Pathology techniques

Pathology techniques developed over the past decade may be successfully applied to the diagnosis of orbital disease. Tissue specimens may be immunophenotyped by immunohistochemical staining and flow cytometry. Immunohistochemical staining provides a qualitative analysis of localization of antigenic determinants and flow cytometry provides a quantitative analysis. The molecular biologic technique of polymerase chain reaction enables detection of minute amounts of material by amplification from DNA primers. Fine-needle aspiration biopsy is an alternative to open biopsy in select cases. The most important concept regarding these techniques is communication with the ophthalmic pathologist regarding the collection and interpretation of the specimen.     

Technetium-99m dimercaptosuccinic acid and ifosfamide tubular dysfunction in children with cancer

Quantitative ^99mTc-dimercaptosuccinic acid (^99mTc-DMSA) renal scintigraphy was used to asses ifosfamide-induced changes in renal function in 11 children who received chemotherapy for various malignancies. Serial measurements of absolute ^99mTc-DMSA renal uptake, calculated on conjugated views, were performed during and after chemotherapy. Data of 37 studies obtained before and at different cumulative dose levels of ifosfamide were analysed in relation to clinical and biochemical parameters. A highly significant relationship between ^99mTc-DMSA uptake and cumulative ifosfamide dose was found (P<0.001). The most frequently observed abnormal pattern on scintigraphic images was decreased kidney uptake together with increased accumulation in bladder. ^99mTc-DMSA uptake was more consistent than ₂-microglobulin values in urine and more sensitive than quantitative hyperaminoaciduria and tubular resorption of phosphate for the detection of ifosfamide-induced tubular dysfunction. ^99mTc-DMSA uptake was decreased in both patients with and patients without clinical toxicity. Persistently reduced ^99mTc-DMSA uptake was observed in four patients during follow-up; in one of them, who was asymptomatic after ifosfamide therapy, sudden onset of Fanconi syndrome was observed when he was retreated with carboplatin 1 year later. It is concluded that ^99mTc-DMSA renal scintigraphy is a suitable method to assess progressive ifosfamide-induced tubular injury whereas scintigraphic imaging is helpful for interpreting renal uptake changes. The test is able to detect subclinical injury and may potentially predict high risk at retreatment.     

A meta-analysis of randomized controlled trials of laparoscopic versus conventional appendectomy

BACKGROUND: Despite many randomized controlled trials, the merits of laparoscopic appendectomy remain unclear. A meta-analysis may provide insights not evident from any individual studies. DATA SOURCES: Systematic literature search yielded 17 trials (1,962 subjects) of true randomized design with usable statistical data comparing laparoscopic and conventional appendectomy in adults. The effect sizes for operating time, hospitalization, postoperative pain, return to normal activity, wound infection, and intra-abdominal abscess were calculated, using the random effects model to allow for heterogeneity. An estimate of the robustness of all positive findings was also calculated. RESULTS: Modest but statistically significant effect sizes were found for four of the six outcome measures. Laparoscopic appendectomy takes 31% longer to perform, but results in less postoperative pain, faster recovery (by 35%), and lower wound infection rates (by 60%). CONCLUSION: Laparoscopic appendectomy offers significant improvement in postoperative outcomes at the cost of a longer operation.     

Predictability of PSA failure in prostate cancer by computerized cytometric assessment of tumoral cell proliferation

OBJECTIVES: To evaluate the relationship of DNA ploidy and cell proliferation (CP) with Gleason score (GS) and clinical outcome in prostate cancer. METHODS: Sixteen patients with benign prostatic hyperplasia (BPH) and 65 patients with prostate cancer classified by GS (four groups: 2 to 4, 5 to 6, 7, and 8 to 10) were studied. All patients with carcinoma underwent prostatectomy and were separated into prostate-specific antigen (PSA) failure and nonfailure groups (failure if PSA 0.1 ng/mL or more three times after surgery). Tumoral CP (Ki-67 inmunostaining and SG2M phase) and DNA ploidy were evaluated by computerized cytometry. RESULTS: BPH were diploid with low CP (8% SG2M cells or less). Carcinomas were either diploid with high CP (greater than 8% SG2M cells) or aneuploid. CP was significantly higher (P <0.001) in tumors with GS 7 or greater than in tumors with GS less than 7 (mean percent Ki-67 cells 18.3% versus 7.8%, respectively). PSA failure increased with GS (7.1% in GS 2 to 4, 21% in GS 5 to 6, 28.6% in GS 7, and 50% in GS 8 to 10), as well as with aneuploidy (18.5% in diploid tumors versus 72.7% in aneuploid tumors). Those experiencing PSA failure had significantly higher (P <0.001) CP than those not failing (mean percent Ki-67 cells 24% and mean percent SG2M 30.4% versus 8.7% and 13.5%, respectively). Cox regression analysis showed GS, DNA ploidy, Ki-67, and SG2M to each be univariately prognostic for time to PSA failure; however, Ki-67 and SG2M were more highly significant (P <0.0001 for both) than GS (P = 0.007) or DNA ploidy (P = 0.002). After adjusting for either SG2M or Ki-67 measures of CP, neither ploidy nor GS contained additional prognostic value. CONCLUSIONS: Tumor CP and DNA ploidy can be reliably determined in prostate cancer by computerized cytometry. On the basis of our preliminary results, CP correlates well with GS and predicts PSA failure better than DNA ploidy or GS.     

Effect of dietary inducer dimethylfumarate on glutathione in cultured human retinal pigment epithelial cells.

PURPOSE: To determine the effect of dimethylfumarate (DMF), an inducer of glutathione (GSH)-dependent detoxification, on intracellular GSH levels in cultured human retinal pigment epithelium (hRPE) cells, its mechanism of action, and its effect on hRPE cells subjected to oxidative injury. METHODS: Established hRPE cell lines were treated with DMF and assayed by high-pressure liquid chromatography for intracellular and extracellular GSH levels. Quantification of gamma-glutamylcysteine synthetase (GLCL) was determined through northern and western blot analyses, and activity was measured. Effects of pretreatment with DMF on GSH redox status of hRPE cells was determined. Sensitivity of hRPE cells to oxidative stress was determined using tert-butylhydroperoxide as the oxidative agent. RESULTS: Dimethylfumarate caused a transient decrease followed by a significant increase in intracellular GSH. Glutathione increased maximally at 24 hours with 100 to 200 microM DMF. The initial decrease could be accounted for by the formation of a DMF-GSH conjugate. Dimethylfumarate treatment increased the steady state mRNA expression of the regulatory subunit of GLCL, but no increase was seen for the catalytic subunit. However, protein levels were increased for both, and the catalytic activity of GLCL was also increased. Whereas the initial decrease in GSH made hRPE cells more susceptible to oxidative damage, pretreatment with DMF under conditions that increased intracellular GSH protected hRPE cells against oxidative damage. CONCLUSIONS: These results suggest a means by which the antioxidant capability of hRPE may be augmented without direct antioxidant supplementation. Specifically, a dietary compound that conjugates with GSH can induce GSH synthesis, increase GSH concentration, and improve protection by GSH-dependent detoxification pathways in hRPE. However, the early depletion of GSH before stimulated synthesis necessitates caution in prevention strategies using dietary inducers.     

Involvement of platelet-activating factor in the modulation of vascular tone in the isolated perfused rabbit kidney

The hypothesis that platelet-activating factor (PAF) plays a role in the modulation of the vasomotor tone and blood pressure was put forward by our group in previous in vivo studies in anaesthetised rabbits. The present study was undertaken to investigate the putative role of this lipid mediator in the vascular reactivity of the renal circulation, using the experimental model of the isolated perfused rabbit kidney. Dose-response curves to noradrenaline-induced vasoconstriction were performed before and after continuous infusions of two different PAF-receptor antagonists (WEB 2086 and yangambin) and of the phospholipase A₂ inhibitor mepacrine. The increases in renal perfusion pressure elicited by noradrenaline were potentiated by all the above-mentioned treatments in a dose-dependent manner. Moreover, prostaglandin F_2α-induced vasoconstriction was also potentiated by the administration of the PAF receptor antagonists and mepacrine. Furthermore, the administration of PAF into the renal circulation induced dose-related and long-lasting vasodilator responses, which were blocked by the PAF receptor antagonists. Nevertheless, PAF-induced renal vasodilation was also abolished by a pretreatment with mepacrine or with the cyclooxygenase inhibitor indomethacin, suggesting that it enhances the secondary formation of vasodilator arachidonic acid metabolites. The data indicate that PAF is involved in the modulation of the vasomotor tone in the renal circulation, through the release of cyclooxygenase products, constituting an additional mechanism of modulation of smooth muscle cell contractility to the ones exerted by well-known vasoactive substances of endothelial origin such as nitric oxide. Received: 20 April 1998 / Accepted: 5 March 1999     

Identification of phase I metabolites of 3-methylindole produced by pig liver microsomes.

A study was conducted to investigate qualitative and quantitative aspects of the phase I metabolism of 3-methylindole (3MI) by porcine liver microsomes. Microsomal suspensions were prepared from the liver of 30 intact (uncastrated) male pigs. Metabolites produced in microsomal incubations were identified and quantitated with HPLC-UV, HPLC-fluorescence, and UV-spectral analysis; liquid chromatography-mass spectrometry (LC-MS) and NMR were used for the identification of a metabolite for which a reference compound was not available. The results showed that seven major metabolites of 3MI are produced by porcine microsomes, three of which had already been identified in pigs (3-OH-3-methyloxindole, 5-OH-3-methylindole, and 6-OH-3-methylindole). The other four major 3MI metabolites identified were 3-OH-3-methylindolenine, 3-methyloxindole, indole-3-carbinol, and 2-aminoacetophenone. On average, the metabolite that was produced in larger amounts was 3-OH-3-methylindolenine (45.1%), followed by the two oxindoles 3-methyloxindole (27.9%) and 3-OH-3-methyloxindole (18.5%). Average percentage of production of 6-OH-3-methylindole was 4.9%, whereas indole-3-carbinol accounted for 2.7% of all metabolites produced; 2-amino-acetophenone and 5-OH-3-methylindole were the metabolites produced in lesser amounts (0.5 and 0.3%, respectively). Large interindividual differences in the rate of production of all metabolites were observed. This variation could be attributed to differences in the activity and/or level of expression of phase I biotransformation enzymes and this issue should be further investigated.     

Alosetron relieves pain and improves bowel function compared with mebeverine in female nonconstipated irritable bowel syndrome patients

BACKGROUND: Irritable bowel syndrome is one of the most common gastrointestinal disorders, yet no therapy convincingly controls the multiple symptoms of this syndrome. AIM: To compare the efficacy and tolerability of the new 5-HT3-receptor antagonist alosetron and the smooth muscle relaxant mebeverine in a double-blind, multicentre, randomized trial. METHODS: Six hundred and twenty-three nonconstipated females with irritable bowel syndrome were randomized to receive alosetron 1 mg twice daily (n=319) or mebeverine 135 mg three times daily (n=304) for 12 weeks, followed by a 4-week post-treatment period. The primary efficacy end-point was monthly responders for adequate relief of irritable bowel syndrome related abdominal pain and discomfort (defined as patients reporting adequate relief on at least 2 out of 4 weeks). Secondary end-points included assessments of bowel function, including urgency, stool frequency and stool consistency. RESULTS: There were significantly more responders in the alosetron group compared with mebeverine at months 2 and 3 (P < 0.01). Compared with mebeverine, the alosetron group experienced significant decreases in proportion of days with urgency and mean stool frequency, and had firmer stools within 1 week of starting treatment. A similar proportion of patients reported adverse events in the two treatment groups. CONCLUSIONS: In nonconstipated female irritable bowel syndrome patients, alosetron is significantly more effective than mebeverine in improving symptoms.