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Cited by 6153D Printing in Biomedical ResearchTissue Engineering and Regenerative MedicineDrug-Induced Hepatotoxicity and Protection

A liver-on-a-chip platform with bioprinted hepatic spheroids

Open access
Bhise, Nupura S, Manoharan, Vijayan, Massa, Solange, Tamayol, Ali, Ghaderi, Masoumeh, Miscuglio, Mario, Lang, Qi, Shrike Zhang, Yu, Shin, Su Ryon, Calzone, Giovanni, Annabi, Nasim, Shupe, Thomas D, Bishop, Colin E, Atala, Anthony, Dokmeci, Mehmet R, Khademhosseini, Ali·2016·Biofabrication
DOI 10.1088/1758-5090/8/1/014101
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Abstract

The inadequacy of animal models in correctly predicting drug and biothreat agent toxicity in humans has resulted in a pressing need for in vitro models that can recreate the in vivo scenario. One of the most important organs in the assessment of drug toxicity is liver. Here, we report the development of a liver-on-a-chip platform for long-term culture of three-dimensional (3D) human HepG2/C3A spheroids for drug toxicity assessment. The bioreactor design allowed for in situ monitoring of the culture environment by enabling direct access to the hepatic construct during the experiment without compromising the platform operation. The engineered bioreactor could be interfaced with a bioprinter to fabricate 3D hepatic constructs of spheroids encapsulated within photocrosslinkable gelatin methacryloyl (GelMA) hydrogel. The engineered hepatic construct remained functional during the 30 days culture period as assessed by monitoring the secretion rates of albumin, alpha-1 antitrypsin, transferrin, and ceruloplasmin, as well as immunostaining for the hepatocyte markers, cytokeratin 18, MRP2 bile canalicular protein and tight junction protein ZO-1. Treatment with 15 mM acetaminophen induced a toxic response in the hepatic construct that was similar to published studies on animal and other in vitro models, thus providing a proof-of-concept demonstration of the utility of this liver-on-a-chip platform for toxicity assessment.

Integrity signals

Registry

Crossref · 10.1088/1758-5090/8/1/014101

Cites

80 of 80 references verified0 citation errors · 0 not checkable

Statistics

Nothing to recompute.

Claims the paper makes

16 of 40 cited sources checked · 5 supported · 0 flagged · accuracy not yet measured

  • Premisecitation

    Conventional 2D hepatic mono- and co-cultures of normal and diseased human cells fail to mimic the complex 3D in vivo microenvironment and extracellular matrix support.

    “However, hepatic models that utilize conventional 2D mono-and co-cultures of normal and diseased human cells fail to mimic the complex threedimensional (3D) in vivo microenvironment and extracellular matrix (ECM) support [16,19,20].”
    Quote in the paperSubstantial claim1 of 3 sources checked
    • cites [16] Khetani, 2008, Microscale culture of human liver cells for drug development, Nat. Biotechnol. source
    • cites [19] Bhatia, 1999, Effect of cell–cell interactions in preservation of cellular phenotype: cocultivation of hepatocytes and nonparenchymal cells, FASEB J. source
    • cites [20] LeCluyse, 2012, Organotypic liver culture models: meeting current challenges in toxicity testing, Crit. Rev. Toxicol. source

      Section 3.3 says traditional 2D monocultures and co-cultures do not typically represent the liver’s three-dimensional cellular configuration or in vivo microenvironments. But Section 2.1 says co-cultures satisfy nearly all biophysical requirements for hepatocytes, including cell–matrix contacts, so the blanket claim that they lack ECM support overstates the source. The cited passages also do not establish the claim specifically for diseased human cells; page numbers were unavailable. (LLM, conf=0.83)

      Experimental — accuracy not yet measured
  • Interpretationcitation

    Increased albumin secretion from spheroids compared to cell monolayers is consistent with observations reported in literature.

    “Increased albumin secretion from spheroids compared to cell monolayers, as seen here, is consistent with observations reported in literature, which indicates that culturing the cells as spheroids may enhance homotypic cell-cell interactions within aggregated hepatocytes and improve functional outcomes of the hepatic construct [25,48].”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [25] Li, 2014, Micropatterned cell–cell interactions enable functional encapsulation of primary hepatocytes in hydrogel microtissues, Tissue Eng. A source

      The “Effect of homotypic contacts on 3D albumin secretion” section reports more than threefold greater albumin secretion from hydrogels containing aggregated hepatocyte pucks than from hydrogels containing unpatterned hepatocytes at day 8. This supports the proposed benefit of aggregation, but the measured comparison was between two 3D hydrogel conditions, not spheroids and cell monolayers. (LLM, conf=0.86)

      Experimental — accuracy not yet measured
    • cites [48] Fraczek, 2013, Primary hepatocyte cultures for pharmaco-toxicological studies: at the busy crossroad of various anti-dedifferentiation strategies, Arch. Toxicol. source
  • Premisecitation

    These cell lines offer a reliable alternative to primary cells by overcoming the above issues, especially for use in proof-of-concept studies in dynamic flow liver-on-a-chip platforms.

    “These cell lines offer a reliable alternative to primary cells by overcoming the above issues, especially for use in proof-of-concept studies in dynamic flow liver-on-achip platforms [20].”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [20] LeCluyse, 2012, Organotypic liver culture models: meeting current challenges in toxicity testing, Crit. Rev. Toxicol. source
    • Rests on an assumption: Primary hepatocytes have limited lifespan, difficulty in obtaining the cell source, and large batch-to-batch variability post-isolation.
  • Interpretationcitation

    By adapting this method, the authors avoided potentially harsh printing conditions including low temperature for a prolonged printing period and undesirable shear stress along the length of the nozzle.

    “By adapting this method, we avoided potentially harsh printing conditions including, low temperature for a prolonged printing period [51] and undesirable shear stress along the length of the nozzle [30].”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [51] Billiet, 2014, The 3D printing of gelatin methacrylamide cell-laden tissue-engineered constructs with high cell viability, Biomaterials source
    • cites [30] Bertassoni, 2014, Direct-write bioprinting of cell-laden methacrylated gelatin hydrogels, Biofabrication source

      Section 3.3 reports that cell viability was not significantly affected and suggests the hydrogel matrix may protect cells from shear stress during dispensing. It also says dispensing required significant stress, so the source supports protection from its effects, not avoidance of shear stress along the nozzle. The low-temperature condition is attached to a different citation in the citing sentence. (LLM, conf=0.84)

      Experimental — accuracy not yet measured
  • Premisecitation

    The oxygen consumption of HepG2/C3A cells can be described with Michaelis-Menten kinetics.

    “The oxygen consumption of HepG2/C3A cells can be described with the Michaelis-Menten kinetics [41,42]:”
    Quote in the paperNot a substantial claim1 of 2 sources checked
    • cites [41] Weise, 2013, Analysis and comparison of oxygen consumption of HepG2 cells in a monolayer and three-dimensional high density cell culture by use of a matrigrid®, Biotechnol. Bioeng. source
    • cites [42] Buchwald, 2009, FEM-based oxygen consumption and cell viability models for avascular pancreatic islets, Theor. Biol. Med. Modelling source

      The Methods section assumes a Michaelis–Menten-type rate for oxygen consumption, and the Discussion’s “Oxygen consumption model” section specifies that this assumption applies to pancreatic islet tissue. The cited paper does not establish that the same kinetics describe HepG2/C3A cells; no page numbers are available in our copy. (LLM, conf=0.88)

      Experimental — accuracy not yet measured
  • May combine two claims. 17
  • Classified as probably not a substantial claim. 14
  • The quote only partly states this claim. 1

  • May combine two claims.

    Premisecitation

    iPSC-derived hepatocyte-like cells are still in nascent stages of development and show characteristics similar to fetal hepatocytes.

    “These iPSC-derived hepatocyte-like cells are still in nascent stages of development and show characteristics similar to fetal hepatocytes [76].”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [76] Bhatia, 2014, Cell and tissue engineering for liver disease, Sci. Translational Med. source

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

    Premisecitation

    Patient-specific hepatocytes derived from autologous iPSCs have recently emerged as an attractive alternative that can overcome issues with primary hepatocytes.

    “Patient-specific hepatocytes derived from autologous iPSCs have recently emerged as an attractive alternative that can overcome these issues [60].”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [60] Shan, 2013, Identification of small molecules for human hepatocyte expansion and iPS differentiation, Nat. Chem. Biol. source

      The paper describes stem cells as an attractive alternative to primary hepatocytes and says iPS cells offer the possibility of patient-specific hepatocyte-like cells (opening passage, page unavailable). It does not establish that these cells overcome the issues with primary hepatocytes: the same passage says their immature phenotype and limited function restrict their use as a renewable source of functional hepatocytes. (LLM, conf=0.84)

      Experimental — accuracy not yet measured

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

    Premisecitation

    The metabolic activity and function of cells fluctuate over time, and similar observations have been reported in the literature.

    “The metabolic activity and function of cells fluctuate over time and similar observations have been reported in the literature [61,62].”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [61] Dash, 2013, Hemodynamic flow improves rat hepatocyte morphology, function, and metabolic activity in vitro, Am. J. Physiol. Cell Physiol. source

      The Abstract reports that hepatocyte-specific function and metabolic enzyme activity decline over days in nonflow cultures. The Discussion (Fig. 7A) likewise describes a decline in enzyme expression and activity over 3–7 days, but says the controlled-hemodynamics system maintains stability. The source supports time-dependent changes in cultured hepatocytes, not a general claim that cells’ activity and function fluctuate over time. (LLM, conf=0.85)

      Experimental — accuracy not yet measured
    • cites [62] Wagner, 2013, A dynamic multi-organ-chip for long-term cultivation and substance testing proven by 3D human liver and skin tissue co-culture, Lab Chip source

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

    Premisecitation

    APAP overdose is a common cause of acute liver failure in the US.

    “APAP overdose is a common cause of acute liver failure in the US [71,72].”
    Quote in the paperNot a substantial claim2 of 2 sources checked
    • cites [71] Blazer, 2009, Nonprescription use of pain relievers by middle-aged and elderly community-living adults: national survey on drug use and health, J. Am. Geriatrics Soc. source

      The Abstract Results report that acetaminophen-containing pain relievers were commonly used without a prescription in the surveyed group, but do not discuss overdose or liver failure. Searches of the full-text copy for acetaminophen overdose and liver failure found no passage addressing whether overdose is a common cause of acute liver failure in the US. (LLM, conf=0.76)

      Experimental — accuracy not yet measured
    • cites [72] Davidson, 1966, Acute liver necrosis following overdose of paracetamol, Br. Med. J. source

      The Summary (page unavailable) describes two fatal cases of acute liver necrosis after paracetamol overdose and suggests paracetamol was the chief cause. The paper does not establish that overdose is a common cause of acute liver failure in the US; it provides no US frequency data. (LLM, conf=0.88)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Premisecitation

    APAP is a well-characterized hepatotoxic drug and its in vivo model data is available in literature.

    “APAP is a wellcharacterized hepatotoxic drug and its in vivo model data is available in literature [73].”
    Quote in the paperSubstantial claim1 of 1 sources checked
    • cites [73] Fey, 2012, Determination of drug toxicity using 3D spheroids constructed from an immortal human hepatocyte cell line, Toxicol. Sci. source

  • May combine two claims.

    Interpretationcitation

    Culturing cells as spheroids may enhance homotypic cell-cell interactions within aggregated hepatocytes and improve functional outcomes of the hepatic construct.

    “Increased albumin secretion from spheroids compared to cell monolayers, as seen here, is consistent with observations reported in literature, which indicates that culturing the cells as spheroids may enhance homotypic cell-cell interactions within aggregated hepatocytes and improve functional outcomes of the hepatic construct [25,48].”
    Quote in the paperSubstantial claim1 of 2 sources checked
    • cites [25] Li, 2014, Micropatterned cell–cell interactions enable functional encapsulation of primary hepatocytes in hydrogel microtissues, Tissue Eng. A source
    • cites [48] Fraczek, 2013, Primary hepatocyte cultures for pharmaco-toxicological studies: at the busy crossroad of various anti-dedifferentiation strategies, Arch. Toxicol. source

  • May combine two claims.

    Premisecitation

    HepG2 cells can cluster over time, but dispersed cells take longer to form cell-cell interactions when encapsulated in a porous scaffold compared to spheroids, which could affect their resistance to drugs.

    “HepG2 cells can cluster over time, but dispersed cells take longer to form these cell-cell interactions when encapsulated in a porous scaffold compared to spheroids, which could affect their resistance to drugs [49].”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [49] Ho, 2010, Incorporation of multicellular spheroids into 3D polymeric scaffolds provides an improved tumor model for screening anticancer drugs, Cancer Sci. source

      The section ‘Spheroids remain intact in scaffolds after seeding’ (page unavailable) compares dispersed cells with spheroids in porous scaffolds and reports greater drug resistance for the spheroid-seeded scaffolds. However, the experiments used U251 glioma cells, not HepG2 cells, and the copy does not establish that dispersed cells take longer than spheroids to form cell-cell interactions in scaffolds. (LLM, conf=0.86)

      Experimental — accuracy not yet measured

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

    Premisecitation

    The observed cell density range is typical for hydrogel encapsulation.

    “which is a typical cell density range for hydrogel encapsulation [7]”
    Quote in the paperNot a substantial claim1 of 1 sources checked
    • cites [7] Akbari, 2014, Composite living fibers for creating tissue constructs using textile techniques, Adv. Funct. Mater. source

      The Experimental Section (Preparation of Cell Suspension in Sodium Alginate; page unavailable) reports using 2–5 × 10^6 cells mL−1 in an alginate suspension for cell-laden hydrogel fibers. This supports the range as an example of hydrogel encapsulation, but the paper does not establish that it is typical across hydrogel encapsulation generally. (LLM, conf=0.82)

      Experimental — accuracy not yet measured

  • May combine two claims.

    Findingown data

    The thickness of the bioprinted GelMA dot is approximately 800 μm, which is thicker than the average diameter of the spheroids (approximately 200 μm).

    “As seen in the side view micrograph of the bioprinted hydrogel with spheroids in figure S2, the thickness of the bioprinted GelMA dot is ∼800 μm, which is thicker than the average diameter of the spheroids (∼200 μm).”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Findingreasoning

    The bioreactor could be unscrewed for disassembly at any time for direct access to the hepatic construct and then resealed, unlike the majority of PDMS-based on-chip systems that are permanently plasma bonded.

    “Unlike the majority of PDMSbased on-chip systems, which are permanently plasma bonded to seal the compartments, the bioreactor could be unscrewed for disassembly at any time for allowing direct access to the hepatic construct and then resealed.”
    Quote in the paperNot a substantial claim

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

    Findingown data

    The dimensions of the bioprinted hydrogel features were much smaller than the 8 mm depth of the chamber.

    “Moreover, the dimensions of the bioprinted hydrogel features were much smaller than the 8 mm depth of the chamber.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    A spheroid size of 200 μm is suitable to avoid necrosis in the core while maintaining the functionality of the cells within the spheroids.

    “Literature suggests that a size of 200 μm is suitable to avoid necrosis in the core, while maintaining the functionality of the cells within the spheroids [46,47].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [46] Fukuda, 2003, Efficacy of a polyurethane foam/spheroid artificial liver by using human hepatoblastoma cell line (Hep G2), Cell Transplant source
    • cites [47] Sakai, 2007, Technique for the control of spheroid diameter using microfabricated chips, Acta. Biomater. source

  • May combine two claims.

    Premisecitation

    The current drug development process suffers from a high failure rate during clinical trials, partly due to inadequacy of animal models to accurately predict toxic side-effects of drugs in humans.

    “The current drug development process suffers from a high failure rate during clinical trials, which is partly due to inadequacy of animal models to accurately predict the toxic side-effects of drugs in humans [1,2].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [1] van Midwoud, 2011, Microfluidic devices for in vitro studies on liver drug metabolism and toxicity, Integr. Biol. (Camb.) source
    • cites [2] Kaplowitz, 2005, Idiosyncratic drug hepatotoxicity, Nat. Rev. Drug Discovery source

  • May combine two claims.

    Premisecitation

    Ex vivo studies using 3D tissue biopsies or slices may be difficult for high-throughput studies and show rapid loss of functionality within a few days in vitro, limiting their application for long-term studies.

    “Ex vivo studies using 3D tissue biopsies or slices have been reported, but they may be difficult to be used for high-throughput studies and show rapid loss of functionality within a few days in vitro, thus limiting their application for long-term studies [21].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [21] van Midwoud, 2010, A microfluidic approach for in vitro assessment of interorgan interactions in drug metabolism using intestinal and liver slices, Lab Chip source

  • Classified as probably not a substantial claim.

    Premisecitation

    The metabolic functions of the liver are highly dependent on its microstructural architecture, fluid flow patterns, zonal ECM composition, and biochemical (oxygen concentration) gradients.

    “The metabolic functions of the liver are highly dependent on its microstructural architecture, fluid flow patterns, zonal ECM composition, and biochemical (oxygen concentration) gradients [74].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [74] Allen, 2003, Formation of steady-state oxygen gradients in vitro: application to liver zonation, Biotechnol. Bioeng. source

  • May combine two claims.

    Interpretationown data

    The acute hepatotoxic dose of APAP causes apoptosis of HepG2 cells within GelMA-encapsulated spheroids, leading to decreased biomarker secretion in the bioreactor system over time.

    “The acute hepatotoxic dose of APAP causes apoptosis of HepG2 cells within GelMA encapsulated spheroids, leading to a decrease in the biomarkers secretion levels in the bioreactor system over time”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    Although relative albumin secreted per cell decreased over time, the level was sustained within the range (0.6-80 pg/cell/day) reported in the literature for albumin production per cell by HepG2 cells/spheroids.

    “This shows that although the relative albumin secreted per cell decreased overtime, the level was sustained within the range (0.6-80 pg/cell/day) reported in the literature for albumin production per cell by HepG2 cells/ spheroids [63,64].”
    Quote in the paperSubstantial claim0 of 2 sources checked
    • cites [63] Torisawa, 2007, A multicellular spheroid array to realize spheroid formation, culture, and viability assay on a chip, Biomaterials source
    • cites [64] Kim, 2007, A microfluidic platform for three-dimensional cell culture and cell-based assays, Biomed. Microdevices. source

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

    Premisecitation

    Supportive interactions with non-parenchymal cells, particularly vascular, stromal and immune cell types, are important for maintaining the high proliferative capacity of hepatocytes in 3D cultures.

    “Moreover, supportive interactions with non-parenchymal cells, particularly the vascular, stromal and immune cell types, are important for maintaining the high proliferative capacity of hepatocytes in 3D cultures [75].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [75] Kostadinova, 2013, A long-term three dimensional liver co-culture system for improved prediction of clinically relevant drug-induced hepatotoxicity, Toxicology Appl. Pharmacology source

  • The quote only partly states this claim.

    Interpretationown data

    The levels of transferrin secreted by HepG2 cells were comparable to the literature range (0.1-17.2 pg/cell/day).

    “transferrin (0.1-17.2 pg/cell/day) [65,67,68]”
    Quote in the paperSubstantial claim0 of 3 sources checked
    • cites [65] Bouma, 1989, Further cellular investigation of the human hepatoblastoma-derived cell line HepG2: morphology and immunocytochemical studies of hepatic-secreted proteins., In Vitro Cell. Developmental Biol. : J. Tissue Cult. Assoc. source
    • cites [67] Carraro, 2008, In vitro analysis of a hepatic device with intrinsic microvascular-based channels, Biomed. Microdevices source
    • cites [68] Iwasa, 1990, Phenotypic variation in human HepG2 hepatoma cells: alterations in cell growth, plasma protein synthesis and heme pathway enzymes, Int. J. Biochem. source

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

    Interpretationreasoning

    The proposed concept of a bioreactor interfaced with bioprinters expands the field of organ-on-a-chip and may be a key step towards fabrication of automated systems for high throughput drug screening.

    “The proposed concept of a bioreactor interfaced with bioprinters expands the field of organ-on-a-chip and may be a key step towards the fabrication of automated systems for high throughput drug screening.”
    Quote in the paperNot a substantial claim

  • Probably not a substantial claim: may not pass S4 (based).

    Study design and conductnone given

    The multilayer design was selected so that any introduced bubble would be washed away without getting trapped within the chambers.

    “The multilayer design was selected in a way that any introduced bubble would be washed away without getting trapped within the chambers.”
    Quote in the paperNot a substantial claim

  • Classified as probably not a substantial claim.

    Interpretationreasoning

    The successful interfacing of a bioprinting platform with the bioreactor might allow for creation of advanced biomimetic architectures by mimicking distribution of ECM-like biomaterials, hepatic cell types, and their ratios.

    “The successful interfacing of a bioprinting platform with the bioreactor, as reported here, might allow for the creation of advanced biomimetic architectures by precisely mimicking the distribution of different ECM-like biomaterials, key hepatic tissue cell types including hepatocytes, stellate, endothelial, and Kupffer cells, and their ratios.”
    Quote in the paperSubstantial claim

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

    Study design and conductnone given

    The same encapsulation conditions were used for control and experimental groups, so that these conditions did not affect the results.

    “Although cell secreted proteins interact with the porous hydrogel scaffold, we used the same encapsulation conditions for control and experimental groups, so that these conditions did not affect the results.”
    Quote in the paperNot a substantial claim

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

    Interpretationcitation

    The levels of ceruloplasmin secreted by HepG2 cells were comparable to the literature range (0.1-0.7 pg/cell/day).

    “ceruloplasmin (0.1-0.7 pg/cell/day) [68] secreted by HepG2 cells.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [68] Iwasa, 1990, Phenotypic variation in human HepG2 hepatoma cells: alterations in cell growth, plasma protein synthesis and heme pathway enzymes, Int. J. Biochem. source

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

    Premisecitation

    Freshly isolated or cryopreserved primary hepatocytes are the preferred cell source for drug metabolism studies.

    “Freshly isolated or cryopreserved primary hepatocytes are the preferred cell source for drug metabolism studies [54][55][56].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [54][55][56]

      Couldn’t be matched to a paper in Crossref, so it can’t be checked.

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

    Study design and conductnone given

    There were no issues encountered while disassembling and reassembling the platform during the APAP assessment.

    “There were no issues encountered while carefully disassembling and reassembling the platform during this assessment.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Interpretationnone given

    The bioprinting approach enabled easy tailoring of other printing parameters such as spheroid concentration per dot and hydrogel composition, as well as controlling multicellular distribution and the geometrical features of the printed construct.

    “Also, this bioprinting approach enabled easy tailoring of other printing parameters such as spheroid concentration per dot and hydrogel composition, as well as controlling multicellular distribution and the geometrical features of the printed construct.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationown data

    The response of the liver-on-a-chip platform to APAP treatment was similar to animal and in vitro models, which confirms the possibility of its application for drug toxicity analysis.

    “The response of the liver-on-achip platform to APAP treatment was similar to animal and in vitro models, which confirms the possibility of its application for drug toxicity analysis.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Interpretationnone given

    The use of bioprinting in the system paved the way for rapid fabrication of the 3D hepatic construct and allowed precise control over the thickness of the hydrogel layer encapsulating the spheroids to maintain consistency in mass diffusion through the hydrogel for all experiments.

    “The use of bioprinting in our system not only paved the way for a rapid fabrication of the 3D hepatic construct, but also allowed for a precise control over the thickness of the hydrogel layer encapsulating the spheroids to maintain consistency in mass diffusion through the hydrogel, for all experiments.”
    Quote in the paperSubstantial claim
29 of 29

  • The authors state this as an assumption.

    The hydrogels can be modeled as a porous medium with a uniform volumetric oxygen consumption rate associated with the total number of encapsulated cells.

    “The hydrogels were modeled as a porous media with uniform volumetric oxygen consumption rate associated with the total number of encapsulated cells.”

  • The authors state this as an assumption.

    The oxygen concentration at the inlet is constant and uniform.

    “The oxygen concentration at the inlet was considered to be constant and uniform.”

  • The authors state this as an assumption.

    All boundaries of the bioreactor can be treated as impermeable surfaces for oxygen transport.

    “Although PDMS is oxygen permeable [40], since the bioreactor is covered by a impermeable PMMA layer, we considered the worst case scenario and all boundaries were treated as impermeable surfaces.”

  • The paper relies on this without support.

    Primary hepatocytes have limited lifespan, difficulty in obtaining the cell source, and large batch-to-batch variability post-isolation.

    “In addition to their limited lifespan, difficulty in obtaining the cell source and large batch-to-batch variability post-isolation are other important challenges associated with the use of primary hepatocytes.”

  • The paper relies on this without support.

    Immortalized cell lines offer a cheap and robust platform for initial toxicity evaluation of an array of drug candidates.

    “The use of immortalized cell lines offers a cheap and robust platform for initial toxicity evaluation of an array of drug candidates.”

  • The paper relies on this without support.

    Issues with cost, availability and batch-to-batch variability make primary hepatocytes a challenging cell source to use.

    “However, issues with cost, availability and batch-to-batch variability make primary hepatocytes a challenging cell source to use.”

  • The paper relies on this without support.

    HepG2/C3A cell lines may not express all metabolic enzymes and cannot capture the genetic variability of the patient population.

    “Although, these cell lines may not express all metabolic enzymes and cannot capture the genetic variability of the patient population”
7 of 7

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 diffusivity differs tenfold between methods and results

    Major

    The methods give an oxygen diffusion coefficient of 3.8 × 10−9 m2/s, but the simulation parameters in the results give 3.8 × 10−8 m2/s. The paper does not make clear which value was used to select the perfusion rate.

  • Reported cells per dot imply fewer dots than were printed

    Minor

    A 7 × 7 array has 49 dots. The figure caption's 400,000 total cells at 16,000 cells per dot instead implies 25 dots; its day-30 figures imply the same count.

  • Drug-treatment figure caption gives a different study duration

    Minor

    The results describe monitoring the APAP-treated construct over six days, while the caption for the corresponding biomarker panels says the treatment lasted 30 days.

  • Simulated oxygen-consumption parameter uses per-cell units

    Minor

    The methods define Vmax as a volumetric consumption rate, with units of mol m−3 s−1. The results instead assign it 4.76 × 10−16 mol s−1, the value and units given for consumption by one cell, leaving the stated simulation input inconsistent with its definition.

4 of 4

Cites

Flagged first, then by citations

Research works printed but not linked: 2

  1. Luiz, 2014, Direct-write bioprinting of cell-laden methacrylated gelatin hydrogels, Biofabrication, 6, 10.1088/1758-5082/6/2/024105 · DOI · Paper page
  2. Nichol, 2010, Cell-laden microengineered gelatin methacrylate hydrogels, Biomaterials, 31, 5536, 10.1016/j.biomaterials.2010.03.064 · DOI · Paper page

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Authors
Bhise, N. S., Manoharan, V., Massa, S., Tamayol, A., Ghaderi, M., Miscuglio, M., Lang, Q., Shrike Zhang, Y., Shin, S. R., Calzone, G., Annabi, N., Shupe, T. D., Bishop, C. E., Atala, A., Dokmeci, M. R., & Khademhosseini, A.
Affiliations
  • Brigham and Women's Hospital (US)
  • Biomaterials Innovation Research Center, Department of Medicine, Brigham and Women’s Hospital, Harvard Medical School, Boston, MA 02115,
  • Wake Forest University Health Sciences (US)
Corresponding author
  • Tamayol, Ali
  • Khademhosseini, Ali
ORCID
Venue
Biofabrication
Publisher
IOP Publishing
Published
2016-01-12
Volume
8
Issue
1
Pages
014101
ISSN
  • 1758-5082 (ISSN-L)
  • 1758-5090
DOI
10.1088/1758-5090/8/1/014101
OpenAlex
W2232645088
Cited by
615
References
78
Citations by year
2026: 22 · 2025: 45 · 2024: 55 · 2023: 65 · 2022: 76 · 2021: 81 · 2020: 73 · 2019: 73 · 2018: 58 · 2017: 43 · 2016: 23
FWCI
31.60
Type
journal article
Language
en
Open access
Licence
Locations
  • Biofabrication (publishedVersion)
  • PubMed (publishedVersion)
  • eScholarship (California Digital Library) (submittedVersion)
Funders
  • National Institutes of Health
  • National Science Foundation
  • Defense Threat Reduction Agency (DTRA) under Space and Naval Warfare Systems Center Pacific
Topics
  • Physical Sciences › Engineering › Biomedical Engineering › 3D Printing in Biomedical Research
  • Health Sciences › Medicine › Surgery › Tissue Engineering and Regenerative Medicine
  • Life Sciences › Pharmacology, Toxicology and Pharmaceutics › Pharmacology › Drug-Induced Hepatotoxicity and Protection
Keywords
liver-on-a-chip, HepG2/C3A cells, 3D bioreactor, GelMA hydrogel, bioprinting, drug toxicity assessment, acetaminophen toxicity, hepatocyte function, albumin secretion, MRP2, ZO-1 tight junction, in vitro toxicity models
MeSH terms
Biological Assay, Equipment Design, Humans, Organ Culture Techniques, Toxicity Tests, Spheroids, Cellular, Liver, Artificial, Equipment Failure Analysis, Chemical and Drug Induced Liver Injury, Lab-On-A-Chip Devices, Hep G2 Cells, Printing, Three-Dimensional

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 2026-10-11.

Classified by jev-1.13-20260917 (gold-guideline-v0.2) on 2026-10-11.

Cited sources are checked by gpt-6-sol.

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.

What counts as a claim ›How we measure confidence ›How we check this ›

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