Expression of EZH2 and Ki-67 in colorectal cancer and associations with treatment response and prognosis.

BACKGROUND: Enhancer of zeste homologue 2 (EZH2) is a member of the Polycomb group of genes that is involved in epigenetic silencing and cell cycle regulation. METHODS: We studied EZH2 expression in 409 patients with colorectal cancer stages II and III. The patients were included in a randomised study, and treated with surgery alone or surgery followed by adjuvant chemotherapy. RESULTS: EZH2 expression was significantly related to increased tumour cell proliferation, as assessed by Ki-67 expression. In colon cancer, strong EZH2 expression (P=0.041) and high proliferation (>or=40%; P=0.001) were both associated with better relapse-free survival (RFS). In contrast, no such associations were found among rectal cancers. High Ki-67 staining was associated with improved RFS in colon cancer patients who received adjuvant chemotherapy (P=0.001), but not among those who were treated by surgery alone (P=0.087). In colon cancers stage III, a significant association between RFS and randomisation group was found in patients with high proliferation (P=0.046), but not in patients with low proliferation (P=0.26). Multivariate analyses of colon cancers showed that stage III (hazard ratio (HR) 4.00) and high histological grade (HR 1.80) were independent predictors of reduced RFS, whereas high proliferation indicated improved RFS (HR 0.55). CONCLUSION: Strong EZH2 expression and high proliferation are associated features and both indicate improved RFS in colon cancer, but not so in rectal cancer.

Studies on the metabolic error in Refsum's disease.

Studies utilizing mevalonic acid-2-(14)C and D(2)O as precursors failed to provide evidence for an appreciable rate of endogenous biosynthesis of phytanic acid in a patient with Refsum's disease. Orally administered tracer doses of phytol-U-(14)C were well absorbed both by seven normal control subjects (61 to 94%) and by two patients with Refsum's disease (74 and 80%). The fraction of the absorbed dose converted to (14)CO(2) in 12 hours was 3.5 and 5.8% in Refsum's disease patients and averaged 20.9% in seven control subjects. Labeled phytanic acid was demonstrated in the plasma of both control subjects and patients given phytol-U-(14)C, establishing phytol in the diet as a potential precursor of phytanic acid. This labeled phytanic acid had disappeared almost completely from the plasma of the seven control subjects by 24 to 48 hours, whereas it persisted at high concentrations in the plasma of the two patients for many days. We conclude that the phytanic acid accumulating in Refsum's disease is primarily of exogenous origin and that patients with Refsum's disease have a relative block in the degradation of phytanic acid and possibly other similar branched-chain compounds. This may relate to a deficiency in mechanisms for release of phytanic acid from stored ester forms or, more probably, to reactions essential to oxidative degradation of the carbon skeleton.

Heredopathia atactica polyneuritiformis phytanic-acid storage disease, Refsum's disease:" a biochemically well-defined disease with a specific dietary treatment.

Heredopathia atactica polyneuritiformis (phytanic-acid storage disease, Refsum's disease) is an inborn error of metabolism, in which the body accumulates exogenous phytanic acid. The severe manifestations of this disease, which include pigmentary retinal degeneration, chronic polyneuropathy, ataxia, impaired hearing, and cardiopathy, can be either kept from worsening or reversed by elimination of foods rich in phytanic acid from patients' diets. This is borne out by a 20-year experience with two patients, whose conditions improved markedly once they stopped eating butter, animal fat, and other foods rich in phytanic acid.

Phytanic acid storage disease: hearing maintained after 15 years of dietary treatment.

Heredopathia atactica polyneuritiformis is a biochemically defined disease with a specific dietary treatment. It is an autosomal inborn error of metabolism. The phytanic acid is of exogenous origin and stems mainly from preformed phytanic acid in foods. In two Norwegian patients, serum phytanic acid has been brought down to normal levels and one of them has been followed for 15 years. During this period of dietary treatment there was no worsening of hearing.

Two cell kinetic methods studied on the rat corneal epithelium.

A stathmokinetic method (using Colcemid) and the [3H]thymidine technique (pulse labelling with tritiated thymidine, [3H]TdR) have been evaluated in the rat corneal epithelium. The dose is not of critical importance for the Colcemid method, thus indicating an all or nothing effect within the dose range studied. A one point estimate is sufficient to calculate the mitotic rate (MR), and in the rat corneal epithelium a 4 h accumulation period is recommended. After administration of [3H]TdR there is an increasing response with increasing dose, followed by a levelling off at higher doses. It seems reasonable to use the lowest maximal effective dose. The labelling index (LI) can be reliably registered 1 h after administration of the drug. For each of the drugs we found corresponding results after topical application and intraperitoneal injection. Hence, topical application of small doses of both Colcemid and [3H]TdR makes interesting in vivo experiments on larger animals and even on human beings possible. Due to the extreme regularity of the corneal epithelium this part of the eye is an interesting organ for cell kinetic studies and provides an excellent tool for evaluating cell kinetic methods.

Cell kinetics of the rat corneal epithelium.

In the rat corneal epithelium the mitotic rate (MR) is almost equal throughout the epithelium in the morning (Haaskjold et al. 1988). The labelling index (LI) shows a marked reduction in the central cornea, which could suggest a lack of uptake of tritiated thymidine via the salvage pathway (Haaskjold et al. 1989). In the present study we have used [3H]deoxycytidine, and [3H]thymidine after prior treatment with a methotrexate regimen to elucidate this discrepancy. Deoxycytidine is incorporated into DNA independent of thymidine kinase, while methotrexate, which depletes the cells of reduced folates, makes the cells completely dependent on the salvage pathway. With both techniques the same pattern of labelling was observed, confirming that in the morning the ratio between the MR and the LI differs throughout the cornea. Based on previous observations, an analysis of the MR/LI ratio during 24 h was performed, showing that these parameters were strongly correlated. This suggests that there may be different circadian variations in the cell proliferation parameters throughout the corneal epithelium. The methotrexate regimen may be a useful tool to investigate the salvage pathway.

Clinical and biochemical heterogeneity in conditions with phytanic acid accumulation.

Phytanic acid accumulation has for more than 20 years been used as a diagnostic criterion of Refsum's disease. Recently, however, phytanic acid has also been found in peroxisomal disorders (Zellweger's syndrome, neonatal adrenoleukodystrophy, infantile Refsum's syndrome, rhizomelic chondrodysplasia punctata). The 17 patients with Refsum's disease in the present study had serum phytanic acid values differing from 73 to less than 0.5 mg/dl (normal). alpha-Oxidation of phytanic acid in skin fibroblast cultures showed a defective capacity in all, with only small differences in residual activity. Phytanic acid determinations in serum from 3 of the 7 patients with peroxisomal disorders showed slightly elevated levels in 2. The alpha-oxidation capacity in the fibroblasts was defective in all, with a residual activity similar to that of Refsum's disease. An assay of the alpha-oxidation capacity may be useful in the diagnosis of both Refsum's disease and the peroxisomal disorders. The distinction between Refsum's disease and the peroxisomal disorders can easily be done on a clinical basis.

Benign familial myoclonus-like movements, partly of early onset.

A family with benign, myoclonus-like involuntary movements of early onset has previously been described by us (Refsum and Sjaastad, 1972, 1973; Sjaastad, 1981). In the present communication electrographical investigations carried out in the proband and his sister will be reported. Additional information will also be reported regarding onset of the abnormal movements.

Inhalation of organic solvents does not promote synovial tissue inflammation in rats.

Organic solvent exposure is known to give rise to a variety of manifestations in the nervous system both in man and in animals. Cardiac and skeletal muscle can also be affected, and strange collagen-like diseases, arthritis and arthralgia have been reported. In this work PVG rats were exposed to inhalation of toluene, commercial hexane, an industrial steel primer, and a cold vulcanizer for 4 months. No signs of synovitis appeared, either macroscopically or histologically, and there were no systemic signs of inflammation as detected by increased acute phase reactants.

Time dependency in the regenerative response to injury of the rat corneal epithelium.

In the present study a central corneal epithelial defect (diameter 3.5 mm) was made in both eyes at 12:00 h in one group of rats and at 24:00 h in another group to see if the regenerative proliferation is influenced by circadian rhythms. The labeling index and the mitotic rate were registered at 4-h intervals in the perilimbal conjunctiva, the limbal area, and different parts of the cornea from the following morning until noon the day after that. The most pronounced regenerative proliferation was seen in the midperipheral and peripheral cornea. The regenerative response occurred in both groups 24-28 h after the injury, but was highly influenced by the normal circadian rhythms, especially with regard to the mitotic rate. The results support the theory that even regeneration is influenced by a circadian proliferative factor.

Pentoxifylline improves resting membrane potential in sepsis.

The resting membrane potential (RMP) of skeletal muscle cells was investigated in a hyperdynamic animal model of sepsis. Reduced membrane potential is known to parallel the clinical course of sepsis and hemorrhagic shock in animal models. Pentoxifylline (PTF), a dimethyl xanthine derivative of the theophylline family, may improve the metabolic effects of sepsis. A cecal ligation and puncture (CL&P) sepsis model in Wistar rats was used in which RMP was measured with differing administration regimes of PTF. The RMP decreased less rapidly in our study when PTF was administered after CL&P and this effect was most noticeable after 18 hours. Administration of PTF before CL&P had little effect on membrane potential fall-off. These results suggest a potential role for PTF in the management of sepsis.

Pentoxifylline improves intracellular electrolytes in sepsis.

Sepsis is associated with a generalised membrane dysfunction leading to an increase in intracellular sodium, chloride, and water. The decrease in extracellular water is thought to act as a nonosmotic stimulus to the secretion of antidiuretic hormone. The resultant hyponatraemia is associated with increased surgical morbidity and mortality. Treatment aimed at improving intracellular electrolytes may improve surgical morbidity and mortality. An animal model of peritonitis was used to evaluate the effect of pentoxifylline. Previously, this dimethyl xanthine derivative was shown to stabilise the cell membrane. Administration of pentoxifylline significantly lowered intracellular sodium and chloride, particularly when given after caecal ligation and puncture. This may have clinical implications in the treatment and prevention of hyponatraemia.

Modulation of TNF alpha and IL-6 in a peritonitis model using pentoxifylline.

Sepsis leads to release of reactants that play an important role in the development of multiple organ failure. The kinetics of two early mediators of the response to sepsis, tumour necrosis factor (TNF alpha) and interleukin 6 (IL-6), and their modulation with pentoxifylline (PTF), were investigated. An established and clinically relevant animal model was employed, and sepsis was induced by cecal ligation and puncture (CLP) in Wistar rats. Six hours after the operation, there was an increase in IL-6 in all animals, which declined toward normal by 18 hours. This early phase of IL-6 production was not influenced by PTF. TNF alpha and IL-6 were significantly higher in the CLP group than in the animals treated with PTF at 24 hours. The blood pressure of the CLP group at 24 hours was significantly lower than that of the shams, and this decrease was not influenced by PTF. This decline in blood pressure may have been the stimulus to TNF production and the second phase of IL-6 production, which appeared to be inhibited by PTF. Pentoxifylline appears to attenuate systemic cytokine production in this model and may have a role in the management of clinical sepsis.

Accentuated oxidative stress following reperfusion injury in diabetic rats.

BACKGROUND: This study investigates whether diabetes mellitus accentuates hind limb ischemia-reperfusion injury. METHODS: Male Wistar rats rendered diabetic (n=40) following injection of streptozotocin were compared to non-diabetic control rats (n=30). Each group was divided into sham, 4 hrs of hind limb ischemia, 4 hrs of ischemia followed by 10, 30 or 60 min of reperfusion. Plasma concentrations of an end-product of lipid peroxidation [malondialdehyde (MDA)] and antioxidants (vitamins A and E) were measured together with the resting membrane potential (RMP) of the gastrocnemius muscle. RESULTS: Following reperfusion, the diabetic group showed greater and more persistent elevation of MDA and greater reduction of antioxidants. This was associated with reduction in the RMP only in the diabetic group. There was significant correlation between MDA level and the RMP in both groups of animals. CONCLUSIONS: These results indicate that oxidative stress following reperfusion injury is greater in the presence of diabetes mellitus. This may lead to a decrease in the RMP and increase in the vascular permeability, which may be associated with more complications.

Skin-reducing mastectomy and one-stage implant reconstruction with a myodermal flap: a safe and effective technique in risk-reducing and therapeutic mastectomy.

INTRODUCTION: Immediate reconstruction following mastectomy for breast cancer has been shown to be oncologically safe and associated with improved psychosocial outcomes for patients. Bostwick described a technique for one-stage implant based reconstruction, combining skin-sparing mastectomy with concurrent reduction of the skin envelope. This report reviews the experience of a single centre using skin-reducing mastectomy and one-stage implant reconstruction in both early stage breast cancer and risk-reducing mastectomy, with specific reference to frequency of complications, implant loss and oncological outcomes. METHODS AND RESULTS: A retrospective review was undertaken to identify women who had undergone skin-reducing mastectomy and one-stage implant reconstruction using a de-epithelialised dermal flap, between October 2008 and October 2012. One hundred and four consecutive mastectomies, with reconstruction, were performed by two surgeons on 64 patients. No complications were seen in 43.8% of patients. At three months, four implants were lost (3.8% of breast reconstructions, 6.3% of patients), due to either peri-implant infection or mastectomy skin flap necrosis. One patient required unplanned return to theatre for evacuation of a haematoma. Minor mastectomy skin flap necrosis was seen in 10 breasts (9.6% of reconstructed breasts) and superficial wound infection in 8 breasts (7.7% of reconstructed breasts). All of these complications were managed conservatively and none required operative intervention. At a median follow up of 35 months (4-53 months) there had been one episode of ipsilateral axillary nodal recurrence. CONCLUSION: One-stage implant reconstruction using a myo-dermal flap technique following skin-reducing mastectomy is safe and should be considered in selected patients. Most complications are minor and will resolve with conservative management. Major complications such as implant failure or immediate reoperation, were relatively uncommon (6.3% patients, 3.8% of reconstructed breasts). Early follow-up suggests that oncological outcomes are satisfactory, but longer follow-up is required to substantiate this.