SARS-CoV-2 immune complex triggers human monocyte necroptosis.

We analyzed the ability of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) itself and SARS-CoV-2-IgG immune complexes to trigger human monocyte necroptosis. SARS-CoV-2 was able to induce monocyte necroptosis dependently of MLKL activation. Necroptosis-associated proteins (RIPK1, RIPK3 and MLKL) were involved in SARS-CoV-2N1 gene expression in monocytes. SARS-CoV-2 immune complexes promoted monocyte necroptosis in a RIPK3- and MLKL-dependent manner, and Syk tyrosine kinase was necessary for SARS-CoV-2 immune complex-induced monocyte necroptosis, indicating the involvement of Fcγ receptors on necroptosis. Finally, we provide evidence that elevated LDH levels as a marker of lytic cell death are associated with COVID-19 pathogenesis.

Inhibition of Pseudomonas aeruginosa LPS-Induced airway inflammation by RIPK3 in human airway.

Although the physiological function of receptor-interacting protein kinase (RIPK) 3 has emerged as a critical mediator of programmed necrosis/necroptosis, the intracellular role it plays as an attenuator in human lungs and human bronchial epithelia remains unclear. Here, we show that the expression of RIPK3 dramatically decreased in the inflamed tissues of human lungs, and moved from the nucleus to the cytoplasm. The overexpression of RIPK3 dramatically increased F-actin formation and decreased the expression of genes for pro-inflammatory cytokines (IL-6 and IL-1ß), but not siRNA-RIPK3. Interestingly, whereas RIPK3 was bound to histone 1b without LPS stimulation, the interaction between them was disrupted after 15 min of LPS treatment. Histone methylation could not maintain the binding of RIPK3 and activated movement towards the cytoplasm. In the cytoplasm, overexpressed RIPK3 continuously attenuated pro-inflammatory cytokine gene expression by inhibiting NF-κB activation, preventing the progression of inflammation during Pseudomonas aeruginosa infection. Our data indicated that RIPK3 is critical for the regulation of the LPS-induced inflammatory microenvironment. Therefore, we suggest that RIPK3 is a potential therapeutic candidate for bacterial infection-induced pulmonary inflammation.

Preparation of the RIPK3 Polyclonal Antibody and Its Application in Immunoassays of Nephropathogenic Infectious Bronchitis Virus-Infected Chickens.

Receptor interacting protein kinase 3 (RIPK3) is a vital serine/threonine kinase in regulating the programmed destruction of infected cells to defend against RNA viruses. Although the role of RIPK3 in viruses in mice is well characterized, it remains unclear where in nephropathogenic infectious bronchitis virus (NIBV) in chickens. Here, we use a self-prepared polyclonal antibody to clarify the abundance of RIPK3 in tissues and define the contributions of RIPK3 in tissue damage caused by NIBV infection in chickens. Western blot analyses showed that RIPK3 polyclonal antibody can specifically recognize RIPK3 in the vital tissues of Hy-Line brown chicks and RIPK3 protein is abundantly expressed in the liver and kidney. Moreover, NIBV significantly upregulated the expression levels of RIPK3 in the trachea and kidney of chicks in a time-dependent manner. In addition, the activation of necroptosis in response to NIBV infection was demonstrated by the coimmunoprecipitation (CoIP) experiments through RIPK3 in the necrosome, which phosphorylates its downstream mixed-spectrum kinase structural domain-like protein (MLKL). Our findings offered preliminary insights into the key role of RIPK3 protein in studying the underlying mechanism of organ failure caused by NIBV infection.