The sodium-dependent di- and tricarboxylate transporter, NaCT, is not responsible for the uptake of D-, L-2-hydroxyglutarate and 3-hydroxyglutarate into neurons

Multi-objective evolutionary algorithms (MOEAs) have become increasingly popular as multi-objective problem solving techniques. Most studies of MOEAs are empirical. Only recently, a few theoretical results have appeared. It is acknowledged that more theoretical research is needed. An important open problem is to understand the role of populations in MOEAs. We present a simple bi-objective problem which emphasizes when populations are needed. Rigorous runtime analysis point out an exponential runtime gap between a population-based algorithm (SEMO) and several single individual-based algorithms on this problem. This means that among the algorithms considered, only the populationbased MOEA is successful and all other algorithms fail

Oksalat – od okoliša do bubrežnih kamenaca

Oxalate urolithiasis (nephrolithiasis) is the most frequent type of kidney stone disease. Epidemiological research has shown that urolithiasis is approximately twice as common in men as in women, but the underlying mechanism of this sex-related prevalence is unclear. Oxalate in the organism partially originate from food (exogenous oxalate) and largely as a metabolic end-product from numerous precursors generated mainly in the liver (endogenous oxalate). Oxalate concentrations in plasma and urine can be modified by various foodstuffs, which can interact in positively or negatively by affecting oxalate absorption, excretion, and/or its metabolic pathways. Oxalate is mostly removed from blood by kidneys and partially via bile and intestinal excretion. In the kidneys, after reaching certain conditions, such as high tubular concentration and damaged integrity of the tubule epithelium, oxalate can precipitate and initiate the formation of stones. Recent studies have indicated the importance of the SoLute Carrier 26 (SLC26) family of membrane transporters for handling oxalate. Two members of this family [Sulfate Anion Transporter 1 (SAT-1; SLC26A1) and Chloride/Formate EXchanger (CFEX; SLC26A6)] may contribute to oxalate transport in the intestine, liver, and kidneys. Malfunction or absence of SAT-1 or CFEX has been associated with hyperoxaluria and urolithiasis. However, numerous questions regarding their roles in oxalate transport in the respective organs and male-prevalent urolithiasis, as well as the role of sex hormones in the expression of these transporters at the level of mRNA and protein, still remain to be answered.Oksalatna urolitijaza (nefrolitijaza) najučestaliji je tip bolesti bubrežnih kamenaca. Rezultati epidemioloških istraživanja pokazali su da je urolitijaza približno dvostruko učestalija u muškaraca nego u žena, ali osnovni mehanizam nastanka ove spolno-ovisne prevalencije nije razjašnjen. Oksalat u organizmu dijelom potječe iz hrane (egzogeni oksalat), a glavninom nastaje kao konačni produkt metabolizma raznih preteča u jetrima (endogeni oksalat). Na koncentraciju oksalata u plazmi i urinu utječu razne tvari iz hrane, koje mogu pozitivno ili negativno djelovati na apsorpciju, metaboličke puteve i/ili izlučivanje oksalata. Oksalat se iz organizma izlučuje u manjem obimu putem žuči u crijevo, a glavninom bubrezima. U bubrezima, pri odgovarajućim uvjetima kao što su visoka koncentracija oksalata i oštećenje epitela bubrežnih kanalića, oksalat može precipitirati i time potaknuti stvaranje kamenaca. Rezultati novih istraživanja upućuju na važnost membranskih prijenosnika otopljenih tvari (SoLute Carriers) iz obitelji 26 (SLC26) za prijenos oksalata u specifičnim organima. Smatra se da dva člana ove obitelji: prijenosnik sulfatnog aniona (Sulfate Anion Transporter 1; SAT-1; SLC26A1) i izmjenjivač klora i mravlje kiseline (Chloride/Formate EXchanger; CFEX; SLC26A6), imaju značajnu ulogu u prijenosu oksalata u crijevima, jetrima i bubrezima; hiperoksalurija i nefrolitijaza utvrđeni su pri slaboj aktivnosti ili nedostatku SAT-1 i CFEX proteina. Međutim, još uvijek postoje brojne nejasnoće glede prijenosa oksalata u navedenim organima, mehanizma nastanka spolnih razlika u nefrolitijazi i utjecaja spolnih hormona na ekspresiju proteina i mRNA za navedene prijenosnike

The Isoquinolone Derived Prolyl Hydroxylase Inhibitor ICA Is a Potent Substrate of the Organic Anion Transporters 1 and 3

Objective: Many cellular responses to hypoxia are mediated by the transcription factor complex hypoxia-inducible factor (HIF). HIF stability is governed by a family of dioxygenases called HIF prolyl hydroxylases (PHDs). Isoquinolone-derived PHD inhibitors, like 2-(1-chloro-4-hydroxyisoquinoline-3-carboxamido) acetate (ICA), which stabilize the intracellular HIF-α have been suggested as a potentially beneficial therapeutic strategy for the treatment of disorders associated with ischemia. To stabilize HIF-α, ICA has to be taken up into proximal tubule cells (PCTs) across the basolateral membrane by one of the organic anion transporters 1, 2 or 3 (OAT1, OAT2 or OAT3). The release into the urine across the luminal membrane may be mediated by OAT4. Method: To demonstrate interaction of ICA with human OAT1, OAT2, OAT3 and OAT4, ICA was tested on these transporters stably transfected in HEK293 cells by using p-aminohippurate (PAH), cGMP and estrone-3-sulfate (ES) as reference substrates, respectively. Results: Uptakes of PAH and ES in OAT1- and OAT3-transfected HEK293 cells were inhibited by ICA with half-maximal inhibition values of 0.29 ± 0.05 and 2.58 ± 0.16 µM, respectively. OAT2 was less sensitive to ICA. Efflux experiments identified ICA as an OAT1 and OAT3 substrate. Preloading OAT4-transfected HEK293 cells with ICA stimulated ES uptake by 18.3 ± 3.8%. Conclusion: The uptake of ICA across the basolateral membrane of PCTs occurs mainly by OAT1 and the efflux into the tubular lumen by OAT4

Role of N-glycosylation in renal betaine transport

The osmolyte and folding chaperone betaine is transported by the renal Na+-coupled GABA (γ-aminobutyric acid) symporter BGT-1 (betaine/GABA transporter 1), a member of the SLC6 (solute carrier 6) family. Under hypertonic conditions, the transcription, translation and plasma membrane (PM) insertion of BGT-1 in kidney cells are significantly increased, resulting in elevated betaine and GABA transport. Re-establishing isotonicity involves PM depletion of BGT-1. The molecular mechanism of the regulated PM insertion of BGT-1 during changes in osmotic stress is unknown. In the present study, we reveal a link between regulated PM insertion and N-glycosylation. Based on homology modelling, we identified two sites (Asn171 and Asn183) in the extracellular loop 2 (EL2) of BGT-1, which were investigated with respect to trafficking, insertion and transport by immunogold-labelling, electron microscopy (EM), mutagenesis and two-electrode voltage clamp measurements in Xenopus laevis oocytes and uptake of radiolabelled substrate into MDCK (Madin–Darby canine kidney) and HEK293 (human embryonic kidney) cells. Trafficking and PM insertion of BGT-1 was clearly promoted by N-glycosylation in both oocytes and MDCK cells. Moreover, association with N-glycans at Asn171 and Asn183 contributed equally to protein activity and substrate affinity. Substitution of Asn171 and Asn183 by aspartate individually caused no loss of BGT-1 activity, whereas the double mutant was inactive, suggesting that N-glycosylation of at least one of the sites is required for function. Substitution by alanine or valine at either site caused a dramatic loss in transport activity. Furthermore, in MDCK cells PM insertion of N183D was no longer regulated by osmotic stress, highlighting the impact of N-glycosylation in regulation of this SLC6 transporter

In female rats, ethylene glycol treatment elevates protein expression of hepatic and renal oxalate transporter sat-1 (Slc26a1) without inducing hyperoxaluria

Aim To investigate whether the sex-dependent expression of hepatic and renal oxalate transporter sat-1 (Slc26a1) changes in a rat model of ethylene glycol (EG)-induced hyperoxaluria. Methods Rats were given tap water (12 males and 12 females; controls) or EG (12 males and 12 females; 0.75% v/v in tap water) for one month. Oxaluric state was confirmed by biochemical parameters in blood plasma, urine, and tissues. Expression of sat-1 and rate-limiting enzymes of oxalate synthesis, alcohol dehydrogenase 1 (Adh1) and hydroxy-acid oxidase 1 (Hao1), was determined by immunocytochemistry (protein) and/or real time reverse transcription polymerase chain reaction (mRNA). Results EG-treated males had significantly higher (in μmol/L; mean ± standard deviation) plasma (59.7 ± 27.2 vs 12.9 ± 4.1, P < 0.001) and urine (3716 ± 1726 vs 241 ± 204, P < 0.001) oxalate levels, and more abundant oxalate crystaluria than controls, while the liver and kidney sat-1 protein and mRNA expression did not differ significantly between these groups. EG-treated females, in comparison with controls had significantly higher (in μmol/L) serum oxalate levels (18.8 ± 2.9 vs 11.6 ± 4.9, P < 0.001), unchanged urine oxalate levels, low oxalate crystaluria, and significantly higher expression (in relative fluorescence units) of the liver (1.59 ± 0.61 vs 0.56 ± 0.39, P = 0.006) and kidney (1.77 ± 0.42 vs 0.69 ± 0.27, P < 0.001) sat-1 protein, but not mRNA. The mRNA expression of Adh1 was femaledominant and that of Hao1 male-dominant, but both were unaffected by EG treatment. Conclusions An increased expression of hepatic and renal oxalate transporting protein sat-1 in EG-treated female rats could protect from hyperoxaluria and oxalate urolithiasis

Spolno-neovisna ekspresija izmjenjivača klora i mravlje kiseline Cfex (Slc26a6) u gušterači, tankom crijevu i jetri štakora i povišena ekspresija u bubrezima mužjaka

Chloride/formate exchanger (CFEX; SLC26A6) mediates oxalate transport in various mammalian organs. Studies in Cfex knockout mice indicated its possible role in development of male-dominant hyperoxaluria and oxalate urolithiasis. Rats provide an important model for studying this pathophysiological condition, but data on Cfex (rCfex) localisation and regulation in their organs are limited. Here we applied the RT-PCR and immunochemical methods to investigate rCfex mRNA and protein expression and regulation by sex hormones in the pancreas, small intestine, liver, and kidneys from intact prepubertal and adult as well as gonadectomised adult rats treated with sex hormones. rCfex cDNA-transfected HEK293 cells were used to confirm the specificity of the commercial anti-CFEX antibody. Various biochemical parameters were measured in 24-h urine collected in metabolic cages. rCfex mRNA and related protein expression varied in all tested organs. Sex-independent expression of the rCfex protein was detected in pancreatic intercalated ducts (apical domain), small intestinal enterocytes (brush-border membrane; duodenum > jejunum > ileum), and hepatocytes (canalicular membrane). In kidneys, the rCfex protein was immunolocalised to the proximal tubule brush-border with segment-specific pattern (S1=S2 jejunum > ileum) i kanalikularnoj membrani hepatocita. U bubrezima je a) prijenosnik rCfex imunolokaliziran u četkastoj membrani proksimalnih kanalića sa segment-specifičnim obrascem (S1=S2 ženke) zbog stimulacijskoga učinka androgena nakon puberteta. Međutim, izlučivanje oksalata urinom nije bilo sukladno ekspresiji bubrežnoga prijenosnika rCfex. Dakle, nejasan je učinak povišene ekspresije prijenosnika rCfex u proksimalnim kanalićima mužjaka na izlučivanje oksalata, a postojanje prijenosnika u kanalikularnoj membrani hepatocita mogući je put izlučivanja oksalata putem žuči

Interaction of Excitatory Amino Acid Transporters 1 – 3 (EAAT1, EAAT2, EAAT3) with N-Carbamoylglutamate and N-Acetylglutamate

Background/Aims: Inborn deficiency of the N-acetylglutamate synthase (NAGS) impairs the urea cycle and causes neurotoxic hyperammonemia. Oral administration of N-carbamoylglutamate (NCG), a synthetic analog of N-acetylglutamate (NAG), successfully decreases plasma ammonia levels in the affected children. Due to structural similarities to glutamate, NCG may be absorbed in the intestine and taken up into the liver by excitatory amino acid transporters (EAATs). Methods: Using Xenopus laevis oocytes expressing either human EAAT1, 2, or 3, or human sodium-dependent dicarboxylate transporter 3 (NaDC3), transport-associated currents of NAG, NCG, and related dicarboxylates were assayed. Results: L-aspartate and L-glutamate produced saturable inward currents with Km values below 30 µM. Whereas NCG induced a small inward current only in EAAT3 expressing oocytes, NAG was accepted by all EAATs. With EAAT3, the NAG-induced current was sodium-dependent and saturable (Km 409 µM). Oxaloacetate was found as an additional substrate of EAAT3. In NaDC3-expressing oocytes, all dicarboxylates induced much larger inward currents than did L-aspartate and L-glutamate. Conclusion: EAAT3 may contribute to intestinal absorption and hepatic uptake of NCG. With respect to transport of amino acids and dicarboxylates, EAAT3 and NaDC3 can complement each other