Pharmacogenetics of Warfarin in a Diverse Patient Population.

INTRODUCTION: Many warfarin-related genotypes have shown to impact the average daily warfarin (ADW) dose requirements; however, information in non-Caucasian populations is limited. OBJECTIVES: To identify the frequencies of 4 warfarin-related gene polymorphisms in an ethnically diverse patient population and to examine their impact with other clinical variables on ADW dose requirements. METHODS: Patients were recruited from 2 anticoagulation clinics in the Los Angeles area. Blood samples were collected and genotyped for vitamin K epoxide reductase (VKORC1), CYP2C9*2, CYP2C9*3, and CYP4F2 after informed consent. Charts were reviewed to collect demographic, clinical, and warfarin dosing data. RESULTS: A total of 291 patients were included (120 Caucasians, 127 Hispanics, and 44 Asians). In patients with wild-type genotypes for VKORC1, CYP2C9*2, CYP2C9*3, and CYP4F2, the highest warfarin requirement was found in Caucasians, lower in Hispanics, and lowest in Asians. Homozygous VKORC1 variant carriers were detected in 15%, 15%, and 79% in Caucasians, Hispanics, and Asians, respectively. Progressive lowering of ADW doses were associated with each VKORC1 variant in Caucasians and Hispanics, but the results in wild-type/ heterozygote Asians were unclear. CYP2C9 variants were associated with lower ADW doses; frequencies of CYP2C9*2 and CYP2C9*3 mutations were higher in Caucasians than in Hispanics but rare to none in Asians. The frequencies of CYP4F2 variant were similar across all ethnicities, but their impact on warfarin dose requirement were insignificant. Clinical factors such as age, body surface area, history of coronary artery disease, deep vein thrombosis or atrial fibrillation, and concomitant amiodarone or HMG-CoA reductase inhibitors had varying impact on the ADW requirements in the ethnicities studied. CONCLUSIONS: Our study demonstrated differences among 3 ethnic groups in terms of ADW dose requirements and the impact of associated clinical variables. The results suggest that a single model for all ethnicities may not provide the best performance in predicting warfarin dose requirements.

Assessment of Urinary Inhibitor or Promoter Activity in Uric Acid Nephrolithiasis.

PURPOSE: We assessed decreased inhibitor activity or increased promoter activity in the urine of idiopathic uric acid stone formers compared to nonstone formers independent of urinary pH. MATERIALS AND METHODS: A total of 30 idiopathic uric acid stone formers, and 9 obese and 12 lean nonstone formers collected 24-hour urine while on a metabolic diet. Three urine aliquots per subject were used to assess spontaneous nucleation (de novo crystal formation), crystal growth using a 0.1 mg/ml anhydrous uric acid seed and steady-state uric acid solubility (the maximum amount of uric acid dissolvable in urine) using a 5 mg/ml uric acid seed. All experiments were performed for 6 hours at a constant pH of 5.0. Uric acid concentration was measured in filtered aliquots at 0, 3 and 6 hours. RESULTS: At baseline 24-hour urinary pH was significantly lower and uric acid saturation was significantly higher in idiopathic uric acid stone formers. No significant spontaneous nucleation developed and similar uric acid steady-state solubility was reached in the 3 groups. Idiopathic uric acid stone formers and lean nonstone formers showed a similar decrease in uric acid concentration during crystal growth. Obese nonstone formers started with a higher uric acid concentration and consequently demonstrated a greater decrease in the uric acid concentration for crystal growth. CONCLUSIONS: This study suggests that there is no significant difference between idiopathic uric acid stone formers and nonstone formers in promoter or inhibitor activity in whole urine against uric acid stone formation when urine pH is maintained constant. The findings suggest that uric acid stone formation is dictated by high urinary saturation with respect to uric acid, which is driven primarily by low urine pH.

Basal levels of CD34 positive cells in peripheral blood differ between individuals and are stable for 18 months.

BACKGROUND: Detection of basal levels of CD34 progenitor cells is a rare event analysis enumerating cells down to 1 cell/µl. A reproducible analytic approach was used in three independent clinical trials in which multiple sequential assays were obtained from the same individual. METHODS: A 4 color panel combining, HLA-DR, CD34, CD45, and CD11b was used in a dual platform analysis to quantify CD34 progenitor cells in peripheral blood, with quality control focused at the lowest measurements (i.e., basal levels), where assay error is greatest. RESULTS: Repeat testing of individuals every 4 h over the course of 6 days provided a unique opportunity to assess the precision of the analytic technique and identified basal differences between individuals. In a second study, the basal levels were stable for 10 weeks while in a third study the individual differences were maintained for 18 months. This approach was then used to monitor the kinetics of mobilization of CD34 cells following G-CSF stimulation every 4 h. CONCLUSIONS: The differences between individuals in basal levels of CD34 were shown to be a biologic constant, stable for 18 months and not a result of the variability of the assay, shown by low coefficients of variation for each individual. These results can be used to augment a quality control program by monitoring individuals over time to establish intra and inter-laboratory assay precision. In addition, the response of six individuals to G-CSF demonstrated differences in absolute numbers of mobilized CD34 progenitor cells but showed identical kinetics, peaking at 80-110 h.

Effects of the renin angiotensin system on vasculogenesis-related progenitor cells.

The current concept is that there are both cells that integrate into the vasculature, true endothelial progenitor cells (EPC), and cells with hematopoietic markers that support neovascularisation. As identification of the EPC is controversial and studies refer cells that might fall into either pools, we will use the term, vasculogenesis-related progenitor cells (VRPC), for this review. VRPC are considered to be an important target for the treatment of cardiovascular diseases (CVD). Angiotensin II is known to be an important player in neovascularisation and the modulation of renin angiotensin system (RAS) is one of the major pharmacotherapeutic strategies for the treatment of CVD. We will review the effects of different components of the RAS on such VRPC under physiological conditions and in CVD. The reviewed research strongly supports a critical role of the RAS in vasculogenesis and vascular regeneration. Therefore, pharmacological intervention on the components of the RAS does not only target directly end-organ remodelling and blood pressure but also influence tissue healing and/or regeneration by influencing specific progenitor cells. Thus, the interrogation of RAS effects on VRPC will be important in the optimisation of RAS intervention or regenerative therapy.

Comparison of semi-empirical and computer derived methods for estimating urinary saturation of calcium oxalate.

PURPOSE: Estimating calcium oxalate saturation in human urine is critical for nephrolithiasis clinical research and practice. The Joint Expert Speciation System (Mayhem Unit Trust and Council for Scientific and Industrial Research, Pretoria, South Africa) computer program has questioned the validity of the widely used EQUIL 2 program in estimating calcium oxalate urinary saturation. To attempt resolution the computer model based supersaturation index (by the Joint Expert Speciation System) and the relative saturation ratio (by EQUIL 2) were compared with the experimentally derived activity product ratio, that is the ratio of activity products before and after incubating urine with synthetic calcium oxalate. MATERIALS AND METHODS: To determine the experimental conditions required to attain calcium oxalate steady state solubility the filtrate concentration product of calcium and oxalate was determined after incubating 8 urine samples with 2 to 15 mg/ml calcium oxalate for various intervals. In 20 urine samples the activity product ratio of calcium oxalate was compared with the relative saturation ratio and the supersaturation index. RESULTS: Steady state solubility occurred after incubating 15 mg calcium oxalate per ml urine for 72 hours. The mean +/- SD supersaturation index of 4.92 +/- 2.57 in the original urine samples closely approximated the activity product ratio of 5.08 +/- 3.10 but the relative saturation ratio of 7.47 +/- 3.89 was significantly higher than the activity product ratio. The supersaturation index was recalculated after omitting soluble complexes recognized by the Joint Expert Speciation System but not by EQUIL 2, including Ca(2)H(2)(PO(4))(2) and (CaCitPO(4))(4-). The corrected supersaturation index was compared with the relative saturation ratio. After correction the supersaturation index increased to 7.28 +/- 3.81, which was not significantly different from the relative saturation ratio. CONCLUSIONS: The Joint Expert Speciation System is more accurate than EQUIL 2 to estimate calcium oxalate saturation, probably by accounting for Ca(2)H(2)(PO(4))(2), (CaCitPO(4))(4-), and other minor complexes of calcium and oxalate.

Angiotensin-(1-7) stimulates hematopoietic progenitor cells in vitro and in vivo.

Effects of angiotensin (Ang)-(1-7), an AngII metabolite, on bone marrow-derived hematopoietic cells were studied. We identified Ang-(1-7) to stimulate proliferation of human CD34(+) and mononuclear cells in vitro. Under in vivo conditions, we monitored proliferation and differentiation of human cord blood mononuclear cells in NOD/SCID mice. Ang-(1-7) stimulated differentially human cells in bone marrow and accumulated them in the spleen. The number of HLA-I(+) and CD34(+) cells in the bone marrow was increased 42-fold and 600-fold, respectively. These results indicate a decisive impact of Ang-(1-7) on hematopoiesis and its promising therapeutic potential in diseases requiring progenitor stimulation.

New methods of assessing crystal growth and saturation of brushite in whole urine: effect of pH, calcium and citrate.

PURPOSE: Brushite crystallization might be important in stone formation and prevention. To explore this question new methods for the saturation and crystal growth of brushite were devised that are applicable to whole urine without any computer program. MATERIALS AND METHODS: The saturation value (concentration-to-product ratio) was determined by dividing the molar concentration product of Ca ([Ca]) and phosphate ([P]), that is [Ca] x [P], of original urine by the steady state solubility obtained after incubating with an excess of brushite (10 mg/ml) for 5 hours. Crystal growth was measured from the depletion of filtrate ([Ca] x [P]) 3 hours after seeding with brushite (0.25 mg/ml). To test the effect of pH, Ca and citrate the saturation value and crystal growth were determined in 24-hour urine samples from 4 normal volunteers and 2 stone formers, and modified artificially to produce 4 ranges of pH, Ca and citrate by adding acid, base, Ca or citrate. RESULTS: The saturation value and crystal growth of brushite increased with an increase in pH or the Ca concentration but they decreased when the citrate concentration increased. The saturation value correlated strongly with crystal growth. CONCLUSIONS: The new methods of brushite saturation value and crystal growth should help discern how abnormalities in urinary pH, Ca and citrate interact to influence the formation of Ca stones in cases of distal renal tubular acidosis and alkali therapy.