[High-dose vitamin therapy as prophylaxis against porphyria cutanea uremica]. / Hochdosierte Vitamingabe zur Verhinderung einer Porphyria cutanea urämica?

50 Patients with chronic renal failure undergoing hemodialysis with or without porphyria cutanea tarda (PCT)-like skin changes were investigated. The total porphyrin amount in erythrocytes, plasma and dialysate and the distribution of porphyrin metabolites in plasma and dialysate were measured. In plasma, the group of patients with skin changes (referred as PCU = porphyria cutanea uremica) showed significantly increased uroporphyrin levels as compared to the non-symptomatic group. In addition, significant differences concerning the ratio uro-/coproporphyrin in plasma were shown: non-symptomatic patients with 0.87, as opposed to the PCU group with 3.7. Considerable differences between the level of vitamin ingestion were identified between the groups. Patients with PCU took distinctly less vitamins C, E and B than patients without symptoms.

Identification of mutations in the uroporphyrinogen III cosynthase gene in German patients with congenital erythropoietic porphyria.

The porphyrias are heterogeneous disorders arising from predominantly inherited catalytic deficiencies of specific enzymes along the heme biosynthetic pathway. Congenital erythropoietic porphyria is a very rare disease that is inherited as an autosomal recessive trait and results from a profound deficiency of uroporphyrinogen III cosynthase, the fourth enzyme in heme biosynthesis. The degree of severity of clinical symptoms mainly depends on the amount of residual uroporphyrinogen III cosynthase activity. In this study, we sought to characterize the molecular basis of congenital erythropoietic porphyria in Germany by studying four patients with congenital erythropoietic porphyria and their families. Using PCR-based techniques, we identified four different mutations: C73R, a well-known hotspot mutation, the promoter mutation -86A that was also described previously, and two novel missense mutations, designated G236V and L237P, the latter one encountered in the homozygous state in one of the patients. Our data from the German population further emphasize the molecular heterogeneity of congenital erythropoietic porphyria as well as the advantages of molecular genetic techniques as a diagnostic tool and for the detection of clinically asymptomatic heterozygous mutation carriers within families.

Cytochrome p450A1 polymorphisms in a Caucasian population with porphyria cutanea tarda.

Porphyria cutanea tarda (PCT) is the most frequent porphyria in humans. The familial type is in contrast to the sporadic type due to an inherited defect of the uroporphyrinogen-II-decarboxylase (URO-D) and both types need additional porphyrinogens to lead to the clinical manifestation of the disease. Various factors such as xenobiotics (i.e. polycyclic aromatic hydrocarbons), alcohol, hormones and viral liver infections (hepatitis B and C) are known to induce porphyria. Cytochrome p450 enzymes play a crucial role in the metabolism of porphyrogens and therefore might have an important influence on the pathogenesis of hepatic porphyrias. Association of CYP1A2 polymorphisms with susceptibility to both types of PCT has already been described in Danish patients. We investigated 65 caucasian patients with PCT in comparison to a healthy control group concerning the tpe of PCT and the cytochrome p4501A1 polymorphisms (m1, m2 and m4) using polymerase chain reaction (PCR) and a restriction fragment length polymorphism. We found an increased incidence of the m4 polymorphism in the familial type of PCT (odds ratio 5.5, P-value 0.01), whereas the m1 and m2 mutations, might be provoked by a higher susceptibility to porphyrogens via the cytochrome p4501A1 m4 polymorphism.

The 5-aminolevulinic acid-induced porphyrin biosynthesis in benign and malignant cells of the skin.

In fluorescence diagnosis and photodynamic therapy of neoplastic tissues 5-aminolevulinic acid is used to synthesize endogenous porphyrins as photosensitizers. The efficacy of neoplastic tissues to fluorescence diagnosis and photodynamic therapy is thought to be dependent on the total level of intralesional formed porphyrins. The available profiles of porphyrin metabolites in normal and in neoplastic cell lines after administration of 5-aminolevulinic acid vary considerably. Thus, this is the first in-vitro study which compares the porphyrin biosynthesis in normal skin cells (HaCaT, fibroblasts) with melanoma cells (Bro, SKMel-23, SKMel-28). After incubation with 1 mM 5-aminolevulinic acid, kinetics of porphyrin levels and metabolites were determined in the cells and the corresponding supernatants. Exogenous 5-aminolevulinic acid induced porphyrin formation in all cells with maximum values after an incubation period of 16-36 h. Increase of porphyrin levels varied from 10- to 80-fold (SKMel-28>HaCaT>fibroblasts>SKMel-23>>Bro) with minimum 1.5 times higher levels of porphyrins in the supernatants than in the cells. In cells and supernatants protoporphyrin and coproporphyrin were the predominantly formed porphyrin metabolites. Metastatic melanoma cells (SKMel-23, SKMel-28) accumulated much higher porphyrin levels than primary melanoma cells (Bro). In conclusion, by optimizing the treatment modalities, especially the light source, topical photodynamic therapy (PDT) could become a treatment alternative of melanoma metastases in progressive disease.

Aminolevulinic acid: pharmacological profile and clinical indication.

The role of aminolevulinic acid hydrochloride (ALA) in photodynamic therapy (PDT) of in situ neoplasias and tumours of epithelial tumours is steadily increasing and it has been shown to be the drug with most clinical use in PDT. In dermatology, topical PDT with ALA is already postulated to be the treatment of choice for actinic keratoses and superficial basal cell carcinomas. In gastroenterology, pulmonology, uro- and nephrology, neurology and gynaecology ALA has an important role as a photosensitiser not only in the diagnosis of neoplastic tissue but as therapy; first experiences have been made with PDT in these organs. Besides the therapeutic efficacy of this technique, the fluorescence of ALA-induced porphyrins can be effectively used to detect and delineate epithelial and endothelial neoplasms. In dermatology, other indications for ALA-treatment are non-tumoural applications, especially psoriasis, viral-induced diseases, or acne vulgaris. ALA is an effective compound in the diagnosis or therapy of various epithelial and endothelial neoplastic lesions.

Treatment of porphyria cutanea tarda with oral thalidomide.

Eight male patients with overt clinical and biochemical features of porphyria cutanea tarda (PCT) were orally treated with 300 mg/day thalidomide for 1 week and with 200 mg/day for 3 more weeks. Already after the first week of treatment no new vesicles and/or bullae could be observed. Spontaneous blisters completely disappeared, increased skin fragility subsided and skin hyperpigmentation receded about 2 months after completion of therapy, whereas hypertrichosis persisted. There was a rapid decrease in the urinary total porphyrin excretion which reached normal levels in all patients by the end of the fourth week of therapy, whereas the posttreament chromatographic pattern of urinary porphyrins revealed a slight reduction of higher carboxylated porphyrin metabolites and an increase in the amount of the excreted coproporphyrin, as compared to the pretreatment period. Somnolence, intermittent constipation and dry mouth occurred in all patients, 2 patients additionally experienced dizziness. No evidence of peripheral neuropathy could be detected and laboratory investigations revealed no abnormalities, as compared to the pretreatment period. During the 16- to 28-month follow-up of the patients, no clinical or biochemical relapse was observed. In view of the encouraging results of the present investigation, further studies are now warranted in order to definitely answer the question whether oral thalidomide may be regarded as an effective alternative approach to the treatment of PCT.

Alterations of rat liver phospholipid composition induced by oral thalidomide.

In an attempt to elucidate the effects of oral thalidomide on liver phospholipid composition, doses of 1 and 3 mg/kg/day of thalidomide were orally administered to two groups of female Wistar rats (7 animals each), respectively, over a period of 60 days. Control animals (n = 7) received corresponding quantities of the vehicle alone. Chromatographic analysis and quantitative determination of the isolated phospholipid classes revealed statistically significant alterations of phospholipid fractions in the liver of the animals treated with the higher thalidomide dose (3 mg/kg/day). These alterations may be associated with changes in the metabolic activity, ionic transport and cell-cell interactions of the hepatic cellular components.

Effect of reconstituting field-dried and early-harvested sorghum grain on the ensiling characteristics of the grain and on growth performance and carcass merit of feedlot heifers.

The objective of this study was to determine whether reconstituting field-dried and early-harvested sorghum grain affected the fermentation characteristics and feed value of the ensiled grain when fed to feedlot heifers. In Trial 1, sorghum grain was harvested at 14% moisture, rolled, and reconstituted to 25, 30, or 35% moisture, then ensiled in laboratory-scale silos. Lactic acid concentration increased (d 5 to 90) and pH decreased more rapidly (d 3 to 90) as moisture level increased (P < .05). Acetic acid concentration increased (P < .05) with moisture and day postfilling. Concentration of ethanol was highest (P < .05) in the 30 and 35% moisture grains from d 1 to 5, but by d 90 the ethanol concentration in the 25% moisture grain exceeded (P < .05) that of the two higher-moisture grains. Ammonia N concentration was lowest (P < .05) in the 25% moisture grain at all sampling times postfilling. In Trial 2, 288 heifers (BW = 286+/-83 kg) were used to compare the feeding value of rolled, ensiled sorghum grain harvested at 25% moisture to the same grain reconstituted to 30 or 35% moisture. A steam-flaked corn (SFC) diet served as the control. Final live weight; ADG; hot carcass weight; backfat depth; marbling score; kidney, pelvic, and heart fat; and liver abscess score were not affected by grain treatment (P > .10). Dry matter intake was highest (P < .10) for heifers fed the 25 or 30% moisture sorghum grain diets and lowest for those fed the SFC diet; DMI for heifers fed the 35% moisture sorghum grain diet was intermediate. Feeding 35% moisture sorghum grain improved gain efficiency (P < .10) compared with feeding 25 or 30% moisture sorghum grain by 9.0 and 5.7%, respectively. We conclude that reconstituting sorghum grain beyond the typical moisture levels of 25 to 30% would enhance the fermentation characteristics of the ensiled grain and improve gain efficiency in feedlot heifers.

Effects of feeding combinations of steam-flaked grain sorghum and steam-flaked, high-moisture, or dry-rolled corn on growth performance and carcass characteristics in feedlot cattle.

The objective of these two feeding trials was to determine the associative effects of feeding steam-flaked grain sorghum (SFGS) in combination with steam-flaked (SFC), dry-rolled (DRC), or high-moisture (HMC) corn on growth performance and carcass characteristics in feedlot cattle. In Trial 1, 200 yearling heifers were blocked by weight, allotted to 25 pens, and fed one of five finishing diets (77% grain, 15% corn silage, and 8% supplement on a DM basis) for an average of 137 d. The grain combinations were 100:0, 75:25, 50:50, 25:75, and 0:100 SFC:SFGS, respectively. Treatment had no effect on DMI (P > .05), but ADG, gain efficiency, and final live and hot carcass weights decreased linearly (P < .05) as the proportion of SFGS increased in the diet. Carcass backfat, quality grade, and liver abscess score were not affected (P > .05) by treatment. In Trial 2, 306 yearling steers were blocked by weight, allotted to 30 pens, and fed diets that contained 74.5% grain, 10% corn silage, 7.5% soybean meal, 4% tallow, and 4% supplement (DM basis) for an average of 139 d. The grain and grain combinations were 100% DRC, HMC, SFC, or SFGS and a 67%:33% combination of SFGS: DRC or SFGS:HMC. For steers fed diets containing a single source of grain, those fed SFC gained 7% more live weight and had a 7% higher gain efficiency (P < .05) than those fed DRC or HMC. Growth performance of steers fed SFGS was intermediate. Feeding grain combinations (67% SFGS:33% HMC or DRC) resulted in a 5 to 6% positive associative effect (P < .05) for ADG and gain efficiency. Carcass characteristics were not affected (P > .05) by treatment. We concluded that there were significant benefits (positive associative effects) when SFGS was fed in combination with DRC or HMC, but the effects were smaller when SFGS was fed in combination with SFC.

Congenital erythropoietic porphyria.

Congenital erythropoietic porphyria (CEP) is one of the rarest autosomal-recessive disorders of the porphyrin metabolism caused by the homozygous defect of uroporphyrinogen III cosynthase. High amounts of uroporphyrin I accumulate in all cells and tissues, reflected by an increased erythrocyte porphyrin concentration and excretion of high porphyrin amounts in urine and feces. Dermal deposits of uroporphyrin frequently induce a dramatic phototoxic oxygen-dependent skin damage with extensive ulcerations and mutilations. Splenomegaly and hemolytic anemia are typical internal symptoms. Skeletal changes such as osteolysis and calcifications are frequent. Up to date 130 cases of CEP have been published. Splenectomy and erythrocyte transfusions showed some beneficial effect. Bone marrow transplantation was performed in 3 patients and stem cell transplantation in 1. The best therapy is the avoidance of sunlight. We give a report on our latest cases of CEP.

Porphyria cutanea tarda.

Porphyria cutanea tarda (PCT) is the most frequent form of porphyria. The underlying enzymatic defect in PCT is a reduced activity of the enzyme uroporphyrinogen decarboxylase (Uro-D). Four different types of Uro-D disturbances are known. Pseudoporphyrias such as porphyria cutanea uraemica or drug-induced PCT-like skin symptoms are distinguished from PCT. Porphyrinogens such as estrogens or alcohol, or other inducers of P450 isoenzymes provoke PCT. Polymorphisms of P450 isoenzymes, iron overload and infection with hepatitis C virus play an important role in the etiopathogenesis of disease manifestation. Dominant clinical symptoms are bullae, increased cutaneous vulnerability, hypertrichosis and elastosis. Biochemically, total porphyrin levels in urine are increased with a predominance of uroporphyrin and heptacarboxylic porphyrin. Isocoproporphyrin is demonstrable in feces. Best therapeutic strategies are the oral administration of chloroquine 125 mg twice a week and repetitive bloodlettings or the combination of both.

Technical and economic feasibility of reusing disposable perfusion cannulas.

OBJECTIVE(S): The reuse of disposable devices is a potential source of significant cost savings to hospitals. Venous and arterial perfusion cannulas under new and reused conditions were selected to identify the clinical, safety, technical, logistic, and economic issues that must be addressed to realize these savings. METHODS: Single- and dual-stage venous and arterial cannulas from two manufacturers were tested when new, after initial clinical use, and after a single clinical use plus up to nine simulated reuses. Reuse was simulated by end-to-end bending, coupling and uncoupling of the connectors, and by two 1-hour soaks in plasma at 4 degrees and 40 degrees C, respectively. Cannulas were decontaminated and then processed by a peracetic acid-based liquid chemical sterilization system after each use/reuse. Sterilization was validated by eliminating Bacillus subtilis spores from the cannulas on each of five consecutive cycles. Cannulas were tested for physical changes, functional integrity, biocompatibility, and in vivo performance in sheep. A cost analysis was also performed. RESULTS: Sterilization was successfully achieved. Mechanical changes were less than 20% on all variables studied and were undetectable by experienced cardiac surgeons in selective evaluation. No clinically important differences were found between new and reused cannulas, even after nine simulated reuses. Reusing cannulas four times would reduce the cost per procedure from $53 to $19 (64%). CONCLUSIONS: Preliminary data suggest that the perfusion cannulas tested can be safely and efficaciously used five times. Limited reuse of these disposable cannulas is technically feasible and cost-effective. Cannula reuse would result in a small incremental savings; however, with more expensive devices and higher-volume sterilization procedures, the savings could be considerably greater. This program provides a model for evaluation of other single-use medical devices for reuse.

Wheat middlings in high-concentrate diets: feedlot performance, carcass characteristics, nutrient digestibilities, passage rates, and ruminal metabolism in finishing steers.

We conducted two experiments to determine the feeding value and effects on diet digestibilities, passage rates, and ruminal metabolism of wheat middlings (WM) fed as a replacement for either the concentrate or roughage components of finishing diets of steers. In Exp. 1, 120 medium-framed steers were blocked by weight and randomly allocated to one of six treatments of high-concentrate diets: control (0%); 5, 10, or 15% pelleted WM replacing dry-rolled corn (DRC); and 5 or 10% pelleted WM replacing chopped alfalfa hay (ALF) components of the diet. Increasing WM replacement of DRC increased DMI (P < .01) and feed:gain ratio (FG; P < .05) linearly. A 9.2% increase in daily DMI and 10.1% increase in FG were observed at 15% of WM. Daily gain and final weight of the steers were not influenced by WM replacement of DRC. The WM replacement of ALF decreased (P < .01) daily DMI linearly, but it had no effect on ADG, final weight, or FG. In Exp. 2, six medium-framed steers, fitted with ruminal cannulas, were used in a 6 x 6 Latin square design with the same treatments as described in Exp. 1. Dry matter, OM, and starch digestibilities decreased (P < .01) by increasing replacement of DRC with WM, and replacing ALF increased DM and OM digestibilities linearly (P < .01). Wheat middlings could replace only up to 5% of DRC without reducing feed conversion efficiency and diet digestibilities, but complete (100%) or partial (50%) replacement of ALF increased digestibilities of DRC finishing diets.

Photodynamic therapy with 5-aminolaevulinic acid-induced porphyrins of an amelanotic melanoma in vivo.

Of particular interest for photodynamic therapy (PDT) are the endogenously formed and photodynamically active porphyrins produced following topical or systemic application of 65-aminolaevulinic acid (ALA), a haem precursor. Having determined the pharmacokinetics and wavelength dependence of PDT with ALA-induced porphyrins, we analysed the porphyrin metabolites in tumour and surrounding skin. The therapeutic efficacy of PDT using ALA-induced porphyrins was investigated. Amelanotic melanomas (A-Mel-3) were implanted subcutaneously in the back of Syrian golden hamsters (body weight (b.w.), 70-80 g). After 5-7 days, tumours with a volume of approximately 150 mm3 were used for PDT (n = 36). ALA (500 mg kg-1 b.w., pH 6.5) was injected intravenously 45, 90, 150 and 300 min before light irradiation (635 nm, 100 mW cm-2, 100 J cm-2). Tumours with light irradiation only served as controls. The tumour volume was measured after PDT for 28 days. The total porphyrin content was determined in the tumours, the surrounding skin and erythrocytes prior to and 45, 90, 180, 240, 300 and 480 min and 24 h following intravenous injection of ALA (500 mg kg-1 b.w.; n = 32). Porphyrin metabolites were separated by high pressure liquid chromatography (HPLC). Tumour growth was significantly delayed when PDT with ALA was performed 45, 90 or 150 min following intravenous administration. At that time, protoporphyrin (1.8 +/- 0.4 nmol g-1), coproporphyrin (2.2 +/- 0.5 nmol g-1) and uroporphyrin (1.7 +/- 1.4 nmol g-1) were the main metabolites in the tumour tissue. Erythrocytes also contained significant amounts of porphyrins (11.8 +/- 1.3 nmol g-1). The tumour and surrounding skin exhibited a different pattern of porphyrin metabolites. Unexpectedly, a single treatment of PDT with ALA-induced porphyrin resulted in only one complete remission out of six amelanotic melanomas when the final therapeutic outcome was assessed after 28 days. The therapeutic efficacy of PDT with ALA-induced porphyrins can be positively correlated with the fluorescence kinetics previously determined. The analysis of the porphyrin metabolites in amelanotic melanoma by HPLC indicates that the porphyrin accumulation is not due to a decreased activity of ferrochelatase. Moreover, the photodynamic effects may not be mediated solely by porphyrins localized in the tumour parenchyma, but also by significant amounts of porphyrins in the microvasculature. PDT with this endogenous photosensitizer failed to induce complete emission of the treated tumours despite irradiation at the time of maximum porphyrin concentration using the optimum therapeutic wavelength. Thus PDT with ALA-induced porphyrins is less effective in our model relative to that observed for the exogenous photosensitizer Photofrin or synthetic porphycenes after a single treatment.

Ex vivo application of delta-aminolevulinic acid induces high and specific porphyrin levels in human skin tumors: possible basis for selective photodynamic therapy.

In photodynamic therapy with topically applied delta-aminolevulinic acid porphyrins are acting as photosensitizers. The profile of porphyrin metabolites in normal or in neoplastic skin after administration of delta-aminolevulinic acid has not been determined in detail yet. Thus, to study porphyrin biosynthesis in human skin an organ culture model was developed. Explant pieces of normal skin, keratoacanthoma, and basal cell carcinoma were incubated with 1 mM delta-aminolevulinic acid for 36 h. Levels of delta-aminolevulinic acid, porphyrins and porphyrin metabolites were measured in tissues and supernatants. After incubation with delta-aminolevulinic acid, higher porphyrin levels were demonstrated in tumors as compared to normal skin. In supernatants, most of formed porphyrins, preferentially highly carboxylated porphyrin metabolites, were measured. The pattern of synthesized porphyrins differed between normal and neoplastic skin explants. In tissues of basal cell carcinomas protoporphyrin was preferentially shown and tissues of keratoacanthomas were characterized by a predominance of coproporphyrin as compared to normal skin. The results show that explant cultures offer an easy approach to examine the porphyrin biosynthesis of various tissues. The tumor-specific delta-aminolevulinic acid metabolism indicates additional porphyrin metabolites such as coproporphyrin apart from protoporphyrin as effective photosensitizers and may offer a novel approach to tumor-selective photodynamic damage.

Effects of clonidine in a primed rat model of acute hepatic porphyria.

Acute hepatic porphyrias can be induced by several drugs and acute attacks of porphyrias are often associated with severe hypertension. Therefore it is important to know if an antihypertensive drug used has porphyrogenic potency or not. As previously demonstrated in normal rats the alpha-receptor blocker clonidine (CAS 4205-90-7) has no significant influence on the porphyrin metabolism. Pretreatment of rats with 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) or allyl-isopropyl-acetamide (AIA) induces hepatic delta-aminolaevulinic acid synthase (ALA-S) and increases the urinary excretion of porphyrin precursors (ALA and PBG) comparable to the latent phase of acute hepatic porphyrias in humans. Clonidine did not induce hepatic ALA-S or urinary excretion of ALA or PBG in normal as well as in DDC or AIA pretreated rats. Moreover the induction of P4501A1 (7-ethoxyresorufin-O-deethylase) by DDC was abolished by simultaneous application of clonidine. From these findings one can probably conclude that clonidine is a safe drug in human acute hepatic porphyria.

Technical and economic feasibility of reusing disposable perfusion cannulae.

The reuse of disposable devices is a potential source of significant cost savings to hospitals. Venous and arterial perfusion cannulae under new and reused conditions were selected to identify the clinical, safety, technical, logistic, and economic issues that must be addressed to realize these savings. Single- and dual-stage venous and arterial cannulae from two manufacturers were tested when new, after initial clinical use, and after a single clinical use plus up to nine simulated reuses. Reuse was simulated by end-to-end bending, coupling and uncoupling the connectors, and by two 1-hour soaks in plasma at 4 degrees C and 40 degrees C, respectively. Cannulae were decontaminated and then sterilized by a peracetic acid based liquid chemical sterilization system following each use/reuse. Sterilization was validated by eliminating Bacillus subtilis spores from the cannulae on each of five consecutive cycles. Cannulae were tested for physical changes, functional integrity, biocompatibility, and in vivo performance in sheep. A cost minimization analysis was also performed. No clinically important differences were found between new and reused cannulae, even after nine simulated reuses. Mechanical changes were less than 20% on all variables studied and were undetectable by experienced cardiac surgeons in selective evaluation. Sterilization was successfully achieved. Reusing cannulae for times would reduce the cost per procedure from $53 to $19 (64%). Perfusion cannulae tested can be safely and efficaciously used five times. This study suggests that reuse would result in a small incremental savings; however, with more expensive devices and higher-volume sterilization procedures, the savings could be exponentially greater. Although this study demonstrates that it may be technically feasible and cost-effective to reuse disposable cannulae, the U.S. Food and Drug Administration does not sanction the reuse of disposable cannulae.

Congenital erythropoietic porphyria.

Congenital erythropoietic porphyria is a rare autosomal-recessive disorder of the porphyrin metabolism caused by the homozygous defect of uroporphyrinogen III cosynthase. High amounts of uroporphyrin I accumulate in all cells and tissues, reflected by an increased erythrocyte porphyrin concentration and excretion of high porphyrin amounts in urine and feces. Dermal deposits of uroporphyrin frequently induce a dramatic phototoxic oxygen-dependent skin damage with extensive ulcerations and mutilations. Splenomegaly and hemolytic anemia are typical internal symptoms. Skeletal changes such as osteolysis and calcifications are frequent. To date 130 cases of congenital erythropoietic porphyria have been published and are summarized here. Splenectomy, erythrocyte transfusions, and bone marrow transplantation have shown some beneficial effect. The best therapy is the avoidance of sunlight. In the two patients with congenital erythropoietic porphyria described here, oral administration of the oxygen quenchers ascorbic acid and alpha-tocopherol resulted in an improvement in the reduced hemoglobin and erythrocyte concentrations.