Plasma-derived from human umbilical cord blood restores ovarian function and improves serum reproductive hormones levels in mice with premature ovarian insufficiency (POI) through cytokines and growth factors.

Premature ovarian insufficiency (POI) patients experience a decline in ovarian function and a reduction in serum reproductive hormones, leading to a significant impact on the outcomes of assisted reproductive technology. Despite the absence of an effective clinical treatment to restore fertility in POI patients, recent research has indicated that cord blood plasma (CBP) derived from human umbilical cord blood (hUCB) may offer therapeutic benefits for various degenerative diseases. The primary aim of this study is to explore approaches for enhancing ovarian function and serum reproductive hormones through the administration of CBP in a murine model. Initially, hUCB was utilized to obtain CBP (CBP), which was subsequently analyzed for cytokine and growth factor profiles in comparison to adult blood plasma (ABP) by use of flow cytometry. Subsequently, POI mouse models were established through the induction of 4-vinylcyclohexene diepoxide, followed by the injection of CBP into the tail. At 7, 14, and 21 days posttreatment, mouse ovaries and blood were collected, and their estrus cycle, body weight, and ovarian weights were evaluated using precise electronic balance. Finally, ovarian morphology and follicle number were assessed through HE staining, while serum levels of anti-Müllerian hormone (AMH), estradiol (E2) and follicle-stimulating hormone (FSH) were determined by ELISA. Our study revealed that individuals with CBP exhibited significantly lower concentrations of proinflammatory cytokines, including IL-ß (p < 0.01) and IL-2 (p < 0.05), while displaying elevated levels of anti-inflammatory cytokines and chemokines, such as IL-2, IL-4, IL-6, IL-8, IL-12P70, IL-17A, IP-10, interferon-γ, and tumor necrosis factor-α (p < 0.01). Furthermore, CBP demonstrated remarkably higher levels of growth factors, including transforming growth factor-ß1, vascular endothelial growth factor, and insulin-like growth factor-1 (p < 0.01) than ABP. Notably, our investigation also revealed that CBP restored the content of serum reproductive hormones, such as AMH, E2, and FSH (p < 0.05), and increased the number of primordial and primary follicles (p < 0.01) and decreased the number of luteal and atretic follicles (p < 0.01) in vivo. Our findings suggested that CBP-secreted cytokines and growth factors could be restored POI ovarian function, enhanced serum reproductive hormones and rescued follicular development in vivo. These findings further support the potential of CBP as a promising strategy in clinical applications for POI related infertility.

Ischemic injury underlies the pathogenesis of aristolochic acid-induced acute kidney injury.

Aristolochic acid nephropathy (AAN) is a progressive tubulointerstitial renal disease caused by aristolochic acid intake. To determine the contribution of renal ischemia to the pathogenesis of AAN, we characterized changes in the expression of angiogenic factors and vasoactive substances, and then we evaluated the expression of a marker of hypoxia in an acute AAN rat model. Rats were orally administrated either a decoction of Aristolochiae manshuriensis that contained 20 mg/kg of aristolochic acid-I or an equal volume of distilled water (control group) once daily for 4 days or 7 days. Renal histology and serum creatinine were assessed. Expression of endothelin-1 (ET-1) and hypoxia inducible factor-1 alpha (HIF-1alpha) mRNA within renal cortex were determined by semiquantitative reverse-transcription polymerase chain reaction. Levels of ET-1, nitric oxide (NO), vascular endothelial growth factor (VEGF), and HIF-1alpha in kidneys were determined by radioimmunoassay, Griess method, Western blot, and immunohistochemistry, respectively. Tubular injury scores and ET-1 mRNA expression were increased in the AA-treated rats at both days 4 and 8, whereas serum creatinine level and ET-1 protein expression was increased only at day 4. In contrast, NO production in AA-treated rats was decreased at day 8 compared with the control group. Similarly, VEGF protein expression was reduced in the AA-treated rats at both days 4 and 8. A dramatic increase in nuclear staining for HIF-1alpha was observed mainly in the tubular cells of tubulointerstitial damage area in the AA-treated rats at day 8. The observed increase in HIF-1alpha protein expression, decrease in VEGF protein expression, and imbalance of vasoactive substances after induction of acute kidney injury by AA suggests that ischemic injury contributes to the pathogenesis of AAN.

Cytotoxicity of phenanthrenes extracted from Aristolochia contorta in human proximal tubular epithelial cell line.

BACKGROUND/AIMS: Aristolochic acid nephropathy, a progressive tubulointerstitial renal disease, is predominantly a result of aristolochic acid I (AA-I) intoxication. However, other unidentified phytotoxins have indeed been postulated as the cause of this unique interstitial nephropathy. The purpose of this study was to investigate the cytotoxicity of other phenanthrene derivatives extracted from Aristolochia contorta in the human proximal tubular epithelial cell line HK-2. METHODS: After HK-2 cells were incubated with an indicated concentration of test compounds for 24 h, cell viability was assessed by lactate dehydrogenase (LDH) leakage assay (cell membrane damage) in combination with MTT assay (metabolic capability). Cellular morphologic assessments were performed with a phase-contrast inverted microscope and transmission electron microscope. RESULTS: In all test compounds at 5 microg/ml, AA-I, 7-methoxy-aristololactam IV and aristololactam IVa showed cytotoxic activity in HK-2 cells in both MTT assay and LDH leakage assay (p < 0.01). At high concentration (5-80 microg/ml), these three compounds caused a dose-dependent decrease in MTT reduction and a dose-dependent increase in LDH leakage compared to non-treated cells (p <0.01). In LDH leakage assay, 40 mug/ml 7-methoxy-aristololactam IV induced a 1.58-fold LDH leakage compared to AA-I at the same concentration (p < 0.01). Moreover, the IC50 of these three compounds were 16.675 microg/ml for AA-I, 4.535 microg/ml for 7-methoxy-aristololactam IV, and 30.244 microg/ml for aristololactam IVa in MTT assay. The cellular morphologic assessments suggest interactions with cell membrane and intracellular structures such as lysosome and mitochondria are likely to be involved in cell injury induced by these three compounds. CONCLUSION: The potency of cytotoxic activity of aristololactam IVa and 7-methoxy-aristololactam IV extracted from A. contorta is similar to or even stronger than that of AA-I.