Hypoxic Sites Research Articles

Pelagic oxycline and damage potential of hypoxia to the Pacific oyster Crassostrea gigas suspended in longline aquaculture systems

AEI Aquaculture Environment Interactions Contact the journal Facebook Twitter RSS Mailing List Subscribe to our mailing list via Mailchimp HomeLatest VolumeAbout the JournalEditorsTheme Sections AEI 9:461-468 (2017) - DOI: https://doi.org/10.3354/aei00245 Pelagic oxycline and damage potential of hypoxia to the Pacific oyster Crassostrea gigas suspended in longline aquaculture systems Sang Jun Lee1, Qtae Jo1,*, Jong-Cheol Han2, Yeong-Cheol Park3, Tae Gyu Park4 1Shellfish Aquaculture Lab, South Sea Fisheries Research Institute, NIFS, Jeonnam 59780, South Korea 2Shellfish Hatchery Center, Southeast Sea Fisheries Research Institute, NIFS, Gyeognam 52440, South Korea 3Environmental Analysis Lab, Southeast Sea Fisheries Research Institute, NIFS, Gyeognam 53085, South Korea 4Harmful Algal Bloom Lab, Southeast Sea Fisheries Research Institute, NIFS, Gyeognam 53085, South Korea *Corresponding author: qtjo@korea.kr ABSTRACT: Depletion in dissolved oxygen (DO) occurred in 2 semi-enclosed shallow bays in Korea in which Crassostrea gigas were suspended for aquaculture. DO depletion started at the bottom and expanded rapidly upwards for days until it was blocked by a temperature-driven density differential. This resulted in the formation of pelagic oxyclines at a depth of 2–4 m in one bay (Hypoxic Site 1, HS-1) and 2–5 m in the other (Hypoxic Site 2, HS-2). Water above the oxycline was normoxic (>4 mg l-1), while water below the oxycline was hypoxic (<1 mg l-1). The oxyclines were accompanied by significant changes in environmental variables and phytoplankton composition, but these changes were not fatal to the oysters. However, the oxyclines themselves caused oyster mortality: complete mortality below the oxyclines and depth-dependent mortality within the oxycline. Interestingly, mortality was observed in a significant number of oysters above the oxyclines compared with the reference site, and greater mortality was observed in HS-1 than HS‑2. These findings suggest the existence of toxic compounds that diffuse up from below the oxycline in shallow waters and exert effects that overshadow those of DO. The higher mortalities in the HS-1 normoxic layer supported this influence from the bottom layer. In a subsequent experiment, we observed additional mortalities among the surviving oysters in the upper normoxic waters after the bays had completely reoxygenated during fall turnover. These data provide useful observations of hypoxia in highly sheltered shallow waters and can be used to guide site selection for oyster longline aquaculture. KEY WORDS: Oxycline · Sheltered shallow coast · Oyster farm · Damage Full text in pdf format PreviousCite this article as: Lee SJ, Jo Q, Han JC, Park YC, Park TG (2017) Pelagic oxycline and damage potential of hypoxia to the Pacific oyster Crassostrea gigas suspended in longline aquaculture systems. Aquacult Environ Interact 9:461-468. https://doi.org/10.3354/aei00245 Export citation RSS - Facebook - Tweet - linkedIn Cited by Published in AEI Vol. 9. Online publication date: November 22, 2017 Print ISSN: 1869-215X; Online ISSN: 1869-7534 Copyright © 2017 Inter-Research.

Molecular and biochemical responses of hypoxia exposure in Atlantic croaker collected from hypoxic regions in the northern Gulf of Mexico.

A major impact of global climate change has been the marked increase worldwide in the incidence of coastal hypoxia (dissolved oxygen, DO<2.0 mg l-1). However, the extent of hypoxia exposure to motile animals such as fish collected from hypoxic waters as well as their molecular and physiological responses to environmental hypoxia exposure are largely unknown. A suite of potential hypoxia exposure biomarkers was evaluated in Atlantic croaker collected from hypoxic and normoxic regions in the northern Gulf of Mexico (nGOM), and in croaker after laboratory exposure to hypoxia (DO: 1.7 mg l-1). Expression of hypoxia-inducible factor-α, hif-α; neuronal nitric oxide synthase, nNOS; and insulin-like growth factor binding protein, igfbp mRNAs and protein carbonyl (PC, an oxidative stress indicator) content were elevated several-fold in brain and liver tissues of croaker collected from nGOM hypoxic sites. All of these molecular and biochemical biomarkers were also upregulated ~3-10-fold in croaker brain and liver tissues within 1–2 days of hypoxia exposure in controlled laboratory experiments. These results suggest that hif-αs, nNOS and igfbp-1 transcripts and PC contents are useful biomarkers of environmental hypoxia exposure and some of its physiological effects, making them important components for improved assessments of long-term impacts of environmental hypoxia on fish populations.

In situ delivery and production system of trastuzumab scFv with Bifidobacterium

A monoclonal antibody targeting human epidermal growth factor receptor-2 (HER2), trastuzumab has become a standard treatment for HER2-positive breast cancer. Recent advancements in antibody engineering have enabled the efficient generation of the trastuzumab single-chain variable fragment (scFv).In this study, we genetically engineered Bifidobacterium, a bacterial strain shown to accumulate safely and selectively in hypoxic tumor sites by intravenous (iv) injection, to express and secrete the trastuzumab scFv. The recombinant scFv bound to cell surface HER2 and inhibited in vitro growth of HER2-positive human cancer cells. Moreover, iv-injected recombinant bacteria specifically localized and secreted trastuzumab scFv in xenografted human HER2-positive tumors and consequently inhibited tumor growth.The development and results of this novel in situ delivery and production system for trastuzumab scFv with Bifidobacterium represents a promising avenue for future application in cancer treatment.

Vascular sprouts induce local attraction of proangiogenic neutrophils.

Angiogenesis, the growth of new blood vessels, is a complex process requiring the orchestration of numerous different cell types, growth factors, and chemokines. Some of the recently acknowledged actors in this process are immune cells. They accumulate at hypoxic sites, but the kinetics, dynamics, and regulation of that trafficking are unknown. In this study, we used intravital and live cell imaging to understand how neutrophils and macrophages migrate and behave at angiogenic sites. We developed two reproducible models of angiogenesis: one by transplanting isolated and hypoxic pancreatic islets into the cremaster muscles of mice, and another by in vitro coculturing of mouse aortic rings with neutrophils. In vivo imaging of the hypoxic site revealed recruitment of neutrophils and macrophages, which occurred in parallel, with depletion of one subset not affecting the accumulation of the other. We found, by cell tracking and statistical analyses, that neutrophils migrated in a directional manner to "angiogenic hotspots" around the islet where endothelial sprouting occurs, which was confirmed in the in vitro model of angiogenesis and is dependent on CXCL12 signaling. Intimate interactions between neutrophils and neovessels were prevalent, and neutrophil depletion greatly hampered vessel growth. Macrophages were less motile and attained supportive positions around the neovessels. Here, we present two novel in vivo and in vitro imaging models to study leukocyte behavior and actions during angiogenesis. These models unveiled that neutrophil migration at a hypoxic site was guided by signals emanating from sprouting endothelium where these immune cells gathered at "angiogenic hotspots" at which vascular growth occurred.

Blocking Surgically Induced Lysyl Oxidase Activity Reduces the Risk of Lung Metastases

SummarySurgery remains the most successful curative treatment for cancer. However, some patients with early-stage disease who undergo surgery eventually succumb to distant metastasis. Here, we show that in response to surgery, the lungs become more vulnerable to metastasis due to extracellular matrix remodeling. Mice that undergo surgery or that are preconditioned with plasma from donor mice that underwent surgery succumb to lung metastases earlier than controls. Increased lysyl oxidase (LOX) activity and expression, fibrillary collagen crosslinking, and focal adhesion signaling contribute to this effect, with the hypoxic surgical site serving as the source of LOX. Furthermore, the lungs of recipient mice injected with plasma from post-surgical colorectal cancer patients are more prone to metastatic seeding than mice injected with baseline plasma. Downregulation of LOX activity or levels reduces lung metastasis after surgery and increases survival, highlighting the potential of LOX inhibition in reducing the risk of metastasis following surgery.

Seasonal hypoxia was a natural feature of the coastal zone in the Little Belt, Denmark, during the past 8 ka

The extent of the hypoxic area in the Baltic Sea has rapidly expanded over the past century. Two previous phases of widespread hypoxia, coinciding with the Holocene Thermal Maximum (HTM; 8–4ka before present; BP) and the Medieval Climate Anomaly (MCA; 2–0.8ka BP), have been identified. Relatively little is known about bottom water redox conditions in the coastal zone of the Baltic Sea during the Holocene, however. Here we studied the geochemical composition of a sediment sequence from a currently seasonally hypoxic site in the Danish coastal zone, the Little Belt, retrieved during Integrated Ocean Drilling Program Expedition 347 (Site M0059). The base of the studied sediment sequence consists of clays low in organic carbon (Corg), molybdenum (Mo) and iron sulfides (Fe-sulfides), and rich in iron oxides (Fe-oxides), indicative of a well-oxygenated, oligotrophic (glacial) meltwater lake. An erosional unconformity separates the glacial lake sediments from sediments that are rich in Corg. The absence of Mo, in combination with high Corg/S values, indicates that these sediments were deposited in a highly productive, well-oxygenated freshwater lake. The transition to modern brackish/marine conditions was very rapid, and subsequent continuous sequestration of Mo in the sediment and high ratios of reactive iron (FeHR) over total Fe (FeTOT) suggest (seasonal) hypoxia occurred over the last ~8ka. Maxima in sediment Corg, Mo and FeHR/FeTOT ratios during the HTM and MCA suggest that the hypoxia intensified. Our results demonstrate that the Little Belt is naturally susceptible to the development of seasonal hypoxia. While periods of climatic warming led to increased deoxygenation of bottom waters, high nutrient availability in combination with density stratification were likely the main drivers of hypoxia in this part of the coastal zone of the Baltic Sea during the Holocene.

Abstract A29: Anti-CTLA-4 antibody scFv producing recombinant Bifidobacterium secretes CTLA-4 blocker specifically inside hypoxic tumor and suppresses tumor growth in syngeneic mice model

Abstract Success of immune checkpoint antibody drugs has provided broad perspective in cancer immunotherapy. Anti-PD-1 antibody and anti-CTLA-4 antibody showed notable efficacy and the combination complimentarily enhanced antitumor benefit. Nevertheless, either single-agent therapy or combination therapy is facing immune-related adverse events (irAEs), which is major cause of treatment discontinuation. Also, still large fraction of cancer patients doesn't respond to the antibody immunotherapy. Since non-specific systemic activation of normal immune system by the therapy is one of major reasons to cause the above problems, approaches to target the hypoxic condition of tumors may help increasing the drug's tumor-targeting specificity, which results in minimizing the irAEs and improving the response rate in patients. In an aim to improve anti-cancer drug delivery efficacy, we have been developing in situ Delivery and Production System (i-DPS) by modifying a non-pathogenic anaerobic bacterium, Bifidobacterium, which localizes and proliferates only in the hypoxic environment like solid tumors after intravenous administration, produces anticancer proteins, enzymes or other pharmacologically active molecules selectively at the tumor site. Here we present anti-human CTLA-4 antibody scFv producing i-DPS in cancer immunotherapy, which could be specifically delivered to and amplified only at the hypoxic sites of solid tumors. A series of in vitro assays has been performed to confirm the stable expression and secretion of anti-hCTLA-4 scFv by recombinant Bifidobacterium, the binding affinity of purified anti-hCTLA-4 scFv and the IC50 competing with hCTLA-4/hCD80 or hCD86 interaction as well. Anti-murine CTLA-4 scFv producing Bifidobacterium as surrogate systemically administered to the syngeneic mice model demonstrated significant tumor growth inhibition. Moreover, the combination therapy of anti-mCTLA-4 scFv producing Bifidobacterium and anti-murine PD-1 antibody was more effective than each of monotherapy. It is noteworthy that scFv was only detected in tumor tissue but not in blood proved by immunoprecipitation assay. Altogether, i-DPS for anti-CTLA-4 antibody provide a new promising immune-therapeutic modality to target hypoxic solid tumors and also provide a unique insight for antibody drug delivery in cancer immunotherapy. Citation Format: Koichiro Shioya, Shiro Kataoka, Li Wang, Tomio Matsumura, Hitomi Shimizu, Yasuyoshi Kanari, Yuji Seki, Yuko Shimatani, Minoru Fujimori, Shun'ichiro Taniguchi. Anti-CTLA-4 antibody scFv producing recombinant Bifidobacterium secretes CTLA-4 blocker specifically inside hypoxic tumor and suppresses tumor growth in syngeneic mice model. [abstract]. In: Proceedings of the AACR Special Conference on Tumor Immunology and Immunotherapy; 2016 Oct 20-23; Boston, MA. Philadelphia (PA): AACR; Cancer Immunol Res 2017;5(3 Suppl):Abstract nr A29.

Hypoxia inducible factors are dispensable for myeloid cell migration into the inflamed mouse eye

Hypoxia inducible factors (HIFs) are ubiquitously expressed transcription factors important for cell homeostasis during dynamic oxygen levels. Myeloid specific HIFs are crucial for aspects of myeloid cell function, including their ability to migrate into inflamed tissues during autoimmune disease. This contrasts with the concept that accumulation of myeloid cells at ischemic and hypoxic sites results from a lack of chemotactic responsiveness. Here we seek to address the role of HIFs in myeloid trafficking during inflammation in a mouse model of human uveitis. We show using mice with myeloid-specific Cre-deletion of HIFs that myeloid HIFs are dispensable for leukocyte migration into the inflamed eye. Myeloid-specific deletion of Hif1a, Epas1, or both together, had no impact on the number of myeloid cells migrating into the eye. Additionally, stabilization of HIF pathways via deletion of Vhl in myeloid cells had no impact on myeloid trafficking into the inflamed eye. Finally, we chemically induce hypoxemia via hemolytic anemia resulting in HIF stabilization within circulating leukocytes to demonstrate the dispensable role of HIFs in myeloid cell migration into the inflamed eye. These data suggest, contrary to previous reports, that HIF pathways in myeloid cells during inflammation and hypoxia are dispensable for myeloid cell tissue trafficking.

Differential expression of survival proteins during decreased intracellular oxygen tension in brain endothelial cells of grey mullets

The brain requires constant oxygen supply to perform its biological functions essential for survival. Because of low oxygen capacity and poor oxygen diffusibility of water, many fish species have evolved various adaptive mechanisms to cope with depleted oxygen. Endothelial cells (EC) are the primary components responsible for controlled environment of brain. Brain homeostasis largely depends on integrity of the EC. To elucidate their adaptive strategy, EC were isolated from the fish brain of Kovalam-control site and Ennore estuary-test/field hypoxic site and were subjected to low oxygen tension in laboratory. Cell viability, 4-hydroxynonenal (4HNE) and total antioxidant capacity (TAC) were analyzed to ascertain stress. Hypoxic insult, cytoprotective role of HSPs and apoptotic effect were analyzed by assessing hypoxia-inducible-factor-α (HIF1α), heat-shock-protein-70 (HSP70), heme-oxygenase 1 (HO-1), and apoptosis signal regulating kinase-1 (ASK1). This study evidenced that HSP70 and HO-1 are the key stress proteins, confer high tolerance to decreased oxygen tension mediated stress.

Multifunctional nanoplatform for enhanced photodynamic cancer therapy and magnetic resonance imaging

Co-delivery of photosensitizers and synergistic agents by one single nanoplatform is interesting for enhancing photodynamic therapy (PDT) of cancer. Here, a multifunctional nanoplatform for enhanced photodynamic therapy and magnetic resonance imaging of cancer was constructed. The poly (lactide-co-glycolide) (PLGA) nanoparticles (NPs) loaded with hematoporphyrin monomethyl ether (HMME) were coated with multifunctional manganese dioxide (MnO2) shells, which were designed as PLGA/HMME@MnO2 NPs. Once the NPs were effectively taken up by tumor cells, the intracellular H2O2 was catalysed by the MnO2 shells to generate O2. Meanwhile, the higher glutathione (GSH) promoted the degradation of MnO2 into Mn2+ ions with the ability of magnetic resonance (MR) imaging. After the degradation of outer layer, the release of photosensitizer was promoted. Under irradiation, the released HMME produced cytotoxic reactive oxygen species (ROS) to damage the tumor cells when the O2 was generated in the hypoxic tumor site. Furthermore, the decreased GSH level further inhibited the consumption of the produced ROS, which greatly enhanced the PDT efficacy. Therefore, this study suggested that this multifunctional system has the potential for enhanced photodynamic therapy and magnetic resonance imaging.

MEMS oxygen transport device for islet transplantation in the subcutaneous site

A new microelectromechanical systems (MEMS) oxygen transporter device to improve the efficacy of islet transplantation to treat type 1 diabetes is described here. The device is fabricated for biocompatibility and designed to maintain islet oxygenation levels throughout the normally hypoxic subcutaneous transplant site. Transplantation of the islets and the device to the subcutaneous location is minimally invasive. Bench-top testing and computational modelling of the device demonstrate that the device can provide sufficient oxygen to prevent hypoxia-induced islet death transplanted subcutaneously. In vitro studies show that the partial oxygen tension is well-maintained using the oxygen transporter in an anoxic environment. In vivo studies using a rat subcutaneous transplant model demonstrate that transplanted islets have more than three-times higher survival using the oxygen transporter compared with transplants not using the transporter, two weeks post-transplant. In addition, histology sections show well-maintained islet structures one month after transplantation for islets transplanted in the presence of the device. Our MEMS oxygen transporter is a novel and promising device to support the efficacy of subcutaneous islet transplantation as a potential cure for type 1 diabetes.

An O2 Self‐Sufficient Biomimetic Nanoplatform for Highly Specific and Efficient Photodynamic Therapy

Conventional oxygen‐dependent photodynamic therapy (PDT) has faced severe challenges because of the non‐specificity of most available photosensitizers (PSs) and the hypoxic nature of tumor tissues. Here, an O2 self‐sufficient cell‐like biomimetic nanoplatform (CAT‐PS‐ZIF@Mem) consisting of the cancer cell membrane (Mem) and a cytoskeleton‐like porous zeolitic imidazolate framework (ZIF‐8) with the embedded catalase (CAT) protein molecules and Al(III) phthalocyanine chloride tetrasulfonic acid (AlPcS4, defined as PS) is developed. Because of the immunological response and homologous targeting abilities of the cancer cell membrane, CAT‐PS‐ZIF@Mem is selectively accumulated at the tumor site and taken up effectively by tumor cells after intravenous injection. After the intracellular H2O2 penetration into the framework, it is catalyzed by CAT to produce O2 at the hypoxic tumor site, facilitating the generation of toxic 1O2 for highly effective PDT in vivo under near‐infrared irradiation. By integrating the immune escape, cell homologous recognition, and O2 self‐sufficiency, this cell‐like biomimetic nanoplatform demonstrates highly specific and efficient PDT against hypoxic tumor cells with much reduced side‐effect on normal tissues.

Monitoring Tumor Hypoxia Using (18)F-FMISO PET and Pharmacokinetics Modeling after Photodynamic Therapy.

Photodynamic therapy (PDT) is an efficacious treatment for some types of cancers. However, PDT-induced tumor hypoxia as a result of oxygen consumption and vascular damage can reduce the efficacy of this therapy. Measuring and monitoring intrinsic and PDT-induced tumor hypoxia in vivo during PDT is of high interest for prognostic and treatment evaluation. In the present study, static and dynamic 18F-FMISO PET were performed with mice bearing either U87MG or MDA-MB-435 tumor xenografts immediately before and after PDT at different time points. Significant difference in tumor hypoxia in response to PDT over time was found between the U87MG and MDA-MB-435 tumors in both static and dynamic PET. Dynamic PET with pharmacokinetics modeling further monitored the kinetics of 18F-FMISO retention to hypoxic sites after treatment. The Ki and k3 parametric analysis provided information on tumor hypoxia by distinction of the specific tracer retention in hypoxic sites from its non-specific distribution in tumor. Dynamic 18F-FMISO PET with pharmacokinetics modeling, complementary to static PET analysis, provides a potential imaging tool for more detailed and more accurate quantification of tumor hypoxia during PDT.

Impact of cable bacteria on sedimentary iron and manganese dynamics in a seasonally-hypoxic marine basin

Cable bacteria have recently been identified in various sedimentary marine settings worldwide. These filamentous microbes mediate electrogenic sulphur oxidation (e-SOx) over centimetre-scale distances, leading to a distinct separation of oxygen- and sulphide-bearing sediment zones. Here we present results of a year-long monthly assessment of the impact of cable bacteria on sedimentary Fe and Mn dynamics at three sites located along a water depth gradient in a seasonally-hypoxic coastal marine lake (Grevelingen, The Netherlands). Fluorescence In Situ Hybridisation (FISH) shows the presence of cable bacteria at two sites in spring. Micro-sensor profiling (O2, pH, H2S) and pore water profiles of dissolved Mn, Fe2+, Ca2+ and SO42− reveal the geochemical signature of e-SOx at these sites, i.e. the development of a broad suboxic zone, characterised by a low pH and acidic dissolution of Ca/Mn carbonates and Fe sulphides. Cable bacteria activity, as reflected by dissolution of FeS in spring, was highest at the deepest and most hypoxic site. In spring, dissolved Mn and Fe2+ released at depth due to e-SOx diffused upwards and was sequestered as Mn- and Fe-(oxyhydr)oxides near the sediment surface, with Mn oxides acting as an oxidant for part of the upward diffusing Fe2+. Strikingly, the thickness of the Fe-(oxyhydr)oxide-bearing surface layer of the sediment was greatest at the most hypoxic site, emphasising the key role of cable bacteria in creating oxidised surface sediments. X-ray absorption fine structure analyses confirm the seasonality in Fe-(oxyhydr)oxide formation and reveal that the sediment Mn oxides were of biogenic (birnessite) and abiotic (hausmannite) origin. Upon the onset of hypoxia in early summer, the sediment Fe-(oxyhydr)oxides were mostly converted to Fe-sulphides but the Mn oxides dissolved and the Mn was lost to the overlying water. After summer hypoxia, Beggiatoaceae mats colonised the sediment with little further change in sediment geochemistry. Our results confirm that cable bacteria act as a key control on the coupled cycling of Fe and Mn in surface sediments of seasonally hypoxic basins.

Glycolysis determines dichotomous regulation of T cell subsets in hypoxia.

Hypoxia occurs in many pathological conditions, including chronic inflammation and tumors, and is considered to be an inhibitor of T cell function. However, robust T cell responses occur at many hypoxic inflammatory sites, suggesting that functions of some subsets are stimulated under low oxygen conditions. Here, we investigated how hypoxic conditions influence human T cell functions and found that, in contrast to naive and central memory T cells (TN and TCM), hypoxia enhances the proliferation, viability, and cytotoxic action of effector memory T cells (TEM). Enhanced TEM expansion in hypoxia corresponded to high hypoxia-inducible factor 1α (HIF1α) expression and glycolytic activity compared with that observed in TN and TCM. We determined that the glycolytic enzyme GAPDH negatively regulates HIF1A expression by binding to adenylate-uridylate-rich elements in the 3'-UTR region of HIF1A mRNA in glycolytically inactive TN and TCM. Conversely, active glycolysis with decreased GAPDH availability in TEM resulted in elevated HIF1α expression. Furthermore, GAPDH overexpression reduced HIF1α expression and impaired proliferation and survival of T cells in hypoxia, indicating that high glycolytic metabolism drives increases in HIF1α to enhance TEM function during hypoxia. This work demonstrates that glycolytic metabolism regulates the translation of HIF1A to determine T cell responses to hypoxia and implicates GAPDH as a potential mechanism for controlling T cell function in peripheral tissue.

Semaphorin 7A Aggravates Pulmonary Inflammation during Lung Injury

The extent of pulmonary inflammation during lung injury ultimately determines patient outcome. Pulmonary inflammation is initiated by the migration of neutrophils into the alveolar space. Recent work has demonstrated that the guidance protein semaphorin 7A (SEMA7A) influences the migration of neutrophils into hypoxic tissue sites, yet, its role during lung injury is not well understood. Here, we report that the expression of SEMA7A is induced in vitro through pro-inflammatory cytokines. SEMA7A itself induces the production of pro-inflammatory cytokines in endothelial and epithelial cells, enhancing pulmonary inflammation. The induction of SEMA7A facilitates the transendothelial migration of neutrophils. In vivo, animals with deletion of SEMA7A expression showed reduced signs of pulmonary inflammatory changes following lipopolysaccharide challenge. We define here the role of SEMA7A in the development of lung injury and identify a potential pathway to interfere with these detrimental changes. Future anti-inflammatory strategies for the treatment of lung injury might be based on this finding.

Endothelial progenitor cell recruitment in a microfluidic vascular model

During vessel injury, endothelial progenitors cells (EPCs) are recruited from bone marrow and directed to the hypoxic injury site. The hypoxic conditions in the damaged blood vessel promote TNF-α, which upregulates intercellular adhesion molecule-1 (ICAM-1). EPCs attach to endothelial cell lining using ICAM-1. Here we aimed to examine EPC attachment to ECs in an injured-blood vessel conditions. We first determined ICAM-1 expression in stimulated HUVECs. We stimulated HUVECs with 21% oxygen (atmospheric), atmospheric with TNF-α-supplemented media, 1% oxygen (hypoxia), and hypoxia with TNF-α-supplemented media and found the highest ECFC attachment on HUVECs stimulated with TNF-α and hypoxia, correlating with the highest ICAM-1 expression. We next designed, fabricated and tested a three-dimensional microbioreactor (3D MBR) system with precise control and monitoring of dissolve oxygen and media flow rate in the cellular environment. We utilized a step-wise seeding approach, producing monolayer of HUVECs on all four walls. When stimulated with both TNF-α and hypoxia, ECFC retention on HUVECs was significantly increased under low shear stress compared to static controls. Overall, the 3D MBR system mimics the pathological oxygen tension and shear stress in the damaged vasculature, providing a platform to model vascular-related disorders.

Endothelial precursor cell-based therapy to target the pathologic angiogenesis and compensate tumor hypoxia

Hypoxia-inducing pathologies as cancer develop pathologic and inefficient angiogenesis which rules tumor facilitating microenvironment, a key target for therapy. As such, the putative ability of endothelial precursor cells (EPCs) to specifically home to hypoxic sites of neovascularization prompted to design optimized, site-specific, cell-mediated, drug-/gene-targeting approach. Thus, EPC lines were established from aorta–gonad–mesonephros (AGM) of murine 10.5 dpc and 11.5 dpc embryo when endothelial repertoire is completed. Lines representing early endothelial differentiation steps were selected: MAgEC10.5 and MagEC11.5. Distinct in maturation, they differently express VEGF receptors, VE-cadherin and chemokine/receptors. MAgEC11.5, more differentiated than MAgEC 10.5, displayed faster angiogenesis in vitro, different response to hypoxia and chemokines. Both MAgEC lines cooperated to tube-like formation with mature endothelial cells and invaded tumor spheroids through a vasculogenesis-like process. In vivo, both MAgEC-formed vessels established blood flow. Intravenously injected, both MAgECs invaded MatrigelTM-plugs and targeted tumors. Here we show that EPCs (MAgEC11.5) target tumor angiogenesis and allow local overexpression of hypoxia-driven soluble VEGF-receptor2 enabling drastic tumor growth reduction. We propose that such EPCs, able to target tumor angiogenesis, could act as therapeutic gene vehicles to inhibit tumor growth by vessel normalization resulting from tumor hypoxia alleviation.

Impaired gamete production and viability in Atlantic croaker collected throughout the 20,000 km2 hypoxic region in the northern Gulf of Mexico

The long-term impacts of recent marked increases in the incidence and extent of hypoxia (dissolved oxygen <2mg/L) in coastal regions worldwide on fisheries and ecosystems are unknown. Reproductive impairment was investigated in Atlantic croaker collected in 2010 from the extensive coastal hypoxic region in the northern Gulf of Mexico. Potential fecundity was significantly lower in croaker collected throughout the ~20,000km2 hypoxic region than in croaker from normoxic sites. In vitro bioassays of gamete viability showed reductions in oocyte maturation and sperm motility in croaker collected from the hypoxic sites in response to reproductive hormones which were accompanied by decreases in gonadal levels of membrane progestin receptor alpha, the receptor regulating these processes. The finding that environmental hypoxia exposure reduces oocyte viability in addition to decreasing oocyte production in croaker suggests that fecundity estimates need to be adjusted to account for the decrease in oocyte maturation.

Endothelial Progenitor Cells in Sprouting Angiogenesis: Proteases Pave the Way.

Sprouting angiogenesis consists of the expansion and remodelling of existing vessels, where the vascular sprouts connect each other to form new vascular loops. Endothelial Progenitor Cells (EPCs) are a subtype of stem cells, with high proliferative potential, able to differentiate into mature Endothelial Cells (ECs) during the neovascularization process. In addition to this direct structural role EPCs improve neovascularization, also secreting numerous pro-angiogenic factors able to enhance the proliferation, survival and function of mature ECs, and other surrounding progenitor cells. While sprouting angiogenesis by mature ECs involves resident ECs, the vasculogenic contribution of EPCs is a high hurdle race. Bone marrowmobilized EPCs have to detach from the stem cell niche, intravasate into bone marrow vessels, reach the hypoxic area or tumour site, extravasate and incorporate into the new vessel lumen, thus complementing the resident mature ECs in sprouting angiogenesis. The goal of this review is to highlight the role of the main protease systems able to control each of these steps. The pivotal protease systems here described, involved in vascular patterning in sprouting angiogenesis, are the matrix-metalloproteinases (MMPs), the serineproteinases urokinase-type plasminogen activator (uPA) associated with its receptor (uPAR) and receptorassociated plasminogen/plasmin, the neutrophil elastase and the cathepsins. Since angiogenesis plays a critical role not only in physiological but also in pathological processes, such as in tumours, controlling the contribution of EPCs to the angiogenic process, through the regulation of the protease systems involved, could yield new opportunities for the therapeutic prospect of efficient control of pathological angiogenesis.