Kidney Ischemia/Reperfusion Injury Induces Changes in the Drug Transporter Expression at the Blood-Brain Barrier in vivo and in vitro.

Ischemia/reperfusion injury is a major cause of acute kidney injury (AKI). AKI is characterized by a sudden decrease in kidney function, systemic inflammation, oxidative stress, and dysregulation of the sodium, potassium, and water channels. While AKI leads to uremic encephalopathy, epidemiological studies have shown that AKI is associated with a subsequent risk for developing stroke and dementia. To get more insights into kidney-brain crosstalk, we have created an in vitro co-culture model based on human kidney cells of the proximal tubule (HK-2) and brain microvascular endothelial cells (BMEC). The HK-2 cell line was grown to confluence on 6-well plates and exposed to oxygen/glucose deprivation (OGD) for 4 h. Control HK-2 cells were grown under normal conditions. The BMEC cell line cerebED was grown to confluence on transwells with 0.4 µm pores. The transwell filters seeded and grown to confluence with cereEND were inserted into the plates with HK-2 cells with or without OGD treatment. In addition, cerebEND were left untreated or treated with uremic toxins, indole-3-acetic acid (IAA) and indoxyl sulfate (IS). The protein and mRNA expression of selected BBB-typical influx transporters, efflux transporters, cellular receptors, and tight junction proteins was measured in BMECs. To validate this in vitro model of kidney-brain interaction, we isolated brain capillaries from mice exposed to bilateral renal ischemia (30 min)/reperfusion injury (24 h) and measured mRNA and protein expression as described above. Both in vitro and in vivo systems showed similar changes in the expression of drug transporters, cellular receptors, and tight junction proteins. Efflux pumps, in particular Abcb1b, Abcc1, and Abcg2, have shown increased expression in our model. Thus, our in vitro co-culture system can be used to study the cellular mechanism of kidney and brain crosstalk in renal ischemia/reperfusion injury.

Is FKBP5 a genetic marker of affective psychosis? A case control study and analysis of disease related traits.

BACKGROUND: A dysregulation of the hypothalamic-pituitary-adrenal (HPA) axis has been proposed as an important pathogenic factor in depression. Genetic variants of FKBP5, a protein of the HPA system modulating the glucocorticoid receptor, have been reported to be genetically associated with improved response to medical treatment and an increase of depressive episodes. METHODS: We examined three single nucleotide polymorphisms (SNPs) in FKBP5, rs4713916 in the proposed promoter region, rs1360780 in the second intron and rs3800373 in the 3'-untranslated region (3'-UTR), in a case-control study of Caucasian origin (affective psychosis: n = 248; controls: n = 188) for genetic association and association with disease related traits. RESULTS: Allele and genotype frequencies of rs4713916, rs1360780 and rs3800373 were not significantly different between cases and controls. Two three-locus haplotypes, G-C-T and A-T-G, accounted for 86.2% in controls. Odds ratios were not increased between cases and controls, except the rare haplotype G-C-G (OR 6.81), representing 2.1% of cases and 0.3% of controls. The frequency of rs4713916AG in patients deviated from expected Hardy-Weinberg equilibrium, the genotype AA at rs4713916 in monopolar depression (P = 0.011), and the two-locus haplotype rs1360780T--rs3800373T in the total sample (overall P = 0.045) were nominally associated with longer continuance of disease. CONCLUSION: Our data do not support a significant genetic contribution of FKBP5 polymorphisms and haplotypes to affective psychosis, and the findings are inconclusive regarding their contribution to disease-related traits.

Systematic mutation analysis of KIAA0767 and KIAA1646 in chromosome 22q-linked periodic catatonia.

BACKGROUND: Periodic catatonia is a familial subtype of schizophrenia characterized by hyperkinetic and akinetic episodes, followed by a catatonic residual syndrome. The phenotype has been evaluated in two independent genome-wide linkage scans with evidence for a major locus on chromosome 15q15, and a second independent locus on chromosome 22qtel. METHODS: In the positional and brain-expressed candidate genes KIAA0767 and KIAA1646, we searched for variants in the complete exons and adjacent splice-junctions as well as in parts of the 5'- and 3'-untranslated regions by means of a systematic mutation screening in individuals from chromosome 22q-linked pedigrees. RESULTS: The mutation scan revealed 24 single nucleotide polymorphisms, among them two rare codon variants (KIAA0767: S159I; KIAA1646: V338G). However, both were neither found segregating with the disease in the respective pedigree nor found at a significant frequency in a case-control association sample. CONCLUSION: Starting from linkage signals at chromosome22qtel in periodic catatonia, we screened two positional brain-expressed candidate genes for genetic variation. Our study excludes genetic variations in the coding and putative promoter regions of KIAA0767 and KIAA1646 as causative factors for periodic catatonia.

Increased symmetrical dimethylarginine in ischemic acute kidney injury as a causative factor of renal L-arginine deficiency.

Availability of L-arginine, the exclusive substrate for nitric oxide synthases, plays an important role in kidney ischemia/reperfusion injury. The endogenous L-arginine derivatives asymmetrical dimethylarginine (ADMA) and symmetrical dimethylarginine (SDMA) block cellular L-arginine uptake competitively, thereby inhibiting the production of nitric oxide. ADMA also blocks nitric oxide synthase activity directly. Here, we investigate the pathomechanistic impact of ADMA and SDMA on ischemic acute kidney injury. Rats were subject to bilateral renal ischemia (60 minutes)/reperfusion (24 hours) injury. Impairment of renal function was determined with inulin clearance (glomerular filtration rate) and para-aminohippurate (PAH) clearance (renal plasma flow). L-arginine, ADMA, and SDMA levels were measured by liquid chromatography-tandem mass spectrometry. L-arginine was extracted from renal tissue and analyzed by enzyme-linked immunosorbent assay, and protein and messenger RNA expressions were determined by Western blot and real-time reverse transcription polymerase chain reaction. Renal function deteriorated severely after ischemia/reperfusion injury, as demonstrated by inulin and PAH clearance. Serum ADMA and SDMA increased, but tissue expression of specific ADMA or SDMA synthesizing and metabolizing enzymes (protein arginine methyltransferases and dimethyl arginine dimethylaminohydrolases) did not alter. Serum L-arginine increased as well, whereas intracellular L-arginine concentration diminished. Renal messenger RNA expression of cationic amino acid transporters, which mediate L-arginine uptake, remained unchanged. In serum, the ratio of L-arginine to ADMA did not alter after ischemia/reperfusion injury, whereas the ratios of L-arginine to SDMA and ADMA to SDMA decreased. A marked increase in serum SDMA, especially when accompanied by a diminished L-arginine-to-SDMA ratio, might reflect competitive inhibition of cellular L-arginine uptake by SDMA. As a consequence, a pathologic renal L-arginine deficiency in ischemic acute kidney injury results.

Family-based study of markers at the 5'-flanking region of the human dopamine transporter gene reveals potential association with schizophrenic psychoses.

The dopamine hypothesis of schizophrenia proposes an inherited or acquired presynaptic hyperactivity of dopaminergic neurons. The human dopamine transporter gene (hSLC6A3; hDAT) represents one major mechanism for the termination of dopaminergic neurotransmission. This study examines the degree of genetic association of the 5'-untranslated region (5'-UTR) of the hSLC6A3 to schizophrenia in a family-based association design. Five single nucleotide polymorphisms (SNPs) derived by a previous systematic mutation scan approximately 1.2 kb of the 5'-UTR of the hSLC6A3 locus were genotyped for transmission disequilibrium between 82 index cases (56 males) with schizophrenia and their biological parents. We observed no preferential transmission of alleles from heterozygous parents to affected offspring. Five estimated haplotypes accounted for a frequency of 90% in the index cases, and were identical in cases and non-transmitted parental control haplotypes. Distinct five-locus-genotypes accumulated in schizophrenia compared to parental controls at P-value 0.0038 with odds-ratio of 2.02 (95% CI 0.99-4.14). In conclusion, our present findings support the genetic involvement of distinct hSLC6A3 genotypes in schizophrenia. We propose replication in extended samples and examination of the functional relevance of the associated genotypes on human dopamine transporter expression.