The hallmark of systemic sclerosis (SSc) is skin fibrosis due to persistent myofibroblast activation and persistence. SSc is associated with reduced cutaneous expression of SIRT1 and SIRT3. The activity of SIRTs is determined by availability of NAD+, which in turn is regulated by the ectoenzyme CD38, NAD+-hydrolase. During chronological aging, up-regulation of CD38 drives cellular NAD+ consumption, resulting in reduced SIRT activity and mitochondrial dysfunction. We hypothesize that biological aging is accelerated in SSc, causing NAD+ depletion, cellular senescence and fibrosis. Genome-wide expression microarrays showed up-regulated CD38 in SSc skin biopsies, while NAD+ biosynthesis enzymes were unaltered. Notably, cutaneous CD38 levels showed strong correlation with the TGF-ß signature (p<0.0001, r=0.42) as well as skin score (p<0.003, r =0.34), and anti-correlation with SIRT activity. The senescent cells were elevated in SSc skin biopsies. In cultured dermal fibroblast, recombinant CD38 reduced SIRT activity and worsened fibrotic responses, while CD38 inhibition mitigated fibrotic responses. Moreover, NAD+ supplementation alone, or combined with CD38 inhibition, abrogated TGF-ß-mediated fibrotic responses and Smad-dependent transcriptional activity. In vivo, NAD+ repletion alone or combined with CD38 inhibition in aged mice attenuated skin fibrosis and inflammation. These anti-fibrotic effects were correlated with increased tissue levels of NAD+. The results are the first to identify altered NAD+ metabolism in fibrosis in SSc and implicate elevated CD38 NADase activity as a pathogenic driver contributing to dermal fibrosis. Targeting CD38 to restore NAD+homeostasis represents a potentially safe and effective approach to mitigate fibrosis in SSc.

Elaine F. Remmers1, Fulya Cosan2, Yohei Kirino1, Michael J. Ombrello1, Neslihan Abaci3, Colleen Satorius1, Julie M. Le1, Barbara Yang4, Benjamin D. Korman1, Aris Cakiris3, Oznur Aglar3, Zeliha Emrence3, Hulya Azakli3, Duran Ustek3, Ilknur Tugal-Tutkun5, Gulsen Akman-Demir6, Wei Chen7, Christopher I. Amos7, Michael B. Dizon4, Afet Akdag Kose8, Gulsevim Azizlerli8, Burak Erer2, Oliver J. Brand9, Virginia G. Kaklamani10, Phaedon Kaklamanis11, Eldad Ben-Chetrit12, Miles Stanford13, Farida Fortune14, Marwen Ghabra15, William E. R. Ollier16, Young-Hun Cho17, Dongsik Bang18, John O'Shea19, Graham R. Wallace20, Massimo Gadina4, Daniel L. Kastner1, and Ahmet Gül2,3 1 Laboratory of Clinical Investigation, National Institute of Arthritis and Musculoskeletal and Skin Diseases, Bethesda, Maryland, USA

Single nucleotide polymorphisms in the STAT4 gene have recently been shown to be associated with rheumatoid arthritis (RA) and systemic lupus erythematosus (SLE). Primary Sjögren's sydrome (pSS) is a related autoimmune disease thought to have a pathogenesis similar to these diseases. To test the hypothesis that the variant haplotype of STAT4 seen in RA and SLE is also associated with pSS, we genotyped rs7574865, the most strongly disease-associated SNP in the variant STAT4 haplotype, in 124 Caucasian pSS subjects and compared them to 1143 Caucasian controls. The disease-associated T allele was more common in chromosomes of the pSS patients (29.6%) than in controls (22.3%), leading to a P-value for association of 0.01. These results implicate polymorphisms in the STAT4 gene in the pathogenesis of pSS.