[Human urine-derived stem cell-derived exosomes improve diabetic erectile dysfunction in rats].

Objective: To investigate the improving effect of human urine-derived stem cell-derived exosomes (USC-Exo) on the endothelial function and erectile function of male rats with diabetic ED (DED) and explore their action mechanism. METHODS: USC-Exo were extracted from the culture medium of USC by ultracentrifugation and identified. Cavernous sinus endothelial cells (CCEC) were collected from SD male rats and cultured in endothelial cell growth medium-2 (EGM-2) (the normal control group), EGM-2 + L-glucose at 25 mM (the high glucose group), EGM-2 + L-glucose at 25 mmol/L) + USC-Exo at 10 µg/ml (the Exo group), and EGM-2 + L-glucose at 25 mmol/L + USC-Exo at 10 µg/ml) + 3-methyladenine at 2 mmol/L (the 3-MA group), respectively. Changes of the autophagic flux in the CCECs transfected with mRFP-GFP-LC3 adenovirus were detected under the fluorescence microscope. The proliferation and tube-forming ability of the cells were assessed by CCK8 and Matrigel assays, respectively. DED was induced by intraperitoneal injection of streptozotocin in 10 of the rats, which were equally and randomly divided into a DED and an Exo group, and another 5 normal male rats were taken as controls. The rats in the normal and DED groups were injected intracavernously with 100 µl of PBS, and those in the Exo group with 100 µl of USC-Exo at the concentration of 1 µg/µl. Four weeks after treatment, the maximum intracavernous pressure (ICPmax) and mean arterial pressure (MAP) were measured, the endothelial marker CD31 detected by immunofluorescence assay, the expressions of the CD31, Beclin1 and LC3 I/II proteins examined by Western blot, and the number of autophagosomes in the cavernous endothelial cells determined under the transmission electron microscope. RESULTS: USC-Exo significantly increased the number of autophagosomes in the CCEC in the high glucose group compared with that in the normal controls (39.5 ± 6.2 vs 12.5 ± 5.4, P < 0.05). The expression of Beclin1 and proliferation of the CCEC were significantly higher in the Exo than in the high glucose group (P < 0.05). The autophagy inhibitor 3-MA evidently reversed the increasing effect of USC-Exo on the proliferation of the CCEC. The tube-forming ability of the CCEC was significantly increased in the Exo group compared with that in the high glucose group (15.3 ± 3.2 vs 6.3 ± 2.1, P < 0.05), which was also reversed in the 3-MA group. Both ICPmax and the ICPmax/MAP ratio were significantly higher in the Exo than in the DED group (ï¼»86.6 ± 12.6ï¼½ vs ï¼»37.9 ± 10.9ï¼½ mmHg, P < 0.05; 89.3 ± 14.1 vs 41.7 ± 11.5, P < 0.05), and so were the expressions of CD31, Beclin1 and LC3 I/II (P< 0.05) and the number of autophagosomes in the cavernosal endothelial cells (3.7 ± 0.6 vs 1.0 ± 1.0, P < 0.05). CONCLUSIONS: USC-Exo can significantly improve the endothelial and erectile functions of DED rats by increasing the autophagy of cavernosal endothelial cells.

[Protective effect of urine-derived stem cells on erectile dysfunction in rats with cavernous nerve injury].

OBJECTIVE: To investigate the protective effect of human urine-derived stem cells (USCs) on erectile function and cavernous structure in rats with cavernous nerve injury (CNI). METHODS: Sixty adult male SD rats with normal sexual function were randomly divided into four groups of equal number: sham operation, bilateral CNI (BCNI) model control, phosphate buffered saline (PBS), and USC. The BCNI model was established in the latter three groups of rats by clamping the bilateral cavernous nerves. After modeling, the rats in the PBS and USC groups were treated by intracavernous injection of PBS at 200 µl and USCs at 1×106/200 µl PBS respectively for 28 days. Then, the maximum intracavernous pressure (mICP) and the ratio of mICP to mean arterial pressure (mICP/MAP) of the rats were calculated by electrical stimulation of the major pelvic ganglions, the proportion of nNOS- or NF200-positive nerve fibers in the total area of penile dorsal nerves determined by immunohistochemical staining, the levels of endothelial cell marker eNOS, smooth muscle marker α-SMA and collagen I detected by Western blot, and the smooth muscle to collagen ratio and the cell apoptosis rate in the corpus cavernosum measured by Masson staining and TUNEL, respectively. RESULTS: After 28 days of treatment, the rats in the USC group, as compared with those in the PBS and BCNI model control groups, showed significant increases in the mICP (ï¼»81 ± 9.9ï¼½ vs ï¼»31 ± 8.3ï¼½ and ï¼»33 ± 4.2ï¼½ mmHg, P <0.05), mICP/MAP ratio (0.72 ± 0.05 vs 0.36 ± 0.03 and 0.35 ± 0.04, P <0.05), the proportions of nNOS-positive nerve fibers (ï¼»11.31 ± 4.22ï¼½% vs ï¼»6.86 ± 3.08ï¼½% and ï¼»7.29 ± 4.84ï¼½% , P <0.05) and NF200-positive nerve fibers in the total area of penile dorsal nerves (ï¼»27.31 ± 3.12ï¼½% vs ï¼»17.38 ± 2.87ï¼½% and ï¼»19.49 ± 4.92ï¼½%, P <0.05), the eNOS/GAPDH ratio (0.52 ± 0.08 vs 0.31 ± 0.06 and 0.33 ± 0.07, P <0.05), and the α-SMA/GAPDH ratio (1.01 ± 0.09 vs 0.36 ± 0.05 and 0.38 ± 0.04, P <0.05), but a remarkable decrease in the collagen I/GAPDH ratio (0.28 ± 0.06 vs 0.68 ± 0.04 and 0.70 ± 0.10, P <0.05). The ratio of smooth muscle to collagen in the corpus cavernosum was significantly higher in the USC than in the PBS and BCNI model control groups (17.91 ± 2.86 vs 7.70 ± 3.12 and 8.21 ± 3.83, P <0.05) while the rate of cell apoptosis markedly lower in the former than in the latter two (3.31 ± 0.83 vs 9.82 ± 0.76, P <0.01; 3.31 ± 0.83 vs 9.75 ± 0.91, P <0.05). CONCLUSIONS: Intracavernous injection of USCs can protect the erectile function of the rat with cavernous nerve injury by protecting the nerves, improving the endothelial function, alleviating fibrosis and inhibiting cell apoptosis in the cavernous tissue.

Alda-1, an ALDH2 activator, protects against hepatic ischemia/reperfusion injury in rats via inhibition of oxidative stress.

Previous studies have proved that activation of aldehyde dehydrogenase two (ALDH2) can attenuate oxidative stress through clearance of cytotoxic aldehydes, and can protect against cardiac, cerebral, and lung ischemia/reperfusion (I/R) injuries. In this study, we investigated the effects of the ALDH2 activator Alda-1 on hepatic I/R injury. Partial warm ischemia was performed in the left and middle hepatic lobes of Sprague-Dawley rats for 1 h, followed by 6 h of reperfusion. Rats received either Alda-1 or vehicle by intravenous injection 30 min before ischemia. Blood and tissue samples of the rats were collected after 6-h reperfusion. Histological injury, proinflammatory cytokines, reactive oxygen species (ROS), cellular apoptosis, ALDH2 expression and activity, 4-hydroxy-trans-2-nonenal (4-HNE) and malondialdehyde (MDA) were measured. BRL-3A hepatocytes were subjected to hypoxia/reoxygenation (H/R). Cell viability, ROS, and mitochondrial membrane potential were determined. Pretreatment with Alda-1 significantly alleviated I/R-induced elevations of alanine aminotransferase and aspartate amino transferase, and significantly blunted the pathological injury of the liver. Moreover, Alda-1 significantly inhibited ROS and proinflammatory cytokines production, 4-HNE and MDA accumulation, and apoptosis. Increased ALDH2 activity was found after Alda-1 administration. No significant changes in ALDH2 expression were observed after I/R. ROS was also higher in H/R cells than in control cells, which was aggravated upon treatment with 4-HNE, and reduced by Alda-1 treatment. Cell viability and mitochondrial membrane potential were inhibited in H/R cells, which was attenuated upon Alda-1 treatment. Activation of ALDH2 by Alda-1 attenuates hepatic I/R injury via clearance of cytotoxic aldehydes.