Maternally supplied S-acyl-transferase is required for crystalloid organelle formation and transmission of the malaria parasite.

Transmission of the malaria parasite from the mammalian host to the mosquito vector requires the formation of adequately adapted parasite forms and stage-specific organelles. Here we show that formation of the crystalloid-a unique and short-lived organelle of the Plasmodium ookinete and oocyst stage required for sporogony-is dependent on the precisely timed expression of the S-acyl-transferase DHHC10. DHHC10, translationally repressed in female Plasmodium berghei gametocytes, is activated translationally during ookinete formation, where the protein is essential for the formation of the crystalloid, the correct targeting of crystalloid-resident protein LAP2, and malaria parasite transmission

Glomerular permeability is not affected by heparan sulfate glycosaminoglycan deficiency in zebrafish embryos

Proteinuria develops when specific components in the glomerular filtration barrier have impaired function. Although the precise components involved in maintaining this barrier have not been fully identified. heparan sulfate proteoglycans are believed to play an essential role in maintaining glomerular filtration. Although in situ studies have shown that a loss of heparan sulfate glycosaminoglycans increases the permeability of the glomerular filtration barrier. recent studies using experimental models have shown that podocyte-specific deletion of heparan sulfate glycosaminoglycan assembly does not lead to proteinuria. However, tubular reabsorption of leaked proteins might have masked an increase in glomerular permeability in these models. Furthermore, not only podocytes but also glomerular endothelial cells are involved in heparan sulfate synthesis in the glomerular filtration barrier. Therefore, we investigated the effect of a global heparan sulfate glycosaminoglycan deficiency on glomerular permeability. We used a zebrafish embryo model carrying a homozygous germline mutation in the ext2 gene. Glomerular permeability was assessed with a quantitative dextran tracer injection method. In this model, we accounted for tubular reabsorption. Loss of anionic sites in the glomerular basement membrane was measured using polyethyleneimine staining. Although mutant animals had significantly fewer negatively charged areas in the glomerular basement membrane. glomerular permeability was unaffected. Moreover, heparan sulfate glycosaminoglycan-deficient embryos had morphologically intact podocyte foot processes. Glomerular filtration remains fully functional despite a global reduction of heparan sulfate.Animal science

Glomerular Function and Structural Integrity Depend on Hyaluronan Synthesis by Glomerular Endothelium

Contains fulltext : 209662.pdf (publisher's version ) (Closed access

Direct Observation of Enhanced Nitric Oxide in a Murine Model of Diabetic Nephropathy.

Uncoupling of nitric oxide synthase (NOS) secondary to redox signaling is a central mechanism in endothelial and macrophage activation. To date studies on the production of nitric oxide (NO) during the development of diabetic complications show paradoxical results. We previously showed that recoupling eNOS by increasing the eNOS cofactor tetrahydrobiopterin (BH4) could restore endothelial function and prevent kidney injury in experimental kidney transplantation. Here, we employed a diabetic mouse model to investigate the effects of diabetes on renal tissue NO bioavailability. For this, we used in vivo NO trapping, followed by electron paramagnetic resonance spectroscopy. In addition, we investigated whether coupling of NOS by supplying the cofactor BH4 could restore glomerular endothelial barrier function. Our data show that overall NO availability at the tissue level is not reduced sixteen weeks after the induction of diabetes in apoE knockout mice, despite the presence of factors that cause endothelial dysfunction, and the presence of the endogenous NOS inhibitor ADMA. Targeting uncoupled NOS with the BH4 precursor sepiapterin further increases NO availability, but did not modify renal glomerular injury. Notably, glomerular heparanase activity as a driver for loss of glomerular barrier function was not reduced, pointing towards NOS-independent mechanisms. This was confirmed by unaltered increased glomerular presence of cathepsin L, the protease that activates heparanase

Increased NO levels affect endothelial glycocalyx non-uniformly.

<p>(A) Representative microscopic images of cationic ferritin (TEM; top), heparanase (HPSE, immunofluorescence; middle) and cathepsin L (CTSL; bottom) in glomeruli of apoE KO mice (apoE), diabetic apoE KO mice (DM) and diabetic apoE KO mice treated with sepiapterin (DM + S). (B) Quantification of endothelial cationic ferritin coverage in 6–8 capillary loops in 9 glomeruli of 3 mice, shown as mean percentage of total capillary length ± SD, (C) Quantification of glomerular heparanase expression, shown as mean area percentage ± SD. (D) Quantification of glomerular cathepsin L expression, shown as mean area percentage ± SD. *P<0.05 compared with ApoE, n = 6–8. Scale bars: 500 nm in TEM images; 20 μm in fluorescent and light microscopic images.</p

Tissue-dependent variation in NO free radical induction.

<p>(A) EPR spectrum of frozen kidney samples. The characteristic triplet structure of the mononitrosyl-iron complex (MNIC, double-headed arrow) centers around g  =  2.035 and represents the formation of local nitric oxide in 334–370 mg tissue. (B-D) Quantification of nitric oxide formation in kidney, liver and heart tissue, shown as mean pmol MNIC / mg wet tissue ± SD, n = 7–9. E) Plasma ADMA concentrations, shows as mean ± SD, n = 8. *P<0.05, compared with ApoE; #P<0.05 compared with DM. ApoE = ApoE KO mice, DM = diabetic apoE KO mice, DM + S = diabetic apoE KO mice + sepiapterin.</p

Experimental set-up for assessment of NO bioavailability.

<p>(A) male ApoE KO mice (<i>B6</i>.<i>129P2- Apoe</i>^{<i>tm1Unc</i>}<i>/J)</i> were injected with citrate buffer ± STZ. Diabetic mice received cholesterol enriched diet and insulin from week 8 onwards. At 18 week of age, diabetic mice were treated with sepiapterin or received normal drinking water for 4 weeks. Urine was collected upon commencing with the experimental procedure and after 2 and 4 weeks of treatment. At 22 weeks, plasma was collected and the mice were sacrificed.</p

Sepiapterin does not reduce albuminuria in diabetic apoE KO mice.

<p>(A) PAS-stained glomeruli of apoE KO mice (apoE), diabetic apoE KO mice (DM) and diabetic apoE KO mice treated with sepiapterin (DM + S), showing heterogeneous diabetic lesions 14 weeks after induction of diabetes with STZ (20). Scale bars: 20 μm. Sepiapterin did not affect mesangial area (B,C), nor blood glucose concentrations (D). Data are shown as mean ± SD, *P<0.05 compared with apoE, n = 8. (E) Albumin-creatinine ratios (ACR) at baseline, 2- and 4 weeks after treatment, as indicated by mean ± SEM, *P<0.05 compared with apoE, n = 14–23.</p