| Background:Mesenchymal stem cells(MSC)have been reported to promote regeneration in both acute subjects with acute kidney injury(AKI)and chronic kidney disease(CKD),but their efficacy remains limited,probably because most of the cells accumulate in the lungs,liver,and spleen after an intravenous infusion.Therefore,ultrasound-guided administration of MSC represents a possible approach to solve this problem.The greater omentum is used to promote cell survival due to its rich vasculature.We hypothesized that ultrasound-guided administration of MSC combined with greater omentum might be more curative than currently available approaches.Methods:In this study,we established an aristolochic acid nephropathy(AAN)model by intraperitoneally administering aristolochic acid I sodium salt(AA-I)at a dose of 5 mg/kg body weight on alternate days for 4 weeks.Regular observation under a light microscope was used to determine whether the cultured red fluorescent protein(RFP-MSC)was contaminated by bacteria or fungi.The method of detection with mycoplasma detection kit and fluorescence microscope observation is used to determine whether the cultured cells are contaminated with mycoplasma.With the help of intravital imaging technology to determine the appropriate single infusion dose of MSC,the number of days the cells can survive between the omentum and the left kidney after infusion,so as to determine the frequency of cell infusion.Subsequently,a laparotomy was performed,and the left kidney from which the capsule had been removed was wrapped with the greater omentum.A dose of 2×10~7MSC was injected into the space between the greater omentum and the left kidney.Equal amounts of MSC were the administered under ultrasound guidance every week for a total of 4 treatments.The body weight of the mice was measured once a week for 8 consecutive weeks to monitor body weight,and blood was taken from the tail vein to measure serum creatinine and blood urea levels to assess renal function.Mice were sacrificed 4 weeks after surgery.Histopathological staining of Periodic Acid–Schiff(PAS)staining,Sirius red staining and Masson,q PCR,Western blotting techniques were used to observe and analyze the renal damage and fibrosis of AAN.Using q PCR,western blot,immunohistochemistry,immunofluorescence technology,transwell cell migration test,dual-light imaging technology to study the anti-fibrosis and anti-inflammatory mechanism of MSC and preliminary exploration of its efficacy through paracrine or cell differentiationResults:The AAN model was successfully established.The cultured cells are healthy and pollution-free.In vivo imaging determined that the appropriate infusion dose of cells was2×10~7/mouse/time.Ultrasound-guided injection of MSC into the greater omentum that surrounds the kidney enriched cells in the kidney region for at least 5 days.The best frequency is one infusion per week.Renal function tests indicated that MSC improved renal function to a great extent,as reflected by decreased blood urea nitrogen and serum creatinine levels.Histological analyses showed that MSC noticeably attenuated kidney injury,as evidenced by the amelioration of tubular necrosis and peritubular interstitial fibrosis.Mitigation of renal interstitial fibrosis was further confirmed by immunohistochemistry,q PCR and western blotting after MSC treatment.Moreover,immunofluorescence staining revealed that MSC alleviated inflammatory responses by increasing the counts of CD206+cells and decreasing the counts of CD68+cells.MSC migration was initiated in response to AA-I-treated renal epithelial cells in an in vitro migration assay.Two-photon microscopy showed that the infused cells did not enter the kidney parenchyma.Conclusions:These findings suggested that administration of MSC into the cavity formed by the injured kidney and the greater omentum under ultrasound guidance improved renal function,attenuated kidney injury,and mitigated renal interstitial fibrosis and inflammatory responses.And the way that MSC exerts its effect is through paracrine function.Thus,local delivery of MSC might be a safe and effective therapy for CKD. |
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