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We thank Dr Manetti for his comments regarding the distinctive CD34 positive dermal stromal cell population and their pathological changes in systemic sclerosis (SSc) skin.1–4 One challenge for the field of SSc dermatopathology is a lack of broad consensus regarding the terminology used to describe CD34+ cells.5 Indeed, CD34+ cells have been described as fibrocytes,6 stromal fibroblastic cells,7 dermal dendritic cells,8 fibroblasts5 9–11 and telocytes.3 4 Our work adds to prior observations by noting that skin samples that exhibit a loss of CD34+ cells, gain expression of alpha-smooth muscle actin (aSMA) in a similar microanatomic distribution, specifically the deeper dermis and subcutaneous fat. This pattern of CD34 and aSMA reversed as patients improved clinically and was associated with a gene signature relevant to clinical improvement.12 Our study was not able to conclude whether this polarisation of aSMA and CD34 staining in scleroderma skin represents aSMA+ cells that transdifferentiated into CD34+ cells over time or aSMA+ cells that died and were replaced by new CD34+ cells. Manetti et al conducted a comprehensive immunophenotyping and structural study of CD34+ cells in SSc and did not observe costaining of CD34 with CD90 or aSMA.3 However, other studies of SSc, scarring, and tissue repair have identified CD90+/CD34+ cells,7 9 CD90+/aSMA+ cells,7 9 and CD34+/aSMA+ cells,10 supportive of the fibroblast transition hypothesis. When examining biopsies with the various aSMA and CD34 staining patterns, we recognise that aSMA+ cells differ morphologically from CD34+ cells. Specifically, aSMA+ cells are larger than the CD34+ cells, exhibiting a rather abundant eosinophilic cytoplasm and vesicular chromatin.
As Dr Manetti highlights, a study limitation includes that we did not perform concurrent CD31 staining which may have resulted in the incorporation of some CD34+ endothelial cells into the CD34 staining score. However, the interstitial intervascular CD34 staining pattern contrasts the well-defined microvascular staining, highlighted by CD34 and CD31. In severe scleroderma, the negative CD34 staining pattern in the intervascular interstitium is in contradistinction to the strongly positively staining vessels, the latter serving as an important positive internal control (figure 1).
In summary, it is increasingly clear that multiple, distinct stromal cell and fibroblast populations play a role in SSc skin, and at least nine types of dermal fibroblasts have been identified by single-cell RNA sequencing in various skin diseases.13 This underscores that a more detailed understanding cellular heterogeneity may be essential for understanding SSc disease pathogenesis and clinical phenotypes. We look forward to future studies using higher dimensional analyses such as single-cell RNA sequencing and multiparameter immunofluorescence staining to further characterise SSc cell types and international collaborative efforts to come to consensus regarding nomenclature.
Handling editor Josef S Smolen
Contributors KS and YZ wrote the first version of the manuscript. All authors read, critically revised and approved the final manuscript. DEO and JKG contributed equally.
Funding This study was supported by the Hospital for Special Surgery Rheumatology Council (JKG) and the Scleroderma Clinical Trials Consortium (SCTC) Working Group Grant (KS). This study uses data from the nilotinib and belimumab trials in systemic sclerosis that were investigator-initiated studies supported by research grants from Novartis and GlaxoSmithKline, respectively.
Competing interests RS reports receiving funds for the following activities: research support: GlaxoSmithKline, Genentech/Roche, Novartis, Corbus Pharmaceuticals, Cytori Therapeutics, Evidera, Actelion Pharmaceuticals, ChemoCentryx, Boehringer Ingelheim Pharmaceuticals, Forbius, InfaRx, Sanofi, Kadmon Corporation; consulting: GlaxoSmithKline, Janssen Pharmaceuticals, Sanofi Aventis, ChemoCentryx, Forbius, CSL Behring; DEO reports receiving funds from Pfizer and personal fees from Astra Zeneca, outside the submitted work; JKG reports receiving funds for the following activities: research support: Corbus Pharmaceuticals, Cumberland Pharmaceuticals, and Eicos Sciences, outside the submitted work.
Patient and public involvement Patients and/or the public were not involved in the design, or conduct, or reporting, or dissemination plans of this research.
Patient consent for publication Not required.
Provenance and peer review Commissioned; internally peer reviewed.
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