in TgKv1.5-/- normalized perfusion and function between LV apex and base (P = NS). Some genetic changes during TTS were reversed by chromonar, suggesting these were independent of TAC and more related to TTS. CONCLUSION: Abnormalities in flow regulation between the LV apex and base cause TTS. When perfusion is normalized between the two regions, normal ventricular function is restored.

The Roles of Bone Marrow-Derived Stem Cells in Coronary Collateral Growth Induced by Repetitive Ischemia.

Many clinical trials have attempted to use stem cells to treat ischemic heart diseases (IHD), but the benefits have been modest. Though coronary collaterals can be a "natural bypass" for IHD patients, the regulation of coronary collateral growth (CCG) and the role of endogenous stem cells in CCG are not fully understood. In this study, we used a bone marrow transplantation scheme to study the role of bone marrow stem cells (BMSCs) in a rat model of CCG. Transgenic GFP rats were used to trace BMSCs after transplantation; GFP bone marrow was harvested or sorted for bone marrow transplantation. After recovering from transplantation, the recipient rats underwent 10 days of repetitive ischemia (RI), with echocardiography before and after RI, to measure cardiac function and myocardial blood flow. At the end of RI, the rats were sacrificed for the collection of bone marrow for flow cytometry or heart tissue for imaging analysis. Our study shows that upon RI stimulation, BMSCs homed to the recipient rat hearts' collateral-dependent zone (CZ), proliferated, differentiated into endothelial cells, and engrafted in the vascular wall for collateral growth. These RI-induced collaterals improved coronary blood flow and cardiac function in the recipients' hearts during ischemia. Depletion of donor CD34+ BMSCs led to impaired CCG in the recipient rats, indicating that this cell population is essential to the process. Overall, these results show that BMSCs contribute to CCG and suggest that regulation of the function of BMSCs to promote CCG might be a potential therapeutic approach for IHD.

Role of endothelial CXCR4 in the development of aortic valve stenosis.

Background: CXCL12/CXCR4 signaling is essential in cardiac development and repair, however, its contribution to aortic valve stenosis (AVS) remains unclear. In this study, we tested the role of endothelial CXCR4 on the development of AVS. Materials and methods: We generated CXCR4 endothelial cell-specific knockout mice (EC CXCR4 KO) by crossing CXCR4fl/fl mice with Tie2-Cre mice to study the role of endothelial cell CXCR4 in AVS. CXCR4fl/fl mice were used as controls. Echocardiography was used to assess the aortic valve and cardiac function. Heart samples containing the aortic valve were stained using Alizarin Red for detection of calcification. Masson's trichrome staining was used for the detection of fibrosis. The apex of the heart samples was stained with wheat germ agglutinin (WGA) to visualize ventricular hypertrophy. Results: Compared with the control group, the deletion of CXCR4 in endothelial cells led to significantly increased aortic valve peak velocity and aortic valve peak pressure gradient, with decreased aortic valve area and ejection fraction. EC CXCR4 KO mice also developed cardiac hypertrophy as evidenced by increased diastolic and systolic left ventricle posterior wall thickness (LVPW), cardiac myocyte size, and heart weight (HW) to body weight (BW) ratio. Our data also confirmed increased microcalcifications, interstitial fibrosis, and thickened valvular leaflets of the EC CXCR4 KO mice. Conclusion: The data collected throughout this study suggest the deletion of CXCR4 in endothelial cells is linked to the development of aortic valve stenosis and left ventricular hypertrophy. The statistically significant parameters measured indicate that endothelial cell CXCR4 plays an important role in aortic valve development and function. We have compiled compelling evidence that EC CXCR4 KO mice can be used as a novel model for AVS.

The essential role for endothelial cell sprouting in coronary collateral growth.

RATIONALE: Coronary collateral growth is a natural bypass for ischemic heart diseases. It offers tremendous therapeutic benefit, but the process of coronary collateral growth isincompletely understood due to limited preclinical murine models that would enable interrogation of its mechanisms and processes via genetic modification and lineage tracing. Understanding the processes by which coronary collaterals develop can unlock new therapeutic strategies for ischemic heart disease. OBJECTIVE: To develop a murine model of coronary collateral growth by repetitive ischemia and investigate whether capillary endothelial cells could contribute to the coronary collateral formation in an adult mouse heart after repetitive ischemia by lineage tracing. METHODS AND RESULTS: A murine model of coronary collateral growth was developed using short episodes of repetitive ischemia. Repetitive ischemia stimulation resulted in robust collateral growth in adult mouse hearts, validated by high-resolution micro-computed tomography. Repetitive ischemia-induced collateral formation compensated ischemia caused by occlusion of the left anterior descending artery. Cardiac function improved during ischemia after repetitive ischemia, suggesting the improvement of coronary blood flow. A capillary-specific Cre driver (Apln-CreER) was used for lineage tracing capillary endothelial cells. ROSA mT/mG reporter mice crossed with the Apln-CreER transgene mice underwent a 17 days' repetitive ischemia protocol for coronary collateral growth. Two-photon and confocal microscopy imaging of heart slices revealed repetitive ischemia-induced coronary collateral growth initiated from sprouting Apelin+ endothelial cells. Newly formed capillaries in the collateral-dependent zone expanded in diameter upon repetitive ischemia stimulation and arterialized with smooth muscle cell recruitment, forming mature coronary arteries. Notably, pre-existing coronary arteries and arterioles were not Apelin+, and all Apelin+ collaterals arose from sprouting capillaries. Cxcr4, Vegfr2, Jag1, Mcp1, and Hif1⍺ mRNA levels in the repetitive ischemia-induced hearts were also upregulated at the early stage of coronary collateral growth, suggesting angiogenic signaling pathways are activated for coronary collaterals formation during repetitive ischemia. CONCLUSIONS: We developed a murine model of coronary collateral growth induced by repetitive ischemia. Our lineage tracing study shows that sprouting endothelial cells contribute to coronary collateral growth in adult mouse hearts. For the first time, sprouting angiogenesis is shown to give rise to mature coronary arteries in response to repetitive ischemia in the adult mouse hearts.

Mitochondrial DNA integrity and function are critical for endothelium-dependent vasodilation in rats with metabolic syndrome.

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.

Mechanism of the switch from NO to H2O2 in endothelium-dependent vasodilation in diabetes.

Coronary microvascular dysfunction is prevalent among people with diabetes and is correlated with cardiac mortality. Compromised endothelial-dependent dilation (EDD) is an early event in the progression of diabetes, but its mechanisms remain incompletely understood. Nitric oxide (NO) is the major endothelium-dependent vasodilatory metabolite in the healthy coronary circulation, but this switches to hydrogen peroxide (H2O2) in coronary artery disease (CAD) patients. Because diabetes is a significant risk factor for CAD, we hypothesized that a similar NO-to-H2O2 switch would occur in diabetes. Vasodilation was measured ex vivo in isolated coronary arteries from wild type (WT) and microRNA-21 (miR-21) null mice on a chow or high-fat/high-sugar diet, and B6.BKS(D)-Leprdb/J (db/db) mice using myography. Myocardial blood flow (MBF), blood pressure, and heart rate were measured in vivo using contrast echocardiography and a solid-state pressure sensor catheter. RNA from coronary arteries, endothelial cells, and cardiac tissues was analyzed via quantitative real-time PCR for gene expression, and cardiac protein expression was assessed via western blot analyses. Superoxide was detected via electron paramagnetic resonance. (1) Ex vivo coronary EDD and in vivo MBF were impaired in diabetic mice. (2) Nω-Nitro-L-arginine methyl ester, an NO synthase inhibitor (L-NAME), inhibited ex vivo coronary EDD and in vivo MBF in WT. In contrast, polyethylene glycol-catalase, an H2O2 scavenger (Peg-Cat), inhibited diabetic mouse EDD ex vivo and MBF in vivo. (3) miR-21 was upregulated in diabetic mouse endothelial cells, and the deficiency of miR-21 prevented the NO-to-H2O2 switch and ameliorated diabetic mouse vasodilation impairments. (4) Diabetic mice displayed increased serum NO and H2O2, upregulated mRNA expression of Sod1, Sod2, iNos, and Cav1, and downregulated Pgc-1α in coronary arteries, but the deficiency of miR-21 reversed these changes. (5) miR-21-deficient mice exhibited increased cardiac PGC-1α, PPARα and eNOS protein and reduced endothelial superoxide. (6) Inhibition of PGC-1α changed the mRNA expression of genes regulated by miR-21, and overexpression of PGC-1α decreased the expression of miR-21 in high (25.5 mM) glucose treated coronary endothelial cells. Diabetic mice exhibit a NO-to-H2O2 switch in the mediator of coronary EDD, which contributes to microvascular dysfunction and is mediated by miR-21. This study represents the first mouse model recapitulating the NO-to-H2O2 switch seen in CAD patients in diabetes.

Hepatocytic Activating Transcription Factor 3 Protects Against Steatohepatitis via Hepatocyte Nuclear Factor 4α.

Activating transcription factor 3 (ATF3) has been shown to play an important role in HDL metabolism; yet, the role of hepatocytic ATF3 in the development of steatohepatitis remains elusive. Here we show that adenoassociated virus-mediated overexpression of human ATF3 in hepatocytes prevents diet-induced steatohepatitis in C57BL/6 mice and reverses steatohepatitis in db/db mice. Conversely, global or hepatocyte-specific loss of ATF3 aggravates diet-induced steatohepatitis. Mechanistically, hepatocytic ATF3 induces hepatic lipolysis and fatty acid oxidation and inhibits inflammation and apoptosis. We further show that hepatocyte nuclear factor 4α (HNF4α) is required for ATF3 to improve steatohepatitis. Thus, the current study indicates that ATF3 protects against steatohepatitis through, at least in part, hepatic HNF4α. Targeting hepatic ATF3 may be useful for treatment of steatohepatitis.

Hepatocyte miR-34a is a key regulator in the development and progression of non-alcoholic fatty liver disease.

OBJECTIVE: Hepatic miR-34a expression is elevated in diet-induced or genetically obese mice and patients with non-alcoholic steatohepatitis (NASH), yet hepatocyte miR-34a's role in the progression of non-alcoholic fatty liver disease (NAFLD) from non-alcoholic fatty liver (NAFL) to NASH remains to be elucidated. METHODS: Mice overexpressing or deficient in hepatocyte miR-34a and control mice were fed a diet enriched in fats, cholesterol, and fructose (HFCF) to induce NASH. C57BL/6 mice with NASH were treated with an miR-34a inhibitor or a scramble control oligo. The effect of miR-34a on the development, progression, and reversal of NAFLD was determined. RESULTS: The hepatocyte-specific expression of miR-34a aggravated HFCF diet-induced NAFLD. In contrast, germline or adult-onset deletion of hepatocyte miR-34a attenuated the development and progression of NAFLD. In addition, pharmacological inhibition of miR-34a reversed HFCF diet-induced steatohepatitis. Mechanistically, hepatocyte miR-34a regulated the development and progression of NAFLD by inducing lipid absorption, lipogenesis, inflammation, and apoptosis but inhibiting fatty acid oxidation. CONCLUSIONS: Hepatocyte miR-34a is an important regulator in the development and progression of NAFLD. MiR-34a may be a useful target for treating NAFLD.

Endothelial TRPV4 channels prevent tumor growth and metastasis via modulation of tumor angiogenesis and vascular integrity.

Transient receptor potential vanilloid 4 (TRPV4) is a ubiquitously expressed polymodally activated ion channel. TRPV4 has been implicated in tumor progression; however, the cell-specific role of TRPV4 in tumor growth, angiogenesis, and metastasis is unknown. Here, we generated endothelial-specific TRPV4 knockout (TRPV4ECKO) mice by crossing TRPV4lox/lox mice with Tie2-Cre mice. Tumor growth and metastasis were significantly increased in a syngeneic Lewis lung carcinoma tumor model of TRPV4ECKO mice compared to TRPV4lox/lox mice. Multiphoton microscopy, dextran leakage, and immunohistochemical analysis revealed increased tumor angiogenesis and metastasis that were correlated with aberrant leaky vessels (increased width and reduced pericyte and VE-cadherin coverage). Mechanistically, increases in VEGFR2, p-ERK, and MMP-9 expression and DQ gelatinase activity were observed in the TRPV4ECKO mouse tumors. Our results demonstrated that endothelial TRPV4 is a critical modulator of vascular integrity and tumor angiogenesis and that deletion of TRPV4 promotes tumor angiogenesis, growth, and metastasis.

Hepatocyte ATF3 protects against atherosclerosis by regulating HDL and bile acid metabolism.

Activating transcription factor (ATF)3 is known to have an anti-inflammatory function, yet the role of hepatic ATF3 in lipoprotein metabolism or atherosclerosis remains unknown. Here we show that overexpression of human ATF3 in hepatocytes reduces the development of atherosclerosis in Western-diet-fed Ldlr-/- or Apoe-/- mice, whereas hepatocyte-specific ablation of Atf3 has the opposite effect. We further show that hepatic ATF3 expression is inhibited by hydrocortisone. Mechanistically, hepatocyte ATF3 enhances high-density lipoprotein (HDL) uptake, inhibits intestinal fat and cholesterol absorption and promotes macrophage reverse cholesterol transport by inducing scavenger receptor group B type 1 (SR-BI) and repressing cholesterol 12α-hydroxylase (CYP8B1) in the liver through its interaction with p53 and hepatocyte nuclear factor 4α, respectively. Our data demonstrate that hepatocyte ATF3 is a key regulator of HDL and bile acid metabolism and atherosclerosis.

Hepatocyte-specific expression of human carboxylesterase 2 attenuates nonalcoholic steatohepatitis in mice.

Human carboxylesterase 2 (CES2) has triacylglycerol hydrolase (TGH) activities and plays an important role in lipolysis. In this study, we aim to determine the role of human CES2 in the progression or reversal of steatohepatitis in diet-induced or genetically obese mice. High-fat/high-cholesterol/high-fructose (HFCF) diet-fed C57BL/6 mice or db/db mice were intravenously injected with an adeno-associated virus expressing human CES2 under the control of an albumin promoter. Human CES2 protected against HFCF diet-induced nonalcoholic fatty liver disease (NAFLD) in C57BL/6J mice and reversed steatohepatitis in db/db mice. Human CES2 also improved glucose tolerance and insulin sensitivity. Mechanistically, human CES2 reduced hepatic triglyceride (T) and free fatty acid (FFA) levels by inducing lipolysis and fatty acid oxidation and inhibiting lipogenesis via suppression of sterol regulatory element-binding protein 1. Furthermore, human CES2 overexpression improved mitochondrial respiration and glycolytic function, and inhibited gluconeogenesis, lipid peroxidation, apoptosis, and inflammation. Our data suggest that hepatocyte-specific expression of human CES2 prevents and reverses steatohepatitis. Targeting hepatic CES2 may be an attractive strategy for treatment of NAFLD.NEW & NOTEWORTHY Human CES2 attenuates high-fat/cholesterol/fructose diet-induced steatohepatitis and reverses steatohepatitis in db/db mice. Mechanistically, human CES2 induces lipolysis, fatty acid and glucose oxidation, and inhibits hepatic glucose production, inflammation, lipid oxidation, and apoptosis. Our data suggest that human CES2 may be targeted for treatment of non-alcoholic steatohepatitis (NASH).

Hepatocyte Nuclear Factor 4α Prevents the Steatosis-to-NASH Progression by Regulating p53 and Bile Acid Signaling (in mice).

BACKGROUND AND AIMS: Hepatocyte nuclear factor 4α (HNF4α) is highly enriched in the liver, but its role in the progression of nonalcoholic liver steatosis (NAFL) to NASH has not been elucidated. In this study, we investigated the effect of gain or loss of HNF4α function on the development and progression of NAFLD in mice. APPROACH AND RESULTS: Overexpression of human HNF4α protected against high-fat/cholesterol/fructose (HFCF) diet-induced steatohepatitis, whereas loss of Hnf4α had opposite effects. HNF4α prevented hepatic triglyceride accumulation by promoting hepatic triglyceride lipolysis, fatty acid oxidation, and VLDL secretion. Furthermore, HNF4α suppressed the progression of NAFL to NASH. Overexpression of human HNF4α inhibited HFCF diet-induced steatohepatitis in control mice but not in hepatocyte-specific p53-/- mice. In HFCF diet-fed mice lacking hepatic Hnf4α, recapitulation of hepatic expression of HNF4α targets cholesterol 7α-hydroxylase and sterol 12α-hydroxylase and normalized hepatic triglyceride levels and attenuated steatohepatitis. CONCLUSIONS: The current study indicates that HNF4α protects against diet-induced development and progression of NAFLD by coordinating the regulation of lipolytic, p53, and bile acid signaling pathways. Targeting hepatic HNF4α may be useful for treatment of NASH.