Biological activities of berries: from antioxidant capacity to anti-cancer effects

Consumption of berries has been implicated with diverse health benefits, such as prevention of stroke, of age-related degenerative diseases and cancer. Some berry constituents have been proven to have cancer preventive actions on chemically induced tumors in vivo and cancer suppressive effects in in vitro studies. Many of these effects were attributed to certain berry phytochemicals with high antioxidative potential that could contribute to, or enhance by induction, the endogenous antioxidant properties of living cells or organisms. Producers and the consumers of berry products need more comprehensive and accurate information on the type and level of health benefits that can be expected from different products. The choice of the chemical or biological test that best predicts specific health benefits of berries is crucial to provide targets for berry breeding programmes or to improve processing methods. The aim of this review is to examine the chemical and biological tests developed to characterize the impact of berries on consumer health.     

Controlled Delivery of Vancomycin via Charged Hydrogels

Surgical site infection (SSI) remains a significant risk for any clean orthopedic surgical procedure. Complications resulting from an SSI often require a second surgery and lengthen patient recovery time. The efficacy of antimicrobial agents delivered to combat SSI is diminished by systemic toxicity, bacterial resistance, and patient compliance to dosing schedules. We submit that development of localized, controlled release formulations for antimicrobial compounds would improve the effectiveness of prophylactic surgical wound antibiotic treatment while decreasing systemic side effects. Our research group developed and characterized oligo(poly(ethylene glycol)fumarate) / sodium methacrylate (OPF/SMA) charged copolymers as biocompatible hydrogel matrices. Here, we report the engineering of this copolymer for use as an antibiotic delivery vehicle in surgical applications. We demonstrate that these hydrogels can be efficiently loaded with vancomycin (over 500 μg drug per mg hydrogel) and this loading mechanism is both time- and charge-dependent. Vancomycin release kinetics are shown to be dependent on copolymer negative charge. In the first 6 hours, we achieved as low as 33.7% release. In the first 24 hours, under 80% of total loaded drug was released. Further, vancomycin release from this system can be extended past four days. Finally, we show that the antimicrobial activity of released vancomycin is equivalent to stock vancomycin in inhibiting the growth of colonies of a clinically derived strain of methicillin-resistant Staphylococcus aureus. In summary, our work demonstrates that OPF/SMA hydrogels are appropriate candidates to deliver local antibiotic therapy for prophylaxis of surgical site infection.     

Application of alkaline elution, Fast Micromethod and flow cytometry in detection of marine contamination.

DNA damage is an inescapable aspect of life in the biosphere. The presented investigations were an attempt to examine the response of a DNA damage as a biomarker of environmental quality in the mussels Mytilus galloprovincialis sampled at differently contaminated areas of Istrian coast, Northern Adriatic. The investigations were performed in order to get information about the genotoxic risk for marine organisms exposed to mixed environmental pollution, as well as the information about the presence of unknown mixture of genotoxic contaminants in the marine environment. Types of DNA damage detected are alkali-labile sites and single-strand breaks measured by Fast Micromethod, interstrand cross-links and DNA protein cross-links by alkaline filter elution and cell cycle disturbation by flow cytometry. The applicability of all three methods for marine quality control is discussed.     

The effectiveness of bone banking in Central Serbia: audit of the first seven years

We analyzed the incidence and predisposing factors for overall discard rate after retrieval of 295 femoral head allografts. The aim of the present study was to evaluate the quality system of institutional bone banking and to ensure that we can provide high standard allografts with low infection rate. Audit of bone banking was conducted on 295 donors and 180 recipients. Of the 295 donated femoral heads 77 were discarded, giving an overall discard rate of 26.1 %. At retrieval, 37 allografts were positive, giving an overall contamination rate of 12.54 %. The organism most commonly identified was Staphylococcus species. Seven (2.37 %) of the 295 allografts failed the blood screening tests. Twelve allografts (4.06 %) were discarded because of suspected damage of the packaging or disuse during surgery. Due to donor death or inability to perform serology retests, 21 (7.11 %) allografts were discarded. In the postoperative survey an infection rate of 2.22 % was found. After 7 years of bone banking, our results show that overall discard rate and allograft related infection rate are in accordance with the international standards. The leading cause of allograft discarding was bacterial contamination influenced by the surgical team. We suggest stringent aseptic allograft handling during harvesting and thawing within highly concentrated antibiotic solution to reduce a possibility of its contamination.     

About and beyond the Henri-Michaelis-Menten rate equation for single-substrate enzyme kinetics

For more than a century the simple single-substrate enzyme kinetics model and related Henri-Michaelis-Menten (HMM) rate equation have been thoroughly explored in various directions. In the present paper we are concerned with a possible generalization of this rate equation recently proposed by F. Kargi (BBRC 382 (2009) 157-159), which is assumed to be valid both in the case that the total substrate or enzyme is in excess and the quasi-steady-state is achieved. We demonstrate that this generalization is grossly inadequate and propose another generalization based on application of the quasi-steady-state condition and conservation equations for both enzyme and substrate. The standard HMM equation is derived by (a) assuming the quasi-steady-state condition, (b) applying the conservation equation only for the enzyme, and (c) assuming that the substrate concentration at quasi-steady-state can be approximated by the total substrate concentration [S](0). In our formula the rate is already expressed through [S](0), and we only assume that when quasi-steady-state is achieved the amount of product formed is negligible compared to [S](0). Numerical simulations show that our formula is generally more accurate than the HMM formula and also can provide a good approximation when the enzyme is in excess, which is not the case for the HMM formula. We show that the HMM formula can be derived from our expression by further assuming that the total enzyme concentration is negligible compared to [S](0).     

Accommodation and repair of a UV photoproduct in DNA at different rotational settings on the nucleosome surface

Cyclobutane-thymine dimers (CTDs), the most common DNA lesion induced by UV radiation, cause 30 degrees bending and 9 degrees unwinding of the DNA helix. We prepared site-specific CTDs within a short sequence bracketed by strong nucleosome-positioning sequences. The rotational setting of CTDs over one turn of the helix near the dyad center on the histone surface was analyzed by hydroxyl radical footprinting. Surprisingly, the position of CTDs over one turn of the helix does not affect the rotational setting of DNA on the nucleosome surface. Gel-shift analysis indicates that one CTD destabilizes histone-DNA interactions by 0.6 or 1.1 kJ/mol when facing away or toward the histone surface, respectively. Thus, 0.5 kJ/mol energy penalty for a buried CTD is not enough to change the rotational setting of sequences with strong rotational preference. The effect of rotational setting on CTD removal by nucleotide excision repair (NER) was examined using Xenopus oocyte nuclear extracts. The NER rates are only 2-3 times lower in nucleosomes and change by only 1.5-fold when CTDs face away or toward the histone surface. Therefore, in Xenopus nuclear extracts, the rotational orientation of CTDs on nucleosomes has surprisingly little effect on rates of repair. These results indicate that nucleosome dynamics and/or chromatin remodeling may facilitate NER in gaining access to DNA damage in nucleosomes.     

Temporal expression of hypoxia-regulated genes is associated with early changes in redox status in irradiated lung

The development of normal lung tissue toxicity after radiation exposure results from multiple changes in cell signaling and communication initiated at the time of the ionizing event. The onset of gross pulmonary injury is preceded by tissue hypoxia and chronic oxidative stress. We have previously shown that development of debilitating lung injury can be mitigated or prevented by administration of AEOL10150, a potent catalytic antioxidant, 24h after radiation. This suggests that hypoxia-mediated signaling pathways may play a role in late radiation injury, but the exact mechanism remains unclear. The purpose of this study was to evaluate changes in the temporal expression of hypoxia-associated genes in irradiated mouse lung and determine whether AEOL10150 alters expression of these genes. A focused oligo array was used to establish a hypoxia-associated gene expression signature for lung tissue from sham-irradiated or irradiated mice treated with or without AEOL10150. Results were further verified by RT-PCR. Forty-four genes associated with metabolism, cell growth, apoptosis, inflammation, oxidative stress, and extracellular matrix synthesis were upregulated after radiation. Elevated expression of 31 of these genes was attenuated in animals treated with AEOL10150, suggesting that expression of a number of hypoxia-associated genes is regulated by early development of oxidative stress after radiation. Genes identified herein could provide insight into the role of hypoxic signaling in radiation lung injury, suggesting novel therapeutic targets, as well as clues to the mechanism by which AEOL10150 confers pulmonary radioprotection.     

The mitochondrial bioenergetic phenotype for protection from cardiac ischemia in SUR2 mutant mice

The sulfonylurea receptor-2 (SUR2) is a subunit of ATP-sensitive potassium channels (K(ATP)) in heart. Mice with the SUR2 gene disrupted (SUR2m) are constitutively protected from ischemia-reperfusion (I/R) cardiac injury. This was surprising because K(ATP), either sarcolemmal or mitochondrial or both, are thought to be important for cardioprotection. We hypothesized that SUR2m mice have an altered mitochondrial phenotype that protects against I/R. Mitochondrial membrane potential (ΔΨ(m)), tolerance to Ca(2+) load, and reactive oxygen species (ROS) generation were studied by fluorescence-based assays, and volumetric changes in response to K(+) were measured by light scattering in isolated mitochondria. For resting SUR2m mitochondria compared with wild type, the ΔΨ(m) was less polarized (46.1 ± 0.4 vs. 51.9 ± 0.6%), tolerance to Ca(2+) loading was increased (163 ± 2 vs. 116 ± 2 μM), and ROS generation was enhanced with complex I [8.5 ± 1.2 vs. 4.9 ± 0.2 arbitrary fluorescence units (afu)/s] or complex II (351 ± 51.3 vs. 166 ± 36.2 afu/s) substrates. SUR2m mitochondria had greater swelling in K(+) medium (30.2 ± 3.1%) compared with wild type (14.5 ± 0.6%), indicating greater K(+) influx. Additionally, ΔΨ(m) decreased and swelling increased in the absence of ATP in SUR2m, but the sensitivity to ATP was less compared with wild type. When the mitochondria were subjected to hypoxia-reoxygenation, the decrease in respiration rates and respiratory control index was less in SUR2m. ΔΨ(m) maintenance in the SUR2m intact myocytes was also more tolerant to metabolic inhibition. In conclusion, the cardioprotection observed in the SUR2m mice is associated with a protected mitochondrial phenotype resulting from enhanced K(+) conductance that partially dissipated ΔΨ(m). These results have implications for possible SUR2 participation in mitochondrial K(ATP).     

Enhanced charge-independent mitochondrial free Ca(2+) and attenuated ADP-induced NADH oxidation by isoflurane: Implications for cardioprotection

Modulation of mitochondrial free Ca(2+) ([Ca(2+)](m)) is implicated as one of the possible upstream factors that initiates anesthetic-mediated cardioprotection against ischemia-reperfusion (IR) injury. To unravel possible mechanisms by which volatile anesthetics modulate [Ca(2+)](m) and mitochondrial bioenergetics, with implications for cardioprotection, experiments were conducted to spectrofluorometrically measure concentration-dependent effects of isoflurane (0.5, 1, 1.5, 2mM) on the magnitudes and time-courses of [Ca(2+)](m) and mitochondrial redox state (NADH), membrane potential (ΔΨ(m)), respiration, and matrix volume. Isolated mitochondria from rat hearts were energized with 10mM Na(+)- or K(+)-pyruvate/malate (NaPM or KPM) or Na(+)-succinate (NaSuc) followed by additions of isoflurane, 0.5mM CaCl(2) (≈200nM free Ca(2+) with 1mM EGTA buffer), and 250μM ADP. Isoflurane stepwise: (a) increased [Ca(2+)](m) in state 2 with NaPM, but not with KPM substrate, despite an isoflurane-induced slight fall in ΔΨ(m) and a mild matrix expansion, and (b) decreased NADH oxidation, respiration, ΔΨ(m), and matrix volume in state 3, while prolonging the duration of state 3 NADH oxidation, respiration, ΔΨ(m), and matrix contraction with PM substrates. These findings suggest that isoflurane's effects are mediated in part at the mitochondrial level: (1) to enhance the net rate of state 2 Ca(2+) uptake by inhibiting the Na(+)/Ca(2+) exchanger (NCE), independent of changes in ΔΨ(m) and matrix volume, and (2) to decrease the rates of state 3 electron transfer and ADP phosphorylation by inhibiting complex I. These direct effects of isoflurane to increase [Ca(2+)](m), while depressing NCE activity and oxidative phosphorylation, could underlie the mechanisms by which isoflurane provides cardioprotection against IR injury at the mitochondrial level.     

Our experiences in treatment of prostate carcinoma in patients over the age of 70

The goal of our study was to present our long-standing experience of the treatment of prostate carcinoma in patients over the age of 70. During the 20 years period (from 1991 to 2010) we diagnosed the prostate carcinoma in 1998 patients. More than 58% of the patients were over 70 years old. The most frequent symptoms of the prostate carcinoma were frequent urination and backache. At the first examination 36% of the patients had both prostate lobes involved, and 27% of them had metastases. The most frequent ones (26%) were those in the bone system (pelvis and spine), while in only 1% metastases were found in solid organs (lungs and liver). According to the TNM classification, T1 and T2 were diagnosed in 818 (71%) patients. Histopathological examination discovered Gleason score 2 in 70% of patients and Gleason score 3 in 24% of them. Most often the combination of castration and antiandrogen therapy (in 68% of the patients) and the combination of castration and Estracyt therapy (in 19% of the patients) were applied. In conclusion, intensified efforts should be made in promoting preventive urological examinations because of the great number of patients (27%) with metastases at the first examination.