Diabetic nephropathy (DN) is a common microvascular complication of diabetes mellitus (DM). Several mechanisms have been proposed for DN pathogenesis; however, it remains a progressive risk for DM patients. For the first time, the current study aimed to investigate the nephroprotective effects of agmatine and pioglitazone through modulating α-klotho, anti-ERS, anti-inflammatory, and antipyroptotic properties in ameliorating DN. A single intraperitoneal injection of streptozotocin (STZ) (52.5 mg/kg), preceded by nicotinamide (NA) (50 mg/kg, i.p.), induced DN. NA provided partial protection to insulin-secreting β-cells against the severe damage caused by STZ. Agmatine (100 mg/kg, i.p.) was injected daily for 6 weeks after modeling DN, either alone or in combination with pioglitazone (5 mg/kg, p.o.). DN rats showed a marked decrease in blood glucose (BG) levels, body weight, serum urea levels, serum creatinine (Cr) levels, and histopathological alterations, besides reducing renal α-klotho content associated with upregulating ATF4/IRE1α/NLRP3 renal content along with increasing HMGB1/NF-κB protein expression. On the contrary, agmatine and pioglitazone improved animals' body weights, BG, serum urea, and Cr levels. Moreover, agmatine and pioglitazone inhibited renal NLRP3 content, reducing oxidative stress and inflammatory cascades. They also upregulated renoprotective α-klotho and decreased renal ATF4/IRE1α/TXNIP/NLRP3 content associated with a decrease in renal HMGB1/NF-κB protein expression. Briefly, besides its antidiabetic effect, agmatine, along with pioglitazone, shows promise as a treatment for DM and DN via mediating α-klotho/ERS/HMGB1/NF-κB/NLRP3 inflammasome signaling pathways.