Primary human colonic mucosal barrier crosstalk with super oxygen-sensitive Faecalibacterium prausnitzii in continuous culture.

BACKGROUND: The gut microbiome plays an important role in human health and disease. Gnotobiotic animal and in vitro cell-based models provide some informative insights into mechanistic crosstalk. However, there is no existing system for a long-term co-culture of a human colonic mucosal barrier with super oxygen-sensitive commensal microbes, hindering the study of human-microbe interactions in a controlled manner. METHODS: Here, we investigated the effects of an abundant super oxygen-sensitive commensal anaerobe, Faecalibacterium prausnitzii, on a primary human mucosal barrier using a Gut-MIcrobiome (GuMI) physiome platform that we designed and fabricated. FINDINGS: Long-term continuous co-culture of F. prausnitzii for two days with colon epithelia, enabled by continuous flow of completely anoxic apical media and aerobic basal media, resulted in a strictly anaerobic apical environment fostering growth of and butyrate production by F. prausnitzii, while maintaining a stable colon epithelial barrier. We identified elevated differentiation and hypoxia-responsive genes and pathways in the platform compared with conventional aerobic static culture of the colon epithelia, attributable to a combination of anaerobic environment and continuous medium replenishment. Furthermore, we demonstrated anti-inflammatory effects of F. prausnitzii through HDAC and the TLR-NFKB axis. Finally, we identified that butyrate largely contributes to the anti-inflammatory effects by downregulating TLR3 and TLR4. CONCLUSIONS: Our results are consistent with some clinical observations regarding F. prausnitzii, thus motivating further studies employing this platform with more complex engineered colon tissues for understanding the interaction between the human colonic mucosal barrier and microbiota, pathogens, or engineered bacteria.

Analyzing the Effectiveness of Different Forms of Cardiopulmonary Resuscitation.

BACKGROUND: In order to simulate a heartbeat in a cardiac arrest patient, cardiopulmonary resuscitation (CPR) requires that chest compressions be delivered with a force of at least 560 N at a rate >100 compressions/min. Many new learners initially use CPR forms that may not meet these parameters sufficiently. We examined three forms of CPR: the form recommended by the American Heart Association (AHA) and two forms that are common among new learners but that are considered incorrect, using a CPR manikin placed on a force plate. Four trained CPR users tested the different methods. DISCUSSION: AHA-recommended CPR is the most effective, delivering a force of 737.2 ± 5.3 N at a rate of 103.2 ± 1.2 compressions/min. Compressions using a bent arms method delivered compressions with a force of 511.8 ± 4.1 N at a rate of 112.8 ± 3.0 compressions/min. Compressions using a different hand position from that recommended by the AHA delivered compressions with a force of 433.3 ± 3.2 N at a rate of 115.2 ± 1.2 compressions/min. CONCLUSIONS: AHA-recommended CPR more effectively compresses a patient's heart than the bent-arms method or the alternate hand-position method, and, of the three methods, only the AHA-recommended form can reliably simulate a patient's heartbeat.

High efficiency hydrodynamic bacterial electrotransformation.

Synthetic biology holds great potential for addressing pressing challenges for mankind and our planet. One technical challenge in tapping into the full potential of synthetic biology is the low efficiency and low throughput of genetic transformation for many types of cells. In this paper, we discuss a novel microfluidic system for improving bacterial electrotransformation efficiency and throughput. Our microfluidic system is comprised of non-uniform constrictions in microchannels to facilitate high electric fields with relatively small applied voltages to induce electroporation. Additionally, the microfluidic device has regions of low electric field to assist in electrophoretic transport of nucleic acids into the cells. The device features hydrodynamically controlled electric fields that allow cells to experience a time dependent electric field that is otherwise difficult to achieve using standard electronics. Results suggest that transformation efficiency can be increased by ∼4×, while throughput can increase by 100-1000× compared to traditional electroporation cuvettes. This work will enable high-throughput and high efficiency genetic transformation of microbes, facilitating accelerated development of genetically engineered organisms.