The levels of the anti-aging protein alpha-Klotho, in its soluble form (s-Klotho), are depressed in the circulation of patients with type 1 diabetes (T1D) or type 2 diabetes (T2D). Gene transfer experiments have suggested a protective role for beta-cell specific expression of alpha-Klotho in murine models of T1D and T1D, but these approaches are not easily translatable to clinical therapy. It is unknown whether systemic s-Klotho protein treatment ameliorates disease in T1D, which is characterized by autoimmune destruction of beta cells. We previously reported from in vitro experiments with beta cells that s-Klotho increases insulin secretion, reduces cells death and promotes beta-cell replication. Here, we investigated s-Klotho protein therapy in NOD mice, which have autoimmune T1D. We observed that diabetic NOD mice have significantly lower plasma levels of s-Klotho, compared to their non-diabetic counterparts. To examine in vivo effects of Klotho, we treated NOD mice with s-Klotho protein, or with a Klotho blocking antibody. Systemic treatment with s-Klotho ameliorated diabetes; notably increasing beta-cell replication and total beta-cell mass. Klotho expression was increased locally in the islets. s-Klotho also markedly reduced immune-cell infiltration of islets (insulitis). In contrast, administration of the Klotho antibody was detrimental, and aggravated the loss of beta-cell mass. Thus, s-Klotho has protective effects in this model of T1D, and this appears to depend on a combination of increased beta-cell replication and reduced insulitis. These findings suggest that s-Klotho might be effective as a new therapeutic agent for T1D. (C) 2020 Elsevier Inc. All rights reserved.