An optimized 3D-printed perfusion bioreactor for homogeneous cell seeding in bone substitute scaffolds for future chairside applications.

A clinical implementation of cell-based bone regeneration in combination with scaffold materials requires the development of efficient, controlled and reproducible seeding procedures and a tailor-made bioreactor design. A perfusion system for efficient, homogeneous, and rapid seeding with human adipogenic stem cells in bone substitute scaffolds was designed. Variants concerning medium inlet and outlet port geometry, i.e. cylindrical or conical diffuser, cell concentration, perfusion mode and perfusion rates were simulated in silico. Cell distribution during perfusion was monitored by dynamic [18F]FDG micro-PET/CT and validated by laser scanning microscopy with three-dimensional image reconstruction. By iterative feedback of the in silico and in vitro experiments, the homogeneity of cell distribution throughout the scaffold was optimized with adjustment of flow rates, cell density and perfusion properties. Finally, a bioreactor with a conical diffusor geometry was developed, that allows a homogeneous cell seeding (hoover coefficient: 0.24) in less than 60 min with an oscillating perfusion mode. During this short period of time, the cells initially adhere within the entire scaffold and stay viable. After two weeks, the formation of several cell layers was observed, which was associated with an osteogenic differentiation process. This newly designed bioreactor may be considered as a prototype for chairside application.

Bronchoalveolar lavage-detected SARS-CoV-2 infection in presence of serial negative nasopharyngeal swabs: a case report.

We describe a case of a SARS coronavirus 2 (SARS-CoV-2) infection in a Swiss 54-years-old immunocompromised patient (lymphoma, therapy with the anti-CD20 antibody Rituximab® ), with initial scarce respiratory symptoms but typical coronavirus disease 2019 (COVID-19) radiological presentation, and symptoms onset during a holiday trip to Texas (USA). Three nasopharyngeal swabs in the 96 hours following hospital admission were negative, despite a CT thorax suggestive for an early stage of infection. COVID-infection was finally confirmed in the bronchoalveolar lavage (BAL) fluid, performed for exclusion of an alternative diagnosis in immunocompromised. In the BAL an increased cellularity with marked lymphocytosis of 35%, a reduced CD4/CD8 ratio of 0.1 and borderline neutrophilia of 3% were found. This finding might be due to the concomitant therapy with anti-CD20 antibodies, but the presence of lymphocytosis in the BAL despite peripheral lymphopenia with decreased CD4/CD8 T-cells ratio are described here for the first time in a SARS-CoV-2 infection. Persistent gastrointestinal symptoms (diarrhea), fever and initially headache were the predominant symptoms. The respiratory symptoms were scarce (variable mild dyspnea mMRC1). The respiratory conditions worsened during the hospital stay, with tachypnea up to 35/min, increased need for supplemental oxygen up to 8 L/min and worsening lung infiltrates on CT thorax on day 5. A therapy with hydroxychloroquine (HCQ) and an immunoglobulin-supplementation were given, with clinical and respiratory improvement, without need for intensive care or any ventilator support, and hospital discharge on day 16. Our case highlights some diagnostic and therapeutical challenges occurring in patients with COVID-19 infection. As take-home message, in the presence of clinical and radiological findings compatible with SARS-CoV-2 infection we outline the importance of treating patients accordingly, also in presence of repeated negative nasopharyngeal swabs. In selected patients as in our case a bronchoscopic BAL should be considered to exclude other infections, but in our opinion not primarily to confirming COVID-19 infection. Our unique finding of a lymphocytosis in the BAL during a COVID-19 infection needs further investigations.