Comparison between indicine and taurine cattle DNA methylation reveals epigenetic variation associated to differences in morphological adaptive traits.

Indicine and taurine subspecies present distinct morphological traits as a consequence of environmental adaptation and artificial selection. Although the two subspecies have been characterized and compared at genome-wide level and at specific loci, their epigenetic diversity has not yet been explored. In this work, Reduced Representation Bisulphite Sequencing (RRBS) profiling of the taurine Angus (A) and indicine Nellore (N) cattle breeds was applied to identify methylation differences between the two subspecies. Genotyping by sequencing (GBS) of the same animals was performed to detect single nucleotide polymorphisms (SNPs) at cytosines in CpG dinucleotides and remove them from the differential methylation analysis. A total of 660,845 methylated cytosines were identified within the CpG context (CpGs) across the 10 animals sequenced (5 N and 5 A). A total of 25,765 of these were differentially methylated (DMCs). Most DMCs clustered in CpG stretches nearby genes involved in cellular and anatomical structure morphogenesis. Also, sequences flanking DMC were enriched in SNPs compared to all other CpGs, either methylated or unmethylated in the two subspecies. Our data suggest a contribution of epigenetics to the regulation and divergence of anatomical morphogenesis in the two subspecies relevant for cattle evolution and sub-species differentiation and adaptation.

Bos taurus haplotypes segregating in Nellore (Bos indicus) cattle.

Brazil is the largest exporter of beef in the world, and most of that beef derives from Nellore cattle. Although considered a zebu breed (Bos indicus), the history of Nellore cattle in Brazil is marked by the importation of bulls from India, the use of a Creole taurine (Bos taurus) maternal lineage to quickly expand the herds and backcrossing to Nellore bulls to recover zebu ancestry. As a consequence, the current Brazilian Nellore population carries an average taurine ancestry of approximately 1%. Although that percentage seems small, some taurine variants deviate substantially from that average, with the better-known cases being the PLAG1-Q haplotype involved with body size variation and the Guarani (PG ) polled variant producing hornless animals. Here, we report taurine haplotypes in 9074 Nellore animals genotyped for 539 657 imputed SNP markers. Apart from PLAG1-Q and PG , our analysis further revealed common taurine haplotypes (>3%) spanning genes related to immunity, growth, reproduction and hair and skin phenotypes. Using data from 22 economically important traits, we showed that many of the major QTL previously reported in the breed are at least partially driven by taurine haplotypes. As B. taurus and B. indicus haplotypes are highly divergent, presenting widely different sets of functional variants, our results provide promising targets for future scrutiny in Nellore cattle.

Association of missense variants in GDF9 with litter size in Entlebucher Mountain dogs.

In the past two decades, average litter size (ALS) in Entlebucher Mountain dogs decreased by approximately 0.8 puppies. We conducted a GWAS for ALS using the single-step methodology to take advantage of 1632 pedigree records, 892 phenotypes and 372 genotypes (173 662 markers) for which only 12% of the dogs had both phenotypes and genotypes available. Our analysis revealed associations towards the growth differentiation factor 9 gene (GDF9), which is known to regulate oocyte maturation. The trait heritability was estimated at 43.1%, from which approximately 15% was accountable by the GDF9 locus alone. Therefore, markers flanking GDF9 explained approximately 6.5% of the variance in ALS. Analysis of WGSs revealed two missense substitutions in GDF9, one of which (g.11:21147009G>A) affected a highly conserved nucleotide in vertebrates. The derived allele A was validated in 111 dogs and shown to be associated with decreased ALS (-0.75 ± 0.22 puppies per litter). The variant was further predicted to cause a proline to serine substitution. The affected residue was immediately followed by a six-residue deletion that is fixed in the canine species but absent in non-canids. We further confirmed that the deletion is prevalent in the Canidae family by sequencing three species of wild canids. Since canids uniquely ovulate oocytes at the prophase stage of the first meiotic division, requiring maturation in the oviduct, we conjecture that the amino acid substitution and the six-residue deletion of GDF9 may serve as a model for insights into the dynamics of oocyte maturation in canids.

Genome-wide mapping of the dominance effects based on breed ancestry for semen traits in admixed Swiss Fleckvieh bulls.

Heterosis is the beneficial deviation of crossbred progeny from the average of parental lines for a particular trait. Heterosis is due to nonadditive genetic effects with dominance and epistatic components. Recent advances in genotyping technology have encouraged researchers to estimate and scan heterosis components for a range of traits in crossbred populations, applying various definitions of such components. In this study, we defined the intralocus (dominance) component of heterosis using local genetic ancestry and performed genome-wide association analysis for admixed Swiss Fleckvieh bulls and their parental populations, Red Holstein Friesian and Swiss Simmental, for semen traits. A linear mixed model for 41,824 SNP, including SNP additive genetic, breed additive, and breed dominance effects on 1,178 bulls (148 Red Holstein Friesian, 213 Swiss Simmental, and 817 Swiss Fleckvieh) with a total of 43,782 measurements was performed. In total, 19 significant regions for breed dominance were identified for volume (2 regions on Bos taurus autosome 10 and 22) and percentage of live spermatozoa (17 regions on Bos taurus autosome 3, 4, 5, 7, 13, 14, and 17), and genes associated with spermatogenesis, sperm motility, and male fertility traits were located there. No significant region for breed dominance was detected for total number of spermatozoa. The signals for breed dominance were relatively wide, most likely due to limited numbers of recombination events in a small number of generations (10-15 generations) of crossbreeding in the recent Swiss Fleckvieh composite.

Genomic clues of the evolutionary history of Bos indicus cattle.

Together with their sister subspecies Bos taurus, zebu cattle (Bos indicus) have contributed to important socioeconomic changes that have shaped modern civilizations. Zebu cattle were domesticated in the Indus Valley 8000 years before present (YBP). From the domestication site, they expanded to Africa, East Asia, southwestern Asia and Europe between 4000 and 1300 YBP, intercrossing with B. taurus to form clinal variations of zebu ancestry across the landmass of Afro-Eurasia. In the past 150 years, zebu cattle reached the Americas and Oceania, where they have contributed to the prosperity of emerging economies. The zebu genome is characterized by two mitochondrial haplogroups (I1 and I2), one Y chromosome haplogroup (Y3) and three major autosomal ancestral groups (Indian-Pakistani, African and Chinese). Phenotypically, zebu animals are recognized by their hump, large ears and excess skin. They are rustic, resilient to parasites and capable of bearing the hot and humid climates of the tropics. Many resources are available to study the zebu genome, including commercial arrays of SNP, reference assemblies and publicly available genotypes and whole-genome sequences. Nevertheless, many of these resources were initially developed to support research and subsidize industrial applications in B. taurus, and therefore they can produce bias in data analysis. The combination of genomics with precision agriculture holds great promise for the identification of genetic variants affecting economically important traits such as tick resistance and heat tolerance, which were naturally selected for millennia and played a major role in the evolution of B. indicus cattle.

Genome-wide association study and heritability estimate for ectopic ureters in Entlebucher mountain dogs.

An ectopic ureter is a congenital anomaly which may lead to urinary incontinence and without a surgical intervention even to end-stage kidney disease. A genetic component contributes to the development of this anomaly in Entlebucher mountain dogs (EMD); however, its nature remains unclear. Using the Illumina CanineHD bead chip, a case-control genome-wide association study was performed to identify SNPs associated with the trait. Six loci on canine chromosomes 3, 17, 27 and 30 were identified with 16 significantly associated SNPs. There was no single outstanding SNP associated with the phenotype, and the association signals were not close to known genes involved in human congenital anomalies of the kidney or lower urinary tract. Additional research will be necessary to elucidate the potential role of the associated genes in the development of ectopic ureters in the EMD breed.

Effects of breed proportion and components of heterosis for semen traits in a composite cattle breed.

The aim of this study was to estimate the non-additive genetic effects of the dominance component of heterosis as well as epistatic loss on semen traits in admixed Swiss Fleckvieh, a composite of Simmental (SI) and Red Holstein Friesian (RHF) cattle. Heterosis is the additional gain in productivity or fitness of cross-bred progeny over the mid-purebred parental populations. Intralocus gene interaction usually has a positive effect, while epistatic loss generally reduces productivity or fitness due to lack of evolutionarily established interactions of genes from different breeds. Genotypic data on 38,205 SNP of 818 admixed, as well as 148 RHF and 213 SI bulls as the parental breeds were used to predict breed origin of alleles. The genomewide locus-specific breed ancestries of individuals were used to calculate effects of breed difference as well as the dominance component of heterosis, while proxies for two definitions of epistatic loss were derived from 100,000 random pairs of loci. The average Holstein Friesian ancestry in admixed bulls was estimated 0.82. Results of fitting different linear mixed models showed including the dominance component of heterosis considerably improved the model adequacy for three of the four traits. Inclusion of epistatic loss increased the accuracy of the models only for our new definition of the epistatic effect for two traits, while the other definition was so highly correlated with the dominance component that statistical separation was impossible.

Dog skin parasite load, TLR-2, IL-10 and TNF-α expression and infectiousness.

Visceral leishmaniosis is a zoonotic disease that is transmitted by Lutzomyia longipalpis sandflies. Dogs are the main peri-urban reservoir of the disease, and progression of canine leishmaniosis is dependent on the type of immune response elaborated against the parasite. Type 1 immunity is characterized by effective cellular response, with production of pro-inflammatory cytokines such as tumour necrosis factor alpha (TNF-α). In contrast, Type 2 immunity is predominantly humoral, associated with progression of the disease and mediated by anti-inflammatory cytokines such as interleukin 10 (IL-10). Although seemly important in the dynamics of leishmaniosis, other gene products such as toll-like receptor 2 (TRL-2) and inducible nitric oxide synthase (iNOS) exert unclear roles in the determination of the type of immune response. Given that the dog skin serves as a micro-environment for the multiplication of Leishmania spp., we investigated the parasite load and the expression of TLR-2, iNOS, IL-10 and TNF-α in the skin of 29 infected and 8 control dogs. We found that increased parasite load leads to upregulation of TLR-2, IL-10 and TNF-α, indicating that abundance of these transcripts is associated with infection. We also performed a xenodiagnosis to demonstrate that increased parasitism is a risk factor for infectiousness to sandflies.

Accuracy of genomic predictions in Gyr (Bos indicus) dairy cattle.

Genomic selection may accelerate genetic progress in breeding programs of indicine breeds when compared with traditional selection methods. We present results of genomic predictions in Gyr (Bos indicus) dairy cattle of Brazil for milk yield (MY), fat yield (FY), protein yield (PY), and age at first calving using information from bulls and cows. Four different single nucleotide polymorphism (SNP) chips were studied. Additionally, the effect of the use of imputed data on genomic prediction accuracy was studied. A total of 474 bulls and 1,688 cows were genotyped with the Illumina BovineHD (HD; San Diego, CA) and BovineSNP50 (50K) chip, respectively. Genotypes of cows were imputed to HD using FImpute v2.2. After quality check of data, 496,606 markers remained. The HD markers present on the GeneSeek SGGP-20Ki (15,727; Lincoln, NE), 50K (22,152), and GeneSeek GGP-75Ki (65,018) were subset and used to assess the effect of lower SNP density on accuracy of prediction. Deregressed breeding values were used as pseudophenotypes for model training. Data were split into reference and validation to mimic a forward prediction scheme. The reference population consisted of animals whose birth year was ≤2004 and consisted of either only bulls (TR1) or a combination of bulls and dams (TR2), whereas the validation set consisted of younger bulls (born after 2004). Genomic BLUP was used to estimate genomic breeding values (GEBV) and reliability of GEBV (R2PEV) was based on the prediction error variance approach. Reliability of GEBV ranged from ∼0.46 (FY and PY) to 0.56 (MY) with TR1 and from 0.51 (PY) to 0.65 (MY) with TR2. When averaged across all traits, R2PEV were substantially higher (R2PEV of TR1 = 0.50 and TR2 = 0.57) compared with reliabilities of parent averages (0.35) computed from pedigree data and based on diagonals of the coefficient matrix (prediction error variance approach). Reliability was similar for all the 4 marker panels using either TR1 or TR2, except that imputed HD cow data set led to an inflation of reliability. Reliability of GEBV could be increased by enlarging the limited bull reference population with cow information. A reduced panel of ∼15K markers resulted in reliabilities similar to using HD markers. Reliability of GEBV could be increased by enlarging the limited bull reference population with cow information.

Correlations between peripheral parasite load and common clinical and laboratory alterations in dogs with visceral leishmaniasis.

Intensity of peripheral parasite infection has an important role in the transmission of Leishmania spp. from one host to another. As parasite load quantification is still an expensive procedure to be used routinely in epidemiological surveillance, the use of surrogate predictors may be an important asset in the identification of dogs with high transmitting ability. The present study examined whether common clinical and laboratory alterations can serve as predictors of peripheral parasitism in dogs naturally infected with Leishmania spp. Thirty-seven dogs were examined in order to establish correlations between parasite load (PL) in multiple peripheral tissues and common clinical and laboratory findings in canine visceral leishmaniasis (CVL). Quantitative polymerase chain reaction was employed to determine PL in conjunctival swabs, ear skin, peripheral blood and buffy coat. Additionally, a series of hematological, biochemical and oxidative stress markers were quantified. Correlations between net peripheral infection and severity of clinical alterations and variation in laboratory parameters were assessed through a new analytical approach, namely Compressed Parasite Load Data (CPLD), which uses dimension reduction techniques from multivariate statistics to summarize PL across tissues into a single variable. The analysis revealed that elevation in PL is positively correlated with severity of clinical sings commonly observed in CVL, such as skin lesions, ophthalmic alterations, onycogriphosis, popliteal lymphadenomegaly and low body mass. Furthermore, increase in PL was found to be followed by intensification of non-regenerative anemia, neutrophilia, eosinopenia, hepatic injury and oxidative imbalance. These results suggest that routinely used clinical and laboratory exams can be predictive of intensity of peripheral parasite infection, which has an important implication in the identification of dogs with high transmitting ability.

Locus-specific ancestry to detect recent response to selection in admixed Swiss Fleckvieh cattle.

Identification of selection signatures is one of the current endeavors of evolutionary genetics. Admixed populations may be used to infer post-admixture selection. We calculated local ancestry for Swiss Fleckvieh, a composite of Simmental (SI) and Red Holstein Friesian (RHF), to infer such signals. Illumina Bovine SNP50 BeadChip data for 300 admixed, 88 SI and 97 RHF bulls were used. The average RHF ancestry across the whole genome was 0.70. To identify regions with high deviation from average, we considered two significance thresholds, based on a permutation test and extreme deviation from normal distribution. Regions on chromosomes 13 (46.3-47.3 Mb) and 18 (18.7-25.9 Mb) passed both thresholds in the direction of increased SI. Extended haplotype homozygosity within (iHS) and between (Rsb) populations was calculated to explore additional patterns of pre- and post-admixture selection signals. The Rsb score of admixed and SI was significant in a wide region of chromosome 18 (6.6-24.6 Mb) overlapped with one area of strong local ancestry deviation. FTO, with pleiotropic effect on milk and fertility, NOD2 on dairy and NKD1 and SALL1 on fertility traits are located there. Genetic differentiation of RHF and SI (Fst ), an alternative indicator of pre-admixture selection in pure populations, was calculated. No considerable overlap of peaks of local ancestry deviations and Fst was observed. We found two regions with significant signatures of post-admixture selection in this very young composite, applying comparatively stringent significance thresholds. The signals cover relatively large genomic areas and did not allow pinpointing of the gene(s) responsible for the apparent shift in ancestry proportions.

Genome-wide association with residual body weight gain in Bos indicus cattle.

Weight gain is a key performance trait for beef cat-tle; however, attention should be given to the production costs for better profitability. Therefore, a feed efficiency trait based on per-formance can be an interesting approach to improve performance without increasing food costs. To identify candidate genes and ge-nomic regions associated with residual body weight gain (RWG), we conducted a genome-wide association study (GWAS) with 720 Nellore cattle using the GRAMMAR-Gamma association test. We identified 30 significant single nucleotide polymorphisms (SNPs), especially on chromosomes 2, 8, 12, and 17. Several genes and quantitative train loci (QTLs) present in the regions identified were appointed; we highlight DMRT2 (doublesex and mab-3 related tran-scription factor 2), IFFO2 (intermediate filament family orphan 2), LNX2 (ligand of numb-protein X 2), MTIF3 (mitochondrial transla-tional initiation factor 3), and TRNAG-CCC (transfer RNA glycine anticodon CCC). The metabolic pathways that can explain part of the phenotypic variation in RWG are related to oxidative stress and muscle control.

A genomewide association mapping study using ultrasound-scanned information identifies potential genomic regions and candidate genes affecting carcass traits in Nellore cattle.

The aim of this study was to identify candidate genes and genomic regions associated with ultrasound-derived measurements of the rib-eye area (REA), backfat thickness (BFT) and rumpfat thickness (RFT) in Nellore cattle. Data from 640 Nellore steers and young bulls with genotypes for 290 863 single nucleotide polymorphisms (SNPs) were used for genomewide association mapping. Significant SNP associations were explored to find possible candidate genes related to physiological processes. Several of the significant markers detected were mapped onto functional candidate genes including ARFGAP3, CLSTN2 and DPYD for REA; OSBPL3 and SUDS3 for BFT; and RARRES1 and VEPH1 for RFT. The physiological pathway related to lipid metabolism (CLSTN2, OSBPL3, RARRES1 and VEPH1) was identified. The significant markers within previously reported QTLs reinforce the importance of the genomic regions, and the other loci offer candidate genes that have not been related to carcass traits in previous investigations.

Genome-wide association study for feedlot average daily gain in Nellore cattle (Bos indicus).

The genome-wide association study (GWAS) results are presented for average daily gain (ADG) in Nellore cattle. Phenotype of 720 male Bos indicus animals with information of ADG in feedlots and 354,147 single-nucleotide polymorphisms (SNPs) obtained from a database added by information from Illumina Bovine HD (777,962 SNPs) and Illumina BovineSNP50 (54,609) by imputation were used. After quality control and imputation, 290,620 SNPs remained in the association analysis, using R package Genome-wide Rapid Association using Mixed Model and Regression method GRAMMAR-Gamma. A genomic region with six significant SNPs, at Bonferroni-corrected significance, was found on chromosome 3. The most significant SNP (rs42518459, BTA3: 85849977, p = 9.49 × 10(-8)) explained 5.62% of the phenotypic variance and had the allele substitution effect of -0.269 kg/day. Important genes such as PDE4B, LEPR, CYP2J2 and FGGY are located near this region, which is overlapped by 12 quantitative trait locus (QTLs) described for several production traits. Other regions with markers with suggestive effects were identified in BTA6 and BTA10. This study showed regions with major effects on ADG in Bos indicus in feedlots. This information may be useful to increase the efficiency of selecting this trait and to understand the physiological processes involved in its regulation.

Difference in coronary artery intima and media calcification in autopsied patients with chronic kidney disease.

BACKGROUND: In patients with chronic kidney disease (CKD), coronary artery calcification occurs at two distinct sites in the vessel wall: the intima and the media. Arterial media calcification (AMC), a nonocclusive condition, affects hemodynamics differently compared to arterial intima calcification (AIC), which occurs in atherosclerotic plaques. Arterial calcification is considered a cell-regulated process resembling intramembranous bone formation. The purpose of this retrospective observational study was to clarify the morphological differences between AIC and AMC and to evaluate the role of vascular smooth muscle cells (VSMCs) and macrophages in AIC and AMC formation. METHODS: We histologically analyzed 14 tissue specimens from 14 autopsies of patients with CKD Stage 5D who underwent hemodialysis and 5 specimens from 5 patients with CKD Stage 2 - 3 (90 ml/min/1.73 m2 > estimated GFR >= 30 ml/min/1.73 m2). We performed immunohistochemical staining of osteopontin (OPN) as a marker for bone matrix protein, alpha-smooth muscle actin (alphaSMA) for VSMCs, Cbfa1/Runx2 as a marker for osteoblastic differentiation of VSMCs, and CD68 for macrophages. RESULTS: In the CKD 2/3 group, we also found AIC and AMC. OPN and CD68 expression in the CKD 2/3 group was similar to that in the CKD 5D group. Although we did not find Cbfa1/Runx2 positive cell expression in the CKD 2/3 group, we did find it in the CKD 5D group. We found CD68-positive cells predominantly in AIC and absent in AMC in both groups. CONCLUSIONS: These findings suggest that the influence of Cbfa1/Runx2 pathway in coronary artery calcification depends on the CKD Stage. Expression of CD68-positive cells depends on the location of the coronary artery calcification.

Effects of probenecid on the pharmacokinetics of mizoribine and co-administration of the two drugs in patients with nephrotic syndrome.

Probenecid (PRB) is an agent that reduces the systemic level of uric acid, and has the ability to inhibit the renal tubular secretion of agents that are co-administered with it. In this study, we evaluated the effects of PRB co-administered with mizoribine (MZR) on the pharmacokinetics (PK) of MZR in 12 patients with nephrotic syndrome. The elimination rate constant (kel) was used as an indicator of changes in the PK of MZR when the secretion of MZR was inhibited by co-administration of PRB, in order to determine the extent to which MZR was influenced by PRB. In 4 of the 12 patients studied, kel decreased and the biological half-life (t1/2) of MZR was prolonged when co-administered with PRB, in comparison with the values when MZR was used alone, thus revealing that the PK of MZR was influenced by PRB. Co-administration of PRB with MZR appears to be effective in prolonging the biological half-life of MZR and enhancing its effect in patients with nephrotic syndrome, although further studies will be required to determine the optimal dosage of PRB and renoprotective effects.

Sodium-sensitive variability of the antiproteinuric efficacy of RAS inhibitors in outpatients with IgA nephropathy.

AIMS: Inhibition of the renin-angiotensin system (RAS) decreases proteinuria in IgA nephropathy and often retards disease progression. However, its antiproteinuric efficacy varies considerably among patients or different stages in a single patient. We sought for the factor(s) underlying the variation in urinary protein excretion in RAS inhibitor-treated outpatients with IgA nephropathy. PATIENTS: 43 patients with biopsy-proven IgA nephropathy, moderate proteinuria (0.5 - 3.5 g/day), normal to moderately-low estimated GFR (eGFR) (28.6 - 114.2 ml/min/1.73 m2) and normal blood pressure, prehypertension or mild hypertension (systolic/diastolic blood pressures < 160/100 mmHg) were placed on RAS inhibitors following diagnosis. METHOD: Excretion of urinary protein (UprV) and sodium (UNaV), estimated protein intake (EPI) and the mean blood pressure (MBP) were determined on 12 consecutive visits for an average duration of 17.6 months. Analyses were performed to determine which factor(s) influenced the variation in UprV. RESULTS: 14 patients (32.6%) showed a significant correlation between UprV and UNaV, whereas UprV correlated significantly with EPI or MBP in 7 (16.3%) and 3 patients (7.0%), respectively. The 14 patients were characterized by lower eGFR and more extensive glomerulosclerosis and tubulointerstitial damage at baseline than the other 29 patients. The UprV-UNaV correlation was significant in 8 of 12 patients (66.7%) with eGFR < 60 ml/min/1.73 m2 and in 6 of 29 patients (19.4%) with eGFR >= 60 ml/min/1.73 m2 (p < 0.05). The UprV/UNaV regression lines were significantly steeper with more extensive glomerulosclerosis (p < 0.05) and tubulointerstitial damage (p < 0.05) at baseline. The lines also tended to be steeper with lower baseline eGFR (p = 0.062). CONCLUSIONS: These results showed that the antiproteinuric effect of RAS inhibitors becomes susceptible to an increase in urinary sodium excretion as renal function and functioning nephron mass decline with the progression of renal histological damage. Stringent dietary sodium restriction is required to maximize the antiproteinuric effect of RAS inhibitors in outpatients with IgA nephropathy.

Effect of oral mizoribine pulse therapy for frequently relapsing steroid-dependent nephrotic syndrome.

AIM: To evaluate the efficacy of oral mizoribine (MZB) pulse therapy given twice a week for frequently relapsing steroid-dependent nephrotic syndrome (FR-SDNS). SUBJECTS: 16 patients with FR-SDNS with a median age of 11.6 years (range 5.1 â 17.8 years) were enrolled in the study. This study was a Phase II trial. METHODS: The dose of MZB was adjusted to achieve a peak blood level of about 3 microg/ml (10.0-19.7 mg/kg/d, maximum total dose 750 mg) in two divided doses given 2 days a week before a meal. The therapeutic benefits of MZB pulse therapy were assessed based on a comparison of the incidence of relapse (times/year) and the required daily dosage of prednisolone (PSL) before and after therapy. RESULTS: The incidence of relapse after therapy was significantly lower than that before therapy (2.4 A+/- 1.6 vs. 3.4 A+/- 1.1 times/year, p < 0.05), and the required daily dosage of PSL after therapy was lower than that before therapy (0.39 A+/- 0.26 vs. 0.47 A+/- 0.24 mg/kg/d; not significant). During the follow-up period, discontinuation of PSL was possible in 6 of 12 patients who showed a decreased rate of relapse after therapy. The age at entry into the study and the peak blood concentration of MZB of these patients were significantly higher than in four patients who did not show a decreased rate of relapse (12.3 A+/- 4.3 vs. 7.9 A+/- 2.6 years, p < 0.05; 3.00 A+/- 0.93 vs. 1.97 A+/- 0.36 microg/ml, p < 0.005, respectively). No adverse effects were observed in any patients. CONCLUSION: Our results show that MZB pulse therapy is effective in decreasing the frequency of relapse and reducing the required PSL dosage in older pediatric patients with FR-SDNS.

T-type calcium channel blockade as a therapeutic strategy against renal injury in rats with subtotal nephrectomy.

T-type calcium channel blockers have been previously shown to protect glomeruli from hypertension by regulating renal arteriolar tone. To examine whether blockade of these channels has a role in protection against tubulointerstitial damage, we used a stereo-selective T-type calcium channel blocker R(-)-efonidipine and studied its effect on the progression of this type of renal injury in spontaneously hypertensive rats that had undergone subtotal nephrectomy. Treatment with racemic efonidipine for 7 weeks significantly reduced systolic blood pressure and proteinuria. The R(-)-enantiomer, however, had no effect on blood pressure but significantly reduced proteinuria compared to vehicle-treated rats. Both agents blunted the increase in tubulointerstitial fibrosis, renal expression of alpha-smooth muscle actin and vimentin along with transforming growth factor-beta (TGF-beta)-induced renal Rho-kinase activity seen in the control group. Subtotal nephrectomy enhanced renal T-type calcium channel alpha1G subunit expression mimicked in angiotensin II-stimulated mesangial cells or TGF-beta-stimulated proximal tubular cells. Our study shows that T-type calcium channel blockade has renal protective actions that depend not only on hemodynamic effects but also pertain to Rho-kinase activity, tubulointerstitial fibrosis, and epithelial-mesenchymal transitions.

Enzyme replacement therapy in Japanese Fabry disease patients: the results of a phase 2 bridging study.

Fabry Disease (alpha-galactosidase A deficiency) is an X-linked hereditary disorder leading to the pathological accumulation of globotriaosylceramide (GL-3) in lysosomes, particularly in the vascular endothelium of the kidney, heart and brain. We report the results of an open-label phase 2 study that was undertaken to evaluate whether ethnic differences exist that would affect agalsidase beta (Fabrazyme) treatment of Fabry patients in the Japanese population, relative to safety and efficacy. The study design mirrored the design of the completed phase 3 clinical trial that led to approval of the product agalsidase beta. The 13 Japanese, male Fabry patients enrolled in the study received the enzyme replacement therapy over a period of 20 weeks as biweekly infusions. All selected efficacy end points showed improvements that were comparable with findings from the phase 3 study. These improvements included reductions of GL-3 accumulation in both kidney and skin capillary endothelial cells to (near) normal levels (92% of patients). Kidney and plasma GL-3 levels decreased by 51.9% and 100%, respectively, by ELISA. Renal function remained normal. Fabry-associated pain, and quality of life, showed improvement over baseline in multiple categories. Related adverse events were mild or moderate in intensity and mostly infusion-associated (fever and rigors). As expected, IgG antibody formation was observed in 85% of the patients, but had no effect on treatment response. These results suggest that treatment with agalsidase beta is safe and effective in Japanese patients with Fabry disease. With regard to safety and efficacy, no differences were observed as compared to the caucasian population.