Preventive carnosine supplementation attenuates painful diabetic neuropathy via restoring mitochondrial function and suppressing inflammasome activation

Figure 1 The rat model of painful diabetic neuropathy established through high-fat feeding combined with low-dose streptozotocin injection. ^aP < 0.05, ^bP < 0.01, ^cP < 0.001 vs the control group; two-way analysis of variance with Sidak’s post-hoc test for Figure 1A and B; Unpaired t-tests, two-tailed for Figure 1C, I, K-M; Linear mixed-effects models were applied for two-group comparisons of data in panels F-H, with animal ID set as a random intercept to account for clustered observations within individual rats; Figure 1A-C: n = 11; Figure 1D-H: n = 3 rats per group, with at least 50 mitochondria counted in 3 random fields per sample. For Figure 1F and G, each field was the statistical unit; for panel H, each individual mitochondrion was the statistical unit for mitochondrial length quantification; I: n = 8; J-M: n = 6. A: Paw withdrawal threshold in painful diabetic neuropathy (PDN) rats; B: Paw withdrawal latency in PDN rats; C: Motor nerve conduction velocity and sensory nerve conduction velocity in PDN rats; D: Ultrastructural analysis of nerve fibers in the dorsal root ganglion (DRG) of PDN rats. Scale bar = 2 μm; E: Examination of mitochondrial structure in the DRG of PDN rats. Scale bar = 500 nm; F: Measurement of the number of mitochondria per field; G: Proportion of abnormal mitochondria to total mitochondria; H: Measurement of mitochondrial length; I: Assessment of mitochondrial membrane potential in the DRG of PDN rats, in which quantitative data were acquired by a fluorescence microplate reader; J-M: Western blotting showing the expression level of proteins associated with mitochondrial fission protein dynamin-related protein 1 and fusion proteins MFN1/MFN2, including representative electrophoresis bands and relative expression levels. PDN: Painful diabetic neuropathy; MNCV: Motor nerve conduction velocity; SNCV: Sensory nerve conduction velocity; STZ: Streptozotocin; Drp1: Dynamin-related protein 1.