Vascular cell adhesion molecule 1 expression by biliary epithelium promotes persistence of inflammation by inhibiting effector T-cell apoptosis

Chronic hepatitis occurs when effector lymphocytes are recruited to the liver from blood and retained in tissue to interact with target cells, such as hepatocytes or bile ducts (BDs). Vascular cell adhesion molecule 1 (VCAM-1; CD106), a member of the immunoglobulin superfamily, supports leukocyte adhesion by binding a4b1 integrins and is critical for the recruitment of monocytes and lymphocytes during inflammation. We detected VCAM-1 on cholangiocytes in chronic liver disease (CLD) and hypothesized that biliary expression of VCAM-1 contributes to the persistence of liver inflammation. Hence, in this study, we examined whether cholangiocyte expression of VCAM-1 promotes the survival of intrahepatic a4b1 expressing effector T cells. We examined interactions between primary human cholangiocytes and isolated intrahepatic T cells ex vivo and in vivo using the Ova-bil antigen-driven murine model of biliary inflammation. VCAM-1 was detected on BDs in CLDs (primary biliary cirrhosis, primary sclerosing cholangitis, alcoholic liver disease, and chronic hepatitis C), and human cholangiocytes expressed VCAM-1 in response to tumor necrosis factor alpha alone or in combination with CD40L or interleukin-17. Liver-derived T cells adhered to cholangiocytes in vitro by a4b1, which resulted in signaling through nuclear factor kappa B p65, protein kinase B1, and p38 mitogen-activated protein kinase phosphorylation. This led to increased mitochondrial B-cell lymphoma 2 accumulation and decreased activation of caspase 3, causing increased cell survival. We confirmed our findings in a murine model of hepatobiliary inflammation where inhibition of VCAM-1 decreased liver inflammation by reducing lymphocyte recruitment and increasing CD8 and T helper 17 CD4 Tcell survival. Conclusions: VCAM-1 expression by cholangiocytes contributes to persistent inflammation by conferring a survival signal to a4b1 expressing proinflammatory T lymphocytes in CLD.

Microstructural Parameters of Bone Evaluated Using HR-pQCT Correlate with the DXA-Derived Cortical Index and the Trabecular Bone Score in a Cohort of Randomly Selected Premenopausal Women.

BACKGROUND Areal bone mineral density is predictive for fracture risk. Microstructural bone parameters evaluated at the appendicular skeleton by high-resolution peripheral quantitative computed tomography (HR-pQCT) display differences between healthy patients and fracture patients. With the simple geometry of the cortex at the distal tibial diaphysis, a cortical index of the tibia combining material and mechanical properties correlated highly with bone strength ex vivo. The trabecular bone score derived from the scan of the lumbar spine by dual-energy X-ray absorptiometry (DXA) correlated ex vivo with the micro architectural parameters. It is unknown if these microstructural correlations could be made in healthy premenopausal women. METHODS Randomly selected women between 20-40 years of age were examined by DXA and HR-pQCT at the standard regions of interest and at customized sub regions to focus on cortical and trabecular parameters of strength separately. For cortical strength, at the distal tibia the volumetric cortical index was calculated directly from HR-pQCT and the areal cortical index was derived from the DXA scan using a Canny threshold-based tool. For trabecular strength, the trabecular bone score was calculated based on the DXA scan of the lumbar spine and was compared with the corresponding parameters derived from the HR-pQCT measurements at radius and tibia. RESULTS Seventy-two healthy women were included (average age 33.8 years, average BMI 23.2 kg/m(2)). The areal cortical index correlated highly with the volumetric cortical index at the distal tibia (R  =  0.798). The trabecular bone score correlated moderately with the microstructural parameters of the trabecular bone. CONCLUSION This study in randomly selected premenopausal women demonstrated that microstructural parameters of the bone evaluated by HR-pQCT correlated with the DXA derived parameters of skeletal regions containing predominantly cortical or cancellous bone. Whether these indexes are suitable for better predictions of the fracture risk deserves further investigation.

Glutamate and nitric oxide as regulatory transmitters in developing rabbit retina

Glutamate is the major excitatory neurotransmitter in the retina and serves as the synaptic messenger for the three classes of neurons which constitute the vertical pathway--the photoreceptors, bipolar cells and ganglion cells. In addition, the glutamate system has been localized morphologically, pharmacologically as well as molecularly during the first postnatal week of development before synaptogenesis occurs. The role which glutamate plays in the maturing visual system is complex but ranges from mediating developmental neurotoxicity to inducing neurite outgrowth.^ Nitric oxide/cGMP is a novel intercellular messenger which is thought to act in concert with the glutamate system in regulating a variety of cellular processes in the brain as well as retina, most notably neurotoxicity. Several developmental activities including programmed cell death, synapse elimination and synaptic reorganization are possible functions of cellular regulation modulated by nitric oxide as well as glutamate.^ The purpose of this thesis is to (1) biochemically characterize the endogenous pools of glutamate and determine what fraction exists extracellularly; (2) examine the morphological expression of NO producing cells in developing retina; (3) test the functional coupling of the NMDA subtype of glutamate receptor to the NO system by examining neurotoxicity which has roles in both the maturing and adult retina.^ Biochemical sampling of perfusates collected from the photoreceptor surface of ex vivo retina demonstrated that although the total pool of glutamate present at birth is relatively modest, a high percentage resides in extracellular pools. As a result, immature neurons without significant synaptic connections survive and develop in a highly glutamatergic environment which has been shown to be toxic in the adult retina.^ The interaction of the glutamate system with the NO system has been postulated to regulate neuronal survival. We therefore examined the developmental expression of the enzyme responsible for producing NO, nitric oxide synthase (NOS), using an antibody to the constitutive form of NOS found in the brain. The neurons thought to produce the majority of NO in the adult retina, a subpopulation of widefield amacrine cells, were not immunoreactive until the end of the second postnatal week. However, a unique developmental expression was observed in the ganglion cell layer and developing outer nuclear layer of the retina during the first postnatal week. We postulate NO producing neurons may not be present in a mature configuration therefore permitting neuronal survival in a highly glutamatergic microenvironment and allowing NO to play a development-specific role at this time.^ The next set of experiments constituted a functional test of the hypothesis that the absence of the prototypic NO producing cells in developing retina protects immature neurons against glutamate toxicity. An explant culture system developed in order to examine cellular responses of immature and adult neurons to glutamate toxicity showed that immature neurons were affected by NMDA but were less responsive to NMDA and NO mediated toxicity. In contrast, adult explants exhibited significant NMDA toxicity which was attenuated by NMDA antagonists, 2-amino-5-phosphonovaleric acid (APV), dextromethorphan (Dex) and N$\rm\sp{G}$-D-methyl arginine (metARG). These results indicated that pan-retinal neurotoxicity via the NMDA receptor and/or NO activation occurred in the adult retina but was not significant in the neonate. (Abstract shortened by UMI.) ^

Rescue from apoptosis in early (CD34 selected) versus late (CD34 unselected) human hematopoietic cells by VLA-4 and VCAM-1 dependent adhesion to bone marrow stromal cells

The interaction of hematopoietic precursor cell with bone marrow stromal cells is assumed to be important to the survival of hematopoietic precursor cells during hematopoietic cell long-term culture. Early hematopoietic stem cells are preferentially found within the stromal adherent cell fraction in primary long-term bone marrow cultures. The purpose of this dissertation was to understand the molecular mechanisms that govern these interactions for the regulation of survival and proliferation of early versus late hematopoietic cells.^ Monoclonal antibodies to the VLA-4 recognize the alpha4 beta1 integrin receptor on human hematopoietic cells. This monoclonal antibody blocks the adhesion between early hematopoietic progenitor cells (CD34 selected cells) and stromal cells when added to cultures of these cells. Addition of the VLA-4 monoclonal antibody to cultures of stromal cells and CD34 selected cells was shown to induce apoptosis of CD34 selected cells in these CD34 selected cell/stromal cell cocultures, as measured by the terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick end-labeling method. In contrast to these experiments with early hematopoietic progenitor cells (CD34+), the level of adhesion between more differentiated cells (unfractionated hematopoietic cells) and stromal cells was not significantly altered by addition of the anti-VLA-4 monoclonal antibody. Similarly, the level of apoptosis of unfractionated hematopoietic cells was not significantly increased by the addition of anti-VLA-4 monoclonal antibody to cultures of the latter cells with stromal cells. The binding of the unfractionated cells is less than that of the CD34 selected. Since there is no difference between the alpha4 beta1 integrin expression level of the early and late myeloid cells, there may be a difference in the functional state of the integrin between the early and late myeloid cells. We also show that CD34+ selected precursor cells proliferate at a higher rate when these cells are plated on recombinant VCAM-1 molecules. These data indicate that the alpha4beta1 integrin receptor (VLA-4) plays a central role in the apoptosis rescue function which results from the anchorage-dependent growth of the CD34 selected early hematopoietic cells on stromal cells. The data suggest that these apoptosis rescue pathways have less significance as the cells mature and become anchorage-independent in their growth. These data should assist in the design of systems for the ex vivo proliferation and transduction of early hematopoietic cells for genetic therapy. ^

Targeting of {\it HER\/}-{\it 2\/}/{\it neu\/} with E1A gene therapy

Amplification or overexpression of HER-2/neu has been demonstrated in human cancers of the ovary, breast, lung and correlated with chemoresistance and poor clinic prognosis. We have previously found that the adenovirus type 5 early region 1A (E1A) gene product can repress the overexpression and suppress the tumorigenic potential of HER-2/neu-overexpressing cancer cells. In addition, E1A has been reported to induce apoptosis and inhibit the metastatic potential of tumor cells. Therefore, E1A could be considered as a tumor suppressor gene in HER-2/neu-overexpressing cancer cells. To develop an efficient HER-2/neu-targeting gene therapy with E1A, adenoviral vector or cationic liposome was used to introduce E1A into human ovarian, breast and lung cancer cells. Successful therapeutic effects were achieved.^ A replication-deficient adenovirus containing the E1A gene, Ad.E1A(+), was used to infect HER-2/neu-overexpressing human ovarian cancer cell line. Ovarian cancer growth in vitro and colony formation in soft agarose were greatly inhibited.^ To examine tumor suppressor function of E1A in breast cancer, we introduced E1A in vitro by adenovirus into both HER-2/neu-overexpressing and low-expressing human breast cancer cell lines. In HER-2/neu-overexpressing cells, E1A greatly inhibited tumor cell growth in vitro and colony formation in soft agarose. However, in low HER-2/neu expressing cancer cell lines, E1A could only reduce colony formation in soft agarose but had no significant effect on cell growth in monolayer, indicating different effects of E1A in these two types of cancer cells. To test the local therapeutic efficacy of E1A, we used either adenovirus- or liposome-mediated E1A gene delivery systems in an orthotopic breast cancer animal model.^ To test the therapeutic efficacy of systemically-delivered E1A in vivo lung cancer, we treated mice bearing intratracheal lung cancer by i.v. tail injections of Ad.E1A(+). As a result, Ad.E1A(+) suppressed HER-2/neu overexpression and inhibited intratracheal lung cancer growth. However, no significant tumor suppression effect of Ad.E1A(+) was observed in mice bearing HER-2/neu low expressing cell line when the same therapeutic procedure was followed. (Abstract shortened by UMI.) ^

64Cu- and 68Ga-labelled [Nle(14),Lys(40)(Ahx-NODAGA)NH2]-exendin-4 for pancreatic beta cell imaging in rats.

PURPOSE Glucagon-like peptide-1 receptor (GLP-1R) is a molecular target for imaging of pancreatic beta cells. We compared the ability of [Nle(14),Lys(40)(Ahx-NODAGA-(64)Cu)NH2]-exendin-4 ([(64)Cu]NODAGA-exendin-4) and [Nle(14),Lys(40)(Ahx-NODAGA-(68)Ga)NH2]-exendin-4 ([(68)Ga]NODAGA-exendin-4) to detect native pancreatic islets in rodents. PROCEDURES The stability, lipophilicity and affinity of the radiotracers to the GLP-1R were determined in vitro. The biodistribution of the tracers was assessed using autoradiography, ex vivo biodistribution and PET imaging. Estimates for human radiation dosimetry were calculated. RESULTS We found GLP-1R-specific labelling of pancreatic islets. However, the pancreas could not be visualised in PET images. The highest uptake of the tracers was observed in the kidneys. Effective dose estimates for [(64)Cu]NODAGA-exendin-4 and [(68)Ga]NODAGA-exendin-4 were 0.144 and 0.012 mSv/MBq, respectively. CONCLUSION [(64)Cu]NODAGA-exendin-4 might be more effective for labelling islets than [(68)Ga]NODAGA-exendin-4. This is probably due to the lower specific radioactivity of [(68)Ga]NODAGA-exendin-4 compared to [(64)Cu]NODAGA-exendin-4. The radiation dose in the kidneys may limit the use of [(64)Cu]NODAGA-exendin-4 as a clinical tracer.

Application of real-time fluorescent PCR for quantitative assessment of Neospora caninum infections in organotypic slice cultures of rat central nervous system tissue

The previously described Nc5-specific PCR test for the diagnosis of Neospora caninum infections was used to develop a quantitative PCR assay which allows the determination of infection intensities within different experimental and diagnostic sample groups. The quantitative PCR was performed by using a dual fluorescent hybridization probe system and the LightCycler Instrument for online detection of amplified DNA. This assay was successfully applied for demonstrating the parasite proliferation kinetics in organotypic slice cultures of rat brain which were infected in vitro with N. caninum tachyzoites. This PCR-based method of parasite quantitation with organotypic brain tissue samples can be regarded as a novel ex vivo approach for exploring different aspects of cerebral N. caninum infection.

CD14-Dependent Monocyte Isolation Enhances Phagocytosis of Listeria monocytogenes by Proinflammatory, GM-CSF-Derived Macrophages

Macrophages are an important line of defence against invading pathogens. Human macrophages derived by different methods were tested for their suitability as models to investigate Listeria monocytogenes (Lm) infection and compared to macrophage-like THP-1 cells. Human primary monocytes were isolated by either positive or negative immunomagnetic selection and differentiated in the presence of granulocyte macrophage colony-stimulating factor (GM-CSF) or macrophage colony-stimulating factor (M-CSF) into pro- or anti-inflammatory macrophages, respectively. Regardless of the isolation method, GM-CSF-derived macrophages (GM-Mφ) stained positive for CD206 and M-CSF-derived macrophages (M-Mφ) for CD163. THP-1 cells did not express CD206 or CD163 following incubation with PMA, M- or GM-CSF alone or in combination. Upon infection with Lm, all primary macrophages showed good survival at high multiplicities of infection whereas viability of THP-1 was severely reduced even at lower bacterial numbers. M-Mφ generally showed high phagocytosis of Lm. Strikingly, phagocytosis of Lm by GM-Mφ was markedly influenced by the method used for isolation of monocytes. GM-Mφ derived from negatively isolated monocytes showed low phagocytosis of Lm whereas GM-Mφ generated from positively selected monocytes displayed high phagocytosis of Lm. Moreover, incubation with CD14 antibody was sufficient to enhance phagocytosis of Lm by GM-Mφ generated from negatively isolated monocytes. By contrast, non-specific phagocytosis of latex beads by GM-Mφ was not influenced by treatment with CD14 antibody. Furthermore, phagocytosis of Lactococcus lactis, Escherichia coli, human cytomegalovirus and the protozoan parasite Leishmania major by GM-Mφ was not enhanced upon treatment with CD14 antibody indicating that this effect is specific for Lm. Based on these observations, we propose macrophages derived by ex vivo differentiation of negatively selected human primary monocytes as the most suitable model to study Lm infection of macrophages.

The ovine cerebral venous system: comparative anatomy, visualization, and implications for translational research.

Cerebrovascular diseases are significant causes of death and disability in humans. Improvements in diagnostic and therapeutic approaches strongly rely on adequate gyrencephalic, large animal models being demanded for translational research. Ovine stroke models may represent a promising approach but are currently limited by insufficient knowledge regarding the venous system of the cerebral angioarchitecture. The present study was intended to provide a comprehensive anatomical analysis of the intracranial venous system in sheep as a reliable basis for the interpretation of experimental results in such ovine models. We used corrosion casts as well as contrast-enhanced magnetic resonance venography to scrutinize blood drainage from the brain. This combined approach yielded detailed and, to some extent, novel findings. In particular, we provide evidence for chordae Willisii and lateral venous lacunae, and report on connections between the dorsal and ventral sinuses in this species. For the first time, we also describe venous confluences in the deep cerebral venous system and an 'anterior condylar confluent' as seen in humans. This report provides a detailed reference for the interpretation of venous diagnostic imaging findings in sheep, including an assessment of structure detectability by in vivo (imaging) versus ex vivo (corrosion cast) visualization methods. Moreover, it features a comprehensive interspecies-comparison of the venous cerebral angioarchitecture in man, rodents, canines and sheep as a relevant large animal model species, and describes possible implications for translational cerebrovascular research.

Minimally invasive photopolymerization in intervertebral disc tissue cavities

Photopolymerized hydrogels are commonly used for a broad range of biomedical applications. As long as the polymer volume is accessible, gels can easily be hardened using light illumination. However, in clinics, especially for minimally invasive surgery, it becomes highly challenging to control photopolymerization. The ratios between polymerization- volume and radiating-surface-area are several orders of magnitude higher than for ex-vivo settings. Also tissue scattering occurs and influences the reaction. We developed a Monte Carlo model for photopolymerization, which takes into account the solid/liquid phase changes, moving solid/liquid-boundaries and refraction on these boundaries as well as tissue scattering in arbitrarily designable tissue cavities. The model provides a tool to tailor both the light probe and the scattering/absorption properties of the photopolymer for applications such as medical implants or tissue replacements. Based on the simulations, we have previously shown that by adding scattering additives to the liquid monomer, the photopolymerized volume was considerably increased. In this study, we have used bovine intervertebral disc cavities, as a model for spinal degeneration, to study photopolymerization in-vitro. The cavity is created by enzyme digestion. Using a custom designed probe, hydrogels were injected and photopolymerized. Magnetic resonance imaging (MRI) and visual inspection tools were employed to investigate the successful photopolymerization outcomes. The results provide insights for the development of novel endoscopic light-scattering polymerization probes paving the way for a new generation of implantable hydrogels.

Retinal Laser Lesion Visibility in Simultaneous Ultra-High Axial Resolution Optical Coherence Tomography

Ex vivo porcine retina laser lesions applied with varying laser power (20 mW–2 W, 10 ms pulse, 196 lesions) are manually evaluated by microscopic and optical coherence tomography (OCT) visibility, as well as in histological sections immediately after the deposition of the laser energy. An optical coherence tomography system with 1.78 um axial resolution specifically developed to image thin retinal layers simultaneously to laser therapy is presented, and visibility thresholds of the laser lesions in OCT data and fundus imaging are compared. Optical coherence tomography scans are compared with histological sections to estimate the resolving power for small optical changes in the retinal layers, and real-time time-lapse scans during laser application are shown and analyzed quantitatively. Ultrahigh-resolution OCT inspection features a lesion visibility threshold 40–50 mW (17 reduction) lower than for visual inspection. With the new measurement system, 42 of the lesions that were invisible using state-of-the-art ophthalmoscopic methods could be detected.

Helicobacter pylori Inhibits Dendritic Cell Maturation via Interleukin-10-Mediated Activation of the Signal Transducer and Activator of Transcription 3 Pathway.

Helicobacter pylori infects the human gastric mucosa causing a chronic infection that is the primary risk factor for gastric cancer development. Recent studies demonstrate that H. pylori promotes tolerogenic dendritic cell (DC) development indicating that this bacterium evades the host immune response. However, the signaling pathways involved in modulating DC activation during infection remain unclear. Here, we report that H. pylori infection activated the signal transducer and activator of transcription 3 (STAT3) pathway in murine bone marrow-derived DCs (BMDCs) and splenic DCs isolated ex vivo. Isogenic cagA-, cagE-, vacA- and urease-mutants exhibited levels of phosphoSTAT3 that were comparable to in the wild-type (WT) parent strain. H. pylori-infected BMDCs produced increased immunosuppressive IL-10, which activated STAT3 in an autocrine/paracrine fashion. Neutralization of IL-10 prevented H. pylori-mediated STAT3 activation in both BMDCs and splenic DCs. In addition, anti-IL-10 treatment of infected H. pylori-BMDCs was associated with increased CD86 and MHC II expression and enhanced proinflammatory IL-1β cytokine secretion. Finally, increased CD86 and MHC II expression was detected in H. pylori-infected STAT3 knockout DCs when compared to WT controls. Together, these results demonstrate that H. pylori infection induces IL-10 secretion in DCs, which activates STAT3, thereby modulating DC maturation and reducing IL-1β secretion. These findings identify a host molecular mechanism by which H. pylori can manipulate the innate immune response to potentially favor chronic infection and promote carcinogenesis. © 2014 S. Karger AG, Basel.

Leukemic stem cell frequency: a strong biomarker for clinical outcome in acute myeloid leukemia.

INTRODUCTION Treatment failure in acute myeloid leukemia is probably caused by the presence of leukemia initiating cells, also referred to as leukemic stem cells, at diagnosis and their persistence after therapy. Specific identification of leukemia stem cells and their discrimination from normal hematopoietic stem cells would greatly contribute to risk stratification and could predict possible relapses. RESULTS For identification of leukemic stem cells, we developed flow cytometric methods using leukemic stem cell associated markers and newly-defined (light scatter) aberrancies. The nature of the putative leukemic stem cells and normal hematopoietic stem cells, present in the same patient's bone marrow, was demonstrated in eight patients by the presence or absence of molecular aberrancies and/or leukemic engraftment in NOD-SCID IL-2Rγ-/- mice. At diagnosis (n=88), the frequency of the thus defined neoplastic part of CD34+CD38- putative stem cell compartment had a strong prognostic impact, while the neoplastic parts of the CD34+CD38+ and CD34- putative stem cell compartments had no prognostic impact at all. After different courses of therapy, higher percentages of neoplastic CD34+CD38- cells in complete remission strongly correlated with shorter patient survival (n=91). Moreover, combining neoplastic CD34+CD38- frequencies with frequencies of minimal residual disease cells (n=91), which reflect the total neoplastic burden, revealed four patient groups with different survival. CONCLUSION AND PERSPECTIVE Discrimination between putative leukemia stem cells and normal hematopoietic stem cells in this large-scale study allowed to demonstrate the clinical importance of putative CD34+CD38- leukemia stem cells in AML. Moreover, it offers new opportunities for the development of therapies directed against leukemia stem cells, that would spare normal hematopoietic stem cells, and, moreover, enables in vivo and ex vivo screening for potential efficacy and toxicity of new therapies.

Successful pacing using a batteryless sunlight-powered pacemaker.

AIMS Today's cardiac pacemakers are powered by batteries with limited energy capacity. As the battery's lifetime ends, the pacemaker needs to be replaced. This surgical re-intervention is costly and bears the risk of complications. Thus, a pacemaker without primary batteries is desirable. The goal of this study was to test whether transcutaneous solar light could power a pacemaker. METHODS AND RESULTS We used a three-step approach to investigate the feasibility of sunlight-powered cardiac pacing. First, the harvestable power was estimated. Theoretically, a subcutaneously implanted 1 cm(2) solar module may harvest ∼2500 µW from sunlight (3 mm implantation depth). Secondly, ex vivo measurements were performed with solar cells placed under pig skin flaps exposed to a solar simulator and real sunlight. Ex vivo measurements under real sunlight resulted in a median output power of 4941 µW/cm(2) [interquartile range (IQR) 3767-5598 µW/cm(2), median skin flap thickness 3.0 mm (IQR 2.7-3.3 mm)]. The output power strongly depended on implantation depth (ρSpearman = -0.86, P < 0.001). Finally, a batteryless single-chamber pacemaker powered by a 3.24 cm(2) solar module was implanted in vivo in a pig to measure output power and to pace. In vivo measurements showed a median output power of >3500 µW/cm(2) (skin flap thickness 2.8-3.84 mm). Successful batteryless VVI pacing using a subcutaneously implanted solar module was performed. CONCLUSION Based on our results, we estimate that a few minutes of direct sunlight (irradiating an implanted solar module) allow powering a pacemaker for 24 h using a suitable energy storage. Thus, powering a pacemaker by sunlight is feasible and may be an alternative energy supply for tomorrow's pacemakers.

Basophils promote innate lymphoid cell responses in inflamed skin.

Type 2 inflammation underlies allergic diseases such as atopic dermatitis, which is characterized by the accumulation of basophils and group 2 innate lymphoid cells (ILC2s) in inflamed skin lesions. Although murine studies have demonstrated that cutaneous basophil and ILC2 responses are dependent on thymic stromal lymphopoietin, whether these cell populations interact to regulate the development of cutaneous type 2 inflammation is poorly defined. In this study, we identify that basophils and ILC2s significantly accumulate in inflamed human and murine skin and form clusters not observed in control skin. We demonstrate that murine basophil responses precede ILC2 responses and that basophils are the dominant IL-4-enhanced GFP-expressing cell type in inflamed skin. Furthermore, basophils and IL-4 were necessary for the optimal accumulation of ILC2s and induction of atopic dermatitis-like disease. We show that ILC2s express IL-4Rα and proliferate in an IL-4-dependent manner. Additionally, basophil-derived IL-4 was required for cutaneous ILC2 responses in vivo and directly regulated ILC2 proliferation ex vivo. Collectively, these data reveal a previously unrecognized role for basophil-derived IL-4 in promoting ILC2 responses during cutaneous inflammation.