Journal: AIDS (London, England)

Article Title: HIV protease inhibitor ritonavir induces renal fibrosis and dysfunction: role of platelet-derived TGF-β1 and intervention via anti-oxidant pathways

doi: 10.1097/QAD.0000000000002516

Figure Lengend Snippet: Pharmacologic levels of the HIV protease RTV induce renal pathology, including fibrosis. Higher levels of renal fibrosis are seen in RTV-exposed wt vs. platelet TGF- β1 deficiency PF4CreTgfb1flox/flox mice, and this can be suppressed by inhaled carbon monoxide. Wt and PF4CreTgfb1flox/flox mice were treated with RTV (10mg/kg) or vehicle (DMSO) for 8 wks. Kidneys were harvested and 4μM sections evaluated for fibrosis by staining with (A) H&E, (B) PAS, and (C) Picrosirius red, and quantification by Aperio whole slide scanning and Aperio image analysis, as described in the legend to Suppl. Fig. 1. In addition, independent evaluation of the extent of fibrosis was performed by four trained observers (E). Widening of Bowman’s spaces (A) with globular eosinophilic material (B), and prominent fibrosis (C-E) was characteristic of RTV exposure of wt mice. Fibrosis was attenuated in PF4CreTgfb1flox/flox mice. Exposure of wt mice to CO daily for 4 hrs (250ppm with each RTV injection) suppressed RTV-induced fibrosis (C-E).

Article Snippet: Total plasma TGF-β1 was quantitated by ELISA using a TGF-β1 DuoSet kit (R&D Systems) after converting latent to active cytokine by acidification followed by neutralization, as previously described [ 13 ].

Techniques: Staining, Injection

Journal: AIDS (London, England)

Article Title: HIV protease inhibitor ritonavir induces renal fibrosis and dysfunction: role of platelet-derived TGF-β1 and intervention via anti-oxidant pathways

doi: 10.1097/QAD.0000000000002516

Figure Lengend Snippet: RTV-induced fibrosis involved over-expression of α-SMA and collagen α1 in kidneys, paralleled by TGF-β1-mediated pSmad2 signaling in wt but not platelet TGF-β1 deficient mice. Paraffin-embedded kidney sections were stained for α-SMA (A), collagen α1 (B), or phosphor-Smad2-specific antibodies. Photographs were taken using a Nikon Eclipse 80i fluorescence microscope with 20X magnification. Quantification of pSmad2 positive nuclei (red dots in blue nuclei counter-stained with DAPI) was determined using the NIH ImageJ program. Platelet TGF-β1 deficient PF4CreTgfb1flox/flox mice had lower levels of pSmad2 in kidney than wt mice after 8 wks of RTV exposure.

Article Snippet: Total plasma TGF-β1 was quantitated by ELISA using a TGF-β1 DuoSet kit (R&D Systems) after converting latent to active cytokine by acidification followed by neutralization, as previously described [ 13 ].

Techniques: Over Expression, Staining, Fluorescence, Microscopy

Journal: AIDS (London, England)

Article Title: HIV protease inhibitor ritonavir induces renal fibrosis and dysfunction: role of platelet-derived TGF-β1 and intervention via anti-oxidant pathways

doi: 10.1097/QAD.0000000000002516

Figure Lengend Snippet: RTV induced renal dysfunction in association with tubular injury in wt mice, its dependence on platelet TGF-β1, and suppression by inhaled CO in association with Nrf2 activation. Cystatin C levels in plasma (A) and KIM-1 levels in urine (B) were assessed by ELISA following 8 wks of RTV exposure. They were significantly elevated in RTV-exposed wt mice. RTV-exposed PF4CreTgfb1flox/flox mice had cystatin C levels equivalent to control wt mice (A). Total plasma TGF-β1 levels correlated with degree of cystatin C elevation (C). Paraffin-embedded kidney sections from wt mice treated with RTV or RTV plus inhaled CO were then stained for Nrf2 (D) and HO-1 (E) and positive areas quantified by confocal scanning and ImageJ analysis. Both proteins, involved in anti-oxidant signaling, were markedly increased following CO exposure.

Article Snippet: Total plasma TGF-β1 was quantitated by ELISA using a TGF-β1 DuoSet kit (R&D Systems) after converting latent to active cytokine by acidification followed by neutralization, as previously described [ 13 ].

Techniques: Activation Assay, Enzyme-linked Immunosorbent Assay, Staining