Abstract
BackgroundThis study was designed to determine the effect of human umbilical cord multipotent mesenchymal stromal cells (hUC-MSC) on acute ischemia/reperfusion (I/R) injury of spermatogenic cells.MethodThe testicular I/R rat model was established through 720° torsion for 1 h. hUC-MSC were intravenously injected 10 min before detorsion. Injury severity of spermatogenic cells was estimated by Johnsen’s score. The proliferating of recipient spermatogonia was measured by the immunostaining of antibodies against Ki67, and all germ cells were detected with DDX4 antibody. And recipient spermatogenesis was assessed by staining spermatozoa with lectin PNA. The levels of inflammatory factors were measured by real-time PCR. And the Selectin-E expression, neutrophil infiltration in the testes was detected by immunostaining. Germ cells apoptosis was tested by TUNEL assay and western blot. Furthermore, the oxidative stress was tested by reactive oxidative species (ROS) levels. In vitro, the condition medium (CM) of hUC-MSC was used to culture human umbilical vein endothelial cells (HUVECs), so as to assess the paracrine effect of hUC-MSC on HUVECs. The protein chip was used to measure the relative concentration of the secretory proteins in the CM of hUC-MSC.ResulthUC-MSC greatly alleviated the testicular injury induced by testis I/R. The levels of proinflammatory factors were downregulated by hUC-MSC in vivo and in vitro. Neutrophil infiltration, ROS, and germ cell apoptosis in testicular tissues were greatly reduced in the group of hUC-MSC. Paracrine factors secreted by hUC-MSC including growth factors, cytokines, and anti-inflammatory cytokine were rich.ConclusionThis study demonstrated that intravenously injected hUC-MSC could protect the spermatogenic cells against I/R injury by reducing the inflammatory response, apoptosis, and acute oxidative injury. Paracrine mechanism of hUC-MSC may contribute to the protection of spermatogenic cells against I/R injury. Therefore, the present study provides a method for clinical treatment of attenuate I/R injury of spermatogenic cells.
Highlights
This study was designed to determine the effect of human umbilical cord multipotent mesenchymal stromal cells on acute ischemia/reperfusion (I/R) injury of spermatogenic cells
To further determine if human umbilical cord multipotent mesenchymal stromal cells (hUC-MSC) could protect recipient spermatogenesis, we analyzed recipient spermatogenesis by staining spermatozoa with lectin PNA conjugated with Alexa FluorTM 594, which selectively binds to acrosome of sperms and can be used as a sperm or haploid spermatid marker [20]
We found that hUC-MSC injected intravenously into rats subjected to testicular torsion and detorsion operation could attenuate I/R injury and promote the proliferation and differentiation of spermatogonia cells resulting in the survival and the regeneration of more spermatogenic cells and sperms (Fig. 1)
Summary
This study was designed to determine the effect of human umbilical cord multipotent mesenchymal stromal cells (hUC-MSC) on acute ischemia/reperfusion (I/R) injury of spermatogenic cells. Testicular torsion is a common type of urological emergency in adolescent males which usually leads to severe acute ischemia injury of the testis. Previous studies suggest that testicular torsion leads to male infertility because of testicular atrophy [1,2,3]. Detorsion operation itself leads to ischemia/reperfusion (I/R) injury when the blood flow returns to normal. Many cellular and molecular mechanisms are investigated in testicular I/R injury. Numerous pharmacological agents have been used as supportive therapy to prevent the adverse effects of I/R injury, including antioxidants, hormones, vitamins, and plant extracts [8]. Most of the agents only targeted to eliminate ROS which is less efficient
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