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Quantifying HIV-1-Mediated Gut CD4+ T Cell Deathin the Lamina Propria Aggregate Culture (LPAC) Model.

Bio-protocol (2020-01-20)
Stephanie M Dillon, Kejun Guo, Moriah J Castleman, Mario L Santiago, Cara C Wilson
ABSTRACT

Gut CD4 T cells are major targets of HIV-1 and are massively depleted early during infection. To better understand the mechanisms governing HIV-1-mediated CD4 T cell death, we developed the physiologically-relevant Lamina Propria Aggregate Culture (LPAC) model. The LPAC model is ideal for studying CD4 T cell death induced by clinically-relevant Transmitted/Founder (TF) HIV-1 strains and is also suitable for studying how enteric microbes and soluble factors (e.g., Type I Interferons) impact LP CD4 T cell death and function. Here, we detail the protocol to establish LP CD4 T cell infection using a process of spinoculation, the subsequent evaluation of infection levels using multicolor flow cytometry and the determination of overall LP CD4 T cell death using absolute LP CD4 T cell counts. We also describe the preparation of virus stocks of Transmitted/Founder (TF) HIV-1 infectious molecular clones that were successfully used in the LPAC model.

MATERIALS
Product Number
Brand
Product Description

Sigma-Aldrich
Calcium chloride, anhydrous, granular, ≤7.0 mm, ≥93.0%