Elizabeth Calamari

Bio: Elizabeth Calamari is an academic researcher from Wyss Institute for Biologically Inspired Engineering. The author has contributed to research in topics: Bone marrow & Microbiome. The author has an hindex of 8, co-authored 10 publications receiving 499 citations. Previous affiliations of Elizabeth Calamari include Worcester Polytechnic Institute.

A Complex Human-Gut Microbiome Cultured in an Anaerobic Intestine-on-a-Chip

Abstract: The diverse bacterial populations that comprise the commensal microbiome of the human intestine play a central role in health and disease. A method that sustains complex microbial communities in direct contact with living human intestinal cells and their overlying mucus layer in vitro would thus enable the investigation of host-microbiome interactions. Here, we show the extended coculture of living human intestinal epithelium with stable communities of aerobic and anaerobic human gut microbiota, using a microfluidic intestine-on-a-chip that permits the control and real-time assessment of physiologically relevant oxygen gradients. When compared to aerobic coculture conditions, the establishment of a transluminal hypoxia gradient in the chip increased intestinal barrier function and sustained a physiologically relevant level of microbial diversity, consisting of over 200 unique operational taxonomic units from 11 different genera and an abundance of obligate anaerobic bacteria, with ratios of Firmicutes and Bacteroidetes similar to those observed in human faeces. The intestine-on-a-chip may serve as a discovery tool for the development of microbiome-related therapeutics, probiotics and nutraceuticals.

Robotic fluidic coupling and interrogation of multiple vascularized organ chips.

Abstract: Organ chips can recapitulate organ-level (patho)physiology, yet pharmacokinetic and pharmacodynamic analyses require multi-organ systems linked by vascular perfusion. Here, we describe an 'interrogator' that employs liquid-handling robotics, custom software and an integrated mobile microscope for the automated culture, perfusion, medium addition, fluidic linking, sample collection and in situ microscopy imaging of up to ten organ chips inside a standard tissue-culture incubator. The robotic interrogator maintained the viability and organ-specific functions of eight vascularized, two-channel organ chips (intestine, liver, kidney, heart, lung, skin, blood-brain barrier and brain) for 3 weeks in culture when intermittently fluidically coupled via a common blood substitute through their reservoirs of medium and endothelium-lined vascular channels. We used the robotic interrogator and a physiological multicompartmental reduced-order model of the experimental system to quantitatively predict the distribution of an inulin tracer perfused through the multi-organ human-body-on-chips. The automated culture system enables the imaging of cells in the organ chips and the repeated sampling of both the vascular and interstitial compartments without compromising fluidic coupling.

On-chip recapitulation of clinical bone marrow toxicities and patient-specific pathophysiology.

Abstract: The inaccessibility of living bone marrow (BM) hampers the study of its pathophysiology under myelotoxic stress induced by drugs, radiation or genetic mutations. Here, we show that a vascularized human BM-on-a-chip (BM chip) supports the differentiation and maturation of multiple blood cell lineages over 4 weeks while improving CD34+ cell maintenance, and that it recapitulates aspects of BM injury, including myeloerythroid toxicity after clinically relevant exposures to chemotherapeutic drugs and ionizing radiation, as well as BM recovery after drug-induced myelosuppression. The chip comprises a fluidic channel filled with a fibrin gel in which CD34+ cells and BM-derived stromal cells are co-cultured, a parallel channel lined by human vascular endothelium and perfused with culture medium, and a porous membrane separating the two channels. We also show that BM chips containing cells from patients with the rare genetic disorder Shwachman–Diamond syndrome reproduced key haematopoietic defects and led to the discovery of a neutrophil maturation abnormality. As an in vitro model of haematopoietic dysfunction, the BM chip may serve as a human-specific alternative to animal testing for the study of BM pathophysiology. A vascularized human bone-marrow-on-a-chip improves the maintenance of patient-derived CD34+ cells, and recapitulates clinically relevant aspects of bone marrow injury as well as key haematopoietic defects of patients with a rare genetic disorder.

Scalable Fabrication of Stretchable, Dual Channel, Microfluidic Organ Chips.

Abstract: A significant number of lead compounds fail in the pharmaceutical pipeline because animal studies often fail to predict clinical responses in human patients. Human Organ-on-a-Chip (Organ Chip) microfluidic cell culture devices, which provide an experimental in vitro platform to assess efficacy, toxicity, and pharmacokinetic (PK) profiles in humans, may be better predictors of therapeutic efficacy and safety in the clinic compared to animal studies. These devices may be used to model the function of virtually any organ type and can be fluidically linked through common endothelium-lined microchannels to perform in vitro studies on human organ-level and whole body-level physiology without having to conduct experiments on people. These Organ Chips consist of two perfused microfluidic channels separated by a permeable elastomeric membrane with organ-specific parenchymal cells on one side and microvascular endothelium on the other, which can be cyclically stretched to provide organ-specific mechanical cues (e.g., breathing motions in lung). This protocol details the fabrication of flexible, dual channel, Organ Chips through casting of parts using 3D printed molds, enabling combination of multiple casting and post-processing steps. Porous poly (dimethyl siloxane) (PDMS) membranes are cast with micrometer sized through-holes using silicon pillar arrays under compression. Fabrication and assembly of Organ Chips involves equipment and steps that can be implemented outside of a traditional cleanroom. This protocol provides researchers with access to Organ Chip technology for in vitro organ- and body-level studies in drug discovery, safety and efficacy testing, as well as mechanistic studies of fundamental biological processes.

Fabrication and characterization of electrospun polycaprolactone and gelatin composite cuffs for tissue engineered blood vessels

Abstract: Sewing cuffs incorporated within tissue-engineered blood vessels (TEBVs) enable graft anastomosis in vivo, and secure TEBVs to bioreactors in vitro. Alternative approaches to cuff design are required to achieve cuff integration with scaffold-free TEBVs during tissue maturation. To create porous materials that promote tissue integration, we used electrospinning to fabricate cuffs from polycaprolactone (PCL), PCL blended with gelatin, and PCL coated with gelatin, and evaluated cuff mechanical properties, porosity, and cellular attachment and infiltration. Gelatin blending significantly decreased cuff ultimate tensile stress and failure strain over PCL alone, but no significant differences were observed in elastic modulus or failure load. Interestingly, gelatin incorporation by blending or coating did not produce significant differences in cellular attachment or pore size. We then created tissue tubes by fusing self-assembled smooth muscle cell rings together with electrospun cuffs on either end. After 7 days, rings and cuffs fused seamlessly, and the resulting tubes were harvested for pull-to-failure tests to measure the strength of cuff-tissue integration. Tubes with gelatin-coated PCL cuffs failed more frequently at the cuff-tissue interface compared to PCL and PCL:gelatin blended groups. This work demonstrates that electrospun cuffs integrated successfully with scaffold-free TEBVs, and that the addition of gelatin did not significantly improve cuff integration over PCL alone for this application. Electrospun cuffs may aid cannulation for dynamic culture and testing of tubular constructs during engineered tissue maturation. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 106B: 817-826, 2018.

Gut Immune Maturation Depends Upon Colonization with Host-Specific Microbiota

Abstract: Gut microbial induction of host immune maturation exemplifies host-microbe mutualism. We colonized germ-free (GF) mice with mouse microbiota (MMb) or human microbiota (HMb) to determine whether small intestinal immune maturation depends on a coevolved host-specific microbiota. Gut bacterial numbers and phylum abundance were similar in MMb and HMb mice, but bacterial species differed, especially the Firmicutes. HMb mouse intestines had low levels of CD4(+) and CD8(+) T cells, few proliferating T cells, few dendritic cells, and low antimicrobial peptide expression--all characteristics of GF mice. Rat microbiota also failed to fully expand intestinal T cell numbers in mice. Colonizing GF or HMb mice with mouse-segmented filamentous bacteria (SFB) partially restored T cell numbers, suggesting that SFB and other MMb organisms are required for full immune maturation in mice. Importantly, MMb conferred better protection against Salmonella infection than HMb. A host-specific microbiota appears to be critical for a healthy immune system.

Modelling cancer in microfluidic human organs-on-chips.

Abstract: One of the problems that has slowed the development and approval of new anticancer therapies is the lack of preclinical models that can be used to identify key molecular, cellular and biophysical features of human cancer progression. This is because most in vitro cancer models fail to faithfully recapitulate the local tissue and organ microenvironment in which tumours form, which substantially contributes to the complex pathophysiology of the disease. More complex in vitro cancer models have been developed, including transwell cell cultures, spheroids and organoids grown within flexible extracellular matrix gels, which better mimic normal and cancerous tissue development than cells maintained on conventional 2D substrates. But these models still lack the tissue-tissue interfaces, organ-level structures, fluid flows and mechanical cues that cells experience within living organs, and furthermore, it is difficult to collect samples from the different tissue microcompartments. In this Review, we outline how recent developments in microfluidic cell culture technology have led to the generation of human organs-on-chips (also known as organ chips) that are now being used to model cancer cell behaviour within human-relevant tissue and organ microenvironments in vitro. Organ chips enable experimentalists to vary local cellular, molecular, chemical and biophysical parameters in a controlled manner, both individually and in precise combinations, while analysing how they contribute to human cancer formation and progression and responses to therapy. We also discuss the challenges that must be overcome to ensure that organ chip models meet the needs of cancer researchers, drug developers and clinicians interested in personalized medicine.

Organs-on-chips: into the next decade

Abstract: Organs-on-chips (OoCs), also known as microphysiological systems or ‘tissue chips’ (the terms are synonymous), have attracted substantial interest in recent years owing to their potential to be informative at multiple stages of the drug discovery and development process. These innovative devices could provide insights into normal human organ function and disease pathophysiology, as well as more accurately predict the safety and efficacy of investigational drugs in humans. Therefore, they are likely to become useful additions to traditional preclinical cell culture methods and in vivo animal studies in the near term, and in some cases replacements for them in the longer term. In the past decade, the OoC field has seen dramatic advances in the sophistication of biology and engineering, in the demonstration of physiological relevance and in the range of applications. These advances have also revealed new challenges and opportunities, and expertise from multiple biomedical and engineering fields will be needed to fully realize the promise of OoCs for fundamental and translational applications. This Review provides a snapshot of this fast-evolving technology, discusses current applications and caveats for their implementation, and offers suggestions for directions in the next decade. Organs-on-chips (OoCs) could be useful at various stages of drug discovery and development, providing insight regarding human organ physiology in both normal and disease contexts, as well as accurately predicting developmental drug safety and efficacy. This Review discusses the advances that have enabled OoCs to demonstrate physiological relevance, and the challenges and opportunities that need to be tackled to tap the full potential of OoC utility for translational research.

Organ-on-a-chip: recent breakthroughs and future prospects

Abstract: The organ-on-a-chip (OOAC) is in the list of top 10 emerging technologies and refers to a physiological organ biomimetic system built on a microfluidic chip. Through a combination of cell biology, engineering, and biomaterial technology, the microenvironment of the chip simulates that of the organ in terms of tissue interfaces and mechanical stimulation. This reflects the structural and functional characteristics of human tissue and can predict response to an array of stimuli including drug responses and environmental effects. OOAC has broad applications in precision medicine and biological defense strategies. Here, we introduce the concepts of OOAC and review its application to the construction of physiological models, drug development, and toxicology from the perspective of different organs. We further discuss existing challenges and provide future perspectives for its application.

Adhesion mechanisms mediated by probiotics and prebiotics and their potential impact on human health.

Abstract: Adhesion ability to the host is a classical selection criterion for potential probiotic bacteria that could result in a transient colonisation that would help to promote immunomodulatory effects, as well as stimulate gut barrier and metabolic functions. In addition, probiotic bacteria have a potential protective role against enteropathogens through different mechanisms including production of antimicrobial compounds, reduction of pathogenic bacterial adhesion and competition for host cell binding sites. The competitive exclusion by probiotic bacteria has a beneficial effect not only on the gut but also in the urogenital tract and oral cavity. On the other hand, prebiotics may also act as barriers to pathogens and toxins by preventing their adhesion to epithelial receptors. In vitro studies with different intestinal cell lines have been widely used along the last decades to assess the adherence ability of probiotic bacteria and pathogen antagonism. However, extrapolation of these results to in vivo conditions still remains unclear, leading to the need of optimisation of more complex in vitro approaches that include interaction with the resident microbiota to address the current limitations. The aim of this mini review is to provide a comprehensive overview on the potential effect of the adhesive properties of probiotics and prebiotics on the host by focusing on the most recent findings related with adhesion and immunomodulatory and antipathogenic effect on human health.