1. Can you please briefly summarize the paper? 

Psoriasis is a chronic, T-cell-mediated inflammatory skin disease with a Th17-skewed immune phenotype. Regulatory T cells (Tregs) play a key role in suppressing the inflammatory immune response and inducing immune homeostasis and self-tolerance. While the role of Tregs in the pathogenesis of psoriasis has been proposed, the exact mechanisms of action and the molecular characterization of skin-homing psoriatic Tregs has not yet been well explored. We performed high-dimensional phenotyping of PBMCs using an extensive 39-marker mass cytometry (CyTOF) panel to profile the circulating immune landscape and to deeply interrogate skin-homing Tregs from 31 psoriasis patients and 32 healthy donors. We further validated these findings with an in-vitro chemokine-mediated Treg migration assay and immunofluorescent imaging of normal and psoriatic skin. We also integrated and analyzed several public single-cell RNA-sequencing psoriatic skin datasets to expand upon our circulating findings into the psoriatic skin microenvironment.

We found that psoriatic PBMCs had an increase in frequency of total Tregs compared to healthy controls. However, we discovered that the psoriatic PBMCs had an overall decrease in CLAhi Tregs and specifically, CLAhiCCR5+ Tregs. Cutaneous lymphocyte-associated antigen (CLA) is an essential skin-homing receptor and is expressed on the surface of most skin-infiltrating T cells. Not only was the frequency of CLAhi Tregs decreased, but the expression was also decreased.

These initial findings hinted that there may be an impairment of skin-homing in Tregs. So, we analyzed additional chemokine receptor expression on total Tregs and also CLAhi Tregs and saw that psoriatic Tregs showed a significant increase in CCR7 and a significant decrease in CXCR3, CCR4 and CCR5, which were also correlated with psoriasis severity. CCR4 is another key skin-homing receptor and its ligands CCL17 and CCL22 are expressed by skin epithelial cells and myeloid cells, respectively. CCR5 is also known to be an important skin-homing receptor and CXCR3 has also been show to be critical in trafficking of Tregs to sites of inflammation. CCR7 on the other hand is involved in recirculation of T cells and migration from peripheral tissues into secondary lymphoid organs. Taken together, an increase in secondary lymphoid migration by increased CCR7 expression and a decrease in skin homing by reduced CXCR3, CCR4 and CCR5 expression suggest a potential skin trafficking property of psoriatic Tregs. To further validate this, we performed an in-vitro Treg migration assay and found that psoriatic Tregs were more sensitive to migration towards CCR7’s ligands CCL19 and CCL21 in a concentration-dependent manner. Furthermore, we performed immunofluorescent imaging of normal and psoriatic skin and found a significant increase in CCR7 expression on psoriatic skin-infiltrated Tregs. While Tregs are continuously recruited to the inflamed skin from circulation, upregulated CCR7 expression may hamper persistent skin trafficking and those Tregs may be more sensitive to recirculation and less capable of persisting and maintaining immune homeostasis in the psoriatic skin lesion.

Integrating and interrogating several public psoriatic skin single-cell RNA seq datasets provided a further detailed landscape of psoriatic lesional and non-lesional skin. The results were consistent with the circulating CyTOF findings showing a decrease in CCR4 gene expression and CCR4-mediated chemokine interactions. Similarly, there was consistent downregulation of CXCR3 and CCR5 gene expression and upregulation of CCR7 gene expression. Of note, cell adhesion molecules involved in immune cell migration and retention within the skin, specifically SELPLG, CD69 and ITGB2 were also reduced in Tregs within the lesional skin compared to non-lesional skin.

In conclusion our study identified differences in the immune phenotypes of circulating Tregs and CLA+ Tregs in psoriatic patients compared to healthy controls. Our integrated results of CyTOF and scRNA-seq data demonstrated an inefficient skin-trafficking property of psoriatic Tregs leading to a suboptimal immune-suppressive environment in controlling ongoing psoriatic inflammation.

2. Can you please tell us the main difficulties you had in the laboratory work and how you overcame them?

With regards to this paper, the in-vitro Treg migration assay and the IF staining of chemokine receptors was a bit difficult. Despite reading over previously published papers and following their exact protocols, the Treg migration assay and IF staining was difficult to set up. In the end, validation required constant trial-and-error along with multi-disciplinary collaboration (from obtaining human blood and skin samples) and reaching out to those with experience in these fields was key to properly setting up these assays.

3. Please introduce your laboratory, university or organization to bio-researchers in Korea.

We are the Lab of Immune Tolerance at Seoul National University College of Medicine (SNU LIT). We leverage cutting-edge multi-omics technologies with immunological expertise to more accurately define human disease and pathogenesis for the development of novel therapies against various autoimmune diseases and cancers. We investigate the role of the immune system, particularly regulatory T cells, which play a central role in maintaining immune homeostasis and tolerance. We research the biology of regulatory T cells in circulation and in inflammatory lesions and in cancers to understand their mechanistic role and dysfunction in disease progression and to identify key targets for restoration of function and return to healthy states.

4. Please tell us your experiences and your thoughts related to research activities abroad.

I am a Korean-American born and raised in the US. I attended the University of North Carolina at Chapel Hill where I researched the role of immune cells, particularly microglia in demyelinating disease and graduated in 2016. I then moved to New York to join the Human Immune Monitoring Center (HIMC) at Mt. Sinai under Dr. Miriam Merad, Dr. Seunghee Kim-Schulze and Dr. Adeeb Rahman. I worked for the first 3 years as a senior associate researcher managing the HIMC’s CyTOF team and platform. I collaborated with PIs and researchers to design and conduct CyTOF experiments for clinical and translational studies.

While working on these large, high-dimensional CyTOF datasets, I realized the importance of being able to code to more deeply interrogate and analyze these datasets. After attending a 6-month intensive data analytics bootcamp at Columbia Engineering I then transitioned into a computational role at the HIMC. I became a computational scientist leading the CyTOF team and platform while developing processing and analytical pipelines for analysis of large-scale, multi-study mass cytometry datasets including the Type1 Diabetes Trialnet/JDRF network, NCI Cancer Immune Monitoring and Analysis Centers (CIMAC) and NIAID-supported IMMuno?Phenotyping?Assessment in a?COVID-19?Cohort (IMPACC)).

The leadership and mentorship under Dr. Miriam Merad, Dr. Seunghee Kim-Schulze and Dr. Adeeb Rahman was truly an amazing learning opportunity and experience. The opportunity they provided at the HIMC to explore and grow and question has truly shaped me into the researcher that I am now. As fate would have it, during my time at Sinai, I met Dr. Hyunje Kim while collaborating on a CyTOF research project. Years later, I made the decision to move to Korea for the first time and conduct my PhD in Dr. Hyunje Kim’s laboratory giving me the great opportunity and experience to collaborate, research and learn under Dr. Tae-Gyun Kim, Dr. Hyunje Kim and Dr. Chung-Gy Park.

5. Can you provide some advice for younger scientists who have plans to study abroad?

For those who plan to study abroad I would say to be constantly inquisitive and curious. Don’t be afraid to speak up and ask questions, it is encouraged. Try to stay open-minded and willing to learn new things. There are so many opportunities and areas of study, so try to learn as much a possible from the experts in the field.

6. Future plan?

I plan on continuing my academic career and hope to become a professor building on previous research to better understand normal, healthy states and the mechanisms involved in pathogenesis of various diseases in order to development therapies to treat these diseases. Not only do I want to identify these novel disease drug targets, but I also want to be involved on the industry side to develop these therapies and bring them into the clinic to treat human disease and improve human health.

7. Do you have anything else that you would like to tell Korean scientists and students?

It has been an amazing experience and opportunity working with Korean scientists and students and I truly hope to collaborate together in the future.