Human type 2 innate lymphoid cells (ILC2s) are identified by coupled detection of CRTH2 and IL7Rα on lineage negative (Lin^–) cells. Type 2 cytokine production by CRTH2^–IL7Rα^– innate lymphoid cells (ILCs) is unknown.

We sought to identify CRTH2^–IL7Rα^– type 2 cytokine–producing ILCs and their disease relevance.

We studied human blood and lung ILCs from asthmatic and control subjects by flow cytometry, ELISA, RNA sequencing, quantitative PCR, adoptive transfer to mice, and measurement of airway hyperreactivity by Flexivent.

We found that IL-5 and IL-13 were expressed not only by CRTH2⁺ but also by CRTH2^–IL7Rα⁺ and CRTH2^–IL7Rα^– (double-negative [DN]) human blood and lung cells. All 3 ILC populations expressed type 2 genes and induced airway hyperreactivity when adoptively transferred to mice. The frequency of type 2 cytokine–positive IL7Rα and DN ILCs were similar to that of CRTH2 ILCs in the blood and lung. Their frequency was higher in asthmatic patients than in disease controls. Transcriptomic analysis of CRTH2, IL7Rα, and DN ILCs confirmed the expression of mRNA for type 2 transcription factors in all 3 populations. Unexpectedly, the mRNA for GATA3 and IL-5 correlated better with mRNA for CD30, TNFR2, ICOS, CCR4, and CD200R1 than for CRTH2. By using a combination of these surface markers, especially CD30/TNFR2, we identified a previously unrecognized ILC2 population.

The commonly used surface markers for human ILC2s leave a majority of type 2 cytokine–producing ILC2s unaccounted for. We identified top GATA3-correlated cell surface–expressed genes in human ILCs by RNA sequencing. These new surface markers, such as CD30 and TNFR2, identified a previously unrecognized human ILC2 population. This ILC2 population is likely to contribute to asthma.