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Cited by 101Tissue Engineering and Regenerative Medicine3D Printing in Biomedical ResearchCancer, Hypoxia, and Metabolism

Oxygen Regulation in Development: Lessons from Embryogenesis towards Tissue Engineering

Open access
Fathollahipour, Shahrzad, Patil, Pritam S., Leipzig, Nic D.·2018·Cells Tissues Organs
DOI 10.1159/000493162PMC PMC6397050
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Abstract

Oxygen is a vital source of energy necessary to sustain and complete embryonic development. Not only is oxygen the driving force for many cellular functions and metabolism, but it is also involved in regulating stem cell fate, morphogenesis, and organogenesis. Low oxygen levels are the naturally preferred microenvironment for most processes during early development and mainly drive proliferation. Later on, more oxygen and also nutrients are needed for organogenesis and morphogenesis. Therefore, it is critical to maintain oxygen levels within a narrow range as required during development. Modulating oxygen tensions is performed via oxygen homeostasis mainly through the function of hypoxia-inducible factors. Through the function of these factors, oxygen levels are sensed and regulated in different tissues, starting from their embryonic state to adult development. To be able to mimic this process in a tissue engineering setting, it is important to understand the role and levels of oxygen in each developmental stage, from embryonic stem cell differentiation to organogenesis and morphogenesis. Taking lessons from native tissue microenvironments, researchers have explored approaches to control oxygen tensions such as hemoglobin-based, perfluorocarbon-based, and oxygen-generating biomaterials, within synthetic tissue engineering scaffolds and organoids, with the aim of overcoming insufficient or nonuniform oxygen levels and nutrient supply.

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Written from the paper’s full text.

TL;DR

This review synthesizes how oxygen levels regulate embryonic development and how those lessons can inform tissue engineering, covering oxygen sensing, angiogenesis, measurement techniques, and oxygen-delivery biomaterials. It argues that mimicking developmental oxygen gradients could improve scaffolds and organoids, but accurate in situ oxygen measurement and safe, tunable delivery remain limiting.

Key findings

  • Oxygen requirements are stage-specific: early development occurs in physiologic hypoxia of 1–5% O2 (8–38 mmHg Po2), while pathologic hypoxia is below 1% or 8 mmHg Po2. Embryonic stem cells require low oxygen (~2%, 15 mmHg Po2) for proliferation and pluripotency; 3–5% O2 preserves the undifferentiated state in vitro versus 21% O2, and bovine blastocysts at 2% O2 had more ESCs in the inner cell mass. Placental cytotrophoblasts prefer ~18 mmHg Po2 (2.4% O2) for undifferentiated proliferation, with higher oxygen triggering differentiation, and placental oxygen rises from 18 to 60 mmHg after maternal vascular connection.
  • Oxygen acts as a morphogen and angiogenesis cue: rat embryos exposed to 60–95% O2 grew to 25 somites, whereas no growth occurred at 20% O2; chick embryos with limited oxygen transfer showed ~10% growth delay. In developing bone, oxygen tension varies from 21 mmHg in pre-hypertrophic regions to 57 mmHg in hypertrophic regions, and angiogenesis is initiated by hypoxia (<25 mmHg Po2). ARNT-deficient mice lacking HIF-1β died by E10.5 with arrested yolk sac vasculature.
  • Tissue engineering oxygen-delivery strategies include hemoglobin-based oxygen carriers, oxygen-generating biomaterials, perfluorocarbon-based carriers, and emerging technologies. Reported examples include HEMOXCell with a P50 of 35 mmHg at 37°C; calcium peroxide silk fibroin scaffolds with Po2 saturation of 10–11 mmHg over 21 days; PTMC/CaO2 releasing oxygen up to 20 days; H2O2-releasing microspheres reaching a maximum oxygen concentration of 18.4% (137 mmHg Po2); PFC nano-platforms transiently releasing 431.0 ± 0.5 mmHg oxygen over 200 sec; and TaOx@PFC-PEG nano-droplets increasing tissue oxygenation by 443 mmHg Po2.
  • Spheroids and organoids face hypoxic core formation, and several strategies aim to improve oxygen diffusion. Gas-permeable PDMS chips maintained an equilibrium oxygen tension of 50–60 mmHg Po2 after 5 days; Oxy chips showed 130 mmHg after 7 days versus 70 mmHg in non-Oxy controls; PDMS-CaO2 disks improved oxygen from 0.16 ± 0.017 mM (~114 mmHg Po2) in week 1 to an average of 0.073 ± 0.007 mM (~52 mmHg Po2) over 4 weeks. Spinning bioreactors, perfusion bioreactors, and microfluidics/bioprinting are also discussed for larger organoid culture.

Methods

  • Narrative review; no primary experiments are reported. It synthesizes literature on oxygen in embryonic development, oxygen homeostasis and sensing, embryonic angiogenesis, tissue oxygen monitoring, oxygen-delivery approaches, and spheroids/organoids.
  • Sources include cited animal and in vitro studies and prior reviews. The paper includes figures summarizing developmental oxygen levels and adult oxygen gradients and Table 1 classifying oxygen-delivery platforms.
  • Main analysis is a qualitative synthesis linking developmental oxygen biology to tissue engineering design criteria.

Limitations

  • Stated: exact tissue oxygen levels are difficult to measure experimentally, so relative changes across developmental stages are not fully known; the field lacks effective, accurate, and straightforward tools for quantifiable oxygen measures, especially within developing tissue.
  • Stated: oxygen-generating materials are affected by pH, temperature, buffer conditions, and catalysts/inhibitors; PFC toxicology has not been fully studied for promising compounds; HbOCs have not been proven safe in humans and may risk myocardial infarction; organoids/spheroids lack vasculature and develop necrotic cores.
  • Evident: as a narrative review, it does not report a systematic search strategy, inclusion criteria, or quantitative meta-analysis; many cited findings come from animal or in vitro models, so translation to human tissue engineering remains uncertain.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1159/000493162

Cites

Statistics

Nothing to recompute.

Claims the paper makes

0 of 205 cited sources checked · 0 supported · 0 flagged

  • Findingreasoning

    Researchers have explored hemoglobin-based, perfluorocarbon-based, and oxygen-generating biomaterials within synthetic tissue engineering scaffolds and organoids to overcome insufficient or non-uniform oxygen and nutrient supply.

    “Taking lessons from native tissue microenvironments, researchers have explored approaches to control oxygen tensions, such as hemoglobin-based, perfluorocarbon-based and oxygen generating biomaterials, within synthetic tissue engineering scaffolds and organoids with the aim of overcoming insufficient or non-uniform oxygen levels and nutrient supply.”
    Quote in the paperSubstantial claim
    • Rests on an assumption: Lessons from native tissue oxygen microenvironments can be translated into synthetic tissue engineering scaffolds and organoids.
  • Findingnone given

    The review found no recent reports that these new oxygen-delivery tools had been expanded into tissue-engineering uses.

    “Interestingly, no recent reports of expansion of these technologies in tissue engineering applications were found.”
    Quote in the paperSubstantial claim
  • Findingcitation

    Toxicological studies have not yet been performed for some of the most promising perfluorocarbon compounds and strategies.

    “Proper toxicological studies are yet to be performed for some of the most promising compounds and strategies”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Serex et al., 2014; Takahashi et al., 2014

      Not checked in the free run.

  • Findingnone given

    There is no consensus on accurate, reliable methods for measuring oxygen in vitro and in vivo.

    “the field is lacking agreed upon accurate and reliable technologies to measure the levels of oxygen both in vitro and in vivo.”
    Quote in the paperSubstantial claim
  • Interpretationnone given

    Later developmental stages require more oxygen and nutrients for organogenesis and morphogenesis.

    “Later, more oxygen, as well as nutrients are needed for organogenesis and morphogenesis.”
    Quote in the paperNot a substantial claim
  • May combine two claims. 99
  • Classified as probably not a substantial claim. 27

  • May combine two claims.

    Findingcitation

    Research has developed synthetic oxygen carriers, oxygen-generating matrices, and perfluorocarbon-associated technologies to facilitate oxygen transport or direct delivery.

    “This approach includes the development and study of synthetic oxygen carriers [Eisenbrey et al., 2015], oxygen generating matrices [Oh et al., 2009] and perfluorocarbon-associated technologies (PFCs) [Geyer, 1988].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Eisenbrey et al., 2015

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    • cites Oh et al., 2009

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    • cites Geyer, 1988

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  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Mitochondrial respiration accounts for the majority of oxygen consumption in humans.

    “mitochondrial respiration accounts for the majority of O2 consumption in humans [Boveris et al., 1999]”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Boveris et al., 1999

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  • Probably not a substantial claim: may not pass S4 (based). May combine two claims.

    Interpretationnone given

    Low oxygen levels are the naturally preferred microenvironment for most early developmental processes and mainly drive proliferation.

    “Low oxygen levels are the naturally preferred microenvironment for most processes during early development and drive mainly proliferation.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Oxygen is essential for sustaining and completing embryonic development and regulates stem cell fate, morphogenesis, and organogenesis.

    “Oxygen is a vital source of energy necessary to sustain and complete embryonic development. Not only is oxygen the preferred driving force for many cellular functions and metabolism, but it also is involved in regulating stem cell fate, morphogenesis, and organogenesis.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    The role and effects of oxygen availability on mammalian embryo morphogenesis in vivo are poorly understood because mammalian embryo oxygenation depends on maternal supply and cannot be easily isolated.

    “Despite these early studies, the role and effects of oxygen availability on mammalian embryo morphogenesis in vivo have not been thoroughly investigated and are poorly understood. This is primarily because mammalian embryo oxygenation depends highly on the maternal supply, and thus the effects of oxygen variation in a mammalian fetus cannot be easily isolated or assessed directly without changing”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Giussani et al., 2007

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    Premisecitation

    The blastocyst serves as the niche and source for embryonic stem cells and maintains the low oxygen microenvironment required at this stage.

    “The blastocyst serves as the niche and source for ESCs, which maintains the required low oxygen microenvironment for this stage of development.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Reduced oxygen tension of 4% is crucial for maintaining pluripotency of human embryonic stem cells and encouraging proliferation in an undifferentiated state in vitro.

    “Likewise, Narva et al. have shown that reduced oxygen tension (4% oxygen, 30 mmHg Po2) is crucial for maintaining the pluripotency of human embryonic stem cells (hESCs) to encourage proliferation in an undifferentiated state in vitro [Narva et al., 2013].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Narva et al., 2013

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    Premisecitation

    In the human embryo, vascular invasion occurs at embryonic day 9, and by day 14-16 almost all small arterioles are formed, allowing optimal oxygen and nutrient transfer to the fetus.

    “In the human embryo, the vascular invasion occurs at embryonic day 9 (E9), and by the end of day 14–16, almost all of the small arterioles are entirely formed, allowing for optimal oxygen and nutrient transfer to the developing fetus [Enders and King, 1991].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Enders and King, 1991

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    Premisecitation

    Chick eggs incubated with 60% oxygen formed significantly heavier eggs on day 18 than controls at 21% oxygen, confirming that chick embryonic growth is regulated by oxygen availability.

    “on day 18 the group incubated with 60% oxygen formed significantly heavier eggs (indicative of embryonic growth) in contrast to the control group exposed to 21% oxygen (160 mmHg Po2),”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Metcalfe et al., 1981

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    Premisereasoning

    Chick embryo development is independent of maternal factors, leading many investigators to focus on oxygen availability in chick embryogenesis.

    “The independence of chick embryo development from maternal factors has led many investigators to focus on the influence of oxygen availability on chick embryogenesis.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Explanted rat embryos at the headfold stage exposed to 60-95% oxygen in watch glass culture grew up to 25 somites, whereas no growth was observed at 20% oxygen.

    “They showed that exposing the explanted rat embryos, at the headfold stage, to 60–95% oxygen (456–722 mmHg Po2) in watch glass culture helped the embryos to grow up to 25 somites. Conversely, no growth was observed in watch glass cultures with 20% oxygen (152 mmHg Po2) [New and Coppola, 1970].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites New and Coppola, 1970

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    Premisereasoning

    Maintaining an undifferentiated state and the eventual fate of placental cytotrophoblasts depends on oxygen availability.

    “Thus, maintaining an undifferentiated state and the eventual fate of placental cytotrophoblasts depends on oxygen availability.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Low oxygen tension in early development results from limited connections with maternal circulation and is required for maintaining pluripotency in embryonic stem cells.

    “This low oxygen tension is the result of limited connections with maternal circulation during early development and is required for maintaining pluripotency in ESCs [Abdollahi et al., 2011].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Abdollahi et al., 2011

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    Premisecitation

    Undifferentiated proliferation of cytotrophoblasts occurs in vitro under low oxygen pressures of 2%, while higher oxygen tensions trigger differentiation into a phenotype required to generate the maternal circulatory system.

    “Studies indicate the undifferentiated proliferation of cytotrophoblasts occurs in vitro under low oxygen pressures (2% oxygen, 15 mmHg Po2). However, higher oxygen tensions trigger these cells to differentiate into a phenotype required to generate the maternal circulatory system [Genbacev et al., 1997; Simon and Keith, 2008].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Genbacev et al., 1997

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    • cites Simon and Keith, 2008

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  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    HIFs, PHDs, FIH-1, AP-1, NF-κB, p53, and c-Myc are the most well-understood factors influencing oxygen regulation in vivo.

    “Hypoxia-inducible factors (HIFs), prolyl hydroxylases (PHDs), factor-inhibiting HIF-1 (FIH-1), activator protein 1 (AP-1), nuclear factor (NF)-κB, p53 and c-Myc are the most well-understood factors that influence oxygen regulation in vivo [Podar and Anderson, 2010].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Podar and Anderson, 2010

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  • Probably not a substantial claim: may not pass S4 (based). May combine two claims.

    Premisenone given

    Oxygen levels directly correlate with cell growth, ECM synthesis, and the success of a tissue engineering approach during growth.

    “As mentioned before, oxygen levels directly correlate with cell growth, ECM synthesis and the success of a tissue engineering approach during growth are essential to monitor.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Fertilized eggs from sea level hens incubated at high altitude around 13% oxygen had increased embryonic mortality and decreased growth at day 20 compared with sea level at 21% oxygen.

    “fertilized eggs laid by sea level hens were incubated in high altitude (~13% oxygen, 100 mmHg Po2) and resulted in increased embryonic mortality as compared to those incubated at the sea level (21% oxygen, 160 mmHg Po2). Decreased growth at the study endpoint (day 20) was also observed in the same conditions”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Giussani et al., 2007

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    Premisenone given

    Common techniques to directly measure oxygen levels include delayed fluorescence of PPIX, intravascular phosphorescent probes, cell-penetrating small molecule phosphorescent nanosensors, and solid-state phosphorescent sensors, all usable for imaging and quantitative measurement.

    “Some of the most common techniques to directly measure the oxygen levels include delayed fluorescence of endogenously overproduced protoporphyrin IX (PPIX), intravascular (cell-impermeable) phosphorescent probes and cell-penetrating small molecule phosphorescent nanosensors, and solid-state phosphorescent sensors - all of which can be used for imaging and to quantitatively measure cell and tissue”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Angiogenesis establishes vasculature and is regulated by tissue demands including sufficient oxygen supply, removal of carbon dioxide and other respiratory gases, and sufficient nutrients.

    “Angiogenesis is the developmental process that establishes vasculature and is further regulated by the individual demands of tissues. These demands include a sufficient oxygen supply, removal of excess carbon dioxide and other respiratory gases, and sufficient nutrients [Rouwkema and Khademhosseini, 2016; Yoon and Jones, 2016].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Rouwkema and Khademhosseini, 2016

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    • cites Yoon and Jones, 2016

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  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisenone given

    Angiogenesis occurs through a sequence of stages: angiogenic stimulus, sprouting of endothelial cells, elongation and branching, lumen formation, and regression.

    “Angiogenesis occurs through a sequence of stages starting with the angiogenic stimulus, sprouting of endothelial cells, elongation and branching, the formation of the lumen and finally regression.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Premisenone given

    Optical fluorescence signals of endogenous PPIX reporter are weak and need enhancement in animal or human tissues to enable oxygen measurement.

    “However, the optical fluorescence signals of the endogenous PPIX reporter is weak and needs to be enhanced in animal or human tissues to enable oxygen measurement.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisenone given

    Enhancement of PPIX signal is usually done by treating tissues with 5-aminolevulinic acid to increase mitochondrial PPIX levels and specific optical signal.

    “This enhancement is usually done by treating tissues with 5-aminolevulinic acid (ALA) to increase mitochondrial PPIX levels and the specific optical signal.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Embryonic angiogenesis is needed for organ development when oxygen levels are insufficient for developmental requirements, whereas adult angiogenesis commonly occurs in response to insufficient blood and oxygen supply in wound healing, reproduction, and exercise.

    “From a fundamental perspective, angiogenesis in adults is different from the embryonic process in the sense that an embryo needs this process for organ development when oxygen levels are not enough to satisfy developmental requirements. In contrast, in an adult, angiogenesis commonly only occurs as a response to insufficient blood and oxygen supply in instances such as wound healing, reproduction,”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Premisenone given

    Direct optical sensing presents moderate selectivity and resolution especially for quantitative oxygen measurement.

    “Despite the diverse applications of this method, it presents moderate selectivity and resolution especially for quantitative oxygen measurement.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    HIFs are typically considered the body's master oxygen sensors and belong to a family of transcriptional factors with many downstream actions.

    “Out of these, HIFs are typically considered as the body’s ‘master oxygen sensors’ and belong to a family of transcriptional factors with many downstream actions [Bryant et al., 2018].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Bryant et al., 2018

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    Premisecitation

    Hypoxia below 5% oxygen triggers endothelial cells to express angiogenic signaling factors including VEGF-A and eNOS, resulting in migration and proliferation of endothelial cells.

    “Accordingly, hypoxia (<5% oxygen, 38 mmHg Po2) triggers endothelial cells to express angiogenic signaling factors, including vascular endothelial growth factor (VEGF)-A and nitric oxide synthase (eNOS) - resulting in subsequent migration and proliferation of endothelial cells [Cross et al., 2003].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Cross et al., 2003

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    Premisecitation

    HIFs largely mediate angiogenesis in hypoxia, and HIF-1 target genes play an essential role in angiogenesis.

    “The HIFs largely mediate angiogenesis in hypoxia and among the HIF family, HIF-1 target genes have been shown to play an essential role in angiogenesis. [Schipani et al., 2009].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Schipani et al., 2009

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    Premisecitation

    Hypoxia triggers endothelial cells to activate HIFs; HIF-α translocates into the nucleus, attaches to HIF-β, and the complex enters the angiogenic signaling pathway modulating capillary and vessel formation.

    “In this regard, as soon as hypoxia is sensed by the endothelial cells, they activate their master oxygen sensors, the HIFs. HIF-α activates and translocates into the endothelial cell nucleus and attaches to HIF-β. The complex then enters the angiogenic signaling pathway modulating the formation of capillaries and vessels [Pugh and Ratcliffe, 2003; Krock et al., 2011].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Pugh and Ratcliffe, 2003

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    • cites Krock et al., 2011

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    Premisecitation

    ARNT (HIF-1β) and HIF-1α are expressed in embryonic stem cells and are required for regulation of hypoxia-responsive genes and angiogenesis.

    “ARNT (aryl hydrocarbon receptor nuclear translocator: HIF-1β) and HIF-1α are both expressed in ESCs and are required for the regulation of hypoxia-responsive genes and angiogenesis.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Premisenone given

    Local oxygen pressures differ within vascular channels based on function, location, and metabolic requirements of adjacent tissue.

    “Local oxygen pressures are different within vascular channels based on their function, location and metabolic requirements of the adjacent tissue, the.”
    Quote in the paperNot a substantial claim

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    Premisecitation

    HIF-deficient mouse embryos develop improper placental structures and impaired vasculature.

    “Furthermore, HIF-deficient mice embryos have been shown to develop improper placental structures as well as impaired vasculature [Dunwoodie, 2009].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Dunwoodie, 2009

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    Premisenone given

    Direct optical sensing of oxygen uses metal-free endogenously produced PPIX as an oxygen-sensing reporter.

    “Direct optical sensing of oxygen is a common method also known as endogenous delayed fluorescence, which uses a metal-free endogenously produced precursor of heme called PPIX as an oxygen-sensing reporter.”
    Quote in the paperNot a substantial claim

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    Premisecitation

    ARNT-deficient mice lacking HIF-1β under low oxygen at 1.5% did not survive past E10.5 due to arrested vasculature in the yolk sac, indicating the importance of HIF expression in embryonic angiogenesis.

    “To test this hypothesis, Maltepe et al. used ARNT-deficient mice (lacking HIF-1β) to investigate the role of ARNT/HIF-1β in the regulation of the genes that are responsive to hypoxia under low oxygen conditions (1.5% oxygen, 12 mmHg Po2) [Maltepe et al., 1997]. The animals did not survive past E10.5, due to the arrested vasculature in the structure of their yolk sac, indicating the importance of”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Maltepe et al., 1997

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    Premisecitation

    Pre-hypertrophic cells at approximately 21 mmHg are more proliferative and stem cell-like, while hypertrophic terminally differentiated cells exist at approximately 57 mmHg.

    “the cell population present in the pre-hypertrophic areas exist at an oxygen tension of approximately 21 mmHg, are more proliferative which aligns with their more stem cell-like nature. The cell type, population, and oxygen level in the pre-hypertrophic region is contrasted by the hypertrophic region that is comprised of a population of terminally differentiated cells existing at a higher oxygen”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites Araldi and Schipani, 2010

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    • cites Rankin et al., 2011

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    • cites Stamati et al., 2011

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    Premisecitation

    Although hypoxia is often considered the driving force for endothelial cells to initiate vascularization, a growing body of literature suggests vessel growth is inhibited by both moderate and severe hypoxic conditions via an unknown mechanism.

    “It is noteworthy to mention that although hypoxia is often considered as the driving force for the endothelial cells to initiate the vascularization process[Krock et al., 2011], there is a growing body of literature suggesting that vessel growth is inhibited by both moderate and severe hypoxic conditions via a still unknown mechanism [Faller, 1999; Hutton and Grayson, 2016].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Krock et al., 2011

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    • cites Faller, 1999

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    • cites Hutton and Grayson, 2016

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    Premisecitation

    Hemoglobin blended with gelatin/fibrinogen scaffolds improves O2 diffusivity and has potential to promote cardiomyogenic differentiation of mesenchymal stem cells.

    “Specifically, hemoglobin blended with gelatin/fibrinogen scaffolds have been shown to improve O2 diffusivity and exhibited the potential to promote cardiomyogenic differentiation of mesenchymal stem cells (MSCs) [Ravichandran et al., 2013].”
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    • cites Ravichandran et al., 2013

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  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Increased oxygen availability causes more inhibition in the reductive mechanism and activation of EF5.

    “Increased oxygen availability in the environment causes more inhibition in the reductive mechanism and activation [Koch, 2002].”
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    • cites Koch, 2002

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    Premisecitation

    Electron paramagnetic resonance oximetry is invasive, requires implantation or intravenous injection of a paramagnetic material and PET imaging, has limited detection range to hypoxic conditions, and is costly.

    “A precise method for measuring oxygen values within tissues is electron paramagnetic resonance (EPR), which is invasive and difficult in that it requires the implantation or intravenous injection of a paramagnetic material as well as imaging via positron emission tomography (PET) scanning. The problem with this technique is it’s a limited range of detection to hypoxic conditions and high costs”
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    • cites Schrey et al., 2010

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    Premisecitation

    Hemoglobin is an iron-containing metalloprotein that exists in tetramer form in red blood cells.

    “Hemoglobin is an iron-containing metalloprotein existing in tetramer form in red blood cells [Jensen et al., 1998].”
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    • cites Jensen et al., 1998

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    Premisenone given

    Indirect oximetry methods include pH measurement, lactate concentration, or glucose consumption.

    “Another common type of oxygen measurement is performed indirectly via oximetry integrating methods such as pH measurement, lactate concentration or glucose consumption.”
    Quote in the paperNot a substantial claim

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    Premisecitation

    Pimonidazole binds to thiol-containing proteins in hypoxic cells, which can be quantitatively detected and converted indirectly to oxygen concentration using point counting and image analysis.

    “Pimidazole can bind to thiol-containing proteins present in hypoxic cells, which can then be quantitatively detected and converted indirectly to oxygen concentration using point counting and image analysis software methods [Varia et al., 1998].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Varia et al., 1998

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    Premisecitation

    In an implantable strategy, sustained controlled oxygen allows time for early neovascularization in vivo to support engineered tissue and maintain cellular metabolic activity and signaling.

    “In an implantable strategy, a sustained and controlled level of oxygen allows enough time for early neovascularization to occur in vivo to support the engineered tissue allowing maintenance of cellular metabolic activity and signaling [Lovett et al., 2009].”
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    • cites Lovett et al., 2009

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    Premisecitation

    Unmodified cell-free hemoglobin or methemoglobin blocks renal tubules causing kidney failure and acts as a pro-inflammatory molecule inducing oxidative toxicity in kidneys, liver, central nervous system, and cardiac tissue.

    “Unfortunately, this unmodified cell-free hemoglobin, or methemoglobin, blocks renal tubules leading to kidney failure, while also acting as a pro-inflammatory molecule that induces oxidative toxicity in kidneys, the liver, the central nervous system and in cardiac tissue [Harrison et al., 1947; Kumar and Bandyopadhyay, 2005].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Harrison et al., 1947

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    • cites Kumar and Bandyopadhyay, 2005

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    Premisereasoning

    MRI with fluorine contrast agents can quantitate tissue oxygen levels by using the 19F relaxation rate to monitor Po2 or create organ oxygen distribution maps.

    “Similarly, magnetic resonance imaging (MRI) techniques using fluorine contrast agents have also been used to quantitate tissue oxygen levels [De Santis and Singer, 2015; Papkovsky and Dmitriev, 2018].”
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    • cites De Santis and Singer, 2015

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    • cites Papkovsky and Dmitriev, 2018

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    Premisecitation

    Pimonidazole has been used clinically as a hypoxia probe to indirectly measure tumor hypoxia.

    “Similar to EF5, pimonidazole has also been used clinically as a hypoxia probe to indirectly measure levels of tumor hypoxia [Nordsmark et al., 2003].”
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    • cites Nordsmark et al., 2003

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    Premisecitation

    Electrochemical oxygen sensors often suffer from invasiveness, oxygen consumption by the electrode, and inability to reflect tissue oxygen heterogeneity.

    “These electrochemical sensors, however, are often faced by challenges such as invasiveness, oxygen consumption by the electrode, and inability to reflect the heterogeneity of tissue oxygen levels [Liu et al., 2011].”
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    • cites Liu et al., 2011

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  • May combine two claims.

    Premisecitation

    Near-infrared spectroscopy of hemoglobin saturation measures mean oxygen levels in large tissue volumes and in blood.

    “A commonly used clinical method for detecting blood oxygen levels uses near-infrared (NIR) spectroscopy of hemoglobin (Hb) saturation to measure mean oxygen levels in large tissue volumes, as well as blood (pulse oximetry).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Scheeren et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    The underlying theme of these hemoglobin modification strategies is to increase size and reduce reactivity to diminish toxicity and renal filtration.

    “The underlying theme behind these strategies is to increase the size and reduce reactivity to diminish the toxicity and renal filtration.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Polarographic oximetry is the gold standard for oxygen measurement, with good reproducibility, accuracy, low sensor deviation, and high detection resolution.

    “the oximetry gold standard presenting advantages such as good reproducibility and accuracy as well as the low deviation between each sensor and high detection resolution [Park et al., 2007].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Park et al., 2007

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    High costs and the need for patient cooperation limit the practical application of fluorine MRI oximetry.

    “High costs of using this technique as well as the requirement for patient cooperation can be considered as main drawbacks that limit it’s practical application [Aydogdu et al., 2012].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Aydogdu et al., 2012

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    EF5/2-nitroimidazole has been employed to study hypoxic tumor environments.

    “One common indirect approach is to use the localization and quantification of an drug that undergoes oxidative metabolism, such as EF5/2-nitroimidazole, (2-(2-nitro-1H-imidazole-1-yl)-N-(2,2,3,3,3-pentafluoropropyl)acetamide), as have been employed to study hypoxic tumor environments [Evans et al., 2000].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Evans et al., 2000

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Cell-free Hb-based artificial blood oxygen carriers have not been proved safe in humans and pose a risk of myocardial infarction.

    “Despite the considerable time and effort spent on the development of Hb-based oxygen carriers, it is important to highlight that cell-free Hb-based artificial blood oxygen carriers to date have not been proved safe in humans, which is concerning because their application poses the risk of myocardial infarction [Natanson et al., 2008].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Natanson et al., 2008

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Electron paramagnetic resonance can detect Po2 within 10-100 mmHg but becomes increasingly inaccurate outside this range.

    “EPR can detect Po2 within the range of 10–100 mmHg, but becomes increasingly inaccurate beyond this range [Presley et al., 2006].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Presley et al., 2006

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisenone given

    Currently available oxygen delivery platforms are broadly classified as hemoglobin-based, perfluorocarbon-based, oxygen generating materials, and a few other recently introduced classes.

    “Currently available platforms for oxygen delivery are broadly classified as hemoglobin-based, perfluorocarbon-based, oxygen generating materials, as well as a few other recently introduced classes (Table 1).”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Table 1

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Intracellular adducts of EF5 can be detected using antibodies, quantified using immunohistochemistry, and correlated with oxygen availability.

    “The intracellular adducts of EF5 can subsequently be detected using antibodies, quantified using immunohistochemistry and correlated with oxygen availability [Evans et al., 2004].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Evans et al., 2004

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Fiber-optic probes for tissue oxygen measurement are limited by invasiveness and probe fragility.

    “Fiber-optic probes are another alternative to measure the tissue oxygen levels which are limited by invasiveness as well as probe fragility [Weyand et al., 2015].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Weyand et al., 2015

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    At standard temperature and pressure, O2 solubility in water is 2.2 mM, and can be up to 44 mM for PFCs like bis(perfluorobutyl)ethene, a 20-fold increase.

    “At standard temperature and pressure, the solubility of O2 in water is 2.2 mM. This value can be much higher, up to 44 mM, for PFCs like bis(perfluorobutyl)ethene, representing a 20-fold increase over O2 solubility in water alone [Lowe et al., 1998].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Lowe et al., 1998

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    HEMOXCell improved oxygenation to hypoxic regions and promoted proliferation of human bone marrow MSCs.

    “In a bone tissue engineering application, HEMOXCell improved oxygenation to the hypoxic regions and promoted the proliferation of human bone marrow MSCs [Le Pape et al., 2018].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Le Pape et al., 2018

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing), S4 (based).

    Premisenone given

    The chemical composition of oxygen-generating materials, such as metal oxides or metal peroxides, determines their oxygenation characteristics.

    “The chemical composition of oxygen-generating materials, such as metal oxides or metal peroxides determines their oxygenation characteristics.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Native hemoglobin solutions show sigmoidal O2 dissociation, whereas colloidal PFC suspensions show a linear relationship between oxygen content and Po2.

    “Solutions of native hemoglobin show sigmoidal O2 dissociation behavior [Lowe et al., 1998], whereas, colloidal suspensions of PFCs, such as perfluorotributylamine, perfluorodecalin, perfluoro-octane, show a linear relationship between oxygen content and oxygen partial pressure (Po2) [Lowe et al., 1998; Spahn, 2000; Khattak et al., 2007].”
    Quote in the paperNot a substantial claim0 of 3 sources checked
    • cites Lowe et al., 1998

      Not checked in the free run.

    • cites Spahn, 2000

      Not checked in the free run.

    • cites Khattak et al., 2007

      Not checked in the free run.

  • May combine two claims.

    Premisenone given

    Stem cells preserve their undifferentiated state and proliferate in hypoxia below 5% oxygen or Po2 below 38 mmHg.

    “As discussed earlier in this review, stem cells are known to preserve their undifferentiated state and proliferate in hypoxia (<5% or Po2 < 38 mmHg)”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Some PFCs have been approved by the FDA as blood substitute products for specific applications, including Fluosol emulsions and perfluorodecalin emulsified with Pluronic F-68 for coronary artery balloon angioplasty and liquid ventilation.

    “Owing to their ability to carry oxygen, some PFCs in the past have been approved by the FDA as blood substitute products for specific applications. These products include Fluosol emulsions, and perfluorodecalin emulsified primarily with the synthetic poloxamer Pluronic F-68, both of which were approved for use in coronary artery balloon angioplasty procedures and as liquid ventilation procedures [”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Waxman, 1986

      Not checked in the free run.

  • May combine two claims.

    Interpretationnone given

    Published literature demonstrates limited understanding of biological interactions with PFCs, and not all PFCs have been studied for short- and long-term toxicity.

    “Despite FDA approvals, published literature demonstrates a limited understanding of biological interactions with PFCs and not all PFCs have been studied for short and long-term toxicity responses.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisereasoning

    Local oxygen concentration in MACF hydrogels leads to enhanced NSPC cell proliferation and neuronal differentiation, suggesting measured oxygen tensions translated to enhanced cellular functions.

    “Further, the local oxygen concentration in MACF hydrogels leads to enhanced NSPC cell proliferation and neuronal differentiation suggesting that the measured oxygen tensions translated to enhanced cellular functions.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Li et al., 2014

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    A poly(trimethylene carbonate) matrix with calcium peroxide particles showed step release of oxygen for up to 20 days and maintained human MSCs under hypoxic conditions with increased mitochondrial activity and enhanced proliferation.

    “Finally, the beneficial effects of a poly(trimethylene carbonate) matrix with calcium peroxide particles (PTMC/CaO2 0.05–0.1 mg/L; 0.55–1.7 mmHg Po2 above control) showed step release of oxygen for up to 20 days. This biomaterial showed beneficial maintenance of human MSCs under hypoxic conditions confirmed by increased mitochondrial activity and enhanced proliferation [Steg et al., 2017].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Steg et al., 2017

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Encapsulated FC-43 emulsion showed no significant improvement on proliferation of pancreatic beta cells (BTC-tet) in hypoxic conditions and exhibited cellular toxicity marked by reduced metabolic activity and viability compared to no PFC controls.

    “In another study, an encapsulated perfluorotributylamine (FC-43) emulsion was shown to have no significant improvement on the proliferation of pancreatic beta cells (BTC-tet) in hypoxic conditions [Goh et al., 2010]. Further, the perfluorotributylamine (FC-43) emulsion exhibited cellular toxicity marked by a reduction in metabolic activity and viability as compared to no PFC controls.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Goh et al., 2010

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Oxygenating biomaterials or carriers such as HEMOXCell can overcome chronic hypoxia around 1% Po2 or 8 mmHg and sustain a mild hypoxic environment.

    “Oxygenating biomaterials or carriers such as HEMOXCell can be used to overcome chronic hypoxia (~1% Po2, Po2 8 mmHg) and sustain a mild hypoxic environment.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Normal arterial and ventricular oxygen tension is about 75-100 mmHg, and CDCs prefer this oxygen microenvironment to maintain their functions.

    “Normal arterial and ventricular oxygen tension is about 75–100 mmHg, and thus it can be concluded that CDCs prefer this oxygen microenvironment to maintain their functions.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Oxysite generates oxygen in situ through calcium peroxide hydrolytic decomposition and mitigates anaerobic glycolysis and preserves and stimulates insulin release in rat pancreatic islet culture.

    “Coronel et al. have introduced Oxysite®, generating oxygen in situ through calcium peroxide hydrolytic decomposition, and showed its ability to mitigate anaerobic glycolysis and preserve and stimulate insulin release in rat pancreatic islet culture (oxygen gradient of 0.8 × 10−4 mM (0.1 mmHg Po2) on the cell side to 0.3 mM (214 mmHg Po2) in Oxysite) [Coronel et al., 2017].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Coronel et al., 2017

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    PFO emulsion loaded microparticles sustained more viable cells compared to controls under hypoxia, suggesting limited direct toxicity.

    “In vitro toxicity response was studied by cell number quantification, which was reduced by the effects of hypoxia, but PFO emulsion loaded MPs were able to sustain more viable cells as compared to controls suggesting limited direct toxicity.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Lee et al., 2015

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Methylated pyridone-derived endoperoxides undergo retro-Diels-Alder reactions in aqueous environments, releasing oxygen in high yields with half-lives up to 13 h.

    “Methylated pyridone-derived endoperoxides undergo retro-Diels-Alder reactions in an aqueous environment releasing oxygen in high yields and with half-lives of up to 13 h, however, this study used a fluorescence-based assay that only provided relative measures of oxygen abundance [Benz et al., 2013].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Benz et al., 2013

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Ali5, Ar5, and PFOC can be attached covalently to a UV-curable methacrylamide chitosan hydrogel (MACF) and used for tunable oxygen delivery to accelerate full thickness acute wound healing in a splinted rat model.

    “Our group has recently shown that pentafluoropropionic anhydride (Ali5, equilibrium 8 ± 1 mmHg Po2 above control), 2,3,4,5,6-pentafluorobenzaldehyde (Ar5, 89 ± 2 mmHg Po2), and pentadecafluoroctanoylchloride (PFOC) (Ali15, 134 ± 3 mmHg Po2) could be attached covalently to a UV-curable methacrylamide chitosan hydrogel (MACF) and used for tunable oxygen delivery to accelerate full thickness acute”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites Wijekoon et al., 2013

      Not checked in the free run.

    • cites Patil et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    At the center of cell seed gels, oxygen tensions were 124 mmHg for aromatic PFC Ar5, 121 mmHg for aliphatic short chain Ali5, and 130 mmHg for aliphatic long-chain Ali15.

    “At the center of cell seed gels, the following oxygen tensions were reported: aromatic PFC Ar5, 124 mmHg, aliphatic short chain Ali5, 121 mmHg and aliphatic long-chain Ali15, 130 mmHg.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Li et al., 2014

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    HEMOXCell reduces cell hypoxia while restoring functions in encapsulated islet cells.

    “HEMOXCell has shown the ability to reduce cell hypoxia while restoring functions in encapsulated islet cells in a [Rodriguez-Brotons et al., 2016].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Rodriguez-Brotons et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    At high O2 conditions (21%, 160 mmHg Po2) NSPCs tend to prefer neuronal differentiation, and at low O2 conditions (2%, 15 mmHg Po2) NSPCs differentiate into glia cells.

    “The role of oxygen tension in NSPC maintenance and fate decisions is supported by other studies including one that demonstrated that at high O2 conditions (21%, 160 mmHg Po2) NSPCs tend to prefer neuronal differentiation and at low O2 conditions (2%, 15 mmHg Po2) NSPCs differentiate into glia cells [Xie et al., 2014].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Xie et al., 2014

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    H2O2-releasing microspheres with bovine liver catalase improved stem cell survival and restored differentiation in cardiosphere-derived cell therapy after myocardial injection.

    “A biomaterial strategy with H2O2−releasing microspheres (maximum oxygen concentration of 18.4%, or 137 mmHg Po2) and bovine liver catalase was created to improve stem cell survival and shown to restore differentiation in the cardiosphere-derived cell (CDC) therapy after myocardial injection [Li et al., 2012].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Li et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    The oxygen releasing system improved cell survival for 7 days and preserved differentiation potential even in hypoxic conditions below 8 mmHg oxygen.

    “In this study, the oxygen releasing system improved cell survival for 7 days and preserved their differentiation potential even in hypoxic conditions (<8 mmHg oxygen environment).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Li et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    A PFC emulsion of perfluorotributylamine (FC-43) is biologically inert, non-toxic, and does not produce toxic metabolic products in rats.

    “In one such study, a PFC emulsion of perfluorotributylamine (FC-43) has shown to be biologically inert, non-toxic and does not produce toxic metabolic products in rats [Chubb, 1985].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Chubb, 1985

      Not checked in the free run.

  • May combine two claims.

    Interpretationcitation

    Covalent immobilization, encapsulation in biocompatible matrices, and coating emulsion droplets with biopolymers/lipids stabilize perfluorocarbons and can greatly reduce bioaccumulation and toxicity.

    “covalent immobilization, encapsulation in the biorthogonal/biocompatible matrices, coating emulsion droplets with biopolymers/lipids allow for the stabilization of perfluorocarbons and can drastically reduce bioaccumulation and toxicity.”
    Quote in the paperSubstantial claim

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Organoids may be useful for replacing organs, modeling disease, discovering drugs, and screening safety.

    “Organoids present promising potential for applications such as organ replacement, modeling of many diseases, drug discovery and safety screening studies”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Hu et al., 2018

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    After 7 days, Oxy-chip cultures reached 130 mmHg oxygen partial pressure, while control cultures fell to 70 mmHg.

    “They observed an oxygen partial pressure of 130 mmHg in their culture system after 7 days using their oxygen chips, whereas this number decreased to 70 mmHg in their non-Oxy chip controls.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Kamoya et al., 2016

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Premisecitation

    Myoglobin has a structure similar to hemoglobin and binds oxygen.

    “Myoglobin is a protein which has a similar structure to hemoglobin and thus binds oxygen to its structure”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Endeward et al., 2010

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Murphy et al.'s observations suggest that MSC spheroid function and hypoxic core formation do not depend on oxygen.

    “These observations indicate that the function of MSC spheroids and their hypoxic cores are non-oxygen-dependent.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Murphy et al. measured oxygen tension across spheroid diameters with oxygen-sensitive microelectrodes and linked the profiles to hypoxic cores.

    “Murphy et al. have utilized oxygen-sensitive microelectrodes to measure the oxygen tension as a function of spheroid diameter and correlated that with the existence of a hypoxic core”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Murphy et al., 2017

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Oxygenating cells directly bypasses diffusion-limited transport constraints and may make cartilage, bone, and cardiac engineering feasible.

    “This direct cell oxygenation avoids the size limitations imposed by diffusion limited mass transport and should enable cartilage, bone, and cardiac tissue engineering.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Without vessels, spheroids form HIFs and, when oxygen shortage lasts, cells in the poorly supplied center often die by apoptosis.

    “Lack of vasculature in spheroids, which triggers the formation of HIFs, often results in cell apoptosis in the central oxygen and nutrient-deficient regions in prolonged cases of oxygen deficiency”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Laschke and Menger, 2017; Lazzari et al., 2017

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Spheroids can model in vivo physiology and permit human cells to be used for studying human responses.

    “Spheroids are great candidates for mimicking these physiologic in vivo conditions towards forming complex organ-like structures and offer the ability to use human cells to better reproduce and study human responses”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Laschke and Menger, 2017

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    FOX molecules were immobilized to graphene oxide to form nano-platforms that act as oxygen reservoirs.

    “In another recent study, 3-pentadecafluoroheptyl,5-perfluorophenyl-1,2,4-oxadiazole (FOX) molecules were immobilized to graphene oxide to form nano-platforms [Maio et al., 2018]. These nano-platforms can act as oxygen reservoirs”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Maio et al., 2018

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Other groups have immobilized linear and branched PFCs to hyaluronic acid, chitosan, and alginate and shown benefits for in vitro culture of mammalian or human cells.

    “In similar approaches, other groups have immobilized linear as well as branched PFCs to hyaluronic acid (HA) [Palumbo et al., 2014], chitosan and alginate [Gattas-Asfura et al., 2012], and have shown benefits for in vitro culture of mammalian or human cells.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites Palumbo et al., 2014

      Not checked in the free run.

    • cites Gattas-Asfura et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Photosynthetic microalgae can be genetically modified to express VEGF, making them a versatile platform for photosynthetic biomaterials in tissue engineering.

    “In addition to oxygen generation, photosynthetic microalgae can be genetically modified to express the angiogenic recombinant protein VEGF (vascular endothelial growth factor) demonstrating its use as a versatile platform for photosynthetic biomaterials for tissue engineering.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Chavez et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In a fully immunocompetent mouse skin defect model, engrafted photosynthetic biomaterials proved safe and effective.

    “They demonstrated the safety and efficacy of photosynthetic biomaterials in vivo after engraftment in a fully immunocompetent mouse skin defect model.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Chavez et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Oxy-chip spheroids kept viable nucleated cores; control spheroids had central necrosis.

    “spheroids cultured on the Oxy chip showed viable nucleated cores, whereas cell necrosis was observed at the center of the non-Oxy chip controls.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Kamoya et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Algae-based photosynthetic biomaterials generated chimeric animal-plant tissues during dermal regeneration and supplied oxygen without needing blood perfusion.

    “photosynthetic biomaterials can produce and provide oxygen (saturation Po2 > 400 mmHg; 18.0 mg/L) independently of blood perfusion by generating chimeric animal-plant tissues during dermal regeneration”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Chavez et al., 2016

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    One PDMS-CaO2 disk produced a reported oxygen concentration of 0.16 ± 0.017 mM (~114 mmHg Po2) in week 1 and 0.073 ± 0.007 mM (~52 mmHg Po2) by week 4, relative to PDMS-only controls.

    “They showed that a single PDMS-CaO2 disk could improve the oxygen concentration in the culture solution from 0.16 ± 0.017 mM (~114 mmHg Po2) on the first week to an average of 0.073 ± 0.007 mM (~52 mmHg Po2) in 4 weeks of these values over baseline controls of PDMS only disks.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Pedraza et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Anada et al. made gas-permeable PDMS chips that oxygenated cells inside spheroids directly; after 5 days, media oxygen tension was 50-60 mmHg Po2 and viability/function were preserved.

    “Anada et al., have utilized 3D culture chips made of gas-permeable polydimethylsiloxane (PDMS) to enable direct oxygen supply to the cells within their spheroids while maintaining the cell viability and function with an equilibrium oxygen tension of 50–60 mmHg Po2 in the culture media after 5 days”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Anada et al., 2012

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Even with small radial oxygen differences, larger spheroids had lower cell metabolism, perhaps from altered matrix deposition and packing.

    “In spite of low radial oxygen fluctuations, the cell metabolism decreased with increasing spheroid size, possibly due to the adaptive changes in matrix deposition and packing density of the spheroids.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Murphy et al., 2017

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Powers et al. reported that spheroids were maintained long-term and formed tissue structures.

    “The results indicated long-term maintenance of the spheroids and the corresponding formation of tissue structures”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Powers et al., 2002

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Larger tissue constructs still need better oxygen diffusion, particularly since organoids cannot form their own blood vessels.

    “developing larger tissue structures is still in need of enhanced oxygen diffusion, especially because organoids cannot grow their own vasculature”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    They made an endothelialized myocardium organoid with controlled anisotropy, suggesting better oxygen movement through the scaffold.

    “they successfully produced an endothelialized myocardium organoid with controlled anisotropy, which indicates enhanced oxygen transport through the scaffold.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zhang et al., 2016

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Interpretationreasoning

    Oxygen-delivery methods remain immature; design rules specific to oxygen in tissue engineering and organoids are still being developed as technologies evolve.

    “These approaches are still in their infancy as oxygen-specific design criteria for tissue engineering and organoid systems are still being formulated while new technologies are constantly emerging and evolving to meet these requirements.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Organoid methods still struggle to manage multiple cell types, supply each type with suitable ECM, and meet each type's oxygen and nutrient needs within one organoid.

    “current organoid technologies face significant challenges, including managing the various cell types in each specific organoid system, while providing the appropriate ECM for each cell type as well as improving the oxygen and nutrient requirements for each particular cell type within the same organoid”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Lou and Leung, 2018

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Poor oxygen diffusion creates a central necrotic core in organoids and blocks normal structural development.

    “oxygen diffusion issues lead to the formation of a necrotic core at the center of the organoid, preventing the normal development of the structure”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Kelava and Lancaster, 2016

      Not checked in the free run.

  • Classified as probably not a substantial claim. May combine two claims.

    Interpretationreasoning

    When growth reaches that limit, adding oxygen may help meet demand, permit more growth, and improve final function.

    “At such a tipping point, a supplemental oxygenating system might be beneficial to support the oxygen requirements of the system to allow further growth and enhance ultimate functions.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Even without vessels, multi-well spinning bioreactors generated forebrain, midbrain, and hypothalamus organoids across 14-84 days, suggesting good oxygen and nutrient movement.

    “Despite the lack of vasculature, these multi-well spinning bioreactors have been able to successfully produce fore-brain, midbrain and hypothalamus organoids over a period of 14 to 84 days, indicating the efficient oxygen and nutrient transport regime.”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Qian et al., 2018

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    In DiStefano et al.'s system, cells proliferated more and differentiated clearly into neurons and S-cone photoreceptors.

    “They showed enhanced the proliferation as well as the well-defined differentiation of these cells into neurons and S-cone photoreceptors.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites DiStefano et al., 2018

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S2 (checkable). May combine two claims.

    Interpretationreasoning

    The measurement gap matters at every embryonic stage and might yield design criteria for adult tissue needs.

    “This is especially important in each stage of embryonic development, which could translate to direct design criteria to satisfy specific requirements of adult tissues to form appropriate tissue engineered structures.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    A 300 μm channel width gave perfusion of 8.2×10-8 mol/cm3/s, which matched the expected hepatocyte oxygen consumption of 3.5×10-8 mol/cm3/s.

    “Bioreactor dimensions, with a channel width of 300 μm, were designed such that achieved perfusion rates of 8.2×10–8 mol/cm3/s met the expected hepatocyte oxygen consumption rates (3.5×10–8 mol/cm3/s).”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Powers et al., 2002

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Changing oxygen availability influences development from early blastocyst stages through morphogenesis and organogenesis, and stays relevant in angiogenesis and adult life.

    “variations in oxygen bioavailability affect embryonic development starting from the very early stages of blastocyst formation to morphogenesis and organogenesis and continue to be important during angiogenesis and into adulthood.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Qian et al. built a spinning bioreactor and grew forebrain organoids from human iPSCs.

    “Qian et al. have developed a spinning bioreactor and successfully cultured forebrain-specific organoids originated from human induced pluripotent stem cells (iPSCs)”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites Qian et al., 2016

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S4 (based). May combine two claims.

    Interpretationnone given

    Oxygen signals proliferation, differentiation, angiogenesis, morphogenesis, and organogenesis, and also supplies energy through anaerobic glycolysis.

    “Oxygen contributes to development by signaling proliferation, differentiation, angiogenesis, morphogenesis, and organogenesis processes as well as providing required energy via anaerobic glycolysis.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationreasoning

    Without vessels, engineered constructs and organoids/spheroids cannot fully reproduce normal development in vitro, and new tissue growth eventually stops.

    “in a tissue-engineered construct or organoid/spheroid, the lack of a vasculature component makes it difficult to mimic the normal developmental process in vitro entirely, and thus inhibits the growth of any newly formed tissue after a certain point.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Microfluidics and bioprinting are exploring artificial vessels in scaffolds to lessen oxygen and nutrient shortages in organoid cultures and permit larger constructs.

    “microfluidics and bioprinting approaches are currently studying how artificial vessels can be added to tissue engineering scaffolds to reduce oxygen/nutrient deficiencies in organoid cultures, thus allowing for the formation of larger tissue constructs”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites Zhang et al., 2017

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationreasoning

    More research on engineered oxygen carriers could yield useful tools for solving oxygen shortage problems.

    “expanding research on engineered oxygen carriers, present valuable new tools to overcome the challenges caused by inadequate oxygen supply.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S2 (checkable).

    Interpretationreasoning

    Knowing the in vivo niche and oxygen levels at each fetal stage can guide tissue-engineering conditions that need precise control of proliferation and differentiation.

    “understanding the specific in vivo microenvironment, as well as the levels of oxygen present at each stage during normal fetal development, helps instruct the environments needed for tissue engineering approaches where fine-tuning of cell proliferation and differentiation are essential.”
    Quote in the paperNot a substantial claim
114 of 114

  • The paper relies on this without support.

    The oxygen levels and gradients observed in embryonic development can serve as design targets for tissue engineering and organoid strategies.

    “which also should be ultimately achieved in any tissue engineering or organoid strategy.”

  • The paper relies on this without support.

    The reported oxygen ranges for embryonic stages are representative enough to guide tissue engineering despite difficulties in measuring exact tissue oxygen levels.

    “Due to the difficulties of measuring the exact tissue oxygen levels experimentally, the relative changes in oxygen levels at the different stages are not fully known [Hemker et al., 2016].”

  • The paper relies on this without support.

    Lessons from native tissue oxygen microenvironments can be translated into synthetic tissue engineering scaffolds and organoids.

    “Taking lessons from native tissue microenvironments, researchers have explored approaches to control oxygen tensions, such as hemoglobin-based, perfluorocarbon-based and oxygen generating biomaterials, within synthetic tissue engineering scaffolds and organoids”

  • The paper relies on this without support.

    A lack of sustained and prolonged oxygen supply through oxygenating strategies can cause primary limitations in tissue engineered constructs to create large, self-sustaining structures.

    “Unless adequately addressed, the lack of sustained and prolonged oxygen supply via oxygenating strategies can cause primary limitations in tissue engineered constructs to create large, self-sustaining structures.”

  • The paper relies on this without support.

    Perfluorocarbons can readily dissolve oxygen in aqueous conditions.

    “Perfluorocarbons (PFCs) can readily dissolve oxygen in aqueous conditions.”

  • The paper relies on this without support.

    Non-immobilized free PFCs that are highly lipophilic tend to exhibit toxic responses to cells and inhibit growth.

    “A few reports show that non-immobilized free PFCs which are highly lipophilic tend to exhibit toxic responses to cells and inhibit growth.”
6 of 6

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • During embryogenesis, physiological oxygen concentration levels are significantly lower, primarily driving cellular proliferation.

    “During embryogenesis, physiological oxygen concentration levels are significantly lower, primarily driving cellular proliferation.”

    From Retinal organoid chip: engineering a physiomimetic oxygen gradient for optimizing long term culture of human retinal organoids (2025) · cites this paper as [35]

    In “Roles of oxygen in early embryonic development,” the paper says the embryo experiences physiological hypoxia at very early stages and that low oxygen is required for embryonic stem cells to initiate proliferation. The claim extends this early-development finding to embryogenesis generally; the “Oxygen as a developmental morphogen” section also describes oxygen as guiding differentiation, not only proliferation. (LLM, conf=0.84)

    Experimental — accuracy not yet measured

  • iPSC-derived organoids may benefit from spatial and temporal presentation of different oxygen tensions to mimic human embryogenesis.

    “This suggests that iPSC-derived organoids may benefit form spatial and temporal presentation of different oxygen tensions to mimic human embryogenesis.35”

    From Retinal organoid chip: engineering a physiomimetic oxygen gradient for optimizing long term culture of human retinal organoids (2025) · cites this paper as [35]

Self-check

Percentages, sums and reported differences re-computed from the paper’s own numbers.

Each contradiction between two passages listed here was confirmed by an independent check.

  • Oxygen-generating approaches are described as both alternatives to and examples of chemical generation

    Minor

    The section introduces oxygen-generating approaches as an alternative to chemical or enzymatic oxygen generation, but then describes chemical oxygen generation as one of those approaches.

1 of 1

The citation lists haven’t been fetched yet

References 185 · Cited by 101

Research works printed but not linked: 2

  1. Park J, Chang J.h., Choi M, Pak JJ, Lee DY, Pak YK (2007) Microfabirated Clark-type Sensor for Measuing Dissolved Oxygen: 2007 IEEE Sensors, pp 1412– 1415.
  2. Wang Y, Zhao S (2010) Vascular Biology of the Placenta: Cell Types of the Placenta. Chapter 4, San Rafael CA, Morgan & Claypool Life Sciences. 21452443

Authors
Fathollahipour, S., Patil, P. S., & Leipzig, N. D.
Affiliations
University of Akron (US)
Corresponding author
Leipzig, Nic D.
ORCID
Venue
Cells Tissues Organs
Publisher
S. Karger AG
Published
2018
Volume
205
Issue
5-6
Pages
350-371
ISSN
  • 1422-6405 (ISSN-L)
  • 1422-6421
DOI
10.1159/000493162
PMC
PMC6397050
OpenAlex
W2895698434
Semantic Scholar
11026fd0bb9870081197d8d159a80b7d3da1fe7f
Cited by
101
References
185
Citations by year
2026: 8 · 2025: 11 · 2024: 16 · 2023: 20 · 2022: 15 · 2021: 17 · 2020: 10 · 2019: 3 · 2016: 1
FWCI
7.79
Type
journal article
Language
en
Open access
Licence
Locations
  • Cells Tissues Organs (publishedVersion)
  • PubMed (publishedVersion)
  • Europe PMC (PubMed Central) (acceptedVersion)
  • PubMed Central (acceptedVersion)
Funders
National Institute of Diabetes and Digestive and Kidney Diseases (R41DK105704)
Topics
  • Health Sciences › Medicine › Surgery › Tissue Engineering and Regenerative Medicine
  • Physical Sciences › Engineering › Biomedical Engineering › 3D Printing in Biomedical Research
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Cancer Research › Cancer, Hypoxia, and Metabolism
Keywords
oxygen regulation, embryonic development, hypoxia-inducible factor, oxygen homeostasis, tissue engineering scaffolds, organoids, hemoglobin-based oxygen carriers, perfluorocarbon oxygen carriers, organogenesis, morphogenesis, stem cell differentiation, nutrient supply
MeSH terms
Animals, Cell Differentiation, Humans, Morphogenesis, Oxygen, Cell Hypoxia, Tissue Engineering, Organogenesis, Embryonic Development, Embryonic Stem Cells

About these checks

Registry, retraction, Cites and statistics results are computed, not judged, with one exception: a printed reference that no index lists is read once by a model before it counts as a citation error. Claims are extracted by a model and kept only when their quote appears verbatim in the text; each cited source is then checked, with a model’s help, for whether it says what the claim relies on. Those verdicts can be wrong — read the claim’s quote and the cited source before relying on one.

Claims read by deepseek-v4.1-flash under definition version 3 (extract-v3) on 2026-10-07.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-07.

Statistics and arithmetic are recomputed, not judged.

Confidence bands are shown only for fields calibrated on our annotated test set. None is calibrated yet, so no band is shown.

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