Human tumor growth in nude mice is associated with decreased plasma cysteine and homocysteine.

The methionine cycle and its metabolites homocysteine and cysteine serve several important functions in cellular metabolism. Abnormalities in metabolism of the methionine cycle have been associated with cancer. We determined plasma levels of methionine, homocysteine and cysteine in nude mice implanted with human cancer cell lines (MDA-MB-435 breast, PC-3 prostate, HT29 colon, BX-PC3 pancreas) over a prolonged period of tumor growth. The data were compared with correspondins values in nontumor-bearing controls. Nude mice were injected s.c. in the right flank with 10(6) cancer cells. Tumor growth was measured over time. Methionine was measured in plasma by HPLC. Cysteine and homocysteine were measured in plasma by recombinant enzyme assays and spectrophotometry to measure products. The concentrations of cysteine and homocysteine in plasma decreased significantly as a result of progression of breast, prostate and the pancreas tumor types implanted in the nude mice at least over a two-month period. Data for the colon tumors were nonsignificant for both cysteine and homocysteine. In the case of methionine, the decrease was significant only due to progression of the breast tumors, grown over a long time period, as compared to the mice without tumors control. The results suggest that sulphur amino acids may be plasma or serum biomarkers for cancer progression.

Talin immunogold density increases in sciatic nerve of diabetic rats after nerve growth factor treatment.

BACKGROUND: Diabetic neuropathy is a debilitating disorder whose causation is poorly understood. Recent studies have shown significant reduction in the activity of nerve growth factor (NGF) and in the amount of talin cytoskeleton protein immunoreactivity in the perineurium in patients with diabetic neuropathy. OBJECTIVE: Since talin is involved in transmembrane connections between extracellular matrix and cytoskeleton, this study investigates the subcellular pattern of talin immunoreactivity and the effect of NGF treatment of diabetic rats on the distribution of talin in the sciatic nerve. MATERIALS AND METHODS: Post-embedding immunogold electron microscopy using monoclonal antibody against talin in combination with quantitative procedures was employed to localize talin-like immunoreactivity in the sciatic nerve of normal, diabetic and NGF treated diabetic rats. RESULTS: We found the highest densities of gold particles in the Schwann cells (139.6+/-5.6 particles/microm2) and in the fibroblasts (127.4+/-4.1 particles/microm2). A moderate amount of immunoreactivity was also present in the endothelial cells of vasa nervosa (32.3+/-9.1 particles/microm2). The myelinated and unmyelinated nerve fibers and the extracellular matrix profiles were not labeled (8.7+/-2.1 particles/microm2, 4.2+/-2.2 particles/microm2, 6.1+/-3.2 particles/microm2, 9.5+/-5.3 particles/microm2, respectively). The immunogold localization of talin in diabetic rats was significantly (p<0.001) reduced in Schwann cells (66.3+/-6.5 particles/microm2) and perineurial and epineurial fibroblasts (56.8+/-3.9 particles/microm2). Diabetic rats treated with NGF for 12 weeks showed significant (p<0.005) increase in talin-like immunogold density in Schwann cells and fibroblasts. Talin immunogold density in Schwann cells and fibroblasts increased approximately 68% and 58%, respectively, after NGF treatment. The endothelial cells of endoneurial and epineurial vessel walls showed no significant change in the talin-like immunogold particle density among control, diabetic and NGF treated diabetic animals. CONCLUSIONS: These results have shown that the administration of exogenous NGF may be essential for inducing functionally significant regenerative mechanisms in diabetic neuropathy through maintaining the permeability of the barrier properties of the peripheral nerve.

Allopurinol provides long-term protection for experimentally induced testicular torsion in a rabbit model.

OBJECTIVE: To assess the effect of five antioxidants on exocrine function of rabbit testes retained in situ for 24 h and 3 months after experimental torsion. MATERIALS AND METHODS: The left testes of peripubertal rabbits were clamped for 60 min, after which the clamps were removed and the testes allowed to reperfuse. The right testes served as internal controls. There were eight rabbits in each of the following experimental groups: (a) sham; (b) 60-min ischaemia followed by reperfusion; (c) 60-min ischaemia followed by left orchidectomy. In five further groups, rabbits were exposed to 60-min ischaemia followed by reperfusion, but received one of the following antioxidants before reperfusion: acetyl salicylic acid, ascorbic acid, allopurinol, quercetin or superoxide dismutase. Both testes were excised at 24 h or 3 months. The degree of lipid peroxidation, a measure of free radical damage, was assessed in testicular tissue homogenates by measuring the tissue levels of malondialdehyde (MDA). The Johnsen score was used to assess the morphological damage at 24 h and 3 months for each group. RESULTS: At 3 months twisted viable testes allowed to reperfuse had higher MDA levels than controls; the left testes of rabbits treated with allopurinol had significantly lower MDA levels than untreated rabbits and rabbits given other antioxidants. Rabbits given quercetin, ascorbic acid or superoxide dismutase had lower (but not significantly) left testicular MDA levels than untreated rabbits, while rabbits given acetyl salicylic acid had even higher levels. Allopurinol-treated rabbits had a Johnsen score of > 7.6 and those given other antioxidants had scores of < 7.6 at 3 months. CONCLUSION: The twisted viable testis treated by orchidopexy contains high free radical levels at 3 months. Of the antioxidants studied, only allopurinol had a beneficial long-term effect, by significantly reducing testicular MDA levels at 3 months.

Significance of determining the point of reperfusion failure in experimental torsion of testis.

BACKGROUND: Experimental studies of the use of free radical scavengers in ischemic/reperfusion (I/R) injury following detorsion of the torted testis have yielded conflicting results due to differences in the period of ischemia used. The authors studied I/R injury in the rabbit model, to define the point beyond which there is reperfusion failure. METHODS: Ischemia/reperfusion injury of the testis was created in 3-6-month-old male New Zealand white rabbits by cross-clamping the left spermatic cord for periods of ischemia lasting 0, 15, 30, 60, 90, 120 and 180 min. There were eight animals per experimental group. The right testis served as internal control. Both testes were harvested after 24 h of reperfusion in four animals and after 3 months in the remaining four animals for each group. Testicular malondialdehyde (MDA), a measure of free radical damage, was determined by using the thiobarbituric acid reaction on testicular homogenates. Johnsen score was used to assess morphological damage caused by the ischemia. RESULTS: After 24 h of reperfusion, the mean testicular MDA in the control right testes at 0, 15, 30, 60, 90, 120 and 180 min was 2.1, 2.5, 2.9, 2.4, 2.1 and 1.9 nmol/mg protein, respectively. The mean left testicular MDA at corresponding ischemic periods was 1.6, 2.0, 3.9, 10.0, 4.4, 6.1 and 1.0 nmol/mg protein, respectively. The maximum left testicular MDA was at 60 min (10.0 nmol/mg protein), following which the level dropped significantly to 1.0 nmol/mg protein at 180 min. At 3 months, the mean Johnsen scores for left testes subjected to 0, 60, 120 and 180 min ischemia were 9.4, 8.8, 2.3, 3.5, respectively. CONCLUSION: The results suggest that following ischemia of up to 60 min in the rabbit testis, adequate reperfusion is possible, but ischemia lasting beyond 60 min results in inadequate reperfusion leading to irreversible damage. Thus, in experiments for assessing the effect of antioxidants on I/R injury of the testis in rabbits, periods up to 60 min of ischemia should be regarded as optimum to observe an effect.

Role of Na+/H+ exchanger isoform-1 in human inflammatory bowel disease.

BACKGROUND: Na+/H+ exchanger (NHE) is responsible for a net uptake of sodium chloride and water from the gastrointestinal tract and maintains electrolyte and water homeostasis. However, its status in human inflammatory bowel disease such as ulcerative colitis (UC) and Crohn's disease (CD) remains poorly understood. OBJECTIVES: To investigate the role of NHE-1 isoform in human CD and UC. METHODS: Expression of NHE-1 protein and messenger ribonucleic acid and sodium pump activity were examined in the colonic biopsy samples taken from UC (n=11) and CD (n=13) patients using enhanced chemiluminescence-Western blot analysis, reverse transcription polymerase chain reaction and spectrophotometry. Subjects presenting with abdominal pain and endoscopically normal colon served as normal controls (n=11). Myeloperoxidase (MPO) activity and histology were performed to confirm tissue inflammation. RESULTS: MPO activity increased significantly (P<0.05) in both UC and CD patients compared with the normal controls. Parallel to MPO activity profile, there was also a significantly higher infiltration of inflammatory cells in both cases. P-nitrophenylphosphatase activity, a marker of the sodium pump, remained unchanged in CD but increased significantly (P<0.05) in UC compared with the normal controls. On the contrary, the level of NHE-1 protein and messenger ribonucleic acid was significantly decreased (P<0.05) in both cases, whereas the internal control, a-actin remained unaltered. CONCLUSIONS: These findings demonstrate a transcriptionally regulated suppression of NHE-1 in both UC and CD. This NHE-1 suppression may reduce an uptake of sodium chloride and water from the inflamed colonic lumen and thus contribute to diarrhea and neuromuscular alterations in these conditions.

Activities of erythrocyte antioxidant enzymes in cancer patients.

We measured the activities of erythrocyte superoxide dismutase (SOD) and glutathione peroxidase (GPX) before therapy in 97 patients with cancer in various sites (gastrointestinal tract (GIT) (n=40), breast (n=30), and others (n=27)), and in 60 matched controls to assess antioxidant enzyme protection. Hemolysate hemoglobin (Hb) was measured spectrophotometrically. The activity of SOD (U/g Hb) was significantly lower in all sites (when all the cancer sites were considered as a group), GIT, breast, and other sites compared to the controls (P<0.0001, P<0.0001, P<0.0001 and P<0.0001, respectively). The activity of GPX (U/g Hb) was significantly decreased in all sites, GIT, and breast cancer sites than in the controls (P=0.024, P=0.033, and P=0.043, respectively). Age showed a weak negative correlation with enzyme activities in controls and patients. There was no significant association between SOD and GPX activities in either the controls or the patients. These results suggest that there may be a greater antioxidant burden for SOD than GPX in cancer, and that a weak association exists between the activities of the two enzymes in antioxidant protection.