Rapid quantification of histamine in human psoriatic plaques using microdialysis and ultra high performance liquid chromatography with fluorescence detection.

Psoriasis is a chronic skin disease resulting from abnormal immune function and is characterized by the presence of scaly psoriatic plaques which are areas of inflammation and excessive skin production. The psoriatic plaques contain mast cells which are increased in number in the uppermost dermis of the psoriatic lesion and which may play a role in the initiation and maintenance of the lesion. These processes are thought to be mediated via the local release of histamine along with other mediators from the mast cells; however their precise role still remains a mystery. Our study involved the development of a rapid and ultra-sensitive liquid chromatographic method for the separation and detection of histamine. To this end a state-of-the-art ultra high pressure liquid chromatography (UHPLC) system incorporating the latest technology in fluorescence detection system was employed which allowed for the rapid and reliable trace level detection of histamine in human derived microdialysate samples. This new reverse phase method utilized a sub-two-micron packed C(18) stationary phase (50 mm × 4.6 mm, 1.8 µm particle size) and a polar mobile phase of ACN:H(2)O:acetic acid (70:30:0.05) (v/v). The column temperature was maintained at (30±2°C), the injection volume was (8 µl), with a flow rate of (1.1 ml/min). Dermal microdialysis was used to collect (20 µl) samples from healthy, peri-lesional and lesional skin regions, in the forearms of a small cohort of subjects (n=6), and the ultra sensitive liquid chromatographic method allowed for nanomolar quantitation of histamine in 6.7 min. To date this represents one of the fastest reported separations of histamine using fluorescence detection with very high chromatographic efficiency (258,000/m) and peak symmetry of (0.88). Prior to sample analysis being performed method linearity, precision and limit of detection (LOD) were investigated. The results showed that intracutaneous histamine measured at 70 min after catheter implantation was (3.44±.52 nmol) (mean±SEM) in non-lesional (control) skin and was not dissimilar to that observed in either lesional (3.10±.76 nmol) or peri-lesional skin (2.24±.20 nmol). A second fraction collected 190 min after implantation also revealed similar levels with no difference in intracutaneous histamine observed between control (2.41±.56 nmol), lesional (2.69±.54 nmol), or peri-lesional skin (2.25±.50 nmol).

Photoprotective behaviour and sunscreen use: impact on vitamin D levels in cutaneous lupus erythematosus.

BACKGROUND: Sun exposure of the skin, independent of dietary sources, may provide sufficient vitamin D in healthy individuals. A recent study of patients with cutaneous lupus erythematosus concluded that over 70% of them restrict their sun exposure. METHODS: We recruited 52 patients with biopsy-proven cutaneous lupus erythematosus to establish whether they are deficient in 25-hydroxyvitamin D [25(OH)D]. We measured their serum 25(OH)D levels during summer months, investigated the effects of several variables on 25(OH)D levels and assessed the role of vitamin D supplementation. RESULTS: An overall mean 25(OH)D level of 63.03 (+/-23.3) nmol/l was obtained. Significantly low values (<25 nmol/l) were recorded in two (3.8%) patients and concentrations below 75 nmol/l were found in 34 (65.4%) patients. 25(OH)D levels were significantly lower among sun avoiders and daily sunscreen users, while significantly higher values were found among those who took cholecalciferol (vitamin D3) supplements. Low values were recorded among those with renal disease despite supplementation with vitamin D3 in some cases. CONCLUSIONS: We suggest that patients with cutaneous lupus erythematous have suboptimal 25(OH)D levels, which are significantly raised by the addition of at least 400 IU/day of cholecalciferol. We recommend supplementation with an active vitamin D analogue in collaboration with a consultant nephrologist, for the subgroup of patients with renal disease.