The novel severe acute respiratory syndrome coronavirus (SARS-CoV-2) is a highly 67 pathogenic virus causing severe respiratory infections, described as coronavirus proinflammatory cytokines and thromboembolic events in various organs (Becker, 83 2020) . Consequently, SARS-CoV-2 is regarded as a pneumotropic virus that infects the 84 patient via the lung but can also cause a systemic infection that affects different organs 85 with a high mortality rate. 86
Little is known about the initial infection process in the alveolar lung tissue, 87 particularly about mechanisms that destroy the lung and mechanisms that allow the 88 virus to affect different organs in the body. Infection models that closely reflect the 89 patient's situation are mainly lacking in part due to the challenges to infect mice and 90 the difficulty of accessing and analyzing infected human alveolar lung cells. Up to 91 now, SARS-CoV-2 infection models have been mainly performed with human airway 92 (non-alveolar) cells or non-human cell lines that naturally express the ACE2 viral 93 receptor, such as the African Green Monkey Vero 76 cell line (Hoffmann et al., 2020) . 94
These cells lack organ-and species-specific characteristics of human lung epithelial 95 cells. For this purpose, cancerous lung epithelial cells (Calu-3 cells), can at least to some 96 extent, reflect the response of the lung epithelium to viral infection (Bestle et al., 2020) . 97
In our study, we present a human-specific in vitro, alveolus-on-a-chip model 98 but also for an adequate immune response. 106
We were able to show that SARS-CoV-2 replicates in the epithelial layer while 107
inducing an acute and robust inflammatory response followed by the destruction of 108 the epithelial layer. Interestingly, in this infection scenario, the endothelial cells were 109 not invaded by SARS-CoV-2 and did not propagate the virus, but nevertheless the 110 epithelial/endothelial barrier integrity was disrupted. 111
Efficient SARS-CoV-2 isolation from patients and propagation in cell-culture 115
To gain fully infectious viral particles for our studies, we collected three respiratory 116 specimens from qRT-PCR-proven COVID-19 patients and performed SARS-CoV-2 117 propagation in cell culture systems (Vero-76 cells). By repeated infection of host cells 118 and viral replication, we were able to isolate high viral titers originating from three 119 different patients. Within our studies, the SARS-CoV-2 isolates SARS-CoV-120 2/hu/Germany/Jena-vi005159/2020 (5159), SARS-CoV-2/hu/Germany/Jena-121 vi005187/2020 (5587) and SARS-CoV-2/hu/Germany/Jena-vi005588/2020 (5588) were 122 employed. Sequencing of virus isolates verified that all three viral strains belong to 123 SARS-CoV-2 (species Severe acute respiratory syndrome-related coronavirus, genus 124
Betacoronavirus) (Gorbalenya et al., 2020) . Phylogenetic analysis revealed a close 125 relationship of SARS-CoV-2 to the SARS-related coronaviruses RaTG13, bat-SL-126
CoVZXC21 and bat-SL-CoVZC45 ( Figure 1A ). Within the SARS-CoV-2 clade, the 127 sequences of strains 5587 and 5588 exhibit two base substitutions T8,782C (nsp1ab: released. These results indicate that SARS-CoV-2 induces replication complexes at ER-153 derived membranes, which were already shown for other types of coronaviruses 154 (Stertz et al., 2007) and also confirm the findings for Vero E6 cells (Ogando et al., 2020) . 155
To better mimic the situation in the human pulmonary alveoli, we performed the 156 infection in Calu-3 cells. In Figure 2B , immunofluorescence measurements compare 157 To further analyze the barrier integrity of the biochip system on a functional level, we 277 performed permeability assays using FITC-dextran to investigate the endothelial and 278 epithelial cell barrier integrity. We were able to show that SARS-CoV-2 significantly 279 increased the tissue permeability with its barrier function severely impaired (Figure 280 Many studies reveal that most severe cases of SARS-CoV-2 infections are not only due 307 to enhanced viral burden, but to a large extent due to aberrant immune responses 308 (Broggi et al., 2020) . 309
In our manuscript, we present an infection model that could be further used to study Netherlands) was used. RNA was thawed on ice. 400 nanogram (ng) RNA were 499 diluted in RNase free water to a volume of 13 μl. 2 μl gDNA removal mix were added 500 to the diluted RNA; followed by incubation at 45°C for 2 min. After incubating the 501 samples for at least 1 min on ice, 5 μl of RT master mix (containing 4 μl Reverse 502
Transcription Mix and 1 μl Reverse Transcription enzyme per sample) were added. 503
The resulting mixture was incubated for 3 min at 25°C, followed by incubation at 45°C 504 for 10 min and an inactivation step at 85°C for 5 min. The cDNA was either directly 505 used for the subsequent experiments or stored at -20°C. 506
qRT-PCRs were performed using the QuantiNova SYBR Green PCR Kit (QIAGEN, 507
Venlo, Netherlands). 1 μl cDNA was added to 19 μl master mix (containing 10 μl SYBR 508
Green, 1.5 μl Forward Primer (10 μM), 1.5 μl Reverse Primer (10 μM) and 6 μL RNase 509 free ddH2O per sample; for primer sequences see table 1), and the real time PCR 510 reaction was started using the following cycle conditions: 95l°C for 2 min, followed by 511 40 cycles of 95°C for 5 sec and 60°C for 10 sec. The qPCR cycle was ended by a stepwise 512 temperature-increase from 60°C to 95°C (1°C every 5 sec). 