Detection of p53 and histopathological classification of skin tumours induced by halogen lamps in hairless mice.

Skin lesions induced by exposure of three strains of female hairless mice to the light emitted by uncovered halogen quartz lamps were subjected to histopathological analysis. We examined 170 representative specimens out of a total of 597 skin lesions, i.e. 38 out of 74 SKH-1 mice, 110 out of 472 MF-1 mice, and 42 out of 51 C3H mice. The results provided evidence of various types of alterations, including preneoplastic changes, such as epidermal hyperplasia, and benign tumours, such as papillomas, as well as tumours with an increasing degree of malignancy, i.e., keratoacanthoma-like tumours, appendage/basal tumours, actinic keratoses/carcinomas in situ, and squamocellular carcinomas. SKH-1 was the most sensitive strain to the far-ultraviolet wavelengths delivered by halogen lamps, as shown not only by the shortest latency time and the highest multiplicity of skin lesions but also by the highest frequency of malignant tumours. Some areas of atypical melanocyte proliferation were only detected in C3H pigmented mice. Eighty-two of the lesions excised from MF-1 mice were additionally examined for p53 protein by immunohistochemical methods. Formalin-fixed, paraffin-embedded sections and frozen sections were analyzed in parallel by using polyclonal CM-1 antibody and monoclonal PAb240 antibody, respectively. A positive response for p53 was only observed in squamocellular carcinomas, and was related to the size of cancers; in fact, six out of 10 cancers of 10-30 mm in diameter were positive, whereas all 16 cancers of 2-9 mm in diameter were negative. All six positive squamocellular carcinomas were detected by using the CM-1 antibody, which recognizes both wild-type and mutant forms of p53 protein, and five of them were also positive with the PAb 240 antibody, which only recognizes the mutant form. Thus, p53 mutation appears to be a late event in the development of halogen-induced skin tumours in hairless mice, requiring a severe degree of malignancy and an advanced stage of the neoplastic mass growth.

Short report: erythema nodosum leprosum lymphadenitis.

A case of isolated erythema nodosum lymphadenitis involving the paravertebral, intercostal, and cervical lymph nodes without concomitant skin involvement is reported in a 62-year-old male patient under treatment for lepromatous leprosy.

beta2-Glycoprotein I - dependent anticardiolipin antibodies in ischemic stroke.

Our study was undertaken to investigate whether beta2-glycoprotein I (GPI) is relevant for enhancing anticardiolipin antibody (aCL) binding in stroke patients, in particular view of vascular risk factors and recurrence of previous cerebral ischemic events. One-hundred and twenty-one sera from patients with ischemic stroke and 154 control sera from patients with non-ischemic neurological disorders (n = 43) and healthy subjects (n = 111) were included in the study. All sera were tested for either GPI-independent aCL and GPI-dependent aCL. GPI-independent aCL were detected in two (1.7%) stroke patients. When GPI was added to the assay system, 13 (10.8%) sera were positive. Of the 43 sera in the neurological control group one was positive for both GPI-independent aCL and GPI-dependent aCL. Multiple linear regression in the stroke group revealed that GPI-dependent aCL are marginally associated with sex, prior TIA/strokes and embolism. More importantly, GPI-dependent aCL were significantly more frequent in stroke patients with a history of prior TIA/strokes compared with patients with a first ischemic stroke (p = 0.029). The present study demonstrates that aCL in stroke patients are of the GPI-dependent type and emphasizes the importance of adding GPI in the immunoassay for the optimal detection of aCL. Furthermore, it supports the recommendation of testing aCL in those patients who experience unexplained recurrent cerebral ischemic events.

Induction of alopecia in mice exposed to cigarette smoke.

Besides being responsible for a high proportion of those chronic degenerative diseases that are the leading causes of death in the population, tobacco smoking has been associated with skin diseases. Smoke genotoxicants are metabolized in hair follicle cells, where they form DNA adducts and cause DNA damage. The suspicion was raised that, in humans, a link may exist between smoking and both premature grey hair and hair loss. In order to check this hypothesis, we carried out a study in C57BL/6 mice exposed whole-body to a mixture of sidestream and mainstream cigarette smoke. After 3 months exposure, most mice developed areas of alopecia and grey hair, while no such lesions occurred either in sham-exposed mice or in smoke-exposed mice receiving the chemopreventive agent N-acetylcysteine with drinking water. Cell apoptosis occurred massively in the hair bulbs at the edge of alopecia areas. Smoke-exposed mice had extensive atrophy of the epidermis, reduced thickness of the subcutaneous tissue, and scarcity of hair follicles. On the whole, exposure to smoke genotoxic components appears to alter the hair cycle with a dystrophic anagen pattern. Although this mechanism is different from that of genotoxic cytostatic drugs, N-acetylcysteine appears to exert protective effects in both conditions.

beta 2-Glycoprotein I-dependence of anticardiolipin antibodies in multibacillary leprosy patients.

This study was undertaken to investigate the influence of beta 2-glycoprotein I (GPI) on anticardiolipin antibody (aCL) titration in leprosy. The study group consisted of 140 sera from patients with multibacillary leprosy (46 borderline, 94 lepromatous). The group included newly diagnosed, previously untreated patients, patients under treatment and patients released from treatment. GPI addition enhanced significantly the aCL titres in sera from lepromatous leprosy but not in those from borderline leprosy. Moreover, when the patients were classified according to their bacteriological status, aCL titres were found to be significantly higher in skin smear positive patients compared to bacteriologically negative patients. Thus, the present study demonstrates that aCL in multibacillary leprosy patients are mainly of the GPI-dependent type and emphasizes the importance of GPI addition for aCL titration in leprosy.

Identification of sequence similarities between Mycobacterium leprae and the myelin proteolipid protein by computational analysis.

This study was undertaken under the assumption that antigenic mimicry plays a role in the pathogenesis of neuropathy in leprosy, a unique feature among mycobacterial diseases. The SWISS-PROT protein sequence databank was scanned using a computer programme based on an identity matrix algorithm, to identify common amino acid regions between human myelin and mycobacterial proteins. The highlighted motifs were back-tested against a database of MHC-binding peptides (MHCPEP). Of the 28 common sequences between mycobacterial and human myelin proteins, only two were found to yield some matches with MHC-presenting peptides. Both motifs were from M.leprae. The myelin proteolipid protein was the human protein containing the identified similarities. We believe that this theoretical approach can provide a way to predict potentially "mimetic" motifs by search for antigenic regions in protein sequence databases without screening a large number of synthetic peptides.

Timed surgery for treatment of angiofibromas in tuberous sclerosis.

Angiofibromas are neoformations frequently occurring on the face of patients affected by tuberous sclerosis. Traditional methods of treating angiofibromas, such as curettage, dermabrasion, chemical peeling, cryosurgery, and laser are not entirely successful as they frequently lead to scarring and pigmentary change. We report the successful use of timed surgery, a novel electrosurgical procedure, in treating multiple angiofibromas. The technique is easy to use and very cost effective.

Age as limiting factor of the efficacy of itraconazole for treatment of onychomycosis.

A clinical study was carried out involving 24 patients between 18 and 64 years of age with disto-lateral onychomycosis of finger and/or toenail in whom intermittent therapy with itraconazole was orally administered for 4 months. A follow-up period of 9 months was instituted to monitor the efficacy of the treatment. Clinical and mycological outcome at the end of the study was correlated with a checklist that included age and sex of the patient, site of infection and species of the causative agent. Nails that were free of disease in both gross and mycological examinations were achieved in 58% of our patients. Age was shown to be the only parameter, among those taken into consideration, that was correlated with the cure rate. The association was found to be statistically significant.

Acrokeratoelastoidosis of Costa: a primary disease of the elastic tissue?

A 24-year-old black woman from Uganda was seen for treatment of multiple papules on her hands and feet. The lesions corresponded microscopically to foci of hyperkeratosis and acanthosis. Acid orcein stain revealed marked elastic fiber fragmentation. Acrokeratoelastoidosis of Costa (AKE) was diagnosed. The same damage to the elastic fibers was also present in an additional specimen from grossly uninvolved skin. On electron microscopy there were pronounced changes of the elastic fibers with elastolysis in both specimens. This case with generalized damage of the dermal elastic tissue supports the view that elastorrhexis is the key feature of AKE. Accordingly, the disease could be regarded as a primary elastic tissue disorder. The marginal acral keratoderma that is seen in AKE patients could represent epidermal changes secondary to chronic trauma.

Antibodies to beta(2)-glycoprotein I in ischemic stroke.

Antibodies to beta(2)-glycoprotein (beta(2)-GPI) have been associated with recurrent thrombotic events in patients with systemic lupus erythematosus. The present study investigated the prevalence of antibodies to beta(2)-GPI in an unselected group of patients with ischemic stroke. One hundred and twenty-one sera from patients with ischemic stroke and 174 control sera from patients with nonischemic neurological disorders (n = 43) and healthy subjects (n = 131) were tested for antibodies to beta(2)-GPI by a solid-phase ELISA. Twenty-nine stroke patients (24%) had antibodies to beta(2)-GPI. Of the 43 patients in the neurological control group, 2 were positive. For comparison between the groups, Fisher's exact test was used for categorical variables and ANOVA for antibody titers. Antibody levels and frequencies of positivity were significantly different between the study groups. None of the sera from the healthy control group had abnormal antibody levels. When risk factors and associated diseases were taken into account, a marginal association was found between the presence of antibodies to beta(2)-GPI and hypertension (p = 0.036). This study demonstrates a significant prevalence of antibodies to beta(2)-GPI in an unselected stroke population.

Effects of fixation on polymerase chain reaction detection of Mycobacterium leprae.

The effects of standard fixatives (10% neutral buffered formalin, ethanol and mercury based) on the detection of Mycobacterium leprae DNA by the polymerase chain reaction (PCR) were studied. Mercury-based fixatives (Zenker's and Carnoy-Lebrun's fluids) strongly inhibited PCR amplification of M. leprae DNA. Ten percent neutral buffered formalin was inhibitory, but significant inhibition was observed only when fixation times exceeded 24 h. Ethanol-based fixatives provided the best medium for holding specimens for subsequent PCR with both free bacilli and skin biopsy specimens containing M. leprae. The M. leprae-specific, 360-bp region of the 18-kDa protein gene could be amplified from paraffin-embedded sections of formalin-fixed skin biopsy specimens from patients with either multibacillary or paucibacillary infections when proper fixation conditions were used. Results of the study demonstrate that tissues properly fixed with two standard fixatives (10% neutral buffered formalin and 50 or 70% ethanol) can be analyzed by PCR for the presence of M. leprae with no loss in specificity and only minimal diminution in sensitivity compared with the specificities and sensitivities obtained by use of freshly prepared, unfixed specimens.

Detection of Mycobacterium leprae and the potential for monitoring antileprosy drug therapy directly from skin biopsies by PCR.

An improved protocol for PCR analysis of Mycobacterium leprae-infected tissues, based on enzymatic lysis, has been developed and used to demonstrate the feasibility of using PCR for detecting M. leprae in routine skin biopsies taken from leprosy patients throughout the clinical spectrum. Of 92 multibacillary patients tested, 99% were PCR-positive using gel electrophoresis or DNA hybridization to detect the amplified product. Similar analysis of paucibacillary patients, in which only one of 27 biopsies had demonstrable AFB microscopically, gave a positivity rate of 74%. No PCR signals were demonstrated from skin biopsies from seven patients with non-leprosy dermatoses and one AIDS patient with a disseminated atypical mycobacteriosis. Evaluation of leprosy patients with antileprosy drug therapy prior to biopsy demonstrated that PCR signals were either greatly diminished or absent after 2 months of continuous antibiotic therapy. PCR was also able to detect the presence of M. leprae in tissues of patients receiving antibacterial therapy when patients were suspected of harbouring drug-resistant M. leprae.

US and MR imaging of peripheral nerves in leprosy.

OBJECTIVE: To analyze peripheral nerves with ultrasonography (US) and magnetic resonance imaging (MR) in leprosy and assess the role of imaging in leprosy patients. DESIGN AND PATIENTS: Fifty-eight nerves with abnormal clinical features or electromyograms were examined in 23 leprosy patients by means of gray-scale US, Doppler US and MR imaging. Image analysis included: measurement of nerve cross-sectional area; assessment of nerve structure and MR signal intensities; identification of nerve compression within osteofibrous tunnels; detection of endoneural color flow signals and Gd-DTPA enhancement. Correlations were made with clinical findings and a control group of 20 subjects. Fourteen nerves in active reversal reaction were followed up after therapy. RESULTS: Leprosy nerves were classified into three groups based on imaging appearance: group I consisted of 17 normal-appearing nerves; group II, of 30 enlarged nerves with fascicular abnormalities; group III, of 11 nerves with absent fascicular structure. Group II nerves were from patients subjected to reversal reactions; 75% of patients with group III nerves had a history of erythema nodosum leprosum. Nerve compression in osteofibrous tunnels was identified in 33% of group II and 18% of group III nerves. Doppler US and MR imaging were 74% and 92% sensitive in identifying active reactions, based on detection of endoneural color flow signals, long T2 and Gd enhancement. In 64% of cases, follow-up studies showed decreased color flow and Gd uptake after steroids and decompressive surgery. CONCLUSIONS: US and MR imaging are able to detect nerves abnormalities in leprosy. Active reversal reactions are indicated by endoneural color flow signals as well as by an increased T2 signal and Gd enhancement. These signs would suggest rapid progression of nerve damage and a poor prognosis unless antireactional treatment is started.