Morphine is the most common drug of choice in clinical pain management; however, morphine tolerance presents a significant clinical challenge. The pathogenesis of morphine tolerance is known to be closely associated with angiotensin II receptor type 1 (AT1R) in microglia. As an AT1R antagonist, candesartan may serve an important role in regulating morphine tolerance. Therefore, the present study aimed to investigate the role of candesartan in morphine tolerance, and to explore the underlying mechanism. To meet this aim, BV2 microglial cells were treated with morphine or candesartan alone, or as a combination, and the expression levels of AT1R in BV2 cells were detected by reverse transcription-quantitative PCR (RT-qPCR) and western blotting. The levels of the inflammatory cytokines tumor necrosis factor-α, interleukin (IL)-1β and IL-6 were subsequently detected by ELISA and western blotting. In addition, immunofluorescence analysis, western blotting and RT-qPCR were used to detect the expression levels of the BV2 cell activation marker, ionized calcium-binding adaptor molecule 1 (IBA-1). Western blotting was also used to detect the expression levels of peroxisome proliferator-activated receptor-γ/AMP-activated protein kinase (PPARγ/AMPK) signaling pathway-associated proteins. Finally, the cells were treated with the PPARγ antagonist GW9662 and the AMPK inhibitor compound C to further explore the mechanism underlying the effects of candesartan on improving morphine tolerance. The expression levels of AT1R were revealed to be significantly increased following morphine induction; however, candesartan treatment inhibited the expression levels of AT1R, the levels of inflammatory cytokines and the protein expression levels of IBA-1 in morphine-induced BV2 cells in a dose-dependent manner. These processes may be associated with activation of the PPARγ/AMPK signaling pathway. Taken together, the present study revealed that treatment with candesartan reduced morphine-induced inflammatory response and cellular activation of BV2 cells via PPARγ/AMPK signaling.

Doxorubicin (DOX) is an efficacious antineoplastic drug; however, its use is limited due to its cardiotoxicity. Cardiomyocyte senescence is considered to be a key factor in the development of DOX-related cardiomyopathy. Complement component 5a (C5a) and the C5a receptor (C5aR) have been reported to play a key role in the process of cellular senescence. However, to the best of our knowledge, the exact role of C5a and C5aR in cellular senescence in the heart remains largely unknown. Reverse transcription-quantitative (RT-q)PCR and western blot assays were used to analyze the expression levels of C5a and C5aR in H9c2 embryonic rat cardiomyocytes and AC16 human cardiomyocyte-like cells. The cells were treated with DOX and a C5aR antagonist (C5aRA). The expression of TNF-α and IFN-γ was determined using ELISA and western blotting. The levels of reactive oxygen species (ROS) were also measured using ELISA. Cellular senescence was determined using senescence-associated β-galactosidase (SA-β-gal) staining and by analyzing the protein expression levels of p53, p16, p21 and insulin-like growth factor-binding protein 3 (IGFBP3). The expression levels of C5a and C5aR were found to be upregulated during the DOX-induced senescence of H9c2 and AC16 cardiomyocytes. Treatment with C5aRA downregulated TNF-α and IFN-γ expression, in addition to ROS levels. Furthermore, C5aRA prevented DOX-induced cellular senescence and decreased the levels of positive SA-β-gal staining in H9c2 and AC16 cardiomyocytes, in addition to downregulating the expression levels of p53, p16, p21 and IGFBP3. C5aRA also increased the telomere length and telomerase activity in H9c2 and AC16 cardiomyocytes following DOX stimulation. In conclusion, the findings of the present study indicated that C5a and C5aR may play a key role in cardiomyocyte senescence, and treatment with C5aRA may be an effective method for preventing DOX-induced cardiomyocyte aging.