Carnosine improves diabetic complications, including diabetic nephropathy, in in vivo models. To further understand the underlying mechanism of nephroprotection, we studied the effect of carnosine under glucose-induced stress on cellular stress response proteins in murine immortalized podocytes, essential for glomerular function. High-glucose stress initiated stress response by increasing intracellular heat shock protein 70 (Hsp70), sirtuin-1 (Sirt-1), thioredoxin (Trx), glutamate-cysteine ligase (gamma-glutamyl cysteine synthetase; γ-GCS) and heme oxygenase-1 (HO-1) in podocytes by 30–50% compared to untreated cells. Carnosine (1 mM) also induced a corresponding upregulation of these intracellular stress markers, which was even more prominent compared to glucose for Hsp70 (21%), γ-GCS and HO-1 (13% and 20%, respectively; all p < 0.001). Co-incubation of carnosine (1 mM) and glucose (25 mM) induced further upregulation of Hsp70 (84%), Sirt-1 (52%), Trx (35%), γ-GCS (90%) and HO-1 (73%) concentrations compared to untreated cells (all p < 0.001). The glucose-induced increase in 4-hydroxy-trans-2-nonenal (HNE) and protein carbonylation was reduced dose-dependently by carnosine by more than 50% (p < 0.001). Although podocytes tolerated high carnosine concentrations (10 mM), high carnosine levels only slightly increased Trx and γ-GCS (10% and 19%, respectively, compared to controls; p < 0.001), but not Hsp70, Sirt-1 and HO-1 proteins (p not significant), and did not modify the glucose-induced oxidative stress response. In podocytes, carnosine induced cellular stress tolerance and resilience pathways and was highly effective in reducing high-glucose-induced glycative and lipoperoxidative stress. Carnosine in moderate concentrations exerted a direct podocyte molecular protective action.

The generation, maturation and remodelling of the extracellular matrix (ECM) are essential for the formation of alveoli during lung development. Alveoli formation is disturbed in preterm infants that develop bronchopulmonary dysplasia (BPD), where collagen fibres are malformed, and perturbations to lung ECM structures may underlie BPD pathogenesis. Malformed ECM structures might result from abnormal protein cross-linking, in part attributable to the increased expression and activity of transglutaminase 2 (TGM2) that have been noted in affected patient lungs, as well as in hyperoxia-based BPD animal models. The objective of the present study was to assess whether TGM2 plays a causal role in normal and aberrant lung alveolarization. Targeted deletion of Tgm2 in C57BL/6J mice increased septal thickness and reduced gas-exchange surface area in otherwise normally developing lungs. During aberrant lung alveolarization that occurred under hyperoxic conditions, collagen structures in Tgm2 mice were partially protected from the impact of hyperoxia, where normal dihydroxylysinonorleucine and hydroxylysylpiridinoline collagen cross-link abundance was restored; however, the lung alveolar architecture remained abnormal. Inhibition of transglutaminases (including TGM2) with cysteamine appreciably reduced transglutaminase activity in vivo, as assessed by N -(γ-l-glutamyl)-l-lysine abundance and TGM catalytic activity, and restored normal dihydroxylysinonorleucine and hydroxylysylpiridinoline collagen cross-link abundance under pathological conditions. Furthermore, a moderate improvement in alveoli size and gas-exchange surface density was noted in cysteamine-treated mouse lungs in which BPD was modelled. These data indicate that TGM2 plays a role in normal lung alveolarization, and contributes to the formation of aberrant ECM structures during disordered lung alveolarization.

Carnosinase 1 (CN1) is encoded by the Cndp1 gene and degrades carnosine and anserine, two natural histidine-containing dipeptides. In vitro and in vivo studies suggest carnosine- and anserine-mediated protection against long-term sequelae of reactive metabolites accumulating, e.g., in diabetes mellitus. We have characterized the metabolic impact of CN1 in 11- and 55-week-old Cndp1-knockout (Cndp1-KO) mice and litter-matched wildtypes (WT). In Cndp1-KO mice, renal carnosine and anserine concentrations were gender-specifically increased 2- to 9-fold, respectively in the kidney and both most abundant in the renal cortex, but remained unchanged in all other organs and in serum. Renal oxidized/reduced glutathione concentrations, renal morphology and function were unaltered. In Cndp1-KO mice at week 11, renal asparagine, serine and glutamine levels and at week 55, renal arginine concentration were reduced. Renal heat-shock-protein 70 (Hspa1a/b) mRNA declined with age in WT but not in Cndp1-KO mice, transcription factor heat-shock-factor 1 was higher in 55-week-old KO mice. Fasting blood glucose concentrations decreased with age in WT mice, but were unchanged in Cndp1-KO mice. Blood glucose response to intraperitoneal insulin was gender- but not genotype-dependent, the response to intraperitoneal glucose injection was similar in all groups. A global Cndp1-KO selectively, age- and gender-specifically, increases renal carnosine and anserine concentrations, alters renal amino acid- and HSP70 profile and modifies systemic glucose homeostasis. Increase of the natural occurring carnosine and anserine levels in the kidney by modulation of CN1 represents a promising therapeutic approach to mitigate or prevent chronic kidney diseases such as diabetic nephropathy.