Chemical characterization of inks in skin reactions to tattoo.

Skin reactions are well described complications of tattooing, usually provoked by red inks. Chemical characterizations of these inks are usually based on limited subjects and techniques. This study aimed to determine the organic and inorganic composition of inks using X-ray fluorescence spectroscopy (XRF), X-ray absorption spectroscopy (XANES) and Raman spectroscopy, in a cohort of patients with cutaneous hypersensitivity reactions to tattoo. A retrospective multicenter study was performed, including 15 patients diagnosed with skin reactions to tattoos. Almost half of these patients developed skin reactions on black inks. XRF identified known allergenic metals - titanium, chromium, manganese, nickel and copper - in almost all cases. XANES spectroscopy distinguished zinc and iron present in ink from these elements in endogenous biomolecules. Raman spectroscopy showed the presence of both reported (azo pigments, quinacridone) and unreported (carbon black, phtalocyanine) putative organic sensitizer compounds, and also defined the phase in which Ti was engaged. To the best of the authors' knowledge, this paper reports the largest cohort of skin hypersensitivity reactions analyzed by multiple complementary techniques. With almost half the patients presenting skin reaction on black tattoo, the study suggests that black modern inks should also be considered to provoke skin reactions, probably because of the common association of carbon black with potential allergenic metals within these inks. Analysis of more skin reactions to tattoos is needed to identify the relevant chemical compounds and help render tattoo ink composition safer.

Vitamin D and Calcium Supplementation Accelerate Vascular Calcification in a Model of Pseudoxanthoma Elasticum.

Arterial calcification is a common feature of pseudoxanthoma elasticum (PXE), a disease characterized by ABCC6 mutations, inducing a deficiency in pyrophosphate, a key inhibitor of calcium phosphate crystallization in arteries. METHODS: we analyzed whether long-term exposure of Abcc6-/- mice (a murine model of PXE) to a mild vitamin D supplementation, with or without calcium, would impact the development of vascular calcification. Eight groups of mice (including Abcc6-/- and wild-type) received vitamin D supplementation every 2 weeks, a calcium-enriched diet alone (calcium in drinking water), both vitamin D supplementation and calcium-enriched diet, or a standard diet (controls) for 6 months. Aorta and kidney artery calcification was assessed by 3D-micro-computed tomography, Optical PhotoThermal IR (OPTIR) spectroscopy, scanning electron microscopy coupled with energy-dispersive X-ray spectroscopy (SEM-EDS) and Yasue staining. RESULTS: at 6 months, although vitamin D and/or calcium did not significantly increase serum calcium levels, vitamin D and calcium supplementation significantly worsened aorta and renal artery calcification in Abcc6-/- mice. CONCLUSIONS: vitamin D and/or calcium supplementation accelerate vascular calcification in a murine model of PXE. These results sound a warning regarding the use of these supplementations in PXE patients and, to a larger extent, patients with low systemic pyrophosphate levels.

Vitamin D and Calcium Supplementation Accelerates Randall's Plaque Formation in a Murine Model.

Most kidney stones are made of calcium oxalate crystals. Randall's plaque, an apatite deposit at the tip of the renal papilla, is considered to at the origin of these stones. Hypercalciuria may promote Randall's plaque formation and growth. We analyzed whether long-term exposure of Abcc6-/- mice (a murine model of Randall's plaque) to vitamin D supplementation, with or without a calcium-rich diet, would accelerate the formation of Randall's plaque. Eight groups of mice (including Abcc6-/- and wild type) received vitamin D alone (100,000 UI/kg every 2 weeks), a calcium-enriched diet alone (calcium gluconate 2 g/L in drinking water), both vitamin D supplementation and a calcium-rich diet, or a standard diet (controls) for 6 months. Kidney calcifications were assessed by 3-dimensional microcomputed tomography, µ-Fourier transform infrared spectroscopy, field emission-scanning electron microscopy, transmission electron microscopy, and Yasue staining. At 6 months, Abcc6-/- mice exposed to vitamin D and calcium supplementation developed massive Randall's plaque when compared with control Abcc6-/- mice (P < 0.01). Wild-type animals did not develop significant calcifications when exposed to vitamin D. Combined administration of vitamin D and calcium significantly accelerates Randall's plaque formation in a murine model. This original model raises concerns about the cumulative risk of vitamin D supplementation and calcium intakes in Randall's plaque formation.

Daily Green Tea Infusions in Hypercalciuric Renal Stone Patients: No Evidence for Increased Stone Risk Factors or Oxalate-Dependent Stones.

Green tea is widely used as a ''healthy'' beverage due to its high level of antioxidant polyphenol compounds. However tea is also known to contain significant amount of oxalate. The objective was to determine, in a cross-sectional observational study among a population of 273 hypercalciuric stone-formers referred to our center for metabolic evaluation, whether daily green tea drinkers (n = 41) experienced increased stone risk factors (especially for oxalate) compared to non-drinkers. Stone risk factors and stone composition were analyzed according to green tea status and sex. In 24-h urine collection, the comparison between green tea drinkers and non-drinkers showed no difference for stone risk factors such as urine oxalate, calcium, urate, citrate, and pH. In females, the prevalence of calcium oxalate dihydrate (COD) and calcium phosphate stones, assessed by infrared analysis (IRS) was similar between green tea drinkers and non-drinkers, whereas prevalence of calcium oxalate monohydrate (COM) stones was strikingly decreased in green tea drinkers (0% vs. 42%, p = 0.04), with data in accordance with a decreased oxalate supersaturation index. In males, stone composition and supersaturation indexes were similar between the two groups. Our data show no evidence for increased stone risk factors or oxalate-dependent stones in daily green tea drinkers.

Fragments and dust after Holmium laser lithotripsy with or without "Moses technology": How are they different?

Urinary stones can be readily disintegrated by Holmium:YAG laser (Holmium laser lithotripsy), resulting in a mixture of small stone dust particles, which will spontaneously evacuate with urine and larger residual fragments (RF) requiring mechanical retrieval. Differences between fragments and dust have not been well characterized. Also, it remains unknown how the recently introduced "Moses technology" may alter stone disintegration products. Three complementary analytical techniques have been used in this study to offer an in-depth characterization of disintegration products after in vitro Holmium laser lithotripsy: stereoscopic microscopy, scanning electron microscopy and Fourier-transform infrared spectroscopy. Dust was separated from fragments based on its floating ability in saline irrigation. Depending on initial crystalline constituents, stone dust either conserved attributes found in larger RFs or showed changes in crystalline organization. These included conversion of calcium oxalate dihydrate towards calcium oxalate monohydrate, changes in carbapatite spectra towards an amorphous phase, changes of magnesium ammonium phosphate towards a differing amorphous and crystalline phase and the appearance of hydroxyapatite on brushite fragments. Comparatively, "Moses technology" produced more pronounced changes. These findings provide new insights suggesting a photothermal effect occurring in Holmium laser lithotripsy. Figure: Appearance of hydroxyapatite hexagons on stone dust collected after Holmium laser lithotripsy of a brushite stone using "Moses technology."

Urothelium proliferation is a trigger for renal crystal deposits in a murine lithogenesis model.

Most mouse kidney stone models induce nephrocalcinosis rather than urolithiasis. The aim of our study was to find an accelerated experimental model in order to study the early events of stone formation, that is, at the time of crystal binding to intrarenal urothelium. C57B6 mice exposed to vitamin D supplements and water containing hydroxyl-L-proline, ammonium chloride and calcium chloride were studied for 42 days. A group receiving urothelial cell mitogen Fibroblast Growth Factor 7 (FGF7) was compared to control group receiving saline. Calcium oxalate monohydrate (COM) crystals were detected in urines by day 2 and within urinary spaces in specialized fornix areas in both groups as soon as day 14 with enhanced deposits in FGF7 group compared to controls at day 21. Urothelial cells proliferation, uroplakin III downregulation and de novo expression of osteopontin receptor CD44 detected in FGF7 group, were delayed in the control group (day 42). Crystal aggregates within specialized fornix areas by day 42 were located in urinary spaces but also within and under a multilayered metaplastic urothelium, simultaneous to macrophages influx. Point of note, administration of a normal diet by day 21 was responsible for a spontaneous crystal clearance. Our data show that under supersaturation conditions, urothelial cell proliferation and calcium oxalate crystal retention occur within specialized fornix areas. Enhanced crystal deposits following FGF7 administration suggest that urothelium proliferation would be a relevant trigger for renal stone formation.

Drug-Induced Kidney Stones and Crystalline Nephropathy: Pathophysiology, Prevention and Treatment.

Drug-induced calculi represent 1-2% of all renal calculi. The drugs reported to produce calculi may be divided into two groups. The first one includes poorly soluble drugs with high urine excretion that favour crystallisation in the urine. Among them, drugs used for the treatment of patients with human immunodeficiency, namely atazanavir and other protease inhibitors, and sulphadiazine used for the treatment of cerebral toxoplasmosis, are the most frequent causes. Besides these drugs, about 20 other molecules may induce nephrolithiasis, such as ceftriaxone or ephedrine-containing preparations in subjects receiving high doses or long-term treatment. Calculi analysis by physical methods including infrared spectroscopy or X-ray diffraction is needed to demonstrate the presence of the drug or its metabolites within the calculi. Some drugs may also provoke heavy intra-tubular crystal precipitation causing acute renal failure. Here, the identification of crystalluria or crystals within the kidney tissue in the case of renal biopsy is of major diagnostic value. The second group includes drugs that provoke the formation of urinary calculi as a consequence of their metabolic effects on urinary pH and/or the excretion of calcium, phosphate, oxalate, citrate, uric acid or other purines. Among such metabolically induced calculi are those formed in patients taking uncontrolled calcium/vitamin D supplements, or being treated with carbonic anhydrase inhibitors such as acetazolamide or topiramate. Here, diagnosis relies on a careful clinical inquiry to differentiate between common calculi and metabolically induced calculi, of which the incidence is probably underestimated. Specific patient-dependent risk factors also exist in relation to urine pH, volume of diuresis and other factors, thus providing a basis for preventive or curative measures against stone formation.

Calcium and vitamin D have a synergistic role in a rat model of kidney stone disease.

Vitamin D supplementation in humans should be accompanied by calcium administration to avoid bone demineralization through vitamin D receptor signaling. Here we analyzed whether long-term exposure of rats to vitamin D supplementation, with or without a calcium-rich diet, would promote kidney stone formation. Four groups of rats received vitamin D alone (100,000 UI/kg/3 weeks), a calcium-enriched diet alone, both vitamin D supplementation and calcium-enriched diet, or a standard diet (controls) for 6 months. Serum and urine parameters and crystalluria were monitored. Kidney stones were assessed by 3-dimensional micro-computed tomography, infrared spectroscopy, von Kossa/Yasue staining, and field emission scanning electron microscopy. Although serum calcium levels were similar in the 4 groups, rats receiving vitamin D had a progressive increase in urinary calcium excretion over time, especially those receiving both calcium and vitamin D. However, oral calcium supplementation alone did not increase urinary calcium excretion. At 6 months, rats exposed to both calcium and vitamin D, but not rats exposed to calcium or vitamin D alone, developed significant apatite kidney calcifications (mean volume, 0.121 mm(3)). Thus, coadministration of vitamin D and increased calcium intake had a synergistic role in tubular calcifications or kidney stone formation in this rat model. Hence, one should be cautious about the cumulative risk of kidney stone formation in humans when exposed to both vitamin D supplementation and high calcium intake.

Characterisation of calcium phosphate crystals on calcified human aortic vascular smooth muscle cells and potential role of magnesium.

BACKGROUND: Cardiovascular disease including vascular calcification (VC) remains the leading cause of death in patients suffering from chronic kidney disease (CKD). The process of VC seems likely to be a tightly regulated process where vascular smooth muscle cells are playing a key role rather than just a mere passive precipitation of calcium phosphate. Characterisation of the chemical and crystalline structure of VC was mainly led in patients or animal models with CKD. Likewise, Mg2+ was found to be protective in living cells although a potential role for Mg2+ could not be excluded on crystal formation and precipitation. In this study, the crystal formation and the role of Mg2+ were investigated in an in vitro model of primary human aortic vascular smooth muscle cells (HAVSMC) with physical techniques. METHODOLOGY/PRINCIPAL FINDINGS: In HAVSMC incubated with increased Ca x Pi medium, only calcium phosphate apatite crystals (CPA) were detected by Micro-Fourier Transform InfraRed spectroscopy (µFTIR) and Field Effect Scanning Electron Microscope (FE-SEM) and Energy Dispersive X-ray spectrometry (EDX) at the cell layer level. Supplementation with Mg2+ did not alter the crystal composition or structure. The crystal deposition was preferentially positioned near or directly on cells as pictured by FE-SEM observations and EDX measurements. Large µFTIR maps revealed spots of CPA crystals that were associated to the cellular layout. This qualitative analysis suggests a potential beneficial effect of Mg2+ at 5 mM in noticeably reducing the number and intensities of CPA µFTIR spots. CONCLUSIONS/SIGNIFICANCE: For the first time in a model of HAVSMC, induced calcification led to the formation of the sole CPA crystals. Our data seems to exclude a physicochemical role of Mg2+ in altering the CPA crystal growth, composition or structure. Furthermore, Mg2+ beneficial role in attenuating VC should be linked to an active cellular role.

The status of strontium in biological apatites: an XANES/EXAFS investigation.

Osteoporosis represents a major public health problem through its association with fragility fractures. The public health burden of osteoporotic fractures will rise in future generations, due in part to an increase in life expectancy. Strontium-based drugs have been shown to increase bone mass in postmenopausal osteoporosis patients and to reduce fracture risk but the molecular mechanisms of the action of these Sr-based drugs are not totally elucidated. The local environment of Sr(2+) cations in biological apatites present in pathological and physiological calcifications in patients without such Sr-based drugs has been assessed. In this investigation, X-ray absorption spectra have been collected for 17 pathological and physiological calcifications. These experimental data have been combined with a set of numerical simulations using the ab initio FEFF9 X-ray spectroscopy program which takes into account possible distortion and Ca/Sr substitution in the environment of the Sr(2+) cations. For selected samples, Fourier transforms of the EXAFS modulations have been performed. The complete set of experimental data collected on 17 samples indicates that there is no relationship between the nature of the calcification (physiological and pathological) and the adsorption mode of Sr(2+) cations (simple adsorption or insertion). Such structural considerations have medical implications. Pathological and physiological calcifications correspond to two very different preparation procedures but are associated with the same localization of Sr(2+) versus apatite crystals. Based on this study, it seems that for supplementation of Sr at low concentration, Sr(2+) cations will be localized into the apatite network.