A genetic strategy involving a glycosyltransferase promoter and a lipid translocating enzyme to eliminate cancer cells

The most common therapeutic strategy for the treatment of cancer uses antimetabolites, which block uncontrolled division of cancer cells and kill them. However, such antimetabolites also kill normal cells, thus yielding detrimental side effects. This emphasizes the need for an alternative therapy, which would have little or no side effects. Our approach involves designing genetic means to alter surface lipid determinants that induce phagocytosis of cancer cells. The specific target of this strategy has been the enzyme activity termed aminophospholipid translocase (APLT) or flippase that causes translocation of phosphatidylserine (PS) from the outer to the inner leaflet of the plasma membrane in viable cells. Efforts to identify the enigmatic, plasma membrane APLT of mammalian cells have led investigators to some P-type ATPases, which have often proven to be the APLT of internal membranes rather than the plasma membrane. By measuring kinetic parameters for the plasma membrane APLT activity, we have shown that the P-type ATPase Atp8a1 is the plasma membrane APLT of the tumorigenic N18 cells, but not the non-tumorigenic HN2 (hippocampal neuron × N18) cells. Targeted knockdown of this enzyme causes PS externalization in the N18 cells, which would trigger phagocytic removal of these cells. But how would we specifically express the mutants or antisense Atp8a1 in the cancer cells? This has brought us to a glycosyltransferase, GnT-V, which is highly expressed in the transformed cells. By using the GnT-V promoter to drive a luciferase reporter gene we have demonstrated a dramatic increase in luciferase expression selectively in tumor cells. The described strategy could be tested for the removal of cancer cells without the use of antimetabolites that often kill normal cells.     

The relative roles of the parasol-like tail and burrow shuttling in thermoregulation of free-ranging Cape ground squirrels,Xerus inauris

As small arid-zone mammals, Cape ground squirrels (Xerus inauris) are unusual in being diurnally active. It is postulated that they remain active during the day by using their parasol-like tails to shade their bodies whilst foraging. However, no studies have continuously measured body temperature to determine the effect of using the tail as a parasol, relative to other thermoregulatory behaviours, such as burrow retreat. We caught four free-ranging Cape ground squirrels (673 ± 36 g) and surgically implanted miniature temperature-sensitive data loggers into their abdomens, to record body temperature every 5 min to an accuracy of 0.04 °C, before they were released back into their home range and observed for two weeks. Mean daily peak black globe temperature was 41 °C, and daily peak body temperature reached 40 °C. Ground squirrels raised their tails significantly more often at globe temperatures above 30 °C, but raising the tail did not decrease body temperature, nor prevent body temperature rising. Ground squirrels retreated to burrows, at 18 °C, significantly more often at high body temperatures and body temperature dropped 1–2 °C before re-emergence. We believe that the tail was raised to provide thermal comfort during high solar radiation exposure, and that burrow retreat was employed to dissipate a heat load and remain active diurnally.     

Inhibition of cysteine peptidase activity in ascitic fluid in pancreatic cancer patients

The work's objective is to answer the question whether there is any possibility of activity inhibition of cysteine peptidases inhibitors playing an important role in key processes accompanying cancer formation, including pancreas. There is a justified speculation that specific inhibitors of these enzymes may inhibit development of cancer processes by inhibiting their activity. In vitro studies confirmed that these enzymes in ascitic fluid were inhibited with egg whites inhibitors even to 90% of their original activity.     

A preliminary study in osteoinduction by a nano-crystalline hydroxyapatite in the mini pig

To test the probable osteoinductive properties of NanoBone, a new highly non-sintered porous nano-crystalline hydroxylapatite bone substitute embedded into a silica gel matrix, granules were implanted subcutaneously and intramuscularly into the back region of 18 mini pigs. After periods of 5 and 10 weeks as well as 4 and 8 months, implantation sites were investigated using histological and histomorphometric procedures. Signs of early osteogenesis could already be detected after 5 weeks. The later periods were characterized by increasing membranous osteogenesis in and around the granules leading to the formation of bone-like structures showing periosteal and tendon-like structures with bone marrow and focal chondrogenesis. Bone formation was better in the subcutaneous than in the intramuscular implantation sites. This ectopic osteogenesis is discussed with regard to the nanoporosity and microporosity of the material, physico-chemical interactions at its surface, the differentiation of osteoblasts, the role of angiogenesis and the probable involvement of growth factors. The results of this preliminary study indicate that this biomaterial has osteoinductive potential and induces the formation of bone structures, mainly in subcutaneous adipose tissue in the pig.     

A tree‐ring reconstruction of wind disturbances in a forest of the Slovakian Tatra Mountains,Western Carpathians

Question: Have past windstorm events influenced the structure and composition of mountain forests in the Tatra Mountains? Can severe and infrequent wind disturbances lead to dynamic coexistence of two tree species with different ecological requirements? Location: Subalpine mixed spruce‐larch forest at 1200‐1300 m a.s.l. in the Slovakian Tatra Mountains. A forested site affected by catastrophic large‐scale windthrow on 19 November 2004. Methods: Sixty‐seven spruce and 30 larch cross‐sections from the oldest cohorts were collected in a regular pattern in a 100‐ha plot. Tree‐ring series were analysed to reconstruct growth releases associated with past windthrows. A boundary‐line release criterion was applied to detect disturbance year. Spatial patterns of release signals were statistically detected with Mantel's test. We compared reconstructed years of disturbance events with historical records. Results: Releases in both species showed three main pulses. More than 85% showed major or moderate releases in 1865‐1879, 48% in 1915‐1924, and 25% in 1940‐1949. All of these disturbance events affected the whole 100‐ha area. Releases were spatially patterned in the first disturbances, but distributed randomly in the last. Releases co‐occurred in time with enhanced production of compression wood, suggesting disturbances were of wind origin. Reconstructed dates of windthrows were confirmed using historical data on storms. Conclusions: At least three windthrows of major and moderate severity took place in the last 150 years on southern slopes of the Tatra Mountains. This disturbance regime may contribute to coexistence of spruce and larch through differences in vulnerability and response to heavy windstorms.     

Didymosphaeria igniaria: a new microorganism useful for the enantioselective reduction of aryl-aliphatic ketones

Didymosphaeria igniaria is a promising biocatalyst in asymmetric reductions of prochiral aromatic-aliphatic ketones such as acetonaphthones, acetophenones, and acetylpyridines. The organism converted the substrates mainly to (S)-alcohols. Excellent results in terms of conversion and enantioselectivity (100% yield, >99% ee) were obtained with acetonaphthones. In case of acetyl pyridines, the optical purity of the product depended on the position of the carbonyl group on the pyridine ring and followed the order 2-acetyl ? 4-acetyl > 3-acetyl-pyridine. Transformation of o-methoxy-acetophenone gave optically pure (S)-(-)-1-(2-methoxyphenyl)-ethanol in 95% yield. The transformation of para-methyl ketone gave (R)-alcohol (81% ee), whereas para-bromo ketone gave (S)-alcohol (98% ee). Monitoring of the biotransformation of these substrates over time led to the conclusion that for both substrates, non-selective carbonyl group reduction occurred in the first step, followed by selective oxidation of the (R)-isomer of p-bromo-phenylethanol and selective oxidation of the (S)-isomer of p-methyl-phenylethanol. D. igniaria exhibited poor enantioselectivity in the reduction of bicyclic aryl-aliphatic ketones such as 1- and 2-tetralones. Only (S)-5-methoxy-1-tetralol was obtained in optically pure (>99% ee) form.     

The effect of oxidative stress on nucleotide-excision repair in colon tissue of newborn piglets

Nucleotide-excision repair (NER) is important for the maintenance of genomic integrity and to prevent the onset of carcinogenesis. Oxidative stress was previously found to inhibit NER in vitro, and dietary antioxidants could thus protect DNA not only by reducing levels of oxidative DNA damage, but also by protecting NER against oxidative stress-induced inhibition. To obtain further insight in the relation between oxidative stress and NER activity in vivo, oxidative stress was induced in newborn piglets by means of intra-muscular injection of iron (200 mg) at day 3 after birth. Indeed, injection of iron significantly increased several markers of oxidative stress, such as 8-oxo-7,8-dihydro-2′-deoxyguanosine (8-oxodG) levels in colon DNA and urinary excretion of 8-oxo-7,8-dihydroguanine (8-oxoGua). In parallel, the influence of maternal supplementation with an antioxidant-enriched diet was investigated in their offspring. Supplementation resulted in reduced iron concentrations in the colon (P = 0.004) at day 7 and a 40% reduction of 8-oxodG in colon DNA (P = 0.044) at day 14 after birth. NER capacity in animals that did not receive antioxidants was significantly reduced to 32% at day 7 compared with the initial NER capacity on day 1 after birth. This reduction in NER capacity was less pronounced in antioxidant-supplemented piglets (69%). Overall, these data indicate that NER can be reduced by oxidative stress in vivo, which can be compensated for by antioxidant supplementation.     

The bi-phased course of electrophysiological response of isolated snail intestine on mechanical stimulation

The transepithelial potential difference and changes of diameter of isolated snail intestine as index of its motility were studied in immersed bath in control conditions and after gentle stimulation by 60 seconds of washing of the intestinal lumen. Immediate depolarization and 20% augmentation of the lumen were observed during the stimulation. After stimulation, additional transient depolarization of the transepithelial potential difference and gradual diminution of intestine lumen back to control values over a period of 20 minutes occurred. The immediate reaction was greatly influenced by the presence of sodium or chloride ion transport inhibitors, however, the late phase of the response was not. It is hypothesized that changes of transepithelial electrogenic ion transport and of intestinal motility during the stimulation mirror the inflow of intestinal content and after completion of stimulation may be related to its storage.     

Direct monitoring of albumin lysine-525 N-homocysteinylation in human serum by liquid chromatography/mass spectrometry

A posttranslational protein modification by homocysteine-thiolactone (N-homocysteinylation) is linked to human vascular and neurodegenerative diseases. Although chemical and immunological methods are available to detect and quantify the extent of protein N-homocysteinylation, the determination of site-specific N-homocysteinylation in vivo remains challenging. Here we describe a liquid chromatography/mass spectrometry method that monitors the extent of N-homocysteinylation at albumin lysine-525 in vivo directly in human serum. Using this method, we found that the extent of lysine-525 N-homocysteinylation was significantly increased in patients with cystathionine β-synthase deficiency.     

Evaluation of temporal bone computed tomography and paranasal sinuses magnetic resonance images in acromegalic patients

OBJECTIVE: Acromegaly is a rare disease, which symptoms are caused by excess secretion of a growth hormone (GH) from the anterior pituitary benign tumor - adenoma. Authors present an evaluation of computed tomography (CT) and magnetic resonance (MR) images of temporal bone and paranasal sinuses of patients with acromegaly. CONCLUSIONS: 1. In all patients with acromegaly, morphological changes in paranasal sinuses were shown. They were mostly pronounced within the maxillary sinuses including the mucosa thickening up to 6 mm and encysted fluid occurrence. 2. CT of temporal bone did not reveal structural changes of internal and median ear. 3. There is a need for further studies on hearing impairment in patients with acromegaly.