Gold: an old drug still working in refractory pemphigus

BACKGROUND: The mainstay of treatment for pemphigus is systemic corticosteroids. Different adjuvants have been used to reduce side-effects of long-term corticotherapy. Gold is an anti-inflammatory drug used in autoimmune diseases, whose use has waned with the advent of new immunosuppressive agents. OBJECTIVE: To study the outcome of the use of intramuscular gold treatment of pemphigus vulgaris refractory to previous therapies. METHODS: Thirteen patients with pemphigus vulgaris who had failed to respond to several prior therapies were treated with aurothiomalate, as a steroid-sparing agent. Patients were monitored to assess disease activity and gold toxicity. RESULTS: Seven patients achieved complete remission. Four patients were able to taper prednisone doses, although pemphigus flared when prednisone was discontinued or reduced. Toxicity was observed in the other two patients. CONCLUSIONS: In 53.4% of the patients, the use of chrysotherapy resulted in the complete clearing of the disease, discontinuation of all systemic therapies and induced a long-term clinical remission. Prednisone doses were able to be reduced in the remaining 46.6%. Any side-effects were reversible with drug discontinuation. Gold therapy showed efficacy as a secondary line treatment in refractory pemphigus vulgaris.     

Folic acid in the treatment of acute watery diarrhoea in children: a double-blind, randomized, controlled trial

One-hundred and six male children aged 6-23 months with a history of acute watery diarrhoea of less than 72 h duration were randomized to receive either folic acid in a dose of 5 mg at 8-h intervals or placebo for 5 d. There were 54 children in the folic acid group and 52 in the placebo group. The admission characteristics were comparable between the two groups. No significant differences were observed in the intake of oral rehydration solution or stool output between the groups. The mean ± SD of total stool output (g kg⁻¹) was 532 ± 476 vs 479 ± 354 and the duration (h) of diarrhoea was 108 ± 68 vs 103 ± 53 in the folic acid vs placebo group, respectively. The findings, therefore, should have a positive influence on preventing the inappropriate use of folic acid in acute diarrhoea.     

Progression and regression of cervical lesions: Review of smears from women followed without initial biopsy or treatment

Cervical smears were reviewed from patients in whom a cytological abnormality was followed, after an interval without interference, either by regression to `negative' or else by progression to invasive carcinoma. Twenty-eight cases were from a previously analysed series with positive smears and an interval of at least two years before investigation, resulting from refusal or failure to trace. Slides were also reviewed from 25 cases in which `positive' smears had regressed to negative without escaping from surveillance, and from 10 patients subsequently developing invasive carcinoma whose previous slides, taken several years earlier, showed abnormalities on review. None of these 63 patients had any biopsy or other surgical procedure to the cervix between the initial smear and the outcome.

Slides showing `superficial cell dyskaryosis' and/or well-differentiated `parabasal cell dyskaryosis' were found only among the groups with subsequent regression. Those showing dissociated poorly differentiated dyskaryotic parabasal cells regressed to negative in two cases and progressed to invasion in nine. This suggests that many examples of spontaneous regression correspond to mild dysplasias which are not precancerous, and overdiagnosis must often have resulted in unnecessary surgical procedures in the past.

`Regressing' and `progressing' groups both included cases in which the spatula had removed coherent pieces of undifferentiated epithelium. These are difficult to interpret cytologically. In nine of them (including four which regressed) the cytological picture was that of carcinoma in situ. The remainder (14 cases) were probably examples of reserve cell hyperplasia, and it is noteworthy that, of the 21 cases subsequently progressing to invasive carcinoma, five were preceded by appearances of this type. It is concluded that cell aggregates suggesting an unusual degree of reserve cell hyperplasia are a danger signal and require careful surveillance.

     

Frequent co-localization of Cox-2 and laminin-5 gamma2 chain at the invasive front of early-stage lung adenocarcinomas

Laminin-5 is an extracellular matrix protein that plays a key role in cell migration and tumor invasion. Cox-2 is an induced isoform of cyclooxygenases that plays an important role in carcinogenesis, suppression of apoptosis, angiogenesis, and metastasis of colon cancer. We report frequent co-expression of cox-2 and laminin-5 at the invasive front of early-stage lung adenocarcinomas. We investigated the expression of cox-2 and laminin-5 immunohistochemically in 102 cases of small-sized lung adenocarcinoma (maximum dimension, 2 cm or less). Cox-2 and laminin-5 were expressed in 97 (95.1%) and 82 (80.4%) cases, respectively. Both were preferentially localized in cancer cells at the cancer-stroma interface, although cox-2 tended to show a diffuse staining pattern in some cases. A comparison of their staining patterns revealed a striking similarity in their distribution in 24 cases, and a partial overlap between their localization in another 20 cases. Moreover, an overall correlation was found between the expression levels of cox-2 and laminin-5 (P = 0.018). To gain insight into the mechanisms that regulate the expression of these proteins, we additionally studied their expression in 58 cases of stage I lung adenocarcinoma, in which p53 status was determined by immunohistochemistry, polymerase chain reaction-single strand conformation polymorphism analysis, and direct sequencing. The results showed that tumors with mutant p53 tended to express more cox-2 than those with wild-type p53 (P = 0.080). Also, tumors that overexpressed p53 had higher levels of cox-2 and laminin-5 than those without p53 overexpression (P = 0.032 and 0.047, respectively). Further immunohistochemical analysis showed that tumors that overexpressed both epidermal growth factor receptor (EGFR) and erbB-2 had higher levels of cox-2 and laminin-5 than those without concomitant overexpression of these proteins (P = 0.014 and P = 0.018, respectively). To see whether EGFR signaling is involved in cox-2 and laminin-5 expression, we further conducted in vitro analyses using six lung adenocarcinoma cell lines (A549, HLC-1, ABC-1, LC-2/ad, VMRC-LCD, and L27). Western blot analyses showed that cox-2 mRNA levels, and to a lesser extent laminin-5 gamma2 mRNA levels, correlated with the expression levels of erbB-2 and the phosphorylated form of MAPK/ERK-1/2 protein. The addition of transforming growth factor-alpha increased both cox-2 and laminin-5 gamma2 mRNA levels in A549, ABC-1, and L27 with different kinetics; the induction of cox-2 occurred earlier than that of laminin-5 gamma2. Finally, the migration of ABC-1 cells was inhibited by MAP kinase kinase inhibitor PD98059 and a selective cox-2 inhibitor NS-398. In contrast, the migration of A549 cells was inhibited by PD98059, but much less effectively by NS-398. These results suggest that co-stimulatory mechanisms may exist that increase the expression of cox-2 and laminin-5 at the invasive front of lung adenocarcinomas and that EGFR signaling could be one of the mechanisms. Further investigations are warranted concerning the role of cox-2 and laminin-5 in cancer cell invasion and the significance of p53 and EGFR signaling in the regulation of cox-2 and laminin-5 expression.     

Expression of vascular endothelial growth factor receptor 3 in blood and lymphatic vessels of lung adenocarcinoma

Vascular endothelial growth factor receptor 3 (VEGFR-3) has been proposed as a marker for lymphatic endothelial cells. This study investigated the expression of VEGFR-3 in the tumour vessels of lung adenocarcinoma and evaluated whether VEGFR-3 staining was useful for identifying lymphatic vessels within the tumour stroma. It also explored whether active growth of lymphatic vessels occurred in lung adenocarcinoma. Formalin-fixed, paraffin-embedded specimens obtained from 60 cases of lung adenocarcinoma, including five cases of pure bronchiolo-alveolar carcinoma (BAC) without stromal, vascular, and pleural invasion, were examined. No VEGFR-3-positive vessels were observed in pure BAC, but varying numbers of VEGFR-3-positive vessels were found in 39 of 55 (70.9%) invasive adenocarcinomas. A comparison of serial sections stained for VEGFR-3, CD31, and laminin-1 showed that most of the VEGFR-3-positive vessels appeared to be blood vessels (CD31-positive, laminin-1-positive), but some had the characteristics of lymphatic vessels (variable staining for CD31, little or no staining for laminin-1). VEGFR-3 staining highlighted lymphatic invasion by cancer cells; this invasion could not be detected by CD31 or haematoxylin and eosin (H&E) staining. Active growth of lymphatic vessels (as indicated by nuclear Ki-67 labelling of the endothelium) was observed in five tumours, four of which showed a high level of lymphatic invasion by cancer cells. It was concluded that VEGFR-3 immunostaining did not discriminate clearly between vascular and lymphatic endothelial cells, since expression of VEGFR-3 can be up-regulated in tumour blood vessels. However, VEGFR-3 staining combined with laminin-1 and CD31 staining would be useful for identifying lymphatic vessels and their invasion by tumour cells in a more objective way. Finally, proliferation of lymphatic endothelial cells may occur in association with lymphatic invasion by cancer cells.     

The GAP-related domain of tuberin, the product of the TSC2 gene, is a target for missense mutations in tuberous sclerosis

Tuberous sclerosis is an autosomal dominant trait in which the dysregulation of cellular proliferation and differentiation results in the development of hamartomatous growths in many organs. The TSC2 gene is one of two genes determining tuberous sclerosis. Inactivating germline mutations of TSC2 in patients with tuberous sclerosis and somatic loss of heterozygosity at the TSC2 locus in the associated hamartomas indicate that TSC2 functions as a tumour suppressor gene and that loss of function is critical to expression of the tuberous sclerosis phenotype. The TSC2 product, tuberin, has a region of homology with the GTPase activating protein rap1GAP and stimulates the GTPase activity of rap1a and rab5a in vitro. Here we show that the region of homology between tuberin and human rap1GAP and the murine GAP mSpa1 is more extensive than previously reported and spans approximately 160 amino acid residues encoded within exons 34-38 of the TSC2 gene. Single strand conformation polymorphism analysis of these exons in 173 unrelated patients with tuberous sclerosis and direct sequencing of variant conformers together with study of additional family members enabled characterisation of disease associated mutations in 14 cases. Missense mutations, which occurred in exons 36, 37 and 38 were identified in eight cases, four of whom shared the same recurrent change P1675L. Each of the five different missense mutations identified was shown to occur de novo in at least one sporadic case of tuberous sclerosis. The high proportion of missense mutations detected in the region of the TSC2 gene encoding the GAP-related domain supports its key role in the regulation of cellular growth.      

Formaldehyde in cryoprotectant propanediol and effect on mouse zygotes

Cryopreservation of human zygotes and embryos has been routinely performed by in-vitro fertilization clinics for many years. Karran and Legge (1996) first reported that formaldehyde (FA) present in the cryoprotective solutions can have a deleterious effect on mouse oocytes. FA is a cytotoxic, carcinogenic and mutagenic chemical. The effect of FA on mouse zygotes was investigated. In addition, the concentrations of FA in propanediol (PROH) obtained from various sources were determined. Pooled 1-cell embryos were dispensed into droplets of modified Ham's F10 or human tubal fluid containing various concentrations of FA. Since bovine serum albumin (BSA) may minimize toxicity additional trials were done as above in the absence of BSA. FA concentration in the standard 1.5 M PROH, from different sources in water, was measured in the same assay using a standard curve of 0-100 microM FA. FA in a complex medium had a significant deleterious effect on embryo development and hatching but only at 1 mM concentration (P < 0.000001; see Tables I-III). There was no significant effect of FA at 100 microM. However, in a simple medium even 50 microM FA decreased embryo hatching. FA was present in 1.5 M PROH from different sources (range 1.0-35.3 microM concentration). It appears that FA concentrations do not increase with storage because FA concentrations were low even after opening and storage for 3 years on the shelf. This suggests that FA is a contaminant during the manufacturing process and may vary from manufacturer to manufacturer and batch to batch. Until further studies are done to confirm the lack of toxicity to embryos during cryopreservation (with or without FA scavengers) it may be prudent to screen all batches of cryoprotectants for FA as part of quality control.