The biological markers and results of treatment in male breast cancer patients. The Cracow experience.

Male breast cancer is a rare form of carcinoma with an incidence rate of approximately 0.5-1% compared with cases of breast carcinoma as a whole. Male breast cancer reacts effectively to endocrine therapy because of a high frequency of hormone receptor expression.The aim of the present study was the assessment of correlations between stage, grade, expression of steroid receptors, basal/mesenchymal markers and proliferation index, as well as analysis of the impact of the above-mentioned parameters on overall (OS) and disease-free survival (DFS) in the group of 32 male breast cancer patients, treated at the Centre of Oncology in Cracow.We showed the significant positive correlation between MIB-1 LI and tumor stage, and hormone receptors (ER or PgR) immunonegativity, and expression of EGFR, vimentin (p<0.05) and P-cadherin (the last at statistical border). The presence of any of basal or masenchymal markers correlated with a more advanced tumor stage. Moreover tumors without vimentin expression were characterised by lower MIB-1 LI and were more frequently EGFR immunonegative.We found that hormone receptor negativity, vimentin immunopositivity and high MIB-1 LI are significant independent indicators of poor OS and DFS for male breast cancer patients (p<0.05).

The impact of potential leakage from the sub-seabed CO2 storage site on the phosphorus transformation in marine sediments - An experimental study.

Carbon Capture and Storage (CCS) in the sub-seabed geological formations is a method of mitigation of carbon dioxide (CO2) emissions to avoid anthropogenic climate change. While CCS can be one of the most promising technologies to reduce atmospheric CO2 in the short and medium term, it raises serious concerns about the potential leakage of gas from storage sites. In the present study, the impact of acidification induced by CO2 leakage from a sub-seabed storage site on geochemical pools, and thus the mobility, of phosphorus (P) in sediment was investigated during laboratory experiments. The experiments were conducted in a hyperbaric chamber at a hydrostatic pressure of 900 kPa, which simulates pressure conditions at a potential sub-seabed CO2 storage site in the southern Baltic Sea. We performed three separate experiments in which the partial pressure of CO2 was: 352 µatm (corresponding pH = 7.7); 1815 µatm (corresponding pH = 7.0), and 9150 µatm (corresponding pH = 6.3). Under pH 7.0 and 6.3, apatite P is transformed into organic and non-apatite inorganic forms, which are less stable than CaP bonds and can be more easily released into the water column. At pH 7.7, P released during mineralization of organic matter and microbial reduction of FeP phases is bound with Ca, and the concentration of this form increases. The obtained results indicate that acidification of bottom water can reduce the efficiency of P burial in marine sediments, which contributes to an increase in P concentration in the water column and promote eutrophication especially in shallow areas.

Heart transplants in interferon-gamma, interleukin 4, and interleukin 10 knockout mice. Recipient environment alters graft rejection.

To study the role of cytokines in long-term cardiac allografts we have used recipient mice with targeted gene deletions (-/-) in IFN-gamma, IL-4, or IL-10. In wild-type and IL-4 -/- recipients immunosuppressed with a 30-d course of anti-CD4 and anti-CD8, graft survival was > 87 d. This time was significantly reduced in IFN-gamma -/- (62 +/- 19 d, P < 0.05) and IL-10 -/- recipients (55 +/- 4 d, P < 0.0001). Histology showed mononuclear cell infiltration, patchy necrosis, fibrosis, and vascular thickening in all groups. Intragraft transcript levels measured by 32P-reverse transcriptase PCR showed different inflammatory patterns. IFN-gamma -/- recipients had higher IL-2 transcripts and selective alteration in macrophage activation that may have contributed to decreased graft survival. Decreased graft survival in IL-10 -/- recipients was associated with increases in iNOS and IFN-gamma-driven responses. Finally, in grafts from IL-4 -/- recipients, there were increases in CD3 transcripts concurrent with TNF-alpha levels. This increase suggests that IL-4 may regulate T cell infiltration through TNF-alpha-mediated inflammatory cell recruitment. Concurrent evaluation of these three isolated cytokine deletions has shown that the recipient environment caused distinct graft modifications.

Predictive value of p53, mdm-2, p21, and mib-1 for chemotherapy response in advanced breast cancer.

p53 is a transcription factor that participates in cell cycle checkpoint processes and apoptosis. The protein product of the murine double minute gene 2 (mdm-2) plays a central role in the regulation of p53. In response to DNA-damaging agents, the wild-type p53-activated fragment 1 (WAF1 also known as p21) is an important downstream effector in the p53-specific growth arrest pathway. In breast cancer patients, it is unclear whether measuring p53, mdm-2, or p21 expression provides information on how patients will respond to chemotherapy. Mib-1 monoclonal antibody recognizes the proliferation-related antigen Ki-67. High tumor proliferation has previously been associated with response to chemotherapy. p53, mdm-2, p21, and mib-1 expression were assessed by immunohistochemical methods in primary tumors derived from 134 patients who took part in a randomized multicenter trial comparing docetaxel to sequential methotrexate and 5-fluorouracil (MF) in advanced breast cancer. Low mib-1 staining correlated with negative p53 staining (P = 0.001), and mdm-2 and p21 stainings correlated positively with each other (P < 0.001). p53, mdm-2, p21, and mib-1 expression were not significantly associated with response to chemotherapy, time to progression, or overall survival in the whole patient population or in the docetaxel group. However, in the MF group, a low mib expression (<25%) and a high mdm-2 expression (> or =10%) predicted a better response (P = 0.014 and P = 0.046, respectively) to treatment and a longer time to progression in both univariate and multivariate analyses. p53 staining status was not associated with response to treatment in either group. Interestingly, tumors with both negative mdm-2 and p21 expression, irrespective of p53 status, had a high response rate to docetaxel but no response to MF. Although highly preliminary, the findings suggest that different tumor biological factors may predict response to different chemotherapy regimens with distinct mechanisms of action. The results of our phenotype analysis also indicate that it is more likely that a panel of tumor biological factors instead of only one single factor may be needed for better prediction of chemotherapy response.

Molecular and functional analysis identifies ALK-1 as the predominant cause of pulmonary hypertension related to hereditary haemorrhagic telangiectasia.

BACKGROUND: Mutations of the transforming growth factor beta (TGFbeta) receptor components ENDOGLIN and ALK-1 cause the autosomal dominant vascular disorder hereditary haemorrhagic telangiectasia (HHT). Heterozygous mutations of the type II receptor BMPR2 underlie familial primary pulmonary hypertension. OBJECTIVE: To investigate kindreds presenting with both pulmonary hypertension and HHT. METHODS: Probands and families were identified by specialist pulmonary hypertension centres in five countries. DNA sequence analysis of ALK-1, ENDOGLIN, and BMPR2 was undertaken. Cellular localisation was investigated by heterologous overexpression of mutant constructs in both BAEC and HeLa cells. The impact of a novel sequence variant was assessed through comparative analysis and computer modelling. RESULTS: Molecular analysis of 11 probands identified eight missense mutations of ALK-1, one of which was observed in two families. Mutations were located within exons 5 to 10 of the ALK-1 gene. The majority of ALK-1 mutant constructs appeared to be retained within the cell cytoplasm, in the endoplasmic reticulum. A novel GS domain mutation, when overexpressed, reached the cell surface but is predicted to disrupt conformational changes owing to loss of a critical hydrogen bond. Two novel missense mutations were identified in ENDOGLIN. CONCLUSIONS: The association of pulmonary arterial hypertension and HHT identifies an important disease complication and appears most common among subjects with defects in ALK-1 receptor signalling. Future studies should focus on detailed molecular analysis of the common cellular pathways disrupted by mutations of ALK-1 and BMPR2 that cause inherited pulmonary vascular disease.

Partial inhibition of allograft arteriosclerosis (chronic rejection) by 15-deoxyspergualin.

15-deoxyspergualin (DSG) is an immunosuppressive drug that suppresses monocyte/macrophage function and/or T cell induction and early differentiation of B lymphocytes. It is as effective as cyclosporine in the prevention of acute rejection. We have investigated the effects of DSG on rat aortic allograft arteriosclerosis (chronic rejection). DSG was administered to the recipient rat at a dose of 0.3-10 mg/kg/day i.p. for 1-3 months, after which recipients were sacrificed. Histological changes were quantitated from paraffin sections. DSG is effective in chronic rejection in rat aortic allografts and reduces all three manifestations in the vascular wall--adventitial inflammation, media necrosis, and intimal thickening. At the dose of 1.0 mg/kg/day we demonstrated significant inhibition of adventitial inflammation from 10.6 point score units (psu) to 4.7 psu (P < 0.05), of media necrosis (P = 0.004) and of intimal thickening from 2.9 psu to 0.8 psu (P = 0.008). The therapeutic window was small in the long-term experiment. Doses of 3-10 mg/kg/day were toxic and 0.3 mg/kg/day was ineffective. In vitro smooth muscle cell proliferation was not inhibited by DSG and in the in vivo carotic denudation model DSG had no inhibitory effects either. These results suggest that DSG works via suppression of the immune/inflammatory response rather than via a direct antiproliferative effect on smooth muscle cells.

Chronic blockade of CD28-B7-mediated T-cell costimulation by CTLA4Ig reduces intimal thickening in MHC class I and II incompatible mouse heart allografts.

BACKGROUND: Chronic rejection develops in MHC class I/II-mismatched mouse allografts with arteriosclerosis and intragraft T-cell activation. Blockade with murine CTLA4Ig was used to study the role of CD28-B7 T-cell costimulation in this model of vascular thickening. METHODS: CBA/CaJ to C57BL/6J vascularized cardiac transplants were treated with murine CTLA4Ig delivered as a single dose (250 microg i.p.) on day 2 or chronically (100 microg i.p. on days 0, 2, and 4 and biweekly). Graft survival, function, and quantitative vessel analysis were compared with those of a reference group treated with anti-CD4 (days 1-4). RESULTS: Day 2 and chronic murine CTLA4Ig treatment prolonged graft survival (mean times and percentage of grafts surviving >75 days) and preserved graft function (measured by palpation scores). However, histology showed that chronic murine CTLA4Ig grafts had little parenchymal infiltration and less prominent vascular occlusion than day 2 murine CTLA4Ig-treated or 4-day anti-CD4-treated grafts. Quantitative analysis showed that the percentage of diseased vessels and the percentage of luminal occlusion were high in the day 2 murine CTLA4Ig group (78+/-20% and 41+/-12%, respectively, n=5) and the anti-CD4 group (94+/-9% and 52+/-17%, respectively, n=9, P=NS). In contrast, the frequency and severity of vessel thickening were significantly reduced in the chronic murine CTLA4Ig group (57+/-13% and 24+/-13%, respectively, n=10, P<0.03). CONCLUSION: In this model with MHC class I and II disparities, day 2 murine CTLA4Ig treatment improved survival and function but did not ameliorate vascular thickening. However, ongoing blockade of CD28-B7 costimulation conferred protection against vascular thickening.

Lack of effect of recombinant human superoxide dismutase on cold ischemia-induced arteriosclerosis in syngeneic rat aortic transplants.

Prolonged cold ischemia time and the generation of free oxygen radicals during reperfusion are risk factors for allograft arteriosclerosis. Growth factors are the main pro-proliferative mediators of smooth muscle cells in classical and in allograft arteriosclerosis. Superoxide dismutase is an enzyme that catalyzes the dismutation of superoxide anions into hydrogen peroxide. This study was designed to investigate which smooth muscle cell growth factor contribute to the formation of arteriosclerosis in syngenic vascular grafts with prolonged ischemia time, and whether perioperative intravenous administration of recombinant human superoxide dismutase (rh-SOD) prevents arteriosclerosis in these grafts. DA aortas were transplanted into DA recipients. One group of transplants was made with a short ex vivo ischemia time (15 min), while the other group transplant grafts was stored for 24 hr in cold saline. In addition to morphometric quantitation of the histological alterations, RNA isolated from grafts with short cold ischemia time in a semiquantitative polymerase chain reaction specific for various known smooth muscle cell growth factors. Syngeneic grafts with prolonged cold ischemia time showed severe intimal thickening and prominent medical necrosis, which were not seen in control groups. Approximately 3-fold levels of insulin-like growth factor-1 were found in ischemic syngeneic grafts compared with non-ischemic syngenic grafts, whereas epidermal growth factor levels were slightly lower. No changes in other growth factor mRNAs were found. Perioperative treatment with rh-SOD did not have significant effect on the extent of intimal thickening nor on the intensity of medial necrosis in grafts with prolonged ischemia time, and administration of rh-SOD did not change the expression level of insulin-like growth factor-1 in the grafts, either.

Redefining peripheral tolerance in the BALB/c to CBA mouse cardiac allograft model: vascular and cytokine analysis after transient CD4 T cell depletion.

BACKGROUND: To evaluate cardiac allografts from recipients that had achieved peripheral tolerance after transient CD4+ T cell depletion, we analyzed cellular infiltrate, cytokine expression, and vascular thickening. Long-surviving cardiac allografts from tolerant recipients were compared with acutely rejecting allografts and isografts. METHODS AND RESULTS: In CBA mice treated with anti-CD4 (GK1.5, 0.5 mg intraperitoneally on days 1-28), BALB/c cardiac allografts survived >100 days. These recipients were tested for tolerance at >70 days, by challenge with donor and third-party (C57BL/6) skin grafts. BALB/c skin grafts survived >30 days, although C57BL/6 skin was rejected in <12 days, reflecting alloantigen-specific peripheral tolerance. When vascular thickening in graft arteries was assessed and computerized measurements performed, heart allografts from tolerant recipients showed significantly increased percentage of luminal occlusion compared with isografts (47% compared with 1.2%). Semiquantitative reverse transcriptase-polymerase chain reaction was used to assess normalized intragraft mRNA transcripts for cytokines and T cell markers, with immunoperoxidase staining of frozen sections to confirmed the presence of protein. Compared with rejecting grafts, well-preserved hearts from tolerant mice had lower levels of macrophage and T cell infiltration and decreased transcription of interferon-gamma, interleukin (IL)-2, IL-10, and inducible nitric oxide synthase. IL-4 expression was similar in both groups. CONCLUSIONS: The degree of tolerance achieved allowed specific acceptance of donor skin grafts, preserved primary graft function, and reduced inflammatory activation. Tolerance did not, however, completely prevent macrophage and T cell infiltration of the graft or the development of vascular lesions typical of chronic rejection.

A vitamin D analog, MC1288, inhibits adventitial inflammation and suppresses intimal lesions in rat aortic allografts.

Certain analogs of vitamin D have been shown to prevent autoimmune diseases and prolong cardiac allograft survival. We transplanted aortic allografts from DA rats to WF rats to investigate the effect of a synthetic vitamin D analog, MC1288, and cyclosporine (CsA), alone or in combination, on acute and chronic rejection (allograft arteriosclerosis) and the mechanism of action of MC1288. The histological changes in the vascular wall were quantitated as point score units (psu). Adventitial inflammation linked with acute rejection at 1 month after transplantation decreased from 10.0+/-0.9 psu to 4.1+/-1.0 psu (P<0.01) when MC1288, 0.1 microg/kg/every other day, and CsA, 5 mg/kg/day, were combined. Intimal thickening decreased from 2.5+/-0.3 psu to 1.1+/-0.4 psu (P<0.05) at 3 months after transplantation. Proliferation of the adventitial lymphoid cells, detected by bromodeoxyuridine labeling, decreased from 140+/-36 to 20+/-19 labeled cells/cross-section. MC1288 alone suppressed interleukin 2 receptor-expressing cells from 156 to 90 positive cells/cross-section. Taken together, MC1288 with CsA effectively suppress T cell proliferation and activation and decrease intimal thickening.

Prognostic significance of telomerase activity in soft tissue sarcomas.

Only few reports on the prognostic significance of telomerase activity in human cancer exist. To find a new prognostic marker in soft tissue tumors, we investigated 60 soft tissue sarcomas of different histology and six benign tumors for telomerase activity. Telomerase activity was measured by using the non-radioactive PCR-based TRAP-assay. PCR products were analyzed on an automated fluorescence sequencer. Tumors of grade-II and grade-III histology showed a significantly poorer prognosis. Both disease-free (p<0.03) and the overall survival (p<0.02) were reduced in the highly malignant sarcoma patients. We found telomerase activity in 38.3% of the cases, there being a correlation with a more aggressive behavior of soft tissue sarcomas. Telomerase activity correlated with the grade of malignancy (p=0.04), but not with sex (p=0.64) or age (p=0. 48) of the patients. The total survival was significantly reduced in patients with telomerase-positive sarcomas (p=0.04). Both of the patients having grade I tumors with telomerase activity died of disease, whereas 10 of 11 patients with telomerase-negative grade I tumors are still alive. Only one of the benign tumors showed telomerase activity. We suggest that telomerase activity is a potential prognostic factor in malignant soft tissue tumors. Despite the histological heterogeneity of soft tissue tumors, single entities should be assessed for telomerase activity.

Risk factors for lung cancer among women in Poland.

A total of 118 women with histologically confirmed lung cancer and 141 healthy controls, were involved in a case-control study conducted in Cracow between 1991 and 1994. The aim of this study was to examine the role of smoking, alcohol consumption, and diet in female lung cancer risk. Multivariate analysis has shown that cigarette smoking was the most strongly active risk factor in female lung cancer. Vodka drinkers showed significantly higher risk than non-drinking women. Frequent intakes of carrots (at least three times a week) significantly lowered the risk. The significant protective effect was also observed in women using margarine on bread. The analysis of dose-response relationship in reference to cigarette smoking, vodka drinking and consumption of carrots also confirmed significant influence of these factors on risk.

MC1288, a vitamin D analog, prevents acute graft-versus-host disease in rat bone marrow transplantation.

The major obstacle to successful bone marrow transplantation (BMT) is graft-versus-host disease (GVHD). Vitamin D analogs have shown their efficacy in solid organ transplantation. The purpose of this study was to investigate the suitability of a novel vitamin D analog, MC1288, in the prevention of acute GVHD in a rat BMT model. Allogeneic BMT were performed from Lewis to BN rats (n = 18). The animals were divided into four groups: an untreated control group, MC1288, cyclosporin A (CsA), and MC1288 + CsA-treated groups. Rats were harvested for histology and immunohistochemistry on day 20 after BMT. Histological changes for GVHD in liver, skin, and spleen were scored. Positivity in immunostaining was quantified as the number of positive cells/high power field. Treatment with MC1288 decreased clinical signs of GVHD compared with untreated or CsA-treated rats. Histological manifestations of GVHD, expressed as mean total increment, were significantly lower (1.4 +/- 0.5) in MC1288 than in untreated (5.0 +/- 1.6) or CsA (3.5 +/- 1.0) groups. Combining MC1288 and CsA further improved histology (1.1 +/- 0.6). The expression of CD4, CD8, MHC class II, interleukin-2 receptor, nitric oxide 2, and NKR-P1A (NK cells) positivity was significantly decreased in the liver and skin of BMT rats by MC1288. MC1288 was effective in preventing clinical and histological signs and symptoms of GVHD. This novel vitamin D analog could be used as an immunomodulating agent in BMT.

Donor and recipient contributions of ICAM-1 and P-selectin in parenchymal rejection and graft arteriosclerosis: insights from double knockout mice.

BACKGROUND: Mice with target gene deletions were used in an immunosuppressed, heterotopic mouse cardiac transplant model to investigate the effects of simultaneous deficiencies of ICAM-1 and P-selectin on late cardiac rejection. METHODS: To determine the contribution of donor sources of ICAM-1 and P-selectin, ICAM-1/P-selectin gene deficient (I/P -/-) (n = 7) or wild type (n = 6) donor hearts were placed into CBA recipients. To study recipient sources of ICAM-1 and P-selectin, wild type donor hearts were placed into I/P -/- (n = 7) or wild type (n = 13) recipients. Recipients received a 30-day course of anti-CD4/8 mAb. RESULTS: I/P -/- donor allografts had prolonged survival (52-57 days) compared with wild type allografts (49-51 days). I/P -/- donor allografts underwent parenchymal rejection with mononuclear cell infiltration and developed alpha-smooth muscle actin positive vascular thickening (30 +/- 7% luminal occlusion, n = 78 vessels). Wild type allografts had parenchymal rejection with vascular medial necrosis and an absence of arteriosclerotic thickening (10 +/- 8%, n = 75, p = 0.008). Using the reverse combination, allografts from I/P -/- or wild type recipients had similar graft survival (50-57 days), comparable but variable degrees of parenchymal rejection, and comparable vascular occlusion (22 +/- 15% vs 28 +/- 19%, p = 0.442). CONCLUSION: We have shown that donor and recipient sources of ICAM-1 and P-selectin may have independent roles in leukocyte trafficking to the graft. Simultaneous interruption of donor ICAM-1 and P-selectin delays onset of parenchymal rejection. However, donor I/P deficiency permits arteriosclerotic development, perhaps by attenuating the alloimmune injury. In contrast, recipient deficiency alone does not altergraft outcomes suggesting that the donor is the critical site of ICAM-1 and P-selectin.

Chronic rejection in rat aortic allografts. IV. Effect of hypercholesterolemia in allograft arteriosclerosis.

Rat aortic allografts transplanted across histoincompatible strains develop arteriosclerotic alterations in the vascular wall that are virtually indistinguishable from those observed in human heart allografts during chronic rejection. In this study we have investigated whether hypercholesterolemia in the recipient rat accelerates allograft arteriosclerosis. Hypercholesterolemia was induced by a 4% cholesterol and 0.5% cholic acid diet, added to the normal rat diet. The cholesterol and cholic acid diet increased the level of serum total cholesterol from 1.3 +/- 0.0 to 4.8 +/- 0.9 (+/- SD) mmol/L and the level of low-density lipoprotein cholesterol from 0.3 +/- 0.0 to 2.6 +/- 1.0 mmol/L (p < 0.05) but caused no change in the level of high-density lipoprotein cholesterol, 1.0 +/- 0.1 versus 0.7 +/- 0.3 mmol/L. The level of plasma triglycerides remained also unchanged. Quantitation of two major chronic rejection-associated eicosanoids from the allograft vascular wall showed a significant increase in the synthesis of thromboxane B2 in the hyperlipidemic animals from 6.0 +/- 5.0 to 8.0 +/- 5.0 ng/mg dry weight and a slight reduction in the synthesis of 6-keto-prostaglandins F1 alpha. In vivo labeling of the recipient rat with tritiated thymidine and autoradiography showed that hypercholesterolemia did not affect the proliferation of inflammatory cells in the allograft adventitia, slightly increased the proliferation of smooth muscle cells in the media from 23 +/- 14 cells to 34 +/- 13 cells (+/- SEM) per cross section (p = ns), but slightly reduced the proliferation of smooth muscle cells in the intima from 13 +/- 6 to 6.2 +/- 1.5 (p = ns). Hypercholesterolemic recipients did not show any significant enhancement but, in fact, showed a delay in the generation of arteriosclerotic changes in the allograft intima. We conclude that although hypercholesterolemia, in the absence of hypertriglyceridemia, induces significant alterations in the eicosanoid metabolism and minor alterations in smooth muscle cell proliferation in the transplant vascular wall, it does not enhance arteriosclerotic alterations in chronically rejecting rat aortic allografts.

Induction and expression of cyclin D3 in human pancreatic cancer.

PURPOSE: Cyclins play a key role in the control and regulation of the cell cycle. The role of cyclins in the pathogenesis of pancreatic cancer is largely unknown. METHODS: Using Northern blot analysis, polymerase chain reaction (PCR) and immunohistochemistry, we examined the expression of cyclins D1, D2, and D3 in human pancreatic cancer and studied the induction of these cyclins by growth factors in pancreatic cancer cell lines. RESULTS: We now report that cyclin D1 and D3 mRNAs are expressed in human pancreatic cancer cell lines, and that the expression of cyclin D3 is enhanced in pancreatic cancer cells by amphiregulin, a member of the epidermal growth factor family. Cyclins D1 and D3 are also expressed in normal and malignant pancreatic tissues. However, while the normal pancreas and pancreatic cancers express cyclin D2 as determined by reverse-transcriptase PCR, we could not detect cyclin D2 mRNA by either Northern blot analysis or reverse transcriptase PCR in the two pancreatic cancer cell lines. Immunohistochemical analysis revealed the expression of cyclin D3 in pancreatic cancer cells. CONCLUSIONS: These findings suggest that D-type cyclins are differentially expressed in pancreatic cancer and that the aberrant activation of the EGF receptor in human pancreatic cancer by amphiregulin may lead to the progression of the cell cycle via induction of cyclin D3 expression, thus contributing to the growth advantage of these transformed cells.

Mycophenolate mofetil (MMF, RS-61443) inhibits inflammation and smooth muscle cell proliferation in rat aortic allografts.

To investigate the impact of mycophenolate mofetil (MMF) on allograft arteriosclerosis (chronic rejection) in rat aortic allograft model, we administrated MMF 20 mg/kg/day from the day of transplantation and sacrificed the rats at 1-12 months afterwards. MMF significantly suppressed all major histological manifestations of allograft arteriosclerosis, i.e. adventitial inflammation, media necrosis and intimal thickening and cellularity. There was a significant decrease in the replication rate (3H-thymidine incorporation) of inflammatory cells in the adventitia and of smooth muscle cells (SMC) in the media. MMF did not have any major effect on mRNA expression of several growth factors, (determined by polymerase chain reaction with inbuilt glyceraldehyde-3-phosphate dehydrogenase control), which have previously been demonstrated to be elevated in nonimmunosuppressed allografts. Immunoperoxidase staining showed a 40% reduction in the number of adventitial interleukin-2 receptors expressing lymphoid cells in MMF-treated allografts. The intensity of SMC alpha-actin staining was also significantly reduced. As the results suggested that MMF may have a direct antiproliferative effect on SMC, this possibility was investigated in primary SMC cultures in vitro and using the carotid denudation model in vivo. Both approaches showed inhibition of SMC proliferation by MMF. Our results indicate that MMF inhibits histopathological changes of chronic rejection by reducing the immune response and possible replication of SMC.

Autoradiography of [14C]paraquat or [14C]diquat in frogs and mice: accumulation in neuromelanin.

The herbicide paraquat has been suggested as a causative agent for Parkinson's disease because of its structural similarity to a metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), which may induce a parkinsonism-like condition. MPTP as well as its metabolite 1-methyl-4-phenylpyridine have melanin affinity, and the parkinsonism-inducing potency of MPTP is much stronger in species with melanin in the nerve cells. Autoradiography of [3H]MPTP in experimental animals has shown accumulation in melanin-containing tissues, including pigmented neurons. In the present whole body autoradiographic study accumulation and retention was seen in neuromelanin in frogs after i.p. injection of [14C]paraquat or [14C]diquat. By means of whole body autoradiography of [14C]diquat in mice (a species with no or very limited amounts of neuromelanin) a low, relatively uniformly distributed level of radioactivity was observed in brain tissue. Accumulation of toxic chemical compounds, such as paraquat, in neuromelanin may ultimately cause lesions in the pigmented nerve cells, leading to Parkinson's disease.

Long-term outcome in patients with unstable angina treated by coronary balloon angioplasty.

UNLABELLED: Comparison of balloon angioplasty results in 472 patients with stable angina (SA) and 158 patients with unstable angina (UA) in 5-year follow-up is reported. Clinical success rate did not differ significantly, while periprocedural complications rate was higher in UA group (22.3 vs. 11.1%, P<0.001). During follow-up UA patients demonstrated higher: restenosis rate (48.5 vs. 30.4%, P<0.001), incidence of myocardial infarction (8.8 vs. 3.0%, P=0.004), although cardiac mortality did not differ significantly (2.2 vs. 1.6%). Reintervention rate in patients with unstable angina resultant from restenosis or significant artherosclerosis progression in coronary vessels, or originating from both of them, was also higher (53.7 vs. 34.1%, P<0.001). Event-free survival was significantly lower in UA patients (43.4 vs. 61.3%, P=0.02). The uni- and multivariate analysis proved that unstable angina was an independent risk factor in restenosis, re-intervention and cardiac events rate, despite perceptible differences in the baseline characteristics. Sub-group analysis of UA patients according to Braunwald classification revealed lower success rate and higher incidence of myocardial infarction during follow-up in post-infarction angina (class C), whereas new onset, no-rest angina (class I) had higher event-free survival in comparison with rest angina (classes II and III). CONCLUSIONS: UA patients treated by balloon angioplasty had higher periprocedural complications rate, as well as restenosis and re-intervention rate. Despite higher cardiovascular events rate during 5-year follow-up in UA group, survival rate in both groups was high and cardiac mortality did not differ significantly. Unstable angina constitutes a strong independent risk factor in adverse long-term outcome.

Pigmentation of kidneys and lymph nodes of mesocolon in rats fed diets containing the laxative danthron.

The tumor-promoting activity of the anthraquinone laxative danthron was studied by giving 3 groups of male rats a single subcutaneous injection of the colon tumor-inducing agent 1,2-dimethylhydrazine (DMH). After 1 week, the animals were fed diets containing 0, 600 or 2400 ppm of danthron for 26 weeks. Two other groups of rats were included in the study; one received no treatment while the other was given danthron only. Altogether 9 tumors were observed among animals given DMA with or without danthron. The incidence of colon tumors was higher in animals receiving DMH and danthron than in those given DMH only (5/60 vs. 0/30), but this difference was not statistically significant. The kidneys and lymph nodes of mesocolon were enlarged and showed a yellowish-red and brown discoloration, respectively. The pigment mostly displayed a PAS-positive reaction but contained no lipid as determined by several staining procedures. The available evidence suggests that the pigment is drug-derived.