Respiratory microphysiological system in respiratory research: recent advances and future prospects.

Respiratory diseases encompass a broad range of conditions that affect the airways, alveolus, and other structures involved in breathing. With respiratory viruses emerging and spreading rapidly, and factors like air pollution, smoking, occupational hazards, and genetics as major risk contributors, respiratory diseases have emerged as a primary cause of illness and mortality around the globe. The development of in vitro respiratory microphysiological systems (MPS), like lung organoids and lung-on-a-chip models, is essential for preventing and treating these diseases effectively.

Generation of ASCL1-mCherry knock-in reporter in human embryonic stem cell line, WAe001-A-2E, using CRISPR/Cas9-based gene targeting.

Achaete-Scute Complex Homolog 1 (ASCL1) is a key regulator in the development and function of the nervous system, particularly in the process of neuronal and neuroendocrine cell differentiation. By employing the CRISPR/Cas9 system, we successfully established an ASCL1-mCherry knock-in human embryonic stem cell (hESC) line by inserting a P2A-mCherry fragment at the ASCL1 locus. The mCherry reporter effectively demonstrated the expression level of endogenous ASCL1 during the process of inducing pulmonary neuroendocrine cells (PNECs) from hESC.

Generation of FOXJ1-EGFP knock-in reporter human embryonic stem cell line, WAe001-A-2D, using CRISPR/Cas9-based gene targeting.

Forkhead box protein J1 (FOXJ1), a member of the forkhead family, is an important transcription factor regulating multiciliated cell differentiation and motile ciliogenic program. Here, we established a FOXJ1- EGFP knock-in human embryonic stem cell (hESC) line by inserting a P2A-EGFP gene cassette of FOXJ1 using CRISPR/Cas9 system. The reporter cell line retained a normal karyotype, expressed comparable pluripotent marker genes, and maintained differentiation potential.

The C and PH domains of CAPS constitute an effective PI(4,5)P2-binding unit essential for Ca-regulated exocytosis.

Ca-dependent activator proteins for secretion (CAPSs) are required for Ca-regulated exocytosis in neurons and neuroendocrine cells. CAPSs contain a pleckstrin homology (PH) domain that binds PI(4,5)P2-membrane. There is also a C domain residing adjacent to the PH domain, but its function remains unclear.