Progranulin Deficiency Promotes Circuit-Specific Synaptic Pruning by Microglia via Complement Activation.

Microglia maintain homeostasis in the brain, but whether aberrant microglial activation can cause neurodegeneration remains controversial. Here, we use transcriptome profiling to demonstrate that deficiency in frontotemporal dementia (FTD) gene progranulin (Grn) leads to an age-dependent, progressive upregulation of lysosomal and innate immunity genes, increased complement production, and enhanced synaptic pruning in microglia. During aging, Grn(-/-) mice show profound microglia infiltration and preferential elimination of inhibitory synapses in the ventral thalamus, which lead to hyperexcitability in the thalamocortical circuits and obsessive-compulsive disorder (OCD)-like grooming behaviors. Remarkably, deleting C1qa gene significantly reduces synaptic pruning by Grn(-/-) microglia and mitigates neurodegeneration, behavioral phenotypes, and premature mortality in Grn(-/-) mice. Together, our results uncover a previously unrecognized role of progranulin in suppressing aberrant microglia activation during aging. These results represent an important conceptual advance that complement activation and microglia-mediated synaptic pruning are major drivers, rather than consequences, of neurodegeneration caused by progranulin deficiency.

Neurotoxic microglia promote TDP-43 proteinopathy in progranulin deficiency.

Aberrant aggregation of the RNA-binding protein TDP-43 in neurons is a hallmark of frontotemporal lobar degeneration caused by haploinsufficiency in the gene encoding progranulin1,2. However, the mechanism leading to TDP-43 proteinopathy remains unclear. Here we use single-nucleus RNA sequencing to show that progranulin deficiency promotes microglial transition from a homeostatic to a disease-specific state that causes endolysosomal dysfunction and neurodegeneration in mice. These defects persist even when Grn-/- microglia are cultured ex vivo. In addition, single-nucleus RNA sequencing reveals selective loss of excitatory neurons at disease end-stage, which is characterized by prominent nuclear and cytoplasmic TDP-43 granules and nuclear pore defects. Remarkably, conditioned media from Grn-/- microglia are sufficient to promote TDP-43 granule formation, nuclear pore defects and cell death in excitatory neurons via the complement activation pathway. Consistent with these results, deletion of the genes encoding C1qa and C3 mitigates microglial toxicity and rescues TDP-43 proteinopathy and neurodegeneration. These results uncover previously unappreciated contributions of chronic microglial toxicity to TDP-43 proteinopathy during neurodegeneration.

Homeostatic, Non-Canonical Role of Macrophage Elastase in Vascular Integrity.

BACKGROUND: Matrix metalloproteinase (MMP)-12 is highly expressed in abdominal aortic aneurysms and its elastolytic function has been implicated in the pathogenesis. This concept is challenged, however, by conflicting data. Here, we sought to revisit the role of MMP-12 in abdominal aortic aneurysm. METHODS: Apoe-/- and Mmp12-/-/Apoe-/- mice were infused with Ang II (angiotensin). Expression of neutrophil extracellular traps (NETs) markers and complement component 3 (C3) levels were evaluated by immunostaining in aortas of surviving animals. Plasma complement components were analyzed by immunoassay. The effects of a complement inhibitor, IgG-FH1-5 (factor H-immunoglobulin G), and macrophage-specific MMP-12 deficiency on adverse aortic remodeling and death from rupture in Ang II-infused mice were determined. RESULTS: Unexpectedly, death from aortic rupture was significantly higher in Mmp12-/-/Apoe-/- mice. This associated with more neutrophils, citrullinated histone H3 and neutrophil elastase, markers of NETs, and C3 levels in Mmp12-/- aortas. These findings were recapitulated in additional models of abdominal aortic aneurysm. MMP-12 deficiency also led to more pronounced elastic laminae degradation and reduced collagen integrity. Higher plasma C5a in Mmp12-/- mice pointed to complement overactivation. Treatment with IgG-FH1-5 decreased aortic wall NETosis and reduced adverse aortic remodeling and death from rupture in Ang II-infused Mmp12-/- mice. Finally, macrophage-specific MMP-12 deficiency recapitulated the effects of global MMP-12 deficiency on complement deposition and NETosis, as well as adverse aortic remodeling and death from rupture in Ang II-infused mice. CONCLUSIONS: An MMP-12 deficiency/complement activation/NETosis pathway compromises aortic integrity, which predisposes to adverse vascular remodeling and abdominal aortic aneurysm rupture. Considering these new findings, the role of macrophage MMP-12 in vascular homeostasis demands re-evaluation of MMP-12 function in diverse settings.

Twist-Angle and Thickness-Ratio Tuning of Plasmon Polaritons in Twisted Bilayer van der Waals Films.

Stacking bilayer structures is an efficient way to tune the topology of polaritons in in-plane anisotropic films, e.g., by leveraging the twist angle (TA). However, the effect of another geometric parameter, the film thickness ratio (TR), on manipulating the plasmon topology in bilayers is elusive. Here, we fabricate bilayer structures of WTe2 films, which naturally host in-plane hyperbolic plasmons in the terahertz range. Plasmon topology is successfully modified by changing the TR and TA synergistically, manifested by the extinction spectra of unpatterned films and the polarization dependence of the plasmon intensity measured in skew ribbon arrays. Such TR- and TA-tunable topological transitions can be well explained based on the effective sheet optical conductivity by adding up those of the two films. Our study demonstrates TR as another degree of freedom for the manipulation of plasmonic topology in nanophotonics, exhibiting promising applications in biosensing, heat transfer, and the enhancement of spontaneous emission.

Improved Interfacial Interactions of Dip Coatings by In Situ Introducing Silica to Enhance Corrosion Resistance and Metal Bonding Strength.

Corrosion is an irreversible phenomenon in nature that has been a major source of metal degradation. We herein provide a unique approach for embedding nanoparticles into epoxy resins via hydrogen bonding adsorption of in situ hydrophilic silica. Based on this adsorption action, a super-anticorrosive epoxy-based Teflon (MEP-PTFE) coating for usage on metals such as aluminum alloys was developed utilizing one-step dip coating, with promising engineering and public applications. It should be noted that the binding strength between the resultant MEP-PTFE coating and the substrate was 13.5 N. This coating had an impedance modulus of over 8 × 109 Ω·cm2 at 0.01 Hz and an impressive corrosion inhibition efficiency of 99.999%. The anticorrosion barrier from the diffusion control to the charge transfer control was revealed for the future good design of resin matrix coatings with excellent corrosion resistance.

FGF-dependent metabolic control of vascular development.

Blood and lymphatic vasculatures are intimately involved in tissue oxygenation and fluid homeostasis maintenance. Assembly of these vascular networks involves sprouting, migration and proliferation of endothelial cells. Recent studies have suggested that changes in cellular metabolism are important to these processes. Although much is known about vascular endothelial growth factor (VEGF)-dependent regulation of vascular development and metabolism, little is understood about the role of fibroblast growth factors (FGFs) in this context. Here we identify FGF receptor (FGFR) signalling as a critical regulator of vascular development. This is achieved by FGF-dependent control of c-MYC (MYC) expression that, in turn, regulates expression of the glycolytic enzyme hexokinase 2 (HK2). A decrease in HK2 levels in the absence of FGF signalling inputs results in decreased glycolysis, leading to impaired endothelial cell proliferation and migration. Pan-endothelial- and lymphatic-specific Hk2 knockouts phenocopy blood and/or lymphatic vascular defects seen in Fgfr1/Fgfr3 double mutant mice, while HK2 overexpression partly rescues the defects caused by suppression of FGF signalling. Thus, FGF-dependent regulation of endothelial glycolysis is a pivotal process in developmental and adult vascular growth and development.

Comparison of acquisition rate and agreement of axial length with two swept-source optical coherence tomographers and a partial coherence interferometer.

PURPOSE: To assess the acquisition rate and agreement of axial length (AL) measurements with the OA-2000, Anterion, and IOLMaster 500 in cataractous patients. METHODS: In total, 298 eyes of 191 cataractous patients were enrolled and scanned with the three devices in random order. The success rate of AL measurements per device was calculated and a chi-square test was utilized to identify the differences in acquisition rate between the three devices. Logistic regression analysis was applied to evaluate the association of different cataract types and severity with the AL measurement acquisition rate. Bland-Altman plots were mapped to appraise the agreement of AL values. RESULTS: AL measurements were successfully achieved in 288 eyes (96.64%) with the OA-2000, in 282 eyes (94.30%) with the Anteiron, and in 246 eyes (82.55%) with the IOLMaster 500. Significant differences in the acquisition rate were found between either of the SS-OCT devices and IOLMaster 500 by chi-square analysis (P < 0.001). No significant difference was noted between OA-2000 and Anterion. Increasing severity of posterior subcapsular cataract was associated with a higher failure rate with the IOLMaster 500. Bland-Altman analysis identified good agreement between the three biometers with narrow 95% limits of agreement. CONCLUSIONS: The OA-2000 and Anterion showed similarly higher acquisition rate of AL measurements than IOLMaster 500 in cataractous patients. Good agreement for AL values was found between the three biometers in cataractous patients.

Repeatability of a new swept-source optical coherence tomographer and agreement with other three optical biometers.

PURPOSE: To investigate the repeatability of Anterion and compare the agreement of ocular biometric measurements and predicted intraocular lens (IOL) powers with other three optical biometers. METHODS: Flat keratometry (Kf), steep keratometry (Ks), J0 and 45 vectors, central cornea thickness (CCT), anterior chamber depth (ACD), and axial length (AL) from the Anterion, IOLMaster 700, Lenstar LS 900, and OA-2000 were recorded. The IOL powers were calculated with the Hoffer Q, Holladay 1, SRK/T, and Haigis formulas. The repeatability was evaluated using the within-subject standard deviation (Sw), repeatability coefficient (RC), coefficient of variation (COV), and intraclass correlation coefficient (ICC). Inter-device agreement between the four biometers was assessed with the 95% limits of agreement. RESULTS: In total, 101 right eyes of 101 participants were enrolled. The Anterion showed good repeatability for all the included biometric parameters with all the CoV ≤ 0.30% and ICC ≥ 0.930 except for J45 with moderate repeatability (ICC was 0.849). Good agreement was found among the four devices for Kf, Ks, J0, J45, ACD, and AL. Generally, wide 95% LoA was found for the predicted IOL powers with the four IOL calculation formulas between the four devices. CONCLUSIONS: The Anterion showed good repeatability of biometric measurements for most parameters. Good agreement among the four optical biometers was achieved for all the parameters except for CCT and the predicted IOL power. The AL values exhibited the best repeatability with Anterion and the best agreement among the biometers in our study.

Loss of HIPK2 Protects Neurons from Mitochondrial Toxins by Regulating Parkin Protein Turnover.

Mitochondria are important sources of energy, but they are also the target of cellular stress, toxin exposure, and aging-related injury. Persistent accumulation of damaged mitochondria has been implicated in many neurodegenerative diseases. One highly conserved mechanism to clear damaged mitochondria involves the E3 ubiquitin ligase Parkin and PTEN-induced kinase 1 (PINK1), which cooperatively initiate the process called mitophagy that identifies and eliminates damaged mitochondria through the autophagosome and lysosome pathways. Parkin is a mostly cytosolic protein, but is rapidly recruited to damaged mitochondria and target them for mitophagy. Moreover, Parkin interactomes also involve signaling pathways and transcriptional machinery critical for survival and cell death. However, the mechanism that regulates Parkin protein level remains poorly understood. Here, we show that the loss of homeodomain interacting protein kinase 2 (HIPK2) in neurons and mouse embryonic fibroblasts (MEFs) has a broad protective effect from cell death induced by mitochondrial toxins. The mechanism by which Hipk2-/- neurons and MEFs are more resistant to mitochondrial toxins is in part due to the role of HIPK2 and its kinase activity in promoting Parkin degradation via the proteasome-mediated mechanism. The loss of HIPK2 leads to higher cytosolic Parkin protein levels at basal conditions and upon exposure to mitochondrial toxins, which protects mitochondria from toxin-induced damage. In addition, Hipk2-/- neurons and MEFs show increased expression of PGC-1α (peroxisome proliferator-activated receptor-γ coactivator 1), a Parkin downstream target that can provide additional benefits via transcriptional activation of mitochondrial genes. Together, these results reveal a previously unrecognized avenue to target HIPK2 in neuroprotection via the Parkin-mediated pathway.SIGNIFICANCE STATEMENT In this study, we provide evidence that homeodomain interacting protein kinase 2 (HIPK2) and its kinase activity promote Parkin degradation via the proteasome-mediated pathway. The loss of HIPK2 increases cytosolic and mitochondrial Parkin protein levels under basal conditions and upon exposure to mitochondrial toxins, which protect mitochondria from toxin-induced damage. In addition, Hipk2-/- neurons and mouse embryonic fibroblasts also show increased expression of PGC-1α (peroxisome proliferator-activated receptor-γ coactivator 1), a Parkin downstream target that can provide additional benefits via transcriptional activation of mitochondrial genes. These results indicate that targeting HIPK2 and its kinase activity can have neuroprotective effects by elevating Parkin protein levels.

Layer-Dependent Pressure Effect on the Electronic Structure of 2D Black Phosphorus.

Through infrared spectroscopy, we systematically study the pressure effect on electronic structures of few-layer black phosphorus (BP) with layer number ranging from 2 to 13. We reveal that the pressure-induced shift of optical transitions exhibits strong layer dependence. In sharp contrast to the bulk counterpart which undergoes a semiconductor to semimetal transition under ∼1.8 GPa, the band gap of 2 L increases with increasing pressure until beyond 2 GPa. Meanwhile, for a sample with a given layer number, the pressure-induced shift also differs for transitions with different indices. Through the tight-binding model in conjunction with a Morse potential for the interlayer coupling, this layer- and transition-index-dependent pressure effect can be fully accounted. Our study paves a way for versatile van der Waals engineering of two-dimensional BP.

Murine knockin model for progranulin-deficient frontotemporal dementia with nonsense-mediated mRNA decay.

Frontotemporal dementia (FTD) is the most common neurodegenerative disorder in individuals under age 60 and has no treatment or cure. Because many cases of FTD result from GRN nonsense mutations, an animal model for this type of mutation is highly desirable for understanding pathogenesis and testing therapies. Here, we generated and characterized GrnR493X knockin mice, which model the most common human GRN mutation, a premature stop codon at arginine 493 (R493X). Homozygous GrnR493X mice have markedly reduced Grn mRNA levels, lack detectable progranulin protein, and phenocopy Grn knockout mice, with CNS microgliosis, cytoplasmic TDP-43 accumulation, reduced synaptic density, lipofuscinosis, hyperinflammatory macrophages, excessive grooming behavior, and reduced survival. Inhibition of nonsense-mediated mRNA decay (NMD) by genetic, pharmacological, or antisense oligonucleotide-based approaches showed that NMD contributes to the reduced mRNA levels in GrnR493X mice and cell lines and in fibroblasts from patients containing the GRNR493X mutation. Moreover, the expressed truncated R493X mutant protein was functional in several assays in progranulin-deficient cells. Together, these findings establish a murine model for in vivo testing of NMD inhibition or other therapies as potential approaches for treating progranulin deficiency caused by the R493X mutation.

Probing the Ferromagnetism and Spin Wave Gap in VI3 by Helicity-Resolved Raman Spectroscopy.

Circularly polarized light carries light spin angular momentum, which may lead helicity-resolved Raman scattering to be sensitive to the electronic spin configuration in magnetic materials. Here, we demonstrate that all Raman modes in the 2D ferromagnet VI3 show different scattering intensities to left and right circularly polarized light at low temperatures, which gives direct evidence of the time-reversal symmetry breaking. By measuring the circular polarization of the dominant Raman mode with respect to the temperature and magnetic field, the ferromagnetic (FM) phase transition and hysteresis behavior can be clearly resolved. Besides the lattice excitations, quasielastic scattering is detected in the paramagnetic phase, and it gradually evolves into the acoustic magnon mode at 18.5 cm-1 in the FM state, which gives the spin wave gap that results from large magnetic anisotropy. Our findings demonstrate that helicity-resolved Raman spectroscopy is an effective tool to directly probe the ferromagnetism in 2D magnets.

From Anomalous to Normal: Temperature Dependence of the Band Gap in Two-Dimensional Black Phosphorus.

The temperature dependence of the band gap is crucial to a semiconductor. Bulk black phosphorus is known to exhibit an anomalous behavior. Through optical spectroscopy, here we show that the temperature effect on black phosphorus band gap gradually evolves with decreasing layer number, eventually turns into a normal one in the monolayer limit, rendering a crossover from the anomalous to the normal. Meanwhile, the temperature-induced shift in optical resonance also differs with different transition indices for the same thickness sample. A comprehensive analysis reveals that the temperature-tunable interlayer coupling is responsible for the observed diverse scenario. Our study provides a key to the apprehension of the anomalous temperature behavior in certain layered semiconductors.

TCF7L2 (Transcription Factor 7-Like 2) Regulation of GATA6 (GATA-Binding Protein 6)-Dependent and -Independent Vascular Smooth Muscle Cell Plasticity and Intimal Hyperplasia.

Objective- TCF7L2 (transcription factor 7-like 2) is a Wnt-regulated transcription factor that maintains stemness and promotes proliferation in embryonic tissues and adult stem cells. Mice with a coronary artery disease-linked mutation in Wnt-coreceptor LRP6 (LDL receptor-related protein 6) exhibit vascular smooth muscle cell dedifferentiation and obstructive coronary artery disease, which are paradoxically associated with reduced TCF7L2 expression. We conducted a comprehensive study to explore the role of TCF7L2 in vascular smooth muscle cell differentiation and protection against intimal hyperplasia. Approach and Results- Using multiple mouse models, we demonstrate here that TCF7L2 promotes differentiation and inhibits proliferation of vascular smooth muscle cells. TCF7L2 accomplishes these effects by stabilization of GATA6 (GATA-binding protein 6) and upregulation of SM-MHC (smooth muscle cell myosin heavy chain) and cell cycle inhibitors. Accordingly, TCF7L2 haploinsufficient mice exhibited increased susceptibility to injury-induced hyperplasia, while mice overexpressing TCF7L2 were protected against injury-induced intimal hyperplasia compared with wild-type littermates. Consequently, the overexpression of TCF7L2 in LRP6 mutant mice rescued the injury-induced intimal hyperplasia. Conclusions- Our novel findings imply cell type-specific functional role of TCF7L2 and provide critical insight into mechanisms underlying the pathogenesis of intimal hyperplasia.

HIPK2-Mediated Transcriptional Control of NMDA Receptor Subunit Expression Regulates Neuronal Survival and Cell Death.

NMDA receptors are critical for neuronal communication. Dysfunction in NMDA receptors has been implicated in neuropsychiatric diseases. While it is well recognized that the composition of NMDA receptors undergoes a GluN2B-to-GluN2A switch in early postnatal life, the mechanism regulating this switch remains unclear. Using transcriptomic and functional analyses in brain tissues from male and female Hipk2+/+ and Hipk2-/- mice, we showed that the HIPK2-JNK-c-Jun pathway is important in suppressing the transcription of Grin2a and Grin2c, which encodes the GluN2A and GluN2C subunits of the NMDA receptors, respectively. Loss of HIPK2 leads to a significant decrease in JNK-c-Jun signaling, which in turn derepresses the transcription of Grin2a and Grin2c mRNA and upregulates GluN2A and GluN2C protein levels. These changes result in a significant increase of GluN2A/GluN2B ratio in synapse and mitochondria, a persistent activation of the ERK-CREB pathway and the upregulation of synaptic activity-regulated genes, which collectively contribute to the resistance of Hipk2-/- neurons to cell death induced by mitochondrial toxins.SIGNIFICANCE STATEMENT We identify HIPK2-JNK-c-Jun signaling as a key mechanism that regulates the transcription of NMDA receptor subunits GluN2A and GluN2C in vivo Our results provide insights into a previously unrecognized molecular mechanism that control the switch of NMDA receptor subunits in early postnatal brain development. Furthermore, we provide evidence that changes in the ratio of NMDA subunits GluN2A/GluN2B can also be detected in the synapse and mitochondria, which contributes to a persistent activation of the prosurvival ERK-CREB pathway and its downstream target genes. Collectively, these changes protect HIPK2 deficient neurons from mitochondrial toxins.

Syndecan 4 controls lymphatic vasculature remodeling during mouse embryonic development.

The role of fluid shear stress in vasculature development and remodeling is well appreciated. However, the mechanisms regulating these effects remain elusive. We show that abnormal flow sensing in lymphatic endothelial cells (LECs) caused by Sdc4 or Pecam1 deletion in mice results in impaired lymphatic vessel remodeling, including abnormal valve morphogenesis. Ablation of either gene leads to the formation of irregular, enlarged and excessively branched lymphatic vessels. In both cases, lymphatic valve-forming endothelial cells are randomly oriented, resulting in the formation of abnormal valves. These abnormalities are much more pronounced in Sdc4-/-; Pecam1-/- double-knockout mice, which develop severe edema. In vitro, SDC4 knockdown human LECs fail to align under flow and exhibit high expression of the planar cell polarity protein VANGL2. Reducing VANGL2 levels in SDC4 knockdown LECs restores their alignment under flow, while VANGL2 overexpression in wild-type LECs mimics the flow alignment abnormalities seen in SDC4 knockdown LECs. SDC4 thus controls flow-induced LEC polarization via regulation of VANGL2 expression.

Endothelial Cell Autonomous Role of Akt1: Regulation of Vascular Tone and Ischemia-Induced Arteriogenesis.

OBJECTIVE: The importance of PI3K/Akt signaling in the vasculature has been demonstrated in several models, as global loss of Akt1 results in impaired postnatal ischemia- and VEGF-induced angiogenesis. The ubiquitous expression of Akt1, however, raises the possibility of cell-type-dependent Akt1-driven actions, thereby necessitating tissue-specific characterization. APPROACH AND RESULTS: Herein, we used an inducible, endothelial-specific Akt1-deleted adult mouse model (Akt1iECKO) to characterize the endothelial cell autonomous functions of Akt1 in the vascular system. Endothelial-targeted ablation of Akt1 reduces eNOS (endothelial nitric oxide synthase) phosphorylation and promotes both increased vascular contractility in isolated vessels and elevated diastolic blood pressures throughout the diurnal cycle in vivo. Furthermore, Akt1iECKO mice subject to the hindlimb ischemia model display impaired blood flow and decreased arteriogenesis. CONCLUSIONS: Endothelial Akt1 signaling is necessary for ischemic resolution post-injury and likely reflects the consequence of NO insufficiency critical for vascular repair.

SLC52A3 expression is activated by NF-κB p65/Rel-B and serves as a prognostic biomarker in esophageal cancer.

The human riboflavin transporter-3 (encoded by SLC52A3) plays a prominent role in riboflavin absorption. Interestingly, abnormal expression patterns of SLC52A3 in multiple types of human cancers have been recently noted. However, the molecular mechanisms underlying its dysregulation remain unclear. In this study, we find that SLC52A3 has two transcript variants that differ in the transcriptional start site, and encode different proteins: SLC52A3a and SLC52A3b. Importantly, aberrant expressions of SLC52A3 are associated with stepwise development of esophageal squamous cell carcinoma (ESCC) as well as the survival rates of ESCC patients. Functionally, SLC52A3a, but not SLC52A3b, strongly promotes the proliferation and colony formation of ESCC cells. Furthermore, SLC52A3 5'-flanking regions contain NF-κB p65/Rel-B-binding sites, which are crucial for mediating SLC52A3 transcriptional activity in ESCC cells. Chromatin immunoprecipitation and electrophoretic mobility shift assay reveal that p65/Rel-B bind to 5'-flanking regions of SLC52A3. Accordingly, NF-κB signaling upregulates SLC52A3 transcription upon TNFα stimulation. Taken together, these results elucidate the mechanisms underlying SLC52A3 overexpression in ESCC. More importantly, our findings identify SLC52A3 as both a predictive and prognostic biomarker for this deadly cancer.

Targeting NCK-Mediated Endothelial Cell Front-Rear Polarity Inhibits Neovascularization.

BACKGROUND: Sprouting angiogenesis is a key process driving blood vessel growth in ischemic tissues and an important drug target in a number of diseases, including wet macular degeneration and wound healing. Endothelial cells forming the sprout must develop front-rear polarity to allow sprout extension. The adaptor proteins Nck1 and 2 are known regulators of cytoskeletal dynamics and polarity, but their function in angiogenesis is poorly understood. Here, we show that the Nck adaptors are required for endothelial cell front-rear polarity and migration downstream of the angiogenic growth factors VEGF-A and Slit2. METHODS AND RESULTS: Mice carrying inducible, endothelial-specific Nck1/2 deletions fail to develop front-rear polarized vessel sprouts and exhibit severe angiogenesis defects in the postnatal retina and during embryonic development. Inactivation of NCK1 and 2 inhibits polarity by preventing Cdc42 and Pak2 activation by VEGF-A and Slit2. Mechanistically, NCK binding to ROBO1 is required for both Slit2- and VEGF-induced front-rear polarity. Selective inhibition of polarized endothelial cell migration by targeting Nck1/2 prevents hypersprouting induced by Notch or Bmp signaling inhibition, and pathological ocular neovascularization and wound healing, as well. CONCLUSIONS: These data reveal a novel signal integration mechanism involving NCK1/2, ROBO1/2, and VEGFR2 that controls endothelial cell front-rear polarity during sprouting angiogenesis.

Interferon-γ-mediated allograft rejection exacerbates cardiovascular disease of hyperlipidemic murine transplant recipients.

RATIONALE: Transplantation, the most effective therapy for end-stage organ failure, is markedly limited by early-onset cardiovascular disease (CVD) and premature death of the host. The mechanistic basis of this increased CVD is not fully explained by known risk factors. OBJECTIVE: To investigate the role of alloimmune responses in promoting CVD of organ transplant recipients. METHODS AND RESULTS: We established an animal model of graft-exacerbated host CVD by combining murine models of atherosclerosis (apolipoprotein E-deficient recipients on standard diet) and of intra-abdominal graft rejection (heterotopic cardiac transplantation without immunosuppression). CVD was absent in normolipidemic hosts receiving allogeneic grafts and varied in severity among hyperlipidemic grafted hosts according to recipient-donor genetic disparities, most strikingly across an isolated major histocompatibility complex class II antigen barrier. Host disease manifested as increased atherosclerosis of the aorta that also involved the native coronary arteries and new findings of decreased cardiac contractility, ventricular dilatation, and diminished aortic compliance. Exacerbated CVD was accompanied by greater levels of circulating cytokines, especially interferon-γ and other Th1-type cytokines, and showed both systemic and intralesional activation of leukocytes, particularly T-helper cells. Serological neutralization of interferon-γ after allotransplantation prevented graft-related atherosclerosis, cardiomyopathy, and aortic stiffening in the host. CONCLUSIONS: Our study reveals that sustained activation of the immune system because of chronic allorecognition exacerbates the atherogenic diathesis of hyperlipidemia and results in de novo cardiovascular dysfunction in organ transplant recipients.