Analysis of local recurrence and optimizing excision margins for cutaneous melanoma.

BACKGROUND: Current guidelines for the treatment of melanoma favour conservatism; however there is still uncertainty regarding best practice for lesions of intermediate thickness. Local recurrence, a measure of treatment adequacy, can be used to determine optimum excision margins and give prognostic information for survival. METHODS: An analysis of the Auckland Melanoma Unit database was performed. Patients with local recurrence were identified and stratified by lesion thickness. Optimum excision margins were derived by regression analysis and evaluated against the database population. Survival and prognostic factors were studied. RESULTS: Eighty-four of 1155 patients (7 per cent) developed local recurrence. Median follow-up was 51 months. Margins predicted to give a local recurrence of zero were: 1 cm for lesions < or = 1 mm thick; 1.5 cm for lesions 1-2 mm thick; and 2 cm for lesions > 2 mm thick. Applied to 1155 patients, there were significant differences in both local recurrence and mortality rates between optimally and suboptimally excised lesions, except for those > 4 mm thick. Thirty-three patients (39 per cent) with local recurrence died. Thickness, local recurrence and ulceration were of prognostic significance. CONCLUSION: Development of local recurrence in melanomas < or = 4 mm thick is due to inadequate treatment. It signifies progressive disease and a poor prognosis. Care must be taken to ensure that all such lesions are optimally excised.

Incidence of malignant melanoma in Auckland, New Zealand: highest rates in the world.

The calculation of incidence rates of melanoma in New Zealand has been hampered in the past by incomplete registration of cases. The aim of this study was to document the incidence of melanoma in the Auckland Caucasian population and to define the pathologic characteristics of these lesions. Data were collected for the Auckland region from the New Zealand Cancer Registry and the Auckland Melanoma Unit database for 1995 and combined with census statistics to give the crude and age-standardized rates for invasive melanoma. The results were analyzed by gender, morphology, body site, and thickness. The crude annual incidence for invasive cutaneous malignant melanoma was 77.7/100,000. The age-standardized annual rate was 56.2/100,000 with no statistically significant differences in the rates for males and females. The cumulative risk of developing melanoma over a lifetime, from age 0 to 74, was 5.7% overall. The age-specific rates steadily increase with advancing age. The lesions were generally thin; 64% were less than 0.76 mm, and only 7% were thicker than 3.00 mm. In conclusion, the Caucasian population in the Auckland region has the highest incidence of melanoma in the world.

Laparoscopic pelvic lymphadenectomy for malignant melanoma.

Therapeutic laparoscopic techniques are well accepted in many areas of general surgery. The use of laparoscopy for the staging of malignancy, in a diagnostic role, is not used widely. Urologists use pelvic lymph node dissection (LPLND) to define those patients who are suitable for curative local therapy in carcinoma of the prostate. We present a case of a patient with metastatic malignant melanoma in whom LPLND was employed as a minimally invasive way of staging and treating her disease.

The sensitization of mucosal mast cells during infections with Trichostrongylus colubriformis or Haemonchus contortus in sheep.

Responses of isolated mucosal mast cells (MMC) during infections with either Trichostrongylus colubriformis or Haemonchus contortus were examined by measuring the release of sheep mast cell protease (SMCP) in a degranulation assay. MMC from sheep immune to T. colubriformis released maximal amounts of SMCP and histamine within 0.5h of incubation with larval antigen whereas maximum secretion of leukotrienes occurred 3h after addition of antigen. It was only after 8 weeks of a primary T. colubriformis infection, that MMC released significantly elevated levels of SMCP (23%); this occurred when the worm burden was being rejected. In contrast, the SMCP release from MMC of immune sheep was significantly higher at 40%, and occurred within 1-4 days after challenge (DAC). The SMCP release peaked at 6-8 DAC at 51%, and declined after 56 DAC to < 25%. MMC isolated from the duodenum and mid-small intestine of immune sheep released 2-3 times higher proportion of SMCP than did cells recovered from the terminal ileum. Mast cell numbers were similar in the 3 regions but the quantity of globule leucocytes (GL) was 2.5 times higher in the duodenum. During infections with H. contortus in the abomasum, MMC isolated from the small intestine released greater levels of SMCP when incubated with larval antigens than did abomasal MMC. There was no increased release during the first 12 weeks of a primary infection although the SMCP release (23%) from immune MMC at 7-10 DAC was significantly enhanced. Once again the release from MMC isolated from the three intestinal regions of sheep immune to H. contortus was lowest in the terminal ileum.(ABSTRACT TRUNCATED AT 250 WORDS)

Changes in inflammatory mediators and larval inhibitory activity in intestinal contents and mucus during primary and challenge infections of sheep with Trichostrongylus colubriformis.

Helminthologically naive merino sheep were given either a single infection of 30,000 or a trickle infection of 6000 Trichostrongylus colubriformis infective larvae (TcL3) per week. Faecal egg counts started to fall after 8 weeks in the single infection and after 11 weeks in the trickle infection. Small intestinal contents were collected from indwelling intestinal fistulae over the next 14 weeks. Concentrations of sheep mast cell protease (SMCP) in these contents increased to highest levels 9-11 weeks and 6-10 weeks after the single infection and from the start of the trickle infection, respectively. Similarly, peptidyl leukotriene (PLT) concentrations were highest at 6 weeks and at 6-9 and 13 weeks, respectively. Histamine concentrations increased slightly after both infections to peak values at 7 weeks and 9 weeks, respectively. Inhibition of migration of larvae in vitro was increased in contents sampled at 8 weeks after the single infection and after 6-10 weeks of the trickle infection. Another 2 groups of sheep were immunised by repeated infections with TcL3. Gut contents from 1 group sampled immediately before and after challenge with 30,000 TcL3 at 0 and 18 days had increased levels of larval migration inhibitory (LMI) activity throughout the 35 day period, especially 7-14 days after challenge (DAC). The mediators SMCP increased significantly 5-7 DAC while PLT increased 7-14 DAC. In the second group of immunised sheep, levels of SMCP and PLT increased rapidly within 1 DAC and further increased 3-14 DAC.(ABSTRACT TRUNCATED AT 250 WORDS)

Laparoscopic extraperitoneal inguinal hernia repair using a balloon dissection technique.

A laparoscopic extraperitoneal approach using a balloon dissection technique was used to repair 40 inguinal hernias in 35 patients. The initial experience with this method is presented. Thirty-four of these hernias were indirect, five direct and one sliding. Three were recurrent hernias. The operation time ranged from 40 to 135 min. Thirty-two of the patients stayed overnight in the hospital; Three stayed 2 nights. Return to normal activities ranged from 3 to 21 days. None of the patients had problems with nerve entrapment and to date there have been no recurrences of the hernias following the repairs. The procedure has been very well-tolerated by all patients.

A new simplified assay for larval migration inhibition.

A simple method is described for the in vitro detection of substances that impair the motility of third-stage larvae of gastro-intestinal nematodes. The test is based on the ability of larvae to freely migrate through selected mesh sizes of nylon sieves and the reduced ability of larvae to migrate after preincubation with, and in the presence of, substances that inhibit or reduce larval motility.

Isolation and degranulation of mucosal mast cells from the small intestine of parasitized sheep.

The isolation of mucosal mast cells and globule leucocytes from the small intestine of sheep immunized with Trichostrongylus colubriformis is described. Sheep mast cell protease was released from these cells in a dose-dependent fashion after incubation with soluble protein from T. colubriformis larvae. Release also occurred with other T. colubriformis antigens whereas non-parasite antigens at comparable protein concentrations evinced only a minimal response. Mucosal mast cells prepared from worm-free sheep also produced a similar minimal response. This is the first report describing the release of sheep mast cell protease from isolated sheep intestinal mucosal mast cells after addition of specific parasite antigens.

Studies of stage-specific immunity against Trichostrongylus colubriformis in sheep: immunization by normal and truncated infections.

Sheep immunized with multiple normal infections of 30,000 Trichostrongylus colubriformis larvae (T.c. L3) suppressed the fecundity, establishment and survival of adoptively transferred adult worms, showing that these parasites were susceptible to the effects of host immunity. When sheep were immunized by four 'truncated' larval infections of 4, 7 or 10 days' (d) duration with 10(5) T.c. L3, animals given 4 x 4d infections were susceptible to challenge, whereas sheep given 4 x 7d and 4 x 10d infections were significantly protected. A serial analysis of the rejection of T. colubriformis from nine sheep given 5 x 7d infections revealed that the challenge larval infection given intraduodenally was expelled within 3 days after challenge (DAC). However, another five of these sheep only rejected around 50% of transferred adult worms by 21 DAC when compared with control animals. The results indicate that stage-specific antigens produced by early L3 and L4 stages of T. colubriformis effectively immunize sheep against a larval challenge but appear less reliably protective against adult worms.

Studies of stage-specific immunity against Trichostrongylus colubriformis in sheep: immunization with adult parasites.

Merino sheep immunized by the adoptive transfer of adult T. colubriformis for 8 weeks were significantly protected against a challenge infection of 20,000 larvae. Two additional groups of sheep received a primary infection of 9000 adult worms which were allowed to persist for 14 weeks before one group was drenched. When both groups were challenged 10 days later with 30,000 larvae, serial necropsies of these and naive sheep revealed that worm rejection did not occur until 7-10 days after challenge. By comparison with the rapid rejection of larval challenges from sheep immunized with normal primary infections, the results suggest that the antigens which elicited rejection in these experiments are stage-specific and were only present or synthesized in sufficient quantities when parasites had developed for 1 week.

A serial study of rejection of Trichostrongylus colubriformis by immune sheep.

Host responses and the rejection of worms were measured at intervals following challenge of immune and susceptible sheep with T. colubriformis infective larvae. Immune sheep rejected most of their larvae within the first day after infection. This early rejection was associated with local appearance of globule leucocytes and increased concentration of T. colubriformis-specific IgG1 and IgG2 in intestinal mucus. Rejection of the remaining worms occurred between 3 and 14 days after infection and was associated with increased T. colubriformis-specific IgA and IgG2 in intestinal mucus, local T cell infiltration, activation, differentiation and epithelial necrosis. Local T cell changes included expansion of the T19- gamma delta+ populations in the villous lamina propria and epithelium.

Demonstration of a range of inflammatory mediators released in trichostrongylosis of sheep.

The levels of inflammatory mediators in the intestinal contents of sheep immunized with Trichostrongylus colubriformis larvae increased in the first 6 days after challenge. These mediators were histamine, leukotriene C4, 6-keto-prostaglandin F1 alpha (from prostacyclin) and thromboxane B2. Leukotriene C4 was released in the greatest quantities. Leukotriene B4 was present but its concentration remained unchanged after challenge. The presence of these particular mediators in the intestinal contents after challenge is consistent with antigen-induced mediator release from the mucosal mast cells found in immune sheep undergoing challenge infection. This is the first sequential analysis of mediator release in sheep that also demonstrates the release of prostacyclin and thromboxane into the intestine during expulsion of a nematode infection.

The response of immune sheep to challenge with Trichostrongylus colubriformis: enteric plasma loss and secretion of biogenic amines.

Sheep were challenged with a single large dose of larvae after vaccination with irradiated Trichostrongylus colubriformis larvae. Worm counts performed on vaccinated sheep 6 or 7 days after challenge (DAC) showed that they were solidly immune and only retarded L3 larvae were recovered at this time. Enteric plasma loss (EPL) in vaccinated animals increased immediately after challenge to peak 4-6 DAC and then decreased to pre-challenge levels. In contrast, a substantial rise in EPL did not occur in unvaccinated sheep until 10 DAC. Secretion of histamine into the duodenum of immunized sheep increased significantly from 2 to 7 DAC with the highest value at 6 DAC which corresponded with a lower duodenal tissue histamine level at this time. Histamine and 5HT secretion into the small intestine of previously uninfected sheep gradually increased during 12 weeks of a trickle infection of 3000 normal T. colubriformis larvae per week. The results indicate that rejection of incoming larvae by immune sheep is accompanied by an intestinal inflammatory response involving secretion of biogenic amines and a concurrent plasma loss.

Histamine and leukotriene concentrations in duodenal tissue and mucus of lambs selected for high and low responsiveness to vaccination and challenge with Trichostrongylus colubriformis.

Lambs selectively bred for high responsiveness or low responsiveness to vaccination with irradiated Trichostrongylus colubriformis were vaccinated and challenged. Duodenal tissue histamine concentrations in both high and low responder lambs were lower at 3 days than at 28 days after challenge. At 3 days after challenge, histamine concentrations were higher in both male and female high responder lambs than in low responder lambs whereas at 28 days concentrations were increased only in high responder females. At 3 days after challenge, histamine concentrations were generally lower in mucus than in tissues, but levels were again higher in mucus from high responder groups. In duodenal tissue at 3 days after challenge, leukotriene C4 and B4 concentrations were similar in high and low responder animals. At the same time, concentrations of both leukotrienes were higher in mucus than in tissues, with high responder female lambs having the highest concentration. It is suggested that increased levels of histamine and leukotrienes in mucus and tissue are associated with larval rejection or exclusion.

A laboratory study of the toxicity of the butanol extract of endod (Phytolacca dodecandra) on two species of freshwater fish and two species of aquatic snails.

The purpose of this study was to evaluate the toxicity of the butanol extract of Endod (Phytolacca dodecandra) on 4 species of aquatic animals. Groups of 10 mosquito fish (Gambusia affinis) and 8 bluegill (Lepomis macrochirus) were exposed to the butanol extract of Endod in 300 ml of water at concentrations of 0.0, 0.3, 0.4, 0.5, 0.6, 0.7, 0.8, 0.9, 1.0, 1.2 or 2.0 ppm. Groups of 10 tropical snails (Biomphalaria glabrata) and 10 pond snails (Physa spp) were also exposed to the crude extract in 50 ml of water at concentrations of 0.0, 1.0, 1.5, 2.0, 2.2, 2.5, 3.0, 4.0 or 5.0 ppm. Following a 24-h exposure period, the test subjects were transferred to extract-free water and observed for an additional 24 h. The number of dead animals was determined after the total 48-h concentration was plotted on logarithmic (probit) graph paper and the concentration of Endod which killed 50% of the test subjects (LC50) was determined. The butanol extract of Endod was lethal to 50% of the fish and snails at relatively low concentrations (less than 3.0 ppm). The results also indicated that fish were approximately 2 to 4 times more sensitive to Endod than snails.

The molluscicidal activity of Phytolacca dodecandra. I. Location of the activating esterase.

A number of methods have been used to extract molluscicidal saponins from the dried berries of Phytolacca dodecandra. The potency of the extract has been determined to depend on the release of an enzyme found only in the seed and breaking the seed is critical to the extraction process. The enzyme is inactivated by heat of alcohol. The highest potency extract is made from a cold water extraction of finely ground dried berries.

The use of fluorescent probes as markers for sheep lymphocyte subpopulations.

Sheep blood lymphocytes were labelled with fluorescent probes and examined under the fluorescence microscope and by the fluorescence-activated cell sorter. A novel probe using fluorescamine, coupled to hexylamine, detected 22.9% of cells, apparently of the B-cell series, counted by fluorescence microscopy. Substitution of fluorescein isothiocyanate (FITC) for the fluorescamine did not label the same subpopulation of cells although the lymphocytes could then be examined in the cell sorter. A larger number of cells (38.8%) formed the brighter cluster but did not behave as B cell when separated on nylon-wool columns. Improvement in discrimination of the cell populations was obtained with FITC-hexadecylamine (C16). This probe detected 38% of cells in the smallest cluster, 44% of cells in the intermediate cluster and 19% of cells in the brightest cluster. The proportion of cells in each cluster appeared to parallel closely the "null", erythrocyte (E) rosette-forming T cells and the B cells detected by conventional markers for blood lymphocytes. Other fluorescent probes, formed from FITC and other amines and amino acids, labelled lymphocyte membranes. Probes with a terminal charge labelled the small cluster particularly well, whereas those that were terminally non-polar labelled the larger cells brigthly, but not to the same intensity as the charged probes in the small cells.