The binding of 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) with the aryl hydrocarbon (AH) receptor and subsequent changes in gene expression have been studied intensively, but the mechanisms by which these lead to toxicity are unclear. We investigated the influence of iron, previously implicated in TCDD-induced hepatic porphyria, in mice with alleles of Ahr that encode receptors with varied affinity for TCDD. The administration of iron to Ahrb-1 C57BL/6J (AH-responsive) mice before a single dose of TCDD (75 micrograms/kg) markedly potentiated not only the hepatic porphyria but also general hepatocellular damage and elevation of plasma hepatic enzymes. The formation of hydroxylated and peroxylated derivatives of uroporphyrins formed from uroporphyrinogen and the induction of a mu-glutathione transferase (GST) were consistent with the operation of an oxidative mechanism. In a comparison of C57BL/6J mice with Ahrb-2 BALB/c (AH-responsive) and Ahrd SWR and DBA/2 (AH-nonresponsive) mice, iron overcame the weak hepatic porphyria and toxicity responses in BALB/c and SWR strains but not in DBA/2. CYP1A isoforms are strongly implicated in the mechanism of porphyria, but activities were lowered by 20-30% with iron treatment, and a comparison of levels between strains did not fully account for the resistance of DBA/2 mice. Studies with the use of gel shift assays and cytosolic aconitase of the capacity of the iron regulatory protein controlling the translation of some iron metabolism proteins showed a significant difference between C57BL/6J and DBA/2 mice after the administration of TCDD. We conclude that iron potentiates both the hepatic porphyria and toxicity of TCDD in susceptible mice in an oxidative process with disturbance of iron regulatory protein capacity. Iron even overcomes the AH-nonresponsive Ahrd allele in the SWR strain but not in DBA/2 mice, which remain resistant.

The isomeric compositions of the heptacarboxylic, hexacarboxylic and pentacarboxylic porphyrinogens formed by incubation of porphobilinogen with human red-cell haemolysates have been analysed and compared with those derived from incubation with chemically prepared uroporphyrinogen III as substrate. The results indicated that when supplied with an excess (3.7 microM) of exogenous uroporphyrinogen III, uroporphyrinogen decarboxylase utilized the substrate at random and a mixture of isomers was produced; whereas with uroporphyrinogen III generated enzymically from porphobilinogen as substrate a clockwise decarboxylation sequence was observed, resulting in the formation of intermediates mainly with the ring-D, rings-AD and rings-ABD acetate groups decarboxylated. Using [14C]uroporphyrinogen III as substrate at low concentrations (0.01-0.5 microM) also led to preferential decarboxylation of the ring-D acetate group. It was concluded that the order of uroporphyrinogen III decarboxylation is substrate-concentration-dependent, and under normal physiological conditions enzymic decarboxylation is most probably orderly and clockwise, starting at the ring-D acetate group.

Introduction: Radix Salviae (Dan-shen in pinyin), a classic Chinese herb, has been extensively used to treat diabetic retinopathy in clinical practice in China for many years. However, the pharmacological mechanisms of Radix Salviae remain vague. The aim of this study was to decrypt the underlying mechanisms of Radix Salviae in the treatment of diabetic retinopathy using a systems pharmacology approach.Methods: A network pharmacology-based strategy was proposed to elucidate the underlying multi-component, multi-target, and multi-pathway mode of action of Radix Salviae against diabetic retinopathy. First, we collected putative targets of Radix Salviae based on the Traditional Chinese Medicine System Pharmacology database and a network of the interactions among the putative targets of Radix Salviae and known therapeutic targets of diabetic retinopathy was built. Then, two topological parameters, "degree" and "closeness certainty" were calculated to identify the major targets in the network. Furthermore, the major hubs were imported to the Database for Annotation, Visualization and Integrated Discovery to perform a pathway enrichment analysis. Results:A total of 130 nodes, including 18 putative targets of Radix Salviae, were observed to be major hubs in terms of topological importance. The results of pathway enrichment analysis indicated that putative targets of Radix Salviae mostly participated in various pathways associated with angiogenesis, protein metabolism, inflammatory response, apoptosis, and cell proliferation. The putative targets of Radix Salviae (vascular endothelial growth factor, matrix metalloproteinases, plasminogen, insulin-like growth factor-1, and cyclooxygenase-2) were recognized as active factors involved in the main biological functions of treatment, which implied that these were involved in the underlying mechanisms of Radix Salviae on diabetic retinopathy. Conclusions:Radix Salviae could alleviate diabetic retinopathy via the molecular mechanisms predicted by network pharmacology. This research demonstrates that the network pharmacology approach can be an effective tool to reveal the mechanisms of traditional Chinese medicine from a holistic perspective.

The isomeric composition of type-III heptacarboxylic porphyrinogens derived from decarbosylation of uroporphyrinogen III by erythrocyte uroporphyringogen decarboxylase was analysed by h.p.l.c. with electrochemical detection. All four possible isomers were identified, and there were little differences in the proportion of isomers formed by erythrocytes from normal subjects and from patients with sporadic porphyria cutanea tarda. The results provide conclusive evidence that the normal decarboxylation pathway is random in nature, and the fourth isomer only increases when enzyme abnormality is found.