Human Kidney Spheroids and Monolayers Provide Insights into SARS-CoV-2 Renal Interactions
Authors: Omer Dorit , Pleniceanu Oren , Gnatek Yehudit , Namestnikov Michael , Cohen-Zontag Osnat , Goldberg Sanja , Friedman Yehudit Eden , Friedman Nehemya , Mandelboim Michal , Vitner Einat B. , Achdout Hagit , Avraham Roy , Zahavy Eran , Israely Tomer , Mayan Haim , Dekel Benjamin 
Published: June 2021

Significance Statement
Coronavirus disease 2019 (COVID-19) commonly results in AKI, but it is unknown whether AKI in patients with COVID-19 stems from direct kidney infection or indirect mechanisms. The authors explored the effects of infection with the causative virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), both in monolayers of primary human kidney cells and in kidney spheroids, a three-dimensional model recapitulating human kidneys in cell composition and function. They demonstrated that the virus effectively infects and replicates in human tubular epithelial cells, but does not cause cytopathic effects. They also provide molecular evidence that activation of a type 1 IFN response may be the underlying mechanism of resistance to SARS-CoV-2–related cytopathic damage. In all, the findings indicate minimal—if any—direct tubular damage by SARS-CoV-2.

Background
Although coronavirus disease 2019 (COVID-19) causes significan t morbidity, mainly from pulmonary involvement, extrapulmonary symptoms are also major componen ts of the disease. Kidney disease, usually presenting as AKI, is particularly severe among patients with COVID-19. It is unknown, however, whether such injury results from direct kidney infection with COVID-19’s causative virus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), or from indirect mechanisms.

Methods
Using ex vivo cell models, we sought to analyze SARS-CoV-2 interactions with kidney tubular cells and assess direct tubular injury. These models comprised primary human kidney epithelial cells (derived from nephrectomies) and grown as either proliferating monolayers or quiescent three-dimensional kidney spheroids.

Results
We demonstrated that viral entry molecules and high baseline levels of type 1 IFN–related molecules were present in monolayers and kidney spheroids. Although both models support viral infection and replication, they did not exhibit a cytopathic effect and cell death, outcomes that were strongly present in SARS-CoV-2–infected controls (African green monkey kidney clone E6 [Vero E6] cultures). A comparison of monolayer and spheroid cultures demonstrated higher infectivity and replication of SARS-CoV-2 in actively proliferating monolayers, although the spheroid cultures exhibited high er levels of ACE2. Monolayers exhibited elevation of some tubular injury molecules—including molecules related to fibrosis (COL1A1 and STAT6) and dedifferentiation (SNAI2)—and a loss of cell identity, evident by reduction in megalin (LRP2). The three-dimensional spheroids were less prone to such injury.

Conclusions
SARS-CoV-2 can infect kidney cells without a cytopathic effect. AKI-induced cellular proliferation may potentially intensify infectivity and tubular damage by SARS-CoV-2, suggesting that early intervention in AKI is warranted to help minimize kidney infection.