Anna Barańczyk-Kuźma

Arginase in patients with breast cancer

The mean arginase activity in breast cancers (n = 80) was significantly higher than in control tissues and it accounted for 0.31 +/- 0.23 U/g wet tissue and 0.083 +/- 0.061 U/g (P < 0.05), respectively. With the cutoff value of 0.1 U/g wet tissue, raised arginase activity was observed in 74% of tumors. The preoperative arginase activity in blood serum from women with breast cancer was 11.2 +/- 7.9 U/l (n = 115), and it was significantly higher than in 70 healthy controls, where it was 5.7 +/- 2.4 U/l (P < 0.05). With the cutoff value for normal serum arginase activity above 8.0 U/l, the activity was raised in 10% of control individuals, and in 63% of women with breast cancer. The sensitivity and specificity of the arginase test in blood serum were 63% and 60%, respectively. Two isoforms immunologically identical to human kidney arginases (L-arginine amidinohydrolase) were found in both normal and cancerous breast tissues. The level of anionic form was similar in control and cancerous tissues, whereas the cationic isoform predominated in breast cancer. The cationic isoform was the only one present in serum of both ill and healthy women, and its level was higher in patients with breast cancer. Thus, it can be concluded that the cationic isoform is responsible for the increase of arginase activity in serum of patients with breast cancer.

Arginase isoforms in human colorectal cancer.

Arginase (EC 3.5.3.1) activity was determined in 54 colorectal tissues obtained from patients with primary colorectal adenocarcinoma as well as in serum of 45 patients and 65 healthy individuals. In patients, the preoperative values of the mean serum arginase activity and the activity in colorectal tumors were much higher than in serum of healthy subjects and control tissues. Two isoforms of arginase, anionic and cationic, were identified in colorectal tissues (normal and cancerogenous), and only one, the cationic form, in serum. These arginases were different from the main human liver cationic arginase (pI 9.3). The anionic colorectal arginase was identical with the human liver anionic isoform (pI 7.7), and the cationic arginase from colorectal tissues and blood serum with the human kidney cationic enzyme (pI 8.9). The total activity and the level of protein of the cationic arginase in colorectal cancer was higher than in control tissue, and it was also higher in serum of patients with colorectal cancer than in healthy subjects. Thus, it can be concluded that the increased arginase activity in blood serum and colorectal cancer in studied patients was due to the raised level of the cationic arginase and this isoform seems to be a discriminating parameter for assessing the presence of colorectal cancer.

Plasma profiling reveals three proteins associated to amyotrophic lateral sclerosis

Amyotrophic lateral sclerosis (ALS) is the most common adult motor neuron disease leading to muscular paralysis and death within 3–5 years from onset. Currently, there are no reliable and sensitive markers able to substantially shorten the diagnosis delay. The objective of the study was to analyze a large number of proteins in plasma from patients with various clinical phenotypes of ALS in search for novel proteins or protein profiles that could serve as potential indicators of disease.

L-arginine as a factor increasing arginase significance in diagnosis of primary and metastatic colorectal cancer.

OBJECTIVE The usefulness of simultaneous L-arginine and arginase determination in diagnosis of primary and metastatic colorectal cancer. METHODS L-arginine and arginase were determined before and after surgery in serum from 43 patients with colorectal cancer (CRC), 24 with colorectal cancer liver metastasis (CRCLM), and 39 control subjects (10 patients with non-malignant diseases and 29 healthy blood donors). RESULTS Preoperative L-arginine concentration in the patient groups was 2-fold higher, whereas arginase activity was over 3- and 6-fold higher in CRC and CRCLM when compared with control. The values of both parameters lowered significantly after surgery. The sensitivity of single parameter in CRC was 79% for L-arginine and 81% for arginase, and in CRCLM it was 83% for each parameter. The combination of L-arginine with arginase improved the sensitivity to 93% and 100% in CRC and CRCLM, respectively. The specificity of L-arginine and arginase calculated for 39 subjects was 87% and 82%. CONCLUSION Simultaneous determination of L-arginine and arginase increases the value of arginase itself in diagnosis and follow up of patients with CRC and CRCLM.

Serum arginase activity in postsurgical monitoring of patients with colorectal carcinoma

Colorectal carcinoma (CRC) is one of the most common malignancies. In the current work, the role of arginase as a diagnostic marker in patients with recurrent CRC and colorectal liver metastases (CRCLM) was studied.

Changes in arginase isoenzymes pattern in human hepatocellular carcinoma.

Hepatocellular carcinoma (HCC) is one of the most common tumors worldwide affecting preferentially patients with liver cirrhosis. The studies were performed on tissues obtained during surgery from 50 patients with HCC, 40 with liver cirrhosis and 40 control livers. It was found that arginase activity in HCC was nearly 5- and 15-fold lower than in cirrhotic and normal livers, respectively. Isoenzymes AI (so-called liver-type arginase) and AII (extrahepatic arginase) were identified by Western blotting in all studied tissues, however the amount of AI, as well as the expression of AI-mRNA were lower in HCC, in comparison with normal liver, and those of AII were significantly higher. Since HCC is arginine-dependent, and arginine is essential for cells growth, the decrease of AI may preserve this amino acid within tumor cells. Concurrently, the rise of AII can increase the level of polyamines, compounds crucial for cells proliferation. Thus, both arginase isoenzymes seem to participate in liver cancerogenesis.

The influence of heroin abuse on glutathione-dependent enzymes in human brain.

Heroin is an illicit narcotic abused by millions of people worldwide. In our earlier studies we have shown that heroin intoxication changes the antioxidant status in human brain. In the present work we continued our studies by estimating the effect of heroin abuse on reduced glutathione (GSH) and enzymes related to this cofactor, such as glutathione S-transferase detoxifying electrophilics (GST) and organic peroxides (as Se-independent glutathione peroxidase-GSHPx), and Se-dependent glutathione peroxidase (Se-GSHPx) specific mainly for hydrogen peroxide. Studies were conducted on human brains obtained from autopsy of 9 heroin abusers and 8 controls. The level of GSH and the activity of glutathione-related enzymes were determined spectrophotometrically. The expression of GST pi on mRNA and protein level was studied by RT-PCR and Western blotting, respectively. The results indicated significant increase of GST and GSHPx activities, unchanged Se-GSHPx activity, and decreased level of GSH in frontal, temporal, parietal and occipital cortex, brain stem, hippocampus, and white matter of heroin abusers. GST pi expression was increased on both mRNA and protein levels, however the increase was lower in brain stem than in other regions. Heroin affects all regions of human brain, and especially brain stem. Its intoxication leads to an increase of organic rather then inorganic peroxides in various brain regions. Glutathione S-transferase plays an important role during heroin intoxication, however its protective effect is lower in brain stem than in brain cortex or hippocampus.

Antioxidant status in different regions of heroin addicts' brain.

Heroin is an illicit narcotic abused by millions of people worldwide. In the present work, we estimated peroxyl radical-trapping capacity (PRTC), oxidative stress markers - malondialdehyde and protein carbonyl groups, as well as antioxidant enzymes - superoxide dismutase and catalase, in different regions of brain. Studies conducted on nine brains from heroin abusers and eight from control subjects revealed a decrease in PRTC in each part of heroin intoxicated brains and an increase in lipid peroxidation in brain cortex, brain stem and white matter but not in hippocampus. Protein oxidation was increased in hippocampus and in brain stem, but it was unchanged in gray and white matters. Superoxide dismutase and catalase activities were unchanged in heroin addicts. We conclude that heroin intoxication changes the antioxidant status in human brain by increasing the amount of organic rather then inorganic peroxides. The most severe condition of oxidative stress occur in brain stem.

Kinesin Expression in the Central Nervous System of Humans and Transgenic hSOD1G93A Mice with Amyotrophic Lateral Sclerosis

Background: Amyotrophic lateral sclerosis is a fatal motor neuron degenerative disease. Most cases are sporadic (SALS), and approximately 10% are familial (FALS) among which over 20% are linked to the SOD1 mutation. Both SALS and FALS have been associated with retrograde axonal transport defects. Kinesins (KIFs) are motor proteins involved mainly in anterograde transport; however, some also participate in retrograde transport. Objective: The purpose of the study was to investigate and compare the expression of kinesins involved in anterograde (KIF5A, 5C) and retrograde (KIFC3/C2) axonal transport in SALS in humans and FALS in mice with the hSOD1G93A mutation. Methods: The studies were conducted on various parts of the CNS from autopsy specimens of SALS patients, and transgenic mice at presymptomatic and symptomatic stages using real-time quantitative PCR and reverse-transcription PCR. Results: All KIF expression in the motor cortex of individual SALS subjects was higher than in the adjacent sensory cortex, in contrast to the expression in control brains. It was also significantly higher in the frontal cortex of symptomatic but not presymptomatic mice compared to wild-type controls. However, the mean KIF expression in the SALS motor and sensory cortexes was lower than in control cortexes. To a lesser extent the decrease in KIF mean expression also occurred in human but not in mouse ALS spinal cords and in both human and mouse cerebella. Conclusion: Disturbances in kinesin expression in the CNS may dysregulate both anterograde and retrograde axonal transports leading to motor neuron degeneration.

Dynactin Deficiency in the CNS of Humans with Sporadic ALS and Mice with Genetically Determined Motor Neuron Degeneration

Dynactin is a complex motor protein involved in the retrograde axonal transport disturbances of which may lead to amyotrophic lateral sclerosis (ALS). Mice with hSOD1G93A mutation develop ALS-like symptoms and are used as a model for the disease studies. Similar symptoms demonstrate Cra1 mice, with Dync1h1 mutation. Dynactin heavy (DCTN1) and light (DCTN3) subunits were studied in the CNS of humans with sporadic ALS (SALS), mice with hSOD1G93A (SOD1/+), Dync1h1 (Cra1/+), and double (Cra1/SOD1) mutation at presymptomatic and symptomatic stages. In SALS subjects, in contrast to control cases, expression of DCTN1-mRNA but not DCTN3-mRNA in the motor cortex was higher than in the sensory cortex. However, the mean levels of DCTN1-mRNA and protein were lower in both SALS cortexes and in the spinal cord than in control structures. DCTN3 was unchanged in brain cortexes but decreased in the spinal cord on both mRNA and protein levels. In all SALS tissues immunohistochemical analyses revealed degeneration and loss of neuronal cells, and poor expression of dynactin subunits. In SOD1/+ mice both subunits expression was significantly lower in the frontal cortex, spinal cord and hippocampus than in wild-type controls, especially at presymptomatic stage. Fewer changes occurred in Cra1/SOD1 and Cra1/+ mice.It can be concluded that in sporadic and SOD1-related ALS the impairment of axonal retrograde transport may be due to dynactin subunits deficiency and subsequent disturbances of the whole dynein/dynactin complex structure and function. The Dync1h1 mutation itself has slight negative effect on dynactin expression and it alleviates the changes caused by SOD1G93A mutation.