Fig 2: Spike increases Rab5a association with ACE2 and junctional proteins. (A) Representative Western blot of coimmunoprecipitation experiments done using Rab5a antibody and probed for different proteins. (B) Mean data indicating fold change in Rab5a association with junctional proteins and ACE2 after S1RBD treatment. n = 6 for each, *P < 0.05 vs. untreated control. (C) Representative Western blot of coimmunoprecipitation experiments done using ACE2 antibody and probed for Rab5a. (D) Mean data indicating fold change in ACE2 association with Rab5a after Spike treatment. n = 6 for each, *P < 0.05 vs. untreated control. (E) Representative Western blot of coimmunoprecipitation experiments in control and diabetic cells after Rab5a knockdown with siRNA. (F) Mean data indicating fold change in ACE2 association with Rab5a after Rab5a siRNA. n = 6 for each, *P < 0.05 vs. control, #P < 0.05 vs. NT siRNA diabetic.

Fig 3: Expression patterns of TMEM206 and ClC-7 in primary bone marrow-derived macrophages.(a) Endogenous TMEM206 co-localizes with early endosomal rab5 and partially with late endosomal rab7. Pearson’s R calculated from 19 cells (Rab5), 27 cells (Rab7) from 2 independent experiments. (b) Expression of human TMEM206 C-terminally tagged with GFP in Tmem206−/− primary macrophages gives similar expression pattern as endogenous TMEM206 protein. (c) ClC-7 co-localizes with late endosome marker rab7. Pearson’s R calculated from 26 cells from 2 independent experiments. Mean ± s.e.m. Scale bars: 10 µm, 1 µm for enlargements. Source numerical data are available in source data. Source data

Fig 4: Remimazolam regulate the cell surface expression of TLR4 after LPS treatment by affecting functions and expressions of RAB5a. (A) (RAW264.7 cells), (C) (BMDM) Western blot analysis of Rab5a expression. (B–D) Protein presence of Rab5a was normalized to GAPDH. (E) Immune-staining of TLR4 with Rab5a in RAW264.7 cells (up panel; Scale bar, 100 μm). Each bar represents the mean ± SD, *p < 0.05, compared with control group; #p < 0.05, compared with LPS group.

Fig 5: SARS-CoV-2 enters into ECs through endocytosis. (A) Representative electron microscopy images (× 80,000) for senescent HUVECs infected with SARS-CoV-2 at 50 MOI (2 hpi). Arrows indicate SARS-CoV-2 virions. Bars: 200 nm. (B) Representative electron microscopy images (× 80,000) for early passage and senescent ECs infected with SARS-CoV-2 at 1 MOI (6 hpi). Arrows indicate SARS-CoV-2 entry via caveolae-like structures. Bars: 200 nm. (C) Representative electron microscopy images (× 80,000) for early passage and senescent ECs infected with SARS-CoV-2 at 1 MOI (24 and 48 hpi). Arrows indicate SARS-CoV-2 virions. Bars: 200 nm. (D) Representative images of immunocytochemistry for SARS-CoV-2 spike protein (red fluorescence) and Rab5A (green fluorescence) in replicative senescent HUVECs infected with SARS-CoV-2 (50 MOI) at 6 hpi. SARS-CoV-2 spike proteins were largely co-localized with Rab5A. Bars: 100 μm. Spike and Rab5A double-positive areas were quantitatively analyzed (n = 6 each). Statistical analysis was performed using Mann Whitney U-test. Data are presented as mean ± SE **P < 0.01.