Fig 1: Effect of PSE-NPs on the GnRhR signaling pathway and expression of meiosis-related proteins SCP3 of GC-1 cells damaged by adriamycin: (A–E) Protein expression of GnRhR, GNAS, p-CREB and SCP3 in GC-1 cells, scale bar 50 µm.
Fig 2: GnRHR expression and cellular uptake of [d-Lys6(FITC)]–GnRH-I on EBC-1 cells. (A) Expression of GnRHR in EBC-1 cells, confirmed by western blot; (B) EBC-1 cells contain a high level of GnRHR (total GnRHR) and a high proportion of these receptors are located in the plasma membrane (membrane GnRHR). Nuclei: blue (DRAQ5), GnRHR: red (Alexa Fluor 594); (C) Time- and concentration-dependent dynamic uptake of [d-Lys6(FITC)]–GnRH-I was confirmed by flow cytometry. Results are presented as mean relative fluorescent intensity ± SD (N = 3); (D) Intracellular localization of [d-Lys6(FITC)]–GnRH-I (green; 10 µM, 24 h) in EBC-1 cells was confirmed by confocal microscopy (nuclei: blue).
Fig 3: Central mechanism in miR-29a/b1 KO mice. (A, B) GnRHR immunoreactivity in pituitary of wild-type and miR-29a/b1 KO females. The receptor was not detectable on the plasma membrane of control. (immunohistochemical: wild-type: 13.43 ± 0.7927, miR-29a/b1 KO: 25.47 ± 0.534, p=0.0249; immunofluorescence: wild-type: 37.79 ± 1.858, miR-29a/b1 KO: 56.64 ± 2.767, p=0.0045, n=3). (C) Serum LH and FSH levels in miR-29a/b1 KO females and controls following ovariectomy (OVX) and sham-operated controls (Sham). (LH: wild-type: p=0.0401, miR-29a/b1 KO: p=0.9249; FSH: wild-type: p=0.0016, miR-29a/b1 KO: p=0.0185, n=6). (D) Expression of Kiss1 and Gnrh1 in hypothalamus (Gnrh1: wild-type: 1 ± 0.1912, miR-29a/b1 KO: 0.7287 ± 0.06234, p=0.1874, Kiss1: wild-type: 1 ± 0.1305, miR-29a/b1 KO: 0.8142 ± 0.0757, p=0.8405, n=15). (E) Normal distribution of GnRH neurons in miR-29a/b1 KO mice compared to control littermates. OVLT, organum vasculosum of the lamina terminalis. Scale bars, 200µm. (wild-type: 9.827 ± 1.547, miR-29a/b1 KO: 8.597 ± 0.8466, p=0.5238, n=3). *p < 0.05, **p < 0.01 and ****p < 0.0001.
Fig 4: Effects of PSE-NPs on sex-hormone levels, GnRh signaling pathway-related proteins in testes, and indices of meiosis in mice: (A) Levels of GnRh, LH and testosterone in the serum of mice in each group. (B) mRNA expression of GnRh signaling pathway-related proteins and meiosis-associated genes in the testicular tissue of mice in each group was measured by RT-qPCR. (C) Protein expression of GnRh signaling pathway and meiosis-associated proteins in the testicular tissue of mice in each group was measured by western blotting. (D) The protein expression of GnRhR and SCP3 in the testicular tissue of mice in each group was measured by Immunofluorescence, scale bar 50 µm.
Fig 5: Intracellular trafficking of crizotinib* and [d-Lys6(crizotinib*)]–GnRH-I. (A) Crizotinib is able to reach the ATP-binding site of c-Met by passive diffusion and to effectively inhibit the c-Met-mediated oncogenic signaling, while only a proportion of the drug accumulates in the lysosomes. (B) The GnRHR-mediated endocytosis leads to the lysosomal sequestration of GnRHR-targeted crizotinib* at the nanomolar concentration range; meanwhile, crizotinib* bypasses c-Met. (C) One possible explanation for the c-Met inhibition and the cell viability inhibition effect of [d-Lys6(crizotinib*)]–GnRH-I at the micromolar concentration range is that during the GnRHR internalization, non–GnRHR-bound conjugates might loaded into the vesicles as “passive passengers”. High lysosomal accumulation of these passive passengers might destabilize the membrane of the lysosomes, leading to drug leakage and cell death.
Supplier Page from Proteintech Group Inc for GNRHR antibody